Up-regulation of proproliferative genes and the ligand/receptor pair placental growth factor and vascular endothelial growth factor receptor 1 in hepatitis C cirrhosis.

PubMed

Huang, Xiao X; McCaughan, Geoffrey W; Shackel, Nicholas A; Gorrell, Mark D

2007-09-01

Cirrhosis can lead to hepatocellular carcinoma (HCC). Non-diseased liver and hepatitis C virus (HCV)-associated cirrhosis with or without HCC were compared. Proliferation pathway genes, immune response genes and oncogenes were analysed by a quantitative real-time reverse transcriptase-polymerase chain reaction (RT-PCR) and immunostaining. Real-time RT-PCR showed up-regulation of genes in HCV cirrhosis including the proliferation-associated genes bone morphogenetic protein 3 (BMP3), placental growth factor 3 (PGF3), vascular endothelial growth factor receptor 1 (VEGFR1) and soluble VEGFR1, the oncogene FYN, and the immune response-associated genes toll-like receptor 9 (TLR9) and natural killer cell transcript 4 (NK4). Expressions of TLR2 and the oncogenes B-cell CLL/lymphoma 9 (BCL9) and PIM2 were decreased in HCV cirrhosis. In addition, PIM2 and TLR2 were increased in HCV cirrhosis with HCC compared with HCV cirrhosis. The ligand/receptor pair PGF and VEGFR1 was intensely expressed by the portal tract vascular endothelium. VEGFR1 was expressed in reactive biliary epithelial structures in fibrotic septum and in some stellate cells and macrophages. PGF and VEGFR1 may have an important role in the pathogenesis of the neovascular response in cirrhosis.

Insulin-like growth factor II messenger RNA-binding protein-3 is an independent prognostic factor in uterine leiomyosarcoma.

PubMed

Yasutake, Nobuko; Ohishi, Yoshihiro; Taguchi, Kenichi; Hiraki, Yuka; Oya, Masafumi; Oshiro, Yumi; Mine, Mari; Iwasaki, Takeshi; Yamamoto, Hidetaka; Kohashi, Kenichi; Sonoda, Kenzo; Kato, Kiyoko; Oda, Yoshinao

2018-04-01

The aim of this study was to identify the prognostic factors of uterine leiomyosarcoma (ULMS). We reviewed 60 cases of surgically resected ULMSs and investigated conventional clinicopathological factors, together with the expression of insulin-like growth factor II messenger RNA-binding protein-3 (IMP3), hormone receptors and cell cycle regulatory markers by immunohistochemistry. Mediator complex subunit 12 (MED12) mutation analysis was also performed. Univariate analyses revealed that advanced stage (P < 0.0001), older age (P = 0.0244) and IMP3 expression (P = 0.0011) were significant predictors of a poor outcome. Multivariate analysis revealed advanced stage (P < 0.0001) and IMP3 (P = 0.0373) as independent predictors of a poor prognosis. Expressions of cell cycle markers and hormone receptors, and MED12 mutations (12% in ULMSs) were not identified as prognostic markers in this study. IMP3 expression in ULMS could be a marker of a poor prognosis. © 2017 John Wiley & Sons Ltd.

Induction of cyclooxygenase-2 expression by prostaglandin E2 stimulation of the prostanoid EP4 receptor via coupling to Gαi and transactivation of the epidermal growth factor receptor in HCA-7 human colon cancer cells.

PubMed

Yoshida, Kenji; Fujino, Hiromichi; Otake, Sho; Seira, Naofumi; Regan, John W; Murayama, Toshihiko

2013-10-15

Increased expressions of cyclooxygenase-2 (COX-2) and its downstream metabolite, prostaglandin E2 (PGE2), are well documented events in the development of colorectal cancer. Interestingly, PGE2 itself can induce the expression of COX-2 thereby creating the potential for positive feedback. Although evidence for such a positive feedback has been previously described, the specific E-type prostanoid (EP) receptor subtype that mediates this response, as well as the relevant signaling pathways, remain unclear. We now report that the PGE2 stimulated induction of COX-2 expression in human colon cancer HCA-7 cells is mediated by activation of the prostanoid EP4 receptor subtype and is followed by coupling of the receptor to Gαi and the activation of phosphatidylinositol 3-kinase. Subsequent activation of metalloproteinases releases membrane bound heparin-binding epidermal growth factor-like growth factor resulting in the transactivation of epidermal growth factor receptors and the activation of the extracellular signal-regulated kinases and induction of COX-2 expression. This induction of COX-2 expression by PGE2 stimulation of the prostanoid EP4 receptor may underlie the upregulation of COX-2 during colorectal cancer and appears to be an early event in the process of tumorigenesis. © 2013 Elsevier B.V. All rights reserved.

Expression of Vascular Endothelial Growth Factor Receptors in Benign Vascular Lesions of the Orbit: A Case Series.

PubMed

Atchison, Elizabeth A; Garrity, James A; Castillo, Francisco; Engman, Steven J; Couch, Steven M; Salomão, Diva R

2016-01-01

Vascular lesions of the orbit, although not malignant, can cause morbidity because of their location near critical structures in the orbit. For the same reason, they can be challenging to remove surgically. Anti-vascular endothelial growth factor (VEGF) drugs are increasingly being used to treat diseases with prominent angiogenesis. Our study aimed to determine to what extent VEGF receptors and their subtypes are expressed on selected vascular lesions of the orbit. Retrospective case series of all orbital vascular lesions removed by one of the authors (JAG) at the Mayo Clinic. A total of 52 patients who underwent removal of vascular orbital lesions. The pathology specimens from the patients were retrieved, their pathologic diagnosis was confirmed, demographic and clinical information were gathered, and sections from vascular tumors were stained with vascular endothelial growth factor receptor (VEGFR), vascular endothelial growth factor receptor type 1 (VEGFR1), vascular endothelial growth factor receptor type 2 (VEGFR2), and vascular endothelial growth factor receptor type 3 (VEGFR3). The existence and pattern of staining with VEGF and its subtypes on these lesions. There were 28 specimens of venous malformations, 4 capillary hemangiomas, 7 lymphatic malformations, and 6 lymphaticovenous malformations. All samples stained with VEGF, 55% stained with VEGFR1, 98% stained with VEGFR2, and 96% stained with VEGFR3. Most (94%) of the VEGFR2 staining was diffuse. Most orbital vascular lesions express VEGF receptors, which may suggest a future target for nonsurgical treatment. Copyright © 2016 American Academy of Ophthalmology. Published by Elsevier Inc. All rights reserved.

Gene expression of growth hormone family and glucocorticoid receptors, osmosensors, and ion transporters in the gill during seawater acclimation of Mozambique tilapia, Oreochromis mossambicus.

PubMed

Breves, Jason P; Fox, Bradley K; Pierce, Andrew L; Hirano, Tetsuya; Grau, E Gordon

2010-08-01

This study characterized endocrine and ionoregulatory responses accompanying seawater (SW) acclimation in Mozambique tilapia (Oreochromis mossambicus). Changes in plasma hormones and gene expression of hormone receptors, putative osmosensors, and ion transporters in the gill were measured. Transfer of freshwater (FW)-acclimated tilapia to SW resulted in a marked elevation in plasma osmolality and a significant rise in plasma growth hormone (GH) levels at 12 hr and 14 days after transfer. Significant reductions in plasma prolactin (PRL(177) and PRL(188)) levels also occurred in SW-transferred fish; no effect of transfer upon plasma cortisol or insulin-like growth factor I was observed. Gene expression of GH receptor increased strongly 6 hr after transfer, whereas PRL receptor was lower than controls at 12 hr. By contrast, mRNA levels of somatolactin and glucocorticoid receptors were unaffected by SW transfer. Osmotic stress transcription factor 1 mRNA levels rose significantly between 3 and 12 hr, whereas the calcium-sensing receptor was unaffected. Aquaporin-3 gene expression was strongly down-regulated during SW acclimation from 12 hr until the conclusion of the experiment. Na(+)/K(+)/2Cl(-) cotransporter gene expression increased significantly 3 hr after transfer, whereas expression of Na(+)/Cl(-) cotransporter, specific to FW-type chloride cells, declined by 6 hr into SW acclimation. The response of Na(+)/H(+) exchanger was less pronounced, but showed a similar pattern to that of the Na(+)/Cl(-) cotransporter. These results suggest that acquisition of hyposmoregulatory mechanisms in Mozambique tilapia entails the coordinated interaction of systemic hormones with local factors in the gill, including hormone receptors, ion transporters, and osmosensors. (c) 2010 Wiley-Liss, Inc.

A tissue microarray study of toll-like receptor 4, decoy receptor 3, and external signal regulated kinase 1/2 expressions in astrocytoma.

PubMed

Lin, Chih-Kung; Ting, Chun-Chieh; Tsai, Wen-Chiuan; Chen, Yuan-Wu; Hueng, Dueng-Yuan

2016-01-01

Decoy receptor 3 (DcR3) functions as a death decoy inhibiting apoptosis mediated by the tumor necrosis factor receptor family. It is highly expressed in many tumors and its expression can be regulated by the MAPK/ERK signaling pathway and ERK is a vital member of this pathway. Toll-like receptor 4 (TLR4) is expressed on immune cells. Increased TLR4 expression has been associated with various types of cancers. The study was conducted to investigate the expression of DcR3, ERK1/2, and TLR4 in astrocytomas and evaluate if they are validating markers for discriminating glioblastoma from anaplastic astrocytoma in limited surgical specimen. Expression of DcR3, ERK1/2, and TLR4 was determined by immunohistochemical staining of tissue microarray from 48 paraffin-embedded tissues. A binary logistic regression method was used to generate functions that discriminate between anaplastic astrocytomas and glioblastomas. The expression of TLR4 and DcR3 was significantly higher in glioblastomas than in anaplastic astrocytomas. DcR3 could discriminate anaplastic astrocytomas from glioblastomas with high sensitivity (93.8%), specificity (90%), and accuracy (92.3%). Our results suggest that DcR3 may be a useful marker for discriminating anaplastic astrocytomas from glioblastomas.

EGFR-driven up-regulation of decoy receptor 3 in keratinocytes contributes to the pathogenesis of psoriasis.

PubMed

Wu, Nan-Lin; Huang, Duen-Yi; Hsieh, Shie-Liang; Hsiao, Cheng-Hsiang; Lee, Te-An; Lin, Wan-Wan

2013-10-01

Decoy receptor 3 (DcR3) is a soluble receptor of Fas ligand (FasL), LIGHT (TNFSF14) and TNF-like molecule 1A (TL1A) and plays pleiotropic roles in many inflammatory and autoimmune disorders and malignant diseases. In cutaneous biology, DcR3 is expressed in primary human epidermal keratinocytes and is upregulated in skin lesions in psoriasis, which is characterized by chronic inflammation and angiogenesis. However, the regulatory mechanisms of DcR3 over-expression in skin lesions of psoriasis are unknown. Here, we demonstrate that DcR3 can be detected in both dermal blood vessels and epidermal layers of psoriatic skin lesions. Analysis of serum samples showed that DcR3 was elevated, but FasL was downregulated in psoriatic patients compared with normal individuals. Additional cell studies revealed a central role of epidermal growth factor receptor (EGFR) in controlling the basal expression of DcR3 in keratinocytes. Activation of EGFR by epidermal growth factor (EGF) and transforming growth factor (TGF)-α strikingly upregulated DcR3 production. TNF-αenhanced DcR3 expression in both keratinocytes and endothelial cells compared with various inflammatory cytokines involved in psoriasis. Additionally, TNF-α-enhanced DcR3 expression in keratinocytes was inhibited when EGFR was knocked down or EGFR inhibitor was used. The NF-κB pathway was critically involved in the molecular mechanisms underlying the action of EGFR and inflammatory cytokines. Collectively, the novel regulatory mechanisms of DcR3 expression in psoriasis, particularly in keratinocytes and endothelial cells, provides new insight into the pathogenesis of psoriasis and may also contribute to the understanding of other diseases that involve DcR3 overexpression. Copyright © 2013 Elsevier B.V. All rights reserved.

NFIL3 suppresses hypoxia-induced apoptotic cell death by targeting the insulin-like growth factor 2 receptor.

PubMed

Lin, Kuan-Ho; Kuo, Chia-Hua; Kuo, Wei-Wen; Ho, Tsung-Jung; Pai, Peiying; Chen, Wei-Kung; Pan, Lung-Fa; Wang, Chien-Cheng; Padma, V Vijaya; Huang, Chih-Yang

2015-06-01

The insulin-like growth factor-II/mannose 6-phosphate receptor (IGF2R) over-expression correlates with heart disease progression. The IGF2R is not only an IGF2 clearance receptor, but it also triggers signal transduction, resulting in cardiac hypertrophy, apoptosis and fibrosis. The present study investigated the nuclear factor IL-3 (NFIL3), a transcription factor of the basic leucine zipper superfamily, and its potential pro-survival effects in cardiomyocytes. NFIL3 might play a key role in heart development and act as a survival factor in the heart, but the regulatory mechanisms are still unclear. IGF2 and IGF2R protein expression were highly increased in rat hearts subjected to hemorrhagic shock. IGF2R protein expression was also up-regulated in H9c2 cells exposed to hypoxia. Over-expression of NFIL3 in H9c2 cardiomyoblast cells inhibited the induction of hypoxia-induced apoptosis and down-regulated IGF2R expression levels. Gel shift assay, double-stranded DNA pull-down assay and chromatin immune-precipitation analyses indicated that NFIL3 binds directly to the IGF2R promoter region. Using a luciferase assay, we further observed NFIL3 repress IGF2R gene promoter activity. Our results demonstrate that NFIL3 is an important negative transcription factor, which through binding to the promoter of IGF2R, suppresses the apoptosis induced by IGF2R signaling in H9c2 cardiomyoblast cells under hypoxic conditions. © 2015 Wiley Periodicals, Inc.

Fibroblast growth factor 8 is expressed at higher levels in lactating human breast and in breast cancer.

PubMed

Zammit, C; Coope, R; Gomm, J J; Shousha, S; Johnston, C L; Coombes, R C

2002-04-08

Fibroblast growth factor 8 can transform NIH3T3 cells and its expression has been found to be associated with breast and prostate cancer. Following our finding that fibroblast growth factor 8 mRNA expression is increased in breast cancer, we have undertaken an immunohistochemistry study of fibroblast growth factor 8 expression in a series of human breast tissues and other normal tissues. Our findings confirm increased expression of fibroblast growth factor 8 in malignant breast tissue but also show significant fibroblast growth factor 8 expression in non-malignant breast epithelial cells. No significant difference in fibroblast growth factor 8 expression was found between different grades of ductal carcinoma, lobular carcinoma and ductal carcinoma in-situ or cancer of different oestrogen receptor, progesterone receptor or nodal status. The highest levels of fibroblast growth factor 8 expression were found in lactating breast tissues and fibroblast growth factor 8 was also detected in human milk. A survey of other normal tissues showed that fibroblast growth factor 8 is expressed in the proliferative cells of the dermis and epithelial cells in colon, ovary fallopian tube and uterus. Fibroblast growth factor 8 appears to be expressed in several organs in man and appears to have an importance in lactation.

Exercise reduces adipose tissue via cannabinoid receptor type 1 which is regulated by peroxisome proliferator-activated receptor-delta.

PubMed

Yan, Zhen Cheng; Liu, Dao Yan; Zhang, Li Li; Shen, Chen Yi; Ma, Qun Li; Cao, Ting Bing; Wang, Li Juan; Nie, Hai; Zidek, Walter; Tepel, Martin; Zhu, Zhi Ming

2007-03-09

Obesity is one major cardiovascular risk factor. We tested effects of endurance exercise on cannabinoid receptor type 1 (CB1) and peroxisome proliferator-activated receptor-delta (PPAR-delta)-dependent pathways in adipose tissue. Male Wistar rats were randomly assigned to standard laboratory chow or a high-fat diet without and with regular endurance exercise. Exercise in rats on high-fat diet significantly reduced visceral fat mass, blood pressure, and adipocyte size (each p<0.05). Adipocyte hypertrophy induced by high-fat diet was accompanied by increased CB1 expression in adipose tissue, whereas exercise significantly reduced CB1 expression (each p<0.05). CB1 receptor expression and adipocyte differentiation were directly regulated by PPAR-delta. Adipocyte hypertrophy induced by high-fat diet was accompanied by reduced PPAR-delta. Furthermore, selective silencing of PPAR-delta by RNA interference in 3T3-L1-preadipocyte cells significantly increased CB1 expression from 1.00+/-0.06 (n=3) to 1.91+/-0.06 (n=3; p<0.01) and increased adipocyte differentiation, whereas adenovirus-mediated overexpression of PPAR-delta significantly reduced CB1 expression to 0.39+/-0.03 (n=3; p<0.01) and reduced adipocyte differentiation. In the presence of the CB1 antagonist rimonabant adipocyte differentiation in stimulated 3T3 L1 preadipocyte cells was significantly reduced. The study indicates that high-fat diet-induced hypertrophy of adipocytes is associated with increased CB1 receptor expression which is directly regulated by PPAR-delta. Both CB1 and PPAR-delta are intimately involved in therapeutic interventions against a most important cardiovascular risk factor.

Activation of Stat1 by mutant fibroblast growth-factor receptor in thanatophoric dysplasia type II dwarfism.

PubMed

Su, W C; Kitagawa, M; Xue, N; Xie, B; Garofalo, S; Cho, J; Deng, C; Horton, W A; Fu, X Y

1997-03-20

The achondroplasia class of chondrodysplasias comprises the most common genetic forms of dwarfism in humans and includes achondroplasia, hypochondroplasia and thanatophoric dysplasia types I and II (TDI and TDII), which are caused by different mutations in a fibroblast growth-factor receptor FGFR3 (ref. 1). The molecular mechanism and the mediators of these FGFR3-related growth abnormalities are not known. Here we show that mutant TDII FGFR3 has a constitutive tyrosine kinase activity which can specifically activate the transcription factor Stat1 (for signal transducer and activator of transcription). Furthermore, expression of TDII FGFR3 induced nuclear translocation of Stat1, expression of the cell-cycle inhibitor p21(WAF1/CIP1), and growth arrest of the cell. Thus, TDII FGFR3 may use Stat1 as a mediator of growth retardation in bone development. Consistent with this, Stat1 activation and increased p21(WAF1/CIP1) expression was found in the cartilage cells from the TDII fetus, but not in those from the normal fetus. Thus, abnormal STAT activation and p21(WAF1/CIP1) expression by the TDII mutant receptor may be responsible for this FGFR3-related bone disease.

Src family kinase expression and subcellular localization in macrophages: implications for their role in CSF-1-induced macrophage migration.

PubMed

Dwyer, Amy R; Mouchemore, Kellie A; Steer, James H; Sunderland, Andrew J; Sampaio, Natalia G; Greenland, Eloise L; Joyce, David A; Pixley, Fiona J

2016-07-01

A major role of colony-stimulating factor-1 is to stimulate the differentiation of mononuclear phagocytic lineage cells into adherent, motile, mature macrophages. The colony-stimulating factor-1 receptor transduces colony-stimulating factor-1 signaling, and we have shown previously that phosphatidylinositol 3-kinase p110δ is a critical mediator of colony-stimulating factor-1-stimulated motility through the colony-stimulating factor-1 receptor pY721 motif. Src family kinases are also implicated in the regulation of macrophage motility and in colony-stimulating factor-1 receptor signaling, although functional redundancy of the multiple SFKs expressed in macrophages makes it challenging to delineate their specific functions. We report a comprehensive analysis of individual Src family kinase expression in macrophage cell lines and primary macrophages and demonstrate colony-stimulating factor-1-induced changes in Src family kinase subcellular localization, which provides clues to their distinct and redundant functions in macrophages. Moreover, expression of individual Src family kinases is both species specific and dependent on colony-stimulating factor-1-induced macrophage differentiation. Hck associated with the activated colony-stimulating factor-1 receptor, whereas Lyn associated with the receptor in a constitutive manner. Consistent with this, inhibitor studies revealed that Src family kinases were important for both colony-stimulating factor-1 receptor activation and colony-stimulating factor-1-induced macrophage spreading, motility, and invasion. Distinct colony-stimulating factor-1-induced changes in the subcellular localization of individual SFKs suggest specific roles for these Src family kinases in the macrophage response to colony-stimulating factor-1. © Society for Leukocyte Biology.

Activation of phosphatidylinositol-3-kinase by platelet-derived growth factor and insulin-like growth factor-1 is inhibited by a transmembrane phosphotyrosine phosphatase.

PubMed

Way, B A; Mooney, R A

1993-12-15

Expression of the transmembrane phosphotyrosine phosphatase (PTPase) CD45 has been shown to inhibit hormone-dependent tyrosine phosphorylation and mitogenesis (Mooney, R. A., Freund, G. G., Way, B. A., and Bordwell, K. L. (1992) J. Biol. Chem. 267, 23443-23446). Here the impact of PTPase expression on insulin-like growth factor-1 (IGF-1) and platelet-derived growth factor- (PDGF) dependent activation of PI-3-K was investigated. In PTPase+ cells, IGF-1 and PDGF-dependent PI-3-K activity in antiphosphotyrosine immunoprecipitates was decreased by 62 +/- 13 and 46 +/- 17%, respectively, compared to control cells. Similar decreases in PI-3-K activity associated with anti-PDGF receptor and anti-insulin receptor substrate-1 (IRS-1) immunoprecipitates were also observed. Association of PI-3-K with the hormone-activated PDGF receptor decreased approximately 55%, paralleling its loss of activation in PTPase+ cells. Tyrosine phosphorylation of the 85-kDa subunit of PI-3-K was also inhibited. Similarly, IGF-1 dependent tyrosine phosphorylation of IRS-1 was decreased by 45%, and its association with PI-3-K was decreased by 65% in PTPase+ cells. Finally, PDGF-dependent tyrosine phosphorylation of phospholipase C-gamma 1 and GTPase-activating protein was reduced by 60-70% in the PTPase+ cells as was tyrosine phosphorylation of the PDGF receptor associated with these proteins. In summary, expression of a transmembrane PTPase decreased hormone-dependent PI-3-K activation, tyrosine phosphorylation of receptor substrates, and their association with signaling complexes. These data support a role for transmembrane PTPases in the regulation of receptor signal transduction pathways.

Interactions of phosphatidylinositol kinase, GTPase-activating protein (GAP), and GAP-associated proteins with the colony-stimulating factor 1 receptor.

PubMed Central

Reedijk, M; Liu, X Q; Pawson, T

1990-01-01

The interactions of the macrophage colony-stimulating factor 1 (CSF-1) receptor with potential targets were investigated after ligand stimulation either of mouse macrophages or of fibroblasts that ectopically express mouse CSF-1 receptors. In Rat-2 cells expressing the mouse CSF-1 receptor, full activation of the receptor and cellular transformation require exogenous CSF-1, whereas NIH 3T3 cells expressing mouse c-fms are transformed by autocrine stimulation. Activated CSF-1 receptors physically associate with a phosphatidylinositol (PI) 3'-kinase. A mutant CSF-1 receptor with a deletion of the kinase insert region was deficient in its ability to bind functional PI 3'-kinase and to induce PI 3'-kinase activity precipitable with antiphosphotyrosine antibodies. In fibroblasts, CSF-1 stimulation also induced the phosphorylation of the GTPase-activating protein (GAP)-associated protein p62 on tyrosine, although GAP itself was a relatively poor substrate. In contrast to PI 3'-kinase association, phosphorylation of p62 and GAP was not markedly affected by deletion of the kinase insert region. These results indicate that the kinase insert region selectively enhances the CSF-1-dependent association of the CSF-1 receptor with active PI 3'-kinase. The insert deletion mutant retains considerable transforming activity in NIH 3T3 cells (G. Taylor, M. Reedijk, V. Rothwell, L. Rohrschneider, and T. Pawson, EMBO J. 8:2029-2037, 1989). This mutant was more seriously impaired in Rat-2 cell transformation, although mutant-expressing Rat-2 cells still formed small colonies in soft agar in the presence of CSF-1. Therefore, phosphorylation of GAP and p62 through activation of the CSF-1 receptor does not result in full fibroblast transformation. The interaction between the CSF-1 receptor and PI 3'-kinase may contribute to c-fms fibroblast transformation and play a role in CSF-1-stimulated macrophages. Images PMID:2172781

The TAM-family receptor Mer mediates production of HGF through the RhoA-dependent pathway in response to apoptotic cells.

PubMed

Park, Hyun-Jung; Baen, Ji-Yeon; Lee, Ye-Ji; Choi, Youn-Hee; Kang, Jihee Lee

2012-08-01

The TAM receptor protein tyrosine kinases Tyro3, Axl, and Mer play important roles in macrophage function. We investigated the roles of the TAM receptors in mediating the induction of hepatocyte growth factor (HGF) during the interaction of macrophages with apoptotic cells. Mer-specific neutralizing antibody, small interfering RNA (siRNA), and a recombinant Mer protein (Mer/Fc) inhibited HGF mRNA and protein expression, as well as activation of RhoA, Akt, and specific mitogen-activated protein (MAP) kinases in response to apoptotic cells. Inhibition of Axl or Tyro3 with specific antibodies, siRNA, or Fc-fusion proteins did not prevent apoptotic cell-induced HGF mRNA and protein expression and did not inhibit activation of the postreceptor signaling molecules RhoA and certain MAP kinases, including extracellular signal-regulated protein kinase and c-Jun NH(2)-terminal kinase. However, Axl- and Tyro3-specific blockers did inhibit the activation of Akt and p38 MAP kinase in response to apoptotic cells. In addition, none of the TAM receptors mediated the effects of apoptotic cells on transforming growth factor-β or epidermal growth factor mRNA expression. However, they were involved in the induction of vascular endothelial growth factor mRNA expression. Our data provide evidence that when macrophages interact with apoptotic cells, only Mer of the TAM-family receptors is responsible for mediating transcriptional HGF production through a RhoA-dependent pathway.

Presence of Functional Neurotrophin TrkB Receptors in the Rat Superior Cervical Ganglion

PubMed Central

Valle-Leija, Pablo; Cancino-Rodezno, Angeles; Sánchez-Tafolla, Berardo M.; Arias, Erwin; Elinos, Diana; Feria, Jessica; Zetina, María E.; Morales, Miguel A.; Cifuentes, Fredy

2017-01-01

Sympathetic neurons express the neurotrophin receptors TrkA, p75NTR, and a non-functional truncated TrkB isoform (TrkB-Tc), but are not thought to express a functional full-length TrkB receptor (TrkB-Fl). We, and others, have demonstrated that nerve growth factor (NGF) and brain derived neurotrophic factor (BDNF) modulate synaptic transmission and synaptic plasticity in neurons of the superior cervical ganglion (SCG) of the rat. To clarify whether TrkB is expressed in sympathetic ganglia and contributes to the effects of BDNF upon sympathetic function, we characterized the presence and activity of the neurotrophin receptors expressed in the adult SCG compared with their presence in neonatal and cultured sympathetic neurons. Here, we expand our previous study regarding the immunodetection of neurotrophin receptors. Immunohistochemical analysis revealed that 19% of adult ganglionic neurons expressed TrkB-Fl immunoreactivity (IR), 82% expressed TrkA-IR, and 51% expressed p75NTR-IR; TrkB-Tc would be expressed in 36% of neurons. In addition, using Western-blotting and reverse transcriptase polymerase chain reaction (RT-PCR) analyses, we confirmed the expression of TrkB-Fl and TrkB-Tc protein and mRNA transcripts in adult SCG. Neonatal neurons expressed significantly more TrkA-IR and TrkB-Fl-IR than p75NTR-IR. Finally, the application of neurotrophin, and high frequency stimulation, induced the activation of Trk receptors and the downstream PI3-kinase (phosphatidyl inositol-3-kinase) signaling pathway, thus evoking the phosphorylation of Trk and Akt. These results demonstrate that SCG neurons express functional TrkA and TrkB-Fl receptors, which may contribute to the differential modulation of synaptic transmission and long-term synaptic plasticity. PMID:28744222

Presence of Functional Neurotrophin TrkB Receptors in the Rat Superior Cervical Ganglion.

PubMed

Valle-Leija, Pablo; Cancino-Rodezno, Angeles; Sánchez-Tafolla, Berardo M; Arias, Erwin; Elinos, Diana; Feria, Jessica; Zetina, María E; Morales, Miguel A; Cifuentes, Fredy

2017-01-01

Sympathetic neurons express the neurotrophin receptors TrkA, p75NTR, and a non-functional truncated TrkB isoform (TrkB-Tc), but are not thought to express a functional full-length TrkB receptor (TrkB-Fl). We, and others, have demonstrated that nerve growth factor (NGF) and brain derived neurotrophic factor (BDNF) modulate synaptic transmission and synaptic plasticity in neurons of the superior cervical ganglion (SCG) of the rat. To clarify whether TrkB is expressed in sympathetic ganglia and contributes to the effects of BDNF upon sympathetic function, we characterized the presence and activity of the neurotrophin receptors expressed in the adult SCG compared with their presence in neonatal and cultured sympathetic neurons. Here, we expand our previous study regarding the immunodetection of neurotrophin receptors. Immunohistochemical analysis revealed that 19% of adult ganglionic neurons expressed TrkB-Fl immunoreactivity (IR), 82% expressed TrkA-IR, and 51% expressed p75NTR-IR; TrkB-Tc would be expressed in 36% of neurons. In addition, using Western-blotting and reverse transcriptase polymerase chain reaction (RT-PCR) analyses, we confirmed the expression of TrkB-Fl and TrkB-Tc protein and mRNA transcripts in adult SCG. Neonatal neurons expressed significantly more TrkA-IR and TrkB-Fl-IR than p75NTR-IR. Finally, the application of neurotrophin, and high frequency stimulation, induced the activation of Trk receptors and the downstream PI3-kinase (phosphatidyl inositol-3-kinase) signaling pathway, thus evoking the phosphorylation of Trk and Akt. These results demonstrate that SCG neurons express functional TrkA and TrkB-Fl receptors, which may contribute to the differential modulation of synaptic transmission and long-term synaptic plasticity.

PPAR-alpha agonist treatment increases trefoil factor family-3 expression and attenuates apoptosis in the liver tissue of bile duct-ligated rats.

PubMed

Karakan, Tarkan; Kerem, Mustafa; Cindoruk, Mehmet; Engin, Doruk; Alper, Murat; Akın, Okan

2013-01-01

Peroxisome proliferators-activated receptor alpha activation modulates cholesterol metabolism and suppresses bile acid synthesis. The trefoil factor family comprises mucin-associated proteins that increase the viscosity of mucins and help protect epithelial linings from insults. We evaluated the effect of short-term administration of fenofibrate, a peroxisome proliferators activated receptor alpha agonist, on trefoil factor family-3 expression, degree of apoptosis, generation of free radicals, and levels of proinflammatory cytokines in the liver tissue of bile duct-ligated rats. Forty male Wistar rats were randomly divided into four groups: 1 = sham operated, 2 = bile duct ligation, 3 = bile duct-ligated + vehicle (gum Arabic), and 4 = bile duct-ligated + fenofibrate (100 mg/kg/day). All rats were sacrificed on the 7 th day after obtaining blood samples and liver tissue. Liver function tests, tumor necrosis factor-alpha and interleukin 1 beta in serum, and trefoil factor family-3 mRNA expression, degree of apoptosis (TUNEL) and tissue malondialdehyde (malondialdehyde, end-product of lipid peroxidation by reactive oxygen species) in liver tissue were evaluated. Fenofibrate administration significantly reduced serum total bilirubin, aspartate aminotransferase, alanine aminotransferase, alkaline phosphatase, gamma-glutamyl transferase, and tumor necrosis factor-alpha and interleukin-1β levels. Apoptosis and malondialdehyde were significantly reduced in the fenofibrate group. Trefoil factor family-3 expression increased with fenofibrate treatment in bile duct-ligated rats. The peroxisome proliferators-activated receptor alpha agonist fenofibrate significantly increased trefoil factor family-3 expression and decreased apoptosis and lipid peroxidation in the liver and attenuated serum levels of proinflammatory cytokines in bile duct-ligated rats. Further studies are needed to determine the protective role of fenofibrate in human cholestatic disorders.

Cloning the promoter for transforming growth factor-beta type III receptor. Basal and conditional expression in fetal rat osteoblasts

NASA Technical Reports Server (NTRS)

Ji, C.; Chen, Y.; McCarthy, T. L.; Centrella, M.

1999-01-01

Transforming growth factor-beta binds to three high affinity cell surface molecules that directly or indirectly regulate its biological effects. The type III receptor (TRIII) is a proteoglycan that lacks significant intracellular signaling or enzymatic motifs but may facilitate transforming growth factor-beta binding to other receptors, stabilize multimeric receptor complexes, or segregate growth factor from activating receptors. Because various agents or events that regulate osteoblast function rapidly modulate TRIII expression, we cloned the 5' region of the rat TRIII gene to assess possible control elements. DNA fragments from this region directed high reporter gene expression in osteoblasts. Sequencing showed no consensus TATA or CCAAT boxes, whereas several nuclear factors binding sequences within the 3' region of the promoter co-mapped with multiple transcription initiation sites, DNase I footprints, gel mobility shift analysis, or loss of activity by deletion or mutation. An upstream enhancer was evident 5' proximal to nucleotide -979, and a silencer region occurred between nucleotides -2014 and -2194. Glucocorticoid sensitivity mapped between nucleotides -687 and -253, whereas bone morphogenetic protein 2 sensitivity co-mapped within the silencer region. Thus, the TRIII promoter contains cooperative basal elements and dispersed growth factor- and hormone-sensitive regulatory regions that can control TRIII expression by osteoblasts.

Fibroblast growth factor receptor signaling is essential for lens fiber cell differentiation.

PubMed

Zhao, Haotian; Yang, Tianyu; Madakashira, Bhavani P; Thiels, Cornelius A; Bechtle, Chad A; Garcia, Claudia M; Zhang, Huiming; Yu, Kai; Ornitz, David M; Beebe, David C; Robinson, Michael L

2008-06-15

The vertebrate lens provides an excellent model to study the mechanisms that regulate terminal differentiation. Although fibroblast growth factors (FGFs) are thought to be important for lens cell differentiation, it is unclear which FGF receptors mediate these processes during different stages of lens development. Deletion of three FGF receptors (Fgfr1-3) early in lens development demonstrated that expression of only a single allele of Fgfr2 or Fgfr3 was sufficient for grossly normal lens development, while mice possessing only a single Fgfr1 allele developed cataracts and microphthalmia. Profound defects were observed in lenses lacking all three Fgfrs. These included lack of fiber cell elongation, abnormal proliferation in prospective lens fiber cells, reduced expression of the cell cycle inhibitors p27(kip1) and p57(kip2), increased apoptosis and aberrant or reduced expression of Prox1, Pax6, c-Maf, E-cadherin and alpha-, beta- and gamma-crystallins. Therefore, while signaling by FGF receptors is essential for lens fiber differentiation, different FGF receptors function redundantly.

Ligand-Independent Epidermal Growth Factor Receptor Overexpression Correlates with Poor Prognosis in Colorectal Cancer.

PubMed

Yun, Sumi; Kwak, Yoonjin; Nam, Soo Kyung; Seo, An Na; Oh, Heung-Kwon; Kim, Duck-Woo; Kang, Sung-Bum; Lee, Hye Seung

2018-01-17

Molecular treatments targeting epidermal growth factor receptors (EGFRs) are important strategies for advanced colorectal cancer (CRC). However, clinicopathologic implications of EGFRs and EGFR ligand signaling have not been fully evaluated. We evaluated the expression of EGFR ligands and correlation with their receptors, clinicopathologic factors, and patients' survival with CRC. The expression of EGFR ligands, including heparin binding epidermal growth factor like growth factor (HBEGF), transforming growth factor (TGF), betacellulin, and epidermal growth factor (EGF), were evaluated in 331 consecutive CRC samples using mRNA in situ hybridization (ISH). We also evaluated the expression status of EGFR, human epidermal growth factor receptor 2 (HER2), HER3, and HER4 using immunohistochemistry and/or silver ISH. Unlike low incidences of TGF (38.1%), betacellulin (7.9%), and EGF (2.1%), HBEGF expression was noted in 62.2% of CRC samples. However, the expression of each EGFR ligand did not reveal significant correlations with survival. The combined analyses of EGFR ligands and EGFR expression indicated that the ligands‒/EGFR+ group showed a significant association with the worst disease-free survival (DFS, p=0.018) and overall survival (OS, p=0.005). It was also an independent, unfavorable prognostic factor for DFS (p=0.026) and OS (p=0.007). Additionally, HER4 nuclear expression, regardless of ligand expression, was an independent, favorable prognostic factor for DFS (p=0.034) and OS (p=0.049), by multivariate analysis. Ligand-independent EGFR overexpression was suggested to have a significant prognostic impact; thus, the expression status of EGFR ligands, in addition to EGFR, might be necessary for predicting patients' outcome in CRC.

Differential expression and role of hyperglycemia induced oxidative stress in epigenetic regulation of β1, β2 and β3-adrenergic receptors in retinal endothelial cells

PubMed Central

2014-01-01

Background Aberrant epigenetic profiles are concomitant with a spectrum of developmental defects and diseases. Role of methylation is an increasingly accepted factor in the pathophysiology of diabetes and its associated complications. This study aims to examine the correlation between oxidative stress and methylation of β1, β2 and β3-adrenergic receptors and to analyze the differential variability in the expression of these genes under hyperglycemic conditions. Methods Human retinal endothelial cells were cultured in CSC complete medium in normal (5 mM) or high (25 mM) glucose to mimic a diabetic condition. Reverse transcription PCR and Western Blotting were performed to examine the expression of β1, β2 and β3-adrenergic receptors. For detections, immunocytochemistry was used. Bisulfite sequencing method was used for promoter methylation analysis. Apoptosis was determined by the terminal deoxynucleotidyl transferase dUTP nick-end labeling (TUNEL) assay. Dichlorodihydrofluorescein diacetate (DCFH-DA) assay was used to measure reactive oxygen species (ROS) production in the cells. Results β1 and β3-adrenergic receptors were expressed in retinal endothelial cells while β2-adrenergic receptor was not detectable both at protein and mRNA levels. Hyperglycemia had no significant effect on β1 and β2-adrenergic receptors methylation and expression however β3-adrenergic receptors showed a significantly higher expression (p < 0.05) and methylation (p < 0.01) in high and low glucose concentration respectively. Apoptosis and oxidative stress were inversely correlated with β3-adrenergic receptors methylation with no significant effect on β1 and β2-adrenergic receptors. β2-adrenergic receptor was hypermethylated with halted expression. Conclusion Our study demonstrates that β1 and β3-adrenergic receptors expressed in human retinal endothelial cells. Oxidative stress and apoptosis are inversely proportional to the extent of promoter methylation, suggesting that methylation loss might be due to oxidative stress-induced DNA damage. PMID:24885710

Captodiamine, a putative antidepressant, enhances hypothalamic BDNF expression in vivo by synergistic 5-HT2c receptor antagonism and sigma-1 receptor agonism.

PubMed

Ring, Rebecca M; Regan, Ciaran M

2013-10-01

The putative antidepressant captodiamine is a 5-HT2c receptor antagonist and agonist at sigma-1 and D3 dopamine receptors, exerts an anti-immobility action in the forced swim paradigm, and enhances dopamine turnover in the frontal cortex. Captodiamine has also been found to ameliorate stress-induced anhedonia, reduce the associated elevations of hypothalamic corticotrophin-releasing factor (CRF) and restore the reductions in hypothalamic BDNF expression. Here we demonstrate chronic administration of captodiamine to have no significant effect on hypothalamic CRF expression through sigma-1 receptor agonism; however, both sigma-1 receptor agonism or 5-HT2c receptor antagonism were necessary to enhance BDNF expression. Regulation of BDNF expression by captodiamine was associated with increased phosphorylation of transcription factor CREB and mediated through sigma-1 receptor agonism but blocked by 5-HT2c receptor antagonism. The existence of two separate signalling pathways was confirmed by immunolocalisation of each receptor to distinct cell populations in the paraventricular nucleus of the hypothalamus. Increased BDNF induced by captodiamine was also associated with enhanced expression of synapsin, but not PSD-95, suggesting induction of long-term structural plasticity between hypothalamic synapses. These unique features of captodiamine may contribute to its ability to ameliorate stress-induced anhedonia as the hypothalamus plays a prominent role in regulating HPA axis activity.

Impact of Parturition on Chemokine Homing Factor Expression in the Vaginal Distention Model of Stress Urinary Incontinence

PubMed Central

Lenis, Andrew T.; Kuang, Mei; Woo, Lynn L.; Hijaz, Adonis; Penn, Marc S.; Butler, Robert S.; Rackley, Raymond; Damaser, Margot S.; Wood, Hadley M.

2015-01-01

Purpose Human childbirth simulated by vaginal distention is known to increase the expression of chemokines and receptors involved in stem cell homing and tissue repair. We hypothesized that pregnancy and parturition in rats contributes to the expression of chemokines and receptors after vaginal distention. Materials and Methods We used 72 age matched female Lewis rats, including virgin rats with and without vaginal distention, and delivered rats with and without vaginal distention. Each rat was sacrificed immediately, or 3 or 7 days after vaginal distention and/or parturition, and the urethra was harvested. Relative expression of chemokines and receptors was determined by real-time polymerase chain reaction. Mixed models were used with the Bonferroni correction for multiple comparisons. Results Vaginal distention up-regulated urethral expression of CCL7 immediately after injury in virgin and postpartum rats. Hypoxia inducible factor-1α and vascular endothelial growth factor were up-regulated only in virgin rats immediately after vaginal distention. CD191 expression was immediately up-regulated in postpartum rats without vaginal distention compared to virgin rats without vaginal distention. CD195 was up-regulated in virgin rats 3 days after vaginal distention compared to virgin rats without vaginal distention. CD193 and CXCR4 showed delayed up-regulation in virgin rats 7 days after vaginal distention. CXCL12 was up-regulated in virgin rats 3 days after vaginal distention compared to immediately after vaginal distention. Interleukin-8 and CD192 showed no differential expression. Conclusions Vaginal distention results in up-regulation of the chemokines and receptors expressed during tissue injury, which may facilitate the spontaneous functional recovery previously noted. Pregnancy and delivery up-regulated CD191 and attenuated the expression of hypoxia inducible factor-1α and vascular endothelial growth factor in the setting of vaginal distention, likely by decreasing hypoxia. PMID:23022009

Sequential expression of cyclooxygenase-2, glutamate receptor-2, and platelet activating factor receptor in rat hippocampal neurons after fluid percussion injury

PubMed Central

Li, Zhiqiang; Shu, Qingming; Li, Lingzhi; Ge, Maolin; Zhang, Yongliang

2014-01-01

Traumatic brain injury causes gene expression changes in different brain regions. Occurrence and development of traumatic brain injury are closely related, involving expression of three factors, namely cyclooxygenase-2, glutamate receptor-2, and platelet activating factor receptor. However, little is known about the correlation of these three factors and brain neuronal injury. In this study, primary cultured rat hippocampal neurons were subjected to fluid percussion injury according to Scott's method, with some modifications. RT-PCR and semi-quantitative immunocytochemical staining was used to measure the expression levels of cyclooxygenase-2, glutamate receptor-2, and platelet activating factor receptor. Our results found that cyclooxygenase-2 expression were firstly increased post-injury, and then decreased. Both mRNA and protein expression levels reached peaks at 8 and 12 hours post-injury, respectively. Similar sequential changes in glutamate receptor 2 were observed, with highest levels mRNA and protein expression at 8 and 12 hours post-injury respectively. On the contrary, the expressions of platelet activating factor receptor were firstly decreased post-injury, and then increased. Both mRNA and protein expression levels reached the lowest levels at 8 and 12 hours post-injury, respectively. Totally, our findings suggest that these three factors are involved in occurrence and development of hippocampal neuronal injury. PMID:25206921

Aberrant expression of decoy receptor 3 in human breast cancer: relevance to lymphangiogenesis.

PubMed

Wu, Qiuwan; Zheng, Yahong; Chen, Donghan; Li, Xiaohong; Lu, Chuanhui; Zhang, Zhiming

2014-05-15

Decoy receptor 3 (DcR3), a decoy receptor against Fas ligand belonging to the tumor necrosis factor receptor superfamily, is overexpressed in some forms of cancer. It was recently reported that DcR3 could protect endothelial cells from apoptosis, implying a potential role in the development of vessels, whereas its role in the lymphangiogenesis remains unclear. In the present study, we studied the DcR3 expression and its relationship with the lymphatic microvessel density (LMVD) to investigate if it played a role in the lymph metastasis of human breast cancer. Real-time polymerase chain reaction and immunohistochemistry were performed to measure the messenger RNA and protein expression of DcR3 in the breast cancer tissues, noncancerous counterparts, and axillary lymph node from 63 patients. LMVD in these specimens was assessed by counting the D2-40 labeled-microvessels. Furthermore, the correlations between DcR3 expression and LMVD and other clinicopathologic parameters were analyzed. DcR3 was overexpressed in the breast cancer tissue of 58 patients (92.1%) and was also expressed in vascular endothelial cells and tumor cells in the lymph nodes. LMVD in cancer tissue and lymph nodes were both positively correlated to the aberrant expression of DcR3. The relevance between DcR3 overexpression and LMVD revealed the existence of possible links between DcR3 and lymphangiogenesis. Based on these findings, it is important to further explore the regulation of lymphangiogenesis operated by the reverse tumor necrosis factor signaling of DcR3. Copyright © 2014 The Authors. Published by Elsevier Inc. All rights reserved.

Repeated social defeat and the rewarding effects of cocaine in adult and adolescent mice: dopamine transcription factors, proBDNF signaling pathways, and the TrkB receptor in the mesolimbic system.

PubMed

Montagud-Romero, Sandra; Nuñez, Cristina; Blanco-Gandia, M Carmen; Martínez-Laorden, Elena; Aguilar, María A; Navarro-Zaragoza, Javier; Almela, Pilar; Milanés, Maria-Victoria; Laorden, María-Luisa; Miñarro, José; Rodríguez-Arias, Marta

2017-07-01

Repeated social defeat (RSD) increases the rewarding effects of cocaine in adolescent and adult rodents. The aim of the present study was to compare the long-term effects of RSD on the conditioned rewarding effects of cocaine and levels of the transcription factors Pitx3 and Nurr1 in the ventral tegmental area (VTA), the dopamine transporter (DAT), the D2 dopamine receptor (D2DR) and precursor of brain-derived neurotrophic factor (proBDNF) signaling pathways, and the tropomyosin-related kinase B (TrkB) receptor in the nucleus accumbens (NAc) in adult and adolescent mice. Male adolescent and young adult OF1 mice were exposed to four episodes of social defeat and were conditioned 3 weeks later with 1 mg/kg of cocaine. In a second set of mice, the expressions of the abovementioned dopaminergic and proBDNF and TrkB receptor were measured in VTA and NAc, respectively. Adolescent mice experienced social defeats less intensely than their adult counterparts and produced lower levels of corticosterone. However, both adult and adolescent defeated mice developed conditioned place preference for the compartment associated with this low dose of cocaine. Furthermore, only adolescent defeated mice displayed diminished levels of the transcription factors Pitx3 in the VTA, without changes in the expression of DAT and D2DR in the NAc. In addition, stressed adult mice showed a decreased expression of proBDNF and the TrkB receptor, while stressed adolescent mice exhibited increased expression of latter without changes in the former. Our findings suggest that dopaminergic pathways and proBDNF signaling and TrkB receptors play different roles in social defeat-stressed mice exposed to cocaine.

Activation‐Induced Killer Cell Immunoglobulin‐like Receptor 3DL2 Binding to HLA–B27 Licenses Pathogenic T Cell Differentiation in Spondyloarthritis

PubMed Central

Ridley, Anna; Hatano, Hiroko; Wong‐Baeza, Isabel; Shaw, Jacqueline; Matthews, Katherine K.; Al‐Mossawi, Hussein; Ladell, Kristin; Price, David A.; Bowness, Paul

2016-01-01

Objective In the spondyloarthritides (SpA), increased numbers of CD4+ T cells express killer cell immunoglobulin‐like receptor 3DL2 (KIR‐3DL2). The aim of this study was to determine the factors that induce KIR‐3DL2 expression, and to characterize the relationship between HLA–B27 and the phenotype and function of KIR‐3DL2–expressing CD4+ T cells in SpA. Methods In total, 34 B27+ patients with SpA, 28 age‐ and sex‐matched healthy controls (20 B27− and 8 B27+), and 9 patients with rheumatoid arthritis were studied. KIR-3DL2 expression and other phenotypic characteristics of peripheral blood and synovial fluid CD4+ T cells were studied by flow cytometry, quantitative polymerase chain reaction, and Western blotting. T cell receptor clonality was determined by template‐switch anchored reverse transcription–polymerase chain reaction and sequencing analysis. Cytokines were measured by enzyme‐linked immunosorbent assay. Results Cellular activation induced KIR‐3DL2 expression on both naive and effector CD4+ T cells. KIR‐3DL2 binding to B27+ cells promoted expression of KIR‐3DL2, the Th17‐specific transcription factor retinoic acid receptor–related orphan nuclear receptor γt, and the antiapoptotic factor B cell lymphoma 2. KIR‐3DL2+CD4+ T cells in patients with ankylosing spondylitis were oligoclonal and enriched for markers of T cell activation and for the gut homing receptor CCR9. In the presence of B27+ antigen‐presenting cells, KIR‐3DL2+CD4+ T cells produced less interleukin‐2 (IL‐2) but more IL‐17. This effect was blocked by HC10, an antibody that inhibits the binding of KIR‐3DL2 to B27 heavy chains. Conclusion KIR‐3DL2 binding to HLA–B27 licenses Th17 cell differentiation in SpA. These findings raise the therapeutic potential of targeting HLA–B27–KIR‐3DL2 interactions for the treatment of B27+ patients with SpA. PMID:26841353

Association of the gene expression variation of tumor necrosis factor-α and expressions changes of dopamine receptor genes in progression of diabetic severe foot ulcers

PubMed Central

Vaseghi, Hajar; Pornour, Majid; Djavid, Gholamreza Esmaeeli; Rigi, Garshasb; Ganji, Shahla Mohammad; Novin, Leila

2017-01-01

Objective(s): Regulation of pro-inflammatory factors such as TNF-α which are secreted by the immune cells through induction of their several receptors including dopamine receptors (especially DRD2 and DRD3) is one of the noticeable problems in diabetic severe foot ulcer healing. This study was conducted to evaluate the alteration of TNF-αin plasma as well as DRD2 and DRD3 changes in PBMCs of diabetics with severe foot ulcers. Materials and Methods: Peripheral blood samples were collected from 31 subjects with ulcers, 29 without ulcers, and 25 healthy individuals. Total mRNA was extracted from PBMCs for the study of DRD2, DRD3, and TNF-α gene expression variations. Expression patterns of these genes were evaluated by real-time PCR. Consequently, concentration of TNF-α was investigated in plasma. Results: Significant decrease in gene expression and plasma concentration of TNF-α in PBMCs was observed in both patient groups at P<0.05. These diminutions are correlated to the decrease in the expression of both DRD2 and DRD3 in PBMCs of both patient groups. Also, the same relationship is present between expressions of two new DRD3 transcripts with TNF-α downturn. Conclusion: We concluded that DRD2 and DRD3 expression alteration and presence of new DRD3 transcripts can be effective in reduction of TNF-α expression as a pro-inflammatory factor. Performing complementary studies, may explain that variations in DRD2 and DRD3 are prognostic and effective markers attributed to the development of diabetes severe foot ulcers. PMID:29299198

Neurotrophins, their receptors and KI-67 in human GH-secreting pituitary adenomas: an immunohistochemical analysis.

PubMed

Artico, M; Bianchi, E; Magliulo, G; De Vincentiis, M; De Santis, E; Orlandi, A; Santoro, A; Pastore, F S; Giangaspero, F; Caruso, R; Re, M; Fumagalli, L

2012-01-01

Pituitary adenomas are a diverse group of tumors arising from the pituitary gland. Typically, they are small, slow-growing, hormonally inactive lesions that come to light as incidental findings on radiologic or postmortem examinations, although some small, slow-growing lesions with excessive hormonal activity may manifest with a clinical syndrome. The family of neurotrophins plays a key role in the development and maintenance of the pituitary endocrine cell function and in the regulation of hypothalamo-pituitary-adrenocortical axis activity. The objective of our experimental study is to investigate the localization of the neurotrophins, their relative receptors and to detect the expression level of Ki-67 to determine whether all these factors participate in the transformation and development of human pituitary adenomas. A very strong expression of Neurotrophin-3 (NT-3) and its receptor TrKC was observed in the extracellular matrix (ECM) and vessel endothelium, together with a clear/marked presence of Brain-derived neurotrophic factor (BDNF), and its receptor TrKB, thus confirming their direct involvement in the progression of pituitary adenomas. On the contrary, NGF (Nerve growth factor) and its receptor TrKA and p75NTR were weakly expressed in the epithelial gland cells and the ECM.

Activation-Induced Killer Cell Immunoglobulin-like Receptor 3DL2 Binding to HLA-B27 Licenses Pathogenic T Cell Differentiation in Spondyloarthritis.

PubMed

Ridley, Anna; Hatano, Hiroko; Wong-Baeza, Isabel; Shaw, Jacqueline; Matthews, Katherine K; Al-Mossawi, Hussein; Ladell, Kristin; Price, David A; Bowness, Paul; Kollnberger, Simon

2016-04-01

In the spondyloarthritides (SpA), increased numbers of CD4+ T cells express killer cell immunoglobulin-like receptor 3DL2 (KIR-3DL2). The aim of this study was to determine the factors that induce KIR-3DL2 expression, and to characterize the relationship between HLA-B27 and the phenotype and function of KIR-3DL2-expressing CD4+ T cells in SpA. In total, 34 B27+ patients with SpA, 28 age- and sex-matched healthy controls (20 B27- and 8 B27+), and 9 patients with rheumatoid arthritis were studied. KIR-3DL2 expression and other phenotypic characteristics of peripheral blood and synovial fluid CD4+ T cells were studied by flow cytometry, quantitative polymerase chain reaction, and Western blotting. T cell receptor clonality was determined by template-switch anchored reverse transcription-polymerase chain reaction and sequencing analysis. Cytokines were measured by enzyme-linked immunosorbent assay. Cellular activation induced KIR-3DL2 expression on both naive and effector CD4+ T cells. KIR-3DL2 binding to B27+ cells promoted expression of KIR-3DL2, the Th17-specific transcription factor retinoic acid receptor-related orphan nuclear receptor γt, and the antiapoptotic factor B cell lymphoma 2. KIR-3DL2+CD4+ T cells in patients with ankylosing spondylitis were oligoclonal and enriched for markers of T cell activation and for the gut homing receptor CCR9. In the presence of B27+ antigen-presenting cells, KIR-3DL2+CD4+ T cells produced less interleukin-2 (IL-2) but more IL-17. This effect was blocked by HC10, an antibody that inhibits the binding of KIR-3DL2 to B27 heavy chains. KIR-3DL2 binding to HLA-B27 licenses Th17 cell differentiation in SpA. These findings raise the therapeutic potential of targeting HLA-B27-KIR-3DL2 interactions for the treatment of B27+ patients with SpA. © 2016 The Authors. Arthritis & Rheumatology published by Wiley Periodicals, Inc. on behalf of the American College of Rheumatology.

β3-Adrenergic receptors, adipokines and neuroendocrine activation during stress induced by repeated immune challenge in male and female rats.

PubMed

Csanova, Agnesa; Hlavacova, Natasa; Hasiec, Malgorzata; Pokusa, Michal; Prokopova, Barbora; Jezova, Daniela

2017-05-01

The main hypothesis of the study is that stress associated with repeated immune challenge has an impact on β 3 -adrenergic receptor gene expression in the brain. Sprague-Dawley rats were intraperitoneally injected with increasing doses of lipopolysaccharide (LPS) for five consecutive days. LPS treatment was associated with body weight loss and increased anxiety-like behavior. In LPS-treated animals of both sexes, β 3 -receptor gene expression was increased in the prefrontal cortex but not the hippocampus. LPS treatment decreased β 3 -receptor gene expression in white adipose tissue with higher values in males compared to females. In the adipose tissue, LPS reduced peroxisome proliferator-activated receptor-gamma, leptin and adiponectin gene expression, but increased interleukin-6 expression, irrespective of sex. Repeated immune challenge resulted in increased concentrations of plasma aldosterone and corticosterone with higher values of corticosterone in females compared to males. Concentrations of dehydroepiandrosterone (DHEA) in plasma were unaffected by LPS, while DHEA levels in the frontal cortex were lower in the LPS-treated animals compared to the controls. Thus, changes of DHEA levels in the brain take place irrespective of the changes of this neurosteroid in plasma. We have provided the first evidence on stress-induced increase in β 3 -adrenergic receptor gene expression in the brain. Greater reduction of β 3 -adrenergic receptor expression in the adipose tissue and of the body weight gain by repeated immune challenge in male than in female rats suggests sex differences in the role of β 3 -adrenergic receptors in the metabolic functions. LPS-induced changes in adipose tissue regulatory factors and hormone concentrations might be important for coping with chronic infections.

Type I Interferon Receptor Expression in Human Pancreatic and Periampullary Cancer Tissue.

PubMed

Booy, Stephanie; Hofland, Leo J; Waaijers, A Marlijn; Croze, Ed; van Koetsveld, Peter M; de Vogel, Lisette; Biermann, Katharina; van Eijck, Casper H J

2015-01-01

Interferons (IFNs) have several anticancer mechanisms. A number of clinical trials have been conducted regarding adjuvant IFN-α therapy in pancreatic cancer. Type I IFNs exert their effect via the type I IFN receptor (IFNAR-1, IFNAR-2c). The aims of the present study were to determine the type I IFN receptor expression in pancreatic and periampullary cancer tissues and to study its relation with clinicopathological factors. Receptor expression was determined by immunohistochemistry in paraffin-embedded cancer tissue of 47 pancreatic and 54 periampullary cancer patients. The results demonstrated that 91.5% of the pancreatic tumors and 88.9% of the periampullary tumors showed expression of IFNAR-1, of which 23.4% and 13.0% were strongly positive, respectively. Regarding IFNAR-2c expression, 68.1% of the pancreatic tumors and 68.5% of the periampullary tumors were positive, of which 4.3% of the pancreatic tumors and none of the periampullary tumors had a strong expression. No statistically significant associations were found between type I IFN receptor expression and clinicopathological factors or survival. Type I IFN receptors are expressed in pancreatic and periampullary cancer tissues although with great intertumoral and intratumoral variability. A small proportion of both tumors showed a strong expression of the IFNAR-1; only a very small percentage of the pancreatic tumors showed strong expression of the IFNAR-2c.

Decoy receptor 3: a pleiotropic immunomodulator and biomarker for inflammatory diseases, autoimmune diseases and cancer.

PubMed

Lin, Wan-Wan; Hsieh, Shie-Liang

2011-04-01

Recently, several decoy molecules belonging to tumor necrosis factor receptor superfamily (TNFRSF) have been identified, including decoy receptor 1 (DcR1), decoy receptor 2 (DcR2), and decoy receptor 3 (DcR3). One of the tumor necrosis factor superfamily (TNFSF) members, TNF-related apoptosis-inducing ligand (TRAIL), binds to DcR1 and DcR2, which are membranous receptors with a truncated cytoplasmic domain, thus unable to transduce TRAIL-mediated signaling. In contrast to DcR1 and DcR2, DcR3 is a soluble receptor capable of neutralizing the biological effects of three other TNFSF members: Fas ligand (FasL/TNFSF6/CD95L), LIGHT (TNFSF14) and TNF-like molecule 1A (TL1A/TNFSF15). Since FasL is a potent apoptosis- and inflammation-inducing factor, LIGHT is involved in apoptosis and inflammation, and TL1A is a T cell costimulator and is involved in gut inflammation, DcR3 can be defined as an immunomodulator on the basis of its neutralizing effects on FasL, LIGHT, and TL1A. Initial studies demonstrated that DcR3 expression is elevated in tumors cells; however, later work showed that DcR3 expression is also upregulated in inflammatory diseases, where serum DcR3 levels correlate with disease progression. In addition to its neutralizing effect, DcR3 also acts as an effector molecule to modulate cell function via 'non-decoy' activities. This review focuses on the immunomodulatory effects of DcR3 via 'decoy' and 'non-decoy' functions, and discusses the potential of DcR3 as a biomarker to predict cancer invasion and inflammation progression. We also discuss the possible utility of recombinant DcR3 as a therapeutic agent to control autoimmune diseases, as well as the potential to attenuate tumor progression by inhibiting DcR3 expression. Copyright © 2011 Elsevier Inc. All rights reserved.

Glucocorticoid and cytokine crosstalk: Feedback, feedforward, and co-regulatory interactions determine repression or resistance

PubMed Central

Shah, Suharsh; Altonsy, Mohammed O.; Gerber, Antony N.

2017-01-01

Inflammatory signals induce feedback and feedforward systems that provide temporal control. Although glucocorticoids can repress inflammatory gene expression, glucocorticoid receptor recruitment increases expression of negative feedback and feedforward regulators, including the phosphatase, DUSP1, the ubiquitin-modifying enzyme, TNFAIP3, or the mRNA-destabilizing protein, ZFP36. Moreover, glucocorticoid receptor cooperativity with factors, including nuclear factor-κB (NF-κB), may enhance regulator expression to promote repression. Conversely, MAPKs, which are inhibited by glucocorticoids, provide feedforward control to limit expression of the transcription factor IRF1, and the chemokine, CXCL10. We propose that modulation of feedback and feedforward control can determine repression or resistance of inflammatory gene expression toglucocorticoid. PMID:28283576

The DNA replication licensing factor miniature chromosome maintenance 7 is essential for RNA splicing of epidermal growth factor receptor, c-Met, and platelet-derived growth factor receptor.

PubMed

Chen, Zhang-Hui; Yu, Yan P; Michalopoulos, George; Nelson, Joel; Luo, Jian-Hua

2015-01-16

Miniature chromosome maintenance 7 (MCM7) is an essential component of DNA replication licensing complex. Recent studies indicate that MCM7 is amplified and overexpressed in a variety of human malignancies. In this report, we show that MCM7 binds SF3B3. The binding motif is located in the N terminus (amino acids 221-248) of MCM7. Knockdown of MCM7 or SF3B3 significantly increased unspliced RNA of epidermal growth factor receptor, platelet-derived growth factor receptor, and c-Met. A dramatic drop of reporter gene expression of the oxytocin exon 1-intron-exon 2-EGFP construct was also identified in SF3B3 and MCM7 knockdown PC3 and DU145 cells. The MCM7 or SF3B3 depleted cell extract failed to splice reporter RNA in in vitro RNA splicing analyses. Knockdown of SF3B3 and MCM7 leads to an increase of cell death of both PC3 and DU145 cells. Such cell death induction is partially rescued by expressing spliced c-Met. To our knowledge, this is the first report suggesting that MCM7 is a critical RNA splicing factor, thus giving significant new insight into the oncogenic activity of this protein. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

Dependence of Relative Expression of NTR1 and EGFR on Cell Density and Extracellular pH in Human Pancreatic Cancer Cell Lines

PubMed Central

Olszewski-Hamilton, Ulrike; Hamilton, Gerhard

2011-01-01

Pancreatic adenocarcinoma is a devastating disease characterized by early dissemination and poor prognosis. These solid tumors express receptors for neuropeptides like neurotensin (NT) or epidermal growth factor (EGF) and exhibit acidic regions when grown beyond a certain size. We previously demonstrated increases in intracellular Ca2+ levels, intracellular pH and interleukin-8 (IL-8) secretion in BxPC-3 and PANC-1 pancreatic cancer cells in response to a stable NT analog. The present study aimed at investigation of the dependence of the relative expression of NT receptor 1 (NTR1) and EGFR in BxPC-3 and MIA PaCa-2 cells on cell density and extracellular pH (pHe). MTT assays revealed the NTR1 inhibitor SR 142948-sensitive Lys8-ψ-Lys9NT (8–13)-induced proliferation in BxPC-3 and PANC-1 cells. Confluent cultures of BxPC3 and HT-29 lines exhibited highest expression of NTR1 and lowest of EGFR and expression of NTR1 was maximal at slightly acidic pHe. IL-8 production was stimulated by Lys8-ψ-Lys9NT (8–13) and even enhanced at both acidic and alkaline pHe in BxPC-3 and PANC-1 cells. In conclusion, our in vitro study suggests that one contributing factor to the minor responses obtained with EGFR-directed therapy may be downregulation of this receptor in tumor cell aggregates, possibly resulting in acquisition of a more aggressive phenotype via other growth factor receptors like NTR1. PMID:24212612

CD86 and beta2-adrenergic receptor signaling pathways, respectively, increase Oct-2 and OCA-B Expression and binding to the 3'-IgH enhancer in B cells.

PubMed

Podojil, Joseph R; Kin, Nicholas W; Sanders, Virginia M

2004-05-28

Stimulation of CD86 (formerly known as B7-2) and/or the beta2-adrenergic receptor on a CD40 ligand/interleukin-4-activated B cell increased the rate of mature IgG1 transcription. To identify the mechanism responsible for this effect, we determined whether CD86 and/or beta2-adrenergic receptor stimulation regulated transcription factor expression and binding to the 3'-IgH enhancer in vitro and in vivo. We showed that CD86 stimulation increased the nuclear localization of NF-kappaB1 (p50) and phosphorylated RelA (p65) and increased Oct-2 expression and binding to the 3'-IgH enhancer, in a protein kinase C-dependent manner. These effects were lost when CD86-deficient or NF-kappaB1-deficient B cells were used. CD86 stimulation also increased the level of IkappaB-alpha phosphorylation but in a protein kinase C-independent manner. Beta2-adrenergic receptor stimulation increased CREB phosphorylation, OCA-B expression, and OCA-B binding to the 3'-IgH enhancer in a protein kinase A-dependent manner, an effect lost when beta2-adrenergic receptor-deficient B cells were used. Also, the beta2-adrenergic receptor-induced increase in the level of mature IgG1 transcript was lost when OCA-B-deficient B cells were used. These data are the first to show that CD86 stimulation up-regulates the expression of the transcription factor Oct-2 in a protein kinase C- and NF-kappaB1-dependent manner, and that beta2-adrenergic receptor stimulation up-regulates the expression of the coactivator OCA-B in a protein kinase A-dependent manner to cooperate with Oct-2 binding to the 3'-IgH enhancer.

The soluble Decoy Receptor 3 is regulated by a PI3K-dependent mechanism and promotes migration and invasion in renal cell carcinoma.

PubMed

Weissinger, Daniel; Tagscherer, Katrin E; Macher-Göppinger, Stephan; Haferkamp, Axel; Wagener, Nina; Roth, Wilfried

2013-10-10

Overexpression of Decoy Receptor 3 (DcR3), a soluble member of the tumor necrosis factor receptor superfamily, is a common event in several types of cancer. In renal cell carcinoma (RCC), DcR3 overexpression is associated with lymph node and distant metastasis as well as a poor prognosis. However, the functional role and regulation of DcR3 expression in RCC is so far unknown. Modulation of DcR3 expression by siRNA and ectopic gene expression, respectively, was performed in ACHN and 769-P RCC cell lines. Functional effects of a modulated DcR3 expression were analyzed with regard to migration, invasion, adhesion, clonogenicity, and proliferation. Furthermore, quantitative RT-PCR and immunoblot analyses were performed to evaluate the expression of downstream mediators of DcR3. In further experiments, luciferase assays, quantitative RT-PCR and immunoblot analyses were applied to study the regulation of DcR3 expression in RCC. Additionally, an ex vivo tissue slice culture technique combined with immunohistochemistry was used to study the regulation of DcR3 expression in human RCC specimens. Here, we show that DcR3 promotes adhesion, migration and invasiveness of RCC cells. The DcR3-dependent increase in cellular invasiveness is accompanied with an up-regulation of integrin alpha 4, matrixmetalloproteinase 7 and urokinase plasminogen activator (uPA). Further, we identified a signaling pathway regulating DcR3 expression in RCC. Using in vitro experiments as well as an ex vivo RCC tissue slice culture model, we demonstrate that expression of DcR3 is regulated in a PI3K/AKT-dependent manner involving the transcription factor nuclear factor of activated T-cells (NFAT). Taken together, our results identify DcR3 as a key driver of tumor cell dissemination and suggest DcR3 as a promising target for rational therapy of RCC.

The Extracellular Domain of p75NTR Is Necessary to Inhibit Neurotrophin-3 Signaling through TrkA*

PubMed Central

Mischel, Paul S.; Smith, Shane G.; Vining, Ella R.; Valletta, Janice S.; Mobley, William C.; Reichardt, Louis F.

2009-01-01

The TrkA receptor is activated primarily by nerve growth factor (NGF), but it can also be activated by high concentrations of neurotrophin 3 (NT-3). The pan-neurotrophin receptor p75NTR strongly inhibits activation of TrkA by NT-3 but not by NGF. To examine the role of p75NTR in regulating the specificity of TrkA signaling, we expressed both receptors in Xenopus oocytes. Application of NGF or NT-3 to oocytes expressing TrkA alone resulted in efflux of 45Ca2+ by a phospholipase C-γ-dependent pathway. Coexpression of p75NTR with TrkA inhibited 45Ca2+ efflux in response to NT-3 but not NGF. The inhibitory effect on NT-3 activation of TrkA increased with increasing expression of p75NTR. Coexpression of a truncated p75NTR receptor lacking all but the first 9 amino acids of the cytoplasmic domain inhibited NT-3 stimulation of 45Ca2+ efflux, whereas coexpression of an epidermal growth factor receptor/p75NTR chimera (extracellular domain of epidermal growth factor receptor with transmembrane and cytoplasmic domains of p75NTR) did not inhibit NT-3 signaling through TrkA. These studies demonstrated that the extracellular domain of p75NTR was necessary to inhibit NT-3 signaling through TrkA. Remarkably, p75NTR binding to NT-3 was not required to prevent signaling through TrkA, since occupying p75NTR with brain-derived neurotrophic factor or anti-p75 antibody (REX) did not rescue the ability of NT-3 to activate 45Ca2+ efflux. These data suggested a physical association between TrkA and p75NTR. Documenting this physical interaction, we showed that p75NTR and TrkA could be coimmunoprecipitated from Xenopus oocytes. Our results suggest that the interaction of these two receptors on the cell surface mediated the inhibition of NT-3-activated signaling through TrkA. PMID:11150291

Nigella sativa Relieves the Altered Insulin Receptor Signaling in Streptozotocin-Induced Diabetic Rats Fed with a High-Fat Diet.

PubMed

Balbaa, Mahmoud; El-Zeftawy, Marwa; Ghareeb, Doaa; Taha, Nabil; Mandour, Abdel Wahab

2016-01-01

The black cumin (Nigella sativa) "NS" or the black seeds have many pharmacological activities such as antioxidant, anticarcinogenic, antihypertensive, and antidiabetic properties. In this work, streptozotocin-induced diabetic rats fed with a high-fat diet were treated daily with NS oil (NSO) in order to study the effect on the blood glucose, lipid profile, oxidative stress parameters, and the gene expression of some insulin receptor-induced signaling molecules. This treatment was combined also with some drugs (metformin and glimepiride) and the insulin receptor inhibitor I-OMe-AG538. The administration of NSO significantly induced the gene expression of insulin receptor compared to rats that did not receive NSO. Also, it upregulated the expression of insulin-like growth factor-1 and phosphoinositide-3 kinase, whereas the expression of ADAM-17 was downregulated. The expression of ADAM-17 is corroborated by the analysis of TIMP-3 content. In addition, the NSO significantly reduced blood glucose level, components of the lipid profile, oxidative stress parameters, serum insulin/insulin receptor ratio, and the tumor necrosis factor-α, confirming that NSO has an antidiabetic activity. Thus, the daily NSO treatment in our rat model indicates that NSO has a potential in the management of diabetes as well as improvement of insulin-induced signaling.

Neuropilin 1 Receptor Is Up-Regulated in Dysplastic Epithelium and Oral Squamous Cell Carcinoma.

PubMed

Shahrabi-Farahani, Shokoufeh; Gallottini, Marina; Martins, Fabiana; Li, Erik; Mudge, Dayna R; Nakayama, Hironao; Hida, Kyoko; Panigrahy, Dipak; D'Amore, Patricia A; Bielenberg, Diane R

2016-04-01

Neuropilins are receptors for disparate ligands, including proangiogenic factors such as vascular endothelial growth factor and inhibitory class 3 semaphorin (SEMA3) family members. Differentiated cells in skin epithelium and cutaneous squamous cell carcinoma highly express the neuropilin-1 (NRP1) receptor. We examined the expression of NRP1 in human and mouse oral mucosa. NRP1 was significantly up-regulated in oral epithelial dysplasia and oral squamous cell carcinoma (OSCC). NRP1 receptor localized to the outer suprabasal epithelial layers in normal tongue, an expression pattern similar to the normal skin epidermis. However, dysplastic tongue epithelium and OSCC up-regulated NRP1 in basal and proliferating epithelial layers, a profile unseen in cutaneous squamous cell carcinoma. NRP1 up-regulation is observed in a mouse carcinogen-induced OSCC model and in human tongue OSCC biopsies. Human OSCC cell lines express NRP1 protein in vitro and in mouse tongue xenografts. Sites of capillary infiltration into orthotopic OSCC tumors correlate with high NRP1 expression. HSC3 xenografts, which express the highest NRP1 levels of the cell lines examined, showed massive intratumoral lymphangiogenesis. SEMA3A inhibited OSCC cell migration, suggesting that the NRP1 receptor was bioactive in OSCC. In conclusion, NRP1 is regulated in the oral epithelium and is selectively up-regulated during epithelial dysplasia. NRP1 may function as a reservoir to sequester proangiogenic ligands within the neoplastic compartment, thereby recruiting neovessels toward tumor cells. Copyright © 2016 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.

Pro-apoptotic signaling induced by Retinoic acid and dsRNA is under the control of Interferon Regulatory Factor-3 in breast cancer cells.

PubMed

Bernardo, Ana R; Cosgaya, José M; Aranda, Ana; Jiménez-Lara, Ana M

2017-07-01

Breast cancer is one of the most lethal malignancies for women. Retinoic acid (RA) and double-stranded RNA (dsRNA) are considered signaling molecules with potential anticancer activity. RA, co-administered with the dsRNA mimic polyinosinic-polycytidylic acid (poly(I:C)), synergizes to induce a TRAIL (Tumor-Necrosis-Factor Related Apoptosis-Inducing Ligand)- dependent apoptotic program in breast cancer cells. Here, we report that RA/poly(I:C) co-treatment, synergically, induce the activation of Interferon Regulatory Factor-3 (IRF3) in breast cancer cells. IRF3 activation is mediated by a member of the pathogen recognition receptors, Toll-like receptor-3 (TLR3), since its depletion abrogates IRF3 activation by RA/poly(I:C) co-treatment. Besides induction of TRAIL, apoptosis induced by RA/poly(I:C) correlates with the increased expression of pro-apoptotic TRAIL receptors, TRAIL-R1/2, and the inhibition of the antagonistic receptors TRAIL-R3/4. IRF3 plays an important role in RA/poly(I:C)-induced apoptosis since IRF3 depletion suppresses caspase-8 and caspase-3 activation, TRAIL expression upregulation and apoptosis. Interestingly, RA/poly(I:C) combination synergizes to induce a bioactive autocrine/paracrine loop of type-I Interferons (IFNs) which is ultimately responsible for TRAIL and TRAIL-R1/2 expression upregulation, while inhibition of TRAIL-R3/4 expression is type-I IFN-independent. Our results highlight the importance of IRF3 and type-I IFNs signaling for the pro-apoptotic effects induced by RA and synthetic dsRNA in breast cancer cells.

Liver X receptor alpha regulates fatty acid synthase expression in chicken.

PubMed

Demeure, O; Duby, C; Desert, C; Assaf, S; Hazard, D; Guillou, H; Lagarrigue, S

2009-12-01

Liver X receptor alpha (LXRalpha), also referred to as nuclear receptor subfamily 1, group H, member 3 is a member of the nuclear hormone receptor superfamily, and has recently been shown to act as a master transcription factor governing hepatic lipogenesis in mammals. Liver X receptor alpha directly regulates both the expression of other lipogenic transcription factors and the expression of lipogenic enzymes, thereby enhancing hepatic fatty acid synthesis (FASN). In birds, like in humans, fatty acid synthesis primarily occurs in the liver. Whether LXRalpha is involved in hepatic regulation of lipogenic genes remained to be investigated in this species. Here we show that fatty acid synthase and the expression of other lipogenic genes (sterol regulatory element binding protein 1 and steroyl coenzyme A desaturase 1) are induced in chicken hepatoma cells in response to a pharmacological liver X receptor agonist, T0901317. A detailed analysis of the chicken FASN promoter revealed a functional liver X response element. These data define the chicken FASN gene as a direct target of LXRalpha and further expand the role of LXRalpha as a regulator of lipid metabolism in this species.

BAG3 promotes tumour cell proliferation by regulating EGFR signal transduction pathways in triple negative breast cancer.

PubMed

Shields, Sarah; Conroy, Emer; O'Grady, Tony; McGoldrick, Alo; Connor, Kate; Ward, Mark P; Useckaite, Zivile; Dempsey, Eugene; Reilly, Rebecca; Fan, Yue; Chubb, Anthony; Matallanas, David Gomez; Kay, Elaine W; O'Connor, Darran; McCann, Amanda; Gallagher, William M; Coppinger, Judith A

2018-03-20

Triple-negative breast cancer (TNBC), is a heterogeneous disease characterised by absence of expression of the estrogen receptor (ER), progesterone receptor (PR) and lack of amplification of human epidermal growth factor receptor 2 (HER2). TNBC patients can exhibit poor prognosis and high recurrence stages despite early response to chemotherapy treatment. In this study, we identified a pro-survival signalling protein BCL2- associated athanogene 3 (BAG3) to be highly expressed in a subset of TNBC cell lines and tumour tissues. High mRNA expression of BAG3 in TNBC patient cohorts significantly associated with a lower recurrence free survival. The epidermal growth factor receptor (EGFR) is amplified in TNBC and EGFR signalling dynamics impinge on cancer cell survival and disease recurrence. We found a correlation between BAG3 and EGFR expression in TNBC cell lines and determined that BAG3 can regulate tumour cell proliferation, migration and invasion in EGFR expressing TNBC cells lines. We identified an interaction between BAG3 and components of the EGFR signalling networks using mass spectrometry. Furthermore, BAG3 contributed to regulation of proliferation in TNBC cell lines by reducing the activation of components of the PI3K/AKT and FAK/Src signalling subnetworks. Finally, we found that combined targeting of BAG3 and EGFR was more effective than inhibition of EGFR with Cetuximab alone in TNBC cell lines. This study demonstrates a role for BAG3 in regulation of distinct EGFR modules and highlights the potential of BAG3 as a therapeutic target in TNBC.

BAG3 promotes tumour cell proliferation by regulating EGFR signal transduction pathways in triple negative breast cancer

PubMed Central

Shields, Sarah; Conroy, Emer; O’Grady, Tony; McGoldrick, Alo; Connor, Kate; Ward, Mark P.; Useckaite, Zivile; Dempsey, Eugene; Reilly, Rebecca; Fan, Yue; Chubb, Anthony; Matallanas, David Gomez; Kay, Elaine W.; O’Connor, Darran; McCann, Amanda; Gallagher, William M.; Coppinger, Judith A.

2018-01-01

Triple-negative breast cancer (TNBC), is a heterogeneous disease characterised by absence of expression of the estrogen receptor (ER), progesterone receptor (PR) and lack of amplification of human epidermal growth factor receptor 2 (HER2). TNBC patients can exhibit poor prognosis and high recurrence stages despite early response to chemotherapy treatment. In this study, we identified a pro-survival signalling protein BCL2- associated athanogene 3 (BAG3) to be highly expressed in a subset of TNBC cell lines and tumour tissues. High mRNA expression of BAG3 in TNBC patient cohorts significantly associated with a lower recurrence free survival. The epidermal growth factor receptor (EGFR) is amplified in TNBC and EGFR signalling dynamics impinge on cancer cell survival and disease recurrence. We found a correlation between BAG3 and EGFR expression in TNBC cell lines and determined that BAG3 can regulate tumour cell proliferation, migration and invasion in EGFR expressing TNBC cells lines. We identified an interaction between BAG3 and components of the EGFR signalling networks using mass spectrometry. Furthermore, BAG3 contributed to regulation of proliferation in TNBC cell lines by reducing the activation of components of the PI3K/AKT and FAK/Src signalling subnetworks. Finally, we found that combined targeting of BAG3 and EGFR was more effective than inhibition of EGFR with Cetuximab alone in TNBC cell lines. This study demonstrates a role for BAG3 in regulation of distinct EGFR modules and highlights the potential of BAG3 as a therapeutic target in TNBC. PMID:29644001

Expression and associations of TRAF1, BMI-1, ALDH1, and Lin28B in oral squamous cell carcinoma.

PubMed

Wu, Tian-Fu; Li, Yi-Cun; Ma, Si-Rui; Bing-Liu; Zhang, Wen-Feng; Sun, Zhi-Jun

2017-04-01

Tumor necrosis factor receptor-associated factor 1, an adaptor protein of tumor necrosis factor 2, is involved in classical nuclear factor (NF)-κB activation and lymphocyte recruitment. However, less is known about the expression and association of tumor necrosis factor receptor-associated factor 1 with cancer stem cell markers in oral squamous cell carcinoma. This study aimed to investigate the expression of tumor necrosis factor receptor-associated factor 1 and stem cell characteristic markers (lin28 homolog B, B cell-specific Moloney murine leukemia virus integration site 1, and aldehyde dehydrogenase 1) in oral squamous cell carcinoma and analyze their relations. Paraffin-embedded tissues of 78 oral squamous cell carcinomas, 39 normal oral mucosa, and 12 oral dysplasia tissues were employed in tissue microarrays, and the expression of tumor necrosis factor receptor-associated factor 1, B cell-specific Moloney murine leukemia virus integration site 1, aldehyde dehydrogenase 1, and lin28 homolog B was measured by immunohistostaining and digital pathological analysis. The expression of tumor necrosis factor receptor-associated factor 1 was higher in the oral squamous cell carcinoma group as compared with the expression in the oral mucosa (p < 0.01) and oral dysplasia (p < 0.001) groups. In addition, the expression of tumor necrosis factor receptor-associated factor 1 was associated with those of B cell-specific Moloney murine leukemia virus integration site 1, aldehyde dehydrogenase 1, and lin28 homolog B (p = 0.032, r 2  = 0.109; p < 0.0001, r 2  = 0.64; and p < 0.001, r 2  = 0.16) in oral squamous cell carcinoma. The patient survival rate was lower in the highly expressed tumor necrosis factor receptor-associated factor 1 group, although the difference was not significant. The clustering analysis showed that tumor necrosis factor receptor-associated factor 1 was most related to aldehyde dehydrogenase 1. These findings suggest that tumor necrosis factor receptor-associated factor 1 has potential direct/indirect regulations with the cancer stem cell markers in oral squamous cell carcinoma, which may help in further analysis of the cancer stem cell characteristics.

Oxidized Low-Density Lipoprotein Suppresses Expression of Prostaglandin E Receptor Subtype EP3 in Human THP-1 Macrophages

PubMed Central

Sui, Xuxia; Liu, Yanmin; Li, Qi; Liu, Gefei; Song, Xuhong; Su, Zhongjing; Chang, Xiaolan; Zhou, Yingbi; Liang, Bin; Huang, Dongyang

2014-01-01

EP3, one of four prostaglandin E2 (PGE2) receptors, is significantly lower in atherosclerotic plaques than in normal arteries and is localized predominantly in macrophages of the plaque shoulder region. However, mechanisms behind this EP3 expression pattern are still unknown. We investigated the underlying mechanism of EP3 expression in phorbol 12-myristate 13-acetate (PMA)-differentiated THP-1 macrophages with oxidized low-density lipoprotein (oxLDL) treatment. We found that oxLDL decreased EP3 expression, in a dose-dependent manner, at both the mRNA and protein levels. Moreover, oxLDL inhibited nuclear factor-κB (NF-κB)-dependent transcription of the EP3 gene by the activation of peroxisome proliferator-activated receptor-γ (PPAR-γ). Finally, chromatin immunoprecipitation revealed decreased binding of NF-κB to the EP3 promoter with oxLDL and PPAR-γ agonist treatment. Our results show that oxLDL suppresses EP3 expression by activation of PPAR-γ and subsequent inhibition of NF-κB in macrophages. These results suggest that down-regulation of EP3 expression by oxLDL is associated with impairment of EP3-mediated anti-inflammatory effects, and that EP3 receptor activity may exert a beneficial effect on atherosclerosis. PMID:25333975

Expression and functional studies of the GDNF family receptor-alpha3 (GFRα3) in the pancreas

PubMed Central

Nivlet, Laure; Herrmann, Joel; Martin, Delia Esteban; Meunier, Aline; Orvain, Christophe; Gradwohl, Gérard

2018-01-01

The generation of therapeutic β-cells from human pluripotent stem cells relies on the identification of growth factors that faithfully mimic pancreatic β-cell development in vitro. In this context, the aim of the study was to determine the expression and function of the Glial cell line derived neurotrophic factor receptor α 3 (GFRα3) and its ligand Artemin in islet cell development and function. GFRα3 and Artn expression were characterized by in situ hybridization, immunochemistry and qRT-PCR. We used GFRα3-deficient mice to study GFRα3 function and generated a transgenic mice overexpressing Artn in the embryonic pancreas to study Artn function. We found that GFRα3 is expressed at the surface of a subset of Ngn3-positive endocrine progenitors as well as of embryonic α- and β-cells, while Artn is found in the pancreatic mesenchyme. Adult β-cells lack GFRα3 but α-cells express the receptor. GFRα3 was also found in parasympathetic and sympathetic intra islets neurons as well as in glial cells in the embryonic and adult pancreas. The loss of GFRα3 or overexpression of Artn has no impact on Ngn3- and islet- cells formation and maintenance in the embryo. Islet organisation and innervation as well as glucose homeostasis is normal in GFRα3-deficient mice suggesting functional redundancy. PMID:26576643

The N-terminal region of the dopamine D2 receptor, a rhodopsin-like GPCR, regulates correct integration into the plasma membrane and endocytic routes

PubMed Central

Cho, DI; Min, C; Jung, KS; Cheong, SY; Zheng, M; Cheong, SJ; Oak, MH; Cheong, JH; Lee, BK; Kim, KM

2012-01-01

BACKGROUND AND PURPOSE Functional roles of the N-terminal region of rhodopsin-like GPCR family remain unclear. Using dopamine D2 and D3 receptors as a model system, we probed the roles of the N-terminal region in the signalling, intracellular trafficking of receptor proteins, and explored the critical factors that determine the functionality of the N-terminal region. EXPERIMENTAL APPROACH The N-terminal region of the D2 receptor was gradually shortened or switched with that of the D3 receptor or a non-specific sequence (FLAG), or potential N-terminal glycosylation sites were mutated. Effects of these manipulations on surface expression, internalization, post-endocytic behaviours and signalling were determined. KEY RESULTS Shortening the N-terminal region of the D2 receptor enhanced receptor internalization and impaired surface expression and signalling; ligand binding, desensitization and down-regulation were not affected but their association with a particular microdomain, caveolae, was disrupted. Replacement of critical residues within the N-terminal region with the FLAG epitope failed to restore surface expression but partially restored the altered internalization and signalling. When the N-terminal regions were switched between D2 and D3 receptors, cell surface expression pattern of each receptor was switched. Mutations of potential N-terminal glycosylation sites inhibited surface expression but enhanced internalization of D2 receptors. CONCLUSIONS AND IMPLICATIONS Shortening of N-terminus or mutation of glycosylation sites located within the N-terminus enhanced receptor internalization but impaired the surface expression of D2 receptors. The N-terminal region of the D2 receptor, in a sequence-specific manner, controls the receptor's conformation and integration into the plasma membrane, which determine its subcellular localization, intracellular trafficking and signalling properties. PMID:22117524

TERATOGENICITY OF 2,3,7,8-TETRACHLORODIBENZO-P-DIOXIN (TCDD) IN MICE LACKING THE EXPRESSION OF EGF AND/OR TGFALPHA

EPA Science Inventory

Abstract
2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) exposure produces hydronephrosis and cleft palate in mice. These responses are correlated with disruption of expression of epidermal growth factor (EGF) receptor ligands, primarily EGF and transforming growth factor-alpha ...

Axonal outgrowth, neuropeptides expression and receptors tyrosine kinase phosphorylation in 3D organotypic cultures of adult dorsal root ganglia

PubMed Central

Alves, Cecília J.; Leitão, Luís; Sousa, Daniela M.; Alencastre, Inês S.; Conceição, Francisco; Lamghari, Meriem

2017-01-01

Limited knowledge from mechanistic studies on adult sensory neuronal activity was generated, to some extent, in recapitulated adult in vivo 3D microenvironment. To fill this gap there is a real need to better characterize the adult dorsal root ganglia (aDRG) organotypic cultures to make these in vitro systems exploitable for different approaches, ranging from basic neurobiology to regenerative therapies, to address the sensory nervous system in adult stage. We conducted a direct head-to-head comparison of aDRG and embryonic DRG (eDRG) organotypic culture focusing on axonal growth, neuropeptides expression and receptors tyrosine kinase (RTK) activation associated with neuronal survival, proliferation and differentiation. To identify alterations related to culture conditions, these parameters were also addressed in retrieved aDRG and eDRG and compared with organotypic cultures. Under similar neurotrophic stimulation, aDRG organotypic cultures displayed lower axonal outgrowth rate supported by reduced expression of growth associated protein-43 and high levels of RhoA and glycogen synthase kinase 3 beta mRNA transcripts. In addition, differential alteration in sensory neuropeptides expression, namely calcitonin gene-related peptide and substance P, was detected and was mainly pronounced at gene expression levels. Among 39 different RTK, five receptors from three RTK families were emphasized: tropomyosin receptor kinase A (TrkA), epidermal growth factor receptors (EGFR, ErbB2 and ErbB3) and platelet-derived growth factor receptor (PDGFR). Of note, except for EGFR, the phosphorylation of these receptors was dependent on DRG developmental stage and/or culture condition. In addition, EGFR and PDGFR displayed alterations in their cellular expression pattern in cultured DRG. Overall we provided valuable information particularly important when addressing in vitro the molecular mechanisms associated with development, maturation and regeneration of the sensory nervous system. PMID:28742111

Regulatory role of tumor necrosis factor receptor-associated factor 6 in breast cancer by activating the protein kinase B/glycogen synthase kinase 3β signaling pathway.

PubMed

Shen, Hongyu; Li, Liangpeng; Yang, Sujin; Wang, Dandan; Zhou, Siying; Chen, Xiu; Tang, Jinhai

2017-08-01

Tumor necrosis factor receptor-associated factor 6 (TRAF6) is an endogenous adaptor of innate and adaptive immune responses, and serves a crucial role in tumor necrosis factor receptor and toll‑like/interleukin‑1 receptor signaling. Although studies have demonstrated that TRAF6 has oncogenic activity, its potential contributions to breast cancer in human remains largely uninvestigated. The present study examined the expression levels and function of TRAF6 in breast carcinoma (n=32) and adjacent healthy (n=25) tissue samples. Compared with adjacent healthy tissues, TRAF6 protein expression levels were significantly upregulated in breast cancer tissues. Reverse transcription‑quantitative polymerase chain reaction analysis revealed a significant upregulation of the cellular proliferative marker Ki‑67 and proliferation cell nuclear antigen expression levels in breast carcinoma specimens. Furthermore, protein expression levels of the accessory molecule, transforming growth factor β‑activated kinase 1 (TAK1), were significantly increased in breast cancer patients, as detected by western blot analysis. As determined by MTT assay, TRAF6 exerted profoundly proliferative effects in the MCF‑7 breast cancer cell line; however, these detrimental effects were ameliorated by TAK1 inhibition. Notably, protein kinase B (AKT)/glycogen synthase kinase (GSK)3β phosphorylation levels were markedly upregulated in breast cancer samples, compared with adjacent healthy tissues. In conclusion, an altered TRAF6‑TAK1 axis and its corresponding downstream AKT/GSK3β signaling molecules may contribute to breast cancer progression. Therefore, TRAF6 may represent a potential therapeutic target for the treatment of breast cancer.

Age-related changes in expression of transforming growth factor-beta and receptors in cells of intervertebral discs.

PubMed

Matsunaga, Shunji; Nagano, Satoshi; Onishi, Toshiyuki; Morimoto, Norio; Suzuki, Shusaku; Komiya, Setsuro

2003-01-01

The authors conducted a study to determine age-related changes in expression of transforming growth factor (TGF)-beta1, -beta2, -beta3, and Type I and Type II receptors in various cells in the nucleus pulposus and anulus fibrosus. Immunolocalization of TGFbetas and Type I and II receptors was examined during the aging process of cervical intervertebral discs in senescence-accelerated mice (SAM). The TGFbeta family has important roles for cellular function of various tissues. Its role in disc aging, however, is unknown. Detailed information on the temporal and spatial localization of TGFbetas and their receptors in discs is required before discussing introduction of them clinically into the intervertebral disc. Three groups of five SAM each were used. The groups of SAM were age 8, 24, and 50 weeks, respectively. Hematoxylin and eosin staining and immunohistochemical study involving specific antibodies for TGFbeta1, -beta2, -beta3, and Types I and II TGF receptors were performed. Intervertebral discs exhibited degenerative change with advancing age. The TGFbetas and their receptors were present in the fibrocartilaginous cells within the anulus fibrosus and notochord-like cells within the nucleus pulposus of young mice. Expression of TGFbetas and Type I and Type II receptors changed markedly in the cells within the anulus fibrosus during the aging process. The TGFbetas and their receptors were present in cells within the nucleus pulposus and the anulus fibrosus of young mice, and their expression decreased with age.

Glucocorticoid and cytokine crosstalk: Feedback, feedforward, and co-regulatory interactions determine repression or resistance.

PubMed

Newton, Robert; Shah, Suharsh; Altonsy, Mohammed O; Gerber, Antony N

2017-04-28

Inflammatory signals induce feedback and feedforward systems that provide temporal control. Although glucocorticoids can repress inflammatory gene expression, glucocorticoid receptor recruitment increases expression of negative feedback and feedforward regulators, including the phosphatase, DUSP1, the ubiquitin-modifying enzyme, TNFAIP3, or the mRNA-destabilizing protein, ZFP36. Moreover, glucocorticoid receptor cooperativity with factors, including nuclear factor-κB (NF-κB), may enhance regulator expression to promote repression. Conversely, MAPKs, which are inhibited by glucocorticoids, provide feedforward control to limit expression of the transcription factor IRF1, and the chemokine, CXCL10. We propose that modulation of feedback and feedforward control can determine repression or resistance of inflammatory gene expression toglucocorticoid. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

Prostaglandin E₂ regulates cellular migration via induction of vascular endothelial growth factor receptor-1 in HCA-7 human colon cancer cells.

PubMed

Fujino, Hiromichi; Toyomura, Kaori; Chen, Xiao-bo; Regan, John W; Murayama, Toshihiko

2011-02-01

An important event in the development of tumors is angiogenesis, or the formation of new blood vessels. Angiogenesis is also known to be involved in tumor cell metastasis and is dependent upon the activity of the vascular endothelial growth factor (VEGF) signaling pathway. Studies of mice in which the EP3 prostanoid receptors have been genetically deleted have shown a role for these receptors in cancer growth and angiogenesis. In the present study, human colon cancer HCA-7 cells were used as a model system to understand the potential role of EP3 receptors in tumor cell migration. We now show that stimulation of HCA-7 cells with PGE₂ enhanced the up-regulation of VEGF receptor-1 (VEGFR-1) expression by a mechanism involving EP3 receptor-mediated activation of phosphatidylinositol 3-kinase and the extracellular signal-regulated kinases. Moreover, the PGE₂ stimulated increase in VEGFR-1 expression was accompanied by an increase in the cellular migration of HCA-7 cells. Given the known involvement of VEGFR-1 in cellular migration, our results suggest that EP3 receptors may contribute to tumor cell metastasis by increasing cellular migration through the up-regulation of VEGFR-1 signaling. Copyright © 2010 Elsevier Inc. All rights reserved.

The effects of abnormalities of glucose homeostasis on the expression and binding of muscarinic receptors in cerebral cortex of rats.

PubMed

Sherin, Antony; Peeyush, Kumar T; Naijil, George; Nandhu, Mohan Sobhana; Jayanarayanan, Sadanandan; Jes, Paul; Paulose, Cheramadathikudiyil Skaria

2011-01-25

Glucose homeostasis in humans is an important factor for the functioning of nervous system. Both hypo and hyperglycemia contributes to neuronal functional deficit. In the present study, effect of insulin induced hypoglycemia and streptozotocin induced diabetes on muscarinic receptor binding, cholinergic enzymes; AChE, ChAT expression and GLUT3 in the cerebral cortex of experimental rats were analysed. Total muscarinic, muscarinic M(1) receptor showed a significant decrease and muscarinic M(3) receptor subtype showed a significant increased binding in the cerebral cortex of hypoglycemic rats compared to diabetic and control. Real-Time PCR analysis of muscarinic M(1), M(3) receptor subtypes confirmed the receptor binding studies. Immunohistochemistry of muscarinic M(1), M(3) receptors using specific antibodies were also carried out. AChE and GLUT3 expression up regulated and ChAT expression down regulated in hypoglycemic rats compared to diabetic and control rats. Our results showed that hypo/hyperglycemia caused impaired glucose transport in neuronal cells as shown by altered expression of GLUT3. Increased AChE and decreased ChAT expression is suggested to alter cortical acetylcholine metabolism in experimental rats along with altered muscarinic receptor binding in hypo/hyperglycemic rats, impair cholinergic transmission, which subsequently lead to cholinergic dysfunction thereby causing learning and memory deficits. We observed a prominent cholinergic functional disturbance in hypoglycemic condition than in hyperglycemia. Hypoglycemia exacerbated the neurochemical changes in cerebral cortex induced by hyperglycemia. These findings have implications for both therapy and identification of causes contributing to neuronal dysfunction in diabetes. Copyright © 2010 Elsevier B.V. All rights reserved.

Pattern of cytokine receptors expressed by human dendritic cells migrated from dermal explants.

PubMed Central

Larregina, A T; Morelli, A E; Kolkowski, E; Sanjuan, N; Barboza, M E; Fainboim, L

1997-01-01

Different reasons account for the lack of information about the expression of cytokine receptors on human dendritic cells (DC): (a) DC are a trace population; (b) the proteolytic treatment used to isolate DC may alter enzyme-sensitive epitopes; and (c) low numbers of receptors per cell. In the present work the expression of cytokine receptors was analysed by flow cytometry on the population of dermal DC (DDC) that spontaneously migrate from short-term culture dermal explants. DDC obtained after dermal culture were CD1alow, CD1b+, CD1c+, human leucocyte antigen (HLA)-DR+, CD11chigh, CD11b+ and CD32+. The DC lineage was confirmed by ultrastructural analysis. DDC expressed interleukin (IL)-1R type 1 (monoclonal antibody (mAb) hIL-1R1-M1; and 6B5); IL-1R type 2 (mAb hIL-1R2-M22); IL-2R alpha chain (mAb anti-Tac; and hIL-2R-M1) and IL-2R gamma chain (mAb 3B5; and AG14C). DDC did not stain for IL-2R beta chain using four mAbs recognizing two different epitopes of IL-2R beta (mAb 2R-B; Mik-beta 1; and CF1; Mik-beta 3, respectively). DDC were also positive for the cytokine binding chains (alpha chains) of IL-3R (mAb 9F5); IL-4R (mAb hIL-4R-M57; and S456C9); and IL-7R (mAb hIL-7R-M20; and R3434). DDC showed low levels of IL-6R alpha chain (mAb B-F19; B-R6; and B-E23) and its signal transducer gp130 (mAb A2; and B1). DDC strongly expressed interferon-gamma receptor (IFN-gamma R) (mAb GIR-208) and were negative for IL-8R (mAb B-G20; and B-F25). All DDC were highly positive for granulocyte-macrophage colony-stimulating factor receptor (GM-CSFR) alpha chain (mAb hGM-CSFR-M1; SC06; SC04, and 8G6) and to a lesser extent for the common beta chain of GM-CSFR, IL-3R and IL-5R (mAb 3D7). On the other hand, reactivity was not found for granulocyte colony-stimulating factor receptor (G-CSFR) (mAb hGCSFR-M1) nor macrophage colony-stimulating factor receptor (M-CSFR) (mAb 7-7A3-17) confirming the DC lineage of DDC. As previously reported for lymphoid DC, DDC expressed tumour necrosis factor receptort (TNFR) 75000 MW (mAb utr-1; hTNFR-M1; and MR2-1) but lacked TNFR 55000 MW (mAb htr-9; MR1-1; and MR1-2). In summary, DDC express receptors for a broad panel of cytokines, even receptors for cytokines whose effects on DC are still unknown (i.e. IL-2R alpha gamma; IL-6R alpha/gp 130; IL-7R alpha gamma). Images Figure 1 PMID:9227332

Sphingosine-1-phosphate receptor expression in cardiac fibroblasts is modulated by in vitro culture conditions.

PubMed

Landeen, Lee K; Aroonsakool, Nakon; Haga, Jason H; Hu, Betty S; Giles, Wayne R

2007-06-01

The bioactive molecule sphingosine-1-phosphate (S1P) binds with high affinity to five recognized receptors (S1P(1-5)) to affect various tissues, including cellular responses of cardiac fibroblasts (CFbs) and myocytes. CFbs are essential components of myocardium, and detailed study of their cell signaling and physiology is required for a number of emerging disciplines. Meaningful studies on CFbs, however, necessitate methods for selective, reproducible cell isolations. Macrophages reside within normal cardiac tissues and often are isolated with CFbs. A protocol was therefore developed that significantly reduces macrophage levels and utilizes more CFb-specific markers (discoidin domain receptor-2) instead of, or in addition to, more commonly used cytoskeletal markers. Our results demonstrate that primary isolated, purified CFbs express predominantly S1P(1-3); however, the relative levels of these receptor subtypes are modulated with time and by culture conditions. In coculture experiments, macrophages altered CFb S1P receptor levels relative to controls. Further investigations using known macrophage-secreted factors showed that S1P and H(2)O(2) had minimal effects on CFb S1P(1-3) expression, whereas transforming growth factor-beta1, TNF-alpha, and PDGF-BB significantly altered all S1P receptor subtypes. Lowering FBS concentrations from 10% to 0.1% increased S1P(2), whereas supplementation with either PDGF-BB or Rho-associated protein kinase inhibitor Y-27632 significantly elevated S1P(3) levels. S1P(2) and S1P(3) receptor levels are known to regulate cell migration. Using cells isolated from either normal or S1P(3)-null mice, we demonstrate that S1P(3) is important and necessary for CFb migration. These results highlight the importance of demonstrating CFb culture purity in functional studies of S1P and also identify conditions that modulate S1P receptor expression in CFbs.

[On the role of selective silencer Freud-1 in the regulation of the brain 5-HT(1A) receptor gene expression].

