淫羊藿苷对急性肾损伤向慢性肾脏病转化小鼠模型的预防性保护作用  

Preventive effect of icariin on transition from acute kidney injury to chronic kidney disease in mouse model

作　　者：谢林 程烨[2] 郑琦敏 张熙 付莉莉[3] 陈敏 王怡[2] 梅长林[3] 谢静远 顾向晨[1,2] XIE Lin;CHENG Ye;ZHENG Qimin;ZHANG Xi;FU Lili;CHEN Min;WANG Yi;MEI Changlin;XIE Jingyuan;GU Xiangchen(Department of Nephrology,Ruijin Hospital,Shanghai Jiao Tong University School of Medicine,Shanghai 200025,China;Department of Nephrology,Yueyang Hospital of Integrated Traditional and Western Medicine,Shanghai University of Traditional Chinese Medicine,Shanghai 200437,China;Department of Nephrology,The Second Affiliated Hospital of Naval Medical University(Shanghai Changzheng Hospital),Shanghai 200003,China)

机构地区：[1]上海交通大学医学院附属瑞金医院肾脏科,上海200025 [2]上海中医药大学附属岳阳中西医结合医院肾脏科,上海200437 [3]海军军医大学第二附属医院(上海长征医院)肾脏科,上海200003

出　　处：《上海交通大学学报（医学版）》2023年第1期8-19,共12页Journal of Shanghai Jiao tong University:Medical Science

基　　金：国家自然科学基金面上项目(81774106);上海市浦江人才计划(18PJD049);上海交通大学医学院“中西医结合研究平台”(2022zxy003);上海中医药大学附属岳阳中西医结合医院院级基金(2021yyjm01)。

摘　　要：目的·探究淫羊藿苷在急性肾损伤(acute kidney disease,AKI)向慢性肾脏病(chronic kidney disease,CKD)转化中的作用并探讨其可能的机制。方法·C57BL/6小鼠腹腔注射马兜铃酸2周,制作AKI向CKD转化模型,并于造模前1周或马兜铃酸注射2周结束时持续灌胃给予淫羊藿苷5周或2周。监测小鼠肾功能和肾脏病理损伤,并用透射电子显微镜观察肾小管上皮细胞线粒体结构,采用荧光定量PCR(qPCR)及免疫组织化学染色法检测脂肪酸氧化代谢通路、纤维化及炎症指标等。采用Transwell细胞小室共培养大鼠肾脏成纤维细胞株和大鼠肾小管上皮细胞株,模拟2种细胞之间的相互作用。先提前1 h在肾小管上皮细胞培养液中加入淫羊藿苷,再加入马兜铃酸干预24 h;去除药物干预后,将肾小管上皮细胞与成纤维细胞共培养24 h。qPCR或Western blotting检测肾脏上皮细胞过氧化物酶体增殖物激活受体α(peroxisome proliferator activated receptorα,Ppara)、促纤维化因子、炎症因子、活化胱天蛋白酶(cleaved caspase-3),以及肾脏成纤维细胞细胞外基质相关基因mRNA或蛋白表达的变化。用Ppara-siRNA转染肾小管上皮细胞,观察淫羊藿苷提前于马兜铃酸干预后PPARα下游线粒体脂肪酸氧化代谢相关指标以及cleaved caspase-3的表达变化。结果·马兜铃酸注射2周结束时和注射结束后2周(造模4周),仅注射马兜铃酸的小鼠血清肌酐及尿素氮水平较对照组均显著升高,肾脏炎症细胞大量浸润,肾小管损伤严重;而提前给予淫羊藿苷干预的小鼠较马兜铃酸组肾功能明显改善,肾脏病理损伤减轻。造模4周时马兜铃酸组肾小管上皮细胞线粒体结构损伤严重,脂肪酸氧化代谢通路相关基因mRNA水平较对照组显著降低,伴巨噬细胞浸润及肾纤维化,提前给予淫羊藿苷干预后上述损伤或指标均明显缓解。在马兜铃酸注射2周结束时给予淫羊藿苷的小鼠血清肌酐和尿�Objective·To investigate the effect of icariin on the transition from acute kidney disease(AKI)to chronic kidney disease(CKD)in mice and the possible mechanism.Methods·C57BL/6 mice were intraperitoneally injected with aristolochic acid for 2 weeks to mimic AKI to CKD transition.Icariin was intragastrically administered for 2 or 5 weeks at the end of 2 weeks of aristolochic acid injection or 1 week before modeling.The renal function and pathological injury of the mice were monitored.The mitochondrial structure of renal tubular epithelial cells was observed by transmission electron microscopy.The indexes related to fatty acid oxidative metabolic pathway,fibrosis,and inflammation were detected by quantitative PCR(qPCR)and immunohistochemical staining.The rat renal fibroblast cell line and the rat renal tubular epithelial cell lines were co-cultured in Transwell cell chambers to simulate the interaction between the two types of cells.The renal tubular epithelial cells were pretreated with icariin for 1 h,and then treated with aristolochic acid for 24 h.After removing the two drugs,the renal tubular epithelial cells and the fibroblasts were co-cultured for 24 h.The mRNA or protein expression levels of peroxisome proliferator-activated receptorα(Ppara),pro-fibrotic factors,inflammatory factors,activated caspase-3(cleaved caspase-3),and extracellular matrix-associated genes in renal fibroblasts were detected by qPCR or Western blotting.The renal tubular epithelial cells were transfected with Ppara-siRNA to observe the expression changes of cleaved caspase-3 and the related indicators of mitochondrial fatty acid oxidative metabolism in the downstream of PPARαafter icariin intervention prior to aristolochic acid.Results·Compared with the control group,the serum creatinine and urea nitrogen levels of the mice injected with aristolochic acid only were significantly higher,with a large number of inflammatory cells infiltrating into the kidney and serious renal tubular injury at the end of 2 weeks of aristolochic acid in

关 键 词：淫羊藿苷 急性肾损伤 慢性肾脏病 线粒体 脂肪酸氧化 肾小管上皮细胞 

分 类 号：R286[医药卫生—中药学]