Defining the role of mesenchymal stromal cells on the regulation of matrix metalloproteinases in skeletal muscle cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Sassoli, Chiara; Nosi, Daniele; Tani, Alessia

Recent studies indicate that mesenchymal stromal cell (MSC) transplantation improves healing of injured and diseased skeletal muscle, although the mechanisms of benefit are poorly understood. In the present study, we investigated whether MSCs and/or their trophic factors were able to regulate matrix metalloproteinase (MMP) expression and activity in different cells of the muscle tissue. MSCs in co-culture with C2C12 cells or their conditioned medium (MSC-CM) up-regulated MMP-2 and MMP-9 expression and function in the myoblastic cells; these effects were concomitant with the down-regulation of the tissue inhibitor of metalloproteinases (TIMP)-1 and -2 and with increased cell motility. In the singlemore » muscle fiber experiments, MSC-CM administration increased MMP-2/9 expression in Pax-7{sup +} satellite cells and stimulated their mobilization, differentiation and fusion. The anti-fibrotic properties of MSC-CM involved also the regulation of MMPs by skeletal fibroblasts and the inhibition of their differentiation into myofibroblasts. The treatment with SB-3CT, a potent MMP inhibitor, prevented in these cells, the decrease of α-smooth actin and type-I collagen expression induced by MSC-CM, suggesting that MSC-CM could attenuate the fibrogenic response through mechanisms mediated by MMPs. Our results indicate that growth factors and cytokines released by these cells may modulate the fibrotic response and improve the endogenous mechanisms of muscle repair/regeneration. - Highlights: • MSC-CM contains paracrine factors that up-regulate MMP expression and function in different skeletal muscle cells. • MSC-CM promotes myoblast and satellite cell migration, proliferation and differentiation. • MSC-CM negatively interferes with fibroblast-myoblast transition in primary skeletal fibroblasts. • Paracrine factors from MSCs modulate the fibrotic response and improve the endogenous mechanisms of muscle regeneration.« less

Dual Effects of Cell Free Supernatants from Lactobacillus acidophilus and Lactobacillus rhamnosus GG in Regulation of MMP-9 by Up-Regulating TIMP-1 and Down-Regulating CD147 in PMA- Differentiated THP-1 Cells

PubMed Central

Maghsood, Faezeh; Mirshafiey, Abbas; Farahani, Mohadese M.; Modarressi, Mohammad Hossein; Jafari, Parvaneh; Motevaseli, Elahe

2018-01-01

Objective Recent studies have reported dysregulated expression of matrix metalloproteinases (MMPs), especially MMP-2, MMP-9, tissue inhibitor of metalloproteinase-1, -2 (TIMP-1, TIMP-2), and extracellular matrix metalloproteinase inducer (EMMPRIN/CD147) in activated macrophages of patients with inflammatory diseases. Therefore, MMP-2, MMP-9, and their regulators may represent a new target for treatment of inflammatory diseases. Probiotics, which are comprised of lactic acid bacteria, have the potential to modulate inflammatory responses. In this experimental study, we investigated the anti-inflammatory effects of cell-free supernatants (CFS) from Lactobacillus acidophilus (L. acidophilus) and L. rhamnosus GG (LGG) in phorbol myristate acetate (PMA)-differentiated THP-1 cells. Materials and Methods In this experimental study, PMA-differentiated THP-1 cells were treated with CFS from L. acidophilus, LGG and uninoculated bacterial growth media (as a control). The expression of MMP-2, MMP-9, TIMP-1, and TIMP-2 mRNAs were determined using real-time quantitative reverse transcription polymerase chain reaction (RT- PCR). The levels of cellular surface expression of CD147 were assessed by flow cytometry, and the gelatinolytic activity of MMP-2 and MMP-9 were determined by zymography. Results Our results showed that CFS from both L. acidophilus and LGG significantly inhibited the gene expression of MMP-9 (P=0.0011 and P=0.0005, respectively), increased the expression of TIMP-1 (P<0.0001), decreased the cell surface expression of CD147 (P=0.0307 and P=0.0054, respectively), and inhibited the gelatinolytic activity of MMP-9 (P=0.0003 and P<0.0001, respectively) in PMA-differentiated THP-1 cells. Although, MMP-2 expression and activity and TIMP-2 expression remained unchanged. Conclusion Our results indicate that CFS from L. acidophilus and LGG possess anti-inflammatory properties and can modulate the inflammatory response. PMID:29105390

ADAM17 targets MMP-2 and MMP-9 via EGFR-MEK-ERK pathway activation to promote prostate cancer cell invasion.

PubMed

Xiao, Li-Jie; Lin, Ping; Lin, Feng; Liu, Xin; Qin, Wei; Zou, Hai-Feng; Guo, Liang; Liu, Wei; Wang, Shu-Juan; Yu, Xiao-Guang

2012-05-01

ADAM17, also known as tumor necrosis factor-α converting enzyme (TACE), is involved in proteolytic ectodomain shedding of cell surface molecules and cytokines. Although aberrant expression of ADAM17 has been shown in various malignancies, the function of ADAM17 in prostate cancer has not been clarified. In the present study, we sought to elucidate whether ADAM17 contributes to prostate cancer cell invasion, as well as the mechanism involved in the process. The expression pattern of ADAM17 was investigated in human prostate cancer cells. The results showed that ADAM17 expression levels are correlated with the invasive ability of androgen-independent prostate cancer cell lines. Further, ADAM17 was overexpressed in cells showing high invasion characteristics, activation of the EGFR-MEK-ERK pathway, up-regulation of MMP-2, MMP-9, and an increased TGF-α release into the supernatant. However, AG1478, PD98059 and antibody against TGF-α deactivating the EGFR-MEK-ERK signaling pathway, abolished up-regulation of MMP-2, MMP-9 and prevented cell invasion. In addition, cells with knockdown of ADAM17 by siRNA exhibited low invasive ability, deactivated EGFR-MEK-ERK signaling pathway, reduced TGF-α released and down-regulation of MMP-2, MMP-9. However, these effects could be reversed by simultaneous addition of TGF-α. These data demonstrated that ADAM17 contributes to androgen-independent prostate cancer cell invasion by shedding of EGFR ligand TGF-α, which subsequently activates the EGFR-MEK-ERK signaling pathway, leading finally to overexpression of MMP-2 and MMP-9. This study suggests that the ADAM17 expression level may be a new predictive biomarker of invasion and metastasis of prostate cancer, and ADAM17 could provide a target for treating metastatic PCa.

NFκB- and AP-1-mediated DNA looping regulates matrix metalloproteinase-9 transcription in TNF-α-treated human leukemia U937 cells.

PubMed

Chen, Ying-Jung; Chang, Long-Sen

2015-10-01

The aim of this study is to explore the spatial association of critical genomic elements in the effect of TNF-α on matrix metalloproteinase-9 (MMP-9) expression in human leukemia U937 cells. TNF-α up-regulated MMP-9 protein expression and mRNA level in U937 cells, and Akt-mediated-NFκB/p65 activation and JNK-mediated c-Jun activation were proven to be involved in TNF-α-induced MMP-9 up-regulation. Promoter luciferase activity assay revealed that NFκB (nt-600) and AP-1 (nt-79) binding sites were crucial for TNF-α-induced transcription of MMP-9 gene. The results of a chromatin immunoprecipitation assay indicated that TNF-α reduced histone deacetylase-1 (HDAC-1) recruitment but increased p300 (a histone acetyltransferase) recruitment to MMP-9 promoter regions surrounding NFκB and AP-1 binding sites. Consistently, TNF-α increased enrichment of the acetylated histone H3 mark on MMP-9 promoter regions. DNA affinity purification assay revealed that p300 and HDAC1 could bind oligonucleotides containing AP-1/c-Jun and NFκB/p65 binding sites. Chromosome conformation capture assay showed that TNF-α stimulated chromosomal loops in the MMP-9 promoter via NFκB/p65 and AP-1/c-Jun. The p300-associated acetyltransferase activity was crucial for p65/c-Jun-mediated DNA looping, and inhibition of HDAC activity increased the level of DNA looping. Reduction in the level of DNA looping eliminated all TNF-α-stimulated MMP-9 up-regulation. Taken together, our data suggest that p65/c-Jun-mediated DNA looping is involved in TNF-α-induced MMP-9 up-regulation and that the recruitment of p300 or HDAC1 to NFκB and AP-1 binding sites modifies the level of DNA looping. Copyright © 2015 Elsevier B.V. All rights reserved.

Interleukin-6 triggers human cerebral endothelial cells proliferation and migration: The role for KDR and MMP-9

DOE Office of Scientific and Technical Information (OSTI.GOV)

Yao, Jianhua S.; Zhai Wenwu; Young, William L.

2006-04-21

Interleukin-6 (IL-6) is involved in angiogenesis. However, the underlying mechanisms are unknown. Using human cerebral endothelial cell (HCEC), we report for First time that IL-6 triggers HCEC proliferation and migration in a dose-dependent manner, specifically associated with enhancement of VEGF expression, up-regulated and phosphorylated VEGF receptor-2 (KDR), and stimulated MMP-9 secretion. We investigated the signal pathway of IL-6/IL-6R responsible for KDR's regulation. Pharmacological inhibitor of PI3K failed to inhibit IL-6-mediated VEGF overexpression, while blocking ERK1/2 with PD98059 could abolish IL-6-induced KDR overexpression. Further, neutralizing endogenous VEGF attenuated KDR expression and phosphorylation, suggesting that IL-6-induced KDR activation is independent of VEGFmore » stimulation. MMP-9 inhibitor GM6001 significantly decreases HCEC proliferation and migration (p < 0.05), indicating the crucial function of MMP-9 in promoting angiogenic changes in HCECs. We conclude that IL-6 triggers VEGF-induced angiogenic activity through increasing VEGF release, up-regulates KDR expression and phosphorylation through activating ERK1/2 signaling, and stimulates MMP-9 overexpression.« less

Physiological loading of joints prevents cartilage degradation through CITED2

PubMed Central

Leong, Daniel J.; Li, Yong H.; Gu, Xiang I.; Sun, Li; Zhou, Zuping; Nasser, Philip; Laudier, Damien M.; Iqbal, Jameel; Majeska, Robert J.; Schaffler, Mitchell B.; Goldring, Mary B.; Cardoso, Luis; Zaidi, Mone; Sun, Hui B.

2011-01-01

Both overuse and disuse of joints up-regulate matrix metalloproteinases (MMPs) in articular cartilage and cause tissue degradation; however, moderate (physiological) loading maintains cartilage integrity. Here, we test whether CBP/p300-interacting transactivator with ED-rich tail 2 (CITED2), a mechanosensitive transcriptional coregulator, mediates this chondroprotective effect of moderate mechanical loading. In vivo, hind-limb immobilization of Sprague-Dawley rats up-regulates MMP-1 and causes rapid, histologically detectable articular cartilage degradation. One hour of daily passive joint motion prevents these changes and up-regulates articular cartilage CITED2. In vitro, moderate (2.5 MPa, 1 Hz) intermittent hydrostatic pressure (IHP) treatment suppresses basal MMP-1 expression and up-regulates CITED2 in human chondrocytes, whereas high IHP (10 MPa) down-regulates CITED2 and increases MMP-1. Competitive binding and transcription assays demonstrate that CITED2 suppresses MMP-1 expression by competing with MMP transactivator, Ets-1 for its coactivator p300. Furthermore, CITED2 up-regulation in vitro requires the p38δ isoform, which is specifically phosphorylated by moderate IHP. Together, these studies identify a novel regulatory pathway involving CITED2 and p38δ, which may be critical for the maintenance of articular cartilage integrity under normal physical activity levels.—Leong, D. J., Li, Y. H., Gu, X. I., Sun, L., Zhou, Z., Nasser, P., Laudier, D. M., Iqbal, J., Majeska, R. J., Schaffler, M. B., Goldring, M. B., Cardoso, L., Zaidi, M., Sun, H. B. Physiological loading of joints prevents cartilage degradation through CITED2. PMID:20826544

Expression of matrix metalloproteinase enzymes in endometrium of women with abnormal uterine bleeding.

PubMed

Grzechocinska, Barbara; Dabrowski, Filip A; Chlebus, Marcin; Gondek, Agata; Czarzasta, Katarzyna; Michalowski, Lukasz; Cudnoch-Jedrzejewska, Agnieszka; Wielgos, Miroslaw

2018-02-01

Abnormal uterine bleeding (AUB) is caused by derangement of physiological processes of tissue growth, shedding and regeneration. It is known that interplay between metalloproteinases (MMP's) and tissue inhibitors of metalloproteinases (TIMP's) may play a crucial role in its occurrence. To define if expression of proMMP-2, MMP-2 and TIMP-1 in endometrium of women with AUB is dependent on steroid sex hormone concentration and histopathological picture. Endometrial scraps were taken from 21 women with AUB and 19 controls. Samples were evaluated in light microscopy by a certified pathologist. Activity of proMMP-2 and MMP-2 proteins levels were evaluated by gelatin zymography and TIMP-1 by reversed zymography. The results has been correlated with serum estradiol and progesterone concentrations in linear regression model. Expression: of proMMP-2 in endometrium of women with AUB is correlated with estradiol concentration and inversely correlated with progesterone levels. It was significantly higher in women with dysfunctional endometrium (p<0.001). Expression of MMP-2 was highest in women with endometrial polyps and longer bleeding (p<0.01), while expression of TIMP-1 was independent from hormone concentration. Lack of correlation between proMMP-2 and MMP-2 levels suggest different pathway of their activation in AUB. ProMMP-2 is up regulated by estradiol and down regulated by progesterone while MMP-2 levels increase with the length of bleeding.

High glucose induces podocyte epithelial-to-mesenchymal transition by demethylation-mediated enhancement of MMP9 expression

PubMed Central

Ling, Li; Chen, Libo; Zhang, Changning; Gui, Shuyan; Zhao, Haiyan; Li, Zhengzhang

2018-01-01

Abnormal expression of matrix metalloproteinase 9 (MMP9) is correlated with podocyte epithelial-to-mesenchymal transition (EMT) in diabetic nephropathy (DN). However, the mechanisms underlying this process are not well defined. Site-specific demethylation may sustain high expression levels of target genes. In the present study, in order to investigate the association between DNA demethylation of MMP9 promoter and podocyte EMT in DN, human podocytes were cultured in high-glucose (HG) medium and a rat model of DN was established by intraperitoneal injection of streptozotocin (STZ) to determine whether site-specific demethylation of the MMP9 promoter was involved in regulating podocyte EMT in DN. The MTT assay was used to assess the effects of HG culture on the growth of podocytes, and the demethylation status of the MMP9 promoter was assessed by bisulfite sequencing polymerase chain reaction. mRNA and protein expression levels of MMP9, α-smooth muscle actin (α-SMA), podocalyxin and fibronectin-1 in podocytes were assessed by reverse transcription-quantitative PCR (RT-qPCR) and western blot analyses. The results demonstrated that HG treatment up regulated the expression of MMP9, α-SMA and fibronectin-1, but down regulated the expression of podocalyxin in podocytes. The MMP9 promoter region was revealed to contain a variety of demethylated CpG sites, and HG treatment reduced the rate of MMP9 promotermethylation, which, in turn, enhanced its promoter activity. In summary, these data suggested that demethylation of the MMP9 promoter may serve an important role in podocyte EMT in DN. The demethylation status of the MMP9 promoter maybe used as an important prognostic marker of DN in clinic. PMID:29436620

Invasive ability of human renal cell carcinoma cell line Caki-2 is accelerated by gamma-aminobutyric acid, via sustained activation of ERK1/2 inducible matrix metalloproteinases.

PubMed

Inamoto, Teruo; Azuma, Haruhito; Sakamoto, Takeshi; Kiyama, Satoshi; Ubai, Takanobu; Kotake, Yatsugu; Watanabe, Masahito; Katsuoka, Yoji

2007-10-01

Gamma-aminobutyric acid (GABA) was first discovered as an inhibitory neurotransmitter in the central nervous system (CNS) and has been reported to have a variety of functions, including regulation of cell division, cell differentiation and maturation, and to be involved in the development of certain cancers outside the CNS. In the present study, using the human renal cell carcinoma cell line Caki-2, we demonstrated that GABA stimulation significantly increased the expression of MMP-2 and -9 and subsequently increased the invasive activity of the cancer cells. Because MAPK signaling is one of the key regulators of MMP expression, we further evaluated MAPK signaling after stimulation with GABA. It was found that GABA stimulation promoted the phosphorylation of MAPKs, including ERK1/2, JNK, and p38. ERK1/2 phosphorylation was sustained for up to 12 h, while phosphorylation of JNK and p38 returned to the endogenous level by 30 min. It was noteworthy that the ras/raf/MEK/ERK pathway inhibitor PD98059 attenuated GABA-induced MMP-9 expression and that both PD98059 and MMP inhibitors attenuated the GABA-induced invasive activity of Caki-2 cells. Moreover, data obtained by depletion of the MEK/ERK pathway using interfering RNA transfection of Caki-2 cells clearly corroborated the above results, as both MMP-9 expression and GABA-induced invasive ability were decreased significantly. We also demonstrated that the GABA-induced increase in invasive ability via ERK1/2 up-regulation was mediated mainly through the GABA-B receptor. These results indicate that GABA stimulation promotes cancer cell invasion and that the effect is partly due to ERK1/2-dependent up-regulation of MMPs.

Isoginkgetin inhibits tumor cell invasion by regulating phosphatidylinositol 3-kinase/Akt-dependent matrix metalloproteinase-9 expression.

PubMed

Yoon, Sang-Oh; Shin, Sejeong; Lee, Ho-Jae; Chun, Hyo-Kon; Chung, An-Sik

2006-11-01

Matrix metalloproteinase (MMP)-9 plays a key role in tumor invasion. Inhibitors of MMP-9 were screened from Metasequoia glyptostroboides (Dawn redwood) and one potent inhibitor, isoginkgetin, a biflavonoid, was identified. Noncytotoxic levels of isoginkgetin decreased MMP-9 production profoundly, but up-regulated the level of tissue inhibitor of metalloproteinase (TIMP)-1, an inhibitor of MMP-9, in HT1080 human fibrosarcoma cells. The major mechanism of Ras-dependent MMP-9 production in HT1080 cells was phosphatidylinositol 3-kinase (PI3K)/Akt/nuclear factor-kappaB (NF-kappaB) activation. Expression of dominant-active H-Ras and p85 (a subunit of PI3K) increased MMP-9 activity, whereas dominant-negative forms of these molecules decreased the level of MMP-9. H-Ras did not increase MMP-9 in the presence of a PI3K inhibitor, LY294002, and a NF-kappaB inhibitor, SN50. Further studies showed that isoginkgetin regulated MMP-9 production via PI3K/Akt/NF-kappaB pathway, as evidenced by the findings that isoginkgetin inhibited activities of both Akt and NF-kappaB. PI3K/Akt is a well-known key pathway for cell invasion, and isoginkgetin inhibited HT1080 tumor cell invasion substantially. Isoginkgetin was also quite effective in inhibiting the activities of Akt and MMP-9 in MDA-MB-231 breast carcinomas and B16F10 melanoma. Moreover, isoginkgetin treatment resulted in marked decrease in invasion of these cells. In summary, PI3K/Akt is a major pathway for MMP-9 expression and isoginkgetin markedly decreased MMP-9 expression and invasion through inhibition of this pathway. This suggests that isoginkgetin could be a potential candidate as a therapeutic agent against tumor invasion.

DNA Methylation of MMP9 Is Associated with High Levels of MMP-9 Messenger RNA in Periapical Inflammatory Lesions.

PubMed

Campos, Kelma; Gomes, Carolina Cavalieri; Farias, Lucyana Conceição; Silva, Renato Menezes; Letra, Ariadne; Gomez, Ricardo Santiago

2016-01-01

Matrix metalloproteinases (MMPs) are the major class of enzymes responsible for degradation of extracellular matrix components and participate in the pathogenesis of periapical inflammatory lesions. MMP expression may be regulated by DNA methylation. The purpose of the present investigation was to analyze the expression of MMP2 and MMP9 in periapical granulomas and radicular cysts and to test the hypothesis that, in these lesions, their transcription may be modulated by DNA methylation. Methylation-specific polymerase chain reaction was used to evaluate the DNA methylation pattern of the MMP2 gene in 13 fresh periapical granuloma samples and 10 fresh radicular cyst samples. Restriction enzyme digestion was used to assess methylation of the MMP9 gene in 12 fresh periapical granuloma samples and 10 fresh radicular cyst samples. MMP2 and MMP9 messenger RNA transcript levels were measured by quantitative real-time polymerase chain reaction. All periapical lesions and healthy mucosa samples showed partial methylation of the MMP2 gene; however, periapical granulomas showed higher MMP2 mRNA expression levels than healthy mucosa (P = .014). A higher unmethylated profile of the MMP9 gene was found in periapical granulomas and radicular cysts compared with healthy mucosa. In addition, higher MMP9 mRNA expression was observed in the periapical lesions compared with healthy tissues. The present study suggests that the unmethylated status of the MMP9 gene in periapical lesions may explain the observed up-regulation of messenger RNA transcription in these lesions. Copyright © 2016 American Association of Endodontists. Published by Elsevier Inc. All rights reserved.

The Role of FAK in the Secretion of MMP9 after CD147 Stimulation in Macrophages.

PubMed

Yu, Chen; Lixia, Yang; Ruiwei, Guo; Yankun, Shi; Jinshan, Ye

2018-03-30

To investigate whether focal adhesion kinase (FAK) can participate in the secretion of matrix metalloproteinase 9 (MMP9) after CD147 stimulation in THP-1 induced macrophages; thus, to explore the potential treatment perspectives for acute coronary syndrome (ACS).Phorbol-12-myristate-13-acetate (PMA) was used to induce THP-1 cells to differentiate into macrophages. To confirm the peak mRNA and protein expression of FAK and MMP9 after the stimulation of CD147, the macrophages were divided into 5 groups (0, 3, 6, 9, and 12 hours), with 0 hours group as control group. To investigate the role of FAK in the secretion of MMP9, with stimulation of CD147 for 9 hours, FAK inhibitor 14 was used to inhibit FAK Y397 phosphorylation. The mRNA and protein expressions were quantified by qRT-PCR and western blotting, respectively. (1) Relative mRNA expression of FAK and MMP9 were both significantly up-regulated (all P < 0.05) after stimulation of CD147, FAK peaked at 9 hours (3.908 ± 0.106 versus 1, P < 0.05), whereas MMP9 peaked at 6 hours (2.522 ± 0.062 versus 1, P < 0.05). (2) Relative protein expression of FAK, pFAK, and MMP9 were all significantly increased after CD147 stimulation (all P < 0.05), FAK (1.930 ± 0.024 versus 1, P < 0.05) and pFAK (1.737 ± 0.021 versus 1, P < 0.05) peaked at 9 hours, whereas MMP9 peaked at 6 hours (1.527 ± 0.033 versus 1, P < 0.05). (3) CD147 up-regulates FAK, pFAK, and MMP9 mRNA and protein expressions in a dose-dependent manner. (4) FAK inhibitor 14 significantly reduced the relative protein expression level of pFAK (0.077 ± 0.012 versus 1, P < 0.05) and MMP9 (0.133 ± 0.012) at 9 hours after CD147 stimulation.The results demonstrated that FAK Y397 phosphorylation was involved in the secretion of MMP9 after CD147 stimulation in macrophages and may play a role in the regulation of ACS.

The involvement of osteopontin and matrix metalloproteinase- 9 in the migration of endometrial epithelial cells in patients with endometriosis.

PubMed

Yang, Mei; Jiang, Chunfan; Chen, Hua; Nian, Yan; Bai, Zhimiao; Ha, Chunfang

2015-08-20

Endometriosis, which shares certain characteristics with cancers, may cause abnormal expression of proteins involved in cell migration. Endometrial epithelial cells (EECs) are believed to play an important role in endometriotic migration. The aim of this study was to investigate the relationship between the expression of osteopontin (OPN) and matrix metalloproteinase-9 (MMP-9) in endometriotic migration. We performed primary culture of EECs and investigated the expression of OPN and MMP-9 in EECs regulated by 17beta-estradiol (E2). OPN-specific siRNA interference was used to down-regulate OPN and to explore the corresponding change in MMP-9 expression. Real-time RT-PCR, western blot analysis and flow cytometry were used to determine the expression levels of OPN and MMP-9. Gelatin zymography was performed to observe the enzymatic activity of MMP-9 in conditioned media. Transwell and wound scratch assays were performed to investigate the migration ability of EECs. The expression levels of OPN and MMP-9 in normal EECs (NEECs) were inferior to those in EECs from patients with endometriosis (EEECs). The expression levels of OPN and MMP-9 from stage III/IV EEECs and secretory-phase EECs were higher than those of stage I/II EEECs or proliferative-phase EECs. The expression levels of OPN and MMP-9 in EEECs were increased by E2 treatment and remarkably decreased by siRNA interference. Active MMP-9 expression increased with E2 treatment and decreased with siRNA treatment in EEECs compared with the same treatments in NEECs. The migratory abilities of EEECs were enhanced after cells were treated with E2; in contrast, these abilities were reduced by siRNA interference. In NEECs, active MMP-9 and cellular migration abilities were only minimally influenced by E2 and siRNA treatment. The present study suggests that the up-regulation of MMP-9 via activation of OPN induced by estrogen may correlate with the migration of endometrial epithelial cells in patients with endometriosis.

MMPs-Mediated ECM Remodeling

PubMed

Mali, Aniket V; Joshi, Asavari A; Hegde, Mahabaleshwar V; Kadam, Shivajirao S

2017-04-01

Background: To enhance their own survival, tumor cells can manipulate their microenvironment through remodeling of the extra cellular matrix (ECM). The urokinase-type plasminogen activator (uPA) system catalyzes plasmin production which further mediates activation of matrix metalloproteinases (MMPs) and plays an important role in breast cancer invasion and metastasis through ECM remodeling. This provides a potential target for therapeutic intervention of breast cancer treatment. Enterolactone (EL) is derived from dietary flax lignans in the human body and is known to have anti-breast cancer activity. We here investigated molecular and cellular mechanisms of EL action on the uPA-plasmin- MMPs system. Methods: MTT and trypan blue dye exclusion assays, anchorage-dependent clonogenic assays and wound healing assays were carried out to study effects on cell proliferation and viability, clonogenicity and migration capacity, respectively. Real-time PCR was employed to study gene expression and gelatin zymography was used to assess MMP-2 and MMP-9 activities. All data were statistically analysed and presented as mean ± SEM values. Results: All the findings collectively demonstrated anticancer and antimetastatic potential of EL with antiproliferative, antimigratory and anticlonogenic cellular mechanisms. EL was found to exhibit multiple control of plasmin activation by down-regulating uPA expression and also up-regulating its natural inhibitor, PAI-1, at the mRNA level. Further, EL was found to down-regulate expression of MMP-2 and MMP-9 genes, and up-regulate TIMP-1 and TIMP-2; natural inhibitors of MMP-2 and MMP-9, respectively. This may be as a consequence of inhibition of plasmin activation, resulting in robust control over migration and invasion of breast cancer cells during metastasis. Conclusions: EL suppresses proliferation, migration and metastasis of MDA-MB-231 breast cancer cells by inhibiting induced ECM remodeling by the ‘uPA-plasmin-MMPs system’. Creative Commons Attribution License

CAPN 7 promotes the migration and invasion of human endometrial stromal cell by regulating matrix metalloproteinase 2 activity.

PubMed

Liu, Hongyu; Jiang, Yue; Jin, Xiaoyan; Zhu, Lihua; Shen, Xiaoyue; Zhang, Qun; Wang, Bin; Wang, Junxia; Hu, Yali; Yan, Guijun; Sun, Haixiang

2013-07-15

Matrix metalloproteinase 2 (MMP-2) has been reported to be an important regulator of cell migration and invasion through degradation of the extracellular matrix (ECM) in many diseases, such as cancer and endometriosis. Here, we found calcium-activated neutral protease 7 (CAPN 7) expression was markedly upregulated in the eutopic endometrium and endometrial stromal cells of women diagnosed with endometriosis. Our studies were carried out to detect the effects of CAPN 7 on human endometrial stromal cell (hESC) migration and invasion. Western blotting and quantitative real-time PCR were used to detect the expression of CAPN 7 in endometriosis patients and normal fertile women. Scratch-wound-healing and invasion chamber assay were used to investigate the role of CAPN 7 in hESC migration and invasion. Western blotting, quantitative real-time PCR and zymography were carried out to detect the effect of CAPN 7 on the expressions and activity of MMP-2. CAPN 7 was markedly up-regulated in endometriosis, thereby promoting the migration and invasion of hESC. CAPN 7 overexpression led to increased expression of MMP-2 and tissue inhibitor of metalloproteinases 2 (TIMP-2); CAPN 7 knockdown reversed these changes. CAPN 7 increased MMP-2 activity by increasing the ratio of MMP-2 to TIMP-2. We also found that OA-Hy (an MMP-2 inhibitor) decreased the effects of CAPN 7 overexpression on hESC migration and invasion by approximately 50% and 55%, respectively. Additionally, a coimmunoprecipitation assay demonstrated that CAPN 7 interacted with activator protein 2α (AP-2α): an important transcription factor of MMP-2. CAPN 7 promotes hESC migration and invasion by increasing the activity of MMP-2 via an increased ratio of MMP-2 to TIMP-2.

Leptin promotes human endometriotic cell migration and invasion by up-regulating MMP-2 through the JAK2/STAT3 signaling pathway.

PubMed

Ahn, Ji-Hye; Choi, Youn Seok; Choi, Jung-Hye

2015-10-01

Despite evidence that leptin may play a role in the pathogenesis of endometriosis, the specific function of leptin in the migration and invasion of endometriotic cells is not well characterized. In this study, we investigated the effect of leptin on the migration, invasion and matrix metalloproteinase (MMP) expression levels of human endometriotic cells. We found that leptin stimulated the migration and invasion of endometriotic cells (11Z, 12Z and 22B) in a dose-dependent manner. Leptin receptor (ObR) siRNA significantly inhibited the migration and invasion induced by leptin in 11Z and 12Z cells. Leptin-induced migration and invasion were significantly attenuated by pretreatment with SB-3CT, a specific gelatinase (MMP-2 and MMP-9) inhibitor. In addition, leptin-induced increases in the mRNA and protein expression and enzyme activity of MMP-2 in 11Z and 12Z cells. Selectively inhibiting MMP-2 using siRNA and an inhibitor (GM6003), impaired the ability of leptin to stimulate the migration and invasion of endometriotic cells, suggesting that MMP-2 plays an essential role in leptin-induced migration and invasion. Janus Kinase 2/Signal Transducer and Activator of Transcription 3 (JAK2/STAT3) inhibitor (AG490) significantly inhibited the migration, invasion and MMP-2 expression induced by leptin in endometriotic cells. Furthermore, the Extracellular signal-Regulated Kinase inhibitor PD98059 neutralized the migration and invasion promoting effects of leptin. Taken together, these results suggest that leptin may contribute to the migration and invasion abilities of endometriotic cells via the up-regulation of MMP-2 through an ObR-dependent JAK2/STAT3 signaling pathway. © The Author 2015. Published by Oxford University Press on behalf of the European Society of Human Reproduction and Embryology. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

MFAP5 promotes tumor progression and bone metastasis by regulating ERK/MMP signaling pathways in breast cancer.

PubMed

Wu, Zhiqiang; Wang, Ting; Fang, Meng; Huang, Wending; Sun, Zhengwang; Xiao, Jianru; Yan, Wangjun

2018-04-06

Breast cancer accounts for about 30% of all cancers in women, while approximately 70% breast cancer patients developed bone metastases throughout the course of their disease, highlighting the importance of exploring new therapeutic targets. Microfibrillar-associated protein 5 (MFAP5) is a component of extracellular elastic microfibril which has been confirmed to function in tissue development and cancer progression. But the role of MFAP5 in breast cancer remains unclear. The present study demonstrated that MFAP5 was up-regulated in breast cancers compared with that in normal breast tissues, and further increased in breast cancer bone metastasis. Functionally, MFAP5 overexpression accelerated breast cancer cell proliferation and migration, while an opposite effect was observed when MFAP5 was knocked down. In addition, up-regulation of MFAP5 increased the expression of MMP2 and MMP9 and activated the ERK signaling pathway. Conversely, inhibition of MFAP5 suppressed the expression of MMP2, MMP9, p-FAK, p-Erk1/2 and p-cJun. These findings may provide a better understanding about the mechanism of breast cancer and suggest that MFAP5 may be a potential prognostic biomarker and therapeutic target for breast cancer, especially for bone metastasis of breast cancer. Copyright © 2018 Elsevier Inc. All rights reserved.

Expression kinetics of metalloproteinases and their tissue inhibitors in experimental murine pulmonary tuberculosis.

PubMed

Ramos-Martínez, Ana G; Enciso-Moreno, José A; Espinosa-Ayala, Israel; Mata-Espinoza, Dulce; Rivas-Santiago, Bruno; Trujillo-Paez, Valentín; Monárrez-Espino, Joel; Hernández-Pando, Rogelio; Serrano, Carmen J

2015-02-01

Explore the temporal expression of metalloproteinases (MMPs) and tissue inhibitors of metalloproteinases (TIMPs) during experimental tuberculosis induced by virulent Mycobacterium tuberculosis strain H37Rv. BALB/c mice were infected via endotracheal instillation with H37Rv. Groups of mice were euthanized at different time points during infection. RNA was isolated from the lungs, and the expression of MMP-3, 8, 9, 10, 12, 13 and TIMP-1-4 was determined by quantitative PCR. Immunohistochemical detection of MMP-3, MMP-9, and MMP-10 was done to determine the cell source. The infection with H37Rv-induced inflammation resulted in maximal up-regulation of MMP-3, 8, 9, 10, 12 and 13 at day 21 postinfection. Additionally, MMP-13 showed another expression peak during late disease at day 60. Airway epithelium and macrophages were the most common MMP-3 and MMP-9 immunopositive cells, while for MMP-10, macrophages and endothelial cells were the most common, particularly at days 14 and 21 in well-formed granulomas. During late disease, vacuolated macrophages in pneumonic areas and bronchial epithelium showed mild MMP immunostaining. MMP-3, 8, 9, 10, 12, and 13 are maximally expressed at the peak of granuloma formation in the mouse tuberculosis model, with no compensation in levels or timing of TIMP expression. This data opens the possibility of participation of these molecules in the granuloma process.

Differentiated thyroid cancer cell invasion is regulated through epidermal growth factor receptor-dependent activation of matrix metalloproteinase (MMP)-2/gelatinase A

PubMed Central

Yeh, Michael W; Rougier, Jean-Philippe; Park, Jin-Woo; Duh, Quan-Yang; Wong, Mariwil; Werb, Zena; Clark, Orlo H

2008-01-01

Mechanisms of invasion in thyroid cancer remain poorly understood. We hypothesized that signaling via the epidermal growth factor receptor (EGFR) stimulates thyroid cancer cell invasion by altering the expression and cleavage of matrix metalloproteinases (MMPs). Papillary and follicular carcinoma cell lines were treated with EGF, the EGFR tyrosine kinase inhibitor AG1478, and the MMP inhibitors GM-6001 and Col-3. Flow cytometry was used to detect EGFR. In vitro invasion assays, gelatin zymography, and quantitative reverse transcription-PCR were used to assess the changes in invasive behavior and MMP expression and activation. All cell lines were found to overexpress functional EGFR. EGF stimulated invasion by thyroid cancer cells up to sevenfold (P<0.0001), a process that was antagonized completely by AG1478 and Col-3, partially by GM-6001, but not by the serine protease inhibitor aprotinin. EGF upregulated expression of MMP-9 (2.64– to 8.89-fold, P<0.0001) and membrane type-1 MMP (MT1-MMP, 1.97- to 2.67-fold, P<0.0001). This effect was blocked completely by AG1478 and partially by Col-3. The activation of MMP-2 paralleled MT1-MMP expression. We demonstrate that MMPs are critical effectors of invasion in the papillary and follicular thyroid cancer cell lines studied. Invasion is regulated by signaling through EGFR, an effect mediated by augmentation of gelatinase expression and activation. MMP inhibitors and growth factor antagonists may be effective tumoristatic agents for the treatment of aggressive thyroid carcinomas. PMID:17158762

The inhibitor of kappa B kinase-epsilon regulates MMP-3 expression levels and can promote lung metastasis.

PubMed

Seccareccia, E; Pinard, M; Wang, N; Li, S; Burnier, J; Dankort, D; Brodt, P

2014-08-18

The factors that determine the ability of metastatic tumor cells to expand and grow in specific secondary site(s) are not yet fully understood. Matrix metalloproteinases (MMP) were identified as potential regulators of the site-specificity of metastasis. We found that lung carcinoma cells ectopically expressing high levels of the receptor for the type I insulin like growth factor receptor (M27(R) cells) had a significant reduction in MMP-3 expression levels and this coincided with reduced metastasis to the lung. We used these cells to further investigate signaling pathways regulating MMP-3 expression and the role that MMP-3 plays in lung metastasis. We show that ectopic IκB kinase ɛ (IKKɛ) expression in these cells partly restored MMP-3 expression levels and also sensitized MMP-3 transcription to induction by phorbol 12-myristate 13-acetate (PMA). This increase in MMP-3 production was due to increased activation of several signal transduction mediators, including protein kinase C alpha, ERK2, Akt and the transcription factor p65. Furthermore, reconstitution of MMP-3 expression in M27(R) cells restored their ability to colonize the lung whereas silencing of MMP-3 in M27 cells reduced metastases. Collectively, our results implicate IKKɛ as a central regulator of PMA-induced cell signaling and MMP-3 expression and identify MMP-3 as an enabler of tumor cell expansion in the lung.Oncogenesis (2014) 3, e116; doi:10.1038/oncsis.2014.28; published online 18 August 2014.

Zinc improves learning and memory abilities of fetal growth restriction rats and promotes trophoblast cell invasion and migration via enhancing STAT3-MMP-2/9 axis activity.

PubMed

Zong, Lu; Wei, Xiaohua; Gou, Wenli; Huang, Pu; Lv, Ye

2017-12-29

Fetal growth restriction (FGR) is a well-known risk factor for cognitive dysfunction, especially for learning and memory abilities. However, knowledge about prevention and treatment methods of learning and memory abilities of fetal are limit. Here, Morris water maze and passive avoidance tests showed zinc supplementation could protect the impairment of the learning and memory abilities caused by FGR. As accumulating evidence suggested that insufficiency of placental trophoblast cell invasion was closely related to FGR fetal neurodevelopmental dysplasia, we further explored the relationship between zinc supplementation during pregnancy and placental trophoblast. Microarray identified 346 differently expressed genes in placental tissues with and without zinc supplementation, and GO and KEGG analyses showed these differently expressed genes were highly enriched in cell invasion and migration and STAT3 pathway. Protein-protein interaction(PPI) analysis found that STAT3 interacted with matrix metalloproteinase-2/9 (MMP-2/9). In vivo , western blot results authenticated that the expression levels of phospho-STAT3, STAT3, MMP-2 and MMP-9 were up-regulated in placental tissues after zinc treatment. To validate whether zinc could promotes trophoblast cell invasion and migration via enhancing STAT3-MMP-2/9 activity. In vitro , Transwell assay was performed, and we observed that abilities of invasion and migration were obviously increased in zinc treated trophoblast cells. And phospho-STAT3, STAT3, MMP-2 and MMP-9 expression levels were correspondingly increased in zinc treated trophoblast cells, which were dose-dependent. Moreover, gain-of-function and loss-of-function of STAT3 confirmed that zinc promotes cell invasion and migration via regulating STAT3 mediated up-regulation of MMP-2/9 activity. We propose that activation of MMP-2/9 mediated by STAT3 may contribute to invasion and migration of trophoblast cells, which improved neurodevelopmental impairment of FGR rats probably via contributing to placental development. Our findings are the first to show a possible mechanism of reversing neurodevelopmental impairment of FGR rats by zinc supplementation, holding promise for the development of novel therapeutic modalities for learning and memory abilities impairment caused by FGR.

Suppression of alpha-tocopherol ether-linked acetic acid in VEGF-induced angiogenesis and the possible mechanisms in human umbilical vein endothelial cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Chuang, Cheng-Hung, E-mail: chchuang@hk.edu.tw; Liu, Chia-Hua; Lu, Ta-Jung

2014-12-15

Alpha-tocopherol ether-linked acetic acid (α-TEA) has been reported to exhibit both anti-tumor and anti-metastatic activities in cell culture and animal studies. However, it is unclear whether α-TEA possesses anti-angiogenic effects. In this study, we investigated the effect of α-TEA on vascular endothelial growth factor (VEGF)-induced angiogenesis and matrix metalloproteinase (MMP) expression both in vitro and ex vivo. We found that the α-TEA inhibited tube formation, invasion, and migration in human umbilical vein endothelial cells (HUVECs) and that such actions were accompanied by reduced expression of MMP-2. α-TEA also inhibited ex vivo angiogenesis, as indicated by chicken egg chorioallantoic membrane assay.more » We further showed that α-TEA attenuated protein expression of VEGF receptor-2 (VEGFR-2)-mediated p38 mitogen-activated protein kinase (p38), phosphorylated p38, and focal adhesion kinase (FAK). Moreover, α-TEA (30 μM) significantly up-regulated protein expression of tissue inhibitors of MMP (TIMP)-2 (by 138%) and the metastasis suppressor gene nm23-H1 (by 54%). These results demonstrate that the anti-angiogenic effect of α-TEA both in vitro and ex vivo and its possible mechanistic action appears to involve the inhibition of MMP-2 level through VEGFR-2-mediated FAK and p38 signaling pathways and through up-regulation of TIMP-2 and nm23-H1 expression. - Graphical abstract: Possible mechanisms of α-TEA on inhibited angiogenesis of human umbilical vein endothelial cells. Brief summary In the present study, we have demonstrated that VEGF-mediated angiogenesis is significantly inhibited by α-TEA, and that this effect involves inhibition of MMP-2 level through VEGFR-2-mediated FAK and p38 signaling pathways related to invasion and migration. - Highlights: • The anti-angiogenic effect and the mechanistic action of α-TEA were investigated. • α-TEA significantly inhibited VEGF-mediated angiogenesis both in vitro and ex vivo. • α-TEA down-regulated MMP-2 via VEGFR-2-mediated FAK and p38 signaling pathways. • α-TEA up-regulated TIMP-2 and nm23-H1 expression in relation to invasion and migration. • Further studies are warranted on the anti-angiogenesis potential of α-TEA.« less

NOV/CCN3 attenuates inflammatory pain through regulation of matrix metalloproteinases-2 and -9

PubMed Central

2012-01-01

Background Sustained neuroinflammation strongly contributes to the pathogenesis of pain. The clinical challenge of chronic pain relief led to the identification of molecules such as cytokines, chemokines and more recently matrix metalloproteinases (MMPs) as putative therapeutic targets. Evidence points to a founder member of the matricial CCN family, NOV/CCN3, as a modulator of these inflammatory mediators. We thus investigated the possible involvement of NOV in a preclinical model of persistent inflammatory pain. Methods We used the complete Freund's adjuvant (CFA)-induced model of persistent inflammatory pain and cultured primary sensory neurons for in vitro experiments. The mRNA expression of NOV and pro-inflammatory factors were measured with real-time quantitative PCR, CCL2 protein expression was assessed using ELISA, MMP-2 and -9 activities using zymography. The effect of drugs on tactile allodynia was evaluated by the von Frey test. Results NOV was expressed in neurons of both dorsal root ganglia (DRG) and dorsal horn of the spinal cord (DHSC). After intraplantar CFA injection, NOV levels were transiently and persistently down-regulated in the DRG and DHSC, respectively, occurring at the maintenance phase of pain (15 days). NOV-reduced expression was restored after treatment of CFA rats with dexamethasone. In vitro, results based on cultured DRG neurons showed that siRNA-mediated inhibition of NOV enhanced IL-1β- and TNF-α-induced MMP-2, MMP-9 and CCL2 expression whereas NOV addition inhibited TNF-α-induced MMP-9 expression through β1 integrin engagement. In vivo, the intrathecal delivery of MMP-9 inhibitor attenuated mechanical allodynia of CFA rats. Importantly, intrathecal administration of NOV siRNA specifically led to an up-regulation of MMP-9 in the DRG and MMP-2 in the DHSC concomitant with increased mechanical allodynia. Finally, NOV intrathecal treatment specifically abolished the induction of MMP-9 in the DRG and, MMP-9 and MMP-2 in the DHSC of CFA rats. This inhibitory effect on MMP is associated with reduced mechanical allodynia. Conclusions This study identifies NOV as a new actor against inflammatory pain through regulation of MMPs thus uncovering NOV as an attractive candidate for therapeutic improvement in pain relief. PMID:22353423

High MMP-9 activity levels in fragile X syndrome are lowered by minocycline.

PubMed

Dziembowska, Magdalena; Pretto, Dalyir I; Janusz, Aleksandra; Kaczmarek, Leszek; Leigh, Mary Jacena; Gabriel, Nielsen; Durbin-Johnson, Blythe; Hagerman, Randi J; Tassone, Flora

2013-08-01

Fragile X syndrome (FXS) is a neurodevelopmental disorder characterized by lack of the FMR1 protein, FMRP, a translational repressor. Its absence leads to up-regulation of locally translated proteins involved in synaptic transmission and plasticity, including the matrix metalloproteinase-9 (MMP-9). In the Fmr1 knock-out (KO), a mouse model of FXS, an abnormal elevated expression of MMP-9 in the brain was pharmacologically down-regulated after treatment with the tetracycline derivative minocycline. Moreover, the rescue of immature dendritic spine morphology and a significant improvement of abnormal behavior were associated with down-regulation of MMP-9. Here, we report on high plasma activity of MMP-9 in individuals with FXS. In addition, we investigate MMP-9 changes in patients with FXS who have gone through a minocycline controlled clinical trial and correlate MMP-9 activity to clinical observations. The results of this study suggest that, in humans, activity levels of MMP-9 are lowered by minocycline and that, in some cases, changes in MMP-9 activity are positively associated with improvement based on clinical measures. Copyright © 2013 Wiley Periodicals, Inc.

Long noncoding RNA LINC00460 targets miR-539/MMP-9 to promote meningioma progression and metastasis.

PubMed

Xing, Hongshun; Wang, Shilei; Li, Quancai; Ma, Yongqian; Sun, Peng

2018-06-12

Emerging evidence reveal that long noncoding RNAs (lncRNAs) participates in the epigenetic regulation of pathophysiological process. However, the deepgoing role of lncRNAs on meningioma is still unclear. In present study, we investigate the roles of lncRNA LINC00460 in meningeoma tissue and uncover its molecular mechanism. Results revealed that LINC00460 expression level was significantly up-regulated in meningeoma tissues and malignant meningeoma cell lines (IOMM-Lee, CH157-MN). Mechanically, loss-of-function assays showed that LINC00460 knockdown significantly suppressed the proliferation ability, increased the apoptosis and decreased the proteins (MMP-2, MMP-9, ZEB1) expression. Bioinformatics tools predicted that miR-539 both targeted with the 3'-UTR of LINC00460 and MMP-9 mRNA, which was confirmed by luciferase reporter assay and western blot analysis. In summary, our study reveals that LINC00460 promotes MMP-9 expression through targeting miR-539, acting as an oncogenic RNA in the meningeoma malignancy and accelerating the proliferation and metastasis of meningeoma. Copyright © 2018 Elsevier Masson SAS. All rights reserved.

Hepatocyte produced matrix metalloproteinases are regulated by CD147 in liver fibrogenesis.

PubMed

Calabro, Sarah R; Maczurek, Annette E; Morgan, Alison J; Tu, Thomas; Wen, Victoria W; Yee, Christine; Mridha, Auvro; Lee, Maggie; d'Avigdor, William; Locarnini, Stephen A; McCaughan, Geoffrey W; Warner, Fiona J; McLennan, Susan V; Shackel, Nicholas A

2014-01-01

The classical paradigm of liver injury asserts that hepatic stellate cells (HSC) produce, remodel and turnover the abnormal extracellular matrix (ECM) of fibrosis via matrix metalloproteinases (MMPs). In extrahepatic tissues MMP production is regulated by a number of mechanisms including expression of the glycoprotein CD147. Previously, we have shown that CD147 is expressed on hepatocytes but not within the fibrotic septa in cirrhosis [1]. Therefore, we investigated if hepatocytes produce MMPs, regulated by CD147, which are capable of remodelling fibrotic ECM independent of the HSC. Non-diseased, fibrotic and cirrhotic livers were examined for MMP activity and markers of fibrosis in humans and mice. CD147 expression and MMP activity were co-localised by in-situ zymography. The role of CD147 was studied in-vitro with siRNA to CD147 in hepatocytes and in-vivo in mice with CCl4 induced liver injury using ãCD147 antibody intervention. In liver fibrosis in both human and mouse tissue MMP expression and activity (MMP-2, -9, -13 and -14) increased with progressive injury and localised to hepatocytes. Additionally, as expected, MMPs were abundantly expressed by activated HSC. Further, with progressive fibrosis there was expression of CD147, which localised to hepatocytes but not to HSC. Functionally significant in-vitro regulation of hepatocyte MMP production by CD147 was demonstrated using siRNA to CD147 that decreased hepatocyte MMP-2 and -9 expression/activity. Further, in-vivo α-CD147 antibody intervention decreased liver MMP-2, -9, -13, -14, TGF-β and α-SMA expression in CCl4 treated mice compared to controls. We have shown that hepatocytes produce active MMPs and that the glycoprotein CD147 regulates hepatocyte MMP expression. Targeting CD147 regulates hepatocyte MMP production both in-vitro and in-vivo, with the net result being reduced fibrotic matrix turnover in-vivo. Therefore, CD147 regulation of hepatocyte MMP is a novel pathway that could be targeted by future anti-fibrogenic agents.

Nucleolin mediated pro-angiogenic role of Hydroxysafflor Yellow A in ischaemic cardiac dysfunction: Post-transcriptional regulation of VEGF-A and MMP-9.

PubMed

Zou, Jiang; Wang, Nian; Liu, Manting; Bai, Yongping; Wang, Hao; Liu, Ke; Zhang, Huali; Xiao, Xianzhong; Wang, Kangkai

2018-05-01

Hydroxysafflor Yellow A (HSYA), a most representative ingredient of Carthamus tinctorius L., had long been used in treating ischaemic cardiovascular diseases in China and exhibited prominently anticoagulant and pro-angiogenic activities, but the underlying mechanisms remained largely unknown. This study aimed to further elucidate the pro-angiogenic effect and mechanism of HSYA on ischaemic cardiac dysfunction. A C57 mouse model of acute myocardial infarction (AMI) was firstly established, and 25 mg/kg HSYA was intraperitoneally injected immediately after operation and given once, respectively, each morning and evening for 2 weeks. It was found that HSYA significantly improved ischaemia-induced cardiac haemodynamics, enhanced the survival rate, alleviated the myocardial injury and increased the expressions of CD31, vascular endothelial growth factor-A (VEGF-A) and nucleolin in the ischaemic myocardium. In addition, HSYA promoted the migration and tube formation of human umbilical vein endothelial cells (HUVECs), enhanced the expressions of nucleolin, VEGF-A and matrix metalloproteinase-9 (MMP-9) in a dose- and time-dependent manner. However, down-regulation of nucleolin expression sharply abrogated the effect mentioned above of HSYA. Further protein-RNA coimmunoprecipitation and immunoprecipitation-RT-PCR assay showed that nucleolin binded to VEGF-A and MMP-9 mRNA and overexpression of nucleolin up-regulated the mRNA expressions of VEGF-A and MMP-9 in the HUVECs through enhancing the stability of VEGF-A and MMP-9 mRNA. Furthermore, HSYA increased the mRNA expressions of VEGF-A and MMP-9 in the extract of antinucleolin antibody-precipitated protein from the heart of AMI mice. Our data revealed that nucleolin mediated the pro-angiogenic effect of HSYA through post-transcriptional regulation of VEGF-A and MMP-9 expression, which contributed to the protective effect of HSYA on ischaemic cardiac dysfunction. © 2018 The Authors. Journal of Cellular and Molecular Medicine published by John Wiley & Sons Ltd and Foundation for Cellular and Molecular Medicine.

Inhibition of pancreatic stellate cell activity by adipose-derived stem cells.

PubMed

Yu, Fu-Xiang; Su, Long-Feng; Dai, Chun-Lei; Wang, Yang; Teng, Yin-Yan; Fu, Jun-Hui; Zhang, Qi-Yu; Tang, Yin-He

2015-04-01

Pancreatic stellate cells (PSCs) play a critical role in the development of pancreatic fibrosis. In this study we used a novel method to isolate and culture rat PSCs and then investigated the inhibitory effects of adipose-derived stem cells (ADSCs) on activation and proliferation of PSCs. Pancreatic tissue was obtained from Sprague-Dawley rats for PSCs isolation. Transwell cell cultures were adopted for co-culture of ADSCs and PSCs. PSCs proliferation and apoptosis were determined using CCK-8 and flow cytometry, respectively. alpha-SMA expressions were analyzed using Western blotting. The levels of cytokines [nerve growth factor (NGF), interleukin-10 (IL-10) and transforming growth factor-beta1 (TGF-beta1)] in conditioned medium were detected by ELISA. Gene expression (MMP-2, MMP-9 and TIMP-1) was analyzed using qRT-PCR. This method produced 17.6+/-6.5X10(3) cells per gram of the body weight with a purity of 90%-95% and a viability of 92%-97%. Co-culture of PSCs with ADSCs significantly inhibited PSCs proliferation and induced PSCs apoptosis. Moreover, alpha-SMA expression was significantly reduced in PSCs+ADSCs compared with that in PSC-only cultures, while expression of fibrinolytic proteins (e.g., MMP-2 and MMP-9) was up-regulated and anti-fibrinolytic protein (TIMP-1) was down-regulated. In addition, NGF expression was up-regulated, but IL-10 and TGF-beta1 expressions were down-regulated in the co-culture conditioned medium compared with those in the PSC-only culture medium. This study provided an easy and reliable technique to isolate PSCs. The data demonstrated the inhibitory effects of ADSCs on the activation and proliferation of PSCs in vitro.

Perfluorooctanoic acid stimulates ovarian cancer cell migration, invasion via ERK/NF-κB/MMP-2/-9 pathway.

PubMed

Li, Xiaozhao; Bao, Chunyu; Ma, Zhinan; Xu, Boqun; Liu, Xiaoqiu; Ying, Xiaoyan; Zhang, Xuesen

2018-05-09

As widely used in consumer products, perfluorooctanoic acid (PFOA) has become a common environmental pollutant, which has been detected in human serum and associated with cancers. Our previous study showed that PFOA is a carcinogen that promotes endometrial cancer cell migration and invasion through activation of ERK/mTOR signaling. Here, we showed that PFOA (≥100 nM) treatment also stimulated A2780 ovarian cancer cell invasion and migration, which correlated with increased matrix metalloproteinases MMP-2/-9 expression, important proteases associated with tumor invasion and migration. Notably, PFOA treatment induced activation of ERK1/2/ NF-κB signaling. Pre-treatment with U0126, an ERK1/2inhibitor；or JSH-23, a NF-kB inhibitor, can reverse the PFOA-induced cell migration and invasion. Consistent with these results, inhibiting ERK1/2 or NF-κB signaling abolished PFOA-induced up-regulation of MMP-2/-9 expression. These results indicate that PFOA can stimulate ovarian cancer cell migration, invasion and MMP-2/-9 expression by up-regulating ERK/NF-κB pathway. Copyright © 2018 Elsevier B.V. All rights reserved.

Unique proliferation response in odontoblastic cells derived from human skeletal muscle stem cells by cytokine-induced matrix metalloproteinase-3

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ozeki, Nobuaki; Hase, Naoko; Kawai, Rie

A pro-inflammatory cytokine mixture (CM: interleukin (IL)-1β, tumor necrosis factor-α and interferon-γ) and IL-1β-induced matrix metalloproteinase (MMP)-3 activity have been shown to increase the proliferation of rat dental pulp cells and murine stem cell-derived odontoblast-like cells. This suggests that MMP-3 may regulate wound healing and regeneration in the odontoblast-rich dental pulp. Here, we determined whether these results can be extrapolated to human dental pulp by investigating the effects of CM-induced MMP-3 up-regulation on the proliferation and apoptosis of purified odontoblast-like cells derived from human skeletal muscle stem cells. We used siRNA to specifically reduce MMP-3 expression. We found that CMmore » treatment increased MMP-3 mRNA and protein levels as well as MMP-3 activity. Cell proliferation was also markedly increased, with no changes in apoptosis, upon treatment with CM and following the application of exogenous MMP-3. Endogenous tissue inhibitors of metalloproteinases were constitutively expressed during all experiments and unaffected by MMP-3. Although treatment with MMP-3 siRNA suppressed cell proliferation, it also unexpectedly increased apoptosis. This siRNA-mediated increase in apoptosis could be reversed by exogenous MMP-3. These results demonstrate that cytokine-induced MMP-3 activity regulates cell proliferation and suppresses apoptosis in human odontoblast-like cells. - Highlights: • Pro-inflammatory cytokines induce MMP-3 activity in human odontoblast-like cells. • Increased MMP-3 activity can promote cell proliferation in odontoblasts. • Specific loss of MMP-3 increases apoptosis in odontoblasts. • MMP-3 has potential as a promising new target for pupal repair and regeneration.« less

Protection against murine osteoarthritis by inhibition of the 26S proteasome and lysine-48 linked ubiquitination.

PubMed

Radwan, Marta; Wilkinson, David J; Hui, Wang; Destrument, Auriane P M; Charlton, Sarah H; Barter, Matt J; Gibson, Beth; Coulombe, Josée; Gray, Douglas A; Rowan, Andrew D; Young, David A

2015-08-01

To determine whether the process of ubiquitination and/or activity of the 26S proteasome are involved in the induction of osteoarthritis (OA). Bovine cartilage resorption assays, chondrocyte cell-line SW1353 and primary human articular chondrocytes were used with the general proteasome inhibitor MG132 or vehicle to identify a role of the ubiquitin-proteasome system (UPS) in cartilage destruction and matrix metalloproteinase-13 (MMP13) expression. In vivo, MG132 or vehicle, were delivered subcutaneously to mice following destabilisation of the medial meniscus (DMM)-induced OA. Subsequently, DMM was induced in Lys-to-Arg (K48R and K63R) mutant ubiquitin (Ub) transgenic mice. Cytokine signalling in SW1353s was monitored by immunoblotting and novel ubiquitinated substrates identified using Tandem Ubiquitin Binding Entities purification followed by mass spectrometry. The ubiquitination of TRAFD1 was assessed via immunoprecipitation and immunoblotting and its role in cytokine signal-transduction determined using RNA interference and real-time RT-PCR for MMP13 and interleukin-6 (IL6). Supplementation with the proteasome inhibitor MG132 protected cartilage from cytokine-mediated resorption and degradation in vivo in mice following DMM-induced OA. Using transgenic animals only K48R-mutated Ub partially protected against OA compared to wild-type or wild-type Ub transgenic mice, and this was only evident on the medial femoral condyle. After confirming ubiquitination was vital for NF-κB signalling and MMP13 expression, a screen for novel ubiquitinated substrates involved in cytokine-signalling identified TRAFD1; the depletion of which reduced inflammatory mediator-induced MMP13 and IL6 expression. Our data for the first time identifies a role for ubiquitination and the proteasome in the induction of OA via regulation of inflammatory mediator-induced MMP13 expression. These data open avenues of research to determine whether the proteasome, or K48-linked ubiquitination, are potential therapeutic targets in OA. Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://group.bmj.com/group/rights-licensing/permissions.

MiR-26b Mimic Inhibits Glioma Proliferation In Vitro and In Vivo Suppressing COX-2 Expression.

PubMed

Chen, Zheng-Gang; Zheng, Chuan-Yi; Cai, Wang-Qing; Li, Da-Wei; Ye, Fu-Yue; Zhou, Jian; Wu, Ran; Yang, Kun

2017-08-11

Glioma is the most common malignant tumor of the nervous system. Studies have shown the microRNA (miR)-26b/cyclooxygenase (COX)-2 axis in the development and progression in many tumor cells. Our study aims to investigate the effect and mechanism of miR-26b/COX-2 axis in glioma. Decreased expression of miR-26b with increased level of COX-2 was found in glioma tissues compared with matched normal tissues. A strong negative correlation was observed between the level of miR-26b and COX-2 in 30 glioma tissues. The miR-26b was then overexpressed by transfecting miR-26b mimic into U-373 cells. The invasive cell number and wounld closing rate were reduced in U-373 cells transfected with miR-26b mimic. Besides, COX2 siRNA enhanced the effect of miR-26b mimic in suppressing the expression of p-ERK1 and p-JNK. Finally, the in vivo experiment revealed that miR-26b mimic transfection strongly reduced the tumor growth, tumor volume and the expression of matrix metalloproteinase (MMP)-2, MMP-9). Taken together, our research indicated a miR-26b/COX-2/ERK/JNK axis in regulating the motility of glioma in vitro and in vivo, providing a new sight for treatment of glioma.

Leptin-promoted human extravillous trophoblast invasion is MMP14 dependent and requires the cross talk between Notch1 and PI3K/Akt signaling.

PubMed

Wang, Huayang; Cheng, Huanhuan; Shao, Qianqian; Dong, Zhaogang; Xie, Qi; Zhao, Lei; Wang, Qingjie; Kong, Beihua; Qu, Xun

2014-04-01

The overexpression of leptin is a crucial feature for the maintenance of pregnancy. The effects of leptin on trophoblast invasion are important to its reproductive function, but the underlying mechanisms remain poorly understood. MMP14 is a member of matrix metalloproteinase (MMP) family that is closely involved in the invasion process. Here, we characterized the importance of MMP14 in the proinvasion effect of leptin on EVT cells and elucidated its molecular mechanisms. Transwell assay revealed that leptin promoted invasion of the immortalized EVT cell line HTR-8/SVneo in a dose- and time-related fashion. Further studies suggested that leptin enhanced HTR-8/SVneo cell invasion by up-regulating MMP14 expression and that knockdown of MMP14 by small interference RNA (siRNA) blocked the proinvasion effect of leptin. Notably, leptin promoted the expression of Notch1 receptor and activated its signaling in HTR-8/SVneo cells, and blocking this pathway by siRNA inhibited both leptin-enhanced MMP14 expression and invasiveness of HTR-8/SVneo cells. Such effects of Notch1 signaling were related with the activation of the PI3K/Akt pathway, which was significantly activated after leptin stimulation and was interfered by Notch1 signaling perturbation. Taken together, our observations suggest that leptin is an effective regulator of MMP14 expression, which consequently plays critical roles in invasion of EVT cells. The promoting effects of leptin on MMP14 require the cross talk between Notch1 and PI3K/Akt signaling pathways.

Age-associated reduction of cellular spreading/mechanical force up-regulates matrix metalloproteinase-1 expression and collagen fibril fragmentation via c-Jun/AP-1 in human dermal fibroblasts.

PubMed

Qin, Zhaoping; Voorhees, John J; Fisher, Gary J; Quan, Taihao

2014-12-01

The dermal compartment of human skin is largely composed of dense collagen-rich fibrils, which provide structural and mechanical support. Skin dermal fibroblasts, the major collagen-producing cells, are interact with collagen fibrils to maintain cell spreading and mechanical force for function. A characteristic feature of aged human skin is fragmentation of collagen fibrils, which is initiated by matrix metalloproteinase 1 (MMP-1). Fragmentation impairs fibroblast attachment and thereby reduces spreading. Here, we investigated the relationship among fibroblast spreading, mechanical force, MMP-1 expression, and collagen fibril fragmentation. Reduced fibroblast spreading due to cytoskeletal disruption was associated with reduced cellular mechanical force, as determined by atomic force microscopy. These reductions substantially induced MMP-1 expression, which led to collagen fibril fragmentation and disorganization in three-dimensional collagen lattices. Constraining fibroblast size by culturing on slides coated with collagen micropatterns also significantly induced MMP-1 expression. Reduced spreading/mechanical force induced transcription factor c-Jun and its binding to a canonical AP-1 binding site in the MMP-1 proximal promoter. Blocking c-Jun function with dominant negative mutant c-Jun significantly reduced induction of MMP-1 expression in response to reduced spreading/mechanical force. Furthermore, restoration of fibroblast spreading/mechanical force led to decline of c-Jun and MMP-1 levels and eliminated collagen fibril fragmentation and disorganization. These data reveal a novel mechanism by which alteration of fibroblast shape/mechanical force regulates c-Jun/AP-1-dependent expression of MMP-1 and consequent collagen fibril fragmentation. This mechanism provides a foundation for understanding the cellular and molecular basis of age-related collagen fragmentation in human skin. © 2014 The Authors. Aging Cell published by the Anatomical Society and John Wiley & Sons Ltd.

SIPA1 promotes invasion and migration in human oral squamous cell carcinoma by ITGB1 and MMP7

DOE Office of Scientific and Technical Information (OSTI.GOV)

Takahara, Toshikazu; Kasamatsu, Atsushi, E-mail: kasamatsua@faculty.chiba-u.jp; Yamatoji, Masanobu

Signal-induced proliferation-associated protein 1 (SIPA1) is known to be a GTPase activating protein. Overexpressed SIPA1 is related to metastatic progression in breast and prostate cancers; however, the relevance of SIPA1 in oral squamous cell carcinoma (OSCC) is still unknown. The aim of this study was to examine SIPA1 expression and its functional mechanisms in OSCC. SIPA1 mRNA and protein expressions were analyzed by quantitative reverse transcriptase-polymerase chain reaction, Western blot analysis, and immunohistochemistry. The expressions of SIPA1 were up-regulated significantly in vitro and in vivo. Moreover, SIPA1 expression was correlated with regional lymph node metastasis. We next assessed the cellularmore » functions associated with tumoral metastasis using SIPA1 knockdown (shSIPA1) cells and analyzed the downstream molecules of SIPA1, i.e., bromodomain containing protein 4(BRD4), integrin beta1 (ITGB1), and matrix metalloproteinase 7 (MMP7). The shSIPA1 cells showed decreased invasiveness and migratory activities, however cellular adhesion ability was maintained at a high level. In addition, ITGB1 expression was greater in shSIPA1 cells, whereas MMP7 expression was lower than in control cells. This research is the first to establish that SIPA1 promotes cancer metastasis by regulating the ITGB1 and MMP7. Therefore, SIPA1 might be a novel therapeutic target for patients with lymph node metastasis of OSCC. - Highlights: • SIPA1 expression was up-regulated in oral squamous cell carcinoma (OSCC). • SIPA1-positive OSCCs were correlated with regional lymph node metastasis. • SIPA1 controlled BRD4 and influenced transcription of ITGB1and MMP7. • SIPA1 induced cellular invasion and migration and decreased cellular adhesion. • SIPA1 might be a potential biomarker of cancer metastasis for OSCC.« less

MicroRNA-21 promotes bone mesenchymal stem cells migration in vitro by activating PI3K/Akt/MMPs pathway.

PubMed

Lv, Chen; Yang, Shengwu; Chen, Xin; Zhu, Xiongbai; Lin, Wenjun; Wang, Lu; Huang, Zhengxiang; Wang, Mingyue; Tu, Guanjun

2017-12-01

MicroRNA-21 (miR-21) contributes to anti-apoptosis in bone marrow mesenchymal stem cells (BMSC), but its role in the migration of BMSCs remains vague. The aim of this study was to determine the possible effect of miR-21 on regulating BMSCs directional migration and the expression of MMP-2/MMP-9 in BMSCs in vitro. BMSCs were successfully infected with miR-21-up lentivirus. Cell migration using Transwell assay indicated that upregulated expression of miR-21 could significantly promote BMSCs migration. Western blot analysis indicated that miR-21 significantly upregulated the expression of MMP-2 and MMP-9, which were related to metastasis-associated genes. GM6001, the specific MMPs inhibitor, abrogated the upregulated expression of MMP-2/MMP-9 and abolished the positive effect of miR-21 on promoting BMSCs migration. Meanwhile, miR-21 significantly enhanced Akt phosphorylation, as measured by Western blot analysis. LY294002, an inhibitor of Akt activation, abrogated the phosphorylation of Akt and abolished the positive effect of miR-21 on promoting BMSCs migration and upregulating MMP-2/MMP-9 expression. These results suggest that miR-21 contributes to BMSCs migration by upregulating MMP-2/MMP-9, potentially via the PI3K/Akt pathway. Copyright © 2017 Elsevier Ltd. All rights reserved.

Steroid and cytokine regulation of matrix metalloproteinase expression in endometriosis and the establishment of experimental endometriosis in nude mice.

PubMed

Bruner-Tran, Kaylon L; Eisenberg, Esther; Yeaman, Grant R; Anderson, Ted A; McBean, Judith; Osteen, Kevin G

2002-10-01

The cyclic expression of matrix metalloproteinases (MMPs) by human endometrium has been suggested to play a role in the invasive process necessary to establish endometriosis. The ability of progesterone exposure to inhibit endometrial MMP-3 and MMP-7 expression requires the local action of TGF beta and may also be linked to the local production of retinoic acid by stromal cells. A continuous expression of several MMPs in endometriotic lesions has been reported, indicating a failure of progesterone or locally produced factors to suppress these enzymes. To address cell-specific MMP regulation associated with endometriosis, we examined expression of MMP-3 and MMP-7 mRNA in eutopic endometrium and endometriotic lesions acquired during the secretory phase of the menstrual cycle. We examined the in vitro regulation of MMP-3 and MMP-7 protein in similar tissues. We also examined the in vitro regulation of MMP secretion by progesterone, retinoic acid, and TGF beta in endometriosis tissues relative to the establishment of experimental disease. Our studies indicate that either eutopic or ectopic tissue from women with endometriosis exhibit patterns of altered MMP regulation in vivo. A lack of responsiveness to progesterone was demonstrated in vitro, associated with a failure to suppress MMP expression and an enhanced ability of the tissue to establish experimental endometriosis. However, in vitro treatments with retinoic acid and TGF beta restored the ability of progesterone to suppress MMPs in vitro and prevented the establishment of experimental disease.

Arctigenin Inhibits Lung Metastasis of Colorectal Cancer by Regulating Cell Viability and Metastatic Phenotypes.

PubMed

Han, Yo-Han; Kee, Ji-Ye; Kim, Dae-Seung; Mun, Jeong-Geon; Jeong, Mi-Young; Park, Sang-Hyun; Choi, Byung-Min; Park, Sung-Joo; Kim, Hyun-Jung; Um, Jae-Young; Hong, Seung-Heon

2016-08-27

Arctigenin (ARC) has been shown to have an anti-cancer effect in various cell types and tissues. However, there have been no studies concerning metastatic colorectal cancer (CRC). In this study, we investigated the anti-metastatic properties of ARC on colorectal metastasis and present a potential candidate drug. ARC induced cell cycle arrest and apoptosis in CT26 cells through the intrinsic apoptotic pathway via MAPKs signaling. In several metastatic phenotypes, ARC controlled epithelial-mesenchymal transition (EMT) through increasing the expression of epithelial marker E-cadherin and decreasing the expressions of mesenchymal markers; N-cadherin, vimentin, β-catenin, and Snail. Moreover, ARC inhibited migration and invasion through reducing of matrix metalloproteinase-2 (MMP-2) and MMP-9 expressions. In an experimental metastasis model, ARC significantly inhibited lung metastasis of CT26 cells. Taken together, our study demonstrates the inhibitory effects of ARC on colorectal metastasis.

Anti-arthritic effects of chlorogenic acid in interleukin-1β-induced rabbit chondrocytes and a rabbit osteoarthritis model.

PubMed

Chen, Wei-Ping; Tang, Jing-Li; Bao, Jia-Peng; Hu, Peng-Fei; Shi, Zhong-Li; Wu, Li-Dong

2011-01-01

Cartilage degradation is one of the pathological changes of osteoarthritis (OA), and accumulating evidence suggests an excess of matrix metalloproteinases (MMPs) plays a role in this cartilage breakdown. Here, we investigated the effects of chlorogenic acid (CGA) on the mRNA and protein expression of MMPs in interleukin (IL)-1β-induced rabbit chondrocytes and evaluated the in vivo effects of CGA in experimental OA induced by anterior cruciate ligament transection (ACLT) in rabbits. Using quantitative real-time PCR and ELISA to investigate the expression levels of MMP-1, MMP-3, MMP-13, and tissue inhibitors of metalloproteinase-1(TIMP-1) in IL-1β-induced rabbit chondrocytes, we showed that CGA inhibits the expression of these MMPs while increasing TIMP-1 expression, at both the mRNA and protein levels. In addition, IL-1β-induced activation of nuclear factor kappa B (NF-κB) and the degradation of inhibitor of κB (IκB)-α were suppressed by CGA. In rabbits, CGA decreased cartilage degradation as assessed by morphological and histological analyses. The down-regulation of MMP-1, MMP-3, and MMP-13 expression and up-regulation of TIMP-1 expression were also detected in CGA-treated cartilage compared with vehicle-treated cartilage, confirming these findings in an in vivo model. Taken together, these findings indicate that CGA may be considered as a possible candidate agent in the treatment of OA. Copyright © 2010 Elsevier B.V. All rights reserved.

PER, a Circadian Clock Component, Mediates the Suppression of MMP-1 Expression in HaCaT Keratinocytes by cAMP.

PubMed

Yeom, Miji; Lee, HansongI; Shin, Seoungwoo; Park, Deokhoon; Jung, Eunsun

2018-03-23

Skin circadian clock system responds to daily changes, thereby regulating skin functions. Exposure of the skin to UV irradiation induces the expression of matrix metalloproteinase-1 (MMP-1) and causes DNA damage. It has been reported both DNA repair and DNA replication are regulated by the circadian clock in mouse skin. However, the molecular link between circadian clock and MMP-1 has little been investigated. We found PERIOD protein, a morning clock component, represses the expression of MMP-1 in human keratinocytes by using a PER-knockdown strategy. Treatment with siPer3 alleviated the suppression of MMP-1 expression induced by forskolin. Results revealed PER3 suppresses the expression of MMP-1 via cAMP signaling pathway. Additionally, we screened for an activator of PER that could repress the expression of MMP-1 using HaCaT cell line containing PER promoter-luciferase reporter gene. Results showed Lespedeza capitate extract (LCE) increased PER promoter activity. LCE inhibited the expression of MMP-1 and its effect of LCE was abolished in knockdown of PER2 or PER3, demonstrating LCE can repress the expression of MMP-1 through PER. Since circadian clock component PER can regulate MMP-1 expression, it might be a new molecular mechanism to develop therapeutics to alleviate skin aging and skin cancer.

Hepatocyte Produced Matrix Metalloproteinases Are Regulated by CD147 in Liver Fibrogenesis

PubMed Central

Morgan, Alison J.; Tu, Thomas; Wen, Victoria W.; Yee, Christine; Mridha, Auvro; Lee, Maggie; d'Avigdor, William; Locarnini, Stephen A.; McCaughan, Geoffrey W.; Warner, Fiona J.; McLennan, Susan V.; Shackel, Nicholas A.

2014-01-01

Background The classical paradigm of liver injury asserts that hepatic stellate cells (HSC) produce, remodel and turnover the abnormal extracellular matrix (ECM) of fibrosis via matrix metalloproteinases (MMPs). In extrahepatic tissues MMP production is regulated by a number of mechanisms including expression of the glycoprotein CD147. Previously, we have shown that CD147 is expressed on hepatocytes but not within the fibrotic septa in cirrhosis [1]. Therefore, we investigated if hepatocytes produce MMPs, regulated by CD147, which are capable of remodelling fibrotic ECM independent of the HSC. Methods Non-diseased, fibrotic and cirrhotic livers were examined for MMP activity and markers of fibrosis in humans and mice. CD147 expression and MMP activity were co-localised by in-situ zymography. The role of CD147 was studied in-vitro with siRNA to CD147 in hepatocytes and in-vivo in mice with CCl4 induced liver injury using ãCD147 antibody intervention. Results In liver fibrosis in both human and mouse tissue MMP expression and activity (MMP-2, -9, -13 and -14) increased with progressive injury and localised to hepatocytes. Additionally, as expected, MMPs were abundantly expressed by activated HSC. Further, with progressive fibrosis there was expression of CD147, which localised to hepatocytes but not to HSC. Functionally significant in-vitro regulation of hepatocyte MMP production by CD147 was demonstrated using siRNA to CD147 that decreased hepatocyte MMP-2 and -9 expression/activity. Further, in-vivo α-CD147 antibody intervention decreased liver MMP-2, -9, -13, -14, TGF-β and α-SMA expression in CCl4 treated mice compared to controls. Conclusion We have shown that hepatocytes produce active MMPs and that the glycoprotein CD147 regulates hepatocyte MMP expression. Targeting CD147 regulates hepatocyte MMP production both in-vitro and in-vivo, with the net result being reduced fibrotic matrix turnover in-vivo. Therefore, CD147 regulation of hepatocyte MMP is a novel pathway that could be targeted by future anti-fibrogenic agents. PMID:25076423

Microarray Analysis Reveals Increased Expression of Matrix Metalloproteases and Cytokines of Interleukin-20 Subfamily in the Kidneys of Neonate Rats Underwent Unilateral Ureteral Obstruction: A Potential Role of IL-24 in the Regulation of Inflammation and Tissue Remodeling.

PubMed

Pap, Domonkos; Sziksz, Erna; Kiss, Zoltán; Rokonay, Réka; Veres-Székely, Apor; Lippai, Rita; Takács, István Márton; Kis, Éva; Fekete, Andrea; Reusz, György; Szabó, Attila J; Vannay, Adam

2017-01-01

Congenital obstructive nephropathy (CON) is the main cause of pediatric chronic kidney diseases leading to renal fibrosis. High morbidity and limited treatment opportunities of CON urge the better understanding of the underlying molecular mechanisms. To identify the differentially expressed genes, microarray analysis was performed on the kidney samples of neonatal rats underwent unilateral ureteral obstruction (UUO). Microarray results were then validated by real-time RT-PCR and bioinformatics analysis was carried out to identify the relevant genes, functional groups and pathways involved in the pathomechanism of CON. Renal expression of matrix metalloproteinase (MMP)-12 and interleukin (IL)-24 were evaluated by real-time RT-PCR, flow cytometry and immunohistochemical analysis. Effect of the main profibrotic factors on the expression of MMP-12 and IL-24 was investigated on HK-2 and HEK-293 cell lines. Finally, the effect of IL-24 treatment on the expression of pro-inflammatory cytokines and MMPs were tested in vitro. Microarray analysis revealed 880 transcripts showing >2.0-fold change following UUO, enriched mainly in immune response related processes. The most up-regulated genes were MMPs and members of IL-20 cytokine subfamily, including MMP-3, MMP-7, MMP-12, IL-19 and IL-24. We found that while TGF-β treatment inhibits the expression of MMP-12 and IL-24, H2O2 or PDGF-B treatment induce the epithelial expression of MMP-12. We demonstrated that IL-24 treatment decreases the expression of IL-6 and MMP-3 in the renal epithelial cells. This study provides an extensive view of UUO induced changes in the gene expression profile of the developing kidney and describes novel molecules, which may play significant role in the pathomechanism of CON. © 2017 The Author(s)Published by S. Karger AG, Basel.

Effect of Bisphenol A on invasion ability of human trophoblastic cell line BeWo.

PubMed

Wang, Zi-Yi; Lu, Jing; Zhang, Yuan-Zhen; Zhang, Ming; Liu, Teng; Qu, Xin-Lan

2015-01-01

Bisphenol A (BPA) is a kind of environmental endocrine disruptors (EEDs) that interfere embryo implantation. Trophoblast invasion plays a crucial role during embryo implantation. In this study, the effects of BPA on invasion ability of human trophoblastic cell line BeWo and its possible mechanism were investigated. BeWo cells were exposed to BPA and co-cultured with human endometrial cells to mimic embryo implantation in transwell model. The proliferation and invasion capability of BeWo cells were detected. The expression of E-cadherin, DNMT1, MMP-2, MMP-9, TIMP-1 and TIMP-2 were also analyzed. The results showed that the invasion capability of BeWo was reduced after daily exposure to BPA. BPA had biphasic effect on E-cadherin expression level in BeWo cells and expression level of DNMT1 was decreased when treated with BPA. Moreover, BPA treatment also changed the balance of MMPs/TIMPs in BeWo cells by down-regulating MMP-2, MMP-9 and up-regulating TIMP-1, TIMP-2 with increasing BPA concentration. Taken together, these results showed that BPA treatment could reduce the invasion ability of BeWo cells and alter the expression level of E-cadherin, DNMT1, TIMP-1, TIMP-2, MMP-2, and MMP-9. Our study would help us to understand the possible mechanism of BPA effect on invasion ability of human trophoblastic cell line BeWo.

Paracrine regulation of matrix metalloproteinase expression in the normal human endometrium.

PubMed

Osteen, K G; Keller, N R; Feltus, F A; Melner, M H

1999-01-01

Endometrial expression of matrix metalloproteinase (MMP)-3, MMP-7 and MMP-11 occurs during menstrual breakdown and subsequent estrogen-mediated growth, but not during the secretory phase. These enzymes are suppressed by progesterone treatment. Paracrine factors, including transforming growth factor-beta (TGF-beta) and retinoic acid, are also critical for MMP regulation in the endometrium. In contrast, inflammatory cytokines such as interleukin-1alpha may block or interfere with steroid-mediated MMP regulation at ectopic sites of growth. Using in vitro models, our laboratory has investigated the complex interactions between progesterone and locally produced cytokines that may affect MMP expression during the development of endometriosis. Our results indicate that targeting the regulation of MMPs may represent an appropriate therapeutic strategy for the treatment of endometriosis. Copyrightz1999S. KargerAG,Basel

CAPN 7 promotes the migration and invasion of human endometrial stromal cell by regulating matrix metalloproteinase 2 activity

PubMed Central

2013-01-01

Background Matrix metalloproteinase 2 (MMP-2) has been reported to be an important regulator of cell migration and invasion through degradation of the extracellular matrix (ECM) in many diseases, such as cancer and endometriosis. Here, we found calcium-activated neutral protease 7 (CAPN 7) expression was markedly upregulated in the eutopic endometrium and endometrial stromal cells of women diagnosed with endometriosis. Our studies were carried out to detect the effects of CAPN 7 on human endometrial stromal cell (hESC) migration and invasion. Methods Western blotting and quantitative real-time PCR were used to detect the expression of CAPN 7 in endometriosis patients and normal fertile women. Scratch-wound-healing and invasion chamber assay were used to investigate the role of CAPN 7 in hESC migration and invasion. Western blotting, quantitative real-time PCR and zymography were carried out to detect the effect of CAPN 7 on the expressions and activity of MMP-2. Results CAPN 7 was markedly up-regulated in endometriosis, thereby promoting the migration and invasion of hESC. CAPN 7 overexpression led to increased expression of MMP-2 and tissue inhibitor of metalloproteinases 2 (TIMP-2); CAPN 7 knockdown reversed these changes. CAPN 7 increased MMP-2 activity by increasing the ratio of MMP-2 to TIMP-2. We also found that OA-Hy (an MMP-2 inhibitor) decreased the effects of CAPN 7 overexpression on hESC migration and invasion by approximately 50% and 55%, respectively. Additionally, a coimmunoprecipitation assay demonstrated that CAPN 7 interacted with activator protein 2α (AP-2α): an important transcription factor of MMP-2. Conclusions CAPN 7 promotes hESC migration and invasion by increasing the activity of MMP-2 via an increased ratio of MMP-2 to TIMP-2. PMID:23855590

Enterolactone Suppresses Proliferation, Migration and Metastasis of MDA-MB-231 Breast Cancer Cells Through Inhibition of uPA Induced Plasmin Activation and MMPs-Mediated ECM Remodeling

PubMed Central

Mali, Aniket V; Joshi, Asavari A; Hegde, Mahabaleshwar V; Kadam, Shivajirao S

2017-01-01

Background: To enhance their own survival, tumor cells can manipulate their microenvironment through remodeling of the extra cellular matrix (ECM). The urokinase-type plasminogen activator (uPA) system catalyzes plasmin production which further mediates activation of matrix metalloproteinases (MMPs) and plays an important role in breast cancer invasion and metastasis through ECM remodeling. This provides a potential target for therapeutic intervention of breast cancer treatment. Enterolactone (EL) is derived from dietary flax lignans in the human body and is known to have anti-breast cancer activity. We here investigated molecular and cellular mechanisms of EL action on the uPA-plasmin-MMPs system. Methods: MTT and trypan blue dye exclusion assays, anchorage-dependent clonogenic assays and wound healing assays were carried out to study effects on cell proliferation and viability, clonogenicity and migration capacity, respectively. Real-time PCR was employed to study gene expression and gelatin zymography was used to assess MMP-2 and MMP-9 activities. All data were statistically analysed and presented as mean ± SEM values. Results: All the findings collectively demonstrated anticancer and antimetastatic potential of EL with antiproliferative, antimigratory and anticlonogenic cellular mechanisms. EL was found to exhibit multiple control of plasmin activation by down-regulating uPA expression and also up-regulating its natural inhibitor, PAI-1, at the mRNA level. Further, EL was found to down-regulate expression of MMP-2 and MMP-9 genes, and up-regulate TIMP-1 and TIMP-2; natural inhibitors of MMP-2 and MMP-9, respectively. This may be as a consequence of inhibition of plasmin activation, resulting in robust control over migration and invasion of breast cancer cells during metastasis. Conclusions: EL suppresses proliferation, migration and metastasis of MDA-MB-231 breast cancer cells by inhibiting induced ECM remodeling by the ‘uPA-plasmin-MMPs system’. PMID:28545187

TGF-β1 stimulates movement of renal proximal tubular epithelial cells in a three-dimensional cell culture via an autocrine TGF-β2 production.

PubMed

Luo, Deyi; Guan, Qiunong; Wang, Kunjie; Nguan, Christopher Y C; Du, Caigan

2017-01-01

TGF-βs are multifunctional cytokines, but their roles in human renal homeostasis are not fully understood. This study investigated the role of TGF-β1 in the movement of human renal proximal tubular epithelial cells (PTECs) in a three-dimensional (3D) model. HKC-8 cells, a human PTEC line, were grown in a 3D collagen culture system. Cell movement was observed under a microscope. The gene expression was examined using PCR Arrays or qRT-PCR, and protein levels by Western blot. Here, we showed that the tight junction structure formed between adjacent cells of a HKC-8 cell colony in 3D cultures, and TGF-β1 stimulated their movement, evidenced by the appearance of fingerlike pseudopodia in the leader cells at the edge of the colonies. The cell movement of these human PTECs was correlated with up-regulation of both MMP2 and MMP9 and down-regulation or inactivation of PLAUR and PTK2B. Analysis of TGF-β signaling targets confirmed autocrine production of TGF-β2 and its cleaving enzyme furin as well as SNAI1 by TGF-β1stimulation. Knockdown of TGF-β2 expression disrupted TGF-β1-stimulated PTEC invasiveness, which was correlated with the down-regulation of MMP2 and MMP9. In conclusion, the activation of TGF-β receptor autocrine signaling by up-regulated TGF-β2 may play a pivotal role in TGF-β1-induced human PTEC movement, which could be mediated at least by both MMP2 and MMP9. Copyright © 2016 Elsevier Inc. All rights reserved.

Preventive effects of β-thujaplicin against UVB-induced MMP-1 and MMP-3 mRNA expressions in skin fibroblasts.

PubMed

Cherng, Jong Yuh; Chen, Li Yin; Shih, Mei Fen

2012-01-01

Solar UV radiation damages human skin by affecting skin tone and resiliency and leads to premature aging (photoaging). The skin damage is caused by the activation of the AP-1 transcription factor, which increases matrix metalloproteinase (MMP) expression and collagen degradation. An increase of interleukin (IL)-6 is also correlated with the activation of MMP-1 expression. β-thujaplicin has shown both acaricidal and antimicrobial activities. Also, β-thujaplicin has been shown to be protective against apoptosis due to the oxidative effects of UV irradiation. However, the effect of β-thujaplicin on UVB-induced MMPs had not been investigated. In this study, after UVB exposure, MMP-1 and IL-6 production in human skin fibroblasts was examined in the presence of β-thujaplicin, vitamin C, and vitamin E. The expression of MMP-1, MMP-3, tissue inhibitor of metalloproteinase (TIMP-1, TIMP-3) and procollagen mRNA was also investigated. Results showed that UVB-induced MMP-1 production was suppressed by the β-thujaplicin treatment in a dose-dependent manner, but not by vitamin C and vitamin E. β-thujaplicin also prevented the up-regulation of MMP-1 and MMP-3 mRNA. Moreover, the UVB-suppressed procollagen gene expression was restored to normal by β-thujaplicin. Neither UVB nor β-thujaplicin affected the mRNA expression of TIMP-1 and TIMP-3. The IL-6 production induced by UVB was lower in β-thujaplicin treated fibroblasts than in the controls. In conclusion, this study shows the capability of β-thujaplicin in preventing MMP-1 production due to UVB irradiation via inhibition of MMP gene expression. Importantly, the UVB-suppressed procollagen gene expression can be restored to normal by β-thujaplicin. These findings indicate that β-thujaplicin is a promising and potent agent to inhibit UVB-induced MMP-1 and MMP-3 gene expression in skin fibroblasts.

Regulation of MMP-3 expression and secretion by the chemokine eotaxin-1 in human chondrocytes.

PubMed

Chao, Pin-Zhir; Hsieh, Ming-Shium; Cheng, Chao-Wen; Lin, Yung-Feng; Chen, Chien-Ho

2011-11-25

Osteoarthritis (OA) is characterized by the degradation of articular cartilage, marked by the breakdown of matrix proteins. Studies demonstrated the involvement of chemokines in this process, and some may potentially serve as diagnostic markers and therapeutic targets; however, the underlying signal transductions are not well understood. We investigated the effects of the CC chemokine eotaxin-1 (CCL11) on the matrix metalloproteinase (MMP) expression and secretion in the human chondrocyte cell line SW1353 and primary chondrocytes. Eotaxin-1 significantly induced MMP-3 mRNA expression in a dose-dependent manner. Inhibitors of extracellular signal-regulated kinase (ERK) and p38 kinase were able to repress eotaxin-1-induced MMP-3 expression. On the contrary, Rp-adenosine-3',5'-cyclic monophosphorothioate (Rp-cAMPs), a competitive cAMP antagonist for cAMP receptors, and H-89, a protein kinase A (PKA) inhibitor, markedly enhanced eotaxin-1-induced MMP-3 expression. These results suggest that MMP-3 expression is specifically mediated by the G protein-coupled eotaxin-1 receptor activities. Interestingly, little amount of MMP-3 protein was detected in the cell lysates of eotaxin-1-treated SW1353 cells, and most of MMP-3 protein was in the culture media. Furthermore we found that the eotaxin-1-dependent MMP-3 protein secretion was regulated by phospholipase C (PLC)-protein kinase C (PKC) cascade and c-Jun N-terminal kinase (JNK)/mitogen-activated protein (MAP) kinase pathways. These data indicate a specific regulation of MMP-3 secretion also by eotaxin-1 receptor activities. Eotaxin-1 not only induces MMP-3 gene expression but also promotes MMP-3 protein secretion through G protein-coupled eotaxin-1 receptor activities. Chemokines, such as eotaxin-1, could be a potential candidate in the diagnosis and treatment of arthritis.

Inhibition of matrix metalloproteinase-9 activity by doxycycline ameliorates RANK ligand-induced osteoclast differentiation in vitro and in vivo

DOE Office of Scientific and Technical Information (OSTI.GOV)

Franco, Gilson C.N.; Department of Pharmacology, FOP/UNICAMP, Piracicaba, SP; Kajiya, Mikihito

2011-06-10

Tetracycline antibiotics, including doxycycli/e (DOX), have been used to treat bone resorptive diseases, partially because of their activity to suppress osteoclastogenesis induced by receptor activator of nuclear factor kappa B ligand (RANKL). However, their precise inhibitory mechanism remains unclear. Therefore, the present study examined the effect of Dox on osteoclastogenesis signaling induced by RANKL, both in vitro and in vivo. Although Dox inhibited RANKL-induced osteoclastogenesis and down-modulated the mRNA expression of functional osteoclast markers, including tartrate-resistant acid phosphatase (TRAP) and cathepsin K, Dox neither affected RANKL-induced MAPKs phosphorylation nor NFATc1 gene expression in RAW264.7 murine monocytic cells. Gelatin zymography andmore » Western blot analyses showed that Dox down-regulated the enzyme activity of RANKL-induced MMP-9, but without affecting its protein expression. Furthermore, MMP-9 enzyme inhibitor also attenuated both RANKL-induced osteoclastogenesis and up-regulation of TRAP and cathepsin K mRNA expression, indicating that MMP-9 enzyme action is engaged in the promotion of RANKL-induced osteoclastogenesis. Finally, Dox treatment abrogated RANKL-induced osteoclastogenesis and TRAP activity in mouse calvaria along with the suppression of MMP9 enzyme activity, again without affecting the expression of MMP9 protein. These findings suggested that Dox inhibits RANKL-induced osteoclastogenesis by its inhibitory effect on MMP-9 enzyme activity independent of the MAPK-NFATc1 signaling cascade.« less

Multifunctional effect of epigallocatechin-3-gallate (EGCG) in downregulation of gelatinase-A (MMP-2) in human breast cancer cell line MCF-7.

PubMed

Sen, Triparna; Moulik, Shuvojit; Dutta, Anindita; Choudhury, Paromita Roy; Banerji, Aniruddha; Das, Shamik; Roy, Madhumita; Chatterjee, Amitava

2009-02-13

The tumor inhibiting property of green tea polyphenol epigallocatechin-3-gallate (EGCG) is well documented. Studies reveal that matrix-metalloproteinases (MMPs) play pivotal roles in tumor invasion through degradation of basement membranes and extracellular matrix (ECM). We studied the effect of EGCG on matrixmetalloproteinases-2 (MMP-2), the factors involved in activation, secretion and signaling molecules that might be involved in the regulation of MMP-2 in human breast cancer cell line, MCF-7. MCF-7 was treated with EGCG (20 muM, 24 h), the effect of EGCG on MMP-2 expression, activity and its regulatory molecules were studied by gelatin zymography, Western blot, quantitative and semi-quantitative real time RT-PCR, immunoflourescence and cell adhesion assay. EGCG treatment reduced the activity, protein expression and mRNA expression level of MMP-2. EGCG treatment reduced the expression of focal adhesion kinase (FAK), membrane type-1-matrix metalloproteinase (MT1-MMP), nuclear factor-kappa B (NF-kB), vascular endothelial growth factor (VEGF) and reduced the adhesion of MCF-7 cells to ECM, fibronectin and vitronectin. Real time RT-PCR revealed a reduced expression of integrin receptors alpha5, beta1, alphav and beta3 due to EGCG treatment. Down regulation of expression of MT1-MMP, NF-kB, VEGF and disruption of functional status of integrin receptors may indicate decreased MMP-2 activation; low levels of FAK expression might indicate disruption in FAK-induced MMP-2 secretion and decrease in activation of phosphatidyl-inositol-3-kinase (PI-3K), extracellular regulated kinase (ERK) indicates probable hindrance in MMP-2 regulation and induction. We propose EGCG as potential inhibitor of expression and activity of pro-MMP-2 by a process involving multiple regulatory molecules in MCF-7.

RANK ligand signaling modulates the matrix metalloproteinase-9 gene expression during osteoclast differentiation

DOE Office of Scientific and Technical Information (OSTI.GOV)

Sundaram, Kumaran; Nishimura, Riko; Senn, Joseph

2007-01-01

Osteoclast differentiation is tightly regulated by receptor activator of NF-{kappa}B ligand (RANKL) signaling. Matrix metalloproteinase-9 (MMP-9), a type IV collagenase is highly expressed in osteoclast cells and plays an important role in degradation of extracellular matrix; however, the molecular mechanisms that regulate MMP-9 gene expression are unknown. In this study, we demonstrate that RANKL signaling induces MMP-9 gene expression in osteoclast precursor cells. We further show that RANKL regulates MMP-9 gene expression through TRAF6 but not TRAF2. Interestingly, blockade of p38 MAPK activity by pharmacological inhibitor, SB203580 increases MMP-9 activity whereas ERK1/2 inhibitor, PD98059 decreases RANKL induced MMP-9 activity inmore » RAW264.7 cells. These data suggest that RANKL differentially regulates MMP-9 expression through p38 and ERK signaling pathways during osteoclast differentiation. Transient expression of MMP-9 gene (+ 1 to - 1174 bp relative to ATG start codon) promoter-luciferase reporter plasmids in RAW264.7 cells and RANKL stimulation showed significant increase (20-fold) of MMP-9 gene promoter activity; however, there is no significant change with respect to + 1 bp to - 446 bp promoter region and empty vector transfected cells. These results indicated that MMP-9 promoter sequence from - 446 bp to - 1174 bp relative to start codon is responsive to RANKL stimulation. Sequence analysis of the mouse MMP-9 gene promoter region further identified the presence of binding motif (- 1123 bp to - 1153 bp) for the nuclear factor of activated T cells 1 (NFATc1) transcription factor. Inhibition of NFATc1 using siRNA and VIVIT peptide inhibitor significantly decreased RANKL stimulation of MMP-9 activity. We further confirm by oligonucleotide pull-down assay that RANKL stimuli enhanced NFATc1 binding to MMP-9 gene promoter element. In addition, over-expression of constitutively active NFAT in RAW264.7 cells markedly increased (5-fold) MMP-9 gene promoter activity in the absence of RANKL. Taken together, our results suggest that RANKL signals through TRAF6 and that NFATc1 is a downstream effector of RANKL signaling to modulate MMP-9 gene expression during osteoclast differentiation.« less

SCARLESS SKIN WOUND HEALING IN FOXN1 DEFICIENT (NUDE) MICE IS ASSOCIATED WITH DISTINCTIVE MATRIX METALLOPROTEINASE EXPRESSION

PubMed Central

Gawronska-Kozak, Barbara

2011-01-01

Similar to mammalian fetuses FOXN1 deficient (nude) mice are able to restore the structure and integrity of injured skin in a scarless healing process by mechanisms independent of the genetic background. Matrix metalloproteinases (MMPs) are required for regular skin wound healing and the distinctive pattern of their expression has been implicated to promote scarless healing. In this study, we analyzed the temporal and spatial expression patterns of these molecules during the incisional skin wounds in adult nude mice. Macroscopic and histological analyses of skin wounds revealed an accelerated wound healing process, minimal granulation tissue formation and markedly diminished scarring in nude mice. Quantitative RT-PCR (Mmp-2,-3,-8,-9,-10,-12,-13,-14 and Timp-1, -2, -3), Western blots (MMP-13) and gelatin zymography (MMP-9) revealed that MMP-9 and MMP-13 showed a unique, bimodal pattern of up-regulation during the early and late phases of wound healing in nude mice. Immunohistochemically MMP-9 and MMP-13 were generally detected in epidermis during the early phase and in dermis during the late (remodeling) phase. Consistent with these in vivo observations, dermal fibroblasts cultured from nude mice expressed higher levels of type I and III collagen, MMP-9 and MMP-13 mRNA levels and higher MMP enzyme activity than wild type controls. Collectively, these finding suggest that the bimodal pattern of MMP-9 and MMP-13 expression during skin repair process in nude mice could be a major component of their ability for scarless healing. PMID:21539913

Insulin-like growth factor binding protein-3 induces angiogenesis through IGF-I- and SphK1-dependent mechanisms.

PubMed

Granata, R; Trovato, L; Lupia, E; Sala, G; Settanni, F; Camussi, G; Ghidoni, R; Ghigo, E

2007-04-01

Angiogenesis is critical for development and repair, and is a prominent feature of many pathological conditions. Based on evidence that insulin-like growth factor binding protein (IGFBP)-3 enhances cell motility and activates sphingosine kinase (SphK) in human endothelial cells, we have investigated whether IGFBP-3 plays a role in promoting angiogenesis. IGFBP-3 potently induced network formation by human endothelial cells on Matrigel. Moreover, it up-regulated proangiogenic genes, such as vascular endothelial growth factor (VEGF) and matrix metalloproteinases (MMP)-2 and -9. IGFBP-3 even induced membrane-type 1 MMP (MT1-MMP), which regulates MMP-2 activation. Decreasing SphK1 expression by small interfering RNA (siRNA), blocked IGFBP-3-induced network formation and inhibited VEGF, MT1-MMP but not IGF-I up-regulation. IGF-I activated SphK, leading to sphingosine-1-phosphate (S1P) formation. The IGF-I effect on SphK activity was blocked by specific inhibitors of IGF-IR, PI3K/Akt and ERK1/2 phosphorylation. The disruption of IGF-I signaling prevented the IGFBP-3 effect on tube formation, SphK activity and VEGF release. Blocking ERK1/2 signaling caused the loss of SphK activation and VEGF and IGF-I up-regulation. Finally, IGFBP-3 dose-dependently stimulated neovessel formation into subcutaneous implants of Matrigel in vivo. Thus, IGFBP-3 positively regulates angiogenesis through involvement of IGF-IR signaling and subsequent SphK/S1P activation.

Role of S100A12 in the pathogenesis of osteoarthritis

DOE Office of Scientific and Technical Information (OSTI.GOV)

Nakashima, Motoshige; Sakai, Tadahiro, E-mail: tadsakai@med.nagoya-u.ac.jp; Hiraiwa, Hideki

Highlights: Black-Right-Pointing-Pointer This is the first report of S100A12 expression in human OA articular cartilages. Black-Right-Pointing-Pointer Exogenous S100A12 increased the production of MMP13 and VEGF in OA chondrocytes. Black-Right-Pointing-Pointer Soluble RAGE suppressed the increased production of MMP13 and VEGF. Black-Right-Pointing-Pointer p38MAPK and NF-{kappa}B inhibitors abrogated S100A12-induced MMP13 and VEGF production. Black-Right-Pointing-Pointer S100A12 may contribute to OA progression by increasing MMP13 and VEGF production. -- Abstract: S100A12 is a member of the S100 protein family, which are intracellular calcium-binding proteins. Although there are many reports on the involvement of S100A12 in inflammatory diseases, its presence in osteoarthritic cartilage has not beenmore » reported. The purpose of this study was to investigate the expression of S100A12 in human articular cartilage in osteoarthritis (OA) and to evaluate the role of S100A12 in human OA chondrocytes. We analyzed S100A12 expression by immunohistochemical staining of cartilage samples obtained from OA and non-OA patients. In addition, chondrocytes were isolated from knee cartilage of OA patients and treated with recombinant human S100A12. Real-time RT-PCR was performed to analyze mRNA expression. Protein production of matrix metalloproteinase 13 (MMP-13) and vascular endothelial growth factor (VEGF) in the culture medium were measured by ELISA. Immunohistochemical analyses revealed that S100A12 expression was markedly increased in OA cartilages. Protein production and mRNA expression of MMP-13 and VEGF in cultured OA chondrocytes were significantly increased by treatment with exogenous S100A12. These increases in mRNA expression and protein production were suppressed by administration of soluble receptor for advanced glycation end products (RAGE). Both p38 mitogen-activated protein kinase (MAPK) and nuclear factor-{kappa}B (NF-{kappa}B) inhibitors also suppressed the increases in mRNA expression and protein production of MMP-13 and VEGF. We demonstrated marked up-regulation of S100A12 expression in human OA cartilages. Exogenous S100A12 increased the production of MMP-13 and VEGF in human OA chondrocytes. Our data indicate the possible involvement of S100A12 in the development of OA by up-regulating MMP-13 and VEGF via p38 MAPK and NF-{kappa}B pathways.« less

Increased CD147 (EMMPRIN) expression in the rat brain following traumatic brain injury.

PubMed

Wei, Ming; Li, Hong; Shang, Yanguo; Zhou, Ziwei; Zhang, Jianning

2014-10-17

The extracellular matrix metalloproteinase inducer (EMMPRIN), or CD147, has been known to play a key regulatory role in vascular permeability and leukocyte activation by inducing the expression of matrix metalloproteinases (MMPs). The effects of traumatic brain injury on the expression of EMMPRIN remain poorly understood. In this study, we investigated changes in EMMPRIN expression in a rat model of fluid percussion injury (FPI) and examined the potential association between EMMPRIN and MMP-9 expression. Adult male rats were subjected to FPI. EMMPRIN expression was markedly up-regulated in the brain tissue surrounding the injured region 6-48 h after TBI, as measured by immunoblot and immunohistochemistry. EMMPRIN expression was localized to inflammatory cells. The increase in EMMPRIN expression was temporally correlated with an increase in MMP-9 levels. These data demonstrate, for the first time, changes in CD147 and MMP-9 expression following TBI. These data also suggest that CD147 and MMP-9 may play a role in vascular injuries after TBI. Copyright © 2014 Elsevier B.V. All rights reserved.

Doxycycline down-regulates matrix metalloproteinase expression and inhibits NF-κB signaling in LPS-induced PC3 cells.

PubMed

Ogut, Deniz; Reel, Buket; Gonen Korkmaz, Ceren; Arun, Mehmet Zuhuri; Cilaker Micili, Serap; Ergur, Bekir Ugur

2016-01-01

Matrix metalloproteinase enzymes (MMPs) play important role in inflammation, malignant cell proliferation, invasion and angiogenesis by mediating extracellular matrix degradation. Doxycycline, a synthetic tetracycline, behaves as a MMP inhibitor at a subantimicrobial dose and inhibits tumor cell proliferation, invasion and angiogenesis. The aberrant activity of nuclear factor kappa B (NF-κB) causes activation of MMPs and thereby proliferation and invasion of cancer cells. The aim of this study was to investigate the effects of doxycycline on the expression of MMPs in lipopolysaccharide (LPS)-induced PC3 human prostate cancer cells and the possible role of NF-κB signaling. PC3 cells were incubated with LPS (0.5 μg/mL) for 24 h in the presence or absence of doxycycline (5 μg/mL). The effects of LPS and doxycycline on the expressions of MMP-2, MMP-8, MMP-9, MMP-10, NF-κB/p65, IκB-α, p-IκB-α, IKK-β were examined by Western blotting and immunohistochemistry in PC3 cells. Furthermore, relative proteinase activities of MMP-2 and MMP-9 were determined by gelatin zymography. LPS increased expression and activity of MMP-9 and expression of MMP-8, MMP-10, NF-κB /p65, p-IκB-α, IKK-β and doxycycline down-regulated its effects with the exception of MMP-10 expression. The expression of MMP-2 and IκB-α was affected by neither LPS nor doxycycline. Our findings indicate that doxycycline inhibits the expression of various MMPs and NF-κB signaling may play a role in the regulation of MMPs expression in LPS-induced PC3 human prostate cancer cells.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Zhan, Yu; Abi Saab, Widian F.; Modi, Nidhi

Mixed lineage kinase 3 (MLK3) is a mitogen-activated protein kinase kinase kinase (MAP3K) that activates MAPK signaling pathways and regulates cellular responses such as proliferation, migration and apoptosis. Here we report high levels of total and phospho-MLK3 in ovarian cancer cell lines in comparison to immortalized nontumorigenic ovarian epithelial cell lines. Using small interfering RNA (siRNA)-mediated gene silencing, we determined that MLK3 is required for the invasion of SKOV3 and HEY1B ovarian cancer cells. Furthermore, mlk3 silencing substantially reduced matrix metalloproteinase (MMP)-1, -2, -9 and -12 gene expression and MMP-2 and -9 activities in SKOV3 and HEY1B ovarian cancer cells.more » MMP-1, -2, -9 and-12 expression, and MLK3-induced activation of MMP-2 and MMP-9 requires both extracellular signal-regulated kinase (ERK) and c-Jun N-terminal kinase (JNK) activities. In addition, inhibition of activator protein-1 (AP-1) reduced MMP-1, MMP-9 and MMP-12 gene expression. Collectively, these findings establish MLK3 as an important regulator of MMP expression and invasion in ovarian cancer cells. -- Highlights: Black-Right-Pointing-Pointer Ovarian cancer cell lines have high levels of total and phosphorylated MLK3. Black-Right-Pointing-Pointer MLK3 is required for MMP expression and activity in ovarian cancer cells. Black-Right-Pointing-Pointer MLK3 is required for invasion of SKOV3 and HEY1B ovarian cancer cells. Black-Right-Pointing-Pointer MLK3-dependent regulation of MMP-2 and MMP-9 activities requires ERK and JNK.« less

Different strains of Propionibacterium acnes modulate differently the cutaneous innate immunity.