PubMed

Naumenko, V S; Osipova, D V; Tsybko, A S

2010-01-01

Selective 5-HT(1A) receptor silencer (Freud-1) is known to be one of the main factors for transcriptional regulation of brain serotonin 5-HT(1A) receptor. However, there is a lack of data on implication of Freud-1 in the mechanisms underlying genetically determined and experimentally altered 5-HT(1A) receptor system state in vivo. In the present study we have found a difference in the 5-HT(1A) gene expression in the midbrain of AKR and CBA inbred mouse strains. At the same time no distinction in Freud-1 expression was observed. We have revealed 90.3% of homology between mouse and rat 5-HT(1A) receptor DRE-element, whereas there was no difference in DRE-element sequence between AKR and CBA mice. This indicates the absence of differences in Freud-1 binding site in these mouse strains. In the model of 5-HT(1A) receptor desensitization produced by chronic 5-HT(1A) receptor agonist administration, a significant reduction of 5-HT(1A) receptor gene expression together with considerable increase of Freud-1 expression were found. These data allow us to conclude that the selective silencer of 5-HT(1A) receptor, Freud-1, is involved in the compensatory mechanisms that modulate the functional state of brain serotonin system, although it is not the only factor for 5-HT(1A) receptor transcriptional regulation.

N-3 polyunsaturated fatty acid regulation of hepatic gene transcription

PubMed Central

Jump, Donald B.

2009-01-01

Purpose of review The liver plays a central role in whole body lipid metabolism and adapts rapidly to changes in dietary fat composition. This adaption involves changes in the expression of genes involved in glycolysis, de-novo lipogenesis, fatty acid elongation, desaturation and oxidation. This review brings together metabolic and molecular studies that help explain n-3 (omega-3) polyunsaturated fatty acid regulation of hepatic gene transcription. Recent findings Dietary n-3 polyunsaturated fatty acid regulates hepatic gene expression by targeting three major transcriptional regulatory networks: peroxisome proliferator-activated receptor α, sterol regulatory element binding protein-1 and the carbohydrate regulatory element binding protein/Max-like factor X heterodimer. 22 : 6,n-3, the most prominent n-3 polyunsaturated fatty acid in tissues, is a weak activator of peroxisome proliferator-activated receptor α. Hepatic metabolism of 22 : 6,n-3, however, generates 20 : 5,n-3, a strong peroxisome proliferator-activated receptor α activator. In contrast to peroxisome proliferator-activated receptor α, 22 : 6,n-3 is the most potent fatty acid regulator of hepatic sterol regulatory element binding protein-1. 22 : 6,n-3 suppresses sterol regulatory element binding protein-1 gene expression while enhancing degradation of nuclear sterol regulatory element binding protein-1 through 26S proteasome and Erk1/2-dependent mechanisms. Both n-3 and n-6 polyunsaturated fatty acid suppress carbohydrate regulatory element binding protein and Max-like factor X nuclear abundance and interfere with glucose-regulated hepatic metabolism. Summary These studies have revealed unique mechanisms by which specific polyunsaturated fatty acids control peroxisome proliferator activated receptor α, sterol regulatory element binding protein-1 and carbohydrate regulatory element binding protein/Max-like factor X function. As such, specific metabolic and signal transduction pathways contribute significantly to the fatty acid regulation of these transcription factors and their corresponding regulatory networks. PMID:18460914

Prognostic value of sex-hormone receptor expression in non-muscle-invasive bladder cancer.

PubMed

Nam, Jong Kil; Park, Sung Woo; Lee, Sang Don; Chung, Moon Kee

2014-09-01

We investigated sex-hormone receptor expression as predicting factor of recurrence and progression in patients with non-muscle invasive bladder cancer. We retrospectively evaluated tumor specimens from patients treated for transitional cell carcinoma of the bladder at our institution between January 2006 and January 2011. Performing immunohistochemistry using a monoclonal androgen receptor antibody and monoclonal estrogen receptor-beta antibody on paraffin-embedded tissue sections, we assessed the relationship of immunohistochemistry results and prognostic factors such as recurrence and progression. A total of 169 patients with bladder cancer were evaluated in this study. Sixty-threepatients had expressed androgen receptors and 52 patients had estrogen receptor beta. On univariable analysis, androgen receptor expression was significant lower in recurrence rates (p=0.001), and estrogen receptor beta expression was significant higher in progression rates (p=0.004). On multivariable analysis, significant association was found between androgen receptor expression and lower recurrence rates (hazard ratio=0.500; 95% confidence interval, 0.294 to 0.852; p=0.011), but estrogen receptor beta expression was not significantly associated with progression rates. We concluded that the possibility of recurrence was low when the androgen receptor was expressed in the bladder cancer specimen and it could be the predicting factor of the stage, number of tumors, carcinoma in situ lesion and recurrence.

Novel synergistic mechanism for sst2 somatostatin and TNFalpha receptors to induce apoptosis: crosstalk between NF-kappaB and JNK pathways.

PubMed

Guillermet-Guibert, J; Saint-Laurent, N; Davenne, L; Rochaix, P; Cuvillier, O; Culler, M D; Pradayrol, L; Buscail, L; Susini, C; Bousquet, C

2007-02-01

Somatostatin is a multifunctional hormone that modulates cell proliferation, differentiation and apoptosis. Mechanisms for somatostatin-induced apoptosis are at present mostly unsolved. Therefore, we investigated whether somatostatin receptor subtype 2 (sst2) induces apoptosis in the nontransformed murine fibroblastic NIH3T3 cells. Somatostatin receptor subtype 2 expression induced an executioner caspase-mediated apoptosis through a tyrosine phosphatase SHP-1 (Src homology domain phosphatase-1)-dependent stimulation of nuclear factor kappa B (NF-kappaB) activity and subsequent inhibition of the mitogen-activated protein kinase JNK. Tumor necrosis factor alpha (TNFalpha) stimulated both NF-kappaB and c-Jun NH2-terminal kinase (JNK) activities, which had opposite action on cell survival. Importantly, sst2 sensitized NIH3T3 cells to TNFalpha-induced apoptosis by (1) upregulating TNFalpha receptor protein expression, and sensitizing to TNFalpha-induced caspase-8 activation; (2) enhancing TNFalpha-mediated activation of NF-kappaB, resulting in JNK inhibition and subsequent executioner caspase activation and cell death. We have here unraveled a novel signaling mechanism for a G protein-coupled receptor, which directly triggers apoptosis and crosstalks with a death receptor to enhance death ligand-induced apoptosis.

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.

The CB₁ cannabinoid receptor signals striatal neuroprotection via a PI3K/Akt/mTORC1/BDNF pathway.

PubMed

Blázquez, C; Chiarlone, A; Bellocchio, L; Resel, E; Pruunsild, P; García-Rincón, D; Sendtner, M; Timmusk, T; Lutz, B; Galve-Roperh, I; Guzmán, M

2015-10-01

The CB1 cannabinoid receptor, the main molecular target of endocannabinoids and cannabis active components, is the most abundant G protein-coupled receptor in the mammalian brain. In particular, the CB1 receptor is highly expressed in the basal ganglia, mostly on terminals of medium-sized spiny neurons, where it plays a key neuromodulatory function. The CB1 receptor also confers neuroprotection in various experimental models of striatal damage. However, the assessment of the physiological relevance and therapeutic potential of the CB1 receptor in basal ganglia-related diseases is hampered, at least in part, by the lack of knowledge of the precise mechanism of CB1 receptor neuroprotective activity. Here, by using an array of pharmacological, genetic and pharmacogenetic (designer receptor exclusively activated by designer drug) approaches, we show that (1) CB1 receptor engagement protects striatal cells from excitotoxic death via the phosphatidylinositol 3-kinase/Akt/mammalian target of rapamycin complex 1 pathway, which, in turn, (2) induces brain-derived neurotrophic factor (BDNF) expression through the selective activation of BDNF gene promoter IV, an effect that is mediated by multiple transcription factors. To assess the possible functional impact of the CB1/BDNF axis in a neurodegenerative-disease context in vivo, we conducted experiments in the R6/2 mouse, a well-established model of Huntington's disease, in which the CB1 receptor and BDNF are known to be severely downregulated in the dorsolateral striatum. Adeno-associated viral vector-enforced re-expression of the CB1 receptor in the dorsolateral striatum of R6/2 mice allowed the re-expression of BDNF and the concerted rescue of the neuropathological deficits in these animals. Collectively, these findings unravel a molecular link between CB1 receptor activation and BDNF expression, and support the relevance of the CB1/BDNF axis in promoting striatal neuron survival.

Fibroblast Growth Factor (FGF-2) and Its Receptors FGFR-2 and FGFR-3 May Be Putative Biomarkers of Malignant Transformation of Potentially Malignant Oral Lesions into Oral Squamous Cell Carcinoma.

PubMed

Nayak, Seema; Goel, Madhu Mati; Makker, Annu; Bhatia, Vikram; Chandra, Saumya; Kumar, Sandeep; Agarwal, S P

2015-01-01

There are several factors like angiogenesis, lymphangiogenesis, genetic alterations, mutational factors that are involved in malignant transformation of potentially malignant oral lesions (PMOLs) to oral squamous cell carcinoma (OSCC). Fibroblast growth factor-2 (FGF-2) is one of the prototypes of the large family of growth factors that bind heparin. FGF-2 induces angiogenesis and its receptors may play a role in synthesis of collagen. FGFs are involved in transmission of signals between the epithelium and connective tissue, and influence growth and differentiation of a wide variety of tissue including epithelia. The present study was undertaken to analyze expression of FGF-2 and its receptors FGFR-2 and FGFR-3 in 72 PMOLs, 108 OSCC and 52 healthy controls, and their role in risk assessment for malignant transformation of Leukoplakia (LKP) and Oral submucous fibrosis (OSMF) to OSCC. Immunohistochemistry was performed using antibodies against FGF-2, FGFR-2 and FGFR-3. IHC results were validated by Real Time PCR. Expression of FGF-2, FGFR-2 and FGFR-3 was upregulated from PMOLs to OSCC. While 90% (9/10) of PMOLs which showed malignant transformation (transformed) expressed FGF-2, only 24.19% cases (15/62) of PMOLs which were not transformed (untransformed) to OSCC expressed FGF-2. Similarly, FGFR-2 expression was seen in 16/62 (25.81%) of untransformed PMOLs and 8/10 (80%) cases of transformed PMOLs. FGFR-3 expression was observed in 23/62 (37.10%) cases of untransformed PMOLs and 6/10 (60%) cases of transformed PMOLs. A significant association of FGF-2 and FGFR-2 expression with malignant transformation from PMOLs to OSCC was observed both at phenotypic and molecular level. The results suggest that FGF-2 and FGFR-2 may be useful as biomarkers of malignant transformation in patients with OSMF and LKP.

Regulation of cytokine receptors by Golgi N-glycan processing and endocytosis.

PubMed

Partridge, Emily A; Le Roy, Christine; Di Guglielmo, Gianni M; Pawling, Judy; Cheung, Pam; Granovsky, Maria; Nabi, Ivan R; Wrana, Jeffrey L; Dennis, James W

2004-10-01

The Golgi enzyme beta1,6 N-acetylglucosaminyltransferase V (Mgat5) is up-regulated in carcinomas and promotes the substitution of N-glycan with poly N-acetyllactosamine, the preferred ligand for galectin-3 (Gal-3). Here, we report that expression of Mgat5 sensitized mouse cells to multiple cytokines. Gal-3 cross-linked Mgat5-modified N-glycans on epidermal growth factor and transforming growth factor-beta receptors at the cell surface and delayed their removal by constitutive endocytosis. Mgat5 expression in mammary carcinoma was rate limiting for cytokine signaling and consequently for epithelial-mesenchymal transition, cell motility, and tumor metastasis. Mgat5 also promoted cytokine-mediated leukocyte signaling, phagocytosis, and extravasation in vivo. Thus, conditional regulation of N-glycan processing drives synchronous modification of cytokine receptors, which balances their surface retention against loss via endocytosis.

Characterization of the growth of murine fibroblasts that express human insulin receptors. II. Interaction of insulin with other growth factors

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

Randazzo, P.A.; Jarett, L.

1990-09-01

The effects of insulin-like growth factor-1 (IGF-1), epidermal growth factor (EGF), platelet-derived growth factor (PDGF), and insulin on DNA synthesis were studied in murine fibroblasts transfected with an expression vector containing human insulin receptor cDNA (NIH 3T3/HIR) and the parental NIH 3T3 cells. In NIH 3T3/HIR cells, individual growth factors in serum-free medium stimulated DNA synthesis with the following relative efficacies: insulin greater than or equal to 10% fetal calf serum greater than PDGF greater than IGF-1 much greater than EGF. In comparison, the relative efficacies of these factors in stimulating DNA synthesis by NIH 3T3 cells were 10% fetalmore » calf serum greater than PDGF greater than EGF much greater than IGF-1 = insulin. In NIH 3T3/HIR cells, EGF was synergistic with 1-10 ng/ml insulin but not with 100 ng/ml insulin or more. Synergy of PDGF or IGF-1 with insulin was not detected. In the parental NIH 3T3 cells, insulin and IGF-1 were found to be synergistic with EGF (1 ng/ml), PDGF (100 ng/ml), and PDGF plus EGF. In NIH 3T3/HIR cells, the lack of interaction of insulin with other growth factors was also observed when the percentage of cells synthesizing DNA was examined. Despite insulin's inducing only 60% of NIH 3T3/HIR cells to incorporate thymidine, addition of PDGF, EGF, or PDGF plus EGF had no further effect. In contrast, combinations of growth factors resulted in 95% of the parental NIH 3T3 cells synthesizing DNA. The independence of insulin-stimulated DNA synthesis from other mitogens in the NIH 3T3/HIR cells is atypical for progression factor-stimulated DNA synthesis and is thought to be partly the result of insulin receptor expression in an inappropriate context or quantity.« less

UDP-Glucuronosyltransferase Expression in Mouse Liver Is Increased in Obesity- and Fasting-Induced Steatosis

PubMed Central

Xu, Jialin; Kulkarni, Supriya R.; Li, Liya

2012-01-01

UDP-glucuronosyltransferases (Ugt) catalyze phase II conjugation reactions with glucuronic acid, which enhances chemical polarity and the elimination from the body. Few studies have addressed whether Ugt expression and activity are affected by liver disease, such as steatosis. The purpose of this study was to determine whether steatosis induced by obesity or fasting could affect liver Ugt mRNA expression and activity. Male C57BL/6J and Lepob/ob (ob/ob) mice were fed ad libitum or food was withheld for 24 h. In steatotic livers of ob/ob mice, Ugt1a1, -1a6, -1a9, -2a3, -3a1, and -3a2 mRNA expression increased. Fasting, which also induced steatosis, increased hepatic Ugt1a1, -1a6, -1a7, -1a9, -2b1, -2b5, -2a3, -3a1, and -3a2 mRNA expression in mouse liver. Likewise, acetaminophen glucuronidation increased by 47% in hepatic microsomes from ob/ob mice compared with that in C57BL/6J mice, but not after fasting. In both steatosis models, Ugt induction was accompanied by increased aryl hydrocarbon receptor, constitutive androstane receptor (CAR), peroxisome proliferator-activated receptor (PPAR)-α, pregnane X receptor, nuclear factor (erythroid-derived 2)-like 2 (Nrf2), and peroxisome proliferator-activated receptor-γ coactivator-1α mRNA expression. In addition, fasting increased CAR, PPAR, and Nrf2 binding activity. The work points to hepatic triglyceride concentrations corresponding with nuclear receptor and Ugt expression. The findings indicate that steatosis significantly alters hepatic Ugt expression and activity, which could have a significant impact on determining circulating hormone levels, drug efficacy, and environmental chemical clearance. PMID:22031624

UDP-glucuronosyltransferase expression in mouse liver is increased in obesity- and fasting-induced steatosis.

PubMed

Xu, Jialin; Kulkarni, Supriya R; Li, Liya; Slitt, Angela L

2012-02-01

UDP-glucuronosyltransferases (Ugt) catalyze phase II conjugation reactions with glucuronic acid, which enhances chemical polarity and the elimination from the body. Few studies have addressed whether Ugt expression and activity are affected by liver disease, such as steatosis. The purpose of this study was to determine whether steatosis induced by obesity or fasting could affect liver Ugt mRNA expression and activity. Male C57BL/6J and Lep(ob/ob) (ob/ob) mice were fed ad libitum or food was withheld for 24 h. In steatotic livers of ob/ob mice, Ugt1a1, -1a6, -1a9, -2a3, -3a1, and -3a2 mRNA expression increased. Fasting, which also induced steatosis, increased hepatic Ugt1a1, -1a6, -1a7, -1a9, -2b1, -2b5, -2a3, -3a1, and -3a2 mRNA expression in mouse liver. Likewise, acetaminophen glucuronidation increased by 47% in hepatic microsomes from ob/ob mice compared with that in C57BL/6J mice, but not after fasting. In both steatosis models, Ugt induction was accompanied by increased aryl hydrocarbon receptor, constitutive androstane receptor (CAR), peroxisome proliferator-activated receptor (PPAR)-α, pregnane X receptor, nuclear factor (erythroid-derived 2)-like 2 (Nrf2), and peroxisome proliferator-activated receptor-γ coactivator-1α mRNA expression. In addition, fasting increased CAR, PPAR, and Nrf2 binding activity. The work points to hepatic triglyceride concentrations corresponding with nuclear receptor and Ugt expression. The findings indicate that steatosis significantly alters hepatic Ugt expression and activity, which could have a significant impact on determining circulating hormone levels, drug efficacy, and environmental chemical clearance.

Clinical validation of nuclear factor kappa B expression in invasive breast cancer.

PubMed

Agrawal, Anil Kumar; Pielka, Ewa; Lipinski, Artur; Jelen, Michal; Kielan, Wojciech; Agrawal, Siddarth

2018-01-01

Breast cancer is the most commonly diagnosed cancer in Polish women. The expression of transcription nuclear factor kappa B, a key inducer of inflammatory response promoting carcinogenesis and cancer progression in breast cancer, is not well-established. We assessed the nuclear factor kappa B expression in a total of 119 invasive breast carcinomas and 25 healthy control samples and correlated this expression pattern with several clinical and pathologic parameters including histologic type and grade, tumor size, lymph node status, estrogen receptor status, and progesterone receptor status. The data used for the analysis were derived from medical records. An immunohistochemical analysis of nuclear factor kappa B, estrogen receptor, and progesterone receptor was carried out and evaluation of stainings was performed. The expression of nuclear factor kappa B was significantly higher than that in the corresponding healthy control samples. No statistical difference was demonstrated in nuclear factor kappa B expression in relation to age, menopausal status, lymph node status, tumor size and location, grade and histologic type of tumor, and hormonal status (estrogen receptor and progesterone receptor). Nuclear factor kappa B is significantly overexpressed in invasive breast cancer tissues. Although nuclear factor kappa B status does not correlate with clinicopathological findings, it might provide important additional information on prognosis and become a promising object for targeted therapy.

Insulin-like growth factor-1 prevents dorsal root ganglion neuronal tyrosine kinase receptor expression alterations induced by dideoxycytidine in vitro.

PubMed

Liu, Huaxiang; Lu, Jing; He, Yong; Yuan, Bin; Li, Yizhao; Li, Xingfu

2014-03-01

Dideoxycytidine (zalcitabine, ddC) produces neurotoxic effects. It is particularly important to understand the toxic effects of ddC on different subpopulations of dorsal root ganglion (DRG) neurons which express distinct tyrosine kinase receptor (Trk) and to find therapeutic factors for prevention and therapy for ddC-induced peripheral sensory neuropathy. Insulin-like growth factor-1 (IGF-1) has been shown to have neurotrophic effects on DRG sensory neurons. However, little is known about the effects of ddC on distinct Trk (TrkA, TrkB, and TrkC) expression in DRG neurons and the neuroprotective effects of IGF-1 on ddC-induced neurotoxicity. Here, we have tested the extent to which the expression of TrkA, TrkB, and TrkC receptors in primary cultured DRG neurons is affected by ddC in the presence or absence of IGF-1. In this experiment, we found that exposure of 5, 25, and 50 μmol/L ddC caused a dose-dependent decrease of the mRNA, protein, and the proportion of TrkA-, TrkB-, and TrkC-expressing neurons. IGF-1 (20 nmol/L) could partially reverse the decrease of TrkA and TrkB, but not TrkC, expression with ddC exposure. The phosphatidylinositol 3-kinase (PI3K) inhibitor LY294002 (10 μmol/L) blocked the effects of IGF-1. These results suggested that the subpopulations of DRG neurons which express distinct TrkA, TrkB, and TrkC receptors were affected by ddC exposure. IGF-1 might relieve the ddC-induced toxicity of TrkA- and TrkB-, but not TrkC-expressing DRG neurons. These data offer new clues for a better understanding of the association of ddC with distinct Trk receptor expression and provide new evidence of the potential therapeutic role of IGF-1 on ddC-induced neurotoxicity.

Expression profiles of relaxin family peptides and their receptors indicate their influence on spermatogenesis in the domestic cat (Felis catus).

PubMed

Braun, B C; Müller, K; Jewgenow, K

2015-07-01

Disturbed spermatogenesis is a common problem in felines. Studying spermatogenesis in the domestic cat can improve the understanding of the biological background and help to counteract fertility problems in other feline species. Here, we analyzed 3 relaxin family peptides (relaxin, relaxin-3, and INSL3) and their receptors (RXFP1, RXFP2, and RXFP3) as potential spermatogenic factors involving their expression in the testis at different stages of its development. It may be concluded from its stage-dependent expression that relaxin, together with RXFP1, appears to be involved in the first stage of spermatogenesis, whereas relaxin-3 via binding to RXFP3 influences spermiogenesis. Furthermore, correlations were observed between relaxin, relaxin-3, RXFP1, RXFP2 and RXFP3 messenger RNA expression, and the relative numbers of haploid cells in testes. The peptide INSL3 was highly expressed at all testis development stages. Because of the low and stage-independent expression of its receptor RXFP2, an auto- and/or paracrine function of INSL3 in spermatogenesis seems unlikely. In the adult testis, messenger RNA expression of relaxin, RXFP1, and RXFP3 predominantly occurs in the tubular testis compartment, whereas INLS3 is mainly expressed in the interstitium. Copyright © 2015 Elsevier Inc. All rights reserved.

Estrogen receptor-α, progesterone receptor, and c-erbB/HER-family receptor mRNA detection and phenotype analysis in spontaneous canine models of breast cancer

PubMed Central

Kabir, Farruk M. Lutful; DeInnocentes, Patricia; Agarwal, Payal; Mill, Christopher P.; Riese, David J.

2017-01-01

Well characterized, stable, p16-defective canine mammary cancer (CMT) cell lines and normal canine mammary epithelial cells were used to investigate expression of the major breast cancer-specific hormone receptors estrogen receptor alpha (ER1) and progesterone receptor (PR) as well as luminal epithelial-specific proto-oncogenes encoding c-erbB-1 (epidermal growth factor receptor/EGFr), c-erbB-2/HER2, c-erbB-3, and c-erbB-4 receptors. The investigation developed and validated quantitative reverse transcriptase polymerase chain reaction assays for each transcript to provide rapid assessment of breast cancer phenotypes for canine cancers, based on ER1, PR, and c-erbB-2/HER2 expressions, similar to those in human disease. Roles for relatively underexplored c-erbB-3 and c-erbB-4 receptor expressions in each of these breast cancer phenotypes were also evaluated. Each quantitative assay was validated by assessment of amplicon size and DNA sequencing following amplification. Differential expression of ER1, PR, and c-erbB-2 in CMT cell lines clearly defined distinct human-like breast cancer phenotypes for a selection of CMT-derived cell lines. Expression profiles for EGFr family genes c-erbB-3 and c-erbB-4 in CMT models also provided an enriched classification of canine breast cancer identifying new extended phenotypes beyond the conventional luminal-basal characterization used in human breast cancer. PMID:27515268

TAM receptors support neural stem cell survival, proliferation and neuronal differentiation.

PubMed

Ji, Rui; Meng, Lingbin; Jiang, Xin; Cvm, Naresh Kumar; Ding, Jixiang; Li, Qiutang; Lu, Qingxian

2014-01-01

Tyro3, Axl and Mertk (TAM) receptor tyrosine kinases play multiple functional roles by either providing intrinsic trophic support for cell growth or regulating the expression of target genes that are important in the homeostatic regulation of immune responses. TAM receptors have been shown to regulate adult hippocampal neurogenesis by negatively regulation of glial cell activation in central nervous system (CNS). In the present study, we further demonstrated that all three TAM receptors were expressed by cultured primary neural stem cells (NSCs) and played a direct growth trophic role in NSCs proliferation, neuronal differentiation and survival. The cultured primary NSCs lacking TAM receptors exhibited slower growth, reduced proliferation and increased apoptosis as shown by decreased BrdU incorporation and increased TUNEL labeling, than those from the WT NSCs. In addition, the neuronal differentiation and maturation of the mutant NSCs were impeded, as characterized by less neuronal differentiation (β-tubulin III+) and neurite outgrowth than their WT counterparts. To elucidate the underlying mechanism that the TAM receptors play on the differentiating NSCs, we examined the expression profile of neurotrophins and their receptors by real-time qPCR on the total RNAs from hippocampus and primary NSCs; and found that the TKO NSC showed a significant reduction in the expression of both nerve growth factor (NGF) and brain-derived neurotrophic factor (BDNF), but accompanied by compensational increases in the expression of the TrkA, TrkB, TrkC and p75 receptors. These results suggest that TAM receptors support NSCs survival, proliferation and differentiation by regulating expression of neurotrophins, especially the NGF.

Inhibitory Effect of Memantine on Streptozotocin-Induced Insulin Receptor Dysfunction, Neuroinflammation, Amyloidogenesis, and Neurotrophic Factor Decline in Astrocytes.

PubMed

Rajasekar, N; Nath, Chandishwar; Hanif, Kashif; Shukla, Rakesh

2016-12-01

Our earlier studies showed that insulin receptor (IR) dysfunction along with neuroinflammation and amyloidogenesis played a major role in streptozotocin (STZ)-induced toxicity in astrocytes. N-methyl-D-aspartate (NMDA) receptor antagonist-memantine shows beneficial effects in Alzheimer's disease (AD) pathology. However, the protective molecular and cellular mechanism of memantine in astrocytes is not properly understood. Therefore, the present study was undertaken to investigate the effect of memantine on insulin receptors, neurotrophic factors, neuroinflammation, and amyloidogenesis in STZ-treated astrocytes. STZ (100 μM) treatment for 24 h in astrocytes resulted significant decrease in brain-derived neurotrophic factor (BDNF), glial cell line-derived neurotrophic factor (GDNF), and insulin-degrading enzyme (IDE) expression in astrocytes. Treatment with memantine (1-10 μM) improved STZ-induced neurotrophic factor decline (BDNF, GDNF) along with IR dysfunction as evidenced by a significant increase in IR protein expression, phosphorylation of IRS-1, Akt, and GSK-3 α/β in astrocytes. Further, memantine attenuated STZ-induced amyloid precursor protein (APP), β-site APP-cleaving enzyme-1 and amyloid-β 1-42 expression and restored IDE expression in astrocytes. In addition, memantine also displays protective effects against STZ-induced astrocyte activation showed by reduction of inflammatory markers, nuclear factor kappa-B translocation, glial fibrillary acidic protein, cyclooxygenase-2, tumor necrosis factor-α level, and oxidative-nitrostative stress. The results suggest that besides the NMDA receptor antagonisic activity, effect on astroglial IR and neurotrophic factor may also be an important factor in the beneficial effect of memantine in AD pathology. Graphical Abstract Novel neuroprotective mechanisms of memenatine in streptozotocin-induced toxicity in astrocytes.

Equine insulin receptor and insulin-like growth factor-1 receptor expression in digital lamellar tissue and insulin target tissues.

PubMed

Kullmann, A; Weber, P S; Bishop, J B; Roux, T M; Norby, B; Burns, T A; McCutcheon, L J; Belknap, J K; Geor, R J

2016-09-01

Hyperinsulinaemia is implicated in the pathogenesis of endocrinopathic laminitis. Insulin can bind to different receptors: two insulin receptor isoforms (InsR-A and InsR-B), insulin-like growth factor-1 receptor (IGF-1R) and InsR/IGF-1R hybrid receptor (Hybrid). Currently, mRNA expression of these receptors in equine tissues and the influence of body type and dietary carbohydrate intake on expression of these receptors is not known. The study objectives were to characterise InsR-A, InsR-B, IGF-1R and Hybrid expression in lamellar tissue (LT) and insulin responsive tissues from horses and examine the effect of dietary nonstructural carbohydrate (NSC) on mRNA expression of these receptors in LT, skeletal muscle, liver and two adipose tissue (AT) depots of lean and obese ponies. In vivo experiment. Lamellar tissue samples were evaluated by quantitative reverse transcription polymerase chain reaction (RT-qPCR) for receptor mRNA expression (n = 8) and immunoblotting for protein expression (n = 3). Archived LT, skeletal muscle, liver and AT from lean and obese mixed-breed ponies fed either a low (~7% NSC as dry matter; 5 lean, 5 obese) or high NSC diet (~42% NSC as dry matter; 6 lean, 6 obese) for 7 days were evaluated by RT-qPCR to determine the effect of body condition and diet on expression of the receptors in different tissues. Significance was set at P≤0.05. Lamellar tissue expresses both InsR isoforms, IGF-1R and Hybrid. LT IGF-1R gene expression was greater than either InsR isoform and InsR-A expression was greater than InsR-B (P≤0.05). Obesity significantly lowered IGF-1R, InsR-A and InsR-B mRNA expression in LT and InsR-A in tailhead AT. High NSC diet lowered expression of all three receptor types in liver; IGF-1R and InsR-A in LT and InsR-A in tailhead AT. Lamellar tissue expresses IGF-1R, InsR isoforms and Hybrids. The functional characteristics of these receptors and their role in endocrinopathic laminitis warrants further investigation. © 2015 EVJ Ltd.

Human TRAF3 adaptor molecule deficiency leads to impaired Toll-like receptor 3 response and susceptibility to herpes simplex encephalitis

PubMed Central

de Diego, Rebeca Pérez; Sancho-Shimizu, Vanessa; Lorenzo, Lazaro; Puel, Anne; Plancoulaine, Sabine; Picard, Capucine; Herman, Melina; Cardon, Annabelle; Durandy, Anne; Bustamante, Jacinta; Vallabhapurapu, Sivakumar; Bravo, Jerónimo; Warnatz, Klaus; Chaix, Yves; Cascarrigny, Françoise; Lebon, Pierre; Rozenberg, Flore; Karin, Michael; Tardieu, Marc; Al-Muhsen, Saleh; Jouanguy, Emmanuelle; Zhang, Shen-Ying; Abel, Laurent; Casanova, Jean-Laurent

2010-01-01

Tumor necrosis factor (TNF) receptor-associated factor 3 (TRAF3) functions downstream of multiple receptors that induce interferon-α (IFN-α), IFN–β and IFN-λ production, including Toll-like receptor 3 (TLR3), which is deficient in some patients with herpes simplex virus-1 encephalitis (HSE). Mice lacking TRAF3 die in the neonatal period, preventing direct investigation of the role of TRAF3 in immune responses and host defenses in vivo. Here we reported the autosomal dominant, human TRAF3 deficiency in a young adult with a history of HSE in childhood. The TRAF3 mutant allele was a loss-of-expression, loss-of-function, dominant-negative phenotype, and was associated with impaired, but not abolished TRAF3-dependent responses upon stimulation of both TNF receptors and receptors that induce IFN production. TRAF3 deficiency was associated with a clinical phenotype limited to HSE resulting from the impairment of TLR3-dependent induction of IFN. Thus, TLR3-mediated immunity against primary infection by HSV-1 in the central nervous system is critically dependent on TRAF3. Highlight sentence Autosomal dominant TRAF3 deficiency is a genetic etiology of herpes simplex encephalitis. Highlight sentence R118W TRAF3 allele is loss-of-function, loss-of-expression, and dominant-negative. Highlight sentence Human TRAF3 deficiency impairs the TLR3-dependent induction of anti-viral interferons. PMID:20832341

Endothelial Snail Regulates Capillary Branching Morphogenesis via Vascular Endothelial Growth Factor Receptor 3 Expression

PubMed Central

Park, Jeong Ae; Kim, Dong Young; Kim, Young-Myeong; Kwon, Young-Guen

2015-01-01

Vascular branching morphogenesis is activated and maintained by several signaling pathways. Among them, vascular endothelial growth factor receptor 2 (VEGFR2) signaling is largely presented in arteries, and VEGFR3 signaling is in veins and capillaries. Recent reports have documented that Snail, a well-known epithelial-to-mesenchymal transition protein, is expressed in endothelial cells, where it regulates sprouting angiogenesis and embryonic vascular development. Here, we identified Snail as a regulator of VEGFR3 expression during capillary branching morphogenesis. Snail was dramatically upregulated in sprouting vessels in the developing retinal vasculature, including the leading-edged vessels and vertical sprouting vessels for capillary extension toward the deep retina. Results from in vitro functional studies demonstrate that Snail expression colocalized with VEGFR3 and upregulated VEGFR3 mRNA by directly binding to the VEGFR3 promoter via cooperating with early growth response protein-1. Snail knockdown in postnatal mice attenuated the formation of the deep capillary plexus, not only by impairing vertical sprouting vessels but also by downregulating VEGFR3 expression. Collectively, these data suggest that the Snail-VEGFR3 axis controls capillary extension, especially in vessels expressing VEGFR2 at low levels. PMID:26147525

The multiple roles of the innate immune system in the regulation of apoptosis and inflammation in the brain.

PubMed

Griffiths, Mark R; Gasque, Philippe; Neal, James W

2009-03-01

Central nervous system (CNS) tissues contain cells (i.e. glia and neurons) that have innate immune functions. These cells express a range of receptors that are capable of detecting and clearing apoptotic cells and regulating inflammatory responses. Phagocytosis of apoptotic cells is a nonphlogistic (i.e. noninflammatory) process that provides immune regulation through anti-inflammatory cytokines andregulatory T cells. Neurons and glia express cellular death signals, including CD95Fas/CD95L, FasL, tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) and tumor necrosis factor receptor 1 (TNFR), through which they can trigger apoptosis in T cells and other infiltrating cells. Microglia, astrocytes, ependymal cells, and neurons express defense collagens and scavenger and phagocytic receptors that recognize apoptotic cells displaying apoptotic cell-associated molecular patterns, which serve as markers of "altered self." Glia also express pentraxins and complement proteins (C1q, C3b, and iC3b) that opsonize apoptotic cells, making them targets for the phagocytic receptors CR3 and CR4. Immunoregulatory molecules such as the complement regulator CD46 are lost from apoptotic cells and stimulate phagocytosis, whereas the expression of CD47 and CD200 is upregulated during apoptosis; this inhibits proinflammatory microglial cytokine expression, thereby reducing the severity of inflammation. This review outlines the cellular pathways used for the detection and phagocytosis of apoptotic cells in vitro and in experimental models of CNS inflammation.

An Insulin-Like Growth Factor 1 Receptor Inhibitor Induces CYP3A4 Expression through a Pregnane X Receptor-Independent, Noncanonical Constitutive Androstane Receptor-Related Mechanism

PubMed Central

Li, Linhao; Sinz, Michael W.; Zimmermann, Kurt

2012-01-01

Inhibition of insulin-like growth factor-1 receptor (IGF-1R) signaling represents an attractive therapeutic strategy for cancer treatment. A first-generation IGF-1R inhibitor (R)-4-(3-(3-chlorophenyl)-3-hydroxypropyl)-3-(4-methyl-6-morpholino-1H-benzo[d]imidazol-2-yl)pyridin-2(1H)-one (BMS-536924), however, was associated with potent CYP3A4 induction mediated by pregnane X receptor (PXR; NR1I2) transactivation. Structural activity-based modification led to the synthesis of 4-(1-(2-(4-((2-(4-chloro-1H-pyrazol-1-yl)ethyl)amino)-2-oxo-1,2-dihydropyridin-3-yl)-4-methyl-1H-benzo[d]imidazol-6-yl)piperidin-4-yl) piperazine-1-carboxylate (BMS-665351) with no PXR activity while maintaining its ability to inhibit IGF-1R. However, BMS-665351 significantly induces CYP3A4 expression in human primary hepatocytes (HPHs). Here, we report a novel nonclassical constitutive androstane receptor (CAR; NR1I3)-related pathway of BMS-665351-mediated CYP3A4 induction. BMS-665351 treatment resulted in the significant induction of CYP3A4 in HPHs and HepG2 cells, but failed to activate either PXR or CAR in cell-based reporter assays. Moreover, BMS-665351 at concentrations that induce CYP3A4 expression was unable to translocate human CAR from the cytoplasm to the nucleus of HPHs, which represents the initial step of CAR activation. Nevertheless, quantitative polymerase chain reaction analysis demonstrated that BMS-665351 significantly enhanced the expression of CYP3A4 in CAR- but not PXR-transfected HepG2 and Huh7 cells. It is noteworthy that BMS-665351 selectively induced the expression of CAR but not PXR in all tested hepatic cell systems. Synergistic induction of CYP3A4 was observed in HPHs cotreated with BMS-665351 and prototypical activators of CAR but not PXR. In summary, our results indicate that BMS-665351-mediated induction of CYP3A4 is CAR-dependent, but BMS-665351 itself is not a typical activator of either CAR or PXR, rather it functions as a selective inducer of CAR expression and increases CYP3A4 through a noncanonical CAR-related mechanism. PMID:22171088

Tumor necrosis factor receptor-associated factor 6 (TRAF6) participates in anti-lipopolysaccharide factors (ALFs) gene expression in mud crab.

PubMed

Sun, Wan-Wei; Zhang, Xin-Xu; Wan, Wei-Song; Wang, Shu-Qi; Wen, Xiao-Bo; Zheng, Huai-Ping; Zhang, Yue-Ling; Li, Sheng-Kang

2017-02-01

Tumor necrosis factor receptor-associated factor 6 (TRAF6) is a key cytoplasm signal adaptor that mediates signals activated by tumor necrosis factor receptor (TNFR) superfamily and the Interleukin-1 receptor/Toll-like receptor (IL-1/TLR) superfamily. The full-length 2492 bp TRAF6 (Sp-TRAF6) from Scylla paramamosain contains 1800 bp of open reading frame (ORF) encoding 598 amino acids, including an N-terminal RING-type zinc finger, two TRAF-type zinc fingers and a conserved C-terminal meprin and TRAF homology (MATH) domain. Multiple alignment analysis shows that the putative amino acid sequence of Sp-TRAf6 has highest identity of 88% with Pt-TRAF6 from Portunus trituberculatus, while the similarity of Sp-TRAF6 with other crustacean sequences was 54-55%. RT-PCR analysis indicated that Sp-TRAF6 transcripts were predominantly expressed in the hepatopancreas and stomach, whereas it was barely detected in the heart and hemocytes in our study. Moreover, Sp-TRAF6 transcripts were significantly up-regulated after Vibrio parahemolyticus and LPS challenges. RNA interference assay was carried out used by siRNA to investigate the genes expression patterns regulated by Sp-TRAF6. The qRT-PCR results showed that silencing Sp-TRAF6 gene could inhibit SpALF1, SpALF2, SpALF5 and SpALF6 expression in hemocytes, while inhibit SpALF1, SpALF3, SpALF4, SpALF5 and SpALF6 expression in hepatopancreas. Taken together, the acute-phase response to immune challenges and the inhibition of SpALFs gene expression indicate that Sp-TRAF6 plays an important role in host defense against pathogen invasions via regulation of ALF gene expression in S. paramamosain. Copyright © 2016. Published by Elsevier Ltd.

Characterization of an ICII82, 780-Induced, Estrogen Receptor (ER)-beta Mediated Apoptotic Pathway in Prostate Cancer Cells and Establishment of (ER)-beta-Regulated Electrophile-Processing Phase II Enzyme Downregulation as a Promotional Factor in Human Prostatic Carcinogenesis

DTIC Science & Technology

2001-05-01

absent in higher-grade cancers. ERP3 expression reappeared in metastatic prostatic carcinomas . The results were also compared to ERMo and androgen...metastatic prostatic carcinomas in bone and lymph nodes. The results were compared to ERax and androgen receptor expressions in those samples. Taken... carcinomas of the peripheral zone were in contrast strongly positive for the 13 receptor whereas its expression in grade 415 cancers was negligible to

Expression of GnRH receptor in the canine corpus luteum, and luteal function following deslorelin acetate-induced puberty delay.

PubMed

Kaya, D; Gram, A; Kowalewski, M P; Schäfer-Somi, S; Kuru, M; Boos, A; Aslan, S

2017-12-01

The goals of this study were as follows: (Experiment 1) to examine the basic capability of canine corpora lutea (CL) to respond to GnRH by assessing expression of gonadotropin-releasing hormone receptor (GnRH-R) in luteal samples collected throughout the luteal lifespan from non-pregnant dogs, and (Experiment 2) to investigate the effects of pre-pubertal application of the GnRH agonist deslorelin acetate on luteal function following the first oestrus. Mature CL were collected during the mid-luteal phase (days 30-45) from treated and control bitches. Transcript levels of several factors were determined: estrogen receptors (ESR1/ERα, ESR2/ERβ), progesterone (P4)-receptor (PGR), prolactin receptor (PRLR), PGE2-synthase (PTGES) and PGE2 receptors (PTGER2/EP2, PTGER4/EP4), vascular endothelial growth factor (VEGFA) and VEGF receptors (VEGFR1 and VEGFR2), cyclooxygenase 2 (COX2/PTGS2), steroidogenic acute regulatory protein (STAR) and 3β-hydroxysteroid dehydrogenase (3βHSD). Additionally, levels of Kisspeptin 1 (Kiss1) and its receptor (KISS1-R) were evaluated. Although generally low, GnRH-R expression was time dependent and was elevated during early dioestrus, with a significant decrease towards luteal regression. In deslorelin-treated and control dogs, its expression was either low or frequently below the detection limit. EP2 and VEGFR1 were higher in the treated group, which could be caused by a feedback mechanism after long-term suppression of reproductive activity. Despite large individual variations, 3βHSD was higher in the deslorelin-treated group. This, along with unchanged STAR expression, was apparently not mirrored in increased luteal functionality, because similar P4 levels were detected in both groups. Finally, the deslorelin-mediated long-term delay of puberty does not have negative carry-over effects on subsequent ovarian functionality in bitches. © 2017 Blackwell Verlag GmbH.

γ-Tocotrienol upregulates aryl hydrocarbon receptor expression and enhances the anticancer effect of baicalein

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

Yamashita, Shuya; Baba, Kiwako; Makio, Akiko

2016-05-13

Previous studies have identified biomolecules that mediate the physiological actions of food factors, such as amino acids, vitamins, fatty acids, minerals, plant polyphenols, and lactobacilli, suggesting that our bodies are equipped with an innate system that senses which food factors are required to maintain our health. However, the effects of environmental factors on food factor sensing (FFS) remains largely unknown. Tocotorienols (T3s), which belongs to the vitamin E family, possess several physiological functions, including cholesterol lowering and neuroprotective effects. Here, we investigated the effects of naturally abundant γ-T3 on FFS-related gene expressions in melanoma using a DNA chip. Our resultsmore » showed that γ-T3 increased the expression level of aryl hydrocarbon receptor (AhR), a sensing molecule to plant polyphenol baicalein. The co-treatment with γ-T3 and baicalein enhanced the anti-proliferative activity of baicalein, accompanied by the downstream events of AhR-activation induced by baicalein. These data suggest that γ-T3 upregulates AhR expression and enhances its sensitivity to baicalein. - Highlights: • γ-T3 upregulated the expression of AhR in mouse melanoma. • Promotion of the binding activity of Sp1 is associated with the increasing effect of γ-T3 on AhR expression. • γ-T3 enhanced the anti-proliferative activity of baicalein that has an AhR ligand activity. • γ-T3 enhanced the inducing activity of baicalein on the expression of AhR target genes.« less

MicroRNA-22 and microRNA-140 suppress NF-{kappa}B activity by regulating the expression of NF-{kappa}B coactivators

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

Takata, Akemi; Otsuka, Motoyuki, E-mail: otsukamo-tky@umin.ac.jp; Kojima, Kentaro

2011-08-12

Highlights: {yields} miRNAs were screened for their ability to regulate NF-{kappa}B activity. {yields} miRNA-22 and miRNA-140-3p suppress NF-{kappa}B activity by regulating coactivators. {yields} miRNA-22 targets nuclear receptor coactivator 1 (NCOA1). {yields} miRNA-140-3p targets nuclear receptor-interacting protein 1 (NRIP1). -- Abstract: Nuclear factor {kappa}B (NF-{kappa}B) is a transcription factor that regulates a set of genes that are critical to many biological phenomena, including liver tumorigenesis. To identify microRNAs (miRNAs) that regulate NF-{kappa}B activity in the liver, we screened 60 miRNAs expressed in hepatocytes for their ability to modulate NF-{kappa}B activity. We found that miRNA-22 and miRNA-140-3p significantly suppressed NF-{kappa}B activity bymore » regulating the expression of nuclear receptor coactivator 1 (NCOA1) and nuclear receptor-interacting protein 1 (NRIP1), both of which are NF-{kappa}B coactivators. Our results provide new information about the roles of miRNAs in the regulation of NF-{kappa}B activity.« less

Brain-derived neurotrophic factor and its receptors in Bergmann glia cells.

PubMed

Poblete-Naredo, Irais; Guillem, Alain M; Juárez, Claudia; Zepeda, Rossana C; Ramírez, Leticia; Caba, Mario; Hernández-Kelly, Luisa C; Aguilera, José; López-Bayghen, Esther; Ortega, Arturo

2011-12-01

Brain-derived neurotrophic factor is an abundant and widely distributed neurotrophin expressed in the Central Nervous System. It is critically involved in neuronal differentiation and survival. The expression of brain-derived neurotrophic factor and that of its catalytic active cognate receptor (TrkB) has been extensively studied in neuronal cells but their expression and function in glial cells is still controversial. Despite of this fact, brain-derived neurotrophic factor is released from astrocytes upon glutamate stimulation. A suitable model to study glia/neuronal interactions, in the context of glutamatergic synapses, is the well-characterized culture of chick cerebellar Bergmann glia cells. Using, this system, we show here that BDNF and its functional receptor are present in Bergmann glia and that BDNF stimulation is linked to the activation of the phosphatidyl-inositol 3 kinase/protein kinase C/mitogen-activated protein kinase/Activator Protein-1 signaling pathway. Accordingly, reverse transcription-polymerase chain reaction (RT-PCR) experiments predicted the expression of full-length and truncated TrkB isoforms. Our results suggest that Bergmann glia cells are able to express and respond to BDNF stimulation favoring the notion of their pivotal role in neuroprotection. Copyright © 2011 Elsevier B.V. All rights reserved.

Identifying Environmental Chemicals as Agonists of the Androgen Receptor by Applying a Quantitative High-throughput Screening Platform

EPA Science Inventory

Background: The androgen receptor (AR, NR3C4) is a nuclear receptor whose main function is acting as a transcription factor regulating gene expression for male sexual development and maintaining accessory sexual organ function. It is also a necessary component of female fertility...

Neurotrophin NT3 promotes ovarian primordial to primary follicle transition

PubMed Central

Nilsson, Eric; Dole, Gretchen; Skinner, Michael K

2017-01-01

Neurotrophins are growth factors that are known to have a role in promoting cell survival and differentiation. The focus of the current study is to examine the role of neurotrophins in regulating ovarian primordial follicle development. Ovaries from 4-day old rats were placed into organ culture and cultured for 10 days in the absence or presence of neurotrophin-3 (NT3), brain-derived neurotrophic factor (BDNF), or nerve growth factor (NGF). Treatment of ovaries with NT3 resulted in a significant (P<0.01) increase in primordial follicle development (i.e. primordial to primary follicle transition). Treatment with BDNF at high doses of 100–250 ng/ml also significantly (P<0.01) increased primordial follicle development, but NGF had no effect. Immunohistochemical studies determined that NT3 was present in granulosa cells, interstitial tissue, and in the oocytes of primordial and primary follicles. The NT3 receptor NTRK3 was present in oocytes at all stages of development. Analysis of ovaries that contain predominantly primordial follicles demonstrated the transcripts for NT3, NTRK3, NGF, and the BDNF/neurotrophin-4 (NT4) receptor NTRK2 are expressed, while BDNF, NT4, and the NGF receptor NTRK1 are not detectable. Inhibition of the NTRK3 receptor with the tyrphostin AG 879 resulted in oocyte death and a significant (P<0.01) reduction in follicle pool size. Inhibition of the NTRK receptors with K252a slowed primordial to primary follicle transition. A microarray analysis demonstrated that a small number of genes were differentially expressed after NT3 treatment. Observations indicate that the neurotrophin NT3, acting through the NTRK3 receptor in oocytes, promotes the primordial to primary follicle transition. PMID:19584175

Minoxidil-induced hair growth is mediated by adenosine in cultured dermal papilla cells: possible involvement of sulfonylurea receptor 2B as a target of minoxidil.

PubMed

Li, M; Marubayashi, A; Nakaya, Y; Fukui, K; Arase, S

2001-12-01

The mechanism by which minoxidil, an adenosine-triphosphate-sensitive potassium channel opener, induces hypertrichosis remains to be elucidated. Minoxidil has been reported to stimulate the production of vascular endothelial growth factor, a possible promoter of hair growth, in cultured dermal papilla cells. The mechanism of production of vascular endothelial growth factor remains unclear, however. We hypothesize that adenosine serves as a mediator of vascular endothelial growth factor production. Minoxidil-induced increases in levels of intracellular Ca(2+) and vascular endothelial growth factor production in cultured dermal papilla cells were found to be inhibited by 8-sulfophenyl theophylline, a specific antagonist for adenosine receptors, suggesting that dermal papilla cells possess adenosine receptors and sulfonylurea receptors, the latter of which is a well-known target receptor for adenosine-triphosphate-sensitive potassium channel openers. The expression of sulfonylurea receptor 2B and of the adenosine A1, A2A, and A2B receptors was detected in dermal papilla cells by means of reverse transcription polymerase chain reaction analysis. In order to determine which of the adenosine receptor subtypes contribute to minoxidil-induced hair growth, the effects of subtype-specific antagonists for adenosine receptors were investigated. Significant inhibition in increase in intracellular calcium level by minoxidil or adenosine was observed as the result of pretreatment with 8-cyclopentyl-1,3-dipropylxanthine, an antagonist for adenosine A1 receptor, but not by 3,7-dimethyl-1-propargyl-xanthine, an antagonist for adenosine A2 receptor, whereas vascular endothelial growth factor production was blocked by both adenosine A1 and A2 receptor antagonists. These results indicate that the effect of minoxidil is mediated by adenosine, which triggers intracellular signal transduction via both adenosine A1 and A2 receptors, and that the expression of sulfonylurea receptor 2B in dermal papilla cells might play a role in the production of adenosine.

alpha1B-Adrenergic receptor phosphorylation and desensitization induced by transforming growth factor-beta.

PubMed Central

Romero-Avila, M Teresa; Flores-Jasso, C Fabián; García-Sáinz, J Adolfo

2002-01-01

Transforming growth factor-beta (TGF-beta) induced alpha(1B)-adrenergic receptor phosphorylation in Rat-1 fibroblasts stably expressing these adrenoceptors. This effect of TGF-beta was rapid, reaching a maximum within 30 min and decreasing thereafter, and concentration-dependent (EC(50) 0.3 pM). The phosphoinositide 3-kinase inhibitors wortmannin and LY294002, and the protein kinase C inhibitors staurosporine, Ro 318220 and bisindolylmaleimide, blocked the effect of this growth factor. alpha(1B)-Adrenergic receptor phosphorylation was associated with desensitization, as indicated by a reduction in the adrenergic-mediated production of [(3)H]inositol phosphates. Phosphorylation of alpha(1B)-adrenergic receptors by TGF-beta was also observed in Cos-1 cells transfected with the receptor. Co-transfection of the dominant-negative mutant of the regulatory subunit of phosphoinositide 3-kinase (Deltap85) inhibited the phosphorylation of alpha(1B)-adrenergic receptors induced by TGF-beta. Our results indicate that activation of TGF-beta receptors induces alpha(1B)-adrenergic receptor phosphorylation and desensitization. The data suggest that phosphoinositide 3-kinase and protein kinase C play key roles in this effect of TGF-beta. PMID:12234252

alpha1B-Adrenergic receptor phosphorylation and desensitization induced by transforming growth factor-beta.

PubMed

Romero-Avila, M Teresa; Flores-Jasso, C Fabián; García-Sáinz, J Adolfo

2002-12-01

Transforming growth factor-beta (TGF-beta) induced alpha(1B)-adrenergic receptor phosphorylation in Rat-1 fibroblasts stably expressing these adrenoceptors. This effect of TGF-beta was rapid, reaching a maximum within 30 min and decreasing thereafter, and concentration-dependent (EC(50) 0.3 pM). The phosphoinositide 3-kinase inhibitors wortmannin and LY294002, and the protein kinase C inhibitors staurosporine, Ro 318220 and bisindolylmaleimide, blocked the effect of this growth factor. alpha(1B)-Adrenergic receptor phosphorylation was associated with desensitization, as indicated by a reduction in the adrenergic-mediated production of [(3)H]inositol phosphates. Phosphorylation of alpha(1B)-adrenergic receptors by TGF-beta was also observed in Cos-1 cells transfected with the receptor. Co-transfection of the dominant-negative mutant of the regulatory subunit of phosphoinositide 3-kinase (Deltap85) inhibited the phosphorylation of alpha(1B)-adrenergic receptors induced by TGF-beta. Our results indicate that activation of TGF-beta receptors induces alpha(1B)-adrenergic receptor phosphorylation and desensitization. The data suggest that phosphoinositide 3-kinase and protein kinase C play key roles in this effect of TGF-beta.

Death Receptor Expression on Blasts in AML Is Associated with Unfavorable Prognosis.

PubMed

Schmohl, Joerg Uwe; Nuebling, Tina; Wild, Julia; Jung, Johannes; Kroell, Tanja; Kanz, Lothar; Salih, Helmut R; Schmetzer, Helga

2015-07-01

Tumor necrosis factor (TNF) receptor family members play a key role in the regulation of biological functions such as differentiation, proliferation and apoptosis of various cell types. We studied co-expression profiles of death receptors from the TNF family [TNF-related apoptosis-inducing ligand receptor (TRAILR) 1 to 3, TNF receptor 1 (TNFR1) and FAS receptor (FAS)] on peripheral blood blasts from 46 patients with acute myeloid leukemia (AML) at first diagnosis by flow cytometry and correlated the obtained specific fluorescence indices (SFI) with morphological, cytogenetic and clinical parameters. We found that the expression of TRAILR2 and R3 was significantly increased in unfavorable risk groups, according to the National Comprehensive Cancer Network. Additionally, cut-off analyses for TRAILR2 and TNFR1 showed significantly shorter overall survival, earlier disease onset, higher proportions of cases with unfavorable prognosis and higher probability of relapse when SFIs were above the established cut-off. We demonstrate that high co-expression of death receptors on blasts is an independent predictor of poor prognosis in AML. Copyright© 2015 International Institute of Anticancer Research (Dr. John G. Delinassios), All rights reserved.

Trefoil factor 3 (TFF3) from human breast milk activates PAR-2 receptors, of the intestinal epithelial cells HT-29, regulating cytokines and defensins.

PubMed

Barrera, G J; Tortolero, G Sanchez

2016-01-01

Trefoil factors are effector molecules in gastrointestinal tract physiology. Each one improves healing of the gastrointestinal tract. Trefoil factors may be grouped into three classes: the gastric peptides (TFF1), spasmolytic peptide (TFF2) and intestinal trefoil factor (TFF3). Significant amounts of TFF3 are present in human breast milk. Previously, we have reported that trefoil factor 3 isolated from human breast milk produces down regulation of cytokines and promotes human beta defensins expression in intestinal epithelial cells. This study aimed to determine the molecular mechanism involved. Here we showed that the presence of TFF3 strongly correlated with protease activated receptors 2 (PAR-2) activation in human intestinal cells. Intracellular calcium ((Ca2+)i)mobilization was induced by the treatment with: 1) TFF3, 2) synthetic PAR-2 agonist peptide. The co-treatment with a synthetic PAR-2 antagonist peptide and TFF3 eliminates the latter's effect. Additionally, we demonstrated the existence of interactions among TFF3 and PAR-2 receptors through far Western blot and co-precipitation. Finally, down regulation of PAR-2 by siRNA resulted in a decrease of TFF3 induced intracellular (Ca2+)i mobilization, cytokine regulation and defensins expression. These findings suggest that TFF3 activates intestinal cells through PAR-2 (Fig. 4, Ref. 19).

Postconditioning: "Toll-erating" mesenteric ischemia-reperfusion injury?

PubMed

Rosero, Olivér; Ónody, Péter; Kovács, Tibor; Molnár, Dávid; Fülöp, András; Lotz, Gábor; Harsányi, László; Szijártó, Attila

2017-04-01

Postconditioning may prove to be a suitable method to decrease ischemia-reperfusion injury of intestine after mesenteric arterial occlusion. Toll-like-receptor-4 is involved in the pathophysiology of organ damage after ischemia-reperfusion; therefore, the aim of our study was to investigate the effect of postconditioning on the mucosal expression of toll-like-receptor-4. Male Wistar rats (n = 10/group) underwent 60 minutes of superior mesenteric artery occlusion followed by 6 hours of reperfusion in 3 groups: sham-operated, ischemia-reperfusion, and a postconditioned group. Postconditioning was performed by 6 alternating cycles of 10 seconds of reperfusion/reocclusion. Blood and tissue samples were collected at the end of reperfusion. Intestinal histopathologic changes and immunohistochemical expression of mucosal caspase-3, antioxidant status, and protein levels of high-mobility group box-1 and toll-like-receptor-4 were assessed. Immunofluorescent labeling and confocal microscopic analysis of toll-like-receptor-4 were performed. Mucosal and serum levels of interleukin-6 and tumor necrosis factor-α protein were measured. Histologic alterations in the postconditioned group were associated with decreased caspase-3 positivity, less toll-like-receptor-4 mRNA, and less protein expression of high-mobility group box-1 and toll-like-receptor-4 in the intestinal villi compared with the ischemia-reperfusion group. Furthermore, a significantly improved antioxidant state of the intestinal mucosa and less mucosal and serum protein levels of interleukin-6 and tumor necrosis factor-α were detected in the postconditioned group. Small intestinal ischemia-reperfusion injury in male Wistar rats caused by the occlusion of the superior mesenteric artery was ameliorated by the use of postconditioning, showing a more favorable inflammatory response, which may be attributed to the decreased mucosal expression of toll-like-receptor-4. Copyright © 2016 Elsevier Inc. All rights reserved.

Hepatic Stellate Cells Express Functional CXCR4: Role in Stromal Cell–Derived Factor-1α–Mediated Stellate Cell Activation

PubMed Central

Hong, Feng; Tuyama, Ana; Lee, Ting Fang; Loke, Johnny; Agarwal, Ritu; Cheng, Xin; Garg, Anita; Fiel, M. Isabel; Schwartz, Myron; Walewski, Jose; Branch, Andrea; Schecter, Alison D.; Bansal, Meena B.

2010-01-01

Chemokine interactions with their receptors have been implicated in hepatic stellate cell (HSC) activation. The hepatic expression of CXCR4 messenger RNA is increased in hepatitis C cirrhotic livers and plasma levels of its endogenous ligand, stromal cell–derived factor-1α (SDF-1α), correlate with increased fibrosis in these patients. The expression of CXCR4 by HSCs has not been reported. We therefore examined whether HSCs express CXCR4 in vivo and in vitro and explored whether SDF-1α/CXCR4 receptor engagement promotes HSC activation, fibrogenesis, and proliferation. The hepatic protein expression of both CXCR4 and SDF-1α is increased in hepatitis C cirrhotic livers and immunoflourescent and immunohistochemical staining confirms that HSCs express CXCR4 in vivo. Immortalized human stellate cells as well as primary human HSCs express CXCR4, and cell surface receptor expression increases with progressive culture-induced activation. Treatment of stellate cells with recombinant SDF-1α increases expression of α-smooth muscle actin and collagen I and stimulates a dose-dependent increase in HSC proliferation. Inhibitor studies suggest that SDF-1α/CXCR4-dependent extracellular signal-regulated kinase 1/2 and Akt phosphorylation mediate effects on collagen I expression and stellate cell proliferation. Conclusion HSCs express CXCR4 receptor in vivo and in vitro. CXCR4 receptor activation by SDF-1α is profibrogenic through its effects on HSC activation, fibrogenesis, and proliferation. Extracellular signal-regulated kinase 1/2 and phosphoinositide 3-kinase pathways mediate SDF-1α–induced effects on HSC expression of collagen I and proliferation. The availability of small molecule inhibitors of CXCR4 make this receptor an appealing target for antifibrotic approaches. PMID:19434726

The Influence of Tumor-Host Interactions in the Stromal Cell-Derived Factor-1/CXCR4 Ligand/Receptor Axis in Determining Metastatic Risk in Breast Cancer

PubMed Central

Hassan, Saima; Ferrario, Cristiano; Saragovi, Uri; Quenneville, Louise; Gaboury, Louis; Baccarelli, Andrea; Salvucci, Ombretta; Basik, Mark

2009-01-01

The chemokine stromal cell-derived factor-1 (SDF-1) may function to attract CXCR4-expressing cancer cells to metastatic organs. We have previously demonstrated that low plasma SDF-1, a host-derived marker, increases distant metastatic risk in breast cancer. We therefore hypothesized that tumors overexpressing the SDF-1 receptor CXCR4 have an enhanced ability to metastasize in patients with low plasma SDF-1 levels. In this study, we determined the prognostic significance of activated CXCR4, or phosphorylated CXCR4 (p-CXCR4), and CXCR7, another receptor for SDF-1. Immunohistochemistry was performed on a tissue microarray built using 237 samples from the same cohort of patients for which we measured plasma SDF-1 levels. We found that the prognostic value of p-CXCR4 expression (hazard ratio or HR, 3.95; P = 0.004) was superior to total CXCR4 expression (HR, 3.20; P = 0.03). The rate of breast cancer-specific mortality was much higher in patients with both high p-CXCR4 expression and low plasma SDF-1 levels (HR, 5.96; P < 0.001) than either low plasma SDF-1 (HR, 3.59; P = 0.01) or high p-CXCR4 expression (HR, 3.83; P = 0.005) alone. The added prognostic value of low plasma SDF-1 was only effective in patients with high p-CXCR4 expression, and as such, provides clinical validation for modulation of the metastatic potential of tumor cells by an inherent host-derived metastatic risk factor. PMID:19497995

Selective inhibition of the platelet-derived growth factor signal transduction pathway by a protein-tyrosine kinase inhibitor of the 2-phenylaminopyrimidine class.

PubMed Central

Buchdunger, E; Zimmermann, J; Mett, H; Meyer, T; Müller, M; Regenass, U; Lydon, N B

1995-01-01

The platelet-derived growth factor (PDGF) receptor is a member of the transmembrane growth factor receptor protein family with intrinsic protein-tyrosine kinase activity. We describe a potent protein-tyrosine kinase inhibitor (CGP 53716) that shows selectivity for the PDGF receptor in vitro and in the cell. The compound shows selectivity for inhibition of PDGF-mediated events such as PDGF receptor autophosphorylation, cellular tyrosine phosphorylation, and c-fos mRNA induction in response to PDGF stimulation of intact cells. In contrast, ligand-induced autophosphorylation of the epidermal growth factor (EGF) receptor, insulin receptor, and the insulin-like growth factor I receptor, as well as c-fos mRNA expression induced by EGF, fibroblast growth factor, and phorbol ester, was insensitive to inhibition by CGP 53716. In antiproliferative assays, the compound was approximately 30-fold more potent in inhibiting PDGF-mediated growth of v-sis-transformed BALB/c 3T3 cells relative to inhibition of EGF-dependent BALB/Mk cells, interleukin-3-dependent FDC-P1 cells, and the T24 bladder carcinoma line. When tested in vivo using highly tumorigenic v-sis- and human c-sis-transformed BALB/c 3T3 cells, CGP 53716 showed antitumor activity at well-tolerated doses. In contrast, CGP 53716 did not show antitumor activity against xenografts of the A431 tumor, which overexpresses the EGF receptor. These findings suggest that CGP 53716 may have therapeutic potential for the treatment of diseases involving abnormal cellular proliferation induced by PDGF receptor activation. Images Fig. 1 Fig. 2 Fig. 3 PMID:7708684

[Expression of AMPA receptors and related protein in immobilization stressed rats and effect of Xiaoyaosan].

PubMed

Yue, Guang-Xin; Wang, Zhu-Feng; Zhang, Qiao-Li; Zhao, Xin; Yue, Li-Feng; Ding, Jie; Chen, Jia-Xu

2008-05-01

To observe protein expression changes of alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptor and related regulatory protein in the hippocampus and amygdala in chronic immobilization stressed rat and Xiaoyaosan's regulatory effect. Rats were tied 3 h per day to establish immobilization stress condition and treatment with Xiaoyaosan. After 7 days and 21 days stress, the protein expression of AMPA receptor subunit (GluR2/3), N-ethylmaleimide sensitive factor (NSF) and protein interacting with C-kinase 1 (PICK1) in hippocampus and amygdala were detected by using Western blot techniques. The expression of GluR2/3, NSF in dentate gyrus (DG) and amygdala were markedly attenuated (P < 0.05) and PICK1 in CA1 region were significantly increased (P < 0.05) in 7 d immobilization stressed rats while 7 days xiaoyaosan treatment showed an effective regulatory result to PICK1's changes. Under 21 days immobilization stressed condition, the expression of GluR2/3, NSF in CA1 region showed an increasing trend, and GluR2/3 showed a markedly increase (P < 0.01), but showed an significantly decreased trend in amygdala, Xiaoyaosan showed an effective result to such changes above (P < 0.05). The expression of PICK1 showed increasing trend in amygdala and xiaoyaosan could lower its expression (P < 0.05). There are different trends of the expression of AMPA receptor in repeat short-term stress versus chronic immobilization stress, and in hippocampal CA1 region versus amygdala. Xiaoyaosan has better regulation effect on the expression of AMPA receptors in the condition of chronic immobilization stress than those of repeat shortterm stress.

Steroid receptor coactivators, HER-2 and HER-3 expression is stimulated by tamoxifen treatment in DMBA-induced breast cancer.

PubMed

Moi, Line L Haugan; Flågeng, Marianne Hauglid; Gjerde, Jennifer; Madsen, Andre; Røst, Therese Halvorsen; Gudbrandsen, Oddrun Anita; Lien, Ernst A; Mellgren, Gunnar

2012-06-15

Steroid receptor coactivators (SRCs) may modulate estrogen receptor (ER) activity and the response to endocrine treatment in breast cancer, in part through interaction with growth factor receptor signaling pathways. In the present study the effects of tamoxifen treatment on the expression of SRCs and human epidermal growth factor receptors (HERs) were examined in an animal model of ER positive breast cancer. Sprague-Dawley rats with DMBA-induced breast cancer were randomized to 14 days of oral tamoxifen 40 mg/kg bodyweight/day or vehicle only (controls). Tumors were measured throughout the study period. Blood samples and tumor tissue were collected at sacrifice and tamoxifen and its main metabolites were quantified using LC-MS/MS. The gene expression in tumor of SRC-1, SRC-2/transcription intermediary factor-2 (TIF-2), SRC-3/amplified in breast cancer 1 (AIB1), ER, HER-1, -2, -3 and HER-4, as well as the transcription factor Ets-2, was measured by real-time RT-PCR. Protein levels were further assessed by Western blotting. Tamoxifen and its main metabolites were detected at high concentrations in serum and accumulated in tumor tissue in up to tenfolds the concentration in serum. Mean tumor volume/rat decreased in the tamoxifen treated group, but continued to increase in controls. The mRNA expression levels of SRC-1 (P = 0.035), SRC-2/TIF-2 (P = 0.002), HER-2 (P = 0.035) and HER-3 (P = 0.006) were significantly higher in tamoxifen treated tumors compared to controls, and the results were confirmed at the protein level using Western blotting. SRC-3/AIB1 protein was also higher in tamoxifen treated tumors. SRC-1 and SRC-2/TIF-2 mRNA levels were positively correlated with each other and with HER-2 (P ≤ 0.001), and the HER-2 mRNA expression correlated with the levels of the other three HER family members (P < 0.05). Furthermore, SRC-3/AIB1 and HER-4 were positively correlated with each other and Ets-2 (P < 0.001). The expression of SRCs and HER-2 and -3 is stimulated by tamoxifen treatment in DMBA-induced breast cancer. Stimulation and positive correlation of coactivators and HERs may represent an early response to endocrine treatment. The role of SRCs and HER-2 and -3 should be further studied in order to evaluate their effects on response to long-term tamoxifen treatment.

Steroid receptor coactivators, HER-2 and HER-3 expression is stimulated by tamoxifen treatment in DMBA-induced breast cancer

PubMed Central

2012-01-01

Background Steroid receptor coactivators (SRCs) may modulate estrogen receptor (ER) activity and the response to endocrine treatment in breast cancer, in part through interaction with growth factor receptor signaling pathways. In the present study the effects of tamoxifen treatment on the expression of SRCs and human epidermal growth factor receptors (HERs) were examined in an animal model of ER positive breast cancer. Methods Sprague-Dawley rats with DMBA-induced breast cancer were randomized to 14 days of oral tamoxifen 40 mg/kg bodyweight/day or vehicle only (controls). Tumors were measured throughout the study period. Blood samples and tumor tissue were collected at sacrifice and tamoxifen and its main metabolites were quantified using LC-MS/MS. The gene expression in tumor of SRC-1, SRC-2/transcription intermediary factor-2 (TIF-2), SRC-3/amplified in breast cancer 1 (AIB1), ER, HER-1, -2, -3 and HER-4, as well as the transcription factor Ets-2, was measured by real-time RT-PCR. Protein levels were further assessed by Western blotting. Results Tamoxifen and its main metabolites were detected at high concentrations in serum and accumulated in tumor tissue in up to tenfolds the concentration in serum. Mean tumor volume/rat decreased in the tamoxifen treated group, but continued to increase in controls. The mRNA expression levels of SRC-1 (P = 0.035), SRC-2/TIF-2 (P = 0.002), HER-2 (P = 0.035) and HER-3 (P = 0.006) were significantly higher in tamoxifen treated tumors compared to controls, and the results were confirmed at the protein level using Western blotting. SRC-3/AIB1 protein was also higher in tamoxifen treated tumors. SRC-1 and SRC-2/TIF-2 mRNA levels were positively correlated with each other and with HER-2 (P ≤ 0.001), and the HER-2 mRNA expression correlated with the levels of the other three HER family members (P < 0.05). Furthermore, SRC-3/AIB1 and HER-4 were positively correlated with each other and Ets-2 (P < 0.001). Conclusions The expression of SRCs and HER-2 and -3 is stimulated by tamoxifen treatment in DMBA-induced breast cancer. Stimulation and positive correlation of coactivators and HERs may represent an early response to endocrine treatment. The role of SRCs and HER-2 and -3 should be further studied in order to evaluate their effects on response to long-term tamoxifen treatment. PMID:22703232

Neuronal expression of fibroblast growth factor receptors in zebrafish.

PubMed

Rohs, Patricia; Ebert, Alicia M; Zuba, Ania; McFarlane, Sarah

2013-12-01

Fibroblast growth factor (FGF) signaling is important for a host of developmental processes such as proliferation, differentiation, tissue patterning, and morphogenesis. In vertebrates, FGFs signal through a family of four fibroblast growth factor receptors (FGFR 1-4), one of which is duplicated in zebrafish (FGFR1). Here we report the mRNA expression of the five known zebrafish fibroblast growth factor receptors at five developmental time points (24, 36, 48, 60, and 72h postfertilization), focusing on expression within the central nervous system. We show that the receptors have distinct and dynamic expression in the developing zebrafish brain, eye, inner ear, lateral line, and pharynx. In many cases, the expression patterns are similar to those of homologous FGFRs in mouse, chicken, amphibians, and other teleosts. Copyright © 2013 Elsevier B.V. All rights reserved.

Thyroid hormone and COUP-TF1 regulate kallikrein-binding protein (KBP) gene expression.

PubMed

Liu, Yan-Yun; Nakatani, Teruyo; Kogai, Takahiko; Mody, Kaizeen; Brent, Gregory A

2011-03-01

Kallikrein-binding protein (KBP) is a component of the kallikrein-kinin system that mediates vasodilation and inhibits tumor growth by antagonizing vascular endothelial growth factor-mediated angiogenesis. We demonstrate that KBP gene expression is repressed by T(3) and modulated by the orphan nuclear receptor, chicken ovalbumin upstream promoter transcription factor 1 (COUP-TF1). In hypothyroid mice, KBP mRNA expression in the testis was increased 2.1-fold compared with euthyroid mice. We have identified two negative thyroid hormone response elements (nTREs) in the mouse KBP gene, nTRE1 located in the 5' flanking region (-53 to -29) and nTRE2, located in the first intron (104-132). We used functional assays, cofactor knockdown, and chromatin immunoprecipitation assays to characterize nTRE1 and nTRE2 in hepatic (HepG2) and testes (GC-1spg) cell lines. Reporter expression directed by both elements was enhanced with addition of thyroid hormone receptor and repressed with the addition of T(3). COUP-TF1 enhanced basal expression of both elements but blunted unliganded thyroid hormone receptor enhancement and T(3) repression of nTRE1 but not nTRE2. Both nTREs bound nuclear corepressor and binding increased in response to T(3). Nuclear corepressor knockdown resulted in loss of T(3) repression of both nTRE1 and nTRE2. COUP-TF1, which usually represses T(3) induction of positive thyroid hormone response elements, reverses T(3) repression mediated by nTRE1 in the mouse KBP gene. Endogenous KBP expression is repressed by T(3) and two functional nTREs, both of which are required, have been characterized in the KBP gene. COUP-TF1 may be an important factor to modulate expression of genes that are repressed by T(3).

Thyroid Hormone and COUP-TF1 Regulate Kallikrein-Binding Protein (KBP) Gene Expression

PubMed Central

Liu, Yan-Yun; Nakatani, Teruyo; Kogai, Takahiko; Mody, Kaizeen

2011-01-01

Kallikrein-binding protein (KBP) is a component of the kallikrein-kinin system that mediates vasodilation and inhibits tumor growth by antagonizing vascular endothelial growth factor-mediated angiogenesis. We demonstrate that KBP gene expression is repressed by T3 and modulated by the orphan nuclear receptor, chicken ovalbumin upstream promoter transcription factor 1 (COUP-TF1). In hypothyroid mice, KBP mRNA expression in the testis was increased 2.1-fold compared with euthyroid mice. We have identified two negative thyroid hormone response elements (nTREs) in the mouse KBP gene, nTRE1 located in the 5′ flanking region (−53 to −29) and nTRE2, located in the first intron (104–132). We used functional assays, cofactor knockdown, and chromatin immunoprecipitation assays to characterize nTRE1 and nTRE2 in hepatic (HepG2) and testes (GC-1spg) cell lines. Reporter expression directed by both elements was enhanced with addition of thyroid hormone receptor and repressed with the addition of T3. COUP-TF1 enhanced basal expression of both elements but blunted unliganded thyroid hormone receptor enhancement and T3 repression of nTRE1 but not nTRE2. Both nTREs bound nuclear corepressor and binding increased in response to T3. Nuclear corepressor knockdown resulted in loss of T3 repression of both nTRE1 and nTRE2. COUP-TF1, which usually represses T3 induction of positive thyroid hormone response elements, reverses T3 repression mediated by nTRE1 in the mouse KBP gene. Endogenous KBP expression is repressed by T3 and two functional nTREs, both of which are required, have been characterized in the KBP gene. COUP-TF1 may be an important factor to modulate expression of genes that are repressed by T3. PMID:21266512

Expression profiling of G-protein-coupled receptors in human urothelium and related cell lines.

PubMed

Ochodnický, Peter; Humphreys, Sian; Eccles, Rachel; Poljakovic, Mirjana; Wiklund, Peter; Michel, Martin C

2012-09-01

What's known on the subject? and What does the study add? Urothelium emerged as a crucial integrator of sensory inputs and outputs in the bladder wall, and urothelial G-protein-coupled receptors (GPCRs) may represent plausible targets for treatment of various bladder pathologies. Urothelial cell lines provide a useful tool to study urothelial receptor function, but their validity as models for native human urothelium remains unclear. We characterize the mRNA expression of genes coding for GPCRs in human freshly isolated urothelium and compare the expression pattern with those in human urothelial cell lines. To characterize the mRNA expression pattern of genes coding for G-protein-coupled receptors (GPCRs) in human freshly isolated urothelium. To compare GPCR expression in human urothelium-derived cell lines to explore the suitability of these cell lines as model systems to study urothelial function. Native human urothelium (commercially sourced) and human urothelium-derived non-cancer (UROtsa and TERT-NHUC) and cancer (J82) cell lines were used. For mRNA expression profiling we used custom-designed real-time polymerase chain reaction array for 40 receptors and several related genes. Native urothelium expressed a wide variety of GPCRs, including α(1A), α(1D) and all subtypes of α(2) and β adrenoceptors. In addition, M(2) and M(3) cholinergic muscarinic receptors, angiotensin II AT(1) receptor, serotonin 5-HT(2A) receptor and all subtypes of bradykinin, endothelin, cannabinoid, tachykinin and sphingosine-1-phosphate receptors were detected. Nerve growth factor and both its low- and high-affinity receptors were also expressed in urothelium. In all cell lines expression of most GPCRs was markedly downregulated, with few exceptions. In UROtsa cells, but much less in other cell lines, the expression of β(2) adrenoceptors, M(3) muscarinic receptors, B(1) and B(2) bradykinin receptors, ET(B) endothelin receptors and several subtypes of sphingosine-1-phosphate receptors was largely retained. Human urothelium expresses a wide range of receptors which enables sensing and integration of various extracellular signals. Human urothelium-derived cell lines, especially UROtsa cells, show comparable mRNA expression to native tissue for several physiologically relevant GPCRs, but lose expression of many other receptors. The use of cell lines as model systems of human urothelium requires careful validation of suitability for the genes of interest. © 2012 BJU INTERNATIONAL.

Tumor necrosis factor-α inhibits angiotensin II receptor type 1 expression in dorsal root ganglion neurons via β-catenin signaling.

PubMed

Yang, Y; Wu, H; Yan, J-Q; Song, Z-B; Guo, Q-L

2013-09-17

Both tumor necrosis factor (TNF)-α and the angiotensin (Ang) II/angiotensin II receptor type 1 (AT1) axis play important roles in neuropathic pain and nociception. In the present study, we explored the interaction between the two systems by examining the mutual effects between TNF-α and the Ang II/AT1 receptor axis in dorsal root ganglion (DRG) neurons. Rat DRG neurons were treated with TNF-α in different concentrations for different lengths of time in the presence or absence of transcription inhibitor actinomycin D, TNF receptor 1 (TNFR1) inhibitor SPD304, β-catenin signaling inhibitor CCT031374, or different kinase inhibitors. TNF-α decreased the AT1 receptor mRNA level as well as the AT1a receptor promoter activity in a dose-dependent manner within 30 h, which led to dose-dependent inhibition of Ang II-binding AT1 receptor level on the cell membrane. Actinomycin D (1 mg/ml), SPD304 (50 μM), p38 mitogen-activated protein kinase (MAPK) inhibitor PD169316 (25 μM), and CCT031374 (50 μM) completely abolished the inhibitory effect of TNF-α on AT1 receptor expression. TNF-α dose-dependently increased soluble β-catenin and phosphorylated GSK-3β levels, which was blocked by SPD304 and PD169316. In DRG neurons treated with AT2 receptor agonist CGP421140, or Ang II with or without AT1 receptor antagonist losartan or AT2 receptor antagonist PD123319 for 30 h, we found that Ang II and Ang II+PD123319 significantly decreased TNF-α expression, whereas CPG421140 and Ang II+losartan increased TNF-α expression. In conclusion, we demonstrate that TNF-α inhibits AT1 receptor expression at the transcription level via TNFR1 in rat DRG neurons by increasing the soluble β-catenin level through the p38 MAPK/GSK-3β pathway. In addition, Ang II appears to inhibit and induce TNF-α expression via the AT1 receptor and the AT2 receptor in DRG neurons, respectively. This is the first evidence of crosstalk between TNF-α and the Ang II/AT receptor axis in DRG neurons. Copyright © 2013 IBRO. Published by Elsevier Ltd. All rights reserved.

The human luteinizing hormone receptor gene promoter: activation by Sp1 and Sp3 and inhibitory regulation.

PubMed

Geng, Y; Tsai-Morris, C H; Zhang, Y; Dufau, M L

1999-09-24

To understand the transcriptional mechanism(s) of human LH receptor (LHR) gene expression, we have identified the dominant functional cis-elements that regulate the activity of the promoter domain (-1 to -176 bp from ATG). Mutagenesis demonstrated that the promoter activity was dependent on two Sp1 domains (-79 bp, -120 bp) in a transformed normal placental cell (PLC) and the choriocarcinoma JAR cell. Both elements interacted with endogenous Sp1 and Sp3 factors but not with Sp2 or Sp4. In Drosophila SL2 cells, the promoter was activated by either Sp1 or Sp3. An ERE half-site (EREhs) at -174 bp was inhibitory (by 100%), but was unresponsive to estradiol and did not bind the estrogen receptor or orphan receptors ERR1 and SF-1. The 5' upstream sequence (-177 to -2056 bp) inhibited promoter activity in PLC by 60%, but only minimally in JAR cells. Activation of the human LHR promoter through Sp1/3 factors is negatively regulated through EREhs and upstream sequences to exert control of gene expression. Copyright 1999 Academic Press.

Sirtuin 3 (SIRT3) maintains bone homeostasis by regulating AMPK-PGC-1β axis in mice

PubMed Central

Huh, Jeong-Eun; Shin, Ji Hye; Jang, Eun Sun; Park, So Jeong; Park, Doo Ri; Ko, Ryeojin; Seo, Dong-Hyun; Kim, Han-Sung; Lee, Seoung Hoon; Choi, Yongwon; Kim, Hyun Seok; Lee, Soo Young

2016-01-01

The mitochondrial sirtuin 3 (SIRT3) is involved in suppressing the onset of multiple pathologies, including cardiovascular disease, fatty liver, age-related hearing loss, and breast cancer. But a physiological role of SIRT3 in bone metabolism is not known. Here we show that SIRT3 is a key regulatory molecule to maintain bone homeostasis. Mice deficient in SIRT3 exhibited severe osteopenia owing to increased numbers of osteoclasts. Osteoclast precursors from Sirt3−/− mice underwent increased osteoclastogenesis in response to receptor activator of nuclear factor-κB ligand (RANKL), an essential cytokine for osteoclast differentiation. SIRT3 expression from RANKL induction depended on the transcription coactivator PGC-1β (peroxisome proliferator-activated receptor-γ co-activator-1β) and the nuclear receptor ERRα (estrogen receptor-related receptor α), and that SIRT3 inhibited the differentiation by interfering with the RANKL-induced expression of PGC-1β. Thus an auto-regulatory feedback mechanism operates to induce its own inhibitor SIRT3 by PGC-1β. Moreover, Sirt3−/− osteoclast precursors reduced AMP-activated protein kinase (AMPK) phosphorylation through down-regulating the expression of AMPK. Our results suggest that a mitochondrial SIRT3 is an intrinsic inhibitor for RANKL-mediated osteoclastogenesis. PMID:26928655

Human Herpesvirus-8-Transformed Endothelial Cells Have Functionally Activated Vascular Endothelial Growth Factor/Vascular Endothelial Growth Factor Receptor

PubMed Central

Masood, Rizwan; Cesarman, Ethel; Smith, D. Lynne; Gill, Parkash S.; Flore, Ornella

2002-01-01

Kaposi’s sarcoma is a vascular tumor commonly associated with human immunodeficiency virus (HIV)-1 and human herpesvirus (HHV-8) also known as Kaposi’s sarcoma-associated herpesvirus. The principal features of this tumor are abnormal proliferation of vascular structures lined with spindle-shaped endothelial cells. HHV-8 may transform a subpopulation of endothelial cells in vitro via viral and cellular gene expression. We hypothesized that among the cellular genes, vascular endothelial growth factors (VEGFs) and their cognate receptors may be involved in viral-mediated transformation. We have shown that HHV-8-transformed endothelial cells (EC-HHV-8) express higher levels of VEGF, VEGF-C, VEGF-D, and PlGF in addition to VEGF receptors-1, -2, and -3. Furthermore, antibodies to VEGF receptor-2 inhibited cell proliferation and viability. Similarly, inhibition of VEGF gene expression with antisense oligonucleotides inhibited EC-HHV-8 cell proliferation/viability. The growth and viability of primary endothelial cells and a fibroblast cell line however were unaffected by either the VEGF receptor-2 antibody or the VEGF antisense oligodeoxynucleotides. VEGF and VEGF receptors are thus induced in EC-HHV-8 and participate in the transformation. Inhibitors of VEGF may thus modulate the disease process during development and progression. PMID:11786394

Molecular and functional characterization of pigeon (Columba livia) tumor necrosis factor receptor-associated factor 3.

PubMed

Zhou, Yingying; Kang, Xilong; Xiong, Dan; Zhu, Shanshan; Zheng, Huijuan; Xu, Ying; Guo, Yaxin; Pan, Zhiming; Jiao, Xinan

2017-04-01

Tumor necrosis factor receptor-associated factor 3 (TRAF3) plays a key antiviral role by promoting type I interferon production. We cloned the pigeon TRAF3 gene (PiTRAF3) according to its predicted mRNA sequence to investigate its function. The 1704-bp full-length open reading frame encodes a 567-amino acid protein. One Ring finger, two TRAF-type Zinc fingers, one Coiled coil, and one MATH domain were inferred. RT-PCR showed that PiTRAF3 was expressed in all tissues, with relatively weak expression in the heart and liver. In HEK293T cells, over-expression of wild-type, △Ring, △Zinc finger, and △Coiled coil PiTRAF3, but not a △MATH form, significantly increased IFN-β promoter activity. Zinc finger and Coiled coil domains were essential for NF-κB activation. In chicken HD11 cells, PiTRAF3 increased IFN-β promoter activity and four domains were all contributing. R848 stimulation of pigeon peripheral blood mononuclear cells and splenocytes significantly increased expression of PiTRAF3 and the inflammatory cytokine genes CCL5, IL-8, and IL-10. These data demonstrate TRAF3's innate immune function and improve understanding of its involvement in poultry antiviral defense. Copyright © 2016 Elsevier Ltd. All rights reserved.

Gene expression analysis of growth factor receptors in human chondrocytes in monolayer and 3D pellet cultures

PubMed Central

Witt, Anika; Salamon, Achim; Boy, Diana; Hansmann, Doris; Büttner, Andreas; Wree, Andreas; Bader, Rainer; Jonitz-Heincke, Anika

2017-01-01

The main goal of cartilage repair is to create functional tissue by enhancing the in vitro conditions to more physiological in vivo conditions. Chondrogenic growth factors play an important role in influencing cartilage homeostasis. Insulin-like growth factor (IGF)-1 and transforming growth factor (TGF)-β1 affect the expression of collagen type II (Col2) and glycosaminoglycans (GAGs) and, therefore, the targeted use of growth factors could make chondrogenic redifferentiation more efficient. In the present study, human chondrocytes were postmortally isolated from healthy articular cartilage and cultivated as monolayer or 3D pellet cultures either under normoxia or hypoxia and stimulated with IGF-1 and/or TGF-β1 to compare the impact of the different growth factors. The mRNA levels of the specific receptors (IGF1R, TGFBR1, TGFBR2) were analyzed at different time points. Moreover, gene expression rates of collagen type 1 and 2 in pellet cultures were observed over a period of 5 weeks. Additionally, hyaline-like Col2 protein and sulphated GAG (sGAG) levels were quantified. Stimulation with IGF-1 resulted in an enhanced expression of IGF1R and TGFBR2 whereas TGF-β1 stimulated TGFBR1 in the monolayer and pellet cultures. In monolayer, the differences reached levels of significance. This effect was more pronounced under hypoxic culture conditions. In pellet cultures, increased amounts of Col2 protein and sGAGs after incubation with TGF-β1 and/or IGF-1 were validated. In summary, constructing a gene expression profile regarding mRNA levels of specific growth factor receptors in monolayer cultures could be helpful for a targeted application of growth factors in cartilage tissue engineering. PMID:28534942

Expression of fibroblast growth factor receptors during development and regression of the bovine corpus luteum.

PubMed

Guerra, D M; Giometti, I C; Price, C A; Andrade, P B; Castilho, A C; Machado, M F; Ripamonte, P; Papa, P C; Buratini, J

2008-01-01

There is evidence that fibroblast growth factors (FGFs) are involved in the regulation of growth and regression of the corpus luteum (CL). However, the expression pattern of most FGF receptors (FGFRs) during CL lifespan is still unknown. The objective of the present study was to determine the pattern of expression of 'B' and 'C' splice variants of FGFRs in the bovine CL. Bovine CL were collected from an abattoir and classed as corpora hemorrhagica (Stage I), developing (Stage II), developed (Stage III) or regressed (Stage IV) CL. Expression of FGFR mRNA was measured by semiquantitative reverse transcription-polymerase chain reaction and FGFR protein was localised by immunohistochemistry. Expression of mRNA encoding the 'B' and 'C' spliced forms of FGFR1 and FGFR2 was readily detectable in the bovine CL and was accompanied by protein localisation. FGFR1C and FGFR2C mRNA expression did not vary throughout CL lifespan, whereas FGFR1B was upregulated in the developed (Stage III) CL. FGFR3B, FGFR3C and FGFR4 expression was inconsistent in the bovine CL. The present data indicate that FGFR1 and FGFR2 splice variants are the main receptors for FGF action in the bovine CL.

Thalidomide inhibits lipopolysaccharide-induced tumor necrosis factor-alpha production via down-regulation of MyD88 expression.

PubMed

Noman, Abu Shadat M; Koide, Naoki; Hassan, Ferdaus; I-E-Khuda, Imtiaz; Dagvadorj, Jargalsaikhan; Tumurkhuu, Gantsetseg; Islam, Shamima; Naiki, Yoshikazu; Yoshida, Tomoaki; Yokochi, Takashi

2009-02-01

The effect of thalidomide on lipopolysaccharide (LPS)-induced tumor necrosis factor (TNF)-alpha production was studied by using RAW 264.7 murine macrophage-like cells. Thalidomide significantly inhibited LPS-induced TNF-alpha production. Thalidomide prevented the activation of nuclear factor (NF)-KB by down-regulating phosphorylation of inhibitory KB factor (IKB), and IKB kinase (IKK)-alpha and IKK-beta Moreover, thalidomide inhibited LPS-induced phosphorylation of AKT, p38 and stress-activated protein kinase (SAPK)/JNK. The expression of myeloid differentiation factor 88 (MyD88) protein and mRNA was markedly reduced in thalidomide-treated RAW 264.7 cells but there was no significant alteration in the expression of interleukin-1 receptor-associated kinase (IRAK) 1 and TNF receptor-associated factor (TRAF) 6 in the cells. Thalidomide did not affect the cell surface expression of Toll-like receptor (TLR) 4 and CD14, suggesting the impairment of intracellular LPS signalling in thalidomide-treated RAW 264.7 cells. Thalidomide significantly inhibited the TNF-alpha production in response to palmitoyl-Cys(RS)-2,3-di(palmitoyloxy) propyl)-Ala-Gly-OH (Pam(3)Cys) as a MyD88-dependent TLR2 ligand. Therefore, it is suggested that thalidomide might impair LPS signalling via down-regulation of MyD88 protein and mRNA and inhibit LPS-induced TNF-alpha production. The putative mechanism of thalidomide-induced MyD88 down-regulation is discussed.

Granulocyte macrophage-colony stimulating factor and interleukin-3 increase expression of type II tumour necrosis factor receptor, increasing susceptibility to tumour necrosis factor-induced apoptosis. Control of leukaemia cell life/death switching.

PubMed

Rae, C; MacEwan, D J

2004-12-01

Tumour necrosis factor (TNF) induces apoptosis in a range of cell types via its two receptors, TNFR1 and TNFR2. Here, we demonstrate that proliferation and TNFR2 expression was increased in human leukaemic TF-1 cells by granulocyte macrophage-colony stimulating factor (GM-CSF) and interleukin-3 (IL-3), with TNFR1 expression unaffected. Consequently, they switch from a proliferative to a TNF-induced apoptotic phenotype. Raised TNFR2 expression and susceptibility to TNF-induced apoptosis was not a general effect of proliferation as IL-1beta and IFN-gamma both proliferated TF-1 cells with no effect on TNFR expression or apoptosis. Although raised TNFR2 expression correlated with the apoptotic phenotype, stimulation of apoptosis in GM-CSF-pretreated cells was mediated by TNFR1, with stimulation of TNFR2 alone insufficient to initiate cell death. However, TNFR2 did play a role in apoptotic and proliferative responses as they were blocked by the presence of an antagonistic TNFR2 antibody. Additionally, coincubation with cycloheximide blocked the mitotic effects of GM-CSF or IL-3, allowing only the apoptotic responses of TNF to persist. TNF life/death was also observed in K562, but not MOLT-4 and HL-60 human leukaemic cell types. These findings show a cooperative role of TNFR2 in the TNF life/death switching phenomenon.

Inhibition of insulin-like growth factor receptor-1 reduces necroptosis-related markers and attenuates LPS-induced lung injury in mice.

PubMed

Lee, Su Hwan; Shin, Ju Hye; Song, Joo Han; Leem, Ah Young; Park, Moo Suk; Kim, Young Sam; Chang, Joon; Chung, Kyung Soo

2018-04-15

Insulin-like growth factor-1 (IGF-1) levels are known to increase in the bronchoalveolar lavage fluid (BALF) of patients with acute respiratory distress syndrome. Herein, we investigated the role of IGF-1 in lipopolysaccharide (LPS)-induced lung injury. In LPS-treated cells, expressions of receptor-interacting protein 3 (RIP3) and phosphorylated mixed lineage kinase domain-like protein (MLKL) were decreased in IGF-1 receptor small interfering RNA (siRNA)-treated cells compared to control cells. The levels of pro-inflammatory cytokines including interleukin (IL)-1β, IL-6, IL-10, tumour necrosis factor-α, and macrophage inflammatory protein 2/C-X-C motif chemokine ligand 2 in the supernatant were significantly reduced in IGF-1 receptor siRNA-treated cells compared to control cells. In LPS-induced murine lung injury model, total cell counts, polymorphonuclear leukocytes counts, and pro-inflammatory cytokine levels in the BALF were significantly lower and histologically detected lung injury was less common in the group treated with IGF-1 receptor monoclonal antibody compared to the non-treated group. On western blotting, RIP3 and phosphorylated MLKL expressions were relatively decreased in the IGF-1 receptor monoclonal antibody group compared to the non-treated group. IGF-1 may be associated with RIP3-mediated necroptosis in vitro, while blocking of the IGF-1 pathway may reduce LPS-induced lung injuries in vivo. Copyright © 2018 Elsevier Inc. All rights reserved.

The effect of melatonin from slow-release implants on basic and TLR-4-mediated gene expression of inflammatory cytokines and their receptors in the choroid plexus in ewes.

PubMed

Kowalewska, M; Herman, A P; Szczepkowska, A; Skipor, J

2017-08-01

The present study concerns the effect of melatonin from slow-release implants on the expression of genes coding interleukin-1β (Il1B), inerleukin-6 (Il6), tumour necrosis factor α (Tnf) and their receptors: IL-1 receptor type I (Il1r1) and type II (Il1r2), IL-6 receptor (Il6r) and signal transducer (Il6st), TNFα receptor type I (Tnfrsf1a) and II (Tnfrsf1b) and retinoid-related orphan receptor α (RorA) and Rev.-erbα in the ovine choroid plexus (CP) under basal and lipopolysaccharide (LPS)-challenged conditions. Studies were performed on four groups: 1) sham-implanted and placebo-treated, 2) melatonin-implanted (Melovine, 18mg) and placebo-treated, 3) sham-implanted and LPS-treated (400ng/kg of body weight) and 4) melatonin-implanted and LPS-treated. Under basal conditions, we observed weak expression of Tnf, low expression of Il1B, Il6 and Il1r2 and intermediate expression of other cytokines receptors. LPS treatment induced (P≤0.05) expression in all cytokines and their receptors, except Il6r 3h after the administration. Melatonin attenuated (P≤0.05) LPS-induced up-regulation of Il6 but had no effect on other cytokines and their receptors and up-regulated (P≤0.05) Rev.-erbα expression under basal conditions. This indicates that melatonin from slow-release implants suppresses TLR4-mediated Il6 expression in the ovine CP via a mechanism likely involving clock genes. Copyright © 2017 Elsevier Ltd. All rights reserved.

Aberrant expression and function of death receptor-3 and death decoy receptor-3 in human cancer.

PubMed

Ge, Zhicheng; Sanders, Andrew J; Ye, Lin; Jiang, Wen G

2011-03-01

Death receptor-3 (DR3) and death decoy receptor-3 (DcR3) are both members of the tumour necrosis factor receptor (TNFR) superfamily. The TNFR superfamily contains eight death domain-containing receptors, including TNFR1 (also called DR1), Fas (also called DR2), DR3, DR4, DR5, DR6, NGFR and EDAR. Upon the binding of these receptors with their corresponding ligands, the death domain recruits various proteins that mediate both the death and proliferation of cells. Receptor function is negatively regulated by decoy receptors (DcR1, DcR2, DcR3 and OPG). DR3/DcR3 are a pair of positive and negative players with which vascular endothelial growth inhibitor (VEGI) interacts. VEGI has been suggested to be a potential tumour suppressor. The inhibitory effects of VEGI on cancer are manifested in three main areas: a direct effect on cancer cells, an anti-angiogenic effect on endothelial cells, and the stimulation of dendritic cell maturation. A recent study indicated that DR3 may be a new receptor for E-selectin, which has been reported to be associated with cancer metastasis. DcR3 is a soluble receptor, highly expressed in various tumours, which lacks an apparent transmembrane segment, prevents cytokine response through ligand binding and neutralization, and is an inhibitor of apoptosis. DcR3 serves as a decoy receptor for FasL, LIGHT and VEGI. The cytokine LIGHT activates various anti-tumour functions and is expected to be a promising candidate for cancer therapy. Certain tumours may escape FasL-dependent immune-cytotoxic attack by expressing DcR3, which blocks FasL function. DR3/DcR3 play profound roles in regulating cell death and proliferation in cancer. The present review briefly discusses DR3/DcR3 and attempts to elucidate the role of these negative and positive players in cancer.

Steroid receptor coactivator-3 regulates glucose metabolism in bladder cancer cells through coactivation of hypoxia inducible factor 1α.

PubMed

Zhao, Wei; Chang, Cunjie; Cui, Yangyan; Zhao, Xiaozhi; Yang, Jun; Shen, Lan; Zhou, Ji; Hou, Zhibo; Zhang, Zhen; Ye, Changxiao; Hasenmayer, Donald; Perkins, Robert; Huang, Xiaojing; Yao, Xin; Yu, Like; Huang, Ruimin; Zhang, Dianzheng; Guo, Hongqian; Yan, Jun

2014-04-18

Cancer cell proliferation is a metabolically demanding process, requiring high glycolysis, which is known as "Warburg effect," to support anabolic growth. Steroid receptor coactivator-3 (SRC-3), a steroid receptor coactivator, is overexpressed and/or amplified in multiple cancer types, including non-steroid targeted cancers, such as urinary bladder cancer (UBC). However, whether SRC-3 regulates the metabolic reprogramming for cancer cell growth is unknown. Here, we reported that overexpression of SRC-3 accelerated UBC cell growth, accompanied by the increased expression of genes involved in glycolysis. Knockdown of SRC-3 reduced the UBC cell glycolytic rate under hypoxia, decreased tumor growth in nude mice, with reduction of proliferating cell nuclear antigen and lactate dehydrogenase expression levels. We further revealed that SRC-3 could interact with hypoxia inducible factor 1α (HIF1α), which is a key transcription factor required for glycolysis, and coactivate its transcriptional activity. SRC-3 was recruited to the promoters of HIF1α-target genes, such as glut1 and pgk1. The positive correlation of expression levels between SRC-3 and Glut1 proteins was demonstrated in human UBC patient samples. Inhibition of glycolysis through targeting HK2 or LDHA decelerated SRC-3 overexpression-induced cell growth. In summary, overexpression of SRC-3 promoted glycolysis in bladder cancer cells through HIF1α to facilitate tumorigenesis, which may be an intriguing drug target for bladder cancer therapy.

Arctigenin induced gallbladder cancer senescence through modulating epidermal growth factor receptor pathway.

PubMed

Zhang, Mingdi; Cai, Shizhong; Zuo, Bin; Gong, Wei; Tang, Zhaohui; Zhou, Di; Weng, Mingzhe; Qin, Yiyu; Wang, Shouhua; Liu, Jun; Ma, Fei; Quan, Zhiwei

2017-05-01

Gallbladder cancer has poor prognosis and limited therapeutic options. Arctigenin, a representative dibenzylbutyrolactone lignan, occurs in a variety of plants. However, the molecular mechanisms involved in the antitumor effect of arctigenin on gallbladder cancer have not been fully elucidated. The expression levels of epidermal growth factor receptor were examined in 100 matched pairs of gallbladder cancer tissues. A positive correlation between high epidermal growth factor receptor expression levels and poor prognosis was observed in gallbladder cancer tissues. Pharmacological inhibition or inhibition via RNA interference of epidermal growth factor receptor induced cellular senescence in gallbladder cancer cells. The antitumor effect of arctigenin on gallbladder cancer cells was primarily achieved by inducing cellular senescence. In gallbladder cancer cells treated with arctigenin, the expression level of epidermal growth factor receptor significantly decreased. The analysis of the activity of the kinases downstream of epidermal growth factor receptor revealed that the RAF-MEK-ERK signaling pathway was significantly inhibited. Furthermore, the cellular senescence induced by arctigenin could be reverted by pcDNA-epidermal growth factor receptor. Arctigenin also potently inhibited the growth of tumor xenografts, which was accompanied by the downregulation of epidermal growth factor receptor and induction of senescence. This study demonstrates arctigenin could induce cellular senescence in gallbladder cancer through the modulation of epidermal growth factor receptor pathway. These data identify epidermal growth factor receptor as a key regulator in arctigenin-induced gallbladder cancer senescence.

PcToll3 was involved in anti-Vibrio response by regulating the expression of antimicrobial peptides in red swamp crayfish, Procambarus clarkii.

PubMed

Lan, Jiang-Feng; Wei, Shun; Wang, Yu-Qing; Dai, Yun-Jia; Tu, Jia-Gang; Zhao, Li-Juan; Li, Xin-Cang; Qin, Qi-Wei; Chen, Nan; Lin, Li

2016-10-01

Tolls and Toll-like receptors (TLRs) play an important role in host immune defenses by regulating the expression of antimicrobial peptides (AMPs) and cytokines, but the functional differences of crustacean Tolls from Drosophila Tolls or Mammal TLRs are largely unknown. A novel Toll receptor, named PcToll3, was identified from red swamp crayfish, Procambarus clarkii. It was widely expressed in all detected tissues, and its transcript in hemocytes was up-regulated at 12 h after Vibrio parahemolyticus (Vibrio) injection or at 24 h post white spot syndrome virus (WSSV) challenge. After knockdown of PcToll3, the activity of bacterial clearance was inhibited, and the expression levels of AMPs including Crustin1 (Cru1), Anti-lippopolysaccharide factor 1 (ALF1), and Lysozymes1 (Lys1), which could be up-regulated by Vibrio, were all affected. Meanwhile, PcToll3 silencing influenced the expression of myeloid differentiation factor 88 (PcMyd88), tumor necrosis factor-associated factor 6 (PcTRAF6), and PcDorsal, which were the counterparts of Drosophila Toll signaling pathway. Interestingly, PcToll3 silencing inhibited translocation of PcDorsal from cytoplasm to nucleus. Furthermore, the knockdown of PcDorsal also impaired the expression of AMPs after Vibrio challenge. Hence, we concluded that, besides participating in antiviral immunity, PcToll3 might also regulate the expression of Cru1 and Lys1 to participate in anti-Vibrio immune responses by promoting PcDorsal translocation into nucleus. Copyright © 2016 Elsevier Ltd. All rights reserved.

