抗纤灵对5/6肾切除诱导的慢性肾纤维化小鼠Akt-mTOR信号通路的影响

Effects of “Kangxianlin ” on Akt/mTOR signal pathway in 5/6 nephrectomized renal fibrosis in mice

目的观察抗纤灵对5/6肾切除诱导的慢性肾脏纤维化小鼠磷酸化蛋白激酶B(p-Akt)、磷酸化哺乳动物雷帕霉素靶蛋白(p-m TOR)及Akt,m TOR mRNA表达的影响,从Akt-m TOR信号通路角度探讨抗纤灵的作用机制。方法 C57BL/6J雄性小鼠采用5/6肾切除制备慢性肾脏纤维化模型。2周后根据肌酐水平将手术组小鼠分为模型组、雷帕霉素组和抗纤灵组,同时设立假手术组,每组10只。雷帕霉素组、抗纤灵组分别给予雷帕霉素(0.8μg/g)、抗纤灵(20 g/kg)每天0.5 ml灌胃治疗,模型组和假手术组灌胃等量蒸馏水。连续用药8周后处死小鼠,检测各组小鼠肌酐(Scr)、尿素氮(BUN)、24小时尿蛋白(24 h UP)含量;采用蛋白质印迹法(Western blotting)、RT-PCR检测肾脏组织中p-Akt、p-m TOR蛋白及Akt、m TOR mRNA的表达。结果与假手术组比较,模型组Scr、BUN、24 h UP、p-Akt、p-m TOR蛋白及Akt、m TOR mRNA显著升高(P<0.05,P<0.01)。与模型组比较,雷帕霉素组、抗纤灵组Scr、UN、24 h UP、p-Akt、p-m TOR蛋白及Akt,m TOR mRNA均有所降低(P<0.05,P<0.01)。雷帕霉素组与抗纤灵组比较,在降低Scr、24 h UP方面,抗纤灵组明显优于雷帕霉素组(P<0.05);在降低p-m TOR蛋白及m TOR mRNA方面,雷帕霉素优于抗纤灵组(P<0.05)。结论抗纤灵可能通过调节Akt-m TOR信号通路,对肾纤维化有一定的改善作用。

Objective To determine effects of ＂Kangxianling Decoction＂ （KXLD） on the Akt-mTOR signaling pathway in 5/6 nephrectomized mice renal fibrosis model and discuss mechanism of it. Methods SPF C57BL/6J male mice were established models of chronic renal failure caused by 5/6 nephrectomized, and two week later after surgery, according to serum creatinine （Scr） level,the mice were divided into model group, KXLD and rapamycin （RAP） group, sham operation group was set at the same time, 10 in each group. KXLD and RAP group were gavaged respectively according to 20 g/kg,RAP 0.8 μg/g,0.5 ml/day. Sham and model groups were administered with the same amount of distilled water. The mice were sacrificed after 8 weeks of treatment, Ser, BUN and 24 h UP were determined, the levels of Akt-mTOR protein and mRNA expression were observed by Western blotting and real-time PCR method. Results Compared with sham group, the Scr, urea nitrogen,24 h UP and p-Akt,p-mTOR levels increased significantly in the model group （P 〈 0.05, P 〈 0.01 ）. Compared with model group, the Scr, urea nitrogen ,24 h UP and p-Akt,p-mTOR protein and Akt, mTORmRNA levels decreased significantly in the KXLD and RAP group （P 〈 0. 01, P 〈 0.05 ）. Compared between KXLD and RAP groups, in Scr, 24 h UP, KXLD superior to RAP, in roTOR protein and mRNA, RAP superior to KXLD （P 〈0.05 ）. Conclusion ＂Kangxianling Decoction＂ is able to protect renal function via Akt-mTOR signaling pathway.