HucMSC-sEVs通过抑制肾小管上皮细胞坏死性凋亡缓解糖尿病肾病大鼠肾损伤OA
HucMSC-sEVs alleviate renal injury in diabetic nephropathy rats by inhibiting necroptosis of renal tubular epithelial cells
目的:探究人脐带间充质干细胞源小细胞外囊泡(human umbilical cord mesenchymal stem cell-derived small extracellular vesicles,HucMSC-sEVs)对糖尿病肾病(diabetic kidney disease,DKD)大鼠肾损伤的干预作用,并研究其潜在的作用机制.方法:分离培养HucMSCs,从其培养上清液中提取HucMSC-sEVs.高脂联合链脲佐菌素(streptozocin,STZ)构建DKD大鼠模型,监测大鼠空腹血糖水平,造模成功后,将大鼠随机分为DKD组和HucMSC-sEVs组,以正常大鼠为对照组.造模8周后通过大鼠尾静脉注射HucMSC-sEVs,24周后收集肾组织.HE染色检测肾组织病变程度,Masson染色观察肾组织纤维样改变;采用qRT-PCR、蛋白质印迹法和免疫组织化学染色等方法检测p-RIPK1/RIPK1,p-RIPK3/RIPK3和p-MLKL/MLKL等坏死性凋亡标志蛋白的表达水平.对大鼠肾小管上皮细胞NRK-52E进行高糖以及HucMSC-sEVs处理,透射电子显微镜观察NRK-52E细胞坏死现象,蛋白质印迹法和qRT-PCR法检测p-RIPK1/RIPK1,p-RIPK3/RIPK3和p-MLKL/MLKL等坏死性凋亡标志蛋白的表达水平.结果:与对照组相比,DKD组大鼠肾小球基底膜明显增厚,间质发生纤维样的改变,坏死性凋亡标志蛋白表达明显增加;经过HucMSC-sEVs治疗后,大鼠肾组织病理损伤和纤维化程度显著缓解,坏死性凋亡标志蛋白表达明显降低.在体外实验中,高糖处理NRK-52E细胞后,细胞呈现线粒体嵴断裂消失、内质网扩张等坏死现象,坏死性凋亡标志蛋白p-RIPK1/RIPK1,p-RIPK3/RIPK3和p-MLKL/MLKL表达水平显著增加,HucMSC-sEVs处理后细胞坏死程度显著降低,坏死性凋亡标志蛋白表达水平明显降低.结论:HucMSC-sEVs能保护肾功能延缓大鼠的DKD进展,其发挥作用的机制可能与抑制肾小管上皮细胞坏死性凋亡有关.
Objective:To investigate the therapeutic effects and underlying mechanisms of human umbilical cord mesenchymal stem cell-derived small extracellular vesicles(HucMSC-sEVs)on renal injury in diabetic kidney disease(DKD)rats.Methods:HucMSCs were isolated,cultured,and used to extract HucMSC-sEVs from their culture supernatant.A DKD rat model was induced by a high-fat diet combined with STZ injection.After successful modeling validation,rats were randomized into the DKD group and the HucMSC-sEVs group,with normal rats as controls.At 8 weeks,HucMSC-sEVs were injected via the tail vein.After 24 weeks,kidney tissues were collected.Renal histopathology was assessed by HE staining,while Masson staining was employed to evaluate collagen deposition.The expression levels of necroptosis markers(p-RIPK1/RIPK1,p-RIPK3/RIPK3,and p-MLKL/MLKL)were quantified using qRT-PCR,Western blotting,and immunohistochemical staining.NRK-52E cells were treated with high glucose and HucMSC-sEVs.Transmission electron microscopy was used to observe necrotic phenomena in NRK-52E cells,and the expression levels of necroptosis marker proteins such as p-RIPK1/RIPK1,p-RIPK3/RIPK3,and p-MLKL/MLKL were detected by Western blotting and qRT-PCR.Results:Compared with the control group,the glomerular basement membrane in the DKD group rats was significantly thickened,and the interstitium exhibited fibrotic characterization,with a marked increase in the expression of necroptosis marker proteins.After treatment with HucMSC-sEVs,the pathological damage and fibrosis degree of the rat kidney tissues were alleviated,and the expression of necroptosis execution proteins was reduced.In vitro experiments revealed that high glucose-treated NRK-52E cells exhibited necrotic features including mitochondrial cristae fragmentation,endoplasmic reticulum dilation,and elevated expression of necroptosis markers(p-RIPK1/RIPK1,p-RIPK3/RIPK3,and p-MLKL/MLKL).Treatment with HucMSC-sEVs significantly attenuated these cellular alterations and reduced the expression levels of necroptosis-related proteins.Conclusion:HucMSC-sEVs can improve renal function and delay the progression of DKD in rats,and the mechanism may be attributed to the inhibition of necroptosis of renal tubular epithelial cells.
薛玲玲;金灿;钱晖
江苏大学医学院,江苏镇江2121013江苏大学医学院,江苏镇江2121013江苏大学医学院,江苏镇江2121013
医药卫生
人脐带间充质干细胞细胞外囊泡糖尿病肾病坏死性凋亡
human umbilical cord mesenchymal stem cellssmall extracellular vesiclesdiabetic kidney diseasenecroptosis
《江苏大学学报（医学版）》 2026 (1)
36-43,8
国家自然科学基金资助项目(82172102)镇江市外泌体基础与转化应用高技术研究重点实验室(SS2018003)
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