IL-6 Overexpression in ERG-Positive Prostate Cancer Is Mediated by Prostaglandin Receptor EP2.

PubMed

Merz, Constanze; von Mässenhausen, Anne; Queisser, Angela; Vogel, Wenzel; Andrén, Ove; Kirfel, Jutta; Duensing, Stefan; Perner, Sven; Nowak, Michael

2016-04-01

Prostate cancer is the most diagnosed cancer in men and multiple risk factors and genetic alterations have been described. The TMPRSS2-ERG fusion event and the overexpression of the transcription factor ERG are present in approximately 50% of all prostate cancer patients, however, the clinical outcome is still controversial. Prostate tumors produce various soluble factors, including the pleiotropic cytokine IL-6, regulating cellular processes such as proliferation and metastatic segregation. Here, we used prostatectomy samples in a tissue microarray format and analyzed the co-expression and the clinicopathologic data of ERG and IL-6 using immunohistochemical double staining and correlated the read-out with clinicopathologic data. Expression of ERG and IL-6 correlated strongly in prostate tissue samples. Forced expression of ERG in prostate tumor cell lines resulted in significantly increased secretion of IL-6, whereas the down-regulation of ERG decreased IL-6 secretion. By dissecting the underlying mechanism in prostate tumor cell lines we show the ERG-mediated up-regulation of the prostanoid receptors EP2 and EP3. The prostanoid receptor EP2 was overexpressed in human prostate cancer tissue. Furthermore, the proliferation rate and IL-6 secretion in DU145 cells was reduced after treatment with EP2-receptor antagonist. Collectively, our study shows that the expression of ERG in prostate cancer is linked to the expression of IL-6 mediated by the prostanoid receptor EP2. Copyright © 2016 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.

Feline leukemia virus infection requires a post-receptor binding envelope-dependent cellular component.

PubMed

Hussain, Naveen; Thickett, Kelly R; Na, Hong; Leung, Cherry; Tailor, Chetankumar S

2011-12-01

Gammaretrovirus receptors have been suggested to contain the necessary determinants to mediate virus binding and entry. Here, we show that murine NIH 3T3 and baby hamster kidney (BHK) cells overexpressing receptors for subgroup A, B, and C feline leukemia viruses (FeLVs) are weakly susceptible (10(1) to 10(2) CFU/ml) to FeLV pseudotype viruses containing murine leukemia virus (MLV) core (Gag-Pol) proteins, whereas FeLV receptor-expressing murine Mus dunni tail fibroblast (MDTF) cells are highly susceptible (10(4) to 10(6) CFU/ml). However, NIH 3T3 cells expressing the FeLV subgroup B receptor PiT1 are highly susceptible to gibbon ape leukemia virus pseudotype virus, which differs from the FeLV pseudotype viruses only in the envelope protein. FeLV resistance is not caused by a defect in envelope binding, low receptor expression levels, or N-linked glycosylation. Resistance is not alleviated by substitution of the MLV core in the FeLV pseudotype virus with FeLV core proteins. Interestingly, FeLV resistance is alleviated by fusion of receptor-expressing NIH 3T3 and BHK cells with MDTF or human TE671 cells, suggesting the absence of an additional cellular component in NIH 3T3 and BHK cells that is required for FeLV infection. The putative FeLV-specific cellular component is not a secreted factor, as MDTF conditioned medium does not alleviate the block to FeLV infection. Together, our findings suggest that FeLV infection requires an additional envelope-dependent cellular component that is absent in NIH 3T3 and BHK cells but that is present in MDTF and TE671 cells.

Expression of receptors for atrial natriuretic peptide on the murine bone marrow-derived stromal cells.

PubMed

Agui, T; Yamada, T; Legros, G; Nakajima, T; Clark, M; Peschel, C; Matsumoto, K

1992-05-01

Atrial natriuretic peptide (ANP) receptors were identified on both murine bone marrow-derived stromal cell lines A-3 and ALC and primary cultured cells using [125I]ANP binding assays and Northern blot analyses. The binding of [125I] ANP to the stromal cells was rapid, saturable, and of high affinity. The dissociation constants between ANP and its receptors on these cells showed no difference among cell types, while maximal binding capacity values were different among cell types. Competitive inhibition of [125I]ANP binding with C-atrial natriuretic factor, specific for ANP clearance receptor (ANPR-C), revealed that most of [125I]ANP-binding sites corresponded to ANPR-C. Northern blotting data corroborated that bone marrow-derived stromal cells expressed ANPR-C. However, in ALC cells, ANP biological receptors (either ANPR-A or ANPR-B), the mol wt of which is approximately 130K, were detected, and cGMP was accumulated after stimulation with ANP. On the other hand, in another stromal cell clone, A-3 cells, the expression of biological receptor was not detected in the affinity cross-linking and competitive inhibition experiments using [125I]ANP. However, A-3 cells accumulated cGMP by responding to ANPR-B-specific ligand, C-type natriuretic peptide. These results suggest that ALC cells equally express ANPR-A and ANPR-B, while A-3 cells express ANPR-B dominantly. Although the physiological roles of these receptors in the bone marrow is still not resolved, ANP is expected to play a role in the regulation of stromal cell functions in bone marrow.

Repetitive acute intermittent hypoxia increases expression of proteins associated with plasticity in the phrenic motor nucleus

PubMed Central

Satriotomo, Irawan; Dale, Erica A.; Dahlberg, Jenny M.; Mitchell, Gordon S.

2015-01-01

Acute intermittent hypoxia (AIH) initiates plasticity in respiratory motor control, including phrenic long term facilitation (pLTF). Since pLTF is enhanced by preconditioning with repetitive exposure to AIH (rAIH), we hypothesized that a rAIH protocol consisting of 3 AIH exposures per week for 10 weeks (3×wAIH; AIH: 10, 5-min episodes of 10.5% O2; 5-min normoxic intervals) would enhance expression of molecules that play key roles in pLTF within the phrenic motor nucleus. Immunohistochemical analyses revealed that 3×wAIH for 10 weeks increased serotonin terminal density in the C4 phrenic motor nucleus and serotonin 2A (5-HT2A) receptor expression in presumptive phrenic motor neurons. Immunoreactive brain derived neurotrophic factor (BDNF) and its high affinity receptor (TrkB) also increased following 3×wAIH. 3×wAIH also increased expression of another hypoxia-sensitive growth factor known to elicit phrenic motor facilitation, vascular endothelial growth factor (VEGF), and its receptor (VEGFR-2). Kinases “downstream” from TrkB and VEGFR-2 were up-regulated in or near presumptive phrenic motor neurons, including phosphorylated extracellular-signal regulated kinase (p-ERK) and protein kinase B (p-AKT). Thus, 3×wAIH up-regulates neurochemicals known to be associated with phrenic motor plasticity. Since 3×wAIH upregulates pro-plasticity molecules without evidence for CNS pathology, it may be a useful therapeutic tool in treating disorders that cause respiratory insufficiency, such as spinal injury or motor neuron disease. PMID:22704858

Basal expression of insulin-like growth factor 1 receptor determines intrinsic resistance of cancer cells to a phosphatidylinositol 3-kinase inhibitor ZSTK474

PubMed Central

Isoyama, Sho; Kajiwara, Gensei; Tamaki, Naomi; Okamura, Mutsumi; Yoshimi, Hisashi; Nakamura, Naoki; Kawamura, Kento; Nishimura, Yumiko; Namatame, Nachi; Yamori, Takao; Dan, Shingo

2015-01-01

Drug resistance often critically limits the efficacy of molecular targeted drugs. Although pharmacological inhibition of phosphatidylinositol 3-kinase (PI3K) is an attractive therapeutic strategy for cancer therapy, molecular determinants for efficacy of PI3K inhibitors (PI3Kis) remain unclear. We previously identified that overexpression of insulin-like growth factor 1 receptor (IGF1R) contributed to the development of drug resistance after long-term exposure to PI3Kis. In this study, we examined the involvement of basal IGF1R expression in intrinsic resistance of drug-naïve cancer cells to PI3Kis and whether inhibition of IGF1R overcomes the resistance. We found that cancer cells highly expressing IGF1R showed resistance to dephosphorylation of Akt and subsequent antitumor effect by ZSTK474 treatment. Knockdown of IGF1R by siRNAs facilitated the dephosphorylation and enhanced the drug efficacy. These cells expressed tyrosine-phosphorylated insulin receptor substrate 1 at high levels, which was dependent on basal IGF1R expression. In these cells, the efficacy of ZSTK474 in vitro and in vivo was improved by its combination with the IGF1R inhibitor OSI-906. Finally, we found a significant correlation between the basal expression level of IGF1R and the inefficacy of ZSTK474 in an in vivo human cancer panel, as well as in vitro. These results suggest that basal IGF1R expression affects intrinsic resistance of cancer cells to ZSTK474, and IGF1R is a promising target to improve the therapeutic efficacy. The current results provide evidence of combination therapy of PI3Kis with IGF1R inhibitors for treating IGF1R-positive human cancers. PMID:25483727

The A3 adenosine receptor agonist CF502 inhibits the PI3K, PKB/Akt and NF-kappaB signaling pathway in synoviocytes from rheumatoid arthritis patients and in adjuvant-induced arthritis rats.

PubMed

Ochaion, A; Bar-Yehuda, S; Cohen, S; Amital, H; Jacobson, K A; Joshi, B V; Gao, Z G; Barer, F; Patoka, R; Del Valle, L; Perez-Liz, G; Fishman, P

2008-08-15

The A(3) adenosine receptor (A(3)AR) is over-expressed in inflammatory cells and was defined as a target to combat inflammation. Synthetic agonists to this receptor, such as IB-MECA and Cl-IB-MECA, exert an anti-inflammatory effect in experimental animal models of adjuvant- and collagen-induced arthritis. In this study we present a novel A(3)AR agonist, CF502, with high affinity and selectivity at the human A(3)AR. CF502 induced a dose dependent inhibitory effect on the proliferation of fibroblast-like synoviocytes (FLS) via de-regulation of the nuclear factor-kappa B (NF-kappaB) signaling pathway. Furthermore, CF502 markedly suppressed the clinical and pathological manifestations of adjuvant-induced arthritis (AIA) in a rat experimental model when given orally at a low dose (100 microg/kg). As is typical of other G-protein coupled receptors, the A(3)AR expression level was down-regulated shortly after treatment with agonist CF502 in paw and in peripheral blood mononuclear cells (PBMCs) derived from treated AIA animals. Subsequently, a decrease in the expression levels of protein kinase B/Akt (PKB/Akt), IkappaB kinase (IKK), I kappa B (IkappaB), NF-kappaB and tumor necrosis factor-alpha (TNF-alpha) took place. In addition, the expression levels of glycogen synthase kinase-3 beta (GSK-3beta), beta-catenin, and poly(ADP-ribose)polymerase (PARP), known to control the level and activity of NF-kappaB, were down-regulated upon treatment with CF502. Taken together, CF502 inhibits FLS growth and the inflammatory manifestations of arthritis, supporting the development of A(3)AR agonists for the treatment of rheumatoid arthritis.

Lipid-sensors, enigmatic-orphan and orphan nuclear receptors as therapeutic targets in breast-cancer.

PubMed

Garattini, Enrico; Bolis, Marco; Gianni', Maurizio; Paroni, Gabriela; Fratelli, Maddalena; Terao, Mineko

2016-07-05

Breast-cancer is heterogeneous and consists of various groups with different biological characteristics. Innovative pharmacological approaches accounting for this heterogeneity are needed. The forty eight human Nuclear-Hormone-Receptors are ligand-dependent transcription-factors and are classified into Endocrine-Receptors, Adopted-Orphan-Receptors (Lipid-sensors and Enigmatic-Orphans) and Orphan-receptors. Nuclear-Receptors represent ideal targets for the design/synthesis of pharmacological ligands. We provide an overview of the literature available on the expression and potential role played by Lipid-sensors, Enigmatic-Orphans and Orphan-Receptors in breast-cancer. The data are complemented by an analysis of the expression levels of each selected Nuclear-Receptor in the PAM50 breast-cancer groups, following re-elaboration of the data publicly available. The major aim is to support the idea that some of the Nuclear-Receptors represent largely unexploited therapeutic-targets in breast-cancer treatment/chemo-prevention. On the basis of our analysis, we conclude that the Lipid-Sensors, NR1C3, NR1H2 and NR1H3 are likely to be onco-suppressors in breast-cancer. The Enigmatic-Orphans, NR1F1 NR2A1 and NR3B3 as well as the Orphan-Receptors, NR0B1, NR0B2, NR1D1, NR2F1, NR2F2 and NR4A3 exert a similar action. These Nuclear-Receptors represent candidates for the development of therapeutic strategies aimed at increasing their expression or activating them in tumor cells. The group of Nuclear-Receptors endowed with potential oncogenic properties consists of the Lipid-Sensors, NR1C2 and NR1I2, the Enigmatic-Orphans, NR1F3, NR3B1 and NR5A2, as well as the Orphan-Receptors, NR2E1, NR2E3 and NR6A1. These oncogenic Nuclear-Receptors should be targeted with selective antagonists, reverse-agonists or agents/strategies capable of reducing their expression in breast-cancer cells.

Membrane Estrogen and HER-2 Receptors in Human Breast Cancer

DTIC Science & Technology

2002-07-01

activation of G-proteins, adenylate cyclase, inositol phosphate, calcium homeostasis and/or MAP kinase. These interactions may promote phosphorylation of ER...of breast cancer cells and interact with transmembrane HER-2 growth factor receptors. Expression of HER-2 receptors occurs in many breast cancers...reports of significant cross-talk and interaction between erb B (HER) pathways and estrogen receptor signaling (3,24,27,34-36). It is generally held

Angiotensin II Receptor Antagonism Reduces Transforming Growth Factor Beta and Smad Signaling in Thoracic Aortic Aneurysm

PubMed Central

Nataatmadja, Maria; West, Jennifer; Prabowo, Sulistiana; West, Malcolm

2013-01-01

ABSTRACT Background The expression of transforming growth factor beta (TGF-β) and Smad3 regulates extracellular matrix homeostasis and inflammation in aortic aneurysms. The expression of Smad3 depends on signaling by angiotensin II (AngII) receptor pathways through TGF-β receptor–dependent and –independent pathways. Methods To determine the expression of AngII type 1 (AT1R) and type 2 receptors (AT2R), TGF-β, and Smad3 in thoracic aortic aneurysms, we performed immunohistochemistry testing on tissue and cultured cells derived from subjects with Marfan syndrome (MFS) and bicuspid aortic valve (BAV) malformation and from normal aortas of subjects who were organ donors. Results MFS and BAV aneurysm tissue showed enhanced accumulation of TGF-β and Smad3 in vascular smooth muscle cells (VSMCs) and in inflammatory cells in the subintimal layer and tunica media. The normal aortic wall exhibited minimal TGF-β and Smad3 staining. Cultured VSMCs from MFS and BAV samples showed nuclear Smad3 and strong cytoplasmic TGF-β expression in the cytoplasmic vesicles. In control cells, Smad3 was located mainly in the cytoplasm, and weak cytoplasmic TGF-β was distributed with a pattern similar to that of the aneurysm-derived cells. Compared to normal aorta cells, AT1R and AT2R expression was increased in both aneurysm types. Treatment of cultured VSMCs with the AT1R antagonist losartan caused both reduced TGF-β vesicle localization and nuclear expression of Smad3. Conclusions Increased TGF-β and Smad3 expression in aneurysm tissue and cultured VSMCs is consistent with aberrant TGF-β expression and the activation of Smad3 signaling. Losartan-mediated reduction in TGF-β expression and the cytoplasmic localization of Smad3 support a role for AT1R antagonism in the inhibition of aneurysm progression. PMID:23532685

Oxygen Modulates Human Decidual Natural Killer Cell Surface Receptor Expression and Interactions with Trophoblasts1

PubMed Central

Wallace, Alison E.; Goulwara, Sonu S.; Whitley, Guy S.; Cartwright, Judith E.

2014-01-01

Decidual natural killer (dNK) cells have been shown to both promote and inhibit trophoblast behavior important for decidual remodeling in pregnancy and have a distinct phenotype compared to peripheral blood NK cells. We investigated whether different levels of oxygen tension, mimicking the physiological conditions of the decidua in early pregnancy, altered cell surface receptor expression and activity of dNK cells and their interactions with trophoblast. dNK cells were isolated from terminated first-trimester pregnancies and cultured in oxygen tensions of 3%, 10%, and 21% for 24 h. Cell surface receptor expression was examined by flow cytometry, and the effects of secreted factors in conditioned medium (CM) on the trophoblast cell line SGHPL-4 were assessed in vitro. SGHPL-4 cells treated with dNK cell CM incubated in oxygen tensions of 10% were significantly more invasive (P < 0.05) and formed endothelial-like networks to a greater extent (P < 0.05) than SGHPL-4 cells treated with dNK cell CM incubated in oxygen tensions of 3% or 21%. After 24 h, a lower percentage of dNK cells expressed CD56 at 21% oxygen (P < 0.05), and an increased percentage of dNK cells expressed NKG2D at 10% oxygen (P < 0.05) compared to other oxygen tensions, with large patient variation. This study demonstrates dNK cell phenotype and secreted factors are modulated by oxygen tension, which induces changes in trophoblast invasion and endovascular-like differentiation. Alterations in dNK cell surface receptor expression and secreted factors at different oxygen tensions may represent regulation of function within the decidua during the first trimester of pregnancy. PMID:25232021

Tissue Factor-Factor VIIa Complex Triggers Protease Activated Receptor 2-Dependent Growth Factor Release and Migration in Ovarian Cancer

PubMed Central

Chanakira, Alice; Westmark, Pamela R.; Ong, Irene M.; Sheehan, John P.

2017-01-01

Objective Enhanced tissue factor (TF) expression in epithelial ovarian cancer (EOC) is associated with aggressive disease. Our objective was to evaluate the role of the TF-factor VIIa-protease-activated receptor-2 (PAR-2) pathway in human EOC. Methods TCGA RNAseq data from EOC databases were analyzed for PAR expression. Cell and microparticle (MP) associated TF protein expression (Western blot) and MP-associated coagulant activity were determined in human EOC (SKOV-3, OVCAR-3 and CaOV-3) and control cell lines. PAR-1 and PAR-2 protein expression were similarly examined. The PAR dependence of VEGF-A release (ELISA) and chemotactic migration in response to FVIIa and cellular proliferation in response to thrombin was evaluated with small molecule antagonists. Results Relative mRNA expression consistently demonstrated PAR-2>PAR-1≫PAR-3/4 in multiple EOC datasets. Human EOC cell line lysates confirmed expression of TF, PAR-1 and PAR-2 proteins. MPs isolated from EOC cell lines demonstrated markedly enhanced (4–10 fold) TF coagulant activity relative to control cell lines. FVIIa induced a dose-dependent increase in VEGF-A release (2.5-3 fold) from EOC cell lines that was abrogated by the PAR-2 antagonist ENMD-1068. FVIIa treatment of CaOV-3 and OVCAR-3 cells resulted in increased chemotactic migration that was abolished by ENMD-1068. Thrombin induced dose-dependent EOC cell line proliferation was completely reversed by the PAR-1 antagonist vorapaxar. Small molecule antagonists had no effect on these phenotypes without protease present. Conclusions Enhanced activity of the TF-FVIIa-PAR-2 axis may contribute to the EOC progression via PAR-2 dependent signaling that supports an angiogenic and invasive phenotype and local thrombin generation supporting PAR-1 dependent proliferation. PMID:28148395

Expression of platelet-derived growth factor and its receptors in proliferative disorders of fibroblastic origin.

PubMed Central

Smits, A.; Funa, K.; Vassbotn, F. S.; Beausang-Linder, M.; af Ekenstam, F.; Heldin, C. H.; Westermark, B.; Nistér, M.

1992-01-01

Platelet-derived growth factor (PDGF) is known to stimulate the proliferation of connective tissue-derived cells in vitro. Less is known about its functions in vivo, and the role of PDGF in the development of human tumors has not been clarified. The authors have investigated the occurrence of PDGF and PDGF receptors in a series of proliferative disorders of fibroblastic origin using immunohistochemical and in situ hybridization techniques. High expression of PDGF beta-receptor mRNA and protein was found in the malignant tumors, and also in some benign lesions, such as dermatofibroma. In all these cases, benign as well as malignant, the PDGF B-chain mRNA, and less clearly, the PDGF A-chain mRNA, were coexpressed with the beta-receptor. In contrast, high expression of PDGF alpha-receptor mRNA was only found in fully malignant lesions, i.e., malignant fibrous histiocytoma. These data indicate that an autocrine growth stimulation via the PDGF beta-receptor could occur in an early phase of tumorigenesis, and may be a necessary but insufficient event for the progression into fully malignant human connective tissue lesions. Images Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 PMID:1372158

Role of Eotaxin-1 (CCL11) and CC chemokine receptor 3 (CCR3) in bleomycin-induced lung injury and fibrosis.

PubMed

Huaux, Francois; Gharaee-Kermani, M; Liu, Tianju; Morel, Valérie; McGarry, Bridget; Ullenbruch, Matt; Kunkel, Steven L; Wang, Jun; Xing, Zhou; Phan, Sem H

2005-12-01

Eotaxin-1/CCL11 and its receptor CCR3 are involved in recruitment of eosinophils to diverse tissues, but their role in eosinophil recruitment in pulmonary fibrosis is unclear. The present study examined the pulmonary expression of CCL11 and CCR3 during bleomycin (blm)-induced lung injury and determined their importance in the recruitment of inflammatory cells and the development of lung fibrosis. In mice, blm induced a marked pulmonary expression of CCL11 and CCR3. Immunostaining for CCR3 revealed that this receptor was not only expressed by eosinophils but also by neutrophils. CCL11-deficient (CCL11(-/-)) mice developed significantly reduced pulmonary fibrosis. Expression of profibrotic cytokines such as transforming growth factor-beta1 was diminished in the absence of CCL11. Furthermore, increased lung expression of CCL11 significantly enhanced blm-induced lung fibrosis and production of profibrotic cytokines. These effects were also associated with an increase of eosinophil and neutrophil pulmonary infiltration. In contrast, mice treated with neutralizing CCR3 antibodies developed significantly reduced pulmonary fibrosis, eosinophilia, neutrophilia, and expression of profibrotic cytokines. Together, these data suggest that CCL11 and CCR3 are important in the pulmonary recruitment of granulocytes and play significant pathogenic roles in blm-induced lung fibrosis.

Role of Eotaxin-1 (CCL11) and CC Chemokine Receptor 3 (CCR3) in Bleomycin-Induced Lung Injury and Fibrosis

PubMed Central

Huaux, Francois; Gharaee-Kermani, M.; Liu, Tianju; Morel, Valérie; McGarry, Bridget; Ullenbruch, Matt; Kunkel, Steven L.; Wang, Jun; Xing, Zhou; Phan, Sem H.

2005-01-01

Eotaxin-1/CCL11 and its receptor CCR3 are involved in recruitment of eosinophils to diverse tissues, but their role in eosinophil recruitment in pulmonary fibrosis is unclear. The present study examined the pulmonary expression of CCL11 and CCR3 during bleomycin (blm)-induced lung injury and determined their importance in the recruitment of inflammatory cells and the development of lung fibrosis. In mice, blm induced a marked pulmonary expression of CCL11 and CCR3. Immunostaining for CCR3 revealed that this receptor was not only expressed by eosinophils but also by neutrophils. CCL11-deficient (CCL11−/−) mice developed significantly reduced pulmonary fibrosis. Expression of profibrotic cytokines such as transforming growth factor-β1 was diminished in the absence of CCL11. Furthermore, increased lung expression of CCL11 significantly enhanced blm-induced lung fibrosis and production of profibrotic cytokines. These effects were also associated with an increase of eosinophil and neutrophil pulmonary infiltration. In contrast, mice treated with neutralizing CCR3 antibodies developed significantly reduced pulmonary fibrosis, eosinophilia, neutrophilia, and expression of profibrotic cytokines. Together, these data suggest that CCL11 and CCR3 are important in the pulmonary recruitment of granulocytes and play significant pathogenic roles in blm-induced lung fibrosis. PMID:16314464

PDGFRα promoter polymorphisms and expression patterns influence risk of development of imatinib-induced thrombocytopenia in chronic myeloid leukemia: A study from India.

PubMed

Guru, Sameer Ahmad; Mir, Rashid; Bhat, Musadiq; Najar, Imtiyaz; Zuberi, Mariyam; Sumi, Mamta; Masroor, Mirza; Gupta, Naresh; Saxena, Alpana

2017-10-01

Platelet-derived growth factor receptor has been implicated in many malignant and non-malignant diseases. Platelet-derived growth factor receptor-α is a tyrosine kinase and a side target for imatinib, a revolutionary drug for the treatment of chronic myeloid leukemia that has dramatically improved the survival of chronic myeloid leukemia patients. Given the importance of platelet-derived growth factor receptor in platelet development and its inhibition by imatinib, it was intriguing to analyze the role of platelet-derived growth factor receptor-α in relation to imatinib treatment in the development of imatinib-induced thrombocytopenia in chronic myeloid leukemia patients. We hypothesized that two known functional polymorphisms, +68GA insertion/deletion and -909C/A, in the promoter region of the platelet-derived growth factor receptor-α gene may affect the susceptibility of chronic myeloid leukemia patients receiving imatinib treatment to the development of thrombocytopenia. A case-control study was conducted among a cohort of chronic myeloid leukemia patients admitted to the Lok Nayak Hospital, New Delhi, India. A set of 100 patients of chronic myeloid leukemia in chronic phase and 100 age- and sex-matched healthy controls were studied. After initiation of imatinib treatment, the hematological response of chronic myeloid leukemia patients was monitored regularly for 2 years, in which the development of thrombocytopenia was the primary end point. Platelet-derived growth factor receptor-α promoter polymorphisms +68GA ins/del and -909C/A were studied by allele-specific polymerase chain reaction. Platelet-derived growth factor receptor-α messenger RNA expression was evaluated by quantitative real-time polymerase chain reaction. The messenger RNA expression results were expressed as 2 -Δct ± standard deviation. The distribution of +68GA ins/del promoter polymorphism genotypes differed significantly between the thrombocytopenic and non-thrombocytopenic chronic myeloid leukemia patient groups (p < 0.0001). Moreover, +68GA del/del and ins/del genotypes in imatinib-treated chronic myeloid leukemia patients were associated with an increased risk of developing thrombocytopenia, with odds ratios 6.5 (95% confidence interval = 2.02-0.89, p = 0.001) and 6.0 (95% confidence interval = 2.26-15.91, p = 0.0002), respectively. Similarly, -909C/A promoter polymorphism genotype distribution also differed significantly between thrombocytopenic and non-thrombocytopenic chronic myeloid leukemia patient groups (p = 0.02), and a significantly increased risk of imatinib-induced thrombocytopenia was associated with -909C/A polymorphism mutant homozygous (AA) genotypes the odds ratio being 7.7 (95% confidence interval 1.50 to 39.91, p = 0.009). However, no significant risk of imatinib-induced thrombocytopenia was found to be associated with heterozygous genotype (-909C/A) with odds ratio 1.9 (95% confidence interval = 0.86-4.56, p = 1.14). Platelet-derived growth factor receptor-α messenger RNA expression was significantly higher in chronic myeloid leukemia patients compared to controls (p = 0.008). Moreover, patients with imatinib-induced thrombocytopenia had a significantly lower platelet-derived growth factor receptor-α messenger RNA expression, compared to patients without thrombocytopenia (p = 0.01). A differential expression of platelet-derived growth factor receptor-α messenger RNA was observed with respect to different +68 GA ins/del and -909C/A polymorphism genotypes. The +68GA deletion allele and -909A allele were significantly associated with lower expression of platelet-derived growth factor receptor-α messenger RNA. The platelet-derived growth factor receptor-α +68GA del/del, +68GA ins/del, and -909AA genotypes are associated with an increased risk of developing thrombocytopenia in imatinib-treated chronic myeloid leukemia patients. A significantly lower platelet-derived growth factor receptor-α messenger RNA expression accompanies the +68GA deletion allele in an allele dose-dependent manner. Platelet-derived growth factor receptor-α -909AA genotype is also associated with lower expression of platelet-derived growth factor receptor-α. The downregulation of platelet-derived growth factor receptor-α expression may play a causative role in imatinib-induced thrombocytopenia, a common side effect, in the subset of chronic myeloid leukemia patients with platelet-derived growth factor receptor-α +68 GA ins/del, +68 GA del/del, and -909C/A genotypes.

Enhanced Expression of Fibroblast Growth Factor Receptor 3 IIIc Promotes Human Esophageal Carcinoma Cell Proliferation

PubMed Central

Ueno, Nobuhiro; Shimizu, Akio; Kanai, Michiyuki; Iwaya, Yugo; Ueda, Shugo; Nakayama, Jun; Seo, Misuzu Kurokawa

2015-01-01

Deregulated expression of fibroblast growth factor receptors (FGFRs) and their ligands plays critical roles in tumorigenesis. The gene expression of an alternatively spliced isoforms of FGFR3, FGFR3IIIc, was analyzed by RT-PCR in samples from patients with esophageal carcinoma (EC), including esophageal squamous cell carcinoma (ESCC) and adenocarcinoma (EAC). The incidence of FGFR3IIIc was higher in EC [12/16 (75%); p=0.073] than in non-cancerous mucosa (NCM) [6/16 (38%)]. Indeed, an immunohistochemical analysis of early-stage ESCC showed that carcinoma cells expressing FGFR3IIIc stained positively with SCC-112, a tumor marker, and Ki67, a cell proliferation marker, suggesting that the expression of FGFR3IIIc promotes cell proliferation. We used EC-GI-10 cells endogenously expressing FGFR3IIIc as a model of ESCC to provide mechanistic insight into the role of FGFR3IIIc in ESCC. The knockdown of endogenous FGFR3 using siRNA treatment significantly abrogated cell proliferation and the overexpression of FGFR3IIIc in cells with enhanced cell proliferation. EC-GI-10 cells and ESCC from patients with EC showed endogenous expression of FGF2, a specific ligand for FGFR3IIIc, suggesting that the upregulated expression of FGFR3IIIc may create autocrine FGF signaling in ESCC. Taken together, FGFR3IIIc may have the potential to be an early-stage tumor marker and a molecular target for ESCC therapy. PMID:26487184

Brain-derived neurotrophic factor inhibits glucose intolerance after cerebral ischemia

PubMed Central

Shu, Xiaoliang; Zhang, Yongsheng; Xu, Han; Kang, Kai; Cai, Donglian

2013-01-01

Brain-derived neurotrophic factor is associated with the insulin signaling pathway and glucose tabolism. We hypothesized that expression of brain-derived neurotrophic factor and its receptor may be involved in glucose intolerance following ischemic stress. To verify this hypothesis, this study aimed to observe the changes in brain-derived neurotrophic factor and tyrosine kinase B receptor expression in glucose metabolism-associated regions following cerebral ischemic stress in mice. At day 1 after middle cerebral artery occlusion, the expression levels of brain-derived neurotrophic factor were significantly decreased in the ischemic cortex, hypothalamus, liver, skeletal muscle, and pancreas. The expression levels of tyrosine kinase B receptor were decreased in the hypothalamus and liver, and increased in the skeletal muscle and pancreas, but remained unchanged in the cortex. Intrahypothalamic administration of brain-derived neurotrophic factor (40 ng) suppressed the decrease in insulin receptor and tyrosine-phosphorylated insulin receptor expression in the liver and skeletal muscle, and inhibited the overexpression of gluconeogenesis-associated phosphoenolpyruvate carboxykinase and glucose-6-phosphatase in the liver of cerebral ischemic mice. However, serum insulin levels remained unchanged. Our experimental findings indicate that brain-derived neurotrophic factor can promote glucose metabolism, reduce gluconeogenesis, and decrease blood glucose levels after cerebral ischemic stress. The low expression of brain-derived neurotrophic factor following cerebral ischemia may be involved in the development of glucose intolerance. PMID:25206547

Regulation of ITAM adaptor molecules and their receptors by inhibition of calcineurin-NFAT signalling during late stage osteoclast differentiation

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

Zawawi, M.S.F.; Discipline of Anatomy and Pathology, School of Medical Sciences, University of Adelaide, Adelaide, SA 5005; Dharmapatni, A.A.S.S.K.

2012-10-19

Highlights: Black-Right-Pointing-Pointer Calcineurin/NFAT inhibitors FK506 and VIVIT treated human PBMC derived osteoclasts in vitro. Black-Right-Pointing-Pointer Differential regulation of ITAM receptors and adaptor molecules by calcineurin/NFAT inhibitors. Black-Right-Pointing-Pointer FK506 and VIVIT suppress ITAM factors during late phase osteoclast differentiation. -- Abstract: Osteoclasts are specialised bone resorptive cells responsible for both physiological and pathological bone loss. Osteoclast differentiation and activity is dependent upon receptor activator NF-kappa-B ligand (RANKL) interacting with its receptor RANK to induce the transcription factor, nuclear factor of activated T-cells, cytoplasmic, calcineurin-dependent 1 (NFATc1). The immunoreceptor tyrosine-based activation motif (ITAM)-dependent pathway has been identified as a co-stimulatory pathway inmore » osteoclasts. Osteoclast-associated receptor (OSCAR) and triggering receptor expressed in myeloid cells (TREM2) are essential receptors that pair with adaptor molecules Fc receptor common gamma chain (FcR{gamma}) and DNAX-activating protein 12 kDa (DAP12) respectively to induce calcium signalling. Treatment with calcineurin-NFAT inhibitors, Tacrolimus (FK506) and the 11R-VIVIT (VIVIT) peptide, reduces NFATc1 expression consistent with a reduction in osteoclast differentiation and activity. This study aimed to investigate the effects of inhibiting calcineurin-NFAT signalling on the expression of ITAM factors and late stage osteoclast genes including cathepsin K (CathK), Beta 3 integrin ({beta}3) and Annexin VIII (AnnVIII). Human peripheral blood mononuclear cells (PBMCs) were differentiated with RANKL and macrophage-colony stimulating factor (M-CSF) over 10 days in the presence or absence of FK506 or VIVIT. Osteoclast formation (as assessed by tartrate resistant acid phosphatase (TRAP)) and activity (assessed by dentine pit resorption) were significantly reduced with treatment. Quantitative real-time polymerase chain reaction (qRT-PCR) analysis demonstrated that FK506 treatment significantly (p < 0.05) reduced the expression of NFATc1, CathK, OSCAR, FcR{gamma}, TREM2 and DAP12 during the terminal stage of osteoclast formation. VIVIT treatment significantly (p < 0.05) decreased CathK, OSCAR, FcR{gamma}, and AnnVIII, gene expression. This data suggest FK506 and VIVIT act differently in targeting the calcineurin-NFAT signalling cascade to suppress key mediators of the ITAM pathway during late stage osteoclast differentiation and this is associated with a reduction in both osteoclast differentiation and activity.« less

Inhibition of Epidermal Growth Factor Receptor and Vascular Endothelial Growth Factor Receptor Phosphorylation on Tumor-Associated Endothelial Cells Leads to Treatment of Orthotopic Human Colon Cancer in Nude Mice1

PubMed Central

Sasaki, Takamitsu; Kitadai, Yasuhiko; Nakamura, Toru; Kim, Jang-Seong; Tsan, Rachel Z; Kuwai, Toshio; Langley, Robert R; Fan, Dominic; Kim, Sun-Jin; Fidler, Isaiah J

2007-01-01

The purpose of our study was to determine whether the dual inhibition of epidermal growth factor receptor (EGFR) and vascular endothelial growth factor receptor (VEGFR) signaling pathways in tumor-associated endothelial cells can inhibit the progressive growth of human colon carcinoma in the cecum of nude mice. SW620CE2 human colon cancer cells growing in culture and orthotopically in the cecum of nude mice expressed a high level of transforming growth factor alpha (TGF-α) and vascular endothelial growth factor (VEGF) but were negative for EGFR, human epidermal growth factor receptor 2 (HER2), and VEGFR. Double immunofluorescence staining revealed that tumor-associated endothelial cells expressed EGFR, VEGFR2, phosphorylated EGFR (pEGFR), and phosphorylated VEGFR (pVEGFR). Treatment of mice with either 7H-pyrrolo [2,3-d]-pyrimidine lead scaffold (AEE788; an inhibitor of EGFR and VEGFR tyrosine kinase) or CPT-11 as single agents significantly inhibited the growth of cecal tumors (P < .01); this decrease was even more pronounced with AEE788 combined with CPT-11 (P < .001). AEE788 alone or combined with CPT-11 also inhibited the expression of pEGFR and pVEGFR on tumor-associated endothelial cells, significantly decreased vascularization and tumor cell proliferation, and increased the level of apoptosis in both tumor-associated endothelial cells and tumor cells. These data demonstrate that targeting EGFR and VEGFR signaling on tumor-associated endothelial cells provides a viable approach for the treatment of colon cancer. PMID:18084614

A platelet-activating factor (PAF) receptor deficiency exacerbates diet-induced obesity but PAF/PAF receptor signaling does not contribute to the development of obesity-induced chronic inflammation.

PubMed

Yamaguchi, Masahiko; Matsui, Masakazu; Higa, Ryoko; Yamazaki, Yasuhiro; Ikari, Akira; Miyake, Masaki; Miwa, Masao; Ishii, Satoshi; Sugatani, Junko; Shimizu, Takao

2015-02-15

Platelet-activating factor (PAF) is a well-known phospholipid that mediates acute inflammatory responses. In the present study, we investigated whether PAF/PAF receptor signaling contributed to chronic inflammation in the white adipose tissue (WAT) of PAF receptor-knockout (PAFR-KO) mice. Body and epididymal WAT weights were higher in PAFR-KO mice fed a high-fat diet (HFD) than in wild-type (WT) mice. TNF-α mRNA expression levels in epididymal WAT and the infiltration of CD11c-positive macrophages into epididymal WAT, which led to chronic inflammation, were also elevated in HFD-fed PAFR-KO mice. HFD-fed PAFR-KO mice had higher levels of fasting serum glucose than HFD-fed WT mice as well as impaired glucose tolerance. Although PAF receptor signaling up-regulated the expression of TNF-α and lipopolysaccharide induced the expression of acyl-CoA:lysophosphatidylcholine acyltransferase 2 (LPCAT2) mRNA in bone marrow-derived macrophages, no significant differences were observed in the expression of LPCAT2 mRNA and PAF levels in epididymal WAT between HFD-fed mice and normal diet-fed mice. In addition to our previous finding in which energy expenditure in PAF receptor (PAFR)-deficient mice was low due to impaired brown adipose tissue function, the present study demonstrated that PAF/PAF receptor signaling up-regulated the expression of Ucp1 mRNA, which is essential for cellular thermogenesis, in 3T3-L1 adipocytes. We concluded that the marked accumulation of abdominal fat due to HFD feeding led to more severe chronic inflammation in WAT, which is associated with glucose metabolism disorders, in PAFR-KO mice than in WT mice, and PAF/PAF receptor signaling may regulate energy expenditure and adiposity. Copyright © 2015 Elsevier Inc. All rights reserved.

Developmentally Regulated Expression of the Nerve Growth Factor Receptor Gene in the Periphery and Brain

NASA Astrophysics Data System (ADS)

Buck, C. R.; Martinez, Humberto J.; Black, Ira B.; Chao, Moses V.

1987-05-01

Nerve growth factor (NGF) regulates development and maintenance of function of peripheral sympathetic and sensory neurons. A potential role for the trophic factor in brain has been detected only recently. The ability of a cell to respond to NGF is due, in part, to expression of specific receptors on the cell surface. To study tissue-specific expression of the NGF receptor gene, we have used sensitive cRNA probes for detection of NGF receptor mRNA. Our studies indicate that the receptor gene is selectively and specifically expressed in sympathetic (superior cervical) and sensory (dorsal root) ganglia in the periphery, and by the septum-basal forebrain centrally, in the neonatal rat in vivo. Moreover, examination of tissues from neonatal and adult rats reveals a marked reduction in steady-state NGF receptor mRNA levels in sensory ganglia. In contrast, a 2- to 4-fold increase was observed in the basal forebrain and in the sympathetic ganglia over the same time period. Our observations suggest that NGF receptor mRNA expression is developmentally regulated in specific areas of the nervous system in a differential fashion.

Region-specific expression and hormonal regulation of the first exon variants of rat prolactin receptor mRNA in rat brain and anterior pituitary gland.

PubMed

Nogami, H; Hoshino, R; Ogasawara, K; Miyamoto, S; Hisano, S

2007-08-01

Recent studies have revealed the occurrence of five first exon variants of the rat prolactin receptor mRNA, suggesting that multiple promoters direct prolactin receptor transcription in response to different regulatory factors. In the present study, regional expression of these first exon variants, as well as two prolactin receptor subtypes generated by alternative splicing, was examined in the brains and anterior pituitary glands of female rats. Expression of the long-form was detected in the choroid plexus, hypothalamus, hippocampus, cerebral cortex and anterior pituitary gland, whereas the short form was detected only in the choroid plexus. E1-3 mRNA, a first exon variant, was detected in the choroid plexus, hypothalamus, and anterior pituitary gland, whereas E1-4 was detected only in the choroid plexus. Other variants were not detectable by the polymerase chain reaction protocol employed in this study. Ovariectomy increased the short form in the choroid plexus and the E1-3 expression in the choroid plexus and pituitary gland, but changes in the long-form and E1-4 expression were minimal. Replacement of oestrogens and prolactin suggest that oestrogens down-regulate E1-3 expression in the choroid plexus and pituitary gland, and that the negative effect of oestrogen is mediated by prolactin in the pituitary gland. The present results revealed the region-specific promoter usage in prolactin receptor mRNA transcription, as well as the involvement of oestrogens in the regulation of E1-3 mRNA expression in the brain and pituitary gland.

The Maternal Maverick/GDF15-like TGF-β Ligand Panda Directs Dorsal-Ventral Axis Formation by Restricting Nodal Expression in the Sea Urchin Embryo.

PubMed

Haillot, Emmanuel; Molina, Maria Dolores; Lapraz, François; Lepage, Thierry

2015-01-01

Specification of the dorsal-ventral axis in the highly regulative sea urchin embryo critically relies on the zygotic expression of nodal, but whether maternal factors provide the initial spatial cue to orient this axis is not known. Although redox gradients have been proposed to entrain the dorsal-ventral axis by acting upstream of nodal, manipulating the activity of redox gradients only has modest consequences, suggesting that other factors are responsible for orienting nodal expression and defining the dorsal-ventral axis. Here we uncover the function of Panda, a maternally provided transforming growth factor beta (TGF-β) ligand that requires the activin receptor-like kinases (Alk) Alk3/6 and Alk1/2 receptors to break the radial symmetry of the embryo and orient the dorsal-ventral axis by restricting nodal expression. We found that the double inhibition of the bone morphogenetic protein (BMP) type I receptors Alk3/6 and Alk1/2 causes a phenotype dramatically more severe than the BMP2/4 loss-of-function phenotype, leading to extreme ventralization of the embryo through massive ectopic expression of nodal, suggesting that an unidentified signal acting through BMP type I receptors cooperates with BMP2/4 to restrict nodal expression. We identified this ligand as the product of maternal Panda mRNA. Double inactivation of panda and bmp2/4 led to extreme ventralization, mimicking the phenotype caused by inactivation of the two BMP receptors. Inhibition of maternal panda mRNA translation disrupted the early spatial restriction of nodal, leading to persistent massive ectopic expression of nodal on the dorsal side despite the presence of Lefty. Phylogenetic analysis indicates that Panda is not a prototypical BMP ligand but a member of a subfamily of TGF-β distantly related to Inhibins, Lefty, and TGF-β that includes Maverick from Drosophila and GDF15 from vertebrates. Indeed, overexpression of Panda does not appear to directly or strongly activate phosphoSmad1/5/8 signaling, suggesting that although this TGF-β may require Alk1/2 and/or Alk3/6 to antagonize nodal expression, it may do so by sequestering a factor essential for Nodal signaling, by activating a non-Smad pathway downstream of the type I receptors, or by activating extremely low levels of pSmad1/5/8. We provide evidence that, although panda mRNA is broadly distributed in the early embryo, local expression of panda mRNA efficiently orients the dorsal-ventral axis and that Panda activity is required locally in the early embryo to specify this axis. Taken together, these findings demonstrate that maternal panda mRNA is both necessary and sufficient to orient the dorsal-ventral axis. These results therefore provide evidence that in the highly regulative sea urchin embryo, the activity of spatially restricted maternal factors regulates patterning along the dorsal-ventral axis.

The Maternal Maverick/GDF15-like TGF-β Ligand Panda Directs Dorsal-Ventral Axis Formation by Restricting Nodal Expression in the Sea Urchin Embryo

PubMed Central

Haillot, Emmanuel; Molina, Maria Dolores; Lapraz, François; Lepage, Thierry

2015-01-01

Specification of the dorsal-ventral axis in the highly regulative sea urchin embryo critically relies on the zygotic expression of nodal, but whether maternal factors provide the initial spatial cue to orient this axis is not known. Although redox gradients have been proposed to entrain the dorsal-ventral axis by acting upstream of nodal, manipulating the activity of redox gradients only has modest consequences, suggesting that other factors are responsible for orienting nodal expression and defining the dorsal-ventral axis. Here we uncover the function of Panda, a maternally provided transforming growth factor beta (TGF-β) ligand that requires the activin receptor-like kinases (Alk) Alk3/6 and Alk1/2 receptors to break the radial symmetry of the embryo and orient the dorsal-ventral axis by restricting nodal expression. We found that the double inhibition of the bone morphogenetic protein (BMP) type I receptors Alk3/6 and Alk1/2 causes a phenotype dramatically more severe than the BMP2/4 loss-of-function phenotype, leading to extreme ventralization of the embryo through massive ectopic expression of nodal, suggesting that an unidentified signal acting through BMP type I receptors cooperates with BMP2/4 to restrict nodal expression. We identified this ligand as the product of maternal Panda mRNA. Double inactivation of panda and bmp2/4 led to extreme ventralization, mimicking the phenotype caused by inactivation of the two BMP receptors. Inhibition of maternal panda mRNA translation disrupted the early spatial restriction of nodal, leading to persistent massive ectopic expression of nodal on the dorsal side despite the presence of Lefty. Phylogenetic analysis indicates that Panda is not a prototypical BMP ligand but a member of a subfamily of TGF-β distantly related to Inhibins, Lefty, and TGF-β that includes Maverick from Drosophila and GDF15 from vertebrates. Indeed, overexpression of Panda does not appear to directly or strongly activate phosphoSmad1/5/8 signaling, suggesting that although this TGF-β may require Alk1/2 and/or Alk3/6 to antagonize nodal expression, it may do so by sequestering a factor essential for Nodal signaling, by activating a non-Smad pathway downstream of the type I receptors, or by activating extremely low levels of pSmad1/5/8. We provide evidence that, although panda mRNA is broadly distributed in the early embryo, local expression of panda mRNA efficiently orients the dorsal-ventral axis and that Panda activity is required locally in the early embryo to specify this axis. Taken together, these findings demonstrate that maternal panda mRNA is both necessary and sufficient to orient the dorsal-ventral axis. These results therefore provide evidence that in the highly regulative sea urchin embryo, the activity of spatially restricted maternal factors regulates patterning along the dorsal-ventral axis. PMID:26352141

Aging alters mRNA expression of amyloid transporter genes at the blood-brain barrier.

PubMed

Osgood, Doreen; Miller, Miles C; Messier, Arthur A; Gonzalez, Liliana; Silverberg, Gerald D

2017-09-01

Decreased clearance of potentially toxic metabolites, due to aging changes, likely plays a significant role in the accumulation of amyloid-beta (Aβ) peptides and other macromolecules in the brain of the elderly and in the patients with Alzheimer's disease (AD). Aging is the single most important risk factor for AD development. Aβ transport receptor proteins expressed at the blood-brain barrier are significantly altered with age: the efflux transporters lipoprotein receptor-related protein 1 and P-glycoprotein are reduced, whereas the influx transporter receptor for advanced glycation end products is increased. These receptors play an important role in maintaining brain biochemical homeostasis. We now report that, in a rat model of aging, gene transcription is altered in aging, as measured by Aβ receptor gene messenger RNA (mRNA) at 3, 6, 9, 12, 15, 20, 30, and 36 months. Gene mRNA expression from isolated cerebral microvessels was measured by quantitative polymerase chain reaction. Lipoprotein receptor-related protein 1 and P-glycoprotein mRNA were significantly reduced in aging, and receptor for advanced glycation end products was increased, in parallel with the changes seen in receptor protein expression. Transcriptional changes appear to play a role in aging alterations in blood-brain barrier receptor expression and Aβ accumulation. Copyright © 2017 Elsevier Inc. All rights reserved.

Expression of Inappropriate Cadherins in Human Breast Carcinomas

DTIC Science & Technology

2000-10-01

fibroblast growth factor receptor ADHERINS constitute a family of transmembrane Hamaguchi et al., 1993). In addition, p120ct", originally...1994. expression is associated with poor prognosis in patients with prostate cancer. Alternative splicing in fibroblast growth factor receptor 2 is... fibroblast growth factor receptor signaling. This year we report that the extracellular domain of N-cadherin is responsible for this

Expression of human factors CD81, claudin-1, scavenger receptor, and occludin in mouse hepatocytes does not confer susceptibility to HCV entry.

PubMed

Hikosaka, Keisuke; Noritake, Hidenao; Kimura, Wataru; Sultana, Nishat; Sharkar, Mohammad T K; Tagawa, Yoh-Ichi; Uezato, Tadayoshi; Kobayashi, Yoshimasa; Wakita, Takaji; Miura, Naoyuki

2011-04-01

No suitable mouse model is available for studying chronic liver disease caused by hepatitis C virus (HCV). CD81, claudin-1, scavenger receptor class B type I, and occludin were recently reported to be the important factors in HCV entry into hepatocytes. We made transgenic mice (Alb-CCSO) expressing the four human proteins and examined whether HCV from a patient serum or HCV pseudoparticles (HCVpp) were capable of infecting them. HCV was not detected in the mouse serum after injecting the mice with HCV from a patient serum. We also found no indications of HCVpp entry into primary hepatocytes from Alb-CCSO mice. In addition, HCV-infectible Hep3B cells were fused with HCV-resistant primary mouse hepatocytes and the fused cells showed 35-fold lower infectivity compared to wild-type Hep3B cells, indicating that primary mouse hepatocytes have the inhibitory factor(s) in HCVpp entry. Our results suggest that the expression of the human factors does not confer susceptibility to HCV entry into the liver.

Follicular expression of pro-inflammatory cytokines tumour necrosis factor-α (TNFα), interleukin 6 (IL6) and their receptors in cattle: TNFα, IL6 and macrophages suppress thecal androgen production in vitro.

PubMed

Samir, Moafaq; Glister, Claire; Mattar, Dareen; Laird, Mhairi; Knight, Phil G

2017-07-01

Pro-inflammatory cytokines secreted by macrophages and other cell types are implicated as intraovarian factors affecting different aspects of ovarian function including follicle and corpus luteum 'turnover', steroidogenesis and angiogenesis. Here, we compared granulosal (GC) and thecal (TC) expression of TNF, IL6 and their receptors (TNFRSF1A, TNFRSF1B and IL6R) during bovine antral follicle development; all five mRNA transcripts were detected in both GC and TC and statistically significant cell-type and follicle stage-related differences were evident. Since few studies have examined cytokine actions on TC steroidogenesis, we cultured TC under conditions that retain a non-luteinized 'follicular' phenotype and treated them with TNFα and IL6 under basal and LH-stimulated conditions. Both TNFα and IL6 suppressed androgen secretion concomitantly with CYP17A1 and LHCGR mRNA expression. In addition, TNFα reduced INSL3, HSD3B1 and NOS3 expression but increased NOS2 expression. IL6 also reduced LHCGR and STAR expression but did not affect HSD3B1, INSL3, NOS2 or NOS3 expression. As macrophages are a prominent source of these cytokines in vivo , we next co-cultured TC with macrophages and observed an abolition of LH-induced androgen production accompanied by a reduction in CYP17A1, INSL3, LHCGR, STAR, CYP11A1 and HSD3B1 expression. Exposure of TC to bacterial lipopolysaccharide also blocked LH-induced androgen secretion, an effect reduced by a toll-like receptor blocker (TAK242). Collectively, the results support an inhibitory action of macrophages on thecal androgen production, likely mediated by their secretion of pro-inflammatory cytokines that downregulate the expression of LHCGR, CYP17A1 and INSL3. Bovine theca interna cells can also detect and respond directly to lipopolysaccharide. © 2017 Society for Reproduction and Fertility.

An essential role for IL-2 receptor in regulatory T cell function

PubMed Central

Levine, Andrew G; Fan, Xiying; Klein, Ulf; Zheng, Ye; Gasteiger, Georg; Feng, Yongqiang; Fontenot, Jason D.; Rudensky, Alexander Y.

2016-01-01

Regulatory T (Treg) cells, expressing abundant amounts of the IL-2 receptor (IL-2R), are reliant on IL-2 produced by activated T cells. This feature implied a key role for a simple network based on IL-2 consumption by Treg cells in their suppressor function. However, congenital deficiency in IL-2R results in reduced expression of the Treg cell lineage specification factor Foxp3, confounding experimental efforts to understand the role of IL-2R expression and signaling in Treg suppressor function. Using genetic gain and loss of function approaches, we demonstrate that IL-2 capture is dispensable for control of CD4+ T cells, but is important for limiting CD8+ T cell activation, and that IL-2R dependent STAT5 transcription factor activation plays an essential role in Treg cell suppressor function separable from T cell receptor signaling. PMID:27595233

Tumor Necrosis Factor Receptor-Associated Factor 5 Interacts with the NS3 Protein and Promotes Classical Swine Fever Virus Replication.

PubMed

Lv, Huifang; Dong, Wang; Guo, Kangkang; Jin, Mingxing; Li, Xiaomeng; Li, Cunfa; Zhang, Yanming

2018-06-05

Classical swine fever, caused by classical swine fever virus (CSFV), is a highly contagious and high-mortality viral disease, causing huge economic losses in the swine industry worldwide. CSFV non-structural protein 3 (NS3), a multifunctional protein, plays crucial roles in viral replication. However, how NS3 exactly exerts these functions is currently unknown. Here, we identified tumor necrosis factor receptor-associated factor 5 (TRAF5) as a novel binding partner of the NS3 protein via yeast two-hybrid, co-immunoprecipitation and glutathione S -transferase pull-down assays. Furthermore, we observed that TRAF5 promoted CSFV replication in porcine alveolar macrophages (PAMs). Additionally, CSFV infection or NS3 expression upregulated TRAF5 expression, implying that CSFV may exploit TRAF5 via NS3 for better growth. Moreover, CSFV infection and TRAF5 expression activated p38 mitogen activated protein kinase (MAPK) activity, and inhibition of p38 MAPK activation by the SB203580 inhibitor suppressed CSFV replication. Notably, TRAF5 overexpression did not promote CSFV replication following inhibition of p38 MAPK activation. Our findings reveal that TRAF5 promotes CSFV replication via p38 MAPK activation. This work provides a novel insight into the role of TRAF5 in CSFV replication capacity.

The alpha1-fetoprotein locus is activated by a nuclear receptor of the Drosophila FTZ-F1 family.

PubMed

Galarneau, L; Paré, J F; Allard, D; Hamel, D; Levesque, L; Tugwood, J D; Green, S; Bélanger, L

1996-07-01

The alpha1-fetoprotein (AFP) gene is located between the albumin and alpha-albumin genes and is activated by transcription factor FTF (fetoprotein transcription factor), presumed to transduce early developmental signals to the albumin gene cluster. We have identified FTF as an orphan nuclear receptor of the Drosophila FTZ-F1 family. FTF recognizes the DNA sequence 5'-TCAAGGTCA-3', the canonical recognition motif for FTZ-F1 receptors. cDNA sequence homologies indicate that rat FTF is the ortholog of mouse LRH-1 and Xenopus xFF1rA. Rodent FTF is encoded by a single-copy gene, related to the gene encoding steroidogenic factor 1 (SF-1). The 5.2-kb FTF transcript is translated from several in-frame initiator codons into FTF isoforms (54 to 64 kDa) which appear to bind DNA as monomers, with no need for a specific ligand, similar KdS (approximately equal 3 x 10(-10) M), and similar transcriptional effects. FTF activates the AFP promoter without the use of an amino-terminal activation domain; carboxy-terminus-truncated FTF exerts strong dominant negative effects. In the AFP promoter, FTF recruits an accessory trans-activator which imparts glucocorticoid reactivity upon the AFP gene. FTF binding sites are found in the promoters of other liver-expressed genes, some encoding liver transcription factors; FTF, liver alpha1-antitrypsin promoter factor LFB2, and HNF-3beta promoter factor UF2-H3beta are probably the same factor. FTF is also abundantly expressed in the pancreas and may exert differentiation functions in endodermal sublineages, similar to SF-1 in steroidogenic tissues. HepG2 hepatoma cells seem to express a mutated form of FTF.

Epidermal growth factor induces G protein-coupled receptor 30 expression in estrogen receptor-negative breast cancer cells.

PubMed

Albanito, Lidia; Sisci, Diego; Aquila, Saveria; Brunelli, Elvira; Vivacqua, Adele; Madeo, Antonio; Lappano, Rosamaria; Pandey, Deo Prakash; Picard, Didier; Mauro, Loredana; Andò, Sebastiano; Maggiolini, Marcello

2008-08-01

Different cellular receptors mediate the biological effects induced by estrogens. In addition to the classical nuclear estrogen receptors (ERs)-alpha and -beta, estrogen also signals through the seven-transmembrane G-protein-coupled receptor (GPR)-30. Using as a model system SkBr3 and BT20 breast cancer cells lacking the classical ER, the regulation of GPR30 expression by 17beta-estradiol, the selective GPR30 ligand G-1, IGF-I, and epidermal growth factor (EGF) was evaluated. Transient transfections with an expression plasmid encoding a short 5'-flanking sequence of the GPR30 gene revealed that an activator protein-1 site located within this region is required for the activating potential exhibited only by EGF. Accordingly, EGF up-regulated GPR30 protein levels, which accumulated predominantly in the intracellular compartment. The stimulatory role elicited by EGF on GPR30 expression was triggered through rapid ERK phosphorylation and c-fos induction, which was strongly recruited to the activator protein-1 site found in the short 5'-flanking sequence of the GPR30 gene. Of note, EGF activating the EGF receptor-MAPK transduction pathway stimulated a regulatory loop that subsequently engaged estrogen through GPR30 to boost the proliferation of SkBr3 and BT20 breast tumor cells. The up-regulation of GPR30 by ligand-activated EGF receptor-MAPK signaling provides new insight into the well-known estrogen and EGF cross talk, which, as largely reported, contributes to breast cancer progression. On the basis of our results, the action of EGF may include the up-regulation of GPR30 in facilitating a stimulatory role of estrogen, even in ER-negative breast tumor cells.

RANKL downregulates cell surface CXCR6 expression through JAK2/STAT3 signaling pathway during osteoclastogenesis.

PubMed

Li, Changhong; Zhao, Jinxia; Sun, Lin; Yao, Zhongqiang; Liu, Rui; Huang, Jiansheng; Liu, Xiangyuan

2012-12-14

The receptor activator of nuclear factor-κB ligand (RANKL), as a member of the tumor necrosis factor (TNF) family, plays an essential role in osteoclast differentiation and function. Chemokines and their receptors have recently been shown to play critical roles in osteoclastogenesis, however, whether CXCL16-CXCR6 plays role in RANKL-mediated osteoclastogenesis is unknown. In this study, we first reported that RANKL decreased CXCR6 in a dose-dependent manner, which may be through deactivation of Akt and STAT3 signaling induced by CXCL16. Interestingly, RANKL-mediated CXCR6 reduction may be associated to the activation of STAT3 by phosphorylation. When STAT3 activation was blocked by JAK2/STAT3 inhibitor AG490, RANKL failed to shut down CXCR6 expression during osteoclastogenesis. However, CXCL16 alone did not augment RANKL-mediated osteoclast differentiation and did not alter RANKL-receptor RANK mRNA expression. These results demonstrate that reduction of CXCL16-CXCR6 is critical in RANKL-mediated osteoclastogenesis, which is mainly through the activation of JAK2/STAT3 signaling. CXCL16-CXCR6 axis may become a novel target for the therapeutic intervention of bone resorbing diseases such as rheumatoid arthritis and osteoporosis. Copyright © 2012 Elsevier Inc. All rights reserved.

[Perspectives of the stomach cancer treatment: the introduction of molecular targeted therapy and the hope for cure].

PubMed

Cheung, Dae Young; Kim, Jae Kwang

2013-03-25

The overall survival of patients with gastric cancer has increased markedly in Korea, even higher than those of developed nations in Western world. It is due to the virtue of Korean National Cancer Screening Program and nowadays more than half of patients are diagnosed at the early stage of gastric cancer. However, for patients with unresectable gastric cancer, the outcomes of traditional cytotoxic chemotherapy regimens stay at a median survival of 9-11 months. The knowledge of cancer biology and the data from gene expression profiling has explosively expanded. Alternations in the expression of receptor tyrosine kinases pathways including Human epidermal growth factor receptor 2 (HER2), epidermal growth factor receptor (EGFR), vascular endothelial growth factor (VEGF), phosphatydyl inositol 3 kinase/mammalian target of rapamycin (PI3K/mTOR), hepatocyte growth factor receptor (HGFR/MET), and fibroblast growth factor receptor (FGFR) were proved to be critical in cancer cell survival and biological agents targeting those altered receptor tyrosine kinases, their ligands and downstream effector molecules are developed for anti-cancer purpose. Until now, only trastuzumab succeeded to significantly increase overall survival of patients with HER2 overexpressing gastric cancer. Other agents including bevacizumab, gefitinib, erlotinib, and lapatinib failed to achieve the efficacy in survival gain over standard chemotherapy. Insights about the variations between regions, races, and individuals call for the effort to find reliable predictive biomarkers for drug efficacy and to design finely stratified clinical trials. Compared to current treatment paradigms, it is hoped that molecularly targeted treatment along with conventional cytotoxic chemotherapy will lead to significant gains in survival.

GSK3 Protein Positively Regulates Type I Insulin-like Growth Factor Receptor through Forkhead Transcription Factors FOXO1/3/4

PubMed Central

Huo, Xiaodong; Liu, Shu; Shao, Ting; Hua, Hui; Kong, Qingbin; Wang, Jiao; Luo, Ting; Jiang, Yangfu

2014-01-01

Glycogen synthase kinase-3 (GSK3) has either tumor-suppressive roles or pro-tumor roles in different types of human tumors. A number of GSK3 targets in diverse signaling pathways have been uncovered, such as tuberous sclerosis complex subunit 2 and β-catenin. The O subfamily of forkhead/winged helix transcription factors (FOXO) is known as tumor suppressors that induce apoptosis. In this study, we find that FOXO binds to type I insulin-like growth factor receptor (IGF-IR) promoter and stimulates its transcription. GSK3 positively regulates the transactivation activity of FOXO and stimulates IGF-IR expression. Although kinase-dead GSK3β cannot up-regulate IGF-IR, the constitutively active GSK3β induces IGF-IR expression in a FOXO-dependent manner. Serum starvation or Akt inhibition leads to an increase in IGF-IR expression, which could be blunted by GSK3 inhibition. GSK3β knockdown or GSK3 inhibitor suppresses IGF-I-induced IGF-IR, Akt, and ERK1/2 phosphorylation. Moreover, knockdown of GSK3β or FOXO1/3/4 leads to a decrease in cellular proliferation and abrogates IGF-I-induced hepatoma cell proliferation. These results suggest that GSK3 and FOXO may positively regulate IGF-I signaling and hepatoma cell proliferation. PMID:25053419

Fibroblast Growth Factor 10-Fibroblast Growth Factor Receptor 2b Mediated Signaling Is Not Required for Adult Glandular Stomach Homeostasis

PubMed Central

Sala, Frederic G.; Ford, Henri R.; Bellusci, Saverio; Grikscheit, Tracy C.

2012-01-01

The signaling pathways that are essential for gastric organogenesis have been studied in some detail; however, those that regulate the maintenance of the gastric epithelium during adult homeostasis remain unclear. In this study, we investigated the role of Fibroblast growth factor 10 (FGF10) and its main receptor, Fibroblast growth factor receptor 2b (FGFR2b), in adult glandular stomach homeostasis. We first showed that mouse adult glandular stomach expressed Fgf10, its receptors, Fgfr1b and Fgfr2b, and most of the other FGFR2b ligands (Fgf1, Fgf7, Fgf22) except for Fgf3 and Fgf20. Fgf10 expression was mesenchymal whereas FGFR1 and FGFR2 expression were mostly epithelial. Studying double transgenic mice that allow inducible overexpression of Fgf10 in adult mice, we showed that Fgf10 overexpression in normal adult glandular stomach increased epithelial proliferation, drove mucous neck cell differentiation, and reduced parietal and chief cell differentiation. Although a similar phenotype can be associated with the development of metaplasia, we found that Fgf10 overexpression for a short duration does not cause metaplasia. Finally, investigating double transgenic mice that allow the expression of a soluble form of Fgfr2b, FGF10's main receptor, which acts as a dominant negative, we found no significant changes in gastric epithelial proliferation or differentiation in the mutants. Our work provides evidence, for the first time, that the FGF10-FGFR2b signaling pathway is not required for epithelial proliferation and differentiation during adult glandular stomach homeostasis. PMID:23133671

Involvement of the orphan nuclear estrogen receptor-related receptor α in osteoclast adhesion and transmigration

PubMed Central

Bonnelye, Edith; Saltel, Frédéric; Chabadel, Anne; Zirngibl, Ralph A; Aubin, Jane E; Jurdic, Pierre

2010-01-01

The orphan nuclear receptor, estrogen receptor-related receptor α (ERRα) is expressed in osteoblasts and osteoclasts (OCs) and has been proposed to be a modulator of estrogen signaling. To determine the role of ERRα in OC biology, we knocked down ERRα activity by transient transfection of an siRNA directed against ERRα in the RAW264.7 monocyte–macrophage cell line that differentiates into OCs in the presence of receptor activator of nuclear factor κB-ligands and macrophage colony-stimulating factor. In parallel, stable RAW cell lines expressing a dominant-negative form of ERRα and green fluorescent protein (RAW-GFP-ERRαΔAF2) were used. Expression of OC markers was assessed by real-time PCR, and adhesion and transmigration tests were performed. Actin cytoskeletal organization was visualized using confocal microscopy. We found that RAW264.7 cells expressing siRNA directed against ERRα and RAW-GFP-ERRαΔAF2 OCs displayed abnormal spreading, and decreased osteopontin and β3 integrin subunit expression compared with the corresponding control cells. Decreased adhesion and the absence of podosome belts concomitant with abnormal localization of c-src were also observed in RAW-GFP-ERRαΔAF2-derived OCs. In addition, RAW-GFP-ERRαΔAF2-derived OCs failed to transmigrate through osteoblast cell layers. Our data show that the impairment of ERRα function does not alter OC precursor proliferation and differentiation but does alter the adhesion/spreading and migration capacities of mature OCs. PMID:20841427

Vasohibin 2 promotes epithelial-mesenchymal transition in human breast cancer via activation of transforming growth factor β 1 and hypoxia dependent repression of GATA-binding factor 3.

PubMed

Tu, Min; Li, Zhanjun; Liu, Xian; Lv, Nan; Xi, Chunhua; Lu, Zipeng; Wei, Jishu; Song, Guoxin; Chen, Jianmin; Guo, Feng; Jiang, Kuirong; Wang, Shui; Gao, Wentao; Miao, Yi

2017-03-01

Vasohibin 2 (VASH2) is identified as an angiogenic factor, and has been implicated in tumor angiogenesis, proliferation and epithelial-mesenchymal transition (EMT). To investigate the EMT role of VASH2 in breast cancer, we overexpressed or knocked down expression of VASH2 in human breast cancer cell lines. We observed that VASH2 induced EMT in vitro and in vivo. The transforming growth factor β1 (TGFβ1) pathway was activated by VASH2, and expression of a dominant negative TGFβ type II receptor could block VASH2-mediated EMT. In clinical breast cancer tissues VASH2 positively correlated with TGFβ1 expression, but negatively correlated with E-cadherin (a marker of EMT) expression. Under hypoxic conditions in vitro or in vivo, we found that down-regulation of estrogen receptor 1 (ESR1) in VASH2 overexpressing ESR1 positive cells suppressed E-cadherin. Correlation coefficient analysis indicated that VASH2 and ESR1 expression were negatively correlated in clinical human breast cancer tissues. Further study revealed that a transcription factor of ESR1, GATA-binding factor 3 (GATA3), was down-regulated by VASH2 under hypoxia or in vivo. These findings suggest that VASH2 drives breast cancer cells to undergo EMT by activation of the TGFβ1 pathway and hypoxia dependent repression GATA3-ESR1 pathway, leading to cancer metastasis. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

Influence of Maternal Undernutrition and Overfeeding on Cardiac Ciliary Neurotrophic Factor Receptor and Ventricular Size in Fetal Sheep

PubMed Central

Dong, Feng; Ford, Stephen P.; Nijland, Mark J.; Nathanielsz, Peter W.; Ren, Jun

2008-01-01

Intrauterine nutrition status is reported to correlate with risk of cardiovascular diseases in adulthood. Either under- or over-nutrition during early to mid gestation contributes to altered fetal growth and ventricular geometry. This study was designed to examine myocardial expression of ciliary neurotrophic factor receptor α (CTNFRα) and its down-stream mediator signal transducer and activator of transcription 3 (STAT3) on maternal under- or over-nutrition-induced changes in fetal heart weight. Multiparous ewes were fed with 50% (nutrient-restricted, NR), 100% (control) or 150% (overfed, OF) of NRC requirements from 28 to 78 days of gestation (dG; Term 148 dG). Ewes were euthanized on day 78, and the gravid uteri and fetuses recovered. Ventricular protein expression of CTNFRα, STAT3, phosphorylated STAT3, insulin-like growth factor I receptor (IGF-1R) and IGF binding protein 3 (IGFBP3) were quantitated using western blot. Plasma cortisol levels were higher in both NR and OF fetuses whereas plasma IGF-1 levels were lower and higher, in NR and OF fetuses. Fetal weights were reduced by 29.9% in NR ewes and were increased by 22.2% in fetuses from OF ewes compared to control group. Nutrient restriction did not affect fetal heart or ventricular weights whereas overfeeding increased heart and ventricular weights. Protein expression of CTNFRα in fetal ventricular tissue was reduced in OF group whereas STAT3 and pSTAT3 levels were reduced in both NR and OF groups. Expression of IGF-1R and IGFBP3 was unaffected in either NR or OF group. These data suggested that compared with maternal undernutrition, intrauterine overfeeding during early to mid gestation is associated with increases fetal blood concentrations of cortisol and IGF-1 in association with ventricular hypertrophy where reduced expression of CNTFRα and STAT3 may play a role. PMID:17869083

Vascular endothelial growth factor A (VEGF-A) decreases expression and secretion of pleiotrophin in a VEGF receptor-independent manner.

PubMed

Poimenidi, Evangelia; Theodoropoulou, Christina; Koutsioumpa, Marina; Skondra, Lamprini; Droggiti, Eirini; van den Broek, Marloes; Koolwijk, Pieter; Papadimitriou, Evangelia

2016-05-01

Vascular endothelial growth factor A (VEGF-A) is a key molecule in angiogenesis acting through VEGF receptors (VEGFRs), ανβ3 integrin, receptor protein tyrosine phosphatase beta/zeta (RPTPβ/ζ) and cell surface nucleolin (NCL). Pleiotrophin (PTN) stimulates endothelial cell migration and limits the angiogenic effects of VEGF-A165 to the levels of its own effect, possibly acting as a VEGF-A165 modifier. Since PTN and VEGF-A165 share receptors and actions on endothelial cells, in the present work we studied whether and how VEGF-A165 affects PTN expression or secretion. VEGF-A165 decreased PTN mRNA and protein levels acting at the transcriptional level. Bevacizumab, a selective VEGFR2 tyrosine kinase inhibitor and down-regulation of VEGFR2 expression by siRNA did not affect this decrease, suggesting that it is VEGFR-independent. VEGF-A121 also decreased PTN mRNA and protein levels, suggesting that heparin binding of VEGF-A165 is not involved. Blockage of cell surface NCL, lack of expression or mutation of β3 integrin and down-regulation of RPTPβ/ζ abolished the inhibitory effect of VEGF-A165 on PTN expression and secretion. Down-regulation of endogenous PTN in endothelial cells enhanced VEGF-A165-induced increase in migration and tube formation on matrigel. Collectively, these data suggest that VEGF-A down-regulates PTN expression and secretion through the RPTPβ/ζ-ανβ3-NCL axis to enhance its own effect on cell migration and further highlight the role of RPTPβ/ζ in VEGF-A actions. Copyright © 2016 Elsevier Inc. All rights reserved.

Activation of phosphatidylinositol-3 kinase by nerve growth factor involves indirect coupling of the trk proto-oncogene with src homology 2 domains.

PubMed

Ohmichi, M; Decker, S J; Saltiel, A R

1992-10-01

Growth factor receptor tyrosine kinases can form stable associations with intracellular proteins that contain src homology (SH) 2 domains, including the p85 regulatory subunit of phosphatidylinositol (PI)-3 kinase. The activation of this enzyme by growth factors is evaluated in PC12 pheochromocytoma cells and NIH 3T3 fibroblasts expressing the pp140c-trk nerve growth factor (NGF) receptor (3T3-c-trk). NGF causes the rapid stimulation of PI-3 kinase activity detected in anti-phosphotyrosine, but not in anti-trk, immunoprecipitates. This effect coincides with the tyrosine phosphorylation of two proteins, with molecular masses of of 100 kd and 110 kd, that coimmunoprecipitate with p85. Similar phosphorylation patterns are induced when an immobilized fusion protein containing the amino-terminal SH2 domain of p85 is used to precipitate tyrosine-phosphorylated proteins. Thus, although NGF produces the rapid activation of PI-3 kinase through a mechanism that involves tyrosine phosphorylation, there is no evidence for tyrosine phosphorylation of p85, or for its ligand-dependent association with the NGF receptor. Perhaps another phosphoprotein may link the NGF receptor to this enzyme.

Altered trafficking and unfolded protein response induction as a result of M3 muscarinic receptor impaired N-glycosylation.

PubMed

Romero-Fernandez, Wilber; Borroto-Escuela, Dasiel O; Alea, Mileidys Perez; Garcia-Mesa, Yoelvis; Garriga, Pere

2011-12-01

The human M(3) muscarinic acetylcholine receptor is present in both the central and peripheral nervous system, and it is involved in the pathophysiology of several neurodegenerative and autoimmune diseases. We suggested a possible N-glycosylation map for the M(3) muscarinic receptor expressed in COS-7 cells. Here, we examined the role that N-linked glycans play in the folding and in the cell surface trafficking of this receptor. The five potential asparagine-linked glycosylation sites in the muscarinic receptor were mutated and transiently expressed in COS-7 cells. The elimination of N-glycan attachment sites did not affect the cellular expression levels of the receptor. However, proper receptor localization to the plasma membrane was affected as suggested by reduced [(3)H]-N-methylscopolamine binding. Confocal microscopy confirmed this observation and showed that the nonglycosylated receptor was primarily localized in the intracellular compartments. The mutant variant showed an increase in phosphorylation of the α-subunit of eukaryote initiation factor 2, and other well-known endoplasmic reticulum stress markers of the unfolded protein response pathway, which further supports the proposal of the improper intracellular accumulation of the nonglycosylated receptor. The receptor devoid of glycans showed more susceptibility to events that culminate in apoptosis reducing cell viability. Our findings suggest up-regulation of pro-apoptotic Bax protein, down-regulation of anti-apoptotic Bcl-2, and cleavage of caspase-3 effectors. Collectively, our data provide experimental evidence of the critical role that N-glycan chains play in determining muscarinic receptor distribution, localization, as well as cell integrity. © The Author 2011. Published by Oxford University Press. All rights reserved.

Vitamin D modulates tissue factor and protease-activated receptor 2 expression in vascular smooth muscle cells.

PubMed

Martinez-Moreno, Julio M; Herencia, Carmen; Montes de Oca, Addy; Muñoz-Castañeda, Juan R; Rodríguez-Ortiz, M Encarnación; Díaz-Tocados, Juan M; Peralbo-Santaella, Esther; Camargo, Antonio; Canalejo, Antonio; Rodriguez, Mariano; Velasco-Gimena, Francisco; Almaden, Yolanda

2016-03-01

Clinical and epidemiologic studies reveal an association between vitamin D deficiency and increased risk of cardiovascular disease. Because vascular smooth muscle cell (VSMC)-derived tissue factor (TF) is suggested to be critical for arterial thrombosis, we investigated whether the vitamin D molecules calcitriol and paricalcitol could reduce the expression of TF induced by the proinflammatory cytokine TNF-α in human aortic VSMCs. We found that, compared with controls, incubation with TNF-α increased TF expression and procoagulant activity in a NF-κB-dependent manner, as deduced from the increased nuclear translocation of nuclear factor κ-light-chain-enhancer of activated B cells protein 65 (p65-NF-κB) and direct interaction of NF-κB to the TF promoter. This was accompanied by the up-regulation of TF signaling mediator protease-activated receptor 2 (PAR-2) expression and by the down-regulation of vitamin D receptor expression in a miR-346-dependent way. However, addition of calcitriol or paricalcitol blunted the TNF-α-induced TF expression and activity (2.01 ± 0.24 and 1.32 ± 0.14 vs. 3.02 ± 0.39 pmol/mg protein, P < 0.05), which was associated with down-regulation of NF-κB signaling and PAR-2 expression, as well as with restored levels of vitamin D receptor and enhanced expression of TF pathway inhibitor. Our data suggest that inflammation promotes a prothrombotic state through the up-regulation of TF function in VSMCs and that the beneficial cardiovascular effects of vitamin D may be partially due to decreases in TF expression and its activity in VSMCs. © FASEB.

Leiomyoma-derived transforming growth factor-β impairs bone morphogenetic protein-2-mediated endometrial receptivity.

PubMed

Doherty, Leo F; Taylor, Hugh S

2015-03-01

To determine whether transforming growth factor (TGF)-β3 is a paracrine signal secreted by leiomyoma that inhibits bone morphogenetic protein (BMP)-mediated endometrial receptivity and decidualization. Experimental. Laboratory. Women with symptomatic leiomyomas. Endometrial stromal cells (ESCs) and leiomyoma cells were isolated from surgical specimens. Leiomyoma-conditioned media (LCM) was applied to cultured ESC. The TGF-β was blocked by two approaches: TGF-β pan-specific antibody or transfection with a mutant TGF-β receptor type II. Cells were then treated with recombinant human BMP-2 to assess BMP responsiveness. Expression of BMP receptor types 1A, 1B, 2, as well as endometrial receptivity mediators HOXA10 and leukemia inhibitory factor (LIF). Enzyme-linked immunosorbent assay showed elevated TGF-β levels in LCM. LCM treatment of ESC reduced expression of BMP receptor types 1B and 2 to approximately 60% of pretreatment levels. Preincubation of LCM with TGF-β neutralizing antibody or mutant TGF receptor, but not respective controls, prevented repression of BMP receptors. HOXA10 and LIF expression was repressed in recombinant human BMP-2 treated, LCM exposed ESC. Pretreatment of LCM with TGF-β antibody or transfection with mutant TGF receptor prevented HOXA10 and LIF repression. Leiomyoma-derived TGF-β was necessary and sufficient to alter endometrial BMP-2 responsiveness. Blockade of TGF-β prevents repression of BMP-2 receptors and restores BMP-2-stimulated expression of HOXA10 and LIF. Blockade of TGF signaling is a potential strategy to improve infertility and pregnancy loss associated with uterine leiomyoma. Copyright © 2015 American Society for Reproductive Medicine. Published by Elsevier Inc. All rights reserved.

Estradiol regulates the insulin-like growth factor-I (IGF-I) signalling pathway: A crucial role of phosphatidylinositol 3-kinase (PI 3-kinase) in estrogens requirement for growth of MCF-7 human breast carcinoma cells

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

Bernard, Laurence; Legay, Christine; Adriaenssens, Eric

2006-12-01

Estrogens can stimulate the proliferation of estrogen-responsive breast cancer cells by increasing their proliferative response to insulin-like growth factors. With a view to investigating the molecular mechanisms implicated, we studied the effect of estradiol on the expression of proteins implicated in the insulin-like growth factor signalling pathway. Estradiol dose- and time-dependently increased the expression of insulin receptor substrate-1 and the p85/p110 subunits of phosphatidylinositol 3-kinase but did not change those of ERK2 and Akt/PKB. ICI 182,780 did not inhibit estradiol-induced IRS-1 and p85 expression. Moreover, two distinct estradiol-BSA conjugate compounds were as effective as estradiol in inducing IRS-1 and p85/p110more » expression indicating the possible implication of an estradiol membrane receptor. Comparative analysis of steroids-depleted and steroids-treated cells showed that IGF-I only stimulates cell growth in the latter condition. Nevertheless, expression of a constitutively active form of PI 3-kinase in steroid-depleted cells triggers proliferation. These results demonstrate that estradiol positively regulates essential proteins of the IGF signalling pathway and put in evidence that phosphatidylinositol 3-kinase plays a central role in the synergistic pro-proliferative action of estradiol and IGF-I.« less

Purinergic receptor ligands stimulate pro-opiomelanocortin gene expression in AtT-20 pituitary corticotroph cells.

PubMed

Zhao, L-F; Iwasaki, Y; Oki, Y; Tsugita, M; Taguchi, T; Nishiyama, M; Takao, T; Kambayashi, M; Hashimoto, K

2006-04-01

Although recent studies have suggested that purinergic receptors are expressed in the anterior pituitary gland, their involvement in the regulation of pituitary hormone gene expression is not completely understood. In the present study, we examined the expression of purinergic receptors and the effects of purinergic receptor ligands on pro-opiomelanocortin (POMC) gene expression, in AtT20 mouse corticotroph cells. We identified the expression of most of the purinergic receptor subtypes (A1, A2, P2X1, 3-7, P2Y1, 2, 4) mRNAs, analysed by the reverse transcriptase-polymerase chain reaction. We also found that adenosine and ATP, two representative and endogenous agonists of A1-3 and P2X/P2Y receptors, respectively, stimulated the 5'-promoter activity of the POMC gene in a dose- and time-related manner. When these ligands were simultaneously used with corticotrophin-releasing hormone (CRH), effects that were more than additive were observed, suggesting an enhancing role of these compounds in CRH-mediated adrenocorticotrophic hormone (ACTH) synthesis. These ligands also stimulated the expression of transcription factors involved in the regulation of the POMC gene, but did not enhance ACTH secretion. Finally, the positive effect of adenosine as well as CRH was completely inhibited by the protein kinase A inhibitor H89, whereas that of ATP was not influenced, indicating that different intracellular signalling pathways mediate these effects. Altogether, our results suggest a stimulatory role for these purinergic receptor ligands in the regulation of POMC gene expression in corticotroph cells. Because adenosine and ATP are known to be produced within the pituitary gland, it is possible they may be acting in an autocrine/paracrine fashion.

Prostate cancer targeting motifs: expression of αν β3, neurotensin receptor 1, prostate specific membrane antigen, and prostate stem cell antigen in human prostate cancer cell lines and xenografts.

PubMed

Taylor, Robert M; Severns, Virginia; Brown, David C; Bisoffi, Marco; Sillerud, Laurel O

2012-04-01

Membrane receptors are frequent targets of cancer therapeutic and imaging agents. However, promising in vitro results often do not translate to in vivo clinical applications. To better understand this obstacle, we measured the expression differences in receptor signatures among several human prostate cancer cell lines and xenografts as a function of tumorigenicity. Messenger RNA and protein expression levels for integrin α(ν) β(3), neurotensin receptor 1 (NTSR1), prostate specific membrane antigen (PSMA), and prostate stem cell antigen (PSCA) were measured in LNCaP, C4-2, and PC-3 human prostate cancer cell lines and in murine xenografts using quantitative reverse transcriptase polymerase chain reaction, flow cytometry, and immunohistochemistry. Stable expression patterns were observed for integrin α(ν) and PSMA in all cells and corresponding xenografts. Integrin β(3) mRNA expression was greatly reduced in C4-2 xenografts and greatly elevated in PC-3 xenografts compared with the corresponding cultured cells. NTSR1 mRNA expression was greatly elevated in LNCaP and PC-3 xenografts. PSCA mRNA expression was elevated in C4-2 xenografts when compared with C4-2 cells cultured in vitro. Furthermore, at the protein level, PSCA was re-expressed in all xenografts compared with cells in culture. The regulation of mRNA and protein expression of the cell-surface target proteins α(ν) β(3), NTSR1, PSMA, and PSCA, in prostate cancer cells with different tumorigenic potential, was influenced by factors of the microenvironment, differing between cell cultures and murine xenotransplants. Integrin α(ν) β(3), NTRS1 and PSCA mRNA expression increased with tumorigenic potential, but mRNA expression levels for these proteins do not translate directly to equivalent expression levels of membrane bound protein. Copyright © 2011 Wiley Periodicals, Inc.

A selective antagonist reveals a potential role of G protein-coupled receptor 55 in platelet and endothelial cell function.

PubMed

Kargl, Julia; Brown, Andrew J; Andersen, Liisa; Dorn, Georg; Schicho, Rudolf; Waldhoer, Maria; Heinemann, Akos

2013-07-01

The G protein-coupled receptor 55 (GPR55) is a lysophosphatidylinositol (LPI) receptor that is also responsive to certain cannabinoids. Although GPR55 has been implicated in several (patho)physiologic functions, its role remains enigmatic owing mainly to the lack of selective GPR55 antagonists. Here we show that the compound CID16020046 ((4-[4-(3-hydroxyphenyl)-3-(4-methylphenyl)-6-oxo-1H,4H,5H,6H-pyrrolo[3,4-c]pyrazol-5-yl] benzoic acid) is a selective GPR55 antagonist. In yeast cells expressing human GPR55, CID16020046 antagonized agonist-induced receptor activation. In human embryonic kidney (HEK293) cells stably expressing human GPR55, the compound behaved as an antagonist on LPI-mediated Ca²⁺ release and extracellular signal-regulated kinases activation, but not in HEK293 cells expressing cannabinoid receptor 1 or 2 (CB₁ or CB₂). CID16020046 concentration dependently inhibited LPI-induced activation of nuclear factor of activated T-cells (NFAT), nuclear factor κ of activated B cells (NF-κB) and serum response element, translocation of NFAT and NF-κB, and GPR55 internalization. It reduced LPI-induced wound healing in primary human lung microvascular endothelial cells and reversed LPI-inhibited platelet aggregation, suggesting a novel role for GPR55 in platelet and endothelial cell function. CID16020046 is therefore a valuable tool to study GPR55-mediated mechanisms in primary cells and tissues.

Selective over-expression of fibroblast growth factor receptors 1 and 4 in clinical prostate cancer.

PubMed

Sahadevan, K; Darby, S; Leung, H Y; Mathers, M E; Robson, C N; Gnanapragasam, V J

2007-09-01

Fibroblast growth factor receptors (FGFRs) mediate the tumourigenic effects of FGFs in prostate cancer. These receptors are therefore potential therapeutic targets in the development of inhibitors to this pathway. To identify the most relevant targets, we simultaneously investigated FGFR1-4 expression using a prostate cancer tissue microarray (TMA) and in laser capture microdissected (LCM) prostate epithelial cells. In malignant prostates (n = 138) we observed significant FGFR1 and FGFR4 protein over-expression in comparison with benign prostates (n = 58; p < 0.0001). FGFR1 was expressed at high levels in the majority of tumours (69% of grade 3 or less, 74% of grade 4 and 70% of grade 5), while FGFR4 was strongly expressed in 83% of grade 5 cancers but in only 25% of grade 1-3 cancers (p < 0.0001). At the transcript level we observed a similar pattern, with FGFR1 and FGFR4 mRNA over-expressed in malignant epithelial cells compared to benign cells (p < 0.0005 and p < 0.05, respectively). While total FGFR2 was increased in some cancers, there was no association between expression and tumour grade or stage. Transcript analysis, however, revealed a switch in the predominant isoform expressed from FGFR2IIIb to FGFR2IIIc among malignant epithelial cells. In contrast, protein and transcript expression of FGFR3 was very similar between benign and cancer biopsies. The functional effect of targeting FGFR4 in prostate cancer cells has not previously been investigated. In in vitro experiments, suppression of FGFR4 by RNA interference effectively blocked prostate cancer cell proliferation (p < 0.0001) and invasion (p < 0.001) in response to exogenous stimulation. This effect was evident regardless of whether the cells expressed the FGFR4 Arg388 or Gly388 allele. In parallel experiments, FGFR3 suppression had no discernible effect on cancer cell behaviour. These results suggest evidence of selective over-expression of FGFR1 and FGFR4 in clinical prostate cancer and support the notion of targeted inhibition of these receptors to disrupt FGF signalling.

Controlled and reversible induction of differentiation and activation of adult human hepatocytes by a biphasic culture technique

PubMed Central

Auth, Marcus K.H.; Boost, Kim A.; Leckel, Kerstin; Beecken, Wolf-Dietrich; Engl, Tobias; Jonas, Dietger; Oppermann, Elsie; Hilgard, Philip; Markus, Bernd H.; Bechstein, Wolf-Otto; Blaheta, Roman A.

2005-01-01

AIM: Clinical application of human hepatocytes (HC) is hampered by the progressive loss of growth and differentiation in vitro. The object of the study was to evaluate the effect of a biphasic culture technique on expression and activation of growth factor receptors and differentiation of human adult HC. METHODS: Isolated HC were sequentially cultured in a hormone enriched differentiation medium (DM) containing nicotinamide, insulin, transferrin, selenium, and dexame-thasone or activation medium (AM) containing hepatocyte growth factor (HGF), epidermal growth factor (EGF), and granulocyte-macrophage colony-stimulating factor (GM-CSF). Expression, distribution and activation of the HC receptors (MET and EGFR) and the pattern of characteristic cytokeratin (CK) filaments were measured by fluorometry, confocal microscopy and Western blotting. RESULTS: In the biphasic culture system, HC underwent repeated cycles of activation (characterized by expression and activation of growth factor receptors) and re-differentiation (illustrated by distribution of typical filaments CK-18 but low or absent expression of CK-19). In AM increased expression of MET and EGFR was associated with receptor translocation into the cytoplasm and induction of atypical CK-19. In DM low expression of MET and EGFR was localized on the cell membrane and CK-19 was reduced. Receptor phosphorylation required embedding of HC in collagen type I gel. CONCLUSION: Control and reversible modulation of growth factor receptor activation of mature human HC can be accomplished in vitro, when defined signals from the extracellular matrix and sequential growth stimuli are provided. The biphasic technique helps overcome de-differentiation, which occurs during continuous stimulation by means of growth factors. PMID:15810072

Controlled and reversible induction of differentiation and activation of adult human hepatocytes by a biphasic culture technique.

PubMed

Auth, Marcus-K H; Boost, Kim A; Leckel, Kerstin; Beecken, Wolf-Dietrich; Engl, Tobias; Jonas, Dietger; Oppermann, Elsie; Hilgard, Philip; Markus, Bernd H; Bechstein, Wolf-Otto; Blaheta, Roman A

2005-04-14

Clinical application of human hepatocytes (HC) is hampered by the progressive loss of growth and differentiation in vitro. The object of the study was to evaluate the effect of a biphasic culture technique on expression and activation of growth factor receptors and differentiation of human adult HC. Isolated HC were sequentially cultured in a hormone enriched differentiation medium (DM) containing nicotinamide, insulin, transferrin, selenium, and dexame-thasone or activation medium (AM) containing hepatocyte growth factor (HGF), epidermal growth factor (EGF), and granulocyte-macrophage colony-stimulating factor (GM-CSF). Expression, distribution and activation of the HC receptors (MET and EGFR) and the pattern of characteristic cytokeratin (CK) filaments were measured by fluorometry, confocal microscopy and Western blotting. In the biphasic culture system, HC underwent repeated cycles of activation (characterized by expression and activation of growth factor receptors) and re-differentiation (illustrated by distribution of typical filaments CK-18 but low or absent expression of CK-19). In AM increased expression of MET and EGFR was associated with receptor translocation into the cytoplasm and induction of atypical CK-19. In DM low expression of MET and EGFR was localized on the cell membrane and CK-19 was reduced. Receptor phosphorylation required embedding of HC in collagen type I gel. Control and reversible modulation of growth factor receptor activation of mature human HC can be accomplished in vitro, when defined signals from the extracellular matrix and sequential growth stimuli are provided. The biphasic technique helps overcome de-differentiation, which occurs during continuous stimulation by means of growth factors.

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

Dewitt, Ann E.; Dong, Jian Y.; Wiley, H S.

Autocrine signaling is important in normal tissue physiology as well as pathological conditions. It is difficult to analyze these systems, however, because they are both self-contained and recursive. To understand how parameters, such as ligand production and receptor expression influence autocrine activity, we investigated a human epidermal growth factor/epidermal growth factor receptor (EGF/EGFR) loop engineered into mouse B82 fibroblasts. We varied the level of ligand production using the tet-off expression system and used metalloprotease inhibitors to modulate ligand release. Receptor expression was varied using antagonistic, blocking antibodies. We compared autocrine ligand release to receptor activation using a microphysiometer-based assay andmore » analyzed our data with a quantitative model of ligand release and receptor dynamics. We found that the activity of our autocrine system could be described in terms of a simple ratio between the rate of ligand production (VL) and the rate of receptor production (VR). At a VL/VR ratio of < 0.3, essentially no ligand was found in the extracellular medium, but a significant number cell receptors (30-40%) were occupied. As the VL/VR ratio increased from 0.3 towards unity, receptor occupancy increased, and significant amounts of ligand now appeared in the medium. Above a VL/VR ratio of 1.0, receptor occupancy approached saturation and most of the released ligand was lost into the medium. Analysis of human mammary epithelial cells showed that a VL/VR ratio of < 5 x 10 -4 was sufficient to evoke >20% of a maximal proliferative response. This suggests that natural autocrine systems are active even when no ligand appears in the extracellular medium; i.e., they operate 'invisibly' to general detection.« less

Increased tumor necrosis factor receptor 1 expression in human colorectal adenomas

PubMed Central

Hosono, Kunihiro; Yamada, Eiji; Endo, Hiroki; Takahashi, Hirokazu; Inamori, Masahiko; Hippo, Yoshitaka; Nakagama, Hitoshi; Nakajima, Atsushi

2012-01-01

AIM: To determine the expression statuses of tumor necrosis factor (TNF)-α, its receptors (TNF-R) and downstream effector molecules in human colorectal adenomas. METHODS: We measured the serum concentrations of TNF-α and its receptors in 62 colorectal adenoma patients and 34 healthy controls. The protein expression of TNF-α, TNF-R1, TNF-R2 and downstream signals of the TNF receptors, such as c-Jun N-terminal kinase (JNK), nuclear factor-κ B and caspase-3, were also investigated in human colorectal adenomas and in normal colorectal mucosal tissues by immunohistochemistry. Immunofluorescence confocal microscopy was used to investigate the consistency of expression of TNF-R1 and phospho-JNK (p-JNK). RESULTS: The serum levels of soluble TNF-R1 (sTNF-R1) in adenoma patients were significantly higher than in the control group (3.67 ± 0.86 ng/mL vs 1.57 ± 0.72 ng/mL, P < 0.001). Receiver operating characteristic analysis revealed the high diagnostic sensitivity of TNF-R1 measurements (AUC was 0.928) for the diagnosis of adenoma, and the best cut-off level of TNF-R1 was 2.08 ng/mL, with a sensitivity of 93.4% and a specificity of 82.4%. There were no significant differences in the serum levels of TNF-α or sTNF-R2 between the two groups. Immunohistochemistry showed high levels of TNF-R1 and p-JNK expression in the epithelial cells of adenomas. Furthermore, a high incidence of co-localization of TNF-R1 and p-JNK was identified in adenoma tissue. CONCLUSION: TNF-R1 may be a promising biomarker of colorectal adenoma, and it may also play an important role in the very early stages of colorectal carcinogenesis. PMID:23082052

Androgen receptor expression in breast cancer in relation to molecular phenotype: results from the Nurses' Health Study.

PubMed

Collins, Laura C; Cole, Kimberly S; Marotti, Jonathan D; Hu, Rong; Schnitt, Stuart J; Tamimi, Rulla M

2011-07-01

Previous studies have demonstrated that androgen receptor is expressed in many breast cancers, but its expression in relation to the various breast cancer subtypes as defined by molecular profiling has not been studied in detail. We constructed tissue microarrays from 3093 breast cancers that developed in women enrolled in the Nurses' Health Study. Tissue microarray sections were immunostained for estrogen receptor (ER), progesterone receptor (PR), human epidermal growth factor receptor 2 (HER2), cytokeratin 5/6, epidermal growth factor receptor (EGFR) and androgen receptor (ER). Immunostain results were used to categorize each cancer as luminal A or B, HER2 and basal like. The relationships between androgen receptor expression and molecular subtype were analyzed. Overall, 77% of the invasive breast carcinomas were androgen receptor positive. Among 2171 invasive cancers, 64% were luminal A, 15% luminal B, 6% HER2 and 11% basal like. The frequency of androgen receptor expression varied significantly across the molecular phenotypes (P<0.0001). In particular, androgen receptor expression was commonly observed in luminal A (91%) and B (68%) cancers, but was less frequently seen in HER2 cancers (59%). Despite being defined by the absence of ER and PR expression and being considered hormonally unresponsive, 32% of basal-like cancers expressed androgen receptor. Among 246 cases of ductal carcinoma in situ, 86% were androgen receptor positive, but the frequency of androgen receptor expression differed significantly across the molecular phenotypes (P=0.001), and high nuclear grade lesions were less likely to be androgen receptor positive compared with lower-grade lesions. Androgen receptor expression is most commonly seen in luminal A and B invasive breast cancers. However, expression of androgen receptor is also seen in approximately one-third of basal-like cancers, providing further evidence that basal-like cancers represent a heterogeneous group. Our findings raise the possibility that targeting the androgen receptor pathway may represent a novel therapeutic approach to the management of patients with basal-like cancers.

Distribution and expression of non-neuronal transient receptor potential (TRPV) ion channels in rosacea.

PubMed

Sulk, Mathias; Seeliger, Stephan; Aubert, Jerome; Schwab, Verena D; Cevikbas, Ferda; Rivier, Michel; Nowak, Pawel; Voegel, Johannes J; Buddenkotte, Jörg; Steinhoff, Martin

2012-04-01

Rosacea is a frequent chronic inflammatory skin disease of unknown etiology. Because early rosacea reveals all characteristics of neurogenic inflammation, a central role of sensory nerves in its pathophysiology has been discussed. Neuroinflammatory mediators and their receptors involved in rosacea are poorly defined. Good candidates may be transient receptor potential (TRP) ion channels of vanilloid type (TRPV), which can be activated by many trigger factors of rosacea. Interestingly, TRPV2, TRPV3, and TRPV4 are expressed by both neuronal and non-neuronal cells. Here, we analyzed the expression and distribution of TRPV receptors in the various subtypes of rosacea on non-neuronal cells using immunohistochemistry, morphometry, double immunoflourescence, and quantitative real-time PCR (qRT-PCR) as compared with healthy skin and lupus erythematosus. Our results show that dermal immunolabeling of TRPV2 and TRPV3 and gene expression of TRPV1 is significantly increased in erythematotelangiectatic rosacea (ETR). Papulopustular rosacea (PPR) displayed an enhanced immunoreactivity for TRPV2, TRPV4, and also of TRPV2 gene expression. In phymatous rosacea (PhR)-affected skin, dermal immunostaining of TRPV3 and TRPV4 and gene expression of TRPV1 and TRPV3 was enhanced, whereas epidermal TRPV2 staining was decreased. Thus, dysregulation of TRPV channels also expressed by non-neuronal cells may be critically involved in the initiation and/or development of rosacea. TRP ion channels may be targets for the treatment of rosacea.

Distribution and Expression of Non-Neuronal Transient Receptor Potential (TRPV) Ion Channels in Rosacea

PubMed Central

Sulk, Mathias; Seeliger, Stephan; Aubert, Jerome; Schwab, Verena D.; Cevikbas, Ferda; Rivier, Michel; Nowak, Pawel; Voegel, Johannes J.; Buddenkotte, Jörg; Steinhoff, Martin

2011-01-01

Rosacea is a frequent chronic inflammatory skin disease of unknown etiology. Because early rosacea reveals all characteristics of neurogenic inflammation, a central role of sensory nerves in its pathophysiology has been discussed. Neuroinflammatory mediators and their receptors involved in rosacea are poorly defined. Good candidates may be transient receptor potential (TRP) ion channels of vanilloid type (TRPV), which can be activated by many trigger factors of rosacea. Interestingly, TRPV2, TRPV3, and TRPV4 are expressed by both neuronal and non-neuronal cells. Here, we analyzed the expression and distribution of TRPV receptors in the various subtypes of rosacea on non-neuronal cells using immunohistochemistry, morphometry, double immunoflourescence, and quantitative real-time PCR (qRT-PCR) as compared with healthy skin and lupus erythematosus. Our results show that dermal immunolabeling of TRPV2 and TRPV3 and gene expression of TRPV1 is significantly increased in erythematotelangiectatic rosacea (ETR). Papulopustular rosacea (PPR) displayed an enhanced immunoreactivity for TRPV2, TRPV4, and also of TRPV2 gene expression. In phymatous rosacea (PhR)-affected skin, dermal immunostaining of TRPV3 and TRPV4 and gene expression of TRPV1 and TRPV3 was enhanced, whereas epidermal TRPV2 staining was decreased. Thus, dysregulation of TRPV channels also expressed by non-neuronal cells may be critically involved in the initiation and/or development of rosacea. TRP ion channels may be targets for the treatment of rosacea. PMID:22189789

Regulation of vacuolar H{sup +}-ATPase in microglia by RANKL

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

Serrano, Eric M.; Ricofort, Ryan D.; Zuo, Jian

2009-11-06

Vacuolar H{sup +}-ATPases (V-ATPases) are large electrogenic proton pumps composed of numerous subunits that play vital housekeeping roles in the acidification of compartments of the endocytic pathway. Additionally, V-ATPases play specialized roles in certain cell types, a capacity that is linked to cell type selective expression of isoforms of some of the subunits. We detected low levels of the a3 isoform of the a-subunit in mouse brain extracts. Examination of various brain-derived cell types by immunoblotting showed a3 was expressed in the N9 microglia cell line and in primary microglia, but not in other cell types. The expression of a3more » in osteoclasts requires stimulation by Receptor Activator of Nuclear Factor {kappa}B-ligand (RANKL). We found that Receptor Activator of Nuclear Factor {kappa}B (RANK) was expressed by microglia. Stimulation of microglia with RANKL triggered increased expression of a3. V-ATPases in microglia were shown to bind microfilaments, and stimulation with RANKL increased the proportion of V-ATPase associated with the detergent-insoluble cytoskeletal fraction and with actin. In summary, microglia express the a3-subunit of V-ATPase. The expression of a3 and the interaction between V-ATPases and microfilaments was modulated by RANKL. These data suggest a novel molecular pathway for regulating microglia.« less

Peroxisome proliferator-activated receptor (PPAR)-gamma expression in human vascular smooth muscle cells: inhibition of growth, migration, and c-fos expression by the peroxisome proliferator-activated receptor (PPAR)-gamma activator troglitazone.

