Diagnostic and Prognostic Significance of Serum and Tissue Galectin 3 Expression in Patients with Carcinoma of the Bladder

PubMed Central

Gendy, Hoda El; Madkour, Bothina; Abdelaty, Sara; Essawy, Fayza; Khattab, Dina; Hammam, Olfat; Nour, Hani H.

2014-01-01

Background Galectins are group of proteins found in the cytoplasm, nucleus, cell surface and extracellular matrix. Galectin 3 (Gal-3) displays pathological expression in a variety of processes such as tumorigenesis. Patients and Method 70 patients classified into the control group, cystitis group, transitional cell carcinoma group, and squamous cell carcinoma group were enrolled in this study which aimed to detect the serum level and the intensity of tissue expression of Gal-3. Results Both serum level and tissue expression of Gal-3 were statistically higher in bladder cancer patients compared to the other groups. Gal-3 level expression increased from low to high grade urothelial tumors, with a statistically significant increase of its level and expression between muscle invasive and non-muscle invasive Ta urothelial tumors. Conclusion The serum Gal-3 level is sensitive and specific for the diagnosis of bladder cancer. The prognostic significance of tissue expression is to be confirmed. PMID:26195948

Podoplanin increases migration and angiogenesis in malignant glioma

PubMed Central

Grau, Stefan J; Trillsch, Fabian; Tonn, Joerg-Christian; Goldbrunner, Roland H; Noessner, Elfriede; Nelson, Peter J; von Luettichau, Irene

2015-01-01

Expression of podoplanin in glial brain tumors is grade dependent. While serving as a marker for tumor progression and modulating invasion in various neoplasms, little is known about podoplanin function in gliomas. Therefore we stably transfected two human glioma cell lines (U373MG and U87MG) with expression plasmids encoding podoplanin. The efficacy of transfection was confirmed by FACS analysis, PCR and immunocytochemistry. Cells were then sorted for highly podoplanin expressing cells (U373Phigh/U87Phigh). Transfection did not influence the production of pro-angiogenic factors including VEGF, VEGF-C and D. Also, expression of VEGF receptors (VEGFR) remained unchanged except for U87Phigh, where a VEGFR3 expression was induced. U373Phigh showed significantly reduced proliferation as compared to mock transfected group. By contrast, podoplanin significantly increased migration and invasion into collagen matrix. Furthermore, conditioned media from Phigh glioma cells strongly induced tube formation on matrigel. In conclusion, podoplanin increased migration of tumor cells and enhanced tube formation activity in endothelial cells independent from VEGF. Thus, podoplanin expression may be an important step in tumor progression. PMID:26339454

Identification of deregulation of apoptosis and cell cycle in neuroendocrine tumors of the lung via NanoString nCounter expression analysis

PubMed Central

Walter, Robert Fred Henry; Werner, Robert; Ting, Saskia; Vollbrecht, Claudia; Theegarten, Dirk; Christoph, Daniel Christian; Schmid, Kurt Werner; Wohlschlaeger, Jeremias; Mairinger, Fabian Dominik

2015-01-01

Background Neuroendocrine tumors of the lung comprise typical (TC) and atypical carcinoids (AC), large-cell neuroendocrine cancer (LCNEC) and small-cell lung cancer (SCLC). Cell cycle and apoptosis are key pathways of multicellular homeostasis and deregulation of these pathways is associated with cancerogenesis. Materials and Methods Sixty representative FFPE-specimens (16 TC, 13 AC, 16 LCNEC and 15 SCLC) were used for mRNA expression analysis using the NanoString technique. Eight genes related to apoptosis and ten genes regulating key points of cell cycle were investigated. Results ASCL1, BCL2, CASP8, CCNE1, CDK1, CDK2, CDKN1A and CDKN2A showed lower expression in carcinoids compared to carcinomas. In contrast, CCNE1 and CDK6 showed elevated expression in carcinoids compared to carcinomas. The calculated BCL2/BAX ratio showed increasing values from TC to SCLC. Between SCLC and LCNEC CDK2, CDKN1B, CDKN2A and PNN expression was significantly different with higher expression in SCLC. Conclusion Carcinoids have increased CDK4/6 and CCND1 expression controlling RB1 phosphorylation via this signaling cascade. CDK2 and CCNE1 were increased in carcinomas showing that these use the opposite way to control RB1. BAX and BCL2 are antagonists in regulating apoptosis. BCL2 expression increased over BAX expression with increasing malignancy of the tumor from TC to SCLC. PMID:26008974

Increased expression of Rab coupling protein in squamous cell carcinoma of the head and neck and its clinical significance

PubMed Central

DAI, YAOZHANG; LIU, YONG; HUANG, DONGHAI; YU, CHANGYUN; CAI, GENGMING; PI, LEIMING; REN, CAIPING; CHEN, GEORGIA Z.; TIAN, YONGQUAN; ZHANG, XIN

2012-01-01

The role of Rab coupling protein (RCP) has not been previously investigated in squamous cell carcinoma of the head and neck (SCCHN). The aim of this study was to explore RCP protein expression and its clinicopathological significance in SCCHN. RCP protein expression in 95 SCCHN samples, 18 vocal nodule epithelia and 16 leukoplakia epithelia samples was analyzed by immunohistochemistry and correlated with clinicopathological parameters and patient outcome. Our data indicated that vocal nodule epithelia, leukoplakia epithelia and SCCHN showed a gradual increase in the expression of RCP protein. RCP overexpression was significantly associated with T classification, clinical staging, lymph node metastasis and recurrence. Survival analysis revealed that a high RCP expression was significantly correlated with shorter overall survival and disease-free survival. In conclusion, RCP protein may contribute to the malignant progression of SCCHN, and serves as a novel prognostic marker in patients with SCCHN. PMID:22783424

Increased expression of Rab coupling protein in squamous cell carcinoma of the head and neck and its clinical significance.

PubMed

Dai, Yaozhang; Liu, Yong; Huang, Donghai; Yu, Changyun; Cai, Gengming; Pi, Leiming; Ren, Caiping; Chen, Georgia Z; Tian, Yongquan; Zhang, Xin

2012-06-01

The role of Rab coupling protein (RCP) has not been previously investigated in squamous cell carcinoma of the head and neck (SCCHN). The aim of this study was to explore RCP protein expression and its clinicopathological significance in SCCHN. RCP protein expression in 95 SCCHN samples, 18 vocal nodule epithelia and 16 leukoplakia epithelia samples was analyzed by immunohistochemistry and correlated with clinicopathological parameters and patient outcome. Our data indicated that vocal nodule epithelia, leukoplakia epithelia and SCCHN showed a gradual increase in the expression of RCP protein. RCP overexpression was significantly associated with T classification, clinical staging, lymph node metastasis and recurrence. Survival analysis revealed that a high RCP expression was significantly correlated with shorter overall survival and disease-free survival. In conclusion, RCP protein may contribute to the malignant progression of SCCHN, and serves as a novel prognostic marker in patients with SCCHN.

Baicalin increases developmental competence of mouse embryos in vitro by inhibiting cellular apoptosis and modulating HSP70 and DNMT expression

PubMed Central

QI, Xiaonan; LI, Huatao; CONG, Xia; WANG, Xin; JIANG, Zhongling; CAO, Rongfeng; TIAN, Wenru

2016-01-01

Scutellaria baicalensis has been effectively used in Chinese traditional medicine to prevent miscarriages. However, little information is available on its mechanism of action. This study is designed specifically to reveal how baicalin, the main effective ingredient of S. baicalensis, improves developmental competence of embryos in vitro, using the mouse as a model. Mouse pronuclear embryos were cultured in KSOM medium supplemented with (0, 2, 4 and 8 μg/ml) baicalin. The results demonstrated that in vitro culture conditions significantly decreased the blastocyst developmental rate and blastocyst quality, possibly due to increased cellular stress and apoptosis. Baicalin (4 µg/ml) significantly increased 2- and 4-cell cleavage rates, morula developmental rate, and blastocyst developmental rate and cell number of in vitro-cultured mouse embryos. Moreover, baicalin increased the expression of Gja1, Cdh1, Bcl-2, and Dnmt3a genes, decreased the expression of Dnmt1 gene, and decreased cellular stress and apoptosis as it decreased the expression of HSP70, CASP3, and BAX and increased BCL-2 expression in blastocysts cultured in vitro. In conclusion, baicalin improves developmental competence of in vitro-cultured mouse embryos through inhibition of cellular apoptosis and HSP70 expression, and improvement of DNA methylation. PMID:27478062

Serglycin as a potential biomarker for glioma: association of serglycin expression, extent of mast cell recruitment and glioblastoma progression

PubMed Central

Roy, Ananya; Attarha, Sanaz; Weishaupt, Holger; Edqvist, Per-Henrik; Swartling, Fredrik J.; Bergqvist, Michael; Siebzehnrubl, Florian A.; Smits, Anja; Pontén, Fredrik; Tchougounova, Elena

2017-01-01

Serglycin is an intracellular proteoglycan with a unique ability to adopt highly divergent structures by glycosylation with variable types of glycosaminoglycans (GAGs) when expressed by different cell types. Serglycin is overexpressed in aggressive cancers suggesting its protumorigenic role. In this study, we explored the expression of serglycin in human glioma and its correlation with survival and immune cell infiltration. We demonstrate that serglycin is expressed in glioma and that increased expression predicts poor survival of patients. Analysis of serglycin expression in a large cohort of low- and high-grade human glioma samples reveals that its expression is grade dependent and is positively correlated with mast cell (MC) infiltration. Moreover, serglycin expression in patient-derived glioma cells is significantly increased upon MC co-culture. This is also accompanied by increased expression of CXCL12, CXCL10, as well as markers of cancer progression, including CD44, ZEB1 and vimentin. In conclusion, these findings indicate the importance of infiltrating MCs in glioma by modulating signaling cascades involving serglycin, CD44 and ZEB1. The present investigation reveals serglycin as a potential prognostic marker for glioma and demonstrates an association with the extent of MC recruitment and glioma progression, uncovering potential future therapeutic opportunities for patients. PMID:28445977

MicroRNA-155 deficiency promotes nephrin acetylation and attenuates renal damage in hyperglycemia-induced nephropathy.

PubMed

Lin, Xu; You, Yanwu; Wang, Jie; Qin, Youling; Huang, Peng; Yang, Fafen

2015-04-01

MiR-155 has been reported to be involved in both innate and adaptive immune responses. But the role of miR-155 in hyperglycemia-induced nephropathy is still unknown. In our current study, 3-month-old male wild-type C57 mice and Mir-155(-/-) mice were used to establish hyperglycemia-induced nephropathy. In our hyperglycemia-induced nephropathy model, the expression of podocyte injury marker desmin was markedly increased in the diabetes group when compared with control. Diabetes also significantly decreased the levels of nephrin and acetylated nephrin, whereas the expression of miR-155 was markedly increased in diabetes group when compared with control. MiR-155(-/-) mice showed significantly increased expression of nephrin, acetylated nephrin, and Wilm's tumor-1 protein (WT-1) when compared with wild-type control. MiR-155 deficiency results in significantly decrease in IL-17A expression both in vivo and in vitro. And the increased expression of WT-1, nephrin, and ac-nephrin was reversed with additional treatment of rmIL-17. Furthermore, we found that the inhibited Th17 differentiation induced by miR-155 deficiency was dependent on increased expression of SOCS1. In conclusion, miR-155 deficiency promotes nephrin acetylation and attenuates renal damage in hyperglycemia-induced nephropathy. This was associated with inhibited IL-17 production through enhancement of SOCS1 expression.

Exercise differentially affects metabolic functions and white adipose tissue in female letrozole- and dihydrotestosterone-induced mouse models of polycystic ovary syndrome.

PubMed

Marcondes, Rodrigo R; Maliqueo, Manuel; Fornes, Romina; Benrick, Anna; Hu, Min; Ivarsson, Niklas; Carlström, Mattias; Cushman, Samuel W; Stenkula, Karin G; Maciel, Gustavo A R; Stener-Victorin, Elisabet

2017-06-15

Here we hypothesized that exercise in dihydrotestosterone (DHT) or letrozole (LET)-induced polycystic ovary syndrome mouse models improves impaired insulin and glucose metabolism, adipose tissue morphology, and expression of genes related to adipogenesis, lipid metabolism, Notch pathway and browning in inguinal and mesenteric fat. DHT-exposed mice had increased body weight, increased number of large mesenteric adipocytes. LET-exposed mice displayed increased body weight and fat mass, decreased insulin sensitivity, increased frequency of small adipocytes and increased expression of genes related to lipolysis in mesenteric fat. In both models, exercise decreased fat mass and inguinal and mesenteric adipose tissue expression of Notch pathway genes, and restored altered mesenteric adipocytes morphology. In conclusion, exercise restored mesenteric adipocytes morphology in DHT- and LET-exposed mice, and insulin sensitivity and mesenteric expression of lipolysis-related genes in LET-exposed mice. Benefits could be explained by downregulation of Notch, and modulation of browning and lipolysis pathways in the adipose tissue. Copyright © 2017 Elsevier B.V. All rights reserved.

Modulation of Progesterone Receptor Isoform Expression in Pregnant Human Myometrium

PubMed Central

2017-01-01

Background. Regulation of myometrial progesterone receptor (PR) expression is an unresolved issue central to understanding the mechanism of functional progesterone withdrawal and initiation of labor in women. Objectives. To determine whether pregnant human myometrium undergoes culture-induced changes in PR isoform expression ex situ and, further, to determine if conditions approaching the in vivo environment stabilise PR isoform expression in culture. Methods. Term nonlaboring human myometrial tissues were cultured under specific conditions: serum supplementation, steroids, stretch, cAMP, PMA, PGF2α, NF-κB inhibitors, or TSA. Following 48 h culture, PR-T, PR-A, and PR-B mRNA levels were determined using qRT-PCR. PR-A/PR-B ratios were calculated. Results. PR-T and PR-A expression and the PR-A/PR-B ratio significantly increased in culture. Steroids prevented the culture-induced increase in PR-T and PR-A expression. Stretch blocked the effects of steroids on PR-T and PR-A expression. PMA further increased the PR-A/PR-B ratio, while TSA blocked culture-induced increases of PR-A expression and the PR-A/PR-B ratio. Conclusion. Human myometrial tissue in culture undergoes changes in PR gene expression consistent with transition toward a laboring phenotype. TSA maintained the nonlaboring PR isoform expression pattern. This suggests that preserving histone and/or nonhistone protein acetylation is critical for maintaining the progesterone dependent quiescent phenotype of human myometrium in culture. PMID:28540297

Hypoxia activates muscle-restricted coiled-coil protein (MURC) expression via transforming growth factor-β in cardiac myocytes.

PubMed

Shyu, Kou-Gi; Cheng, Wen-Pin; Wang, Bao-Wei; Chang, Hang

2014-03-01

The expression of MURC (muscle-restricted coiled-coil protein), a hypertrophy-regulated gene, increases during pressure overload. Hypoxia can cause myocardial hypertrophy; however, how hypoxia affects the regulation of MURC in cardiomyocytes undergoing hypertrophy is still unknown. The aim of the present study was to test the hypothesis that hypoxia induces MURC expression in cardiomyocytes during hypertrophy. The expression of MURC was evaluated in cultured rat neonatal cardiomyocytes subjected to hypoxia and in an in vivo model of AMI (acute myocardial infarction) to induce myocardial hypoxia in adult rats. MURC protein and mRNA expression were significantly enhanced by hypoxia. MURC proteins induced by hypoxia were significantly blocked after the addition of PD98059 or ERK (extracellular-signal-regulated kinase) siRNA 30 min before hypoxia. Gel-shift assay showed increased DNA-binding activity of SRF (serum response factor) after hypoxia. PD98059, ERK siRNA and an anti-TGF-β (transforming growth factor-β) antibody abolished the SRF-binding activity enhanced by hypoxia or exogenous administration of TGF-β. A luciferase promoter assay demonstrated increased transcriptional activity of SRF in cardiomyocytes by hypoxia. Increased βMHC (β-myosin heavy chain) and BNP (B-type natriuretic peptide) protein expression and increased protein synthesis was identified after hypoxia with the presence of MURC in hypertrophic cardiomyocytes. MURC siRNA inhibited the hypertrophic marker protein expression and protein synthesis induced by hypoxia. AMI in adult rats also demonstrated increased MURC protein expression in the left ventricular myocardium. In conclusion, hypoxia in cultured rat neonatal cardiomyocytes increased MURC expression via the induction of TGF-β, SRF and the ERK pathway. These findings suggest that MURC plays a role in hypoxia-induced hypertrophy in cardiomyocytes.

Immunophenotyping of Monocytes During Human Sepsis Shows Impairment in Antigen Presentation: A Shift Toward Nonclassical Differentiation and Upregulation of FcγRi-Receptor.

PubMed

Ferreira da Mota, Nadijane Valeria; Brunialti, Milena Karina Colo; Santos, Sidneia Sousa; Machado, Flavia Ribeiro; Assunçao, Murillo; de Azevedo, Luciano Cesar Pontes; Salomao, Reinaldo

2017-12-05

Monocytes and macrophages are pivotal in the host response to sepsis, recognizing the infecting microorganism and triggering an inflammatory response. These functions are, at least in part, modulated by the expression of cell surface receptors. We aimed to characterize the monocyte phenotype from septic patients during an ongoing sepsis process and its association with clinical outcomes. Sixty-one septic patients and 31 healthy volunteers (HVs) were enrolled in the study. Samples were obtained from patients at baseline (D0, N = 61), and after 7 (D7, N = 36) and 14 days of therapy (D14, N = 22). Monocytes from septic patients presented decreased expression of CD86, HLA-DR, CD200R, CCR2, CXCR2, and CD163 compared with HV monocytes. In contrast, the PD-1, PD-L1, CD206, CD64, and CD16 expression levels were upregulated in patients. HLA-DR, CD64, PD-1, and PD-L1 expression levels were higher in survivors than in nonsurvivors. Increased CD86, HLA-DR, and CXCR2 expression levels were observed in follow-up samples; in contrast, CD64 and CD16 GMFI decreased over time. In conclusion, monocytes from septic patients show antigen presentation impairment as characterized by decreased HLA-DR and costimulatory CD86 expression and increased PD-1 and PD-L1 expression. On the contrary, increased monocyte inflammatory and phagocytic activities may be inferred by the increased CD16 and CD64 expression. We found conflicting results regarding differentiation toward the M2 phenotype, with increased CD206 expression and decreased CD163 expression on monocytes from septic patients, whereas the subset of nonclassical monocytes was demonstrated by increased CD16.

Estrogen Receptor Expression in Atypical Hyperplasia: Lack of Association with Breast Cancer

PubMed Central

Barr Fritcher, Emily G.; Degnim, Amy C.; Hartmann, Lynn C.; Radisky, Derek C.; Boughey, Judy C.; Anderson, Stephanie S.; Vierkant, Robert A.; Frost, Marlene H.; Visscher, Daniel W.; Reynolds, Carol

2011-01-01

Background Estrogen receptor (ER) is expressed in normal and malignant breast epithelium, and expression levels have been found to increase with age in normal breast epithelium but not in atypical hyperplasia (AH) and carcinoma in situ. Here we assess ER expression in AH and its association with later breast cancer. Methods ER expression was assessed immunohistochemically in archival sections from 246 women with AH who had open benign breast biopsy from 1967–1991. The ACISRIII (Dako, Carpinteria, CA) was utilized to calculate ER expression in all atypical foci. Using multivariate linear regression, we examined associations of ER expression with age at biopsy, indication for biopsy, type of atypia, number of atypical foci, involution status, and family history. Breast cancer risk across levels of ER expression was also assessed compared to the Iowa SEER control population. Results Among 246 women, 87 (35%) had atypical ductal hyperplasia (ADH), 141 (57%) had atypical lobular hyperplasia (ALH), and 18 (7%) had both. Forty-nine (20%) developed breast cancer (median follow-up of 14.4 years). Multivariate analysis indicated that type of atypia and age at diagnosis were significantly associated with ER percent staining and intensity [p<0.05]. ER expression was increased in women with ADH and/or those over age 55. ER expression did not significantly impact breast cancer risk in patients diagnosed with atypia. Conclusion We found increasing ER expression in atypical hyperplasia with increasing age. ER expression in atypical hyperplasia does not further discriminate breast cancer risk in women with atypia. PMID:21209395

Protease Activated Receptor-2 Expression and Function in Asthmatic Bronchial Smooth Muscle

PubMed Central

Gilbert, Guillaume; Carvalho, Gabrielle; Trian, Thomas; Ozier, Annaig; Gillibert-Duplantier, Jennifer; Ousova, Olga; Maurat, Elise; Thumerel, Matthieu; Quignard, Jean-François; Girodet, Pierre-Olivier; Marthan, Roger; Berger, Patrick

2014-01-01

Asthmatic bronchial smooth muscle (BSM) is characterized by structural remodeling associated with mast cell infiltration displaying features of chronic degranulation. Mast cell-derived tryptase can activate protease activated receptor type-2 (PAR-2) of BSM cells. The aims of the present study were (i) to evaluate the expression of PAR-2 in both asthmatic and non asthmatic BSM cells and, (ii) to analyze the effect of prolonged stimulation of PAR-2 in asthmatic BSM cells on cell signaling and proliferation. BSM cells were obtained from both 33 control subjects and 22 asthmatic patients. PAR-2 expression was assessed by flow cytometry, western blot and quantitative RT-PCR. Calcium response, transduction pathways and proliferation were evaluated before and following PAR-2 stimulation by SLIGKV-NH2 or trypsin for 1 to 3 days. Asthmatic BSM cells expressed higher basal levels of functional PAR-2 compared to controls in terms of mRNA, protein expression and calcium response. When PAR-2 expression was increased by means of lentivirus in control BSM cells to a level similar to that of asthmatic cells, PAR-2-induced calcium response was then similar in both types of cell. However, repeated PAR-2 stimulations increased the proliferation of asthmatic BSM cells but not that of control BSM cells even following lentiviral over-expression of PAR-2. Such an increased proliferation was related to an increased phosphorylation of ERK in asthmatic BSM cells. In conclusion, we have demonstrated that asthmatic BSM cells express increased baseline levels of functional PAR-2. This higher basal level of PAR-2 accounts for the increased calcium response to PAR-2 stimulation, whereas the increased proliferation to repeated PAR-2 stimulation is related to increased ERK phosphorylation. PMID:24551046

Erythropoietin over-expression protects against diet-induced obesity in mice through increased fat oxidation in muscles.

PubMed

Hojman, Pernille; Brolin, Camilla; Gissel, Hanne; Brandt, Claus; Zerahn, Bo; Pedersen, Bente Klarlund; Gehl, Julie

2009-06-12

Erythropoietin can be over-expressed in skeletal muscles by gene electrotransfer, resulting in 100-fold increase in serum EPO and significant increases in haemoglobin levels. Earlier studies have suggested that EPO improves several metabolic parameters when administered to chronically ill kidney patients. Thus we applied the EPO over-expression model to investigate the metabolic effect of EPO in vivo.At 12 weeks, EPO expression resulted in a 23% weight reduction (P<0.01) in EPO transfected obese mice; thus the mice weighed 21.9+/-0.8 g (control, normal diet,) 21.9+/-1.4 g (EPO, normal diet), 35.3+/-3.3 g (control, high-fat diet) and 28.8+/-2.6 g (EPO, high-fat diet). Correspondingly, DXA scanning revealed that this was due to a 28% reduction in adipose tissue mass.The decrease in adipose tissue mass was accompanied by a complete normalisation of fasting insulin levels and glucose tolerance in the high-fat fed mice. EPO expression also induced a 14% increase in muscle volume and a 25% increase in vascularisation of the EPO transfected muscle. Muscle force and stamina were not affected by EPO expression. PCR array analysis revealed that genes involved in lipid metabolism, thermogenesis and inflammation were increased in muscles in response to EPO expression, while genes involved in glucose metabolism were down-regulated. In addition, muscular fat oxidation was increased 1.8-fold in both the EPO transfected and contralateral muscles.In conclusion, we have shown that EPO when expressed in supra-physiological levels has substantial metabolic effects including protection against diet-induced obesity and normalisation of glucose sensitivity associated with a shift to increased fat metabolism in the muscles.

Erythropoietin Over-Expression Protects against Diet-Induced Obesity in Mice through Increased Fat Oxidation in Muscles

PubMed Central

Hojman, Pernille; Brolin, Camilla; Gissel, Hanne; Brandt, Claus; Zerahn, Bo; Pedersen, Bente Klarlund; Gehl, Julie

2009-01-01

Erythropoietin can be over-expressed in skeletal muscles by gene electrotransfer, resulting in 100-fold increase in serum EPO and significant increases in haemoglobin levels. Earlier studies have suggested that EPO improves several metabolic parameters when administered to chronically ill kidney patients. Thus we applied the EPO over-expression model to investigate the metabolic effect of EPO in vivo. At 12 weeks, EPO expression resulted in a 23% weight reduction (P<0.01) in EPO transfected obese mice; thus the mice weighed 21.9±0.8 g (control, normal diet,) 21.9±1.4 g (EPO, normal diet), 35.3±3.3 g (control, high-fat diet) and 28.8±2.6 g (EPO, high-fat diet). Correspondingly, DXA scanning revealed that this was due to a 28% reduction in adipose tissue mass. The decrease in adipose tissue mass was accompanied by a complete normalisation of fasting insulin levels and glucose tolerance in the high-fat fed mice. EPO expression also induced a 14% increase in muscle volume and a 25% increase in vascularisation of the EPO transfected muscle. Muscle force and stamina were not affected by EPO expression. PCR array analysis revealed that genes involved in lipid metabolism, thermogenesis and inflammation were increased in muscles in response to EPO expression, while genes involved in glucose metabolism were down-regulated. In addition, muscular fat oxidation was increased 1.8-fold in both the EPO transfected and contralateral muscles. In conclusion, we have shown that EPO when expressed in supra-physiological levels has substantial metabolic effects including protection against diet-induced obesity and normalisation of glucose sensitivity associated with a shift to increased fat metabolism in the muscles. PMID:19521513

CXCR4 expression in papillary thyroid carcinoma: induction by nitric oxide and correlation with lymph node metastasis

PubMed Central

Yasuoka, Hironao; Kodama, Rieko; Hirokawa, Mitsuyoshi; Takamura, Yuuki; Miyauchi, Akira; Sanke, Tokio; Nakamura, Yasushi

2008-01-01

Background Metastasis to regional lymph nodes is a common step in the progression of cancer. Recent evidence suggests that tumor production of CXCR4 promotes lymph node metastasis. Nitric oxide (NO) may also increase metastatic ability in human cancers. Methods Nitrite/nitrate levels and functional CXCR4 expression were assessed in K1 and B-CPAP papillary thyroid carcinoma (PTC) cells after induction and/or inhibition of NO synthesis. CXCR4 expression was also analyzed in primary human PTC. The relationship between nitrotyrosine levels, which are a biomarker for peroxynitrate formation from NO in vivo, CXCR4 expression, and lymph node status was also analyzed. Results Production of nitrite/nitrate and functional CXCR4 expression in both cell lines was increased by treatment with the NO donor DETA NONOate. The NOS inhibitor L-NAME eliminated this increase. Positive CXCR4 immunostaining was observed in 60.7% (34/56) of PTCs. CXCR4 expression was significantly correlated with nitrotyrosine levels and lymph node metastasis in human PTC. Conclusion Our data indicate that NO stimulates CXCR4 expression in vitro. Formation of the NO biomarker nitrotyrosine was also correlated with CXCR4 expression and lymph node metastasis in human PTC. NO may induce lymph node metastasis via CXCR4 induction in papillary thyroid carcinoma. PMID:18826577

Elevated expression of pleiotrophin in pilocarpine-induced seizures of immature rats and in pentylenetetrazole-induced hippocampal astrocytes in vitro.

PubMed

Zhang, Shuqin; Liang, Feng; Wang, Bing; Le, Yuan; Wang, Hua

2014-03-01

Pleiotrophin (PTN) is a secreted extracellular matrix (ECM)-associated cytokine that has emerged as an important neuromodulator with multiple neuronal functions. In the present study, we detected and compared the dynamic expression of PTN in the hippocampus and adjacent cortex of immature rats with pilocarpine-induced epilepsy. Moreover, we also confirmed the results by examining PTN expression in hippocampal astrocytes cultured in the presence of pentylenetetrazole (PTZ). Immunohistochemistry showed faint immunostaining of PTN in the control hippocampus and adjacent cortex. Notably, PTN immunoreactivity began to increase in relatively small cells in the hippocampus and adjacent cortex at 2h and 3 weeks after seizures, and the labeling intensity reached the maximum level in the hippocampus and adjacent cortex at 8 weeks after seizures. Furthermore, we also found that PTZ treatment significantly reduced astrocytic viability in a dose-dependent manner and time-dependently increased expression levels of PTN in hippocampal astrocytes. In conclusion, our data suggest that increased expression of PTN in the brain tissues may be involved in epileptogenesis. Copyright © 2013 Elsevier GmbH. All rights reserved.

Dietary nickel chloride induces oxidative stress, apoptosis and alters Bax/Bcl-2 and caspase-3 mRNA expression in the cecal tonsil of broilers.

PubMed

Wu, Bangyuan; Cui, Hengmin; Peng, Xi; Fang, Jing; Zuo, Zhicai; Deng, Junliang; Huang, Jianying

2014-01-01

The purpose of this study was to investigate the effects of dietary NiCl2 on antioxidant function, apoptosis, and the protein expression, mRNA expression and contents of the bcl-2, bax and caspase-3 in the cecal tonsil of broilers. 280 one-day-old avian broilers were divided into four groups and fed on a corn-soybean basal diet as control diet or the same basal diet supplemented with 300, 600 and 900 mg/kg of NiCl2 for 42 days. The activities of SOD, CAT and GSH-Px, and the ability to inhibit hydroxy radical, and GSH content were significantly decreased in all experimental groups. MDA content was significantly increased. The protein expression, mRNA expression and contents of bcl-2 were decreased, and bax and caspase-3 were increased in all experimental groups. The percentages of apoptotic lymphocytes were significantly increased. In conclusion, dietary NiCl2 in excess of 300 mg/kg caused oxidative stress, and then induced decreased the protein expression, mRNA expression and the contents of bcl-2, and increased protein expression, mRNA expression and the contents of bax and caspase-3 proteins in the cecal tonsil. The local intestinal mucosal immunity could finally be impaired due to the oxidative stress and apoptosis in the cecal tonsil caused by NiCl2. Copyright © 2013 Elsevier Ltd. All rights reserved.

Study of the expression and function of ACY1 in patients with colorectal cancer.

PubMed

Yu, Bing; Liu, Xuezhong; Cao, Xiuzhen; Zhang, Mingyue; Chang, Hong

2017-04-01

Aminoacylase 1 (ACY1) is important for regulating the proliferation of numerous types of cancer. However, the expression and mechanisms underlying the function of ACY1 in colorectal cancer remain unclear. In order to investigate the expression and function of ACY1 in colorectal cancer, tumor tissue and blood samples were collected for analysis from 132 patients diagnosed with colorectal cancer. Reverse transcription-quantitative polymerase chain reaction analysis and western blotting identified significantly increased expression of ACY1 mRNA in colorectal tumor tissue (P<0.05 vs. adjacent normal tissue) and notably increased ACY1 protein levels. This ACY1 mRNA expression was found to be positively correlated with tumor stage. In addition, plasma ACY1 concentration was increased in patients with colorectal cancer compared with healthy controls. Furthermore, in vitro knockdown of ACY1 in human colorectal cancer HT-29 cells was shown to inhibit proliferation and increase apoptosis. This effect was found to be associated with the activation of ERK1 and TGF-β1 signaling. In conclusion, the results of the present study suggest that ACY1 promotes tumor progression, and thus may be a potential target for the diagnosis and treatment of colorectal cancer.

Transcriptional bursting explains the noise–versus–mean relationship in mRNA and protein levels

DOE PAGES

Dar, Roy; Shaffer, Sydney M.; Singh, Abhyudai; ...

2016-07-28

Recent analysis demonstrates that the HIV-1 Long Terminal Repeat (HIV LTR) promoter exhibits a range of possible transcriptional burst sizes and frequencies for any mean-expression level. However, these results have also been interpreted as demonstrating that cell-tocell expression variability (noise) and mean are uncorrelated, a significant deviation from previous results. Here, we re-examine the available mRNA and protein abundance data for the HIV LTR and find that noise in mRNA and protein expression scales inversely with the mean along analytically predicted transcriptional burst-size manifolds. We then experimentally perturb transcriptional activity to test a prediction of the multiple burst-size model: thatmore » increasing burst frequency will cause mRNA noise to decrease along given burst-size lines as mRNA levels increase. In conclusion, the data show that mRNA and protein noise decrease as mean expression increases, supporting the canonical inverse correlation between noise and mean.« less

Nickel chloride (NiCl2) in hepatic toxicity: apoptosis, G2/M cell cycle arrest and inflammatory response

PubMed Central

Guo, Hongrui; Cui, Hengmin; Fang, Jing; Zuo, Zhicai; Deng, Junliang; Wang, Xun; Zhao, Ling; Chen, Kejie; Deng, Jie

2016-01-01

Up to now, the precise mechanism of Ni toxicology is still indistinct. Our aim was to test the apoptosis, cell cycle arrest and inflammatory response mechanism induced by NiCl2 in the liver of broiler chickens. NiCl2 significantly increased hepatic apoptosis. NiCl2 activated mitochondria-mediated apoptotic pathway by decreasing Bcl-2, Bcl-xL, Mcl-1, and increasing Bax, Bak, caspase-3, caspase-9 and PARP mRNA expression. In the Fas-mediated apoptotic pathway, mRNA expression levels of Fas, FasL, caspase-8 were increased. Also, NiCl2 induced ER stress apoptotic pathway by increasing GRP78 and GRP94 mRNA expressions. The ER stress was activated through PERK, IRE1 and ATF6 pathways, which were characterized by increasing eIF2α, ATF4, IRE1, XBP1 and ATF6 mRNA expressions. And, NiCl2 arrested G2/M phase cell cycle by increasing p53, p21 and decreasing cdc2, cyclin B mRNA expressions. Simultaneously, NiCl2 increased TNF-α, IL-1β, IL-6, IL-8 mRNA expressions through NF-κB activation. In conclusion, NiCl2 induces apoptosis through mitochondria, Fas and ER stress-mediated apoptotic pathways and causes cell cycle G2/M phase arrest via p53-dependent pathway and generates inflammatory response by activating NF-κB pathway. PMID:27824316

Scrutinizing the Expression and Blockade of Inhibitory Molecules Expressed on T Cells from Acute Myeloid Leukemia Patients.

PubMed

Abdolmaleki, Mohsen; Mojtabavi, Nazanin; Zavvar, Mahdi; Vaezi, Mohammad; Noorbakhsh, Farshid; Nicknam, Mohammad Hossein

2018-06-01

T cell exhaustion is an immunosuppressive mechanism which occurs in chronic viral infections, solid tumors and hematologic malignancies. Exhausted T cell has increased the expression of inhibitory receptors, and functional impairment. In this study, we investigated the expression from some of those inhibitory receptors being Programmed death 1 (PD-1), T cell immunoglobulin and mucin domain containing molecules 3 (TIM-3) and CD244 on T cells from Iranian acute myeloid leukemia (AML) patients. Peripheral blood samples were collected from Iranian newly diagnosed AML patients and flow cytometric analysis was accomplished for cell surface expression of PD-1, TIM-3, and CD244 on T lymphocytes. Functionality and proliferation assay were done in the presence of anti-PD-1 and anti-CD244 blocking antibodies. Immunophenotyping of T cells showed a significant increase of PD-1 and CD244 expression on CD4+ and CD8+ T cells of AML patients. Whereas blockade of PD1 and CD244 increased the proliferation of CD4+ and CD8+ T lymphocytes of AML patients but IFN-γ production was not significantly increased. In conclusion, our data indicate that CD4+ and CD8+ T cells from AML patients appeared to be exhausted and blockade of some immune checkpoints can improve the proliferation of those cells.

Inhibin removes the inhibitory effects of activin on steroid enzyme expression and androgen production by normal ovarian thecal cells

PubMed Central

Young, J M; McNeilly, A S

2012-01-01

Activin and inhibin are important local modulators of theca cell steroidogenesis in the ovary. Using a serum-free primary theca cell culture system, this study investigated the effects of inhibin on theca cell androgen production and expression of steroidogenic enzymes. Androstenedione secretion from theca cells cultured in media containing activin, inhibin and follistatin was assessed by RIA over 144 h. Activin (1–100 ng/ml) suppressed androstenedione production. Inhibin (1–100 ng/ml) blocked the suppressive effects of added activin, but increased androstenedione production when added alone, suggesting it was blocking endogenous activin produced by theca cells. Addition of SB-431542 (activin receptor inhibitor) and follistatin (500 ng/ml) increased androstenedione production, supporting this concept. Infection of theca cells with adenoviruses expressing inhibitory Smad6 or 7 increased androstenedione secretion, confirming that the suppressive effects of activin required activation of the Smad2/3 pathway. Activin decreased the expression levels of steroidogenic acute regulatory protein (STAR), whereas STAR expression was increased by inhibin and SB-431542, alone and in combination. CYP11A was unaffected. The expression of CYP17 encoding 17α-hydroxylase was unaffected by activin but increased by inhibin and SB-431542, and when added in combination the effect was further enhanced. The expression of 3β-hydroxysteroid dehydrogenase (3β-HSD) was significantly decreased by activin, while inhibin alone and in combination with SB-431542 both potently increased the expression of 3β-HSD. In conclusion, activin suppressed theca cell androstenedione production by decreasing the expression of STAR and 3β-HSD. Inhibin and other blockers of activin action reversed this effect, supporting the concept that endogenous thecal activin modulates androgen production in theca cells. PMID:22082494

Manganese Superoxide Dismutase Is a Promising Target for Enhancing Chemosensitivity of Basal-Like Breast Carcinoma

PubMed Central

Kumar, Alan Prem; Loo, Ser Yue; Shin, Sung Won; Tan, Tuan Zea; Eng, Chon Boon; Singh, Rajeev; Putti, Thomas Choudary; Ong, Chee Wee; Salto-Tellez, Manuel; Goh, Boon Cher; Park, Joo In; Thiery, Jean Paul; Pervaiz, Shazib

2014-01-01

Abstract Aims: Although earlier reports highlighted a tumor suppressor role for manganese superoxide dismutase (MnSOD), recent evidence indicates increased expression in a variety of human cancers including aggressive breast carcinoma. In the present article, we hypothesized that MnSOD expression is significantly amplified in the aggressive breast carcinoma basal subtype, and targeting MnSOD could be an attractive strategy for enhancing chemosensitivity of this highly aggressive breast cancer subtype. Results: Using MDA-MB-231 and BT549 as a model of basal breast cancer cell lines, we show that knockdown of MnSOD decreased the colony-forming ability and sensitized the cells to drug-induced cell death, while drug resistance was associated with increased MnSOD expression. In an attempt to develop a clinically relevant approach to down-regulate MnSOD expression in patients with basal breast carcinoma, we employed activation of the peroxisome proliferator-activated receptor gamma (PPARγ) to repress MnSOD expression; PPARγ activation significantly reduced MnSOD expression, increased chemosensitivity, and inhibited tumor growth. Moreover, as a proof of concept for the clinical use of PPARγ agonists to decrease MnSOD expression, biopsies derived from breast cancer patients who had received synthetic PPARγ ligands as anti-diabetic therapy had significantly reduced MnSOD expression. Finally, we provide evidence to implicate peroxynitrite as the mechanism involved in the increased sensitivity to chemotherapy induced by MnSOD repression. Innovation and Conclusion: These data provide evidence to link increased MnSOD expression with the aggressive basal breast cancer, and underscore the judicious use of PPARγ ligands for specifically down-regulating MnSOD to increase the chemosensitivity of this subtype of breast carcinoma. Antioxid. Redox Signal. 20, 2326–2346. PMID:23964924

Differential Expression of Ccn4 and Other Genes Between Metastatic and Non-metastatic EL4 Mouse Lymphoma Cells

PubMed Central

S. CHAHAL, MANPREET; TERESA KU, H.; ZHANG, ZHIHONG; M. LEGASPI, CHRISTIAN; LUO, ANGELA; M. HOPKINS, MANDI; E. MEIER, KATHRYN

2016-01-01

Background: Previous work characterized variants of the EL4 murine lymphoma cell line. Some are non-metastatic, and others metastatic, in syngenic mice. In addition, metastatic EL4 cells were stably transfected with phospholipase D2 (PLD2), which further enhanced metastasis. Materials and Methods: Microarray analyses of mRNA expression was performed for non-metastatic, metastatic, and PLD2-expressing metastatic EL4 cells. Results: Many differences were observed between non-metastatic and metastatic cell lines. One of the most striking new findings was up-regulation of mRNA for the matricellular protein WNT1-inducible signaling pathway protein 1 (CCN4) in metastatic cells; increased protein expression was verified by immunoblotting and immunocytochemistry. Other differentially expressed genes included those for reproductive homeobox 5 (Rhox5; increased in metastatic) and cystatin 7 (Cst7; decreased in metastatic). Differences between PLD2-expressing and parental cell lines were limited but included the signaling proteins Ras guanyl releasing protein 1 (RGS18; increased with PLD2) and suppressor of cytokine signaling 2 (SOCS2; decreased with PLD2). Conclusion: The results provide insights into signaling pathways potentially involved in conferring metastatic ability on lymphoma cells. PMID:27807066

Reduced expression of brain cannabinoid receptor 1 (Cnr1) is coupled with an increased complementary micro-RNA (miR-26b) in a mouse model of fetal alcohol spectrum disorders

PubMed Central

2013-01-01

Background Prenatal alcohol exposure is known to result in fetal alcohol spectrum disorders, a continuum of physiological, behavioural, and cognitive phenotypes that include increased risk for anxiety and learning-associated disorders. Prenatal alcohol exposure results in life-long disorders that may manifest in part through the induction of long-term gene expression changes, potentially maintained through epigenetic mechanisms. Findings Here we report a decrease in the expression of Canabinoid receptor 1 (Cnr1) and an increase in the expression of the regulatory microRNA miR-26b in the brains of adult mice exposed to ethanol during neurodevelopment. Furthermore, we show that miR-26b has significant complementarity to the 3’-UTR of the Cnr1 transcript, giving it the potential to bind and reduce the level of Cnr1 expression. Conclusions These findings elucidate a mechanism through which some genes show long-term altered expression following prenatal alcohol exposure, leading to persistent alterations to cognitive function and behavioural phenotypes observed in fetal alcohol spectrum disorders. PMID:23915435

Dyslipidemia and Diabetes Increase the OPG/TRAIL Ratio in the Cardiovascular System.

PubMed

Toffoli, Barbara; Fabris, Bruno; Bartelloni, Giacomo; Bossi, Fleur; Bernardi, Stella

2016-01-01

Background . Dyslipidemia and diabetes are two of the most well established risk factors for the development of cardiovascular disease (CVD). Both of them usually activate a complex range of pathogenic pathways leading to organ damage. Here we hypothesized that dyslipidemia and diabetes could affect osteoprotegerin (OPG) and TNF-related apoptosis-inducing ligand (TRAIL) expression in the vessels and the heart. Materials and Methods . Gene and protein expression of OPG, TRAIL, and OPG/TRAIL ratio were quantified in the aorta and the hearts of control mice, dyslipidemic mice, and diabetic mice. Results . Diabetes significantly increased OPG and the OPG/TRAIL ratio expression in the aorta, while dyslipidemia was the major determinant of the changes observed in the heart, where it significantly increased OPG and reduced TRAIL expression, thus increasing cardiac OPG/TRAIL ratio. Conclusions . This work shows that both dyslipidemia and diabetes affect OPG/TRAIL ratio in the cardiovascular system. This could contribute to the changes in circulating OPG/TRAIL which are observed in patients with diabetes and CVD. Most importantly, these changes could mediate/contribute to atherosclerosis development and cardiac remodeling.

Temporal Alterations in Vascular Angiotensin Receptors and Vasomotor Response in Offspring of Protein-restricted Rat Dams

PubMed Central

SATHISHKUMAR, Kunju; BALAKRISHNAN, Meena; CHINNATHAMBI, Vijayakumar; GAO, Haijun; YALLAMPALLI, Chandra

2012-01-01

Objective Examine temporal alterations in vascular angiotensin II (ANG II) receptors (AT1R and AT2R) and determine vascular response to ANG II in growth-restricted offspring. Study design Offspring of pregnant rats fed low-protein (6%) and control (20%) diet were compared. Results Prenatal protein restriction reprogrammed AT1aR mRNA expression in males’ mesenteric arteries to cause 1.7- and 2.3-fold increases at 3 and 6 months of age associated with arterial pressure increases of 10 and 33 mmHg, respectively; however, in females, increased AT1aR expression (2-fold) and arterial pressure (15 mmHg) occurred only at 6 months. Prenatal protein restriction did not affect AT2R expression. Losartan abolished hypertension, suggesting that AT1aR plays a primary role in arterial pressure elevation. Vasoconstriction to ANG II was exaggerated in all protein-restricted offspring, with greater potency and efficacy in males. Conclusion Prenatal protein restriction increased vascular AT1R expression and vasoconstriction to ANG II, possibly contributing to programmed hypertension. PMID:22537420

The enhancement of stress-related memory by glucocorticoids depends on synapsin-Ia/Ib

PubMed Central

Revest, J-M; Kaouane, N; Mondin, M; Le Roux, A; Rougé-Pont, F; Vallée, M; Barik, J; Tronche, F; Desmedt, A; Piazza, P V

2010-01-01

The activation of glucocorticoid receptors (GR) by glucocorticoids increases stress-related memory through the activation of the MAPK signaling pathway and the downstream transcription factor Egr-1. Here, using converging in vitro and in vivo approaches, respectively, GR-expressing cell lines, culture of hippocampal neurons, and GR genetically modified mice (GRNesCre), we identified synapsin-Ia/Ib as one of the effectors of the glucocorticoid signaling cascade. Stress and glucocorticoid-induced activation of the GR modulate synapsin-Ia/Ib through two complementary mechanisms. First, glucocorticoids driving Egr-1 expression increase the expression of synapsin-Ia/Ib, and second, glucocorticoids driving MAPK activation increase its phosphorylation. Finally, we showed that blocking fucosylation of synapsin-Ia/Ib in the hippocampus inhibits its expression and prevents the glucocorticoid-mediated increase in stress-related memory. In conclusion, our data provide a complete molecular pathway (GR/Egr-1/MAPK/Syn-Ia/Ib) through which stress and glucocorticoids enhance the memory of stress-related events and highlight the function of synapsin-Ia/Ib as molecular effector of the behavioral effects of stress. PMID:20368707

Macrophage differentiation increases expression of the ascorbate transporter (SVCT2)

PubMed Central

Qiao, Huan; May, James M.

2013-01-01

To determine whether macrophage differentiation involves increased uptake of vitamin C, or ascorbic acid, we assessed the expression and function of its transporter SVCT2 during phorbol ester-induced differentiation of human-derived THP-1 monocytes. Induction of THP-1 monocyte differentiation by phorbol 12-myristate 13-acetate (PMA) markedly increased SVCT2 mRNA, protein, and function. When ascorbate was present during PMA-induced differentiation, the increase in SVCT2 protein expression was inhibited, but differentiation was enhanced. PMA-induced SVCT2 protein expression was blocked by inhibitors of protein kinase C (PKC), with most of the affect due to the PKCβI and βII isoforms. Activation of MEK/ERK was sustained up to 48 h after PMA treatment, and the inhibitors completely blocked PMA-stimulated SVCT2 protein expression, indicating an exclusive role for the classical MAP kinase pathway. However, inhibitors of NF-κB activation, NADPH oxidase inhibitors, and several antioxidants also partially prevented SVCT2 induction, suggesting diverse distal routes for control of SVCT2 transcription. Both known promoters for the SVCT2 were involved in these effects. In conclusion, PMA-induced monocyte-macrophage differentiation is enhanced by ascorbate and associated with increased expression and function of the SVCT2 protein through a pathway involving sustained activation of PKCβI/II, MAP kinase, NADPH oxidase, and NF-κB. PMID:19232538

Employing conservation of co-expression to improve functional inference

PubMed Central

Daub, Carsten O; Sonnhammer, Erik LL

2008-01-01

Background Observing co-expression between genes suggests that they are functionally coupled. Co-expression of orthologous gene pairs across species may improve function prediction beyond the level achieved in a single species. Results We used orthology between genes of the three different species S. cerevisiae, D. melanogaster, and C. elegans to combine co-expression across two species at a time. This led to increased function prediction accuracy when we incorporated expression data from either of the other two species and even further increased when conservation across both of the two other species was considered at the same time. Employing the conservation across species to incorporate abundant model organism data for the prediction of protein interactions in poorly characterized species constitutes a very powerful annotation method. Conclusion To be able to employ the most suitable co-expression distance measure for our analysis, we evaluated the ability of four popular gene co-expression distance measures to detect biologically relevant interactions between pairs of genes. For the expression datasets employed in our co-expression conservation analysis above, we used the GO and the KEGG PATHWAY databases as gold standards. While the differences between distance measures were small, Spearman correlation showed to give most robust results. PMID:18808668

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.

Functional Erythropoietin Autocrine Loop in Melanoma

PubMed Central

Kumar, Suresh M.; Acs, Geza; Fang, Dong; Herlyn, Meenhard; Elder, David E.; Xu, Xiaowei

2005-01-01

Although erythropoietin (Epo) is a known stimulator of erythropoiesis, recent evidence suggests that its biological functions are not confined to hematopoietic cells. To elucidate the role of Epo and erythropoietin receptor (EpoR) in melanoma, we examined the expression and function of these proteins in melanocytes and melanoma cells. We found increased expression of Epo in melanoma cells compared to melanocyte in vitro. EpoR was also strongly expressed in all of the melanoma cell lines and two of the three melanocyte cell lines examined. Epo expression was significantly higher in melanoma than in benign nevi as determined by immunohistochemistry. Although melanoma cells secreted Epo in normoxic condition in vitro, hypoxia and CoCl2 treatment increased Epo secretion. EpoR in melanoma cells was functional, because exogenous Epo increased melanoma resistance to hypoxic stress, pretreatment of melanoma cells with Epo significantly increased resistance to dacarbazine treatment, and Epo increased the phosphorylation of EpoR, RAF, and MEK. In conclusion, we demonstrated constitutive expression of Epo and EpoR as well as autonomous secretion of Epo by melanoma cells, indicating a novel autocrine loop of Epo in melanoma. The results suggest that the autocrine and paracrine functions of Epo might play a role in malignant transformation of melanocytes and in the survival of melanoma cells in hypoxia and other adverse conditions. PMID:15743794

Epithelial-derived IL-33 promotes intestinal tumorigenesis in Apc Min/+ mice.

PubMed

He, Zhengxiang; Chen, Lili; Souto, Fabricio O; Canasto-Chibuque, Claudia; Bongers, Gerold; Deshpande, Madhura; Harpaz, Noam; Ko, Huaibin M; Kelley, Kevin; Furtado, Glaucia C; Lira, Sergio A

2017-07-14

Increased expression of Interleukin (IL)-33 has been detected in intestinal samples of patients with ulcerative colitis, a condition associated with increased risk for colon cancer, but its role in the development of colorectal cancer has yet to be fully examined. Here, we investigated the role of epithelial expressed IL-33 during development of intestinal tumors. IL-33 expression was detected in epithelial cells in colorectal cancer specimens and in the Apc Min/+ mice. To better understand the role of epithelial-derived IL-33 in the intestinal tumorigenesis, we generated transgenic mice expressing IL-33 in intestinal epithelial cells (V33 mice). V33 Apc Min/+ mice, resulting from the cross of V33 with Apc Min/+ mice, had increased intestinal tumor burden compared with littermate Apc Min/+ mice. Consistently, Apc Min/+ mice deficient for IL-33 receptor (ST2), had reduced polyp burden. Mechanistically, overexpression of IL-33 promoted expansion of ST2 + regulatory T cells, increased Th2 cytokine milieu, and induced alternatively activated macrophages in the gut. IL-33 promoted marked changes in the expression of antimicrobial peptides, and antibiotic treatment of V33 Apc Min/+ mice abrogated the tumor promoting-effects of IL-33 in the colon. In conclusion, elevated IL-33 signaling increases tumor development in the Apc Min/+ mice.

Skin barrier disruption by sodium lauryl sulfate-exposure alters the expressions of involucrin, transglutaminase 1, profilaggrin, and kallikreins during the repair phase in human skin in vivo.

PubMed

Törmä, Hans; Lindberg, Magnus; Berne, Berit

2008-05-01

Detergents are skin irritants affecting keratinocytes. In this study, healthy volunteers were exposed to water (vehicle) and 1% sodium lauryl sulfate (SLS) under occlusive patch tests for 24 hours. The messenger RNA (mRNA) expression of keratinocyte differentiation markers and of enzymes involved in corneodesmosome degradation was examined in skin biopsies (n=8) during the repair phase (6 hours to 7 days postexposure) using real-time reverse-transcription PCR. It was found that the expression of involucrin was increased at 6 hours, but then rapidly normalized. The expression of transglutaminase 1 exhibited a twofold increase after 24 hours in the SLS-exposed skin. Profilaggrin was decreased after 6 hours. Later (4-7 days), the expression in SLS-exposed areas was >50% above than in control areas. An increased and altered immunofluorescence pattern of involucrin, transglutaminase 1, and filaggrin was also found (n=4). At 6 hours post-SLS exposure, the mRNA expression of kallikrein-7 (KLK-7) and kallikrein-5 (KLK-5) was decreased by 50 and 75%, respectively, as compared with control and water-exposed areas. Thereafter, the expression pattern of KLK-7 and KLK-5 was normalized. Changes in protein expression of KLK-5 were also found. In conclusion, SLS-induced skin barrier defects induce altered mRNA expression of keratinocyte differentiation markers and enzymes degrading corneodesmosomes.

Insulin secretion and GLUT-2 expression in undernourished neonate rats.

PubMed

Lopes Da Costa, Célia; Sampaio De Freitas, Marta; Sanchez Moura, Anibal

2004-04-01

In previous studies, we verified increased insulin sensitivity in adult male offspring of lactating rats readjusting to lack of insulin secretion reduction brought about by protein restriction during lactation. The present study aims to evaluate the effects of maternal protein undernutrition during lactation on glucose-induced insulin secretion and GLUT-2 expression in beta-cells of neonate male and female rats. Lactating Wistar rats were given a protein-free diet during the first 10 days and a normal diet (22% of protein) until weaning. The neonates were separated at birth by sex and diet and studied at 4, 8 and 21 days of lactation. Glucose-induced insulin secretion by pancreatic islets was analyzed by radioimmunoassay and GLUT-2 expression in beta-cells by Western blot. Glucose-induced insulin secretion of the undernourished groups was higher than in the control groups except among females. When comparing the male and female groups and the control and undernourished groups, female neonates showed significantly greater insulin secretion than the male group. Also it was noted that undernutrition induced greater GLUT-2 expression. For instance, comparing the undernourished male and female neonates there was an increase in female GLUT-2 expression on day 4. On the other hand, in undernourished male neonates a GLUT-2 expression increased later in lactation. In conclusion, during a short term, maternal undernutrition induces an increase of the glucose-induced insulin secretion only in male neonates and is associated with an increase in GLUT-2 expression in the beta-cell.

Anti-oxidizing effect of the dichloromethane and hexane fractions from Orostachys japonicus in LPS-stimulated RAW 264.7 cells via upregulation of Nrf2 expression and activation of MAPK signaling pathway.

PubMed

Lee, Hyeong-Seon; Lee, Gyeong-Seon; Kim, Seon-Hee; Kim, Hyun-Kyung; Suk, Dong-Hee; Lee, Dong-Seok

2014-02-01

Orostachys japonicus shows various biological activities. However, the molecular mechanisms remain unknown in LPS-stimulated macrophages. Here, we investigated the anti-oxidizing effect of the dichloromethane (DCM) and hexane fractions from O. japonicus (OJD and OJH) against oxidative stress in RAW 264.7 cells stimulated by LPS. OJD and OJH significantly increased the expression of heme oxygenase-1 (HO-1) in a dose- and time-dependent manner. Additionally, it was found that the expression of HO-1 was stimulated by Nrf2 activated via degradation of Keap1. ERK and p38 inhibitors repressed HO-1 induced by OJD and OJH in LPS-stimulated cells, respectively. In conclusion, these results suggest that OJD and OJH may block oxidative damage stimulated by LPS, via increasing the expression of HO-1 and Nrf2, and MAPK signaling pathway.

Expression of fas protein on CD4+T cells irradiated by low level He-Ne

NASA Astrophysics Data System (ADS)

Nie, Fan; Zhu, Jing; Zhang, Hui-Guo

2005-07-01

Objective: To investigate the influence on the Expression of Fas protein on CD4+ T cells irradiated by low level He-Ne laser in the cases of psoriasis. Methods:the expression of CD4+ T Fas protein was determined in the casee of psoriasis(n=5) pre and post-low level laser irradiation(30 min、60min and 120min)by flow cytometry as compared withthe control(n=5). Results:In the cases of psoriasis,the expression of CD4+T FAS protein 21.4+/-3.1% was increased significantly than that of control group 16.8+/-2.1% pre-irradiation, p<0.05in the control,there is no difference between pre and post- irradiation,p>0.05in the cases , the expression of CD4+T Fas protein wae positively corelated to the irradiation times, when the energy density arrived to 22.92J/cm2（60 minutes）and 45.84J/cm2（120minutes）, the expression of CD4+ T Fas protein was increased significantly as compared with pre-irradiation,p<0.05.Conclusion: The expression of CD4+T Fas protein may be increased by low level He-Ne laser irradiation ,the uncontrolled status of apoptosis could be corrected.

Chronic crude garlic-feeding modified adult male rat testicular markers: mechanisms of action

PubMed Central

Hammami, Imen; Amara, Souheila; Benahmed, Mohamed; El May, Michèle V; Mauduit, Claire

2009-01-01

Background Garlic or Allium sativum (As) shows therapeutic effects such as reduction of blood pressure or hypercholesterolemia but side-effects on reproductive functions remain poorly investigated. Because of garlic's chemical complexity, the processing methods and yield in preparations differ in efficacy and safety. In this context, we clarify the mechanisms of action of crushed crude garlic on testicular markers. Methods During one month of treatment, 24 male rats were fed 5%, 10% and 15% crude garlic. Results We showed that crude garlic-feeding induced apoptosis in testicular germ cells (spermatocytes and spermatids). This cell death process was characterized by increased levels of active CASP3 but not CASP6. Expression of the caspase inhibitors BIRC3 and BIRC2 was increased at all doses of As while expression of XIAP and BIRC5 was unchanged. Moreover, expression of the IAP inhibitor DIABLO was increased at doses 10% and 15% of As. The germ cell death process induced by As might be related to a decrease in testosterone production because of the reduced expression of steroidogenic enzymes (Star, Cyp11a, Hsd3b5 and Hsd17b). Evaluation of Sertoli markers showed that TUBB3 and GSTA2 expression was unchanged. In contrast, AMH, RHOX5 and CDKN1B expression was decreased while GATA4 expression was increased. Conclusion In summary, we showed that feeding with crude garlic inhibited Leydig steroidogenic enzyme expression and Sertoli cell markers. These alterations might induce apoptosis in testicular germ cells. PMID:19552815

Mechanism Governing Human Kappa-Opioid Receptor Expression under Desferrioxamine-Induced Hypoxic Mimic Condition in Neuronal NMB Cells

PubMed Central

Babcock, Jennifer; Herrera, Alberto; Coricor, George; Karch, Christopher; Liu, Alexander H.; Rivera-Gines, Aida; Ko, Jane L.

2017-01-01

Cellular adaptation to hypoxia is a protective mechanism for neurons and relevant to cancer. Treatment with desferrioxamine (DFO) to induce hypoxia reduced the viability of human neuronal NMB cells. Surviving/attached cells exhibited profound increases of expression of the human kappa-opioid receptor (hKOR) and hypoxia inducible factor-1α (HIF-1α). The functional relationship between hKOR and HIF-1α was investigated using RT-PCR, Western blot, luciferase reporter, mutagenesis, siRNA and receptor-ligand binding assays. In surviving neurons, DFO increased HIF-1α expression and its amount in the nucleus. DFO also dramatically increased hKOR expression. Two (designated as HIFC and D) out of four potential HIF response elements of the hKOR gene (HIFA–D) synergistically mediated the DFO response. Mutation of both elements completely abolished the DFO-induced effect. The CD11 plasmid (containing HIFC and D with an 11 bp spacing) produced greater augmentation than that of the CD17 plasmid (HIFC and D with a 17 bp-spacing), suggesting that a proper topological interaction of these elements synergistically enhanced the promoter activity. HIF-1α siRNA knocked down the increase of endogenous HIF-1α messages and diminished the DFO-induced increase of hKOR expression. Increased hKOR expression resulted in the up-regulation of hKOR protein. In conclusion, the adaptation of neuronal hKOR under hypoxia was governed by HIF-1, revealing a new mechanism of hKOR regulation. PMID:28117678

Immunological and pathological changes in the placenta during infection with Listeria monocytogenes in pregnant guinea pigs.

PubMed

Irvin, Elizabeth Ann; Williams, Denita; Hamler, Sarah E; Smith, Mary Alice

2008-10-01

Exposure to Listeria monocytogenes during pregnancy can result in spontaneous abortion and stillbirths; however, the mechanisms are unknown. Our objective was to determine the effects of infection on specific inflammatory and anti-inflammatory cytokine mRNA expression and apoptosis in the placenta after infection with L. monocytogenes. Pregnant guinea pigs were treated on gestation day (gd) 35 with 10(8) colony forming units L. monocytogenes and sacrificed on gd 37, 41, 44, or 55. At gd 41, IFN-gamma and IL-2 mRNA expression was significantly decreased in placentas from treated dams (0.0012-fold and 0.131-fold, respectively). At gd 55, TNF-alpha mRNA expression was significantly decreased (0.19-fold), while IFN-gamma mRNA expression was significantly increased (32-fold), and apoptosis was detected in 100% of placentas from treated dams. In conclusion, inflammatory cytokine mRNA expression is altered and apoptosis is increased in the placenta after treatment with L. monocytogenes, and these changes may contribute to fetal death.

Effects of Anger Awareness and Expression Training versus Relaxation Training on Headaches: A Randomized Trial

PubMed Central

Slavin-Spenny, Olga; Lumley, Mark A.; Thakur, Elyse R.; Nevedal, Dana C.; Hijazi, Alaa M.

2013-01-01

Background and purpose Stress contributes to headaches, and effective interventions for headaches routinely include relaxation training (RT) to directly reduce negative emotions and arousal. Yet, suppressing negative emotions, particularly anger, appears to augment pain, and experimental studies suggest that expressing anger may reduce pain. Therefore, we developed and tested anger awareness and expression training (AAET) on people with headaches. Methods Young adults with headaches (N = 147) were randomized to AAET, RT, or a wait-list control. We assessed affect during sessions, and process and outcome variables at baseline and 4 weeks after treatment. Results On process measures, both interventions increased self-efficacy to manage headaches, but only AAET reduced alexithymia and increased emotional processing and assertiveness. Yet, both interventions were equally effective at improving headache outcomes relative to controls. Conclusions Enhancing anger awareness and expression may improve chronic headaches, although not more than RT. Researchers should study which patients are most likely to benefit from emotional expression versus emotional reduction approaches to chronic pain. PMID:23620190

Estradiol, acting through ERα, induces endothelial non-classic renin-angiotensin system increasing angiotensin 1-7 production.

PubMed

Mompeón, Ana; Lázaro-Franco, Macarena; Bueno-Betí, Carlos; Pérez-Cremades, Daniel; Vidal-Gómez, Xavier; Monsalve, Elena; Gironacci, Mariela M; Hermenegildo, Carlos; Novella, Susana

2016-02-15

Intracellular renin-angiotensin system (RAS) can operate independently of the circulating RAS. Estrogens provide protective effects by modulating the RAS. Our aim was to investigate the effect of estradiol (E2) on angiotensin converting enzymes (ACE) 1 and ACE2 expression and activities in human endothelial cells (HUVEC), and the role of estrogen receptors (ER). The results confirmed the presence of active intracellular RAS in HUVEC. Physiological concentrations of E2 induced a concentration-dependent increase of ACE1 and ACE2 mRNA expression and ACE1, but not ACE2, protein levels. ACE1 and ACE2 enzymatic activities were also induced with E2. These effects were mediated through ERα activation, since ER antagonists ICI 182780 and MPP completely abolished the effect of E2. Moreover, the ERα agonist PPT mirrored the E2 effects on ACE1 and ACE2 protein expression and activity. Exposure of endothelial cells to E2 significantly increased Ang-(1-7) production. In conclusion, E2 increases Ang-(1-7) production, through ERα, involving increased ACE1 and ACE2 mRNA expression and activity and ACE1 protein levels. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

Constitutive androstane receptor activation evokes the expression of glycolytic genes

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

Yarushkin, Andrei A.; Kazantseva, Yuliya A.; Prokopyeva, Elena A.

It is well-known that constitutive androstane receptor (CAR) activation by 1,4-bis[2-(3,5-dichloropyridyloxy)]benzene (TCPOBOP) increases the liver-to-body weight ratio. CAR-mediated liver growth is correlated with increased expression of the pleiotropic transcription factor cMyc, which stimulates cell cycle regulatory genes and drives proliferating cells into S phase. Because glycolysis supports cell proliferation and cMyc is essential for the activation of glycolytic genes, we hypothesized that CAR-mediated up-regulation of cMyc in mouse livers might play a role in inducing the expression of glycolytic genes. The aim of the present study was to examine the effect of long-term CAR activation on glycolytic genes in amore » mouse model not subjected to metabolic stress. We demonstrated that long-term CAR activation by TCPOBOP increases expression of cMyc, which was correlated with reduced expression of gluconeogenic genes and up-regulation of glucose transporter, glycolytic and mitochondrial pyruvate metabolising genes. These changes in gene expression after TCPOBOP treatment were strongly correlated with changes in levels of glycolytic intermediates in mouse livers. Moreover, we demonstrated a significant positive regulatory effect of TCPOBOP-activated CAR on both mRNA and protein levels of Pkm2, a master regulator of glucose metabolism and cell proliferation. Thus, our findings provide evidence to support the conclusion that CAR activation initiates a transcriptional program that facilitates the coordinated metabolic activities required for cell proliferation. - Highlights: • CAR-mediated liver growth is correlated with increased expression of cMyc. • CAR activation increased the expression of glycolytic genes in mouse livers. • CAR activation increased the level of Pkm2 in mouse livers.« less

Association of IRF5 polymorphisms with activation of the interferon α pathway

PubMed Central

Rullo, Ornella J; Woo, Jennifer M P; Wu, Hui; Hoftman, Alice D C; Maranian, Paul; Brahn, Brittany A; McCurdy, Deborah; Cantor, Rita M; Tsao, Betty P

2011-01-01

Objective The genetic association of interferon regulatory factor 5 (IRF5) with systemic lupus erythematosus (SLE) susceptibility has been convincingly established. To gain understanding of the effect of IRF5 variation in individuals without SLE, a study was undertaken to examine whether such genetic variation predisposes to activation of the interferon α (IFNα) pathway. Methods Using a computer simulated approach, 14 single nucleotide polymorphisms (SNPs) and haplotypes of IRF5 were tested for association with mRNA expression levels of IRF5, IFNα and IFN-inducible genes and chemokines in lymphoblastoid cell lines (LCLs) from individuals of European (CEU), Han Chinese (CHB), Japanese (JPT) and Yoruba Nigerian (YRI) backgrounds. IFN-inducible gene expression was assessed in LCLs from children with SLE in the presence and absence of IFNα stimulation. Results The major alleles of IRF5 rs13242262 and rs2280714 were associated with increased IRF5 mRNA expression levels in the CEU, CHB+JPT and YRI samples. The minor allele of IRF5 rs10488631 was associated with increased IRF5, IFNα and IFN-inducible chemokine expression in CEU (pc=0.0005, 0.01 and 0.04, respectively). A haplotype containing these risk alleles of rs13242262, rs10488631 and rs2280714 was associated with increased IRF5, IFNα and IFN-inducible chemokine expression in CEU LCLs. In vitro studies showed specific activation of IFN-inducible genes in LCLs by IFNα. Conclusions SNPs of IRF5 in healthy individuals of a number of ethnic groups were associated with increased mRNA expression of IRF5. In European-derived individuals, an IRF5 haplotype was associated with increased IRF5, IFNα and IFN-inducible chemokine expression. Identifying individuals genetically predisposed to increased IFN-inducible gene and chemokine expression may allow early detection of risk for SLE. PMID:19854706

Differential expression of poliovirus receptor, regulator of G-protein signaling 11 and erythrocyte protein band 4.1-like 3 in human granulosa cells during follicular growth and maturation.

PubMed

Barzilay, Eran; Yung, Yuval; Shapira, Lev; Haas, Jigal; Ophir, Libby; Yerushalmi, Gil M; Maman, Ettie; Hourvitz, Ariel

2014-09-01

Poliovirus receptor (PVR), regulator of G-protein signaling-11 (RGS11), and erythrocyte protein band-4.1-like 3 (EPB41L3) have been proposed to function in follicular maturation in mouse models. We have examined their expression in human mural (mGCs) and cumulus granulosa cells (CCs). Expression of PVR and RGS11 in mGCs decreased in medium-sized follicles compared to small follicles of IVM cycles and increased again in large follicles. Luteinization caused decreased expression of both PVR and RGS11. In vitro incubation of mGCs with progesterone-rich conditioned media decreased expression of RGS11 without affecting PVR levels. Inhibition of progesterone signaling enhanced expression of both RGS11 and PVR. Expression in CCs was examined by means of global transcriptome sequencing analysis RGS11 and EPB41L3 increased in CCs during follicular maturation while PVR levels did not change. In conclusion, during human follicular maturation there are significant changes in expression of PVR, RGS11 and EPB41L3, possibly regulated by progesterone.

Histomorphometry and expression of Cdc47 and caspase-3 in hyperthyroid rat uteri and placentas during gestation and postpartum associated with fetal development.

PubMed

Freitas, E S; Leite, E D; Souza, C A; Ocarino, N M; Ferreira, E; Cassali, G D; Gomes, M G; Serakides, R

2007-01-01

In two different experiments, the effects of hyperthyroidism on the histomorphometry and expression of Cdc47 and caspase-3 were evaluated in the uteri and placentas during gestation and postpartum. Fetal development was also evaluated during gestation. In the first experiment, 36 adult female Wistar rats were divided into two groups of 18 animals each: (1) hyperthyroid; and (2) euthyroid (control). Female rats were mated and killed at 7, 14 and 19 days of gestation. Uteri and placentas were weighed and subjected to histomorphometric and immunohistochemical evaluation to determine the expression of Cdc47 and caspase-3. Ovaries were also evaluated for weight and subjected to morphometric analysis. Fetuses were quantified and weighed individually. In the second experiment, 12 adult female Wistar rats were divided into two groups of six animals each: (1) hyperthyroid; and (2) euthyroid (control). Female rats were mated and killed 2 days postpartum. Uteri were evaluated in the same way as for the first experiment. Hyperthyroidism increased ovulation and conception rates without disturbing the size and viability of the fetuses. In the pregnant uteri, hyperthyroidism did not change the thickness of the layers or the expression of Cdc47 and caspase-3. However, in the placentas, hyperthyroidism increased the medium diameter of trophoblast cells, as well as the thickness and the expression of Cdc47 of spongiotrophoblast cells, at 14 days of gestation. During uterine involution, hyperthyroidism significantly increased the expression of Cdc47 and reduced the expression of caspase-3 in the uterine layers. In conclusion, hyperthyroidism increased the conception rate because of an ovulation gain, induced significant placental changes during pregnancy and, in the uterus, increased Cdc47 expression and decreased caspase-3 expression after parturition.

Increased expression of 78 kD glucose-regulated protein promotes cardiomyocyte apoptosis in a rat model of liver cirrhosis

PubMed Central

Zhang, Lili; Zhang, Huiying; Lv, Minli; Jia, Jiantao; Fan, Yimin; Tian, Xiaoxia; Li, Xujiong; Li, Baohong; Ji, Jingquan; Wang, Limin; Zhao, Zhongfu; Han, Dewu; Ji, Cheng

2015-01-01

Aims: This study was to investigate the role and underlying mechanism of 78 kD glucose-regulated protein (GRP78) in cardiomyocyte apoptosis in a rat model of liver cirrhosis. Methods: A rat model of liver cirrhosis was established with multiple pathogenic factors. A total of 42 male SD rats were randomly divided into the liver cirrhosis group and control group. Cardiac structure analysis was performed to assess alterations in cardiac structure. Cardiomyocytes apoptosis was detected by TdT-mediated dUTP nick end labeling method. Expression of GRP78, CCAAT/enhancer-binding protein homologous protein (CHOP), caspase-12, nuclear factor kappa-light-chain-enhancer of activated B cells p65 subunit (NF-κB p65) and B cell lymphoma-2 (Bcl-2) was detected by immunohistochemical staining. Results: The ratios of left ventricular wall thickness to heart weight and heart weight to body weight were significantly increased with the progression of liver cirrhosis (P < 0.05). Apoptosis index of cardiomyocytes was significantly increased with the progression of liver cirrhosis (P < 0.05). The expression levels of GRP78, CHOP and caspase-12 were significantly increased in the progression of liver cirrhosis (P < 0.05). The expression levels of NF-κB p65 and Bcl-2 were highest in the 4-wk liver cirrhosis, and they were decreased in the 6-wk and 8-wk in the progression of liver cirrhosis. GRP78 expression levels were positively correlated with apoptosis index, CHOP and caspase-12 expression levels (P < 0.05). CHOP expression levels were negatively correlated with NF-κB p65 and Bcl-2 expression levels (P < 0.05). Conclusion: Increased expression of GRP78 promotes cardiomyocyte apoptosis in rats with cirrhotic cardiomyopathy. PMID:26464674

A Central Role for Heme Oxygenase-1 in the Control of Intestinal Epithelial Chemokine Expression.

PubMed

Onyiah, Joseph C; Schaefer, Rachel E M; Colgan, Sean P

2018-05-23

In mucosal inflammatory disorders, the protective influence of heme oxygenase-1 (HO-1) and its metabolic byproducts, carbon monoxide (CO) and biliverdin, is a topic of significant interest. Mechanisms under investigation include the regulation of macrophage function and mucosal cytokine expression. While there is an increasing recognition of the importance of epithelial-derived factors in the maintenance of intestinal mucosal homeostasis, the contribution of intestinal epithelial cell (IEC) HO-1 on inflammatory responses has not previously been investigated. We examined the influence of modulating HO-1 expression on the inflammatory response of human IECs. Engineered deficiency of HO-1 in Caco-2 and T84 IECs led to increased proinflammatory chemokine expression in response to pathogenic bacteria and inflammatory cytokine stimulation. Crosstalk with activated leukocytes also led to increased chemokine expression in HO-1-deficient cells in an IL-1β dependent manner. Treatment of Caco-2 cells with a pharmacological inducer of HO-1 led to the inhibition of chemokine expression. Mechanistic studies suggest that HO-1 and HO-1-related transcription factors, but not HO-1 metabolic products, are partly responsible for the influence of HO-1 on chemokine expression. In conclusion, our data identify HO-1 as a central regulator of IEC chemokine expression that may contribute to homeo-stasis in the intestinal mucosa. © 2018 S. Karger AG, Basel.

Induction of Proteinases in the Human Preovulatory Follicle of the Menstrual Cycle by hCG

PubMed Central

Rosewell, Katherine L.; Al-Alem, Linah; Zakerkish, Farnosh; McCord, Lauren; Akin, James W.; Chaffin, Charles L.; Brännström, Mats; Curry, Thomas E.

2014-01-01

Objective To explore the temporal expression in granulosa and theca cells of key members of the MMP and ADAMTS families across the periovulatory period in women in order to gain insight into their possible roles during ovulation and early luteinization. Design Experimental prospective clinical study and laboratory-based investigation. Setting University Medical Center and private IVF center. Animal and Patient(s) Thirty eight premenopausal women undergoing surgery for tubal ligation and 6 premenopausal women undergoing ART. Intervention(s) Administration of hCG and harvesting of follicles by laparoscopy and collection of granulosa-lutein cells at oocyte retrieval. Main Outcome Measure(s) Expression of mRNA for MMPs and ADAMTSs in human granulosa cells and theca cells collected across the periovulatory period of the menstrual cycle and in cultured granulosa-lutein cells after hCG. Localization of MMPs and ADAMTSs by immunohistochemistry. Result(s) Expression of MMP1 and MMP19 mRNA increased in both granulosa and theca cells after hCG administration. ADAMTS1 and ADAMTS 9 mRNA increased in granulosa cells after hCG treatment, however thecal cell expression for ADAMTS1 was unchanged while ADAMTS9 expression was decreased. Expression of MMP8 and MMP13 mRNA was unchanged. Immunohistochemistry confirmed the localization of MMP1, MMP19, ADAMTS1 and ADAMTS9 to the granulosa and thecal cell layers. Conclusion(s) The collection of the dominant follicle throughout the periovulatory period has allowed the identification of proteolytic remodeling enzymes in the granulosa and theca compartments that may be critically involved in human ovulation. These proteinases may work in concert to regulate breakdown of the follicular wall and release of the oocyte. PMID:25516084

Changes in mitochondrial function and mitochondria associated protein expression in response to 2-weeks of high intensity interval training

PubMed Central

Vincent, Grace; Lamon, Séverine; Gant, Nicholas; Vincent, Peter J.; MacDonald, Julia R.; Markworth, James F.; Edge, Johann A.; Hickey, Anthony J. R.

2015-01-01

Purpose: High-intensity short-duration interval training (HIT) stimulates functional and metabolic adaptation in skeletal muscle, but the influence of HIT on mitochondrial function remains poorly studied in humans. Mitochondrial metabolism as well as mitochondrial-associated protein expression were tested in untrained participants performing HIT over a 2-week period. Methods: Eight males performed a single-leg cycling protocol (12 × 1 min intervals at 120% peak power output, 90 s recovery, 4 days/week). Muscle biopsies (vastus lateralis) were taken pre- and post-HIT. Mitochondrial respiration in permeabilized fibers, citrate synthase (CS) activity and protein expression of peroxisome proliferator-activated receptor gamma coactivator (PGC-1α) and respiratory complex components were measured. Results: HIT training improved peak power and time to fatigue. Increases in absolute oxidative phosphorylation (OXPHOS) capacities and CS activity were observed, but not in the ratio of CCO to the electron transport system (CCO/ETS), the respiratory control ratios (RCR-1 and RCR-2) or mitochondrial-associated protein expression. Specific increases in OXPHOS flux were not apparent after normalization to CS, indicating that gross changes mainly resulted from increased mitochondrial mass. Conclusion: Over only 2 weeks HIT significantly increased mitochondrial function in skeletal muscle independently of detectable changes in mitochondrial-associated and mitogenic protein expression. PMID:25759671

Study of the expression and function of ACY1 in patients with colorectal cancer

PubMed Central

Yu, Bing; Liu, Xuezhong; Cao, Xiuzhen; Zhang, Mingyue; Chang, Hong

2017-01-01

Aminoacylase 1 (ACY1) is important for regulating the proliferation of numerous types of cancer. However, the expression and mechanisms underlying the function of ACY1 in colorectal cancer remain unclear. In order to investigate the expression and function of ACY1 in colorectal cancer, tumor tissue and blood samples were collected for analysis from 132 patients diagnosed with colorectal cancer. Reverse transcription-quantitative polymerase chain reaction analysis and western blotting identified significantly increased expression of ACY1 mRNA in colorectal tumor tissue (P<0.05 vs. adjacent normal tissue) and notably increased ACY1 protein levels. This ACY1 mRNA expression was found to be positively correlated with tumor stage. In addition, plasma ACY1 concentration was increased in patients with colorectal cancer compared with healthy controls. Furthermore, in vitro knockdown of ACY1 in human colorectal cancer HT-29 cells was shown to inhibit proliferation and increase apoptosis. This effect was found to be associated with the activation of ERK1 and TGF-β1 signaling. In conclusion, the results of the present study suggest that ACY1 promotes tumor progression, and thus may be a potential target for the diagnosis and treatment of colorectal cancer. PMID:28454420

Ancient Mars: wet in many places.

PubMed

Paige, David A

2005-03-11

New results from the Mars Express Orbiter mission reveal multiple deposits of minerals formed in the presence of liquid water. They reinforce the conclusion that ancient Mars was warmer and wetter than it is today, and increase the number of promising localities to search for evidence of past life.

Upregulation of Cyclooxygenase-2 Expression in Porcine Macula Densa With Chronic Nitric Oxide Synthase Inhibition

PubMed Central

Kommareddy, M.; McAllister, R. M.; Ganjam, V. K.; Turk, J. R.; Laughlin, M. Harold

2012-01-01

The objective of this study was to investigate the effects of chronic inhibition of nitric oxide synthase (NOS) on cyclooxygenase-2 (COX-2) expression in the macula densa (MD) of swine, as well as the effects on expression of related proteins. Adult female Yucatan swine were given either tap water (control, n = 6) or water with NG-nitro-l-arginine methyl ester (L-NAME, 100 mg/liter, n = 5) for a minimum of 30 days. Duplicate samples of kidney were fixed or snap frozen. There was a significant (P = .0082) upregulation of COX-2 mRNA expression in the MD of L-NAME, as well as an apparent increase in COX-2 protein. Plasma renin activity also increased with L-NAME treatment (control, 0.34 ± 0.08 ng/ml; L-NAME, 1.26 ± 0.03 ng/ml; P = .00000003). There were no differences between groups in expression of either inducible NOS or renin protein or in serum electrolyte concentrations. In conclusion, with chronic inhibition of NOS, COX-2 in MD is upregulated, perhaps to compensate for loss of nitric oxide. Increases in COX-2 products may counteract renal arteriolar constriction and sustain renin release. PMID:21160023

Upregulation of cyclooxygenase-2 expression in porcine macula densa with chronic nitric oxide synthase inhibition.

PubMed

Kommareddy, M; McAllister, R M; Ganjam, V K; Turk, J R; Laughlin, M Harold

2011-11-01

The objective of this study was to investigate the effects of chronic inhibition of nitric oxide synthase (NOS) on cyclooxygenase-2 (COX-2) expression in the macula densa (MD) of swine, as well as the effects on expression of related proteins. Adult female Yucatan swine were given either tap water (control, n = 6) or water with N (G)-nitro-L-arginine methyl ester (L-NAME, 100 mg/liter, n = 5) for a minimum of 30 days. Duplicate samples of kidney were fixed or snap frozen. There was a significant (P = .0082) upregulation of COX-2 mRNA expression in the MD of L-NAME, as well as an apparent increase in COX-2 protein. Plasma renin activity also increased with L-NAME treatment (control, 0.34 ± 0.08 ng/ml; L-NAME, 1.26 ± 0.03 ng/ml; P = .00000003). There were no differences between groups in expression of either inducible NOS or renin protein or in serum electrolyte concentrations. In conclusion, with chronic inhibition of NOS, COX-2 in MD is upregulated, perhaps to compensate for loss of nitric oxide. Increases in COX-2 products may counteract renal arteriolar constriction and sustain renin release.

Panax ginseng induces the expression of CatSper genes and sperm hyperactivation

PubMed Central

Park, Eun Hwa; Kim, Do Rim; Kim, Ha Young; Park, Seong Kyu; Chang, Mun Seog

2014-01-01

The cation channel of sperm (CatSper) protein family plays important roles in male reproduction and infertility. The four members of this family are expressed exclusively in the testis and are localized differently in sperm. To investigate the effects of Panax ginseng treatment on the expression of CatSper genes and sperm hyperactivation in male mice, sperm motility and CatSper gene expression were assessed using a computer-assisted semen analysis system, a Fluoroskan Ascent microplate fluorometer to assess Ca2+ influx, real-time polymerase chain reaction, Western blotting and immunofluorescence. The results suggested that the Ca2+ levels of sperm cells treated with P. ginseng were increased significantly compared with the normal group. The P. ginseng-treated groups showed increased sperm motility parameters, such as the curvilinear velocity and amplitude of lateral head displacement. Taken together, the data suggest that CatSper messenger ribonucleic acid levels were increased significantly in mouse testes in the P. ginseng-treated group, as was the protein level, with the exception of CatSper2. In conclusion, P. ginseng plays an important role in improving sperm hyperactivation via CatSper gene expression. PMID:24969054

Perivascular Delivery of Notch 1 siRNA Inhibits Injury-Induced Arterial Remodeling

PubMed Central

Redmond, Eileen M.; Liu, Weimin; Hamm, Katie; Hatch, Ekaterina; Cahill, Paul A.; Morrow, David

2014-01-01

Objectives To determine the efficacy of perivascular delivery of Notch 1 siRNA in preventing injury-induced arterial remodeling. Methods and Results Carotid artery ligation was performed to induce arterial remodeling. After 14 days, morphometric analysis confirmed increased vSMC growth and subsequent media thickening and neointimal formation. Laser capture microdissection, quantitative qRT-PCR and immunoblot analysis of medial tissue revealed a significant increase in Notch1 receptor and notch target gene, Hrt 1 and 2 expression in the injured vessels. Perivascular delivery of Notch 1 siRNA by pluronic gel inhibited the injury-induced increase in Notch 1 receptor and target gene expression when compared to scrambled siRNA controls while concomitantly reducing media thickening and neointimal formation to pre-injury, sham-operated levels. Selective Notch 1 knockdown also reversed the injury-induced inhibition of pro-apoptotic Bax expression while decreasing injury-induced anti-apoptotic Bcl-XL expression to sham-operated control levels. In parallel experiments, proliferative cyclin levels, as measured by PCNA expression, were reversed to sham-operated control levels following selective Notch 1 knockdown. Conclusion These results suggest that injury-induced arterial remodeling can be successfully inhibited by localized perivascular delivery of Notch 1 siRNA. PMID:24416200

ROCK inhibitor Y-27632 enhances the survivability of dissociated buffalo (Bubalus bubalis) embryonic stem cell-like cells.

PubMed

Sharma, Ruchi; George, Aman; Chauhan, Manmohan S; Singla, Suresh; Manik, Radhey S; Palta, Prabhat

2013-01-01

This study investigated the effects of supplementation of culture medium with 10 μM Y-27632, a specific inhibitor of Rho kinase activity, for 6 days on self-renewal of buffalo embryonic stem (ES) cell-like cells at Passage 50-80. Y-27632 increased mean colony area (P<0.05) although it did not improve their survival. It decreased OCT4 expression (P<0.05), increased NANOG expression (P<0.05), but had no effect on SOX2 expression. It also increased expression of anti-apoptotic gene BCL-2 (P<0.05) and decreased that of pro-apoptotic genes BAX and BID (P<0.05). It increased plating efficiency of single-cell suspensions of ES cells (P<0.05). Following vitrification, the presence of Y-27632 in the vitrification solution or thawing medium or both did not improve ES cell colony survival. However, following seeding of clumps of ES cells transfected with pAcGFP1N1 carrying green fluorescent protein (GFP), Y-27632 increased colony formation rate (P<0.01). ES cell colonies that formed in all Y-27632-supplemented groups were confirmed for expression of pluripotency markers alkaline phosphatase, SSEA-4 and TRA-1-60, and for their ability to generate embryoid bodies containing cells that expressed markers of ectoderm, mesoderm and endoderm. In conclusion, Y-27632 improves survival of buffalo ES cells under unfavourable conditions such as enzymatic dissociation to single cells or antibiotic-assisted selection after transfection, without compromising their pluripotency.

Uremic Conditions Drive Human Monocytes to Pro-Atherogenic Differentiation via an Angiotensin-Dependent Mechanism

PubMed Central

Trojanowicz, Bogusz; Ulrich, Christof; Seibert, Eric; Fiedler, Roman; Girndt, Matthias

2014-01-01

Aims Elevated expression levels of monocytic-ACE have been found in haemodialysis patients. They are not only epidemiologically linked with increased mortality and cardiovascular disease, but may also directly participate in the initial steps of atherosclerosis. To further address this question we tested the role of monocytic-ACE in promotion of atherosclerotic events in vitro under conditions mimicking those of chronic renal failure. Methods and Results Treatment of human primary monocytes or THP-1 cells with uremic serum as well as PMA-induced differentiation led to significantly up-regulated expression of ACE, further increased by additional treatment with LPS. Functionally, these monocytes revealed significantly increased adhesion and transmigration through endothelial monolayers. Overexpression of ACE in transfected monocytes or THP-1 cells led to development of more differentiated, macrophage-like phenotype with up-regulated expression of Arg1, MCSF, MCP-1 and CCR2. Expression of pro-inflammatory cytokines TNFa and IL-6 were also noticeably up-regulated. ACE overexpression resulted in significantly increased adhesion and transmigration properties. Transcriptional screening of ACE-overexpressing monocytes revealed noticeably increased expression of Angiotensin II receptors and adhesion- as well as atherosclerosis-related ICAM-1 and VCAM1. Inhibition of monocyte ACE or AngII-receptor signalling led to decreased adhesion potential of ACE-overexpressing cells. Conclusions Taken together, these data demonstrate that uremia induced expression of monocytic-ACE mediates the development of highly pro-atherogenic cells via an AngII-dependent mechanism. PMID:25003524

Nocturnal Light Exposure Alters Hepatic Pai-1 Expression by Stimulating the Adrenal Pathway in C3H Mice

PubMed Central

Aoshima, Yoshiki; Sakakibara, Hiroyuki; Suzuki, Taka-aki; Yamazaki, Shunsuke; Shimoi, Kayoko

2014-01-01

Recent studies have suggested the possibility that nocturnal light exposure affects many biological processes in rodents, especially the circadian rhythm, an endogenous oscillation of approximately 24 h. However, there is still insufficient information about the physiological effects of nocturnal light exposure. In this study, we examined the changes in gene expression and serum levels of plasminogen activator inhibitor-1 (PAI-1), a major component of the fibrinolytic system that shows typical circadian rhythmicity, in C3H/He mice. Zeitgeber time (ZT) was assessed with reference to the onset of light period (ZT0). Exposure to fluorescent light (70 lux) for 1 h in the dark period (ZT14) caused a significant increase in hepatic Pai-1 gene expression at ZT16. Serum PAI-1 levels also tended to increase, albeit not significantly. Expression levels of the typical clock genes Bmal1, Clock, and Per1 were significantly increased at ZT21, ZT16, and ZT18, respectively. Exposure to nocturnal light significantly increased plasma adrenalin levels. The effects of nocturnal light exposure on Pai-1 expression disappeared in adrenalectomized mice, although the changes in clock genes were still apparent. In conclusion, our results suggest that nocturnal light exposure, even for 1 h, alters hepatic Pai-1 gene expression by stimulating the adrenal pathway. Adrenalin secreted from the adrenal gland may be an important signaling mediator of the change in Pai-1 expression in response to nocturnal light exposure. PMID:25077763

Central adiponectin administration reveals new regulatory mechanisms of bone metabolism in mice

PubMed Central

Wu, Yuwei; Tu, Qisheng; Valverde, Paloma; Zhang, Jin; Murray, Dana; Dong, Lily Q.; Cheng, Jessica; Jiang, Hua; Rios, Maribel; Morgan, Elise; Tang, Zhihui

2014-01-01

Adiponectin (APN), the most abundant adipocyte-secreted adipokine, regulates energy homeostasis and exerts well-characterized insulin-sensitizing properties. The peripheral or central effects of APN regulating bone metabolism are beginning to be explored but are still not clearly understood. In the present study, we found that APN-knockout (APN-KO) mice fed a normal diet exhibited decreased trabecular structure and mineralization and increased bone marrow adiposity compared with wild-type (WT) mice. APN intracerebroventricular infusions decreased uncoupling protein 1 (UCP1) expression in brown adipose tissue, epinephrine and norepinephrine serum levels, and osteoclast numbers, whereas osteoblast osteogenic marker expression and trabecular bone mass increased in APN-KO and WT mice. In addition, centrally administered APN increased hypothalamic tryptophan hydroxylase 2 (TPH2), cocaine- and amphetamine-regulated transcript (CART), and 5-hydroxytryptamine (serotonin) receptor 2C (Htr2C) expressions but decreased hypothalamic cannabinoid receptor-1 expression. Treatment of immortalized mouse neurons with APN demonstrated that APN-mediated effects on TPH2, CART, and Htr2C expression levels were abolished by downregulating adaptor protein containing pleckstrin homology domain, phosphotyrosine domain, and leucine zipper motif (APPL)-1 expression. Pharmacological increase in sympathetic activity stimulated adipogenic differentiation of bone marrow stromal cells (BMSC) and reversed APN-induced expression of the lysine-specific demethylases involved in regulating their commitment to the osteoblastic lineage. In conclusion, we found that APN regulates bone metabolism via central and peripheral mechanisms to decrease sympathetic tone, inhibit osteoclastic differentiation, and promote osteoblastic commitment of BMSC. PMID:24780611

Expression of the hMLH1 and hMSH2 proteins in normal tissues: relationship to cancer predisposition in hereditary non-polyposis colon cancer.

PubMed

Plevová, Pavlína; Sedláková, Eva; Zapletalová, Jana; Krepelová, Anna; Skýpalová, Petra; Kolár, Zdenek

2005-02-01

The majority of tumours in patients with hereditary non-polyposis colon cancer (HNPCC) occur in large intestine and endometrium; also, other tissues are at increased risk. We studied expression of hMLH1 and hMSH2 proteins in 148 normal samples of various tissues from non-HNPCC patients and in 14 normal colon tissues from HNPCC patients. Immunohistochemical technique was used. Intensity of nuclear staining, percentage of stained cells and H-scores were calculated. Tissues were divided into groups. Groups A, B and C included tissues with increased risk of cancer in HNPCC A) stomach, small and large bowel; (B) endometrium; (C) ovary, ureter, urinary bladder, kidney and liver. Group D tissues were without increased risk. Expression of the proteins was significantly higher in groups A, B and C compared with group D (P<0.0001, P=0.0004 for hMSH2 in C versus D). The expression was highest in testis. In colons of HNPCC patients, expression of the mutated gene product was significantly lower than in non-HNPCC patients. In conclusion, hMLH1/hMSH2 protein expression is constitutively higher in certain cell types of certain tissues, including the majority of tissues that are at increased risk of cancer in HNPCC. However, association of strong hMLH1/hMSH2 expression with cancer risk is not strictly valid.

Na+/H+ exchanger isoform 1-induced osteopontin expression facilitates cardiac hypertrophy through p90 ribosomal S6 kinase.

PubMed

Abdulrahman, Nabeel; Jaspard-Vinassa, Beatrice; Fliegel, Larry; Jabeen, Aayesha; Riaz, Sadaf; Gadeau, Alain-Pierre; Mraiche, Fatima

2018-05-01

Cardiovascular diseases are the leading cause of death worldwide. One in three cases of heart failure is due to dilated cardiomyopathy. The Na + /H + exchanger isoform 1 (NHE1), a multifunctional protein and the key pH regulator in the heart, has been demonstrated to be increased in this condition. We have previously demonstrated that elevated NHE1 activity induced cardiac hypertrophy in vivo. Furthermore, the overexpression of active NHE1 elicited modulation of gene expression in cardiomyocytes including an upregulation of myocardial osteopontin (OPN) expression. To determine the role of OPN in inducing NHE1-mediated cardiomyocyte hypertrophy, double transgenic mice expressing active NHE1 and OPN knockout were generated and assessed by echocardiography and the cardiac phenotype. Our studies showed that hearts expressing active NHE1 exhibited cardiac remodeling indicated by increased systolic and diastolic left ventricular internal diameter and increased ventricular volume. Moreover, these hearts demonstrated impaired function with decreased fractional shortening and ejection fraction. Atrial natriuretic peptide (ANP) and brain natriuretic peptide (BNP) mRNA was upregulated, and there was an increase in heart cell cross-sectional area confirming the cardiac hypertrophic effect. Moreover, NHE1 transgenic mice also showed increased collagen deposition, upregulation of CD44 and phosphorylation of p90 ribosomal s6 kinase (RSK), effects that were regressed in OPN knockout mice. In conclusion, we developed an interesting comparative model of active NHE1 transgenic mouse lines which express a dilated hypertrophic phenotype expressing CD44 and phosphorylated RSK, effects which were regressed in absence of OPN.

Assessing the Effect of High Performance Inulin Supplementation via KLF5 mRNA Expression in Adults with Type 2 Diabetes: A Randomized Placebo Controlled Clinical Trail.

PubMed

Ghavami, Abed; Roshanravan, Neda; Alipour, Shahriar; Barati, Meisam; Mansoori, Behzad; Ghalichi, Faezeh; Nattagh-Eshtivan, Elyas; Ostadrahimi, Alireza

2018-03-01

Purpose: The worldwide prevalence of metabolic disorders such as diabetes is increasing rapidly. Currently, the complications of diabetes are the major health concern. The aim of this study was to investigate the effect of high performance (HP) inulin supplementation on glucose homeostasis via KLF5 mRNA expression in adults with type 2 diabetes. Methods: In the present clinical trial conducted for a duration of 6 weeks, 46 volunteers diabetic patients referring to diabetes clinic in Tabriz, Iran, were randomly assigned into intervention (n= 23, consuming 10 gr/d HP inulin) and control groups (n= 23, consuming 10 gr/ d starch). We assessed glycemic and anthropometric indices, blood lipids and plasmatic level of miR-375 as well as KLF5 mRNA expression before and after the intervention. Results: Findings indicated that inulin supplementation significantly decreased fasting plasma glucose (FPG) in comparison to the placebo group (P<0.001). Also Intra-group and between group results showed that inulin supplementation resulted in significant decrease in KLF5 mRNA expression in peripheral blood mononuclear cells (PBMCs) (Fold change: 0.61± 0.11; P-value= 0.001) and significant increase in plasmatic level of miR-375 (Fold change: 3.75± 0.70; P-value=0.004). Conclusion: Considering the improvements of FPG level in diabetic patients, it seems that HP inulin supplementation may be beneficial in controlling diabetes via the expression of some genes. However, further studies are needed to achieve concise conclusions.

Assessing the Effect of High Performance Inulin Supplementation via KLF5 mRNA Expression in Adults with Type 2 Diabetes: A Randomized Placebo Controlled Clinical Trail

PubMed Central

Ghavami, Abed; Roshanravan, Neda; Alipour, Shahriar; Barati, Meisam; Mansoori, Behzad; Ghalichi, Faezeh; Nattagh- Eshtivan, Elyas; Ostadrahimi, Alireza

2018-01-01

Purpose: The worldwide prevalence of metabolic disorders such as diabetes is increasing rapidly. Currently, the complications of diabetes are the major health concern. The aim of this study was to investigate the effect of high performance (HP) inulin supplementation on glucose homeostasis via KLF5 mRNA expression in adults with type 2 diabetes. Methods: In the present clinical trial conducted for a duration of 6 weeks, 46 volunteers diabetic patients referring to diabetes clinic in Tabriz, Iran, were randomly assigned into intervention (n= 23, consuming 10 gr/d HP inulin) and control groups (n= 23, consuming 10 gr/ d starch). We assessed glycemic and anthropometric indices, blood lipids and plasmatic level of miR-375 as well as KLF5 mRNA expression before and after the intervention. Results: Findings indicated that inulin supplementation significantly decreased fasting plasma glucose (FPG) in comparison to the placebo group (P<0.001). Also Intra-group and between group results showed that inulin supplementation resulted in significant decrease in KLF5 mRNA expression in peripheral blood mononuclear cells (PBMCs) (Fold change: 0.61± 0.11; P-value= 0.001) and significant increase in plasmatic level of miR-375 (Fold change: 3.75± 0.70; P-value=0.004). Conclusion: Considering the improvements of FPG level in diabetic patients, it seems that HP inulin supplementation may be beneficial in controlling diabetes via the expression of some genes. However, further studies are needed to achieve concise conclusions. PMID:29670837

Quercetin suppresses immune cell accumulation and improves mitochondrial gene expression in adipose tissue of diet‐induced obese mice

PubMed Central

Takahashi, Yumiko; Sakurai, Mutsumi; Akimoto, Yukari; Tsushida, Tojiro; Oike, Hideaki; Ippoushi, Katsunari

2015-01-01

Scope To examine the effect of dietary quercetin on the function of epididymal adipose tissue (EAT) in Western diet‐induced obese mice. Methods and results C57BL/6J mice were fed a control diet; a Western diet high in fat, cholesterol, and sucrose; or the same Western diet containing 0.05% quercetin for 18 weeks. Supplementation with quercetin suppressed the increase in the number of macrophages, the decrease in the ratio of CD4+ to CD8+ T cells in EAT, and the elevation of plasma leptin and tumor necrosis factor α levels in mice fed the Western diet. Comprehensive gene expression analysis revealed that quercetin suppressed gene expression associated with the accumulation and activation of immune cells, including macrophages and lymphocytes in EAT. It also improved the expression of the oxidative stress‐sensitive transcription factor NFκB, NADPH oxidases, and antioxidant enzymes. Quercetin markedly increased gene expression associated with mitochondrial oxidative phosphorylation and mitochondrial DNA content. Conclusion Quercetin most likely universally suppresses the accumulation and activation of immune cells, including antiinflammatory cells, whereas it specifically increased gene expression associated with mitochondrial oxidative phosphorylation. Suppression of oxidative stress and NFκB activity likely contributed to the prevention of the accumulation and activation of immune cells and resulting chronic inflammation. PMID:26499876

The barley MATE gene, HvAACT1, increases citrate efflux and Al3+ tolerance when expressed in wheat and barley

PubMed Central

Zhou, Gaofeng; Delhaize, Emmanuel; Zhou, Meixue; Ryan, Peter R.

