基于mTOR/p70S6K信号通路探究紫丁香苷对高糖诱导的小鼠足细胞MPC5损伤的影响

doi:10.3969/j.issn.1006-6233.2023.01.012

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摘要 目的: 探究紫丁香苷对高糖诱导的小鼠足细胞MPC5损伤及哺乳动物雷帕霉素靶蛋白(mTOR)/p70核糖体蛋白S6激酶(p70S6K)信号通路的影响。方法: 将小鼠足细胞MPC5分为:对照组(5.5mmoL/L葡萄糖)、高糖诱导组(30mmoL/L葡萄糖)、紫丁香苷低浓度组(2.5μmoL/L紫丁香苷+30mmoL/L葡萄糖)、紫丁香苷中浓度组(5μmoL/L紫丁香苷+30mmoL/L葡萄糖)、紫丁香苷高浓度组(10μmoL/L紫丁香苷+30mmoL/L葡萄糖)。各组向RPMI 1640培养基中加入上述浓度的相应试剂或药物,培养72h。酶标仪测定各组MPC5细胞吸光度值,计算细胞活力;流式细胞仪测定各组MPC5细胞凋亡情况;鬼笔环肽染色观察各组MPC5细胞骨架形态;荧光定量PCR和蛋白印迹法分别测定各组MPC5细胞mTOR、p70S6K信使核糖核酸(mRNA)和蛋白表达。结果: 对照组MPC5细胞骨架形态正常,细胞长轴呈线性分布,荧光染色明显;与对照组比较,高糖诱导组MPC5细胞骨架断裂,细胞萎缩,荧光染色黯淡,吸光度值、细胞活力显著降低,凋亡率、mTOR、p70S6K mRNA和蛋白表达水平显著升高(P<0.05);与高糖诱导组比较,紫丁香苷低、中、高浓度组MPC5细胞骨架结构恢复清晰,荧光强度逐渐变亮,足突趋于明显,吸光度值、细胞活力依次升高,凋亡率、mTOR、p70S6K mRNA和蛋白表达水平依次降低(P<0.05)。结论: 紫丁香苷对高糖诱导的MPC5细胞损伤具有保护作用,能明显促进MPC5细胞增殖,抑制其凋亡;其机制可能与紫丁香苷抑制高糖状态下MPC5细胞中mTOR/p70S6K通路的激活有关。

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关键词 ：紫丁香苷, 高糖, 小鼠足细胞MPC5, 哺乳动物雷帕霉素靶蛋白/p70核糖体蛋白S6激酶信号通路

Abstract：Objective: To investigate the effects of syringin on the injury of mouse podocyte MPC5 induced by high glucose and mammalian target of rapamycin (mTOR)/p70 ribosomal protein S6 kinase (p70S6K) signal pathway. Methods: MPC5 of mouse podocytes were divided into control group (5.5 mmol/L glucose), high glucose induced group (30mmol/L glucose), syringin low concentration group (2.5μmol/L syringin+30mmol/L glucose), syringin medium concentration group (5μmol/L syringin+30mmol/L glucose), syringin high concentration group (10μmol/L syringin+30mmol/L glucose). Each group added the corresponding reagent or drug of the above concentration to RPMI 1640 medium and cultured for 72h. The absorbance of MPC5 cells in each group was measured by microplate reader, and the cell viability was calculated; the apoptosis of MPC5 cells in each group was determined by flow cytometry; the cytoskeleton morphology of MPC5 cells in each group was observed by phalloidin staining; the expression of mTOR, p70S6K messenger RNA (mRNA) and protein in MPC5 cells of each group were determined by fluorescence quantitative PCR and Western blot. Results: The cytoskeleton morphology of MPC5 cells in the control group was normal, the long axis of the cells was linearly distributed, and the fluorescence staining was obvious; compared with the control group, the cytoskeleton of MPC5 cells in the high glucose induced group was broken, the cells were atrophied, the fluorescence staining was dim, the absorbance value and cell viability were significantly decreased, and the apoptosis rate, mTOR, p70S6K mRNA and protein expression levels were significantly increased (P<0.05); compared with the high glucose induced group, the cytoskeleton structure of MPC5 cells in the syringin low, medium and high concentration groups recovered clearly, the fluorescence intensity gradually became bright, the peduncle tended to be obvious, the absorbance value and cell viability increased in turn, and the apoptosis rate, mTOR, p70S6K mRNA and protein expression levels were decreased in turn (P<0.05). Conclusion: Syringin has a protective effect on MPC5 cells damage induced by high glucose, which can significantly promote MPC5 cells proliferation and inhibit its apoptosis. Its mechanism may be related to the inhibition of syringin on the activation of MPC5 cells mTORC1/p70S6K pathway under high glucose state.

Key words： Syringin High glucose Mouse podocyte MPC5 Mammalian target of rapamycin/p70 ribosomal protein S6 kinase signal pathway

基金资助:湖北省卫生计生委中医药、中西医结合科研指导性项目,(编号:鄂卫生计生通[2017]20号)

引用本文:

姚春红, 李国红, 周波, 徐佳. 基于mTOR/p70S6K信号通路探究紫丁香苷对高糖诱导的小鼠足细胞MPC5损伤的影响[J]. 河北医学, 2023, 29(1): 65-70.
YAO Chunhong, LI Guohong, ZHOU Bo, et al. Probing the Effect of Syringin on High Glucose-Induced MPC5 Damage in Mouse Podocyte Based on the mTOR/p70S6K Signaling Pathway. HeBei Med, 2023, 29(1): 65-70.

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http://www.hbyxzzs.cn/CN/10.3969/j.issn.1006-6233.2023.01.012 或 http://www.hbyxzzs.cn/CN/Y2023/V29/I1/65

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