PubMed

Jasson, Fiona; Nagy, Istvan; Knol, Anne Chantal; Zuliani, Thomas; Khammari, Amir; Dréno, Brigitte

2013-09-01

Acne is a chronic inflammatory illness of the pilosebaceous follicle where innate immunity plays a central role. In acne, the density of Propionibacterium acnes is increased in the pilosebaceous unit. We hypothesized that the severity of acne is not only dependent on the proliferation of P. acnes but also dependent on the pro-inflammatory potential of P. acnes strains and consequently constitutes potential triggering factor for acne scarring. We investigated pro-inflammatory potential of five different strains of P. acnes and P. avidum in skin explants and the preventive effect of zinc gluconate. The expression of immune markers was studied by immunohistochemistry, RT-qPCR and ELISA. P. acnes strains modulate differently the expression of immune markers both at gene and at protein levels. P. acnes type III had the highest pro-inflammatory potential by up-regulating the expression of PAR-2, TNF-alpha, MMP-13 and TIMP-2, whereas P. avidum had the weakest by up-regulating only MMP-13 and TIMP-2. Preincubation of zinc gluconate, which is a modulator of innate immunity, down-regulates the expression of most immune markers induced by P. acnes, PAR-2, TIMP-2, up-regulates MMP-1, TIMP-1. Our results demonstrate that different P. acnes strains have different inflammatory potential targeting markers of cutaneous innate immunity, and that inflammatory potential can be down-regulated by zinc gluconate. As such, the inflammatory potential of P. acnes strains on acne skin may influence the severity of inflammatory acne lesions and scars. © 2013 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Epigenetic regulation of matrix metalloproteinase expression in ameloblastoma

PubMed Central

2012-01-01

Background An ameloblastoma is a benign odontogenic neoplasm with aggressive behaviour and high recurrence rates. The increased expression of matrix metalloproteinases (MMPs) has been reported in ameloblastomas. In the present study, we hypothesised that epigenetic alterations may regulate MMP expression in ameloblastomas. Methods We investigated the methylation status of the genes MMP-2 and MMP-9 in addition to mRNA transcription and protein expression in ameloblastomas. Methylation analysis was performed by both methylation-specific polymerase chain reaction (MSP-PCR) and restriction enzyme digestion to evaluate the methylation profile of MMP-2 and MMP-9 in 12 ameloblastoma samples and 12 healthy gingiva fragments, which were included as controls. Furthermore, we investigated the transcription levels of the genes by quantitative reverse-transcription PCR (qRT-PCR). Zymography was performed to verify protein expression in ameloblastomas. Results The ameloblastomas showed a high frequency of unmethylated MMP-2 and MMP-9, whereas the healthy gingival samples presented a sharp prevalence of methylated MMPs. Higher expression levels of MMP-9 were found in ameloblastomas compared to healthy gingiva. However, no significant differences in the MMP-2 mRNA expression between groups was found. All ameloblastomas showed positive expression of MMP-2 and MMP-9 proteins. Conclusions Our findings suggest that expression of MMP-9 is increased in ameloblastomas and is possibly modulated by unmethylation of the gene. PMID:22866959

Expression of matrix metalloproteinases (MMP-1 and -2) and their inhibitors (TIMP-1, -2 and -3) in oral lichen planus, dysplasia, squamous cell carcinoma and lymph node metastasis.

PubMed Central

Sutinen, M.; Kainulainen, T.; Hurskainen, T.; Vesterlund, E.; Alexander, J. P.; Overall, C. M.; Sorsa, T.; Salo, T.

1998-01-01

Although matrix metalloproteinases (MMPs) are among the potential key mediators of cancer invasion, their involvement in premalignant lesions and conditions is not clarified. Therefore, we studied, using in situ hybridization, immunohistochemistry and zymography the expression and distribution of MMP-1 and -2, and their tissue inhibitors (TIMPs -1, -2 and -3) in oral squamous cell carcinomas (SCC) and lymph node metastases as well as in oral lichen planus, epithelial dysplasias and normal buccal mucosa. In oral SCC and lymph node metastasis, MMP-1 mRNA was detected in fibroblastic cells of tumoral stroma. In two out of ten carcinomas studied, the peripheral cells of neoplastic islands were also positive. MMP-2 mRNA expression was noted in fibroblasts surrounding the carcinoma cells, and no signal in carcinoma cells was detected. A clear TIMP-3 mRNA expression was seen in stromal cells surrounding the neoplastic islands in all SCCs and lymph node metastases studied. TIMP-1 mRNA was detected in some stromal cells surrounding the neoplastic islands, whereas the mRNA expression for TIMP-2 was negligible. On the other hand, expression of MMPs and TIMPs was consistently low in oral epithelial dysplasias, lichen planus and normal mucosa. In certain epithelial dysplasias and lichen planus, MMP-1 and -2 mRNA expressions were detected in few fibroblasts under the basement membrane zone, but normal mucosa was completely negative. In SCC and lymph node metastasis, a detectable immunostaining for MMP-1 in stromal cells and in some carcinoma cells was observed. MMP-2 immunoreactivity was detected in the peripheral cell layer in neoplastic islands and in some fibroblast-like cells of tumoral stroma. Immunostaining for TIMP-3 was detected in stromal cells surrounding the neoplastic islands. A weak positive staining for TIMP-1 was located in tumoral stroma, whereas the immunostaining for TIMP-2 was negative. Using zymography, elevated levels of MMP-2 and MMP-9 were observed in carcinoma samples in comparison with lichen planus or normal oral mucosa. Our results indicate that the studied MMPs and TIMPs are clearly up-regulated during invasion in oral SCC. However, there was also a clear, although weak, up-regulation of the expression of the MMPs but not TIMPs in some of the lichen planus and dysplastic lesions. Images Figure 1 Figure 2 Figure 3 Figure 4 PMID:9649139

Sirtuin 1 regulates matrix metalloproteinase-13 expression induced by Porphyromonas endodontalis lipopolysaccharide via targeting nuclear factor–κB in osteoblasts

PubMed Central

Qu, Liu; Yu, Yaqiong; Qiu, Lihong; Yang, Di; Yan, Lu; Guo, Jiajie; Jahan, Rabita

2017-01-01

ABSTRACT Porphyromonas endodontalis lipopolysaccharide (P.e LPS) is an important initiating factor for periapical inflammation and bone destruction. Matrix metalloproteinase-13 (MMP-13) has been shown to participate in the formation and diffusion of periapical bone lesion in chronic apical periodontitis. Sirtuin 1 (SIRT1) is a key regulator of inflammation in mammalian cells which suppresses the release of inflammatory mediators. This study aimed to explore the role of SIRT1 in regulating MMP-13 expression induced by P.e LPS in osteoblasts. P.e LPS stimulated MMP-13 expression in MC3T3-E1 cells. Knockdown of SIRT1 reinforced the increase of MMP-13mRNA expression induced by P.e LPS. SIRT1 activator resveratrol significantly reduced the expression of MMP-13 and SIRT1 inhibitor EX-527 enhanced the expression of MMP-13. Moreover, SIRT1 activation with resveratrol inhibited acetylation of NF-κB p65 and NF-κB transcriptional activity, which were enhanced by P.e LPS. In addition, NF-κB p65 was involved in P.e LPS-induced MMP-13 expression via directly binding to the MMP-13 promoter. However, SIRT1 activation significantly interfered with this binding. These findings strongly suggest that P.e LPS induces MMP-13 expression in osteoblasts, and SIRT1 suppresses this expression of MMP-13 through targeting NF-κB p65. This provides new insights into understanding the actions of SIRT1 on anti-inflammatory and anti-bone resorption activity. PMID:28473882

Sirtuin 1 regulates matrix metalloproteinase-13 expression induced by Porphyromonas endodontalis lipopolysaccharide via targeting nuclear factor-κB in osteoblasts.

PubMed

Qu, Liu; Yu, Yaqiong; Qiu, Lihong; Yang, Di; Yan, Lu; Guo, Jiajie; Jahan, Rabita

2017-01-01

Porphyromonas endodontalis lipopolysaccharide (P.e LPS) is an important initiating factor for periapical inflammation and bone destruction. Matrix metalloproteinase-13 (MMP-13) has been shown to participate in the formation and diffusion of periapical bone lesion in chronic apical periodontitis. Sirtuin 1 (SIRT1) is a key regulator of inflammation in mammalian cells which suppresses the release of inflammatory mediators. This study aimed to explore the role of SIRT1 in regulating MMP-13 expression induced by P.e LPS in osteoblasts. P.e LPS stimulated MMP-13 expression in MC3T3-E1 cells. Knockdown of SIRT1 reinforced the increase of MMP-13mRNA expression induced by P.e LPS. SIRT1 activator resveratrol significantly reduced the expression of MMP-13 and SIRT1 inhibitor EX-527 enhanced the expression of MMP-13. Moreover, SIRT1 activation with resveratrol inhibited acetylation of NF-κB p65 and NF-κB transcriptional activity, which were enhanced by P.e LPS. In addition, NF-κB p65 was involved in P.e LPS-induced MMP-13 expression via directly binding to the MMP-13 promoter. However, SIRT1 activation significantly interfered with this binding. These findings strongly suggest that P.e LPS induces MMP-13 expression in osteoblasts, and SIRT1 suppresses this expression of MMP-13 through targeting NF-κB p65. This provides new insights into understanding the actions of SIRT1 on anti-inflammatory and anti-bone resorption activity.

Rg1 inhibits high glucose-induced mesenchymal activation and fibrosis via regulating miR-2113/RP11-982M15.8/Zeb1 pathway.

PubMed

Xue, Li-Ping; Fu, Xiao-Lin; Hu, Min; Zhang, Li-Wei; Li, Ya-Di; Peng, Ya-Li; Ding, Peng

2018-07-02

Recent study has showed that Ginsenoside Rg1, the mian active compound of Panax ginseng, could ameliorate oxidative stress and myocardial apoptosis in diabetes mellitus. However, the roles and mechanisms of Rg1 in proliferative diabetic retinopathy (PDR) are still unclear. In the present study, we aimed to investigate the effects of Rg1 on mesenchymal activation of high-glucose (HG) cultured müller cells. High glucose conditions up-regulate MMP-2, MMP-9 and down-regulate TIMP-2, and promote mesenchymal activation in Müller cells. And Rg1 inhibits the HG-induced mesenchymal activation and HG-increased MMP-2 and MMP-9 and HG-decreased TIMP-2 in Müller cells. HG up-regulates Zeb1 and lncRNA RP11-982M15.8, and down-regulates miR-2113, and Rg1 inhibits these effects of HG. Both inhibition of miR-2113 and over-expression of RP11-982M15.8 significantly restored the HG induced mesenchymal activasion. Taken together, our findings suggested that Rg1 inhibited HG-induced mesenchymal activation and fibrosis via regulating miR-2113/RP11-982M15.8/Zeb1 pathway. Copyright © 2018. Published by Elsevier Inc.

Interleukin-6 increases matrix metalloproteinase-14 (MMP-14) levels via down-regulation of p53 to drive cancer progression.

PubMed

Cathcart, Jillian M; Banach, Anna; Liu, Alice; Chen, Jun; Goligorsky, Michael; Cao, Jian

2016-09-20

Matrix metalloproteinases (MMPs) play critical roles in cancer invasion and metastasis by digesting basement membrane and extracellular matrix (ECM). Much attention has focused on the enzymatic activities of MMPs; however, the regulatory mechanism of MMP expression remains elusive. By employing bioinformatics analysis, we identified a potential p53 response element within the MMP-14 promoter. Experimentally, we found that p53 can repress MMP-14 promoter activity, whereas deletion of this p53 response element abrogated this effect. Furthermore, we found that p53 expression decreases MMP-14 mRNA and protein levels and attenuates MMP-14-mediated cellular functions. Additional promoter analysis and chromatin immunoprecipitation studies identified a mechanism of regulation of MMP-14 expression by which p53 and transcription factor Sp1 competitively bind to the promoter. As the correlation between inflammation and cancer aggressiveness is well described, we next sought to evaluate if inflammatory cytokines could differentially affect p53 and MMP-14 levels. We demonstrate that interleukin-6 (IL-6) down-regulates p53 protein levels and thus results in a concomitant increase in MMP-14 expression, leading to enhanced cancer cell invasion and metastasis. Our data collectively indicate a novel mechanism of regulation of MMP-14 by a cascade of IL-6 and p53, demonstrating that the tumor microenvironment directly stimulates molecular changes in cancer cells to drive an invasive phenotype.

NFATc3 promotes Ca(2+) -dependent MMP3 expression in astroglial cells.

PubMed

Neria, Fernando; del Carmen Serrano-Perez, María; Velasco, Patricia; Urso, Katia; Tranque, Pedro; Cano, Eva

2013-07-01

Increase in intracellular calcium ([Ca(2+) ]i ) is a key mediator of astrocyte signaling, important for activation of the calcineurin (CN)/nuclear factor of activated T cells (NFAT) pathway, a central mediator of inflammatory events. We analyzed the expression of matrix metalloproteinase 3 (Mmp3) in response to increases in [Ca(2+) ]i and the role of the CN/NFAT pathway in this regulation. Astrocyte Mmp3 expression was induced by overexpression of a constitutively active form of NFATc3, whereas other MMPs and tissue inhibitor of metalloproteinases (TIMP) were unaffected. Mmp3 mRNA and protein expression was also induced by calcium ionophore (Io) and 2'(3')-O-(4-benzoylbenzoyl) adenosine 5'-triphosphate (Bz-ATP) and Mmp3 upregulation was prevented by the CN inhibitor cyclosporin A (CsA). Ca(2+) -dependent astrocyte Mmp3 expression was also inhibited by actinomycin D, and a Mmp3 promoter luciferase reporter was efficiently activated by increased [Ca(2+) ]i , indicating regulation at the transcriptional level. Furthermore, Ca(2+) /CN/NFAT dependent Mmp3 expression was confirmed in pure astrocyte cultures derived from neural stem cells (Ast-NSC), demonstrating that the induced Mmp3 expression occurs in astrocytes, and not microglial cells. In an in vivo stab-wound model of brain injury, MMP3 expression was detected in NFATc3-positive scar-forming astrocytes. Because [Ca(2+) ]i increase is an early event in most brain injuries, these data support an important role for Ca(2+) /CN/NFAT-induced astrocyte MMP3 expression in the early neuroinflammatory response. Understanding the molecular pathways involved in this regulation could provide novel therapeutic targets and approaches to promoting recovery of the injured brain. Copyright © 2013 Wiley Periodicals, Inc.

Azilsartan increases levels of IL-10, down-regulates MMP-2, MMP-9, RANKL/RANK, Cathepsin K and up-regulates OPG in an experimental periodontitis model.

PubMed

Araújo, Aurigena Antunes de; Varela, Hugo; Brito, Gerly Anne de Castro; Medeiros, Caroline Addison Carvalho Xavier de; Araújo, Lorena de Souza; do Nascimento, José Heriberto Oliveira; de Araújo Júnior, Raimundo Fernandes

2014-01-01

The aim of this study was to evaluate the effects of azilsartan (AZT) on bone loss, inflammation, and the expression of matrix metallo proteinases (MMPs), receptor activator of nuclear factor κB ligand (RANKL), receptor activator of nuclear factor κB (RANK), osteoprotegerin (OPG), cyclooxygenase-2 (COX-2), and cathepsin K in periodontal tissue in a rat model of ligature-induced periodontitis. Male Wistar albino rats were randomly divided into 5 groups of 10 rats each: (1) nonligated, water; (2) ligated, water; (3) ligated, 1 mg/kg AZT; (4) ligated, 5 mg/kg AZT; and (5) ligated, 10 mg/kg AZT. All groups were treated with saline or AZT for 10 days. Periodontal tissues were analyzed by histopathology and immunohistochemical detection of MMP-2, MMP-9, COX-2, RANKL, RANK, OPG, and cathepsin K. Levels of IL-1β, IL-10, TNF-α, myeloperoxidase (MPO), and glutathione (GSH) were determined by ELISA. Treatment with 5 mg/kg AZT resulted in reduced MPO (p<0.05) and IL-1β (p<0.05), increased levels of IL-10 (p<0.05), and reduced expression of MMP-2, MMP-9, COX-2, RANK, RANKL, cathepsin K, and increased expression of OPG. These findings reveal that AZT increases anti-inflammatory cytokines and GSH and decreases bone loss in ligature-induced periodontitis in rats.

Matrix metalloproteinase-9 plays a role in protecting zebrafish from lethal infection with Listeria monocytogenes by enhancing macrophage migration.

PubMed

Shan, Ying; Zhang, Yikai; Zhuo, Xunhui; Li, Xiaoliang; Peng, Jinrong; Fang, Weihuan

2016-07-01

Zebrafish could serve as an alternative animal model for pathogenic bacteria in multiple infectious routes. Our previous study showed that immersion infection in zebrafish with Listeria monocytogenes did not cause lethality but induced transient expression of several immune response genes. We used an Affymetrix gene chip to examine the expression profiles of genes of zebrafish immersion-infected with L. monocytogenes. A total of 239 genes were up-regulated and 56 genes down-regulated compared with uninfected fish. Highest expression (>20-fold) was seen with the mmp-9 gene encoding the matrix metalloproteinase-9 (Mmp-9) known to degrade the extracellular matrix proteins. By morpholino knockdown of mmp-9, we found that the morphants showed rapid death with much higher bacterial load after intravenous or intraventricular (brain ventricle) infection with L. monocytogenes. Macrophages in mmp-9-knockdown morphants had significant defect in migrating to the brain cavity upon intraventricular infection. Decreased migration of murine macrophages with knockdown of mmp-9 and cd44 was also seen in transwell inserts with 8-μm pore polycarbonate membrane, as compared with the scrambled RNA. These findings suggest that Mmp-9 is a protective molecule against infection by L. monocytogenes by engaging in migration of zebrafish macrophages to the site of infection via a non-proteolytic role. Further work is required on the molecular mechanisms governing Mmp-9-driven macrophage migration in zebrafish. Copyright © 2016 Elsevier Ltd. All rights reserved.

Regulation of MMP-3 expression and secretion by the chemokine eotaxin-1 in human chondrocytes

PubMed Central

2011-01-01

Background Osteoarthritis (OA) is characterized by the degradation of articular cartilage, marked by the breakdown of matrix proteins. Studies demonstrated the involvement of chemokines in this process, and some may potentially serve as diagnostic markers and therapeutic targets; however, the underlying signal transductions are not well understood. Methods We investigated the effects of the CC chemokine eotaxin-1 (CCL11) on the matrix metalloproteinase (MMP) expression and secretion in the human chondrocyte cell line SW1353 and primary chondrocytes. Results Eotaxin-1 significantly induced MMP-3 mRNA expression in a dose-dependent manner. Inhibitors of extracellular signal-regulated kinase (ERK) and p38 kinase were able to repress eotaxin-1-induced MMP-3 expression. On the contrary, Rp-adenosine-3',5'-cyclic monophosphorothioate (Rp-cAMPs), a competitive cAMP antagonist for cAMP receptors, and H-89, a protein kinase A (PKA) inhibitor, markedly enhanced eotaxin-1-induced MMP-3 expression. These results suggest that MMP-3 expression is specifically mediated by the G protein-coupled eotaxin-1 receptor activities. Interestingly, little amount of MMP-3 protein was detected in the cell lysates of eotaxin-1-treated SW1353 cells, and most of MMP-3 protein was in the culture media. Furthermore we found that the eotaxin-1-dependent MMP-3 protein secretion was regulated by phospholipase C (PLC)-protein kinase C (PKC) cascade and c-Jun N-terminal kinase (JNK)/mitogen-activated protein (MAP) kinase pathways. These data indicate a specific regulation of MMP-3 secretion also by eotaxin-1 receptor activities. Conclusions Eotaxin-1 not only induces MMP-3 gene expression but also promotes MMP-3 protein secretion through G protein-coupled eotaxin-1 receptor activities. Chemokines, such as eotaxin-1, could be a potential candidate in the diagnosis and treatment of arthritis. PMID:22114952

Proteinase-activated receptor (PAR)-2 activation impacts bone resorptive properties of human osteoarthritic subchondral bone osteoblasts.

PubMed

Amiable, Nathalie; Tat, Steeve Kwan; Lajeunesse, Daniel; Duval, Nicolas; Pelletier, Jean-Pierre; Martel-Pelletier, Johanne; Boileau, Christelle

2009-06-01

In osteoarthritis (OA), the subchondral bone undergoes a remodelling process involving several factors synthesized by osteoblasts. In this study, we investigated the expression, production, modulation, and role of PAR-2 in human OA subchondral bone osteoblasts. PAR-2 expression and production were determined by real-time PCR and flow cytometry, respectively. PAR-2 modulation was investigated in OA subchondral bone osteoblasts treated with IL-1 beta (100 pg/ml), TNF-alpha (5 ng/ml), TGF-beta1 (10 ng/ml), PGE(2) (500 nM), IL-6 (10 ng/ml) and IL-17 (10 ng/ml). Membranous RANKL protein was assessed by flow cytometry, and OPG, MMP-1, MMP-9, MMP-13, IL-6 and intracellular signalling pathways by specific ELISAs. Bone resorptive activity was measured by using a co-culture model of human PBMC and OA subchondral bone osteoblasts. PAR-2 expression and production (p<0.05) were markedly increased when human OA subchondral bone osteoblasts were compared to normal. On OA osteoblasts, PAR-2 production was significantly increased by IL-1 beta, TNF-alpha and PGE(2). Activation of PAR-2 with a specific agonist, SLIGKV-NH(2), induced a significant up-regulation of MMP-1, MMP-9, IL-6, and membranous RANKL, but had no effect on MMP-13 or OPG production. Interestingly, bone resorptive activity was also significantly enhanced following PAR-2 activation. The PAR-2 effect was mediated by activation of the MAP kinases Erk1/2 and JNK. This study is the first to demonstrate that PAR-2 activation plays a role in OA subchondral bone resorption via an up-regulation of major bone remodelling factors. These results shed new light on the potential of PAR-2 as a therapeutic target in OA.

IL-1β-induced matrix metalloproteinase-13 is activated by a disintegrin and metalloprotease-28-regulated proliferation of human osteoblast-like cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ozeki, Nobuaki; Kawai, Rie; Yamaguchi, Hideyuki

2014-04-15

We reported previously that matrix metalloproteinase (MMP)-13 accelerates bone remodeling in oral periradicular lesions, and indicated a potentially unique role for MMP-13 in wound healing and regeneration of alveolar bone. The ADAM (a disintegrin and metalloprotease) family is a set of multifunctional cell surface and secreted glycoproteins, of which ADAM-28 has been localized in bone and bone-like tissues. In this study, we show that interleukin (IL)-1β induces the expression of MMP-13 and ADAM-28 in homogeneous α7 integrin-positive human skeletal muscle stem cell (α7{sup +}hSMSC)-derived osteoblast-like (α7{sup +}hSMSC-OB) cells, and promotes proliferation while inhibiting apoptosis in these cells. At higher concentrations,more » however, IL-1β failed to induce the expression of these genes and caused an increase in apoptosis. We further employed ADAM-28 small interfering RNA (siRNA) to investigate whether IL-1β-induced MMP-13 expression is linked to this IL-1β-mediated changes in cell proliferation and apoptosis. Silencing ADAM-28 expression potently suppressed IL-1β-induced MMP-13 expression and activity, decreased cell proliferation and increased apoptosis in α7{sup +}hSMSC-OB cells. In contrast, MMP-13 siRNA had no effect on ADAM-28 expression, suggesting ADAM-28 regulates MMP-13. Exogenous MMP-13 induced α7{sup +}hSMSC-OB cell proliferation and could rescue ADAM-28 siRNA-induced apoptosis, and we found that proMMP-13 is partially cleaved into its active form by ADAM-28 in vitro. Overall, our results suggest that IL-1β-induced MMP-13 expression and changes in cell proliferation and apoptosis in α7{sup +}hSMSC-OB cells are regulated by ADAM-28. - Highlights: • IL-1β induces the MMP-13 and ADAM-28 expression in human osteoblast-like cells. • IL-1β-induced MMP-13 expression increases proliferation and decreased apoptosis. • MMP-13 expression induced by IL-1β is regulated by ADAM-28. • proMMP-13 appears to be cleaved into its active form via ADAM-28.« less

Rosmarinic acid induces rabbit articular chondrocyte differentiation by decreases matrix metalloproteinase-13 and inflammation by upregulating cyclooxygenase-2 expression.

PubMed

Eo, Seong-Hui; Kim, Song Ja

2017-09-18

Matrix metalloproteinases (MMPs) are known to play an important role in the degradation of the extracellular matrix and the pathological progression of osteoarthritis (OA). The natural polyphenolic compound rosmarinic acid (Ros. A) has been shown to suppress the inhibitory activity of matrix metalloproteinases (MMPs). However, the effects of Ros. A on OA have not been investigated. In the current study, primary articular chondrocytes were cultured from rabbit articular cartilage and treated with Ros. A. Phenotypic characterization was performed by western blotting to assess specific markers, prostaglandin E 2 (PGE 2 ) assays, and alcian blue staining to measure sulfated-proteoglycan production. We report that in rabbit articular chondrocytes, Ros. A increased type II collagen, sulfated-proteoglycan, cyclooxygenase-2 (COX-2), and PGE 2 production in a dose- and time-dependent manner. Furthermore, Ros. A suppressed the expression of MMP-13. In addition, treatment with Ros A activated extracellular signal-regulated kinase (ERK)-1/2 and p38 kinase signaling pathways. Inhibition of MMP-13 enhanced Ros. A-induced type II collagen expression and sulfated-proteoglycan synthesis but COX-2 and PGE 2 production were unchanged. Ros. A-mediated up-regulation of ERK phosphorylation was abolished by the MEK inhibitor, PD98059, which prevented induction of the associated inflammatory response. Inhibition of p38 kinase with SB203580 enhanced the increase in type II collagen expression via Ros. A-mediated down-regulation of MMP-13. Results suggest that ERK-1/2 regulates Ros. A-induced inflammation and that p38 regulates differentiation by inhibiting MMP-13 in rabbit articular chondrocytes.

Epigenetic regulation of metalloproteinases and their inhibitors in rotator cuff tears

PubMed Central

Caires dos Santos, Leonardo; Martins de Oliveira, Adrielle; Santoro Belangero, Paulo; Antônio Figueiredo, Eduardo; Cohen, Carina; de Seixas Alves, Felipe; Hiromi Yanaguizawa, Wânia; Vicente Andreoli, Carlos; de Castro Pochini, Alberto; Ejnisman, Benno; Cardoso Smith, Marília; de Seixas Alves, Maria Teresa; Cohen, Moises

2017-01-01

Rotator cuff tear is a common orthopedic condition. Metalloproteinases (MMP) and their inhibitors (TIMP) seem to play a role in the development of joint injuries and in the failure of tissue healing. However, the mechanisms of regulation of gene expression in tendons are still unknown. Epigenetic mechanisms, such as DNA methylation and microRNAs regulation, are involved in the dynamic control of gene expression. Here, the mRNA expression and DNA methylation status of MMPs (MMP1, MMP2, MMP3, MMP9, MMP13, and MMP14) and TIMPs (TIMP1-3) and the expression of miR-29 family members in ruptured supraspinatus tendons were compared with non-injured tendons of individuals without this lesion. Additionally, the gene expression and methylation status at the edge of the ruptured tendon were compared with macroscopically non-injured rotator cuff tendon samples from the anterior and posterior regions of patients with tendon tears. Moreover, the possible associations between the molecular alterations and the clinical and histologic characteristics were investigated. Dysregulated expression and DNA methylation of MMP and TIMP genes were found across the rotator cuff tendon samples of patients with supraspinatus tears. These alterations were influenced at least in part by age at surgery, sex, smoking habit, tear size, and duration of symptoms. Alterations in the studied MMP and TIMP genes may contribute to the presence of microcysts, fissures, necrosis, and neovascularization in tendons and may thus be involved in the tendon healing process. In conclusion, MMPs and their inhibitors are regulated by epigenetic modifications and may play a role in rotator cuff tears. PMID:28902861

EMMPRIN/CD147 up-regulates urokinase-type plasminogen activator: implications in oral tumor progression

PubMed Central

2012-01-01

Backgrounds An elevated level of EMMPRIN in cancer tissues have been correlated with tumor invasion in numerous cancers including oral cavity and larynx. Although EMMPRIN's effect has been generally attributed to its MMP inducing activity, we have previously demonstrated in breast cancer model that EMMPRIN can also enhance invasion by upregulating uPA. In this study, the role of EMMPRIN in regulating uPA and invasion was investigated in oral squamous cell carcinoma (OSCC) progression. Methods Precancerous and invasive oral tumoral tissues were used as well as the corresponding cell lines, DOK and SCC-9 respectively. The paracrine regulation of uPA by EMMPRIN was investigated by treating culture cells with EMMPRIN-enriched membrane vesicles. UPA expression was analyzed by qPCR and immunostaining and the consequence on the invasion capacity was studied using modified Boyden chamber assay, in the presence or absence of EMMPRIN blocking antibody, the uPA inhibitor amiloride or the MMP inhibitor marimastat. Results OSCC tumors were shown to express more EMMPRIN and uPA compared to dysplastic lesions. The corresponding cell models, SCC-9 and DOK cells, displayed similar expression pattern. In both cell types EMMPRIN upregulated the expression of uPA as well as that of MMP-2 and MMP-9. EMMPRIN treatment led to a significant increase in cell invasion both in the invasive SCC-9 and in the less invasive dysplastic DOK cells, in an MMP and uPA dependent manner. Conclusions Our results suggest that the upregulation of uPA contributes to EMMPRIN's effect in promoting oral tumor invasion. PMID:22443116

EMMPRIN/CD147 up-regulates urokinase-type plasminogen activator: implications in oral tumor progression.

PubMed

Lescaille, Géraldine; Menashi, Suzanne; Cavelier-Balloy, Bénédicte; Khayati, Farah; Quemener, Cathy; Podgorniak, Marie Pierre; Naïmi, Benyoussef; Calvo, Fabien; Lebbe, Céleste; Mourah, Samia

2012-03-23

An elevated level of EMMPRIN in cancer tissues have been correlated with tumor invasion in numerous cancers including oral cavity and larynx. Although EMMPRIN's effect has been generally attributed to its MMP inducing activity, we have previously demonstrated in breast cancer model that EMMPRIN can also enhance invasion by upregulating uPA. In this study, the role of EMMPRIN in regulating uPA and invasion was investigated in oral squamous cell carcinoma (OSCC) progression. Precancerous and invasive oral tumoral tissues were used as well as the corresponding cell lines, DOK and SCC-9 respectively. The paracrine regulation of uPA by EMMPRIN was investigated by treating culture cells with EMMPRIN-enriched membrane vesicles. UPA expression was analyzed by qPCR and immunostaining and the consequence on the invasion capacity was studied using modified Boyden chamber assay, in the presence or absence of EMMPRIN blocking antibody, the uPA inhibitor amiloride or the MMP inhibitor marimastat. OSCC tumors were shown to express more EMMPRIN and uPA compared to dysplastic lesions. The corresponding cell models, SCC-9 and DOK cells, displayed similar expression pattern. In both cell types EMMPRIN upregulated the expression of uPA as well as that of MMP-2 and MMP-9. EMMPRIN treatment led to a significant increase in cell invasion both in the invasive SCC-9 and in the less invasive dysplastic DOK cells, in an MMP and uPA dependent manner. Our results suggest that the upregulation of uPA contributes to EMMPRIN's effect in promoting oral tumor invasion.

Activation of G protein coupled estrogen receptor (GPER) promotes the migration of renal cell carcinoma via the PI3K/AKT/MMP-9 signals.

PubMed

Guan, Bao-Zhang; Yan, Rui-Ling; Huang, Jian-Wei; Li, Fo-Lan; Zhong, Ying-Xue; Chen, Yu; Liu, Fan-Na; Hu, Bo; Huang, Si-Bo; Yin, Liang-Hong

2018-03-04

Renal cell carcinoma (RCC) is the third most frequent malignancy within urological oncology. However, the mechanisms responsible for RCC metastasis are still needed further illustration. Our present study revealed that a seven-transmembrane receptor G-protein coupled estrogen receptor (GPER) was highly detected in various RCC cell lines such as ACHN, OS-RC-2 and SW839. The activation of GPER by its specific agonist G-1 significantly promoted the in vitro migration and invasion of ACHN and OS-RC-2 cells. G-1 also up regulated the expression of matrix metalloproteinase-2 (MMP-2) and MMP-9. The inhibitor of MMP-9 (Cat-444278), but not MMP-2 (Sc-204092), abolished G-1 induced cell migration, which suggested that MMP-9 is the key molecule mediating G-1 induced RCC progression. Further, G-1 treatment resulted in phosphorylation of AKT and ERK in RCC cells. PI3K/AKT inhibitor (LY294002), while not ERK inhibitor (PD98059), significantly abolished G-1 induced up regulation of MMP-9 in both AHCN and OS-RC-2 cells. Generally, our data revealed that activation of GPER by its specific agonist G-1 promoted the metastasis of RCC cells through PI3K/AKT/MMP-9 signals, which might be a promising new target for drug discovery of RCC patients.

Collagenolytic protease expression in cranial cruciate ligament and stifle synovial fluid in dogs with cranial cruciate ligament rupture.

PubMed

Muir, Peter; Danova, Nichole A; Argyle, David J; Manley, Paul A; Hao, Zhengling

2005-01-01

To determine expression of collagenolytic genes and collagen degradation in stifle tissues of dogs with ruptured cranial cruciate ligament (CCL). Six dogs with CCL rupture and 11 dogs with intact CCL. Gene expression in CCL tissue and synovial fluid cells was studied using reverse transcriptase-polymerase chain reaction (RT-PCR). Collagen degradation was studied using CCL explant cultures and a synovial fluid bioassay. Expression of matrix metalloproteases (MMP) was not found in young Beagles with intact CCL; however, increased expression of MMP-3 was found in CCL tissue from older hounds with intact CCL, when compared with young Beagles. In dogs with ruptured CCL, expression of MMP-2 and -9 was increased in stifle tissues, when compared with dogs with intact CCL. Similar to MMP-9, expression of tartrate-resistant acid phosphatase (TRAP) and cathepsin S was only found in stifle tissues from dogs with ruptured CCL; in contrast, expression of cathepsin K was found in all ruptured and intact CCL. Collagen degradation was increased in ruptured CCL, when compared with intact CCL. Rupture of the CCL is associated with up-regulation of expression of MMP-2 and -9 (gelatinase A and B), TRAP, and cathepsin S, and increased degradation of collagen. These findings suggest that MMP-2, -9, cathepsin S, and TRAP may be important mediators of progressive joint destruction in dogs with CCL rupture. These genes are markers for macrophages and dendritic cells. MMP and cathepsin S pathways may offer novel targets for anti-inflammatory medical therapy aimed at ameliorating joint degradation associated with inflammatory arthritis.

Interleukin-6 increases matrix metalloproteinase-14 (MMP-14) levels via down-regulation of p53 to drive cancer progression

PubMed Central

Cathcart, Jillian M.; Banach, Anna; Liu, Alice; Chen, Jun; Goligorsky, Michael; Cao, Jian

2016-01-01

Matrix metalloproteinases (MMPs) play critical roles in cancer invasion and metastasis by digesting basement membrane and extracellular matrix (ECM). Much attention has focused on the enzymatic activities of MMPs; however, the regulatory mechanism of MMP expression remains elusive. By employing bioinformatics analysis, we identified a potential p53 response element within the MMP-14 promoter. Experimentally, we found that p53 can repress MMP-14 promoter activity, whereas deletion of this p53 response element abrogated this effect. Furthermore, we found that p53 expression decreases MMP-14 mRNA and protein levels and attenuates MMP-14-mediated cellular functions. Additional promoter analysis and chromatin immunoprecipitation studies identified a mechanism of regulation of MMP-14 expression by which p53 and transcription factor Sp1 competitively bind to the promoter. As the correlation between inflammation and cancer aggressiveness is well described, we next sought to evaluate if inflammatory cytokines could differentially affect p53 and MMP-14 levels. We demonstrate that interleukin-6 (IL-6) down-regulates p53 protein levels and thus results in a concomitant increase in MMP-14 expression, leading to enhanced cancer cell invasion and metastasis. Our data collectively indicate a novel mechanism of regulation of MMP-14 by a cascade of IL-6 and p53, demonstrating that the tumor microenvironment directly stimulates molecular changes in cancer cells to drive an invasive phenotype. PMID:27531896

MDA-MB-231 and 8701BC breast cancer lines promote the migration and invasiveness of ECV304 cells on 2D and 3D type-I collagen matrix.

PubMed

Saladino, Silvia; Salamone, Monica; Ghersi, Giulio

2017-09-01

Tumor angiogenesis is a multiphasic process, having the extracellular matrix remodeling as critical step. Different classes of proteolytic enzymes in matrix digestion/remodeling are involved. The role of lytic enzymes and their activation mode have not been completely elucidated. Herein, the crosstalk between endothelia and tumor cells, by realization of bi- and three-dimensional endothelial and breast cancer cells co-cultures, were studied in vitro. Particularly, the effects of two tumor conditioned media (TCM) were assessed about endothelial proliferation, migration, and invasiveness. An increase in expression of pro-MMP9 was detected when endothelial cells were cultured in the presence of both TCM; such as an up-regulation of MMP1 and MMP14 and a down-regulation of MMP7. Moreover the increased MMP2 gene expression from one of them and the stimulation MMP3 synthesis from the other one were observed; an increases of β3-integrin, VEGFA, and DPP4 molecules were detected when endothelia cells are cultured with both TCM. The selection/characterization of elements present in conditioned media from breast cancer cells differently affect endothelial cells, make them potential effectors useful in breast cancer treatment. © 2017 International Federation for Cell Biology.

Matrix Metalloproteinase-9 Expression Is Enhanced in Renal Parietal Epithelial Cells of Zucker Diabetic Fatty Rats and Is Induced by Albumin in In Vitro Primary Parietal Cell Culture

PubMed Central

Zhang, Yuanyuan; George, Jasmine; Li, Yun; Olufade, Rebecca; Zhao, Xueying

2015-01-01

As a subfamily of matrix metalloproteinases (MMPs), gelatinases including MMP-2 and MMP-9 play an important role in remodeling and homeostasis of the extracellular matrix. However, conflicting results have been reported regarding their expression level and activity in the diabetic kidney. This study investigated whether and how MMP-9 expression and activity were changed in glomerular epithelial cells upon albumin overload. In situ zymography, immunostaining and Western blot for renal MMP gelatinolytic activity and MMP-9 protein expression were performed in Zucker lean and Zucker diabetic rats. Confocal microscopy revealed a focal increase in gelatinase activity and MMP-9 protein in the glomeruli of diabetic rats. Increased glomerular MMP-9 staining was mainly observed in hyperplastic parietal epithelial cells (PECs) expressing claudin-1 in the diabetic kidneys. Interestingly, increased parietal MMP-9 was often accompanied by decreased staining for podocyte markers (nephrin and podocalyxin) in the sclerotic area of affected glomeruli in diabetic rats. Additionally, urinary excretion of podocyte marker proteins was significantly increased in association with the levels of MMP-9 and albumin in the urine of diabetic animals. To evaluate the direct effect of albumin on expression and activity of MMP-9, primary cultured rat glomerular PECs were incubated with rat serum albumin (0.25 - 1 mg/ml) for 24 - 48 hrs. MMP-9 mRNA levels were significantly increased following albumin treatment. Meanwhile, albumin administration resulted in a dose-dependent increase in MMP-9 protein and activity in culture supernatants of PECs. Moreover, albumin activated p44/42 mitogen-activated protein kinase (MAPK) in PECs. Inhibition of p44/42 MAPK suppressed albumin-induced MMP-9 secretion from glomerular PECs. Taken together, we have demonstrated that an up-regulation of MMP-9 in activated parietal epithelium is associated with a loss of adjacent podocytes in progressive diabetic nephropathy. Albumin overload may induce MMP-9 expression and secretion by PECs via the activation of p44/42 MAPK pathway. PMID:25849723

Thrombin-induced Migration and Matrix Metalloproteinase-9 Expression Are Regulated by MAPK and PI3K Pathways in C6 Glioma Cells

PubMed Central

Kim, Jiyoung; Lee, Jae-Won; Kim, Song-In; Choi, Yong-Joon; Lee, Won-Ki; Jeong, Myung-Ja; Cha, Sang-Hoon; Lee, Hee Jae; Chun, Wanjoo

2011-01-01

Glioblastoma multiforme is one of the most common and aggressive tumors in central nervous system. It often possesses characteristic necrotic lesions with hemorrhages, which increase the chances of exposure to thrombin. Thrombin has been known as a regulator of MMP-9 expression and cancer cell migration. However, the effects of thrombin on glioma cells have not been clearly understood. In the present study, influences of thrombin on glioma cell migration were examined using Boyden chamber migration assay and thrombin-induced changes in MMP-9 expression were measured using zymography, semi-quantitative RT-PCR, and Western blotting. Furthermore, underlying signaling pathways by which thrombin induces MMP-9 expression were examined. Thrombin-induced migration and MMP-9 expression were significantly potentiated in the presence of wortmannin, a PI3K inhibitor, whereas MAPK inhibitors suppressed thrombin-induced migration and MMP-9 expression in C6 glioma cells. The present data strongly demonstrate that MAPK and PI3K pathways evidently regulate thrombin-induced migration and MMP-9 expression of C6 glioma cells. Therefore, the control of these pathways might be a beneficial therapeutic strategy for treatment of invasive glioblastoma multiforme. PMID:21994479

Matrix Metallopeptidase 14 Plays an Important Role in Regulating Tumorigenic Gene Expression and Invasion Ability of HeLa Cells.

PubMed

Zhang, Ying-Hui; Wang, Juan-Juan; Li, Min; Zheng, Han-Xi; Xu, Lan; Chen, You-Guo

2016-03-01

The objectives of this study were to investigate the functional effect of matrix metallopeptidase 14 (MMP14) on cell invasion in cervical cancer cells (HeLa line) and to study the underlying molecular mechanisms. Expression vector of short hairpin RNA targeting MMP14 was treated in HeLa cells, and then, transfection efficiency was verified by a florescence microscope. Transwell assay was used to investigate cell invasion ability in HeLa cells. Quantitative polymerase chain reaction and Western blotting analysis were used to detect the expression of MMP14 and relative factors in messenger RNA and protein levels, respectively. Matrix metallopeptidase 14 short hairpin RNA expression vector transfection obviously decreased MMP14 expression in messenger RNA and protein levels. Down-regulation of MMP14 suppressed invasion ability of HeLa cells and reduced transforming growth factor β1 and vascular endothelial growth factor B expressions. Furthermore, MMP14 knockdown decreased bone sialoprotein and enhanced forkhead box protein L2 expression in both RNA and protein levels. Matrix metallopeptidase 14 plays an important role in regulating invasion of HeLa cells. Matrix metallopeptidase 14 knockdown contributes to attenuating the malignant phenotype of cervical cancer cell.

Effects of rosuvastatin on the production and activation of matrix metalloproteinase-2 and migration of cultured rat vascular smooth muscle cells induced by homocysteine.

PubMed

Shi, Ya-fei; Chi, Ju-fang; Tang, Wei-liang; Xu, Fu-kang; Liu, Long-bin; Ji, Zheng; Lv, Hai-tao; Guo, Hang-yuan

2013-08-01

To test the influence of homocysteine on the production and activation of matrix metalloproteinase-2 (MMP-2) and tissue inhibitors of matrix metalloproteinase-2 (TIMP-2) and on cell migration of cultured rat vascular smooth muscle cells (VSMCs). Also, to explore whether rosuvastatin can alter the abnormal secretion and activation of MMP-2 and TIMP-2 and migration of VSMCs induced by homocysteine. Rat VSMCs were incubated with different concentrations of homocysteine (50-5000 μmol/L). Western blotting and gelatin zymography were used to investigate the expressions and activities of MMP-2 and TIMP-2 in VSMCs in culture medium when induced with homocysteine for 24, 48, and 72 h. Transwell chambers were employed to test the migratory ability of VSMCs when incubated with homocysteine for 48 h. Different concentrations of rosuvastatin (10(-9)-10(-5) mol/L) were added when VSMCs were induced with 1000 μmol/L homocysteine. The expressions and activities of MMP-2 and TIMP-2 were examined after incubating for 24, 48, and 72 h, and the migration of VSMCs was also examined after incubating for 48 h. Homocysteine (50-1000 μmol/L) increased the production and activation of MMP-2 and expression of TIMP-2 in a dose-dependent manner. However, when incubated with 5000 μmol/L homocysteine, the expression of MMP-2 was up-regulated, but its activity was down-regulated. Increased homocysteine-induced production and activation of MMP-2 were reduced by rosuvastatin in a dose-dependent manner whereas secretion of TIMP-2 was not significantly altered by rosuvastatin. Homocysteine (50-5000 μmol/L) stimulated the migration of VSMCs in a dose-dependent manner, but this effect was eliminated by rosuvastatin. Homocysteine (50-1000 μmol/L) significantly increased the production and activation of MMP-2, the expression of TIMP-2, and the migration of VSMCs in a dose-dependent manner. Additional extracellular rosuvastatin can decrease the excessive expression and activation of MMP-2 and abnormal migration of VSMCs induced by homocysteine.

Cancer-associated fibroblasts affect breast cancer cell gene expression, invasion and angiogenesis.

PubMed

Eiro, Noemi; González, Lucía; Martínez-Ordoñez, Anxo; Fernandez-Garcia, Belen; González, Luis O; Cid, Sandra; Dominguez, Francisco; Perez-Fernandez, Román; Vizoso, Francisco J

2018-03-01

It has been reported that stromal cell features may affect the clinical outcome of breast cancer patients. Cancer associated fibroblasts (CAFs) represent one of the most abundant cell types within the breast cancer stroma. Here, we aimed to explore the influence of CAFs on breast cancer gene expression, as well as on invasion and angiogenesis. qRT-PCR was used to evaluate the expression of several cancer progression related genes (S100A4, TGFβ, FGF2, FGF7, PDGFA, PDGFB, VEGFA, IL-6, IL-8, uPA, MMP2, MMP9, MMP11 and TIMP1) in the human breast cancer-derived cell lines MCF-7 and MDA-MB-231, before and after co-culture with CAFs. Stromal mononuclear inflammatory cell (MIC) MMP11 expression was used to stratify primary tumors. In addition, we assessed the in vitro effects of CAFs on both MDA-MB-231 breast cancer cell invasion and endothelial cell (HUVEC) tube formation. We found that the expression levels of most of the genes tested were significantly increased in both breast cancer-derived cell lines after co-culture with CAFs from either MMP11+ or MMP11- MIC tumors. IL-6 and IL-8 showed an increased expression in both cancer-derived cell lines after co-culture with CAFs from MMP11+ MIC tumors. We also found that the invasive and angiogenic capacities of, respectively, MDA-MB-231 and HUVEC cells were increased after co-culture with CAFs, especially those from MMP11+ MIC tumors. Our data indicate that tumor-derived CAFs can induce up-regulation of genes involved in breast cancer progression. Our data additionally indicate that CAFs, especially those derived from MMP11+ MIC tumors, can promote breast cancer cell invasion and angiogenesis.

[Overexpressed miRNA-134b inhibits proliferation and invasion of CD133+ U87 glioma stem cells].

PubMed

Liu, Yifeng; Zhang, Baochao; Wen, Changming; Wen, Gongling; Zhou, Guoping; Zhang, Jingwei; He, Haifa; Wang, Ning; Li, Wei

2017-05-01

Objective To investigate the role of microRNA-134b (miR-134b) in the tumorigenesis of glioma stem cells (GSCs) and the possible molecular mechanism. Methods Real-time quantitative PCR (qRT-PCR) was used to evalate the expression of miR-134b in CD133 + and CD133 - U87 GSCs. A lentiviral vector overexpressing miR-134b in U87 GSCs was constructed, and the effect of miR-134b overexpression on matrix metalloproteinase-2 (MMP-2), MMP-9 and MMP-12 expressions at both mRNA and protein levels were detected by qRT-PCR and Western blotting, respectively. Transwell TM assay was performed to determine the effect of miR-134b overexpression on GSCs invasion ability. Tumor xenograft models in nude mice were established to evaluate the effect of miR-134b overexpression on tumorgenesis in vivo. Results The qRT-PCR showed that, compared with CD133 - cells, miR-134b was significantly down-regulated in CD133 + cells. Cell line over-expressing miR-134b was successfully established, and miR-134b was up-regulated significantly compared with empty vector control. Overexpression of miR-134b remarkably inhibited the invasion of U87 GSCs and the expression of MMP-12. However, overexpression of miR-134b did not affect MMP-2 and MMP-9 expressions. miR-134b also suppressed U87 GSCs xenograft growth in vivo. Tumor volume in tumor xenograft model group was significantly lower than that in control group, and tumor weight decreased by 42% in the former group. Conclusion Overexpression of miR-134b inhibits the growth and invasion of CD133 + GSCs.

Azilsartan Increases Levels of IL-10, Down-Regulates MMP-2, MMP-9, RANKL/RANK, Cathepsin K and Up-Regulates OPG in an Experimental Periodontitis Model

PubMed Central

Brito, Gerly Anne de Castro; de Medeiros, Caroline Addison Carvalho Xavier; Araújo, Lorena de Souza; do Nascimento, José Heriberto Oliveira; de Araújo Júnior, Raimundo Fernandes

2014-01-01

Aims The aim of this study was to evaluate the effects of azilsartan (AZT) on bone loss, inflammation, and the expression of matrix metallo proteinases (MMPs), receptor activator of nuclear factor κB ligand (RANKL), receptor activator of nuclear factor κB (RANK), osteoprotegerin (OPG), cyclooxygenase-2 (COX-2), and cathepsin K in periodontal tissue in a rat model of ligature-induced periodontitis. Materials and Methods Male Wistar albino rats were randomly divided into 5 groups of 10 rats each: (1) nonligated, water; (2) ligated, water; (3) ligated, 1 mg/kg AZT; (4) ligated, 5 mg/kg AZT; and (5) ligated, 10 mg/kg AZT. All groups were treated with saline or AZT for 10 days. Periodontal tissues were analyzed by histopathology and immunohistochemical detection of MMP-2, MMP-9, COX-2, RANKL, RANK, OPG, and cathepsin K. Levels of IL-1β, IL-10, TNF-α, myeloperoxidase (MPO), and glutathione (GSH) were determined by ELISA. Results Treatment with 5 mg/kg AZT resulted in reduced MPO (p<0.05) and IL-1β (p<0.05), increased levels of IL-10 (p<0.05), and reduced expression of MMP-2, MMP-9, COX-2, RANK, RANKL, cathepsin K, and increased expression of OPG. Conclusions These findings reveal that AZT increases anti-inflammatory cytokines and GSH and decreases bone loss in ligature-induced periodontitis in rats. PMID:24819928

S100A8/A9 regulates MMP-2 expression and invasion and migration by carcinoma cells

PubMed Central

Silva, Emmanuel J.; Argyris, Prokopios P.; Zou, Xianqiong; Ross, Karen F.; Herzberg, Mark C.

2014-01-01

Intracellular calprotectin (S100A8/A9) functions in the control of the cell cycle checkpoint at G2/M. Dysregulation of S100A8/A9 appears to cause loss of the checkpoint, which frequently characterizes head and neck squamous cell carcinoma (HNSCC). In the present study, we analyzed carcinoma cells for other S100A8/A9-directed changes in malignant phenotype. Using a S100A8/A9-negative human carcinoma cell line (KB), transfection to express S100A8 and S100A9 caused selective down-regulation of MMP-2 and inhibited in vitro invasion and migration. Conversely, silencing of endogenous S100A8 and S100A9 expression in TR146 cells, a well-differentiated HNSCC cell line, increased MMP-2 activity and in vitro invasion and migration. When MMP-2 expression was silenced, cells appeared to assume a less malignant phenotype. To more closely model the architecture of cell growth in vivo, cells were grown in a 3D collagen substrate, which was compared to 2D. Growth on 3D substrates caused greater MMP-2 expression. Whereas hypermethylation of CpG islands occurs frequently in HNSCC, S100A8/A9-dependent regulation of MMP-2 could not be explained by modification of the upstream promoters of MMP2 or TIMP2. Collectively, these results suggest that intracellular S100A8/A9 contributes to the cancer cell phenotype by modulating MMP-2 expression and activity to regulate cell migration and mobility. PMID:25236491

Leptin signaling enhances cell invasion and promotes the metastasis of human pancreatic cancer via increasing MMP-13 production.

PubMed

Fan, Yingchao; Gan, Yu; Shen, Yuling; Cai, Xiaojin; Song, Yanfang; Zhao, Fangyu; Yao, Ming; Gu, Jianren; Tu, Hong

2015-06-30

Emerging evidence has suggested that leptin, an adipokine related to energy homeostasis, plays a role in cancer growth and metastasis. However, its impact on pancreatic cancer is rarely studied. In this study, we found that leptin's functional receptor Ob-Rb was expressed in pancreatic cancer cell lines. Treatment with leptin enhanced the migration and invasion of pancreatic cancer cells but did not affect the proliferation of human pancreatic cancer cells. Leptin up-regulated the expression of matrix metalloproteinase-13 (MMP-13) via the JAK2/STAT3 signaling pathway. The overexpression of leptin was shown to significantly promote tumor growth and lymph node metastasis in a subcutaneous model and an orthotopic model of human pancreatic cancer, respectively. Furthermore, in human pancreatic cancer tissues, the expression of Ob-Rb was positively correlated with the MMP-13 level. The increased expression of either Ob-Rb or MMP-13 was significantly associated with lymph node metastasis and tended to be associated with the TNM stage in patients with pancreatic cancer. Our findings suggest that leptin enhances the invasion of pancreatic cancer through the increase in MMP-13 production, and targeting the leptin/MMP-13 axis could be an attractive therapeutic strategy for pancreatic cancer.

Leptin signaling enhances cell invasion and promotes the metastasis of human pancreatic cancer via increasing MMP-13 production

PubMed Central

Shen, Yuling; Cai, Xiaojin; Song, Yanfang; Zhao, Fangyu; Yao, Ming; Gu, Jianren; Tu, Hong

2015-01-01

Emerging evidence has suggested that leptin, an adipokine related to energy homeostasis, plays a role in cancer growth and metastasis. However, its impact on pancreatic cancer is rarely studied. In this study, we found that leptin's functional receptor Ob-Rb was expressed in pancreatic cancer cell lines. Treatment with leptin enhanced the migration and invasion of pancreatic cancer cells but did not affect the proliferation of human pancreatic cancer cells. Leptin up-regulated the expression of matrix metalloproteinase-13 (MMP-13) via the JAK2/STAT3 signaling pathway. The overexpression of leptin was shown to significantly promote tumor growth and lymph node metastasis in a subcutaneous model and an orthotopic model of human pancreatic cancer, respectively. Furthermore, in human pancreatic cancer tissues, the expression of Ob-Rb was positively correlated with the MMP-13 level. The increased expression of either Ob-Rb or MMP-13 was significantly associated with lymph node metastasis and tended to be associated with the TNM stage in patients with pancreatic cancer. Our findings suggest that leptin enhances the invasion of pancreatic cancer through the increase in MMP-13 production, and targeting the leptin/MMP-13 axis could be an attractive therapeutic strategy for pancreatic cancer. PMID:25948792

Matrix metalloproteinase-9 and vascular endothelial growth factor expression change in experimental retinal neovascularization.

PubMed

Di, Yu; Nie, Qing-Zhu; Chen, Xiao-Long

2016-01-01

To investigate the signal transduction mechanism of matrix metalloproteinase-9 (MMP-9) mediated- vascular endothelial growth factor (VEGF) expression and retinal neovascularization (RNV) in oxygen-induced retinopathy (OIR) model. C57BL/6J mice were divided into four groups: control group, OIR group, OIR control group (phosphate-buffered saline by intravitreal injection) and treated group [tissue inhibitor of matrix metalloproteinase-1 (TIMP-1) by intravitreal injection]. OIR model was established in C57BL/6J mice exposed to 75%±2% oxygen for 5d. mRNA level and protein expression of MMP-9, TIMP-1 and VEGF were measured by real-time polymerase chain reaction and Western blotting, and located by immunohistochemistry. Levels of MMP-9 and VEGF in retina were significantly increased in animals with OIR and OIR control group. Levels of TIMP-1 in retina was significantly reduced in animals with OIR and OIR control group. Furthermore, a significant correlation was found between MMP-9 and VEGF. Intravitreal injection of TIMP-1 significantly reduced MMP-9 and VEGF expression of the OIR mouse model (all P<0.05). These results demonstrate that MMP-9-mediated up-regulation of VEGF promotes RNV in retinopathy of prematurity (ROP). TIMP-1 may be a potential target for the prevention and treatment of ROP.

AT1-receptor blockade attenuates outward aortic remodeling associated with diet-induced obesity in mice.

PubMed

Krueger, Friedrich; Kappert, Kai; Foryst-Ludwig, Anna; Kramer, Frederike; Clemenz, Markus; Grzesiak, Aleksandra; Sommerfeld, Manuela; Paul Frese, Jan; Greiner, Andreas; Kintscher, Ulrich; Unger, Thomas; Kaschina, Elena

2017-08-01

The renin-angiotensin system (RAS) and obesity have been implicated in vascular outward remodeling, including aneurysms, but the precise mechanisms are not yet understood. We investigated the effect of the angiotensin receptor type 1 (AT1-receptor) antagonist telmisartan on aortic outward remodeling in a diet-induced obesity model in mice. C57/Black6J mice were fed either a low-fat diet (LFD) or a high-fat diet (HFD) for 14 weeks. One group of HFD mice was additionally exposed to telmisartan (3 mg/kg per day) for the last 4 weeks. HFD led to aortic outward remodeling, characterized by increased proteolysis, along with structural changes, such as fragmentation of elastic fibers and decreased elastin content. Vascular damage was associated with up-regulation of matrix metalloproteinase (MMP)-2 (MMP-2), MMP-3, MMP-12, cathepsin D, and cathepsin B. HFD aortae exhibited an enhanced inflammatory status, characterized by tumor necrosis factor α (TNF-α) and interleukin-1β (IL-1β) colocalized with adipocytes in the adventitia. HFD resulted in a significant increase in aortic dimensions, evident by ultrasound measurements. Telmisartan abolished aortic dilatation and preserved elastin content. HFD induced enhanced expression of aortic MMP-2, MMP-9, and TNF-α was abrogated by telmisartan. Adventitial proteolytic and inflammatory factors were also examined in samples from human abdominal aneurysms. The expression of TNF-α, IL-1β, and MMP-9 was higher in the adventitial fat of diseased vessels compared with healthy tissues. Finally, adipocytes treated with TNF-α showed enhanced MMP-2, MMP-3, and cathepsin D, which was prevented by telmisartan. Taken together, HFD in mice induced aortic dilatation with up-regulation of matrix degrading and inflammatory pathways similar to those seen in human aortic aneurysmatic tissue. The HFD-induced vascular pathology was reduced by AT1-receptor antagonist telmisartan. © 2017 The Author(s). Published by Portland Press Limited on behalf of the Biochemical Society.

Effects of tumor necrosis factor-alpha and interferon-gamma on expressions of matrix metalloproteinase-2 and -9 in human bladder cancer cells.

PubMed

Shin, K Y; Moon, H S; Park, H Y; Lee, T Y; Woo, Y N; Kim, H J; Lee, S J; Kong, G

2000-10-31

We have investigated the effects of tumor necrosis factor-alpha (TNF-alpha) and interferon (INF-gamma), the potent Bacillus Calmette-Guerin (BCG)-induced cytokines on the production of MMP-2, MMP-9, TIMP-1, TIMP-2 and MT1-MMP in high grade human bladder cancer cell lines, T-24, J-82 and HT-1376 cell lines. MMP-2 expression and activity were decreased in T-24 cells treated with both cytokines in a dose dependent manner. However, J-82 cells treated with TNF-alpha and INF-gamma revealed dose dependent increases of MMP-9 expression and activity with similar baseline expression and activity of MMP-2. HT-1376 cells after exposure to TNF-alpha only enhanced the expression and activity of MMP-9. These results indicate that TNF-alpha and INF-gamma could regulate the production of MMP-2 or MMP-9 on bladder cancer cells and their patterns of regulation are cell specific. Furthermore, this diverse response of bladder cancer cells to TNF-alpha and INF-gamma suggests that BCG immunotherapy may enhance the invasiveness of bladder cancer in certain conditions with induction of MMPs.

Interlukin-18 Is a Pivot Regulatory Factor on Matrix Metalloproteinase-13 Expression and Brain Astrocytic Migration.

PubMed

Chen, Jia-Hong; Tsai, Chon-Haw; Lin, Hsiao-Yun; Huang, Chien-Fang; Leung, Yuk-Man; Lai, Sheng-Wei; Tsai, Cheng-Fang; Chang, Pei-Chun; Lu, Dah-Yuu; Lin, Chingju

2016-11-01

The expression of matrix metalloproteinase-13 (MMP-13) has been shown to be elevated in some pathophysiological conditions and is involved in the degradation of extracellular matrix in astrocytes. In current study, the function of MMP-13 was further investigated. The conditioned medium (CM) collected from activated microglia increased interleukin (IL)-18 production and enhanced MMP-13 expression in astrocytes. Furthermore, treatment with recombinant IL-18 increased MMP-13 protein and mRNA levels in astrocytes. Recombinant IL-18 stimulation also increased the enzymatic activity of MMP-13 and the migratory activity of astrocytes, while administration of MMP-13 or pan-MMP inhibitors antagonized IL-18-induced migratory activity of astrocytes. In addition, administration of recombinant IL-18 to astrocytes led to the phosphorylation of JNK, Akt, or PKCδ, and treatment of astrocytes with JNK, PI3 kinase/Akt, or PKCδ inhibitors significantly decreased the IL-18-induced migratory activity. Taken together, the results suggest that IL-18-induced MMP-13 expression in astrocytes is regulated by JNK, PI3 kinase/Akt, and PKCδ signaling pathways. These findings also indicate that IL-18 is an important regulator leading to MMP-13 expression and cell migration in astrocytes.

Wip1 is associated with tumorigenity and metastasis through MMP-2 in human intrahepatic cholangiocarcinoma

PubMed Central

Liu, Sulai; Jiang, Bo; Li, Hao; He, Zili; Lv, Pin; Peng, Chuang; Wang, Yonggang; Cheng, Wei; Xu, Zhengquan; Chen, Wei; Liu, Zhengkai; Zhang, Bao; Shen, Shengqian; Xiang, Shuanglin

2017-01-01

Wip1 has been shown to correlate with the metastasis/invasion of several tumors. This study was designed to investigate the clinical significance and biological function of Wip1 in intrahepatic cholangiocarcinoma (ICC). The expression of Wip1 was investigated in sixty human ICC biopsy samples by immunohistochemistry. Transient and stable knockdown of Wip1 in two human ICC cells (ICC-9810 and SSP25) were established using short hairpin RNA expression vector. Immunohistochemistry revealed that Wip1 was up-regulated in human ICC tissues (47/60, 78.3%). High levels of Wip1 in human ICC correlated with metastasis to the lymph metastasis (P=0.022). Genetic depletion of Wip1 in ICC cells resulted in significantly inhibited proliferation and invasion compared with controls. Most importantly, Wip1 down-regulation impaired tumor migration capacity of ICC cells in vivo. Subsequent investigations revealed that matrix metalloproteinase-2 (MMP-2) is an important target of Wip1. Consistently, in human ICC tissues, Wip1 level was positively correlated with MMP-2 expression. Taken together, our founding indicates that Wip1 may be a crucial regulator in the tumorigenicity and invasion of human ICC, Wip1 exerts its pro-invasion function at least in part through the MMP-2 signaling pathway, suggesting Wip1 as a potential therapeutic target for ICC. PMID:28915621

The effect of newly synthesized progesterone derivatives on apoptotic and angiogenic pathway in MCF-7 breast cancer cells.

PubMed

Yahya, Shaymaa M M; Abdelhamid, Abdou O; Abd-Elhalim, Mervat M; Elsayed, Ghada H; Eskander, Emad F

2017-10-01

Due to its high potency and selectivity, anticancer agents consisting of combined molecules have gained great interests. The current study introduces newly synthesized progesterone derivatives of promising anticancer effect. Moreover, the pro-apoptotic and anti-angiogenic effects of these compounds were studied extensively. Several thiazole, pyridine, pyrazole, thiazolopyridine and pyrazolopyridine progesterone derivatives were synthesized. The structure of the novel progesterone derivatives was elucidated and confirmed using the analytical and spectral data. This novel derivatives were tested for their cytotoxic effect against human breast cancer cells (MCF-7) using neutral red uptake assay. Tested compounds showed anticancer activity against MCF-7 cancer cell line in the descending order of 7>2>3>8>6>9>4. The expression levels of Bcl-2, survivin, CCND1, CDC2, P53 and P21, VEGF, Hif-1α, MMP-2, MMP-9, Ang-1, Ang-2, and FGF-1 genes were investigated using QRT-PCR (Quantitative real time-polymerase chain reaction). The study clarified that compounds 2, 3, 4, 6, 7, 8 and 9 showed significant pro-apoptotic effect through the down regulation of Bcl-2., besides, survivin and CCND1 expression levels were down regulated by compounds 3, 4, 6, 7, 8, 9. However, Compound 4 may exert this pro-apoptotic effect through the up-regulation of P53 gene expression. On the other hand, the anti-angiogenic effect of these newly synthesized derivatives was due to their down regulation of VEGF, Ang-2, MMP-9 and FGF-1; and the up-regulation of HIF-1α and ang-1. This study recommended promising pro-apoptotic and anti-angiogenic anticancer agents acting through the regulation of key regulators of apoptosis, cell cycle genes, and pro-angiogenic genes. Copyright © 2017 Elsevier Inc. All rights reserved.

Analysis of expression of FLI1 and MMP1 in American cutaneous leishmaniasis caused by Leishmania braziliensis infection

PubMed Central

Almeida, Lucas; Silva, Juliana A.; Andrade, Viviane M.; Machado, Paulo; Jamieson, Sarra E.; Carvalho, Edgar M.

2017-01-01

FLI1 (Friend leukemia virus integration 1) and IL6 (interleukin 6; IL-6) are associated with Leishmania braziliensis susceptibility. Cutaneous lesions show exaggerated matrix metalloproteinase 1 (MMP1). In other skin diseases, FLI1 promoter methylation reduces FLI1 expression, and low FLI1 down-regulates MMP1. IL-6 increases FLI1 expression. We hypothesized that epigenetic regulation of FLI1 in cutaneous leishmaniasis, together with IL-6, might determine MMP1 expression. While generally low (<10%), percent FLI1 promoter methylation was lower (P=0.001) in lesion biopsies than normal skin. Contrary to expectation, a strong positive correlation occurred between FLI1 methylation and gene expression in lesions (r=0.98, P=0.0005) and in IL-6-treated L. braziliensis-infected macrophages (r=0.99, P=0.0004). In silico analysis of the FLI1 promoter revealed co-occurring active H3K27ac and repressive DNA methylation marks to enhance gene expression. FLI1 expression was enhanced between 3 and 24 hours post infection in untreated (P=0.0002) and IL-6-treated (P=0.028) macrophages. MMP1 was enhanced in lesion biopsies (P=0.0002), induced (P=0.007) in infected macrophages, but strongly inhibited by IL-6. No correlations occurred between FLI1 and MMP1 expression in lesions or infected macrophages (with/without IL-6). We conclude that MMP1 is regulated by factors other than FLI1, and that the influence of IL-6 on MMP1 was independent of its effect on FLI1. PMID:28119029

Requirement of matrix metalloproteinase-1 for intestinal homeostasis in the adult Drosophila midgut

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lee, Shin-Hae; Park, Joung-Sun; Kim, Young-Shin

Stem cells are tightly regulated by both intrinsic and extrinsic signals as well as the extracellular matrix (ECM) for tissue homeostasis and regenerative capacity. Matrix metalloproteinases (MMPs), proteolytic enzymes, modulate the turnover of numerous substrates, including cytokine precursors, growth factors, and ECM molecules. However, the roles of MMPs in the regulation of adult stem cells are poorly understood. In the present study, we utilize the Drosophila midgut, which is an excellent model system for studying stem cell biology, to show that Mmp1 is involved in the regulation of intestinal stem cells (ISCs). The results showed that Mmp1 is expressed inmore » the adult midgut and that its expression increases with age and with exposure to oxidative stress. Mmp1 knockdown or Timp-overexpressing flies and flies heterozygous for a viable, hypomorphic Mmp1 allele increased ISC proliferation in the gut, as shown by staining with an anti-phospho-histone H3 antibody and BrdU incorporation assays. Reduced Mmp1 levels induced intestinal hyperplasia, and the Mmp1depletion-induced ISC proliferation was rescued by the suppression of the EGFR signaling pathway, suggesting that Mmp1 regulates ISC proliferation through the EGFR signaling pathway. Furthermore, adult gut-specific knockdown and whole-animal heterozygotes of Mmp1 increased additively sensitivity to paraquat-induced oxidative stress and shortened lifespan. Our data suggest that Drosophila Mmp1 is involved in the regulation of ISC proliferation for maintenance of gut homeostasis. -- Highlights: Black-Right-Pointing-Pointer Mmp1 is expressed in the adult midgut. Black-Right-Pointing-Pointer Mmp1 is involved in the regulation of ISC proliferation activity. Black-Right-Pointing-Pointer Mmp1-related ISC proliferation is associated with EGFR signaling. Black-Right-Pointing-Pointer Mmp1 in the gut is required for the intestinal homeostasis and longevity.« less

Differential effects of tumor necrosis factor-α on matrix metalloproteinase-2 expression in human myometrial and uterine leiomyoma smooth muscle cells.

PubMed

Wang, Yuebing; Feng, Guowei; Wang, Jiyuan; Zhou, Yu; Liu, Yixin; Shi, Yiquan; Zhu, Yingjun; Lin, Wanjun; Xu, Yang; Li, Zongjin

2015-01-01

Does tumor necrosis factor-α (TNF-α) differentially regulate matrix metalloproteinase-2 (MMP-2) expression in leiomyomas compared with normal myometrium? TNF-α up-regulates MMP-2 expression and stimulates cell migration through the activation of extracellular signal-regulated kinase (ERK) signaling pathway in leiomyoma smooth muscle cells (SMCs), but not in normal myometrial SMCs. Uterine leiomyoma, the benign smooth muscle cell tumor, is the single most common indication for hysterectomy. High expression of MMPs or TNF-α has been reported in uterine leiomyomas; however, the molecular mechanism underlying these observations remains unknown. Samples were obtained between 2009 and 2013 from 12 women of reproductive age at the proliferative phase of the menstrual cycle by hysterectomy. Leiomyomas and matched normal myometrium from each woman were analyzed in vitro. Western blot, RT-qPCR and a wound-healing assay were used to investigate the effects of TNF-α on MMP-2 expression and intracellular signal transduction in cultured SMCs from leiomyomas and matched myometrium. Western blot and RT-qPCR analyses using tissues from clinical patients showed that the levels of MMP-2 protein (P = 0.008) and mRNA (P = 0.009) were significantly higher in uterine leiomyomas compared with their matched myometrium. Treatment with TNF-α significantly up-regulated the protein (P = 0.039) and mRNA (P = 0.037) levels of MMP-2 in cultured leiomyoma SMCs but not in matched myometrial SMCs. The extracellular signal-regulated kinase (ERK) and nuclear factor-kappa B (NF-κB) pathways were activated by TNF-α in leiomyoma SMCs. Specific inhibitors of the ERK or NF-κB pathway (PD98059 or Bay11-7082) suppressed TNF-α-induced MMP-2 expression in leiomyoma SMCs. The wound-healing assay revealed that TNF-α promoted the migration of cultured leiomyoma SMCs (P = 0.036); however, PD98059 compromised the cell migration triggered by TNF-α. This study is descriptive and although we observed clear differential regulation of MMP-2 by TNF-α at mRNA and protein levels in leiomyoma, future studies are needed to identify why the difference in TNF-α response exists between human leiomyoma tissue and normal myometrium. Including some of the experiments such as transfection studies for TNF-α and MMP-2 promoter mapping could have added more insight as to why this difference exists. In addition, further studies in vivo are needed to verify the results obtained from primary cultured SMCs. Considering the positive effect of TNF-α on leiomyoma SMC migration, strategies targeting TNF-α, in parallel with the production of more specific inhibitors of MMPs, may provide alternative therapeutic approaches for the treatment of leiomyoma. This work was partially supported by grants from the Program for New Century Excellent Talents in University (NCET-12-0282), National Natural Science Foundation of China (81371620) and Tianjin Natural Science Foundation (12JCZDJC24900). The authors have no conflicts of interest to declare. © The Author 2014. Published by Oxford University Press on behalf of the European Society of Human Reproduction and Embryology. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

Impressic acid from Acanthopanax koreanum, possesses matrix metalloproteinase-13 down-regulating capacity and protects cartilage destruction.

PubMed

Lim, Hyun; Min, Dong Suk; Yun, Han Eul; Kim, Kil Tae; Sun, Ya Nan; Dat, Le Duc; Kim, Young Ho; Kim, Hyun Pyo

2017-09-14

Acanthopanax koreanum (Araliaceae) has been used in traditional medicine for enhancing vitality, rheumatism, and bone-related pains. But its activity on cartilage protection has not been known yet. Matrix metalloproteinase (MMP)-13 has an important role in degrading cartilage materials under pathologic conditions such as arthritis. The present study was designed to find the inhibitory activity of impressic acid on MMP-13 expression and cartilage protective action. 70% ethanol extract of Acanthopanax koreanum leaves and impressic acid, a major constituent isolated from the same plant materials, were examined on MMP-13 down-regulating capacity in IL-1β-treated human chondrocyte cell line (SW1353) and rabbit cartilage explants. In IL-1β-treated SW1353 cells, impressic acid significantly and concentration-dependently inhibited MMP-13 expression at 0.5-10μM. Impressic acid was found to be able to inhibit MMP-13 expression by blocking the phosphorylation of signal transducer and activator of transcription-1/-2 (STAT-1/-2) and activation of c-Jun and c-Fos among the cellular signaling pathways involved. Further, impressic acid was found to inhibit the expression of MMP-13 mRNA (47.7% inhibition at 10μM), glycosaminoglycan release (42.2% reduction at 10μM) and proteoglycan loss in IL-1-treated rabbit cartilage explants culture. In addition, a total of 21 lupane-type triterpenoids structurally-related to impressic acid were isolated from the same plant materials and their suppressive activities against MMP-13 expression were also examined. Among these derivatives, compounds 2, 3, 16, and 18 clearly down-regulated MMP-13 expression. However, impressic acid was more potent than these derivatives in down-regulating MMP-13 expression. Impressic acid, its related triterpenoids, and A. koreanum extract have potential as therapeutic agents to prevent cartilage degradation by inhibiting matrix protein degradation. Copyright © 2017 Elsevier Ireland Ltd. All rights reserved.

Local Inflammation Alters MMP-2 and MMP-9 Gelatinase Expression Associated with the Severity of Nifedipine-Induced Gingival Overgrowth: a Rat Model Study.

PubMed

Li, Wu-Li; Wu, Cheng-Hai; Yang, Jun; Tang, Min; Chen, Long-Jie; Zhao, Shou-Liang

2015-08-01

Nifedipine-induced gingival overgrowth (NIGO) is characterized by cell proliferation and extracellular matrix (ECM) component accumulation in gingival connective tissues, with varying degrees of inflammation and fibrosis. Impaired collagen and ECM homeostasis may be among the underlying molecular mechanisms that lead to the fibrotic changes that occur in drug-induced gingival overgrowth (DIGO). Because matrix metalloproteinases (MMPs) play vital roles in regulating collagen and ECM metabolism, many studies have been performed to reveal the relationship between MMPs and DIGO. It is thought that the gelatinases MMP-2 and MMP-9, both type IV collagenases, are involved in the development of tissue inflammation and organ fibrosis. However, the few studies regarding gelatinase expression in DIGO are controversial. Recent studies have demonstrated the inhibitory effect of cyclosporine A (CsA) on gelatinase expression and/or activity; however, similar changes have yet to be detected in Nif-treated gingival tissues. In this study, we verified that Nif treatment could lead to gingival overgrowth in rats and that gingival inflammation played a pro-proliferative role in NIGO development. Additionally, we examined the temporal expression of gelatinases on days 0, 7, 14, 21, 30, and 40 during NIGO development. The aim was to investigate whether MMP-2 and MMP-9 played significant roles in regulating NIGO development and progression. MMP-2 gene expression was not altered by Nif treatment alone but was significantly inhibited by Nif treatment for 30 days in the presence of local inflammation. However, no significant alterations in MMP-2 protein expression were detected in the Nif-treated gingival tissue, regardless of the presence or absence of local inflammation. Moreover, Nif treatment could lead to transient and significant increases in MMP-9 gene and protein expression levels in the presence of local inflammation. In particular, active MMP-9 expression increased significantly in the gingival tissue that received the combined effect of Nif and ligation treatment; besides, a temporal, but not significant, change was also observed in the gingival tissue that received Nif treatment alone. Taken together, these results provided evidence that temporal changes in MMP-2 and MMP-9 expression occurred during NIGO development. Nif treatment accompanied by local inflammation regulated MMP-2 and MMP-9 expression, primarily MMP-9, which was most likely associated with NIGO severity. Thus, MMP-9 is a potential contributing factor in the process of NIGO development.

S100A8/A9 regulates MMP-2 expression and invasion and migration by carcinoma cells.

PubMed

Silva, Emmanuel J; Argyris, Prokopios P; Zou, Xianqiong; Ross, Karen F; Herzberg, Mark C

2014-10-01

Intracellular calprotectin (S100A8/A9) functions in the control of the cell cycle checkpoint at G2/M. Dysregulation of S100A8/A9 appears to cause loss of the checkpoint, which frequently characterizes head and neck squamous cell carcinoma (HNSCC). In the present study, we analyzed carcinoma cells for other S100A8/A9-directed changes in malignant phenotype. Using a S100A8/A9-negative human carcinoma cell line (KB), transfection to express S100A8 and S100A9 caused selective down-regulation of MMP-2 and inhibited in vitro invasion and migration. Conversely, silencing of endogenous S100A8 and S100A9 expression in TR146 cells, a well-differentiated HNSCC cell line, increased MMP-2 activity and in vitro invasion and migration. When MMP-2 expression was silenced, cells appeared to assume a less malignant phenotype. To more closely model the architecture of cell growth in vivo, cells were grown in a 3D collagen substrate, which was compared to 2D. Growth on 3D substrates caused greater MMP-2 expression. Whereas hypermethylation of CpG islands occurs frequently in HNSCC, S100A8/A9-dependent regulation of MMP-2 could not be explained by modification of the upstream promoters of MMP2 or TIMP2. Collectively, these results suggest that intracellular S100A8/A9 contributes to the cancer cell phenotype by modulating MMP-2 expression and activity to regulate cell migration and mobility. Published by Elsevier Ltd.

Matrine Exerts a Strong Anti-Arthritic Effect on Type II Collagen-Induced Arthritis in Rats by Inhibiting Inflammatory Responses

PubMed Central

Pu, Jiang; Fang, Fan-Fu; Li, Xiu-Qing; Shu, Zhi-Heng; Jiang, Yi-Ping; Han, Ting; Peng, Wei; Zheng, Cheng-Jian

2016-01-01

To investigate anti-arthritic effects of matrine isolated from the roots of S. flavescens on type II collagen-induced arthritis (CIA) in rats and to explore its related potential mechanisms, CIA rats were established and administered with matrine (20, 40 or 80 mg/kg/days, for 30 days). Subsequently, blood was collected to determine serum levels of TNF-α, IL-1β, IL-6, IL-8, IL-17A, IL-10, MMP-2, MMP-3 and MMP-9, and hind paws and knee joints were collected for histopathological examination. Furthermore, indices of the thymus and spleen were determined, and synovial tissues were collected to determine the protein expressions of p-IκB, IκB, Cox-2 and iNOS. Our results indicated that matrine significantly suppressed inflammatory reactions and synovial tissue destruction. Matrine inhibited paw swelling, arthritis indices and weight loss in CIA rats. Additionally, matrine decreased the levels of TNF-α, IL-1β, IL-6, IL-8, IL-17A, MMP-2, MMP-3 and MMP-9. Matrine also down-regulated expressions of p-IκB, Cox-2, and iNOS but up-regulated IκB in synovial tissues in CIA rats. The results suggested matrine possesses an anti-arthritic effect in CIA rats via inhibiting the release of pro-inflammatory cytokines and proteins that promote the NF-κB pathway. PMID:27571073

Ethanol increases matrix metalloproteinase-12 expression via NADPH oxidase-dependent ROS production in macrophages

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kim, Mi Jin; Nepal, Saroj; Lee, Eung-Seok

2013-11-15

Matrix metalloproteinase-12 (MMP-12), an enzyme responsible for degradation of extracellular matrix, plays an important role in the progression of various diseases, including inflammation and fibrosis. Although most of those are pathogenic conditions induced by ethanol ingestion, the effect of ethanol on MMP-12 has not been explored. In the present study, we investigated the effect of ethanol on MMP-12 expression and its potential mechanisms in macrophages. Here, we demonstrated that ethanol treatment increased MMP-12 expression in primary murine peritoneal macrophages and RAW 264.7 macrophages at both mRNA and protein levels. Ethanol treatment also significantly increased the activity of nicotinamide adenine dinucleotidemore » (NADPH) oxidase and the expression of NADPH oxidase-2 (Nox2). Pretreatment with an anti-oxidant (N-acetyl cysteine) or a selective inhibitor of NADPH oxidase (diphenyleneiodonium chloride (DPI)) prevented ethanol-induced MMP-12 expression. Furthermore, knockdown of Nox2 by small interfering RNA (siRNA) prevented ethanol-induced ROS production and MMP-12 expression in RAW 264.7 macrophages, indicating a critical role for Nox2 in ethanol-induced intracellular ROS production and MMP-12 expression in macrophages. We also showed that ethanol-induced Nox2 expression was suppressed by transient transfection with dominant negative IκB-α plasmid or pretreatment with Bay 11-7082, a selective inhibitor of NF-κB, in RAW 264.7 macrophages. In addition, ethanol-induced Nox2 expression was also attenuated by treatment with a selective inhibitor of p38 MAPK, suggesting involvement of p38 MAPK/NF-κB pathway in ethanol-induced Nox2 expression. Taken together, these results demonstrate that ethanol treatment elicited increase in MMP-12 expression via increase in ROS production derived from Nox2 in macrophages. - Highlights: • Ethanol increases ROS production through up-regulation of Nox2 in macrophages. • Enhanced oxidative stress contributes to ethanol-induced MMP-12 expression. • p38 MAPK/NF-κB signaling pathway modulates ethanol-induced Nox2 expression.« less

Regulation of Membrane-Type 4 Matrix Metalloproteinase by SLUG Contributes to Hypoxia-Mediated Metastasis12

PubMed Central

Huang, Chi-Hung; Yang, Wen-Hao; Chang, Shyue-Yih; Tai, Shyh-Kuan; Tzeng, Cheng-Hwei; Kao, Jung-Yie; Wu, Kou-Juey; Yang, Muh-Hwa

2009-01-01

The hypoxic tumor environment has been shown to be critical to cancer metastasis through the promotion of angiogenesis, induction of epithelial-mesenchymal transition (EMT), and acquisition of invasive potential. However, the impact of hypoxia on the expression profile of the proteolytic enzymes involved in invasiveness is relatively unknown. Membrane-type 4 matrix metalloproteinase (MT4-MMP) is a glycosyl-phosphatidyl inositol-anchored protease that has been shown to be overexpressed in human cancers. However, detailed mechanisms regarding the regulation and function of MT4-MMP expression in tumor cells remain unknown. Here, we demonstrate that hypoxia or overexpression of hypoxia-inducible factor-1α (HIF-1α) induced MT4-MMP expression in human cancer cells. Activation of SLUG, a transcriptional factor regulating the EMT process of human cancers, by HIF-1α was critical for the induction of MT4-MMP under hypoxia. SLUG regulated the transcription of MT4-MMP through direct binding to the E-box located in its proximal promoter. Short-interference RNA-mediated knockdown of MT4-MMP attenuated in vitro invasiveness and in vivo pulmonary colonization of tumor cells without affecting cell migratory ability. MT4-MMP promoted invasiveness and pulmonary colonization through modulation of the expression profile of MMPs and angiogenic factors. Finally, coexpression of HIF-1α and MT4-MMP in human head and neck cancer was predictive of a worse clinical outcome. These findings establish a novel signaling pathway for hypoxia-mediated metastasis and elucidate the underlying regulatory mechanism and functional significance of MT4-MMP in cancer metastasis. PMID:20019845

Matrix Metalloproteinase Expression in the Rat Myometrium During Pregnancy, Term Labor, and Postpartum1

PubMed Central

Nguyen, Tina Tu-Thu Ngoc; Shynlova, Oksana; Lye, Stephen J.

2016-01-01

Pregnancy, spontaneous term labor (TL), and postpartum (PP) involution are associated with changes in the cellular and extracellular matrix composition of the uterus. Both the uterine smooth muscle (myometrium) and the infiltrating peripheral blood leukocytes involved in the activation of labor secrete extracellular matrix-degrading enzymes (matrix metalloproteinases, MMPs) that can modulate cellular behavior and barrier function. MMP expression is induced by mechanical stretch in several tissues. We hypothesized that the expression and activity of myometrial MMPs and their tissue inhibitors (TIMPs) are modulated in preparation for TL and PP involution and are regulated by mechanical stretch of uterine walls imposed by the growing fetus. Myometrial tissues were collected from bilaterally and unilaterally pregnant rats across gestation, TL, and PP. Total RNA and proteins were subjected to real-time PCR and immunoblotting, respectively, and tissue localization and activity was examined by immunohistochemistry and in situ zymography. We found that Mmp7, Mmp11, and Mmp12 mRNA levels were upregulated during TL and PP, while Mmp2, Mmp3, Mmp8, Mmp9, Mmp10, and Mmp13 mRNAs were only upregulated during PP. Timp1–Timp4 were stably expressed throughout gestation with some fluctuations PP. Active MMP2 was induced in the empty uterine horn during gestation and in the gravid PP uterus, suggesting negative regulation by biological mechanical stretch. We conclude that specific subsets of uterine MMPs are differentially regulated in the rat myometrium in preparation for two major events: TL and PP uterine involution. PMID:27251092

Alcohol modulation of cardiac matrix metalloproteinases (MMPs) and tissue inhibitors of MMPs favors collagen accumulation.

PubMed

El Hajj, Elia C; El Hajj, Milad C; Voloshenyuk, Tetyana G; Mouton, Alan J; Khoutorova, Elena; Molina, Patricia E; Gilpin, Nicholas W; Gardner, Jason D

2014-02-01

Chronic alcohol consumption has been shown in human and animal studies to result in collagen accumulation, myocardial fibrosis, and heart failure. Cardiac fibroblasts produce collagen and regulate extracellular matrix (ECM) homeostasis through the synthesis and activity of matrix metalloproteinases (MMPs) and tissue inhibitors of MMPs (TIMPs), with the balance of MMPs/TIMPs determining the rate of collagen turnover. Dynamic changes of MMP and TIMP expression were reported in alcohol-induced hepatic fibrosis; however, the effect of alcohol on MMP/TIMP balance in the heart and cardiac fibroblasts is unknown. We hypothesized that alcohol exposure alters cardiac fibroblast MMP and TIMP expression to promote collagen accumulation in the heart. Cardiac fibroblasts isolated from adult rats were cultured in the presence of alcohol (12.5 to 200 mM) for 48 hours. MMP, TIMP, and collagen type I and III expression were assayed by Western blot analysis. Hydroxyproline (HPro) was used as a marker of collagen production. The in vivo cardiac effects of ethanol (EtOH) were determined using rats exposed to EtOH vapor for 2 weeks, resulting in blood alcohol levels of 150 to 200 mg/dl. Cardiac collagen volume fraction (CVF), as well as MMP, TIMP, and collagen expression, was assessed. EtOH-exposed rats exhibited up-regulation of TIMP-1, TIMP-3 and TIMP-4 in the heart, with no significant increases in MMPs. Cardiac fibroblasts exhibited transformation to a profibrotic phenotype following exposure to alcohol. These changes were reflected by increased α-smooth muscle actin and collagen I and III expression, as well as increased collagen secretion. In vivo EtOH exposure also produced fibrosis, indicated by increased CVF and expression of collagens. Alcohol exposure modulates cardiac fibroblast MMP/TIMP expression favoring a profile associated with collagen accumulation. Our data suggest that this disrupted MMP/TIMP profile may contribute to the development of myocardial fibrosis and cardiac dysfunction resulting from chronic alcohol abuse. Copyright © 2013 by the Research Society on Alcoholism.

IL-1β-induced, matrix metalloproteinase-3-regulated proliferation of embryonic stem cell-derived odontoblastic cells is mediated by the Wnt5 signaling pathway

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ozeki, Nobuaki; Hase, Naoko; Hiyama, Taiki

2014-10-15

We previously established a method for differentiating induced pluripotent stem cells and embryonic stem (ES) cells into α2 integrin-positive odontoblast-like cells. We also reported that interleukin (IL)-1β induces matrix metalloproteinase (MMP)-3-regulated cell proliferation and suppresses apoptosis in these cells, suggesting that MMP-3 plays a potentially unique physiological role in the regeneration of odontoblast-like cells. Here, we examined whether up-regulation of MMP-3 activity by IL-1β was mediated by Wnt signaling and led to increased proliferation of odontoblast-like cells. IL-1β increased mRNA and protein levels of Wnt5a, Wnt5b and the Wnt receptor Lrp5. Exogenous Wnt5a and Wnt5b were found to increase MMP-3more » mRNA, protein and activity, and interestingly the rate of proliferation in these cells. Treatment with siRNAs against Wnt5a, Wnt5b and Lrp5 suppressed the IL-1β-induced increase in MMP-3 expression and suppressed cell proliferation, an effect rescued by application of exogenous Wnt5. These results demonstrate the sequential involvement of Wnt5, Lrp5 and MMP-3 in effecting IL-1β-induced proliferation of ES cell-derived odontoblast-like cells. - Highlights: • IL-1β induces Wnt5, Lrp5/Fzd9 and MMP-3 in ES cell-derived odontoblast-like cells. • IL-1β-induced Wnt5 expression results in increased cell proliferation. • Exogenous Wnt5 increases MMP-3 activity and cell proliferation. • Exogenous Wnt5 rescues IL-1β-driven proliferation with anti-Wnt5 siRNA suppression. • IL-1β-induced cell proliferation involves Wnt5, Lrp5, and MMP-3 sequentially.« less

Analysis of expression of FLI1 and MMP1 in American cutaneous leishmaniasis caused by Leishmania braziliensis infection.

PubMed

Almeida, Lucas; Silva, Juliana A; Andrade, Viviane M; Machado, Paulo; Jamieson, Sarra E; Carvalho, Edgar M; Blackwell, Jenefer M; Castellucci, Léa C

2017-04-01

FLI1 (Friend leukemia virus integration 1) and IL6 (interleukin 6; IL-6) are associated with Leishmania braziliensis susceptibility. Cutaneous lesions show exaggerated matrix metalloproteinase 1 (MMP1). In other skin diseases, FLI1 promoter methylation reduces FLI1 expression, and low FLI1 down-regulates MMP1. IL-6 increases FLI1 expression. We hypothesized that epigenetic regulation of FLI1 in cutaneous leishmaniasis, together with IL-6, might determine MMP1 expression. While generally low (<10%), percent FLI1 promoter methylation was lower (P=0.001) in lesion biopsies than normal skin. Contrary to expectation, a strong positive correlation occurred between FLI1 methylation and gene expression in lesions (r=0.98, P=0.0005) and in IL-6-treated L. braziliensis-infected macrophages (r=0.99, P=0.0004). In silico analysis of the FLI1 promoter revealed co-occurring active H3K27ac and repressive DNA methylation marks to enhance gene expression. FLI1 expression was enhanced between 3 and 24hour post infection in untreated (P=0.0002) and IL-6-treated (P=0.028) macrophages. MMP1 was enhanced in lesion biopsies (P=0.0002), induced (P=0.007) in infected macrophages, but strongly inhibited by IL-6. No correlations occurred between FLI1 and MMP1 expression in lesions or infected macrophages (with/without IL-6). We conclude that MMP1 is regulated by factors other than FLI1, and that the influence of IL-6 on MMP1 was independent of its effect on FLI1. Copyright © 2017 Elsevier B.V. All rights reserved.

ING2 is upregulated in colon cancer and increases invasion by enhanced MMP13 expression

PubMed Central

Kumamoto, Kensuke; Fujita, Kaori; Kurotani, Reiko; Saito, Motonobu; Unoki, Motoko; Hagiwara, Nobutoshi; Shiga, Hideaki; Bowman, Elise D.; Yanaihara, Nozomu; Okamura, Shu; Nagashima, Makoto; Miyamoto, Kotaro; Takenoshita, Seiichi; Yokota, Jun; Harris, Curtis C.

2009-01-01

Inhibitor of growth 2 (ING2) is associated with chromatin remodeling and regulation of gene expression by binding to a methylated histone H3K4 residue and recruiting HDAC complexes to the region. The aim of our study is to investigate the regulation of ING2 expression and the clinical significance of upregulated ING2 in colon cancer. Here, we show that the ING2 mRNA level in colon cancer tissue increased to more than twice than that in normal mucosa in the 45% of colorectal cancer cases that we examined. A putative NF-κB binding site was found in the ING2 promoter region. We confirmed that NF-κB could bind to the ING2 promoter by EMSA and luciferase assays. Subsequent microarray analyses revealed that ING2 upregulates expression of matrix metalloproteinase 13 (MMP13), which enhances cancer invasion and metastasis. ING2 regulation of MMP13 expression was confirmed in both ING2 overexpression and knock down experiments. MMP13 expression was further induced by coexpression of ING2 with HDAC1 or with mSin3A, suggesting that the ING2-HDAC1-mSin3A complex members regulates expression of MMP13. In vitro invasion assay was performed to determine functional significance of ING2 upregulation. ING2 overexpressed cells exhibited greater invasive potential. Taken together, upregulation of ING2 was associated with colon cancer and MMP13-dependent cellular invasion, indicating that ING2 expression might be involved with cancer invasion and metastasis. PMID:19437536

Newly generated heparanase knock-out mice unravel co-regulation of heparanase and matrix metalloproteinases.

PubMed

Zcharia, Eyal; Jia, Juan; Zhang, Xiao; Baraz, Lea; Lindahl, Ulf; Peretz, Tamar; Vlodavsky, Israel; Li, Jin-Ping

2009-01-01

Heparanase, a mammalian endo-beta-D-glucuronidase, specifically degrades heparan sulfate proteoglycans ubiquitously associated with the cell surface and extracellular matrix. This single gene encoded enzyme is over-expressed in most human cancers, promoting tumor metastasis and angiogenesis. We report that targeted disruption of the murine heparanase gene eliminated heparanase enzymatic activity, resulting in accumulation of long heparan sulfate chains. Unexpectedly, the heparanase knockout (Hpse-KO) mice were fertile, exhibited a normal life span and did not show prominent pathological alterations. The lack of major abnormalities is attributed to a marked elevation in the expression of matrix metalloproteinases, for example, MMP2 and MMP14 in the Hpse-KO liver and kidney. Co-regulation of heparanase and MMPs was also noted by a marked decrease in MMP (primarily MMP-2,-9 and 14) expression following transfection and over-expression of the heparanase gene in cultured human mammary carcinoma (MDA-MB-231) cells. Immunostaining (kidney tissue) and chromatin immunoprecipitation (ChIP) analysis (Hpse-KO mouse embryonic fibroblasts) suggest that the newly discovered co-regulation of heparanase and MMPs is mediated by stabilization and transcriptional activity of beta-catenin. The lack of heparanase expression and activity was accompanied by alterations in the expression level of MMP family members, primarily MMP-2 and MMP-14. It is conceivable that MMP-2 and MMP-14, which exert some of the effects elicited by heparanase (i.e., over branching of mammary glands, enhanced angiogenic response) can compensate for its absence, in spite of their different enzymatic substrate. Generation of viable Hpse-KO mice lacking significant abnormalities may provide a promising indication for the use of heparanase as a target for drug development.

Ovarian Expression and Regulation of the Stromelysins During the Periovulatory Period in the Human and the Rat1

PubMed Central

McCord, Lauren A.; Li, Feixue; Rosewell, Katherine L.; Brännström, Mats; Curry, Thomas E.

2011-01-01

ABSTRACT The matrix metalloproteinases (MMPs) are postulated to facilitate follicular rupture. In the present study, expression of the stromelysins (MMP3, MMP10, MMP11) was analyzed in the periovulatory human and rat ovary. Human granulosa and theca cells were collected from the dominant follicle at various times after human chorionic gonadotropin (hCG). Intact rat ovaries, granulosa cells, and residual tissue (tissue remaining after granulosa cell collection) were isolated from equine CG (eCG)-hCG-primed animals. Mmp10 mRNA was highly induced in human granulosa and theca cells and intact rat ovaries, granulosa cells, and residual tissue. Localization of MMP10 to granulosa and theca cells in both human and rat ovarian follicles was confirmed by immunohistochemistry. Mmp3 mRNA was unchanged in human cells and rat granulosa cells, but increased in intact rat ovaries and residual tissue. Mmp11 mRNA decreased following hCG treatment in human granulosa and theca cells as well as rat granulosa cells. Regulation of Mmp10 in cultured rat granulosa cells revealed that the EGF inhibitor AG1478 and the progesterone receptor antagonist RU486 suppressed the induction of Mmp10 mRNA, whereas the prostaglandin inhibitor NS398 had no effect. Studies on the Mmp10 promoter demonstrated that forskolin plus PMA stimulated promoter activity, which was dependent upon a proximal AP1 site. In conclusion, there are divergent patterns of stromelysin expression associated with ovulation, with a marked induction of Mmp10 mRNA and a decrease in Mmp11 mRNA, yet a species-dependent pattern on Mmp3 mRNA expression. The induction of Mmp10 expression suggests an important role for this MMP in the follicular changes associated with ovulation and subsequent luteinization. PMID:22116802

Ovarian expression and regulation of the stromelysins during the periovulatory period in the human and the rat.

PubMed

McCord, Lauren A; Li, Feixue; Rosewell, Katherine L; Brännström, Mats; Curry, Thomas E

2012-03-01

The matrix metalloproteinases (MMPs) are postulated to facilitate follicular rupture. In the present study, expression of the stromelysins (MMP3, MMP10, MMP11) was analyzed in the periovulatory human and rat ovary. Human granulosa and theca cells were collected from the dominant follicle at various times after human chorionic gonadotropin (hCG). Intact rat ovaries, granulosa cells, and residual tissue (tissue remaining after granulosa cell collection) were isolated from equine CG (eCG)-hCG-primed animals. Mmp10 mRNA was highly induced in human granulosa and theca cells and intact rat ovaries, granulosa cells, and residual tissue. Localization of MMP10 to granulosa and theca cells in both human and rat ovarian follicles was confirmed by immunohistochemistry. Mmp3 mRNA was unchanged in human cells and rat granulosa cells, but increased in intact rat ovaries and residual tissue. Mmp11 mRNA decreased following hCG treatment in human granulosa and theca cells as well as rat granulosa cells. Regulation of Mmp10 in cultured rat granulosa cells revealed that the EGF inhibitor AG1478 and the progesterone receptor antagonist RU486 suppressed the induction of Mmp10 mRNA, whereas the prostaglandin inhibitor NS398 had no effect. Studies on the Mmp10 promoter demonstrated that forskolin plus PMA stimulated promoter activity, which was dependent upon a proximal AP1 site. In conclusion, there are divergent patterns of stromelysin expression associated with ovulation, with a marked induction of Mmp10 mRNA and a decrease in Mmp11 mRNA, yet a species-dependent pattern on Mmp3 mRNA expression. The induction of Mmp10 expression suggests an important role for this MMP in the follicular changes associated with ovulation and subsequent luteinization.

Reciprocal regulation by hypoxia-inducible factor-2α and the NAMPT-NAD(+)-SIRT axis in articular chondrocytes is involved in osteoarthritis.

PubMed

Oh, H; Kwak, J-S; Yang, S; Gong, M-K; Kim, J-H; Rhee, J; Kim, S K; Kim, H-E; Ryu, J-H; Chun, J-S

2015-12-01

Hypoxia-inducible factor-2α (HIF-2α) transcriptionally upregulates Nampt in articular chondrocytes. NAMPT, which exhibits nicotinamide phosphoribosyltransferase activity, in turn causes osteoarthritis (OA) in mice by stimulating the expression of matrix-degrading enzymes. Here, we sought to elucidate whether HIF-2α activates the NAMPT-NAD(+)-SIRT axis in chondrocytes and thereby contributes to the pathogenesis of OA. Assays of NAD levels, SIRT activity, reporter gene activity, mRNA, and protein levels were conducted in primary cultured mouse articular chondrocytes. Experimental OA in mice was induced by intra-articular (IA) injection of adenovirus expressing HIF-2α (Ad-Epas1) or NAMPT (Ad-Nampt). The functions of SIRT in OA were examined by IA co-injection of SIRT inhibitors or adenovirus expressing individual SIRT isoforms or shRNA targeting specific SIRT isoforms. HIF-2α activated the NAMPT-NAD(+)-SIRT axis in chondrocytes by upregulating NAMPT, which stimulated NAD(+) synthesis and thereby activated SIRT family members. The activated NAMPT-SIRT pathway, in turn, promoted HIF-2α protein stability by negatively regulating its hydroxylation and 26S proteasome-mediated degradation, resulting in increased HIF-2α transcriptional activity. Among SIRT family members (SIRT1-7), SIRT2 and SIRT4 were positively associated with HIF-2α stability and transcriptional activity in chondrocytes. This reciprocal regulation was required for the expression of catabolic matrix metalloproteinases (MMP3, MMP12, and MMP13) and OA cartilage destruction caused by IA injection of Ad-Epas1 Ad-Nampt. The reciprocal regulation of HIF-2α and the NAMPT-NAD(+)-SIRT axis in articular chondrocytes is involved in OA cartilage destruction caused by HIF-2α or NAMPT. Copyright © 2015 The Authors. Published by Elsevier Ltd.. All rights reserved.

[Expression of Id1 and Id3 in endometrial carcinoma and their roles in regulating biological behaviors of endometrial carcinoma cells in vitro].

PubMed

Sun, Lili; Li, Xuenong; Liu, Guobing

2013-06-01

To investigate the expression of inhibitor of DNA differentiation/DNA binding 1 (Id1) and Id3 in endometrial carcinoma and explore their roles in regulating the proliferation, invasion, migration and adhesion of endometrial carcinoma cells in vitro. Id1 and Id3 expression in 4 fresh endometrial cancer tissue specimens and matched adjacent tissues were detected using Western blotting. Two endometrial cancer cell lines, HEC-1-B and RL-952, were both divided into 4 groups, namely the untreated group, blank virus group, promoter group and Id1/Id3 double-knockdown group, and their expressions of MMP2, CXCR4 and P21 were detected by qRT-PCR and Western blotting. The proliferation, invasion, migration and adhesion of the cells were evaluated with MTT, Transwell, wound-healing, and adhesion assays. Endometrial carcinoma tissues showed significantly higher Id1 and Id3 expression than the adjacent tissues (P<0.05). In the two endometrial carcinoma cell lines, Id1/Id3 double-knockdown significantly decreased MMP2 and CXCR4 expression and increased P21 expression at both mRNA and protein levels (P<0.05), and resulted in suppressed cell proliferation, invasion, migration and adhesion. Id1 and Id3 expressions are up-regulated in endometrial carcinoma to promote the proliferation, invasion, migration and adhesion of the tumor cells by increasing MMP2 and CXCR4 expression and reducing P21 expression. Therapies targeting Id1/Id3 can be a novel strategy for treatment of endometrial carcinoma.

Oleic acid-induced ANGPTL4 enhances head and neck squamous cell carcinoma anoikis resistance and metastasis via up-regulation of fibronectin.

PubMed

Shen, Chih-Jie; Chan, Shih-Hung; Lee, Chung-Ta; Huang, Wan-Chen; Tsai, Jhih-Peng; Chen, Ben-Kuen

2017-02-01

Obese patients have higher levels of free fatty acids (FFAs) in their plasma and a higher risk of cancer than their non-obese counterparts. However, the mechanisms involved in the regulation of cancer metastasis by FFAs remain unclear. In this study, we found that oleic acid (OA) induced angiopoietin-like 4 (ANGPTL4) protein expression and secretion and conferred anoikis resistance to head and neck squamous cell carcinomas (HNSCCs). The autocrine production of OA-induced ANGPTL4 further promoted HNSCC migration and invasion. In addition, the expression of peroxisome proliferator-activated receptor (PPAR) was essential for the OA-induced ANGPTL4 expression and invasion. The levels of OA-induced epithelial-mesenchymal transition markers, such as vimentin, MMP-9, and fibronectin and its downstream effectors Rac1/Cdc42, were significantly reduced in ANGPTL4-depleted cells. Knocking down fibronectin inhibited the expression of MMP-9 and repressed OA- and recombinant ANGPTL4-induced HNSCC invasion. On the other hand, ANGPTL4 siRNA inhibited OA-induced MMP-9 expression, which was reversed in fibronectin-overexpressing cells. Furthermore, the depletion of ANGPTL4 impeded the OA-primed metastatic seeding of tumor cells in the lungs. These results demonstrate that OA enhances HNSCC metastasis through the ANGPTL4/fibronectin/Rac1/Cdc42 and ANGPTL4/fibronectin/MMP-9 signaling axes. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

Garlic Supplementation Ameliorates UV-Induced Photoaging in Hairless Mice by Regulating Antioxidative Activity and MMPs Expression.

PubMed

Kim, Hye Kyung

2016-01-08

UV exposure is associated with oxidative stress and is the primary factor in skin photoaging. UV-induced reactive oxygen species (ROS) cause the up-regulation of metalloproteinase (MMPs) and the degradation of dermal collagen and elastic fibers. Garlic and its components have been reported to exert antioxidative effects. The present study investigated the protective effect of garlic on UV-induced photoaging and MMPs regulation in hairless mice. Garlic was supplemented in the diet, and Skh-1 hairless mice were exposed to UV irradiation five days/week for eight weeks. Mice were divided into four groups; Non-UV, UV-irradiated control, UV+1% garlic powder diet group, and UV+2% garlic powder diet group. Chronic UV irradiation induced rough wrinkling of the skin with hyperkeratosis, and administration of garlic diminished the coarse wrinkle formation. UV-induced dorsal skin and epidermal thickness were also ameliorated by garlic supplementation. ROS generation, skin and serum malondialdehyde levels were significantly increased by UV exposure and were ameliorated by garlic administration although the effects were not dose-dependent. Antioxidant enzymes such as superoxide dismutase and catalase activities in skin tissues were markedly reduced by UV irradiation and garlic treatment increased these enzyme activities. UV-induced MMP-1 and MMP-2 protein levels were suppressed by garlic administration. Furthermore, garlic supplementation prevented the UV-induced increase of MMP-1 mRNA expression and the UV-induced decrease of procollagen mRNA expression. These results suggest that garlic may be effective for preventing skin photoaging accelerated by UV irradiation through the antioxidative system and MMP regulation.

Dietary Zinc Modulates Matrix Metalloproteinases in Traumatic Brain Injury.

PubMed

Scrimgeour, Angus; Carrigan, Christopher; Condlin, Michelle Lynn; Urso, Maria L; van den Berg, Roland M; van Helden, Herman P M; Montain, Scott J; Joosen, Marloes J A

2018-05-18

Animal models of mild traumatic brain injury (mTBI) provide opportunity to examine the extent to which dietary interventions can be used to improve recovery after injury. Animal studies also suggest that matrix metalloproteinases (MMPs) play a role in tissue remodeling post-TBI. Because dietary zinc (Zn) improved recovery in non-blast mTBI models, and the MMPs are Zn-requiring enzymes, we evaluated the effects of low- and adequate Zn diets on MMP expression and behavioral responses, following exposure to a single blast. MMP mRNA expression in soleus muscle and frontal cortex tissues were quantified at 48h and 14d post-blast. In muscle, blast resulted in significant upregulation of MT-MMP, MMP-2, TIMP-1 and TIMP-2 at 48h post-injury in rats consuming adequate Zn diets (AdZn). At 14d post-blast, there were no blast- or dietary-effects observed on MMP levels in muscle, supporting the existence of a Zn-responsive, functional repair and remodeling mechanism. In contrast, blast resulted in a significant down-regulation of MT-MMP, TIMP-1 and TIMP-2, and a significant up-regulation of MMP-3 levels at 48h post-injury in cortex tissue; while at 14d post-blast, MT-MMP, MMP-2 and TIMP-2 were all down-regulated in response to blast, independent of diet, and TIMP-1 were significantly increased in rats fed AdZn diets despite the absence of elevated MMPs. Because the blast injuries occurred while animals were under general anesthesia, the increased immobility observed post-injury in rats consuming LoZn diets, suggest that blast mTBI can, in the absence of any psychological stressor, induce PTSD-related traits that are chronic, but responsive to diet. Taken together, our results support a relationship between marginally Zn-deficient status and a compromised regenerative response post-injury in muscle, likely through the MMP pathway. However, in neuronal tissue changes in MMP/TIMP levels following blast indicate a variable response to marginally Zn-deficient diets that may help explain compromised repair mechanism(s) previously associated with the systemic hypozincemia that develops in patients with TBI.

miR-125b inhibits keratinocyte proliferation and promotes keratinocyte apoptosis in oral lichen planus by targeting MMP-2 expression through PI3K/Akt/mTOR pathway.

PubMed

Wang, Jing; Luo, Hong; Xiao, Yan; Wang, Luyao

2016-05-01

Oral lichen planus (OLP) is a chronic inflammatory mucosal disease that involves the degeneration of keratinocytes. However, the etiology and mechanisms of OLP pathogenesis have not been fully elucidated. In this study, we used keratinocytes HaCaT stimulated with lipopolysaccharide (LPS) to mimic a local OLP immune environment, and investigated the regulatory role of miR-125b in keratinocyte proliferation and apoptosis under OLP conditions. Immunohistochemical analysis and quantitative real-time PCR (qRT-PCR) assay showed that MMP-2 expression was up-regulated and miR-125b expression was down-regulated in both OLP mucosa tissues and LPS-incubated HaCaT cells. Western blot analysis indicated that miR-125b overexpression suppressed LPS-induced MMP-2 expression in HaCaT cells. Molecularly, our results confirmed that MMP-2 is a target gene of miR-125b in HaCaT cells. The effect of miR-125b on cell proliferation was revealed by CCK-8 assay, BrdU assay and cell cycle analysis, which illustrated that miR-125b overexpression impeded LPS-induced HaCaT cell proliferation. Flow cytometry analysis further demonstrated that miR-125b overexpression promoted HaCaT cell apoptosis. Moreover, these effects were involved in PI3K/Akt/mTOR activation, as miR-125b overexpression inhibited LPS-enhanced expression of p-Akt and p-mTOR proteins. Taken together, these data confirm that miR-125b might inhibit keratinocyte proliferation and promote keratinocyte apoptosis in OLP pathogenesis by targeting MMP-2 through PI3K/Akt/mTOR pathway. Copyright © 2016 Elsevier Masson SAS. All rights reserved.