PubMed

Benson, S; Wu, J; Padmanabhan, S; Kurtz, T W; Pershadsingh, H A

2000-01-01

This study was conducted to determine whether cultured human coronary artery and aorta vascular smooth muscle (VSM) cells express the nuclear transcription factor peroxisome proliferator-activated receptor-gamma (PPARgamma); whether the thiazolidinedione troglitazone, a ligand for PPARgamma, would inhibit c-fos expression by these cells; and whether troglitazone would inhibit proliferation and migration induced in these cells by mitogenic growth factors. Using immunoblotting and reverse-transcriptase polymerase chain reaction (RT-PCR) techniques, we show that both human aorta and coronary artery VSM cell lines expressed PPARgamma protein and mRNA for both PPARgamma isoforms, PPARgamma1 and PPARgamma2. Immunocytochemical staining localized the PPARgamma protein primarily within the nucleus. Troglitazone inhibited basic fibroblast growth factor and platelet-derived growth factor-BB induced DNA synthesis in a dose-dependent manner and downregulated the growth-factor-induced expression of c-fos. Troglitazone also inhibited the migration of coronary artery VSM cells along a platelet-derived growth factor-BB concentration gradient. These findings demonstrate for the first time the expression and nuclear localization of PPARgamma in human coronary artery and aorta VSM cells. The data also suggest that the downregulation of c-fos expression, growth-factor-induced proliferation, and migration by VSM may, in part, be mediated by activation of the PPARgamma receptor.

Biochemical characterization of individual human glycosylated pro-insulin-like growth factor (IGF)-II and big-IGF-II isoforms associated with cancer.

PubMed

Greenall, Sameer A; Bentley, John D; Pearce, Lesley A; Scoble, Judith A; Sparrow, Lindsay G; Bartone, Nicola A; Xiao, Xiaowen; Baxter, Robert C; Cosgrove, Leah J; Adams, Timothy E

2013-01-04

Insulin-like growth factor II (IGF-II) is a major embryonic growth factor belonging to the insulin-like growth factor family, which includes insulin and IGF-I. Its expression in humans is tightly controlled by maternal imprinting, a genetic restraint that is lost in many cancers, resulting in up-regulation of both mature IGF-II mRNA and protein expression. Additionally, increased expression of several longer isoforms of IGF-II, termed "pro" and "big" IGF-II, has been observed. To date, it is ambiguous as to what role these IGF-II isoforms have in initiating and sustaining tumorigenesis and whether they are bioavailable. We have expressed each individual IGF-II isoform in their proper O-glycosylated format and established that all bind to the IGF-I receptor and both insulin receptors A and B, resulting in their activation and subsequent stimulation of fibroblast proliferation. We also confirmed that all isoforms are able to be sequestered into binary complexes with several IGF-binding proteins (IGFBP-2, IGFBP-3, and IGFBP-5). In contrast to this, ternary complex formation with IGFBP-3 or IGFBP-5 and the auxillary protein, acid labile subunit, was severely diminished. Furthermore, big-IGF-II isoforms bound much more weakly to purified ectodomain of the natural IGF-II scavenging receptor, IGF-IIR. IGF-II isoforms thus possess unique biological properties that may enable them to escape normal sequestration avenues and remain bioavailable in vivo to sustain oncogenic signaling.

Differential expression of appetite-regulating genes in avian models of anorexia and obesity.

PubMed

Yi, J; Yuan, J; Gilbert, E R; Siegel, P B; Cline, M A

2017-08-01

Chickens from lines that have been selected for low (LWS) or high (HWS) juvenile body weight for more than 57 generations provide a unique model by which to research appetite regulation. The LWS display different severities of anorexia, whereas all HWS become obese. In the present study, we measured mRNA abundance of various factors in appetite-associated nuclei in the hypothalamus. The lateral hypothalamus (LHA), paraventricular nucleus (PVN), ventromedial hypothalamus (VMH), dorsomedial nucleus (DMN) and arcuate nucleus (ARC) were collected from 5 day-old chicks that were fasted for 180 minutes or provided with continuous access to food. Fasting increased neuropeptide Y receptor subtype 1 (NPYR1) mRNA in the LHA and c-Fos in the VMH, at the same time as decreasing c-Fos in the LHA, neuropeptide Y receptor subtype 5 and ghrelin in the PVN, and neuropeptide Y receptor subtype 2 in the ARC. Fasting increased melanocortin receptor subtype 3 (MC3R) expression in the DMN and NPY in the ARC of LWS but not HWS chicks. Expression of NPY was greater in LWS than HWS in the DMN. neuropeptide Y receptor subtype 5 mRNA was greater in LWS than HWS in the LHA, PVN and ARC. Expression of orexin was greater in LWS than HWS in the LHA. There was greater expression of NPYR1, melanocortin receptor subtype 4 and cocaine- and amphetamine-regulated transcript in HWS than LWS and mesotocin in LWS than HWS in the PVN. In the ARC, agouti-related peptide and MC3R were greater in LWS than HWS and, in the VMH, orexin receptor 2 and leptin receptor were greater in LWS than HWS. Greater mesotocin in the PVN, orexin in the LHA and ORXR2 in the VMH of LWS may contribute to their increased sympathetic tone and anorexic phenotype. The results of the present study also suggest that an increased hypothalamic anorexigenic tone in the LWS over-rides orexigenic factors such as NPY and AgRP that were more highly expressed in LWS than HWS in several nuclei. Published 2017. This article is a U.S. Government work and is in the public domain in the USA.

The effect of very-low-calorie diet on mRNA expression of inflammation-related genes in subcutaneous adipose tissue and peripheral monocytes of obese patients with type 2 diabetes mellitus.

PubMed

Mraz, M; Lacinova, Z; Drapalova, J; Haluzikova, D; Horinek, A; Matoulek, M; Trachta, P; Kavalkova, P; Svacina, S; Haluzik, M

2011-04-01

Low-grade inflammation links obesity, type 2 diabetes mellitus (T2DM), and cardiovascular diseases. To explore the expression profile of genes involved in inflammatory pathways in adipose tissue and peripheral monocytes (PM) of obese patients with and without T2DM at baseline and after dietary intervention. Two-week intervention study with very-low-calorie diet (VLCD). University hospital. Twelve obese females with T2DM, 8 obese nondiabetic females (OB) and 15 healthy age-matched females. Two weeks of VLCD (2500 kJ/d). Metabolic parameters, circulating cytokines, hormones, and mRNA expression of 39 genes in sc adipose tissue (SCAT) and PM. Both T2DM and OB group had significantly increased serum concentrations of circulating proinflammatory factors (C-reactive protein, TNFα, IL-6, IL-8), mRNA expression of macrophage antigen CD68 and proinflammatory chemokines (CCL-2, -3, -7, -8, -17, -22) in SCAT and complementary chemokine receptors (CCR-1, -2, -3, -5) and other proinflammatory receptors (toll-like receptor 2 and 4, TNF receptor superfamily 1A and 1B, IL-6R) in PM, with OB group showing less pronounced chemoattracting and proinflammatory profile compared to T2DM group. In T2DM patients VLCD decreased body weight, improved metabolic profile, and decreased mRNA expression of up-regulated CCRs in PM and chemokines [CCL 8, chemokine (C-X-C motif) ligand 10] in SCAT. VLCD markedly increased mRNA expression of T-lymphocyte attracting chemokine CCL-17 in SCAT. Obese patients with and without T2DM have increased mRNA expression of chemotactic and proinflammatory factors in SCAT and expression of corresponding receptors in PM. Two weeks of VLCD significantly improved this profile in T2DM patients.

Keratin 17 is overexpressed and predicts poor survival in estrogen receptor-negative/human epidermal growth factor receptor-2-negative breast cancer.

PubMed

Merkin, Ross D; Vanner, Elizabeth A; Romeiser, Jamie L; Shroyer, A Laurie W; Escobar-Hoyos, Luisa F; Li, Jinyu; Powers, Robert S; Burke, Stephanie; Shroyer, Kenneth R

2017-04-01

Clinicopathological features of breast cancer have limited accuracy to predict survival. By immunohistochemistry (IHC), keratin 17 (K17) expression has been correlated with triple-negative status (estrogen receptor [ER]/progesterone receptor/human epidermal growth factor receptor-2 [HER2] negative) and decreased survival, but K17 messenger RNA (mRNA) expression has not been evaluated in breast cancer. K17 is a potential prognostic cancer biomarker, targeting p27, and driving cell cycle progression. This study compared K17 protein and mRNA expression to ER/progesterone receptor/HER2 receptor status and event-free survival. K17 IHC was performed on 164 invasive breast cancers and K17 mRNA was evaluated in 1097 breast cancers. The mRNA status of other keratins (16/14/9) was evaluated in 113 ER - /HER2 - ductal carcinomas. IHC demonstrated intense cytoplasmic and membranous K17 localization in myoepithelial cells of benign ducts and lobules and tumor cells of ductal carcinoma in situ. In ductal carcinomas, K17 protein was detected in most triple-negative tumors (28/34, 82%), some non-triple-negative tumors (52/112, 46%), but never in lobular carcinomas (0/15). In ductal carcinomas, high K17 mRNA was associated with reduced 5-year event-free survival in advanced tumor stage (n = 149, hazard ratio [HR] = 3.68, P = .018), and large (n = 73, HR = 3.95, P = .047), triple-negative (n = 103, HR = 2.73, P = .073), and ER - /HER2 - (n = 113, HR = 2.99, P = .049) tumors. There were significant correlations among keratins 17, 16, 14, and 9 mRNA levels suggesting these keratins (all encoded on chromosome 17) could be coordinately expressed in breast cancer. Thus, K17 is expressed in a subset of triple-negative breast cancers, and is a marker of poor prognosis in patients with advanced stage and ER - /HER2 - breast cancer. Copyright © 2016 Elsevier Inc. All rights reserved.

Glucocorticoid receptor is involved in the differential expression of hepatic 3β-hydroxysteroid dehydrogenase between barrows and boars at finishing stage.

PubMed

Li, Xian; Cong, Rihua; Yao, Wen; Jia, Yimin; Li, Runsheng; Sun, Zhiyuan; Li, Xi; Zhao, Ruqian

2018-01-01

The enzyme 3β-hydroxysteroid dehydrogenase (3β-HSD) plays an important role in androstenone metabolism in pig liver, and its defective expression is related to the development of boar taint. Early age castration is a common practice in many countries to avoid boar taint, yet whether and how castration affects porcine hepatic 3β-HSD expression are still poorly understood. In this study, we aimed to compare the expression of 3β-HSD between intact (boars) and castrated (barrows) male pigs, and to explore the potential factors regulating 3β-HSD transcription. Compared to barrows, boars showed worse carcass quality. Boars had significantly higher levels of serum androstenone (P < 0.01), testosterone (P < 0.01) and hepatic cortisol (P < 0.05), which were contrary to significantly lower expression of 3β-HSD messenger RNA (P < 0.01) and protein (P < 0.01) in the liver. Significant differences were detected for the hepatic expression of androgen receptor (AR) and CCAAT/enhancer binding protein β (C/EBPβ). Chromatin immunoprecipitation (ChIP) assay demonstrated reduced histone H3 acetylation (P < 0.05) but increased glucocorticoid receptor (GR) binding to 3β-HSD gene promoter in boars (P < 0.05). These results indicate that GR binding to 3β-HSD promoter is involved in the differential hepatic 3β-HSD expression between boars and barrows. © 2017 Japanese Society of Animal Science.

17β-estradiol-induced regulation of the novel 5-HT1A-related transcription factors NUDR and Freud-1 in SH SY5Y cells.

PubMed

Adeosun, Samuel O; Albert, Paul R; Austin, Mark C; Iyo, Abiye H

2012-05-01

Nuclear deformed epidermal autoregulatory factor-1 (NUDR/Deaf-1) and five prime repressor element under dual repression (Freud-1) are novel transcriptional regulators of the 5-HT(1A) receptor, a receptor that has been implicated in the pathophysiology of various psychiatric illnesses. The antidepressant effect of 17β-Estradiol (17βE(2)) is purported to involve the downregulation of this receptor. We investigated the possible role of NUDR and Freud-1 in 17βE(2)-induced downregulation of the 5-HT(1A) receptor in the neuroblastoma cell line SH SY5Y. Cells were treated with 10 nM of 17βE(2) for 3 or 48 h, followed by a 24-h withdrawal period. Proteins were isolated and analyzed by western blotting. 17βE(2) treatment increased NUDR immunoreactivity while Freud-1 and the 5-HT(1A) receptor showed significant decreases. Upon withdrawal of 17βE(2), protein expression returned to control levels, except for NUDR, which remained significantly elevated in the 3-h treatment. Taken together, these data support a non-genomic downregulation of 5-HT(1A) receptor protein by 17βE(2), which does not involve NUDR and Freud-1. Rather, changes in both transcription factors seem to be compensatory/homeostatic responses to changes in 5-HT(1A) receptor induced by 17βE(2). These observations further highlight the importance of NUDR and Freud-1 in regulating 5-HT(1A) receptor expression.

Mitogenic signals and transforming potential of Nyk, a newly identified neural cell adhesion molecule-related receptor tyrosine kinase.

PubMed Central

Ling, L; Kung, H J

1995-01-01

Nyk/Mer is a recently identified receptor tyrosine kinase with neural cell adhesion molecule-like structure (two immunoglobulin G-like domains and two fibronectin III-like domains) in its extracellular region and belongs to the Ufo/Axl family of receptors. The ligand for Nyk/Mer is presently unknown, as are the signal transduction pathways mediated by this receptor. We constructed and expressed a chimeric receptor (Fms-Nyk) composed of the extracellular domain of the human colony-stimulating factor 1 receptor (Fms) and the transmembrane and cytoplasmic domains of human Nyk/Mer in NIH 3T3 fibroblasts in order to investigate the mitogenic signaling and biochemical properties of Nyk/Mer. Colony-stimulating factor 1 stimulation of the Fms-Nyk chimeric receptor in transfected NIH 3T3 fibroblasts leads to a transformed phenotype and generates a proliferative response in the absence of other growth factors. We show that phospholipase C gamma, phosphatidylinositol 3-kinase/p70 S6 kinase, Shc, Grb2, Raf-1, and mitogen-activated protein kinase are downstream components of the Nyk/Mer signal transduction pathways. In addition, Nyk/Mer weakly activates p90rsk, while stress-activated protein kinase, Ras GTPase-activating protein (GAP), and GAP-associated p62 and p190 proteins are not activated or tyrosine phosphorylated by Nyk/Mer. An analysis comparing the Nyk/Mer signal cascade with that of the epidermal growth factor receptor indicates substrate preferences by these two receptors. Our results provide a detailed description of the Nyk/Mer signaling pathways. Given the structural similarity between the Ufo/Axl family receptors, some of the information may also be applied to other members of this receptor tyrosine kinase family. PMID:8524223

Stimulation by thyroid-stimulating hormone and Grave's immunoglobulin G of vascular endothelial growth factor mRNA expression in human thyroid follicles in vitro and flt mRNA expression in the rat thyroid in vivo.

PubMed

Sato, K; Yamazaki, K; Shizume, K; Kanaji, Y; Obara, T; Ohsumi, K; Demura, H; Yamaguchi, S; Shibuya, M

1995-09-01

To elucidate the pathogenesis of thyroid gland hypervascularity in patients with Graves' disease, we studied the expression of mRNAs for vascular endothelial growth factor (VEGF) and its receptor, Flt family, using human thyroid follicles in vitro and thiouracil-fed rats in vivo. Human thyroid follicles, cultured in the absence of endothelial cells, secreted de novo-synthesized thyroid hormone in response to thyroid-stimulating hormone (TSH) and Graves' IgG. The thyroid follicles produced VEGF mRNA but not flt-1 mRNA. The expression of VEGF mRNA was enhanced by insulin, tumor-promoting phorbol ester, calcium ionophore, dibutyryl cAMP, TSH, and Graves' IgG. When rats were fed thiouracil for 4 wk, their serum levels of TSH were increased at day 3. VEGF mRNA was also increased on day 3, accompanied by an increase in flt family (flt-1 and KDR/ flk-1) mRNA expression. These in vitro and in vivo findings suggest that VEGF is produced by thyroid follicles in response to stimulators of TSH receptors, via the protein kinase A and C pathways. VEGF, a secretable angiogenesis factor, subsequently stimulates Flt receptors on endothelial cells in a paracrine manner, leading to their proliferation and producing hypervascularity of the thyroid gland, as seen in patients with Graves' disease.

Gene expression of tumour necrosis factor and insulin signalling-related factors in subcutaneous adipose tissue during the dry period and in early lactation in dairy cows.

PubMed

Sadri, H; Bruckmaier, R M; Rahmani, H R; Ghorbani, G R; Morel, I; van Dorland, H A

2010-10-01

Gene expression of adipose factors, which may be part of the mechanisms that underlie insulin sensitivity, were studied in dairy cows around parturition. Subcutaneous fat biopsies and blood samples were taken from 27 dairy cows in week 8 antepartum (a.p.), on day 1 postpartum (p.p.) and in week 5 p.p. In the adipose tissue samples, mRNA was quantified by real-time reverse transcription polymerase chain reaction for tumour necrosis factor alpha (TNFα), insulin-independent glucose transporter (GLUT1), insulin-responsive glucose transporter (GLUT4), insulin receptor, insulin receptor substrate 1 (IRS1), insulin receptor substrate 2 (IRS2), regulatory subunit of phosphatidylinositol-3 kinase (p85) and catalytic subunit of phosphatidylinositol-3 kinase. Blood plasma was assayed for concentrations of glucose, β-hydroxybutyric acid, non-esterified fatty acids (NEFA) and insulin. Plasma parameters followed a pattern typically observed in dairy cows. Gene expression changes were observed, but there were no changes in TNFα concentrations, which may indicate its local involvement in catabolic adaptation of adipose tissue. Changes in GLUT4 and GLUT1 mRNA abundance may reflect their involvement in reduced insulin sensitivity and in sparing glucose for milk synthesis in early lactation. Unchanged gene expression of IRS1, IRS2 and p85 over time may imply a lack of their involvement in terms of insulin sensitivity dynamics. Alternatively, it may indicate that post-transcriptional modifications of these factors came into play and may have concealed an involvement. © 2010 Blackwell Verlag GmbH.

VEGF and Ki-67 Overexpression in Predicting Poor Overall Survival in Adenoid Cystic Carcinoma.

PubMed

Park, Seongyeol; Nam, Soo Jeong; Keam, Bhumsuk; Kim, Tae Min; Jeon, Yoon Kyung; Lee, Se-Hoon; Hah, J Hun; Kwon, Tack-Kyun; Kim, Dong-Wan; Sung, Myung-Whun; Heo, Dae Seog; Bang, Yung-Jue

2016-04-01

The purpose of this study was to evaluate potential prognostic factors in patients with adenoid cystic carcinoma (ACC). A total of 68 patients who underwent curative surgery and had available tissue were enrolled in this study. Their medical records and pathologic slides were reviewed and immunohistochemistry for basic fibroblast growth factor, fibroblast growth factor receptor (FGFR) 2, FGFR3, c-kit, Myb proto-oncogene protein, platelet-derived growth factor receptor beta, vascular endothelial growth factor (VEGF), and Ki-67 was performed. Univariate and multivariate analysis was performed for determination of disease-free survival (DFS) and overall survival (OS). In univariate analyses, primary site of nasal cavity and paranasal sinus (p=0.022) and Ki-67 expression of more than 7% (p=0.001) were statistically significant factors for poor DFS. Regarding OS, perineural invasion (p=0.032), high expression of VEGF (p=0.033), and high expression of Ki-67 (p=0.007) were poor prognostic factors. In multivariate analyses, primary site of nasal cavity and paranasal sinus (p=0.028) and high expression of Ki-67 (p=0.004) were independent risk factors for poor DFS, and high expression of VEGF (p=0.011) and Ki-67 (p=0.011) showed independent association with poor OS. High expression of VEGF and Ki-67 were independent poor prognostic factors for OS in ACC.

A Transcriptional Regulatory Network Containing Nuclear Receptors and Long Noncoding RNAs Controls Basal and Drug-Induced Expression of Cytochrome P450s in HepaRG Cells.

PubMed

Chen, Liming; Bao, Yifan; Piekos, Stephanie C; Zhu, Kexin; Zhang, Lirong; Zhong, Xiao-Bo

2018-07-01

Cytochrome P450 (P450) enzymes are responsible for metabolizing drugs. Expression of P450s can directly affect drug metabolism, resulting in various outcomes in therapeutic efficacy and adverse effects. Several nuclear receptors are transcription factors that can regulate expression of P450s at both basal and drug-induced levels. Some long noncoding RNAs (lncRNAs) near a transcription factor are found to participate in the regulatory functions of the transcription factors. The aim of this study is to determine whether there is a transcriptional regulatory network containing nuclear receptors and lncRNAs controlling both basal and drug-induced expression of P450s in HepaRG cells. Small interfering RNAs or small hairpin RNAs were applied to knock down four nuclear receptors [hepatocyte nuclear factor 1 α (HNF1 α ), hepatocyte nuclear factor 4 α (HNF4 α ), pregnane X receptor (PXR), and constitutive androstane receptor (CAR)] as well as two lncRNAs [HNF1 α antisense RNA 1 (HNF1 α -AS1) and HNF4 α antisense RNA 1 (HNF4 α -AS1)] in HepaRG cells with or without treatment of phenobarbital or rifampicin. Expression of eight P450 enzymes was examined in both basal and drug-induced levels. CAR and PXR mainly regulated expression of specific P450s. HNF1 α and HNF4 α affected expression of a wide range of P450s as well as other transcription factors. HNF1 α and HNF4 α controlled the expression of their neighborhood lncRNAs, HNF1 α -AS1 and HNF4 α -AS1, respectively. HNF1 α -AS1 and HNF4 α -AS1 was also involved in the regulation of P450s and transcription factors in diverse manners. Altogether, our study concludes that a transcription regulatory network containing the nuclear receptors and lncRNAs controls both basal and drug-induced expression of P450s in HepaRG cells. Copyright © 2018 by The American Society for Pharmacology and Experimental Therapeutics.

Transcriptional response to muscarinic acetylcholine receptor stimulation: regulation of Egr-1 biosynthesis by ERK, Elk-1, MKP-1, and calcineurin in carbachol-stimulated human neuroblastoma cells.

PubMed

Rössler, Oliver G; Henss, Isabell; Thiel, Gerald

2008-02-01

Carbachol-mediated activation of type M(3) muscarinic acetylcholine receptors induces the biosynthesis of the transcription factor Egr-1 in human SH-SY5Y neuroblastoma cells involving an activation of extracellular signal-regulated protein kinase. Carbachol triggered the phosphorylation of the ternary complex factor Elk-1, a key transcriptional regulator of serum response element-driven gene transcription, and strikingly enhanced the transcriptional activation potential of Elk-1. Chromatin immunoprecipitation experiments revealed that Elk-1 binds in vivo to the 5'-upstream region of the Egr-1 gene in carbachol-stimulated neuroblastoma cells. Together, these data indicate that Elk-1 connects the intracellular signaling cascade elicited by activation of M(3) muscarinic acetylcholine receptors with the transcription of the Egr-1 gene. Lentiviral-mediated expression of either MAP kinase phosphatase-1 (MKP-1) or a constitutively active mutant of calcineurin A inhibited Egr-1 biosynthesis following carbachol stimulation, indicating that these phosphatases function as shut-off devices of muscarinic acetylcholine receptor signaling. Additionally, carbachol stimulation increased transcription of a chromatin-embedded collagenase promoter/reporter gene, showing that AP-1 activity is enhanced in carbachol-stimulated neuroblastoma. Expression experiments revealed that both MKP-1 and a constitutively active mutant of calcineurin A impaired carbachol-induced upregulation of AP-1 activity. The fact that carbachol stimulation of neuroblastoma cells activates the transcription factors Egr-1 and AP-1 suggests that changes in the gene expression pattern are an integral part of muscarinic acetylcholine receptor signaling.

Neomycin is a platelet-derived growth factor (PDGF) antagonist that allows discrimination of PDGF alpha- and beta-receptor signals in cells expressing both receptor types.

PubMed

Vassbotn, F S; Ostman, A; Siegbahn, A; Holmsen, H; Heldin, C H

1992-08-05

The aminoglycoside neomycin has recently been found to affect certain platelet-derived growth factor (PDGF) responses in C3H/10T1/2 C18 fibroblasts. Using porcine aortic endothelial cells transfected with PDGF alpha- or beta-receptors, we explored the possibility that neomycin interferes with the interaction between the different PDGF isoforms and their receptors. We found that neomycin (5 mM) inhibited the binding of 125I-PDGF-BB to the alpha-receptor with only partial effect on the binding of 125I-PDGF-AA; in contrast, the binding of 125I-PDGF-BB to the beta-receptor was not affected by the aminoglycoside. Scatchard analyses showed that neomycin (5 mM) decreased the number of binding sites for PDGF-BB on alpha-receptor-expressing cells by 87%. Together with cross-competition studies with 125I-labeled PDGF homodimers, the effect of neomycin indicates that PDGF-AA and PDGF-BB bind to both common and unique structures on the PDGF alpha-receptor. Neomycin specifically inhibited the autophosphorylation of the alpha-receptor by PDGF-BB, with less effect on the phosphorylation induced by PDGF-AA and no effect on the phosphorylation of the beta-receptor by PDGF-BB. Thus, neomycin is a PDGF isoform- and receptor-specific antagonist that provides a possibility to compare the signal transduction pathways of alpha- and beta-receptors in cells expressing both receptor types. This approach was used to show that activation of PDGF beta-receptors by PDGF-BB mediated a chemotactic response in human fibroblasts, whereas activation of alpha-receptors by the same ligand inhibited chemotaxis.

Suppression of transient receptor potential melastatin 4 expression promotes conversion of endothelial cells into fibroblasts via transforming growth factor/activin receptor-like kinase 5 pathway.

PubMed

Echeverría, Cesar; Montorfano, Ignacio; Cabello-Verrugio, Claudio; Armisén, Ricardo; Varela, Diego; Simon, Felipe

2015-05-01

To study whether transient receptor potential melastatin 4 (TRPM4) participates in endothelial fibrosis and to investigate the underlying mechanism. Primary human endothelial cells were used and pharmacological and short interfering RNA-based approaches were used to test the transforming growth factor beta (TGF-β)/activin receptor-like kinase 5 (ALK5) pathway participation and contribution of TRPM7 ion channel. Suppression of TRPM4 expression leads to decreased endothelial protein expression and increased expression of fibrotic and extracellular matrix markers. Furthermore, TRPM4 downregulation increases intracellular Ca levels as a potential condition for fibrosis. The underlying mechanism of endothelial fibrosis shows that inhibition of TRPM4 expression induces TGF-β1 and TGF-β2 expression, which act through their receptor, ALK5, and the nuclear translocation of the profibrotic transcription factor smad4. TRPM4 acts to maintain endothelial features and its loss promotes fibrotic conversion via TGF-β production. The regulation of TRPM4 levels could be a target for preserving endothelial function during inflammatory diseases.

Screening a mouse liver gene expression Compendium Identifies Effectors of the Aryl Hydrocarbon receptor (AhR)

EPA Science Inventory

The aryl hydrocarbon receptor (AhR) is a ligand-activated transcription factor that mediates the biological and toxic effects of 2,3, 7 ,8-tetrachlorodibenzo-p-dioxin {TCDD), dioxin-like compounds (DLC) as well as some drugs and endogenous tryptophan metabolites. Short-term act...

Two signaling molecules share a phosphotyrosine-containing binding site in the platelet-derived growth factor receptor.

PubMed

Nishimura, R; Li, W; Kashishian, A; Mondino, A; Zhou, M; Cooper, J; Schlessinger, J

1993-11-01

Autophosphorylation sites of growth factor receptors with tyrosine kinase activity function as specific binding sites for Src homology 2 (SH2) domains of signaling molecules. This interaction appears to be a crucial step in a mechanism by which receptor tyrosine kinases relay signals to downstream signaling pathways. Nck is a widely expressed protein consisting exclusively of SH2 and SH3 domains, the overexpression of which causes cell transformation. It has been shown that various growth factors stimulate the phosphorylation of Nck and its association with autophosphorylated growth factor receptors. A panel of platelet-derived growth factor (PDGF) receptor mutations at tyrosine residues has been used to identify the Nck binding site. Here we show that mutation at Tyr-751 of the PDGF beta-receptor eliminates Nck binding both in vitro and in living cells. Moreover, the Y751F PDGF receptor mutant failed to mediate PDGF-stimulated phosphorylation of Nck in intact cells. A phosphorylated Tyr-751 is also required for binding of phosphatidylinositol-3 kinase to the PDGF receptor. Hence, the SH2 domains of p85 and Nck share a binding site in the PDGF receptor. Competition experiments with different phosphopeptides derived from the PDGF receptor suggest that binding of Nck and p85 is influenced by different residues around Tyr-751. Thus, a single tyrosine autophosphorylation site is able to link the PDGF receptor to two distinct SH2 domain-containing signaling molecules.

Neuropeptides activate human mast cell degranulation and chemokine production

PubMed Central

Kulka, Marianna; Sheen, Cecilia H; Tancowny, Brian P; Grammer, Leslie C; Schleimer, Robert P

2008-01-01

During neuronal-induced inflammation, mast cells may respond to stimuli such as neuropeptides in an FcεRI-independent manner. In this study, we characterized human mast cell responses to substance P (SP), nerve growth factor (NGF), calcitonin gene-related peptide (CGRP) and vasoactive intestinal polypeptide (VIP) and compared these responses to human mast cell responses to immunoglobulin E (IgE)/anti-IgE and compound 48/80. Primary cultured mast cells, generated from CD34+ progenitors in the presence of stem cell factor and interleukin-6 (IL-6), and human cultured mast cells (LAD2) were stimulated with these and other stimuli (gastrin, concanavalin A, radiocontrast media, and mannitol) and their degranulation and chemokine production was assessed. VIP and SP stimulated primary human mast cells and LAD cells to degranulate; gastrin, concanavalin A, radiocontrast media, mannitol, CGRP and NGF did not activate degranulation. While anti-IgE stimulation did not induce significant production of chemokines, stimulation with VIP, SP or compound 48/80 potently induced production of monocyte chemoattractant protein-1, inducible protein-10, monokine induced by interferon-γ (MIG), RANTES (regulated on activation, normal, T-cell expressed, and secreted) and IL-8. VIP, SP and compound 48/80 also activated release of tumour necrosis factor, IL-3 and granulocyte–macrophage colony-stimulating factor, but not IL-4, interferon-γ or eotaxin. Human mast cells expressed surface neurokinin 1 receptor (NK1R), NK2R, NK3R and VIP receptor type 2 (VPAC2) but not VPAC1 and activation of human mast cells by IgE/anti-IgE up-regulated expression of VPAC2, NK2R, and NK3R. These studies demonstrate the pattern of receptor expression and activation of mast cell by a host of G-protein coupled receptor ligands and suggest that SP and VIP activate a unique signalling pathway in human mast cells. These results are likely to have direct relevance to neuronally induced inflammatory diseases. PMID:17922833

Hypotonic stress induces RANKL via transient receptor potential melastatin 3 (TRPM3) and vaniloid 4 (TRPV4) in human PDL cells.

PubMed

Son, G Y; Yang, Y M; Park, W S; Chang, I; Shin, D M

2015-03-01

Bone remodeling occurs in response to various types of mechanical stress. The periodontal ligament (PDL) plays an important role in mechanical stress-mediated alveolar bone remodeling. However, the underlying mechanism at the cellular level has not been extensively studied. In this study, we investigated the effect of shear stress on the expression of bone remodeling factors, including receptor activator of nuclear factor-kappa B (NF-κB) ligand (RANKL) and osteoprotegerin (OPG), as well as its upstream signaling pathway in primary human PDL cells. We applied hypotonic stress to reproduce shear stress to PDL cells. Hypotonic stress induced the messenger RNA (mRNA) and protein expression of RANKL but not OPG. It also increased intracellular Ca(2+) concentration ([Ca(2+)]i). Extracellular Ca(2+) depletion and nonspecific plasma membrane Ca(2+) channel blockers completely inhibited the increase in both [Ca(2+)]i and RANKL mRNA expression. We identified the expression and activation of transient receptor potential melastatin 3 (TRPM3) and vaniloid 4 (TRPV4) channels in PDL cells. Pregnenolone sulfate (PS) and 4α-phorbol 12, 13-didecanoate (4α-PDD), which are agonists of TRPM3 and TRPV4, augmented Ca(2+) influx and RANKL mRNA expression. Both pharmacological (2-aminoethoxydiphenyl borate [2-APB], ruthenium red [RR], ononetin [Ono], and HC 067047 [HC]) and genetic (small interfering RNA [siRNA]) inhibitors of TRPM3 and TRPV4 reduced the hypotonic stress-mediated increase in [Ca(2+)]i and RANKL mRNA expression. Our study shows that hypotonic stress induced RANKL mRNA expression via TRPM3- and TRPV4-mediated extracellular Ca(2+) influx and RANKL expression. This signaling pathway in PDL cells may play a critical role in mechanical stress-mediated alveolar bone remodeling. © International & American Associations for Dental Research 2015.

Expression of Functional Interleukin-3 Receptors on Hodgkin and Reed-Sternberg Cells

PubMed Central

Aldinucci, Donatella; Poletto, Dalisa; Gloghini, Annunziata; Nanni, Paola; Degan, Massimo; Perin, Tiziana; Ceolin, Paola; Rossi, Francesca Maria; Gattei, Valter; Carbone, Antonino; Pinto, Antonio

2002-01-01

The human interleukin-3 receptor (IL-3R) is a heterodimeric complex consisting of an IL-3-specific α chain (IL-3Rα) and a common β chain (βc), this latter shared with the receptors for granulocyte-macrophage colony-stimulating factor and IL-5. Despite extensive research on cytokine circuitries regulating proliferation and survival of tumor cells in Hodgkin’s disease (HD) the functional expression of IL-3Rs in this pathobiological entity has not yet been investigated. In the present study, we demonstrate that the great majority (>90%) of malignant Hodgkin and Reed-Sternberg cells of classic HD (19 of 19 analyzed cases) express IL-3Rα by immunostaining of frozen sections and cell suspensions from involved lymph nodes. Accordingly, HD cell lines (L428, KMH2, HDLM2, L1236) expressed the α and β chains of IL-3R both at the mRNA and protein level, with a molecular size of IL-3Rα identical (70 kd) to that expressed by human myeloid cells. Exogenous IL-3 promoted the growth of cultured Hodgkin and Reed-Sternberg cells, such effect being potentiated by IL-9 co-stimulation, and was able to partially rescue tumor cells from apoptosis induced by serum deprivation. This data suggests an involvement of IL-3/IL-3R interactions in the cellular growth of HD through paracrine mechanisms. PMID:11839579

Decoy receptor 3: an endogenous immunomodulator in cancer growth and inflammatory reactions.

PubMed

Hsieh, Shie-Liang; Lin, Wan-Wan

2017-06-19

Decoy receptor 3 (DcR3), also known as tumor necrosis factor receptor (TNFR) superfamily member 6b (TNFRSF6B), is a soluble decoy receptor which can neutralize the biological functions of three members of tumor necrosis factor superfamily (TNFSF): Fas ligand (FasL), LIGHT, and TL1A. In addition to 'decoy' function, recombinant DcR3.Fc is able to modulate the activation and differentiation of dendritic cells (DCs) and macrophages via 'non-decoy' action. DcR3-treated DCs skew T cell differentiation into Th2 phenotype, while DcR3-treated macrophages behave M2 phenotype. DcR3 is upregulated in various cancer cells and several inflammatory tissues, and is regarded as a potential biomarker to predict inflammatory disease progression and cancer metastasis. However, whether DcR3 is a pathogenic factor or a suppressor to attenuate inflammatory reactions, has not been discussed comprehensively yet. Because mouse genome does not have DcR3, it is not feasible to investigate its physiological functions by gene-knockout approach. However, DcR3-mediated effects in vitro are determined via overexpressing DcR3 or addition of recombinant DcR3.Fc fusion protein. Moreover, CD68-driven DcR3 transgenic mice are used to investigate DcR3-mediated systemic effects in vivo. Upregulation of DcR3 during inflammatory reactions exerts negative-feedback to suppress inflammation, while tumor cells hijack DcR3 to prevent apoptosis and promote tumor growth and invasion. Thus, 'switch-on' of DcR3 expression may be feasible for the treatment of inflammatory diseases and enhance tissue repairing, while 'switch-off' of DcR3 expression can enhance tumor apoptosis and suppress tumor growth in vivo.

Taste bud-derived BDNF maintains innervation of a subset of TrkB-expressing gustatory nerve fibers

PubMed Central

Tang, Tao; Rios-Pilier, Jennifer; Krimm, Robin

2018-01-01

Taste receptor cells transduce different types of taste stimuli and transmit this information to gustatory neurons that carry it to the brain. Taste receptor cells turn over continuously in adulthood, requiring constant new innervation from nerve fibers. Therefore, the maintenance of innervation to taste buds is an active process mediated by many factors, including brain-derived neurotrophic factor (BDNF). Specifically, 40% of taste bud innervation is lost when Bdnf is removed during adulthood. Here we speculated that not all gustatory nerve fibers express the BDNF receptor, TrkB, resulting in subsets of neurons that vary in their response to BDNF. However, it is also possible that the partial loss of innervation occurred because the Bdnf gene was not effectively removed. To test these possibilities, we first determined that not all gustatory nerve fibers express the TrkB receptor in adult mice. We then verified the efficiency of Bdnf removal specifically in taste buds of K14-CreER:Bdnf mice and found that Bdnf expression was reduced to 1%, indicating efficient Bdnf gene recombination. BDNF removal resulted in a 55% loss of TrkB-expressing nerve fibers, which was greater than the loss of P2X3-positive fibers (39%), likely because taste buds were innervated by P2X3+/TrkB− fibers that were unaffected by BDNF removal. We conclude that gustatory innervation consists of both TrkB-positive and TrkB-negative taste fibers and that BDNF is specifically important for maintaining TrkB-positive innervation to taste buds. In addition, although taste bud size was not affected by inducible Bdnf removal, the expression of the γ subunit of the ENaC channel was reduced. So, BDNF may regulate expression of some molecular components of taste transduction pathways. PMID:28600222

TRAIL Death Receptor-4 Expression Positively Correlates With the Tumor Grade in Breast Cancer Patients With Invasive Ductal Carcinoma

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

Sanlioglu, Ahter D.; Department of Medical Biology and Genetics, Akdeniz University Faculty of Medicine, Antalya; Korcum, Aylin F.

2007-11-01

Purpose: Tumor necrosis factor-related apoptosis inducing ligand (TRAIL) selectively induces apoptosis in cancer cells but not in normal cells, and a number of clinical trials have recently been initiated to test the safety and antitumoral potential of TRAIL in cancer patients. Four different receptors have been identified to interact with TRAIL: two are death-inducing receptors (TRAIL-R1 [DR4] and TRAIL-R2 [DR5]), whereas the other two (TRAIL-R3 [DcR1] and TRAIL-R4 [DcR2]) do not induce death upon ligation and are believed to counteract TRAIL-induced cytotoxicity. Because high levels of DcR2 expression have recently been correlated with carcinogenesis in the prostate and lung, thismore » study investigated the importance of TRAIL and TRAIL receptor expression in breast cancer patients with invasive ductal carcinoma, taking various prognostic markers into consideration. Methods and Materials: Immunohistochemical analyses were performed on 90 breast cancer patients with invasive ductal carcinoma using TRAIL and TRAIL receptor-specific antibodies. Age, menopausal status, tumor size, lymph node status, tumor grade, lymphovascular invasion, perineural invasion, extracapsular tumor extension, presence of an extensive intraductal component, multicentricity, estrogen and progesterone receptor status, and CerbB2 expression levels were analyzed with respect to TRAIL/TRAIL receptor expression patterns. Results: The highest TRAIL receptor expressed in patients with invasive ductal carcinoma was DR4. Although progesterone receptor-positive patients exhibited lower DR5 expression, CerbB2-positive tissues displayed higher levels of both DR5 and TRAIL expressions. Conclusions: DR4 expression positively correlates with the tumor grade in breast cancer patients with invasive ductal carcinoma.« less

Amplification of Genomic DNA for Decoy Receptor 3 Predicts Post-Resection Disease Recurrence in Breast Cancer Patients.

PubMed

Kanbayashi, Chizuko; Koyama, Yu; Ichikawa, Hiroshi; Sakata, Eiko; Hasegawa, Miki; Toshikawa, Chie; Manba, Naoko; Ikarashi, Mayuko; Kobayashi, Takashi; Minagawa, Masahiro; Kosugi, Shin-Ichi; Wakai, Toshifumi

2014-02-01

Decoy receptor 3 (DcR3), a member of the tumor necrosis factor receptor (TNFR) superfamily, shows inhibitory effects on Fas-mediated apoptosis. Currently, data are lacking on the correlation between DcR3 and the recurrence of breast cancer. The authors examined DcR3 mRNA expression and genomic amplification in breast cancer, and investigated the effect of DcR3 gene amplification on prognosis of patients. A total of 95 patients formed the basis of the current retrospective study. DcR3 mRNA expression in breast cancer tissues was examined by RNase protection assay and in situ hybridization. DcR3 gene amplification was examined by quantitative polymerase chain reaction. The correlation between DcR3 gene amplification status and clinicopathological factors was examined and also the relationship between DcR3-Amp and relapse and survival. The relative copy numbers of DcR3 genomic DNA correlated significantly with the levels of DcR3 mRNA expression (ρ = 0.755, P = 0.0067). In addition, lymphatic invasion correlated significantly with DcR3 gene amplification (P = 0.012). However, there was no correlation between the remaining clinicopathological factors and DcR3 gene amplification. In the univariate analysis, the recurrence-free survival (RFS) rate of patients who were positive for DcR3 gene amplification was significantly lower than that of patients who were negative for DcR3 gene amplification (P = 0.0271). Multivariate analysis showed that DcR3 gene amplification (P = 0.028) and disease stage (P < 0.001) remained significant independent predictors of RFS. DcR3 gene amplification was significantly correlated with lymphatic invasion, and also DcR3 gene amplification predicts recurrence after resection, which may be an important prognostic factor in breast cancer patients.

Vascular Endothelial Growth Factor and Angiopoietin are Required for Prostate Regeneration.

PubMed Central

Wang, Gui-min; Kovalenko, Bruce; Huang, Yili; Moscatelli, David

2007-01-01

BACKGROUND The regulation of the prostate size by androgens may be partly the result of androgen effects on the prostatic vasculature. We examined the effect of changes in androgen levels on the expression of a variety of angiogenic factors in the mouse prostate and determined if vascular endothelial growth factor (VEGF)-A and the angiopoietins are involved in the vascular response to androgens. METHODS Expression of angiogenic factors in prostate was quantitated using real-time PCR at different times after castration and after administration of testosterone to castrated mice. Angiopoietins were localized in prostate by immunohistochemistry and in situ hybridization. The roles of VEGF and the angiopoietins in regeneration of the prostate were examined in mice inoculated with cells expressing soluble VEGF receptor-2 or soluble Tie-2. RESULTS Castration resulted in a decrease in VEGF-A, VEGF-B, VEGF-C, placenta growth factor, FGF-2, and FGF-8 expression after one day. In contrast, VEGF-D mRNA levels increased. No changes in angiopoietin-1 (Ang-1), angiopoietin-2 (Ang-2), hepatocyte growth factor, VEGF receptor-1, VEGF receptor-2 or tie-2 mRNA levels were observed. Administration of testosterone to castrated mice had the opposite effect on expression of these angiogenic factors. Ang-2 was expressed predominately in prostate epithelial cells whereas Ang-1 was expressed in epithelium and smooth muscle. Inoculation of mice with cells expressing soluble VEGF receptor-2 or Tie-2 blocked the increase in vascular density normally observed after administration of testosterone to castrated mice. The soluble receptors also blocked the increase in prostate weight and proliferation of prostatic epithelial cells. CONCLUSION VEGF-A and angiopoietins are required for the vascular response to androgens and for the ability of the prostate to regenerate in response to androgens. PMID:17221843

Synergistic action of cisplatin and echistatin in MDA-MB-231 breast cancer cells.

PubMed

Czarnomysy, Robert; Surażyński, Arkadiusz; Popławska, Bożena; Rysiak, Edyta; Pawłowska, Natalia; Czajkowska, Anna; Bielawski, Krzysztof; Bielawska, Anna

2017-03-01

The aim of our study was to determine whether the use of cisplatin in the presence echistatin in MDA-MB-231 breast cancer cells leads to a reduction of toxic effects associated with the use of cisplatin. The expression of β 1 -integrin and insulin-like growth factor 1 receptor (IGF-IR), signaling pathway protein expression: protein kinase B (AKT), mitogen-activated protein kinases (ERK1/ERK2), nuclear factor kappa B (NFκB), and caspase-3 and -9 activity was measured after 24 h of incubation with tested compounds to explain detailed molecular mechanism of induction of apoptosis. The viability of MDA-MB-231 breast cancer cells was determined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. Annexin V-FITC/propidium iodide staining assay was performed to detect the induction of apoptosis. Inhibition DNA biosynthesis was determined by [ 3 H]thymidine incorporation into DNA. The expression of of β 1 -integrin, IGF-IR, AKT, ERK1/ERK2, NFκB, caspase-3 and -9 was evaluated using Western blot. The results suggest that treatment of MDA-MB-231 breast cancer cells for 24 h cisplatin plus echistatin severely inhibits cell growth and activates apoptosis by upregulation of caspase-3 and -9 expressions. The effect was stronger than treatment cisplatin and echistatin alone. In this study, we have found that cisplatin plus echistatin treatment decreases collagen biosynthesis in MDA-MB-231 breast cancer cells stronger than the individual compounds. The inhibition was found to be dependent on the β 1 -integrin and IGF receptor activation. A significant reduction of ERK1/ERK2, AKT expression in cancer cells after cisplatin plus echistatin treatment was also found. The cancer cells treated by echistatin, cisplatin, and in particular the combination of both compounds drastically increased expression of NFκB transcription factor. Our results suggest that combined therapy cisplatin plus echistatin is a possible way to improve selectiveness of cisplatin. This mechanism probably is due to downregulation of expression of β 1 -integrin and IGF-IR receptors, and the signaling pathway proteins induced by these receptors. Our results suggest that therapy cisplatin plus echistatin is a possible way to improve selectiveness of cisplatin.

Direct effects of hypoxia and nitric oxide on ecdysone secretion by insect prothoracic glands.

PubMed

DeLalio, Leon J; Dion, Sara M; Bootes, Abigail M; Smith, Wendy A

2015-05-01

Insect molting and metamorphosis are controlled by the molt stimulating hormone ecdysone. A recent study suggests that reduced tissue oxygenation correlates with the size-sensing mechanism responsible for triggering molting. When reared in hypoxia, larvae of Manduca sexta and Drosophila melanogaster initiate molting at lower weights than do larvae reared in normoxia. Furthermore, in Drosophila, the signaling gas nitric oxide (NO) appears to be required for normal developmental timing. As observed in Drosophila, NO signaling targets the nuclear hormone receptor beta fushi tarazu transcription factor 1 (βFTZ-F1) through activation of Drosophila hormone receptor 3 (DHR3), two key regulators of ecdysone production and metamorphic tissue progression. We set out to directly examine the effects of hypoxia and NO on ecdysone secretion using prothoracic glands from feeding fifth (last) larval stage M. sexta. Our results indicate that in vitro treatment of prothoracic glands with hypoxia (2% oxygen) or the NO donor DETA-NONOate significantly inhibit ecdysone secretion. Protein markers of glandular activity were also in keeping with an initial inhibition, measured a decrease in phosphorylated ERK (extracellular signal regulated kinase) and an increase in non-phosphorylated 4EBP (eukaryotic initiation factor 4E binding protein). Additionally, gene expression levels of Manduca hormone receptor 3 (mhr3), βftz-f1, nitric oxide synthase (nos), and the PTTH receptor torso, were quantified using real-time PCR. NO treatment increased mhr3 expression and decreased nos expression. Hypoxia increased mhr3 transcription after 2 hr, but decreased transcription after 12 hr, with no effect on nos expression. Both NO and hypoxia had small effects on βftz-f1 expression, yet strongly increased torso transcription. Our results demonstrate that, in isolated prothoracic glands, hypoxia and NO signaling directly inhibit ecdysteroid secretion, but at the same time alter aspects of prothoracic gland function that may enhance secretory response. Copyright © 2015 Elsevier Ltd. All rights reserved.

Ephrin reverse signaling mediates palatal fusion and epithelial-to-mesenchymal transition independently of Tgfß3.

PubMed

Serrano, Maria J; Liu, Jingpeng; Svoboda, Kathy K H; Nawshad, Ali; Benson, M Douglas

2015-12-01

The mammalian secondary palate forms from shelves of epithelia-covered mesenchyme that meet at midline and fuse. The midline epithelial seam (MES) is thought to degrade by apoptosis, epithelial-to-mesenchymal transition (EMT), or both. Failure to degrade the MES blocks fusion and causes cleft palate. It was previously thought that transforming growth factor ß3 (Tgfß3) is required to initiate fusion. Members of the Eph tyrosine kinase receptor family and their membrane-bound ephrin ligands are expressed on the MES. We demonstrated that treatment of mouse palates with recombinant EphB2/Fc to activate ephrin reverse signaling (where the ephrin acts as a receptor and transduces signals from its cytodomain) was sufficient to cause mouse palatal fusion when Tgfß3 signaling was blocked by an antibody against Tgfß3 or by an inhibitor of the TgfßrI serine/threonine receptor kinase. Cultured palatal epithelial cells traded their expression of epithelial cell markers for that of mesenchymal cells and became motile after treatment with EphB2/Fc. They concurrently increased their expression of the EMT-associated transcription factors Snail, Sip1, and Twist1. EphB2/Fc did not cause apoptosis in these cells. These data reveal that ephrin reverse signaling directs palatal fusion in mammals through a mechanism that involves EMT but not apoptosis and activates a gene expression program not previously associated with ephrin reverse signaling. © 2015 Wiley Periodicals, Inc.

The receptor tyrosine kinase ERBB4 is expressed in skin keratinocytes and influences epidermal proliferation.

PubMed

Hoesl, Christine; Röhrl, Jennifer M; Schneider, Marlon R; Dahlhoff, Maik

2018-04-01

The epidermal growth factor receptor (EGFR) and associated receptors ERBB2 and ERBB3 are important for skin development and homeostasis. To date, ERBB4 could not be unambiguously identified in the epidermis. The aim of this study was to analyze the ERBB-receptor family with a special focus on ERBB4 in vitro in human keratinocytes and in vivo in human and murine epidermis. We compared the transcript levels of all ERBB-receptors and the seven EGFR-ligands in HaCaT and A431 cells. ERBB-receptor activity was analyzed after epidermal growth factor (EGF) stimulation by Western blot analysis. The location of the receptors was investigated by immunofluorescence in human keratinocytes and skin. Finally, we investigated the function of ERBB4 in the epidermis of skin-specific ERBB4-knockout mice. After EGF stimulation, all ligands were upregulated except for epigen. Expression levels of EGFR were unchanged, but all other ERBB-receptors were down-regulated after EGF stimulation, although all ERBB-receptors were phosphorylated. We detected ERBB4 at mRNA and protein levels in both human epidermal cell lines and in the basal layer of human and murine epidermis. Skin-specific ERBB4-knockout mice revealed a significantly reduced epidermal thickness with a decreased proliferation rate. ERBB4 is expressed in the basal layer of human epidermis and cultured keratinocytes as well as in murine epidermis. Moreover, ERBB4 is phosphorylated in HaCaT cells due to EGF stimulation, and its deletion in murine epidermis affects skin thickness by decreasing proliferation. ERBB4 is expressed in human keratinocytes and plays a role in murine skin homeostasis. Copyright © 2018 Elsevier B.V. All rights reserved.

A comprehensive data mining study shows that most nuclear receptors act as newly proposed homeostasis-associated molecular pattern receptors.

PubMed

Wang, Luqiao; Nanayakkara, Gayani; Yang, Qian; Tan, Hongmei; Drummer, Charles; Sun, Yu; Shao, Ying; Fu, Hangfei; Cueto, Ramon; Shan, Huimin; Bottiglieri, Teodoro; Li, Ya-Feng; Johnson, Candice; Yang, William Y; Yang, Fan; Xu, Yanjie; Xi, Hang; Liu, Weiqing; Yu, Jun; Choi, Eric T; Cheng, Xiaoshu; Wang, Hong; Yang, Xiaofeng

2017-10-24

Nuclear receptors (NRs) can regulate gene expression; therefore, they are classified as transcription factors. Despite the extensive research carried out on NRs, still several issues including (1) the expression profile of NRs in human tissues, (2) how the NR expression is modulated during atherosclerosis and metabolic diseases, and (3) the overview of the role of NRs in inflammatory conditions are not fully understood. To determine whether and how the expression of NRs are regulated in physiological/pathological conditions, we took an experimental database analysis to determine expression of all 48 known NRs in 21 human and 17 murine tissues as well as in pathological conditions. We made the following significant findings: (1) NRs are differentially expressed in tissues, which may be under regulation by oxygen sensors, angiogenesis pathway, stem cell master regulators, inflammasomes, and tissue hypo-/hypermethylation indexes; (2) NR sequence mutations are associated with increased risks for development of cancers and metabolic, cardiovascular, and autoimmune diseases; (3) NRs have less tendency to be upregulated than downregulated in cancers, and autoimmune and metabolic diseases, which may be regulated by inflammation pathways and mitochondrial energy enzymes; and (4) the innate immune sensor inflammasome/caspase-1 pathway regulates the expression of most NRs. Based on our findings, we propose a new paradigm that most nuclear receptors are anti-inflammatory homeostasis-associated molecular pattern receptors (HAMPRs). Our results have provided a novel insight on NRs as therapeutic targets in metabolic diseases, inflammations, and malignancies.

Regulation of ERRα Gene Expression by Estrogen Receptor Agonists and Antagonists in SKBR3 Breast Cancer Cells: Differential Molecular Mechanisms Mediated by G Protein-Coupled Receptor GPR30/GPER-1

PubMed Central

Li, Yin; Birnbaumer, Lutz; Teng, Christina T.

2010-01-01

In selected tissues and cell lines, 17β-estradiol (E2) regulates the expression of estrogen-related receptor α (ERRα), a member of the orphan nuclear receptor family. This effect is thought to be mediated by the estrogen receptor α (ERα). However in the ERα- and ERβ-negative SKBR3 breast cancer cell line, physiological levels of E2 also stimulate ERRα expression. Here, we explored the molecular mechanism that mediates estrogen action in ER-negative breast cancer cells. We observed that E2, the ERα agonist, as well as the ERα antagonists ICI 182,780 and tamoxifen (TAM), a selective ER modulator, stimulate the transcriptional activity of the ERRα gene and increase the production of ERRα protein in SKBR3 cells. Moreover, the ERRα downstream target genes expression and cellular proliferation are also increased. We show further that the G protein-coupled receptor GPR30/GPER-1 (GPER-1) mediates these effects. The GPER-1 specific ligand G-1 mimics the actions of E2, ICI 182,780, and TAM on ERRα expression, and changing the levels of GPER-1 mRNA by overexpression or small interfering RNA knockdown affected the expression of ERRα accordingly. Utilizing inhibitors, we delineate a different downstream pathway for ER agonist and ER antagonist-triggered signaling through GPER-1. We also find differential histone acetylation and transcription factor recruitment at distinct nucleosomes of the ERRα promoter, depending on whether the cells are activated with E2 or with ER antagonists. These findings provide insight into the molecular mechanisms of GPER-1/ERRα-mediated signaling and may be relevant to what happens in breast cancer cells escaping inhibitory control by TAM. PMID:20211987

Nitric oxide donor restores lung growth factor and receptor expression in hyperoxia-exposed rat pups.

PubMed

Lopez, Emmanuel; Boucherat, Olivier; Franco-Montoya, Marie-Laure; Bourbon, Jacques R; Delacourt, Christophe; Jarreau, Pierre-Henri

2006-06-01

Exposure of newborn rats to hyperoxia impairs alveolarization. Nitric oxide (NO) may prevent this evolution. Angiogenesis and factors involved in this process, but also other growth factors (GFs) involved in alveolar development, are likely potential therapeutic targets for NO. We studied the effects of the NO donor, [Z]-1-[N-(2-aminoethyl)-N-(2-ammonioethyl)aminio]diazen-1-ium-1, 2-diolate, also termed DETANONOate (D-NO), on hyperoxia-induced changes in key regulatory factors of alveolar development in neonatal rats, and its possible preventive effect on the physiologic consequences of hyperoxia. Newborn rat pups were randomized at birth to hyperoxia (> 95% O2) or room air exposure for 6 or 10 d, while receiving D-NO or its diluent. On Day 6, several GFs and their receptors were studied at pre- and/or post-translational levels. Elastin transcript determination on Day 6, and elastin deposition in tissue and morphometric analysis of the lungs on Day 10, were also performed. Hyperoxia decreased the expression of vascular endothelial growth factor (VEGF) receptor (VEGFR) 2, fibroblast growth factor (FGF)-18, and FGF receptors (FGFRs) FGFR3 and FGFR4, increased mortality, and impaired alveolarization and capillary growth. D-NO treatment of hyperoxia-exposed pups restored the expression level of FGF18 and FGFR4, induced an increase of both VEGF mRNA and protein, enhanced elastin expression, and partially restored elastin deposition in alveolar walls. Although, under the present conditions, D-NO failed to prevent the physiologic consequences of hyperoxia in terms of survival and lung alveolarization, our findings demonstrate molecular effects of NO on GFs involved in alveolar development that may have contributed to the protective effects previously reported for NO.

Altered subcellular localization of heat shock protein 90 is associated with impaired expression of the aryl hydrocarbon receptor pathway in dogs.

PubMed

van Steenbeek, Frank G; Spee, Bart; Penning, Louis C; Kummeling, Anne; van Gils, Ingrid H M; Grinwis, Guy C M; Van Leenen, Dik; Holstege, Frank C P; Vos-Loohuis, Manon; Rothuizen, Jan; Leegwater, Peter A J

2013-01-01

The aryl hydrocarbon receptor (AHR) mediates biological responses to toxic chemicals. An unexpected role for AHR in vascularization was suggested when mice lacking AHR displayed impaired closure of the ductus venosus after birth, as did knockout mice for aryl hydrocarbon receptor interacting protein (AIP) and aryl hydrocarbon receptor nuclear translocator (ARNT). The resulting intrahepatic portosystemic shunts (IHPSS) are frequently diagnosed in specific dog breeds, such as the Irish wolfhound. We compared the expression of components of the AHR pathway in healthy Irish wolfhounds and dogs with IHPSS. To this end, we analyzed the mRNA expression in the liver of AHR,AIP, ARNT, and other genes involved in this pathway, namely, those for aryl hydrocarbon receptor nuclear translocator 2 (ARNT2), hypoxia inducible factor 1alpha (HIF1A), heat shock protein 90AA1 (HSP90AA1), cytochromes P450 (CYP1A1, CYP1A2, and CYP1B1), vascular endothelial growth factor A (VEGFA), nitric oxide synthesase 3 (NOS3), and endothelin (EDN1). The observed low expression of AHR mRNA in the Irish wolfhounds is in associated with a LINE-1 insertion in intron 2, for which these dogs were homozygous. Down regulation in Irish wolfhounds was observed for AIP, ARNT2, CYP1A2, CYP1B1 and HSP90AA1 expression, whereas the expression of HIF1A was increased. Immunohistochemistry revealed lower levels of AHR, HIF1A, and VEGFA protein in the nucleus and lower levels of ARNT and HSP90AA1 protein in the cytoplasm of the liver cells of Irish wolfhounds. The impaired expression of HSP90AA1 could trigger the observed differences in mRNA and protein levels and therefore explain the link between two very different functions of AHR: regulation of the closure of the ductus venosus and the response to toxins.

Comprehensive analysis of fibroblast growth factor receptor expression patterns during chick forelimb development.

PubMed

Sheeba, Caroline J; Andrade, Raquel P; Duprez, Delphine; Palmeirim, Isabel

2010-01-01

Specific interactions between fibroblast growth factors (Fgf1-22) and their tyrosine kinase receptors (FgfR1-4) activate different signalling pathways that are responsible for the biological processes in which Fgf signalling is implicated during embryonic development. In the chick, several Fgf ligands (Fgf2, 4, 8, 9, 10, 12, 13 and 18) and the four FgfRs (FgfR 1, 2, 3 and 4) have been reported to be expressed in the developing limb. The precise spatial and temporal expression of these transcripts is important to guide the limb bud to develop into a wing/leg. In this paper, we present a detailed and systematic analysis of the expression patterns of FgfR1, 2, 3 and 4 throughout chick wing development, by in situ hybridisation on whole mounts and sections. Moreover, we characterize for the first time the different isoforms of FGFR1-3 by analysing their differential expression in limb ectoderm and mesodermal tissues, using RT-PCR and in situ hybridisation on sections. Finally, isoform-specific sequences for FgfR1IIIb, FgfR1IIIc, FgfR3IIIb and FgfR3IIIc were determined and deposited in GenBank with the following accession numbers: GU053725, GU065444, GU053726, GU065445, respectively.

The role of cortistatin in the human immune system.

PubMed

van Hagen, P Martin; Dalm, Virgil A; Staal, Frank; Hofland, Leo J

2008-05-14

Cortistatin (CST) is a recently described neuropeptide that shares high homology with somatostatin (somatotropin release-inhibiting factor, SRIF) and binds with high affinity to all somatostatin (sst) receptor subtypes. CST is currently known to have a widespread distribution in many human organs including the immune system. The activities specific to CST may be partially attributable to its binding to the growth hormone secretagogue (GHS)-receptor (GHS-R) and the orphan G-protein-coupled receptor MrgX2. Human immune cells produce CST, whereas macrophage lineage and activated endothelium express sst2, and human lymphocytes express sst3. The human thymus expresses sst1, 2, 3, MrgX2 and almost all immune cells express GHS-R. Moreover, at this very moment promising research with CST in experimental animal models is being performed. On the basis of these promising results, studies aiming to further evaluate the possibilities of CST as a therapeutic agent in human immune-mediated inflammatory diseases are warranted.

Biochemical characterization and analysis of the transforming potential of the FLT3/FLK2 receptor tyrosine kinase.

PubMed

Maroc, N; Rottapel, R; Rosnet, O; Marchetto, S; Lavezzi, C; Mannoni, P; Birnbaum, D; Dubreuil, P

1993-04-01

We recently cloned an additional member of the receptor type tyrosine kinase class III. This new gene, called Flt3 by our group [Rosnet, O., Matteï, M.G., Marchetto, S. & Birnbaum, D. (1991). Genomics, 9, 380-385; Rosnet, O., Marchetto, S., deLapeyriere, O. & Birnbaum, D. (1991). Oncogene, 6, 1641-1650] and Flk2 by others [Matthews, W., Jordan, C.T., Wieg, G.W., Pardoll, D. & Lemischka, I.R. (1991). Cell, 65, 1143-1152] is strongly related to the important developmental genes Kit, Fms and Pdgfr. The murine 3.2-kb full-length cDNA, when introduced into COS-1 cells, shows the expression of two polypeptides with apparent molecular weights of 155 kDa and 132 kDa. Treatment of cells with N-linked glycosylation inhibitors results in the expression of a 110-kDa protein. We have shown that FLT3 contains an intrinsic tyrosine kinase activity. A point mutation in a highly conserved residue within the phosphoryltransferase domain inactivates the catalytic function of this receptor, whereas activation by way of a chimeric molecule between the ligand-binding domain of colony-stimulating factor type 1 (CSF-1) receptor (CSF-1R) and the kinase domain of FLT3 results, in the presence of CSF-1, in the development of the transforming activity of this receptor as shown by anchorage-independent cell growth. Finally, expression analysis of the FLT3 protein shows that, in addition to the hematopoietic system, FLT3 is strongly expressed in neural, gonadal, hepatic and placental tissues in the mouse.

Nuclear translocation of the 1,25D{sub 3}-MARRS (membrane associated rapid response to steroids) receptor protein and NF{kappa}B in differentiating NB4 leukemia cells

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

Wu, Wenqing; Beilhartz, Greg; Roy, Yvette

2010-04-15

1,25 Dihydroxyvitamin D{sub 3} (1,25D{sub 3}) primes NB4 promyelocytic leukemia cells to differentiate along the monocyte/macrophage lineage through a non-genomic mechanism. Here we show that NB4 cells express high levels of the recently identified membrane receptor for 1,25D{sub 3}, which is a distinct gene product from the classical nuclear vitamin D receptor. This 57 kDa protein, named 1,25D{sub 3}-MARRS (Membrane Activated Rapid Response to Steroids)/ERp57/PIA3 appears to associate in a complex with the transcription factor, nuclear factor kappa B (NF{kappa}B). In unstimulated cells, 1,25D{sub 3}-MARRS can be co-immunoprecipitated with antibodies directed at NF{kappa}B, and NF{kappa}B is co-precipitated when antibodies againstmore » 1,25D{sub 3}-MARRS or ERp57 are used. Confocal microscopy and subcellular fractionation studies demonstrate that both 1,25D{sub 3}-MARRS and NF{kappa}B begin translocating to the nucleus within minutes of co-stimulation with 1,25D{sub 3} and phorbol ester. The predominant nuclear localization of both proteins precedes the expression of the monocyte/macrophage phenotype and suggests that this event may be critical to the differentiation pathway. This suggests a role for 1,25D{sub 3}-MARRS in the nucleus as a regulator of gene expression. Here it may also regulate the activity of NF{kappa}B and other factors with which it may be interacting.« less

Microvascular pericytes express platelet-derived growth factor-beta receptors in human healing wounds and colorectal adenocarcinoma.

PubMed Central

Sundberg, C.; Ljungström, M.; Lindmark, G.; Gerdin, B.; Rubin, K.

1993-01-01

The expression of platelet-derived growth factor- beta (PDGF-beta) receptors in the microvasculature of human healing wounds and colorectal adenocarcinoma was investigated. Frozen sections were subjected to double immunofluorescence staining using monoclonal antibodies (MAbs) specific for pericytes (MAb 225.28 recognizing the high-molecular weight-melanoma-associated antigen, expressed by activated pericytes during angiogenesis), endothelial cells (MAb PAL-E), laminin, as well as PDGF-beta receptors (MAb PDGFR-B2) and its ligand PDGF-B chain (MAb PDGF 007). Stained sections were analyzed by computer-aided imaging processing that allowed for a numerical quantification of the degree of colocalization of the investigated antigens. An apparent background colocalization, varying between 23 and 35%, between markers for cells not expected to co-localize was recorded. This background could be due to limitations of camera resolution, to out-of-focus fluorescence, and to interdigitations of the investigated structures. In all six tumor specimens, co-localization of PDGF-beta receptors and PAL-E was not different from the background co-localization, whereas that of PDGF-beta receptors and high-molecular weight-melanoma-associated antigen was significantly higher with mean values between 57 and 71%. Qualitatively, the same pattern was obtained in the two investigated healing wounds. PDGF-B chain did not co-localize with either PAL-E or high-molecular weight-melanoma-associated antigen, but PDGF-B chain-expressing cells were, however, frequently found juxtaposed to the microvasculature. The expression of PDGF-beta receptors on pericytes in activated microvessels and the presence of PDGF-B chain-expressing cells in close proximity to the microvasculature of healing wounds and colorectal adenocarcinoma is compatible with a role for PDGF in the physiology of the microvasculature in these conditions. Images Figure 1 p1381-a Figure 3 Figure 4 PMID:8238254

Selenoprotein W controls epidermal growth factor receptor surface expression, activation and degradation via receptor ubiquitination

USDA-ARS?s Scientific Manuscript database

Epidermal growth factor (EGF) receptor (EGFR) is the founding member of the ErbB family of growth factor receptors that modulate a complex network of intracellular signaling pathways controlling growth, proliferation and differentiation. Selenoprotein W (SEPW1) is a diet-regulated, highly conserved...

Platelets Express Activated P2Y12 Receptor in Patients With Diabetes Mellitus.

PubMed

Hu, Liang; Chang, Lin; Zhang, Yan; Zhai, Lili; Zhang, Shenghui; Qi, Zhiyong; Yan, Hongmei; Yan, Yan; Luo, Xinping; Zhang, Si; Wang, Yiping; Kunapuli, Satya P; Ye, Hongying; Ding, Zhongren

2017-08-29

Platelets from patients with diabetes mellitus are hyperactive. Hyperactivated platelets may contribute to cardiovascular complications and inadequate responses to antiplatelet agents in the setting of diabetes mellitus. However, the underlying mechanism of hyperactivated platelets is not completely understood. We measured P2Y 12 expression on platelets from patients with type 2 diabetes mellitus and on platelets from rats with diabetes mellitus. We also assayed platelet P2Y 12 activation by measuring cAMP and VASP phosphorylation. The antiplatelet and antithrombotic effects of AR-C78511 and cangrelor were compared in rats. Finally, we explored the role of the nuclear factor-κB pathway in regulating P2Y 12 receptor expression in megakaryocytes. Platelet P2Y 12 levels are 4-fold higher in patients with type 2 diabetes mellitus compared with healthy subjects. P2Y 12 expression correlates with ADP-induced platelet aggregation (r=0.89, P <0.01). P2Y 12 in platelets from patients with diabetes mellitus is constitutively activated. Although both AR-C78511, a potent P2Y 12 inverse agonist, and cangrelor have similar antiplatelet efficacy on platelets from healthy subjects, AR-C78511 exhibits more powerful antiplatelet effects on diabetic platelets than cangrelor (aggregation ratio 36±3% versus 49±5%, respectively, P <0.05). Using a FeCl 3 -injury mesenteric arteriole thrombosis model in rats and an arteriovenous shunt thrombosis model in rats, we found that the inverse agonist AR-C78511 has greater antithrombotic effects on GK rats with diabetes mellitus than cangrelor (thrombus weight 4.9±0.3 mg versus 8.3±0.4 mg, respectively, P <0.01). We also found that a pathway involving high glucose-reactive oxygen species-nuclear factor-κB increases platelet P2Y 12 receptor expression in diabetes mellitus. Platelet P2Y 12 receptor expression is significantly increased and the receptor is constitutively activated in patients with type 2 diabetes mellitus, which contributes to platelet hyperactivity and limits antiplatelet drug efficacy in type 2 diabetes mellitus. © 2017 American Heart Association, Inc.

Differential expression of FGF receptors and of myogenic regulatory factors in primary cultures of satellite cells originating from fast (EDL) and slow (Soleus) twitch rat muscles.

PubMed

Martelly, I; Soulet, L; Bonnavaud, S; Cebrian, J; Gautron, J; Barritault, D

2000-11-01

In the rat, the fast and slow twitch muscles respectively Extensor digitorum longus (EDL) and Soleus present differential characteristics during regeneration. This suggests that their satellite cells responsible for muscle growth and repair represent distinct cellular populations. We have previously shown that satellite cells dissociated from Soleus and grown in vitro proliferate more readily than those isolated from EDL muscle. Fibroblast growth factors (FGFs) are known as regulators of myoblast proliferation and several studies have revealed a relationship between the response of myoblasts to FGF and the expression of myogenic regulatory factors (MRF) of the MyoD family by myoblasts. Therefore, we presently examined the possibility that the satellite cells isolated from EDL and Soleus muscles differ in the expression of FGF receptors (FGF-R) and of MRF expression. FGF-R1 and -R4 were strongly expressed in proliferating cultures whereas FGF-R2 and R3 were not detected in these cultures. In differentiating cultures, only -R1 was present in EDL satellite cells while FGF-R4 was also still expressed in Soleus cells. Interestingly, the unconventional receptor for FGF called cystein rich FGF receptor (CFR), of yet unknown function, was mainly detected in EDL satellite cell cultures. Soleus and EDL satellite cell cultures also differed in the expression MRFs. These results are consistent with the notion that satellite cells from fast and slow twitch muscles belong to different types of myogenic cells and suggest that satellite cells might play distinct roles in the formation and diversification of fast and slow fibres.

Toll-like receptor-mediated inhibition of Gas6 and ProS expression facilitates inflammatory cytokine production in mouse macrophages

PubMed Central

Deng, Tingting; Zhang, Yue; Chen, Qiaoyuan; Yan, Keqin; Han, Daishu

2012-01-01

Activation of Toll-like receptors (TLRs) triggers rapid inflammatory cytokine production in various cell types. The exogenous product of growth-arrest-specific gene 6 (Gas6) and Protein S (ProS) inhibit the TLR-triggered inflammatory responses through the activation of Tyro3, Axl and Mer (TAM) receptors. However, regulation of the Gas6/ProS-TAM system remains largely unknown. In the current study, mouse macrophages are shown to constitutively express Gas6 and ProS, which synergistically suppress the basal and TLR-triggered production of inflammatory cytokines, including those of tumour necrosis factor-α, interleukin-6 and interleukin-1β, by the macrophages in an autocrine manner. Notably, TLR signalling markedly decreases Gas6 and ProS expression in macrophages through the activation of the nuclear factor-κB. Further, the down-regulation of Gas6 and ProS by TLR signalling facilitates the TLR-mediated inflammatory cytokine production in mouse macrophages. These results describe a self-regulatory mechanism of TLR signalling through the suppression of Gas6 and ProS expression. PMID:22043818

Human microRNA-1245 down-regulates the NKG2D receptor in natural killer cells and impairs NKG2D-mediated functions

PubMed Central

Espinoza, J. Luis; Takami, Akiyoshi; Yoshioka, Katsuji; Nakata, Katsuya; Sato, Tokiharu; Kasahara, Yoshihito; Nakao, Shinji

2012-01-01

Background NKG2D is an activating receptor expressed by natural killer and T cells, which have crucial functions in tumor and microbial immunosurveillance. Several cytokines have been identified as modulators of NKG2D receptor expression. However, little is known about NKG2D gene regulation. In this study, we found that microRNA 1245 attenuated the expression of NKG2D in natural killer cells. Design and Methods We investigated the potential interactions between the 3′-untranslated region of the NKG2D gene and microRNA as well as their functional roles in the regulation of NKG2D expression and cytotoxicity in natural killer cells. Results Transforming growth factor-β1, a major negative regulator of NKG2D expression, post-transcriptionally up-regulated mature microRNA-1245 expression, thus down-regulating NKG2D expression and impairing NKG2D-mediated immune responses in natural killer cells. Conversely, microRNA-1245 down-regulation significantly increased the expression of NKG2D expression in natural killer cells, resulting in more efficient NKG2D-mediated cytotoxicity. Conclusions These results reveal a novel NKG2D regulatory pathway mediated by microRNA-1245, which may represent one of the mechanisms used by transforming growth factor-β1 to attenuate NKG2D expression in natural killer cells. PMID:22491735

Expression of IGF-I, IGF-I receptor and IGF binding proteins-1, -2, -3, -4 and -5 in human atherectomy specimens.

PubMed

Grant, M B; Wargovich, T J; Ellis, E A; Tarnuzzer, R; Caballero, S; Estes, K; Rossing, M; Spoerri, P E; Pepine, C

1996-12-17

The molecular and cellular processes that induce rapid atherosclerotic plaque progression in patients with unstable angina and initiate restenosis following coronary interventional procedures are uncertain. We examined primary (de novo) and restenotic lesions retrieved at the time of directional coronary atherectomy for expression of insulin-like-growth factor-I (IGF-I). IGF-I receptor, and five IGF binding proteins (IGFBPs), IGFBP-1, IGFBP-2, IGFBP-3, IGFBP-4, and IGFBP-5 in smooth muscle cells (SMCs) using colloidal gold immunocytochemistry. IGF-1, its receptor and binding proteins were not detected in SMCs of normal coronary arteries. IGF-I localized primarily in synthetic smooth muscle cells (sSMCs) in both de novo and restenotic plaques. IGF-I receptor localized on sSMCs and their processes and colocalized with IGF-I. Although morphometric analysis of IGF-I and IGF-I receptor immunoreactivity in sSMCs of de novo and restenotic lesions showed comparable levels of IGF-I (3.2 +/- 1.0 and 2.9 +/- 0.9, respectively). IGF-I receptor was significantly higher in de novo lesions as compared to restenotic lesions (10.7 +/- 2.5 and 4.2 +/- 1.3, P < 0.05, respectively). IGFBP-1, IGFBP-2, IGFBP-3, IGFBP-4 and IGFBP-5 localized in the cytoplasm of sSMCs and in the extracellular matrix. Quantitative reverse transcription polymerase chain reaction (QRT-PCR) performed on de novo atherectomy specimens identified mRNA for IGF-I, IGF-I receptor, IGFBP-1, IGFBP-2, IGFBP-4, IGFBP-5 levels and detected mRNA for IGFBP-3. The expression of IGF-I, IGF-I receptor, and IGFBPs in atherectomy plaques suggests that the development of coronary obstructive lesions may be a result of changes in the IGF system.

GATA3 expression in triple-negative breast cancers.

PubMed

Byrne, David J; Deb, Siddhartha; Takano, Elena A; Fox, Stephen B

2017-07-01

GATA-binding protein 3 (GATA3) is a well-studied transcription factor found to be essential in the development of luminal breast epithelium and has been identified in a variety of tumour types, including breast and urothelial carcinomas, making it a useful immunohistochemistry marker in the diagnosis of both primary and metastatic disease. We investigated GATA3 protein expression in a 106 primary triple-negative breast carcinomas (100 basal-like, six non-basal-like) using Cell Marque mouse monoclonal anti-GATA3 (L50-823). Reverse transcription-quantitative polymerase chain reaction (RT-qPCR) was used to quantify mRNA expression in 22 triple-negative breast cancers (TNBCs) (20 primary and two cell lines), four luminal (three primary and one cell line) and five human epidermal growth factor receptor 2 (HER2) (four primary and one cell line) amplified tumours. In 98 TNBCs where IHC was assessable, 47 (48%) had a 1+ or greater staining with 20 (21%) having high GATA3 expression when using a weighted scoring. Our study has demonstrated that GATA3 expression is common in primary triple-negative breast carcinomas. It also suggests that although GATA3 is an oestrogen receptor (ER) regulated gene, it still proves useful in differentiating between primary and metastatic tumours in patients with a history of breast cancer regardless of its molecular subtype. © 2017 John Wiley & Sons Ltd.

Effects of Toll-like receptor 3 on herpes simplex virus type-1-infected mouse neural stem cells.

PubMed

Sun, Xiuning; Shi, Lihong; Zhang, Haoyun; Li, Ruifang; Liang, Ruiwen; Liu, Zhijun

2015-03-01

In this study, we aimed to investigate the effect of herpes simplex virus type-1 (HSV-1) infection on the phosphorylation of interferon regulatory factor 3 (IRF3) and the expression of interferon-β (IFN-β), as well as to clarify the functions of toll-like receptor 3 (TLR3) in mouse neural stem cells (NSCs) infected with HSV-1. In HSV-1-infected cultured NSCs, immunofluorescence, reverse transcription - polymerase chain reaction, Western blot, and ELISA were performed to reveal the expression patterns of TLR3, IRF3, and IFN-β. Then, lentivirus-mediated RNA interference (RNAi) was used to block the expression of TLR3, and its effect on host resistance to HSV-1 infection was investigated. Under uninfected conditions, NSCs expressed TLR3 and phosphorylated IRF3, but after infection, the expression level of TLR3 was upregulated and the phosphorylation level of IRF3 in the nucleus was significantly enhanced, while IFN-β was also expressed. After TLR3 expression was blocked by lentivirus-mediated RNAi, IRF3 phosphorylation and IFN-β expression were downregulated. Therefore, HSV-1 upregulated the expression of TLR3 in NSCs and promoted nuclear translocation after IRF3 was phosphorylated to induce IFN-β expression. TLR3 exhibited an anti-HSV-1 infection capacity via innate immune functions.

The Wnt receptor Ryk controls specification of GABAergic neurons versus oligodendrocytes during telencephalon development

PubMed Central

Zhong, Jingyang; Kim, Hyoung-Tai; Lyu, Jungmook; Yoshikawa, Kazuaki; Nakafuku, Masato; Lu, Wange

2011-01-01

GABAergic neurons and oligodendrocytes originate from progenitors within the ventral telencephalon. However, the molecular mechanisms that control neuron-glial cell-fate segregation, especially how extrinsic factors regulate cell-fate changes, are poorly understood. We have discovered that the Wnt receptor Ryk promotes GABAergic neuron production while repressing oligodendrocyte formation in the ventral telencephalon. We demonstrate that Ryk controls the cell-fate switch by negatively regulating expression of the intrinsic oligodendrogenic factor Olig2 while inducing expression of the interneuron fate determinant Dlx2. In addition, we demonstrate that Ryk is required for GABAergic neuron induction and oligodendrogenesis inhibition caused by Wnt3a stimulation. Furthermore, we showed that the cleaved intracellular domain of Ryk is sufficient to regulate the cell-fate switch by regulating the expression of intrinsic cell-fate determinants. These results identify Ryk as a multi-functional receptor that is able to transduce extrinsic cues into progenitor cells, promote GABAergic neuron formation, and inhibit oligodendrogenesis during ventral embryonic brain development. PMID:21205786

Delayed Parturition and Altered Myometrial Progesterone Receptor Isoform A Expression in Mice Null for Kruppel-like Factor 9

USDA-ARS?s Scientific Manuscript database

Pre-term and delayed labor conditions are devastating health problems, with currently unknown etiologies. We previously showed that the transcription factor Krüppel-like factor 9 (KLF9) influences the expression and/or transcriptional activity of receptors for estrogen and progesterone in endometria...

No Detectable Hypoxia in Malignant Salivary Gland Tumors: Preliminary Results

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

Wijffels, Karien; Hoogsteen, Ilse J.; Lok, Jasper

2009-04-01

Purpose: Hypoxia is detected in most solid tumors and is associated with malignant progression and adverse treatment outcomes. However, the oxygenation status of malignant salivary gland tumors has not been previously studied. The aim of this study was to investigate the potential clinical relevance of hypoxia in this tumor type. Methods and Materials: Twelve patients scheduled for surgical resection of a salivary gland tumor were preoperatively injected with the hypoxia marker pimonidazole and the proliferation marker iododeoxyuridine. Tissue samples of the dissected tumor were immunohistochemically stained for blood vessels, pimonidazole, carbonic anhydrase-IX, glucose transporters-1 and -3 (Glut-1, Glut-3), hypoxia-inducible factor-1{alpha},more » iododeoxyuridine, and epidermal growth factor receptor. The tissue sections were quantitatively assessed by computerized image analysis. Results: The tissue material from 8 patients was of sufficient quality for quantitative analysis. All tumors were negative for pimonidazole binding, as well as for carbonic anhydrase-IX, Glut-1, Glut-3, and hypoxia-inducible factor-1{alpha}. The vascular density was high, with a median value of 285 mm{sup -2} (range, 209-546). The iododeoxyuridine-labeling index varied from <0.1% to 12.2% (median, 2.2%). Epidermal growth factor receptor expression levels were mostly moderate to high. In one-half of the cases, nuclear expression of epidermal growth factor receptor was observed. Conclusion: The absence of detectable pimonidazole binding, as well as the lack of expression of hypoxia-associated proteins in all tumors, indicates that malignant salivary gland tumors are generally well oxygenated. It is unlikely that hypoxia is a relevant factor for their clinical behavior and treatment responsiveness.« 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.

Integrated analysis reveals that STAT3 is central to the crosstalk between HER/ErbB receptor signaling pathways in human mammary epithelial cells

DOE PAGES

Gong, Chunhong; Zhang, Yi; Shankaran, Harish; ...

2014-10-02

Human epidermal growth factor receptors (HER, also known as ErbB) drive cellular proliferation, pro-survival and stress responses by activating several downstream kinases, in particular ERK, p38, JNK (SAPK), the PI3K/AKT, as well as various transcriptional regulators such as STAT3. When co-expressed, first three members of HER family (HER1-3) can form homo- and hetero-dimers. Based on the considerable evidence which suggest that every receptor dimer activates intracellular signaling pathways differentially, we hypothesized that the HER dimerization pattern is a better predictor of downstream signaling than the total receptor activation levels. We validated our hypothesis using a combination of model-based analysis tomore » quantify the HER dimerization patterns and multi-factorial experiments where HER dimerization patterns and signaling crosstalk were rationally perturbed. We have measured the activation of HER1-3 receptors and of the sentinel signaling proteins ERK, AKT, p38, JNK, STAT3 as a function of time in a panel of human mammary epithelial (HME) cells expressing different levels of HER1-3 stimulated with various ligand combinations. Our analysis using multiple ways of clustering the activation data has confirmed that the HER receptor dimer is a better predictor of the signaling through p38, ERK and AKT pathways than the total HER receptor expression and activation levels. Targeted inhibition studies to identify the causal effects allowed us to obtain a consensus regulatory interaction model, which revealed that STAT3 occupies a central role in the crosstalk between the studied pathways.« less

EFFECTS OF EPIDERMAL GROWTH FACTOR (EGF), TRANSFORMING GROWTH FACTOR- (TGF), AND 2,3,7,8-TETRACHLORODIBENZO-P-DIOXIN ON FUSION OF EMBRYONIC PALATES IN SERUM-FREE ORGAN CULTURE USING WILD-TYPE, EGF KNOCKOUT, AND TGF KNOCKOUT MOUSE STRAINS

EPA Science Inventory

Backround: 2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) is teratogenic in mice, producing cleft palate (CP). TCDD exposure disrupts expression of epidermal growth factor (EGF) receptor, EGF, and transforming growth factor- (TGF) in the palate and affects proliferation and different...

Reduced Sympathetic Innervation in Endometriosis is Associated to Semaphorin 3C and 3F Expression.

PubMed

Scheerer, Claudia; Frangini, Sergio; Chiantera, Vito; Mechsner, Sylvia

2017-09-01

Endometriosis is a chronic inflammatory disease and one of the most common causes of pelvic pain. The mechanisms underlying pain emergence or chronic inflammation during endometriosis remain unknown. Several chronic inflammatory diseases including endometriosis show reduced amounts of noradrenergic nerve fibers. The source of the affected innervation is still unclear. Semaphorins represent potential elicitors, due to their known role as axonal guidance cues, and are suggested as nerve repellent factors in different chronic inflammatory diseases. Therefore, semaphorins might influence the progress of neuroinflammatory mechanisms during endometriosis. Here, we analyzed the noradrenergic innervation and the expression of the specific semaphorins and receptors possibly involved in the neuroimmunomodulation in endometriosis. Our studies revealed an affected innervation and a significant increase of semaphorins and their receptors in peritoneal endometriotic tissue. Thereby, the expression of the receptors was identified on the membrane of noradrenergic nerve fibers and vessels. Macrophages and activated fibroblasts were found in higher density levels and additionally express semaphorins in peritoneal endometriotic tissue. Inflammation leads to an increased release of immune cells, which secrete a variety of inflammatory factors capable of affecting innervation. Therefore, our data suggests that the chronic inflammatory condition in endometriosis might contribute to the increase of semaphorins, which could possibly affect the innervation in peritoneal endometriosis.

Role of atypical chemokine receptor ACKR2 in experimental oral squamous cell carcinogenesis.

PubMed

da Silva, Janine Mayra; Dos Santos, Tálita Pollyanna Moreira; Saraiva, Adriana Machado; Fernandes de Oliveira, Ana Laura; Garlet, Gustavo Pompermaier; Batista, Aline Carvalho; de Mesquita, Ricardo Alves; Russo, Remo Castro; da Silva, Tarcília Aparecida

2018-03-14

Chemokines and chemokine receptors are critical in oral tumourigenesis. The atypical chemokine receptor ACKR2 is a scavenger of CC chemokines controlling the availability of these molecules at tumour sites, but the role of ACKR2 in the context of oral carcinogenesis is unexplored. In this study, wild-type (WT) and ACKR2 deficient mice (ACKR2 -/- ) were treated with chemical carcinogen 4-nitroquinoline-1-oxide (4NQO) for induction of oral carcinogenesis. Tongues were collected for macro and microscopic analysis and to evaluate the expression of ACKRs, CC chemokines and its receptors, inflammatory cytokines, angiogenic factors, adhesion molecules and extracellular matrix components. An increased expression of ACKR2 in squamous cell carcinoma (SCC) lesions of 4NQO-treated WT mice was observed. No significant differences were seen in the ACKR1, ACKR3 and ACKR4 mRNA expression comparing SCC lesions from WT and ACKR2 -/- treated mice. Significantly higher expression of CCL2, IL-6 and IL-17 was detected in ACKR2 -/- treated mice. In contrast, the expression of other CC-chemokines, and receptors, angiogenic factors, adhesion molecules and extracellular matrix components were similarly increased in SCC lesions of both groups. Clinical and histopathological analysis revealed no differences in inflammatory cell recruitment and in the SCC incidence comparing WT and ACKR2 -/- treated mice. The results suggest that ACKR2 expression regulates inflammation in tumour-microenvironment but the absence of ACKR2 does not impact chemically-induced oral carcinogenesis. Copyright © 2018 Elsevier Ltd. All rights reserved.

Can insulin-like growth factor 1 (IGF-1), IGF-1 receptor connective tissue growth factor and Ki-67 labelling index have a prognostic role in pulmonary carcinoids?

PubMed

Kanakis, Georgios A; Grimelius, Lars; Papaioannou, Dimitrios; Kaltsas, Gregory; Tsolakis, Apostolos V

2018-04-27

Altered expression of Insulin-like Growth Factor-1 (IGF-1), its receptor (IGF-1R), Connective Tissue Growth Factor (CTGF) and Hypoxia Inducible Factor-1 (HIF-1), has been implicated in tumorigenesis. So far, these factors have not been studied systematically in Pulmonary Carcinoids (PCs). To examine IGF-1, IGF-1R, CTGF and HIF-1 expression in PCs, and assess their prognostic value over established factors. Retrospective study of 121 PCs (104 Typical and 17 Atypical). The expression of growth factors was studied immunohistochemically and tumors were considered positive if immunoreactivity appeared in >50% of cells. All studied parameters were expressed in the majority of tumors (IGF-1, IGF-1R, CTGF and HIF-1, in 78.5%, 67%, 72% and 78%, respectively). Their expression tended to be more frequent in TCs and in tumors with Ki-67≤2% (significant only for HIF-1; 82 vs. 53%; p=0.023 and 83 vs. 63%; p=0.025 respectively). CTGF was the only factor correlated with more extensive disease (larger size; presence of lymph node and distant metastases). According to logistic regression analysis, only advanced age, Ki-67≥3.4% and lymph node involvement could predict the development of distant metastases. IGF-1, IGF-1R, CTGF and HIF-1 are avidly expressed in PCs; however, their presence did not appear to be of statistically significant value over established prognostic factors.

Vascular Endothelial Growth Factor Modulates Skeletal Myoblast Function

PubMed Central

Germani, Antonia; Di Carlo, Anna; Mangoni, Antonella; Straino, Stefania; Giacinti, Cristina; Turrini, Paolo; Biglioli, Paolo; Capogrossi, Maurizio C.

2003-01-01

Vascular endothelial growth factor (VEGF) expression is enhanced in ischemic skeletal muscle and is thought to play a key role in the angiogenic response to ischemia. However, it is still unknown whether, in addition to new blood vessel growth, VEGF modulates skeletal muscle cell function. In the present study immunohistochemical analysis showed that, in normoperfused mouse hindlimb, VEGF and its receptors Flk-1 and Flt-1 were expressed mostly in quiescent satellite cells. Unilateral hindlimb ischemia was induced by left femoral artery ligation. At day 3 and day 7 after the induction of ischemia, Flk-1 and Flt-1 were expressed in regenerating muscle fibers and VEGF expression by these fibers was markedly enhanced. Additional in vitro experiments showed that in growing medium both cultured satellite cells and myoblast cell line C2C12 expressed VEGF and its receptors. Under these conditions, Flk-1 receptor exhibited constitutive tyrosine phosphorylation that was increased by VEGF treatment. During myogenic differentiation Flk-1 and Flt-1 were down-regulated. In a modified Boyden Chamber assay, VEGF enhanced C2C12 myoblasts migration approximately fivefold. Moreover, VEGF administration to differentiating C2C12 myoblasts prevented apoptosis, while inhibition of VEGF signaling either with selective VEGF receptor inhibitors (SU1498 and CB676475) or a neutralizing Flk-1 antibody, enhanced cell death approximately 3.5-fold. Finally, adenovirus-mediated VEGF165 gene transfer inhibited ischemia-induced apoptosis in skeletal muscle. These results support a role for VEGF in myoblast migration and survival, and suggest a novel autocrine role of VEGF in skeletal muscle repair during ischemia. PMID:14507649

Inhibition of LPS binding to MD-2 co-receptor for suppressing TLR4-mediated expression of inflammatory cytokine by 1-dehydro-10-gingerdione from dietary ginger

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

Park, Sun Hong; Kyeong, Min Sik; Hwang, Yuri

Highlights: Black-Right-Pointing-Pointer 1-Dehydro-10-gingerdione (1D10G) from ginger inhibits LPS binding to MD-2. Black-Right-Pointing-Pointer 1D10G suppresses MyD88- or TRIF-dependent signaling in LPS-activated macrophages. Black-Right-Pointing-Pointer 1D10G down-regulates the expression of NF-{kappa}B-, AP1- or IRF3-target genes. Black-Right-Pointing-Pointer MD-2 is a molecular target in the anti-inflammatory action of 1D10G. -- Abstract: Myeloid differentiation protein 2 (MD-2) is a co-receptor of toll-like receptor 4 (TLR4) for innate immunity. Here, we delineated a new mechanism of 1-dehydro-10-gingerdione (1D10G), one of pungent isolates from ginger (Zingiber officinale), in the suppression of lipopolysaccharide (LPS)-induced gene expression of inflammatory cytokines. 1D10G inhibited LPS binding to MD-2 with higher affinity thanmore » gingerol and shogaol from dietary ginger. Moreover, 1D10G down-regulated TLR4-mediated expression of nuclear factor-{kappa}B (NF-{kappa}B) or activating protein 1 (AP1)-target genes such as tumor necrosis factor {alpha} (TNF-{alpha}) and interleukin-1{beta}, as well as those of interferon (IFN) regulatory factor 3 (IRF3)-target IFN-{beta} gene and IFN-{gamma} inducible protein 10 (IP-10) in LPS-activated macrophages. Taken together, MD-2 is a molecular target in the anti-inflammatory action of 1D10G.« less

Toll-like Receptor 2 Signalling and the Lysosomal Machinery in Barrett's Esophagus.

PubMed

Verbeek, Romy E; Siersema, Peter D; Vleggaar, Frank P; Ten Kate, Fiebo J; Posthuma, George; Souza, Rhonda F; de Haan, Judith; van Baal, Jantine W P M

2016-09-01

Inflammation plays an important role in the development of esophageal adenocarcinoma and its metaplastic precursor lesion, Barrett's esophagus. Toll-like receptor (TLR) 2 signalling and lysosomal function have been linked to inflammation-associated carcinogenesis. We examined the expression of TLR2 in the esophagus and the effect of long-term TLR2 activation on morphological changes and expression of factors involved in lysosomal function in a Barrett's esophagus epithelium cell line. TLR2 expression in normal squamous esophagus, reflux esophagitis, Barrett's esophagus and esophageal adenocarcinoma biopsies was assessed with Q-RT-PCR, in situ hybridization and immunohistochemistry. Barrett's esophagus epithelium cells (BAR-T) were incubated with acid and bile salts in the presence or absence of the TLR2 agonist Pam3CSK4 for a period up to 4 weeks. Morphological changes were assessed with electron microscopy, while Q-RT-PCR was used to determine the expression of lysosomal enzymes (Cathepsin B and C) and factors involved in endocytosis (LAMP-1 and M6PR) and autophagy (LC3 and Rab7). TLR2 was expressed in normal squamous esophagus, reflux esophagitis, Barrett's esophagus but was most prominent in esophageal adenocarcinoma. Long-term TLR2 activation in acid and bile salts exposed BAR-T cells resulted in more and larger lysosomes, more mitochondria and increased expression of LAMP-1, M6PR, Cathepsin B and C when compared to BAR-T cells incubated with acid and bile salts but no TLR2 agonist. Factors associated with autophagy (LC3 and Rab7) expression remained largely unchanged. Activation of TLR2 in acid and bile salts exposed Barrett epithelium cells resulted in an increased number of mitochondria and lysosomes and increased expression of lysosomal enzymes and factors involved in endocytosis.

Expression of CD73/ecto-5'-nucleotidase on human gingival fibroblasts and contribution to the inhibition of interleukin-1alpha-induced granulocyte-macrophage colony stimulating factor production.

PubMed

Nemoto, Eiji; Kunii, Ryotaro; Tada, Hiroyuki; Tsubahara, Taisuke; Ishihata, Hiroshi; Shimauchi, Hidetoshi

2004-02-01

CD73/5'-nucleotidase (5'-NT) is an ectoenzyme that participates in immune/inflammatory reactions. We examined the possible expression of CD73/5'-NT on human gingival fibroblasts (hGF), which are important to the immune/inflammatory system in periodontal tissue. We demonstrated that CD73/5'-NT was expressed on hGF by flow cytometry. We found that pre-treatment of hGF with 5'-AMP induced marked inhibition of granulocyte-macrophage colony-stimulating factor (GM-CSF) production from hGF upon stimulation with interleukin-1alpha (IL-1alpha) by enzyme-linked immunosorbent assay (ELISA). A specific inhibitor of 5'-NT, adenosine 5'-[alpha,beta-methylene] diphosphate blocked the inhibition of GM-CSF production, suggesting that adenosine converted from 5'-AMP acts on the inhibitory effects. The GM-CSF inhibition suggested that A3 receptor might be involved. The rank order of agonists was found to be (N6-benzyl-5'-N-ethylcarboxamidoadenosine) A3 receptor agonist > or = (2-chloroadenosine) non-selective agonist > (CGS-21680) A2A receptor agonist > adenosine > or = (N6-cyclohexyladenosine) A1 agonist. Further support for the main role of A3 receptor was the binding A3 antagonist [9-chloro-2-(2-furanyl)-5-([phenylacetyl]amino)[1,2,4]-triazolo[1,5-c]quinazdine] reversed the effect of adenosine, but no significant reverse was observed by A1 (1,3-dipropyl-8-cyclopentylxanthine), A2 [3,7-dimethyl-1-(2-propargyl)xanthine], A2A[8-(3-chlorostyryl)caffeine], and A2B (alloxazine) antagonists. The CD73/5'-NT expression was increased upon stimulation with gamma-interferon, but not other stimulants such as tumor necrosis factor-alpha, IL-4, lipopolysaccharide from Porphyromonas gingivalis and Escherichia coli, and fimbriae from P. gingivalis, and this increase was correlated with the enhanced GM-CSF inhibition by 5'-AMP but not adenosine. These findings suggested that CD73/5'-NT on hGF exerts an anti-inflammatory effects in periodontal disease by conversion from 5'-AMP to adenosine.

Biochemical Characterization of Individual Human Glycosylated pro-Insulin-like Growth Factor (IGF)-II and big-IGF-II Isoforms Associated with Cancer

PubMed Central

Greenall, Sameer A.; Bentley, John D.; Pearce, Lesley A.; Scoble, Judith A.; Sparrow, Lindsay G.; Bartone, Nicola A.; Xiao, Xiaowen; Baxter, Robert C.; Cosgrove, Leah J.; Adams, Timothy E.

2013-01-01

Insulin-like growth factor II (IGF-II) is a major embryonic growth factor belonging to the insulin-like growth factor family, which includes insulin and IGF-I. Its expression in humans is tightly controlled by maternal imprinting, a genetic restraint that is lost in many cancers, resulting in up-regulation of both mature IGF-II mRNA and protein expression. Additionally, increased expression of several longer isoforms of IGF-II, termed “pro” and “big” IGF-II, has been observed. To date, it is ambiguous as to what role these IGF-II isoforms have in initiating and sustaining tumorigenesis and whether they are bioavailable. We have expressed each individual IGF-II isoform in their proper O-glycosylated format and established that all bind to the IGF-I receptor and both insulin receptors A and B, resulting in their activation and subsequent stimulation of fibroblast proliferation. We also confirmed that all isoforms are able to be sequestered into binary complexes with several IGF-binding proteins (IGFBP-2, IGFBP-3, and IGFBP-5). In contrast to this, ternary complex formation with IGFBP-3 or IGFBP-5 and the auxillary protein, acid labile subunit, was severely diminished. Furthermore, big-IGF-II isoforms bound much more weakly to purified ectodomain of the natural IGF-II scavenging receptor, IGF-IIR. IGF-II isoforms thus possess unique biological properties that may enable them to escape normal sequestration avenues and remain bioavailable in vivo to sustain oncogenic signaling. PMID:23166326

Gper and ESRs are expressed in rat round spermatids and mediate oestrogen-dependent rapid pathways modulating expression of cyclin B1 and Bax.

PubMed

Chimento, A; Sirianni, R; Zolea, F; Bois, C; Delalande, C; Andò, S; Maggiolini, M; Aquila, S; Carreau, S; Pezzi, V

2011-10-01

Spermatogenesis is a precisely controlled and timed process, comprising mitotic divisions of spermatogonia, meiotic divisions of spermatocytes, maturation and differentiation of haploid spermatids giving rise to spermatozoa. It is well known that the maintenance of spermatogenesis is controlled by gonadotrophins and testosterone, the effects of which are modulated by a complex network of locally produced factors, including oestrogens. However, it remains uncertain whether oestrogens are able to activate rapid signalling pathways directly in male germ cells. Classically, oestrogens act by binding to oestrogen receptors (ESRs) 1 and 2. Recently, it has been demonstrated that rapid oestrogen action can also be mediated by the G-protein-coupled oestrogen receptor 1 (Gper). The aim of the present study was to investigate ESRs and Gper expression in primary cultures of adult rat round spermatids (RS) and define if oestradiol (E2) is able to activate, through these receptors, pathways involved in the regulation of genes controlling rat RS apoptosis and/or maturation. In this study, we demonstrated that rat RS express ESR1, ESR2 and Gper. Short-time treatment of RS with E2, the selective Gper agonist G1 and the selective ESR1 and ERβ agonists, 4,4',4"-(4-propyl-[1H]pyrazole-1,3,5-triyl) trisphenol (PPT) and 2,3-bis(4-hydroxyphenyl)-propionitrile (DPN), respectively, determined activation of Extra-cellular signal-regulated kinase (ERK1/2) through the involvement of epidermal growth factor receptor transactivation. In addition, we investigated the effects of ESRs and Gper pathway activation on factors involved in RS maturation. Expression of cyclin B1 mRNA was downregulated by E2, G1 and PPT, but not by DPN. A concomitant and inverse regulation of the pro-apoptotic factor Bax mRNA expression was observed in the same conditions, with DPN being the only one determining an increase in this factor expression. Collectively, these data demonstrate that E2 activates, through ESRs and Gper, pathways involved in the regulation of genes controlling rat RS apoptosis and differentiation such as cyclin B1 and Bax. © 2010 The Authors. International Journal of Andrology © 2011 European Academy of Andrology.

F-prostanoid receptor regulation of fibroblast growth factor 2 signaling in endometrial adenocarcinoma cells.