2013-01-01

Background and Aims Aluminium is toxic in acid soils because the soluble Al3+ inhibits root growth. A mechanism of Al3+ tolerance discovered in many plant species involves the release of organic anions from root apices. The Al3+-activated release of citrate from the root apices of Al3+-tolerant genotypes of barley is controlled by a MATE gene named HvAACT1 that encodes a citrate transport protein located on the plasma membrane. The aim of this study was to investigate whether expressing HvAACT1 with a constitutive promoter in barley and wheat can increase citrate efflux and Al3+ tolerance of these important cereal species. Methods HvAACT1 was over-expressed in wheat (Triticum aestivum) and barley (Hordeum vulgare) using the maize ubiquitin promoter. Root apices of transgenic and control lines were analysed for HvAACT1 expression and organic acid efflux. The Al3+ tolerance of transgenic and control lines was assessed in both hydroponic solution and acid soil. Key Results and Conclusions Increased HvAACT1 expression in both cereal species was associated with increased citrate efflux from root apices and enhanced Al3+ tolerance, thus demonstrating that biotechnology can complement traditional breeding practices to increase the Al3+ tolerance of important crop plants. PMID:23798600

Abdominal Manual Therapy Repairs Interstitial Cells of Cajal and Increases Colonic c-Kit Expression When Treating Bowel Dysfunction after Spinal Cord Injury

PubMed Central

Zhang, Wenyi; Zhu, Zhaojin; Xie, Bin; Yu, Jun

2017-01-01

Background This study aimed to evaluate the therapeutic effects of abdominal manual therapy (AMT) on bowel dysfunction after spinal cord injury (SCI), investigating interstitial cells of Cajal (ICCs) and related c-kit expression. Methods Model rats were divided as SCI and SCI with drug treatment (intragastric mosapride), low-intensity (SCI + LMT; 50 g, 50 times/min), and high-intensity AMT (SCI + HMT; 100 g, 150 times/min). After 14 days of treatment, weight, improved Basso-Beattie-Bresnahan (BBB) locomotor score, and intestinal movement were evaluated. Morphological structure of spinal cord and colon tissues were examined. Immunostaining, RT-PCR, and western blot were used to assess c-kit expression. Results In SCI rats, AMT could not restore BBB, but it significantly increased weight, shortened time to defecation, increased feces amounts, and improved fecal pellet traits and colon histology. AMT improved the number, distribution, and ultrastructure of colonic ICCs, increasing colonic c-kit mRNA and protein levels. Compared with the SCI + Drug and SCI + LMT groups, the SCI + HMT group showed better therapeutic effect in improving intestinal transmission function and promoting c-kit expression. Conclusions AMT is an effective therapy for recovery of intestinal transmission function. It could repair ICCs and increase c-kit expression in colon tissues after SCI, in a frequency-dependent and pressure-dependent manner. PMID:29349063

Surface expression of squamous cell carcinoma antigen (SCCA) can be increased by the preS1(21-47) sequence of hepatitis B virus.

PubMed

Ruvoletto, M G; Tono, N; Carollo, D; Vilei, T; Trentin, L; Muraca, M; Marino, M; Gatta, A; Fassina, G; Pontisso, P

2004-03-01

A variant of the serpin squamous cell carcinoma antigen (SCCA) has been identified as a hepatitis B virus binding protein and high expression of SCCA has recently been found in hepatocarcinoma. Since HBV is involved in liver carcinogenesis, experiments were carried out to examine the effect of HBV preS1 envelope protein on SCCA expression. Surface and intracellular staining for SCCA was assessed by FACS analysis. Preincubation of HepG2 cells and primary human hepatocytes with preS1 protein or with preS1(21-47) tetrameric peptide significantly increased the surface expression of SCCA, without modification of its overall cellular burden, suggesting a surface redistribution of the serpin. An increase in HBV binding and internalization was observed after pre-incubation of the cells with preS1 preparations, compared to cells preincubated with medium alone. Pretreatment of cells with DMSO, while not influencing SCCA basal expression, was responsible for an increase in the efficiency of HBV internalization and this effect was additive to that obtained after incubation with preS1 preparations. In conclusion, the HBV preS1(21-47) sequence is able to induce overexpression of SCCA at the cell surface facilitating virus internalization, while the increased efficiency of HBV entry following DMSO addition is not mediated by SCCA.

XIAP and Ki-67: A Correlation Between Antiapoptotic and Proliferative Marker Expression in Benign and Malignant Tumours of Salivary Gland: An Immunohistochemical Study

PubMed Central

Gawande, Madhuri; Chaudhary, Minal; Gadbail, Amol Ramchandra; Patil, Swati; Bagulkar, Smita

2015-01-01

Background: Impaired balance between cell proliferation and apoptosis is crucial to the development of malignant neoplasm. The purpose of this study was to evaluate and compare the expression of X-Linked inhibitor of apoptotic protein (XIAP) (antiapoptotic marker) and Ki-67 (proliferative marker) expression in benign and malignant salivary gland (SG) tumours. Materials and Methods: The study consisted of 40 cases of benign SG tumours and 50 cases of malignant SG tumours. The immunohistochemistry was carried out by using Ki-67 antibody (clone MIB-1) and XIAP antibody in all the groups. Results: XIAP expression was significantly higher in malignant SG tumours than benign SG tumours (p = 0.016). Ki-67 LI was significantly higher in malignant SG tumours than benign SG tumours (p = 0.0002). Statistically significant positive correlation between Ki-67 count and XIAP expression was noted in benign and malignant SG tumours (p = 0.000). Conclusion: As the expression of an antiapoptotic marker (XIAP) increases, the expression of a proliferative marker (Ki-67) also increases from benign to malignant SG tumours. Thus, targeted therapy of XIAP may play a future role in the management of SG malignancy. PMID:25859460

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

Zhu Lingling; Zaidi, Samir; Peng Yuanzhen

Strontium ranelate, a new agent for the treatment of osteoporosis, has been shown stimulate bone formation in various experimental models. This study examines the effect of strontium ranelate on gene expression in osteoblasts, as well as the formation of mineralized (von Kossa-positive) colony-forming unit-osteoblasts (CFU-obs). Bone marrow-derived stromal cells cultured for 21 days under differentiating conditions, when exposed to strontium ranelate, displayed a significant time- and concentration-dependent increase in the expression of the master gene, Runx2, as well as bone sialoprotein (BSP), but interestingly without effects on osteocalcin. This was associated with a significant increase in the formation of CFU-obsmore » at day 21 of culture. In U-33 pre-osteoblastic cells, strontium ranelate significantly enhanced the expression of Runx2 and osteocalcin, but not BSP. Late, more mature osteoblastic OB-6 cells showed significant elevations in BSP and osteocalcin, but with only minimal effects on Runx2. In conclusion, strontium ranelate stimulates osteoblast differentiation, but the induction of the program of gene expression appears to be cell type-specific. The increased osteoblastic differentiation is the likely basis underlying the therapeutic bone-forming actions of strontium ranelate.« less

Thermo-chemotherapy Induced miR-218 upregulation inhibits the invasion of gastric cancer via targeting Gli2 and E-cadherin.

PubMed

Ruan, Qiang; Fang, Zhi-Yuan; Cui, Shu-Zhong; Zhang, Xiang-Liang; Wu, Yin-Bing; Tang, Hong-Sheng; Tu, Yi-Nuo; Ding, Yan

2015-08-01

Thermo-chemotherapy has been proven to reduce the invasion capability of cancer cells. However, the molecular mechanism underlying this anti-invasion effect is still unclear. In this study, the role of thermo-chemotherapy in the inhibition of tumor invasion was studied. The results demonstrated that expression of miR-218 was downregulated in gastric cancer tissues, which had a positive correlation with tumor invasion and metastasis. In vitro thermo-chemotherapy increased miR-218 expression in SGC7901 cells and inhibited both proliferation and invasion of cancer cells. Gli2 was identified as a downstream target of miR-218, and its expression was negatively regulated by miR-218. The thermo-chemotherapy induced miR-218 upregulation was also accompanied by increasing of E-cadherin expression. In conclusion, the present study indicates that thermo-chemotherapy can effectively decrease the invasion capability of cancer cells and increase cell-cell adhesion. miR-218 and its downstream target Gli2, as well as E-cadherin, participate in the anti-invasion process.

Differential action of glucocorticoids on apolipoprotein E gene expression in macrophages and hepatocytes

PubMed Central

Trusca, Violeta Georgeta; Fuior, Elena Valeria; Fenyo, Ioana Madalina; Kardassis, Dimitris; Simionescu, Maya

2017-01-01

Apolipoprotein E (apoE) has anti-atherosclerotic properties, being involved in the transport and clearance of cholesterol-rich lipoproteins as well as in cholesterol efflux from cells. We hypothesized that glucocorticoids may exert anti-inflammatory properties by increasing the level of macrophage-derived apoE. Our data showed that glucocorticoids increased apoE expression in macrophages in vitro as well as in vivo. Dexamethasone increased ~6 fold apoE mRNA levels in cultured peritoneal macrophages and RAW 264.7 cells. Administered to C57BL/6J mice, dexamethasone induced a two-fold increase in apoE expression in peritoneal macrophages. By contrast, glucocorticoids did not influence apoE expression in hepatocytes, in vitro and in vivo. Moreover, dexamethasone enhanced apoE promoter transcriptional activity in RAW 264.7 macrophages, but not in HepG2 cells, as tested by transient transfections. Analysis of apoE proximal promoter deletion mutants, complemented by protein-DNA interaction assays demonstrated the functionality of a putative glucocorticoid receptors (GR) binding site predicted by in silico analysis in the -111/-104 region of the human apoE promoter. In hepatocytes, GR can bind to their specific site within apoE promoter but are not able to modulate the gene expression. The modulatory blockade in hepatocytes is a consequence of partial involvement of transcription factors and other signaling molecules activated through MEK1/2 and PLA2/PLC pathways. In conclusion, our study indicates that glucocorticoids (1) differentially target apoE gene expression; (2) induce a significant increase in apoE level specifically in macrophages. The local increase of apoE gene expression in macrophages at the level of the atheromatous plaque may have therapeutic implications in atherosclerosis. PMID:28355284

Increased expression of pro-angiogenic factors and vascularization in thyroid hyperfunctioning adenomas with and without TSH receptor activating mutations.

PubMed

Celano, Marilena; Sponziello, Marialuisa; Tallini, Giovanni; Maggisano, Valentina; Bruno, Rocco; Dima, Mariavittoria; Di Oto, Enrico; Redler, Adriano; Durante, Cosimo; Sacco, Rosario; Filetti, Sebastiano; Russo, Diego

2013-02-01

Autonomously functioning thyroid nodules (AFTN) are known to receive an increased blood influx necessary to sustain their high rate of growth and hormone production. Here, we investigated the expression of hematic and lymphatic vases in a series of 20 AFTN compared with the contralateral non-tumor tissues of the same patients, and the transcript levels of proteins involved in the control of vascular proliferation, including the vascular endothelial growth factor (VEGF) and platelet-derived growth factors (PDGF) and their receptors and the endothelial nitric oxide synthase (eNOS). In parallel, the expression of the differentiation markers sodium/iodide symporter (NIS), thyroperoxidase (TPO), thyroglobulin (Tg), and TSH receptor (TSHR) was also investigated. The data were further analyzed comparing subgroups of tumors with or without mutations in the TSHR gene. Analysis by means of CD31 and D2-40 immunostaining showed in AFTN an increased number of hematic, but not lymphatic, vessels in parallel with an enhanced proliferation rate shown by increased Ki67 staining. Quantitative RT-PCR analysis revealed an increase of VEGF, VEGFR1 and 2, PDGF-A, PDGF-B, and eNOS expression in tumor versus normal tissues. Also, higher transcript levels of NIS, TPO, and Tg were detected. Comparison of the two subgroups of samples revealed only few differences in the expression of the genes examined. In conclusion, these data demonstrate an increased expression of angiogenesis-related factors associated with an enhanced proliferation of hematic, but not lymphatic, vessels in AFTNs. In this context, the presence of TSHR mutations may only slightly influence the expression of pro-angiogenic growth factors.

Tributyltin increases the expression of apoptosis- and adipogenesis-related genes in rat ovaries

PubMed Central

Lee, Hyojin; Lim, Sojeong; Yun, Sujin; Yoon, Ayoung; Park, Gayoung

2012-01-01

Objective Tributyltin (TBT), an endocrine disrupting chemical, has been reported to decrease ovarian function by causing apoptosis in the ovary, but the mechanism is not fully understood. Therefore, we examined whether TBT increases the expression of adipogenesis-related genes in the ovary and the increased expression of these genes is associated with apoptosis induction. Methods Three-week-old Sprague-Dawley rats were orally administered TBT (1 or 10 mg/kg body weight) or sesame oil as a control for 7 days. The ovaries were obtained and weighed on day 8, and then they were fixed for terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) or frozen for RNA extraction. Using the total RNA of the ovaries, adipogenesis- and apoptosis-related genes were analyzed by real-time polymerase chain reaction (PCR). Results The ovarian weight was significantly decreased in rats administered 10 mg/kg TBT compared to that in control rats. As determined by the TUNEL assay, the number of apoptotic follicles in ovary was significantly increased in rats administered 10 mg/kg TBT. The real-time PCR results showed that the expression of adipogenesis-related genes such as PPARγ, aP2, CD36, and PEPCK was increased after TBT administration. In addition, apoptosis-related genes such as TNFα and TNFR1 were expressed more in the TBT-administered rats compared with the control rats. Conclusion The present study demonstrates that TBT induces the expression of adipogenesis- and apoptosis-related genes in the ovary leading to apoptosis in the ovarian follicles. These results suggest that the increased expression of adipogenesis-related genes in the ovary by TBT exposure might induce apoptosis resulting in a loss of ovarian function. PMID:22563546

Effect of polysaccharide of dendrobium candidum on proliferation and apoptosis of human corneal epithelial cells in high glucose

PubMed Central

Li, Qiangxiang; Chen, Jing; Li, Yajia; Chen, Ting; Zou, Jing; Wang, Hua

2017-01-01

Abstract Background: The aim of the study was to observe the effect of polysaccharide of dendrobium candidum (PDC) and high glucose on proliferation, apoptosis of human corneal epithelial cells (HCEC). Methods: The MTT method was used to screen and take the optimal high-glucose concentration, treatment time, and PDC concentration using HCEC and divide it into 4 groups: control group (C), high glucose group (HG), PDC group, and HG + PDC group. We observed and compared the effect of the 4 groups on HCEC proliferation by MTT, apoptosis by Annexin V-FITC/PI double fluorescent staining and flow cytometry (FCM), and expression of bax mRNA and bcl-2 mRNA by RT-qPCR. Results: Compared with the control group, proliferative activity of HCEC cells was reduced; the cells apoptosis ratio was increased; the expression of bax mRNA was increased, and the expression of bcl-2 mRNA was reduced in the HG group. Proliferative activity of HCEC cells in the PDC group was increased, and the expression of bcl-2 mRNA was increased but that of bax mRNA was decreased. Proliferative activity of HCEC cells in the HG + PDC group was increased, but it could not restore to the normal level; the expression of bax mRNA was significantly decreased but the expression of bcl-2 mRNA was significantly increased. Conclusions: Our results demonstrate that high glucose can inhibit proliferative activity and induce apoptosis of HCEC. PDC can improve the proliferative activity of HCEC cells under the high glucose environment and reduce the apoptosis of cells by regulating the expression of bax and bcl-2. PDC play a very important role on protecting and repairing of corneal epithelial cells damage in high glucose. PMID:28796073

Arsenic exposure from drinking water is associated with decreased gene expression and increased DNA methylation in peripheral blood

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

Ameer, Syeda Shegufta

Background: Exposure to inorganic arsenic increases the risk of cancer and non-malignant diseases. Inefficient arsenic metabolism is a marker for susceptibility to arsenic toxicity. Arsenic may alter gene expression, possibly by altering DNA methylation. Objectives: To elucidate the associations between arsenic exposure, gene expression, and DNA methylation in peripheral blood, and the modifying effects of arsenic metabolism. Methods: The study participants, women from the Andes, Argentina, were exposed to arsenic via drinking water. Arsenic exposure was assessed as the sum of arsenic metabolites in urine (U-As), using high performance liquid-chromatography hydride-generation inductively-coupled-plasma-mass-spectrometry, and arsenic metabolism efficiency was assessed by themore » urinary fractions (%) of the individual metabolites. Genome-wide gene expression (N = 80 women) and DNA methylation (N = 93; 80 overlapping with gene expression) in peripheral blood were measured using Illumina DirectHyb HumanHT-12 v4.0 and Infinium Human-Methylation 450K BeadChip, respectively. Results: U-As concentrations, ranging 10–1251 μg/L, was associated with decreased gene expression: 64% of the top 1000 differentially expressed genes were down-regulated with increasing U-As. U-As was also associated with hypermethylation: 87% of the top 1000 CpGs were hypermethylated with increasing U-As. The expression of six genes and six individual CpG sites were significantly associated with increased U-As concentration. Pathway analyses revealed enrichment of genes related to cell death and cancer. The pathways differed somewhat depending on arsenic metabolism efficiency. We found no overlap between arsenic-related gene expression and DNA methylation for individual genes. Conclusions: Increased arsenic exposure was associated with lower gene expression and hypermethylation in peripheral blood, but with no evident overlap. - Highlights: • Women exposed to inorganic arsenic were studied for molecular responses in blood. • Arsenic is associated with decreased gene expression and increased DNA methylation. • Arsenic related pathways differed to some extent due to arsenic metabolism efficiency.« less

Annexin A1: A new immunohistological marker of cholangiocarcinoma

PubMed Central

Hongsrichan, Nuttanan; Rucksaken, Rucksak; Chamgramol, Yaovalux; Pinlaor, Porntip; Techasen, Anchalee; Yongvanit, Puangrat; Khuntikeo, Narong; Pairojkul, Chawalit; Pinlaor, Somchai

2013-01-01

AIM: To evaluate a new immunohistological marker, annexin A1 (ANXA1), in cholangiocarcinoma (CCA) and hepatocellular carcinoma (HCC). METHODS: Expression of ANXA1 protein was investigated in liver tissues from patients with CCA and HCC by immunohistochemistry. Its expression on differences stages of tumor development was investigated in hamster CCA tissues induced by Opisthorchis viverrini and N-nitrosodimethylamine. Moreover, mRNA expression of ANXA1 was assessed in CCA cell lines by quantitative real-time polymerase chain reaction and silencing of ANXA1 gene expression using small interfering RNA. RESULTS: In human CCA tissue arrays, immunohistochemical analysis revealed that the positive expression of ANXA1 was 94.1% (64/68 cases) consisting of a high expression (66.2%, 45/68 cases) and a low expression (33.8%, 23/68 cases). However, expression of ANXA1 protein was negative in all histologic patterns for HCC (46/46 cases) and healthy individuals (6/6 cases). In hamster with opisthorchiasis-associated CCA, the expression of ANXA1 was observed in the cytoplasm of inflammatory cells, bile duct epithelia and tumor cells. Grading scores of ANXA1 expression were significantly increased with tumor progression. In addition, mRNA expression of ANXA1 significantly increased in all of the various CCA cell lines tested compared to an immortalized human cholangiocyte cell line (MMNK1). Suppressing the ANXA1 gene significantly reduced the matrix metalloproteinase (MMP) 2 and MMP9, and transforming growth factor-β genes, but increased nuclear factor-κB gene expression. CONCLUSION: ANXA1 is highly expressed in CCA, but low in HCC, suggesting it may serve as a new immunohistochemical marker of CCA. ANXA1 may play a role in opisthorchiasis-associated cholangiocarcinogenesis. PMID:23674846

Decreased expression of fibulin-4 in aortic wall of aortic dissection.

PubMed

Huawei, P; Qian, C; Chuan, T; Lei, L; Laing, W; Wenlong, X; Wenzhi, L

2014-02-01

In this research, we will examine the expression of Fibulin-4 in aortic wall to find out its role in aortic dissection development. The samples of aortic wall were obtained from 10 patients operated for acute ascending aortic dissection and five patients for chronic ascending aortic dissection. Another 15 pieces of samples from patients who had coronary artery bypass were as controls. The aortic samples were stained with aldehyde magenta dyeing to evaluate the arrangement of elastic fibers. The Fibulin-4 protein and mRNA expression were both determined by Western blot and realtime quantitative polymerase chain reaction. Compared with the control group, both in acute and chronic ascending aortic dissection, elastic fiber fragments increased and the expression of fibulin-4 protein significantly decreased (P= 0.045 < 0.05). The level of fibulin-4 mRNA decreased in acute ascending aortic dissection (P= 0.034 < 0.05), while it increased in chronic ascending aortic dissection (P=0.004 < 0.05). The increased amounts of elastic fiber fragments were negatively correlated with the expression of fibulin-4 mRNA in acute ascending aortic dissection. In conclusion, in aortic wall of ascending aortic dissection, the expression of fibulin-4 protein decreased and the expression of fibulin-4 mRNA was abnormal. Fibulin-4 may play an important role in the pathogenesis of aortic dissection.

Mechanical Loading of Articular Cartilage Reduces IL-1-Induced Enzyme Expression

PubMed Central

Torzilli, P. A.; Bhargava, M.; Chen, C. T.

2011-01-01

Objective: Exposure of articular cartilage to interleukin-1 (IL-1) results in increased synthesis of matrix degrading enzymes. Previously mechanical load applied together with IL-1 stimulation was found to reduce aggrecan cleavage by ADAMTS-4 and 5 and MMP-1, -3, -9, and -13 and reduce proteoglycan loss from the extracellular matrix. To further delineate the inhibition mechanism the gene expression of ADAMTS-4 and 5; MMP-1, -3, -9, and -13; and TIMP-1, -2, and -3 were measured. Design: Mature bovine articular cartilage was stimulated with a 0.5 MPa compressive stress and 10 ng/ml of IL-1α for 3 days and then allowed to recover without stimulation for 1 additional day. The media was assayed for proteoglycan content on a daily basis, while chondrocyte gene expression (mRNA) was measured during stimulation and 1 day of recovery. Results: Mechanical load alone did not change the gene expression for ADAMTS, MMP, or TIMP. IL-1 caused an increase in gene expression for all enzymes after 1 day of stimulation while not affecting the TIMP levels. Load applied together with IL-1 decreased the expression levels of ADAMTS-4 and -5 and MMP-1 and -3 and increased TIMP-3 expression. Conclusions: A mechanical load appears to modify cartilage degradation by IL-1 at the cellular level by reducing mRNA. PMID:22039566

Swim training and the genetic expression of adipokines in monosodium glutamate-treated obese rats.

PubMed

Svidnicki, Paulo Vinicius; Leite, Nayara Carvalho; Vicari, Marcelo Ricardo; Almeida, Mara Cristina de; Artoni, Roberto Ferreira; Favero, Giovani Marino; Grassiolli, Sabrina; Nogaroto, Viviane

2015-06-01

The aim of this study was to evaluate the genetic expression of adipokines in the adipocytes of monosodium glutamate (MSG)-treated obese rats submitted to physical activity. Obesity was induced by neonatal MSG administration. Exercised rats (MSG and control) were subjected to swim training for 30 min for 10 weeks, whereas their respective controls remained sedentary. Total RNA was obtained from sections of the mesenteric adipose tissue of the rats. mRNA levels of adiponectin (Adipoq), tumor necrosis factor alpha (Tnf), peroxisome proliferator-activated receptor alpha (Ppara), and peroxisome proliferator-activated receptor gamma (Pparg) adipokines were quantified by quantitative Real-Time Polymerase Chain Reaction (qRT-PCR). In the exercise-trained control group, the expression of Adipoq increased compared to the sedentary control, which was not observed in the MSG-obese rats. Increased levels of Tnf in MSG-obese rats were not reversed by the swim training. The expression of Ppara was higher in sedentary MSG-obese rats compared to the sedentary control. Swimming increased this adipokine expression in the exercise-trained control rats compared to the sedentary ones. mRNA levels of Pparg were higher in the sedentary MSG-rats compared to the sedentary control; however, the exercise did not influenced its expression in the groups analyzed. In conclusion, regular physical activity was not capable to correct the expression of proinflammatory adipokines in MSG-obese rat adipocytes.

Increased ERp57 Expression in HBV-Related Hepatocellular Carcinoma: Possible Correlation and Prognosis.

PubMed

Liu, Miao; Du, Lingyao; He, Zhiliang; Yan, Libo; Shi, Ying; Shang, Jin; Tang, Hong

2017-01-01

Aim. ERp57 is involved in virus induced endoplasmic reticulum stress (ERS) and plays an important role in tumorigenesis. This study aimed to find whether HBV infection altered ERp57 expression and whether ERp57 regulation was involved in hepatitis B virus-related hepatocellular carcinoma (HBV-HCC) genesis. Materials and Methods. HBV-HCC tissues, chronic hepatitis B (CHB) liver tissues, and normal liver tissues were acquired. ERp57 expressions in these tissues were detected through immunohistochemistry (IHC). And ERp57 expression in liver cell line L02, HBV replicative liver cell line L02-pHBV4.1, and HCC cell lines were detected through western blot for verification. Then medical data on patients providing HCC tissues were collected and analyzed along with ERp57 expression. Results. Higher ERp57 expression was found in HCC and CHB tissues ( p < 0.001). And HCC cell lines and L02-pHBV4.1 presented higher ERp57 expression as well. In patients, ERp57 expression showed significant differences between death and survival groups ( p = 0.037). And cumulative survival in patients with higher ERp57 (score ⩾ 8.75) is significantly lower ( p = 0.009). Conclusion. Our study found increased expression of ERp57 in HBV-HCC. Such altered expression could be related to HBV infection and high ERp57 expression may lead to poor prognosis of HBV-HCC patients.

Paeonol protects rat vascular endothelial cells from ox-LDL-induced injury in vitro via downregulating microRNA-21 expression and TNF-α release

PubMed Central

Liu, Ya-rong; Chen, Jun-jun; Dai, Min

2014-01-01

Aim: Paeonol (2′-hydroxy-4′-methoxyacetophenone) from Cortex moutan root is a potential therapeutic agent for atherosclerosis. This study sought to investigate the mechanisms underlying anti-inflammatory effects of paeonol in rat vascular endothelial cells (VECs) in vitro. Methods: VECs were isolated from rat thoracic aortas. The cells were pretreated with paeonol for 24 h, and then stimulated with ox-LDL for another 24 h. The expression of microRNA-21 (miR-21) and PTEN in VECs was analyzed using qRT-PCR. The expression of PTEN protein was detected by Western blotting. TNF-α release by VECs was measured by ELISA. Results: Ox-LDL treatment inhibited VEC growth in dose- and time-dependent manners (the value of IC50 was about 20 mg/L at 24 h). Furthermore, ox-LDL (20 mg/L) significantly increased miR-21 expression and inhibited the expression of PTEN, one of downstream target genes of miR-21 in VECs. In addition, ox-LDL (20 mg/L) significantly increased the release of TNF-α from VECs. Pretreatment with paeonol increased the survival rate of ox-LDL-treated VECs in dose- and time-dependent manners. Moreover, paeonol (120 μmol/L) prevented ox-LDL-induced increases in miR-21 expression and TNF-α release, and ox-LDL-induced inhibition in PTEN expression. A dual-luciferase reporter assay showed that miR-21 bound directly to PTEN's 3′-UTR, thus inhibiting PTEN expression. In ox-LDL treated VECs, transfection with a miR-21 mimic significantly increased miR-21 expression and inhibited PTEN expression, and attenuated the protective effects of paeonol pretreatment, whereas transfection with an miR-21 inhibitor significantly decreased miR-21 expression and increased PTEN expression, thus enhanced the protective effects of paeonol pretreatment. Conclusion: miR-21 is an important target of paeonol for its protective effects against ox-LDL-induced VEC injury, which may play critical roles in development of atherosclerosis. PMID:24562307

Amphetamine as a social drug: Effects of d-amphetamine on social processing and behavior

PubMed Central

Wardle, Margaret C.; Garner, Matthew J.; Munafò, Marcus R.; de Wit, Harriet

2012-01-01

Rationale Drug users often report using drugs to enhance social situations, and empirical studies support the idea that drugs increase both social behavior and the value of social interactions. One way drugs may affect social behavior is by altering social processing, for example by decreasing perceptions of negative emotion in others. Objectives We examined effects of d-amphetamine on processing of emotional facial expressions, and on the social behavior of talking. We predicted amphetamine would enhance attention, identification and responsivity to positive expressions, and that this in turn would predict increased talkativeness. Methods Over three sessions, 36 healthy normal adults received placebo, 10mg, and 20mg d-amphetamine under counterbalanced double-blind conditions. At each session we measured processing of happy, fearful, sad and angry expressions using an attentional visual probe task, a dynamic emotion identification task, and measures of facial muscle activity. We also measured talking. Results Amphetamine decreased the threshold for identifying all emotions, increased negative facial responses to sad expressions, and increased talkativeness. Contrary to our hypotheses, amphetamine did not alter attention to, identification of or facial responses to positive emotions specifically. Interestingly, the drug decreased the threshold to identify all emotions, and this effect was uniquely related to increased talkativeness, even after controlling for overall sensitivity to amphetamine. Conclusions The results suggest that amphetamine may encourage sociability by increasing sensitivity to subtle emotional expressions. These findings suggest novel social mechanisms that may contribute to the rewarding effects of amphetamine. PMID:22526538

9- and 13-HODE regulate fatty acid binding protein-4 in human macrophages, but does not involve HODE/GPR132 axis in PPAR-γ regulation of FABP4

PubMed Central

Shashidhar, Venkatesh; Collier, Fiona; Hodge, Jason; Rush, Catherine; Malabu, Usman; Baune, Bernhard; Kennedy, Richard Lee

2018-01-01

Background: Both activation of monocytes and increased serum fatty acid binding protein-4 (FABP4) occur in diabetes and are associated with increased atherosclerosis. The oxidized lipid, 9-hydroxyoctadecadienoic acid (9-HODE) increases FABP4 in macrophages, and is a ligand for G protein-coupled receptor 132 (GPR132). We investigated the involvement of GPR132 in mediating the 9-, 13-HODE stimulation of FABP4 secretion, and whether GPR132 expression is increased in monocytes from patients with type 2 diabetes. Methods: The effects of siRNA silencing of GPR132 gene and of the PPAR-γ antagonist T0070907 were studied in THP-1 cells. Serum levels of FABP4 and other adipokines were measured in patients with diabetes, and monocyte subpopulations were analyzed using flow cytometry. GPR132 mRNA was quantified in isolated CD14+ cells. Results: 9-HODE and 13-HODE increased FABP4 expression in THP-1 monocytes and macrophages, and also increased GPR132 expression. Silencing of GPR132 did not influence the increase in FABP4 with 9-HODE, 13-HODE, or rosiglitazone (ROSI). By contrast, T0070907 inhibited the effect of all three ligands on FABP4 expression. Diabetic subjects had increased serum FABP4, and activated monocytes. They also expressed higher levels of GPR132 mRNA in CD14+ cells. Conclusions: We conclude that GPR132 is an independent monocyte activation marker in diabetes, but does not contribute to PPAR-γ-mediated induction of FABP4 by HODEs.

Bupivacaine-induced apoptosis independently of WDR35 expression in mouse neuroblastoma Neuro2a cells

PubMed Central

2012-01-01

Background Bupivacaine-induced neurotoxicity has been shown to occur through apoptosis. Recently, bupivacaine was shown to elicit reactive oxygen species (ROS) production and induce apoptosis accompanied by activation of p38 mitogen-activated protein kinase (MAPK) in a human neuroblastoma cell line. We have reported that WDR35, a WD40-repeat protein, may mediate apoptosis through caspase-3 activation. The present study was undertaken to test whether bupivacaine induces apoptosis in mouse neuroblastoma Neuro2a cells and to determine whether ROS, p38 MAPK, and WDR35 are involved. Results Our results showed that bupivacaine induced ROS generation and p38 MAPK activation in Neuro2a cells, resulting in apoptosis. Bupivacaine also increased WDR35 expression in a dose- and time-dependent manner. Hydrogen peroxide (H2O2) also increased WDR35 expression in Neuro2a cells. Antioxidant (EUK-8) and p38 MAPK inhibitor (SB202190) treatment attenuated the increase in caspase-3 activity, cell death and WDR35 expression induced by bupivacaine or H2O2. Although transfection of Neuro2a cells with WDR35 siRNA attenuated the bupivacaine- or H2O2-induced increase in expression of WDR35 mRNA and protein, in contrast to our previous studies, it did not inhibit the increase in caspase-3 activity in bupivacaine- or H2O2-treated cells. Conclusions In summary, our results indicated that bupivacaine induced apoptosis in Neuro2a cells. Bupivacaine induced ROS generation and p38 MAPK activation, resulting in an increase in WDR35 expression, in these cells. However, the increase in WDR35 expression may not be essential for the bupivacaine-induced apoptosis in Neuro2a cells. These results may suggest the existence of another mechanism of bupivacaine-induced apoptosis independent from WDR35 expression in Neuro2a cells. PMID:23227925

Cycle Training Increased GLUT4 and Activation of mTOR in Fast Twitch Muscle Fibers

PubMed Central

Stuart, Charles A.; Howell, Mary E.A.; Baker, Jonathan D.; Dykes, Rhesa J.; Duffourc, Michelle M.; Ramsey, Michael W.; Stone, Michael H.

2009-01-01

Purpose To determine if cycle training of sedentary subjects would increase the expression of the principle muscle glucose transporters, six volunteers completed six weeks of progressively increasing intensity stationary cycle cycling. Methods In vastus lateralis muscle biopsies, changes in expression of GLUT1, GLUT4, GLUT5, and GLUT12 were compared using quantitative immunoblots with specific protein standards. Regulatory pathway components were evaluated by immunoblots of muscle homogenates and immunohistochemistry of microscopic sections. Results GLUT1 was unchanged, GLUT4 increased 66%, GLUT12 increased 104%, and GLUT5 decreased 72%. A mitochondrial marker (cytochrome c) and regulators of mitochondrial biogenesis (PGC-1α and phospho-AMPK) were unchanged, but the muscle hypertrophy pathway component, phospho-mTOR increased 83% after the exercise program. In baseline biopsies, GLUT4 by immunohistochemical techniques was 37% greater in Type I (slow twitch, red) muscle fibers, but the exercise training increased GLUT4 expression in Type II (fast twitch, white) fibers by 50%, achieving parity with the Type I fibers. Baseline phospho-mTOR expression was 50% higher in Type II fibers and increased more in Type II fibers (62%) with training, but also increased in Type I fibers (34%). Conclusion Progressive intensity stationary cycle training of previously sedentary subjects increased muscle insulin-responsive glucose transporters (GLUT4 and GLUT12) and decreased the fructose transporter (GLUT5). The increase in GLUT4 occurred primarily in Type II muscle fibers and this coincided with activation of the mTOR muscle hypertrophy pathway. There was little impact on Type I fiber GLUT4 expression and no evidence of change in mitochondrial biogenesis. PMID:20010125

Testosterone Administration Inhibits Hepcidin Transcription and is Associated with Increased Iron Incorporation into Red Blood Cells

PubMed Central

Guo, Wen; Bachman, Eric; Li, Michelle; Roy, Cindy N.; Blusztajn, Jerzy; Wong, Siu; Chan, Stephen Y.; Serra, Carlo; Jasuja, Ravi; Travison, Thomas G.; Muckenthaler, Martina U.; Nemeth, Elizabeta; Bhasin, Shalender

2013-01-01

Testosterone administration increases hemoglobin levels and has been used to treat anemia of chronic disease. Erythrocytosis is the most frequent adverse event associated with testosterone therapy of hypogonadal men, especially older men. However, the mechanisms by which testosterone increases hemoglobin remain unknown. Testosterone administration in male and female mice was associated with a greater increase in hemoglobin and hematocrit, reticulocyte count, reticulocyte hemoglobin concentration, and serum iron and transferring saturation than placebo. Testosterone downregulated hepatic hepcidin mRNA expression, upregulated renal erythropoietin mRNA expression, and increased erythropoietin levels. Testosterone-induced suppression of hepcidin expression was independent of its effects on erythropoietin or hypoxia-sensing mechanisms. Transgenic mice with liver-specific constitutive hepcidin over-expression failed to exhibit the expected increase in hemoglobin in response to testosterone administration. Testosterone upregulated splenic ferroportin expression and reduced iron retention in spleen. After intravenous administration of transferrin-bound 58Fe, the amount of 58Fe incorporated into red blood cells was significantly greater in testosterone-treated mice than in placebo-treated mice. Serum from testosterone-treated mice stimulated hemoglobin synthesis in K562 erythroleukemia cells more than that from vehicle-treated mice. Testosterone administration promoted the association of androgen receptor (AR) with Smad1 and Smad4 to reduce their binding to BMP-response elements in hepcidin promoter in the liver. Ectopic expression of AR in hepatocytes suppressed hepcidin transcription; this effect was blocked dose-dependently by AR antagonist flutamide. Testosterone did not affect hepcidin mRNA stability. Conclusion: Testosterone inhibits hepcidin transcription through its interaction with BMP-Smad signaling. Testosterone administration is associated with increased iron incorporation into red blood cells. PMID:23399021

11β-HSD1 reduces metabolic efficacy and adiponectin synthesis in hypertrophic adipocytes.

PubMed

Koh, Eun Hee; Kim, Ah-Ram; Kim, Hyunshik; Kim, Jin Hee; Park, Hye-Sun; Ko, Myoung Seok; Kim, Mi-Ok; Kim, Hyuk-Joong; Kim, Bum Joong; Yoo, Hyun Ju; Kim, Su Jung; Oh, Jin Sun; Woo, Chang-Yun; Jang, Jung Eun; Leem, Jaechan; Cho, Myung Hwan; Lee, Ki-Up

2015-06-01

Mitochondrial dysfunction in hypertrophic adipocytes can reduce adiponectin synthesis. We investigated whether 11β-hydroxysteroid dehydrogenase type 1 (11β-HSD1) expression is increased in hypertrophic adipocytes and whether this is responsible for mitochondrial dysfunction and reduced adiponectin synthesis. Differentiated 3T3L1 adipocytes were cultured for up to 21 days. The effect of AZD6925, a selective 11β-HSD1 inhibitor, on metabolism was examined. db/db mice were administered 600 mg/kg AZD6925 daily for 4 weeks via gastric lavage. Mitochondrial DNA (mtDNA) content, mRNA expression levels of 11 β -H sd1 and mitochondrial biogenesis factors, adiponectin synthesis, fatty acid oxidation (FAO), oxygen consumption rate and glycolysis were measured. Adipocyte hypertrophy in 3T3L1 cells exposed to a long duration of culture was associated with increased 11 β -Hsd1 mRNA expression and reduced mtDNA content, mitochondrial biogenesis factor expression and adiponectin synthesis. These cells displayed reduced mitochondrial respiration and increased glycolysis. Treatment of these cells with AZD6925 increased adiponectin synthesis and mitochondrial respiration. Inhibition of FAO by etomoxir blocked the AZD6925-induced increase in adiponectin synthesis, indicating that 11β-HSD1-mediated reductions in FAO are responsible for the reduction in adiponectin synthesis. The expression level of 11 β -Hsd1 was higher in adipose tissues of db/db mice. Administration of AZD6925 to db/db mice increased the plasma adiponectin level and adipose tissue FAO. In conclusion, increased 11β-HSD1 expression contributes to reduced mitochondrial respiration and adiponectin synthesis in hypertrophic adipocytes. © 2015 Society for Endocrinology.

Association between macrophage migration inhibitory factor in the endometrium and estrogen in endometriosis

PubMed Central

ZHANG, XIAO; MU, LIN

2015-01-01

Recent studies have shown that macrophage migration inhibitory factor (MIF) has a possible role in endometriosis-related pain and infertility, yet it has not been explored whether the mRNA level of MIF is altered in endometrial tissues from patients with endometriosis. The aim of the present study was to compare the expression of MIF in endometrial tissues from women with and without endometriosis, and to analyze the association between endometrial MIF expression and 17β-estradiol (E2). The protein and mRNA expression of MIF in the human endometrial tissue was assessed by western blotting and reverse transcription-polymerase chain reaction analysis, respectively. The MIF expression of women with endometriosis was found to be significantly higher than that of the controls. A positive correlation was noted between the serum E2 level and MIF expression. In endometrial cells from women with endometriosis, the level of E2-induced MIF upregulation was significantly higher than that in cells from women without endometriosis. In conclusion, this study demonstrated a significant increase in MIF expression in the endometrial tissues of women with endometriosis and an association between MIF expression and E2 level. MIF expression in endometrial cells from patients with endometriosis showed an increased sensitivity to stimulation by E2. PMID:26622394

Human cytomegalovirus UL76 induces chromosome aberrations

PubMed Central

2009-01-01

Background Human cytomegalovirus (HCMV) is known to induce chromosome aberrations in infected cells, which can lead to congenital abnormalities in infected fetuses. HCMV UL76 belongs to a conserved protein family from herpesviruses. Some reported roles among UL76 family members include involvement in virulence determination, lytic replication, reactivation of latent virus, modulation of gene expression, induction of apoptosis, and perturbation of cell cycle progression, as well as potential nuclease activity. Previously, we have shown that stable expression of UL76 inhibits HCMV replication in glioblastoma cells. Methods To examine chromosomal integrity and the DNA damage signal γ-H2AX in cells constitutively expressing UL76, immunofluorescent cell staining and Western blotting were performed. The comet assay was employed to assess DNA breaks in cells transiently expressing UL76. Results We report that stably transfected cells expressing UL76 developed chromosome aberrations including micronuclei and misaligned chromosomes, lagging and bridging. In mitotic cells expressing UL76, aberrant spindles were increased compared to control cells. However, cells with supernumerary centrosomes were marginally increased in UL76-expressing cells relative to control cells. We further demonstrated that UL76-expressing cells activated the DNA damage signal γ-H2AX and caused foci formation in nuclei. In addition, the number of cells with DNA breaks increased in proportion to UL76 protein levels. Conclusion Our findings suggest that the virus-associated protein UL76 induces DNA damage and the accumulation of chromosome aberrations. PMID:19930723

Odontogenic ameloblast-associated protein (ODAM) inhibits growth and migration of human melanoma cells and elicits PTEN elevation and inactivation of PI3K/AKT signaling

PubMed Central

2013-01-01

Background The Odontogenic Ameloblast-associated Protein (ODAM) is expressed in a wide range of normal epithelial, and neoplastic tissues, and we have posited that ODAM serves as a novel prognostic biomarker for breast cancer and melanoma. Transfection of ODAM into breast cancer cells yields suppression of cellular growth, motility, and in vivo tumorigenicity. Herein we have extended these studies to the effects of ODAM on cultured melanoma cell lines. Methods The A375 and C8161 melanoma cell lines were stably transfected with ODAM and assayed for properties associated with tumorigenicity including cell growth, motility, and extracellular matrix adhesion. In addition, ODAM–transfected cells were assayed for signal transduction via AKT which promotes cell proliferation and survival in many neoplasms. Results ODAM expression in A375 and C8161 cells strongly inhibited cell growth and motility in vitro, increased cell adhesion to extracellular matrix, and yielded significant cytoskeletal/morphologic rearrangement. Furthermore, AKT activity was downregulated by ODAM expression while an increase was noted in expression of the PTEN (phosphatase and tensin homolog on chromosome 10) tumor suppressor gene, an antagonist of AKT activation. Increased PTEN in ODAM-expressing cells was associated with increases in PTEN mRNA levels and de novo protein synthesis. Silencing of PTEN expression yielded recovery of AKT activity in ODAM-expressing melanoma cells. Similar PTEN elevation and inhibition of AKT by ODAM was observed in MDA-MB-231 breast cancer cells while ODAM expression had no effect in PTEN-deficient BT-549 breast cancer cells. Conclusions The apparent anti-neoplastic effects of ODAM in cultured melanoma and breast cancer cells are associated with increased PTEN expression, and suppression of AKT activity. This association should serve to clarify the clinical import of ODAM expression and any role it may serve as an indicator of tumor behavior. PMID:23648148

Vitamin D receptor expression and potential role of vitamin D on cell proliferation and steroidogenesis in goat ovarian granulosa cells.

PubMed

Yao, Xiaolei; Zhang, Guomin; Guo, Yixuan; Ei-Samahy, Mohamed; Wang, Shuting; Wan, Yongjie; Han, Le; Liu, Zifei; Wang, Feng; Zhang, Yanli

2017-10-15

This study aimed to investigate the expression of the vitamin D receptor (VDR) in goat follicles and to determine the effects of Vit D 3 supplementation on goat granulosa cells (GCs) function linked to follicular development. The results demonstrated that VDR was prominently localized in GCs, with expression increasing with follicle diameter. Addition of Vit D 3 (1α,25-(OH) 2 VD 3 ; 10 nM) to GCs caused an increase in VDR and in steroidogenic acute regulator (StAR) and 3β-hydroxysteroid dehydrogenase (3β-HSD) mRNA expression. Additionally, Vit D 3 increased the cyclic adenosine monophosphate (cAMP), estradiol (E 2 ), and progesterone (P 4 ) levels, while it decreased anti-müllerian hormone receptor (AMHR) and follicle-stimulating hormone receptor (FSHR) mRNA expression (P < 0.05). Addition of FSH remarkably increased E 2, P 4 , and cAMP levels (P < 0.05), and Vit D 3 further enhanced the E 2 and cAMP levels in the presence of FSH (P < 0.05). Vit D 3 significantly induced the mRNA expression of CDK4 and CyclinD1, and downregulated P21 gene expression (P < 0.05). In addition, Vit D 3 significantly decreased reactive oxygen species (ROS) production and increased the mRNA and protein expression of superoxide dismutase 2 (SOD2) and catalase (CAT) (P < 0.05). In conclusion, VDR is expressed in GCs of the goat ovaries and Vit D 3 might play an important role in GCs proliferation by regulating cellular oxidative stress and cell cycle-related genes. Meanwhile, Vit D 3 enhances the E 2 and P 4 output of GCs by regulating the expression of 3β-HSD and StAR and the level of cAMP, which regulate steroidogenesis, supporting a potential role for Vit D 3 in follicular development. Copyright © 2017 Elsevier Inc. All rights reserved.

Analysis of TP53 gene expression and p53 level of human hypopharyngeal FaDu (HTB-43) head and neck cancer cell line after microRNA-181a inhibition.

PubMed

Cheah, Y K; Cheng, R W; Yeap, S K; Khoo, C H; See, H S

2014-03-17

The identification of new biomarkers for early detection of highly recurrent head and neck cancer is urgently needed. MicroRNAs (miRNAs) are small and non-coding RNAs that regulate cancer-related gene expression, such as tumor protein 53 (TP53) gene expression. This study was carried out to analyze TP53 gene expression using real-time PCR and to determine changes in intracellular p53 level by flow cytometry after downregulation of miRNA-181a miRNA inhibitor in the FaDu cell line. TP53 gene expression showed a 3-fold increment and the p53 protein level was also increased in the miRNA-181a-treated cells. In conclusion, miRNA-181a binds to the TP53 gene and inhibits its expression, decreasing the synthesis of p53.

Albumin Overload and PINK1/Parkin Signaling-Related Mitophagy in Renal Tubular Epithelial Cells.

PubMed

Tan, Jin; Xie, Qi; Song, Shuling; Miao, Yuyang; Zhang, Qiang

2018-03-01

BACKGROUND Albumin, as a major urinary protein component, is a risk factor for chronic kidney disease progression. Mitochondrial dysfunction is one of the main causes of albumin-induced proximal tubule cells injury. Mitophagy is considered as a pivotal protective mechanism for the elimination of dysfunctional mitochondria. The objective of this research was to determine whether albumin overload-induced mitochondrial dysfunction can activate PINK1/Parkin-mediated mitophagy in renal tubular epithelial cells (TECs). MATERIAL AND METHODS Immunofluorescence assay and Western blot assay were used to detect the effects of albumin overload on autophagy marker protein LC3. Transmission electron microscopy and Western blot assay were used to investigate the role of albumin in mitochondrial injury. Western blot assay and co-localization of acidic lysosomes and mitochondria assay were employed to detect the activation of mitophagy induced by albumin. Finally, we explored the role of PINK1/Parkin signaling in albumin-induced mitophagy by inhibiting mitophagy by knockdown of PARK2 (Parkin) level. RESULTS Immunofluorescence and Western blot results showed that the expression level of LC3-II increased, and the maximum increase point was observed after 8 h of albumin treatment. Transmission electron microscopy results demonstrated that albumin overload-induced mitochondrial injury and quantity of autophagosomes increased. Additionally, expression of PINK1 and cytosolic cytochrome C increased and mitochondria cytochrome C decreased in the albumin group. The co-localization of acidic lysosomes and mitochondria demonstrated that the number of albumin overload-induced mitophagy-positive dots increased. The transient transfection of PARK2 siRNA result showed knockdown of the expression level of PARK2 can inhibit mitophagy induced by albumin. CONCLUSIONS In conclusion, our study suggests that mitochondrial dysfunction activates the PINK1/Parkin signaling and mitophagy in renal tubular epithelial cells under albumin overload condition.

The expression and significance of HIF-1alpha and GLUT-3 in glioma.

PubMed

Liu, Yang; Li, Yun-ming; Tian, Rui-feng; Liu, Wei-ping; Fei, Zhou; Long, Qian-fa; Wang, Xiao-an; Zhang, Xiang

2009-12-22

HIF-1alpha plays an indispensable role in tumor formation and histogenesis. Target genes involved in glucose transport are acutely transactivated by HIF-1alpha. GLUT-3 protein is the rate-limiting factor related to glucose transport, which is classified as brain-type glucose transporter. This study was the initial one aiming to probe into the co-expression and clinical significance of HIF-1alpha and GLUT-3 in glioma. One hundred and twenty cases of glioma tissues and ten human normal cerebral tissues decompressed in glioma excision were examined using immunohistochemistry and Western blot. The expression of HIF-1alpha and GLUT-3 increased gradually with the increase of pathological grade of glioma, respectively. There was significant difference in the expression of HIF-1alpha and GLUT-3 in every two groups, respectively. There was a positive correlation between HIF-1alpha and GLUT-3. In conclusion, the expression of HIF-1alpha and GLUT-3 in glioma was correlated significantly with tumors' pathological grade, which can be taken as a pair of useful markers for predicting the biological behavior of glioma.

Retinoic acid has different effects on UCP1 expression in mouse and human adipocytes

PubMed Central

2013-01-01

Background Increased adipose thermogenesis is being considered as a strategy aimed at preventing or reversing obesity. Thus, regulation of the uncoupling protein 1 (UCP1) gene in human adipocytes is of significant interest. Retinoic acid (RA), the carboxylic acid form of vitamin A, displays agonist activity toward several nuclear hormone receptors, including RA receptors (RARs) and peroxisome proliferator-activated receptor δ (PPARδ). Moreover, RA is a potent positive regulator of UCP1 expression in mouse adipocytes. Results The effects of all-trans RA (ATRA) on UCP1 gene expression in models of mouse and human adipocyte differentiation were investigated. ATRA induced UCP1 expression in all mouse white and brown adipocytes, but inhibited or had no effect on UCP1 expression in human adipocyte cell lines and primary human white adipocytes. Experiments with various RAR agonists and a RAR antagonist in mouse cells demonstrated that the stimulatory effect of ATRA on UCP1 gene expression was indeed mediated by RARs. Consistently, a PPARδ agonist was without effect. Moreover, the ATRA-mediated induction of UCP1 expression in mouse adipocytes was independent of PPARγ coactivator-1α. Conclusions UCP1 expression is differently affected by ATRA in mouse and human adipocytes. ATRA induces UCP1 expression in mouse adipocytes through activation of RARs, whereas expression of UCP1 in human adipocytes is not increased by exposure to ATRA. PMID:24059847

The role of CD147 expression in prostate cancer: a systematic review and meta-analysis.

PubMed

Ye, Yun; Li, Su-Liang; Wang, Yao; Yao, Yang; Wang, Juan; Ma, Yue-Yun; Hao, Xiao-Ke

2016-01-01

There are a number of studies which show that expression of CD147 is increased significantly in prostate cancer (PCa). However, conflicting conclusions have also been reported by other researchers lately. In order to arrive at a clear conclusion, a meta-analysis of eligible studies was conducted. We searched PubMed, MEDLINE, Cochrane Library, and the China National Knowledge Infrastructure databases to identify all the published case-control studies on the relationship between the expression of CD147 and PCa until February 2016. In the end, a total of 930 patients in eight studies were included in the meta-analysis. CD147 expression in the PCa patients increased significantly (odds ratio [OR], 4.65; 95% confidence interval [CI], 3.52-6.14; Z=10.79; P<0.05), but there was obvious heterogeneity between studies (I (2)=92.9%, P<0.05). Subgroup analysis showed that positive expression of CD147 was associated with PCa among the Asian population (OR, 21.01; 95% CI, 12.88-34.28; Z=12.19; P<0.05). Furthermore, it was significantly related to TNM stage (OR, 0.24; 95% CI, 0.17-0.35; Z=7.74; P<0.05), Gleason score (OR, 0.41; 95% CI, 0.31-0.56; Z=5.62; P<0.05), differentiation grade (OR, 0.27; 95% CI, 0.13-0.56; Z=3.47; P<0.05), and pretreatment serum prostate-specific antigen level (OR, 0.07; 95% CI, 0.03-0.16; Z=6.47; P<0.05). Positive expression of CD147 was related to PCa, significant heterogeneity was not found between Asian studies, and the result became more significant. The positive expression of CD147 was significantly related to the clinicopathological characteristics of PCa. This suggests that CD147 plays an essential role in poor prognosis and recurrence prediction.

Increase in apoptosis by combination of metformin with silibinin in human colorectal cancer cells

PubMed Central

Tsai, Cheng-Chia; Chuang, Tang-Wei; Chen, Li-Jen; Niu, Ho-Shan; Chung, Kun-Ming; Cheng, Juei-Tang; Lin, Kao-Chang

2015-01-01

AIM: To investigate the effect of metformin on silibinin-induced apoptosis in human colorectal cancer (COLO 205) cells. METHODS: MTT assays were performed to quantify cell viability. Western blot assays were applied to identify the expression of signaling proteins. RESULTS: The combined treatment of COLO 205 cells with metformin and silibinin decreased cell survival at a dose insufficient to influence the non-malignant cells [Human colonic epithelial cells (HCoEpiC)]. Silibinin and metformin increased phosphatase and tensin homolog and 5’-adenosine monophosphate-activated protein kinase expression in COLO 205 cells and inhibited the phosphorylation of mammol/Lalian target of rapamycin. This combined treatment resulted in an increase in the expression of activated caspase 3 and apoptosis inducing factor, indicating apoptosis. CONCLUSION: The combined treatment of human colorectal cancer cells with silibinin and metformin may induce apoptosis at a dose that does not affect HCoEpiC. This finding reveals a potential therapeutic strategy for the treatment of colorectal cancer. PMID:25892866

Regulation of protein expression and function of octn2 in forskolin-induced syncytialization in BeWo Cells.

PubMed

Huang, F-D; Kung, F-L; Tseng, Y-C; Chen, M-R; Chan, H-S; Lin, C-J

2009-02-01

Placental OCTN2 is a high-affinity carnitine transporter that can interact with a number of therapeutic agents. The process of syncytialization is associated with the expression of a variety of genes. However, the association between syncytialization and OCTN2 expression is not yet clear. Given that forskolin induces BeWo cells to undergo biochemical and morphological differentiation, the purpose of the present study was to investigate whether the function and expression of OCTN2 are influenced by forskolin treatment during syncytialization. The forskolin-induced differentiation of BeWo cells was validated by secretion of beta-human chorionic gonadotropin (beta-hCG) and syncytin expression. Cellular localization of OCTN2 was analyzed by confocal microscopy. Expression of OCTN2 and the modular proteins PDZK1, PDZK2, NHERF1 and NHERF2 was analyzed by Western blotting and carnitine uptake by BeWo cells was estimated and the kinetic properties of uptake measured. The results showed that forskolin treatment increased beta-hCG secretion and syncytin expression, suggesting induction of syncytialization. Confocal images of BeWo cells showed the localization of OCTN2 in the brush-border membrane. OCTN2 protein expression was upregulated in isolated brush-border membranes by long-term forskolin treatment, but the V(m) for carnitine uptake was unchanged, although the K(m) increased. PDZK1, NHERF1 and NHERF2 protein expression in the brush-border membrane was downregulated by forskolin treatment, whereas PDZK2 levels remained unchanged. In conclusion, protein expression and function of OCTN2 in BeWo cells can be regulated by forskolin treatment. While the presence of forskolin results in an increase in OCTN2 protein expression, the increase in uptake capacity may be compensated by the decreased expression of PDZK1, NHERF1 or NHERF2.

The tumour-associated glycoprotein podoplanin is expressed in fibroblast-like synoviocytes of the hyperplastic synovial lining layer in rheumatoid arthritis

PubMed Central

2011-01-01

Introduction Activated fibroblast-like synoviocytes (FLSs) in rheumatoid arthritis (RA) share many characteristics with tumour cells and are key mediators of synovial tissue transformation and joint destruction. The glycoprotein podoplanin is upregulated in the invasive front of several human cancers and has been associated with epithelial-mesenchymal transition, increased cell migration and tissue invasion. The aim of this study was to investigate whether podoplanin is expressed in areas of synovial transformation in RA and especially in promigratory RA-FLS. Methods Podoplanin expression in human synovial tissue from 18 RA patients and nine osteoarthritis (OA) patients was assessed by immunohistochemistry and confirmed by Western blot analysis. The expression was related to markers of synoviocytes and myofibroblasts detected by using confocal immunofluoresence microscopy. Expression of podoplanin, with or without the addition of proinflammatory cytokines and growth factors, in primary human FLS was evaluated by using flow cytometry. Results Podoplanin was highly expressed in cadherin-11-positive cells throughout the synovial lining layer in RA. The expression was most pronounced in areas with lining layer hyperplasia and high matrix metalloproteinase 9 expression, where it coincided with upregulation of α-smooth muscle actin (α-sma). The synovium in OA was predominantly podoplanin-negative. Podoplanin was expressed in 50% of cultured primary FLSs, and the expression was increased by interleukin 1β, tumour necrosis factor α and transforming growth factor β receptor 1. Conclusions Here we show that podoplanin is highly expressed in FLSs of the invading synovial tissue in RA. The concomitant upregulation of α-sma and podoplanin in a subpopulation of FLSs indicates a myofibroblast phenotype. Proinflammatory mediators increased the podoplanin expression in cultured RA-FLS. We conclude that podoplanin might be involved in the synovial tissue transformation and increased migratory potential of activated FLSs in RA. PMID:21385358

Effects of the duration of expressions on the recognition of microexpressions*

PubMed Central

Shen, Xun-bing; Wu, Qi; Fu, Xiao-lan

2012-01-01

Objective: The purpose of this study was to investigate the effects of the duration of expressions on the recognition of microexpressions, which are closely related to deception. Methods: In two experiments, participants were briefly (from 20 to 300 ms) shown one of six basic expressions and then were asked to identify the expression. Results: The results showed that the participants’ performance in recognition of microexpressions increased with the duration of the expressions, reaching a turning point at 200 ms before levelling off. The results also indicated that practice could improve the participants’ performance. Conclusions: The results of this study suggest that the proper upper limit of the duration of microexpressions might be around 1/5 of a second and confirmed that the ability to recognize microexpressions can be enhanced with practice. PMID:22374615

Effect of Relaxin Expressing Adenovirus on Scar Remodeling: A Preliminary Study

PubMed Central

Jung, Bok Ki; Lee, Won Jai; Kang, Eunhye; Ahn, Hyo Min; Kim, Yong Oock; Rah, Dong Kyun; Yun, Chae-Ok

2017-01-01

Background Relaxin is a transforming growth factor β1 antagonist. To determine the effects of relaxin on scar reduction, we investigated the scar remodeling process by injecting relaxin-expressing adenoviruses using a pig scar model. Methods Scars with full thickness were generated on the backs of Yorkshire pigs. Scars were divided into two groups (relaxin [RLX] and Control). Adenoviruses were injected into the RLX (expressing relaxin) and Control (not expressing relaxin) groups. Changes in the surface areas, color index and pliability of scars were compared. Results Fifty days after treatment, the surface areas of scars decreased, the color of scars was normalized, and the pliability of scars increased in RLX group. Conclusion Relaxin-expressing adenoviruses improved the surface area, color, and pliability of scars. The mechanism of therapeutic effects on scar formation should be further investigated. PMID:28913296

Effects of Forskolin on Trefoil factor 1 expression in cultured ventral mesencephalic dopaminergic neurons.

PubMed

Jensen, P; Ducray, A D; Widmer, H R; Meyer, M

2015-12-03

Trefoil factor 1 (TFF1) belongs to a family of secreted peptides that are mainly expressed in the gastrointestinal tract. Notably, TFF1 has been suggested to operate as a neuropeptide, however, its specific cellular expression, regulation and function remain largely unknown. We have previously shown that TFF1 is expressed in developing and adult rat ventral mesencephalic tyrosine hydroxylase-immunoreactive (TH-ir) dopaminergic neurons. Here, we investigated the expression of TFF1 in rat ventral mesencephalic dopaminergic neurons (embryonic day 14) grown in culture for 5, 7 or 10 days in the absence (controls) or presence of either glial cell line-derived neurotrophic factor (GDNF), Forskolin or the combination. No TFF1-ir cells were identified at day 5 and only a few at day 7, whereas TH was markedly expressed at both time points. At day 10, several TFF1-ir cells were detected, and their numbers were significantly increased after the addition of GDNF (2.2-fold) or Forskolin (4.1-fold) compared to controls. Furthermore, the combination of GDNF and Forskolin had an additive effect and increased the number of TFF1-ir cells by 5.6-fold compared to controls. TFF1 expression was restricted to neuronal cells, and the percentage of TH/TFF1 co-expressing cells was increased to the same extent in GDNF and Forskolin-treated cultures (4-fold) as compared to controls. Interestingly, the combination of GDNF and Forskolin resulted in a significantly increased co-expression (8-fold) of TH/TFF1, which could indicate that GDNF and Forskolin targeted different subpopulations of TH/TFF1 neurons. Short-term treatment with Forskolin resulted in an increased number of TFF1-ir cells, and this effect was significantly reduced by the MEK1 inhibitor PD98059 or the protein kinase A (PKA) inhibitor H89, suggesting that Forskolin induced TFF1 expression through diverse signaling pathways. In conclusion, distinct populations of cultured dopaminergic neurons express TFF1, and their numbers can be increased by factors known to influence survival and differentiation of dopaminergic cells. Copyright © 2015 IBRO. Published by Elsevier Ltd. All rights reserved.

Suppressed heat shock protein response in the kidney of exercise-trained diabetic rats.

PubMed

Lappalainen, J; Oksala, N K J; Laaksonen, D E; Khanna, S; Kokkola, T; Kaarniranta, K; Sen, C K; Atalay, M

2018-07-01

Impaired expression of heat shock proteins (HSPs) and increased oxidative stress may contribute to the pathophysiology of diabetes by disrupted tissue protection. Acute exercise induces oxidative stress, whereas exercise training up-regulates endogenous antioxidant defenses and HSP expression. Although diabetic nephropathy is a major contributor to diabetic morbidity, information regarding the effect of HSPs on kidney protection is limited. This study evaluated the effects of eight-week exercise training on kidney HSP expression and markers of oxidative stress at rest and after acute exercise in rats with or without streptozotocin-induced diabetes. Induction of diabetes increased DNA-binding activity of heat shock factor-1, but decreased the expression of HSP72, HSP60, and HSP90. The inflammatory markers IL-6 and TNF-alpha were increased in the kidney tissue of diabetic animals. Both exercise training and acute exercise increased HSP72 and HSP90 protein levels only in non-diabetic rats. On the other hand, exercise training appeared to reverse the diabetes-induced histological changes together with decreased expression of TGF-beta as a key inducer of glomerulosclerosis, and decreased levels of IL-6 and TNF-alpha. Notably, HSP72 and TGF-beta were negatively correlated. In conclusion, impaired HSP defense seems to contribute to kidney injury vulnerability in diabetes and exercise training does not up-regulate kidney HSP expression despite the improvements in histopathological and inflammatory markers. © 2018 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

Evidence against Resveratrol as a viable therapy for the rescue of defective ΔF508 CFTR

PubMed Central

Jai, Ying; Shah, Kalpit; Bridges, Robert J.; Bradbury, Neil A.

2015-01-01

BACKGROUND Resveratrol, a natural phenolic compound, has been reported to rescue mutant ΔF508 CFTR in expression systems and primary epithelial cells. Although this implies a therapeutic benefit to patients with CF, investigations were performed using resveratrol concentrations greatly in excess of those achievable in plasma. We evaluated the efficacy of resveratrol as a CFTR corrector in relevant primary airway cells, using physiologically achievable resveratrol concentrations. METHODS Cells expressing wt or ΔF508 CFTR were exposed to chronic or acute resveratrol. CFTR mRNA and protein expression were monitored. The effects of resveratrol on primary ΔF508 human airway cells were evaluated by equivalent current analysis using modified Ussing chambers. RESULTS Consistent with previously published data in heterologous expression systems, high doses of resveratrol increased CFTR expression; however physiologically relevant concentrations were without effect. In contrast to heterologous expression systems, resveratrol was unable to increase mutant CFTR channel activity in primary airway cells. Elevated amiloride-sensitive currents, indicative of sodium transport and characteristically elevated in CF airway cells, were also unaffected by resveratrol CONCLUSIONS High concentrations of resveratrol can increase CFTR mRNA and protein in some cell types. In addition, acute resveratrol exposure can stimulate CFTR mediated chloride secretion, probably by increasing cellular cAMP levels. Resveratrol at physiologically achievable levels yielded no benefit in primary ΔF508 airway cells, either in terms of amiloride-sensitive currents of CFTR currents. PMID:26342647

Hepatitis C virus core protein potentiates proangiogenic activity of hepatocellular carcinoma cells.

PubMed

Shao, Yu-Yun; Hsieh, Min-Shu; Wang, Han-Yu; Li, Yong-Shi; Lin, Hang; Hsu, Hung-Wei; Huang, Chung-Yi; Hsu, Chih-Hung; Cheng, Ann-Lii

2017-10-17

Increased angiogenic activity has been demonstrated in hepatitis C virus (HCV)-related hepatocellular carcinoma (HCC), but the mechanism was unclear. To study the role of HCV core protein, we used tube formation and Matrigel plug assays to assess the proangiogenic activity of an HCC cell line, HuH7, and 2 of its stable clones-HuH7-core-high and HuH7-core-low, with high and low HCV core protein expression, respectively. In both assays, HuH7-core-high and HuH7-core-low cells dose-dependently induced stronger angiogenesis than control cells. HuH7 cells with HCV core protein expression showed increased mRNA and protein expression of vascular endothelial growth factor (VEGF). VEGF inhibition by bevacizumab reduced the proangiogenic activity of HuH7-core-high cells. The promotor region of VEGF contains the binding site of activator protein-1 (AP-1). Compared with controls, HuH7-core-high cells had an increased AP-1 activity and nuclear localization of phospho-c-jun. AP-1 inhibition using either RNA knockdown or AP-1 inhibitors reduced the VEGF mRNA expression and the proangiogenic activity of HuH7-core-high cells. Among 131 tissue samples from HCC patients, HCV-related HCC revealed stronger VEGF expression than did hepatitis B virus-related HCC. In conclusion, increased VEGF expression through AP-1 activation is a crucial mechanism underlying the proangiogenic activity of the HCV core protein in HCC cells.

Hepatitis C virus core protein potentiates proangiogenic activity of hepatocellular carcinoma cells

PubMed Central

Shao, Yu-Yun; Hsieh, Min-Shu; Wang, Han-Yu; Li, Yong-Shi; Lin, Hang; Hsu, Hung-Wei; Huang, Chung-Yi; Hsu, Chih-Hung; Cheng, Ann-Lii

2017-01-01

Increased angiogenic activity has been demonstrated in hepatitis C virus (HCV)-related hepatocellular carcinoma (HCC), but the mechanism was unclear. To study the role of HCV core protein, we used tube formation and Matrigel plug assays to assess the proangiogenic activity of an HCC cell line, HuH7, and 2 of its stable clones—HuH7-core-high and HuH7-core-low, with high and low HCV core protein expression, respectively. In both assays, HuH7-core-high and HuH7-core-low cells dose-dependently induced stronger angiogenesis than control cells. HuH7 cells with HCV core protein expression showed increased mRNA and protein expression of vascular endothelial growth factor (VEGF). VEGF inhibition by bevacizumab reduced the proangiogenic activity of HuH7-core-high cells. The promotor region of VEGF contains the binding site of activator protein-1 (AP-1). Compared with controls, HuH7-core-high cells had an increased AP-1 activity and nuclear localization of phospho-c-jun. AP-1 inhibition using either RNA knockdown or AP-1 inhibitors reduced the VEGF mRNA expression and the proangiogenic activity of HuH7-core-high cells. Among 131 tissue samples from HCC patients, HCV-related HCC revealed stronger VEGF expression than did hepatitis B virus-related HCC. In conclusion, increased VEGF expression through AP-1 activation is a crucial mechanism underlying the proangiogenic activity of the HCV core protein in HCC cells. PMID:29156827

Enhancing solubility of deoxyxylulose phosphate pathway enzymes for microbial isoprenoid production

PubMed Central

2012-01-01

Background Recombinant proteins are routinely overexpressed in metabolic engineering. It is well known that some over-expressed heterologous recombinant enzymes are insoluble with little or no enzymatic activity. This study examined the solubility of over-expressed homologous enzymes of the deoxyxylulose phosphate pathway (DXP) and the impact of inclusion body formation on metabolic engineering of microbes. Results Four enzymes of this pathway (DXS, ISPG, ISPH and ISPA), but not all, were highly insoluble, regardless of the expression systems used. Insoluble dxs (the committed enzyme of DXP pathway) was found to be inactive. Expressions of fusion tags did not significantly improve the solubility of dxs. However, hypertonic media containing sorbitol, an osmolyte, successfully doubled the solubility of dxs, with the concomitant improvement in microbial production of the metabolite, DXP. Similarly, sorbitol significantly improved the production of soluble and functional ERG12, the committed enzyme in the mevalonate pathway. Conclusion This study demonstrated the unanticipated findings that some over-expressed homologous enzymes of the DXP pathway were highly insoluble, forming inclusion bodies, which affected metabolite formation. Sorbitol was found to increase both the solubility and function of some of these over-expressed enzymes, a strategy to increase the production of secondary metabolites. PMID:23148661

Transgenic Muscle-Specific Nor-1 Expression Regulates Multiple Pathways That Effect Adiposity, Metabolism, and Endurance

PubMed Central

Pearen, Michael A.; Goode, Joel M.; Fitzsimmons, Rebecca L.; Eriksson, Natalie A.; Thomas, Gethin P.; Cowin, Gary J.; Wang, S.-C. Mary; Tuong, Zewen K.

2013-01-01

The mRNA encoding Nor-1/NR4A3 is rapidly and strikingly induced by β2-adrenergic signaling in glycolytic and oxidative skeletal muscle. In skeletal muscle cells, Nor-1 expression is important for the regulation of oxidative metabolism. Transgenic skeletal muscle-specific expression of activated Nor-1 resulted in the acquisition of an endurance phenotype, an increase in type IIA/X oxidative muscle fibers, and increased numbers of mitochondria. In the current study, we used dual-energy x-ray absorptiometry and magnetic resonance imaging analysis to demonstrate decreased adiposity in transgenic (Tg) Nor-1 mice relative to that in wild-type littermates. Furthermore, the Tg-Nor-1 mice were resistant to diet-induced weight gain and maintained fasting glucose at normoglycemic levels. Expression profiling and RT-quantitative PCR analysis revealed significant increases in genes involved in glycolysis, the tricarboxylic acid cycle, oxidative phosphorylation, fatty acid oxidation, and glycogen synthesis, in concordance with the lean phenotype. Moreover, expression profiling identified several Z-disc and sarcomeric binding proteins that modulate fiber type phenotype and endurance, eg, α-actinin-3. In addition, we demonstrated that the Tg-Nor-1 mouse line has significantly higher glycogen content in skeletal muscle relative to that in wild-type littermates. Finally, we identified a decreased NAD+/NADH ratio with a concordant increase in peroxisome proliferator-activated receptor γ coactivator-1α1 protein/mRNA expression. Increased NADH was associated with an induction of the genes involved in the malate-aspartate shuttle and a decrease in the glycerol 3-phosphate shuttle, which maximizes aerobic ATP production. In conclusion, skeletal muscle-specific Nor-1 expression regulates genes and pathways that regulate adiposity, muscle fiber type metabolic capacity, and endurance. PMID:24065705

Albumin-induced apoptosis of tubular cells is modulated by BASP1

PubMed Central

Sanchez-Niño, M D; Fernandez-Fernandez, B; Perez-Gomez, M V; Poveda, J; Sanz, A B; Cannata-Ortiz, P; Ruiz-Ortega, M; Egido, J; Selgas, R; Ortiz, A

2015-01-01

Albuminuria promotes tubular injury and cell death, and is associated with faster progression of chronic kidney disease (CKD) to end-stage renal disease. However, the molecular mechanisms regulating tubular cell death in response to albuminuria are not fully understood. Brain abundant signal protein 1 (BASP1) was recently shown to mediate glucose-induced apoptosis in tubular cells. We have studied the role of BASP1 in albumin-induced tubular cell death. BASP1 expression was studied in experimental puromycin aminonucleoside-induced nephrotic syndrome in rats and in human nephrotic syndrome. The role of BASP1 in albumin-induced apoptosis was studied in cultured human HK2 proximal tubular epithelial cells. Puromycin aminonucleoside induced proteinuria and increased total kidney BASP1 mRNA and protein expression. Immunohistochemistry localized the increased BASP1 to tubular cells. BASP1 expression colocalized with deoxynucleotidyl-transferase-mediated dUTP nick-end labeling staining for apoptotic cells. Increased tubular BASP1 expression was observed in human proteinuric nephropathy by immunohistochemistry, providing evidence for potential clinical relevance. In cultured tubular cells, albumin induced apoptosis and increased BASP1 mRNA and protein expression at 6–48 h. Confocal microscopy localized the increased BASP1 expression in albumin-treated cells mainly to the perinuclear area. A peripheral location near the cell membrane was more conspicuous in albumin-treated apoptotic cells, where it colocalized with actin. Inhibition of BASP1 expression by a BASP1 siRNA protected from albumin-induced apoptosis. In conclusion, albumin-induced apoptosis in tubular cells is BASP1-dependent. This information may be used to design novel therapeutic approaches to slow CKD progression based on protection of tubular cells from the adverse consequences of albuminuria. PMID:25675304

4-PBA improves lithium-induced nephrogenic diabetes insipidus by attenuating ER stress.

PubMed

Zheng, Peili; Lin, Yu; Wang, Feifei; Luo, Renfei; Zhang, Tiezheng; Hu, Shan; Feng, Pinning; Liang, Xinling; Li, Chunling; Wang, Weidong

2016-10-01

Endoplasmic reticulum (ER) stress has been implicated in some types of glomerular and tubular disorders. The objectives of this study were to elucidate the role of ER stress in lithium-induced nephrogenic diabetes insipidus (NDI) and to investigate whether attenuation of ER stress by 4-phenylbutyric acid (4-PBA) improves urinary concentrating defect in lithium-treated rats. Wistar rats received lithium (40 mmol/kg food), 4-PBA (320 mg/kg body wt by gavage every day), or no treatment (control) for 2 wk, and they were dehydrated for 24 h before euthanasia. Lithium treatment resulted in increased urine output and decreased urinary osmolality, which was significantly improved by 4-PBA. 4-PBA also prevented reduced protein expression of aquaporin-2 (AQP2), pS256-AQP2, and pS261-AQP2 in the inner medulla of kidneys from lithium-treated rats after 24-h dehydration. Lithium treatment resulted in increased expression of ER stress markers in the inner medulla, which was associated with dilated cisternae and expansion of ER in the inner medullary collecting duct (IMCD) principal cells. Confocal immunofluorescence studies showed colocalization of a molecular chaperone, binding IgG protein (BiP), with AQP2 in principal cells. Immunohistochemistry demonstrated increased intracellular expression of BiP and decreased AQP2 expression in IMCD principal cells of kidneys from lithium-treated rats. 4-PBA attenuated expression of ER stress markers and recovered ER morphology. In IMCD suspensions isolated from lithium-treated rats, 4-PBA incubation was also associated with increased AQP2 expression and ameliorated ER stress. In conclusion, in experimental lithium-induced NDI, 4-PBA improved the urinary concentrating defect and increased AQP2 expression, likely via attenuating ER stress in IMCD principal cells. Copyright © 2016 the American Physiological Society.

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

Cheng, Xingguo, E-mail: chengx@stjohns.edu; Vispute, Saurabh G.; Liu, Jie

The toxic effects of dioxins, such as 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), mainly through activation of the aryl hydrocarbon receptor (AhR) are well documented. Fibroblast growth factor (Fgf) 21 plays critical roles in metabolic adaptation to fasting by increasing lipid oxidation and ketogenesis in the liver. The present study was performed to determine whether activation of the AhR induces Fgf21 expression. In mouse liver, TCDD increased Fgf21 mRNA in both dose- and time-dependent manners. In addition, TCDD markedly increased Fgf21 mRNA expression in cultured mouse and human hepatocytes. Moreover, TCDD increased mRNA (in liver) and protein levels (in both liver and serum) ofmore » Fgf21 in wild-type mice, but not in AhR-null mice. Chromatin immunoprecipitation assays showed that TCDD increased AhR protein binding to the Fgf21 promoter (− 105/+ 1 base pair). Fgf21-null mice administered 200 μg/kg of TCDD died within 20 days, whereas wild-type mice receiving the same treatment were still alive at one month after administration. This indicates that TCDD-induced Fgf21 expression protects against TCDD toxicity. Diethylhexylphthalate (DEHP) pretreatment attenuated TCDD-induced Fgf21 expression in mouse liver and white adipose tissue, which may explain a previous report that DEHP pretreatment decreases TCDD-induced wasting. In conclusion, Fgf21 appears to be a target gene of AhR-signaling pathway in mouse and human liver. - Highlights: • TCDD induced Fgf21 expression at both mRNA and protein levels. • Fgf21 induction by TCDD is AhR-dependent. • DEHP attenuated TCDD-induced Fgf21 expression.« less

Mucin (MUC) expression in EUS-FNA specimens is a useful prognostic factor in pancreatic ductal adenocarcinoma

PubMed Central

Higashi, Michiyo; Yokoyama, Seiya; Yamamoto, Takafumi; Goto, Yuko; Kitazono, Ikumi; Hiraki, Tsubasa; Taguchi, Hiroki; Hashimoto, Shinichi; Fukukura, Yoshihiko; Koriyama, Chihaya; Mataki, Yuko; Maemura, Kosei; Shinchi, Hiroyuki; Jain, Maneesh; Batra, Surinder K.; Yonezawa, Suguru

2015-01-01

Objectives The aim of this study was to further examine the utility of mucin expression profiles as prognostic factors in PDAC. Methods Mucin (MUC) expression was examined by immunohistochemistry (IHC) analysis in endoscopic ultrasound-guided fine-needle aspiration (EUS-FNA) specimens obtained from 114 patients with PDAC. The rate of expression of each mucin was compared with clinicopathologic features. Results The expression rates of mucins in cancer lesions were MUC1, 87.7%; MUC2, 0.8%; MUC4, 93.0%; MUC5AC, 78.9%; MUC6, 24.6%; and MUC16, 67.5%. MUC1 and MUC4 were positive and MUC2 was negative in most PDACs. Patients with advanced stage of PDAC with MUC5AC expression had a significantly better outcome than those who were MUC5AC-negative (P=0.002).With increasing clinical stage, total MUC6 expression decreased (P for trend=0.001) and MUC16 cytoplasmic expression increased (P for trend=0.02). The prognosis of patients with MUC16 cytoplasmic expression was significantly poorer than those without this expression. Multivariate survival analysis revealed that MUC16 cytoplasmic expression was a significant independent predictor of a poor prognosis after adjusting for the effects of other prognostic factors (P=0.002). Conclusion Mucin expression profiles in EUS-FNA specimens have excellent diagnostic utility and are useful predictors of outcome in patients with PDAC. PMID:25906442

Korean Red Ginseng inhibits apoptosis in neuroblastoma cells via estrogen receptor β-mediated phosphatidylinositol-3 kinase/Akt signaling

PubMed Central

Nguyen, Cuong Thach; Luong, Truc Thanh; Kim, Gyu-Lee; Pyo, Suhkneung; Rhee, Dong-Kwon

2014-01-01

Background Ginseng has been shown to exert antistress effects both in vitro and in vivo. However, the effects of ginseng on stress in brain cells are not well understood. This study investigated how Korean Red Ginseng (KRG) controls hydrogen peroxide-induced apoptosis via regulation of phosphatidylinositol-3 kinase (PI3K)/Akt and estrogen receptor (ER)-β signaling. Methods Human neuroblastoma SK-N-SH cells were pretreated with KRG and subsequently exposed to H2O2. The ability of KRG to inhibit oxidative stress-induced apoptosis was assessed in MTT cytotoxicity assays. Apoptotic protein expression was examined by Western blot analysis. The roles of ER-β, PI3K, and p-Akt signaling in KRG regulation of apoptosis were studied using small interfering RNAs and/or target antagonists. Results Pretreating SK-N-SH cells with KRG decreased expression of the proapoptotic proteins p-p53 and caspase-3, but increased expression of the antiapoptotic protein BCL2. KRG pretreatment was also associated with increased ER-β, PI3K, and p-Akt expression. Conversely, ER-β inhibition with small interfering RNA or inhibitor treatment increased p-p53 and caspase-3 levels, but decreased BCL2, PI3K, and p-Akt expression. Moreover, inhibition of PI3K/Akt signaling diminished p-p53 and caspase-3 levels, but increased BCL2 expression. Conclusion Collectively, the data indicate that KRG represses oxidative stress-induced apoptosis by enhancing PI3K/Akt signaling via upregulation of ER-β expression. PMID:25535479

Living high training low induces physiological cardiac hypertrophy accompanied by down-regulation and redistribution of the renin-angiotensin system

PubMed Central

Shi, Wei; Meszaros, J Gary; Zeng, Shao-ju; Sun, Ying-yu; Zuo, Ming-xue

2013-01-01

Aim: Living high training low” (LHTL) is an exercise-training protocol that refers living in hypoxia stress and training at normal level of O2. In this study, we investigated whether LHTL caused physiological heart hypertrophy accompanied by changes of biomarkers in renin-angiotensin system in rats. Methods: Adult male SD rats were randomly assigned into 4 groups, and trained on living low-sedentary (LLS, control), living low-training low (LLTL), living high-sedentary (LHS) and living high-training low (LHTL) protocols, respectively, for 4 weeks. Hematological parameters, hemodynamic measurement, heart hypertrophy and plasma angiotensin II (Ang II) level of the rats were measured. The gene and protein expression of angiotensin-converting enzyme (ACE), angiotensinogen (AGT) and angiotensin II receptor I (AT1) in heart tissue was assessed using RT-PCR and immunohistochemistry, respectively. Results: LLTL, LHS and LHTL significantly improved cardiac function, increased hemoglobin concentration and RBC. At the molecular level, LLTL, LHS and LHTL significantly decreased the expression of ACE, AGT and AT1 genes, but increased the expression of ACE and AT1 proteins in heart tissue. Moreover, ACE and AT1 protein expression was significantly increased in the endocardium, but unchanged in the epicardium. Conclusion: LHTL training protocol suppresses ACE, AGT and AT1 gene expression in heart tissue, but increases ACE and AT1 protein expression specifically in the endocardium, suggesting that the physiological heart hypertrophy induced by LHTL is regulated by region-specific expression of renin-angiotensin system components. PMID:23377552

Central adiponectin administration reveals new regulatory mechanisms of bone metabolism in mice.

PubMed

Wu, Yuwei; Tu, Qisheng; Valverde, Paloma; Zhang, Jin; Murray, Dana; Dong, Lily Q; Cheng, Jessica; Jiang, Hua; Rios, Maribel; Morgan, Elise; Tang, Zhihui; Chen, Jake

2014-06-15

Adiponectin (APN), the most abundant adipocyte-secreted adipokine, regulates energy homeostasis and exerts well-characterized insulin-sensitizing properties. The peripheral or central effects of APN regulating bone metabolism are beginning to be explored but are still not clearly understood. In the present study, we found that APN-knockout (APN-KO) mice fed a normal diet exhibited decreased trabecular structure and mineralization and increased bone marrow adiposity compared with wild-type (WT) mice. APN intracerebroventricular infusions decreased uncoupling protein 1 (UCP1) expression in brown adipose tissue, epinephrine and norepinephrine serum levels, and osteoclast numbers, whereas osteoblast osteogenic marker expression and trabecular bone mass increased in APN-KO and WT mice. In addition, centrally administered APN increased hypothalamic tryptophan hydroxylase 2 (TPH2), cocaine- and amphetamine-regulated transcript (CART), and 5-hydroxytryptamine (serotonin) receptor 2C (Htr2C) expressions but decreased hypothalamic cannabinoid receptor-1 expression. Treatment of immortalized mouse neurons with APN demonstrated that APN-mediated effects on TPH2, CART, and Htr2C expression levels were abolished by downregulating adaptor protein containing pleckstrin homology domain, phosphotyrosine domain, and leucine zipper motif (APPL)-1 expression. Pharmacological increase in sympathetic activity stimulated adipogenic differentiation of bone marrow stromal cells (BMSC) and reversed APN-induced expression of the lysine-specific demethylases involved in regulating their commitment to the osteoblastic lineage. In conclusion, we found that APN regulates bone metabolism via central and peripheral mechanisms to decrease sympathetic tone, inhibit osteoclastic differentiation, and promote osteoblastic commitment of BMSC. Copyright © 2014 the American Physiological Society.

Hormonal regulation of β-myosin heavy chain expression in the mouse left ventricle.

PubMed

Patrizio, Mario; Musumeci, Marco; Piccone, Ambra; Raggi, Carla; Mattei, Elisabetta; Marano, Giuseppe

2013-03-01

We investigated the influence of sex hormones on the expression of α- and β-cardiac myosin heavy chain isoforms (α-MHC and β-MHC) in C57bl/6 mice of both sexes under physiological and pathological conditions. In the left ventricles (LVs) of fertile female mice, β-MHC expression was tenfold higher compared with the age-matched males, whereas no difference was found in α-MHC expression. These differences disappeared after ovariectomy or in immature mice. We also found a sex-related difference in expression of β-adrenoceptors (β1-AR), as mRNA levels of this gene were 40% lower in fertile females compared with males of the same age but did not differ in prepubertal or ovariectomized animals. Interestingly, the deletion of both β1- and β2-ARs abolished sex difference of β-MHC expression, as mRNA levels in the LVs of knockout males were increased and reached values comparable to those of knockout females. Moreover, the β1-AR antagonist metoprolol induced about a threefold increase in β-MHC expression in adult male mice. The capability of gender to regulate β-MHC expression was also evaluated in the presence of hemodynamic overload. Thoracic aortic coarctation (TAC) produced cardiac hypertrophy along with a 12-fold increase in β-MHC and a 50% decrease in β1-AR expression in males but not in females, thus abolishing the gender difference observed in sham animals for such genes. By contrast, TAC did not change β2-AR expression. In conclusion, our results show that the expression of β-MHC and β1-AR in the LVs undergo gender-related and correlated changes under both physiological and pathological conditions and suggest a role of β1-AR-mediated signaling.

Expression of Immune Genes on Chromosome 6p21.3-22.1 in Schizophrenia

PubMed Central

Sinkus, Melissa L.; Adams, Catherine E.; Logel, Judith; Freedman, Robert; Leonard, Sherry

2013-01-01

Schizophrenia is a common mental illness with a large genetic component. Three genome-wide association studies have implicated the major histocompatibility complex gene region on chromosome 6p21.3-22.1 in schizophrenia. In addition, nicotine, which is commonly abused in schizophrenia, affects the expression of central nervous system immune genes. Messenger RNA levels for genes in the 6p21.3-22.1 region were measured in human postmortem hippocampus of 89 subjects. The effects of schizophrenia diagnosis, smoking and systemic inflammatory illness were compared. Cell-specific expression patterns for the class I major histocompatibility complex gene HLA-A were explored utilizing in situ hybridization. Expression of five genes was altered in schizophrenic subjects. Messenger RNA levels for the class I major histocompatibility complex antigen HLA-B were increased in schizophrenic nonsmokers, while levels for smokers were indistinguishable from those of controls. β2 microglobulin, HLA-A and Notch4 were all expressed in a pattern where inflammatory illness was associated with increased expression in controls but not in subjects with schizophrenia. Schizophrenia was also associated with increased expression of Butyrophilin 2A2. HLA-A was expressed in glutamatergic and GABAergic neurons in the dentate gyrus, hilus, and the stratum pyramidale of the CA1-CA4 regions of the hippocampus, but not in astrocytes. In conclusion, the expression of genes from the major histocompatibility complex region of chromosome 6 with likely roles in synaptic development is altered in schizophrenia. There were also significant interactions between schizophrenia diagnosis and both inflammatory illness and smoking. PMID:23395714

Periostin in Mature Stage Localized Scleroderma

PubMed Central

Kim, Min-Woo; Park, Jung Tae; Kim, Jung Ho; Koh, Seong-Joon; Yoon, Hyun-Sun; Cho, Soyun

2017-01-01

Background Periostin is a novel matricellular protein expressed in many tissues, including bone, periodontal ligament, and skin. Although its expression is prominent in various fibrotic conditions, studies of periostin in localized scleroderma are rare. Objective To investigate the expression of periostin and other molecules in localized scleroderma. Methods A retrospective study of 14 patients with confirmed mature stage localized scleroderma was undertaken. Fourteen age-matched and biopsy site-matched subjects with normal skin were included as controls. Collagen fiber deposition, periostin, procollagen, transforming growth factor-β, and matrix metalloproteinase (MMP)-1 expression were assessed and compared between the two groups. Co-localization of α-smooth muscle actin and periostin was evaluated using confocal microscopy. Results Periostin was predominantly expressed along the dermo-epidermal junction in the controls. Conversely, patients with localized scleroderma demonstrated increased collagen fiber deposition and periostin expression that was more widely distributed along the entire dermis. MMP-1 staining showed increased expression in the epidermis and dermis of patients compared to scanty expression in the controls. A semi-quantitative evaluation showed a higher proportion of excessive collagen bundle deposition (57.1% vs. 7.1%, p=0.013), diffuse periostin positivity (42.9% vs. 0%, p=0.016), and moderate MMP-1 positivity (71.4% vs. 7.1%, p=0.001) in patients than in the controls. Conclusion Compared to the controls, patients with localized scleroderma had enhanced periostin expression corresponding to increased collagen fiber deposition and unexpected overexpression of MMP-1. The results of this human in vivo study may implicate the pathogenesis of localized scleroderma. PMID:28566901

Tissue Factor Inflammatory Response Regulated by Promoter Genotype and p38 MAPK in Neonatal vs. Adult Microvascular Endothelial Cells

PubMed Central

Buzby, Jeffrey S.; Williams, Shirley A.; Imfeld, Karen L.; Kunicki, Thomas J.; Nugent, Diane J.

2014-01-01

Objective and design Variable tissue factor (TF) expression by human microvascular endothelial cells (HMVEC) may be regulated by two promoter haplotypes, distinguished by an 18 base pair deletion (D) or insertion (I) at -1208. We sought to determine the relationship between these haplotypes and interleukin-1 (IL-1α)-induced TF expression in neonatal versus adult HMVEC. Results IL-1-stimulated TF mRNA, protein, and activity were significantly higher in neonatal compared to adult D/D donors. IL-1-stimulated HMVEC from neonatal D/D donors expressed 3-fold higher levels of TF mRNA, 2-fold higher TF protein, and 4-fold increased TF activity compared to HMVEC from adult D/D donors. These results indicate that homozygosity for the D haplotype is characterized by increased response to IL-1 in neonates but not adults. IL-1 induced increased phosphorylation of p38 mitogen-activated protein kinase (MAPK), which was significantly greater in neonatal compared to adult HMVEC. Moreover, inhibition of the p38 MAPK pathway reduced IL-1-stimulated TF mRNA expression in D/D neonatal but not adult HMVEC. Conclusions Up-regulation of D/D neonatal HMVEC TF expression by IL-1 is mediated through the p38 MAPK pathway. This heightened response of D/D neonatal HMVEC to inflammatory stimuli may contribute to increased microvascular coagulopathies in susceptible newborn infants. PMID:24385191

Cigarette smoke increases Toll-like receptor 4 and modifies lipopolysaccharide-mediated responses in airway epithelial cells

PubMed Central

Pace, Elisabetta; Ferraro, Maria; Siena, Liboria; Melis, Mario; Montalbano, Angela M; Johnson, Malcolm; Bonsignore, Maria R; Bonsignore, Giovanni; Gjomarkaj, Mark

2008-01-01

Airway epithelium is emerging as a regulator of innate immune responses to a variety of insults including cigarette smoke. The main goal of this study was to explore the effects of cigarette smoke extracts (CSE) on Toll-like receptor (TLR) expression and activation in a human bronchial epithelial cell line (16-HBE). The CSE increased the expression of TLR4 and the lipopolysaccharide (LPS) binding, the nuclear factor-κB (NF-κB) activation, the release of interleukin-8 (IL-8) and the chemotactic activity toward neutrophils. It did not induce TLR2 expression or extracellular signal-regulated signal kinase 1/2 (ERK1/2) activation. The LPS increased the expression of TLR4 and induced both NF-κB and ERK1/2 activation. The combined exposure of 16-HBE to CSE and LPS was associated with ERK activation rather than NF-κB activation and with a further increase of IL-8 release and of chemotactic activity toward neutrophils. Furthermore, CSE decreased the constitutive interferon-inducible protein-10 (IP-10) release and counteracted the effect of LPS in inducing both the IP-10 release and the chemotactic activity toward lymphocytes. In conclusion, cigarette smoke, by altering the expression and the activation of TLR4 via the preferential release of IL-8, may contribute to the accumulation of neutrophils within the airways of smokers. PMID:18217953

Effects of the antipsychotic paliperidone on stress-induced changes in the endocannabinoid system in rat prefrontal cortex.

PubMed

MacDowell, Karina S; Sayd, Aline; García-Bueno, Borja; Caso, Javier R; Madrigal, José L M; Leza, Juan Carlos

2017-09-01

Objectives There is a need to explore novel mechanisms of action of existing/new antipsychotics. One potential candidate is the endocannabinoid system (ECS). The present study tried to elucidate the effects of the antipsychotic paliperidone on stress-induced ECS alterations. Methods Wister rats were submitted to acute/chronic restraint stress. Paliperidone (1 mg/kg) was given prior each stress session. Cannabinoid receptors and endocannabinoids (eCBs) synthesis and degradation enzymes were measured in prefrontal cortex (PFC) samples by RT-PCR and Western Blot. Results In the PFC of rats exposed to acute stress, paliperidone increased CB1 receptor (CB1R) expression. Furthermore, paliperidone increased the expression of the eCB synthesis enzymes N-acylphosphatidylethanolamine- hydrolysing phospholipase D and DAGLα, and blocked the stress-induced increased expression of the degrading enzyme fatty acid amide hydrolase. In chronic conditions, paliperidone prevented the chronic stress-induced down-regulation of CB1R, normalised DAGLα expression and reverted stress-induced down-regulation of the 2-AG degrading enzyme monoacylglycerol lipase. ECS was analysed also in periphery. Acute stress decreased DAGLα expression, an effect prevented by paliperidone. Contrarily, chronic stress increased DAGLα and this effect was potentiated by paliperidone. Conclusions The results obtained described a preventive effect of paliperidone on stress-induced alterations in ECS. Considering the diverse alterations on ECS described in psychotic disease, targeting ECS emerges as a new therapeutic possibility.

Increased Retinal Expression of the Pro-Angiogenic Receptor GPR91 via BMP6 in a Mouse Model of Juvenile Hemochromatosis

PubMed Central

Arjunan, Pachiappan; Gnanaprakasam, Jaya P.; Ananth, Sudha; Romej, Michelle A.; Rajalakshmi, Veeranan-Karmegam; Prasad, Puttur D.; Martin, Pamela M.; Gurusamy, Mariappan; Thangaraju, Muthusamy; Bhutia, Yangzom D.; Ganapathy, Vadivel

2016-01-01

Purpose Hemochromatosis, an iron-overload disease, occurs as adult and juvenile types. Mutations in hemojuvelin (HJV), an iron-regulatory protein and a bone morphogenetic protein (BMP) coreceptor, underlie most of the juvenile type. Hjv−/− mice accumulate excess iron in retina and exhibit aberrant vascularization and angiomas. A succinate receptor, GPR91, is pro-angiogenic in retina. We hypothesized that Hjv−/− retinas have increased BMP signaling and increased GPR91 expression as the basis of angiomas. Methods Expression of GPR91 was examined by qPCR, immunofluorescence, and Western blot in wild-type and Hjv−/− mouse retinas and pRPE cells. Influence of excess iron and BMP6 on GPR91 expression was investigated in ARPE-19 cells, and wild-type and Hjv−/− pRPE cells. Succinate was used to activate GPR91 and determine the effects of GPR91 signaling on VEGF expression. Signaling of BMP6 was studied by the expression of Smad1/5/8 and pSmad4, and the BMP-target gene Id1. The interaction of pSmad4 with GPR91 promoter was studied by ChIP. Results Expression of GPR91 was higher in Hjv−/− retinas and RPE than in wild-type counterparts. Unexpectedly, BMP signaling was increased, not decreased, in Hjv−/− retinas and RPE. Bone morphogenetic protein 6 induced GPR91 in RPE, suggesting that increased BMP signaling in Hjv−/− retinas was likely responsible for GPR91 upregulation. Exposure of RPE to excess iron and succinate as well as BMP6 and succinate increased VEGF expression. Bone morphogenetic protein 6 promoted the interaction of pSmad4 with GPR91 promoter in RPE. Conclusions G-protein-coupled receptor 91 is a BMP6 target and Hjv deletion enhances BMP signaling in retina, thus underscoring a role for excess iron and hemochromatosis in abnormal retinal vascularization. PMID:27046124

Increased DNA methylation of scavenger receptor class B type I contributes to inhibitory effects of prenatal caffeine ingestion on cholesterol uptake and steroidogenesis in fetal adrenals

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

Wu, Dong-Mei; He, Zheng; Ma, Liang-Peng

Steroid hormones synthesized from cholesterol in the fetal adrenal are crucial for fetal development. We have observed the inhibited fetal adrenal corticosterone synthesis and increased intrauterine growth retardation (IUGR) rate in rats under prenatal caffeine ingestion. The aim of this study is to evaluate the effects of prenatal caffeine ingestion on cholesterol supply in fetal adrenal steroidogenesis in rats and explore the underlying epigenetic mechanisms. Pregnant Wistar rats were treated with 60 mg/kg·d caffeine from gestational day (GD) 7 to GD17. Histological changes of fetal adrenals and increased IUGR rates were observed in the caffeine group. There were significantly decreasedmore » steroid hormone contents and cholesterol supply in caffeine-treated fetal adrenals. Data from the gene expression array suggested that prenatal caffeine ingestion caused increased expression of genes related to DNA methylation and decreased expression of genes related to cholesterol uptake. The following conjoint analysis of DNA methylation array with these differentially expressed genes suggested that scavenger receptor class B type I (SR-BI) may play an important role in caffeine-induced cholesterol supply deficiency. Moreover, real-time RT-PCR and immunohistochemical detection certified the inhibitory effects of caffeine on both mRNA expression and protein expression of SR-BI in the fetal adrenal. And the increased DNA methylation frequency in the proximal promoter of SR-BI was confirmed by bisulfite-sequencing PCR. In conclusion, prenatal caffeine ingestion can induce DNA hypermethylation of the SR-BI promoter in the rat fetal adrenal. These effects may lead to decreased SR-BI expression and cholesterol uptake, which inhibits steroidogenesis in the fetal adrenal. - Highlights: • Prenatal caffeine ingestion inhibits steroid hormone production in the fetal adrenal. • Prenatal caffeine ingestion inhibits cholesterol uptake in the fetal adrenal. • Prenatal caffeine ingestion inhibits the expression of SR-BI. • Prenatal caffeine ingestion induces increased DNA methylation of SR-BI promoter.« less

The Proteins from Sika deer antler as potential modulators on cisplatin-induced cytotoxicity in human embryonic kidney 293 cells.