Relationship among LRP1 expression, Pyk2 phosphorylation and MMP-9 activation in left ventricular remodelling after myocardial infarction.

PubMed

Revuelta-López, Elena; Soler-Botija, Carol; Nasarre, Laura; Benitez-Amaro, Aleyda; de Gonzalo-Calvo, David; Bayes-Genis, Antoni; Llorente-Cortés, Vicenta

2017-09-01

Left ventricular (LV) remodelling after myocardial infarction (MI) is a crucial determinant of the clinical course of heart failure. Matrix metalloproteinase (MMP) activation is strongly associated with LV remodelling after MI. Elucidation of plasma membrane receptors related to the activation of specific MMPs is fundamental for treating adverse cardiac remodelling after MI. The aim of current investigation was to explore the potential association between the low-density lipoprotein receptor-related protein 1 (LRP1) and MMP-9 and MMP-2 spatiotemporal expression after MI. Real-time PCR and Western blot analyses showed that LRP1 mRNA and protein expression levels, respectively, were significantly increased in peri-infarct and infarct zones at 10 and 21 days after MI. Confocal microscopy demonstrated high colocalization between LRP1 and the fibroblast marker vimentin, indicating that LRP1 is mostly expressed by cardiac fibroblasts in peri-infarct and infarct areas. LRP1 also colocalized with proline-rich tyrosine kinase 2 (pPyk2) and MMP-9 in cardiac fibroblasts in ischaemic areas at 10 and 21 days after MI. Cell culture experiments revealed that hypoxia increases LRP1, pPyk2 protein levels and MMP-9 activity in fibroblasts, without significant changes in MMP-2 activity. MMP-9 activation by hypoxia requires LRP1 and Pyk2 phosphorylation in fibroblasts. Collectively, our in vivo and in vitro data support a major role of cardiac fibroblast LRP1 levels on MMP-9 up-regulation associated with ventricular remodelling after MI. © 2017 The Authors. Journal of Cellular and Molecular Medicine published by John Wiley & Sons Ltd and Foundation for Cellular and Molecular Medicine.

The protective effect of 18β-Glycyrrhetinic acid against UV irradiation induced photoaging in mice.

PubMed

Kong, Song-Zhi; Chen, Hai-Ming; Yu, Xiu-Ting; Zhang, Xie; Feng, Xue-Xuan; Kang, Xin-Huang; Li, Wen-Jie; Huang, Na; Luo, Hui; Su, Zi-Ren

2015-01-01

It has been confirmed that repeated exposure of skin to ultraviolet (UV) radiation results in cutaneous oxidative stress and inflammation, which act in concert to cause premature skin aging, well known as photoaging. 18β-Glycyrrhetinic acid (GA), widely used to treat various tissue inflammations, is the main active component of licorice root, and has also been shown to possess favorable anti-oxidative property and modulating immunity function. In the present study, we investigated the potential protective effect of GA on UV-induced skin photoaging in a mouse model. During the experimental period of ten consecutive weeks, the dorsal depilated skin of mice was treated with topical GA for 2 hours prior to UV irradiation. The results showed that GA pretreatment significantly alleviated the macroscopic and histopathological damages in mice skin caused by UV. Meanwhile, the data also indicated that GA markedly up-regulated the activities of the antioxidant enzymes (SOD, GSH-Px), and increased the content of skin collagen, while obviously decreased malonaldehyde level and inhibited high expressions of matrix metalloproteinase-1 (MMP-1) and -3 (MMP-3), as well as down-regulated the expression of inflammatory cytokines such as IL-6, TNF-α and IL-10. Taken together, these findings amply demonstrate that GA observably attenuates UV-induced skin photoaging mainly by virtue of its antioxidative and anti-inflammatory properties, as well as regulating the abnormal expression of MMP-1 and MMP-3. Copyright © 2014 Elsevier Inc. All rights reserved.

A Transcriptional Regulatory Role for the Membrane Type-1 Matrix Metalloproteinase in Carcinogen-Induced Inflammasome Gene Expression.

PubMed

Sheehy, Samuel; Annabi, Borhane

2017-01-01

Signal-transducing functions driven by the cytoplasmic domain of membrane type-1 matrix metalloproteinase (MT1-MMP) are believed to regulate many inflammation-associated cancer cell functions including migration, proliferation, and survival. Aside from upregulation of the inflammation biomarker cyclooxygenase-2 (COX-2) expression, MT1-MMP's role in relaying intracellular signals triggered by extracellular pro-inflammatory cues remains poorly understood. Here, we triggered inflammation in HT1080 fibrosarcoma cells with phorbol-12-myristate-13-acetate (PMA), an inducer of COX-2 and of MT1-MMP. To assess the global transcriptional regulatory role that MT1-MMP may exert on inflammation biomarkers, we combined gene array screens with a transient MT1-MMP gene silencing strategy. Expression of MT1-MMP was found to exert both stimulatory and repressive transcriptional control of several inflammasome-related biomarkers such as interleukin (IL)-1B, IL-6, IL-12A, and IL-33, as well as of transcription factors such as EGR1, ELK1, and ETS1/2 in PMA-treated cells. Among the signal-transducing pathways explored, the silencing of MT1-MMP prevented PMA from phosphorylating extracellular signal-regulated kinase, inhibitor of κB, and p105 nuclear factor κB (NF-κB) intermediates. We also found a signaling axis linking MT1-MMP to MMP-9 transcriptional regulation. Altogether, our data indicate a significant involvement of MT1-MMP in the transcriptional regulation of inflammatory biomarkers consolidating its contribution to signal transduction functions in addition to its classical hydrolytic activity.

Mixomics analysis of breast cancer: Long non-coding RNA linc01561 acts as ceRNA involved in the progression of breast cancer.

PubMed

Jiang, Rui; Zhao, Chunming; Gao, Binbin; Xu, Jiawen; Song, Wei; Shi, Peng

2018-06-08

This study aimed at finding the long non-coding RNA (lncRNA), miRNA and mRNA which played critical roles in breast cancer (BrCa) by using mixOmics R package. The BrCa dataset were obtained from TCGA and then analyzed using "DESeq2" R package. Multivariate analyses were performed with the "mixOmics" R package and the first component of the stacked partial least-Squares discriminant analysis results were used for searching the interested lncRNA, miRNA and mRNA. qRT-PCR was applied to identify the bioinformatics results in four BrCa cell lines (MCF7, BT-20, ZR-75-1, and MX-1) and the breast epithelial cell line MCF-10 A. Then cells (MCF-1 and MX-1) were transfected with si-linc01561, miR-145-5p mimics and si-MMP11 to further investigate the effects of linc01561, miR-145-5p and MMP11 on the BrCa cells proliferation and apoptosis. MixOmics results showed that linc01561, miR-145-5p and MMP11 might play important roles in BrCa. qRT-PCR results identified that in BrCa cell lines, linc01561 and MMP11 were higher expressed while miR-145-5p was lower expressed compared with those in epithelial cell line. The linc01561 inhibition elevated miR-145-5p expression and then suppressed MMP11 expression. Moreover, linc01561 inhibition suppressed the BrCa cells proliferation and promoted the apoptosis, which was realized by up-regulating expression of miR-145-5p and down-regulating expression of MMP11. In summary, the findings of this study, based on ceRNA theory, combining the research foundation of miR-145-5p and MMP11, and taking linc01561 as a new study point, provide new insight into molecular-level reversing proliferation and apoptosis of BrCa. Copyright © 2018 Elsevier Ltd. All rights reserved.

Matrix metalloproteinases-2 and -9 in Campylobacter jejuni-induced paralytic neuropathy resembling Guillain-Barré syndrome in chickens.

PubMed

Nyati, Kishan Kumar; Prasad, Kashi Nath; Agrawal, Vinita; Husain, Nuzhat

2017-10-01

Inflammation in Guillain-Barré syndrome (GBS) is manifested by changes in matrix metalloproteinase (MMP) and pro-inflammatory cytokine expression. We investigated the expression of MMP-2, -9 and TNF-α and correlated it with pathological changes in sciatic nerve tissue from Campylobacter jejuni-induced chicken model for GBS. Campylobacter jejuni and placebo were fed to chickens and assessed for disease symptoms. Sciatic nerves were examined by histopathology and immunohistochemistry. Expressions of MMPs and TNF-α, were determined by real-time PCR, and activities of MMPs by zymography. Diarrhea developed in 73.3% chickens after infection and 60.0% of them developed GBS like neuropathy. Pathology in sciatic nerves showed perinodal and/or patchy demyelination, perivascular focal lymphocytic infiltration and myelin swelling on 10th- 20th post infection day (PID). MMP-2, -9 and TNF-α were up-regulated in progressive phase of the disease. Enhanced MMP-2, -9 and TNF-α production in progressive phase correlated with sciatic nerve pathology in C. jejuni-induced GBS chicken model. Copyright © 2017 Elsevier Ltd. All rights reserved.

Localization of matrix metalloproteinases, (MMPs) their tissue inhibitors, and vascular endothelial growth factor (VEGF) in growth plates of children and adolescents indicates a role for MMPs in human postnatal growth and skeletal maturation.

PubMed

Haeusler, G; Walter, I; Helmreich, M; Egerbacher, M

2005-05-01

Numerous studies have focused on the expression, regulation, and biological significance of matrix metalloproteinases (MMPs) in the growth plate. Findings in mouse knockout models and in vitro data from various species indicate that MMPs not only degrade extracellular matrix components but may regulate the activity of local growth factors. In this study we investigated the presence, distribution, and activity of various MMPs and inhibitors, tissue transglutaminase (tTG or TG2) and vascular endothelial growth factor (VEGF) in the human child and adolescent growth plates by means of immunohistochemistry and gelatin zymography. Tissue was derived during orthopedic surgery (epiphysiodesis) in two prepubertal and four pubertal patients.MMP-2 and MMP-14 were present in reserve cell chondrocytes. MMP-14 was the most prominent MMP within all zones of the growth plate including proliferating chondrocytes. MMP-1 and MMP-13 (collagenases 1 and 3), MMP-9 (gelatinases B), MMP-10, and MMP-11 (stromelysins) and VEGF were positive in hypertrophic chondrocytes and osteoblasts. MMP-2 showed the same expression pattern but was negative in osteoblasts. Osteoclasts stained positive for MMP-9, MMP-2, and TG2. Tissue inhibitor of MMP (TIMP)-1 was present in all zones of the growth plate, osteoblasts, and osteoclasts; TIMP-2 was found in hypertrophic chondrocytes and osteoblasts. In summary, the presence of MMPs, TIMPs, TG2, and VEGF in our study indicated that the MMPs are relevant in growth plate physiology during the postnatal period in humans. The specific location of MMP expression within the growth plate may be the basis for further studies on the role of MMPs in the local regulation of chondrocyte differentiation, proliferation, and ossification at the chondroosseus junction.

Regulation of matrix metalloproteinase-9 expression between gingival fibroblast cells from old and young rats

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kim, Su-Jung; Chung, Yong-Koo; Chung, Tae-Wook

2009-01-09

Gingival fibroblast cells (rGF) from aged rats have an age-related decline in proliferative capacity compared with young rats. We investigated G1 phase cell cycle regulation and MMP-9 expression in both young and aged rGF. G1 cell cycle protein levels and activity were significantly reduced in response to interleukin-1{beta} (IL-1{beta}) stimulation with increasing in vitro age. Tumor necrosis factor-{alpha} (TNF-{alpha})-induced matrix metalloproteinase-9 (MMP-9) expression was also decreased in aged rGF in comparison with young rGF. Mutational analysis and gel shift assays demonstrated that the lower MMP-9 expression in aged rGF is associated with lower activities of transcription factors NF-{kappa}B and AP-1.more » These results suggest that cell cycle dysregulation and down-regulation of MMP-9 expression in rGF may play a role in gingival remodeling during in vitro aging.« less

Tumour-associated neutrophils and loss of epithelial PTEN can promote corticosteroid-insensitive MMP-9 expression in the chronically inflamed lung microenvironment

PubMed Central

Vannitamby, Amanda; Seow, Huei Jiunn; Anderson, Gary; Vlahos, Ross; Thompson, Michelle; Steinfort, Daniel; Irving, Louis B; Bozinovski, Steven

2017-01-01

Matrix metalloproteinase-9 (MMP-9) is increased in a number of pathological lung conditions, where the proteinase contributes to deleterious remodelling of the airways. While both lung cancer and COPD are associated with increased MMP-9 expression, the cellular and molecular drivers of MMP-9 remain unresolved. In this study, MMP-9 transcript measured within the tumour region from patients with non-small-cell lung cancer (NSCLC) and coexisting COPD was found to be uniformly increased relative to adjacent tumour-free tissue. MMP-9 gene expression and immunohistochemistry identified tumour-associated neutrophils, but not macrophages, as a predominant source of this proteinase. In addition, PTEN gene expression was significantly reduced in tumour and there was evidence of epithelial MMP-9 expression. To explore whether PTEN can regulate epithelial MMP-9 expression, a small interfering (si)RNA knockdown strategy was used in Beas-2B bronchial epithelial cells. PTEN knockdown by siRNA selectively increased MMP-9 expression in response to lipopolysaccharide in a corticosteroid-insensitive manner. In summary, tumour-associated neutrophils represent an important source of MMP-9 in NSCLC, and loss of epithelial PTEN may further augment steroid-insensitive expression. PMID:28202627

Schwann Cell Migration Induced by Earthworm Extract via Activation of PAs and MMP2/9 Mediated through ERK1/2 and p38

PubMed Central

Chang, Yung-Ming; Shih, Ying-Ting; Chen, Yueh-Sheng; Liu, Chien-Liang; Fang, Wen-Kuei; Tsai, Chang-Hai; Tsai, Fuu-Jen; Kuo, Wei-Wen; Lai, Tung-Yuan; Huang, Chih-Yang

2011-01-01

The earthworm, which has stasis removal and wound-healing functions, is a widely used Chinese herbal medicine in China. Schwann cell migration is critical for the regeneration of injured nerves. Schwann cells provide an essentially supportive activity for neuron regeneration. However, the molecular migration mechanisms induced by earthworms in Schwann cells remain unclear. Here, we investigate the roles of MAPK (ERK1/2, JNK and p38) pathways for earthworm-induced matrix-degrading proteolytic enzyme (PAs and MMP2/9) production in Schwann cells. Moreover, earthworm induced phosphorylation of ERK1/2 and p38, but not JNK, activate the downstream signaling expression of PAs and MMPs in a time-dependent manner. Earthworm-stimulated ERK1/2 and p38 phosphorylation was attenuated by pretreatment with U0126 and SB203580, resulting in migration and uPA-related signal pathway inhibition. The results were confirmed using small interfering ERK1/2 and p38 RNA. These results demonstrated that earthworms can stimulate Schwann cell migration and up-regulate PAs and MMP2/9 expression mediated through the MAPK pathways, ERK1/2 and p38. Taken together, our data suggests the MAPKs (ERK1/2, p38)-, PAs (uPA, tPA)-, MMP (MMP2, MMP9) signaling pathway of Schwann cells regulated by earthworms might play a major role in Schwann cell migration and nerve regeneration. PMID:19808845

MMP1 bimodal expression and differential response to inflammatory mediators is linked to promoter polymorphisms

PubMed Central

2011-01-01

Background Identifying the functional importance of the millions of single nucleotide polymorphisms (SNPs) in the human genome is a difficult challenge. Therefore, a reverse strategy, which identifies functionally important SNPs by virtue of the bimodal abundance across the human population of the SNP-related mRNAs will be useful. Those mRNA transcripts that are expressed at two distinct abundances in proportion to SNP allele frequency may warrant further study. Matrix metalloproteinase 1 (MMP1) is important in both normal development and in numerous pathologies. Although much research has been conducted to investigate the expression of MMP1 in many different cell types and conditions, the regulation of its expression is still not fully understood. Results In this study, we used a novel but straightforward method based on agglomerative hierarchical clustering to identify bimodally expressed transcripts in human umbilical vein endothelial cell (HUVEC) microarray data from 15 individuals. We found that MMP1 mRNA abundance was bimodally distributed in un-treated HUVECs and showed a bimodal response to inflammatory mediator treatment. RT-PCR and MMP1 activity assays confirmed the bimodal regulation and DNA sequencing of 69 individuals identified an MMP1 gene promoter polymorphism that segregated precisely with the MMP1 bimodal expression. Chromatin immunoprecipation (ChIP) experiments indicated that the transcription factors (TFs) ETS1, ETS2 and GATA3, bind to the MMP1 promoter in the region of this polymorphism and may contribute to the bimodal expression. Conclusions We describe a simple method to identify putative bimodally expressed RNAs from transcriptome data that is effective yet easy for non-statisticans to understand and use. This method identified bimodal endothelial cell expression of MMP1, which appears to be biologically significant with implications for inflammatory disease. (271 Words) PMID:21244711

The role of annexin A1 in expression of matrix metalloproteinase-9 and invasion of breast cancer cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kang, Hyereen; Ko, Jesang; Jang, Sung-Wuk, E-mail: swjang@amc.seoul.kr

2012-06-22

Highlights: Black-Right-Pointing-Pointer We evaluated the effect of ANXA1 on promoting migration and invasion in MDA-MB-231 cells. Black-Right-Pointing-Pointer ANXA1 siRNA inhibits invasion and migration. Black-Right-Pointing-Pointer ANXA1 regulates MMP-9 expression and activity. Black-Right-Pointing-Pointer ANX-1 siRNA inhibits the activation of NF-{kappa}B in MDA-MB-231 cells. -- Abstract: Matrix metalloproteinase-9 (MMP-9) plays an important role in the invasion and metastasis of cancer cells. However, the regulatory mechanism of MMP-9 expression and its biological effects on breast cancer development remain obscure. In the current study, we examined the potential role of annexin A1 (ANXA1) in regulating migration and invasion in breast cancer cell lines. Both ANXA1more » mRNA and protein are expressed in the highly invasive, hormone-insensitive human breast cancer cell lines MDA-MB-231 and SKBr3, but not in the hormone-responsive cell lines MCF-7 and T47D. Downregulation of ANXA1 expression with specific small interfering RNAs (ANXA1 siRNA) in MDA-MB-231 cells resulted in decreased cancer cell migration and invasion. Ablation of ANXA1 expression decreases the expression of MMP-9 at both the mRNA and protein levels and also reduces the proteolytic activity of MMP-9 in MDA-MB-231 cells. Moreover, silencing ANXA1 also decreases the transcriptional activity of MMP-9 by the suppression of nuclear factor kappa-B (NF-{kappa}B) activity. Collectively, these results indicate that ANXA1 functions as a positive regulator of MMP-9 expression and invasion of breast cancer cells through specific activation of the NF-{kappa}B signaling pathway.« less

The role of extracellular matrix metalloproteinase inducer (EMMPRIN) in the regulation of bovine endometrial cell functions.

PubMed

Mishra, Birendra; Kizaki, Keiichiro; Sato, Takashi; Ito, Akira; Hashizume, Kazuyoshi

2012-06-01

Extracellular matrix metalloproteinase inducer (EMMPRIN) is a cell surface glycoprotein that stimulates the production of several matrix metalloproteinases (MMPs) for tissue remodeling. Previously, we detected EMMPRIN in the bovine endometrium, and it is mainly expressed in the luminal and glandular epithelium whereas MMPs are expressed in the underlying stroma. From this expression pattern, we hypothesized that EMMPRIN may regulate stromal MMPs in endometrial cell functions. To test this hypothesis, a coculture of epithelial and stromal cells was performed using a transwell system. In the coculture, epithelial cells were cultured on the insert membrane and stromal cell on the surface of well plates. Expression of stromal MMP-2 and MMP-14 was significantly higher in coculture with epithelial cell. Further, with the addition of anti-EMMPRIN antibody into the epithelial cell compartment, the expression of stromal EMMPRIN and MMP-2 and MMP-14 was decreased. To identify the active site of EMMPRIN for the augmentation of MMPs, EMMPRIN synthetic peptides that correspond to the extracellular loop domain-I (EM1, EM2, EM3, and EM4) were added into the epithelial cell compartment, and only EM2 at a higher dose interfered with EMMPRIN-mediated expression of MMP-14. Next, we examined the effects of progesterone and/or estrogen on the expression of EMMPRIN, MMP-2, and MMP-14. Progesterone (300 nM) significantly stimulated the expression of EMMPRIN but had no effects on any of the MMPs. These results suggest that EMMPRIN derived from epithelial cells regulates MMPs in the endometrium under progesterone-rich conditions and may thereby modulate bovine endometrial cell functions during gestation.

Increased expression of matrix metalloproteinases mediates thromboxane A2-induced invasion in lung cancer cells.

PubMed

Li, Xiuling; Tai, Hsin-Hsiung

2012-07-01

Thromboxane A(2) receptor (TP) has been shown to play an important role in multiple aspects of cancer development including regulation of tumor growth, survival and metastasis. Here we report that TP mediates cancer cell invasion by inducing expression of matrix metalloproteinases (MMPs). TP agonist, I-BOP, significantly elevated MMP-1, MMP-3, MMP-9 and MMP-10 mRNA levels in A549 human lung adenocarcinoma cells overexpressing TPα or TPβ. The secretion of MMP-1 and MMP-9 in conditioned media was determined using Western blot analysis and zymographic assay. Signaling pathways of I-BOP-induced MMP-1 expression were examined in further detail as a model system for MMPs induction. Signaling molecules involved in I-BOP-induced MMP-1 expression were identified by using specific inhibitors including small interfering (si)-RNAs of signaling molecules and promoter reporter assay. The results indicate that I-BOP-induced MMP-1 expression is mediated by protein kinase C (PKC), extracellular signal-regulated kinase (ERK)-activator protein-1(AP-1) and ERK-CCAAT/enhancer-binding protein β (C/EBPβ) pathways. I-BOP-induced cellular invasiveness of A549 cells expressing TPα or TPβ was determined by invasion assay. GM6001, a general inhibitor of MMPs, decreased basal and I-BOP-induced cell invasion. Knockdown of MMP-1 and MMP-9 by their respective siRNA partially reduced I-BOP-stimulated cell invasion suggesting that other MMPs induced by I-BOP were also involved. Our studies establish the relationship between TP and MMPs in cancer cell invasion and suggest that the thromboxane A(2) (TXA(2))-TP signaling is a potential therapeutic target for cancer invasion and metastasis.

NADPH oxidase/ROS-dependent PYK2 activation is involved in TNF-α-induced matrix metalloproteinase-9 expression in rat heart-derived H9c2 cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Yang, Chuen-Mao, E-mail: chuenmao@mail.cgu.edu.tw; Heart Failure Center, Division of Cardiology, Department of Internal Medicine, Chang Gung Memorial Hospital at Keelung, Keelung, Taiwan; Lee, I-Ta

TNF-α plays a mediator role in the pathogenesis of chronic heart failure contributing to cardiac remodeling and peripheral vascular disturbances. The implication of TNF-α in inflammatory responses has been shown to be mediated through up-regulation of matrix metalloproteinase-9 (MMP-9). However, the detailed mechanisms of TNF-α-induced MMP-9 expression in rat embryonic-heart derived H9c2 cells are largely not defined. We demonstrated that in H9c2 cells, TNF-α induced MMP-9 mRNA and protein expression associated with an increase in the secretion of pro-MMP-9. TNF-α-mediated responses were attenuated by pretreatment with the inhibitor of ROS (N-acetyl-L-cysteine, NAC), NADPH oxidase [apocynin (APO) or diphenyleneiodonium chloride (DPI)],more » MEK1/2 (U0126), p38 MAPK (SB202190), JNK1/2 (SP600125), NF-κB (Bay11-7082), or PYK2 (PF-431396) and transfection with siRNA of TNFR1, p47{sup phox}, p42, p38, JNK1, p65, or PYK2. Moreover, TNF-α markedly induced NADPH oxidase-derived ROS generation in these cells. TNF-α-enhanced p42/p44 MAPK, p38 MAPK, JNK1/2, and NF-κB (p65) phosphorylation and in vivo binding of p65 to the MMP-9 promoter were inhibited by U0126, SB202190, SP600125, NAC, DPI, or APO. In addition, TNF-α-mediated PYK2 phosphorylation was inhibited by NAC, DPI, or APO. PYK2 inhibition could reduce TNF-α-stimulated MAPKs and NF-κB activation. Thus, in H9c2 cells, we are the first to show that TNF-α-induced MMP-9 expression is mediated through a TNFR1/NADPH oxidase/ROS/PYK2/MAPKs/NF-κB cascade. We demonstrated that NADPH oxidase-derived ROS generation is involved in TNF-α-induced PYK2 activation in these cells. Understanding the regulation of MMP-9 expression and NADPH oxidase activation by TNF-α on H9c2 cells may provide potential therapeutic targets of chronic heart failure. - Highlights: • TNF-α induces MMP-9 secretion and expression via a TNFR1-dependent pathway. • TNF-α induces ROS/PYK2-dependent MMP-9 expression in H9c2 cells. • TNF-α induces MMP-9 expression via a NADPH oxidase/ROS-dependent NF-κB signaling. • TNF-α activates MAPK phosphorylation through NADPH oxidase/ROS generation.« less

Up-Regulation and Profibrotic Role of Osteopontin in Human Idiopathic Pulmonary Fibrosis

PubMed Central

Pardo, Annie; Gibson, Kevin; Cisneros, José; Richards, Thomas J; Yang, Yinke; Becerril, Carina; Yousem, Samueal; Herrera, Iliana; Ruiz, Victor; Selman, Moisés; Kaminski, Naftali

2005-01-01

Background Idiopathic pulmonary fibrosis (IPF) is a progressive and lethal disorder characterized by fibroproliferation and excessive accumulation of extracellular matrix in the lung. Methods and Findings Using oligonucleotide arrays, we identified osteopontin as one of the genes that significantly distinguishes IPF from normal lungs. Osteopontin was localized to alveolar epithelial cells in IPF lungs and was also significantly elevated in bronchoalveolar lavage from IPF patients. To study the fibrosis-relevant effects of osteopontin we stimulated primary human lung fibroblasts and alveolar epithelial cells (A549) with recombinant osteopontin. Osteopontin induced a significant increase of migration and proliferation in both fibroblasts and epithelial cells. Epithelial growth was inhibited by the pentapeptide Gly-Arg-Gly-Asp-Ser (GRGDS) and antibody to CD44, while fibroproliferation was inhibited by GRGDS and antibody to αvβ3 integrin. Fibroblast and epithelial cell migration were inhibited by GRGDS, anti-CD44, and anti-αvβ3. In fibroblasts, osteopontin up-regulated tissue inhibitor of metalloprotease-1 and type I collagen, and down-regulated matrix metalloprotease-1 (MMP-1) expression, while in A549 cells it caused up-regulation of MMP-7. In human IPF lungs, osteopontin colocalized with MMP-7 in alveolar epithelial cells, and application of weakest link statistical models to microarray data suggested a significant interaction between osteopontin and MMP-7. Conclusions Our results provide a potential mechanism by which osteopontin secreted from the alveolar epithelium may exert a profibrotic effect in IPF lungs and highlight osteopontin as a potential target for therapeutic intervention in this incurable disease. PMID:16128620

[Tissue collagenase MMP-14 and endogenous regulators of its activity in the corpus uteri in squamous cell carcinoma of the cervix].

PubMed

Timoshenko, O S; Gureeva, T A; Kugaevskaya, E V; Zavalishina, L E; Andreeva, Yu Yu; Solovyeva, N I

to investigate the expression of the membrane-bound matrix metalloproteinase MT1-MMP (MMP-14), its tissue inhibitor TIMP-2, and the proMMP-14 activator furin in the corpus uteri from the vaginal wall to the bottom of the uterine cavity in squamous cell carcinoma of the cervix (SCCC). Hysterectomy material was examined in patients with SCCC. Reverse transcriptase polymerase chain reaction (RT-PCR), immunohistochemistry (IHC), and enzyme assays were used. In SCCC, higher levels of MMP-14 expression were established in tumor cells, as evidenced by IHC (+3) and RT-PCR. IHC showed that the expression of MMP-14 was absent or insignificant in the normal uterine endometrial and myometrial tissues. However, that of MMP-14 mRNA was also found in the normal tissues to the bottom of the uterine cavity. Furin activity in the tumor was much higher than that in normal tissues. IHC indicated that TIMP-2 expression was low or absent in both the tumor and normal tissues. The expression of TIMP-2 mRNA was sufficiently obvious in both the tumor and normal tissues to the bottom of the uterine cavity. In SCCC, MMP-14 expression was substantially increased in tumors. The expression of MMP-14 and regulators of its activity is aimed at enhancing the tumor destructive (invasive) potential in the pericellular space and can occur (be induced) in the morphologically normal uterine tissue apparently with involvement of signaling through the epithelial-mesenchymal interaction. Data are important for understanding the role of MMP-14 in the development of a multistage process of carcinogenesis and may have prognostic value and an impact on therapeutic strategy for the patient.

Extracellular matrix remodeling and matrix metalloproteinases (ajMMP-2 like and ajMMP-16 like) characterization during intestine regeneration of sea cucumber Apostichopus japonicus.

PubMed

Miao, Ting; Wan, Zixuan; Sun, Lina; Li, Xiaoni; Xing, Lili; Bai, Yucen; Wang, Fang; Yang, Hongsheng

2017-10-01

Remodeling of extracellular matrix (ECM) regulated by matrix metalloproteinases (MMPs) is essential for tissue regeneration. In the present study, we used immunohistochemistry (IHC) techniques against ECM components to reveal changes of ECM during intestine regeneration of Apostichopus japonicus. The expression of collagen I and laminin reduced apparently from the eviscerated intestine, while fibronectin exhibited continuous expression in all regeneration stages observed. Meanwhile, we cloned two MMP genes from A. japonicus by RACE PCR. The full-length cDNA of ajMMP-2 like is 2733bp and contains a predicted open reading frame (ORF) of 1716bp encoding 572 amino acids. The full-length cDNA of ajMMP-16 like is 2705bp and contains an ORF of 1452bp encoding 484 amino acids. The predicted protein sequences of each MMP contain two conserved domains, ZnMc_MMP and HX. Homology and phylogenetic analysis revealed that ajMMP-2 like and ajMMP-16 like share high sequence similarity with MMP-2 and MMP-16 from Strongylocentrotus purpuratus, respectively. Then we investigated spatio-temporal expression of ajMMP-2 like and ajMMP-16 like during different regeneration stages by qRT-PCR and IHC. The expression pattern of them showed a roughly opposite trend from that of ECM components. According to our results, a fibronectin-dominate temporary matrix is created in intestine regeneration, and it might provide structural integrity for matrix and promote cell movement. We also hypothesize that ajMMP-2 like and ajMMP-16 like could accelerate cell migration and regulate interaction between ECM components and growth factors. This work provides new evidence of ECM and MMPs involvement in sea cucumber regeneration. Copyright © 2017 Elsevier Inc. All rights reserved.

Dexamethasone ameliorates H₂S-induced acute lung injury by alleviating matrix metalloproteinase-2 and -9 expression.

PubMed

Wang, Jun; Zhang, Huazhong; Su, Chenglei; Chen, Junjie; Zhu, Baoli; Zhang, Hengdong; Xiao, Hang; Zhang, Jinsong

2014-01-01

Acute lung injury (ALI) is one of the fatal outcomes after exposure to high levels of hydrogen sulfide (H2S), and the matrix metalloproteinases (MMPs) especially MMP-2 and MMP-9 are believed to be involved in the development of ALI by degrading the extracellular matrix (ECM) of blood-air barrier. However, the roles of MMP-2 and MMP-9 in H2S-induced ALI and the mechanisms of dexamethasone (DXM) in treating ALI in clinical practice are still largely unknown. The present work was aimed to investigate the roles of MMP-2 and MMP-9 in H2S-induced ALI and the protective effects of DXM. In our study, SD rats were exposed to H2S to establish the ALI model and in parallel, A549 cells were incubated with NaHS (a H2S donor) to establish cell model. The lung HE staining, immunohistochemisty, electron microscope assay and wet/dry ratio were used to identify the ALI induced by H2S, then the MMP-2 and MMP-9 expression in both rats and A549 cells were detected. Our results revealed that MMP-2 and MMP-9 were obviously increased in both mRNA and protein level after H2S exposure, and they could be inhibited by MMP inhibitor doxycycline (DOX) in rat model. Moreover, DXM significantly ameliorated the symptoms of H2S-induced ALI including alveolar edema, infiltration of inflammatory cells and the protein leakage in BAFL via up-regulating glucocorticoid receptor(GR) to mediate the suppression of MMP-2 and MMP-9. Furthermore, the protective effects of DXM in vivo and vitro study could be partially blocked by co-treated with GR antagonist mifepristone (MIF). Our results, taken together, demonstrated that MMP-2 and MMP-9 were involved in the development of H2S-induced ALI and DXM exerted protective effects by alleviating the expression of MMP-2 and MMP-9. Therefore, MMP-2 and MMP-9 might represent novel pharmacological targets for the treatment of H2S and other hazard gases induced ALI.

Nobiletin inhibits human osteosarcoma cells metastasis by blocking ERK and JNK-mediated MMPs expression

PubMed Central

Cheng, Hsin-Lin; Hsieh, Ming-Ju; Yang, Jia-Sin; Lin, Chiao-Wen; Lue, Ko-Haung; Lu, Ko-Hsiu; Yang, Shun-Fa

2016-01-01

Nobiletin, a polymethoxyflavone, has a few pharmacological activities, including anti-inflammation and anti-cancer effects. However, its effect on human osteosarcoma progression remains uninvestigated. Therefore, we examined the effectiveness of nobiletin against cellular metastasis of human osteosarcoma and the underlying mechanisms. Nobiletin, up to 100 μM without cytotoxicity, significantly decreased motility, migration and invasion as well as enzymatic activities, protein levels and mRNA expressions of matrix metalloproteinase (MMP)-2 and MMP-9 in U2OS and HOS cells. In addition to inhibition of extracellular signal-regulated kinase (ERK) and c-Jun N-terminal kinase (JNK), the inhibitory effect of nobiletin on the DNA-binding activity of the transcription factor nuclear factor-kappa B (NF-κB), cAMP response element-binding protein (CREB), and specificity protein 1 (SP-1) in U2OS and HOS cells. Co-treatment with ERK and JNK inhibitors and nobiletin further reduced U2OS cells migration and invasion. These results indicated that nobiletin inhibits human osteosarcoma U2OS and HOS cells motility, migration and invasion by down-regulating MMP-2 and MMP-9 expressions via ERK and JNK pathways and through the inactivation of downstream NF-κB, CREB, and SP-1. Nobiletin has the potential to serve as an anti-metastatic agent for treating osteosarcoma. PMID:27144433

CXCL4-induced plaque macrophages can be specifically identified by co-expression of MMP7+S100A8+ in vitro and in vivo.

PubMed

Erbel, Christian; Tyka, Mirjam; Helmes, Christian M; Akhavanpoor, Mohmmadreza; Rupp, Gregor; Domschke, Gabriele; Linden, Fabian; Wolf, Antonia; Doesch, Andreas; Lasitschka, Felix; Katus, Hugo A; Gleissner, Christian A

2015-04-01

Macrophage heterogeneity in human atherosclerotic plaques has been recognized; however, markers for unequivocal identification of some subtypes are lacking. We found that the platelet chemokine CXCL4 induces a unique macrophage phenotype, which we proposed to call 'M4'. Here, we sought to identify suitable markers that identify M4 macrophages in vitro and in vivo. Using a stringent algorithm, we identified a set of potential markers from transcriptomic data derived from polarized macrophages. We specifically focused on matrix metalloproteinase (MMP)7 and S100A8, the co-expression of which has not been described in any macrophage type thus far. We found dose- and time-dependent MMP7 and S100A8 expression in M4 macrophages at the gene and protein levels. CXCL4-induced up-regulation of both MMP7 and S100A8 was curbed in the presence of heparin, which binds to CXCL4 and glycosaminoglycans, most likely representing the macrophage receptor for CXCL4. Immunofluorescence of post-mortem atherosclerotic coronary arteries identified CD68(+)MMP7(+), CD68(+)MMP7(-), CD68(+)S100A8(+) and CD68(+)S100A8(-) macrophages. A small proportion of MMP7(+)S100A8(+) macrophages most likely represent M4 macrophages. In summary, we have identified co-expression of MMP7 and S100A8 to be a marker combination exclusively found in M4 macrophages. This finding may allow further dissection of the role of M4 macrophages in atherosclerosis and other pathologic conditions. © The Author(s) 2014 Reprints and permissions: sagepub.co.uk/journalsPermissions.nav.

Matrix Metalloproteinase-9–Null Mice Are Resistant to TGF-β–Induced Anterior Subcapsular Cataract Formation

PubMed Central

Korol, Anna; Pino, Giuseppe; Dwivedi, Dhruva; Robertson, Jennifer V.; Deschamps, Paula A.; West-Mays, Judith A.

2015-01-01

Epithelial-mesenchymal transition (EMT) is associated with fibrotic diseases in the lens, such as anterior subcapsular cataract (ASC) formation. Often mediated by transforming growth factor (TGF)-β, EMT in the lens involves the transformation of lens epithelial cells into a multilayering of myofibroblasts, which manifest as plaques beneath the lens capsule. TGF-β–induced EMT and ASC have been associated with the up-regulation of two matrix metalloproteinases (MMPs): MMP-2 and MMP-9. The current study used MMP-2 and MMP-9 knockout (KO) mice to further determine their unique roles in TGF-β–induced ASC formation. Adenoviral injection of active TGF-β1 into the anterior chamber of all wild-type and MMP-2 KO mice led to the formation of distinct ASC plaques that were positive for α-smooth muscle actin, a marker of EMT. In contrast, only a small proportion of the MMP-9 KO eyes injected with adenovirus-expressing TGF-β1 exhibited ASC plaques. Isolated lens epithelial explants from wild-type and MMP-2 KO mice that were treated with TGF-β exhibited features indicative of EMT, whereas those from MMP-9 KO mice did not acquire a mesenchymal phenotype. MMP-9 KO mice were further bred onto a TGF-β1 transgenic mouse line that exhibits severe ASC formation, but shows a resistance to ASC formation in the absence of MMP-9. These findings suggest that MMP-9 expression is more critical than MMP-2 in mediating TGF-β–induced ASC formation. PMID:24814605

Membrane Type 1 Matrix Metalloproteinase Regulates Monocyte Migration and Collagen Destruction in Tuberculosis.

PubMed

Sathyamoorthy, Tarangini; Tezera, Liku B; Walker, Naomi F; Brilha, Sara; Saraiva, Luisa; Mauri, Francesco A; Wilkinson, Robert J; Friedland, Jon S; Elkington, Paul T

2015-08-01

Tuberculosis (TB) remains a global pandemic and drug resistance is rising. Multicellular granuloma formation is the pathological hallmark of Mycobacterium tuberculosis infection. The membrane type 1 matrix metalloproteinase (MT1-MMP or MMP-14) is a collagenase that is key in leukocyte migration and collagen destruction. In patients with TB, induced sputum MT1-MMP mRNA levels were increased 5.1-fold compared with matched controls and correlated positively with extent of lung infiltration on chest radiographs (r = 0.483; p < 0.05). M. tuberculosis infection of primary human monocytes increased MT1-MMP surface expression 31.7-fold and gene expression 24.5-fold. M. tuberculosis-infected monocytes degraded collagen matrix in an MT1-MMP-dependent manner, and MT1-MMP neutralization decreased collagen degradation by 73%. In human TB granulomas, MT1-MMP immunoreactivity was observed in macrophages throughout the granuloma. Monocyte-monocyte networks caused a 17.5-fold increase in MT1-MMP surface expression dependent on p38 MAPK and G protein-coupled receptor-dependent signaling. Monocytes migrating toward agarose beads impregnated with conditioned media from M. tuberculosis-infected monocytes expressed MT1-MMP. Neutralization of MT1-MMP activity decreased this M. tuberculosis network-dependent monocyte migration by 44%. Taken together, we demonstrate that MT1-MMP is central to two key elements of TB pathogenesis, causing collagen degradation and regulating monocyte migration. Copyright © 2015 The Authors.

Assessment of MMP-2/-9 expression by fluorescence endoscopy for evaluation of anastomotic healing in a murine model of anastomotic leakage

PubMed Central

Twardy, Vanessa; Becker, Felix; Geyer, Christiane; Schwegmann, Katrin; Mohr, Annika; Faust, Andreas

2018-01-01

Background Disturbance of intestinal wound closure leads to insufficient anastomotic healing and is associated with considerable morbidity following colorectal resections. Matrix metalloproteinases (MMPs) play a crucial role in regulation of wound closure. Here fluorescence endoscopy was evaluated for assessment of MMP-2/-9 expression during failed intestinal anastomotic healing. Methods Distal colonic anastomoses were performed as a model for disturbed healing in 36 Balb/c mice. Healing was evaluated endoscopically, macroscopically, and histologically after 1, 3 and 5 days. For detection of MMP-2/-9 expression fluorescence endoscopy (FE) was used following i.v.-administration of a Cy5.5-labeled MMP-2/-9 specific tracer. FE was complemented by quantification of the fluorescence signal using the MS-FX PRO Optical Imaging System. An overall leakage score was calculated and correlated with the results of FE. Results With increasing incidence of anastomotic leakage from POD1 (17%) to POD5 (83%) the uptake of the MMP tracer gradually increased (signal-to-noise ratio (SNR), POD1: 17.91 ± 1.251 vs. POD3: 30.56 ± 3.03 vs. POD5: 44.8 ± 4.473, P<0.0001). Mice with defective anastomotic healing showed significantly higher uptake compared to non-defective (SNR: 37.37± 3.63 vs. 26.16± 3.635, P = 0.0369). White light endoscopy and FE allowed evaluation of anastomotic healing and visualization of mucosal MMPs in vivo. Using FE based detection of MMPs in the anastomosis, an overall positive predictive value of 71.4% and negative predictive value of 66.6% was calculated for detection of anastomotic leakage. Conclusion During disturbed anastomotic healing increased expression of MMP-2/-9 was observed in the anastomotic tissue. Fluorescence endoscopy for detection of MMP-2/-9 during the healing process might be a promising tool for early identification of anastomotic leakage. PMID:29566031

Assessment of MMP-2/-9 expression by fluorescence endoscopy for evaluation of anastomotic healing in a murine model of anastomotic leakage.

PubMed

Neumann, Philipp-Alexander; Twardy, Vanessa; Becker, Felix; Geyer, Christiane; Schwegmann, Katrin; Mohr, Annika; Faust, Andreas; Lenz, Philipp; Rijcken, Emile

2018-01-01

Disturbance of intestinal wound closure leads to insufficient anastomotic healing and is associated with considerable morbidity following colorectal resections. Matrix metalloproteinases (MMPs) play a crucial role in regulation of wound closure. Here fluorescence endoscopy was evaluated for assessment of MMP-2/-9 expression during failed intestinal anastomotic healing. Distal colonic anastomoses were performed as a model for disturbed healing in 36 Balb/c mice. Healing was evaluated endoscopically, macroscopically, and histologically after 1, 3 and 5 days. For detection of MMP-2/-9 expression fluorescence endoscopy (FE) was used following i.v.-administration of a Cy5.5-labeled MMP-2/-9 specific tracer. FE was complemented by quantification of the fluorescence signal using the MS-FX PRO Optical Imaging System. An overall leakage score was calculated and correlated with the results of FE. With increasing incidence of anastomotic leakage from POD1 (17%) to POD5 (83%) the uptake of the MMP tracer gradually increased (signal-to-noise ratio (SNR), POD1: 17.91 ± 1.251 vs. POD3: 30.56 ± 3.03 vs. POD5: 44.8 ± 4.473, P<0.0001). Mice with defective anastomotic healing showed significantly higher uptake compared to non-defective (SNR: 37.37± 3.63 vs. 26.16± 3.635, P = 0.0369). White light endoscopy and FE allowed evaluation of anastomotic healing and visualization of mucosal MMPs in vivo. Using FE based detection of MMPs in the anastomosis, an overall positive predictive value of 71.4% and negative predictive value of 66.6% was calculated for detection of anastomotic leakage. During disturbed anastomotic healing increased expression of MMP-2/-9 was observed in the anastomotic tissue. Fluorescence endoscopy for detection of MMP-2/-9 during the healing process might be a promising tool for early identification of anastomotic leakage.

Anti-tumor effect and mechanism of cyclooxygenase-2 inhibitor through matrix metalloproteinase 14 pathway in PANC-1 cells.

PubMed

Li, Siyuan; Gu, Zhuoyu; Xiao, Zhiwei; Zhou, Ting; Li, Jun; Sun, Kan

2015-01-01

To investigate whether celecoxib, a selective cyclooxygenase-2 (COX-2) inhibitor, can attenuate proliferation, migration, invasion and MMP-14 expression in pancreatic cancer cells PANC-1 and the possible anti-tumor mechanism of celecoxib. Human pancreatic cancer cell line PANC-1 cells were treated with diverse concentrations of celecoxib (20, 60, 100 μmol/L). Cell proliferation, invasion and migration capabilities were measured by MTT colorimetry, transwell invasion assay, and scratch assay separately. At the same time, the protein expression of COX-2 and MMP-14 was assessed by ELISA. The capabilities of proliferation, invasion and migration in PANC-1 cells were attenuated in a concentration-dependent manner after treated with celecoxib, followed by the down-regulation of the protein expression of COX-2 and MMP-14. In addition, MMP-14 expression was significantly positively correlated with COX-2 expression. COX-2 inhibitor celecoxib can inhibit the proliferation, invasion and migration of PANC-1 cells via down-regulating the expression of MMP-14 in a concentration-dependent manner, thus contributing to its anti-tumor effect in pancreatic cancer.

MicroRNA 203 Modulates Glioma Cell Migration via Robo1/ERK/MMP-9 Signaling

PubMed Central

Dontula, Ranadheer; Dinasarapu, Ashok; Chetty, Chandramu; Pannuru, Padmavathi; Herbert, Engelhard; Ozer, Howard

2013-01-01

Glioblastoma (GBM) is the most common and malignant primary adult brain cancer. Allelic deletion on chromosome 14q plays an important role in the pathogenesis of GBM, and this site was thought to harbor multiple tumor suppressor genes associated with GBM, a region that also encodes microRNA-203 (miR-203). In this study, we sought to identify the role of miR-203 as a tumor suppressor in the pathogenesis of GBM. We analyzed the miR-203 expression data of GBM patients in 10 normal and 495 tumor tissue samples derived from The Cancer Genome Atlas data set. Quantitative real-time PCR and in situ hybridization in 10 high-grade GBM and 10 low-grade anaplastic astrocytoma tumor samples showed decreased levels of miR-203 expression in anaplastic astrocytoma and GBM tissues and cell lines. Exogenous expression of miR-203 using a plasmid expressing miR-203 precursor (pmiR-203) suppressed glioma cell proliferation, migration, and invasion. We determined that one relevant target of miR-203 was Robo1, given that miR-203 expression decreased mRNA and protein levels as determined by RT-PCR and Western blot analysis. Moreover, cotransfection experiments using a luciferase-based transcription reporter assay have shown direct regulation of Robo1 by miR-203. We also show that Robo1 mediates miR-203 mediated antimigratory functions as up-regulation of Robo1 abrogates miR-203 mediated antimigratory effects. We also show that miR-203 expression suppressed ERK phosphorylation and MMP-9 expression in glioma cells. Furthermore, we demonstrate that miR-203 inhibits migration of the glioma cells by disrupting the Robo1/ERK/MMP-9 signaling axis. Taken together, these studies demonstrate that up-regulation of Robo1 in response to the decrease in miR-203 in glioma cells is responsible for glioma tumor cell migration and invasion. PMID:24167656

Neutrophil-Derived MMP-8 Drives AMPK-Dependent Matrix Destruction in Human Pulmonary Tuberculosis.

PubMed

Ong, Catherine W M; Elkington, Paul T; Brilha, Sara; Ugarte-Gil, Cesar; Tome-Esteban, Maite T; Tezera, Liku B; Pabisiak, Przemyslaw J; Moores, Rachel C; Sathyamoorthy, Tarangini; Patel, Vimal; Gilman, Robert H; Porter, Joanna C; Friedland, Jon S

2015-05-01

Pulmonary cavities, the hallmark of tuberculosis (TB), are characterized by high mycobacterial load and perpetuate the spread of M. tuberculosis. The mechanism of matrix destruction resulting in cavitation is not well defined. Neutrophils are emerging as key mediators of TB immunopathology and their influx are associated with poor outcomes. We investigated neutrophil-dependent mechanisms involved in TB-associated matrix destruction using a cellular model, a cohort of 108 patients, and in separate patient lung biopsies. Neutrophil-derived NF-kB-dependent matrix metalloproteinase-8 (MMP-8) secretion was up-regulated in TB and caused matrix destruction both in vitro and in respiratory samples of TB patients. Collagen destruction induced by TB infection was abolished by doxycycline, a licensed MMP inhibitor. Neutrophil extracellular traps (NETs) contain MMP-8 and are increased in samples from TB patients. Neutrophils lined the circumference of human pulmonary TB cavities and sputum MMP-8 concentrations reflected TB radiological and clinical disease severity. AMPK, a central regulator of catabolism, drove neutrophil MMP-8 secretion and neutrophils from AMPK-deficient patients secrete lower MMP-8 concentrations. AMPK-expressing neutrophils are present in human TB lung biopsies with phospho-AMPK detected in nuclei. These data demonstrate that neutrophil-derived MMP-8 has a key role in the immunopathology of TB and is a potential target for host-directed therapy in this infectious disease.

Neutrophil-Derived MMP-8 Drives AMPK-Dependent Matrix Destruction in Human Pulmonary Tuberculosis

PubMed Central

Ong, Catherine W. M.; Elkington, Paul T.; Brilha, Sara; Ugarte-Gil, Cesar; Tome-Esteban, Maite T.; Tezera, Liku B.; Pabisiak, Przemyslaw J.; Moores, Rachel C.; Sathyamoorthy, Tarangini; Patel, Vimal; Gilman, Robert H.; Porter, Joanna C.; Friedland, Jon S.

2015-01-01

Pulmonary cavities, the hallmark of tuberculosis (TB), are characterized by high mycobacterial load and perpetuate the spread of M. tuberculosis. The mechanism of matrix destruction resulting in cavitation is not well defined. Neutrophils are emerging as key mediators of TB immunopathology and their influx are associated with poor outcomes. We investigated neutrophil-dependent mechanisms involved in TB-associated matrix destruction using a cellular model, a cohort of 108 patients, and in separate patient lung biopsies. Neutrophil-derived NF-kB-dependent matrix metalloproteinase-8 (MMP-8) secretion was up-regulated in TB and caused matrix destruction both in vitro and in respiratory samples of TB patients. Collagen destruction induced by TB infection was abolished by doxycycline, a licensed MMP inhibitor. Neutrophil extracellular traps (NETs) contain MMP-8 and are increased in samples from TB patients. Neutrophils lined the circumference of human pulmonary TB cavities and sputum MMP-8 concentrations reflected TB radiological and clinical disease severity. AMPK, a central regulator of catabolism, drove neutrophil MMP-8 secretion and neutrophils from AMPK-deficient patients secrete lower MMP-8 concentrations. AMPK-expressing neutrophils are present in human TB lung biopsies with phospho-AMPK detected in nuclei. These data demonstrate that neutrophil-derived MMP-8 has a key role in the immunopathology of TB and is a potential target for host-directed therapy in this infectious disease. PMID:25996154

Matrix Metalloproteinase-2 Cleavage of the β1 Integrin Ectodomain Facilitates Colon Cancer Cell Motility*

PubMed Central

Kryczka, Jakub; Stasiak, Marta; Dziki, Lukasz; Mik, Michał; Dziki, Adam; Cierniewski, Czesław S.

2012-01-01

Cancer cell invasion is a key element in metastasis that requires integrins for adhesion/de-adhesion, as well as matrix metalloproteinases (MMPs) for focalized proteolysis. Herein we show that MMP-2 is up-regulated in resected colorectal tumors and degrades β1 integrins with the release of fragments containing the β1 I-domain. The β1 cleavage pattern is similar to that produced by digestion of α5β1 and α2β1 with MMP-2. Two such fragments, at 25 and 75 kDa, were identified after immunoprecipitation, with monoclonal antibody BD610468 reacting with the NH2-terminal I-like ectodomain followed by SDS-PAGE and microsequencing using electrospray (ISI-Q-TOF-Micromass) spectrometry. Cleavage of the β1 integrin can be abolished by inhibition of MMP-2 activity; it can be induced by up-regulation of MMP-2 expression, as exemplified by HT29 colon cancer cells transfected with pCMV6-XL5-MMP-2. Co-immunoprecipitation studies of colon cancer cells showed that the β1 integrin subunit is associated with MMP-2. The MMP-2-mediated shedding of the I-like domain from β1 integrins resulted in decreased adhesion of colon cancer cells to collagen and fibronectin, thus abolishing their receptivity. Furthermore, such cells showed enhanced motility as evaluated by a “wound healing-like” assay and time-lapse microscopy, indicating their increased invasiveness. Altogether, our data demonstrate that MMP-2 amplifies the motility of colon cancer cells, not only by digesting the extracellular matrix components in the vicinity of cancer cells but also by inactivating their major β1 integrin receptors. PMID:22898815

Polysaccharides of Aloe vera induce MMP-3 and TIMP-2 gene expression during the skin wound repair of rat.

PubMed

Tabandeh, Mohammad Reza; Oryan, Ahmad; Mohammadalipour, Adel

2014-04-01

Polysaccharides are the main macromolecules of Aloe vera gel but no data about their effect on extracellular matrix (ECM) elements are available. Here, mannose rich Aloe vera polysaccharides (AVP) with molecular weight between 50 and 250 kDa were isolated and characterized. Open cutaneous wounds on the back of 45 rats (control and treated) were daily treated with 25mg (n=15) and 50 mg (n=15) AVP for 30 days. The levels of MMP-3 and TIMP-2 gene expression were analyzed using real time PCR. The levels of n-acetyl glucosamine (NAGA), n-acetyl galactosamine (NAGLA) and collagen contents were also measured using standard biochemical methods. Faster wound closure was observed at day 15 post wounding in AVP treated animals in comparison with untreated group. At day 10 post wounding, AVP inhibited MMP-3 gene expression, while afterwards MMP-3 gene expression was upregulated. AVP enhanced TIMP-2 gene expression, collagen, NAGLA and NAGA synthesis in relation to untreated wounds. Our results suggest that AVP has positive effects on the regulation of ECM factor synthesis, which open up new perspectives for the wound repair activity of Aloe vera polysaccharide at molecular level. Copyright © 2014 Elsevier B.V. All rights reserved.

Increased expression of matrix metalloproteinase-1 in systemic vessels of preeclamptic women: a critical mediator of vascular dysfunction.

PubMed

Estrada-Gutierrez, Guadalupe; Cappello, Renato E; Mishra, Nikita; Romero, Roberto; Strauss, Jerome F; Walsh, Scott W

2011-01-01

This study was conducted to determine the following: (1) whether matrix metalloproteinase-1 (MMP-1) is increased in systemic vessels of preeclamptic women, (2) whether this increase might be mediated by neutrophils, and (3) whether MMP-1 could be responsible for vascular dysfunction. Omental arteries and plasma were collected from healthy pregnant and preeclamptic women. Omental arteries were evaluated for gene and protein expression of MMP-1, collagen type 1α, tissue inhibitor of metalloproteinase-1, and vascular reactivity to MMP-1. Gene and protein expression levels were also evaluated in human vascular smooth muscle cells (VSMCs) co-cultured with activated neutrophils, reactive oxygen species, or tumor necrosis factor α. Vessel expression of MMP-1 and circulating MMP-1 levels were increased in preeclamptic women, whereas vascular expression of collagen or tissue inhibitor of metalloproteinase-1 were down-regulated or unchanged. In cultured VSMCs, the imbalance in collagen-regulating genes of preeclamptic vessels was reproduced by treatment with neutrophils, tumor necrosis factor α, or reactive oxygen species. Chemotaxis studies with cultured cells revealed that MMP-1 promoted recruitment of neutrophils via vascular smooth muscle release of interleukin-8. Furthermore, MMP-1 induced vasoconstriction via protease-activated receptor-1, whose expression was significantly increased in omental arteries of preeclamptic women and in VSMCs co-cultured with neutrophils. Collectively, these findings disclose a novel role for MMP-1 as a mediator of vasoconstriction and vascular dysfunction in preeclampsia. Copyright © 2011 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.

Acute and long-term transcriptional responses in sulfur mustard-exposed SKH-1 hairless mouse skin.

PubMed

Vallet, V; Poyot, T; Cléry-Barraud, C; Coulon, D; Sentenac, C; Peinnequin, A; Boudry, I

2012-03-01

Sulfur mustard (HD) ranks among the alkylating chemical warfare agents. Skin contact with HD produces an inflammatory response that evolves into separation at the epidermal-dermal junction conducting to blistering and epidermis necrosis. Up to now, current treatment strategies of HD burns have solely consisted in symptomatic management of skin damage. Therapeutic efficacy studies are still being conducted; classically using appropriate animal skin toxicity models. In order to substantiate the use of SKH-1 hairless mouse as an appropriate model for HD-induced skin lesions, we investigate the time-dependent quantitative gene expression of various selected transcripts associated to the dorsal skin exposure to HD saturated vapors. Using quantitative real time polymerase chain reaction (RT-qPCR), the expression of interleukins (IL-1β and IL-6), tumor necrosis factor (TNF)-α, macrophage inflammatory proteins (MIP)-2α (also called Cxcl2) and MIP-1αR (also called Ccr1), matrix metalloproteases (MMP-9 and MMP-2), laminin γ2 monomer (Lamc2) and keratin (K)1 was determined up to 21 days after HD challenge in order to allow enough time for wound repair to begin. Specific transcript RT-qPCR analysis demonstrated that IL-6, IL-1β, Ccr1, Cxcl2 mRNA levels increased as early as 6 h in HD-exposed skins and remained up-regulated over a 14-day period. Topical application of HD also significantly up-regulated MMP-9, TNF-α, and Lamc2 expression at specific time points. In contrast, MMP-2 mRNA levels remained unaffected by HD over the time-period considered, whereas that long-term study revealed that K1 mRNA level significantly increased only 21 days after HD challenge. Our study hereby provides first-hand evidence to substantiate a long period variation expression in the inflammatory cytokine, MMPs and structural components following cutaneous HD exposure in hairless mouse SKH-1. Our data credit the use of SKH-1 for investigating mechanisms of HD-induced skin toxicity and for the development of pharmacological countermeasures.

Caveolin-1 mediates tissue plasminogen activator-induced MMP-9 up-regulation in cultured brain microvascular endothelial cells.

PubMed

Jin, Xinchun; Sun, Yanyun; Xu, Ji; Liu, Wenlan

2015-03-01

Thrombolysis with tissue plasminogen activator (tPA) increases matrix metalloproteinase-9 (MMP-9) activity in the ischemic brain, which exacerbates blood-brain barrier injury and increases the risk of symptomatic cerebral hemorrhage. The mechanism through which tPA enhances MMP-9 activity is not well understood. Here we report an important role of caveolin-1 in mediating tPA-induced MMP-9 synthesis. Brain microvascular endothelial cell line bEnd3 cells were incubated with 5 or 20 μg/ml tPA for 24 hrs before analyzing MMP-9 levels in the conditioned media and cellular extracts by gelatin zymography. tPA at a dose of 20 μg/mL tPA, but not 5 μg/mL, significantly increased MMP-9 level in cultured media while decreasing it in cellular extracts. Concurrently, tPA treatment induced a 2.3-fold increase of caveolin-1 protein levels in endothelial cells. Interestingly, knockdown of Cav-1 with siRNA inhibited tPA-induced MMP-9 mRNA up-regulation and MMP-9 increase in the conditioned media, but did not affect MMP-9 decrease in cellular extracts. These results suggest that caveolin-1 critically contributes to tPA-mediated MMP-9 up-regulation, but may not facilitate MMP-9 secretion in endothelial cells. Thrombolysis with tissue plasminogen activator (tPA) increases matrix metalloproteinase-9 (MMP-9) activity in the ischemic brain, which exacerbates ischemic blood brain barrier (BBB) injury and increases the risk of symptomatic cerebral hemorrhage. Our results suggest a novel mechanism underlying this tPA-MMP 9 axis. In response to tPA treatment, caveolin-1 protein levels increased in endothelial cells, which mediate MMP-9 mRNA up-regulation and its secretion into extracellular space. Caveolin-1 may, however, not facilitate MMP-9 secretion in endothelial cells. Our data suggest caveolin-1 as a novel therapeutic target for protecting the BBB against ischemic damage. The schematic outlines tPA-induced MMP-9 upreguation. © 2015 International Society for Neurochemistry.

Regulation of matrix metalloproteinases (MMPs) expression and secretion in MDA-MB-231 breast cancer cells by LIM and SH3 protein 1 (LASP1).

PubMed

Endres, Marcel; Kneitz, Susanne; Orth, Martin F; Perera, Ruwan K; Zernecke, Alma; Butt, Elke

2016-09-27

The process of tumor invasion requires degradation of extracellular matrix by proteolytic enzymes. Cancer cells form protrusive invadopodia, which produce and release matrix metalloproteinases (MMPs) to degrade the basement membrane thereby enabling metastasis. We investigated the effect of LASP1, a newly identified protein in invadopodia, on expression, secretion and activation of MMPs in invasive breast tumor cell lines.By analyzing microarray data of in-house generated control and LASP1-depleted MDA-MB-231 breast cancer cells, we observed downregulation of MMP1, -3 and -9 upon LASP1 depletion. This was confirmed by Western blot analysis. Conversely, rescue experiments restored in part MMP expression and secretion. The regulatory effect of LASP1 on MMP expression was also observed in BT-20 breast cancer cells as well as in prostate and bladder cancer cell lines.In line with bioinformatic FunRich analysis of our data, which mapped a high regulation of transcription factors by LASP1, public microarray data analysis detected a correlation between high LASP1 expression and enhanced c-Fos levels, a protein that is part of the transcription factor AP-1 and known to regulate MMP expression. Compatibly, in luciferase reporter assays, AP-1 showed a decreased transcriptional activity after LASP1 knockdown.Zymography assays and Western blot analysis revealed an additional promotion of MMP secretion into the extracellular matrix by LASP1, thus, most likely, altering the microenvironment during cancer progression.The newly identified role of LASP1 in regulating matrix degradation by affecting MMP transcription and secretion elucidated the migratory potential of LASP1 overexpressing aggressive tumor cells in earlier studies.

Fibroblast extracellular matrix gene expression in response to keratinocyte-releasable stratifin.

PubMed

Ghaffari, Abdi; Li, Yunyaun; Karami, Ali; Ghaffari, Mazyar; Tredget, Edward E; Ghahary, Aziz

2006-05-15

Termination of wound-healing process requires a fine balance between connective tissue deposition and its hydrolysis. Previously, we have demonstrated that keratinocyte-releasable stratifin, also known as 14-3-3 sigma protein, stimulates collagenase (MMP-1) expression in dermal fibroblasts. However, role of extracellular stratifin in regulation of extracellular matrix (ECM) factors and other matrix metalloproteinases (MMPs) in dermal fibroblast remains unexplored. To address this question, large-scale ECM gene expression profile were analyzed in human dermal fibroblasts co-cultured with keratinocytes or treated with recombinant stratifin. Superarray pathway-specific microarrays were utilized to identify upregulation or downregulation of 96 human ECM and adhesion molecule genes. RT-PCR and Western blot were used to validate microarray expression profiles of selected genes. Comparison of gene profiles with the appropriate controls showed a significant (more than twofold) increase in expression of collagenase-1, stromelysin-1 and -2, neutrophil collagenase, and membrane type 5 MMP in dermal fibroblasts treated with stratifin or co-cultured with keratinocytes. Expression of type I collagen and fibronectin genes decreased in the same fibroblasts. The results of a dose-response experiment showed that stratifin stimulates the expression of stromelysin-1 (MMP-3) mRNA by dermal fibroblasts in a concentration-dependent fashion. Furthermore, Western blot analysis of fibroblast-conditioned medium showed a peak in MMP-3 protein levels 48 h following treatment with recombinant stratifin. In a lasting-effect study, MMP-3 protein was detected in fibroblast-condition medium for up to 72 h post removal of stratifin. In conclusion, our results suggest that keratinocyte-releasable stratifin plays a major role in induction of ECM degradation by dermal fibroblasts through stimulation of key MMPs, such as MMP-1 and MMP-3. Therefore, stratifin protein may prove to be a useful target for clinical intervention in controlling excessive wound healing in fibrotic conditions. Copyright 2006 Wiley-Liss, Inc.

C-type natriuretic peptide suppresses mesangial proliferation and matrix expression via a MMPs/TIMPs-independent pathway in vitro.

PubMed

Huang, Bao Yu; Hu, Peng; Zhang, Dong Dong; Jiang, Guang Mei; Liu, Si Yan; Xu, Yao; Wu, Yang Fang; Xia, Xun; Wang, Ya

2017-08-01

C-type natriuretic peptide (CNP) acts mainly in a local, paracrine fashion to regulate vascular tone and cell proliferation. Although several in vivo studies have demonstrated that CNP exerts an inhibitory effect on mesangial matrix generation, a limited number of reports exist about the anti-extracellular matrix (ECM) accumulation effect of CNP and its underlying mechanisms in mesangial cells (MCs) in vitro. In this study, human MCs were incubated in serum-containing medium in the absence or presence of CNP (0, 10 and 100 pM) for 24, 48 and 72 h, respectively. CNP administration significantly suppresses MCs proliferation and collagen (Col)-IV expression in a time- and dose-dependent manner. In addition, the study presented herein was designed as a first demonstration of the regulative effects of CNP on the metabolisms of matrix metalloproteinases (MMPs) and tissue inhibitors of metalloproteinases (TIMPs) in MCs in vitro, and found that: (1) CNP administration significantly decreased the secretion and expression of MMP-2 and MMP-9 in the cultured MCs; (2) the secretion and expression of TIMP-1 progressively elevated after treatment with CNP for 24 and 48 h, whereas declined at later time point; (3) CNP expression was negatively correlated with MMP-2 and MMP-9 expression; (4) the balance of MMPs/TIMPs was shifted toward the reduction in MMP-2 and MMP-9 activity and/or the increment in TIMP-1 expression, which could not account for the down-regulation of Col-IV expression in CNP-treated MCs. In conclusion, CNP suppresses mesangial proliferation and ECM expression via a MMPs/TIMPs-independent pathway in vitro.

Activation of Toll-like receptor-9 promotes cellular migration via up-regulating MMP-2 expression in oral squamous cell carcinoma.

PubMed

Ruan, Min; Zhang, Zun; Li, Siyi; Yan, Min; Liu, Shengwen; Yang, Wenjun; Wang, Lizheng; Zhang, Chenping

2014-01-01

Activation of Toll like receptors (TLRs) signaling has been implicated in promoting malignant cell invasion and metastatic potential. Previously we demonstrated that increased TLR-9 expression predicted poor survival in oral cancer patients. The objective of this study is to further investigate the roles and potential molecular mechanisms of TLR-9 signaling in human oral cancer cell invasion. Cell migration, invasion and protein expression were detected by wound healing assay, Transwell chambers model and western blot. The secretion and activity levels of metalloproteinases-2/9 were quantified by ELISA and Gelatin zymography. EMSA and ChIP assays were employed to detect the activity of AP-1signal pathway. TLR-9 siRNA transfection was used to regulate the expression and activity of TLR-9 in oral cancer cell line HB cells. The results of both wound healing assay and in vitro Transwell assay revealed that activation of TLR-9 induced dose- and time- dependent migration and invasion of HB cells. An increased expression, secretion and activity of MMP-2 were observed upon the treatment of CpG-ODN. The TLR-9 signaling-mediated MMP-2 expression appeared to be a consequence of AP-1 activation, because that their DNA binding activity was enhanced by CpG-ODN treatment. All these influences were efficiently repressed by the knockdown of TLR-9 through siRNA or pretreatment of an AP-1 inhibitor. Activation of TLR-9 signaling could promote human oral cancer HB cells invasion with the induction of MMP-2 presentation by attenuating AP-1 binding activity, suggesting a novel anti-metastatic application for TLR-9 targeted therapy in oral cancer in the future.

Co-ordinated expression of MMP-2 and its putative activator, MT1-MMP, in human placentation.

PubMed

Bjørn, S F; Hastrup, N; Lund, L R; Danø, K; Larsen, J F; Pyke, C

1997-08-01

The spatial expression of mRNA for matrix metalloproteinase 2 (MMP-2), its putative activator, the membrane-type 1 matrix metalloproteinase (MT1-MMP), and the MMP-2 substrate type IV collagen was investigated in human placentas of both normal and tubal ectopic pregnancies and in cyclic endometrium using in-situ hybridization. Cytokeratin staining applied to adjacent sections was used to identify epithelial and trophoblast cells. In both normal and tubal pregnancies MT1-MMP, MMP-2 and type IV collagen mRNA were highly expressed and co-localized in the extravillous cytotrophoblasts of anchoring villi, in cytotrophoblasts that had penatrated into the placental bed and in cytotrophoblastic cell islands. In addition, the decidual cells of normal pregnancies in some areas co-expressed MT1-MMP and MMP-2 mRNA, with moderate signals for both components. Fibroblast-like stromal cells in tubal pregnancies were positive for MMP-2 mRNA but generally negative for MT1-MMP mRNA. The consistent co-localization of MT1-MMP with MMP-2 and type IV collagen in the same subset of cytotrophoblasts strongly suggests that all three components co-operate in the tightly regulated fetal invasion process. The co-expression of MT1-MMP and MMP-2 mRNA in some of the decidual cells indicates that these cells are also actively involved in the placentation process.

Curcumin synergistically increases effects of β-interferon and retinoic acid on breast cancer cells in vitro and in vivo by up-regulation of GRIM-19 through STAT3-dependent and STAT3-independent pathways.

PubMed

Ren, Min; Wang, Ying; Wu, Xiaodong; Ge, Suxia; Wang, Benzhong

2017-03-01

The study aimed to investigate the effects of combination treatment of curcumin and β-interferon (IFN-β)/retinoic acid (RA) on breast cancer cells, including cell viability, apoptosis and migration, and to determine the mechanisms related to GRIM-19 through STAT3-dependent and STAT3-independent pathways. The following groups were used for the in vitro experiment: control siRNA, GRIM-19 siRNA, IFN-β/RA and IFN-β/RA + curcumin. Cell viability is by the MTT method, cell apoptosis by flow cytometry and cell migration by wound healing experiment; GRIM-19, STAT3, survivin, Bcl-2, GADD153 and COX-2 expression was measured by Western blot. In vivo experiment, MCF-7 cells were subcutaneously injected into nude mice. GRIM-19 siRNA promoted MCF-7 cell proliferation and migration; inhibited cell apoptosis; and promoted the expression of STAT3, survivin, Bcl-2 and MMP-9. IFN-β/RA inhibited cell proliferation and migration; promoted cell apoptosis; up-regulated GRIM-19; and inhibited the expression of STAT3, survivin, Bcl-2 and MMP-9. Combination treatment of curcumin and IFN-β/RA had a stronger effect than that of the IFN-β/RA group. In addition, curcumin and IFN-β/RA combination inhibited the expression of COX-2 and up-regulated GADD153. Curcumin synergistically increases the effects of IFN-β/RA on breast cancer cells. The mechanism may be related to the up-regulation of GRIM-19 through STAT3-dependent and STAT3-independent pathways.

Coronin 3 promotes gastric cancer metastasis via the up-regulation of MMP-9 and cathepsin K.

PubMed

Ren, Gui; Tian, Qifei; An, Yanxin; Feng, Bin; Lu, Yuanyuan; Liang, Jie; Li, Kai; Shang, Yulong; Nie, Yongzhan; Wang, Xin; Fan, Daiming

2012-09-14

Coronins are a family of highly evolutionary conserved proteins reportedly involved in the regulation of actin cytoskeletal dynamics, although only coronin 3 has been shown to be related to cancer cell migration. In glioblastoma cells, the knockdown of coronin 3 inhibits cell proliferation and invasion. Coronin 3 is also associated with the aggression and metastasis of hepatocellular carcinoma. In this paper, we analyze the migration, invasion and metastasis abilities of gastric cancer cells after up- or down-regulation of coronin 3, and explore the mechanism of coronin 3 in the process of gastric cancer metastasis. The expression of coronin 3 was higher in the highly metastatic sub-cell line MKN28-M, which we established in our laboratory. We also demonstrated that the expression of coronin 3 was remarkably higher in lymph lode metastases than in primary gastric cancer tissues, and over-expression of coronin 3 was correlated with the increased clinical stage and lymph lode metastasis. Recombinant lentiviral vectors encoding shRNAs were designed to down-regulate coronin 3 expression in gastric cancer cell lines. Stable knockdown of coronin 3 by this lentiviral vector could efficiently inhibit the migration and invasion of MKN45 gastric cancer cells. In contrast, up-regulation of coronin 3 significantly enhanced migration and invasion of MKN28-NM cells. In addition, knockdown of coronin 3 significantly reduced liver metastasis in mice after tail vein injection of gastric cancer cells. The Human Tumor Metastasis PCR Array was used to screen the metastasis-associated genes identified by the down-regulation of coronin 3, and the results suggested that, following the knockdown of coronin 3, the tumor cell migration and invasion were inhibited by the reduced expression of MMP-9 and cathepsin K. Coronin 3 is highly expressed in gastric cancer metastases and can promote the metastatic behaviors of gastric cancer cells, including their migration and invasion.

BAG3 increases the invasiveness of uterine corpus carcinoma cells by suppressing miR-29b and enhancing MMP2 expression.

PubMed

Habata, Shutaro; Iwasaki, Masahiro; Sugio, Asuka; Suzuki, Miwa; Tamate, Masato; Satohisa, Seiro; Tanaka, Ryoichi; Saito, Tsuyoshi

2015-05-01

Approximately 30% of uterine corpus carcinomas are diagnosed at an advanced stage and have a poor prognosis. Our previous study indicated that BCL2-associated athanogene 3 (BAG3) enhances matrix metalloproteinase-2 (MMP2) expression and binds to MMP2 to positively regulate the process of cell invasion in ovarian cancer cells. Recently, altered miRNA expression patterns were observed in several groups of patients with endometrial cancers. One of the altered miRNAs, miR-29b, reportedly reduces tumor invasiveness by suppressing MMP2 expression. Our aim in the present study was to examine the relationships among BAG3, miR-29b and MMP2 in endometrioid adenocarcinoma cells. We found that BAG3 suppresses miR-29b expression and enhances MMP2 expression, which in turn increases cell motility and invasiveness. Moreover, restoration of miR-29b through BAG3 knockdown reduced MMP2 expression, as well as cell motility and invasiveness. Collectively, our findings indicate that BAG3 enhances MMP2 expression by suppressing miR-29b, thereby increasing the metastatic potential of endometrioid adenocarcinomas.

[Anti-sense miRNA-21 oligonucleotide inhibits Tb 3.1 human tongue squamous cell carcinoma growth in vitro].

PubMed

Tao, Ying-jie; Ren, Yu; Dong, Jia-bin; Zhang, Lun; Cheng, Jun-ping; Zhou, Xuan

2011-02-01

To investigate the effect of micro RNA-21 (miRNA-21) knocking on the Tb3.1 human tongue squamous cell carcinoma growth. Anti-sense miRNA-21 oligonucleotide was delivered with oligofectamine to suppress Tb 3.1 tongue cancer cell growth in vitro. Real-time polymerase chain reaction (PCR) was conducted to detect the miRNA-21 expression after transfection. Methyl thiazolyl tetrazolium (MTT) assay was used to determine Tb 3.1 cell survival rate. Apoptosis were examined by flow-cytometry. Matrigel matrix and transwell assay were used to determine Tb 3.1 cell colony formation and migration ability. Antigen KI-67 (Ki67), B cell lymphoma (Bcl-2), phosphatase and tensin homolog (PTEN), matrirx metalloproteinase 2 (MMP-2, MMP-9) and tissue inhibitor of metalloproteinase 1 (TIMP-1) protein expression in Tb 3.1 cell were measured by Western blotting. miRNA-21 expression was decreased in miRNA-21 antisense oligonucleotide (ASODN) group. The survival rate of Tb 3.1 cells with AS-miRNA-21 transfection was significantly suppressed (F = 27.02, P = 0.00) and early phase apoptosis (F = 26.641, P = 0.001) induced in Tb 3.1 cell. Ki67, Bcl-2, MMP-2 and MMP-9 protein were down regulated while PTEN and TIMP-1 protein expression was increased. Blocking miRNA-21 expression in Tb3.1 cell could suppress cancer cell growth in vitro and miRNA-21 can serve as a novel target candidate for human tongue cancer gene therapy.

Association of MMP7 -181A→G Promoter Polymorphism with Gastric Cancer Risk: INFLUENCE OF NICOTINE IN DIFFERENTIAL ALLELE-SPECIFIC TRANSCRIPTION VIA INCREASED PHOSPHORYLATION OF cAMP-RESPONSE ELEMENT-BINDING PROTEIN (CREB).

PubMed

Kesh, Kousik; Subramanian, Lakshmi; Ghosh, Nillu; Gupta, Vinayak; Gupta, Arnab; Bhattacharya, Samir; Mahapatra, Nitish R; Swarnakar, Snehasikta

2015-06-05

Elevated expression of matrix metalloproteinase7 (MMP7) has been demonstrated to play a pivotal role in cancer invasion. The -181A→G (rs11568818) polymorphism in the MMP7 promoter modulates gene expression and possibly affects cancer progression. Here, we evaluated the impact of -181A→G polymorphism on MMP7 promoter activity and its association with gastric cancer risk in eastern Indian case-control cohorts (n = 520). The GG genotype as compared with the AA genotype was predisposed (p = 0.02; odds ratio = 1.9, 95% confidence interval = 1.1-3.3) to gastric cancer risk. Stratification analysis showed that tobacco addiction enhanced gastric cancer risk in GG subjects when compared with AA subjects (p = 0.03, odds ratio = 2.46, and 95% confidence interval = 1.07-5.68). Meta-analysis revealed that tobacco enhanced the risk for cancer more markedly in AG and GG carriers. Activity and expression of MMP7 were significantly higher in GG than in AA carriers. In support, MMP7 promoter-reporter assays showed greater transcriptional activity toward A to G transition under basal/nicotine-induced/cAMP-response element-binding protein (CREB) overexpressed conditions in gastric adenocarcinoma cells. Moreover, nicotine (a major component of tobacco) treatment significantly up-regulated MMP7 expression due to enhanced CREB phosphorylation followed by its nuclear translocation in gastric adenocarcinoma cells. Furthermore, chromatin immunoprecipitation experiments revealed higher binding of phosphorylated CREB with the -181G than the -181A allele. Altogether, specific binding of phosphorylated CREB to the G allele-carrying promoter enhances MMP7 gene expression that is further augmented by nicotine due to increased CREB phosphorylation and thereby increases the risk for gastric cancer. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

Curcumin exhibits anti-tumor effect and attenuates cellular migration via Slit-2 mediated down-regulation of SDF-1 and CXCR4 in endometrial adenocarcinoma cells.

PubMed

Sirohi, Vijay Kumar; Popli, Pooja; Sankhwar, Pushplata; Kaushal, Jyoti Bala; Gupta, Kanchan; Manohar, Murli; Dwivedi, Anila

2017-06-01

Although curcumin shows anti-proliferative and anti-inflammatory activities in various cancers, the effect of curcumin on cellular migration in endometrial adenocarcinoma cells remains to be understood. The current investigation was aimed to explore the anti-proliferative and anti-migratory effects of curcumin and its mechanism of action in endometrial cancer cells. Our in-vitro and in-vivo experimental studies showed that curcumin inhibited the proliferation of endometrial cancer cells and suppressed the tumor growth in Ishikawa xenograft mouse model. Curcumin induced ROS-mediated apoptosis in endometrial cancer cells. Curcumin suppressed the migration rate of Ishikawa and Hec-1B cells as analyzed by scratch wound assay. In transwell migration studies, knock down of Slit-2 reversed the anti-migratory effect of curcumin in these cell lines. Curcumin significantly up-regulated the expression of Slit-2 in Ishikawa, Hec-1B and primary endometrial cancer cells while it down-regulated the expression of stromal cell-derived factor-1 (SDF-1) and CXCR4 which in turn, suppressed the expression of matrix metallopeptidases (MMP) 2 and 9, thus attenuating the migration of endometrial cancer cells. In summary, we have demonstrated that curcumin has inhibitory effect on cellular migration via Slit-2 mediated down-regulation of CXCR4, SDF-1, and MMP2/MMP9 in endometrial carcinoma cells. These findings helped explore the role of Slit-2 in endometrial cancer cells. Copyright © 2017 Elsevier Inc. All rights reserved.

Effects of PTHrP on expression of MMP9 and MMP13 in sika deer antler chondrocytes.

PubMed

Wang, Shou-Tang; Gao, Ying-Jie; Duan, Cui-Cui; Li, Dang-Dang; Tian, Xue-Chao; Zhang, Qiao-Ling; Guo, Bin; Yue, Zhan-Peng

2013-12-01

Deer antlers are the only mammalian appendages to display an annual cycle of full regeneration. However, little is known about the molecular mechanisms of antler regeneration. Our previous study has demonstrated that parathyroid hormone-related peptide (PTHrP) can promote proliferation of antler chondrocytes and inhibit its differentiation, but the mechanism underlying such regulation is not fully understood. We have determined the role of PTHrP on the mRNA expression of matrix metalloproteinase-9 (MMP9) and MMP13 in the antler chondrocytes. The possible pathways that transduce PTHrP effects were examined. In situ hybridization showed that MMP9 and MMP13 were mainly localized in the dermal fibroblasts, perichondrium, and cartilage in the sika deer antler, of which MMP9 and MMP13 were highly expressed in the chondrocytes. Exogenous PTHrP could inhibit the expression of MMP9 and MMP13 in the antler chondrocytes. The inhibitory effect of PTHrP on MMP9 was abolished by JNK inhibitor, SP600125, while P38MAPK inhibitor SB203850 and PKC inhibitor GF109203X could rescue the inhibitory effect of PTHrP on MMP13. The results suggest that PTHrP can inhibit MMP9 expression by JNK signaling pathway and MMP13 expression by p38MAPK and PKC signaling pathways in the antler chondrocytes. Thus PTHrP is involved in the control of antler chondrocytes maturation and cartilage matrix degradation. © 2013 International Federation for Cell Biology.

Migration of bone marrow and cord blood mesenchymal stem cells in vitro is regulated by stromal-derived factor-1-CXCR4 and hepatocyte growth factor-c-met axes and involves matrix metalloproteinases.

PubMed

Son, Bo-Ra; Marquez-Curtis, Leah A; Kucia, Magda; Wysoczynski, Marcin; Turner, A Robert; Ratajczak, Janina; Ratajczak, Mariusz Z; Janowska-Wieczorek, Anna

2006-05-01

Human mesenchymal stem cells (MSCs) are increasingly being considered in cell-based therapeutic strategies for regeneration of various organs/tissues. However, the signals required for their homing and recruitment to injured sites are not yet fully understood. Because stromal-derived factor (SDF)-1 and hepatocyte growth factor (HGF) become up-regulated during tissue/organ damage, in this study we examined whether these factors chemoattract ex vivo-expanded MSCs derived from bone marrow (BM) and umbilical cord blood (CB). Specifically, we investigated the expression by MSCs of CXCR4 and c-met, the cognate receptors of SDF-1 and HGF, and their functionality after early and late passages of MSCs. We also determined whether MSCs express matrix metalloproteinases (MMPs), including membrane type 1 (MT1)-MMP, matrix-degrading enzymes that facilitate the trafficking of hematopoietic stem cells. We maintained expanded BM- or CB-derived MSCs for up to 15-18 passages with monitoring of the expression of 1) various tissue markers (cardiac and skeletal muscle, neural, liver, and endothelial cells), 2) functional CXCR4 and c-met, and 3) MMPs. We found that for up to 15-18 passages, both BM- and CB-derived MSCs 1) express mRNA for cardiac, muscle, neural, and liver markers, as well as the vascular endothelial (VE) marker VE-cadherin; 2) express CXCR4 and c-met receptors and are strongly attracted by SDF-1 and HGF gradients; 3) express MMP-2 and MT1-MMP transcripts and proteins; and 4) are chemo-invasive across the reconstituted basement membrane Matrigel. These in vitro results suggest that the SDF-1-CXCR4 and HGF-c-met axes, along with MMPs, may be involved in recruitment of expanded MSCs to damaged tissues.

Tumour-associated neutrophils and loss of epithelial PTEN can promote corticosteroid-insensitive MMP-9 expression in the chronically inflamed lung microenvironment.

PubMed

Vannitamby, Amanda; Seow, Huei Jiunn; Anderson, Gary; Vlahos, Ross; Thompson, Michelle; Steinfort, Daniel; Irving, Louis B; Bozinovski, Steven

2017-12-01

Matrix metalloproteinase-9 (MMP-9) is increased in a number of pathological lung conditions, where the proteinase contributes to deleterious remodelling of the airways. While both lung cancer and COPD are associated with increased MMP-9 expression, the cellular and molecular drivers of MMP-9 remain unresolved. In this study, MMP-9 transcript measured within the tumour region from patients with non-small-cell lung cancer (NSCLC) and coexisting COPD was found to be uniformly increased relative to adjacent tumour-free tissue. MMP-9 gene expression and immunohistochemistry identified tumour-associated neutrophils, but not macrophages, as a predominant source of this proteinase. In addition, PTEN gene expression was significantly reduced in tumour and there was evidence of epithelial MMP-9 expression. To explore whether PTEN can regulate epithelial MMP-9 expression, a small interfering (si)RNA knockdown strategy was used in Beas-2B bronchial epithelial cells. PTEN knockdown by siRNA selectively increased MMP-9 expression in response to lipopolysaccharide in a corticosteroid-insensitive manner. In summary, tumour-associated neutrophils represent an important source of MMP-9 in NSCLC, and loss of epithelial PTEN may further augment steroid-insensitive expression. Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://www.bmj.com/company/products-services/rights-and-licensing/.

Fluid shear promotes chondrosarcoma cell invasion by activating matrix metalloproteinase 12 via IGF-2 and VEGF signaling pathways

PubMed Central

Wang, P; Chen, S-H; Hung, W-C; Paul, C; Zhu, F; Guan, P-P; Huso, DL; Kontrogianni-Konstantopoulos, A; Konstantopoulos, K

2015-01-01

Interstitial fluid flow in and around the tumor tissue is a physiologically relevant mechanical signal that regulates intracellular signaling pathways throughout the tumor. Yet, the effects of interstitial flow and associated fluid shear stress on the tumor cell function have been largely overlooked. Using in vitro bioengineering models in conjunction with molecular cell biology tools, we found that fluid shear (2 dyn/cm2) markedly upregulates matrix metalloproteinase 12 (MMP-12) expression and its activity in human chondrosarcoma cells. MMP-12 expression is induced in human chondrocytes during malignant transformation. However, the signaling pathway regulating MMP-12 expression and its potential role in human chondrosarcoma cell invasion and metastasis have yet to be delineated. We discovered that fluid shear stress induces the synthesis of insulin growth factor-2 (IGF-2) and vascular endothelial growth factor (VEGF) B and D, which in turn transactivate MMP-12 via PI3-K, p38 and JNK signaling pathways. IGF-2-, VEGF-B- or VEGF-D-stimulated chondrosarcoma cells display markedly higher migratory and invasive potentials in vitro, which are blocked by inhibiting MMP-12, PI3-K, p38 or JNK activity. Moreover, recombinant human MMP-12 or MMP-12 overexpression can potentiate chondrosarcoma cell invasion in vitro and the lung colonization in vivo. By reconstructing and delineating the signaling pathway regulating MMP-12 activation, potential therapeutic strategies that interfere with chondrosarcoma cell invasion may be identified. PMID:25435370

Fluid shear promotes chondrosarcoma cell invasion by activating matrix metalloproteinase 12 via IGF-2 and VEGF signaling pathways.

PubMed

Wang, P; Chen, S-H; Hung, W-C; Paul, C; Zhu, F; Guan, P-P; Huso, D L; Kontrogianni-Konstantopoulos, A; Konstantopoulos, K

2015-08-27

Interstitial fluid flow in and around the tumor tissue is a physiologically relevant mechanical signal that regulates intracellular signaling pathways throughout the tumor. Yet, the effects of interstitial flow and associated fluid shear stress on the tumor cell function have been largely overlooked. Using in vitro bioengineering models in conjunction with molecular cell biology tools, we found that fluid shear (2 dyn/cm(2)) markedly upregulates matrix metalloproteinase 12 (MMP-12) expression and its activity in human chondrosarcoma cells. MMP-12 expression is induced in human chondrocytes during malignant transformation. However, the signaling pathway regulating MMP-12 expression and its potential role in human chondrosarcoma cell invasion and metastasis have yet to be delineated. We discovered that fluid shear stress induces the synthesis of insulin growth factor-2 (IGF-2) and vascular endothelial growth factor (VEGF) B and D, which in turn transactivate MMP-12 via PI3-K, p38 and JNK signaling pathways. IGF-2-, VEGF-B- or VEGF-D-stimulated chondrosarcoma cells display markedly higher migratory and invasive potentials in vitro, which are blocked by inhibiting MMP-12, PI3-K, p38 or JNK activity. Moreover, recombinant human MMP-12 or MMP-12 overexpression can potentiate chondrosarcoma cell invasion in vitro and the lung colonization in vivo. By reconstructing and delineating the signaling pathway regulating MMP-12 activation, potential therapeutic strategies that interfere with chondrosarcoma cell invasion may be identified.

Matrix metalloproteinase-1 facilitates MSC migration via cleavage of IGF-2/IGFBP2 complex.

PubMed

Guan, Shou P; Lam, Alan T L; Newman, Jennifer P; Chua, Kevin L M; Kok, Catherine Y L; Chong, Siao T; Chua, Melvin L K; Lam, Paula Y P

2018-01-01

The specific mechanism underlying the tumor tropism of human mesenchymal stem cells (MSCs) for cancer is not well defined. We previously showed that the migration potential of MSCs correlated with the expression and protease activity of matrix metalloproteinase (MMP)-1. Furthermore, highly tumor-tropic MSCs expressed higher levels of MMP-1 and insulin-like growth factor (IGF)-2 than poorly migrating MSCs. In this study, we examined the functional roles of IGF-2 and MMP-1 in mediating the tumor tropism of MSCs. Exogenous addition of either recombinant IGF-2 or MMP-1 could stimulate MSC migration. The correlation between IGF-2, MMP-1 expression, and MSC migration suggests that MMP-1 may play a role in regulating MSC migration via the IGF-2 signaling cascade. High concentrations of IGF binding proteins (IGFBPs) can inhibit IGF-stimulated functions by blocking its binding to its receptors and proteolysis of IGFBP is an important mechanism for the regulation of IGF signaling. We thus hypothesized that MMP-1 acts as an IGFBP2 proteinase, resulting in the cleavage of IGF-2/IGFBP2 complex and extracellular release of free IGF-2. Indeed, our results showed that conditioned media from highly migrating MSCs, which expressed high levels of MMP-1, cleaved the IGF-2/IGFBP2 complex. Taken together, these results showed that the MMP-1 secreted by highly tumor-tropic MSCs cleaved IGF-2/IGFBP2 complex. Free IGF-2 released from the complex may facilitate MSC migration toward tumor.

Expression of modulators of extracellular matrix structure after anterior cruciate ligament injury.

PubMed

Haslauer, Carla M; Proffen, Benedikt L; Johnson, Victor M; Murray, Martha M

2014-01-01

The ability of the anterior cruciate ligament (ACL) to heal after injury declines within the first 2 weeks after ACL rupture. To begin to explore the mechanism behind this finding, we quantified the expression of genes for collagen I and III, decorin, tenascin-C, and alpha smooth muscle actin, as well as matrix metalloproteinase (MMP)-1 and -13 gene expression within multiple tissues of the knee joint after ACL injury in a large animal model over a 2-week postinjury period. Gene expression of collagen I and III, decorin, and MMP-1 was highest in the synovium, whereas the highest MMP-13 gene expression levels were found in the ACL. The gene expression for collagen and decorin increased over the 2 weeks to levels approaching that in the ligament and synovium; however, no significant increase in either of the MMPs was found in the provisional scaffold. This suggests that although the ACL and synovium up-regulate both anabolic and catabolic factors, the provisional scaffold is primarily anabolic in function. The relative lack of provisional scaffold formation within the joint environment may thus be one of the key reasons for ACL degradation after injury. © 2014 by the Wound Healing Society.

The Significance of Notch1 Compared with Notch3 in High Metastasis and Poor Overall Survival in Hepatocellular Carcinoma

PubMed Central

Li, Qingjun; Sun, Wei; Zhang, Yong; Wang, Desheng; Dou, Kefeng

2013-01-01

Background The prognosis for patients with hepatocellular carcinoma (HCC) is poor, and the mechanisms underlying the development of HCC remain unclear. Notch1 and Notch3 may be involved in malignant transformation, although their roles remain unknown. Materials and Methods HCC tissues were stained with anti-Notch1 or -Notch3 antibody. The migration and invasion capacities of the cells were measured with transwell cell culture chambers. RT-PCR was used to measure the expression of Notch1 and Notch3 mRNA. Additionally, western blot analysis was used to assess the protein expression of Notch1, Notch3, CD44v6, E-cadherin, matrix metalloproteinase-2 (MMP-2), MMP-9, and urokinase-type plasminogen activator (uPA). RNA interference was used to down-regulate the expression of Notch1 and Notch3. Cell viability was assessed using MTT. Results Based on immunohistochemistry, high Notch1 expression was correlated with tumor size, tumor grade, metastasis, venous invasion and AJCC TNM stage. High Notch3 expression was only strongly correlated with metastasis, venous invasion and satellite lesions. Kaplan-Meier curves demonstrated that patients with high Notch1 or Notch3 expression were at a significantly increased risk for shortened survival time. In vitro, the down-regulation of Notch1 decreased the migration and invasion capacities of HCC cells by regulating CD44v6, E-cadherin, MMP-2, MMP-9, and uPA via the COX-2 and ERK1/2 pathways. Down-regulation of Notch3 only decreased the invasion capacity of HCC cells by regulating MMP-2 and MMP-9 via the ERK1/2 pathway. Conclusions Based on the migration and invasion of HCC, we hypothesize that targeting Notch1 may be more useful than Notch3 for designing novel preventive and therapeutic strategies for HCC in the near future. PMID:23468978

Extracellular redox state regulates features associated with prostate cancer cell invasion.

PubMed

Chaiswing, Luksana; Zhong, Weixiong; Cullen, Joseph J; Oberley, Larry W; Oberley, Terry D

2008-07-15

We have examined the possible role of extracellular reduction-oxidation (redox) state in regulation of biological/biochemical features associated with prostate cancer cell invasion. DU145, PC-3, and RWPE1-derived human prostate cancer (WPE1-NB26) cell lines were used for the present in vitro analysis. Increasing levels of nitric oxide using S-nitroso-N-acetylpenicillamine resulted in a decrease in cell invasion ability, whereas increasing levels of extracellular superoxide radical (O(2)(*-)) using xanthine/xanthine oxidase resulted in an increase in cell invasion ability in these three cell lines. WPE1-NB26 cells exhibited an increased glutathione/glutathione disulfide ratio in the medium in comparison with RWPE1 cells (immortalized but nonmalignant prostate epithelial cells), suggesting an alteration of extracellular redox state of WPE1-NB26 cells. We hypothesized that O(2)(*-) production at or near the plasma membrane or in the adjacent extracellular matrix at least partially regulated prostate cancer cell invasion. Using adenovirus-mediated extracellular superoxide dismutase (EC-SOD) gene transduction to enzymatically decrease O(2)(*-) levels, we showed that in the presence of heparin, adenovirus EC-SOD gene transduction resulted in an increase in the expression of EC-SOD outside the cells with resultant inhibition of cell invasion ability. This inhibition correlated with reduced metalloproteinase [matrix metalloproteinase (MMP) 2/membrane type 1-MMP] activities and increased levels of extracellular nitrite. Our results suggest a prominent role of extracellular redox status in regulation of cell invasion, which may provide opportunities for therapeutic interventions.

Factors of transforming growth factor beta signalling are co-regulated in human hepatocellular carcinoma.

PubMed

Longerich, Thomas; Breuhahn, Kai; Odenthal, Margarete; Petmecky, Katharina; Schirmacher, Peter

2004-12-01

Transforming growth factor beta (TGFbeta) is a central mitoinhibitory factor for epithelial cells, and alterations of TGFbeta signalling have been demonstrated in many different human cancers. We have analysed human hepatocellular carcinomas (HCCs) for potential pro-tumourigenic alterations in regard to expression of Smad4 and mutations and expression changes of the pro-oncogenic transcriptional co-repressors Ski and SnoN, as well as mRNA levels of matrix metalloproteinase-2 (MMP2), which is transcriptionally regulated by TGFbeta. Smad4 mRNA was detected in all HCCs; while, using immunohistology, loss of Smad4 expression was found in 10% of HCCs. Neither mutations in the transformation-relevant sequences nor significant pro-tumourigenic expression changes of the Ski and SnoN genes were detected. In HCC cell lines, expression of both genes was regulated, potentially involving phosphorylation. Ski showed a distinct nuclear speckled pattern, indicating recruitment to active transcription complexes. MMP2 mRNA levels were increased in 19% of HCCs, whereas MMP2 mRNA was not detectable in HCC cell lines, suggesting that MMP2 was derived only from tumour stroma cells. Transcript levels of Smad4, Ski, SnoN and MMP2 correlated well. These data argue against a significant role of Ski and SnoN in human hepatocarcinogenesis and suggest that, in the majority of HCCs, the analysed factors are co-regulated by an upstream mechanism, potentially by TGFbeta itself.

Changes of MMP-1 and collagen type Ialpha1 by UVA, UVB and IRA are differentially regulated by Trx-1.

PubMed

Buechner, Nicole; Schroeder, Peter; Jakob, Sascha; Kunze, Kerstin; Maresch, Tanja; Calles, Christian; Krutmann, Jean; Haendeler, Judith

2008-07-01

Exposure of human skin to solar radiation, which includes ultraviolet (UV) radiation (UVA and UVB) visible light and infrared radiation, induces skin aging. The effects of light have been attributed to irradiation-induced reactive oxygen species (ROS) formation, but the specific signaling pathways are not well understood. Detrimental effects of solar radiation are dermal diseases and photoaging. Exposure of cultured human dermal fibroblasts to UVA, UVB or IRA increased ROS formation in vitro. One important redox regulator is the oxidoreductase thioredoxin-1 (Trx). Trx is ubiquitously expressed and has anti-oxidative and anti-apoptotic properties. Besides its function to reduce H(2)O(2), Trx binds to and regulates transcription factors. The aim of this study was to investigate whether Trx influences the regulation of MMP-1 and collagen Ialpha1 by UVA, UVB and IRA. We irradiated human dermal fibroblasts with UVA, UVB and IRA. UVA, UVB and IRA upregulated MMP-1 expression. Trx inhibited UVA-induced MMP-1 upregulation in a NFkappaB dependent manner. UVA, UVB and IRA reduced collagen Ialpha1 expression. Incubation with Trx inhibited the effects of UVB and IRA on collagen Ialpha1 expression. In conclusion, MMP-1 and collagen Ialpha1, which play important roles in aging processes, seems to be regulated by different transcriptional mechanisms and Trx can only influence distinct signaling pathways induced by UVA, UVB and probably IRA. Thus, Trx may serve as an important contributor to an "anti-aging therapeutic cocktail".

Matrix Metalloproteinase (MMP) 9 Transcription in Mouse Brain Induced by Fear Learning*

PubMed Central

Ganguly, Krishnendu; Rejmak, Emilia; Mikosz, Marta; Nikolaev, Evgeni; Knapska, Ewelina; Kaczmarek, Leszek

2013-01-01

Memory formation requires learning-based molecular and structural changes in neurons, whereas matrix metalloproteinase (MMP) 9 is involved in the synaptic plasticity by cleaving extracellular matrix proteins and, thus, is associated with learning processes in the mammalian brain. Because the mechanisms of MMP-9 transcription in the brain are poorly understood, this study aimed to elucidate regulation of MMP-9 gene expression in the mouse brain after fear learning. We show here that contextual fear conditioning markedly increases MMP-9 transcription, followed by enhanced enzymatic levels in the three major brain structures implicated in fear learning, i.e. the amygdala, hippocampus, and prefrontal cortex. To reveal the role of AP-1 transcription factor in MMP-9 gene expression, we have used reporter gene constructs with specifically mutated AP-1 gene promoter sites. The constructs were introduced into the medial prefrontal cortex of neonatal mouse pups by electroporation, and the regulation of MMP-9 transcription was studied after contextual fear conditioning in the adult animals. Specifically, −42/-50- and −478/-486-bp AP-1 binding motifs of the mouse MMP-9 promoter sequence have been found to play a major role in MMP-9 gene activation. Furthermore, increases in MMP-9 gene promoter binding by the AP-1 transcription factor proteins c-Fos and c-Jun have been demonstrated in all three brain structures under investigation. Hence, our results suggest that AP-1 acts as a positive regulator of MMP-9 transcription in the brain following fear learning. PMID:23720741

Matrix metalloproteinase (MMP) 9 transcription in mouse brain induced by fear learning.

PubMed

Ganguly, Krishnendu; Rejmak, Emilia; Mikosz, Marta; Nikolaev, Evgeni; Knapska, Ewelina; Kaczmarek, Leszek

2013-07-19

Memory formation requires learning-based molecular and structural changes in neurons, whereas matrix metalloproteinase (MMP) 9 is involved in the synaptic plasticity by cleaving extracellular matrix proteins and, thus, is associated with learning processes in the mammalian brain. Because the mechanisms of MMP-9 transcription in the brain are poorly understood, this study aimed to elucidate regulation of MMP-9 gene expression in the mouse brain after fear learning. We show here that contextual fear conditioning markedly increases MMP-9 transcription, followed by enhanced enzymatic levels in the three major brain structures implicated in fear learning, i.e. the amygdala, hippocampus, and prefrontal cortex. To reveal the role of AP-1 transcription factor in MMP-9 gene expression, we have used reporter gene constructs with specifically mutated AP-1 gene promoter sites. The constructs were introduced into the medial prefrontal cortex of neonatal mouse pups by electroporation, and the regulation of MMP-9 transcription was studied after contextual fear conditioning in the adult animals. Specifically, -42/-50- and -478/-486-bp AP-1 binding motifs of the mouse MMP-9 promoter sequence have been found to play a major role in MMP-9 gene activation. Furthermore, increases in MMP-9 gene promoter binding by the AP-1 transcription factor proteins c-Fos and c-Jun have been demonstrated in all three brain structures under investigation. Hence, our results suggest that AP-1 acts as a positive regulator of MMP-9 transcription in the brain following fear learning.

Human disc degeneration is associated with increased MMP 7 expression.

PubMed

Le Maitre, C L; Freemont, A J; Hoyland, J A

2006-01-01

During intervertebral disc (IVD) degeneration, normal matrix synthesis decreases and degradation of disc matrix increases. A number of proteases that are increased during disc degeneration are thought to be involved in its pathogenesis. Matrix metalloproteinase 7 (MMP 7) (Matrilysin, PUMP-1) is known to cleave the major matrix molecules found within the IVD, i.e., the proteoglycan aggrecan and collagen type II. To date, however, it is not known how its expression changes with degeneration or its exact location. We investigated the localization of MMP 7 in human, histologically graded, nondegenerate, degenerated and prolapsed discs to ascertain whether MMP 7 is up-regulated during disc degeneration. Samples of human IVD tissue were fixed in neutral buffered formalin, embedded in paraffin, and sections stained with hematoxylin and eosin to score the degree of morphological degeneration. Immunohistochemistry was performed to localize MMP 7 in 41 human IVDs with varying degrees of degeneration. We found that the chondrocyte-like cells of the nucleus pulposus and inner annulus fibrosus were MMP 7 immunopositive; little immunopositivity was observed in the outer annulus. Nondegenerate discs showed few immunopositive cells. A significant increase in the proportion of MMP 7 immunopositive cells was seen in the nucleus pulposus of discs classified as showing intermediate levels of degeneration and a further increase was seen in discs with severe degeneration. Prolapsed discs showed more MMP 7 immunopositive cells compared to nondegenerated discs, but fewer than those seen in cases of severe degeneration.

[Effect of LPXN Overexpression on the Proliferation, Adhesion and Invasion of THP-1 Cells and Its Mechamisms].

PubMed

Dai, Hai-Ping; Zhu, Guo-Hua; Wu, Li-Li; Wang, Qian; Yao, Hong; Wang, Qin-Rong; Wen, Li-Jun; Qiu, Hui-Ying; Shen, Qun; Chen, Su-Ning; Wu, De-Pei

2017-06-01

To explore the effect of LPXN overexpression on the proliferation, adhesion and invasion of THP-1 cells and its possible mechanism. A THP-1 cell line with stable overexpression of LPXN was constucted by using a lentivirus method, CCK-8 was used to detect the proliferation of cells, adhesion test was used to evaluate adhesion ablity of cells to Fn. Transwell assay was used to detect the change of invasion capability. Western blot was used to detect expression of LPXN, ERK, pERK and integrin α4, α5, β1, the Gelatin zymography was applied to detect activity of MMP2/MMP9 secreted by the THP-1 cells. Successful establishment of THP-1 cells with LPXN overexpression (THP-1 LPXN) was confirmed with Western blot. THP-1 LPXN cells were shown to proliferate faster than the control THP-1 vector cells. Adhesion to Fn and expression of ERK, integrin α4, α5 and β1 in the THP-1 LPXN cells were higher than that in the control cells. Invasion across matrigel and enhanced activity of MMP2 could be detected both in the THP-1 LPXN cells as compared with the control cells. Ectopically ovexpression of LPXN may promote proliferation of THP-1 cells through up-regulation of ERK; promote adhesion of THP-1 cells through up-regulating the integrin α4/β1 as well as integrin α5/β1 complex; promote invasion of THP-1 cells through activating MMP2.

Mangiferin regulates proliferation and apoptosis in glioma cells by induction of microRNA-15b and inhibition of MMP-9 expression.

PubMed

Xiao, Jinsong; Liu, Li; Zhong, Zian; Xiao, Cheng; Zhang, Junjian

2015-06-01

Mangiferin, a flavonoid extracted from the leaves of the Anacardiaceae plant, the mango tree, has physiological activity and pharmacological effects in many aspects. The present study aimed to clarify the effect of mangiferin on proliferation and apoptosis of glioma cells and the mechanism of these curative effects of mangiferin. In this experiment, we detected the proliferation using 3-(4,5-dimethylthylthiazol-2-yl)-2,5 diphenyltetrazolium bromide (MTT) assay. Then, cell apoptosis of U87 glioma cells was measured with the Annexin V-FITC/propidium iodide (PI) apoptosis detection kit, DAPI staining assay and the caspase-3 and caspase-9 activity assay kit. Next, quantitative real-time PCR and gelatin zymography were used to analyze the expression of microRNA-15b (miR-15b) and matrix metalloproteinase-9 (MMP-9), respectively. MMP-9 agonist, miR-15b mimics and anti-miR-15b mimics were added to the U87 glioma cells for elucidating the mechanisms involved in the curative effects of mangiferin. In the present study, mangiferin notably restrained the proliferation and increased the apoptosis of the U87 glioma cells. Meanwhile, mangiferin specifically promoted the expression of miR-15b and suppressed the level of MMP-9 in the U87 glioma cells. miR-15b regulated the expression of MMP-9 in the U87 glioma cells. MMP-9 agonist and anti-miR‑15b reduced the curative effects of mangiferin in the U87 glioma cells. In summary, mangiferin regulates proliferation and apoptosis in glioma cells by induction of miR-15b and inhibition of MMP-9 expression.

MMP-7 expression may influence the rate of distant recurrences and disease-specific survival in HPV-positive oropharyngeal squamous cell carcinoma.

PubMed

Vento, Seija I; Jouhi, Lauri; Mohamed, Hesham; Haglund, Caj; Mäkitie, Antti A; Atula, Timo; Hagström, Jaana; Mäkinen, Laura K

2018-05-02

The objective of this study was to determine if matrix metalloproteinase-7 (MMP-7) expression is related to human papilloma virus (HPV) status, clinical parameters, and outcome in oropharyngeal squamous cell carcinoma (OPSCC). Tumor tissue specimens from 201 OPSCC patients treated with curative intent were available for immunohistochemistry, and the samples were stained with monoclonal MMP-7 antibody. All the patients were followed up at least 3 years or until death. MMP-7 expression did not differ between HPV-positive and HPV-negative patients. MMP-7 was not prognostic among patients with HPV-negative OPSCC. In the HPV-positive subgroup, patients with moderate, high, or very high MMP-7 expression had significantly worse 5-year disease-specific survival (DSS) (56.6%) than patients with absent, or low MMP-7 expression (77.2%), and MMP-7 expression appeared as a prognostic factor in the multivariate analysis. In addition, among HPV-positive OPSCC with moderate, high, or very high MMP-7 expression, the 5-year distant recurrence-free survival was significantly lower (69.6%) than in those who had low or absent MMP-7 expression (97.5%). Our results suggest that among HPV-positive OPSCC patients, high MMP-7 expression is related to worse 5-year DSS and increased rate of distant recurrences.

