沉默结缔组织生长因子对大鼠肝纤维化及细胞外基质积聚的影响  

Effect of silencing connective tissue growth factor on rat liver fibrosis and the accumulation of extraceiinlar matrix

作　　者：李光明[1] 李定国[1] 谢青[2] 史毅[3] 姜山[2] 周慧娟[2] 陆汉明[1] 金由辛[3] 

机构地区：[1]上海交通大学医学院附属新华医院消化科,200092 [2]上海交通大学医学院附属瑞金医院感染科 [3]中国科学院上海生命研究院、生物化学与细胞生物学研究所、分子生物学国家重点实验室

出　　处：《中华肝脏病杂志》2008年第3期188-192,共5页Chinese Journal of Hepatology

摘　　要：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细胞外基质积聚的影响。方法雄性SD大鼠30只，随机分成5组。模型组：皮下注射CCl4及经门静脉注射等渗盐水，3d1次，连续6周；预防组：皮下注射CCh及经门静脉注射CTGFsiRNA，连续6周；治疗组：皮下注射CCl4 2周，随后给予CTGFsiRNA及CCl4 4周；对照siRNA干预组：皮下注射CCla及经门静脉注射对照siRNA，连续6周；空白对照组：经门静脉注射等渗盐水6周。用HE和SiNusred染色评估大鼠肝组织学变化，RTPCR或（和）Westernblot检测肝组织CTGF、Ⅰ与Ⅲ型胶原及层黏连蛋白的表达，放射免疫法检测外周血Ⅲ型前胶原及透明质酸含量。结果模型组及对照siRNA组大鼠肝组织CTGF、Ⅰ与Ⅲ型胶原及层黏连蛋白基因表达显著上调；与模型组相比，预防组及治疗组由CCl4诱导的CTGFmRNA和蛋白表达及Ⅰ与Ⅲ型胶原、层黏连蛋白mRNA表达分别下调76%±8％、95％±2％、74%±8％、78％±8％、31％±7％和80％±3％、93％±3％、57％±6%、59％±10％、43％±9％（F值分别为68．630，21．234，24．219，16．315和9．716，P值均〈0．01），肝纤维化程度明显减轻，大鼠外周血Ⅲ型前胶原和透明质酸含量也显著降低。结论经门静脉注射CTGF siRNA能显著抑制实验大鼠肝CTGF基因表达，由此通过阻止细胞外基质积聚而缓解肝纤维化。Objective To investigate the anti-fibrogenesis property of intraportal vein injection of small interfering RNA targeting connective tissue growth factor （CTGF） in a rat model of liver fibrosis and its effect on the accumulation of extracellular matrix （ECM）. Methods Thirty male rats were randomly divided into five groups. Some rats received CCL subcutaneously every three days for 6 consecutive weeks, and in the meantime they also received either siRNA targeting CTGF （preventive group）, saline （model group） or siRNA （siRNA control group） by intraportal vein injections. Other rats received CCL by subcutaneous injection for 2 weeks, followed by CCL and CTGF siRNA intraportal vein injection for 4 more weeks （as treatment group）. The expressions of CTGF and type Ⅰ and Ⅲ collagen genes were detected by means of reverse transcriptionpolymerase chain reaction （RT-PCR） and/or Western blot respectively. Hepatic histology was evaluated by HE and Sirius red stained sections. The collagen staining areas were measured quantitatively using a computer-aided manipulator with slight modifications. Serum procollagen type Ⅲ and hyaluronic acid were determined by radioimmunoassay. Results Six weeks after CC14 injection, prominent upregulations of gene expressions of CTGF, type I and In collagen, and laminin in saline or siRNA-treated rat livers were observed. The expressions of CTGF at mRNA and protein level and type I and In collagen at mRNA level were mark- edly reduced in rats with CTGF siRNA treated for four or six weeks. Expressions of CTGF at mRNA and protein levels decreased by 76%±8%, 80% ± 3% （F = 68.630, P 〈 0.01） and 95% ± 2%, 93% ± 3% （F = 21.234, P 〈 0.01）; type I and In collagen and laminin at mRNA levels decreased by 74%± 8%, 78% ± 8%, 31% ± 7% and 57% ± 6%, 59% ±10%, 43% ± 9% （F = 24.219, 16.315, 9.716, P 〈 0.01） compared with rats in the model group at 72 h. The CTGF siRNA treatment markedly reduced serum levels of procollagen type In and hyaluronic acid a

关 键 词：肝纤维化 结缔组织生长因子 小干扰RNA 细胞外基质 

分 类 号：R285.5[医药卫生—中药学]