PubMed

Yang, Huihai; Li, Wei; Wang, Lulu; He, Xiaofeng; Sun, Hang; Zhang, Jing

2017-07-31

Our study aimed to investigate the protective role of SDAPR on cisplatin-induced cytotoxicity and its' possible mechanism in HEK293 cells. Cell viability was measured by MTT assay. Oxidative stress (SOD, GSH, LDH and MDA), inflammatory factors (TNF-α and IL-6) and apoptosis-related proteins (caspase-3, Bax, Bcl-2) expression were measured. The apoptotic cells were observed by TUNEL staining. Our study results indicated that non-cytotoxic levels of SDAPR significantly increased viability rate (LD 50 value of cisplatin is 20 μM), which improved antioxidant defence, attenuated apoptosis by decreasing expression levels of cleaved-caspase-3 and Bax, increasing Bcl-2 expression and inhibiting apoptotic positive cells in HEK 293 cells. In addition, SDAPR treatment markedly inhibited the levels of TNF-α and IL-6. In conclusion, Sika deer antler protein, a potential modulator, could alleviate cisplatin-induced cytotoxicity in HEK 293 cells.

Human Alpha Defensin 5 Expression in the Human Kidney and Urinary Tract

PubMed Central

Porter, Edith; Bevins, Charles L.; DiRosario, Julianne; Becknell, Brian; Wang, Huanyu

2012-01-01

Background The mechanisms that maintain sterility in the urinary tract are incompletely understood. Recent studies have implicated the importance of antimicrobial peptides (AMP) in protecting the urinary tract from infection. Here, we characterize the expression and relevance of the AMP human alpha-defensin 5 (HD5) in the human kidney and urinary tract in normal and infected subjects. Methodology/Principal Findings Using RNA isolated from human kidney, ureter, and bladder tissue, we performed quantitative real-time PCR to show that DEFA5, the gene encoding HD5, is constitutively expressed throughout the urinary tract. With pyelonephritis, DEFA5 expression significantly increased in the kidney. Using immunoblot analysis, HD5 production also increased with pyelonephritis. Immunostaining localized HD5 to the urothelium of the bladder and ureter. In the kidney, HD5 was primarily produced in the distal nephron and collecting tubules. Using immunoblot and ELISA assays, HD5 was not routinely detected in non-infected human urine samples while mean urinary HD5 production increased with E.coli urinary tract infection. Conclusions/Significance DEFA5 is expressed throughout the urinary tract in non-infected subjects. Specifically, HD5 is expressed throughout the urothelium of the lower urinary tract and in the collecting tubules of the kidney. With infection, HD5 expression increases in the kidney and levels become detectable in the urine. To our knowledge, our findings represent the first to quantitate HD5 expression and production in the human kidney. Moreover, this is the first report to detect the presence of HD5 in infected urine samples. Our results suggest that HD5 may have an important role in maintaining urinary tract sterility. PMID:22359618

Loss of Hfe Leads to Progression of Tumor Phenotype in Primary Retinal Pigment Epithelial Cells

PubMed Central

Gnana-Prakasam, Jaya P.; Veeranan-Karmegam, Rajalakshmi; Coothankandaswamy, Veena; Reddy, Sushma K.; Martin, Pamela M.; Thangaraju, Muthusamy; Smith, Sylvia B.; Ganapathy, Vadivel

2013-01-01

Purpose. Hemochromatosis is a disorder of iron overload arising mostly from mutations in HFE. HFE is expressed in retinal pigment epithelium (RPE), and Hfe−/− mice develop age-related iron accumulation and retinal degeneration associated with RPE hyperproliferation. Here, the mechanism underlying the hyperproliferative phenotype in RPE was investigated. Methods. Cellular senescence was monitored by β-galactosidase activity. Gene expression was monitored by real-time PCR. Survivin was analyzed by Western blot and immunofluorescence. Migration and invasion were monitored using appropriate kits. Glucose transporters (GLUTs) were monitored by 3-O-methyl-D-glucose uptake. Histone deacetylases (HDACs) were studied by monitoring catalytic activity and acetylation status of histones H3/H4. Results. Hfe−/− RPE cells exhibited slower senescence rate and higher survivin expression than wild type cells. Hfe−/− cells migrated faster and showed greater glucose uptake and increased expression of GLUTs. The expression of HDACs and DNA methyltransferase (DNMTs) also was increased. Similarly, RPE cells from hemojuvelin (Hjv)-knockout mice, another model of hemochromatosis, also had increased expression of GLUTs, HDACs, and DNMTs. The expression of Slc5a8 was decreased in Hfe−/− RPE cells, but treatment with a DNA methylation inhibitor restored the transporter expression, indicating involvement of DNA methylation in the silencing of Slc5a8 in Hfe−/− cells. Conclusions. RPE cells from iron-overloaded mice exhibit several features of tumor cells: decreased senescence, enhanced migration, increased glucose uptake, and elevated levels of HDACs and DNMTs. These features are seen in Hfe−/− RPE cells as well as in Hjv−/− RPE cells, providing a molecular basis for the hyperproliferative phenotype of Hfe−/− and Hjv−/− RPE cells. PMID:23169885

Effect of KCNJ5 Mutations on Gene Expression in Aldosterone-Producing Adenomas and Adrenocortical Cells

PubMed Central

Monticone, Silvia; Hattangady, Namita G.; Nishimoto, Koshiro; Mantero, Franco; Rubin, Beatrice; Cicala, Maria Verena; Pezzani, Raffaele; Auchus, Richard J.; Ghayee, Hans K.; Shibata, Hirotaka; Kurihara, Isao; Williams, Tracy A.; Giri, Judith G.; Bollag, Roni J.; Edwards, Michael A.; Isales, Carlos M.

2012-01-01

Context: Primary aldosteronism is a heterogeneous disease that includes both sporadic and familial forms. A point mutation in the KCNJ5 gene is responsible for familial hyperaldosteronism type III. Somatic mutations in KCNJ5 also occur in sporadic aldosterone producing adenomas (APA). Objective: The objective of the study was to define the effect of the KCNJ5 mutations on gene expression and aldosterone production using APA tissue and human adrenocortical cells. Methods: A microarray analysis was used to compare the transcriptome profiles of female-derived APA samples with and without KCNJ5 mutations and HAC15 adrenal cells overexpressing either mutated or wild-type KCNJ5. Real-time PCR validated a set of differentially expressed genes. Immunohistochemical staining localized the KCNJ5 expression in normal adrenals and APA. Results: We report a 38% (18 of 47) prevalence of KCNJ5 mutations in APA. KCNJ5 immunostaining was highest in the zona glomerulosa of NA and heterogeneous in APA tissue, and KCNJ5 mRNA was 4-fold higher in APA compared with normal adrenals (P < 0.05). APA with and without KCNJ5 mutations displayed slightly different gene expression patterns, notably the aldosterone synthase gene (CYP11B2) was more highly expressed in APA with KCNJ5 mutations. Overexpression of KCNJ5 mutations in HAC15 increased aldosterone production and altered expression of 36 genes by greater than 2.5-fold (P < 0.05). Real-time PCR confirmed increases in CYP11B2 and its transcriptional regulator, NR4A2. Conclusions: KCNJ5 mutations are prevalent in APA, and our data suggest that these mutations increase expression of CYP11B2 and NR4A2, thus increasing aldosterone production. PMID:22628608

Differential expression of glucose-metabolizing enzymes in multiple sclerosis lesions.

PubMed

Nijland, Philip G; Molenaar, Remco J; van der Pol, Susanne M A; van der Valk, Paul; van Noorden, Cornelis J F; de Vries, Helga E; van Horssen, Jack

2015-12-04

Demyelinated axons in multiple sclerosis (MS) lesions have an increased energy demand in order to maintain conduction. However, oxidative stress-induced mitochondrial dysfunction likely alters glucose metabolism and consequently impairs neuronal function in MS. Imaging and pathological studies indicate that glucose metabolism is altered in MS, although the underlying mechanisms and its role in neurodegeneration remain elusive. We investigated expression patterns of key enzymes involved in glycolysis, tricarboxylic acid (TCA) cycle and lactate metabolism in well-characterized MS tissue to establish which regulators of glucose metabolism are involved in MS and to identify underlying mechanisms. Expression levels of glycolytic enzymes were increased in active and inactive MS lesions, whereas expression levels of enzymes involved in the TCA cycle were upregulated in active MS lesions, but not in inactive MS lesions. We observed reduced expression and production capacity of mitochondrial α-ketoglutarate dehydrogenase (αKGDH) in demyelinated axons, which correlated with signs of axonal dysfunction. In inactive lesions, increased expression of lactate-producing enzymes was observed in astrocytes, whereas lactate-catabolising enzymes were mainly detected in axons. Our results demonstrate that the expression of various enzymes involved in glucose metabolism is increased in both astrocytes and axons in active MS lesions. In inactive MS lesions, we provide evidence that astrocytes undergo a glycolytic shift resulting in enhanced astrocyte-axon lactate shuttling, which may be pivotal for the survival of demyelinated axons. In conclusion, we show that key enzymes involved in energy metabolism are differentially expressed in active and inactive MS lesions. Our findings imply that, in addition to reduced oxidative phosphorylation activity, other bioenergetic pathways are affected as well, which may contribute to ongoing axonal degeneration in MS.

Ste20-like kinase, SLK, activates the heat shock factor 1 - Hsp70 pathway.

PubMed

Cybulsky, Andrey V; Guillemette, Julie; Papillon, Joan

2016-09-01

Expression and activation of SLK increases during renal ischemia-reperfusion injury. When highly expressed, SLK signals via c-Jun N-terminal kinase and p38 to induce apoptosis, and it exacerbates apoptosis induced by ischemia-reperfusion injury. Overexpression of SLK in glomerular epithelial cells (GECs)/podocytes in vivo induces injury and proteinuria. In response to various stresses, cells enhance expression of chaperones or heat shock proteins (e.g. Hsp70), which are involved in the folding and maturation of newly synthesized proteins, and can refold denatured or misfolded proteins. We address the interaction of SLK with the heat shock factor 1 (HSF1)-Hsp70 pathway. Increased expression of SLK in GECs (following transfection) induced HSF1 transcriptional activity. Moreover, HSF1 transcriptional activity was increased by in vitro ischemia-reperfusion injury (chemical anoxia/recovery) and heat shock, and in both instances was amplified further by SLK overexpression. HSF1 binds to promoters of target genes, such as Hsp70 and induces their transcription. By analogy to HSF1, SLK stimulated Hsp70 expression. Hsp70 was also enhanced by anoxia/recovery and was further amplified by SLK overexpression. Induction of HSF1 and Hsp70 was dependent on the kinase activity of SLK, and was mediated via polo-like kinase-1. Transfection of constitutively active HSF1 enhanced Hsp70 expression and inhibited SLK-induced apoptosis. Conversely, the proapoptotic action of SLK was augmented by HSF1 shRNA, or the Hsp70 inhibitor, pifithrin-μ. In conclusion, increased expression/activity of SLK activates the HSF1-Hsp70 pathway. Hsp70 attenuates the primary proapoptotic effect of SLK. Modulation of chaperone expression may potentially be harnessed as cytoprotective therapy in renal cell injury. Copyright © 2016 Elsevier B.V. All rights reserved.

Epigenetic regulation of matrix metalloproteinase expression in ameloblastoma

PubMed Central

2012-01-01

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

Glut-1 as a prognostic biomarker in oral squamous cell carcinoma

PubMed Central

Harshani, Jyotsna M; Yeluri, Sivaranjani; Guttikonda, Venkateswara Rao

2014-01-01

Introduction: Glut-1 is a glucose transporter protein, the expression of which is upregulated in malignant cells which show increased glucose uptake. Alterations in expression of Glut-1 have been reported in several pre-malignant and malignant lesions. The objectives of the present study were to compare the expression of Glut-1 in normal persons and in patients with oral squamous cell carcinoma (OSCC), to correlate the expression of Glut-1 with respect to clinical staging of OSCC and to evaluate the expression of Glut-1 with respect to different histopathological grades of OSCC. Materials and Methods: Thirty cases of OSCC were staged clinically and graded histopathologically. Immunohistochemical method was used to detect the expression of Glut-1 in OSCC and the same was compared with the normal subjects. The scores were compared using the chi-square test. Results: Glut-1 expression was detected in all grades of OSCC. A significant correlation with a P value of 0.00004 was found in immunostaining between normal and OSCC. The expression of Glut-1 was significant when compared with different clinical stages with significant P value of 0.0004 and in different histopathological grades of OSCC with a P value of 0.00001. Conclusion: Higher immunohistochemical staining scores were obtained with increased clinical staging and histopathological grades of OSCC. High expression of Glut-1 may be related to poor prognosis in OSCC. PMID:25948991

Molecular Mechanisms of Increased Heart Rate in Shenxianshengmai-treated Bradycardia Rabbits

PubMed Central

Liu, Zhou-Ying; Huang, Jian; Liu, Na-Na; Zheng, Min; Zhao, Tao; Zhao, Bu-Chang; Wang, Yi-Min; Pu, Jie-Lin

2017-01-01

Background: The molecular mechanisms of Shenxianshengmai (SXSM), a traditional Chinese medicine, on bradycardia have been incompletely understood. The study tried to investigate the gene expression profile and proteomics of bradycardia rabbits’ hearts after SXSM treatment. Methods: Twenty-four adult rabbits were randomly assigned in four groups: sham, model, model plus SXSM treatment, and sham plus SXSM treatment groups. Heart rate was recorded in all rabbits. Then, total RNA of atria and proteins of ventricle were isolated and quantified, respectively. Gene expression profiling was conducted by gene expression chip, and quantitative real-time reverse transcription-polymerase chain reaction (RT-PCR) was performed to confirm the results of gene expression chip. We used isobaric tags for elative and absolute quantitation and Western blotting to identify altered proteins after SXSM treatment. Results: There was a constant decrease in the mean heart rate (32%, from 238 ± 6 beats/min to 149 ± 12 beats/min) after six weeks in model compared with that in sham group. This effect was partially reversed by 4-week SXSM treatment. Complementary DNA microarray demonstrated that the increased acetylcholinesterase and reduced nicotinic receptor were take responsibility for the increased heart rate. In addition, proteins involved in calcium handling and signaling were affected by SXSM treatment. Real-time RT-PCR verified the results from gene chip. Results from proteomics demonstrated that SXSM enhanced oxidative phosphorylation and tricarboxylic acid (TCA) cycle in ventricular myocardium to improve ATP generation. Conclusions: Long-term SXSM stimulates sympathetic transmission by increasing the expression of acetylcholinesterase and reduces the expression of nicotinic receptor to increase heart rate. SXSM also restored the calcium handling genes and altered genes involved in signaling. In addition, SXSM improves the ATP supply of ventricular myocardium by increasing proteins involved in TCA cycle and oxidation-respiratory chain. PMID:28091410

Effect of patchouli alcohol on the regulation of heat shock-induced oxidative stress in IEC-6 cells.

PubMed

Liu, Xiaoxi; Jiang, Linshu; Liu, Fenghua; Chen, Yuping; Xu, Lei; Li, Deyin; Ma, Yunfei; Li, Huanrong; Xu, Jianqin

2016-08-01

Purpose Patchouli alcohol (PA) is used to treat gastrointestinal dysfunction. The purpose of this study was to ascertain the function of PA in the regulated process of oxidative stress in rat intestinal epithelial cells (IEC-6). Materials and methods Oxidative stress was stimulated by exposing IEC-6 cells to heat shock (42 °C for 3 h). IEC-6 cells in treatment groups were pretreated with various concentrations of PA (10, 40, and 80 ng/mL) for 3 h before heat shock. Results Heat shock caused damage to the morphology of IEC-6 cells, and increased reactive oxygen species (ROS) level and malondialdehyde (MDA) content. Moreover, mRNA and protein expression by target genes related to oxidative stress in heat shock were also altered. Specifically, the mRNA expression by HSP70, HSP90, GSH-px, NRF2 nd HO-1were all increased, and Nrf2 and Keap1 protein expression were increased after heat shock. However, pretreatment with PA weakened the level of damage to the cellular morphology, and decreased the MDA content caused by heat shock, indicating PA had cytoprotective activities. Pretreatment with PA at high dose significantly increased generation of intracellular ROS. Compared with the heat shock group alone, PA pretreatment significantly decreased the mRNA expression by HSP70, HSP90, SOD, CAT, GSH-px, KEAP1 and HO-1. Furthermore, the high dose of PA significantly increased Nrf2 protein expression, while both the intermediate and high dose of PA significantly increased HO-1 protein expression. Conclusion Heat-shock-induced oxidative stress in IEC-6 cells, and PA could alleviate the Nrf2-Keap1 cellular oxidative stress responses.

Protein disulfide isomerase-mediated apoptosis and proliferation of vascular smooth muscle cells induced by mechanical stress and advanced glycosylation end products result in diabetic mouse vein graft atherosclerosis.

PubMed

Ping, Suning; Liu, Shuying; Zhou, Yuhuan; Li, Ziqing; Li, Yuhuang; Liu, Kefeng; Bardeesi, Adham Sa; Wang, Linli; Chen, Jingbo; Deng, Lie; Wang, Jingjing; Wang, Hong; Chen, Dadi; Zhang, Zhengyu; Sheng, Puyi; Li, Chaohong

2017-05-25

Protein disulfide isomerase (PDI) involves cell survival and death. Whether PDI mediates mechanical stretch stress (SS) and/or advanced glycosylation end products (AGEs) -triggered simultaneous increases in proliferation and apoptosis of vascular smooth muscle cells (VSMCs) is unknown. Here, we hypothesized that different expression levels of PDI trigger completely opposite cell fates among the different VSMC subtypes. Mouse veins were grafted into carotid arteries of non-diabetic and diabetic mice for 8 weeks; the grafted veins underwent simultaneous increases in proliferation and apoptosis, which triggered vein graft arterializations in non-diabetic or atherosclerosis in diabetic mice. A higher rate of proliferation and apoptosis was seen in the diabetic group. SS and/or AGEs stimulated the quiescent cultured VSMCs, resulting in simultaneous increases in proliferation and apoptosis; they could induce increased PDI activation and expression. Both in vivo and in vitro, the proliferating VSMCs indicated weak co-expression of PDI and SM-α-actin while apoptotic or dead cells showed strong co-expression of both. Either SS or AGEs rapidly upregulated the expression of PDI, NOX1 and ROS, and their combination had synergistic effects. Inhibiting PDI simultaneously suppressed the proliferation and apoptosis of VSMCs, while inhibition of SM-α-actin with cytochalasin D led to increased apoptosis and cleaved caspases-3 but had no effect on proliferation. In conclusion, different expression levels of PDI in VSMCs induced by SS and/or AGEs triggered a simultaneous increase in proliferation and apoptosis, accelerated vein graft arterializations or atherosclerosis, leading us to propose PDI as a novel target for the treatment of vascular remodeling and diseases.

Protein disulfide isomerase-mediated apoptosis and proliferation of vascular smooth muscle cells induced by mechanical stress and advanced glycosylation end products result in diabetic mouse vein graft atherosclerosis

PubMed Central

Ping, Suning; Liu, Shuying; Zhou, Yuhuan; Li, Ziqing; Li, Yuhuang; Liu, Kefeng; Bardeesi, Adham SA; Wang, Linli; Chen, Jingbo; Deng, Lie; Wang, Jingjing; Wang, Hong; Chen, Dadi; Zhang, Zhengyu; Sheng, Puyi; Li, Chaohong

2017-01-01

Protein disulfide isomerase (PDI) involves cell survival and death. Whether PDI mediates mechanical stretch stress (SS) and/or advanced glycosylation end products (AGEs) -triggered simultaneous increases in proliferation and apoptosis of vascular smooth muscle cells (VSMCs) is unknown. Here, we hypothesized that different expression levels of PDI trigger completely opposite cell fates among the different VSMC subtypes. Mouse veins were grafted into carotid arteries of non-diabetic and diabetic mice for 8 weeks; the grafted veins underwent simultaneous increases in proliferation and apoptosis, which triggered vein graft arterializations in non-diabetic or atherosclerosis in diabetic mice. A higher rate of proliferation and apoptosis was seen in the diabetic group. SS and/or AGEs stimulated the quiescent cultured VSMCs, resulting in simultaneous increases in proliferation and apoptosis; they could induce increased PDI activation and expression. Both in vivo and in vitro, the proliferating VSMCs indicated weak co-expression of PDI and SM-α-actin while apoptotic or dead cells showed strong co-expression of both. Either SS or AGEs rapidly upregulated the expression of PDI, NOX1 and ROS, and their combination had synergistic effects. Inhibiting PDI simultaneously suppressed the proliferation and apoptosis of VSMCs, while inhibition of SM-α-actin with cytochalasin D led to increased apoptosis and cleaved caspases-3 but had no effect on proliferation. In conclusion, different expression levels of PDI in VSMCs induced by SS and/or AGEs triggered a simultaneous increase in proliferation and apoptosis, accelerated vein graft arterializations or atherosclerosis, leading us to propose PDI as a novel target for the treatment of vascular remodeling and diseases. PMID:28542133

Preventive effect of Ibrolipim on suppressing lipid accumulation and increasing lipoprotein lipase in the kidneys of diet-induced diabetic minipigs

PubMed Central

2011-01-01

Background The role of renal lipoprotein lipase (LPL) per se in kidney diseases is still controversial and obscure. The purpose of this study was to observe the preventive effects of Ibrolipim, a LPL activator, on lipid accumulation and LPL expression in the kidneys of minipigs fed a high-sucrose and high-fat diet (HSFD). Methods Male Chinese Bama minipigs were fed a control diet or HSFD with or without 0.1 g/kg/day Ibrolipim for 5 months. Body weight, plasma glucose, insulin, lipids, LPL activity, and urinary microalbumin were measured. Renal tissue was obtained for detecting LPL activity and contents of triglyceride and cholesterol, observing the renal lipid accumulation by Oil Red O staining, and examining the mRNA and protein expression of LPL by real time PCR, Western Blot and immunohistochemistry. Results Feeding HSFD to minipigs caused weight gain, hyperglycemia, hyperinsulinemia, hyperlipidemia and microalbuminuria. HSFD increased plasma LPL activity while it decreased the mRNA and protein expression and activity of LPL in the kidney. The increases in renal triglyceride and cholesterol contents were associated with the decrease in renal LPL activity of HSFD-fed minipigs. In contrast, supplementing Ibrolipim into HSFD lowered body weight, plasma glucose, insulin, triglyceride and urinary albumin concentrations while it increased plasma total cholesterol and HDL-C. Ibrolipim suppressed the renal accumulation of triglyceride and cholesterol, and stimulated the diet-induced down-regulation of LPL expression and activity in the kidney. Conclusions Ibrolipim exerts renoprotective and hypolipidemic effects via the increase in renal LPL activity and expression, and thus the increased expression and activity of renal LPL play a vital role in suppressing renal lipid accumulation and ameliorating proteinuria in diet-induced diabetic minipigs. PMID:21762526

Changes in the Expression and Distribution of Claudins, Increased Epithelial Apoptosis, and a Mannan-Binding Lectin-Associated Immune Response Lead to Barrier Dysfunction in Dextran Sodium Sulfate-Induced Rat Colitis

PubMed Central

Yuan, Bosi; Zhou, Shuping; Lu, Youke; Liu, Jiong; Jin, Xinxin; Wan, Haijun; Wang, Fangyu

2015-01-01

Background/Aims This animal study aimed to define the underlying cellular mechanisms of intestinal barrier dysfunction. Methods Rats were fed 4% with dextran sodium sulfate (DSS) to induce experimental colitis. We analyzed the sugars in 24-hour urine output by high pressure liquid chromatography. The expression of claudins, mannan-binding lectin (MBL), and MBL-associated serine proteases 2 (MASP-2) were detected in the colonic mucosa by immunohistochemistry; and apoptotic cells in the colonic epithelium were detected by the terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick-end labeling method assay. Results The lactulose and sucralose excretion levels in the urine of rats with DSS-induced colitis were significantly higher than those in the control rats. Mannitol excretion was lower and lactulose/mannitol ratios and sucralose/mannitol ratios were significantly increased compared with those in the control group (p<0.05). Compared with the controls, the expression of sealing claudins (claudin 3, claudin 5, and claudin 8) was significantly decreased, but that of claudin 1 was increased. The expression of pore-forming claudin 2 was upregulated and claudin 7 was downregulated in DSS-induced colitis. The epithelial apoptotic ratio was 2.8%±1.2% in controls and was significantly increased to 7.2%±1.2% in DSS-induced colitis. The expression of MBL and MASP-2 in the intestinal mucosa showed intense staining in controls, whereas there was weak staining in the rats with colitis. Conclusions There was increased intestinal permeability in DSS-induced colitis. Changes in the expression and distribution of claudins, increased epithelial apoptosis, and the MASP-2-induced immune response impaired the intestinal epithelium and contributed to high intestinal permeability. PMID:25717051

Protease-activated receptor 2 agonist increases cell proliferation and invasion of human pancreatic cancer cells

PubMed Central

XIE, LIQUN; DUAN, ZEXING; LIU, CAIJU; ZHENG, YANMIN; ZHOU, JING

2015-01-01

The aim of this study was to determine the expression of protease-activated receptor 2 (PAR-2) in the human pancreatic cancer cell line SW1990, and to evaluate its effect on cell proliferation and invasion. The expression of PAR-2 protein and mRNA in SW1990 cells was determined by immunocytochemistry and reverse transcription polymerase chain reaction (PCR), respectively. MTT and cell invasion and migration assays, as well as semi-quantitative PCR and zymography analysis, were additionally performed. PAR-2 mRNA was significantly upregulated in the cells treated with trypsin or the PAR-2 activating peptide Ser-Leu-Ile-Gly-Lys-Val (SLIGKV) (P<0.01), but not in the Val-Lys-Gly-Ile-Leu-Ser group (P>0.05). Trypsin and SLIGKV significantly promoted SW1990 cell proliferation in a dose- and time-dependent manner (P<0.05). Compared with the control group, trypsin and SLIGKV significantly increased the mRNA expression (P<0.01) and gelatinolytic activity (P<0.01) of matrix metalloproteinase (MMP)-2. In conclusion, PAR-2 is expressed in SW1990 cells. PAR-2 activation may promote the invasion and migration of human pancreatic cancer cells by increasing MMP-2 expression. PMID:25452809

Effects of extracellular modulation through hypoxia on the glucose metabolism of human breast cancer stem cells

NASA Astrophysics Data System (ADS)

Yustisia, I.; Jusman, S. W. A.; Wanandi, S. I.

2017-08-01

Cancer stem cells have been reported to maintain stemness under certain extracellular changes. This study aimed to analyze the effect of extracellular O2 level modulation on the glucose metabolism of human CD24-/CD44+ breast cancer stem cells (BCSCs). The primary BCSCs (CD24-/CD44+ cells) were cultured under hypoxia (1% O2) for 0.5, 4, 6, 24 and 48 hours. After each incubation period, HIF1α, GLUT1 and CA9 expressions, as well as glucose metabolism status, including glucose consumption, lactate production, O2 consumption and extracellular pH (pHe) were analyzed using qRT-PCR, colorimetry, fluorometry, and enzymatic reactions, respectively. Hypoxia caused an increase in HIF1α mRNA expressions and protein levels and shifted the metabolic states to anaerobic glycolysis, as demonstrated by increased glucose consumption and lactate production, as well as decreased O2 consumption and pHe. Furthermore, we demonstrated that GLUT1 and CA9 mRNA expressions simultaneously increased, in line with HIF1α expression. In conclusion, modulation of the extracellular environment of human BCSCs through hypoxia shifedt the metabolic state of BCSCs to anaerobic glycolysis, which might be associated with GLUT1 and CA9 expressions regulated by HIFlα transcription factor.

Vitamin D increases programmed death receptor-1 expression in Crohn’s disease

PubMed Central

Bendix, Mia; Greisen, Stinne; Dige, Anders; Hvas, Christian L.; Bak, Nina; Jørgensen, Søren P.; Dahlerup, Jens F.; Deleuran, Bent; Agnholt, Jørgen

2017-01-01

Background: Vitamin D modulates inflammation in Crohns disease (CD). Programmed death (PD)-1 receptor contributes to the maintenance of immune tolerance. Vitamin D might modulate PD-1 signalling in CD. Aim: To investigate PD-1 expression on T cell subsets in CD patients treated with vitamin D or placebo. Methods: We included 40 CD patients who received 1200 IU vitamin D3 for 26 weeks or placebo and eight healthy controls. Peripheral blood mononuclear cells (PBMCs) and plasma were isolated at baseline and week 26. The expressions of PD-1, PD-L1, and surface activation markers were analysed by flow cytometry. Soluble PD-1 plasma levels were measured by ELISA. Results: PD-1 expression upon T cell stimulation was increased in CD4+CD25+int T cells in vitamin D treated CD patients from 19% (range 10 39%) to 29% (11 79%)(p = 0.03) compared with placebo-treated patients. Vitamin D treatment, but not placebo, decreased the expression of the T cell activation marker CD69 from 42% (31 62%) to 33% (19 - 54%)(p = 0.01). Soluble PD-1 levels were not influenced by vitamin D treatment. Conclusions: Vitamin D treatment increases CD4+CD25+int T cells ability to up-regulate PD-1 in response to activation and reduces the CD69 expression in CD patients. PMID:28412753

Cytokine expression in mice exposed to diesel exhaust particles by inhalation. Role of tumor necrosis factor

PubMed Central

Saber, Anne T; Jacobsen, Nicklas R; Bornholdt, Jette; Kjær, Sanna L; Dybdahl, Marianne; Risom, Lotte; Loft, Steffen; Vogel, Ulla; Wallin, Håkan

2006-01-01

Background Particulate air pollution has been associated with lung and cardiovascular disease, for which lung inflammation may be a driving mechanism. The pro-inflammatory cytokine, tumor necrosis factor (TNF) has been suggested to have a key-role in particle-induced inflammation. We studied the time course of gene expression of inflammatory markers in the lungs of wild type mice and Tnf-/- mice after exposure to diesel exhaust particles (DEPs). Mice were exposed to either a single or multiple doses of DEP by inhalation. We measured the mRNA level of the cytokines Tnf and interleukin-6 (Il-6) and the chemokines, monocyte chemoattractant protein (Mcp-1), macrophage inflammatory protein-2 (Mip-2) and keratinocyte derived chemokine (Kc) in the lung tissue at different time points after exposure. Results Tnf mRNA expression levels increased late after DEP-inhalation, whereas the expression levels of Il-6, Mcp-1 and Kc increased early. The expression of Mip-2 was independent of TNF if the dose was above a certain level. The expression levels of the cytokines Kc, Mcp-1 and Il-6, were increased in the absence of TNF. Conclusion Our data demonstrate that Tnf is not important in early DEP induced inflammation and rather exerts negative influence on Mcp-1 and Kc mRNA levels. This suggests that other signalling pathways are important, a candidate being one involving Mcp-1. PMID:16504008

Decrease of PECAM-1-gene-expression induced by proinflammatory cytokines IFN-γ and IFN-α is reversed by TGF-β in sinusoidal endothelial cells and hepatic mononuclear phagocytes

PubMed Central

Neubauer, Katrin; Lindhorst, Alexander; Tron, Kyrylo; Ramadori, Giuliano; Saile, Bernhard

2008-01-01

Background and aim The mechanisms of transmigration of inflammatory cells through the sinusoids are still poorly understood. This study aims to identify in vitro conditions (cytokine treatment) which may allow a better understanding of the changes in PECAM (platelet endothelial cell adhesion molecule)-1-gene-expression observed in vivo. Methods and results In this study we show by immunohistochemistry, that there is an accumulation of ICAM-1 (intercellular cell adhesion molecule-1) and ED1 positive cells in necrotic areas of livers of CCl4-treated rats, whereas there are few PECAM-1 positive cells observable. After the administration of CCl4, we could detect an early rise of levels of IFN-γ followed by an enhanced TGF-β protein level. As shown by Northern blot analysis and surface protein expression analysed by flow cytometry, IFN-γ-treatment decreased PECAM-1-gene-expression in isolated SECs (sinusoidal endothelial cells) and mononuclear phagocytes (MNPs) in parallel with an increase in ICAM-1-gene-expression in a dose and time dependent manner. In contrast, TGF-β-treatment increased PECAM-1-expression. Additional administration of IFN-γ to CCl4-treated rats and observations in IFN-γ-/- mice confirmed the effect of IFN-γ on PECAM-1 and ICAM-1-expression observed in vitro and increased the number of ED1-expressing cells 12 h after administration of the toxin. Conclusion The early decrease of PECAM-1-expression and the parallel increase of ICAM-1-expression following CCl4-treatment is induced by elevated levels of IFN-γ in livers and may facilitate adhesion and transmigration of inflammatory cells. The up-regulation of PECAM-1-expression in SECs and MNPs after TGF-β-treatment suggests the involvement of PECAM-1 during the recovery after liver damage. PMID:18466611

Interleukin 31 mediates MAP kinase and STAT1/3 activation in intestinal epithelial cells and its expression is upregulated in inflammatory bowel disease

PubMed Central

Dambacher, Julia; Beigel, Florian; Seiderer, Julia; Haller, Dirk; Göke, Burkhard; Auernhammer, Christoph J; Brand, Stephan

2007-01-01

Background/aim Interleukin 31 (IL31), primarily expressed in activated lymphocytes, signals through a heterodimeric receptor complex consisting of the IL31 receptor alpha (IL31Rα) and the oncostatin M receptor (OSMR). The aim of this study was to analyse IL31 receptor expression, signal transduction, and specific biological functions of this cytokine system in intestinal inflammation. Methods Expression studies were performed by RT‐PCR, quantitative PCR, western blotting, and immunohistochemistry. Signal transduction was analysed by western blotting. Cell proliferation was measured by MTS assays, cell migration by restitution assays. Results Colorectal cancer derived intestinal epithelial cell (IEC) lines express both IL31 receptor subunits, while their expression in unstimulated primary murine IEC was low. LPS and the proinflammatory cytokines TNF‐α, IL1β, IFN‐γ, and sodium butyrate stimulation increased IL31, IL31Rα, and OSMR mRNA expression, while IL31 itself enhanced IL8 expression in IEC. IL31 mediates ERK‐1/2, Akt, STAT1, and STAT3 activation in IEC resulting in enhanced IEC migration. However, at low cell density, IL31 had significant antiproliferative capacities (p<0.005). IL31 mRNA expression was not increased in the TNFΔARE mouse model of ileitis but in inflamed colonic lesions compared to non‐inflamed tissue in patients with Crohn's disease (CD; average 2.4‐fold increase) and in patients with ulcerative colitis (UC; average 2.6‐fold increase) and correlated with the IL‐8 expression in these lesions (r = 0.564 for CD; r = 0.650 for UC; total number of biopsies analysed: n = 88). Conclusion IEC express the functional IL31 receptor complex. IL31 modulates cell proliferation and migration suggesting a role in the regulation of intestinal barrier function particularly in intestinal inflammation. PMID:17449633

UV-induced inhibition of adipokine production in subcutaneous fat aggravates dermal matrix degradation in human skin.

PubMed

Kim, Eun Ju; Kim, Yeon Kyung; Kim, Min-Kyoung; Kim, Sungsoo; Kim, Jin Yong; Lee, Dong Hun; Chung, Jin Ho

2016-05-10

Ultraviolet (UV) exposure to the human skin reduces triglycerides contents and lipid synthesis in the subcutaneous (SC) fat. Because adiponectin and leptin are the most abundant adipokines from the SC fat, we aim to investigate how they interact with UV exposure and skin aging. The expressions of adiponectin and leptin were significantly decreased in SC fat of sun-exposed forearm skin, in comparison with that of sun-protected buttock skin of the same elderly individuals, indicating that chronic UV exposure decreases both adipokines. Acute UV irradiation also decreased the expressions of adiponectin and leptin in SC fat. The expressions of adiponectin receptor 1/2 and leptin receptor were significantly decreased in the dermis as well as in SC fat. Moreover, while exogenous adiponectin and leptin administration prevented UV- and TNF-α induced matrix metalloproteinase (MMP)-1 expression, they also increased UV- and TNF-α induced reduction of type 1 procollagen production. Silencing of adiponectin, leptin or their receptors led to an increased MMP-1 and a decreased type 1 procollagen expression, which was reversed by treatment with recombinant human adiponectin or leptin. In conclusion, UV exposure decreases the expression of adiponectin and leptin, leading to the exacerbation of photoaging by stimulating MMP-1 expression and inhibiting procollagen synthesis.

Prognostic Indications of Elevated MCT4 and CD147 across Cancer Types: A Meta-Analysis

PubMed Central

Bovenzi, Cory D.; Hamilton, James; Tassone, Patrick; Johnson, Jennifer; Cognetti, David M.; Luginbuhl, Adam; Keane, William M.; Zhan, Tingting; Tuluc, Madalina; Bar-Ad, Voichita; Martinez-Outschoorn, Ubaldo; Curry, Joseph M.

2015-01-01

Background. Metabolism in the tumor microenvironment can play a critical role in tumorigenesis and tumor aggression. Metabolic coupling may occur between tumor compartments; this phenomenon can be prognostically significant and may be conserved across tumor types. Monocarboxylate transporters (MCTs) play an integral role in cellular metabolism via lactate transport and have been implicated in metabolic synergy in tumors. The transporters MCT1 and MCT4 are regulated via expression of their chaperone, CD147. Methods. We conducted a meta-analysis of existing publications on the relationship between MCT1, MCT4, and CD147 expression and overall survival and disease-free survival in cancer, using hazard ratios derived via multivariate Cox regression analyses. Results. Increased MCT4 expressions in the tumor microenvironment, cancer cells, or stromal cells were all associated with decreased overall survival and decreased disease-free survival (p < 0.001 for all analyses). Increased CD147 expression in cancer cells was associated with decreased overall survival and disease-free survival (p < 0.0001 for both analyses). Few studies were available on MCT1 expression; MCT1 expression was not clearly associated with overall or disease-free survival. Conclusion. MCT4 and CD147 expression correlate with worse prognosis across many cancer types. These results warrant further investigation of these associations. PMID:26779534

Cilostazol Upregulates Autophagy via SIRT1 Activation: Reducing Amyloid-β Peptide and APP-CTFβ Levels in Neuronal Cells.

PubMed

Lee, Hye Rin; Shin, Hwa Kyoung; Park, So Youn; Kim, Hye Young; Bae, Sun Sik; Lee, Won Suk; Rhim, Byung Yong; Hong, Ki Whan; Kim, Chi Dae

2015-01-01

Autophagy is a vital pathway for the removal of β-amyloid peptide (Aβ) and the aggregated proteins that cause Alzheimer's disease (AD). We previously found that cilostazol induced SIRT1 expression and its activity in neuronal cells, and thus, we hypothesized that cilostazol might stimulate clearances of Aβ and C-terminal APP fragment β subunit (APP-CTFβ) by up-regulating autophagy.When N2a cells were exposed to soluble Aβ1-42, protein levels of beclin-1, autophagy-related protein5 (Atg5), and SIRT1 decreased significantly. Pretreatment with cilostazol (10-30 μM) or resveratrol (20 μM) prevented these Aβ1-42 evoked suppressions. LC3-II (a marker of mammalian autophagy) levels were significantly increased by cilostazol, and this increase was reduced by 3-methyladenine. To evoke endogenous Aβ overproduction, N2aSwe cells (N2a cells stably expressing human APP containing the Swedish mutation) were cultured in medium with or without tetracycline (Tet+ for 48 h and then placed in Tet- condition). Aβ and APP-CTFβ expressions were increased after 12~24 h in Tet- condition, and these increased expressions were significantly reduced by pretreating cilostazol. Cilostazol-induced reductions in the expressions of Aβ and APP-CTFβ were blocked by bafilomycin A1 (a blocker of autophagosome to lysosome fusion). After knockdown of the SIRT1 gene (to ~40% in SIRT1 protein), cilostazol failed to elevate the expressions of beclin-1, Atg5, and LC3-II, indicating that cilostazol increases these expressions by up-regulating SIRT1. Further, decreased cell viability induced by Aβ was prevented by cilostazol, and this inhibition was reversed by 3-methyladenine, indicating that the protective effect of cilostazol against Aβ induced neurotoxicity is, in part, ascribable to the induction of autophagy. In conclusion, cilostazol modulates autophagy by increasing the activation of SIRT1, and thereby enhances Aβ clearance and increases cell viability.

Precision-cut liver slices as a model for the early onset of liver fibrosis to test antifibrotic drugs

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

Westra, Inge M.; Oosterhuis, Dorenda; Groothuis, Geny M.M.

Induction of fibrosis during prolonged culture of precision-cut liver slices (PCLS) was reported. In this study, the use of rat PCLS was investigated to further characterize the mechanism of early onset of fibrosis in this model and the effects of antifibrotic compounds. Rat PCLS were incubated for 48 h, viability was assessed by ATP and gene expression of PDGF-B and TGF-β1 and the fibrosis markers Hsp47, αSma and Pcol1A1 and collagen1 protein expressions were determined. The effects of the antifibrotic drugs imatinib, sorafenib and sunitinib, PDGF-pathway inhibitors, and perindopril, valproic acid, rosmarinic acid, tetrandrine and pirfenidone, TGFβ-pathway inhibitors, were determined.more » After 48 h of incubation, viability of the PCLS was maintained and gene expression of PDGF-B was increased while TGF-β1 was not changed. Hsp47, αSma and Pcol1A1 gene expressions were significantly elevated in PCLS after 48 h, which was further increased by PDGF-BB and TGF-β1. The increased gene expression of fibrosis markers was inhibited by all three PDGF-inhibitors, while TGFβ-inhibitors showed marginal effects. The protein expression of collagen 1 was inhibited by imatinib, perindopril, tetrandrine and pirfenidone. In conclusion, the increased gene expression of PDGF-B and the down-regulation of fibrosis markers by PDGF-pathway inhibitors, together with the absence of elevated TGF-β1 gene expression and the limited effect of the TGFβ-pathway inhibitors, indicated the predominance of the PDGF pathway in the early onset of fibrosis in PCLS. PCLS appear a useful model for research of the early onset of fibrosis and for testing of antifibrotic drugs acting on the PDGF pathway. - Highlights: • During culture, fibrosis markers increased in precision-cut liver slices (PCLS). • Gene expression of PDGF-β was increased, while TGFβ was not changed in rat PCLS. • PDGF-pathway inhibitors down-regulated this increase of fibrosis markers. • TGFβ-pathway inhibitors had only minor effects on fibrosis markers. • Rat PCLS can be used to study the early onset of fibrosis.« less

Mechanical Force-induced TGFB1 Increases Expression of SOST/POSTN by hPDL Cells.

PubMed

Manokawinchoke, J; Limjeerajarus, N; Limjeerajarus, C; Sastravaha, P; Everts, V; Pavasant, P

2015-07-01

The aim of this study was to investigate the response of human periodontal ligament (hPDL) fibroblasts to an intermittent compressive force and its effect on the expression of SOST, POSTN, and TGFB1. A computerized cell compressive force loading apparatus was introduced, and hPDL cells were subjected to intermittent compressive force. The changes in messenger RNA (mRNA) and protein expression were monitored by real-time polymerase chain reaction and Western blot analysis, respectively. An increased expression of SOST, POSTN, and TGFB1 was observed in a time-dependent fashion. Addition of cycloheximide, a transforming growth factor (TGF)-β inhibitor (SB431542), or a neutralizing antibody against TGF-β1 attenuated the force-induced expression of SOST and POSTN as well as sclerostin and periostin, indicating a role of TGF-β1 in the pressure-induced expression of these proteins. Enzyme-linked immunosorbent assay analysis revealed an increased level of TGF-β1 in the cell extracts but not in the medium, suggesting that intermittent compressive force promoted the accumulation of TGF-β1 in the cells or their surrounding matrix. In conclusion, an intermittent compressive force regulates SOST/POSTN expression by hPDL cells via the TGF-β1 signaling pathway. Since these proteins play important roles in the homeostasis of the periodontal tissue, our results indicate the importance of masticatory forces in this process. © International & American Associations for Dental Research 2015.

Correlation of High-Risk Soft Tissue Sarcoma Biomarker Expression Patterns with Outcome following Neoadjuvant Chemoradiation

PubMed Central

Magliocco, Anthony; Zhang, Qiang; Wang, Dian; Klimowicz, Alex; Harris, Jonathan; Simko, Jeff; DeLaney, Thomas; Kraybill, William; Kirsch, David G.

2018-01-01

Background Sarcoma mortality remains high despite adjuvant chemotherapy. Biomarker predictors of treatment response and outcome could improve treatment selection. Methods Tissue microarrays (TMAs) were created using pre- and posttreatment tumor from two prospective trials (MGH pilot and RTOG 9514) of neoadjuvant/adjuvant MAID chemotherapy and preoperative radiation. Biomarkers were measured using automated computerized imaging (AQUA or ACIS). Expression was correlated with disease-free survival (DFS), distant disease-free survival (DDFS), and overall survival (OS). Results Specimens from 60 patients included 23 pretreatment (PRE), 40 posttreatment (POST), and 12 matched pairs (MPs). In the MP set, CAIX, GLUT1, and PARP1 expression significantly decreased following neoadjuvant therapy, but p53 nuclear/cytoplasmic (N/C) ratio increased. In the PRE set, no biomarker expression was associated with DFS, DDFS, or OS. In the POST set, increased p53 N/C ratio was associated with a significantly decreased DFS and DDFS (HR 4.13, p=0.017; HR 4.16, p=0.016), while increased ERCC1 and XPF expression were associated with an improved DFS and DDFS. No POST biomarkers were associated with OS. Conclusions PRE biomarker expression did not predict survival outcomes. Expression pattern changes after neoadjuvant chemoradiation supports the concepts of tumor reoxygenation, altered HIF-1α signaling, and a p53 nuclear accumulation DNA damage response. Clinical Trial Registration NRG Oncology RTOG 9514 is registered with ClinicalTrials.gov. The ClinicalTrials.gov Identifier is NCT00002791. PMID:29681762

Vascular endothelial growth factor C promotes cervical cancer cell invasiveness via regulation of microRNA-326/cortactin expression.

PubMed

Cheng, Yang; Jiang, Shuyi; Yuan, Jin; Liu, Junxiu; Simoncini, Tommaso

2018-04-16

Vascular endothelial growth factor C (VEGF-C) accelerates cervical cancer metastasis, while the detailed mechanism remains largely unknown. Recent evidence indicates that microRNA play a crucial role in controlling cancer cell invasiveness. In the present study, we investigated the role of miR-326 in VEGF-C-induced cervical cancer cell invasion. VEGF-C expression was higher and miR-326 was much lower in primary cervical cancer specimens than that in non-cancerous specimens, and a negative correlation between VEGF-C and miR-326 was found. On cervical carcinoma cell line SiHa cells, treatment with VEGF-C downregulated miR-326 level and increased cortactin protein expression. Transfection with miR-326 mimic reversed cortactin expression induced by VEGF-C, suggesting that VEGF-C increased cortactin via downregulation of miR-326. VEGF-C activated c-Src and c-Src inhibitor PP2 abolished VEGF-C effect on miR-326 and cortactin expression, implying that VEGF-C regulated miR-326/cortactin via c-Src signaling. VEGF-C promoted SiHa cell invasion index, which was largely inhibited by transfection with miR-326 antagonist or by siRNA against cortactin. In conclusion, our findings implied that VEGF-C reduced miR-326 expression and increased cortactin expression through c-Src signaling, leading to enhanced cervical cancer invasiveness. This may shed light on potential therapeutic strategies for cervical cancer therapy.

Suppression of MicroRNA let-7a Expression by Agmatine Regulates Neural Stem Cell Differentiation

PubMed Central

Song, Juhyun; Oh, Yumi; Kim, Jong Youl; Cho, Kyoung Joo

2016-01-01

Purpose Neural stem cells (NSCs) effectively reverse some severe central nervous system (CNS) disorders, due to their ability to differentiate into neurons. Agmatine, a biogenic amine, has cellular protective effects and contributes to cellular proliferation and differentiation in the CNS. Recent studies have elucidated the function of microRNA let-7a (let-7a) as a regulator of cell differentiation with roles in regulating genes associated with CNS neurogenesis. Materials and Methods This study aimed to investigate whether agmatine modulates the expression of crucial regulators of NSC differentiation including DCX, TLX, c-Myc, and ERK by controlling let-7a expression. Results Our data suggest that high levels of let-7a promoted the expression of TLX and c-Myc, as well as repressed DCX and ERK expression. In addition, agmatine attenuated expression of TLX and increased expression of ERK by negatively regulating let-7a. Conclusion Our study therefore enhances the present understanding of the therapeutic potential of NSCs in CNS disorders. PMID:27593875

Elevated mu-opioid receptor expression in the nucleus of the solitary tract accompanies attenuated withdrawal signs after chronic low dose naltrexone in opiate-dependent rats.

PubMed

Van Bockstaele, E J; Rudoy, C; Mannelli, P; Oropeza, V; Qian, Y

2006-02-15

We previously described a decrease in withdrawal behaviors in opiate-dependent rats that were chronically treated with very low doses of naltrexone in their drinking water. Attenuated expression of withdrawal behaviors correlated with decreased c-Fos expression and intracellular signal transduction elements [protein kinase A regulatory subunit II (PKA) and phosphorylated cAMP response element binding protein (pCREB)] in brainstem noradrenergic nuclei. In this study, to determine whether similar cellular changes occurred in forebrain nuclei associated with drug reward, expressions of PKA and pCREB were analyzed in the ventral tegmental area, frontal cortex, striatum, and amygdala of opiate-treated rats that received low doses of naltrexone in their drinking water. No significant difference in PKA or pCREB was detected in these regions following drug treatment. To examine further the cellular mechanisms in noradrenergic nuclei that could underlie attenuated withdrawal behaviors following low dose naltrexone administration, the nucleus of the solitary tract (NTS) and locus coeruleus (LC) were examined for opioid receptor (OR) protein expression. Results showed a significant increase in muOR expression in the NTS of morphine-dependent rats that received low doses of naltrexone in their drinking water, and increases in muOR expression were also found to be dose dependent. Protein expression of muOR in the LC and deltaOR in either brain region remained unchanged. In conclusion, our previously reported decreases in c-Fos and PKA expression in the NTS following pretreatment with low doses of naltrexone may be partially explained by a greater inhibition of NTS neurons resulting from increased muOR expression in this region.

Developmental expression of the receptor for advanced glycation end-products (RAGE) and its response to hyperoxia in the neonatal rat lung

PubMed Central

Lizotte, Pierre-Paul; Hanford, Lana E; Enghild, Jan J; Nozik-Grayck, Eva; Giles, Brenda-Louise; Oury, Tim D

2007-01-01

Background The receptor for advanced glycation end products (mRAGE) is associated with pathology in most tissues, while its soluble form (sRAGE) acts as a decoy receptor. The adult lung is unique in that it expresses high amounts of RAGE under normal conditions while other tissues express low amounts normally and up-regulate RAGE during pathologic processes. We sought to determine the regulation of the soluble and membrane isoforms of RAGE in the developing lung, and its expression under hyperoxic conditions in the neonatal lung. Results Fetal (E19), term, 4 day, 8 day and adult rat lung protein and mRNA were analyzed, as well as lungs from neonatal (0–24 hrs) 2 day and 8 day hyperoxic (95% O2) exposed animals. mRAGE transcripts in the adult rat lung were 23% greater than in neonatal (0–24 hrs) lungs. On the protein level, rat adult mRAGE expression was 2.2-fold higher relative to neonatal mRAGE expression, and adult sRAGE protein expression was 2-fold higher compared to neonatal sRAGE. Fetal, term, 4 day and 8 day old rats had a steady increase in both membrane and sRAGE protein expression evaluated by Western Blot and immunohistochemistry. Newborn rats exposed to chronic hyperoxia showed significantly decreased total RAGE expression compared to room air controls. Conclusion Taken together, these data show that rat pulmonary RAGE expression increases with age beginning from birth, and interestingly, this increase is counteracted under hyperoxic conditions. These results support the emerging concept that RAGE plays a novel and homeostatic role in lung physiology. PMID:17343756

KLF5 regulates infection- and inflammation-induced pro-labour mediators in human myometrium.

PubMed

Lappas, Martha

2015-05-01

The transcription factor Kruppel-like factor 5 (KLF5) has been shown to associate with nuclear factor kappa B (NFκB) to regulate genes involved in inflammation. However, there are no studies on the expression and regulation of KLF5 in the processes of human labour and delivery. Thus, the aims of this study were to determine the effect of i) human labour on KLF5 expression in both foetal membranes and myometrium; ii) the pro-inflammatory cytokine interleukin 1 beta (IL1β), bacterial product flagellin and the viral dsRNA analogue poly(I:C) on KLF5 expression and iii) KLF5 knockdown by siRNA in human myometrial primary cells on pro-inflammatory and pro-labour mediators. In foetal membranes, there was no effect of term or preterm labour on KLF5 expression. In myometrium, the term labour was associated with an increase in nuclear KLF5 protein expression. Moreover, KLF5 expression was also increased in myometrial cells treated with IL1β, flagellin or poly(IC), likely factors contributing to preterm birth. KLF5 silencing in myometrial cells significantly decreased IL1β-induced cytokine expression (IL6 and IL8 mRNA expression and release), COX2 mRNA expression, and subsequent release of prostaglandins PGE2 and PGF2 α. KLF5 silencing also significantly reduced flagellin- and poly(I:C)-induced IL6 and IL8 mRNA expression. Lastly, IL1β-, flagellin- and poly(I:C)-stimulated NFκB transcriptional activity was significantly suppressed in KLF5-knockout myometrial cells. In conclusion, this study describes novel data in which KLF5 is increased in labouring myometrium, and KLF5 silencing decreased inflammation- and infection-induced pro-labour mediators. © 2015 Society for Reproduction and Fertility.

Androgen receptor expression in the human vagina under different physiological and treatment conditions.

PubMed

Baldassarre, M; Perrone, A M; Giannone, F A; Armillotta, F; Battaglia, C; Costantino, A; Venturoli, S; Meriggiola, M C

2013-01-01

Recent data report an important role of testosterone (T) in modulating female sexual responses, but little is known about the expression and distribution of androgen receptor (AR) in the human vagina. Therefore, the aims of our study were to evaluate the expression of AR in the human vagina in premenopausal (PrM) and menopausal (M) women and in T-treated women. Vaginal biopsies were obtained from PrM and postmenopausal women and from women with gender identity disorder (female to male (FtM)) receiving exogenous T. AR gene and protein expression levels in vaginal tissues were determined by real-time PCR and western blot analysis, respectively, whereas the localization of AR in vaginal mucosa and stroma was performed by immunohistochemistry. ARs were detected by immunostaining both in the mucosa and stroma. In vaginal mucosa, AR density score decreases with age but does not change with T administration. In stromal tissue, AR density score does not change with age but significantly increases with T administration (P<0.01). AR protein expression was significantly increased in FtM subjects (P<0.001). The expression of AR messenger RNA (mRNA) evaluated by Real-time PCR showed a significantly higher mRNA expression in FtM versus M patients (P<0.01) and in PrM versus M subjects (P<0.05). In conclusion, we found AR protein and mRNA expression both in the epithelium and stroma of the human vagina in all groups of women. A negative correlation exists between age and AR expression in the vaginal mucosa. T administration increases AR expression in both the mucosa and stroma.

Intra-tubular hydrodynamic forces influence tubulo-interstitial fibrosis in the kidney

PubMed Central

Rohatgi, Rajeev; Flores, Daniel

2010-01-01

Purpose of review Renal epithelial cells respond to mechanical stimuli with immediate transduction events (e.g., activation of ion channels), intermediate biological responses (e.g., changes in gene expression), and long term cellular adaptation (e.g., protein expression). Progressive renal disease is characterized by disturbed glomerular hydrodynamics that contributes to glomerulosclerosis, but, how intra-tubular biomechanical forces contribute to tubulo-interstital inflammation and fibrosis is poorly understood. Recent findings In vivo and in vitro models of obstructive uropathy demonstrate that tubular stretch induces robust expression of transforming growth factor β-1 (TGFβ-1), activation of tubular apoptosis, and induction of NF-κB signaling which contribute to the inflammatory and fibrotic milieu. Non-obstructive structural kidney diseases associated with nephron loss follow a course characterized by compensatory increases of single nephron glomerular filtration rate and tubular flow rate. Resulting increases in tubular fluid shear stress (FSS) reduce tissue-plasminogen activator and urokinase enzymatic activity which diminishes breakdown of extracellular matrix. In models of high dietary Na intake, which increase tubular flow, urinary TGFβ-1 concentrations and renal mitogen activated protein kinase activity are increased. Summary In conclusion, intra-tubular biomechanical forces, stretch and FSS, generate changes in intracellular signaling and gene expression that contribute to the pathobiology of obstructive, and non-obstructive kidney disease. PMID:19851105

Rilmenidine improves hepatic steatosis through p38-dependent pathway to higher the expression of farnesoid X receptor.

PubMed

Yang, Po-Sheng; Wu, Hung-Tsung; Chung, Hsien-Hui; Chen, Chun-Ta; Chi, Chin-Wen; Yeh, Ching-Hua; Cheng, Juei-Tang

2012-01-01

The nuclear receptor farnesoid X receptor (FXR) regulates pathways in lipid, glucose, and energy metabolism. Activation of FXR in mice significantly improved high-fat diet-induced hepatic steatosis. It has been reported that activation of imidazoline I-1 receptor by rilmenidine increases the expression of FXR in human hepatoma cell line, Hep G2 cell, to regulate the target genes relating to lipid metabolism; activation of FXR by rilmenidine exerts an antihyperlipidemic action. However, signals for this action of rilmenidine are still unknown. In the present study, hepatic steatosis induced in mice by high-fat diet was improved by rilmenidine after intraperitoneal injection at 1 mg/kg daily for 12 weeks. Also, mediation of I-1 receptors was identified using the specific antagonist efaroxan. Moreover, rilmenidine decreased the oleic acid-induced lipid accumulation in Hep G2 cells. Otherwise, rilmenidine increased the phosphorylation of p38 to increase the expression of FXR. Deletion of calcium ions by BAPTA-AM reversed the rilmenidine-induced p38 phosphorylation. In conclusion, we suggest that rilmenidine activates I-1 receptor to increase intracellular calcium ions that may enhance the phosphorylation of p38 to higher the expression of FXR for improvement of hepatic steatosis in both animals and cells.

Estradiol induces endothelial cell migration and proliferation through estrogen receptor-enhanced RhoA/ROCK pathway.

PubMed

Oviedo, Pilar J; Sobrino, Agua; Laguna-Fernandez, Andrés; Novella, Susana; Tarín, Juan J; García-Pérez, Miguel-Angel; Sanchís, Juan; Cano, Antonio; Hermenegildo, Carlos

2011-03-30

Migration and proliferation of endothelial cells are involved in re-endothelialization and angiogenesis, two important cardiovascular processes that are increased in response to estrogens. RhoA, a small GTPase which controls multiple cellular processes, is involved in the control of cell migration and proliferation. Our aim was to study the role of RhoA on estradiol-induced migration and proliferation and its dependence on estrogen receptors activity. Human umbilical vein endothelial cells were stimulated with estradiol, in the presence or absence of ICI 182780 (estrogen receptors antagonist) and Y-27632 (Rho kinase inhibitor). Estradiol increased Rho GEF-1 gene expression and RhoA (gene and protein expression and activity) in an estrogen receptor-dependent manner. Cell migration, stress fiber formation and cell proliferation were increased in response to estradiol and were also dependent on the estrogen receptors and RhoA activation. Estradiol decreased p27 levels, and significantly raised the expression of cyclins and CDK. These effects were counteracted by the use of either ICI 182780 or Y-27632. In conclusion, estradiol enhances the RhoA/ROCK pathway and increases cell cycle-related protein expression by acting through estrogen receptors. This results in an enhanced migration and proliferation of endothelial cells. Copyright © 2010 Elsevier Ireland Ltd. All rights reserved.

Na+-glucose cotransporter SGLT1 protein in salivary glands: potential involvement in the diabetes-induced decrease in salivary flow.

PubMed

Sabino-Silva, R; Freitas, H S; Lamers, M L; Okamoto, M M; Santos, M F; Machado, U F

2009-03-01

Oral health complications in diabetes include decreased salivary secretion. The SLC5A1 gene encodes the Na(+)-glucose cotransporter SGLT1 protein, which not only transports glucose, but also acts as a water channel. Since SLC5A1 expression is altered in kidneys of diabetic subjects, we hypothesize that it could also be altered in salivary glands, contributing to diabetic dysfunction. The present study shows a diabetes-induced decrease (p < 0.001) in salivary secretion, which was accompanied by enhanced (p < 0.05) SGLT1 mRNA expression in parotid (50%) and submandibular (30%) glands. Immunohistochemical analysis of parotid gland of diabetic rats revealed that SGLT1 protein expression increased in the luminal membrane of ductal cells, which can stimulate water reabsorption from primary saliva. Furthermore, SGLT1 protein was reduced in myoepithelial cells of the parotid from diabetic animals, and that, by reducing cellular contractile activity, might also be related to reduced salivary flux. Six-day insulin-treated diabetic rats reversed all alterations. In conclusion, diabetes increases SLC5A1 gene expression in salivary glands, increasing the SGLT1 protein content in the luminal membrane of ductal cells, which, by increasing water reabsorption, might explain the diabetes-induced decrease in salivary secretion.

Uropathogenic E.coli (UPEC) Infection Induces Proliferation through Enhancer of Zeste Homologue 2 (EZH2)

PubMed Central

Penna, Frank; Samiei, Alaleh Najdi; Sidler, Martin; Jiang, Jia-Xin; Ibrahim, Fadi; Tolg, Cornelia; Delgado-Olguin, Paul; Rosenblum, Norman; Bägli, Darius J.

2016-01-01

Host-pathogen interactions can induce epigenetic changes in the host directly, as well as indirectly through secreted factors. Previously, uropathogenic Escherichia coli (UPEC) was shown to increase DNA methyltransferase activity and expression, which was associated with methylation-dependent alterations in the urothelial expression of CDKN2A. Here, we showed that paracrine factors from infected cells alter expression of another epigenetic writer, EZH2, coordinate with proliferation. Urothelial cells were inoculated with UPEC, UPEC derivatives, or vehicle (mock infection) at low moi, washed, then maintained in media with Gentamycin. Urothelial conditioned media (CM) and extracellular vesicles (EV) were isolated after the inoculations and used to treat naïve urothelial cells. EZH2 increased with UPEC infection, inoculation-induced CM, and inoculation-induced EV vs. parallel stimulation derived from mock-inoculated urothelial cells. We found that infection also increased proliferation at one day post-infection, which was blocked by the EZH2 inhibitor UNC1999. Inhibition of demethylation at H3K27me3 had the opposite effect and augmented proliferation. CONCLUSION: Uropathogen-induced paracrine factors act epigenetically by altering expression of EZH2, which plays a key role in early host cell proliferative responses to infection. PMID:26964089

Chronic Heat Stress Impairs the Quality of Breast-Muscle Meat in Broilers by Affecting Redox Status and Energy-Substance Metabolism.

PubMed

Lu, Zhuang; He, Xiaofang; Ma, Bingbing; Zhang, Lin; Li, Jiaolong; Jiang, Yun; Zhou, Guanghong; Gao, Feng

2017-12-27

We investigated the molecular mechanisms by which chronic heat stress impairs the breast-meat quality of broilers. Broilers were assigned to three groups: the normal control (NC) group, heat-stress (HS) group, and pair-fed (PF) group. After 7 days of heat exposure (32 °C), the high temperature had caused oxidative stress; elevated the activity of citrate synthase (CS), the mRNA expression of M-CPT1, and the phosphorylation level of AMPKα; and reduced the mRNA expression of avUCP. After 14 days of heat exposure, the heat stress had increased the lightness and drip loss and decreased the pH and shear force of the breast meat. Additionally, the heat exposure had increased the mRNA expressions of FAS, ACC, and PDK4; the content of lipids; and the activities of lactic dehydrogenase and pyruvate kinase, and it had decreased the mRNA expression of M-CPT1 and the activity of CS. In conclusion, chronic heat stress impairs meat quality by causing mitochondria to malfunction and affecting energy-substance aerobic metabolism, resulting in increased glycolysis and intramuscular fat deposition.

Cyp1b1 deletion and retinol deficiency coordinately suppress mouse liver lipogenic genes and hepcidin expression during post-natal development

PubMed Central

Maguire, Meghan; Larsen, Michele Campaigne; Foong, Yee Hoon; Tanumihardjo, Sherry; Jefcoate, Colin R.

2018-01-01

Cyp1b1 deletion and gestational vitamin A deficiency (GVAD) redirect adult liver gene expression. A matched sufficient pre- and post-natal diet, which has high carbohydrate and normal iron content (LF12), increased inflammatory gene expression markers in adult livers that were suppressed by GVAD and Cyp1b1 deletion. At birth on the LF12 diet, Cyp1b1 deletion and GVAD each suppress liver expression of the iron suppressor, hepcidin (Hepc), while increasing stellate cell activation markers and suppressing post-natal increases in lipogenesis. Hepc was less suppressed in Cyp1b1−/− pups with a standard breeder diet, but was restored by iron supplementation of the LF12 diet. Conclusions The LF12 diet delivered low post-natal iron and attenuated Hepc. Hepc decreases in Cyp1b1−/− and GVAD mice resulted in stellate activation and lipogenesis suppression. Endothelial BMP6, a Hepc stimulant, is a potential coordinator and Cyp1b1 target. These neonatal changes in Cyp1b1−/− mice link to diminished adult obesity and liver inflammation. PMID:28583802

Overexpression of BID in thyroids of transgenic mice increases sensitivity to iodine-induced autoimmune thyroiditis

PubMed Central

2014-01-01

Background BID functions as a bridge molecule between death-receptor and mitochondrial related apoptotic pathways to amplify apoptotic signaling. Our previous studies have demonstrated a substantial increase in BID expression in primary normal thyroid epithelia cells treated with inflammatory cytokines, including the combination of IFNγ and IL-1β or IFNγ and TNFα. The aim of this study was to determine whether an increase in BID expression in thyroid can induce autoimmune thyroiditis. Methods A transgenic mouse line that expresses human BID in thyroid cells was established by fusing a mouse thyroglobulin (Tg) promoter upstream of human BID (Tg-BID). We tested whether the increased expression of pro-apoptotic BID in thyroid would induce autoimmune thyroiditis, both in the presence and absence of 0.3% iodine water. Results Our data show that Tg-BID mice in a CBA/J (H-2 k) background do not spontaneously develop autoimmune thyroiditis for over a year. However, upon ingestion of iodine in the drinking water, autoimmune thyroiditis does develop in Tg-BID transgenic mice, as shown by a significant increase in anti-Tg antibody and mononuclear cell infiltration in the thyroid glands in 30% of mice tested. Serum T4 levels, however, were similar between iodine-treated Tg-BID transgenic mice and the wild type mice. Conclusions Our data demonstrate that increased thyroid expression of BID facilitates the development of autoimmune thyroiditis induced by iodine uptake. However, the overexpression of BID itself is not sufficient to initiate thyroiditis in CBA/J (H-2 k) mice. PMID:24957380

The conclusiveness of less-invasive imaging techniques (computer tomography, X-ray) with regard to their identification of bone diseases in a primate model (Callithrix jacchus).

PubMed

Grohmann, J; Taetzner, S; Theuss, T; Kuehnel, F; Buchwald, U; Einspanier, A

2012-04-01

Although common marmosets seem to be appropriate animal models to examine bone diseases, no data about the conclusiveness of less-invasive techniques are available. Therefore, the aim was to combine different techniques to analyse changes in bone metabolism of common marmosets with bone diseases. Five monkeys were examined by X-ray, computer tomography (CT), histology and immunohistochemistry (IHC). Monkeys with lowest bone mineral density (BMD) showed increased bone marrow, decreased cancellous bone and decreased contrast in X-ray. Highest alkaline phosphatase (AP)-levels were detected in bones with low elastic modulus. Expression of osteopontin (OPN), osteocalcin (OC) and runt-related transcriptions factor 2 (RUNX 2) was detected in bones with high modulus. No expression was present in bones with lower modulus. Collagen type I and V were found in every bone. In conclusion, CT, X-ray and AP are useful techniques to detect bone diseases in common marmosets. These observations could be confirmed by IHC. © 2012 John Wiley & Sons A/S.

Evaluation of acetylcholinesterase in an animal model of maple syrup urine disease.

PubMed

Scaini, Giselli; de Rochi, Natália; Jeremias, Isabela C; Deroza, Pedro F; Zugno, Alexandra I; Pereira, Talita C B; Oliveira, Giovanna M T; Kist, Luiza W; Bogo, Maurício R; Schuck, Patrícia F; Ferreira, Gustavo C; Streck, Emilio L

2012-04-01

Maple syrup urine disease is an inherited metabolic disease predominantly characterized by neurological dysfunction. However, the mechanisms underlying the neuropathology of this disease are still not defined. Therefore, the aim of this study was to investigate the effect of acute and chronic administration of a branched-chain amino acids (BCAA) pool (leucine, isoleucine, and valine) on acetylcholinesterase (AChE) activity and gene expression in the brain and serum of rats and to assess if antioxidant treatment prevented the alterations induced by BCAA administration. Our results show that the acute administration of a BCAA pool in 10- and 30-day-old rats increases AChE activity in the cerebral cortex, striatum, hippocampus, and serum. Moreover, chronic administration of the BCAA pool also increases AChE activity in the structures studied, and antioxidant treatment prevents this increase. In addition, we show a significant decrease in the mRNA expression of AChE in the hippocampus following acute administration in 10- and 30-day-old rats. On the other hand, AChE expression increased significantly after chronic administration of the BCAA pool. Interestingly, the antioxidant treatment was able to prevent the increased AChE activity without altering AChE expression. In conclusion, the results from the present study demonstrate a marked increase in AChE activity in all brain structures following the administration of a BCAA pool. Moreover, the increased AChE activity is prevented by the coadministration of N-acetylcysteine and deferoxamine as antioxidants.

Polyamine and methionine adenosyltransferase 2A crosstalk in human colon and liver cancer

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

Tomasi, Maria Lauda; USC Research Center for Liver Diseases, Keck School of Medicine of University of Southern California, Los Angeles, CA 90033; The Southern California Research Center for Alcoholic and Pancreatic Diseases and Cirrhosis, Keck School of Medicine of University of Southern California, Los Angeles, CA 90033

Methionine adenosyltransferase (MAT) is an essential enzyme that is responsible for the biosynthesis of S-adenosylmethionine (SAMe), the principal methyl donor and precursor of polyamines. MAT1A is expressed in normal liver and MAT2A is expressed in all extrahepatic tissues. MAT2A expression is increased in human colon cancer and in colon cancer cells treated with mitogens, whereas silencing MAT2A resulted in apoptosis. The aim of the current work was to examine the mechanism responsible for MAT2A-dependent growth and apoptosis. We found that in RKO (human adenocarcinoma cell line) cells, MAT2A siRNA treatment lowered cellular SAMe and putrescine levels by 70–75%, increased apoptosismore » and inhibited growth. Putrescine supplementation blunted significantly MAT2A siRNA-induced apoptosis and growth suppression. Putrescine treatment (100 pmol/L) raised MAT2A mRNA level to 4.3-fold of control, increased the expression of c-Jun and c-Fos and binding to an AP-1 site in the human MAT2A promoter and the promoter activity. In human colon cancer specimens, the expression levels of MAT2A, ornithine decarboxylase (ODC), c-Jun and c-Fos are all elevated as compared to adjacent non-tumorous tissues. Overexpression of ODC in RKO cells also raised MAT2A mRNA level and MAT2A promoter activity. ODC and MAT2A are also overexpressed in liver cancer and consistently, similar MAT2A-ODC-putrescine interactions and effects on growth and apoptosis were observed in HepG2 cells. In conclusion, there is a crosstalk between polyamines and MAT2A. Increased MAT2A expression provides more SAMe for polyamines biosynthesis; increased polyamine (putrescine in this case) can activate MAT2A at the transcriptional level. This along with increased ODC expression in cancer all feed forward to further enhance the proliferative capacity of the cancer cell. -- Highlights: • MAT2A knockdown depletes putrescine and leads to apoptosis. • Putrescine attenuates MAT2A knockdown-induced apoptosis and growth suppression. • Putrescine induces AP-1, which activates MAT2A promoter to increase its expression. • Putrescine increases ornithine decarboxylase expression, which induce MAT2A promoter. • Expression of MAT2A correlates with that of ornithine decarboxylase in colon cancer.« less

MUC Expression in Gallbladder Epithelial Tissues in Cholesterol-Associated Gallbladder Disease

PubMed Central

Yoo, Kyo-Sang; Choi, Ho Soon; Jun, Dae Won; Lee, Hang Lak; Lee, Oh Young; Yoon, Byung Chul; Lee, Kyeong Geun; Paik, Seung Sam; Kim, Yong Seok; Lee, Jin

2016-01-01

Background/Aims Gallstone pathogenesis is linked to mucin hypersecretion and bacterial infection. Several mucin genes have been identified in gallbladder epithelial cells (GBECs). We investigated MUC expression in cholesterol-associated gallbladder disease and evaluated the relationship between mucin and bacterial infection. Methods The present study involved 20 patients with cholesterol stones with cholecystitis, five with cholesterol stones with cholesterolosis, six with cholesterol polyps, two with gallbladder cancer, and six controls. Canine GBECs treated with lipopolysaccharide were also studied. MUC3, MUC5AC, MUC5B, and MUC6 antibodies were used for dot/slot immunoblotting and immunohistochemical studies of the gallbladder epithelial tissues, canine GBECs, and bile. Reverse-transcription polymerase chain reaction was performed to evaluate MUC3 and MUC5B expression. Results MUC3, MUC5AC, MUC5B, and MUC6 were expressed in the normal gallbladder epithelium, and of those, MUC3 and MUC5B exhibited the highest expression levels. Greatly increased levels of MUC3 and MUC5B expression were observed in the cholesterol stone group, and slightly increased levels were observed in the cholesterol polyp group; MUC3 and MUC5B mRNA was also upregulated in those groups. Canine GBECs treated with lipopolysaccharide also showed upregulation of MUC3 and MUC5B. Conclusions The mucin genes with the highest expression levels in gallbladder tissue in cholesterol-associated diseases were MUC3 and MUC5B. Cholesterol stones and gallbladder infections were associated with increased MUC3 and MUC5B expression. PMID:27563024

Regulation of CCN2/CTGF Expression in the Nucleus Pulposus of the Intervertebral Disc: Role of Smad and AP1 Signaling

PubMed Central

Tran, Cassie M.; Markova, Dessislava; Smith, Harvey E.; Susarla, Bala; Ponnappan, Ravi Kumar; Anderson, D Greg; Symes, Aviva; Shapiro, Irving M.; Risbud, Makarand V.

2011-01-01

Objective To investigate TGFβ regulation of CTGF expression in cells of the nucleus pulposus. Methods Real Time RT-PCR and Western blot analysis was used to measure CTGF expression in the nucleus pulposus. Transfections were used to measure the effect of Smad2/3/7 and AP1on TGFβ mediated CTGF promoter activity. Results CTGF expression was lower in the neonatal disc compared with the skeletally mature rat disc. An increase in CTGF expression and promoter activity was observed in nucleus pulposus cells after TGFβ treatment. Deletion analysis indicated that promoter constructs lacking smad and AP1 motifs were unresponsive to treatment. Analysis showed that full-length Smad3 and the Smad3-MH2 domain alone increased CTGF activity. Further evidence of Smad3 and AP1 involvement was seen when DN-Smad3, SiRNA-Smad3, smad7 and DN-AP1 suppressed TGFβ mediated activation of the CTGF promoter. When either Smad3 or AP1 sites were mutated, CTGF promoter induction by TGFβ was suppressed. We also observed a decrease in expression of CTGF in discs of Smad3 null mice compared to the wild type. Analysis of human nucleus pulposus indicated a trend of increasing CTGF and TGFβ expression in the degenerate state. Conclusion TGFβ, through Smad3 and AP1, serves as a positive regulator of CTGF expression in the nucleus pulposus. We propose that CTGF is a part of the limited reparative response of the degenerate disc. PMID:20222112

Increased expression of G-protein-coupled receptor kinases 3 and 4 in hyperfunctioning thyroid nodules.

PubMed

Voigt, Carsten; Holzapfel, Hans-Peter; Meyer, Silke; Paschke, Ralf

2004-07-01

G-protein-coupled receptor kinases (GRKs) are implicated in the pathophysiology of human diseases such as arterial hypertension, heart failure and rheumatoid arthritis. While G-protein-coupled receptor kinases 2 and 5 have been shown to be involved in the desensitization of the rat thyrotropin receptor (TSHR), their role in the pathophysiology of hyperfunctioning thyroid nodules (HTNs) is unknown. Therefore, we analyzed the expression pattern of the known GRKs in human thyroid tissue and investigated their function in the pathology of HTNs. The expression of different GRKs in human thyroid and HTNs was measured by Western blotting. The influence of GRK expression on TSHR function was analyzed by coexpression experiments in HEK 293 cells. We demonstrate that in addition to GRKs 2, 5 and 6, GRKs 3 and 4 are also expressed in the human thyroid. GRKs 2, 3, 5 and 6 are able to desensitize the TSHR in vitro. This GRK-induced desensitization is amplified by the additional over-expression of beta-arrestin 1 or 2. We did not find any mutations in the GRKs 2, 3 and 5 from 14 HTNs without TSHR mutations and Gsalpha mutations. The expression of GRKs 3 and 4 was increased in HTNs independently from the existence of TSHR mutations or Gsalpha mutations. In conclusion, the increased expression of GRK 3 in HTNs and the ability of GRK 3 to desensitize the TSHR in vitro, suggest a potential role for GRK 3 as a negative feedback regulator for the constitutively activated cAMP pathway in HTNs.

Effects of bromopride on expression of metalloproteinases and interleukins in left colonic anastomoses: an experimental study

PubMed Central

Silva, S.M.; Jerônimo, M.S.; Silva-Pereira, I.; Bocca, A.L.; Sousa, J.B.

2014-01-01

Anastomotic dehiscence is the most severe complication of colorectal surgery. Metalloproteinases (MMPs) and interleukins (ILs) can be used to analyze the healing process of anastomosis. To evaluate the effects of bromopride on MMP and cytokine gene expression in left colonic anastomoses in rats with or without induced abdominal sepsis, 80 rats were divided into two groups for euthanasia on the third or seventh postoperative day (POD). They were then divided into subgroups of 20 rats for sepsis induction or not, and then into subgroups of 10 rats for administration of bromopride or saline. Left colonic anastomosis was performed and abdominal sepsis was induced by cecal ligation and puncture. A colonic segment containing the anastomosis was removed for analysis of gene expression of MMP-1α, MMP-8, MMP-13, IL-β, IL-6, IL-10, tumor necrosis factor-α (TNF-α), and interferon-γ (IFN-γ). On the third POD, bromopride was associated with increased MMP-1α, MMP-13, IL-6, IFN-γ, and IL-10 gene expression. On the seventh POD, all MMP transcripts became negatively modulated and all IL transcripts became positively modulated. In the presence of sepsis, bromopride administration increased MMP-8 and IFN-γ gene expression and decreased MMP-1, TNF-α, IL-6, and IL-10 gene expression on the third POD. On the seventh POD, we observed increased expression of MMP-13 and all cytokines, except for TNF-α. In conclusion, bromopride interferes with MMP and IL gene expression during anastomotic healing. Further studies are needed to correlate these changes with the healing process. PMID:25140813

Effects of bromopride on expression of metalloproteinases and interleukins in left colonic anastomoses: an experimental study.

PubMed

Silva, S M; Jerônimo, M S; Silva-Pereira, I; Bocca, A L; Sousa, J B

2014-10-01

Anastomotic dehiscence is the most severe complication of colorectal surgery. Metalloproteinases (MMPs) and interleukins (ILs) can be used to analyze the healing process of anastomosis. To evaluate the effects of bromopride on MMP and cytokine gene expression in left colonic anastomoses in rats with or without induced abdominal sepsis, 80 rats were divided into two groups for euthanasia on the third or seventh postoperative day (POD). They were then divided into subgroups of 20 rats for sepsis induction or not, and then into subgroups of 10 rats for administration of bromopride or saline. Left colonic anastomosis was performed and abdominal sepsis was induced by cecal ligation and puncture. A colonic segment containing the anastomosis was removed for analysis of gene expression of MMP-1α, MMP-8, MMP-13, IL-β, IL-6, IL-10, tumor necrosis factor-α (TNF-α), and interferon-γ (IFN-γ). On the third POD, bromopride was associated with increased MMP-1α, MMP-13, IL-6, IFN-γ, and IL-10 gene expression. On the seventh POD, all MMP transcripts became negatively modulated and all IL transcripts became positively modulated. In the presence of sepsis, bromopride administration increased MMP-8 and IFN-γ gene expression and decreased MMP-1, TNF-α, IL-6, and IL-10 gene expression on the third POD. On the seventh POD, we observed increased expression of MMP-13 and all cytokines, except for TNF-α. In conclusion, bromopride interferes with MMP and IL gene expression during anastomotic healing. Further studies are needed to correlate these changes with the healing process.

CONNECTIVE TISSUE GROWTH FACTOR IS A TARGET OF NOTCH SIGNALING IN CELLS OF THE OSTEOBLASTIC LINEAGE

PubMed Central

Canalis, Ernesto; Zanotti, Stefano; Smerdel-Ramoya, Anna

2014-01-01

Connective tissue growth factor (Ctgf) or CCN2 is a protein synthesized by osteoblasts necessary for skeletal homeostasis, although its overexpression inhibits osteogenic signals and bone formation. Ctgf is induced by bone morphogenetic proteins, transforming growth factor β and Wnt; and in the present studies, we explored whether Notch regulated Ctgf expression in osteoblasts. We employed RosaNotch mice, where the Notch intracellular domain (NICD) is expressed following the excision of a STOP cassette, placed between the Rosa26 promoter and NICD. Notch was activated by transduction of adenoviral vectors expressing Cre recombinase (Ad-CMV-Cre). Notch induced Ctgf mRNA levels in a time dependent manner and increased Ctgf heterogeneous nuclear RNA. Notch also destabilized Ctgf mRNA shortening its half-life from 13 h to 3 h. The effect of Notch on Ctgf expression was lost following Rbpjκ downregulation, demonstrating that it was mediated by Notch canonical signaling. However, downregulation of the classic Notch target genes Hes1, Hey1 and Hey2 did not modify the effect of Notch on Ctgf expression. Wild type osteoblasts exposed to immobilized Delta-like 1 displayed enhanced Notch signaling and increased Ctgf expression. In addition to the effects of Notch in vitro, Notch induced Ctgf in vivo, and calvariae and femurs from RosaNotch mice mated with transgenics expressing the Cre recombinase in cells of the osteoblastic lineage exhibited increased expression of Ctgf. In conclusion, Ctgf is a target of Notch canonical signaling in osteoblasts, and may act in concert with Notch to regulate skeletal homeostasis. PMID:24792956

Liarozole inhibits transforming growth factor-β3–mediated extracellular matrix formation in human three-dimensional leiomyoma cultures

PubMed Central

Levy, Gary; Malik, Minnie; Britten, Joy; Gilden, Melissa; Segars, James; Catherino, William H.

2014-01-01

Objective To investigate the impact of liarozole on transforming growth factor-β3 (TGF-β3) expression, TGF-β3 controlled profibrotic cytokines, and extracellular matrix formation in a three-dimensional (3D) leiomyoma model system. Design Molecular and immunohistochemical analysis in a cell line evaluated in a three-dimensional culture. Setting Laboratory study. Patient(s) None. Intervention(s) Treatment of leiomyoma and myometrial cells with liarozole and TGF-β3 in a three-dimensional culture system. Main Outcome Measure(s) Quantitative real-time reverse-transcriptase polymerase chain reaction and Western blotting to assess fold gene and protein expression of TGF-β3 and TGF-β3 regulated fibrotic cytokines: collagen 1A1 (COL1A1), fibronectin, and versican before and after treatment with liarozole, and confirmatory immunohistochemical stains of treated three-dimensional cultures. Result(s) Both TGF-β3 gene and protein expression were elevated in leiomyoma cells compared with myometrium in two-dimensional and 3D cultures. Treatment with liarozole decreased TGF-β3 gene and protein expression. Extracellular matrix components versican, COL1A1, and fibronectin were also decreased by liarozole treatment in 3D cultures. Treatment of 3D cultures with TGF-β3 increased gene expression and protein production of COL1A1, fibronectin, and versican. Conclusion(s) Liarozole decreased TGF-β3 and TGF-β3–mediated extracellular matrix expression in a 3D uterine leiomyoma culture system. PMID:24825427

Estrogen Enhances the Expression of the Multidrug Transporter Gene ABCG2—Increasing Drug Resistance of Breast Cancer Cells through Estrogen Receptors

PubMed Central

Chang, Fung-Wei; Fan, Hueng-Chuen; Liu, Jui-Ming; Fan, Tai-Ping; Jing, Jin; Yang, Chia-Ling; Hsu, Ren-Jun

2017-01-01

Background: Multidrug resistance is a major obstacle in the successful therapy of breast cancer. Studies have proved that this kind of drug resistance happens in both human cancers and cultured cancer cell lines. Understanding the molecular mechanisms of drug resistance is important for the reasonable design and use of new treatment strategies to effectively confront cancers. Results: In our study, ATP-binding cassette sub-family G member 2 (ABCG2), adenosine triphosphate (ATP) synthase and cytochrome c oxidase subunit VIc (COX6C) were over-expressed more in the MCF-7/MX cell line than in the normal MCF7 cell line. Therefore, we believe that these three genes increase the tolerance of MCF7 to mitoxantrone (MX). The data showed that the high expression of COX6C made MCF-7/MX have more stable on mitochondrial membrane potential (MMP) and reactive oxygen species (ROS) expression than normal MCF7 cells under hypoxic conditions. The accumulation of MX was greater in the ATP-depleted treatment MCF7/MX cells than in normal MCF7/MX cells. Furthermore, E2 increased the tolerance of MCF7 cells to MX through inducing the expression of ABCG2. However, E2 could not increase the expression of ABCG2 after the inhibition of estrogen receptor α (ERα) in MCF7 cells. According to the above data, under the E2 treatment, MDA-MB231, which lacks ER, had a higher sensitivity to MX than MCF7 cells. Conclusions: E2 induced the expression of ABCG2 through ERα and the over-expressed ABCG2 made MCF7 more tolerant to MX. Moreover, the over-expressed ATP synthase and COX6c affected mitochondrial genes and function causing the over-expressed ABCG2 cells pumped out MX in a concentration gradient from the cell matrix. Finally lead to chemoresistance. PMID:28098816

Skeletal muscle deiodinase type 2 regulation during illness in mice.

PubMed

Kwakkel, J; van Beeren, H C; Ackermans, M T; Platvoet-Ter Schiphorst, M C; Fliers, E; Wiersinga, W M; Boelen, A

2009-11-01

We have previously shown that skeletal muscle deiodinase type 2 (D2) mRNA (listed as Dio2 in MGI Database) is upregulated in an animal model of acute illness. However, human studies on the expression of muscle D2 during illness report conflicting data. Therefore, we evaluated the expression of skeletal muscle D2 and D2-regulating factors in two mouse models of illness that differ in timing and severity of illness: 1) turpentine-induced inflammation, and 2) Streptococcus pneumoniae infection. During turpentine-induced inflammation, D2 mRNA and activity increased compared to pair-fed controls, most prominently at day 1 and 2, whereas after S. pneumoniae infection D2 mRNA decreased. We evaluated the association of D2 expression with serum thyroid hormones, (de-)ubiquitinating enzymes ubiquitin-specific peptidase 33 and WD repeat and SOCS box-containing 1 (Wsb1), cytokine expression and activation of inflammatory pathways and cAMP pathway. During chronic inflammation the increased muscle D2 expression is associated with the activation of the cAMP pathway. The normalization of D2 5 days after turpentine injection coincides with increased Wsb1 and tumor necrosis factor alpha expression. Muscle interleukin-1beta (Il1b) expression correlated with decreased D2 mRNA expression after S. pneumoniae infection. In conclusion, muscle D2 expression is differentially regulated during illness, probably related to differences in the inflammatory response and type of pathology. D2 mRNA and activity increases in skeletal muscle during the acute phase of chronic inflammation compared to pair-fed controls probably due to activation of the cAMP pathway. In contrast, muscle D2 mRNA decreases 48 h after a severe bacterial infection, which is associated with local Il1b mRNA expression and might also be due to diminished food-intake.

Relationship between serum levels of triglycerides and vascular inflammation, measured as COX-2, in arteries from diabetic patients: a translational study

PubMed Central

2013-01-01

Background Inflammation is a common feature in the majority of cardiovascular disease, including Diabetes Mellitus (DM). Levels of pro-inflammatory markers have been found in increasing levels in serum from diabetic patients (DP). Moreover, levels of Cyclooxygenase-2 (COX-2) are increased in coronary arteries from DP. Methods Through a cross-sectional design, patients who underwent CABG were recruited. Vascular smooth muscle cells (VSMC) were cultured and COX-2 was measured by western blot. Biochemical and clinical data were collected from the medical record and by blood testing. COX-2 expression was analyzed in internal mammary artery cross-sections by confocal microscopy. Eventually, PGI2 and PGE2 were assessed from VSMC conditioned media by ELISA. Results Only a high glucose concentration, but a physiological concentration of triglycerides exposure of cultured human VSMC derived from non-diabetic patients increased COX-2 expression .Diabetic patients showed increasing serum levels of glucose, Hb1ac and triglycerides. The bivariate analysis of the variables showed that triglycerides was positively correlated with the expression of COX-2 in internal mammary arteries from patients (r2 = 0.214, P < 0.04). Conclusions We conclude that is not the glucose blood levels but the triglicerydes leves what increases the expression of COX-2 in arteries from DP. PMID:23642086

Lung fibrosis-associated soluble mediators and bronchoalveolar lavage from idiopathic pulmonary fibrosis patients promote the expression of fibrogenic factors in subepithelial lung myofibroblasts.

PubMed

Bouros, Evangelos; Filidou, Eirini; Arvanitidis, Konstantinos; Mikroulis, Dimitrios; Steiropoulos, Paschalis; Bamias, George; Bouros, Demosthenes; Kolios, George

2017-10-01

Idiopathic pulmonary fibrosis (IPF) is characterized by infiltration of inflammatory cells, excessive collagen production and accumulation of myofibroblasts. We explored the possible role of subepithelial lung myofibroblasts (SELMs) in the development of fibrosis in IPF. SELMs, isolated from surgical specimens of healthy lung tissue, were cultured with pro-inflammatory factors or bronchoalveolar lavage fluid (BALF) from patients with IPF or idiopathic non-specific interstitial pneumonia (iNSIP) and their fibrotic activity was assessed. Stimulation of SELMs with pro-inflammatory factors induced a significant increase of Tissue Factor (TF) and Tumor necrosis factor-Like cytokine 1 A (TL1A) expression and collagen production in culture supernatants. Stimulation with BALF from IPF patients with mild to moderate, but not severe disease, and from iNSIP patients induced a significant increase of TF expression. BALF from all IPF patients induced a significant increase of TL1A expression and collagen production, while BALF from iNSIP patients induced a significant increase of TL1A, but not of collagen production. Interestingly, TGF-β1 and BALF from all IPF, but not iNSIP patients, induced a significant increase in SELMs migration. In conclusion, BALF from IPF patients induces fibrotic activity in lung myofibroblasts, similar to mediators associated with lung fibrosis, indicating a key role of SELMs in IPF. Copyright © 2017 Elsevier Ltd. All rights reserved.

Overexpression of xeroderma pigmentosum group C decreases the chemotherapeutic sensitivity of colorectal carcinoma cells to cisplatin.

PubMed

Zhang, Yi; Cao, Jia; Meng, Yanni; Qu, Chunying; Shen, Feng; Xu, Leiming

2018-05-01

Xeroderma pigmentosum group C (XPC) is a DNA-damage-recognition gene active at the early stage of DNA repair. XPC also participates in regulation of cell-cycle checkpoint and DNA-damage-induced apoptosis. In the present study, the expression levels of genes involved in nucleotide excision repair (NER) were assessed in human colorectal cancer (CRC) tissue. This analysis revealed that expression of XPC mRNA significantly increased in colorectal carcinoma tissues compared with matched normal controls. Expression of XPC gradually increased along with the degree of progression of CRC. In vitro , an XTT assay demonstrated that small interfering RNA (siRNA) targeting XPC significantly increased the sensitivity of CRC SW480 cells to cisplatin, whereas cells transfected with a XPC-overexpression plasmid became more resistant to cisplatin. Furthermore, flow cytometry revealed that the proportion of apoptotic cells significantly increased in XPC-knockdown cells upon cisplatin treatment. However, the overexpression XPC significantly increased the resistance of cells to cisplatin. In vivo , tumor growth was significantly reduced in tumor-bearing mice when the XPC gene was knocked down. Upregulation of the expression of pro-apoptotic Bcl-associated X and downregulation of the anti-apoptotic B-cell lymphoma 2 proteins was observed in the implanted tumor tissue. In conclusion, XPC serves a key role in chemotherapeutic sensitivity of CRC to cisplatin, meaning that it may be a potential target for chemotherapy of CRC.

IL-6 mediates differentiation disorder during spermatogenesis in obesity-associated inflammation by affecting the expression of Zfp637 through the SOCS3/STAT3 pathway.

PubMed

Huang, Guizhen; Yuan, Miao; Zhang, Jie; Li, Jun; Gong, Di; Li, Yanyan; Zhang, Jie; Lin, Ping; Huang, Lugang

2016-06-22

Zfp637 is a recently identified zinc finger protein, and its functions remain largely unknown. Here, we innovatively demonstrate the effects of Zfp637 on the differentiation of mouse spermatogonia and on its downstream target gene SOX2 in vitro. Obesity has been recognized as a chronic inflammatory disease that leads to decreased sexual function and sexual development disorders. We observed higher levels of IL-6 in serum and testis homogenates from obese mice compared with control mice. We also demonstrated that high levels of IL-6 inhibited Zfp637 expression, and we elucidated the underlying mechanisms. SOCS3 overexpression and STAT3 phosphorylation inhibitor (AG490) were used to investigate the function of the SOCS3/STAT3 pathway during this process. Our results showed that exposure of mouse spermatogonial cells to high levels of IL-6 inhibited Zfp637 expression by increasing SOCS3 expression and inhibiting the phosphorylation of STAT3, further reducing cellular differentiation. Consistent with the in vitro results, we observed increasing expression levels of SOCS3 and SOX2, but a reduction of Zfp637 expression, in obese mouse testes. In conclusion, Zfp637 plays a crucial role in spermatogenesis by downregulating SOX2 expression, and IL-6 can decrease the expression of Zfp637 through the SOCS3/STAT3 signaling pathway.

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.

Study of NGEP expression in androgen sensitive prostate cancer cells: A potential target for immunotherapy

PubMed Central

Mohsenzadegan, Monireh; Tajik, Nader; Madjd, Zahra; Shekarabi, Mehdi; Farajollahi, Mohammad M

2015-01-01

Background: Prostate cancer is one of the leading causes of cancer deaths among men. New gene expressed in prostate (NGEP), is a prostate-specific gene expressed only in normal prostate and prostate cancer tissue. Because of its selective expression in prostate cancer cell surface, NGEP is a potential immunotherapeutic target. To target the NGEP in prostate cancer, it is essential to investigate its expression in prostate cancer cells. Methods: In the present study, we investigated NGEP expression in LNCaP and DU145 cells by real time and RT-PCR, flow cytometric and immunocytochemical analyses. Results: Real time and RT-PCR analyses of NGEP expression showed that NGEP was expressed in the LNCaP cells but not in DU145 cells. The detection of NGEP protein by flow cytometric and immunocytochemistry analyses indicated that NGEP protein was weakly expressed only in LNCaP cell membrane. Conclusion: Our results demonstrate that LNCaP cell line is more suitable than DU145 for NGEP expression studies; however, its low-level expression is a limiting issue. NGEP expression may be increased by androgen supplementation of LNCaP cell culture medium. PMID:26000254

Intermittent Fasting Alleviates the Increase of Lipoprotein Lipase Expression in Brain of a Mouse Model of Alzheimer's Disease: Possibly Mediated by β-hydroxybutyrate.

PubMed

Zhang, Jingzhu; Li, Xinhui; Ren, Yahao; Zhao, Yue; Xing, Aiping; Jiang, Congmin; Chen, Yanqiu; An, Li

2018-01-01

Intermittent fasting has been demonstrated to protect against Alzheimer's disease (AD), however, the mechanism is unclear. Histone acetylation and lipoprotein lipase (LPL) are involved in AD progression. Importantly, LPL has been documented to be regulated by histone deacetylases (HDACs) inhibitors (increase histone acetylation level) in adipocyte and mesenchymal stem cells, or by fasting in adipose and muscle tissues. In brain, however, whether histone acetylation or fasting regulates LPL expression is unknown. This study was designed to demonstrate intermittent fasting may protect against AD through increasing β-hydroxybutyrate, a HDACs inhibitor, to regulate LPL. We also investigated microRNA-29a expression associating with regulation of LPL and histone acetylation. The results showed LPL mRNA expression was increased and microRNA-29a expression was decreased in the cerebral cortex of AD model mice (APP/PS1), which were alleviated by intermittent fasting. No significant differences were found in the total expression of LPL protein (brain-derived and located in capillary endothelial cells from peripheral tissues) in the cerebral cortex of APP/PS1 mice. Further study indicated that LPL located in capillary endothelial cells was decreased in the cerebral cortex of APP/PS1 mice, which was alleviated by intermittent fasting. LPL and microRNA-29a expression were separately increased and down-regulated in 2 μM Aβ 25-35 -exposed SH-SY5Y cells, but respectively decreased and up-regulated in 10 μM Aβ 25-35 -exposed cells, which were all reversed by β-hydroxybutyrate. The increase of HDAC2/3 expression and the decrease of acetylated H3K9 and H4K12 levels were alleviated in APP/PS1 mice by intermittent fasting treatment, as well in 2 or 10 μM Aβ 25-35 -exposed cells by β-hydroxybutyrate treatment. These findings above suggested the results from APP/PS1 mice were consistent with those from cells treated with 2 μM Aβ 25-35 . Interestingly, LPL expression was reduced (0.2-folds) and microRNA-29a expression was up-regulated (1.7-folds) in HDAC2-silenced cells, but respectively increased (1.3-folds) and down-regulated (0.8-folds) in HDAC3-silenced cells. Furthermore, LPL expression was decreased in cells treated with microRNA-29a mimic and increased with inhibitor treatment. In conclusion, intermittent fasting inhibits the increase of brain-derived LPL expression in APP/PS1 mice partly through β-hydroxybutyrate-mediated down-regulation of microRNA-29a expression. HDAC2/3 may be implicated in the effect of β-hydroxybutyrate on microRNA-29a expression.

The influence of fluid shear stress on the expression of Cbfa1 in MG-63 cells cultured under different gravitational conditions

NASA Astrophysics Data System (ADS)

Zhang, S.; Wang, B.; Cao, X. S.; Yang, Z.; Sun, X. Q.

2008-12-01

AuthorPurposeThis study was aimed to explore the effect of flow shear stress on the expression of Cbfa1 in human osteosarcoma cells and to survey its functional alteration in simulated microgravity. After culture for 48 h in two different gravitational environments, i.e. 1 G terrestrial gravitational condition and simulated microgravity condition, human osteosarcoma cells (MG-63) were treated with 0.5 or 1.5 Pa fluid shear stress (FSS) in a flow chamber for 15, 30, and 60 min, respectively. The total RNA in cells was isolated. RT-PCR analysis was made to examine the gene expression of Cbfa1. The total protein of cells was extracted and the expression of Cbfa1 protein was detected by means of Western blotting. ResultsMG-63 cells cultured in 1 G condition reacted to FSS treatment with an enhanced expression of Cbfa1. Compared with no-FSS control group, Cbfa1 mRNA expression increased significantly at 30 and 60 min with the treatment of FSS ( P < 0.01). And there was remarkable difference on the Cbfa1 mRNA expression between the treatments of 0.5 and 1.5 Pa FSS at 30 or 60 min ( P < 0.01). Cbfa1 protein expressions had a trend to increase at 30 min with the treatment of FSS and they increased significantly at 60 min with the treatment of 0.5 or 1.5 Pa FSS ( P < 0.05). As to the cells cultured in simulated microgravity by using clinostat, the expression of Cbfa1 was significantly different between 1 G and simulated microgravity conditions at each test time ( P < 0.05). Compared with no-FSS control group cultured in simulated microgravity, Cbfa1 mRNA expression increased significantly at 30 and 60 min with the treatment of FSS ( P < 0.05). And Cbfa1 protein expression increased significant at 60 min with the treatment of 1.5 Pa FSS under simulated microgravity conditions ( P < 0.05). ConclusionsFSS can significantly increase the gene and protein expression of Cbfa1 in human osteosarcoma cells. And this inducible function of FSS was adversely affected by simulated microgravity.

Increased expression of transforming growth factor beta s after acute oedematous pancreatitis in rats suggests a role in pancreatic repair.

