shRNA重组慢病毒沉默SMAD3对大鼠肝纤维化的抑制作用  

The effect of lentivirus expressing shRNA targeted to rat SMAD3 on liver fibrosis in rats

作　　者：张莹[1] 孔贺利[2] 张建军[3] 刘梅[2] 任锋[2] 刘霜[2] 陈煜[2] 段钟平[2] 郑素军[2] 

机构地区：[1]解放军第302医院重症监护中心,北京市100039 [2]首都医科大学附属北京佑安医院 [3]四川省广元市第一人民医院

出　　处：《实用肝脏病杂志》2014年第3期267-270,共4页Journal of Practical Hepatology

基　　金："艾滋病和病毒性肝炎等重大传染病防治"科技重大专项(No.2013ZX10002002-006-001);国家自然科学基金(No.30800979;81270532);北京市自然科学基金(No.7102085);北京市科技新星基金(No.2007B055);四川省卫生厅科研项目(No.080137);北京市卫生系统高层次卫生技术人才(No.2011-3-083)

摘　　要：目的观察SMAD3 shRNA重组慢病毒对大鼠肝纤维化的影响。方法随机将Wistar大鼠60只分为生理盐水对照组(20只)、shRNA对照组(20只)和SMAD3 shRNA组(20只)。对shRNA对照组和SMAD3 shRNA组大鼠,通过脾脏注射法给予慢病毒1.0×108TU/只,生理盐水对照组则给予等体积500μl生理盐水,给药1w后开始制备大鼠四氯化碳肝纤维化模型。在造模4w和8w时,采用Real-Time PCR法检测肝组织SMAD3表达;采用ELISA法检测血清I型和III型胶原水平。结果在造模4w和8w时,与生理盐水对照组和shRNA对照组比,SMAD3 shRNA组肝组织SMAD3 mRNA水平显著降低(4w:P=0.000,P=0.001;8w:P=0.001,P=0.009);肝组织学检查显示,在造模4w和8w时,SMAD3 shRNA组大鼠肝纤维化程度减轻;在造模4w时,SMAD3 shRNA组动物血清I型胶原[(3.33±1.60)ng/ml]和III型胶原[(1.32±0.56)ng/ml]明显低于生理盐水对照组[(69.4±13.67)ng/ml,(3.90±1.41)ng/ml],也低于shRNA对照组[(66.8±3.50)ng/ml和(3.80±0.93)ng/ml,均P<0.01];在8w时,各组间胶原水平比较无明显差异(P>0.05)。结论 SMAD3 shRNA在大鼠体内能明显减轻肝纤维化程度。Objective To investigate the effects of lentivirus expressing shRNA targeted to rat SMAD3 on liver fibrosis in rats. Methods A total of 60 rats were randomly divided into three groups,i.e. SMAD3 shRNA group （n=20）,shRNA control group （n=20） and normal saline group （n=20）. The lentivirus were intrasplenicly injected at the dose of 1.0＆#215;108 TU per rat in experimental group,and in normal saline groups,the rats were giv-en the same volume of normal saline. The liver fibrosis in rat were established by intraperitoneal injection of car-bon tetrachloride. The rats were sacrificed at week 4 and week 8 after CCL4 injection,respectively. The SMAD3 mRNA levels in liver tissues was detected by using real-time PCR. The serum collagen I and collagen III were detected by ELISA. Results SMAD3 shRNA decreased the SMAD3 mRNA levels obviously at week 4 and 8 af-ter CCL4 injection（at week 4：P=0.000,P=0.001;at week 8：P=0.001,P=0.009）,respectively,as compared with normal saline group and shRNA control group;the liver fibrosis in SMAD3 shRNA-treated rats improved at week 4 and 8;serum collagen I levels in SMAD3 shRNA-treated group decreased at week 4 （3.33±1.60）ng/ml in SMAD3 shRNA,（69.4±13.67） ng/ml in normal saline and （66.8±3.50）ng/ml in shRNA control group （all P〈0.01）;serum collagen III in SMAD3 shRNA group decreased too at week 4,However,there were no significant differences among the 3 groups at week 8（P〉0.05）. Conclusions Blocking SMAD3 expression can significantly reduced the degree of liver fibrosis in rats.

关 键 词：肝纤维化 SMAD3 SHRNA 慢病毒 大鼠 

分 类 号：R575.2[医药卫生—消化系统]