AIM：To research the effect of Y-27632,a selective Rhoassociated coiled-coil kinase（ROCK） inhibitor,on TGF-β1/Smad2,3 signal transduction in ocular Tenon＇s capsule fi broblasts（OTFs）.METHODS：Primary ocular Teno...AIM：To research the effect of Y-27632,a selective Rhoassociated coiled-coil kinase（ROCK） inhibitor,on TGF-β1/Smad2,3 signal transduction in ocular Tenon＇s capsule fi broblasts（OTFs）.METHODS：Primary ocular Tenon＇s capsule fibroblasts had been cultured in vitro.The effect of Y27632 on proliferation of OTF stimulated by lysophosphatidic acid（LPA） was evaluated by MTT colorimetric assay so as to sift out the proper concentrations range of Y-27632 for the next experiment.Real time-polymerase chain reactor（RT-PCR） was to analyze the changes of Smad2 and Smad3 genes of cells affected by Y-27632,though unaffected by transforming growth factorbeta1（TGF-β1）.Proteins of Smad2,Smad3,phosphorylated Smad2（Ser245/250/255）,and phosphorylated Smad3（Ser423/425/203） were respectively quantifi ed by Western blot after OTFs were successively incubated by TGF-β1 and Y-27632.Meanwhile,α-smooth muscular actin（α-SMA） protein was also quantified after the small intervening gene fragments of human Smad2 and Smad3 were designed,synthesized,and then transfected to OTFs.RESULTS：Y-27632 signifi cantly inhibited OTFs proliferation stimulated by LPA.Also Y-27632 signifi cantly suppressed the expressions of Smad2 m RNA,Smad2,3 proteins expressions,Smad3 phosphorylation at the carboxylic terminals of Ser423/425/203 which had been radically promoted by TGF-β1.Si RNA-Smad2,3 suppressed α-SMA expressions,but less effectively than Y-27632.CONCLUSION：The inhibition of ROCK signaling may be a potential therapeutic candidate for the treatment of the fi ltration channel fi brosis.展开更多
In order to elucidate the effect of dexamethasone on the expression of transforming growth factor-betal （TGF-β1） in ciliary pigment epithelial （CPE） cells cultured in vitro, rabbit CPE cells were cultured in vitr...In order to elucidate the effect of dexamethasone on the expression of transforming growth factor-betal （TGF-β1） in ciliary pigment epithelial （CPE） cells cultured in vitro, rabbit CPE cells were cultured in vitro, treated with DMEM medium containing 0, 1 × 10^-8 , 5 × 10^-8 , 10 × 10^-8 and 50 × 10^-8 mol/L dexamethasone respectively for 5 days. The TGF-131 expression was detected by immunohistochemistry Supervision methods and analyzed semi-quantitatively by HMIAS-2000 image system. As opposed to in vivo, rabbit CPE cells expressed TGF-131 under cul- tured circumstance in vitro. The gray scales of the positive yellow staining in the groups of 1 × 10^-8 , 5 × 10^-8 , 10 × 10^-8 and 50 × 10^-8 mol/L dexamethasone were 136.57 ± 4.43, 140.20 ± 6. 10, 142. 98± 2. 99, 146.80±1.68 and 150.05 × 1.94 respectively. When the concentrations of dexamethasone were equal to or higher than 5 × 10^-8mol/L and, the expression of TGF-β1 was inhibited. 10^-7 mol/L dexamethasone showed a significant inhibition. It was suggested that CPE cells possess the potential ability of synthesizing and expressing TGF-β1. The inhibition of TGF-β1 expression by dexamethasone may be beneficial to the treatment of proliferative vitroretinopathy, also exert some influence on the secretion of aqueous humor and ciliary inflammation.展开更多
To investigate the effects of IL-5 on the expression of TGF-β1 in eosinophils in vitro, eosinophils were incubated in the presence of the same concentrations of IL-4, IL-5 and IFNγ, different concentrations of IL-5 ...To investigate the effects of IL-5 on the expression of TGF-β1 in eosinophils in vitro, eosinophils were incubated in the presence of the same concentrations of IL-4, IL-5 and IFNγ, different concentrations of IL-5 in yitro and changes of eosinophil viability were assessed by trypan blue exclusion. Non-cytokine was employed as a negative control. 16 h after the cultivation, supernatants and cells were assayed by using TGF-β1 specific ELISA and RT-PCR. The mRNA expression and protein expresssion of TGF-β1 in eosinophils stimulated with different eytokines was observed. The expression of TGF-β1 protein in eosinophils was increased significantly by IL-4 （433. 67 ±9.86 vs 228.9±2.87） and IL-5 （403.7±7.60 vs 228.9±2.87, P〈0.05）, while decreased by IFNγ （178. 47±2. 60 vs 228.9±2.87）. At the same time, the results demonstrated that the basal level of TGF expression was enhanced by IL-5 in all samples （P〈0.05）. The expression of TGF-β1 mR- NA was 1.42, 1.70, 1.76-folds higher than that of the non-stimulated controls. It is concluded that IL-5 can up-regulate the expression of TGF-β1 in eosinophils in vitro, which might have effect in eosinophil-associated chronic rejection.展开更多
