RNA干扰技术下调大鼠血管紧张素Ⅱ1型受体

RNA Interference Reduced Rat Angiotensin Ⅱ Type 1 Receptors Expression

目的:研究RNA干扰技术对体内外大鼠血管紧张素Ⅱ1型受体(AT1R)基因的沉默作用。方法:人胚肾293细胞扩增Ad5-AT1R-shRNA-EGFP并感染体外培养的C6细胞株,进行RT-PCR和Western-blot检测AT1R mRNA和蛋白的表达。Ad5-AT1R-shRNA-EGFP尾静脉注射法转染SHR,免疫组织化学法测定SHR主要器官:心脏、肝脏、肾脏、肾上腺和主动脉AT1R表达。结果:Ad5-AT1R-shRNA-EGFP显著抑制C6细胞AT1R mRNA及蛋白表达,并显著抑制SHR心脏、肝脏、肾脏、肾上腺、主动脉的AT1R表达。结论:靶向大鼠AT1R基因mRNA的短发夹RNA腺病毒载体,在体外和体内环境下高效特异性抑制靶基因的表达,能达到基因干扰目的。

Objective： Research on gene silence by RNA interference technology, through transfection with constructed Aden- ovirus - based shRNA expression systems which target against the rat angiotensin Ⅱ receptor gene. Methods： Use human embryo kidney 293 cell to culture Ad5- ATIR- shRNA -EGFP. And then recombinant adenoviral vectors were transfected into rat glioma cells. The cultured cells were collected at different phases. RT - PCR and Western blot were performed. SHR were transfected with recombinant adenoviral vectors by caudal vein injection method. ATI R expression of major organ： include heart, liver, kidney, adrenal gland and aorta was detected by immunohistoehemical method. Results： Ads - ATIR - shRNA - EGFP were transfected into C6 in vitro and SHR in vivo successfully. C6 ATIR mRNA and protein expression could be inhibited significantly. Immunohistochemieal method showed that AT IR expression of major organ decreased considerably. Conclusion： Both in vitro and in vivo, Adenovirus - based shRNA expression systems targeting against the rat angiotensin Ⅱ receptor gene enables the effective silence of gene expres- sion to the target mRNA and leads to effective inhibition of translation of proteins.