PubMed Central

Riesle, E; Friess, H; Zhao, L; Wagner, M; Uhl, W; Baczako, K; Gold, L I; Korc, M; Büchler, M W

1997-01-01

BACKGROUND: Transforming growth factor beta isoforms (TGF beta s) belong to a family of multifunctional regulators of cellular growth and differentiation. They are mitogenic and chemotactic for fibroblasts and are potent stimulators of extracellular matrix production (collagen) and deposition. Upregulation of TGF beta transcription has been reported for several in vivo systems during repair after injury. AIMS: To study the expression of the three mammalian isoforms of TGF beta (TGF beta 1-3) and their relation to collagen expression as a marker for fibroblast response in acute oedematous pancreatitis in rats. METHODS: Using northern blot analysis and immunohistochemistry, the expression and localisation of TGF beta isoforms, collagen, and amylase were analysed during the course of acute oedematous pancreatitis in rats, experimentally induced by intravenous caerulein infusion. RESULTS: Induction of acute pancreatitis resulted in a biphasic peak pattern of expression of TGF beta 1, beta 2, and beta 3 mRNA, with a pronounced increase from day 1 to day 3 (sixfold, 2.5-fold, fivefold, respectively) and again from day 5 to day 7 (three-fold, 2.3-fold, 3.5-fold, respectively). The temporal changes in TGF beta mRNA identically paralleled the expression in collagen mRNA. In contrast, amylase mRNA expression, used as a general indicator of acinar cell integrity, was slightly decreased after induction of acute pancreatitis. Immunohistochemical analysis of pancreatitis tissue showed that increased expression of TGF beta s was mainly present in the pancreatic acinar and ductal cells; this was evident within one day after pancreatitis induction. CONCLUSION: Overexpression of TGF beta s after induction of acute pancreatitis suggests a role for these proteins in pancreatic repair and remodelling. The increased levels of TGF beta s may help suppress immune activation, and may contribute to the increase in the extracellular matrix including collagen and to the repair of the pancreatic parenchyma. Images PMID:9155579

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

Kim, Kook Hwan; Samsung Advanced Institute for Health Sciences and Technology, Sungkyunkwan University School of Medicine, 50 Irwon-dong Gangnam-gu, Seoul 135-710; Jeong, Yeon Taek

Highlights: •Metformin induces FGF21 expression in an AMPK independent manner. •Metformin enhances FGF21 expression by inhibiting mitochondrial complex I activity. •The PERK-eIF2α-ATF4 axis is required for metformin-induced FGF21 expression. •Metformin activates the ATF4-FGF21 axis in the liver of mouse. •Metformin increases serum FGF21 level in diabetic human subjects. -- Abstract: Fibroblast growth factor 21 (FGF21) is an endocrine hormone that exhibits anti-obesity and anti-diabetes effects. Because metformin is widely used as a glucose-lowering agent in patients with type 2 diabetes (T2D), we investigated whether metformin modulates FGF21 expression in cell lines, and in mice or human subjects. We found thatmore » metformin increased the expression and release of FGF21 in a diverse set of cell types, including rat hepatoma FaO, primary mouse hepatocytes, and mouse embryonic fibroblasts (MEFs). Intriguingly, AMP-activated protein kinase (AMPK) was dispensable for the induction of FGF21 by metformin. Mammalian target of rapamycin complex 1 (mTORC1) and peroxisome proliferator-activated receptor α (PPARα), which are additional targets of metformin, were not involved in metformin-induced FGF21 expression. Importantly, inhibition of mitochondrial complex I activity by metformin resulted in FGF21 induction through PKR-like ER kinase (PERK)-eukaryotic translation factor 2α (eIF2α)-activating transcription factor 4 (ATF4). We showed that metformin activated ATF4 and increased FGF21 expression in the livers of mice, which led to increased serum levels of FGF21. We also found that serum FGF21 level was increased in human subjects with T2D after metformin therapy for 6 months. In conclusion, our results indicate that metformin induced expression of FGF21 through an ATF4-dependent mechanism by inhibiting mitochondrial respiration independently of AMPK. Therefore, FGF21 induction by metformin might explain a portion of the beneficial metabolic effects of metformin.« less

Uric acid upregulates the adiponectin-adiponectin receptor 1 pathway in renal proximal tubule epithelial cells

PubMed Central

Yang, Qingmei; Fu, Chensheng; Xiao, Jing; Ye, Zhibin

2018-01-01

Adiponectin (APN) is a protein hormone that is primarily derived from adipocytes. It can also be secreted by renal cells. Hypoadiponectinemia has been documented in patients with hyperuricemia, however, whether soluble uric acid (SUA) regulates the expression of APN and APN receptor 1 (AdipoR1) in renal proximal tubule epithelial cells (PTECs) remains to be elucidated. The present study investigated the expression of APN and AdipoR1 in cultured PTECs that were exposed to SUA through immunofluorescence and western blot analysis. In addition, Sprague-Dawley rats with oxonic acid-induced hyperuricemia (HUA) with or without febuxostat treatment were employed as an animal model to measure 24 h urine protein, serum creatinine, urea nitrogen, uric acid and homeostasis model assessment of insulin resistance. Renal pathology was evaluated using hematoxylin and eosin and immunohistochemical staining. APN and AdipoR1 expression in the renal cortex were evaluated by western blotting. The results demonstrated that, in PTECs, the expression of APN and AdipoR1 was constant and increased upon SUA exposure. Similar observations were made within the proximal renal tubules of rats, and the oxonic acid-induced increases in APN and AdipoR1 were offset by febuxostat treatment. Furthermore, SUA-treated PTECs exhibited an increase in the expression of NLR family pyrin domain-containing (NLRP) 3, which was dose-dependent. NLRP3 expression was also significantly increased in the renal cortex of HUA rats compared with control and febuxostat-treated rats. In conclusion, SUA enhanced the expression of APN and AdipoR1 in PTECs, which was associated with an increase in NLRP3 expression. The APN-AdipoR1 pathway was demonstrated to have an important role in in vitro and in vivo models of renal proximal tubule inflammatory injury. Therefore, this pathway may be a potential therapy target in urate nephropathy. PMID:29359786

INCREASED EXPRESSION OF GLIAL FIBRILLARY ACIDIC PROTEIN IN CEREBELLUM AND HIPPOCAMPUS: DIFFERENTIAL EFFECTS ON NEONATAL BRAIN REGIONAL ACETYLCHOLINESTERASE FOLLOWING MATERNAL EXPOSURE TO COMBINED CHLORPYRIFOS AND NICOTINE. (R829399)

EPA Science Inventory

The perspectives, information and conclusions conveyed in research project abstracts, progress reports, final reports, journal abstracts and journal publications convey the viewpoints of the principal investigator and may not represent the views and policies of ORD and EPA. Concl...

CD44 Promotes Inflammation and Extracellular Matrix Production During Arteriovenous Fistula Maturation

PubMed Central

Kuwahara, Go; Hashimoto, Takuya; Tsuneki, Masayuki; Yamamoto, Kota; Assi, Roland; Foster, Trenton R; Hanisch, Jesse J; Bai, Hualong; Hu, Haidi; Protack, Clinton D; Hall, Michael R; Schardt, John S; Jay, Steven M; Madri, Joseph A; Kodama, Shohta; Dardik, Alan

2017-01-01

Objective Arteriovenous fistulae (AVF) remain the optimal conduit for hemodialysis access but continue to demonstrate poor patency and poor rates of maturation. We hypothesized that CD44, a widely expressed cellular adhesion molecule that serves as a major receptor for extracellular matrix (ECM) components, promotes wall thickening and ECM deposition during AVF maturation. Approach and Results AVF were created via needle puncture in wild-type (WT) C57BL/6J and CD44 knockout (KO) mice. CD44 mRNA and protein expression was increased in WT AVF. CD44 KO mice showed no increase in AVF wall thickness (8.9 μm vs. 26.8 μm; P = 0.0114), collagen density, and hyaluronic acid density, but similar elastin density when compared to control AVF. CD44 KO mice also showed no increase in VCAM-1 expression, ICAM-1 expression and MCP-1 expression in the AVF compared to controls; there were also no increased M2 macrophage markers (TGM2: 81.5 fold, P = 0.0015; IL-10: 7.6 fold, P = 0.0450) in CD44 KO mice. Delivery of MCP-1 to CD44 KO mice rescued the phenotype with thicker AVF walls (27.2 μm vs. 14.7 μm; P = 0.0306), increased collagen density (2.4 fold; P = 0.0432), and increased number of M2 macrophages (2.1 fold; P = 0.0335). Conclusions CD44 promotes accumulation of M2 macrophages, ECM deposition and wall thickening during AVF maturation. These data show the association of M2 macrophages with wall thickening during AVF maturation and suggest that enhancing CD44 activity may be a strategy to increase AVF maturation. PMID:28450292

Association of BDNF and BMPR1A with clinicopathologic parameters in benign and malignant gallbladder lesions

PubMed Central

2013-01-01

Background Neurotrophic factors such as brain derived neurotrophic factor (BDNF) are synthesized in a variety of neural and non-neuronal cell types and regulate survival, proliferation and apoptosis. In addition, bone morphogenetic proteins (BMPs) inhibit the proliferation of pulmonary large carcinoma cells bone morphogenetic protein receptor, type IA (BMPR1A). Little is known about the expression of BDNF or BMPR1A in malignant gall bladder lesions. This study was to evaluate BDNF and BMPR1A expression and evaluate the clinicopathological significance in benign and malignant lesions of the gallbladder. Methods The BDNF and BMPR1A expression of gallbladder adenocarcinoma, peritumoral tissues, adenoma, polyp and chronic cholecystitis were Immunohistochemically determined. Results BDNF expression was significantly higher in gallbladder adenocarcinoma than in peritumoral tissues, adenoma, polyps and chronic cholecystitis samples. However, BMPR1A expression was significantly lower in gallbladder adenocarcinoma than in peritumoral tissues, adenomas, polyps and chronic cholecystitis tissues. The specimens with increased expression of BDNF in the benign lesions exhibited moderate- or severe-dysplasia of gallbladder epithelium. BDNF expression was significantly lower in well-differentiated adenocarcinomas with maximum tumor diameter <2 cm, no metastasis to lymph nodes, and no invasion of regional tissues compared to poorly-differentiated adenocarcinomas with maximal tumor diameter >2 cm, metastasis of lymph node, and invasiveness of regional tissues in gallbladder adenocarcinoma. BMPR1A expression were significantly higher in the well-differentiated adenocarcinoma with maximal tumor diameter <2 cm, no metastasis of lymph node, and no invasion of regional tissues compared to poorly-differentiated adenocarcinomas with maximal tumor diameter >2 cm, metastasis of lymph node, and invasiveness of regional tissues in gallbladder. Univariate Kaplan-Meier analysis indicated increased expression of BDNF or decreased expression of BMPR1A was associated with decreased disease specific survival (DSS) rates. Similarly, multivariate Cox regression analysis showed increased expression of BDNF or decreased expression of BMPR1A are independent predictors of poor DSS rates in gallbladder adenocarcinoma. Conclusions In gallbladder malignancies, the increased expression of BDNF and decreased expression of BMPR1A were associated with increased risk of metastasis, regional invasion and mortality. They might serve as novel indicators of gallbladder adenocarcinoma outcomes, which may prove valuable for the development of personalized therapeutic paradigms. PMID:23531103

Coordinated Regulation Among Progesterone, Prostaglandins, and EGF-Like Factors in Human Ovulatory Follicles

PubMed Central

Choi, Yohan; Wilson, Kalin; Hannon, Patrick R.; Rosewell, Katherine L.; Brännström, Mats; Akin, James W.; Curry, Thomas E.

2017-01-01

Context: In animal models, the luteinizing hormone surge increases progesterone (P4) and progesterone receptor (PGR), prostaglandins (PTGs), and epidermal growth factor (EGF)–like factors that play essential roles in ovulation. However, little is known about the expression, regulation, and function of these key ovulatory mediators in humans. Objective: To determine when and how these key ovulatory mediators are induced after the luteinizing hormone surge in human ovaries. Design and Participants: Timed periovulatory follicles were obtained from cycling women. Granulosa/lutein cells were collected from in vitro fertilization patients. Main Outcome Measures: The in vivo and in vitro expression of PGR, PTG synthases and transporters, and EGF-like factors were examined at the level of messenger RNA and protein. PGR binding to specific genes was assessed. P4 and PTGs in conditioned media were measured. Results: PGR, PTGS2, and AREG expressions dramatically increased in ovulatory follicles at 12 to 18 hours after human chorionic gonadotropin (hCG). In human granulosa/lutein cell cultures, hCG increased P4 and PTG production and the expression of PGR, specific PTG synthases and transporters, and EGF-like factors, mimicking in vivo expression patterns. Inhibitors for P4/PGR and EGF-signaling pathways reduced hCG-induced increases in PTG production and the expression of EGF-like factors. PGR bound to the PTGS2, PTGES, and SLCO2A1 genes. Conclusions: This report demonstrated the time-dependent induction of PGR, AREG, and PTGS2 in human periovulatory follicles. In vitro studies indicated that collaborative actions of P4/PGR and EGF signaling are required for hCG-induced increases in PTG production and potentiation of EGF signaling in human periovulatory granulosa cells. PMID:28323945

Highly Purified Eicosapentaenoic Acid Increases Interleukin-10 Levels of Peripheral Blood Monocytes in Obese Patients With Dyslipidemia

PubMed Central

Satoh-Asahara, Noriko; Shimatsu, Akira; Sasaki, Yousuke; Nakaoka, Hidenori; Himeno, Akihiro; Tochiya, Mayu; Kono, Shigeo; Takaya, Tomohide; Ono, Koh; Wada, Hiromichi; Suganami, Takayoshi; Hasegawa, Koji; Ogawa, Yoshihiro

2012-01-01

OBJECTIVE It has recently been highlighted that proinflammatory (M1) macrophages predominate over anti-inflammatory (M2) macrophages in obesity, thereby contributing to obesity-induced adipose inflammation and insulin resistance. A recent clinical trial revealed that highly purified eicosapentaenoic acid (EPA) reduces the incidence of major coronary events. In this study, we examined the effect of EPA on M1/M2-like phenotypes of peripheral blood monocytes in obese dyslipidemic patients. RESEARCH DESIGN AND METHODS Peripheral blood monocytes were prepared from 26 obese patients without and 90 obese patients with dyslipidemia. Of the latter 90 obese patients with dyslipidemia, 82 patients were treated with or without EPA treatment (1.8 g daily) for 3 months. RESULTS Monocytes in obese patients with dyslipidemia showed a significantly lower expression of interleukin-10 (IL-10), an M2 marker, than those without dyslipidemia. EPA significantly increased serum IL-10 and EPA levels, the EPA/arachidonic acid (AA) ratio, and monocyte IL-10 expression and decreased the pulse wave velocity (PWV), an index of arterial stiffness, compared with the control group. After EPA treatment, the serum EPA/AA ratio was significantly correlated with monocyte IL-10 expression. Only increases in monocyte IL-10 expression and serum adiponectin were independent determinants of a decreased PWV by EPA. Furthermore, EPA significantly increased the expression and secretion of IL-10 in human monocytic THP-1 cells through a peroxisome proliferator–activated receptor (PPAR)γ-dependent pathway. CONCLUSIONS This study is the first to show that EPA increases the monocyte IL-10 expression in parallel with decrease of arterial stiffness, which may contribute to the antiatherogenic effect of EPA in obese dyslipidemic patients. PMID:22912426

Macrophages of multiple sclerosis patients display deficient SHP-1 expression and enhanced inflammatory phenotype.

PubMed

Christophi, George P; Panos, Michael; Hudson, Chad A; Christophi, Rebecca L; Gruber, Ross C; Mersich, Akos T; Blystone, Scott D; Jubelt, Burk; Massa, Paul T

2009-07-01

Recent studies in mice have demonstrated that the protein tyrosine phosphatase SHP-1 is a crucial negative regulator of proinflammatory cytokine signaling, TLR signaling, and inflammatory gene expression. Furthermore, mice genetically lacking SHP-1 (me/me) display a profound susceptibility to inflammatory CNS demyelination relative to wild-type mice. In particular, SHP-1 deficiency may act predominantly in inflammatory macrophages to increase CNS demyelination as SHP-1-deficient macrophages display coexpression of inflammatory effector molecules and increased demyelinating activity in me/me mice. Recently, we reported that PBMCs of multiple sclerosis (MS) patients have a deficiency in SHP-1 expression relative to normal control subjects indicating that SHP-1 deficiency may play a similar role in MS as to that seen in mice. Therefore, it became essential to examine the specific expression and function of SHP-1 in macrophages from MS patients. Herein, we document that macrophages of MS patients have deficient SHP-1 protein and mRNA expression relative to those of normal control subjects. To examine functional consequences of the lower SHP-1, the activation of STAT6, STAT1, and NF-kappaB was quantified and macrophages of MS patients showed increased activation of these transcription factors. In accordance with this observation, several STAT6-, STAT1-, and NF-kappaB-responsive genes that mediate inflammatory demyelination were increased in macrophages of MS patients following cytokine and TLR agonist stimulation. Supporting a direct role of SHP-1 deficiency in altered macrophage function, experimental depletion of SHP-1 in normal subject macrophages resulted in an increased STAT/NF-kappaB activation and increased inflammatory gene expression to levels seen in macrophages of MS patients. In conclusion, macrophages of MS patients display a deficiency of SHP-1 expression, heightened activation of STAT6, STAT1, and NF-kappaB and a corresponding inflammatory profile that may be important in controlling macrophage-mediated demyelination in MS.

Grape seed extract has superior beneficial effects than vitamin E on oxidative stress and apoptosis in the hippocampus of streptozotocin induced diabetic rats.

PubMed

Yonguc, Goksin Nilufer; Dodurga, Yavuz; Adiguzel, Esat; Gundogdu, Gulsah; Kucukatay, Vural; Ozbal, Seda; Yilmaz, Ismail; Cankurt, Ulker; Yilmaz, Yusuf; Akdogan, Ilgaz

2015-01-25

We aimed to investigate the effects of grape seed extract (GSE) and vitamin E (Vit E) on oxidative stress and apoptosis in the hippocampus of streptozotocin-induced diabetic rats. In Control, Diabetic, and Diabetic treated with GSE (Diabetic+GSE) and vitamin E (Diabetic+Vit E) groups, oxidative stress index (OSI), TUNEL staining and Bcl-2, Bcl-XL, Bax, caspase-3, -9, and -8, Cyt-c, TNF-α, and NF-κB gene expressions were evaluated. OSI was significantly increased in the plasma and hippocampus of the Diabetic compared to Control group and decreased in Diabetic+GSE and Diabetic+Vit E groups compared to Diabetic. TUNEL positive neurons significantly increased in the hippocampus of the Diabetic group compared to Control and decreased in Diabetic+GSE (more prominently) and Diabetic+Vit E groups compared to Diabetic. In the hippocampus of the Diabetic group, Bcl-2 and Bcl-XL gene expressions were significantly decreased; Bax, caspase-3, -9, and -8, Cyt-c, TNF-α, and NF-κB gene expressions were significantly increased compared to Control. In Diabetic+GSE and Diabetic+Vit E groups, Bcl-2 gene expressions were significantly increased; Bcl-XL gene expressions did not differ compared to the Diabetic group. The expression of Bax, caspase-3, -9, and -8, Cyt-c, TNF-α, and NF-κB genes in the Diabetic+GSE group and the expression of caspase-3 and -9, TNF-α, and NF-κB genes in the Diabetic+Vit E group were significantly decreased compared to Diabetic. In conclusion, GSE (more prominently) and vitamin E decreased oxidative stress and neuronal apoptosis occurring in the hippocampus of diabetic rats. Copyright © 2014 Elsevier B.V. All rights reserved.

Complement Factor H Is Expressed in Adipose Tissue in Association With Insulin Resistance

PubMed Central

Moreno-Navarrete, José María; Martínez-Barricarte, Rubén; Catalán, Victoria; Sabater, Mònica; Gómez-Ambrosi, Javier; Ortega, Francisco José; Ricart, Wifredo; Blüher, Mathias; Frühbeck, Gema; Rodríguez de Cordoba, Santiago; Fernández-Real, José Manuel

2010-01-01

OBJECTIVE Activation of the alternative pathway of the complement system, in which factor H (fH; complement fH [CFH]) is a key regulatory component, has been suggested as a link between obesity and metabolic disorders. Our objective was to study the associations between circulating and adipose tissue gene expressions of CFH and complement factor B (fB; CFB) with obesity and insulin resistance. RESEARCH DESIGN AND METHODS Circulating fH and fB were determined by enzyme-linked immunosorbent assay in 398 subjects. CFH and CFB gene expressions were evaluated in 76 adipose tissue samples, in isolated adipocytes, and in stromovascular cells (SVC) (n = 13). The effects of weight loss and rosiglitazone were investigated in independent cohorts. RESULTS Both circulating fH and fB were associated positively with BMI, waist circumference, triglycerides, and inflammatory parameters and negatively with insulin sensitivity and HDL cholesterol. For the first time, CFH gene expression was detected in human adipose tissue (significantly increased in subcutaneous compared with omental fat). CFH gene expression in omental fat was significantly associated with insulin resistance. In contrast, CFB gene expression was significantly increased in omental fat but also in association with fasting glucose and triglycerides. The SVC fraction was responsible for these differences, although isolated adipocytes also expressed fB and fH at low levels. Both weight loss and rosiglitazone led to significantly decreased circulating fB and fH levels. CONCLUSIONS Increased circulating fH and fB concentrations in subjects with altered glucose tolerance could reflect increased SVC-induced activation of the alternative pathway of complement in omental adipose tissue linked to insulin resistance and metabolic disturbances. PMID:19833879

Filgrastim (RHG-CSF) related modulation of the inflammatory response in patients at risk of sepsis or with sepsis.

PubMed

Weiss, M; Gross-Weege, W; Harms, B; Schneider, E M

1996-03-01

Over a period of 14 days a longitudinal analysis was performed on the effects of filgrastim (recombinant human granulocyte colony stimulating factor, rhG-CSF) administered to 20 postoperative/posttraumatic patients at risk of or with sepsis. The following parameters were determined: leukocyte counts, serum cytokine levels and the surface expression of functional antigens and adhesion molecules. Filgrastim (1 mu g/kg.day) was infused continuously on the first 3 days and tapered to 0.5 mu g/kg.day on the following 4 days or until discharge from the surgical intensive care unit. During infusion of filgrastim, G-CSF levels increased in 16 out of the 20 patients within 48 h. In these 16 patients, leukocyte counts increased in 15 out of 16 patients. Expression of CD64 was upregulated within 24 h. The expression of CD32 was upregulated in 8 out of 9 patients with an initial expression < 55%. LAM-1 expression was downregulated in all patients revealing an initial expression of LAM-1 > 40%. Soluble ICAM increased in 9 out of 11 patients. IL-8 decreased in all 6 patients presenting initial values of IL-8 > 90 pg/ml. IL-1RA increased in 10 patients. Filgrastim had no effect on the expression of CD14, CD16 and CD34 and on the levels of TNF-alpha and sTNF-R type I (p55). In conclusion, infusion of filgrastim in postoperative/post traumatic patients at risk of and with sepsis resulted in improved generation and function of neutrophils and appeared to counterregulate hyperactivation of proinflammatory processes.

hnRNP K plays a protective role in TNF-α-induced apoptosis in podocytes.

PubMed

Zhao, Shili; Feng, Junxia; Wang, Qi; Tian, Lu; Zhang, Yunfang; Li, Hongyan

2018-06-29

Apoptosis of podocytes contributes to proteinuria in many chronic kidney diseases. The cytokine, tumor necrosis factor-α (TNF-α) is thought to be involved in podocyte apoptosis, but the underlying mechanism is not understood. In our study, we established a model of TNF-α-induced apoptosis by isolating primary podocytes from mice. After exposing cells to TNF-α, we determined the expression levels of heterogeneous nuclear ribonucleoprotein K (hnRNP K) and cellular FLICE-inhibitory protein (c-FLIP) and the phosphorylation levels of glycogen synthase kinase β (GSK3β) and extracellular signal-regulated kinase (ERK). We then knocked down or overexpressed the levels of hnRNP K and observed its effects on the expressions of c-FLIP, caspase-8, caspase-3, and the phosphorylation of GSK3β and ERK. In addition, we examined the percentage of cells undergoing apoptosis and studied cell cycle distribution. We found that TNF-α induced apoptosis in podocytes and that the expressions of hnRNP K and c-FLIP were significantly decreased, whereas the phosphorylations of GSK3β and ERK were significantly increased. Both gene knockdown and overexpression of hnRPN K resulted in varied expressions/phosphorylations of c-FLIP, GSK3β, and ERK. Moreover, decreased hnRPN K expression contributed to increased levels of caspase-8 and capase-3, as well as an increase in cell apoptosis and G0/G1 arrest. In conclusion, down-regulated expression of hnRNP K by TNF-α resulted in a decrease in the expression of c-FLIP as well as increases in phosphorylated GSK3β, ERK, caspase-8, and caspase-3, and then critically contributed to the podocyte apoptosis. © 2018 The Author(s).

A HER2-targeting antibody-drug conjugate, trastuzumab deruxtecan (DS-8201a), enhances antitumor immunity in a mouse model.

PubMed

Iwata, Tomomi Nakayama; Ishii, Chiaki; Ishida, Saori; Ogitani, Yusuke; Wada, Teiji; Agatsuma, Toshinori

2018-04-27

Trastuzumab deruxtecan (DS-8201a), a HER2-targeting antibody-drug conjugate with a topoisomerase I inhibitor exatecan derivative (DX-8951 derivative, DXd), has been reported to exert potent antitumor effects in xenograft mouse models and clinical trials. In this study, the immune system-activating ability of DS-8201a was assessed. DS-8201a significantly suppressed tumor growth in an immunocompetent mouse model with human HER2-expressing CT26.WT (CT26.WT-hHER2) cells. Cured immunocompetent mice rejected not only re-challenged CT26.WT-hHER2 cells, but also CT26.WT-mock cells. Splenocytes from the cured mice responded to both CT26.WT-hHER2 and CT26.WT-mock cells. Further analyses revealed that DXd up-regulated CD86 expression on bone marrow-derived DCs in vitro, and that DS-8201a increased tumor-infiltrating DCs and up-regulated their CD86 expression in vivo. DS-8201a also increased tumor-infiltrating CD8+ T cells and enhanced PD-L1 and MHC class I expression on tumor cells. Furthermore, combination therapy with DS-8201a and anti-PD-1 antibody was more effective than either monotherapy. In conclusion, DS-8201a enhanced antitumor immunity, as evidenced by the increased expression of DC markers, augmented expression of MHC class I in tumor cells, and rejection of re-challenged tumor cells by adaptive immune cells, suggesting that DS-8201a enhanced tumor recognition by T cells. Furthermore, DS-8201a treatment benefited from combination with anti-PD-1 antibody, possibly due to increased T cell activity and up-regulated PD-L1 expression induced by DS-8201a. Copyright ©2018, American Association for Cancer Research.

Smad, PI3K/Akt, and Wnt-dependent signaling pathways are involved in BMP-4-induced ESC self-renewal.

PubMed

Lee, Min Young; Lim, Hyun Woo; Lee, Sang Hun; Han, Ho Jae

2009-08-01

It is known that bone morphogenetic protein 4 (BMP-4) has a diverse effect on ESCs. However, its precise mechanism in mouse ESCs is not fully understood. We evaluated the effect of BMP-4 on ESC proliferation and its related signal cascades in this study. BMP-4 significantly increased the level of [(3)H]-thymidine incorporation in time- (> or =8 hours) and dose- (> or =10 ng/ml) dependent manners. Additionally, BMP-4 increased cyclin D1 and decreased p27(kip1) expression values in a time-dependent manner. The increases in BMP-4-induced [(3)H]-thymidine incorporation and cyclin D1 expression were inhibited by the BMP-4 receptor antagonist noggin. BMP-4 increased Wnt1 expression. Wnt1 expression was attenuated by Smad4 small interfering RNA (siRNA), and BMP-4-induced cyclin D1 expression was inhibited by Smad4 and Wnt1 siRNAs. BMP-4 also activated beta-catenin, which was blocked by Smad4 and Wnt1 siRNAs. In addition, BMP-4 induced Akt phosphorylation. BMP-4-induced beta-catenin activation and cyclin D1 expression were attenuated by phosphatidyl inositol 3-kinase (PI3K) siRNA and Akt inhibitor. Additionally, downregulation of Smad4, Wnt1, and PI3K expression by siRNA decreased the levels of pluripotency marker mRNAs of ESCs, including Oct4, Sox2, and FoxD3. Our results suggested that BMP-4-induced [(3)H]-thymidine incorporation was significantly attenuated by Smad4, Wnt1, and PI3K knockdown. In conclusion, BMP-4 contributed to the maintenance of cell proliferation and the pluripotent state by Smad, PI3K/Akt, and Wnt1/beta-catenin in mouse ESCs.

Quercetin attenuates toosendanin-induced hepatotoxicity through inducing the Nrf2/GCL/GSH antioxidant signaling pathway.

PubMed

Jin, Yao; Huang, Zhen-Lin; Li, Li; Yang, Yang; Wang, Chang-Hong; Wang, Zheng-Tao; Ji, Li-Li

2018-06-19

Toosendanin (TSN) is the main active compound in Toosendan Fructus and Meliae Cortex, two commonly used traditional Chinese medicines. TSN has been reported to induce hepatotoxicity, but its mechanism remains unclear. In this study, we demonstrated the critical role of nuclear factor erythroid 2-related factor 2 (Nrf2) in protecting against TSN-induced hepatotoxicity in mice and human normal liver L-02 cells. In mice, administration of TSN (10 mg/kg)-induced acute liver injury evidenced by increased serum alanine/aspartate aminotransferase (ALT/AST) and alkaline phosphatase (ALP) activities, and total bilirubin (TBiL) content as well as the histological changes. Furthermore, TSN markedly increased liver reactive oxygen species (ROS) and malondialdehyde (MDA) levels, and decreased liver glutathione (GSH) content and Nrf2 expression. In L-02 cells, TSN (2 μM) time-dependently reduced glutamate-cysteine ligase (GCL) activity and cellular expression of the catalytic/modify subunit of GCL (GCLC/GCLM). Moreover, TSN reduced cellular GSH content and the increased ROS formation, and time-dependently decreased Nrf2 expression and increased the expression of the Nrf2 inhibitor protein kelch-like ECH-associated protein-1 (Keap1). Pre-administration of quercetin (40, 80 mg/kg) effectively inhibited TSN-induced liver oxidative injury and reversed the decreased expression of Nrf2 and GCLC/GCLM in vivo and in vitro. In addition, the quercetin-provided protection against TSN-induced hepatotoxicity was diminished in Nrf2 knock-out mice. In conclusion, TSN decreases cellular GSH content by reducing Nrf2-mediated GCLC/GCLM expression via decreasing Nrf2 expression. Quercetin attenuates TSN-induced hepatotoxicity by inducing the Nrf2/GCL/GSH antioxidant signaling pathway. This study implies that inducing Nrf2 activation may be an effective strategy to prevent TSN-induced hepatotoxicity.

Reduced Expression of Adipose Triglyceride Lipase Enhances Tumor Necrosis Factor α-induced Intercellular Adhesion Molecule-1 Expression in Human Aortic Endothelial Cells via Protein Kinase C-dependent Activation of Nuclear Factor-κB*

PubMed Central

Inoue, Tomoaki; Kobayashi, Kunihisa; Inoguchi, Toyoshi; Sonoda, Noriyuki; Fujii, Masakazu; Maeda, Yasutaka; Fujimura, Yoshinori; Miura, Daisuke; Hirano, Ken-ichi; Takayanagi, Ryoichi

2011-01-01

We examined the effects of adipose triglyceride lipase (ATGL) on the initiation of atherosclerosis. ATGL was recently identified as a rate-limiting triglyceride (TG) lipase. Mutations in the human ATGL gene are associated with neutral lipid storage disease with myopathy, a rare genetic disease characterized by excessive accumulation of TG in multiple tissues. The cardiac phenotype, known as triglyceride deposit cardiomyovasculopathy, shows massive TG accumulation in both coronary atherosclerotic lesions and the myocardium. Recent reports show that myocardial triglyceride content is significantly higher in patients with prediabetes or diabetes and that ATGL expression is decreased in the obese insulin-resistant state. Therefore, we investigated the effect of decreased ATGL activity on the development of atherosclerosis using human aortic endothelial cells. We found that ATGL knockdown enhanced monocyte adhesion via increased expression of TNFα-induced intercellular adhesion molecule-1 (ICAM-1). Next, we determined the pathways (MAPK, PKC, or NFκB) involved in ICAM-1 up-regulation induced by ATGL knockdown. Both phosphorylation of PKC and degradation of IκBα were increased in ATGL knockdown human aortic endothelial cells. In addition, intracellular diacylglycerol levels and free fatty acid uptake via CD36 were significantly increased in these cells. Inhibition of the PKC pathway using calphostin C and GF109203X suppressed TNFα-induced ICAM-1 expression. In conclusion, we showed that ATGL knockdown increased monocyte adhesion to the endothelium through enhanced TNFα-induced ICAM-1 expression via activation of NFκB and PKC. These results suggest that reduced ATGL expression may influence the atherogenic process in neutral lipid storage diseases and in the insulin-resistant state. PMID:21828047

Effect of heat stress and recovery on viability, oxidative damage, and heat shock protein expression in hepatic cells of grass carp (Ctenopharyngodon idellus).

PubMed

Cui, Yanting; Liu, Bo; Xie, Jun; Xu, Pao; Habte-Tsion, H-Michael; Zhang, Yuanyuan

2014-06-01

In this study, we investigated the effects of hyperthermia and recovery on cell viability, lactate dehydrogenase (LDH) activity, superoxide dismutase (SOD) activity, malondialdehyde (MDA), total antioxidant capacity (T-AOC), and heat shock protein (HSP60, 70, and 90) mRNA expression in the hepatic cells of the grass carp, Ctenopharyngodon idellus. Triplicate groups of cultured cells were exposed to 30, 32, or 34 °C for 0.5 h and then immediately incubated at 27 °C in 5 % CO2 for 6, 12, 24, or 48 h. Hyperthermia stress greatly reduced cell viability and increased LDH release. Cell damage declined after recovery. Hyperthermia stress increased the lipid peroxide levels and reduced the antioxidant capacity (e.g., reduced SOD and T-AOC) of the cells. However, oxidative damage declined as the recovery period increased, and the levels of MDA, SOD, and T-AOC were restored. After cells were exposed to 32 °C, the expression of HSP60 after recovery for 1, 2, and 4 h (P < 0.05), the expression of HSP70 after recovery for 0.5 and 1 h (P < 0.01), and the expression of HSP90 throughout recovery were significantly higher (P < 0.01) than the prestress levels. During the recovery period, the variations in HSP gene expression reflected the transition period from a state of cellular growth to one of the cellular repairs. In conclusion, hyperthermia depresses cell viability, induces oxidative damage, and increases HSP expression, which plays an important role during hyperthermic stress in grass carp hepatic cells.

High glucose impaired estrogen receptor alpha signaling via β-catenin in osteoblastic MC3T3-E1.

PubMed

Wang, Rui; Gao, Dong; Zhou, Yin; Chen, Lu; Luo, Bin; Yu, Yanrong; Li, Hao; Hu, Jiawei; Huang, Qiren; He, Ming; Peng, Weijie; Luo, Dan

2017-11-01

Diabetic Mellitus is a risk factor for osteoporosis. It has been suggested that altered estrogen or estrogen receptor α/β (ERα/β) signaling may be involved in diabetic osteoporosis. The present study is to investigate the effects of high glucose on ERα/β signaling in osteoblastic MC3T3-E1 and how the altered signaling of ERα/β affect osteoblastic bone formation. ERα/β signaling was demonstrated as ERα/β protein expression (Western Blotting) and ER transcription activity (Luciferase Reporter assays). Proliferation (WSK-1 assaying), differentiation (ALP staining) and mineralization (Alizalard Red staining) of MC3T3-E1 were examined to evaluate bone formation function. It has been found that high glucose increased ERα/β expression dose-dependently and time-dependently, but high glucose (33mM) decreased ERα transcription activity. 17β-estradiol increased the ERα/β expression dose-dependently in normal medium, but decreased the ERα/β expression dose-dependently in medium with high glucose (33mM). High glucose decreased bone formation and also decreased the osteogenic effects of 17β-estradiol (10 -8 M). High glucose decreased β-catenin expression dose-dependently and time-dependently. LiCl, an inhibitor of β-catenin degradation, decreased ERα expression but increased ERα transcription activity. When compared with high glucose treatment, LiCl (5mM) increased ALP activity and calcified nodes. Besides, high glucose also decreased the protein expression PI-3K, pAKT/AKT, GSK-3β. In conclusion, the present study suggested that high glucose may impair ERα transcription activity by inhibiting β-catenin signaling in osteoblastic MC3T3-E1, leading decreased bone formation ligand-dependently or ligand-independently. Copyright © 2017 Elsevier Ltd. All rights reserved.

Maternal Obesity Induces Sustained Inflammation in Both Fetal and Offspring Large Intestine of Sheep

PubMed Central

Yan, Xu; Huang, Yan; Wang, Hui; Du, Min; Hess, Bret W.; Ford, Stephen P.; Nathanielsz, Peter W.; Zhu, Mei-Jun

2010-01-01

Background Both maternal obesity and inflammatory bowel diseases (IBDs) are increasing. It was hypothesized that maternal obesity induces an inflammatory response in the fetal large intestine, predisposing offspring to IBDs. Methods Nonpregnant ewes were assigned to a control (Con, 100% of National Research Council [NRC] recommendations) or obesogenic (OB, 150% of NRC) diet from 60 days before conception. The large intestine was sampled from fetuses at 135 days (term 150 days) after conception and from offspring lambs at 22.5 ± 0.5 months of age. Results Maternal obesity enhanced mRNA expression tumor necrosis factor (TNF)α, interleukin (IL)1α, IL1β, IL6, IL8, and monocyte/macrophage chemotactic protein-1 (MCP1), as well as macrophage markers, CD11b, CD14, and CD68 in fetal gut. mRNA expression of Toll-like receptor (TLR) 2 and TLR4 was increased in OB versus Con fetuses; correspondingly, inflammatory NF-κB and JNK signaling pathways were also upregulated. Both mRNA expression and protein content of transforming growth factor (TGF) β was increased. The IL-17A mRNA expression and protein content was higher in OB compared to Con samples, which was associated with fibrosis in the large intestine of OB fetuses. Similar inflammatory responses and enhanced fibrosis were detected in OB compared to Con offspring. Conclusions Maternal obesity induced inflammation and enhanced expression of proinflammatory cytokines in fetal and offspring large intestine, which correlated with increased TGFβ and IL17 expression. These data show that maternal obesity may predispose offspring gut to IBDs. PMID:21674707

Aging and a long-term diabetes mellitus increase expression of 1 α-hydroxylase and vitamin D receptors in the rat liver.

PubMed

Vuica, Ana; Ferhatović Hamzić, Lejla; Vukojević, Katarina; Jerić, Milka; Puljak, Livia; Grković, Ivica; Filipović, Natalija

2015-12-01

Diabetes mellitus (DM) is a metabolic disorder associated with serious liver complications. As a metabolic chronic disease, DM is very common in the elderly. Recent studies suggest ameliorating effects of vitamin D on metabolic and oxidative stress in the liver tissue in an experimental model of DM. The aim of this study was to investigate the expression of vitamin D receptors (VDRs) and 1α-hydroxylase, the key enzyme for the production of active vitamin D form (calcitriol) in the liver during long-term diabetes mellitus type 1 (DM1) in aging rats. We performed immunohistochemical analysis of liver expression of 1α-hydroxylase and VDRs during aging in long-term streptozotocin-induced DM1. 1α-Hydroxylase was identified in the monocyte/macrophage system of the liver. In addition to the nuclear expression, we also observed the expression of VDR in membranes of lipid droplets within hepatocytes. Aging and long-term DM1 resulted in significant increases in the number of 1α-hydroxylase immunoreactive cells, as well as the percentage of strongly positive VDR hepatocytes. In conclusion, the liver has the capacity for active vitamin D synthesis in its monocyte/macrophage system that is substantially increased in aging and long-term diabetes mellitus. These conditions are also characterized by significant increases in vitamin D receptor expression in hepatocytes. The present study suggests that VDR signaling system could be a potential target in prevention of liver complications caused by diabetes and aging. Copyright © 2015 Elsevier Inc. All rights reserved.

ZEB1 is Estrogen Responsive In Vitro in Human Foreskin Cells and is Over Expressed in Penile Skin in Patients With Severe Hypospadias

PubMed Central

Qiao, Liang; Tasian, Gregory E.; Zhang, Haiyang; Cunha, Gerald R.; Baskin, Laurence

2012-01-01

Purpose We determined the effect of estrogen on ZEB1 in vitro and tested the hypothesis that ZEB1 is over expressed in the penile skin of subjects with hypospadias. Materials and Methods Hs68 cells, a fibroblast cell line derived from human foreskin, were exposed to 0, 1, 10 and 100 nM estrogen, and the expression level of ZEB1 was assessed using reverse transcription real-time polymerase chain reaction, Western blot and immunocytochemical analysis. Next, preputial skin was prospectively collected from case and control subjects at hypospadias repair (37 cases) and circumcision (11). Hypospadias was classified as severe (13 cases) or mild (24) based on the position of the urethral meatus. ZEB1 expression was quantified using reverse transcription real-time polymerase chain reaction, Western blot and immunohistochemical analysis. Results Estrogen increased ZEB1 expression at the mRNA and protein levels in Hs68 cells in a concentration dependent fashion (p <0.01). Subjects with severe hypospadias had significantly higher ZEB1 mRNA levels and protein expression compared to controls or subjects with mild hypospadias (both p <0.01). Subjects with severe hypospadias had increased expression of ZEB1 in the basal layers of the preputial epidermis. Conclusions Estrogen increases ZEB1 expression in a human foreskin fibroblast cell line in vitro. Furthermore, ZEB1 is significantly over expressed in the penile skin of subjects with severe hypospadias. We propose that ZEB1 overexpression may contribute to development of hypospadias and may mediate the effect of estrogen on developing external male genitalia. PMID:21421232

OmpR and RcsB abolish temporal and spatial changes in expression of flhD in Escherichia coli biofilm.

PubMed

Samanta, Priyankar; Clark, Emily R; Knutson, Katie; Horne, Shelley M; Prüß, Birgit M

2013-08-02

Biofilms are communities of bacteria that are characterized by specific phenotypes, including an increased resistance towards anti-microbials and the host immune system. This calls for the development of novel biofilm prevention and treatment options to combat infectious disease. In Escherichia coli, numerous global regulators have been implicated in the control of biofilm associated cell surface organelles. These include the flagellar regulator FlhD/FlhC, the osmoregulator EnvZ/OmpR, and the colanic acid activator RcsCDB. Using flow cell technology and fluorescence microscopy, we determined the temporal expression from flhD::gfp, ompR::gfp, and rcsB::gfp in E. coli biofilm, as well as the impact of the negative regulation of flhD by OmpR and RcsB. Spatial gene expression was investigated from flhD::gfp. The temporal gene expression profile for flhD yielded an early peak at 12 h, a minimum of expression at 35 h, and a second increase in expression towards 51 h of biofilm development. In contrast, the ompR profile showed a peak at 35 h. A mutation in ompR abolished time dependence of flhD expression after the initial growth period of 12 h. Intriguingly, rcsB expression did not correlate inversely with flhD expression, yet a mutation in rcsB abolished time dependence of flhD expression as well. Spatially, expression of flhD was highest in the outermost layer of the biofilm in the parent strain. In ompR and rcsB mutants, flhD was expressed throughout the biofilm. Mutations in both, ompR and rcsB increased flhD expression throughout all temporal and spatial experiments. This increase was paralleled by reductions in biofilm amounts at four tested time points. Our data lead to the conclusion that FlhD/FlhC and its regulation by OmpR and RcsB may be our first target mechanism for the development of novel biofilm prevention and treatment techniques.

Induction of Cyclooxygenase-2 Expression by Hepatitis B Virus Depends on Demethylation-associated Recruitment of Transcription Factors to the Promoter

PubMed Central

2011-01-01

Background The hepatitis B virus (HBV) is a major etiological factor of inflammation and damage to the liver resulting in hepatocellular carcinoma. Transcription factors play important roles in the disordered gene expression and liver injury caused by HBV. However, the molecular mechanisms behind this observation have not been defined. Results In this study, we observed that circulating prostaglandin (PGE) 2 synthesis was increased in patients with chronic hepatitis B infection, and detected elevated cyclooxygenase (COX)-2 expression in HBV- and HBx-expressing liver cells. Likewise, the association of HBx with C/EBPβ contributed to the induction of COX-2. The COX-2 promoter was hypomethylated in HBV-positive cells, and specific demethylation of CpG dinucleotides within each of the two NF-AT sites in the COX-2 promoter resulted in the increased binding affinity of NF-AT to the cognate sites in the promoter, followed by increased COX-2 expression and PGE2 accumulation. The DNA methylatransferase DNMT3B played a key role in the methylation of the COX-2 promoter, and its decreased binding to the promoter was responsible for the regional demethylation of CpG sites, and for the increased binding of transcription factors in HBV-positive cells. Conclusion Our results indicate that upregulation of COX-2 by HBV and HBx is mediated by both demethylation events and recruitment of multiple transcription factors binding to the promoter. PMID:21401943

Evaluation of the effect of follicular stimulating hormone on the in vitro bovine spermatogonial stem cells self-renewal: An experimental study

PubMed Central

Jabarpour, Masoome; Tajik, Parviz

2017-01-01

Background: Spermatogonial stem cells (SSCs) are undifferentiated cells which are highly reproducible and expandable. Several studies have been conducted to reproduce these cells in culture. They used growth factors, hormones and different feeder cells to improve survival and proliferation of SSCs. Objective: This study was conducted to evaluate the effects of follicular stimulating hormone (FSH) on gene expression of fibroblast growth factor (FGF2) and glial cell-derived neurotrophic factor (GDNF) in Sertoli cells. Materials and Methods: Sertoli cells and SSCs were isolated from 3-5 month-old calves. Bovine testicular cells were cultured for 15 days with or without FSH. Identification of these cells was confirmed by immunocytochemistry analysis. Colony formation of SSCs was evaluated using an inverted microscope. The gene expression of FGF2 and GDNF and the gene markers bcl6b, thy-1, and C-kit were evaluated using the quantitative RT-PCR technique. Results: The results indicated that FSH increased colonization of SSCs. the expression of GDNF, FGF2, and markers of undifferentiated spermatogonia was increased following culture in control and FSH groups (p<0.05), this increase was more in FSH group. Conversely, the expression of C-kit was decreased in both groups (p<0.05). Conclusion: The results showed that FSH can increase the self-renewal of SSCs in vitro via upregulation of GDNF and FGF2 expression in Sertoli cells. PMID:29492477

Inflammation, Impaired Motility, and Permeability in a Guinea Pig Model of Postoperative Ileus

PubMed Central

Lee, Yoo Jin; Hussain, Zahid; Huh, Cheal Wung; Lee, Young Ju; Park, Hyojin

2018-01-01

Background/Aims Postoperative ileus (POI) is characterized by impaired propulsive function of the gastrointestinal tract after surgery. Although inflammation is considered to be an important pathogenesis of POI, significant data are lacking. We aim to correlate the recovery time of postoperative dysmotility with that of inflammation and mucosal permeability. Methods An experimental POI model of guinea pig was used. Contractile activity of the circular muscle of the stomach, jejunum, ileum, and proximal colon was measured through a tissue bath study. Inflammatory cells were counted, and the expression of calprotectin and tryptase were analyzed. The expression of protease-activated receptor 2 (PAR-2), claudin-1, and claudin-2 were analyzed with immunofluorescence. Results The small bowel and colon showed decreased contractile amplitude in the POI groups compared to control. In contrast to the colon, the contractile amplitude of the small bowel significantly recovered in the POI group at 6 hours after the operation compared to the control group. Inflammation was highly significant in the POI groups compared to the control and sham groups, especially in the colon. Immunofluorescence showed increased PAR-2 expression in the POI groups compared to sham. The decreased claudin-1 expression and increased claudin-2 expression may suggest increased mucosal permeability of the small bowel and colon in the POI groups. Conclusions Increased inflammation and mucosal permeability may play an important role in the differential recovery stages in POI. These data may provide further insights into the pathophysiology and potential new therapeutic prospects of POI. PMID:29291615

A quality improvement project to improve the rate of early breast milk expression in mothers of preterm infants.

PubMed

Murphy, Lindsey; Warner, Diane D; Parks, Jessica; Whitt, Jenny; Peter-Wohl, Sigal

2014-11-01

Providing breast milk is challenging for non-nursing mothers of premature infants. Early breast milk expression results in successful and longer lactation in mothers of very low birth weight (VLBW) infants. This quality improvement initiative sought to increase the rate of early milk expression in mothers of VLBW infants and increase the proportion of infants receiving maternal breast milk (MBM) at 28 days of age and at discharge. Phase 1 (n = 45) occurred between April 1, 2012, and August 31, 2012. Phase 2 (n = 58) occurred between September 1, 2012, and February 28, 2013. Pre-phase 2 actions included increased lactation consultant workforce, early lactation consultation, tracking of MBM supply, and physician education. Inborn infants < 1500 grams were eligible. Primary outcomes were the time of first maternal milk expression (TFME) and infant feeding type at 28 days of age and at discharge. The median TFME decreased from 9 (25th, 75th percentile; 6, 16) hours to 6 (5, 11) hours after implementation (P = .06). The proportion of infants receiving exclusive MBM at 28 days and at discharge was 64% and 74%, respectively (P = .40), and the proportion of infants receiving exclusive MBM at discharge increased from 37% to 59% (P = .046). In conclusion, a multidisciplinary initiative aimed at improving the rate of early milk expression was associated with more VLBW infants receiving exclusive MBM at discharge. © The Author(s) 2014.

Induction of extracellular ATP mediates increase in intracellular thioredoxin in RAW264.7 cells exposed to low-dose γ-rays.

PubMed

Ohshima, Yasuhiro; Kitami, Akihiro; Kawano, Ayumi; Tsukimoto, Mitsutoshi; Kojima, Shuji

2011-09-15

We previously showed that low doses (0.25-0.5 Gy) of γ-rays elevated thioredoxin (Trx-1) in various organs of mice after whole-body irradiation. Also, it is reported that extracellular ATP, which is released in response to various stresses, regulates the expression of intracellular antioxidants through activation of P2 receptors. We have recently found that low-dose γ-rays induce ATP release from the exposed cells. However, it is not yet clear whether the radiation-induced extracellular ATP modulates the cellular redox balance. Here, we investigated whether γ-ray irradiation-induced release of extracellular ATP contributes to the induction of the cellular antioxidant Trx-1, using mouse macrophage-like RAW264.7 cells. Irradiation with γ-rays or exogenously added ATP increased the expression of Trx-1, and in both cases the increase was blocked by pretreatment with an ectonucleotidase, apyrase. Then, the involvement of ATP-dependent reactive oxygen species (ROS) generation in the increase in antioxidant capacity was examined. ATP stimulation promoted the generation of intracellular ROS and also increased Trx-1 expression. The increase in Trx-1 expression was significantly suppressed by pretreatment of the cells with antioxidants. In conclusion, the γ-ray irradiation-induced release of extracellular ATP may, at least in part, contribute to the production of ROS via purinergic signaling, leading to promotion of intracellular antioxidants as an adaptive response to an oxidative stress. Copyright © 2011 Elsevier Inc. All rights reserved.

Research in Undergraduate Instruction: A Biotech Lab Project for Recombinant DNA Protein Expression in Bacteria

NASA Astrophysics Data System (ADS)

Brockman, Mark; Ordman, Alfred B.; Campbell, A. Malcolm

1996-06-01

In the sophomore-level Molecular Biology and Biotechnology course at Beloit College, students learn basic methods in molecular biology in the context of pursuing a semester-long original research project. We are exploring how DNA sequence affects expression levels of proteins. A DNA fragment encoding all or part of the guanylate monokinase (gmk) sequence is cloned into pSP73 and expressed in E. coli. A monoclonal antibody is made to gmk. The expression level of gmk is determined by SDS gel elctrophoresis, a Western blot, and an ELISA assay. Over four years, an increase in enrollment in the course from 9 to 34 students, the 85% of majors pursuing advanced degrees, and course evaluations all support the conclusion that involving students in research during undergraduate courses encourages them to pursue careers in science.

DREAM Mediated Regulation of GCM1 in the Human Placental Trophoblast

PubMed Central

Baczyk, Dora; Kibschull, Mark; Mellstrom, Britt; Levytska, Khrystyna; Rivas, Marcos; Drewlo, Sascha; Lye, Stephen J.; Naranjo, Jose R.; Kingdom, John C. P.

2013-01-01

The trophoblast transcription factor glial cell missing-1 (GCM1) regulates differentiation of placental cytotrophoblasts into the syncytiotrophoblast layer in contact with maternal blood. Reduced placental expression of GCM1 and abnormal syncytiotrophoblast structure are features of hypertensive disorder of pregnancy – preeclampsia. In-silico techniques identified the calcium-regulated transcriptional repressor – DREAM (Downstream Regulatory Element Antagonist Modulator) - as a candidate for GCM1 gene expression. Our objective was to determine if DREAM represses GCM1 regulated syncytiotrophoblast formation. EMSA and ChIP assays revealed a direct interaction between DREAM and the GCM1 promoter. siRNA-mediated DREAM silencing in cell culture and placental explant models significantly up-regulated GCM1 expression and reduced cytotrophoblast proliferation. DREAM calcium dependency was verified using ionomycin. Furthermore, the increased DREAM protein expression in preeclamptic placental villi was predominantly nuclear, coinciding with an overall increase in sumolylated DREAM and correlating inversely with GCM1 levels. In conclusion, our data reveal a calcium-regulated pathway whereby GCM1-directed villous trophoblast differentiation is repressed by DREAM. This pathway may be relevant to disease prevention via calcium-supplementation. PMID:23300953

Proteomic profiling of proteins associated with the rejuvenation of Sequoia sempervirens (D. Don) Endl

PubMed Central

2010-01-01

Background Restoration of rooting competence is important for rejuvenation in Sequoia sempervirens (D. Don) Endl and is achieved by repeatedly grafting Sequoia shoots after 16 and 30 years of cultivation in vitro. Results Mass spectrometry-based proteomic analysis revealed three proteins that differentially accumulated in different rejuvenation stages, including oxygen-evolving enhancer protein 2 (OEE2), glycine-rich RNA-binding protein (RNP), and a thaumatin-like protein. OEE2 was found to be phosphorylated and a phosphopeptide (YEDNFDGNSNVSVMVpTPpTDK) was identified. Specifically, the protein levels of OEE2 increased as a result of grafting and displayed a higher abundance in plants during the juvenile and rejuvenated stages. Additionally, SsOEE2 displayed the highest expression levels in Sequoia shoots during the juvenile stage and less expression during the adult stage. The expression levels also steadily increased during grafting. Conclusion Our results indicate a positive correlation between the gene and protein expression patterns of SsOEE2 and the rejuvenation process, suggesting that this gene is involved in the rejuvenation of Sequoia sempervirens. PMID:21143964

Phlebotomy-induced anemia alters hippocampal neurochemistry in neonatal mice

PubMed Central

Wallin, Diana J.; Tkac, Ivan; Stucker, Sara; Ennis, Kathleen M.; Sola-Visner, Martha; Rao, Raghavendra; Georgieff, Michael K.

2015-01-01

Background Phlebotomy-induced anemia (PIA) is common in preterm infants. The hippocampus undergoes rapid differentiation during late fetal/early neonatal life and relies on adequate oxygen and iron to support oxidative metabolism necessary for development. Anemia shortchanges these two critical substrates, potentially altering hippocampal development and function. Methods PIA (hematocrit <25%) was induced in neonatal mice pups from postnatal day (P)3 to P14. Neurochemical concentrations in the hippocampus were determined using in vivo 1H NMR spectroscopy at 9.4T and compared with control animals at P14. Gene expression was assessed using qRT-PCR. Results PIA decreased brain iron concentration, increased hippocampal lactate and creatine concentrations, and decreased phosphoethanolamine (PE) concentration and the phosphocreatine/creatine ratio. Hippocampal transferrin receptor (Tfrc) gene expression was increased, while the expression of calcium/calmodulin-dependent protein kinase type II alpha (CamKIIα) was decreased in PIA mice. Conclusion This clinically relevant model of neonatal anemia alters hippocampal energy and phospholipid metabolism and gene expression during a critical developmental period. Low target hematocrits for preterm neonates in the NICU may have potential adverse neural implications. PMID:25734245

The effect of sebocytes cultured from nevus sebaceus on hair growth.

PubMed

Lee, Weon Ju; Cha, Hyun Wuk; Lim, Hyun Jung; Lee, Seok-Jong; Kim, Do Won

2012-10-01

Sebaceous glands are known to affect hair growth. Nevus sebaceus, a sebaceous gland hamartomas, presents as hairless patches. In this study, cultures of nevus sebaceus sebocytes (NSS) and normal scalp hair follicle sebocytes (NS) were used in performance of microarray, RT-PCR, western blot assay and immunofluorescence staining. NSS- and NS-conditioned media were also added to the culture of outer root sheath cells (ORSCs), dermal papilla cells (DPCs) or normal scalp hair follicle sebocytes. Results of this study showed a decrease in the survival rate of ORSCs and DPCs and hair growth in the NSS-conditioned medium-treated group, compared with the control and NS-conditioned medium-treated groups. An increase in expression of fibroblast growth factor (FGF)-5, Dickkopf-1 and inflammatory cytokines and a decrease in expression of Wnt10b and Lef1 were observed. In conclusion, NSS showed an increase in expression of hair growth-suppressing bioactive factors, including FGF-5, and a decrease in expression of hair growth-stimulating factors. © 2012 John Wiley & Sons A/S.

Seven mutations in the human insulin gene linked to permanent neonatal/infancy-onset diabetes mellitus

PubMed Central

Colombo, Carlo; Porzio, Ottavia; Liu, Ming; Massa, Ornella; Vasta, Mario; Salardi, Silvana; Beccaria, Luciano; Monciotti, Carla; Toni, Sonia; Pedersen, Oluf; Hansen, Torben; Federici, Luca; Pesavento, Roberta; Cadario, Francesco; Federici, Giorgio; Ghirri, Paolo; Arvan, Peter; Iafusco, Dario; Barbetti, Fabrizio

2008-01-01

Permanent neonatal diabetes mellitus (PNDM) is a rare disorder usually presenting within 6 months of birth. Although several genes have been linked to this disorder, in almost half the cases documented in Italy, the genetic cause remains unknown. Because the Akita mouse bearing a mutation in the Ins2 gene exhibits PNDM associated with pancreatic β cell apoptosis, we sequenced the human insulin gene in PNDM subjects with unidentified mutations. We discovered 7 heterozygous mutations in 10 unrelated probands. In 8 of these patients, insulin secretion was detectable at diabetes onset, but rapidly declined over time. When these mutant proinsulins were expressed in HEK293 cells, we observed defects in insulin protein folding and secretion. In these experiments, expression of the mutant proinsulins was also associated with increased Grp78 protein expression and XBP1 mRNA splicing, 2 markers of endoplasmic reticulum stress, and with increased apoptosis. Similarly transfected INS-1E insulinoma cells had diminished viability compared with those expressing WT proinsulin. In conclusion, we find that mutations in the insulin gene that promote proinsulin misfolding may cause PNDM. PMID:18451997

Transcription elongation factors are involved in programming hormone production in pituitary neuroendocrine GH4C1 cells.

PubMed

Fujita, Toshitsugu; Piuz, Isabelle; Schlegel, Werner

2010-05-05

Transcription elongation of many eukaryotic genes is regulated. Two negative transcription elongation factors, 5,6-dichloro-1-beta-D-ribofuranosylbenzimidazole (DRB) sensitivity-inducing factor (DSIF) and negative elongation factor (NELF) are known to stall collaboratively RNA polymerase II promoter proximally. We discovered that DSIF and NELF are linked to hormone expression in rat pituitary GH4C1 cells. When NELF-E, a subunit of NELF or Spt5, a subunit of DSIF was stably knocked-down, prolactin (PRL) expression was increased both at the mRNA and protein levels. In contrast, stable knock-down of only Spt5 abolished growth hormone (GH) expression. Transient NELF-E knock-down increased coincidentally PRL expression and enhanced transcription of a PRL-promoter reporter gene. However, no direct interaction of NELF with the PRL gene could be demonstrated by chromatin immuno-precipitation. Thus, NELF suppressed PRL promoter activity indirectly. In conclusion, transcription regulation by NELF and DSIF is continuously involved in the control of hormone production and may contribute to neuroendocrine cell differentiation. Copyright 2010 Elsevier Ireland Ltd. All rights reserved.

Proteasome inhibitors enhance endothelial thrombomodulin expression via induction of Krüppel-like transcription factors

PubMed Central

Hiroi, Toyoko; Deming, Clayton B.; Zhao, Haige; Hansen, Baranda S.; Arkenbout, Elisabeth K.; Myers, Thomas J.; McDevitt, Michael A.; Rade, Jeffrey J.

2009-01-01

Objective Impairment of the thrombomodulin-protein C anticoagulant pathway has been implicated in pathologic thrombosis associated with malignancy. Patients who receive proteasome inhibitors as part of their chemotherapeutic regimen appear to be at decreased risk for thromboembolic events. We investigated the effects of proteasome inhibitors on endothelial thrombomodulin expression and function. Methods and Results Proteasome inhibitors as a class markedly induced the expression thrombomodulin and enhanced the protein C activating capacity of endothelial cells. Thrombomodulin upregulation was independent of NF-κB signaling, a principal target of proteasome inhibitors, but was instead a direct consequence of increased expression of the Krüppel-like transcription factors, KLF2 and KLF4. These effects were confirmed in vivo, where systemic administration of a proteasome inhibitor enhanced thrombomodulin expression that was paralleled by changes in the expression of KLF2 and KLF4. Conclusions These findings identify a novel mechanism of action of proteasome inhibitors that may help to explain their clinically observed thromboprotective effects. PMID:19661484

Macronutrients and the FTO gene expression in hypothalamus; a systematic review of experimental studies.

PubMed

Doaei, Saeid; Kalantari, Naser; Mohammadi, Nastaran Keshavarz; Tabesh, Ghasem Azizi; Gholamalizadeh, Maryam

The various studies have examined the relationship between FTO gene expression and macronutrients levels. In order to obtain better viewpoint from this interactions, all of existing studies were reviewed systematically. All published papers have been obtained and reviewed using standard and sensitive keywords from databases such as CINAHL, Embase, PubMed, PsycInfo, and the Cochrane, from 1990 to 2016. The results indicated that all of 6 studies that met the inclusion criteria (from a total of 428 published article) found FTO gene expression changes at short-term follow-ups. Four of six studies found an increased FTO gene expression after calorie restriction, while two of them indicated decreased FTO gene expression. The effect of protein, carbohydrate and fat were separately assessed and suggested by all of six studies. In Conclusion, The level of FTO gene expression in hypothalamus is related to macronutrients levels. Future research should evaluate the long-term impact of dietary interventions. Copyright © 2017. Published by Elsevier B.V.

The homeodomain transcription factor Cdx1 does not behave as an oncogene in normal mouse intestine.

PubMed

Crissey, Mary Ann S; Guo, Rong-Jun; Fogt, Franz; Li, Hong; Katz, Jonathan P; Silberg, Debra G; Suh, Eun Ran; Lynch, John P

2008-01-01

The Caudal-related homeobox genes Cdx1 and Cdx2 are intestine-specific transcription factors that regulate differentiation of intestinal cell types. Previously, we have shown Cdx1 to be antiproliferative and to promote cell differentiation. However, other studies have suggested that Cdx1 may be an oncogene. To test for oncogenic behavior, we used the murine villin promoter to ectopically express Cdx1 in the small intestinal villi and colonic surface epithelium. No changes in intestinal architecture, cell differentiation, or lineage selection were observed with expression of the transgene. Classic oncogenes enhance proliferation and induce tumors when ectopically expressed. However, the Cdx1 transgene neither altered intestinal proliferation nor induced spontaneous intestinal tumors. In a murine model for colitis-associated cancer, the Cdx1 transgene decreased, rather than increased, the number of adenomas that developed. In the polyps, the expression of the endogenous and the transgenic Cdx1 proteins was largely absent, whereas endogenous Villin expression was retained. This suggests that transgene silencing was specific and not due to a general Villin inactivation. In conclusion, neither the ectopic expression of Cdx1 was associated with changes in intestinal cell proliferation or differentiation nor was there increased intestinal cancer susceptibility. Our results therefore suggest that Cdx1 is not an oncogene in normal intestinal epithelium.

Tumour-derived alkaline phosphatase regulates tumour growth, epithelial plasticity and disease-free survival in metastatic prostate cancer

PubMed Central

Rao, S R; Snaith, A E; Marino, D; Cheng, X; Lwin, S T; Orriss, I R; Hamdy, F C; Edwards, C M

2017-01-01

Background: Recent evidence suggests that bone-related parameters are the main prognostic factors for overall survival in advanced prostate cancer (PCa), with elevated circulating levels of alkaline phosphatase (ALP) thought to reflect the dysregulated bone formation accompanying distant metastases. We have identified that PCa cells express ALPL, the gene that encodes for tissue nonspecific ALP, and hypothesised that tumour-derived ALPL may contribute to disease progression. Methods: Functional effects of ALPL inhibition were investigated in metastatic PCa cell lines. ALPL gene expression was analysed from published PCa data sets, and correlated with disease-free survival and metastasis. Results: ALPL expression was increased in PCa cells from metastatic sites. A reduction in tumour-derived ALPL expression or ALP activity increased cell death, mesenchymal-to-epithelial transition and reduced migration. Alkaline phosphatase activity was decreased by the EMT repressor Snail. In men with PCa, tumour-derived ALPL correlated with EMT markers, and high ALPL expression was associated with a significant reduction in disease-free survival. Conclusions: Our studies reveal the function of tumour-derived ALPL in regulating cell death and epithelial plasticity, and demonstrate a strong association between ALPL expression in PCa cells and metastasis or disease-free survival, thus identifying tumour-derived ALPL as a major contributor to the pathogenesis of PCa progression. PMID:28006818

Development of Urinary Bladder Pre-Neoplasia by Schistosoma haematobium Eggs and Chemical Carcinogen in Mice

PubMed Central

Chala, Bayissa; Choi, Min-Ho; Moon, Kyung Chul; Kim, Hyung Suk; Kwak, Cheol; Hong, Sung-Tae

2017-01-01

Schistosoma haematobium is a biocarcinogen of human urinary bladder (UB). The present study investigated developing UB cancer mouse model by injecting S. haematobium eggs into the bladder wall and introduction of chemical carcinogens. Histopathological findings showed mild hyperplasia to epithelial vacuolar change, and high grade dysplasia. Squamous metaplasia was observed in the S. haematobium eggs+NDMA group at week 12 but not in other groups. Immunohistochemistry revealed significantly high expression of Ki-67 in urothelial epithelial cells of the S. haematobium eggs+BBN group at week 20. The qRT-PCR showed high expression of p53 gene in S. haematobium eggs group at week 4 and S. haematobium eggs+BBN group at week 20. E-cadherin and vimentin showed contrasting expression in S. haematobium eggs+BBN group. Such inverse expression of E-cadherin and vimentin may indicate epithelial mesenchymal transition in the UB tissue. In conclusion, S. haematobium eggs and nitrosamines may transform UB cells into squamous metaplasia and dysplasia in correlation with increased expression of Ki-67. Marked decrease in E-cadherin and increase in p53 and vimentin expressions may support the transformation. The present study introduces a promising modified animal model for UB cancer study using S. haematobium eggs. PMID:28285503

Expression variability of co-regulated genes differentiates Saccharomyces cerevisiae strains

PubMed Central

2011-01-01

Background Saccharomyces cerevisiae (Baker's yeast) is found in diverse ecological niches and is characterized by high adaptive potential under challenging environments. In spite of recent advances on the study of yeast genome diversity, little is known about the underlying gene expression plasticity. In order to shed new light onto this biological question, we have compared transcriptome profiles of five environmental isolates, clinical and laboratorial strains at different time points of fermentation in synthetic must medium, during exponential and stationary growth phases. Results Our data unveiled diversity in both intensity and timing of gene expression. Genes involved in glucose metabolism and in the stress response elicited during fermentation were among the most variable. This gene expression diversity increased at the onset of stationary phase (diauxic shift). Environmental isolates showed lower average transcript abundance of genes involved in the stress response, assimilation of nitrogen and vitamins, and sulphur metabolism, than other strains. Nitrogen metabolism genes showed significant variation in expression among the environmental isolates. Conclusions Wild type yeast strains respond differentially to the stress imposed by nutrient depletion, ethanol accumulation and cell density increase, during fermentation of glucose in synthetic must medium. Our results support previous data showing that gene expression variability is a source of phenotypic diversity among closely related organisms. PMID:21507216

(−)-Epicatechin Prevents Blood Pressure Increase and Reduces Locomotor Hyperactivity in Young Spontaneously Hypertensive Rats

PubMed Central

Berenyiova, A.; Drobna, M.; Lukac, S.

2016-01-01

This study investigated the effects of subchronic (−)-epicatechin (Epi) treatment on locomotor activity and hypertension development in young spontaneously hypertensive rats (SHR). Epi was administered in drinking water (100 mg/kg/day) for 2 weeks. Epi significantly prevented the development of hypertension (138 ± 2 versus 169 ± 5 mmHg, p < 0.001) and reduced total distance traveled in the open-field test (22 ± 2 versus 35 ± 4 m, p < 0.01). In blood, Epi significantly enhanced erythrocyte deformability, increased total antioxidant capacity, and decreased nitrotyrosine concentration. In the aorta, Epi significantly increased nitric oxide (NO) synthase (NOS) activity and elevated the NO-dependent vasorelaxation. In the left heart ventricle, Epi increased NOS activity without altering gene expressions of nNOS, iNOS, and eNOS. Moreover, Epi reduced superoxide production in the left heart ventricle and the aorta. In the brain, Epi increased nNOS gene expression (in the brainstem and cerebellum) and eNOS expression (in the cerebellum) but had no effect on overall NOS activity. In conclusion, Epi prevented the development of hypertension and reduced locomotor hyperactivity in young SHR. These effects resulted from improved cardiovascular NO bioavailability concurrently with increased erythrocyte deformability, without changes in NO production in the brain. PMID:27885334

Anger Arousal and Behavioral Anger Regulation in Everyday Life among Patients with Chronic Low Back Pain: Relationships to Patient Pain and Function

PubMed Central

Burns, John W.; Gerhart, James I.; Bruehl, Stephen; Peterson, Kristina M.; Smith, David A.; Porter, Laura S.; Schuster, Erik; Kinner, Ellen; Buvanendran, Asokumar; Marie Fras, Anne; Keefe, Francis J.

2015-01-01

Objective To determine the degree to which patient anger arousal and behavioral anger regulation (expression, inhibition) occurring in the course of daily life was related to patient pain and function as rated by patients and their spouses. Method Married couples (N = 105) (one spouse with chronic low back pain) completed electronic daily diaries, with assessments 5 times/day for 14 days. Patients completed items on their own state anger, behavioral anger expression and inhibition, and pain-related factors. Spouses completed items on their observations of patient pain-related factors. Hierarchical linear modeling was used to test concurrent and lagged relationships. Results Patient-reported increases in state anger were related to their reports of concurrent increases in pain and pain interference and to spouse reports of patient pain and pain behavior. Patient-reported increases in behavioral anger expression were related to lagged increases in pain intensity and interference and decreases in function. Most of these relationships remained significant with state anger controlled. Patient-reported increases in behavioral anger inhibition were related to concurrent increases in pain interference and decreases in function, which also remained significant with state anger controlled. Patient-reported increases in state anger were related to lagged increases in spouse reports of patient pain intensity and pain behaviors. Conclusions Results indicate that in patients with chronic pain, anger arousal and both behavioral anger expression and inhibition in everyday life are related to elevated pain intensity and decreased function as reported by patients. Spouse ratings show some degree of concordance with patient reports. PMID:25110843

G protein, phosphorylated-GATA4 and VEGF expression in the hearts of transgenic mice overexpressing β1- and β2-adrenergic receptors

PubMed Central

Tae, Hyun-Jin; Petrashevskaya, Natalia; Kim, In Hye; Park, Joon Ha; Lee, Jae-Chul; Won, Moo-Ho; Kim, Yang Hee; Ahn, Ji Hyeon; Park, Jinseu; Choi, Soo Young; Jeon, Yong Hwan

2017-01-01

β1- and β2-adrenergic receptors (ARs) regulate cardiac contractility, calcium handling and protein phosphorylation. The present study aimed to examine the expression levels of vascular endothelial growth factor A (VEGF-A) and several G proteins, and the phosphorylation of transcription factor GATA binding protein 4 (GATA4), by western blot analysis, using isolated hearts from 6 month-old transgenic (TG) mice that overexpress β1AR or β2AR. Cardiac contractility/relaxation and heart rate was increased in both β1AR TG and β2AR TG mouse hearts compared with wild type; however, no significant differences were observed between the β1- and β2AR TG mouse hearts. Protein expression levels of inhibitory guanine nucleotide-binding protein (Gi) 2, Gi3 and G-protein-coupled receptor kinase 2 were upregulated in both TG mice, although the upregulation of Gi2 was more prominent in the β2AR TG mice. VEGF-A expression levels were also increased in both TG mice, and were highest in the β1AR TG mice. In addition, the levels of phosphorylated-GATA4 expression were increased in β1- and β2AR TG mice. In conclusion, the present study demonstrated that cardiac contractility/relaxation and heart rate is increased in β1AR TG and β2AR TG mice, and indicated that this increase may be related to the overexpression of G proteins and G-protein-associated proteins. PMID:28487987

Fluid shear stress enhances the cell volume decrease of osteoblast cells by increasing the expression of the ClC-3 chloride channel

PubMed Central

LIU, LI; CAI, SIYI; QIU, GUIXING; LIN, JIN

2016-01-01

ClC-3 is a volume-sensitive chloride channel that is responsible for cell volume adjustment and regulatory cell volume decrease (RVD). In order to evaluate the effects of fluid shear stress (FSS) stimulation on the osteoblast ClC-3 chloride channel, MC3T3-E1 cells were stimulated by FSS in the experimental group. Fluorescence quantitative polymerase chain reaction was used to detect changes in ClC-3 mRNA expression, the chloride ion fluorescent probe N-(ethoxycarbonylmethyl)-6-methoxyquinolinium bromide (MQAE) was used to detect the chloride channel activity, and whole-cell patch clamping was used to monitor the changes in the volume-sensitive chloride current activated by a hypotonic environment following mechanical stimulation. The results show that the expression of the osteoblast chloride channel ClC-3 was significantly higher in the FSS group compared with the control group. MQAE fluorescence intensity was significantly reduced in the FSS group compared to the control group, suggesting that mechanical stimulation increased chloride channel activity and increased the efflux of intracellular chloride ions. Image analysis of osteoblast volume changes showed that osteoblast RVD was enhanced by mechanical stimulation. Whole-cell patch clamping showed that the osteoblast volume-sensitive chloride current was larger in the stimulated group compared to the control group, suggesting that elevated ClC-3 chloride channel expression results in an increased volume-sensitive chloride current. In conclusion, FSS stimulation enhances the RVD of osteoblast cell by increasing the expression of the ClC-3 and enhancing the chloride channel activity. PMID:27073622

Expression and Function of Methylthioadenosine Phosphorylase in Chronic Liver Disease

PubMed Central

Czech, Barbara; Dettmer, Katja; Valletta, Daniela; Saugspier, Michael; Koch, Andreas; Stevens, Axel P.; Thasler, Wolfgang E.; Müller, Martina; Oefner, Peter J.; Bosserhoff, Anja-Katrin; Hellerbrand, Claus

2013-01-01

To study expression and function of methylthioadenosine phosphorylase (MTAP), the rate-limiting enzyme in the methionine and adenine salvage pathway, in chronic liver disease. Design MTAP expression was analyzed by qRT-PCR, Western blot and immunohistochemical analysis. Levels of MTA were determined by liquid chromatography-tandem mass spectrometry. Results MTAP was downregulated in hepatocytes in murine fibrosis models and in patients with chronic liver disease, leading to a concomitant increase in MTA levels. In contrast, activated hepatic stellate cells (HSCs) showed strong MTAP expression in cirrhotic livers. However, also MTA levels in activated HSCs were significantly higher than in hepatocytes, and there was a significant correlation between MTA levels and collagen expression in diseased human liver tissue indicating that activated HSCs significantly contribute to elevated MTA in diseased livers. MTAP suppression by siRNA resulted in increased MTA levels, NFκB activation and apoptosis resistance, while overexpression of MTAP caused the opposite effects in HSCs. The anti-apoptotic effect of low MTAP expression and high MTA levels, respectively, was mediated by induced expression of survivin, while inhibition of survivin abolished the anti-apoptotic effect of MTA on HSCs. Treatment with a DNA demethylating agent induced MTAP and reduced survivin expression, while oxidative stress reduced MTAP levels but enhanced survivin expression in HSCs. Conclusion MTAP mediated regulation of MTA links polyamine metabolism with NFκB activation and apoptosis in HSCs. MTAP and MTAP modulating mechanisms appear as promising prognostic markers and therapeutic targets for hepatic fibrosis. PMID:24324622

Cervical HSV-2 infection causes cervical remodeling and increases risk for ascending infection and preterm birth.

PubMed

McGee, Devin; Smith, Arianna; Poncil, Sharra; Patterson, Amanda; Bernstein, Alison I; Racicot, Karen

2017-01-01

Preterm birth (PTB), or birth before 37 weeks gestation, is the leading cause of neonatal mortality worldwide. Cervical viral infections have been established as risk factors for PTB in women, although the mechanism leading to increased risk is unknown. Using a mouse model of pregnancy, we determined that intra-vaginal HSV2 infection caused increased rates of preterm birth following an intra-vaginal bacterial infection. HSV2 infection resulted in histological changes in the cervix mimicking cervical ripening, including significant collagen remodeling and increased hyaluronic acid synthesis. Viral infection also caused aberrant expression of estrogen and progesterone receptor in the cervical epithelium. Further analysis using human ectocervical cells demonstrated a role for Src kinase in virus-mediated changes in estrogen receptor and hyaluronic acid expression. In conclusion, HSV2 affects proteins involved in tissue hormone responsiveness, causes significant changes reminiscent of premature cervical ripening, and increases risk of preterm birth. Studies such as this improve our chances of identifying clinical interventions in the future.

Cervical HSV-2 infection causes cervical remodeling and increases risk for ascending infection and preterm birth

PubMed Central

McGee, Devin; Poncil, Sharra; Patterson, Amanda

2017-01-01

Preterm birth (PTB), or birth before 37 weeks gestation, is the leading cause of neonatal mortality worldwide. Cervical viral infections have been established as risk factors for PTB in women, although the mechanism leading to increased risk is unknown. Using a mouse model of pregnancy, we determined that intra-vaginal HSV2 infection caused increased rates of preterm birth following an intra-vaginal bacterial infection. HSV2 infection resulted in histological changes in the cervix mimicking cervical ripening, including significant collagen remodeling and increased hyaluronic acid synthesis. Viral infection also caused aberrant expression of estrogen and progesterone receptor in the cervical epithelium. Further analysis using human ectocervical cells demonstrated a role for Src kinase in virus-mediated changes in estrogen receptor and hyaluronic acid expression. In conclusion, HSV2 affects proteins involved in tissue hormone responsiveness, causes significant changes reminiscent of premature cervical ripening, and increases risk of preterm birth. Studies such as this improve our chances of identifying clinical interventions in the future. PMID:29190738

Elevated toll-like receptor 4 expression and signaling in muscle from insulin-resistant subjects.

PubMed

Reyna, Sara M; Ghosh, Sangeeta; Tantiwong, Puntip; Meka, C S Reddy; Eagan, Phyllis; Jenkinson, Christopher P; Cersosimo, Eugenio; Defronzo, Ralph A; Coletta, Dawn K; Sriwijitkamol, Apiradee; Musi, Nicolas

2008-10-01

OBJECTIVE- Tall-like receptor (TLR)4 has been implicated in the pathogenesis of free fatty acid (FFA)-induced insulin resistance by activating inflammatory pathways, including inhibitor of kappaB (IkappaB)/nuclear factor kappaB (NFkappaB). However, it is not known whether insulin-resistant subjects have abnormal TLR4 signaling. We examined whether insulin-resistant subjects have abnormal TLR4 expression and TLR4-driven (IkappaB/NFkappaB) signaling in skeletal muscle. RESEARCH DESIGN AND METHODS- TLR4 gene expression and protein content were measured in muscle biopsies in 7 lean, 8 obese, and 14 type 2 diabetic subjects. A primary human myotube culture system was used to examine whether FFAs stimulate IkappaB/NFkappaB via TLR4 and whether FFAs increase TLR4 expression/content in muscle. RESULTS- Obese and type 2 diabetic subjects had significantly elevated TLR4 gene expression and protein content in muscle. TLR4 muscle protein content correlated with the severity of insulin resistance. Obese and type 2 diabetic subjects also had lower IkappaBalpha content, an indication of elevated IkappaB/NFkappaB signaling. The increase in TLR4 and NFkappaB signaling was accompanied by elevated expression of the NFkappaB-regulated genes interleukin (IL)-6 and superoxide dismutase (SOD)2. In primary human myotubes, acute palmitate treatment stimulated IkappaB/NFkappaB, and blockade of TLR4 prevented the ability of palmitate to stimulate the IkappaB/NFkappaB pathway. Increased TLR4 content and gene expression observed in muscle from insulin-resistant subjects were reproduced by treating myotubes from lean, normal-glucose-tolerant subjects with palmitate. Palmitate also increased IL-6 and SOD2 gene expression, and this effect was prevented by inhibiting NFkappaB. CONCLUSIONS- Abnormal TLR4 expression and signaling, possibly caused by elevated plasma FFA levels, may contribute to the pathogenesis of insulin resistance in humans.

FcγRIIIa Expression on Monocytes in Rheumatoid Arthritis: Role in Immune-Complex Stimulated TNF Production and Non-Response to Methotrexate Therapy

PubMed Central

Cooper, Dawn L.; Martin, Stephen G.; Robinson, James I.; Mackie, Sarah L.; Charles, Christopher J.; Nam, Jackie; Consortium, YEAR; Isaacs, John D.; Emery, Paul; Morgan, Ann W.

2012-01-01

Objective The expression of FcγRIIIa/CD16 may render monocytes targets for activation by IgG-containing immune complexes (IC). We investigated whether FcγRIIIa/CD16 was upregulated in rheumatoid arthritis (RA), associated with TNF production in response to IC-stimulation, and if this predicted response to methotrexate therapy. Methods FcγRIIIa/CD16 expression on CD14low and CD14++ monocytes was measured by flow cytometry in healthy controls and RA patients (early and long-standing disease). Intracellular TNF-staining was carried out after in vitro LPS or heat-aggregated immunoglobulin (HAG) activation. FcγRIIIa/CD16 expression pre- and post-steroid/methotrexate treatment was examined. Results Increased FcγRIIIa/CD16 expression on CD14++ monocytes in long-standing RA patients compared to controls was demonstrated (p = 0.002) with intermediate levels in early-RA patients. HAG-induced TNF-production in RA patients was correlated with the percentage of CD14++ monocytes expressing FcγRIIIa/CD16 (p<0.001). The percentage of CD14++ monocytes expressing FcγRIIIa/CD16 at baseline in early DMARD-naïve RA patients was negatively correlated with DAS28-ESR improvement 14-weeks post-methotrexate therapy (p = 0.003) and was significantly increased in EULAR non-responders compared to moderate (p = 0.01) or good responders (p = 0.003). FcγRIIIa/CD16 expression was not correlated with age, presence of systemic inflammation or autoantibody titers. Conclusion Increased FcγRIIIa/CD16 expression on CD14++ monocytes in RA may result in a cell that has increased responsiveness to IC-stimulation. This monocyte subset may contribute to non-response to methotrexate therapy. PMID:22235253

Short-term in vivo inhibition of nitric oxide synthase with L-NAME influences the contractile function of single left ventricular myocytes in rats.

PubMed

Lunz, Wellington; Natali, Antônio José; Carneiro, Miguel Araújo; Dos Santos Aggum Capettini, Luciano; Baldo, Marcelo Perim; de Souza, Matheus Ornelas; Quintão, Judson Fonseca; Bozi, Luiz Henrique Marchesi; Lemos, Virginia Soares; Mill, José Geraldo

2011-04-01

The main purpose of this study was to investigate the effects of short-term L-NAME treatment on the contractile function of left ventricle (LV) myocytes and the expression of proteins related to Ca(2+) homeostasis. Data from Wistar rats treated with L-NAME (L group, n = 20; 0.7 g/L in drinking water; 7 days) were compared with results from untreated controls (C group, n = 20). Cardiomyocytes from the L group showed increased (p < 0.05) fractional shortening (23%) and maximum rate of shortening (20%) compared with the C group. LV from the L group also showed increased (p < 0.05) expression of the ryanodine receptor 2 and Na(+)/Ca(2+) exchanger proteins (76% and 83%, respectively; p < 0.05). However, the L and C groups showed similar in vivo hemodynamic parameters of cardiac function. In conclusion, short-term NOS inhibition determines an increased expression of Ca(2+) regulatory proteins, which contributes to improving cardiomyocyte contractile function, preserving left ventricular function.

Elevated HERV-K Expression in Soft Tissue Sarcoma Is Associated with Worsened Relapse-Free Survival.

PubMed

Giebler, Maria; Staege, Martin S; Blauschmidt, Sindy; Ohm, Lea I; Kraus, Matthias; Würl, Peter; Taubert, Helge; Greither, Thomas

2018-01-01

A wide variety of endogenous retroviral sequences has been demonstrated in the human genome so far, divided into several different families according to the sequence homology to viral strains. While increased expression of human endogenous retrovirus (HERV) elements has already been linked to unfavorable prognosis in hepatocellular carcinoma, breast cancer, and ovarian carcinoma yet less is known about the impact of the expression of different HERV elements on sarcomagenesis in general as well as the outcome of soft tissue sarcoma (STS) patients. Therefore, in this study the association between expression of HERV-K and HERV-F and the clinicopathological characteristics in a cohort of STSs as well as the patients' prognosis was evaluated. HERV-K and HERV-F expression was assessed by quantitative real-time PCR in 120 patient specimens. HERV-K and HERV-F expression was significantly correlated ( r S = 0.5; p = 6.4 × 10 -9 ; Spearman's rank bivariate correlation). Also, tumor diameter exhibited a significant negative association to HERV-K and HERV-F expression. Levels of several hypoxia-related RNAs like HIF-1α and miR-210 showed a significant positive correlation with both HERV-K and HERV-F expression. Although in survival analyses no impact of HERV expression on disease-specific survival could be detected, patients with elevated HERV-K expression had a significantly shorter relapse-free survival ( p = 0.014, log-rank analysis). In conclusion, we provide evidence for the first time that the increased expression of HERV-K in tumors is associated with STS patients' prognosis.

Effect of Acupotomy on FAK-PI3K Signaling Pathways in KOA Rabbit Articular Cartilages

PubMed Central

Xie, Zhan-guo; Guo, Yan; Yu, Jia-Ni; Lu, Juan; Zhang, Wei; Wang, Li-Juan; Xu, Jing; Zhao, Rui-Li; Zhou, Shuai

2017-01-01

Objective By observing the needle-knife of KOA rabbit morphology, knee joint cartilage p-FAK, p-PI3K, Aggrecan gene, and protein expression, to study the effect of needle-knife to promote cartilage cell synthesis metabolism mechanism. Method 49 male New Zealand rabbits, randomly divided into normal group (Z), model group (M), model-inhibitors (MP), needle-knife group (D), needle-knife inhibitors group (DP), electroacupuncture group (E), and electroacupuncture inhibitors (EP). RT-PCR and Western Blot were used to test each animal cartilage p-FAK, p-PI3K, and Aggrecan gene and protein expression level. Results Compared with N group, p-FAK and p-PI3K protein and mRNA expression of M group, D group, and E group increased (P < 0.05), while the protein and mRNA expression of Aggrecan reduced (P < 0.05). Compared with M group, p-FAK, p-PI3K, Aggrecan protein, and mRNA of E and D group increased (P < 0.05). Compared with E group, p-FAK, p-PI3K, Aggrecan protein, and mRNA expression of D group increased (P < 0.05); after adding inhibitors, p-FAK, p-PI3K, Aggrecan protein, and mRNA expression reduced (P < 0.05). Conclusion Needle-knife therapy can promote the repairment of cartilage cells by activating FAK-PI3K signaling pathways, promoting the synthesis of cartilage cell metabolism. PMID:29234400

Expression and regulation of glucocorticoid-induced leucine zipper in the developing anterior pituitary gland.

PubMed

Ellestad, Laura E; Malkiewicz, Stefanie A; Guthrie, H David; Welch, Glenn R; Porter, Tom E

2009-02-01

The expression profile of glucocorticoid-induced leucine zipper (GILZ) in the anterior pituitary during the second half of embryonic development in the chick is consistent with in vivo regulation by circulating corticosteroids. However, nothing else has been reported about the presence of GILZ in the neuroendocrine system. We sought to characterize expression and regulation of GILZ in the chicken embryonic pituitary gland and determine the effect of GILZ overexpression on anterior pituitary hormone levels. Pituitary GILZ mRNA levels increased during embryogenesis to a maximum on the day of hatch, and decreased through the first week after hatch. GILZ expression was rapidly upregulated by corticosterone in embryonic pituitary cells. To determine whether GILZ regulates hormone gene expression in the developing anterior pituitary, we overexpressed GILZ in embryonic pituitary cells and measured mRNA for the major pituitary hormones. Exogenous GILZ increased prolactin mRNA above basal levels, but not as high as that in corticosterone-treated cells, indicating that GILZ may play a small role in lactotroph differentiation. The largest effect we observed was a twofold increase in FSH beta subunit in cells transfected with GILZ but not treated with corticosterone, suggesting that GILZ may positively regulate gonadotroph development in a manner not involving glucocorticoids. In conclusion, this is the first report to characterize avian GILZ and examine its regulation in the developing neuroendocrine system. We have shown that GILZ is upregulated by glucocorticoids in the embryonic pituitary gland and may regulate expression of several pituitary hormones.

COX2 Inhibition Reduces Aortic Valve Calcification In Vivo

PubMed Central

Wirrig, Elaine E.; Gomez, M. Victoria; Hinton, Robert B.; Yutzey, Katherine E.

2016-01-01

Objective Calcific aortic valve disease (CAVD) is a significant cause of morbidity and mortality, which affects approximately 1% of the US population and is characterized by calcific nodule formation and stenosis of the valve. Klotho-deficient mice were used to study the molecular mechanisms of CAVD as they develop robust aortic valve (AoV) calcification. Through microarray analysis of AoV tissues from klotho-deficient and wild type mice, increased expression of the gene encoding cyclooxygenase 2/COX2 (Ptgs2) was found. COX2 activity contributes to bone differentiation and homeostasis, thus the contribution of COX2 activity to AoV calcification was assessed. Approach and Results In klotho-deficient mice, COX2 expression is increased throughout regions of valve calcification and is induced in the valvular interstitial cells (VICs) prior to calcification formation. Similarly, COX2 expression is increased in human diseased AoVs. Treatment of cultured porcine aortic VICs with osteogenic media induces bone marker gene expression and calcification in vitro, which is blocked by inhibition of COX2 activity. In vivo, genetic loss of function of COX2 cyclooxygenase activity partially rescues AoV calcification in klotho-deficient mice. Moreover, pharmacologic inhibition of COX2 activity in klotho-deficient mice via celecoxib-containing diet reduces AoV calcification and blocks osteogenic gene expression. Conclusions COX2 expression is upregulated in CAVD and its activity contributes to osteogenic gene induction and valve calcification in vitro and in vivo. PMID:25722432

Cyclooxygenase-2/carbonic anhydrase-IX up-regulation promotes invasive potential and hypoxia survival in colorectal cancer cells

PubMed Central

Sansone, Pasquale; Piazzi, Giulia; Paterini, Paola; Strillacci, Antonio; Ceccarelli, Claudio; Minni, Francesco; Biasco, Guido; Chieco, Pasquale; Bonafè, Massimiliano

2009-01-01

Inflammation promotes colorectal carcinogenesis. Tumour growth often generates a hypoxic environment in the inner tumour mass. We here report that, in colon cancer cells, the expression of the pro-inflammatory enzyme cyclooxygenase-2 (COX-2) associates with that of the hypoxia response gene carbonic anhydrase-IX (CA-IX). The COX-2 knockdown, achieved by the stable infection of a COX-2 specific short harpin RNA interference (shCOX-2), down-regulates CA-IX gene expression. In colorectal cancer (CRC) cells, PGE2, the main COX-2 gene products, promotes CA-IX gene expression by ERK1/2 activation. In normoxic environment, shCOX-2 infected/CA-IX siRNA transfected CRC cells show a reduced level of active metalloproteinase-2 (MMP-2) that associates with a decreased extracellular matrix invasion capacity. In presence of hypoxia, COX-2 gene expression and PGE2 production increase. The knockdown of COX-2/CA-IX blunts the survival capability of CRC cells in hypoxia. At a high cell density, a culture condition that creates a mild pericellular hypoxic environment, the expression of COX-2/CA-IX genes is increased and triggers the invasive potential of colon cancer cells. In human colon cancer tissues, COX-2/CA-IX protein expression levels, assessed by Western blot and immunohistochemistry, correlate each other and increase with tumour stage. In conclusion, these data indicate that COX-2/CA-IX interplay promotes the aggressive behaviour of CRC cells. PMID:19017360

Hyperphosphatemia induces cellular senescence in human aorta smooth muscle cells through integrin linked kinase (ILK) up-regulation.

PubMed

Troyano, Nuria; Nogal, María Del; Mora, Inés; Diaz-Naves, Manuel; Lopez-Carrillo, Natalia; Sosa, Patricia; Rodriguez-Puyol, Diego; Olmos, Gemma; Ruiz-Torres, María P

2015-12-01

Aging is conditioned by genetic and environmental factors. Hyperphosphatemia is related to some pathologies, affecting to vascular cells behavior. This work analyze whether high concentration of extracellular phosphate induces vascular smooth muscle cells senescence, exploring the intracellular mechanisms and highlighting the in vivo relevance of this phenomenon. Human aortic smooth muscle cells treated with β-Glycerophosphate (BGP, 10mM) suffered cellular senescence by increasing p53, p21 and p16 expression and the senescence associated β-galactosidase activity. In parallel, BGP induced ILK overexpression, dependent on the IGF-1 receptor activation, and oxidative stress. Down-regulating ILK expression prevented BGP-induced senescence and oxidative stress. Aortic rings from young rats treated with 10mM BGP for 48h, showed increased p53, p16 and ILK expression and SA-β-gal activity. Seven/eight nephrectomized rats feeding a hyperphosphatemic diet and fifteenth- month old mice showed hyperphosphatemia and aortic ILK, p53 and p16 expression. In conclusion, we demonstrated that high extracellular concentration of phosphate induced senescence in cultured smooth muscle through the activation of IGF-1 receptor and ILK overexpression and provided solid evidences for the in vivo relevance of these results since aged animals showed high levels of serum phosphate linked to increased expression of ILK and senescence genes. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

Immunogenicity and immunomodulatory properties of umbilical cord lining mesenchymal stem cells.

PubMed

Deuse, Tobias; Stubbendorff, Mandy; Tang-Quan, Karis; Phillips, Neil; Kay, Mark A; Eiermann, Thomas; Phan, Thang T; Volk, Hans-Dieter; Reichenspurner, Hermann; Robbins, Robert C; Schrepfer, Sonja

2011-01-01

We here present an immunologic head-to-head comparison between human umbilical cord lining mesenchymal stem cells (clMSCs) and adult bone marrow MSCs (bmMSCs) from patients >65 years of age. clMSCs had significantly lower HLA class I expression, higher production of tolerogenic TGF-β and IL-10, and showed significantly faster proliferation. In vitro activation of allogeneic lymphocytes and xenogeneic in vivo immune activation was significantly stronger with bmMSCs, whereas immune recognition of clMSCs was significantly weaker. Thus, bmMSCs were more quickly rejected in immunocompetent mice. IFN-γ at 25 ng/ml increased both immunogenicity by upregulation of HLA class I/ HLA-DR expression and tolerogenicity by increasing intracellular HLA-G and surface HLA-E expression, augmenting TGF-β and IL-10 release, and inducing indoleamine 2,3-dioxygenase (IDO) expression. Higher concentrations of IFN-γ (>50 ng/ml) further enhanced the immunosuppressive phenotype of clMSCs, more strongly downregulating HLA-DR expression and further increasing IDO production (at 500 ng/ml). The net functional immunosuppressive efficacy of MSCs was tested in mixed lymphocyte cultures. Although both clMSCs and bmMSCs significantly reduced in vitro immune activation, clMSCs were significantly more effective than bmMSCs. The veto function of both MSC lines was enhanced in escalating IFN-γ environments. In conclusion, clMSCs show a more beneficial immunogeneic profile and stronger overall immunosuppressive potential than aged bmMSCs.

Gene expression information improves reliability of receptor status in breast cancer patients

PubMed Central

Kenn, Michael; Schlangen, Karin; Castillo-Tong, Dan Cacsire; Singer, Christian F.; Cibena, Michael; Koelbl, Heinz; Schreiner, Wolfgang

2017-01-01

Immunohistochemical (IHC) determination of receptor status in breast cancer patients is frequently inaccurate. Since it directs the choice of systemic therapy, it is essential to increase its reliability. We increase the validity of IHC receptor expression by additionally considering gene expression (GE) measurements. Crisp therapeutic decisions are based on IHC estimates, even if they are borderline reliable. We further improve decision quality by a responsibility function, defining a critical domain for gene expression. Refined normalization is devised to file any newly diagnosed patient into existing data bases. Our approach renders receptor estimates more reliable by identifying patients with questionable receptor status. The approach is also more efficient since the rate of conclusive samples is increased. We have curated and evaluated gene expression data, together with clinical information, from 2880 breast cancer patients. Combining IHC with gene expression information yields a method more reliable and also more efficient as compared to common practice up to now. Several types of possibly suboptimal treatment allocations, based on IHC receptor status alone, are enumerated. A ‘therapy allocation check’ identifies patients possibly miss-classified. Estrogen: false negative 8%, false positive 6%. Progesterone: false negative 14%, false positive 11%. HER2: false negative 2%, false positive 50%. Possible implications are discussed. We propose an ‘expression look-up-plot’, allowing for a significant potential to improve the quality of precision medicine. Methods are developed and exemplified here for breast cancer patients, but they may readily be transferred to diagnostic data relevant for therapeutic decisions in other fields of oncology. PMID:29100391

MALAT1-miR-124-RBG2 axis is involved in growth and invasion of HR-HPV-positive cervical cancer cells.

PubMed

Liu, Shikai; Song, Lili; Zeng, Saitian; Zhang, Liang

2016-01-01

Metastasis-associated lung adenocarcinoma transcript 1 (MALAT 1) is a large, infrequently spliced non-coding RNA aberrantly expressed in cervical cancer. But the molecular mechanisms of its oncogenic role are still not quite clear. The present study explored whether there is a competing endogenous RNAs (ceRNAs) mechanism involved in the oncogenic effect of MALAT1. MALAT1 expression was firstly verified in high-risk human papillomavirus (HR-HPV)-positive tumor tissues and cell lines. Its regulation over miR-124 and the downstream target of miR-124 in regulation of growth, invasion, and apoptosis of the cancer cells are also studied. Findings of this study confirmed higher MALAT1 expression in HR-HPV (+) cervical cancer. Knockdown of endogenous MALAT1 significantly reduced cell growth rate and invasion and increased cell apoptosis of Hela and siHa cells. Besides, knockdown of MALAT1 increased the expression of miRNA-124, while ectopic expression of miR-124 decreased MALAT1 expression. In addition, we also verified a direct interaction between miR-124 and 3'UTR of GRB2. MALAT1 can indirectly modulate GRB2 expression via competing miR-124. Knockdown of GRB2 reduced cell invasion and increased cell apoptosis. In conclusion, MALAT1 can promote HR-HPV (+) cancer cell growth and invasion at least partially through the MALAT1-miR-124-RBG2 axis. This finding might provide some useful evidence about the lncRNA interaction regulatory network in tumorigenesis cervical cancer.

Aromatase inhibitor regulates let-7 expression and let-7f induced cell migration in endometrial cells from women with endometriosis

PubMed Central

Cho, SiHyun; Mutlu, Levent; Zhou, Yuping; Taylor, Hugh S.

