基于脂噬介导的mTOR/TFEB信号通路探讨当归红芪超滤物干预糖尿病肾病作用机制  

Discussion on the Mechanism of Intervention of Dangui Hongqi Ultrafiltration in Diabetic Nephropathy Based on Lipophage Mediated mTOR/TFEB Signaling Pathway

作　　者：高婷 李荣科[1] 万生芳[1] 张磊[1] 张小琳 徐庆全 GAO Ting;LI Rongke;WAN Shengfang;ZHANG Lei;ZHANG Xiaolin;XU Qingquan(Gansu University of Chinese Medicine,Lanzhou 730000,China)

机构地区：[1]甘肃中医药大学,甘肃兰州730000

出　　处：《中国中医药信息杂志》2024年第6期66-72,共7页Chinese Journal of Information on Traditional Chinese Medicine

基　　金：国家自然科学基金(82060914);甘肃中医药大学中医学一级学科“岐黄英才”导师专项基金(2022年);甘肃省高等学校青年博士基金(2022QB-096);甘肃省省级科技计划青年科技基金(21JR1RA274)。

摘　　要：目的以当归红芪超滤物干预糖尿病肾病模型大鼠,探讨其对糖尿病肾病脂噬的效应机制。方法50只SPF级雄性Wistar大鼠随机选取7只为空白组,其余43只采用一次性大剂量腹腔注射链脲佐菌素联合高脂高糖饲料喂养制备糖尿病肾病模型。将成模大鼠随机分为模型组、厄贝沙坦组(17.9 mg/kg)和中药低、中、高剂量组(1.5、3、6g/kg),分别灌胃相应溶液,连续12周。检测大鼠随机血糖、体质量及24h尿蛋白含量,生化检测糖化血清蛋白(GSP)、血肌酐(SCr)、血尿素氮(BUN)、三酰甘油(TG)、总胆固醇(TC)、高密度脂蛋白胆固醇(HDL-C)、低密度脂蛋白胆固醇(LDL-C)、游离脂肪酸(FFA)含量,HE染色观察肾组织形态变化,Masson染色观察肾组织纤维化情况,透射电镜观察肾组织超微结构,RT-qPCR检测肾组织哺乳动物雷帕霉素靶蛋白(mTOR)、微管相关蛋白1轻链3(LC3B)mRNA表达,Westernblot检测肾组织mTOR、转录因子EB(TFEB)、p-TFEB、LC3B蛋白表达。结果与空白组比较,模型组大鼠随机血糖显著升高(P<0.01),体质量显著减少(P<0.01),24h尿蛋白含量显著升高(P<0.01),GSP、SCr、BUN、TG、TC、LDL-C、FFA含量显著升高(P<0.01),HDL-C含量显著降低(P<0.01);肾小管上皮细胞空泡变性、胞质空泡化、有大量炎性细胞浸润,胶原纤维增多且分布杂乱,脂滴数量增加,自噬体数量减少;肾组织mTORmRNA表达显著升高(P<0.01),LC3BmRNA表达显著降低(P<0.01),mTOR、p-TFEB蛋白表达显著升高(P<0.01),TFEB、LC3BⅡ蛋白表达显著降低(P<0.01)。与模型组比较,各给药组大鼠给药12周随机血糖、24h尿蛋白含量降低,体质量增加,GSP、SCr、BUN、TG、TC、LDL-C、FFA含量降低,HDL-C含量升高;肾组织炎性细胞浸润、纤维化、脂滴沉积有不同程度缓解,肾组织mTORmRNA表达降低,LC3BmRNA表达升高,mTOR、p-TFEB蛋白表达降低,TFEB、LC3BⅡ蛋白表达升高,其中厄贝沙坦组和中药高剂量组差异均有统计学意义(P<Objective To explore the effect mechanism of lipophagy in diabetic nephropathy model rats based on intervention of Dangui Hongqi ultrafiltration in diabetic nephropathy model rats.Methods Fifty male Wistar rats of SPF grade were randomly selected 7 rats as the blank group,and the other 43 rats were given a one-time high-dose intraperitoneal injection of streptozotocin combined with high-fat and high-sugar diet to prepare diabetes nephropathy model.The modeling rats were randomly divided into model group,irbesartan group(17.9 mg/kg),and TCM low-,medium-,and high-dosage groups(1.5,3,6 g/kg).The rats were continuously gavaged by corresponding drugs for 12 weeks.Randomized blood glucose,body mass,and 24 h urine protein content were detected;biochemical tests were used for GSP,SCr,BUN,TG,TC,HDL-C,LDL-C,FFA content;HE staining was used to observe morphology changes in renal tissue,Masson staining was used to observe fibrosis changes in renal tissue,renal tissue microstructure changes were observed under transmisson electron microscopy,RT-qPCR was used to detect the mRNA expressions of mTOR and LC3B in renal tissue,and Western blot was used to detect the protein expressions of mTOR,TFEB,p-TFEB,and LC3B in renal tissue.Results Compared with the blank group,the model group rats showed a significant increase in randomized blood glucose(P<0.01),a significant decrease in body mass(P<0.01),a significant increase in 24 h urine protein content(P<0.01),a significant increase in GSP,SCr,BUN,TG,TC,LDL-C,FFA content(P<0.01),and a significant decrease in HDL-C content(P<0.01);renal tubular epithelial cells underwent vacuolar degeneration,cytoplasmic vacuolization,and extensive infiltration of inflammatory cells,collagen fibers increased and were distributed randomly,the number of lipid droplets increased,and autophagosomes decreased;the mTOR mRNA expressions in renal tissue significantly increased(P<0.01),the LC3B mRNA expressions significantly decreased(P<0.01),the mTOR and p-TFEB protein expressions significantly increased(P<0.01)

关 键 词：糖尿病肾病 当归红芪超滤物 脂噬 mTOR/TFEB信号通路 大鼠 

分 类 号：R285.5[医药卫生—中药学]