Aryl hydrocarbon receptor-dependent up-regulation of the heterodimeric amino acid transporter LAT1 (SLC7A5)/CD98hc (SLC3A2) by diesel exhaust particle extract in human bronchial epithelial cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Le Vee, Marc; Jouan, Elodie; Lecureur, Valérie

The heterodimeric L-type amino acid transporter (LAT) 1/CD98hc is overexpressed in lung cancers with a poor prognosis factor. Factors that contribute to LAT1/CD98hc overexpression in lung cells remain however to be determined, but the implication of atmospheric pollution can be suspected. The present study was therefore designed to analyze the effects of diesel exhaust particle (DEP) extract (DEPe) on LAT1/CD98hc expression in bronchial epithelial BEAS-2B cells. Exposure to DEPe up-regulated LAT1 and CD98hc mRNA levels in a concentration-dependent manner, with DEPe EC{sub 50} values (around 0.2 μg/mL) relevant to environmental situations. DEPe concomitantly induced LAT1/CD98hc protein expression and LAT1-mediated leucinemore » accumulation in BEAS-2B cells. Inhibition of the aryl hydrocarbon receptor (AhR) pathway through the use of a chemical AhR antagonist or the siRNA-mediated silencing of AhR expression was next found to prevent DEPe-mediated induction of LAT1/CD98hc, indicating that this regulation depends on AhR, known to be activated by major chemical DEP components like polycyclic aromatic hydrocarbons. DEPe exposure was finally shown to induce mRNA expression and activity of matrix metalloproteinase (MMP)-2 in BEAS-2B cells, in a CD98hc/focal adhesion kinase (FAK)/extracellular regulated kinase (ERK) manner, thus suggesting that DEPe-mediated induction of CD98hc triggers activation of the integrin/FAK/ERK signaling pathway known to be involved in MMP-2 regulation. Taken together, these data demonstrate that exposure to DEPe induces functional overexpression of the amino acid transporter LAT1/CD98hc in lung cells. Such a regulation may participate to pulmonary carcinogenic effects of DEPs, owing to the well-documented contribution of LAT1 and CD98hc to cancer development. - Highlights: • The amino acid transporter LAT1/CD98hc is up-regulated in DEPe-treated lung cells. • The aryl hydrocarbon receptor is involved in DEPe-triggered induction of LAT1/CD98hc. • DEPe enhances LAT1-mediated accumulation of leucine in lung cells. • DEPe increases MMP-2 expression and activity in a CD98hc/FAK/ERK-dependent manner.« less

DOE Office of Scientific and Technical Information (OSTI.GOV)

Otani, Satoshi; Kakinuma, Sei, E-mail: skakinuma.gast@tmd.ac.jp; Department for Liver Disease Control, Tokyo Medical and Dental University, Tokyo

Fetal hepatic stem/progenitor cells, called hepatoblasts, play central roles in liver development; however, the molecular mechanisms regulating the phenotype of these cells have not been completely elucidated. Matrix metalloproteinase (MMP)-14 is a type I transmembrane proteinase regulating pericellular proteolysis of the extracellular matrix and is essential for the activation of several MMPs and cytokines. However, the physiological functions of MMP-14 in liver development are unknown. Here we describe a functional role for MMP-14 in hepatic and biliary differentiation of mouse hepatoblasts. MMP-14 was upregulated in cells around the portal vein in perinatal stage liver. Formation of bile duct-like structures inmore » MMP-14–deficient livers was significantly delayed compared with wild-type livers in vivo. In vitro biliary differentiation assays showed that formation of cholangiocytic cysts derived from MMP-14–deficient hepatoblasts was completely impaired, and that overexpression of MMP-14 in hepatoblasts promoted the formation of bile duct-like cysts. In contrast, the expression of molecules associated with metabolic functions in hepatocytes, including hepatic nuclear factor 4α and tryptophan 2,3-dioxygenase, were significantly increased in MMP-14–deficient livers. Expression of the epidermal growth factor receptor and phosphorylation of mitogen-activated protein kinases were significantly upregulated in MMP-14–deficient livers. We demonstrate that MMP-14–mediated signaling in fetal hepatic progenitor cells promotes biliary luminal formation around the portal vein and negatively controls the maturation of hepatocytes. - Highlights: • Loss of MMP-14 delayed formation of bile duct-like structures in perinatal liver. • Overexpression of MMP-14 in hepatobalsts promoted the biliary formation in vitro. • Loss of MMP-14 promoted hepatocyte maturation of hepatoblasts in vivo. • MMP-14–mediated signaling regulates terminal differentiation of hepatoblasts.« less

Matrix Metalloproteinases Are Differentially Regulated and Responsive to Compression Therapy in a Red Duroc Model of Hypertrophic Scar.

PubMed

Travis, Taryn E; Ghassemi, Pejhman; Prindeze, Nicholas J; Moffatt, Lauren T; Carney, Bonnie C; Alkhalil, Abdulnaser; Ramella-Roman, Jessica C; Shupp, Jeffrey W

2018-01-01

Objective: Proteins of the matrix metalloproteinases family play a vital role in extracellular matrix maintenance and basic physiological processes in tissue homeostasis. The function and activities of matrix metalloproteinases in response to compression therapies have yet to be defined. Here, a swine model of hypertrophic scar was used to profile the transcription of all known 26 matrix metalloproteinases in scars treated with a precise compression dose. Methods: Full-thickness excisional wounds were created. Wounds underwent healing and scar formation. A subset of scars underwent 2 weeks of compression therapy. Biopsy specimens were preserved, and microarrays, reverse transcription-polymerase chain reaction, Western blotting, and immunohistochemistry were performed to characterize the transcription and expression of various matrix metalloproteinase family members. Results: Microarray results showed that 13 of the known 26 matrix metalloproteinases were differentially transcribed in wounds relative to the preinjury skin. The predominant upregulation of these matrix metalloproteinases during early wound-healing stages declined gradually in later stages of wound healing. The use of compression therapy reduced this decline in 10 of the 13 differentially regulated matrix metalloproteinases. Further investigation of MMP7 using reverse transcription-polymerase chain reaction confirmed the effect of compression on transcript levels. Assessment of MMP7 at the protein level using Western blotting and immunohistochemistry was concordant. Conclusions: In a swine model of hypertrophic scar, the application of compression to hypertrophic scar attenuated a trend of decreasing levels of matrix metalloproteinases during the process of hypertrophic wound healing, including MMP7, whose enzyme regulation was confirmed at the protein level.

Apoptosis Signal-Regulating Kinase 1 Is Involved in Brain-Derived Neurotrophic Factor (BDNF)-Enhanced Cell Motility and Matrix Metalloproteinase 1 Expression in Human Chondrosarcoma Cells

PubMed Central

Lin, Chih-Yang; Chang, Sunny Li-Yun; Fong, Yi-Chin; Hsu, Chin-Jung; Tang, Chih-Hsin

2013-01-01

Chondrosarcoma is the primary malignancy of bone that is characterized by a potent capacity to invade locally and cause distant metastasis, and is therefore associated with poor prognoses. Chondrosarcoma further shows a predilection for metastasis to the lungs. The brain-derived neurotrophic factor (BDNF) is a small molecule in the neurotrophin family of growth factors that is associated with the disease status and outcome of cancers. However, the effect of BDNF on cell motility in human chondrosarcoma cells is mostly unknown. Here, we found that human chondrosarcoma cell lines had significantly higher cell motility and BDNF expression compared to normal chondrocytes. We also found that BDNF increased cell motility and expression of matrix metalloproteinase-1 (MMP-1) in human chondrosarcoma cells. BDNF-mediated cell motility and MMP-1 up-regulation were attenuated by Trk inhibitor (K252a), ASK1 inhibitor (thioredoxin), JNK inhibitor (SP600125), and p38 inhibitor (SB203580). Furthermore, BDNF also promoted Sp1 activation. Our results indicate that BDNF enhances the migration and invasion activity of chondrosarcoma cells by increasing MMP-1 expression through a signal transduction pathway that involves the TrkB receptor, ASK1, JNK/p38, and Sp1. BDNF thus represents a promising new target for treating chondrosarcoma metastasis. PMID:23892595

Metalloproteinases and atherothrombosis: MMP-10 mediates vascular remodeling promoted by inflammatory stimuli.

PubMed

Rodriguez, Jose A; Orbe, Josune; Martinez de Lizarrondo, Sara; Calvayrac, Olivier; Rodriguez, Cristina; Martinez-Gonzalez, Jose; Paramo, Jose A

2008-01-01

Atherosclerosis is the common pathophysiological substrate of ischemic vascular diseases and their thrombotic complications. The unbalance between matrix metalloproteinases (MMPs) and their inhibitors (TIMPs) has been hypothesized to be involved in the growth, destabilization, and eventual rupture of atherosclerotic lesions. Different MMPs have been assigned relevant roles in the pathology of vascular diseases and MMP-10 (stromelysin-2) has been involved in vascular development and atherogenesis. This article examines the pathophysiological role of MMPs, particularly MMP-10, in the onset and progression of vascular diseases and their regulation by pro-inflammatory stimuli. MMP-10 over-expression has been shown to compromise vascular integrity and it has been associated with aortic aneurysms. MMP-10 is induced by C-reactive protein in endothelial cells, and it is over-expressed in atherosclerotic lesions. Additionally, higher MMP-10 serum levels are associated with inflammatory markers, increased carotid intima-media thickness and the presence of atherosclerotic plaques. We have cloned the promoter region of the MMP-10 gene and studied the effect of inflammatory stimuli on MMP-10 transcriptional regulation, providing evidences further supporting the involvement of MMP-10 in the pathophysiology of atherothrombosis.

LncRNA FAS-AS1 promotes the degradation of extracellular matrix of cartilage in osteoarthritis.

PubMed

Zhu, J-K; He, T-D; Wei, Z-X; Wang, Y-M

2018-05-01

To investigate the expression of long non-coding RNA (lncRNA) FAS-AS1 in osteoarthritis cartilage and to explore its effect on articular cartilage cells. A total of 20 tissue samples of primary knee joint osteoarthritis and 20 tissue samples of knee joint cartilage after traumatic amputation were collected. Fluorescence quantitative polymerase chain reaction (PCR) was performed to detect the expression of FAS-AS1, MMP1, MMP13, and COL2A1 in cartilage. FAS-AS1 small interfering RNA (siRNA) was transfected to chondrocytes transiently to observe its effects on proliferation, apoptosis of chondrocytes, and the expressions of MMP1, MMP13, and COL2A1. The expressions of FAS-AS1, MMP1, and MMP13 in osteoarthritis tissues increased significantly, while COL2A1 presented a low expression. Reducing the expression of FAS-AS1 inhibited cell apoptosis and promote cell proliferation. Additionally, in vitro experiments showed that low expression of FAS-AS1 decreased the expressions of MMP1 and MMP13, but increased the expression of COL2A1. The expression of FAS-AS1 was increased in osteoarthritis, and FAS-AS1 could be involved in the development of the disease by regulating the proliferation, apoptosis of chondrocytes and promoting the degradation of extracellular matrix.

Messenger RNA for membrane-type 2 matrix metalloproteinase, MT2-MMP, is expressed in human placenta of first trimester.

PubMed

Bjørn, S F; Hastrup, N; Larsen, J F; Lund, L R; Pyke, C

2000-01-01

An intimately regulated cell surface activation of matrix metalloproteinases (MMPs) is believed to be of critical importance for the control of trophoblast invasion. A histological investigation of the expression and localization of three different MMPs, the membrane-type matrix metalloproteinases 1 and 2 (MT1-MMP, MT2-MMP) and matrix metalloproteinase 2 (MMP-2/gelatinase A) was performed by in situ hybridization on consecutive sections from human placentae of first trimester pregnancies. Cytokeratin immunostaining identified trophoblast cells. Both normal and tubal implantation sites were studied. We observed a high degree of coexpression of MT2-MMP, MT1-MMP and MMP-2 mRNAs in single extravillous cytotrophoblasts that had invaded the endometrium and tubal wall. Furthermore, mRNAs for all three genes were also seen in cytotrophoblasts of cell islands. In contrast to this coexpression pattern, MT2-MMP expression was absent from cell columns and decidual cells, in which signals for MT1-MMP and MMP-2 mRNAs were seen. The present data on the cellular expression of MT2-MMP mRNA in placenta extend our knowledge of the proteolytic events that take place during early pregnancy. The data suggest that MT2-MMP, capable of activating MMP-2 in vitro, is involved in the invasion of extravillous cytotrophoblast, possibly related to the physiological activation of MMP-2. Copyright 2000 Harcourt Publishers Ltd.

Transcription Factor SOX5 Promotes the Migration and Invasion of Fibroblast-Like Synoviocytes in Part by Regulating MMP-9 Expression in Collagen-Induced Arthritis.

PubMed

Shi, Yumeng; Wu, Qin; Xuan, Wenhua; Feng, Xiaoke; Wang, Fang; Tsao, Betty P; Zhang, Miaojia; Tan, Wenfeng

2018-01-01

Fibroblast-like synoviocytes (FLS) exhibit a unique aggressive phenotype in rheumatoid arthritis (RA). Increased FLS migration and subsequent invasion of the extracellular matrix are essential to joint destruction in RA. Our previous research reported that transcription factor SOX5 was highly expressed in RA-FLS. Here, the effects of SOX5 in RA-FLS migration and invasion will be investigated. The migration and invasion of RA-FLS were evaluated using a transwell chamber assay. The expression of several potential SOX5-targeted genes, including matrix metalloproteinases (MMP-1, 2, 3 and 9), chemokines (CCL4, CCL2, CCR5 and CCR2), and pro-inflammatory cytokines (TNF-α and IL-6), were examined in RA-FLS using SOX5 gain- and loss-of-function study. The molecular mechanisms of SOX5-mediated MMP-9 expressions were assayed by luciferase reporter gene and chromatin immunoprecipitation (ChIP) studies. The in vivo effect of SOX5 on FLS migration and invasion was examined using collagen-induced arthritis (CIA) in DBA/1J mice. Knockdown SOX5 decreased lamellipodium formation, migration, and invasion of RA-FLS. The expression of MMP-9 was the only gene tested to be concomitantly affected by silencing or overexpressing SOX5. ChIP assay revealed that SOX5 was bound to the MMP-9 promoter in RA-FLS. The overexpression of SOX5 markedly enhanced the MMP-9 promoter activity, and specific deletion of a putative SOX5-binding site in MMP-9 promoter diminished this promoter-driven transcription in FLS. Locally knocked down SOX5 inhibited MMP-9 expression in the joint tissue and reduced pannus migration and invasion into the cartilage in CIA mice. SOX5 plays a novel role in mediating migration and invasion of FLS in part by regulating MMP-9 expression in RA.

IGF-1R Regulates the Extracellular Level of Active MMP-2, Pathological Neovascularization, and Functionality in Retinas of OIR Mouse Model.

PubMed

Lorenc, Valeria E; Subirada Caldarone, Paula V; Paz, María C; Ferrer, Darío G; Luna, José D; Chiabrando, Gustavo A; Sánchez, María C

2018-02-01

In ischemic proliferative diseases such as retinopathies, persistent hypoxia leads to the release of numerous neovascular factors that participate in the formation of abnormal vessels and eventually cause blindness. The upregulation and activation of metalloproteinases (MMP-2 and MMP-9) represent a final common pathway in this process. Although many regulators of the neovascular process have been identified, the complete role of the insulin-like growth factor 1 (IGF-1) and its receptor (IGF-1R) appears to be significantly more complex. In this study, we used an oxygen-induced retinopathy (OIR) mouse model as well as an in vitro model of hypoxia to study the role of MMP-2 derived from Müller glial cells (MGCs) and its relation with the IGF-1/IGF-1R system. We demonstrated that MMP-2 protein expression increased in P17 OIR mice, which coincided with the active phase of the neovascular process. Also, glutamine synthetase (GS)-positive cells were also positive for MMP-2, whereas IGF-1R was expressed by GFAP-positive cells, indicating that both proteins were expressed in MGCs. In addition, in the OIR model a single intravitreal injection of the IGF-1R blocking antibody (αIR3) administered at P12 effectively prevented pathologic neovascularization, accelerated physiological revascularization, and improved retinal functionality at P17. Finally, in MGC supernatants, the blocking antibody abolished the IGF-1 effect on active MMP-2 under normoxic and hypoxic conditions without affecting the extracellular levels of pro-MMP-2. These results demonstrate, for the first time, that the IGF-1/IGF-1R system regulates active MMP-2 levels in MGCs, thus contributing to MEC remodeling during the retinal neovascular process.

PARP-1 depletion in combination with carbon ion exposure significantly reduces MMPs activity and overall increases TIMPs expression in cultured HeLa cells.

PubMed

Ghorai, Atanu; Sarma, Asitikantha; Chowdhury, Priyanka; Ghosh, Utpal

2016-09-22

Hadron therapy is an innovative technique where cancer cells are precisely killed leaving surrounding healthy cells least affected by high linear energy transfer (LET) radiation like carbon ion beam. Anti-metastatic effect of carbon ion exposure attracts investigators into the field of hadron biology, although details remain poor. Poly(ADP-ribose) polymerase-1 (PARP-1) inhibitors are well-known radiosensitizer and several PARP-1 inhibitors are in clinical trial. Our previous studies showed that PARP-1 depletion makes the cells more radiosensitive towards carbon ion than gamma. The purpose of the present study was to investigate combining effects of PARP-1 inhibition with carbon ion exposure to control metastatic properties in HeLa cells. Activities of matrix metalloproteinases-2, 9 (MMP-2, MMP-9) were measured using the gelatin zymography after 85 MeV carbon ion exposure or gamma irradiation (0- 4 Gy) to compare metastatic potential between PARP-1 knock down (HsiI) and control cells (H-vector - HeLa transfected with vector without shRNA construct). Expression of MMP-2, MMP-9, tissue inhibitor of MMPs such as TIMP-1, TIMP-2 and TIMP-3 were checked by immunofluorescence and western blot. Cell death by trypan blue, apoptosis and autophagy induction were studied after carbon ion exposure in each cell-type. The data was analyzed using one way ANOVA and 2-tailed paired-samples T-test. PARP-1 silencing significantly reduced MMP-2 and MMP-9 activities and carbon ion exposure further diminished their activities to less than 3 % of control H-vector. On the contrary, gamma radiation enhanced both MMP-2 and MMP-9 activities in H-vector but not in HsiI cells. The expression of MMP-2 and MMP-9 in H-vector and HsiI showed different pattern after carbon ion exposure. All three TIMPs were increased in HsiI, whereas only TIMP-1 was up-regulated in H-vector after irradiation. Notably, the expressions of all TIMPs were significantly higher in HsiI than H-vector at 4 Gy. Apoptosis was the predominant mode of cell death and no autophagic death was observed. Our study demonstrates for the first time that PARP-1 inhibition in combination with carbon ion synergistically decreases MMPs activity along with overall increase of TIMPs. These data open up the possibilities of improvement of carbon ion therapy with PARP-1 inhibition to control highly metastatic cancers.

Chymotrypsin C (Caldecrin) Is Associated with Enamel Development

PubMed Central

Lacruz, R.S.; Smith, C.E.; Smith, S.M.; Hu, P.; Bringas, P.; Sahin-Tóth, M.; Moradian-Oldak, J.; Paine, M.L.

2011-01-01

Two main proteases cleave enamel extracellular matrix proteins during amelogenesis. Matrix metalloprotease-20 (Mmp20) is the predominant enzyme expressed during the secretory stage, while kallikrein-related peptidase-4 (Klk4) is predominantly expressed during maturation. Mutations to both Mmp20 and Klk4 result in abnormal enamel phenotypes. During a recent whole-genome microarray analysis of rat incisor enamel organ cells derived from the secretory and maturation stages of amelogenesis, the serine protease chymotrypsin C (caldecrin, Ctrc) was identified as significantly up-regulated (> 11-fold) during enamel maturation. Prior reports indicate that Ctrc expression is pancreas-specific, albeit low levels were also noted in brain. We here report on the expression of Ctrc in the enamel organ. Quantitative PCR (qPCR) and Western blot analysis were used to confirm the expression of Ctrc in the developing enamel organ. The expression profile of Ctrc is similar to that of Klk4, increasing markedly during the maturation stage relative to the secretory stage, although levels of Ctrc mRNA are lower than for Klk4. The discovery of a new serine protease possibly involved in enamel development has important implications for our understanding of the factors that regulate enamel biomineralization. PMID:21828354

Inhibitory effect of midazolam on MMP-9, MMP-1 and MMP-13 expression in PMA-stimulated human chondrocytes via recovery of NF-κB signaling.

PubMed

Wang, Jen-Jui; Huan, Steven Kuan-Hua; Hsieh, Kuo-Hsien; Chou, Hsiu-Chu; Hsiao, George; Jayakumar, Thanasekaran; Sheu, Joen-Rong

2013-04-20

Midazolam, a benzodiazepine, has a hypnotic effect and is widely used as an intravenous sedative. Past studies have clearly established that midazolam has beneficial effects in attenuating ischemia-reperfusion injury more than other currently used sedative drugs. However, the role of midazolam on chondroprotection via inhibition of matrix metalloproteinases (MMPs) is warrant investigation. The aim of this study was to examine the mechanisms of action of midazolam on MMP expression via nuclear factor κB (NF-κB) signaling in activated chondrosarcoma cells maintained in vitro. Chondrocytes, SW1353 cells, were stimulated with phorbol 12-myristate 13-acetate (PMA) in the absence or presence of various concentrations of midazolam (5-20 µM). Release of MMP-9 into the culture media was determined by gelatin zymography. The expressions of MMP-1, MMP-9 and MMP-13, phosphorylation of extracellular signal-regulated kinase (ERK), p38 mitogen-activated protein kinases and degradation of IκB-α were determined by western blotting assay. Midazolam significantly down-regulated PMA-induced MMP-9 protein expression at concentrations of 5, 10 and 20 µM, the values were 1.95 ±0.09 (p < 0.01), 1.71 ±0.12 (p < 0.01) and 1.35 ±0.20 (p < 0.001), respectively. At concentrations of 5, 10 and 20 µM, it was significantly inhibited the PMA-induced expressions of MMP-1 (2.27 ±0.10, 1.98 ±0.11 and 1.56 ±0.15; p < 0.001) and MMP-13 (0.89 ±0.04, 0.81 ±0.07, and 0.74 ±0.09; p < 0.001), respectively. Midazolam at concentrations of 10 and 20 µM for 15 min significantly reversed the rate of degradation (0.895 ±0.051; p < 0.05 and 0.926 ±0.060; p < 0.01, respectively) of IκB-α in PMA-chondrocyte cells. In addition, this sedative drug inhibited PMA-induced levels of phos-ERK (1.243 ±0.12, 1.108 ±0.16 and 0.903 ±0.19, respectively) and phos-p38 (1.146 ±0.10, 1.063 ±0.13 and 0.946 ±0.18, at concentrations of (5, 10 and 20 µM), respectively. These results are important for understanding the mechanism of midazolam in inhibiting PMA-induced MMP expression through the signaling pathways of either NF-κB or ERK/p38 MAPKs down-regulation.

Methanol extract of Codium fragile inhibits tumor necrosis factor-α-induced matrix metalloproteinase-9 and invasiveness of MDA-MB-231 cells by suppressing nuclear factor-κB activation.

PubMed

Dilshara, Matharage Gayani; Jayasooriya, Rajapaksha Gedara Prasad Tharanga; Kang, Chang-Hee; Choi, Yung-Hyun; Kim, Gi-Young

2016-06-01

To evaluate whether the methanol extract of Codium fragile (MECF) regulates tumor necrosis factor-α (TNF-α)-induced invasion of human breast cancer MDA-MB-231 cells by suppressing matrix metalloproteinase-9 (MMP-9). Reverse transcription-polymerase chain reaction (RT-PCR) and western blot analysis were performed to analyze the expression of MMP-9 and nuclear factor-κB (NF-κB) subunits, p65 and p50, and IκB in MDA-MB-231 cells. 3-(4,5-Dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) assay was used for cell viability. MMP-9 activity and invasion were measured by gelatin zymography and a matrigel invasion assay, respectively. NF-κB activity was measured by an electrophoretic mobility shift assay and luciferase activity. MECF had no effect on cell viability up to a concentration of 100 μg/mL in human breast cancer MDA-MB-231 cells regardless of the presence of TNF-α. MDA-MB-231 cells that were stimulated with TNF-α showed a marked increase of invasion compared to the untreated control, whereas pretreatment with MECF downregulated the TNF-α-induced invasion of MDA-MB-231 cells. Additionally, zymography, western blot analysis, and RT-PCR confirmed that MECF decreased TNF-α-induced MMP-9 expression and activity which is a key regulator for cancer invasion. According to an electrophoretic morbidity shift assay, pretreatment with MECF in MDA-MB-231 cells significantly decreased the TNF-α-induced DNA-binding activity of NF-κB, which is an important transcription factor for regulating cancer invasion-related genes such as MMP-9. Furthermore, treatment with MECF sustained the expression of p65 and p50 in response to TNF-α in the cytosolic compartment. The luciferase assay demonstrated that MECF attenuated TNF-α-induced NF-κB luciferase activity. MECF exhibited its anti-invasive capability by downregulating TNF-α-induced MMP-9 expression, resulting from the suppression of NF-κB activity in the human breast cancer cell line MDA-MB-231. Copyright © 2016 Hainan Medical College. Production and hosting by Elsevier B.V. All rights reserved.

Osteoactivin regulates head and neck squamous cell carcinoma invasion by modulating matrix metalloproteases.

PubMed

Arosarena, Oneida A; Barr, Eric W; Thorpe, Ryan; Yankey, Hilary; Tarr, Joseph T; Safadi, Fayez F

2018-01-01

Nearly 60% of patients with head and neck squamous cell carcinoma (HNSCC) die of metastases or locoregional recurrence. Metastasis is mediated by cancer cell migration and invasion, which are in part dependent on extracellular matrix degradation by matrix metalloproteinases. Osteoactivin (OA) overexpression plays a role in metastases in several malignancies, and has been shown to upregulate matrix metalloproteinase (MMP) expression and activity. To determine how OA modulates MMP expression and activity in HNSCC, and to investigate OA effects on cell invasion, we assessed effects of OA treatment on MMP mRNA and protein expression, as well as gelatinase and caseinolytic activity in HNSCC cell lines. We assessed the effects of OA gene silencing on MMP expression, gelatinase and caseinolytic activity, and cell invasion. OA treatment had differential effects on MMP mRNA expression. OA treatment upregulated MMP-10 expression in UMSCC14a (p = 0.0431) and SCC15 (p < 0.0001) cells, but decreased MMP-9 expression in UMSCC14a cells (p = 0.0002). OA gene silencing decreased MMP-10 expression in UMSCC12 cells (p = 0.0001), and MMP-3 (p = 0.0005) and -9 (p = 0.0036) expression in SCC25 cells. In SCC15 and SCC25 cells, OA treatment increased MMP-2 (p = 0.0408) and MMP-9 gelatinase activity (p < 0.0001), respectively. OA depletion decreased MMP-2 (p = 0.0023) and -9 (p < 0.0001) activity in SCC25 cells. OA treatment increased 70 kDa caseinolytic activity in UMSCC12 cells consistent with tissue type plasminogen activator (p = 0.0078). OA depletion decreased invasive capacity of UMSCC12 cells (p < 0.0001). OA's effects on MMP expression in HNSCC are variable, and may promote cancer cell invasion. © 2017 Wiley Periodicals, Inc.

Silibinin inhibits triple negative breast cancer cell motility by suppressing TGF-β2 expression.

PubMed

Kim, Sangmin; Han, Jeonghun; Jeon, Myeongjin; You, Daeun; Lee, Jeongmin; Kim, Hee Jung; Bae, Sarang; Nam, Seok Jin; Lee, Jeong Eon

2016-08-01

Transforming growth factor-beta (TGF-β) is a multifunctional cytokine that regulates many biological events including cell motility and angiogenesis. Here, we investigated the role of elevated TGF-β2 level in triple negative breast cancer (TNBC) cells and the inhibitory effect of silibinin on TGF-β2 action in TNBC cells. Breast cancer patients with high TGF-β2 expression have a poor prognosis. The levels of TGF-β2 expression increased significantly in TNBC cells compared with those in non-TNBC cells. In addition, cell motility-related genes such as fibronectin (FN) and matrix metalloproteinase-2 (MMP-2) expression also increased in TNBC cells. Basal FN, MMP-2, and MMP-9 expression levels decreased in response to LY2109761, a dual TGF-β receptor I/II inhibitor, in TNBC cells. TNBC cell migration also decreased in response to LY2109761. Furthermore, we observed that TGF-β2 augmented the FN, MMP-2, and MMP-9 expression levels in a time- and dose-dependent manner. In contrast, TGF-β2-induced FN, MMP-2, and MMP-9 expression levels decreased significantly in response to LY2109761. Interestingly, we found that silibinin decreased TGF-β2 mRNA expression level but not that of TGF-β1 in TNBC cells. Cell migration as well as basal FN and MMP-2 expression levels decreased in response to silibinin. Furthermore, silibinin significantly decreased TGF-β2-induced FN, MMP-2, and MMP-9 expression levels and suppressed the lung metastasis of TNBC cells. Taken together, these results suggest that silibinin suppresses metastatic potential of TNBC cells by inhibiting TGF-β2 expression in TNBC cells. Thus, silibinin may be a promising therapeutic drug to treat TNBC.

PPARdelta inhibits IL-1beta-stimulated proliferation and migration of vascular smooth muscle cells via up-regulation of IL-1Ra.

PubMed

Kim, H J; Kim, M Y; Hwang, J S; Kim, H J; Lee, J H; Chang, K C; Kim, J-H; Han, C W; Kim, J-H; Seo, H G

2010-06-01

Activation of peroxisome proliferator-activated receptor (PPAR) delta by GW501516, a specific PPARdelta ligand, significantly inhibited interleukin (IL)-1beta-induced proliferation and migration of vascular smooth muscle cells (VSMCs). This effect of GW501516 was dependent on transforming growth factor-beta, and was mediated through the up-regulation of IL-1 receptor antagonist. The inhibitory effect of GW501516 on VSMC proliferation was associated with cell cycle arrest at the G1 to S phase transition, which was accompanied by the induction of p21 and p53 along with decreased cyclin-dependent kinase 4 expression. Inhibition of cell migration by GW501516 was associated with the down-regulation of matrix metalloproteinase (MMP)-2 and MMP-9 in IL-1beta-treated VSMCs. Inhibition of extracellular signal-regulated kinase significantly reduced the GW501516-mediated inhibition of IL-1beta-stimulated VSMC proliferation. These results suggest that PPARdelta plays an important role in the pathophysiology of diseases associated with the proliferation and migration of VSMCs.

Occludin is regulated by epidermal growth factor receptor activation in brain endothelial cells and brains of mice with acute liver failure.

PubMed

Chen, Feng; Hori, Tomohide; Ohashi, Norifumi; Baine, Ann-Marie; Eckman, Christopher B; Nguyen, Justin H

2011-04-01

Mechanisms of brain edema in acute liver failure (ALF) are not completely understood. We recently demonstrated that matrix metalloproteinase 9 (MMP-9) induces significant alterations to occludin in brain endothelial cells in vitro and in brains of mice with experimental ALF (Hepatology 2009;50:1914). In this study we show that MMP-9-induced transactivation of epidermal growth factor receptor (EGFR) and p38 MAPK/NFκB (mitogen-activated protein kinase/nuclear factor-kappa B) signals participate in regulating brain endothelial occludin level. Mouse brain endothelial bEnd3 cells were exposed to MMP-9 or p38 MAPK up-regulation in the presence and absence of EGFR inhibitor, p38 MAPK inhibitor, NFκB inhibitor, and/or appropriate small interfering RNA. Reverse-transcription polymerase chain reaction (RT-PCR) and western blotting were used for messenger RNA and protein expression analyses. Immunohistochemical staining and confocal microscopy were used to demonstrate cellular EGFR activation. Intraperitoneal azoxymethane was use to induce ALF in mice. Brains of comatose ALF mice were processed for histological and biochemical analyses. When bEnd3 cells were exposed to MMP-9, EGFR was significantly transactivated, followed by p38 MAPK activation, I-kappa B alpha (IκBα) degradation, NFκB activation, and suppression of occludin synthesis and expression. Similar EGFR activation and p38 MAPK/NFκB activation were found in the brains of ALF mice, and these changes were attenuated with GM6001 treatment. EGFR activation with p38 MAPK/NFκB signaling contributes to the regulation of tight junction integrity in ALF. EGFR activation may thus play an important role in vasogenic brain edema in ALF. 2011 American Association for the Study of Liver Diseases.

A Comparison of the Inflammatory and Proteolytic Effects of Dung Biomass and Cigarette Smoke Exposure in the Lung

PubMed Central

Mehra, Divya; Geraghty, Patrick M.; Hardigan, Andrew A.; Foronjy, Robert

2012-01-01

Rationale Biomass is the energy source for cooking and heating for billions of people worldwide. Despite their prevalent use and their potential impact on global health, the effects of these fuels on lung biology and function remain poorly understood. Methods We exposed human small airway epithelial cells and C57BL/6 mice to dung biomass smoke or cigarette smoke to compare how these exposures impacted lung signaling and inflammatory and proteolytic responses that have been linked with disease pathogenesis. Results The in vitro exposure and siRNA studies demonstrated that biomass and cigarette smoke activated ERK to up regulate IL-8 and MMP-1 expression in human airway epithelial cells. In contrast to cigarette smoke, biomass also activated p38 and JNK within these lung cells and lowered the expression of tissue inhibitor of matrix metalloproteinase-1 (TIMP-1). Similarly, in the lungs of mice, both biomass and cigarette smoke exposure increased macrophages, activated ERK and p38 and up regulated MMP-9 and MMP-12 expression. The main differences seen in the exposure studies was that mice exposed to biomass exhibited more perivascular inflammation and had higher G-CSF and GM-CSF lavage fluid levels than mice exposed identically to cigarette smoke. Conclusion Biomass activates similar pathogenic processes seen in cigarette smoke exposure that are known to result in the disruption of lung structure. These findings provide biological evidence that public health interventions are needed to address the harm associated with the use of this fuel source. PMID:23285217

Inhibition of miR-155 attenuates abdominal aortic aneurysm in mice by regulating macrophage-mediated inflammation.

PubMed

Zhang, Zhidong; Liang, Kai; Zou, Gangqiang; Chen, Xiaosan; Shi, Shuaitao; Wang, Guoquan; Zhang, Kewei; Li, Kun; Zhai, Shuiting

2018-06-29

The aim of the present study was to identify abdominal aortic aneurysms (AAA)-associated miR-155 contributing to AAA pathology by regulating macrophage-mediated inflammation. Angiotensin II (AngII)-infused apolipoprotein E-deficient (ApoE-/-) mice and THP-1 cells model of miR-155 overexpression and deficiency were used in the experiments. The expression of miR-155 was detected by quantitative reverse transcription polymerase chain reaction (qRT-PCR). Cytokines were evaluated using enzyme-linked immunoabsorbent assay (ELISA). Western blotting was used to measure the levels of MMP-2, MMP-9, iNOS, and monocyte chemoattractant protein (MCP)-1 proteins. Immunostaining and transwell were used to determine CD68, elastic collagen, proliferation, and migration of vascular smooth muscle cells (VSMCs). The results showed that miR-155 and cytokines were up-regulated in AAA patients or ApoE-/- mice. Overexpression of miR-155 enhanced MMP-2, MMP-9, iNOS, and MCP-1 levels, and stimulated the proliferation and migration of VSMCs. Meanwhile, inhibition of miR-155 had the opposite effect. In addition, histology demonstrated accumulation of CD68 and elastic collagen-positive areas significantly decreased in miR-155 antagomir injection group. In conclusion, the results of the present study suggest that inhibiting miR-155 is crucial to prevent the development of AAA by regulating macrophage inflammation. © 2018 The Author(s).

The pVHL172 isoform is not a tumor suppressor and up-regulates a subset of pro-tumorigenic genes including TGFB1 and MMP13

PubMed Central

Hascoet, Pauline; Chesnel, Franck; Jouan, Florence; Goff, Cathy Le; Couturier, Anne; Darrigrand, Eric; Mahe, Fabrice; Rioux-Leclercq, Nathalie; Goff, Xavier Le; Arlot-Bonnemains, Yannick

2017-01-01

The von Hippel-Lindau (VHL) tumor suppressor gene is often deleted or mutated in ccRCC (clear cell renal cell carcinoma) producing a non-functional protein. The gene encodes two mRNA, and three protein isoforms (pVHL213, pVHL160 and pVHL172). The pVHL protein is part of an E3 ligase complex involved in the ubiquitination and proteasomal degradation of different proteins, particularly hypoxia inducible factors (HIF) that drive the transcription of genes involved in the regulation of cell proliferation, angiogenesis or extracellular matrix remodelling. Other non-canonical (HIF-independent) pVHL functions have been described. A recent work reported the expression of the uncharacterized protein isoform pVHL172 which is translated from the variant 2 by alternative splicing of the exon 2. This splice variant is sometimes enriched in the ccRCCs and the protein has been identified in the respective samples of ccRCCs and different renal cell lines. Functional studies on pVHL have only concerned the pVHL213 and pVHL160 isoforms, but no function was assigned to pVHL172. Here we show that pVHL172 stable expression in renal cancer cells does not regulate the level of HIF, exacerbates tumorigenicity when 786-O-pVHL172 cells were xenografted in mice. The pVHL172-induced tumors developed a sarcomatoid phenotype. Moreover, pVHL172 expression was shown to up regulate a subset of pro-tumorigenic genes including TGFB1, MMP1 and MMP13. In summary we identified that pVHL172 is not a tumor suppressor. Furthermore our findings suggest an antagonistic function of this pVHL isoform in the HIF-independent aggressiveness of renal tumors compared to pVHL213. PMID:29100286

Involvement of Smad3 pathway in atrial fibrosis induced by elevated hydrostatic pressure.

PubMed

Wei, Wei; Rao, Fang; Liu, Fangzhou; Xue, Yumei; Deng, Chunyu; Wang, Zhaoyu; Zhu, Jiening; Yang, Hui; Li, Xin; Zhang, Mengzhen; Fu, Yongheng; Zhu, Wensi; Shan, Zhixin; Wu, Shulin

2018-06-01

Hypertension is a main risk factor for atrial fibrillation, but the direct effects of hydrostatic pressure on the atrial fibrosis are still unknown. The present study investigated whether hydrostatic pressure is responsible for atrial fibrosis, and addressed a potential role of the Smad pathway in this pathology. Biochemical assays were used to study regulation and expression of fibrotic factors in spontaneously hypertensive rats (SHRs) and Wistar rats, and in cardiac fibroblasts (CFs) cultured under standard (0 mmHg) and elevated (20, 40 mmHg) hydrostatic pressure. Levels of atrial fibrosis and protein expression of fibrotic factors Col-1A1/-3A1, TGF-β1, and MMP-2 in SHRs' left atrial tissues were higher than those in Wistar rats. Exposure to elevated pressure was associated with the proliferation of CFs. The protein expression of Col-1A1/-3A1, TGF-β1, and MMP-2 in CFs was also up-regulated in a pressure-dependent manner. The proliferation of CFs and increased expressions of fibrotic markers induced by elevated hydrostatic pressure could be reversed by the Smad3 inhibitor naringenin. The activation of Smad3 pathway was also stimulated by elevated hydrostatic pressure. These results demonstrate that CF secretory function and proliferation can be up-regulated by exposure to elevated pressure, and that Smad3 may modulate CF activation induced by high hydrostatic pressure. © 2017 Wiley Periodicals, Inc.

The emerging role of MMP14 in brain tumorigenesis and future therapeutics.

PubMed

Ulasov, Ilya; Yi, Ruiyang; Guo, Donna; Sarvaiya, Purvaba; Cobbs, Charles

2014-08-01

Glioblastoma is a malignant brain tumor of glial origin. These tumors are thought to be derived from astrocytic cells that undergo malignant transformation. A growing body of evidence suggests that upregulation of MMP expression plays a significant role in promoting glioma pathogenesis. Elevated expression of MMP14 not only promotes glioma invasion and tumor cell proliferation but also plays a role in angiogenesis. Despite the fact that levels of MMP14 correlate with breast cancer progression, the controversial role of MMP14 in gliomagenesis needs to be elucidated. In the present review, we discuss the role of MMP14 in glioma progression as well as the mechanisms of MMP14 regulation in the context of future therapeutic manipulations. Copyright © 2014 Elsevier B.V. All rights reserved.

Gasdermin C is induced by ultraviolet light and contributes to MMP-1 expression via activation of ERK and JNK pathways.

PubMed

Kusumaningrum, Novi; Lee, Dong Hun; Yoon, Hyun-Sun; Kim, Yeon Kyung; Park, Chi-Hyun; Chung, Jin Ho

2018-05-01

Ultraviolet (UV) radiation plays important roles in various skin diseases including premature aging and cancer. UV has been shown to regulate the expressions of many genes including matrix metalloproteinases (MMPs). Gasdermin C (GSDMC) belongs to Gasdermin family and is known to be expressed in the epithelial cells of many tissues including the skin. However, the functions of GSDMC remain poorly understood. We aimed to investigate the role of GSDMC in UV-induced MMP-1, MMP-3, and MMP-9 expressions in human skin keratinocytes. Primary human skin keratinocytes and an immortalized human skin keratinocyte cell line (HaCaT cells) were irradiated with UV. Knockdown and overexpression of GSDMC were performed to study the effect of GSDMC. The mRNA and protein levels were analyzed by quantitative real-time polymerase chain reaction (qRT-PCR) and western blotting, respectively. We found that GSDMC expression is increased by UV irradiation in human skin keratinocytes. Further studies showed that GSDMC expression is increased at relatively late time points after UV irradiation and that this GSDMC induction plays important roles in the expressions of MMP-1, but not of MMP-3 and MMP-9, and the activations of ERK and JNK induced by UV. In addition, we found that overexpression of GSDMC increases the MMP-1 expression and the activities of ERK and JNK and that GSDMC-induced MMP-1 expression is suppressed by inhibition of ERK or JNK activities. Our results suggest that GSDMC is increased by UV radiation and contributes to UV-induced MMP-1 expression through the activation of ERK and JNK pathways. Copyright © 2018 Japanese Society for Investigative Dermatology. Published by Elsevier B.V. All rights reserved.

Nitric Oxide Promotes Airway Epithelial Wound Repair through Enhanced Activation of MMP-9

PubMed Central

Bove, Peter F.; Wesley, Umadevi V.; Greul, Anne-Katrin; Hristova, Milena; Dostmann, Wolfgang R.; van der Vliet, Albert

2007-01-01

The airway epithelium provides a protective barrier against inhaled environmental toxins and microorganisms, and epithelial injury initiates a number of processes to restore its barrier integrity, including activation of matrix metalloproteinases such as MMP-9 (92-kD gelatinase B). Airway epithelial cells continuously produce nitric oxide (NO), which has been linked to cell migration and MMP-9 regulation in several cell types, but the importance of epithelial NO in mediating airway epithelial repair or MMP-9 activation is unknown. Using primary or immortalized human bronchial epithelial cells, we demonstrate that low concentrations of NO promote epithelial cell migration and wound repair in an in vitro wound assay, which was associated with increased localized expression and activation of MMP-9. In addition, in HBE1 cells that were stably transfected with inducible NOS (NOS2), to mimic constitutive epithelial NOS2 expression in vivo, NOS inhibition decreased epithelial wound repair and MMP-9 expression. The stimulatory effects of NO on epithelial wound repair and MMP-9 expression were dependent on cGMP-mediated pathways and were inhibited by 1H-[1,2,4]oxadiazolo-[4,3-a]quinoxalin-1-one (ODQ), an inhibitor of soluble guanylyl cyclase. Inhibition of cGMP-dependent protein kinase (PKG) attenuated NO-mediated epithelial wound closure, but did not affect MMP-9 expression. However, pharmacologic MMP inhibition and siRNA knockdown of MMP-9 expression demonstrated the contribution of MMP-9 to NO-mediated wound closure. Overall, our results demonstrate that NOS2-derived NO contributes to airway epithelial repair by both PKG-dependent and -independent mechanisms, and involves NO-dependent expression and activation of MMP-9. PMID:16980554

Resistin promotes tumor metastasis by down-regulation of miR-519d through the AMPK/p38 signaling pathway in human chondrosarcoma cells

PubMed Central

Huang, Ho-Ning; Hung, Chih-Hung; Hsu, Chin-Jung; Fong, Yi-Chin; Hsu, Horng-Chaung; Huang, Yuan-Li; Tang, Chih-Hsin

2015-01-01

Resistin is a recently discovered adipocyte-secreting adipokine, which may play a critical role in modulating cancer pathogenesis. Chondrosarcoma is a highly malignant tumor known to frequently metastasize; however, the role of resistin in the metastasis of human chondrosarcoma is largely unknown. Here, we found that the expression of resistin was higher in chondrosarcoma biopsy tissues than in normal cartilage. Moreover, treatment with resistin increased matrix metalloproteinase (MMP)-2 expression and promoted cell migration in human chondrosarcoma cells. Co-transfection with microRNA (miR)-519d mimic resulted in reversed resistin-mediated cell migration and MMP-2 expression. Additionally, AMP-activated protein kinase (AMPK) and p38 inhibitors or siRNAs reduced the resistin-increased cell migration and miR-519d suppression, and inhibition of resistin expression resulted in suppression of MMP-2 expression and lung metastasis in vivo. Taken together, our results indicate that resistin promotes chondrosarcoma metastasis and MMP-2 expression through activation of the AMPK/p38 signaling pathway and down-regulation of miR-519d expression. Therefore, resistin may represent a potential novel molecular therapeutic target in chondrosarcoma metastasis. PMID:25404641

Expression of extracellular matrix metalloproteinase inducer (EMMPRIN) and its expected roles in the bovine endometrium during gestation.

PubMed

Mishra, B; Kizaki, K; Koshi, K; Ushizawa, K; Takahashi, T; Hosoe, M; Sato, T; Ito, A; Hashizume, K

2012-02-01

Extracellular matrix metalloproteinase inducer (EMMPRIN) and its induced matrix metalloproteinases (MMPs) play a crucial role in tissue remodeling during the peri-implantation period. However, the role of EMMPRIN in the bovine placenta is still unclear. We have postulated that EMMPRIN might play a regulatory role in trophoblastic cell functions during gestation by itself or through the regulation of MMP expression. In this study, EMMPRIN mRNA was detected in the bovine placentome and interplacentome throughout gestation, and its expression was significantly higher in the cotyledon during late gestation. In situ hybridization showed that EMMPRIN mRNA was expressed in the caruncular epithelium and the cotyledonary epithelium, including binucleate cells. Western blot analysis detected a band representing a protein of approximately 65 kDa in the caruncular and cotyledonary tissues, and the intensity of its expression was increased in both of these tissues during late gestation. The expression levels of MMP-2 and MMP-14 in the bovine placenta were higher during late gestation, as was observed for EMMPRIN. Therefore, EMMPRIN might regulate trophoblastic cell functions, especially those of binucleate cells, through MMP expression in the bovine placenta. Copyright © 2012 Elsevier Inc. All rights reserved.

Full-length soluble CD147 promotes MMP-2 expression and is a potential serological marker in detection of hepatocellular carcinoma

PubMed Central

2014-01-01

Background As a surface glycoprotein, CD147 is capable of stimulating the production of matrix metalloproteinases (MMPs) from neighboring fibroblasts. The aim of the present study is to explore the role of soluble CD147 on MMPs secretion from hepatocellular carcinoma (HCC) cells, and to investigate the diagnostic value of serum soluble CD147 in the HCC detection. Methods We identified the form of soluble CD147 in cell culture supernate of HCC cells and serum of patients with HCC, and explored the role of soluble CD147 on MMPs secretion. Serum CD147 levels were detected by the enzyme-linked immunosorbent assay, and the value of soluble CD147 as a marker in HCC detection was analyzed. Results Full length soluble CD147 was presented in the culture medium of HCC cells and serum of patients with HCC. The extracellular domain of soluble CD147 promoted the expression of CD147 and MMP-2 from HCC cells. Knockdown of CD147 markedly diminished the up-regulation of CD147 and MMP-2 which induced by soluble CD147. Soluble CD147 activated ERK, FAK, and PI3K/Akt pathways, leading to the up-regulation of MMP-2. The level of soluble CD147 in serum of patients with HCC was significantly elevated compared with healthy individuals (P < 0.001). Soluble CD147 levels were found to be associated with HCC tumor size (P = 0.007) and Child-Pugh grade (P = 0.007). Moreover, soluble CD147 showed a better performance in distinguishing HCC compared with alpha-fetoprotein. Conclusions The extracellular domain of soluble CD147 enhances the secretion of MMP-2 from HCC cells, requiring the cooperation of membrane CD147 and activation of ERK, FAK, and PI3K/Akt signaling. The measurement of soluble CD147 may offer a useful approach in diagnosis of HCC. PMID:24996644

Full-length soluble CD147 promotes MMP-2 expression and is a potential serological marker in detection of hepatocellular carcinoma.

PubMed

Wu, Jiao; Hao, Zhi-Wei; Zhao, You-Xu; Yang, Xiang-Min; Tang, Hao; Zhang, Xin; Song, Fei; Sun, Xiu-Xuan; Wang, Bin; Nan, Gang; Chen, Zhi-Nan; Bian, Huijie

2014-07-04

As a surface glycoprotein, CD147 is capable of stimulating the production of matrix metalloproteinases (MMPs) from neighboring fibroblasts. The aim of the present study is to explore the role of soluble CD147 on MMPs secretion from hepatocellular carcinoma (HCC) cells, and to investigate the diagnostic value of serum soluble CD147 in the HCC detection. We identified the form of soluble CD147 in cell culture supernate of HCC cells and serum of patients with HCC, and explored the role of soluble CD147 on MMPs secretion. Serum CD147 levels were detected by the enzyme-linked immunosorbent assay, and the value of soluble CD147 as a marker in HCC detection was analyzed. Full length soluble CD147 was presented in the culture medium of HCC cells and serum of patients with HCC. The extracellular domain of soluble CD147 promoted the expression of CD147 and MMP-2 from HCC cells. Knockdown of CD147 markedly diminished the up-regulation of CD147 and MMP-2 which induced by soluble CD147. Soluble CD147 activated ERK, FAK, and PI3K/Akt pathways, leading to the up-regulation of MMP-2. The level of soluble CD147 in serum of patients with HCC was significantly elevated compared with healthy individuals (P < 0.001). Soluble CD147 levels were found to be associated with HCC tumor size (P = 0.007) and Child-Pugh grade (P = 0.007). Moreover, soluble CD147 showed a better performance in distinguishing HCC compared with alpha-fetoprotein. The extracellular domain of soluble CD147 enhances the secretion of MMP-2 from HCC cells, requiring the cooperation of membrane CD147 and activation of ERK, FAK, and PI3K/Akt signaling. The measurement of soluble CD147 may offer a useful approach in diagnosis of HCC.

Dimerization of Matrix Metalloproteinase-2 (MMP-2)

PubMed Central

Koo, Bon-Hun; Kim, Yeon Hyang; Han, Jung Ho; Kim, Doo-Sik

2012-01-01

Matrix metalloproteinase-2 (MMP-2) functions in diverse biological processes through the degradation of extracellular and non-extracellular matrix molecules. Because of its potential for tissue damage, there are several ways to regulate MMP-2 activity, including gene expression, compartmentalization, zymogen activation, and enzyme inactivation by extracellular inhibitors. Enzyme regulation through zymogen activation is important for the regulation of MMP-2 activity. In our previous studies, we showed that thrombin directly cleaved the propeptide of MMP-2 at specific sites for enzyme activation. We also demonstrated that heparan sulfate was required for thrombin-mediated activation of pro-MMP-2 by binding to thrombin, presumably through conformational changes at the active site of the enzyme. This suggests a regulatory mechanism for thrombin-mediated activation of pro-MMP-2. In this study, we found that MMP-2 formed a reduction-sensitive homodimer in a controlled manner and that Ca2+ ion was essential for homodimerization of MMP-2. Homodimerization was not associated with protein kinase C-mediated phosphorylation of MMP-2. MMP-2 formed a homodimer through an intermolecular disulfide bond between Cys102 and the neighboring Cys102. Homodimerization of MMP-2 enhanced thrombin-mediated activation of pro-MMP-2. Moreover, the MMP-2 homodimer could cleave a small peptide substrate without removal of the propeptide. Taken together, our experimental data suggest a novel regulatory mechanism for pro-MMP-2 activation that is modulated through homodimerization of MMP-2. PMID:22577146

Serum Levels of IL-1β, IL-6, TGF-β, and MMP-9 in Patients Undergoing Carotid Artery Stenting and Regulation of MMP-9 in a New In Vitro Model of THP-1 Cells Activated by Stenting

PubMed Central

Zhang, Rongrong; Jiang, Fan; Chen, Cindy Si; Wang, Tianzhu; Feng, Jinzhou; Tao, Tao; Qin, Xinyue

2015-01-01

Inflammation plays an important role in the pathophysiological process after carotid artery stenting (CAS). Monocyte is a significant source of inflammatory cytokines in vascular remodeling. Telmisartan could reduce inflammation. In our study, we first found that, after CAS, the serum IL-1β, IL-6, TGF-β, and MMP-9 levels were significantly increased, but only MMP-9 level was elevated no less than 3 months. Second, we established a new in vitro model, where THP-1 monocytes were treated with the supernatants of human umbilical vein endothelial cells (HUVECs) that were scratched by pipette tips, which mimics monocytes activated by mechanical injury of stenting. The treatment enhanced THP-1 cell adhesion, migration and invasion ability, and the phosphorylation of ERK1/2 and Elk-1 and MMP-9 expression were significantly increased. THP-1 cells pretreated with PD98095 (ERK1/2 inhibitor) attenuated the phosphorylation of ERK1/2 and Elk-1 and upregulation of MMP-9, while pretreatment with telmisartan merely decreased the phosphorylation of Elk-1 and MMP-9 expression. These results suggested that IL-1β, IL-6, TGF-β, and MMP-9 participate in the pathophysiological process after CAS. Our new in vitro model mimics monocytes activated by stenting. MMP-9 expression could be regulated through ERK1/2/Elk-1 pathway, and the protective effects of telmisartan after stenting are partly attributed to its MMP-9 inhibition effects via suppression of Elk-1. PMID:26113783

Expression of SLP-2 was associated with invasion of esophageal squamous cell carcinoma.

PubMed

Cao, Wenfeng; Zhang, Bin; Ding, Fang; Zhang, Weiran; Sun, Baocun; Liu, Zhihua

2013-01-01

Stomatin-like protein 2 (SLP-2), a member of the Stomatin superfamily, has been identified as an oncogenic-related protein and found to be up-regulated in multi-cancers. Nonetheless, the expression pattern and regulation of SLP-2 in human esophageal squamous cell carcinoma (ESCC) remain unexplored. Immunohistochemistry and immunofluorescence staining analysis were performed to show SLP-2 expression and location. RNAi method was used to inhibit specific protein expression. Transwell assay was done to investigate cells invasive capability. RT-PCR and Western blot analysis were used to detect mRNA and protein expression levels. Immunohistochemical analysis showed that up-regulation of SLP-2 was found in invasive front compared with cancer central tissue in ESCC. Inhibition of SLP-2 by SLP-2 siRNA can decrease ESCC cells invasive capability through MMP-2 dependent manner. Up-regulation of SLP-2 was effectively abrogated by the ERK1/2 inhibitors either PD98059 or U0126, but no effect was showed by the treatment of AKT inhibitors either LY294002 or MK-2206. So the regulation of SLP-2 was involved in activation of the MAPK/ERK pathway. We found that PMA/EGF could induce the up-regulated expression of SLP-2 probably through activating ERK signalling. The current study suggests that SLP-2 may represent an important molecular hallmark that is clinically relevant to the invasion of ESCC.

Yin Yang 1 Is a Critical Repressor of Matrix Metalloproteinase-9 Expression in Brain Neurons*

PubMed Central

Rylski, Marcin; Amborska, Renata; Zybura, Katarzyna; Mioduszewska, Barbara; Michaluk, Piotr; Jaworski, Jacek; Kaczmarek, Leszek

2008-01-01

Membrane depolarization controls long lasting adaptive neuronal changes in brain physiology and pathology. Such responses are believed to be gene expression-dependent. Notably, however, only a couple of gene repressors active in nondepolarized neurons have been described. In this study, we show that in the unstimulated rat hippocampus in vivo, as well as in the nondepolarized brain neurons in primary culture, the transcriptional regulator Yin Yang 1 (YY1) is bound to the proximal Mmp-9 promoter and strongly represses Mmp-9 transcription. Furthermore, we demonstrate that monoubiquitinated and CtBP1 (C-terminal binding protein 1)-bound YY1 regulates Mmp-9 mRNA synthesis in rat brain neurons controlling its transcription apparently via HDAC3-dependent histone deacetylation. In conclusion, our data suggest that YY1 exerts, via epigenetic mechanisms, a control over neuronal expression of MMP-9. Because MMP-9 has recently been shown to play a pivotal role in physiological and pathological neuronal plasticity, YY1 may be implicated in these phenomena as well. PMID:18940814

Collagenase IV plays an important role in regulating hair cycle by inducing VEGF, IGF-1, and TGF-β expression

PubMed Central

Hou, Chun; Miao, Yong; Wang, Jin; Wang, Xue; Chen, Chao-Yue; Hu, Zhi-Qi

2015-01-01

Background It has been reported that collagenases (matrix metalloproteinase 2 [MMP-2] and matrix metalloproteinase 9 [MMP-9]) are associated with hair cycle, whereas the mechanism of the association is largely unknown. Methods The mice were randomly allocated into four groups: saline, and 5, 10, and 15 nM SB-3CT. Immunohistochemical analysis was employed to examine MMP-2 and MMP-9 protein. Real-time polymerase chain reaction and enzyme-linked immunosorbent assay were performed to determine mRNA and protein levels of VEGF, IGF-1, TGF-β, and GAPDH. Growing hair follicles from anagen phase III–IV were scored based on hematoxylin and eosin staining. Hair regrowth was also evaluated. Results Results showed that mRNA expressions of enzymes changed with a peak at late anagen and a trough at telogen after depilation. Immunostaining showed that the highest expression of MMP-2 was more than that of MMP-9, and the highest expression of enzymes changed during anagen. The localizations of MMP-2 changed from dermal papilla, keratinocyte strand, out of root sheath, and basal plate at early anagen, to hair bulb, inner root sheath, and outer root sheath at late anagen. The localization of MMP-9 changed from partial keratinocyte to dermal papilla at early anagen and to outer root sheath at late anagen. VEGF, IGF-1, and TGF-β have been shown to regulate hair growth. We found mRNA and protein expressions of VEGF and IGF-1 fluctuated with a peak at anagen and a decrease at catagen to telogen. In contrast, mRNA and protein expressions of TGF-β changed with highest and lowest levels at anagen and telogen, respectively. With selective inhibitor of collagenase IV, SB-3CT, mice showed significant suppressed hair growth and decreased expression of VEGF, IGF-1, and TGF-β. The MMPs agonist also significantly increased expression of VEGF, IGF-1, and TGF-β. Meanwhile, SB-3CT treatment significantly suppressed hair growth. Conclusion All these data suggest that the type IV collagenases, MMP-2 and MMP-9, play important roles in hair cycle, and this could be mediated by induced expression of VEGF, IGF-1, and TGF-β. PMID:26451090

Gel structure has an impact on pericellular and extracellular matrix deposition, which subsequently alters metabolic activities in chondrocyte-laden PEG hydrogels.

PubMed

Nicodemus, G D; Skaalure, S C; Bryant, S J

2011-02-01

While designing poly(ethylene glycol) hydrogels with high moduli suitable for in situ placement is attractive for cartilage regeneration, the impact of a tighter crosslinked structure on the organization and deposition of the matrix is not fully understood. The objectives of this study were to characterize the composition and spatial organization of new matrix as a function of gel crosslinking and study its impact on chondrocytes in terms of anabolic and catabolic gene expression and catabolic activity. Bovine articular chondrocytes were encapsulated in hydrogels with three crosslinking densities (compressive moduli 60, 320 and 590 kPa) and cultured for 25 days. Glycosaminoglycan production increased with culture time and was greatest in the gels with lowest crosslinking. Collagens II and VI, aggrecan, link protein and decorin were localized to pericellular regions in all gels, but their presence decreased with increasing gel crosslinking. Collagen II and aggrecan expression were initially up-regulated in gels with higher crosslinking, but increased similarly up to day 15. Matrix metalloproteinase (MMP)-1 and MMP-13 expression were elevated (∼25-fold) in gels with higher crosslinking throughout the study, while MMP-3 was unaffected by gel crosslinking. The presence of aggrecan and collagen degradation products confirmed MMP activity. These findings indicate that chondrocytes synthesized the major cartilage components within PEG hydrogels, however, gel structure had a significant impact on the composition and spatial organization of the new tissue and on how chondrocytes responded to their environment, particularly with respect to their catabolic expression. Copyright © 2010 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

Leptin-mediated regulation of MT1-MMP localization is KIF1B dependent and enhances gastric cancer cell invasion.

PubMed

Dong, Zhaogang; Xu, Xiaofei; Du, Lutao; Yang, Yongmei; Cheng, Huanhuan; Zhang, Xin; Li, Zewu; Wang, Lili; Li, Juan; Liu, Hui; Qu, Xun; Wang, Chuanxin

2013-05-01

Leptin overexpression is closely correlated with gastric cancer (GC) invasion, but its exact effect and the underlying mechanism in tumorigenesis remain poorly understood. Membrane type 1-matrix metalloproteinase (MT1-MMP), a surface-anchored 'master switch' proteinase, is overexpressed and plays crucial roles in tumor invasion. Here, we characterized the influence of leptin on the generation and surface localization of MT1-MMP in GC and elucidated its molecular mechanisms. Our results revealed that leptin promoted GC cell invasion in vitro by upregulating MT1-MMP expression. Furthermore, cell surface biotinylation assay and flow cytometry demonstrated that the surface expression of MT1-MMP was also enhanced by leptin, and knockdown of kinesin family member 1B (KIF1B, a microtubule plus end-directed monomeric motor protein) by small interference RNA inhibited this process. Notably, coimmunoprecipitation analysis indicated that leptin enhanced the interaction of MT1-MMP with KIF1B in a time-dependent manner, which consequently contributed to GC cell invasion. Moreover, leptin increased MT1-MMP or KIF1B expression by the protein kinase B (AKT) pathway and extracellular signal-regulated kinase 1/2 partially participated in this process. However, only AKT was implicated in the leptin-mediated membrane localization of MT1-MMP. Immunohistochemistry analysis revealed that leptin, MT1-MMP and KIF1B are overexpressed in GC tissues, and they positively correlated with clinical stage and lymph node metastasis. These observations indicate that this regulatory network exists in vivo. Taken together, our findings suggest that leptin is an effective intracellular stimulator of MT1-MMP and that leptin-enhanced cell surface localization of MT1-MMP is dependent on KIF1B, which consequently plays a critical role in GC invasion.

EMMPRIN (basigin/CD147) is involved in the morphogenesis of tooth germ in mouse molars.

PubMed

Xie, Ming; Jiao, Ting; Chen, Yuqin; Xu, Chun; Li, Jing; Jiang, Xinquan; Zhang, Fuqiang

2010-05-01

The pattern of gene expression for extracellular matrix metalloproteinase inducer (EMMPRIN) was revealed in the tooth germ of mouse mandibular molars using quantitative real-time PCR. In situ hybridization and immunohistochemical study demonstrated the characteristic distribution of EMMPRIN in the different stages of tooth germ development. To investigate the functional role played by EMMPRIN in tooth germ development, EMMPRIN siRNA interference approach was carried out in cultured mouse mandibles at embryonic day 11.0 (E11.0). The results showed that EMMPRIN siRNA-treated explants exhibited a marked growth inhibition of tooth germ compared to the control and scrambled siRNA-treated explants. Meanwhile, a significant increase in MT1-MMP mRNA expression and a reduction in MMP-2, MMP-3, MMP-9, MMP-13 and MT2-MMP mRNA expression were observed in the mouse mandibles following EMMPRIN abrogation. The current results indicate that EMMPRIN could thus be involved in the early stage of tooth germ development and morphogenesis, possibly by regulating the expression of MMP genes.

Stromal matrix metalloproteinase 2 regulates collagen expression and promotes the outgrowth of experimental metastases.

PubMed

Bates, Andreia L; Pickup, Michael W; Hallett, Miranda A; Dozier, E Ashley; Thomas, Stacy; Fingleton, Barbara

2015-04-01

Breast cancer survival rates decrease from 99% for patients with local disease to 25% for those with distant metastases. Matrix metalloproteinases (MMPs), including MMP2, are associated with metastatic progression. We found that loss of host MMP2 reduces the proliferation of experimental metastases in the lungs and identified fibroblasts in tumour-bearing lungs as the major source of MMP2. In vitro, spheroidal mammary tumour growth was increased by co-culture with control fibroblasts isolated from tumour-bearing lungs, but not when fibroblasts with stable Mmp2 knockdown were used. This result prompted us to assess whether MMP2 was responsible for a tumour-proliferative, activated fibroblast phenotype. To test this, we evaluated: (a) fibroblasts from wild-type tumour-bearing lungs, with or without shRNA-mediated MMP2 knockdown; and (b) normal, quiescent fibroblasts isolated from either WT or Mmp2(-/-) mice. Quantitative PCR revealed that Mmp2 knockdown attenuated expression of two markers of activation (α-smooth muscle actin and vimentin), but there was minimal expression in quiescent WT or Mmp2(-/-) fibroblasts, as expected. Placing quiescent fibroblasts under activating conditions led to increases in activation-associated transcripts in WT but not Mmp2(-/-) fibroblasts. Additionally, Mmp2 knockdown fibroblasts showed significantly decreased expression of the matrix transcripts collagen I, collagen IV and fibronectin. Addition of active TGFβ was sufficient to rescue the MMP2-dependent collagen I and IV expression, while MMP2-induced collagen expression was blocked by the addition of TGFβ1-neutralizing antibody. Gene expression data in stromal cells of human breast cancers reveal that MMP2 expression is also positively correlated with activation and matrix transcripts. Thus, we present a model whereby MMP2 production in tumour fibroblasts is important for TGFβ1 activity and subsequent activation of fibroblasts to a matrix-producing, proliferation-supportive phenotype. Overall, our results reveal a previously undefined role for MMP2 in metastatic outgrowth mediated by fibroblasts, and extend the mechanisms by which MMPs contribute to tumour progression. Copyright © 2014 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

The zinc-finger transcription factor Hindsight regulates ovulation competency of Drosophila follicles

PubMed Central

Deady, Lylah D; Li, Wei

2017-01-01

Follicle rupture, the final step in ovulation, utilizes conserved molecular mechanisms including matrix metalloproteinases (Mmps), steroid signaling, and adrenergic signaling. It is still unknown how follicles become competent for follicle rupture/ovulation. Here, we identify a zinc-finger transcription factor Hindsight (Hnt) as the first transcription factor regulating follicle’s competency for ovulation in Drosophila. Hnt is not expressed in immature stage-13 follicle cells but is upregulated in mature stage-14 follicle cells, which is essential for follicle rupture/ovulation. Hnt upregulates Mmp2 expression in posterior follicle cells (essential for the breakdown of the follicle wall) and Oamb expression in all follicle cells (the receptor for receiving adrenergic signaling and inducing Mmp2 activation). Hnt’s role in regulating Mmp2 and Oamb can be replaced by its human homolog Ras-responsive element-binding protein 1 (RREB-1). Our data suggest that Hnt/RREB-1 plays conserved role in regulating follicle maturation and competency for ovulation. PMID:29256860

Immunohistochemical expression of matrix metalloproteinase 13 in chronic periodontitis.

PubMed

Nagasupriya, Alapati; Rao, Donimukkala Bheemalingeswara; Ravikanth, Manyam; Kumar, Nalabolu Govind; Ramachandran, Cinnamanoor Rajmani; Saraswathi, Thillai Rajashekaran

2014-01-01

The extracellular matrix is a complex integrated system responsible for the physiologic properties of connective tissue. Collagen is the major extracellular component that is altered in pathologic conditions, mainly periodontitis. The destruction involves proteolytic enzymes, primarily matrix metalloproteinases (MMPs), which play a key role in mediating and regulating the connective tissue destruction in periodontitis. The study group included 40 patients with clinically diagnosed chronic periodontitis. The control group included 20 patients with clinically normal gingiva covering impacted third molars undergoing extraction or in areas where crown-lengthening procedures were performed. MMP-13 expression was demonstrated using immunohistochemistry in all the gingival biopsies, and the data were analyzed statistically. MMP-13 expression was observed more in chronic periodontitis when compared with normal gingiva. MMP-13 expression was expressed by fibroblasts, lymphocytes, macrophages, plasma cells, and basal cells of the sulcular epithelium. Comparative evaluation of all the clinical and histologic parameters with MMP-13 expression showed high statistical significance with Spearman correlation coefficient. Elevated levels of MMP-13 may play a role in the pathogenesis of chronic periodontitis. There is a direct correlation of increased expression of MMP-13 with various clinical and histologic parameters in disease severity.

HuR (Elavl1) and HuB (Elavl2) Stabilize Matrix Metalloproteinase-9 mRNA During Seizure-Induced Mmp-9 Expression in Neurons

PubMed Central

Zybura-Broda, Katarzyna; Wolder-Gontarek, Malgorzata; Ambrozek-Latecka, Magdalena; Choros, Artur; Bogusz, Agnieszka; Wilemska-Dziaduszycka, Joanna; Rylski, Marcin

2018-01-01

Matrix metalloproteinase-9 (Mmp-9) is involved in different general and cell-type–specific processes, both in neuronal and non-neuronal cells. Moreover, it is implicated in an induction or progression of various human disorders, including diseases of the central nervous system. Mechanisms regulating activity-driven Mmp-9 expression in neurons are still not fully understood. Here, we show that stabilization of Mmp-9 mRNA is one of the factors responsible for the neuronal activity-evoked upregulation of Mmp-9 mRNA expression in hippocampal neurons. Furthermore, we demonstrate that the molecular mechanism related to this stabilization is dependent on the neuronal seizure-triggered transiently increased binding of the mRNA stability-inducing protein, HuR, to ARE1 and ARE4 motifs of the 3′UTR for Mmp-9 mRNA as well as the stably augmented association of another mRNA-stabilizing protein, HuB, to the ARE1 element of the 3′UTR. Intriguingly, we demonstrate further that both HuR and HuB are crucial for an incidence of Mmp-9 mRNA stabilization after neuronal activation. This study identifies Mmp-9 mRNA as the first HuB target regulated by mRNA stabilization in neurons. Moreover, these results are the first to describe an existence of HuR-dependent mRNA stabilization in neurons of the brain. PMID:29686606

Stimulation of nuclear receptor REV-ERBs regulates tumor necrosis factor-induced expression of proinflammatory molecules in C6 astroglial cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Morioka, Norimitsu, E-mail: mnori@hiroshima-u.ac.jp; Tomori, Mizuki; Zhang, Fang Fang

Under physiological conditions, astrocytes maintain homeostasis in the CNS. Following inflammation and injury to the CNS, however, activated astrocytes produce neurotoxic molecules such as cytokines and chemokines, amplifying the initial molecular-cellular events evoked by inflammation and injury. Nuclear receptors REV-ERBα and REV-ERBβ (REV-ERBs) are crucial in the regulation of inflammation- and metabolism-related gene transcription. The current study sought to elucidate a role of REV-ERBs in rat C6 astroglial cells on the expression of inflammatory molecules following stimulation with the neuroinflammatory cytokine tumor necrosis factor (TNF). Stimulation of C6 cells with TNF (10 ng/ml) significantly increased the mRNA expression of CCL2, interleukin-6more » (IL-6), inducible nitric oxide synthase (iNOS), and matrix metalloprotease (MMP)-9, but not fibroblast growth factor-2 (FGF-2), cyclooxygenase-2 (COX-2) and MMP-2. Treatment with either REV-ERB agonists GSK4112 or SR9009 significantly blocked TNF-induced upregulation of CCL2 mRNA and MMP-9 mRNA, but not IL-6 mRNA and iNOS mRNA expression. Furthermore, treatment with RGFP966, a selective histone deacetylase 3 (HDAC3) inhibitor, potently reversed the inhibitory effects of GSK4112 on TNF-induced expression of MMP-9 mRNA, but not CCL2 mRNA. Expression of Rev-erbs mRNA in C6 astroglial cells, primary cultured rat cortical and spinal astrocytes was confirmed by reverse transcription polymerase chain reaction. Together, the findings demonstrate an anti-inflammatory effect, downregulating of MMP-9 and CCL2 transcription, of astroglial REV-ERBs activation through HDAC3-dependent and HDAC3-independent mechanisms. - Highlights: • Rev-erbα mRNA and Rev-erbβ mRNA are expressed in C6 astroglial cells. • TNF increases the expression of CCL2, IL-6, MMP-9 and iNOS mRNA. • REV-ERB activation inhibits CCL2 mRNA and MMP-9 mRNA expression. • HDAC3 activity is involved in the inhibitory effect of REV-ERB on MMP-9 induction.« less

Mechanical and IL-1β Responsive miR-365 Contributes to Osteoarthritis Development by Targeting Histone Deacetylase 4.

PubMed

Yang, Xu; Guan, Yingjie; Tian, Shaoqi; Wang, Yuanhe; Sun, Kang; Chen, Qian

2016-03-23

Mechanical stress plays an important role in the initiation and progression of osteoarthritis. Studies show that excessive mechanical stress can directly damage the cartilage extracellular matrix and shift the balance in chondrocytes to favor catabolic activity over anabolism. However, the underlying mechanism remains unknown. MicroRNAs (miRNAs) are emerging as important regulators in osteoarthritis pathogenesis. We have found that mechanical loading up-regulated microRNA miR-365 in growth plate chondrocytes, which promotes chondrocyte differentiation. Here, we explored the role of the mechanical responsive microRNA miR-365 in pathogenesis of osteoarthritis (OA). We found that miR-365 was up-regulated by cyclic loading and IL-1β stimulation in articular chondrocytes through a mechanism that involved the transcription factor NF-κB. miR-365 expressed significant higher level in rat anterior cruciate ligament (ACL) surgery induced OA cartilage as well as human OA cartilage from primary OA patients and traumatic OA Patients. Overexpression of miR-365 in chondrocytes increases gene expression of matrix degrading enzyme matrix metallopeptidase 13 (MMP13) and collagen type X (Col X). The increase in miR-365 expression in OA cartilage and in response to IL-1 may contribute to the abnormal gene expression pattern characteristic of OA. Inhibition of miR-365 down-regulated IL-1β induced MMP13 and Col X gene expression. We further showed histone deacetylase 4 (HDAC4) is a direct target of miR-365, which mediates mechanical stress and inflammation in OA pathogenesis. Thus, miR-365 is a critical regulator of mechanical stress and pro-inflammatory responses, which contributes cartilage catabolism. Manipulation of the expression of miR-365 in articular chondrocytes by miR-365 inhibitor may be a potent therapeutic target for the prevention and treatment of osteoarthritis.

Three-dimensional culture using a radial flow bioreactor induces matrix metalloprotease 7-mediated EMT-like process in tumor cells via TGFbeta1/Smad pathway.

PubMed

Shibata, Shun-Ichi; Marushima, Hideki; Asakura, Tadashi; Matsuura, Tomokazu; Eda, Homare; Aoki, Katsuhiko; Matsudaira, Hiroshi; Ueda, Kazu; Ohkawa, Kiyoshi

2009-05-01

To confirm the usefulness of the radial flow type bioreactor (RFB) for a three-dimensional (3D) culture system, which provides a tissue architecture and molecular function mimicking the in vivo environment, molecular expression in the A431 human squamous carcinoma cell line during culture were analyzed under the physically different environments of 3D culture in the RFB, 2D culture in a monolayer as well as in nude mice. Time-dependent accumulation of autocrine transforming growth factor (TGF) beta1 was found in spent culture media obtained only from 3D cultured A431 cancer cells, which grew well with a stratified-sheet morphology. Cells in the RFB overexpressed matrix metalloproteinase 7 (MMP7) and showed an increased release of soluble 80-kDa fragments of E-cadherin into the media time-dependently, resulting in the reduction of E-cadherin protein at the cell surface without down-regulation of the mRNA. beta-Catenin and its nuclear partner, LEF1, were up-regulated and Wnt protein secretion was also accelerated. Additional up-regulation of the transcriptional factors, HMGA2 and down-stream Slug, was noted. TGFbeta1-dependent, MMP7-mediated up-regulation of beta-catenin/LEF1 signaling and TGFbeta1-activated HMGA2 pathways consequently converged with Slug overexpression, due to disassembly and further repression of E-cadherin expression, which was reproducible in the epithelial mesenchymal transition process without any manipulation. Other transcriptional factors, Notch/HEY1 and NF-kappaB, were also up-regulated in 3D-cultured cells. These signals recruited molecules related to extracellular matrix-cell remodeling and angiogenesis. Expression of several representative molecules in the 3D cultured cells was parallel with that in xenotransplanted A431 tumor tissues in nude mice. 3D culture of tumor cells in the RFB is a useful tool for cancer experimental biology and evaluation of cancer therapeutic-like systems in nude mice.

14,15-EET induces the infiltration and tumor-promoting function of neutrophils to trigger the growth of minimal dormant metastases

PubMed Central

Luo, Chao; Wang, Yu; Li, Dong; Shu, Yu; Wang, Shan-Shan; Qin, Jian; Li, Yong-Chao; Zou, Jiu-Ming; Tian, De-An; Zhang, Gui-Mei; Feng, Zuo-Hua

2016-01-01

Infiltrating neutrophils are known to promote in thedevelopment of tumor. However, it is unclear whether and how neutrophils areinvolved in triggering the growth of dormant metastases. Here we show that14,15-epoxyeicosatrienoic acid (14,15-EET) can trigger the growth of dormantmicrometastases by inducing neutrophilic infiltration and converting neutrophilfunction. 14,15-EET triggered neutrophil infiltration in metastatic lesions byactivating STAT3 and JNK pathways to induce the expression of human IL-8 andmurine CXCL15 in corresponding tumor cells. The continuous expression ofhIL-8/mCXCL15 was maintained by the sustained and enhanced activation of JNKpathway. 14,15-EET up-regulated miR-155 expression by activating STAT3 and JNKpathways. miR-155 in turn down-regulated the expression of SHIP1 and DET1, thusaugmenting the activation of JNK and c-Jun. Moreover, the function ofneutrophils was converted from tumor-suppressing to tumor-promoting by14,15-EET in vivo. By inducing the production of G-CSF/IL-6 in vivo, 14,15-EET induced the enhancement of STAT3 activation in neutrophilsto increase MMP-9 expression and decrease TRAIL expression. Neutrophil-derivedMMP-9 was required for 14,15-EET to induce angiogenesis during the growth ofdormant micrometastases. Depleting neutrophils or inhibiting hIL-8/mCXCL15up-regulation resulted in the failure of 14,15-EET to promote the developmentof micrometastases. These findings reveal a mechanism through which theinfiltration and tumor-promoting function of neutrophils could be induced totrigger the growth of dormant metastases, which might be a driving force forthe tumor recurrence based on dormant metastases. PMID:27270316

The Annexin a2 Promotes Development in Arthritis through Neovascularization by Amplification Hedgehog Pathway.

PubMed

Yi, Jun; Zhu, Yan; Jia, Yin; Jiang, Hongdie; Zheng, Xin; Liu, Dejing; Gao, Shunxiang; Sun, Mingjuan; Hu, Bo; Jiao, Binghua; Wang, Lianghua; Wang, Kaihui

2016-01-01

The neovascularization network of pannus formation plays a crucial role in the development of rheumatoid arthritis (RA). Annexin a2 (Axna2) is an important mediating agent that induces angiogenesis in vascular diseases. The correlation between Axna2 and pannus formation has not been studied. Here, we provided evidence that compared to osteoarthritis (OA) patients and healthy people, the expression of Axna2 and Axna2 receptor (Axna2R) were up-regulated in patients with RA. Joint swelling, inflammation and neovascularization were increased significantly in mice with collagen-induced arthritis (CIA) that were exogenously added Axna2. Cell experiments showed that Axna2 promoted HUVEC proliferation by binding Axna2R, and could activate Hedgehog (HH) signaling and up-regulate the expression of Ihh and Gli. Besides, expression of Ihh, Patched (Ptc), Smoothened (Smo) and Gli and matrix metalloproteinase-2 (MMP-2), vascular endothelial growth factor (VEGF) and angiopoietin-2 (Ang-2), angiogenic growth factor of HH signaling downstream, were down-regulated after inhibition of expression Axna2R on HUVEC. Together, our research definitely observed that over-expression of Axna2 could promote the development of CIA, especially during the process of pannus formation for the first time. Meanwhile, Axna2 depended on combining Axna2R to activate and enlarge HH signaling and the expression of its downstream VEGF, Ang-2 and MMP-2 to promote HUVEC proliferation, and eventually caused to angiogenesis. Therefore, the role of Axna2 is instructive for understanding the development of RA, suppress the effect of Axna2 might provide a new potential measure for treatment of RA.

Effect of ginseng extract on the TGF-β1 signaling pathway in CCl4-induced liver fibrosis in rats.

PubMed

Hafez, Mohamed M; Hamed, Sherifa S; El-Khadragy, Manal F; Hassan, Zeinab K; Al Rejaie, Salim S; Sayed-Ahmed, Mohamed M; Al-Harbi, Naif O; Al-Hosaini, Khalid A; Al-Harbi, Mohamed M; Alhoshani, Ali R; Al-Shabanah, Othman A; Alsharari, Shakir Dekhal

2017-01-13

Liver diseases are major global health problems. Ginseng extract has antioxidant, immune-modulatory and anti-inflammatory activities. This study investigated the effect of ginseng extract on carbon tetrachloride (CCl4)-induced liver fibrosis in rats. Male Wistar rats were divided into four groups: control group, ginseng group, CCl 4 group and CCl 4  + ginseng group. Liver injury was induced by the intraperitoneal (I.P) injection of 3 ml/kg CCl 4 (30% in olive oil) weekly for 8 weeks. The control group was I.P injected with olive oil. The expression of genes encoding transforming growth factor beta (TGF-β), type I TGF-β receptor (TβR-1), type II TGF-β receptor (TβR-II), mothers against decapentaplegic homolog 2 (Smad2), Smad3, Smad4, matrix metalloproteinase 2 (MMP2), MMP9, tissue inhibitor matrix metalloproteinase-1 (TIMP-1), Collagen 1a2 (Col1a2), Collagen 3a1 (Col3a1), interleukin-8 (IL-8) and interleukin -10 (IL-10) were measured by real-time PCR. Treatment with ginseng extract decreased hepatic fat deposition and lowered hepatic reticular fiber accumulation compared with the CCl 4 group. The CCl 4 group showed a significant increase in hepatotoxicity biomarkers and up-regulation of the expression of genes encoding TGF-β, TβR-I, TβR-II, MMP2, MMP9, Smad-2,-3, -4, and IL-8 compared with the control group. However, CCl 4 administration resulted in the significant down-regulation of IL-10 mRNA expression compared with the control group. Interestingly, ginseng extract supplementation completely reversed the biochemical markers of hepatotoxicity and the gene expression alterations induced by CCl 4 . ginseng extract had an anti-fibrosis effect via the regulation of the TGF-β1/Smad signaling pathway in the CCl 4 -induced liver fibrosis model. The major target was the inhibition of the expression of TGF-β1, Smad2, and Smad3.

Differential expression of folate receptor 1 in medulloblastoma and the correlation with clinicopathological characters and target therapeutic potential.

PubMed

Liu, Hailong; Sun, Qianwen; Zhang, Mingshan; Zhang, Zhihua; Fan, Xinyi; Yuan, Hongyu; Li, Cheng; Guo, Yuduo; Ning, Weihai; Sun, Youliang; Song, Yongmei; Yu, Chunjiang

2017-04-04

Medulloblastoma is the most common malignant brain tumor in children. Folate receptor 1 (Folr1) was abundantly expressed in some epithelial malignancies. However the expression profile and the role of clinicopathological significance and therapeutic target potential in medulloblastoma still remain elusive. Currently we detected the expression of Folr1 in medulloblastoma and identified the diagnostic application by evaluating the clinical, pathological and neuroimaging values. Then we developed a target therapeutic compound with Folr1, which exhibited promising efficiency in treatment of medulloblastoma. Folr1 expression was up-regulated in medulloblastoma and positively correlated with percentage of Ki-67 and MMP9 labeling, pathological subtypes, serum Folr1 levels and CSF spreading on MRI. The level of serum Folr1 showed rational sensitivity and specificity in predicting histological subgroups. Strong Folr1 expression was recommended as the independent value regarding the prognosis of patients with medulloblastoma. Folr1 targeted therapy attenuated the tumor growth and metastasis with down-regulation of MMPs proteins and activation of apoptosis. Immunostaining analysis in the xenograft samples showed the decreased Ki-67 and MMP9 index providing the strong evidences that Folr1 targeted application can suppress the proliferation and invasion. Our findings uncovered in Folr1 a predictive candidate and therapeutic target for medulloblastoma.

Keratin 8 and 18 Loss in Epithelial Cancer Cells Increases Collective Cell Migration and Cisplatin Sensitivity through Claudin1 Up-regulation*

PubMed Central

Fortier, Anne-Marie; Asselin, Eric; Cadrin, Monique

2013-01-01

Keratins 8 and 18 (K8/18) are simple epithelial cell-specific intermediate filament proteins. Keratins are essential for tissue integrity and are involved in intracellular signaling pathways that regulate cell response to injuries, cell growth, and death. K8/18 expression is maintained during tumorigenesis; hence, they are used as a diagnostic marker in tumor pathology. In recent years, studies have provided evidence that keratins should be considered not only as markers but also as regulators of cancer cell signaling. The loss of K8/18 expression during epithelial-mesenchymal transition (EMT) is associated with metastasis and chemoresistance. In the present study, we investigated whether K8/18 expression plays an active role in EMT. We show that K8/18 stable knockdown using shRNA increased collective migration and invasiveness of epithelial cancer cells without modulating EMT markers. K8/18-depleted cells showed PI3K/Akt/NF-κB hyperactivation and increased MMP2 and MMP9 expression. K8/18 deletion also increased cisplatin-induced apoptosis. Increased Fas receptor membrane targeting suggests that apoptosis is enhanced via the extrinsic pathway. Interestingly, we identified the tight junction protein claudin1 as a regulator of these processes. This is the first indication that modulation of K8/18 expression can influence the phenotype of epithelial cancer cells at a transcriptional level and supports the hypothesis that keratins play an active role in cancer progression. PMID:23449973

Repeated whiskey binges promote liver injury in rats fed a choline-deficient diet.

PubMed

Nieto, Natalia; Rojkind, Marcos

2007-02-01

Alcoholic liver disease is associated with nutritional deficiency and it may aggravate within the context of fatty liver. We investigated the relationship between alcohol intake (whiskey binge drinking) and a choline-deficient diet (CD) and assessed whether stellate cells could contribute to liver injury in this model. Rats fed the CD diet plus whiskey showed increased liver damage compared to rats fed the CD diet, as demonstrated by H&E staining, elevated transaminases, steatosis, TNF-alpha levels, enhanced CYP2E1 activity, impaired antioxidant defense, elevated lipid peroxidation, and protein carbonyls. The combined treatment triggered an apoptotic response as determined by elevated Bax, caspase-3 activity, cytochrome-c release, and decreased Bcl-2 and Bcl-XL. Stellate cells were activated as increased expression of alpha-Sma was observed over that by the CD diet alone. The combined treatment shifted extracellular matrix remodeling towards a pro-fibrogenic response due to up-regulation of collagen I, TIMP1, and Hsp47 proteins, along with down-regulation of MMP13, MMP2, and MMP9 expression, proteases which degrade collagen I. These events were accompanied by increased phosphorylation of p38, a kinase that elevates collagen I. Repeated alcohol binges in the context of mild steatosis may promote activation of stellate cells and contribute to liver injury.

Matrilysin (Matrix Metalloproteinase-7) Regulates Anti-Inflammatory and Antifibrotic Pulmonary Dendritic Cells That Express CD103 (αEβ7-Integrin)

PubMed Central

Manicone, Anne M.; Huizar, Isham; McGuire, John K.

2009-01-01

The E-cadherin receptor CD103 (αEβ7-integrin) is expressed on specific populations of pulmonary dendritic cells (DC) and T cells. However, CD103 function in the lung is not well understood. Matrilysin (MMP-7) expression is increased in lung injury and cleaves E-cadherin from injured lung epithelium. Thus, to assess matrilysin effects on CD103-E-cadherin interactions in lung injury, wild-type, CD103−/−, and Mmp7−/− mice, in which E-cadherin isn’t cleaved in the lung, were treated with bleomycin or bleomycin with nFMLP to reverse the defect in acute neutrophil influx seen in Mmp7−/− mice. Pulmonary CD103+ DC were significantly increased in injured wild-type compared with Mmp7−/− mice, and CD103+ leukocytes showed significantly enhanced interaction with E-cadherin on injured wild-type epithelium than with Mmp7−/− epithelium in vitro and in vivo. Bleomycin-treated CD103−/− mice had persistent neutrophilic inflammation, increased fibrosis, and increased mortality compared with wild-type mice, a phenotype that was partially recapitulated in bleomycin/nFMLP-treated Mmp7−/− mice. Soluble E-cadherin increased IL-12 and IL-10 and reduced IL-6 mRNA expression in wild-type bone marrow-derived DC but not in CD103−/− bone marrow-derived DC. Similar mRNA patterns were seen in lungs of bleomycin-injured wild-type, but not CD103−/− or Mmp7−/−, mice. In conclusion, matrilysin regulates pulmonary localization of DC that express CD103, and E-cadherin cleavage may activate CD103+ DC to limit inflammation and inhibit fibrosis. PMID:19893044

Roles of the cyclooxygenase 2 matrix metalloproteinase 1 pathway in brain metastasis of breast cancer.

PubMed

Wu, Kerui; Fukuda, Koji; Xing, Fei; Zhang, Yingyu; Sharma, Sambad; Liu, Yin; Chan, Michael D; Zhou, Xiaobo; Qasem, Shadi A; Pochampally, Radhika; Mo, Yin-Yuan; Watabe, Kounosuke

2015-04-10

Brain is one of the major sites of metastasis in breast cancer; however, the pathological mechanism of brain metastasis is poorly understood. One of the critical rate-limiting steps of brain metastasis is the breaching of blood-brain barrier, which acts as a selective interface between the circulation and the central nervous system, and this process is considered to involve tumor-secreted proteinases. We analyzed clinical significance of 21 matrix metalloproteinases on brain metastasis-free survival of breast cancer followed by verification in brain metastatic cell lines and found that only matrix metalloproteinase 1 (MMP1) is significantly correlated with brain metastasis. We have shown that MMP1 is highly expressed in brain metastatic cells and is capable of degrading Claudin and Occludin but not Zo-1, which are key components of blood-brain barrier. Knockdown of MMP1 in brain metastatic cells significantly suppressed their ability of brain metastasis in vivo, whereas ectopic expression of MMP1 significantly increased the brain metastatic ability of the cells that are not brain metastatic. We also found that COX2 was highly up-regulated in brain metastatic cells and that COX2-induced prostaglandins were directly able to promote the expression of MMP1 followed by augmenting brain metastasis. Furthermore, we found that COX2 and prostaglandin were able to activate astrocytes to release chemokine (C-C motif) ligand 7 (CCL7), which in turn promoted self-renewal of tumor-initiating cells in the brain and that knockdown of COX2 significantly reduced the brain metastatic ability of tumor cells. Our results suggest the COX2-MMP1/CCL7 axis as a novel therapeutic target for brain metastasis. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

Roles of the Cyclooxygenase 2 Matrix Metalloproteinase 1 Pathway in Brain Metastasis of Breast Cancer*

PubMed Central

Wu, Kerui; Fukuda, Koji; Xing, Fei; Zhang, Yingyu; Sharma, Sambad; Liu, Yin; Chan, Michael D.; Zhou, Xiaobo; Qasem, Shadi A.; Pochampally, Radhika; Mo, Yin-Yuan; Watabe, Kounosuke

2015-01-01

Brain is one of the major sites of metastasis in breast cancer; however, the pathological mechanism of brain metastasis is poorly understood. One of the critical rate-limiting steps of brain metastasis is the breaching of blood-brain barrier, which acts as a selective interface between the circulation and the central nervous system, and this process is considered to involve tumor-secreted proteinases. We analyzed clinical significance of 21 matrix metalloproteinases on brain metastasis-free survival of breast cancer followed by verification in brain metastatic cell lines and found that only matrix metalloproteinase 1 (MMP1) is significantly correlated with brain metastasis. We have shown that MMP1 is highly expressed in brain metastatic cells and is capable of degrading Claudin and Occludin but not Zo-1, which are key components of blood-brain barrier. Knockdown of MMP1 in brain metastatic cells significantly suppressed their ability of brain metastasis in vivo, whereas ectopic expression of MMP1 significantly increased the brain metastatic ability of the cells that are not brain metastatic. We also found that COX2 was highly up-regulated in brain metastatic cells and that COX2-induced prostaglandins were directly able to promote the expression of MMP1 followed by augmenting brain metastasis. Furthermore, we found that COX2 and prostaglandin were able to activate astrocytes to release chemokine (C-C motif) ligand 7 (CCL7), which in turn promoted self-renewal of tumor-initiating cells in the brain and that knockdown of COX2 significantly reduced the brain metastatic ability of tumor cells. Our results suggest the COX2-MMP1/CCL7 axis as a novel therapeutic target for brain metastasis. PMID:25691572

Differential expression of extracellular matrix constituents and cell adhesion molecules between malignant pleural mesothelioma and mesothelial hyperplasia.

PubMed

Alì, Greta; Borrelli, Nicla; Riccardo, Giannini; Proietti, Agnese; Pelliccioni, Serena; Niccoli, Cristina; Boldrini, Laura; Lucchi, Marco; Mussi, Alfredo; Fontanini, Gabriella

2013-11-01

Malignant pleural mesothelioma (MPM) is a highly aggressive neoplasm associated with asbestos exposure. Currently, the molecular mechanisms that induce MPM development are still unknown. The purpose of this study was to identify new molecular biomarkers for mesothelial carcinogenesis. We analyzed a panel of 84 genes involved in extracellular matrix remodeling and cell adhesion by polymerase chain reaction (PCR) array in 15 samples of epithelioid mesothelioma and 10 samples of reactive mesothelial hyperplasia (MH; 3 of 25 samples were inadequate for mRNA analysis). To validate the differentially expressed genes identified by PCR array, we analyzed 27 more samples by immunohistochemistry, in addition to the 25 samples already studied. Twenty-five genes were differentially expressed in MPM and MH by PCR array. Of these we studied matrix metalloproteinase 7 (MMP7), MMP14, CD44, and integrin, alpha3 expression by immunohistochemistry in 26 epithelioid MPM and 26 MH samples from the entire series of 52 cases. We observed higher MMP14 and integrin, alpha3 expression in MPM samples compared with MH samples (p = 0.000002 and p = 0.000002, respectively). Conversely, CD44 expression was low in most (57.7%) mesothelioma samples but only in 11.5% of the MH samples (p = 0.0013). As regards MMP7, we did not observe differential expression between MH and MPM samples. We have extensively studied genes involved in cell adhesion and extracellular matrix remodeling in MPM and MH samples, gaining new insight into the pathophysiology of mesothelioma. Moreover, our data suggest that these factors could be potential biomarkers for MPM.

Root extractive from Daphne genkwa benefits in wound healing of anal fistula through up-regulation of collagen genes in human skin fibroblasts.

PubMed

Yang, Dong; Xu, Jun-Hua; Shi, Ren-Jie

2017-04-30

Wound healing is the main problem in the therapy of anal fistula (AF). Daphne genkwa root has been traditionally used as an agent to soak sutures in operation of AF patients, but its function in wound healing remains largely unclear. The aim of the present study was to illuminate mechanisms of D. genkwa root treatment on AF. In the present study, 60 AF patients after surgery were randomly divided into two groups, external applied with or without the D. genkwa extractive. Wound healing times were compared and granulation tissues were collected. In vitro , we constructed damaged human skin fibroblasts (HSFs) with the treatment of TNF-α (10 μg/ml). Cell Count Kit-8 (CCK-8) and flow cytometry analysis were used to determine the effects of D. genkwa root extractive on cell viability, cell cycle and apoptosis of damaged HSFs. Furthermore, protein levels of TGF-β, COL1A1, COL3A1, Timp-1 , matrix metalloproteinase (MMP)-3 ( MMP-3 ) and MEK/ERK signalling pathways were investigated both in vivo and in vitro Results showed that D. genkwa root extractive greatly shortens the wound healing time in AF patients. In granulation tissues and HSFs, treatment with the extractive significantly elevated the expressions of COL1A1, COL3A1, Timp-1, c-fos and Cyclin D1 , while reduced the expression of MMP-3 Further detection presented that MEK/ERK signalling was activated after the stimulation of extractive in HSFs. Our study demonstrated that extractive from D. genkwa root could effectively improve wound healing in patients with AF via the up-regulation of fibroblast proliferation and expressions of COL1A1 and COL3A1 . © 2017 The Author(s).

Effect of fluoroquinolones on the expression of matrix metalloproteinase in debrided cornea of rats.

PubMed

Sharma, Charu; Velpandian, Thirumurthy; Baskar Singh, Sundararajan; Ranjan Biswas, Nihar; Bihari Vajpayee, Rasik; Ghose, Supriyo

2011-01-01

Matrix metalloproteinases (MMPs) are implicated in regenerative and healing processes in corneal injuries. Based upon reports that topical fluoroquinolones (FQs) may cause perforations during corneal healing by modulating MMPs, this study evaluated the comparative effects of commercially available FQs eye drops on the expression of MMP-2 and MMP-9 in the cornea after ethanol injury. Uniform corneal epithelial defects were created using 70% ethanol in the right eye of the rats (n = 6). The groups studied were (I) sham, (II) normal saline with benzalkonium chloride (NS-BKC), (III) norfloxacin 0.3%, (IV) ciprofloxacin 0.3%, (V) lomefloxacin 0.3%, (VI) sparfloxacin 0.3%, (VII) gatifloxacin 0.3%, and (VIII) moxifloxacin 0.5%. Each treatment was instilled six times/day up to 48 h and rats were sacrificed using excess of anesthesia. The corneas were excised to study the expression of MMP-2 and MMP-9 using gelatin zymography and real-time PCR. All the FQs significantly increased the expression of MMP-2 and MMP-9 as compared to the sham and NS-BKC-treated group. NS-BKC did not show a significant effect on MMPs expression compared to the sham group. Among the studied FQs, ciprofloxacin was observed to exhibit maximal induction of MMP-2 and MMP-9, whereas lomefloxacin exhibited an equivocal effect on both MMP-2 and MMP-9 expression. Findings of the present study demonstrate that topical application of FQs may induce the expression of MMP-2 and MMP-9 in debrided corneal epithelium and, therefore, may delay corneal wound healing. Thus, it can be concluded that selecting a FQ for ophthalmic use having minimal effect on MMPs may impact wound healing in injured or vulnerable cornea.

Cytokine expression profile in the bone-anchored hearing system: 12-week results from a prospective randomized, controlled study.

PubMed

Calon, Tim George Ate; van Tongeren, Joost; Omar, Omar; Johansson, Martin Lars; Stokroos, Robert-Jan

2018-04-27

To study the effect of implanting the percutaneous bone-anchored hearing system (BAHS) itself and inflammation of the peri-abutment skin warrant clarification. In this study, we aimed to acquire further insight into the immune responses related to BAHS surgery and peri-implant skin inflammation. During surgery and 12 weeks post-implantation, skin biopsies were obtained. If applicable, additional biopsies were taken during cases of inflammation. The mRNA expression of IL-1β, IL-6, IL-8, TNFα, IL-17, IL-10, TGF-ß, MIP-1α, MMP-9, TIMP-1, COL1α1, VEGF-A, FGF-2 TLR-2, and TLR-4 was quantified using qRT-PCR. Thirty-five patients agreed to the surgery and 12-week biopsy. Twenty-two patients had mRNA of sufficient quality for analysis. Ten were fitted with a BAHS using the minimally invasive Ponto surgery technique. Twelve were fitted with a BAHS using the linear incision technique with soft-tissue preservation. Five biopsies were obtained during episodes of inflammation. The post-implantation mRNA expression of IL-1β (P = .002), IL-8 (P = .003), MMP9 (P = .005), TIMP-1 (P = .002), and COL1α1 (P < .001) was significantly up-regulated. IL-6 (P = .009) and FGF-2 (P = .004) mRNA expression was significantly down-regulated after implantation. Within patients, no difference between post-implantation mRNA expression (at 12 weeks) and when inflammation was observed. Between patients, the expression of IL-1β (P = .015) and IL-17 (P = .02) was higher during cases of inflammation compared with patients who had no inflammation at 12-week follow-up. As part of a randomized, prospective, clinical trial, the present study reports the molecular profile of selected cytokines in the soft tissue around BAHS. Within the limit of this study, the results showed that 12 weeks after BAHS implantation the gene expression of some inflammatory cytokines (IL-8 and IL-1β) is still relatively high compared with the baseline, steady-state, expression. The up-regulation of anabolic (COL1α1) and tissue-remodeling (MMP-9 and TIMP1) genes indicates an ongoing remodeling process after 12 weeks of implantation. The results suggest that IL-1β, IL-17, and TNF-α may be interesting markers associated with inflammation. © 2018 The Authors. Clinical Implant Dentistry and Related Research published by Wiley Periodicals, Inc.

Hydrostatic pressure influences HIF-2 alpha expression in chondrocytes.

PubMed

Inoue, Hiroaki; Arai, Yuji; Kishida, Tsunao; Terauchi, Ryu; Honjo, Kuniaki; Nakagawa, Shuji; Tsuchida, Shinji; Matsuki, Tomohiro; Ueshima, Keiichirou; Fujiwara, Hiroyoshi; Mazda, Osam; Kubo, Toshikazu

2015-01-05

Hypoxia-inducible factor (HIF)-2α is considered to play a major role in the progression of osteoarthritis. Recently, it was reported that pressure amplitude influences HIF-2α expression in murine endothelial cells. We examined whether hydrostatic pressure is involved in expression of HIF-2α in articular chondrocytes. Chondrocytes were cultured and stimulated by inflammation or hydrostatic pressure of 0, 5, 10, or 50 MPa. After stimulation, heat shock protein (HSP) 70, HIF-2α, nuclear factor kappa B (NF-κB), matrix metalloproteinase (MMP)-13, MMP-3, and vascular endothelial growth factor (VEGF) gene expression were evaluated. The levels of all gene expression were increased by inflammatory stress. When chondrocytes were exposed to a hydrostatic pressure of 5 MPa, HIF-2α, MMP-13, and MMP-3 gene expression increased significantly although those of HSP70 and NF-κB were not significantly different from the control group. In contrast, HIF-2α gene expression did not increase under a hydrostatic pressure of 50 MPa although HSP70 and NF-κB expression increased significantly compared to control. We considered that hydrostatic pressure of 5 MPa could regulate HIF-2α independent of NF-κB, because the level of HIF-2α gene expression increased significantly without upregulation of NF-κB expression at 5 MPa. Hydrostatic pressure may influence cartilage degeneration, inducing MMP-13 and MMP-3 expression through HIF-2α.

Hsa-miR-134 suppresses non-small cell lung cancer (NSCLC) development through down-regulation of CCND1

PubMed Central

Sun, Cheng-Cao; Li, Shu-Jun; Li, De-Jia

2016-01-01

Hsa-miRNA-134 (miR-134) has recently been discovered to have anticancer efficacy in different organs. However, the role of miR-134 on non-small cell lung cancer (NSCLC) is still ambiguous. In this study, we investigated the role of miR-134 on the development of NSCLC. The results indicated that miR-134 was significantly down-regulated in primary tumor tissues and very low levels were found in NSCLC cell lines. Ectopic expression of miR-134 in NSCLC cell lines significantly suppressed cell growth as evidenced by cell viability assay, colony formation assay and BrdU staining, through inhibition of cyclin D1, cyclin D2, CDK4 and up-regulation of p57(Kip2) and p21(Waf1/Cip1). In addition, miR-134 induced apoptosis, as indicated by concomitantly with up-regulation of key apoptosis protein cleaved caspase-3, and down-regulation of anti-apoptosis protein Bcl2. Moreover, miR-134 inhibited cellular migration and invasiveness through inhibition of matrix metalloproteinases (MMP)-7 and MMP-9. Further, oncogene CCND1 was revealed to be a putative target of miR-134, which was inversely correlated with miR-134 expression in NSCLC. Taken together, our results demonstrated that miR-134 played a pivotal role on NSCLC through inhibiting cell proliferation, migration, invasion, and promoting apoptosis by targeting oncogenic CCND1. PMID:27166267

[The expression of MMP-2 and MMP-9 in adenoid cystic carcinoma of lacrimal gland].

PubMed

Zhang, Lei; Zhang, Hong; Song, Guo-xiang; Lin, Ting-ting; Xu, Guang-chang; Zhu, Li-min

2013-01-01

To investigate the expression of matrix metal proteinase (MMP)-2 and MMP-9 in adenoid cystic carcinoma of lacrimal gland as well as their relation with biological behaviour of adenoid cystic carcinoma. Experimental study. The research objects were 60 cases of adenoid cystic carcinoma of lacrimal gland which were collected from No.2 Hospital of Tianjin Medical University from January 1991 to Jule 2011. There were 25 males and 35 females aged from 29 to 42 years. Based on histological revision, there were 36 cases of cribriform-tubular subtype and 24 cases of solid subtype. Forty-five cases were primary lesions and 15 cases were recurrent lesions. Ten samples of normal lacrimal gland around polymorphic adenoma were selected as the control group. The expression of CD105, MMP-2 and MMP-9 were evaluated by immunohistochemistry. The microvessel density (MVD) was defined by expression of CD105. One way ANOVA, χ(2)-test and spearman correlation test were used to analyzed the data. The number of MVD [(17.71 ± 5.63)/100 folds field of vision] and the positive rates of MMP-2 (45.0%, 27/60) and MMP-9 (55.0%, 33/60) in the samples of adenoid cystic carcinoma of lacrimal gland were higher than those in the normal lacrimal gland [the number of MVD was (0.70 ± 0.95)/100 folds field of vision, the expressions of MMP-2 and MMP-9 were negative] (t' = 2.039, P < 0.05; χ(2) = 5.550, P < 0.05; χ(2) = 8.315, P < 0.01), the solid subtypes had more MVD [(26.12 ± 5.32)/100 folds field of vision] and higher positive rates of MMP-2 (62.5%, 15/24) and MMP-9 (79.2%, 19/24) than the cribriform-tubular subtypes (t' = 2.060, P < 0.05; χ(2) = 4.950, P < 0.05; χ(2) = 9.439, P < 0.05); the recurrent lesions had more MVD and higher positive rate of MMP-2 and MMP-9 than the primary lesions (t' = 2.129, P < 0.05; χ(2) = 9.899, P < 0.05; χ(2) = 8.103, P < 0.05). The number of MVD in ACC of lacrimal gland patients was correlated with the positive rate of MMP-2 and MMP-9 respectively (rs = 0.636, P < 0.05; rs = 0.524, P < 0.05). The number of MVD and the expression of MMP-2 and MMP-9 are higher level in adenoid cystic carcinoma of lacrimal gland and are significantly correlated with pathological type and recurrence. Detecting the number of MVD and the expression of MMP-2 and MMP-9 may become biological indexes for malignancy, recurrence and metastasis of adenoid cystic carcinoma of lacrimal gland.

Proliferative effects of apical, but not basal, matrix metalloproteinase-7 activity in polarized MDCK cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Harrell, Permila C.; McCawley, Lisa J.; Fingleton, Barbara

2005-02-15

Matrix metalloproteinase-7 (MMP-7) is primarily expressed in glandular epithelium. Therefore, its mechanism of action may be influenced by its regulated vectorial release to either the apical and/or basolateral compartments, where it would act on its various substrates. To gain a better understanding of where MMP-7 is released in polarized epithelium, we have analyzed its pattern of secretion in polarized MDCK cells expressing stably transfected human MMP-7 (MDCK-MMP-7), and HCA-7 and Caco2 human colon cancer cell lines. In all cell lines, latent MMP-7 was secreted to both cellular compartments, but was 1.5- to 3-fold more abundant in the basolateral compartment asmore » compared to the apical. However, studies in the MDCK system demonstrated that MMP-7 activity was 2-fold greater in the apical compartment of MDCK-MMP-7{sup HIGH}-polarized monolayers, which suggests the apical co-release of an MMP-7 activator. In functional assays, MMP-7 over-expression increased cell saturation density as a result of increased cell proliferation with no effect on apoptosis. Apical MMP-7 activity was shown to be responsible for the proliferative effect, which occurred, as demonstrated by media transfer experiments, through cleavage of an apical substrate and not through the generation of a soluble factor. Taken together, our findings demonstrate the importance of MMP-7 secretion in relation to its mechanism of action when expressed in a polarized epithelium.« less

Microarray Analysis of Gene Expression Alteration in Human Middle Ear Epithelial Cells Induced by Asian Sand Dust.

PubMed

Go, Yoon Young; Park, Moo Kyun; Kwon, Jee Young; Seo, Young Rok; Chae, Sung-Won; Song, Jae-Jun

2015-12-01

The primary aim of this study is to evaluate the gene expression profile of Asian sand dust (ASD)-treated human middle ear epithelial cell (HMEEC) using microarray analysis. The HMEEC was treated with ASD (400 µg/mL) and total RNA was extracted for microarray analysis. Molecular pathways among differentially expressed genes were further analyzed. For selected genes, the changes in gene expression were confirmed by real-time polymerase chain reaction. A total of 1,274 genes were differentially expressed by ASD. Among them, 1,138 genes were 2 folds up-regulated, whereas 136 genes were 2 folds down-regulated. Up-regulated genes were mainly involved in cellular processes, including apoptosis, cell differentiation, and cell proliferation. Down-regulated genes affected cellular processes, including apoptosis, cell cycle, cell differentiation, and cell proliferation. The 10 genes including ADM, CCL5, EDN1, EGR1, FOS, GHRL, JUN, SOCS3, TNF, and TNFSF10 were identified as main modulators in up-regulated genes. A total of 11 genes including CSF3, DKK1, FOSL1, FST, TERT, MMP13, PTHLH, SPRY2, TGFBR2, THBS1, and TIMP1 acted as main components of pathway associated with 2-fold down regulated genes. We identified the differentially expressed genes in ASD-treated HMEEC. Our work indicates that air pollutant like ASD, may play an important role in the pathogenesis of otitis media.

