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Objective To investigate the effects of hepatocyte growth factor (HGF) on proliferation and transdifferentiation of human Tenon capsule fibroblasts induced by transforming growth factor-β1(TGF-β1) in vitro.Methods Human Tenon capsule fibroblasts were cultured and divided into blank control group,TGF-β1 treated group and different concentrations HGF+TGF-β1 groups.The TGF-β1 at 10 μg/L was added into culture medium of the TGF-β1 treated group,and different concentrations of HGF (25,50,100,200 μg/L) and 10 μg/L TGF-β1 were added into culture medium of the HGF25 μg/L +TGF-β1,HGF50 μg/L +TGF-β1,HGF100 μg/L +TGF-β1,HGF200 μg/L +TGF-β1 group respectively,Methyl thiazolyl tetrazolium (MTT) was employed to measure the cell proliferation (absorbance at 560 nm).Immunofluorescence staining was used to evaluate and locate the expression of α-smooth muscle action (α-SMA) in the cells.The expression of α-SMA protein in the cells was detected by Western blot assay.Results Cultured cells showed fusiform in shape with the positive response for vimentin.The proliferation value of the cells was 0.203±0.025,0.497 ± 0.101,0.426 ± 0.062,0.354 ± 0.040,0.272 ± 0.084,0.241 ± 0.011 in the blank control group,TGF-β1 treated group,HGF25 μg/L +TGF-β1 group,HGF50 μg/L +TGF-β1 group,HGF100 μg/L +TGF-β group and HGF200 μg/L + TGF-β1 group,respectively,showing a significant difference among the groups (F =9.21 0,P =0.003).Compared with the TGF-β1 treated group,the proliferation values of the cells were significantly reduced in the blank control group and HGF+TGF-β1 group (all at P＜0.05).Immunofiuorescence staining showed that α-SMA protein mainly located in cytoplasm with the strong red fluorescence in the cells of the TGF-β1 treated group and weak red fluorescence in HGF+TGF-β1 group,and the expression of α-SMA was absent in the blank control group.The percentage of α-SMA-positive cells was (60.0±4.7)％ in the TGF-β1 treated group and (14.3±3.1)％ in the HGF+TGF-β1 group,with significant difference between the two groups (t =19.856,P＜0.001).The relative expression levels of the α-SMA protein in the cells were 0.642 ±0.032,1.330± 0.069 and 0.884 ±0.040 in the blank control group,TGF-β1 group and HGF100μg/L +TGF-β1 group,respectively,showing a significant difference among the groups (F =13.370,P＜ 0.001),and relative expression levels of the α-SMA protein in the cells were significantly lower in the blank controlgroup and HGF100 μg/L+TGF-β1 group than that in the TGF-β1 treated group (all at P＜0.05).Conclusions HGF can inhibit the proliferation of human Tenon capsule fibroblasts,down-regulate the expression of α-SMA protein induced by TGF-β1 and arrest the phenotype transformation of fibroblasts in vitro.
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Objective To investigate the effects of hepatocyte growth factor ( HGF ) on proliferation and transdifferentiation of human Tenon capsule fibroblasts induced by transforming growth factor .β1(TGF.β1) in vitro. Methods Human Tenon capsule fibroblasts were cultured and divided into blank control group, TGF.β1 treated group and different concentrations HGF+TGF.β1 groups. The TGF.β1 at 10 μg/L was added into culture medium of the TGF.β1 treated group,and different concentrations of HGF (25,50,100,200 μg/L) and 10μg/L TGF.β1 were added into culture medium of the HGF25μg/L+TGF.β1 ,HGF50μg/L+TGF.β1 ,HGF100μg/L+TGF.β1 ,HGF200μg/L+TGF.β1 group respectively,Methyl thiazolyl tetrazolium ( MTT) was employed to measure the cell proliferation ( absorbance at 560 nm) . Immunofluorescence staining was used to evaluate and locate the expression of α.smooth muscle action (α.SMA) in the cells. The expression ofα.SMA protein in the cells was detected by Western blot assay. Results Cultured cells showed fusiform in shape with the positive response for vimentin. The proliferation value of the cells was 0. 203±0. 025,0. 497±0. 101,0. 426±0. 062,0. 354±0. 040,0. 272±0. 084,0. 241±0. 011 in the blank control group, TGF.β1 treated group,HGF25μg/L+TGF.β1 group,HGF50μg/L+TGF.β1 group,HGF100μg/L+TGF.β1 group and HGF200μg/L+TGF.β1 group,respectively,showing a significant difference among the groups (F=9. 210,P=0. 003). Compared with the TGF.β1 treated group,the proliferation values of the cells were significantly reduced in the blank control group and HGF+TGF.β1 group ( all at P<0. 05 ) . Immunofluorescence staining showed that α.SMA protein mainly located in cytoplasm with the strong red fluorescence in the cells of the TGF.β1 treated group and weak red fluorescence in HGF+TGF.β1 group,and the expression of α.SMA was absent in the blank control group. The percentage of α.SMA.positive cells was ( 60. 0 ± 4. 7 )％ in the TGF.β1 treated group and ( 14. 3 ± 3. 1 )％ in the HGF+TGF.β1 group, with significant difference between the two groups (t=19. 856,P<0. 001). The relative expression levels of the α.SMA protein in the cells were 0. 642±0. 032,1. 330±0. 069 and 0. 884±0. 040 in the blank control group,TGF.β1 group and HGF100μg/L+TGF.β1 group, respectively, showing a significant difference among the groups ( F=13. 370, P<0. 001),and relative expression levels of the α.SMA protein in the cells were significantly lower in the blank control group and HGF100μg/L+TGF.β1 group than that in the TGF.β1 treated group (all at P<0. 05). Conclusions HGF can inhibit the proliferation of human Tenon capsule fibroblasts, down.regulate the expression of α.SMA protein induced by TGF.β1 and arrest the phenotype transformation of fibroblasts in vitro.
