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浙江大学学报（医学版）
浙江大学学报（医学版）  2011, Vol. 40 Issue (6): 603-608    DOI: 10.3785/j.issn.1008-9292.2011.06.006
专题报道     
结缔组织生长因子特异性小分子干扰RNA的筛选及其在抗肝纤维化中的作用
毛小荣，岳伟，袁宏，陈红，薛苗
兰州大学第一医院传染科，甘肃 兰州 730000
Inhibition effect of small interfering RNA targeting connective tissue growth factor on liver fibrosis in rats
MAO Xiao-rong,YUE Wei,YUAN Hong,CHEN Hong，XUE Miao
Department of Infectious Diseases,The First Affiliated Hospital of Lanzhou University,Lanzhou 730000, China
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摘要： 目的：设计和合成针对抗肝纤维化关键基因结缔组织生长因子（connective tissue growth factor，CTGF）的特异性小干扰RNA（siRNA），并筛选高效的CTGFsiRNA抗肝纤维化序列，探讨其通过尾静脉注射是否具有抗大鼠肝纤维化作用。
方法：①筛选高效的CTGF siRNA序列。②干预肝纤维化模型大鼠。选取雄性SD大鼠30只，随机分成5组，分别为空白对照组尾静脉注射生理盐水8周；模型组腹腔注射40%CCl4(3 ml/kg)及尾静脉注射生理盐水，1次/3d，连续8周；预防组腹腔注射40%CCl4 及尾静脉注射CTGF siRNA（0.1 mg/kg），1次/3 d，连续8周；2周治疗组腹腔注射40%CCl4 2周，后给予CTGF siRNA及CCl4 6周；4周治疗组腹腔注射40%CCl4 4周，后给予CTGF siRNA及CCl4 4周。于最后一次CCl4注射3天后取血及组织标本，检测肝纤维化指标，应用Real-Time PCR及Western印迹法检测CTGF mRNA及蛋白质在大鼠肝组织表达，应用Masson染色检测肝组织纤维化。
结果：与模型组(0.544±0.019)相比，预防组（0.105±0.003）及治疗组(0.190±0.006)大鼠肝组织CTGF mRNA和蛋白质表达显著下调（P<0.05），肝组织炎症和坏死及纤维化明显减轻，肝纤维化指标显著降低（P<0.05）。4周治疗组与模型组相比各指标降低，与预防组及2周治疗组相比各指标相对升高（P<0.05）。
结论：成功筛选出高效的CTGF siRNA，且经尾静脉注射CTGF siRNA能显著抑制大鼠体内肝脏CTGF基因表达，并能有效防治大鼠肝纤维化，对于病程较长的肝纤维化也有一定的治疗作用，提示CTGF siRNA在抗肝纤维化治疗中有重要作用。
关键词： RNA;  小分子干扰/药理学;  胞间信号肽类和蛋白质类/生物合成;  肝硬化/病理学;  疾病模型;  动物;  结缔组织生长因子;  小分子干扰RNA;  肝纤维化    
Abstract: Objective： To design and synthesize small interfering RNA (siRNA) targeting connective tissue growth factor (CTGF) and to investigate its effect on liver fibrosis.
Methods： The interference sequence of CTGF was designed and synthesized.Rat hepatic fibrosis model was induced by intraperitoneal injection of 40 % CCl4(3 ml/kg).Thirty male rats were randomly divided into 5 groups:in normal control and model groups rats received tail vein injection of normal saline every 3 days for 8 consecutive weeks; in preventive group rats received tail vein injection of CTGF siRNA (0.1 mg/kg) every 3 days for 8 weeks；in 2-w treatment group CTGF siRNA was given for 6 weeks starting from two weeks after CCl4 injection; in 4-w treatment group CTGF siRNA was given for 4 weeks starting 4 weeks after CCl4 injection.The serum and hepatic tissue samples were harvested 3 days after the last CCl4 injection.Hepatic fibrosis indices were measured.Expression of CTGF mRNA and protein in the liver was evaluated by RT-PCR and Western blot,respectively.Fibrosis in rat liver was analyzed by Masson staining.
Results： Compared with model group (0.544 0.019),the expression of CTGF mRNA and protein in liver of both preventive（0.105±0.003）and 2-w treatment groups (0.190±0.006) were markedly down-regulated（P<0.05）.Inflammation,necrosis and fibrosis in hepatic tissue were significantly attenuated.In addition,the serum ration of liver fibrosis indices was greatly reduced（P<0.05）.Compared with preventive and 2-w treatment groups,the expression of CTGF mRNA and protein in liver in4 weeks of treatment group were up-regulated（P<0.05）; inflammation,necrosis and fibrosis in hepatic tissue were relative increased; and the serum concentrations of liver fibrosis indices were relatively higher（P<0.05）.
Conclusion： The highly effective CTGF siRNA has been successfully synthesized,which can inhibit CTGF expression in liver,prevent hepatic fibrosis and its progress in rats.
Key words: RNA,small interfering/pharmacol    Intercellular signaling peptides and proteins/biosyn    Liver cirrhosis/pathol    Disease models,animal    Connective tissue growth factor    siRNA    Liver fibrosis
出版日期: 2011-11-25
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毛小荣;岳伟;袁宏;陈红;薛苗. 结缔组织生长因子特异性小分子干扰RNA的筛选及其在抗肝纤维化中的作用[J]. 浙江大学学报（医学版）, 2011, 40(6): 603-608.

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https://www.zjujournals.com/med/CN/Y2011/V40/I6/603

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