Background： Transforming growth factor-beta 1 （TGF-β 1） and gene variants have been extensively studied in various human diseases. For example, TGF-β1 polymorphisms were associated with fibrosis and pneumoconiosi...Background： Transforming growth factor-beta 1 （TGF-β 1） and gene variants have been extensively studied in various human diseases. For example, TGF-β1 polymorphisms were associated with fibrosis and pneumoconiosis, but the data remained controversial. The aim of this meta-analysis was to assess the association between TGF-β1-509 C〉T [rs 1800469], ＋869 T〉C [rs 1800470], and ＋915 G〉C [rs 1800471 ] polymorphisms and pneumoconiosis, Methods： A comprehensive literature search was conducted through searching in PubMed, Embase, the Chinese Biomedical Database, and the Wei Pu （Chinese） Database by the end of April 2016. Eleven publications with 21 studies were included in this recta-analysis, covering a total of 4333 patients with pneumoconiosis and 3478 controls. Study quality was assessed, and heterogeneity and publication bias were measured. All statistical analyses were performed using STATA version 12.0 （StataCorp, College Station, TX, USA） software. Results： The data showed significant associations between TGF-β1-509 C〉T polymorphism and the risk ofpneumoconiosis development （T vs. C, odds ratio [OR] = 1.35, 95% confidence interval [CI]： 1.00-1.81, P = 0.046）; between TGF-fll ＋915 G〉C polymorphism and the pneumoconiosis risk （C vs. G, OR = 1.69, 95% CI： 1.19-2.40, P = 0.004; CG vs. GG, OR = 1.79, 95% CI： 1.23-2.60, P = 0.002; CC＋CG vs. GG, OR = 1.80, 95% CI： 1.24-2.61, P = 0.002）. In addition, the subgroup analysis of ethnicity versus pneumoconiosis types indicated a significant association of silicosis among Asian populations but not that of coal workers＇ pneumoconiosis in Caucasian populations. In contrast, no significant association was exhibited between TGF-β1 ＋869 T〉C polymorphism and risk ofpneumoconiosis. Conclusion： The polymorphisms of both TGF-β1 -509 C〉T and ＋915 G〉C are associated with increased risk of pneumoconiosis. Key words： Meta analysis; Pneumoconiosis; Polymorphism; Transforming Growth Factor-betal展开更多
Background Hepatic fibrosis is the key stage of the pathological progress from hepatic injury to cirrhosis. Ursodeoxycholic acid （UDCA） has been known as having significant clinical therapeutic effects on chronic li...Background Hepatic fibrosis is the key stage of the pathological progress from hepatic injury to cirrhosis. Ursodeoxycholic acid （UDCA） has been known as having significant clinical therapeutic effects on chronic liver diseases. Our research aimed to study the effect of UDCA on the signaling pathway of transforming growth factor beta1 （TGFβ1）/Smad and discuss its possible molecular mechanisms of inhibiting hepatic fibrosis. Methods Rat hepatic stellate cells were cultured in vitro and randomly assigned to 4 groups. Group A was control group with only DMEM culture medium applied, and groups B, C, D were experimental groups, with different doses of UDCA （1.0 mmol/L, 0.5 mmol/L and 0.25 mmol/L respectively） added into their DMEM culture medium for further culture of 24 hours and 48 hours. The protein expressions of TGFβ1, TGF type 1 receptor, Smad3, Smad4 and Smad7 were measured by Western blotting, as well as the expressions of TGFβ1, Smad3, Smad7 and cAMP response element （CREB） binding protein （CBP） mRNA by real-time PCR. SPSS 11.5 statistical package was adopted for data analyses. Results Compared with control group, the mRNA expressions of TGFβ1 in the high and middle UDCA dose groups for 24 hours and 48 hours significantly decreased （P 〈0.05）, the protein expressions of TGFβ1 in the two above groups for 48 hours and in the high dose group for 24 hours significantly decreased （P 〈0.05）. The protein and mRNA expressions of Smad3 in each UDCA dose group for 24 hours and 48 hours significantly decreased, with significant difference among different UDCA dose groups and between that of 24 hours and 48 hours observed （P 〈0.05）. The protein and mRNA expressions of Smad7 in the high and middle UDCA dose groups for 24 hours and 48 hours significantly increased. The CBP mRNA expression in each UDCA dose group for 24 hours and 48 hours significantly decreased （P 〈0.05）, with significant difference among different UDCA dose groups observed （P 〈0.05）. Conclusion UDCA could curb the development of hepatic fibrosis through affecting the signaling pathway of TGFβ1/Smad by inhibiting the expressions of TGFβ1, Smad3 and CBP and increasing the expression of Smad7.展开更多
