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目的探讨经门静脉注射抗结缔组织生长因子(CTGF)小分子干扰RNA(siRNA)是否能在体内抑制大鼠肝CTGF基因表达及防治大鼠肝纤维化。方法雄性SD大鼠24只,均分成4组,模型组皮下注射40%CCl4(3ml/kg)及门静脉注射生理盐水,每3d1次,连续6周;预防组皮下注射CCl4及门静脉注射抗CTGFsiRNA(0.1mg/kg),每3d1次,连续6周;治疗组皮下注射CCl42周,随后再给予抗CTGFsiRNA及CCl44周;对照组门静脉注射生理盐水6周。于最后1次CCl4注射后3d行门静脉测压,并取血及组织标本,检测血清转氨酶、透明质酸(HA)及Ⅲ型前胶原(PⅢNP)浓度,应用RTPCR及Western印迹法检测CTGFmRNA及蛋白质在大鼠肝组织表达,应用HE及Siriusred胶原染色检测肝组织炎症及纤维化,应用偏光扫描对肝组织胶原染色面积进行定量。结果与模型组相比,预防组及治疗组大鼠肝组织CTGFmRNA及蛋白质表达显著下调,肝组织炎症、坏死及纤维化明显减轻,血清转氨酶、HA及PⅢNP浓度显著降低;预防组、模型组和治疗组的肝组织胶原染色面积分别为5.8%±0.8%、12.3%±0.8%和7.2%±0.9%(P<0.01);门静脉压力分别为(14.2±2.3)cmH2O、(20.6±5.8)cmH2O和(15.1±3.6)cmH2O(P<0.05),明显降低。结论经门静脉注射抗CTGFsiRNA能在体内显著抑制大鼠肝CTGF基因表达并能有效防治大鼠肝纤维化,提示抗CTGFsiRNA有潜力成为防治肝纤维化的一种新策略。
Objective To investigate whether transfection of anti-connective tissue growth factor (CTGF) small interfering RNA (siRNA) via portal vein can inhibit hepatic CTGF gene expression in rat and prevent and cure hepatic fibrosis in rats. Methods Twenty-four male Sprague-Dawley rats were randomly divided into four groups. The model group was subcutaneously injected with 40% CCl4 (3ml / kg) and the portal vein with physiological saline for 6 weeks. The preventive group received CCl4 subcutaneously and portal vein injection of anti-CTGFsiRNA 0.1mg / kg) every 3 days for 6 weeks. In the treatment group, CCl42 was given subcutaneously for 42 weeks, followed by anti-CTGF siRNA and CCl44 for 24 weeks. The control group was given saline for 6 weeks. The portal vein manometry was performed 3d after the last injection of CCl4, and the concentrations of serum transaminase, HA and PⅢNP were measured. The levels of CTGF mRNA and protein were detected by RTPCR and Western blotting The expression of hepatic tissue was detected by HE and Siriusred collagen staining in liver tissue. Fibrosis was quantified by polarized light scanning. Results Compared with the model group, the expression of CTGF mRNA and protein in the liver tissue of the prevention group and the treatment group were significantly decreased, the inflammation, necrosis and fibrosis of the liver tissue were significantly reduced, and the levels of serum aminotransferase, HA and PⅢNP were significantly decreased. Collagen staining area of the treated group was 5.8% ± 0.8%, 12.3% ± 0.8% and 7.2% ± 0.9% (P <0.01), and portal pressure was (14.2 ± 2.3) cmH2O and (20.6 ± 5.8) And (15.1 ± 3.6) cmH2O (P <0.05), significantly decreased. Conclusion Intravenous injection of anti-CTGFsiRNA can significantly inhibit hepatic CTGF gene expression in rat and effectively prevent and treat hepatic fibrosis in rats, indicating that anti-CTGFsiRNA has the potential to become a new strategy for prevention and treatment of hepatic fibrosis.