2018-01-01

Objectives To evaluate associations between aromatase inhibitor (AI) treatment and let-7 family microRNA expression in endometriosis. Design In vitro study using Ishikawa cells and human endometrial stromal cells (HESC) obtained from patients with endometriosis Setting University research center. Patients Women undergoing laparoscopic surgery for endometriosis Interventions HESCs and Ishikawa cells treated with various letrozol concentrations and transfected with a mimic of let-7 subtypes of interest Main Outcome Measures microRNAs let7a-f and aromatase expression were evaluated. Migration potential after transfection with a let-7f mimic were analyzed. Results After letrozole treatment for 48 hours, all let-7 subtypes showed a trend toward increased expression in a dose dependent manner in Ishikawa cells, and significant differences were found in let-7b and let-7f between controls and the 20 μmol/L treated groups. Further, let-7f showed significant differences between control and 1.0 μmol/L treatment group, a typical therapeutic level, in HESCs. Transfection of a let-7f mimic decreased aromatase expression in both Ishikawa cells and HESC, and led to a significant decrease in number of migrating cells in both cell types. Conclusions AI treatment significantly increased expression of let-7f in Ishikawa cells and HESCs from patient with endometriosis; increased lef-7f expression effectively reduced the migration of endometrial cells. Modulation of miRNAs involved in the pathogenesis of endometriosis may have therapeutic potential for endometriosis. PMID:27320036

Co-expression of five genes in E coli for L-phenylalanine in Brevibacterium flavum

PubMed Central

Wu, Yong-Qing; Jiang, Pei-Hong; Fan, Chang-Sheng; Wang, Jian-Gang; Shang, Liang; Huang, Wei-Da

2003-01-01

AIM: To study the effect of co-expression of ppsA, pckA, aroG, pheA and tyrB genes on the production of L-phenylalanine, and to construct a genetic engineering strain for L-phenylalanine. METHODS: ppsA and pckA genes were amplified from genomic DNA of E. coli by polymerase chain reaction, and then introduced into shuttle vectors between E coli and Brevibacterium flavum to generate constructs pJN2 and pJN5. pJN2 was generated by inserting ppsA and pckA genes into vector pCZ; whereas pJN5 was obtained by introducing ppsA and pckA genes into pCZ-GAB, which was originally constructed for co-expression of aroG, pheA and tyrB genes. The recombinant plasmids were then introduced into B. flavum by electroporation and the transformants were used for L-phenylalanine fermentation. RESULTS: Compared with the original B. flavum cells, all the transformants were showed to have increased five enzyme activities specifically, and have enhanced L-phenylalanine biosynthesis ability variably. pJN5 transformant was observed to have the highest elevation of L-phenylalanine production by a 3.4-fold. Co-expression of ppsA and pckA increased activity of DAHP synthetase significantly. CONCLUSION: Co-expression of ppsA and pckA genes in B. flavum could remarkably increase the expression of DAHP synthetase; Co-expression of ppsA, pckA, aroG, pheA and tyrB of E. coli in B. flavum was a feasible approach to construct a strain for phenylalanine production. PMID:12532463

Subchronic exposure to arsenic through drinking water alters expression of cancer-related genes in rat liver.

PubMed

Cui, Xing; Li, Song; Shraim, Amjad; Kobayashi, Yayoi; Hayakawa, Toru; Kanno, Sanae; Yamamoto, Megumi; Hirano, Seishiro

2004-01-01

Although arsenic exposure causes liver disease and/or hepatoma, little is known about molecular mechanisms of arsenic-induced liver toxicity or carcinogenesis. We investigated the effects of arsenic on expression of cancer-related genes in a rat liver following subchronic exposure to sodium arsenate (1, 10, 100 ppm in drinking water), by using real-time quantitative RT-PCR and immunohistochemical analyses. Arsenic accumulated in the rat liver dose-dependently and caused hepatic histopathological changes, such as disruption of hepatic cords, sinusoidal dilation, and fatty infiltration. A 1-month exposure to arsenic significantly increased hepatic mRNA levels of cyclin D1 (10 ppm), ILK (1 ppm), and p27(Kip1) (10 ppm), whereas it reduced mRNA levels of PTEN (1 ppm) and beta-catenin (100 ppm). In contrast, a 4-month arsenic exposure showed increased mRNA expression of cyclin D1 (100 ppm), ILK (1 ppm), and p27(Kip1) (1 and 10 ppm), and decreased expression of both PTEN and beta-catenin at all 3 doses. An immunohistochemical study revealed that each protein expression accords closely with each gene expression of mRNA level. In conclusion, subchronic exposure to inorganic arsenate caused pathological changes and altered expression of cyclin D1, p27(Kip1), ILK, PTEN, and beta-catenin in the liver. This implies that arsenic liver toxicity involves disturbances of some cancer-related molecules.

Role of leptin in delayed embryonic development in the Indian short-nosed fruit bat, Cynopterus sphinx.

PubMed

Banerjee, A; Meenakumari, K J; Krishna, A

2010-08-01

An adiposity-associated rise in leptin occurs at the time of delayed embryonic development in Cynopterus sphinx. The aim of present study was to examine the mechanism by which leptin may inhibit progesterone, and therefore could be responsible for delayed development. The study showed a significant increase in circulating leptin level during the period of increased fat accumulation, which coincided with significant decrease in serum progesterone level and delayed embryonic development in C. sphinx. The study showed increased Ob-R expression in the corpus luteum and in the utero-embryonic unit during the period of delayed embryonic development. The in vitro study showed suppressive effect of leptin on progesterone synthesis. The effect of high dose of leptin on ovarian steroidogenesis was found to be mediated through decreased expression of StAR and LH-R proteins in the ovary. The treatment with leptin caused increased expression of STAT 3 and iNOS proteins in the ovary, which correlated with decreased expression of StAR protein in the ovary. The inhibitory effects of leptin on progesterone synthesis in the ovary are thus mediated through STAT 3 and iNOS-NO signaling pathways. This study further demonstrated low expression of PCNA coinciding with the increased concentration of the leptin receptor in the utero-embryonic unit and high circulating leptin level during November. In conclusion, adiposity associated increased leptin level during November-December might play role in suppressing progesterone synthesis in the corpus luteum as well as suppressing the rate of cell-proliferation in the utero-embryonic unit thereby causing delayed embryonic development in C. sphinx. Copyright 2010 Elsevier Inc. All rights reserved.

Lactobacillus acidophilus stimulates the expression of SLC26A3 via a transcriptional mechanism

PubMed Central

Raheja, Geetu; Singh, Varsha; Ma, Ke; Boumendjel, Redouane; Borthakur, Alip; Gill, Ravinder K.; Saksena, Seema; Alrefai, Waddah A.; Ramaswamy, Krishnamurthy

2010-01-01

Clinical efficacy of probiotics in treating various forms of diarrhea has been clearly established. However, mechanisms underlying antidiarrheal effects of probiotics are not completely defined. Diarrhea is caused either by decreased absorption or increased secretion of electrolytes and solutes in the intestine. In this regard, the electroneutral absorption of two major electrolytes, Na+ and Cl−, occurs mainly through the coupled operation of Na+/H+ exchangers and Cl−/OH− exchangers. Previous studies from our laboratory have shown that Lactobacillus acidophilus (LA) acutely stimulated Cl−/OH− exchange activity via an increase in the surface levels of the apical anion exchanger SLC26A3 (DRA). However, whether probiotics influence SLC26A3 expression and promoter activity has not been examined. The present studies were, therefore, undertaken to investigate the long-term effects of LA on SLC26A3 expression and promoter activity. Treatment of Caco-2 cells with LA for 6–24 h resulted in a significant increase in Cl−/OH− exchange activity. DRA mRNA levels were also significantly elevated in response to LA treatment starting as early as 8 h. Additionally, the promoter activity of DRA was increased by more than twofold following 8 h LA treatment of Caco-2 cells. Similar to the in vitro studies, in vivo studies using mice gavaged with LA also showed significantly increased DRA mRNA (∼4-fold) and protein expression in the colonic regions as assessed by Western blot analysis and immunofluorescence. In conclusion, increase in DRA promoter activity and expression may contribute to the upregulation of intestinal electrolyte absorption and might underlie the potential antidiarrheal effects of LA. PMID:20044511

Haplodeficiency of Klotho Gene Causes Arterial Stiffening via Upregulation of Scleraxis Expression and Induction of Autophagy.

PubMed

Chen, Kai; Zhou, Xiaoli; Sun, Zhongjie

2015-11-01

The prevalence of arterial stiffness increases with age, whereas the level of the aging-suppressor protein klotho decreases with age. The objective of this study is to assess whether haplodeficiency of klotho gene causes arterial stiffness and to investigate the underlying mechanism. Pulse wave velocity, a direct measure of arterial stiffness, was increased significantly in klotho heterozygous (klotho(+/-)) mice versus their age-matched wild-type (WT) littermates, suggesting that haplodeficiency of klotho causes arterial stiffening. Notably, plasma aldosterone levels were elevated significantly in klotho(+/-) mice. Treatment with eplerenone (6 mg/kg per day IP), an aldosterone receptor blocker, abolished klotho deficiency-induced arterial stiffening in klotho(+/-) mice. Klotho deficiency was associated with increased collagen and decreased elastin contents in the media of aortas. In addition, arterial matrix metalloproteinase-2, matrix metalloproteinase-9, and transforming growth factor-β1 expression and myofibroblast differentiation were increased in klotho(+/-) mice. These klotho deficiency-related changes can be blocked by eplerenone. Protein expression of scleraxis, a transcription factor for collagen synthesis, and LC3-II/LC3-I, an index of autophagy, were upregulated in aortas of klotho(+/-) mice, which can be abolished by eplerenone. In cultured mouse aortic smooth muscle cells, aldosterone increased collagen-1 expression that can be completely eliminated by small interfering RNA knockdown of scleraxis. Interestingly, aldosterone decreased elastin levels in smooth muscle cells, which can be abolished by small interfering RNA knockdown of Beclin-1, an autophagy-related gene. In conclusion, this study demonstrated for the first time that klotho deficiency-induced arterial stiffening may involve aldosterone-mediated upregulation of scleraxis and induction of autophagy, which led to increased collagen-1 expression and decreased elastin levels, respectively. © 2015 American Heart Association, Inc.

Soya protein stimulates bile acid excretion by the liver and intestine through direct and indirect pathways influenced by the presence of dietary cholesterol.

PubMed

Arellano-Martínez, Gloria Leticia; Granados, Omar; Palacios-González, Berenice; Torres, Nimbe; Medina-Vera, Isabel; Tovar, Armando R

2014-06-28

Several studies using different animal models have demonstrated that the consumption of soya protein (SP) reduces serum cholesterol concentrations by increasing the excretion of bile acids (BA). However, the mechanism by which SP enhances BA excretion is not fully understood. Therefore, the aim of the present study was to determine whether the consumption of SP regulates the expression of key enzymes involved in hepatic BA synthesis and the transporters involved in reverse cholesterol transport (RCT) via fibroblast growth factor 15 (FGF15) and/or small heterodimer protein (SHP) in rats. To achieve this aim, four groups of rats were fed experimental diets containing 20 % casein (C) or SP with or without the addition of 0·2 % cholesterol and the expression of hepatic genes involved in BA synthesis and the ileal and hepatic RCT was measured. Rats fed the SP diet had higher concentrations of ileal FGF15 and hepatic FGF15 receptor (FGFR4) and increased expression of SHP and liver receptor homolog 1 (LRH1) than those fed the C diet; as a result, the excretion of faecal BA was greater. The addition of cholesterol to the diet repressed the protein abundance of FGF15 and FGFR4; however, SP increased the expression of SHP and LRH1 to a lesser extent. Nonetheless, the expression of ABCG5/8 was increased in the intestine of rats fed the SP diet, and the effect was enhanced by the addition of cholesterol to the diet. In conclusion, SP in the presence of cholesterol increases BA synthesis via the repressions of FGF15 and SHP and accelerates BA excretion to prevent cholesterol overload in the enterocytes by increasing RCT.

Alteration in cardiac uncoupling proteins and eNOS gene expression following high-intensity interval training in favor of increasing mechanical efficiency

PubMed Central

Fallahi, Ali Asghar; Shekarfroush, Shahnaz; Rahimi, Mostafa; Jalali, Amirhossain; Khoshbaten, Ali

2016-01-01

Objective(s): High-intensity interval training (HIIT) increases energy expenditure and mechanical energy efficiency. Although both uncoupling proteins (UCPs) and endothelial nitric oxide synthase (eNOS) affect the mechanical efficiency and antioxidant capacity, their effects are inverse. The aim of this study was to determine whether the alterations of cardiac UCP2, UCP3, and eNOS mRNA expression following HIIT are in favor of increased mechanical efficiency or decreased oxidative stress. Materials and Methods: Wistar rats were divided into five groups: control group (n=12), HIIT for an acute bout (AT1), short term HIIT for 3 and 5 sessions (ST3 and ST5), long-term training for 8 weeks (LT) (6 in each group). The rats of the training groups were made to run on a treadmill for 60 min in three stages: 6 min running for warm-up, 7 intervals of 7 min running on treadmill with a slope of 5° to 20° (4 min with an intensity of 80-110% VO2max and 3 min at 50-60% VO2max), and 5-min running for cool-down. The control group did not participate in any exercise program. Rats were sacrificed and the hearts were extracted to analyze the levels of UCP2, UCP3 and eNOS mRNA by RT-PCR. Results: UCP3 expression was increased significantly following an acute training bout. Repeated HIIT for 8 weeks resulted in a significant decrease in UCPs mRNA and a significant increase in eNOS expression in cardiac muscle. Conclusion: This study indicates that Long term HIIT through decreasing UCPs mRNA and increasing eNOS mRNA expression may enhance energy efficiency and physical performance. PMID:27114795

VEGF improves survival of mesenchymal stem cells in infarcted hearts

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

Pons, Jennifer; Huang Yu; Arakawa-Hoyt, Janice

2008-11-14

Bone marrow-derived mesenchymal stem cells (MSC) are a promising source for cell-based treatment of myocardial infarction (MI), but existing strategies are restricted by low cell survival and engraftment. We examined whether vascular endothelial growth factor (VEGF) improve MSC viability in infracted hearts. We found long-term culture increased MSC-cellular stress: expressing more cell cycle inhibitors, p16{sup INK}, p21 and p19{sup ARF}. VEGF treatment reduced cellular stress, increased pro-survival factors, phosphorylated-Akt and Bcl-xL expression and cell proliferation. Co-injection of MSCs with VEGF to MI hearts increased cell engraftment and resulted in better improvement of cardiac function than that injected with MSCs ormore » VEGF alone. In conclusion, VEGF protects MSCs from culture-induce cellular stress and improves their viability in ischemic myocardium, which results in improvements of their therapeutic effect for the treatment of MI.« less

Increased TET1 Expression in Inflammatory Microenvironment of Hyperinsulinemia Enhances the Response of Endometrial Cancer to Estrogen by Epigenetic Modulation of GPER

PubMed Central

Lv, Qiao-Ying; Xie, Bing-Ying; Yang, Bing-Yi; Ning, Cheng-Cheng; Shan, Wei-Wei; Gu, Chao; Luo, Xue-Zhen; Chen, Xiao-Jun; Zhang, Zhen-Bo; Feng, You-Ji

2017-01-01

Background: Insulin resistance (IR) has been well studied in the initiation and development of endometrial endometrioid carcinoma (EEC). As yet, it has been largely neglected for estrogen sensitivity in local endometrium in hyperinsulinemia-induced systemic microenvironment. The aim of this study was to investigate the role of insulin in regulating estrogen sensitivity and explore the potential mechanisms in insulin-driven inflammatory microenvironment. Methods: We first investigated the effect of insulin on estradiol-driven endometrial cancer cells proliferation in vitro to address the roles of insulin in modulating estrogen sensitivity. Then GPER, ERα and TET1 in EEC samples with or without insulin resistance were screened by immunohistochemistry to confirm whether insulin resistance regulates estrogen receptors. Further mechanism analysis was carried out to address whether TET1 was mediated epigenetic modulation of GPER in insulin-induced microenvironment. Results: Insulin enhanced estradiol-driven endometrial cancer cells proliferation by up-regulating G-protein-coupled estrogen receptor (GPER) expression, but not ERα or ERβ. Immunohistochemistry of EEC tissues showed that GPER expression was greatly increased in endometrial tissues from EEC subjects with insulin resistance and was positively correlated with Ten-eleven-translocation 1 (TET1) expression. Mechanistically, insulin up-regulates TET1 expression, and the latter, an important DNA hydroxymethylase, could up-regulate GPER expression through epigenetic modulation. Conclusion: This study identified TET1 as the upstream regulator of GPER expression and provides a possible mechanism that insulin-induced positive regulation of estrogen sensitivity in endometrial cancer cells. Increasing expression of GPER through TET1-mediated epigenetic modulation may emerge as the main regulator to enhance the response of endometrial cancer to estrogen in insulin-driven inflammatory microenvironment. PMID:28382153

Increased TET1 Expression in Inflammatory Microenvironment of Hyperinsulinemia Enhances the Response of Endometrial Cancer to Estrogen by Epigenetic Modulation of GPER.

PubMed

Lv, Qiao-Ying; Xie, Bing-Ying; Yang, Bing-Yi; Ning, Cheng-Cheng; Shan, Wei-Wei; Gu, Chao; Luo, Xue-Zhen; Chen, Xiao-Jun; Zhang, Zhen-Bo; Feng, You-Ji

2017-01-01

Background: Insulin resistance (IR) has been well studied in the initiation and development of endometrial endometrioid carcinoma (EEC). As yet, it has been largely neglected for estrogen sensitivity in local endometrium in hyperinsulinemia-induced systemic microenvironment. The aim of this study was to investigate the role of insulin in regulating estrogen sensitivity and explore the potential mechanisms in insulin-driven inflammatory microenvironment. Methods: We first investigated the effect of insulin on estradiol-driven endometrial cancer cells proliferation in vitro to address the roles of insulin in modulating estrogen sensitivity. Then GPER, ERα and TET1 in EEC samples with or without insulin resistance were screened by immunohistochemistry to confirm whether insulin resistance regulates estrogen receptors. Further mechanism analysis was carried out to address whether TET1 was mediated epigenetic modulation of GPER in insulin-induced microenvironment. Results: Insulin enhanced estradiol-driven endometrial cancer cells proliferation by up-regulating G-protein-coupled estrogen receptor (GPER) expression, but not ERα or ERβ. Immunohistochemistry of EEC tissues showed that GPER expression was greatly increased in endometrial tissues from EEC subjects with insulin resistance and was positively correlated with Ten-eleven-translocation 1 (TET1) expression. Mechanistically, insulin up-regulates TET1 expression, and the latter, an important DNA hydroxymethylase, could up-regulate GPER expression through epigenetic modulation. Conclusion: This study identified TET1 as the upstream regulator of GPER expression and provides a possible mechanism that insulin-induced positive regulation of estrogen sensitivity in endometrial cancer cells. Increasing expression of GPER through TET1-mediated epigenetic modulation may emerge as the main regulator to enhance the response of endometrial cancer to estrogen in insulin-driven inflammatory microenvironment.

MicroRNA-30b-Mediated Regulation of Catalase Expression in Human ARPE-19 Cells

PubMed Central

Haque, Rashidul; Chun, Eugene; Howell, Jennifer C.; Sengupta, Trisha; Chen, Dan; Kim, Hana

2012-01-01

Background Oxidative injury to retinal pigment epithelium (RPE) and retinal photoreceptors has been linked to a number of retinal diseases, including age-related macular degeneration (AMD). Reactive oxygen species (ROS)-mediated gene expression has been extensively studied at transcriptional levels. Also, the post-transcriptional control of gene expression at the level of translational regulation has been recently reported. However, the microRNA (miRNA/miR)-mediated post-transcriptional regulation in human RPE cells has not been thoroughly looked at. Increasing evidence points to a potential role of miRNAs in diverse physiological processes. Methodology/Principal Findings We demonstrated for the first time in a human retinal pigment epithelial cell line (ARPE-19) that the post-transcriptional control of gene expression via miRNA modulation regulates human catalase, an important and potent component of cell's antioxidant defensive network, which detoxifies hydrogen peroxide (H2O2) radicals. Exposure to several stress-inducing agents including H2O2 has been reported to alter miRNA expression profile. Here, we demonstrated that a sublethal dose of H2O2 (200 µM) up-regulated the expression of miR-30b, a member of the miR-30 family, which inhibited the expression of endogenous catalase both at the transcript and protein levels. However, antisense (antagomirs) of miR-30b was not only found to suppress the miR-30b mimics-mediated inhibitions, but also to dramatically increase the expression of catalase even under an oxidant environment. Conclusions/Significance We propose that a microRNA antisense approach could enhance cytoprotective mechanisms against oxidative stress by increasing the antioxidant defense system. PMID:22880027

RELM-β promotes human pulmonary artery smooth muscle cell proliferation via FAK-stimulated surviving

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

Lin, Chunlong, E-mail: lclmd@sina.com; Li, Xiaohui; Luo, Qiong

Resistin-like molecule-β (RELM-β), focal adhesion kinase (FAK), and survivin may be involved in the proliferation of cultured human pulmonary artery smooth muscle cells (HPAMSCs), which is involved in pulmonary hypertension. HPAMSCs were treated with human recombinant RELM-β (rhRELM-β). siRNAs against FAK and survivin were transfected into cultured HPASMCs. Expression of FAK and survivin were examined by RT-PCR and western blot. Immunofluorescence was used to localize FAK. Flow cytometry was used to examine cell cycle distribution and cell death. Compared to the control group, all rhRELM-β-treated groups demonstrated significant increases in the expression of FAK and survivin (P<0.05). rhRELM-β significantly increasedmore » the proportion of HPASMCs in the S phase and decreased the proportion in G0/G1. FAK siRNA down-regulated survivin expression while survivin siRNA did not affect FAK expression. FAK siRNA effectively inhibited FAK and survivin expression in RELM-β-treated HPASMCs and partially suppressed cell proliferation. RELM-β promoted HPASMC proliferation and upregulated FAK and survivin expression. In conclusion, results suggested that FAK is upstream of survivin in the signaling pathway mediating cell proliferation. FAK seems to be important in RELM-β-induced HPASMC proliferation, partially by upregulating survivin expression. - Highlights: • rhRELM-β increased the expression of FAK and survivin. • rhRELM-β increased the proportion of HPASMCs in the S phase. • FAK is upstream of survivin in the signaling pathway mediating cell proliferation. • FAK is important in RELM-β-induced HPASMC proliferation, partly via survivin.« less

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

Mucin gene expression in the intestine of ulcerative colitis patients: a systematic review and meta-analysis.

PubMed

Niv, Yaron

2016-11-01

The mucus layer of the colon is the main barrier between luminal microbes and the mucosa, and plays a significant role in the body defense mechanisms. Several studies have examined mucin gene (MUC) expression in ulcerative colitis (UC) without conclusive results. The aim of the study was to establish the knowledge of mucin expression in UC as a basis for further investigation. English medical literature searches were performed for mucin expression in the colonic mucosa of UC patients in comparison with controls. Case-control studies were included. A meta-analysis was carried out using 'Comprehensive meta-analysis' software. Pooled odds ratios (ORs) and 95% confidence intervals were calculated. Altogether, we found 311 eligible studies. Only 10 case-control studies from five countries fulfilled the inclusion criteria. A moderate heterogeneity was found in the studies included: Q=52.703, d.f. (Q)=15.000, I=71.539%. OR for mucin expression in UC patients versus healthy controls was 1.868 with a 95% confidence interval (CI) 1.263-2.764, P=0.002. Thus, we could find a significant increase of 87% of mucin expression in UC patients. OR for MUC2 was 2.520, 95% CI 1.320-4.809, P<0.001. MUC3 was also increased with OR 2.599, 95% CI 1.389-4.861, P=0.003. Funnel plot did not indicate a significant publication bias. We found a global increase in mucin expression in UC patients, specifically in MUC2 and MUC3. Further studies are needed, especially in patients treated with biologics for mucosal healing, to understand the role of mucin expression in the natural history of UC.

ERAP1 Reduces Accumulation of Aberrant and Disulfide-Linked Forms of HLA-B27 on the Cell Surface

PubMed Central

Tran, Tri; Hong, Sohee; Edwan, Jehad; Colbert, Robert A.

2016-01-01

Objective Endoplasmic reticulum (ER) aminopeptidase 1 (ERAP1) variants contribute to the risk of ankylosing spondylitis in HLA-B27 positive individuals, implying a disease-related interaction between these gene products. The aim of this study was to determine whether reduced ERAP1 expression would alter the cell surface expression of HLA-B27 and the formation of aberrant disulfide-linked forms that have been implicated in the pathogenesis of spondyloarthritis. Methods ERAP1 expression was knocked down in monocytic U937 cells expressing HLA-B27 and endogenous HLA class I. The effect of ERAP1 knockdown on the accumulation HLA-B alleles (B18, B51, and B27) was assessed using immunoprecipitation, isoelectric focusing, and immunoblotting, as well as flow cytometry with antibodies specific for different forms of HLA-B27. Cell surface expression of aberrant disulfide-linked HLA-B27 dimers was assessed by immunoprecipitation and electrophoresis on non-reducing polyacrylamide gels. Results ERAP1 knockdown increased the accumulation of HLA-B27 on the cell surface including disulfide-linked dimers, but had no effect on levels of HLA-B18 or -B51. Antibodies with unique specificity for HLA-B27 confirmed increased cell surface expression of complexes shown previously to contain long peptides. IFN-γ treatment resulted in striking increases in the expression of disulfide-linked HLA-B27 heavy chains, even in cells with normal ERAP1 expression. Conclusions Our results suggest that normal levels of ERAP1 reduce the accumulation of aberrant and disulfide-linked forms of HLA-B27 in monocytes, and thus help to maintain the integrity of cell surface HLA-B27 complexes. PMID:27107845

Immunohistochemichal Assessment of the CrkII Proto-oncogene Expression in Common Malignant Salivary Gland Tumors and Pleomorphic Adenoma.

PubMed

Askari, Mitra; Darabi, Masoud; Jahanzad, Esa; Mostakhdemian Hosseini, Zahra; Musavi Chavoshi, Marjan; Darabi, Maryam

2015-01-01

Background and aims. Various morphologies are seen in different salivary gland tumorsor within an individual tumor, and the lesions show divers biological behaviors. Experimental results support the hypothesis that increased CrkII proto-oncogene is associated with cytokine-induced tumor initiation and progression by altering cell motility signaling pathway. The aim of this study was to assess the CrkII expression in common malignant salivary gland tumors and pleomorphic ade-noma. Materials and methods. Immunohistochemical analysis of CrkII expression was performed on paraffin blocks of 64 car-cinomas of salivary glands, 10 pleomorphic adenomas, and 10 normal salivary glands. Biopsies were subjected to immu-nostaining with EnVision detection system using monoclonal anti-CrkII. Evaluation of immunoreactivity of CrkII was based on the immunoreaction intensity and percentage of stained tumor cells which were scored semi-quantitatively on a scale with four grades 0 to 3. Kruskal-wallis test and additional Mann-Whitney statistical test were used for analysis of CrkII expression levels. Results. Increased expression of CrkII was seen (P=0.005) in malignant tumors including: mucoepidermoid carcinoma, adenoid cystic carcinoma, and carcinoma ex pleomorphic adenoma, but CrkII expression in acinic cell carcinoma was weak. CrkII expression in pleomorphic adenoma was weak or negative. A weak staining was sparsely seen in normal acinar serous cell. Conclusion. Increased expression of CrkII and its higher intensity of staining in tumors with more aggressive biologic behavior in carcinomas of salivary gland is consistent with a role for this proto-oncogene in salivary gland tumorigenesis and cancer progression.

Overexpression of hyaluronan synthase 2 and gonadotropin receptors in cumulus cells of goats subjected to one-shot eCG/FSH hormonal treatment for ovarian stimulation.

PubMed

Santos, Juliana D R; Batista, Ribrio I T P; Magalhães, Livia C; Paula, Alexandre R; Souza, Samara S; Salamone, Daniel F; Bhat, Maajid H; Teixeira, Dárcio I A; Freitas, Vicente J F; Melo, Luciana M

2016-07-01

Hormonal ovarian stimulation may affect transcripts in somatic cells of cumulus-oocyte complexes (COCs) and affect the resulting oocyte quality. Here, in parallel with morphological classification and in vitro maturation (IVM) rate analysis, we investigated the expression of hyaluronan synthase 2 (HAS2), gonadotropic receptors (FSHR and LHR) and connexin 43 (GJA1) in cumulus cells (CCs) from goat COCs after multi-dose FSH (MD) or one-shot FSH/eCG (OS) treatments, using bovine COCs as control groups. The MD treatment produced more large follicles, and the resulting COCs had a better morphology and IVM rate than were obtained with OS. The OS treatment produced COCs with increased HAS2, FSHR, LHR and GJA1 expression. This gene expression pattern was also observed in the CCs of COCs that showed poor morphological characteristics. On the other hand, the mRNA levels were more similar between groups after IVM; FSHR and LHR were the main genes that showed decreased expression. Some events that occurred in bovine CCs during IVM, such as cell expansion, increased HAS2 expression and decreased GJA1 expression, were less evident or did not occur in goat COCs. In conclusion, increasing HAS2, FSHR, LHR and GJA1 expression in goat COCs does not confer greater meiotic competence to oocytes. Instead, it may result from poor regulation of gene expression in CCs by lower quality oocytes. Finally, cumulus expansion, together with HAS2 upregulation and GJA1 downregulation, seems to be more important for bovine COCs than for goat COCs. Additional studies are needed to investigate the importance of other HAS isoforms and connexins in goat COCs. Copyright © 2016 Elsevier B.V. All rights reserved.

GDF-15 is abundantly expressed in plexiform lesions in patients with pulmonary arterial hypertension and affects proliferation and apoptosis of pulmonary endothelial cells

PubMed Central

2011-01-01

Background Growth-differentiation factor-15 (GDF-15) is a stress-responsive, transforming growth factor-β-related cytokine, which has recently been reported to be elevated in serum of patients with idiopathic pulmonary arterial hypertension (IPAH). The aim of the study was to examine the expression and biological roles of GDF-15 in the lung of patients with pulmonary arterial hypertension (PAH). Methods GDF-15 expression in normal lungs and lung specimens of PAH patients were studied by real-time RT-PCR and immunohistochemistry. Using laser-assisted micro-dissection, GDF-15 expression was further analyzed within vascular compartments of PAH lungs. To elucidate the role of GDF-15 on endothelial cells, human pulmonary microvascular endothelial cells (HPMEC) were exposed to hypoxia and laminar shear stress. The effects of GDF-15 on the proliferation and cell death of HPMEC were studied using recombinant GDF-15 protein. Results GDF-15 expression was found to be increased in lung specimens from PAH patients, com-pared to normal lungs. GDF-15 was abundantly expressed in pulmonary vascular endothelial cells with a strong signal in the core of plexiform lesions. HPMEC responded with marked upregulation of GDF-15 to hypoxia and laminar shear stress. Apoptotic cell death of HPMEC was diminished, whereas HPMEC proliferation was either increased or decreased depending of the concentration of recombinant GDF-15 protein. Conclusions GDF-15 expression is increased in PAH lungs and appears predominantly located in vascular endothelial cells. The expression pattern as well as the observed effects on proliferation and apoptosis of pulmonary endothelial cells suggest a role of GDF-15 in the homeostasis of endothelial cells in PAH patients. PMID:21548946

MYC, FBXW7 and TP53 copy number variation and expression in Gastric Cancer

PubMed Central

2013-01-01

Background MYC deregulation is a common event in gastric carcinogenesis, usually as a consequence of gene amplification, chromosomal translocations, or posttranslational mechanisms. FBXW7 is a p53-controlled tumor-suppressor that plays a role in the regulation of cell cycle exit and reentry via MYC degradation. Methods We evaluated MYC, FBXW7, and TP53 copy number, mRNA levels, and protein expression in gastric cancer and paired non-neoplastic specimens from 33 patients and also in gastric adenocarcinoma cell lines. We also determined the invasion potential of the gastric cancer cell lines. Results MYC amplification was observed in 51.5% of gastric tumor samples. Deletion of one copy of FBXW7 and TP53 was observed in 45.5% and 21.2% of gastric tumors, respectively. MYC mRNA expression was significantly higher in tumors than in non-neoplastic samples. FBXW7 and TP53 mRNA expression was markedly lower in tumors than in paired non-neoplastic specimens. Moreover, deregulated MYC and FBXW7 mRNA expression was associated with the presence of lymph node metastasis and tumor stage III-IV. Additionally, MYC immunostaining was more frequently observed in intestinal-type than diffuse-type gastric cancers and was associated with MYC mRNA expression. In vitro studies showed that increased MYC and reduced FBXW7 expression is associated with a more invasive phenotype in gastric cancer cell lines. This result encouraged us to investigate the activity of the gelatinases MMP-2 and MMP-9 in both cell lines. Both gelatinases are synthesized predominantly by stromal cells rather than cancer cells, and it has been proposed that both contribute to cancer progression. We observed a significant increase in MMP-9 activity in ACP02 compared with ACP03 cells. These results confirmed that ACP02 cells have greater invasion capability than ACP03 cells. Conclusion In conclusion, FBXW7 and MYC mRNA may play a role in aggressive biologic behavior of gastric cancer cells and may be a useful indicator of poor prognosis. Furthermore, MYC is a candidate target for new therapies against gastric cancer. PMID:24053468

Increasing the Performance of a Sliding Discharge Actuator Through the Application of Multiple Potentials

DTIC Science & Technology

2011-09-01

75 4.5 Summary . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . 85 vi Page V. Conclusions...Map Results . . . . . . . . . . . . . . . . 75 4.24 Case 13 Velocity Contour Map Results . . . . . . . . . . . . . . . . 76 ix Figure Page 4.25 Case 14...expressed in this materail are those of the author and do not necessarily reflect the views of AFOSR. LCDR Steven D Seney Jr v Table of Contents Page

Emodin enhances the chemosensitivity of endometrial cancer by inhibiting ROS-mediated Cisplatin-resistance.

PubMed

Ding, Ning; Zhang, Hong; Su, Shan; Ding, Yumei; Yu, Xiaohui; Tang, Yujie; Wang, Qingfang; Liu, Peishu

2017-12-18

Background Endometrial cancer is a common cause of death in gynecological malignancies. Cisplatin is a clinically chemotherapeutic agent. However, drug-resistance is the primary cause of treatment failure. Objective Emodin is commonly used clinically to increase the sensitivity of chemotherapeutic agents, yet whether Emodin promotes the role of Cisplatin in the treatment of endometrial cancer has not been studied. Method CCK-8 kit was utilized to determine the growth of two endometrial cancer cell lines, Ishikawa and HEC-IB. The apoptosis level of Ishikawa and HEC-IB cells was detected by Annexin V / propidium iodide double-staining assay. ROS level was detected by DCFH-DA and NADPH oxidase expression. Expressions of drug-resistant genes were examined by real-time PCR and Western blotting. Results Emodin combined with Cisplatin reduced cell growth and increased the apoptosis of endometrial cancer cells. Co-treatment of Emodin and Cisplatin increased chemosensitivity by inhibiting the expression of drug-resistant genes through reducing the ROS levels in endometrial cancer cells. In an endometrial cancer xenograft murine model, the tumor size was reduced and animal survival time was increased by co-treatment of Emodin and Cisplatin. Conclusion This study demonstrates that Emodin enhances the chemosensitivity of Cisplatin on endometrial cancer by inhibiting ROS-mediated expression of drug-resistance genes. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

Possible Mitochondria-Associated Enzymatic Role in Non-Hodgkin Lymphoma Residual Disease

PubMed Central

Kusao, Ian; Troelstrup, David; Shiramizu, Bruce

2009-01-01

Background The mechanisms responsible for resistant or recurrent disease in childhood non-Hodgkin lymphoma (NHL) are not yet fully understood. A unique mechanism suggesting the role of the mitochondria as the key energy source responsible for residual cells has been assessed in the clinical setting on specimens from patients on therapy were found to have increased copies of mitochondrial DNA (mtDNA) associated with positive minimal residual disease and/or persistent disease (MRD/PD) status. The potential role of mtDNA in MRD/PD emphasizes queries into the contributions of relevant enzymatic pathways responsible for MRD/PD. This study hypothesized that in an in-vitro model, recovering or residual cells from chemotoxicity will exhibit an increase in both citrate synthase and isocitrate dehydrogenase expression and decrease in succinate dehydrogenase expression. Procedure Ramos cells (Burkitt lymphoma cell line) were exposed to varying concentrations of doxorubicin and vincristine for 1 hr; and allowing for recovery in culture over a 7-day period. cDNA was extracted on days 1 and 7 of the cell culture period to assess the relative expression of the aforementioned genes. Results Increase citrate synthase, increase isocitrate dehydrogenase and decrease succinate dehydrogenase expressions were found in recovering Ramos cells. Conclusion Recovering lymphoma cells appear to compensate by regulating enzymatic levels of appropriate genes in the Krebs Cycle suggesting an important role of the mitochondria in the presence of residual cells. PMID:19936279

Effect of low oxygen tension on the biological characteristics of human bone marrow mesenchymal stem cells.

PubMed

Kim, Dae Seong; Ko, Young Jong; Lee, Myoung Woo; Park, Hyun Jin; Park, Yoo Jin; Kim, Dong-Ik; Sung, Ki Woong; Koo, Hong Hoe; Yoo, Keon Hee

2016-11-01

Culture of mesenchymal stem cells (MSCs) under ambient conditions does not replicate the low oxygen environment of normal physiological or pathological states and can result in cellular impairment during culture. To overcome these limitations, we explored the effect of hypoxia (1 % O 2 ) on the biological characteristics of MSCs over the course of different culture periods. The following biological characteristics were examined in human bone marrow-derived MSCs cultured under hypoxia for 8 weeks: proliferation rate, morphology, cell size, senescence, immunophenotypic characteristics, and the expression levels of stemness-associated factors and cytokine and chemokine genes. MSCs cultured under hypoxia for approximately 2 weeks showed increased proliferation and viability. During long-term culture, hypoxia delayed phenotypic changes in MSCs, such as increased cell volume, altered morphology, and the expression of senescence-associated-β-gal, without altering their characteristic immunophenotypic characteristics. Furthermore, hypoxia increased the expression of stemness and chemokine-related genes, including OCT4 and CXCR7, and did not decrease the expression of KLF4, C-MYC, CCL2, CXCL9, CXCL10, and CXCR4 compared with levels in cells cultured under normoxia. In conclusion, low oxygen tension improved the biological characteristics of MSCs during ex vivo expansion. These data suggest that hypoxic culture could be a useful method for increasing the efficacy of MSC cell therapies.

Altered Left Ventricular Ion Channel Transcriptome in a High-Fat-Fed Rat Model of Obesity: Insight into Obesity-Induced Arrhythmogenesis

PubMed Central

Yon, Marianne; Pickavance, Lucy; Yanni Gerges, Joseph; Davis, Gershan; Wilding, John; Jian, Kun; Hart, George; Boyett, Mark

2016-01-01

Introduction. Obesity is increasingly common and is associated with an increased prevalence of cardiac arrhythmias. The aim of this study was to see whether in obesity there is proarrhythmic gene expression of ventricular ion channels and related molecules. Methods and Results. Rats were fed on a high-fat diet and compared to control rats on a normal diet (n = 8). After 8 weeks, rats on the high-fat diet showed significantly greater weight gain and higher adiposity. Left ventricle samples were removed at 8 weeks and mRNA expression of ion channels and other molecules was measured using qPCR. Obese rats had significant upregulation of Cav1.2, HCN4, Kir2.1, RYR2, NCX1, SERCA2a, and RYR2 mRNA and downregulation of ERG mRNA. In the case of HCN4, it was confirmed that there was a significant increase in protein expression. The potential effects of the mRNA changes on the ventricular action potential and intracellular Ca2+ transient were predicted using computer modelling. Modelling predicted prolongation of the ventricular action potential and an increase in the intracellular Ca2+ transient, both of which would be expected to be arrhythmogenic. Conclusion. High-fat diet causing obesity results in arrhythmogenic cardiac gene expression of ion channels and related molecules. PMID:27747100

Cinnamaldehyde supplementation prevents fasting-induced hyperphagia, lipid accumulation, and inflammation in high-fat diet-fed mice.

PubMed

Khare, Pragyanshu; Jagtap, Sneha; Jain, Yachna; Baboota, Ritesh K; Mangal, Priyanka; Boparai, Ravneet K; Bhutani, Kamlesh K; Sharma, Shyam S; Premkumar, Louis S; Kondepudi, Kanthi K; Chopra, Kanwaljit; Bishnoi, Mahendra

2016-01-01

Cinnamaldehyde, a bioactive component of cinnamon, is increasingly gaining interest for its preventive and therapeutic effects against metabolic complications like type-2 diabetes. This study is an attempt to understand the effect of cinnamaldehyde in high-fat diet (HFD)-associated increase in fasting-induced hyperphagia and related hormone levels, adipose tissue lipolysis and inflammation, and selected cecal microbial count in mice. Cinnamaldehyde, at 40 µM dose, prevented lipid accumulation and altered gene expression toward lipolytic phenotype in 3T3-L1 preadipocyte cell lines. In vivo, cinnamaldehyde coadministration prevented HFD-induced body weight gain, decreased fasting-induced hyperphagia, as well as circulating leptin and leptin/ghrelin ratio. In addition to that, cinnamaldehyde altered serum biochemical parameters related to lipolysis, that is, glycerol and free fatty acid levels. At transcriptional level, cinnamaldehyde increased anorectic gene expression in hypothalamus and lipolytic gene expression in visceral white adipose tissue. Furthermore, cinnamaldehyde also decreased serum IL-1β and inflammatory gene expression in visceral white adipose tissue. However, cinnamaldehyde did not modulate the population of selected gut microbial (Lactobacillus, Bifidibaceria, and Roseburia) count in cecal content. In conclusion, cinnamaldehyde increased adipose tissue lipolysis, decreased fasting-induced hyperphagia, normalized circulating levels of leptin/ghrelin ratio, and reduced inflammation in HFD-fed mice, which augurs well for its antiobesity role. © 2016 International Union of Biochemistry and Molecular Biology.

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

PubMed

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

2017-01-01

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

Chronic Vitamin C Deficiency Promotes Redox Imbalance in the Brain but Does Not Alter Sodium-Dependent Vitamin C Transporter 2 Expression

PubMed Central

Paidi, Maya D.; Schjoldager, Janne G.; Lykkesfeldt, Jens; Tveden-Nyborg, Pernille

2014-01-01

Vitamin C (VitC) has several roles in the brain acting both as a specific and non-specific antioxidant. The brain upholds a very high VitC concentration and is able to preferentially retain VitC even during deficiency. The accumulation of brain VitC levels much higher than in blood is primarily achieved by the sodium dependent VitC transporter (SVCT2). This study investigated the effects of chronic pre-and postnatal VitC deficiency as well as the effects of postnatal VitC repletion, on brain SVCT2 expression and markers of oxidative stress in young guinea pigs. Biochemical analyses demonstrated significantly decreased total VitC and an increased percentage of dehydroascorbic acid, as well as increased lipid oxidation (malondialdehyde), in the brains of VitC deficient animals (p < 0.0001) compared to controls. VitC repleted animals were not significantly different from controls. No significant changes were detected in either gene or protein expression of SVCT2 between groups or brain regions. In conclusion, chronic pre-and postnatal VitC deficiency increased brain redox imbalance but did not increase SVCT2 expression. Our findings show potential implications for VitC deficiency induced negative effects of redox imbalance in the brain and provide novel insight to the regulation of VitC in the brain during deficiency. PMID:24787032

Increased Expression of Fatty-Acid and Calcium Metabolism Genes in Failing Human Heart

PubMed Central

Rodríguez-Penas, Diego; Feijóo-Bandín, Sandra; Noguera-Moreno, Teresa; Calaza, Manuel; Álvarez-Barredo, María; Mosquera-Leal, Ana; Parrington, John; Brugada, Josep; Portolés, Manuel; Rivera, Miguel; González-Juanatey, José Ramón; Lago, Francisca

2012-01-01

Background Heart failure (HF) involves alterations in metabolism, but little is known about cardiomyopathy-(CM)-specific or diabetes-independent alterations in gene expression of proteins involved in fatty-acid (FA) uptake and oxidation or in calcium-(Ca2+)-handling in the human heart. Methods RT-qPCR was used to quantify mRNA expression and immunoblotting to confirm protein expression in left-ventricular myocardium from patients with HF (n = 36) without diabetes mellitus of ischaemic (ICM, n = 16) or dilated (DCM, n = 20) cardiomyopathy aetiology, and non-diseased donors (CTL, n = 6). Results Significant increases in mRNA of genes regulating FA uptake (CD36) and intracellular transport (Heart-FA-Binding Protein (HFABP)) were observed in HF patients vs CTL. Significance was maintained in DCM and confirmed at protein level, but not in ICM. mRNA was higher in DCM than ICM for peroxisome-proliferator-activated-receptor-alpha (PPARA), PPAR-gamma coactivator-1-alpha (PGC1A) and CD36, and confirmed at the protein level for PPARA and CD36. Transcript and protein expression of Ca2+-handling genes (Two-Pore-Channel 1 (TPCN1), Two-Pore-Channel 2 (TPCN2), and Inositol 1,4,5-triphosphate Receptor type-1 (IP3R1)) increased in HF patients relative to CTL. Increases remained significant for TPCN2 in all groups but for TPCN1 only in DCM. There were correlations between FA metabolism and Ca2+-handling genes expression. In ICM there were six correlations, all distinct from those found in CTL. In DCM there were also six (all also different from those found in CTL): three were common to and three distinct from ICM. Conclusion DCM-specific increases were found in expression of several genes that regulate FA metabolism, which might help in the design of aetiology-specific metabolic therapies in HF. Ca2+-handling genes TPCN1 and TPCN2 also showed increased expression in HF, while HF- and CM-specific positive correlations were found among several FA and Ca2+-handling genes. PMID:22701570

Differential expression of type X collagen in a mechanically active 3-D chondrocyte culture system: a quantitative study

PubMed Central

Yang, Xu; Vezeridis, Peter S; Nicholas, Brian; Crisco, Joseph J; Moore, Douglas C; Chen, Qian

2006-01-01

Objective Mechanical loading of cartilage influences chondrocyte metabolism and gene expression. The gene encoding type X collagen is expressed specifically by hypertrophic chondrocytes and up regulated during osteoarthritis. In this study we tested the hypothesis that the mechanical microenvironment resulting from higher levels of local strain in a three dimensional cell culture construct would lead to an increase in the expression of type X collagen mRNA by chondrocytes in those areas. Methods Hypertrophic chondrocytes were isolated from embryonic chick sterna and seeded onto rectangular Gelfoam sponges. Seeded sponges were subjected to various levels of cyclic uniaxial tensile strains at 1 Hz with the computer-controlled Bio-Stretch system. Strain distribution across the sponge was quantified by digital image analysis. After mechanical loading, sponges were cut and the end and center regions were separated according to construct strain distribution. Total RNA was extracted from the cells harvested from these regions, and real-time quantitative RT-PCR was performed to quantify mRNA levels for type X collagen and a housing-keeping gene 18S RNA. Results Chondrocytes distributed in high (9%) local strain areas produced more than two times type X collagen mRNA compared to the those under no load conditions, while chondrocytes located in low (2.5%) local strain areas had no appreciable difference in type X collagen mRNA production in comparison to non-loaded samples. Increasing local strains above 2.5%, either in the center or end regions of the sponge, resulted in increased expression of Col X mRNA by chondrocytes in that region. Conclusion These findings suggest that the threshold of chondrocyte sensitivity to inducing type X collagen mRNA production is more than 2.5% local strain, and that increased local strains above the threshold results in an increase of Col X mRNA expression. Such quantitative analysis has important implications for our understanding of mechanosensitivity of cartilage and mechanical regulation of chondrocyte gene expression. PMID:17150098

The antiproliferative effect of Moringa oleifera crude aqueous leaf extract on cancerous human alveolar epithelial cells

PubMed Central

2013-01-01

Background The incidence of lung cancer is expected to increase due to increases in exposure to airborne pollutants and cigarette smoke. Moringa oleifera (MO), a medicinal plant found mainly in Asia and South Africa is used in the traditional treatment of various ailments including cancer. This study investigated the antiproliferative effect of MO leaf extract (MOE) in cancerous A549 lung cells. Methods A crude aqueous leaf extract was prepared and the cells were treated with 166.7 μg/ml MOE (IC50) for 24 h and assayed for oxidative stress (TBARS and Glutathione assays), DNA fragmentation (comet assay) and caspase (3/7 and 9) activity. In addition, the expression of Nrf2, p53, Smac/DIABLO and PARP-1 was determined by Western blotting. The mRNA expression of Nrf2 and p53 was assessed using qPCR. Results A significant increase in reactive oxygen species with a concomitant decrease in intracellular glutathione levels (p < 0.001) in MOE treated A549 cells was observed. MOE showed a significant reduction in Nrf2 protein expression (1.89-fold, p < 0.05) and mRNA expression (1.44-fold). A higher level of DNA fragmentation (p < 0.0001) was seen in the MOE treated cells. MOE’s pro-apoptotic action was confirmed by the significant increase in p53 protein expression (1.02-fold, p < 0.05), p53 mRNA expression (1.59-fold), caspase-9 (1.28-fold, p < 0.05), caspase-3/7 (1.52-fold) activities and an enhanced expression of Smac/DIABLO. MOE also caused the cleavage and activation of PARP-1 into 89 KDa and 24 KDa fragments (p < 0.0001). Conclusion MOE exerts antiproliferative effects in A549 lung cells by increasing oxidative stress, DNA fragmentation and inducing apoptosis. PMID:24041017

Low-level overexpression of p53 promotes warfarin-induced calcification of porcine aortic valve interstitial cells by activating Slug gene transcription.

PubMed

Gao, Li; Ji, Yue; Lu, Yan; Qiu, Ming; Shen, Yejiao; Wang, Yaqing; Kong, Xiangqing; Shao, Yongfeng; Sheng, Yanhui; Sun, Wei

2018-03-09

The most frequently used oral anti-coagulant warfarin has been implicated in inducing calcification of aortic valve interstitial cells (AVICs), whereas the mechanism is not fully understood. The low-level activation of p53 is found to be involved in osteogenic transdifferentiation and calcification of AVICs. Whether p53 participates in warfarin-induced AVIC calcification remains unknown. In this study, we investigated the role of low-level p53 overexpression in warfarin-induced porcine AVIC (pAVIC) calcification. Immunostaining, quantitative PCR, and Western blotting revealed that p53 was expressed in human and pAVICs and that p53 expression was slightly increased in calcific human aortic valves compared with non-calcific valves. Terminal deoxynucleotidyltransferase-mediated dUTP nick end labeling staining indicated that apoptosis slightly increased in calcific aortic valves than in non-calcific valves. Warfarin treatment led to a low-level increase of p53 mRNA and protein in both pAVICs and mouse aortic valves. Low-level overexpression of p53 in pAVICs via an adenovirus vector did not affect pAVIC apoptosis but promoted warfarin-induced calcium deposition and expression of osteogenic markers. shRNA-mediated p53 knockdown attenuated the pAVIC calcium deposition and osteogenic marker expression. Moreover, ChIP and luciferase assays showed that p53 was recruited to the slug promoter and activated slug expression in calcific pAVICs. Of note, overexpression of Slug increased osteogenic marker Runx2 expression, but not pAVIC calcium deposition, and Slug knockdown attenuated pAVIC calcification and p53-mediated pAVIC calcium deposition and expression of osteogenic markers. In conclusion, we found that p53 plays an important role in warfarin induced pAVIC calcification, and increased slug transcription by p53 is required for p53-mediated pAVIC calcification. © 2018 by The American Society for Biochemistry and Molecular Biology, Inc.

Monoamine oxidase A expression is suppressed in human cholangiocarcinoma via coordinated epigenetic and IL-6-driven events

PubMed Central

Huang, Li; Frampton, Gabriel; Rao, Arundhati; Zhang, Kun-song; Chen, Wei; Lai, Jia-ming; Yin, Xiao-yu; Walker, Kimberly; Culbreath, Brianne; Leyva-Illades, Dinorah; Quinn, Matthew; McMillin, Matthew; Bradley, Michelle; Liang, Li-Jian; DeMorrow, Sharon

2014-01-01

Objectives The secretion of dopamine and serotonin is increased in cholangiocarcinoma, which has growth-promoting effects. Monoamine oxidase A (MAOA), the degradation enzyme of serotonin and dopamine, is suppressed in cholangiocarcinoma via an unknown mechanism. The aims of this study were to (i) correlate MAOA immunoreactivity with pathophysiological parameters of cholangiocarcinoma, (ii) determine the mechanism by which MAOA expression is suppressed and (iii) evaluate the consequences of restored MAOA expression in cholangiocarcinoma. Design MAOA expression was assessed in cholangiocarcinoma and non-malignant controls. The control of MAOA expression by promoter hypermethylation was evaluated and the contribution of IL-6 signaling to the suppression of MAOA expression was determined. The effects of MAOA overexpression on cholangiocarcinoma growth and invasion were also assessed. Results MAOA expression is correlated with differentiation, invasion and survival in cholangiocarcinoma. The MAOA promoter was hypermethylated immediately upstream of the start codon in cholangiocarcinoma samples and cell lines but not in non-malignant counterparts. IL-6 signaling also decreased MAOA expression via a mechanism independent of hypermethylation, involving the regulation of the balance between SP-1 transcriptional activity and its inhibitor, R1 repressor. Inhibition of both IL-6 signaling and DNA methylation restored MAOA levels to those observed in cholangiocytes. Forced MAOA overexpression inhibited cholangiocarcinoma growth and invasion. Conclusions MAOA expression is suppressed by the coordinated control of promoter hypermethylation and IL-6 signaling. MAOA may be a useful prognostic marker in the management of cholangiocarcinoma, and therapies designed to increase MAOA expression might prove beneficial in the treatment of cholangiocarcinoma. PMID:22906985

Exposure to radio-frequency electromagnetic waves alters acetylcholinesterase gene expression, exploratory and motor coordination-linked behaviour in male rats.

PubMed

Obajuluwa, Adejoke Olukayode; Akinyemi, Ayodele Jacob; Afolabi, Olakunle Bamikole; Adekoya, Khalid; Sanya, Joseph Olurotimi; Ishola, Azeez Olakunle

2017-01-01

Humans in modern society are exposed to an ever-increasing number of electromagnetic fields (EMFs) and some studies have demonstrated that these waves can alter brain function but the mechanism still remains unclear. Hence, this study sought to investigate the effect of 2.5 Ghz band radio-frequency electromagnetic waves (RF-EMF) exposure on cerebral cortex acetylcholinesterase (AChE) activity and their mRNA expression level as well as locomotor function and anxiety-linked behaviour in male rats. Animals were divided into four groups namely; group 1 was control (without exposure), group 2-4 were exposed to 2.5 Ghz radiofrequency waves from an installed WI-FI device for a period of 4, 6 and 8 weeks respectively. The results revealed that WiFi exposure caused a significant increase in anxiety level and affect locomotor function. Furthermore, there was a significant decrease in AChE activity with a concomitant increase in AChE mRNA expression level in WiFi exposed rats when compared with control. In conclusions, these data showed that long term exposure to WiFi may lead to adverse effects such as neurodegenerative diseases as observed by a significant alteration on AChE gene expression and some neurobehavioral parameters associated with brain damage.

IL-17 Promotes Angiogenic Factors IL-6, IL-8, and Vegf Production via Stat1 in Lung Adenocarcinoma.

PubMed

Huang, Qi; Duan, Limin; Qian, Xin; Fan, Jinshuo; Lv, Zhilei; Zhang, Xiuxiu; Han, Jieli; Wu, Feng; Guo, Mengfei; Hu, Guorong; Du, Jiao; Chen, Caiyun; Jin, Yang

2016-11-07

Inflammation and angiogenesis are two hallmarks of carcinoma. The proinflammatory cytokine interleukin-17 (IL-17) facilitates angiogenesis in lung cancer; however, the underlying mechanism is not fully understood. In this study, tumour microvessel density (MVD) was positively associated with IL-17, interleukin-6 (IL-6), interleukin-8 (IL-8), and vascular endothelial cell growth factor (VEGF) expression in human lung adenocarcinoma tissues, and it was increased in tumour tissues of A549-IL-17 cell-bearing nude mice. Importantly, positive correlations were also detected between IL-17 expression and IL-6, IL-8 and VEGF expression in human lung adenocarcinoma tissues. Furthermore, IL-6, IL-8 and VEGF production, as well as STAT1 phosphorylation, were increased in tumour tissues of A549-IL-17 cell-bearing nude mice in vivo and in A549 and H292 cells following IL-17 stimulation in vitro. In addition, STAT1 knockdown using an inhibitor and siRNA attenuated the IL-17-mediated increases in IL-6, IL-8 and VEGF expression in A549 and H292 cells. In conclusion, IL-17 may promote the production of the angiogenic inducers IL-6, IL-8 and VEGF via STAT1 signalling in lung adenocarcinoma.

Down-regulation of miR-181a can reduce heat stress damage in PBMCs of Holstein cows.

PubMed

Chen, Kun-Lin; Fu, Yuan-Yuan; Shi, Min-Yan; Li, Hui-Xia

2016-09-01

Heat stress can weaken the immune system and even increase livestock's susceptibility to disease. MicroRNA (miR) is short non-coding RNA that functions in post-transcriptional regulation of gene expression and some phenotypes. Our recent study found that miR-181a is highly expressed in the serum of heat-stressed Holstein cows, but the potential function of miR-181a is still not clarified. In this study, peripheral blood mononuclear cells (PBMCs), isolated from Holstein cows' peripheral blood, were used to investigate the effects of miR-181a inhibitor on heat stress damage. Our results showed that significant apoptosis and oxidative damage were induced by heat stress in PBMCs. However, with apoptosis, the levels of reactive oxygen species (ROS) and content of malondialdehyde (MDA) were reduced, while the content of glutathione (GSH) and the activity of superoxide dismutase (SOD) were increased even under heat stress conditions after transfecting miR-181a inhibitors to PBMCs. Meanwhile, mRNA expression of bax and caspase-3 was significantly decreased, but mRNA expression of bcl-2 was increased in transfected PBMCs. In conclusion, our results demonstrated that down-regulation of miR-181a can reduce heat stress damage in PBMCs of Holstein cows.

Effects of nerve growth factor (NGF) on blood vessels area and expression of the angiogenic factors VEGF and TGFbeta1 in the rat ovary

PubMed Central

Julio-Pieper, Marcela; Lara, Hernán E; Bravo, Javier A; Romero, Carmen

2006-01-01

Background Angiogenesis is a crucial process in follicular development and luteogenesis. The nerve growth factor (NGF) promotes angiogenesis in various tissues. An impaired production of this neurotrophin has been associated with delayed wound healing. A variety of ovarian functions are regulated by NGF, but its effects on ovarian angiogenesis remain unknown. The aim of this study was to elucidate if NGF modulates 1) the amount of follicular blood vessels and 2) ovarian expression of two angiogenic factors: vascular endothelial growth factor (VEGF) and transforming growth factor beta 1 (TGFbeta1), in the rat ovary. Results In cultured neonatal rat ovaries, NGF increased VEGF mRNA and protein levels, whereas TGFbeta1 expression did not change. Sectioning of the superior ovarian nerve, which increases ovarian NGF protein content, augmented VEGF immunoreactivity and the area of capillary vessels in ovaries of prepubertal rats compared to control ovaries. Conclusion Results indicate that NGF may be important in the maintenance of the follicular and luteal vasculature in adult rodents, either indirectly, by increasing the expression of VEGF in the ovary, or directly via promoting the proliferation of vascular cells. This data suggests that a disruption on NGF regulation could be a component in ovarian disorders related with impaired angiogenesis. PMID:17096853

Evaluation of the reversal of multidrug resistance by MDR1 ribonucleic acid interference in a human colon cancer model using a Renilla luciferase reporter gene and coelenterazine.

PubMed

Jeon, Yong Hyun; Bae, Seon-ae; Lee, Yong Jin; Lee, You La; Lee, Sang-Woo; Yoon, Ghil-Suk; Ahn, Byeong-Cheol; Ha, Jeoung-Hee; Lee, Jaetae

2010-12-01

The reversal effect of multidrug resistance (MDR1) gene expression by adenoviral vector-mediated MDR1 ribonucleic acid interference was assessed in a human colon cancer animal model using bioluminescent imaging with Renilla luciferase (Rluc) gene and coelenterazine, a substrate for Rluc or MDR1 gene expression. A fluorescent microscopic examination demonstrated an increased green fluorescent protein signal in Ad-shMDR1- (recombinant adenovirus that coexpressed MDR1 small hairpin ribonucleic acid [shRNA] and green fluorescent protein) infected HCT-15/Rluc cells in a virus dose-dependent manner. Concurrently, with an increasing administered virus dose (0, 15, 30, 60, and 120 multiplicity of infection), Rluc activity was significantly increased in Ad-shMDR1-infected HCT-15/Rluc cells in a virus dose-dependent manner. In vivo bioluminescent imaging showed about 7.5-fold higher signal intensity in Ad-shMDR1-infected tumors than in control tumors (p < .05). Immunohistologic analysis demonstrated marked reduction of P-glycoprotein expression in infected tumor but not in control tumor. In conclusion, the reversal of MDR1 gene expression by MDR1 shRNA was successfully evaluated by bioluminescence imaging with Rluc activity using an in vivo animal model with a multidrug resistance cancer xenograft.

Metallothionein-1 and nitric oxide expression are inversely correlated in a murine model of Chagas disease

PubMed Central

Gonzalez-Mejia, Martha Elba; Torres-Rasgado, Enrique; Porchia, Leonardo M; Salgado, Hilda Rosas; Totolhua, José-Luis; Ortega, Arturo; Hernández-Kelly, Luisa Clara Regina; Ruiz-Vivanco, Guadalupe; Báez-Duarte, Blanca G; Pérez-Fuentes, Ricardo

2014-01-01

Chagas disease, caused by Trypanosoma cruzi, represents an endemic among Latin America countries. The participation of free radicals, especially nitric oxide (NO), has been demonstrated in the pathophysiology of seropositive individuals with T. cruzi. In Chagas disease, increased NO contributes to the development of cardiomyopathy and megacolon. Metallothioneins (MTs) are efficient free radicals scavengers of NO in vitro and in vivo. Here, we developed a murine model of the chronic phase of Chagas disease using endemic T. cruzi RyCH1 in BALB/c mice, which were divided into four groups: infected non-treated (Inf), infected N-monomethyl-L-arginine treated (Inf L-NAME), non-infected L-NAME treated and non-infected vehicle-treated. We determined blood parasitaemia and NO levels, the extent of parasite nests in tissues and liver MT-I expression levels. It was observed that NO levels were increasing in Inf mice in a time-dependent manner. Inf L-NAME mice had fewer T. cruzi nests in cardiac and skeletal muscle with decreased blood NO levels at day 135 post infection. This affect was negatively correlated with an increase of MT-I expression (r = -0.8462, p < 0.0001). In conclusion, we determined that in Chagas disease, an unknown inhibitory mechanism reduces MT-I expression, allowing augmented NO levels. PMID:24676665

The Skeletal Site-Specific Role of Connective Tissue Growth Factor in Prenatal Osteogenesis

PubMed Central

Lambi, Alex G.; Pankratz, Talia L.; Mundy, Christina; Gannon, Maureen; Barbe, Mary F.; Richtsmeier, Joan T.; Popoff, Steven N.

2013-01-01

Background Connective tissue growth factor (CTGF/CCN2) is a matricellular protein that is highly expressed during bone development. Mice with global CTGF ablation (knockout, KO) have multiple skeletal dysmorphisms and perinatal lethality. A quantitative analysis of the bone phenotype has not been conducted. Results We demonstrated skeletal site-specific changes in growth plate organization, bone microarchitecture, and shape and gene expression levels in CTGF KO compared with wild-type mice. Growth plate malformations included reduced proliferation zone and increased hypertrophic zone lengths. Appendicular skeletal sites demonstrated decreased metaphyseal trabecular bone, while having increased mid-diaphyseal bone and osteogenic expression markers. Axial skeletal analysis showed decreased bone in caudal vertebral bodies, mandibles, and parietal bones in CTGF KO mice, with decreased expression of osteogenic markers. Analysis of skull phenotypes demonstrated global and regional differences in CTGF KO skull shape resulting from allometric (size-based) and nonallometric shape changes. Localized differences in skull morphology included increased skull width and decreased skull length. Dysregulation of the transforming growth factor-β-CTGF axis coupled with unique morphologic traits provides a potential mechanistic explanation for the skull phenotype. Conclusions We present novel data on a skeletal phenotype in CTGF KO mice, in which ablation of CTGF causes site-specific aberrations in bone formation. PMID:23073844

Regulation of 5alpha-reductase isoforms by oxytocin in the rat ventral prostate.

PubMed

Assinder, S J; Johnson, C; King, K; Nicholson, H D

2004-12-01

Oxytocin (OT) is present in the male reproductive tract, where it is known to modulate contractility, cell growth, and steroidogenesis. Little is known about how OT regulates these processes. This study describes the localization of OT receptor in the rat ventral prostate and investigates if OT regulates gene expression and/or activity of 5alpha-reductase isoforms I and II. The ventral prostates of adult male Wistar rats were collected following daily sc administration of saline (control), OT, a specific OT antagonist or both OT plus antagonist for 3 d. Expression of the OT receptor was identified in the ventral prostate by RT-PCR and Western blot, and confirmed to be a single active binding site by radioreceptor assay. Immunohistochemistry localized the receptor to the epithelium of prostatic acini and to the stromal tissue. Real-time RT-PCR determined that OT treatment significantly reduced expression of 5alpha-reductase I but significantly increased 5alpha-reductase II expression in the ventral prostate. Activity of both isoforms of 5alpha-reductase was significantly increased by OT, resulting in increased concentration of prostatic dihydrotestosterone. In conclusion, OT is involved in regulating conversion of testosterone to the biologically active dihydrotestosterone in the rat ventral prostate. It does so by differential regulation of 5alpha-reductase isoforms I and II.

Progesterone inhibits contraction and increases TREK-1 potassium channel expression in late pregnant rat uterus

PubMed Central

Yin, Zongzhi; Li, Yun; He, Wenzhu; Li, Dan; Li, Hongyan; Yang, Yuanyuan; Shen, Bing; Wang, Xi; Cao, Yunxia; Khalil, Raouf A.

2018-01-01

Objective The aim of this study was to investigate the effect and mechanism by which progesterone regulates uterine contraction in late pregnant rats Results Progesterone caused concentration-dependent relaxation of uterine strips that was enhanced compared with control nontreated uterine strips. Uterine strips incubated with progesterone showed a significant increase in TREK-1 mRNA expression and protein level. TREK-1 inhibitor L-methionine partly reversed uterine relaxation caused by the progesterone, while TREK-1 activator arachidonic acid did not cause significant change in progesterone-induced relaxation. Conclusions Progesterone inhibits uterine contraction and induces uterine relaxation in late pregnancy. The progesterone-induced inhibition of uterine contraction appears to partly involve increased potassium channel TREK-1 expression/activity. Materials and Methods Uterus from late-pregnant rats (gestational day 19) was isolated, and uterine strips were prepared for isometric contraction measurement. Oxytocin-induced contraction was compared in uterine strips pretreated with different concentration of progesterone. TREK-1 potassium channel inhibitor L-methionine and TREK-1 agonist arachidonic acid were used to determine whether the changes caused by progesterone involve changes in TREK-1 activity. The mRNA and protein expression of TREK-1 in uterine tissues were measured using qPCR and Western blot. PMID:29416642

Prognostic value of estrogen receptor and progesterone receptor tumor expression in Danish ovarian cancer patients: from the 'MALOVA' ovarian cancer study.

PubMed

Høgdall, Estrid V S; Christensen, Lise; Høgdall, Claus K; Blaakaer, Jan; Gayther, Simon; Jacobs, Ian J; Christensen, Ib Jarle; Kjaer, Susanne K

2007-11-01

Estrogen and progesterone are important hormones secreted by the ovary acting through specific receptors. Tumor tissue expression profiles of these have demonstrated prognostic value in malignancies such as breast, uterine and prostate cancer. In this study, including tissue samples from 773 Danish patients with an ovarian tumor, we evaluated whether estrogen receptor (ER) and progesterone receptor (PR) expression correlated with clinico-pathological parameters, and a possible prognostic impact on ovarian cancer (OC) patients was investigated. Using tissue array and immunohistochemistry, we analyzed the ER and PR expression levels in tissues from 582 women with OC and 191 women with low malignancy potential (LMP) ovarian tumors. Our results demonstrated that ER was expressed in 30 of the 191 LMP tumors (16%) and in 207 of the 582 OC (36%). PR was expressed in 38 LMP tumors (20%) and in 115 OC (20%). For both tumor types an excess of positive tumors was found in the serous compared to the mucinous subtype (p< or =0.00001). The frequency of ER expression-positive OC increased with increasing FIGO stage (p=0.0003), and the frequency of PR-positive tumors increased with increasing histological grade (p=0.0006). In a Cox survival analysis, a tissue ER and PR expression 10% or higher was found to imply an independent significant advantageous course of patient disease-specific survival (ER: hazard ratio (HR), 0.80; 95% confidence interval (CI), 0.63-0.99; PR: HR, 0.69; 95% CI, 0.51-0.94) together with FIGO stage, residual tumor after primary surgery, age at diagnosis and other histological types vs. serous adenocarcinoma. The histological grade of tumor was found to have no independent prognostic value. The prognostic value of ER and PR was found additive with a HR for patients with high ER and PR expression of 0.48 (95% CI, 0.31-0.74) compared to patients with <10% expression for both receptors. In conclusion, our results predict that an elevated expression of ER and PR, alone and in combination, point to a favorable outcome for patients with OC.

17β-estradiol-induced growth of triple-negative breast cancer cells is prevented by the reduction of GPER expression after treatment with gefitinib.

PubMed

Girgert, Rainer; Emons, Günter; Gründker, Carsten

2017-02-01

Triple-negative breast cancers (TNBCs) are neither susceptible to endocrine therapy due to a lack of estrogen receptor α expression nor trastuzumab. TNBCs frequently overexpress epidermal growth factor receptor (EGFR) and membrane bound estrogen receptor, GPER. To a certain extent the growth of TNBCs is stimulated by 17β-estradiol via GPER. We analyzed whether inhibition of EGFR by gefitinib reduces the expression of GPER and subsequent signal transduction in TNBC cells. Dependence of proliferation on 17β-estradiol was determined using Alamar Blue assay. Expression of GPR30 and activation of c-src, EGFR and cAMP-responsive element binding (CREB) protein by 17β-estradiol was analyzed by western blotting. Expression of c-fos, cyclin D1 and aromatase was determined using RT-PCR. Gefitinib reduced GPER expression concentration‑ and time‑dependently. In HCC70 cells, GPER expression was reduced to 15±11% (p<0.05) after treatment with 200 nM gefitinib for four days, and in HCC1806 cells GPER expression was reduced to 39±5% (p<0.01) of the control. 17β-estradiol significantly increased the percentage of HCC1806 cells within 7 days to 145±29% of the control (HCC70, 110±8%). This increase in cell growth was completely prevented in both TNBC cell lines after GPR30 expression was downregulated by treatment with 200 nM gefitinib. In HCC1806 cells, activation of c-src was increased by 17β-estradiol to 350±50% (p<0.01), and gefitinib reduced src activation to 110%. Similar results were obtained in the HCC70 cells. Phosphorylation of EGFR increased to 240±40% (p<0.05) in the HCC1806 cells treated with 17β-estradiol (HCC70, 147±25%). Gefitinib completely prevented this activation. Phosphorylation of CREB and induction of c-fos, cyclin D1 and aromatase expression by 17β-estradiol were all prevented by gefitinib. These experiments conclusively show that reduction of GPER expression is a promising therapeutic approach for TNBC.

Soy isoflavones interfere with thyroid hormone homeostasis in orchidectomized middle-aged rats

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

Šošić-Jurjević, Branka, E-mail: brankasj@ibiss.bg.ac.rs; Filipović, Branko; Wirth, Eva Katrin

We previously reported that genistein (G) and daidzein (D) administered subcutaneously (10 mg/kg) induce changes in the angio-follicular units of the thyroid gland, reduce concentration of total thyroid hormones (TH) and increase thyrotropin (TSH) in serum of orchidectomized middle-aged (16-month-old) rats. To further investigate these effects, we now examined expression levels of the thyroglobulin (Tg), thyroperoxidase (Tpo), vascular endothelial growth factor A (Vegfa) and deiodinase type 1 (Dio 1) genes in the thyroid; in the pituitary, genes involved in TH feedback control (Tsh β, Dio 1, Dio 2, Trh receptor); and in the liver and kidney, expression of T{sub 3}-activatedmore » genes Dio 1 and Spot 14, as well as transthyretin (Ttr), by quantitative real-time PCR. We also analyzed TPO-immunopositivity and immunofluorescence of T{sub 4} bound to Tg, determined thyroid T{sub 4} levels and measured deiodinase enzyme activities in examined organs. Decreased expression of Tg and Tpo genes (p < 0.05) correlated with immunohistochemical staining results, and together with decreased serum total T{sub 4} levels, indicates decreased Tg and TH synthesis following treatments with both isoflavones. However, expression of Spot 14 (p < 0.05) gene in liver and kidney was up-regulated, and liver Dio 1 expression and activity (p < 0.05) increased. At the level of pituitary, no significant change in gene expression levels, or Dio 1 and 2 enzyme activities was observed. In conclusion, both G and D impaired Tg and TH synthesis, but at the same time increased tissue availability of TH in peripheral tissues of Orx middle-aged rats. - Highlights: • We tested how genistein and daidzein interfere with thyroid hormone homeostasis. • Thyroid: decreased expression of Tg and TPO genes correlated with IHC results. • Serum: total T{sub 4} reduced and TSH increased. • Liver and kidney: expression of Spot 14 and liver Dio 1 activity increased. • Pituitary: expression of T{sub 3}-regulated genes and Dio 1 and 2 activities unchanged.« less

Cpt1a gene expression in peripheral blood mononuclear cells as an early biomarker of diet-related metabolic alterations

PubMed Central

Díaz-Rúa, Rubén; Palou, Andreu; Oliver, Paula

2016-01-01

Background Research on biomarkers that provide early information about the development of future metabolic alterations is an emerging discipline. Gene expression analysis in peripheral blood mononuclear cells (PBMC) is a promising tool to identify subjects at risk of developing diet-related diseases. Objective We analysed PBMC expression of key energy homeostasis-related genes in a time-course analysis in order to find out early markers of metabolic alterations due to sustained intake of high-fat (HF) and high-protein (HP) diets. Design We administered HF and HP diets (4 months) to adult Wistar rats in isocaloric conditions to a control diet, mainly to avoid overweight associated with the intake of hyperlipidic diets and, thus, to be able to characterise markers of metabolically obese normal-weight (MONW) syndrome. PBMC samples were collected at different time points of dietary treatment and expression of relevant energy homeostatic genes analysed by real-time reverse transcription-polymerase chain reaction. Serum parameters related with metabolic syndrome, as well as fat deposition in liver, were also analysed. Results The most outstanding results were those obtained for the expression of the lipolytic gene carnitine palmitoyltransferase 1a (Cpt1a). Cpt1a expression in PBMC increased after only 1 month of exposure to both unbalanced diets, and this increased expression was maintained thereafter. Interestingly, in the case of the HF diet, Cpt1a expression was altered even in the absence of increased body weight but correlated with alterations such as higher insulin resistance, alteration of serum lipid profile and, particularly, increased fat deposition in liver, a feature characteristic of metabolic syndrome, which was even observed in animals fed with HP diet. Conclusions We propose Cpt1a gene expression analysis in PBMC as an early biomarker of metabolic alterations associated with MONW phenotype due to the intake of isocaloric HF diets, as well as a marker of increased risk of metabolic diseases associated with the intake of HF or HP diets. PMID:27885970

Bile acid-FXRα pathways regulate male sexual maturation in mice

PubMed Central

Vega, Aurélie; Sédes, Lauriane; Rouaisnel, Betty; de Haze, Angélique; Baron, Silvère; Schoonjans, Kristina; Caira, Françoise; Volle, David H.

2016-01-01

The bile acid receptor Farnesol-X-Receptor alpha (FRXα) is a member of the nuclear receptor superfamily. FRXα is expressed in the interstitial compartment of the adult testes, which contain the Leydig cells. In adult, short term treatment (12 hours) with FRXα agonist inhibits the expression of steroidogenic genes via the induction of the Small heterodimer partner (SHP). However the consequences of FRXα activation on testicular pathophysiology have never been evaluated. We demonstrate here that mice fed a diet supplemented with bile acid during pubertal age show increased incidence of infertility. This is associated with altered differentiation and increase apoptosis of germ cells due to lower testosterone levels. At the molecular level, next to the repression of basal steroidogenesis via the induction expression of Shp and Dax-1, two repressors of steroidogenesis, the main action of the BA-FRXα signaling is through lowering the Leydig cell sensitivity to the hypothalamo-pituitary axis, the main regulator of testicular endocrine function. In conclusion, BA-FRXα signaling is a critical actor during sexual maturation. PMID:26848619

Transcriptional Changes That Characterize the Immune Reactions of Leprosy

PubMed Central

Dupnik, Kathryn M.; Bair, Thomas B.; Maia, Andressa O.; Amorim, Francianne M.; Costa, Marcos R.; Keesen, Tatjana S. L.; Valverde, Joanna G.; Queiroz, Maria do Carmo A. P.; Medeiros, Lúcio L.; de Lucena, Nelly L.; Wilson, Mary E.; Nobre, Mauricio L.; Johnson, Warren D.; Jeronimo, Selma M. B.

2015-01-01

Background. Leprosy morbidity is increased by 2 pathologic immune reactions, reversal reaction (RR) and erythema nodosum leprosum (ENL). Methods. To discover host factors related to immune reactions, global transcriptional profiles of peripheral blood mononuclear cells were compared between 11 RR, 11 ENL, and 19 matched control patients, with confirmation by quantitative polymerase chain reaction. Encoded proteins were investigated in skin biopsy specimens by means of immunohistochemistry. Results. There were 275 genes differentially expressed in RR and 517 differentially expressed in ENL on the microarray. Pathway analysis showed immunity-related pathways represented in RR and ENL transcriptional profiles, with the “complement and coagulation” pathway common to both. Interferon γ was identified as a significant upstream regulator of the expression changes for RR and ENL. Immunohistochemical staining of skin lesions showed increased C1q in both RR and ENL. Conclusions. These data suggest a previously underrecognized role for complement in the pathogenesis of both RR and ENL, and we propose new hypotheses for reaction pathogenesis. PMID:25398459

Carbamylated erythropoietin protects the kidneys from ischemia-reperfusion injury without stimulating erythropoiesis

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

Imamura, Ryoichi; Isaka, Yoshitaka; Ichimaru, Naotsugu

Several studies have shown that erythropoietin (EPO) can protect the kidneys from ischemia-reperfusion injury and can raise the hemoglobin (Hb) concentration. Recently, the EPO molecule modified by carbamylation (CEPO) has been identified and was demonstrated to be able to protect several organs without increasing the Hb concentration. We hypothesized that treatment with CEPO would protect the kidneys from tubular apoptosis and inhibit subsequent tubulointerstitial injury without erythropoiesis. The therapeutic effect of CEPO was evaluated using a rat ischemia-reperfusion injury model. Saline-treated kidneys exhibited increased tubular apoptosis with interstitial expression of {alpha}-smooth muscle actin ({alpha}-SMA), while EPO treatment inhibited tubular apoptosismore » and {alpha}-SMA expression to some extent. On the other hand, CEPO-treated kidneys showed minimal tubular apoptosis with limited expression of {alpha}-SMA. Moreover, CEPO significantly promoted tubular epithelial cell proliferation without erythropoiesis. In conclusion, we identified a new therapeutic approach using CEPO to protect kidneys from ischemia-reperfusion injury.« less

Nandrolone decanoate is able to modulate proliferation and adhesion of myoblasts.

PubMed

Oliveira, E N; Fernandes, K P; Silva, C A; Oliveira, T S; Junior, J A; Bussadori, S K; Renno, A C; Mesquita-Ferrari, R A

2014-07-01

The search for a more efficient repair process of muscle injuries has become evident in clinical practice. The aim of the present study was to evaluate the effect of nandrolone decanoate (ND) on the proliferation, adhesion, and expression of myogenic regulatory factors (MRFs) in C2C12 cells.Methods. Cell proliferation and adhesion were assessed using an MTT assay. The expression of MRFs was assessed by real-time PCR.Results. ND applied at 10 or 25 µM concentration induced after 60 min an increase in adhesion, at 5 µM concentration induced after 5 days an increase in cell proliferation, and ND at 50 µM concentration led after 5 days to a decrease in cell proliferation in comparison with other groups. The steroid did not alter the expression of MRFs.Conclusions. The positive effects of ND regarding the proliferation and adhesion of C2C12 cells suggest that this steroid may have positive effects following a muscle injury.

Distinct effects of calorie restriction on adipose tissue cytokine and angiogenesis profiles in obese and lean mice

PubMed Central

2012-01-01

Background Obesity associates with low-grade inflammation and adipose tissue remodeling. Using sensitive high-throughput protein arrays we here investigated adipose tissue cytokine and angiogenesis-related protein profiles from obese and lean mice, and in particular, the influence of calorie restriction (CR). Methods Tissue samples from visceral fat were harvested from obese mice fed with a high-fat diet (60% of energy), lean controls receiving low-fat control diet as well as from obese and lean mice kept under CR (energy intake 70% of ad libitum intake) for 50 days. Protein profiles were analyzed using mouse cytokine and angiogenesis protein array kits. Results In obese and lean mice, CR was associated with 11.3% and 15.6% reductions in body weight, as well as with 4.0% and 4.6% reductions in body fat percentage, respectively. Obesity induced adipose tissue cytokine expressions, the most highly upregulated cytokines being IL-1ra, IL-2, IL-16, MCP-1, MIG, RANTES, C5a, sICAM-1 and TIMP-1. CR increased sICAM-1 and TIMP-1 expression both in obese and lean mice. Overall, CR showed distinct effects on cytokine expressions; in obese mice CR largely decreased but in lean mice increased adipose tissue cytokine expressions. Obesity was also associated with increased expressions of angiogenesis-related proteins, in particular, angiogenin, endoglin, endostatin, endothelin-1, IGFBP-3, leptin, MMP-3, PAI-1, TIMP-4, CXCL16, platelet factor 4, DPPIV and coagulation factor III. CR increased endoglin, endostatin and platelet factor 4 expressions, and decreased IGFBP-3, NOV, MMP-9, CXCL16 and osteopontin expressions both in obese and lean mice. Interestingly, in obese mice, CR decreased leptin and TIMP-4 expressions, whereas in lean mice their expressions were increased. CR decreased MMP-3 and PAI-1 only in obese mice, whereas CR decreased FGF acidic, FGF basic and coagulation factor III, and increased angiogenin and DPPIV expression only in lean mice. Conclusions CR exerts distinct effects on adipocyte cytokine and angiogenesis profiles in obese and lean mice. Our study also underscores the importance of angiogenesis-related proteins and cytokines in adipose tissue remodeling and development of obesity. PMID:22748184

Expression of multidrug resistance proteins in retinoblastoma

PubMed Central

Shukla, Swati; Srivastava, Arpna; Kumar, Sunil; Singh, Usha; Goswami, Sandeep; Chawla, Bhavna; Bajaj, Mandeep Singh; Kashyap, Seema; Kaur, Jasbir

2017-01-01

AIM To elucidate the mechanism of multidrug resistance in retinoblastoma, and to acquire more insights into in vivo drug resistance. METHODS Three anticancer drug resistant Y79 human RB cells were generated against vincristine, etoposide or carboplatin, which are used for conventional chemotherapy in RB. Primary cultures from enucleated eyes after chemotherapy (PCNC) were also prepared. Their chemosensitivity to chemotherapeutic agents (vincristine, etoposide and carboplatin) were measured using MTT assay. Western blot analysis was performed to evaluate the expression of p53, Bcl-2 and various multidrug resistant proteins in retinoblastoma cells. RESULTS Following exposure to chemotherapeutic drugs, PCNC showed less sensitivity to drugs. No significant changes observed in the p53 expression, whereas Bcl-2 expression was found to be increased in the drug resistant cells as well as in PCNC. Increased expression of P-glycoprotein (P-gp) was observed in drug resistant Y79 cells; however there was no significant change in the expression of P-gp found between primary cultures of primarily enucleated eyes and PCNC. Multidrug resistance protein 1 (Mrp-1) expression was found to be elevated in the drug resistant Y79 cells as well as in PCNC. No significant change in the expression of lung resistance associated protein (Lrp) was observed in the drug resistant Y79 cells as well as in PCNC. CONCLUSION Our results suggest that multidrug resistant proteins are intrinsically present in retinoblastoma which causes treatment failure in managing retinoblastoma with chemotherapy. PMID:29181307

[Adipose-derived stem cell transplantation promotes the expression of netrin-1 in the rat cortex after focal cerebral ischemia].

PubMed

Wang, Jiehua; Hong, Zhuquan; Pan, Ying; Li, Guoqian

2017-01-01

Objective To observe the effect of adipose-derived stem cells (ADSCs) transplantation on the expression of netrin-1 in rats after focal cerebral ischemia. Methods Male SD rats were randomly divided into control group, model group and ADSC group. ADSCs were harvested and purified. Focal cerebral ischemia models were established in rats by the suture method. ADSCs were injected into the lateral ventricle of ADSC group rats and the same does of PBS was given to model group rats. At day 4, 7 and 14 after reperfusion, six rats were sacrificed to remove the brain tissues at each time point. The expression of netrin-1 was detected by reverse-transcription PCR, Western blotting and immunohistochemistry. Results Compared with the control group, the expression of netrin-1 in the brain tissues of the model group increased after focal cerebral ischemia, reached the peak at 4 days, and the expression of netrin-1 was significantly higher than that of the control group at each time point. Compared with the model group, the expression of netrin-1 in the ADSC group increased further, reached the peak at 7 days, and the expression of netrin-1 in the ADSC group was significantly higher than that of the model group at each time point. Conclusion ADSC transplantation could up-regulate the expression of netrin-1, and promote axon regeneration and the recovery of neurological functions.

Autoantigens targeted in scleroderma patients with vascular disease are enriched in endothelial lineage cells

PubMed Central

McMahan, Zsuzsanna H.; Cottrell, Tricia R.; Wigley, Fredrick M.; Antiochos, Brendan; Zambidis, Elias T.; Park, Tea Soon; Halushka, Marc K.; Gutierrez-Alamillo, Laura; Cimbro, Raffaello; Rosen, Antony; Casciola-Rosen, Livia

2016-01-01

Objective Scleroderma patients with autoantibodies to centromere proteins (CENPs) and/or interferon-inducible protein 16 (IFI16) are at increased risk of severe vascular complications. We set out to define whether these autoantigens are enriched in cells of the vasculature. Methods Successive stages of embryoid bodies (EBs) as well as vascular progenitors were used to evaluate the expression of scleroderma autoantigens IFI16 and CENP by immunoblotting. CD31 was included to mark early blood vessels. IFI16 and CD31 expression were defined in skin paraffin sections from scleroderma patients and from healthy controls. IFI16 expression was determined by flow cytometry in circulating endothelial cells (CECs) and circulating progenitor cells (CPCs). Results Expression of CENP-A, IFI16 and CD31 was enriched in EBs at days 10 and 12 of differentiation, and particularly in cultures enriched in vascular progenitors (IFI16, CD31, CENPs A and-B). This pattern was distinct from that of comparator autoantigens. Immunohistochemical staining of skin paraffin sections showed enrichment of IFI16 in CD31-positive vascular endothelial cells in biopsies from scleroderma patients and normal controls. Flow cytometry analysis revealed IFI16 expression in CPCs, but minimal expression in CECs. Conclusion Expression of scleroderma autoantigens IFI16 and CENPs, which are associated with severe vascular disease, is increased in vascular progenitors and mature endothelial cells. High level, lineage-enriched expression of autoantigens may explain the striking association between clinical phenotypes and the immune targeting of specific autoantigens. PMID:27159521

Mechanical versus humoral determinants of brain death-induced lung injury

PubMed Central

Dewachter, Laurence; Rorive, Sandrine; Remmelink, Myriam; Weynand, Birgit; Melot, Christian; Hupkens, Emeline; Dewachter, Céline; Creteur, Jacques; Mc Entee, Kathleen; Naeije, Robert; Rondelet, Benoît

2017-01-01

Background The mechanisms of brain death (BD)-induced lung injury remain incompletely understood, as uncertainties persist about time-course and relative importance of mechanical and humoral perturbations. Methods Brain death was induced by slow intracranial blood infusion in anesthetized pigs after randomization to placebo (n = 11) or to methylprednisolone (n = 8) to inhibit the expression of pro-inflammatory mediators. Pulmonary artery pressure (PAP), wedged PAP (PAWP), pulmonary vascular resistance (PVR) and effective pulmonary capillary pressure (PCP) were measured 1 and 5 hours after Cushing reflex. Lung tissue was sampled to determine gene expressions of cytokines and oxidative stress molecules, and pathologically score lung injury. Results Intracranial hypertension caused a transient increase in blood pressure followed, after brain death was diagnosed, by persistent increases in PAP, PCP and the venous component of PVR, while PAWP did not change. Arterial PO2/fraction of inspired O2 (PaO2/FiO2) decreased. Brain death was associated with an accumulation of neutrophils and an increased apoptotic rate in lung tissue together with increased pro-inflammatory interleukin (IL)-6/IL-10 ratio and increased heme oxygenase(HO)-1 and hypoxia inducible factor(HIF)-1 alpha expression. Blood expressions of IL-6 and IL-1β were also increased. Methylprednisolone pre-treatment was associated with a blunting of increased PCP and PVR venous component, which returned to baseline 5 hours after BD, and partially corrected lung tissue biological perturbations. PaO2/FiO2 was inversely correlated to PCP and lung injury score. Conclusions Brain death-induced lung injury may be best explained by an initial excessive increase in pulmonary capillary pressure with increased pulmonary venous resistance, and was associated with lung activation of inflammatory apoptotic processes which were partially prevented by methylprednisolone. PMID:28753621

Effects of Dietary Eicosapentaenoic Acid (EPA) Supplementation in High-Fat Fed Mice on Lipid Metabolism and Apelin/APJ System in Skeletal Muscle

PubMed Central

Wanecq, Estelle; Rancoule, Chloé; Batut, Aurélie; Deleruyelle, Simon; Lionetti, Lillà; Valet, Philippe; Castan-Laurell, Isabelle

2013-01-01

Various studies have shown that eicosapentaenoic acid (EPA) has beneficial effects on obesity and associated disorders. Apelin, the ligand of APJ receptor also exerts insulin-sensitizing effects especially by improving muscle metabolism. EPA has been shown to increase apelin production in adipose tissue but its effects in muscle have not been addressed. Thus, the effects of EPA supplementation (36 g/kg EPA) in high-fat diet (HFD) (45% fat, 20% protein, 35% carbohydrate) were studied in mice with focus on muscle lipid metabolism and apelin/APJ expression. Compared with HFD mice, HFD+EPA mice had significantly less weight gain, fat mass, lower blood glucose, insulinemia and hepatic steatosis after 10 weeks of diet. In addition, EPA prevented muscle metabolism alterations since intramuscular triglycerides were decreased and β-oxidation increased. In soleus muscles of HFD+EPA mice, apelin and APJ expression were significantly increased compared to HFD mice. However, plasma apelin concentrations in HFD and HFD+EPA mice were similar. EPA-induced apelin expression was confirmed in differentiated C2C12 myocytes but in this model, apelin secretion was also increased in response to EPA treatment. In conclusion, EPA supplementation in HFD prevents obesity and metabolic alterations in mice, especially in skeletal muscle. Since EPA increases apelin/APJ expression in muscle, apelin may act in a paracrine/autocrine manner to contribute to these benefical effects. PMID:24244380

Increased hepatic mitochondrial FA oxidation reduces plasma and liver TG levels and is associated with regulation of UCPs and APOC-III in rats

PubMed Central

Lindquist, Carine; Bjørndal, Bodil; Rossmann, Christine Renate; Tusubira, Deusdedit; Svardal, Asbjørn; Røsland, Gro Vatne; Tronstad, Karl Johan; Hallström, Seth; Berge, Rolf Kristian

2017-01-01

Hepatic mitochondrial function, APOC-III, and LPL are potential targets for triglyceride (TG)-lowering drugs. After 3 weeks of dietary treatment with the compound 2-(tridec-12-yn-1-ylthio)acetic acid (1-triple TTA), the hepatic mitochondrial FA oxidation increased more than 5-fold in male Wistar rats. Gene expression analysis in liver showed significant downregulation of APOC-III and upregulation of LPL and the VLDL receptor. This led to lower hepatic (53%) and plasma (73%) TG levels. Concomitantly, liver-specific biomarkers related to mitochondrial biogenesis and function (mitochondrial DNA, citrate synthase activity, and cytochrome c and TFAM gene expression) were elevated. Interestingly, 1-triple TTA lowered plasma acetylcarnitine levels, whereas the concentration of β-hydroxybutyrate was increased. The hepatic energy state was reduced in 1-triple TTA-treated rats, as reflected by increased AMP/ATP and decreased ATP/ADP ratios, whereas the energy state remained unchanged in muscle and heart. The 1-triple TTA administration induced gene expression of uncoupling protein (UCP)2 and UCP3 in liver. In conclusion, the 1-triple TTA-mediated clearance of blood TG may result from lowered APOC-III production, increased hepatic LPL gene expression, mitochondrial FA oxidation, and (re)uptake of VLDL facilitating drainage of FAs to the liver for β-oxidation and production of ketone bodies as extrahepatic fuel. The possibility that UCP2 and UCP3 mediate a moderate degree of mitochondrial uncoupling should be considered. PMID:28473603

Effect of Pioglitazone on Cardio-Metabolic Risk in Patients with Obstructive Sleep Apnea

PubMed Central

Liu, Alice; Abbasi, Fahim; Kim, Sun H.; Ariel, Danit; Lamendola, Cindy; Cardell, James; Xu, Shiming; Patel, Shailja; Tomasso, Vanessa; Mojaddidi, Hafasa; Grove, Kaylene; Tsao, Philip S.; Kushida, Clete A.; Reaven, Gerald M.

2017-01-01

Prevalence of insulin resistance is increased in patients with obstructive sleep apnea (OSA). Since insulin resistance is an independent predictor of cardiovascular disease (CVD), this study was initiated to see if pioglitazone administration would improve insulin sensitivity, and thereby decrease risk of CVD in overweight/obese, nondiabetic, insulin-resistant patients with untreated OSA. Patients (n=30) were administered pioglitazone (45 mg/day) for 8 weeks, and measurements were made before and after intervention of insulin action (insulin-mediated glucose uptake by the insulin suppression test), C-reactive protein, lipid/lipoprotein profile, and gene expression profile of peri-umbilical subcutaneous fat tissue. Insulin sensitivity increased 31% (p<0.001) among pioglitazone-treated individuals, associated with a decrease in C-reactive protein concentration (p≤ 0.001), a decrease in plasma triglyceride and increase in high-density lipoprotein cholesterol concentrations (p≤ 0.001), accompanied by significant changes in apolipoprotein A1 and B concentrations and lipoprotein subclasses known to decrease CVD risk. In addition, subcutaneous adipose tissue gene expression profile showed a 1.6-fold (p<0.01) increase in GLUT4 expression, as well as decreased expression in 5 of 9 inflammatory genes (p<0.05). In conclusion, enhanced insulin sensitivity can significantly decrease multiple cardio-metabolic risk factors in patients with untreated OSA, consistent with the view that coexisting insulin resistance plays an important role in the association between OSA and increased risk of CVD. PMID:28219664

Overexpression of xeroderma pigmentosum group C decreases the chemotherapeutic sensitivity of colorectal carcinoma cells to cisplatin

PubMed Central

Zhang, Yi; Cao, Jia; Meng, Yanni; Qu, Chunying; Shen, Feng; Xu, Leiming

2018-01-01

Xeroderma pigmentosum group C (XPC) is a DNA-damage-recognition gene active at the early stage of DNA repair. XPC also participates in regulation of cell-cycle checkpoint and DNA-damage-induced apoptosis. In the present study, the expression levels of genes involved in nucleotide excision repair (NER) were assessed in human colorectal cancer (CRC) tissue. This analysis revealed that expression of XPC mRNA significantly increased in colorectal carcinoma tissues compared with matched normal controls. Expression of XPC gradually increased along with the degree of progression of CRC. In vitro, an XTT assay demonstrated that small interfering RNA (siRNA) targeting XPC significantly increased the sensitivity of CRC SW480 cells to cisplatin, whereas cells transfected with a XPC-overexpression plasmid became more resistant to cisplatin. Furthermore, flow cytometry revealed that the proportion of apoptotic cells significantly increased in XPC-knockdown cells upon cisplatin treatment. However, the overexpression XPC significantly increased the resistance of cells to cisplatin. In vivo, tumor growth was significantly reduced in tumor-bearing mice when the XPC gene was knocked down. Upregulation of the expression of pro-apoptotic Bcl-associated X and downregulation of the anti-apoptotic B-cell lymphoma 2 proteins was observed in the implanted tumor tissue. In conclusion, XPC serves a key role in chemotherapeutic sensitivity of CRC to cisplatin, meaning that it may be a potential target for chemotherapy of CRC. PMID:29616110

Effect of biotin on activity and gene expression of biotin-dependent carboxylases in the liver of dairy cows.

PubMed

Ferreira, G; Weiss, W P

2007-03-01

Biotin is a cofactor of the gluconeogenic enzymes pyruvate carboxylase (PC) and propionyl-coenzyme A carboxylase (PCC). We hypothesized that biotin supplementation increases the activity and gene expression of PC and PCC and the gene expression of phosphoenol-pyruvate carboxykinase (PEPCK) in the liver of lactating dairy cows. Eight multiparous Holstein cows (40 +/- 2 kg/d of milk yield and 162 +/- 35 d in milk) were randomly assigned to 1 of 2 diet sequences in a crossover design with two 22-d periods. Treatments consisted of a basal diet (60% concentrate) containing 0 or 0.96 mg/kg of supplemental biotin. On d 21 of each period, liver tissue was collected by percutaneous liver biopsy. Activities of PC and PCC were determined by measuring the fixation of [14C]O2 in liver homogenates. Abundance of mRNA for PCC, PC, and PEPCK was determined by quantitative reverse-transcription PCR. Biotin supplementation did not affect milk production or composition. Biotin supplementation increased the activity of PC but had no effect on PCC activity. Biotin supplementation did not affect the gene expression of PC, PCC, and PEPCK. The increased activity of PC without changes in mRNA abundance may have been caused by increased activation of the apoenzymes by holocarboxylase synthetase. In conclusion, biotin supplementation affected the activity of PC in the liver of lactating dairy cows, but whether biotin supplementation increases glucose production in the liver remains to be determined.

Dexamethasone upregulates ANP C-receptor protein in human mesangial cells without affecting mRNA.

PubMed

Ardaillou, N; Blaise, V; Placier, S; Amestoy, F; Ardaillou, R

1996-03-01

The objective of this study was to examine the role of dexamethasone on the expression of natriuretic peptide B-type and C-type receptors (ANPR-B and ANPR-C) in cultured human mesangial cells, which only possess these two subtypes. Dexamethasone caused concentration- and time-dependent increases in 125I-labeled ANP binding, which were prevented by glucocorticoid receptor inhibition with RU-38486. A lag time of 24 h and a concentration of dexamethasone of at least 1 nmol/l were necessary for this effect to occur. Dexamethasone-induced upregulation of 125I-ANP binding resulted from increased receptor density. No change in dissociation constant (Kd) was observed. Only ANPR-C were affected by dexamethasone. Indeed, dexamethasone did not modify C-type natriuretic peptide (i.e., CNP)-dependent cGMP production by mesangial cells. Moreover, dexamethasone upregulated ANPR-C protein expression as shown by Western blot analysis and by an increase in ANPR-C immunoreactivity at the cell surface. In contrast, dexamethasone did not modify ANPR-C mRNA expression. In conclusion, glucocorticoids increase ANPR-C density on mesangial cells through a mechanism implying, successively, interaction with the glucocorticoid receptor and increase of ANPR-C protein synthesis at a posttranscriptional stage. Thus dexamethasone may influence availability of natriuretic peptides at their glomerular target sites.

Increased Atherogenesis during Streptococcus mutans Infection in ApoE-null Mice

PubMed Central

Kesavalu, L.; Lucas, A.R.; Verma, R.K.; Liu, L.; Dai, E.; Sampson, E.; Progulske-Fox, A.

2012-01-01

Streptococcus mutans, a dental caries pathogen, also causes endocarditis and is detected in atheroscelerotic plaque. We investigated the potential for an invasive strain of S. mutans, OMZ175, to accelerate plaque growth in apolipoprotein E deficient (ApoEnull) mice without and with balloon angioplasty (BA) injury, a model of restenosis. ApoEnull mice were divided into 4 groups (N = 10), 2 with and 2 without BA. One each of the BA and non-BA groups was infected with S. mutans (Sm). S. mutans DNA, plaque area, inflammatory cell invasion, and Toll-like receptor (TLR) expression were measured at 6-20 weeks post-infection. S. mutans genomic DNA was detected in the aorta, liver, spleen, and heart. Plaque growth was significantly increased in infected mice with BA (Sm+BA) vs. those in the non-infected groups (p < 0.03). Plaque size was increased after infection without BA (Sm), but did not reach significance. Aortic specimens from both S. mutans and Sm+BA groups displayed increased numbers of macrophages, and TLR4 expression was increased in BA mice. In conclusion, S. mutans infection accelerated plaque growth, macrophage invasion, and TLR4 expression after angioplasty. S. mutans may also be associated with atherosclerotic plaque growth in non-injured arteries. PMID:22262633

The Regulation of Vascular Endothelial Growth Factor by Hypoxia and Prostaglandin F2α during Human Endometrial Repair

PubMed Central

Maybin, Jacqueline A.; Hirani, Nikhil; Brown, Pamela; Jabbour, Henry N.

