降脂通络软胶囊调控SIRT1/FoxO3通路对膜性肾病大鼠细胞凋亡的影响

Jiangzhi Tongluo Soft Capsules Regulate SIRT1/FoxO3 Pathway to Affect Cell Apoptosis in Rats with Membranous Nephropathy

目的:通过观察降脂通络软胶囊对膜性肾病(MN)大鼠肾组织中沉默信息调节因子2相关酶1(SIRT1)/叉头转录因子O3(FoxO3)蛋白表达及足细胞凋亡的影响,探讨其治疗MN的可能分子机制。方法:将60只雄性SD大鼠随机分为6组,每组10只,随机选取1组作为正常组,其余5组分别为模型组、盐酸贝那普利(西药)组(10 mg·kg^(-1))和降脂通络软胶囊低、中、高剂量(中药低、中、高剂量)组(25、50、100 mg·kg^(-1)),并通过尾静脉注射阳离子化牛血清白蛋白(C-BSA)的方法造模。造模成功后,各组按照相应比例剂量连续4周灌胃给药,并在第4周末留取肾组织,采用电镜、免疫荧光观察肾脏病理学变化,蛋白免疫印迹法(Western blot)检测大鼠肾脏SIRT1、FoxO3蛋白的表达情况,免疫组化(IHC)检测B细胞淋巴瘤-2(Bcl-2)、Bcl-2相关X蛋白(Bax)、死亡启动子重组蛋白(Bad)、足细胞裂孔膜蛋白(Nephrin)、膜蛋白(Podocin)的蛋白表达情况。结果:与正常组比较,模型组大鼠肾组织中促凋亡因子Bax、Bad、FoxO3蛋白表达均明显增加(P<0.05),抗凋亡因子Bcl-2、SIRT1、Nephrin、Podocin蛋白表达均降低(P<0.05);与模型组比较,各治疗组肾组织中Bax、Bad、FoxO3蛋白表达明显减少(P<0.05),Bcl-2、SIRT1、Nephrin、Podocin表达明显升高(P<0.05)。结论:降脂通络软胶囊对MN的肾保护作用可能与其调控SIRT1/FoxO3通路,减少足细胞凋亡,维护足细胞结构稳定有关。

Objective:To observe the effect of Jiangzhi Tongluo soft capsule on the protein levels of silent mating-type information regulation 2 homolog 1(SIRT1)and forkhead transcription factor FoxO3 and podocyte apoptosis in the renal tissue of rats with membranous nephropathy and to reveal the underlying molecular mechanisms for the treatment of MN.Method:Sixty male SD rats were randomly assigned into 6 groups with 10 rats each.The six groups included a normal group,a model group,benazepril hydrochloride group,and Jiangzhi Tongluo soft capsule groups of low,medium and high doses(25,50,100 mg·kg^(-1),respectively).The model rats were established by injection with cationized bovine serum albumin into the tail vein.After modeling,the rats were administrated with corresponding agents by gavage for 4 weeks.At the end of the 4th week,an electron microscope was used to observe the pathological changes in the kidney.Western blot was employed to detect the protein levels of SIRT1 and FoxO3 protein in rat kidney,and immunohistochemistry to detect the expression of B lymphocytoma-2(Bcl-2),Bcl-2-associated X protein(Bax),Bcl-2-associated death promoter(Bad),and podocyte split diaphragm proteins nephrin and podocin.Result:Compared with normal group,the expression of pro-apoptotic factors Bax,Bad,and FoxO3 in the kidney was up-regulated(P<0.05),while that of anti-apoptotic factors Bcl-2,SIRT1,nephrin,and podocin was down-regulated(P<0.05)after modeling.Compared with the model group,the treatments down-regulated the expression of Bax,Bad,and FoxO3(P<0.05)and up-regulated that of Bcl-2,SIRT1,nephrin,and podocin(P<0.05).Conclusion:Jiangzhi Tongluo soft capsule may regulate the SIRT1/FoxO3 pathway to reduce podocyte apoptosis and maintain podocyte structure stability,thereby exerting the renal protection effect.