PubMed

Sales, Kurt J; Boddy, Sheila C; Williams, Alistair R W; Anderson, Richard A; Jabbour, Henry N

2007-08-01

Prostaglandin (PG) F(2alpha) is a potent bioactive lipid in the female reproductive tract, and exerts its function after coupling with its heptahelical G-protein-coupled receptor [F-series-prostanoid (FP) receptor] to initiate cell signaling and target gene transcription. In the present study, we found elevated expression of fibroblast growth factor (FGF) 2, FGF receptor 1 (FGFR1), and FP receptor, colocalized within the neoplastic epithelial cells of endometrial adenocarcinomas. We investigated a role for PGF(2alpha)-FP receptor interaction in modulating FGF2 expression and signaling using an endometrial adenocarcinoma cell line stably expressing the FP receptor to the levels detected in endometrial adenocarcinomas (FPS cells) and endometrial adenocarcinoma tissue explants. PGF(2alpha)-FP receptor activation rapidly induced FGF2 mRNA expression, and elevated FGF2 protein expression and secretion into the culture medium in FPS cells and endometrial adenocarcinoma explants. The effect of PGF(2alpha) on the expression and secretion of FGF2 could be abolished by treatment of FPS cells and endometrial tissues with an FP receptor antagonist (AL8810) and inhibitor of ERK (PD98059). Furthermore, we have shown that FGF2 can promote the expression of FGF2 and cyclooxygenase-2, and enhance proliferation of endometrial adenocarcinoma cells via the FGFR1 and ERK pathways, thereby establishing a positive feedback loop to regulate neoplastic epithelial cell function in endometrial adenocarcinomas.

Bergamottin Promotes Adipocyte Differentiation and Inhibits Tumor Necrosis Factor-α-induced Inflammatory Cytokines Induction in 3T3-L1 Cells.

PubMed

Mizuno, Hideya; Hatano, Tomoko; Taketomi, Ayako; Kawabata, Mami; Nakabayashi, Toshikatsu

2017-01-01

Nowadays, a lot of food ingredients are marketed as dietary supplements for health. Because the effectiveness and mechanisms of these compounds have not been fully characterized, they might have unknown functions. Therefore, we investigated the effect of several food ingredients (Bergamottin, Chrysin, L-Citrulline and β-Carotene) known as health foods on adipocyte differentiation by using 3T3-L1 preadipocytes. In this study, we found that Bergamottin, a furanocoumarin isolated from grapefruit juice, promotes adipocyte differentiation. In addition, Bergamottin increases the expression of adiponectin, an anti-inflammatory adipokine, and peroxisome proliferator activated receptor γ (PPARγ), a nuclear receptor regulating adipocyte differentiation. Furthermore, the anti-inflammatory activity of Bergamottin was demonstrated by its inhibition of the activation of nuclear factor-κB (NF-κB), an inflammatory transcription factor. Stimulation of mature 3T3-L1 adipocytes by tumor necrosis factor-α (TNF-α) decreased the expression of the endogeneous NF-κB inhibitor, IκBα. Treatment with Bergamottin further decreased the TNF-α-induced change in IκBα expression, suggesting that Bergamottin mediated the inhibition of NF-κB activation. In addition, Bergamottin decreased the TNF-α-induced increase in the mRNA levels of pro-inflammatory adipokines, monocyte chemoattractant protein-1 and interleukin-6. Taken together, our results show that Bergamottin treatment could inhibit inflammatory activity through promoting adipocyte differentiation, which in turn suggests that Bergamottin has the potential to minimize the risk factors of metabolic syndrome.

RGS16 inhibits breast cancer cell growth by mitigating phosphatidylinositol 3-kinase signaling.

PubMed

Liang, Genqing; Bansal, Geetanjali; Xie, Zhihui; Druey, Kirk M

2009-08-07

Aberrant activity of the phosphatidylinositol 3-kinase (PI3K) pathway supports growth of many tumors including those of breast, lung, and prostate. Resistance of breast cancer cells to targeted chemotherapies including tyrosine kinase inhibitors (TKI) has been linked to persistent PI3K activity, which may in part be due to increased membrane expression of epidermal growth factor (EGF) receptors (HER2 and HER3). Recently we found that proteins of the RGS (regulator of G protein signaling) family suppress PI3K activity downstream of the receptor by sequestering its p85alpha subunit from signaling complexes. Because a substantial percentage of breast tumors have RGS16 mutations and reduced RGS16 protein expression, we investigated the link between regulation of PI3K activity by RGS16 and breast cancer cell growth. RGS16 overexpression in MCF7 breast cancer cells inhibited EGF-induced proliferation and Akt phosphorylation, whereas shRNA-mediated extinction of RGS16 augmented cell growth and resistance to TKI treatment. Exposure to TKI also reduced RGS16 expression in MCF7 and BT474 cell lines. RGS16 bound the amino-terminal SH2 and inter-SH2 domains of p85alpha and inhibited its interaction with the EGF receptor-associated adapter protein Gab1. These results suggest that the loss of RGS16 in some breast tumors enhances PI3K signaling elicited by growth factors and thereby promotes proliferation and TKI evasion downstream of HER activation.

Expression of the vascular endothelial growth factor receptor neuropilin-1 at the human embryo-maternal interface.

PubMed

Baston-Buest, Dunja M; Porn, Anne C; Schanz, Andrea; Kruessel, Jan-S; Janni, Wolfgang; Hess, Alexandra P

2011-02-01

Angiogenesis is required for successful implantation of the invading blastocyst. Vascular endothelial growth factor (VEGF) is an important key player in angiogenesis and vascular remodeling during the implantation process. Besides its well-characterized receptors VEGFR1 and VEGFR2, neuropilin-1 (NRP-1) has been shown to play an additional role in the signaling process of angiogenesis in human endometrium during the menstrual cycle, as a co-receptor of VEGF. These findings led to the hypothesis that NRP-1 might play a role in the vascular remodeling process during embryo implantation and the establishment of a pregnancy. NRP-1 mRNA transcript and protein expression were investigated in human choriocarcinoma cell lines (JEG-3, Jar and BeWo) aiming to evaluate the expression of NRP-1 in vitro, as well as in human decidua of all three trimesters of pregnancy, by western blot analysis (three samples of each trimester of pregnancy). The localization of NRP-1 in human decidua of all three trimesters of pregnancy was analyzed by immunohistochemistry (five samples of each trimester of pregnancy). NRP-1 transcript and protein were expressed in all cell lines examined. Corresponding to the analysis of human tissue by western blot and the localization by immunohistochemistry, NRP-1 protein higher expressed in samples of early pregnancy in comparison to the end of pregnancy. NRP-1 was expressed in the decidua, villi and invading cytotrophoblast of all samples investigated. This is the first study clearly showing the expression of NRP-1 in human decidua and trophoblast, suggesting an important role for the VEGF co-receptor NRP-1 besides the established receptor VEGFR2 at the embryo-maternal interface during embryonic implantation and placentation. Copyright © 2010 Elsevier Ireland Ltd. All rights reserved.

Effects of Secreted Mast Cell Mediators on Retinal Pigment Epithelial Cells: Focus on Mast Cell Tryptase.

PubMed

Arai, Rei; Usui-Ouchi, Ayumi; Ito, Yosuke; Mashimo, Keitaro; Murakami, Akira; Ebihara, Nobuyuki

2017-01-01

Numerous mast cells are present in the choroid, but the effects of mast cell mediators on retinal pigment epithelial (RPE) cells are not well understood. We investigated the influence of mast cell mediators on RPE cells in vitro, focusing on tryptase. Expression of receptors was examined by the reverse transcription polymerase chain reaction. We also assessed production of interleukin 8 and vascular endothelial growth factor (VEGF) after RPE cells were stimulated with mast cell mediators by using an antibody array and enzyme-linked immunosorbent assay. Furthermore, we investigated the influence of tryptase on RPE cell migration and integrity by the scratch assay and the transepithelial resistance. RPE cells expressed protease-activated receptor 2 (PAR2), histamine receptor 1, tumor necrosis factor- α (TNF- α ) receptor 1, and CCR 1, 3, 4, 8, and 11. Tryptase, PAR2 agonists, histamine, and TNF- α all enhanced interleukin 8 production by RPE cells, while only tryptase enhanced VEGF production. Tryptase also enhanced expression of phosphorylated extracellular signal-regulated kinases 1/2, resulting in increased migration of RPE cells. However, tryptase did not alter epithelial integrity or the expression of zonula occludens-1 and junctional adhesion molecule-A by RPE cells. Mast cell mediators, especially tryptase, may influence RPE cell inflammation.

Early activation of quorum sensing in Pseudomonas aeruginosa reveals the architecture of a complex regulon.

PubMed

Schuster, Martin; Greenberg, E Peter

2007-08-22

Quorum-sensing regulation of gene expression in Pseudomonas aeruginosa is complex. Two interconnected acyl-homoserine lactone (acyl-HSL) signal-receptor pairs, 3-oxo-dodecanoyl-HSL-LasR and butanoyl-HSL-RhlR, regulate more than 300 genes. The induction of most of the genes is delayed during growth of P. aeruginosa in complex medium, cannot be advanced by addition of exogenous signal, and requires additional regulatory components. Many of these late genes can be induced by addition of signals early by using specific media conditions. While several factors super-regulate the quorum receptors, others may co-regulate target promoters or may affect expression posttranscriptionally. To better understand the contributions of super-regulation and co-regulation to quorum-sensing gene expression, and to better understand the general structure of the quorum sensing network, we ectopically expressed the two receptors (in the presence of their cognate signals) and another component that affects quorum sensing, the stationary phase sigma factor RpoS, early in growth. We determined the effect on target gene expression by microarray and real-time PCR analysis. Our results show that many target genes (e.g. lasB and hcnABC) are directly responsive to receptor protein levels. Most genes (e.g. lasA, lecA, and phnAB), however, are not significantly affected, although at least some of these genes are directly regulated by quorum sensing. The majority of promoters advanced by RhlR appeared to be regulated directly, which allowed us to build a RhlR consensus sequence. The direct responsiveness of many quorum sensing target genes to receptor protein levels early in growth confirms the role of super-regulation in quorum sensing gene expression. The observation that the induction of most target genes is not affected by signal or receptor protein levels indicates that either target promoters are co-regulated by other transcription factors, or that expression is controlled posttranscriptionally. This architecture permits the integration of multiple signaling pathways resulting in quorum responses that require a "quorum" but are otherwise highly adaptable and receptive to environmental conditions.

Endocrine gland-derived vascular endothelial growth factor (EG-VEGF) and its receptor PROKR2 are associated to human colorectal cancer progression and peritoneal carcinomatosis.

PubMed

Benlahfid, Mohammed; Traboulsi, Wael; Sergent, Frederic; Benharouga, Mohamed; Elhattabi, Khalid; Erguibi, Driss; Karkouri, Mehdi; Elattar, Hicham; Fadil, Abdelaziz; Fahmi, Yassine; Aboussaouira, Touria; Alfaidy, Nadia

2018-02-06

The highest risk factor for mortality among malignant tumors is metastasis. Endocrine gland-derived vascular endothelial growth factor (EG-VEGF) is an angiogenic factor which biological activity is mediated via two G protein-coupled receptors, prokineticin receptor1 (PROKR1) and PROKR2. Recent studies suggested that EG-VEGF expression is deregulated in multiple cancers including colorectal cancer (CRC). Using distinctive CRC and peritoneal carcinomatosis (PC) cohorts and a corresponding control cohort, we determined the circulating levels of EG-VEGF and its in situ expression, and that of its related receptors. Circulating EG-VEGF levels were significantly increased in patients with metastatic PC compared to CRC and control patients (p< 0.05). Furthermore, according to clinicopathologic examinations, local EG-VEGF expression correlated with higher tumor and nodal stages (p< 0.001) of CRC. EG-VEGF and PROKR2 were highly expressed in colorectal primary lesions compared to positive controls. PROKR1 expression was lower and did not change in tumor specimens. Also, EG-VEGF and its receptor PROKR2 were differentially expressed in the colorectal primary lesions and in the control groups. Altogether these findings suggest that EG-VEGF/receptors system might be an important actor in the CRC progression into PC and might be involved in the ability of tumor cells to invade other organs. Circulating EG-VEGF could be proposed as a prognostic marker in human CRC and its progression into PC.

Transient Receptor Potential Melastatin-3 (TRPM3) Mediates Nociceptive-Like Responses in Hydra vulgaris

PubMed Central

Malafoglia, Valentina; Traversetti, Lorenzo; Del Grosso, Floriano; Scalici, Massimiliano; Lauro, Filomena; Russo, Valeria; Persichini, Tiziana; Salvemini, Daniela; Mollace, Vincenzo; Fini, Massimo; Raffaeli, William

2016-01-01

The ability of mammals to feel noxious stimuli lies in a heterogeneous group of primary somatosensory neurons termed nociceptors, which express specific membrane receptors, such as the Transient Receptor Potential (TRP) family. Here, we show that one of the most important nociceptive-like pathways is conserved in the freshwater coelenterate Hydra vulgaris, the most primitive organism possessing a nervous system. In particular, we found that H. vulgaris expresses TRPM3, a nociceptor calcium channel involved in the detection of noxious heat in mammals. Furthermore, we detected that both heat shock and TRPM3 specific agonist (i.e., pregnenolone sulfate) induce the modulation of the heat shock protein 70 (HSP70) and the nitric oxide synthase (NOS), two genes activated by TRP-mediated heat painful stimuli in mammals. As expected, these effects are inhibited by a TRPM3 antagonist (i.e., mefenamic acid). Interestingly, the TRPM3 agonist and heat shock also induce the expression of nuclear transcription erythroid 2-related factor (Nrf2) and superoxide dismutase (SOD), known markers of oxidative stress; noteworthy gene expression was also inhibited by the TRPM3 antagonist. As a whole, our results demonstrate the presence of conserved molecular oxidative/nociceptive-like pathways at the primordial level of the animal kingdom. PMID:26974325

Transient Receptor Potential Melastatin-3 (TRPM3) Mediates Nociceptive-Like Responses in Hydra vulgaris.

PubMed

Malafoglia, Valentina; Traversetti, Lorenzo; Del Grosso, Floriano; Scalici, Massimiliano; Lauro, Filomena; Russo, Valeria; Persichini, Tiziana; Salvemini, Daniela; Mollace, Vincenzo; Fini, Massimo; Raffaeli, William; Muscoli, Carolina; Colasanti, Marco

2016-01-01

The ability of mammals to feel noxious stimuli lies in a heterogeneous group of primary somatosensory neurons termed nociceptors, which express specific membrane receptors, such as the Transient Receptor Potential (TRP) family. Here, we show that one of the most important nociceptive-like pathways is conserved in the freshwater coelenterate Hydra vulgaris, the most primitive organism possessing a nervous system. In particular, we found that H. vulgaris expresses TRPM3, a nociceptor calcium channel involved in the detection of noxious heat in mammals. Furthermore, we detected that both heat shock and TRPM3 specific agonist (i.e., pregnenolone sulfate) induce the modulation of the heat shock protein 70 (HSP70) and the nitric oxide synthase (NOS), two genes activated by TRP-mediated heat painful stimuli in mammals. As expected, these effects are inhibited by a TRPM3 antagonist (i.e., mefenamic acid). Interestingly, the TRPM3 agonist and heat shock also induce the expression of nuclear transcription erythroid 2-related factor (Nrf2) and superoxide dismutase (SOD), known markers of oxidative stress; noteworthy gene expression was also inhibited by the TRPM3 antagonist. As a whole, our results demonstrate the presence of conserved molecular oxidative/nociceptive-like pathways at the primordial level of the animal kingdom.

MyD88 expression in the rat dental follicle: Implications for osteoclastogenesis and tooth eruption

PubMed Central

Liu, Dawen; Yao, Shaomian; Wise, Gary E.

2010-01-01

Myeloid differentiation factor 88 (MyD88) is a key adaptor molecule in the interleukin-1 (IL-1) and IL-18 Toll-like receptor signaling pathway. Because it is present in dental follicle (DF) cells in vitro, the purpose of this study was to determine its chronological expression in vivo, as well as its possible role in osteoclastogenesis and tooth eruption. An oligo DNA microarray was used to determine gene expression of MyD88 in vivo in the DFs from the first mandibular molars of postnatal rats from days 1–11. The results showed that MyD88 was expressed maximally at day 3. Using siRNA to knock down MyD88 expression in the DF cells also reduced the gene expression of nuclear factor-kappa B-1 (NFKB1) and monocyte chemoattractant protein 1 (MCP-1). IL-1α up-regulated the expression of NFKB1, MCP-1 and receptor activator of nuclear factor kappa B ligand (RANKL), but knockdown of MyD88 nullified this IL-1α effect. Conditioned medium from DF cells with MyD88 knocked down reduced chemotactic activity for mononuclear cells and reduced osteoclastogenesis as opposed to controls. In conclusion, the maximal expression of MyD88 at day 3 in the DF may contribute to the major burst of osteoclastogenesis needed for eruption by up-regulating MCP-1 and RANKL expression. PMID:20662905

Binding of the Ras activator son of sevenless to insulin receptor substrate-1 signaling complexes.

PubMed

Baltensperger, K; Kozma, L M; Cherniack, A D; Klarlund, J K; Chawla, A; Banerjee, U; Czech, M P

1993-06-25

Signal transmission by insulin involves tyrosine phosphorylation of a major insulin receptor substrate (IRS-1) and exchange of Ras-bound guanosine diphosphate for guanosine triphosphate. Proteins containing Src homology 2 and 3 (SH2 and SH3) domains, such as the p85 regulatory subunit of phosphatidylinositol-3 kinase and growth factor receptor-bound protein 2 (GRB2), bind tyrosine phosphate sites on IRS-1 through their SH2 regions. Such complexes in COS cells were found to contain the heterologously expressed putative guanine nucleotide exchange factor encoded by the Drosophila son of sevenless gene (dSos). Thus, GRB2, p85, or other proteins with SH2-SH3 adapter sequences may link Sos proteins to IRS-1 signaling complexes as part of the mechanism by which insulin activates Ras.

[Influence of macrophages on the expression of vascular endothelial growth factor receptor mRNA, homeobox B2 mRNA, and integrin alpha nu beta3 in vascular endothelial strain].

PubMed

Liu, Liang; Liu, Chang; Zhang, Xiao-qi; Ming, Jia; Liu, Xu-sheng; Xu, Hui; Cheng, Tian-min

2005-06-01

To investigate the influence of macrophages on the expression of the vascular endothelial growth factor (VEGF) receptor (KDR) mRNA, homeobox B2 (HOXB2) mRNA, and integrin alpha nu beta3 in vitro in vascular endothelial strain. Human umbilical vein cells (ECV304) were cultured in vitro and divided into 4 groups, i.e. (1) ECV304 group, (2) ECV304 + conA group [with conA (25 microg/ml in culture) added to ECV304], (3) ECV304 + U937 group (with 1 x 10(5)/ml of U937 cells added to 1 x 10(5)/ml ECV 304), (4) ECV304 + U937 + conA group [with 1 x 10(5)/ml of U937 cells and conA (25 microg/ml in culture)] groups. Forty-eight hours after culturing, the expression of integrin receptor alpha nu beta3 and the changes in the expression of KDR mRNA and HOXB2 mRNA in each group were determined by immunofluorescent technique and RT-PCR, respectively. The expression of integrin receptor alpha nu beta3, KDR mRNA, and HOXB2 mRNA in ECV304 group were 6.7 +/- 1.5, 0.633 +/- 0.012, and 0.674 +/- 0.004, respectively, while those in ECV304 + U937 + conA group (10.2 +/- 1.7, 0.879 +/- 0.003, 0.947 +/- 0.003) were obviously more upregulated when compared with those in ECV304 group (P < 0.01). No difference in the above indices was found between ECV304 and ECV304 + conA, ECV304 + U937 groups (P > 0.05). Macrophages activated by ConA can accelerate the proliferation, migration and adhesion to the basement membrane matrix of vascular endothelial cells through the influence on the expression of KDR mRNA, HOXB2 mRNA and integrin alpha nu beta3, and through this pathway the angiogenesis is modulated.

The orphan receptor hepatic nuclear factor 4 functions as a transcriptional activator for tissue-specific and hypoxia-specific erythropoietin gene expression and is antagonized by EAR3/COUP-TF1.

PubMed

Galson, D L; Tsuchiya, T; Tendler, D S; Huang, L E; Ren, Y; Ogura, T; Bunn, H F

1995-04-01

The erythropoietin (Epo) gene is regulated by hypoxia-inducible cis-acting elements in the promoter and in a 3' enhancer, both of which contain consensus hexanucleotide hormone receptor response elements which are important for function. A group of 11 orphan nuclear receptors, transcribed and translated in vitro, were screened by the electrophoretic mobility shift assay. Of these, hepatic nuclear factor 4 (HNF-4), TR2-11, ROR alpha 1, and EAR3/COUP-TF1 bound specifically to the response elements in the Epo promoter and enhancer and, except for ROR alpha 1, formed DNA-protein complexes that had mobilities similar to those observed in nuclear extracts of the Epo-producing cell line Hep3B. Moreover, both anti-HNF-4 and anti-COUP antibodies were able to supershift complexes in Hep3B nuclear extracts. Like Epo, HNF-4 is expressed in kidney, liver, and Hep3B cells but not in HeLa cells. Transfection of a plasmid expressing HNF-4 into HeLa cells enabled an eightfold increase in the hypoxic induction of a luciferase reporter construct which contains the minimal Epo enhancer and Epo promoter, provided that the nuclear hormone receptor consensus DNA elements in both the promoter and the enhancer were intact. The augmentation by HNF-4 in HeLa cells could be abrogated by cotransfection with HNF-4 delta C, which retains the DNA binding domain of HNF-4 but lacks the C-terminal activation domain. Moreover, the hypoxia-induced expression of the endogenous Epo gene was significantly inhibited in Hep3B cells stably transfected with HNF-4 delta C. On the other hand, cotransfection of EAR3/COUP-TF1 and the Epo reporter either with HNF-4 into HeLa cells or alone into Hep3B cells suppressed the hypoxia induction of the Epo reporter. These electrophoretic mobility shift assay and functional experiments indicate that HNF-4 plays a critical positive role in the tissue-specific and hypoxia-inducible expression of the Epo gene, whereas the COUP family has a negative modulatory role.

Reduced Chrna7 expression in C3H mice is associated with increases in hippocampal parvalbumin and glutamate decarboxylase-67 (GAD67) as well as altered levels of GABAA receptor subunits

PubMed Central

Bates, Ryan C.; Stith, Bradley J.; Stevens, Karen E.; Adams, Catherine E.

2014-01-01

Decreased expression of CHRNA7, the gene encoding the α7* subtype of nicotinic receptor, may contribute to the cognitive dysfunction observed in schizophrenia by disrupting the inhibitory/excitatory balance in the hippocampus. C3H mice with reduced Chrna7 expression have significant reductions in hippocampal α7* receptor density, deficits in hippocampal auditory gating, increased hippocampal activity as well as significant decreases in hippocampal glutamate decarboxylase-65 (GAD65) and γ-aminobutyric acid-A (GABAA) receptor levels. The current study investigated whether altered Chrna7 expression is associated with changes in the levels of parvalbumin, GAD67 and/or GABAA receptor subunits in hippocampus from male and female C3H Chrna7 wildtype, C3H Chrna7 heterozygous and C3H Chrna7 knockout mice using quantitative western immunoblotting. Reduced Chrna7 expression was associated with significant increases in hippocampal parvalbumin and GAD67 and with complex alterations in GABAA receptor subunits. A decrease in α3 subunit protein was seen in both female C3H Chrna7 Het and KO mice while a decrease in α4 subunit protein was also detected in C3H Chrna7 KO mice with no sex difference. In contrast, an increase in δ subunit protein was observed in C3H Chrna7 Het mice while a decrease in this subunit was observed in C3H Chrna7 KO mice, with δ subunit protein levels being greater in males than in females. Finally, an increase in γ2 subunit protein was found in C3H Chrna7 KO mice with the levels of this subunit again being greater in males than in females. The increases in hippocampal parvalbumin and GAD67 observed in C3H Chrna7 mice are contrary to reports of reductions in these proteins in postmortem hippocampus from schizophrenic individuals. We hypothesize that the disparate results may occur because of the influence of factors other than CHRNA7 that have been found to be abnormal in schizophrenia. PMID:24836856

Regulatory T cells protect mice against coxsackievirus-induced myocarditis through the transforming growth factor beta-coxsackie-adenovirus receptor pathway.

PubMed

Shi, Yu; Fukuoka, Masahiro; Li, Guohua; Liu, Youan; Chen, Manyin; Konviser, Michael; Chen, Xin; Opavsky, Mary Anne; Liu, Peter P

2010-06-22

Coxsackievirus B3 infection is an excellent model of human myocarditis and dilated cardiomyopathy. Cardiac injury is caused either by a direct cytopathic effect of the virus or through immune-mediated mechanisms. Regulatory T cells (Tregs) play an important role in the negative modulation of host immune responses and set the threshold of autoimmune activation. This study was designed to test the protective effects of Tregs and to determine the underlying mechanisms. Carboxyfluorescein diacetate succinimidyl ester-labeled Tregs or naïve CD4(+) T cells were injected intravenously once every 2 weeks 3 times into mice. The mice were then challenged with intraperitoneal coxsackievirus B3 immediately after the last cell transfer. Transfer of Tregs showed higher survival rates than transfer of CD4(+) T cells (P=0.0136) but not compared with the PBS injection group (P=0.0589). Interestingly, Tregs also significantly decreased virus titers and inflammatory scores in the heart. Transforming growth factor-beta and phosphorylated AKT were upregulated in Tregs-transferred mice and coxsackie-adenovirus receptor expression was decreased in the heart compared with control groups. Transforming growth factor-beta decreased coxsackie-adenovirus receptor expression and inhibited coxsackievirus B3 infection in HL-1 cells and neonatal cardiac myocytes. Splenocytes collected from Treg-, CD4(+) T-cell-, and PBS-treated mice proliferated equally when stimulated with heat-inactivated virus, whereas in the Treg group, the proliferation rate was reduced significantly when stimulated with noninfected heart tissue homogenate. Adoptive transfer of Tregs protected mice from coxsackievirus B3-induced myocarditis through the transforming growth factor beta-coxsackie-adenovirus receptor pathway and thus suppresses the immune response to cardiac tissue, maintaining the antiviral immune response.

Type 1 receptor tyrosine kinases are differentially phosphorylated in mammary carcinoma and differentially associated with steroid receptors.

PubMed Central

Bacus, S. S.; Chin, D.; Yarden, Y.; Zelnick, C. R.; Stern, D. F.

1996-01-01

The neu/erbB-2/HER-2 proto-oncogene is amplified and/or overexpressed in up to 30% of mammary carcinomas and has been variably correlated with poor prognosis. The signaling activity of the encoded receptor tyrosine kinase is regulated by interactions with other type 1 receptors and their ligands. We have used a novel approach, phosphorylation-sensitive anti-Neu antibodies, to quantify signaling by Neu and epidermal growth factor receptor in a panel of frozen sections of mammary carcinoma specimens. We also determined the relationship of Neu, phosphorylated Neu (and epidermal growth factor receptor), and phosphotyrosine to the expression of Neu-related receptors (epidermal growth factor receptor, HER-3, and HER-4) and to prognostic factors (estrogen and progesterone receptor). We found that tyrosine phosphorylation of Neu (and hence signaling activity) is highly variable among mammary carcinomas. Neu and HER-4 were associated with divergent correlates, suggesting that they have profoundly different biological activities. These results have implications for etiology of mammary carcinoma for clinical evaluation of mammary carcinoma patients, and for development of Neu-targeted therapeutic strategies. Images Figure 1 Figure 2 PMID:8579117

IRS-1: essential for insulin- and IL-4-stimulated mitogenesis in hematopoietic cells.

PubMed

Wang, L M; Myers, M G; Sun, X J; Aaronson, S A; White, M; Pierce, J H

1993-09-17

Although several interleukin-3 (IL-3)-dependent cell lines proliferate in response to IL-4 or insulin, the 32D line does not. Insulin and IL-4 sensitivity was restored to 32D cells by expression of IRS-1, the principal substrate of the insulin receptor. Although 32D cells possessed receptors for both factors, they lacked the IRS-1--related protein, 4PS, which becomes phosphorylated by tyrosine in insulin- or IL-4--responsive lines after stimulation. These results indicate that factors that bind unrelated receptors can use similar mitogenic signaling pathways in hematopoietic cells and that 4PS and IRS-1 are functionally similar proteins that are essential for insulin- and IL-4--induced proliferation.

Keratinocyte growth factor is a growth factor for mammary epithelium in vivo. The mammary epithelium of lactating rats is resistant to the proliferative action of keratinocyte growth factor.

PubMed Central

Ulich, T. R.; Yi, E. S.; Cardiff, R.; Yin, S.; Bikhazi, N.; Biltz, R.; Morris, C. F.; Pierce, G. F.

1994-01-01

Keratinocyte growth factor (KGF) is a member of the fibroblast growth factor (FGF) family. KGF is secreted by stromal cells and affects epithelial but not mesenchymal cell proliferation. KGF injected intravenously was found to cause dramatic proliferation of mammary epithelium in the mammary glands of rats. KGF causes ductal neogenesis and intraductal epithelial hyperplasia but not lobular differentiation in nulliparous female rats. KGF causes ductal and lobular epithelial hyperplasia in male rats. KGF causes proliferation of ductal and acinar cells in the mammary glands of pregnant rats. On the other hand, the ductal epithelium of lactating postpartum rats is resistant to the proliferative action of KGF. The mammary glands of lactating rats did not express less KGF receptor mRNA than the glands of pregnant rats, suggesting that the resistance of the ductal epithelium to KGF during lactation is not related to KGF receptor mRNA down-regulation. The mammary glands of both pregnant and postpartum lactating rats express KGF mRNA with more KGF present in the glands of lactating rats. In conclusion, the KGF and KGF receptor genes are expressed in rat mammary glands and recombinant KGF is a potent growth factor for mammary epithelium. Images Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 PMID:8178937

Poly(ADP-ribose) polymerase inhibitors sensitize cancer cells to death receptor-mediated apoptosis by enhancing death receptor expression.

PubMed

Meng, X Wei; Koh, Brian D; Zhang, Jin-San; Flatten, Karen S; Schneider, Paula A; Billadeau, Daniel D; Hess, Allan D; Smith, B Douglas; Karp, Judith E; Kaufmann, Scott H

2014-07-25

Recombinant human tumor necrosis factor-α-related apoptosis inducing ligand (TRAIL), agonistic monoclonal antibodies to TRAIL receptors, and small molecule TRAIL receptor agonists are in various stages of preclinical and early phase clinical testing as potential anticancer drugs. Accordingly, there is substantial interest in understanding factors that affect sensitivity to these agents. In the present study we observed that the poly(ADP-ribose) polymerase (PARP) inhibitors olaparib and veliparib sensitize the myeloid leukemia cell lines ML-1 and K562, the ovarian cancer line PEO1, non-small cell lung cancer line A549, and a majority of clinical AML isolates, but not normal marrow, to TRAIL. Further analysis demonstrated that PARP inhibitor treatment results in activation of the FAS and TNFRSF10B (death receptor 5 (DR5)) promoters, increased Fas and DR5 mRNA, and elevated cell surface expression of these receptors in sensitized cells. Chromatin immunoprecipitation demonstrated enhanced binding of the transcription factor Sp1 to the TNFRSF10B promoter in the presence of PARP inhibitor. Knockdown of PARP1 or PARP2 (but not PARP3 and PARP4) not only increased expression of Fas and DR5 at the mRNA and protein level, but also recapitulated the sensitizing effects of the PARP inhibition. Conversely, Sp1 knockdown diminished the PARP inhibitor effects. In view of the fact that TRAIL is part of the armamentarium of natural killer cells, these observations identify a new facet of PARP inhibitor action while simultaneously providing the mechanistic underpinnings of a novel therapeutic combination that warrants further investigation.

Platelet-derived growth factor receptors differentially inform intertumoral and intratumoral heterogeneity

PubMed Central

Kim, Youngmi; Kim, Eunhee; Wu, Qiulian; Guryanova, Olga; Hitomi, Masahiro; Lathia, Justin D.; Serwanski, David; Sloan, Andrew E.; Weil, Robert J.; Lee, Jeongwu; Nishiyama, Akiko; Bao, Shideng; Hjelmeland, Anita B.; Rich, Jeremy N.

2012-01-01

Growth factor-mediated proliferation and self-renewal maintain tissue-specific stem cells and are frequently dysregulated in cancers. Platelet-derived growth factor (PDGF) ligands and receptors (PDGFRs) are commonly overexpressed in gliomas and initiate tumors, as proven in genetically engineered models. While PDGFRα alterations inform intertumoral heterogeneity toward a proneural glioblastoma (GBM) subtype, we interrogated the role of PDGFRs in intratumoral GBM heterogeneity. We found that PDGFRα is expressed only in a subset of GBMs, while PDGFRβ is more commonly expressed in tumors but is preferentially expressed by self-renewing tumorigenic GBM stem cells (GSCs). Genetic or pharmacological targeting of PDGFRβ (but not PDGFRα) attenuated GSC self-renewal, survival, tumor growth, and invasion. PDGFRβ inhibition decreased activation of the cancer stem cell signaling node STAT3, while constitutively active STAT3 rescued the loss of GSC self-renewal caused by PDGFRβ targeting. In silico survival analysis demonstrated that PDGFRB informed poor prognosis, while PDGFRA was a positive prognostic factor. Our results may explain mixed clinical responses of anti-PDGFR-based approaches and suggest the need for integration of models of cancer as an organ system into development of cancer therapies. PMID:22661233

Gefitinib enhances human colon cancer cells to TRAIL-induced apoptosis of via autophagy- and JNK-mediated death receptors upregulation.

PubMed

Chen, Lei; Meng, Yue; Guo, Xiaoqing; Sheng, Xiaotong; Tai, Guihua; Zhang, Fenglei; Cheng, Hairong; Zhou, Yifa

2016-11-01

Tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) is a potent cancer cell-specific apoptosis-inducing cytokine with little toxicity to most normal cells. Here, we report that gefitinib and TRAIL in combination produce a potent synergistic effect on TRAIL-sensitive human colon cancer HCT116 cells and an additive effect on TRAIL-resistant HT-29 cells. Interestingly, gefitinib increases the expression of cell surface receptors DR4 and DR5, possibly explaining the synergistic effect. Knockdown of DR4 and DR5 by siRNA significantly decreases gefitinib- and TRAIL-mediated cell apoptosis, supporting this idea. Because the inhibition of gefitinib-induced autophagy by 3-MA significantly decreases DR4 and DR5 upregulation, as well as reduces gefitinib- and TRAIL-induced apoptosis, we conclude that death receptor upregulation is autophagy mediated. Furthermore, our results indicate that death receptor expression may also be regulated by JNK activation, because pre-treatment of cells with JNK inhibitor SP600125 significantly decreases gefitinib-induced death receptor upregulation. Interestingly, SP600125 also inhibits the expression CHOP, yet CHOP has no impact on death receptor expressions. We also find here that phosphorylation of Akt and ERK might also be required for TRAIL sensitization. In summary, our results indicate that gefitinib effectively enhances TRAIL-induced apoptosis, likely via autophagy and JNK- mediated death receptor expression and phosphorylation of Akt and ERK.

Nck-2, a Novel Src Homology2/3-containing Adaptor Protein That Interacts with the LIM-only Protein PINCH and Components of Growth Factor Receptor Kinase-signaling Pathways

PubMed Central

Tu, Yizeng; Li, Fugang; Wu, Chuanyue

1998-01-01

Many of the protein–protein interactions that are essential for eukaryotic intracellular signal transduction are mediated by protein binding modules including SH2, SH3, and LIM domains. Nck is a SH3- and SH2-containing adaptor protein implicated in coordinating various signaling pathways, including those of growth factor receptors and cell adhesion receptors. We report here the identification, cloning, and characterization of a widely expressed, Nck-related adaptor protein termed Nck-2. Nck-2 comprises primarily three N-terminal SH3 domains and one C-terminal SH2 domain. We show that Nck-2 interacts with PINCH, a LIM-only protein implicated in integrin-linked kinase signaling. The PINCH-Nck-2 interaction is mediated by the fourth LIM domain of PINCH and the third SH3 domain of Nck-2. Furthermore, we show that Nck-2 is capable of recognizing several key components of growth factor receptor kinase-signaling pathways including EGF receptors, PDGF receptor-β, and IRS-1. The association of Nck-2 with EGF receptors was regulated by EGF stimulation and involved largely the SH2 domain of Nck-2, although the SH3 domains of Nck-2 also contributed to the complex formation. The association of Nck-2 with PDGF receptor-β was dependent on PDGF activation and was mediated solely by the SH2 domain of Nck-2. Additionally, we have detected a stable association between Nck-2 and IRS-1 that was mediated primarily via the second and third SH3 domain of Nck-2. Thus, Nck-2 associates with PINCH and components of different growth factor receptor-signaling pathways via distinct mechanisms. Finally, we provide evidence indicating that a fraction of the Nck-2 and/or Nck-1 proteins are associated with the cytoskeleton. These results identify a novel Nck-related SH2- and SH3-domain–containing protein and suggest that it may function as an adaptor protein connecting the growth factor receptor-signaling pathways with the integrin-signaling pathways. PMID:9843575

Hepatitis C virus utilizes VLDLR as a novel entry pathway.

PubMed

Ujino, Saneyuki; Nishitsuji, Hironori; Hishiki, Takayuki; Sugiyama, Kazuo; Takaku, Hiroshi; Shimotohno, Kunitada

2016-01-05

Various host factors are involved in the cellular entry of hepatitis C virus (HCV). In addition to the factors previously reported, we discovered that the very-low-density lipoprotein receptor (VLDLR) mediates HCV entry independent of CD81. Culturing Huh7.5 cells under hypoxic conditions significantly increased HCV entry as a result of the expression of VLDLR, which was not expressed under normoxic conditions in this cell line. Ectopic VLDLR expression conferred susceptibility to HCV entry of CD81-deficient Huh7.5 cells. Additionally, VLDLR-mediated HCV entry was not affected by the knockdown of cellular factors known to act as HCV receptors or HCV entry factors. Because VLDLR is expressed in primary human hepatocytes, our results suggest that VLDLR functions in vivo as an HCV receptor independent of canonical CD81-mediated HCV entry.

Heregulin/ErbB3 Signaling Enhances CXCR4-Driven Rac1 Activation and Breast Cancer Cell Motility via Hypoxia-Inducible Factor 1α

PubMed Central

Lopez-Haber, Cynthia; Barrio-Real, Laura; Casado-Medrano, Victoria

2016-01-01

The growth factor heregulin (HRG), a ligand of ErbB3 and ErbB4 receptors, contributes to breast cancer development and the promotion of metastatic disease, and its expression in breast tumors has been associated with poor clinical outcome and resistance to therapy. In this study, we found that breast cancer cells exposed to sustained HRG treatment show markedly enhanced Rac1 activation and migratory activity in response to the CXCR4 ligand SDF-1/CXCL12, effects mediated by P-Rex1, a Rac-guanine nucleotide exchange factor (GEF) aberrantly expressed in breast cancer. Notably, HRG treatment upregulates surface expression levels of CXCR4, a G protein-coupled receptor (GPCR) implicated in breast cancer metastasis and an indicator of poor prognosis in breast cancer patients. A detailed mechanistic analysis revealed that CXCR4 upregulation and sensitization of the Rac response/motility by HRG are mediated by the transcription factor hypoxia-inducible factor 1α (HIF-1α) via ErbB3 and independently of ErbB4. HRG caused prominent induction in the nuclear expression of HIF-1α, which transcriptionally activates the CXCR4 gene via binding to a responsive element located in positions −1376 to −1372 in the CXCR4 promoter, as revealed by mutagenesis analysis and chromatin immunoprecipitation (ChIP). Our results uncovered a novel function for ErbB3 in enhancing breast cancer cell motility and sensitization of the P-Rex1/Rac1 pathway through HIF-1α-mediated transcriptional induction of CXCR4. PMID:27185877

Differential activity of 2-methylene-19-nor vitamin D analogs on growth factor gene expression in rhino mouse skin and comparison to all-trans retinoic acid.

PubMed

Ahrens, Jamie M; Jones, James D; Nieves, Nirca J; Mitzey, Ann M; DeLuca, Hector F; Clagett-Dame, Margaret

2017-01-01

While all 2-methylene-19-nor analogs of 1α,25-dihydroxyvitamin D3 (1α,25(OH)2D3) tested produce an increase in epidermal thickness in the rhino mouse, only a subset reduce utricle size (comedolysis). All-trans retinoic acid (atRA) also causes epidermal thickening and a reduction in utricle size in the rhino mouse. We now report that 2-methylene-19-nor-(20S)-1α-hydroxybishomopregnacalciferol (2MbisP), a comedolytic analog, increases epidermal thickening more rapidly than does atRA, while both reduce utricle area at an equal rate. Whereas unlike atRA, 2MbisP does not alter the epidermal growth factor receptor ligand, heparin-binding epidermal growth factor-like growth factor, it does increase the expression of both amphiregulin and epigen mRNA, even after a single dose. In situ hybridization reveals an increase in these transcripts throughout the closing utricle as well as in the interfollicular epidermis. The mRNAs for other EGFR ligands including betacellulin and transforming growth factor-α, as well as the epidermal growth factor receptor are largely unaffected by 2MbisP. Another analog, 2-methylene-19-nor-(20S)-26,27-dimethylene-1α,25-dihydroxyvitamin D3 (CAGE-3), produces epidermal thickening but fails to reduce utricle size or increase AREG mRNA levels. CAGE-3 modestly increases epigen mRNA levels, but only after 5 days of dosing. Thus, 2-MbisP produces unique changes in epidermal growth factor receptor ligand mRNAs that may be responsible for both epidermal proliferation and a reduction in utricle size.

Correlation between VEGFR-2 receptor kinase domain-containing receptor (KDR) mRNA and angiotensin II receptor type 1 (AT1-R) mRNA in endometrial cancer.

PubMed

Piastowska-Ciesielska, Agnieszka W; Płuciennik, Elżbieta; Wójcik-Krowiranda, Katarzyna; Bieńkiewicz, Andrzej; Nowakowska, Magdalena; Pospiech, Karolina; Bednarek, Andrzej K; Domińska, Kamila; Ochędalski, Tomasz

2013-02-01

Angiogenesis, a multistep process that results in new blood vessel formation from preexisting vasculature is essential for both the growth of solid tumour and for metastasis. Stimulation of vascular endothelial growth factor receptor (VEGFR), a transmembrane glycoprotein, results in mitogenesis. Within this family of receptors, VEGFR 2/kinase-insert-domain containing receptor appears to be principally upregulated during tumorigenesis. The aim of this study was to determine the expression of VEGFR-2/kinase-insert-domain containing receptor (KDR) and its correlation with angiotensin receptor type 1 (AT1-R) and clinical factors in endometrial carcinoma. The expression of KDR and AT1-R was studied in endometrial carcinoma and normal endometrium by Real-time RT-PCR and Western blot analysis in 136 samples. The expression profile was correlated with the clinicopathological characteristics of endometrial adenocarcinoma. We noted a significant correlation between the expression of KDR and AT1-R in tumour grade G1, G2 and G3 (R(s)=0.50; p=0.002, R(s)=0.69; p=0.0001, R(s)=0.52; p=0.005, respectively). In stage I and stage II carcinoma, a significant correlation was also found between the expression of KDR and AT1-R (R(s)=0.70, p=0.0001, R(s)=0.67; p=0.001, respectively). Moreover significant correlation was observed between both KDR and AT1-R in tissue with different myometrial invasion (R(s)=0.54, p=0.0001, R(s)=0.68; p=0.0001; respectively for tumours with invasion into the inner half and invasion into the outer half). Basing on received correlation between AT1-R and KDR expression and previous results we speculate that angiotensin through AT1-R modulates KDR expression and thus have influence on local VEGF level. However, further studies are required to clarify the biological interaction between KDR, AT1-R and other hormonal regulators in endometrial carcinoma. Copyright © 2012 Elsevier Ltd. All rights reserved.

Taxonomy of breast cancer based on normal cell phenotype predicts outcome

PubMed Central

Santagata, Sandro; Thakkar, Ankita; Ergonul, Ayse; Wang, Bin; Woo, Terri; Hu, Rong; Harrell, J. Chuck; McNamara, George; Schwede, Matthew; Culhane, Aedin C.; Kindelberger, David; Rodig, Scott; Richardson, Andrea; Schnitt, Stuart J.; Tamimi, Rulla M.; Ince, Tan A.

2014-01-01

Accurate classification is essential for understanding the pathophysiology of a disease and can inform therapeutic choices. For hematopoietic malignancies, a classification scheme based on the phenotypic similarity between tumor cells and normal cells has been successfully used to define tumor subtypes; however, use of normal cell types as a reference by which to classify solid tumors has not been widely emulated, in part due to more limited understanding of epithelial cell differentiation compared with hematopoiesis. To provide a better definition of the subtypes of epithelial cells comprising the breast epithelium, we performed a systematic analysis of a large set of breast epithelial markers in more than 15,000 normal breast cells, which identified 11 differentiation states for normal luminal cells. We then applied information from this analysis to classify human breast tumors based on normal cell types into 4 major subtypes, HR0–HR3, which were differentiated by vitamin D, androgen, and estrogen hormone receptor (HR) expression. Examination of 3,157 human breast tumors revealed that these HR subtypes were distinct from the current classification scheme, which is based on estrogen receptor, progesterone receptor, and human epidermal growth factor receptor 2. Patient outcomes were best when tumors expressed all 3 hormone receptors (subtype HR3) and worst when they expressed none of the receptors (subtype HR0). Together, these data provide an ontological classification scheme associated with patient survival differences and provides actionable insights for treating breast tumors. PMID:24463450

Transcriptional Modulation of the Immune Response by Peroxisome Proliferator-Activated Receptor-α Agonists in Autoimmune Disease1

PubMed Central

Gocke, Anne R.; Hussain, Rehana Z.; Yang, Yuhong; Peng, Haiyan; Weiner, Jeffrey; Ben, Li-Hong; Drew, Paul D.; Stuve, Olaf; Lovett-Racke, Amy E.; Racke, Michael K.

2010-01-01

Peroxisome proliferator-activated receptor-α (PPARα) agonists have been shown to have a therapeutic benefit in experimental autoimmune encephalomyelitis (EAE), an animal model for multiple sclerosis (MS). In this study, we investigated the mechanism by which the PPARα agonist gemfibrozil induces immune deviation and protects mice from EAE. We demonstrated that treatment with gemfibrozil increases expression of the Th2 transcription factor GATA-3 and decreases expression of the Th1 transcription factor T-bet in vitro and directly ex vivo. These changes correlated with an increase in nuclear PPARα expression. Moreover, the protective effects of PPARα agonists in EAE were shown to be partially dependent on IL-4 and to occur in a receptor-dependent manner. PPARα was demonstrated, for the first time, to regulate the IL-4 and IL-5 genes and to bind the IL-4 promoter in the presence of steroid receptor coactivator-1, indicating that PPARα can directly transactivate the IL-4 gene. Finally, therapeutic administration of PPARα agonists ameliorated clinically established EAE, suggesting that PPARα agonists may provide a treatment option for immune-mediated inflammatory diseases. PMID:19299749

Transcriptomics study of neurodegenerative disease: emphasis on synaptic dysfunction mechanism in Alzheimer's disease.

PubMed

Karim, Sajjad; Mirza, Zeenat; Ansari, Shakeel A; Rasool, Mahmood; Iqbal, Zafar; Sohrab, Sayed S; Kamal, Mohammad A; Abuzenadah, Adel M; Al-Qahtani, Mohammed H

2014-01-01

Alzheimer's disease (AD) is a common neurodegenerative disorder primarily affecting memory and thinking ability; caused by progressive degeneration and death of nerve cells. In this study, we integrated multiple dataset retrieved from the National Center for Biotechnology Information's Gene Expression Omnibus database, and took a systems-biology approach to compare and distinguish the molecular network based synaptic dysregulation associated with AD in particular and neurodegenerative diseases in general. We first identified 832 differentially expressed genes using cut off P value <0.5 and fold change > 2, followed by gene ontology study to identify genes associated with synapse (n=95) [membrane associated guanylate kinase, 2, amyloid beta precursor protein, neurotrophic tyrosine kinase, receptor, type 2], synapse part [γ-aminobutyric acid A receptor, γ1], synaptic vesicle [glutamate receptor, ionotropic, α-amino-3-hydroxy-5- methyl-4-isoxazole propionic acid receptor 2, synaptoporin], pre- and post-synaptic density [neuronal calcium sensor 1, glutamate receptor, metabotropic 3]. We integrated these data with known pathways using Ingenuity Pathway Analysis tool and found following synapse associated pathways to be most affected; γ-aminobutyric acid receptor signaling, synaptic long term potentiation/depression, nuclear factor-erythroid 2-related factor 2-mediated oxidative stress response, huntington's disease signaling and Reelin signaling in neurons. In conclusion, synaptic dysfunction is tightly associated with the development and progression of neurodegenerative diseases like AD.

Cellular localization of the activated EGFR determines its effect on cell growth in MDA-MB-468 cells

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

Hyatt, Dustin C.; Ceresa, Brian P.

2008-11-01

The epidermal growth factor (EGF) receptor (EGFR) is a ubiquitously expressed receptor tyrosine kinase that regulates diverse cell functions that are dependent upon cell type, the presence of downstream effectors, and receptor density. In addition to activating biochemical pathways, ligand stimulation causes the EGFR to enter the cell via clathrin-coated pits. Endocytic trafficking influences receptor signaling by controlling the duration of EGFR phosphorylation and coordinating the receptor's association with downstream effectors. To better understand the individual contributions of cell surface and cytosolic EGFRs on cell physiology, we used EGF that was conjugated to 900 nm polystyrene beads (EGF-beads). EGF-beads canmore » stimulate the EGFR and retain the activated receptor at the plasma membrane. In MDA-MB-468 cells, a breast cancer cell line that over-expresses the EGFR, only internalized, activated EGFRs stimulate caspase-3 and induce cell death. Conversely, signaling cascades triggered from activated EGFR retained at the cell surface inhibit caspase-3 and promote cell proliferation. Thus, through endocytosis, the activated EGFR can differentially regulate cell growth in MDA-MB-468 cells.« less

Pioglitazone inhibits LOX-1 expression in human coronary artery endothelial cells by reducing intracellular superoxide radical generation.

PubMed

Mehta, Jawahar L; Hu, Bo; Chen, Jiawei; Li, Dayuan

2003-12-01

LOX-1, a novel lectin-like receptor for oxidized LDL (ox-LDL), is expressed in response to ox-LDL, angiotensin II (Ang II), tumor necrosis factor (TNF)-alpha, and other stress stimuli. It is highly expressed in atherosclerotic tissues. Peroxisome proliferator-activated receptor (PPAR)-gamma ligands, such as pioglitazone, exert antiatherosclerotic effects. This study examined the regulation of LOX-1 expression in human coronary artery endothelial cells (HCAECs) by pioglitazone. Fourth generation HCAECs were treated with ox-LDL, Ang II, or TNF-alpha with or without pioglitazone pretreatment. All 3 stimuli upregulated LOX-1 expression (mRNA and protein). Pioglitazone, in a concentration-dependent manner, reduced LOX-1 expression (P<0.01 versus ox-LDL, Ang II, or TNF-alpha alone). Ox-LDL, Ang II, and TNF-alpha each enhanced intracellular superoxide radical generation, and pioglitazone pretreatment reduced superoxide generation (P<0.01 versus ox-LDL, Ang II, or TNF-alpha). Furthermore, all 3 stimuli upregulated the expression of the transcription factors nuclear factor-kappaB and activator protein-1 (determined by electrophoretic mobility shift assay), and pioglitazone pretreatment reduced this expression (P<0.01 versus ox-LDL, Ang II, or TNF-alpha). To determine the biological significance of pioglitazone-mediated downregulation of LOX-1, we studied monocyte adhesion to ox-LDL-treated HCAECs. Pioglitazone reduced the adhesion of monocytes to activated HCAECs in a fashion similar to that produced by antisense to LOX-1 mRNA. These observations suggest that the PPAR-gamma ligand pioglitazone reduces intracellular superoxide radical generation and subsequently reduces the expression of transcription factors, expression of the LOX-1 gene, and monocyte adhesion to activated endothelium. The salutary effect of PPAR-gamma ligands in atherogenesis may involve the inhibition of LOX-1 and the adhesion of monocytes to endothelium.

Significance of decoy receptor 3 (Dcr3) and external-signal regulated kinase 1/2 (Erk1/2) in gastric cancer.

PubMed

Yang, Donghai; Fan, Xin; Yin, Ping; Wen, Qiang; Yan, Feng; Yuan, Sibo; Liu, Bin; Zhuang, Guohong; Liu, Zhongchen

2012-06-06

Decoy receptor 3 (DcR3), a member of the tumor necrosis factor receptor (TNFR) superfamily, is associated with anti-tumor immunity suppression. It is highly expressed in many tumors, and its expression can be regulated by the MAPK/MEK/ERK signaling pathway. The MAPK/MEK/ERK pathway has been reported to be a regulator in tumor occurrence, development and clonal expansion. External-signal regulated kinase (ERK) is a vital member of this pathway. The expression of DcR3 and ERK1/2 in tumor tissues of gastric cancer patients was significantly higher than the non-cancerous group (P < 0.05). There was no statistical difference among tumor tissues from patients with different ages or gender, and even of different differentiation (P > 0.05). However, in patients with stage I gastric cancer, the DcR3 and ERK1/2 levels were significantly lower than patients with more advanced stages. DcR3 and ERK1/2 play a vital role in the development of gastric cancer, and they may be new markers for indicating the efficiency of gastric cancer treatment in the future.

Role of Insulin-Like Growth Factor-1 Signaling Pathway in Cisplatin-Resistant Lung Cancer Cells

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

Sun Yunguang; Zheng Siyuan; Torossian, Artour

2012-03-01

Purpose: The development of drug-resistant phenotypes has been a major obstacle to cisplatin use in non-small-cell lung cancer. We aimed to identify some of the molecular mechanisms that underlie cisplatin resistance using microarray expression analysis. Methods and Materials: H460 cells were treated with cisplatin. The differences between cisplatin-resistant lung cancer cells and parental H460 cells were studied using Western blot, MTS, and clonogenic assays, in vivo tumor implantation, and microarray analysis. The cisplatin-R cells were treated with human recombinant insulin-like growth factor (IGF) binding protein-3 and siRNA targeting IGF-1 receptor. Results: Cisplatin-R cells illustrated greater expression of the markers CD133more » and aldehyde dehydrogenase, more rapid in vivo tumor growth, more resistance to cisplatin- and etoposide-induced apoptosis, and greater survival after treatment with cisplatin or radiation than the parental H460 cells. Also, cisplatin-R demonstrated decreased expression of insulin-like growth factor binding protein-3 and increased activation of IGF-1 receptor signaling compared with parental H460 cells in the presence of IGF-1. Human recombinant IGF binding protein-3 reversed cisplatin resistance in cisplatin-R cells and targeting of IGF-1 receptor using siRNA resulted in sensitization of cisplatin-R-cells to cisplatin and radiation. Conclusions: The IGF-1 signaling pathway contributes to cisplatin-R to cisplatin and radiation. Thus, this pathway represents a potential target for improved lung cancer response to treatment.« less

Neurotrophin regulation of sodium and calcium channels in human neuroblastoma cells.

PubMed

Urbano, F J; Buño, W

2000-01-01

Neurotrophins, acting through tyrosine kinase family genes, are essential for neuronal differentiation. The expression of tyrosine kinase family genes is prognostic in neuroblastoma, and neurotrophins reduce proliferation and induce differentiation, indicating that neuroblastomas are regulated by neurotrophins. We tested the effects of nerve growth factor and brain-derived neurotrophic factor on Na(+) and Ca(2+) currents, using the whole-cell patch-clamp technique, in human neuroblastoma NB69 cells. Control cells exhibited a slow tetrodotoxin-resistant (IC(50)=98 nM) Na(+) current and a high-voltage-activated Ca(2+) current. Exposure to nerve growth factor (50 ng/ml) and/or brain-derived neurotrophic factor (5 ng/ml) produced the expression of a fast tetrodotoxin-sensitive (IC(50)=10 nM) Na(+) current after day 3, and suppressed the slow tetrodotoxin-resistant variety. The same type of high-voltage-activated Ca(2+) current was expressed in control and treated cells. The treatment increased the surface density of both Na(+) and Ca(2+) currents with time after plating, from 17 pA/pF at days 3-5 and 1-5 to 34 and 30 pA/pF after days 6-10, respectively. Therefore, both nerve growth factor and brain-derived neurotrophic factor, acting through different receptors of the tyrosine kinase family and also possibly the tumor necrosis factor receptor-II, were able to regulate differentiation and the expression of Na(+) and Ca(2+) channels, partially reproducing the modifications induced by diffusible astroglial factors. We show that neurotrophins induced differentiation to a neuronal phenotype and modified the expression of Na(+) and Ca(2+) currents, partially reproducing the effects of diffusible astroglial factors.

Identification of bone morphogenetic protein 9 (BMP9) as a novel profibrotic factor in vitro.

PubMed

Muñoz-Félix, José M; Cuesta, Cristina; Perretta-Tejedor, Nuria; Subileau, Mariela; López-Hernández, Francisco J; López-Novoa, José M; Martínez-Salgado, Carlos

2016-09-01

Upregulated synthesis of extracellular matrix (ECM) proteins by myofibroblasts is a common phenomenon in the development of fibrosis. Although the role of TGF-β in fibrosis development has been extensively studied, the involvement of other members of this superfamily of cytokines, the bone morphogenetic proteins (BMPs) in organ fibrosis has given contradictory results. BMP9 is the main ligand for activin receptor-like kinase-1 (ALK1) TGF-β1 type I receptor and its effect on fibrosis development is unknown. Our purpose was to study the effect of BMP9 in ECM protein synthesis in fibroblasts, as well as the involved receptors and signaling pathways. In cultured mice fibroblasts, BMP9 induces an increase in collagen, fibronectin and connective tissue growth factor expression, associated with Smad1/5/8, Smad2/3 and Erk1/2 activation. ALK5 inhibition with SB431542 or ALK1/2/3/6 with dorsomorphin-1, inhibition of Smad3 activation with SIS3, and inhibition of the MAPK/Erk1/2 with U0126, demonstrates the involvement of these pathways in BMP9-induced ECM synthesis in MEFs. Whereas BMP9 induced Smad1/5/8 phosphorylation through ALK1, it also induces Smad2/3 phosphorylation through ALK5 but only in the presence of ALK1. Summarizing, this is the first study that accurately identifies BMP9 as a profibrotic factor in fibroblasts that promotes ECM protein expression through ALK1 and ALK5 receptors. Copyright © 2016 Elsevier Inc. All rights reserved.

Reconstruction of nuclear receptor network reveals that NR2E3 is a novel upstream regulator of ESR1 in breast cancer

PubMed Central

Park, Yun-Yong; Kim, Kyounghyun; Kim, Sang-Bae; Hennessy, Bryan T; Kim, Soo Mi; Park, Eun Sung; Lim, Jae Yun; Li, Jane; Lu, Yiling; Gonzalez-Angulo, Ana Maria; Jeong, Woojin; Mills, Gordon B; Safe, Stephen; Lee, Ju-Seog

2012-01-01

ESR1 is one of the most important transcription factors and therapeutic targets in breast cancer. By applying systems-level re-analysis of publicly available gene expression data, we uncovered a potential regulator of ESR1. We demonstrated that orphan nuclear receptor NR2E3 regulates ESR1 via direct binding to the ESR1 promoter with concomitant recruitment of PIAS3 to the promoter in breast cancer cells, and is essential for physiological cellular activity of ESR1 in estrogen receptor (ER)-positive breast cancer cells. Moreover, expression of NR2E3 was significantly associated with recurrence-free survival and a favourable response to tamoxifen treatment in women with ER-positive breast cancer. Our results provide mechanistic insights on the regulation of ESR1 by NR2E3 and the clinical relevance of NR2E3 in breast cancer. PMID:22174013

DcR3 induces epithelial-mesenchymal transition through activation of the TGF-β3/SMAD signaling pathway in CRC.

PubMed

Liu, Yan-Ping; Zhu, Hui-Fang; Liu, Ding-Li; Hu, Zhi-Yan; Li, Sheng-Nan; Kan, He-Ping; Wang, Xiao-Yan; Li, Zu-Guo

2016-11-22

Decoy receptor 3 (DcR3), a novel member of the tumor necrosis factor receptor (TNFR) family, was recently reported to be associated with tumorigenesis and metastasis. However, the role of DcR3 in human colorectal cancer (CRC) has not been fully elucidated. In this study, we found that DcR3 expression was significantly higher in human colorectal cancer tissues than in paired normal tissues, and that DcR3 expression was strongly correlated with tumor invasion, lymph node metastases and poor prognoses. Moreover, DcR3 overexpression significantly enhanced CRC cell proliferation and migration in vitro and tumorigenesis in vivo. Conversely, DcR3 knockdown significantly repressed CRC cell proliferation and migration in vitro, and DcR3 deficiency also attenuated CRC tumorigenesis and metastasis in vivo. Functionally, DcR3 was essential for TGF-β3/SMAD-mediated epithelial-mesenchymal transition (EMT) of CRC cells. Importantly, cooperation between DcR3 and TGF-β3/SMAD-EMT signaling-related protein expression was correlated with survival and survival time in CRC patients. In conclusion, our results demonstrate that DcR3 may be a prognostic biomarker for CRC and that this receptor facilitates CRC development and metastasis by participating in TGF-β3/SMAD-mediated EMT of CRC cells.

Bromocriptine modulates the expression of PTHrP receptor, Indian hedgehog, and Runx2 proteins in the growth plate of lactating rats.

PubMed

Wongdee, Kannikar; Thonapan, Natchayaporn; Saengamnart, Wasana; Krishnamra, Nateetip; Charoenphandhu, Narattaphol

2013-09-01

In lactating rats, the endochondral bone growth is markedly enhanced, leading to the lengthening of long bone. This lactation-induced bone elongation could be abolished by a dopaminergic D2 receptor agonist bromocriptine, but how bromocriptine altered the expression of major chondroregulatory proteins in the growth plate cartilage was elusive. Here, we performed a quantitative immunohistochemical analysis to determine the expression of various peptides and transcription factors known to control the growth plate chondrocyte proliferation and differentiation [i.e., parathyroid hormone-related protein (PTHrP), PTHrP receptor, Indian hedgehog (Ihh), and runt-related transcription factor 2 (Runx2)], in bromocriptine-treated lactating rats. The results showed that bromocriptine markedly increased Ihh expression in hypertrophic chondrocytes during early and mid-lactation, while the expression of PTHrP receptor, but not its ligand PTHrP, was upregulated in the proliferative and hypertrophic zones during mid and late lactation. In contrast, the expression of Runx2, an important transcription factor for chondrocyte differentiation, was suppressed in the hypertrophic chondrocytes of bromocriptine-treated rats. In conclusion, bromocriptine increased Ihh and PTHrP receptor expressions and decreased Runx2 expression, which might, in turn, enhance chondrocyte proliferation and delay chondrocyte hypertrophy, thereby slowing down endochondral bone growth. This finding could explain how bromocriptine compromised the lactation-induced bone elongation.

The Metastasis Suppressor, N-MYC Downstream-regulated Gene-1 (NDRG1), Down-regulates the ErbB Family of Receptors to Inhibit Downstream Oncogenic Signaling Pathways*

PubMed Central

Kovacevic, Zaklina; Menezes, Sharleen V.; Sahni, Sumit; Kalinowski, Danuta S.; Bae, Dong-Hun; Lane, Darius J. R.; Richardson, Des R.

2016-01-01

N-MYC downstream-regulated gene-1 (NDRG1) is a potent growth and metastasis suppressor that acts through its inhibitory effects on a wide variety of cellular signaling pathways, including the TGF-β pathway, protein kinase B (AKT)/PI3K pathway, RAS, etc. To investigate the hypothesis that its multiple effects could be regulated by a common upstream effector, the role of NDRG1 on the epidermal growth factor receptor (EGFR) and other members of the ErbB family, namely human epidermal growth factor receptor 2 (HER2) and human epidermal growth factor receptor 3 (HER3), was examined. We demonstrate that NDRG1 markedly decreased the expression and activation of EGFR, HER2, and HER3 in response to the epidermal growth factor (EGF) ligand, while also inhibiting formation of the EGFR/HER2 and HER2/HER3 heterodimers. In addition, NDRG1 also decreased activation of the downstream MAPKK in response to EGF. Moreover, novel anti-tumor agents of the di-2-pyridylketone class of thiosemicarbazones, namely di-2-pyridylketone 4,4-dimethyl-3-thiosemicarbazone and di-2-pyridylketone 4-cyclohexyl-4-methyl-3-thiosemicarbazone, which markedly up-regulate NDRG1, were found to inhibit EGFR, HER2, and HER3 expression and phosphorylation in cancer cells. However, the mechanism involved appeared dependent on NDRG1 for di-2-pyridylketone 4,4-dimethyl-3-thiosemicarbazone, but was independent of this metastasis suppressor for di-2-pyridylketone 4-cyclohexyl-4-methyl-3-thiosemicarbazone. This observation demonstrates that small structural changes in thiosemicarbazones result in marked alterations in molecular targeting. Collectively, these results reveal a mechanism for the extensive downstream effects on cellular signaling attributed to NDRG1. Furthermore, this study identifies a novel approach for the treatment of tumors resistant to traditional EGFR inhibitors. PMID:26534963

Distinct actions of ancestral vinclozolin and juvenile stress on neural gene expression in the male rat.

PubMed

Gillette, Ross; Miller-Crews, Isaac; Skinner, Michael K; Crews, David

2015-01-01

Exposure to the endocrine disrupting chemical vinclozolin during gestation of an F0 generation and/or chronic restraint stress during adolescence of the F3 descendants affects behavior, physiology, and gene expression in the brain. Genes related to the networks of growth factors, signaling peptides, and receptors, steroid hormone receptors and enzymes, and epigenetic related factors were measured using quantitative polymerase chain reaction via Taqman low density arrays targeting 48 genes in the central amygdaloid nucleus, medial amygdaloid nucleus, medial preoptic area (mPOA), lateral hypothalamus (LH), and the ventromedial nucleus of the hypothalamus. We found that growth factors are particularly vulnerable to ancestral exposure in the central and medial amygdala; restraint stress during adolescence affected neural growth factors in the medial amygdala. Signaling peptides were affected by both ancestral exposure and stress during adolescence primarily in hypothalamic nuclei. Steroid hormone receptors and enzymes were strongly affected by restraint stress in the mPOA. Epigenetic related genes were affected by stress in the ventromedial nucleus and by both ancestral exposure and stress during adolescence independently in the central amygdala. It is noteworthy that the LH showed no effects of either manipulation. Gene expression is discussed in the context of behavioral and physiological measures previously published.

PANC-1 pancreatic cancer cell growth inhibited by cucurmosin alone and in combination with an epidermal growth factor receptor-targeted drug.

PubMed

Wang, Congfei; Yang, Aiqin; Zhang, Baoming; Yin, Qiang; Huang, Heguang; Chen, Minghuang; Xie, Jieming

2014-03-01

To investigate the inhibition of PANC-1 pancreatic cancer cell growth by cucurmosin (CUS) and its possible mechanism. We observed the inhibition of PANC-1 cell growth by sulforhodamine B and colony-forming experiments in vitro and established nonobese diabetic/severe combined immunodeficiency mouse subcutaneous tumor models in vivo. We used Western blot to analyze protein levels related to apoptosis and epidermal growth factor receptor (EGFR) signaling pathways after drug intervention, whereas the messenger RNA expression of EGFR was analyzed by quantitative real-time polymerase chain reaction. Sulforhodamine B and colony-forming experiments indicated that CUS inhibited PANC-1 cell proliferation in a dose- and time-dependent manner. A stronger inhibitory effect was observed when CUS was combined with gefitinib. The subcutaneous tumor growth was also inhibited. Western blot showed that all the examined proteins decreased, except for 4E-BP1 and the active fragments of caspase 3 and caspase 9 increased. Epidermal growth factor receptor expression did not change significantly in quantitative real-time polymerase chain reaction. Cucurmosin can strongly inhibit the growth of PANC-1 cells in vitro and in vivo. Cucurmosin can down-regulate EGFR protein expression, but not at the messenger RNA level. Cucurmosin can also inhibit the ras/raf and phosphatidylinositol 3-kinase/Akt downstream signaling pathways and enhance the sensitivity of the EGFR-targeted drug gefitinib.

Dual Role of Cyanidin-3-glucoside on the Differentiation of Bone Cells.

PubMed

Park, K H; Gu, D R; So, H S; Kim, K J; Lee, S H

2015-12-01

Cyanidin-3-glucoside (C3G) is one of the major components of anthocyanin, a water-soluble phytochemical. Recent studies demonstrated the chemopreventive and chemotherapeutic activities of C3G in various conditions, including cancer, although the precise effects of C3G on osteoclast and osteoblast differentiation remain unclear. Here, we investigated the role of C3G in the differentiation of bone-associated cells and its underlying mechanism. C3G inhibited receptor activator of nuclear factor kappa-B ligand (RANKL)-mediated osteoclast differentiation and formation in a dose-dependent manner and downregulated the expression of osteoclast differentiation marker genes. Pretreatment with C3G considerably reduced the induction of extracellular signal-regulated kinase, c-Jun N-terminal kinase, and p38 mitogen-activated kinases activation by RANKL in osteoclast precursor cells. Furthermore, C3G dramatically inhibited the expression of c-Fos and nuclear factor of activated T-cells, cytoplasmic 1, which are important transcription factors for osteoclast differentiation and activation. The formation of osteoclasts in coculture of bone marrow cells and calvaria-derived osteoblasts was also inhibited by C3G treatment, although the expression of macrophage colony-stimulating factor and RANKL (master factors for osteoclast differentiation and formation) and osteoprotegerin (a decoy receptor for RANKL) on osteoblasts was unaffected. The inhibitory effect of C3G on osteoclastogenesis is therefore targeted specifically to osteoclasts but not osteoblasts. Moreover, analysis of the expression levels of osteoblast differentiation marker genes and alizarin red staining showed that osteoblast differentiation and matrix formation increased after C3G treatment. Taken together, these results strongly suggest that C3G has a dual role in bone metabolism, as an effective inhibitor of osteoclast differentiation but also as an activator of osteoblast differentiation. Therefore, C3G may be used as a potent preventive or therapeutic agent for bone-related diseases, such as osteoporosis, rheumatoid arthritis, and periodontitis. © International & American Associations for Dental Research 2015.

Novel targeted approaches to treating biliary tract cancer: the dual epidermal growth factor receptor and ErbB-2 tyrosine kinase inhibitor NVP-AEE788 is more efficient than the epidermal growth factor receptor inhibitors gefitinib and erlotinib.

PubMed

Wiedmann, Marcus; Feisthammel, Jürgen; Blüthner, Thilo; Tannapfel, Andrea; Kamenz, Thomas; Kluge, Annett; Mössner, Joachim; Caca, Karel

2006-08-01

Aberrant activation of the epidermal growth factor receptor is frequently observed in neoplasia, notably in tumors of epithelial origin. Attempts to treat such tumors with epidermal growth factor receptor antagonists resulted in remarkable success in recent studies. Little is known, however, about the efficacy of this therapy in biliary tract cancer. Protein expression of epidermal growth factor receptor, ErbB-2, and vascular endothelial growth factor receptor-2 was assessed in seven human biliary tract cancer cell lines by immunoblotting. In addition, histological sections from 19 patients with extrahepatic cholangiocarcinoma were analyzed for epidermal growth factor receptor, ErbB-2 and vascular endothelial growth factor receptor-2 expression by immunohistochemistry. Moreover, we sequenced the cDNA products representing the entire epidermal growth factor receptor coding region of the seven cell lines, and searched for genomic epidermal growth factor receptor amplifications and polysomy by fluorescence in-situ hybridization. Cell growth inhibition by gefitinib erlotinib and NVP-AEE788 was studied in vitro by automated cell counting. In addition, the anti-tumoral effect of erlotinib and NVP-AEE788 was studied in a chimeric mouse model. The anti-tumoral drug mechanism in this model was assessed by MIB-1 antibody staining, terminal deoxynucleotidyl transfer-mediated dUTP nick end-labelling assay, von Willebrand factor staining, and immunoblotting for p-p42/44 (p-Erk1/2, p-MAPK) and p-AKT. Immunoblotting revealed expression of epidermal growth factor receptor, ErbB-2, and vascular endothelial growth factor receptor-2 in all biliary tract cancer cell lines. EGFR was detectable in six of 19 (32%) extrahepatic human cholangiocarcinoma tissue samples, ErbB-2 in 16 of 19 (84%), and vascular endothelial growth factor receptor-2 in nine of 19 (47%). Neither epidermal growth factor receptor mutations nor amplifications or polysomy were found in the seven biliary tract cancer cell lines. Gefitinib, erlotinib and NVP-AEE788 caused a significant growth inhibition in vitro; however, there was a significant difference in efficacy (NVP-AEE788>erlotinib>gefitinib). After 14 days of in-vivo treatment, using the chimeric mouse model, tumors had a significantly reduced volume and mass after NVP-AEE788, but not after erlotinib treatment, as compared with placebo. Reduction of proliferation (signalling via the mitogen-activated protein kinase pathway), induction of apoptosis and inhibition of angiogenesis were the main mechanisms of drug action. No significant reduction of anti-apoptotic AKT phosphorylation, however, occurred, which may be a possible counter mechanism of the tumor. Epidermal growth factor receptor, ErbB-2, and vascular endothelial growth factor receptor-2 expression was detectable in biliary tract cancer, and receptor inhibition exerts marked effects on tumor growth in vitro and in vivo, which was strongest for the dual EGFR/ErbB-2 inhibitor NVP-AEE788. Therefore, further clinical evaluation of this new drug for the treatment of biliary tract cancer is recommended.

The Synthetic Cannabinoid R(+)WIN55,212-2 Augments Interferon-β Expression via Peroxisome Proliferator-activated Receptor-α*

PubMed Central

Downer, Eric J.; Clifford, Eileen; Amu, Sylvie; Fallon, Padraic G.; Moynagh, Paul N.

2012-01-01

We have demonstrated that R(+)WIN55,212-2, a synthetic cannabinoid that possesses cannabimimetic properties, acts as a novel regulator of Toll-like receptor 3 (TLR3) signaling to interferon (IFN) regulatory factor 3 (IRF3) activation and IFN-β expression, and this is critical for manifesting its protective effects in a murine multiple sclerosis model. Here we investigated the role of peroxisome proliferator-activated receptor-α (PPARα) in mediating the effects of R(+)WIN55,212-2 on this pathway. Data herein demonstrate that the TLR3 agonist poly(I:C) promotes IFN-β expression and R(+)WIN55,212-2 enhances TLR3-induced IFN-β expression in a stereoselective manner via PPARα. R(+)WIN55,212-2 promotes increased transactivation and expression of PPARα. Using the PPARα antagonist GW6471, we demonstrate that R(+)WIN55,212-2 acts via PPARα to activate JNK, activator protein-1, and positive regulatory domain IV to transcriptionally regulate the IFN-β promoter. Furthermore, GW6471 ameliorated the protective effects of R(+)WIN55,212-2 during the initial phase of experimental autoimmune encephalomyelitis. Overall, these findings define PPARα as an important mediator in manifesting the effects of R(+)WIN55,212-2 on the signaling cascade regulating IFN-β expression. The study adds to our molecular appreciation of potential therapeutic effects of R(+)WIN55,212-2 in multiple sclerosis. PMID:22654113

Immunohistochemical study of the growth factors, aFGF, bFGF, PDGF-AB, VEGF-A and its receptor (Flk-1) during arteriogenesis.

PubMed

Wu, Song; Wu, Xiaoqiong; Zhu, Wu; Cai, Wei-Jun; Schaper, Jutta; Schaper, Wolfgang

2010-10-01

Growth factors are viewed as main arteriogenic stimulators for collateral vessel growth. However, the information about their native expression and distribution in collateral vessels is still limited. This study was designed to profile expression of acidic and basic FGF, platelet-derived growth factor (PDGF-AB) and vascular endothelial growth factor (VEGF-A) and its receptor, fetal liver kinase-1 (Flk-1) during arteriogenesis by confocal immunofluorescence in both dog ameroid constrictor model and rabbit arteriovenous shunt model of arteriogenesis. We found that: (1) in normal arteries (NA) in dog heart, aFGF, bFGF, and PDGF-AB all were mainly expressed in endothelial cells (EC) and media smooth muscle cells (SMC), but the expression of aFGF was very weak, with those of the other two being moderate; (2) in collateral arteries (CAs), aFGF, bFGF, and PDGF-AB all were significantly upregulated (P < 0.05); they were present in all the layers of the vascular wall and were 2.1, 1.7, and 1.9 times higher than that in NA, respectively; and (3) in NA in rabbit hind limb, VEGF-A was absent, Flk-1 was only weakly present in endothelial cells, but in one week CAs VEGF-A and Flk-1 were significantly increased in both shunt and ligation sides; this was more evident in the shunt-side CAs, 2.3, and 2 times higher than that in the ligation side, respectively. In conclusion, our data demonstrate for the first time that growth factors, aFGF, bFGF, and PDGF-AB are significantly upregulated in collateral vessels in dog heart, and enhanced VEGF-A and its receptor, Flk-1, are associated with rapid and lasting increased shear stress. These findings suggest that endogenous production of growth factors could be an important factor promoting collateral vessel growth.

Esculetin attenuates receptor activator of nuclear factor kappa-B ligand-mediated osteoclast differentiation through c-Fos/nuclear factor of activated T-cells c1 signaling pathway

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

Baek, Jong Min; Park, Sun-Hyang; Cheon, Yoon-Hee

Esculetin exerts various biological effects on anti-oxidation, anti-tumors, and anti-inflammation. However, the involvement of esculetin in the bone metabolism process, particularly osteoclast differentiation has not yet been investigated. In the present study, we first confirmed the inhibitory effect of esculetin on receptor activator of nuclear factor-κB ligand (RANKL)-induced osteoclast formation. We then revealed the relationship between esculetin and the expression of osteoclast-specific molecules to elucidate its underlying mechanisms. Esculetin interfered with the expression of c-Fos and nuclear factor of activated T cell c1 (NFATc1) both at the mRNA and protein level with no involvement in osteoclast-associated early signaling pathways, suppressingmore » the expression of various transcription factors exclusively expressed in osteoclasts such as tartrate-resistant acid phosphatase (Trap), osteoclast-associated receptor (Oscar), dendritic cell-specific transmembrane protein (Dcstamp), osteoclast stimulatory transmembrane protein (Ocstamp), cathepsin K, αvβ3 integrin, and calcitonin receptor (Ctr). Additionally, esculetin inhibited the formation of filamentous actin (F-actin) ring-positive osteoclasts during osteoclast differentiation. However, the development of F-actin structures and subsequent bone resorbing activity of mature osteoclasts, which are observed in osteoclast/osteoblast co-culture systems were not affected by esculetin. Taken together, our results indicate for the first time that esculetin inhibits RANKL-mediated osteoclastogenesis via direct suppression of c-Fos and NFATc1 expression and exerts an inhibitory effect on actin ring formation during osteoclastogenesis. - Highlights: • We first investigated the effects of esculetin on osteoclast differentiation and function. • Our data demonstrate for the first time that esculetin can suppress osteoclastogenesis in vitro. • Esculetin acts as an inhibitor of c-Fos and NFATc1 activation. • Esculetin acts a negative regulator of actin ring formation during osteoclast differentiation. • Esculetin deserves new evaluation as a potential treatment target in various bone diseases.« less

Signal Peptide and Denaturing Temperature are Critical Factors for Efficient Mammalian Expression and Immunoblotting of Cannabinoid Receptors*

PubMed Central

WANG, Chenyun; WANG, Yingying; WANG, Miao; CHEN, Jiankui; YU, Nong; SONG, Shiping; KAMINSKI, Norbert E.; ZHANG, Wei

2013-01-01

Summary Many researchers employed mammalian expression system to artificially express cannabinoid receptors, but immunoblot data that directly prove efficient protein expression can hardly be seen in related research reports. In present study, we demonstrated cannabinoid receptor protein was not able to be properly expressed with routine mammalian expression system. This inefficient expression was rescued by endowing an exogenous signal peptide ahead of cannabinoid receptor peptide. In addition, the artificially synthesized cannabinoid receptor was found to aggregate under routine sample denaturing temperatures (i.e., ≥95°C), forming a large molecular weight band when analyzed by immunoblotting. Only denaturing temperatures ≤75°C yielded a clear band at the predicted molecular weight. Collectively, we showed that efficient mammalian expression of cannabinoid receptors need a signal peptide sequence, and described the requirement for a low sample denaturing temperature in immunoblot analysis. These findings provide very useful information for efficient mammalian expression and immunoblotting of membrane receptors. PMID:22528237

Central insulin-mediated regulation of hepatic glucose production [Review].

PubMed

Inoue, Hiroshi

2016-01-01

Insulin controls hepatic glucose production (HGP) and maintains glucose homeostasis through the direct action of hepatic insulin receptors, as well as the indirect action of insulin receptors in the central nervous system. Insulin acts on insulin receptors in the hypothalamic arcuate nucleus, activates ATP-sensitive potassium channels in a phosphoinositide 3-kinase (PI3K)-dependent manner, induces hyperpolarization of the hypothalamic neurons, and regulates HGP via the vagus nerve. In the liver, central insulin action augments IL-6 expression in Kupffer cells and activates STAT3 transcription factors in hepatocytes. Activated STAT3 suppresses the gene expression of gluconeogenic enzymes, thereby reducing HGP. It has become evident that nutrients such as glucose, fatty acids, and amino acids act upon the hypothalamus together with insulin, affecting HGP. On the other hand, HGP control by central insulin action is impeded in obesity and impeded by insulin resistance due to disturbance of PI3K signaling and inflammation in the hypothalamus or inhibition of STAT3 signaling in the liver. Although the mechanism of control of hepatic gluconeogenic gene expression by central insulin action is conserved across species, its importance in human glucose metabolism has not been made entirely clear and its elucidation is anticipated in the future.

The role of protease-activated receptors PAR-1 and PAR-2 in the repair of 16HBE 14o(-) epithelial cell monolayers in vitro.

PubMed

Ewen, D; Clarke, S L; Smith, J R; Berger, C; Salmon, G; Trevethick, M; Shute, J K

2010-03-01

We recently reported that repair following mechanical wounding of epithelial cell layers in vitro is dependent on fibrin formation and the activity of locally expressed coagulation cascade proteins. Serine proteases of the coagulation cascade are an important group of protease-activated receptor (PAR) activators and PAR-1 to 4 are expressed by the normal bronchial epithelium. We tested the hypothesis that activation of PAR-1 and PAR-2 by coagulation cascade proteases stimulates epithelial repair via effects on fibrin formation. Using mechanically wounded 16HBE 14o(-) epithelial cell layers in culture, we investigated the effect of PAR-1 and PAR-2 agonist peptides, control partially scrambled peptides and PAR-neutralizing antibodies on the rate of repair and fibrin formation. Coagulation factors in culture supernatants were measured by immunoblot. RT-PCR was used to investigate PAR-1, PAR-2 and PGE2 receptor (EP-1 to EP-4) expression in this model and qRT-PCR to quantify responses to wounding. Additionally, we investigated the effect of exogenously added factor Xa (FXa) and neutrophil elastase and the influence of PGE2 and indomethacin on the repair response. PAR-1 and PAR-2 peptide agonists stimulated the rate of repair and enhanced the formation of a fibrin provisional matrix to support the repair process. Conversely, PAR-neutralizing antibodies inhibited repair. Under serum-free culture conditions, 16HBE 14o(-) cells expressed EP-2 and EP-3, but not EP-1 or EP-4, receptors. Wounding induced an increased expression of EP-3 but did not alter EP-2, PAR-1 or PAR-2 expression. In the absence of PAR agonists, there was no evidence for a role for PGE2 in fibrin formation or the repair process. Indomethacin attenuated fibrin formation in wounded cultures only in the presence of the PAR-2 peptide. FXa stimulated epithelial repair while neutrophil elastase reduced the levels of coagulation factors and inhibited repair. Locally expressed serine proteases of the coagulation cascade activate PAR-1 and PAR-2 to enhance fibrin formation and bronchial epithelial repair.

Immunocytochemical assessment of sigma-1 receptor and human sterol isomerase in breast cancer and their relationship with a series of prognostic factors

PubMed Central

Simony-Lafontaine, J; Esslimani, M; Bribes, E; Gourgou, S; Lequeux, N; Lavail, R; Grenier, J; Kramar, A; Casellas, P

2000-01-01

The purpose of this study was to immunocytochemically investigate two new markers, the sigma-1 receptor and the human sterol isomerase (hSI), in comparison with a series of clinicopathological and immunocytochemical prognostic factors in a trial including 95 patients with operable primary breast cancers. Our results showed no statistically significant relationship between these two markers and the age of the patients, their menopausal status, the tumour size and its histological grade, the nodal status and the expression of the Ki-67 proliferative marker. However, we evidenced a close correlation between the sigma-1 receptor expression and the hormonal receptor positivity (P = 0.008), essentially due to a link with the progesterone receptor status (P = 0.01). By contrast there was an inverse relationship between hSI expression and the oestrogen receptor and/or progesterone receptor positivity (P = 0.098). A significant relationship was shown between both the sigma-1 receptor, hSI expressions and Bcl2 expression, with P = 0.017 and 0.035 respectively. We also assessed whether the expression of the sigma-1 receptor or hSI might be linked with disease-free survival (DFS) and found that the presence of hSI and the absence of sigma-1 receptor expression were associated with a poorer disease-free survival (P = 0.007). Altogether these results suggest that in primary breast carcinomas in association with the evaluation of the steroid receptor status, the sigma-1 receptor and hSI may be interesting new markers useful to identify those patients who might be able to benefit from an adjuvant therapy. © 2000 Cancer Research Campaign PMID:10864204

Selective Glucocorticoid Receptor modulators.

PubMed

De Bosscher, Karolien

2010-05-31

The ancient two-faced Roman god Janus is often used as a metaphor to describe the characteristics of the Glucocorticoid Receptor (NR3C1), which exhibits both a beneficial side, that serves to halt inflammation, and a detrimental side responsible for undesirable effects. However, recent developments suggest that the Glucocorticoid Receptor has many more faces with the potential to express a range of different functionalities, depending on factors that include the tissue type, ligand type, receptor variants, cofactor surroundings and target gene promoters. This behavior of the receptor has made the development of safer ligands, that trigger the expression program of only a desirable subset of genes, a real challenge. Thus more knowledge-based fundamental research is needed to ensure the design and development of selective Glucocorticoid Receptor modulators capable of reaching the clinic. Recent advances in the characterization of novel selective Glucocorticoid Receptor modulators, specifically in the context of anti-inflammatory strategies, will be described in this review. 2010 Elsevier Ltd. All rights reserved.

Sildenafil prevents the up-regulation of transient receptor potential canonical channels in the development of cardiomyocyte hypertrophy

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

Kiso, Hironori; Ohba, Takayoshi; Iino, Kenji

2013-07-05

Highlights: •Transient receptor potential canonical (TRPC1, 3 and 6) are up-regulated by ET-1. •Sildenafil inhibited hypertrophic responses (BNP, Ca entry, NFAT activation). •Sildenafil suppressed TRPC1, 3 and 6 expression. -- Abstract: Background: Transient receptor potential canonical (TRPCs) channels are up-regulated in the development of cardiac hypertrophy. Sildenafil inhibits TRPC6 activation and expression, leading to the prevention of cardiac hypertrophy. However, the effects of sildenafil on the expression of other TRPCs remain unknown. We hypothesized that in addition to its effects of TRPC6, sildenafil blocks the up-regulation of other TRPC channels to suppress cardiomyocyte hypertrophy. Methods and results: In cultured neonatalmore » rat cardiomyocytes, a 48 h treatment with 10 nM endothelin (ET)-1 induced hypertrophic responses characterized by nuclear factor of activated T cells activation and enhancement of brain natriuretic peptide expression and cell surface area. Co-treatment with sildenafil (1 μM, 48 h) inhibited these ET-1-induced hypertrophic responses. Although ET-1 enhanced the gene expression of TRPCs, sildenafil inhibited the enhanced gene expression of TRPC1, C3 and C6. Moreover, co-treatment with sildenafil abolished the augmentation of SOCE in the hypertrophied cardiomyocytes. Conclusions: These results suggest that sildenafil inhibits cardiomyocyte hypertrophy by suppressing the up-regulation of TRPC expression.« less

Inositol 1,4,5-trisphosphate receptor regulates replication, differentiation, infectivity and virulence of the parasitic protist Trypanosoma cruzi.

PubMed

Hashimoto, Muneaki; Enomoto, Masahiro; Morales, Jorge; Kurebayashi, Nagomi; Sakurai, Takashi; Hashimoto, Tetsuo; Nara, Takeshi; Mikoshiba, Katsuhiko

2013-03-01

In animals, inositol 1,4,5-trisphosphate receptors (IP3 Rs) are ion channels that play a pivotal role in many biological processes by mediating Ca(2+) release from the endoplasmic reticulum. Here, we report the identification and characterization of a novel IP3 R in the parasitic protist, Trypanosoma cruzi, the pathogen responsible for Chagas disease. DT40 cells lacking endogenous IP3 R genes expressing T. cruzi IP3 R (TcIP3 R) exhibited IP3 -mediated Ca(2+) release from the ER, and demonstrated receptor binding to IP3 . TcIP3 R was expressed throughout the parasite life cycle but the expression level was much lower in bloodstream trypomastigotes than in intracellular amastigotes or epimastigotes. Disruption of two of the three TcIP3 R gene loci led to the death of the parasite, suggesting that IP3 R is essential for T. cruzi. Parasites expressing reduced or increased levels of TcIP3 R displayed defects in growth, transformation and infectivity, indicating that TcIP3 R is an important regulator of the parasite's life cycle. Furthermore, mice infected with T. cruzi expressing reduced levels of TcIP3 R exhibited a reduction of disease symptoms, indicating that TcIP3 R is an important virulence factor. Combined with the fact that the primary structure of TcIP3 R has low similarity to that of mammalian IP3 Rs, TcIP3 R is a promising drug target for Chagas disease. © 2013 Blackwell Publishing Ltd.

Molecular characterization of human thyroid hormone receptor β isoform 4.

PubMed

Moriyama, Kenji; Yamamoto, Hiroyuki; Futawaka, Kumi; Atake, Asami; Kasahara, Masato; Tagami, Tetsuya

2016-01-01

Thyroid hormone exerts a pleiotropic effect on development, differentiation, and metabolism through thyroid hormone receptor (TR). A novel thyroid hormone receptor β isoform (TRβ4) was cloned using PCR from a human pituitary cDNA library as a template. We report here the characterization of TRβ4 from a molecular basis. Temporal expression of TRβ4 during the fetal period is abundant in the brain and kidney, comparable with the adult pattern. Western blot analysis revealed that TRs are ubiquitination labile proteins, while TRβ1 is potentially stable. TRβ1, peroxisome proliferator-activated receptors (PPAR), and vitamin D receptor (VDR), which belong to class II transcription factors that function via the formation of heterodimeric complexes with retinoid X receptor (RXR), were suppressed by TRβ4 in a dose-dependent manner. Thus, TRβ4 exhibits ligand-independent transcriptional silencing, possibly as a substitute for dimerized RXR. In this study, TRβ1 and TRβ4 transcripts were detected in several cell lines. Quantitative RT-PCR assay showed that the expression of TRβ4 in human embryonic carcinoma cells of the testis was suppressed by sex hormone in a reciprocal manner to TRβ1. In contrast, TRβ4 was expressed under a high dose of triiodothyronine (T3) in a reciprocal manner to TRβ1. Finally, in transiently transfected NIH-3T3 cells, green fluorescence protein (GFP)-tagged TRβ4 was mostly nuclear in both the absence and the presence of T3. By mutating defined regions of both TRβs, we found that both TRβ1 and TRβ4 had altered nuclear/cytoplasmic distribution as compared with wild-type, and different to T3 and the nuclear receptor corepressor (NCoR). Thus, site-specific DNA binding is not essential for maintaining TRβs within the nucleus.

NR4A nuclear receptors support memory enhancement by histone deacetylase inhibitors

PubMed Central

Hawk, Joshua D.; Bookout, Angie L.; Poplawski, Shane G.; Bridi, Morgan; Rao, Allison J.; Sulewski, Michael E.; Kroener, Brian T.; Manglesdorf, David J.; Abel, Ted

2012-01-01

The formation of a long-lasting memory requires a transcription-dependent consolidation period that converts a short-term memory into a long-term memory. Nuclear receptors compose a class of transcription factors that regulate diverse biological processes, and several nuclear receptors have been implicated in memory formation. Here, we examined the potential contribution of nuclear receptors to memory consolidation by measuring the expression of all 49 murine nuclear receptors after learning. We identified 13 nuclear receptors with increased expression after learning, including all 3 members of the Nr4a subfamily. These CREB-regulated Nr4a genes encode ligand-independent “orphan” nuclear receptors. We found that blocking NR4A activity in memory-supporting brain regions impaired long-term memory but did not impact short-term memory in mice. Further, expression of Nr4a genes increased following the memory-enhancing effects of histone deacetylase (HDAC) inhibitors. Blocking NR4A signaling interfered with the ability of HDAC inhibitors to enhance memory. These results demonstrate that the Nr4a gene family contributes to memory formation and is a promising target for improving cognitive function. PMID:22996661

Expression of a transmembrane phosphotyrosine phosphatase inhibits cellular response to platelet-derived growth factor and insulin-like growth factor-1.

PubMed

Mooney, R A; Freund, G G; Way, B A; Bordwell, K L

1992-11-25

Tyrosine phosphorylation is a mechanism of signal transduction shared by many growth factor receptors and oncogene products. Phosphotyrosine phosphatases (PTPases) potentially modulate or counter-regulate these signaling pathways. To test this hypothesis, the transmembrane PTPase CD45 (leukocyte common antigen) was expressed in the murine cell line C127. Hormone-dependent autophosphorylation of the platelet-derived growth factor (PDGF) and insulin-like growth factor-1 (IGF-1) receptors was markedly reduced in cells expressing the transmembrane PTPase. Tyrosine phosphorylation of other PDGF-dependent phosphoproteins (160, 140, and 55 kDa) and IGF-1-dependent phosphoproteins (145 kDa) was similarly decreased. Interestingly, the pattern of growth factor-independent tyrosine phosphorylations was comparable in cells expressing the PTPase and control cells. This suggests a selectivity or accessibility of the PTPase limited to a subset of cellular phosphotyrosyl proteins. The maximum mitogenic response to PDGF and IGF-1 in cells expressing the PTPase was decreased by 67 and 71%, respectively. These results demonstrate that a transmembrane PTPase can both affect the tyrosine phosphorylation state of growth factor receptors and modulate proximal and distal cellular responses to the growth factors.

Effects of heparin-binding epidermal growth factor-like growth factor on cell repopulation and signal transduction in periodontal ligament cells after scratch wounding in vitro.

PubMed

Lee, J S; Kim, J M; Hong, E K; Kim, S-O; Yoo, Y-J; Cha, J-H

2009-02-01

A growing amount of attention has been placed on periodontal regeneration and wound healing for periodontal therapy. This study was conducted in an effort to determine the effects of heparin-binding epidermal growth factor-like growth factor on cell repopulation and signal transduction in periodontal ligament cells after scratch wounding in vitro. Human periodontal ligament cells were acquired from explant tissue of human healthy periodontal ligament. After the wounding of periodontal ligament cells, the change in expression of heparin-binding epidermal growth factor-like growth factor and epidermal growth factor receptors 1-4 mRNA was assessed. The effects of heparin-binding epidermal growth factor-like growth factor on periodontal ligament cell proliferation and repopulation were assessed in vitro via the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and by photographing the injuries, respectively. Extracellular signal-regulated kinase (Erk)1/2, p38 and Akt phosphorylation was characterized via western blotting. Scratch wounding resulted in a significant up-regulation of heparin-binding epidermal growth factor-like growth factor mRNA expression, whereas wounding had no effect on the expression levels of epidermal growth factor receptors 1-4. Interestingly, no expression of epidermal growth factor receptors 2 and 4 was detectable prior to or after wounding. Heparin-binding epidermal growth factor-like growth factor treatment promoted the proliferation and repopulation of periodontal ligament cells. The scratch wounding also stimulated the phosphorylation of Erk1/2 and p38, but not of Akt, in periodontal ligament cells, and heparin-binding epidermal growth factor-like growth factor treatment applied after wounding amplified and extended the activations of Erk1/2 and p38, but not of Akt. Furthermore, Erk1/2 inhibition blocked the process of cell repopulation induced by heparin-binding epidermal growth factor-like growth factor, whereas the inhibition of p38 delayed the process. These results indicate that heparin-binding epidermal growth factor-like growth factor may constitute a critical factor in the wound healing of human periodontal ligament cells by a mechanism that requires the activation of Erk1/2 via specific interaction with epidermal growth factor receptor 1.

Modeling the Effects of HER/ErbB1-3 Coexpression on Receptor Dimerization and Biological Response

PubMed Central

Shankaran, Harish; Wiley, H. Steven; Resat, Haluk

2006-01-01

The human epidermal growth factor receptor (HER/ErbB) system comprises the epidermal growth factor receptor (EGFR/HER1) and three other homologs, namely HERs 2–4. This receptor system plays a critical role in cell proliferation and differentiation and receptor overexpression has been associated with poor prognosis in cancers of the epithelium. Here, we examine the effect of coexpressing varying levels of HERs 1–3 on the receptor dimerization patterns using a detailed kinetic model for HER/ErbB dimerization and trafficking. Our results indicate that coexpression of EGFR with HER2 or HER3 biases signaling to the cell surface and retards signal downregulation. In addition, simultaneous coexpression of HERs 1–3 leads to an abundance of HER2-HER3 heterodimers, which are known to be potent inducers of cell growth and transformation. Our new approach to use parameter dependence analysis in experimental design reveals that measurements of HER3 phosphorylation and HER2 internalization ratio may prove to be especially useful for the estimation of critical model parameters. Further, we examine the effect of receptor dimerization patterns on biological response using a simple phenomenological model. Results indicate that coexpression of EGFR with HER2 and HER3 at low to moderate levels may enable cells to match the response of a high HER2 expresser. PMID:16533841

Protein-disulfide Isomerase Regulates the Thyroid Hormone Receptor-mediated Gene Expression via Redox Factor-1 through Thiol Reduction-Oxidation*

PubMed Central

Hashimoto, Shoko; Imaoka, Susumu

2013-01-01

Protein-disulfide isomerase (PDI) is a dithiol/disulfide oxidoreductase that regulates the redox state of proteins. We previously found that overexpression of PDI in rat pituitary tumor (GH3) cells suppresses 3,3′,5-triiodothyronine (T3)-stimulated growth hormone (GH) expression, suggesting the contribution of PDI to the T3-mediated gene expression via thyroid hormone receptor (TR). In the present study, we have clarified the mechanism of regulation by which TR function is regulated by PDI. Overexpression of wild-type but not redox-inactive mutant PDI suppressed the T3-induced GH expression, suggesting that the redox activity of PDI contributes to the suppression of GH. We considered that PDI regulates the redox state of the TR and focused on redox factor-1 (Ref-1) as a mediator of the redox regulation of TR by PDI. Interaction between Ref-1 and TRβ1 was detected. Overexpression of wild-type but not C64S Ref-1 facilitated the GH expression, suggesting that redox activity of Cys-64 in Ref-1 is involved in the TR-mediated gene expression. Moreover, PDI interacted with Ref-1 and changed the redox state of Ref-1, suggesting that PDI controls the redox state of Ref-1. Our studies suggested that Ref-1 contributes to TR-mediated gene expression and that the redox state of Ref-1 is regulated by PDI. Redox regulation of PDI via Ref-1 is a new aspect of PDI function. PMID:23148211

The Interleukin 3 Gene (IL3) Contributes to Human Brain Volume Variation by Regulating Proliferation and Survival of Neural Progenitors

PubMed Central

Huang, Liang; Nho, Kwangsik; Deng, Min; Chen, Qiang; Weinberger, Daniel R.; Vasquez, Alejandro Arias; Rijpkema, Mark; Mattay, Venkata S.; Saykin, Andrew J.; Shen, Li; Fernández, Guillén; Franke, Barbara; Chen, Jing-chun; Chen, Xiang-ning; Wang, Jin-kai; Xiao, Xiao; Qi, Xue-bin; Xiang, Kun; Peng, Ying-Mei; Cao, Xiang-yu; Li, Yi; Shi, Xiao-dong; Gan, Lin; Su, Bing

2012-01-01

One of the most significant evolutionary changes underlying the highly developed cognitive abilities of humans is the greatly enlarged brain volume. In addition to being far greater than in most other species, the volume of the human brain exhibits extensive variation and distinct sexual dimorphism in the general population. However, little is known about the genetic mechanisms underlying normal variation as well as the observed sex difference in human brain volume. Here we show that interleukin-3 (IL3) is strongly associated with brain volume variation in four genetically divergent populations. We identified a sequence polymorphism (rs31480) in the IL3 promoter which alters the expression of IL3 by affecting the binding affinity of transcription factor SP1. Further analysis indicated that IL3 and its receptors are continuously expressed in the developing mouse brain, reaching highest levels at postnatal day 1–4. Furthermore, we found IL3 receptor alpha (IL3RA) was mainly expressed in neural progenitors and neurons, and IL3 could promote proliferation and survival of the neural progenitors. The expression level of IL3 thus played pivotal roles in the expansion and maintenance of the neural progenitor pool and the number of surviving neurons. Moreover, we found that IL3 activated both estrogen receptors, but estrogen didn’t directly regulate the expression of IL3. Our results demonstrate that genetic variation in the IL3 promoter regulates human brain volume and reveals novel roles of IL3 in regulating brain development. PMID:23226269

Quantification of STAT3 and VEGF expression for molecular diagnosis of lymph node metastasis in breast cancer

PubMed Central

Chen, Yujuan; Liu, Ya; Wang, Yu; Li, Wen; Wang, Xiaolu; Liu, Xuejuan; Chen, Yao; Ouyang, Chibin; Wang, Jing

2017-01-01

Abstract Background: Axillary lymph node metastasis is associated with increased risk of regional recurrence, distant metastasis, and poor survival in breast malignant neoplasm. Expression of signal transducer and activator of transcription 3 (STAT3) is significantly associated with tumor formation, migration, and invasion in various cancers. In addition, vascular endothelial growth factor (VEGF) expression could promote angiogenesis and increase the risk of tumorigenesis. To determine correlations among STAT3 expression, VEGF, and clinicopathological data on lymph node involvement in breast cancer patients after surgery. Methods: The mRNA expression levels of STAT3 and VEGFs were measured in 45 breast invasive ductal carcinoma tissues, 45 peritumoral tissues, and 45 adjacent nontumor tissues by real-time quantitative reverse transcription-polymerase chain reaction (RT-qPCR). Postoperative pathological examination revealed explicit axillary lymph node involvement in all patients. Results: Average mRNA levels of STAT3 and VEGFs were the highest in breast invasive ductal carcinoma tissues, followed by peritumoral tissues. High expression of STAT3 showed significant positive correlation with high axillary lymph node involvement and progesterone receptor (PR), VEGF-C, VEGF-D, and vascular endothelial growth factor receptor (VEGFR)-3 expression. The expression levels of STAT3, VEGF-C, and VEGFR-3 were significantly higher in the tumor tissues of patients with axillary lymph node metastasis than in those of patients without the metastasis. Expression levels of VEGF-C and VEGFR-3 were also significantly higher in peritumoral tissues of patients with axillary lymph node metastasis. Positive correlations were found between STAT3 and VEGF-C/-D mRNA levels. Conclusion: These data suggest that STAT3/VEGF-C/VEGFR-3 signaling pathway plays an important role in carcinogenesis and lymph-angiogenesis. Our findings suggest that STAT3 may be a potential molecular biomarker for predicting the involvement of axillary lymph nodes in breast cancer, and therapies targeting STAT3 may be important for preventing breast cancer metastasis. PMID:29137038

Changes in expression and secretion patterns of fibroblast growth factor 8 and Sonic Hedgehog signaling pathway molecules during murine neural stem/progenitor cell differentiation in vitro☆

PubMed Central

Lu, Jiang; Lu, Kehuan; Li, Dongsheng

2012-01-01

In the present study, we investigated the dynamic expression of fibroblast growth factor 8 and Sonic Hedgehog signaling pathway related factors in the process of in vitro hippocampal neural stem/progenitor cell differentiation from embryonic Sprague-Dawley rats or embryonic Kunming species mice, using fluorescent quantitative reverse transcription-PCR and western blot analyses. Results demonstrated that the dynamic expression of fibroblast growth factor 8 was similar to fibroblast growth factor receptor 1 expression but not to other fibroblast growth factor receptors. Enzyme-linked immunosorbent assay demonstrated that fibroblast growth factor 8 and Sonic Hedgehog signaling pathway protein factors were secreted by neural cells into the intercellular niche. Our experimental findings indicate that fibroblast growth factor 8 and Sonic Hedgehog expression may be related to the differentiation of neural stem/progenitor cells. PMID:25624789

Development of EMab-51, a Sensitive and Specific Anti-Epidermal Growth Factor Receptor Monoclonal Antibody in Flow Cytometry, Western Blot, and Immunohistochemistry.