2011-01-01

Context: The human endometrium has an exceptional capacity for repeated repair after menses, but its regulation remains undefined. Premenstrually, progesterone levels fall and prostaglandin (PG) F2α synthesis increases, causing spiral arteriole constriction. We hypothesized that progesterone withdrawal, PGF2α, and hypoxia increase vascular endothelial growth factor (VEGF), an endometrial repair factor. Design and Results: Endometrial biopsies were collected (n = 47) with ethical approval and consent. VEGF mRNA, quantified by quantitative RT-PCR, was increased during menstruation (P < 0.01).VEGF protein was maximally secreted from proliferative endometrial explants. Treatment of an endometrial epithelial cell line and primary human endometrial stromal cells with 100 nm PGF2α or hypoxia (0.5% O2) resulted in significant increases in VEGF mRNA and protein. VEGF was maximal when cells were cotreated with PGF2α and hypoxia simultaneously (P < 0.05–0.001). Secretory-phase endometrial explants also showed an increase in VEGF with cotreatment (P < 0.05). However, proliferative-phase explants showed no increase in VEGF on treatment with PGF2α and/or hypoxia. Proliferative tissue was induced to increase VEGF mRNA expression when exposed to progesterone and its withdrawal in vitro but only in the presence of hypoxia and PG. Hypoxia-inducible factor-1α (HIF-1α) silencing with RNA interference suppressed hypoxia-induced VEGF expression in endometrial cells but did not alter PGF2α-induced VEGF expression. Conclusions: Endometrial VEGF is increased at the time of endometrial repair. Progesterone withdrawal, PGF2α, and hypoxia are necessary for this perimenstrual VEGF expression. Hypoxia acts via HIF-1α to increase VEGF, whereas PGF2α acts in a HIF-1α-independent manner. Hence, two pathways regulate the expression of VEGF during endometrial repair. PMID:21677035

Regulation of aromatase expression in the anterior amygdala of the developing mouse brain depends on ERβ and sex chromosome complement.

PubMed

Cisternas, Carla Daniela; Cabrera Zapata, Lucas Ezequiel; Arevalo, María Angeles; Garcia-Segura, Luis Miguel; Cambiasso, María Julia

2017-07-13

During development sex differences in aromatase expression in limbic regions of mouse brain depend on sex chromosome factors. Genes on the sex chromosomes may affect the hormonal regulation of aromatase expression and this study was undertaken to explore that possibility. Male E15 anterior amygdala neuronal cultures expressed higher levels of aromatase (mRNA and protein) than female cultures. Furthermore, treatment with oestradiol (E2) or dihydrotestosterone (DHT) increased Cyp19a1 expression and aromatase protein levels only in female neuronal cultures. The effect of E2 on aromatase expression was not imitated by oestrogen receptor (ER) α agonist PPT or the GPER agonist G1, but it was fully reproduced by DPN, a specific ligand of ERβ. By contrast, the effect of DHT on aromatase expression was not blocked by the anti-androgen flutamide, but completely abrogated by the ERβ antagonist PHTPP. Experiments using the four core genotype model showed a sex chromosome effect in ERβ expression (XY > XX) and regulation by E2 or DHT (only XX respond) in amygdala neurons. In conclusion, sex chromosome complement governs the hormonal regulation of aromatase expression through activation of ERβ in developing mouse brain.

Over-expression of IQGAP1 indicates poor prognosis in head and neck squamous cell carcinoma.

PubMed

Wu, Cong-Cong; Li, Hao; Xiao, Yao; Yang, Lei-Lei; Chen, Lei; Deng, Wei-Wei; Wu, Lei; Zhang, Wen-Feng; Sun, Zhi-Jun

2018-05-30

IQ-domain GTPase-activating protein 1 (IQGAP1) is associated with the development and progression of many human cancers. We aimed to investigate the expression and clinicopathological significances of IQGAP1 in head and neck squamous cell carcinoma (HNSCC). In this study, immunohistochemical staining of IQGAP1, co-inhibitory immune checkpoint molecules and macrophage markers were performed in human HNSCC samples to analyze the expression and correlation with clinicopathological characteristics. Immunoreactivity of IQGAP1 was also detected in immunocompetent mouse HNSCC tissue. We found that IQGAP1 expression level was significantly increased in human HNSCC compared with dysplasia and normal mucosa, and the expression of IQGAP1 in HNSCC was positively associated with advanced lymph node status. Besides, our data indicated that patients with higher IQGAP1 expression exhibited poor overall survival compared with patients with lower IQGAP1 expression. Furthermore, this study demonstrated that IQGAP1 expression was positively associated with TIM3, Galectin-9 (TIM3 ligand), B7H4, macrophage markers CD68 and CD163. In conclusion, these findings suggest that over-expression of IQGAP1 in human HNSCC may indicate poor prognosis.

Are Brain Dead Individuals Dead? Grounds for Reasonable Doubt

PubMed Central

Brugger, E. Christian

2016-01-01

According to the biological definition of death, a human body that has not lost the capacity to holistically organize itself is the body of a living human individual. Reasonable doubt against the conclusion that it has lost the capacity exists when the body appears to express it and no evidence to the contrary is sufficient to rule out reasonable doubt against the conclusion that the apparent expression is a true expression (i.e., when the conclusion that what appears to be holistic organization is in fact holistic organization remains a reasonable explanatory hypothesis in light of the best evidence to the contrary). This essay argues that the evidence and arguments against the conclusion that the signs of complex bodily integration exhibited in ventilated brain dead bodies are true expressions of somatic integration are unpersuasive; that is, they are not adequate to exclude reasonable doubt against the conclusion that BD bodies are dead. Since we should not treat as corpses what for all we know might be living human beings, it follows that we have an obligation to treat BD individuals as if they were living human beings. PMID:27075192

Interactions between ethylene and auxin are crucial to the control of grape (Vitis vinifera L.) berry ripening

PubMed Central

2013-01-01

Background Fruit development is controlled by plant hormones, but the role of hormone interactions during fruit ripening is poorly understood. Interactions between ethylene and the auxin indole-3-acetic acid (IAA) are likely to be crucial during the ripening process, since both hormones have been shown to be implicated in the control of ripening in a range of different fruit species. Results Grapevine (Vitis vinifera L.) homologues of the TRYPTOPHAN AMINOTRANSFERASE RELATED (TAR) and YUCCA families, functioning in the only characterized pathway of auxin biosynthesis, were identified and the expression of several TAR genes was shown to be induced by the pre-ripening application of the ethylene-releasing compound Ethrel. The induction of TAR expression was accompanied by increased IAA and IAA-Asp concentrations, indicative of an upregulation of auxin biosynthesis and conjugation. Exposure of ex planta, pre-ripening berries to the ethylene biosynthesis inhibitor aminoethoxyvinylglycine resulted in decreased IAA and IAA-Asp concentrations. The delayed initiation of ripening observed in Ethrel-treated berries might therefore represent an indirect ethylene effect mediated by increased auxin concentrations. During berry development, the expression of three TAR genes and one YUCCA gene was upregulated at the time of ripening initiation and/or during ripening. This increase in auxin biosynthesis gene expression was preceded by high expression levels of the ethylene biosynthesis genes 1-aminocyclopropane-1-carboxylate synthase and 1-aminocyclopropane-1-carboxylate oxidase. Conclusions In grape berries, members of both gene families involved in the two-step pathway of auxin biosynthesis are expressed, suggesting that IAA is produced through the combined action of TAR and YUCCA proteins in developing berries. The induction of TAR expression by Ethrel applications and the developmental expression patterns of auxin and ethylene biosynthesis genes indicate that elevated concentrations of ethylene prior to the initiation of ripening might lead to an increased production of IAA, suggesting a complex involvement of this auxin and its conjugates in grape berry ripening. PMID:24364881

Wnt signaling inhibits cementoblast differentiation and promotes proliferation.

PubMed

Nemoto, Eiji; Koshikawa, Yohei; Kanaya, Sousuke; Tsuchiya, Masahiro; Tamura, Masato; Somerman, Martha J; Shimauchi, Hidetoshi

2009-05-01

Cementoblasts, tooth root lining cells, are responsible for laying down cementum on the root surface, a process that is indispensable for establishing a functional periodontal ligament. Cementoblasts share phenotypical features with osteoblasts. Wnt signaling has been implicated in increased bone formation by controlling mesenchymal stem cell or osteoblastic cell functions; however the role of Wnt signaling on cementogenesis has not been examined. In this study, we have identified a consistent expression profile of Wnt signaling molecules in cementoblasts, in vitro by RT-PCR. Exposure of cells to LiCl, which promotes canonical Wnt signaling by inhibiting GSK-3beta, increased beta-catenin nuclear translocation and up-regulated the transcriptional activity of a canonical Wnt-responsive promoters, suggesting that an endogenous canonical Wnt pathway functions in cementoblasts. Activation of endogenous canonical Wnt signaling with LiCl suppressed alkaline phosphatase (ALP) activity and expression of genes associated with cementum function; ALP, bone sialoprotein (BSP), and osteocalcin (OCN). Exposure to Wnt3a, as a representative canonical Wnt member, also inhibited the expression of ALP, BSP, and OCN gene. This effect was accompanied by decreased gene expression of Runx2 and Osterix and by increased gene expression of lymphoid enhancer factor-1. Pretreatment with Dickkopf (Dkk)-1, a potent canonical Wnt antagonist, which binds to a low-density lipoprotein-receptor-related protein (LRP)-5/6 co-receptor, attenuated the suppressive effects of Wnt3a on mRNA expression of Runx2 and OCN on cementoblasts. These findings suggest that canonical Wnt signaling inhibits cementoblast differentiation via regulation of expression of selective transcription factors. Wnt3a also increased the expression of cyclin D1, known as a cell cycle regulator, as well as cell proliferation. In conclusion, these observations suggest that Wnt signaling inhibits cementoblast differentiation and promotes cell proliferation. Elucidating the role of Wnt in controlling cementoblast function will provide new tools needed to improve on existing periodontal regeneration therapies.

MicroRNA-15b deteriorates hypoxia/reoxygenation-induced cardiomyocyte apoptosis by downregulating Bcl-2 and MAPK3.

PubMed

Liu, Yaling; Yang, Liqun; Yin, Jiemin; Su, Diansan; Pan, Zhiying; Li, Peiying; Wang, Xiaodong

2018-01-01

To investigate the role of miRNA-15b in cardiomyocyte apoptosis after ischemia reperfusion injury in acute myocardial infarction (AMI), we conducted the AMI rat model by using left anterior descending ligation and performed hypoxia/reoxygenation experiments in H9c2 cells. MiRNA-15b was measured by quantitative reverse transcription PCR (qRT-PCR). Cardiomyocyte apoptosis was determined by terminal deoxynucleotide transferase dUTP nick end labeling staining. Synthesized miRNA-15b mimic and inhibitor were transfected into H9c2 cells by Lipofectamine regent. RNA expression of B cell lymphoma/leukemia-2 (Bcl-2) and mitogen-activated protein kinase 3 (MAPK3) was examined by qRT-PCR and their protein expression was determined by western blot. Ischemia reperfusion increased miRNA-15b expression in the ischemic rat heart and resulted more severe cardiomyocytes apoptosis. In H9c2 cells, hypoxia/reoxygenation induced increased miRNA-15b expression and augmented cardiomyocyte apoptosis observed at 24 hours after 24-hour hypoxia. Compared with the vehicle group, miRNA-15b mimic further raised miRNA-15b level and increased cardiomyocyte apoptosis, whereas miRNA-15b inhibitor suppressed miRNA-15b expression and protected cardiomyocytes from apoptosis. Although the mRNA expression of the target genes Bcl-2 and MAPK3 was not changed significantly, the protein expression of these two genes were markedly reduced after miRNA-15b mimic treatment and significantly increased after transfected with miRNA-15b inhibitors. In conclusion, miRNA-15b deteriorates cardiomyocyte apoptosis by post-transcriptionally downregulating the expression of Bcl-2 and MAPK3. © American Federation for Medical Research (unless otherwise stated in the text of the article) 2018. All rights reserved. No commercial use is permitted unless otherwise expressly granted.

Analysis of the temporal expression of chemokines and chemokine receptors during experimental granulomatous inflammation: role and expression of MIP-1α and MCP-1

PubMed Central

Carollo, Maria; Hogaboam, Cory M; Kunkel, Stephen L; Delaney, Stephen; Christie, Mark I; Perretti, Mauro

2001-01-01

Chemokine expression and function was monitored in an experimental model of granulomatous tissue formation after injection of croton oil in complete Freund's adjuvant (CO/CFA) into mouse dorsal air-pouches up to 28 days. In the first week, mast cell degranulation and leukocyte influx (mononuclear cell, MNC, and polymorphonuclear cell, PMN) were associated with CXCR2, KC and macrophage inflammatory protein (MIP)-2 mRNA expression, as determined by TaqMan® reverse transcriptase-polymerase chain reaction. KC (∼400 pg mg protein−1, n=12) and MIP-2 (∼800 pg mg protein−1, n=12) proteins peaked at day 7, together with myeloperoxidase (MPO) activity. Highest MIP-1α (>1 ng mg protein−1, n=12) levels were measured at day 3. After day 7, a gradual increase in CCR2 and CCR5 mRNA, monocyte chemoattractant protein (MCP)-1 mRNA and protein expression was measured. MCP-1 protein peaked at day 21 (∼150 pg mg protein−1, n=12) and was predominantly expressed by mast cells. A gradual increase in N-acetyl-β-D-glucosaminidase (NAG) activity (maximal at 28 days) was also measured. An antiserum against MIP-1α did not modify the inflammatory response measured at day 7 (except for a 50% reduction in MIP-1α levels), but provoked a significant increase in MPO, NAG and MCP-1 levels as measured at day 21 (n=6, P<0.05). An antiserum to MCP-1 reduced NAG activity at day 21 but increased MPO activity values (n=8, P<0.05). In conclusion, we have shown that CO/CFA initiates a complex inflammatory reaction in which initial expression of MIP-1α serves a protective role whereas delayed expression of MCP-1 seems to have a genuine pro-inflammatory role. PMID:11704636

Increased SHP-1 Protein Expression by High Glucose Levels Reduces Nephrin Phosphorylation in Podocytes*

PubMed Central

Denhez, Benoit; Lizotte, Farah; Guimond, Marie-Odile; Jones, Nina; Takano, Tomoko; Geraldes, Pedro

2015-01-01

Nephrin, a critical podocyte membrane component that is reduced in diabetic nephropathy, has been shown to activate phosphotyrosine signaling pathways in human podocytes. Nephrin signaling is important to reduce cell death induced by apoptotic stimuli. We have shown previously that high glucose level exposure and diabetes increased the expression of SHP-1, causing podocyte apoptosis. SHP-1 possesses two Src homology 2 domains that serve as docking elements to dephosphorylate tyrosine residues of target proteins. However, it remains unknown whether SHP-1 interacts with nephrin and whether its elevated expression affects the nephrin phosphorylation state in diabetes. Here we show that human podocytes exposed to high glucose levels exhibited elevated expression of SHP-1, which was associated with nephrin. Coexpression of nephrin-CD16 and SHP-1 reduced nephrin tyrosine phosphorylation in transfected human embryonic kidney 293 cells. A single tyrosine-to-phenylalanine mutation revealed that rat nephrin Tyr1127 and Tyr1152 are required to allow SHP-1 interaction with nephrin. Overexpression of dominant negative SHP-1 in human podocytes prevented high glucose-induced reduction of nephrin phosphorylation. In vivo, immunoblot analysis demonstrated that nephrin expression and phosphorylation were decreased in glomeruli of type 1 diabetic Akita mice (Ins2+/C96Y) compared with control littermate mice (Ins2+/+), and this was associated with elevated SHP-1 and cleaved caspase-3 expression. Furthermore, immunofluorescence analysis indicated increased colocalization of SHP-1 with nephrin in diabetic mice compared with control littermates. In conclusion, our results demonstrate that high glucose exposure increases SHP-1 interaction with nephrin, causing decreased nephrin phosphorylation, which may, in turn, contribute to diabetic nephropathy. PMID:25404734

MiR-199a-3p enhances cisplatin sensitivity of cholangiocarcinoma cells by inhibiting mTOR signaling pathway and expression of MDR1.

PubMed

Li, Qiang; Xia, Xuefeng; Ji, Jie; Ma, Jianghui; Tao, Liang; Mo, Linjun; Chen, Wei

2017-05-16

Several studies have reported reduced miRNA-199a-3p (miR-199a-3p) in different human malignancies, however, little is known about miR-199a-3p in cholangiocarcinoma cells. In this study, we demonstrate the essential role and mechanism of miR-199a-3p in regulating cisplatin sensitivity in cholangiocarcinoma cell lines. Using a CCK-8 cell counting assay we found that expression of miR-199a-3p was positively correlated with cisplatin sensitivity in cholangiocarcinoma cell lines. MiR-199a-3p overexpression could decrease the proliferation rate and increase apoptosis of cholangiocarcinoma cells in the presence of cisplatin, while miR-199a-3p inhibition had the opposite effect. Further study demonstrated that mTOR was the target gene of miR-199a-3p, and that miR-199a-3p mimics could inhibit expression of mTOR, which consequently reduced the phosphorylation of its downstream proteins 4EBP1 and p70s6k. Rescue experiments proved that miR-199a-3p could increase the cisplatin sensitivity of cholangiocarcinoma cell lines by regulating mTOR expression. Moreover, we also found that miR-199a-3p overexpression could reduce cisplatin induced MDR1 expression by decreasing the synthesis and increasing the degradation of MDR1, thus enhancing the effectiveness of cisplatin in cholangiocarcinoma. In conclusion, miR-199a-3p could increase cisplatin sensitivity of cholangiocarcinoma cell lines by inhibiting the activity of the mTOR signaling pathway and decreasing the expression of MDR1.

Overexpression of L-Type Amino Acid Transporter 1 (LAT1) and 2 (LAT2): Novel Markers of Neuroendocrine Tumors

PubMed Central

Barollo, Susi; Bertazza, Loris; Watutantrige-Fernando, Sara; Censi, Simona; Cavedon, Elisabetta; Galuppini, Francesca; Pennelli, Gianmaria; Fassina, Ambrogio; Citton, Marilisa; Rubin, Beatrice; Pezzani, Raffaele; Benna, Clara; Opocher, Giuseppe; Iacobone, Maurizio; Mian, Caterina

2016-01-01

Background 6-18F-fluoro-L-3,4-dihydroxyphenylalanine (18F-FDOPA) PET is a useful tool in the clinical management of pheochromocytoma (PHEO) and medullary thyroid carcinoma (MTC). 18F-FDOPA is a large neutral amino acid biochemically resembling endogenous L-DOPA and taken up by the L-type amino acid transporters (LAT1 and LAT2). This study was conducted to examine the expression of the LAT system in PHEO and MTC. Methods Real-time PCR and Western blot analyses were used to assess LAT1 and LAT2 gene and protein expression in 32 PHEO, 38 MTC, 16 normal adrenal medulla and 15 normal thyroid tissue samples. Immunohistochemistry method was applied to identify the proteins’ subcellular localization. Results LAT1 and LAT2 were overexpressed in both PHEO and MTC by comparison with normal tissues. LAT1 presented a stronger induction than LAT2, and their greater expression was more evident in PHEO (15.1- and 4.1-fold increases, respectively) than in MTC (9.9- and 4.1-fold increases, respectively). Furthermore we found a good correlation between LAT1/2 and GLUT1 expression levels. A positive correlation was also found between urinary noradrenaline and adrenaline levels and LAT1 gene expression in PHEO. The increased expression of LAT1 is also confirmed at the protein level, in both PHEO and MTC, with a strong cytoplasmic localization. Conclusions The present study is the first to provide experimental evidence of the overexpression in some NET cancers (such as PHEO or MTC) of L-type amino acid transporters, and the LAT1 isoform in particular, giving the molecular basis to explain the increase of the DOPA uptake seen in such tumor cells. PMID:27224648

Inhibition of nitric oxide production reverses diabetes-induced Kupffer cell activation and Klebsiella pneumonia liver translocation

PubMed Central

Wu, Ying-Ying; Fung, Chang-Phone; Hsu, Ching-Mei

2017-01-01

Klebsiella pneumoniae (KP) is the most common pathogen of pyogenic liver abscess in East and Southeast Asia and diabetes mellitus (DM) is a major risk factor. The effect and mechanism of diabetes on KP liver abscess was examined in streptozotocin-induced diabetic mice and Akita mice (C57BL/6J-Ins2Akita). KP translocation to liver and plasma alaine transaminase levels were increased and liver clearance of KP was decreased in DM mice. Diabetic mice exhibited overgrowth of Enterococcus as well as E.coli and decreased lactobacilli/bifidas growth in intestine, increased intestinal iNOS protein and nitrite levels in portal vein, and increased IL-1β and TNF-α expression of Kupffer cells. Fructooligosaccharides (FOS) or dead L. salivarius (dLac) supplementation reversed diabetes-induced enteric dysbiosis, NO levels in portal vein, and KP translocation to liver. L-NAME treatment decreased intestinal iNOS protein expression as well as Kupffer cell activation and increased liver clearance of KP in DM mice. Dead E.coli (2×108 CFU/ml) feeding for one week induced iNOS and TLR4 expression of intestine in germ-free (GF) mice. Dead bacteria feeding induced IL-1β and TNF-α expression of Kupffer cells in GF mice but not in GF TLR4-/- mice. In conclusion, balance of intestinal microflora is important for preventing intestinal iNOS expression, Kupffer cell activation, and KP liver translocation in diabetes. Reversal of diabetes-induced enteric dysbiosis with FOS or dead L. salivarius decreases diabetes-induced intestinal iNOS expression and KP liver translocation. Diabetes induces Kupffer cell activation and KP liver translocation through enteric dysbiosis and nitric oxide production. PMID:28493939

Effect of scuba diving on the oxidant/antioxidant status, SIRT1 and SIRT3 expression in recreational divers after a winter nondive period.

PubMed

Perović, Antonija; Sobočanec, Sandra; Dabelić, Sanja; Balog, Tihomir; Dumić, Jerka

2018-02-01

The aim of this study was to examine the effects of scuba diving on oxidative damage markers in erythrocytes and plasma, antioxidant system in peripheral blood mononuclear cells (PBMCs), as well as sirtuin 1 (SIRT1) and sirtuin 3 (SIRT3) gene expressions in recreational divers after a winter nondive period (at least 5 months). For that purpose, 17 male recreational divers performed an immersion at a depth of 30 m for 30 min. Blood samples were collected immediately before and after diving, 3 and 6 h after diving. Erythrocyte lipid peroxidation measured by thiobarbituric-reactive substances (TBARS) method was significantly increased immediately after diving, but returned to the baseline 6 h after diving, while no significant change was found for plasma TBARS and protein carbonyl derivates in both plasma and erythrocytes. Diving-induced catalase (CAT), superoxide dismutase 2 (SOD2), and consequently total superoxide dismutase (SOD) activities in the PBMC samples (significantly increased immediately after diving, reached the maximum activities 3 h after diving, while 6 h after diving only CAT activity remained significantly increased). No significant change was observed for SOD1 activity and gene expression, as well as SOD2 expression, while CAT and SIRT1 expressions were slightly decreased immediately after diving and 3 h after diving. Interestingly, SIRT3 expression was significantly increased 6 h after diving. In conclusion, after the first dive to 30 m after a nondive season, activation of antioxidant defence was not sufficient to prevent oxidative damage, while SIRT3 upregulation could be a step towards an adaptive response to the diving.

Heat Shock Factor 1 Epigenetically Stimulates Glutaminase-1-Dependent mTOR Activation to Promote Colorectal Carcinogenesis.

PubMed

Li, Jiaqiu; Song, Ping; Jiang, Tingting; Dai, Dongjun; Wang, Hanying; Sun, Jie; Zhu, Liyuan; Xu, Wenxia; Feng, Lifeng; Shin, Vivian Y; Morrison, Helen; Wang, Xian; Jin, Hongchuan

2018-04-14

Heat shock factor 1 (HSF1) generally exhibits its properties under stress conditions. In tumors, HSF1 has a pleiotropic feature in regulating growth, survival, and aggressiveness of cancer cells. In this study, we found HSF1 was increased in colorectal cancer (CRC) and had a positive correlation with shorter disease-free survival (DFS). Knockdown of HSF1 in CRC cells attenuated their growth while inhibiting mTOR activation and glutamine metabolism. HSF1 inhibited the expression of microRNA137 (MIR137), which targeted GLS1 (glutaminase 1), thus stimulating GLS1 protein expression to promote glutaminolysis and mTOR activation. HSF1 bound DNA methyltransferase DNMT3a and recruited it to the promoter of lncRNA MIR137 host gene (MIR137HG), suppressing the generation of primary MIR137. The chemical inhibitor of HSF1 also reduced cell growth, increased apoptosis, and impaired glutamine metabolism in vitro. Moreover, both chemical inhibition and genetic knockout of HSF1 succeeded in increasing MIR137 expression, reducing GLS1 expression, and alleviating colorectal tumorigenesis in azoxymethane (AOM)/dextran sulfate sodium (DSS) mice. In conclusion, HSF1 expression was increased and associated with poor prognosis in CRC. By recruiting DNMT3a to suppress the expression of MIR137 that targets GLS1 mRNA, HSF1 stimulated GLS1-dependent mTOR activation to promote colorectal carcinogenesis. Therefore, targeting HSF1 to attenuate glutaminolysis and mTOR activation could be a promising approach for CRC treatment. Copyright © 2018 The American Society of Gene and Cell Therapy. Published by Elsevier Inc. All rights reserved.

Methionine Deprivation Induces a Targetable Vulnerability in Triple-negative Breast Cancer Cells by Enhancing TRAIL Receptor-2 Expression

PubMed Central

Strekalova, Elena; Malin, Dmitry; Good, David M.; Cryns, Vincent L.

2015-01-01

Purpose Many neoplasms are vulnerable to methionine deficiency by mechanisms that are poorly understood. Because gene profiling studies have revealed that methionine depletion increases TNF-related apoptosis-inducing ligand receptor-2 (TRAIL-R2) mRNA, we postulated that methionine stress sensitizes breast cancer cells to proapoptotic TRAIL-R2 agonists. Experimental Design Human triple (ER/PR/HER2)-negative breast carcinoma cell lines were cultured in control or methionine-free media. The effects of methionine depletion on TRAIL receptor expression and sensitivity to chemotherapy or a humanized agonistic TRAIL-R2 monoclonal antibody (lexatumumab) were determined. The melanoma-associated antigen MAGED2 was silenced to delineate its functional role in sensitizing TNBC cells to methionine stress. An orthotopic TNBC model was utilized to evaluate the effects of dietary methionine deficiency, lexatumumab or the combination. Results Methionine depletion sensitized TNBC cells to lexatumumab-induced caspase activation and apoptosis by increasing TRAIL-R2 mRNA and cell surface expression. MCF-10A cells transformed by oncogenic H-Ras, but not untransformed cells, and matrix-detached TNBC cells were highly sensitive to the combination of lexatumumab and methionine depletion. Proteomics analyses revealed that MAGED2, which has been reported to reduce TRAIL-R2 expression, was suppressed by methionine stress. Silencing MAGED2 recapitulated features of methionine deprivation, including enhanced mRNA and cell surface expression of TRAIL receptors and increased sensitivity to TRAIL receptor agonists. Dietary methionine deprivation enhanced the antitumor effects of lexatumumab in an orthotopic metastatic TNBC model. Conclusion Methionine depletion exposes a targetable defect in TNBC cells by increasing TRAIL-R2 expression. Our findings provide the foundation for a clinical trial combining dietary methionine restriction and TRAIL-R2 agonists. PMID:25724522

PDGF-α stimulates intestinal epithelial cell turnover after massive small bowel resection in a rat.

PubMed

Sukhotnik, Igor; Mogilner, Jorge G; Pollak, Yulia; Blumenfeld, Shiri; Bejar, Jacob; Coran, Arnold G

2012-06-01

Numerous cytokines have been shown to affect epithelial cell differentiation and proliferation through epithelial-mesenchymal interaction. Growing evidence suggests that platelet-derived growth factor (PDGF) signaling is an important mediator of these interactions. The purpose of this study was to evaluate the effect of PDGF-α on enterocyte turnover in a rat model of short bowel syndrome (SBS). Male rats were divided into four groups: Sham rats underwent bowel transection, Sham-PDGF-α rats underwent bowel transection and were treated with PDGF-α, SBS rats underwent a 75% bowel resection, and SBS-PDGF-α rats underwent bowel resection and were treated with PDGF-α. Parameters of intestinal adaptation, enterocyte proliferation and apoptosis were determined at euthanasia. Illumina's Digital Gene Expression analysis was used to determine PDGF-related gene expression profiling. PDGF-α and PDGF-α receptor (PDGFR-α) expression was determined by real-time PCR. Western blotting was used to determine p-ERK, Akt1/2/3, bax, and bcl-2 protein levels. SBS rats demonstrated a significant increase in PDGF-α and PDGFR-α expression in jejunum and ileum compared with sham animals. SBS-PDGF-α rats demonstrated a significant increase in bowel and mucosal weight, villus height, and crypt depth in jejunum and ileum compared with SBS animals. PDGF-α receptor expression in crypts increased in SBS rats (vs. sham) and was accompanied by an increased cell proliferation following PDGF-α administration. A significant decrease in cell apoptosis in this group was correlated with lower bax protein levels. In conclusion, in a rat model of SBS, PDGF-α stimulates enterocyte turnover, which is correlated with upregulated PDGF-α receptor expression in the remaining small intestine.

Antimicrobial Barrier of an in vitro Oral Epithelial Model

PubMed Central

Kimball, Janet R.; Nittayananta, Wipawee; Klausner, Mitchell; Chung, Whasun O.; Dale, Beverly A.

2008-01-01

Objective Oral epithelia function as a microbial barrier and are actively involved in recognizing and responding to bacteria. Our goal was to examine a tissue engineered model of buccal epithelium for its response to oral bacteria and proinflammatory cytokines and compare the tissue responses with those of a submerged monolayer cell culture. Design The tissue model was characterized for keratin and β-defensin expression. Altered expression of β-defensins was evaluated by RT-PCR after exposure of the apical surface to oral bacteria and after exposure to TNF-α in the medium. These were compared to the response in traditional submerged oral epithelial cell culture. Results The buccal model showed expression of differentiation specific keratin 13, hBD1 and hBD3 in the upper half of the tissue; hBD2 was not detected. hBD1 mRNA was constitutively expressed, while hBD2 mRNA increased 2-fold after exposure of the apical surface to three oral bacteria tested and hBD3 mRNA increased in response to the non-pathogenic bacteria tested. In contrast, hBD2 mRNA increased 3–600 fold in response to bacteria in submerged cell culture. HBD2 mRNA increased over 100 fold in response to TNF-α in the tissue model and 50 fold in submerged cell culture. Thus, the tissue model is capable of upregulating hBD2, however, the minimal response to bacteria suggests that the tissue has an effective antimicrobial barrier due to its morphology, differentiation, and defensin expression. Conclusions The oral mucosal model is differentiated, expresses hBD1 and hBD3, and has an intact surface with a functional antimicrobial barrier. PMID:16815238

Angiopoietin-like 7 Secretion Is Induced by Glaucoma Stimuli and Its Concentration Is Elevated in Glaucomatous Aqueous Humor

PubMed Central

Kuchtey, John; Källberg, Maria E.; Gelatt, Kirk N.; Rinkoski, Tommy; Komàromy, András M.; Kuchtey, Rachel W.

2010-01-01

Purpose To investigate the possibility that Angiopoietin-like 7 (ANGPTL7) protein is involved in the pathogenesis of glaucoma. Methods Primary human trabecular meshwork (TM) cells and corneoscleral explants were stimulated with either dexamethasone (DEX) or transforming growth factor β (TGFβ), and ANGPTL7 protein secreted into culture medium was determined by Western blot analysis. The effect of stable overexpression of ANGPTL7 in transfected immortalized TM cell lines on collagen expression was investigated by immunocytochemistry. Localization of ANGPTL7 protein in human eyes was determined by immunohistochemistry. The concentration of ANGPTL7 protein in aqueous humor (AH) from patients with glaucoma and control patients was compared by Western blot analysis. The beagle model of primary open-angle glaucoma (POAG) was used to correlate ANGPTL7 protein levels in canine AH with disease progression. Results TGFβ and DEX stimulated secretion of ANGPTL7 protein by TM cells and corneoscleral explants. Overexpression of ANGPTL7 by immortalized TM cell lines increased expression of type I collagen. Expression of ANGPTL7 protein was located in the corneal stroma, near the limbus, and throughout the sclera, with lower expression in the TM. In the lamina cribrosa, ANGPTL7 expression was associated with the cribriform plates. The concentration of ANGPTL7 protein was elevated in AH from patients with glaucoma and increased as disease progressed in POAG beagle dogs. Conclusions Induction of ANGPTL7 secretion by glaucoma stimuli and increased concentration of ANGPTL7 in glaucomatous AH suggest that ANGPTL7 is overexpressed in glaucoma. Since overexpression of ANGPTL7 increases collagen expression, a potential disease mechanism, ANGPTL7 could have a pathogenic role in glaucoma, and may serve as a potential therapeutic target. PMID:18421092

Curcumin regulates gene expression of insulin like growth factor, B-cell CLL/lymphoma 2 and antioxidant enzymes in streptozotocin induced diabetic rats

PubMed Central

2013-01-01

Background The effects of curcumin on the activities and gene expression of antioxidant enzymes, superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPX), glutathione-S-transferase (G-ST), B-cell CLL/lymphoma 2 (Bcl-2) and insulin like growth factor-1 (IGF-1) in diabetic rats were studied. Methods Twenty four rats were assigned to three groups (8 rats for each). Rats of first group were non diabetic and rats of the second group were rendered diabetic by streptozotocin (STZ). Both groups received vehicle, corn oil only (5 ml/kg body weight) and served as negative and positive controls, respectively. Rats of the third group were rendered diabetic and received oral curcumin dissolved in corn oil at a dose of 15 mg/5 ml/kg body weight for 6 weeks. Results Diabetic rats showed significant increase of blood glucose, thiobarbituric acid reactive substances (TBARS) and activities of all antioxidant enzymes with significant reduction of reduced glutathione (GSH) compare to the control non diabetic group. Gene expression of Bcl2, SOD, CAT, GPX and GST was increased significantly in diabetic untreated rats compare to the control non diabetic group. The administration of curcumin to diabetic rats normalized significantly their blood sugar level and TBARS values and increased the activities of all antioxidant enzymes and GSH concentration. In addition, curcumin treated rats showed significant increase in gene expression of IGF-1, Bcl2, SOD and GST compare to non diabetic and diabetic untreated rats. Conclusion Curcumin was antidiabetic therapy, induced hypoglycemia by up-regulation of IGF-1 gene and ameliorate the diabetes induced oxidative stress via increasing the availability of GSH, increasing the activities and gene expression of antioxidant enzymes and Bcl2. Further studies are required to investigate the actual mechanism of action of curcumin regarding the up regulation of gene expression of examined parameters. PMID:24364912

PHEX Mimetic (SPR4-Peptide) Corrects and Improves HYP and Wild Type Mice Energy-Metabolism

PubMed Central

Zelenchuk, Lesya V.; Hedge, Anne-Marie; Rowe, Peter S. N.

2014-01-01

Context PHEX or DMP1 mutations cause hypophosphatemic-rickets and altered energy metabolism. PHEX binds to DMP1-ASARM-motif to form a complex with α5β3 integrin that suppresses FGF23 expression. ASARM-peptides increase FGF23 by disrupting the PHEX-DMP1-Integrin complex. We used a 4.2 kDa peptide (SPR4) that binds to ASARM-peptide/motif to study the DMP1-PHEX interaction and to assess SPR4 for the treatment of energy metabolism defects in HYP and potentially other bone-mineral disorders. Design Subcutaneously transplanted osmotic pumps were used to infuse SPR4-peptide or vehicle (VE) into wild-type mice (WT) and HYP-mice (PHEX mutation) for 4 weeks. Results SPR4 partially corrected HYP mice hypophosphatemia and increased serum 1.25(OH)2D3. Serum FGF23 remained high and PTH was unaffected. WT-SPR4 mice developed hypophosphatemia and hypercalcemia with increased PTH, FGF23 and 1.25(OH)2D3. SPR4 increased GAPDH HYP-bone expression 60× and corrected HYP-mice hyperglycemia and hypoinsulinemia. HYP-VE serum uric-acid (UA) levels were reduced and SPR4 infusion suppressed UA levels in WT-mice but not HYP-mice. SPR4 altered leptin, adiponectin, and sympathetic-tone and increased the fat mass/weight ratio for HYP and WT mice. Expression of perlipin-2 a gene involved in obesity was reduced in HYP-VE and WT-SPR4 mice but increased in HYP-SPR4 mice. Also, increased expression of two genes that inhibit insulin-signaling, ENPP1 and ESP, occurred with HYP-VE mice. In contrast, SPR4 reduced expression of both ENPP1 and ESP in WT mice and suppressed ENPP1 in HYP mice. Increased expression of FAM20C and sclerostin occurred with HYP-VE mice. SPR4 suppressed expression of FAM20C and sclerostin in HYP and WT mice. Conclusions ASARM peptides and motifs are physiological substrates for PHEX and modulate osteocyte PHEX-DMP1-α5β3-integrin interactions and thereby FGF23 expression. These interactions also provide a nexus that regulates bone and energy metabolism. SPR4 suppression of sclerostin and/or sequestration of ASARM-peptides improves energy metabolism and may have utility for treating familial rickets, osteoporosis, obesity and diabetes. PMID:24839967

Reduced Expression of SARM in Mouse Spleen during Polymicrobial Sepsis.

PubMed

Gong, Yu; Zou, Lin; Cen, Dongzhi; Chao, Wei; Chen, Dunjin

2016-12-01

Objective Immune dysfunction, including prominent apoptosis of immune cells and decreased functioning of the remaining immune cells, plays a central role in the pathogenesis of sepsis. Sterile α and HEAT/armadillo motif-containing protein (SARM) is implicated in the regulation of immune cell apoptosis. This study aimed to elucidate SARM contributes to sepsis-induced immune cell death and immunosuppression. Methods A mouse model of polymicrobial sepsis was generated by cecum ligation and puncture (CLP). SARM gene and protein expression, caspase 3 cleavage and intracellular ATP production were measured in the mouse spleens. Results CLP-induced polymicrobial sepsis specifically attenuated both the gene and protein expression of SARM in the spleens. Moreover, the attenuation of SARM expression synchronized with splenocyte apoptosis, as evidenced by increased caspase 3 cleavage and ATP depletion. Conclusions These findings suggest that SARM is a potential regulator of sepsis-induced splenocyte apoptosis.

The mRNA expression of amino acid and sugar transporters, aminopeptidase, as well as the di- and tri-peptide transporter PepT1 in the intestines of Eimeria infected broiler chickens.

PubMed

Miska, K B; Fetterer, R H

2017-02-01

Coccidiosis in chickens is caused by infection of gut epithelial cells with protozoan parasites of the genus Eimeria This disease causes losses to the poultry industry since infected birds fail to gain weight as rapidly as non-infected birds and efficiency of feed conversion is compromised. For the present study the effect of Eimeria on expression of components of amino acid and sugar uptake mechanisms was determined. Broiler chicks were infected with Eimeria maxima, which infects the jejunum; Eimeria acervulina, which infects the duodenum; or Eimeria tenella, which infects the ceca. Sections of the jejunum, duodenum, and ceca (depending on species of Eimeria) were taken at several time points between d zero and 14 post infection (PI) for mRNA expression analysis. Genes examined included one digestive enzyme, 7 peptide and amino acid transporters located on the brush border, 8 transporters located at the basolateral surface of the gut epithelium, and 5 sugar transporters. All 3 Eimeria species examined caused decrease in expression of brush border transporters particularly at d 5 to 7 PI, which corresponds to the time when pathology is greatest. The same pattern was seen in expression of sugar transporters. However, the expression of basolateral transporters differed among species. Eimeria tenella infection resulted in decreased expression of all basolateral transporters, while E. maxima infection caused increased expression of 2 genes and slight decrease in expression of the remaining 5 genes. Infection with E. acervulina resulted in increased expression at the height of infection of all but one basolateral transporter. In conclusion, Eimeria infection causes a general decrease in gene expression of sugar transporter and brush border AATs at the height of infection. However the expression of basolateral transporters is increased in E. maxima and E. acervulina infected birds. It is possible that decreased expression of brush border transporters in combination with increased expression of basolateral transporters leads to decrease of nutrients available for the parasite, thus limiting parasite reproduction. Published by Oxford University Press on behalf of Poultry Science Association 2016. This work is written by (a) US Government employee(s) and is in the public domain in the US.

Expression and regulation of enzymes in the ceramide metabolic pathway in human retinal pigment epithelial cells and their relevance to retinal degeneration.

PubMed

Zhu, DanHong; Sreekumar, Parameswaran G; Hinton, David R; Kannan, Ram

2010-03-31

Ceramide and its metabolic derivatives are important modulators of cellular apoptosis and proliferation. Dysregulation or imbalance of their metabolic pathways may promote the development of retinal degeneration. The aim of this study was to identify the expression and regulation of key enzymes of the ceramide pathway in retinal pigment epithelial (RPE) cells. RT-PCR was used to screen the enzymes involved in ceramide metabolism that are expressed in RPE. Over-expression of neutral sphingomyelinase-2 (SMPD3) or sphingosine kinase 1 (Sphk1) in ARPE-19 cells was achieved by transient transfection of SMPD3 or Sphk1 cDNA subcloned into an expression vector. The number of apoptotic or proliferating cells was determined using TUNEL and BrdU assays, respectively. Neutral sphingomyelinase-1, neutral sphingomyelinase-2, acidic ceramidase, ceramide kinase, SphK1 and Sphk2 were expressed in both ARPE-19 and early passage human fetal RPE (fRPE) cells, while alkaline ceramidase 2 was only expressed in fRPE cells. Over-expression of SMPD3 decreased RPE cell proliferation and increased cell apoptosis. The percentage of apoptotic cells increased proportionally with the amount of transfected SMPD3 DNA. Over-expression of SphK1 promoted cell proliferation and protected ARPE-19 cells from ceramide-induced apoptosis. The effect of C(2) ceramide on induction of apoptosis was evaluated in polarized vs. non-polarized RPE cultures; polarization of RPE was associated with much reduced apoptosis in response to ceramide. In conclusion, RPE cells possess the synthetic machinery for the production of ceramide, sphingosine, ceramide-1-phosphate (C1P), and sphingosine-1-phosphate (S1P). Over-expression of SMPD3 may increase cellular ceramide levels, leading to enhanced cell death and arrested cell proliferation. The selective induction of apoptosis in non-polarized RPE cultures by C(2) ceramide suggests that increased ceramide levels will preferentially affect non-polarized RPE, as are found in late age-related macular degeneration lesions, and may spare the normal RPE monolayer. SphK1 over-expression increased cellular S1P, which promoted cell proliferation and protected RPE from ceramide-induced apoptosis. Understanding the relationship between the metabolism of sphingolipids and their effects in RPE cell survival/death may help us to develop effective and efficient therapies for retinal degeneration. Copyright 2009 Elsevier Ltd. All rights reserved.

VEGF165 Stimulates Vessel Density and Vessel Diameter Differently in Angiogenesis and Lymphangiogenesis

NASA Technical Reports Server (NTRS)

Parsons-Wingerter, Patricia; Radhakrishnan, Krishnan; DiCorleto, Paul E.; Leontiev, Dmitry; Anand-Apte, Bela; Albarran, Brian; Farr, Andrew G.

2005-01-01

Vascular endothelial growth factor-165 (VEGF(sub 165)) stimulated angiogenesis in the quail chorioallantoic membrane (CAM) by vessel expansion from the capillary network. However, lymphangiogenesis was stimulated by the filopodial guidance of tip cells located on blind-ended lymphatic sprouts. As quantified by fractal/generational branching analysis using the computer code VESGEN, vascular density increased maximally at low VEGF concentrations, and vascular diameter increased most at high VEGF concentrations. Increased vascular density and diameter were statistically independent events (r(sub s), -0.06). By fluorescence immunohistochemistry of VEGF receptors VEGFR-1 and VEGFR-2, alpha smooth muscle actin ((alpha) SMA) and a vascular/lymphatic marker, VEGF(sub 165) increased the density and diameter of sprouting lymphatic vessels guided by tip cells (accompanied by the dissociation of lymphatics from blood vessels). Isolated migratory cells expressing (alpha)SMA were recruited to blood vessels, whereas isolated cells expressing VEGFR-2 were recruited primarily to lymphatics. In conclusion, VEGF(sub 165) increased lymphatic vessel density by lymphatic sprouting, but increased blood vessel density by vascular expansion from the capillary network.

Peroxisome proliferator-activated receptor {alpha} agonist-induced down-regulation of hepatic glucocorticoid receptor expression in SD rats

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

Chen Xiang; Li Ming; Sun Weiping

2008-04-18

It was reported that glucocorticoid production was inhibited by fenofibrate through suppression of type-1 11{beta}-hydroxysteroid dehydrogenase gene expression in liver. The inhibition might be a negative-feedback regulation of glucocorticoid receptor (GR) activity by peroxisome proliferator-activated receptor alpha (PPAR{alpha}), which is quickly induced by glucocorticoid in the liver. However, it is not clear if GR expression is changed by fenofibrate-induced PPAR{alpha} activation. In this study, we tested this possibility in the liver of Sprague-Dawley rats. GR expression was reduced by fenofibrate in a time- and does-dependent manner. The inhibition was observed in liver, but not in fat and muscle. The corticosteronemore » level in the blood was increased significantly by fenofibrate. These effects of fenofibrate were abolished by PPAR{alpha} inhibitor MK886, suggesting that fenofibrate activated through PPAR{alpha}. In conclusion, inhibition of GR expression may represent a new molecular mechanism for the negative feedback regulation of GR activity by PPAR{alpha}.« less

RGC32 induces epithelial-mesenchymal transition by activating the Smad/Sip1 signaling pathway in CRC.

PubMed

Wang, Xiao-Yan; Li, Sheng-Nan; Zhu, Hui-Fang; Hu, Zhi-Yan; Zhong, Yan; Gu, Chuan-Sha; Chen, Shi-You; Liu, Teng-Fei; Li, Zu-Guo

2017-05-04

Response gene to complement 32 (RGC32) is a transcription factor that regulates the expression of multiple genes involved in cell growth, viability and tissue-specific differentiation. However, the role of RGC32 in tumorigenesis and tumor progression in colorectal cancer (CRC) has not been fully elucidated. Here, we showed that the expression of RGC32 was significantly up-regulated in human CRC tissues versus adjacent normal tissues. RGC32 expression was significantly correlated with invasive and aggressive characteristics of tumor cells, as well as poor survival of CRC patients. We also demonstrated that RGC32 overexpression promoted proliferation, migration and tumorigenic growth of human CRC cells in vitro and in vivo. Functionally, RGC32 facilitated epithelial-mesenchymal transition (EMT) in CRC via the Smad/Sip1 signaling pathway, as shown by decreasing E-cadherin expression and increasing vimentin expression. In conclusion, our findings suggested that overexpression of RGC32 facilitates EMT of CRC cells by activating Smad/Sip1 signaling.

[CD69 expression on T cell surface in patients with obstructive sleep apnea hypopnea syndrome].

PubMed

Chen, Y H; Wang, P F; Wang, H J; Hu, X; Li, Z Y; Xiong, H Q

2017-02-20

Objective: To investigate the detection and significance of T cell CD69 expression in peripheral blood of patients with obstructive sleep apnea hypopnea syndrome (OSAHS). Method: According to AHI, 81 OSAHS patients diagnosed by PSG were divided into 3 groups: light, medium and heavy, with 27 cases in each group; 27 patients without OSAHS as control group. Flow cytometry was used to detect the expression rate of CD69 in T cells, to analyze the correlation between the expression rate of CD69 on T cells and the gender, age, BMI, and PSG index. Result: ①The CD69 expression rate of T cells in peripheral blood of OSAHS patients with snoring degree increases gradually ( P < 0.05); Comparison between the two shows that there was no significant difference in CD69 expression rate on T cells between the control group and the mild group ( t = 1.649, P > 0.05); there were significant differences between the other groups ( P < 0.05). ②The CD69 expression rate of T cells in peripheral blood of OSAHS patients has no correlation with BMI, age and gender ( P > 0.05), were positively correlated with AHI, negatively correlated with LSaO₂ ( P < 0.01). ③The CD69 expression rate of T cells and AHI in 27 cases of severe OSAHS patients with a comprehensive treatment has significantly reduced, LSaO₂ increased significantly ( P < 0.01). Conclusion: Increased expression of CD69 in peripheral blood T cells may be one of the mechanisms of OSAHS complicated with cardiovascular disease. Detection of CD69 expression rate in T cells for reflecting the degree of disease in patients with OSAHS, assessment of risk of cardiovascular damage, have certain clinical significance. Copyright© by the Editorial Department of Journal of Clinical Otorhinolaryngology Head and Neck Surgery.

Effects of apocynin on oxidative stress and expression of apoptosis-related genes in testes of diabetic rats.

PubMed

Li, Mingchao; Liu, Zhuo; Zhuan, Li; Wang, Tao; Guo, Shuiming; Wang, Shaogang; Liu, Jihong; Ye, Zhangqun

2013-01-01

Reactive oxygen species (ROS) are important in the development of diabetic testicular dysfunction. Overproduction of ROS promotes the process of apoptosis, which shows that there is a crosstalk between oxidative stress and apoptosis. Recent research has suggested that NADPH oxidase is the main source of ROS. In this study, we investigated whether the NADPH oxidase inhibitor, apocynin, can improve diabetes‑induced testicular dysfunction by suppressing oxidative stress. The streptozocin (STZ)-induced diabetic rats were administered apocynin, and the mRNA and protein expression of Bax, Bcl-2, p47phox and p67phox was examined by real-time PCR (RT-PCR) and western blot analysis. Production of ROS was measured by thiobarbituric acid reactive substances (TBARS) assay. Terminal-deoxynucleoitidyl transferase mediated nick end-labeling (TUNEL) assay was used to detect apoptosis and ELISA was used to detect total testosterone levels. The mRNA and protein expression of Bcl-2 was significantly reduced, and that of Bax, p47phox and p67phox was significantly increased in the diabetic rats compared to the control group. Apocynin significantly increased the expression of Bcl-2 and decreased the expression of Bax, p47phox and p67phox at both the mRNA and protein levels. The production of ROS and apoptotic cells significantly increased in the diabetic group compared to the control group. Apocynin significantly reduced the production of ROS and apoptotic cells and increased the total testosterone level. In conclusion, apocynin is capable of ameliorating testicular dysfunction.

Oral Leucine Supplementation Is Sensed by the Brain but neither Reduces Food Intake nor Induces an Anorectic Pattern of Gene Expression in the Hypothalamus

PubMed Central

Zampieri, Thais T.; Pedroso, João A. B.; Furigo, Isadora C.; Tirapegui, Julio; Donato, Jose

2013-01-01

Leucine activates the intracellular mammalian target of the rapamycin (mTOR) pathway, and hypothalamic mTOR signaling regulates food intake. Although central infusion of leucine reduces food intake, it is still uncertain whether oral leucine supplementation is able to affect the hypothalamic circuits that control energy balance. We observed increased phosphorylation of p70s6k in the mouse hypothalamus after an acute oral gavage of leucine. We then assessed whether acute oral gavage of leucine induces the activation of neurons in several hypothalamic nuclei and in the brainstem. Leucine did not induce the expression of Fos in hypothalamic nuclei, but it increased the number of Fos-immunoreactive neurons in the area postrema. In addition, oral gavage of leucine acutely increased the 24 h food intake of mice. Nonetheless, chronic leucine supplementation in the drinking water did not change the food intake and the weight gain of ob/ob mice and of wild-type mice consuming a low- or a high-fat diet. We assessed the hypothalamic gene expression and observed that leucine supplementation increased the expression of enzymes (BCAT1, BCAT2 and BCKDK) that metabolize branched-chain amino acids. Despite these effects, leucine supplementation did not induce an anorectic pattern of gene expression in the hypothalamus. In conclusion, our data show that the brain is able to sense oral leucine intake. However, the food intake is not modified by chronic oral leucine supplementation. These results question the possible efficacy of leucine supplementation as an appetite suppressant to treat obesity. PMID:24349566

PPAR ligands improve impaired metabolic pathways in fetal hearts of diabetic rats.

PubMed

Kurtz, Melisa; Capobianco, Evangelina; Martinez, Nora; Roberti, Sabrina Lorena; Arany, Edith; Jawerbaum, Alicia

2014-10-01

In maternal diabetes, the fetal heart can be structurally and functionally affected. Maternal diets enriched in certain unsaturated fatty acids can activate the nuclear receptors peroxisome proliferator-activated receptors (PPARs) and regulate metabolic and anti-inflammatory pathways during development. Our aim was to investigate whether PPARα expression, lipid metabolism, lipoperoxidation, and nitric oxide (NO) production are altered in the fetal hearts of diabetic rats, and to analyze the putative effects of in vivo PPAR activation on these parameters. We found decreased PPARα expression in the hearts of male but not female fetuses of diabetic rats when compared with controls. Fetal treatments with the PPARα ligand leukotriene B4 upregulated the expression of PPARα and target genes involved in fatty acid oxidation in the fetal hearts. Increased concentrations of triglycerides, cholesterol, and phospholipids were found in the hearts of fetuses of diabetic rats. Maternal treatments with diets supplemented with 6% olive oil or 6% safflower oil, enriched in unsaturated fatty acids that can activate PPARs, led to few changes in lipid concentrations, but up-regulated PPARα expression in fetal hearts. NO production, which was increased in the hearts of male and female fetuses in the diabetic group, and lipoperoxidation, which was increased in the hearts of male fetuses in the diabetic group, was reduced by the maternal treatments supplemented with safflower oil. In conclusion, impaired PPARα expression, altered lipid metabolism, and increased oxidative and nitridergic pathways were evidenced in hearts of fetuses of diabetic rats and were regulated in a gender-dependent manner by treatments enriched with PPAR ligands. © 2014 Society for Endocrinology.

Cytochrome c oxidase is activated by the oncoprotein Ras and is required for A549 lung adenocarcinoma growth

PubMed Central

2012-01-01

Background Constitutive activation of Ras in immortalized bronchial epithelial cells increases electron transport chain activity, oxygen consumption and tricarboxylic acid cycling through unknown mechanisms. We hypothesized that members of the Ras family may stimulate respiration by enhancing the expression of the Vb regulatory subunit of cytochrome c oxidase (COX). Results We found that the introduction of activated H-RasV12 into immortalized human bronchial epithelial cells increased eIF4E-dependent COX Vb protein expression simultaneously with an increase in COX activity and oxygen consumption. In support of the regulation of COX Vb expression by the Ras family, we also found that selective siRNA-mediated inhibition of K-Ras expression in A549 lung adenocarcinoma cells reduced COX Vb protein expression, COX activity, oxygen consumption and the steady-state concentration of ATP. We postulated that COX Vb-mediated activation of COX activity may be required for the anchorage-independent growth of A549 cells as soft agar colonies or as lung xenografts. We transfected the A549 cells with COX Vb small interfering or shRNA and observed a significant reduction of their COX activity, oxygen consumption, ATP and ability to grow in soft agar and as poorly differentiated tumors in athymic mice. Conclusion Taken together, our findings indicate that the activation of Ras increases COX activity and mitochondrial respiration in part via up-regulation of COX Vb and that this regulatory subunit of COX may have utility as a Ras effector target for the development of anti-neoplastic agents. PMID:22917272

Attenuation of β-amyloid-induced tauopathy via activation of CK2α/SIRT1: targeting for cilostazol.

PubMed

Lee, Hye Rin; Shin, Hwa Kyoung; Park, So Youn; Kim, Hye Young; Lee, Won Suk; Rhim, Byung Yong; Hong, Ki Whan; Kim, Chi Dae

2014-02-01

β-Amyloid (Aβ) deposits and hyperphosphorylated tau aggregates are the chief hallmarks in the Alzheimer's disease (AD) brains, but the strategies for controlling these pathological events remain elusive. We hypothesized that CK2-coupled SIRT1 activation stimulated by cilostazol suppresses tau acetylation (Ac-tau) and tau phosphorylation (P-tau) by inhibiting activation of P300 and GSK3β. Aβ was endogenously overproduced in N2a cells expressing human APP Swedish mutation (N2aSwe) by exposure to medium containing 1% fetal bovine serum for 24 hr. Increased Aβ accumulation was accompanied by increased Ac-tau and P-tau levels. Concomitantly, these cells showed increased P300 and GSK3β P-Tyr216 expression; their expressions were significantly reduced by treatment with cilostazol (3-30 μM) and resveratrol (20 μM). Moreover, decreased expression of SIRT1 and its activity by Aβ were significantly reversed by cilostazol as by resveratrol. In addition, cilostazol strongly stimulated CK2α phosphorylation and its activity, and then stimulated SIRT1 phosphorylation. These effects were confirmed by using the pharmacological inhibitors KT5720 (1 μM, PKA inhibitor), TBCA (20 μM, inhibitor of CK2), and sirtinol (20 μM, SIRT1 inhibitor) as well as by SIRT1 gene silencing and overexpression techniques. In conclusion, increased cAMP-dependent protein kinase-linked CK2/SIRT1 expression by cilostazol can be a therapeutic strategy to suppress the tau-related neurodegeneration in the AD brain. Copyright © 2013 Wiley Periodicals, Inc.

Pharmacologic Suppression of Hepcidin Increases Macrophage Cholesterol Efflux and Reduces Foam Cell Formation and Atherosclerosis

PubMed Central

Saeed, Omar; Otsuka, Fumiyuki; Polavarapu, Rohini; Karmali, Vinit; Weiss, Daiana; Davis, Talina; Rostad, Brad; Pachura, Kimberly; Adams, Lila; Elliott, John; Taylor, W. Robert; Narula, Jagat; Kolodgie, Frank; Virmani, Renu; Hong, Charles C.; Finn, Aloke V.

2012-01-01

Objectives We recently reported that lowering of macrophage free intracellular iron increases expression of cholesterol efflux transporters ABCA1 and ABCG1 by reducing generation of reactive oxygen species. In this study, we explore whether reducing macrophage intracellular iron levels via pharmacologic suppression of hepcidin can increase macrophage-specific expression of cholesterol efflux transporters and reduce atherosclerosis. Methods and Results To suppress hepcidin, increase expression of the iron exporter ferroportin (FPN), and reduce macrophage intracellular iron, we used a small molecule inhibitor of BMP signaling, LDN 193189 (LDN). LDN (10 mg/kg i.p. bid) was administered to mice and its effects on atherosclerosis, intracellular iron, oxidative stress, lipid efflux, and foam cell formation were measured in plaques and peritoneal macrophages. Long-term LDN administration to Apo E (-/-) mice increased ABCA1 immunoreactivity within intraplaque macrophages by 3.7-fold (n=8; p=0.03), reduced oil-red-o positive lipid area by 50% (n=8; p=0.02) and decreased total plaque area by 43% (n=8; p=0.001). LDN suppressed liver hepcidin transcription and increased macrophage FPN, lowering intracellular iron and hydrogen peroxide production. LDN treatment increased macrophage ABCA1 and ABCG1 expression, significantly raised cholesterol efflux to ApoA-1 and decreased foam cell formation. All preceding LDN-induced effects on cholesterol efflux were reversed by exogenous hepcidin administration, suggesting that modulation of intracellular iron levels within macrophages as the mechanism by which LDN triggers these effects. Conclusion These data suggest that pharmacologic manipulation of iron homeostasis may be a promising target to increase macrophage reverse cholesterol transport and limit atherosclerosis. PMID:22095982

Liver X receptor agonists augment human islet function through activation of anaplerotic pathways and glycerolipid/free fatty acid cycling.

PubMed

Ogihara, Takeshi; Chuang, Jen-Chieh; Vestermark, George L; Garmey, James C; Ketchum, Robert J; Huang, Xiaolun; Brayman, Kenneth L; Thorner, Michael O; Repa, Joyce J; Mirmira, Raghavendra G; Evans-Molina, Carmella

2010-02-19

Recent studies in rodent models suggest that liver X receptors (LXRs) may play an important role in the maintenance of glucose homeostasis and islet function. To date, however, no studies have comprehensively examined the role of LXRs in human islet biology. Human islets were isolated from non-diabetic donors and incubated in the presence or absence of two synthetic LXR agonists, TO-901317 and GW3965, under conditions of low and high glucose. LXR agonist treatment enhanced both basal and stimulated insulin secretion, which corresponded to an increase in the expression of genes involved in anaplerosis and reverse cholesterol transport. Furthermore, enzyme activity of pyruvate carboxylase, a key regulator of pyruvate cycling and anaplerotic flux, was also increased. Whereas LXR agonist treatment up-regulated known downstream targets involved in lipogenesis, we observed no increase in the accumulation of intra-islet triglyceride at the dose of agonist used in our study. Moreover, LXR activation increased expression of the genes encoding hormone-sensitive lipase and adipose triglyceride lipase, two enzymes involved in lipolysis and glycerolipid/free fatty acid cycling. Chronically, insulin gene expression was increased after treatment with TO-901317, and this was accompanied by increased Pdx-1 nuclear protein levels and enhanced Pdx-1 binding to the insulin promoter. In conclusion, our data suggest that LXR agonists have a direct effect on the islet to augment insulin secretion and expression, actions that should be considered either as therapeutic or unintended side effects, as these agents are developed for clinical use.

[Knockdown of DNA-PKcs inhibits cell cycle and its mechanism of drug-resistant Bel7402/5-Fu hepatocellular carcinoma cells].

PubMed

Li, Dayu; Liu, Yun; Yu, Chunbo; Liu, Xiping; Fan, Fang

2017-12-01

Objective To study the effect of the knock-down of the DNA-dependent protein kinase catalytic subunit (DNA-PKcs) on the cell cycle of the multidrug-resistant (MDR) Bel7402/5-Fu hepatocellular carcinoma cells and its MDR mechanism. Methods After cationic liposome-mediated siDNA-PKcs oligonucleotide transfection, the drug sensitivity of Bel7402/5-Fu cells to 5-fluorouracil (5-Fu) and adriamycin (ADM) was determined by MTT assay; the cell cycle were detected by flow cytometry; meanwhile, the protein expressions of cell cycle-related proteins P21, cell cycle protein B1 (cyclin B1), cell cycle division protein 2 (CDC2) were tested by Western blotting; the expressions of ataxia telangiectasia mutated (ATM) and p53 at both mRNA and protein levels were detected by real-time PCR and Western blot analysis. Results The MTT results showed siDNA-PKcs increased the chemotherapeutic sensitivity of Bel7402/5-Fu cells to 5-Fu and ADM. The flow cytometric analysis showed siDNA-PKcs decreased the percentage of S-phase cells but increased the percentage of G2/M phase cells. Western blotting showed siDNA-PKcs increased the protein expression of P21 but decreased cyclinB1 and CDC2 proteins. In addition, siDNA-PKcs also increased the expressions of ATM and p53. Conclusion DNA-PKcs silencing increases P21 while decreases cyclin B1 and CDC2 expressions, and finally induces G2/M phase arrest in Bel7402/5-Fu cells, which may be related to ATM-p53 signaling pathway.

Electrical stimulation modulates Wnt signaling and regulates genes for the motor endplate and calcium binding in muscle of rats with spinal cord transection

PubMed Central

2013-01-01

Background Spinal cord injury (SCI) results in muscle atrophy and a shift of slow oxidative to fast glycolytic fibers. Electrical stimulation (ES) at least partially restores muscle mass and fiber type distribution. The objective of this study was to was to characterize the early molecular adaptations that occur in rat soleus muscle after initiating isometric resistance exercise by ES for one hour per day for 1, 3 or 7 days when ES was begun 16 weeks after SCI. Additionally, changes in mRNA levels after ES were compared with those induced in soleus at the same time points after gastrocnemius tenotomy (GA). Results ES increased expression of Hey1 and Pitx2 suggesting increased Notch and Wnt signaling, respectively, but did not normalize RCAN1.4, a measure of calcineurin/NFAT signaling, or PGC-1ß mRNA levels. ES increased PGC-1α expression but not that of slow myofibrillar genes. Microarray analysis showed that after ES, genes coding for calcium binding proteins and nicotinic acetylcholine receptors were increased, and the expression of genes involved in blood vessel formation and morphogenesis was altered. Of the 165 genes altered by ES only 16 were also differentially expressed after GA, of which 12 were altered in the same direction by ES and GA. In contrast to ES, GA induced expression of genes related to oxidative phosphorylation. Conclusions Notch and Wnt signaling may be involved in ES-induced increases in the mass of paralyzed muscle. Molecular adaptations of paralyzed soleus to resistance exercise are delayed or defective compared to normally innervated muscle. PMID:23914941

Exposure to welding fumes and lower airway infection with Streptococcus pneumoniae

PubMed Central

Suri, Reetika; Periselneris, Jimstan; Lanone, Sophie; Zeidler-Erdely, Patti C.; Melton, Geoffrey; Palmer, Keith T.; Andujar, Pascal; Antonini, James M.; Cohignac, Vanessa; Erdely, Aaron; Jose, Ricardo J.; Mudway, Ian; Brown, Jeremy; Grigg, Jonathan

2015-01-01

Background Welders are at increased risk of pneumococcal pneumonia. The mechanism for this association is not known. The capacity of pneumococci to adhere to and infect lower airway cells is mediated by host-expressed platelet-activating factor receptor (PAFR). Objective We sought to assess the effect of mild steel welding fumes (MS-WF) on PAFR-dependent pneumococcal adhesion and infection to human airway cells in vitro and on pneumococcal airway infection in a mouse model. Methods The oxidative potential of MS-WF was assessed by their capacity to reduce antioxidants in vitro. Pneumococcal adhesion and infection of A549, BEAS-2B, and primary human bronchial airway cells were assessed by means of quantitative bacterial culture and expressed as colony-forming units (CFU). After intranasal instillation of MS-WF, mice were infected with Streptococcus pneumoniae, and bronchoalveolar lavage fluid (BALF) and lung CFU values were determined. PAFR protein levels were assessed by using immunofluorescence and immunohistochemistry, and PAFR mRNA expression was assessed by using quantitative PCR. PAFR was blocked by CV-3988, and oxidative stress was attenuated by N-acetylcysteine. Results: MS-WF exhibited high oxidative potential. In A549 and BEAS-2B cells MS-WF increased pneumococcal adhesion and infection and PAFR protein expression. Both CV-3988 and N-acetylcysteine reduced MS-WF–stimulated pneumococcal adhesion and infection of airway cells. MS-WF increased mouse lung PAFR mRNA expression and increased BALF and lung pneumococcal CFU values. In MS-WF–exposed mice CV-3988 reduced BALF CFU values. Conclusions Hypersusceptibility of welders to pneumococcal pneumonia is in part mediated by the capacity of welding fumes to increase PAFR-dependent pneumococcal adhesion and infection of lower airway cells. PMID:26277596

The Chinese Herb Yi-Qi-Huo-Xue Protects Cardiomyocyte Function in Diabetic Cardiomyopathy.

PubMed

Wang, Xiangsheng; Huang, Jing; Wang, Shengyi; Ni, Qing

2018-01-01

Aims. To study the effect of the Chinese herb Yi-qi-huo-xue on cardiomyopathy in diabetic rats. Methods . Rats were fed a high fat and high glucose diet and injected with 50 ml/kg streptozotocin (STZ) to induce diabetic cardiomyopathy (DCM), followed by treatment with Yi-qi-huo-xue for 4 weeks. We measured the rats' heart weight index, observed the myocardial morphology using hematoxylin eosin (HE) staining, and determined the content of collagen types I and III in the myocardium using enzyme-linked immunosorbent assay (ELISA). We determined Bcl-2, Bax, and P53 protein expression by Western blot analysis and the cardiomyocyte apoptosis rate via a flow cytometry assay. Results. Compared with the rats in the control group, the diabetic rats gained weight and had increased blood sugar levels, an enhanced heart weight index, and increased myocardial pathophysiological damage. There was a decrease in their Bcl-2 expression, and their Bax and P53 expression increased. The Bcl-2/Bax ratio was enhanced, and there was an increase in the content of collagen types I and III in the myocardium. After treatment with Yi-qi-huo-xue, all levels listed above returned to normal. Conclusion. The Chinese herb Yi-qi-huo-xue degraded the myocardial interstitial collagen types I and III to protect the myocardium of the diabetic rats, thus delaying the role of myocardial fibrosis. Yi-qi-huo-xue could play an important role in protecting the myocardium of DCM rats by enhancing the expression of the Bcl-2 protein, inhibiting the expression of the Bax and P53 proteins, increasing the ratio of Bcl-2/Bax, and inhibiting the apoptosis of cardiomyocytes.

The farnesoid X receptor agonist obeticholic acid upregulates biliary excretion of asymmetric dimethylarginine via MATE-1 during hepatic ischemia/reperfusion injury

PubMed Central

Berardo, Clarissa; Siciliano, Veronica; Rizzo, Vittoria; Adorini, Luciano; Richelmi, Plinio

2018-01-01

Background We previously showed that increased asymmetric dimethylarginine (ADMA) biliary excretion occurs during hepatic ischemia/reperfusion (I/R), prompting us to study the effects of the farnesoid X receptor (FXR) agonist obeticholic acid (OCA) on bile, serum and tissue levels of ADMA after I/R. Material and methods Male Wistar rats were orally administered 10mg/kg/day of OCA or vehicle for 5 days and were subjected to 60 min partial hepatic ischemia or sham-operated. After a 60 min reperfusion, serum, tissue and bile ADMA levels, liver mRNA and protein expression of ADMA transporters (CAT-1, CAT-2A, CAT-2B, OCT-1, MATE-1), and enzymes involved in ADMA synthesis (protein-arginine-N-methyltransferase-1, PRMT-1) and metabolism (dimethylarginine-dimethylaminohydrolase-1, DDAH-1) were measured. Results OCA administration induced a further increase in biliary ADMA levels both in sham and I/R groups, with no significant changes in hepatic ADMA content. A reduction in CAT-1, CAT-2A or CAT-2B transcripts was found in OCA-treated sham-operated rats compared with vehicle. Conversely, OCA administration did not change CAT-1, CAT-2A or CAT-2B expression, already reduced by I/R. However, a marked decrease in OCT-1 and increase in MATE-1 expression was observed. A similar trend occurred with protein expression. Conclusion The reduced mRNA expression of hepatic CAT transporters suggests that the increase in serum ADMA levels is probably due to decreased liver uptake of ADMA from the systemic circulation. Conversely, the mechanism involved in further increasing biliary ADMA levels in sham and I/R groups treated with OCA appears to be MATE-1-dependent. PMID:29346429

Granulocyte and monocyte CD11b expression during plasma separation is dependent on complement factor 5 (C5) - an ex vivo study with blood from a C5-deficient individual.

PubMed

Hardersen, Randolf; Enebakk, Terje; Christiansen, Dorte; Bergseth, Grethe; Brekke, Ole-Lars; Mollnes, Tom Eirik; Lappegård, Knut Tore; Hovland, Anders

2018-04-01

The aim of the study was to investigate the role of complement factor 5 (C5) in reactions elicited by plasma separation using blood from a C5-deficient (C5D) individual, comparing it to C5-deficient blood reconstituted with C5 (C5DR) and blood from healthy donors. Blood was circulated through an ex vivo plasma separation model. Leukocyte CD11b expression and leukocyte-platelet conjugates were measured by flow cytometry during a 30-min period. Other markers were assessed during a 240-min period. Granulocyte and monocyte CD11b expression did not increase in C5D blood during plasma separation. In C5DR samples granulocytes CD11b expression, measured by mean fluorescence intensity (MFI), increased from 10481 ± 6022 (SD) to 62703 ± 4936, and monocytes CD11b expression changed from 13837 ± 7047 to 40063 ± 713. Granulocyte-platelet conjugates showed a 2.5-fold increase in the C5DR sample compared to the C5D sample. Monocyte-platelet conjugates increased independently of C5. In the C5D samples, platelet count decreased from 210 × 10 9 /L (201-219) (median and range) to 51 × 10 9 /L (50-51), and C3bc increased from 14 CAU/mL (21-7) to 198 CAU/mL (127-269), whereas TCC formation was blocked during plasma separation. In conclusion, up-regulation of granulocyte and monocyte CD11b during plasma separation was C5-dependent. The results also indicate C5 dependency in granulocyte-platelet conjugates formation. © 2018 APMIS. Published by John Wiley & Sons Ltd.

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

PubMed Central

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

2009-01-01

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

Interactions between IGF-I, estrogen receptor-α (ERα), and ERβ in regulating growth/apoptosis of MCF-7 human breast cancer cells.

PubMed

Mendoza, Rhone A; Enriquez, Marlene I; Mejia, Sylvia M; Moody, Emily E; Thordarson, Gudmundur

2011-01-01

Understanding of the interactions between estradiol (E₂) and IGF-I is still incomplete. Cell lines derived from the MCF-7 breast cancer cells were generated with suppressed expression of the IGF-I receptor (IGF-IR), termed IGF-IR.low cells, by stable transfection using small interfering RNA (siRNA) expression vector. Vector for control cells carried sequence generating noninterfering RNA. Concomitant with reduction in the IGF-IR levels, the IGF-IR.low cells also showed a reduction in estrogen receptor α (ERα) and progesterone receptor expressions, and an elevation in the expression of ERβ. The number of the IGF-IR.low cells was reduced in response to IGF-I and human GH plus epidermal growth factor, but E₂ did not cause an increase in the number of the IGF-IR.low cells compared to controls. The proliferation rate of IGF-IR.low cells was only reduced in response to E₂ compared to controls, whereas their basal and hormone-stimulated apoptosis rate was increased. Phosphorylation of p38 mitogen-activated protein kinase (p38 MAPK) was increased in the IGF-IR.low cells after treatment with E₂, without affecting control cells. Furthermore, phosphorylation of the tumor suppressor protein p53 was elevated in the IGF-IR.low cells compared to the controls. In conclusion, suppressing IGF-IR expression decreased the level of ERα but increased the level of ERβ. Overall growth rate of the IGF-IR.low cells was reduced mostly through an increase in apoptosis without affecting proliferation substantially. We hypothesize that a decreased ERα:ERβ ratio triggered a rapid phosphorylation of p38 MAPK, which in turn phosphorylated the p53 tumor suppressor and accelerated apoptosis rate.

In silico analysis of stomach lineage specific gene set expression pattern in gastric cancer

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

Pandi, Narayanan Sathiya, E-mail: sathiyapandi@gmail.com; Suganya, Sivagurunathan; Rajendran, Suriliyandi

Highlights: •Identified stomach lineage specific gene set (SLSGS) was found to be under expressed in gastric tumors. •Elevated expression of SLSGS in gastric tumor is a molecular predictor of metabolic type gastric cancer. •In silico pathway scanning identified estrogen-α signaling is a putative regulator of SLSGS in gastric cancer. •Elevated expression of SLSGS in GC is associated with an overall increase in the survival of GC patients. -- Abstract: Stomach lineage specific gene products act as a protective barrier in the normal stomach and their expression maintains the normal physiological processes, cellular integrity and morphology of the gastric wall. However,more » the regulation of stomach lineage specific genes in gastric cancer (GC) is far less clear. In the present study, we sought to investigate the role and regulation of stomach lineage specific gene set (SLSGS) in GC. SLSGS was identified by comparing the mRNA expression profiles of normal stomach tissue with other organ tissue. The obtained SLSGS was found to be under expressed in gastric tumors. Functional annotation analysis revealed that the SLSGS was enriched for digestive function and gastric epithelial maintenance. Employing a single sample prediction method across GC mRNA expression profiles identified the under expression of SLSGS in proliferative type and invasive type gastric tumors compared to the metabolic type gastric tumors. Integrative pathway activation prediction analysis revealed a close association between estrogen-α signaling and SLSGS expression pattern in GC. Elevated expression of SLSGS in GC is associated with an overall increase in the survival of GC patients. In conclusion, our results highlight that estrogen mediated regulation of SLSGS in gastric tumor is a molecular predictor of metabolic type GC and prognostic factor in GC.« less

JC Virus Mediates Invasion and Migration in Colorectal Metastasis

PubMed Central

Link, Alexander; Shin, Sung Kwan; Nagasaka, Takeshi; Balaguer, Francesc; Koi, Minoru; Jung, Barbara; Boland, C. Richard; Goel, Ajay

2009-01-01

Introduction JC Virus (JCV), a human polyomavirus, is frequently present in colorectal cancers (CRCs). JCV large T-Ag (T-Ag) expressed in approximately half of all CRC's, however, its functional role in CRC is poorly understood. We hypothesized that JCV T-Ag may mediate metastasis in CRC cells through increased migration and invasion. Material and Methods CRC cell lines (HCT116 and SW837) were stably transfected with JCV early transcript sequences cloned into pCR3 or empty vectors. Migration and invasion assays were performed using Boyden chambers. Global gene expression analysis was performed to identify genetic targets and pathways altered by T-Ag expression. Microarray results were validated by qRT-PCR, protein expression analyses and immunohistochemistry. Matching primary CRCs and liver metastases from 33 patients were analyzed for T-Ag expression by immunohistochemistry. Results T-Ag expressing cell lines showed 2 to 3-fold increase in migration and invasion compared to controls. JCV T-Ag expression resulted in differential expression of several genetic targets, including genes that mediate cell migration and invasion. Pathway analysis suggested a significant involvement of these genes with AKT and MAPK signaling. Treatment with selective PI3K/AKT and MAPK pathway inhibitors resulted in reduced migration and invasion. In support of our in-vitro results, immunohistochemical staining of the advanced stage tumors revealed frequent JCV T-Ag expression in metastatic primary tumors (92%) as well as in their matching liver metastasis (73%). Conclusion These data suggest that JCV T-Ag expression in CRC associates with a metastatic phenotype, which may partly be mediated through the AKT/MAPK signaling pathway. Frequent expression of JCV T-Ag in CRC liver metastasis provides further clues supporting a mechanistic role for JCV as a possible mediator of cellular motility and invasion in CRC. PMID:19997600

The daily timing of gene expression and physiology in mammals

PubMed Central

Schibler, Ueli

2007-01-01

Mammalian behavior and physiology undergo daily rhythms that are coordinated by an endogenous circadian timing system. This system has a hierarchical structure, in that a master pacemaker, residing in the suprachiasmatic nucleus of the ventral hypothalamus, synchronizes peripheral oscillators in virtually all body cells. While the basic molecular mechanisms generating the daily rhythms are similar in aIl cells, most clock out-puts are cell-specific. This conclusion is based on genomewide transcriptome profiling studies in several tissues that have revealed hundreds of rhythmically expressed genes. Cyclic gene expression in the various organs governs overt rhythms in behavior and physiology, encompassing sleep-wake cycles, metabolism, xenobiotic detoxification, and cellularproliferation. As a consequence, chronic perturbation of this temporal organization may lead to increased morbidity and reduced lifespan. PMID:17969863

Cubilin expression is monoallelic and epigenetically augmented via PPARs

PubMed Central

2013-01-01

Background Cubilin is an endocytic receptor that is necessary for renal and intestinal absorption of a range of ligands. Endocytosis mediated by cubilin and its co-receptor megalin is the principal mechanism for proximal tubule reabsorption of proteins from the glomerular filtrate. Cubilin is also required for intestinal endocytosis of intrinsic factor-vitamin B12 complex. Despite its importance, little is known about the regulation of cubilin expression. Results Here we show that cubilin expression is under epigenetic regulation by at least two processes. The first process involves inactivation of expression of one of the cubilin alleles. This monoallelic expression state could not be transformed to biallelic by inhibiting DNA methylation or histone deacetylation. The second process involves transcriptional regulation of cubilin by peroxisome proliferator-activated receptor (PPAR) transcription factors that are themselves regulated by DNA methylation and histone deacetylation. This is supported by findings that inhibitors of DNA methylation and histone deacetylation, 5Aza and TSA, increase cubilin mRNA and protein in renal and intestinal cell lines. Not only was the expression of PPARα and γ inducible by 5Aza and TSA, but the positive effects of TSA and 5Aza on cubilin expression were also dependent on both increased PPAR transcription and activation. Additionally, 5Aza and TSA had similar effects on the expression of the cubilin co-receptor, megalin. Conclusions Together, these findings reveal that cubilin and megalin mRNA expression is under epigenetic control and thus point to new avenues for overcoming pathological suppression of these genes through targeting of epigenetic regulatory processes. PMID:23773363

Corruption of the Fas Pathway Delays the Pulmonary Clearance of Murine Osteosarcoma Cells, Enhances Their Metastatic Potential, and Reduces the Effect of Aerosol Gemcitabine

PubMed Central

Gordon, Nancy; Koshkina, Nadezhda V.; Jia, Shu-Fang; Khanna, Chand; Mendoza, Arnulfo; Worth, Laura L.; Kleinerman, Eugenie S.

2015-01-01

Purpose Pulmonary metastases continue to be a significant problem in osteosarcoma. Apoptosis dysfunction is known to influence tumor development. Fas (CD95, APO-1)/FasL is one of the most extensively studied apoptotic pathways. Because FasL is constitutively expressed in the lung, cells that express Fas should be eliminated by lung endothelium. Cells with low or no cell surface Fas expression may be able to evade this innate defense mechanism. The purpose of these studies was to evaluate Fas expression in osteosarcoma lung metastases and the effect of gemcitabine on Fas expression and tumor growth. Experimental Design and Results Using the K7M2 murine osteosarcoma model, Fas expression was quantified using immunohistochemistry. High levels of Fas were present in primary tumors, but no Fas expression was present in actively growing lung metastases. Blocking the Fas pathway using Fas-associated death domain dominant-negative delayed tumor cell clearance from the lung and increased metastatic potential. Treatment of mice with aerosol gemcitabine resulted in increased Fas expression and subsequent tum or regression. Conclusions We conclude that corruption of the Fas pathway is critical to the ability of osteosarcoma cells to grow in the lung. Agents such as gemcitabine that up-regulate cell surface Fas expression may therefore be effective in treating osteosarcoma lung metastases. These data also suggest that an additional mechanism by which gemcitabine induces regression of osteosarcoma lung metastases is mediated by enhancing the sensitivity of the tumor cells to the constitutive FasL in the lung. PMID:17671136

Associations between expression levels of nucleotide excision repair proteins in lymphoblastoid cells and risk of squamous cell carcinoma of the head and neck.

PubMed

Han, Peng; Liu, Hongliang; Shi, Qiong; Liu, Zhensheng; Troy, Jesse D; Lee, Walter T; Zevallos, Jose P; Li, Guojun; Sturgis, Erich M; Wei, Qingyi

2018-06-01

Squamous cell carcinoma of head and neck (SCCHN) is one of the most common malignancies worldwide, and nucleotide excision repair (NER) is involved in SCCHN susceptibility. In this analysis of 349 newly diagnosed SCCHN patients and 295 cancer-free controls, we investigated whether expression levels of eight core NER proteins were associated with risk of SCCHN. We quantified NER protein expression levels in cultured peripheral lymphocytes using a reverse-phase protein microarray. Compared with the controls, SCCHN patients had statistically significantly lower expression levels of ERCC3 and XPA (P = 0.001 and 0.001, respectively). After dividing the subjects by controls' median values of expression levels, we found a dose-dependent association between an increased risk of SCCHN and low expression levels of ERCC3 (adjusted OR, 1.75, and 95% CI: 1.26-2.42; P trend  = 0.008) and XPA (adjusted OR, 1.88; 95% CI, 1.35-2.60; P trend  = 0.001). We also identified a significant multiplicative interaction between smoking status and ERCC3 expression levels (P = 0.014). Finally, after integrating demographic and clinical variables, we found that the addition of ERCC3 and XPA expression levels to the model significantly improved the sensitivity of the expanded model on SCCHN risk. In conclusion, reduced protein expression levels of ERCC3 and XPA were associated with an increased risk of SCCHN. However, these results need to be confirmed in additional large studies. © 2018 Wiley Periodicals, Inc.

Cadmium-induced ethylene production and responses in Arabidopsis thaliana rely on ACS2 and ACS6 gene expression

PubMed Central

2014-01-01

Background Anthropogenic activities cause metal pollution worldwide. Plants can absorb and accumulate these metals through their root system, inducing stress as a result of excess metal concentrations inside the plant. Ethylene is a regulator of multiple plant processes, and is affected by many biotic and abiotic stresses. Increased ethylene levels have been observed after exposure to excess metals but it remains unclear how the increased ethylene levels are achieved at the molecular level. In this study, the effects of cadmium (Cd) exposure on the production of ethylene and its precursor 1-aminocyclopropane-1-carboxylic acid (ACC), and on the expression of the ACC Synthase (ACS) and ACC Oxidase (ACO) multigene families were investigated in Arabidopsis thaliana. Results Increased ethylene release after Cd exposure was directly measurable in a system using rockwool-cultivated plants; enhanced levels of the ethylene precursor ACC together with higher mRNA levels of ethylene responsive genes: ACO2, ETR2 and ERF1 also indicated increased ethylene production in hydroponic culture. Regarding underlying mechanisms, it was found that the transcript levels of ACO2 and ACO4, the most abundantly expressed members of the ACO multigene family, were increased upon Cd exposure. ACC synthesis is the rate-limiting step in ethylene biosynthesis, and transcript levels of both ACS2 and ACS6 showed the highest increase and became the most abundant isoforms after Cd exposure, suggesting their importance in the Cd-induced increase of ethylene production. Conclusions Cadmium induced the biosynthesis of ACC and ethylene in Arabidopsis thaliana plants mainly via the increased expression of ACS2 and ACS6. This was confirmed in the acs2-1acs6-1 double knockout mutants, which showed a decreased ethylene production, positively affecting leaf biomass and resulting in a delayed induction of ethylene responsive gene expressions without significant differences in Cd contents between wild-type and mutant plants. PMID:25082369

Intoxication- and withdrawal-dependent expression of central and peripheral cytokines following initial ethanol exposure

PubMed Central

Doremus-Fitzwater, Tamara L.; Buck, Hollin M.; Bordner, Kelly A.; Richey, Laura; Jones, Megan E.; Deak, Terrence

2016-01-01

Background Evidence has emerged demonstrating that ethanol influences cytokine expression within the CNS, although most studies have examined long-term exposure. Thus, the cytokine response to an acute ethanol challenge was investigated, in order to characterize profiles of cytokine changes following acute exposure. Methods Rats pups were injected intraperitoneally (i.p.) with 2-g/kg ethanol and IL-1 mRNA and protein assessed 0, 60, 120, 180, and 240 min post-injection (Exp. 1). In Exps. 2-5, the expression of several cytokines was examined in adult male rats during acute intoxication (3 hr after 4-g/kg ethanol), as well as withdrawal (18 hr post-injection), after i.p. and intragastric (i.g.) ethanol administration. Results Early in ontogeny, acute ethanol significantly decreased brain IL-1 mRNA and protein. Subsequently, when adult rats were examined, significant and temporally dynamic alterations in central and peripheral cytokines were observed following acute i.p. ethanol exposure (4-g/kg). Although cytokine- and region-dependent, central IL-6 expression was generally increased and TNFα decreased during intoxication, whereas IL-1 expression exhibited increases during withdrawal. In the periphery, acute i.p. ethanol elevated expression of all cytokines, with the response growing in magnitude as the time post-injection increased. Following acute i.g. ethanol (4-g/kg), intoxication-related increases in IL-6 expression were again observed in the PVN, although to a lesser extent. Long-term, voluntary, intermittent ethanol consumption resulted in tolerance to the effects of an i.g. ethanol challenge (4-g/kg) on PVN IL-6 expression, whereas these same elevations in IL-6 expression were still seen in the amygdala in rats with a history of moderate ethanol intake. Treatment with minocycline did not significantly attenuate i.p. or i.g. ethanol-induced changes in central cytokine expression. Conclusions Together, these studies provide a foundation for understanding fluctuations in central and peripheral cytokines following acute ethanol as potential contributors to the constellation of neural and behavioral alterations observed during ethanol intoxication and withdrawal. PMID:25156612

Post-Translational Regulation of Polycystin-2 Protein Expression as a Novel Mechanism of Cholangiocyte Reaction and Repair from Biliary Damage

PubMed Central

Spirli, Carlo; Villani, Ambra; Mariotti, Valeria; Fabris, Luca; Fiorotto, Romina; Strazzabosco, Mario

2015-01-01

Polycystin-2 (PC2 /TRPP2), a member of the transient receptor potential channels (TRP) family, is a non-selective calcium channel. Mutations in PC2/TRPP2 are associated with Polycystic Liver Diseases. PC2-defective cholangiocytes shows increased production of cAMP, PKA-dependent activation of the ERK1/2 pathway, HIF1α-mediated VEGF production, and stimulation of cyst growth and progression. Activation of the ERK/HIF1α/VEGF pathway in cholangiocytes plays a key role during repair from biliary damage. We hypothesized that PC2 levels are modulated during biliary damage/repair, resulting in activation of the ERK/HIF1α/VEGF pathway. Results PC2 protein expression, but not its gene expression, was significantly reduced in mouse livers with biliary damage (Mdr2−/−-KO, bile duct ligation, DDC-treatment). Treatment of colangiocytes with pro-inflammatory cytokines, nitric oxide (NO) donors and ER stressors), increased ERK1/2 phosphorylation, HIF1α transcriptional activity, secretion of VEGF, VEGFR2 phosphorylation and downregulated PC2 protein expression without affecting PC2 gene expression. Expression of Herp and NEK, ubiquitin-like proteins that promote proteosomal PC2 degradation was increased. Pre-treatment with the proteasome inhibitor MG-132 restored the expression of PC2 in cells treated with cytokines but not in cells treated with NO donors or with ER stressors. In these conditions, PC2 degradation was instead inhibited by interfering with the autophagy pathway. Treatment of DDC-mice and of Mdr2−/−-mice with the proteasome inhibitor bortezomib, restored PC2 expression and significantly reduced the ductular reaction, fibrosis and p-ERK1/2. In conclusion, in response to biliary damage, PC2 expression is modulated post-translationally by the proteasome or the autophagy pathways. PC2-dowregulation is associated with activation of ERK1/2 and increase of HIF1α-mediated VEGF secretion. Treatments able to restore PC2 expression and to reduce ductular reaction and fibrosis may represent a new therapeutic approach in biliary diseases. PMID:26313562

Prominin-2 expression increases protrusions, decreases caveolae and inhibits Cdc42 dependent fluid phase endocytosis

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

Singh, Raman Deep, E-mail: Takhter.Ramandeep@mayo.edu; Schroeder, Andreas S.; Scheffer, Luana

Highlights: •Prominin-2 expression induced protrusions that co-localized with lipid raft markers. •Prominin-2 expression decreased caveolae, caveolar endocytosis and increased pCav1. •Prominin-2 expression inhibited fluid phase endocytosis by inactivation of Cdc42. •These endocytic effects can be reversed by adding exogenous cholesterol. •Caveolin1 knockdown restored fluid phase endocytosis in Prominin2 expressing cells. -- Abstract: Background: Membrane protrusions play important roles in biological processes such as cell adhesion, wound healing, migration, and sensing of the external environment. Cell protrusions are a subtype of membrane microdomains composed of cholesterol and sphingolipids, and can be disrupted by cholesterol depletion. Prominins are pentaspan membrane proteins thatmore » bind cholesterol and localize to plasma membrane (PM) protrusions. Prominin-1 is of great interest as a marker for stem and cancer cells, while Prominin-2 (Prom2) is reportedly restricted to epithelial cells. Aim: To characterize the effects of Prom-2 expression on PM microdomain organization. Methods: Prom2-fluorescent protein was transfected in human skin fibroblasts (HSF) and Chinese hamster ovary (CHO) cells for PM raft and endocytic studies. Caveolae at PM were visualized using transmission electron microscopy. Cdc42 activation was measured and caveolin-1 knockdown was performed using siRNAs. Results: Prom2 expression in HSF and CHO cells caused extensive Prom2-positive protrusions that co-localized with lipid raft markers. Prom2 expression significantly decreased caveolae at the PM, reduced caveolar endocytosis and increased caveolin-1 phosphorylation. Prom2 expression also inhibited Cdc42-dependent fluid phase endocytosis via decreased Cdc42 activation. Effects on endocytosis were reversed by addition of cholesterol. Knockdown of caveolin-1 by siRNA restored Cdc42 dependent fluid phase endocytosis in Prom2-expressing cells. Conclusions: Prom2 protrusions primarily localize to lipid rafts and recruit cholesterol into protrusions and away from caveolae, leading to increased phosphorylation of caveolin-1, which inhibits Cdc42-dependent endocytosis. This study provides a new insight for the role for prominins in the regulation of PM lipid organization.« less

Piper betle L. Modulates Senescence-Associated Genes Expression in Replicative Senescent Human Diploid Fibroblasts

PubMed Central

Durani, Lina Wati; Tan, Jen Kit; Chua, Kien Hui

2017-01-01

Piper betle (PB) is a traditional medicine that is widely used to treat different diseases around Asian region. The leaf extracts contain various bioactive compounds, which were reported to have antidiabetic, antibacterial, anti-inflammatory, antioxidant, and anticancer effects. In this study, the effect of PB aqueous extracts on replicative senescent human diploid fibroblasts (HDFs) was investigated by determining the expressions of senescence-associated genes using quantitative PCR. Our results showed that PB extracts at 0.4 mg/ml can improve cell proliferation of young (143%), presenescent (127.3%), and senescent (157.3%) HDFs. Increased expressions of PRDX6, TP53, CDKN2A, PAK2, and MAPK14 were observed in senescent HDFs compared to young and/or presenescent HDFs. Treatment with PB extracts modulates the transcriptional profile changes in senescent HDFs. By contrast, expressions of SOD1 increased, whereas GPX1, PRDX6, TP53, CDKN2A, PAK2, and MAPK14 were decreased in PB-treated senescent HDFs compared to untreated senescent HDFs. In conclusion, this study indicates the modulation of PB extracts on senescence-associated genes expression of replicative senescent HDFs. Further studies warrant determining the mechanism of PB in modulating replicative senescence of HDFs through these signaling pathways. PMID:28596968

Piper betle L. Modulates Senescence-Associated Genes Expression in Replicative Senescent Human Diploid Fibroblasts.

PubMed

Durani, Lina Wati; Khor, Shy Cian; Tan, Jen Kit; Chua, Kien Hui; Mohd Yusof, Yasmin Anum; Makpol, Suzana

2017-01-01

Piper betle (PB) is a traditional medicine that is widely used to treat different diseases around Asian region. The leaf extracts contain various bioactive compounds, which were reported to have antidiabetic, antibacterial, anti-inflammatory, antioxidant, and anticancer effects. In this study, the effect of PB aqueous extracts on replicative senescent human diploid fibroblasts (HDFs) was investigated by determining the expressions of senescence-associated genes using quantitative PCR. Our results showed that PB extracts at 0.4 mg/ml can improve cell proliferation of young (143%), presenescent (127.3%), and senescent (157.3%) HDFs. Increased expressions of PRDX6 , TP53 , CDKN2A , PAK2 , and MAPK14 were observed in senescent HDFs compared to young and/or presenescent HDFs. Treatment with PB extracts modulates the transcriptional profile changes in senescent HDFs. By contrast, expressions of SOD1 increased, whereas GPX1 , PRDX6 , TP53 , CDKN2A , PAK2 , and MAPK14 were decreased in PB-treated senescent HDFs compared to untreated senescent HDFs. In conclusion, this study indicates the modulation of PB extracts on senescence-associated genes expression of replicative senescent HDFs. Further studies warrant determining the mechanism of PB in modulating replicative senescence of HDFs through these signaling pathways.

Shear stress reduces protease activated receptor-1 expression in human endothelial cells

NASA Technical Reports Server (NTRS)

Nguyen, K. T.; Eskin, S. G.; Patterson, C.; Runge, M. S.; McIntire, L. V.

2001-01-01

Shear stress has been shown to regulate several genes involved in the thrombotic and proliferative functions of endothelial cells. Thrombin receptor (protease-activated receptor-1: PAR-1) increases at sites of vascular injury, which suggests an important role for PAR-1 in vascular diseases. However, the effect of shear stress on PAR-1 expression has not been previously studied. This work investigates effects of shear stress on PAR-1 gene expression in both human umbilical vein endothelial cells (HUVECs) and microvascular endothelial cells (HMECs). Cells were exposed to different shear stresses using a parallel plate flow system. Northern blot and flow cytometry analysis showed that shear stress down-regulated PAR-1 messenger RNA (mRNA) and protein levels in both HUVECs and HMECs but with different thresholds. Furthermore, shear-reduced PAR-1 mRNA was due to a decrease of transcription rate, not increased mRNA degradation. Postshear stress release of endothelin-1 in response to thrombin was reduced in HUVECs and HMECs. Moreover, inhibitors of potential signaling pathways applied during shear stress indicated mediation of the shear-decreased PAR-1 expression by protein kinases. In conclusion, shear stress exposure reduces PAR-1 gene expression in HMECs and HUVECs through a mechanism dependent in part on protein kinases, leading to altered endothelial cell functional responses to thrombin.

4PBA strongly attenuates endoplasmic reticulum stress, fibrosis, and mitochondrial apoptosis markers in cyclosporine treated human gingival fibroblasts.

PubMed

Ranga Rao, Suresh; Subbarayan, Rajasekaran; Ajitkumar, Supraja; Murugan Girija, Dinesh

2018-01-01

Cyclosporine induces overgrowth of human gingiva. Previously we have shown (i) cyclosporine-inducing ER stress in human gingival fibroblasts (HGF), (ii) increased matrix protein expression, and (iii) interference with mitochondrial pro- and anti-apoptotic factors. This study was undertaken to assess the effects of melatonin (an antioxidant), 4PBA (an ER stress inhibitor), and simvastatin on the expression of ER Stress markers as well as on matrix and mitochondrial markers. HGF incubated with cyclosporine, or without melatonin/4PBA/statin. After 24 hr of incubation, mRNA expression of ER stress markers (GRP78, CHOP, XBP1, and XBPs) and matrix protein markers (like α-SMA, VEGF, TGF-β, CTGF), and mitochondrial apoptosis markers estimated and compared with housekeeping gene GAPDH. Compared to the control cyclosporine significantly augmented ER Stress and matrix proteins, which decreased significantly with the use of melatonin, 4PBA, and simvastatin. The mitochondrial proapoptotic molecule cyclophilin D, as well as Bcl2 expression also decreased after PBA treatment, paralleling an increase in cytochrome c expression. The effect of 4PBA was much more pronounced than the influence of other two. In conclusion, 4PBA could be a viable therapeutic option for drug-induced gingival overgrowth. © 2017 Wiley Periodicals, Inc.

Clinical characteristics of inflammation-associated depression: Monocyte gene expression is age-related in major depressive disorder.

PubMed

Grosse, Laura; Carvalho, Livia A; Wijkhuijs, Annemarie J M; Bellingrath, Silja; Ruland, Tillmann; Ambrée, Oliver; Alferink, Judith; Ehring, Thomas; Drexhage, Hemmo A; Arolt, Volker

2015-02-01

Increased inflammatory activation might only be present in a subgroup of depressed individuals in which immune processes are especially relevant to disease development. We aimed to analyze demographic, depression, and trauma characteristics of major depressive disorder (MDD) patients with regard to inflammatory monocyte gene expression. Fifty-six naturalistically treated MDD patients (32 ± 12 years) and 57 healthy controls (HC; 31 ± 11 years) were analyzed by the Inventory of Depressive Symptomatology (IDS) and by the Childhood Trauma Questionnaire (CTQ). We determined the expression of 38 inflammatory and immune activation genes including the glucocorticoid receptor (GR)α and GRβ genes in purified CD14(+) monocytes using quantitative-polymerase chain reaction (RT-qPCR). Monocyte gene expression was age-dependent, particularly in MDD patients. Increased monocyte gene expression and decreased GRα/β ratio were only present in MDD patients aged ⩾ 28 years. Post hoc analyses of monocyte immune activation in patients <28 years showed two subgroups: a subgroup with a severe course of depression (recurrent type, onset <15 years) - additionally characterized by panic/arousal symptoms and childhood trauma - that had a monocyte gene expression similar to HC, and a second subgroup with a milder course of the disorder (73% first episode depression, onset ⩾15 years) - additionally characterized by the absence of panic symptoms - that exhibited a strongly reduced inflammatory monocyte activation compared to HC. In conclusion, monocyte immune activation was not uniformly raised in MDD patients but was increased only in patients of 28 years and older. Copyright © 2014 Elsevier Inc. All rights reserved.