基金Supported by Scientific and Technological Project of Shaanxi Province,China(No.2016SF-010)
文摘AIM：To research the effect of Y-27632,a selective Rhoassociated coiled-coil kinase（ROCK） inhibitor,on TGF-β1/Smad2,3 signal transduction in ocular Tenon＇s capsule fi broblasts（OTFs）.METHODS：Primary ocular Tenon＇s capsule fibroblasts had been cultured in vitro.The effect of Y27632 on proliferation of OTF stimulated by lysophosphatidic acid（LPA） was evaluated by MTT colorimetric assay so as to sift out the proper concentrations range of Y-27632 for the next experiment.Real time-polymerase chain reactor（RT-PCR） was to analyze the changes of Smad2 and Smad3 genes of cells affected by Y-27632,though unaffected by transforming growth factorbeta1（TGF-β1）.Proteins of Smad2,Smad3,phosphorylated Smad2（Ser245/250/255）,and phosphorylated Smad3（Ser423/425/203） were respectively quantifi ed by Western blot after OTFs were successively incubated by TGF-β1 and Y-27632.Meanwhile,α-smooth muscular actin（α-SMA） protein was also quantified after the small intervening gene fragments of human Smad2 and Smad3 were designed,synthesized,and then transfected to OTFs.RESULTS：Y-27632 signifi cantly inhibited OTFs proliferation stimulated by LPA.Also Y-27632 signifi cantly suppressed the expressions of Smad2 m RNA,Smad2,3 proteins expressions,Smad3 phosphorylation at the carboxylic terminals of Ser423/425/203 which had been radically promoted by TGF-β1.Si RNA-Smad2,3 suppressed α-SMA expressions,but less effectively than Y-27632.CONCLUSION：The inhibition of ROCK signaling may be a potential therapeutic candidate for the treatment of the fi ltration channel fi brosis.
文摘In order to elucidate the effect of dexamethasone on the expression of transforming growth factor-betal （TGF-β1） in ciliary pigment epithelial （CPE） cells cultured in vitro, rabbit CPE cells were cultured in vitro, treated with DMEM medium containing 0, 1 × 10^-8 , 5 × 10^-8 , 10 × 10^-8 and 50 × 10^-8 mol/L dexamethasone respectively for 5 days. The TGF-131 expression was detected by immunohistochemistry Supervision methods and analyzed semi-quantitatively by HMIAS-2000 image system. As opposed to in vivo, rabbit CPE cells expressed TGF-131 under cul- tured circumstance in vitro. The gray scales of the positive yellow staining in the groups of 1 × 10^-8 , 5 × 10^-8 , 10 × 10^-8 and 50 × 10^-8 mol/L dexamethasone were 136.57 ± 4.43, 140.20 ± 6. 10, 142. 98± 2. 99, 146.80±1.68 and 150.05 × 1.94 respectively. When the concentrations of dexamethasone were equal to or higher than 5 × 10^-8mol/L and, the expression of TGF-β1 was inhibited. 10^-7 mol/L dexamethasone showed a significant inhibition. It was suggested that CPE cells possess the potential ability of synthesizing and expressing TGF-β1. The inhibition of TGF-β1 expression by dexamethasone may be beneficial to the treatment of proliferative vitroretinopathy, also exert some influence on the secretion of aqueous humor and ciliary inflammation.
文摘To investigate the effects of IL-5 on the expression of TGF-β1 in eosinophils in vitro, eosinophils were incubated in the presence of the same concentrations of IL-4, IL-5 and IFNγ, different concentrations of IL-5 in yitro and changes of eosinophil viability were assessed by trypan blue exclusion. Non-cytokine was employed as a negative control. 16 h after the cultivation, supernatants and cells were assayed by using TGF-β1 specific ELISA and RT-PCR. The mRNA expression and protein expresssion of TGF-β1 in eosinophils stimulated with different eytokines was observed. The expression of TGF-β1 protein in eosinophils was increased significantly by IL-4 （433. 67 ±9.86 vs 228.9±2.87） and IL-5 （403.7±7.60 vs 228.9±2.87, P〈0.05）, while decreased by IFNγ （178. 47±2. 60 vs 228.9±2.87）. At the same time, the results demonstrated that the basal level of TGF expression was enhanced by IL-5 in all samples （P〈0.05）. The expression of TGF-β1 mR- NA was 1.42, 1.70, 1.76-folds higher than that of the non-stimulated controls. It is concluded that IL-5 can up-regulate the expression of TGF-β1 in eosinophils in vitro, which might have effect in eosinophil-associated chronic rejection.
