摘要
目的探讨肾疏宁干预乙型肝炎病毒(HBV)诱导人肾小管上皮细胞(HK-2)转分化,延缓肾间质纤维化作用的可能机制。方法通过75只C57BL/6小鼠的中药肾疏宁灌胃,制备含药血清;使用Lipofectamine转染HBV至HK-2的方法造模;将细胞分为:空白血清对照组、空质粒组、HBV组、贝那普利组、肾疏宁低浓度组、肾疏宁中浓度组、肾疏宁高浓度组。运用酶联免疫吸附实验(ELISA)、实时荧光定量PCR法、蛋白质印迹(Western blot)法检测HK-2细胞中HBsAg、HBeAg、TGF-β1、p-Smad2、p-Smad3、Smad7、Col-Ⅲ、ColⅣ、FN的表达。结果ELISA检测结果:HBV组与空质粒组比较,HBsAg、HBeAg、TGF-β1表达上调(P<0.01);与HBV组比较,贝那普利组、肾疏宁低浓度组、肾疏宁中浓度组、肾疏宁高浓度组TGF-β1蛋白表达下降(P<0.001);与贝那普利组比较,肾疏宁低浓度组TGF-β1的表达相对较高(P<0.05)。实时荧光定量PCR检测结果:与HBV组比较,贝那普利组、肾疏宁各干预组ColⅢ、Col-Ⅳ、FN mRNA表达明显下降(P<0.001)。Western blot检测结果:与HBV组比较,贝那普利组、肾疏宁各干预组p-Smad2、p-Smad3蛋白表达明显下降(P<0.001)。结论肾疏宁可通过干预TGF-β1/Smad信号通路减轻HBV诱导的人肾小管上皮细胞转分化,并减轻细胞外基质的沉积,从而延缓纤维化的进程。
Objective To explore the possible mechanism of Shen Shu Ning(Kidney-tonifying and Liver-soothing Formula,SSN)in the intervention of HBV-induced HK-2 transdifferentiation and delay of renal interstitial fibrosis.Methods The drug-containing serum was prepared by intragastric administration of 75 C57 BL/6 mice and models were made by using lipofectamineto transfect HBV to HK-2.The cells were divided into:blank serum control group,empty plasmid group,HBV group,benazepril group,SSN low concentration group,SSN middle concentration group,and SSN high concentration group.The expressions of HBsAg,HBeAg,TGF-β1,pSmad2,pSmad3,Smad7,Col-III,Col IV and FN in HK-2 cells were detected by enzyme-linked immunosorbent assay(ELISA),real-time fluorescent quantitative PCR and Western blot.Results ELISA detection indicated that expressions of HBsAg,HBeAg and TGF-β1 were significantly increased in HBV group compared with the empty plasmid group(P<0.01).Compared with the HBV group,the expression of TGF-β1 protein was significantly decreased in the benazepril group,SSN low concentration group,SSN middle concentration group,and SSN high concentration group(P<0.001).Compared with the benazepril group,the expression of TGF-β1 in the SSN low concentration group was relatively high(P<0.05);qPCR detection showed that,compared with the HBV group,in the benapril group,and all SSN intervention groups the expressions of Col III,Col-IV and FN mRNA were significantly decreased(P<0.001).Western blot:Compared with the HBV group,the expressions of pSmad2 and pSmad3 in the benazepril group and the SSN groups were significantly decreased(P<0.001).Conclusion SSN could possibly alleviate HBV-induced transdifferentiation of human renal tubular epithelial cells and reduce the deposition of extracellular matrix by interfering with TGF-β1/Smad signaling pathway,thereby delaying the progression of fibrosis.
作者
杨熙凯
徐冰
王耀光
Yang Xikai;Xu Bing;Wang Yaoguang(First Teaching Hospital of Tianjin University of Traditional Chinese Medicine,Tianjin 300381,China;Tianjin University of Traditional Chinese Medicine,Tianjin 301617,China)
出处
《北京中医药大学学报》
CAS
CSCD
北大核心
2019年第10期841-846,共6页
Journal of Beijing University of Traditional Chinese Medicine
基金
国家自然科学基金面上项目(No.81573888)~~