Valproic Acid Increases Expression of Neuronal Stem/Progenitor Cell in Spinal Cord Injury

PubMed Central

Bang, Woo-Seok; Cho, Dae-Chul; Kim, Hye-Jeong; Sung, Joo-Kyung

2013-01-01

Objective This study investigates the effect of valproic acid (VPA) on expression of neural stem/progenitor cells (NSPCs) in a rat spinal cord injury (SCI) model. Methods Adult male rats (n=24) were randomly and blindly allocated into three groups. Laminectomy at T9 was performed in all three groups. In group 1 (sham), only laminectomy was performed. In group 2 (SCI-VPA), the animals received a dose of 200 mg/kg of VPA. In group 3 (SCI-saline), animals received 1.0 mL of the saline vehicle solution. A modified aneurysm clip with a closing force of 30 grams was applied extradurally around the spinal cord at T9, and then rapidly released with cord compression persisting for 2 minutes. The rats were sacrificed and the spinal cord were collected one week after SCI. Immunohistochemistry (IHC) and western blotting sample were obtained from 5 mm rostral region to the lesion and prepared. We analyzed the nestin immunoreactivity from the white matter of ventral cord and the ependyma of central canal. Nestin and SOX2 were used for markers for NSPCs and analyzed by IHC and western blotting, respectively. Results Nestin and SOX2 were expressed significantly in the SCI groups but not in the sham group. Comparing SCI groups, nestin and SOX2 expression were much stronger in SCI-VPA group than in SCI-saline group. Conclusion Nestin and SOX2 as markers for NSPCs showed increased expression in SCI-VPA group in comparison with SCI-saline group. This result suggests VPA increases expression of spinal NSPCs in SCI. PMID:24044073

Ischemic postconditioning provides cardioprotective and antiapoptotic effects against ischemia-reperfusion injury through iNOS inhibition in hyperthyroid rats.

PubMed

Zaman, Jalal; Jeddi, Sajad; Daneshpour, Maryam Sadat; Zarkesh, Maryam; Daneshian, Zahra; Ghasemi, Asghar

2015-10-10

Ischemic postconditioning (IPost) is a strategy to provide protection against ischemia-reperfusion (IR) injury. The cardioprotective effects of IPost in cases of ischemic heart disease along with co-morbidities like hyperthyroidism remain unknown. The aim of this study was to investigate the effects of IPost on expression of eNOS, iNOS, Bax, and Bcl-2 genes in hyperthyroid male rats, subjected to myocardial IR. Hyperthyroidism was induced by adding thyroxine to drinking water for a period of 21 days. Using the Langendorff device hearts were perfused, then subjected to a 30-minute global ischemia which was followed by 120 min of reperfusion; subsequently IPost was induced immediately after ischemia. Results indicated that following IR, expression of eNOS and Bcl-2 decreased, whereas expression of iNOS and Bax increased in both the control and hyperthyroid groups. In hyperthyroid animals, IPost significantly increased expression of eNOS by 3.19 fold and Bcl-2 by 3.66 fold; it also decreased expression of Bax by 51%, and reduced IR-induced DNA laddering pattern and infarct size (45.7 ± 1.82% vs. 59.3 ± 1.83%, p<0.05) in the presence of aminoguanidine (AG), a selective iNOS inhibitor. In conclusion, IPost per se could not provide cardioprotection against myocardial ischemia in hyperthyroid rats, a loss of which however was restored by the combination of IPost and iNOS inhibition that acts by a decrease in Bax and an increase in both eNOS and Bcl-2 expression. Copyright © 2015 Elsevier B.V. All rights reserved.

Immunolocalization of osteopontin in dysplasias and squamous cell carcinomas arising from oral epithelium

PubMed Central

Aravind, Thara; Janardhanan, Mahija; Rakesh, S; Savithri, Vindhya; Unnikrishnan, U G

2017-01-01

Background: Early detection of oral squamous cell carcinoma (OSCC) remains one of the most efficient ways to ensure patient survival and improved quality of life. Although specific biomarkers related to OSCC have been investigated, a useful biomarker that assesses the transition potential of potentially malignant lesion to OSCC remains to be found. Osteopontin (OPN) has been recognized as an important factor in tumorigenesis and their expression in OSCC have been investigated earlier. In the present study, evaluation of OPN expression in premalignant and malignant lesions has been carried out to assess their possible role as a biomarker in the early diagnosis and prognosis of OSCC. Objectives: The objective of this study is to evaluate the role of OPN as a biomarker in the diagnosis and prognosis of OSCC. Materials and Methods: The study group consisted of archival paraffin-embedded blocks of ten cases each of varying grades of OSCC, oral epithelial dysplasias and epithelial hyperplasias. Sections were subjected to immunohistochemical staining for the biomarker OPN. Results: A positive OPN expression was noticed in epithelial dysplasias and SCC arising from the oral epithelium. A progressive increase in the intensity of staining was seen with increasing grades of dysplasias and a decrease in OPN expression with an increase in grades was observed in OSCC. Conclusion: The expression of OPN in full thickness of epithelium in severe dysplasias, carcinoma in situ, and in the superficial epithelium of OSCC suggest the possibility of considering OPN expression in full epithelial thickness in dysplasias as an indicator for malignant transformation. PMID:28479681

Low-Intensity Extracorporeal Shock Wave Therapy Enhances Brain-Derived Neurotrophic Factor Expression through PERK/ATF4 Signaling Pathway.

PubMed

Wang, Bohan; Ning, Hongxiu; Reed-Maldonado, Amanda B; Zhou, Jun; Ruan, Yajun; Zhou, Tie; Wang, Hsun Shuan; Oh, Byung Seok; Banie, Lia; Lin, Guiting; Lue, Tom F

2017-02-16

Low-intensity extracorporeal shock wave therapy (Li-ESWT) is used in the treatment of erectile dysfunction, but its mechanisms are not well understood. Previously, we found that Li-ESWT increased the expression of brain-derived neurotrophic factor (BDNF). Here we assessed the underlying signaling pathways in Schwann cells in vitro and in penis tissue in vivo after nerve injury. The result indicated that BDNF were significantly increased by the Li-ESWT after nerve injury, as well as the expression of BDNF in Schwann cells (SCs, RT4-D6P2T) in vitro. Li-ESWT activated the protein kinase RNA-like endoplasmic reticulum (ER) kinase (PERK) pathway by increasing the phosphorylation levels of PERK and eukaryotic initiation factor 2a (eIF2α), and enhanced activating transcription factor 4 (ATF4) in an energy-dependent manner. In addition, GSK2656157-an inhibitor of PERK-effectively inhibited the effect of Li-ESWT on the phosphorylation of PERK, eIF2α, and the expression of ATF4. Furthermore, silencing ATF4 dramatically attenuated the effect of Li-ESWT on the expression of BDNF, but had no effect on hypoxia-inducible factor (HIF)1α or glial cell-derived neurotrophic factor (GDNF) in Schwann cells. In conclusion, our findings shed new light on the underlying mechanisms by which Li-ESWT may stimulate the expression of BDNF through activation of PERK/ATF4 signaling pathway. This information may help to refine the use of Li-ESWT to further improve its clinical efficacy.

Low-Intensity Extracorporeal Shock Wave Therapy Enhances Brain-Derived Neurotrophic Factor Expression through PERK/ATF4 Signaling Pathway

PubMed Central

Wang, Bohan; Ning, Hongxiu; Reed-Maldonado, Amanda B.; Zhou, Jun; Ruan, Yajun; Zhou, Tie; Wang, Hsun Shuan; Oh, Byung Seok; Banie, Lia; Lin, Guiting; Lue, Tom F.

2017-01-01

Low-intensity extracorporeal shock wave therapy (Li-ESWT) is used in the treatment of erectile dysfunction, but its mechanisms are not well understood. Previously, we found that Li-ESWT increased the expression of brain-derived neurotrophic factor (BDNF). Here we assessed the underlying signaling pathways in Schwann cells in vitro and in penis tissue in vivo after nerve injury. The result indicated that BDNF were significantly increased by the Li-ESWT after nerve injury, as well as the expression of BDNF in Schwann cells (SCs, RT4-D6P2T) in vitro. Li-ESWT activated the protein kinase RNA-like endoplasmic reticulum (ER) kinase (PERK) pathway by increasing the phosphorylation levels of PERK and eukaryotic initiation factor 2a (eIF2α), and enhanced activating transcription factor 4 (ATF4) in an energy-dependent manner. In addition, GSK2656157—an inhibitor of PERK—effectively inhibited the effect of Li-ESWT on the phosphorylation of PERK, eIF2α, and the expression of ATF4. Furthermore, silencing ATF4 dramatically attenuated the effect of Li-ESWT on the expression of BDNF, but had no effect on hypoxia-inducible factor (HIF)1α or glial cell-derived neurotrophic factor (GDNF) in Schwann cells. In conclusion, our findings shed new light on the underlying mechanisms by which Li-ESWT may stimulate the expression of BDNF through activation of PERK/ATF4 signaling pathway. This information may help to refine the use of Li-ESWT to further improve its clinical efficacy. PMID:28212323

Aquaporin Expression and Water Transport Pathways inside Leaves Are Affected by Nitrogen Supply through Transpiration in Rice Plants

PubMed Central

Ding, Lei; Li, Yingrui; Gao, Limin; Lu, Zhifeng; Wang, Min; Ling, Ning; Shen, Qirong; Guo, Shiwei

2018-01-01

The photosynthetic rate increases under high-N supply, resulting in a large CO2 transport conductance in mesophyll cells. It is less known that water movement is affected by nitrogen supply in leaves. This study investigated whether the expression of aquaporin and water transport were affected by low-N (0.7 mM) and high-N (7 mM) concentrations in the hydroponic culture of four rice varieties: (1) Shanyou 63 (SY63), a hybrid variant of the indica species; (2) Yangdao 6 (YD6), a variant of indica species; (3) Zhendao 11 (ZD11), a hybrid variant of japonica species; and (4) Jiuyou 418 (JY418), another hybrid of the japonica species. Both the photosynthetic and transpiration rate were increased by the high-N supply in the four varieties. The expressions of aquaporins, plasma membrane intrinsic proteins (PIPs), and tonoplast membrane intrinsic protein (TIP) were higher in high-N than low-N leaves, except in SY63. Leaf hydraulic conductance (Kleaf) was lower in high-N than low-N leaves in SY63, while Kleaf increased under high-N supply in the YD6 variant. Negative correlations were observed between the expression of aquaporin and the transpiration rate in different varieties. Moreover, there was a significant negative correlation between transpiration rate and intercellular air space. In conclusion, the change in expression of aquaporins could affect Kleaf and transpiration. A feedback effect of transpiration would regulate aquaporin expression. The present results imply a coordination of gas exchange with leaf hydraulic conductance. PMID:29337869

Leupaxin acts as a mediator in prostate carcinoma progression through deregulation of p120catenin expression.

PubMed

Kaulfuss, S; von Hardenberg, S; Schweyer, S; Herr, A M; Laccone, F; Wolf, S; Burfeind, P

2009-11-12

Recently, we could show that the focal adhesion protein leupaxin (LPXN) is expressed in human prostate carcinomas (PCa) and induces invasiveness of androgen-independent PCa cells. In this study we show that LPXN enhanced the progression of existing PCa in vivo by breeding transgenic mice with prostate-specific LPXN expression and TRAMP mice (transgenic adenocarcinoma of mouse prostate). Double transgenic LPXN/TRAMP mice showed a significant increase in poorly differentiated PCa and distant metastases as compared with control TRAMP mice. Additional studies on primary PCa cells generated from both transgenic backgrounds confirmed the connection regarding LPXN overexpression and increased motility and invasiveness of PCa cells. One mediator of LPXN-induced invasion was found to be the cell-cell adhesion protein p120catenin (p120CTN). Both in vitro and in vivo experiments revealed that p120CTN expression negatively correlates with LPXN expression, followed by a redistribution of beta-catenin. Downregulation of LPXN using small interfering RNAs (siRNAs) resulted in a membranous localization of beta-catenin, whereas strong nuclear accumulation of beta-catenin was observed in p120CTN knockdown cells leading to enhanced transcription of the beta-catenin target gene matrix metalloprotease-7. In conclusion, the present results indicate that LPXN enhances the progression of PCa through downregulation of p120CTN expression and that LPXN could function as a marker for aggressive PCa in the future.

IL-8 as mediator in the microenvironment-leukaemia network in acute myeloid leukaemia.

PubMed

Kuett, Alexander; Rieger, Christina; Perathoner, Deborah; Herold, Tobias; Wagner, Michaela; Sironi, Silvia; Sotlar, Karl; Horny, Hans-Peter; Deniffel, Christian; Drolle, Heidrun; Fiegl, Michael

2015-12-17

The bone marrow microenvironment is physiologically hypoxic with areas being as low as 1% O2, e.g. the stem cell niche. Acute myeloid leukaemia (AML) blasts misuse these bone marrow niches for protection by the local microenvironment, but also might create their own microenvironment. Here we identify IL-8 as a hypoxia-regulated cytokine in both AML cell lines and primary AML samples that is induced within 48 hours of severe hypoxia (1% O2). IL-8 lacked effects on AML cells but induced migration in mesenchymal stromal cells (MSC), an integral part of the bone marrow. Accordingly, MSC were significantly increased in AML bone marrow as compared to healthy bone marrow. Interestingly, mononuclear cells obtained from healthy bone marrow displayed both significantly lower endogenous and hypoxia-induced production of IL-8. IL-8 mRNA expression in AML blasts from 533 patients differed between genetic subgroups with significantly lower expression of IL-8 in acute promyelocytic leukaemia (APL), while in non APL-AML patients with FLT ITD had the highest IL-8 expression. In this subgroup, high IL-8 expression was also prognostically unfavourable. In conclusion, hypoxia as encountered in the bone marrow specifically increases IL-8 expression of AML, which in turn impacts niche formation. High IL-8 expression might be correlated with poor prognosis in certain AML subsets.

Inverse Relationship of the CMKLR1 Relative Expression and Chemerin Serum Levels in Obesity with Dysmetabolic Phenotype and Insulin Resistance

PubMed Central

Corona-Meraz, Fernanda-Isadora; Navarro-Hernández, Rosa-Elena; Ruíz-Quezada, Sandra-Luz; Madrigal-Ruíz, Perla-Monserrat; Castro-Albarrán, Jorge; Chavarría-Ávila, Efraín; Guzmán-Ornelas, Milton-Omar; Gómez-Bañuelos, Eduardo; Petri, Marcelo-Herón; Ramírez-Cedano, Joel-Isidro; Aguilar-Aldrete, María-Elena; Ríos-Ibarra, Clara; Vázquez-Del Mercado, Mónica

2016-01-01

Background. In obesity there is a subclinical chronic low-grade inflammatory response where insulin resistance (IR) may develop. Chemerin is secreted in white adipose tissue and promotes low-grade inflammatory process, where it expressed CMKLR1 receptor. The role of chemerin and CMKLR1 in inflammatory process secondary to obesity is not defined yet. Methods. Cross-sectional study with 134 individuals classified as with and without obesity by body mass index (BMI) and IR. Body fat storage measurements and metabolic and inflammatory markers were measured by routine methods. Soluble chemerin and basal levels of insulin by ELISA and relative expression of CMKLR1 were evaluated with qPCR and 2−ΔΔCT method. Results. Differences (P < 0.05) were observed between obesity and lean individuals in body fat storage measurements and metabolic-inflammatory markers. Both CMKLR1 expression and chemerin levels were increased in obesity without IR. Soluble chemerin levels correlate with adiposity and metabolic markers (r = 8.8% to 38.5%), P < 0.05. Conclusion. The increment of CMKLR1 expression was associated with insulin production. Increased serum levels of chemerin in obesity were observed, favoring a dysmetabolic response. The results observed in this study suggest that both chemerin and CMKLR1 have opposite expression in the context of low-grade inflammatory response manifested in the development of IR. PMID:27239101

The Homeodomain Transcription Factor Cdx1 Does Not Behave as an Oncogene in Normal Mouse Intestine1

PubMed Central

Crissey, Mary Ann S; Guo, Rong-Jun; Fogt, Franz; Li, Hong; Katz, Jonathan P; Silberg, Debra G; Suh, Eun Ran; Lynch, John P

2008-01-01

The Caudal-related homeobox genes Cdx1 and Cdx2 are intestine-specific transcription factors that regulate differentiation of intestinal cell types. Previously, we have shown Cdx1 to be antiproliferative and to promote cell differentiation. However, other studies have suggested that Cdx1 may be an oncogene. To test for oncogenic behavior, we used the murine villin promoter to ectopically express Cdx1 in the small intestinal villi and colonic surface epithelium. No changes in intestinal architecture, cell differentiation, or lineage selection were observed with expression of the transgene. Classic oncogenes enhance proliferation and induce tumors when ectopically expressed. However, the Cdx1 transgene neither altered intestinal proliferation nor induced spontaneous intestinal tumors. In a murine model for colitis-associated cancer, the Cdx1 transgene decreased, rather than increased, the number of adenomas that developed. In the polyps, the expression of the endogenous and the transgenic Cdx1 proteins was largely absent, whereas endogenous Villin expression was retained. This suggests that transgene silencing was specific and not due to a general Villin inactivation. In conclusion, neither the ectopic expression of Cdx1 was associated with changes in intestinal cell proliferation or differentiation nor was there increased intestinal cancer susceptibility. Our results therefore suggest that Cdx1 is not an oncogene in normal intestinal epithelium. PMID:18231635

Altered Osteocyte-Specific Protein Expression in Bone after Childhood Solid Organ Transplantation

PubMed Central

Pereira, Renata C.; Valta, Helena; Tumber, Navdeep; Salusky, Isidro B.; Jalanko, Hannu

2015-01-01

Background Bone fragility is common post solid organ transplantation but little is known about bone pathology on a tissue level. Abnormal osteocytic protein expression has been linked to compromised bone health in chronic kidney disease (CKD) and immunosuppressant medications may impact osteocyte function. Methods Transiliac bone biopsies were obtained from 22 pediatric solid organ allograft recipients (average age 15.6 years) an average of 6.3 ± 1.2 years after transplantation and from 12 pediatric pre-dialysis CKD patients (average age 13.2 years). Histomorphometry and immunohistochemistry for FGF23, DMP1, sclerostin, and osteopontin were performed on all biopsies. Results FGF23 and sclerostin were increased in transplant recipients relative to non-transplant CKD, regardless of the type of allograft received and despite, in the case of liver and heart recipients, a higher GFR. Bone DMP1 expression was higher in liver or heart than in kidney recipients, concomitant with higher serum phosphate values. Osteopontin expression was higher in CKD than in transplant recipients (p<0.01). Bone FGF23 and sclerostin correlated directly (r = 0.38, p<0.05); bone FGF23 expression and osteoid thickness correlated inversely (r = - 0.46, p<0.01). Conclusions Solid-organ transplantation is associated with increased FGF23 and sclerostin expression. The contribution of these findings to compromised bone health post transplantation warrants further evaluation. PMID:26390291

Luteinizing hormone/chorionic gonadotrophin receptor overexpressed in granulosa cells from polycystic ovary syndrome ovaries is functionally active.

PubMed

Kanamarlapudi, Venkateswarlu; Gordon, Uma D; López Bernal, Andrés

2016-06-01

Polycystic ovarian syndrome (PCOS) is associated with anovulatory infertility. Luteinizing hormone/chorionic gonadotrophin receptor (LHCGR), which is critical for ovulation, has been suggested to be expressed prematurely in the ovarian follicles of women with PCOS. This study aimed to analyse the expression and activity of LHCGR in ovarian granulosa cells from PCOS patients and the involvement of ARF6 small GTPase in LHCGR internalization. Granulosa cells (GC) isolated from follicular fluid collected during oocyte retrieval from normal women (n = 19) and women with PCOS (n = 17) were used to study differences in LHCGR protein expression and activity between normal and PCOS patients. LHCGR expression is up-regulated in GC from PCOS women. LHCGR in PCOS GC is functionally active, as shown by increased cAMP production upon human gonadotrophin (HCG)-stimulation. Moreover, ARF6 is highly expressed in GC from PCOS patients and HCG-stimulation increases the concentrations of active ARF6. The inhibition of ARF6 activation attenuates HCG-induced LHCGR internalization in both normal and PCOS GC, indicating that there are no alterations in LHCGR internalisation in GC from PCOS. In conclusion, the expression and activation of LHCGR and ARF6 are up-regulated in GC from PCOS women but the mechanism of agonist-induced LHCGR internalization is unaltered. Copyright © 2016 Reproductive Healthcare Ltd. Published by Elsevier Ltd. All rights reserved.

Hybrid proline-rich proteins: novel players in plant cell elongation?

PubMed Central

Dvořáková, Lenka; Srba, Miroslav; Opatrny, Zdenek; Fischer, Lukas

2012-01-01

Background and Aims Hybrid proline-rich proteins (HyPRPs) represent a large family of putative cell-wall proteins characterized by the presence of a variable N-terminal domain and a conserved C-terminal domain that is related to non-specific lipid transfer proteins. The function of HyPRPs remains unclear, but their widespread occurrence and abundant expression patterns indicate that they may be involved in a basic cellular process. Methods To elucidate the cellular function of HyPRPs, we modulated the expression of three HyPRP genes in tobacco (Nicotiana tabacum) BY-2 cell lines and in potato (Solanum tuberosum) plants. Key Results In BY-2 lines, over-expression of the three HyPRP genes with different types of N-terminal domains resulted in similar phenotypic changes, namely increased cell elongation, both in suspension culture and on solid media where the over-expression resulted in enhanced calli size. The over-expressing cells showed increased plasmolysis in a hypertonic mannitol solution and accelerated rate of protoplast release, suggesting loosening of the cell walls. In contrast to BY-2 lines, no phenotypic changes were observed in potato plants over-expressing the same or analogous HyPRP genes, presumably due to more complex compensatory mechanisms in planta. Conclusions Based on the results from BY-2 lines, we propose that HyPRPs, more specifically their C-terminal domains, represent a novel group of proteins involved in cell expansion. PMID:22028464

Impact of STAT/SOCS mRNA Expression Levels after Major Injury

PubMed Central

Brumann, M.; Matz, M.; Kusmenkov, T.; Stegmaier, J.; Biberthaler, P.; Kanz, K.-G.; Mutschler, W.; Bogner, V.

2014-01-01

Background. Fulminant changes in cytokine receptor signalling might provoke severe pathological alterations after multiple trauma. The aim of this study was to evaluate the posttraumatic imbalance of the innate immune system with a special focus on the STAT/SOCS family. Methods. 20 polytraumatized patients were included. Blood samples were drawn 0 h–72 h after trauma; mRNA expression profiles of IL-10, STAT 3, SOCS 1, and SOCS 3 were quantified by qPCR. Results. IL-10 mRNA expression increased significantly in the early posttraumatic period. STAT 3 mRNA expressions showed a significant maximum at 6 h after trauma. SOCS 1 levels significantly decreased 6 h–72 h after trauma. SOCS 3 levels were significantly higher in nonsurvivors 6 h after trauma. Conclusion. We present a serial, sequential investigation in human neutrophil granulocytes of major trauma patients evaluating mRNA expression profiles of IL-10, STAT 3, SOCS 1, and SOCS 3. Posttraumatically, immune disorder was accompanied by a significant increase of IL-10 and STAT 3 mRNA expression, whereas SOCS 1 mRNA levels decreased after injury. We could demonstrate that death after trauma was associated with higher SOCS 3 mRNA levels already at 6 h after trauma. To support our results, further investigations have to evaluate protein levels of STAT/SOCS family in terms of posttraumatic immune imbalance. PMID:24648661

Nestin is highly expressed in fetal spinal cord isolated from placenta previa patients and promotes inflammation by enhancing NF-κB activity.

PubMed

Li, Ling; Zhang, Jing; Gao, Huahe; Ma, Yuyan

2018-04-26

Purpose Nestin is expressed in various tissues of the embryo in patients with placenta previa, while the regulatory mechanism still unknown. Materials and methods All participants terminated pregnancy. Among them, 75 patients with placenta previa were assigned to the case group and 80 healthy pregnant women with normal placenta were assigned to the control group. Expression of nestin and CDK5 in fetal spinal cord tissues was detected by Western Blot and RT-qPCR methods. The enzyme-linked immunosorbent assay (ELISA) was used to determine the serum expression of some pro-inflammatory cytokines in placenta previa patients. The interaction between nestin and CDK5 was evaluated by immunoprecipitation and siRNA inhibition of nestin was performed to estimate its effect on NF-κB activity in fetal spinal cord tissues. Results Along with increased expression of nestin and CDK5 in fetal spinal cord tissues in the case group, IL-1β, IL-6, TNF-α, and IFN-γ were increased in the serum of placenta previa patients. siRNA inhibition analysis indicated that nestin interacted with CDK5 and regulated NF-κB activity in fetal spinal cord tissues. Conclusions Nestin is highly expressed and the interaction between nestin and CDK5 might lead to the progress of placenta previa through its regulation on NF-κB.

Role of emmprin in endometrial cancer

PubMed Central

2012-01-01

Background Extracellular matrix metalloproteinase inducer (Emmprin/CD147) is a transmembrane glycoprotein that belongs to the immunoglobulin superfamily. Enriched on the surface of many tumor cells, emmprin promotes tumor growth, invasion, metastasis and angiogenesis. We evaluated the clinical importance of emmprin and investigated its role in endometrial cancer. Methods Emmprin expression was examined in uterine normal endometrium, endometrial hyperplasia and cancer specimens by immunohistochemistry. In addition, the biological functions and inhibitory effects of an emmprin knockdown were investigated in HEC-50B and KLE endometrial cancer cell lines. Results The levels of emmprin expression were significantly increased in the endometrial cancer specimens compared with the normal endometrium and endometrial hyperplasia specimens (p < 0.05). The disease-free survival (DFS) and overall survival (OS) rates of patients with high emmprin expression were significantly higher than those of patients with low emmprin expression (DFS: p < 0.001; OS: p < 0.001). Emmprin knockdown by the siRNA led to cell proliferation, migration and invasion through TGF-β, EGF, NF-κB, VEGF, MMP-2, and MMP-9 expression, which in turn resulted in increased levels of E-cadherin and reduced levels of Vimentin and Snail in endometrial cancer. Conclusions The present findings suggest that low emmprin expression might be a predictor of favorable prognosis in endometrial cancer patients, and that emmprin may represent a potential therapeutic target for endometrial cancer. PMID:22640183

Glucagon Like Peptide-1 Receptor Expression in the Human Thyroid Gland

PubMed Central

Gier, Belinda; Butler, Peter C.; Lai, Chi K.; Kirakossian, David; DeNicola, Matthew M.

2012-01-01

Background: Glucagon like peptide-1 (GLP-1) mimetic therapy induces medullary thyroid neoplasia in rodents. We sought to establish whether C cells in human medullary thyroid carcinoma, C cell hyperplasia, and normal human thyroid express the GLP-1 receptor. Methods: Thyroid tissue samples with medullary thyroid carcinoma (n = 12), C cell hyperplasia (n = 9), papillary thyroid carcinoma (n = 17), and normal human thyroid (n = 15) were evaluated by immunofluorescence for expression of calcitonin and GLP-1 receptors. Results: Coincident immunoreactivity for calcitonin and GLP-1 receptor was consistently observed in both medullary thyroid carcinoma and C cell hyperplasia. GLP-1 receptor immunoreactivity was also detected in 18% of papillary thyroid carcinoma (three of 17 cases). Within normal human thyroid tissue, GLP-1 receptor immunoreactivity was found in five of 15 of the examined cases in about 35% of the total C cells assessed. Conclusions: In humans, neoplastic and hyperplastic lesions of thyroid C cells express the GLP-1 receptor. GLP-1 receptor expression is detected in 18% papillary thyroid carcinomas and in C cells in 33% of control thyroid lobes. The consequence of long-term pharmacologically increased GLP-1 signaling on these GLP-1 receptor-expressing cells in the thyroid gland in humans remains unknown, but appropriately powered prospective studies to exclude an increase in medullary or papillary carcinomas of the thyroid are warranted. PMID:22031513

Selectively enhanced expression of prophenoloxidase activating enzyme 1 (PPAE1) at a bacteria clearance site in the white shrimp, Litopenaeus vannamei

PubMed Central

2011-01-01

Background The prophenoloxidase-activating (PO activating) system plays an important role in the crustacean innate immunity, particularly in wound healing and pathogen defense. A key member of this system is prophenoloxidase-activating enzyme (PPAE), which is the direct activator of prophenoloxidase (proPO). Despite their importance in crustacean PO activating system, the studies on them remain limited. Results Here we report on a PPAE of white shrimp, Litopenaeus vannamei (lvPPAE1), which showed 94% similarity to PPAE1 of Penaeus monodon. We found that lvPPAE1 in fluid hemocytes was down regulated after challenge by Vibrio harveyi but was enhanced when shrimps were exposed to a bacteria-rich environment for long-term. In vivo gene silence of lvPPAE1 by RNAi can significantly reduce the phenoloxidase activity (PO) and increase the susceptibility of shrimps to V. harveyi. Although lvPPAE1 was down-regulated in fluid hemocytes by Vibrio challenge, its expression increased significantly in gill after bacteria injection, which is the primary bacteria-clearance tissue. Conclusion Suppressed expression in fluid hemocytes and enhanced expression in gill indicates selectively enhanced expression at the bacterial clearance site. This is a novel feature for PPAE expression. The results will contribute to our understanding of the PO activating system in crustaceans. PMID:22208405

Effects of metoprolol therapy on cardiac gap junction remodelling and conduction in human chronic atrial fibrillation

PubMed Central

Dhein, S; Rothe, S; Busch, A; Rojas Gomez, DM; Boldt, A; Reutemann, A; Seidel, T; Salameh, A; Pfannmüller, B; Rastan, A; Kostelka, M; Mohr, FW

2011-01-01

BACKGROUND AND PURPOSE We investigated the influence of metoprolol on gap junction proteins connexin43 (Cx43) and connexin40 (Cx40) in atrial tissue from patients with/without atrial fibrillation (AF). EXPERIMENTAL APPROACH Left atrial tissue samples from 160 patients with AF or sinus rhythm (SR) with or without metoprolol (mean daily dose: 65.2 ± 9.1 mg·day−1) were analysed for Cx43 and Cx40 by Western blot and immunohistology. Transverse and longitudinal conduction velocities were determined by 64 multi-electrode mapping. KEY RESULTS Both Cx43 and Cx40 expression were significantly increased in patients with AF versus SR. Cx43-expression in AF was significantly higher in patients receiving metoprolol, while Cx40 expression was unaffected by metoprolol treatment. In AF, the ratio of lateral/polar expression of Cx43 and Cx40 was enhanced due to increased expression at the sides of the cells (lateral) and a loss at the cell poles. This AF-induced increase in lateral/polar expression of Cx43, but not of Cx40, was significantly antagonized by metoprolol treatment. Functionally, in AF patients, transverse conduction velocity in atrial samples was significantly enhanced and this change was also significantly antagonized by metoprolol. CONCLUSIONS AND IMPLICATIONS AF induced enhanced lateral expression of Cx43 and Cx40 together with enhanced transverse conduction velocity in left atrial tissue. Alterations in localization of Cx43 and conduction changes were both antagonized by metoprolol, showing that pharmacological modulation of gap junction remodelling seems, in principle, possible. This finding may open new approaches to the development of anti-arrythmic drugs. PMID:21542828

Increased Expression of Cathelicidin by Direct Activation of Protease-Activated Receptor 2: Possible Implications on the Pathogenesis of Rosacea

PubMed Central

Kim, Ji Young; Kim, Yoon Jee; Lim, Beom Jin; Sohn, Hyo Jung; Shin, Dongyun

2014-01-01

Purpose Recent findings of increased cathelicidin protein and its proteolytic fragments in rosacea suggest a pathogenic role for cathelicidin in this disease. The relationship between cathelicidin and protease-activated receptor 2 (PAR-2) is therefore of interest, as PAR-2, expressed principally in keratinocytes, regulates pro-inflammatory cytokine expression in the skin. The purpose of this study was to determine the relationship between expression of PAR-2 and cathelicidin in rosacea and to test the effect of direct PAR-2 activation on cathelicidin expression in keratinocytes. Materials and Methods Samples from 40 patients with clinicopathologic diagnosis of rosacea and facial skin tissue samples from 20 patients with no specific findings or milium without inflammation were retrieved. Intensities of immunohistochemical staining for PAR-2 and cathelicidin were compared between normal and rosacea-affected skin tissues. Additionally, correlations between PAR-2 and cathelicidin staining intensities within rosacea patients were analyzed. In cultured keratinocytes, changes in PAR-2, cathelicidin, and vascular endothelial growth factor (VEGF) mRNA and protein were analyzed after treatment with PAR-2 activating peptide (AP). Results Cathelicidin expression was significantly higher in rosacea skin tissues than in normal tissues (p<0.001), while PAR-2 expression was not significantly higher in rosacea tissues than in normal skin tissues. A positive correlation between PAR-2 and cathelicidin within rosacea samples was observed (R=0.330, p=0.037). After treatment of PAR-2 AP, both mRNA and protein levels for PAR-2, cathelicidin, and VEGF significantly increased in cultured keratinocytes, compared with PAR-2 control peptide treatment. Conclusion PAR-2 may participate in the pathogenesis of rosacea through activation of cathelicidin LL-37, a mediator of innate immune responses in the skin. PMID:25323904

Changes in the renin angiotensin system during the development of colorectal cancer liver metastases

PubMed Central

2010-01-01

Background Blockade of the renin angiotensin system (RAS) via angiotensin I converting enzyme (ACE) inhibition reduces growth of colorectal cancer (CRC) liver metastases in a mouse model. In this work we defined the expression of the various components of the RAS in both tumor and liver during the progression of this disease. Methods Immunohistochemistry and quantitative RT-PCR was used to examine RAS expression in a mouse CRC liver metastases model. CRC metastases and liver tissue was assessed separately at key stages of CRC liver metastases development in untreated (control) mice and in mice treated with the ACE inhibitor captopril (750 mg/kg/day). Non-tumor induced (sham) mice indicated the effect of tumors on normal liver RAS. The statistical significance of multiple comparisons was determined using one-way analysis of variance followed by Bonferroni adjustment with SAS/STAT software. Results Reduced volume of CRC liver metastases with captopril treatment was evident. Local RAS of CRC metastases differed from the surrounding liver, with lower angiotensin II type 1 receptor (AT1R) expression but increased ANG-(1-7) receptor (MasR) compared to the liver. The AT1R localised to cancer and stromal infiltrating cells, while other RAS receptors were detected in cancer cells only. Tumor induction led to an initial increase in AT1R and ACE expression while captopril treatment significantly increased ACE expression in the final stages of tumor growth. Conversely, captopril treatment decreased expression of AT1R and angiotensinogen. Conclusions These results demonstrate significant changes in RAS expression in the tumor-bearing captopril treated liver and in CRC metastases. The data suggests the existence of a tumor-specific RAS that can be independently targeted by RAS blockade. PMID:20380732

Anti-Obesity and Anti-Diabetic Effects of Acacia Polyphenol in Obese Diabetic KKAy Mice Fed High-Fat Diet

PubMed Central

Ikarashi, Nobutomo; Toda, Takahiro; Okaniwa, Takehiro; Ito, Kiyomi; Ochiai, Wataru; Sugiyama, Kiyoshi

2011-01-01

Acacia polyphenol (AP) extracted from the bark of the black wattle tree (Acacia meansii) is rich in unique catechin-like flavan-3-ols, such as robinetinidol and fisetinidol. The present study investigated the anti-obesity/anti-diabetic effects of AP using obese diabetic KKAy mice. KKAy mice received either normal diet, high-fat diet or high-fat diet with additional AP for 7 weeks. After the end of administration, body weight, plasma glucose and insulin were measured. Furthermore, mRNA and protein expression of obesity/diabetic suppression-related genes were measured in skeletal muscle, liver and white adipose tissue. As a result, compared to the high-fat diet group, increases in body weight, plasma glucose and insulin were significantly suppressed for AP groups. Furthermore, compared to the high-fat diet group, mRNA expression of energy expenditure-related genes (PPARα, PPARδ, CPT1, ACO and UCP3) was significantly higher for AP groups in skeletal muscle. Protein expressions of CPT1, ACO and UCP3 for AP groups were also significantly higher when compared to the high-fat diet group. Moreover, AP lowered the expression of fat acid synthesis-related genes (SREBP-1c, ACC and FAS) in the liver. AP also increased mRNA expression of adiponectin and decreased expression of TNF-α in white adipose tissue. In conclusion, the anti-obesity actions of AP are considered attributable to increased expression of energy expenditure-related genes in skeletal muscle, and decreased fatty acid synthesis and fat intake in the liver. These results suggest that AP is expected to be a useful plant extract for alleviating metabolic syndrome. PMID:21799697

Maternal nutrient deprivation induces sex-specific changes in thyroid hormone receptor and deiodinase expression in the fetal guinea pig brain

PubMed Central

Chan, Shiao Y; Andrews, Marcus H; Lingas, Rania; McCabe, Chris J; Franklyn, Jayne A; Kilby, Mark D; Matthews, Stephen G

2005-01-01

Thyroid hormone deprivation during fetal life has been implicated in neurodevelopmental morbidity. In humans, poor growth in utero is also associated with fetal hypothyroxinaemia. In guinea pigs, a short period (48 h) of maternal nutrient deprivation at gestational day (gd) 50 results in fetuses with hypothyroxinaemia and increased brain/body weight ratios. Thyroid hormone action is mediated by nuclear thyroid hormone receptors (TRs) and is dependent upon the prereceptor regulation of supply of triiodothyronine (T3) by deiodinase enzymes. Examination of fetal guinea pig brains using in situ hybridization demonstrated widespread expression of mRNAs encoding TRα1, α2 and β1, with regional colocalization of deiodinase type 2 (D2) mRNA in the developing forebrain, limbic structures, brainstem and cerebellum at gd52. With maternal nutrient deprivation, TRα1 and β1 mRNA expression was significantly increased in the male, but decreased in the female fetal hippocampus and cerebellum and other areas showing high TR expression under euthyroid conditions. Maternal nutrient deprivation resulted in elevated D2 mRNA expression in males and females. Deiodinase type 3 (D3) mRNA expression was confined to the shell of the nucleus accumbens, the posterior amygdalohippocampal area, brainstem and cerebellum, and did not change with maternal nutrient deprivation. In conclusion, maternal nutrient deprivation resulted in sex-specific changes in TR mRNA expression and a generalized increase in D2 mRNAs within the fetal brain. These changes may represent a protective mechanism to maintain appropriate thyroid hormone action in the face of fetal hypothyroxinaemia in order to optimize brain development. PMID:15878952

Variation in branchial expression among insulin-like growth-factor binding proteins (igfbps) during Atlantic salmon smoltification and seawater exposure

USGS Publications Warehouse

Breves, Jason P.; Fujimoto, Chelsea K.; Phipps-Costin, Silas K.; Einarsdottir, Ingibjörg E.; Björnsson, Björn Thrandur; McCormick, Stephen

2017-01-01

BackgroundIn preparation for migration from freshwater to marine habitats, Atlantic salmon (Salmo salar L.) undergo smoltification, a transformation that includes the acquisition of hyposmoregulatory capacity. The growth hormone (Gh)/insulin-like growth-factor (Igf) axis promotes the development of branchial ionoregulatory functions that underlie ion secretion. Igfs interact with a suite of Igf binding proteins (Igfbps) that modulate hormone activity. In Atlantic salmon smolts, igfbp4,−5a,−5b1,−5b2,−6b1 and−6b2 transcripts are highly expressed in gill. We measured mRNA levels of branchial and hepatic igfbps during smoltification (March, April, and May), desmoltification (July) and following seawater (SW) exposure in March and May. We also characterized parallel changes in a broad suite of osmoregulatory (branchial Na+/K+-ATPase (Nka) activity, Na+ /K + /2Cl − cotransporter 1 (nkcc1) and cystic fibrosis transmembrane regulator 1 (cftr1) transcription) and endocrine (plasma Gh and Igf1) parameters.ResultsIndicative of smoltification, we observed increased branchial Nka activity, nkcc1 and cftr1 transcription in May. Branchial igfbp6b1 and -6b2 expression increased coincidentally with smoltification. Following a SW challenge in March, igfbp6b1 showed increased expression while igfbp6b2 exhibited diminished expression. igfbp5a,−5b1 and−5b2 mRNA levels did not change during smolting, but each had lower levels following a SW exposure in March.ConclusionsSalmonids express an especially large suite of igfbps. Our data suggest that dynamic expression of particular igfbps accompanies smoltification and SW challenges; thus, transcriptional control of igfbps may provide a mechanism for the local modulation of Igf activity in salmon gill.

Expression and activity of Rac1 is negatively affected in the dehydroepiandrosterone induced polycystic ovary of mouse

PubMed Central

2014-01-01

Background Polycystic ovarian syndrome (PCOS) is characterized by the presence of multiple follicular cysts, giving rise to infertility due to anovulation. This syndrome affects about 10% of women, worldwide. The exact molecular mechanism leading to PCOS remains obscure. RhoGTPase has been associated with oogenesis, but its role in PCOS remains unexplored. Therefore, we attempted to elucidate the Vav-Rac1 signaling in PCOS mice model. Methods We generated a PCOS mice model by injecting dehydroepiandrosterone (DHEA) for a period of 20 days. The expression levels of Rac1, pRac1, Vav, pVav and Caveolin1 were analyzed by employing immuno-blotting and densitometry. The association between Vav and Rac1 proteins were studied by immuno-precipitation. Furthermore, we analyzed the activity of Rac1 and levels of inhibin B and 17β-estradiol in ovary using biochemical assays. Results The presence of multiple follicular cysts in ovary were confirmed by histology. The activity of Rac1 (GTP bound state) was significantly reduced in the PCOS ovary. Similarly, the expression levels of Rac1 and its phosphorylated form (pRac1) were decreased in PCOS in comparison to the sham ovary. The expression level and activity (phosphorylated form) of guanine nucleotide exchanger of Rac1, Vav, was moderately down-regulated. We observed comparatively increased expressions of Caveolin1, 17β-estradiol, and inhibin B in the polycystic ovary. Conclusion We conclude that hyperandrogenization (PCOS) by DHEA diminishes ovarian Rac1 and Vav expression and activity along with an increase in expression of Caveolin1. This is accompanied by an increase in the intra-ovarian level of '17 β-estradiol and inhibin B. PMID:24628852

Shikonin Inhibites Migration and Invasion of Thyroid Cancer Cells by Downregulating DNMT1

PubMed Central

Zhang, Yue; Sun, Bin; Huang, Zhi

2018-01-01

Background Shikonin is a component of Chinese herbal medicine. The aim of this study was to investigate the effects of shikonin on cell migration of papillary thyroid cancer cells of the TPC-1 cell line in vitro and expression levels of the phosphate and tensin homolog deleted on chromosome 10 (PTEN) and DNA methyltransferase 1 (DNMT1) genes. Material/Methods The Cell Counting Kit-8 (CCK-8) assay was performed to evaluate the proliferation of TPC-1 papillary thyroid cancer cells, and the normal thyroid cells, HTori-3, in vitro. A transwell motility assay was used to analyze the migration of TPC-1 cells. Western blot was performed to determine the expression levels of PTEN and DNMT1 genes. A methylation-specific polymerase chain reaction (PCR) (MSP) assay was used to evaluate the methylation of PTEN. Results Following treatment with shikonin, the cell survival rate of TPC-1 cells decreased in a dose-dependent manner; the inhibitory effects on HTori-3 cells were less marked. Shikonin inhibited TPC-1 cell migration and invasion in a dose-dependent manner. The methylation of PTEN was suppressed by shikonin, which also reduced the expression of DNMT1 in a dose-dependent manner, and increased the expression of PTEN. Overexpression of DNMT1 promoted the migration of TPC-1 cells and the methylation of PTEN. Levels of protein expression of PTEN in TPC-1 cells treated with shikonin decreased, and were increased by DNMT1 knockdown. Conclusions Shikonin suppressed the expression of DNMT1, reduced PTEN gene methylation, and increased PTEN protein expression, leading to the inhibition of TPC-1 cell migration. PMID:29389913

Differential expression of cysteine desulfurases in soybean

PubMed Central

2011-01-01

Background Iron-sulfur [Fe-S] clusters are prosthetic groups required to sustain fundamental life processes including electron transfer, metabolic reactions, sensing, signaling, gene regulation and stabilization of protein structures. In plants, the biogenesis of Fe-S protein is compartmentalized and adapted to specific needs of the cell. Many environmental factors affect plant development and limit productivity and geographical distribution. The impact of these limiting factors is particularly relevant for major crops, such as soybean, which has worldwide economic importance. Results Here we analyze the transcriptional profile of the soybean cysteine desulfurases NFS1, NFS2 and ISD11 genes, involved in the biogenesis of [Fe-S] clusters, by quantitative RT-PCR. NFS1, ISD11 and NFS2 encoding two mitochondrial and one plastid located proteins, respectively, are duplicated and showed distinct transcript levels considering tissue and stress response. NFS1 and ISD11 are highly expressed in roots, whereas NFS2 showed no differential expression in tissues. Cold-treated plants showed a decrease in NFS2 and ISD11 transcript levels in roots, and an increased expression of NFS1 and ISD11 genes in leaves. Plants treated with salicylic acid exhibited increased NFS1 transcript levels in roots but lower levels in leaves. In silico analysis of promoter regions indicated the presence of different cis-elements in cysteine desulfurase genes, in good agreement with differential expression of each locus. Our data also showed that increasing of transcript levels of mitochondrial genes, NFS1/ISD11, are associated with higher activities of aldehyde oxidase and xanthine dehydrogenase, two cytosolic Fe-S proteins. Conclusions Our results suggest a relationship between gene expression pattern, biochemical effects, and transcription factor binding sites in promoter regions of cysteine desulfurase genes. Moreover, data show proportionality between NFS1 and ISD11 genes expression. PMID:22099069

Development of an optimized tetracycline-inducible expression system to increase the accumulation of interleukin-10 in tobacco BY-2 suspension cells

PubMed Central

2012-01-01

Background Plant cell suspension cultures can be used for the production of valuable pharmaceutical and industrial proteins. When the recombinant protein is secreted into the culture medium, restricting expression to a defined growth phase can improve both the quality and quantity of the recovered product by minimizing proteolytic activity. Temporal restriction is also useful for recombinant proteins whose constitutive expression affects cell growth and viability, such as viral interleukin-10 (vIL-10). Results We have developed a novel, tetracycline-inducible system suitable for tobacco BY-2 suspension cells which increases the yields of vIL-10. The new system is based on a binary vector that is easier to handle than conventional vectors, contains an enhanced inducible promoter and 5′-UTR to improve yields, and incorporates a constitutively-expressed visible marker gene to allow the rapid and straightforward selection of the most promising transformed clones. Stable transformation of BY-2 cells with this vector, without extensive optimization of the induction conditions, led to a 3.5 fold increase in vIL-10 levels compared to constitutive expression in the same host. Conclusions We have developed an effective and straightforward molecular farming platform technology that improves both the quality and the quantity of recombinant proteins produced in plant cells, particularly those whose constitutive expression has a negative impact on plant growth and development. Although we tested the platform using vIL-10 produced in BY-2 cells, it can be applied to other host/product combinations and is also useful for basic research requiring strictly controlled transgene expression. PMID:22784336

Pancreatic stellate cells and CX3CR1: occurrence in normal pancreas, acute and chronic pancreatitis and effect of their activation by a CX3CR1 agonist

PubMed Central

Uchida, Masahiko; Ito, Tetsuhide; Nakamura, Taichi; Hijioka, Masayuki; Igarashi, Hisato; Oono, Takamasa; Kato, Masaki; Nakamura, Kazuhiko; Suzuki, Koichi; Takayanagi, Ryoichi; Jensen, Robert T.

2014-01-01

Objectives Numerous studies suggest important roles of the chemokine, fractalkine (CX3CL1) in acute/chronic pancreatitis, however the possible mechanisms of the effects are unclear. Pancreatic stellate cells (PSCs) can play important roles in pancreatitis, secreting inflammatory cytokines/chemokines, as well as proliferation. Therefore, we investigated CX3CL1 receptor (CX3CR1) occurrence in normal pancreas and pancreatitis (acute/chronic) tissues, and the effects of CX3CL1 on activated-PSCs. Methods CX3CR1 expression/localization in normal pancreas and pancreatitis (acute/chronic) tissues were evaluated with immunohistochemical analysis. CX3CR1 expression and effects of CX3CL1 on activated-PSCs were examined with realtime-PCR, BrdU assays and Western Blotting. Results In normal pancreas, acinar cells expressed CX3CR1 within granule-like-formations in the cytoplasm, whereas in acute/chronic pancreatitis, acinar, ductal and activated-PSCs expressed CX3CR1 on cell membranes. With activation of normal PSCs, CX3CR1 is increased. CX3CL1 activated multiple signaling cascades in PSCs. CX3CL1, did not induce inflammatory-genes expression in activated-PSCs, but induced proliferation. Conclusions CX3CR1s are expressed in normal pancreas. Expression is increased in acute/chronic pancreatitis and the CX3CR1s are activated. CX3CL1 induces proliferation of activated-PSCs without increasing release of inflammatory-mediators. These results suggest that CX3CR1 activation of PSCs could be important in their effects in pancreatitis, especially to PSCs proliferation in pancreatitis where CX3CL1 levels are elevated. PMID:24681877

Relationship Between Quantitative GRB7 RNA Expression and Recurrence after Adjuvant Anthracycline Chemotherapy in Triple Negative Breast Cancer

PubMed Central

Sparano, Joseph A.; Goldstein, Lori J.; Childs, Barrett H.; Shak, Steven; Brassard, Diana; Badve, Sunil; Baehner, Frederick L.; Bugarini, Roberto; Rowley, Steve; Perez, Edith; Shulman, Lawrence N.; Martino, Silvana; Davidson, Nancy E.; Kenny, Paraic A.; Sledge, George W.; Gray, Robert

2012-01-01

Purpose To perform an exploratory analysis of the relationship between gene expression and recurrence in patients with operable triple negative breast cancer (TNBC) treated with adjuvant doxorubicin-containing chemotherapy. Experimental design RNA was extracted from archived tumor samples derived from 246 patients with stage I-III TNBC treated with adjuvant doxorubicin-containing chemotherapy, and was analyzed by quantitative RT-PCR for a panel of 374 genes. The relationship between gene expression and recurrence was evaluated using weighted Cox proportional hazards model score tests. Results GRB7 was the only gene for which higher expression was significantly associated with increased recurrence in TNBC (Korn’s adjusted p value=0.04). In a Cox proportional hazards model adjusted for clinicopathologic features, higher GRB7 expression was associated with an increased recurrence risk (HR 2.31, p=0.04 using the median as the split). The 5-year recurrence rates were 10.5% (95% confidence intervals [CI] 7.8%, 14.1%) in the low and 20.4% (95% CI 16.5%, 25.0%) in the high GRB7 groups. External validation in other datasets indicated that GRB7 expression was not prognostic in two adjuvant trials including variable systemic therapy, but in two other trials showed that high GBR7 expression was associated with resistance to neoadjuvant doxorubicin and taxane therapy. Conclusions GRB7 was associated with an increased risk of recurrence in TNBC, suggesting that GRB7 or GRB7-dependent pathways may serve as potential biomarkers for therapeutic targets. Therapeutic targeting of one or more factors identified which function as interaction nodes or effectors should also be considered. PMID:21933890

Sweet Taste Receptor Expressed in Pancreatic β-Cells Activates the Calcium and Cyclic AMP Signaling Systems and Stimulates Insulin Secretion

PubMed Central

Nakagawa, Yuko; Nagasawa, Masahiro; Yamada, Satoko; Hara, Akemi; Mogami, Hideo; Nikolaev, Viacheslav O.; Lohse, Martin J.; Shigemura, Noriatsu; Ninomiya, Yuzo; Kojima, Itaru

2009-01-01

Background Sweet taste receptor is expressed in the taste buds and enteroendocrine cells acting as a sugar sensor. We investigated the expression and function of the sweet taste receptor in MIN6 cells and mouse islets. Methodology/Principal Findings The expression of the sweet taste receptor was determined by RT–PCR and immunohistochemistry. Changes in cytoplasmic Ca2+ ([Ca2+]c) and cAMP ([cAMP]c) were monitored in MIN6 cells using fura-2 and Epac1-camps. Activation of protein kinase C was monitored by measuring translocation of MARCKS-GFP. Insulin was measured by radioimmunoassay. mRNA for T1R2, T1R3, and gustducin was expressed in MIN6 cells. In these cells, artificial sweeteners such as sucralose, succharin, and acesulfame-K increased insulin secretion and augmented secretion induced by glucose. Sucralose increased biphasic increase in [Ca2+]c. The second sustained phase was blocked by removal of extracellular calcium and addition of nifedipine. An inhibitor of inositol(1, 4, 5)-trisphophate receptor, 2-aminoethoxydiphenyl borate, blocked both phases of [Ca2+]c response. The effect of sucralose on [Ca2+]c was inhibited by gurmarin, an inhibitor of the sweet taste receptor, but not affected by a Gq inhibitor. Sucralose also induced sustained elevation of [cAMP]c, which was only partially inhibited by removal of extracellular calcium and nifedipine. Finally, mouse islets expressed T1R2 and T1R3, and artificial sweeteners stimulated insulin secretion. Conclusions Sweet taste receptor is expressed in β-cells, and activation of this receptor induces insulin secretion by Ca2+ and cAMP-dependent mechanisms. PMID:19352508

Clinical Significance of SASH1 Expression in Glioma

PubMed Central

Yang, Liu; Zhang, Haitao; Yao, Qi; Yan, Yingying; Wu, Ronghua; Liu, Mei

2015-01-01

Objective. SAM and SH3 domain containing 1 (SASH1) is a recently discovered tumor suppressor gene. The role of SASH1 in glioma has not yet been described. We investigated SASH1 expression in glioma cases to determine its clinical significance on glioma pathogenesis and prognosis. Methods. We produced tissue microarrays using 121 patient-derived glioma samples and 30 patient-derived nontumor cerebral samples. Immunohistochemistry and Western blotting were used to evaluate SASH1 expression. We used Fisher's exact tests to determine relationships between SASH1 expression and clinicopathological characteristics; Cox regression analysis to evaluate the independency of different SASH1 expression; Kaplan-Meier analysis to determine any correlation of SASH1 expression with survival rate. Results. SASH1 expression was closely correlated with the WHO glioma grade. Of the 121 cases, 66.9% with low SASH1 expression were mostly grade III-IV cases, whereas 33.1% with high SASH1 expression were mostly grades I-II. Kaplan-Meier analysis revealed a significant positive correlation between SASH1 expression and postoperative survival. Conclusions. SASH1 was widely expressed in normal and low-grade glioma tissues. SASH1 expression strongly correlated with glioma grades, showing higher expression at a lower grade, which decreased significantly as grade increased. Furthermore, SASH1 expression was positively correlated with better postoperative survival in patients with glioma. PMID:26424902

Characterization and Expression of Glutamate Dehydrogenase in Response to Acute Salinity Stress in the Chinese Mitten Crab, Eriocheir sinensis

PubMed Central

Wang, Yueru; Li, Erchao; Yu, Na; Wang, Xiaodan; Cai, Chunfang; Tang, Boping; Chen, Liqiao; Van Wormhoudt, Alain

2012-01-01

Background Glutamate dehydrogenase (GDH) is a key enzyme for the synthesis and catabolism of glutamic acid, proline and alanine, which are important osmolytes in aquatic animals. However, the response of GDH gene expression to salinity alterations has not yet been determined in macro-crustacean species. Methodology/Principal Findings GDH cDNA was isolated from Eriocheir sinensis. Then, GDH gene expression was analyzed in different tissues from normal crabs and the muscle of crabs following transfer from freshwater (control) directly to water with salinities of 16‰ and 30‰, respectively. Full-length GDH cDNA is 2,349 bp, consisting of a 76 bp 5′- untranslated region, a 1,695 bp open reading frame encoding 564 amino acids and a 578 bp 3′- untranslated region. E. sinensis GDH showed 64–90% identity with protein sequences of mammalian and crustacean species. Muscle was the dominant expression source among all tissues tested. Compared with the control, GDH expression significantly increased at 6 h in crabs transferred to 16‰ and 30‰ salinity, and GDH expression peaked at 48 h and 12 h, respectively, with levels approximately 7.9 and 8.5 fold higher than the control. The free amino acid (FAA) changes in muscle, under acute salinity stress (16‰ and 30‰ salinities), correlated with GDH expression levels. Total FAA content in the muscle, which was based on specific changes in arginine, proline, glycine, alanine, taurine, serine and glutamic acid, tended to increase in crabs following transfer to salt water. Among these, arginine, proline and alanine increased significantly during salinity acclimation and accounted for the highest proportion of total FAA. Conclusions E. sinensis GDH is a conserved protein that serves important functions in controlling osmoregulation. We observed that higher GDH expression after ambient salinity increase led to higher FAA metabolism, especially the synthesis of glutamic acid, which increased the synthesis of proline and alanine to meet the demand of osmoregulation at hyperosmotic conditions. PMID:22615974

Factors Expressed in an Animal Model of Anteroinferior Glenohumeral Instability

PubMed Central

Mulcahey, Mary K.; Marshall, Mindy; Gallacher, Stacey E.; Kaback, Lee A.; Blaine, Theodore A.

2015-01-01

Background: There is little information on the molecular factors important in healing and changes that occur in the glenoid labrum in response to injury. Using a novel animal model of acute anterior shoulder dislocation, this study characterizes the factors expressed in the glenoid labrum in response to injury and correlates their expression to glenohumeral stability. Purpose: To study the response of the glenoid labrum to injury both biomechanically and with immunohistochemical testing. Methods: An injury to the anteroinferior labrum was surgically induced in 50 male Lewis rats. Rats were sacrificed at 3, 7, 14, 28, or 42 days. Immunolocalization experiments were performed to localize the expression of growth factors and cytokines. For biomechanical testing, dynamic stiffness for anterior and posterior laxity, load to failure, stiffness, and maximum load were recorded. Statistical differences were determined at P < .05. Study Design: Descriptive laboratory study. Results: Expression of interleukin–1 beta (IL-1β), transforming growth factor–beta 1 (TGF-β1), matrix metalloproteinase 3 (MMP3), and matrix metalloproteinase 13 (MMP13) were increased in injured compared with uninjured specimens. Collagen III expression was increased early and decreased with time. Biomechanical testing verified instability by demonstrating increased anterior displacement and decreased stiffness in injured shoulders at all time points. Conclusion: This novel animal model of acute anterior shoulder dislocation showed increased expression of IL-1β, TGF-β1, MMP3, MMP13, and collagen III in the injured labral tissue at early time points. Increased anterior laxity and decreased stiffness and maximum load to failure were seen after anterior labral injury, supporting the model’s ability to re-create anterior glenohumeral instability. These data provide important information on the temporal changes occurring in a rat model of anterior glenohumeral dislocation. Clinical Relevance: Identification of factors expressed in the anterior capsule and glenoid labrum in response to injury may lead to the development of novel agents that can be used to augment glenoid labrum healing and ultimately improve both surgical and nonsurgical treatment of this common shoulder injury. PMID:26535392

Patients with Encapsulating Peritoneal Sclerosis Have Increased Peritoneal Expression of Connective Tissue Growth Factor (CCN2), Transforming Growth Factor-β1, and Vascular Endothelial Growth Factor

PubMed Central

Abrahams, Alferso C.; Habib, Sayed M.; Dendooven, Amélie; Riser, Bruce L.; van der Veer, Jan Willem; Toorop, Raechel J.; Betjes, Michiel G. H.; Verhaar, Marianne C.; Watson, Christopher J. E.; Nguyen, Tri Q.; Boer, Walther H.

2014-01-01

Introduction Encapsulating peritoneal sclerosis (EPS) is a devastating complication of peritoneal dialysis (PD). The pathogenesis is not exactly known and no preventive strategy or targeted medical therapy is available. CCN2 has both pro-fibrotic and pro-angiogenic actions and appears an attractive target. Therefore, we studied peritoneal expression of CCN2, as well as TGFβ1 and VEGF, in different stages of peritoneal fibrosis. Materials and methods Sixteen PD patients were investigated and compared to 12 hemodialysis patients and four pre-emptively transplanted patients. Furthermore, expression was investigated in 12 EPS patients in comparison with 13 PD and 12 non-PD patients without EPS. Peritoneal tissue was taken during kidney transplantation procedure or during EPS surgery. In a subset of patients, CCN2 protein levels in peritoneal effluent and plasma were determined. Samples were examined by qPCR, histology, immunohistochemistry, and ELISA. Results Peritoneal CCN2 expression was 5-fold higher in PD patients compared to pre-emptively transplanted patients (P<0.05), but did not differ from hemodialysis patients. Peritoneal expression of TGFβ1 and VEGF were not different between the three groups; neither was peritoneal thickness. Peritoneum of EPS patients exhibited increased expression of CCN2 (35-fold, P<0.001), TGFβ1 (24-fold, P<0.05), and VEGF (77-fold, P<0.001) compared to PD patients without EPS. In EPS patients, CCN2 protein was mainly localized in peritoneal endothelial cells and fibroblasts. CCN2 protein levels were significantly higher in peritoneal effluent of EPS patients compared to levels in dialysate of PD patients (12.0±4.5 vs. 0.91±0.92 ng/ml, P<0.01), while plasma CCN2 levels were not increased. Conclusions Peritoneal expression of CCN2, TGFβ1, and VEGF are significantly increased in EPS patients. In early stages of peritoneal fibrosis, only CCN2 expression is slightly increased. Peritoneal CCN2 overexpression in EPS patients is a locally driven response. The potential of CCN2 as biomarker and target for CCN2-inhibiting agents to prevent or treat EPS warrants further study. PMID:25384022

Elevation of Il6 is associated with disturbed let-7 biogenesis in a genetic model of depression

PubMed Central

Wei, Y B; Liu, J J; Villaescusa, J C; Åberg, E; Brené, S; Wegener, G; Mathé, A A; Lavebratt, C

2016-01-01

Elevation of the proinflammatory cytokine IL-6 has been implicated in depression; however, the mechanisms remain elusive. MicroRNAs (miRNAs) are small non-coding RNAs that inhibit gene expression post-transcriptionally. The lethal-7 (let-7) miRNA family was suggested to be involved in the inflammation process and IL-6 was shown to be one of its targets. In the present study, we report elevation of Il6 in the prefrontal cortex (PFC) of a genetic rat model of depression, the Flinders Sensitive Line (FSL) compared to the control Flinders Resistant Line. This elevation was associated with an overexpression of LIN28B and downregulation of let-7 miRNAs, the former an RNA-binding protein that selectively represses let-7 synthesis. Also DROSHA, a key enzyme in miRNA biogenesis was downregulated in FSL. Running was previously shown to have an antidepressant-like effect in the FSL rat. We found that running reduced Il6 levels and selectively increased let-7i and miR-98 expression in the PFC of FSL, although there were no differences in LIN28B and DROSHA expression. Pri-let-7i was upregulated in the running FSL group, which associated with increased histone H4 acetylation. In conclusion, the disturbance of let-7 family biogenesis may underlie increased proinflammatory markers in the depressed FSL rats while physical activity could reduce their expression, possibly through regulating primary miRNA expression via epigenetic mechanisms. PMID:27529677

Elevation of Il6 is associated with disturbed let-7 biogenesis in a genetic model of depression.

PubMed

Wei, Y B; Liu, J J; Villaescusa, J C; Åberg, E; Brené, S; Wegener, G; Mathé, A A; Lavebratt, C

2016-08-16

Elevation of the proinflammatory cytokine IL-6 has been implicated in depression; however, the mechanisms remain elusive. MicroRNAs (miRNAs) are small non-coding RNAs that inhibit gene expression post-transcriptionally. The lethal-7 (let-7) miRNA family was suggested to be involved in the inflammation process and IL-6 was shown to be one of its targets. In the present study, we report elevation of Il6 in the prefrontal cortex (PFC) of a genetic rat model of depression, the Flinders Sensitive Line (FSL) compared to the control Flinders Resistant Line. This elevation was associated with an overexpression of LIN28B and downregulation of let-7 miRNAs, the former an RNA-binding protein that selectively represses let-7 synthesis. Also DROSHA, a key enzyme in miRNA biogenesis was downregulated in FSL. Running was previously shown to have an antidepressant-like effect in the FSL rat. We found that running reduced Il6 levels and selectively increased let-7i and miR-98 expression in the PFC of FSL, although there were no differences in LIN28B and DROSHA expression. Pri-let-7i was upregulated in the running FSL group, which associated with increased histone H4 acetylation. In conclusion, the disturbance of let-7 family biogenesis may underlie increased proinflammatory markers in the depressed FSL rats while physical activity could reduce their expression, possibly through regulating primary miRNA expression via epigenetic mechanisms.

MRP1 and glucosylceramide are coordinately over expressed and enriched in rafts during multidrug resistance acquisition in colon cancer cells.

PubMed

Klappe, Karin; Hinrichs, John W J; Kroesen, Bart-Jan; Sietsma, Hannie; Kok, Jan Willem

2004-07-01

Previously we have described a novel multidrug-resistant cell line, HT29(col), which displayed over expression of the multidrug-resistance protein 1 (MRP1) and an altered sphingolipid composition, including enhanced levels of glucosylceramide (GlcCer; Kok JW, Veldman RJ, Klappe K, Koning H, Filipeanu C, Muller M. Int J Cancer 2000;87:172-8). In our study, long-term screening revealed that, during colchicine-induced acquisition of multidrug resistance in a new HT29(col) cell line, increases in GlcCer occurred concomitantly with upregulation of MRP1 expression. Both MRP1 and GlcCer were found enriched in Lubrol-insoluble membrane domains. The expression of MRP1 and GlcCer were tightly correlated, as indicated also by a reversal of both at the later stage of colchicine consolidation. Resistance to colchicine was determined by MRP1, while glucosylceramide synthase (GCS) did not contribute: 1). Resistance was fully inhibited by MK571. 2). GCS expression and activity were not upregulated in HT29(col) cells. 3). Inhibition of GCS did not affect MRP1-mediated efflux function or sensitivity to colchicine. Instead, overall sphingolipid metabolism was upregulated through an increased rate of ceramide biosynthesis. In conclusion, upregulation of MRP1 occurs in concert with upregulation of GlcCer during multidrug-resistance acquisition, and both are enriched in rafts. The increased GlcCer pool does not directly modulate MRP1 function and cell survival. Copyright 2004 Wiley-Liss, Inc.

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

He, Xiao-dong; Tobo, Masayuki; Mogi, Chihiro

Highlights: Black-Right-Pointing-Pointer Glucocorticoid (GC) induced the expression of proton-sensing TDAG8 in macrophages. Black-Right-Pointing-Pointer GC enhanced acidic pH-induced cAMP accumulation and inhibition of TNF-{alpha} production. Black-Right-Pointing-Pointer The enhancement of the GC-induced actions was lost by TDAG8 deficiency. Black-Right-Pointing-Pointer GC-induced anti-inflammatory actions are partly mediated by TDAG8 expression. -- Abstract: Dexamethasone (DEX), a potent glucocorticoid, increased the expression of T-cell death associated gene 8 (TDAG8), a proton-sensing G protein-coupled receptor, which is associated with the enhancement of acidic pH-induced cAMP accumulation, in peritoneal macrophages. We explored the role of increased TDAG8 expression in the anti-inflammatory actions of DEX. The treatment of macrophagesmore » with either DEX or acidic pH induced the cell death of macrophages; however, the cell death was not affected by TDAG8 deficiency. While DEX inhibited lipopolysaccharide-induced production of tumor necrosis factor-{alpha}, an inflammatory cytokine, which was independent of TDAG8, at neutral pH, the glucocorticoid enhanced the acidic pH-induced inhibition of tumor necrosis factor-{alpha} production in a manner dependent on TDAG8. In conclusion, the DEX-induced increase in TDAG8 expression is in part involved in the glucocorticoid-induced anti-inflammatory actions through the inhibition of inflammatory cytokine production under the acidic pH environment. On the other hand, the role of TDAG8 in the DEX-induced cell death is questionable.« less

Effects of castration on expression of lipid metabolism genes in the liver of korean cattle.

PubMed

Baik, Myunggi; Nguyen, Trang Hoa; Jeong, Jin Young; Piao, Min Yu; Kang, Hyeok Joong

2015-01-01

Castration induces the accumulation of body fat and deposition of intramuscular fat in Korean cattle, resulting in improved beef quality. However, little is known about the metabolic adaptations in the liver following castration. To understand changes in lipid metabolism following castration, hepatic expression levels of lipid metabolism genes were compared between Korean bulls and steers. Steers had higher (p<0.001) hepatic lipids contents and higher (p<0.01) mRNA levels of lipogenic acetyl-CoA carboxylase. This differential gene expression may, in part, contribute to increased hepatic lipid content following the castration of bulls. However, we found no differences in the hepatic expression levels of genes related to triglyceride synthesis (mitochondrial glycerol-3-phosphate acyltransferase, diacylglycerol O-acyltransferase 1 and 2) and fatty acid (FA) oxidation (carnitine palmitoyltransferase 1A, C-4 to C-12 straight chain acyl-CoA dehydrogenase, very long chain acyl-CoA dehydrogenase) between bulls and steers. No differences in gene expression for very-low-density lipoprotein (VLDL) secretion, including apolipoprotein B mRNA and microsomal triglyceride transfer protein (MTTP) protein, were observed in the liver although MTTP mRNA levels were higher in steers compared to bulls. In conclusion, FA synthesis may contribute to increased hepatic lipid deposition in steers following castration. However, hepatic lipid metabolism, including triglyceride synthesis, FA oxidation, and VLDL secretion, was not significantly altered by castration. Our results suggest that hepatic lipid metabolism does not significantly contribute to increased body fat deposition in steers following castration.

Increased Intraepithelial Vα24 Invariant NKT Cells in the Celiac Duodenum

PubMed Central

Montalvillo, Enrique; Bernardo, David; Martínez-Abad, Beatriz; Allegretti, Yessica; Fernández-Salazar, Luis; Calvo, Carmen; Chirdo, Fernando G.; Garrote, José A.; Arranz, Eduardo

2015-01-01

Celiac Disease (CD) is an interferon (IFN)γ-mediated duodenal hypersensitivity to wheat gluten occurring in genetically predisposed individuals. Gluten-free diet (GFD) leads to a complete remission of the disease. Vα24-restricted invariant NKT (iNKT) cells are important to maintain immune homeostasis in the gut mucosa because of their unique capacity to rapidly produce large quantities of both T-helper (Th)1 and Th2 cytokines upon stimulation. We studied the presence of these cells in the CD duodenum. Duodenal biopsies were obtained from 45 untreated-CD patients (uCD), 15 Gluten Free Diet-CD patients (GFD-CD), 44 non-inflamed non-CD controls (C-controls) and 15 inflamed non-CD controls (I-controls). Two populations from Spain and Argentina were recruited. Messenger RNA (mRNA) expression of Vα24-Jα18 (invariant TCRα chain of human iNKT cells), IFNγ and intracellular transcription factor Forkhead Box P3 (Foxp3), and flow cytometry intraepithelial lymphocyte (IEL) profile were determined. Both uCD and GFD-CD patients had higher Vα24-Jα18 mRNA levels than non-CD controls (I and C-controls). The expression of Vα24-Jα18 correlated with Marsh score for the severity of mucosal lesion and also with increased mRNA IFNγ levels. uCD and GFD-CD patients had decreased mRNA expression of FoxP3 but increased expression of Vα24-Jα18, which revealed a CD-like molecular profile. Increased numbers of iNKT cells were confirmed by flow cytometry within the intraepithelial lymphocyte compartment of uCD and GFD-CD patients and correlated with Vα24-Jα18 mRNA expression. In conclusion, we have found an increased number of iNKT cells in the duodenum from both uCD and GFD-CD patients, irrespective of the mucosal status. A CD-like molecular profile, defined by an increased mRNA expression of Vα24-Jα18 together with a decreased expression of FoxP3, may represent a pro-inflammatory signature of the CD duodenum. PMID:26529008

Increased Intraepithelial Vα24 Invariant NKT Cells in the Celiac Duodenum.

PubMed

Montalvillo, Enrique; Bernardo, David; Martínez-Abad, Beatriz; Allegretti, Yessica; Fernández-Salazar, Luis; Calvo, Carmen; Chirdo, Fernando G; Garrote, José A; Arranz, Eduardo

2015-10-30

Celiac Disease (CD) is an interferon (IFN)γ-mediated duodenal hypersensitivity to wheat gluten occurring in genetically predisposed individuals. Gluten-free diet (GFD) leads to a complete remission of the disease. Vα24-restricted invariant NKT (iNKT) cells are important to maintain immune homeostasis in the gut mucosa because of their unique capacity to rapidly produce large quantities of both T-helper (Th)1 and Th2 cytokines upon stimulation. We studied the presence of these cells in the CD duodenum. Duodenal biopsies were obtained from 45 untreated-CD patients (uCD), 15 Gluten Free Diet-CD patients (GFD-CD), 44 non-inflamed non-CD controls (C-controls) and 15 inflamed non-CD controls (I-controls). Two populations from Spain and Argentina were recruited. Messenger RNA (mRNA) expression of Vα24-Jα18 (invariant TCRα chain of human iNKT cells), IFNγ and intracellular transcription factor Forkhead Box P3 (Foxp3), and flow cytometry intraepithelial lymphocyte (IEL) profile were determined. Both uCD and GFD-CD patients had higher Vα24-Jα18 mRNA levels than non-CD controls (I and C-controls). The expression of Vα24-Jα18 correlated with Marsh score for the severity of mucosal lesion and also with increased mRNA IFNγ levels. uCD and GFD-CD patients had decreased mRNA expression of FoxP3 but increased expression of Vα24-Jα18, which revealed a CD-like molecular profile. Increased numbers of iNKT cells were confirmed by flow cytometry within the intraepithelial lymphocyte compartment of uCD and GFD-CD patients and correlated with Vα24-Jα18 mRNA expression. In conclusion, we have found an increased number of iNKT cells in the duodenum from both uCD and GFD-CD patients, irrespective of the mucosal status. A CD-like molecular profile, defined by an increased mRNA expression of Vα24-Jα18 together with a decreased expression of FoxP3, may represent a pro-inflammatory signature of the CD duodenum.

Human umbilical cord mesenchymal stromal cells suppress MHC class II expression on rat vascular endothelium and prolong survival time of cardiac allograft

PubMed Central

Qiu, Ying; Yun, Mark M; Han, Xia; Zhao, Ruidong; Zhou, Erxia; Yun, Sheng

2014-01-01

Background: Human umbilical cord mesenchymal stromal cells (UC-MSCs) have low immunogenicity and immune regulation. To investigate immunomodulatory effects of human UC-MSCs on MHC class II expression and allograft, we transplanted heart of transgenic rats with MHC class II expression on vascular endothelium. Methods: UC-MSCs were obtained from human umbilical cords and confirmed with flow cytometry analysis. Transgenic rat line was established using the construct of human MHC class II transactivator gene (CIITA) under mouse ICAM-2 promoter control. The induced MHC class II expression on transgenic rat vascular endothelial cells (VECs) was assessed with immunohistological staining. And the survival time of cardiac allograft was compared between the recipients with and without UC-MSC transfusion. Results: Flow cytometry confirmed that the human UC-MSCs were positive for CD29, CD44, CD73, CD90, CD105, CD271, and negative for CD34 and HLA-DR. Repeated infusion of human UC-MSCs reduced MHC class II expression on vascular endothelia of transplanted hearts, and increased survival time of allograft. The UC-MSCs increased regulatory cytokines IL10, transforming growth factor (TGF)-β1 and suppressed proinflammatory cytokines IL2 and IFN-γ in vivo. The UC-MSC culture supernatant had similar effects on cytokine expression, and decreased lymphocyte proliferation in vitro. Conclusions: Repeated transfusion of the human UC-MSCs reduced MHC class II expression on vascular endothelia and prolonged the survival time of rat cardiac allograft. PMID:25126177

Characterization of the Regulation and Function of Zinc-Dependent Histone Deacetylases During Mouse Liver Regeneration

PubMed Central

Huang, Jiansheng; Barr, Emily; Rudnick, David A.

2013-01-01

The studies reported here were undertaken to define the regulation and functional importance of zinc-dependent histone deacetylase (Zn-HDAC) activity during liver regeneration using the mouse partial hepatectomy (PH) model. The results showed that hepatic HDAC activity was significantly increased in nuclear and cytoplasmic fractions following PH. Further analyses showed isoform-specific effects of PH on HDAC mRNA and protein expression, with increased expression of the class I HDACs, 1 and 8, and class II HDAC4 in regenerating liver. Hepatic expression of (class II) HDAC5 was unchanged after PH; however HDAC5 exhibited transient nuclear accumulation in regenerating liver. These changes in hepatic HDAC expression, subcellular localization, and activity coincided with diminished histone acetylation in regenerating liver. The significance of these events was investigated by determining the effects of suberoylanilide hydroxyamic acid (SAHA, a specific inhibitor of Zn-HDAC activity) on hepatic regeneration. The results showed that SAHA-treatment suppressed the effects of PH on histone deacetylation and hepatocellular BrdU incorporation. Further examination showed that SAHA blunted hepatic expression and activation of cell cycle signals downstream of induction of cyclin D1 expression in mice subjected to PH. Conclusion The data reported here demonstrate isoform-specific regulation of Zn-HDAC expression, subcellular localization, and activity in regenerating liver. These studies also indicate that HDAC activity promotes liver regeneration by regulating hepatocellular cell cycle progression at a step downstream of cyclin D1 induction. PMID:23258575

Differential expression and prognostic value of the chemokine receptor CXCR4 in bronchopulmonary neuroendocrine neoplasms

PubMed Central

Specht, Elisa; Wirtz, Ralph M.; Sayeg, Manal; Baum, Richard P.; Schulz, Stefan; Lupp, Amelie

2015-01-01

Introduction For many tumors, the overexpression of the chemokine receptor CXCR4 is associated with increased malignancy and poor patient outcomes. However, comprehensive data for neuroendocrine neoplasms of the lung are still lacking. Methods CXCR4 expression was evaluated in a panel of bronchopulmonary neuroendocrine neoplasms (BP-NEN) comprising typical carcinoids (n = 26), atypical carcinoids (n = 30), and small cell lung cancers (SCLC, n = 34). Samples were analyzed by immunohistochemistry using the novel monoclonal rabbit anti-human CXCR4 antibody UMB-2 and by qRT-PCR. The expression was correlated with clinical data and overall patient survival. Results CXCR4 was predominantly localized at the plasma membrane of the tumor cells. CXCR4 was expressed with a high intensity in almost all of the 30 SCLC samples. In contrast, it was detected infrequently and with low intensity in the typical carcinoid and atypical carcinoid samples. There was a significant correlation between the immunohistochemistry and qRT-PCR data. Additionally, there was a significant negative relationship between CXCR4 expression and overall survival. Conclusions With increasing malignancy, BP-NEN clearly differ in the extent of CXCR4 expression. As in other tumor entities, CXCR4 overexpression significantly correlates with negative patient outcome. Due to its particular high expression rate in SCLC, CXCR4 may serve as a promising new target for diagnostic and pharmacological intervention as well as for peptide receptor-based radionuclide therapy. PMID:25671300

Correlation of matrix metalloproteinase (MMP)-1, -2, -3, and -9 expressions with demographic and radiological features in primary lumbar intervertebral disc disease.

PubMed

Basaran, Recep; Senol, Mehmet; Ozkanli, Seyma; Efendioglu, Mustafa; Kaner, Tuncay

2017-07-01

Degeneration of IVD is a progressive and irreversible process and can be evaluated with immunohistochemical examination or radiological grading. MMPs are a family of proteolytic enzymes and involved in the degradation of the matrix components of the IVD. We aimed to compare MMP-1, -2, -3, and -9 expressions with demographic features, visual analogue scale (VAS), Oswestry Disability Index (ODI) and radiological (MRI) grades. The study involved 60 participants. We recorded data about age, complaint, radiological imaging, expression levels of MMP-1, -2, -3, and -9, ODI and VAS for back pain retrospectively. Intervertebral disc degeneration was graded on a 0-5 scale according to the Pfirrmann classification. As a result of the study, the median age was 52.09±12.74years. There were statistical significances between age and MMP-1, and MMP-2. There was a close correlation between grade and MMP-9. We found correlation between the VAS and the MMP-9 expression. In addition, there was relationship between expression of MMP-2 and MMP-1, MMP-3, MMP-9. In conclusion, the expressions of MMP-1 and -2 are increased with aging. There was no relationship between radiological evaluation of IVDD and aging. Increased expression of MMPs affected IVDD positively. The relationship with MMPs is not explained. This study adds to our understanding of the interaction between MMPs and IVDD. Copyright © 2017 Elsevier Ltd. All rights reserved.

Effect of Antisense Oligodeoxynucleotides Glucose Transporter-1 on Enhancement of Radiosensitivity of Laryngeal Carcinoma

PubMed Central

Yan, Sen-Xiang; Luo, Xing-Mei; Zhou, Shui-Hong; Bao, Yang-Yang; Fan, Jun; Lu, Zhong-Jie; Liao, Xin-Biao; Huang, Ya-Ping; Wu, Ting-Ting; Wang, Qin-Ying

2013-01-01

Purpose: Laryngeal carcinomas always resist to radiotherapy. Hypoxia is an important factor in radioresistance of laryngeal carcinoma. Glucose transporter-1 (GLUT-1) is considered to be a possible intrinsic marker of hypoxia in malignant tumors. We speculated that the inhibition of GLUT-1 expression might improve the radiosensitivity of laryngeal carcinoma. Methods: We assessed the effect of GLUT-1 expression on radioresistance of laryngeal carcinoma and the effect of GLUT-1 expressions by antisense oligodeoxynucleotides (AS-ODNs) on the radiosensitivity of laryngeal carcinoma in vitro and in vivo. Results: After transfection of GLUT-1 AS-ODNs: MTS assay showed the survival rates of radiation groups were reduced with the prolongation of culture time (p<0.05); Cell survival rates were significantly reduced along with the increasing of radiation dose (p<0.05). There was significant difference in the expression of GLUT-1mRNA and protein in the same X-ray dose between before and after X-ray radiation (p<0.05). In vivo, the expressions of GLUT-1 mRNA and protein after 8Gy radiation plus transfection of GLUT-1 AS-ODNs were significant decreased compared to 8Gy radiation alone (p<0.001). Conclusion: Radioresistance of laryngeal carcinoma may be associated with increased expression of GLUT-1 mRNA and protein. GLUT-1 AS-ODNs may enhance the radiosensitivity of laryngeal carcinoma mainly by inhibiting the expression of GLUT-1. PMID:23983599

Overexpression of connexin 43 reduces melanoma proliferative and metastatic capacity

PubMed Central

Tittarelli, A; Guerrero, I; Tempio, F; Gleisner, M A; Avalos, I; Sabanegh, S; Ortíz, C; Michea, L; López, M N; Mendoza-Naranjo, A; Salazar-Onfray, F

2015-01-01

Background: Alterations in connexin 43 (Cx43) expression and/or gap junction (GJ)-mediated intercellular communication are implicated in cancer pathogenesis. Herein, we have investigated the role of Cx43 in melanoma cell proliferation and apoptosis sensitivity in vitro, as well as metastatic capability and tumour growth in vivo. Methods: Connexin 43 expression levels, GJ coupling and proliferation rates were analysed in four different human melanoma cell lines. Furthermore, tumour growth and lung metastasis of high compared with low Cx43-expressing FMS cells were evaluated in vivo using a melanoma xenograft model. Results: Specific inhibition of Cx43 channel activity accelerated melanoma cell proliferation, whereas overexpression of Cx43 increased GJ coupling and reduced cell growth. Moreover, Cx43 overexpression in FMS cells increased basal and tumour necrosis factor-α-induced apoptosis and resulted in decreased melanoma tumour growth and lower number and size of metastatic foci in vivo. Conclusions: Our findings reveal an important role for Cx43 in intrinsically controlling melanoma growth, death and metastasis, and emphasise the potential use of compounds that selectively enhance Cx43 expression on melanoma in the future chemotherapy and/or immunotherapy protocols. PMID:26135897

Methyl jasmonate affects phenolic metabolism and gene expression in blueberry (Vaccinium corymbosum).

PubMed

Cocetta, Giacomo; Rossoni, Mara; Gardana, Claudio; Mignani, Ilaria; Ferrante, Antonio; Spinardi, Anna

2015-02-01

Blueberry (Vaccinium corymbosum) is a fruit very much appreciated by consumers for its antioxidant potential and health-promoting traits. Its beneficial potential properties are mainly due to a high content of anthocyanins and their amount can change after elicitation with methyl jasmonate. The aim of this work is to evaluate the changes in expression of several genes, accumulation of phenolic compounds and alterations in antioxidant potential in two different blueberry cultivars ('Duke' and 'Blueray') in response to methyl jasmonate (0.1 mM). Results showed that 9 h after treatment, the expression of phenylalanine ammonium lyase, chalcone synthase and anthocyanidin synthase genes was stimulated more in the 'Blueray' variety. Among the phenols measured an increase was recorded also for epicatechin and anthocyanin concentrations. 'Duke' is a richer sourche of anthocyanins compared to 'Blueray', treatment with methyl jasmonate promoted in 'Blueray' an increase in pigments as well as in the antioxidant potential, especially in fully ripe berries, but treated 'Duke' berries had greater levels, which were not induced by methyl jasmonate treatment. In conclusion, methyl jasmonate was, in some cases, an effective elicitor of phenolic metabolism and gene expression in blueberry, though with different intensity between cultivars. © 2014 Scandinavian Plant Physiology Society.

Sex differences in body composition, fat storage, and gene expression profile in Caenorhabditis elegans in response to dietary restriction.

PubMed

Miersch, Claudia; Döring, Frank

2013-07-02

The metabolic and health-promoting effects of dietary restriction (DR) have been extensively studied in several species. The response to DR with respect to sex is essentially unknown. To address this question, we used the model organism Caenorhabditis elegans to analyze body composition and gene expression in males and hermaphrodites in response to DR. Unexpectedly, DR increased the fat-to-fat-free mass ratio and enlarged lipid droplets in both sexes to a similar extent. These effects were linked to a downregulation of the lipase-like 5 (lipl-5) gene in both sexes at two developmental stages. By contrast, the reductions in body size, protein content, and total RNA content in response to DR were more pronounced in hermaphrodites than in males. Functional enrichment analysis of gene expression data showed a DR-induced downregulation of several embryogenesis-associated genes concomitant with an ongoing expression of sperm-associated genes in hermaphrodites. In conclusion, DR increases fat stores in both sexes of C. elegans in the form of large and possibly lipolysis-resistant lipid droplets and markedly alters the reproductive program in hermaphrodites but not in males.

The Biological Properties of OGI Surfaces Positively Act on Osteogenic and Angiogenic Commitment of Mesenchymal Stem Cells

PubMed Central

Bressan, Eriberto; Gardin, Chiara; Ferroni, Letizia; Soldini, Maria Costanza; Mandelli, Federico; Soldini, Claudio

2017-01-01

Osteogenesis process displays a fundamental role during dental implant osteointegration. In the present work, we studied the influence of Osteon Growth Induction (OGI) surface properties on the angiogenic and osteogenic behaviors of Mesenchymal Stem cells (MSC). MSC derived from dental pulp and HUVEC (Human Umbilical Vein Endothelial Cells) were grown in on OGI titanium surfaces, and cell proliferation and DNA synthesis were evaluated by MTT [3-(4,5-dimethylthiazol-2yl)-2,5-diphenyltetrazolium bromide] test and DNA quantification. Gene expression has been performed in order to evaluate the presence of mRNA related to endothelial and osteogenesis markers. Moreover, morphological and biochemical analyses of osteogenesis commitments has been performed. On OGI surfaces, MSC and HUVEC are able to proliferate. Gene expression profiler confirms that MSC on OGI surfaces are able to express endothelial and osteogenic markers, and that these expression are higher compared the expression on control surfaces. In conclusion On OGI surfaces proliferation, expression and morphological analyses of angiogenesis-associated markers in MSC are promoted. This process induces an increasing on their osteogenesis commitment. PMID:29149082

Lhx2 Determines Odorant Receptor Expression Frequency in Mature Olfactory Sensory Neurons

PubMed Central

Zhang, Guangfan; Titlow, William B.; Biecker, Stephanie M.; Stromberg, Arnold J.

2016-01-01

Abstract A developmental program of epigenetic repression prepares each mammalian olfactory sensory neuron (OSN) to strongly express one allele from just one of hundreds of odorant receptor (OR) genes, but what completes this process of OR gene choice by driving the expression of this allele is incompletely understood. Conditional deletion experiments in mice demonstrate that Lhx2 is necessary for normal expression frequencies of nearly all ORs and all trace amine-associated receptors, irrespective of whether the deletion of Lhx2 is initiated in immature or mature OSNs. Given previous evidence that Lhx2 binds OR gene control elements, these findings indicate that Lhx2 is directly involved in driving OR expression. The data also support the conclusion that OR expression is necessary to allow immature OSNs to complete differentiation and become mature. In contrast to the robust effects of conditional deletion of Lhx2, the loss of Emx2 has much smaller effects and more often causes increased expression frequencies. Lhx2:Emx2 double mutants show opposing effects on Olfr15 expression that reveal independent effects of these two transcription factors. While Lhx2 is necessary for OR expression that supports OR gene choice, Emx2 can act differently; perhaps by helping to control the availability of OR genes for expression. PMID:27822500

Activating Nrf-2 Signaling Depresses Unilateral Ureteral Obstruction-Evoked Mitochondrial Stress-Related Autophagy, Apoptosis and Pyroptosis in Kidney

PubMed Central

Chung, Shue Dong; Lai, Ting Yu; Chien, Chiang Ting; Yu, Hong Jen

2012-01-01

Exacerbated oxidative stress and inflammation may induce three types of programmed cell death, autophagy, apoptosis and pyroptosis in unilateral ureteral obstruction (UUO) kidney. Sulforaphane activating NF-E2-related nuclear factor erythroid-2 (Nrf-2) signaling may ameliorate UUO-induced renal damage. UUO was induced in the left kidney of female Wistar rats. The level of renal blood flow, cortical and medullary oxygen tension and reactive oxygen species (ROS) was evaluated. Fibrosis, ED-1 (macrophage/monocyte) infiltration, oxidative stress, autophagy, apoptosis and pyroptosis were evaluated by immunohistochemistry and Western blot in UUO kidneys. Effects of sulforaphane, an Nrf-2 activator, on Nrf-2- and mitochondrial stress-related proteins and renal injury were examined. UUO decreased renal blood flow and oxygen tension and increased renal ROS, 3-nitrotyrosine stain, ED-1 infiltration and fibrosis. Enhanced renal tubular Beclin-1 expression started at 4 h UUO and further enhanced at 3d UUO, whereas increased Atg-5-Atg12 and LC3-II expression were found at 3d UUO. Increased renal Bax/Bcl-2 ratio, caspase 3 and PARP fragments, apoptosis formation associated with increased caspase 1 and IL-1β expression for pyroptosis formation were started from 3d UUO. UUO reduced nuclear Nrf-2 translocation, increased cytosolic and inhibitory Nrf-2 expression, increased cytosolic Bax translocation to mitochondrial and enhanced mitochondrial Cytochrome c release into cytosol of the UUO kidneys. Sulforaphane significantly increased nuclear Nrf-2 translocation and decreased mitochondrial Bax translocation and Cytochrome c release into cytosol resulting in decreased renal injury. In conclusion, sulforaphane via activating Nrf-2 signaling preserved mitochondrial function and suppressed UUO-induced renal oxidative stress, inflammation, fibrosis, autophagy, apoptosis and pyroptosis. PMID:23071780

Simultaneous Increases in Proliferation and Apoptosis of Vascular Smooth Muscle Cells Accelerate Diabetic Mouse Venous Atherosclerosis

PubMed Central

Liu, Shuying; Zhang, Zhengyu; Wang, Jingjing; Zhou, Yuhuan; Liu, Kefeng; Huang, Jintao; Chen, Dadi; Wang, Junmei; Li, Chaohong

2015-01-01

Aims This study was designed to demonstrate simultaneous increases in proliferation and apoptosis of vascular smooth muscle cells (VSMCs) leading to accelerated vein graft remodeling and to explore the underlying mechanisms. Methods Vein grafts were performed in non-diabetic and diabetic mice. The cultured quiescent VSMCs were subjected to mechanical stretch stress (SS) and/or advanced glycosylation end products (AGEs). Harvested vein grafts and treated VSMCs were used to detect cell proliferation, apoptosis, mitogen-activated protein kinases (MAPKs) activation and SM-α-actin expression. Results Significantly thicker vessel walls and greater increases in proliferation and apoptosis were observed in diabetic vein grafts than those in non-diabetic. Both SS and AGEs were found to induce different activation of three members of MAPKs and simultaneous increases in proliferation and apoptosis of VSMCs, and combined treatment with both had a synergistic effect. VSMCs with strong SM-α-actin expression represented more activated JNKs or p38MAPK, and cell apoptosis, while the cells with weak SM-α-actin expression demonstrated preferential activation of ERKs and cell proliferation. In contrast, inhibition of MAPKs signals triggered significant decreases in VSMC proliferation, and apoptosis. Treatment of the cells with RNA interference of receptor of AGEs (RAGE) also resulted in significant decreases in both proliferation and apoptosis. Conclusions Increased pressure-induced SS triggers simultaneous increases in proliferation and apoptosis of VSMCs in the vein grafts leading to vein arterializations, which can be synergistically accelerated by high glucose-induced AGEs resulting in vein graft atherosclerosis. Either SS or AGEs and their combination induce simultaneous increases in proliferation and apoptosis of VSMCs via different activation of three members of MAPKs resulting from different VSMC subtypes classified by SM-α-actin expression levels. PMID:26488175

Age-Related Susceptibility to Apoptosis in Human Retinal Pigment Epithelial Cells Is Triggered by Disruption of p53–Mdm2 Association

PubMed Central

Bhattacharya, Sujoy; Chaum, Edward; Johnson, Dianna A.; Johnson, Leonard R.

2012-01-01

Purpose. Relatively little is known about the contribution of p53/Mdm2 pathway in apoptosis of retinal pigment epithelial (RPE) cells or its possible link to dysfunction of aging RPE or to related blinding disorders such as age-related macular degeneration (AMD). Methods. Age-associated changes in p53 activation were evaluated in primary RPE cultures from human donor eyes of various ages. Apoptosis was evaluated by activation of caspases and DNA fragmentation. Gene-specific small interfering RNA was used to knock down expression of p53. Results. We observed that the basal rate of p53-dependent apoptosis increased in an age-dependent manner in human RPE. The age-dependent increase in apoptosis was linked to alterations in several aspects of the p53 pathway. p53 phosphorylation Ser15 was increased through the stimulation of ATM-Ser1981. p53 acetylation Lys379 was increased through the inhibition of SIRT1/2. These two posttranslational modifications of p53 blocked the sequestration of p53 by Mdm2, thus resulting in an increase in free p53 and of p53 stimulation of apoptosis through increased expression of PUMA (p53 upregulated modulator of apoptosis) and activation of caspase-3. Aged RPE also had reduced expression of antiapoptotic Bcl-2, which contributed to the increase in apoptosis. Of particular interest in these studies was that pharmacologic treatments to block p53 phosphorylation, acetylation, or expression were able to protect RPE cells from apoptosis. Conclusions. Our studies suggest that aging in the RPE leads to alterations of specific checkpoints in the apoptotic pathway, which may represent important molecular targets for the treatment of RPE-related aging disorders such as AMD. PMID:23139272

Differential expression of microRNAs in omental adipose tissue from gestational diabetes mellitus subjects reveals miR-222 as a regulator of ERα expression in estrogen-induced insulin resistance.

PubMed

Shi, Zhonghua; Zhao, Chun; Guo, Xirong; Ding, Hongjuan; Cui, Yugui; Shen, Rong; Liu, Jiayin

2014-05-01

Omental adipose tissue plays a central role in insulin resistance in gestational diabetes mellitus (GDM), and the molecular mechanisms leading to GDM remains vague. Evidence demonstrates that maternal hormones, such as estradiol, contribute to insulin resistance in GDM. In this study we determined the differential expression patterns of microRNAs (miRNAs) in omental adipose tissues from GDM patients and pregnant women with normal glucose tolerance using AFFX miRNA expression chips. MiR-222, 1 of 17 identified differentially expressed miRNAs, was found to be significantly up-regulated in GDM by quantitative real-time PCR (P < .01), and its expression was closely related with serum estradiol level (P < .05). Furthermore, miR-222 expression was significantly increased in 3T3-L1 adipocytes with a high concentration of 17β-estradiol stimulation (P < .01), whereas the expressions of estrogen receptor (ER)-α protein and insulin-sensitive membrane transporter glucose transporter 4 (GLUT4) protein (P < .01) were markedly reduced. In addition, ERα was shown to be a direct target of miR-222 in 3T3-L1 adipocytes by using the luciferase assay. Finally, antisense oligonucleotides of miR-222 transfection was used to silence miR-222 in 3T3-L1 adipocytes. The results showed that the expressions of ERα and GLUT4, the insulin-stimulated translocation of GLUT4 from the cytoplasm to the cell membrane and glucose uptake in mature adipocytes were dramatically increased (P < .01). In conclusion, miR-222 is a potential regulator of ERα expression in estrogen-induced insulin resistance in GDM and might be a candidate biomarker and therapeutic target for GDM.

The gut microbiota reduces leptin sensitivity and the expression of the obesity-suppressing neuropeptides proglucagon (Gcg) and brain-derived neurotrophic factor (Bdnf) in the central nervous system.

PubMed

Schéle, Erik; Grahnemo, Louise; Anesten, Fredrik; Hallén, Anna; Bäckhed, Fredrik; Jansson, John-Olov

2013-10-01

The gut microbiota contributes to fat mass and the susceptibility to obesity. However, the underlying mechanisms are not completely understood. To investigate whether the gut microbiota affects hypothalamic and brainstem body fat-regulating circuits, we compared gene expression of food intake-regulating neuropeptides between germ-free and conventionally raised (CONV-R) mice. We found that CONV-R mice had decreased expression of the antiobesity neuropeptide glucagon-like peptide-1 (GLP-1) precursor proglucagon (Gcg) in the brainstem. Moreover, in both the hypothalamus and the brainstem, CONV-R mice had decreased expression of the antiobesity neuropeptide brain-derived neurotrophic factor (Bdnf). CONV-R mice had reduced expression of the pro-obesity peptides neuropeptide-Y (Npy) and agouti-related protein (Agrp), and increased expression of the antiobesity peptides proopiomelanocortin (Pomc) and cocaine- and amphetamine-regulated transcript (Cart) in the hypothalamus. The latter changes in neuropeptide expression could be secondary to elevated fat mass in CONV-R mice. Leptin treatment caused less weight reduction and less suppression of orexigenic Npy and Agrp expression in CONV-R mice compared with germ-free mice. The hypothalamic expression of leptin resistance-associated suppressor of cytokine signaling 3 (Socs-3) was increased in CONV-R mice. In conclusion, the gut microbiota reduces the expression of 2 genes coding for body fat-suppressing neuropeptides, Gcg and Bdnf, an alteration that may contribute to fat mass induction by the gut microbiota. Moreover, the presence of body fat-inducing gut microbiota is associated with hypothalamic signs of Socs-3-mediated leptin resistance, which may be linked to failed compensatory body fat reduction.

Hsp27 (HSPB1) differential expression in normal salivary glands and pleomorphic adenomas and association with an increased Bcl2/Bax ratio.

PubMed

Siqueira, Elisa C de; Souza, Fabrício T A; Diniz, Marina G; Gomez, Ricardo S; Gomes, Carolina C

2015-01-01

Pleomorphic adenoma (PA) is the most common salivary gland neoplasm. The Hsp27 (HSPB1) is an antiapoptotic protein whose synthesis follows cytotoxic stresses and result in a transient increase in tolerance to subsequent cell injury. Although Hsp27 is expressed in a range of normal tissues and neoplasms, a wide variation in its expression exists among different cells and tissues types. In certain tumours of glandular origin (such as oesophageal adenocarcinomas), the level of Hsp27 is decreased. In the present study, Hsp27 protein levels were evaluated by enzyme-linked immunosorbent assay (ELISA) in a set of 18 fresh PA and 12 normal salivary gland samples. In addition, we tested if Hsp27 protein levels correlated with p53 expression and cell proliferation index, as well as with the transcriptional levels of Bcl-2-associated X protein (BAX), B cell lymphoma 2 (BCL2) and Caspase 3 in PA. We further tested the association between Hsp27 expression and PA tumour size. While all normal salivary gland samples expressed Hsp27 protein, only half of the PA samples expressed it, resulting in a reduced expression of Hsp27 in PA when compared with normal salivary glands (P = 0.003). The expression levels of this protein correlated positively with a higher messenger ribonucleic acid (mRNA) ratio of Bcl2/Bax (R = 0.631; P = 0.01). In conclusion, a decreased Hsp27 protein expression level in PA was found. In addition, Hsp27 levels correlated positively with the Bcl2/Bax mRNA ratio, suggesting an antiapoptotic effect.