文摘Background： Transforming growth factor-beta 1 （TGF-β 1） and gene variants have been extensively studied in various human diseases. For example, TGF-β1 polymorphisms were associated with fibrosis and pneumoconiosis, but the data remained controversial. The aim of this meta-analysis was to assess the association between TGF-β1-509 C〉T [rs 1800469], ＋869 T〉C [rs 1800470], and ＋915 G〉C [rs 1800471 ] polymorphisms and pneumoconiosis, Methods： A comprehensive literature search was conducted through searching in PubMed, Embase, the Chinese Biomedical Database, and the Wei Pu （Chinese） Database by the end of April 2016. Eleven publications with 21 studies were included in this recta-analysis, covering a total of 4333 patients with pneumoconiosis and 3478 controls. Study quality was assessed, and heterogeneity and publication bias were measured. All statistical analyses were performed using STATA version 12.0 （StataCorp, College Station, TX, USA） software. Results： The data showed significant associations between TGF-β1-509 C〉T polymorphism and the risk ofpneumoconiosis development （T vs. C, odds ratio [OR] = 1.35, 95% confidence interval [CI]： 1.00-1.81, P = 0.046）; between TGF-fll ＋915 G〉C polymorphism and the pneumoconiosis risk （C vs. G, OR = 1.69, 95% CI： 1.19-2.40, P = 0.004; CG vs. GG, OR = 1.79, 95% CI： 1.23-2.60, P = 0.002; CC＋CG vs. GG, OR = 1.80, 95% CI： 1.24-2.61, P = 0.002）. In addition, the subgroup analysis of ethnicity versus pneumoconiosis types indicated a significant association of silicosis among Asian populations but not that of coal workers＇ pneumoconiosis in Caucasian populations. In contrast, no significant association was exhibited between TGF-β1 ＋869 T〉C polymorphism and risk ofpneumoconiosis. Conclusion： The polymorphisms of both TGF-β1 -509 C〉T and ＋915 G〉C are associated with increased risk of pneumoconiosis. Key words： Meta analysis; Pneumoconiosis; Polymorphism; Transforming Growth Factor-betal
文摘Background Hepatic fibrosis is the key stage of the pathological progress from hepatic injury to cirrhosis. Ursodeoxycholic acid （UDCA） has been known as having significant clinical therapeutic effects on chronic liver diseases. Our research aimed to study the effect of UDCA on the signaling pathway of transforming growth factor beta1 （TGFβ1）/Smad and discuss its possible molecular mechanisms of inhibiting hepatic fibrosis. Methods Rat hepatic stellate cells were cultured in vitro and randomly assigned to 4 groups. Group A was control group with only DMEM culture medium applied, and groups B, C, D were experimental groups, with different doses of UDCA （1.0 mmol/L, 0.5 mmol/L and 0.25 mmol/L respectively） added into their DMEM culture medium for further culture of 24 hours and 48 hours. The protein expressions of TGFβ1, TGF type 1 receptor, Smad3, Smad4 and Smad7 were measured by Western blotting, as well as the expressions of TGFβ1, Smad3, Smad7 and cAMP response element （CREB） binding protein （CBP） mRNA by real-time PCR. SPSS 11.5 statistical package was adopted for data analyses. Results Compared with control group, the mRNA expressions of TGFβ1 in the high and middle UDCA dose groups for 24 hours and 48 hours significantly decreased （P 〈0.05）, the protein expressions of TGFβ1 in the two above groups for 48 hours and in the high dose group for 24 hours significantly decreased （P 〈0.05）. The protein and mRNA expressions of Smad3 in each UDCA dose group for 24 hours and 48 hours significantly decreased, with significant difference among different UDCA dose groups and between that of 24 hours and 48 hours observed （P 〈0.05）. The protein and mRNA expressions of Smad7 in the high and middle UDCA dose groups for 24 hours and 48 hours significantly increased. The CBP mRNA expression in each UDCA dose group for 24 hours and 48 hours significantly decreased （P 〈0.05）, with significant difference among different UDCA dose groups observed （P 〈0.05）. Conclusion UDCA could curb the development of hepatic fibrosis through affecting the signaling pathway of TGFβ1/Smad by inhibiting the expressions of TGFβ1, Smad3 and CBP and increasing the expression of Smad7.