[Impact of siRNA-mediated down-regulation of CD147 on human breast cancer cells].

PubMed

Li, Zhenqian; Li, Daoming; Li, Jiangwei; Huang, Pei; Qin, Hui

2015-10-01

To investigate the influence of siRNA-mediated down-regulation of CD147 on growth, proliferation and movement of human breast cancer cell line MDA-MB-231. The protein expression of CD147, MMP-2 and TIMP-2 of the MDA-MB-231 cells were analyzed by ABC. Lentiviral expression vector of CD147 gene was constructed and transfected into MDA-MB-231 cells. RT-PCR and Western blot were used to detect the mRNA and protein level changes of CD147 genes to identify the optimal time point, followed by detection of changes of mRNA and protein expression of MMP-2 and TIMP-2 genes. CCK-8 reagent method and cell scratch test were used to detect the proliferation and migration change of MDA-MB-231 cells. The nude mouse model of breast cancer by hypodermic injection with MDA-MB-231 cells was established to document the effect of CD147 siRNA on the tumor transplants. After transfection of lentiviral expression vector of CD147 gene, protein of CD147, MMP-2 and TIMP-2 were weakly or negative expressed, significantly weaker than those of control group (P < 0.01). After 72 hours of transfection, average down-regulation rate of CD147 and MMP-2 were 96.03% ± 0.84% and 96.03% ± 0.84%, respectively. Both CD147 mRNA and MMP-2 mRNA expression were down-regulated (P < 0.05), while TIMP-2 mRNA expression showed no significant deference (P > 0.05). No less than 2 days after transfection, cell growth of MDA-MB-231 cell line was found significantly inhibited (P < 0.05). After 24 hours of transection, average migration distance of MDA-MB-231 cell line and control group were (0.64 ± 0.12) mm and (4.69 ± 0.85) mm, respectively, which indicated a lower migrate speed. Down regulation of CD147 led to reduction of volume and mass of nude mouses. The growth of the carcinoma transplant was inhibited upon siRNA-mediated down-regulation of CD147 (P < 0.05), with an average tumor mass of (1.85 ± 0.98) g and both reduction of tumor size and tumor mass. CD147 may alter the MMP-2/TIMP-2 balance in MDA-MB-231 cells. CD147 gene silencing inhibits the proliferation and migration of MDA-MB-231 cells and the growth of carcinoma transplants in nude mice.

Expression of SLP-2 Was Associated with Invasion of Esophageal Squamous Cell Carcinoma

PubMed Central

Cao, Wenfeng; Zhang, Bin; Ding, Fang; Zhang, Weiran; Sun, Baocun; Liu, Zhihua

2013-01-01

Introduction Stomatin-like protein 2 (SLP-2), a member of the Stomatin superfamily, has been identified as an oncogenic-related protein and found to be up-regulated in multi-cancers. Nonetheless, the expression pattern and regulation of SLP-2 in human esophageal squamous cell carcinoma (ESCC) remain unexplored. Methods Immunohistochemistry and immunofluorescence staining analysis were performed to show SLP-2 expression and location. RNAi method was used to inhibit specific protein expression. Transwell assay was done to investigate cells invasive capability. RT-PCR and Western blot analysis were used to detect mRNA and protein expression levels. Results Immunohistochemical analysis showed that up-regulation of SLP-2 was found in invasive front compared with cancer central tissue in ESCC. Inhibition of SLP-2 by SLP-2 siRNA can decrease ESCC cells invasive capability through MMP-2 dependent manner. Up-regulation of SLP-2 was effectively abrogated by the ERK1/2 inhibitors either PD98059 or U0126, but no effect was showed by the treatment of AKT inhibitors either LY294002 or MK-2206. So the regulation of SLP-2 was involved in activation of the MAPK/ERK pathway. Conclusions We found that PMA/EGF could induce the up-regulated expression of SLP-2 probably through activating ERK signalling. The current study suggests that SLP-2 may represent an important molecular hallmark that is clinically relevant to the invasion of ESCC. PMID:23667687

Wogonin suppresses TNF-{alpha}-induced MMP-9 expression by blocking the NF-{kappa}B activation via MAPK signaling pathways in human aortic smooth muscle cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lee, Syng-Ook; Jeong, Yun-Jeong; Yu, Mi Hee

2006-12-08

Matrix metalloproteinase-9 (MMP-9) plays a major role in the pathogenesis of atherosclerosis and restenosis by regulating both migration and proliferation of vascular smooth muscle cells (VSMC) after an arterial injury. In this study, we examined the inhibitory effect of three major flavonoids in Scutellariae Radix, baicalin, baicalein, and wogonin, on TNF-{alpha}-induced MMP-9 expression in human aortic smooth muscle cells (HASMC). Wogonin, but not baicalin and baicalein, significantly and selectively suppressed TNF-{alpha}-induced MMP-9 expression in HASMC. Reporter gene, electrophoretic mobility shift, and Western blotting assays showed that wogonin inhibits MMP-9 gene transcriptional activity by blocking the activation of NF-{kappa}B via MAPKmore » signaling pathways. Moreover, the Matrigel migration assay showed that wogonin reduced TNF-{alpha}-induced HASMC migration. These results suggest that wogonin effectively suppresses TNF-{alpha}-induced HASMC migration through the selective inhibition of MMP-9 expression and represents a potential agent for the prevention of vascular disorders related to the migration of VSMC.« less

Effect of TGF-beta1 on MMP/TIMP and TGF-beta1 receptors in great saphenous veins and its significance on chronic venous insufficiency.

PubMed

Serralheiro, Pedro; Cairrão, Elisa; Maia, Cláudio J; João, Marina; Almeida, Carlos M Costa; Verde, Ignacio

2017-06-01

Objectives Transforming growth factor-beta1 (TGF-β1) may participate in local chronic inflammatory processes in varicose veins and in venous wall structure modifications through regulation of matrix metalloproteinases (MMP) and their inhibitors (tissue inhibitor of metalloproteinase (TIMP)). The aim of this study was to analyze the effect of TGF-β1 in the vein wall, namely on the gene expression of selected MMP, TIMP and TGF-β1 receptors. Methods Healthy vein samples were harvested from eight subjects who underwent coronary bypass graft surgery with great saphenous vein. Each vein sample was divided into two segments, which were cultivated separately in vitro (one of the segments had TGF-β1 added) and then submitted to gene expression analysis. Results In the TGF-β1 supplemented group, there was a general increase in the mean gene expression. Specifically, expression of MMP9, MMP12, TIMP1 and TIMP2 were statistically significant. Conclusion The results of this study demonstrate that the gene expression of MMP9, MMP12, TIMP1 and TIMP2 was influenced by the addition of TGF-β1. These results may be translated to chronic venous insufficiency framework and suggest involvement of TGF-β1 in the vein wall pathology.

Curcuma DMSO extracts and curcumin exhibit an anti-inflammatory and anti-catabolic effect on human intervertebral disc cells, possibly by influencing TLR2 expression and JNK activity

PubMed Central

2012-01-01

Background As proinflammatory cytokines seem to play a role in discogenic back pain, substances exhibiting anti-inflammatory effects on intervertebral disc cells may be used as minimal-invasive therapeutics for intradiscal/epidural injection. The purpose of this study was to investigate the anti-inflammatory and anti-catabolic potential of curcuma, which has been used in the Indian Ayurvedic medicine to treat multiple ailments for a long time. Methods Human disc cells were treated with IL-1β to induce an inflammatory/catabolic cascade. Different extracts of curcuma as well as curcumin (= a component selected based on results with curcuma extracts and HPLC/MS analysis) were tested for their ability to reduce mRNA expression of proinflammatory cytokines and matrix degrading enzymes after 6 hours (real-time RT-PCR), followed by analysis of typical inflammatory signaling mechanisms such as NF-κB (Western Blot, Transcription Factor Assay), MAP kinases (Western Blot) and Toll-like receptors (real-time RT-PCR). Quantitative data was statistically analyzed using a Mann Whitney U test with a significance level of p < 0.05 (two-tailed). Results Results indicate that the curcuma DMSO extract significantly reduced levels of IL-6, MMP1, MMP3 and MMP13. The DMSO-soluble component curcumin, whose occurrence within the DMSO extract was verified by HPLC/MS, reduced levels of IL-1β, IL-6, IL-8, MMP1, MMP3 and MMP13 and both caused an up-regulation of TNF-α. Pathway analysis indicated that curcumin did not show involvement of NF-κB, but down-regulated TLR2 expression and inhibited the MAP kinase JNK while activating p38 and ERK. Conclusions Based on its anti-inflammatory and anti-catabolic effects, intradiscal injection of curcumin may be an attractive treatment alternative. However, whether the anti-inflammatory properties in vitro lead to analgesia in vivo will need to be confirmed in an appropriate animal model. PMID:22909087

Inflammatory response in rat lungs with recurrent exposure to welding fumes: a transcriptomic approach.

PubMed

Oh, Jung-Hwa; Yang, Mi-Jin; Heo, Jeong-Doo; Yang, Young-Su; Park, Han-Jin; Park, Se-Myo; Kwon, Myung-Sang; Song, Chang-Woo; Yoon, Seokjoo; Yu, Il Je

2012-04-01

As chronic exposure to welding fumes causes pulmonary diseases, such as pneumoconiosis, public concern has increased regarding continued exposure to these hazardous gases in the workplace. In a previous study, the inflammatory response to welding fume exposure was analysed in rat lungs in the case of recurrent exposure and recovery periods. Thus using lung samples, well-annotated by histological observation and biochemical analysis, this study examines the gene expression profiles to identify phenotype-anchored genes corresponding to lung inflammation and the repair phenomenon after recurrent welding fume exposure. Seven genes (Mmp12, Cd5l, LOC50101, LOC69183, Spp1, and Slc26a4) were found to be significantly up-regulated according to the severity of the lung injury. In addition, the transcription and translation of Trem2, which was up-regulated in response to the repair process, were validated using a real-time polymerase chain reaction, Western blotting, and immunohistochemistry. The differentially expressed genes in the exposure and recovery groups were also classified using k-means and hierarchical clustering, plus their toxicological function and canonical pathways were further analysed using Ingenuity Pathways Analysis Software. As a result, this comprehensive and integrative analysis of the transcriptional changes that occur during repeated exposure provides important information on the inflammation and repair processes after welding-fume-induced lung injury.

Matrix Metalloproteinase-9 Is a Predictive Factor for Systematic Hypertension and Heart Dysfunction in Patients with Obstructive Sleep Apnea Syndrome

PubMed Central

Wang, Shuhui; Li, Shisheng; Wang, Bin; Liu, Jiajia

2018-01-01

Patients with obstructive sleep apnea syndrome (OSAS) showed higher prevalence in cardiovascular diseases due to aberrant hypoxia and oxidative stress. However, not all OSAS patients end up with cardiovascular disorders, and identification of novel biomarker will be invaluable for differentiating patients at risk. Here we tested the serum matrix metalloproteinase-9 (MMP-9) levels in 47 untreated OSAS patients and found that the MMP-9 level was positively correlated with severity of OSAS, which was consistent with hypoxia degree and duration. Besides, the MMP-9 level was higher in patients complicated with systematic hypertension (P < 0.001). Furthermore, we selected those OSAS patients without any cardiovascular dysfunction (n = 35) and followed up for up to five years. By the end of follow-up, 12 patients had hypertension onset and 3 patients had left ventricular hypertrophy. By analyzing the clinical outcomes with MMP-9 expression, we demonstrated that high serum MMP-9 in OSAS patients was a risk factor for occurrence of cardiovascular diseases. In addition, we cultured the vascular endothelial cells (VEC) from rat aorta in hypoxia condition to investigate whether MMP-9 was elevated due to hypoxia in OSAS patients. Cellular results revealed that the expression, secretion, and activity of MMP-9 were all upregulated by hypoxia and can cleave the beta2-adrenergic receptor (β2AR) on VEC surface. Our results not only determined MMP-9 as a risk factor for cardiovascular diseases in OSAS patients, but also showed the possible involvement of hypoxia-MMP-9-β2AR signaling axis. PMID:29693002

Urtica dioica extract suppresses miR-21 and metastasis-related genes in breast cancer.

PubMed

Mansoori, Behzad; Mohammadi, Ali; Hashemzadeh, Shahriar; Shirjang, Solmaz; Baradaran, Ali; Asadi, Milad; Doustvandi, Mohammad Amin; Baradaran, Behzad

2017-09-01

Breast cancer has a high prevalence among women worldwide. Tumor invasion and metastasis still remains an open issue that causes most of the therapeutic failures and remains the prime cause of patient mortality. Hence, there is an unmet need to develop the most effective therapeutic approach with the lowest side effects and highest cytotoxicity that will effectively arrest or eradicate metastasis. An MTT assay and scratch test were used to assess the cytotoxicity and migration effects of Urtica dioica on the breast cancer cells. The QRT-PCR was used to study the expression levels of miR-21, MMP1, MMP9, MMP13, CXCR4, vimentin, and E-cadherin. The results of gene expression in tumoral groups confirmed the overexpression of miR-21, MMP1, MMP9, MMP13, vimentin, and CXCR4, and the lower expression of E-cadherin compared to control groups (P<0.05). Moreover, the results of the MTT assay show that Urtica dioica significantly inhibited breast cancer cell proliferation. Moreover, findings from the scratch assay exhibited the inhibitory effects of Urtica dioica on the migration of breast cancer cell lines. Urtica dioica extract could inhibit cancer cell migration by regulating miR-21, MMP1, MMP9, MMP13, vimentin, CXCR4, and E-Cadherin. Moreover, our findings demonstrated that the extract could decrease miR-21 expression, which substantially lessens the overexpressed MMP1, MMP9, MMP13, vimentin, and CXCR4 and increases E-cadherin in the tumoral group. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

BMP2 induces PANC-1 cell invasion by MMP-2 overexpression through ROS and ERK.

PubMed

Liu, Jun; Ben, Qi-Wen; Yao, Wei-Yan; Zhang, Jian-Jun; Chen, Da-Fan; He, Xiang-Yi; Li, Lei; Yuan, Yao-Zong

2012-06-01

The emerging roles of bone morphogenetic proteins (BMPs) in the initiation and progression of multiple cancers have drawn great attention in cancer research. We hypothesized that BMP2 promotes cancer metastasis by modulating MMP-2 secretion and activity through intracellular ROS regulation and ERK activation in human pancreatic cancer. Our data show that stimulation of PANC-1 cells with BMP2 induced MMP-2 secretion and activation, associated with decreased E-cadherin expression, resulting in epithelial-to-mesenchymal transformation (EMT) and cell invasion. Blockade of ROS by the ROS scavenger, 2-MPG, abolished cell invasion, inhibited the EMT process and decreased MMP-2 expression, suggesting ROS accumulation caused an increase in MMP-2 expression in BMP2-stimulated PANC-1 cell invasion. Furthermore, treatment of PANC-1 cells with 2-MPG or ERK inhibitor PD98059 reduced the phosphorylation of ERK, resulting in attenuation of BMP2-induced cell invasion and MMP-2 activation. Taken together, these results suggest that BMP2 induces the cell invasion of PANC-1 cells by enhancing MMP-2 secretion and acting through ROS accumulation and ERK activation.

MT2-MMP-dependent release of collagen IV NC1 domains regulates submandibular gland branching morphogenesis.

PubMed

Rebustini, Ivan T; Myers, Christopher; Lassiter, Keyonica S; Surmak, Andrew; Szabova, Ludmila; Holmbeck, Kenn; Pedchenko, Vadim; Hudson, Billy G; Hoffman, Matthew P

2009-10-01

Proteolysis is essential during branching morphogenesis, but the roles of MT-MMPs and their proteolytic products are not clearly understood. Here, we discover that decreasing MT-MMP activity during submandibular gland branching morphogenesis decreases proliferation and increases collagen IV and MT-MMP expression. Specifically, reducing epithelial MT2-MMP profoundly decreases proliferation and morphogenesis, increases Col4a2 and intracellular accumulation of collagen IV, and decreases the proteolytic release of collagen IV NC1 domains. Importantly, we demonstrate the presence of collagen IV NC1 domains in developing tissue. Furthermore, recombinant collagen IV NC1 domains rescue branching morphogenesis after MT2-siRNA treatment, increasing MT-MMP and proproliferative gene expression via beta1 integrin and PI3K-AKT signaling. Additionally, HBEGF also rescues MT2-siRNA treatment, increasing NC1 domain release, proliferation, and MT2-MMP and Hbegf expression. Our studies provide mechanistic insight into how MT2-MMP-dependent release of bioactive NC1 domains from collagen IV is critical for integrating collagen IV synthesis and proteolysis with epithelial proliferation during branching morphogenesis.

Neisseria gonorrhoeae Challenge Increases Matrix Metalloproteinase-8 Expression in Fallopian Tube Explants.

PubMed

Juica, Natalia E; Rodas, Paula I; Solar, Paula; Borda, Paula; Vargas, Renato; Muñoz, Cristobal; Paredes, Rodolfo; Christodoulides, Myron; Velasquez, Luis A

2017-01-01

Background: Neisseria gonorrhoeae (Ngo) is the etiological agent of gonorrhea, a sexually transmitted infection that initially infects the female lower genital tract. In untreated women, the bacteria can ascend to the upper genital reproductive tract and infect the fallopian tube (FTs), which is associated with salpingitis and can lead to impaired FT function and infertility. The extracellular matrix (ECM) plays an important role in cell migration and differentiation in the female genital tract, and some pathogens modify the ECM to establish successful infections. The ECM is regulated by matrix metalloproteinases (MMPs) and tissue inhibitors of metalloproteinases (TIMPs), their endogenous inhibitors; MMP deregulation causes pathological conditions in a variety of tissues. Results: The aim of this work was to analyze the expression and localization of MMP-3, MMP-8, MMP-9, and TIMP-1 in FT explants during Ngo infection using real-time PCR, immunohistochemistry, zymography and ELISA. No significant variations in MMP-3, MMP-9, and TIMP-1 transcript levels were observed. In contrast, a significant increase ( p < 0.05) was observed for MMP-8 expression and was accompanied by stromal immunoreactivity in infected explants. ELISA results supported these findings and showed that MMP-8 release increased upon gonococcal infection. Conclusions: Our results indicate that gonococcal infection induces increased MMP-8 expression, which might contribute to FT damage during infection.

Matrix metalloproteinase-9 activates TGF-β and stimulates fibroblast contraction of collagen gels.

PubMed

Kobayashi, Tetsu; Kim, HuiJung; Liu, Xiangde; Sugiura, Hisatoshi; Kohyama, Tadashi; Fang, Qiuhong; Wen, Fu-Qiang; Abe, Shinji; Wang, Xingqi; Atkinson, Jeffrey J; Shipley, James M; Senior, Robert M; Rennard, Stephen I

2014-06-01

Matrix metalloproteinase-9 (MMP-9) is a matrix-degrading enzyme implicated in many biological processes, including inflammation. It is produced by many cells, including fibroblasts. When cultured in three-dimensional (3D) collagen gels, fibroblasts contract the surrounding matrix, a function that is thought to model the contraction that characterizes both normal wound repair and fibrosis. The current study was designed to evaluate the role of endogenously produced MMP-9 in fibroblast contraction of 3D collagen gels. Fibroblasts from mice lacking expression of MMP-9 and human lung fibroblasts (HFL-1) transfected with MMP-9 small-interfering RNA (siRNA) were used. Fibroblasts were cast into type I collagen gels and floated in culture medium with or without transforming growth factor (TGF)-β1 for 5 days. Gel size was determined daily using an image analysis system. Gels made from MMP-9 siRNA-treated human fibroblasts contracted less than control fibroblasts, as did fibroblasts incubated with a nonspecific MMP inhibitor. Similarly, fibroblasts cultured from MMP-9-deficient mice contracted gels less than did fibroblasts from control mice. Transfection of the MMP-9-deficient murine fibroblasts with a vector expressing murine MMP-9 restored contractile activity to MMP-9-deficient fibroblasts. Inhibition of MMP-9 reduced active TGF-β1 and reduced several TGF-β1-driven responses, including activity of a Smad3 reporter gene and production of fibronectin. Because TGF-β1 also drives fibroblast gel contraction, this suggests the mechanism for MMP-9 regulation of contraction is through the generation of active TGF-β1. This study provides direct evidence that endogenously produced MMP-9 has a role in regulation of tissue contraction of 3D collagen gels mediated by fibroblasts. Copyright © 2014 the American Physiological Society.

Inhibition of EMMPRIN and MMP-9 Expression by Epigallocatechin-3-Gallate through 67-kDa Laminin Receptor in PMA-Induced Macrophages.

PubMed

Wang, Qi-Ming; Wang, Hao; Li, Ya-Fei; Xie, Zhi-Yong; Ma, Yao; Yan, Jian-Jun; Gao, Yi Fan Wei; Wang, Ze-Mu; Wang, Lian-Sheng

2016-01-01

It is well documented that overexpression of EMMPRIN (extracellular matrix metalloproteinase inducer) and MMPs (matrix metalloproteinases) by monocytes/macrophages plays an important role in atherosclerotic plaque rupture. Green tea polyphenol epigallocatechin-3-gallate (EGCG) has a variety of pharmacological properties and exerts cardiovascular protective effects. Recently, the 67-kD laminin receptor (67LR) has been identified as a cell surface receptor of EGCG. The aim of the present study was to evaluate the effects of EGCG on the expression of EMMPRIN and MMP-9 in PMA-induced macrophages, and the potential mechanisms underlying its effects. Human monocytic THP-1 cells were induced to differentiate into macrophages with phorbol 12-myristate 13-acetate (PMA). Protein expression and MMP-9 activity were assayed by Western blot and Gelatin zymography, respectively. Real-time PCR was used to examine EMMPRIN and MMP-9 mRNA expression. We showed that EGCG (10-50µmol/L) significantly inhibited the expression of EMMPRIN and MMP-9 and activation of extracellular signal-regulated kinase 1/2 (ERK1/2), p38 and c-Jun N-terminal kinase (JNK) in PMA-induced macrophages. Downregulation of EMMPRIN by gene silencing hindered PMA-induced MMP-9 secretion and expression, indicating an important role of EMMPRIN in the inhibition of MMP-9 by EGCG. Moreover, 67LR was involved in EGCG-mediated suppression of EMMPRIN and MMP-9 expression. Anti-67LR antibody treatment led to abrogation of the inhibitory action of EGCG on the expression of EMMPRIN and MMP-9 and activation of ERK1/2, p38, and JNK. Our results indicate that EGCG restrains EMMPRIN and MMP-9 expression via 67LR in PMA-induced macrophages, which also suggests that EGCG may be a possible therapeutic agent for stabilizing atherosclerotic plaque. © 2016 The Author(s) Published by S. Karger AG, Basel.

Effect of Thrombin on Human Amnion Mesenchymal Cells, Mouse Fetal Membranes, and Preterm Birth*

PubMed Central

Mogami, Haruta; Keller, Patrick W.; Shi, Haolin; Word, R. Ann

2014-01-01

Here, we investigated the effects of thrombin on matrix metalloproteinases (MMPs) and prostaglandin (PG) synthesis in fetal membranes. Thrombin activity was increased in human amnion from preterm deliveries. Treatment of mesenchymal, but not epithelial, cells with thrombin resulted in increased MMP-1 and MMP-9 mRNA and enzymatic activity. Thrombin also increased COX2 mRNA and PGE2 in these cells. Protease-activated receptor-1 (PAR-1) was localized to amnion mesenchymal and decidual cells. PAR-1-specific inhibitors and activating peptides indicated that thrombin-induced up-regulation of MMP-9 was mediated via PAR-1. In contrast, thrombin-induced up-regulation of MMP-1 and COX-2 was mediated through Toll-like receptor-4, possibly through thrombin-induced release of soluble fetal fibronectin. In vivo, thrombin-injected pregnant mice delivered preterm. Mmp8, Mmp9, and Mmp13, and PGE2 content was increased significantly in fetal membranes from thrombin-injected animals. These results indicate that thrombin acts through multiple mechanisms to activate MMPs and PGE2 synthesis in amnion. PMID:24652285

Epithelial expression of extracellular matrix metalloproteinase inducer/CD147 and matrix metalloproteinase-2 in neoplasms and precursor lesions derived from cutaneous squamous cells: An immunohistochemical study.

PubMed

Ayva, Sebnem Kupana; Karabulut, Ayse Anil; Akatli, Ayşe Nur; Atasoy, Pinar; Bozdogan, Onder

2013-10-01

Extracellular matrix metalloproteinase inducer (CD147) is a transmembrane glycoprotein involved in the regulation of matrix metalloproteinases (MMPs). The study investigated CD147 and MMP-2 expression in epidermis of cutaneous squamous lesions. CD147 and MMP-2 expressions were evaluated immunohistochemically in 44 specimens: 18 actinic keratoses (AK), 6 squamous cell carcinomas in situ (SCCIS), 13 squamous cell carcinomas (SCC; peritumoral and invasive portions assessed), and 7 normal skins. Patterns of expression were assessed, with MMP-2 in nuclei (MMP-2n) and cytoplasm (MMP-2c) evaluated separately. The expression of each marker was quantified using a calculated immunohistochemical/histologic score (H-score). Correlations were analyzed for the marker H-scores in each study group. Associations between H-scores and histopathologic parameters were also evaluated. CD147 H-score was the highest in SCC (invasive islands), followed by AK, SCCIS, and control specimens, respectively. MMP-2n and MMP-2c H-scores were the highest in AK, followed by SCCIS, SCC, and control specimens, respectively. MMP-2c and MMP-2n H-scores were significantly higher in peritumoral epidermis than in invasive islands of SCC. MMP-2c and CD147 H-scores were positively correlated in the peritumoral SCCs. CD147 H-score was positively correlated with tumor differentiation in SCC. The findings suggest that overexpression of CD147 plays a role in the development of SCC. Copyright © 2013 Elsevier GmbH. All rights reserved.

Anthrax Toxin Receptor 2 Functions in ECM Homeostasis of the Murine Reproductive Tract and Promotes MMP Activity

PubMed Central

Reeves, Claire V.; Wang, Xing; Charles-Horvath, Pelisa C.; Vink, Joy Y.; Borisenko, Valeriya Y.; Young, John A. T.; Kitajewski, Jan K.

2012-01-01

Anthrax Toxin Receptor proteins function as receptors for anthrax toxin, however physiological activity remains unclear. To evaluate the biological role of Antxr2, we generated Antxr2−/− mice. Antxr2−/− mice were viable, however Antxr2 is required for parturition in young females and for preserving fertility in older female mice. Histological analysis of the uterus and cervix revealed aberrant deposition of extracellular matrix proteins such as type I collagen, type VI collagen and fibronectin. A marked disruption of both the circular and longitudinal myometrial cell layers was evident in Antxr2−/− mice. These changes progressed as the mice aged, resulting in a thickened, collagen dense, acellular stroma and the disappearance of normal uterine architecture. To investigate the molecular mechanism underlying the uterine fibrosis we performed immunoblotting for MMP2 using uterine lysates and zymography using conditioned medium from Antxr2−/− mouse embryonic fibroblasts and found reduced levels of activated MMP2 in both. This prompted us to investigate MT1-MMP status, as MMP2 processing is regulated by MT1-MMP. We found MT1-MMP activity, as measured by MMP2 processing and activation, was enhanced by expression of either ANTXR1 or ANTXR2. We identified an ANTXR2/MT1-MMP complex and demonstrated that MT1-MMP activity is dependent on ANTXR2 expression levels in cells. Thus, we have discovered that ANTXR1 and ANTXR2 function as positive regulators of MT1-MMP activity. PMID:22529944

Membrane Type 1–Matrix Metalloproteinase/Akt Signaling Axis Modulates TNF-α-Induced Procoagulant Activity and Apoptosis in Endothelial Cells

PubMed Central

Ohkawara, Hiroshi; Ishibashi, Toshiyuki; Sugimoto, Koichi; Ikeda, Kazuhiko; Ogawa, Kazuei; Takeishi, Yasuchika

2014-01-01

Membrane type 1–matrix metalloproteinase (MT1-MMP) functions as a signaling molecule in addition to a proteolytic enzyme. Our hypothesis was that MT1-MMP cooperates with protein kinase B (Akt) in tumor necrosis factor (TNF)-α-induced signaling pathways of vascular responses, including tissue factor (TF) procoagulant activity and endothelial apoptosis, in cultured human aortic endothelial cells (ECs). TNF-α (10 ng/mL) induced a decrease in Akt phosphorylation within 60 minutes in ECs. A chemical inhibitor of MMP, TIMP-2 and selective small interfering RNA (siRNA)-mediated suppression of MT1-MMP reversed TNF-α-triggered transient decrease of Akt phosphorylation within 60 minutes, suggesting that MT1-MMP may be a key regulator of Akt phosphorylation in TNF-α-stimulated ECs. In the downstream events, TNF-α increased TF antigen and activity, and suppressed the expression of thrombomodulin (TM) antigen. Inhibition of Akt markedly enhanced TNF-α-induced expression of TF antigen and activity, and further reduced the expression of TM antigen. Silencing of MT1-MMP by siRNA also reversed the changed expression of TF and TM induced by TNF-α. Moreover, TNF-α induced apoptosis of ECs through Akt- and forkhead box protein O1 (FoxO1)-dependent signaling pathway and nuclear factor-kB (NF-kB) activation. Knockdown of MT1-MMP by siRNA reversed apoptosis of ECs by inhibiting TNF-α-induced Akt-dependent regulation of FoxO1 in TNF-α-stimulated ECs. Immunoprecipitation demonstrated that TNF-α induced the changes in the associations between the cytoplasmic fraction of MT1-MMP and Akt in ECs. In conclusion, we show new evidence that MT1-MMP/Akt signaling axis is a key modifier for TNF-α-induced signaling pathways for modulation of procoagulant activity and apoptosis of ECs. PMID:25162582

Curcumin inhibits EMMPRIN and MMP-9 expression through AMPK-MAPK and PKC signaling in PMA induced macrophages.

PubMed

Cao, Jiatian; Han, Zhihua; Tian, Lei; Chen, Kan; Fan, Yuqi; Ye, Bozhi; Huang, Weijian; Wang, Changqian; Huang, Zhouqing

2014-09-21

In coronary arteries, plaque disruption, the major acute clinical manifestations of atherosclerosis, leads to a subsequent cardiac event, such as acute myocardial infarction (AMI) and unstable angina pectoris (UA). Numerous reports have shown that high expression of MMP-9 (matrix metalloproteinase-9), MMP-13 (matrix metalloproteinase-13) and EMMPRIN (extracellular matrix metalloproteinase induce) in monocyte/macrophage results in the plaque progression and destabilization. Curcumin exerts well-known anti-inflammatory and antioxidant effects and probably has a protective role in the atherosclerosis. The purpose of our study was to investigate the molecular mechanisms by which curcumin affects MMP-9, MMP13 and EMMPRIN in PMA (phorbol 12-myristate 13-acetate) induced macrophages. Human monocytic cells (THP-1 cells) were pretreated with curcumin or compound C for 1 h, and then induced by PMA for 48 h. Total RNA and proteins were collected for real-time PCR and Western blot analysis, respectively. In the present study, the exposure to curcumin resulted in attenuated JNK, p38, and ERK activation and decreased expression of MMP-9, MMP-13 and EMMPRIN in PMA induced macrophages. Moreover, we demonstrated that AMPK (AMP-activated protein kinase) and PKC (Protein Kinase C) was activated by PMA during monocyte/macrophage differentiation. Furthermore, curcumin reversed PMA stimulated PKC activation and suppressed the chronic activation of AMPK, which in turn reduced the expression of MMP-9, MMP-13 and EMMPRIN. Therefore, it is suggested that curcumin by inhibiting AMPK-MAPK (mitogen activated protein kinase) and PKC pathway may led to down-regulated EMMPRIN, MMP-9 and MMP-13 expression in PMA-induced THP-1 cells.

Selenium modulates MMP2 expression through the TGFβ1/Smad signalling pathway in human umbilical vein endothelial cells and rabbits following lipid disturbance.

PubMed

Xu, Chenggui; Lu, Guihua; Li, Qinglang; Zhang, Juhong; Huang, Zhibin; Gao, Xiuren

2017-07-01

A high-fat diet is a major risk factor for coronary heart diseases. Matrix metalloprotease (MMP) expression is changed in many cardiovascular diseases. Selenium, which is an important trace element in animals, has a close relationship with cardiovascular diseases. The TGFβ1/Smad signalling pathway is ubiquitous in diverse tissues and cells, and it is also associated with the occurrence and development of cardiovascular diseases. Therefore, in this study, we aimed to determine selenium's effect on lipid metabolism, atherosclerotic plaque formation, and MMP2 expression, as well as the underlying functional mechanism. In vivo tests: 24 male New Zealand white rabbits were randomly divided into 4 groups: regular diet, high-fat diet, high-fat diet+selenium and regular diet+selenium groups. The high-fat diet induced the lipid disturbances of rabbits at week 12. Selenium supplementation lowered total cholesterol (TC), low-density lipoprotein cholesterol (LDL-C) and triglyceride (TG) levels (p<0.01). Selenium supplementation also suppressed MMP2 over-expression in thoracic aortas. In vitro tests: Human umbilical vein endothelial cells (HUVECs) were treated with different concentrations of selenium or ox-LDL. Ox-LDL promoted MMP2 expression by increasing TGFβ1, pSmad2, pSmad3 and Smad3 expression (p<0.01). Selenium attenuated MMP2 over-expression by regulating the TGFβ1/Smad signalling pathway. Selenium suppressed high-fat diet-induced MMP2 over-expression in vivo by improving lipid metabolism. In vitro, selenium attenuated MMP2 over-expression through the TGFβ1/Smad signalling pathway. Copyright © 2017 Elsevier GmbH. All rights reserved.

Tomato Sl3-MMP, a member of the Matrix metalloproteinase family, is required for disease resistance against Botrytis cinerea and Pseudomonas syringae pv. tomato DC3000.

PubMed

Li, Dayong; Zhang, Huijuan; Song, Qiuming; Wang, Lu; Liu, Shixia; Hong, Yongbo; Huang, Lei; Song, Fengming

2015-06-14

Matrix metalloproteinases (MMPs) are a family of zinc-dependent endopeptidases. MMPs have been characterized in detail in mammals and shown to play key roles in many physiological and pathological processes. Although MMPs in some plant species have been identified, the function of MMPs in biotic stress responses remains elusive. A total of five MMP genes were identified in tomato genome. qRT-PCR analysis revealed that expression of Sl-MMP genes was induced with distinct patterns by infection of Botrytis cinerea and Pseudomonas syringae pv. tomato (Pst) DC3000 and by treatment with defense-related hormones such as salicylic acid, jasmonic acid and ethylene precursor 1-amino cyclopropane-1-carboxylic acid. Virus-induced gene silencing (VIGS)-based knockdown of individual Sl-MMPs and disease assays indicated that silencing of Sl3-MMP resulted in reduced resistance to B. cinerea and Pst DC3000, whereas silencing of other four Sl-MMPs did not affect the disease resistance against these two pathogens. The Sl3-MMP-silenced tomato plants responded with increased accumulation of reactive oxygen species and alerted expression of defense genes after infection of B. cinerea. Transient expression of Sl3-MMP in leaves of Nicotiana benthamiana led to an enhanced resistance to B. cinerea and upregulated expression of defense-related genes. Biochemical assays revealed that the recombinant mature Sl3-MMP protein had proteolytic activities in vitro with distinct preferences for specificity of cleavage sites. The Sl3-MMP protein was targeted onto the plasma membrane of plant cells when transiently expressed in onion epidermal cells. VIGS-based knockdown of Sl3-MMP expression in tomato and gain-of-function transient expression of Sl3-MMP in N. benthamiana demonstrate that Sl3-MMP functions as a positive regulator of defense response against B. cinerea and Pst DC3000.

Effects of different omega-3 sources, fish oil, krill oil, and green-lipped mussel against cytokine-mediated canine cartilage degradation.

PubMed

Buddhachat, Kittisak; Siengdee, Puntita; Chomdej, Siriwadee; Soontornvipart, Kumpanart; Nganvongpanit, Korakot

2017-05-01

Our purpose was to evaluate the protective effect of three marine omega-3 sources, fish oil (FO), krill oil (KO), and green-lipped mussel (GLM) against cartilage degradation. Canine cartilage explants were stimulated with either 10 ng/mL interleukin-1β (IL-1β) or IL-1β/oncostatin M (10 ng/mL each) and then treated with various concentrations of docosahexaenoic acid (DHA), eicosapentaenoic acid (EPA; 3 and 30 μg/mL), FO, KO, or GLM (250, 500, and 1000 μg/mL) for 28 days. Gene expression was then investigated in primary canine chondrocytes. Our results showed that DHA and EPA as well as omega-3 sources could suppress matrix degradation in cytokine-induced cartilage explants by significantly reducing the increase of sulfated glycosaminoglycans (s-GAGs) and preserving uronic acid and hydroxyproline content (except GLM). These agents were not able to reduce IL-1β-induced IL1B and TNFA expression but were able to down-regulate the expression of the catabolic genes MMP1, MMP3, and MMP13 and up-regulate the anabolic genes AGG and COL2A1; FO and KO were especially effective. Our findings indicated that FO and KO were superior to GLM for their protective effect against proteoglycan and collagen degradation. Hence, FO and KO could serve as promising sources of chondroprotective agents.

The protective effects of fucosterol against skin damage in UVB-irradiated human dermal fibroblasts.

PubMed

Hwang, Eunson; Park, Sang-Yong; Sun, Zheng-wang; Shin, Heon-Sub; Lee, Don-Gil; Yi, Tae Hoo

2014-06-01

Exposure to ultraviolet (UV) light causes matrix metalloproteinase (MMP) overexpression and extracellular matrix depletion, leading to skin photoaging. The activation of MMP is related to increased interlukin-6 (IL-6) and type I procollagen production, which is regulated by transforming growth factor-β1 (TGF-β1). Activator protein-1 (AP-1) activation induces MMP-1 production and reduces type I procollagen secretion. Fucosterol, which is extracted and purified from the brown algae Hizikia fusiformis, is a phytosterol. We assessed the effects of fucosterol on photodamage and investigated its molecular mechanism of action in UVB-irradiated normal human dermal fibroblasts by using enzyme-linked immunosorbent assay, Western blot analysis, and reverse transcription-polymerase chain reaction. Our results showed that fucosterol significantly decreased the UVB-induced expression of MMP-1, IL-6, p-c-Jun, and p-c-Fos. Additionally, fucosterol markedly increased the UVB-induced production of type I procollagen and TGF-β1. Our results indicate that fucosterol regulates MMP-1 and type I procollagen expression by modulating AP-1 and TGF-β1 signaling and that MMP-1 activation is correlated with IL-6. These data suggest that fucosterol is a promising botanical agent to protect against skin photodamage.

6-Gingerol inhibits hair cycle via induction of MMP2 and MMP9 expression.

PubMed

Hou, Chun; Miao, Yong; Ji, Hang; Wang, Susheng; Liang, Gang; Zhang, Zhihua; Hong, Weijin

2017-01-01

6-Gingerol is the major active constituent of ginger. In the current study, we aimed to investigate the mechanisms underlying the effects of 6-Gingerol on hair growth. Mice were randomly divided into five groups; after hair depilation (day 0), mice were treated with saline, or different concentrations of 6-Gingerol for 11 days. The histomorphological characteristics of the growing hair follicles were examined after hematoxylin and eosin staining. The results indicated that 6-Gingerol significantly suppressed hair growth compared with that in the control group. And choose the concentration of 6-Gingerol at 1 mg/mL to treated with mice. Moreover, 6-Gingerol (1 mg/mL) significantly reduced hair re-growth ratio, hair follicle number, and hair follicle length, which were associated with increased expression of MMP2 and MMP9. Furthermore, the growth factors, such as EGF, KGF, VEGF, IGF-1 and TGF-β participate in the hair follicle cycle regulation and regulate hair growth. We then measured the concentrations of them using ELISA assays, and the results showed that 6-Gingerol decreased EGF, KGF, VEGF, and IGF-1 concentrations, and increased TGF-β concentration. Thus, this study showed that 6-Gingerol might act as a hair growth suppressive drug via induction of MMP2 and MMP9 expression, which could interfere with the hair cycle.

Cleaved high molecular weight kininogen, a novel factor in the regulation of matrix metalloproteinases in vascular smooth muscle cells.

PubMed

Vosgerau, Uwe; Lauer, Diljara; Unger, Thomas; Kaschina, Elena

2010-01-15

We previously reported that Brown Norway Katholiek rats, which feature a deficiency of plasma kininogens, develop severe abdominal aortic aneurysm. Increased activity of matrix metalloproteinases (MMPs) in the aortic wall, leading to degradation of extracellular matrix components, is considered to play a crucial role in aneurysm formation. Using an in vitro model of vascular smooth muscle cells (VSMCs), cultured from the rat aorta, we investigated whether the cleaved form of high molecular weight kininogen, designated HKa, affects the expression of MMP-9 and MMP-2 and their tissue inhibitors (TIMPs). Treatment of VSMCs with HKa reduced in a concentration-dependent manner IL-1alpha-induced release of MMP-9 and MMP-2, associated with decreased MMP enzymatic activity levels in conditioned media, as demonstrated by gelatin zymography and fluorescein-labeled gelatin substrate assay, respectively. Real-time PCR revealed that HKa reduced corresponding MMP-9 mRNA levels. Further investigations showed that this effect did not result from a modified rate of MMP-9 mRNA degradation. TIMP-1 mRNA levels, already increased as a result of cytokine-stimulation, were significantly enhanced by HKa. Furthermore, we found elevated basal mRNA expression levels of MMP-2 and TIMP-2 in VSMCs derived from kininogen-deficient Brown Norway Katholiek rats. These results demonstrate for the first time that HKa affects the regulation of MMPs in VSMCs.

Naringenin targets ERK2 and suppresses UVB-induced photoaging.

PubMed

Jung, Sung Keun; Ha, Su Jeong; Jung, Chang Hwa; Kim, Yun Tai; Lee, Hoo-Keun; Kim, Myoung Ok; Lee, Mee-Hyun; Mottamal, Madhusoodanan; Bode, Ann M; Lee, Ki Won; Dong, Zigang

2016-05-01

A number of natural phytochemicals have anti-photoaging properties that appear to be mediated through the inhibition of matrix metalloproteinase-1 (MMP-1) expression, but their direct target molecule(s) and mechanism(s) remain unclear. We investigated the effect of naringenin, a major flavonoid found in citrus, on UVB-induced MMP-1 expression and identified its direct target. The HaCaT human skin keratinocyte cell line and 3-dimensional (3-D) human skin equivalent cultures were treated or not treated with naringenin for 1 hr before exposure to UVB. The mechanism and target(s) of naringenin were analysed by kinase assay and multiplex molecular assays. Dorsal skins of hairless mice were exposed to UVB 3 times per week, with a dose of irradiation that was increased weekly by 1 minimal erythema dose (MED; 45 mJ/cm(2)) to 4 MED over 15 weeks. Wrinkle formation, water loss and water content were then assessed. Naringenin suppressed UVB-induced MMP-1 expression and AP-1 activity, and strongly suppressed UVB-induced phosphorylation of Fos-related antigen (FRA)-1 at Ser265. Importantly, UVB irradiation-induced FRA1 protein stability was reduced by treatment with naringenin, as well as with a mitogen-activated protein kinase (MEK) inhibitor. Naringenin significantly suppressed UVB-induced extracellular signal-regulated kinase 2 (ERK2) activity and subsequently attenuated UVB-induced phosphorylation of p90(RSK) by competitively binding with ATP. Constitutively active MEK (CA-MEK) increased FRA1 phosphorylation and expression and also induced MMP-1 expression, whereas dominant-negative ERK2 (DN-ERK2) had opposite effects. U0126, a MEK inhibitor, also decreased FRA1 phosphorylation and expression as well as MMP-1 expression. The photoaging data obtained from mice clearly demonstrated that naringenin significantly inhibited UVB-induced wrinkle formation, trans-epidermal water loss and MMP-13 expression. Naringenin exerts potent anti-photoaging effects by suppressing ERK2 activity and decreasing FRA1 stability, followed by down-regulation of AP-1 transactivation and MMP-1 expression. © 2016 The Authors. Journal of Cellular and Molecular Medicine published by John Wiley & Sons Ltd and Foundation for Cellular and Molecular Medicine.

The Complement C3a-C3aR Axis Promotes Development of Thoracic Aortic Dissection via Regulation of MMP2 Expression.

PubMed

Ren, Weihong; Liu, Yan; Wang, Xuerui; Piao, Chunmei; Ma, Youcai; Qiu, Shulan; Jia, Lixin; Chen, Boya; Wang, Yuan; Jiang, Wenjian; Zheng, Shuai; Liu, Chang; Dai, Nan; Lan, Feng; Zhang, Hongjia; Song, Wen-Chao; Du, Jie

2018-03-01

Thoracic aortic dissection (TAD), once ruptured, is devastating to patients, and no effective pharmaceutical therapy is available. Anaphylatoxins released by complement activation are involved in a variety of diseases. However, the role of the complement system in TAD is unknown. We found that plasma levels of C3a, C4a, and C5a were significantly increased in patients with TAD. Elevated circulating C3a levels were also detected in the developmental process of mouse TAD, which was induced by β-aminopropionitrile monofumarate (BAPN) treatment, with enhanced expression of C1q and properdin in mouse dissected aortas. These findings indicated activation of classical and alternative complement pathways. Further, expression of C3aR was obviously increased in smooth muscle cells of human and mouse dissected aortas, and knockout of C3aR notably inhibited BAPN-induced formation and rupture of TAD in mice. C3aR antagonist administered pre- and post-BAPN treatment attenuated the development of TAD. We found that C3aR knockout decreased matrix metalloproteinase 2 (MMP2) expression in BAPN-treated mice. Additionally, recombinant C3a stimulation enhanced MMP2 expression and activation in smooth muscle cells that were subjected to mechanical stretch. Finally, we generated MMP2-knockdown mice by in vivo MMP2 short hairpin RNA delivery using recombinant adeno-associated virus and found that MMP2 deficiency significantly reduced the formation of TAD. Therefore, our study suggests that the C3a - C3aR axis contributes to the development of TAD via regulation of MMP2 expression. Targeting the C3a-C3aR axis may represent a strategy for inhibiting the formation of TAD. Copyright © 2018 by The American Association of Immunologists, Inc.

Naringenin regulates production of matrix metalloproteinases in the knee-joint and primary cultured articular chondrocytes and alleviates pain in rat osteoarthritis model.

PubMed

Wang, C C; Guo, L; Tian, F D; An, N; Luo, L; Hao, R H; Wang, B; Zhou, Z H

2017-03-23

Inflammation of cartilage is a primary symptom for knee-joint osteoarthritis. Matrix metalloproteinases (MMPs) are known to play an important role in the articular cartilage destruction related to osteoarthritis. Naringenin is a plant-derived flavonoid known for its anti-inflammatory properties. We studied the effect of naringenin on the transcriptional expression, secretion and enzymatic activity of MMP-3 in vivo in the murine monosodium iodoacetate (MIA) osteoarthritis model. The assessment of pain behavior was also performed in the MIA rats. The destruction of knee-joint tissues was analyzed microscopically. Moreover, the effect of naringenin was also studied in vitro in IL-1β activated articular chondrocytes. The transcriptional expression of MMP-3, MMP-1, MMP-13, thrombospondin motifs (ADAMTS-4) and ADAMTS-5 was also studied in primary cultured chondrocytes of rats. Naringenin caused significant reduction in pain behavior and showed marked improvement in the tissue morphology of MIA rats. Moreover, a significant inhibition of MMP-3 expression in MIA rats was observed upon treatment with naringenin. In the in vitro tests, naringenin caused a significant reduction in the transcriptional expression, secretion and enzymatic activity of the studied degradative enzymes. The NF-κB pathway was also found to be inhibited upon treatment with naringenin in vitro. Overall, the study suggests that naringenin alleviated pain and regulated the production of matrix-metalloproteinases via regulation of NF-κB pathway. Thus, naringenin could be a potent therapeutic option for the treatment of osteoarthritis.

Hydrostatic Pressure Influences HIF-2 Alpha Expression in Chondrocytes

PubMed Central

Inoue, Hiroaki; Arai, Yuji; Kishida, Tsunao; Terauchi, Ryu; Honjo, Kuniaki; Nakagawa, Shuji; Tsuchida, Shinji; Matsuki, Tomohiro; Ueshima, Keiichirou; Fujiwara, Hiroyoshi; Mazda, Osam; Kubo, Toshikazu

2015-01-01

Hypoxia-inducible factor (HIF)-2α is considered to play a major role in the progression of osteoarthritis. Recently, it was reported that pressure amplitude influences HIF-2α expression in murine endothelial cells. We examined whether hydrostatic pressure is involved in expression of HIF-2α in articular chondrocytes. Chondrocytes were cultured and stimulated by inflammation or hydrostatic pressure of 0, 5, 10, or 50 MPa. After stimulation, heat shock protein (HSP) 70, HIF-2α, nuclear factor kappa B (NF-κB), matrix metalloproteinase (MMP)-13, MMP-3, and vascular endothelial growth factor (VEGF) gene expression were evaluated. The levels of all gene expression were increased by inflammatory stress. When chondrocytes were exposed to a hydrostatic pressure of 5 MPa, HIF-2α, MMP-13, and MMP-3 gene expression increased significantly although those of HSP70 and NF-κB were not significantly different from the control group. In contrast, HIF-2α gene expression did not increase under a hydrostatic pressure of 50 MPa although HSP70 and NF-κB expression increased significantly compared to control. We considered that hydrostatic pressure of 5 MPa could regulate HIF-2α independent of NF-κB, because the level of HIF-2α gene expression increased significantly without upregulation of NF-κB expression at 5 MPa. Hydrostatic pressure may influence cartilage degeneration, inducing MMP-13 and MMP-3 expression through HIF-2α. PMID:25569085

Deletion of the transforming growth factor β receptor type II gene in articular chondrocytes leads to a progressive osteoarthritis-like phenotype in mice.

PubMed

Shen, Jie; Li, Jia; Wang, Baoli; Jin, Hongting; Wang, Meina; Zhang, Yejia; Yang, Yunzhi; Im, Hee-Jeong; O'Keefe, Regis; Chen, Di

2013-12-01

While transforming growth factor β (TGFβ) signaling plays a critical role in chondrocyte metabolism, the TGFβ signaling pathways and target genes involved in cartilage homeostasis and the development of osteoarthritis (OA) remain unclear. Using an in vitro cell culture method and an in vivo mouse genetic approach, we undertook this study to investigate TGFβ signaling in chondrocytes and to determine whether Mmp13 and Adamts5 are critical downstream target genes of TGFβ signaling. TGFβ receptor type II (TGFβRII)-conditional knockout (KO) (TGFβRII(Col2ER)) mice were generated by breeding TGFβRII(flox/flox) mice with Col2-CreER-transgenic mice. Histologic, histomorphometric, and gene expression analyses were performed. In vitro TGFβ signaling studies were performed using chondrogenic rat chondrosarcoma cells. To determine whether Mmp13 and Adamts5 are critical downstream target genes of TGFβ signaling, TGFβRII/matrix metalloproteinase 13 (MMP-13)- and TGFβRII/ADAMTS-5-double-KO mice were generated and analyzed. Inhibition of TGFβ signaling (deletion of the Tgfbr2 gene in chondrocytes) resulted in up-regulation of Runx2, Mmp13, and Adamts5 expression in articular cartilage tissue and progressive OA development in TGFβRII(Col2ER) mice. Deletion of the Mmp13 or Adamts5 gene significantly ameliorated the OA-like phenotype induced by the loss of TGFβ signaling. Treatment of TGFβRII(Col2ER) mice with an MMP-13 inhibitor also slowed OA progression. Mmp13 and Adamts5 are critical downstream target genes involved in the TGFβ signaling pathway during the development of OA. Copyright © 2013 by the American College of Rheumatology.

Taspine derivative 12k suppressed A549 cell migration through the Wnt/β-catenin and EphrinB2 signaling pathway.

PubMed

Dai, Bingling; Ma, Yujiao; Yang, Tianfeng; Wang, Wenjie; Zhang, Yanmin

2017-03-01

12k, a taspine derivative, has been demonstrated to have the potent anti-tumor activity in lung cancer and colorectal cancer. The study aims to further explore the underlying mechanisms of 12k on A549 cell migration in vitro. Our data demonstrated that 12k negatively regulated Wnt signaling pathway by suppressing the phosphorylation of LRP5/6, and inhibiting the expression and nuclear translocation of β-catenin. 12k was shown to downregulate MMP3 and MMP7 expression which regulated by β-catenin interacts with TCF/LEF in the nucleus, and effectively impaired the related migration protein expression of MMP2 and MMP9 in A549 cells. In addition, 12k repressed the EphrinB2 and its PDZ protein, impairing the VEGFR2 and VEGFR3 expression in A549 cells, as well as inhibited the downstream of VEGFR2 included PI3K/AKT/mTOR and ERK/MAPK signaling pathways. Taken together, our findings revealed that 12k suppressed migration of A549 cells through the Wnt/β-catenin signaling pathway and EphrinB2 related signaling pathway. Copyright © 2016 Elsevier Masson SAS. All rights reserved.

Resveratrol protects primary cortical neuron cultures from transient oxygen-glucose deprivation by inhibiting MMP-9.

PubMed

Gao, Dakuan; Huang, Tao; Jiang, Xiaofan; Hu, Shijie; Zhang, Lei; Fei, Zhou

2014-06-01

It was recently shown that resveratrol exerts neuroprotective effects against cerebral ischemia in mice. The aim of the present study was to further confirm these effects in in vitro primary cortical neuron cultures with transient oxygen-glucose deprivation (OGD), and to investigate whether these effects are due to the inhibition of matrix metalloproteinase-9 (MMP-9) and of cell apoptosis. Neuronal primary cultures of cerebral cortex were prepared from BALB/c mice embryos (13-15 days). Cells from 14- to 16-day cultures were subjected to OGD for 3 h, followed by 21 h of reoxygenation to simulate transient ischemia. Different doses of resveratrol were added into the culture medium during the simulation of transient ischemia. The effect of the extracellular signal-regulated kinase (ERK) inhibitor U0126 was studied by adding U0126 (5 µg/µl, 4 µl) into the culture medium during transient ischemia; as a control, we used treatment of cells with 50 µM of resveratrol. Cell viability was investigated using the 3-[4,5-dimethylthiazol-2-yl]-2,5 diphenyl tetrazolium bromide (MTT) reduction assay. Cell apoptosis was assessed by flow cytometry. The effects of resveratrol on the expression of MMP-9 were analyzed by western blotting and reverse transcription-polymerase chain reaction (RT-PCR), while the levels of ERK, phosphorylated (p)-ERK, cleaved caspase-3, Bax and Bcl-2 were measured by western blotting. The results of the MTT assay showed that cell viability is significantly reduced by transient OGD. OGD induced cell apoptosis, the expression of Bax and the activation of caspase-3 and ERK, inhibited the expression of Bcl-2 and increased the expression of MMP-9, while these effects were reversed by treatment with resveratrol. The therapeutic efficacy of resveratrol was shown to be dose-dependent, with the most suitable dose range determined at 50-100 µM. Treatment with U0126 inhibited MMP-9 and Bax expression and caspase-3 activation, while it further promoted the expression of the anti-apoptotic molecule Bcl-2, suggesting that resveratrol inhibits MMP-9 expression and cell apoptosis by attenuating the activation of ERK1/2. In conclusion, OGD can induce apoptosis through canonical apoptotic signals and by regulating the expression of MMP-9; the anti-apoptotic activity of resveratrol and its inhibitory effect on MMP-9 expression contribute in the reduced activation of ERK.

MMP16 promotes tumor metastasis and indicates poor prognosis in hepatocellular carcinoma

PubMed Central

Shen, Zhenghai; Wang, Xing; Yu, Xiaotian; Zhang, Yun; Qin, Lei

2017-01-01

Matrix metalloproteinase (MMPs) participates in multiple biological behaviors and plays an important role in regulating tumor invasion. However, the functions of MMP16 in hepatocellular carcinoma (HCC) remains unknown. The prognostic value of MMP16 was studied in TCGA database and validation cohort. MMP16-silencing HCC cells (HepG2 and HCCLM3) were used for evaluating cell proliferation and invasion by CCK-8 and Transwell assays. Our results showed that the MMP16 was a predictor for overall survival in patients with HCC (HR: 1.169, 95% CI: 1.034–1.321, P = 0.013) in TCGA database. In validation cohort, MMP16 expression was an independent predictor for survival in both univariate and multivariate analysis (P < 0.05). Furthermore, knockdown MMP16 weakened the cell invasive potential by inhibiting epithelial-mesenchymal transition (EMT) process. Therefore, our findings showed that MMP16 was a prognostic factor in HCC, ectopic MMP16 expression promoted invasion of HCC cells by inducing EMT process, suggesting a tumor oncogenic function in HCC and provides the potential therapeutic target for the treatment of HCC. PMID:29069779

Generation of Soluble Receptor Activator of NF-KappaB Ligand Is Critical for Osteolytic Bone Metastasis

DTIC Science & Technology

2010-10-01

including breast (15), head and neck squa- mous carcinoma (18), melanoma, and chondrosarcoma (42). A previous report suggests involvement of MMP13 in the...2003;88: 1318–26. 42. Uria JA, Balbin M, Lopez JM, et al. Collagenase-3 (MMP-13) expres- sion in chondrosarcoma cells and its regulation by basic

Differential free fatty acid receptor-1 (FFAR1/GPR40) signalling is associated with gene expression or gelatinase granule release in bovine neutrophils.

PubMed

Mena, Sandra J; Manosalva, Carolina; Carretta, Maria D; Teuber, Stefanie; Olmo, Iván; Burgos, Rafael A; Hidalgo, Maria A

2016-08-01

Fatty acids have been recognized as regulators of immune function in addition to their known metabolic role. Long-chain fatty acids bind free fatty acid receptor (FFAR)-1/GPR40, which is expressed on bovine neutrophils, and increase responses such as granule release and gene expression. In this study, we investigated the molecular mechanisms governing the up-regulation of cyclooxygenase-2 (COX-2) and IL-8, as well as matrix metalloproteinase (MMP)-9 granule release in FFAR1/GPR40 agonist-stimulated neutrophils. Our results showed that natural (oleic and linoleic acid) and synthetic (GW9508) FFAR1/GPR40 agonists increased ERK1/2, p38 MAPK and Akt phosphorylation, and that the FFAR1/GPR40 antagonist GW1100 reduced these responses. We evaluated the levels of IκBα, a component of the classical activation pathway of the transcription factor NF-κB, and we observed IκBα reduction after stimulation with FFAR1/GPR40 agonists, an effect that was inhibited by GW1100 or the inhibitors UO126, SB203580 or LY294002. FFAR1/GPR40 agonists increased COX-2 and IL-8 expression, which was inhibited by GW1100 and an NF-κB inhibitor. Finally, the FFAR1/GPR40 agonist-induced MMP-9 granule release was reduced by GW1100 and UO126. In conclusion, FFAR1/GPR40 agonists differentially stimulate neutrophil functions; COX-2 and IL-8 are expressed after FFAR1/GPR40 activation via NF-κB, IκBα reduction is FFAR1/GPR40- and PI3K/MAPK-dependent, and MMP-9 granule release is FFAR1/GPR40- and ERK1/2-dependent. © The Author(s) 2016.

Hsa-miR-326 targets CCND1 and inhibits non-small cell lung cancer development

PubMed Central

Li, Shujun; Yang, Cuili; Xi, Yongyong; Wang, Liang; Zhang, Feng; Fu, Yunfeng; Li, Dejia

2016-01-01

Hsa-miRNA-326 (miR-326) has recently been discovered having anticancer efficacy in different organs. However, the role of miR-326 on non-small cell lung cancer (NSCLC) is still ambiguous. In this study, we investigated the role of miR-326 on the development of NSCLC. The results indicated that miR-326 was significantly down-regulated in primary tumor tissues and very low levels were found in NSCLC cell lines. Ectopic expression of miR-326 in NSCLC cell lines significantly suppressed cell growth as evidenced by cell viability assay, colony formation assay and BrdU staining, through inhibition of cyclin D1, cyclin D2, CDK4 and up-regulation of p57(Kip2) and p21(Waf1/Cip1). In addition, miR-326 induced apoptosis, as indicated by concomitantly with up-regulation of key apoptosis protein cleaved caspase-3, and down-regulation of anti-apoptosis protein Bcl2. Moreover, miR-326 inhibited cellular migration and invasiveness through inhibition of matrix metalloproteinases (MMP)-7 and MMP-9. Further, oncogene CCND1 was revealed to be a putative target of miR-326, which was inversely correlated with miR-326 expression in NSCLC. Taken together, our results demonstrated that miR-326 played a pivotal role on NSCLC through inhibiting cell proliferation, migration, invasion, and promoting apoptosis by targeting oncogenic CCND1. PMID:26840018

Local and systemic gene expression responses of Atlantic salmon (Salmo salar L.) to infection with the salmon louse (Lepeophtheirus salmonis)

PubMed Central

Skugor, Stanko; Glover, Kevin Alan; Nilsen, Frank; Krasnov, Aleksei

2008-01-01

Background The salmon louse (SL) is an ectoparasitic caligid crustacean infecting salmonid fishes in the marine environment. SL represents one of the major challenges for farming of salmonids, and veterinary intervention is necessary to combat infection. This study addressed gene expression responses of Atlantic salmon infected with SL, which may account for its high susceptibility. Results The effects of SL infection on gene expression in Atlantic salmon were studied throughout the infection period from copepodids at 3 days post infection (dpi) to adult lice (33 dpi). Gene expression was analyzed at three developmental stages in damaged and intact skin, spleen, head kidney and liver, using real-time qPCR and a salmonid cDNA microarray (SFA2). Rapid detection of parasites was indicated by the up-regulation of immunoglobulins in the spleen and head kidney and IL-1 receptor type 1, CD4, beta-2-microglobulin, IL-12β, CD8α and arginase 1 in the intact skin of infected fish. Most immune responses decreased at 22 dpi, however, a second activation was observed at 33 dpi. The observed pattern of gene expression in damaged skin suggested the development of inflammation with signs of Th2-like responses. Involvement of T cells in responses to SL was witnessed with up-regulation of CD4, CD8α and programmed death ligand 1. Signs of hyporesponsive immune cells were seen. Cellular stress was prevalent in damaged skin as seen by highly significant up-regulation of heat shock proteins, other chaperones and mitochondrial proteins. Induction of the major components of extracellular matrix, TGF-β and IL-10 was observed only at the adult stage of SL. Taken together with up-regulation of matrix metalloproteinases (MMP), this classifies the wounds afflicted by SL as chronic. Overall, the gene expression changes suggest a combination of chronic stress, impaired healing and immunomodulation. Steady increase of MMP expression in all tissues except liver was a remarkable feature of SL infected fish. Conclusion SL infection in Atlantic salmon is associated with a rapid induction of mixed inflammatory responses, followed by a period of hyporesponsiveness and delayed healing of injuries. Persistent infection may lead to compromised host immunity and tissue self-destruction. PMID:18945374

An intracellular matrix metalloproteinase-2 isoform induces tubular regulated necrosis: implications for acute kidney injury.

PubMed

Ceron, Carla S; Baligand, Celine; Joshi, Sunil; Wanga, Shaynah; Cowley, Patrick M; Walker, Joy P; Song, Sang Heon; Mahimkar, Rajeev; Baker, Anthony J; Raffai, Robert L; Wang, Zhen J; Lovett, David H

2017-06-01

Acute kidney injury (AKI) causes severe morbidity, mortality, and chronic kidney disease (CKD). Mortality is particularly marked in the elderly and with preexisting CKD. Oxidative stress is a common theme in models of AKI induced by ischemia-reperfusion (I-R) injury. We recently characterized an intracellular isoform of matrix metalloproteinase-2 (MMP-2) induced by oxidative stress-mediated activation of an alternate promoter in the first intron of the MMP-2 gene. This generates an NH 2 -terminal truncated MMP-2 (NTT-MMP-2) isoform that is intracellular and associated with mitochondria. The NTT-MMP-2 isoform is expressed in kidneys of 14-mo-old mice and in a mouse model of coronary atherosclerosis and heart failure with CKD. We recently determined that NTT-MMP-2 is induced in human renal transplants with delayed graft function and correlated with tubular cell necrosis. To determine mechanism(s) of action, we generated proximal tubule cell-specific NTT-MMP-2 transgenic mice. Although morphologically normal at the light microscopic level at 4 mo, ultrastructural studies revealed foci of tubular epithelial cell necrosis, the mitochondrial permeability transition, and mitophagy. To determine whether NTT-MMP-2 expression enhances sensitivity to I-R injury, we performed unilateral I-R to induce mild tubular injury in wild-type mice. In contrast, expression of the NTT-MMP-2 isoform resulted in a dramatic increase in tubular cell necrosis, inflammation, and fibrosis. NTT-MMP-2 mice had enhanced expression of innate immunity genes and release of danger-associated molecular pattern molecules. We conclude that NTT-MMP-2 "primes" the kidney to enhanced susceptibility to I-R injury via induction of mitochondrial dysfunction. NTT-MMP-2 may be a novel AKI treatment target.

Rosmarinic acid counteracts activation of hepatic stellate cells via inhibiting the ROS-dependent MMP-2 activity: Involvement of Nrf2 antioxidant system

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lu, Changfang; Zou, Yu; Liu, Yuzhang

Recently, oxidative stress is involved in hepatofibrogenesis. Matrix metalloproteinase-2 (MMP-2) is required for activation of hepatic stellate cells (HSCs) in response to reactive oxygen species (ROS). This study was designed to explore the hypothesis that the inhibitory effect of rosmarinic acid (RA) on HSCs activation might mainly result from its antioxidant capability by increasing the synthesis of glutathione (GSH) involved in nuclear factor kappa B (NF-κB)-dependent inhibition of MMP-2 activity. Here, we demonstrate that RA reverses activated HSCs to quiescent cells. Concomitantly, RA inhibits MMP-2 activity. RNA interference-imposed knockdown of NF-κB abolished down-regulation of MMP-2 by RA. RA-mediated inactivation ofmore » NF-κB could be blocked by the diphenyleneiodonium chloride (DPI; a ROS inhibitor). Conversely, transfection of dominant-negative (DN) mutant of extracellular signal-regulated kinases 2 (ERK2), c-Jun N-terminal kinase 1 (JNK1), or p38α kinase had no such effect. Simultaneously, RA suppresses ROS generation and lipid peroxidation (LPO) whereas increases cellular GSH in HSC-T6 cells. Furthermore, RA significantly increased antioxidant response element (ARE)-mediated luciferase activity, nuclear translocation of nuclear factor erythroid 2-related factor 2 (Nrf2) and catalytic subunits from glutamate cysteine ligase (GCLc) expression, but not modulatory subunits from GCL (GCLm). RA-mediated up-regulation of GClc is inhibited by the shRNA-induced Nrf2 knockdown. The knocking down of Nrf2 or buthionine sulfoximine (a GCL inhibitor) abolished RA-mediated inhibition of ROS. Collectively, these results provide novel insights into the mechanisms of RA as an antifibrogenic candidate in the prevention and treatment of liver fibrosis. - Highlights: • RA reverses activated HSCs to quiescent cells. • RA suppresses MMP-2 activity through a NF-κB-dependent pathway. • Inhibition of oxidative stress by RA is dependent on nuclear translocation of Nrf2. • RA-mediated down-regulation of MMP-2 was ROS-dependent.« less

Artesunate modulates expression of matrix metalloproteinases and their inhibitors as well as collagen-IV to attenuate pulmonary fibrosis in rats.

PubMed

Wang, Y; Huang, G; Mo, B; Wang, C

2016-06-03

The aim of this study was to determine the effect of artesunate on extracellular matrix (ECM) accumulation and the expression of collagen-IV, matrix metalloproteinase (MMP), and tissue inhibitor of matrix metalloproteinase (TIMP) to understand the pharmacological role of artesunate in pulmonary fibrosis. Eighty Sprague-Dawley rats were randomly assigned to four groups that were administered saline alone, bleomycin (BLM) alone, BLM + artesunate, or artesunate alone for 28 days. Lung tissues from 10 rats in each group were used to obtain lung fibroblast (LF) primary cells, and the rest were used to analyze protein expression. The mRNA expression of collagen-IV, MMP-2, MMP-9, TIMP-1, and TIMP-2 in lung fibroblasts was detected by real-time quantitative reverse transcriptase polymerase chain reaction. The protein levels of collagen-IV, MMP-2, MMP-9, TIMP-1, and TIMP-2 protein in lung tissues were analyzed by western blotting. Artesunate treatment alleviated alveolitis and pulmonary fibrosis induced by bleomycin in rats, as indicated by a decreased lung coefficient and improvement of lung tissue morphology. Artesunate treatment also led to decreased collagen-IV protein levels, which might be a result of its downregulated expression and increased MMP-2 and MMP-9 protein and mRNA levels. Increased TIMP-1 and TIMP- 2 protein and mRNA levels were detected after artesunate treatment in lung tissues and primary lung fibroblast cells and may contribute to enhanced activity of MMP-2 and -9. These findings suggested that artesunate attenuates alveolitis and pulmonary fibrosis by regulating expression of collagen-IV, TIMP-1 and 2, as well as MMP-2 and -9, to reduce ECM accumulation.

Protease Expression Levels in Prostate Cancer Tissue Can Explain Prostate Cancer-Associated Seminal Biomarkers-An Explorative Concept Study.

PubMed

Neuhaus, Jochen; Schiffer, Eric; Mannello, Ferdinando; Horn, Lars-Christian; Ganzer, Roman; Stolzenburg, Jens-Uwe

2017-05-04

Previously, we described prostate cancer (PCa) detection (83% sensitivity; 67% specificity) in seminal plasma by CE-MS/MS. Moreover, advanced disease was distinguished from organ-confined tumors with 80% sensitivity and 82% specificity. The discovered biomarkers were naturally occurring fragments of larger seminal proteins, predominantly semenogelin 1 and 2, representing endpoints of the ejaculate liquefaction. Here we identified proteases putatively involved in PCa specific protein cleavage, and examined gene expression and tissue protein levels, jointly with cell localization in normal prostate (nP), benign prostate hyperplasia (BPH), seminal vesicles and PCa using qPCR, Western blotting and confocal laser scanning microscopy. We found differential gene expression of chymase (CMA1), matrix metalloproteinases (MMP3, MMP7), and upregulation of MMP14 and tissue inhibitors (TIMP1 and TIMP2) in BPH. In contrast tissue protein levels of MMP14 were downregulated in PCa. MMP3/TIMP1 and MMP7/TIMP1 ratios were decreased in BPH. In seminal vesicles, we found low-level expression of most proteases and, interestingly, we also detected TIMP1 and low levels of TIMP2. We conclude that MMP3 and MMP7 activity is different in PCa compared to BPH due to fine regulation by their inhibitor TIMP1. Our findings support the concept of seminal plasma biomarkers as non-invasive tool for PCa detection and risk stratification.

PEP-1-SIRT2-induced matrix metalloproteinase-1 and -13 modulates type II collagen expression via ERK signaling in rabbit articular chondrocytes.

PubMed

Eo, Seong-Hui; Choi, Soo Young; Kim, Song Ja

2016-11-01

Matrix metalloproteinases (MMPs) are critical for the degradation of the extracellular matrix (ECM), which includes cartilage-specific collagen types I, II and XI. We previously found that PEP-1-sirtuin (SIRT)2 could induce dedifferentiation of articular chondrocytes; however, the underlying mechanisms remains unclear. We addressed this in the present study by examining the association between PEP-1-SIRT2 and the expression of MMP-1 and MMP-13 and type II collagen in rabbit articular chondrocytes. We found that PEP-1-SIRT2 increased MMP-1 and -13 expression in a dose- and time-dependent manner, as determined by western blotting. A similar trend in MMP-1 and -13 levels was observed in cultures during expansion to four passages. Pharmacological inhibition of MMP-1 and -13 blocked the PEP-1-SIRT2-induced decrease in type II collagen level. Phosphorylation of extracellular regulated kinase (ERK) was increased by PEP-1-SIRT2; however, treatment with the mitogen-activated protein kinase inhibitor PD98059 suppressed PEP-1-SIRT2-induced MMP-1 and -13 expression and dedifferentiation while restoring type II collagen expression in passage 2 cells. These results suggest that PEP-1-SIRT2 promotes MMP-induced dedifferentiation via ERK signaling in articular chondrocytes. Copyright © 2016 Elsevier Inc. All rights reserved.

Hepatocyte growth factor/scatter factor enhances the invasion of mesothelioma cell lines and the expression of matrix metalloproteinases

PubMed Central

Harvey, P; Clark, I M; Jaurand, M-C; Warn, R M; Edwards, D R

2000-01-01

Hepatocyte growth factor/scatter factor (HGF/SF) is a multifunctional factor involved both in development and tissue repair, as well as pathological processes such as cancer and metastasis. It has been identified in vivo in many types of tumours together with its tyrosine kinase receptor, Met. We show here that exogenous HGF/SF acts as a strong chemoattractant for human mesothelioma cell lines. The factor also enhanced cell adhesion to and invasion into Matrigel. The mesothelioma cell lines synthesized a panel of matrix metalloproteinases critical for tumour progression such as MMP-1, 2, 3, 9 and membrane-bound MT1-MMP. HGF/SF stimulated the expression of MMP-1, 9 and MT1-MMP and had a slight effect on expression of the MMP inhibitor TIMP-1 but not TIMP-2. However, there was no simple correlation between the levels of MMPs and TIMPs of the cell lines and their different invasion properties or between HGF/SF stimulatory effects on MMP expression and invasion. In addition, effects of protease inhibitors on invasion suggested that serine proteases were also expressed in human mesothelioma cell lines and were involved in HGF/SF-induced invasion. The results show a predominant role for HGF/SF in mesothelioma cell invasion, stimulating simultaneously adhesion, motility, invasion and regulation of MMP and TIMP levels. © 2000 Cancer Research Campaign PMID:11027427

Retroviral gene transfer of an antisense construct against membrane type 1 matrix metalloproteinase reduces the invasiveness of rheumatoid arthritis synovial fibroblasts.

PubMed

Rutkauskaite, Edita; Volkmer, Dagmar; Shigeyama, Yukio; Schedel, Jörg; Pap, Geza; Müller-Ladner, Ulf; Meinecke, Ingmar; Alexander, Dorothea; Gay, Renate E; Drynda, Susanne; Neumann, Wolfram; Michel, Beat A; Aicher, Wilhelm K; Gay, Steffen; Pap, Thomas

2005-07-01

Membrane type 1 matrix metalloproteinase (MT1-MMP) is expressed prominently in rheumatoid arthritis synovial fibroblasts (RASFs), but the specific contribution of MT1-MMP to fibroblast-mediated destruction of articular cartilage is incompletely understood. This study used gene transfer of an antisense expression construct to assess the effects of MT1-MMP inhibition on the invasiveness of RASFs. Retroviral gene transfer of a pLXIN vector-based antisense RNA expression construct (MT1-MMPalphaS) to MT1-MMP was used to stably transduce RASFs. Levels of MT1-MMP RNA and protein were determined by quantitative polymerase chain reaction, Western blotting, and immunocytochemistry in MT1-MMPalphaS-transduced RASFs as well as in control cells, with monitoring for 60 days. The effects of MT1-MMPalphaS on the invasiveness of RASFs were analyzed in the SCID mouse co-implantation model of RA. MT1-MMPalphaS-transduced RASFs produced high levels of antisense RNA that exceeded endogenous levels of MT1-MMP messenger RNA by 15-fold and resulted in a down-regulation of MT1-MMP at the protein level. Inhibition of MT1-MMP production was maintained for 60 days and significantly reduced the invasiveness of RASFs in the SCID mouse model. Whereas prominent invasion into cartilage by non-transduced and mock-transduced RASFs was observed (mean invasion scores 3.0 and 3.1, respectively), MT1-MMPalphaS-transduced cells showed only moderate invasiveness (mean invasion score 1.8; P < 0.05). The data demonstrate that an antisense RNA expression construct against MT1-MMP can be generated and expressed in RASFs for at least 60 days. Inhibition of MT1-MMP significantly reduces the cartilage degradation by RASFs.

Activation of Nrf2 Reduces UVA-Mediated MMP-1 Upregulation via MAPK/AP-1 Signaling Cascades: The Photoprotective Effects of Sulforaphane and Hispidulin

PubMed Central

Chaiprasongsuk, Anyamanee; Lohakul, Jinaphat; Soontrapa, Kitipong; Sampattavanich, Somponnat; Akarasereenont, Pravit

2017-01-01

UVA irradiation plays a role in premature aging of the skin through triggering oxidative stress-associated stimulation of matrix metalloproteinase-1 (MMP-1) responsible for collagen degradation, a hallmark of photoaged skin. Compounds that can activate nuclear factor E2-related factor 2 (Nrf2), a transcription factor regulating antioxidant gene expression, should therefore serve as effective antiphotoaging agents. We investigated whether genetic silencing of Nrf2 could relieve UVA-mediated MMP-1 upregulation via activation of mitogen-activated protein kinase (MAPK)/activator protein 1 (AP-1) signaling using human keratinocyte cell line (HaCaT). Antiphotoaging effects of hispidulin (HPD) and sulforaphane (SFN) were assessed on their abilities to activate Nrf2 in controlling MMP-1 and collagen expressions in association with phosphorylation of MAPKs (extracellular signal-regulated kinase, c-Jun N-terminal kinase, and p38), c-Jun, and c-Fos, using the skin of BALB/c mice subjected to repetitive UVA irradiation. Our findings suggested that depletion of Nrf2 promoted both mRNA expression and activity of MMP-1 in the UVA-irradiated HaCaT cells. Treatment of Nrf2 knocked-down HaCaT cells with MAPK inhibitors significantly suppressed UVA-induced MMP-1 and AP-1 activities. Moreover, pretreatment of the mouse skin with HPD and SFN, which could activate Nrf2, provided protective effects against UVA-mediated MMP-1 induction and collagen depletion in correlation with the decreased levels of phosphorylated MAPKs, c-Jun, and c-Fos in the mouse skin. In conclusion, Nrf2 could influence UVA-mediated MMP-1 upregulation through the MAPK/AP-1 signaling cascades. HPD and SFN may therefore represent promising antiphotoaging candidates. PMID:28011874

Cationic star-shaped polymer as an siRNA carrier for reducing MMP-9 expression in skin fibroblast cells and promoting wound healing in diabetic rats.

PubMed

Li, Na; Luo, Heng-Cong; Yang, Chuan; Deng, Jun-Jie; Ren, Meng; Xie, Xiao-Ying; Lin, Diao-Zhu; Yan, Li; Zhang, Li-Ming

2014-01-01

Excessive expression of matrix metalloproteinase-9 (MMP-9) is deleterious to the cutaneous wound-healing process in the context of diabetes. The aim of the present study was to explore whether a cationic star-shaped polymer consisting of β-cyclodextrin (β-CD) core and poly(amidoamine) dendron arms (β-CD-[D₃]₇) could be used as the gene carrier of small interfering RNA (siRNA) to reduce MMP-9 expression for enhanced diabetic wound healing. The cytotoxicity of β-CD-(D₃)₇ was investigated by 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay (MMT) method in the rat CRL1213 skin fibroblast cell line. The transfection efficiency of β-CD-(D₃)₇/MMP-9-small interfering RNA (siRNA) complexes was determined by confocal microscopy and flow cytometry. Quantitative real time (RT) polymerase chain reaction was performed to measure the gene expression of MMP-9 after the transfection by β-CD-(D₃)₇/MMP-9-siRNA complexes. The β-CD-(D₃)₇/MMP-9-siRNA complexes were injected on the wounds of streptozocin-induced diabetic rats. Wound closure was measured on days 4 and 7 post-wounding. β-CD-(D₃)₇ exhibited low cytotoxicity in fibroblast cells, and easily formed the complexes with MMP-9-siRNA. The β-CD-(D₃)₇/MMP-9-siRNA complexes were readily taken up by fibroblast cells, resulting in the downregulation of MMP-9 gene expression (P<0.01). Animal experiments revealed that the treatment by β-CD-(D₃)₇/MMP-9-siRNA complexes enhanced wound closure in diabetic rats on day 7 post-wounding (P<0.05). β-CD-(D₃)₇ may be used as an efficient carrier for the delivery of MMP-9-siRNA to reduce MMP-9 expression in skin fibroblast cells and promote wound healing in diabetic rats.

Hsa-miR-875-5p exerts tumor suppressor function through down-regulation of EGFR in colorectal carcinoma (CRC)

PubMed Central

Li, Qi; Xue, Peng; Chen, Zhixiao; Dong, Xiao; Xue, Ying

2016-01-01

Hsa-miRNA-875-5p (miR-875-5p) has recently been discovered to have anticancer efficacy in different organs. However, the role of miR-875-5p on colorectal carcinoma (CRC) is still ambiguous. In this study, we investigated the role of miR-875-5p on the development of CRC. The results indicated that miR-875-5p was significantly down-regulated in primary tumor tissues and very low levels were found in CRC cell lines. Ectopic expression of miR-875-5p in CRC cell lines significantly suppressed cell growth as evidenced by cell viability assay, colony formation assay and BrdU staining, through inhibition of cyclin D1, cyclin D2, CDK4 and up-regulation of p57(Kip2) and p21(Waf1/Cip1). In addition, miR-875-5p induced apoptosis, as indicated by concomitantly with up-regulation of key apoptosis protein cleaved caspase-3, and down-regulation of anti-apoptosis protein Bcl2. Moreover, miR-875-5p inhibited cellular migration and invasiveness through inhibition of matrix metalloproteinases (MMP)-7 and MMP-9. Further, oncogene EGFR was revealed to be a putative target of miR-875-5p, which was inversely correlated with miR-875-5p expression in CRC. Taken together, our results demonstrated that miR-875-5p played a pivotal role on CRC through inhibiting cell proliferation, migration, invasion, and promoting apoptosis by targeting oncogenic EGFR. PMID:27302926

Hsa-miR-875-5p exerts tumor suppressor function through down-regulation of EGFR in colorectal carcinoma (CRC).

PubMed

Zhang, Tiening; Cai, Xun; Li, Qi; Xue, Peng; Chen, Zhixiao; Dong, Xiao; Xue, Ying

2016-07-05

Hsa-miRNA-875-5p (miR-875-5p) has recently been discovered to have anticancer efficacy in different organs. However, the role of miR-875-5p on colorectal carcinoma (CRC) is still ambiguous. In this study, we investigated the role of miR-875-5p on the development of CRC. The results indicated that miR-875-5p was significantly down-regulated in primary tumor tissues and very low levels were found in CRC cell lines. Ectopic expression of miR-875-5p in CRC cell lines significantly suppressed cell growth as evidenced by cell viability assay, colony formation assay and BrdU staining, through inhibition of cyclin D1, cyclin D2, CDK4 and up-regulation of p57(Kip2) and p21(Waf1/Cip1). In addition, miR-875-5p induced apoptosis, as indicated by concomitantly with up-regulation of key apoptosis protein cleaved caspase-3, and down-regulation of anti-apoptosis protein Bcl2. Moreover, miR-875-5p inhibited cellular migration and invasiveness through inhibition of matrix metalloproteinases (MMP)-7 and MMP-9. Further, oncogene EGFR was revealed to be a putative target of miR-875-5p, which was inversely correlated with miR-875-5p expression in CRC. Taken together, our results demonstrated that miR-875-5p played a pivotal role on CRC through inhibiting cell proliferation, migration, invasion, and promoting apoptosis by targeting oncogenic EGFR.

miR-346 and miR-582-3p-regulated EG-VEGF expression and trophoblast invasion via matrix metalloproteinases 2 and 9.

PubMed

Su, Mei-Tsz; Tsai, Pei-Yin; Tsai, Hui-Ling; Chen, Yi-Chi; Kuo, Pao-Lin

2017-03-01

Endocrine gland-derived vascular endothelial growth factor (EG-VEGF) is an important regulator for embryo implantation and placental development, and is clinically associated with several obstetric disorders related to insufficient or inappropriate trophoblast invasion, such as recurrent abortion, preeclampsia, and intrauterine fetal growth restriction. This study was performed to identify the microRNAs targeting EG-VEGF, and evaluate the regulatory effect on trophoblast biology. miR-346 and miR-582-3p were initially identified via bioinformatic tools, and their specific binding sites on the EG-VEGF 3'UTR were further confirmed using dual luciferase and a co-transfection assays. miR-346 and miR-582-3p were demonstrated not only to suppress EG-VEGF expression, but also inhibit trophoblast invasion and migration in the JAR and HTR-8/SVneo cell lines. We further evaluated the effect of microRNAs in HTR-8/SVneo cells coexpressing EG-VEGF and miR-346 or miR-582-3p on matrix metalloproteinase (MMP 2 and MMP 9) and the tissue inhibitors of metalloproteinase (TIMP 1 and TIMP 2) using RT-PCR, western blotting and gelatin zymography. TIMP 1 and TIMP 2 were not affected by the two microRNAs, whereas the expressions and activities of MMP 2 and MMP 9 were significantly downregulated, which in turn inhibited the invasion ability of trophoblasts. In conclusion, miR-346 and miR-582-3p regulate EG-VEGF-induced trophoblast invasion through repressing MMP 2 and MMP 9, and may become novel diagnostic biomarkers or therapeutic targets for EG-VEGF-related obstetric disorders. © 2016 BioFactors, 43(2):210-219, 2017. © 2016 International Union of Biochemistry and Molecular Biology.

MT2-MMP-dependent release of collagen IV NC1 domains regulates submandibular gland branching morphogenesis

PubMed Central

Rebustini, Ivan T.; Myers, Christopher; Lassiter, Keyonica S.; Surmak, Andrew; Szabova, Ludmila; Holmbeck, Kenn; Pedchenko, Vadim; Hudson, Billy G.; Hoffman, Matthew P.

2009-01-01

Summary Proteolysis is essential during branching morphogenesis, but the roles of MT-MMPs and their proteolytic products are not clearly understood. Here we discover that decreasing MT-MMP activity during submandibular gland branching morphogenesis decreases proliferation and increases collagen IV and MT-MMP expression. Importantly, reducing epithelial MT2-MMP profoundly decreases proliferation and morphogenesis, increases Col4a2 and intracellular accumulation of collagen IV, and decreases the proteolytic release of collagen IV NC1 domains. Importantly, we demonstrate the presence of collagen IV NC1 domains in developing tissue. Furthermore, recombinant collagen IV NC1 domains rescue branching morphogenesis after MT2-siRNA-treatment, increasing MT-MMP and pro-proliferative gene expression via β1 integrin and PI3K-AKT signaling. Additionally, HBEGF also rescues MT2-siRNA-treatment, increasing NC1 domain release, proliferation, and MT2-MMP and Hbegf expression. Our studies provide mechanistic insight into how MT2-MMP-dependent release of bioactive NC1 domains from collagen IV is critical for integrating collagen IV synthesis and proteolysis with epithelial proliferation during branching morphogenesis. PMID:19853562

Poly(I:C) induces expressions of MMP-1, -2, and -3 through various signaling pathways including IRF3 in human skin fibroblasts.

PubMed

Yao, Cheng; Lee, Dong Hun; Oh, Jang-Hee; Kim, Min-Kyoung; Kim, Kyu Han; Park, Chi-Hyun; Chung, Jin Ho

2015-10-01

Ultraviolet (UV) irradiation can result in premature skin aging (photoaging) which is characterized by decreased expression of collagen and increased expression of matrix metalloproteinases (MMPs). Double-stranded RNAs (dsRNAs) can be generated at various conditions including virally infected cells or UV-damaged skin cells. Recent studies have shown that a synthetic dsRNA, polyinosinic-polycytidylic acid (poly(I:C)), can reduce procollagen expression in human skin fibroblasts. However, little is known about the effect of poly(I:C) on the expression of MMPs in skin fibroblasts and its underlying mechanisms. We examined the effect of poly(I:C) on MMP-1, -2, and -3 expressions in human skin fibroblasts. Then, we further explored the underlying signaling pathways involved in the processes. Human skin fibroblasts were treated with poly(I:C) for the indicated times in the presence or the absence of various chemical inhibitors or small interfering RNAs (siRNAs) at the indicated concentrations. Protein and mRNA levels of various target molecules were examined by Western blotting and quantitative real-time PCR, respectively. Poly(I:C) induced MMP-1, -2, and -3 expressions, which were dependent on TLR3. Poly(I:C) also induced activations of the mitogen-activated protein kinases (MAPKs), the nuclear factor-kappaB (NF-κB) and the interferon regulatory factor 3 (IRF3) pathways. By using specific inhibitors, we found that poly(I:C)-induced expressions of MMP-1, -2, and -3 were differentially regulated by these signaling pathways. In particular, we found that the inhibition of IRF3 signaling pathways attenuated poly(I:C)-induced expressions of all the three MMPs. Our data show that the expressions of MMP-1, -2, and -3 are induced by poly(I:C) through various signaling pathways in human skin fibroblasts and suggest that TLR3 and/or IRF3 may be good targets for regulating the expressions of MMP-1, -2, and -3 induced by dsRNAs. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

Isolation and characterization of chicken bile matrix metalloproteinase.

PubMed

Packialakshmi, B; Liyanage, R; Rasaputra, K S; Lay, Jackson O; Rath, N C

2014-06-01

Avian bile is rich in matrix metalloproteinases (MMP), the enzymes that cleave extracellular matrix proteins such as collagens and proteoglycans. Changes in bile MMP expression have been correlated with hepatic and gall bladder pathologies, but the significance of their expression in normal, healthy bile is not understood. We hypothesized that the MMP in bile may aid the digestion of native collagens that are resistant to conventional gastric proteases. Hence, the objective of this study was to characterize the bile MMP and check its regulation in association with dietary factors. We used substrate zymography, azocoll protease assay, and gelatin affinity chromatography to identify and purify the MMP from chicken bile. Using zymography and SDS PAGE, 5 bands at 70, 64, 58, 50, and 42 kDa were detected. The bands corresponding to 64, 50, and 42 kDa were identified as MMP2 using trypsin in-gel digestion and matrix-assisted laser desorption time-of-flight mass spectrometry and peptide mass fingerprinting. Chickens fed diets containing gelatin supplements showed higher levels of MMP expression in the bile by both azocoll assay and zymography. We conclude that the bile MMP may be associated with the digestion of collagens and other extracellular matrix proteins in avian diets. Poultry Science Association Inc.

Downregulation of CD9 in Keratinocyte Contributes to Cell Migration via Upregulation of Matrix Metalloproteinase-9

PubMed Central

Jiang, Xu-pin; Zhang, Dong-xia; Teng, Miao; Zhang, Qiong; Zhang, Jia-ping; Huang, Yue-sheng

2013-01-01

Tetraspanin CD9 has been implicated in various cellular and physiological processes, including cell migration. In our previous study, we found that wound repair is delayed in CD9-null mice, suggesting that CD9 is critical for cutaneous wound healing. However, many cell types, including immune cells, endothelial cells, keratinocytes and fibroblasts undergo marked changes in gene expression and phenotype, leading to cell proliferation, migration and differentiation during wound repair, whether CD9 regulates kerationcytes migration directly remains unclear. In this study, we showed that the expression of CD9 was downregulated in migrating keratinocytes during wound repair in vivo and in vitro. Recombinant adenovirus vector for CD9 silencing or overexpressing was constructed and used to infect HaCaT cells. Using cell scratch wound assay and cell migration assay, we have also demonstrated that downregulation of CD9 promoted keratinocyte migration in vitro, whereas CD9 overexpression inhibited cell migration. Moreover, CD9 inversely regulated the activity and expression of MMP-9 in keratinocytes, which was involved in CD9-regulated keratinocyte migration. Importantly, CD9 silencing-activated JNK signaling was accompanied by the upregulation of MMP-9 activity and expression. Coincidentally, we found that SP600125, a JNK pathway inhibitor, decreased the activity and expression of MMP-9 of CD9-silenced HaCaT cells. Thus, our results suggest that CD9 is downregulated in migrating keratinocytes in vivo and in vitro, and a low level of CD9 promotes keratinocyte migration in vitro, in which the regulation of MMP-9 through the JNK pathway plays an important role. PMID:24147081

Mmp1 processing of the PDF neuropeptide regulates circadian structural plasticity of pacemaker neurons.

PubMed

Depetris-Chauvin, Ana; Fernández-Gamba, Agata; Gorostiza, E Axel; Herrero, Anastasia; Castaño, Eduardo M; Ceriani, M Fernanda

2014-10-01

In the Drosophila brain, the neuropeptide PIGMENT DISPERSING FACTOR (PDF) is expressed in the small and large Lateral ventral neurons (LNvs) and regulates circadian locomotor behavior. Interestingly, PDF immunoreactivity at the dorsal terminals changes across the day as synaptic contacts do as a result of a remarkable remodeling of sLNv projections. Despite the relevance of this phenomenon to circuit plasticity and behavior, the underlying mechanisms remain poorly understood. In this work we provide evidence that PDF along with matrix metalloproteinases (Mmp1 and 2) are key in the control of circadian structural remodeling. Adult-specific downregulation of PDF levels per se hampers circadian axonal remodeling, as it does altering Mmp1 or Mmp2 levels within PDF neurons post-developmentally. However, only Mmp1 affects PDF immunoreactivity at the dorsal terminals and exerts a clear effect on overt behavior. In vitro analysis demonstrated that PDF is hydrolyzed by Mmp1, thereby suggesting that Mmp1 could directly terminate its biological activity. These data demonstrate that Mmp1 modulates PDF processing, which leads to daily structural remodeling and circadian behavior.

Mmp1 Processing of the PDF Neuropeptide Regulates Circadian Structural Plasticity of Pacemaker Neurons

PubMed Central

Depetris-Chauvin, Ana; Fernández-Gamba, Ágata; Gorostiza, E. Axel; Herrero, Anastasia; Castaño, Eduardo M.; Ceriani, M. Fernanda

2014-01-01

In the Drosophila brain, the neuropeptide PIGMENT DISPERSING FACTOR (PDF) is expressed in the small and large Lateral ventral neurons (LNvs) and regulates circadian locomotor behavior. Interestingly, PDF immunoreactivity at the dorsal terminals changes across the day as synaptic contacts do as a result of a remarkable remodeling of sLNv projections. Despite the relevance of this phenomenon to circuit plasticity and behavior, the underlying mechanisms remain poorly understood. In this work we provide evidence that PDF along with matrix metalloproteinases (Mmp1 and 2) are key in the control of circadian structural remodeling. Adult-specific downregulation of PDF levels per se hampers circadian axonal remodeling, as it does altering Mmp1 or Mmp2 levels within PDF neurons post-developmentally. However, only Mmp1 affects PDF immunoreactivity at the dorsal terminals and exerts a clear effect on overt behavior. In vitro analysis demonstrated that PDF is hydrolyzed by Mmp1, thereby suggesting that Mmp1 could directly terminate its biological activity. These data demonstrate that Mmp1 modulates PDF processing, which leads to daily structural remodeling and circadian behavior. PMID:25356918

Decursin inhibits UVB-induced MMP expression in human dermal fibroblasts via regulation of nuclear factor-κB.

PubMed

Hwang, Bo-Mi; Noh, Eun-Mi; Kim, Jong-Suk; Kim, Jeong-Mi; Hwang, Jin-Ki; Kim, Hye-Kyung; Kang, Jae-Seon; Kim, Do-Sung; Chae, Han-Jung; You, Yong-Ouk; Kwon, Kang-Beom; Lee, Young-Rae

2013-02-01

Decursin, a coumarin compound, was originally isolated from the roots of Angelica gigas almost four decades ago, and it was found to exhibit cytotoxicity against various types of human cancer cells and anti-amnesic activity in vivo through the inhibition of AChE activity. However, the anti-skin photoaging effects of decursin have not been reported to date. In the present study, we investigated the inhibitory effects of decursin on the expression of matrix metalloproteinase (MMP)-1 and MMP-3 in human dermal fibroblast (HDF) cells. Western blot analysis and real-time PCR revealed that decursin inhibited the ultraviolet (UV)B-induced expression of MMP-1 and MMP-3 in a dose-dependent manner. Decursin significantly blocked the UVB-induced activation of nuclear factor-κB (NF-κB). However, decursin showed no effect on MAPK or AP-1 activity. In this study, decursin prevented the UVB-induced expression of MMPs via the inhibition of NF-κB activation. In conclusion, decursin may be a potential agent for the prevention and treatment of skin photoaging.

MMP20 Overexpression Disrupts Molar Ameloblast Polarity and Migration.

PubMed

Shin, M; Chavez, M B; Ikeda, A; Foster, B L; Bartlett, J D

2018-07-01

Ameloblasts responsible for enamel formation express matrix metalloproteinase 20 (MMP20), an enzyme that cleaves enamel matrix proteins, including amelogenin (AMELX) and ameloblastin (AMBN). Previously, we showed that continuously erupting incisors from transgenic mice overexpressing active MMP20 had a massive cell infiltrate present within their enamel space, leading to enamel mineralization defects. However, effects of MMP20 overexpression on mouse molars were not analyzed, although these teeth more accurately represent human odontogenesis. Therefore, MMP20-overexpressing mice ( Mmp20 +/+ Tg + ) were assessed by multiscale analyses, combining several approaches from high-resolution micro-computed tomography to enamel organ immunoblots. During the secretory stage at postnatal day 6 (P6), Mmp20 +/+ Tg + mice had a discontinuous ameloblast layer and, unlike incisors, molar P12 maturation stage ameloblasts abnormally migrated away from the enamel layer into the stratum intermedium/stellate reticulum. TOPflash assays performed in vitro demonstrated that MMP20 expression promoted β-catenin nuclear localization and that MMP20 expression promoted invasion through Matrigel-coated filters. However, for both assays, significant differences were eliminated in the presence of the β-catenin inhibitor ICG-001. This suggests that MMP20 activity promotes cell migration via the Wnt pathway. In vivo, the unique molar migration of amelogenin-expressing ameloblasts was associated with abnormal deposition of ectopic calcified nodules surrounding the adherent enamel layer. Enamel content was assessed just prior to eruption at P15. Compared to wild-type, Mmp20 +/+ Tg + molars exhibited significant reductions in enamel thickness (70%), volume (60%), and mineral density (40%), and MMP20 overexpression resulted in premature cleavage of AMBN, which likely contributed to the severe defects in enamel mineralization. In addition, Mmp20 +/+ Tg + mouse molar enamel organs had increased levels of inactive p-cofilin, a protein that regulates cell polarity. These data demonstrate that increased MMP20 activity in molars causes premature degradation of ameloblastin and inactivation of cofilin, which may contribute to pathological Wnt-mediated cell migration away from the enamel layer.

Metalloproteinases and their associated genes contribute to the functional integrity and noise-induced damage in the cochlear sensory epithelium

PubMed Central

Hu, Bo Hua; Cai, Qunfeng; Hu, Zihua; Patel, Minal; Bard, Jonathan; Jamison, Jennifer; Coling, Donald

2012-01-01

Matrix metalloproteinases (MMPs) and their related gene products regulate essential cellular functions. An imbalance in MMPs has been implicated in various neurological disorders, including traumatic injuries. Here, we report a role for MMPs and their related gene products in the modulation of cochlear responses to acoustic trauma in rats. The normal cochlea was shown to be enriched in MMP enzymatic activity, and this activity was reduced in a time-dependent fashion after traumatic noise injury. The analysis of gene expression by RNA-seq and qRT-PCR revealed the differential expression of MMPs and their related genes between functionally specialized regions of the sensory epithelium. The expression of these genes was dynamically regulated between the acute and chronic phases of noise-induced hearing loss. Moreover, noise-induced expression changes in two endogenous MMP inhibitors, Timp1 and Timp2, in sensory cells were dependent upon the stage of nuclear condensation, suggesting a specific role for MMP activity in sensory cell apoptosis. A short-term application of doxycycline, a broad-spectrum inhibitor of MMPs, prior to noise exposure reduced noise-induced hearing loss and sensory cell death. By contrast, a 7-day treatment compromised hearing sensitivity and potentiated noise-induced hearing loss. This detrimental effect of the long-term inhibition of MMPs on noise-induced hearing loss was further confirmed using targeted Mmp7 knockout mice. Together, these observations suggest that MMPs and their related genes participate in the regulation of cochlear responses to acoustic overstimulation and that the modulation of MMP activity can serve as a novel therapeutic target for the reduction of noise-induced cochlear damage. PMID:23100416

Leptin modulates the expression of catabolic genes in rat nucleus pulposus cells through the mitogen-activated protein kinase and Janus kinase 2/signal transducer and activator of transcription 3 pathways.

PubMed

Miao, Daoyi; Zhang, Lingzhou

2015-08-01

Obesity has been demonstrated to be involved in the progress of intervertebral disc degeneration (IDD). However, the associated mechanisms remain to be elucidated. The purpose the present study was to examine the effect of leptin on the expression of degeneration-associated genes in rat nucleus pulposus (NP) cells, and determine the possible mechanism. Normal NP cells, obtained from Sprague Dawley rats, were identified using immunocytochemistry for the expression of collagen II and CA125, and treated with leptin and/or interleukin (IL)-β. Subsequently, the mRNA expression levels of matrix metalloproteinase (MMP)-1, MMP-3, MMP-9, MMP-13, a disintegrin and metalloproteinase with thrombospondin motifs (ADAMTS)-4, ADAMTS-5, aggrecan and COL2A1 were detected by reverse transcription-quantitative polymerase chain reaction (RT-q-PCR). Alcian staining and immunocytochemistry were used to examine the expression levels of proteoglycan and collagen II. The pathway activation was investigated using western blotting, and inhibitors of the pathways were used to reveal the effect of these pathways on the NP cells. The results of the RT-qPCR demonstrated that leptin alone upregulated the mRNA expression levels of MMP-1, MMP-13, ADAMTS-4, ADAMTS-5 and COL2A1. Synergy of leptin and IL-β was found in the increased expression levels of MMP-1, MMP-3 and ADAMTS-5. The leptin-treated NP cells exhibited decreased expression of collagen II. The mitrogen-activated protein kinase (MAPK) pathway (c-Jun-N-terminal kinase, phosphorylated extracellular signal-regulated kinase and p38), phosphatidylinositol 3-kinase (PI3K)/Akt pathway and Janus kinase (JAK)2/signal transducer and activator of transcription 3 pathway were all activated by leptin, however, inhibitors of all the pathways, with the exception of the PI3K/Akt pathway, reversed the expression levels of MMP-1 and MMP-13. These results suggested that leptin promoted catabolic metabolism in the rat NP cells via the MAPK and JAK2/STAT3 pathways, which may be the mechanism mediating the association between obesity and IDD.

Leptin modulates the expression of catabolic genes in rat nucleus pulposus cells through the mitogen-activated protein kinase and Janus kinase 2/signal transducer and activator of transcription 3 pathways

PubMed Central

MIAO, DAOYI; ZHANG, LINGZHOU

2015-01-01

Obesity has been demonstrated to be involved in the progress of intervertebral disc degeneration (IDD). However, the associated mechanisms remain to be elucidated. The purpose the present study was to examine the effect of leptin on the expression of degeneration-associated genes in rat nucleus pulposus (NP) cells, and determine the possible mechanism. Normal NP cells, obtained from Sprague Dawley rats, were identified using immunocytochemistry for the expression of collagen II and CA125, and treated with leptin and/or interleukin (IL)-β. Subsequently, the mRNA expression levels of matrix metalloproteinase (MMP)-1, MMP-3, MMP-9, MMP-13, a disintegrin and metalloproteinase with thrombospondin motifs (ADAMTS)-4, ADAMTS-5, aggrecan and COL2A1 were detected by reverse transcription-quantitative polymerase chain reaction (RT-q-PCR). Alcian staining and immunocytochemistry were used to examine the expression levels of proteoglycan and collagen II. The pathway activation was investigated using western blotting, and inhibitors of the pathways were used to reveal the effect of these pathways on the NP cells. The results of the RT-qPCR demonstrated that leptin alone upregulated the mRNA expression levels of MMP-1, MMP-13, ADAMTS-4, ADAMTS-5 and COL2A1. Synergy of leptin and IL-β was found in the increased expression levels of MMP-1, MMP-3 and ADAMTS-5. The leptin-treated NP cells exhibited decreased expression of collagen II. The mitrogen-activated protein kinase (MAPK) pathway (c-Jun-N-terminal kinase, phosphorylated extracellular signal-regulated kinase and p38), phosphatidylinositol 3-kinase (PI3K)/Akt pathway and Janus kinase (JAK)2/signal transducer and activator of transcription 3 pathway were all activated by leptin, however, inhibitors of all the pathways, with the exception of the PI3K/Akt pathway, reversed the expression levels of MMP-1 and MMP-13. These results suggested that leptin promoted catabolic metabolism in the rat NP cells via the MAPK and JAK2/STAT3 pathways, which may be the mechanism mediating the association between obesity and IDD. PMID:25892402

Down-regulation of Gab1 inhibits cell proliferation and migration in hilar cholangiocarcinoma.

PubMed

Sang, Haiquan; Li, Tingting; Li, Hangyu; Liu, Jingang

2013-01-01

Hilar cholangiocarcinoma is a highly aggressive malignancy originating from the hilar biliary duct epithelium. Due to few effective comprehensive treatments, the prognosis of hilar cholangiocarcinoma is poor. In this study, immunohistochemistry was first used to detect and analyze the expression of Gab1, VEGFR-2, and MMP-9 in hilar cholangiocarcinoma solid tumors and the relationships to the clinical pathological features. Furthermore, Gab1 and VEGFR-2 siRNA were used to interfere the hilar cholangiocarcinoma cell line ICBD-1 and then detect the PI3K/Akt signaling pathway, MMP-9 levels and malignant biological behaviors of tumor cells. The data showed that 1. Gab1, VEGFR-2, and MMP-9 were highly expressed and positively correlated with each other in hilar cholangiocarcinoma tissues, which were related to lymph node metastasis and differentiation. 2. After Gab1 or VEGFR-2 siRNA interference, PI3K/Akt pathway activity and MMP-9 levels were decreased in ICBD-1 cells. At the same time, cell proliferation decreased, cell cycle arrested in G1 phase, apoptosis increased and invasion decreased. These results suggest that the expression of Gab1, VEGFR-2, and MMP-9 are significantly related to the malignant biological behavior of hilar cholangiocarcinoma. Gab1 regulates growth, apoptosis and invasion through the VEGFR-2/Gab1/PI3K/Akt signaling pathway in hilar cholangiocarcinoma cells and influences the invasion of tumor cells via MMP-9.

Down-Regulation of Gab1 Inhibits Cell Proliferation and Migration in Hilar Cholangiocarcinoma

PubMed Central

Sang, Haiquan; Li, Tingting; Li, Hangyu; Liu, Jingang

2013-01-01

Hilar cholangiocarcinoma is a highly aggressive malignancy originating from the hilar biliary duct epithelium. Due to few effective comprehensive treatments, the prognosis of hilar cholangiocarcinoma is poor. In this study, immunohistochemistry was first used to detect and analyze the expression of Gab1, VEGFR-2, and MMP-9 in hilar cholangiocarcinoma solid tumors and the relationships to the clinical pathological features. Furthermore, Gab1 and VEGFR-2 siRNA were used to interfere the hilar cholangiocarcinoma cell line ICBD-1 and then detect the PI3K/Akt signaling pathway, MMP-9 levels and malignant biological behaviors of tumor cells. The data showed that 1. Gab1, VEGFR-2, and MMP-9 were highly expressed and positively correlated with each other in hilar cholangiocarcinoma tissues, which were related to lymph node metastasis and differentiation. 2. After Gab1 or VEGFR-2 siRNA interference, PI3K/Akt pathway activity and MMP-9 levels were decreased in ICBD-1 cells. At the same time, cell proliferation decreased, cell cycle arrested in G1 phase, apoptosis increased and invasion decreased. These results suggest that the expression of Gab1, VEGFR-2, and MMP-9 are significantly related to the malignant biological behavior of hilar cholangiocarcinoma. Gab1 regulates growth, apoptosis and invasion through the VEGFR-2/Gab1/PI3K/Akt signaling pathway in hilar cholangiocarcinoma cells and influences the invasion of tumor cells via MMP-9. PMID:24312291

The MMP-9 -1562 C/T Polymorphism in the Presence of Metabolic Syndrome Increases the Risk of Clinical Events in Patients with Coronary Artery Disease

PubMed Central

Opstad, Trine B.; Arnesen, Harald; Pettersen, Alf Å.; Seljeflot, Ingebjørg

2014-01-01

Background and Objectives Elevated levels of matrix metalloproteinase (MMP)-9 have been associated with the metabolic syndrome (MetS) and cardiovascular events. The MMP-9 −1562 C/T polymorphism has furthermore been shown as a risk factor for coronary artery disease (CAD). The non-favourable cardiometabolic state in MetS may increase the risk. We aimed to investigate the influence of MMP-9 −1562 C/T polymorphism in subjects with CAD and MetS. Methods Patients (n = 1000) with verified CAD stratified in Mets +/− (n = 244/756), were analyzed for the MMP-9 −1562 C/T polymorphism and related to clinical events after 2 years follow-up. Serum levels of total MMP-9 and tissue inhibitor of matrix metalloproteinases (TIMP)-1were analyzed in all, whereas MMP-9 activity, extracellular matrix metalloproteinase inducer (EMMPRIN), and expression of the two genes were analyzed in a subset of 240 randomly selected patients. Results Totally, 106 clinical endpoints were recorded. In MetS; the T-allele associated with 5.5 fold increase in event rate (p<0.0001), increased with number of MetS components, a 117% increase in total MMP-9 levels (TT homozygous, p = 0.05), significantly higher total- and endogenous active MMP-9 and TIMP-1 levels (p<0.01 all), and EMMPRIN was inversely correlated with pro- and endogenous active MMP-9 (p<0.05, both). In non-MetS; the T-allele was not associated with new events, nor higher MMP-9 levels. EMMPRIN was significantly correlated with total MMP-9 and TIMP-1 (p<0.01, both) and the two genes were inter-correlated (p<0.001). Conclusion In CAD patients with MetS, the MMP-9 T-allele increased the risk of clinical events, probably mediated through elevated MMP-9 levels and altered MMP-9 regulation. PMID:25191702

The MMP-9 -1562 C/T polymorphism in the presence of metabolic syndrome increases the risk of clinical events in patients with coronary artery disease.

