CAJ | 학술논문

目的研究转化生长因子(TGF) β1的小干扰RNA (siRNA)对CCl4诱导肝纤维化大鼠TGFβ1/Smad信号通路的影响.方法以前期研究的TGFβ1siRNA治疗前后肝纤维化大鼠肝脏组织作为研究对象,分为正常组、模型组、TGF β 1 siRNA 0.125 mg/kg组、TGFβ1 siRNA 0.25 mg/kg组和阴性对照组(0.25 mg/kg非特异性靶序列).用免疫组织化学和Western blot检测Smad2、3、4、7的蛋白表达;Real-time PCR法检测Smad2、3、4、7的mRNA表达.多组间数据比较用方差分析,两两比较采用q检验. 结果免疫组织化学结果显示TGF-β1 siRNA 0.25mg/kg组及0.125 mg/kg组Smad2/3、4的蛋白表达量较模型组、阴性对照组明显减少(F值分别为115.538,117.178,P＜ 0.01),TGFβ 1 siRNA 0.25 mg/kg组表达量较0.125 mg/kg组少(P＜0.05).TGFβ 1 siRNA 0.25 mg/kg组及0.125 mg/kg组Smad7的蛋白表达量较模型组、阴性对照组增加(F=125.34,P＜ 0.01),TGFβ 1 siRNA 0.25mg/kg组表达量较0.125 mg/kg组多(P＜0.05).Western blot结果与之相似.Real-time PCR结果显示,与模型组和阴性对照组比较,TGFβ1 siRNA可以明显抑制Smad 2、3、4基因的表达,且TGFβ1 siRNA 0.25 mg/kg组Smad2、3、4的mRNA表达量(相对表达量分别为2.870±0.705、2.904±0.414、2.667±0.466)较TGF β1siRNA 0.125 mg/kg组(相对表达量分别为5.998±0.329、5.827±0.781、5.102±0.102)明显减少(P＜0.05).TGFβ1 siRNA 0.25mg/kg组及0.125 mg/kg组Smad7的mRNA表达量较模型组、阴性对照组增加(F=29.615,P＜0.01),TGFβ1 siRNA 0.25 mg/kg组表达量较0.125mg/kg组多(P＜0.05).结论 TGFβ1 siRNA干扰大鼠TGFβ1表达后,Smad2、3、4表达量明显降低,而Smad7表达仍维持在较高水平,这利于肝纤维化的改善.
목적 연구전화생장인자(TGF) β1적소간우RNA (siRNA)대CCl4유도간섬유화대서TGFβ1/Smad신호통로적영향.방법 이전기연구적TGFβ1siRNA치료전후간섬유화대서간장조직작위연구대상,분위정상조、모형조、TGF β 1 siRNA 0.125 mg/kg조、TGFβ1 siRNA 0.25 mg/kg조화음성대조조(0.25 mg/kg비특이성파서렬).용면역조직화학화Western blot검측Smad2、3、4、7적단백표체;Real-time PCR법검측Smad2、3、4、7적mRNA표체.다조간수거비교용방차분석,량량비교채용q검험. 결과 면역조직화학결과현시TGF-β1 siRNA 0.25mg/kg조급0.125 mg/kg조Smad2/3、4적단백표체량교모형조、음성대조조명현감소(F치분별위115.538,117.178,P＜ 0.01),TGFβ 1 siRNA 0.25 mg/kg조표체량교0.125 mg/kg조소(P＜0.05).TGFβ 1 siRNA 0.25 mg/kg조급0.125 mg/kg조Smad7적단백표체량교모형조、음성대조조증가(F=125.34,P＜ 0.01),TGFβ 1 siRNA 0.25mg/kg조표체량교0.125 mg/kg조다(P＜0.05).Western blot결과여지상사.Real-time PCR결과현시,여모형조화음성대조조비교,TGFβ1 siRNA가이명현억제Smad 2、3、4기인적표체,차TGFβ1 siRNA 0.25 mg/kg조Smad2、3、4적mRNA표체량(상대표체량분별위2.870±0.705、2.904±0.414、2.667±0.466)교TGF β1siRNA 0.125 mg/kg조(상대표체량분별위5.998±0.329、5.827±0.781、5.102±0.102)명현감소(P＜0.05).TGFβ1 siRNA 0.25mg/kg조급0.125 mg/kg조Smad7적mRNA표체량교모형조、음성대조조증가(F=29.615,P＜0.01),TGFβ1 siRNA 0.25 mg/kg조표체량교0.125mg/kg조다(P＜0.05).결론 TGFβ1 siRNA간우대서TGFβ1표체후,Smad2、3、4표체량명현강저,이Smad7표체잉유지재교고수평,저리우간섬유화적개선.
Objective To investigate the changes in Smad 2,3,4 and 7 of the transforming growth factor-beta 1 (TGF-βl)/Smad signaling pathways in carbon tetrachloride (CCLA)-induced hepatic fibrosis rats treated with TGF-β1 small interferring (si)RNA.Methods Rats were randomly divided among five groups:non-fibrotic (normal); fibrosis-induced (model); fibrotic treated with 0.125 mg/kg TGF-β 1 siRNA;fibrotic treated with 0.250 mg/kg TGF-β1 siRNA; and fibrotic treated with negative control TGF-β1 siRNA,The expression of Smad 2,3,4 and 7 was detected by real-time polymerase chain reaction (for mRNA),immunohistoehemistry and Western blotting (for protein).Results The mRNA and protein levels of Smad 2,3and 4 were significantly lower in the the fibrotie rats treated with either 0.250 mg/kg or 0.125 mg/kg TGF-β1siRNA than in the fibrotic model or the negative control TGF-β1 siRNA rats (P ＜ 0.01).Moreover,the mRNA and protein expression levels of Smad 2,3 and 4 were significantly lower in the 0.250 mg/kg TGF-β1 siRNA group than in the 0.125 mg/kg group (P ＜ 0.05).Comparing the 0.250 mg/kg and 0.125 mg/kg TGF-β1 siRNA groups to the model group and the TGF-β1 siRNA negative control group showed significantly increased levels of mRNA and protein expression of Smad 7 (P ＜ 0.01).In addition,the expression levels of Smad 7were significantly higher in the 0.250 mg/kg TGF-β1 siRNA group than in the 0.125 mg/kg group (P ＜ 0.05).Conclusions siRNA-mediated silencing of TGF-β1 in rats led to significantly reduced expression of Smad 2,3and 4,but significantly increased expression of Smad 7.TGF-β1 regulation of Smad signaling molecules may contribute to hepatic fibrosis in rats and represent a target of future therapeutic intervention.