加味三仁汤对免疫球蛋白A肾病大鼠PI3K/AKT/NF-κB信号通路的影响

Effect of Jiawei Sanren Decoction on PI3K/Akt/NF-κB Signaling Pathway in Immunoglobulin A Nephropathy Rats

目的:研究加味三仁汤对免疫球蛋白A肾病(immunoglobulinA nephropathy,IgAN)大鼠肾脏的保护作用及其对PI3K/AKT/NF-κB信号通路的调控作用,探讨加味三仁汤治疗IgA肾病的机制。方法:采用牛血清白蛋白(BSA)400 mg/kg灌胃+四氯化碳(CCl;)0.1 mL皮下注射+脂多糖(LPS)0.05 mg尾静脉注射方法建立实验性IgA肾病(IgAN)大鼠模型。造模成功大鼠随机分为模型对照组、贝那普利0.01 g/kg组、加味三仁汤10.35 g/kg组,另取正常大鼠作为对照。各组给予相应的药物干预,灌胃体积8 mL/kg,1次/d,连续给药4 w后检测大鼠24 h尿蛋白(24-UTP)、血清肌酐(SCr)、尿素氮(BUN)、白蛋白(ALB)含量,同时检测炎性因子白介素-1β(IL-1β)、白介素-18(IL-18)、单核细胞趋化蛋白-1(MCP-1)含量;苏木精-伊红(HE)染色,观察肾脏组织病理形态学变化;免疫组化法、蛋白质印迹法(Western blot)和反转录·聚合酶链反应法(RT-PCR)分别检测PI3K/AKT/NF-κB通路相关蛋白和基因的表达。结果:与正常对照组比较,模型对照组大鼠24 h-UTP显著升高,血清白蛋白含量显著降低,IL-1β、IL-18、MCP-1含量均显著升高(P<0.01)。肾小球基底膜增厚,细胞外基质增多,系膜扩张;PI3K/AKT/NF-κB信号通路相关蛋白及基因表达显著改变(P<0.01)。与模型对照组比较,加味三仁汤10.35 g/kg组24 h-UTP显著降低(P<0.01),血清白蛋白含量明显升高(P<0.05),IL-1β、IL-18水平显著降低(P<0.01),MCP-1含量明显降低(P<0.05);肾脏组织病变明显改善,明显抑制肾组织中PI3K、AKT蛋白磷酸化及下调mRNA,下调NF-κB蛋白及mRNA表达,同时上调PTEN蛋白及mRNA表达。结论:加味三仁汤可能通过调控PI3K/AKT/NF-κB信号通路改善IgA肾病大鼠的肾组织损伤状况,延缓IgA肾病的发生发展。

Objective:To study the protective effect of Jiawei Sanren Decoction on kidney in immunoglobulin A nephropathy(IgAN)rats and its regulatory effect on the PI3 K/Akt/NF-κB signaling pathway,and investigate its mechanism on IgAN.Methods:The IgAN model was induced by intragastric administration of 400 mg/kg bovine serum albumin(BSA),subcutaneous injection of 0.1 mL of carbon tetrachloride(CCL;)and caudal intravenous injection of 0.05 mg of lipopolysaccharide(LPS)in rats.The model rats were randomly divided into a model group,a benazepril group(0.01 g/kg),and a Jiawei Sanren Decoction group(10.35 g/kg).Normal rats were selected and assigned to the control group.Rats were treated correspondingly by gavage at 8 mL/kg,once per day.After 4 weeks of continuous administration,the content of 24-hour urine total protein(24-UTP),serum creatinine(Scr),blood urea nitrogen(BUN),and albumin(ALB)was measured.Inflammatory factors,such as interleukin 1 beta(IL-1β),interleukin 18(IL-18),and monocyte chemotactic protein 1(MCP-1)were detected.Hematoxylin-eosin(HE)staining was used to observe the pathological changes of renal tissues.Immunohistochemistry assay,Western blot and RT-PCR were used to detect protein and gene expression levels in the PI3 K/Akt/NF-κB signaling pathway.Results:Compared with the control group,the model group showed elevated content of 24 h-UTP,IL-1β,IL-18,and MCP-1(P<0.01),decreased content of ALB(P<0.01),thickened glomerular basement membrane,increased extracellular matrix,expanded mesenterium,and changed protein and gene expression in the PI3 K/Akt/NF-κB signaling pathway(P<0.01).Compared with the model group,the Jiawei Sanren Decoction group showed decreased content of 24 h-UTP,IL-1β,IL-18,and MCP-1(P<0.01 or P<0.05),increased content of ALB(P<0.05),improved pathological changes of renal tissues,inhibited phosphorylation of PI3 K and Akt,up regulated protein and mRNA expression level of PTEN,and down-regulated protein and mRNA expression level of NF-κB.Conclusion:Jiawei Sanren Decoction may ameliorate the kidney injury of IgAN rats by inhibiting the PI3 K/Akt/NF-κB signaling pathway and slowing down the development of IgAN.