益气化瘀清热方及其拆方对IgA肾病大鼠肾组织mTOR信号通路的影响

Effects of Yiqi Huayu Qingre Formula and its compositions on mTOR signaling pathway in nephrogenic tissue of rats with IgA nephropathy

目的:探讨益气化瘀清热方及其拆方干预IgA肾病的作用机制及IgA肾病的发生发展机制。方法:将90只雄性Wistar大鼠随机分为6组,即空白组、模型组、益气组、化瘀组、清热组、复方组,采用牛血清白蛋白+四氯化碳+脂多糖的方法制备IgA肾病模型;运用免疫组化技术检测mTOR信号通路下游信号蛋白磷酸化核糖体蛋白S6(pS6)、磷酸化真核细胞启动因子4E结合蛋白1(p-4EBP1)的表达部位,并对其进行半定量分析;运用Western blot检测各组大鼠肾皮质pS6、p-4EBP1的蛋白表达量。结果:pS6、p-4EBP1主要在IgA肾病模型大鼠肾组织中肾小球足细胞及壁层上皮细胞中表达,肾小管中散在表达。免疫组化结果显示,pS6表达量模型组最高(P﹤0.05),益气组次之(P﹤0.05)。各实验组p-4EBP1蛋白表达量模型组最高(P﹤0.05),复方组、空白组最低(P﹤0.05)。Western blot结果同免疫组化结果一致。结论:益气化瘀清热方及其拆方可通过抑制mTOR下游信号蛋白pS6、p-4EBP1的表达发挥治疗作用。

Objective： To explore the mechanism of Yiqi Huayu Qingre Formula and its compositions in IgA nephropathy （IgAN） and the pathogenesis of IgAN. Methods： Ninety male Wistar rats were randomly divided into 6 groups： blank group, model group, Yiqi group, Huayu group, Qingre group, combination group. The IgAN model was prepared by BSA＋CCI4＋LPS method. The expression of pS6 and p-4EBP1 in the downstream of mTOR signaling pathway was detected by immunohistochemistry, then was analyzed by Semi-quantitative; the expression levels of pS6 and p-4EBP1 in renal cortex of each group were detected by Western blot. Results： pS6 and p-4EBP1 were mainly expressed in glomerular podocytes and parietal epithelial cells in the renal tissues of IgAN model rats, and scattered in the renal tubules. The results of immunohistochemistry showed that the pS6 expression model group was the highest （P〈0.05）, the Yiqi group was the second （P〈0.05）. The model group of p-4EBP1 protein expression was the highest in each experimental group （P〈0.05）, and the lowest in combination group and blank group （P〈0.05）. Western blot results were consistent with immunohistochemistry results. Conclusion： Yiqi Huayu Qingre Formula and its decomposed formula can play a therapeutic role by inhibiting the expression of mTOR downstream signaling proteins pS6 and p-4EBP1.