PubMed

Itai, Shunsuke; Kaneko, Mika K; Fujii, Yuki; Yamada, Shinji; Nakamura, Takuro; Yanaka, Miyuki; Saidoh, Noriko; Handa, Saori; Chang, Yao-Wen; Suzuki, Hiroyoshi; Harada, Hiroyuki; Kato, Yukinari

2017-10-01

The epidermal growth factor receptor (EGFR) is a member of the human epidermal growth factor receptor (HER) family of receptor tyrosine kinases and is involved in cell growth and differentiation. EGFR homodimers or heterodimers with other HER members, such as HER2 and HER3, activate downstream signaling cascades in many cancers. In this study, we developed novel anti-EGFR monoclonal antibodies (mAbs) and characterized their efficacy in flow cytometry, Western blot, and immunohistochemical analyses. First, we expressed the full-length or ectodomain of EGFR in LN229 glioblastoma cells and then immunized mice with LN229/EGFR or ectodomain of EGFR, and performed the first screening using enzyme-linked immunosorbent assays. Subsequently, we selected mAbs according to their efficacy in flow cytometry (second screening), Western blot (third screening), and immunohistochemical (fourth screening) analyses. Among 100 mAbs, only one clone EMab-51 (IgG 1 , kappa) reacted with EGFR in Western blot analysis. Finally, immunohistochemical analyses with EMab-51 showed sensitive and specific reactions against oral cancer cells, warranting the use of EMab-51 to detect EGFR in pathological analyses of EGFR-expressing cancers.

Gene therapy of murine teratocarcinoma: separate functions for insulin-like growth factors I and II in immunogenicity and differentiation.

PubMed Central

Trojan, J; Johnson, T R; Rudin, S D; Blossey, B K; Kelley, K M; Shevelev, A; Abdul-Karim, F W; Anthony, D D; Tykocinski, M L; Ilan, J

1994-01-01

Teratocarcinoma is a germ-line carcinoma giving rise to an embryoid tumor with structures derived from the three embryonic layers: mesoderm, endoderm, and ectoderm. Teratocarcinoma is widely used as an in vitro model system to study regulation of cell determination and differentiation during mammalian embryogenesis. Murine embryonic carcinoma (EC) PCC3 cells express insulin-like growth factor I(IGF-I) and its receptor, while all derivative tumor structures express IGF-I and IGF-II and their receptors. Therefore the system lends itself to dissect the role of these two growth factors during EC differentiation. With an episomal antisense strategy, we define a role for IGF-I in tumorigenicity and evasion of immune surveillance. Antisense IGF-I EC transfectants are shown to elicit a curative anti-tumor immune response with tumor regression at distal sites. In contrast, IGF-II is shown to drive determination and differentiation in EC cells. Since IGF-I and IGF-II bind to type I receptor and antisense sequence used for IGF-II cannot form duplex with endogenous IGF-I transcripts, it follows that this receptor is not involved in determination and differentiation. Images PMID:8016120

Effects of glucose, insulin and triiodothyroxine on leptin and leptin receptor expression and the effects of leptin on activities of enzymes related to glucose metabolism in grass carp (Ctenopharyngodon idella) hepatocytes.

PubMed

Lu, Rong-Hua; Zhou, Yi; Yuan, Xiao-Chen; Liang, Xu-Fang; Fang, Liu; Bai, Xiao-Li; Wang, Min; Zhao, Yu-Hua

2015-08-01

Leptin is an important regulator of appetite and energy expenditure in mammals, but its role in fish metabolism control is poorly understood. Our previous studies demonstrated that leptin has an effect on the regulation of food intake and energy expenditure as well as lipid metabolism (stimulation of lipolysis and inhibition of adipogenesis) in the grass carp Ctenopharyngodon idella. To further investigate the role of leptin in fish, the effects of glucose, insulin and triiodothyroxine (T3) on the expression levels of leptin and leptin receptor (Lepr) and the effects of leptin on the activities of critical glucose metabolism enzymes in grass carp hepatocytes were evaluated in the present study. Our data indicated that leptin gene expression was induced by glucose in a dose-dependent manner, while Lepr gene expression exhibited a biphasic change. A high dose of insulin (100 ng/mL) significantly up-regulated the expression of leptin and Lepr. Leptin expression was markedly up-regulated by a low concentration of T3 but inhibited by a high concentration of T3. T3 up-regulated Lepr expression in a dose-dependent manner. Together, these data suggest that leptin had a close relationship with three factors (glucose, insulin and T3) and might participate in the regulation of glucose metabolism in grass carp. In addition, we also found that leptin affected the activities of key enzymes that are involved in glucose metabolism, which might be mediated by insulin receptor substrate-phosphoinositol 3-kinase signaling.

Airborne fine particulate matter causes murine bronchial hyperreactivity via MAPK pathway-mediated M3 muscarinic receptor upregulation.

PubMed

Wang, Rong; Xiao, Xue; Shen, Zhenxing; Cao, Lei; Cao, Yongxiao

2017-02-01

Regarding the human health effects, airborne fine particulate matter 2.5 (PM 2.5 ) is an important environmental risk factor. However, the underlying molecular mechanisms are largely unknown. The present study examined the hypothesis that PM 2.5 causes bronchial hyperreactivity by upregulated muscarinic receptors via the mitogen-activated protein kinase (MAPK) pathway. The isolated rat bronchi segments were cultured with different concentration of PM 2.5 for different time. The contractile response of the bronchi segments were recorded by a sensitive myograph. The mRNA and protein expression levels of M 3 muscarinic receptors were studied by quantitative real-time PCR and immunohistochemistry, respectively. The muscarinic receptors agonist, carbachol induced a remarkable contractile response on fresh and DMSO cultured bronchial segments. Compared with the fresh or DMSO culture groups, 1.0 µg/mL of PM 2.5 cultured for 24 h significantly enhanced muscarinic receptor-mediated contractile responses in bronchi with a markedly increased maximal contraction. In addition, the expression levels of mRNA and protein for M 3 muscarinic receptors in bronchi of PM 2.5 group were higher than that of fresh or DMSO culture groups. SB203580 (p38 inhibitor) and U0126 (MEK1/2 inhibitor) significantly inhibited the PM 2.5 -induced enhanced contraction and increased mRNA and protein expression of muscarinic receptors. However, JNK inhibitor SP600125 had no effect on PM 2.5 -induced muscarinic receptor upregulation and bronchial hyperreactivity. In conclusion, airborne PM 2.5 upregulates muscarinic receptors, which causes subsequently bronchial hyperreactivity shown as enhanced contractility in bronchi. This process may be mediated by p38 and MEK1/2 MAPK pathways. © 2016 Wiley Periodicals, Inc. Environ Toxicol 32: 371-381, 2017. © 2016 Wiley Periodicals, Inc.

Growth factors FGF8 and FGF2 and their receptor FGFR1, transcriptional factors Msx-1 and MSX-2, and apoptotic factors p19 and RIP5 participate in the early human limb development.

PubMed

Becic, Tina; Kero, Darko; Vukojevic, Katarina; Mardesic, Snjezana; Saraga-Babic, Mirna

2018-04-01

The expression pattern of fibroblast growth factors FGF8 and FGF2 and their receptor FGFR1, transcription factors MSX-1 and MSX-2, as well as cell proliferation (Ki-67) and cell death associated caspase-3, p19 and RIP5 factors were analyzed in histological sections of eight 4th-9th-weeks developing human limbs by immunohistochemistry and semi-thin sectioning. Increasing expression of all analyzed factors (except FGF8) characterized both the multilayered human apical ectodermal ridge (AER), sub-ridge mesenchyme (progress zone) and chondrocytes in developing human limbs. While cytoplasmic co-expression of MSX-1 and MSX-2 was observed in both limb epithelium and mesenchyme, p19 displayed strong cytoplasmic expression in non-proliferating cells. Nuclear expression of Ki-67 proliferating cells, and partly of MSX-1 and MSX-2 was detected in the whole limb primordium. Strong expression of factors p19 and RIP5, both in the AER and mesenchyme of human developing limbs indicates their possible involvement in control of cell senescence and cell death. In contrast to animal studies, expression of FGFR1 in the surface ectoderm and p19 in the whole limb primordium might reflect interspecies differences in limb morphology. Expression of FGF2 and downstream RIP5 gene, and transcription factors Msx-1 and MSX-2 did not show human-specific changes in expression pattern. Based on their spatio-temporal expression during human limb development, our study indicates role of FGFs and Msx genes in stimulation of cell proliferation, limb outgrowth, digit elongation and separation, and additionally MSX-2 in control of vasculogenesis. The cascade of orchestrated gene expressions, including the analyzed developmental factors, jointly contribute to the complex human limb development. Copyright © 2018 Elsevier GmbH. All rights reserved.

Apoptosis gene expression and death receptor signaling in mitomycin-C-treated human tenon capsule fibroblasts.

PubMed

Crowston, Jonathan G; Chang, Lydia H; Constable, Peter H; Daniels, Julie T; Akbar, Arne N; Khaw, Peng T

2002-03-01

To examine the effect of mitomycin-C on the expression of apoptosis genes in human Tenon capsule fibroblasts and to evaluate whether death receptor signaling modulates mitomycin-C cytotoxicity. Bcl-2, Bax, Bcl-x, Fas (CD95) and tumor necrosis factor (TNF) receptor expression was determined by flow cytometry in control and mitomycin-C-treated Tenon fibroblasts. Fibroblast death was quantified using a lactate dehydrogenase release assay. The effect of Fas and TNF-receptor signaling was evaluated using Fas-specific antibodies and soluble TNF-alpha. Tenon fibroblasts constitutively express Bcl-2, Bax, and Bcl-x in culture. Mitomycin-C (0.4 mg/mL) induced a small but consistent increase in the expression of all three proteins. Tenon fibroblasts express low levels of Fas but are resistant to the effects of Fas-receptor ligation. Mitomycin-C (0.01-1.0 mg/mL) led to a significant increase in Fas expression at all concentrations tested (P < 0.01). Pretreatment with mitomycin-C (0.4 mg/mL) rendered fibroblasts susceptible to agonistic anti-Fas monoclonal IgM antibodies (50-500 ng/mL) and led to a further 50% reduction in viable fibroblasts at 48 hours, compared with mitomycin-C alone (P < 0.05). Antibodies that block the Fas receptor did not inhibit mitomycin-C-induced apoptosis. Mitomycin-C alters apoptosis gene expression and primes fibroblasts to the effects of Fas receptor ligation. Factors other than the level of Fas receptor expression modulate the response to Fas receptor signaling. Determining the signals that regulate fibroblast apoptosis may help to refine therapeutic strategies for switching off the subconjunctival healing response and maintaining intraocular pressure control.

Anti-sense suppression of epidermal growth factor receptor expression alters cellular proliferation, cell-adhesion and tumorigenicity in ovarian cancer cells.

PubMed

Alper, O; De Santis, M L; Stromberg, K; Hacker, N F; Cho-Chung, Y S; Salomon, D S

2000-11-15

Over-expression of epidermal growth factor receptor (EGFR) in ovarian cancer has been well documented. Human NIH:OVCAR-8 ovarian carcinoma cells were transfected with an expression vector containing the anti-sense orientation of truncated human EGFR cDNA. EGFR anti-sense over-expression resulted in decreased EGFR protein and mRNA expression, cell proliferation and tumor formation in nude mice. In accordance with the reduced levels of EGFR in EGFR anti-sense-expressing cells, tyrosine phosphorylation of EGFR was decreased compared to untransfected parental cells treated with EGF. In EGFR anti-sense-transfected cells, expression of erbB-3, but not erbB-2, was increased. In addition, basal and heregulin-beta 1-stimulated tyrosine phosphorylation of erbB-3 was higher in EGFR anti-sense vector-transfected cells. A morphological alteration in EGFR anti-sense gene-expressing cells was correlated with a decrease in the expression of E-cadherin, alpha-catenin and, to a lesser extent, beta-catenin. Changes in the expression of these proteins were associated with a reduction in complex formation among E-cadherin, beta-catenin and alpha-catenin and between beta-catenin and EGFR in EGFR anti-sense-expressing cells compared to sense-transfected control cells. These results demonstrate that EGFR expression in ovarian carcinoma cells regulates expression of cell adhesion proteins that may enhance cell growth and invasiveness. Copyright 2000 Wiley-Liss, Inc.

NEUROTROPHIN SELECTIVITY IN ORGANIZING TOPOGRAPHIC REGENERATION OF NOCICEPTIVE AFFERENTS

PubMed Central

Kelamangalath, Lakshmi; Tang, Xiaoqing; Bezik, Kathleen; Sterling, Noelle; Son, Young-Jin; Smith, George M.

2015-01-01

Neurotrophins represent some of the best candidates to enhance regeneration. In the current study, we investigated the effects of artemin, a member of the glial derived neurotrophic factor (GDNF) family, on sensory axon regeneration following a lumbar dorsal root injury and compared these effects with that observed after either NGF or GDNF expression in the rat spinal cord. Unlike previously published data, artemin failed to induce regeneration of large-diameter myelinated sensory afferents when expressed within either the spinal cord or DRG. However, artemin or NGF induced regeneration of calcitonin gene related peptide positive (CGRP+) axons only when expressed within the spinal cord. Accordingly, artemin or NGF enhanced recovery of only nociceptive behavior and showed a cFos distribution similar to the topography of regenerating axons. Artemin and GDNF signaling requires binding to different co-receptors (GFRα3 or GFRα1, respectively) prior to binding to the signaling receptor, cRet. Approximately 70% of DRG neurons express cRet, but only 35% express either co-receptor. To enhance artemin-induced regeneration, we co-expressed artemin with either GFRα3 or GDNF. Co-expression of artemin and GFRα3 only slightly enhanced regeneration of IB4+ non-peptidergic nociceptive axons, but not myelinated axons. Interestingly, this co-expression also disrupted the ability of artemin to produce topographic targeting and lead to significant increases in cFos immunoreactivity within the deep dorsal laminae. This study failed to demonstrate artemin-induced regeneration of myelinated axons, even with co-expression of GFR-α3, which only promoted mistargeted regeneration. PMID:26054884

Neurotrophin selectivity in organizing topographic regeneration of nociceptive afferents.

PubMed

Kelamangalath, Lakshmi; Tang, Xiaoqing; Bezik, Kathleen; Sterling, Noelle; Son, Young-Jin; Smith, George M

2015-09-01

Neurotrophins represent some of the best candidates to enhance regeneration. In the current study, we investigated the effects of artemin, a member of the glial derived neurotrophic factor (GDNF) family, on sensory axon regeneration following a lumbar dorsal root injury and compared these effects with that observed after either NGF or GDNF expression in the rat spinal cord. Unlike previously published data, artemin failed to induce regeneration of large-diameter myelinated sensory afferents when expressed within either the spinal cord or DRG. However, artemin or NGF induced regeneration of calcitonin gene related peptide positive (CGRP(+)) axons only when expressed within the spinal cord. Accordingly, artemin or NGF enhanced recovery of only nociceptive behavior and showed a cFos distribution similar to the topography of regenerating axons. Artemin and GDNF signaling requires binding to different co-receptors (GFRα3 or GFRα1, respectively) prior to binding to the signaling receptor, cRet. Approximately 70% of DRG neurons express cRet, but only 35% express either co-receptor. To enhance artemin-induced regeneration, we co-expressed artemin with either GFRα3 or GDNF. Co-expression of artemin and GFRα3 only slightly enhanced regeneration of IB4(+) non-peptidergic nociceptive axons, but not myelinated axons. Interestingly, this co-expression also disrupted the ability of artemin to produce topographic targeting and lead to significant increases in cFos immunoreactivity within the deep dorsal laminae. This study failed to demonstrate artemin-induced regeneration of myelinated axons, even with co-expression of GFRα3, which only promoted mistargeted regeneration. Copyright © 2015 Elsevier Inc. All rights reserved.

C2C12 myotubes inhibit the proliferation and differentiation of 3T3-L1 preadipocytes by reducing the expression of glucocorticoid receptor gene

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

Chu, Weiwei; Wei, Wei; Yu, Shigang

Obesity is a well-established risk factor to health for its relationship with insulin resistance, diabetes and metabolic syndrome. Myocyte-adipocyte crosstalk model plays a significant role in studying the interaction of muscle and adipose development. Previous related studies mainly focus on the effects of adipocytes on the myocytes activity, however, the influence of myotubes on the preadipocytes development remains unclear. The present study was carried out to settle this issue. Firstly, the co-culture experiment showed that the proliferation, cell cycle, and differentiation of 3T3-L1 preadipocytes were arrested, and the apoptosis was induced, by differentiated C2C12 myotubes. Next, the sensitivity of 3T3-L1more » preadipocytes to glucocorticoids (GCs), which was well known as cell proliferation, differentiation, apoptosis factor, was decreased after co-cultured with C2C12 myotubes. What's more, our results showed that C2C12 myotubes suppressed the mRNA and protein expression of glucocorticoid receptor (GR) in 3T3-L1 preadipocytes, indicating the potential mechanism of GCs sensitivity reduction. Taken together, we conclude that C2C12 myotubes inhibited 3T3-L1 preadipocytes proliferation and differentiation by reducing the expression of GR. These data suggest that decreasing GR by administration of myokines may be a promising therapy for treating patients with obesity or diabetes. - Highlights: • C2C12 myotubes inhibited proliferation and differentiation of 3T3-L1 preadipocytes. • C2C12 myotubes arrested cell cycle of 3T3-L1 preadipocytes. • C2C12 myotubes induced apoptosis of 3T3-L1 preadipocytes. • C2C12 inhibit 3T3-L1 cells by reducing the expression of glucocorticoid receptor gene.« less

Lenalidomide enhances the function of chimeric antigen receptor T cells against the epidermal growth factor receptor variant III by enhancing immune synapses.

PubMed

Kuramitsu, S; Ohno, M; Ohka, F; Shiina, S; Yamamichi, A; Kato, A; Tanahashi, K; Motomura, K; Kondo, G; Kurimoto, M; Senga, T; Wakabayashi, T; Natsume, A

2015-10-01

The epidermal growth factor receptor variant III (EGFRvIII) is exclusively expressed on the cell surface in ~50% of glioblastoma multiforme (GBM). This variant strongly and persistently activates the phosphatidylinositol 3-kinase-Akt signaling pathway in a ligand-independent manner resulting in enhanced tumorigenicity, cellular motility and resistance to chemoradiotherapy. Our group generated a recombinant single-chain variable fragment (scFv) antibody specific to the EGFRvIII, referred to as 3C10-scFv. In the current study, we constructed a lentiviral vector transducing the chimeric antigen receptor (CAR) that consisted of 3C10-scFv, CD3ζ, CD28 and 4-1BB (3C10-CAR). The 3C10-CAR-transduced peripheral blood mononuclear cells (PBMCs) and CD3(+) T cells specifically lysed the glioma cells that express EGFRvIII. Moreover, we demonstrated that CAR CD3(+) T cells migrated to the intracranial xenograft of GBM in the mice treated with 3C10-CAR PBMCs. An important and novel finding of our study was that a thalidomide derivative lenalidomide induced 3C10-CAR PBMC proliferation and enhanced the persistent antitumor effect of the cells in vivo. Lenalidomide also exhibited enhanced immunological synapses between the effector cells and the target cells as determined by CD11a and F-actin polymerization. Collectively, lentiviral-mediated transduction of CAR effectors targeting the EGFRvIII showed specific efficacy, and lenalidomide even intensified CAR cell therapy by enhanced formation of immunological synapses.

Expression of the peroxisome proliferator-activated receptor γ (PPARγ) in human atherosclerosis and regulation in macrophages by colony stimulating factors and oxidized low density lipoprotein

PubMed Central

Ricote, Mercedes; Huang, Jannet; Fajas, Luis; Li, Andrew; Welch, John; Najib, Jamila; Witztum, Joseph L.; Auwerx, Johan; Palinski, Wulf; Glass, Christopher K.

1998-01-01

The peroxisome proliferator-activated receptor γ (PPARγ) is a ligand-dependent transcription factor that has been demonstrated to regulate fat cell development and glucose homeostasis. PPARγ is also expressed in a subset of macrophages and negatively regulates the expression of several proinflammatory genes in response to natural and synthetic ligands. We here demonstrate that PPARγ is expressed in macrophage foam cells of human atherosclerotic lesions, in a pattern that is highly correlated with that of oxidation-specific epitopes. Oxidized low density lipoprotein (oxLDL) and macrophage colony-stimulating factor, which are known to be present in atherosclerotic lesions, stimulated PPARγ expression in primary macrophages and monocytic cell lines. PPARγ mRNA expression was also induced in primary macrophages and THP-1 monocytic leukemia cells by the phorbol ester 12-O-tetradecanoylphorbol 13-acetate (TPA). Inhibition of protein kinase C blocked the induction of PPARγ expression by TPA, but not by oxLDL, suggesting that more than one signaling pathway regulates PPARγ expression in macrophages. TPA induced the expression of PPARγ in RAW 264.7 macrophages by increasing transcription from the PPARγ1 and PPARγ3 promoters. In concert, these observations provide insights into the regulation of PPARγ expression in activated macrophages and raise the possibility that PPARγ ligands may influence the progression of atherosclerosis. PMID:9636198

Taste bud-derived BDNF maintains innervation of a subset of TrkB-expressing gustatory nerve fibers.

PubMed

Tang, Tao; Rios-Pilier, Jennifer; Krimm, Robin

2017-07-01

Taste receptor cells transduce different types of taste stimuli and transmit this information to gustatory neurons that carry it to the brain. Taste receptor cells turn over continuously in adulthood, requiring constant new innervation from nerve fibers. Therefore, the maintenance of innervation to taste buds is an active process mediated by many factors, including brain-derived neurotrophic factor (BDNF). Specifically, 40% of taste bud innervation is lost when Bdnf is removed during adulthood. Here we speculated that not all gustatory nerve fibers express the BDNF receptor, TrkB, resulting in subsets of neurons that vary in their response to BDNF. However, it is also possible that the partial loss of innervation occurred because the Bdnf gene was not effectively removed. To test these possibilities, we first determined that not all gustatory nerve fibers express the TrkB receptor in adult mice. We then verified the efficiency of Bdnf removal specifically in taste buds of K14-CreER:Bdnf mice and found that Bdnf expression was reduced to 1%, indicating efficient Bdnf gene recombination. BDNF removal resulted in a 55% loss of TrkB-expressing nerve fibers, which was greater than the loss of P2X3-positive fibers (39%), likely because taste buds were innervated by P2X3+/TrkB- fibers that were unaffected by BDNF removal. We conclude that gustatory innervation consists of both TrkB-positive and TrkB-negative taste fibers and that BDNF is specifically important for maintaining TrkB-positive innervation to taste buds. In addition, although taste bud size was not affected by inducible Bdnf removal, the expression of the γ subunit of the ENaC channel was reduced. So, BDNF may regulate expression of some molecular components of taste transduction pathways. Copyright © 2017. Published by Elsevier Inc.

Steroid hormone and epidermal growth factor receptors in meningiomas.

PubMed

Horsfall, D J; Goldsmith, K G; Ricciardelli, C; Skinner, J M; Tilley, W D; Marshall, V R

1989-11-01

A prospective study of steroid hormone and epidermal growth factor receptor expression in 57 meningiomas is presented. Scatchard analysis of radioligand binding identified 20% of meningiomas as expressing classical oestrogen receptors (ER) at levels below that normally accepted for positivity, the remainder being negative. ER could not be visualized in any meningioma using immunocytochemistry. Alternatively, 74% of meningiomas demonstrated the presence of progesterone receptors (PR) by Scatchard analysis, the specificity of which could not be attributed to glucocorticoid or androgen receptors. Confirmation of classical PR presence was determined by immunocytochemical staining. The presence of epidermal growth factor receptor (EGFR) was demonstrated in 100% of meningiomas using immunocytochemical staining. These data are reviewed in the context of previously reported results and are discussed in relation to the potential for medical therapy as an adjunct to surgery.

Neurotrophin/receptor expression in urinary bladder of mice with overexpression of NGF in urothelium.

PubMed

Girard, Beatrice M; Malley, Susan E; Vizzard, Margaret A

2011-02-01

Urothelium-specific overexpression of nerve growth factor (NGF) in the urinary bladder of transgenic mice stimulates neuronal sprouting in the urinary bladder, produces increased voiding frequency, and results in increased referred somatic hypersensitivity. Additional NGF-mediated pleiotropic changes might contribute to the increased voiding frequency and pelvic hypersensitivity observed in these transgenic mice, such as modulation of other growth factor/receptor systems. Chronic overexpression of NGF in the urothelium was achieved through the use of a highly urothelium-specific uroplakin II promoter. In the present study, we examined NGF, brain-derived neurotrophic factor (BDNF), and associated receptor [p75(NTR), tyrosine kinase (Trk)A, TrkB] transcript and protein expression in urothelium and detrusor smooth muscle of NGF-overexpressing (OE) and littermate wild-type mice, using real-time quantitative reverse transcription-polymerase chain reaction, ELISAs, and semiquantitation of immunohistochemistry. We focused on these growth factor/receptors given the established roles of NGF/TrkA, NGF/p75(NTR), and BDNF/TrkB systems in bladder function. Increased voiding frequency in NGF-OE mice was confirmed by examining urination patterns. BDNF, TrkA, and TrkB protein expression was significantly (P ≤ 0.01) reduced and p75(NTR) protein expression was significantly (P ≤ 0.01) increased in urinary bladder of NGF-OE mice. The NGF-OE-induced changes in neurotrophic factor/receptor expression in urinary bladder may represent compensatory changes to reduce voiding frequency in the NGF-OE mouse.

Insulin-Like Growth Factors Are Expressed in the Taste System, but Do Not Maintain Adult Taste Buds.

PubMed

Biggs, Bradley T; Tang, Tao; Krimm, Robin F

2016-01-01

Growth factors regulate cell growth and differentiation in many tissues. In the taste system, as yet unknown growth factors are produced by neurons to maintain taste buds. A number of growth factor receptors are expressed at greater levels in taste buds than in the surrounding epithelium and may be receptors for candidate factors involved in taste bud maintenance. We determined that the ligands of eight of these receptors were expressed in the E14.5 geniculate ganglion and that four of these ligands were expressed in the adult geniculate ganglion. Of these, the insulin-like growth factors (IGF1, IGF2) were expressed in the ganglion and their receptor, insulin-like growth factor receptor 1 (IGF1R), were expressed at the highest levels in taste buds. To determine whether IGF1R regulates taste bud number or structure, we conditionally eliminated IGF1R from the lingual epithelium of mice using the keratin 14 (K14) promoter (K14-Cre::Igf1rlox/lox). While K14-Cre::Igf1rlox/lox mice had significantly fewer taste buds at P30 compared with control mice (Igf1rlox/lox), this difference was not observed by P80. IGF1R removal did not affect taste bud size or cell number, and the number of phospholipase C β2- (PLCβ2) and carbonic anhydrase 4- (Car4) positive taste receptor cells did not differ between genotypes. Taste buds at the back of the tongue fungiform taste field were larger and contained more cells than those at the tongue tip, and these differences were diminished in K14-Cre::Igf1rlox/lox mice. The epithelium was thicker at the back versus the tip of the tongue, and this difference was also attenuated in K14-Cre::Igf1rlox/lox mice. We conclude that, although IGFs are expressed at high levels in the taste system, they likely play little or no role in maintaining adult taste bud structure. IGFs have a potential role in establishing the initial number of taste buds, and there may be limits on epithelial thickness in the absence of IGF1R signaling.

The cannabinoid receptor CB1 modulates the signaling properties of the lysophosphatidylinositol receptor GPR55.

PubMed

Kargl, Julia; Balenga, Nariman; Parzmair, Gerald P; Brown, Andrew J; Heinemann, Akos; Waldhoer, Maria

2012-12-28

The G protein-coupled receptor (GPCR) 55 (GPR55) and the cannabinoid receptor 1 (CB1R) are co-expressed in many tissues, predominantly in the central nervous system. Seven transmembrane spanning (7TM) receptors/GPCRs can form homo- and heteromers and initiate distinct signaling pathways. Recently, several synthetic CB1 receptor inverse agonists/antagonists, such as SR141716A, AM251, and AM281, were reported to activate GPR55. Of these, SR141716A was marketed as a promising anti-obesity drug, but was withdrawn from the market because of severe side effects. Here, we tested whether GPR55 and CB1 receptors are capable of (i) forming heteromers and (ii) whether such heteromers could exhibit novel signaling patterns. We show that GPR55 and CB1 receptors alter each others signaling properties in human embryonic kidney (HEK293) cells. We demonstrate that the co-expression of FLAG-CB1 receptors in cells stably expressing HA-GPR55 specifically inhibits GPR55-mediated transcription factor activation, such as nuclear factor of activated T-cells and serum response element, as well as extracellular signal-regulated kinases (ERK1/2) activation. GPR55 and CB1 receptors can form heteromers, but the internalization of both receptors is not affected. In addition, we observe that the presence of GPR55 enhances CB1R-mediated ERK1/2 and nuclear factor of activated T-cell activation. Our data provide the first evidence that GPR55 can form heteromers with another 7TM/GPCR and that this interaction with the CB1 receptor has functional consequences in vitro. The GPR55-CB1R heteromer may play an important physiological and/or pathophysiological role in tissues endogenously co-expressing both receptors.

The Cannabinoid Receptor CB1 Modulates the Signaling Properties of the Lysophosphatidylinositol Receptor GPR55*

PubMed Central

Kargl, Julia; Balenga, Nariman; Parzmair, Gerald P.; Brown, Andrew J.; Heinemann, Akos; Waldhoer, Maria

2012-01-01

The G protein-coupled receptor (GPCR) 55 (GPR55) and the cannabinoid receptor 1 (CB1R) are co-expressed in many tissues, predominantly in the central nervous system. Seven transmembrane spanning (7TM) receptors/GPCRs can form homo- and heteromers and initiate distinct signaling pathways. Recently, several synthetic CB1 receptor inverse agonists/antagonists, such as SR141716A, AM251, and AM281, were reported to activate GPR55. Of these, SR141716A was marketed as a promising anti-obesity drug, but was withdrawn from the market because of severe side effects. Here, we tested whether GPR55 and CB1 receptors are capable of (i) forming heteromers and (ii) whether such heteromers could exhibit novel signaling patterns. We show that GPR55 and CB1 receptors alter each others signaling properties in human embryonic kidney (HEK293) cells. We demonstrate that the co-expression of FLAG-CB1 receptors in cells stably expressing HA-GPR55 specifically inhibits GPR55-mediated transcription factor activation, such as nuclear factor of activated T-cells and serum response element, as well as extracellular signal-regulated kinases (ERK1/2) activation. GPR55 and CB1 receptors can form heteromers, but the internalization of both receptors is not affected. In addition, we observe that the presence of GPR55 enhances CB1R-mediated ERK1/2 and nuclear factor of activated T-cell activation. Our data provide the first evidence that GPR55 can form heteromers with another 7TM/GPCR and that this interaction with the CB1 receptor has functional consequences in vitro. The GPR55-CB1R heteromer may play an important physiological and/or pathophysiological role in tissues endogenously co-expressing both receptors. PMID:23161546

β5 Integrin Up-Regulation in Brain-Derived Neurotrophic Factor Promotes Cell Motility in Human Chondrosarcoma

PubMed Central

Li, Te-Mao; Fong, Yi-Chin; Liu, Shan-Chi; Chen, Po-Chun; Tang, Chih-Hsin

2013-01-01

Chondrosarcoma is a primary malignant bone cancer, with a potent capacity to invade locally and cause distant metastasis; it has a poor prognosis and shows a predilection for metastasis to the lungs. Brain derived neurotrophic factor (BDNF) is a small-molecule protein from the neurotrophin family of growth factors that is associated with the disease status and outcomes of cancers. However, the effect of BDNF on migration activity in human chondrosarcoma cells is mostly unknown. Here, we found that human chondrosarcoma tissues showed significant expression of BDNF, which was higher than that in normal cartilage and primary chondrocytes. We also found that BDNF increased the migration and expression of β5 integrin in human chondrosarcoma cells. In addition, knockdown of BDNF expression markedly inhibited migratory activity. BDNF-mediated migration and β5 integrin up-regulation were attenuated by antibody, inhibitor, or siRNA against the TrkB receptor. Pretreatment of chondrosarcoma cells with PI3K, Akt, and NF-κB inhibitors or mutants also abolished BDNF-promoted migration and integrin expression. The PI3K, Akt, and NF-κB signaling pathway was activated after BDNF treatment. Taken together, our results indicate that BDNF enhances the migration of chondrosarcoma by increasing β5 integrin expression through a signal transduction pathway that involves the TrkB receptor, PI3K, Akt, and NF-κB. BDNF thus represents a promising new target for treating chondrosarcoma metastasis. PMID:23874483

Serotonin regulates brain-derived neurotrophic factor expression in select brain regions during acute psychological stress

PubMed Central

Jiang, De-guo; Jin, Shi-li; Li, Gong-ying; Li, Qing-qing; Li, Zhi-ruo; Ma, Hong-xia; Zhuo, Chuan-jun; Jiang, Rong-huan; Ye, Min-jie

2016-01-01

Previous studies suggest that serotonin (5-HT) might interact with brain-derived neurotrophic factor (BDNF) during the stress response. However, the relationship between 5-HT and BDNF expression under purely psychological stress is unclear. In this study, one hour before psychological stress exposure, the 5-HT1A receptor agonist 8-OH-DPAT or antagonist MDL73005, or the 5-HT2A receptor agonist DOI or antagonist ketanserin were administered to rats exposed to psychological stress. Immunohistochemistry and in situ hybridization revealed that after psychological stress, with the exception of the ventral tegmental area, BDNF protein and mRNA expression levels were higher in the 5-HT1A and the 5-HT2A receptor agonist groups compared with the solvent control no-stress or psychological stress group in the CA1 and CA3 of the hippocampus, prefrontal cortex, central amygdaloid nucleus, dorsomedial hypothalamic nucleus, dentate gyrus, shell of the nucleus accumbens and the midbrain periaqueductal gray. There was no significant difference between the two agonist groups. In contrast, after stress exposure, BDNF protein and mRNA expression levels were lower in the 5-HT1A and 5-HT2A receptor antagonist groups than in the solvent control non-stress group, with the exception of the ventral tegmental area. Our findings suggest that 5-HT regulates BDNF expression in a rat model of acute psychological stress. PMID:27857753

The aryl hydrocarbon receptor and estrogen receptor alpha differentially modulate nuclear factor erythroid-2-related factor 2 transactivation in MCF-7 breast cancer cells

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

Lo, Raymond; Matthews, Jason, E-mail: jason.matthews@utoronto.ca

2013-07-15

Nuclear factor erythroid-2-related factor 2 (NRF2; NFE2L2) plays an important role in mediating cellular protection against reactive oxygen species. NRF2 signaling is positively modulated by the aryl hydrocarbon receptor (AHR) but inhibited by estrogen receptor alpha (ERα). In this study we investigated the crosstalk among NRF2, AHR and ERα in MCF-7 breast cancer cells treated with the NRF2 activator sulforaphane (SFN), a dual AHR and ERα activator, 3,3′-diindolylmethane (DIM), 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) or 17β-estradiol (E2). SFN-dependent increases in NADPH-dependent oxidoreductase 1 (NQO1) and heme oxygenase I (HMOX1) mRNA levels were significantly reduced after co-treatment with E2. E2-dependent repression of NQO1 andmore » HMOX1 was associated with increased ERα but reduced p300 recruitment and reduced histone H3 acetylation at both genes. In contrast, DIM + SFN or TCDD + SFN induced NQO1 and HMOX1 mRNA expression to levels higher than SFN alone, which was prevented by RNAi-mediated knockdown of AHR. DIM + SFN but not TCDD + SFN also induced recruitment of ERα to NQO1 and HMOX1. However, the presence of AHR at NQO1 and HMOX1 restored p300 recruitment and histone H3 acetylation, thereby reversing the ERα-dependent repression of NRF2. Taken together, our study provides further evidence of functional interplay among NRF2, AHR and ERα signaling pathways through altered p300 recruitment to NRF2-regulated target genes. - Highlights: • We examined crosstalk among ERα, AHR, and NRF2 in MCF-7 breast cancer cells. • AHR enhanced the mRNA expression levels of two NRF2 target genes – HMOX1 and NQO1. • ERα repressed HMOX1 and NQO1 expression via decreased histone acetylation. • AHR prevented ERα-dependent repression of HMOX1 and NQO1.« less

Expression and significance of Tie-1 and Tie-2 receptors, and angiopoietins-1, 2 and 4 in colorectal adenocarcinoma: Immunohistochemical analysis and correlation with clinicopathological factors

PubMed Central

Nakayama, Toshiyuki; Hatachi, Go; Wen, Chun-Yang; Yoshizaki, Ayumi; Yamazumi, Kazuyuki; Niino, Daisuke; Sekine, Ichiro

2005-01-01

AIM: There is strong evidence that tyrosine kinases are involved in the regulation of tumor progression, cellular growth and differentiation. Recently, many kinds of tyrosine kinase receptors have been reported, among them Tie-1 and Tie-2 receptors constitute a major class. Angiopoietin (Ang)-1 is known as a ligand of Tie-2 tyrosine kinase receptor. The objective of this study was to establish a comprehensive Tie-1 and Tie-2 and Ang-1, 2 and 4 expression profile in human colorectal adenocarcinomas. METHODS: We examined 96 cases of surgically resected human colorectal adenocarcinoma by immunohistochemistry and investigated the statistical correlation between the expressions of Ties and Angs and clinicopathological factors. RESULTS: Among the 96 cases of adenocarcinoma, 87 (90.6%), 92 (95.8%), 83 (86.5%), 89 (92.7%), and 76 cases (79.2%) showed positive staining in the cytoplasm of carcinoma cells for the Tie-1 and Tie-2 and Ang-1, 2 and 4 proteins, respectively. Histologically, the expressions of Ties and Angs were variable. The expressions of Ties and Angs were correlated with several clinicopathological factors, but did not correlate with the presence of lymph node metastasis. Ties and Angs were highly expressed in human colorectal adenocarcinoma cells. CONCLUSION: These findings suggest that the Tie-Ang receptor-ligand complex is one of the factors involved in the cellular differentiation and progression of human colorectal adenocarcinoma. PMID:15742397

Growth Factors and COX2 Expression in Canine Perivascular Wall Tumors.

PubMed

Avallone, G; Stefanello, D; Boracchi, P; Ferrari, R; Gelain, M E; Turin, L; Tresoldi, E; Roccabianca, P

2015-11-01

Canine perivascular wall tumors (PWTs) are a group of subcutaneous soft tissue sarcomas developing from vascular mural cells. Mural cells are involved in angiogenesis through a complex crosstalk with endothelial cells mediated by several growth factors and their receptors. The evaluation of their expression may have relevance since they may represent a therapeutic target in the control of canine PWTs. The expression of vascular endothelial growth factor (VEGF) and receptors VEGFR-I/II, basic fibroblast growth factor (bFGF) and receptor Flg, platelet-derived growth factor B (PDGFB) and receptor PDGFRβ, transforming growth factor β1 (TGFβ1) and receptors TGFβR-I/II, and cyclooxygenase 2 (COX2) was evaluated on frozen sections of 40 PWTs by immunohistochemistry and semiquantitatively scored to identify their potential role in PWT development. Statistical analysis was performed to analyze possible correlations between Ki67 labeling index and the expression of each molecule. Proteins of the VEGF-, PDGFB-, and bFGF-mediated pathways were highly expressed in 27 (67.5%), 30 (75%), and 19 (47.5%) of 40 PWTs, respectively. Proteins of the TGFβ1- and COX2-mediated pathways were highly expressed in 4 (10%) and 14 (35%) of 40 cases. Statistical analysis identified an association between VEGF and VEGFR-I/II (P = .015 and .003, respectively), bFGF and Flg (P = .038), bFGF and PDGFRβ (P = .003), and between TGFβ1 and COX2 (P = .006). These findings were consistent with the mechanisms that have been reported to play a role in angiogenesis and in tumor development. No association with Ki67 labeling index was found. VEGF-, PDGFB-, and bFGF-mediated pathways seem to have a key role in PWT development and growth. Blockade of tyrosine kinase receptors after surgery could represent a promising therapy with the aim to reduce the PWT relapse rate and prolong the time to relapse. © The Author(s) 2015.

Influence of Panax ginseng on Alpha-Adrenergic Receptor of Benign Prostatic Hyperplasia

PubMed Central

Kim, Su Kang; Chung, Joo-Ho; Lee, Byung-Cheol; Lee, Sang Won; Lee, Kang Hyo

2014-01-01

Purpose Benign prostatic hyperplasia (BPH) is the most common prostate problem in older men. The present study aimed to investigate the inhibitory effect of Panax ginseng C.A. Meyer (P. ginseng) on a rat model of testosterone-induced BPH. Methods The rats were divided into 3 groups (each group, n=10): control, testosterone-induced BPH (20 mg/kg, subcutaneous injection), and P. ginseng (200 mg/kg, orally) groups. After 4 weeks, all animals were sacrificed to examine the blood biochemical profiles, prostate volume, weight, histopathological changes, alpha-1D adrenergic receptor (Adra1d) mRNA expression, and epidermal growth factor receptor (EGFR) and B-cell CLL/lymphoma 2 (BCL2) protein expression. Results The group treated with P. ginseng showed significantly lesser prostate size and weight than the testosterone-induced BPH group. In addition, P. ginseng decreased the mRNA expression of Adra1d as well as the expression of EGFR and BCL2 in prostate tissue. Conclusions These results suggest that P. ginseng may inhibit the alpha-1-adrenergic receptor to suppress the development of BPH. PMID:25558416

Estrogen receptor 1 (ESR1) regulates VEGFA in adipose tissue.

PubMed

Fatima, L A; Campello, R S; Santos, R de Souza; Freitas, H S; Frank, A P; Machado, U F; Clegg, D J

2017-12-01

Vascular endothelial growth factor A (VEGFA) is a key factor in the regulation of angiogenesis in adipose tissue. Poor vascularization during adipose tissue proliferation causes fibrosis and local inflammation, and is associated with insulin resistance. It is known that 17-beta estradiol (E2) regulates adipose tissue function and VEGFA expression in other tissues; however, the ability of E2 to regulate VEGFA in adipose tissue is currently unknown. In this study, we showed that, in 3T3-L1 cells, E2 and the estrogen receptor 1 (ESR1) agonist PPT induced VEGFA expression, while ESR1 antagonist (MPP), and selective knockdown of ESR1 using siRNA decreased VEGFA and prevented the ability of E2 to modulate its expression. Additionally, we found that E2 and PPT induced the binding of hypoxia inducible factor 1 alpha subunit (HIF1A) in the VEGFA gene promoter. We further found that VEGFA expression was lower in inguinal and gonadal white adipose tissues of ESR1 total body knockout female mice compared to wild type mice. In conclusion, our data provide evidence of an important role for E2/ESR1 in modulating adipose tissue VEGFA, which is potentially important to enhance angiogenesis, reduce inflammation and improve adipose tissue function.

Gene Expression Analysis Implicates a Death Receptor Pathway in Schizophrenia Pathology

PubMed Central

Catts, Vibeke Sørensen; Shannon Weickert, Cynthia

2012-01-01

An increase in apoptotic events may underlie neuropathology in schizophrenia. By data-mining approaches, we identified significant expression changes in death receptor signaling pathways in the dorsolateral prefrontal cortex (DLPFC) of patients with schizophrenia, particularly implicating the Tumor Necrosis Factor Superfamily member 6 (FAS) receptor and the Tumor Necrosis Factor [ligand] Superfamily member 13 (TNFSF13) in schizophrenia. We sought to confirm and replicate in an independent tissue collection the noted mRNA changes with quantitative real-time RT-PCR. To test for regional and diagnostic specificity, tissue from orbital frontal cortex (OFC) was examined and a bipolar disorder group included. In schizophrenia, we confirmed and replicated significantly increased expression of TNFSF13 mRNA in the DLPFC. Also, a significantly larger proportion of subjects in the schizophrenia group had elevated FAS receptor expression in the DLPFC relative to unaffected controls. These changes were not observed in the bipolar disorder group. In the OFC, there were no significant differences in TNFSF13 or FAS receptor mRNA expression. Decreases in BH3 interacting domain death agonist (BID) mRNA transcript levels were found in the schizophrenia and bipolar disorder groups affecting both the DLPFC and the OFC. We tested if TNFSF13 mRNA expression correlated with neuronal mRNAs in the DLPFC, and found significant negative correlations with interneuron markers, parvalbumin and somatostatin, and a positive correlation with PPP1R9B (spinophilin), but not DLG4 (PSD-95). The expression of TNFSF13 mRNA in DLPFC correlated negatively with tissue pH, but decreasing pH in cultured cells did not cause increased TNFSF13 mRNA nor did exogenous TNFSF13 decrease pH. We concluded that increased TNFSF13 expression may be one of several cell-death cytokine abnormalities that contribute to the observed brain pathology in schizophrenia, and while increased TNFSF13 may be associated with lower brain pH, the change is not necessarily causally related to brain pH. PMID:22545112

Innate immune responses induced by lipopolysaccharide and lipoteichoic acid in primary goat mammary epithelial cells.

PubMed

Bulgari, Omar; Dong, Xianwen; Roca, Alfred L; Caroli, Anna M; Loor, Juan J

2017-01-01

Innate immune responses induced by in vitro stimulation of primary mammary epithelial cells (MEC) using Gram-negative lipopolysaccharide (LPS) and Gram-positive lipoteichoic acid (LTA) bacterial cell wall components are well- characterized in bovine species. The objective of the current study was to characterize the downstream regulation of the inflammatory response induced by Toll-like receptors in primary goat MEC (pgMEC). We performed quantitative real-time RT-PCR (qPCR) to measure mRNA levels of 9 genes involved in transcriptional regulation or antibacterial activity: Toll-like receptor 2 ( TLR2 ), Toll-like receptor 4 ( TLR4 ), prostaglandin-endoperoxide synthase 2 ( PTGS2 ), interferon induced protein with tetratricopeptide repeats 3 ( IFIT3 ), interferon regulatory factor 3 ( IRF3 ), myeloid differentiation primary response 88 ( MYD88 ), nuclear factor of kappa light polypeptide gene enhancer in B-cells 1 ( NFKB1 ), Toll interacting protein ( TOLLIP ), and lactoferrin ( LTF ). Furthermore, we analyzed 7 cytokines involved in Toll-like receptor signaling pathways: C-C motif chemokine ligand 2 ( CCL2 ), C-C motif chemokine ligand 5 ( CCL5 ), C-X-C motif chemokine ligand 6 ( CXCL6 ), interleukin 8 ( CXCL8 ), interleukin 1 beta ( IL1B ), interleukin 6 ( IL6 ), and tumor necrosis factor alpha ( TNF ). Stimulation of pgMEC with LPS for 3 h led to an increase in expression of CCL2 , CXCL6 , IL6 , CXCL8 , PTGS2 , IFIT3 , MYD88 , NFKB1 , and TLR4 ( P  < 0.05). Except for IL6 , and PTGS2 , the same genes had greater expression than controls at 6 h post-LPS ( P  < 0.05). Expression of CCL5 , PTGS2 , IFIT3 , NFKB1 , TLR4 , and TOLLIP was greater than controls after 3 h of incubation with LTA ( P  < 0.05). Compared to controls, stimulation with LTA for 6 h led to greater expression of PTGS2 , IFIT3 , NFKB1 , and TOLLIP ( P  < 0.05) whereas the expression of CXCL6 , CXCL8 , and TLR4 was lower ( P  < 0.05). At 3 h incubation with both toxins compared to controls a greater expression ( P  < 0.05) of CCL2 , CCL5 , CXCL6 , CXCL8 , IL6 , PTGS2 , IFIT3 , IRF3 , MYD88 , and NFKB1 was detected. After 6 h of incubation with both toxins, the expression of CCL2 , CXCL6 , IFIT3 , MYD88 , NFKB1 , and TLR4 was higher than the controls ( P  < 0.05). Data indicate that in the goat MEC, LTA induces a weaker inflammatory response than LPS. This may be related to the observation that gram-positive bacteria cause chronic mastitis more often than gram-negative infections.

β3-adrenergic receptor activation induces TGFβ1 expression in cardiomyocytes via the PKG/JNK/c-Jun pathway.

PubMed

Xu, Zhongcheng; Wu, Jimin; Xin, Junzhou; Feng, Yenan; Hu, Guomin; Shen, Jing; Li, Mingzhe; Zhang, Youyi; Xiao, Han; Wang, Li

2018-06-05

In heart failure, the expression of cardiac β 3 -adrenergic receptors (β 3 -ARs) increases. However, the precise role of β 3 -AR signaling within cardiomyocytes remains unclear. Transforming growth factor β1 (TGFβ1) is a crucial cytokine mediating the cardiac remodeling that plays a causal role in the progression of heart failure. Here, we set out to determine the effect of β 3 -AR activation on TGFβ1 expression in rat cardiomyocytes and examine the underlying mechanism. The selective β 3 -AR agonist BRL37344 induced an increase in TGFβ1 expression and the phosphorylation of c-Jun N-terminal kinase (JNK) and c-Jun in β 3 -AR-overexpressing cardiomyocytes. Those effects of BRL37344 were suppressed by a β 3 -AR antagonist. Moreover, the inhibition of JNK and c-Jun activity by a JNK inhibitor and c-Jun siRNA blocked the increase in TGFβ1 expression upon β 3 -AR activation. A protein kinase G (PKG) inhibitor also attenuated β 3 -AR-agonist-induced TGFβ1 expression and the phosphorylation of JNK and c-Jun. In conclusion, the β 3 -AR activation in cardiomyocytes increases the expression of TGFβ1 via the PKG/JNK/c-Jun pathway. These results help us further understand the role of β 3 -AR signaling in heart failure. Copyright © 2018 Elsevier Inc. All rights reserved.

The Transfection of BDNF to Dopamine Neurons Potentiates the Effect of Dopamine D3 Receptor Agonist Recovering the Striatal Innervation, Dendritic Spines and Motor Behavior in an Aged Rat Model of Parkinson’s Disease

PubMed Central

Razgado-Hernandez, Luis F.; Espadas-Alvarez, Armando J.; Reyna-Velazquez, Patricia; Sierra-Sanchez, Arturo; Anaya-Martinez, Veronica; Jimenez-Estrada, Ismael; Bannon, Michael J.; Martinez-Fong, Daniel; Aceves-Ruiz, Jorge

2015-01-01

The progressive degeneration of the dopamine neurons of the pars compacta of substantia nigra and the consequent loss of the dopamine innervation of the striatum leads to the impairment of motor behavior in Parkinson’s disease. Accordingly, an efficient therapy of the disease should protect and regenerate the dopamine neurons of the substantia nigra and the dopamine innervation of the striatum. Nigral neurons express Brain Derived Neurotropic Factor (BDNF) and dopamine D3 receptors, both of which protect the dopamine neurons. The chronic activation of dopamine D3 receptors by their agonists, in addition, restores, in part, the dopamine innervation of the striatum. Here we explored whether the over-expression of BDNF by dopamine neurons potentiates the effect of the activation of D3 receptors restoring nigrostriatal innervation. Twelve-month old Wistar rats were unilaterally injected with 6-hydroxydopamine into the striatum. Five months later, rats were treated with the D3 agonist 7-hydroxy-N,N-di-n-propy1-2-aminotetralin (7-OH-DPAT) administered i.p. during 4½ months via osmotic pumps and the BDNF gene transfection into nigral cells using the neurotensin-polyplex nanovector (a non-viral transfection) that selectively transfect the dopamine neurons via the high-affinity neurotensin receptor expressed by these neurons. Two months after the withdrawal of 7-OH-DPAT when rats were aged (24 months old), immunohistochemistry assays were made. The over-expression of BDNF in rats receiving the D3 agonist normalized gait and motor coordination; in addition, it eliminated the muscle rigidity produced by the loss of dopamine. The recovery of motor behavior was associated with the recovery of the nigral neurons, the dopamine innervation of the striatum and of the number of dendritic spines of the striatal neurons. Thus, the over-expression of BDNF in dopamine neurons associated with the chronic activation of the D3 receptors appears to be a promising strategy for restoring dopamine neurons in Parkinson’s disease. PMID:25693197

Poly(ADP-ribose) Polymerase Inhibitors Sensitize Cancer Cells to Death Receptor-mediated Apoptosis by Enhancing Death Receptor Expression*

PubMed Central

Meng, X. Wei; Koh, Brian D.; Zhang, Jin-San; Flatten, Karen S.; Schneider, Paula A.; Billadeau, Daniel D.; Hess, Allan D.; Smith, B. Douglas; Karp, Judith E.; Kaufmann, Scott H.

2014-01-01

Recombinant human tumor necrosis factor-α-related apoptosis inducing ligand (TRAIL), agonistic monoclonal antibodies to TRAIL receptors, and small molecule TRAIL receptor agonists are in various stages of preclinical and early phase clinical testing as potential anticancer drugs. Accordingly, there is substantial interest in understanding factors that affect sensitivity to these agents. In the present study we observed that the poly(ADP-ribose) polymerase (PARP) inhibitors olaparib and veliparib sensitize the myeloid leukemia cell lines ML-1 and K562, the ovarian cancer line PEO1, non-small cell lung cancer line A549, and a majority of clinical AML isolates, but not normal marrow, to TRAIL. Further analysis demonstrated that PARP inhibitor treatment results in activation of the FAS and TNFRSF10B (death receptor 5 (DR5)) promoters, increased Fas and DR5 mRNA, and elevated cell surface expression of these receptors in sensitized cells. Chromatin immunoprecipitation demonstrated enhanced binding of the transcription factor Sp1 to the TNFRSF10B promoter in the presence of PARP inhibitor. Knockdown of PARP1 or PARP2 (but not PARP3 and PARP4) not only increased expression of Fas and DR5 at the mRNA and protein level, but also recapitulated the sensitizing effects of the PARP inhibition. Conversely, Sp1 knockdown diminished the PARP inhibitor effects. In view of the fact that TRAIL is part of the armamentarium of natural killer cells, these observations identify a new facet of PARP inhibitor action while simultaneously providing the mechanistic underpinnings of a novel therapeutic combination that warrants further investigation. PMID:24895135

FGFR3 gene mutation plus GRB10 gene duplication in a patient with achondroplasia plus growth delay with prenatal onset.

PubMed

Yuan, Haiming; Huang, Linhuan; Hu, Xizi; Li, Qian; Sun, Xiaofang; Xie, Yingjun; Kong, Shu; Wang, Xiaoman

2016-07-02

Achondroplasia is a well-defined and common bone dysplasia. Genotype- and phenotype-level correlations have been found between the clinical symptoms of achondroplasia and achondroplasia-specific FGFR3 mutations. A 2-year-old boy with clinical features consistent with achondroplasia and Silver-Russell syndrome-like symptoms was found to carry a mutation in the fibroblast growth factor receptor-3 (FGFR3) gene at c.1138G > A (p.Gly380Arg) and a de novo 574 kb duplication at chromosome 7p12.1 that involved the entire growth-factor receptor bound protein 10 (GRB10) gene. Using quantitative real-time PCR analysis, GRB10 was over-expressed, and, using enzyme-linked immunosorbent assays for IGF1 and IGF-binding protein-3 (IGFBP3), we found that IGF1 and IGFBP3 were low-expressed in this patient. We demonstrate that a combination of uncommon, rare and exceptional molecular defects related to the molecular bases of particular birth defects can be analyzed and diagnosed to potentially explain the observed variability in the combination of molecular defects.

Insights into GABA receptor signalling in TM3 Leydig cells.

PubMed

Doepner, Richard F G; Geigerseder, Christof; Frungieri, Monica B; Gonzalez-Calvar, Silvia I; Calandra, Ricardo S; Raemsch, Romi; Fohr, Karl; Kunz, Lars; Mayerhofer, Artur

2005-01-01

Gamma-aminobutyric acid (GABA) is an emerging signalling molecule in endocrine organs, since it is produced by endocrine cells and acts via GABA(A) receptors in a paracrine/autocrine fashion. Testicular Leydig cells are producers and targets for GABA. These cells express GABA(A) receptor subunits and in the murine Leydig cell line TM3 pharmacological activation leads to increased proliferation. The signalling pathway of GABA in these cells is not known in this study. We therefore attempted to elucidate details of GABA(A) signalling in TM3 and adult mouse Leydig cells using several experimental approaches. TM3 cells not only express GABA(A )receptor subunits, but also bind the GABA agonist [(3)H]muscimol with a binding affinity in the range reported for other endocrine cells (K(d) = 2.740 +/- 0.721 nM). However, they exhibit a low B(max) value of 28.08 fmol/mg protein. Typical GABA(A) receptor-associated events, including Cl(-) currents, changes in resting membrane potential, intracellular Ca(2+) or cAMP, were not measurable with the methods employed in TM3 cells, or, as studied in part, in primary mouse Leydig cells. GABA or GABA(A) agonist isoguvacine treatment resulted in increased or decreased levels of several mRNAs, including transcription factors (c-fos, hsf-1, egr-1) and cell cycle-associated genes (Cdk2, cyclin D1). In an attempt to verify the cDNA array results and because egr-1 was recently implied in Leydig cell development, we further studied this factor. RT-PCR and Western blotting confirmed a time-dependent regulation of egr-1 in TM3. In the postnatal testis egr-1 was seen in cytoplasmic and nuclear locations of developing Leydig cells, which bear GABA(A) receptors and correspond well to TM3 cells. Thus, GABA acts via an atypical novel signalling pathway in TM3 cells. Further details of this pathway remain to be elucidated. Copyright (c) 2005 S. Karger AG, Basel.

Extrinsic factors regulate partial agonist efficacy of strychnine-sensitive glycine receptors

PubMed Central

Farroni, Jeffrey S; McCool, Brian A

2004-01-01

Background Strychnine-sensitive glycine receptors in many adult forebrain regions consist of alpha2 + beta heteromeric channels. This subunit composition is distinct from the alpha1 + beta channels found throughout the adult spinal cord. Unfortunately, the pharmacology of forebrain alpha2beta receptors are poorly defined compared to 'neonatal' alpha2 homomeric channels or 'spinal' alpha1beta heteromers. In addition, the pharmacologic properties of native alpha2beta glycine receptors have been generally distinct from receptors produced by heterologous expression. To identify subtype-specific pharmacologic tools for the forebrain alpha2beta receptors, it is important to identify a heterologous expression system that closely resembles these native glycine-gated chloride channels. Results While exploring pharmacological properties of alpha2beta glycine receptors compared to alpha2-homomers, we found that distinct heterologous expression systems appeared to differentially influence partial agonist pharmacology. The β-amino acid taurine possessed 30–50% efficacy for alpha2-containing receptor isoforms when expressed in HEK 293 cells. However, taurine efficacy was dramatically reduced in L-cell fibroblasts. Similar results were obtained for β-alanine. The efficacy of these partial agonists was also strongly reduced by the beta subunit. There were no significant differences in apparent strychnine affinity values calculated from concentration-response data between expression systems or subunit combinations. Nor did relative levels of expression correlate with partial agonist efficacy when compared within or between several different expression systems. Finally, disruption of the tubulin cytoskeleton reduced the efficacy of partial agonists in a subunit-dependent, but system-independent, fashion. Conclusions Our results suggest that different heterologous expression systems can dramatically influence the agonist pharmacology of strychnine-sensitive glycine receptors. In the systems examine here, these effects are independent of both absolute expression level and any system-related alterations in the agonist binding site. We conclude that complex interactions between receptor composition and extrinsic factors may play a significant role in determining strychnine-sensitive glycine receptor partial agonist pharmacology. PMID:15301692

Extrinsic factors regulate partial agonist efficacy of strychnine-sensitive glycine receptors.

PubMed

Farroni, Jeffrey S; McCool, Brian A

2004-08-09

Strychnine-sensitive glycine receptors in many adult forebrain regions consist of alpha2 + beta heteromeric channels. This subunit composition is distinct from the alpha1 + beta channels found throughout the adult spinal cord. Unfortunately, the pharmacology of forebrain alpha2beta receptors are poorly defined compared to 'neonatal' alpha2 homomeric channels or 'spinal' alpha1beta heteromers. In addition, the pharmacologic properties of native alpha2beta glycine receptors have been generally distinct from receptors produced by heterologous expression. To identify subtype-specific pharmacologic tools for the forebrain alpha2beta receptors, it is important to identify a heterologous expression system that closely resembles these native glycine-gated chloride channels. While exploring pharmacological properties of alpha2beta glycine receptors compared to alpha2-homomers, we found that distinct heterologous expression systems appeared to differentially influence partial agonist pharmacology. The beta-amino acid taurine possessed 30-50% efficacy for alpha2-containing receptor isoforms when expressed in HEK 293 cells. However, taurine efficacy was dramatically reduced in L-cell fibroblasts. Similar results were obtained for beta-alanine. The efficacy of these partial agonists was also strongly reduced by the beta subunit. There were no significant differences in apparent strychnine affinity values calculated from concentration-response data between expression systems or subunit combinations. Nor did relative levels of expression correlate with partial agonist efficacy when compared within or between several different expression systems. Finally, disruption of the tubulin cytoskeleton reduced the efficacy of partial agonists in a subunit-dependent, but system-independent, fashion. Our results suggest that different heterologous expression systems can dramatically influence the agonist pharmacology of strychnine-sensitive glycine receptors. In the systems examine here, these effects are independent of both absolute expression level and any system-related alterations in the agonist binding site. We conclude that complex interactions between receptor composition and extrinsic factors may play a significant role in determining strychnine-sensitive glycine receptor partial agonist pharmacology.

The type I BMP receptors, Bmpr1a and Acvr1, activate multiple signaling pathways to regulate lens formation

PubMed Central

Rajagopal, Ramya; Huang, Jie; Dattilo, Lisa K.; Kaartinen, Vesa; Mishina, Yuji; Deng, Chu-Xia; Umans, Lieve; Zwijsen, An; Roberts, Anita B.; Beebe, David C.

2009-01-01

BMPs play multiple roles in development and BMP signaling is essential for lens formation. However, the mechanisms by which BMP receptors function in vertebrate development are incompletely understood. To determine the downstream effectors of BMP signaling and their functions in the ectoderm that will form the lens, we deleted the genes encoding the type I BMP receptors, Bmpr1a and Acvr1, and the canonical transducers of BMP signaling, Smad4, Smad1 and Smad5. Bmpr1a and Acvr1 regulated cell survival and proliferation, respectively. Absence of both receptors interfered with the expression of proteins involved in normal lens development and prevented lens formation, demonstrating that BMPs induce lens formation by acting directly on the prospective lens ectoderm. Remarkably, the canonical Smad signaling pathway was not needed for most of these processes. Lens formation, placode cell proliferation, the expression of FoxE3, a lens-specific transcription factor, and the lens protein, αA-crystallin were regulated by BMP receptors in a Smad-independent manner. Placode cell survival was promoted by R-Smad signaling, but in a manner that did not involve Smad4. Of the responses tested, only maintaining a high level of Sox2 protein, a transcription factor expressed early in placode formation, required the canonical Smad pathway. A key function of Smad-independent BMP receptor signaling may be reorganization of actin cytoskeleton to drive lens invagination. PMID:19733164

Genomic characterization and regulation of CYP3a13: role of xenobiotics and nuclear receptors.

PubMed

Anakk, Sayeepriyadarshini; Kalsotra, Auinash; Shen, Qi; Vu, Mary T; Staudinger, Jeffrey L; Davies, Peter J A; Strobel, Henry W

2003-09-01

We report that CYP3a13 gene, located on mouse chromosome 5, spans 27.5 Kb and contains 13 exons. The transcription start site is 35 bp upstream of the coding region and results in a 109 bp 5' untranslated region. CYP3a13 promoter shows putative binding sites for retinoid X receptor, pregnane X receptor, and estrogen receptor. CYP3a13 shows a broad tissue distribution with predominant expression in liver. Although CYP3a13 shares 92% nucleotide identity with the female-specific rat CYP3A9, its expression does not exhibit sexual dimorphism. Ligand activation of peroxisomal proliferator-activated receptor-gamma and retinoid X receptor inhibit expression of CYP3a13 at the transcription level in a tissue-specific manner. Another novel finding is hepatic induction of CYP3a13 by dexamethasone occurring only in pregnane X receptor null mice. We also report that pregnane X receptor is essential to maintain robust in vivo basal levels of CYP3a13 in contrast to CYP3a11. CYP3a13 protein expressed in vitro can metabolize clinically active drugs ethylmorphine and erythromycin, as well as benzphetamine. We conclude that CYP3a13 is regulated differentially by various nuclear receptors. In humans this may lead to altered drug metabolism, as many of the newly synthesized ligands/drugs targeted toward these nuclear receptors could influence CYP3A gene expression.

The Neurotensin Receptor-1 Pathway Contributes to Human Ductal Breast Cancer Progression

PubMed Central

Dupouy, Sandra; Viardot-Foucault, Véronique; Alifano, Marco; Souazé, Frédérique; Plu-Bureau, Geneviève; Chaouat, Marc; Lavaur, Anne; Hugol, Danielle; Gespach, Christian

2009-01-01

Background The neurotensin (NTS) and its specific high affinity G protein coupled receptor, the NT1 receptor (NTSR1), are considered to be a good candidate for one of the factors implicated in neoplastic progression. In breast cancer cells, functionally expressed NT1 receptor coordinates a series of transforming functions including cellular migration and invasion. Methods and Results we investigated the expression of NTS and NTSR1 in normal human breast tissue and in invasive ductal breast carcinomas (IDCs) by immunohistochemistry and RT-PCR. NTS is expressed and up-regulated by estrogen in normal epithelial breast cells. NTS is also found expressed in the ductal and invasive components of IDCs. The high expression of NTSR1 is associated with the SBR grade, the size of the tumor, and the number of metastatic lymph nodes. Furthermore, the NTSR1 high expression is an independent factor of prognosis associated with the death of patients. Conclusion these data support the activation of neurotensinergic deleterious pathways in breast cancer progression. PMID:19156213

The neurotensin receptor-1 pathway contributes to human ductal breast cancer progression.

PubMed

Dupouy, Sandra; Viardot-Foucault, Véronique; Alifano, Marco; Souazé, Frédérique; Plu-Bureau, Geneviève; Chaouat, Marc; Lavaur, Anne; Hugol, Danielle; Gespach, Christian; Gompel, Anne; Forgez, Patricia

2009-01-01

The neurotensin (NTS) and its specific high affinity G protein coupled receptor, the NT1 receptor (NTSR1), are considered to be a good candidate for one of the factors implicated in neoplastic progression. In breast cancer cells, functionally expressed NT1 receptor coordinates a series of transforming functions including cellular migration and invasion. we investigated the expression of NTS and NTSR1 in normal human breast tissue and in invasive ductal breast carcinomas (IDCs) by immunohistochemistry and RT-PCR. NTS is expressed and up-regulated by estrogen in normal epithelial breast cells. NTS is also found expressed in the ductal and invasive components of IDCs. The high expression of NTSR1 is associated with the SBR grade, the size of the tumor, and the number of metastatic lymph nodes. Furthermore, the NTSR1 high expression is an independent factor of prognosis associated with the death of patients. these data support the activation of neurotensinergic deleterious pathways in breast cancer progression.

Immunohistochemical examination of effects of kefir, koumiss and commercial probiotic capsules on platelet derived growth factor-c and platelet derived growth factor receptor-alpha expression in mouse liver and kidney.

PubMed

Bakir, B; Sari, E K; Aydin, B D; Yildiz, S E

2015-04-01

We investigated using immunohistochemistry the effects of kefir, koumiss and commercial probiotic capsules on the expression of platelet derived growth factor-c (PDGF-C) and platelet derived growth factor receptor-alpha (PDGFR-α) in mouse liver and kidney. Mice were assigned to four groups: group 1 was given commercial probiotic capsules, group 2 was given kefir, group 3 was given koumiss and group 4 was untreated. After oral administration for 15 days, body weights were recorded and liver and kidney tissue samples were obtained. Hematoxylin and eosin staining was used to examine histology. PDGF-C and PDGFR-α in liver and kidney were localized using the streptavidin-biotin peroxidase complex method (ABC). We found that the weights of the mice in the kefir, koumiss and commercial probiotic capsules groups increased compared to the control group. No differences in liver and kidney histology were observed in any of the experimental groups. Kefir, koumiss and the commercial probiotic preparation increased PDGF-C and PDGFR-α expression.

Insidious adrenocortical insufficiency underlies neuroendocrine dysregulation in TIF-2 deficient mice.

PubMed

Patchev, Alexandre V; Fischer, Dieter; Wolf, Siegmund S; Herkenham, Miles; Götz, Franziska; Gehin, Martine; Chambon, Pierre; Patchev, Vladimir K; Almeida, Osborne F X

2007-01-01

The transcription-intermediary-factor-2 (TIF-2) is a coactivator of the glucocorticoid receptor (GR), and its disruption would be expected to influence glucocorticoid-mediated control of the hypothalamo-pituitary-adrenal (HPA) axis. Here, we show that its targeted deletion in mice is associated with altered expression of several glucocorticoid-dependent components of HPA regulation (e.g., corticotropin-releasing hormone, vasopressin, ACTH, glucocorticoid receptors), suggestive of hyperactivity under basal conditions. At the same time, TIF-2(-/-) mice display significantly lower basal corticosterone levels and a sluggish and blunted initial secretory response to brief emotional and prolonged physical stress. Subsequent analysis revealed this discrepancy to result from pronounced aberrations in the structure and function of the adrenal gland, including the cytoarchitectural organization of the zona fasciculata and basal and stress-induced expression of key elements of steroid hormone synthesis, such as the steroidogenic acute regulatory (StAR) protein and 3beta-hydroxysteroid dehydrogenase (3beta-HSD). In addition, altered expression levels of two nuclear receptors, DAX-1 and steroidogenic factor 1 (SF-1), in the adrenal cortex strengthen the view that TIF-2 deletion disrupts adrenocortical development and steroid biosynthesis. Thus, hyperactivity of the hypothalamo-pituitary unit is ascribed to insidious adrenal insufficiency and impaired glucocorticoid feedback.

The Antimicrobial Peptide Human Beta-Defensin-3 Is Induced by Platelet-Released Growth Factors in Primary Keratinocytes

PubMed Central

Lammel, Justus; Tohidnezhad, Mersedeh; Lippross, Sebastian; Behrendt, Peter; Klüter, Tim; Pufe, Thomas; Cremer, Jochen; Jahr, Holger; Rademacher, Franziska; Gläser, Regine; Harder, Jürgen

2017-01-01

Platelet-released growth factors (PRGF) and its related clinically used formulations (e.g., Vivostat Platelet-Rich Fibrin (PRF®)) contain a variety of chemokines, cytokines, and growth factors and are therefore used to support healing of chronic, hard-to-heal, or infected wounds. Human beta-defensin-3 (hBD-3) is an antimicrobial peptide inducibly expressed in human keratinocytes especially upon wounding. The potent antimicrobial activity of hBD-3 together with its wound closure-promoting activities suggests that hBD-3 may play a crucial role in wound healing. Therefore, we analyzed the influence of PRGF on hBD-3 expression in human primary keratinocytes in vitro. In addition, we investigated the influence of Vivostat PRF on hBD-3 expression in artificially generated human skin wounds in vivo. PRGF treatment of primary keratinocytes induced a significant, concentration- and time-dependent increase in hBD-3 gene expression which was partially mediated by the epidermal growth factor receptor (EGFR). In line with these cell culture data, in vivo experiments revealed an enhanced hBD-3 expression in experimentally produced human wounds after the treatment with Vivostat PRF. Thus, the induction of hBD-3 may contribute to the beneficial effects of thrombocyte concentrate lysates in the treatment of chronic or infected wounds. PMID:28811680

Uncommon endocytic and trafficking pathway of the natural killer cell CD94/NKG2A inhibitory receptor.

PubMed

Masilamani, Madhan; Narayanan, Sriram; Prieto, Martha; Borrego, Francisco; Coligan, John E

2008-06-01

The CD94/NKG2A inhibitory receptor, expressed by natural killer and T cells, is constantly exposed to its HLA-E ligand expressed by surrounding cells. Ligand exposure often induces receptor downregulation. For CD94/NKG2A, this could potentiate activation receptor(s) induced responses to normal bystander cells. We investigated CD94/NKG2A endocytosis and found that it occurs by an amiloride-sensitive, Rac1-dependent macropinocytic-like process; however, it does not require clathrin, dynamin, ADP ribosylation factor-6, phosphoinositide-3 kinase or the actin cytoskeleton. Once endocytosed, CD94/NKG2A traffics to early endosomal antigen 1(+), Rab5(+) early endosomes. It does appear in Rab4(+) early/sorting endosome, but, in the time period examined, fails to reach Rab11(+) recycling or Rab7(+) late endosomes or lysosome-associated membrane protein-1(+) lysosomes. These results indicate that CD94/NKG2A utilizes a previously undescribed endocytic mechanism coupled with an abbreviated trafficking pattern, perhaps to insure surface expression.

Posttranscriptional mRNA processing as a mechanism for regulation of human A1 adenosine receptor expression.

PubMed Central

Ren, H; Stiles, G L

1994-01-01

The human A1 adenosine receptor gene contains six exons with exons 1, 2, 3, 4, and part of 5 representing 5' untranslated regions. Reverse transcription-PCR with exon-specific primers showed two distinct transcripts containing either exons 3, 5, and 6 or exons 4, 5, and 6, with exons 3 and 4 being mutually exclusive. No mature mRNAs containing exons 1 and 2 have been detected. All human tissues that express any A1 receptors contain mRNA with exons 4, 5, and 6. Tissues which express high levels of A1 receptors contain mRNA with exons 3, 5, and 6. Exon 4 contains two upstream ATG codons whereas exon 3 contains none. COS cells transfected with expression vectors containing exon 4 (exons 1-6, 3-6, or Ex4-6) express much lower levels of A1 receptors than vectors without exon 4 (exons 3, 5, and 6). Mutation of upstream ATG codons in exon 4 leads to 3- to 7-fold increased A1 receptor expression, up to the level seen with the construct containing exons 3, 5, and 6. Thus, in human tissues "basal" levels of A1 receptors can be expressed by use of mRNA containing exons 4, 5, and 6, but when high levels are needed, alternative transcripts with exons 3, 5, and 6 are produced. Images PMID:8197148

Effects of specific and prolonged expression of zebrafish growth factors, Fgf2 and Lif in primordial germ cells in vivo

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

Wong, Ten-Tsao, E-mail: wong20@purdue.edu; Collodi, Paul

2013-01-04

Highlights: Black-Right-Pointing-Pointer We discovered that nanos3 3 Prime UTR prolonged PGC-specific protein expression up to 26 days. Black-Right-Pointing-Pointer Expression of Fgf2 in PGCs significantly increased PGC number at later developmental stages. Black-Right-Pointing-Pointer Expression of Lif in PGCs resulted in a significant disruption of PGC migration. Black-Right-Pointing-Pointer Lif illicited its effect on PGC migration through Lif receptor a. Black-Right-Pointing-Pointer Our approach could be used to achieve prolonged PGC-specific expression of other proteins. -- Abstract: Primordial germ cells (PGCs), specified early in development, proliferate and migrate to the developing gonad before sexual differentiation occurs in the embryo and eventually give rise tomore » spermatogonia or oogonia. In this study, we discovered that nanos3 3 Prime UTR, a common method used to label PGCs, not only directed PGC-specific expression of DsRed but also prolonged this expression up to 26 days post fertilization (dpf) when DsRed-nanos3 3 Prime UTR hybrid mRNAs were introduced into 1- to 2-cell-stage embryos. As such, we employed this knowledge to express zebrafish leukemia inhibitory factor (Lif), basic fibroblast growth factor (Fgf2) and bone morphogenetic protein 4 (Bmp4) in the PGCs and evaluate their effects on PGC development in vivo for over a period of 3 weeks. The results show that expression of Fgf2 significantly increased PGC number at 14- and 21-dpf while Bmp4 resulted in severe ventralization and death of the embryos by 3 days. Expression of Lif resulted in a significant disruption of PGC migration. Mopholino knockdown experiments indicated that Lif illicited its effect on PGC migration through Lif receptor a (Lifra) but not Lifrb. The general approach described in this study could be used to achieve prolonged PGC-specific expression of other proteins to investigate their roles in germ cell and gonad development. The results also indicate that zebrafish PGCs have a mechanism to stabilize and prolong the expression of mRNA that carries nanos3 3 Prime UTR. Understanding this mechanism may make it possible to achieve prolonged RNA expression in other cell types.« less

RANKL downregulates cell surface CXCR6 expression through JAK2/STAT3 signaling pathway during osteoclastogenesis

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

Li, Changhong; Zhao, Jinxia; Sun, Lin

Highlights: Black-Right-Pointing-Pointer CXCR6 is down-regulated during RANKL-induced osteoclastogenesis in RAW264.7 cells. Black-Right-Pointing-Pointer CXCR6 reduction was nearly reversed by inhibition of JAK2/STAT3 signaling pathway. Black-Right-Pointing-Pointer CXCL16 alone does not positively regulate osteoclastogenesis. -- Abstract: The receptor activator of nuclear factor-{kappa}B ligand (RANKL), as a member of the tumor necrosis factor (TNF) family, plays an essential role in osteoclast differentiation and function. Chemokines and their receptors have recently been shown to play critical roles in osteoclastogenesis, however, whether CXCL16-CXCR6 plays role in RANKL-mediated osteoclastogenesis is unknown. In this study, we first reported that RANKL decreased CXCR6 in a dose-dependent manner, which maymore » be through deactivation of Akt and STAT3 signaling induced by CXCL16. Interestingly, RANKL-mediated CXCR6 reduction may be associated to the activation of STAT3 by phosphorylation. When STAT3 activation was blocked by JAK2/STAT3 inhibitor AG490, RANKL failed to shut down CXCR6 expression during osteoclastogenesis. However, CXCL16 alone did not augment RANKL-mediated osteoclast differentiation and did not alter RANKL-receptor RANK mRNA expression. These results demonstrate that reduction of CXCL16-CXCR6 is critical in RANKL-mediated osteoclastogenesis, which is mainly through the activation of JAK2/STAT3 signaling. CXCL16-CXCR6 axis may become a novel target for the therapeutic intervention of bone resorbing diseases such as rheumatoid arthritis and osteoporosis.« less

Altered expressions of fibroblast growth factor receptors and alveolarization in neonatal mice exposed to 85% oxygen.

PubMed

Park, Min Soo; Rieger-Fackeldey, Esther; Schanbacher, Brandon L; Cook, Angela C; Bauer, John A; Rogers, Lynette K; Hansen, Thomas N; Welty, Stephen E; Smith, Charles V

2007-12-01

In the present study, we tested the hypothesis that exposure of newborn mice to sublethal hyperoxia would alter lung development and expressions of fibroblast growth factor receptors (FGFRs)-3 and FGFR-4. Newborn FVB mice were exposed to 85% O2 or maintained in room air for up to 14 d. No animal mortality was observed, and body weight gains were not affected by hyperoxia. At postnatal d 7 and 14 (P7, P14), lungs of mice exposed to 85% O2 showed fewer alveolar secondary crests and larger alveoli or terminal air spaces than did mice in room air. In pups kept in room air, lung levels of FGFR-3 and FGFR-4 mRNA were greater at P3 than at P1, but similar increases were not observed in hyperoxic mice. Immunoreactivity of FGFR-3 and FGFR-4 was lower in lungs of hyperoxic mice than in controls at P14. In pups kept in room air, lung fibroblast growth factor (FGF)-7 mRNA levels were greater at P14 than at P1, but similar changes were not observed in hyperoxic mice. The temporally and spatially specific alterations in the expressions of FGFR-3, FGFR-4, and FGF-7 in the mice exposed to hyperoxia may contribute to aberrant lung development.

Tyr721 regulates specific binding of the CSF-1 receptor kinase insert to PI 3'-kinase SH2 domains: a model for SH2-mediated receptor-target interactions.

PubMed Central

Reedijk, M; Liu, X; van der Geer, P; Letwin, K; Waterfield, M D; Hunter, T; Pawson, T

1992-01-01

Efficient binding of active phosphatidylinositol (PI) 3'-kinase to the autophosphorylated macrophage colony stimulating factor receptor (CSF-1R) requires the noncatalytic kinase insert (KI) region of the receptor. To test whether this region could function independently to bind PI 3'-kinase, the isolated CSF-1R KI was expressed in Escherichia coli, and was inducibly phosphorylated on tyrosine. The tyrosine phosphorylated form of the CSF-1R KI bound PI 3'-kinase in vitro, whereas the unphosphorylated form had no binding activity. The p85 alpha subunit of PI 3'-kinase contains two Src homology (SH)2 domains, which are implicated in the interactions of signalling proteins with activated receptors. Bacterially expressed p85 alpha SH2 domains complexed in vitro with the tyrosine phosphorylated CSF-1R KI. Binding of the CSF-1R KI to PI 3'-kinase activity, and to the p85 alpha SH2 domains, required phosphorylation of Tyr721 within the KI domain, but was independent of phosphorylation at Tyr697 and Tyr706. Tyr721 was also critical for the association of activated CSF-1R with PI 3'-kinase in mammalian cells. Complex formation between the CSF-1R and PI 3'-kinase can therefore be reconstructed in vitro in a specific interaction involving the phosphorylated receptor KI and the SH2 domains of p85 alpha. Images PMID:1314163

Expression of cholinergic, insulin, vitamin D receptors and GLUT 3 in the brainstem of streptozotocin induced diabetic rats: effect of treatment with vitamin D₃.

PubMed

Peeyush Kumar, T; Paul, Jes; Antony, Sherin; Paulose, C S

2011-11-01

Complications arising from diabetes mellitus include cognitive deficits, neurophysiological and structural changes in the brain. The current study investigated the expression of cholinergic, insulin, Vitamin D receptor and GLUT 3 in the brainstem of streptozotocin-induced diabetic rats. Radioreceptor binding assays and gene expression were done in the brainstem of male Wistar rats. Our results showed that B(max) of total muscarinic, muscarinic M3 receptors was increased and muscarinic M1 receptor was decreased in diabetic rats compared to control. A significant increase in gene expression of muscarinic M3, α7 nicotinic acetylcholine, insulin, Vitamin D₃ receptors, acetylcholine esterase, choline acetyl transferase and GLUT 3 were observed in the brainstem of diabetic rats. Immunohistochemistry studies of muscarinic M1, M3 and α7 nicotinic acetylcholine receptors confirmed the gene expression at protein level. Vitamin D₃ and insulin treatment reversed diabetes-induced alterations to near control. This study provides an evidence that diabetes can alter the expression of cholinergic, insulin, Vitamin D receptors and GLUT 3 in brainstem. We found that Vitamin D₃ treatment could modulate the Vitamin D receptors and plays a pivotal role in maintaining the glucose transport and expressional level of cholinergic receptors in the brainstem of diabetic rats. Thus, our results suggest a therapeutic role of Vitamin D₃ in managing neurological disorders associated with diabetes.

Niclosamide inhibits epithelial-mesenchymal transition and tumor growth in lapatinib-resistant human epidermal growth factor receptor 2-positive breast cancer.

PubMed

Liu, Junjun; Chen, Xiaosong; Ward, Toby; Mao, Yan; Bockhorn, Jessica; Liu, Xiaofei; Wang, Gen; Pegram, Mark; Shen, Kunwei

2016-02-01

Acquired resistance to lapatinib, a human epidermal growth factor receptor 2 kinase inhibitor, remains a clinical problem for women with human epidermal growth factor receptor 2-positive advanced breast cancer, as metastasis is commonly observed in these patients. Niclosamide, an anti-helminthic agent, has recently been shown to exhibit cytotoxicity to tumor cells with stem-like characteristics. This study was designed to identify the mechanisms underlying lapatinib resistance and to determine whether niclosamide inhibits lapatinib resistance by reversing epithelial-mesenchymal transition. Here, two human epidermal growth factor receptor 2-positive breast cancer cell lines, SKBR3 and BT474, were exposed to increasing concentrations of lapatinib to establish lapatinib-resistant cultures. Lapatinib-resistant SKBR3 and BT474 cells exhibited up-regulation of the phenotypic epithelial-mesenchymal transition markers Snail, vimentin and α-smooth muscle actin, accompanied by activation of nuclear factor-кB and Src and a concomitant increase in stem cell marker expression (CD44(high)/CD24(low)), compared to naive lapatinib-sensitive SKBR3 and BT474 cells, respectively. Interestingly, niclosamide reversed epithelial-mesenchymal transition, induced apoptosis and inhibited cell growth by perturbing aberrant signaling pathway activation in lapatinib-resistant human epidermal growth factor receptor 2-positive cells. The ability of niclosamide to alleviate stem-like phenotype development and invasion was confirmed. Collectively, our results demonstrate that lapatinib resistance correlates with epithelial-mesenchymal transition and that niclosamide inhibits lapatinib-resistant cell viability and epithelial-mesenchymal transition. These findings suggest a role of niclosamide or derivatives optimized for more favorable bioavailability not only in reversing lapatinib resistance but also in reducing metastatic potential during the treatment of human epidermal growth factor receptor 2-positive breast cancer. Copyright © 2015 Elsevier Ltd. All rights reserved.

[99mTc] HYNIC-hEGF, a potential agent for imaging of EGF receptors in vivo: preparation and pre-clinical evaluation.

PubMed

Babaei, Mohammad Hossein; Almqvist, Ylva; Orlova, Anna; Shafii, Mohammad; Kairemo, Kalevi; Tolmachev, Vladimir

2005-06-01

Expression of epidermal growth factor receptors (EGFR) has prognostic and predictive value in many kinds of tumors. Imaging of expression of EGFR in vivo may give valuable diagnostic information. The epidermal growth factor (EGF), a natural ligand, is a possible candidate for the targeting of EGFR. The present study describes a method for preparation of (99m)Tc-EGF via the hydrazinopyridine-3-carboxylic acid (HYNIC) conjugation using tricine and ethylenediamine-N,N'-diacetic acid (EDDA) as co-ligands. Both conjugates bound EGFR expressing cells with nanomolar affinity, and demonstrated good intracellular retention. The complex with EDDA demonstrated much higher stability in blood serum and during cysteine challenge. Biodistribution of (99m)Tc-EDDA-HYNIC-EGF in normal mice demonstrated fast blood clearance of conjugate, and its ability to bind EGFR in vivo. (99m)Tc-EDDA-HYNIC-EGF is a promising candidate for visualization of EGFR expression in vivo.

Effect of acute and continuous morphine treatment on transcription factor expression in subregions of the rat caudate putamen. Marked modulation by D4 receptor activation.

PubMed

Gago, Belén; Suárez-Boomgaard, Diana; Fuxe, Kjell; Brené, Stefan; Reina-Sánchez, María Dolores; Rodríguez-Pérez, Luis M; Agnati, Luigi F; de la Calle, Adelaida; Rivera, Alicia

2011-08-17

Acute administration of the dopamine D(4) receptor (D(4)R) agonist PD168,077 induces a down-regulation of the μ opioid receptor (MOR) in the striosomal compartment of the rat caudate putamen (CPu), suggesting a striosomal D(4)R/MOR receptor interaction in line with their high co-distribution in this brain subregion. The present work was designed to explore if a D(4)R/MOR receptor interaction also occurs in the modulation of the expression pattern of several transcription factors in striatal subregions that play a central role in drug addiction. Thus, c-Fos, FosB/ΔFosB and P-CREB immunoreactive profiles were quantified in the rat CPu after either acute or continuous (6-day) administration of morphine and/or PD168,077. Acute and continuous administration of morphine induced different patterns of expression of these transcription factors, effects that were time-course and region dependent and fully blocked by PD168,077 co-administration. Moreover, this effect of the D(4)R agonist was counteracted by the D(4)R antagonist L745,870. Interestingly, at some time-points, combined treatment with morphine and PD168,077 substantially increased c-Fos, FosB/ΔFosB and P-CREB expression. The results of this study give indications for a general antagonistic D(4)R/MOR receptor interaction at the level of transcription factors. The change in the transcription factor expression by D(4)R/MOR interactions in turn suggests a modulation of neuronal activity in the CPu that could be of relevance for drug addiction. Copyright © 2011 Elsevier B.V. All rights reserved.

IRE1α links Nck1 deficiency to attenuated PTP1B expression in HepG2 cells.

PubMed

Li, Hui; Li, Bing; Larose, Louise

2017-08-01

PTP1B, a prototype of the non-receptor subfamily of the protein tyrosine phosphatase superfamily, plays a key role in regulating intracellular signaling from various receptor and non-receptor protein tyrosine kinases. Previously, we reported that silencing Nck1 in human hepatocellular carcinoma HepG2 cells enhances basal and growth factor-induced activation of the PI3K-Akt pathway through attenuating PTP1B expression. However, the underlying mechanism by which Nck1 depletion represses PTP1B expression remains unclear. In this study, we found that silencing Nck1 attenuates PTP1B expression in HepG2 cells through down-regulation of IRE1α. Indeed, we show that silencing Nck1 in HepG2 cells leads to decreased IRE1α expression and signaling. Accordingly, IRE1α depletion using siRNA in HepG2 cells enhances PI3K-dependent basal and growth factor-induced Akt activation, reproducing the effects of silencing Nck1 on activation of this pathway. In addition, depletion of IRE1α also leads to reduced PTP1B expression, which was rescued by ectopic expression of IRE1α in Nck1-depleted cells. Mechanistically, we found that silencing either Nck1 or IRE1α in HepG2 cells decreases PTP1B mRNA levels and stability. However, despite miR-122 levels, a miRNA targeting PTP1B 3' UTR and inducing PTP1B mRNA degradation in HepG2 cells, are increased in both Nck1- and IRE1α-depleted HepG2 cells, a miR-122 antagomir did not rescue PTP1B expression in these cells. Overall, this study highlights an important role for Nck1 in fine-tuning IRE1α expression and signaling that regulate PTP1B expression and subsequent activation of the PI3K-Akt pathway in HepG2 cells. Copyright © 2017 Elsevier Inc. All rights reserved.

Prolactin Signaling through the Short Form of Its Receptor Represses Forkhead Transcription Factor FOXO3 and Its Target Gene Galt Causing a Severe Ovarian Defect

PubMed Central

Halperin, Julia; Devi, Sangeeta Y.; Elizur, Shai; Stocco, Carlos; Shehu, Aurora; Rebourcet, Diane; Unterman, Terry G.; Leslie, Nancy D.; Le, Jamie; Binart, Nadine; Gibori, Geula

2008-01-01

Prolactin (PRL) is a hormone with over 300 biological activities. Although the signaling pathway downstream of the long form of its receptor (RL) has been well characterized, little is known about PRL actions upon activation of the short form (RS). Here, we show that mice expressing only RS exhibit an ovarian phenotype of accelerated follicular recruitment followed by massive follicular death leading to premature ovarian failure. Consequently, RS-expressing ovaries of young adults are depleted of functional follicles and formed mostly by interstitium. We also show that activation of RS represses the expression of the transcription factor Forkhead box O3 (FOXO3) and that of the enzyme galactose-1-phosphate uridyltransferase (Galt), two proteins known to be essential for normal follicular development. Our finding that FOXO3 regulates the expression of Galt and enhances its transcriptional activity indicates that it is the repression of FOXO3 by PRL acting through RS that prevents Galt expression in the ovary and causes follicular death. Coexpression of RL with RS prevents PRL inhibition of Galt, and the ovarian defect is no longer seen in RS transgenic mice that coexpress RL, suggesting that RL prevents RS-induced ovarian impairment. In summary, we show that PRL signals through RS and causes, in the absence of RL, a severe ovarian pathology by repressing the expression of FOXO3 and that of its target gene Galt. We also provide evidence of a link between the premature ovarian failure seen in mice expressing RS and in mice with FOXO3 gene deletion as well as in human with Galt mutation. PMID:17975019

Individual receptor profiling as a novel tool to support diagnosis of bladder pain syndrome/interstitial cystitis (BPS/IC).

PubMed

Neuhaus, Jochen; Schulte-Baukloh, Heinrich; Stolzenburg, Jens-Uwe; Speroni di Fenizio, Pietro; Horn, Lars-Christian; Rüffert, Henrik; Hartenstein, Siegurd; Burger, Maximilian; Schulze, Matthias; Schwalenberg, Thilo

2012-10-01

Dysregulation of neurotransmitter receptors may contribute to bladder overactivity (OAB) symptoms. To address the question whether specific receptor expression patterns are associated with bladder pain syndrome/interstitial cystitis (BPS/IC), we examined the expression of muscarinic, purinergic and histamine receptors in the detrusor. Detrusor receptor expression was investigated in bladder biopsies of female BPS/IC patients (n = 44; age 60.64 ± 13.78, mean ± SD) and carcinoma patients (n = 11; age 58.91 ± 12.72) undergoing cystectomy. Protein expression of muscarinic (M2, M3), purinergic (P2X1-3) and histamine receptors (H1, H2) was analysed by confocal immunofluorescence, and gene expression was quantified by real-time polymerase chain reaction (qPCR). M2, P2X1, P2X2 and H1 receptor immunoreactivity (-IR) was significantly enhanced in BPS/IC compared to the control group, while there was no difference for M3-, P2X3- and H2-IR. We calculated a score, which separated BPS/IC from control patients with an AUC of 89.46%, showing 84.09% sensitivity and 90.91% specificity. Patients had a 9.25 times enhanced calculated risk for BPS/IC. In addition, two patient subgroups (M2 > M3 and M3 > M2) were observed, which differed in associated purinergic and histamine receptor expression. M2, P2X1, P2X2 and H1 were significantly upregulated in BPS/IC patients, and H2 was occasionally highly overexpressed. There was no significant correlation between receptor protein and gene expression, implying posttranslational mechanisms being responsible for the altered receptor expressions. On the basis of individual receptor profiles, upregulated receptors could be targeted by monotherapy or combination therapy with already approved receptor inhibitors, thereby promoting tailored therapy for patients suffering from BPS/IC-like symptoms.

A screen for transcription factor targets of glycogen synthase kinase-3 highlights an inverse correlation of NFκB and androgen receptor signaling in prostate cancer.

PubMed

Campa, Victor M; Baltziskueta, Eder; Bengoa-Vergniory, Nora; Gorroño-Etxebarria, Irantzu; Wesołowski, Radosław; Waxman, Jonathan; Kypta, Robert M

2014-09-30

Expression of Glycogen Synthase Kinase-3 (GSK-3) is elevated in prostate cancer and its inhibition reduces prostate cancer cell proliferation, in part by reducing androgen receptor (AR) signaling. However, GSK-3 inhibition can also activate signals that promote cell proliferation and survival, which may preclude the use of GSK-3 inhibitors in the clinic. To identify such signals in prostate cancer, we screened for changes in transcription factor target DNA binding activity in GSK-3-silenced cells. Among the alterations was a reduction in AR DNA target binding, as predicted from previous studies, and an increase in NFκB DNA target binding. Consistent with the latter, gene silencing of GSK-3 or inhibition using the GSK-3 inhibitor CHIR99021 increased basal NFκB transcriptional activity. Activation of NFκB was accompanied by an increase in the level of the NFκB family member RelB. Conversely, silencing RelB reduced activation of NFκB by CHIR99021. Furthermore, the reduction of prostate cancer cell proliferation by CHIR99021 was potentiated by inhibition of NFκB signaling using the IKK inhibitor PS1145. Finally, stratification of human prostate tumor gene expression data for GSK3 revealed an inverse correlation between NFκB-dependent and androgen-dependent gene expression, consistent with the results from the transcription factor target DNA binding screen. In addition, there was a correlation between expression of androgen-repressed NFκB target genes and reduced survival of patients with metastatic prostate cancer. These findings highlight an association between GSK-3/AR and NFκB signaling and its potential clinical importance in metastatic prostate cancer.

A screen for transcription factor targets of Glycogen Synthase Kinase-3 highlights an inverse correlation of NFκB and Androgen Receptor Signaling in Prostate Cancer

PubMed Central

Campa, Victor M.; Baltziskueta, Eder; Bengoa-Vergniory, Nora; Gorroño-Etxebarria, Irantzu; Wesołowski, Radosław; Waxman, Jonathan; Kypta, Robert M.

2014-01-01

Expression of Glycogen Synthase Kinase-3 (GSK-3) is elevated in prostate cancer and its inhibition reduces prostate cancer cell proliferation, in part by reducing androgen receptor (AR) signaling. However, GSK-3 inhibition can also activate signals that promote cell proliferation and survival, which may preclude the use of GSK-3 inhibitors in the clinic. To identify such signals in prostate cancer, we screened for changes in transcription factor target DNA binding activity in GSK-3-silenced cells. Among the alterations was a reduction in AR DNA target binding, as predicted from previous studies, and an increase in NFκB DNA target binding. Consistent with the latter, gene silencing of GSK-3 or inhibition using the GSK-3 inhibitor CHIR99021 increased basal NFκB transcriptional activity. Activation of NFκB was accompanied by an increase in the level of the NFκB family member RelB. Conversely, silencing RelB reduced activation of NFκB by CHIR99021. Furthermore, the reduction of prostate cancer cell proliferation by CHIR99021 was potentiated by inhibition of NFκB signaling using the IKK inhibitor PS1145. Finally, stratification of human prostate tumor gene expression data for GSK3 revealed an inverse correlation between NFκB-dependent and androgen-dependent gene expression, consistent with the results from the transcription factor target DNA binding screen. In addition, there was a correlation between expression of androgen-repressed NFκB target genes and reduced survival of patients with metastatic prostate cancer. These findings highlight an association between GSK-3/AR and NFκB signaling and its potential clinical importance in metastatic prostate cancer. PMID:25327559

Contribution of vascular endothelial growth factor receptor-2 sialylation to the process of angiogenesis.

PubMed

Chiodelli, P; Rezzola, S; Urbinati, C; Federici Signori, F; Monti, E; Ronca, R; Presta, M; Rusnati, M

2017-11-23

Vascular endothelial growth factor receptor-2 (VEGFR2) is the main pro-angiogenic receptor expressed by endothelial cells (ECs). Using surface plasmon resonance, immunoprecipitation, enzymatic digestion, immunofluorescence and cross-linking experiments with specific sugar-binding lectins, we demonstrated that VEGFR2 bears both α,1-fucose and α(2,6)-linked sialic acid (NeuAc). However, only the latter is required for VEGF binding to VEGFR2 and consequent VEGF-dependent VEGFR2 activation and motogenic response in ECs. Notably, downregulation of β-galactoside α(2,6)-sialyltransferase expression by short hairpin RNA transduction inhibits VEGFR2 α(2,6) sialylation that is paralleled by an increase of β-galactoside α(2,3)-sialyltransferase expression. This results in an ex-novo α(2,3)-NeuAc sialylation of the receptor that functionally replaces the lacking α(2,6)-NeuAc, thus allowing VEGF/VEGFR2 interaction. In keeping with the role of VEGFR2 sialylation in angiogenesis, the α(2,6)-NeuAc-binding lectin Sambucus nigra (SNA) prevents VEGF-dependent VEGFR2 autophosphorylation and EC motility, proliferation and motogenesis. In addition, SNA exerts a VEGF-antagonist activity in tridimensional angiogenesis models in vitro and in the chick-embryo chorioallantoic membrane neovascularization assay and mouse matrigel plug assay in vivo. In conclusion, VEGFR2-associated NeuAc plays an important role in modulating VEGF/VEGFR2 interaction, EC pro-angiogenic activation and neovessel formation. VEGFR2 sialylation may represent a target for the treatment of angiogenesis-dependent diseases.

Dehydroepiandrosterone Activation of G-protein-coupled Estrogen Receptor Rapidly Stimulates MicroRNA-21 Transcription in Human Hepatocellular Carcinoma Cells*

PubMed Central

Teng, Yun; Radde, Brandie N.; Litchfield, Lacey M.; Ivanova, Margarita M.; Prough, Russell A.; Clark, Barbara J.; Doll, Mark A.; Hein, David W.; Klinge, Carolyn M.

2015-01-01

Little is known about the regulation of the oncomiR miR-21 in liver. Dehydroepiandrosterone (DHEA) regulates gene expression as a ligand for a G-protein-coupled receptor and as a precursor for steroids that activate nuclear receptor signaling. We report that 10 nm DHEA increases primary miR-21 (pri-miR-21) transcription and mature miR-21 expression in HepG2 cells in a biphasic manner with an initial peak at 1 h followed by a second, sustained response from 3–12 h. DHEA also increased miR-21 in primary human hepatocytes and Hep3B cells. siRNA, antibody, and inhibitor studies suggest that the rapid DHEA-mediated increase in miR-21 involves a G-protein-coupled estrogen receptor (GPER/GPR30), estrogen receptor α-36 (ERα36), epidermal growth factor receptor-dependent, pertussis toxin-sensitive pathway requiring activation of c-Src, ERK1/2, and PI3K. GPER antagonist G-15 attenuated DHEA- and BSA-conjugated DHEA-stimulated pri-miR-21 transcription. Like DHEA, GPER agonists G-1 and fulvestrant increased pri-miR-21 in a GPER- and ERα36-dependent manner. DHEA, like G-1, increased GPER and ERα36 mRNA and protein levels. DHEA increased ERK1/2 and c-Src phosphorylation in a GPER-responsive manner. DHEA increased c-Jun, but not c-Fos, protein expression after 2 h. DHEA increased androgen receptor, c-Fos, and c-Jun recruitment to the miR-21 promoter. These results suggest that physiological concentrations of DHEA activate a GPER intracellular signaling cascade that increases pri-miR-21 transcription mediated at least in part by AP-1 and androgen receptor miR-21 promoter interaction. PMID:25969534

The distribution of the orphan bombesin receptor subtype-3 in the rat CNS.

PubMed

Jennings, C A; Harrison, D C; Maycox, P R; Crook, B; Smart, D; Hervieu, G J

2003-01-01

Bombesin receptor subtype 3 (BRS-3) is an orphan G-protein coupled receptor that shares between 47 and 51% homology with other known bombesin receptors. The natural ligand for BRS-3 is currently unknown and little is known about the mechanisms regulating BRS-3 gene expression. Unlike other mammalian bombesin receptors that have been shown to be predominantly expressed in the CNS and gastrointestinal tract, expression of the BRS-3 receptor in the rat brain has previously not been observed. To gain further understanding of the biology of BRS-3, we have studied the distribution of BRS-3 mRNA and protein in the rat CNS. The mRNA expression pattern was studied using reverse transcription followed by quantitative polymerase chain reaction. Using immunohistological techniques, the distribution of BRS-3 protein in the rat brain was investigated using a rabbit affinity-purified polyclonal antiserum raised against an N-terminal peptide. The BRS-3 receptor was found to be widely expressed in the rat brain at both mRNA and protein levels. Particularly strong immunosignals were observed in the cerebral cortex, hippocampal formation, hypothalamus and thalamus. Other regions of the brain such as the basal ganglia, midbrain and reticular formation were also immunopositive for BRS-3. In conclusion, our neuroanatomical data provide evidence that BRS-3 is as widely expressed in the rat brain as other bombesin-like peptide receptors and suggest that this receptor may also have important roles in the CNS, mediating the functions of a so far unidentified ligand.