Baicalin increases VEGF expression and angiogenesis by activating the ERRα/PGC-1α pathway

PubMed Central

Zhang, Keqiang; Lu, Jianming; Mori, Taisuke; Smith-Powell, Leslie; Synold, Timothy W.; Chen, Shiuan; Wen, Wei

2011-01-01

Aims Baicalin is the major component found in Scutellaria baicalensis root, a widely used herb in traditional Chinese medicine. Although it has been used for thousands of years to treat stroke, the mechanisms of action of S. baicalensis have not been clearly elucidated. In this report, we studied the modulation of angiogenesis as one possible mechanism by investigating the effects of these agents on expression of vascular endothelial growth factor (VEGF), a critical factor for angiogenesis. Methods and results The effects of baicalin and an extract of S. baicalensis on VEGF expression were tested in several cell lines. Both agents induced VEGF expression in all cells without increasing expression of hypoxia-inducible factor-1α (HIF-1α). The expression of reporter genes was also activated under the control of the VEGF promoter containing either a functional or a defective HIF response element (HRE). Only minimal effects were observed on reporter activation under the HRE promoter. Instead, both agents significantly induced oestrogen-related receptor (ERRα) expression as well as the activity of reporter genes under the control of ERRα-binding element. Their ability to induce VEGF expression was suppressed once ERRα expression was knocked down by siRNA or ERRα-binding sites were deleted in the VEGF promoter. We also found that both agents stimulated cell migration and vessel sprout formation from the aorta. Conclusion Our results implicate baicalin and S. baicalensis in angiogenesis by inducing VEGF expression through the activation of the ERRα pathway. These data may facilitate a better understanding of the potential health benefits of these agents in the treatment of cardiovascular diseases. PMID:20851810

Corticosteroid-induced gene expression in allergen-challenged asthmatic subjects taking inhaled budesonide

PubMed Central

Kelly, MM; King, EM; Rider, CF; Gwozd, C; Holden, NS; Eddleston, J; Zuraw, B; Leigh, R; O'Byrne, PM; Newton, R

2012-01-01

BACKGROUND AND PURPOSE Inhaled corticosteroids (ICS) are the cornerstone of asthma pharmacotherapy and, acting via the glucocorticoid receptor (GR), reduce inflammatory gene expression. While this is often attributed to a direct inhibitory effect of the GR on inflammatory gene transcription, corticosteroids also induce the expression of anti-inflammatory genes in vitro. As there are no data to support this effect in asthmatic subjects taking ICS, we have assessed whether ICS induce anti-inflammatory gene expression in subjects with atopic asthma. EXPERIMENTAL APPROACH Bronchial biopsies from allergen-challenged atopic asthmatic subjects taking inhaled budesonide or placebo were subjected to gene expression analysis using real-time reverse transcriptase-PCR for the corticosteroid-inducible genes (official gene symbols with aliases in parentheses): TSC22D3 [glucocorticoid-induced leucine zipper (GILZ)], dual-specificity phosphatase-1 (MAPK phosphatase-1), both anti-inflammatory effectors, and FKBP5 [FK506-binding protein 51 (FKBP51)], a regulator of GR function. Cultured pulmonary epithelial and smooth muscle cells were also treated with corticosteroids before gene expression analysis. KEY RESULTS Compared with placebo, GILZ and FKBP51 mRNA expression was significantly elevated in budesonide-treated subjects. Budesonide also increased GILZ expression in human epithelial and smooth muscle cells in culture. Immunostaining of bronchial biopsies revealed GILZ expression in the airways epithelium and smooth muscle of asthmatic subjects. CONCLUSIONS AND IMPLICATIONS Expression of the corticosteroid-induced genes, GILZ and FKBP51, is up-regulated in the airways of allergen-challenged asthmatic subjects taking inhaled budesonide. Consequently, the biological effects of corticosteroid-induced genes should be considered when assessing the actions of ICS. Treatment modalities that increase or decrease GR-dependent transcription may correspondingly affect corticosteroid efficacy. PMID:21827450

Expression of Y-box-binding protein YB-1 allows stratification into long- and short-term survivors of head and neck cancer patients

PubMed Central

Kolk, A; Jubitz, N; Mengele, K; Mantwill, K; Bissinger, O; Schmitt, M; Kremer, M; Holm, P S

2011-01-01

Background: Histology-based classifications and clinical parameters of head and neck squamous cell carcinoma (HNSCC) are limited in their clinical capacity to provide information on prognosis and treatment choice of HNSCC. The primary aim of this study was to analyse Y-box-binding protein-1 (YB-1) protein expression in different grading groups of HNSCC patients, and to correlate these findings with the disease-specific survival (DSS). Methods: We investigated the expression and cellular localisation of the oncogenic transcription/translation factor YB-1 by immunohistochemistry on tissue micro arrays in a total of 365 HNSCC specimens and correlated expression data with clinico-pathological parameters including DSS. Results: Compared with control tissue from healthy individuals, a significantly (P<0.01) increased YB-1 protein expression was observed in high-grade HNSCC patients. By univariate survival data analysis, HNSCC patients with elevated YB-1 protein expression had a significantly (P<0.01) decreased DSS. By multivariate Cox regression analysis, high YB-1 expression and nuclear localisation retained its significance as a statistically independent (P<0.002) prognostic marker for DSS. Within grade 2 group of HNSCC patients, a subgroup defined by high nuclear and cytoplasmic YB-1 levels (co-expression pattern) in the cells of the tumour invasion front had a significantly poorer 5-year DSS rate of only 38% compared with overall 55% for grade 2 patients. Vice versa, the DSS rate was markedly increased to 74% for grade 2 cancer patients with low YB-1 protein expression at the same localisation. Conclusion: Our findings point to the fact that YB-1 expression in combination with histological classification in a double stratification strategy is superior to classical grading in the prediction of tumour progression in HNSCC. PMID:22095225

Beneficial Effects of cART Initiated during Primary and Chronic HIV-1 Infection on Immunoglobulin-Expression of Memory B-Cell Subsets

PubMed Central

Pensieroso, Simone; Tolazzi, Monica; Chiappetta, Stefania; Nozza, Silvia; Lazzarin, Adriano; Tambussi, Giuseppe; Scarlatti, Gabriella

2015-01-01

Introduction During HIV-1 infection the B-cell compartment undergoes profound changes towards terminal differentiation, which are only partially restored by antiretroviral therapy (cART). Materials and Methods To investigate the impact of infection as early as during primary HIV-1 infection (PHI) we assessed distribution of B-cell subsets in 19 PHI and 25 chronic HIV-1-infected (CHI) individuals before and during 48 weeks of cART as compared to healthy controls (n = 23). We also analysed Immunoglobulin-expression of memory B-cell subsets to identify alterations in Immunoglobulin-maturation. Results Determination of B-cell subsets at baseline showed that total and Naive B-cells were decreased whereas Activated Memory (AM), Tissue-like Memory (TLM) B-cells and Plasma cells were increased in both PHI and CHI patients. After 4 weeks of cART total B-cells increased, while AM, TLM B-cells and Plasma cells decreased, although without reaching normal levels in either group of individuals. This trend was maintained until week 48, though only total B-cells normalized in both PHI and CHI. Resting Memory (RM) B-cells were preserved since baseline. This subset remained stable in CHI, while was expanded by an early initiation of cART during PHI. Untreated CHI patients showed IgM-overexpression at the expenses of switched (IgM-IgD-) phenotypes of the memory subsets. Interestingly, in PHI patients a significant alteration of Immunoglobulin-expression was evident at BL in TLM cells, and after 4 weeks, despite treatment, in AM and RM subsets. After 48 weeks of therapy, Immunoglobulin-expression of AM and RM almost normalized, but remained perturbed in TLM cells in both groups. Conclusions In conclusion, aberrant activated and exhausted B-cell phenotypes rose already during PHI, while most of the alterations in Ig-expression seen in CHI appeared later, despite 4 weeks of effective cART. After 48 weeks of cART B-cell subsets distribution improved although without full normalization, while Immunoglobulin-expression normalized among AM and RM, remaining perturbed in TLM B-cells of PHI and CHI. PMID:26474181

Overexpression of SerpinE2/protease nexin-1 Contribute to Pathological Cardiac Fibrosis via increasing Collagen Deposition

PubMed Central

Li, Xuelian; Zhao, Dandan; Guo, Zhenfeng; Li, Tianshi; Qili, Muge; Xu, Bozhi; Qian, Ming; Liang, Haihai; E, Xiaoqiang; Chege Gitau, Samuel; Wang, Lu; Huangfu, Longtao; Wu, Qiuxia; Xu, Chaoqian; Shan, Hongli

2016-01-01

Although increases in cardiovascular load (pressure overload) are known to elicit ventricular remodeling including cardiomyocyte hypertrophy and interstitial fibrosis, the molecular mechanisms of pressure overload or AngII -induced cardiac interstitial fibrosis remain elusive. In this study, serpinE2/protease nexin-1 was over-expressed in a cardiac fibrosis model induced by pressure-overloaded via transverse aortic constriction (TAC) in mouse. Knockdown of serpinE2 attenuates cardiac fibrosis in a mouse model of TAC. At meantime, the results showed that serpinE2 significantly were increased with collagen accumulations induced by AngII or TGF-β stimulation in vitro. Intriguingly, extracellular collagen in myocardial fibroblast was reduced by knockdown of serpinE2 compared with the control in vitro. In stark contrast, the addition of exogenous PN-1 up-regulated the content of collagen in myocardial fibroblast. The MEK1/2- ERK1/2 signaling probably promoted the expression of serpinE2 via transcription factors Elk1 in myocardial fibroblast. In conclusion, stress-induced the ERK1/2 signaling pathway activation up-regulated serpinE2 expression, consequently led accumulation of collagen protein, and contributed to cardiac fibrosis. PMID:27876880

Carvacrol Exerts Neuroprotective Effects Via Suppression of the Inflammatory Response in Middle Cerebral Artery Occlusion Rats.

PubMed

Li, Zhenlan; Hua, Cong; Pan, Xiaoqiang; Fu, Xijia; Wu, Wei

2016-08-01

Increasing evidence demonstrates that inflammation plays an important role in cerebral ischemia. Carvacrol, a monoterpenic phenol, is naturally occurring in various plants belonging to the family Lamiaceae and exerts protective effects in a mice model of focal cerebral ischemia/reperfusion injury by reducing infarct volume and decreasing the expression of cleaved caspase-3. However, the anti-inflammatory mechanisms by which carvacrol protect the brain have yet to be fully elucidated. We investigated the effects of carvacrol on inflammatory reaction and inflammatory mediators in middle cerebral artery occlusion rats. The results of the present study showed that carvacrol inhibited the levels of inflammatory cytokines and myeloperoxidase (MPO) activity, as well as the expression of iNOS and COX-2. It also increased SOD activity and decreased MDA level in ischemic cortical tissues. In addition, carvacrol treatment suppressed the ischemia/reperfusion-induced increase in the protein expression of nuclear NF-kB p65. In conclusion, we have shown that carvacrol inhibits the inflammatory response via inhibition of the NF-kB signaling pathway in a rat model of focal cerebral ischemia. Therefore, carvacrol may be a potential therapeutic agent for the treatment of cerebral ischemia injury.

Inflamed In Vitro Retina: Cytotoxic Neuroinflammation and Galectin-3 Expression

PubMed Central

Bauer, Patrik Maximilian; Zalis, Marina Castro; Abdshill, Hodan; Deierborg, Tomas; Johansson, Fredrik; Englund-Johansson, Ulrica

2016-01-01

Background Disease progression in retinal neurodegeneration is strongly correlated to immune cell activation, which may have either a neuroprotective or neurotoxic effect. Increased knowledge about the immune response profile and retinal neurodegeneration may lead to candidate targets for treatments. Therefore, we have used the explanted retina as a model to explore the immune response and expression of the immune modulator galectin-3 (Gal-3), induced by the cultivation per se and after additional immune stimulation with lipopolysaccharide (LPS), and how this correlates with retinal neurotoxicity. Methods Post-natal mouse retinas were cultured in a defined medium. One group was stimulated with LPS (100 ng/ml, 24 h). Retinal architecture, apoptotic cell death, and micro- and macroglial activity were studied at the time of cultivation (0 days in vitro (DIV)) and at 3, 4 and 7 DIV using morphological staining, biochemical- and immunohistochemical techniques. Results Our results show that sustained activation of macro- and microglia, characterized by no detectable cytokine release and limited expression of Gal-3, is not further inducing apoptosis additional to the axotomy-induced apoptosis in innermost nuclear layer. An elevated immune response was detected after LPS stimulation, as demonstrated primarily by release of immune mediators (i.e. interleukin 2 (IL-2), IL-6, KC/GRO (also known as CLCX1) and tumour necrosis factor-α (TNF-α)), increased numbers of microglia displaying morphologies of late activation stages as well as Gal-3 expression. This was accompanied with increased apoptosis in the two additional nuclear layers, and damage to retinal gross architecture. Conclusion We demonstrate that an immune response characterized by sustained and increased release of cytokines, along with an increase in Gal-3 expression, is accompanied by significant increased neurotoxicity in the explanted retina. Further investigations using the current setting may lead to increased understanding on the mechanisms involved in neuronal loss in retinal neurodegenerations. PMID:27612287

Increased expression of the yeast multidrug resistance ABC transporter Pdr18 leads to increased ethanol tolerance and ethanol production in high gravity alcoholic fermentation

PubMed Central

2012-01-01

Background The understanding of the molecular basis of yeast tolerance to ethanol may guide the design of rational strategies to increase process performance in industrial alcoholic fermentations. A set of 21 genes encoding multidrug transporters from the ATP-Binding Cassette (ABC) Superfamily and Major Facilitator Superfamily (MFS) in S. cerevisiae were scrutinized for a role in ethanol stress resistance. Results A yeast multidrug resistance ABC transporter encoded by the PDR18 gene, proposed to play a role in the incorporation of ergosterol in the yeast plasma membrane, was found to confer resistance to growth inhibitory concentrations of ethanol. PDR18 expression was seen to contribute to decreased 3 H-ethanol intracellular concentrations and decreased plasma membrane permeabilization of yeast cells challenged with inhibitory ethanol concentrations. Given the increased tolerance to ethanol of cells expressing PDR18, the final concentration of ethanol produced during high gravity alcoholic fermentation by yeast cells devoid of PDR18 was lower than the final ethanol concentration produced by the corresponding parental strain. Moreover, an engineered yeast strain in which the PDR18 promoter was replaced in the genome by the stronger PDR5 promoter, leading to increased PDR18 mRNA levels during alcoholic fermentation, was able to attain a 6 % higher ethanol concentration and a 17 % higher ethanol production yield than the parental strain. The improved fermentative performance of yeast cells over-expressing PDR18 was found to correlate with their increased ethanol tolerance and ability to restrain plasma membrane permeabilization induced throughout high gravity fermentation. Conclusions PDR18 gene over-expression increases yeast ethanol tolerance and fermentation performance leading to the production of highly inhibitory concentrations of ethanol. PDR18 overexpression in industrial yeast strains appears to be a promising approach to improve alcoholic fermentation performance for sustainable bio-ethanol production. PMID:22839110

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.

Effects of PCB 126 and PCB 153 on secretion of steroid hormones and mRNA expression of steroidogenic genes (STAR, HSD3B, CYP19A1) and estrogen receptors (ERα, ERβ) in prehierarchical chicken ovarian follicles.

PubMed

Sechman, Andrzej; Batoryna, Marta; Antos, Piotr A; Hrabia, Anna

2016-12-15

The objective of this study was to assess the in vitro effects of dioxin-like PCB 126 and non-dioxin-like PCB 153 on basal and ovine LH (oLH)-stimulated testosterone (T) and estradiol (E2) secretion and expression of steroidogenic genes (STAR, HSD3B and CYP19A1) and estrogen receptors α (ERα) and β (ERβ) in white (WF) and yellowish (YF) prehierarchical follicles of the hen ovary. Steroid concentrations in a medium and gene expression in follicles following 6h of exposition were determined by RIA and real-time qPCR, respectively. Both PCBs increased basal and oLH-stimulated T secretion by the WF follicles. PCB 126 reduced basal E2 secretion by the WF follicles. PCB 153 elevated but PCB 126 reduced oLH-stimulated E2 secretion by the prehierarchical follicles. PCB 126 increased basal STAR and HSD3B and reduced CYP19A1 mRNA expression in these follicles. PCB 153 increased basal expression of STAR and HSD3B in YF follicles, but diminished HSD3B mRNA levels in the WF. The studied PCBs had an opposite effect on basal and oLH-stimulated CYP19A1 mRNA expression in prehierarchical follicles. Both PCBs modulated basal and inhibited oLH-stimulated ERα and ERβ gene expression in the prehierarchical follicles. In conclusion, data of the current study demonstrate the congener-specific effects of PCBs on sex steroid secretion by prehierarchical follicles of the chicken ovary, which are at least partly related to STAR, HSD3B and CYP19A1 gene expression. It is suggested that PCBs, by influencing follicular steroidogenesis and expression of estrogen receptors, may impair development and selection of yellowish follicles to the preovulatory hierarchy. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

Endothelial Activation by Platelets from Sickle Cell Anemia Patients

PubMed Central

Proença-Ferreira, Renata; Brugnerotto, Ana Flávia; Garrido, Vanessa Tonin; Dominical, Venina Marcela; Vital, Daiana Morelli; Ribeiro, Marilene de Fátima Reis; dos Santos, Melissa Ercolin; Traina, Fabíola; Olalla-Saad, Sara T.; Costa, Fernando Ferreira; Conran, Nicola

2014-01-01

Sickle cell anemia (SCA) is associated with a hypercoagulable state. Increased platelet activation is reported in SCA and SCA platelets may present augmented adhesion to the vascular endothelium, potentially contributing to the vaso-occlusive process. We sought to observe the effects of platelets (PLTs) from healthy control (CON) individuals and SCA individuals on endothelial activation, in vitro. Human umbilical vein endothelial cells (HUVEC) were cultured, in the presence, or not, of washed PLTs from CON or steady-state SCA individuals. Supernatants were reserved for cytokine quantification, and endothelial adhesion molecules (EAM) were analyzed by flow cytometry; gene expressions of ICAM1 and genes of the NF-κB pathway were analyzed by qPCR. SCA PLTs were found to be more inflammatory, displaying increased adhesive properties, an increased production of IL-1β and a tendency towards elevated expressions of P-selectin and activated αIIbβ3. Following culture in the presence of SCA PLTs, HUVEC presented significant augmentations in the expressions of the EAM, ICAM-1 and E-selectin, as well as increased IL-8 production and increased ICAM1 and NFKB1 (encodes p50 subunit of NF-κB) gene expressions. Interestingly, transwell inserts abolished the effects of SCA PLTs on EAM expression. Furthermore, an inhibitor of the NF-κB pathway, BAY 11-7082, also prevented the induction of EAM expression on the HUVEC surface by SCA PLTs. In conclusion, we find further evidence to indicate that platelets circulate in an activated state in sickle cell disease and are capable of stimulating endothelial cell activation. This effect appears to be mediated by direct contact, or even adhesion, between the platelets and endothelial cells and via NFκB-dependent signaling. As such, activated platelets in SCD may contribute to endothelial activation and, therefore, to the vaso-occlusive process. Results provide further evidence to support the use of anti-platelet approaches in association with other therapies for SCD. PMID:24551209

Nitric Oxide Increases Arterial Endotheial Permeability through Mediating VE-Cadherin Expression during Arteriogenesis.

PubMed

Yang, Baolin; Cai, Baizhen; Deng, Panyue; Wu, Xiaoqiong; Guan, Yinglu; Zhang, Bin; Cai, Weijun; Schaper, Jutta; Schaper, Wolfgang

2015-01-01

Macrophage invasion is an important event during arteriogenesis, but the underlying mechanism is still only partially understood. The present study tested the hypothesis that nitric oxide (NO) and VE-cadherin, two key mediators for vascular permeability, contribute to this event in a rat ischemic hindlimb model. In addition, the effect of NO on expression of VE-caherin and endothelial permeability was also studied in cultured HUVECs. We found that: 1) in normal arteriolar vessels (NAV), eNOS was moderately expressed in endothelial cells (EC) and iNOS was rarely detected. In contrast, in collateral vessels (CVs) induced by simple femoral artery ligation, both eNOS and iNOS were significantly upregulated (P<0.05). Induced iNOS was found mainly in smooth muscle cells, but also in other vascular cells and macrophages; 2) in NAV VE-cadherin was strongly expressed in EC. In CVs, VE-cadherin was significantly downregulated, with a discontinuous and punctate pattern. Administration of nitric oxide donor DETA NONOate (NONOate) further reduced the amounts of Ve-cadherin in CVs, whereas NO synthase inhibitor L-NAME inhibited downregulation of VE-cadherin in CVs; 3) in normal rats Evans blue extravasation (EBE) was low in the musculus gracilis, FITC-dextron leakage was not detected in the vascular wall and few macrophages were observed in perivascular space. In contrast, EBE was significantly increased in femoral artery ligation rats, FITC-dextron leakage and increased amounts of macrophages were detected in CVs, which were further enhanced by administration of NONOate, but inhibited by L-NAME supplement; 4) in vitro experiments confirmed that an increase in NO production reduced VE-cadherin expression, correlated with increases in the permeability of HUVECs. In conclusion, our data for the first time reveal the expression profile of VE-cadherin and alterations of vascular permeability in CVs, suggesting that NO-mediated VE-cadherin pathway may be one important mechanism responsible, at least in part, for macrophage invasion during arteriogenesis.

Nitric Oxide Increases Arterial Endotheial Permeability through Mediating VE-Cadherin Expression during Arteriogenesis

PubMed Central

Wu, Xiaoqiong; Guan, Yinglu; Zhang, Bin; Cai, Weijun; Schaper, Jutta; Schaper, Wolfgang

2015-01-01

Macrophage invasion is an important event during arteriogenesis, but the underlying mechanism is still only partially understood. The present study tested the hypothesis that nitric oxide (NO) and VE-cadherin, two key mediators for vascular permeability, contribute to this event in a rat ischemic hindlimb model. In addition, the effect of NO on expression of VE-caherin and endothelial permeability was also studied in cultured HUVECs. We found that: 1) in normal arteriolar vessels (NAV), eNOS was moderately expressed in endothelial cells (EC) and iNOS was rarely detected. In contrast, in collateral vessels (CVs) induced by simple femoral artery ligation, both eNOS and iNOS were significantly upregulated (P<0.05). Induced iNOS was found mainly in smooth muscle cells, but also in other vascular cells and macrophages; 2) in NAV VE-cadherin was strongly expressed in EC. In CVs, VE-cadherin was significantly downregulated, with a discontinuous and punctate pattern. Administration of nitric oxide donor DETA NONOate (NONOate) further reduced the amounts of Ve-cadherin in CVs, whereas NO synthase inhibitor L-NAME inhibited downregulation of VE-cadherin in CVs; 3) in normal rats Evans blue extravasation (EBE) was low in the musculus gracilis, FITC-dextron leakage was not detected in the vascular wall and few macrophages were observed in perivascular space. In contrast, EBE was significantly increased in femoral artery ligation rats, FITC-dextron leakage and increased amounts of macrophages were detected in CVs, which were further enhanced by administration of NONOate, but inhibited by L-NAME supplement; 4) in vitro experiments confirmed that an increase in NO production reduced VE-cadherin expression, correlated with increases in the permeability of HUVECs. In conclusion, our data for the first time reveal the expression profile of VE-cadherin and alterations of vascular permeability in CVs, suggesting that NO-mediated VE-cadherin pathway may be one important mechanism responsible, at least in part, for macrophage invasion during arteriogenesis. PMID:26133549

Fetal growth restriction and the programming of heart growth and cardiac insulin-like growth factor 2 expression in the lamb.

PubMed

Wang, Kimberley C W; Zhang, Lei; McMillen, I Caroline; Botting, Kimberley J; Duffield, Jaime A; Zhang, Song; Suter, Catherine M; Brooks, Doug A; Morrison, Janna L

2011-10-01

Reduced growth in fetal life together with accelerated growth in childhood, results in a ~50% greater risk of coronary heart disease in adult life. It is unclear why changes in patterns of body and heart growth in early life can lead to an increased risk of cardiovascular disease in adulthood. We aimed to investigate the role of the insulin-like growth factors in heart growth in the growth-restricted fetus and lamb. Hearts were collected from control and placentally restricted (PR) fetuses at 137-144 days gestation and from average (ABW) and low (LBW) birth weight lambs at 21 days of age. We quantified cardiac mRNA expression of IGF-1, IGF-2 and their receptors, IGF-1R and IGF-2R, using real-time RT-PCR and protein expression of IGF-1R and IGF-2R using Western blotting. Combined bisulphite restriction analysis was used to assess DNA methylation in the differentially methylated region (DMR) of the IGF-2/H19 locus and of the IGF-2R gene. In PR fetal sheep, IGF-2, IGF-1R and IGF-2R mRNA expression was increased in the heart compared to controls. LBW lambs had a greater left ventricle weight relative to body weight as well as increased IGF-2 and IGF-2R mRNA expression in the heart, when compared to ABW lambs. No changes in the percentage of methylation of the DMRs of IGF-2/H19 or IGF-2R were found between PR and LBW when compared to their respective controls. In conclusion, a programmed increased in cardiac gene expression of IGF-2 and IGF-2R may represent an adaptive response to reduced substrate supply (e.g. glucose and/or oxygen) in order to maintain heart growth and may be the underlying cause for increased ventricular hypertrophy and the associated susceptibility of cardiomyocytes to ischaemic damage later in life.

Chronic effects of soft drink consumption on the health state of Wistar rats: A biochemical, genetic and histopathological study.

PubMed

Alkhedaide, Adel; Soliman, Mohamed Mohamed; Salah-Eldin, Alaa-Eldin; Ismail, Tamer Ahmed; Alshehiri, Zafer Saad; Attia, Hossam Fouad

2016-06-01

The present study was performed to examine the effects of chronic soft drink consumption (SDC) on oxidative stress, biochemical alterations, gene biomarkers and histopathology of bone, liver and kidney. Free drinking water of adult male Wistar rats was substituted with three different soft drinks: Coca‑Cola, Pepsi and 7‑Up, for three consecutive months. The serum and organs were collected for examining the biochemical parameters associated with bone, liver and kidney functions. Semi‑quantitative reverse transcription polymerase chain reaction was used to observe the changes in the expression of genes in the liver and kidney, which are associated with oxidative stress resistance. Histopathological investigations were performed to determine the changes in bone, liver and kidney tissues using hematoxylin and eosin stains. SDC affected liver, kidney and bone function biomarkers. Soft drinks increased oxidative stress, which is represented by an increase in malondialdehyde and a decrease in antioxidant levels. SDC affected serum mineral levels, particularly calcium and phosphorus. Soft drinks downregulated the expression levels of glutathione‑S‑transferase and super oxide dismutase in the liver compared with that of control rats. Rats administered Coca‑Cola exhibited a hepatic decrease in the mRNA expression of α2‑macroglobulin compared with rats administered Pepsi and 7‑Up. On the other hand, SDC increased the mRNA expression of α1‑acid glycoprotein. The present renal studies revealed that Coca‑Cola increased the mRNA expression levels of desmin, angiotensinogen and angiotensinogen receptor compared with the other groups, together with mild congestion in renal histopathology. Deleterious histopathological changes were reported predominantly in the bone and liver of the Coca‑Cola and Pepsi groups. In conclusion, a very strict caution must be considered with SDC due to the increase in oxidative stress biomarkers and disruption in the expression of certain genes associated with the bio‑vital function of both the liver and kidney.

Inhibition of Mdm2 Sensitizes Human Retinal Pigment Epithelial Cells to Apoptosis

PubMed Central

Ray, Ramesh M.; Chaum, Edward; Johnson, Dianna A.; Johnson, Leonard R.

2011-01-01

Purpose. Because recent studies indicate that blocking the interaction between p53 and Mdm2 results in the nongenotoxic activation of p53, the authors sought to investigate whether the inhibition of p53-Mdm2 binding activates p53 and sensitizes human retinal epithelial cells to apoptosis. Methods. Apoptosis was evaluated by the activation of caspases and DNA fragmentation assays. The Mdm2 antagonist Nutlin-3 was used to dissociate p53 from Mdm2 and, thus, to increase p53 activity. Knockdown of p53 expression was accomplished by using p53 siRNA. Results. ARPE-19 and primary RPE cells expressed high levels of the antiapoptotic proteins Bcl-2 and Bcl-xL. Exposure of these cells to camptothecin (CPT) or TNF-α/ cycloheximide (CHX) failed to induce apoptosis. In contrast, treatment with the Mdm2 antagonist Nutlin-3 in the absence of CPT or TNF-α/CHX increased apoptosis. Activation of p53 in response to Nutlin-3 also increased levels of Noxa, p53-upregulated modulator of apoptosis (PUMA), and Siva-1, decreased expression of Bcl-2 and Bcl-xL, and simultaneously increased caspases-9 and -3 activities and DNA fragmentation. Knockdown of p53 decreased the basal expression of p21Cip1 and Bcl-2, inhibited the Nutlin-3–induced upregulation of Siva-1 and PUMA expression, and consequently inhibited caspase-3 activation. Conclusions. These results indicate that the normally available pool of intracellular p53 is predominantly engaged in the regulation of cell cycle checkpoints by p21Cip1 and does not trigger apoptosis in response to DNA-damaging agents. However, the blockage of p53 binding to Mdm2 frees a pool of p53 that is sufficient, even in the absence of DNA-damaging agents, to increase the expression of proapoptotic targets and to override the resistance of RPE cells to apoptosis. PMID:21345989

Chronic effects of soft drink consumption on the health state of Wistar rats: A biochemical, genetic and histopathological study

PubMed Central

ALKHEDAIDE, ADEL; SOLIMAN, MOHAMED MOHAMED; SALAH-ELDIN, ALAA-ELDIN; ISMAIL, TAMER AHMED; ALSHEHIRI, ZAFER SAAD; ATTIA, HOSSAM FOUAD

2016-01-01

The present study was performed to examine the effects of chronic soft drink consumption (SDC) on oxidative stress, biochemical alterations, gene biomarkers and histopathology of bone, liver and kidney. Free drinking water of adult male Wistar rats was substituted with three different soft drinks: Coca-Cola, Pepsi and 7-Up, for three consecutive months. The serum and organs were collected for examining the biochemical parameters associated with bone, liver and kidney functions. Semi-quantitative reverse transcription polymerase chain reaction was used to observe the changes in the expression of genes in the liver and kidney, which are associated with oxidative stress resistance. Histopathological investigations were performed to determine the changes in bone, liver and kidney tissues using hematoxylin and eosin stains. SDC affected liver, kidney and bone function biomarkers. Soft drinks increased oxidative stress, which is represented by an increase in malondialdehyde and a decrease in antioxidant levels. SDC affected serum mineral levels, particularly calcium and phosphorus. Soft drinks downregulated the expression levels of glutathione-S-transferase and super oxide dismutase in the liver compared with that of control rats. Rats administered Coca-Cola exhibited a hepatic decrease in the mRNA expression of α2-macroglobulin compared with rats administered Pepsi and 7-Up. On the other hand, SDC increased the mRNA expression of α1-acid glycoprotein. The present renal studies revealed that Coca-Cola increased the mRNA expression levels of desmin, angiotensinogen and angiotensinogen receptor compared with the other groups, together with mild congestion in renal histopathology. Deleterious histopathological changes were reported predominantly in the bone and liver of the Coca-Cola and Pepsi groups. In conclusion, a very strict caution must be considered with SDC due to the increase in oxidative stress biomarkers and disruption in the expression of certain genes associated with the bio-vital function of both the liver and kidney. PMID:27121771

Glutathione S-Transferase Pi Isoform (GSTP1) Expression in Murine Retina Increases with Developmental Maturity

PubMed Central

Lee, Wen-Hsiang; Joshi, Pratibha; Wen, Rong

2014-01-01

Background and Aims Glutathione S-transferase pi isoform (GSTP1) is an intracellular detoxification enzyme that catalyzes reduction of chemically reactive electrophiles and is a zeaxanthin-binding protein in the human macula. We have previously demonstrated that GSTP1 levels are decreased in human age-related macular degeneration (AMD) retina compared to normal controls [1]. We also showed that GSTP1 levels parallel survival of human retinal pigment epithelial (RPE) cells exposed to UV light, and GSTP1 over-expression protects them against UV light damage [2]. In the present work, we determined the developmental time course of GSTP1 expression in murine retina and in response to light challenge. Methods Eyes from BALB/c mice at post-natal day 20, 1 month, and 2 months of age were prepared for retinal protein extraction and cryo sectioning, and GSTP1 levels in the retina were analyzed by Western blot and immunohistochemistry (IHC). Another group of BALB/c mice with the same age ranges was exposed to 1000 lux of white fluorescent light for 24 hours, and their retinas were analyzed for GSTP1 expression by Western blot and IHC in a similar manner. Results GSTP1 levels in the murine retina increased in ascending order from post-natal day 20, 1 month, and 2 months of age. Moreover, GSTP1 expression in murine retina at post-natal day 20, 1 month, and 2 months of age increased in response to brief light exposure compared to age-matched controls under normal condition. Conclusions GSTP1 expression in retina increases with developmental age in mice and accompanies murine retinal maturation. Brief exposure to light induces GSTP1 expression in the murine retina across various developmental ages. GSTP1 induction may be a protective response to light-induced oxidative damage in the murine retina. PMID:24664677

SMAD-PI3K-Akt-mTOR Pathway Mediates BMP-7 Polarization of Monocytes into M2 Macrophages

PubMed Central

Rocher, Crystal; Singla, Dinender K.

2013-01-01

Previously we demonstrated that bone morphogenetic protein-7 (BMP-7) treatment polarizes monocytes into M2 macrophages and increases the expression of anti-inflammatory cytokines. Despite these findings, the mechanisms for the observed BMP-7 induced monocyte polarization into M2 macrophages are completely unknown. In this study, we demonstrate the mechanisms involved in the polarization of monocytes into M2 macrophages. Apoptotic conditioned media (ACM) was generated to mimic the stressed conditions, inducing monocyte polarization. Monocytes were treated with ACM along with BMP-7 and/or its inhibitor, follistatin, for 48 hours. Furthermore, an inhibitor of the PI3K pathway, LY-294002, was also studied. Our data show that BMP-7 induces polarization of monocytes into M2 macrophages while significantly increasing the expression of anti-inflammatory markers, arginase-1 and IL-10, and significantly (p<0.05) decreasing the expression of pro-inflammatory markers iNOS, IL-6, TNF-α and MCP-1; (p<0.05). Moreover, addition of the PI3K inhibitor, LY-294002, significantly (p<0.05) decreases upregulation of IL-10 and arginase-1, suggesting involvement of the PI3K pathway in M2 macrophage polarization. Next, following BMP-7 treatment, a significant (p<0.05) increase in p-SMAD1/5/8 and p-PI3K expression resulting in downstream activation of p-Akt and p-mTOR was observed. Furthermore, expression of p-PTEN, an inhibitor of the PI3K pathway, was significantly (p<0.05) increased in the ACM group. However, BMP-7 treatment inhibited its expression, suggesting involvement of the PI3K-Akt-mTOR pathway. In conclusion, we demonstrate that BMP-7 polarizes monocytes into M2 macrophages and enhances anti-inflammatory cytokine expression which is mediated by the activated SMAD-PI3K-Akt-mTOR pathway. PMID:24376781

Perinatally administered losartan augments renal ACE2 expression but not cardiac or renal Mas receptor in spontaneously hypertensive rats.

PubMed

Klimas, Jan; Olvedy, Michael; Ochodnicka-Mackovicova, Katarina; Kruzliak, Peter; Cacanyiova, Sona; Kristek, Frantisek; Krenek, Peter; Ochodnicky, Peter

2015-08-01

Since the identification of the alternative angiotensin converting enzyme (ACE)2/Ang-(1-7)/Mas receptor axis, renin-angiotensin system (RAS) is a new complex target for a pharmacological intervention. We investigated the expression of RAS components in the heart and kidney during the development of hypertension and its perinatal treatment with losartan in young spontaneously hypertensive rats (SHR). Expressions of RAS genes were studied by the RT-PCR in the left ventricle and kidney of rats: normotensive Wistar, untreated SHR, SHR treated with losartan since perinatal period until week 9 of age (20 mg/kg/day) and SHR treated with losartan only until week 4 of age and discontinued until week 9. In the hypertrophied left ventricle of SHR, cardiac expressions of Ace and Mas were decreased while those of AT1 receptor (Agtr1a) and Ace2 were unchanged. Continuous losartan administration reduced LV weight (0.43 ± 0.02; P < 0.05 versus SHR) but did not influence altered cardiac RAS expression. Increased blood pressure in SHR (149 ± 2 in SHR versus 109 ± 2 mmHg in Wistar; P < 0.05) was associated with a lower renal expressions of renin, Agtr1a and Mas and with an increase in ACE2. Continuous losartan administration lowered blood pressure to control levels (105 ± 3 mmHg; P < 0.05 versus SHR), however, only renal renin and ACE2 were significantly up-regulated (for both P < 0.05 versus SHR). Conclusively, prevention of hypertension and LV hypertrophy development by losartan was unrelated to cardiac or renal expression of Mas. Increased renal Ace2, and its further increase by losartan suggests the influence of locally generated Ang-(1-7) in organ response to the developing hypertension in SHRs. © 2015 The Authors. Journal of Cellular and Molecular Medicine published by John Wiley & Sons Ltd and Foundation for Cellular and Molecular Medicine.

Perinatally administered losartan augments renal ACE2 expression but not cardiac or renal Mas receptor in spontaneously hypertensive rats

PubMed Central

Klimas, Jan; Olvedy, Michael; Ochodnicka-Mackovicova, Katarina; Kruzliak, Peter; Cacanyiova, Sona; Kristek, Frantisek; Krenek, Peter; Ochodnicky, Peter

2015-01-01

Since the identification of the alternative angiotensin converting enzyme (ACE)2/Ang-(1-7)/Mas receptor axis, renin-angiotensin system (RAS) is a new complex target for a pharmacological intervention. We investigated the expression of RAS components in the heart and kidney during the development of hypertension and its perinatal treatment with losartan in young spontaneously hypertensive rats (SHR). Expressions of RAS genes were studied by the RT-PCR in the left ventricle and kidney of rats: normotensive Wistar, untreated SHR, SHR treated with losartan since perinatal period until week 9 of age (20 mg/kg/day) and SHR treated with losartan only until week 4 of age and discontinued until week 9. In the hypertrophied left ventricle of SHR, cardiac expressions of Ace and Mas were decreased while those of AT1 receptor (Agtr1a) and Ace2 were unchanged. Continuous losartan administration reduced LV weight (0.43 ± 0.02; P < 0.05 versus SHR) but did not influence altered cardiac RAS expression. Increased blood pressure in SHR (149 ± 2 in SHR versus 109 ± 2 mmHg in Wistar; P < 0.05) was associated with a lower renal expressions of renin, Agtr1a and Mas and with an increase in ACE2. Continuous losartan administration lowered blood pressure to control levels (105 ± 3 mmHg; P < 0.05 versus SHR), however, only renal renin and ACE2 were significantly up-regulated (for both P < 0.05 versus SHR). Conclusively, prevention of hypertension and LV hypertrophy development by losartan was unrelated to cardiac or renal expression of Mas. Increased renal Ace2, and its further increase by losartan suggests the influence of locally generated Ang-(1-7) in organ response to the developing hypertension in SHRs. PMID:25766467

Cytokine-mediated FOXO3a phosphorylation suppresses FasL expression in hemopoietic cell lines: investigations of the role of Fas in apoptosis due to cytokine starvation.

PubMed

Behzad, Hayedeh; Jamil, Sarwat; Denny, Trisha A; Duronio, Vincent

2007-05-01

We have investigated phosphatidylinositol 3-kinase (PI3K)-dependent survival signalling pathways using several cytokines in three different hemopoietic cell lines, MC/9, FDC-P1, and TF-1. Cytokines caused PI3K- and PKB-dependent phosphorylation of FOXO3a (previously known as FKHRL1) at three distinct sites. Following cytokine withdrawal or PI3K inhibition, both of which are known to lead to apoptosis, there was a loss of FOXO3a phosphorylation, and a resulting increase in forkhead transcriptional activity, along with increased expression of Fas Ligand (FasL), which could be detected at the cell surface. Concurrently, an increase in cell surface expression of Fas was also detected. Despite the presence of both FasL and Fas, there was no detectable evidence that activation of Fas-mediated apoptotic events was contributing to apoptosis resulting from cytokine starvation or inhibition of PI3K activity. Thus, inhibition of FOXO3a activity is mediated by the PI3K-PKB pathway, but regulation of FasL is not the primary means by which cell survival is regulated in cytokine-dependent hemopoietic cells. We were also able to confirm increased expression of known FOXO3a targets, Bim and p27kip1. Together, these results support the conclusion that mitochondrial-mediated signals play the major role in apoptosis of hemopoietic cells due to loss of cytokine signalling.

Development and implementation of an online program to improve how patients communicate emotional concerns to their oncology providers

PubMed Central

Porter, Laura S.; Pollak, Kathryn I.; Farrell, David; Cooper, Meredith; Arnold, Robert M.; Jeffreys, Amy S.; Tulsky, James A.

2015-01-01

Purpose Patients often struggle to express their emotional concerns to their oncology providers, and may therefore experience unmet needs. This paper describes the development and implementation of an online program that teaches patients how to communicate their emotions to their oncology providers. Methods The intervention was developed by a multidisciplinary team consisting of palliative care physicians, psychologists, and an intervention software developer, and included input from patients. It incorporated elements of Social Cognitive Theory and validated cognitive behavioral strategies for communication skills training. Strategies to increase intervention adherence were implemented midway through the study. Results The intervention consists of four interactive, online modules to teach patients strategies for expressing emotional concerns to their providers and asking for support. In addition to skill-building, the intervention was designed to raise patients’ expectations that expressing emotional concerns to providers would be helpful, to enhance their self-efficacy for doing so, and to help them overcome barriers to having these conversations. After implementing strategies to improve adherence, usage rates increased from 47% to 64%. Conclusions This intervention addresses an unmet educational need for patients with advanced cancer. Strategies to increase adherence led to improvements in usage rates in this population of older patients. We are currently evaluating the intervention in a randomized clinical trial to determine its efficacy in increasing patient expression of emotional concerns and requests for support. If successful, this intervention could serve as a model for future online patient education programs. PMID:25701437

The histone methyltransferase EZH2 as a novel prosurvival factor in clinically aggressive chronic lymphocytic leukemia.

PubMed

Papakonstantinou, Nikos; Ntoufa, Stavroula; Chartomatsidou, Elisavet; Kotta, Konstantia; Agathangelidis, Andreas; Giassafaki, Lefki; Karamanli, Tzeni; Bele, Panagiota; Moysiadis, Theodoros; Baliakas, Panagiotis; Sutton, Lesley Ann; Stavroyianni, Niki; Anagnostopoulos, Achilles; Makris, Antonios M; Ghia, Paolo; Rosenquist, Richard; Stamatopoulos, Kostas

2016-06-14

The histone methyltransferase EZH2 induces gene repression through trimethylation of histone H3 at lysine 27 (H3K27me3). EZH2 overexpression has been reported in many types of cancer and associated with poor prognosis. Here we investigated the expression and functionality of EZH2 in chronic lymphocytic leukemia (CLL). Aggressive cases with unmutated IGHV genes (U-CLL) displayed significantly higher EZH2 expression compared to indolent CLL cases with mutated IGHV genes (M-CLL); furthermore, in U-CLL EZH2 expression was upregulated with disease progression. Within U-CLL, EZH2high cases harbored significantly fewer (p = 0.033) TP53 gene abnormalities compared to EZH2low cases. EZH2high cases displayed high H3K27me3 levels and increased viability suggesting that EZH2 is functional and likely confers a survival advantage to CLL cells. This argument was further supported by siRNA-mediated downmodulation of EZH2 which resulted in increased apoptosis. Notably, at the intraclonal level, cell proliferation was significantly associated with EZH2 expression. Treatment of primary CLL cells with EZH2 inhibitors induced downregulation of H3K27me3 levels leading to increased cell apoptosis. In conclusion, EZH2 is overexpressed in adverse-prognosis CLL and associated with increased cell survival and proliferation. Pharmacologic inhibition of EZH2 catalytic activity promotes apoptosis, highlighting EZH2 as a novel potential therapeutic target for specific subgroups of patients with CLL.

The histone methyltransferase EZH2 as a novel prosurvival factor in clinically aggressive chronic lymphocytic leukemia

PubMed Central

Chartomatsidou, Elisavet; Kotta, Konstantia; Agathangelidis, Andreas; Giassafaki, Lefki; Karamanli, Tzeni; Bele, Panagiota; Moysiadis, Theodoros; Baliakas, Panagiotis; Sutton, Lesley Ann; Stavroyianni, Niki; Anagnostopoulos, Achilles; Makris, Antonios M.; Ghia, Paolo; Rosenquist, Richard; Stamatopoulos, Kostas

2016-01-01

The histone methyltransferase EZH2 induces gene repression through trimethylation of histone H3 at lysine 27 (H3K27me3). EZH2 overexpression has been reported in many types of cancer and associated with poor prognosis. Here we investigated the expression and functionality of EZH2 in chronic lymphocytic leukemia (CLL). Aggressive cases with unmutated IGHV genes (U-CLL) displayed significantly higher EZH2 expression compared to indolent CLL cases with mutated IGHV genes (M-CLL); furthermore, in U-CLL EZH2 expression was upregulated with disease progression. Within U-CLL, EZH2high cases harbored significantly fewer (p = 0.033) TP53 gene abnormalities compared to EZH2low cases. EZH2high cases displayed high H3K27me3 levels and increased viability suggesting that EZH2 is functional and likely confers a survival advantage to CLL cells. This argument was further supported by siRNA-mediated downmodulation of EZH2 which resulted in increased apoptosis. Notably, at the intraclonal level, cell proliferation was significantly associated with EZH2 expression. Treatment of primary CLL cells with EZH2 inhibitors induced downregulation of H3K27me3 levels leading to increased cell apoptosis. In conclusion, EZH2 is overexpressed in adverse-prognosis CLL and associated with increased cell survival and proliferation. Pharmacologic inhibition of EZH2 catalytic activity promotes apoptosis, highlighting EZH2 as a novel potential therapeutic target for specific subgroups of patients with CLL. PMID:27191993

Detection and characterisation of multi-drug resistance protein 1 (MRP-1) in human mitochondria

PubMed Central

Roundhill, E A; Burchill, S A

2012-01-01

Background: Overexpression of plasma membrane multi-drug resistance protein 1 (MRP-1) can lead to multidrug resistance. In this study, we describe for the first time the expression of mitochondrial MRP-1 in untreated human normal and cancer cells and tissues. Methods: MRP-1 expression and subcellular localisation in normal and cancer cells and tissues was examined by differential centrifugation and western blotting, and immunofluorescence microscopy. Viable mitochondria were isolated and MRP-1 efflux activity measured using the calcein-AM functional assay. MRP-1 expression was increased using retroviral infection and specific overexpression confirmed by RNA array. Cell viability was determined by trypan blue exclusion and annexin V-propidium iodide labelling of cells. Results: MRP-1 was detected in the mitochondria of cancer and normal cells and tissues. The efflux activity of mitochondrial MRP-1 was more efficient (55–64%) than that of plasma membrane MRP-1 (11–22% P<0.001). Induced MRP-1 expression resulted in a preferential increase in mitochondrial MRP-1, suggesting selective targeting to this organelle. Treatment with a non-lethal concentration of doxorubicin (0.85 n, 8 h) increased mitochondrial and plasma membrane MRP-1, increasing resistance to MRP-1 substrates. For the first time, we have identified MRP-1 with efflux activity in human mitochondria. Conclusion: Mitochondrial MRP-1 may be an exciting new therapeutic target where historically MRP-1 inhibitor strategies have limited clinical success. PMID:22353810

Inhibition of Prolyl Hydroxylase Attenuates Fas Ligand-Induced Apoptosis and Lung Injury in Mice.

PubMed

Nagamine, Yusuke; Tojo, Kentaro; Yazawa, Takuya; Takaki, Shunsuke; Baba, Yasuko; Goto, Takahisa; Kurahashi, Kiyoyasu

2016-12-01

Alveolar epithelial injury and increased alveolar permeability are hallmarks of acute respiratory distress syndrome. Apoptosis of lung epithelial cells via the Fas/Fas ligand (FasL) pathway plays a critical role in alveolar epithelial injury. Activation of hypoxia-inducible factor (HIF)-1 by inhibition of prolyl hydroxylase domain proteins (PHDs) is a possible therapeutic approach to attenuate apoptosis and organ injury. Here, we investigated whether treatment with dimethyloxalylglycine (DMOG), an inhibitor of PHDs, could attenuate Fas/FasL-dependent apoptosis in lung epithelial cells and lung injury. DMOG increased HIF-1α protein expression in vitro in MLE-12 cells, a murine alveolar epithelial cell line. Treatment of MLE-12 cells with DMOG significantly suppressed cell surface expression of Fas and attenuated FasL-induced caspase-3 activation and apoptotic cell death. Inhibition of the HIF-1 pathway by echinomycin or small interfering RNA transfection abolished these antiapoptotic effects of DMOG. Moreover, intraperitoneal injection of DMOG in mice increased HIF-1α expression and decreased Fas expression in lung tissues. DMOG treatment significantly attenuated caspase-3 activation, apoptotic cell death in lung tissue, and the increase in alveolar permeability in mice instilled intratracheally with FasL. In addition, inflammatory responses and histopathological changes were also significantly attenuated by DMOG treatment. In conclusion, inhibition of PHDs protects lung epithelial cells from Fas/FasL-dependent apoptosis through HIF-1 activation and attenuates lung injury in mice.

The flavonoid hesperidin exerts anti-photoaging effect by downregulating matrix metalloproteinase (MMP)-9 expression via mitogen activated protein kinase (MAPK)-dependent signaling pathways.

PubMed

Lee, Hee Jeong; Im, A-Rang; Kim, Su-Man; Kang, Hyung-Sik; Lee, Jae Dong; Chae, Sungwook

2018-01-30

Hesperidin is a flavonoid with antioxidant, anti-inflammatory, and immune modulatory activities. Photoaging is a consequence of chronic exposure to the sun and ultraviolet (UV) radiation. This study was designed to evaluate the efficacy of hesperidin against photoaging of dorsal skin in hairless mice. Hairless male mice (6-week-old) were divided into three groups (n = 7): control, UVB-treated vehicle, and UVB-treated hesperidin groups. UVB-irradiated mice from hesperidin group were orally administered 0.1 mL of water containing 100 mg/kg body weight per day hesperidin. The mean length and depth of wrinkles in the UVB-treated hesperidin group significantly improved after the oral administration of hesperidin, which significantly inhibited the increase in epidermal thickness and epidermal hypertrophy (P < 0.05). UVB irradiation of mice induced epidermal barrier dysfunction including an increase in the transepidermal water loss (TEWL); however, hesperidin decreased the TEWL. UVB irradiation increased the expression of MMP-9 and pro-inflammatory cytokines whereas UVB-treated hesperidin group showed reduced expression. These results indicate that hesperidin showed anti-photoaging activity in the UVB-irradiated hairless mice. In conclusion, hesperidin inhibited the UVB-induced increase in skin thickness, wrinkle formation, and collagen fiber loss in male hairless mice. These results suggest that hesperidin shows potent anti-photoaging activity by regulating MMP-9 expression through the suppression of MAPK-dependent signaling pathways.

Zirconium Ions Up-Regulate the BMP/SMAD Signaling Pathway and Promote the Proliferation and Differentiation of Human Osteoblasts

PubMed Central

Chen, Yongjuan; Roohani-Esfahani, Seyed-Iman; Lu, ZuFu; Zreiqat, Hala; Dunstan, Colin R.

2015-01-01

Zirconium (Zr) is an element commonly used in dental and orthopedic implants either as zirconia (ZrO2) or in metal alloys. It can also be incorporated into calcium silicate-based ceramics. However, the effects of in vitro culture of human osteoblasts (HOBs) with soluble ionic forms of Zr have not been determined. In this study, primary culture of human osteoblasts was conducted in the presence of medium containing either ZrCl4 or Zirconium (IV) oxynitrate (ZrO(NO3)2) at concentrations of 0, 5, 50 and 500 µM, and osteoblast proliferation, differentiation and calcium deposition were assessed. Incubation of human osteoblast cultures with Zr ions increased the proliferation of human osteoblasts and also gene expression of genetic markers of osteoblast differentiation. In 21 and 28 day cultures, Zr ions at concentrations of 50 and 500 µM increased the deposition of calcium phosphate. In addition, the gene expression of BMP2 and BMP receptors was increased in response to culture with Zr ions and this was associated with increased phosphorylation of SMAD1/5. Moreover, Noggin suppressed osteogenic gene expression in HOBs co-treated with Zr ions. In conclusion, Zr ions appear able to induce both the proliferation and the differentiation of primary human osteoblasts. This is associated with up-regulation of BMP2 expression and activation of BMP signaling suggesting this action is, at least in part, mediated by BMP signaling. PMID:25602473

Changes in protein and gene expression of angiotensin II receptors (AT1 and AT2) in aorta of diabetic and hypertensive rats.

PubMed

Romero-Nava, R; Rodriguez, J E; Reséndiz-Albor, A A; Sánchez-Muñoz, F; Ruiz-Hernandéz, A; Huang, F; Hong, E; Villafaña, S

2016-01-01

Diabetes and hypertension have been associated with cardiovascular diseases and stroke. Some reports have related the coexistence of hypertension and diabetes with increase in the risk of developing vascular complications. Recently some studies have shown results suggesting that in the early stages of diabetes and hypertension exist a reduced functional response to vasopressor agents like angiotensin II (Ang II), which plays an important role in blood pressure regulation mechanism through the activation of its AT1 and AT2 receptors. For that reason, the aim of this work was to study the gene and protein expression of AT1 and AT2 receptors in aorta of diabetic SHR and WKY rats. Diabetes was induced by the administration of streptozotocin (60 mg/kg i.p.). After 4 weeks of the onset of diabetes, the protein expression was obtained by western blot and the mRNA expression by RT-PCR. Our results showed that the hypertensive rats have a higher mRNA and protein expression of AT1 receptors than normotensive rats while the AT2 expression remained unchanged. On the other hand, the combination of diabetes and hypertension increased the mRNA and protein expression of AT1 and AT2 receptors significantly. In conclusion, our results suggest that diabetes with hypertension modifies the mRNA and protein expression of AT1 and AT2 receptors. However, the overexpression of AT2 could be associated with the reduction in the response to Ang II in the early stage of diabetes.

Isotretinoin therapy changes the expression of antimicrobial peptides in acne vulgaris.

PubMed

Borovaya, Alena; Dombrowski, Yvonne; Zwicker, Stephanie; Olisova, Olga; Ruzicka, Thomas; Wolf, Ronald; Schauber, Jürgen; Sárdy, Miklós

2014-10-01

In acne vulgaris, antimicrobial peptides (AMPs) could play a dual role; i.e., protective by acting against Propionibacterium acnes, pro-inflammatory by acting as signalling molecules. The cutaneous expression of 15 different AMPs was investigated in acne patients; furthermore, the impact of isotretinoin therapy on AMP expression was analysed in skin biopsies from 13 patients with acne vulgaris taken before, during and after a 6-month treatment cycle with isotretinoin using quantitative real-time polymerase chain reaction. Cutaneous expression of the AMPs cathelicidin, human β-defensin-2 (HBD-2), lactoferrin, lysozyme, psoriasin (S100A7), koebnerisin (S100A15), and RNase 7 was upregulated in untreated acne vulgaris, whereas α-defensin-1 (HNP-1) was downregulated compared to controls. While relative expression levels of cathelicidin, HBD-2, lactoferrin, psoriasin (S100A7), and koebnerisin (S100A15) decreased during isotretinoin treatment, only those of cathelicidin and koebnerisin returned to normal after 6 months of isotretinoin therapy. The increased expression of lysozyme and RNase 7 remained unaffected by isotretinoin treatment. The levels of granulysin, RANTES (CCL5), perforin, CXCL9, substance P, chromogranin B, and dermcidin were not regulated in untreated acne patients and isotretinoin had no effect on these AMPs. In conclusion, the expression of various AMPs is altered in acne vulgaris. Isotretinoin therapy normalizes the cutaneous production of distinct AMPs while the expression of others is still increased in healing acne. Considering the antimicrobial and pro-inflammatory role of AMPs, these molecules could serve as specific targets for acne therapy and maintenance of clinical remission.

Lymphatic function is regulated by a coordinated expression of lymphangiogenic and anti-lymphangiogenic cytokines

PubMed Central

Zampell, Jamie C.; Avraham, Tomer; Yoder, Nicole; Fort, Nicholas; Yan, Alan; Weitman, Evan S.

2012-01-01

Lymphangiogenic cytokines such as vascular endothelial growth factor-C (VEGF-C) are critically required for lymphatic regeneration; however, in some circumstances, lymphatic function is impaired despite normal or elevated levels of these cytokines. The recent identification of anti-lymphangiogenic molecules such as interferon-γ (IFN-γ), transforming growth factor-β1, and endostatin has led us to hypothesize that impaired lymphatic function may represent a dysregulated balance in the expression of pro/anti-lymphangiogenic stimuli. We observed that nude mice have significantly improved lymphatic function compared with wild-type mice in a tail model of lymphedema. We show that gradients of lymphatic fluid stasis regulate the expression of lymphangiogenic cytokines (VEGF-A, VEGF-C, and hepatocyte growth factor) and that paradoxically the expression of these molecules is increased in wild-type mice. More importantly, we show that as a consequence of T-cell-mediated inflammation, these same gradients also regulate expression patterns of anti-lymphangiogenic molecules corresponding temporally and spatially with impaired lymphatic function in wild-type mice. We show that neutralization of IFN-γ significantly increases inflammatory lymph node lymphangiogenesis independently of changes in VEGF-A or VEGF-C expression, suggesting that alterations in the balance of pro- and anti-lymphangiogenic cytokine expression can regulate lymphatic vessel formation. In conclusion, we show that gradients of lymphatic fluid stasis regulate not only the expression of pro-lymphangiogenic cytokines but also potent suppressors of lymphangiogenesis as a consequence of T-cell inflammation and that modulation of the balance between these stimuli can regulate lymphatic function. PMID:21940662

Elevated Expression of Immunoreceptor Tyrosine-Based Inhibitory Motif (TIGIT) on T Lymphocytes is Correlated with Disease Activity in Rheumatoid Arthritis.

PubMed

Luo, Qing; Deng, Zhen; Xu, Chuxin; Zeng, Lulu; Ye, Jianqing; Li, Xue; Guo, Yang; Huang, Zikun; Li, Junming

2017-03-10

BACKGROUND It is well known that lymphocytes play an important role in rheumatoid arthritis (RA). T cell immunoreceptors with immunoglobulin (Ig) and immunoreceptor tyrosine-based inhibitory motif (TIGIT) have immunosuppressive co-stimulatory molecules that mediate inhibitory effects, but their roles in RA are poorly understood. MATERIAL AND METHODS Were recruited 76 patients with RA and 33 healthy controls (HC). Clinical manifestations, laboratory measurements, physical examination, and medical history of RA patients were recorded. The expression of TIGIT on CD3+ T lymphocytes, B lymphocytes, monocytes, neutrophils, CD3+CD4+ T lymphocytes, and CD3+CD8+ T lymphocytes was determined using flow cytometry. The expression of TIGIT on T lymphocytes in patients with RA was further analyzed to investigate its correlations with markers of autoimmune response, inflammation, and disease activity in RA. RESULTS Compared with HC, the expression levels of TIGIT on CD3+CD4+ T lymphocytes and CD3+CD8+ T lymphocytes were significantly increased in patients with RA (P < 0.01). The frequency of TIGIT-expressing CD3+CD4+ T lymphocytes was positively correlated with RF, increased ACPA, ESR, and CRP levels. The frequency of TIGIT-expressing CD3+CD8+ T lymphocytes was positively correlated with RF and ESR levels. Furthermore, the expression level of TIGIT on CD3+CD4+ T lymphocytes was positively correlated with the DAS28 score in RA. CONCLUSIONS The expression levels of TIGIT on T lymphocytes were elevated and correlated with disease activity in RA.

Differential Expression of Mitochondrial Manganese Superoxide Dismutase (SOD) in Triticum aestivum Exposed to Silver Nitrate and Silver Nanoparticles.

PubMed

Karimi, Javad; Mohsenzadeh, Sasan; Niazi, Ali; Moghadam, Ali

2017-01-01

Background: The increasing use of nanoparticles (NPs) may have negative impacts on both organisms and the environment. Objectives: The differential expression of mitochondrial manganese superoxide dismutase ( MnSOD ) gene in wheat in response to silver nitrate nanoparticles (AgNPs) and AgNO 3 was investigated. Materials and Methods: A quantitative Real-Time RT-PCR experiment was carried out with MnSOD gene using RNAs isolated from wheat shoots treated for 0, 2, 6, 12, and 24 h with 100 mg.L -1 of either AgNO 3 or AgNPs. Results: The results of this study showed that both treatments cause changes in the expression pattern of the MnSOD gene. While 2 and 6 h following the beginning of the stress, MnSOD expression was up-regulated significantly, in response to AgNO 3 (1.4 and 2.8 fold, respectively), in response to AgNPs, it was up-regulated significant only after 6 h (1.6 fold), compared with the control. The gene expression, after 12 h in response to AgNO 3 and AgNPs were downregulated significantly (0.7 and 0.8 fold, respectively), and in the next 12 h , the expression appeared to be similar to the control. Conclusion: Exposure to both AgNPs and Ag ions led to a significant increase in MnSOD expression, but AgNO 3 changed the MnSOD expression faster than AgNPs. Therefore, it is suggested that AgNO 3 has greater penetrability and effectiveness.

Suppression of thymosin β10 increases cell migration and metastasis of cholangiocarcinoma

PubMed Central

2013-01-01

Background Thymosin β10 (Tβ10) expression is associated with malignant phenotypes in many cancers. However, the role and mechanisms of Tβ10 in liver fluke-associated cholangiocarcinoma (CCA) are not fully understood. In this study, we investigated the expression of Tβ10 in CCA tumor tissues and cell lines as well as molecular mechanisms of Tβ10 in tumor metastasis of CCA cell lines. Methods Tβ10 expression was determined by real time RT-PCR or immunocytochemistry. Tβ10 silence or overexpression in CCA cells was achieved using gene delivery techniques. Cell migration was assessed using modified Boyden chamber and wound healing assay. The effect of silencing Tβ10 on CCA tumor metastasis was determined in nude mice. Phosphorylation of ERK1/2 and the expression of EGR1, Snail and matrix metalloproteinases (MMPs) were studied. Results Ten pairs of CCA tissues (primary and metastatic tumors) and 5 CCA cell lines were studied. With real time RT-PCR and immunostaining analysis, Tβ10 was highly expressed in primary tumors of CCA; while it was relatively low in the metastatic tumors. Five CCA cell lines showed differential expression levels of Tβ10. Silence of Tβ10 significantly increased cell migration, invasion and wound healing of CCA cells in vitro; reversely, overexpression of Tβ10 reduced cell migration compared with control cells (P<0.05). In addition, silence of Tβ10 in CCA cells increased liver metastasis in a nude mouse model of CCA implantation into the spleen. Furthermore, silence of Tβ10 activated ERK1/2 and increased the expression of Snail and MMPs in CCA cell lines. Ras-GTPase inhibitor, FPT inhibitor III, effectively blocked Tβ10 silence-associated ERK1/2 activation, Snail expression and cell migration. Conclusions Low expression of Tβ10 is associated with metastatic phenotype of CCA in vitro and in vivo, which may be mediated by the activation of Ras, ERK1/2 and upregulation of Snail and MMPs. This study suggests a new molecular pathway of CCA pathogenesis and a novel strategy to treat or prevent CCA metastasis. PMID:24053380

The effect of clozapine on mRNA expression for genes encoding G protein-coupled receptors and the protein components of clathrin-mediated endocytosis

PubMed Central

Hu, Ying; Weymer, Jon F.; Rizig, Mie; McQuillin, Andrew; Hunt, Stephen P.; Gurling, Hugh M.D.

2013-01-01

Objectives Clathrin-mediated endocytosis (CME) is an intracellular trafficking mechanism for packaging cargo, including G protein-coupled receptors (GPCRs), into clathrin-coated vesicles (CCVs). The antipsychotic chlorpromazine inhibits CCV assembly of adaptor protein AP2 whereas clozapine increases serotonin2A receptor internalization. We hypothesized that clozapine alters the expression of CME genes modulating vesicle turnover and GPCR internalization. Materials and methods SH-SY5Y human neuroblastoma cells were incubated with clozapine (1–20 µmol/l) for 24–72 h. GPCR and CME-related gene mRNA expression was measured using RT-PCR. We quantified changes in the same genes using expression data from a microarray study of mice brains after 12 weeks of treatment with 12 mg/kg/day clozapine. Results The expression of genes encoding adaptor and clathrin assembly proteins, AP2A2, AP2B1, AP180, CLINT1, HIP1, ITSN2, and PICALM, increased relative to the control in SH-SY5Y cells incubated with 5–10 µmol/l clozapine for 24–72 h. The microarray study showed significantly altered expression of the above CME-related genes, with a marked 641-fold and 17-fold increase in AP180 and the serotonin1A GPCR, respectively. The expression of three serotonergic receptor and lysophosphatidic acid receptor 2 (EDG4) GPCR genes was upregulated in SH-SY5Y cells incubated with 5 µmol/l clozapine for 24 h. EDG4 expression was also increased with 10–20 µmol/l clozapine treatment at 48–72 h. Clozapine significantly decreased the expression of β-arrestin, involved in GPCR desensitization, both in vitro and vivo. Conclusion The changes we report in CME and GPCR mRNAs implicate CCV-mediated internalization of GPCRs and the serotonergic system in clozapine’s mechanism of action, which may be useful in the design of more effective and less toxic antipsychotic therapies. PMID:23811784

Effects of high-pressure oxygen therapy on brain tissue water content and AQP4 expression in rabbits with cerebral hemorrhage.

PubMed

Wu, Jing; Chen, Jiong; Guo, Hua; Peng, Fang

2014-12-01

To investigate the effects of different atmosphere absolutes (ATA) of high-pressure oxygen (HPO) on brain tissue water content and Aquaporin-4 (AQP4) expression in rabbits with cerebral hemorrhage. 180 New Zealand white rabbits were selected and randomly divided into normal group (n = 30), control group (n = 30) and cerebral hemorrhage group (n = 120), and cerebral hemorrhage group was divided into group A, B, C and D with 30 rabbits in each group. The groups received 1.0, 1.8, 2.0 and 2.2 ATA of HPO treatments, respectively. Ten rabbits in each group were killed at first, third and fifth day to detect the brain tissue water content and change of AQP4 expression. In cerebral hemorrhage group, brain tissue water content and AQP4 expression after model establishment were first increased, then decreased and reached the maximum on third day (p < 0.05). Brain tissue water content and AQP4 expression in control group and cerebral hemorrhage group were significantly higher than normal group at different time points (p < 0.05). In contrast, brain tissue water content and AQP4 expression in group C were significantly lower than in group A, group B, group D and control group (p < 0.05). In control group, AQP4-positive cells significantly increased after model establishment, which reached maximum on third day, and positive cells in group C were significantly less than in group A, group B and group D. We also found that AQP4 expression were positively correlated with brain tissue water content (r = 0.719, p < 0.05) demonstrated by significantly increased AQP4 expression along with increased brain tissue water content. In conclusion, HPO can decrease AQP4 expression in brain tissue of rabbits with cerebral hemorrhage to suppress the progression of brain edema and promote repairing of injured tissue. 2.0 ATA HPO exerts best effects, which provides an experimental basis for ATA selection of HPO in treating cerebral hemorrhage.

TNF-{alpha} mediates the stimulation of sclerostin expression in an estrogen-deficient condition

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

Kim, Beom-Jun; Bae, Sung Jin; Lee, Sun-Young

Highlights: Black-Right-Pointing-Pointer Estrogen deprivation stimulates the bony sclerostin levels with reversal by estrogen. Black-Right-Pointing-Pointer TNF-{alpha} increases the activity and expression of MEF2 in UMR-106 cells. Black-Right-Pointing-Pointer TNF-{alpha} blocker prevents the stimulation of bony sclerostin expression by ovariectomy. Black-Right-Pointing-Pointer No difference in bony sclerostin expression between sham-operated and ovariectomized nude mice. -- Abstract: Although recent clinical studies have suggested a possible role for sclerostin, a secreted Wnt antagonist, in the pathogenesis of postmenopausal osteoporosis, the detailed mechanisms how estrogen deficiency regulates sclerostin expression have not been well-elucidated. Bilateral ovariectomy or a sham operation in female C57BL/6 mice and BALB/c nude micemore » was performed when they were seven weeks of age. The C57BL/6 mice were intraperitoneally injected with phosphate-buffered serum (PBS), 5 {mu}g/kg {beta}-estradiol five times per week for three weeks, or 10 mg/kg TNF-{alpha} blocker three times per week for three weeks. Bony sclerostin expression was assessed by immunohistochemistry staining in their femurs. The activity and expression of myocyte enhancer factors 2 (MEF2), which is essential for the transcriptional activation of sclerostin, in rat UMR-106 osteosarcoma cells were determined by luciferase reporter assay and western blot analysis, respectively. Bony sclerostin expression was stimulated by estrogen deficiency and it was reversed by estradiol supplementation. When the UMR-106 cells were treated with well-known, estrogen-regulated cytokines, only TNF-{alpha}, but not IL-1 and IL-6, increased the MEF2 activity. Consistently, TNF-{alpha} also increased the nuclear MEF2 expression. Furthermore, the TNF-{alpha} blocker prevented the stimulation of bony sclerostin expression by ovariectomy. We also found that there was no difference in sclerostin expression between ovariectomized nude mice and sham-operated nude mice. In conclusion, these results suggest that TNF-{alpha} originating from T cells may be at least in part responsible for stimulating the sclerostin expression observed in an estrogen-deficient condition.« less

Plasma and adipose tissue level of angiopoietin-like 7 (ANGPTL7) are increased in obesity and reduced after physical exercise

PubMed Central

Cherian, Preethi; Al-Khairi, Irina; Madhu, Dhanya; Tiss, Ali; Warsam, Samia; Alhubail, Asma; Sriraman, Devarajan; Al-Refaei, Faisal; Abubaker, Jehad

2017-01-01

Objective ANGPTL7 is a member of the Angiopoietin-like (ANGPTL) protein family that is composed of eight proteins (1–8). Increasing evidence is associating ANGPTL proteins to obesity and insulin resistance. The biological role of ANGPTL7 is yet to be understood except for a recently proposed role in the pathophysiology of glaucoma. This study was designed to shed light on the function of ANGPTL7 in obesity and its modulation by physical exercise as well as its potential association with lipid profile. Methods A total of 144 subjects were enrolled in this study and finished three months of physical exercise. The participants were classified based on their BMI, 82 subjects were non-obese and 62 obese. ANGPTL7 levels in plasma and adipose tissue were measured by ELISA, RT-PCR and immunohistochemistry. Results In this study, we showed that ANGPTL7 level was increased in the plasma of obese subjects (1249.05± 130.39 pg/mL) as compared to non-obese (930.34 ± 87.27 pg/mL) (p-Value = 0.032). ANGPTL7 Gene and protein expression levels in adipose tissue also showed over two fold increase. Physical exercise reduced circulating level of ANGPTL7 in the obese subjects to 740.98± 127.18 pg/mL, (p-Value = 0.007). ANGPTL7 expression in adipose tissue was also reduced after exercise. Finally, ANGPTL7 circulating level showed significant association with TG level in the obese subjects (R2 = 0.183, p-Value = 0.03). Conclusion In conclusion, our data shows for the first time that obesity increases the level of ANGPTL7 in both plasma and adipose tissue. Increased expression of ANGPTL7 might play a minor role in the regulation of TG level in obese subjects either directly or through interaction with other ANGPTL protein members. Physical exercise reduced the level of ANGPTL7 highlighting the potential for targeting this protein as a therapeutic target for regulating dyslipidemia. PMID:28264047

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

Brown, Kyle E.; Division of Gastroenterology-Hepatology, University of Iowa Roy J. and Lucille A. Carver College of Medicine; Program in Free Radical and Radiation Biology, University of Iowa Roy J. and Lucille A. Carver College of Medicine, Iowa City, IA

Introduction:: Oxidative stress can trigger a cellular stress response characterized by induction of antioxidants, acute phase reactants (APRs) and heat shock proteins (HSPs), which are presumed to play a role in limiting tissue damage. In rodents, hepatic iron overload causes oxidative stress that results in upregulation of antioxidant defenses with minimal progressive liver injury. The aim of this study was to determine whether iron overload modulates expression of other stress-responsive proteins such as APRs and HSPs that may confer protection against iron-induced damage in rodent liver. Methods:: Male rats received repeated injections of iron dextran or dextran alone over amore » 6-month period. Hepatic transcript levels for a panel of APRs and HSPs were quantitated by real-time PCR and protein expression was evaluated by Western blot and immunohistochemistry. Results:: Hepatic iron concentrations were increased > 50-fold in the iron-loaded rats compared to controls. Iron loading resulted in striking increases in mRNAs for Hsp32 (heme oxygenase-1; 12-fold increase vs. controls) and metallothionein-1 and -2 (both increased {approx} 6-fold). Transcripts for {alpha}1-acid glycoprotein, the major rat APR, were increased {approx} 3-fold, while expression of other classical APRs was unaltered. Surprisingly, although mRNA levels for the HSPs were not altered by iron, the abundance of Hsp25, Hsp70 and Hsp90 proteins was uniformly reduced in the iron-loaded livers, as were levels of NAD(P)H:quinone oxidoreductase 1, an Hsp70 client protein. Conclusions:: Chronic iron administration elicits a unique pattern of stress protein expression. These alterations may modulate hepatic responses to iron overload, as well as other injury processes.« less

Fluoxetine prevents the development of depressive-like behavior in a mouse model of cancer related fatigue.

PubMed

Norden, Diana M; Devine, Raymond; Bicer, Sabahattin; Jing, Runfeng; Reiser, Peter J; Wold, Loren E; Godbout, Jonathan P; McCarthy, Donna O

2015-03-01

Cancer patients frequently suffer from fatigue, a complex syndrome associated with tiredness and depressed mood. Cancer-related fatigue (CRF) can be present at the time of diagnosis, escalates during treatment, and can persist for years after treatment. CRF negatively influences quality of life, limits functional independence, and is associated with decreased survival in patients with incurable disease. We have previously shown that increased pro-inflammatory cytokine expression in the brain contributes to depressive- and fatigue-like behaviors in a mouse model of CRF. Inflammatory cytokines increase the activity of indoleamine 2,3-dioxygenase (IDO) and kynurenine 3-monooxygenase (KMO), which competitively reduce serotonin synthesis. Reduced serotonin availability in the brain and increased production of alternative neuroactive metabolites of tryptophan are thought to contribute to the development of depression and fatigue. The purpose of this study was to determine the effects of fluoxetine, a selective serotonin reuptake inhibitor (SSRI), on brain cytokines and behavioral measures of fatigue and depression in tumor-bearing mice. Here we show that tumor growth increased brain expression of pro-inflammatory cytokines and KMO. Treatment with fluoxetine had no effect on tumor growth, muscle wasting, fatigue behavior, or cytokine expression in the brain. Fluoxetine, however, reduced depressive-like behaviors in tumor bearing mice. In conclusion, our data confirm that increased brain expression of pro-inflammatory cytokines is associated with tumor-induced fatigue- and depressive-like behaviors. However, it is possible to separate the effects of tumor growth on mood and fatigue-like behaviors using SSRIs such as fluoxetine. Copyright © 2014 Elsevier Inc. All rights reserved.

Toll-like receptor 4 increases intestinal permeability through up-regulation of membrane PKC activity in alcoholic steatohepatitis.

PubMed

Li, Xin; Wang, Chen; Nie, Jiao; Lv, Dong; Wang, Tianyi; Xu, Youqing

2013-09-01

Intestinal hyperpermeability is a causal factor for the development of alcoholic endotoxemia and steatohepatitis. However, the mechanisms governing this link remain unknown. The purpose of this study was to determine whether toll-like receptor 4 (TLR4) is involved in ethanol's deleterious effects on the intestinal barrier. Caco-2 cells were incubated in vitro with 1-10% ethanol. The results indicated that ethanol had a dose-dependent effect in increasing TLR4 expression and intercellular permeability. Then the effects of TLR4 on protein kinase C (PKC) and the intercellular junction protein occludin were assessed with and without pretreatment with a TLR4 inhibitor. The results indicated that TLR4 increased nonspecific PKC activity and reduced the expression of phosphorylated occludin in the membrane, which increased intercellular permeability. These effects were prevented by pretreatment with TLR4 mAb. Wild-type C57BL/6 mice were fed an ethanol or isocaloric liquid diet for 6 weeks. Hepatitis was diagnosed by the presence of an associated elevated blood endotoxin level. Chronic ethanol treatment significantly elevated blood endotoxin levels, intestinal permeability, and the expression of TLR4 in the ileum and colon. Moreover, ethanol exposure reduced the distribution of phosphorylated occludin in the intestinal epithelium because of PKC activation. In conclusion, chronic ethanol exposure induces a high response of TLR4 to lipopolysaccharide (LPS), and TLR4 increases intestinal permeability through down-regulation of phosphorylated occludin expression in the intestinal epithelial barrier, accompanied by membrane PKC hyperactivity. Copyright © 2013 Elsevier Inc. All rights reserved.

Renal papillary tip extract stimulates BNP production and excretion from cardiomyocytes

PubMed Central

Hashizume, Ryotaro; Suzuki, Noboru; Ito, Rie; Yamanaka, Keiichi; Saito, Hiromitsu; Kiyonari, Hiroshi; Tawara, Isao; Kageyama, Yuki; Ogihara, Yoshito; Ali, Yusuf; Yamada, Norikazu; Katayama, Naoyuki; Ito, Masaaki

2018-01-01

Background Brain natriuretic peptide (BNP) is an important biomarker for patients with cardiovascular diseases, including heart failure, hypertension and cardiac hypertrophy. It is also known that BNP levels are relatively higher in patients with chronic kidney disease and no heart disease; however, the mechanism remains unclear. Methods and results We developed a BNP reporter mouse and occasionally found that this promoter was activated specifically in the papillary tip of the kidneys, and its activation was not accompanied by BNP mRNA expression. No evidence was found to support the existence of BNP isoforms or other nucleotide expression apart from BNP and tdTomato. The pBNP-tdTomato-positive cells were interstitial cells and were not proliferative. Unexpectedly, both the expression and secretion of BNP increased in primary cultured neonatal cardiomyocytes after their treatment with an extract of the renal papillary tip. Intraperitoneal injection of the extract of the papillary tips reduced blood pressure from 210 mmHg to 165 mmHg, the decrease being accompanied by an increase in serum BNP and urinary cGMP production in stroke-prone spontaneously hypertensive (SHR-SP) rats. Furthermore the induction of BNP by the papillary extract from rats with heart failure due to myocardial infarction was increased in cardiomyocytes. Conclusions These results suggested that the papillary tip express a substance that can stimulate BNP production and secretion from cardiomyocytes. PMID:29734386

Sulforaphane protection against the development of doxorubicin-induced chronic heart failure is associated with Nrf2 Upregulation.

PubMed

Bai, Yang; Chen, Qiang; Sun, Yun-Peng; Wang, Xuan; Lv, Li; Zhang, Li-Ping; Liu, Jin-Sha; Zhao, Song; Wang, Xiao-Lu

2017-10-01

Doxorubicin (DOX) is an anthracycline antitumor drug. However, its clinical use is limited by dose-dependent cardiotoxicity and even progresses to chronic heart failure (CHF). This study aims to investigate whether the Nrf2 activator, sulforaphane (SFN), can prevent DOX-induced CHF. Male Sprague-Dawley rats which received treatment for 6 weeks were divided into four groups (n=30 per group): control, SFN, DOX and DOX plus SFN group. Results revealed that DOX induced progressive cardiac damage as indicated by increased cardiac injury markers, cardiac inflammation, fibrosis and oxidative stress. SFN significantly prevented DOX-induced progressive cardiac dysfunction between 2-6 weeks and prevented DOX-induced cardiac function deterioration. Furthermore, it significantly decreased ejection fraction and increased the expression of brain natriuretic peptide. SFN also almost completely prevented DOX-induced cardiac oxidative stress, inflammation and fibrosis. SFN upregulated NF-E2-related factor 2 (Nrf2) expression and transcription activity, which was reflected by the increased mRNA expression of Nrf2 and its downstream genes. Furthermore, in cultured H9c2 cardiomyocytes, the protective effect of SFN against DOX-induced fibrotic and inflammatory responses was abolished by Nrf2 silencing. We arrived at the conclusion that DOX-induced CHF can be prevented by SFN through the upregulation of Nrf2 expression and transcriptional function. © 2017 John Wiley & Sons Ltd.

Tisp40 deficiency attenuates renal ischemia reperfusion injury induced apoptosis of tubular epithelial cells.

PubMed

Qin, Cong; Xiao, Chengcheng; Su, Yang; Zheng, Haizhou; Xu, Tao; Lu, Jingxiao; Luo, Pengcheng; Zhang, Jie

2017-10-01

Renal ischemia reperfusion (IR) is a major cause of acute kidney injury (AKI) and no effective treatments have been established. Tisp40 is a transcription factor of the CREB/ATF family and involves in cell apoptosis, proliferation and differentiation, but its role in renal IR remains unknown. Here, we investigated the role of Tisp40 in renal IR injury. In vivo, Tisp40 knockout (KO) and wild-type (WT) mice were subjected to thirty minutes of bilateral renal ischemia and 48h reperfusion, the blood and kidneys were harvested for analysis. In vitro, Tisp40 overexpression and vector cells were subjected to hypoxia/reoxygenation (HR), the apoptosis rate and the expressions of related proteins were measured. Following IR, the expressions of Tisp40 protein, serum creatinine (sCr), blood urea nitrogen (BUN) and apoptosis of tubular cells were significantly increased in WT mice. However, Tisp40 deficiency significantly attenuated the increase of sCr, BUN and apoptosis of tubular cells. Following HR, apoptosis of tubular cells was increased in Tisp40 overexpression cells compared with vector cells. Mechanistically, Tisp40 promoted the expressions of C/EBP homologous protein (CHOP), Bax and Cleaved caspase3 and suppressed the expression of Bcl-2 in renal IR injury. In conclusion, Tisp40 aggravates tubular cells apoptosis in renal IR injury. Copyright © 2017 Elsevier Inc. All rights reserved.

Sulforaphane Ameliorates Bladder Dysfunction through Activation of the Nrf2-ARE Pathway in a Rat Model of Partial Bladder Outlet Obstruction

PubMed Central

Liu, Chong; Xu, Huan; Fu, Shi; Chen, Yanbo; Chen, Qi; Cai, Zhikang; Zhou, Juan; Wang, Zhong

2016-01-01

Purpose. We evaluated the effect of sulforaphane (SFN) treatment on the function and changes of expression of Nrf2-ARE pathway in the bladder of rats with bladder outlet obstruction (BOO). Materials and Methods. A total of 18 male Sprague-Dawley rats at age of 8 weeks were divided into 3 groups (6 of each): the sham operated group, the BOO group, and the BOO+SFN group. We examined histological alterations and the changes of oxidative stress markers and the protein expression of the Nrf2-ARE pathway. Results. We found that SFN treatment could prolong micturition interval and increase bladder capacity and bladder compliance. However, the peak voiding pressure was lower than BOO group. SFN treatment can ameliorate the increase of collagen fibers induced by obstruction. SFN treatment also increased the activity of SOD, GSH-Px, and CAT compared to the other groups. The level of bladder cell apoptosis was decreased in BOO rats with SFN treatment. Moreover, SFN could reduce the ratio of Bax/Bcl-2 expression. Furthermore, SFN could activate the Nrf2 expression with elevation of its target antioxidant proteins. Conclusions. The sulforaphane-mediated decrease of oxidative stress and activation of the Nrf2-ARE pathway may ameliorate bladder dysfunction caused by bladder outlet obstruction. PMID:27433291

Novel taspine derivative 12k inhibits cell growth and induces apoptosis in lung cell carcinoma.

PubMed

Dai, Bingling; Wang, Wenjie; Liu, Rui; Wang, Hongying; Zhang, Yanmin

2015-03-01

Taspine is an active compound in anticancer agent development. 12k was synthesized with taspine as lead compound bearing biphenyl scaffold and showed potent anticancer activity. Here, we investigated the effect of taspine derivative 12k on A549 lung cells. We showed that 12k not only decreased significantly A549 cell viability, A549 cell colony formation but also impaired A549 cell migration. Moreover, 12k treatment blocked cell cycle progression by increasing cell number in S phase to 42.80% for 6 μmol/L vs. 28.86% for control while decreasing cell number in G1 phase. Accordingly, this was associated with an increase protein expression of cyclin E and a decrease protein expression of cyclin D1, cyclin B1 and its associated CDK1 (cdc2). Meanwhile, we found that 12k induced A549 cell apoptosis, which was closely associated with the effect of the Bcl-2 family. Increase of Bad, Bak and Bax expression levels, decrease of Bcl-2 and Mcl-1 expression levels were observed. SiRNA knockdown of c-myc in A549 cells significantly attenuated tumor inhibition effects of 12k. In conclusion, our results demonstrate that 12k has an inhibitory effect on growth of A549 cell by inducing cell cycle arrest and apoptosis. Copyright © 2015 Elsevier Masson SAS. All rights reserved.

Prenatal Polycyclic Aromatic Hydrocarbon, Adiposity, Peroxisome Proliferator-Activated Receptor (PPAR) γ Methylation in Offspring, Grand-Offspring Mice

PubMed Central

Yan, Zhonghai; Zhang, Hanjie; Maher, Christina; Arteaga-Solis, Emilio; Champagne, Frances A.; Wu, Licheng; McDonald, Jacob D.; Yan, Beizhan; Schwartz, Gary J.; Miller, Rachel L.

2014-01-01

Rationale Greater levels of prenatal exposure to polycyclic aromatic hydrocarbon (PAH) have been associated with childhood obesity in epidemiological studies. However, the underlying mechanisms are unclear. Objectives We hypothesized that prenatal PAH over-exposure during gestation would lead to weight gain and increased fat mass in offspring and grand-offspring mice. Further, we hypothesized that altered adipose gene expression and DNA methylation in genes important to adipocyte differentiation would be affected. Materials and Methods Pregnant dams were exposed to a nebulized PAH mixture versus negative control aerosol 5 days a week, for 3 weeks. Body weight was recorded from postnatal day (PND) 21 through PND60. Body composition, adipose cell size, gene expression of peroxisome proliferator-activated receptor (PPAR) γ, CCAAT/enhancer-binding proteins (C/EBP) α, cyclooxygenase (Cox)-2, fatty acid synthase (FAS) and adiponectin, and DNA methylation of PPAR γ, were assayed in both the offspring and grand-offspring adipose tissue. Findings Offspring of dams exposed to greater PAH during gestation had increased weight, fat mass, as well as higher gene expression of PPAR γ, C/EBP α, Cox2, FAS and adiponectin and lower DNA methylation of PPAR γ. Similar differences in phenotype and DNA methylation extended through the grand-offspring mice. Conclusions Greater prenatal PAH exposure was associated with increased weight, fat mass, adipose gene expression and epigenetic changes in progeny. PMID:25347678

Secreted HoxA3 Promotes Epidermal Proliferation and Angiogenesis in Genetically Modified Three-Dimensional Composite Skin Constructs

PubMed Central

Kuo, Jennifer H.; Cuevas, Ileana; Chen, Amy; Dunn, Ashley; Kuri, Mauricio; Boudreau, Nancy

2014-01-01

Objective: Homeobox (HOX) transcription factors coordinate gene expression in wound repair and angiogenesis. Previous studies have shown that gene transfer of HoxA3 to wounds of diabetic mice accelerates wound healing, increasing angiogenesis and keratinocyte migration. In this study, we examined whether HoxA3 can also improve angiogenesis, epidermal integrity, and viability of composite skin grafts. Approach: To determine the effects of HoxA3 on composite skin grafts, we constructed bilayered composite grafts incorporating fibroblasts engineered to constitutively secrete HoxA3. We then transplanted these composite grafts in vivo. Results: The composite grafts produced a stratified epidermal layer after seventeen days in culture and following transplantation in vivo, these grafts exhibit normal epidermal differentiation and reduced contraction compared to controls. In addition, HoxA3 grafts showed increased angiogenesis. Quantitative polymerase chain reaction (PCR) analyses of HoxA3 graft tissue reveal an increase in the downstream HoxA3 target genes MMP-14 and uPAR expression, as well as a reduction in CCL-2 and CxCl-12. Innovation: Expression of secreted HoxA3 in composite grafts represents a comprehensive approach that targets both keratinocytes and endothelial cells to promote epidermal proliferation and angiogenesis. Conclusion: Secreted HoxA3 improves angiogenesis, reduces expression of inflammatory mediators, and prolongs composite skin graft integrity. PMID:25302136

Prevalence and outcomes of breast milk expressing in women with healthy term infants: a systematic review

PubMed Central

2013-01-01

Background Expressing breast milk has become increasingly prevalent, particularly in some developed countries. Concurrently, breast pumps have evolved to be more sophisticated and aesthetically appealing, adapted for domestic use, and have become more readily available. In the past, expressed breast milk feeding was predominantly for those infants who were premature, small or unwell; however it has become increasingly common for healthy term infants. The aim of this paper is to systematically explore the literature related to breast milk expressing by women who have healthy term infants, including the prevalence of breast milk expressing, reported reasons for, methods of, and outcomes related to, expressing. Methods Databases (Medline, CINAHL, JSTOR, ProQuest Central, PsycINFO, PubMed and the Cochrane library) were searched using the keywords milk expression, breast milk expression, breast milk pumping, prevalence, outcomes, statistics and data, with no limit on year of publication. Reference lists of identified papers were also examined. A hand-search was conducted at the Australian Breastfeeding Association Lactation Resource Centre. Only English language papers were included. All papers about expressing breast milk for healthy term infants were considered for inclusion, with a focus on the prevalence, methods, reasons for and outcomes of breast milk expression. Results A total of twenty two papers were relevant to breast milk expression, but only seven papers reported the prevalence and/or outcomes of expressing amongst mothers of well term infants; all of the identified papers were published between 1999 and 2012. Many were descriptive rather than analytical and some were commentaries which included calls for more research, more dialogue and clearer definitions of breastfeeding. While some studies found an association between expressing and the success and duration of breastfeeding, others found the opposite. In some cases these inconsistencies were compounded by imprecise definitions of breastfeeding and breast milk feeding. Conclusions There is limited evidence about the prevalence and outcomes of expressing breast milk amongst mothers of healthy term infants. The practice of expressing breast milk has increased along with the commercial availability of a range of infant feeding equipment. The reasons for expressing have become more complex while the outcomes, when they have been examined, are contradictory. PMID:24246046

Fasting Induces IL-1 Resistance and Free-Fatty Acid-Mediated Up-Regulation of IL-1R2 and IL-1RA

PubMed Central

Joesting, Jennifer J.; Moon, Morgan L.; Gainey, Stephen J.; Tisza, Brittany L.; Blevins, Neil A.; Freund, Gregory G.

2014-01-01

Objective: Weight-loss is a near societal obsession and many diet programs use significant calorie restriction including fasting/short term starvation to generate rapid effects. Fasting is also a well-recognized cause of immunosuppression especially within the innate immune system. In this study, we sought to determine if the IL-1 arm of the neuroimmune system was down-regulated by a 24 h fast and how fasting might generate this effect. Design: Mice were allowed ad libitum access to food or had food withheld for 24 h. Expression of the endogenous IL-1 antagonists, IL-1 receptor type 2 (IL-1R2), and IL-1 receptor antagonist (IL-1RA) was determined as were sickness behaviors before and after IL-1β administration. Results: Fasting markedly increased gene expression of IL-1R2 (83-fold in adipose tissue, 9.5-fold in liver) and IL-1RA (68-fold in liver). Fasted mice were protected from IL-1β-induced weight-loss, hypoglycemia, loss of locomotor, and social anxiety. These protections were coupled to a large positive interaction of fasting and IL-1β on IL-1R2 gene expression in adipose tissue and liver (2.6- and 1.6-fold, respectively). Fasting not only increased IL-1RA and IL-1R2 protein 2.5- and 3.2-fold, respectively, in liver but also increased IL-1R2 1.8-fold in adipose tissue. Fasting, in turn, triggered a 2.4-fold increase in plasma free-fatty acids (FFAs) and a 2.1-fold increase in plasma corticosterone. Inhibition, of glucocorticoid action with mifepristone did not impact fasting-dependent IL-1R2 or IL-1RA gene expression. Administration of the FFA, palmitate, to mice increased liver IL-1R2 and IL-1RA gene expression by 14- and 11-fold, respectively. Conclusion: These findings indicate that fasting augments expression of endogenous IL-1 antagonists inducing IL-1 resistance. Fasting-induced increases in plasma FFAs appears to be a signal that drives immunosuppression during fasting/short term starvation. PMID:25071776

Glucocorticoid receptor expression on circulating leukocytes in healthy and asthmatic adolescents in response to exercise

PubMed Central

Lu, Kim D.; Cooper, Dan; Haddad, Fadia; Zaldivar, Frank; Kraft, Monica; Radom-Aizik, Shlomit

2017-01-01

Background Poor aerobic fitness is associated with worsening of asthma symptoms and fitness training may improve asthma control. The mechanism linking fitness with asthma is not known. We hypothesized that repeated bouts of exercise would lead to a downregulation of glucocorticoid receptor (GR) expression on circulating leukocytes reflecting a reduced responsiveness to stress. Methods In a prospective exercise training intervention of healthy and asthmatic adolescents, GR expression in leukocytes was measured using flow cytometry in response to a brief exercise challenge before and after the training intervention. PBMC gene expression of GR, GRβ, HSP70, and TGFβ1, 2 were determined using RT-PCR. Results Peak V̇O2 increased by 14.6 ± 2.3% indicating an effective training (p<0.01). There was a significant difference in GR expression among leukocyte subtypes, with highest expression in eosinophils. Following the training intervention, there was a significant decrease in baseline GR expression (p<0.05) in leukocyte and monocyte subtypes in both healthy and asthmatic adolescents. Conclusions This is the first study in adolescents to show that exercise training reduces GR expression on circulating leukocytes. We speculate that exercise training downregulates the stress response in general, manifested by decreased GR expression, and may explain why improving fitness improves asthma health. PMID:28796240

Hepcidin suppression in β-thalassemia is associated with the down-regulation of atonal homolog 8.

PubMed

Upanan, Supranee; McKie, Andrew T; Latunde-Dada, Gladys O; Roytrakul, Sittiruk; Uthaipibull, Chairat; Pothacharoen, Peraphan; Kongtawelert, Prachya; Fucharoen, Suthat; Srichairatanakool, Somdet

2017-08-01

Atonal homolog 8 (ATOH8) is defined as a positive regulator of hepcidin transcription, which links erythropoietic activity with iron-sensing molecules. In the present study, we investigated the association between hepcidin and ATOH8 expression in β-thalassemia. We found that inhibition of hepcidin expression in β-thalassemia is correlated with reduced ATOH8 expression. Hepatic hepcidin 1 (Hamp1) and Atoh8 mRNA expression were down-regulated in β-thalassemic mice. Hepcidin (HAMP) and ATOH8 mRNA expression were consistently suppressed in Huh7 cells cultured in medium supplemented with β-thalassemia patient serum. The Huh7 cells, which were transfected with ATOH8-FLAG expression plasmid and cultured in the supplemented medium, exhibited increased levels of ATOH8 mRNA, ATOH8-FLAG protein, pSMAD1,5,8, and HAMP mRNA. Interestingly, over-expression of ATOH8 reversed the effects of hepcidin suppression induced by the β-thalassemia patient sera. In conclusion, hepcidin suppression in β-thalassemia is associated with the down-regulation of ATOH8 in response to anemia. We, therefore, suggest that ATOH8 is an important transcriptional regulator of hepcidin in β-thalassemia.

Evaluating the consistency of gene sets used in the analysis of bacterial gene expression data.

PubMed

Tintle, Nathan L; Sitarik, Alexandra; Boerema, Benjamin; Young, Kylie; Best, Aaron A; Dejongh, Matthew

2012-08-08

Statistical analyses of whole genome expression data require functional information about genes in order to yield meaningful biological conclusions. The Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) are common sources of functionally grouped gene sets. For bacteria, the SEED and MicrobesOnline provide alternative, complementary sources of gene sets. To date, no comprehensive evaluation of the data obtained from these resources has been performed. We define a series of gene set consistency metrics directly related to the most common classes of statistical analyses for gene expression data, and then perform a comprehensive analysis of 3581 Affymetrix® gene expression arrays across 17 diverse bacteria. We find that gene sets obtained from GO and KEGG demonstrate lower consistency than those obtained from the SEED and MicrobesOnline, regardless of gene set size. Despite the widespread use of GO and KEGG gene sets in bacterial gene expression data analysis, the SEED and MicrobesOnline provide more consistent sets for a wide variety of statistical analyses. Increased use of the SEED and MicrobesOnline gene sets in the analysis of bacterial gene expression data may improve statistical power and utility of expression data.

Silencing of BAG3 inhibits the epithelial-mesenchymal transition in human cervical cancer.

PubMed

Song, Fei; Wang, Geng; Ma, Zhifang; Ma, Yuebing; Wang, Yingying

2017-11-10

Bcl2-associated athanogene 3 (BAG3) has been reported to be involved in aggressive progression of many tumors. In the present study, we examined the expression of BAG3 in human cervical cancer (CC) tissues and investigated the role of BAG3 in SiHa and HeLa cell growth, migration, and invasion. Here, we found that most of CC tissues highly expressed the protein and mRNA of BAG3, while their expression was obviously lower in paired normal tissues (all p<0.001). BAG3 expression was associated with FIGO stage and metastasis (all p<0.05). In-vitro analysis demonstrated that BAG3 siRNAs inhibited SiHa and HeLa cell growth, invasion and migration. Mechanically, BAG3 siRNAs inhibited the expression of EMT-regulating markers, involving MMP2, Slug and N-cadherin, and increased the expression of E-cadherin. In a xenograft nude model, BAG3 siRNAs inhibited tumor growth and the expression of EMT biomarkers. In conclusion, BAG3 is involved in the EMT process, including cell growth, invasion and migration in the development of CC. Thus, BAG3 target might be recommended as a novel therapeutic approach.

Silencing of BAG3 inhibits the epithelial-mesenchymal transition in human cervical cancer

PubMed Central

Song, Fei; Wang, Geng; Ma, Zhifang; Ma, Yuebing; Wang, Yingying

2017-01-01

Bcl2-associated athanogene 3 (BAG3) has been reported to be involved in aggressive progression of many tumors. In the present study, we examined the expression of BAG3 in human cervical cancer (CC) tissues and investigated the role of BAG3 in SiHa and HeLa cell growth, migration, and invasion. Here, we found that most of CC tissues highly expressed the protein and mRNA of BAG3, while their expression was obviously lower in paired normal tissues (all p<0.001). BAG3 expression was associated with FIGO stage and metastasis (all p<0.05). In-vitro analysis demonstrated that BAG3 siRNAs inhibited SiHa and HeLa cell growth, invasion and migration. Mechanically, BAG3 siRNAs inhibited the expression of EMT-regulating markers, involving MMP2, Slug and N-cadherin, and increased the expression of E-cadherin. In a xenograft nude model, BAG3 siRNAs inhibited tumor growth and the expression of EMT biomarkers. In conclusion, BAG3 is involved in the EMT process, including cell growth, invasion and migration in the development of CC. Thus, BAG3 target might be recommended as a novel therapeutic approach. PMID:29221135

Lipocalin-2 Promotes Endoplasmic Reticulum Stress and Proliferation by Augmenting Intracellular Iron in Human Pulmonary Arterial Smooth Muscle Cells

PubMed Central

Wang, Guoliang; Liu, Shenghua; Wang, Li; Meng, Liukun; Cui, Chuanjue; Zhang, Hao; Hu, Shengshou; Ma, Ning; Wei, Yingjie

2017-01-01

Endoplasmic reticulum (ER) stress, a feature of many conditions associated with pulmonary hypertension (PH), is increasingly recognized as a common response to promote proliferation in the walls of pulmonary arteries. Increased expression of Lipocalin-2 in PH led us to test the hypothesis that Lipocalin-2, a protein known to sequester iron and regulate it intracellularly, might facilitate the ER stress and proliferation in pulmonary arterial smooth muscle cells (PASMCs). In this study, we observed greatly increased Lcn2 expression accompanied with increased ATF6 cleavage in a standard rat model of pulmonary hypertension induced by monocrotaline. In cultured human PASMCs, Lcn2 significantly promoted ER stress (determined by augmented cleavage and nuclear localization of ATF6, up-regulated transcription of GRP78 and NOGO, increased expression of SOD2, and mild augmented mitochondrial membrane potential) and proliferation (assessed by Ki67 staining and BrdU incorporation). Lcn2 promoted ER stress accompanied with augmented intracellular iron levels in human PASMCs. Treatment human PASMCs with FeSO4 induced the similar ER stress and proliferation response and iron chelator (deferoxamine) abrogated the ER stress and proliferation induced by Lcn2 in cultured human PASMCs. In conclusion, Lcn2 significantly promoted human PASMC ER stress and proliferation by augmenting intracellular iron. The up-regulation of Lcn2 probably involved in the pathogenesis and progression of PH. PMID:28255266