PubMed

Opstad, Trine B; Arnesen, Harald; Pettersen, Alf Å; Seljeflot, Ingebjørg

2014-01-01

Elevated levels of matrix metalloproteinase (MMP)-9 have been associated with the metabolic syndrome (MetS) and cardiovascular events. The MMP-9 -1562 C/T polymorphism has furthermore been shown as a risk factor for coronary artery disease (CAD). The non-favourable cardiometabolic state in MetS may increase the risk. We aimed to investigate the influence of MMP-9 -1562 C/T polymorphism in subjects with CAD and MetS. Patients (n = 1000) with verified CAD stratified in Mets +/- (n = 244/756), were analyzed for the MMP-9 -1562 C/T polymorphism and related to clinical events after 2 years follow-up. Serum levels of total MMP-9 and tissue inhibitor of matrix metalloproteinases (TIMP)-1 were analyzed in all, whereas MMP-9 activity, extracellular matrix metalloproteinase inducer (EMMPRIN), and expression of the two genes were analyzed in a subset of 240 randomly selected patients. Totally, 106 clinical endpoints were recorded. In MetS; the T-allele associated with 5.5 fold increase in event rate (p<0.0001), increased with number of MetS components, a 117% increase in total MMP-9 levels (TT homozygous, p = 0.05), significantly higher total- and endogenous active MMP-9 and TIMP-1 levels (p<0.01 all), and EMMPRIN was inversely correlated with pro- and endogenous active MMP-9 (p<0.05, both). In non-MetS; the T-allele was not associated with new events, nor higher MMP-9 levels. EMMPRIN was significantly correlated with total MMP-9 and TIMP-1 (p<0.01, both) and the two genes were inter-correlated (p<0.001). In CAD patients with MetS, the MMP-9 T-allele increased the risk of clinical events, probably mediated through elevated MMP-9 levels and altered MMP-9 regulation.

Regulation of invadopodia formation and activity by CD147

PubMed Central

Grass, G. Daniel; Bratoeva, Momka; Toole, Bryan P.

2012-01-01

A defining feature of malignant tumor progression is cellular penetration through the basement membrane and interstitial matrices that separate various cellular compartments. Accumulating evidence supports the notion that invasive cells employ specialized structures termed invadopodia to breach these structural barriers. Invadopodia are actin-based, lipid-raft-enriched membrane protrusions containing membrane-type-1 matrix metalloproteinase (MT1-MMP; also known as matrix metalloproteinase 14; MMP14) and several signaling proteins. CD147 (emmprin, basigin), an immunoglobulin superfamily protein that is associated with tumor invasion and metastasis, induces the synthesis of various matrix metalloproteinases in many systems. In this study we show that upregulation of CD147 is sufficient to induce MT1-MMP expression, invasiveness and formation of invadopodia-like structures in non-transformed, non-invasive, breast epithelial cells. We also demonstrate that CD147 and MT1-MMP are in close proximity within these invadopodia-like structures and co-fractionate in membrane compartments with the properties of lipid rafts. Moreover, manipulation of CD147 levels in invasive breast carcinoma cells causes corresponding changes in MT1-MMP expression, invasiveness and invadopodia formation and activity. These findings indicate that CD147 regulates invadopodia formation and activity, probably through assembly of MT1-MMP-containing complexes within lipid-raft domains of the invadopodia. PMID:22389410

An ex vivo ruminal ovine model to study the immediate immune response in the context of bacterial lipopolysaccharide.

PubMed

Abouhajer, Fathi; El-Ashram, Saeed; Karama, Musafiri; Huang, Shujian; Liu, Jian-Feng

2018-05-01

We have set up an ex vivo ovine ruminal model, which can mimic the multicellular process to explore the early steps in Salmonella typhimurium lipopolysaccharide (LPS) stimulation using RNA-seq technology. Ovine ruminal explants were collected for histological and transcriptional analysis and supernatants collected to quantitate lactate dehydrogenase (LDH) enzymes. A total of 8 and 523 genes were significantly over-expressed between LPS-treated and control tissues at 6 and 12 h, respectively. However, six and seven hundred and thirteen genes were substantially repressed between the aforementioned tissues, correspondingly. Key genes up-regulated in response to the addition of LPS were tumor necrosis factor (TNF), interlukin (IL)-1 beta(b), IL-6, IL-8, IL-17B, IL-19, MMP-1, MMP-3, and integrin alpha 2 (ITGA8, 9). This study shows for the first time that galectin-1 is up-regulated in an ex vivo ruminal segment model exposed to bacterial lipopolysaccharide following 6 h of incubation. The ruminal segment model has been shown to be a suitable tool to study the bacterial lipopolysaccharide effects on the ovine ruminal tissues prior to in vivo assessment.

Cell surface chondroitin sulfate glycosaminoglycan in melanoma: role in the activation of pro-MMP-2 (pro-gelatinase A)

PubMed Central

Iida, Joji; Wilhelmson, Krista L.; Ng, Janet; Lee, Peter; Morrison, Charlotte; Tam, Eric; Overall, Christopher M.; McCarthy, James B.

2007-01-01

We previously reported that CS (chondroitin sulfate) GAG (glycosaminoglycan), expressed on MCSP (melanoma-specific CS proteoglycan), is important for regulating MT3-MMP [membrane-type 3 MMP (matrix metalloproteinase)]-mediated human melanoma invasion and gelatinolytic activity in vitro. In the present study, we sought to determine if CS can directly enhance MT3-MMP-mediated activation of pro-MMP-2. Co-immunoprecipitation studies suggest that MCSP forms a complex with MT3-MMP and MMP-2 on melanoma cell surface. When melanoma cells were treated with βDX (p-nitro-β-D-xylopyranoside) to inhibit coupling of CS on the core protein, both active form and proform of MMP-2 were no longer co-immunoprecipitated with either MCSP or MT3-MMP, suggesting a model in which CS directly binds to MMP-2 and presents the gelatinase to MT3-MMP to be activated. By using recombinant proteins, we determined that MT3-MMP directly activates pro-MMP-2 and that this activation requires the interaction of the C-terminal domain of pro-MMP-2 with MT3-MMP. Activation of pro-MMP-2 by suboptimal concentrations of MT3-MMP is also significantly enhanced in the presence of excess C4S (chondroitin 4-sulfate), whereas C6S (chondroitin 6-sulfate) or low-molecular-mass hyaluronan was ineffective. Affinity chromatography studies using CS isolated from aggrecan indicate that the catalytic domain of MT3-MMP and the C-terminal domain of MMP-2 directly bind to the GAG. Thus the direct binding of pro-MMP-2 with CS through the C-domain would present the catalytic domain of pro-MMP-2 to MT3-MMP, which facilitates the generation of the active form of MMP-2. These results suggest that C4S, which is expressed on tumour cell surface, can function to bind to pro-MMP-2 and facilitate its activation by MT3-MMP-expressing tumour cells to enhance invasion and metastasis. PMID:17217338

Failure of matrix metalloproteinase-9 dimer induction by phorbol 12-myristate 13-acetate in normal human cell lines.

PubMed

Waheed Roomi, Mohd; Kalinovsky, Tatiana; Rath, Matthias; Niedzwiecki, Aleksandra

2015-06-01

Increasing experimental and clinical data has identified an association between increased levels of matrix metalloproteinase (MMP)-9 and shortened patient survival, cancer progression and metastasis. MMP-9 has a significant role in tumor cell invasion and metastasis, as it digests the basement membrane and components of the extracellular matrix. MMP-9 is secreted in either a monomeric or dimeric form. Although limited evidence exists concerning MMP-9 dimers, certain studies have demonstrated that the dimer is associated with aggressive tumor progression. This is believed to be due to the fact that cellular migration depends upon the MMP-9 dimer, and not the monomer. Our previous study revealed that cancer cell MMP-9 dimer secretion patterns could be divided into different categories, and that high MMP-9 and MMP-9 dimer secretion levels were correlated with the most aggressive cancer cell lines. It has been established that signal transduction pathways and cytokines, including those activated by phorbol 12-myristate 13-acetate (PMA), regulate the expression of MMPs. The aim of the present study was to analyze the expression patterns of MMP-2, MMP-9 and MMP-9 dimer in normal human cells from a number of tissues treated with PMA. Muscle, epithelial and connective tissues were selected for use in the present study, since adenosarcomas, carcinomas and sarcomas are derived from these tissue types, respectively. The cell lines were first cultured in 24-well tissue culture plates containing recommended media that was supplemented with 10% fetal bovine serum and antibiotics. When at confluency, the cells were washed and fresh medium was added. In addition, a parallel set of cultures was treated with PMA. Subsequent to a 24-h incubation period, the media were collected and analyzed using gelatinase zymography for the expression of MMP-2 and MMP-9 monomer and dimer forms. The results revealed that the cellular expression of MMP-2 and MMP-9 was dependent upon the primary tissue subtype. All cell lines, regardless of tissue origin, expressed MMP-2. PMA induced the expression of MMP-9 in muscle tissue, glandular epithelia and supportive connective tissue cell lines. By contrast, cell lines of endothelial origin and proper connective tissue were insensitive to treatment with PMA. MMP-9 dimer secretion was not observed in any of the cell lines, which indicated that cellular migration is not supported by these cells.

miR-132 Regulates Dendritic Spine Structure by Direct Targeting of Matrix Metalloproteinase 9 mRNA.

PubMed

Jasińska, Magdalena; Miłek, Jacek; Cymerman, Iwona A; Łęski, Szymon; Kaczmarek, Leszek; Dziembowska, Magdalena

2016-09-01

Mir-132 is a neuronal activity-regulated microRNA that controls the morphology of dendritic spines and neuronal transmission. Similar activities have recently been attributed to matrix metalloproteinase-9 (MMP-9), an extrasynaptic protease. In the present study, we provide evidence that miR-132 directly regulates MMP-9 mRNA in neurons to modulate synaptic plasticity. With the use of luciferase reporter system, we show that miR-132 binds to the 3'UTR of MMP-9 mRNA to regulate its expression in neurons. The overexpression of miR-132 in neurons reduces the level of endogenous MMP-9 protein secretion. In synaptoneurosomes, metabotropic glutamate receptor (mGluR)-induced signaling stimulates the dissociation of miR-132 from polyribosomal fractions and shifts it towards the messenger ribonucleoprotein (mRNP)-containing fraction. Furthermore, we demonstrate that the overexpression of miR-132 in the cultured hippocampal neurons from Fmr1 KO mice that have increased synaptic MMP-9 level provokes enlargement of the dendritic spine heads, a process previously implicated in enhanced synaptic plasticity. We propose that activity-dependent miR-132 regulates structural plasticity of dendritic spines through matrix metalloproteinase 9.

Cyclic tensile strain and cyclic hydrostatic pressure differentially regulate expression of hypertrophic markers in primary chondrocytes.

PubMed

Wong, Marcy; Siegrist, Mark; Goodwin, Kelly

2003-10-01

Endochondral ossification is regulated by many factors, including mechanical stimuli, which can suppress or accelerate chondrocyte maturation. Mathematical models of endochondral ossification have suggested that tension (or shear stress) can accelerate the formation of endochondral bone, while hydrostatic stress preserves the cartilage phenotype. The goal of this study was to test this hypothesis by examining the expression of hypertrophic chondrocyte markers (transcription factor Cbfa1, MMP-13, type X collagen, VEGF, CTGF) and cartilage matrix proteins under cyclic tension and cyclic hydrostatic pressure. Chondrocyte-seeded alginate constructs were exposed to one of the two loading modes for a period of 3 h per day for 3 days. Gene expression was analyzed using real-time RT-PCR. Cyclic tension upregulated the expression of Cbfa1, MMP-13, CTGF, type X collagen and VEGF and downregulated the expression of TIMP-1. Cyclic tension also upregulated the expression of type 2 collagen, COMP and lubricin, but did not change the expression of SOX9 and aggrecan. Cyclic hydrostatic pressure downregulated the expression of MMP-13 and type I collagen and upregulated expression of TIMP-1 compared to the unloaded controls. Hydrostatic pressure may slow chondrocyte differentiation and have a chondroprotective, anti-angiogenic influence on cartilage tissue. Our results suggest that cyclic tension activates the Cbfa1/MMP-13 pathway and increases the expression of terminal differentiation hypertrophic markers. Mammalian chondrocytes appear to have evolved complex mechanoresponsive mechanisms, the effects of which can be observed in the histomorphologic establishment of the cartilaginous skeleton during development and maturation.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Xue, Gang; Zou, Xi; Zhou, Jin-Yong

Highlights: •Raddeanin A is a triterpenoid saponin in herb medicine Anemone raddeana Regel. •Raddeanin A can inhibit 3 kinds of gastric cancer cells’ proliferation and invasion. •Caspase-cascades’ activation indicates apoptosis induced by Raddeanin A. •MMPs, RECK, Rhoc and E-cad are involved in Raddeanin A-induced invasion inhibition. -- Abstract: Raddeanin A is one of the triterpenoid saponins in herbal medicine Anemone raddeana Regel which was reported to suppress the growth of liver and lung cancer cells. However, little was known about its effect on gastric cancer (GC) cells. This study aimed to investigate its inhibitory effect on three kinds of differentmore » differentiation stage GC cells (BGC-823, SGC-7901 and MKN-28) in vitro and the possible mechanisms. Proliferation assay and flow cytometry demonstrated Raddeanin A’s dose-dependent inhibitory effect and determined its induction of cells apoptosis, respectively. Transwell assay, wounding heal assay and cell matrix adhesion assay showed that Raddeanin A significantly inhibited the abilities of the invasion, migration and adhesion of the BGC-823 cells. Moreover, quantitative real time PCR and Western blot analysis found that Raddeanin A increased Bax expression while reduced Bcl-2, Bcl-xL and Survivin expressions and significantly activated caspase-3, caspase-8, caspase-9 and poly-ADP ribose polymerase (PARP). Besides, Raddeanin A could also up-regulate the expression of reversion inducing cysteine rich protein with Kazal motifs (RECK), E-cadherin (E-cad) and down-regulate the expression of matrix metalloproteinases-2 (MMP-2), MMP-9, MMP-14 and Rhoc. In conclusion, Raddeanin A inhibits proliferation of human GC cells, induces their apoptosis and inhibits the abilities of invasion, migration and adhesion, exhibiting potential to become antitumor drug.« less

Regulation of gene expression in intervertebral disc cells by low and high hydrostatic pressure.

PubMed

Neidlinger-Wilke, Cornelia; Würtz, Karin; Urban, Jill P G; Börm, Wolfgang; Arand, Markus; Ignatius, Anita; Wilke, Hans-Joachim; Claes, Lutz E

2006-08-01

Intervertebral disc structures are exposed to wide ranges of intradiscal hydrostatic pressure during different loading exercises and are at their minimum during lying or relaxed sitting and at maximum during lifting weights with a round back. We hypothesize that these different loading magnitudes influence the intervertebral disc (IVD) by alteration of disc matrix turnover depending on their magnitudes. Therefore the aim of this study was to assess changes in gene expression of human nucleus cells after the application of low hydrostatic pressure (0.25 MPa) and high hydrostatic pressure (2.5 MPa). IVD cells isolated from the nucleus of human (n = 18) and bovine (n = 24 from four animals) disc biopsies were seeded into three-dimensional collagen type-I matrices and exposed to the different loading magnitudes by specially developed pressure chambers. The lower pressure range (0.25 MPa, 30 min, 0.1 Hz) was applied with a recently published device by using an external compression cylinder. For the application of higher loads (2.5 MPa, 30 min, 0.1 Hz) the cell-loaded collagen gels were sealed into sterile bags with culture medium and stimulated in a newly developed water-filled compression cylinder by using a loading frame. These methods allowed the comparison of loading regimes in a wide physiological range under an equal three-dimensional culture conditions. Cells were harvested 24 h after the end of stimulation and changes in the expression of genes known to influence IVD matrix turnover (collagen-I, collagen-II, aggrecan, MMP1, MMP2, MMP3, MMP13) were analyzed by real-time RT-PCR. A Wilcoxon signed-rank test(1) and a Wilcoxon 2-sample test(2) were performed to detect differences between the stimulated and control samples(1) and differences between low and high hydrostatic pressure(2). Multiple testing was considered by adjusting the p value appropriately. Both regimes of hydrostatic pressure influenced gene expression in nucleus cells with opposite tendencies for the matrix forming proteins aggrecan and collagen type-I in response to the two different pressure magnitudes: Low hydrostatic-pressure (0.25 MPa) tended to increase collagen-I and aggrecan expression of human nucleus cells (P < 0.05) but only to a small degree. High hydrostatic pressure (2.5 MPa) tended to decrease gene expression of all anabolic proteins with significant effects on aggrecan expression of nucleus cells (P = 0.004). Low hydrostatic pressure had no influence on the expression of matrix metalloproteinases (MMP1, MMP2, MMP3 and MMP13). In contrast, high hydrostatic pressure tended to increase the expression of MMP1, MMP3 and MMP13 of human nucleus cells with high individual-individual variations. The decreased expression of aggrecan (P = 0.008) and collagen type II (P = 0.023) and the increased MMP3 expression (P = 0.008) in response to high hydrostatic pressure could be confirmed in additional experiments with bovine nucleus cells. These results suggest that hydrostatic pressure as one of the physiological stimuli of the IVD may influence matrix turnover in a magnitude dependent way. Low hydrostatic pressure (0.25 MPa) has quite small influences with a tendency to anabolic effects, whereas high hydrostatic pressure (2.5 MPa) tends to decrease the matrix protein expression with a tendency to increase some matrix-turnover enzymes. Therefore, hydrostatic pressure may regulate disc matrix turnover in a dose-dependent way.

Src promotes cutaneous wound healing by regulating MMP-2 through the ERK pathway.

PubMed

Wu, Xue; Yang, Longlong; Zheng, Zhao; Li, Zhenzhen; Shi, Jihong; Li, Yan; Han, Shichao; Gao, Jianxin; Tang, Chaowu; Su, Linlin; Hu, Dahai

2016-03-01

Wound healing is a highly orchestrated, multistep process, and delayed wound healing is a significant symptomatic clinical problem. Keratinocyte migration and re-epithelialization play the most important roles in wound healing, as they determine the rate of wound healing. In our previous study, we found that Src, one of the oldest proto‑oncogenes encoding a membrane-associated, non-receptor protein tyrosine kinase, promotes keratinocyte migration. We therefore hypothesized that Src promotes wound healing through enhanced keratinocyte migration. In order to test this hypothesis, vectors for overexpressing Src and small interfering RNAs (siRNAs) for silencing of Src were used in the present study. We found that the overexpression of Src accelerated keratinocyte migration in vitro and promoted wound healing in vivo without exerting a marked effect on cell proliferation. The extracellular signal-regulated kinase (ERK) and c-Jun N-terminal kinase (JNK) signaling pathways play important roles in Src-accelerated keratinocyte migration. Further experiments demonstrated that Src induced the protein expression of matrix metalloproteinase-2 (MMP-2) and decreased the protein expression of E-cadherin. We suggest that ERK signaling is involved in the Src-mediated regulation of MMP-2 expression. The present study provided evidence that Src promotes keratinocyte migration and cutaneous wound healing, in which the regulation of MMP-2 through the ERK pathway plays an important role, and thus we also demonstrated a potential therapeutic role for Src in cutaneous wound healing.

Heme Oxygenase-1 Regulation of Matrix Metalloproteinase-1 Expression Underlies Distinct Disease Profiles in Tuberculosis

PubMed Central

Andrade, Bruno B.; Kumar, Nathella Pavan; Amaral, Eduardo P.; Riteau, Nicolas; Mayer-Barber, Katrin D.; Tosh, Kevin W.; Maier, Nolan; Conceição, Elisabete L.; Kubler, Andre; Sridhar, Rathinam; Banurekha, Vaithilingam V.; Jawahar, Mohideen S.; Barbosa, Theolis; Manganiello, Vincent C.; Moss, Joel; Fontana, Joseph R.; Marciano, Beatriz E.; Sampaio, Elizabeth P.; Olivier, Kenneth N.; Holland, Steven M.; Jackson, Sharon H.; Moayeri, Mahtab; Leppla, Stephen; Sereti, Irini; Barber, Daniel L.; Nutman, Thomas B.; Babu, Subash; Sher, Alan

2015-01-01

Pulmonary tuberculosis (TB) is characterized by oxidative stress and lung tissue destruction by matrix metalloproteinases (MMP). The interplay between these distinct pathological processes and the implications for TB diagnosis and disease staging are poorly understood. Heme oxygenase-1 (HO-1) levels have been shown to distinguish active from latent as well as successfully treated Mycobacterium tuberculosis (Mtb) infection. MMP-1 expression is also associated with active TB. Here, we measured plasma levels of these two important biomarkers in distinct TB cohorts from India and Brazil. Patients with active TB expressed either very high levels of HO-1 and low levels of MMP-1 or the converse. Moreover, TB patients with either high HO-1 or MMP-1 levels displayed distinct clinical presentations as well as plasma inflammatory marker profiles. In contrast, in an exploratory North American study, inversely correlated expression of HO-1 and MMP-1 was not observed in patients with other non-tuberculous lung diseases. To assess possible regulatory interactions in the biosynthesis of these two enzymes at the cellular level, we studied expression of HO-1 and MMP-1 in Mtb-infected human and murine macrophages. We found that infection of macrophages with live virulent Mtb is required for robust induction of high levels of HO-1, but not MMP-1. In addition, we observed that carbon monoxide, a product of Mtb induced HO-1 activity, inhibits MMP-1 expression by suppressing c-Jun/AP-1 activation. These findings reveal a mechanistic link between oxidative stress and tissue remodeling that may find applicability in the clinical staging of TB patients. PMID:26268658

Pinus densiflora extract protects human skin fibroblasts against UVB-induced photoaging by inhibiting the expression of MMPs and increasing type I procollagen expression.

PubMed

Jung, Hoe-Yune; Shin, Jae-Cheon; Park, Seon-Min; Kim, Na-Ri; Kwak, Wonjung; Choi, Bo-Hwa

2014-01-01

Exposure to ultraviolet (UV) light can cause skin photoaging, which is associated with upregulation of matrix metalloproteinases (MMPs) and downregulation of collagen synthesis. It has been reported that MMPs, especially MMP-1, MMP-3 and MMP-9, decrease the elasticity of the dermis by degrading collagen. In this study, we assessed the effects of Pinus densiflora extract (PDE) on photoaging and investigated its mechanism of action in human skin fibroblast (Hs68) cells after UVB exposure using real-time polymerase chain reaction, Western blot analysis, and enzymatic activity assays. PDE exhibited an antioxidant activity and inhibited elastase activities in vitro. We also found that PDE inhibited UVB-induced cytotoxicity, MMP-1 production and expression of MMP-1, -3 and -9 mRNA in Hs68 cells. In addition, PDE decreased UVB-induced MMP-2 activity and MMP-2 mRNA expression. Moreover, PDE prevented the decrease of type I procollagen mediated by exposure to UVB irradiation, an effect that is linked to the upregulation and downregulation of Smad3 and Smad7, respectively. Another effect of UV irradiation is to stimulate activator protein 1 (AP-1) activity via overexpression of c-Jun/c-Fos, which, in turn, upregulates MMP-1, -3, and -9. In this study, we found that PDE suppressed UV-induced c-Jun and c-Fos mRNA expression. Taken together, these results demonstrate that PDE regulates UVB-induced expression of MMPs and type I procollagen synthesis by inhibiting AP-1 activity and restoring impaired Smad signaling, suggesting that PDE may be useful as an effective anti-photoaging agent.

Chronic exposure of interleukin-13 suppress the induction of matrix metalloproteinase-1 by tumour necrosis factor α in normal and scleroderma dermal fibroblasts through protein kinase B/Akt.

PubMed

Brown Lobbins, M L; Shivakumar, B R; Postlethwaite, A E; Hasty, K A

2018-01-01

Peripheral blood mononuclear cells taken from patients with scleroderma express increased levels of interleukin (IL)-13. Moreover, the expression of matrix metalloproteinase-1 (MMP-1) from involved scleroderma skin fibroblasts is refractory to stimulation by tumour necrosis factor (TNF)-α. To elucidate the mechanism(s) involved, we examined the effect of IL-13 on TNF-α-induced MMP-1 expression in normal and scleroderma human dermal fibroblast lines and studied the involvement of serine/threonine kinase B/protein kinase B (Akt) in this response. Dermal fibroblast lines were stimulated with TNF-α in the presence of varying concentrations of IL-13. Total Akt and pAkt were quantitated using Western blot analyses. Fibroblasts were treated with or without Akt inhibitor VIII in the presence of IL-13 followed by TNF-α stimulation. MMP-1 expression was analysed by real-time polymerase chain reaction (PCR) and enzyme-linked immunosorbent assay (ELISA). Statistical analysis was performed using analysis of variance (anova) or Student's t-test. Upon TNF-α stimulation, normal dermal fibroblasts secrete more MMP-1 than systemic sclerosis (SSc) fibroblasts. This increase in MMP-1 is lost when fibroblasts are co-incubated with IL-13 and TNF-α. IL-13 induced a significant increase in levels of pAkt in dermal fibroblasts, while Akt inhibitor VIII reversed the suppressive effects of IL-13 on the response of cultured fibroblasts to TNF-α, increasing their expression of MMP-1. We show that IL-13 suppresses MMP-1 in TNF-α-stimulated normal and scleroderma dermal fibroblast. Akt inhibitor VIII is able to reverse the suppressive effect of IL-13 on MMP-1 expression and protein synthesis. Our data suggest that IL-13 regulates MMP-1 expression in response to TNF-α through an Akt-mediated pathway and may play a role in fibrotic diseases such as scleroderma. © 2017 British Society for Immunology.

Cyclophilin A (CypA) is associated with the inflammatory infiltration and alveolar bone destruction in an experimental periodontitis

DOE Office of Scientific and Technical Information (OSTI.GOV)

Liu, Lihua; Li, Chengzhang, E-mail: l56cz@hotmail.com; Department of Periodontology, School and Hospital of Stomatology, Wuhan University, 237 Luo Yu Road, Hongshan District, Wuhan 430079

2010-01-01

Background and objective: CypA is able to regulate inflammatory responses and MMPs production via interaction with its cell surface receptor, EMMPRIN. This study aimed to address the possible association of CypA with pathological inflammation and destruction of periodontal tissues, and whether CypA-EMMPRIN interaction exists in periodontitis. Materials and methods: Experimental periodontitis was induced by ligation according to our previous method. Histological and radiographic examinations were performed. Western blot was used to detect CypA and EMMPRIN expressions in gingival tissues. Immunohistochemistry was applied for CypA, EMMPRIN, MMP-1, MMP-2, MMP-9, as well as cell markers of macrophage, lymphocyte and neutrophil. CypA expression,more » alveolar bone loss, and inflammatory infiltrations were quantified followed by correlation analyses. Results: Western blot revealed that CypA and EMMRPIN expressions were dramatically elevated in inflamed gingival tissues (ligature group) as compared to healthy gingival tissues (control group). The enhanced CypA and EMMPRIN expressions were highly consistent in cell localization on seriate sections. They were permanently co-localized in infiltrating macrophages and lymphocytes, as well as osteoclasts and osteoblasts in interradicular bone, but rarely expressed by infiltrating neutrophils. MMP-1, MMP-2, and MMP-9 expressions were also sharply increased in inflamed gingiva. MMP-2 and MMP-9 were mainly over-expressed by macrophages, while MMP-1 was over-produced by fibroblasts and infiltrating cells. The number of CypA-positive cells was strongly correlated with the ACJ-AC distance (r = 0.839, p = 0.000), the number of macrophages (r = 0.972, p = 0.000), and the number of lymphocytes (r = 0.951, p = 0.000). Conclusion: CypA is associated with the inflammatory infiltration and alveolar bone destruction of periodontitis. CypA-EMMPRIN interaction may exist in these pathological processes.« less

PTP1B promotes aggressiveness of breast cancer cells by regulating PTEN but not EMT.

PubMed

Liu, Xue; Chen, Qian; Hu, Xu-Gang; Zhang, Xian-Chao; Fu, Ti-Wei; Liu, Qing; Liang, Yan; Zhao, Xi-Long; Zhang, Xia; Ping, Yi-Fang; Bian, Xiu-Wu

2016-10-01

Metastasis is a complicated, multistep process and remains the major cause of cancer-related mortality. Exploring the molecular mechanisms underlying tumor metastasis is crucial for development of new strategies for cancer prevention and treatment. In this study, we found that protein tyrosine phosphatase 1B (PTP1B) promoted breast cancer metastasis by regulating phosphatase and tensin homolog (PTEN) but not epithelial-mesenchymal transition (EMT). By detecting PTP1B expression of the specimens from 128 breast cancer cases, we found that the level of PTP1B was higher in breast cancer tissues than the corresponding adjacent normal tissues. Notably, PTP1B was positively associated with lymph node metastasis (LNM) and estrogen receptor (ER) status. In vitro, disturbing PTP1B expression obviously attenuated cell migration and invasion. On the contrary, PTP1B overexpression significantly increased migration and invasion of breast cancer cells. Mechanistically, PTP1B knockdown upregulated PTEN, accompanied with an abatement of AKT phosphorylation and the expression of matrix metalloproteinase 2 (MMP2) and MMP7. Conversely, forced expression of PTP1B reduced PTEN and increased AKT phosphorylation as well as the expression of MMP2 and MMP7. Notably, neither EMT nor stemness of breast cancer cells was regulated by PTP1B. We also found that PTP1B acted as an independent prognostic factor and predicted poor prognosis in ER-positive breast cancer patients. Taken together, our findings provide advantageous evidence for the development of PTP1B as a potential therapeutic target for breast cancer, especially for ER-positive breast cancer patients.

Functional self-assembled peptide scaffold inhibits tumor necrosis factor-alpha-induced inflammation and apoptosis in nucleus pulposus cells by suppressing nuclear factor-κB signaling.

PubMed

Li, Xiaochuan; Cheng, Shi; Wu, Yaohong; Ying, Jingwei; Wang, Chaofeng; Wen, Tianyong; Bai, Xuedong; Ji, Wei; Wang, Deli; Ruan, Dike

2018-04-01

Although nucleus pulposus (NP) tissue engineering has achieved tremendous success, researches still face the huge obstacles in maintaining cell survival and function. A novel functional self-assembled peptide RADA-KPSS was constructed by conjugating BMP-7 short active fragment (KPSS) to the C-terminus of RADA16-I that displays anti-inflammatory and anti-apoptosis effects. However, whether this functional self-assembled RADA-KPSS peptide can alleviate inflammation and NPC apoptosis induced by tumor necrosis factor-alpha (TNF-α) has not been studied. Therefore, we cultured NPCs treated with TNF-α for 48 h with the RADA-KPSS peptide, and compared the results to those with RADA16-I peptide. The cell apoptosis rate, inflammatory mediator secretion, expression of matrix-degrading enzymes, and extracellular matrix (ECM) protein levels were evaluated. The expression of nuclear factor-κB-p65 (NF-κB-p65) protein was also tested. TNF-α-treated NPCs cultured with the RADA16-I peptide showed up-regulated gene expression for matrix-degrading enzymes, such as matrix metalloproteinases-3 (MMP-3), MMP-9, and a disintegrin and metalloproteinase with thrombospondin motifs (ADAMTS-4), and down-regulated gene expression for ECM proteins such as aggrecan, collagen II, and Sox-9. The RADA-KPSS peptide could attenuate the expression of MMP-3, MMP-9, and ADAMTS-4, promote accumulation of ECM proteins, and increase secretion of glycosaminoglycan as compared with the RADA16-I peptide. Moreover, the TNF-α-damaged NPCs was further demonstrated to inhibit NF-κB-p65, IL-1, IL-6, and prostaglandin E-2 proteins and decrease cell apoptosis in RADA-KPSS peptide. In conclusion, the functional self-assembled RADA-KPSS peptides have anti-inflammatory and anti-apoptotic effects by promoting anabolic processes and inhibiting catabolic processes in intervertebral disk degeneration. These peptides may be feasible for clinical applications in NP tissue engineering. © 2017 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 106A: 1082-1091, 2018. © 2017 Wiley Periodicals, Inc.

Involvement of matrix metalloproteinases in chronic Q fever.

PubMed

Jansen, A F M; Schoffelen, T; Textoris, J; Mege, J L; Bleeker-Rovers, C P; Roest, H I J; Wever, P C; Joosten, L A B; Netea, M G; van de Vosse, E; van Deuren, M

2017-07-01

Chronic Q fever is a persistent infection with the intracellular Gram-negative bacterium Coxiella burnetii, which can lead to complications of infected aneurysms. Matrix metalloproteinases (MMPs) cleave extracellular matrix and are involved in infections as well as aneurysms. We aimed to study the role of MMPs in the pathogenesis of chronic Q fever. We investigated gene expression of MMPs through microarray analysis and MMP production with ELISA in C. burnetii-stimulated peripheral blood mononuclear cells (PBMCs) of patients with chronic Q fever and healthy controls. Twenty single nucleotide polymorphisms (SNPs) of MMP and tissue inhibitor of MMP genes were genotyped in 139 patients with chronic Q fever and 220 controls with similar cardiovascular co-morbidity. Additionally, circulating MMPs levels in patients with chronic Q fever were compared with those in cardiovascular controls with and without a history of past Q fever. In healthy controls, the MMP pathway involving four genes (MMP1, MMP7, MMP10, MMP19) was significantly up-regulated in C. burnetii-stimulated but not in Escherichia coli lipopolysaccharide -stimulated PBMCs. Coxiella burnetii induced MMP-1 and MMP-9 production in PBMCs of healthy individuals (both p<0.001), individuals with past Q fever (p<0.05, p<0.01, respectively) and of patients with chronic Q fever (both p<0.001). SNPs in MMP7 (rs11568810) (p<0.05) and MMP9 (rs17576) (p<0.05) were more common in patients with chronic Q fever. Circulating MMP-7 serum levels were higher in patients with chronic Q fever (median 33.5 ng/mL, interquartile range 22.3-45.7 ng/mL) than controls (20.6 ng/mL, 15.9-33.8 ng/mL). Coxiella burnetii-induced MMP production may contribute to the development of chronic Q fever. Copyright © 2017 European Society of Clinical Microbiology and Infectious Diseases. Published by Elsevier Ltd. All rights reserved.

VANGL2 interacts with integrin αv to regulate matrix metalloproteinase activity and cell adhesion to the extracellular matrix.

PubMed

Jessen, Tammy N; Jessen, Jason R

2017-12-15

Planar cell polarity (PCP) proteins are implicated in a variety of morphogenetic processes including embryonic cell migration and potentially cancer progression. During zebrafish gastrulation, the transmembrane protein Vang-like 2 (VANGL2) is required for PCP and directed cell migration. These cell behaviors occur in the context of a fibrillar extracellular matrix (ECM). While it is thought that interactions with the ECM regulate cell migration, it is unclear how PCP proteins such as VANGL2 influence these events. Using an in vitro cell culture model system, we previously showed that human VANGL2 negatively regulates membrane type-1 matrix metalloproteinase (MMP14) and activation of secreted matrix metalloproteinase 2 (MMP2). Here, we investigated the functional relationship between VANGL2, integrin αvβ3, and MMP2 activation. We provide evidence that VANGL2 regulates cell surface integrin αvβ3 expression and adhesion to fibronectin, laminin, and vitronectin. Inhibition of MMP14/MMP2 activity suppressed the cell adhesion defect in VANGL2 knockdown cells. Furthermore, our data show that MMP14 and integrin αv are required for increased proteolysis by VANGL2 knockdown cells. Lastly, we have identified integrin αvβ3 as a novel VANGL2 binding partner. Together, these findings begin to dissect the molecular underpinnings of how VANGL2 regulates MMP activity and cell adhesion to the ECM. Copyright © 2017 The Authors. Published by Elsevier Inc. All rights reserved.

E6/E7 oncoproteins of high risk HPV-16 upregulate MT1-MMP, MMP-2 and MMP-9 and promote the migration of cervical cancer cells.

PubMed

Zhu, Dingjun; Ye, Mei; Zhang, Wei

2015-01-01

E6 and E7 of high risk human papillomavirus 16 (HPV16) were reported to correlate with the cervical cancer (CC). And the presence of matrix metalloproteinases (MMPs) has also been indicated to be associated with CC. The present study investigated the expression of MMPs (MT1-MMP, MMP-2 and MMP-9) in CC cells with HPV16-E6/E7 oncoprotein(s) negative or positive, and then determined the regulation of HPV16-E6/E7 oncoproteins on the expression of MMPs (MT1-MMP, MMP-2 and MMP-9) and the migration of cervical cancer Caski and SiHa cells with RNAi technology. It was demonstrated that the overexpression or the knockdown of HPV16-E6/E7 promoted or reduced MT1-MMP, MMP-2 and MMP-9 in CC cells. And the HPV16-E6, -E7 or -E6E7 influenced the migration of CC cells. The overexpression or the knockdown of them promoted or inhibited the migration of C33A or Caski/SiHa cells. Moreover, the chemical inhibition of MMP-2 or MMP-9 significantly reduced the migration of CC Caski or SiHa cells. Our results demonstrated that the E6-HPV16 or E7-HPV16 promoted the activity of MMP-2/9, and contributed to the migration of cervical cells.

Caspase inhibition augmented oridonin-induced cell death in murine fibrosarcoma l929 by enhancing reactive oxygen species generation.

PubMed

Wu, Jin-Nan; Huang, Jian; Yang, Jia; Tashiro, Shin-Ichi; Onodera, Satoshi; Ikejima, Takashi

2008-09-01

Oridonin, a diterpenoid isolated from Rabdosia rubescences, has been reported to have antitumor effects. In this study, the growth-inhibitory activity of oridonin for L929 cells was exerted in a time-and dose-dependent manner. After treatment with oridonin for 24 h, L929 cells underwent both apoptosis and necrosis as measured by an lactate dehydrogenase (LDH) activity-based assay. A rapid generation of reactive oxygen species (ROS) was triggered by oridonin, and subsequently up-regulation of phospho-p53 (ser 15) expression and an increased expression ratio of Bax/Bcl-2 was observed. Furthermore, there was a significant fall in mitochondrial membrane potential (MMP) and increase in caspase-3 activity after exposure to oridonin for 24 h. Surprisingly, the pan-caspase inhibitor z-VAD-fmk and caspase3 inhibitor z-DEVD-fmk rendered L929 cells more sensitive to oridonin, rather than preventing oridonin-induced cell death. Oridonin and z-VAD-fmk co-treatment not only resulted in an even higher ROS production, but also made a more significant reduction in the MMP. Pretreatment of ROS scavenger N-acetylcysteine (NAC) led to a complete inhibition of oridonin-induced cell death, intracellular ROS generation, and MMP collapse. NAC treatment also reversed the potentiation of cell death by the pan-caspase inhibitor z-VAD-fmk. Taken together, these observations showed that oridonin-induced cell death in L929 cells involved intracellular ROS generation, activation of phospho-p53 (ser 15), and up-regulation of the Bax/Bcl-2 ratio; and the augmented cell death by z-VAD-fmk was dependent on an increased ROS production.

In vitro anticancer activities of osthole against renal cell carcinoma cells.

PubMed

Liu, Lei; Mao, Jun; Wang, Qifei; Zhang, Zhiwei; Wu, Guangzhen; Tang, Qizhen; Zhao, Bin; Li, Lianhong; Li, Quanlin

2017-10-01

Renal cell carcinoma (RCC) is a common urinary malignancy that is resistant to chemotherapy and radiotherapy. Osthole, a monomer compound extracted from a traditional Chinese herb, has potent anti-tumor effects on various types of cancer cells. However, the therapeutic effects of osthole on RCC remain unclear. In our study, osthole could suppress the proliferation and colony formation of two RCC cell lines, ACHN and 786-O cells, in a dose-dependent manner. Treatment with osthole resulted in a significant, dose-dependent increase in the expression of pro-apoptotic proteins (cleaved caspase-3 and Bax) and decreased expression of anti-apoptotic proteins (Bcl-2 and survivin), which were consistent with evidence of apoptotic nuclear morphology revealed by DAPI staining. Pre-treatment with osthole attenuated the migratory and invasive abilities of RCC cells in a dose-dependent manner, as evidenced by a reduction in migrating cells in a Transwell assay and a decreased wound closure ratio in a scratch assay as compared with the control. Additionally, osthole down-regulated the expression of migration/invasion-related proteins matrix metalloproteinase (MMP)-2 and MMP-9. Osthole significantly up-regulated epithelial biomarkers (E-cadherin and beta-catenin) and down-regulated mesenchymal biomarkers (N-cadherin and vimentin). Furthermore, our results suggest that osthole suppressed the expression of epithelial-mesenchymal transition transcriptional factors Smad-3, Snail-1, and Twist-1. Taken together, the results of this study suggest that osthole might be a potential novel herbal agent against RCC. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

Peripheral blood gene expression profile of atherosclerotic coronary artery disease in patients of different ethnicity in Malaysia.

PubMed

Abdullah, Mohd Hafiz Ngoo; Othman, Zulhabri; Noor, Hamdan Mohd; Arshad, Siti Suri; Yusof, Ahmad Khairuddin Mohd; Jamal, Rahman; Rahman, Abdul Rashid Abdul

2012-09-01

The molecular basis of coronary artery disease (CAD) has been widely studied in the western world but there is no published work on the Malaysian population. This study looked at the global gene expression profiling of the peripheral blood of patients with CAD from the 3 main ethnic groups in Malaysia. Male subjects selected were based on angiographically confirmed CAD (≥50% stenosis) and normal control subjects (0% stenosis) with age range of 55.6±5.3 and 51.0±5.5 years, respectively. The global gene expression of 12 angiographically documented CAD patients and 11 matched control subjects were performed. The combined group samples identified 6 up regulated differential expression (DE) genes (GHRL, LTA, CBS, HP, ITGA2B, and OLR1) and 12 down regulated DE genes (IL18R1, ITGA2B, IL18RAP, HP, OLR1, SOD2 ITGB3, IL1B, MMP9, PLA2G7, UTS2, and CBS) to be involved in CAD at the fold change of 1.3 with fault discovery rate (FDR) of 1%. Three genes, MMP9, IL1B, and SOD2 were down regulated in all the 3 ethnic groups making them potential biomarker candidates for CAD across all three ethnicities. Further verification in a cohort study is needed. Copyright © 2012 Japanese College of Cardiology. Published by Elsevier Ltd. All rights reserved.

Ang-(1-7) exerts protective role in blood-brain barrier damage by the balance of TIMP-1/MMP-9.

PubMed

Wu, Jitao; Zhao, Duo; Wu, Shuang; Wang, Dan

2015-02-05

Cerebrovascular disease (CVD) ranks as the top three health risks, specially cerebral ischemia characterized with the damage of blood-brain barrier (BBB). The angiotensin Ang-(1-7) was proven to have a protective effect on cerebrovascular diseases. However, its role on blood-brain barrier and the underlying molecular mechanism remains unclear. In this study, Ang-(1-7) significantly relieved damage of ischemia reperfusion injury on blood-brain barrier in cerebral ischemia reperfusion injury (IRI) rats. Furthermore, its treatment attenuated BBB permeability and brain edema. Similarly, Ang-(1-7) also decreased the barrier permeability of brain endothelial cell line RBE4. Further analysis showed that Ang-(1-7) could effectively restore tight junction protein (claudin-5 and zonula occludens ZO-1) expression levels both in IRI-rats and hypoxia-induced RBE4 cells. Furthermore, Ang-(1-7) stimulation down-regulated hypoxia-induced matrix metalloproteinase-9 (MMP-9) levels, whose silencing with (matrix metalloproteinase-9 hemopexin domain) MMP9-PEX inhibitor significantly increased the expression of claudin-5 and ZO-1. Further mechanism analysis demonstrated that Ang-(1-7) might junction protein levels by tissue inhibitor of metalloproteinase 1 (TIMP1)-MMP9 pathway, because Ang-(1-7) enhanced TIMP1 expression, whose silencing obviously attenuated the inhibitor effect of Ang-(1-7) on MMP-9 levels and decreased Ang-(1-7)-triggered increase in claudin-5 and ZO-1. Together, this study demonstrated a protective role of Ang-(1-7) in IRI-induced blood-brain barrier damage by TIMP1-MMP9-regulated tight junction protein expression. Accordingly, Ang-(1-7) may become a promising therapeutic agent against IRI and its complications. Copyright © 2014 Elsevier B.V. All rights reserved.

Chemopreventive effects of a curcumin-like diarylpentanoid [2,6-bis(2,5-dimethoxybenzylidene)cyclohexanone] in cellular targets of rheumatoid arthritis in vitro.

PubMed

Lee, Ka-Heng; Abas, Faridah; Mohamed Alitheen, Noorjahan Banu; Shaari, Khozirah; Lajis, Nordin Haji; Israf, Daud Ahmad; Syahida, Ahmad

2015-07-01

Synovial fibroblast has emerged as a potential cellular target in progressive joint destruction in rheumatoid arthritis development. In this study, BDMC33 (2,6-bis[2,5-dimethoxybenzylidene]cyclohexanone), a curcumin analogue with enhanced anti-inflammatory activity has been synthesized and the potency of BDMC33 on molecular and cellular basis of synovial fibroblasts (SF) were evaluated in vitro. Synovial fibroblast cells (HIG-82) were cultured in vitro and induced by phorbol-12-myristate acetate (PMA) to stimulate the expression of matrix metalloproteinase (MMPs) and pro-inflammatory cytokines. The protective effects of BDMC33 were evaluated toward MMP activities, pro-inflammatory cytokine expression and nuclear factor kappa-B (NF-κB) activation by using various bioassay methods, including zymography, Western blotting, reverse transcription polymerase chain reaction, immunofluorescense microscopy and electrophoretic mobility shift assay. The results showed that BDMC33 significantly inhibited the pro-gelatinase B (pro-MMP-9) and collagenase activities via suppression of MMP-1 in activated SF. In addition, BDMC33 strongly suppressed MMP-3 gene expression as well as inhibited COX-2 and IL-6 pro-inflammatory gene expression. We also demonstrated that BDMC33 abolished the p65 NF-κB nuclear translocation and NF-κB DNA binding activity in PMA-stimulated SF. BDMC33 represents an effective chemopreventive agent and could be used as a promising lead compound for further development of rheumatoid arthritis therapeutic intervention. © 2014 Asia Pacific League of Associations for Rheumatology and Wiley Publishing Asia Pty Ltd.

Matrix Metalloproteinase-2 Activity is Associated with Divergent Regulation of Calponin-1 in Conductance and Resistance Arteries in Hypertension-induced Early Vascular Dysfunction and Remodelling.

PubMed

Parente, Juliana M; Pereira, Camila A; Oliveira-Paula, Gustavo H; Tanus-Santos, José E; Tostes, Rita C; Castro, Michele M

2017-10-01

Matrix metalloproteinase (MMP)-2 participates in hypertension-induced maladaptive vascular remodelling by degrading extra- and intracellular proteins. The consequent extracellular matrix rearrangement and phenotype switch of vascular smooth muscle cells (VSMCs) lead to increased cellular migration and proliferation. As calponin-1 degradation by MMP-2 may lead to VSMC proliferation during hypertension, the hypothesis of this study is that increased MMP-2 activity contributes to early hypertension-induced maladaptive remodelling in conductance and resistance arteries via regulation of calponin-1. The main objective was to analyse whether MMP-2 exerts similar effects on the structure and function of the resistance and conductance arteries during early hypertension. Two-kidney, one-clip (2K-1C) hypertensive male rats and corresponding controls were treated with doxycycline (30 mg/kg/day) or water until reaching one week of hypertension. Systolic blood pressure was increased in 2K-1C rats, and doxycycline did not reduce it. Aortas and mesenteric arteries were analysed. MMP-2 activity and expression were increased in both arteries, and doxycycline reduced it. Significant hypertrophic remodelling and VSMC proliferation were observed in aortas but not in mesenteric arteries of 2K-1C rats. The contractility of mesenteric arteries to phenylephrine was increased in 2K-1C rats, and doxycycline prevented this alteration. The potency of phenylephrine to contract aortas of 2K-1C rats was increased, and doxycycline decreased it. Whereas calponin-1 expression was increased in 2K-1C mesenteric arteries, calponin-1 was reduced in aortas. Doxycycline treatment reverted changes in calponin-1 expression. MMP-2 contributes to hypertrophic remodelling in aortas by decreasing calponin-1 levels, which may result in VSMC proliferation. On the other hand, MMP-2-dependent increased calponin-1 in mesenteric arteries may contribute to vascular hypercontractility in 2K-1C rats. Divergent regulation of calponin-1 by MMP-2 may be an important mechanism that leads to maladaptive vascular effects in hypertension. © 2017 Nordic Association for the Publication of BCPT (former Nordic Pharmacological Society).

Human neuronal uncoupling proteins 4 and 5 (UCP4 and UCP5): structural properties, regulation, and physiological role in protection against oxidative stress and mitochondrial dysfunction.

PubMed

Ramsden, David B; Ho, Philip W-L; Ho, Jessica W-M; Liu, Hui-Fang; So, Danny H-F; Tse, Ho-Man; Chan, Koon-Ho; Ho, Shu-Leong

2012-07-01

Uncoupling proteins (UCPs) belong to a large family of mitochondrial solute carriers 25 (SLC25s) localized at the inner mitochondrial membrane. UCPs transport protons directly from the intermembrane space to the matrix. Of five structural homologues (UCP1 to 5), UCP4 and 5 are principally expressed in the central nervous system (CNS). Neurons derived their energy in the form of ATP that is generated through oxidative phosphorylation carried out by five multiprotein complexes (Complexes I-V) embedded in the inner mitochondrial membrane. In oxidative phosphorylation, the flow of electrons generated by the oxidation of substrates through the electron transport chain to molecular oxygen at Complex IV leads to the transport of protons from the matrix to the intermembrane space by Complex I, III, and IV. This movement of protons to the intermembrane space generates a proton gradient (mitochondrial membrane potential; MMP) across the inner membrane. Complex V (ATP synthase) uses this MMP to drive the conversion of ADP to ATP. Some electrons escape to oxygen-forming harmful reactive oxygen species (ROS). Proton leakage back to the matrix which bypasses Complex V resulting in a major reduction in ROS formation while having a minimal effect on MMP and hence, ATP synthesis; a process termed "mild uncoupling." UCPs act to promote this proton leakage as means to prevent excessive build up of MMP and ROS formation. In this review, we discuss the structure and function of mitochondrial UCPs 4 and 5 and factors influencing their expression. Hypotheses concerning the evolution of the two proteins are examined. The protective mechanisms of the two proteins against neurotoxins and their possible role in regulating intracellular calcium movement, particularly with regard to the pathogenesis of Parkinson's disease are discussed.

Silibinin downregulates MMP2 expression via Jak2/STAT3 pathway and inhibits the migration and invasive potential in MDA-MB-231 cells.

PubMed

Byun, Hyo Joo; Darvin, Pramod; Kang, Dong Young; Sp, Nipin; Joung, Youn Hee; Park, Jong Hwan; Kim, Sun Jin; Yang, Young Mok

2017-06-01

Worldwide, breast cancer (BCa) is the most common cancer in women. Among its subtypes, triple-negative breast cancer (TNBC) is an aggressive form associated with diminished survival. TNBCs are characterized by their absence, or minimal expression, of the estrogen and progesterone receptors, as well as the human epidermal growth factor receptor 2 (i.e. ER-/-, PR-/-, Her2-/Low). Consequently, treatment for this subtype of BCa remains problematic. Silibinin, a derivative of the flavonoid silymarin, is reported to have anticancer activities against hepatic and non-small cell lung cancers. We hypothesized that silibinin might inhibit cell-extracellular matrix interactions via the regulation, expression, and activation of STAT3 in TNBCs, which could directly inhibit metastasis in silibinin-treated BCa cells. Using proliferation assays, we found that exposure to silibinin at a concentration of 200 µM inhibited the proliferation of breast cancer (BCa) cells; this concentration also inhibited phosphorylation of STAT3 and its principal upstream kinase, Jak2. Furthermore, we found that silibinin inhibited the nuclear translocation of STAT3, as well as its binding to the MMP2 gene promoter. The ability of silibinin to inhibit metastasis was further studied using an in vitro invasion assay. The results confirm the role of STAT3 as a critical mediator in the invasive potential of BCa cells, and STAT3 knock-down resulted in inhibition of invasion. The invasion ability of silibinin-treated BCa cells was studied in detail with the expression of MMP2. Prevention of STAT3 activation also resulted in the inhibition of MMP2 expression. Use of a small interfering RNA to knock down STAT3 (siSTAT3) allowed us to confirm the role of STAT3 in regulating MMP2 expression, as well as the mechanism of action of silibinin in inhibiting MMP2. Taken together, we found that silibinin inhibits the Jak2/STAT3/MMP2 signaling pathway, and inhibits the proliferation, migration, and invasion of triple-negative BCa cells.

Molecular pathology of brain matrix metalloproteases, claudin5, and aquaporins in forensic autopsy cases with special regard to methamphetamine intoxication.

PubMed

Wang, Qi; Ishikawa, Takaki; Michiue, Tomomi; Zhu, Bao-Li; Guan, Da-Wei; Maeda, Hitoshi

2014-05-01

Methamphetamine (METH) is a highly addictive drug of abuse and toxic to the brain. Recent studies indicated that besides direct damage to dopamine and 5-HT terminals, neurotoxicity of METH may also result from its ability to modify the structure of blood-brain barrier (BBB). The present study investigated the postmortem brain mRNA and immunohistochemical expressions of matrix metalloproteases (MMPs), claudin5 (CLDN5), and aquaporins (AQPs) in forensic autopsy cases of carbon monoxide (n = 14), METH (n = 21), and phenobarbital (n = 17) intoxication, compared with mechanical asphyxia (n = 15), brain injury (n = 11), non-brain injury (n = 21), and sharp instrument injury (n = 15) cases. Relative mRNA quantification using Taqman real-time PCR assay demonstrated higher expression of AQP4 and MMP9, lower expression of CLDN5 in METH intoxication cases and lower expression of MMP2 in phenobarbital intoxication cases. Immunostaining results showed substantial interindividual variations in each group, showing no evident differences in distribution or intensity among all the causes of death. These findings suggest that METH may increase BBB permeability by altering CLDN5 and MMP9, and the self-protective system maybe activated to eliminate accumulating water from the extracellular space of the brain by up-regulating AQP4. Systematic analysis of gene expressions using real-time PCR may be a useful procedure in forensic death investigation.

Ureaplasma isolates stimulate pro-inflammatory CC chemokines and matrix metalloproteinase-9 in neonatal and adult monocytes

PubMed Central

Silwedel, Christine; Fehrholz, Markus; Henrich, Birgit; Waaga-Gasser, Ana Maria; Claus, Heike; Speer, Christian P.

2018-01-01

Being generally regarded as commensal bacteria, the pro-inflammatory capacity of Ureaplasma species has long been debated. Recently, we confirmed Ureaplasma–driven pro-inflammatory cytokine responses and a disturbance of cytokine equilibrium in primary human monocytes in vitro. The present study addressed the expression of CC chemokines and matrix metalloproteinase-9 (MMP-9) in purified term neonatal and adult monocytes stimulated with serovar 8 of Ureaplasma urealyticum (Uu) and serovar 3 of U. parvum (Up). Using qRT-PCR and multi-analyte immunoassay, we assessed mRNA and protein expression of the monocyte chemotactic proteins 1 and 3 (MCP-1/3), the macrophage inflammatory proteins 1α and 1β (MIP-1α/β) as well as MMP-9. For the most part, both isolates stimulated mRNA expression of all given chemokines and MMP-9 in cord blood and adult monocytes (p<0.05 and p<0.01). These results were paralleled by Uu and Up-induced secretion of MCP-1 protein in both cells (neonatal: p<0.01, adult: p<0.05 and p<0.01). Release of MCP-3, MIP-1α, MIP-1β and MMP-9 was enhanced upon exposure to Up (neonatal: p<0.05, p<0.01 and p<0.001, respectively; adult: p<0.05). Co-stimulation of LPS-primed monocytes with Up increased LPS-induced MCP-1 release in neonatal cells (p<0.05) and aggravated LPS-induced MMP-9 mRNA in both cell subsets (neonatal: p<0.05, adult: p<0.01). Our results document considerable expression of pro-inflammatory CC chemokines and MMP-9 in human monocytes in response to Ureaplasma isolates in vitro, adding to our previous data. Findings from co-stimulated cells indicate that Ureaplasma may modulate monocyte immune responses to a second stimulus. PMID:29558521

Biological Mechanisms Underlying the Ultraviolet Radiation-Induced Formation of Skin Wrinkling and Sagging II: Over-Expression of Neprilysin Plays an Essential Role

PubMed Central

Imokawa, Genji; Nakajima, Hiroaki; Ishida, Koichi

2015-01-01

Our previous studies strongly indicated that the up-regulated activity of skin fibroblast-derived elastase plays a pivotal role in wrinkling and/or sagging of the skin via the impairment of elastic fiber configuration and the subsequent loss of skin elasticity. Fortunately, we succeeded in identifying human skin fibroblast-derived elastase as a previously known enzyme, neprilysin or neutral endopeptidase (NEP). We have also characterized epithelial-mesenchymal paracrine cytokine interactions between UVB-exposed-keratinocytes and dermal fibroblasts and found that interleukin-1α and granulocyte macrophage colony stimulatory factor (GM-CSF) are intrinsic cytokines secreted by UVB-exposed keratinocytes that stimulate the expression of neprilysin by fibroblasts. On the other hand, direct UVA exposure of human fibroblasts significantly stimulates the secretion of IL-6 and also elicits a significant increase in the gene expression of matrix metallo-protease(MMP)-1 as well as neprilysin (to a lesser extent), which is followed by distinct increases in their protein and enzymatic activity levels. Direct UVA exposure of human keratinocytes also stimulates the secretion of IL-6, IL-8 and GM-CSF but not of IL-1 and endothelin-1. These findings suggest that GM-CSF secreted by UVA-exposed keratinocytes as well as IL-6 secreted by UVA-exposed dermal fibroblasts play important and additional roles in UVA-induced sagging and wrinkling by up-regulation of neprilysin and MMP-1, respectively, in dermal fibroblasts. PMID:25856676

Identification of key microRNAs and genes in preeclampsia by bioinformatics analysis

PubMed Central

Luo, Shouling; Cao, Nannan; Tang, Yao; Gu, Weirong

2017-01-01

Preeclampsia is a leading cause of perinatal maternal–foetal mortality and morbidity. The aim of this study is to identify the key microRNAs and genes in preeclampsia and uncover their potential functions. We downloaded the miRNA expression profile of GSE84260 and the gene expression profile of GSE73374 from the Gene Expression Omnibus database. Differentially expressed miRNAs and genes were identified and compared to miRNA-target information from MiRWalk 2.0, and a total of 65 differentially expressed miRNAs (DEMIs), including 32 up-regulated miRNAs and 33 down-regulated miRNAs, and 91 differentially expressed genes (DEGs), including 83 up-regulated genes and 8 down-regulated genes, were identified. The pathway enrichment analyses of the DEMIs showed that the up-regulated DEMIs were enriched in the Hippo signalling pathway and MAPK signalling pathway, and the down-regulated DEMIs were enriched in HTLV-I infection and miRNAs in cancers. The gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes pathway (KEGG) enrichment analyses of the DEGs were performed using Multifaceted Analysis Tool for Human Transcriptome. The up-regulated DEGs were enriched in biological processes (BPs), including the response to cAMP, response to hydrogen peroxide and cell-cell adhesion mediated by integrin; no enrichment of down-regulated DEGs was identified. KEGG analysis showed that the up-regulated DEGs were enriched in the Hippo signalling pathway and pathways in cancer. A PPI network of the DEGs was constructed by using Cytoscape software, and FOS, STAT1, MMP14, ITGB1, VCAN, DUSP1, LDHA, MCL1, MET, and ZFP36 were identified as the hub genes. The current study illustrates a characteristic microRNA profile and gene profile in preeclampsia, which may contribute to the interpretation of the progression of preeclampsia and provide novel biomarkers and therapeutic targets for preeclampsia. PMID:28594854

Matrix Metalloproteinase-8 Facilitates Neutrophil Migration through the Corneal Stromal Matrix by Collagen Degradation and Production of the Chemotactic Peptide Pro-Gly-Pro

PubMed Central

Lin, Michelle; Jackson, Patricia; Tester, Angus M.; Diaconu, Eugenia; Overall, Christopher M.; Blalock, J. Edwin; Pearlman, Eric

2008-01-01

Matrix metalloproteinase (MMP)-8 and MMP-9 play several roles in inflammation, including degradation of extracellular matrix (ECM) components and regulation of cytokine activity. To determine the roles of MMP-8 and MMP-9 in a neutrophil-dependent inflammatory response, we used a murine model of corneal inflammation in which LPS is injected into the corneal stroma. In contrast to wild-type mice, we found that i) lipopolysaccharide (LPS)-injected CXCR2−/− corneas had impaired neutrophil infiltration and did not express either MMP-8 or MMP-9; ii) neutrophil migration through the central cornea was impaired in Mmp8−/−, but not Mmp9−/−, mice; iii) neutrophil migration was inhibited in collagenase-resistant mice; iv) the chemotactic Pro-Gly-Pro (PGP) tripeptide that binds CXCR2 was decreased in CXCR2−/− mice; v) PGP production was impaired in Mmp8−/− corneas; and vi) neutralizing anti-PGP antibody did not inhibit neutrophil infiltration in Mmp8−/− mice. We found no effects of MMP-8 on LPS-induced CXC chemokine (LIX, or CXCL5)-induced neutrophil recruitment or on LPS-induced CXC chemokine production. Together, these studies indicate that neutrophils contribute to the production of both MMP-8 and MMP-9 in LPS-injected corneas and that MMP-8 regulates neutrophil migration through the dense collagenous ECM of the corneal stroma by generating chemotactic PGP during inflammation. PMID:18556780

Up-regulation of CCL11 and CCL26 is associated with activated eosinophils in bullous pemphigoid

PubMed Central

Günther, C; Wozel, G; Meurer, M; Pfeiffer, C

2011-01-01

Eosinophils contribute to the pathogenesis of bullous pemphigoid (BP) by secretion of proinflammatory cytokines and proteases. Trafficking of eosinophils into tissue in animal models and asthma depends on interleukin-5 and a family of chemokines named eotaxins, comprising CCL11, CCL24 and CCL26. Up-regulation of CCL11 has been described in BP, but the expression of the other two members of the eotaxin-family, CCL24 and CCL26, has not been investigated. In addition to these chemokines, expression of adhesion molecules associated with eosinophil migration to the skin should be analysed. We demonstrate that similar to CCL11, the concentration of CCL26 was up-regulated in serum and blister fluid of BP patients. In contrast, the concentration of CCL24 was not elevated in sera and blister fluid of the same BP patients. In lesional skin, CCL11 and CCL26 were detected in epidermis and dermis by immunohistochemistry. In contrast to CCL11, CCL26 was expressed strongly by endothelial cells. In line with these findings, eosinophils represented the dominating cell population in BP lesional skin outnumbering other leucocytes. The percentage of eosinophils expressing very late antigen (VLA): VLA-4 (CD49d) and CD11c correlated with their quantity in tissue. Macrophage antigen (MAC)-1 (CD11b/CD18) was expressed constitutively by tissue eosinophils. In conclusion, these data link the up-regulation of the eosinophil chemotactic factor CCL26 in BP to the lesional accumulation of activated eosinophils in the skin. Thereby they broaden the understanding of BP pathogenesis and might indicate new options for therapeutic intervention. PMID:21985360

Whey peptide Isoleucine-Tryptophan inhibits expression and activity of matrix metalloproteinase-2 in rat aorta.

PubMed

Kopaliani, Irakli; Martin, Melanie; Zatschler, Birgit; Müller, Bianca; Deussen, Andreas

2016-08-01

Aortic stiffness is an independent risk factor for development of cardiovascular diseases. Activation of renin-angiotensin-aldosterone system (RAAS) including angiotensin converting enzyme (ACE) activity leads to overproduction of angiotensin II (ANGII) from its precursor angiotensin I (ANGI). ANGII leads to overexpression and activation of matrix metalloproteinase-2 (MMP2), which is critically associated with pathophysiology of aortic stiffness. We previously reported that the whey peptide Isoleucine-Tryptophan (IW) acts as a potent ACE inhibitor. Herein, we critically elucidate the mechanism of action by which IW causes inhibition of expression and activity of MMP2 in aortic tissue. Effects of IW on expression and activity of MMP2 were assessed on endothelial and smooth muscle cells (ECs and SMCs) in vitro and ex vivo (isolated rat aorta). As controls we used the pharmaceutical ACE inhibitor - captopril and the ANGII type 1 receptor blocker - losartan. In vitro, both ANGII and ANGI stimulation significantly (P<0.01) increased expression of MMP2 assessed with western blot. Similarly, to captopril IW significantly (P<0.05) inhibited ANGI, but not ANGII mediated increase in expression of MMP2, while losartan also blocked effects of ANGII. Signaling pathways regulating MMP2 expression in ECs and SMCs were similarly inhibited after treatment with IW or captopril. In ECs IW significantly (P<0.05) inhibited JNK pathway, whereas in SMCs JAK2/STAT3 pathway, assessed with western blot. In vitro findings were fully consistent with results in isolated rat aorta ex vivo. Moreover, IW not only inhibited the MMP2 expression, but also its activation assessed with gelatin zymography. Our findings demonstrate that IW effectively inhibits expression and activation of MMP2 in rat aorta by decreasing local conversion of ANGI to ANGII. Thus, similar to pharmaceutical ACE inhibitor captopril the dipeptide IW may effectively inhibit ACE activity and prevent the age and hypertension associated rise of aortic stiffness. Copyright © 2016 Elsevier Inc. All rights reserved.

Matrix metalloproteinase-1 inhibitory activities of Morinda citrifolia seed extract and its constituents in UVA-irradiated human dermal fibroblasts.

PubMed

Masuda, Megumi; Murata, Kazuya; Naruto, Shunsuke; Uwaya, Akemi; Isami, Fumiyuki; Matsuda, Hideaki

2012-01-01

The objective of this study was to examine whether a 50% ethanolic extract (MCS-ext) of the seeds of Morinda citrifolia (noni) and its constituents have matrix metalloproteinase-1 (MMP-1) inhibitory activity in UVA-irradiated normal human dermal fibroblasts (NHDFs). The MCS-ext (10 μg/mL) inhibited MMP-1 secretion from UVA-irradiated NHDFs, without cytotoxic effects, at 48 h after UV exposure. The ethyl acetate-soluble fraction of MCS-ext was the most potent inhibitor of MMP-1 secretion. Among the constituents of the fraction, a lignan, 3,3'-bisdemethylpinoresinol (1), inhibited the MMP-1 secretion at a concentration of 0.3 μM without cytotoxic effects. Furthermore, 1 (0.3 μM) reduced the level of intracellular MMP-1 expression. Other constituents, namely americanin A (2), quercetin (3) and ursolic acid (4), were inactive. To elucidate inhibition mechanisms of MMP-1 expression and secretion, the effect of 1 on mitogen-activated protein kinases (MAPKs) phosphorylation was examined. Western blot analysis revealed that 1 (0.3 μM) reduced the phosphorylations of p38 and c-Jun-N-terminal kinase (JNK). These results suggested that 1 suppresses intracellular MMP-1 expression, and consequent secretion from UVA-irradiated NHDFs, by down-regulation of MAPKs phosphorylation.

Bone marrow-derived mesenchymal stem cells effectively regenerate fibrotic liver in bile duct ligation rat model

PubMed Central

Elswefy, Sahar E; Rashed, Laila A; Younis, Nahla N; Shaheen, Mohamed A; Ghanim, Amal MH

2016-01-01

Mesenchymal stem cells (MSCs) have attracted lots of attention for the treatment of acute liver failure and end-stage liver diseases. This study aimed at investigating the fundamental mechanism by which bone marrow-derived MSCs (BM-MSCs) induce liver regeneration of fibrotic liver in rats. Rats underwent bile duct ligation (BDL) surgery and four weeks later they were treated with either BM-MSCs (3 × 106 cells /rat, once, tail vein injection) or silymarin (100 mg/kg, daily, orally) for four weeks. Liver function tests and hepatic oxidative stress were determined. Hepatic injury and fibrosis were assessed by H and E, Sirus red staining and immunohistochemical expression of α-smooth muscle actin (α-SMA). Hepatocyte growth factor (HGF) and the gene expression of cytokeratin-19 (CK-19) and matrix metalloproteinase-2 (MMP-2) in liver tissue were determined. BDL induced cholestatic liver injury characterized by elevated ALT and AST activities, bilirubin and decreased albumin. The architecture damage was staged as Metavir score: F3, A3. Fibrosis increased around proliferating bile duct as indicated by sirus red staining and α-SMA immunostaining. Fibrogenesis was favored over fibrolysis and confirmed by decreased HGF with increased expression of CK-19, but decreased MMP-2 expression. BM-MSCs treatment restored deteriorated liver functions and restored the histological changes, resolved fibrosis by improving liver regenerative capabilities (P < 0.001), increases in HGF and MMP-2 mRNA and downregulating CK-19 mRNA. Sliymarin, however, induced similar but less prominent effects compared to BM-MSCs. In conclusion, liver regenerative capabilities can be stimulated by BM-MSCs via augmentation of HGF that subsequently up-regulate MMP-2 mRNA while downregulating CK-19 mRNA. PMID:26811102

Bone marrow-derived mesenchymal stem cells effectively regenerate fibrotic liver in bile duct ligation rat model.

PubMed

Mohamed, Hoda E; Elswefy, Sahar E; Rashed, Laila A; Younis, Nahla N; Shaheen, Mohamed A; Ghanim, Amal M H

2016-03-01

Mesenchymal stem cells (MSCs) have attracted lots of attention for the treatment of acute liver failure and end-stage liver diseases. This study aimed at investigating the fundamental mechanism by which bone marrow-derived MSCs (BM-MSCs) induce liver regeneration of fibrotic liver in rats. Rats underwent bile duct ligation (BDL) surgery and four weeks later they were treated with either BM-MSCs (3 × 10(6) cells /rat, once, tail vein injection) or silymarin (100 mg/kg, daily, orally) for four weeks. Liver function tests and hepatic oxidative stress were determined. Hepatic injury and fibrosis were assessed by H and E, Sirus red staining and immunohistochemical expression of α-smooth muscle actin (α-SMA). Hepatocyte growth factor (HGF) and the gene expression of cytokeratin-19 (CK-19) and matrix metalloproteinase-2 (MMP-2) in liver tissue were determined. BDL induced cholestatic liver injury characterized by elevated ALT and AST activities, bilirubin and decreased albumin. The architecture damage was staged as Metavir score: F3, A3. Fibrosis increased around proliferating bile duct as indicated by sirus red staining and α-SMA immunostaining. Fibrogenesis was favored over fibrolysis and confirmed by decreased HGF with increased expression of CK-19, but decreased MMP-2 expression. BM-MSCs treatment restored deteriorated liver functions and restored the histological changes, resolved fibrosis by improving liver regenerative capabilities (P < 0.001), increases in HGF and MMP-2 mRNA and downregulating CK-19 mRNA. Sliymarin, however, induced similar but less prominent effects compared to BM-MSCs. In conclusion, liver regenerative capabilities can be stimulated by BM-MSCs via augmentation of HGF that subsequently up-regulate MMP-2 mRNA while downregulating CK-19 mRNA. © 2016 by the Society for Experimental Biology and Medicine.

Acetylcholine-induced activation of M3 muscarinic receptors stimulates robust matrix metalloproteinase gene expression in human colon cancer cells.

PubMed

Xie, Guofeng; Cheng, Kunrong; Shant, Jasleen; Raufman, Jean-Pierre

2009-04-01

Previously, we showed that ACh-induced proliferation of human colon cancer cells is mediated by transactivation of epidermal growth factor (EGF) receptors (EGFRs). In the present study, we elucidate the molecular mechanism underlying this action. ACh-induced proliferation of H508 colon cancer cells, which express exclusively M3 muscarinic receptors (M3Rs), was attenuated by anti-EGFR ligand binding domain antibody, a broad-spectrum matrix metalloproteinase (MMP) inhibitor, anti-MMP7 antibody, a diphtheria toxin analog that blocks release of an EGFR ligand [heparin-binding EGF-like growth factor (HBEGF)], and anti-HBEGF antibody. Conditioned media from ACh-treated H508 cells induced proliferation of SNU-C4 colon cancer cells that express EGFR but not M3R. These actions were attenuated by an EGFR inhibitor and by anti-EGFR and anti-HBEGF antibodies. In H508, but not SNU-C4, colon cancer cells, ACh caused a striking dose- and time-dependent increase in levels of MMP7 mRNA and MMP7 protein. Similarly, ACh induced robust MMP1 and MMP10 gene transcription. ACh-induced MMP1, MMP7, and MMP10 gene transcription was attenuated by atropine, anti-EGFR antibody, and chemical inhibitors of EGFR and ERK activation. In contrast, inhibitors of phosphatidylinositol 3-kinase and NF-kappaB activation did not alter MMP gene transcription. Collectively, these findings indicate that MMP7-catalyzed release of HBEGF mediates ACh-induced transactivation of EGFR and consequent proliferation of colon cancer cells. ACh-induced activation of EGFR and downstream ERK signaling also regulates transcriptional activation of MMP7, thereby identifying a novel feed-forward mechanism for neoplastic cell proliferation.

Acetylcholine-induced activation of M3 muscarinic receptors stimulates robust matrix metalloproteinase gene expression in human colon cancer cells

PubMed Central

Xie, Guofeng; Cheng, Kunrong; Shant, Jasleen; Raufman, Jean-Pierre

2009-01-01

Previously, we showed that ACh-induced proliferation of human colon cancer cells is mediated by transactivation of epidermal growth factor (EGF) receptors (EGFRs). In the present study, we elucidate the molecular mechanism underlying this action. ACh-induced proliferation of H508 colon cancer cells, which express exclusively M3 muscarinic receptors (M3Rs), was attenuated by anti-EGFR ligand binding domain antibody, a broad-spectrum matrix metalloproteinase (MMP) inhibitor, anti-MMP7 antibody, a diphtheria toxin analog that blocks release of an EGFR ligand [heparin-binding EGF-like growth factor (HBEGF)], and anti-HBEGF antibody. Conditioned media from ACh-treated H508 cells induced proliferation of SNU-C4 colon cancer cells that express EGFR but not M3R. These actions were attenuated by an EGFR inhibitor and by anti-EGFR and anti-HBEGF antibodies. In H508, but not SNU-C4, colon cancer cells, ACh caused a striking dose- and time-dependent increase in levels of MMP7 mRNA and MMP7 protein. Similarly, ACh induced robust MMP1 and MMP10 gene transcription. ACh-induced MMP1, MMP7, and MMP10 gene transcription was attenuated by atropine, anti-EGFR antibody, and chemical inhibitors of EGFR and ERK activation. In contrast, inhibitors of phosphatidylinositol 3-kinase and NF-κB activation did not alter MMP gene transcription. Collectively, these findings indicate that MMP7-catalyzed release of HBEGF mediates ACh-induced transactivation of EGFR and consequent proliferation of colon cancer cells. ACh-induced activation of EGFR and downstream ERK signaling also regulates transcriptional activation of MMP7, thereby identifying a novel feed-forward mechanism for neoplastic cell proliferation. PMID:19221016

Down-regulation of cancer/testis antigen OY-TES-1 attenuates malignant behaviors of hepatocellular carcinoma cells in vitro.

PubMed

Fu, Jun; Luo, Bin; Guo, Wen-Wen; Zhang, Qing-Mei; Shi, Lei; Hu, Qi-Ping; Chen, Fang; Xiao, Shao-Wen; Xie, Xiao-Xun

2015-01-01

Cancer/testis (CT) antigens are normally expressed in testis and overexpressed in various tumor types. However, their biological function is largely unknown. OY-TES-1, one of cancer/testis (CT) antigens, is reported overexpression in hepatocellular carcinoma (HCC). And we assumed that OY-TES-1 contribute to oncogenesis and progression of HCC. In this study, we knocked down OY-TES-1 by small interference RNA (siRNA) in HCC cell lines (HepG2 and BEL-7404) to verify this assumption and evaluate its potential as therapeutic targets for HCC. We showed that down regulation of OY-TES-1 decreased cell growth, induced the G0/G1 arrest and apoptosis, and prevented migration and invasion in the two HCC cell lines. Further analysis revealed that down regulation of OY-TES-1 increased expression of apoptosis-regulated protein caspase-3, and decreased expression of cell cycle-regulated protein cyclin E, migration/invasion-regulated proteins MMP2 and MMP9. These findings may shed light on the gene therapy about the OY-TES-1 expression in HCC cells.

Down-regulation of cancer/testis antigen OY-TES-1 attenuates malignant behaviors of hepatocellular carcinoma cells in vitro

PubMed Central

Fu, Jun; Luo, Bin; Guo, Wen-Wen; Zhang, Qing-Mei; Shi, Lei; Hu, Qi-Ping; Chen, Fang; Xiao, Shao-Wen; Xie, Xiao-Xun

2015-01-01

Cancer/testis (CT) antigens are normally expressed in testis and overexpressed in various tumor types. However, their biological function is largely unknown. OY-TES-1, one of cancer/testis (CT) antigens, is reported overexpression in hepatocellular carcinoma (HCC). And we assumed that OY-TES-1 contribute to oncogenesis and progression of HCC. In this study, we knocked down OY-TES-1 by small interference RNA (siRNA) in HCC cell lines (HepG2 and BEL-7404) to verify this assumption and evaluate its potential as therapeutic targets for HCC. We showed that down regulation of OY-TES-1 decreased cell growth, induced the G0/G1 arrest and apoptosis, and prevented migration and invasion in the two HCC cell lines. Further analysis revealed that down regulation of OY-TES-1 increased expression of apoptosis-regulated protein caspase-3, and decreased expression of cell cycle-regulated protein cyclin E, migration/invasion-regulated proteins MMP2 and MMP9. These findings may shed light on the gene therapy about the OY-TES-1 expression in HCC cells. PMID:26339343

Interaction between the Wnt/β-catenin signaling pathway and the EMMPRIN/MMP-2, 9 route in periodontitis.

PubMed

Liu, X; Zhang, Z; Pan, S; Shang, S; Li, C

2018-06-14

In periodontitis, the Wnt/β-catenin signaling pathway is related to the metabolism of the alveolar bone; further, extracellular matrix metalloproteinase inducer (EMMPRIN) expression is correlated with matrix metalloproteinases (MMPs) expression and inflammation severity. The aim of this study was to perform a preliminary investigation of the interaction between the Wnt/β-catenin signaling pathway and the EMMPRIN/MMPs route in periodontitis. Chronic periodontitis and healthy gingival tissues were obtained to detect the expression of Wnt3a, β-catenin, EMMPRIN and MMP-2, 9 by using immunohistochemical analysis. The human immortalized oral epithelial cell/human gingival fibroblast direct co-culture model was treated with 10 μg/mL Porphyromonas gingivalis lipopolysaccharide (Pg. LPS). Anti-EMMPRIN antibody was used to block the effect of EMMPRIN. Dickkopf-1 (DKK-1) and Wnt3a were used as the inhibitor and activator of the Wnt/β-catenin signaling pathway, respectively. Immunofluorescence was performed to visualize the localization of β-catenin and EMMPRIN. Expression of the EMMPRIN, MMP-2, 9 and Wnt pathway's components was confirmed by western blotting and quantitative real-time polymerase chain reaction. Higher levels of Wnt3a, β-catenin, EMMPRIN and MMP-2, 9 were observed in chronic periodontitis gingival tissues compared with controls. Pg. LPS significantly enhanced β-catenin, p-GSK-3β, EMMPRIN and MMP-2, 9 inductions in the human immortalized oral epithelial cell/human gingival fibroblast co-culture model. Anti-EMMPRIN antibody markedly reduced the expression of MMP-2, 9 only in the presence of Pg. LPS. Co-expression of β-catenin and EMMPRIN was detected in the co-culture model. DKK-1 inhibited Wnt pathway, but upregulated the EMMPRIN/MMP-2, 9 routes. In contrast, activating Wnt pathway by Wnt3a repressed the EMMPRIN/MMP-2, 9 routes. The promotion effect of DKK-1 on MMP-2, 9 expressions was partially inhibited by the anti-EMMPRIN antibody. In addition, anti-EMMPRIN antibody led to a drastic decrease in β-catenin and p-GSK-3β. In periodontitis, EMMPRIN regulates MMP-2, 9 expressions, the activation of Wnt/β-catenin signaling pathway downregulates the EMMPRIN/MMP-2, 9 routes and the blockade of EMMPRIN attenuates Wnt/β-catenin signaling pathway. © 2018 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

FGF-1-induced matrix metalloproteinase-9 expression in breast cancer cells is mediated by increased activities of NF-kappaB and activating protein-1.

PubMed

Lungu, Gina; Covaleda, Lina; Mendes, Odete; Martini-Stoica, Heidi; Stoica, George

2008-06-01

Matrix metalloproteinase-9 (MMP-9) plays a critical role in tumor invasion and metastasis. Here, we investigate the effect of fibroblast growth factor-1 (FGF-1) on the expression of MMP-9 in ENU1564, an ethyl-N-nitrosourea-induced rat mammary adenocarcinoma cell line. We observed that FGF-1 induces a dose-dependent increase in MMP-9 mRNA, protein, and activity in ENU1564 cells. To gain insight into the molecular mechanism of MMP-9 regulation by FGF-1, we investigated the role of components of PI3K-Akt and MEK1/2-ERK signaling pathways in our system since NF-kappaB and AP-1 transcription factor binding sites have been characterized in the upstream region of the MMP-9 gene. We demonstrated that FGF-1 increases Akt phosphorylation, triggers nuclear translocation of NF-kappaBp65, and enhances degradation of cytoplasmic IkappaBalpha. Pretreatment of cells with LY294002, a PI3K inhibitor, significantly inhibited MMP-9 protein expression in FGF-1-treated cells. Conversely, our data show that FGF-1 increases ERK phosphorylation in ENU1564 cells, increases c-jun and c-fos mRNA expression in a time-dependent manner, and triggers nuclear translocation of c-jun. Pretreatment of cells with PD98059, a MEK1/2 inhibitor significantly inhibited MMP-9 protein expression in FGF-1 treated cells. Finally, we observed increased DNA binding of NF-kappaB and AP-1 in FGF-1-treated cells and that mutation of either NF-kappaB or AP-1 response elements prevented MMP-9 promoter activation by FGF-1. Taken together, these results demonstrated that FGF-1-induced MMP-9 expression in ENU1564 cells is associated with increasing DNA binding activities of NF-kappaB and AP-1 and involve activation of a dual signaling pathway, PI3K-Akt and MEK1/2-ERK. (c) 2007 Wiley-Liss, Inc.

Peroxisome proliferator-activated receptor δ inhibits Porphyromonas gingivalis lipopolysaccharide-induced activation of matrix metalloproteinase-2 by downregulating NADPH oxidase 4 in human gingival fibroblasts.

PubMed

Yoo, T; Ham, S A; Hwang, J S; Lee, W J; Paek, K S; Oh, J W; Kim, J H; Do, J T; Han, C W; Kim, J H; Seo, H G

2016-10-01

We investigated the roles of peroxisome proliferator-activated receptor δ (PPARδ) in Porphyromonas gingivalis-derived lipopolysaccharide (Pg-LPS)-induced activation of matrix metalloproteinase 2 (MMP-2). In human gingival fibroblasts (HGFs), activation of PPARδ by GW501516, a specific ligand of PPARδ, inhibited Pg-LPS-induced activation of MMP-2 and generation of reactive oxygen species (ROS), which was associated with reduced expression of NADPH oxidase 4 (Nox4). These effects were significantly smaller in the presence of small interfering RNA targeting PPARδ or the specific PPARδ inhibitor GSK0660, indicating that PPARδ is involved in these events. In addition, modulation of Nox4 expression by small interfering RNA influenced the effect of PPARδ on MMP-2 activity, suggesting a mechanism in which Nox4-derived ROS modulates MMP-2 activity. Furthermore, c-Jun N-terminal kinase and p38, but not extracellular signal-regulated kinase, mediated PPARδ-dependent inhibition of MMP-2 activity in HGFs treated with Pg-LPS. Concomitantly, PPARδ-mediated inhibition of MMP-2 activity was associated with the restoration of types I and III collagen to levels approaching those in HGFs not treated with Pg-LPS. These results indicate that PPARδ-mediated downregulation of Nox4 modulates cellular redox status, which in turn plays a critical role in extracellular matrix homeostasis through ROS-dependent regulation of MMP-2 activity. © 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

PAI-1, a target gene of miR-143, regulates invasion and metastasis by upregulating MMP-13 expression of human osteosarcoma.

PubMed

Hirahata, Mio; Osaki, Mitsuhiko; Kanda, Yusuke; Sugimoto, Yui; Yoshioka, Yusuke; Kosaka, Nobuyoshi; Takeshita, Fumitaka; Fujiwara, Tomohiro; Kawai, Akira; Ito, Hisao; Ochiya, Takahiro; Okada, Futoshi

2016-05-01

Despite recent improvements in the therapy for osteosarcoma, 30-40% of osteosarcoma patients die of this disease, mainly due to its lung metastasis. We have previously reported that intravenous injection of miR-143 significantly suppresses lung metastasis of human osteosarcoma cells (143B) in a mouse model. In this study, we examined the biological role and mechanism of miR-143 in the metastasis of human osteosarcoma cells. We identified plasminogen activator inhibitor-1 (PAI-1) as a direct target gene of miR-143. To determine the role of PAI-1 in human osteosarcoma cells, siRNA was transfected into 143B cells for knockdown of PAI-1 expression. An in vitro study showed that downregulation of PAI-1 suppressed cell invasion activity, but not proliferation. Moreover, injection of PAI-1 siRNA into a primary lesion in the osteosarcoma mouse model inhibited lung metastasis compared to control siRNA-injected mice, without influencing the proliferative activity of the tumor cells. Subsequent examination using 143B cells revealed that knockdown of PAI-1 expression resulted in downregulation of the expression and secretion of matrix metalloproteinase-13 (MMP-13), which is also a target gene of miR-143 and a proteolytic enzyme that regulates tumor-induced osteolysis. Immunohistochemical analysis using clinical samples showed that higher miR-143 expressing cases showed poor expression of PAI-1 in the primary tumor cells. All such cases belonged to the lung metastasis-negative group. Moreover, the frequency of lung metastasis-positive cases was significantly higher in PAI-1 and MMP-13 double-positive cases than in PAI-1 or MMP-13 single-positive or double-negative cases (P < 0.05). These results indicated that PAI-1, a target gene of miR-143, regulates invasion and lung metastasis via enhancement of MMP-13 expression and secretion in human osteosarcoma cells, suggesting that these molecules could be potential therapeutic target genes for preventing lung metastasis in osteosarcoma patients. © 2016 The Authors. Cancer Medicine published by John Wiley & Sons Ltd.

Gallic Acid Inhibited Matrix Invasion and AP-1/ETS-1-Mediated MMP-1 Transcription in Human Nasopharyngeal Carcinoma Cells

PubMed Central

S. Pang, Jong-Hwei; Yen, Jia-Hau; Wu, Hsiao-Ting; Huang, Sheng-Teng

2017-01-01

Gallic acid is a trihydroxybenzoic acid found in natural herbal plants. Gallic acid has been reported to inhibit the migration and invasive capability of various cancers. Little is known about the underlying mechanisms of invasion responsible for cancer metastasis via gallic acid. The present study was intended to investigate the anti-invasive effect of gallic acid on human nasopharyngeal carcinoma cells (NPC-BM1) and its related mechanism. Gallic acid inhibited the invasion of NPC-BM1 cells dose- and time-dependently without significant cytotoxic effect. Affymetrix oligonucleotide microarray analysis revealed matrix metalloproteinase-1 (MMP-1) as the most down-regulated gene in NPC-BM1 cells by gallic acid. The cytosolic and secreted MMP-1 levels were both found to be inhibited by gallic acid as demonstrated by western blot analysis and ELISA respectively. The mRNA expression and transcription of MMP-1 gene was also down-regulated as determined by RT/real-time PCR and promoter activity assay. The expression of two major transcription binding factors in the MMP-1 promoter, AP-1 and ETS-1, were demonstrated to be reduced by gallic acid in NPC-BM1 cells. The effect of gallic acid was associated with the inhibition of p38 MAPK signaling pathway. In addition, gallic acid enhanced the gene expression of tissue inhibitor of matrix metalloproteinase-1 (TIMP-1) which further suppressed the MMP-1 activity. These findings may be useful to develop a novel chemotherapeutic agent to inhibit the metastasis of nasopharyngeal cancer. PMID:28672814

Gallic Acid Inhibited Matrix Invasion and AP-1/ETS-1-Mediated MMP-1 Transcription in Human Nasopharyngeal Carcinoma Cells.

PubMed

Pang, Jong-Hwei S; Yen, Jia-Hau; Wu, Hsiao-Ting; Huang, Sheng-Teng

2017-06-24

Gallic acid is a trihydroxybenzoic acid found in natural herbal plants. Gallic acid has been reported to inhibit the migration and invasive capability of various cancers. Little is known about the underlying mechanisms of invasion responsible for cancer metastasis via gallic acid. The present study was intended to investigate the anti-invasive effect of gallic acid on human nasopharyngeal carcinoma cells (NPC-BM1) and its related mechanism. Gallic acid inhibited the invasion of NPC-BM1 cells dose- and time-dependently without significant cytotoxic effect. Affymetrix oligonucleotide microarray analysis revealed matrix metalloproteinase-1 (MMP-1) as the most down-regulated gene in NPC-BM1 cells by gallic acid. The cytosolic and secreted MMP-1 levels were both found to be inhibited by gallic acid as demonstrated by western blot analysis and ELISA respectively. The mRNA expression and transcription of MMP-1 gene was also down-regulated as determined by RT/real-time PCR and promoter activity assay. The expression of two major transcription binding factors in the MMP-1 promoter, AP-1 and ETS-1, were demonstrated to be reduced by gallic acid in NPC-BM1 cells. The effect of gallic acid was associated with the inhibition of p38 MAPK signaling pathway. In addition, gallic acid enhanced the gene expression of tissue inhibitor of matrix metalloproteinase-1 (TIMP-1) which further suppressed the MMP-1 activity. These findings may be useful to develop a novel chemotherapeutic agent to inhibit the metastasis of nasopharyngeal cancer.

MicroRNA-188-5p regulates contribution of bone marrow-derived cells to choroidal neovascularization development by targeting MMP-2/13.

PubMed

Hou, Huiyuan; Gao, Fan; Liang, Hongliang; Lv, Yang; Li, Manhong; Yao, Libo; Zhang, Jian; Dou, Guorui; Wang, Yusheng

2018-06-14

Our previous investigations have shown that bone marrow-derived cells (BMCs), including mesenchymal stem cells (MSCs), contribute to the development of choroidal neovascularization (CNV) as sources of cells and angiogenic factors. Two main steps for circulating BMCs to integrate into CNV lesions are extracellular matrix remodeling and consequential cell migration. MicroRNAs (miRNAs) were found to be involved in CNV development; however, little is known about whether miRNAs regulate the contribution of BMCs to CNV. In the present study, we found that the expression of miR-188-5p was decreased in cultured hypoxic MSCs and BMCs within laser-induced CNV in mice. Matrix metalloproteinase 2 (MMP-2) and MMP-13 were both discovered as targets of miR-188-5p by bioinformatics predictions and dual-luciferase reporter system. Accordingly, increased expression of MMP-2/13 was found in hypoxic MSCs and BMCs in CNV lesions. Furthermore, miR-188-5p mimic transfection caused downregulation of MMP-2/13 in hypoxic MSCs and decreased tube formation of co-cultured vascular endothelial cells. Intravitreal injections of a miR-188-5p agomir attenuated the severity of CNV and inhibited the migration of BMCs into CNV lesions in mice. Our study suggests that miR-188-5p regulates the contribution of BMCs to CNV development by targeting MMP-2/13-mediated extracellular matrix degeneration, and miR-188-5p serves as a therapeutic target to treat CNV-related diseases. Copyright © 2018. Published by Elsevier Ltd.

Matrix metalloproteinase-9 and -2 enhance the ligand sensitivity of photoreceptor cyclic nucleotide-gated channels.

PubMed

Meighan, Peter C; Meighan, Starla E; Rich, Elizabeth D; Brown, R Lane; Varnum, Michael D

2012-01-01

Photoreceptor cyclic nucleotide-gated (CNG) channels are the principal ion channels responsible for transduction of the light-induced change in cGMP concentration into an electrical signal. The ligand sensitivity of photoreceptor CNG channels is subject to regulation by intracellular signaling effectors, including calcium-calmodulin, tyrosine kinases and phosphoinositides. Little is known, however, about regulation of channel activity by modification to extracellular regions of CNG channel subunits. Extracellular proteases MMP9 and -2 are present in the interphotoreceptor matrix adjacent to photoreceptor outer segments. Given that MMPs have been implicated in retinal dysfunction and degeneration, we hypothesized that MMP activity may alter the functional properties of photoreceptor CNG channels. For heterologously expressed rod and cone CNG channels, extracellular exposure to MMPs dramatically increased the apparent affinity for cGMP and the efficacy of cAMP. These changes to ligand sensitivity were not prevented by destabilization of the actin cytoskeleton or by disruption of integrin mediated cell adhesion, but could be attenuated by inhibition of MMP catalytic activity. MMP-mediated gating changes exhibited saturable kinetic properties consistent with enzymatic processing of the CNG channels. In addition, exposure to MMPs decreased the abundance of full-length expressed CNGA3 subunits, with a concomitant increase in putative degradation products. Similar gating effects and apparent proteolysis were observed also for native rod photoreceptor CNG channels. Furthermore, constitutive apparent proteolysis of retinal CNGA1 and retinal MMP9 levels were both elevated in aged mice compared with young mice. Together, these results provide evidence that MMP-mediated proteolysis can regulate the ligand sensitivity of CNG channels.

Matrix metalloproteinase-9 and -2 enhance the ligand sensitivity of photoreceptor cyclic nucleotide-gated channels

PubMed Central

Meighan, Peter C.; Meighan, Starla E.; Rich, Elizabeth D.; Brown, R. Lane; Varnum, Michael D.

2012-01-01

Photoreceptor cyclic nucleotide-gated (CNG) channels are the principal ion channels responsible for transduction of the light-induced change in cGMP concentration into an electrical signal. The ligand sensitivity of photoreceptor CNG channels is subject to regulation by intracellular signaling effectors, including calcium-calmodulin, tyrosine kinases and phosphoinositides. Little is known, however, about regulation of channel activity by modification to extracellular regions of CNG channel subunits. Extracellular proteases MMP9 and -2 are present in the interphotoreceptor matrix adjacent to photoreceptor outer segments. Given that MMPs have been implicated in retinal dysfunction and degeneration, we hypothesized that MMP activity may alter the functional properties of photoreceptor CNG channels. For heterologously expressed rod and cone CNG channels, extracellular exposure to MMPs dramatically increased the apparent affinity for cGMP and the efficacy of cAMP. These changes to ligand sensitivity were not prevented by destabilization of the actin cytoskeleton or by disruption of integrin mediated cell adhesion, but could be attenuated by inhibition of MMP catalytic activity. MMP-mediated gating changes exhibited saturable kinetic properties consistent with enzymatic processing of the CNG channels. In addition, exposure to MMPs decreased the abundance of full-length expressed CNGA3 subunits, with a concomitant increase in putative degradation products. Similar gating effects and apparent proteolysis were observed also for native rod photoreceptor CNG channels. Furthermore, constitutive apparent proteolysis of retinal CNGA1 and retinal MMP9 levels were both elevated in aged mice compared with young mice. Together, these results provide evidence that MMP-mediated proteolysis can regulate the ligand sensitivity of CNG channels. PMID:22699690

Recanalization and flow regulate venous thrombus resolution and matrix metalloproteinase expression in vivo.

PubMed

Chabasse, Christine; Siefert, Suzanne A; Chaudry, Mohammed; Hoofnagle, Mark H; Lal, Brajesh K; Sarkar, Rajabrata

2015-01-01

We examined the role of thrombus recanalization and ongoing blood flow in the process of thrombus resolution by comparing two murine in vivo models of deep venous thrombosis. In CD1 mice, we performed surgical inferior vena cava ligation (stasis thrombosis), stenosis (thrombosis with recanalization), or sham procedure. We analyzed thrombus weight over time as a measure of thrombus resolution and quantified the messenger RNA and protein levels of membrane-type matrix metalloproteinases (MT-MMPs) as well as effectors of the plasmin complex at days 4, 8, and 12 after surgery. Despite similar initial thrombus size, the presence of ongoing blood flow (stenosis model) was associated with a 45.91% subsequent improvement in thrombus resolution at day 8 and 12.57% at day 12 compared with stasis thrombosis (ligation model). Immunoblot and real-time polymerase chain reaction analysis demonstrated a difference in MMP-2 and MMP-9 activity at day 8 between the two models (P = .03 and P = .006, respectively) as well as a difference in MT2-MMP gene expression at day 8 (P = .044) and day 12 (P = .03) and MT1-MMP protein expression at day 4 (P = .021). Histologic analyses revealed distinct areas of recanalization in the thrombi of the stenosis model compared with the ligation model as well as the recruitment of inflammatory cells, especially macrophages, and a focal pattern of localized expression of MT1-MMP and MT3-MMP proteins surrounding the areas of recanalization in the stenosis model. Recanalization and ongoing blood flow accelerate deep venous thrombus resolution in vivo and are associated with distinct patterns of MT1-MMP and MT3-MMP expression and macrophage localization in areas of intrathrombus recanalization. Copyright © 2015 Society for Vascular Surgery. Published by Elsevier Inc. All rights reserved.

Isolation and characterization of novel RECK tumor suppressor gene splice variants

PubMed Central

Trombetta-Lima, Marina; Winnischofer, Sheila Maria Brochado; Demasi, Marcos Angelo Almeida; Filho, Renato Astorino; Carreira, Ana Claudia Oliveira; Wei, Beiyang; de Assis Ribas, Thais; Konig, Michelle Silberspitz; Bowman-Colin, Christian; Oba-Shinjo, Sueli Mieko; Marie, Suely Kazue Nagahashi; Stetler-Stevenson, William; Sogayar, Mari Cleide

2015-01-01

Glioblastoma multiforme is the most common and lethal of the central nervous system glial-derived tumors. RECK suppresses tumor invasion by negatively regulating at least three members of the matrix metalloproteinase family: MMP-9, MMP-2, and MT1-MMP. A positive correlation has been observed between the abundance of RECK expression in tumor samples and a more favorable prognosis for patients with several types of tumors. In the present study, novel alternatively spliced variants of the RECK gene: RECK-B and RECK-I were isolated by RT-PCR and sequenced. The expression levels and profiles of these alternative RECK transcripts, as well as canonical RECK were determined in tissue samples of malignant astrocytomas of different grades and in a normal tissue RNA panel by qRT-PCR. Our results show that higher canonical RECK expression, accompanied by a higher canonical to alternative transcript expression ratio, positively correlates with higher overall survival rate after chemotherapeutic treatment of GBM patients. U87MG and T98G cells over-expressing the RECK-B alternative variant display higher anchorage-independent clonal growth and do not display modulation of, respectively, MMP-2 and MMP-9 expression. Our findings suggest that RECK transcript variants might have opposite roles in GBM biology and the ratio of their expression levels may be informative for the prognostic outcome of GBM patients. PMID:26431549

Screening of differentially expressed genes between multiple trauma patients with and without sepsis.

PubMed

Ji, S C; Pan, Y T; Lu, Q Y; Sun, Z Y; Liu, Y Z

2014-03-17

The purpose of this study was to identify critical genes associated with septic multiple trauma by comparing peripheral whole blood samples from multiple trauma patients with and without sepsis. A microarray data set was downloaded from the Gene Expression Omnibus (GEO) database. This data set included 70 samples, 36 from multiple trauma patients with sepsis and 34 from multiple trauma patients without sepsis (as a control set). The data were preprocessed, and differentially expressed genes (DEGs) were then screened for using packages of the R language. Functional analysis of DEGs was performed with DAVID. Interaction networks were then established for the most up- and down-regulated genes using HitPredict. Pathway-enrichment analysis was conducted for genes in the networks using WebGestalt. Fifty-eight DEGs were identified. The expression levels of PLAU (down-regulated) and MMP8 (up-regulated) presented the largest fold-changes, and interaction networks were established for these genes. Further analysis revealed that PLAT (plasminogen activator, tissue) and SERPINF2 (serpin peptidase inhibitor, clade F, member 2), which interact with PLAU, play important roles in the pathway of the component and coagulation cascade. We hypothesize that PLAU is a major regulator of the component and coagulation cascade, and down-regulation of PLAU results in dysfunction of the pathway, causing sepsis.

Tumor Necrosis Factor-α Regulates Triggering Receptor Expressed on Myeloid Cells-1-dependent Matrix Metalloproteinases in the Carotid Plaques of Symptomatic Patients with Carotid Stenosis

PubMed Central

Rao, Velidi H.; Rai, Vikrant; Stoupa, Samantha; Subramanian, Saravanan; Agrawal, Devendra K

2016-01-01

Objective To determine the relationship between increased triggering receptor expressed on myeloid cells (TREM)-1 and plaque stability in atherosclerotic carotid stenosis. Methods The mRNA transcripts and protein for TREM-1, MMP-1, MMP-9, collagen type I (COL1A1) and collagen type III (COL3A1) were analyzed by qPCR and immunofluorescence in both tissues and VSMCs isolated from atherosclerotic carotid plaques of symptomatic and asymptomatic patients with carotid stenosis. Results The TREM-1, MMP-1 and MMP-9 mRNA transcripts were significantly increased (TREM-1, p<0.01; MMP-1, p<0.01 and MMP-9, p<0.001) while COL1A1 and COL3A1 mRNA transcripts were decreased (p<0.001) in VSMCs isolated from carotid plaques of symptomatic (S) than asymptomatic (AS) patients. Stimulation of cells with TNF-α further increased the mRNA transcripts of TREM-1, MMPs, COL1A1 and COL3A1. Modulation of TREM-1 by treatment with TREM-1 decoy receptor rTREM-1/Fc, and either TREM-1 antibodies or TREM-1 siRNA attenuated the TNF-α induced expression of MMP-1 and MMP-9 (p<0.01) and COL1A1 and COL3A1 (p<0.01) in S compared to AS VSMCs isolated from carotid plaques. Inhibition of NF-kB (BAY 11-7085), JNK (SP600125) and PI3K (LY294002) signaling pathways decreased the expression of TREM-1 (p<0.01), MMP-1 (p<0.001) and MMP-9 (p<0.01) in TNF-α treated VSMCs isolated from S carotid plaques compared to AS patients. Conclusion Increased expression of TREM-1 in S compared to AS patients involving MMP-1 and MMP-9 suggest a potential role of TREM-1 in plaque destabilization. Selective blockade of TREM-1 may contribute to the development of new therapies and promising targets for stabilizing vulnerable atherosclerotic plaques. PMID:27017522

Social defeat stress promotes tumor growth and angiogenesis by upregulating vascular endothelial growth factor/extracellular signal-regulated kinase/matrix metalloproteinase signaling in a mouse model of lung carcinoma.

PubMed

Wu, Xiao; Liu, Bao-Jun; Ji, Shumeng; Wu, Jing-Feng; Xu, Chang-Qing; Du, Yi-Jie; You, Xiao-Fang; Li, Bei; Le, Jing-Jing; Xu, Hai-Lin; Duan, Xiao-Hong; Dong, Jing-Cheng

2015-07-01

Numerous epidemiological and experimental animal studies have indicated that chronic psychological stress may promote tumor development. However, the underlying molecular mechanisms by which chronic stress promotes tumorigenesis remain to be fully elucidated and animal models have not yet been well established. In the present study, an established mouse model of repeated social defeat stress (RSDS), was generated and used to investigate the effect of stress on tumor growth and metastasis. C57BL/6 mice were exposed to RSDS for 10 days, followed by subcutaneousl inoculation with Lewis lung carcinoma cells for seven days. The tumor weight and volume as well as the number of the lung metastatic nodules were then determined. Vascular endothelial growth factor (VEGF) serum levels were measured using ELISAs. In addition, expression levels of VEGF receptor (VEGFR) and L1 cell adhesion molecule (L1CAM) messenger (m)RNA were confirmed using reverse transcription quantitative polymerase chain reaction. Furthermore, protein expression levels of phosphorlyated extracellular signal-regulated kinase (pERK), matrix metalloproteinase (MMP)-2 and MMP-9 were examined using western blot analysis. The results showed that RSDS significantly increased the weight and the volume of the primary tumor as well as the number of the lung metastatic nodules. Serum VEGF levels were significantly higher in the tumor-stress group compared with those of the unstressed tumor mice. In addition, tumors in stressed animals demonstrated markedly enhanced expression of VEGFR-2 and L1CAM mRNA as well as pERK, MMP-2 and MMP-9 protein expression. In conclusion, these results suggested that RSDS contributed to lung cancer progression, angiogenesis and metastasis, which was partially associated with increased VEGF secretion and therefore the activation of the ERK signaling pathway, resulting in the induction of MMP-2 and MMP-9 protein expression.

Matrix metalloproteinase-2 and -9 are induced differently by metal nanoparticles in human monocytes: The role of oxidative stress and protein tyrosine kinase activation

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wan Rong; Department of Pathology, Fujian Medical University, Fujian; Mo Yiqun

2008-12-01

Recently, many studies have shown that nanoparticles can translocate from the lungs to the circulatory system. As a particulate foreign body, nanoparticles could induce host responses such as reactive oxygen species (ROS) generation, inflammatory cytokine and matrix metalloproteinase (MMP) release which play a major role in tissue destruction and remodeling. However, the direct effects of nanoparticles on leukocytes, especially monocytes, are still unclear. The objective of the present study was to compare the ability of Nano-Co and Nano-TiO{sub 2} to cause alteration of transcription and activity of MMPs and to explore possible mechanisms. We hypothesized that non-toxic doses of somemore » transition metal nanoparticles stimulate an imbalance of MMP/TIMP that cause MMP production that may contribute to their health effects. To test this hypothesis, U937 cells were treated with Nano-Co and Nano-TiO{sub 2} and cytotoxic effects and ROS generation were measured. The alteration of MMP-2 and MMP-9 expression and activity of MMP-2 and MMP-9 after exposure to these metal nanoparticles were subsequently determined. To investigate the potential signaling pathways involved in the Nano-Co-induced MMP activation, the ROS scavengers or inhibitors, AP-1 inhibitor, and protein tyrosine kinase (PTK) inhibitors were also used to pre-treat U937 cells. Our results demonstrated that exposure of U937 cells to Nano-Co, but not to Nano-TiO{sub 2}, at a dose that does not cause cytotoxicity, resulted in ROS generation and up-regulation of MMP-2 and MMP-9 mRNA expression{sub ..} Our results also showed dose- and time-related increases in pro-MMP-2 and pro-MMP-9 gelatinolytic activities in conditioned media after exposure of U937 cells to Nano-Co, but not to Nano-TiO{sub 2}. Nano-Co-induced pro-MMP-2 and pro-MMP-9 activity increases were inhibited by pre-treatment with ROS scavengers or inhibitors. We also demonstrated dose- and time-related decreases in tissue inhibitors of metalloproteinases 2 (TIMP-2) in U937 cells after exposure to Nano-Co, but not to Nano-TiO{sub 2}. However, neither Nano-Co nor Nano-TiO{sub 2} exposure led to any transcriptional change of TIMP-1. The decrease of TIMP-2 after exposure to Nano-Co was also inhibited by pre-treatment with ROS scavengers or inhibitors. Our results also showed that pre-treatment of U937 cells with AP-1 inhibitor, curcumin, or the PTK specific inhibitor, herbimycin A or genistein, prior to exposure to Nano-Co, significantly abolished Nano-Co-induced pro-MMP-2 and-9 activity. Our results suggest that Nano-Co causes an imbalance between the expression and activity of MMPs and their inhibitors which is mediated by the AP-1 and tyrosine kinase pathways due to oxidative stress.« less

Therapeutic potential of fibroblast growth factor-2 for hypertrophic scars: upregulation of MMP-1 and HGF expression.