Subchronic treatment with fluoxetine and ketanserin increases hippocampal brain-derived neurotrophic factor, β-catenin and antidepressant-like effects.

PubMed

Pilar-Cuéllar, F; Vidal, R; Pazos, A

2012-02-01

5-HT(2A) receptor antagonists improve antidepressant responses when added to 5-HT-selective reuptake inhibitors (SSRIs) or tricyclic antidepressants. Here, we have studied the involvement of neuroplasticity pathways and/or the 5-hydroxytryptaminergic system in the antidepressant-like effect of this combined treatment, given subchronically. Expression of brain-derived neurotrophic factor (BDNF) and its receptor (TrkB), 5-bromo-2'-deoxyuridine (BrdU) incorporation, and β-catenin protein expression in different cellular fractions, as well as 5-HT(1A) receptor function were measured in the hippocampus of rats treated with fluoxetine, ketanserin and fluoxetine + ketanserin for 7 days, followed by a forced swimming test (FST) to analyse antidepressant efficacy. mRNA for BDNF was increased in the CA3 field and dentate gyrus of the hippocampus by combined treatment with fluoxetine + ketanserin. Expression of β-catenin was increased in total hippocampal homogenate and in the membrane fraction, but unchanged in the nuclear fraction after combined treatment with fluoxetine + ketanserin. These effects were paralleled by a decreased immobility time in the FST. There were no changes in BrdU incorporation, TrkB expression and 5-HT(1A) receptor function in any of the groups studied. The antidepressant-like effect induced by subchronic co-treatment with a SSRI and a 5-HT(2A) receptor antagonist may mainly be because of modifications in hippocampal neuroplasticity (BDNF and membrane-associated β-catenin), without a significant role for other mechanisms involved in chronic antidepressant response, such as hippocampal neuroproliferation or 5-HT(1A) receptor desensitization in the dorsal raphe nucleus. © 2011 The Authors. British Journal of Pharmacology © 2011 The British Pharmacological Society.

DcR3 induces epithelial-mesenchymal transition through activation of the TGF-β3/SMAD signaling pathway in CRC

PubMed Central

Hu, Zhi-Yan; Li, Sheng-Nan; Kan, He-Ping; Wang, Xiao-Yan; Li, Zu-Guo

2016-01-01

Decoy receptor 3 (DcR3), a novel member of the tumor necrosis factor receptor (TNFR) family, was recently reported to be associated with tumorigenesis and metastasis. However, the role of DcR3 in human colorectal cancer (CRC) has not been fully elucidated. In this study, we found that DcR3 expression was significantly higher in human colorectal cancer tissues than in paired normal tissues, and that DcR3 expression was strongly correlated with tumor invasion, lymph node metastases and poor prognoses. Moreover, DcR3 overexpression significantly enhanced CRC cell proliferation and migration in vitro and tumorigenesis in vivo. Conversely, DcR3 knockdown significantly repressed CRC cell proliferation and migration in vitro, and DcR3 deficiency also attenuated CRC tumorigenesis and metastasis in vivo. Functionally, DcR3 was essential for TGF-β3/SMAD-mediated epithelial-mesenchymal transition (EMT) of CRC cells. Importantly, cooperation between DcR3 and TGF-β3/SMAD-EMT signaling-related protein expression was correlated with survival and survival time in CRC patients. In conclusion, our results demonstrate that DcR3 may be a prognostic biomarker for CRC and that this receptor facilitates CRC development and metastasis by participating in TGF-β3/SMAD-mediated EMT of CRC cells. PMID:27764793

The expression of Fc and complement receptors in young, adult and aged mice.

PubMed Central

Vĕtvicka, V; Fornůsek, L; Zídková, J

1985-01-01

Age-dependent changes in the expression of Fc receptors (FcR) for different isotypes of immunoglobulins and receptors for C3b, C5b and C3bi fragments of complement on the membranes of peritoneal macrophages were studied with mice of different ages. An age-related increase in expression of Fc receptors for IgM, IgE, IgA, IgG2b and IgG3, and a decrease in the expression of Fc receptors for IgG1 was observed. The expression of FcR on macrophages of donors of different ages corresponded with Fc-receptor mediated phagocytosis. The highest number of C3b-binding macrophages was found in aged mice, in contrast to low numbers of C3bi-binding macrophages at this age. The percentage of C5b-binding macrophages was lowest in adult animals. We also observed effective inhibition of binding of the C3b component of complement by preincubation of macrophages with aggregated IgG and vice versa. These observations suggest that fluctuation in expression of Fc but not C receptors may be important to the generalized changes that occur in macrophage function during development and ageing. PMID:2931351

Requirement of Vascular Integrin α_vβ_3 for Angiogenesis

NASA Astrophysics Data System (ADS)

Brooks, Peter C.; Clark, Richard A. F.; Cheresh, David A.

1994-04-01

Angiogenesis depends on the adhesive interactions of vascular cells. The adhesion receptor integrin α_vβ_3 was identified as a marker of angiogenic vascular tissue. Integrin α_vβ_3 was expressed on blood vessels in human wound granulation tissue but not in normal skin, and it showed a fourfold increase in expression during angiogenesis on the chick chorioallantoic membrane. In the latter assay, a monoclonal antibody to α_vβ_3 blocked angiogenesis induced by basic fibroblast growth factor, tumor necrosis factor-α, and human melanoma fragments but had no effect on preexisting vessels. These findings suggest that α_vβ_3 may be a useful therapeutic target for diseases characterized by neovascularization.

The GDNF System Is Altered in Diverticular Disease – Implications for Pathogenesis

PubMed Central

Böttner, Martina; Barrenschee, Martina; Hellwig, Ines; Harde, Jonas; Egberts, Jan-Hendrik; Becker, Thomas; Zorenkov, Dimitri; Schäfer, Karl-Herbert; Wedel, Thilo

2013-01-01

Background & Aims Absence of glial cell line-derived neurotrophic factor (GDNF) leads to intestinal aganglionosis. We recently demonstrated that patients with diverticular disease (DD) exhibit hypoganglionosis suggesting neurotrophic factor deprivation. Thus, we screened mRNA expression pattern of the GDNF system in DD and examined the effects of GDNF on cultured enteric neurons. Methods Colonic specimens obtained from patients with DD (n = 21) and controls (n = 20) were assessed for mRNA expression levels of the GDNF system (GDNF, GDNF receptors GFRα1 and RET). To identify the tissue source of GDNF and its receptors, laser-microdissected (LMD) samples of human myenteric ganglia and intestinal muscle layers were analyzed separately by qPCR. Furthermore, the effects of GDNF treatment on cultured enteric neurons (receptor expression, neuronal differentiation and plasticity) were monitored. Results mRNA expression of GDNF and its receptors was significantly down-regulated in the muscularis propria of patients with DD. LMD samples revealed high expression of GDNF in circular and longitudinal muscle layers, whereas GDNF receptors were also expressed in myenteric ganglia. GDNF treatment of cultured enteric neurons increased mRNA expression of its receptors and promoted neuronal differentiation and plasticity revealed by synaptophysin mRNA and protein expression. Conclusions Our results suggest that the GDNF system is compromised in DD. In vitro studies demonstrate that GDNF enhances expression of its receptors and promotes enteric neuronal differentiation and plasticity. Since patients with DD exhibit hypoganglionosis, we propose that the observed enteric neuronal loss in DD may be due to lacking neurotrophic support mediated by the GDNF system. PMID:23805210

Roles of cell-cell adhesion-dependent tyrosine phosphorylation of Gab-1.

PubMed

Shinohara, M; Kodama, A; Matozaki, T; Fukuhara, A; Tachibana, K; Nakanishi, H; Takai, Y

2001-06-01

Gab-1 is a multiple docking protein that is tyrosine phosphorylated by receptor tyrosine kinases such as c-Met, hepatocyte growth factor/scatter factor receptor, and epidermal growth factor receptor. We have now demonstrated that cell-cell adhesion also induces marked tyrosine phosphorylation of Gab-1 and that disruption of cell-cell adhesion results in its dephosphorylation. An anti-E-cadherin antibody decreased cell-cell adhesion-dependent tyrosine phosphorylation of Gab-1, whereas the expression of E-cadherin specifically induced tyrosine phosphorylation of Gab-1. A relatively selective inhibitor of Src family kinases reduced cell-cell adhesion-dependent tyrosine phosphorylation of Gab-1, whereas expression of a dominant-negative mutant of Csk increased it. Disruption of cell-cell adhesion, which reduced tyrosine phosphorylation of Gab-1, also reduced the activation of mitogen-activated protein kinase and Akt in response to cell-cell adhesion. These results indicate that E-cadherin-mediated cell-cell adhesion induces tyrosine phosphorylation by a Src family kinase of Gab-1, thereby regulating the activation of Ras/MAP kinase and phosphatidylinositol 3-kinase/Akt cascades.

Regulation of the ovarian inflammatory response at ovulation by nuclear progesterone receptor.

PubMed

Akison, Lisa K; Robertson, Sarah A; Gonzalez, Macarena B; Richards, JoAnne S; Smith, C Wayne; Russell, Darryl L; Robker, Rebecca L

2018-06-01

The nuclear progesterone receptor (PGR) transcription factor is essential for ovulation; however, the exact mechanisms by which PGR controls ovulation are not known. The aim of this study was to determine whether PGR regulates inflammatory mediators in the ovary. Ovaries from mice lacking PGR (PRKO) and heterozygous PR+/- littermates were subjected to microarray analysis of a large panel of inflammatory genes. Immune cell subsets were detected by gene expression; and neutrophils by immunohistochemistry and chemotaxis assay. PRKO ovaries exhibited dysregulated expression of vasodilator (Edn1), cytokine (Il-6, Tgfb1), adhesion receptor (Cd34), apoptotic factor (Bax) and transcription factors (Nfkb2, Socs1, Stat3). Ptgs2 was also reduced in PRKO ovaries, but mRNA and protein were not different in granulosa cells. There were reduced neutrophils in ovaries of PRKO mice at ovulation; however, chemotaxis assays showed PRKO neutrophils migrate normally and that PRKO ovarian extracts exhibit chemotactic properties in vitro. Specific inflammatory mediators are altered in the ovaries of PRKO mice indicating that progesterone regulates features of inflammation at ovulation. © 2018 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Tendencies for higher co-expression of EGFR and HER2 and downregulation of HER3 in prostate cancer lymph node metastases compared with corresponding primary tumors.

PubMed

Carlsson, J; Shen, L; Xiang, J; Xu, J; Wei, Q

2013-01-01

The epidermal growth factor receptor (EGFR) family members are potential targets for therapy using extra-cellular domain receptor binding agents, such as the antibodies trastuzumab and cetuximab, or antibodies labeled with therapeutically useful radionuclides or toxins. This is especially the case when the tumor cells are resistant to chemotherapy and tyrosine kinase inhibitors. Studies concerning the expression of these receptors in prostate cancer vary in the literature, possibly due to differences in patient inclusion, sample preparations and scoring criteria. In our study, EGFR, HER2 and HER3 expression was analyzed in prostate cancer samples from primary tumors and corresponding lymph node metastases from 12 patients. The expression of HER2 and EGFR was scored from immunohistochemical preparations and the HercepTest criteria (0, 1+, 2+ or 3+), while HER3 expression was scored as no, weak or strong staining. There were 5 EGFR-positive (2+ or 3+) primary tumors and 6 EGFR-positive lymph node metastases, and there was EGFR upregulation in one metastasis. Only 4 of the 12 patients had marked HER2 expression (2+ or 3+) in their primary tumors and there was one downregulation and 5 cases of upregulation in the metastases. Thus, a total of 8 out of 12 analyzed metastases were HER2-positive. Of the 12 primary tumors, 9 expressed HER3 while only 2 of the lymph node metastases expressed recognizable HER3 staining, so 7 metastases appeared to have downregulated HER3 expression. In one of the primary tumors there was positive co-expression of EGFR and HER2, while this co-expression was observed in 4 of the metastases. Thus, there were tendencies for upregulation of HER2, increased co-expression of EGFR and HER2 and downregulation of HER3 in the prostate cancer lymph node metastases in comparison to the primary tumors. The results are encouraging for studies involving more patients. Possible strategies for EGFR- and HER2-targeted therapy are briefly discussed in the present study, especially with regard to the expression and co-expression of EGFR and HER2 in metastases.

Gab3-deficient mice exhibit normal development and hematopoiesis and are immunocompetent.

PubMed

Seiffert, Martina; Custodio, Joseph M; Wolf, Ingrid; Harkey, Michael; Liu, Yan; Blattman, Joseph N; Greenberg, Philip D; Rohrschneider, Larry R

2003-04-01

Gab proteins are intracellular scaffolding and docking molecules involved in signaling pathways mediated by various growth factor, cytokine, or antigen receptors. Gab3 has been shown to act downstream of the macrophage colony-stimulating factor receptor, c-Fms, and to be important for macrophage differentiation. To analyze the physiological role of Gab3, we used homologous recombination to generate mice deficient in Gab3. Gab3(-/-) mice develop normally, are visually indistinguishable from their wild-type littermates, and are healthy and fertile. To obtain a detailed expression pattern of Gab3, we generated Gab3-specific monoclonal antibodies. Immunoblotting revealed a predominant expression of Gab3 in lymphocytes and bone marrow-derived macrophages. However, detailed analysis demonstrated that hematopoiesis in mice lacking Gab3 is not impaired and that macrophages develop in normal numbers and exhibit normal function. The lack of Gab3 expression during macrophage differentiation is not compensated for by increased levels of Gab1 or Gab2 mRNA. Furthermore, Gab3-deficient mice have no major immune deficiency in T- and B-lymphocyte responses to protein antigens or during viral infection. In addition, allergic responses in Gab3-deficient mice appeared to be normal. Together, these data demonstrate that loss of Gab3 does not result in detectable defects in normal mouse development, hematopoiesis, or immune system function.

Biochemical and mass spectrometric characterization of the human CB2 cannabinoid receptor expressed in Pichia pastoris--importance of correct processing of the N-terminus.

PubMed

Zhang, Rundong; Kim, Tae-Kang; Qiao, Zhaun-Hong; Cai, Jian; Pierce, William M; Song, Zhao-Hui

2007-10-01

This study was conducted to optimize the expression of human CB2 cannabinoid receptors in methylotrophic yeast Pichia pastoris (P. pastoris). Two major species of expressed CB2 proteins were seen on Western blot, i.e., a 42 kDa band which matches the calculated molecular weight for tagged CB2, and a 52/55 kDa doublet. Treatment of membranes with N-glycosidase F or inclusion of tunicamycin in the culture medium during induction resulted in the disappearance of the 55 kDa, but not the 52 kDa band, suggesting that the 3 kDa extra in the 55 kDa band is due to N-glycosylation, but the 10 kDa extra in the 52 kDa band is not due to N-glycosylation. Anti-FLAG M1 antibody had a much higher preference for the 42 kDa band over the 52/55 kDa doublet, and a 10 kDa fragment recognized by anti-FLAG M2 antibody was generated by CNBr digestion of the 52/55 doublet. These data strongly support the hypothesis that the 10 kDa increase in molecular weight was due to unprocessed alpha-factor sequence. This conclusion was further validated by finding several peptide sequences for alpha-factor fragments at the N-terminal of the CB2 receptor using pepsin/chymotrypsin digestion and LC/MS/MS approaches. Importantly, unprocessed alpha-factor was found to be associated with poor ligand binding. In addition, controlling the level of CB2 protein expression was found to be critical for minimizing the presence of unprocessed alpha-factor sequence. The information gained from this study should aid the proper expression of not only CB2 receptor but also other members of the GPCR family in P. pastoris.

Epidermal growth factor increases LRF/Pokemon expression in human prostate cancer cells.

PubMed

Aggarwal, Himanshu; Aggarwal, Anshu; Agrawal, Devendra K

2011-10-01

Leukemia/lymphoma related factor/POK erythroid myeloid ontogenic factor (LRF/Pokemon) is a member of the POK family of proteins that promotes oncogenesis in several forms of cancer. Recently, we found higher LRF expression in human breast and prostate carcinomas compared to the corresponding normal tissues. The aim of this study was to examine the regulation of LRF expression in human prostate cells. Epidermal growth factor (EGF) and its receptors mediate several tumorigenic cascades that regulate cell differentiation, proliferation, migration and survival of prostate cancer cells. There was significantly higher level of LRF expression in the nucleus of LNCaP and PC-3 cells than RWPE-1 cells. A significant increase in LRF expression was observed with increasing doses of EGF in more aggressive and androgen-sensitive prostate cancer cells suggesting that EGF signaling pathway is critical in upregulating the expression of LRF/Pokemon to promote oncogenesis. Copyright © 2011 Elsevier Inc. All rights reserved.

Epidermal Growth Factor Increases LRF/Pokemon Expression in Human Prostate Cancer Cells

PubMed Central

Aggarwal, Himanshu; Aggarwal, Anshu; Agrawal, Devendra K.

2011-01-01

Leukemia/lymphoma related factor/POK erythroid myeloid ontogenic factor (LRF/Pokemon) is a member of the POK family of proteins that promotes oncogenesis in several forms of cancer. Recently, we found higher LRF expression in human breast and prostate carcinomas compared to the corresponding normal tissues. The aim of this study was to examine the regulation of LRF expression in human prostate cells. Epidermal growth factor (EGF) and its receptors mediate several tumorigenic cascades that regulate cell differentiation, proliferation, migration and survival of prostate cancer cells. There was significantly higher level of LRF expression in the nucleus of LNCaP and PC-3 cells than RWPE-1 cells. A significant increase in LRF expression was observed with increasing doses of EGF in more aggressive and androgen-sensitive prostate cancer cells suggesting that EGF signaling pathway is critical in upregulating the expression of LRF/Pokemon to promote oncogenesis. PMID:21640721

Biological responses to PDGF-BB versus PDGF-DD in human mesangial cells.

PubMed

van Roeyen, C R C; Ostendorf, T; Denecke, B; Bokemeyer, D; Behrmann, I; Strutz, F; Lichenstein, H S; LaRochelle, W J; Pena, C E; Chaudhuri, A; Floege, J

2006-04-01

Platelet-derived growth factor (PDGF)-BB and PDGF-DD mediate mesangial cell proliferation in vitro and in vivo. While PDGF-BB is a ligand for the PDGF alpha- and beta-receptor chains, PDGF-DD binds more selectively to the beta-chain, suggesting potential differences in the biological activities. Signal transduction and regulation of gene expression induced by PDGF-BB and -DD were compared in primary human mesangial cells (HMCs), which expressed PDGF alpha- and beta-receptor subunits. The growth factor concentrations used were chosen based on their equipotency in inducing HMCs proliferation and binding to the betabeta-receptor. Both growth factors, albeit at different concentrations induced phosphorylation and activation of extracellular signal-regulated kinase 1 (ERK1) and ERK2. In addition, PDGFs led to the phosphorylation and activation of signal transducers and activators of transcription 1 (STAT1) and STAT3. HMCs proliferation induced by either PDGF-BB or -DD could be blocked by signal transduction inhibitors of the mitogen-activated protein kinase-, Janus kinase (JAK)/STAT-, or phosphatidyl-inositol 3-kinase pathways. Using a gene chip array and subsequent verification by real-time reverse transcriptase (RT)-polymerase chain reaction, we found that in HMC genes for matrix metalloproteinase 13 (MMP-13) and MMP-14 and, to a low extent, cytochrome B5 and cathepsin L were exclusively regulated by PDGF-BB, whereas no exclusive gene regulation was detected by PDGF-DD. However, at the protein level, both MMP-13 and -14 were equally induced by PDGF-BB and -DD. PDGF-BB and -DD effect similar biological responses in HMCs albeit at different potencies. Rare apparently differential gene regulation did not result in different protein expression, suggesting that in HMCs both PDGFs exert their biological activity almost exclusively via the PDGF beta-receptor.

Functional Expression of Two Neuronal Nicotinic Acetylcholine Receptors from cDNA Clones Identifies a Gene Family

NASA Astrophysics Data System (ADS)

Boulter, Jim; Connolly, John; Deneris, Evan; Goldman, Dan; Heinemann, Steven; Patrick, Jim

1987-11-01

A family of genes coding for proteins homologous to the α subunit of the muscle nicotinic acetylcholine receptor has been identified in the rat genome. These genes are transcribed in the central and peripheral nervous systems in areas known to contain functional nicotinic receptors. In this paper, we demonstrate that three of these genes, which we call alpha3, alpha4, and beta2, encode proteins that form functional nicotinic acetylcholine receptors when expressed in Xenopus oocytes. Oocytes expressing either alpha3 or alpha4 protein in combination with the beta2 protein produced a strong response to acetylcholine. Oocytes expressing only the alpha4 protein gave a weak response to acetylcholine. These receptors are activated by acetylcholine and nicotine and are blocked by Bungarus toxin 3.1. They are not blocked by α -bungarotoxin, which blocks the muscle nicotinic acetylcholine receptor. Thus, the receptors formed by the alpha3, alpha4, and beta2 subunits are pharmacologically similar to the ganglionic-type neuronal nicotinic acetylcholine receptor. These results indicate that the alpha3, alpha4, and beta2 genes encode functional nicotinic acetylcholine receptor subunits that are expressed in the brain and peripheral nervous system.

Identification of embryonic precursor cells that differentiate into thymic epithelial cells expressing autoimmune regulator

PubMed Central

Takizawa, Nobukazu; Miyauchi, Maki; Yanai, Hiromi; Tateishi, Ryosuke; Shinzawa, Miho; Yoshinaga, Riko; Kurihara, Masaaki; Yasuda, Hisataka; Sakamoto, Reiko; Yoshida, Nobuaki

2016-01-01

Medullary thymic epithelial cells (mTECs) expressing autoimmune regulator (Aire) are critical for preventing the onset of autoimmunity. However, the differentiation program of Aire-expressing mTECs (Aire+ mTECs) is unclear. Here, we describe novel embryonic precursors of Aire+ mTECs. We found the candidate precursors of Aire+ mTECs (pMECs) by monitoring the expression of receptor activator of nuclear factor-κB (RANK), which is required for Aire+ mTEC differentiation. pMECs unexpectedly expressed cortical TEC molecules in addition to the mTEC markers UEA-1 ligand and RANK and differentiated into mTECs in reaggregation thymic organ culture. Introduction of pMECs in the embryonic thymus permitted long-term maintenance of Aire+ mTECs and efficiently suppressed the onset of autoimmunity induced by Aire+ mTEC deficiency. Mechanistically, pMECs differentiated into Aire+ mTECs by tumor necrosis factor receptor-associated factor 6-dependent RANK signaling. Moreover, nonclassical nuclear factor-κB activation triggered by RANK and lymphotoxin-β receptor signaling promoted pMEC induction from progenitors exhibiting lower RANK expression and higher CD24 expression. Thus, our findings identified two novel stages in the differentiation program of Aire+ mTECs. PMID:27401343

Growth Factor Receptor-Directed Therapy in Human Breast Cancer

DTIC Science & Technology

1997-12-01

related more to acquired rather than to intrinsic drug resistance. 3) To define the role of HER-2 and heregulin gene expression in antiestrogen... treatment strategies in affected patients. 3) Role of HER-2 and heregulin gene expression in antiestrogen resistance. The hypothesis that heregulins may be a...native amplification/overexpression of the HER-2/neu gene and sion. Finally, to avoid the possibility that any observed are shown as positive controls

Overexpression of androgen receptor and forkhead-box A1 protein in apocrine breast carcinoma.

PubMed

Sasahara, Manami; Matsui, Akira; Ichimura, Yoshiko; Hirakata, Yuuko; Murata, Yuuya; Marui, Eiji

2014-03-01

Apocrine breast carcinoma often lacks estrogen receptor (ER), progesterone receptor (PgR), and human epidermal growth factor receptor type-2 (HER2) expression. Accordingly, development of a new treatment strategy is important for this type of cancer. The growth stimulus through the androgen receptor (AR) can be a candidate for targeted treatment. Therefore, we examined the factors related to AR transcription. We immunohistochemically evaluated 54 apocrine cancer lesions for ER, PgR, AR, HER2, Ki-67, forkhead-box protein A1 (FOXA1), and prostate-specific antigen (PSA) expression. ER, PgR, and HER2 were expressed at a low level, thus 44 out of 54 (81.4%) cases were of triple-negative breast cancer. AR, PSA and FOXA1 were expressed in 100% (54/54), 48% (26/54) and 93% (50/54) of cases, respectively. Most of apocrine breast carcinomas were immunohistochemically-positive for AR and FOXA1. Anti-androgenic therapies can potentially serve as a cancer-targeting therapy for apocrine breast carcinoma.

Differential expression of VEGF ligands and receptors in prostate cancer.

PubMed

Woollard, David J; Opeskin, Kenneth; Coso, Sanja; Wu, Di; Baldwin, Megan E; Williams, Elizabeth D

2013-05-01

Prostate cancer disseminates to regional lymph nodes, however the molecular mechanisms responsible for lymph node metastasis are poorly understood. The vascular endothelial growth factor (VEGF) ligand and receptor family have been implicated in the growth and spread of prostate cancer via activation of the blood vasculature and lymphatic systems. The purpose of this study was to comprehensively examine the expression pattern of VEGF ligands and receptors in the glandular epithelium, stroma, lymphatic vasculature and blood vessels in prostate cancer. The localization of VEGF-A, VEGF-C, VEGF-D, VEGF receptor (VEGFR)-1, VEGFR-2, and VEGFR-3 was examined in cancerous and adjacent benign prostate tissue from 52 subjects representing various grades of prostate cancer. Except for VEGFR-2, extensive staining was observed for all ligands and receptors in the prostate specimens. In epithelial cells, VEGF-A and VEGFR-1 expression was higher in tumor tissue compared to benign tissue. VEGF-D and VEGFR-3 expression was significantly higher in benign tissue compared to tumor in the stroma and the endothelium of lymphatic and blood vessels. In addition, the frequency of lymphatic vessels, but not blood vessels, was lower in tumor tissue compared with benign tissue. These results suggest that activation of VEGFR-1 by VEGF-A within the carcinoma, and activation of lymphatic endothelial cell VEGFR-3 by VEGF-D within the adjacent benign stroma may be important signaling mechanisms involved in the progression and subsequent metastatic spread of prostate cancer. Thus inhibition of these pathways may contribute to therapeutic strategies for the management of prostate cancer. Copyright © 2012 Wiley Periodicals, Inc.

Neurotrophins and tonsillar hypertrophy in children with obstructive sleep apnea.

PubMed

Goldbart, Aviv D; Mager, Edward; Veling, Maria C; Goldman, Julie L; Kheirandish-Gozal, Leila; Serpero, Laura D; Piedimonte, Giovanni; Gozal, David

2007-10-01

Enlarged adenotonsillar tissue (AT) is a major determinant of obstructive sleep apnea (OSA) severity in children; however, mechanisms of AT proliferation are poorly understood. We hypothesized that early exposure to respiratory syncytial virus (RSV) may modify AT proliferation through up-regulation of nerve growth factor (NGF)-neurokinin 1 (NK1) receptor dependent pathways. AT harvested from 34 children with OSA and 25 children with recurrent tonsillitis (RI) were examined for mRNA expression of multiple growth factors and their receptors. In addition, NK1 receptor expression and location, and substance P tissue concentrations were compared in AT from OSA and RI children. NGF mRNA and its high-affinity tyrosine kinase receptor (trkA) expression were selectively increased in OSA (p<0.001). NK1 receptor mRNA and protein expression were also enhanced in OSA (p<0.01), and substance P concentrations in OSA patients were higher than in RI (p<0.0001). AT from OSA children exhibit distinct differences in the expression of NGF and trkA receptors, NK1 receptors, and substance P. The homology between these changes and those observed in the lower airways following RSV infection suggests that RSV may have induced neuro-immunomodulatory changes within AT, predisposing them to increased proliferation, and ultimately contribute to emergence of OSA.

Neurotrophins and Tonsillar Hypertrophy in Children With Obstructive Sleep Apnea

PubMed Central

GOLDBART, AVIV D.; MAGER, EDWARD; VELING, MARIA C.; GOLDMAN, JULIE L.; KHEIRANDISH-GOZAL, LEILA; SERPERO, LAURA D.; PIEDIMONTE, GIOVANNI; GOZAL, DAVID

2013-01-01

Enlarged adenotonsillar tissue (AT) is a major determinant of obstructive sleep apnea (OSA) severity in children; however, mechanisms of AT proliferation are poorly understood. We hypothesized that early exposure to respiratory syncytial virus (RSV) may modify AT proliferation through up-regulation of nerve growth factor (NGF)-neurokinin 1 (NK1) receptor dependent pathways. AT harvested from 34 children with OSA and 25 children with recurrent tonsillitis (RI) were examined for mRNA expression of multiple growth factors and their receptors. In addition, NK1 receptor expression and location, and substance P tissue concentrations were compared in AT from OSA and RI children. NGF mRNA and its high-affinity tyrosine kinase receptor (trkA) expression were selectively increased in OSA (p < 0.001). NK1 receptor mRNA and protein expression were also enhanced in OSA (p < 0.01), and substance P concentrations in OSA patients were higher than in RI (p < 0.0001). AT from OSA children exhibit distinct differences in the expression of NGF and trkA receptors, NK1 receptors, and substance P. The homology between these changes and those observed in the lower airways following RSV infection suggests that RSV may have induced neuro-immunomodulatory changes within AT, predisposing them to increased proliferation, and ultimately contribute to emergence of OSA. PMID:17667845

Long term effects of neonatal hypoglycaemia on pancreatic function.

PubMed

Anju, T R; Paulose, C S

2015-02-01

Low blood glucose in neonates predisposes to long term pancreatic damage. We focused on evaluating long term consequences of neonatal hypoglycaemia in pancreatic functions. Pancreatic function was analysed by measuring DNA/protein synthesis, glucose/ATP uptake in vitro. Gene expression of Pdx1, NeuroD1, Pax4, Bax, caspase 3, Beclin1 were done. Muscarinic receptors were analysed by radio receptor assay. Overall pancreatic efficiency was reduced in one-month-old rats exposed to neonatal hypoglycaemia as indicated by decreased DNA/protein synthesis and glucose/ATP uptake in vitro. Both Pdx1 and Neuro D1 expression were significantly down-regulated whereas Pax4 was up-regulated. Up-regulated Bax, caspase 3 and beclin1 along with reduced muscarinic receptors accounts for activation of cell death pathways. The study revealed a drastic reduction in pancreatic functions along with activation of apoptotic factors in one month old rats exposed to neonatal hypoglycaemia.

Smad2-dependent glycosaminoglycan elongation in aortic valve interstitial cells enhances binding of LDL to proteoglycans.

PubMed

Osman, Narin; Grande-Allen, K Jane; Ballinger, Mandy L; Getachew, Robel; Marasco, Silvana; O'Brien, Kevin D; Little, Peter J

2013-01-01

Calcific aortic valve disease is a progressive condition that shares some common pathogenic features with atherosclerosis. Transforming growth factor-β1 is a recognized mediator of atherosclerosis and is expressed in aortic valve lesions. Transforming growth factorβ1 stimulates glycosaminoglycan elongation of proteoglycans that is associated with increased lipid binding. We investigated the presence of transforming growth factor-β1 and downstream signaling intermediates in diseased human aortic valves and the effects of activated transforming growth factor-β1 receptor signaling on aortic valve interstitial cell proteoglycan synthesis and lipid binding as a possible mechanism for the initiation of the early lesion of calcific aortic valve disease. Diseased human aortic valve leaflets demonstrated strong immunohistochemical staining for transforming growth factor-β1 and phosphorylated Smad2/3. In primary porcine aortic valve interstitial cells, Western blots showed that transforming growth factor-β1 stimulated phosphorylation in both the carboxy and linker regions of Smad2/3, which was inhibited by the transforming growth factor-β1 receptor inhibitor SB431542. Gel electrophoresis and size exclusion chromatography demonstrated that SB431542 decreased transforming growth factor-β1-mediated [(35)S]-sulfate incorporation into proteoglycans in a dose-dependent manner. Further, in proteoglycans derived from transforming growth factor-β1-treated valve interstitial cells, gel mobility shift assays demonstrated that inhibition of transforming growth factor-β1 receptor signaling resulted in decreased lipid binding. Classic transforming growth factor-β1 signaling is present in human aortic valves in vivo and contributes to the modification of proteoglycans expressed by valve interstitial cells in vitro. These findings suggest that transforming growth factor-β1 may promote increased low-density lipoprotein binding in the early phases of calcific aortic valve disease. Copyright © 2013 Elsevier Inc. All rights reserved.

Expression of Inapproptriate Cadherins in Human Breast Carcinomas

DTIC Science & Technology

2000-08-01

fibroblast growth factor receptor signaling. * We showed that cadherin 11 acts in a manner... fibroblast growth factor receptor signaling; and that cadherin 11 promotes epithelial cell motility in a manner similar to N-cadherin. 28 N-Cadherin...levels of E-cadherin; and that N- cadherin-dependent motility may be mediated by fibroblast growth factor receptor signaling. 14. SUBJECT TERMS

Endocannabinoids Stimulate Human Melanogenesis via Type-1 Cannabinoid Receptor*

PubMed Central

Pucci, Mariangela; Pasquariello, Nicoletta; Battista, Natalia; Di Tommaso, Monia; Rapino, Cinzia; Fezza, Filomena; Zuccolo, Michela; Jourdain, Roland; Finazzi Agrò, Alessandro; Breton, Lionel; Maccarrone, Mauro

2012-01-01

We show that a fully functional endocannabinoid system is present in primary human melanocytes (normal human epidermal melanocyte cells), including anandamide (AEA), 2-arachidonoylglycerol, the respective target receptors (CB1, CB2, and TRPV1), and their metabolic enzymes. We also show that at higher concentrations AEA induces normal human epidermal melanocyte apoptosis (∼3-fold over controls at 5 μm) through a TRPV1-mediated pathway that increases DNA fragmentation and p53 expression. However, at lower concentrations, AEA and other CB1-binding endocannabinoids dose-dependently stimulate melanin synthesis and enhance tyrosinase gene expression and activity (∼3- and ∼2-fold over controls at 1 μm). This CB1-dependent activity was fully abolished by the selective CB1 antagonist SR141716 or by RNA interference of the receptor. CB1 signaling engaged p38 and p42/44 mitogen-activated protein kinases, which in turn activated the cyclic AMP response element-binding protein and the microphthalmia-associated transcription factor. Silencing of tyrosinase or microphthalmia-associated transcription factor further demonstrated the involvement of these proteins in AEA-induced melanogenesis. In addition, CB1 activation did not engage the key regulator of skin pigmentation, cyclic AMP, showing a major difference compared with the regulation of melanogenesis by α-melanocyte-stimulating hormone through melanocortin 1 receptor. PMID:22431736

Discovery and characterization of Isofistularin-3, a marine brominated alkaloid, as a new DNA demethylating agent inducing cell cycle arrest and sensitization to TRAIL in cancer cells

PubMed Central

Florean, Cristina; Schnekenburger, Michael; Lee, Jin-Young; Kim, Kyung Rok; Mazumder, Aloran; Song, Sungmi; Kim, Jae-Myun; Grandjenette, Cindy; Kim, Jeoung-Gyun; Yoon, Ah-Young; Dicato, Mario; Kim, Kyu-Won; Christov, Christo; Han, Byung-Woo; Proksch, Peter; Diederich, Marc

2016-01-01

We characterized the brominated alkaloid Isofistularin-3 (Iso-3), from the marine sponge Aplysina aerophoba, as a new DNA methyltransferase (DNMT)1 inhibitor. Docking analysis confirmed our in vitro DNMT inhibition data and revealed binding of Iso-3 within the DNA binding site of DNMT1. Subsequent increased expression of tumor suppressor gene aryl hydrocarbon receptor (AHR) could be correlated to decreased methylation of CpG sites within the essential Sp1 regulatory region of its promoter. Iso-3 induced growth arrest of cancer cells in G0/G1 concomitant with increased p21 and p27 expression and reduced cyclin E1, PCNA and c-myc levels. Reduced proliferation was accompanied by morphological changes typical of autophagy revealed by fluorescent and transmission electron microscopy and validated by LC3I-II conversion. Furthermore, Iso-3 strongly synergized with tumor-necrosis-factor related apoptosis inducing ligand (TRAIL) in RAJI [combination index (CI) = 0.22] and U-937 cells (CI = 0.21) and increased TRAIL-induced apoptosis via a mechanism involving reduction of survivin expression but not of Bcl-2 family proteins nor X-linked inhibitor of apoptosis protein (XIAP). Iso-3 treatment decreased FLIPL expression and triggered activation of endoplasmatic reticulum (ER) stress with increased GRP78 expression, eventually inducing TRAIL receptor death receptor (DR)5 surface expression. Importantly, as a potential candidate for further anticancer drug development, Iso-3 reduced the viability, colony and in vivo tumor forming potential without affecting the viability of PBMCs from healthy donors or zebrafish development. PMID:27006469

Discovery and characterization of Isofistularin-3, a marine brominated alkaloid, as a new DNA demethylating agent inducing cell cycle arrest and sensitization to TRAIL in cancer cells.

PubMed

Florean, Cristina; Schnekenburger, Michael; Lee, Jin-Young; Kim, Kyung Rok; Mazumder, Aloran; Song, Sungmi; Kim, Jae-Myun; Grandjenette, Cindy; Kim, Jeoung-Gyun; Yoon, Ah-Young; Dicato, Mario; Kim, Kyu-Won; Christov, Christo; Han, Byung-Woo; Proksch, Peter; Diederich, Marc

2016-04-26

We characterized the brominated alkaloid Isofistularin-3 (Iso-3), from the marine sponge Aplysina aerophoba, as a new DNA methyltransferase (DNMT)1 inhibitor. Docking analysis confirmed our in vitro DNMT inhibition data and revealed binding of Iso-3 within the DNA binding site of DNMT1. Subsequent increased expression of tumor suppressor gene aryl hydrocarbon receptor (AHR) could be correlated to decreased methylation of CpG sites within the essential Sp1 regulatory region of its promoter. Iso-3 induced growth arrest of cancer cells in G0/G1 concomitant with increased p21 and p27 expression and reduced cyclin E1, PCNA and c-myc levels. Reduced proliferation was accompanied by morphological changes typical of autophagy revealed by fluorescent and transmission electron microscopy and validated by LC3I-II conversion. Furthermore, Iso-3 strongly synergized with tumor-necrosis-factor related apoptosis inducing ligand (TRAIL) in RAJI [combination index (CI) = 0.22] and U-937 cells (CI = 0.21) and increased TRAIL-induced apoptosis via a mechanism involving reduction of survivin expression but not of Bcl-2 family proteins nor X-linked inhibitor of apoptosis protein (XIAP). Iso-3 treatment decreased FLIPL expression and triggered activation of endoplasmatic reticulum (ER) stress with increased GRP78 expression, eventually inducing TRAIL receptor death receptor (DR)5 surface expression. Importantly, as a potential candidate for further anticancer drug development, Iso-3 reduced the viability, colony and in vivo tumor forming potential without affecting the viability of PBMCs from healthy donors or zebrafish development.

Transition from androgenic to neurosteroidal action of 5α-androstane-3α, 17β-diol through the type A γ-aminobutyric acid receptor in prostate cancer progression.

PubMed

Xia, Ding; Lai, Doan V; Wu, Weijuan; Webb, Zachary D; Yang, Qing; Zhao, Lichao; Yu, Zhongxin; Thorpe, Jessica E; Disch, Bryan C; Ihnat, Michael A; Jayaraman, Muralidharan; Dhanasekaran, Danny N; Stratton, Kelly L; Cookson, Michael S; Fung, Kar-Ming; Lin, Hsueh-Kung

2018-04-01

Androgen ablation is the standard of care prescribed to patients with advanced or metastatic prostate cancer (PCa) to slow down disease progression. Unfortunately, a majority of PCa patients under androgen ablation progress to castration-resistant prostate cancer (CRPC). Several mechanisms including alternative intra-prostatic androgen production and androgen-independent androgen receptor (AR) activation have been proposed for CRPC progression. Aldo-keto reductase family 1 member C3 (AKR1C3), a multi-functional steroid metabolizing enzyme, is specifically expressed in the cytoplasm of PCa cells; and positive immunoreactivity of the type A γ-aminobutyric acid receptor (GABA A R), an ionotropic receptor and ligand-gated ion channel, is detected on the membrane of PCa cells. We studied a total of 72 radical prostatectomy cases by immunohistochemistry, and identified that 21 cases exhibited positive immunoreactivities for both AKR1C3 and GABA A R. In the dual positive cancer cases, AKR1C3 and GABA A R subunit α 1 were either expressed in the same cells or in neighboring cells. Among several possible substrates, AKR1C3 reduces 5α-dihydrotesterone (DHT) to form 5α-androstane-3α, 17β-diol (3α-diol). 3α-diol is a neurosteroid that acts as a positive allosteric modulator of the GABA A R in the central nervous system (CNS). We examined the hypothesis that 3α-diol-regulated pathological effects in the prostate are GABA A R-dependent, but are independent of the AR. In GABA A R-positive, AR-negative human PCa PC-3 cells, 3α-diol significantly stimulated cell growth in culture and the in ovo chorioallantoic membrane (CAM) xenograft model. 3α-diol also up-regulated expression of the epidermal growth factor (EGF) family of growth factors and activation of EGF receptor (EGFR) and Src as measured by quantitative polymerase chain reaction and immunoblotting, respectively. Inclusion of GABA A R antagonists reversed 3α-diol-stimulated tumor cell growth, expression of EGF family members, and activation of EGFR and Src to the level observed in untreated cells. Results from the present study suggest that 3α-diol may act as an alternative intra-prostatic neurosteroid that activates AR-independent PCa progression. The involvement of AKR1C3-mediated steroid metabolisms in modulating GABA A R activation and promoting PCa progression requires continued studies. Copyright © 2017 Elsevier Ltd. All rights reserved.

Methamphetamine-induced stereotypy correlates negatively with patch-enhanced prodynorphin and arc mRNA expression in the rat caudate putamen: the role of mu opioid receptor activation.

PubMed

Horner, Kristen A; Noble, Erika S; Gilbert, Yamiece E

2010-06-01

Amphetamines induce stereotypy, which correlates with patch-enhanced c-Fos expression the patch compartment of caudate putamen (CPu). Methamphetamine (METH) treatment also induces patch-enhanced expression of prodynorphin (PD), arc and zif/268 in the CPu. Whether patch-enhanced activation of any of these genes correlates with METH-induced stereotypy is unknown, and the factors that contribute to this pattern of expression are poorly understood. Activation of mu opioid receptors, which are expressed by the neurons of the patch compartment, may underlie METH-induced patch-enhanced gene expression and stereotypy. The current study examined whether striatal mu opioid receptor blockade altered METH-induced stereotypy and patch-enhanced gene expression, and if there was a correlation between the two responses. Animals were intrastriatally infused with the mu antagonist CTAP (10 microg/microl), treated with METH (7.5 mg/kg, s.c.), placed in activity chambers for 3h, and then sacrificed. CTAP pretreatment attenuated METH-induced increases in PD, arc and zif/268 mRNA expression and significantly reduced METH-induced stereotypy. Patch-enhanced PD and arc mRNA expression in the dorsolateral CPu correlated negatively with METH-induced stereotypy. These data indicate that mu opioid receptor activation contributes to METH-induced gene expression in the CPu and stereotypy, and that patch-enhanced PD and arc expression may be a homeostatic response to METH treatment. Copyright 2010 Elsevier Inc. All rights reserved.

Methamphetamine-Induced Stereotypy Correlates Negatively with Patch-Enhanced Prodynorphin and ARC mRNA Expression in the Rat Caudate Putamen: The Role of Mu Opioid Receptor Activation

PubMed Central

Horner, Kristen A.; Noble, Erika S.; Gilbert, Yamiece E.

2010-01-01

Amphetamines induce stereotypy, which correlates with patch-enhanced c-Fos expression the patch compartment of caudate putamen (CPu). Methamphetamine (METH) treatment also induces patch-enhanced expression of prodynorphin (PD), arc and zif/268 in the CPu. Whether patch-enhanced activation of any of these genes correlates with METH-induced stereotypy is unknown, and the factors that contribute to this pattern of expression are poorly understood. Activation of mu opioid receptors, which are expressed by the neurons of the patch compartment, may underlie METH-induced patch-enhanced gene expression and stereotypy. The current study examined whether striatal mu opioid receptor blockade altered METH-induced stereotypy and patch-enhanced gene expression, and if there was a correlation between the two responses. Animals were intrastriatally infused with the mu antagonist CTAP (10 μg/μl), treated with METH (7.5 mg/kg, s.c.), placed in activity chambers for 3h, and then sacrificed. CTAP pretreatment attenuated METH-induced increases in PD, arc and zif/268 mRNA expression and significantly reduced METH-induced stereotypy. Patch-enhanced PD and arc mRNA expression in the dorsolateral CPu correlated negatively with METH-induced stereotypy. These data indicate that mu opioid receptor activation contributes to METH-induced gene expression in the CPu and stereotypy, and that patch-enhanced PD and arc expression may be a homeostatic response to METH treatment. PMID:20298714

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

Baker, Anna D.; Malur, Anagha; Barna, Barbara P.

Peroxisome proliferator-activated receptor-gamma (PPAR{gamma}) is a nuclear transcription factor involved in lipid metabolism that is constitutively expressed in the alveolar macrophages of healthy individuals. PPAR{gamma} has recently been implicated in the catabolism of surfactant by alveolar macrophages, specifically the cholesterol component of surfactant while the mechanism remains unclear. Studies from other tissue macrophages have shown that PPAR{gamma} regulates cholesterol influx, efflux, and metabolism. PPAR{gamma} promotes cholesterol efflux through the liver X receptor-alpha (LXR{alpha}) and ATP-binding cassette G1 (ABCG1). We have recently shown that macrophage-specific PPAR{gamma} knockout (PPAR{gamma} KO) mice accumulate cholesterol-laden alveolar macrophages that exhibit decreased expression of LXR{alpha} andmore » ABCG1 and reduced cholesterol efflux. We hypothesized that in addition to the dysregulation of these cholesterol efflux genes, the expression of genes involved in cholesterol synthesis and influx was also dysregulated and that replacement of PPAR{gamma} would restore regulation of these genes. To investigate this hypothesis, we have utilized a Lentivirus expression system (Lenti-PPAR{gamma}) to restore PPAR{gamma} expression in the alveolar macrophages of PPAR{gamma} KO mice. Our results show that the alveolar macrophages of PPAR{gamma} KO mice have decreased expression of key cholesterol synthesis genes and increased expression of cholesterol receptors CD36 and scavenger receptor A-I (SRA-I). The replacement of PPAR{gamma} (1) induced transcription of LXR{alpha} and ABCG1; (2) corrected suppressed expression of cholesterol synthesis genes; and (3) enhanced the expression of scavenger receptors CD36. These results suggest that PPAR{gamma} regulates cholesterol metabolism in alveolar macrophages.« less

Establishment of primary mixed cell cultures from spontaneous canine mammary tumors: Characterization of classic and new cancer-associated molecules

PubMed Central

Gentile, Luciana B.; Nagamine, Marcia K.; Biondi, Luiz R.; Sanches, Daniel S.; Toyota, Fábio; Giovani, Tatiane M.; de Jesus, Isis P.; da Fonseca, Ivone I. M.; Queiroz-Hazarbassanov, Nicolle; Diaz, Bruno L.; Salles Gomes, Cristina de O. Massoco

2017-01-01

There are many factors which make canine cancer like cancer in humans. The occurrence of spontaneous mammary tumors in pet dogs, tumor genetics, molecular targets and exposure to the same environmental risk factors are among these factors. Therefore, the study of canine cancer can provide useful information to the oncology field. This study aimed to establish and characterize a panel of primary mixed cell cultures obtained from spontaneous canine mammary tumors. Eight established cell cultures obtained from one normal mammary gland, one complex adenoma, one mixed adenoma, two complex carcinomas and two mixed carcinomas were analyzed. The gene expression levels of classic molecular cancer players such as fibroblast growth factor receptor (FGFR) 2, breast cancer (BRCA) 1, BRCA2 and estrogen receptor (ESR) 1 were evaluated. For the first time, three orphan nuclear receptors, estrogen-related receptors (ERRs) α, β and γ were studied in canine mammary cancer. The highest expression level of ERRα was observed in complex carcinoma-derived cell culture, while the highest levels of ERRβ and γ were observed in cells derived from a mixed carcinoma. Meanwhile, complex carcinomas presented the highest levels of expression of ESR1, BRCA1 and FGFR2 among all samples. BRCA2 was found exclusively in complex adenoma. The transcription factor GATA3 had its highest levels in mixed carcinoma samples and its lowest levels in complex adenoma. Proliferation assays were also performed to evaluate the mixed cell cultures response to ER ligands, genistein and DES, both in normoxia and hypoxic conditions. Our results demonstrate that morphological and functional studies of primary mixed cell cultures derived from spontaneous canine mammary tumors are possible and provide valuable tool for the study of various stages of mammary cancer development. PMID:28945747

How glucocorticoid receptors modulate the activity of other transcription factors: a scope beyond tethering.

PubMed

Ratman, Dariusz; Vanden Berghe, Wim; Dejager, Lien; Libert, Claude; Tavernier, Jan; Beck, Ilse M; De Bosscher, Karolien

2013-11-05

The activity of the glucocorticoid receptor (GR), a nuclear receptor transcription factor belonging to subclass 3C of the steroid/thyroid hormone receptor superfamily, is typically triggered by glucocorticoid hormones. Apart from driving gene transcription via binding onto glucocorticoid response elements in regulatory regions of particular target genes, GR can also inhibit gene expression via transrepression, a mechanism largely based on protein:protein interactions. Hereby GR can influence the activity of other transcription factors, without contacting DNA itself. GR is known to inhibit the activity of a growing list of immune-regulating transcription factors. Hence, GCs still rule the clinic for treatments of inflammatory disorders, notwithstanding concomitant deleterious side effects. Although patience is a virtue when it comes to deciphering the many mechanisms GR uses to influence various signaling pathways, the current review is testimony of the fact that groundbreaking mechanistic work has been accumulating over the past years and steadily continues to grow. Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.

Molecular pathways mediating differential responses to lipopolysaccharide between human and baboon arterial endothelial cells.

PubMed

Shi, Qiang; Cox, Laura A; Glenn, Jeremy; Tejero, Maria E; Hondara, Vida; Vandeberg, John L; Wang, Xing Li

2010-02-01

1. Vascular inflammation plays a critical role in atherogenesis. Previously, we showed that baboon arterial endothelial cells (BAEC) were hyporesponsive to lipopolysaccharide (LPS) compared with human arterial endothelial cells (HAEC). 2. In the present study, we investigated mechanisms underlying differential responses between HAEC and BAEC to tumour necrosis factor (TNF)-alpha and LPS. 3. Both HAEC and BAEC responded similarly to TNF-alpha. However, BAEC showed retarded responses to LPS in expression of E-selectin, intercellular adhesion molecule-1, monocyte chemotactic protein-1 and interleukin-8 (P < 0.05). These changes were confirmed at the mRNA level. Tumour necrosis factor-alpha activated nuclear factor-kappaB members such as p50, p52, p65, c-rel and RelB in both HAEC and BAEC. In contrast, LPS activated p50 and p65 only in HAEC. Using microarray assays, we found that TNF receptor-associated factor 2 (TRAF-2), TNF receptor superfamily, member 1A-associated via death domain (TRADD) and nuclear factors such as nuclear factor of kappa in B-cells inhibitor, alpha (NFKBIA) and nuclear factor of kappa in B-cells inhibitor, beta (NFKBIB) were upregulated by LPS only in HAEC. Although the baseline expression of Toll-like receptor (TLR) 4 was low in both HAEC and BAEC, TNF-alpha activated TLR4 expression in both cell types. Although LPS increased TLR4 expression only in HAEC, human and baboon peripheral blood mononuclear cells exhibited similar TLR4 expression and response to LPS. Transfecting BAEC with TLR4/myeloid differentiation protein-2 overexpression vector conferred BAEC responsiveness to LPS. 4. The findings of the present study indicate that an altered TLR4 system may be responsible for the resistance of baboon endothelial cells to LPS. Given the importance of TLR4 in human immune responses and vascular diseases, the natural resistance of baboons to LPS/TLR4-initiated inflammation could make the baboon a valuable animal model in which to study how inflammation affects atherogenesis.

Upregulation of Nicotinic Acetylcholine Receptor alph4+beta2 through a Ligand-Independent PI3Kbeta Mechanism That Is Enhanced by TNFalpha and the Jak2/p38Mapk Pathways.

PubMed

Rogers, Scott W; Gahring, Lorise C

2015-01-01

High affinity nicotine-binding sites in the mammalian brain are neuronal nicotinic acetylcholine receptors (nAChR) assembled from at least alpha4 and beta2 subunits into pentameric ion channels. When exposed to ligands such as nicotine, these receptors respond by undergoing upregulation, a correlate of nicotine addiction. Upregulation can be measured using HEK293 (293) cells that stably express alpha4 and beta2 subunits using quantification of [3H]epibatidine ([3H]Eb) binding to measure mature receptors. Treatment of these cells with choline also produces upregulation through a hemicholinium3 (HC3)-sensitive (choline kinase) and an HC3-insensitive pathway which are both independent of the mechanism used by nicotine for upregulation. In both cases, upregulation is significantly enhanced by the pro-inflammatory cytokine tumor necrosis factor alpha (TNFα) which signals through its receptor Tnfr1 to activate p38Mapk. Here we report that the inhibition of class1 phosphoinositide 3-kinases isoform PI3Kbeta using the selective antagonist PI828 is alone sufficient to produce upregulation and enhance both nicotine and choline HC3-sensitive mediated upregulation. Further, these processes are impacted upon by an AG-490 sensitive Jak2-associated pathway. Both PI3Kbeta (negative) and Jak2 (positive) modulation of upregulation converge through p38Mapk and both overlap with TNFalpha enhancement of this process. Upregulation through the PI3Kbeta pathway did not require Akt. Collectively these findings support upregulation of endogenous alpha4beta2 as a balance among cellular signaling networks that are highly responsive to multiple environmental, inflammatory and metabolic agents. The findings also suggest how illness and metabolic stress could alter the expression of this important nicotinic receptor and novel avenues to intercede in modifying its expression.

Characterization of germ cell-specific expression of the orphan nuclear receptor, germ cell nuclear factor.

PubMed

Katz, D; Niederberger, C; Slaughter, G R; Cooney, A J

1997-10-01

Nuclear receptors, such as those for androgens, estrogens, and progesterones, control many reproductive processes. Proteins with structures similar to these receptors, but for which ligands have not yet been identified, have been termed orphan nuclear receptors. One of these orphans, germ cell nuclear factor (GCNF), has been shown to be germ cell specific in the adult and, therefore, may also participate in the regulation of reproductive functions. In this paper, we examine more closely the expression patterns of GCNF in germ cells to begin to define spatio-temporal domains of its activity. In situ hybridization showed that GCNF messenger RNA (mRNA) is lacking in the testis of hypogonadal mutant mice, which lack developed spermatids, but is present in the wild-type testis. Thus, GCNF is, indeed, germ cell specific in the adult male. Quantitation of the specific in situ hybridization signal in wild-type testis reveals that GCNF mRNA is most abundant in stage VII round spermatids. Similarly, Northern analysis and specific in situ hybridization show that GCNF expression first occurs in testis of 20-day-old mice, when round spermatids first emerge. Therefore, in the male, GCNF expression occurs postmeiotically and may participate in the morphological changes of the maturing spermatids. In contrast, female expression of GCNF is shown in growing oocytes that have not completed the first meiotic division. Thus, GCNF in the female is expressed before the completion of meiosis. Finally, the nature of the two different mRNAs that hybridize to the GCNF complementary DNA was studied. Although both messages contain the DNA binding domain, only the larger message is recognized by a probe from the extreme 3' untranslated region. In situ hybridization with these differential probes demonstrates that both messages are present in growing oocytes. In addition, the coding region and portions of the 3' untranslated region of the GCNF complementary DNA are conserved in the rat.

Pattern of somatostatin receptors expression in normal and bladder cancer tissue samples.

PubMed

Karavitakis, Markos; Msaouel, Pavlos; Michalopoulos, Vassilis; Koutsilieris, Michael

2014-06-01

Known risks factors for bladder cancer progression and recurrence are limited regarding their prognostic ability. Therefore identification of molecular determinants of disease progression could provide with more specific prognostic information and could be translated into new approaches for biomarker development. In the present study we evaluated, the expression patterns of somatostatin receptors 1-5 (SSTRs) in normal and tumor bladder tissues. The expression of SSTR1-5 was characterized in 45 normal and bladder cancer tissue samples using reverse transcriptase-polymerase chain reaction (RT-PCR). SSTR1 was expressed in 24 samples, SSTR2 in 15, SSTR3 in 23, SSTR4 in 16 and SSTR5 in all but one sample. Bladder cancer tissue samples expressed lower levels of SSTR3. Co-expression of SSTRs was associated with superficial disease. Our results demonstrate, for the first time, that there is expression of SSTR in normal and bladder cancer urothelium. Further studies are required to evaluate the prognostic and therapeutic significance of these findings. Copyright© 2014 International Institute of Anticancer Research (Dr. John G. Delinassios), All rights reserved.

Expression of death decoy receptor-3 (DcR3) in human breast cancer and its functional effects on breast cancer cells in vitro.

PubMed

Ge, Zhicheng; Sanders, Andrew J; Ye, Lin; Wang, Yu; Jiang, Wen G

2011-01-01

Death Decoy Receptor-3 (DcR3), otherwise known as tumour necrosis factor receptor superfamily member 6b, is suggested to be involved in the progression and immune evasion of malignant tumours. Its ligands include FASL and LIGHT (Tumour necrosis factor ligand superfamily member 14). DcR3 has been found to be amplified in certain solid tumours. However, its role in breast tumours remains unclear. In the present study, we examined the role played by DcR3 in MCF7 and MDA-MB-231 cell lines. The expression of DcR3 was examined in MCF7 and MDA-MB-231 cell lines using immunocytochemical staining and RT-PCR. Anti-DcR3 hammerhead ribozyme transgenes were constructed and transfected into cells to create DcR3 knock-down cell sublines. The biological impact of modifying DcR3 expression in breast cancer cells was evaluated using a variety of in vitro assays, including growth, adhesion, migration and invasion models. MCF7 and MDA-MB-231 cells, usually expressing DcR3, were transfected with the anti-DcR3 ribozyme transgene. Stable transfectants containing the DcR3 ribozyme transgene (MCF7DcR3KO, MDA-MB-231DcR3KO) displayed a reduction of DcR3 expression at mRNA and protein levels. DcR3 knockdown in MCF7 cells was found to significantly reduce invasive capacity compared to pEF6 control cell lines (30.78 +/- 6.40 vs.151.67 +/- 17.67 P < 0.001). The rate of migration in MCF7DcR3KO was significantly lower than MCF7pEF6 (P < 0.001). In contrast, no such significant differences was seen between MDA-MB-231DcR3KO and MDA-MB-231pEF6. Suppressing DcR3 expression was found to have an inhibitory effect on cellular invasion and migration in MCF7 breast cancer cells. This suggests that the invasion and migration capacity of this breast cancer cell line may, at least partly, depend on DcR3. DcR3 may be regarded as a negative regulator for aggressiveness during the development and progression of certain types of breast cancer.

[Steroid and xenobiotic receptor (SXR), multidrug resistance gene (MDR1) and GSTs, SULTs and CYP polymorphism expression in invasive bladder cancer, analysis of their expression and correlation with other prognostic factors].

PubMed

Rioja Zuazu, J; Bandrés Elizalde, E; Rosell Costa, D; Rincón Mayans, A; Zudaire Bergera, J; Gil Sanz, M J; Rioja Sanz, L A; García Foncillas, J; Berián Polo, J M

2007-01-01

Steroid and Xenobiotic Receptor (SXR) has demonstrated its activation by numerous drugs, including cytochrome P450 potent inducers like rifampicina or cotrimazol. The role of SXR is well known, and lies regulating in a positive manner cytochrome P450 3A4 (CYP3A4) transcription and the multidrug resistance gene (MDR1), it's considered a key in the xenobiotic detoxification mechanism, being involved in all phases of the detoxification process. Enzymes involved in Policyclic Aromatic hidrocarbures (PAH) metabolism and degradation are polymorphic in humans, including glutation S-transferases (GSTs), N-acetiltransferases (NATs), sulfotransferases (SULTs)1A1 and cytochrome p450 (CYP)1B1. The objectives we've planned are: 1. Analyze the expression of the transcription factor SXR and MDR1 in bladder by means of RT-PCR real time, both in normal bladder and in tumoral bladder. 2. Analyze the relation between clinical and pathological factors with the expression of SXR and MDR1. 3. Analyze the expression of the polymorphims CYP1B1, GSTM1 GSTT1 and SULT1A1 and their correlation with different clinic-pathological and molecular factors. In a prospective way the size of the sample was estimated. In 67 patients from two institutions (Hospital Universitario Miguel Servet (49 HUMS) and Clinica Universitaria de Navarra (18 CUN)), diagnosed of invasive bladder cancer and treated by means of radical cystectomy, were determined the expression of both SXR and MDR1 by means of real time PCR, as well as the polymorphisms CYP1B1, GSTM1 GSTT1 y SULT1A1 by means of RFLP (Restriction fragment length polymorphism). Correlations with other prognostic factors by contingency tables were performed. Average follow up was 23.7 months with a median of 28.26 months. Of the 67 patients studied, 31 patients (46.3) presented disease progression, in form of local recurrence or in distant metastasis or both. With a average time to progression of 12.4 months and a median of 10 months, with a range of 1.1 month to 31.9 month. 36 patients (53.7%) did not have any evidence of disease progression during follow up. The Steroid and Xenobiotic Receptor as well as the Multidrug Resistance Gene (MDR1) are expressed in both normal bladder (0.94DeltaCt y 0.94DeltaCt) and tumoral bladder in the cystectomy specimen (1.09 DeltaCt y 0.45 DeltaCt). We've analyzed their expression in a quantitative manner and in a qualitative manner. The expression of SXR correlates with the presence of ca. in situ (p=0.024), vasculo-lymphatic invasion (p=0.05) mean while MDR1 correlates with presence of vasculo-lymphatic invasion (p=0.05) Both factors are correlate between each others (p=0.011). Polymorphisms: CYP1B1, GSTM1, GSTT1 and SULT1A1, are expressed in these patients but their expression doesn't correlates with any prognostic factor Both SXR and MDR1 are expressed in normal bladder as well as in tumoral bladder. And their expression correlates with different prognostic factors with influence in the survival described in the literature.

S100A8/A9 is associated with estrogen receptor loss in breast cancer.

PubMed

Bao, Y I; Wang, Antao; Mo, Juanfen

2016-03-01

S100A8 and S100A9 are calcium-binding proteins that are secreted primarily by granulocytes and monocytes, and are upregulated during the inflammatory response. S100A8 and S100A9 have been identified to be expressed by epithelial cells involved in malignancy. In the present study, the transcriptional levels of S100A8 and S100A9 were investigated in various subtypes of breast cancer (BC), and the correlation with estrogen receptor 1 (ESR1) and GATA binding protein 3 (GATA3) gene expression was evaluated using microarray datasets. The expression of S100A8 and S100A9 in BC cells was assessed by reverse transcription-polymerase chain reaction (RT-PCR). The regulation of ESR1 and GATA3 by administration of recombinant S100A8/A9 was examined in the BC MCF-7 cell line using quantitative (q)PCR. The association between S100A8 and S100A9 and overall survival (OS) was investigated in GeneChip® data of BC. The expression levels of S100A8 and S100A9 were higher in human epidermal growth factor receptor 2 (Her2)-amplified and basal-like BC. The messenger (m)RNA levels of S100A8 and S100A9 were inversely correlated with ESR1 and GATA3 expression. S100A8/A9 induced a 10-fold decrease in the mRNA levels of ESR1 in MCF-7 cells. Poor OS was associated with high expression levels of S100A9, but not with high expression levels of S100A8 in BC. In conclusion, strong expression and secretion of S100A8/A9 may be associated with the loss of estrogen receptor in BC, and may be involved in the poor prognosis of Her2+/basal-like subtypes of BC.

S100A8/A9 is associated with estrogen receptor loss in breast cancer

PubMed Central

BAO, YI; WANG, ANTAO; MO, JUANFEN

2016-01-01

S100A8 and S100A9 are calcium-binding proteins that are secreted primarily by granulocytes and monocytes, and are upregulated during the inflammatory response. S100A8 and S100A9 have been identified to be expressed by epithelial cells involved in malignancy. In the present study, the transcriptional levels of S100A8 and S100A9 were investigated in various subtypes of breast cancer (BC), and the correlation with estrogen receptor 1 (ESR1) and GATA binding protein 3 (GATA3) gene expression was evaluated using microarray datasets. The expression of S100A8 and S100A9 in BC cells was assessed by reverse transcription-polymerase chain reaction (RT-PCR). The regulation of ESR1 and GATA3 by administration of recombinant S100A8/A9 was examined in the BC MCF-7 cell line using quantitative (q)PCR. The association between S100A8 and S100A9 and overall survival (OS) was investigated in GeneChip® data of BC. The expression levels of S100A8 and S100A9 were higher in human epidermal growth factor receptor 2 (Her2)-amplified and basal-like BC. The messenger (m)RNA levels of S100A8 and S100A9 were inversely correlated with ESR1 and GATA3 expression. S100A8/A9 induced a 10-fold decrease in the mRNA levels of ESR1 in MCF-7 cells. Poor OS was associated with high expression levels of S100A9, but not with high expression levels of S100A8 in BC. In conclusion, strong expression and secretion of S100A8/A9 may be associated with the loss of estrogen receptor in BC, and may be involved in the poor prognosis of Her2+/basal-like subtypes of BC. PMID:26998104

Hypoxia-inducing factors as master regulators of stemness properties and altered metabolism of cancer- and metastasis-initiating cells

PubMed Central

Mimeault, Murielle; Batra, Surinder K

2013-01-01

Accumulating lines of experimental evidence have revealed that hypoxia-inducible factors, HIF-1α and HIF-2α, are key regulators of the adaptation of cancer- and metastasis-initiating cells and their differentiated progenies to oxygen and nutrient deprivation during cancer progression under normoxic and hypoxic conditions. Particularly, the sustained stimulation of epidermal growth factor receptor (EGFR), insulin-like growth factor-1 receptor (IGF-1R), stem cell factor (SCF) receptor KIT, transforming growth factor-β receptors (TGF-βRs) and Notch and their downstream signalling elements such as phosphatidylinositol 3′-kinase (PI3K)/Akt/molecular target of rapamycin (mTOR) may lead to an enhanced activity of HIFs. Moreover, the up-regulation of HIFs in cancer cells may also occur in the hypoxic intratumoral regions formed within primary and secondary neoplasms as well as in leukaemic cells and metastatic prostate and breast cancer cells homing in the hypoxic endosteal niche of bone marrow. The activated HIFs may induce the expression of numerous gene products such as induced pluripotency-associated transcription factors (Oct-3/4, Nanog and Sox-2), glycolysis- and epithelial-mesenchymal transition (EMT) programme-associated molecules, including CXC chemokine receptor 4 (CXCR4), snail and twist, microRNAs and angiogenic factors such as vascular endothelial growth factor (VEGF). These gene products in turn can play critical roles for high self-renewal ability, survival, altered energy metabolism, invasion and metastases of cancer cells, angiogenic switch and treatment resistance. Consequently, the targeting of HIF signalling network and altered metabolic pathways represents new promising strategies to eradicate the total mass of cancer cells and improve the efficacy of current therapies against aggressive and metastatic cancers and prevent disease relapse. PMID:23301832

Comparative gene expression profiling of ADAMs, MMPs, TIMPs, EMMPRIN, EGF-R and VEGFA in low grade meningioma.

PubMed

Rooprai, Harcharan K; Martin, Andrew J; King, Andrew; Appadu, Usha D; Jones, Huw; Selway, Richard P; Gullan, Richard W; Pilkington, Geoffrey J

2016-12-01

MMPs (matrix metalloproteinases), ADAMs (a disintegrin and metalloproteinase) and TIMPs (tissue inhibitors of metalloproteinases) are implicated in invasion and angiogenesis: both are tissue remodeling processes involving regulated proteolysis of the extracellular matrix, growth factors and their receptors. The expression of these three groups and their correlations with clinical behaviour has been reported in gliomas but a similar comprehensive study in meningiomas is lacking. In this study, we aimed to evaluate the patterns of expression of 23 MMPs, 4 TIMPs, 8 ADAMs, selective growth factors and their receptors in 17 benign meningiomas using a quantitative real-time polymerase chain reaction (qPCR). Results indicated very high gene expression of 13 proteases, inhibitors and growth factors studied: MMP2 and MMP14, TIMP-1, -2 and -3, ADAM9, 10, 12, 15 and 17, EGF-R, EMMPRIN and VEGF-A, in almost every meningioma. Expression pattern analysis showed several positive correlations between MMPs, ADAMs, TIMPs and growth factors. Furthermore, our findings suggest that expression of MMP14, ADAM9, 10, 12, 15 and 17, TIMP-2, EGF-R and EMMPRIN reflects histological subtype of meningioma such that fibroblastic subtype had the highest mRNA expression, transitional subtype was intermediate and meningothelial type had the lowest expression. In conclusion, this is the first comprehensive study characterizing gene expression of 8 ADAMs in meningiomas. These neoplasms, although by histological definition benign, have invasive potential. Taken together, the selected elevated gene expression pattern may serve to identify targets for therapeutic intervention or indicators of biological progression and recurrence.

The postnatal 5-HT1A receptor regulates adult anxiety and depression differently via multiple molecules.

PubMed

Ishikawa, Chihiro; Shiga, Takashi

2017-08-01

Serotonin (5-HT) and the 5-HT 1A receptor during development are known to modulate anxiety and depression in later life. However, the brain mechanisms linking the postnatal 5-HT system and adult behavior remain unknown. Here, we examined the effects of pharmacological 5-HT 1A receptor activation during the postnatal period on anxiety and depression-like behavior in adult BALB/c male mice. To elucidate the underlying mechanisms, we measured mRNA expression of the 5-HT 1A receptor, brain-derived neurotrophic factor (BDNF), GABA A receptor subunits, and AMPA receptor subunits in the medial prefrontal cortex (mPFC), amygdala, and hippocampus. Treatment with the selective 5-HT reuptake inhibitor (fluoxetine) and 5-HT 1A receptor agonist (8-OH-DPAT) during the postnatal period decreased anxiety-like behavior in adulthood, whereas only 8-OH-DPAT treatment increased depression-like behavior. Concomitantly with the behavioral effects, postnatal treatment with fluoxetine and 8-OH-DPAT decreased the mRNA expression of the GABA A receptor α3 subunit in the mPFC and ventral hippocampus in adulthood, while 8-OH-DPAT, but not fluoxetine, decreased the mRNA expression of the 5-HT 1A receptor and BDNF in the mPFC and the GABA A receptor α2 subunit in the mPFC and ventral hippocampus. On the basis of the correlative changes between behavior and mRNA expression, these results suggest that the GABA A receptor α3 subunit in the mPFC and ventral hippocampus may regulate anxiety-like behavior. In contrast, depression-like behavior may be regulated by the 5-HT 1A receptor and BDNF in the mPFC and by the GABA A receptor α2 subunit in the mPFC and ventral hippocampus. In summary, activation of the 5-HT 1A receptor during the postnatal period may reduce anxiety levels, but increase depression levels during adulthood via different multiple molecules in the mPFC and ventral hippocampus. Copyright © 2017 Elsevier Inc. All rights reserved.

An ancient neurotrophin receptor code; a single Runx/Cbfβ complex determines somatosensory neuron fate specification in zebrafish

PubMed Central

Condon, Logan; Raible, David W.

2017-01-01

In terrestrial vertebrates such as birds and mammals, neurotrophin receptor expression is considered fundamental for the specification of distinct somatosensory neuron types where TrkA, TrkB and TrkC specify nociceptors, mechanoceptors and proprioceptors/mechanoceptors, respectively. In turn, Runx transcription factors promote neuronal fate specification by regulating neurotrophin receptor and sensory receptor expression where Runx1 mediates TrkA+ nociceptor diversification while Runx3 promotes a TrkC+ proprioceptive/mechanoceptive fate. Here, we report in zebrafish larvae that orthologs of the neurotrophin receptors in contrast to terrestrial vertebrates mark overlapping and distinct subsets of nociceptors suggesting that TrkA, TrkB and TrkC do not intrinsically promote nociceptor, mechanoceptor and proprioceptor/mechanoceptor neuronal fates, respectively. While we find that zebrafish Runx3 regulates nociceptors in contrast to terrestrial vertebrates, it shares a conserved regulatory mechanism found in terrestrial vertebrate proprioceptors/mechanoceptors in which it promotes TrkC expression and suppresses TrkB expression. We find that Cbfβ, which enhances Runx protein stability and affinity for DNA, serves as an obligate cofactor for Runx in neuronal fate determination. High levels of Runx can compensate for the loss of Cbfβ, indicating that in this context Cbfβ serves solely as a signal amplifier of Runx activity. Our data suggests an alteration/expansion of the neurotrophin receptor code of sensory neurons between larval teleost fish and terrestrial vertebrates, while the essential roles of Runx/Cbfβ in sensory neuron cell fate determination while also expanded are conserved. PMID:28708822

An ancient neurotrophin receptor code; a single Runx/Cbfβ complex determines somatosensory neuron fate specification in zebrafish.

PubMed

Gau, Philia; Curtright, Andrew; Condon, Logan; Raible, David W; Dhaka, Ajay

2017-07-01

In terrestrial vertebrates such as birds and mammals, neurotrophin receptor expression is considered fundamental for the specification of distinct somatosensory neuron types where TrkA, TrkB and TrkC specify nociceptors, mechanoceptors and proprioceptors/mechanoceptors, respectively. In turn, Runx transcription factors promote neuronal fate specification by regulating neurotrophin receptor and sensory receptor expression where Runx1 mediates TrkA+ nociceptor diversification while Runx3 promotes a TrkC+ proprioceptive/mechanoceptive fate. Here, we report in zebrafish larvae that orthologs of the neurotrophin receptors in contrast to terrestrial vertebrates mark overlapping and distinct subsets of nociceptors suggesting that TrkA, TrkB and TrkC do not intrinsically promote nociceptor, mechanoceptor and proprioceptor/mechanoceptor neuronal fates, respectively. While we find that zebrafish Runx3 regulates nociceptors in contrast to terrestrial vertebrates, it shares a conserved regulatory mechanism found in terrestrial vertebrate proprioceptors/mechanoceptors in which it promotes TrkC expression and suppresses TrkB expression. We find that Cbfβ, which enhances Runx protein stability and affinity for DNA, serves as an obligate cofactor for Runx in neuronal fate determination. High levels of Runx can compensate for the loss of Cbfβ, indicating that in this context Cbfβ serves solely as a signal amplifier of Runx activity. Our data suggests an alteration/expansion of the neurotrophin receptor code of sensory neurons between larval teleost fish and terrestrial vertebrates, while the essential roles of Runx/Cbfβ in sensory neuron cell fate determination while also expanded are conserved.

Direct activation of RIP3/MLKL-dependent necrosis by herpes simplex virus 1 (HSV-1) protein ICP6 triggers host antiviral defense

PubMed Central

Wang, Xing; Li, Yun; Liu, Shan; Yu, Xiaoliang; Li, Lin; Shi, Cuilin; He, Wenhui; Li, Jun; Xu, Lei; Hu, Zhilin; Yu, Lu; Yang, Zhongxu; Chen, Qin; Ge, Lin; Zhang, Zili; Zhou, Biqi; Jiang, Xuejun; Chen, She; He, Sudan

2014-01-01

The receptor-interacting kinase-3 (RIP3) and its downstream substrate mixed lineage kinase domain-like protein (MLKL) have emerged as the key cellular components in programmed necrotic cell death. Receptors for the cytokines of tumor necrosis factor (TNF) family and Toll-like receptors (TLR) 3 and 4 are able to activate RIP3 through receptor-interacting kinase-1 and Toll/IL-1 receptor domain-containing adapter inducing IFN-β, respectively. This form of cell death has been implicated in the host-defense system. However, the molecular mechanisms that drive the activation of RIP3 by a variety of pathogens, other than the above-mentioned receptors, are largely unknown. Here, we report that human herpes simplex virus 1 (HSV-1) infection triggers RIP3-dependent necrosis. This process requires MLKL but is independent of TNF receptor, TLR3, cylindromatosis, and host RIP homotypic interaction motif-containing protein DNA-dependent activator of IFN regulatory factor. After HSV-1 infection, the viral ribonucleotide reductase large subunit (ICP6) interacts with RIP3. The formation of the ICP6–RIP3 complex requires the RHIM domains of both proteins. An HSV-1 ICP6 deletion mutant failed to cause effective necrosis of HSV-1–infected cells. Furthermore, ectopic expression of ICP6, but not RHIM mutant ICP6, directly activated RIP3/MLKL-mediated necrosis. Mice lacking RIP3 exhibited severely impaired control of HSV-1 replication and pathogenesis. Therefore, this study reveals a previously uncharacterized host antipathogen mechanism. PMID:25316792

Improved decision making for prioritizing tumor targeting antibodies in human xenografts: Utility of fluorescence imaging to verify tumor target expression, antibody binding and optimization of dosage and application schedule.

PubMed

Dobosz, Michael; Haupt, Ute; Scheuer, Werner

2017-01-01

Preclinical efficacy studies of antibodies targeting a tumor-associated antigen are only justified when the expression of the relevant antigen has been demonstrated. Conventionally, antigen expression level is examined by immunohistochemistry of formalin-fixed paraffin-embedded tumor tissue section. This method represents the diagnostic "gold standard" for tumor target evaluation, but is affected by a number of factors, such as epitope masking and insufficient antigen retrieval. As a consequence, variances and discrepancies in histological staining results can occur, which may influence decision-making and therapeutic outcome. To overcome these problems, we have used different fluorescence-labeled therapeutic antibodies targeting human epidermal growth factor receptor (HER) family members and insulin-like growth factor-1 receptor (IGF1R) in combination with fluorescence imaging modalities to determine tumor antigen expression, drug-target interaction, and biodistribution and tumor saturation kinetics in non-small cell lung cancer xenografts. For this, whole-body fluorescence intensities of labeled antibodies, applied as a single compound or antibody mixture, were measured in Calu-1 and Calu-3 tumor-bearing mice, then ex vivo multispectral tumor tissue analysis at microscopic resolution was performed. With the aid of this simple and fast imaging method, we were able to analyze the tumor cell receptor status of HER1-3 and IGF1R, monitor the antibody-target interaction and evaluate the receptor binding sites of anti-HER2-targeting antibodies. Based on this, the most suitable tumor model, best therapeutic antibody, and optimal treatment dosage and application schedule was selected. Predictions drawn from obtained imaging data were in excellent concordance with outcome of conducted preclinical efficacy studies. Our results clearly demonstrate the great potential of combined in vivo and ex vivo fluorescence imaging for the preclinical development and characterization of monoclonal antibodies.

LGR4 and Its Ligands, R-Spondin 1 and R-Spondin 3, Regulate Food Intake in the Hypothalamus of Male Rats

PubMed Central

Li, Ji-Yao; Chai, Biaoxin; Zhang, Weizhen; Fritze, Danielle M.; Zhang, Chao

2014-01-01

The hypothalamus plays a key role in the regulation of feeding behavior. Several hypothalamic nuclei, including the arcuate nucleus (ARC), paraventricular nucleus, and ventromedial nucleus of the hypothalamus (VMH), are involved in energy homeostasis. Analysis of microarray data derived from ARC revealed that leucine-rich repeat-containing G protein-coupled receptor 4 (LGR4) is highly expressed. LGR4, LGR5, and LGR6 form a subfamily of closely related receptors. Recently, R-spondin (Rspo) family proteins were identified as ligands of the LGR4 subfamily. In the present study, we investigated the distribution and function of LGR4–LGR6 and Rspos (1–4) in the brain of male rat. In situ hybridization showed that LGR4 is expressed in the ARC, VMH, and median eminence of the hypothalamus. LGR4 colocalizes with neuropeptide Y, proopiomelanocortin, and brain-derived neurotrophic factor neurons. LGR5 is not detectable with in situ hybridization; LGR6 is only expressed in the epithelial lining of the lower portion of the third ventricle and median eminence. Rspo1 is expressed in the VMH and down-regulated with fasting. Rspo3 is expressed in the paraventricular nucleus and also down-regulated with fasting. Rspos 1 and 3 colocalize with the neuronal marker HuD, indicating that they are expressed by neurons. Injection of Rspo1 or Rspo3 into the third brain ventricle inhibited food intake. Rspo1 decreased neuropeptide Y and increased proopiomelanocortin expression in the ARC. Rspo1 and Rspo3 mRNA is up-regulated by insulin. These data indicate that Rspo1 and Rspo3 and their receptor LGR4 form novel circuits in the brain to regulate energy homeostasis. PMID:24280058

LGR4 and its ligands, R-spondin 1 and R-spondin 3, regulate food intake in the hypothalamus of male rats.

PubMed

Li, Ji-Yao; Chai, Biaoxin; Zhang, Weizhen; Fritze, Danielle M; Zhang, Chao; Mulholland, Michael W

2014-02-01

The hypothalamus plays a key role in the regulation of feeding behavior. Several hypothalamic nuclei, including the arcuate nucleus (ARC), paraventricular nucleus, and ventromedial nucleus of the hypothalamus (VMH), are involved in energy homeostasis. Analysis of microarray data derived from ARC revealed that leucine-rich repeat-containing G protein-coupled receptor 4 (LGR4) is highly expressed. LGR4, LGR5, and LGR6 form a subfamily of closely related receptors. Recently, R-spondin (Rspo) family proteins were identified as ligands of the LGR4 subfamily. In the present study, we investigated the distribution and function of LGR4-LGR6 and Rspos (1-4) in the brain of male rat. In situ hybridization showed that LGR4 is expressed in the ARC, VMH, and median eminence of the hypothalamus. LGR4 colocalizes with neuropeptide Y, proopiomelanocortin, and brain-derived neurotrophic factor neurons. LGR5 is not detectable with in situ hybridization; LGR6 is only expressed in the epithelial lining of the lower portion of the third ventricle and median eminence. Rspo1 is expressed in the VMH and down-regulated with fasting. Rspo3 is expressed in the paraventricular nucleus and also down-regulated with fasting. Rspos 1 and 3 colocalize with the neuronal marker HuD, indicating that they are expressed by neurons. Injection of Rspo1 or Rspo3 into the third brain ventricle inhibited food intake. Rspo1 decreased neuropeptide Y and increased proopiomelanocortin expression in the ARC. Rspo1 and Rspo3 mRNA is up-regulated by insulin. These data indicate that Rspo1 and Rspo3 and their receptor LGR4 form novel circuits in the brain to regulate energy homeostasis.

Spatial pattern of receptor expression in the olfactory epithelium.

PubMed Central

Nef, P; Hermans-Borgmeyer, I; Artières-Pin, H; Beasley, L; Dionne, V E; Heinemann, S F

1992-01-01

A PCR-based strategy for amplifying putative receptors involved in murine olfaction was employed to isolate a member (OR3) of the seven-transmembrane-domain receptor superfamily. During development, the first cells that express OR3 appear adjacent to the wall of the telencephalic vesicle at embryonic day 10. The OR3 receptor is uniquely expressed in a subset of olfactory cells that have a characteristic bilateral symmetry in the adult olfactory epithelium. This receptor and its specific pattern of expression may serve a functional role in odor coding or, alternatively, may play a role in the development of the olfactory system. Images PMID:1384038

Angiogenesis and Vascular Architecture in Pheochromocytomas

PubMed Central

Favier, Judith; Plouin, Pierre-François; Corvol, Pierre; Gasc, Jean-Marie

2002-01-01

Angiogenesis is a critical step in tumor growth and metastatic invasion. We here report the study of the vascular status of 10 benign and 9 malignant pheochromocytomas. We examined the vascular architecture after immunostaining endothelial cells (CD34) and vascular smooth muscle cells (α-actin) and identified a vascular pattern characteristic of malignant lesions. To define a gene expression profile indicative of the invasive phenotype, we studied by in situ hybridization the expression of genes encoding several pro- and anti-angiogenic factors [hypoxia-inducible factor (HIF-1α), EPAS1, vascular endothelial growth factor (VEGF), VEGF receptors, angiopoietins and their receptor Tie2, five genes of the endothelin system, and thrombospondin 1]. A semiquantitative evaluation of the labeling revealed an induction of genes encoding EPAS1, VEGF, VEGFR-1, VEGFR-2, endothelin receptor, type B (ETB) and endothelin receptor, type A (ETA) in malignant pheochromocytomas as compared to benign tumors. These differences were observed in tumor cells, in endothelial cells, or in both. Quantification by real-time reverse-transcriptase polymerase chain reaction showed an increase of EPAS1, VEGF, and ETB transcripts of 4.5-, 3.5-, and 10-fold, respectively, in malignant versus benign tumors. Furthermore, we observed a strong correlation between the expression of EPAS1 and VEGF in tumoral tissue and between EPAS1 and ETB in endothelial cells. Altogether, our observations show that analysis of angiogenesis provides promising new criteria for the diagnosis of malignant pheochromocytomas. PMID:12368197

Signaling of Prostaglandin E Receptors, EP3 and EP4 Facilitates Wound Healing and Lymphangiogenesis with Enhanced Recruitment of M2 Macrophages in Mice.

PubMed

Hosono, Kanako; Isonaka, Risa; Kawakami, Tadashi; Narumiya, Shuh; Majima, Masataka

2016-01-01

Lymphangiogenesis plays an important role in homeostasis, metabolism, and immunity, and also occurs during wound-healing. Here, we examined the roles of prostaglandin E2 (PGE2) receptor (EP) signaling in enhancement of lymphangiogenesis in wound healing processes. The hole-punch was made in the ears of male C57BL/6 mice using a metal ear punch. Healing process and lymphangiogenesis together with macrophage recruitment were analyzed in EP knockout mice. Lymphangiogenesis was up-regulated in the granulation tissues at the margins of punched-hole wounds in mouse ears, and this increase was accompanied by increased expression levels of COX-2 and microsomal prostaglandin E synthase-1. Administration of celecoxib, a COX-2 inhibitor, suppressed lymphangiogenesis in the granulation tissues and reduced the induction of the pro-lymphangiogenic factors, vascular endothelial growth factor (VEGF) -C and VEGF-D. Topical applications of selective EP receptor agonists enhanced the expressions of lymphatic vessel endothelial hyaluronan receptor-1 and VEGF receptor-3. The wound-healing processes and recruitment of CD11b-positive macrophages, which produced VEGF-C and VEGF-D, were suppressed under COX-2 inhibition. Mice lacking either EP3 or EP4 exhibited reduced wound-healing, lymphangiogenesis and recruitment of M2 macrophages, compared with wild type mice. Proliferation of cultured human lymphatic endothelial cells was not detected under PGE2 stimulation. Lymphangiogenesis and recruitment of M2 macrophages that produced VEGF-C/D were suppressed in mice treated with a COX-2 inhibitor or lacking either EP3 or EP4 during wound healing. COX-2 and EP3/EP4 signaling may be novel targets to control lymphangiogenesis in vivo.

Prognostic significance of epidermal growth factor receptor in surgically treated squamous cell lung cancer patients.

PubMed

Niemiec, Joanna; Kołodziejski, Leszek; Dyczek, Sonia; Gasińska, Anna

2004-01-01

Epidermal growth factor receptor (EGFR) is one of signalling pathways activated during premalignant proliferative changes in the airway epithelium. However there is no agreement about prognostic significance of EGFR expression in non-small cell lung cancer (NSCLC). Facts mentioned above prompted us to study EGFR expression in the group of 78 surgically treated squamous cell lung cancer (SqCLC) patients. The EGFR expression was visualized in formalin-fixed, paraffin-embedded sections, using immunohistochemistry. Three methods of assessment of EGFR expression were applied: percentage of cells with membranous EGFR expression--EGFR labellig index (EGFR LI), percentage of fields with membranous EGFR staining (PS%) and staining intensity (absent, weak or strong) in the whole specimen (SI). Mean EGFR LI and PS% values were 30.4 +/- 3.5% and 51.6 +/- 3.9%, respectively. Patients with higher EGFR expression (EGFR LI, PS%, SI) were significantly younger than those with low EGFR expression. EGFR LI was higher in pT3 tumours than in pT1+pT2 tumours, moreover, EGFR expression (EGFR LI, PS%, SI) was significantly higher in G1+G2 tumours than in G3 tumours. There were significant correlations between parameters used for assessment of EGFR expression. PS% < or = 50 indicated shorter disease-specific survival than PS% > 50. However, patients with tumours with both very low and very high EGFR LI (13% > or = EGFR LI > 80%) showed significantly shorter survival than those with medium EGFR LI (13% < GFR LI < or = 80%). Additionally, pTNM and pN significantly influenced patients' survival. In multivariate analysis, EGFR LI and pTNM were independent prognostic parameters influencing disease-specific survival of patients.

Phosphorylation of hepatocyte growth factor receptor and epidermal growth factor receptor of human hepatocytes can be maintained in a (3D) collagen sandwich culture system.

PubMed

Engl, Tobias; Boost, Kim A; Leckel, Kerstin; Beecken, Wolf-Dietrich; Jonas, Dietger; Oppermann, Elsie; Auth, Marcus K H; Schaudt, André; Bechstein, Wolf-Otto; Blaheta, Roman A

2004-08-01

In vitro culture models that employ human liver cells could be potent tools for predictive studies on drug toxicity and metabolism in the pharmaceutical industry. However, an adequate receptor responsiveness is necessary to allow intracellular signalling and metabolic activity. We tested the ability of three-dimensionally arranged human hepatocytes to respond to the growth factors hepatocyte growth factor (HGF) or epidermal growth factor (EGF). Isolated adult human hepatocytes were cultivated within a three-dimensional collagen gel (sandwich) or on a two-dimensional collagen matrix. Cells were treated with HGF or EGF and expression and phosphorylative activity of HGF receptors (HGFr, c-met) or EGF receptors (EGFr) were measured by flow cytometry and Western blot. Increasing HGFr and EGFr levels were detected in hepatocytes growing two-dimensionally. However, both receptors were not activated in presence of growth factors. In contrast, when hepatocytes were plated within a three-dimensional matrix, HGFr and EGFr levels remained constantly low. However, both receptors became strongly phosphorylated by soluble HGF or EGF. We conclude that cultivation of human hepatocytes in a three-dimensionally arranged in vitro system allows the maintenance of specific functional activities. The necessity of cell dimensionality for HGFr and EGFr function should be considered when an adequate in vitro system has to be introduced for drug testing.

Combined caveolin-1 and epidermal growth factor receptor expression as a prognostic marker for breast cancer.

PubMed

Liang, Ya-Nan; Liu, Yu; Wang, Letian; Yao, Guodong; Li, Xiaobo; Meng, Xiangning; Wang, Fan; Li, Ming; Tong, Dandan; Geng, Jingshu

2018-06-01

Previous studies have indicated that caveolin-1 (Cav-1) is able to bind the signal transduction factor epidermal growth factor receptor (EGFR) to regulate its tyrosine kinase activity. The aim of the present study was to evaluate the clinical significance of Cav-1 gene expression in association with the expression of EGFR in patients with breast cancer. Primary breast cancer samples from 306 patients were analyzed for Cav-1 and EGFR expression using immunohistochemistry, and clinical significance was assessed using multivariate Cox regression analysis, Kaplan-Meier estimator curves and the log-rank test. Stromal Cav-1 was downregulated in 38.56% (118/306) of tumor tissues, whereas cytoplasmic EGFR and Cav-1 were overexpressed in 53.92% (165/306) and 44.12% (135/306) of breast cancer tissues, respectively. EGFR expression was positively associated with cytoplasmic Cav-1 and not associated with stromal Cav-1 expression in breast cancer samples; however, low expression of stromal Cav-1 was negatively associated with cytoplasmic Cav-1 expression in total tumor tissues, and analogous results were identified in the chemotherapy group. Multivariate Cox's proportional hazards model analysis revealed that, for patients in the estrogen receptor (ER)(+) group, the expression of stromal Cav-1 alone was a significant prognostic marker of breast cancer. However, in the chemotherapy, human epidermal growth factor receptor 2 (HER-2)(-), HER-2(+) and ER(-) groups, the use of combined markers was more effective prognostic marker. Stromal Cav-1 has a tumor suppressor function, and the combined marker stromal Cav-1/EGFR expression was identified as an improved prognostic marker in the diagnosis of breast cancer. Parenchymal expression of Cav-1 is able to promote EGFR signaling in breast cancer, potentially being required for EGFR-mediated initiation of mitosis.

Fluid shear stress regulates vascular remodeling via VEGFR-3 activation, although independently of its ligand, VEGF-C, in the uterus during pregnancy

PubMed Central

Park, Yang-Gyu; Choi, Jawun; Jung, Hye-Kang; Song, In Kyu; Shin, Yongwhan; Park, Sang-Youel; Seol, Jae-Won

2017-01-01

Early pregnancy is characterized by an increase in the blood volume of the uterus for embryonic development, thereby exerting fluid shear stress (FSS) on the vascular walls. The uterus experiences vascular remodeling to accommodate the increased blood flow. The blood flow-induced FSS elevates the expression of vascular endothelial growth factors (VEGFs) and their receptors, and regulates vascular remodeling through the activation of VEGF receptor-3 (VEGFR-3). However, the mechanisms responsible for FSS-induced VEGFR-3 expression in the uterus during pregnancy are unclear. In this study, we demonstrate that vascular remodeling in the uterus during pregnancy is regulated by FSS-induced VEGFR-3 expression. We examined the association between VEGFR-3 and FSS through in vivo and in vitro experiments. In vivo experiments revealed VEGFR-3 expression in the CD31-positive region of the uterus of pregnant mice; VEGF-C (ligand for VEGFR-3) was undetected in the uterus. These results confirmed that VEGFR-3 expression in the endometrium is independent of its ligand. In vitro studies experiments revealed that FSS induced morphological changes and increased VEGFR-3 expression in human uterine microvascular endothelial cells. Thus, VEGFR-3 activation by FSS is associated with vascular remodeling to allow increased blood flow in the uterus during pregnancy. PMID:28849193

Fluid shear stress regulates vascular remodeling via VEGFR-3 activation, although independently of its ligand, VEGF-C, in the uterus during pregnancy.

PubMed

Park, Yang-Gyu; Choi, Jawun; Jung, Hye-Kang; Song, In Kyu; Shin, Yongwhan; Park, Sang-Youel; Seol, Jae-Won

2017-10-01

Early pregnancy is characterized by an increase in the blood volume of the uterus for embryonic development, thereby exerting fluid shear stress (FSS) on the vascular walls. The uterus experiences vascular remodeling to accommodate the increased blood flow. The blood flow‑induced FSS elevates the expression of vascular endothelial growth factors (VEGFs) and their receptors, and regulates vascular remodeling through the activation of VEGF receptor-3 (VEGFR-3). However, the mechanisms responsible for FSS-induced VEGFR-3 expression in the uterus during pregnancy are unclear. In this study, we demonstrate that vascular remodeling in the uterus during pregnancy is regulated by FSS-induced VEGFR-3 expression. We examined the association between VEGFR-3 and FSS through in vivo and in vitro experiments. In vivo experiments revealed VEGFR-3 expression in the CD31-positive region of the uterus of pregnant mice; VEGF-C (ligand for VEGFR‑3) was undetected in the uterus. These results confirmed that VEGFR-3 expression in the endometrium is independent of its ligand. In vitro studies experiments revealed that FSS induced morphological changes and increased VEGFR-3 expression in human uterine microvascular endothelial cells. Thus, VEGFR-3 activation by FSS is associated with vascular remodeling to allow increased blood flow in the uterus during pregnancy.

Bradykinin-induced growth inhibition of normal rat kidney (NRK) cells is paralleled by a decrease in epidermal-growth-factor receptor expression.

PubMed Central

Van Zoelen, E J; Peters, P H; Afink, G B; Van Genesen, S; De Roos, D G; Van Rotterdam, W; Theuvenet, A P

1994-01-01

Normal rat kidney fibroblasts, grown to density arrest in the presence of epidermal growth factor (EGF), can be induced to undergo phenotypic transformation by treatment with transforming growth factor beta or retinoic acid. Here we show that bradykinin blocks this growth-stimulus-induced loss of density-dependent growth arrest by a specific receptor-mediated mechanism. The effects of bradykinin are specific, and are not mimicked by other phosphoinositide-mobilizing agents such as prostaglandin F2 alpha. Northern-blot analysis and receptor-binding studies demonstrate that bradykinin also inhibits the retinoic acid-induced increase in EGF receptor levels in these cells. These studies provide additional evidence that EGF receptor levels modulate EGF-induced expression of the transformed phenotype in these cells. Images Figure 5 PMID:8135739

Insulin-like growth factor 1 signaling is essential for mitochondrial biogenesis and mitophagy in cancer cells.

PubMed

Lyons, Amy; Coleman, Michael; Riis, Sarah; Favre, Cedric; O'Flanagan, Ciara H; Zhdanov, Alexander V; Papkovsky, Dmitri B; Hursting, Stephen D; O'Connor, Rosemary

2017-10-13

Mitochondrial activity and metabolic reprogramming influence the phenotype of cancer cells and resistance to targeted therapy. We previously established that an insulin-like growth factor 1 (IGF-1)-inducible mitochondrial UTP carrier (PNC1/SLC25A33) promotes cell growth. This prompted us to investigate whether IGF signaling is essential for mitochondrial maintenance in cancer cells and whether this contributes to therapy resistance. Here we show that IGF-1 stimulates mitochondrial biogenesis in a range of cell lines. In MCF-7 and ZR75.1 breast cancer cells, IGF-1 induces peroxisome proliferator-activated receptor γ coactivator 1β (PGC-1β) and PGC-1α-related coactivator (PRC). Suppression of PGC-1β and PRC with siRNA reverses the effects of IGF-1 and disrupts mitochondrial morphology and membrane potential. IGF-1 also induced expression of the redox regulator nuclear factor-erythroid-derived 2-like 2 (NFE2L2 alias NRF-2). Of note, MCF-7 cells with acquired resistance to an IGF-1 receptor (IGF-1R) tyrosine kinase inhibitor exhibited reduced expression of PGC-1β, PRC, and mitochondrial biogenesis. Interestingly, these cells exhibited mitochondrial dysfunction, indicated by reactive oxygen species expression, reduced expression of the mitophagy mediators BNIP3 and BNIP3L, and impaired mitophagy. In agreement with this, IGF-1 robustly induced BNIP3 accumulation in mitochondria. Other active receptor tyrosine kinases could not compensate for reduced IGF-1R activity in mitochondrial protection, and MCF-7 cells with suppressed IGF-1R activity became highly dependent on glycolysis for survival. We conclude that IGF-1 signaling is essential for sustaining cancer cell viability by stimulating both mitochondrial biogenesis and turnover through BNIP3 induction. This core mitochondrial protective signal is likely to strongly influence responses to therapy and the phenotypic evolution of cancer. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

AhR mediates an anti-inflammatory feedback mechanism in human Langerhans cells involving FcεRI and IDO.

PubMed

Koch, S; Stroisch, T J; Vorac, J; Herrmann, N; Leib, N; Schnautz, S; Kirins, H; Förster, I; Weighardt, H; Bieber, T

2017-11-01

Aryl hydrocarbon receptor (AhR), an important regulator of immune responses, is activated by UVB irradiation in the skin. Langerhans cells (LC) in the epidermis of patients with atopic dermatitis (AD) carry the high-affinity receptor for IgE, FcεRI, and are crucially involved in the pathogenesis of AD by inducing inflammatory responses and regulating tolerogenic processes. We investigated AhR and AhR repressor (AhRR) expression and functional consequences of AhR activation in human ex vivo skin cells and in in vitro-generated LC. Epidermal cells from healthy skin were analyzed for their expression of AhR and AhRR. LC generated from CD34 + hematopoietic stem cells (CD34LC) were treated with the UV photoproduct and AhR ligand 6-formylindolo[3,2-b]carbazole (FICZ). Cell surface receptors, transcription factors, and the tolerogenic tryptophan-degrading enzyme indoleamine 2,3-dioxygenase (IDO) were analyzed using flow cytometry and quantitative PCR. Epidermal LC and CD34LC express AhR and AhRR. AhR was also found in keratinocytes, which lack AhRR. AhR activation of LC by FICZ caused downregulation of FcεRI in CD34LC without affecting their maturation. AhR-mediated regulation of FcεRI did not involve any known transcription factors related to this receptor. Furthermore, we could show upregulation of IDO mediated by AhR engagement. Our study shows that AhR activation by FICZ reduces FcεRI and upregulates IDO expression in LC. This AhR-mediated anti-inflammatory feedback mechanism may dampen the allergen-induced inflammation in AD. © 2017 EAACI and John Wiley and Sons A/S. Published by John Wiley and Sons Ltd.

Evaluation of the osteoclastogenic process associated with RANK / RANK-L / OPG in odontogenic myxomas

PubMed Central

González-Galván, María del Carmen; Mosqueda-Taylor, Adalberto; Bologna-Molina, Ronell; Setien-Olarra, Amaia; Marichalar-Mendia, Xabier; Aguirre-Urizar, José-Manuel

2018-01-01

Background Odontogenic myxoma (OM) is a benign intraosseous neoplasm that exhibits local aggressiveness and high recurrence rates. Osteoclastogenesis is an important phenomenon in the tumor growth of maxillary neoplasms. RANK (Receptor Activator of Nuclear Factor κappa B) is the signaling receptor of RANK-L (Receptor activator of nuclear factor kappa-Β ligand) that activates the osteoclasts. OPG (osteoprotegerin) is a decoy receptor for RANK-L that inhibits pro-osteoclastogenesis. The RANK / RANKL / OPG system participates in the regulation of osteolytic activity under normal conditions, and its alteration has been associated with greater bone destruction, and also with tumor growth. Objectives To analyze the immunohistochemical expression of OPG, RANK and RANK-L proteins in odontogenic myxomas (OMs) and their relationship with the tumor size. Material and Methods Eighteen OMs, 4 small (<3 cm) and 14 large (> 3cm) and 18 dental follicles (DF) that were included as control were studied by means of standard immunohistochemical procedure with RANK, RANKL and OPG antibodies. For the evaluation, 5 fields (40x) of representative areas of OM and DF were selected where the expression of each antibody was determined. Descriptive and comparative statistical analyses were performed with the obtained data. Results There are significant differences in the expression of RANK in OM samples as compared to DF (p = 0.022) and among the OMSs and OMLs (p = 0.032). Also a strong association is recognized in the expression of RANK-L and OPG in OM samples. Conclusions Activation of the RANK / RANK-L / OPG triad seems to be involved in the mechanisms of bone balance and destruction, as well as associated with tumor growth in odontogenic myxomas. Key words:Odontogenic myxoma, dental follicle, RANK, RANK-L, OPG, osteoclastogenesis. PMID:29680857

GATA-3 and FOXA1 expression is useful to differentiate breast carcinoma from other carcinomas.

