芪地糖肾方调控Akt1/HIF-1α/Bcl-xl信号通路提高糖尿病肾病足细胞自噬的机制  

Qidi Tangshen Prescription(QDTS)Regulate Akt1/HIF-1α/Bcl-xl Signaling Pathway to Improve Podocyte Autophagy in Diabetic Nephropathy

作　　者：高飞 谢惠迪[3] 于博睿 周盈 史扬 张先慧 柳红芳[2] GAO Fei;XIE Huidi;YU Borui;ZHOU Ying;SHI Yang;ZHANG Xianhui;LIU Hongfang(Beijing Chaoyang District Hospital of Traditional Chinese Medicine,Beijing 100020,China;Dongzhimen Hospital,Beijing University of Chinese Medicine,Beijing 100700,China;First Medical Center of Chinese PLA General Hospital,Beijing 100853,China;Beijing Puren Hospital,Beijing 100062,China)

机构地区：[1]北京市朝阳区中医医院,北京100020 [2]北京中医药大学东直门医院,北京100700 [3]中国人民解放军总医院第一医学中心,北京100853 [4]北京市普仁医院,北京100062

出　　处：《中国实验方剂学杂志》2024年第15期90-97,共8页Chinese Journal of Experimental Traditional Medical Formulae

基　　金：国家自然科学基金项目(81774273);北京中医药大学“揭榜挂帅”项目(2023-JYBJBZD-023)。

摘　　要：目的:探讨芪地糖肾方(QDTS)减轻糖尿病肾病(DKD)足细胞损伤,降低尿蛋白的作用机制。方法:使用网络药理学方法,收集QDTS中的化学成分和靶标,以及DKD相关靶标,构建QDTS治疗DKD的药物-成分-靶标-疾病交互网络,系统阐释其作用机制。体内实验将db/db小鼠分为模型组、芪地糖肾方(3.34 g·kg^(-1))和缬沙坦胶囊(10.29 mg·kg^(-1))组,db/m小鼠作为正常组,每组8只,连续干预8周。末次给药后处死小鼠,检查小鼠尿微量白蛋白排泄率(UAER)、肾脏病理变化及蛋白激酶B1(Akt1)、低氧诱导因子-1α(HIF-1α)、磷酸化B细胞淋巴瘤放大因子(p-Bcl-xl)和自噬活性指标微管相关蛋白1轻链3(LC3)、p62、自噬关键分子酵母Atg6同系物(Beclin1)蛋白表达水平。体外实验将小鼠足细胞分为正常糖组(5.5 mmol·L^(-1)葡萄糖),高糖组(35 mmol·L^(-1)),二甲基亚砜(DMSO)组(35 mmol·L^(-1)葡萄糖+200 mg·L^(-1)DMSO)和QDTS组(35 mmol·L^(-1)葡萄糖+200 mg·L^(-1)QDTS冻干粉),干预48 h后检测足细胞中Akt1、HIF-1α、磷酸化(p)-Bcl-xl、LC3、p62、Beclin1蛋白表达水平。结果:QDTS治疗DKD的有效成分有34种,作用靶点143个,其中与自噬相关的靶点有55个,Akt1、HIF-1α和Bcl-xl是其关键靶点。与正常组比较,模型组小鼠UAER显著升高,小鼠肾小球肥大,蓝色胶原纤维沉积,肾小球基底膜增厚,部分节段伴有明显足细胞足突融合,小鼠肾脏中p-Akt1、HIF-1α、p62蛋白表达增强,p-Bcl-xl、LC3和Beclin1蛋白表达明显减弱(P<0.05);与模型组比较,QDTS组和缬沙坦组小鼠UAER含量明显降低(P<0.05),小鼠肾脏病理损伤明显减轻,小鼠肾脏中p-Akt1、HIF-1α和p62蛋白表达减弱,p-Bcl-xl、LC3和Beclin1蛋白表达增强(P<0.05);与正常糖组比较,高糖组小鼠足细胞中p-Akt1、HIF-1α和p62蛋白表达增强,p-Bcl-xl、LC3和Beclin1蛋白表达减弱(P<0.05);与高糖组比较,QDTS组小鼠足细胞中p-Akt1、HIF-1α和p62蛋白表达减弱,p-Bcl-xl、LC3和Beclin1蛋Objective:To explore the mechanism of Qidi Tangshen prescription(QDTS)in alleviating podocyte injury and reducing urinary protein in diabetic nephropathy(DN).Method:Using network pharmacology methods,we collected the chemical components and targets of QDTS,as well as the targets related to DN.Subsequently,we constructed a"drug-ingredient-target-disease"network for QDTS in the treatment of DN to systematically elucidate the mechanism.The db/db mice were assigned into the model,QDTS(3.34 g·kg^(-1)),and losartan capsules(10.29 mg·kg^(-1))groups,and db/m mice served as the normal group.Each group consisted of 8 mice,and they underwent continuous intervention for 8 weeks.After the last administration,mice were euthanized,and the urinary albumin excretion rate(UAER)and renal pathological changes were measured and observed.The expression levels of protein kinase B1(Akt1),hypoxia-inducible factor-1 alpha(HIF-1α),phosphorylated B-cell lymphoma-extra-large(p-Bcl-xl),as well as autophagy-related indicators microtubule-associated protein 1 light chain 3(LC3),ubiquitin-binding protein p62(p62),and autophagy-related gene 6 homolog(Beclin1),were determined.Furthermore,mouse podocytes were divided into the normal glucose(5.5 mmol·L^(-1)),high glucose(35 mmol·L^(-1)),DMSO(35 mmol·L^(-1) glucose+200 mg·L^(-1) DMSO),and QDTS(35 mmol·L^(-1) glucose+200 mg·L^(-1) QDTS freeze-dried powder)groups.After 48 h of intervention,the protein levels of Akt1,HIF-1α,p-Bcl-xl,LC3,p62,and Beclin1 in podocytes were measured.Result:QDTS had 34 active components acting on 143 targets in the treatment of DN,and 55 targets were related to autophagy,in which Akt1,HIF-1α,and Bcl-xl were the key targets.Compared with the normal group,mice in the model group exhibited significantly increased UAER,glomerular hypertrophy,deposition of blue collagen fibers,thickening of the glomerular basement membrane,and noticeable fusion of podocyte foot processes in some segments.Furthermore,the modeling up-regulated the protein levels of p-Akt1,HIF-1α,and p62

关 键 词：芪地糖肾方 糖尿病肾病 足细胞损伤 自噬 信号通路 分子机制 

分 类 号：R2-0[医药卫生—中医学] R33R289R575.1R692