沉默结缔组织生长因子对大鼠肝纤维化的影响

Effect of scilencing connective tissue growth factor on the liver fibrosis in rats

目的 研究结缔组织生长因子（CTGF）小干扰RNA（siRNA）在CCl4诱导的大鼠肝纤维化模型中的抗肝纤维化作用及其对肝星状细胞（HSC）激活的影响.方法 雄性SD大鼠24只,随机分成4组.模型组：皮下注射CCl4及经门静脉注射等渗盐水,每3 d 1次,连续6周（给药频率与时间,下同）;CTGF siRNA干预组：皮下注射CCl4及经门静脉注射CTGF siRNA ;对照siRNA干预组：皮下注射CCl4及经门静脉注射对照siRNA;正常对照组：经门静脉注射等渗盐水.用HE染色和Sirius red染色评估大鼠肝组织学变化,Western blot检测肝组织CTGF及α平滑肌肌动蛋白（α-SMA）的表达,免疫组织化学染色评估肝组织α-SMA阳性细胞数量.对实验数据用单因素方差分析或Kruskal-Wallis检验分析.结果 模型组及对照siRNA组大鼠肝组织CTGF及α-SMA蛋白表达显著上调,α-SMA染色阳性细胞数量明显增加.与模型组相比,CTGF siRNA组CTGF及α-SMA蛋白表达分别下调95%±2%和86%±11%（F值分别为21.234和12.473,P值均＜0.01）;肝组织α-SMA染色阳性细胞数减少76%±9%（F=9.179,P＜0.01）;组织学检查显示肝纤维化程度明显减轻.结论 siRNA沉默CTGF基因能显著减轻大鼠实验性肝纤维化,减少活化HSC数量可能是其改善肝纤维化的主要机制之一.

Objective To investigate the anti-fibrogenesis property of intraportal vein small interfering RNA（siRNA） injection targeting connective tissue growth factor （CTGF） in a rat model of liver fibrosis induced by carbon tetrachloride （CCl4） and its effect on hepatic stellate cell （HSC） activation. Methods 24 male rats were randomly divided into four group, rats received CC14 by subcutaneous injections every three days for 6 consecutive weeks, and meantime they also obtained either siRNA targeting CTGF （as CTGF siRNA group）, saline （as model group） or a control siRNA （as control siRNA group） by intraportal vein injection to rats liver at the same approach. Other rats received saline intraportal vein injection for 6 weeks （as normal control group）. The expressions of CTGF and α -SMA protein were detected by Western blot. Hepatic histology was evaluated by HE staining and Sirius red staining. The collagen staining areas were measured quantitatively using a computer-aided manipulator with slight modifications. The number of active HSC were evaluated by immunohistochemisty. Results Six weeks after CCl4 injection, prominent upregulations were observed in the expressions of CTGF and α -SMA protein in saline or control siRNAtreated rats livers. In rats with CTGF siRNA treatment, the protein expressions of CTGF and α -SMA in liver decreased by 95% ± 2% and 86% ± 11% （F = 21.234 and 12.473, P 〈 0.01） respectively, the number of active HSC in liver decreased by 76% ± 9% （F = 9.179, P 〈 0.01） as compared to the model group. The