PubMed

Eto, Hitomi; Suga, Hirotaka; Aoi, Noriyuki; Kato, Harunosuke; Doi, Kentaro; Kuno, Shinichiro; Tabata, Yasuhiko; Yoshimura, Kotaro

2012-02-01

Although hypertrophic scars (HTSs) and keloids are challenging problems, their pathogenesis is not well understood, making therapy difficult. We showed that matrix metalloproteinase (MMP)-1 expression was downregulated in HTS compared with normal skin from the same patients, whereas type 1 and 3 collagen and transforming growth factor-β (TGF-β) were upregulated. These differences, however, were not seen in cultured fibroblasts, suggesting the involvement of microenvironmental factors in the pathogenesis of HTS. Fibroblast growth factor-2 (FGF-2) highly upregulated the expression of MMP-1 and hepatocyte growth factor (HGF) in both HTS-derived and control fibroblasts; the upregulation was reversed by extracellular signal-regulated kinase (ERK) and c-Jun N-terminal kinase (JNK) inhibitors. An animal study using human HTS tissue implanted into nude mice indicated that controlled-release FGF-2 resulted in significantly less weight and decreased hydroxyproline content in HTS. Degradation of collagen fibers in FGF-2-treated HTS was also confirmed histologically. Western blotting showed that FGF-2-treated HTS expressed significantly higher MMP-1 protein than control. Decreased MMP-1 expression may be an important transcriptional change in HTS, and its reversal as well as upregulation of HGF by FGF-2 could be a new therapeutic approach for HTS.

PI16 is a shear stress and inflammation-regulated inhibitor of MMP2

PubMed Central

Hazell, Georgina G. J.; Peachey, Alasdair M. G.; Teasdale, Jack E.; Sala-Newby, Graciela B.; Angelini, Gianni D.; Newby, Andrew C.; White, Stephen J.

2016-01-01

Raised endothelial shear stress is protective against atherosclerosis but such protection may be lost at sites of inflammation. We found that four splice variants of the peptidase inhibitor 16 (PI16) mRNA are among the most highly shear stress regulated transcripts in human coronary artery endothelial cells (HCAECs), in vitro but that expression is reduced by inflammatory mediators TNFα and IL-1β. Immunohistochemistry demonstrated that PI16 is expressed in human coronary endothelium and in a subset of neointimal cells and medial smooth muscle cells. Adenovirus-mediated PI16 overexpression inhibits HCAEC migration and secreted matrix metalloproteinase (MMP) activity. Moreover, PI16 inhibits MMP2 in part by binding an exposed peptide loop above the active site. Our results imply that, at high endothelial shear stress, PI16 contributes to inhibition of protease activity; protection that can be reversed during inflammation. PMID:27996045

Overexpression of membrane sialic acid-specific sialidase Neu3 inhibits matrix metalloproteinase-9 expression in vascular smooth muscle cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Moon, Sung-Kwon; Cho, Seung-Hak; Kim, Kyung-Woon

2007-05-11

The ganglioside-specific sialidase Neu3 has been suggested to participate in cell growth, migration, and differentiation. Recent reports suggest that sialidase may be involved in intimal thickening, an early stage in the development of atherosclerosis. However, the role of the Neu3 gene in vascular smooth muscle cells (VSMC) responses has not yet been elucidated. To determine whether a Neu3 is able to modulate VSMC growth, the effect of overexpression of the Neu3 gene on cell proliferation was examined. However, the results show that the overexpression of this gene has no effect on DNA synthesis and ERK phosphorylation in cultured VSMC inmore » the presence of TNF-{alpha}. Because atherogenic effects need not be limited to proliferation, we decided to examine whether Neu3 exerted inhibitory effects on matrix metalloproteinase-9 (MMP-9) activity in TNF-{alpha}-induced VSMC. The expression of the Neu3 gene led to the inhibition of TNF-{alpha}-induced matrix metalloproteinase-9 (MMP-9) expression in VSMC as determined by zymography and immunoblot. Furthermore, Neu3 gene expression strongly decreased MMP-9 promoter activity in response to TNF-{alpha}. This inhibition was characterized by the down-regulation of MMP-9, which was transcriptionally regulated at NF-{kappa}B and activation protein-1 (AP-1) sites in the MMP-9 promoter. These findings suggest that the Neu3 gene represents a physiological modulator of VSMC responses that may contribute to plaque instability in atherosclerosis.« less

Quercetin Inhibits the Migration and Invasion of HCCLM3 Cells by Suppressing the Expression of p-Akt1, Matrix Metalloproteinase (MMP) MMP-2, and MMP-9.

PubMed

Lu, Jun; Wang, Zhiqiang; Li, Shuyan; Xin, Qi; Yuan, Miaomiao; Li, Huanping; Song, Xiaoxia; Gao, Haijun; Pervaiz, Nabeel; Sun, Xudong; Lv, Wei; Jing, Tao; Zhu, Yanmei

2018-04-27

BACKGROUND Quercetin is a natural bioactive flavonoid that is present in a wide variety of vegetables and fruits and exhibits a promising anti-metastasis property in various human cancer cells. However, the effect of quercetin on human HCCLM3 cells is unclear. MATERIAL AND METHODS In the current study, a wound-healing assay was performed using quercetin-treated HCCLM3 cells to further explore whether quercetin affects the motility of human HCCLM3 cells. Transwell assay was used to explore the potential effect of quercetin in HCCLM3 cells on cell migration and cell invasion. Western blotting analysis was used to explore the expression of p-Akt1, MMP-2, and MMP-9 in quercetin-treated HCCLM3 cells. RESULTS The wound-healing time was delayed in quercetin-treated HCCLM3 cells, and the ability to migrate and invade was inhibited in quercetin-treated human HCCLM3 cells. Moreover, the protein levels of p-Akt1, MMP-2, and MMP-9 were down-regulated in quercetin-treated HCCLM3 cells, as detected by Western blotting. CONCLUSIONS Our data show that quercetin attenuated cell migration and invasion by suppressing the protein levels of p-Akt1, MMP-2, and MMP-9 in HCCLM3 cells.

Collagenase-3 expression in periapical lesions: an immunohistochemical study.

PubMed

Bhalla, G; Astekar, M S; Ramesh, G; Kaur, P; Sowmya, G V

2014-08-01

Collagenase-3 (matrix metalloproteinase-13) is a metalloproteinase (MMP) that is associated with bone lesions and exhibits variable expression patterns in odontogenic cysts; it may play a role in regulating focal proliferation and maturation of jaw cyst epithelium. We studied the localization, staining intensity and distribution of collagenase-3 in 13 periapical granulomas with epithelium, 16 periapical granulomas without epithelium and 10 radicular cysts using archived formalin fixed, paraffin embedded tissues. A monoclonal antibody against human collagenase-3 was used to evaluate its expression. Immunohistochemical staining intensities of collagenase-3 in all periapical lesions were (-), 4 (10%); (+), 1 (3%); (++), 22 (56%) and (+++), 12 (31%); differences were not statistically significant. Immunohistochemical distribution of collagenase-3 in epithelial cells was (-), 17 (44%); (+), 17 (44%); (++), 5 (13%); in fibroblasts it was (-), 8 (20%); (+), 23 (59%); (++), 8 (21%); in plasma cells it was (-), 7 (18%); (+), 22 (56%); (++), 10 (26%); in macrophages it was (-), 7 (18%); (+), and 15 (38%); and (++), 17 (44%). Statistically significant differences were found in epithelial cells (p = 0.00) and fibroblasts (p = 0.02), whereas differences were not statistically significant for plasma cells and macrophages. Collagenase-3 may play a role in the conversion of a periapical granuloma with epithelium to radicular cyst. MMP's influence not only epithelial rest cell migration, but also invasion of various stromal cells into granulomatous tissue.

Activation of Peroxisome Proliferator-Activated Receptor Alpha Improves Aged and UV-Irradiated Skin by Catalase Induction.

PubMed

Shin, Mi Hee; Lee, Se-Rah; Kim, Min-Kyoung; Shin, Chang-Yup; Lee, Dong Hun; Chung, Jin Ho

2016-01-01

Peroxisome proliferator-activated receptor alpha (PPARα) is a nuclear hormone receptor involved in the transcriptional regulation of lipid metabolism, fatty acid oxidation, and glucose homeostasis. Its activation stimulates antioxidant enzymes such as catalase, whose expression is decreased in aged human skin. Here we investigated the expression of PPARα in aged and ultraviolet (UV)-irradiated skin, and whether PPARα activation can modulate expressions of matrix metalloproteinase (MMP)-1 and procollagen through catalase regulation. We found that PPARα mRNA level was significantly decreased in intrinsically aged and photoaged human skin as well as in UV-irradiated skin. A PPARα activator, Wy14643, inhibited UV-induced increase of MMP-1 and decrease of procollagen expression and caused marked increase in catalase expression. Furthermore, production of reactive oxygen species (ROS) was suppressed by Wy14643 in UV-irradiated and aged dermal fibroblasts, suggesting that the PPARα activation-induced upregulation of catalase leads to scavenging of ROS produced due to UV irradiation or aging. PPARα knockdown decreased catalase expression and abolished the beneficial effects of Wy14643. Topical application of Wy14643 on hairless mice restored catalase activity and prevented MMP-13 and inflammatory responses in skin. Our findings indicate that PPARα activation triggers catalase expression and ROS scavenging, thereby protecting skin from UV-induced damage and intrinsic aging.

Local blockage of EMMPRIN impedes pressure ulcers healing in a rat model.

PubMed

Zhao, Xi-Lan; Luo, Xiao; Wang, Ze-Xin; Yang, Guo-Li; Liu, Ji-Zhong; Liu, Ya-Qiong; Li, Ming; Chen, Min; Xia, Yong-Mei; Liu, Jun-Jie; Qiu, Shu-Ping; Gong, Xiao-Qing

2015-01-01

Excessive extracellular matrix degradation caused by the hyperfunction of matrix metalloproteinases (MMPs) has been implicated in the failure of pressure ulcers healing. EMMPRIN, as a widely expressed protein, has emerged as an important regulator of MMP activity. We hypothesize that EMMPRIN affects the process of pressure ulcer healing by modulating MMP activity. In the rat pressure ulcer model, the expression of EMMPRIN in ulcers detected by Western blot was elevated compared with that observed in normal tissue. To investigate the role of EMMPRIN in regulating ulcer healing, specific antibodies against EMMPRIN were used via direct administration on the pressure ulcer. Local blockage of EMMPRIN resulted in a poor ulcer healing process compared with control ulcers, which was the opposite of our expectation. Furthermore, inhibiting EMMPRIN minimally impacted MMP activity. However, the collagen content in the pressure ulcer was reduced in the EMMPRIN treated group. Angiogenesis and the expression of angiogenic factors in pressure ulcers were also reduced by EMMPRIN local blockage. The results in the present study indicate a novel effect of EMMPRIN in the regulation of pressure ulcer healing by controlling the collagen contents and angiogenesis rather than MMPs activity.

Induction of Proteinases in the Human Preovulatory Follicle of the Menstrual Cycle by hCG

PubMed Central

Rosewell, Katherine L.; Al-Alem, Linah; Zakerkish, Farnosh; McCord, Lauren; Akin, James W.; Chaffin, Charles L.; Brännström, Mats; Curry, Thomas E.

2014-01-01

Objective To explore the temporal expression in granulosa and theca cells of key members of the MMP and ADAMTS families across the periovulatory period in women in order to gain insight into their possible roles during ovulation and early luteinization. Design Experimental prospective clinical study and laboratory-based investigation. Setting University Medical Center and private IVF center. Animal and Patient(s) Thirty eight premenopausal women undergoing surgery for tubal ligation and 6 premenopausal women undergoing ART. Intervention(s) Administration of hCG and harvesting of follicles by laparoscopy and collection of granulosa-lutein cells at oocyte retrieval. Main Outcome Measure(s) Expression of mRNA for MMPs and ADAMTSs in human granulosa cells and theca cells collected across the periovulatory period of the menstrual cycle and in cultured granulosa-lutein cells after hCG. Localization of MMPs and ADAMTSs by immunohistochemistry. Result(s) Expression of MMP1 and MMP19 mRNA increased in both granulosa and theca cells after hCG administration. ADAMTS1 and ADAMTS 9 mRNA increased in granulosa cells after hCG treatment, however thecal cell expression for ADAMTS1 was unchanged while ADAMTS9 expression was decreased. Expression of MMP8 and MMP13 mRNA was unchanged. Immunohistochemistry confirmed the localization of MMP1, MMP19, ADAMTS1 and ADAMTS9 to the granulosa and thecal cell layers. Conclusion(s) The collection of the dominant follicle throughout the periovulatory period has allowed the identification of proteolytic remodeling enzymes in the granulosa and theca compartments that may be critically involved in human ovulation. These proteinases may work in concert to regulate breakdown of the follicular wall and release of the oocyte. PMID:25516084

Induction of proteinases in the human preovulatory follicle of the menstrual cycle by human chorionic gonadotropin.

PubMed

Rosewell, Katherine L; Al-Alem, Linah; Zakerkish, Farnosh; McCord, Lauren; Akin, James W; Chaffin, Charles L; Brännström, Mats; Curry, Thomas E

2015-03-01

To explore the temporal expression in granulosa and theca cells of key members of the MMP and ADAMTS families across the periovulatory period in women to gain insight into their possible roles during ovulation and early luteinization. Experimental prospective clinical study and laboratory-based investigation. University medical center and private IVF center. Thirty-eight premenopausal women undergoing surgery for tubal ligation and six premenopausal women undergoing assisted reproductive techniques. Administration of hCG and harvesting of follicles by laparoscopy and collection of granulosa-lutein cells at oocyte retrieval. Expression of mRNA for matrix metalloproteinase (MMPs) and the A disintegrin and metalloproteinase with thrombospondin-like motifs (ADAMTS) in human granulosa cells and theca cells collected across the periovulatory period of the menstrual cycle and in cultured granulosa-lutein cells after hCG. Localization of MMPs and ADAMTSs by immunohistochemistry. Expression of MMP1 and MMP19 mRNA increased in both granulosa and theca cells after hCG administration. ADAMTS1 and ADAMTS9 mRNA increased in granulosa cells after hCG treatment, however, thecal cell expression for ADAMTS1 was unchanged, while ADAMTS9 expression was decreased. Expression of MMP8 and MMP13 mRNA was unchanged. Immunohistochemistry confirmed the localization of MMP1, MMP19, ADAMTS1, and ADAMTS9 to the granulosa and thecal cell layers. The collection of the dominant follicle throughout the periovulatory period has allowed the identification of proteolytic remodeling enzymes in the granulosa and theca compartments that may be critically involved in human ovulation. These proteinases may work in concert to regulate breakdown of the follicular wall and release of the oocyte. Copyright © 2015 American Society for Reproductive Medicine. Published by Elsevier Inc. All rights reserved.

Genome-Wide Expression Profiling of Complex Regional Pain Syndrome

PubMed Central

Jin, Eun-Heui; Zhang, Enji; Ko, Youngkwon; Sim, Woo Seog; Moon, Dong Eon; Yoon, Keon Jung; Hong, Jang Hee; Lee, Won Hyung

2013-01-01

Complex regional pain syndrome (CRPS) is a chronic, progressive, and devastating pain syndrome characterized by spontaneous pain, hyperalgesia, allodynia, altered skin temperature, and motor dysfunction. Although previous gene expression profiling studies have been conducted in animal pain models, there genome-wide expression profiling in the whole blood of CRPS patients has not been reported yet. Here, we successfully identified certain pain-related genes through genome-wide expression profiling in the blood from CRPS patients. We found that 80 genes were differentially expressed between 4 CRPS patients (2 CRPS I and 2 CRPS II) and 5 controls (cut-off value: 1.5-fold change and p<0.05). Most of those genes were associated with signal transduction, developmental processes, cell structure and motility, and immunity and defense. The expression levels of major histocompatibility complex class I A subtype (HLA-A29.1), matrix metalloproteinase 9 (MMP9), alanine aminopeptidase N (ANPEP), l-histidine decarboxylase (HDC), granulocyte colony-stimulating factor 3 receptor (G-CSF3R), and signal transducer and activator of transcription 3 (STAT3) genes selected from the microarray were confirmed in 24 CRPS patients and 18 controls by quantitative reverse transcription-polymerase chain reaction (qRT-PCR). We focused on the MMP9 gene that, by qRT-PCR, showed a statistically significant difference in expression in CRPS patients compared to controls with the highest relative fold change (4.0±1.23 times and p = 1.4×10−4). The up-regulation of MMP9 gene in the blood may be related to the pain progression in CRPS patients. Our findings, which offer a valuable contribution to the understanding of the differential gene expression in CRPS may help in the understanding of the pathophysiology of CRPS pain progression. PMID:24244504

Abrogation of epithelial BMP2 and BMP4 causes Amelogenesis Imperfecta by reducing MMP20 and KLK4 expression.

PubMed

Xie, Xiaohua; Liu, Chao; Zhang, Hua; Jani, Priyam H; Lu, Yongbo; Wang, Xiaofang; Zhang, Bin; Qin, Chunlin

2016-05-05

Amelogenesis Imperfecta (AI) can be caused by the deficiencies of enamel matrix proteins, molecules responsible for the transportation and secretion of enamel matrix components, and proteases processing enamel matrix proteins. In the present study, we discovered the double deletion of bone morphogenetic protein 2 (Bmp2) and bone morphogenetic protein 4 (Bmp4) in the dental epithelium by K14-cre resulted in hypoplastic enamel and reduced density in X-ray radiography as well as shortened enamel rods under scanning electron microscopy. Such enamel phenotype was consistent with the diagnosis of hypoplastic amelogenesis imperfecta. Histological and molecular analyses revealed that the removal of matrix proteins in the mutant enamel was drastically delayed, which was coincided with the greatly reduced expression of matrix metalloproteinase 20 (MMP20) and kallikrein 4 (KLK4). Although the expression of multiple enamel matrix proteins was down-regulated in the mutant ameloblasts, the cleavage of ameloblastin was drastically impaired. Therefore, we attributed the AI primarily to the reduction of MMP20 and KLK4. Further investigation found that BMP/Smad4 signaling pathway was down-regulated in the K14-cre;Bmp2(f/f);Bmp4(f/f)ameloblasts, suggesting that the reduced MMP20 and KLK4 expression may be due to the attenuated epithelial BMP/Smad4 signaling.

Abrogation of epithelial BMP2 and BMP4 causes Amelogenesis Imperfecta by reducing MMP20 and KLK4 expression

PubMed Central

Xie, Xiaohua; Liu, Chao; Zhang, Hua; Jani, Priyam H.; Lu, Yongbo; Wang, Xiaofang; Zhang, Bin; Qin, Chunlin

2016-01-01

Amelogenesis Imperfecta (AI) can be caused by the deficiencies of enamel matrix proteins, molecules responsible for the transportation and secretion of enamel matrix components, and proteases processing enamel matrix proteins. In the present study, we discovered the double deletion of bone morphogenetic protein 2 (Bmp2) and bone morphogenetic protein 4 (Bmp4) in the dental epithelium by K14-cre resulted in hypoplastic enamel and reduced density in X-ray radiography as well as shortened enamel rods under scanning electron microscopy. Such enamel phenotype was consistent with the diagnosis of hypoplastic amelogenesis imperfecta. Histological and molecular analyses revealed that the removal of matrix proteins in the mutant enamel was drastically delayed, which was coincided with the greatly reduced expression of matrix metalloproteinase 20 (MMP20) and kallikrein 4 (KLK4). Although the expression of multiple enamel matrix proteins was down-regulated in the mutant ameloblasts, the cleavage of ameloblastin was drastically impaired. Therefore, we attributed the AI primarily to the reduction of MMP20 and KLK4. Further investigation found that BMP/Smad4 signaling pathway was down-regulated in the K14-cre;Bmp2f/f;Bmp4f/fameloblasts, suggesting that the reduced MMP20 and KLK4 expression may be due to the attenuated epithelial BMP/Smad4 signaling. PMID:27146352

Polysaccharide of Black cumin (Nigella sativa) modulates molecular signaling cascade of gastric ulcer pathogenesis.

PubMed

Manjegowda, Srikanta Belagihalli; Rajagopal, Harsha Mysore; Dharmesh, Shylaja Mallaiah

2017-08-01

Gastric ulcer is a multi-step disease and healing requires a complex process including repair and re-architecture of gastric mucosa with the involvement of molecular events. Current study was designed to understand the gastric ulcer healing mechanism of rhamnogalacturonan-I type pectic polysaccharide of black cumin (BCPP) utilizing acetic acid induced gastric ulcers in rats. BCPP fed groups at 200mg/kg b.w. for 10days showed up to 85% healing of gastric ulcers with modulation of key molecular events involved in ulcer healing process such as increase in gastric mucin content, cyclooxygenase-2 (Cox-2) and prostaglandin E 2 (PGE 2 ). The increase in extracellular signal-regulated kinase-2 (ERK-2) indicated that, BCPP could induce PGE-2 synthesis by increasing ERK-2 mediated COX-2 activity. Increase in matrix metalloproteinase-2 (MMP-2) and decrease in MMP-9 levels in BCPP treated groups indicated differential regulation of MMP-2 and 9, an essential event required for gastric mucosal re-modulation. BCPP containing bound phenolics (26mg/g) might have also played a role in increasing speed and quality of ulcer healing by inhibiting H + , K + -ATPase and decreasing free radical mediated oxidation and cellular damages. Overall, studies showed that the polysaccharide can mediate ulcer healing by modulating signaling pathways involved in either ulcer aggravation or healing process. Copyright © 2017. Published by Elsevier B.V.

Overexpression of Shox2 Leads to Congenital Dysplasia of the Temporomandibular Joint in Mice

PubMed Central

Li, Xihai; Liang, Wenna; Ye, Hongzhi; Weng, Xiaping; Liu, Fayuan; Liu, Xianxiang

2014-01-01

Our previous study reported that inactivation of Shox2 led to dysplasia and ankylosis of the temporomandibular joint (TMJ), and that replacing Shox2 with human Shox partially rescued the phenotype with a prematurely worn out articular disc. However, the mechanisms of Shox2 activity in TMJ development remain to be elucidated. In this study, we investigated the molecular and cellular basis for the congenital dysplasia of TMJ in Wnt1-Cre; pMes-stop Shox2 mice. We found that condyle and glenoid fossa dysplasia occurs primarily in the second week after the birth. The dysplastic TMJ of Wnt1-Cre; pMes-stop Shox2 mice exhibits a loss of Collagen type I, Collagen type II, Ihh and Gli2. In situ zymography and immunohistochemistry further demonstrate an up-regulation of matrix metalloproteinases (MMPs), MMP9 and MMP13, accompanied by a significantly increased cell apoptosis. In addition, the cell proliferation and expressions of Sox9, Runx2 and Ihh are no different in the embryonic TMJ between the wild type and mutant mice. Our results show that overexpression of Shox2 leads to the loss of extracellular matrix and the increase of cell apoptosis in TMJ dysplasia by up-regulating MMPs and down-regulating the Ihh signaling pathway. PMID:25062348

MMP-1 and Pro-MMP-10 as potential urinary pharmacodynamic biomarkers of FGFR3-targeted therapy in patients with bladder cancer.

PubMed

Du, Xiangnan; Lin, Benjamin C; Wang, Qian-Rena; Li, Hao; Ingalla, Ellen; Tien, Janet; Rooney, Isabelle; Ashkenazi, Avi; Penuel, Elicia; Qing, Jing

2014-12-15

The aim of this study was to identify noninvasive pharmacodynamic biomarkers of FGFR3-targeted therapies in bladder cancer to facilitate the clinical development of experimental agent targeting FGFR3. Potential soluble pharmacodynamic biomarkers of FGFR3 were identified using a combination of transcriptional profiling and biochemical analyses in preclinical models. Two matrix metalloproteinases (MMP), MMP-1 and MMP-10, were selected for further studies in human bladder cancer xenograft models treated with a specific anti-FGFR3 monoclonal antibody, R3Mab. Serum and urinary levels of MMP-1 and MMP-10 were determined in healthy donors and patients with bladder cancer. The modulation of MMP-1 and MMP-10 by R3Mab in patients with bladder cancer was further evaluated in a phase I dose-escalation study. MMP-1 and MMP-10 mRNA and protein were downmodulated by FGFR3 shRNA and R3Mab in bladder cancer cell lines. FGFR3 signaling promoted the expression and secretion of MMP-1 and pro-MMP-10 in a MEK-dependent fashion. In bladder cancer xenograft models, R3Mab substantially blocked tumor progression and reduced the protein levels of human MMP-1 and pro-MMP-10 in tumor tissues as well as in mouse serum. Furthermore, both MMP-1 and pro-MMP-10 were elevated in the urine of patients with advanced bladder cancer. In a phase I dose-escalation trial, R3Mab administration resulted in an acute reduction of urinary MMP-1 and pro-MMP-10 levels in patients with bladder cancer. These findings reveal a critical role of FGFR3 in regulating MMP-1 and pro-MMP-10 expression and secretion, and identify urinary MMP-1 and pro-MMP-10 as potential pharmacodynamic biomarkers for R3Mab in patients with bladder cancer. ©2014 American Association for Cancer Research.

The role of cyclooxygenase-2, interleukin-1β and fibroblast growth factor-2 in the activation of matrix metalloproteinase-1 in sheared-chondrocytes and articular cartilage.

PubMed

Guan, Pei-Pei; Guo, Jing-Wen; Yu, Xin; Wang, Yue; Wang, Tao; Konstantopoulos, Konstantinos; Wang, Zhan-You; Wang, Pu

2015-05-20

MMP-1 expression is detected in fluid shear stress (20 dyn/cm(2))-activated and osteoarthritic human chondrocytes, however, the precise mechanisms underlying shear-induced MMP-1 synthesis remain unknown. Using primary chondrocytes and T/C-28a2 chondrocytic cells as model systems, we report that prolonged application of high fluid shear to human chondrocytes induced the synthesis of cyclooxygenase-2 (COX-2), interleukin-1β (IL-1β) and fibroblast growth factor-2 (FGF-2), which led to a marked increase in MMP-1 expression. IL-1β, COX-2-dependent PGE2 activated the PI3-K/AKT and p38 signaling pathways, which were in turn responsible for MMP-1 synthesis via NF-κB- and c-Jun-transactivating pathways. Prolonged shear stress exposure (>12 h) induced 15-Deoxy-Δ(12,14)-prostaglandin J2 (15d-PGJ2) synthesis. Although 15d-PGJ2 suppressed PI3-K/AKT and p38 signaling pathways, it stimulated MMP-1 expression via activating heme oxygenase 1 (HO-1). The critical role of COX-2 in regulating MMP-1 expression in articular cartilage in vivo was demonstrated using COX-2(+/-) transgenic mice in the absence or presence of rofecoxib oral administration. These findings provide novel insights for developing therapeutic strategies to combat OA.

The Role of Cyclooxygenase-2, Interleukin-1β and Fibroblast Growth Factor-2 in the Activation of Matrix Metalloproteinase-1 in Sheared-Chondrocytes and Articular Cartilage

PubMed Central

Guan, Pei-Pei; Guo, Jing-Wen; Yu, Xin; Wang, Yue; Wang, Tao; Konstantopoulos, Konstantinos; Wang, Zhan-You; Wang, Pu

2015-01-01

MMP-1 expression is detected in fluid shear stress (20 dyn/cm2)-activated and osteoarthritic human chondrocytes, however, the precise mechanisms underlying shear-induced MMP-1 synthesis remain unknown. Using primary chondrocytes and T/C-28a2 chondrocytic cells as model systems, we report that prolonged application of high fluid shear to human chondrocytes induced the synthesis of cyclooxygenase-2 (COX-2), interleukin-1β (IL-1β) and fibroblast growth factor-2 (FGF-2), which led to a marked increase in MMP-1 expression. IL-1β, COX-2-dependent PGE2 activated the PI3-K/AKT and p38 signaling pathways, which were in turn responsible for MMP-1 synthesis via NF-κB- and c-Jun-transactivating pathways. Prolonged shear stress exposure (>12 h) induced 15-Deoxy-Δ12,14-prostaglandin J2 (15d-PGJ2) synthesis. Although 15d-PGJ2 suppressed PI3-K/AKT and p38 signaling pathways, it stimulated MMP-1 expression via activating heme oxygenase 1 (HO-1). The critical role of COX-2 in regulating MMP-1 expression in articular cartilage in vivo was demonstrated using COX-2+/− transgenic mice in the absence or presence of rofecoxib oral administration. These findings provide novel insights for developing therapeutic strategies to combat OA. PMID:25992485

HOXC6 promotes gastric cancer cell invasion by upregulating the expression of MMP9.

PubMed

Chen, Shi-Wei; Zhang, Qing; Xu, Zhi-Feng; Wang, Hai-Ping; Shi, Yi; Xu, Feng; Zhang, Wen-Jian; Wang, Ping; Li, Yong

2016-10-01

Previous studies have demonstrated that the homoebox C6 (HOXC6) gene is highly expressed in gastric cancer tissues and is associated with the depth of tumor invasion, and is associated with poor prognosis of gastric cancer patients expressing HOXC6. The present study investigated the effect and underlying mechanism of HOXC6 on the proliferation and metastasis of gastric cancer cells in vitro. Reverse transcription‑quantitative polymerase chain (PCR) reaction was used to investigate the expression levels of HOXC6 in different gastric cancer cell lines and the effect of different levels of expression on the proliferation of gastric cancer cells was determined by cell growth curve and plate colony formation. The effect of HOXC6 on the anchorage‑independent proliferation of gastric cancer cells was determined by soft agar colony formation assay while the Transwell invasion assay was used to investigate the effect of different levels of HOXC6 expression on the invasive and metastatic abilities of gastric cancer cells. Semi‑quantitative PCR was used to detect the effect of different levels of HOXC6 expression on the expression of matrix metalloproteinase (MMP)2 and MMP9 in gastric cancer cells. Immunoblotting was used to assess MMP9 signaling in the gastric cancer cells. The HOXC6 gene is highly expressed in the majority of the gastric cancer cell lines. Overexpression of HOXC6 promoted gastric cancer cell proliferation and colony formation ability while HOXC6 downregulation inhibited cell proliferation and clone forming ability. HOXC6 overexpression also enhanced the soft agar colony formation ability of gastric cancer cells while HOXC6 downregulation decreased the colony formation ability. Upregulated HOXC6 increased the migration and invasion abilities of gastric cancer cells while interfering with HOXC6 expression inhibited the migration and invasion of the gastric cancer cells. The expression of MMP9 was enhanced with an upregulation of HOXC6 expression while HOXC6 downregulation lowered MMP9 gene expression levels. Increased expression of HOXC6 in gastric cancer cell lines significantly activated extracellular signal‑regulated kinase signaling and upregulated MMP9. The HOXC6 gene promotes the proliferation of gastric cancer cells while upregulation of MMP9 promotes migration and invasion of gastric cancer cells.

Sivelestat sodium hydrate attenuates acute lung injury by decreasing systemic inflammation in a rat model of severe burns.

PubMed

Xiao, X-G; Zu, H-G; Li, Q-G; Huang, P

2016-01-01

Patients with severe burns often develop acute lung injury (ALI), systemic inflammatory response syndrome (SIRS) often complicates with ALI. Sivelestat sodium hydrate is an effective drug against ALI. However, the mechanisms of this beneficial effect are still poorly understood. In the current study, we evaluate the effects of sivelestat sodium hydrate on systemic and local inflammatory parameters (neutrophil elastase [NE], interleukin [IL]-8, matrix metalloproteinase [MMP] 2 and 9) in a rat model of severe burns and ALI. And to analyze the correlations between expression of NE and IL-8 and acute lung injury. 48 Sprague-Dawley (SD) rats were divided into 3 groups: normal control group, severe burns injury group and severe burns treated with sivelestat sodium hydrate group (SSI). The lung water content and PaO2 were detected in each group. Pathological manifestations in each group were observed for pathology scoring in SD rats with acute lung injury. ELISA was used for detecting expression of NE and IL-8 in serum and BAL specimens of SD rats in each group. RT-PCR was used to detect mRNA expression of NE and IL-8 in lung tissues of each group. Western blotting was used for detecting protein expression of MMP-2 and MMP-9 in lung tissues of each group. SPSS 18.0 was used for statistical analysis. The PaO2 was significantly increased after sivelestat sodium hydrate intravenous injection. Pathological score and water content of lung tissue were significantly decreased in SSI group compared with severe burns injury group, slightly higher than that normal control group. NE and IL-8 levels significantly decreased in serum, BAL and lung tissue specimens after sivelestat sodium hydrate intravenous injection; Expression of MMP-2 and MMP-9 were significantly up-regulated in severe burns group and showed no significantly changed after sivelestat sodium hydrate intravenous injection. In a rat model of severe burns and ALI, administration of sivelestat sodium hydrate improved symptoms of ALI and significantly decreased inflammatory parameters NE and IL-8.

Paeoniflorin Suppressed High Glucose-Induced Retinal Microglia MMP-9 Expression and Inflammatory Response via Inhibition of TLR4/NF-κB Pathway Through Upregulation of SOCS3 in Diabetic Retinopathy.

PubMed

Zhu, Su-Hua; Liu, Bing-Qian; Hao, Mao-Juan; Fan, Yi-Xin; Qian, Cheng; Teng, Peng; Zhou, Xiao-Wei; Hu, Liang; Liu, Wen-Tao; Yuan, Zhi-Lan; Li, Qing-Ping

2017-10-01

Diabetic retinopathy (DR) is a serious-threatening complication of diabetes and urgently needed to be treated. Evidence has accumulated indicating that microglia inflammation within the retina plays a critical role in DR. Microglial matrix metalloproteinase 9 (MMP-9) has an important role in the destruction of the integrity of the blood-retinal barrier (BRB) associated with the development of DR. MMP-9 was also considered important for regulating inflammatory responses. Paeoniflorin, a monoterpene glucoside, has a potent immunomodulatory effect on microglia. We hypothesized that paeoniflorin could significantly suppress microglial MMP-9 activation induced by high glucose and further relieve DR. BV2 cells were used to investigate the effects and mechanism of paeoniflorin. The activation of MMP-9 was measured by gelatin zymography. Cell signaling was measured by western blot assay and immunofluorescence assay. High glucose increased the activation of MMP-9 in BV2 cells, which was abolished by HMGB1, TLR4, p38 MAPK, and NF-κB inhibition. Phosphorylation of p38 MAPK induced by high glucose was decreased by TLR4 inhibition in BV2 cells. Paeoniflorin induced suppressor of cytokine signaling 3 (SOCS3) expression and reduced MMP-9 activation in BV2 cells. The effect of paeoniflorin on SOCS3 was abolished by the TLR4 inhibitor. In streptozotocin (STZ)-induced diabetes mice, paeoniflorin induced SOCS3 expression and reduced MMP-9 activation. Paeoniflorin suppressed STZ-induced IBA-1 and IL-1β expression and decreased STZ-induced high blood glucose level. In conclusion, paeoniflorin suppressed high glucose-induced retinal microglia MMP-9 expression and inflammatory response via inhibition of the TLR4/NF-κB pathway through upregulation of SOCS3 in diabetic retinopathy.

The Effect of Gestational Age on Angiogenic Gene Expression in the Rat Placenta

PubMed Central

Vaswani, Kanchan; Hum, Melissa Wen-Ching; Chan, Hsiu-Wen; Ryan, Jennifer; Wood-Bradley, Ryan J.; Nitert, Marloes Dekker; Mitchell, Murray D.; Armitage, James A.; Rice, Gregory E.

2013-01-01

The placenta plays a central role in determining the outcome of pregnancy. It undergoes changes during gestation as the fetus develops and as demands for energy substrate transfer and gas exchange increase. The molecular mechanisms that coordinate these changes have yet to be fully elucidated. The study performed a large scale screen of the transcriptome of the rat placenta throughout mid-late gestation (E14.25–E20) with emphasis on characterizing gestational age associated changes in the expression of genes invoved in angiogenic pathways. Sprague Dawley dams were sacrificed at E14.25, E15.25, E17.25 and E20 (n = 6 per group) and RNA was isolated from one placenta per dam. Changes in placental gene expression were identifed using Illumina Rat Ref-12 Expression BeadChip Microarrays. Differentially expressed genes (>2-fold change, <1% false discovery rate, FDR) were functionally categorised by gene ontology pathway analysis. A subset of differentially expressed genes identified by microarrays were confirmed using Real-Time qPCR. The expression of thirty one genes involved in the angiogenic pathway was shown to change over time, using microarray analysis (22 genes displayed increased and 9 gene decreased expression). Five genes (4 up regulated: Cd36, Mmp14, Rhob and Angpt4 and 1 down regulated: Foxm1) involved in angiogenesis and blood vessel morphogenesis were subjected to further validation. qPCR confirmed late gestational increased expression of Cd36, Mmp14, Rhob and Angpt4 and a decrease in expression of Foxm1 before labour onset (P<0.0001). The observed acute, pre-labour changes in the expression of the 31 genes during gestation warrant further investigation to elucidate their role in pregnancy. PMID:24391823

Matrix metalloproteinase-9 is involved in chronic lymphocytic leukemia cell response to fludarabine and arsenic trioxide.

PubMed

Amigo-Jiménez, Irene; Bailón, Elvira; Ugarte-Berzal, Estefanía; Aguilera-Montilla, Noemí; García-Marco, José A; García-Pardo, Angeles

2014-01-01

Matrix metalloproteinase-9 (MMP-9) contributes to chronic lymphocytic leukemia (CLL) pathology by regulating cell migration and preventing spontaneous apoptosis. It is not known if MMP-9 is involved in CLL cell response to chemotherapy and we address this in the present study, using arsenic trioxide (ATO) and fludarabine as examples of cytotoxic drugs. We used primary cells from the peripheral blood of CLL patients and MEC-1 cells stably transfected with an empty vector or a vector containing MMP-9. The effect of ATO and fludarabine was determined by flow cytometry and by the MTT assay. Expression of mRNA was measured by RT-PCR and qPCR. Secreted and cell-bound MMP-9 was analyzed by gelatin zymography and flow cytometry, respectively. Protein expression was analyzed by Western blotting and immunoprecipitation. Statistical analyses were performed using the two-tailed Student's t-test. In response to ATO or fludarabine, CLL cells transcriptionally upregulated MMP-9, preceding the onset of apoptosis. Upregulated MMP-9 primarily localized to the membrane of early apoptotic cells and blocking apoptosis with Z-VAD prevented MMP-9 upregulation, thus linking MMP-9 to the apoptotic process. Culturing CLL cells on MMP-9 or stromal cells induced drug resistance, which was overcome by anti-MMP-9 antibodies. Accordingly, MMP-9-MEC-1 transfectants showed higher viability upon drug treatment than Mock-MEC-1 cells, and this effect was blocked by silencing MMP-9 with specific siRNAs. Following drug exposure, expression of anti-apoptotic proteins (Mcl-1, Bcl-xL, Bcl-2) and the Mcl-1/Bim, Mcl-1/Noxa, Bcl-2/Bax ratios were higher in MMP-9-cells than in Mock-cells. Similar results were obtained upon culturing primary CLL cells on MMP-9. Our study describes for the first time that MMP-9 induces drug resistance by modulating proteins of the Bcl-2 family and upregulating the corresponding anti-apoptotic/pro-apoptotic ratios. This is a novel role for MMP-9 contributing to CLL progression. Targeting MMP-9 in combined therapies may thus improve CLL response to treatment.

Matrix Metalloproteinase-9 Is Involved in Chronic Lymphocytic Leukemia Cell Response to Fludarabine and Arsenic Trioxide

PubMed Central

Amigo-Jiménez, Irene; Bailón, Elvira; Ugarte-Berzal, Estefanía; Aguilera-Montilla, Noemí; García-Marco, José A.; García-Pardo, Angeles

2014-01-01

Background Matrix metalloproteinase-9 (MMP-9) contributes to chronic lymphocytic leukemia (CLL) pathology by regulating cell migration and preventing spontaneous apoptosis. It is not known if MMP-9 is involved in CLL cell response to chemotherapy and we address this in the present study, using arsenic trioxide (ATO) and fludarabine as examples of cytotoxic drugs. Methods We used primary cells from the peripheral blood of CLL patients and MEC-1 cells stably transfected with an empty vector or a vector containing MMP-9. The effect of ATO and fludarabine was determined by flow cytometry and by the MTT assay. Expression of mRNA was measured by RT-PCR and qPCR. Secreted and cell-bound MMP-9 was analyzed by gelatin zymography and flow cytometry, respectively. Protein expression was analyzed by Western blotting and immunoprecipitation. Statistical analyses were performed using the two-tailed Student's t-test. Results In response to ATO or fludarabine, CLL cells transcriptionally upregulated MMP-9, preceding the onset of apoptosis. Upregulated MMP-9 primarily localized to the membrane of early apoptotic cells and blocking apoptosis with Z-VAD prevented MMP-9 upregulation, thus linking MMP-9 to the apoptotic process. Culturing CLL cells on MMP-9 or stromal cells induced drug resistance, which was overcome by anti-MMP-9 antibodies. Accordingly, MMP-9-MEC-1 transfectants showed higher viability upon drug treatment than Mock-MEC-1 cells, and this effect was blocked by silencing MMP-9 with specific siRNAs. Following drug exposure, expression of anti-apoptotic proteins (Mcl-1, Bcl-xL, Bcl-2) and the Mcl-1/Bim, Mcl-1/Noxa, Bcl-2/Bax ratios were higher in MMP-9-cells than in Mock-cells. Similar results were obtained upon culturing primary CLL cells on MMP-9. Conclusions Our study describes for the first time that MMP-9 induces drug resistance by modulating proteins of the Bcl-2 family and upregulating the corresponding anti-apoptotic/pro-apoptotic ratios. This is a novel role for MMP-9 contributing to CLL progression. Targeting MMP-9 in combined therapies may thus improve CLL response to treatment. PMID:24956101

Diverse matrix metalloproteinase functions regulate cancer amoeboid migration

PubMed Central

Orgaz, Jose L.; Pandya, Pahini; Viros, Amaya; Albrengues, Jean; Nestle, Frank O.; Ridley, Anne J.; Gaggioli, Cedric; Marais, Richard; Karagiannis, Sophia N.; Sanz-Moreno, Victoria

2014-01-01

Rounded-amoeboid cancer cells use actomyosin contractility driven by Rho-ROCK and JAK-STAT3 to migrate efficiently. It has been suggested that rounded-amoeboid cancer cells do not require matrix metalloproteinases (MMPs) to invade. Here we compare MMP levels in rounded-amoeboid and elongated-mesenchymal melanoma cells. Surprisingly, we find that rounded-amoeboid melanoma cells secrete higher levels of several MMPs, including collagenase MMP-13 and gelatinase MMP-9. As a result, rounded-amoeboid melanoma cells degrade collagen I more efficiently than elongated-mesenchymal cells. Furthermore, using a non-catalytic mechanism, MMP-9 promotes rounded-amoeboid 3D migration through regulation of actomyosin contractility via CD44 receptor. MMP-9 is upregulated in a panel of rounded-amoeboid compared with elongated-mesenchymal melanoma cell lines and its levels are controlled by ROCK-JAK-STAT3 signalling. MMP-9 expression increases during melanoma progression and it is particularly prominent in the invasive fronts of lesions, correlating with cell roundness. Therefore, rounded-amoeboid cells use both catalytic and non-catalytic activities of MMPs for invasion. PMID:24963846

Expression of MMP-9 decreases metastatic potential of Chondrosarcoma: an immunohistochemical study.

PubMed

Malcherczyk, Dominik; Heyse, Thomas J; El-Zayat, Bilal F; Kunzke, Vanessa; Moll, Roland; Fuchs-Winkelmann, Susanne; Paletta, Jürgen R J

2018-01-09

Chondrosarcoma is the second most common primary malignant bone tumor. Because of their heterogeneity, with differences in invasive and metastatic behavior, it is important to identify biological markers that will allow for a more accurate estimation of prognosis in patients with these tumors. Matrix metalloproteinases (MMP) play a crucial role in tumor progression, invasion and metastasis. The mechanism of tumor progression dependent of MMPs is complex and influences malignant transformation, angiogenesis and tumor growth at the primary and metastatic sites. The purpose of this study was to investigate immunohistochemicaly the influence of MMP-1, MMP-3, MMP-9 and MMP-13 expression on prognostic parameter in chondrosarcoma. We investigated tissue samples of 28 patients with chondrosarcoma. Immunohistochemical staining to evaluate the expression of MMP-1, MMP-3, MMP-9 and MMP-13 was performed. Subsequently, the expression level was correlated with metastatic potential, histological grading and overall survival in patients with this neoplasm. In consideration of semi quantitative scoring 64% of chondrosarcoma were scored as positive for MMP-1, 46% for MMP-3, 61% for MMP-9. The specimens had shown no expression of MMP-13. High expression of MMP-9 was associated with better histological differentiation, decreased metastatic potential and favourable overall survival. No correlation was found for expression of MMP-1, MMP-3 or MMP-13. MMP-1, MMP-3 and MMP-9 are expressed in chondrosarcoma. Our findings suggest that the expression of MMP-9 is associated with clinical outcome parameters in chondrosarcoma.

Matrix metalloproteinase-2 and -9 in the cerebellum of teleost fish: Functional implications for adult neurogenesis.

PubMed

Sîrbulescu, Ruxandra F; Ilieş, Iulian; Zupanc, Günther K H

2015-09-01

Matrix metalloproteinases (MMPs) are a family of highly conserved zinc-dependent proteases involved in both development and pathogenesis. The present study examines the role of MMP-2 (gelatinase A) and MMP-9 (gelatinase B) in adult neurogenesis, using the corpus cerebelli, a subdivision of the cerebellum, of knifefish (Apteronotus leptorhynchus) as a model system. Transcripts of five isoforms of these gelatinases were identified in the central nervous system of this species. Sequence similarity analysis and homology modeling indicated that functionally and structurally critical elements were highly conserved in knifefish gelatinases. Immunohistochemical staining revealed a differential distribution of MMP-2 and MMP-9 at both the cellular and subcellular level. MMP-2 expression was found mainly in Sox2-immunopositive stem/progenitor cells, both quiescent and mitotically active; and was localized in both the cytoplasmic compartment and the nucleus. By contrast, MMP-9 immunoreactivity was absent in neurogenic niches and displayed a more homogenous distribution, with low to moderate intensity levels, in the molecular and granular layers. MMP-9 expression appeared to be restricted to the extracellular space. In situ zymography indicated that gelatinase activity matched the cellular and subcellular distributions of the two MMPs. The observed patterns of gelatinase activity and expression support the hypothesis that MMP-2 is primarily involved in regulation of the activity of stem/progenitor cells that give rise to new granule neurons, whereas MMP-9 facilitates migration of the progeny of these cells by proteolysis of extracellular matrix proteins. Copyright © 2015 Elsevier Inc. All rights reserved.

Presence of intestinal Mycobacterium avium subspecies paratuberculosis (MAP) DNA is not associated with altered MMP expression in ulcerative colitis

PubMed Central

2011-01-01

Background Mycobacterium avium subspecies paratuberculosis (MAP) is suspected to be a causative agent in human Crohn's disease (CD). Recent evidence suggests that pathogenic mycobacteria and MAP can induce the expression of Matrix Metalloproteinases (MMP), which are the main proteases in the pathogenesis of mucosal ulcerations in inflammatory bowel disease (IBD). Within this study we assessed the prevalence of intestinal MAP specific DNA in patients with Crohn's disease, ulcerative colitis (UC), and healthy controls. We further analysed regulation patterns of MMPs in mucosal tissues of UC patients with and without intestinal MAP DNA detection. Methods Colonic biopsy samples were obtained from 63 Norwegian and German IBD patients and 21 healthy controls. RNA was quantified by quantitative real-time polymerase chain reaction (PCR) to study MMP gene expression in both pathological and healthy mucosal specimens. The presence of MAP DNA in colonic mucosa was examined using MAP specific PCR. Results MAP DNA was detected in 20% of UC patients and 33% of healthy controls but only in 7% of patients with CD. UC patients treated with corticosteroids exhibited a significantly increased frequency of intestinal MAP DNA compared to those not receiving corticosteroids. Expression of MMP-1, -2, -7, -9, -13, -19, -28 and TNF-α did not differ between UC patients with presence of intestinal MAP DNA compared to those without. MMP-2, MMP-9 and MMP-13 were significantly decreased in UC patients receiving corticosteroids. Conclusions The presence of intestinal MAP specific DNA is not associated with altered MMP expression in UC in vivo. Corticosteroids are associated with increased detection of intestinal MAP DNA and decreased expression of certain MMPs. Frequent detection of MAP DNA in healthy controls might be attributable to the wide environmental distribution of MAP and its presence in the food-chain. PMID:21477272

ADAM17 is associated with EMMPRIN and predicts poor prognosis in patients with uterine cervical carcinoma.

PubMed

Xu, Qin; Ying, Mingang; Chen, Guilin; Lin, Ang; Xie, Yunqing; Ohara, Noriyuki; Zhou, Dongmei

2014-08-01

Metalloproteinase activities of a disintegrin and metalloproteinase 17 (ADAM17), amphiregulin (AREG), extracellular matrix metalloproteinase inducer (EMMPRIN), and matrix metalloproteinases (MMPs) are involved in tumor biology. In patients with uterine cervical carcinoma, the expression and prognostic significance of ADAM17 remain to be fully elucidated. The expression of ADAM17, AREG, EMMPRIN, phospho-epidermal growth factor receptor (p-EGFR), phospho-extracellular signal-regulated kinase (p-ERK), MMP-2, and MMP-9 was assessed by immunohistochemistry and/or Western blotting from cervical carcinoma cell lines, SiHa and HeLa cells, and cervical carcinoma tissues. AREG activity was measured by ELISA assay. The correlation of ADAM17, AREG, EMMPRIN, and MMP-9 expression with patients' survival rates was assessed by Kaplan-Meier and Cox regression analyses. RNA interference (RNAi) experiment was performed using small interfering mRNA to ADAM17 and EMMPRIN. ADAM17, EMMPRIN, and MMP-9 protein content was overexpressed in cervical carcinoma tissues compared with normal cervical tissues (P < 0.05). Strong expression of ADAM17, AREG, EMMPRIN, and MMP-9 was significantly associated with stages, lymph node metastasis, differentiation, and parametrium invasion (P < 0.05). Overexpression of ADAM17, AREG, EMMPRIN, and MMP-9 was significantly correlated with short progression-free survival and overall survival (P < 0.05). Multivariate analysis suggested that lymph node metastasis, parametrium invasion, and ADAM17 expression were independent prognostic indicators for cervical cancer. ADAM17 RNAi decreased EMMPRIN, p-EGFR, p-ERK, MMP-2, and MMP-9 proteins in SiHa and HeLa cells. ELISA assay revealed that AREG activity was stimulated by ADAM17 and was reversed by ADAM17 RNAi in SiHa and HeLa cells. Our data suggest that the increased expression of ADAM17 in cervical cancer is significantly associated with aggressive progression and poor prognosis. ADAM17 may be a molecular marker for predicting the progression and prognosis in cervical cancer.

Functional Roles of N-Linked Glycosylation of Human Matrix Metalloproteinase 9.

PubMed

Duellman, Tyler; Burnett, John; Yang, Jay

2015-10-01

Matrix metalloproteinase-9 (MMP-9) is a secreted endoproteinase with a critical role in the regulation of the extracellular matrix and proteolytic activation of signaling molecules. Human (h)MMP-9 has two well-defined N-glycosylation sites at residues N38 and N120; however, their role has remained mostly unexplored partly because expression of the N-glycosylation-deficient N38S has been difficult due to a recently discovered single nucleotide polymorphism-dependent miRNA-mediated inhibitory mechanism. hMMP-9 cDNA encoding amino acid substitutions at residues 38 (modified-S38, mS38) or 120 (N120S) were created in the background of a miRNA-binding site disrupted template and expressed by transient transfection. hMMP-9 harboring a single mS38 replacement secreted well, whereas N120S, or a double mS38/N120S hMMP-9 demonstrated much reduced secretion. Imaging indicated endoplasmic reticulum (ER) retention of the non-secreted variants and co-immunoprecipitation confirmed an enhanced strong interaction between the non-secreted hMMP-9 and the ER-resident protein calreticulin (CALR). Removal of N-glycosylation at residue 38 revealed an amino acid-dependent strong interaction with CALR likely preventing unloading of the misfolded protein from the ER chaperone down the normal secretory pathway. As with other glycoproteins, N-glycosylation strongly regulates hMMP-9 secretion. This is mediated, however, through a novel mechanism of cloaking an N-glycosylation-independent strong interaction with the ER-resident CALR. © 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Methamphetamine and HIV-1 gp120 Effects on Lipopolysaccharide Stimulated Matrix Metalloproteinase-9 Production by Human Monocyte-Derived Macrophages

PubMed Central

Reynolds, Jessica L.; Mahajan, Supriya D.; Aalinkeel, Ravikumar; Nair, Bindukumar; Sykes, Donald E.; Schwartz, Stanley A.

2011-01-01

Monocytes/macrophages are a primary source of human immunodeficiency virus (HIV-1) in the central nervous system (CNS). Macrophages infected with HIV-1 produce a plethora of factors, including matrix metalloproteinase-9 (MMP-9) that may contribute to the development of HIV-1-associated neurocognitive disorders (HAND). MMP-9 plays a pivotal role in the turnover of the extracellular matrix (ECM) and functions to remodel cellular architecture. We have investigated the role of methamphetamine and HIV-1 gp120 in the regulation of lipopolysaccaride (LPS) induced-MMP-9 production in monocyte-derived macrophages (MDM). Here, we show that LPS-induced MMP-9 gene expression and protein secretion are potentiated by incubation with methamphetamine alone and gp120 alone. Further, concomitant incubation with gp120 and methamphetamine potentiated LPS-induced MMP-9 expression and biological activity in MDM. Collectively methamphetamine and gp120 effects on MMPs may modulate remodeling of the extracellular environment enhancing migration of monocytes/macrophages to the CNS. PMID:21425912

Study on the relationship between the methylation of the MMP-9 gene promoter region and diabetic nephropathy.

PubMed

Yang, Xiao-Hui; Feng, Shi-Ya; Yu, Yang; Liang, Zhou

2018-01-01

This study aims to explore the relationship between the methylation of matrix metalloproteinase (MMP)-9 gene promoter region and diabetic nephropathy (DN) through the detection of the methylation level of MMP-9 gene promoter region in the peripheral blood of patients with DN in different periods and serum MMP-9 concentration. The methylation level of the MMP-9 gene promoter region was detected by methylation-specific polymerase chain reaction (MSP), and the content of MMP-9 in serum was determined by enzyme-linked immunosorbent assay (ELISA). Results of the statistical analysis revealed that serum MMP-9 protein expression levels gradually increased in patients in the simple diabetic group, early diabetic nephropathy group and clinical diabetic nephropathy group, compared with the control group; and the difference was statistically significant (P < 0.05). Compared with the control group, the methylation levels of MMP-9 gene promoter regions gradually decreased in patients in the simple diabetic group, early diabetic nephropathy group, and clinical diabetic nephropathy group; and the difference was statistically significant (P < 0.05). Furthermore, correlation analysis results indicated that the demethylation levels of the MMP-9 gene promoter region was positively correlated with serum protein levels, urinary albumin to creatinine ratio (UACR), urea and creatinine; and was negatively correlated with GFR. The demethylation of the MMP-9 gene promoter region may be involved in the occurrence and development of diabetic nephropathy by regulating the expression of MMP-9 protein in serum.

Prostanoid-induced expression of matrix metalloproteinase-1 messenger ribonucleic acid in rat osteosarcoma cells

NASA Technical Reports Server (NTRS)

Clohisy, J. C.; Connolly, T. J.; Bergman, K. D.; Quinn, C. O.; Partridge, N. C.

1994-01-01

Individual prostanoids have distinct potencies in activating intracellular signaling pathways and regulating gene expression in osteoblastic cells. The E-series prostaglandins (PGs) are known to stimulate matrix metalloproteinase-1 (MMP-1) synthesis and secretion in certain rodent and human osteoblastic cells, yet the intracellular events involved remain unclear. To further characterize this response and its signal transduction pathway(s), we examined prostanoid-induced expression of the MMP-1 gene in the rat osteoblastic osteosarcoma cell line UMR 106-01. Northern blot analysis demonstrated that prostaglandin E2 (PGE2) and PGE1 were very potent stimulators (40-fold) of MMP-1 transcript abundance, PGF2 alpha and prostacyclin were weak stimulators (4-fold), and thromboxane-B2 had no effect. The marked increase in MMP-1 transcript abundance after PGE2 treatment was first detected at 2 h, became maximal at 4 h, and persisted beyond 24 h. This response was dose dependent and elicited maximal and half-maximal effects with concentrations of 10(-6) and 0.6 x 10(-7) M, respectively. Cycloheximide, a protein synthesis inhibitor, completely blocked this effect of PGE2, suggesting that the expression of other genes is required. Nuclear run-on experiments demonstrated that PGE2 rapidly activates MMP-1 gene transcription, with a maximal increase at 2-4 h. The second messenger analog, 8-bromo-cAMP, mimicked the effects of PGE2 by stimulating a dose-dependent increase in MMP-1 messenger RNA (mRNA) levels, with a maximal effect quantitatively similar to that observed with PGE2. Thus, in UMR 106-01 cells, different prostanoids have distinct potencies in stimulating MMP-1 mRNA abundance. Our data suggest that PGE2 stimulation of MMP-1 synthesis is due to activation of MMP-1 gene transcription and a subsequent marked increase in MMP-1 mRNA abundance. This effect is dependent on de novo protein synthesis and is mimicked by protein kinase-A activation.

EGCG Inhibits Proliferation, Invasiveness and Tumor Growth by Up-Regulation of Adhesion Molecules, Suppression of Gelatinases Activity, and Induction of Apoptosis in Nasopharyngeal Carcinoma Cells

PubMed Central

Fang, Chih-Yeu; Wu, Chung-Chun; Hsu, Hui-Yu; Chuang, Hsin-Ying; Huang, Sheng-Yen; Tsai, Ching-Hwa; Chang, Yao; Tsao, George Sai-Wah; Chen, Chi-Long; Chen, Jen-Yang

2015-01-01

(−)-Epigallocatechin-3-gallate (EGCG), a major green tea polyphenol, has been shown to inhibit the proliferation of a variety of tumor cells. Epidemiological studies have shown that drinking green tea can reduce the incidence of nasopharyngeal carcinoma (NPC), yet the underlying mechanism is not well understood. In this study, the inhibitory effect of EGCG was tested on a set of Epstein Barr virus-negative and -positive NPC cell lines. Treatment with EGCG inhibited the proliferation of NPC cells but did not affect the growth of a non-malignant nasopharyngeal cell line, NP460hTert. Moreover, EGCG treated cells had reduced migration and invasive properties. The expression of the cell adhesion molecules E-cadherin and β-catenin was found to be up-regulated by EGCG treatment, while the down-regulation of matrix metalloproteinases (MMP)-2 and MMP-9 were found to be mediated by suppression of extracellular signal-regulated kinase (ERK) phosphorylation and AP-1 and Sp1 transactivation. Spheroid formation by NPC cells in suspension was significantly inhibited by EGCG. Oral administration of EGCG was capable of suppressing tumor growth in xenografted mice bearing NPC tumors. Treatment with EGCG was found to elevate the expression of p53 and p21, and eventually led to apoptosis of NPC cells via caspase 3 activation. The nuclear translocation of NF-κB and β-catenin was also suppressed by EGCG treatment. These results indicate that EGCG can inhibit the proliferation and invasiveness, and induce apoptosis, of NPC cells, making it a promising agent for chemoprevention or adjuvant therapy of NPC. PMID:25625511

Selective glucocorticoid receptor modulator compound A, in contrast to prednisolone, does not induce leptin or the leptin receptor in human osteoarthritis synovial fibroblasts.

PubMed

Malaise, Olivier; Relic, Biserka; Quesada-Calvo, Florence; Charlier, Edith; Zeddou, Mustapha; Neuville, Sophie; Gillet, Philippe; Louis, Edouard; de Seny, Dominique; Malaise, Michel G

2015-06-01

Glucocorticoids are powerful anti-inflammatory compounds that also induce the expression of leptin and leptin receptor (Ob-R) in synovial fibroblasts through TGF-βsignalling and Smad1/5 phosphorylation. Compound A (CpdA), a selective glucocorticoid receptor agonist, reduces inflammation in murine arthritis models and does not induce diabetes or osteoporosis, thus offering an improved risk:benefit ratio in comparison with glucocorticoids. Due to the detrimental role of leptin in OA pathogenesis, we sought to determine whether CpdA also induced leptin and Ob-R protein expression as observed with prednisolone. Human synovial fibroblasts and chondrocytes were isolated from the synovium and cartilage of OA patients after joint surgery. The cells were treated with prednisolone, TGF-β1, TNF-α and/or CpdA. Levels of leptin, IL-6, IL-8, MMP-1 and MMP-3 were measured by ELISA and expression levels of Ob-R phospho-Smad1/5, phospho-Smad2, α-tubulin and glyceraldehyde 3-phosphate dehydrogenase were analysed by western blotting. CpdA, unlike prednisolone, did not induce leptin secretion or Ob-R protein expression in OA synovial fibroblasts. Moreover, CpdA decreased endogenous Ob-R expression and down-regulated prednisolone-induced leptin secretion and Ob-R expression. Mechanistically, CpdA, unlike prednisolone, did not induce Smad1/5 phosphorylation. CpdA, similarly to prednisolone, down-regulated endogenous and TNF-α-induced IL-6, IL-8, MMP-1 and MMP-3 protein secretion. The dissociative effect of CpdA was confirmed using chondrocytes with no induction of leptin secretion, but with a significant decrease in IL-6, IL-8, MMP-1 and MMP-3 protein secretion. CpdA, unlike prednisolone, did not induce leptin or Ob-R in human OA synovial fibroblasts, thereby demonstrating an improved risk:benefit ratio. © The Author 2014. Published by Oxford University Press on behalf of the British Society for Rheumatology. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

Expression of most matrix metalloproteinase family members in breast cancer represents a tumor-induced host response.

PubMed Central

Heppner, K. J.; Matrisian, L. M.; Jensen, R. A.; Rodgers, W. H.

1996-01-01

Matrix metalloproteinase (MMP) family members have been associated with advanced-stage cancer and contribute to tumor progression, invasion, and metastasis as determined by inhibitor studies. In situ hybridization was performed to analyze the expression and localization of all known MMPs in a series of human breast cancer biopsy specimens. Most MMPs were localized to tumor stroma, and all MMPs had very distinct expression patterns. Matrilysin was expressed by morphologically normal epithelial ducts within tumors and in tissue from reduction mammoplasties, and by epithelial-derived tumor cells. Many family members, including stromelysin-3, gelatinase A, MT-MMP, interstitial collagenase, and stromelysin-1 were localized to fibroblasts of tumor stroma of invasive cancers but in quite distinct, and generally widespread, patterns. Gelatinase B, collagenase-3, and metalloelastase expression were more focal; gelatinase B was primarily localized to endothelial cells, collagenase-3 to isolated tumor cells, and metalloelastase to cytokeratin-negative, macrophage-like cells. The MMP inhibitor, TIMP-1, was expressed in both stromal and tumor components in most tumors, and neither stromelysin-2 nor neutrophil collagenase were detected in any of the tumors. These results indicate that there is very tight and complex regulation in the expression of MMP family members in breast cancer that generally represents a host response to the tumor and emphasize the need to further evaluate differential functions for MMP family members in breast tumor progression. Images Figure 1 Figure 2 Figure 3 PMID:8686751

Rheumatoid Factor Positivity Is Associated with Increased Joint Destruction and Upregulation of Matrix Metalloproteinase 9 and Cathepsin K Gene Expression in the Peripheral Blood in Rheumatoid Arthritic Patients Treated with Methotrexate

PubMed Central

Tchetina, Elena V.; Demidova, Natalia V.; Karateev, Dmitry E.; Nasonov, Eugeny L.

2013-01-01

We evaluated changes in gene expression of mTOR, p21, caspase-3, ULK1, TNFα, matrix metalloproteinase (MMP)-9, and cathepsin K in the whole blood of rheumatoid arthritic (RA) patients treated with methotrexate (MTX) in relation to their rheumatoid factor status, clinical, immunological, and radiological parameters, and therapeutic response after a 24-month follow-up. The study group consisted of 35 control subjects and 33 RA patients without previous history of MTX treatment. Gene expression was measured using real-time RT-PCR. Decreased disease activity in patients at the end of the study was associated with significant downregulation of TNFα expression. Downregulation of mTOR was observed in seronegative patients, while no significant changes in the expression of p21, ULK1, or caspase-3 were noted in any RA patients at the end of the study. The increase in erosion numbers observed in the seropositive patients at the end of the follow-up was accompanied by upregulation of MMP-9 and cathepsin K, while seronegative patients demonstrated an absence of significant changes in MMP-9 and cathepsin K expression and no increase in the erosion score. Our results suggest that increased expression of MMP-9 and cathepsin K genes in the peripheral blood might indicate higher bone tissue destruction activity in RA patients treated with methotrexate. The clinical study registration number is 0120.0810610. PMID:24348567

[IL-23 promotes invasion of esophageal squamous cell carcinoma cells by activating DLL4/Notch1 signaling pathway].

PubMed

Li, Wei; Zhou, Yuepeng; Su, Yuting; Ouyang, Yibo; Xie, Xiaodong; Wu, Yingying; Mao, Chaoming; Chen, Deyu

2015-06-01

To investigate the role of interlukin-23 (IL-23) in the invasion of human esophageal squamous cell carcinoma (ESCC) cells and the related mechanism. IL-23 expression in tumor and adjacent tissues from 10 ESCC patients were detected by immunohistochemistry. Real-time fluorescent PCR was used to examine the expressions of Notch1 and Foxn4 mRNAs in different concentration IL-23-treated TE-1 cells. After Notch pathway was blocked with γ-secretase inhibitor DAPT, expressions of Notch intracellular domain (NICD), Delta-like 4 (DLL4), hairy enhancer of split 1 (Hes1), matrix metalloproteinase 9 (MMP-9) in IL-23-treated TE-1 cells were measured by Western blotting. And the migration of IL-23-treated TE-1 cells was studied by TranswellTM migration assay. Compared with adjacent tissues, IL-23 was highly expressed in ESCC tissues. IL-23 treatment up-regulated significantly the expressions of NICD, DLL4, Hes1 and MMP-9 in TE-1 cells. The blockade of Notch1 pathway inhibited the expressions induced by IL-23. Migration assay revealed that IL-23 treatment significantly enhanced the migration of TE-1 cells. IL-23 could promote migration of human ESCC cells by activating DLL4/Notch1 signaling pathway.

Recanalization and flow regulate venous thrombus resolution and Matrix metalloproteinases expression in vivo

PubMed Central

Chabasse, Christine; Siefert, Suzanne A.; Chaudry, Mohammed; Hoofnagle, Mark H.; Lal, Brajesh K.; Sarkar, Rajabrata

2016-01-01

Objective We examined the role of thrombus recanalization and ongoing blood flow in the process of thrombus resolution by comparing two murine in vivo models of deep venous thrombosis. Design of study In CD1 mice, we performed surgical inferior vena cava (IVC) ligation (stasis thrombosis), stenosis (thrombosis with recanalization) or sham procedure. We analyzed thrombus weight over time as a measure of thrombus resolution, and quantified the mRNA and protein levels of Membrane-Type Matrix Metalloproteinases (MT-MMPs) as well as effectors of the plasmin complex at day 4, 8 and 12 post-surgery. Results Despite similar initial thrombus size, the presence of ongoing blood flow (stenosis model) was associated with a 45.91% subsequent improvement in thrombus resolution at day 8, and 12.57% at day 12, as compared with stasis thrombosis (ligation model). Immunoblot and real-time PCR demonstrated a difference in MMP-2 and MMP-9 activity at day 8 between the two models (P=.03 and P=.006 respectively), as well as a difference in MT2-MMP gene expression at day 8 (P=.044) and day 12 (P=0.03) and MT1-MMP protein expression at day 4 (P=.021). Histological analyses revealed distinct areas of recanalization in the thrombi of the stenosis model compared to the ligation model, as well as the recruitment of inflammatory cells, especially macrophages, and a focal pattern of localized expression of MT1-MMP and MT3-MMP proteins surrounding the areas of recanalization in the stenosis model. Conclusions Recanalization and ongoing blood flow accelerate deep venous thrombus resolution in vivo, and are associated with distinct patterns of MT1- and MT3-MMP expression and macrophages localization in areas of intra-thrombus recanalization. PMID:26993683

Angiotensin receptors as sensitive markers of acute bronchiole injury after lung transplantation.

PubMed

Nataatmadja, Maria; Passmore, Margaret; Russell, Fraser D; Prabowo, Sulistiana; Corley, Amanda; Fraser, John F

2014-08-01

Although lung transplantation is the only means of survival for patients with end-stage pulmonary disease, outcomes from this intervention are inferior to other solid organ transplants. The reason for the poor outcomes may be linked to an early reaction, such as primary graft dysfunction, and associated with marked inflammatory response, bronchiole injury, and later fibrotic responses. Mediators regulating these effects include angiotensin II and matrix metalloproteinases (MMPs). We investigated changes to these mediators over the course of cardiopulmonary bypass (CPB) and up to 72 h after lung transplantation, using immunohistochemistry, Western blot, and ELISA techniques. We found 4- and 16-fold increases in plasma angiotensin II and MMP-9, respectively, from pre-CPB to post-CPB. MMP-9 levels remained elevated 1 h after transplantation. MMP-2 levels were elevated 6-24 h after lung transplantation. Type 2 angiotensin II receptor (ATR2) expression was 3.5-fold higher in bronchoalveolar cells 1-6 h after transplantation than in controls. The study suggests that the combination of cardiopulmonary bypass and lung transplantation is associated with early changes in the angiotensin II receptor system and in MMPs, and that altered expression of these mediators may be a useful marker to examine pathological changes that occur in lungs during transplant surgery.

TGF-beta1 modulates matrix metalloproteinase-13 expression in hepatic stellate cells by complex mechanisms involving p38MAPK, PI3-kinase, AKT, and p70S6k.

PubMed

Lechuga, Carmen G; Hernández-Nazara, Zamira H; Domínguez Rosales, José-Alfredo; Morris, Elena R; Rincón, Ana Rosa; Rivas-Estilla, Ana María; Esteban-Gamboa, Andrés; Rojkind, Marcos

2004-11-01

Transforming growth factor-beta1 (TGF-beta1), the main cytokine involved in liver fibrogenesis, induces expression of the type I collagen genes in hepatic stellate cells by a transcriptional mechanism, which is hydrogen peroxide and de novo protein synthesis dependent. Our recent studies have revealed that expression of type I collagen and matrix metalloproteinase-13 (MMP-13) mRNAs in hepatic stellate cells is reciprocally modulated. Because TGF-beta1 induces a transient elevation of alpha1(I) collagen mRNA, we investigated whether this cytokine was able to induce the expression of MMP-13 mRNA during the downfall of the alpha1(I) collagen mRNA. In the present study, we report that TGF-beta1 induces a rapid decline in steady-state levels of MMP-13 mRNA at the time that it induces the expression of alpha1(I) collagen mRNA. This change in MMP-13 mRNA expression occurs within the first 6 h postcytokine administration and is accompanied by a twofold increase in gene transcription and a fivefold decrease in mRNA half-life. This is followed by increased expression of MMP-13 mRNA, which reaches maximal values by 48 h. Our results also show that this TGF-beta1-mediated effect is de novo protein synthesis-dependent and requires the activity of p38MAPK, phosphatidylinositol 3-kinase, AKT, and p70(S6k). Altogether, our data suggest that regulation of MMP-13 by TGF-beta1 is a complex process involving transcriptional and posttranscriptional mechanisms.

Osthole regulates TGF-β1 and MMP-2/9 expressions via activation of PPARα/γ in cultured mouse cardiac fibroblasts stimulated with angiotensin II.

PubMed

Chen, Rong; Xue, Jie; Xie, Meilin

2013-01-01

Our previous studies have demonstrated that osthole, an active constituent isolated from the fruit of Cnidium monnieri (L.) Cusson, can prevent isoprenaline-induced myocardial fibrosis in mice, but the underlying mechanism is still unclear. The mouse cardiac fibroblasts (CFs) stimulated with angiotensin II (Ang II) were cultured and treated with different concentrations of osthole. The mRNA expressions of peroxisome proliferator-activated receptor (PPAR) α/γ, transforming growth factor β1 (TGF-β1), and matrix metalloproteinase (MMP)-2/9 were detected by reverse transcription polymerase chain reaction method, and the protein expressions of nuclear factor-κB (NF-κB) and TGF-β1 were detected by Western blot method, respectively. Following treatment of cells with osthole at 2.5, 5, 10 and 20 μg/mL, the NF-κB and TGF-β1 expressions were dose-dependently decreased, while the expressions of PPARα/γ and MMP-2/9 were dose-dependently increased. After the cells were preincubated with PPARα antagonist (MK886) or/and PPARγ antagonist (GW9662), the inhibitions of osthole on the NF-κB and TGF-β1 expressions were decreased or completely halted and the increment of osthole on the MMP-2/9 expressions were also decreased or completely cancelled. Osthole could inhibit the NF-κB and TGF-β1 expressions by activation of PPARα/γ, and subsequently enhance the MMP-2/9 expressions in cultured CFs, and these effects of osthole may play the beneficial roles in the prevention and treatment of myocardial fibrosis.

Ellagitannins of the fruit rind of pomegranate (Punica granatum) antagonize in vitro the host inflammatory response mechanisms involved in the onset of malaria.

PubMed

Dell'agli, Mario; Galli, Germana V; Bulgari, Michela; Basilico, Nicoletta; Romeo, Sergio; Bhattacharya, Deepak; Taramelli, Donatella; Bosisio, Enrica

2010-07-19

The sun-dried rind of the immature fruit of pomegranate (Punica granatum) is presently used as a herbal formulation (OMARIA, Orissa Malaria Research Indigenous Attempt) in Orissa, India, for the therapy and prophylaxis of malaria. The pathogenesis of cerebral malaria, a complication of the infection by Plasmodium falciparum, is an inflammatory cytokine-driven disease associated to an up-regulation and activity of metalloproteinase-9 and to the increase of TNF production. The in vitro anti-plasmodial activity of Punica granatum (Pg) was recently described. The aim of the present study was to explore whether the anti-malarial effect of OMARIA could also be sustained via other mechanisms among those associated to the host immune response. From the methanolic extract of the fruit rind, a fraction enriched in tannins (Pg-FET) was prepared. MMP-9 secretion and expression were evaluated in THP-1 cells stimulated with haemozoin or TNF. The assays were conducted in the presence of the Pg-FET and its chemical constituents ellagic acid and punicalagin. The effect of urolithins, the ellagitannin metabolites formed by human intestinal microflora, was also investigated. Pg-FET and its constituents inhibited the secretion of MMP-9 induced by haemozoin or TNF. The effect occurred at transcriptional level since MMP-9 mRNA levels were lower in the presence of the tested compounds. Urolithins as well inhibited MMP-9 secretion and expression. Pg-FET and pure compounds also inhibited MMP-9 promoter activity and NF-kB-driven transcription. The beneficial effect of the fruit rind of Punica granatum for the treatment of malarial disease may be attributed to the anti-parasitic activity and the inhibition of the pro-inflammatory mechanisms involved in the onset of cerebral malaria.

HSPA6 augments garlic extract-induced inhibition of proliferation, migration, and invasion of bladder cancer EJ cells; Implication for cell cycle dysregulation, signaling pathway alteration, and transcription factor-associated MMP-9 regulation

PubMed Central

Hwang, Byungdoo; Noh, Dae-Hwa; Park, Sung Lyea; Kim, Won Tae; Park, Sung-Soo; Kim, Wun-Jae; Moon, Sung-Kwon

2017-01-01

Although recent studies have demonstrated the anti-tumor effects of garlic extract (GE), the exact molecular mechanism is still unclear. In this study, we investigated the molecular mechanism associated with the inhibitory action of GE against bladder cancer EJ cell responses. Treatment with GE significantly inhibited proliferation of EJ cells dose-dependently through G2/M-phase cell cycle arrest. This G2/M-phase cell cycle arrest by GE was due to the activation of ATM and CHK2, which appears to inhibit phosphorylation of Cdc25C (Ser216) and Cdc2 (Thr14/Tyr15), this in turn was accompanied by down-regulation of cyclin B1 and up-regulation of p21WAF1. Furthermore, GE treatment was also found to induce phosphorylation of MAPK (ERK1/2, p38MAPK, and JNK) and AKT. In addition, GE impeded the migration and invasion of EJ cells via inhibition of MMP-9 expression followed by decreased binding activities of AP-1, Sp-1, and NF-κB motifs. Based on microarray datasets, we selected Heat shock protein A6 (HSPA6) as the most up-regulated gene responsible for the inhibitory effects of GE. Interestingly, overexpression of HSPA6 gene resulted in an augmentation effect with GE inhibiting proliferation, migration, and invasion of EJ cells. The augmentation effect of HSPA6 was verified by enhancing the induction of G2/M-phase-mediated ATM-CHK2-Cdc25C-p21WAF1-Cdc2 cascade, phosphorylation of MAPK and AKT signaling, and suppression of transcription factor-associated MMP-9 regulation in response to GE in EJ cells. Overall, our novel results indicate that HSPA6 reinforces the GE-mediated inhibitory effects of proliferation, migration, and invasion of EJ cells and may provide a new approach for therapeutic treatment of malignancies. PMID:28187175

Tumor necrosis factor-{alpha} induces MMP-9 expression via p42/p44 MAPK, JNK, and nuclear factor-{kappa}B in A549 cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lin, C.-C.; Tseng, Hsiao-Wei; Hsieh, Hsi-Lung

2008-06-15

Matrix metalloproteinases (MMPs), in particular MMP-9, have been shown to be induced by cytokines including tumor necrosis factor-{alpha} (TNF-{alpha}) and contributes to airway inflammation. However, the mechanisms underlying MMP-9 expression induced by TNF-{alpha} in human A549 cells remain unclear. Here, we showed that TNF-{alpha} induced production of MMP-9 protein and mRNA is determined by zymographic, Western blotting, RT-PCR and ELISA assay, which were attenuated by inhibitors of MEK1/2 (U0126), JNK (SP600125), and NF-{kappa}B (helenalin), and transfection with dominant negative mutants of ERK2 ({delta}ERK) and JNK ({delta}JNK), and siRNAs for MEK1, p42 and JNK2. TNF-{alpha}-stimulated phosphorylation of p42/p44 MAPK and JNKmore » were attenuated by pretreatment with the inhibitors U0126 and SP600125 or transfection with dominant negative mutants of {delta}ERK and {delta}JNK. Furthermore, the involvement of NF-{kappa}B in TNF-{alpha}-induced MMP-9 production was consistent with that TNF-{alpha}-stimulated degradation of I{kappa}B-{alpha} and translocation of NF-{kappa}B into the nucleus which were blocked by helenalin, but not by U0126 and SP600125, revealed by immunofluorescence staining. The regulation of MMP-9 gene transcription by MAPKs and NF-{kappa}B was further confirmed by gene luciferase activity assay. MMP-9 promoter activity was enhanced by TNF-{alpha} in A549 cells transfected with wild-type MMP-9-Luc, which was inhibited by helenalin, U0126, or SP600125. In contrast, TNF-{alpha}-stimulated MMP-9 luciferase activity was totally lost in cells transfected with mutant-NF-{kappa}B MMP-9-luc. Moreover, pretreatment with actinomycin D and cycloheximide attenuated TNF-{alpha}-induced MMP-9 expression. These results suggest that in A549 cells, phosphorylation of p42/p44 MAPK, JNK, and transactivation of NF-{kappa}B are essential for TNF-{alpha}-induced MMP-9 gene expression.« less

Phenotype anchoring in zebrafish reveals a potential role for matrix metalloproteinases (MMPs) in tamoxifen's effects on skin epithelium

DOE Office of Scientific and Technical Information (OSTI.GOV)

Bugel, Sean M., E-mail: Sean.Bugel@oregonstate.edu; Wehmas, Leah C., E-mail: wehmasl@onid.oregonstate.edu; La Du, Jane K., E-mail: Jane.LaDu@oregonstate.edu

The zebrafish is a powerful alternative model used to link phenotypes with molecular effects to discover drug mode of action. Using a zebrafish embryo-larval toxicity bioassay, we evaluated the effects of tamoxifen — a widely used anti-estrogen chemotherapeutic. Zebrafish exposed to ≥ 10 μM tamoxifen exhibited a unique necrotic caudal fin phenotype that was rapidly induced regardless of developmental life-stage when treatment was applied. To define tamoxifen's bioactivity resulting in this phenotype, targeted gene expression was used to evaluate 100 transcripts involved in tissue remodeling, calcium signaling, cell cycle and cell death, growth factors, angiogenesis and hypoxia. The most robustlymore » misregulated transcripts in the tail were matrix metalloproteinases mmp9 and mmp13a, induced 127 and 1145 fold, respectively. Expression of c-fos, c-jun, and ap1s1 were also moderately elevated (3–7 fold), consistent with AP-1 activity — a transcription factor that regulates MMP expression. Immunohistochemistry confirmed high levels of induction for MMP13a in affected caudal fin skin epithelial tissue. The necrotic caudal fin phenotype was significantly attenuated or prevented by three functionally unique MMP inhibitors: EDTA (metal chelator), GM 6001 (broad MMP inhibitor), and SR 11302 (AP-1 transcription factor inhibitor), suggesting MMP-dependence. SR 11302 also inhibited induction of mmp9, mmp13a, and a putative MMP target, igfbp1a. Overall, our studies suggest that tamoxifen's effect is the result of perturbation of the MMP system in the skin leading to ectopic expression, cytotoxicity, and the necrotic caudal fin phenotype. These studies help advance our understanding of tamoxifen's non-classical mode of action and implicate a possible role for MMPs in tissues such as skin. - Highlights: • Tamoxifen rapidly induced a unique necrotic caudal fin phenotype in zebrafish. • Apoptosis co-localized temporally and spatially in the necrotic tail. • The necrotic fin phenotype was p53, GPER and ER independent. • The necrotic fin phenotype was dependent on ectopic MMP induction and activity in the skin. • The necrotic fin phenotype occurred at concentrations exceeding anti-estrogenic effects.« less

Protein arginine methyltransferase 7 promotes breast cancer cell invasion through the induction of MMP9 expression

PubMed Central

Baldwin, R. Mitchell; Haghandish, Nasim; Daneshmand, Manijeh; Amin, Shahrier; Paris, Geneviève; Falls, Theresa J.; Bell, John C.; Islam, Shahidul; Côté, Jocelyn

2015-01-01

Recent evidence points to the protein arginine methyltransferase (PRMT) family of enzymes playing critical roles in cancer. PRMT7 has been identified in several gene expression studies to be associated with increased metastasis and decreased survival in breast cancer patients. However, this has not been extensively studied. Here we report that PRMT7 expression is significantly upregulated in both primary breast tumour tissues and in breast cancer lymph node metastases. We have demonstrated that reducing PRMT7 levels in invasive breast cancer cells using RNA interference significantly decreased cell invasion in vitro and metastasis in vivo. Conversely, overexpression of PRMT7 in non-aggressive MCF7 cells enhanced their invasiveness. Furthermore, we show that PRMT7 induces the expression of matrix metalloproteinase 9 (MMP9), a well-known mediator of breast cancer metastasis. Importantly, we significantly rescued invasion of aggressive breast cancer cells depleted of PRMT7 by the exogenous expression of MMP9. Our results demonstrate that upregulation of PRMT7 in breast cancer may have a significant role in promoting cell invasion through the regulation of MMP9. This identifies PRMT7 as a novel and potentially significant biomarker and therapeutic target for breast cancer. PMID:25605249

Protein arginine methyltransferase 7 promotes breast cancer cell invasion through the induction of MMP9 expression.

PubMed

Baldwin, R Mitchell; Haghandish, Nasim; Daneshmand, Manijeh; Amin, Shahrier; Paris, Geneviève; Falls, Theresa J; Bell, John C; Islam, Shahidul; Côté, Jocelyn

2015-02-20

Recent evidence points to the protein arginine methyltransferase (PRMT) family of enzymes playing critical roles in cancer. PRMT7 has been identified in several gene expression studies to be associated with increased metastasis and decreased survival in breast cancer patients. However, this has not been extensively studied. Here we report that PRMT7 expression is significantly upregulated in both primary breast tumour tissues and in breast cancer lymph node metastases. We have demonstrated that reducing PRMT7 levels in invasive breast cancer cells using RNA interference significantly decreased cell invasion in vitro and metastasis in vivo. Conversely, overexpression of PRMT7 in non-aggressive MCF7 cells enhanced their invasiveness. Furthermore, we show that PRMT7 induces the expression of matrix metalloproteinase 9 (MMP9), a well-known mediator of breast cancer metastasis. Importantly, we significantly rescued invasion of aggressive breast cancer cells depleted of PRMT7 by the exogenous expression of MMP9. Our results demonstrate that upregulation of PRMT7 in breast cancer may have a significant role in promoting cell invasion through the regulation of MMP9. This identifies PRMT7 as a novel and potentially significant biomarker and therapeutic target for breast cancer.

Role of PTPα in the Destruction of Periodontal Connective Tissues

PubMed Central

Rajshankar, Dhaarmini; Sima, Corneliu; Wang, Qin; Goldberg, Stephanie R.; Kazembe, Mwayi; Wang, Yongqiang; Glogauer, Michael; Downey, Gregory P.; McCulloch, Christopher A.

2013-01-01

IL-1β contributes to connective tissue destruction in part by up-regulating stromelysin-1 (MMP-3), which in fibroblasts is a focal adhesion-dependent process. Protein tyrosine phosphatase-α (PTPα) is enriched in and regulates the formation of focal adhesions, but the role of PTPα in connective tissue destruction is not defined. We first examined destruction of periodontal connective tissues in adult PTPα+/+ and PTPα−/− mice subjected to ligature-induced periodontitis, which increases the levels of multiple cytokines, including IL-1β. Three weeks after ligation, maxillae were processed for morphometry, micro-computed tomography and histomorphometry. Compared with unligated controls, there was ∼1.5–3 times greater bone loss as well as 3-fold reduction of the thickness of the gingival lamina propria and 20-fold reduction of the amount of collagen fibers in WT than PTPα−/− mice. Immunohistochemical staining of periodontal tissue showed elevated expression of MMP-3 at ligated sites. Second, to examine mechanisms by which PTPα may regulate matrix degradation, human MMP arrays were used to screen conditioned media from human gingival fibroblasts treated with vehicle, IL-1β or TNFα. Although MMP-3 was upregulated by both cytokines, only IL-1β stimulated ERK activation in human gingival fibroblasts plated on fibronectin. TIRF microscopy and immunoblotting analyses of cells depleted of PTPα activity with the use of various mutated constructs or with siRNA or PTPαKO and matched wild type fibroblasts were plated on fibronectin to enable focal adhesion formation and stimulated with IL-1β. These data showed that the catalytic and adaptor functions of PTPα were required for IL-1β-induced focal adhesion formation, ERK activation and MMP-3 release. We conclude that inflammation-induced connective tissue degradation involving fibroblasts requires functionally active PTPα and in part is mediated by IL-1β signaling through focal adhesions. PMID:23940616

REV-ERBs agonism suppresses osteoclastogenesis and prevents ovariectomy-induced bone loss partially via FABP4 upregulation.

PubMed

Song, Chao; Tan, Peng; Zhang, Zheng; Wu, Wei; Dong, Yonghui; Zhao, Liming; Liu, Huiyong; Guan, Hanfeng; Li, Feng

2018-01-22

REV-ERBs (REV-ERBα and REV-ERBβ) are transcription repressors and circadian regulators. Previous investigations have shown that REV-ERBs repress the expression of target genes, including MMP9 and CX3CR1, in macrophages. Because MMP9 and CX3CR1 reportedly participate in receptor activator of nuclear factor-κB ligand (RANKL)-induced osteoclastogenesis, we inferred that REV-ERBs might play a role in osteoclastogenesis. In the present study, we found that the REV-ERBα level decreased significantly during RANKL-induced osteoclast differentiation from primary bone marrow-derived macrophages (BMMs). REV-ERBα knockdown by small interfering RNA in BMMs resulted in the enhanced formation of osteoclasts, whereas REV-ERBβ knockdown showed no effect on osteoclast differentiation. Moreover, the REV-ERB agonist SR9009 inhibited osteoclast differentiation and bone resorption. Intraperitoneal SR9009 administration prevented ovariectomy-induced bone loss; this effect was accompanied by decreased serum RANKL and C-terminal telopeptide of type I collagen levels and increased osteoprotegerin levels. Further investigation revealed that NF-κB and MAPK activation and nuclear factor of activated T cells, cytoplasmic 1, and c-fos expression were suppressed by SR9009. The level of reactive oxygen species was also decreased by SR9009, with NADPH oxidase subunits also being down-regulated. In addition, an expression microarray showed that FABP4, an intracellular lipid-binding protein, was up-regulated by REV-ERB agonism. BMS309403, an inhibitor of FABP4, partially prevented the suppression of osteoclastogenesis by SR9009 through stabilizing phosphorylation of p65. To summarize, our results proved that the REV-ERB agonism inhibited osteoclastogenesis partially via FABP4 up-regulation.-Song, C., Tan, P., Zhang, Z., Wu, W., Dong, Y., Zhao, L., Liu, H., Guan, H., Li, F. REV-ERBs agonism suppresses osteoclastogenesis and prevents ovariectomy-induced bone loss partially via FABP4 upregulation.

Vitamin D Impacts the Expression of Runx2 Target Genes and Modulates Inflammation, Oxidative Stress and Membrane Vesicle Biogenesis Gene Networks in 143B Osteosarcoma Cells

PubMed Central

Garimella, Rama; Tadikonda, Priyanka; Tawfik, Ossama; Gunewardena, Sumedha; Rowe, Peter; Van Veldhuizen, Peter

2017-01-01

Osteosarcoma (OS) is an aggressive malignancy of bone affecting children, adolescents and young adults. Understanding vitamin D metabolism and vitamin D regulated genes in OS is an important aspect of vitamin D/cancer paradigm, and in evaluating vitamin D as adjuvant therapy for human OS. Vitamin D treatment of 143B OS cells induced significant and novel changes in the expression of genes that regulate: (a) inflammation and immunity; (b) formation of reactive oxygen species, metabolism of cyclic nucleotides, sterols, vitamins and mineral (calcium), quantity of gap junctions and skeletogenesis; (c) bone mineral density; and (d) cell viability of skeletal cells, aggregation of bone cancer cells and exocytosis of secretory vesicles. Ingenuity pathway analysis revealed significant reduction in Runx2 target genes such as fibroblast growth factor -1, -12 (FGF1 and FGF12), bone morphogenetic factor-1 (BMP1), SWI/SNF related, matrix associated actin dependent regulator of chromatin subfamily a, member 4 (SMARCA4), Matrix extracellular phosphoglycoprotein (MEPE), Integrin, β4 (ITGBP4), Matrix Metalloproteinase -1, -28 (MMP1 and MMP28), and signal transducer and activator of transcription-4 (STAT4) in vitamin D treated 143B OS cells. These genes interact with the inflammation, oxidative stress and membrane vesicle biogenesis gene networks. Vitamin D not only inhibited the expression of Runx2 target genes MMP1, MMP28 and kallikrein related peptidase-7 (KLK7), but also migration and invasion of 143B OS cells. Vitamin D regulated Runx2 target genes or their products represent potential therapeutic targets and laboratory biomarkers for applications in translational oncology. PMID:28300755

Vitamin D Impacts the Expression of Runx2 Target Genes and Modulates Inflammation, Oxidative Stress and Membrane Vesicle Biogenesis Gene Networks in 143B Osteosarcoma Cells.

PubMed

Garimella, Rama; Tadikonda, Priyanka; Tawfik, Ossama; Gunewardena, Sumedha; Rowe, Peter; Van Veldhuizen, Peter

2017-03-16

Osteosarcoma (OS) is an aggressive malignancy of bone affecting children, adolescents and young adults. Understanding vitamin D metabolism and vitamin D regulated genes in OS is an important aspect of vitamin D/cancer paradigm, and in evaluating vitamin D as adjuvant therapy for human OS. Vitamin D treatment of 143B OS cells induced significant and novel changes in the expression of genes that regulate: (a) inflammation and immunity; (b) formation of reactive oxygen species, metabolism of cyclic nucleotides, sterols, vitamins and mineral (calcium), quantity of gap junctions and skeletogenesis; (c) bone mineral density; and (d) cell viability of skeletal cells, aggregation of bone cancer cells and exocytosis of secretory vesicles. Ingenuity pathway analysis revealed significant reduction in Runx2 target genes such as fibroblast growth factor -1, -12 ( FGF1 and FGF12 ), bone morphogenetic factor-1 ( BMP1 ), SWI/SNF related, matrix associated actin dependent regulator of chromatin subfamily a, member 4 ( SMARCA4 ), Matrix extracellular phosphoglycoprotein ( MEPE ), Integrin, β4 ( ITGBP4 ), Matrix Metalloproteinase -1, -28 ( MMP1 and MMP28 ), and signal transducer and activator of transcription-4 ( STAT4 ) in vitamin D treated 143B OS cells. These genes interact with the inflammation, oxidative stress and membrane vesicle biogenesis gene networks. Vitamin D not only inhibited the expression of Runx2 target genes MMP1 , MMP28 and kallikrein related peptidase-7 ( KLK7 ), but also migration and invasion of 143B OS cells. Vitamin D regulated Runx2 target genes or their products represent potential therapeutic targets and laboratory biomarkers for applications in translational oncology.

Inflammation increases NOTCH1 activity via MMP9 and is counteracted by Eicosapentaenoic Acid-free fatty acid in colon cancer cells

PubMed Central

Fazio, Chiara; Piazzi, Giulia; Vitaglione, Paola; Fogliano, Vincenzo; Munarini, Alessandra; Prossomariti, Anna; Milazzo, Maddalena; D’Angelo, Leonarda; Napolitano, Manuela; Chieco, Pasquale; Belluzzi, Andrea; Bazzoli, Franco; Ricciardiello, Luigi

2016-01-01

Aberrant NOTCH1 signalling is critically involved in multiple models of colorectal cancer (CRC) and a prominent role of NOTCH1 activity during inflammation has emerged. Epithelial to Mesenchymal Transition (EMT), a crucial event promoting malignant transformation, is regulated by inflammation and Metalloproteinase-9 (MMP9) plays an important role in this process. Eicosapentaenoic Acid (EPA), an omega-3 polyunsaturated fatty acid, was shown to prevent colonic tumors in different settings. We recently found that an extra-pure formulation of EPA as Free Fatty Acid (EPA-FFA) protects from colon cancer development in a mouse model of Colitis-Associated Cancer (CAC) through modulation of NOTCH1 signalling. In this study, we exposed colon cancer cells to an inflammatory stimulus represented by a cytokine-enriched Conditioned Medium (CM), obtained from THP1-differentiated macrophages. We found, for the first time, that CM strongly up-regulated NOTCH1 signalling and EMT markers, leading to increased invasiveness. Importantly, NOTCH1 signalling was dependent on MMP9 activity, upon CM exposure. We show that a non-cytotoxic pre-treatment with EPA-FFA antagonizes the effect of inflammation on NOTCH1 signalling, with reduction of MMP9 activity and invasiveness. In conclusion, our data suggest that, in CRC cells, inflammation induces NOTCH1 activity through MMP9 up-regulation and that this mechanism can be counteracted by EPA-FFA. PMID:26864323

Inflammation increases NOTCH1 activity via MMP9 and is counteracted by Eicosapentaenoic Acid-free fatty acid in colon cancer cells.

PubMed

Fazio, Chiara; Piazzi, Giulia; Vitaglione, Paola; Fogliano, Vincenzo; Munarini, Alessandra; Prossomariti, Anna; Milazzo, Maddalena; D'Angelo, Leonarda; Napolitano, Manuela; Chieco, Pasquale; Belluzzi, Andrea; Bazzoli, Franco; Ricciardiello, Luigi

2016-02-11

Aberrant NOTCH1 signalling is critically involved in multiple models of colorectal cancer (CRC) and a prominent role of NOTCH1 activity during inflammation has emerged. Epithelial to Mesenchymal Transition (EMT), a crucial event promoting malignant transformation, is regulated by inflammation and Metalloproteinase-9 (MMP9) plays an important role in this process. Eicosapentaenoic Acid (EPA), an omega-3 polyunsaturated fatty acid, was shown to prevent colonic tumors in different settings. We recently found that an extra-pure formulation of EPA as Free Fatty Acid (EPA-FFA) protects from colon cancer development in a mouse model of Colitis-Associated Cancer (CAC) through modulation of NOTCH1 signalling. In this study, we exposed colon cancer cells to an inflammatory stimulus represented by a cytokine-enriched Conditioned Medium (CM), obtained from THP1-differentiated macrophages. We found, for the first time, that CM strongly up-regulated NOTCH1 signalling and EMT markers, leading to increased invasiveness. Importantly, NOTCH1 signalling was dependent on MMP9 activity, upon CM exposure. We show that a non-cytotoxic pre-treatment with EPA-FFA antagonizes the effect of inflammation on NOTCH1 signalling, with reduction of MMP9 activity and invasiveness. In conclusion, our data suggest that, in CRC cells, inflammation induces NOTCH1 activity through MMP9 up-regulation and that this mechanism can be counteracted by EPA-FFA.

Activation of Peroxisome Proliferator-Activated Receptor Alpha Improves Aged and UV-Irradiated Skin by Catalase Induction

PubMed Central

Shin, Mi Hee; Lee, Se-Rah; Kim, Min-Kyoung; Shin, Chang-Yup

2016-01-01

Peroxisome proliferator-activated receptor alpha (PPARα) is a nuclear hormone receptor involved in the transcriptional regulation of lipid metabolism, fatty acid oxidation, and glucose homeostasis. Its activation stimulates antioxidant enzymes such as catalase, whose expression is decreased in aged human skin. Here we investigated the expression of PPARα in aged and ultraviolet (UV)-irradiated skin, and whether PPARα activation can modulate expressions of matrix metalloproteinase (MMP)-1 and procollagen through catalase regulation. We found that PPARα mRNA level was significantly decreased in intrinsically aged and photoaged human skin as well as in UV-irradiated skin. A PPARα activator, Wy14643, inhibited UV-induced increase of MMP-1 and decrease of procollagen expression and caused marked increase in catalase expression. Furthermore, production of reactive oxygen species (ROS) was suppressed by Wy14643 in UV-irradiated and aged dermal fibroblasts, suggesting that the PPARα activation-induced upregulation of catalase leads to scavenging of ROS produced due to UV irradiation or aging. PPARα knockdown decreased catalase expression and abolished the beneficial effects of Wy14643. Topical application of Wy14643 on hairless mice restored catalase activity and prevented MMP-13 and inflammatory responses in skin. Our findings indicate that PPARα activation triggers catalase expression and ROS scavenging, thereby protecting skin from UV-induced damage and intrinsic aging. PMID:27611371

Gab1 regulates proliferation and migration through the PI3K/Akt signaling pathway in intrahepatic cholangiocarcinoma.

PubMed

Sang, Haiquan; Li, Tingting; Li, Hangyu; Liu, Jingang

2015-11-01

Intrahepatic cholangiocarcinoma is the second most common primary malignant tumor of the liver, and it originates from the intrahepatic biliary duct epithelium. Prognosis is poor due to lack of effective comprehensive treatments. In this study, we assessed the expression of Gab1, VEGFR-2, and MMP-9 in intrahepatic cholangiocarcinoma solid tumors by immunohistochemistry and determined whether their expression was associated with clinical and pathological features. We found that expression of Gab1, VEGFR-2, and MMP-9 was highly and positively correlated with each other and with lymph node metastasis and TNM stage in intrahepatic cholangiocarcinoma tissues. Interference of Gab1 and VEGFR-2 expression via siRNA in the intrahepatic cholangiocarcinoma cell line RBE resulted in decreased PI3K/Akt pathway activity. Inhibition of Gab1 and VEGFR-2 expression also caused decreased cell proliferation, cell cycle arrested in G1 phase, increased apoptosis, and decreased invasion in RBE cells. These results suggest that Gab1, VEGFR-2, and MMP-9 contribute significantly to the highly malignant behavior of intrahepatic cholangiocarcinoma. The regulation of growth, apoptosis, and invasion by Gab1 through the VEGFR-2/Gab1/PI3K/Akt signaling pathway may represent potential targets for improving the treatment of intrahepatic cholangiocarcinoma.

CD36 Is a Matrix Metalloproteinase-9 Substrate That Stimulates Neutrophil Apoptosis and Removal During Cardiac Remodeling.

PubMed

DeLeon-Pennell, Kristine Y; Tian, Yuan; Zhang, Bai; Cates, Courtney A; Iyer, Rugmani Padmanabhan; Cannon, Presley; Shah, Punit; Aiyetan, Paul; Halade, Ganesh V; Ma, Yonggang; Flynn, Elizabeth; Zhang, Zhen; Jin, Yu-Fang; Zhang, Hui; Lindsey, Merry L

2016-02-01

After myocardial infarction, the left ventricle undergoes a wound healing response that includes the robust infiltration of neutrophils and macrophages to facilitate removal of dead myocytes as well as turnover of the extracellular matrix. Matrix metalloproteinase (MMP)-9 is a key enzyme that regulates post-myocardial infarction left ventricular remodeling. Infarct regions from wild-type and MMP-9 null mice (n=8 per group) analyzed by glycoproteomics showed that of 541 N-glycosylated proteins quantified, 45 proteins were at least 2-fold upregulated or downregulated with MMP-9 deletion (all P<0.05). Cartilage intermediate layer protein and platelet glycoprotein 4 (CD36) were identified as having the highest fold increase in MMP-9 null mice. By immunoblotting, CD36 but not cartilage intermediate layer protein decreased steadily during the time course post-myocardial infarction, which identified CD36 as a candidate MMP-9 substrate. MMP-9 was confirmed in vitro and in vivo to proteolytically degrade CD36. In vitro stimulation of day 7 post-myocardial infarction macrophages with MMP-9 or a CD36-blocking peptide reduced phagocytic capacity. Dual immunofluorescence revealed concomitant accumulation of apoptotic neutrophils in the MMP-9 null group compared with wild-type group. In vitro stimulation of isolated neutrophils with MMP-9 decreased neutrophil apoptosis, indicated by reduced caspase-9 expression. Our data reveal a new cell-signaling role for MMP-9 through CD36 degradation to regulate macrophage phagocytosis and neutrophil apoptosis. © 2015 American Heart Association, Inc.

Protective effect of Disporum sessile D.Don extract against UVB-induced photoaging via suppressing MMP-1 expression and collagen degradation in human skin cells.

PubMed

Mohamed, Mohamed Antar Aziz; Jung, Mira; Lee, Sang Min; Lee, Tae Hoon; Kim, Jiyoung

2014-04-05

In the present study, we report that Disporum sessile D.Don herbal extract (DDE) possesses anti-skin photoaging effect through inhibition of MMP-1 mRNA and protein expression levels and increase collagen production in UVB-irradiated human dermal fibroblast cells (NHDF). To delineate the molecular mechanism by which DDE inhibited MMP-1 expression, immortal human keratinocytes cells (HaCaT) have been used. We have found that DDE inhibited UVB-induced MMP-1 mRNA and protein expression levels in HaCaT cells through inhibition of UVB-induced activation of NF-κB in HaCaT cells. Inhibitors of NF-κB (Bay11-7082), and mitogen-activated protein kinases such as extracellular regulated kinase (PD98059), c-Jun N-terminal kinase (SP600125), and p38 (SB203580) suppressed expression of MMP-1, and phosphorylation of these signaling molecules were attenuated by DDE. DDE also inhibited phosphorylation of IKKα and IκBα, and reduced nuclear translocation of NF-κB. Our results also demonstrated that DDE inhibited NF-κB driven expression of luciferase reporter gene and the DNA binding of NF-κB to its cognate binding site in UV-irradiated cells. Therefore, these results strongly suggest that DDE can be utilized as a potential agent for prevention and treatment of skin photoaging. Copyright © 2014 Elsevier B.V. All rights reserved.

Matrix metalloproteinase-9 is up-regulated by CCL21/CCR7 interaction via extracellular signal-regulated kinase-1/2 signaling and is involved in CCL21-driven B-cell chronic lymphocytic leukemia cell invasion and migration.

PubMed

Redondo-Muñoz, Javier; José Terol, María; García-Marco, José A; García-Pardo, Angeles

2008-01-01

B-cell chronic lymphocytic leukemia (B-CLL) progression is frequently accompanied by clinical lymphadenopathy, and the CCL21 chemokine may play an important role in this process. Indeed, CCR7 (the CCL21 receptor), as well as matrix metalloproteinase-9 (MMP-9), are overexpressed in infiltrating B-CLL cells. We have studied whether MMP-9 is regulated by CCL21 and participates in CCL21-dependent migration. CCL21 significantly increased B-CLL MMP-9 production, measured by gelatin zymography. This was inhibited by blocking extracellular signal-regulated kinase-1/2 (ERK1/2) activity or by cell transfection with CCR7-siRNA. Accordingly, CCL21/CCR7 interaction activated the ERK1/2/c-Fos pathway and increased MMP-9 mRNA. CCL21-driven B-CLL cell migration through Matrigel or human umbilical vein endothelial cells (HUVEC) was blocked by anti-CCR7 antibodies, CCR7-siRNA transfection, or the ERK1/2 inhibitor U0126, as well as by anti-MMP-9 antibodies or tissue inhibitor of metalloproteinase 1 (TIMP-1). These results strongly suggest that MMP-9 is involved in B-CLL nodal infiltration and expand the roles of MMP-9 and CCR7 in B-CLL progression. Both molecules could thus constitute therapeutic targets for this disease.

Matrix Metalloproteinases in Normal Pregnancy and Preeclampsia

PubMed Central

Chen, Juanjuan; Khalil, Raouf A.

2017-01-01

Normal pregnancy is associated with marked hemodynamic and uterine changes that allow adequate uteroplacental blood flow and uterine expansion for the growing fetus. These pregnancy-associated changes involve significant uteroplacental and vascular remodeling. Matrix metalloproteinases (MMPs) are important regulators of vascular and uterine remodeling. Increases in MMP-2 and MMP-9 have been implicated in vasodilation, placentation and uterine expansion during normal pregnancy. The increases in MMPs could be induced by the increased production of estrogen and progesterone during pregnancy. MMP expression/activity may be altered during complications of pregnancy. Decreased vascular MMP-2 and MMP-9 may lead to decreased vasodilation, increased vasoconstriction, hypertensive pregnancy and preeclampsia. Abnormal expression of uteroplacental integrins, cytokines and MMPs may lead to decreased maternal tolerance, apoptosis of invasive trophoblast cells, inadequate remodeling of spiral arteries, and reduced uterine perfusion pressure (RUPP). RUPP may cause imbalance between the anti-angiogenic factors soluble fms-like tyrosine kinase-1 and soluble endoglin and the pro-angiogenic vascular endothelial growth factor and placental growth factor, or stimulate the release of inflammatory cytokines, hypoxia-inducible factor, reactive oxygen species, and angiotensin AT1 receptor agonistic autoantibodies. These circulating factors could target MMPs in the extracellular matrix as well as endothelial and vascular smooth muscle cells, causing generalized vascular dysfunction, increased vasoconstriction and hypertension in pregnancy. MMP activity can also be altered by endogenous tissue inhibitors of metalloproteinases (TIMPs) and changes in the MMP/TIMP ratio. In addition to their vascular effects, decreases in expression/activity of MMP-2 and MMP-9 in the uterus could impede uterine growth and expansion and lead to premature labor. Understanding the role of MMPs in uteroplacental and vascular remodeling and function could help design new approaches for prediction and management of preeclampsia and premature labor. PMID:28662830

Long noncoding RNA BC200 regulates cell growth and invasion in colon cancer.

PubMed

Wu, Kaiming; Xu, Kaiwu; Liu, Kuanzhi; Huang, Jiehong; Chen, Jianhui; Zhang, Jian; Zhang, Ning

2018-06-01

Colon cancer is the third most commonly diagnosed and deadly cancer worldwide. Efforts have been made to characterize its pathological mechanisms and to explore new therapeutic targets of this disease. Aberrant expression of long noncoding RNAs (lncRNAs) has been associated with the pathogenesis of colon cancer. In the current study, we aimed to define the biological mechanism of the lncRNA BC200 in colon cancer. Here, we found that expression of BC200 was up-regulated in colon cancer tissues as compared with adjacent non-cancerous tissues. The BC200 level was positively correlated with advanced TNM stage. The Kaplan-Meier method indicated that the cumulative survival rate was significantly lower in patients with high BC200 expression than in those with low BC200 expression. Interestingly, we found that knockdown of BC200 inhibited proliferation of HCT-116 and HT29 colon cancer cell lines and reduce the expression of cell proliferation markers, such as Ki-67 and PCNA. In addition, silencing of BC200 could induce obvious G0/G1 arrest and cause apoptosis in HCT-116 and HT29 cells and reduced the expression of cyclin D1, cyclin E, and c-Myc through inhibiting the expression of β-catenin. Importantly, we found that knockdown of BC200 reduced invasion of HCT-116 and HT29 cells and epithelial-mesenchymal transition (EMT) by reducing the expression of MMP-2 and MMP-9. Mechanistically, silencing of BC200 significantly reduced the phosphorylation of STAT3. Overall, the findings presented here suggest that lncRNA BC200 may serve as a novel oncogene and a new therapeutic target for colon cancer. Copyright © 2018. Published by Elsevier Ltd.

The role of lipopolysaccharide in infectious bone resorption of periapical lesion.

