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Objective To investigate whether the interleukin-10 (IL-10) gene therapy has the effect of anti liver fibrosis in mice and its mechanism.Methods Liver fibrosis was induced by long-term thioacetamide administration in mice.Human IL-10 expression plasmid was delivered via electroporation after liver fibrosis was established.The immunohistochemistry was used to study the expression of IL-10 in liver.Sircol collagen determination method was used to detect the contents of collagen in the liver.The reverse transcription polymerase chain reaction (RT-PCR) was used to detect the expressions of liver fibrosis-related genes,including transforming growth factor-β1 (TGF-β1),collagen α1,tumor necrosis factor-α (TNF-α),intercellular adhesion molecule-1 (ICAM-1),fibronectin,vascular cell adhesion molecule 1 (VCAM-1),and matrix metalloproteinase-inhibiting factor (TIMP-1,TIMP-2).Results Immunohistochemical results showed IL-10 gene therapy reversed hepatic fibrosis.Sircol collagen assay showed that IL-10 gene therapy reduced the content of collagen fibers(P ＜ 0.05).RT-PCR revealed IL-10 gene therapy reduced liver TGF-β1,TNF-α,collagen α1,cell adhesion molecule,and TIMPs mRNA upregulation.Conclusions Electroporative IL-10 gene therapy might be an effective therapeutic reagent for liver fibrosis with potential future clinical applications.
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Objective To construct transforming growth factor β receptor Ⅱ (TGFβRⅡ) siRNA expression vector, and investigate the inhibitory effect on TGFβRⅡ in hepatic stellate cells (HSC-T6) ,thus offer a preliminary study for RNAi therapy in liver fibrosis. Methods The two sites of RNAi action were selected in TGFβRⅡ cDNA through online primer design software of Ambion company. The corresponding double-stranded DNA sequence was constructed into pSilencer-U6 plasmid, which could transcribe small interference RNA, the plasmid was transfected into HSC-T6 cells with Lipofectamine 2000. The expression of TGFβRⅡ mRNA and protein were detected by reverse transcription polymerase chain reaction (RT-PCR) and western blotting. Results Expression siRNA plasmids of siRNA-a and siRNA-b that target TGFβRⅡ were successfully constructed. Compared with control group, the expression of TGFβRⅡ mRNA was reduced by 0. 89 ± 0. 06 and 0. 25 ± 0. 03 in two siRNA groups, and expression of TGFβRⅡ protein reduced to 0. 86± 0. 05 and 0. 23 ± 0. 02, respectively. TGFβRⅡ expression was inhibited at mRNA and protein levels after transfected with the siRNA-b plasmid( P <0. 01 ).. There was no difference in the TGFβRⅡ expression after transfected with the siRNA-a plasmid( P > 0. 05). Conclusion TGFβRⅡ-siRNA can effective inhibit expression of TGFβRⅡ in HSC-T6.
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Objectives The purpose of this study is to observe the effects of valsartan on hepapetic fibrosis. Methods Thirty male Sprague-Dawley rats were randomly divided into three groups: valsartan -prevetive group (A), modle group of hepatic fibrosis (B)and valsar-tan-treating group (C). The model of hepatic fibrosis in rats was induced by intraperitoneai injection of dimethylnitrosamine (DMN) for 4 weeks(2ml/kg everyday, three times a week). Valsartan (10mg/kg everyday) was given together with injection of DMN per intrngastric (Ig) in group A for 8 weeks. After stop injection of DMN, the S valsartan(10mg/kg, everyday)was given per Ig in group C for 4 weeks. After modeling, normal saline were given per Ig everyday in group B. At the end of eighth week, the histomorphylogic structure of the liver was ob-served with light microscope. Immunohistoebemical staining was used to evaluate the expression of a-SMA. Results In group B, there was a large necrotic area and a number of pesudolobes appeared in the liver tissue. In group A, there were normal hepatic cords. In the group C, there was fibrosis interval formation and portal area expansion and fibrotie intervals extending to the lobule. The quantitative analysis of Mas-son staining showed that the collagen quantities in group B was higher than that of other group(P<0.01). The collagen quantities in group A was lower than that of group C(P<0.05). The results of immanohistochemical staining showed that the expression of a-SMA in group B was strong positive, middle positive in group C, and weak positive in group A (P<0.05). Conclusion The valsartan has preventive and treatment effects on hepatic fibrosis in rats of hepatic fibrosis model induced by DMN, and the preventive effect of valsartan is better than its treatment effect. The valsartan can ameliorate the hver cirrhosis by partly suppressing the activation of HSC.