中药复方益糖康颗粒通过AGE-RAGE轴介导SIRT1调控PI3K/Akt/FoxO1信号通路促进足细胞自噬的机制  

Mechanism of Yitangkang Granule in Promoting Podocyte Autophagy Through Regulation of PI3K/Akt/FoxO1 Signaling Pathway Mediated by SIRT1 via AGE-RAGE Axis

作　　者：程玥凤 于嘉祥 张瀚文[1] 曲超 霍易飞 张笑蕊 石岩[1] 张文顺 CHENG Yuefeng;YU Jiaxiang;ZHANG Hanwen;QU Chao;HUO Yifei;ZHANG Xiaorui;SHI Yan;ZHANG Wenshun(Liaoning University of Traditional Chinese Medicine(TCM),Shenyang 110847,China;The Second Affiliated Hospital of Liaoning University of TCM(Liaoning Institute of TCM),Shenyang 110034,China)

机构地区：[1]辽宁中医药大学,沈阳110847 [2]辽宁省中医药研究院/辽宁中医药大学附属第二医院,沈阳110034

出　　处：《中国实验方剂学杂志》2024年第17期113-121,共9页Chinese Journal of Experimental Traditional Medical Formulae

基　　金：国家重点基础研究发展计划(973计划)项目(2013CB532004);辽宁省“兴辽英才计划”青年拔尖人才项目(XLYC2007121);2023年辽宁省应用基础研究计划项目(2023JH2/101300094);辽宁省科技厅博士科研启动课题(2021-BS-183,2023-BS-141);中医脏象理论及应用教育部重点实验室2021年度开放基金项目(zyzx2109);辽宁省科技厅联合基金项目(2023-MSLH-184)。

摘　　要：目的:通过观察中药复方益糖康(YTK)颗粒对晚期糖基化终末产物(AGE)-晚期糖基化终产物受体(RAGE)轴介导沉默信息调节因子1(SIRT1)调控磷脂酰肌醇3-激酶(PI3K)/蛋白激酶B(Akt)/叉头框蛋白O1(FoxO1)信号通路改善糖尿病肾脏疾病(DKD)大鼠足细胞自噬的影响,探讨YTK治疗DKD的可能作用机制。方法:选取8周龄SPF级健康雄性Wistar大鼠96只,使用随机数字表法分为空白组、模型组、YTK高、中、低剂量组(40、20、10 g·kg^(-1))、西药组(氯沙坦片,20 mg·kg^(-1)),采用高脂饲料喂养联合腹腔注射链脲佐菌素建立DKD大鼠模型。造模成功后,各组按照相应比例剂量连续给药8周后取材。严格按照试剂盒所示方法检测超氧化物歧化酶(SOD)、丙二醛(MDA)、谷胱甘肽过氧化物酶(GSH-Px)、过氧化氢酶(CAT)的水平;苏木素-伊红(HE)染色观察肾脏组织病理形态学变化;免疫组化法检测α-平滑肌肌动蛋白(α-SMA)、纤维连接蛋白(FN)、肌间质蛋白(Desmin)、裂孔膜蛋白(Nephrin)的平均积分吸光度值;蛋白免疫印迹法(Western blot)分析DKD大鼠肾组织中滑膜PI3K、磷酸化磷脂酰肌醇3-激酶(p-PI3K)、Akt、磷酸化蛋白激酶B(p-Akt)、RAGE、SIRT1、胱天蛋白酶-3(Caspase-3)、FoxO1的蛋白表达水平。结果:与空白组比较,模型组SOD、GSH-Px、CAT表达水平明显降低,MDA显著升高(P<0.01);大鼠呈现严重肾损现象;足细胞标志蛋白α-SMA、FN、Desmin阳性表达显著增多,Nephrin、足突蛋白质(Podocin)显著下降(P<0.01);肾组织中PI3K、p-PI3K、Akt、p-Akt、RAGE、Caspase-3蛋白表达水平显著升高,SIRT1、FoxO1蛋白表达水平显著降低(P<0.01)。与模型组比较,YTK各剂量组大鼠血清中SOD、GSH-Px、CAT水平显著升高,MDA显著下降(P<0.01);肾损程度均发生了不同程度减轻;足细胞标志蛋白α-SMA、FN、Desmin平均积分吸光度值均明显降低,Nephrin、Podocin显著升高(P<0.01);肾组织中PI3K、p-PI3K、Akt、p-Akt、RAGE、CaObjective:To explore the underlying mechanism by which the Chinese medicine compound Yitangkang granule(YTK)treats diabetic kidney disease(DKD)by observing its effects on podocyte autophagy through the regulation of phosphatidylinositol 3-kinase(PI3K)/protein kinase B(Akt)/forkhead transcription factor O1(FoxO1)signaling pathway mediated by silent information regulator 1(SIRT1)via advanced glycation end products(AGE)/receptor for AGE(RAGE)axis.Method:Ninety-six 8-week-old healthy male SPF-grade Wistar rats were selected and randomly divided into blank control group(B),model control group,high-dose YTK(40 g·kg^(-1)),medium-dose YTK(20 g·kg^(-1)),low-dose YTK(10 g·kg^(-1)),and Western medicine control(20 mg·kg^(-1)losartan)groups.The DKD rat model was established by high-fat diet feeding combined with intraperitoneal injection of streptozotocin.After successful modeling,the rats in each group received the corresponding treatments for eight weeks.The levels of superoxide dismutase(SOD),malondialdehyde(MDA),glutathione peroxidase(GSH-Px),and catalase(CAT)were measured according to the instructions of the respective assay kits.Hematoxylin and eosin(HE)staining was used to observe pathological changes in kidney tissues.Immunohistochemistry was employed to detect the average optical density values ofα-smooth muscle actin(α-SMA),fibronectin(FN),desmin,and nephrin.Western blot analysis was used to measure the expression levels of PI3K,phosphorylated PI3K(p-PI3K),Akt,phosphorylated Akt(p-Akt),RAGE,SIRT1,Caspase-3,and FoxO1 proteins in kidney tissues of DKD rats.Result:Compared with the blank control group,the model group showed significantly lower levels of SOD,GSH-Px,and CAT,and significantly higher levels of MDA(P<0.01).The rats exhibited severe kidney damage.The positive expression of podocyte marker proteinsα-SMA,FN,and desmin increased significantly,while nephrin and podocin significantly decreased(P<0.01).The expression levels of PI3K,p-PI3K,Akt,p-Akt,RAGE,and Caspase-3 proteins were significantly elevated,whi

关 键 词：糖尿病肾脏疾病 中药复方益糖康颗粒 晚期糖基化终末产物(AGE)-晚期糖基化终产物受体(RAGE)轴 沉默信息调节因子1(SIRT1) 磷脂酰肌醇3-激酶(PI3K)/蛋白激酶B(Akt)/叉头框蛋白O1(FoxO1)信号通路 足细胞自噬 

分 类 号：R284[医药卫生—中药学] R285[医药卫生—中医学] R289R287R22R2-031R33R24