PubMed

Hong, Chi-Yuan; Lin, Sze-Kwan; Kok, Sang-Heng; Cheng, Shih-Jung; Lee, Ming-Shu; Wang, Tong-Mei; Chen, Chuan-Shuo; Lin, Li-Deh; Wang, Juo-Song

2004-03-01

The role of lipopolysaccharide (LPS) in periapical lesion-induced bone resorption was investigated. Polymyxin B (PMB), a specific inhibitor of LPS, was evaluated to treat the apical lesion. Lipopolysaccharide isolated from two common endodontic pathogens, Fusobacterium nucleatum and Porphyromonas endodontalis, stimulated mouse macrophage (J774) to release interleukin-1alpha (IL-1 alpha) and tumor necrosis factor-alpha (TNF-alpha) in a time-dependent manner. Combination of LPS further enhanced the stimulation. PMB inhibited these effects significantly. LPS also stimulated matrix metalloproteinase-1 (MMP-1) gene expression in J774, whereas anti-IL-1 alpha and anti-TNF-alpha antibodies, as well as PMB, diminished this effect. A disease model of periapical lesion was established in Wistar rat. Administration of PMB reduced the extent of lesion-associated bone resorption by 76% to approximately 80%, and simultaneously reduced the numbers of MMP-1-producing macrophages. It is suggested that LPS released from the infected root canal triggers the synthesis of IL-1 alpha and TNF-alpha from macrophages. These pro-inflammatory cytokines up-regulate the production of MMP-1 by macrophages to promote periapical bone resorption.

Heat shock proteins HSP70 and MRJ cooperatively regulate cell adhesion and migration through urokinase receptor.

PubMed

Lin, Yuli; Peng, Nana; Zhuang, Hongqin; Zhang, Di; Wang, Yao; Hua, Zi-Chun

2014-08-30

The urokinase-type plasminogen activator receptor (uPAR) is an important regulator of ECM proteolysis, cell-ECM interactions and cell signaling. uPAR and heat shock proteins HSP70 and MRJ (DNAJB6) have been implicated in tumor growth and metastasis. We have reported recently that MRJ (DNAJB6, a heat shock protein) can interact with uPAR and enhance cell adhesion. Here, we identified another heat shock protein HSP70 as a novel uPAR-interacting protein. We performed co-immunoprecipitation in human embryonic kidney (HEK) 293 and colon cancer HCT116 cells as well as immunofluorence assays in HEK293 cells stably transfected with uPAR to investigate the association of suPAR with HSP70/MRJ. To understand the biological functions of the triple complex of suPAR/HSP70/MRJ, we determined whether HSP70 and/or MRJ regulated uPAR-mediated cell invasion, migration, adhesion to vitronectin and MAPK pathway in two pair of human tumor cells (uPAR negative HEK293 cells vs HEK293 cells stably transfected with uPAR and HCT116 cells stably transfected with antisense-uPAR vs HCT116 mock cells transfected with vector only) using transwell assay, wound healing assay, quantitative RT-PCR analyzing mmp2 and mmp9 transcription levels, cell adhesion assay and Western blotting assay. HSP70 and MRJ formed a triple complex with uPAR and over-expression of MRJ enhanced the interaction between HSP70 and uPAR, while knockdown of MRJ decreased soluble uPAR in HCT116 cells (P < 0.05) and reduced the formation of the triple complex, suggesting that MRJ may act as an uPAR-specific adaptor protein to link uPAR to HSP70. Further experiments showed that knockdown of HSP70 and/or MRJ by siRNA inhibited uPAR-mediated cell adhesion to vitronectin as well as suppressed cell invasion and migration. Knockdown of HSP70 and/or MRJ inhibited expression of invasion related genes mmp2 and mmp9. Finally, HSP70 and/or MRJ up-regulated phosphorylation levels of ERK1/2 and FAK suggesting MAPK pathway was involved. All the biological function experiments in cell level showed an additive effect when HSP70 and MRJ were regulated simultaneously indicating their collaborated regulation effects on uPAR. These findings may offer a novel insight into the interactions between uPAR and HSP70/MRJ and their functions in cell adhesion and migration may provide more understanding of the roles in regulating cancer metastasis.

A HER2-targeting antibody-drug conjugate, trastuzumab deruxtecan (DS-8201a), enhances antitumor immunity in a mouse model.

PubMed

Iwata, Tomomi Nakayama; Ishii, Chiaki; Ishida, Saori; Ogitani, Yusuke; Wada, Teiji; Agatsuma, Toshinori

2018-04-27

Trastuzumab deruxtecan (DS-8201a), a HER2-targeting antibody-drug conjugate with a topoisomerase I inhibitor exatecan derivative (DX-8951 derivative, DXd), has been reported to exert potent antitumor effects in xenograft mouse models and clinical trials. In this study, the immune system-activating ability of DS-8201a was assessed. DS-8201a significantly suppressed tumor growth in an immunocompetent mouse model with human HER2-expressing CT26.WT (CT26.WT-hHER2) cells. Cured immunocompetent mice rejected not only re-challenged CT26.WT-hHER2 cells, but also CT26.WT-mock cells. Splenocytes from the cured mice responded to both CT26.WT-hHER2 and CT26.WT-mock cells. Further analyses revealed that DXd up-regulated CD86 expression on bone marrow-derived DCs in vitro, and that DS-8201a increased tumor-infiltrating DCs and up-regulated their CD86 expression in vivo. DS-8201a also increased tumor-infiltrating CD8+ T cells and enhanced PD-L1 and MHC class I expression on tumor cells. Furthermore, combination therapy with DS-8201a and anti-PD-1 antibody was more effective than either monotherapy. In conclusion, DS-8201a enhanced antitumor immunity, as evidenced by the increased expression of DC markers, augmented expression of MHC class I in tumor cells, and rejection of re-challenged tumor cells by adaptive immune cells, suggesting that DS-8201a enhanced tumor recognition by T cells. Furthermore, DS-8201a treatment benefited from combination with anti-PD-1 antibody, possibly due to increased T cell activity and up-regulated PD-L1 expression induced by DS-8201a. Copyright ©2018, American Association for Cancer Research.

The Plasma Membrane Sialidase NEU3 Regulates the Malignancy of Renal Carcinoma Cells by Controlling β1 Integrin Internalization and Recycling*

PubMed Central

Tringali, Cristina; Lupo, Barbara; Silvestri, Ilaria; Papini, Nadia; Anastasia, Luigi; Tettamanti, Guido; Venerando, Bruno

2012-01-01

The human plasma membrane sialidase NEU3 is a key enzyme in the catabolism of membrane gangliosides, is crucial in the regulation of cell surface processes, and has been demonstrated to be significantly up-regulated in renal cell carcinomas (RCCs). In this report, we show that NEU3 regulates β1 integrin trafficking in RCC cells by controlling β1 integrin recycling to the plasma membrane and controlling activation of the epidermal growth factor receptor (EGFR) and focal adhesion kinase (FAK)/protein kinase B (AKT) signaling. NEU3 silencing in RCC cells increased the membrane ganglioside content, in particular the GD1a content, and changed the expression of key regulators of the integrin recycling pathway. In addition, NEU3 silencing up-regulated the Ras-related protein RAB25, which directs internalized integrins to lysosomes, and down-regulated the chloride intracellular channel protein 3 (CLIC3), which induces the recycling of internalized integrins to the plasma membrane. In this manner, NEU3 silencing enhanced the caveolar endocytosis of β1 integrin, blocked its recycling and reduced its levels at the plasma membrane, and, consequently, inhibited EGFR and FAK/AKT. These events had the following effects on the behavior of RCC cells: they (a) decreased drug resistance mediated by the block of autophagy and the induction of apoptosis; (b) decreased metastatic potential mediated by down-regulation of the metalloproteinases MMP1 and MMP7; and (c) decreased adhesion to collagen and fibronectin. Therefore, our data identify NEU3 as a key regulator of the β1 integrin-recycling pathway and FAK/AKT signaling and demonstrate its crucial role in RCC malignancy. PMID:23139422

Qishenyiqi Dropping Pill attenuates myocardial fibrosis in rats by inhibiting RAAS-mediated arachidonic acid inflammation.

PubMed

Wang, Jing; Lu, Linghui; Wang, Yong; Wu, Yan; Han, Jing; Wang, Wei; Li, Chun; Tu, Pengfei

2015-12-24

In China, Qishenyiqi Dropping Pill (QSDP), a Chinese medicine formula containing Astragalus membranaceus (Fisch.) Bunge, Salvia miltiorrhiza Bunge, Panax notoginseng (Burkill) F.H.Chen and Dalbergia odorifera T.C.Chen, has been used frequently in traditional folk medicine for treatment of coronary heart diseases (CHD) and heart failure (HF). Previous study has shown that QSDP has definite therapeutic effects on promoting the heart function on CHD patients. The present study was designed to study the anti-fibrosis effects of QSDP on HF rats and to explore the underlying molecular mechanisms. HF rat model was induced by left anterior descending (LAD) coronary artery ligation. Two-dimensional (2D) echocardiography was adopted to evaluate heart functions. Immunohistochemical (IHC) method and Western-blot were used to detect expression of critical proteins in renin-angiotensin-aldosterone system (RAAS) or arachidonic acid (AA) metabolic pathway. Heart functions were seriously injured in the model group. Expressions of fibrotic markers, such as collagen Ⅰ, collagen Ⅲ, matrix metallopeptidase 2 (MMP2) and MMP9 were elevated in the model group. RAAS pathway was activated. Interestingly, AA pathway was also up-regulated in the model group and it was down-regulated by angiotensin converting enzyme inhibitors (ACEIs) drug Captopril. Expressions of the important signal-transuding proteins, including NF-κB, JAK1/STAT3 and Akt, all increased remarkably in the model group. Treatment with QSDP could attenuate myocardial fibrosis by inhibiting RAAS-activated pathway, as indicated by decreased angiotensin type 1 receptor (AT1) and increased AT2 expression. Expressions of phospholipase A2 (PLA2), cyclooxygenase 1 (COX1) and COX2 were also down-regulated in the QSDP-treated group. In addition, "therapeutic" QSDP administration seemed to down-regulate expressions of NF-κB, JAK1/ STAT3 and Akt which may play important roles in myocardial fibrosis. QSDP can exert anti-fibrosis effect by down-regulating RAAS pathway, and subsequently inhibiting expressions of proteins in AA pathway. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

UV radiation-induced skin aging in hairless mice is effectively prevented by oral intake of sea buckthorn (Hippophae rhamnoides L.) fruit blend for 6 weeks through MMP suppression and increase of SOD activity.

PubMed

Hwang, In Sik; Kim, Ji Eun; Choi, Sun Il; Lee, Hye Ryun; Lee, Young Ju; Jang, Min Ju; Son, Hong Ju; Lee, Hee Seob; Oh, Chung Hun; Kim, Bae Hwan; Lee, Sang Hak; Hwang, Dae Youn

2012-08-01

Oxidative stress and oxidative photodamage induced by UV radiation can cause serious skin damage that is characterized by wrinkling, roughness, laxity and pigmentation. The effects of a sea buckthorn (Hippophae rhamnoides L.) fruit blend (SFB) containing sea buckthorn fruit extract, blueberry extract and collagen on UV-induced skin aging were examined by treating hairless mice for 6 weeks with UV irradiation and SFB administered orally. The effects of SFB were measured in the skin of these mice by phenotypical and histological analysis and western blotting. According to wrinkle formation analysis, the oral intake of SFB induced a decrease in wrinkle formation in the damaged skin of UV-irradiated mice. The thickness of the epidermis and dermis in the vitamin extracts (Vit)- and SFB-treated group was lower than that in the vehicle-treated group, but the group treated with SFB50 was the most effective group. The mice treated with the Vit- or SFB solution maintained a normal moisture content through the inhibition of transdermal water loss (TEWL) and an increase in skin moisture content. Furthermore, the levels of matrix metalloproteinase (MMP) and collagen protein expression were assessed in five groups to examine the mechanisms underlying the effects of SFB oral intake. The application of SFB induced a decrease in MMP-1 and -9 expression to the levels observed in the vehicle-treated group, but MMP-9 expression showed a much larger decrease than MMP-1. Furthermore, the expression of collagen-1 in the skin corresponded to MMP expression except for the SFB30-treated group, whereas the superoxide dismutase (SOD) activity was increased dramatically in the SFB50-treated group. These results suggest that SFB has potential as a protective and therapeutic drug candidate against skin aging that functions by regulating the moisture content, MMP expression levels and SOD activity.

Profiling analysis of FOX gene family members identified FOXE1 as potential regulator of NSCLC development.

PubMed

Ji, G H; Cui, Y; Yu, H; Cui, X B

2016-09-30

Lung cancer is one of the most malignant tumors worldwide with a high mortality rate, which has not been improved since several decades ago. FOX gene family members have been reported to play extensive roles in regulating many biological processes and disorders. In order to clarify the contribution of FOX gene family members in lung cancer biology, we performed expression profiling analysis of FOX gene family members from FOXA to FOXR in lung cancer cell lines and tissue specimens by Real-time PCR, western blot and immunohistochemistry analysis. We found that FOXE1 was the only gene which was over-expressed in six out of eight lung cancer cell lines and human cancer tissue specimens (28 out of 35 cases with higher expression and 7 out of 35 cases with moderate expression). Further investigation showed that MMP2 gene was up-regulated, and autophagy markers such as LC3B, ATG5, ATG12 and BECLIN1, were down-regulated concomitant with the increase of FOXE1. These results implicated that FOXE1 may be an important regulator by targeting autophagy and MMPs pathways in lung cancer development.

p-Coumaric Acid Attenuates UVB-Induced Release of Stratifin from Keratinocytes and Indirectly Regulates Matrix Metalloproteinase 1 Release from Fibroblasts

PubMed Central

Seok, Jin Kyung

2015-01-01

Ultraviolet (UV) radiation-induced loss of dermal extracellular matrix is associated with skin photoaging. Recent studies demonstrated that keratinocyte-releasable stratifin (SFN) plays a critical role in skin collagen metabolism by inducing matrix metalloproteinase 1 (MMP1) expression in target fibroblasts. In the present study, we examined whether SFN released from UVB-irradiated epidermal keratinocytes increases MMP1 release from dermal fibroblasts, and whether these events are affected by p-coumaric acid (p-CA), a natural phenolic compound with UVB-shielding and antioxidant properties. HaCaT cells were exposed to UVB in the absence and presence of p-CA, and the conditioned medium was used to stimulate fibroblasts in medium transfer experiments. The cells and media were analyzed to determine the expressions/releases of SFN and MMP1. UVB exposure increased SFN release from keratinocytes into the medium. The conditioned medium of UVB-irradiated keratinocytes increased MMP1 release from fibroblasts. The depletion of SFN using a siRNA rendered the conditioned medium of UVB-irradiated keratinocytes ineffective at stimulating fibroblasts to release MMP1. p-CA mitigated UVB-induced SFN expression in keratinocytes, and attenuated the MMP1 release by fibroblasts in medium transfer experiments. In conclusion, the present study demonstrated that the use of UV absorbers such as p-CA would reduce UV-induced SFN-centered signaling events involved in skin photoaging. PMID:25954129

p-Coumaric Acid Attenuates UVB-Induced Release of Stratifin from Keratinocytes and Indirectly Regulates Matrix Metalloproteinase 1 Release from Fibroblasts.

PubMed

Seok, Jin Kyung; Boo, Yong Chool

2015-05-01

Ultraviolet (UV) radiation-induced loss of dermal extracellular matrix is associated with skin photoaging. Recent studies demonstrated that keratinocyte-releasable stratifin (SFN) plays a critical role in skin collagen metabolism by inducing matrix metalloproteinase 1 (MMP1) expression in target fibroblasts. In the present study, we examined whether SFN released from UVB-irradiated epidermal keratinocytes increases MMP1 release from dermal fibroblasts, and whether these events are affected by p-coumaric acid (p-CA), a natural phenolic compound with UVB-shielding and antioxidant properties. HaCaT cells were exposed to UVB in the absence and presence of p-CA, and the conditioned medium was used to stimulate fibroblasts in medium transfer experiments. The cells and media were analyzed to determine the expressions/releases of SFN and MMP1. UVB exposure increased SFN release from keratinocytes into the medium. The conditioned medium of UVB-irradiated keratinocytes increased MMP1 release from fibroblasts. The depletion of SFN using a siRNA rendered the conditioned medium of UVB-irradiated keratinocytes ineffective at stimulating fibroblasts to release MMP1. p-CA mitigated UVB-induced SFN expression in keratinocytes, and attenuated the MMP1 release by fibroblasts in medium transfer experiments. In conclusion, the present study demonstrated that the use of UV absorbers such as p-CA would reduce UV-induced SFN-centered signaling events involved in skin photoaging.

Human neuronal uncoupling proteins 4 and 5 (UCP4 and UCP5): structural properties, regulation, and physiological role in protection against oxidative stress and mitochondrial dysfunction

PubMed Central

Ramsden, David B; Ho, Philip W-L; Ho, Jessica W-M; Liu, Hui-Fang; So, Danny H-F; Tse, Ho-Man; Chan, Koon-Ho; Ho, Shu-Leong

2012-01-01

Uncoupling proteins (UCPs) belong to a large family of mitochondrial solute carriers 25 (SLC25s) localized at the inner mitochondrial membrane. UCPs transport protons directly from the intermembrane space to the matrix. Of five structural homologues (UCP1 to 5), UCP4 and 5 are principally expressed in the central nervous system (CNS). Neurons derived their energy in the form of ATP that is generated through oxidative phosphorylation carried out by five multiprotein complexes (Complexes I–V) embedded in the inner mitochondrial membrane. In oxidative phosphorylation, the flow of electrons generated by the oxidation of substrates through the electron transport chain to molecular oxygen at Complex IV leads to the transport of protons from the matrix to the intermembrane space by Complex I, III, and IV. This movement of protons to the intermembrane space generates a proton gradient (mitochondrial membrane potential; MMP) across the inner membrane. Complex V (ATP synthase) uses this MMP to drive the conversion of ADP to ATP. Some electrons escape to oxygen-forming harmful reactive oxygen species (ROS). Proton leakage back to the matrix which bypasses Complex V resulting in a major reduction in ROS formation while having a minimal effect on MMP and hence, ATP synthesis; a process termed “mild uncoupling.” UCPs act to promote this proton leakage as means to prevent excessive build up of MMP and ROS formation. In this review, we discuss the structure and function of mitochondrial UCPs 4 and 5 and factors influencing their expression. Hypotheses concerning the evolution of the two proteins are examined. The protective mechanisms of the two proteins against neurotoxins and their possible role in regulating intracellular calcium movement, particularly with regard to the pathogenesis of Parkinson's disease are discussed. PMID:22950050

Acute transcriptional up-regulation specific to osteoblasts/osteoclasts in medaka fish immediately after exposure to microgravity

PubMed Central

Chatani, Masahiro; Morimoto, Hiroya; Takeyama, Kazuhiro; Mantoku, Akiko; Tanigawa, Naoki; Kubota, Koji; Suzuki, Hiromi; Uchida, Satoko; Tanigaki, Fumiaki; Shirakawa, Masaki; Gusev, Oleg; Sychev, Vladimir; Takano, Yoshiro; Itoh, Takehiko; Kudo, Akira

2016-01-01

Bone loss is a serious problem in spaceflight; however, the initial action of microgravity has not been identified. To examine this action, we performed live-imaging of animals during a space mission followed by transcriptome analysis using medaka transgenic lines expressing osteoblast and osteoclast-specific promoter-driven GFP and DsRed. In live-imaging for osteoblasts, the intensity of osterix- or osteocalcin-DsRed fluorescence in pharyngeal bones was significantly enhanced 1 day after launch; and this enhancement continued for 8 or 5 days. In osteoclasts, the signals of TRAP-GFP and MMP9-DsRed were highly increased at days 4 and 6 after launch in flight. HiSeq from pharyngeal bones of juvenile fish at day 2 after launch showed up-regulation of 2 osteoblast- and 3 osteoclast- related genes. Gene ontology analysis for the whole-body showed that transcription of genes in the category “nucleus” was significantly enhanced; particularly, transcription-regulators were more up-regulated at day 2 than at day 6. Lastly, we identified 5 genes, c-fos, jun-B-like, pai-1, ddit4 and tsc22d3, which were up-regulated commonly in the whole-body at days 2 and 6, and in the pharyngeal bone at day 2. Our results suggested that exposure to microgravity immediately induced dynamic alteration of gene expression levels in osteoblasts and osteoclasts. PMID:28004797

Temperature oscillations drive cycles in the activity of MMP-2,9 secreted by a human trabecular meshwork cell line.

PubMed

Li, Stanley Ka-Lok; Banerjee, Juni; Jang, Christopher; Sehgal, Amita; Stone, Richard A; Civan, Mortimer M

2015-02-05

Aqueous humor inflow falls 50% during sleeping hours without proportional fall in IOP, partly reflecting reduced outflow facility. The mechanisms underlying outflow facility cycling are unknown. One outflow facility regulator is matrix metalloproteinase (MMP) release from trabecular meshwork (TM) cells. Because anterior segment temperature must oscillate due to core temperature cycling and eyelid closure during sleep, we tested whether physiologically relevant temperature oscillations drive cycles in the activity of secreted MMP. Temperature of transformed normal human TM cells (hTM5 line) was fixed or alternated 12 hours/12 hours between 33°C and 37°C. Activity of secreted MMP-2 and MMP-9 was measured by zymography, and gene expression by RT-PCR and quantitative PCR. Raising temperature to 37°C increased, and lowering to 33°C reduced, activity of secreted MMP. Switching between 37°C and 33°C altered MMP-9 by 40% ± 3% and MMP-2 by 22% ± 2%. Peripheral circadian clocks did not mediate temperature-driven cycling of MMP secretion because MMP-release oscillations did not persist at constant temperature after 3 to 6 days of alternating temperatures, and temperature cycles did not entrain clock-gene expression in these cells. Furthermore, inhibiting heat shock transcription factor 1, which links temperature and peripheral clock-gene oscillations, inhibited MMP-9 but not MMP-2 temperature-driven MMP cycling. Inhibition of heat-sensitive TRPV1 channels altered total MMP secretion but not temperature-induced modulations. Inhibiting cold-sensitive TRPM-8 channels had no effect. Physiologically relevant temperature oscillations drive fluctuations of secreted MMP-2 and MMP-9 activity in hTM5 cells independent of peripheral clock genes and temperature-sensitive TRP channels. Copyright 2015 The Association for Research in Vision and Ophthalmology, Inc.

Doxycycline blocks gastric ulcer by regulating matrix metalloproteinase-2 activity and oxidative stress

PubMed Central

Singh, Laishram Pradeepkumar; Mishra, Amartya; Saha, Debjit; Swarnakar, Snehasikta

2011-01-01

AIM: To examine the effect of doxycycline on the activity of matrix metalloproteinases (MMPs) and oxidative stress in gastric tissues of rats following gastric injury. METHODS: Gastric ulcers were generated in rats by administration of 70% ethanol, and activity of doxycycline was tested by administration 30 min prior to ethanol. Similarly, the effect of doxycycline was tested in an indomethacin-induced gastric ulcer model. The activities and expression of MMPs were examined by zymography and Western blot analysis. RESULTS: Gastric injury in rats as judged by elevated ulcer indices following exposure to ulcerogen, either indomethacin or ethanol, was reversed significantly by doxycycline. Indomethacin-induced ulcerated gastric tissues exhibited about 12-fold higher proMMP-9 activity and about 5-fold higher proMMP-3 activity as compared to control tissues. Similarly, ethanol induced about 22-fold and about 6-fold higher proMMP-9 and proMMP-3 activities, respectively, in rat gastric tissues. Both proMMP-9 and MMP-3 activities were markedly decreased by doxycycline in ulcerogen treated rat gastric tissues. In contrast, the reduced MMP-2 activity in ulcerated tissues was increased by doxycycline during ulcer prevention. On the other hand, doxycycline inhibited significantly proMMP-9, -2 and -3 activities in vitro. In addition, doxycycline reduced oxidative load in gastric tissues and scavenged H2O2 in vitro. Our results suggest a novel regulatory role of doxycycline on MMP-2 activity in addition to inhibitory action on MMP-9 and MMP-3 during prevention of gastric ulcers. CONCLUSION: This is the first demonstration of dual action of doxycycline, that is, regulation of MMP activity and reduction of oxidative stress in arresting gastric injury. PMID:21876619

Identification of Caspase-6 as a New Regulator of Alternatively Activated Macrophages*

PubMed Central

Yao, Yongfang; Shi, Qian; Chen, Bing; Wang, Qingsong; Li, Xinda; Li, Long; Huang, Yahong; Ji, Jianguo; Shen, Pingping

2016-01-01

Alternatively activated macrophages (AAMs) play essential roles in the promotion of tissue remodeling, vasculogenesis, and tumor progression; however, the detailed mechanisms underlying the activation of AAMs remain largely unknown. Here, by using quantitative proteomic analysis, we identified 62 proteins that were up-regulated in IL-4-induced macrophages. Among these, Caspase-6 was increased significantly. Caspase-6 is important in the apoptotic signaling pathway; however, its role in non-apoptosis is also reported. Here, we first examined the non-apoptotic role of Caspase-6 in the alternative activation of macrophages after administration of IL-4, 4T1 tumor conditional medium, or co-culture with 4T1 cells. Both treatments promoted alternative activation of RAW264.7 cells and primary macrophages, whereas disruption of caspase-6 expression and activity could markedly suppress the biomarker levels of AAMs. Overexpression of Caspase-6 could significantly promote the activation of AAMs. Importantly, we further present evidence that caspase-6 could regulate breast cancer cell invasion by modulating MMP-2 and MMP-9 expression in 4T1 tumor-associated macrophages, as ablation of protein levels or activity of caspase-6 suppressed tumor cell invasion in vitro. In conclusion, the observed results markedly expanded our views of the dynamic changes in protein composition during alternative activation of macrophages, and they revealed a critical new role of caspase-6 in regulating this cellular biological process, which suggested that caspase-6 might be a key nod molecule to regulate immunological steady-state and be a therapeutic candidate for tumor immunotherapy. PMID:27325699

α2 Integrin, extracellular matrix metalloproteinase inducer, and matrix metalloproteinase-3 act sequentially to induce differentiation of mouse embryonic stem cells into odontoblast-like cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ozeki, Nobuaki; Kawai, Rie; Hase, Naoko

We previously reported that interleukin 1β acts via matrix metalloproteinase (MMP)-3 to regulate cell proliferation and suppress apoptosis in α2 integrin-positive odontoblast-like cells differentiated from mouse embryonic stem (ES) cells. Here we characterize the signal cascade underpinning odontoblastic differentiation in mouse ES cells. The expression of α2 integrin, extracellular matrix metalloproteinase inducer (Emmprin), and MMP-3 mRNA and protein were all potently increased during odontoblastic differentiation. Small interfering RNA (siRNA) disruption of the expression of these effectors potently suppressed the expression of the odontoblastic biomarkers dentin sialophosphoprotein, dentin matrix protein-1 and alkaline phosphatase, and blocked odontoblast calcification. Our siRNA, western blotmore » and blocking antibody analyses revealed a unique sequential cascade involving α2 integrin, Emmprin and MMP-3 that drives ES cell differentiation into odontoblasts. This cascade requires the interaction between α2 integrin and Emmprin and is potentiated by exogenous MMP-3. Finally, although odontoblast-like cells potently express α2, α6, αV, β1, and β3, integrins, we confirmed that β1 integrin acts as the trigger for ES cell differentiation, apparently in complex with α2 integrin. These results demonstrate a unique and unanticipated role for an α2 integrin-, Emmprin-, and MMP-3-mediated signaling cascade in driving mouse ES cell differentiation into odontoblast-like cells. - Highlights: • Odontoblast differentiation requires activation of α2 integrin, Emmprin and MMP-3. • α2 integrin, Emmprin and MMP-3 form a sequential signaling cascade. • β1 integrin acts a specific trigger for odontoblast differentiation. • The role of these effectors is highly novel and unanticipated.« less

Redox regulation of MMP-3/TIMP-1 ratio in intestinal myofibroblasts: effect of N-acetylcysteine and curcumin.

PubMed

Fontani, Filippo; Marcucci, Tommaso; Picariello, Lucia; Tonelli, Francesco; Vincenzini, Maria Teresa; Iantomasi, Teresa

2014-04-15

Matrix metalloproteinases (MMPs) play a critical role in inflammation and ulcerations in gut of Crohn׳s disease (CD) patients. Intestinal subepithelial myofibroblasts (ISEMFs) secrete MMPs in response to inflammatory stimuli. Previous data showed in CD-ISEMFs increased oxidative status. The aim of this study was to investigate the role of ISEMFs in modulating the production of MMP-3 and TIMP-1, an inhibitor of MMPs activity. A relationship among oxidative stress, activity of antioxidants and MMP-3/TIMP-1 was also studied. ISEMFs isolated from CD patient colon and human colonic cell line of myofibroblasts (18Co) were used. Oxidative state was modulated by buthionine sulfoximine, an inhibitor of glutathione (GSH) synthesis, and N-acetylcysteine (NAC), GSH precursor. An up-regulation of MMP-3 due to increased oxidative state was found in CD-ISEMFs. Stimulation by tumor necrosis factor (TNF)α increased further MMP-3 levels. On the contrary, no change in TIMP-1 production was determined. NAC treatment decreased MMP-3 production in CD-ISMEFs and removed the enhancement due to TNFα. Similar effects were observed in 18Co cells treated with curcumin, antioxidant with anti-inflammatory properties. The involvement of MAPKs on MMP-3 redox regulation was also shown. This study demonstrates the involvement of ISEMFs and high oxidative state in the increased MMP-3 production found in intestinal mucosa of CD patients. NAC and curcumin normalize MMP-3 levels mainly in TNFα stimulated cells. A modulation of MMP-3 production by NAC and curcumin due to their direct action on transcriptional factors has been also suggested. Therefore, they could have a therapeutic use for the prevention and treatment of fistulaes in CD. Copyright © 2014 Elsevier Inc. All rights reserved.

Matrix Metalloproteinase-20 Over-Expression Is Detrimental to Enamel Development: A Mus musculus Model

PubMed Central

Shin, Masashi; Hu, Yuanyuan; Tye, Coralee E.; Guan, Xiaomu; Deagle, Craig C.; Antone, Jerry V.; Smith, Charles E.; Simmer, James P.; Bartlett, John D.

2014-01-01

Background Matrix metalloproteinase-20 (Mmp20) ablated mice have enamel that is thin and soft with an abnormal rod pattern that abrades from the underlying dentin. We asked if introduction of transgenes expressing Mmp20 would revert this Mmp20 null phenotype back to normal. Unexpectedly, for transgenes expressing medium or high levels of Mmp20, we found opposite enamel phenotypes depending on the genetic background (Mmp20−/− or Mmp20+/+) in which the transgenes were expressed. Methodology/Principal Findings Amelx-promoter-Mmp20 transgenic founder mouse lines were assessed for transgene expression and those expressing low, medium or high levels of Mmp20 were selected for breeding into the Mmp20 null background. Regardless of expression level, each transgene brought the null enamel back to full thickness. However, the high and medium expressing Mmp20 transgenes in the Mmp20 null background had significantly harder more mineralized enamel than did the low transgene expresser. Strikingly, when the high and medium expressing Mmp20 transgenes were present in the wild-type background, the enamel was significantly less well mineralized than normal. Protein gel analysis of enamel matrix proteins from the high and medium expressing transgenes present in the wild-type background demonstrated that greater than normal amounts of cleavage products and smaller quantities of higher molecular weight proteins were present within their enamel matrices. Conclusions/Significance Mmp20 expression levels must be within a specific range for normal enamel development to occur. Creation of a normally thick enamel layer may occur over a wider range of Mmp20 expression levels, but acquisition of normal enamel hardness has a narrower range. Since over-expression of Mmp20 results in decreased enamel hardness, this suggests that a balance exists between cleaved and full-length enamel matrix proteins that are essential for formation of a properly hardened enamel layer. It also suggests that few feedback controls are present in the enamel matrix to prevent excessive MMP20 activity. PMID:24466234

Inhibition of matrix metalloproteinase-2 by PARP inhibitors

DOE Office of Scientific and Technical Information (OSTI.GOV)

Nicolescu, Adrian C.; Holt, Andrew; Kandasamy, Arulmozhi D.

2009-10-02

Matrix metalloproteinase-2 (MMP-2), a ubiquitously expressed zinc-dependent endopeptidase, and poly(ADP-ribosyl) polymerase (PARP), a nuclear enzyme regulating DNA repair, are activated by nitroxidative stress associated with various pathologies. As MMP-2 plays a detrimental role in heart injuries resulting from enhanced nitroxidative stress, where PARP and MMP inhibitors are beneficial, we hypothesized that PARP inhibitors may affect MMP-2 activity. Using substrate degradation assays to determine MMP-2 activity we found that four PARP inhibitors (3-AB, PJ-34, 5-AIQ, and EB-47) inhibited 64 kDa MMP-2 in a concentration-dependent manner. The IC{sub 50} values of PJ-34 and 5-AIQ were in the high micromolar range and comparablemore » to those of known MMP-2 inhibitors doxycycline, minocycline or o-phenanthroline, whereas those for 3-AB and EB-47 were in the millimolar range. Co-incubation of PARP inhibitors with doxycycline showed an additive inhibition of MMP-2 that was significant for 3-AB alone. These data demonstrate that the protective effects of some PARP inhibitors may include inhibition of MMP-2 activity.« less

Sera from remitting and secondary progressive multiple sclerosis patients disrupt the blood-brain barrier.

PubMed

Shimizu, Fumitaka; Tasaki, Ayako; Sano, Yasuteru; Ju, Mihua; Nishihara, Hideaki; Oishi, Mariko; Koga, Michiaki; Kawai, Motoharu; Kanda, Takashi

2014-01-01

Pathological destruction of blood-brain barrier (BBB) has been thought to be the initial key event in the process of developing multiple sclerosis (MS). The purpose of the present study was to clarify the possible molecular mechanisms responsible for the malfunction of BBB by sera from relapse-remitting MS (RRMS) and secondary progressive MS (SPMS) patients. We evaluated the effects of sera from the patients in the relapse phase of RRMS (RRMS-R), stable phase of RRMS (RRMS-S) and SPMS on the expression of tight junction proteins and vascular cell adhesion protein-1 (VCAM-1), and on the transendothelial electrical resistance (TEER) in human brain microvascular endothelial cells (BMECs). Sera from the RRMS-R or SPMS patients decreased the claudin-5 protein expression and the TEER in BMECs. In RRMS-R, this effect was restored after adding an MMP inhibitor, and the MMP-2/9 secretion by BMECs was significantly increased after the application of patients' sera. In SPMS, the immunoglobulin G (IgG) purified from patients' sera also decreased the claudin-5 protein expression and the TEER in BMECs. The sera and purified IgG from all MS patients increased the VCAM-1 protein expression in BMECs. The up-regulation of autocrine MMP-2/9 by BMECs after exposure to sera from RRMS-R patients or the autoantibodies against BMECs from SPMS patients can compromise the BBB. Both RRMS-S and SPMS sera increased the VCAM-1 expression in the BBB, thus indicating that targeting the VCAM-1 in the BBB could represent a possible therapeutic strategy for even the stable phase of MS and SPMS.

Glucose-derived AGEs promote migration and invasion of colorectal cancer by up-regulating Sp1 expression.

PubMed

Deng, Ruyuan; Wu, Huo; Ran, Hui; Kong, Xiang; Hu, Lei; Wang, Xiao; Su, Qing

2017-05-01

It is well established that the risk of colorectal cancer (CRC) is significantly increased in diabetic patients. As one of main forms of the advanced glycation end products (AGEs) that accumulate in vivo, glucose-derived AGEs play an important role in the pathogenesis of diabetic complications and may contribute to CRC progression. However, to date, both the contribution of glucose-derived AGEs to the course of CRC and the underlying mechanism are unclear. In the present study, the concentration of glucose-derived AGEs in the serum and tumor tissue of patients with CRC increased. A clinical data analysis demonstrated that the expression of the receptor for AGEs (RAGE), Specificity Protein 1 (Sp1), and matrix metallopeptidase -2 (MMP2) was significantly higher in cancerous tissues compared with non-tumor tissue in Chinese Han patients with CRC and that RAGE expression was closely associated with lymph node metastasis and TNM stage. Furthermore, in vivo and in vitro experiments showed that AGEs promoted invasion and migration of colorectal cancer, and the AGEs treatment increased the expression of RAGE, Sp1, and MMP2 in a dose-dependent manner. A RAGE blocking antibody and an Sp1-specific siRNA attenuated the AGE-induced effects. Moreover, the AGEs treatment increased the phosphorylation of ERK, and reducing the phosphorylation level of ERK by MEK1/2 inhibitor decreased the expression of Sp1. In conclusion, glucose-derived AGEs promote the invasion and metastasis of CRC partially through the RAGE/ERK/SP1/MMP2 cascade. These findings may provide an explanation for the poor prognoses of colorectal cancer in diabetic patients. Copyright © 2017 Elsevier B.V. All rights reserved.

Contrasting expression of membrane metalloproteinases, MT1-MMP and MT3-MMP, suggests distinct functions in skeletal development.

PubMed

Yang, Maozhou; Zhang, Bingbing; Zhang, Liang; Gibson, Gary

2008-07-01

Membrane-type 1 matrix metalloproteinase (MT1-MMP) is the most ubiquitous and widely studied of the membrane-type metalloproteinases (MT-MMPs). It was thus surprising to find no published data on chicken MT1-MMP. We report here the characterization of the chicken gene. Its low sequence identity with the MT1-MMP genes of other species, high GC content, and divergent catalytic domain explains the absence of data and our difficulties in characterizing the gene. The absence of structural features in the chicken gene that have been suggested to be critical for the activation of MMP-2 by MT1-MMP; for the effect of MT1-MMP on cell migration and for the recycling of MT1-MMP suggest these features are either not essential or that MT1-MMP does not perform these functions in chickens. Comparison of the expression of chicken MT1-MMP with MT3-MMP and with MMP-2 and MMP-13 has confirmed the previously recognized co-expression of MT1-MMP with MMP-2 and MMP-13 in fibrous and vascular tissues, particularly those surrounding the developing long bones in other species. By contrast, MT3-MMP expression differs markedly from that of MT1-MMP and of both MMP-2 and MMP-13. MT3-MMP is expressed by chondrocytes of the developing articular surface. Similar expression patterns of this group of MT-MMPs and MMPs have been observed in mouse embryos and suggest distinct and specific functions for MT1-MMP and MT3-MMP in skeletal development.

Long non-coding RNA PICART1 suppresses proliferation and promotes apoptosis in lung cancer cells by inhibiting JAK2/STAT3 signaling.

PubMed

Zhao, J M; Cheng, W; He, X G; Liu, Y L; Wang, F F; Gao, Y F

2018-06-26

Lung cancer remains the most common cause of tumor-related death worldwide. Recent studies have revealed that long non-coding RNAs (lncRNAs) are involved in the development of various cancers, including lung cancer. This study aimed to investigate the effect and the molecular basis of lncRNA PICART1 on lung cancer. We first assessed the PICART1 expression in lung cancer in vitro and vivo by qRT-PCR. Then the expression of PICART1 in SPC-A-1 and NCI-H1975 cell lines was inhibited and overexpressed by transient transfections. Thereafter, cell viability, cell cycle, migration and apoptosis were respectively measured by MTT, Transwell and flow cytometry assay. Furthermore, qRT-PCR and western blot analysis were mainly performed to assess the expression levels of apoptosis- and migration-related proteins and JAK2/STAT3 pathway proteins. Tumor formation was measured by xenograft tumor model assay in vivo. PICART1 expression was down-regulated in human lung cancer tissues and cell lines. Knockdown of PICART1 increased cell viability of lung cancer cell lines. However, PICART1 overexpression inhibited cell cycle progression and promoted apoptosis in SPC-A-1 and NCI-H1975 cell lines. PICART1 overexpression also inhibited migration, as evidenced by up-regulation of E-cadherin, and down-regulation of Twist1, MMP2 and MMP9. Furthermore, we found PICART1 inhibition may regulate cell apoptosis and migration through activating JAK2/STAT3 pathway. In vivo experiments revealed that PICART1 knockdown significantly promoted tumor formation.This study demonstrates that PICART1 overexpression represents an anti-growth and anti-metastasis role in lung cancer cells. Additionally, PICART1 acts as a tumor suppressor may be via regulation of JAK2/STAT3 pathway.

3′3-Diindolylmethane inhibits migration, invasion and metastasis of hepatocellular carcinoma by suppressing FAK signaling

PubMed Central

Li, Wen-Xue; Chen, Li-Ping; Sun, Min-Ying; Li, Jun-Tao; Liu, Hua-Zhang; Zhu, Wei

2015-01-01

Late stage hepatocellular carcinoma (HCC) usually has a low survival rate because it has high potential of metastases and there is no effective cure. 3′3-Diindolylmethane (DIM) is the major product of the acid-catalyzed oligomerization of indole-3-carbinol present in cruciferous vegetables. DIM has been proved to exhibit anticancer properties. In this study, we explored the effects and molecular mechanisms of anti-metastasis of DIM on HCC cells both in vitro and in vivo. We chose two HCC cell lines SMMC-7721 and MHCC-97H that have high potential of invasion. The results showed that DIM inhibited the proliferation, migration and invasion of these two cell lines in vitro. In addition, in vivo study demonstrated that DIM significantly decreased the volumes of SMMC-7721 orthotopic liver tumor and suppressed lung metastasis in nude mice. Focal Adhesion Kinase (FAK) is found over activated in HCC cells. We found that DIM decreased the level of phospho-FAK (Tyr397) both in vitro and in vivo. DIM inhibition of phospho-FAK (Tyr397) led to down-regulation of MMP2/9 and decreased potential of metastasis. DIM also repressed the migration and invasion induced by vitronectin through inactivation of FAK pathway and down-regulation of MMP2/9 in vitro. We also found that pTEN plays a role in down-regulation of FAK by DIM. These results demonstrated that DIM blocks HCC cell metastasis by suppressing tumor cell migration and invasion. The anti-metastasis effect of DIM could be explained to be its down-regulated expression and activation of MMP2/9 partly induced by up-regulation of pTEN and inhibition of phospho-FAK (Tyr397). PMID:26068982

ERK1/2 signalling pathway is involved in CD147-mediated gastric cancer cell line SGC7901 proliferation and invasion.

PubMed

Chen, Liping; Pan, Yuqin; Gu, Ling; Nie, Zhenlin; He, Bangshun; Song, Guoqi; Li, Rui; Xu, Yeqiong; Gao, Tianyi; Wang, Shukui

2013-08-01

This study aimed to investigate the role of CD147 in the progression of gastric cancer and the signalling pathway involved in CD147-mediated gastric cancer cell line SGC7901 proliferation and invasion. Short hairpin RNA (shRNA) expression vectors targeting CD147 were constructed to silence CD147, and the expression of CD147 was monitored by quantitative realtime reverse transcriptase polymerase chain reaction and Western blot and further confirmed by immunohistochemistry in vivo. Cell proliferation was determined by Cell Counting Kit-8 assay, the activities of matrix metalloproteinase (MMP)-2 and MMP-9 were determined by gelatin zymography, and the invasion of SGC7901 was determined by invasion assay. The phosphorylation and non-phosphorylation of the mitogen-activated protein kinases, extracellular signal-regulated kinase1/2 (ERK1/2), P38 and c-Jun NH2-terminal kinase were examined by Western blot. Additionally, the ERK1/2 inhibitor U0126 were used to confirm the signalling pathway involved in CD147-mediated SGC7901 progression. The BALB/c nude mice were used to study tumour progression in vivo. The results revealed that CD147 silencing inhibited the proliferation and invasion of SGC7901 cells, and down-regulated the activities of MMP-2 and MMP-9 and the phosphorylation of the ERK1/2 in SGC7901 cells. ERK1/2 inhibitor U0126 decreased the proliferation, and invasion of SGC7901 cells, and down-regulated the MMP-2 and MMP-9 activities. In a nude mouse model of subcutaneous xenografts, the tumour volume was significantly smaller in the SGC7901/shRNA group compared to the SGC7901 and SGC7901/snc-RNA group. Immunohistochemistry analysis showed that CD147 and p-ERK1/2 protein expressions were down-regulated in the SGC7901/shRNA2 group compared to the SGC7901 and SGC7901/snc-RNA group. These results suggest that ERK1/2 pathway involves in CD147-mediated gastric cancer growth and invasion. These findings further highlight the importance of CD147 in cancer progression, indicating that CD147 would be an attractive therapeutic target for gastric cancer.

Expressions of Matrix Metalloproteinases (MMP-2, MMP-7, and MMP-9) and Their Inhibitors (TIMP-1, TIMP-2) in Inflammatory Bowel Diseases

PubMed Central

Jakubowska, Katarzyna; Pryczynicz, Anna; Iwanowicz, Piotr; Niewiński, Andrzej; Maciorkowska, Elżbieta; Hapanowicz, Jerzy; Jagodzińska, Dorota; Kemona, Andrzej; Guzińska-Ustymowicz, Katarzyna

2016-01-01

Crohn's disease (CD) and ulcerative colitis (UC) belong to a group of inflammatory bowel diseases (IBD). The aim of our study was to evaluate the expression of MMP-2, MMP-7, MMP-9, TIMP-1, and TIMP-2 in ulcerative colitis and Crohn's disease. The study group comprised 34 patients with UC and 10 patients with CD. Evaluation of MMP-2, MMP-7, MMP-9, TIMP-1, and TIMP-2 expression in tissue samples was performed using immunohistochemistry. The overexpression of MMP-9 and TIMP-1 was dominant in both the glandular epithelium and inflammatory infiltration in UC patients. In contrast, in CD subjects the positive expression of MMP-2 and TIMP-1 was in glandular tubes while mainly MMP-7 and TIMP-2 expression was in inflammatory infiltration. Metalloproteinases' expression was associated with the presence of erosions, architectural tissue changes, and inflammatory infiltration in the lamina propria of UC patients. The expression of metalloproteinase inhibitors correlated with the presence of eosinophils and neutrophils in UC and granulomas in CD patients. Our studies indicate that the overexpression of metalloproteinases and weaker expression of their inhibitors may determine the development of IBD. It appears that MMP-2, MMP-7, and MMP-9 may be a potential therapeutic target and the use of their inhibitors may significantly reduce UC progression. PMID:27034654

Expressions of Matrix Metalloproteinases (MMP-2, MMP-7, and MMP-9) and Their Inhibitors (TIMP-1, TIMP-2) in Inflammatory Bowel Diseases.

PubMed

Jakubowska, Katarzyna; Pryczynicz, Anna; Iwanowicz, Piotr; Niewiński, Andrzej; Maciorkowska, Elżbieta; Hapanowicz, Jerzy; Jagodzińska, Dorota; Kemona, Andrzej; Guzińska-Ustymowicz, Katarzyna

2016-01-01

Crohn's disease (CD) and ulcerative colitis (UC) belong to a group of inflammatory bowel diseases (IBD). The aim of our study was to evaluate the expression of MMP-2, MMP-7, MMP-9, TIMP-1, and TIMP-2 in ulcerative colitis and Crohn's disease. The study group comprised 34 patients with UC and 10 patients with CD. Evaluation of MMP-2, MMP-7, MMP-9, TIMP-1, and TIMP-2 expression in tissue samples was performed using immunohistochemistry. The overexpression of MMP-9 and TIMP-1 was dominant in both the glandular epithelium and inflammatory infiltration in UC patients. In contrast, in CD subjects the positive expression of MMP-2 and TIMP-1 was in glandular tubes while mainly MMP-7 and TIMP-2 expression was in inflammatory infiltration. Metalloproteinases' expression was associated with the presence of erosions, architectural tissue changes, and inflammatory infiltration in the lamina propria of UC patients. The expression of metalloproteinase inhibitors correlated with the presence of eosinophils and neutrophils in UC and granulomas in CD patients. Our studies indicate that the overexpression of metalloproteinases and weaker expression of their inhibitors may determine the development of IBD. It appears that MMP-2, MMP-7, and MMP-9 may be a potential therapeutic target and the use of their inhibitors may significantly reduce UC progression.

Dihydroartiminisin inhibits the growth and metastasis of epithelial ovarian cancer.

PubMed

Wu, Buchu; Hu, Ke; Li, Shu; Zhu, Jing; Gu, Liying; Shen, Haoran; Hambly, Brett D; Bao, Shisan; Di, Wen

2012-01-01

Dihydroartiminisin (DHA), the active component of a Chinese herb (Artemisia annua), has been utilised as an anti-malarial drug since ancient China. DHA has also been shown to inhibit proliferation of cancer in vitro. However, the capacity of DHA to inhibit the development of ovarian cancer is still unclear. The adhesion, invasion, and migration of human ovarian cancer cell line (HO8910PM) was determined following DHA treatment in vitro, using Matrigel coated plate, transwell membrane chamber, and wound healing models, respectively. A mouse ovarian cancer model was established by orthotopic inoculation of HO8910PM cell line in nude mice. The growth and metastasis in vivo was determined 8 weeks post-implantation in response to DHA treatment. The expression of phosphorylated focal adhesion kinase (pFAK) and matrix metalloproteinases (MMP-2 and MMP-9) was evaluated using Western blotting. The expression of Von Willebrand factor (vWF) and infiltration of macrophages were determined, using immunohistochemistry. DHA inhibits ovarian cancer cell proliferation, adhesion, migration and invasion in vitro in a dose-dependent manner, consistent with decreased expression of pFAK and MMP-2, but not MMP-9. DHA inhibited metastasis significantly in vivo, associated with reduced vWF expression and macrophage infiltration. In conclusion, DHA inhibits the development of ovarian cancer, in part via down-regulating pFAK, MMP-2, vWF and macrophage infiltration.

Interleukin-1β and cyclic AMP mediate the invasion of sheared chondrosarcoma cells via a matrix metalloproteinase-1-dependent mechanism.

PubMed

Wang, Pu; Guan, Pei-Pei; Wang, Tao; Yu, Xin; Guo, Jian-Jun; Konstantopoulos, Konstantinos; Wang, Zhan-You

2014-05-01

Matrix metalloproteinase-1 (MMP-1) is a potential biomarker for chondrosarcoma that is overexpressed at the invading edges of articular cartilage, and its expression correlates with poor survival rates. However, the molecular mechanisms of MMP-1 regulation and its potential contribution to chondrosarcoma cell invasion have yet to be elucidated, especially in shear-activated cells. Using molecular biology tools and an in vitro fluid shear model, we report that shear stress upregulates cyclic AMP (cAMP) and interleukin-1β (IL-1β) release, which in turn promotes the invasion of chondrosarcoma cells via the induction of MMP-1 in a phosphoinositide 3-kinase (PI3-K)- and ERK1/2-dependent manner. Activated PI3-K and ERK1/2 signaling pathways phosphorylate c-Jun, which in turn transactivates MMP-1 in human chondrosarcoma cells. Collectively, fluid shear stress upregulates matrix MMP-1 expression, which is responsible for the enhanced invasion of human chondrosarcoma cells. Copyright © 2014 Elsevier B.V. All rights reserved.

Cell-Responsive Hydrogel for Encapsulation of Vascular Cells

PubMed Central

Kraehenbuehl, Thomas P.; Ferreira, Lino S.; Zammaretti, Prisca; Hubbell, Jeffrey A.; Langer, Robert

2014-01-01

The in vitro potential of a synthetic matrix metalloproteinase (MMP)-responsive polyethylene glycol) (PEG)-based hydrogel as a bioactive co-encapsulation system for vascular cells and a small bioactive peptide, thymosin β4 (Tp4), was examined. We show that the physical incorporation of Tβ4 in this bioactive matrix creates a three-dimensional (3D) environment conducive for human umbilical vein endothelial cell (HUVEC) adhesion, survival, migration and organization. Gels with entrapped Tβ4 increased the survival of HUVEC compared to gels without Tp4, and significantly up-regulated the endothelial genes vascular endothelial-cadherin and angiopoietin-2, whereas von Willebrand factor was significantly down-regulated. Incorporation of Tβ4 significantly increased MMP-2 and MMP-9 secretion of encapsulated HUVEC. The gel acts as a controlled Tβ4-release system, as MMP-2 and MMP-9 enzymes trigger the release. In addition, Tβ4 facilitated HUVEC attachment and induced vascular-like network formation upon the PEG-hydrogels. These MMP-responsive PEG-hydrogels may thus serve as controlled co-encapsulation system of vascular cells and bioactive factors for in situ regeneration of ischemic tissues. PMID:19500842

Stem cell factor supports migration in canine mesenchymal stem cells.

PubMed

Enciso, Nathaly; Ostronoff, Luciana L K; Mejías, Guillermo; León, Leticia G; Fermín, María Luisa; Merino, Elena; Fragio, Cristina; Avedillo, Luis; Tejero, Concepción

2018-03-01

Adult Mesenchymal Stem Cells (MSC) are cells that can be defined as multipotent cells able to differentiate into diverse lineages, under appropriate conditions. These cells have been widely used in regenerative medicine, both in preclinical and clinical settings. Initially discovered in bone marrow, MSC can now be isolated from a wide spectrum of adult and foetal tissues. Studies to evaluate the therapeutic potential of these cells are based on their ability to arrive to damaged tissues. In this paper we have done a comparative study analyzing proliferation, surface markers and OCT4, SOX9, RUNX2, PPARG genes expression in MSC cells from Bone marrow (BMMSC) and Adipose tissue (ASC). We also analyzed the role of Stem Cell Factor (SCF) on MSC proliferation and on ASCs metalloproteinases MMP-2, MMP-9 secretion. Healthy dogs were used as BMMSC donors, and ASC were collected from omentum during elective ovariohysterectomy surgery. Both cell types were cultured in IMDM medium with or without SCF, 10% Dog Serum (DS), and incubated at 38 °C with 5% CO2. Growth of BMMSCs and ASCs was exponential until 25-30 days. Flow citometry of MSCs revealed positive results for CD90 and negative for CD34, CD45 and MCH-II. Genes were evaluated by RT-PCR and metalloproteinases by zymografy. Our findings indicate morphological and immunological similarities as well as expression of genes from both origins on analyzed cells. Furthermore, SCF did not affect proliferation of MSCs, however it up-regulated MMP-2 and MMP-9 secretion in ASCs. These results suggest that metalloproteinases are possibly essential molecules pivoting migration.

GPNMB/OA protein increases the invasiveness of human metastatic prostate cancer cell lines DU145 and PC3 through MMP-2 and MMP-9 activity

DOE Office of Scientific and Technical Information (OSTI.GOV)

Fiorentini, Chiara; Bodei, Serena; Bedussi, Francesca

2014-04-15

Non-metastatic glycoprotein melanoma protein B (GPNMB), also known as osteoactivin (OA) is expressed in a wide array of tumors and represents an emerging target for drug development. In this study, we investigated the role of GPNMB/OA in the progression of human metastatic DU145 and PC3 prostate cancer cells. GPNMB/OA contribution in PCa malignant phenotype has been analyzed by small interfering RNA-induced GPNMB/OA silencing. We found that following GPNMB/OA silencing the migration capability of both DU145 and PC3 cells, evaluated by using in vitro invasivity assay, as well as the metalloproteinases MMP-2 and MMP-9 activity were equally strongly inhibited. By contrastmore » knocking down GPNMB/OA weakly attenuated cell proliferation rate of DU145, an effect that paralleled with an increase number of apoptotic cells. However, PC3 cell growth seems to be not affected by GPNMB/OA. Together, these data reveal that GPNMB/OA acts as a critical molecular mediator promoting the acquisition of the more aggressive, pro-metastatic phenotype distinctive of human DU145 and PC3 cell lines. - Highlights: • GPNMB/OA expression correlates with DU145 and PC3 cells malignant phenotype. • GPNMB/OA silencing affects the migration capability of both DU145 and PC3 cells. • GPNMB/OA increases invasiveness by up-regulating MMPs activity. • GPNMB/OA promotes DU145 and PC3 cells progression into a more aggressive phenotype.« less

The epigenetic regulation of SOX9 by miR-145 in human chondrosarcoma.

PubMed

Mak, Isabella W Y; Singh, Shalini; Turcotte, Robert; Ghert, Michelle

2015-01-01

Chondrosarcoma is the most common primary bone malignancy in the adult population with a high rate of pulmonary metastasis. Chondrosarcoma is managed with surgical excision as the tumors do not respond well to conventional chemotherapy or radiation therapy. Thus, there exists a dire need to develop systemic treatment options to target chondrosarcoma cells for metastatic spread. We hypothesized that the expression of miR-145 is low in chondrosarcoma, leading to decreased transcriptional control of SOX9 (the master regulator of chondrogenesis), and downstream activation of the transcription factor ETV5. We have previously shown that ETV5 activates MMP-2 expression in chondrosarcoma, which in turn increases local bone matrix resorption. In this study, we confirm high expression of SOX9 in human chondrosarcoma using real-time PCR, Western blotting, and immunofluorescence. An ETV5 promoter-reporter plasmid was transfected into chondrosarcoma cells to determine if SOX9 directly regulates the expression of ETV5. Co-transfection of the ETV5 promoter-plasmid with SOX9 lentivirus significantly increased the luciferase activity derived from the ETV5 promoter, from which the regulatory relationship between SOX9 and ETV5 is established. MiR-145 was found to be down-regulated in chondrosarcoma cell lines, patient samples, and further confirmed with a public sarcoma database. After stable miR-145 lentiviral transfection, the subsequent mRNA expression levels of SOX9, ETV5, and MMP-2 were significantly decreased in chondrosarcoma cells. The results generated by this study may have important clinical significance in the treatment of patients with chondrosarcoma in that targeted miRNA may have the potential to downregulate the upstream activators of proteases such as MMP-2. © 2014 Wiley Periodicals, Inc.

Effects of non-ablative fractional erbium glass laser treatment on gene regulation in human three-dimensional skin models.

PubMed

Amann, Philipp M; Marquardt, Yvonne; Steiner, Timm; Hölzle, Frank; Skazik-Voogt, Claudia; Heise, Ruth; Baron, Jens M

2016-04-01

Clinical experiences with non-ablative fractional erbium glass laser therapy have demonstrated promising results for dermal remodelling and for the indications of striae, surgical scars and acne scars. So far, molecular effects on human skin following treatment with these laser systems have not been elucidated. Our aim was to investigate laser-induced effects on skin morphology and to analyse molecular effects on gene regulation. Therefore, human three-dimensional (3D) organotypic skin models were irradiated with non-ablative fractional erbium glass laser systems enabling qRT-PCR, microarray and histological studies at same and different time points. A decreased mRNA expression of matrix metalloproteinases (MMPs) 3 and 9 was observed 3 days after treatment. MMP3 also remained downregulated on protein level, whereas the expression of other MMPs like MMP9 was recovered or even upregulated 5 days after irradiation. Inflammatory gene regulatory responses measured by the expression of chemokine (C-X-C motif) ligands (CXCL1, 2, 5, 6) and interleukin expression (IL8) were predominantly reduced. Epidermal differentiation markers such as loricrin, filaggrin-1 and filaggrin-2 were upregulated by both tested laser optics, indicating a potential epidermal involvement. These effects were also shown on protein level in the immunofluorescence analysis. This novel standardised laser-treated human 3D skin model proves useful for monitoring time-dependent ex vivo effects of various laser systems on gene expression and human skin morphology. Our study reveals erbium glass laser-induced regulations of MMP and interleukin expression. We speculate that these alterations on gene expression level could play a role for dermal remodelling, anti-inflammatory effects and increased epidermal differentiation. Our finding may have implications for further understanding of the molecular mechanism of erbium glass laser-induced effects on human skin.

Enhanced cyclooxygenase-2 expression levels and metalloproteinase 2 and 9 activation by Hexachlorobenzene in human endometrial stromal cells.

PubMed

Chiappini, Florencia; Bastón, Juan Ignacio; Vaccarezza, Agustina; Singla, José Javier; Pontillo, Carolina; Miret, Noelia; Farina, Mariana; Meresman, Gabriela; Randi, Andrea

2016-06-01

Hexachlorobenzene (HCB) is an organochlorine pesticide that induces toxic reproductive effects in laboratory animals. It is a dioxin-like compound and a weak ligand of the aryl hydrocarbon receptor (AhR). Endometriosis is characterized by the presence of functional endometrial tissues outside the uterine cavity. Experimental studies indicate that exposure to organochlorines can interfere with both hormonal regulation and immune function to promote endometriosis. Altered expression of metalloproteinases (MMPs) in patients with endometriosis, suggests that MMPs may play a critical role. In the endometriotic lesions, prostaglandin E2 (PGE2) produced by cyclooxygenase-2 (COX-2), binds to its EP4 receptor (EP4), and via c-Src kinase induces MMPs activation, promoting endometriosis. We examined the HCB action on MMP-2 and MMP-9 activities and expression, COX-2 levels, PGE2 signaling, and the AhR involvement in HCB-induced effects. We have used different in vitro models: (1) human endometrial stromal cell line T-HESC, (2) primary cultures of Human Uterine Fibroblast (HUF), and (3) primary cultures of endometrial stromal cells from eutopic endometrium of control (CESC) and subjects with endometriosis (EESC). Our results show that HCB enhances MMP-2 and MMP-9 activities in T-HESC, HUF and ESC cells. The MMP-9 levels were elevated in all models, while the MMP-2 expression only increased in ESC cells. HCB enhanced COX-2 and EP4 expression, PGE2 secretion and the c-Src kinase activation in T-HESC. Besides, we observed that AhR is implicated in these HCB-induced effects. In conclusion, our results show that HCB exposure could contribute to endometriosis development, affecting inflammation and invasion parameters of human endometrial cells. Copyright © 2016 Elsevier Inc. All rights reserved.

Disentangling the multifactorial contributions of fibronectin, collagen and cyclic strain on MMP expression and extracellular matrix remodeling by fibroblasts.

PubMed

Zhang, Yang; Lin, Zhe; Foolen, Jasper; Schoen, Ingmar; Santoro, Alberto; Zenobi-Wong, Marcy; Vogel, Viola

2014-11-01

Early wound healing is associated with fibroblasts assembling a provisional fibronectin-rich extracellular matrix (ECM), which is subsequently remodeled and interlaced by type I collagen. This exposes fibroblasts to time-variant sets of matrices during different stages of wound healing. Our goal was thus to gain insight into the ECM-driven functional regulation of human foreskin fibroblasts (HFFs) being either anchored to a fibronectin (Fn) or to a collagen-decorated matrix, in the absence or presence of cyclic mechanical strain. While the cells reoriented in response to the onset of uniaxial cyclic strain, cells assembled exogenously added Fn with a preferential Fn-fiber alignment along their new orientation. Exposure of HFFs to exogenous Fn resulted in an increase in matrix metalloproteinase (MMP) expression levels, i.e. MMP-15 (RT-qPCR), and MMP-9 activity (zymography), while subsequent exposure to collagen slightly reduced MMP-15 expression and MMP-9 activity compared to Fn-exposure alone. Cyclic strain upregulated Fn fibrillogenesis and actin stress fiber formation, but had comparatively little effect on MMP activity. We thus propose that the appearance of collagen might start to steer HFFs towards homeostasis, as it decreased both MMP secretion and the tension of Fn matrix fibrils as assessed by Fluorescence Resonance Energy Transfer. These results suggest that HFFs might have a high ECM remodeling or repair capacity in contact with Fn alone (early event), which is reduced in the presence of Col1 (later event), thereby down-tuning HFF activity, a processes which would be required in a tissue repair process to finally reach tissue homeostasis. Copyright © 2014. Published by Elsevier B.V.

Cardiotrophin-1 Induces Matrix Metalloproteinase-1 in Human Aortic Endothelial Cells

PubMed Central

Tokito, Akinori; Jougasaki, Michihisa; Ichiki, Tomoko; Hamasaki, Shuichi

2013-01-01

Rupture of an atherosclerotic plaque is a key event in the development of cardiovascular disorders, in which matrix metalloproteinase-1 (MMP-1) plays a crucial role by degradation of extracellular matrix resulting in plaque instability. Cardiotrophin-1 (CT-1), a member of interleukin-6-type proinflammatory cytokines, has potent cardiovascular actions and is highly expressed in vascular endothelium, however its role in atherosclerosis has not been fully elucidated to date. The present study was designed to investigate whether CT-1 induces MMP-1 in human aortic endothelial cells (HAECs). Ribonuclease protection assay demonstrated that MMP-1 gene level in HAECs was enhanced by the treatment of CT-1 in a dose- and time-dependent manner. Immunocytochemical staining, Western immunoblot analysis and enzyme-linked immunosorbent assay revealed that CT-1 augmented MMP-1 protein synthesis and secretion. MMP-1 activity assay revealed that MMP-1 present in the supernatant of HAECs was exclusively precursor form. Casein zymography disclosed proteolytic activity in the supernatant of HAECs, which was enhanced by CT-1 treatment. Furthermore, pharmacological inhibitor study indicated the important roles of extracellular signal-regulated kinase (ERK) 1/2, p38 mitogen-activated protein (MAP) kinase, c-Jun N-terminal kinase (JNK) and Janus kinase/signal transducers and activators of transcription (JAK/STAT) signaling pathways in mediating CT-1-induced MMP-1 gene and protein expression. These data reveal for the first time that CT-1 induces the proteolytic potential in HAECs by upregulating MMP-1 expression through ERK1/2, p38 MAP kinase, JNK and JAK/STAT pathways, and suggest that CT-1 may play an important role in the pathophysiology of atherosclerosis and plaque instability. PMID:23935888

Changes in matrix metalloproteinase network in a spontaneous autoimmune uveitis model.

PubMed

Hofmaier, Florian; Hauck, Stefanie M; Amann, Barbara; Degroote, Roxane L; Deeg, Cornelia A

2011-04-08

Autoimmune uveitis is a sight-threatening disease in which autoreactive T cells cross the blood-retinal barrier. Molecular mechanisms contributing to the loss of eye immune privilege in this autoimmune disease are not well understood. In this study, the authors investigated the changes in the matrix metalloproteinase network in spontaneous uveitis. Matrix metalloproteinase (MMP) MMP2, MMP9, and MMP14 expression and tissue inhibitor of metalloproteinase (TIMP)-2 and lipocalin 2 (LCN2) expression were analyzed using Western blot quantification. Enzyme activities were examined with zymography. Expression patterns of network candidates were revealed with immunohistochemistry, comparing physiological appearance and changes in a spontaneous recurrent uveitis model. TIMP2 protein expression was found to be decreased in both the vitreous and the retina of a spontaneous model for autoimmune uveitis (equine recurrent uveitis [ERU]), and TIMP2 activity was significantly reduced in ERU vitreous. Functionally associated MMPs such as MMP2, MMP14, and MMP9 were found to show altered or shifted expression and activity. Although MMP2 decreased in ERU vitreous, MMP9 expression and activity were found to be increased. These changes were reflected by profound changes within uveitic target tissue, where TIMP2, MMP9, and MMP14 decreased in expression, whereas MMP2 displayed a shifted expression pattern. LCN2, a potential stabilizer of MMP9, was found prominently expressed in equine healthy retina and displayed notable changes in expression patterns accompanied by significant upregulation in autoimmune conditions. Invading cells expressed MMP9 and LCN2. This study implicates a dysregulation or a change in functional protein-protein interactions in this TIMP2-associated protein network, together with altered expression of functionally related MMPs.

Single nucleotide polymorphism-specific regulation of matrix metalloproteinase-9 by multiple miRNAs targeting the coding exon

PubMed Central

Duellman, Tyler; Warren, Christopher; Yang, Jay

2014-01-01

Microribonucleic acids (miRNAs) work with exquisite specificity and are able to distinguish a target from a non-target based on a single nucleotide mismatch in the core nucleotide domain. We questioned whether miRNA regulation of gene expression could occur in a single nucleotide polymorphism (SNP)-specific manner, manifesting as a post-transcriptional control of expression of genetic polymorphisms. In our recent study of the functional consequences of matrix metalloproteinase (MMP)-9 SNPs, we discovered that expression of a coding exon SNP in the pro-domain of the protein resulted in a profound decrease in the secreted protein. This missense SNP results in the N38S amino acid change and a loss of an N-glycosylation site. A systematic study demonstrated that the loss of secreted protein was due not to the loss of an N-glycosylation site, but rather an SNP-specific targeting by miR-671-3p and miR-657. Bioinformatics analysis identified 41 SNP-specific miRNA targeting MMP-9 SNPs, mostly in the coding exon and an extension of the analysis to chromosome 20, where the MMP-9 gene is located, suggesting that SNP-specific miRNAs targeting the coding exon are prevalent. This selective post-transcriptional regulation of a target messenger RNA harboring genetic polymorphisms by miRNAs offers an SNP-dependent post-transcriptional regulatory mechanism, allowing for polymorphic-specific differential gene regulation. PMID:24627221

Suppressed invasive and migratory behaviors of SW1353 chondrosarcoma cells through the regulation of Src, Rac1 GTPase, and MMP13.

PubMed

Xu, Wenxiao; Wan, Qiaoqiao; Na, Sungsoo; Yokota, Hiroki; Yan, Jing-Long; Hamamura, Kazunori

2015-12-01

Chondrosarcoma is the second frequent type of primary bone cancer. In response to stress to the endoplasmic reticulum, activation of eIF2α-mediated signaling is reported to induce apoptosis. However, its effects on invasive and migratory behaviors of chondrosarcoma have not been understood. Focusing on potential roles of Src kinase, Rac1 GTPase, and MMP13, we investigated eIF2α-driven regulation of SW1353 chondrosarcoma cells. In particular, we employed two chemical agents (salubrinal, Sal; and guanabenz, Gu) that elevate the level of eIF2α phosphorylation. The result revealed that both Sal and Gu reduced invasion and motility of SW1353 chondrosarcoma cells in a dose dependent manner. Live imaging using a fluorescent resonance energy transfer (FRET) technique showed that Sal and Gu downregulated activities of Src kinase as well as Rac1 GTPase in an eIF2α dependent manner. RNA interference experiments supported an eIF2α-mediated regulatory network in the inhibitory role of Sal and Gu. Partial silencing of MMP13 also suppressed malignant phenotypes of SW1353 chondrosarcoma cells. However, MMP13 was not regulated via eIF2α since administration of Sal but not Gu reduced expression of MMP13. In summary, we demonstrate that eIF2α dependent and independent pathways regulate invasion and motility of SW1353 chondrosarcoma cells, and inactivation of Src, Rac1, and MMP13 by Sal could provide a potential adjuvant therapy for combating metastatic chondrosarcoma cells. Copyright © 2015 Elsevier Inc. All rights reserved.

Insights on ornithine decarboxylase silencing as a potential strategy for targeting retinoblastoma.

PubMed

Muthukumaran, Sivashanmugam; Bhuvanasundar, Renganathan; Umashankar, Vetrivel; Sulochana, K N

2018-02-01

Ornithine Decarboxylase (ODC) is a key enzyme involved in polyamine synthesis and is reported to be up regulated in several cancers. However, the effect of ODC gene silencing in retinoblastoma is to be understood for utilization in therapeutic applications. Hence, in this study, a novel siRNA (small interference RNA) targeting ODC was designed and validated in Human Y79 retinoblastoma cells for its effects on intracellular polyamine levels, Matrix Metalloproteinase 2 & 9 activity and Cell cycle. The designed siRNA showed efficient silencing of ODC mRNA expression and protein levels in Y79 cells. It also showed significant reduction of intracellular polyamine levels and altered levels of oncogenic LIN28b expression. By this study, a regulatory loop is proposed, wherein, ODC silencing in Y79 cells to result in decreased polyamine levels, thereby, leading to altered protein levels of Lin28b, MMP-2 and MMP-9, which falls in line with earlier studies in neuroblastoma. Thus, by this study, we propose ODC silencing as a prospective strategy for targeting retinoblastoma. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

Homocysteine enhances MMP-9 production in murine macrophages via ERK and Akt signaling pathways

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lee, Seung Jin; Lee, Yi Sle; Seo, Kyo Won

2012-04-01

Homocysteine (Hcy) at elevated levels is an independent risk factor of cardiovascular diseases, including atherosclerosis. In the present study, we investigated the effect of Hcy on the production of matrix metalloproteinases (MMP) in murine macrophages. Among the MMP known to regulate the activities of collagenase and gelatinase, Hcy exclusively increased the gelatinolytic activity of MMP-9 in J774A.1 cells as well as in mouse peritoneal macrophages. Furthermore, this activity was found to be correlated with Western blot findings in J774A.1 cells, which showed that MMP-9 expression was concentration- and time-dependently increased by Hcy. Inhibition of the ERK and Akt pathways ledmore » to a significant decrease in Hcy-induced MMP-9 expression, and combined treatment with inhibitors of the ERK and Akt pathways showed an additive effects. Activity assays for ERK and Akt showed that Hcy increased the phosphorylation of both, but these phosphorylation were not affected by inhibitors of the Akt and ERK pathways. In line with these findings, the molecular inhibition of ERK and Akt using siRNA did not affect the Hcy-induced phosphorylation of Akt and ERK, respectively. Taken together, these findings suggest that Hcy enhances MMP-9 production in murine macrophages by separately activating the ERK and Akt signaling pathways. -- Highlights: ► Homocysteine (Hcy) induced MMP-9 production in murine macrophages. ► Hcy induced MMP-9 production through ERK and Akt signaling pathways. ► ERK and Akt signaling pathways were activated by Hcy in murine macrophages. ► ERK and Akt pathways were additively act on Hcy-induced MMP-9 production. ► Hcy enhances MMP-9 production in macrophages via activation of ERK and Akt signaling pathways in an independent manner.« less

Effects of mechanical strain on human mesenchymal stem cells and ligament fibroblasts in a textured poly(L-lactide) scaffold for ligament tissue engineering.

PubMed

Kreja, Ludwika; Liedert, Astrid; Schlenker, Heiter; Brenner, Rolf E; Fiedler, Jörg; Friemert, Benedikt; Dürselen, Lutz; Ignatius, Anita

2012-10-01

The purpose of this study was to prove the effect of cyclic uniaxial intermittent strain on the mRNA expression of ligament-specific marker genes in human mesenchymal stem cells (MSC) and anterior cruciate ligament-derived fibroblasts (ACL-fibroblasts) seeded onto a novel textured poly(L-lactide) scaffold (PLA scaffold). Cell-seeded scaffolds were mechanically stimulated by cyclic uniaxial stretching. The expression of ligament matrix gene markers: collagen types I and III, fibronectin, tenascin C and decorin, as well as the proteolytic enzymes matrix metalloproteinase MMP-1 and MMP-2 and their tissue specific inhibitors TIMP-1 and TIMP-2 was investigated by analysing the mRNA expression using reverse transcriptase polymerase chain reaction and related to the static control. In ACL-fibroblasts seeded on PLA, mechanical load induced up-regulation of collagen types I and III, fibronectin and tenascin C. No effect of mechanical stimulation on the expression of ligament marker genes was found in undifferentiated MSC seeded on PLA. The results indicated that the new textured PLA scaffold could transfer the mechanical load to the ACL-fibroblasts and improved their ligament phenotype. This scaffold might be suitable as a cell-carrying component of ACL prostheses.

Impaired CD23 and CD62L expression and tissue inhibitors of metalloproteinases secretion by eosinophils in adults with atopic dermatitis.

PubMed

de Oliveira Titz, T; Orfali, R L; de Lollo, C; Dos Santos, V G; da Silva Duarte, A J; Sato, M N; Aoki, V

2016-12-01

Eosinophils are multifunctional, polymorphonuclear leucocytes that secrete proteins within cytoplasmic granules, such as cytokines, chemokines, metalloproteinases (MMPs) and metalloproteinases tissue inhibitors (TIMPs). Although eosinophilia is a hallmark of atopic dermatitis (AD), several functional aspects of eosinophils remain unknown. We aimed to evaluate the phenotype and functional response of eosinophils under staphylococcal enterotoxin B (SEB) and Toll-like receptor (TLR)-2/6 (FSL-1) stimulation in the secretion of CCL5, MMPs and TIMPs in adults with AD. Forty-one adult patients with AD and 45 healthy controls enrolled for the study. Phenotype of eosinophils from granulocytes of peripheral blood was analysed by flow cytometry. We performed evaluation of CCL5 (cytometric bead array), MMP and TIMP (ELISA) secretion, in culture supernatants of purified eosinophils stimulated with SEB or TLR2/6 agonist (FSL-1). We found a higher frequency of LIN1 - CCR3 + eosinophils, and decreased expression of CD23 and CD62L receptors in eosinophils of AD patients. There was no difference in MMP and TIMP serum levels between the evaluated groups. However, we detected decreased basal levels of TIMP-1, TIMP-2 and CCL5 in culture supernatants from purified, unstimulated eosinophils from AD patients. In adults with AD, phenotypical features of eosinophils reveal decreased expression of early activation and L-selectin receptors. Regarding the functional profile of purified eosinophils related to tissue remodelling in atopic dermatitis, innate immune stimulation (TLR2/6 agonist and SEB) did not affect the ratio of MMP/TIMPs secretion in AD. Our findings reinforce the potential breakdown in tissue remodelling process mediated by eosinophils in AD. © 2016 European Academy of Dermatology and Venereology.

Stimulation of epithelial cell matrix metalloproteinase (MMP-2, -9, -13) and interleukin-8 secretion by fusobacteria.

PubMed

Gursoy, U K; Könönen, E; Uitto, V-J

2008-10-01

Bacterial pathogens involved in periodontal diseases exert their destructive effects primarily by stimulating the host cells to increase their secretion of proinflammatory cytokines and matrix metalloproteinases (MMPs). This study aimed to determine the epithelial cell matrix metalloproteinase and interleukin-8 (IL-8) secretion upon exposure to fusobacteria. Eight different oral and non-oral Fusobacterium strains were incubated with HaCaT epithelial cells. Gelatin zymography and Western blot analysis were performed to detect collagenase 3 (MMP-13), gelatinase A (MMP-2), gelatinase B (MMP-9), and IL-8 secretion by epithelial cells. All Fusobacterium strains, especially Fusobacterium necrophorum ATCC 25286, Fusobacterium nucleatum ATCC 25586, and Fusobacterium varium ATCC 51644, increased MMP-9 and MMP-13 secretion. Fusobacterium simiae ATCC 33568, and to a lesser extent F. nucleatum and F. necrophorum, increased epithelial MMP-2 secretion. F. nucleatum and F. necrophorum also increased IL-8 secretion. F. varium ATCC 27725, a strain that only weakly stimulated MMP production, strongly increased the IL-8 production, suggesting that their expression is differently regulated. We conclude that the pathogenic potential of fusobacteria may partly result from their ability to stimulate secretion of MMP-9, MMP-13, and IL-8 from epithelial cells.

MMP-9 in translation: from molecule to brain physiology, pathology, and therapy.

PubMed

Vafadari, Behnam; Salamian, Ahmad; Kaczmarek, Leszek

2016-10-01

Matrix metalloproteinase-9 (MMP-9) is a member of the metzincin family of mostly extracellularly operating proteases. Despite the fact that all of these enzymes might be target promiscuous, with largely overlapping catalogs of potential substrates, MMP-9 has recently emerged as a major and apparently unique player in brain physiology and pathology. The specificity of MMP-9 may arise from its very local and time-restricted actions, even when released in the brain from cells of various types, including neurons, glia, and leukocytes. In fact, the quantity of MMP-9 is very low in the naive brain, but it is markedly activated at the levels of enzymatic activity, protein abundance, and gene expression following various physiological stimuli and pathological insults. Neuronal MMP-9 participates in synaptic plasticity by controlling the shape of dendritic spines and function of excitatory synapses, thus playing a pivotal role in learning, memory, and cortical plasticity. When improperly unleashed, MMP-9 contributes to a large variety of brain disorders, including epilepsy, schizophrenia, autism spectrum disorder, brain injury, stroke, neurodegeneration, pain, brain tumors, etc. The foremost mechanism of action of MMP-9 in brain disorders appears to be its involvement in immune/inflammation responses that are related to the enzyme's ability to process and activate various cytokines and chemokines, as well as its contribution to blood-brain barrier disruption, facilitating the extravasation of leukocytes into brain parenchyma. However, another emerging possibility (i.e., the control of MMP-9 over synaptic plasticity) should not be neglected. The translational potential of MMP-9 has already been recognized in both the diagnosis and treatment domains. The most striking translational aspect may be the discovery of MMP-9 up-regulation in a mouse model of Fragile X syndrome, quickly followed by human studies and promising clinical trials that have sought to inhibit MMP-9. With regard to diagnosis, suggestions have been made to use MMP-9 alone or combined with tissue inhibitor of matrix metalloproteinase-1 or brain-derived neurotrophic factor as disease biomarkers. MMP-9, through cleavage of specific target proteins, plays a major role in synaptic plasticity and neuroinflammation, and by those virtues contributes to brain physiology and a host of neurological and psychiatric disorders. This article is part of the 60th Anniversary special issue. © 2016 International Society for Neurochemistry.

Effects of Detergents on Catalytic Activity of Human Endometase/Matrilysin-2, a Putative Cancer Biomarker†

PubMed Central

Park, Hyun I.; Lee, Seakwoo; Ullah, Asad; Cao, Qiang; Sang, Qing-Xiang Amy

2009-01-01

Matrix metalloproteinases (MMPs) are a family of hydrolytic enzymes that play significant roles in development, morphogenesis, inflammation, and cancer invasion. Endometase (matrilysin 2 or MMP-26) is a putative early biomarker for human carcinomas. The effects of the ionic and nonionic detergents on catalytic activity of endometase were investigated. The hydrolytic activity of endometase was detergent concentration-dependent exhibiting a bell-shaped curve with its maximum activity near the critical micelle concentration (CMC) of nonionic detergents tested. The effect of Brij-35 on human gelatinase B (MMP-9), matirilysin (MMP-7), and membrane-type 1 MMP (MT1-MMP) was further explored. Their maximum catalysis was observed near the CMC of Brij-35 (~90 μM). Their IC50 values were above the CMC. The inhibition mechanism of MMP-7, MMP-9, and MT1-MMP by Brij-35 was mixed-type as determined by Dixon’s plot, however, that of endometase was non-competitive with a Ki value of 240 μM. The catalytic activities of MMPs are influenced by detergents. Monomer of detergents may activate and stabilize MMPs to enhance catalysis, but micelle of detergents may sequester enzyme and block substrate binding site to impede catalysis. Under physiological conditions lipid or membrane microenvironment may regulate enzymatic activity. PMID:19818727

Expression of matrix metalloproteinase-1, -2 and -3 in squamous cell carcinoma and actinic keratosis

PubMed Central

Tsukifuji, R; Tagawa, K; Hatamochi, A; Shinkai, H

1999-01-01

Matrix metalloproteinase (MMP) plays an important role in extracellular matrix degradation associated with cancer invasion. An expression of MMP-1 (interstitial collagenase), MMP-2 (72-kDa type IV collagenase) and MMP-3 (stromelysin-1) was investigated in squamous cell carcinoma (SCC) and its precancerous condition, actinic keratosis (AK), using in situ hybridization techniques. MMP-1 mRNA was detected in tumour cells and/or in stromal cells in all cases of SCC, four of six AKs adjacent to SCC and four of 16 AKs. MMP-2 and MMP-3 mRNAs were detected in SCC but not in AK. The expression of MMP-3 correlated to that of MMP-1 (P = 0.03) localized at the tumour mass and stroma of the invasive area, while MMP-2 mRNA was detected widely throughout the stroma independent of MMP-1 expression. Our results indicated that the expression of MMP-1, -2 and -3 showed different localization patterns, suggesting a unique role of each MMP in tumour progression. Moreover, MMP-1 expression could be an early event in the development of SCC, and AK demonstrating MMP-1 mRNA, might be in a more advanced dysplastic state, progressing to SCC. © 1999 Cancer Research Campaign PMID:10362121

miR-26a regulates mouse hepatocyte proliferation via directly targeting the 3' untranslated region of CCND2 and CCNE2.

PubMed

Zhou, Jian; Ju, Wei-Qiang; Yuan, Xiao-Peng; Zhu, Xiao-Feng; Wang, Dong-Ping; He, Xiao-Shun

2016-02-01

The deficiency of liver regeneration needs to be addressed in the fields of liver surgery, split liver transplantation and living donor liver transplantation. Researches of microRNAs would broaden our understandings on the mechanisms of various diseases. Our previous research confirmed that miR-26a regulated liver regeneration in mice; however, the relationship between miR-26a and its target, directly or indirectly, remains unclear. Therefore, the present study further investigated the mechanism of miR-26a in regulating mouse hepatocyte proliferation. An established mouse liver cell line, Nctc-1469, was transfected with Ad5-miR-26a-EGFP, Ad5-anti-miR-26a-EGFP or Ad5-EGFP vector. Cell proliferation was assessed by MTS, cell apoptosis and cell cycle by flow cytometry, and gene expression by Western blotting and quantitative real-time PCR. Dual-luciferase reporter assays were used to test targets of miR-26a. Compared with the Ad5-EGFP group, Ad5-anti-miR-26a-EGFP down-regulated miR-26a and increased proliferation of hepatocytes, with more cells entering the G1 phase of cell cycle (82.70%+/-1.45% vs 75.80%+/-3.92%), and decreased apoptosis (5.50%+/-0.35% vs 6.73%+/-0.42%). CCND2 and CCNE2 were the direct targeted genes of miR-26a. miR-26a down-regulation up-regulated CCND2 and CCNE2 expressions and down-regulated p53 expression in Nctc-1469 cells. On the contrary, miR-26a over-expression showed the opposite results. miR-26a regulated mouse hepatocyte proliferation by directly targeting the 3' untranslated regions of cyclin D2/cyclin E2; miR-26a also regulated p53-mediated apoptosis. Our data suggested that miR-26a may be a promising regulator in liver regeneration.

Regulation of MT1-MMP and MMP-2 by leptin in cardiac fibroblasts involves Rho/ROCK-dependent actin cytoskeletal reorganization and leads to enhanced cell migration.

PubMed

Schram, Kristin; Ganguly, Riya; No, Eun Kyung; Fang, Xiangping; Thong, Farah S L; Sweeney, Gary

2011-05-01

Altered leptin action has been implicated in the pathophysiology of heart failure in obesity, a hallmark of which is extracellular matrix remodeling. Here, we characterize the direct influence of leptin on matrix metalloproteinase (MMP) activity in primary adult rat cardiac fibroblasts and focus on elucidating the molecular mechanisms responsible. Leptin increased expression and cell surface localization of membrane type 1 (MT1)-MMP, measured by cell surface biotinylation assay and antibody-based colorimetric detection of an exofacial epitope in intact cells. Coimmunoprecipitation analysis showed that leptin also induced the formation of a cluster of differentiation 44/MT1-MMP complex. Qualitative analysis using rhodamine-conjugated phalloidin immunofluorescence indicated that leptin stimulated actin cytoskeletal reorganization and enhanced stress fiber formation. Hence, we analyzed activation of Ras homolog gene family (Rho), member A GTPase activity and found a rapid increase in response to leptin that corresponded with increased phosphorylation of cofilin. Quantitative analysis of cytoskeleton reorganization upon separation of globular and filamentous actin by differential centrifugation confirmed the significant increase in filamentous to globular actin ratio in response to leptin, which was prevented by pharmacological inhibition of Rho (C3 transferase) or its downstream effector kinase Rho-associated coiled-coil-forming protein kinase (ROCK) (Y-27632). Inhibition of Rho or ROCK also attenuated leptin-stimulated increases in cell surface MT1-MMP content. Pro-MMP-2 is a known MT1-MMP substrate, and we observed that enhanced cell surface MT1-MMP in response to leptin resulted in enhanced extracellular activation of pro-MMP-2 measured by gelatin zymography, which was again attenuated by inhibition of Rho or ROCK. Using wound scratch assays, we observed enhanced cell migration, but not proliferation, measured by 5-bromo2'-deoxy-uridine incorporation, in response to leptin, again via a Rho-dependent signaling mechanism. Our results suggest that leptin regulates myocardial matrix remodeling by regulating the cell surface localization of MT1-MMP in adult cardiac fibroblasts via Rho/ROCK-dependent actin cytoskeleton reorganization. Subsequent pro-MMP-2 activation then contributes to stimulation of cell migration.

Aesculin inhibits matrix metalloproteinase-9 expression via p38 mitogen activated protein kinase and activator protein 1 in lipopolysachride-induced RAW264.7 cells.

PubMed

Choi, Hee-Jung; Chung, Tae-Wook; Kim, Jai-Eun; Jeong, Han-Sol; Joo, Myungsoo; Cha, Jaeho; Kim, Cheorl-Ho; Ha, Ki-Tae

2012-11-01

Expression of matrix metalloproteinase 9 (MMP-9) may contribute to inflammatory conditions such as arthritis, hepatitis, atherosclerosis, and pulmonary fibrosis, which involves the destruction of the extracellular matrix (ECM). Macrophages stimulated with lipopolysaccharide (LPS) express MMP-9 through the nuclear factor-kappa B (NF-κB) and activator protein 1 (AP-1) signaling pathways. Aesculin, a 6,7-dihydroxycoumarin-6-O-beta-glucopyranoside, has been highlighted for its anti-hepatotoxic, hypouricemic, antioxidative, photo-protective, and anti-apoptotic properties. In this study, we investigated the effects of aesculin on LPS-stimulated MMP-9 production and its regulatory mechanism by using murine macrophage RAW264.7 cells. Aesculin did not trigger any significant cytotoxic effect on RAW264.7 cells at concentration up to 150 μM. Secretion and expression levels of MMP-9, which were highly elevated by LPS treatment, were reduced by the addition of aesculin in a dose-dependent manner. However, gelatinolytic activity of MMP-9 was not reduced by aesculin. Luciferase activity assays and electrophoretic mobility shift assays using RAW264.7 cells showed that the inhibition of MMP-9 expression by aesculin was mediated by AP-1 rather than NF-κB. In addition, aesculin inhibited phosphorylation of p38 MAPK and subsequent activation of c-fos, a component of AP-1 transcription factor, but not JNK, ERK1/2, and c-jun. These findings suggest that aesculin is a potent drug candidate that protects against the inflammatory destruction of ECM. Copyright © 2012 Elsevier B.V. All rights reserved.

Matrix Metalloproteinases in Non-Neoplastic Disorders

PubMed Central

Tokito, Akinori; Jougasaki, Michihisa

2016-01-01

The matrix metalloproteinases (MMPs) are zinc-dependent endopeptidases belonging to the metzincin superfamily. There are at least 23 members of MMPs ever reported in human, and they and their substrates are widely expressed in many tissues. Recent growing evidence has established that MMP not only can degrade a variety of components of extracellular matrix, but also can cleave and activate various non-matrix proteins, including cytokines, chemokines and growth factors, contributing to both physiological and pathological processes. In normal conditions, MMP expression and activity are tightly regulated via interactions between their activators and inhibitors. Imbalance among these factors, however, results in dysregulated MMP activity, which causes tissue destruction and functional alteration or local inflammation, leading to the development of diverse diseases, such as cardiovascular disease, arthritis, neurodegenerative disease, as well as cancer. This article focuses on the accumulated evidence supporting a wide range of roles of MMPs in various non-neoplastic diseases and provides an outlook on the therapeutic potential of inhibiting MMP action. PMID:27455234

Human telomerase reverse transcriptase is a promising target for cancer inhibition in squamous cell carcinomas.

PubMed

Park, Young-Jin; Kim, Eun-Kyoung; Moon, Sook; Hong, Doo-Pyo; Bae, Jung Yoon; Kim, Jin

2014-11-01

The present study aimed to investigate whether the down-regulation of human telomerase reverse transcriptase (hTERT) may induce an anti-invasive effect in oral squamous cell cancer cell lines. A genetically-engineered squamous carcinoma cell line overexpressing hTERT in immortalized oral keratinocytes transfected by human papilloma virus (HPV)-16 E6/E7 (IHOK) was used. In vivo tumorigenicity was examined using an orthotopic xenograft model of nude mice. For evaluating anti-invasive activity by knockdown of hTERT expression, transwell invasion assay and real-time polymerase chain reaction (PCR) for matrix metalloproteinases (MMP) were employed. The down-regulation of hTERT expression reduced the invasive activity and MMP expression. This result was re-confirmed in the HSC3 oral squamous carcinoma cell line. Targeting hTERT may lead to novel therapeutic approaches. Copyright© 2014 International Institute of Anticancer Research (Dr. John G. Delinassios), All rights reserved.

Fraxinus rhynchophylla ethanol extract attenuates carbon tetrachloride-induced liver fibrosis in rats via down-regulating the expressions of uPA, MMP-2, MMP-9 and TIMP-1.

PubMed

Peng, Wen-Huang; Tien, Yun-Chen; Huang, Chih-Yang; Huang, Tai-Hung; Liao, Jung-Chun; Kuo, Chao-Lin; Lin, Ying-Chih

2010-02-17

To investigate the effect of Fraxinus rhynchophylla ethanol extract (FR(EtOH)) on liver fibrosis induced by carbon tetrachloride (CCl(4)) in rats. Rat hepatic fibrosis was induced by oral administration of CCl(4). Sixty SD rats were divided randomly into 6 groups: control, CCl(4) group, silymarin group and three FR(EtOH)-treated groups. Except for the rats in control group, all rats were administered orally with CCl(4) (20%, 0.2 mL/100g body weight) twice a week for 8 weeks. Rats in FR(EtOH) groups were treated daily with FR(EtOH) (0.1, 0.5 and 1.0 g/kg, p.o.) throughout the whole experimental period. Liver function parameters (such as activities of serum GOT and GPT levels), activities of liver anti-oxidant enzymes (such as catalase, SOD, GPx) and expressions of uPA, tPA, MMP-2, MMP-9 and TIMP-1, -2, -3, -4 in the liver fibrosis pathway were detected. The results showed that FR(EtOH) (0.1, 0.5 and 1.0 g/kg BW) significantly reduced the elevated activities of sGOT and sGPT caused by CCl(4). FR(EtOH) (0.1 and 0.5 g/kg BW) and significantly increased the activities of GSH-Px. The histopathological study showed that FR(EtOH) (0.1 and 0.5 g/kg BW) reduced the incidence of liver lesions, including hepatic cells cloudy swelling, lymphocytes infiltration, cytoplasm vacuolization hepatic necrosis and fibrous connective tissue proliferated induced by CCl(4) in rats. In our study it was showed that CCl(4)-treated group significantly increased the protein levels of uPA, MMP-2, MMP-9 and TIMP-1. FR(EtOH) (0.1 and 0.5 g/kg BW) could inhibit the protein levels of uPA, MMP-2, MMP-9 and TIMP-1. Finally, the amount of esculetin in the FR(EtOH) was 33.54 mg/g extract. Oral administration of FR(EtOH) significantly reduces CCl(4)-induced hepatic fibrosis in rats, probably by exerting a protective effect against hepatocellular fibrosis by its free radical scavenging ability. FR(EtOH) down-regulated the expressions of uPA, MMP-2 and MMP-9 in CCl(4)-induced liver fibrosis in rats. Copyright 2009 Elsevier Ireland Ltd. All rights reserved.

Membrane type 1-matrix metalloproteinase cleaves off the NH2-terminal portion of heparin-binding epidermal growth factor and converts it into a heparin-independent growth factor.

PubMed

Koshikawa, Naohiko; Mizushima, Hiroto; Minegishi, Tomoko; Iwamoto, Ryo; Mekada, Eisuke; Seiki, Motoharu

2010-07-15

Epidermal growth factor (EGF) receptors (ErbB) and EGF family members represent promising targets for cancer therapy. Heparin-binding EGF (HB-EGF) is a member of the EGF family and is an important target for therapy in some types of human cancers. Processing of HB-EGF by proprotein convertases, and successively, by ADAM family proteases, generates a soluble growth factor that requires heparin as a cofactor. Although heparin potentiates HB-EGF activity in vitro, it is not clear how the heparin-binding activity of HB-EGF is regulated. Here, we show that membrane type 1-matrix metalloproteinase (MT1-MMP; MMP14), a potent invasion-promoting protease, markedly enhances HB-EGF-dependent tumor formation in mice. MT1-MMP additionally cleaves HB-EGF and removes the NH(2)-terminal 20 amino acids that are important for binding heparin. Consequently, the processing of HB-EGF by MT1-MMP converts HB-EGF into a heparin-independent growth factor with enhanced mitogenic activity, and thereby, expression of both proteins costimulates tumor cell growth in vitro and in vivo. The ErbB family of receptors expressed in human gastric carcinoma cells play a role in mediating enhanced HB-EGF activity by MT1-MMP during invasive cell growth in collagen. Thus, we shed light on a new mechanism whereby HB-EGF activity is regulated that should be considered when designing HB-EGF-targeted cancer therapy. (c)2010 AACR.

Leptin regulates MMP-2, TIMP-1 and collagen synthesis via p38 MAPK in HL-1 murine cardiomyocytes.

PubMed

Schram, Kristin; De Girolamo, Sabrina; Madani, Siham; Munoz, Diana; Thong, Farah; Sweeney, Gary

2010-12-01

A clear association between obesity and heart failure exists and a significant role for leptin, the product of the obese gene, has been suggested. One aspect of myocardial remodeling which characterizes heart failure is a disruption in the balance of extracellular matrix synthesis and degradation. Here we investigated the effects of leptin on matrix metalloproteinase (MMP) activity, tissue inhibitor of metalloproteinase (TIMP) expression, as well as collagen synthesis in HL-1 cardiac muscle cells. Gelatin zymographic analysis of MMP activity in conditioned media showed that leptin enhanced MMP-2 activity in a dose- and time-dependent manner. Leptin is known to stimulate phosphorylation of p38 MAPK in cardiac cells and utilization of the p38 MAPK inhibitor, SB203580, demonstrated that this kinase also plays a role in regulating several extracellular matrix components, such that inhibition of p38 MAPK signaling prevented the leptin-induced increase in MMP-2 activation. We also observed that leptin enhanced collagen synthesis determined by both proline incorporation and picrosirius red staining of conditioned media. Pro-collagen type-I and pro-collagen type-III expression, measured by real-time PCR and Western blotting were also increased by leptin, effects which were again attenuated by SB203580. In summary, these results demonstrate the potential for leptin to play a role in mediating myocardial ECM remodeling and that the p38 MAPK pathway plays an important role in mediating these effects.

Elongation factor-2 kinase regulates TG2/β1 integrin/Src/uPAR pathway and epithelial–mesenchymal transition mediating pancreatic cancer cells invasion

PubMed Central

Ashour, Ahmed A; Gurbuz, Nilgun; Alpay, Sultan Neslihan; Abdel-Aziz, Abdel-Aziz H; Mansour, Ahmed M; Huo, Longfei; Ozpolat, Bulent

2014-01-01

Pancreatic ductal adenocarcinoma is one of the lethal cancers with extensive local tumour invasion, metastasis, early systemic dissemination and poorest prognosis. Thus, understanding the mechanisms regulating invasion/metastasis and epithelial–mesenchymal transition (EMT), is the key for developing effective therapeutic strategies for pancreatic cancer (PaCa). Eukaryotic elongation factor-2 kinase (eEF-2K) is an atypical kinase that we found to be highly up-regulated in PaCa cells. However, its role in PaCa invasion/progression remains unknown. Here, we investigated the role of eEF-2K in cellular invasion, and we found that down-regulation of eEF-2K, by siRNA or rottlerin, displays impairment of PaCa cells invasion/migration, with significant decreases in the expression of tissue transglutaminase (TG2), the multifunctional enzyme implicated in regulation of cell attachment, motility and survival. These events were associated with reductions in β1 integrin/uPAR/MMP-2 expressions as well as decrease in Src activity. Furthermore, inhibition of eEF-2K/TG2 axis suppresses the EMT, as demonstrated by the modulation of the zinc finger transcription factors, ZEB1/Snail, and the tight junction proteins, claudins. Importantly, while eEF-2K silencing recapitulates the rottlerin-induced inhibition of invasion and correlated events, eEF-2K overexpression, by lentivirus-based expression system, suppresses such rottlerin effects and potentiates PaCa cells invasion/migration capability. Collectively, our results show, for the first time, that eEF-2K is involved in regulation of the invasive phenotype of PaCa cells through promoting a new signalling pathway, which is mediated by TG2/β1 integrin/Src/uPAR/MMP-2, and the induction of EMT biomarkers which enhance cancer cell motility and metastatic potential. Thus, eEF-2K could represent a novel potential therapeutic target in pancreatic cancer. PMID:25215932

TGF-beta1 induces the different expressions of lysyl oxidases and matrix metalloproteinases in anterior cruciate ligament and medial collateral ligament fibroblasts after mechanical injury.

PubMed

Xie, Jing; Wang, Chunli; Huang, Dong-Yue; Zhang, Yanyan; Xu, Jianwen; Kolesnikov, Stanislav S; Sung, K L Paul; Zhao, Hucheng

2013-03-15

The anterior cruciate ligament (ACL) is known to have a poor self-healing ability. In contrast, the medial collateral ligament (MCL) can heal relatively well and restore the joint function. Transforming growth factor-beta1 (TGF-β1) is considered to be an important chemical mediator in the wound healing of the ligaments. While the role of TGF-β1-induced expressions of the lysyl oxidases (LOXs) and matrix metalloproteinases (MMPs), which respectively facilitate the extracellular matrix (ECM) repair and degradation, is poorly understood. In this study, we used equibiaxial stretch chamber to mimic mechanical injury of ACL and MCL fibroblasts, and aimed to determine the intrinsic differences between ACL and MCL by characterizing the differential expressions of LOXs and MMPs in response to TGF-β1 after mechanical injury. By using semi-quantitative PCR, quantitative real-time PCR, western blot and zymography, we found TGF-β1 induced injured MCL to express more LOXs than injured ACL (up to 1.85-fold in LOX, 2.21-fold in LOXL-1, 1.71-fold in LOXL-2, 2.52-fold in LOXL-3 and 3.32-fold in LOXL-4). Meanwhile, TGF-β1 induced injured ACL to express more MMPs than injured MCL fibroblasts (up to 2.33-fold in MMP-1, 2.45-fold in MMP-2, 1.89-fold in MMP-3 and 1.50-fold in MMP-12). The further protein results were coincident with the gene expressions above. The different expressions of LOXs and MMPs inferred the intrinsic differences between ACL and MCL, and the intrinsic differences could help to explain their differential healing abilities. Copyright © 2012 Elsevier Ltd. All rights reserved.

3-Hydroxyflavone inhibits human osteosarcoma U2OS and 143B cells metastasis by affecting EMT and repressing u-PA/MMP-2 via FAK-Src to MEK/ERK and RhoA/MLC2 pathways and reduces 143B tumor growth in vivo.

PubMed

Lu, Ko-Hsiu; Chen, Pei-Ni; Hsieh, Yi-Hsien; Lin, Chin-Yin; Cheng, Fu-Yuan; Chiu, Peng-Chou; Chu, Shu-Chen; Hsieh, Yih-Shou

2016-11-01

Many natural flavonoids have cytostatic and apoptotic properties; however, we little know whether the effect of synthetic 3-hydroxyflavone on metastasis and tumor growth of human osteosarcoma. Here, we tested the hypothesis that 3-hydroxyflavone suppresses human osteosarcoma cells metastasis and tumor growth. 3-hydroxyflavone, up to 50 μM without cytotoxicity, inhibited U2OS and 143B cells motility, invasiveness and migration by reducing matrix metalloproteinase (MMP)-2 and urokinase-type plasminogen activator (u-PA) and also impaired cell adhesion to gelatin. 3-hydroxyflavone significantly reduced p-focal adhesion kinase (FAK) Tyr397, p-FAK Tyr925, p-steroid receptor coactivator (Src), p-mitogen/extracellular signal-regulated kinase (MEK)1/2, p-myosin light chain (MLC)2 Ser19, epithelial cell adhesion molecule, Ras homolog gene family (Rho)A and fibronectin expressions. 3-hydroxyflavone also affected the epithelial-mesenchymal transition (EMT) by down-regulating expressions of Vimentin and α-catenin with activation of the transcription factor Slug. In nude mice xenograft model and tail vein injection model showed that 3-hydroxyflavone reduced 143B tumor growth and lung metastasis. 3-hydroxyflavone possesses the anti-metastatic activity of U2OS and 143B cells by affecting EMT and repressing u-PA/MMP-2 via FAK-Src to MEK/ERK and RhoA/MLC2 pathways and suppresses 143B tumor growth in vivo. This may lead to clinical trials of osteosarcoma chemotherapy to confirm the promising result in the future. Copyright © 2016 Elsevier Ltd. All rights reserved.

Magnolol suppresses the proliferation and invasion of cholangiocarcinoma cells via inhibiting the NF-κB signaling pathway.

PubMed

Zhang, Fu-Hui; Ren, Hong-Yue; Shen, Jin-Xing; Zhang, Xiao-Yun; Ye, Hui-Ming; Shen, Dong-Yan

2017-10-01

Magnolol has shown the potential anticancer properties against a variety of cancers. However, the role of magnolol in cholangiocarcinoma (CCA) cells is unknown. In this study, we assessed the effect of magnolol on the CCA cells. CCA cells were treated with magnolol in the absence or presence of TNFα, the activator for NF-κB. After co-incubation with magnolol, cell proliferation and growth were examined by MTT, colony formation and xenograft tumors; cell cycle was analyzed by flow cytometry; cell migration and invasion were detected by wound healing and transwell assays; the expression of PCNA, Ki67, CyclinD1, MMP-2, MMP-7 and MMP-9 and NF-κB pathway were evaluated by using Western blot. Magnolol inhibited the abilities of CCA cell growth, migration and invasion accompanying with a decreased expression of PCNA, Ki67, MMP-2, MMP-7 and MMP-9 (all P<0.05). with magnolol induced cell cycle arrest in G1 phase with a downregulation of cell cycle protein CyclinD1 (all P<0.05). In addition, magnolol suppressed the expression of p-IκBα and p-P65 and the effect of magnolol on CCA cells could be inhibited by TNFα. Magnolol could inhibit the growth, migration and invasion of CCA cells through regulation of NF-κB pathway, and these data indicate that magnolol is a potential candidate for treating of CCA. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

Cigarette smoke-induced hypercapnic emphysema in C3H mice is associated with increases of macrophage metalloelastase and substance P in the lungs.

PubMed

Xu, J; Xu, F; Wang, R; Seagrave, Jc; Lin, Y; March, T H

2007-01-01

The authors tested whether macrophage metalloelastase (MMP-12) and substance P (SP) were increased in the cigarette smoke (CS)-exposed female C3H/HeN mice with hypercapnic emphysema. The authors found that as compared to control (filtered air), 16 weeks of CS exposure significantly up-regulated mRNA and protein levels of MMP-12, the ratio of MMP-12/tissue inhibitor of matrix metalloproteinase-1, and SP/preprotachykinin-A (a precursor to SP) in the lungs. Importantly, a significant correlation was found between MMP-12 and SP, and between MMP-12/SP and the degrees of hypoxemia/hypercapnia denoted in CS-exposed mice. These data suggest a possible involvement of SP and MMP-12 in the pathogenesis of severe COPD.

Glioma-mediated microglial activation promotes glioma proliferation and migration: roles of Na+/H+ exchanger isoform 1

PubMed Central

Zhu, Wen; Carney, Karen E.; Pigott, Victoria M.; Falgoust, Lindsay M.; Clark, Paul A.; Kuo, John S.; Sun, Dandan

2016-01-01

Microglia play important roles in extracellular matrix remodeling, tumor invasion, angiogenesis, and suppression of adaptive immunity in glioma. Na+/H+ exchanger isoform 1 (NHE1) regulates microglial activation and migration. However, little is known about the roles of NHE1 in intratumoral microglial activation and microglia–glioma interactions. Our study revealed up-regulation of NHE1 protein expression in both glioma cells and tumor-associated Iba1+ microglia in glioma xenografts and glioblastoma multiforme microarrays. Moreover, we observed positive correlation of NHE1 expression with Iba1 intensity in microglia/macrophages. Glioma cells, via conditioned medium or non-contact glioma-microglia co-cultures, concurrently upregulated microglial expression of NHE1 protein and other microglial activation markers (iNOS, arginase-1, TGF-β, IL-6, IL-10 and the matrix metalloproteinases MT1-MMP and MMP9). Interestingly, glioma-stimulated microglia reciprocally enhanced glioma proliferation and migration. Most importantly, inhibition of microglial NHE1 activity via small interfering RNA (siRNA) knockdown or the potent NHE1-specific inhibitor HOE642 significantly attenuated microglial activation and abolished microglia-stimulated glioma migration and proliferation. Taken together, our findings provide the first evidence that NHE1 function plays an important role in glioma–microglia interactions, enhancing glioma proliferation and invasion by stimulating microglial release of soluble factors. NHE1 upregulation is a novel marker of the glioma-associated microglial activation phenotype. Inhibition of NHE1 represents a novel glioma therapeutic strategy by targeting tumor-induced microglial activation. PMID:27287871

Macrophage Metalloelastase (MMP-12) Deficiency Mitigates Retinal Inflammation and Pathological Angiogenesis in Ischemic Retinopathy

PubMed Central

Li, Jingming; Wang, Joshua J.; Peng, Qisheng; Chen, Chen; Humphrey, Mary Beth; Heinecke, Jay; Zhang, Sarah X.

2012-01-01

Pathological angiogenesis is a major cause of vision loss in ischemic and inflammatory retinal diseases. Recent evidence implicates macrophage metalloelastase (MMP-12), a macrophage-derived elastinolytic protease in inflammation, tissue remodeling and angiogenesis. However, little is known about the role of MMP-12 in retinal pathophysiology. The present study aims to explore the enzyme’s contributions to retinal angiogenesis in oxygen-induced retinopathy (OIR) using MMP-12 knockout (KO) mice. We find that MMP-12 expression was upregulated in OIR, accompanied by elevated macrophage infiltration and increased inflammatory markers. Compared to wildtype mice, MMP-12 KO mice had decreased levels of adhesion molecule and inflammatory cytokines and reduced vascular leakage in OIR. Concomitantly, these mice had markedly reduced macrophage content in the retina with impaired macrophage migratory capacity. Significantly, loss of MMP-12 attenuated retinal capillary dropout in early OIR and mitigated pathological retinal neovascularization (NV). Similar results were observed in the study using MMP408, a pharmacological inhibitor of MMP-12. Intriguingly, in contrast to reducing pathological angiogenesis, lack of MMP-12 accelerated revascularization of avascular retina in OIR. Taken together, we conclude that MMP-12 is a key regulator of macrophage infiltration and inflammation, contributing to retinal vascular dysfunction and pathological angiogenesis. PMID:23285156