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Objective@#To investigate the effect of curcumin on intestinal mucosal mechanical barrier in rats with non-alcoholic fatty liver disease.@*Methods@#A total of 30 male Sprague-Dawley rats were equally divided into normal control group, model group, and curcumin intervention group. The rats in the model group and the curcumin intervention group were given high-fat feed for 16 weeks, and those in the curcumin intervention group were given curcumin 200 mg/kg/day by gavage once a day after 8 weeks of high-fat feeding. The rats were sacrificed at the end of week 16. A light microscope was used to observe pathological changes in the liver, an electron microscope was used to observe the tight junction of the intestinal mucosa, an automatic biochemical analyzer was used to measure the serum levels of alanine aminotransferase (ALT) and aspartate aminotransferase (AST), chromogenic substrate Limulus amebocyte lysate assay was used to measure plasma lipopolysaccharide (LPS) level, spectrophotometric method was used to measure the activity of serum diamine oxidase, ELISA was used to measure the serum level of tumor necrosis factor-α (TNFα), and immunohistochemistry was used to measure the expression of the tight junction protein occludin. One-way ANOVA test and SNK-q test were used for statistical analysis.@*Results@#Under the light microscope, the control group had no hepatocyte steatosis, the model group had significant hepatocyte steatosis and inflammatory cell infiltration, and the curcumin intervention group had reduced hepatocyte steatosis and inflammatory cell infiltration. Under the electron microscope, the control group had a clear and complete structure of the tight junction of the intestinal mucosa and normal structures of mitochondria and endoplasmic reticulum; in the model group, the structure of the tight junction of the intestinal mucosa was destroyed, the intercellular space was widened, the desmosomes had a loose structure, there was edema in some mitochondria, and the endoplasmic reticulum was dilated; the curcumin intervention group had improvements in the structure of tight junction of the intestinal mucosa, intercellular space, edema in the mitochondria, and dilation of the endoplasmic reticulum. Compared with the control group, the model group had significant increases in the serum levels of AST, ALT, DAO, TNFα, and LPS (q = -15.918, -14.402, -33.700, -8.944, and -10.832, P < 0.05); compared with the model group, the curcumin intervention group had significant reductions in the serum levels of AST, ALT, DAO, TNFα, and LPS (q = 10.457, 7.752, 18.802, 5.202, and 4.279, P < 0.05). In the control group, occludin showed a linear distribution along the top of small intestinal mucosal epithelial cells. The model group had a significant reduction in positive staining compared with the control group, and the curcumin intervention group had a significant increase in positive staining compared with the model group. The relative expression of occludin was 0.29±0.03 in the control group, 0.12±0.02 in the model group, and 0.21±0.02 in the curcumin intervention group (P < 0.05).@*Conclusion@#Intestinal mucosal mechanical barrier is impaired in rats with NAFLD. Curcumin can reduce such damage, and its mechanism of action may be related to up-regulating the expression of occludin in the intestinal mucosa and reducing the levels of TNFα and LPS.
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Objective To explore the related risk factors for diabetic retinopathy(DR)in type 2diabetes.Methods The clinical data of 412 type 2 diabetes patients,diagnosed between 2003 and 2010,were analyzed retrospectively.The diagnosis of DR and proliferative diabetic retinopathy(PDR)was confirmed by ophthalmoloseopy and fundus fluorescein angiography.Glycated hemoglobin Alc,glucose,insulin,and C-peptide of fasting plasma,and 1,2 and 3 hours postprandial plasma were measured.According to the above-mentioned data,get the fluctuation of glucose,insulin and C-peptide of 1,2 and 3hour postprandial plasma.Results The morbidity of DR and PDR increased following the longer disease duration.Age,diabetic duration,body mass index(BMI),hypertension grade,HbAlC,fasting plasma insulin and C-peptide,2 and 3 hours postprandial plasma glucose,1 and 2 hours postprandial plasma insulin,1,2 and 3 hour postprandial plasma C-peptide,1,2 and 3 hours postprandial plasma glucose,insulin and C-peptide fluctuation are different statistically among non-DR group,non-PDR group and PDR group(P＜0.05).3 hours postprandial plasma glucose and fasting plasma insulin were risk factors of DR (P＜0.05).Conclusions Postprandial plasma glucose and fasting plasma insulin were risk factors of DR.Nevertheless,postprandial insulin,fasting and postprandial C-peptide,postprandial plasma glucose,insulin and C-peptide fluctuation were useful for DR diagnosis.
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Objective To investigate the change of epithelium growth factor(EGF), tumor necrosis factor (TNF), superoxide dismutese(SOD) and hemorrheology (HL) in patients with diabetic retinopathy.Methods EGF、TNF、SOD levels and HL changes have been measured in 30 NIDDM patients with retinopathy (DR group), 29 NIDDM patients without retinopathy (NDR group), and 29 patients with senile cataract and without diabetes as control (C group). Results The mean levels of EGF、TNF and blood viscosity were significantly higher in both DR and NDR group than those in C group; the mean levels of SOD was significantly lower in both DR and NDR group than that in C group. The EGF、TNF and blood viscosity was significantly higher in proliferative type DR than in background DR; The SOD was significantly lower in proliferative type DR than in background DR.Conclusion The development of diabetic retinopathy is related to the increase of EGF、TNF、blood viscosity and decrease of serum SOD.