PubMed

Davis, Drew G; Siddiqui, Momin T; Oprea-Ilies, Gabriela; Stevens, Keith; Osunkoya, Adeboye O; Cohen, Cynthia; Li, Xiaoxian Bill

2016-01-01

GATA-3, a member of the GATA family of zinc-finger DNA binding proteins, and FOXA1, a member of the forkhead transcription factor family, are both associated with estrogen receptor expression. Both GATA-3 and FOXA1 are useful markers for breast carcinoma, but their expression in the different breast cancer subtypes and other neoplasms has not been thoroughly evaluated. We examined the expression of GATA-3 and FOXA1 in estrogen receptor-positive, Her2/neu-positive, and triple-negative breast carcinomas as well as in 10 other common carcinomas, including hepatocellular, colonic, pancreatic, gastric, endometrial (endometrioid), lung, prostatic, renal cell, urothelial, and ovarian serous carcinomas. Primary and metastatic melanomas and mesotheliomas were also evaluated. GATA-3 and FOXA1 staining of estrogen receptor-positive breast carcinomas was seen in 96.6% and 96.2%, respectively. In triple-negative breast carcinomas, GATA-3 and FOXA1 staining was seen in 21.6% and 15.9%, respectively. Among the other tumors, GATA-3 staining was only seen in urothelial carcinoma (70.9%) and FOXA1 staining was only seen in prostatic (87.5%), urothelial (5.1%) carcinomas, and mesotheliomas (40.0%). In conclusion, GATA-3 and FOXA1 are excellent breast carcinoma markers; however, their utility is limited in the triple-negative subtype. The utility of FOXA1 in diagnosing prostatic carcinoma and mesothelioma warrants further investigation. Copyright © 2015 Elsevier Inc. All rights reserved.

Expression changes of serotonin receptor gene subtype 5HT3a in peripheral blood mononuclear cells from schizophrenic patients treated with haloperidol and Olanzapin.

PubMed

Shariati, Gholam Reza; Ahangari, Ghasem; Hossein-nezhad, Arash; Asadi, Seyed Mohammad; Pooyafard, Farzaneh; Ahmadkhaniha, Hamid Reza

2009-09-01

Serotonin receptors are involved in pathophysiology of schizophrenia and may mediate other neurotransmitter effects. We investigated serotonin receptors gene expression in peripheral blood mononuclear cells (PBMC) of naïve schizophrenic patients, before and after treatment. Also serotonin receptor gene expression was compared in two treatment groups including Haloperidol and Olanzapine. The PBMC was separated from whole blood by Ficoll-hypaque. The total cellular RNA was extracted and the cDNA was synthesized. This process was followed by real-time PCR using primer pairs specific for 5HT(3a) serotonin receptor mRNA and beta-actin as internal control. The results showed the presence of subtype of serotonin receptor in lymphocytes. Serotonin gene expression showed significant changes in Olanzapine treatment group which correlated with Clinical Global Impression (CGI) score improvement. In conclusion, the present study has shown that human PBMC express serotonin receptors 5HT(3a). Moreover, clinical symptom improvement of Olanzapin may be demonstrated by a change in serotonin receptor gene expression.

Mechanisms of resistance to anti-human epidermal growth factor receptor 2 agents in breast cancer.

PubMed

Mukohara, Toru

2011-01-01

Approximately 20% of breast cancers are characterized by overexpression of human epidermal growth factor receptor 2 (HER2) protein and associated gene amplification, and the receptor tyrosine kinase is believed to play a critical role in the pathogenesis of these tumors. The development and implementation of trastuzumab, a humanized monoclonal antibody against the extracellular domain of HER2 protein, has significantly improved treatment outcomes in patients with HER2-overexpressing breast cancer. However, despite this clinical usefulness, unmet needs for better prediction of trastuzumab's response and overcoming primary and acquired resistance remain. In this review, we discuss several potential mechanisms of resistance to trastuzumab that have been closely studied over the last decade. Briefly, these mechanisms include: impaired access of trastuzumab to HER2 by expression of extracellular domain-truncated HER2 (p95 HER2) or overexpression of MUC4; alternative signaling from insulin-like growth factor-1 receptor, other epidermal growth factor receptor family members, or MET; aberrant downstream signaling caused by loss of phosphatase and tensin homologs deleted from chromosome 10 (PTEN), PIK3CA mutation, or downregulation of p27; or FCGR3A polymorphisms. In addition, we discuss potential strategies for overcoming resistance to trastuzumab. Specifically, the epidermal growth factor receptor/HER2 tyrosine kinase inhibitor lapatinib partially overcame trastuzumab resistance in a clinical setting, so its efficacy results and limited data regarding potential mechanisms of resistance to the drug are also discussed. © 2010 Japanese Cancer Association.

The combination of decoy receptor 3 and soluble triggering receptor expressed on myeloid cells-1 for the diagnosis of nosocomial bacterial meningitis.

PubMed

Liu, Yong-Juan; Shao, Li-Hua; Zhang, Jian; Fu, Shan-Ji; Wang, Gang; Chen, Feng-Zhe; Zheng, Feng; Ma, Rui-Ping; Liu, Hai-Hong; Dong, Xiao-Meng; Ma, Li-Xian

2015-03-23

Early diagnosis and appropriate antibiotic treatment can significantly reduce mortality of nosocomial bacterial meningitis. However, it is a challenge for clinicians to make an accurate and rapid diagnosis of bacterial meningitis. This study aimed at determining whether combined biomarkers can provide a useful tool for the diagnosis of bacterial meningitis. A retrospective study was carried out. Cerebrospinal fluid (CSF) levels of decoy receptor 3 (DcR3) and soluble triggering receptor expressed on myeloid cells-1 (sTREM-1) were detected by enzyme-linked immunosorbent assay (ELISA). The patients with bacterial meningitis had significantly elevated levels of the above mentioned biomarkers. The two biomarkers were all risk factors with bacterial meningitis. The biomarkers were constructed into a "bioscore". The discriminative performance of the bioscore was better than that of each biomarker, with an area under the receiver operating characteristic (ROC) curve (AUC) of 0.842 (95% confidence intervals (CI) 0.770-0.914; p< 0.001). Combined measurement of CSF DcR3 and sTREM-1 concentrations improved the prediction of nosocomial bacterial meningitis. The combined strategy is of interest and the validation of that improvement needs further studies.

Developmental Programming: Does Prenatal Steroid Excess Disrupt the Ovarian VEGF System in Sheep?1

PubMed Central

Ortega, Hugo Héctor; Veiga-Lopez, Almudena; Sreedharan, Shilpa; del Luján Velázquez, Melisa María; Salvetti, Natalia Raquel; Padmanabhan, Vasantha

2015-01-01

Prenatal testosterone (T), but not dihydrotestosterone (DHT), excess disrupts ovarian cyclicity and increases follicular recruitment and persistence. We hypothesized that the disruption in the vascular endothelial growth factor (VEGF) system contributes to the enhancement of follicular recruitment and persistence in prenatal T-treated sheep. The impact of T/DHT treatments from Days 30 to 90 of gestation on VEGFA, VEGFB, and their receptor (VEGFR-1 [FLT1], VEGFR-2 [KDR], and VEGFR-3 [FLT4]) protein expression was examined by immunohistochemistry on Fetal Days 90 and 140, 22 wk, 10 mo (postpubertal), and 21 mo (adult) of age. Arterial morphometry was performed in Fetal Day 140 and postpubertal ovaries. VEGFA and VEGFB expression were found in granulosa cells at all stages of follicular development with increased expression in antral follicles. VEGFA was present in theca interna, while VEGFB was present in theca interna/externa and stromal cells. All three receptors were expressed in the granulosa, theca, and stromal cells during all stages of follicular development. VEGFR-3 increased with follicular differentiation with the highest level seen in the granulosa cells of antral follicles. None of the members of the VEGF family or their receptor expression were altered by age or prenatal T/DHT treatments. At Fetal Day 140, area, wall thickness, and wall area of arteries from the ovarian hilum were larger in prenatal T- and DHT-treated females, suggestive of early androgenic programming of arterial differentiation. This may facilitate increased delivery of endocrine factors and thus indirectly contribute to the development of the multifollicular phenotype. PMID:26178718

Granulocyte-macrophage and macrophage colony-stimulating factors differentially regulate alpha v integrin expression on cultured human macrophages.

PubMed

De Nichilo, M O; Burns, G F

1993-03-15

The colony-stimulating factors (CSFs) greatly influence mature macrophage function in vitro: macrophage (M)-CSF induces maturation of monocytes and enhances differentiated cell function; granulocyte-macrophage (GM)-CSF stimulates a variety of antimicrobial functions. In vivo M-CSF is thought to promote differentiation, and GM-CSF is thought to potentiate the inflammatory response. One mechanism by which these differential effects may be achieved is through the receptor-mediated interaction of macrophages with their extracellular matrix. Here we show that M-CSF induces specifically the expression of the alpha v beta 5 integrin receptor, whereas GM-CSF rapidly induces mRNA and surface expression of the alpha v beta 3 integrin. The M-CSF-treated cells acquire a flattened epitheloid phenotype, and on vitronectin the alpha v beta 5 is located in adhesion plaques. These cells do not bind collagen or laminin. In contrast, cells treated with GM-CSF adopt an elongated phenotype on a number of substrates, including collagen and laminin, and express alpha v beta 3 at the leading edge of cells on vitronectin. These results suggest that a primary means by which the CSFs exert their individual effects on mature cells may be through regulating integrin expression.

ARF6 Activated by the LHCG Receptor through the Cytohesin Family of Guanine Nucleotide Exchange Factors Mediates the Receptor Internalization and Signaling*

PubMed Central

Kanamarlapudi, Venkateswarlu; Thompson, Aiysha; Kelly, Eamonn; López Bernal, Andrés

2012-01-01

The luteinizing hormone chorionic gonadotropin receptor (LHCGR) is a Gs-coupled GPCR that is essential for the maturation and function of the ovary and testis. LHCGR is internalized following its activation, which regulates the biological responsiveness of the receptor. Previous studies indicated that ADP-ribosylation factor (ARF)6 and its GTP-exchange factor (GEF) cytohesin 2 regulate LHCGR internalization in follicular membranes. However, the mechanisms by which ARF6 and cytohesin 2 regulate LHCGR internalization remain incompletely understood. Here we investigated the role of the ARF6 signaling pathway in the internalization of heterologously expressed human LHCGR (HLHCGR) in intact cells using a combination of pharmacological inhibitors, siRNA and the expression of mutant proteins. We found that human CG (HCG)-induced HLHCGR internalization, cAMP accumulation and ARF6 activation were inhibited by Gallein (βγ inhibitor), Wortmannin (PI 3-kinase inhibitor), SecinH3 (cytohesin ARF GEF inhibitor), QS11 (an ARF GAP inhibitor), an ARF6 inhibitory peptide and ARF6 siRNA. However, Dynasore (dynamin inhibitor), the dominant negative mutants of NM23-H1 (dynamin activator) and clathrin, and PBP10 (PtdIns 4,5-P2-binding peptide) inhibited agonist-induced HLHCGR and cAMP accumulation but not ARF6 activation. These results indicate that heterotrimeric G-protein, phosphatidylinositol (PI) 3-kinase (PI3K), cytohesin ARF GEF and ARF GAP function upstream of ARF6 whereas dynamin and clathrin act downstream of ARF6 in the regulation of HCG-induced HLHCGR internalization and signaling. In conclusion, we have identified the components and molecular details of the ARF6 signaling pathway required for agonist-induced HLHCGR internalization. PMID:22523074

Molecular identification and functional expression of mu 3, a novel alternatively spliced variant of the human mu opiate receptor gene.

PubMed

Cadet, Patrick; Mantione, Kirk J; Stefano, George B

2003-05-15

Studies from our laboratory have revealed a novel mu opiate receptor, mu 3, which is expressed in both vascular tissues and leukocytes. The mu 3 receptor is selective for opiate alkaloids and is insensitive to opioid peptides. We now identify the mu 3 receptor at the molecular level using a 441-bp conserved region of the mu 1 receptor. Sequence analysis of the isolated cDNA suggests that it is a novel, alternatively spliced variant of the mu opiate receptor gene. To determine whether protein expressed from this cDNA exhibits the biochemical characteristics expected of the mu 3 receptor, the cDNA clone was expressed in a heterologous system. At the functional level, COS-1 cells transfected with the mu 3 receptor cDNA exhibited dose-dependent release of NO following treatment with morphine, but not opioid peptides (i.e., Met-enkephalin). Naloxone was able to block the effect of morphine on COS-1 transfected cells. Nontransfected COS-1 cells did not produce NO in the presence of morphine or the opioid peptides at similar concentrations. Receptor binding analysis with [(3)H]dihydromorphine further supports the opiate alkaloid selectivity and opioid peptide insensitivity of this receptor. These data suggest that this new mu opiate receptor cDNA encodes the mu 3 opiate receptor, since it exhibits biochemical characteristics known to be unique to this receptor (opiate alkaloid selective and opioid peptide insensitive). Furthermore, using Northern blot, RT-PCR, and sequence analysis, we have demonstrated the expression of this new mu variant in human vascular tissue, mononuclear cells, polymorphonuclear cells, and human neuroblastoma cells.

c-erbA and v-erbA modulate growth and gene expression of a mouse glial precursor cell line.

PubMed

Iglesias, T; Llanos, S; López-Barahona, M; Pérez-Aranda, A; Rodríguez-Peña, A; Bernal, J; Höhne, A; Seliger, B; Muñoz, A

1994-07-01

The c-erbA alpha protooncogene coding for the thyroid hormone (T3) receptor (TR alpha 1) and the viral, mutated v-erbA oncogene were expressed in an immortal mouse glial cell line (B3.1) using retroviral vectors. c-erbA alpha expression led to a decrease in cell proliferation in high and low serum conditions, both in the presence and in the absence of T3. In serum-free medium, c-erbA-expressing cells (B3.1 + TR alpha 1) were completely arrested, whereas cells expressing v-erbA (B3.1 + v-erbA) showed a higher DNA synthesis rate than normal B3.1 cells. Although proliferation of all three cell types was stimulated by platelet-derived growth factor and basic fibroblast growth factor, differences were also observed in the response to these agents. B3.1 + TR alpha 1 cells were more sensitive to platelet-derived growth factor than B3.1 and B3.1 + v-erbA cells. In contrast, B3.1 cells responded to basic fibroblast growth factor better than B3.1 + TR alpha 1 or B3.1 + v-erbA cells. Insulin-like growth factor I potentiated the action of platelet-derived growth factor and basic fibroblast growth factor. Again, different responses to treatment with insulin-like growth factor I alone were observed; B3.1 + TR alpha 1 cells did not respond to it, whereas B3.1 + v-erbA cells showed a dramatic stimulation by this agent. Interestingly, in the presence of T3, the blockade in B3.1 + TR alpha 1 cell proliferation was accompanied by the down-regulation of the typical astrocytic genes, glial fibrillary acidic protein and vimentin. These hormone effects were not found in v-erbA-expressing cells. In addition, v-erbA inhibited the basal expression of the cyclic nucleotide phosphodiesterase gene, an oligodendrocytic marker.(ABSTRACT TRUNCATED AT 250 WORDS)

Effects of age and insulin-like growth factor-1 on rat neurotrophin receptor expression after nerve injury.

PubMed

Luo, T David; Alton, Timothy B; Apel, Peter J; Cai, Jiaozhong; Barnwell, Jonathan C; Sonntag, William E; Smith, Thomas L; Li, Zhongyu

2016-10-01

Neurotrophin receptors, such as p75(NTR) , direct neuronal response to injury. Insulin-like growth factor-1 receptor (IGF-1R) mediates the increase in p75(NTR) during aging. The aim of this study was to examine the effect of aging and insulin-like growth factor-1 (IGF-1) treatment on recovery after peripheral nerve injury. Young and aged rats underwent tibial nerve transection with either local saline or IGF-1 treatment. Neurotrophin receptor mRNA and protein expression were quantified. Aged rats expressed elevated baseline IGF-1R (34% higher, P = 0.01) and p75(NTR) (68% higher, P < 0.01) compared with young rats. Post-injury, aged animals expressed significantly higher p75(NTR) levels (68.5% above baseline at 4 weeks). IGF-1 treatment suppressed p75(NTR) gene expression at 4 weeks (17.2% above baseline, P = 0.002) post-injury. Local IGF-1 treatment reverses age-related declines in recovery after peripheral nerve injuries by suppressing p75(NTR) upregulation and pro-apoptotic complexes. IGF-1 may be considered a viable adjuvant therapy to current treatment modalities. Muscle Nerve 54: 769-775, 2016. © 2016 Wiley Periodicals, Inc.

The chemokine receptor CXCR3 and its splice variant are expressed in human airway epithelial cells.

PubMed

Kelsen, Steven G; Aksoy, Mark O; Yang, Yi; Shahabuddin, Syed; Litvin, Judith; Safadi, Fayez; Rogers, Thomas J

2004-09-01

Activation of the chemokine receptor CXCR3 by its cognate ligands induces several differentiated cellular responses important to the growth and migration of a variety of hematopoietic and structural cells. In the human respiratory tract, human airway epithelial cells (HAEC) release the CXCR3 ligands Mig/CXCL9, IP-10/CXCL10, and I-TAC/CXCL11. Simultaneous expression of CXCR3 by HAEC would have important implications for the processes of airway inflammation and repair. Accordingly, in the present study we sought to determine whether HAEC also express the classic CXCR3 chemokine receptor CXCR3-A and its splice variant CXCR3-B and hence may respond in autocrine fashion to its ligands. We found that cultured HAEC (16-HBE and tracheocytes) constitutively expressed CXCR3 mRNA and protein. CXCR3 mRNA levels assessed by expression array were approximately 35% of beta-actin expression. In contrast, CCR3, CCR4, CCR5, CCR8, and CX3CR1 were <5% beta-actin. Both CXCR3-A and -B were expressed. Furthermore, tracheocytes freshly harvested by bronchoscopy stained positively for CXCR3 by immunofluorescence microscopy, and 68% of cytokeratin-positive tracheocytes (i.e., the epithelial cell population) were positive for CXCR3 by flow cytometry. In 16-HBE cells, CXCR3 receptor density was approximately 78,000 receptors/cell when assessed by competitive displacement of 125I-labeled IP-10/CXCL10. Finally, CXCR3 ligands induced chemotactic responses and actin reorganization in 16-HBE cells. These findings indicate constitutive expression by HAEC of a functional CXC chemokine receptor, CXCR3. Our data suggest the possibility that autocrine activation of CXCR3 expressed by HAEC may contribute to airway inflammation and remodeling in obstructive lung disease by regulating HAEC migration.

Cellular density-dependent down-regulation of EP4 prostanoid receptors via the up-regulation of hypoxia-inducible factor-1α in HCA-7 human colon cancer cells.

PubMed

Otake, Sho; Yoshida, Kenji; Seira, Naofumi; Sanchez, Christopher M; Regan, John W; Fujino, Hiromichi; Murayama, Toshihiko

2015-02-01

Increases in prostaglandin E2 (PGE2) and cyclooxygenase-2 (COX-2) levels are features of colon cancer. Among the different E-type prostanoid receptor subtypes, EP4 receptors are considered to play a crucial role in carcinogenesis by, for example, inducing COX-2 when stimulated with PGE2. However, EP4 receptor levels and PGE2-induced cellular responses are inconsistent among the cellular conditions. Therefore, the connections responsible for the expression of EP4 receptors were investigated in the present study by focusing on cell density-induced hypoxia-inducible factor-1α (HIF-1α). The expression of EP4 receptors was examined using immunoblot analysis, quantitative polymerase chain reaction, and reporter gene assays in HCA-7 human colon cancer cells with different cellular densities. The involvement of HIF-1α and its signaling pathways were also examined by immunoblot analysis, reporter gene assays, and with siRNA. We here demonstrated that EP4 receptors as well as EP4 receptor-mediated COX-2 expression levels decreased with an increase in cellular density. In contrast, HIF-1α levels increased in a cellular density-dependent manner. The knockdown of HIF-1α by siRNA restored the expression of EP4 receptors and EP4 receptor-mediated COX-2 in cells at a high density. Thus, the cellular density-dependent increase observed in HIF-1α expression levels reduced the expression of COX-2 by decreasing EP4 receptor levels. This novel regulation mechanism for the expression of EP4 receptors by HIF-1α may provide an explanation for the inconsistent actions of PGE2. The expression levels of EP4 receptors may vary depending on cellular density, which may lead to the differential activation of their signaling pathways by PGE2. Thus, cellular density-dependent PGE2-mediated signaling may determine the fate/stage of cancer cells, i.e., the surrounding environments could define the fate/stage of malignancies associated with colon cancer.

Cellular density-dependent down-regulation of EP4 prostanoid receptors via the up-regulation of hypoxia-inducible factor-1α in HCA-7 human colon cancer cells

PubMed Central

Otake, Sho; Yoshida, Kenji; Seira, Naofumi; Sanchez, Christopher M; Regan, John W; Fujino, Hiromichi; Murayama, Toshihiko

2015-01-01

Increases in prostaglandin E2 (PGE2) and cyclooxygenase-2 (COX-2) levels are features of colon cancer. Among the different E-type prostanoid receptor subtypes, EP4 receptors are considered to play a crucial role in carcinogenesis by, for example, inducing COX-2 when stimulated with PGE2. However, EP4 receptor levels and PGE2-induced cellular responses are inconsistent among the cellular conditions. Therefore, the connections responsible for the expression of EP4 receptors were investigated in the present study by focusing on cell density-induced hypoxia-inducible factor-1α (HIF-1α). The expression of EP4 receptors was examined using immunoblot analysis, quantitative polymerase chain reaction, and reporter gene assays in HCA-7 human colon cancer cells with different cellular densities. The involvement of HIF-1α and its signaling pathways were also examined by immunoblot analysis, reporter gene assays, and with siRNA. We here demonstrated that EP4 receptors as well as EP4 receptor-mediated COX-2 expression levels decreased with an increase in cellular density. In contrast, HIF-1α levels increased in a cellular density-dependent manner. The knockdown of HIF-1α by siRNA restored the expression of EP4 receptors and EP4 receptor-mediated COX-2 in cells at a high density. Thus, the cellular density-dependent increase observed in HIF-1α expression levels reduced the expression of COX-2 by decreasing EP4 receptor levels. This novel regulation mechanism for the expression of EP4 receptors by HIF-1α may provide an explanation for the inconsistent actions of PGE2. The expression levels of EP4 receptors may vary depending on cellular density, which may lead to the differential activation of their signaling pathways by PGE2. Thus, cellular density-dependent PGE2-mediated signaling may determine the fate/stage of cancer cells, i.e., the surrounding environments could define the fate/stage of malignancies associated with colon cancer. PMID:25692008

Modulation of tumor necrosis factor (TNF) receptor expression during monocytic differentiation by glucocorticoids.

PubMed

Goppelt-Struebe, M; Reiser, C O; Schneider, N; Grell, M

1996-10-01

Regulation of tumor necrosis factor receptors by glucocorticoids was investigated during phorbol ester-induced monocytic differentiation. As model system the human monocytic cell lines U937 and THP-1, which express both types of TNF receptors (TNF-R60 and TNF-R80), were differentiated with tetradecanoyl phorbol-13-acetate (TPA, 5 x 10(-9) M) in the presence or absence of dexamethasone (10(-9) - 10(-6) M). Expression of TNF receptors was determined at the mRNA level by Northern blot analysis and at the protein level by FACS analysis. During differentiation, TNF-R60 mRNA was down-regulated, whereas TNF-R80 mRNA levels were increased. Dexamethasone had no effect on TNF-R60 mRNA expression but attenuated TNF-R80 mRNA expression in both cell lines. Cell surface expression of TNF-R60 protein remained essentially unchanged during differentiation of THP-1 cells, whereas a rapid down-regulation of TNF-R80 was observed that was followed by a slow recovery. Surface expression of TNF-R80 was not affected by dexamethasone, whereas TNF-R60 expression was reduced by about 25%. These results indicate differential regulation of the two types of TNF receptors at the mRNA and protein level during monocytic differentiation. Glucocorticoids interfered with mRNA expression of TNF-R80 and protein expression of TNF-R60, but the rather limited effect leaves the question of its functional relevance open. In contrast to other cytokine systems, TNF receptors do not appear to be major targets of glucocorticoid action.

Identification of SR1078, a synthetic agonist for the orphan nuclear receptors RORα and RORγ.

PubMed

Wang, Yongjun; Kumar, Naresh; Nuhant, Philippe; Cameron, Michael D; Istrate, Monica A; Roush, William R; Griffin, Patrick R; Burris, Thomas P

2010-11-19

The retinoic acid receptor-related receptors (RORs) are members of the nuclear receptor (NR) superfamily of transcription factors. Several NRs are still characterized as orphan receptors because ligands have not yet been identified for these proteins. Here, we describe the identification of a synthetic RORα/RORγ ligand, SR1078. SR1078 modulates the conformation of RORγ in a biochemical assay and activates RORα and RORγ driven transcription. Furthermore, SR1078 stimulates expression of endogenous ROR target genes in HepG2 cells that express both RORα and RORγ. Pharmacokinetic studies indicate that SR1078 displays reasonable exposure following injection into mice, and consistent with SR1078 functioning as a RORα/RORγ agonist, expression of two ROR target genes, glucose-6-phosphatase and fibroblast growth factor 21, were stimulated in the liver. Thus, we have identified the first synthetic RORα/γ agonist, and this compound can be utilized as a chemical tool to probe the function of these receptors both in vitro and in vivo.

Identification of a Synthetic Agonist for the Orphan Nuclear Receptors RORα and RORγ, SR1078

PubMed Central

Wang, Yongjun; Kumar, Naresh; Nuhant, Philippe; Cameron, Michael D.; Istrate, Monica A.; Roush, William R.; Griffin, Patrick R.; Burris, Thomas P.

2010-01-01

The retinoic acid receptor-related receptors (RORs) are members of the nuclear receptor (NR) superfamily of transcription factors. Several NRs are still characterized as orphan receptors since ligands have not yet been identified for these proteins. Here, we describe the identification of a synthetic RORα/RORγ ligand, SR1078. SR1078 modulates the conformation of RORγ in a biochemical assay and activates RORα and RORγ driven transcription. Furthermore, SR1078 stimulates expression of endogenous ROR target genes in HepG2 cells that express both RORα and RORγ. Pharmacokinetic studies indicate that SR1078 displays reasonable exposure following injection into mice and consistent with SR1078 functioning as a RORα/RORγ agonist, expression of two ROR target genes, glucose-6-phosphatase and fibroblast growth factor 21, were stimulated in the liver. Thus, we have identified the first synthetic RORα/γ agonist and this compound can be utilized as a chemical tool to probe the function of these receptors both in vitro and in vivo. PMID:20735016

Supplementation of conventional freezing medium with a combination of catalase and trehalose results in better protection of surface molecules and functionality of hematopoietic cells.

PubMed

Sasnoor, Lalita M; Kale, Vaijayanti P; Limaye, Lalita S

2003-10-01

Our previous studies had shown that a combination of the bio-antioxidant catalase and the membrane stabilizer trehalose in the conventional freezing mixture affords better cryoprotection to hematopoietic cells as judged by clonogenic assays. In the present investigation, we extended these studies using several parameters like responsiveness to growth factors, expression of growth factor receptors, adhesion assays, adhesion molecule expression, and long-term culture-forming ability. Cells were frozen with (test cells) or without additives (control cells) in the conventional medium containing 10% dimethylsulfoxide (DMSO). Experiments were done on mononuclear cells (MNC) from cord blood/fetal liver hematopoietic cells (CB/FL) and CD34(+) cells isolated from frozen MNC. Our results showed that the responsiveness of test cells to the two early-acting cytokines, viz. interleukin-3 (IL-3) and stem cell factor (SCF) in CFU assays was better than control cells as seen by higher colony formation at limiting concentrations of these cytokines. We, therefore, analyzed the expression of these two growth factor receptors by flow cytometry. We found that in cryopreserved test MNC, as well as CD34(+) cells isolated from them, the expression of both cytokine receptors was two- to three-fold higher than control MNC and CD34(+) cells isolated from them. Adhesion assays carried out with CB/FL-derived CD34(+) cells and KG1a cells showed significantly higher adherence of test cells to M210B4 than respective control cells. Cryopreserved test MNC as well as CD34(+) cells isolated from them showed increased expression of adhesion molecules like CD43, CD44, CD49d, and CD49e. On isolated CD34(+) cells and KG1a cells, there was a two- to three-fold increase in a double-positive population expressing CD34/L-selectin in test cells as compared to control cells. Long-term cultures (LTC) were set up with frozen MNC as well as with CD34(+) cells. Clonogenic cells from LTC were enumerated at the end of the fifth week. There was a significantly increased formation of CFU from test cells than from control cells, indicating better preservation of early progenitors in test cells. Our results suggest that use of a combination of catalase and trehalose as a supplement in the conventional freezing medium results in better protection of growth factor receptors, adhesion molecules, and functionality of hematopoietic cells, yielding a better graft quality.

Pattern-Recognition Receptor Signaling Regulator mRNA Expression in Humans and Mice, and in Transient Inflammation or Progressive Fibrosis

PubMed Central

Günthner, Roman; Kumar, Vankayala Ramaiah Santhosh; Lorenz, Georg; Anders, Hans-Joachim; Lech, Maciej

2013-01-01

The cell type-, organ-, and species-specific expression of the pattern-recognition receptors (PRRs) are well described but little is known about the respective expression profiles of their negative regulators. We therefore determined the mRNA expression levels of A20, CYLD, DUBA, ST2, CD180, SIGIRR, TANK, SOCS1, SOCS3, SHIP, IRAK-M, DOK1, DOK2, SHP1, SHP2, TOLLIP, IRF4, SIKE, NLRX1, ERBIN, CENTB1, and Clec4a2 in human and mouse solid organs. Humans and mice displayed significant differences between their respective mRNA expression patterns of these factors. Additionally, we characterized their expression profiles in mononuclear blood cells upon bacterial endotoxin, which showed a consistent induction of A20, SOCS3, IRAK-M, and Clec4a2 in human and murine cells. Furthermore, we studied the expression pattern in transient kidney ischemia-reperfusion injury versus post-ischemic atrophy and fibrosis in mice. A20, CD180, ST2, SOCS1, SOCS3, SHIP, IRAK-M, DOK1, DOK2, IRF4, CENTB1, and Clec4a2 were all induced, albeit at different times of injury and repair. Progressive fibrosis was associated with a persistent induction of these factors. Thus, the organ- and species-specific expression patterns need to be considered in the design and interpretation of studies related to PRR-mediated innate immunity, which seems to be involved in tissue injury, tissue regeneration and in progressive tissue scarring. PMID:24009023

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

Hypothalamic transcriptional expression of the kisspeptin system and sex steroid receptors differs among polycystic ovary syndrome rat models with different endocrine phenotypes.

PubMed

Marcondes, Rodrigo Rodrigues; Carvalho, Kátia Cândido; Giannocco, Gisele; Duarte, Daniele Coelho; Garcia, Natália; Soares-Junior, José Maria; da Silva, Ismael Dale Cotrim Guerreiro; Maliqueo, Manuel; Baracat, Edmund Chada; Maciel, Gustavo Arantes Rosa

2017-08-01

Polycystic ovary syndrome is a heterogeneous endocrine disorder that affects reproductive-age women. The mechanisms underlying the endocrine heterogeneity and neuroendocrinology of polycystic ovary syndrome are still unclear. In this study, we investigated the expression of the kisspeptin system and gonadotropin-releasing hormone pulse regulators in the hypothalamus as well as factors related to luteinizing hormone secretion in the pituitary of polycystic ovary syndrome rat models induced by testosterone or estradiol. A single injection of testosterone propionate (1.25 mg) (n=10) or estradiol benzoate (0.5 mg) (n=10) was administered to female rats at 2 days of age to induce experimental polycystic ovary syndrome. Controls were injected with a vehicle (n=10). Animals were euthanized at 90-94 days of age, and the hypothalamus and pituitary gland were used for gene expression analysis. Rats exposed to testosterone exhibited increased transcriptional expression of the androgen receptor and estrogen receptor-β and reduced expression of kisspeptin in the hypothalamus. However, rats exposed to estradiol did not show any significant changes in hormone levels relative to controls but exhibited hypothalamic downregulation of kisspeptin, tachykinin 3 and estrogen receptor-α genes and upregulation of the gene that encodes the kisspeptin receptor. Testosterone- and estradiol-exposed rats with different endocrine phenotypes showed differential transcriptional expression of members of the kisspeptin system and sex steroid receptors in the hypothalamus. These differences might account for the different endocrine phenotypes found in testosterone- and estradiol-induced polycystic ovary syndrome rats.

The Role of Cholecystokinin in Peripheral Taste Signaling in Mice

PubMed Central

Yoshida, Ryusuke; Shin, Misa; Yasumatsu, Keiko; Takai, Shingo; Inoue, Mayuko; Shigemura, Noriatsu; Takiguchi, Soichi; Nakamura, Seiji; Ninomiya, Yuzo

2017-01-01

Cholecystokinin (CCK) is a gut hormone released from enteroendocrine cells. CCK functions as an anorexigenic factor by acting on CCK receptors expressed on the vagal afferent nerve and hypothalamus with a synergistic interaction between leptin. In the gut, tastants such as amino acids and bitter compounds stimulate CCK release from enteroendocrine cells via activation of taste transduction pathways. CCK is also expressed in taste buds, suggesting potential roles of CCK in taste signaling in the peripheral taste organ. In the present study, we focused on the function of CCK in the initial responses to taste stimulation. CCK was coexpressed with type II taste cell markers such as Gα-gustducin, phospholipase Cβ2, and transient receptor potential channel M5. Furthermore, a small subset (~30%) of CCK-expressing taste cells expressed a sweet/umami taste receptor component, taste receptor type 1 member 3, in taste buds. Because type II taste cells are sweet, umami or bitter taste cells, the majority of CCK-expressing taste cells may be bitter taste cells. CCK-A and -B receptors were expressed in both taste cells and gustatory neurons. CCK receptor knockout mice showed reduced neural responses to bitter compounds compared with wild-type mice. Consistently, intravenous injection of CCK-Ar antagonist lorglumide selectively suppressed gustatory nerve responses to bitter compounds. Intravenous injection of CCK-8 transiently increased gustatory nerve activities in a dose-dependent manner whereas administration of CCK-8 did not affect activities of bitter-sensitive taste cells. Collectively, CCK may be a functionally important neurotransmitter or neuromodulator to activate bitter nerve fibers in peripheral taste tissues. PMID:29163209

Fibroblast Growth Factor Receptors (FGFRs) in Human Sperm: Expression, Functionality and Involvement in Motility Regulation.

PubMed

Saucedo, Lucía; Buffa, Gabriela N; Rosso, Marina; Guillardoy, Tomás; Góngora, Adrian; Munuce, María J; Vazquez-Levin, Mónica H; Marín-Briggiler, Clara

2015-01-01

Fibroblast growth factors receptors (FGFRs) have been widely characterized in somatic cells, but there is scarce evidence of their expression and function in mammalian gametes. The objective of the present study was to evaluate the expression of FGFRs in human male germ cells, to determine sperm FGFR activation by the FGF2 ligand and their participation in the regulation of sperm motility. The expression of FGFR1, 2, 3 and 4 mRNAs and proteins in human testis and localization of these receptors in germ cells of the seminiferous epithelium was demonstrated. In ejaculated sperm, FGFRs were localized to the acrosomal region and flagellum. Sperm exposure to FGF2 caused an increase in flagellar FGFR phosphorylation and activation of extracellular signal-regulated kinase (ERK) and protein kinase B (PKB or Akt) signaling pathways. Incubation with FGF2 led to a significant increase in the percentage of total and progressive sperm motility, as well as in sperm kinematics. All responses were prevented by sperm preincubation with BGJ398, a specific inhibitor of FGFR tyrosine kinase activity. In addition to confirming the expression of FGFRs in germ cells of the human testis, our study describes for the first time the presence, localization and functionality of human sperm FGFRs, and provides evidence of the beneficial effect of FGF2 upon sperm motility.

Fibroblast Growth Factor Receptors (FGFRs) in Human Sperm: Expression, Functionality and Involvement in Motility Regulation

PubMed Central

Saucedo, Lucía; Buffa, Gabriela N.; Rosso, Marina; Guillardoy, Tomás; Góngora, Adrian; Munuce, María J.

2015-01-01

Fibroblast growth factors receptors (FGFRs) have been widely characterized in somatic cells, but there is scarce evidence of their expression and function in mammalian gametes. The objective of the present study was to evaluate the expression of FGFRs in human male germ cells, to determine sperm FGFR activation by the FGF2 ligand and their participation in the regulation of sperm motility. The expression of FGFR1, 2, 3 and 4 mRNAs and proteins in human testis and localization of these receptors in germ cells of the seminiferous epithelium was demonstrated. In ejaculated sperm, FGFRs were localized to the acrosomal region and flagellum. Sperm exposure to FGF2 caused an increase in flagellar FGFR phosphorylation and activation of extracellular signal-regulated kinase (ERK) and protein kinase B (PKB or Akt) signaling pathways. Incubation with FGF2 led to a significant increase in the percentage of total and progressive sperm motility, as well as in sperm kinematics. All responses were prevented by sperm preincubation with BGJ398, a specific inhibitor of FGFR tyrosine kinase activity. In addition to confirming the expression of FGFRs in germ cells of the human testis, our study describes for the first time the presence, localization and functionality of human sperm FGFRs, and provides evidence of the beneficial effect of FGF2 upon sperm motility. PMID:25970615

Localization and signaling patterns of vascular endothelial growth factors and receptors in human periapical lesions.

PubMed

Virtej, Anca; Løes, Sigbjørn S; Berggreen, Ellen; Bletsa, Athanasia

2013-05-01

Vascular endothelial growth factors (VEGFs) and their receptors (VEGFRs) are key players in vasculogenesis and are also involved in pathologic conditions with bone destruction. Vasculogenesis is critical for disease progression, and bone resorption is a hallmark of apical periodontitis. However, the localization of VEGFs and VEGFRs and their gene signaling pathways in human apical periodontitis have not been thoroughly investigated. The aim of this study was to localize VEGFs and VEGFRs and analyze their gene expression as well as signaling pathways in human periapical lesions. Tissue was collected after endodontic surgery from patients diagnosed with chronic apical periodontitis. Periodontal ligament samples from extracted healthy wisdom teeth was also collected and used as control tissue. In lesion cryosections, VEGFs/VEGFRs were identified by immunohistochemistry/double immunofluorescence by using specific antibodies. A human VEGF signaling polymerase chain reaction array system was used for gene expression analysis comparing lesions with periodontal ligament samples. The histologic evaluation revealed heterogeneous morphology of the periapical lesions with various degrees of inflammatory infiltrates. In the lesions, all investigated factors and receptors were identified in blood vessels and various immune cells. No lymphatic vessels were detected. Gene expression analysis revealed up-regulation of VEGF-A and VEGFR-3, although not significant. Phosphatidylinositol-3-kinases, protein kinase C, mitogen-activated protein kinases, and phospholipases, all known to be involved in VEGF-mediated angiogenic activity, were significantly up-regulated. The cellular and vascular expressions of VEGFs and VEGFRs in chronic apical periodontitis, along with significant alterations of genes mediating VEGF-induced angiogenic responses, suggest ongoing vascular remodeling in established chronic periapical lesions. Copyright © 2013 American Association of Endodontists. Published by Elsevier Inc. All rights reserved.

Steroid Receptor Coactivator 3 Contributes to Host Defense against Enteric Bacteria by Recruiting Neutrophils via Upregulation of CXCL2 Expression.

PubMed

Chen, Wenbo; Lu, Xuqiang; Chen, Yuan; Li, Ming; Mo, Pingli; Tong, Zhangwei; Wang, Wei; Wan, Wei; Su, Guoqiang; Xu, Jianming; Yu, Chundong

2017-02-15

Steroid receptor coactivator 3 (SRC-3) is a transcriptional coactivator that interacts with nuclear receptors and some other transcription factors to enhance their effects on target gene transcription. We reported previously that SRC-3-deficient (SRC-3 -/- ) mice are extremely susceptible to Escherichia coli-induced septic peritonitis as a result of uncontrolled inflammation and a defect in bacterial clearance. In this study, we observed significant upregulation of SRC-3 in colonic epithelial cells in response to Citrobacter rodentium infection. Based on these findings, we hypothesized that SRC-3 is involved in host defense against attaching and effacing bacterial infection. We compared the responses of SRC-3 -/- and wild-type mice to intestinal C. rodentium infection. We found that SRC-3 -/- mice exhibited delayed clearance of C. rodentium and more severe tissue pathology after oral infection with C. rodentium compared with wild-type mice. SRC-3 -/- mice expressed normal antimicrobial peptides in the colons but exhibited delayed recruitment of neutrophils into the colonic mucosa. Accordingly, SRC-3 -/- mice showed a delayed induction of CXCL2 and CXCL5 in colonic epithelial cells, which are responsible for neutrophil recruitment. At the molecular level, we found that SRC-3 can activate the NF-κB signaling pathway to promote CXCL2 expression at the transcriptional level. Collectively, we show that SRC-3 contributes to host defense against enteric bacteria, at least in part via upregulating CXCL2 expression to recruit neutrophils. Copyright © 2017 by The American Association of Immunologists, Inc.

Insulin-Like Growth Factors Are Expressed in the Taste System, but Do Not Maintain Adult Taste Buds

PubMed Central

Biggs, Bradley T.; Tang, Tao; Krimm, Robin F.

2016-01-01

Growth factors regulate cell growth and differentiation in many tissues. In the taste system, as yet unknown growth factors are produced by neurons to maintain taste buds. A number of growth factor receptors are expressed at greater levels in taste buds than in the surrounding epithelium and may be receptors for candidate factors involved in taste bud maintenance. We determined that the ligands of eight of these receptors were expressed in the E14.5 geniculate ganglion and that four of these ligands were expressed in the adult geniculate ganglion. Of these, the insulin-like growth factors (IGF1, IGF2) were expressed in the ganglion and their receptor, insulin-like growth factor receptor 1 (IGF1R), were expressed at the highest levels in taste buds. To determine whether IGF1R regulates taste bud number or structure, we conditionally eliminated IGF1R from the lingual epithelium of mice using the keratin 14 (K14) promoter (K14-Cre::Igf1rlox/lox). While K14-Cre::Igf1rlox/lox mice had significantly fewer taste buds at P30 compared with control mice (Igf1rlox/lox), this difference was not observed by P80. IGF1R removal did not affect taste bud size or cell number, and the number of phospholipase C β2- (PLCβ2) and carbonic anhydrase 4- (Car4) positive taste receptor cells did not differ between genotypes. Taste buds at the back of the tongue fungiform taste field were larger and contained more cells than those at the tongue tip, and these differences were diminished in K14-Cre::Igf1rlox/lox mice. The epithelium was thicker at the back versus the tip of the tongue, and this difference was also attenuated in K14-Cre::Igf1rlox/lox mice. We conclude that, although IGFs are expressed at high levels in the taste system, they likely play little or no role in maintaining adult taste bud structure. IGFs have a potential role in establishing the initial number of taste buds, and there may be limits on epithelial thickness in the absence of IGF1R signaling. PMID:26901525

Mouse glucocorticoid-induced tumor necrosis factor receptor ligand is costimulatory for T cells

PubMed Central

Tone, Masahide; Tone, Yukiko; Adams, Elizabeth; Yates, Stephen F.; Frewin, Mark R.; Cobbold, Stephen P.; Waldmann, Herman

2003-01-01

Recently, agonist antibodies to glucocorticoid-induced tumor necrosis factor receptor (GITR) (tumor necrosis factor receptor superfamily 18) have been shown to neutralize the suppressive activity of CD4+CD25+ regulatory T cells. It was anticipated that this would be the role of the physiological ligand. We have identified and expressed the gene for mouse GITR ligand and have confirmed that its interaction with GITR reverses suppression by CD4+CD25+ T cells. It also, however, provides a costimulatory signal for the antigen-driven proliferation of naïve T cells and polarized T helper 1 and T helper 2 clones. RT-PCR and mAb staining revealed mouse GITR ligand expression in dendritic cells, macrophages, and B cells. Expression was controlled by the transcription factor NF-1 and potentially by alternative splicing of mRNA destabilization sequences. PMID:14608036

Expression of VEGFR and PDGFR-α/-β in 187 canine nasal carcinomas.

PubMed

Gramer, I; Killick, D; Scase, T; Chandry, D; Marrington, M; Blackwood, L

2017-09-01

Radiotherapy represents the standard of care for intranasal carcinomas. Responses to tyrosine kinase inhibitors (TKIs) have been reported but data on expression of target receptor tyrosine kinases (rTKs) is limited. This study characterizes the expression of vascular endothelial growth factor receptor (VEGFR), platelet-derived growth factor receptor (PDGFR)-α and PDGFR-β in canine intranasal carcinomas. Histological samples from 187 dogs were retrieved. Immunohistochemistry was performed using commercially available antibodies. Expression of rTKs was classified into weak, moderate or intense and additionally recorded as cytoplasmic, membranous, cytoplasmic-membranous, nuclear or stromal. VEGFR was expressed in 158 dogs with predominantly moderate expression (36.9%) and a cytoplasmic-membranous expression pattern (70.9%). PDGFR-α was detected in 133 with predominantly weak expression (57.9%) and cytoplasmic pattern (87.9%). PDGFR-β was identified in 74 patients with a predominantly moderate expression (17.6%) and cytoplasmic expression pattern (63.5%). Co-expression of rTKs was common. These results confirm expression of VEGFR, PDGFR-α and PDGFR-β in canine intranasal carcinomas and support the utility of TKIs. © 2016 John Wiley & Sons Ltd.

The combination of Bifidobacterium breve with non-digestible oligosaccharides suppresses airway inflammation in a murine model for chronic asthma.

PubMed

Sagar, Seil; Vos, Arjan P; Morgan, Mary E; Garssen, Johan; Georgiou, Niki A; Boon, Louis; Kraneveld, Aletta D; Folkerts, Gert

2014-04-01

Over the last decade, there has been a growing interest in the use of interventions that target the intestinal microbiota as a treatment approach for asthma. This study is aimed at exploring the therapeutic effects of long-term treatment with a combination of Bifidobacterium breve with non-digestible oligosaccharides on airway inflammation and remodeling. A murine ovalbumin-induced chronic asthma model was used. Pulmonary airway inflammation; mRNA expression of pattern recognition receptors, Th-specific cytokines and transcription factors in lung tissue; expression of Foxp3 in blood Th cells; in vitro T cell activation; mast cell degranulation; and airway remodeling were examined. The combination of B. breve with non-digestible oligosaccharides suppressed pulmonary airway inflammation; reduced T cell activation and mast cell degranulation; modulated expression of pattern recognition receptors, cytokines and transcription factors; and reduced airway remodeling. The treatment induced regulatory T cell responses, as shown by increased Il10 and Foxp3 transcription in lung tissue, and augmented Foxp3 protein expression in blood CD4+CD25+Foxp3+ T cells. This specific combination of beneficial bacteria with non-digestible oligosaccharides has strong anti-inflammatory properties, possibly via the induction of a regulatory T cell response, resulting in reduced airway remodeling and, therefore, may be beneficial in the treatment of chronic inflammation in allergic asthma. Copyright © 2014 Elsevier B.V. All rights reserved.

Leptospira santorosai Serovar Shermani detergent extract induces an increase in fibronectin production through a Toll-like receptor 2-mediated pathway.

PubMed

Tian, Ya-Chung; Hung, Cheng-Chieh; Li, Yi-Jung; Chen, Yung-Chang; Chang, Ming-Yang; Yen, Tzung-Hai; Hsu, Hsiang-Hao; Wu, Mai-Szu; Phillips, Aled; Yang, Chih-Wei

2011-03-01

Leptospirosis can activate inflammatory responses through Toll-like receptors (TLRs) and may cause renal tubulointerstitial fibrosis characterized by the accumulation of extracellular matrix (ECM). We have previously demonstrated that Leptospira santorosai serovar Shermani detergent extract stimulates ECM accumulation in vitro. The aim of this study was to examine the mechanistic basis of these previous observations and, in particular, to examine the potential involvement of TLRs. The addition of serovar Shermani detergent extract led to an increase in fibronectin gene expression and production. Inhibition of TLR2 but not TLR4 expression abrogated serovar Shermani detergent extract-mediated increases in fibronectin production. This response was also blocked by the knockdown of the gene expression of the TLR2 downstream transducers myeloid differentiation factor 88 (MyD88) and tumor necrosis factor receptor-associated factor 6 (TRAF6). Serovar Shermani detergent extract also activated nuclear factor-κB, and its inhibition by curcumin-attenuated serovar Shermani detergent extract induced increases in fibronectin production. These effects were also mimicked by the specific TLR2 agonist, Pam(3)CsK(4), a response that was also abrogated by the knockdown of MyD88 and TRAF6. Similarly, the administration of live leptospires to cells also induced fibronectin production that was blocked by inhibition of TLR2 and MyD88 expression. In conclusion, serovar Shermani detergent extract can induce fibronectin production through the TLR2-associated cascade, providing evidence of an association between TLRs and leptospirosis-mediated ECM deposition.

Fumaric acid attenuates the eotaxin-1 expression in TNF-α-stimulated fibroblasts by suppressing p38 MAPK-dependent NF-κB signaling.

PubMed

Roh, Kyung-Baeg; Jung, Eunsun; Park, Deokhoon; Lee, Jongsung

2013-08-01

Eotaxin-1 is a potent chemoattractant for eosinophils and a critical mediator during the development of eosinophilic inflammation. Fumaric acid is an intermediate product of the citric acid cycle, which is source of intracellular energy. Although fumaric acid ameliorates psoriasis and multiple sclerosis, its involvement in eotaxin-1-mediated effects has not been assessed. In this study, we investigated the effects of fumaric acid on eotaxin-1 expression in a mouse fibroblast cell line. We found that fumaric acid significantly inhibited tumor necrosis factor-α (TNF-α-induced eotaxin-1 expression. This fumaric acid effect was mediated through the inhibition of p38 mitogen-activated protein kinase (MAPK)-dependent nuclear factor (NF)-κB signaling. We also found that fumaric acid operates downstream of MEKK3 during TNF-α-induced NF-κB signaling, which upregulated eotaxin-1 expression. In addition, fumaric acid attenuated expression of CC-chemokine receptor 3 (CCR3), an eotaxin-1 receptor, and adhesion molecules that play important roles in eosinophil binding to induce allergic inflammation. Taken together, these findings indicate that inhibiting TNF-α-induced eotaxin-1 expression by fumaric acid occurs primarily through suppression of NF-κB signaling, which is mediated by inhibiting p38 MAPK and suggest that fumaric acid may be used as a complementary treatment option for eotaxin-1-mediated diseases. Copyright © 2013 Elsevier Ltd. All rights reserved.

Phosphorylation and Internalization of Lysophosphatidic Acid Receptors LPA1, LPA2, and LPA3

PubMed Central

Alcántara-Hernández, Rocío; Hernández-Méndez, Aurelio; Campos-Martínez, Gisselle A.; Meizoso-Huesca, Aldo; García-Sáinz, J. Adolfo

2015-01-01

Results The lysophosphatidic acid receptors LPA1, LPA2, and LPA3 were individually expressed in C9 cells and their signaling and regulation were studied. Agonist-activation increases intracellular calcium concentration in a concentration-dependent fashion. Phorbol myristate acetate markedly inhibited LPA1- and LPA3-mediated effect, whereas that mediated by LPA2 was only partially diminished; the actions of the phorbol ester were inhibited by bisindolylmaleimide I and by overnight incubation with the protein kinase C activator, which leads to down regulation of this protein kinase. Homologous desensitization was also observed for the three LPA receptors studied, with that of LPA2 receptors being consistently of lesser magnitude; neither inhibition nor down-regulation of protein kinase C exerted any effect on homologous desensitization. Activation of LPA1–3 receptors induced ERK 1/2 phosphorylation; this effect was markedly attenuated by inhibition of epidermal growth factor receptor tyrosine kinase activity, suggesting growth factor receptor transactivation in this effect. Lysophosphatidic acid and phorbol myristate acetate were able to induce LPA1–3 phosphorylation, in time- and concentration-dependent fashions. It was also clearly observed that agonists and protein kinase C activation induced internalization of these receptors. Phosphorylation of the LPA2 subtype required larger concentrations of these agents and its internalization was less intense than that of the other subtypes. Conclusion Our data show that these three LPA receptors are phosphoproteins whose phosphorylation state is modulated by agonist-stimulation and protein kinase C-activation and that differences in regulation and cellular localization exist, among the subtypes. PMID:26473723

Improvement in verbal memory following SSRI augmentation of antipsychotic treatment is associated with changes in the expression of mRNA encoding for the GABA-A receptor and BDNF in PMC of schizophrenic patients.

PubMed

Silver, Henry; Mandiuk, Nina; Einoch, Reef; Susser, Ehud; Danovich, Lena; Bilker, Warren; Youdim, Moussa; Weinreb, Orly

2015-05-01

Verbal memory impairment in schizophrenia is associated with abnormalities in gamma-aminobutyric acid (GABA)-ergic and brain-derived neurotrophic factor (BDNF) systems. Recent evidence from animal and clinical studies that adding fluvoxamine to antipsychotics alters the expression of transcripts encoding for the GABA-A receptor and BDNF led us to postulate that fluvoxamine augmentation may improve memory in schizophrenia. To test this, we examined the effect of add-on fluvoxamine on verbal memory and other cognitive functions and related it to the expression of mRNA coding for the GABA-A receptor and BDNF in peripheral mononuclear cells (PMC) of schizophrenic patients. Twenty-nine patients completed a 6-week study in which fluvoxamine (100 mg/day) was added to ongoing antipsychotic treatment. Verbal memory, abstraction working memory, object and face recognition, and psychomotor speed and clinical symptoms were assessed at baseline and after 3 and 6 weeks of treatment. Blood samples were taken at baseline and weeks 1, 3, and 6 and PMC was assayed for the GABA-A beta3 receptor and BDNF mRNA by quantitative real-time reverse transcription-PCR. Associative and logical verbal memory improved significantly and showed a significant correlation with changes in PMC BDNF and GABA-A beta3 receptor mRNA, which increased during treatment. Abstraction and object recognition improved, but this did not correlate with PMC measures. Negative and positive symptoms improved significantly; the latter showed significant correlations with changes in PMC measures. Addition of fluvoxamine to antipsychotics improves verbal memory. It is postulated that the mechanism involves enhanced GABA-A receptor/BDNF-dependent synaptic plasticity in the hippocampus.

Piracy of PGE2/EP receptor mediated signaling by Kaposi’s sarcoma associated herpes virus (KSHV/HHV-8) for latency gene expression: Strategy of a successful pathogen

PubMed Central

Paul, Arun George; Sharma-Walia, Neelam; Kerur, Nagaraj; White, Carl; Chandran, Bala

2010-01-01

KSHV is implicated in the pathogenesis of KS, a chronic inflammation associated malignancy. COX-2 and its metabolite PGE2, two pivotal proinflammatory/oncogeneic molecules, are proposed to play roles in the expression of major KSHV latency associated nuclear antigen-1 (LANA-1). Microsomal prostaglandin E2 synthase (mPGES), PGE2 and its receptors (EP1, EP2, EP3, and EP4) were detected in KS lesions with the distinct staining of EP2/EP4 in KS lesions. In latently infected endothelial TIVE-LTC cells, EP receptor antagonists down-regulated LANA-1 expression as well as Ca2+, p-Src, p-PI3K, p-PKCζ/λ, and p-NF-κB, which are also some of the signal molecules proposed to be important in KS pathogenesis. Exogenous PGE2 and EP receptor agonists induced the LANA-1 promoter in 293 cells, and YY1, Sp1, Oct-1, Oct-6, C/EBP and c-Jun transcription factors appear to be involved in this induction. PGE2/EP receptor induced LANA-1 promoter activity was down-regulated significantly by the inhibition of Ca2+, p-Src, p-PI3K, p-PKCζ/λ, and p-NF-κB. These findings implicate the inflammatory PGE2/EP receptors and the associated signal molecules in herpes virus latency and uncover a novel paradigm that demonstrates the evolution of KSHV genome plasticity to utilize inflammatory response for its survival advantage of maintaining latent gene expression. This data also suggests that potential use of anti-COX-2 and anti-EP receptor therapy may not only ameliorate the chronic inflammation associated with KS but could also lead to elimination of the KSHV latent infection and the associated KS lesions. PMID:20388794

Identification of modulators of the nuclear receptor peroxisome proliferator-activated receptor α (PPARα) in a mouse liver gene expression compendium.

PubMed

Oshida, Keiyu; Vasani, Naresh; Thomas, Russell S; Applegate, Dawn; Rosen, Mitch; Abbott, Barbara; Lau, Christopher; Guo, Grace; Aleksunes, Lauren M; Klaassen, Curtis; Corton, J Christopher

2015-01-01

The nuclear receptor family member peroxisome proliferator-activated receptor α (PPARα) is activated by therapeutic hypolipidemic drugs and environmentally-relevant chemicals to regulate genes involved in lipid transport and catabolism. Chronic activation of PPARα in rodents increases liver cancer incidence, whereas suppression of PPARα activity leads to hepatocellular steatosis. Analytical approaches were developed to identify biosets (i.e., gene expression differences between two conditions) in a genomic database in which PPARα activity was altered. A gene expression signature of 131 PPARα-dependent genes was built using microarray profiles from the livers of wild-type and PPARα-null mice after exposure to three structurally diverse PPARα activators (WY-14,643, fenofibrate and perfluorohexane sulfonate). A fold-change rank-based test (Running Fisher's test (p-value ≤ 10(-4))) was used to evaluate the similarity between the PPARα signature and a test set of 48 and 31 biosets positive or negative, respectively for PPARα activation; the test resulted in a balanced accuracy of 98%. The signature was then used to identify factors that activate or suppress PPARα in an annotated mouse liver/primary hepatocyte gene expression compendium of ~1850 biosets. In addition to the expected activation of PPARα by fibrate drugs, di(2-ethylhexyl) phthalate, and perfluorinated compounds, PPARα was activated by benzofuran, galactosamine, and TCDD and suppressed by hepatotoxins acetaminophen, lipopolysaccharide, silicon dioxide nanoparticles, and trovafloxacin. Additional factors that activate (fasting, caloric restriction) or suppress (infections) PPARα were also identified. This study 1) developed methods useful for future screening of environmental chemicals, 2) identified chemicals that activate or suppress PPARα, and 3) identified factors including diets and infections that modulate PPARα activity and would be hypothesized to affect chemical-induced PPARα activity.

Identification of Modulators of the Nuclear Receptor Peroxisome Proliferator-Activated Receptor α (PPARα) in a Mouse Liver Gene Expression Compendium

PubMed Central

Oshida, Keiyu; Vasani, Naresh; Thomas, Russell S.; Applegate, Dawn; Rosen, Mitch; Abbott, Barbara; Lau, Christopher; Guo, Grace; Aleksunes, Lauren M.; Klaassen, Curtis; Corton, J. Christopher

2015-01-01

The nuclear receptor family member peroxisome proliferator-activated receptor α (PPARα) is activated by therapeutic hypolipidemic drugs and environmentally-relevant chemicals to regulate genes involved in lipid transport and catabolism. Chronic activation of PPARα in rodents increases liver cancer incidence, whereas suppression of PPARα activity leads to hepatocellular steatosis. Analytical approaches were developed to identify biosets (i.e., gene expression differences between two conditions) in a genomic database in which PPARα activity was altered. A gene expression signature of 131 PPARα-dependent genes was built using microarray profiles from the livers of wild-type and PPARα-null mice after exposure to three structurally diverse PPARα activators (WY-14,643, fenofibrate and perfluorohexane sulfonate). A fold-change rank-based test (Running Fisher’s test (p-value ≤ 10-4)) was used to evaluate the similarity between the PPARα signature and a test set of 48 and 31 biosets positive or negative, respectively for PPARα activation; the test resulted in a balanced accuracy of 98%. The signature was then used to identify factors that activate or suppress PPARα in an annotated mouse liver/primary hepatocyte gene expression compendium of ~1850 biosets. In addition to the expected activation of PPARα by fibrate drugs, di(2-ethylhexyl) phthalate, and perfluorinated compounds, PPARα was activated by benzofuran, galactosamine, and TCDD and suppressed by hepatotoxins acetaminophen, lipopolysaccharide, silicon dioxide nanoparticles, and trovafloxacin. Additional factors that activate (fasting, caloric restriction) or suppress (infections) PPARα were also identified. This study 1) developed methods useful for future screening of environmental chemicals, 2) identified chemicals that activate or suppress PPARα, and 3) identified factors including diets and infections that modulate PPARα activity and would be hypothesized to affect chemical-induced PPARα activity. PMID:25689681

Inhibiting receptor for advanced glycation end product (AGE) and oxidative stress involved in the protective effect mediated by glucagon-like peptide-1 receptor on AGE induced neuronal apoptosis.

PubMed

Chen, Song; Yin, Lei; Xu, Zheng; An, Feng-Mao; Liu, Ai-Ran; Wang, Ying; Yao, Wen-Bing; Gao, Xiang-Dong

2016-01-26

Our previous study has demonstrated that glucagon-like peptide-1 (GLP-1) receptor agonist could protect neurons from advanced glycation end products (AGEs) toxicity in vitro. However, further studies are still needed to clarify the molecular mechanism of this GLP-1 receptor -dependent action. The present study mainly focused on the effect of GLP-1 receptor agonists against the receptor for advanced glycation end products (RAGE) signal pathway and the mechanism underlying this effect of GLP-1. Firstly the data based on the SH-GLP-1R(+) and SH-SY5Y cells confirmed our previous finding that GLP-1 receptor could mediate the protective effect against AGEs. The assays of the protein activity and of the mRNA level revealed that apoptosis-related proteins such as caspase-3, caspase-9, Bax and Bcl-2 were involved. Additionally, we found that both GLP-1 and exendin-4 could reduce AGEs-induced reactive oxygen species (ROS) accumulation by suppressing the activity of nicotinamide adenine dinucleotide phosphate-oxidase. Interestingly, we also found that GLP-1 receptor activation could attenuate the abnormal expression of the RAGE in vitro and in vivo. Furthermore, based on the analysis of the protein expression and translocation level of transcription factor nuclear factor-κB (NF-κB), and the use of GLP-1 receptor antagonist exendin(9-39) and NF-κB inhibitor pyrrolidine dithiocarbamate, we found that the effect mediated by GLP-1 receptor could alleviate the over expression of RAGE induced by ligand via the suppression of NF-κB. In summary, the results indicated that inhibiting RAGE/oxidative stress was involved in the protective effect of GLP-1 on neuron cells against AGEs induced apoptosis. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

Vascular endothelial growth factor c/vascular endothelial growth factor receptor 3 signaling regulates chemokine gradients and lymphocyte migration from tissues to lymphatics.

PubMed

Iwami, Daiki; Brinkman, C Colin; Bromberg, Jonathan S

2015-04-01

Circulation of leukocytes via blood, tissue and lymph is integral to adaptive immunity. Afferent lymphatics form CCL21 gradients to guide dendritic cells and T cells to lymphatics and then to draining lymph nodes (dLN). Vascular endothelial growth factor C and vascular endothelial growth factor receptor 3 (VEGFR-3) are the major lymphatic growth factor and receptor. We hypothesized these molecules also regulate chemokine gradients and lymphatic migration. CD4 T cells were injected into the foot pad or ear pinnae, and migration to afferent lymphatics and dLN quantified by flow cytometry or whole mount immunohistochemistry. Vascular endothelial growth factor receptor 3 or its signaling or downstream actions were modified with blocking monoclonal antibodies (mAbs) or other reagents. Anti-VEGFR-3 prevented migration of CD4 T cells into lymphatic lumen and significantly decreased the number that migrated to dLN. Anti-VEGFR-3 abolished CCL21 gradients around lymphatics, although CCL21 production was not inhibited. Heparan sulfate (HS), critical to establish CCL21 gradients, was down-regulated around lymphatics by anti-VEGFR-3 and this was dependent on heparanase-mediated degradation. Moreover, a Phosphoinositide 3-kinase (PI3K)α inhibitor disrupted HS and CCL21 gradients, whereas a PI3K activator prevented the effects of anti-VEGFR-3. During contact hypersensitivity, VEGFR-3, CCL21, and HS expression were all attenuated, and anti-heparanase or PI3K activator reversed these effects. Vascular endothelial growth factor C/VEGFR-3 signaling through PI3Kα regulates the activity of heparanase, which modifies HS and CCL21 gradients around lymphatics. The functional and physical linkages of these molecules regulate lymphatic migration from tissues to dLN. These represent new therapeutic targets to influence immunity and inflammation.

The overexpressed human 46-kDa mannose 6-phosphate receptor mediates endocytosis and sorting of. beta. -glucuronidase

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

Watanabe, H.; Grubb, J.H.; Sly, W.S.

1990-10-01

The authors studied the function of the human small (46-kDa) mannose 6-phosphate receptor (SMPR) in transfected mouse L cells that do not express the larger insulin-like growth factor II/mannose 6-phosphate receptor. Cells overexpressing human SMPR were studied for enzyme binding to cell surface receptors, for binding to intracellular receptors in permeabilized cells, and for receptor-mediated endocytosis of recombinant human {beta}-glucuronidase. Specific binding to human SMPR in permeabilized cells showed a pH optimum between pH 6.0 and pH 6.5. Binding was significant in the present of EDTA but was enhanced by added divalent cations. Up to 2.3{percent} of the total functionalmore » receptor could be detected on the cell surface by enzyme binding. They present experiments showing that at very high levels of overexpression, and at pH 6.5, human SMPR mediated the endocytosis of {beta}-glucuronidase. At pH 7.5, the rate of endocytosis was only 14{percent} the rate seen at pH 6.5. Cells overexpressing human SMPR also showed reduced secretion of newly synthesized {beta}-glucuronidase when compared to cells transfected with vector only, suggesting that overexpressed human SMPR can participate in sorting of newly synthesized {beta}-glucuronidase and partially correct the sorting defect in mouse L cells that do not express the insulin-like growth factor II/mannose 6-phosphate receptor.« less

The Oxidized Low-Density Lipoprotein Receptor Mediates Vascular Effects of Inhaled Vehicle Emissions

PubMed Central

Lucero, JoAnn; Harman, Melissa; Madden, Michael C.; McDonald, Jacob D.; Seagrave, Jean Clare; Campen, Matthew J.

2011-01-01

Rationale: To determine vascular signaling pathways involved in inhaled air pollution (vehicular engine emission) exposure–induced exacerbation of atherosclerosis that are associated with onset of clinical cardiovascular events. Objectives: To elucidate the role of oxidized low-density lipoprotein (oxLDL) and its primary receptor on endothelial cells, the lectin-like oxLDL receptor (LOX-1), in regulation of endothelin-1 expression and matrix metalloproteinase activity associated with inhalational exposure to vehicular engine emissions. Methods: Atherosclerotic apolipoprotein E knockout mice were exposed by inhalation to filtered air or mixed whole engine emissions (250 μg particulate matter [PM]/m3 diesel + 50 μg PM/m3 gasoline exhausts) 6 h/d for 7 days. Concurrently, mice were treated with either mouse IgG or neutralizing antibodies to LOX-1 every other day. Vascular and plasma markers of oxidative stress and expression proatherogenic factors were assessed. In a parallel study, healthy human subjects were exposed to either 100 μg PM/m3 diesel whole exhaust or high-efficiency particulate air and charcoal-filtered “clean” air (control subjects) for 2 hours, on separate occasions. Measurements and Main Results: Mixed emissions exposure increased oxLDL and vascular reactive oxygen species, as well as LOX-1, matrix metalloproteinase-9, and endothelin-1 mRNA expression and also monocyte/macrophage infiltration, each of which was attenuated with LOX-1 antibody treatment. In a parallel study, diesel exhaust exposure in volunteer human subjects induced significant increases in plasma-soluble LOX-1. Conclusions: These findings demonstrate that acute exposure to vehicular source pollutants results in up-regulation of vascular factors associated with progression of atherosclerosis, endothelin-1, and matrix metalloproteinase-9, mediated through oxLDL–LOX-1 receptor signaling, which may serve as a novel target for future therapy. PMID:21493736

Disruption of transforming growth factor-beta signaling by curcumin induces gene expression of peroxisome proliferator-activated receptor-gamma in rat hepatic stellate cells.

PubMed

Zheng, Shizhong; Chen, Anping

2007-01-01

Activation of hepatic stellate cells (HSC), the major effectors of hepatic fibrogenesis, is coupled with sequential alterations in gene expression, including an increase in receptors for transforming growth factor-beta (TGF-beta) and a dramatic reduction in the peroxisome proliferator-activated receptor-gamma (PPAR-gamma). The relationship between them remains obscure. We previously demonstrated that curcumin induced gene expression of PPAR-gamma in activated HSC, leading to reducing cell proliferation, inducing apoptosis and suppressing expression of extracellular matrix genes. The underlying molecular mechanisms are largely unknown. We recently observed that stimulation of PPAR-gamma activation suppressed gene expression of TGF-beta receptors in activated HSC, leading to the interruption of TGF-beta signaling. This observation supported our assumption of an antagonistic relationship between PPAR-gamma activation and TGF-beta signaling in HSC. In this study, we further hypothesize that TGF-beta signaling might negatively regulate gene expression of PPAR-gamma in activated HSC. The present report demonstrates that exogenous TGF-beta1 inhibits gene expression of PPAR-gamma in activated HSC, which is eliminated by the pretreatment with curcumin likely by interrupting TGF-beta signaling. Transfection assays further indicate that blocking TGF-beta signaling by dominant negative type II TGF-beta receptor increases the promoter activity of PPAR-gamma gene. Promoter deletion assays, site-directed mutageneses, and gel shift assays localize two Smad binding elements (SBEs) in the PPAR-gamma gene promoter, acting as curcumin response elements and negatively regulating the promoter activity in passaged HSC. The Smad3/4 protein complex specifically binds to the SBEs. Overexpression of Smad4 dose dependently eliminates the inhibitory effects of curcumin on the PPAR-gamma gene promoter and TGF-beta signaling. Taken together, these results demonstrate that the interruption of TGF-beta signaling by curcumin induces gene expression of PPAR-gamma in activated HSC in vitro. Our studies provide novel insights into the molecular mechanisms of curcumin in the induction of PPAR-gamma gene expression and in the inhibition of HSC activation.

STAT3 Regulates Uterine Epithelial Remodeling and Epithelial-Stromal Crosstalk During Implantation

PubMed Central

Pawar, Sandeep; Starosvetsky, Elina; Orvis, Grant D.; Behringer, Richard R.; Bagchi, Indrani C.

2013-01-01

Embryo implantation is regulated by a variety of endometrial factors, including cytokines, growth factors, and transcription factors. Earlier studies identified the leukemia inhibitory factor (LIF), a cytokine produced by uterine glands, as an essential regulator of implantation. LIF, acting via its cell surface receptor, activates the signal transducer and activator of transcription 3 (STAT3) in the uterine epithelial cells. However, the precise mechanism via which activated STAT3 promotes uterine function during implantation remains unknown. To identify the molecular pathways regulated by STAT3, we created SWd/d mice in which Stat3 gene is conditionally inactivated in uterine epithelium. The SWd/d mice are infertile due to a lack of embryo attachment to the uterine luminal epithelium and consequent implantation failure. Gene expression profiling of uterine epithelial cells of SWd/d mice revealed dysregulated expression of specific components of junctional complexes, including E-cadherin, α- and β-catenin, and several claudins, which critically regulate epithelial junctional integrity and embryo attachment. In addition, uteri of SWd/d mice exhibited markedly reduced stromal proliferation and differentiation, indicating that epithelial STAT3 controls stromal function via a paracrine mechanism. The stromal defect arose from a drastic reduction in the production of several members of the epidermal growth factor family in luminal epithelium of SWd/d uteri and the resulting lack of activation of epidermal growth factor receptor signaling and mitotic activity in the stromal cells. Collectively, our results uncovered an intricate molecular network operating downstream of STAT3 that regulates uterine epithelial junctional reorganization, and stromal proliferation, and differentiation, which are critical determinants of successful implantation. PMID:24100212

Estrogen receptor (α and β) but not androgen receptor expression is correlated with recurrence, progression and survival in post prostatectomy T3N0M0 locally advanced prostate cancer in an urban Greek population

PubMed Central

Megas, Georgios; Chrisofos, Michael; Anastasiou, Ioannis; Tsitlidou, Aida; Choreftaki, Theodosia; Deliveliotis, Charalampos

2015-01-01

The objective of this study was to evaluate the expression of estrogen receptors (ER(α) and ER(β)) and androgen receptors (ARs) as prognostic factors for biochemical recurrence, disease progression and survival in patients with pT3N0M0 prostate cancer (PCa) in an urban Greek population. A total of 100 consecutive patients with pT3N0M0 PCa treated with radical prostatectomy participated in the study. The mean age and follow-up were 64.2 and 6 years, respectively. The HSCORE was used for semi-quantitative analysis of the immunoreactivity of the receptors. The prognostic value of the ER(α) and ER(β) and AR was assessed in terms of recurrence, progression, and survival. AR expression was not associated with any of the above parameters; however, both ERs correlated with the prognosis. A univariate Cox regression analysis showed that ER(α) positive staining was significantly associated with a greater hazard for all outcomes. Increased ER(β) staining was significantly associated with a lower hazard for all outcomes in the univariate analysis. When both ER HSCORES were used for the analysis, it was found that patients with high ER(α) or low ER(β) HSCORES compared with patients with negatively stained ER(α) and >1.7 hSCORE ER(β) had 6.03, 10.93, and 10.53 times greater hazard for biochemical disease recurrence, progression of disease and death, respectively. Multiple Cox proportional hazard analyses showed that the age, preoperative prostate specific antigen, Gleason score and ERs were independent predictors of all outcomes. ER expression is an important prognosticator after radical prostatectomy in patients with pT3N0M0 PCa. By contrast, AR expression has limited prognostic value. PMID:25219910

Estrogen receptor (α and β) but not androgen receptor expression is correlated with recurrence, progression and survival in post prostatectomy T3N0M0 locally advanced prostate cancer in an urban Greek population.

PubMed

Megas, Georgios; Chrisofos, Michael; Anastasiou, Ioannis; Tsitlidou, Aida; Choreftaki, Theodosia; Deliveliotis, Charalampos

2015-01-01

The objective of this study was to evaluate the expression of estrogen receptors (ER(α) and ER(β)) and androgen receptors (ARs) as prognostic factors for biochemical recurrence, disease progression and survival in patients with pT3N0M0 prostate cancer (PCa) in an urban Greek population. A total of 100 consecutive patients with pT3N0M0 PCa treated with radical prostatectomy participated in the study. The mean age and follow-up were 64.2 and 6 years, respectively. The HSCORE was used for semi-quantitative analysis of the immunoreactivity of the receptors. The prognostic value of the ER(α) and ER(β) and AR was assessed in terms of recurrence, progression, and survival. AR expression was not associated with any of the above parameters; however, both ERs correlated with the prognosis. A univariate Cox regression analysis showed that ER(α) positive staining was significantly associated with a greater hazard for all outcomes. Increased ER(β) staining was significantly associated with a lower hazard for all outcomes in the univariate analysis. When both ER HSCORES were used for the analysis, it was found that patients with high ER(α) or low ER(β) HSCORES compared with patients with negatively stained ER(α) and >1.7 hSCORE ER(β) had 6.03, 10.93, and 10.53 times greater hazard for biochemical disease recurrence, progression of disease and death, respectively. Multiple Cox proportional hazard analyses showed that the age, preoperative prostate specific antigen, Gleason score and ERs were independent predictors of all outcomes. ER expression is an important prognosticator after radical prostatectomy in patients with pT3N0M0 PCa. By contrast, AR expression has limited prognostic value.

Toll-Like Receptor 3 Signaling Contributes to Regional Neutrophil Recruitment in Cultured Human Glomerular Endothelial Cells.

PubMed

Liu, Qiang; Imaizumi, Tadaatsu; Kawaguchi, Shogo; Aizawa, Tomomi; Matsumiya, Tomoh; Watanabe, Shojiro; Tsugawa, Koji; Yoshida, Hidemi; Tsuruga, Kazushi; Joh, Kensuke; Kijima, Hiroshi; Tanaka, Hiroshi

2018-05-23

Given the importance of neutrophil recruitment in the pathogenesis of glomerulonephritis (GN), the representative neutrophil chemoattractant C-X-C motif chemokine 1 (CXCL1)/GROα and the adhesion molecule E-selectin in glomerular endothelial cells (GECs) play a pivotal role in the development of GN. Endothelial Toll-like receptor 3 (TLR3) is thought to be involved in the inflammatory response via innate immunity. However, the role of endothelial TLR3 signaling in the expression of neutrophil chemoattractants and adhesion molecules remains to be elucidated. Thus, we aimed to examine this issue. We treated normal human GECs with polyinosinic-polycytidylic acid (poly IC), an authentic double-stranded RNA, and analyzed the expressions of CXCL1 and E-selectin using quantitative real-time reverse transcription-polymerase chain reaction, western blotting, and enzyme-linked immunosorbent assay. To further elucidate the poly IC-induced signaling pathway, we subjected the cells to RNA interference against TLR3, interferon (IFN)-β, nuclear factor (NF)-κB p65, and IFN regulatory factor (IRF) 3. We also used immunofluorescence to examine the endothelial expression of CXCL1 in biopsy specimens from patients with crescentic and non-crescentic purpura nephritis (PN). We found that the activation of TLR3 induced the endothelial expression of CXCL1 and E-selectin, and that this involved TLR3, -NF-κB, IRF3, and IFN-β. Intense endothelial CXCL1 expression was observed in biopsy specimens from patients with crescentic PN. These findings support a role for glomerular antiviral innate immunity in the pathogenesis of GN. Intervention of glomerular TLR3 signaling may therefore be a suitable therapeutic strategy for treating GN in the future. © 2018 S. Karger AG, Basel.

Low molecular weight glycosaminoglycan C3 attenuates AF64A-stimulated, low-affinity nerve growth factor receptor-immunoreactive axonal varicosities in the rat septum.

PubMed

Dudas, Bertalan; Rose, Michael; Cornelli, Umberto; Hanin, Israel

2005-02-01

Glycosaminoglycans (GAGs) play a pivotal role in the pathogenesis of Alzheimer's disease (AD). Although, as we have shown earlier, a low molecular weight GAG, C3, protects against ethylcholine aziridinium (AF64A)-induced cholinergic damage, and against A(beta)-induced tau-2-immunoreactivity (IR), the mechanism of the neuroprotective effect of GAGs is not yet known. Several clues exist. Previous studies in rats revealed that continuous NGF infusion (icv) after AF64A injection increases septal ChAT and AChE activities. Moreover, C3 increases axonal outgrowth in the rat hippocampus, raising the possibility of a NGF-receptor mediated neuroprotection. Furthermore, it has been reported that NGF expression is increased in the septum following AF64A administration. To study the question regarding the mechanism of neuroprotective action of GAGs, AF64A, a selective cholinotoxin, was administered stereotaxically, bilaterally, into the lateral ventricles of Fischer albino male rats (1 nmol/2 microl/side). In order to establish the effect of C3 on the expression of the NGF receptor-IR elements, C3 was administered orally (25 mg/kg, once a day), by gavage, 7 days before, and 7 days after the AF64A injection. NGF receptor immunohistochemistry revealed that AF64A induced the appearance of NGF-receptor-IR axonal varicosities in the rat medial septum. These varicose fibers were attenuated by 14 days' administration of C3. The possible explanation of our data may be that C3 increases NGF synthesis in the lateral septum. The increased level of NGF could suppress the increased, AF64A-induced NGF receptor expression in the medial septal nucleus. These results further accentuate our earlier observations that C3 may have potential as a therapeutic agent in AD and other neurodegenerative disorders.

Osteopontin inhibits osteoblast responsiveness through the down-regulation of focal adhesion kinase mediated by the induction of low-molecular weight protein tyrosine phosphatase.

PubMed

Kusuyama, Joji; Bandow, Kenjiro; Ohnishi, Tomokazu; Hisadome, Mitsuhiro; Shima, Kaori; Semba, Ichiro; Matsuguchi, Tetsuya

2017-05-15

Osteopontin (OPN) is an osteogenic marker protein. Osteoblast functions are affected by inflammatory cytokines and pathological conditions. OPN is highly expressed in bone lesions such as those in rheumatoid arthritis. However, local regulatory effects of OPN on osteoblasts remain ambiguous. Here we examined how OPN influences osteoblast responses to mechanical stress and growth factors. Expression of NO synthase 1 ( Nos1 ) and Nos2 was increased by low-intensity pulsed ultrasound (LIPUS) in MC3T3-E1 cells and primary osteoblasts. The increase of Nos1/2 expression was abrogated by both exogenous OPN overexpression and recombinant OPN treatment, whereas it was promoted by OPN-specific siRNA and OPN antibody. Moreover, LIPUS-induced phosphorylation of focal adhesion kinase (FAK), a crucial regulator of mechanoresponses, was down-regulated by OPN treatments. OPN also attenuated hepatocyte growth factor-induced vitamin D receptor ( Vdr ) expression and platelet-derived growth factor-induced cell mobility through the repression of FAK activity. Of note, the expression of low-molecular weight protein tyrosine phosphatase (LMW-PTP), a FAK phosphatase, was increased in both OPN-treated and differentiated osteoblasts. CD44 was a specific OPN receptor for LWW-PTP induction. Consistently, the suppressive influence of OPN on osteoblast responsiveness was abrogated by LMW-PTP knockdown. Taken together, these results reveal novel functions of OPN in osteoblast physiology. © 2017 Kusuyama et al. This article is distributed by The American Society for Cell Biology under license from the author(s). Two months after publication it is available to the public under an Attribution–Noncommercial–Share Alike 3.0 Unported Creative Commons License (http://creativecommons.org/licenses/by-nc-sa/3.0).

FADD and the NF-κB family member Bcl-3 regulate complementary pathways to control T-cell survival and proliferation

PubMed Central

Rangelova, Svetla; Kirschnek, Susanne; Strasser, Andreas; Häcker, Georg

2008-01-01

Fas-associated protein with death domain/mediator of receptor induced toxicity (FADD/MORT1) was first described as a transducer of death receptor signalling but was later recognized also to be important for proliferation of T cells. B-cell lymphoma 3 (Bcl-3) is a relatively little understood member of the nuclear factor (NF)-κB family of transcription factors. We recently found that Bcl-3 is up-regulated in T cells from mice where FADD function is blocked by a dominant negative transgene (FADD-DN). To understand the importance of this, we generated FADD-DN/bcl-3−/− mice. Here, we report that T cells from these mice show massive cell death and severely reduced proliferation in response to T-cell receptor (TCR) stimulation in vitro. Transgenic co-expression of Bcl-2 (FADD-DN/bcl-3−/−/vav-bcl-2 mice) rescued the survival but not the proliferation of T cells. FADD-DN/bcl-3−/− mice had normal thymocyte numbers but reduced numbers of peripheral T cells despite an increase in cycling T cells in vivo. However, activation of the classical NF-κB and extracellular regulated kinase (ERK) pathways and expression of interleukin (IL)-2 mRNA upon stimulation were normal in T cells from FADD-DN/bcl-3−/− mice. These data suggest that FADD and Bcl-3 regulate separate pathways that both contribute to survival and proliferation in mouse T cells. PMID:18557791

Role of ERRF, a Novel ER-Related Nuclear Factor, in the Growth Control of ER-Positive Human Breast Cancer Cells

PubMed Central

Su, Dan; Fu, Xiaoying; Fan, Songqing; Wu, Xiao; Wang, Xin-Xin; Fu, Liya; Dong, Xue-Yuan; Ni, Jianping Jenny; Fu, Li; Zhu, Zhengmao; Dong, Jin-Tang

2012-01-01

Whereas estrogen–estrogen receptor α (ER) signaling plays an important role in breast cancer growth, it is also necessary for the differentiation of normal breast epithelial cells. How this functional conversion occurs, however, remains unknown. Based on a genome-wide sequencing study that identified mutations in several breast cancer genes, we examined some of the genes for mutations, expression levels, and functional effects on cell proliferation and tumorigenesis. We present the data for C1orf64 or ER-related factor (ERRF) from 31 cell lines and 367 primary breast cancer tumors. Whereas mutation of ERRF was infrequent (1 of 79 or 1.3%), its expression was up-regulated in breast cancer, and the up-regulation was more common in lower-stage tumors. In addition, increased ERRF expression was significantly associated with ER and/or progesterone receptor (PR) positivity, which was still valid in human epidermal growth factor receptor 2 (HER2)–negative tumors. In ER-positive tumors, ERRF expression was inversely correlated with HER2 status. Furthermore, higher ERRF protein expression was significantly associated with better disease-free survival and overall survival, particularly in ER- and/or PR-positive and HER2-negative tumors (luminal A subtype). Functionally, knockdown of ERRF in two ER-positive breast cancer cell lines, T-47D and MDA-MB-361, suppressed cell growth in vitro and tumorigenesis in xenograft models. These results suggest that ERRF plays a role in estrogen-ER–mediated growth of breast cancer cells and could, thus, be a potential therapeutic target. PMID:22341523

Differential distribution of fibroblast growth factor receptors (FGFRs) on foveal cones: FGFR-4 is an early marker of cone photoreceptors.

PubMed

Cornish, Elisa E; Natoli, Riccardo C; Hendrickson, Anita; Provis, Jan M

2004-01-08

Relatively little is known of the expression and distribution of FGF receptors (FGFR) in the primate retina. We investigated expression of FGFRs in developing and adult Macaca monkey retina, paying particular attention to the cone rich, macular region. One fetal human retina was used for diagnostic PCR using primers designed for FGFR1, FGFR2, FGFR3, FGFR4, and FGFR like-protein 1 (FGFrl1) and for probe design to FGFR3, FGFR4, and FGFrl1. Rat cDNA was used to synthesize probes for FGFR1 and FGFR2 with 90% and 93% homology to human, respectively. Paraffin sections of retina from macaque fetuses sacrificed at fetal days (Fd) 64, 73, 85, 105, 115, 120, and 165, and postnatal ages 2.5 and 11 years were used to detect FGF receptors by immunohistochemistry and in situ hybridization. PCR showed each of the FGF receptors are expressed in fetal human retina. In situ hybridization indicated that mRNA for each receptor is expressed in all retinal cell layers during development, but most intensely in the ganglion cell layer (GCL). FGFR2 mRNA is reduced in the adult inner (INL) and outer (ONL) nuclear layers, while FGFrl1 mRNA is virtually absent from the adult ONL. FGFR4 mRNA is particularly intense in fetal and adult cone photoreceptors. Immunoreactivity to FGFR1-FGFR4 was detected in the interphotoreceptor matrix in what appeared to be RPE microvilli associated with developing photoreceptor outer segments, and generally is high in the GCL and low in the INL. Different patterns of FGFR3 and FGFR4 immunoreactivities in the outer plexiform layer (OPL) suggest localization of FGFR3 to horizontal cell processes, with FGFR4 being expressed by both horizontal and bipolar cell processes. FGFR1, FGFR3, and FGFR4 immunoreactivities are present in the inner segments and somata of adult cones. The pedicles of developing and adult cones are FGFR1 and FGFR3 immunoreactive, and the basal, synaptic region is FGFR4 immunoreactive. FGFR4 labels cones almost in their entirety from early in development and is not detected in rods. The fibers of Henle are intensely FGFR4 immunoreactive in adult cones. The results show high levels of FGF receptor expression in developing and adult retina. Differential distribution of FGF receptors across developing and adult photoreceptors suggests specific roles for FGF signalling in development and maintenance of photoreceptors, particularly the specialized cones of the fovea.

Harnessing tumor necrosis factor receptors to enhance antitumor activities of drugs.

PubMed

Muntané, Jordi

2011-10-17

Cancer is the second-leading cause of death in the U.S. behind heart disease and over stroke. The hallmarks of cancer comprise six biological capabilities acquired during the multistep development of human tumors. The inhibition of cell death pathways is one of these tumor characteristics which also include sustained proliferative signaling, evading growth suppressor signaling, replicative immortality, angiogenesis, and promotion of invasion and metastasis. Cell death is mediated through death receptor (DR) stimulation initiated by specific ligands that transmit signaling to the cell death machinery or through the participation of mitochondria. Cell death involving DR is mediated by the superfamily of tumor necrosis factor receptor (TNF-R) which includes TNF-R type I, CD95, DR3, TNF-related apoptosis-inducing ligand (TRAIL) receptor-1 (TRAIL-R1) and -2 (TRAIL-R2), DR6, ectodysplasin A (EDA) receptor (EDAR), and the nerve growth factor (NGF) receptor (NGFR). The expression of these receptors in healthy and tumor cells induces treatment side effects that limit the systemic administration of cell death-inducing therapies. The present review is focused on the different therapeutic strategies such as targeted antibodies or small molecules addressed to selective stimulated DR-mediated apoptosis or reduce cell proliferation in cancer cells.

Enhancement of high glucose-induced PINK1 expression by melatonin stimulates neuronal cell survival: Involvement of MT2 /Akt/NF-κB pathway.

PubMed

Onphachanh, Xaykham; Lee, Hyun Jik; Lim, Jae Ryong; Jung, Young Hyun; Kim, Jun Sung; Chae, Chang Woo; Lee, Sei-Jung; Gabr, Amr Ahmed; Han, Ho Jae

2017-09-01

Hyperglycemia is a representative hallmark and risk factor for diabetes mellitus (DM) and is closely linked to DM-associated neuronal cell death. Previous investigators reported on a genome-wide association study and showed relationships between DM and melatonin receptor (MT), highlighting the role of MT signaling by assessing melatonin in DM. However, the role of MT signaling in DM pathogenesis is unclear. Therefore, we investigated the role of mitophagy regulators in high glucose-induced neuronal cell death and the effect of melatonin against high glucose-induced mitophagy regulators in neuronal cells. In our results, high glucose significantly increased PTEN-induced putative kinase 1 (PINK1) and LC-3B expressions; as well it decreased cytochrome c oxidase subunit 4 expression and Mitotracker™ fluorescence intensity. Silencing of PINK1 induced mitochondrial reactive oxygen species (ROS) accumulation and mitochondrial membrane potential impairment, increased expressions of cleaved caspases, and increased the number of annexin V-positive cells. In addition, high glucose-stimulated melatonin receptor 1B (MTNR1B) mRNA and PINK1 expressions were reversed by ROS scavenger N-acetyl cysteine pretreatment. Upregulation of PINK1 expression in neuronal cells is suppressed by pretreatment with MT 2 receptor-specific inhibitor 4-P-PDOT. We further showed melatonin stimulated Akt phosphorylation, which was followed by nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) phosphorylation and nuclear translocation. Silencing of PINK1 expression abolished melatonin-regulated mitochondrial ROS production, cleaved caspase-3 and caspase-9 expressions, and the number of annexin V-positive cells. In conclusion, we have demonstrated the melatonin stimulates PINK1 expression via an MT 2 /Akt/NF-κB pathway, and such stimulation is important for the prevention of neuronal cell apoptosis under high glucose conditions. © 2017 The Authors. Journal of Pineal Research Published by John Wiley & Sons Ltd.

Early growth response 2 and Egr3 are unique regulators in immune system.

PubMed

Taefehshokr, Sina; Key, Yashar Azari; Khakpour, Mansour; Dadebighlu, Pourya; Oveisi, Amin

2017-01-01

The immune system is evolved to defend the body against pathogens and is composed of thousands of complicated and intertwined pathways, which are highly controlled by processes such as transcription and repression of cellular genes. Sometimes the immune system malfunctions and a break down in self-tolerance occurs. This lead to the inability to distinguish between self and non-self and cause attacks on host tissues, a condition also known as autoimmunity, which can result in chronic debilitating diseases. Early growth response genes are family of transcription factors comprising of four members, Egr1, Egr2, Egr3 and Egr4. All of which contain three cyc2-His2 zinc fingers. Initially, Egr2 function was identified in the regulation of peripheral nerve myelination, hindbrain segmentation. Egr3, on the other hand, is highly expressed in muscle spindle development. Egr2 and Egr3 are induced due to the antigen stimulation and this signaling is implemented through the B and T cell receptors in the adaptive immunity. T cell receptor signaling plays a key role in Egr 2 and 3 expressions via their interaction with NFAT molecules. Egr 2 and 3 play a crucial role in regulation of the immune system and their involvement in B and T cell activation, anergy induction and preventing the autoimmune disease has been investigated. The deficiency of these transcription factors has been associated to deficient Cbl-b expression, a resistant to anergy phenotype, and expression of effector and activated T cells.

Stat3-induced S1PR1 expression is critical for persistent Stat3 activation in tumors

PubMed Central

Lee, Heehyoung; Deng, Jiehui; Kujawski, Maciej; Yang, Chunmei; Liu, Yong; Herrmann, Andreas; Kortylewski, Marcin; Horne, David; Somlo, George; Forman, Stephen; Jove, Richard; Yu, Hua

2011-01-01

IL-6/Jak2 signaling is viewed critical for persistent Stat3 activation in cancer. However, IL-6-induced Stat3 activity is transient in normal physiology. Here we identify a mechanism important for persistent Stat3 activation in tumor cells and the tumor microenvironment. We show that sphingosine-1-phosphate receptor 1 (S1PR1), a G-protein-coupled receptor for lysophospholipid sphingosine-1-phosphate (S1P), is elevated in Stat3-positive tumors. Stat3 is a transcription factor for the S1pr1 gene. Enhanced S1pr1 expression activates Stat3 and upregulates Il6 gene expression, thereby accelerating tumor growth and metastasis. Conversely, silencing S1pr1 in tumor cells or immune cells inhibits tumor Stat3 activity, tumor growth and metastasis. S1P/S1PR1-induced Stat3 activation is persistent, in contrast to transient Stat3 activation by IL-6. S1PR1 activates Stat3 in part by upregulating Jak2 tyrosine kinase activity. We demonstrate that Stat3-induced S1pr1 expression, as well as S1P/S1PR1 pathway, is important for persistent Stat3 activation in cancer cells and the tumor microenvironment and for malignant progression. PMID:21102457

Expression Profiling of Nuclear Receptors Identifies Key Roles of NR4A Subfamily in Uterine Fibroids

PubMed Central

Yin, Hanwei; Lo, Jay H.; Kim, Ji-Young; Marsh, Erica E.; Kim, J. Julie; Ghosh, Asish K.; Bulun, Serdar

2013-01-01

Uterine fibroids (UFs), also known as uterine leiomyomas, are benign, fibrotic smooth muscle tumors. Although the GnRH analog leuprolide acetate that suppresses gonadal steroid hormones is used as a treatment, it has significant side effects, thereby limiting its use. Availability of more effective therapy is limited because of a lack of understanding of molecular underpinnings of the disease. Although ovarian steroid hormones estrogen and progesterone and their receptors are clearly involved, the role of other nuclear receptors (NRs) in UFs is not well defined. We used quantitative real-time PCR to systematically profile the expression of 48 NRs and identified several NRs that were aberrantly expressed in UFs. Among others, expression of NR4A subfamily members including NGFIB (NR4A1), NURR1 (NR4A2), and NOR1 (NR4A3) were dramatically suppressed in leiomyoma compared with the matched myometrium. Restoration of expression of each of these NR4A members in the primary leiomyoma smooth muscle cells decreased cell proliferation. Importantly, NR4As regulate expressions of the profibrotic factors including TGFβ3 and SMAD3, and several collagens that are key components of the extracellular matrix. Finally, we identify NR4A members as targets of leuprolide acetate treatment. Together, our results implicate several NRs including the NR4A subfamily in leiomyoma etiology and identify NR4As as potential therapeutic targets for treating fibrotic diseases. PMID:23550059

Orphan nuclear receptor small heterodimer partner inhibits transforming growth factor-beta signaling by repressing SMAD3 transactivation.

PubMed

Suh, Ji Ho; Huang, Jiansheng; Park, Yun-Yong; Seong, Hyun-A; Kim, Dongwook; Shong, Minho; Ha, Hyunjung; Lee, In-Kyu; Lee, Keesook; Wang, Li; Choi, Hueng-Sik

2006-12-22

Orphan nuclear receptor small heterodimer partner (SHP) is an atypical member of the nuclear receptor superfamily; SHP regulates the nuclear receptor-mediated transcription of target genes but lacks a conventional DNA binding domain. In this study, we demonstrate that SHP represses transforming growth factor-beta (TGF-beta)-induced gene expression through a direct interaction with Smad, a transducer of TGF-beta signaling. Transient transfection studies demonstrate that SHP represses Smad3-induced transcription. In vivo and in vitro protein interaction assays revealed that SHP directly interacts with Smad2 and Smad3 but not with Smad4. Mapping of domains mediating the interaction between SHP and Smad3 showed that the entire N-terminal domain (1-159 amino acids) of SHP and the linker domain of Smad3 are involved in this interaction. In vitro glutathione S-transferase pulldown competition experiments revealed the SHP-mediated repression of Smad3 transactivation through competition with its co-activator p300. SHP also inhibits the activation of endogenous TGF-beta-responsive gene promoters, the p21, Smad7, and plasminogen activator inhibitor-1 (PAI-1) promoters. Moreover, adenovirus-mediated overexpression of SHP decreases PAI-1 mRNA levels, and down-regulation of SHP by a small interfering RNA increases both the transactivation of Smad3 and the PAI-1 mRNA levels. Finally, the PAI-1 gene is expressed in SHP(-/-) mouse hepatocytes at a higher level than in normal hepatocytes. Taken together, these data indicate that SHP is a novel co-regulator of Smad3, and this study provides new insights into regulation of TGF-beta signaling.

Role of decoy molecules in neuronal ischemic preconditioning

PubMed Central

Panneerselvam, Mathivadhani; Patel, Piyush M.; Roth, David M.; Kidd, Michael W.; Chin-Lee, Blake; Head, Brian P.; Niesman, Ingrid R.; Inoue, Satoki; Patel, Hemal H.; Davis, Daniel P.

2011-01-01

Decoy receptors bind with TNF related apoptosis inducing ligands (TRAIL) but do not contain the cytoplasmic domains necessary to transduce apoptotic signals. We hypothesized that decoy receptors may confer neuronal protection against lethal ischemia after ischemic preconditioning (IPC). Mixed cortical neurons were exposed to IPC one day prior to TRAIL treatment or lethal ischemia. IPC increased decoy receptor but reduced death receptor expression compared to lethal ischemia. IPC-induced increase in decoy receptor expression was reduced by prior treatment with CAPE, a nuclear factor-kappa B inhibitor (NFκB). Expression of decoy molecules, dependent on NFκB, may mediate neuronal survival induced by IPC. PMID:21315738

Hypoxia attenuates purinergic P2X receptor-induced inflammatory gene expression in brainstem microglia

PubMed Central

Smith, Stephanie MC; Mitchell, Gordon S; Friedle, Scott A; Sibigtroth, Christine M; Vinit, Stéphane; Watters, Jyoti J

2013-01-01

Hypoxia and increased extracellular nucleotides are frequently coincident in the brainstem. Extracellular nucleotides are potent modulators of microglial inflammatory gene expression via P2X purinergic receptor activation. Although hypoxia is also known to modulate inflammatory gene expression, little is known about how hypoxia or P2X receptor activation alone affects inflammatory molecule production in brainstem microglia, nor how hypoxia and P2X receptor signaling interact when they occur together. In the study reported here, we investigated the ability of a brief episode of hypoxia (2 hours) in the presence and absence of the nonselective P2X receptor agonist 2′(3′)-O-(4-benzoylbenzoyl)adenosine-5′-triphosphate (BzATP) to promote inflammatory gene expression in brainstem microglia in adult rats. We evaluated inducible nitric oxide synthase (iNOS), tumor necrosis factor alpha (TNFα), and interleukin (IL)-6 messenger RNA levels in immunomagnetically isolated brainstem microglia. While iNOS and IL-6 gene expression increased with hypoxia and BzATP alone, TNFα expression was unaffected. Surprisingly, BzATP-induced inflammatory effects were lost after hypoxia, suggesting that hypoxia impairs proinflammatory P2X-receptor signaling. We also evaluated the expression of key P2X receptors activated by BzATP, namely P2X1, P2X4, and P2X7. While hypoxia did not alter their expression, BzATP upregulated P2X4 and P2X7 mRNAs; these effects were ablated in hypoxia. Although both P2X4 and P2X7 receptor expression correlated with increased microglial iNOS and IL-6 levels in microglia from normoxic rats, in hypoxia, P2X7 only correlated with IL-6, and P2X4 correlated only with iNOS. In addition, correlations between P2X7 and P2X4 were lost following hypoxia, suggesting that P2X4 and P2X7 receptor signaling differs in normoxia and hypoxia. Together, these data suggest that hypoxia suppresses P2X receptor-induced inflammatory gene expression, indicating a potentially immunosuppressive role of extracellular nucleotides in brainstem microglia following exposure to hypoxia. PMID:24377098

GDNF family receptor α-1 in the catfish: Possible implication to brain dopaminergic activity.

PubMed

Mamta, Sajwan-Khatri; Senthilkumaran, Balasubramanian

2018-05-31

Glial cell line-derived neurotrophic factor (GDNF)is a potent trophic factor that preferentially binds to GDNF family receptor α-1 (GFRα-1)by regulating dopaminergic (DA-ergic) neuronsin brain. Present study aimed to evaluate the significance of GFRα-1 expression during early brain development in catfish. Initially, the full-length cDNA of GFRα-1 was cloned from adult brain which showed high homology with other vertebrate counterparts. Quantitative PCR analysis of tissue distribution revealed ubiquitous expression of GFRα-1 in the tissues analyzed with high levels in female brain and ovary. Significant high expression was evident in brain at 75 and 100 days post hatch females than the respective age-match males. Expression of GFRα-1 was high in brain during the spawning phase when compared to other reproductive phases. Localization of GFRα-1 revealed its presence in preoptic area-hypothalamus which correlated well with the expression profile in discrete areas of brain in adult catfish. Transient silencing of GFRα-1through siRNA lowered expression levels of GFRα-1, which further down regulated the expression of certain brain-specific genes. Expression of GFRα-1 in brain declined significantly upon treatment with the 1-methyl-1,2,3,6-tetrahydropyridinecausing neurodegeneration which further correlated with catecholamines (CA), L-3,4-dihydroxyphenylalanine, DA and norepinephrine levels. Taken together, GFRα-1 plausibly entrains gonadotropin-releasing hormone and gonadotropin axiseither directly or indirectly, at least by partially targeting CA-ergic activity. Copyright © 2018 Elsevier Inc. All rights reserved.

The epidermal growth factor receptor (EGFR) promotes uptake of influenza A viruses (IAV) into host cells.

PubMed

Eierhoff, Thorsten; Hrincius, Eike R; Rescher, Ursula; Ludwig, Stephan; Ehrhardt, Christina

2010-09-09

Influenza A viruses (IAV) bind to sialic-acids at cellular surfaces and enter cells by using endocytotic routes. There is evidence that this process does not occur constitutively but requires induction of specific cellular signals, including activation of PI3K that promotes virus internalization. This implies engagement of cellular signaling receptors during viral entry. Here, we present first indications for an interplay of IAV with receptor tyrosine kinases (RTKs). As representative RTK family-members the epidermal growth factor receptor (EGFR) and the c-Met receptor were studied. Modulation of expression or activity of both RTKs resulted in altered uptake of IAV, showing that these receptors transmit entry relevant signals upon virus binding. More detailed studies on EGFR function revealed that virus binding lead to clustering of lipid-rafts, suggesting that multivalent binding of IAV to cells induces a signaling platform leading to activation of EGFR and other RTKs that in turn facilitates IAV uptake.