摘要
目的探讨Smad7基因对大鼠肾小球系膜细胞(MsC)Ⅰ、Ⅲ型胶原(ColⅠ、ColⅢ)表达的影响,为试图运用Smad7对肾纤维化进行基因治疗提供实验依据。方法经脂质体介导将含有Smad7重组表达质粒转染大鼠MsC,用G418筛选及Northern blot、Western blot法鉴定;又分别采用RT-PCR和Western blot法,检测转染阳性MsC克隆ColⅠ、ColⅢ表达改变。结果成功建立稳定高表达Smad7的阳性MsC克隆(S-22与S-26),并证实两阳性MsC克隆ColⅠ及ColⅢmRNA及蛋白的表达均被明显抑制,其中S-22克隆ColⅠ及ColⅢmRNA表达分别降低47%和56%,其蛋白表达分别降低65%和54%。结论Smad7可能通过抑制组织内ColⅠ及ColⅢ的生成而起到减轻肾纤维化进展的作用。
Objective To investigate the changes of type Ⅰ ,Ⅲ collagen（Col Ⅰ , Col Ⅲ ） expression on cultured rat glomerular mesangial cells （MsC） transfected with Smad 7 cDNA in order to provide experimental proofs for blocking renal fibrosis with Smad 7 gene therapy. Methods Lipofectin method was used to transfect Smad 7 cDNA into MsC, and transfected cells were selected with G418. Smad 7 rnRNA and protein expression was detected by Northern blot and Western blot analysis, respectively. The expression of type Ⅰ ,Ⅲ collagen on MsC were determined by reverse transcription-polymerase chain reaction （RT-PCR） and Western blot analysis. Results Overexpression of Smad 7 on two MsC clones （S-22, S-26） was successfully established. Two MsC clones showed decreased expression of Col Ⅰ , Col Ⅲ mRNA and their proteins. The level of ColⅠ , Col Ⅲ mRNA expression on S-22 clone decreased 47% and 56% respectively. However, the level of their protein expression decreased by 65 % and 54%, respectively. Conclusions It is suggested that Smad 7 could alleviate the progression of renal fibrosis by down-regulating the expression of Col Ⅰ and Col Ⅲ at rnRNA and protein level.
出处
《复旦学报（医学版）》
CAS
CSCD
北大核心
2008年第3期436-440,共5页
Fudan University Journal of Medical Sciences
基金
上海市科委发展基金重点资助项目(01JC14018)
