摘要
目的探讨间充质干细胞外泌体(MSC-EVs)通过调节哺乳动物雷帕霉素靶点(mTOR)/p70核糖体蛋白S6激酶(S6K1)/盘卷肌球蛋白样Bcl-2-相互作用蛋白(Beclin 1)通路改善糖尿病肾病(DN)大鼠肾损伤的机制。方法采用高脂饮食联合腹腔链脲佐菌素(STZ)注射的方法建立SD大鼠DN模型,随机分为模型组、MSC-EVs组、MSC-EVs+MHY1485(mTOR激活剂)组,每组12只。另取12只SD大鼠以正常饲料喂养6周后,腹腔注射同等剂量柠檬酸钠溶液作为对照。以MSC-EVs和MHY1485分组处理后,检测大鼠血糖及肾功能指标[血尿素氮(BUN)、血清肌酐(Scr)、尿微量白蛋白(UmALB)]水平。采用苏木精-伊红(HE)染色检测各组大鼠肾组织病理形态;采用免疫组化检测各组大鼠肾组织mTOR/S6K1/Beclin 1通路相关蛋白表达;采用蛋白免疫印迹法检测各组大鼠肾组织mTOR/S6K1/Beclin 1通路及自噬相关蛋白表达。结果与对照组相比,模型组大鼠肾组织形态发生损伤,血糖、BUN、Scr、UmALB、p-mTOR及p-S6K1相对阳性表达、p-mTOR/mTOR、p-S6K1/S6K1显著升高(P<0.05),微管相关蛋白1A/1B轻链3(LC3)Ⅱ/LC3Ⅰ、Beclin 1蛋白表达、Beclin 1相对阳性表达显著降低(P<0.05);与模型组相比,MSC-EVs组大鼠肾组织形态损伤减轻,血糖、BUN、Scr、UmALB、p-mTOR及p-S6K1相对阳性表达、p-mTOR/mTOR、p-S6K1/S6K1显著降低(P<0.05),LC3Ⅱ/LC3Ⅰ、Beclin 1蛋白表达、Beclin 1相对阳性表达显著升高(P<0.05);与MSC-EVs组相比,MSC-EVs+MHY1485组大鼠肾组织形态损伤加重,血糖、BUN、Scr、UmALB、p-mTOR及p-S6K1相对阳性表达、p-mTOR/mTOR、p-S6K1/S6K1显著升高(P<0.05),LC3Ⅱ/LC3Ⅰ、Beclin 1蛋白表达、Beclin 1相对阳性表达显著降低(P<0.05)。结论MSC-EVs可增强自噬,降低DN大鼠血糖、SCr、BUN和尿蛋白水平,减轻肾组织损伤,其机制可能与抑制mTOR/S6K1/Beclin 1通路激活有关。
Objective To explore the mechanisms of mesenchymal stem cell-derived extracellular vesicles(MSC-EVs)in improvement of renal injury in rats with diabetic nephropathy(DN)by regulating mammalian target of rapamycin(mTOR)/p70 ribosome protein S6 kinase(S6K1)/coiled-coil myosin-like Bcl-2-interacting protein(Beclin 1)pathway.Methods The model of SD rats with DN was established by a method of high-fat diet combined with intraperitoneal injection of streptozotocin,and they were randomly divided into model group,MSC-EVs group,and MSC-EVs+MHY1485(mTOR activator)group,with 12 rats in each group.Another 12 SD rats were normally fed for 6 weeks and then intraperitoneally injected with an equal dose of sodium citrate solution as controls.After grouping with MSC-EVs and MHY1485,blood glucose and levels of renal function indicators[blood urea nitrogen(BUN),serum creatinine(Scr),and urinary microalbumin(UmALB)]in rats were detected.HE staining was used to detect the pathological morphology of renal tissue in rats of each group;immunohistochemistry was used to detect the expression of mTOR/S6K1/Beclin 1 pathway related proteins in the renal tissues of rats in each group;the Western blot was used to detect the mTOR/S6K1/Beclin 1 pathway and autophagy-related protein expression in the renal tssues of rats in each group.Results Compared with the control group,the renal tissue morphology of rats in the model group were impaired,and the blood glucose,BUN,Scr,UmALB,relative positive expressions of p-mTOR and p-S6K1,p-mTOR/mTOR,p-S6K1/S6K1 increased significantly(P<0.05),while microtubule associated protein 1A/1B light chain 3(LC3)I/LC3 I,expression of Beclin 1 protein,and relative positive expression of Beclin 1 significantly reduced(P<0.05);compared with the model group,the MSC-EVs group had less impairment in the renal tissue morphology,and the blood glucose,BUN,Scr,UmALB,relative positive expressions of p-mTOR and p-S6K1,p-mTOR/mTOR,and p-S6K1/S6K1 decreased significantly(P<0.05),while expression of LC3 I/LC3 I,Beclin 1 protein,and relative positive expression of Beclin 1 increased significantly(P<0.05);compared with the MSC-EVs group,the impairment of the renal tissue morphology was worse in the MSC-EVs+MHY1485 group,and the blood glucose,BUN,Scr,UmALB,relative positive expressions of p-mTOR and p-S6K1,expression of p-mTOR/mTOR,and p-S6K1/S6K1 increased significantly(P<0.05),while the expression of LC3I/LC3I,Beclin 1 protein,and relative positive expression of Beclin 1 decreased significantly(P<0.05).Conclusion MSC-EVs can enhance autophagy,reduce levels of blood glucose,Scr,BUN and urinary protein in rats with DN,alleviate renal tissue damage,and the mechanism may be achieved by inhibiting the activation of the mTOR/S6K1/Beclin 1 pathway.
作者
吴莉莉
林景涛
张远成
钟佩敏
唐劲松
王海波
WU Lili;LIN Jingtao;ZHANG Yuancheng;ZHONG Peimin;TANG Jinsong;WANG Haibo(Department of Laboratory,Dongguan Dalang Hospital Guangdong Province,Dongguan,Guangdong,523770;Guangdong Medical University,Dongguan,Guangdong,523000)
出处
《实用临床医药杂志》
CAS
2024年第10期51-57,共7页
Journal of Clinical Medicine in Practice
基金
广东省东莞市社会科技发展重点项目(20221800905322)。
