SIRT3抑制线粒体自噬并减轻高糖加重的神经元缺氧再灌注损伤  

SIRT3 Inhibits Mitophagy and Alleviates Neuronal Oxygen Deprivation/Reoxygenation Injury Aggravated by High Glucose

作　　者：张晨阳 黑常春[2] 袁仕林 周玉佳 曹美玲 秦亦欣 杨笑[3] ZHANG Chen-yang;HEI Chang-chun;YUAN Shi-lin;ZHOU Yu-jia;CAO Mei-ling;QIN Yi-xin;YANG Xiao(School of Clinical Medicine,Ningxia Medical University,Yinchuan 750004,China;Department of Human Anatomy,Histology and Embryology,Ningxia Medical University,Yinchuan 750004,China;Neuroscience Center,General Hospital of Ningxia Medical University,Yinchuan 750004,China)

机构地区：[1]宁夏医科大学临床医学院,银川750004 [2]宁夏医科大学基础医学院人体解剖与组织胚胎学系,银川750004 [3]宁夏医科大学总医院神经内科,银川750004

出　　处：《中国生物工程杂志》2021年第11期1-13,共13页China Biotechnology

基　　金：国家自然科学基金(82060237、81560226、81860250);宁夏自然科学基金(2018AAC03134、2021AAC03371)资助项目。

摘　　要：目的:探讨SIRT3调控的线粒体自噬对高糖加重神经元缺氧再灌注损伤的影响及机制。方法:高糖(50 mmol/L)干预HT22细胞后,构建细胞缺氧/复氧模型,利用SIRT3抑制剂3-TYP抑制SIRT3表达。倒置显微镜观察细胞形态改变,CCK8法检测细胞存活率,流式细胞术检测细胞凋亡率,TMRE荧光试剂盒检测细胞线粒体膜电位,RT-qPCR、Western blot检测相关分子的基因和蛋白质表达。结果:高糖使神经元缺氧再灌注后的细胞碎片进一步增加,细胞存活率降低,细胞凋亡率升高(P<0.05)。此外,高糖降低了神经元缺氧再灌注后的线粒体膜电位(P<0.05)。进一步研究发现,高糖上调神经元缺氧再灌注后线粒体分裂相关蛋白DRP1的表达水平,降低了线粒体融合相关蛋白OPA1和线粒体外膜蛋白TOM20的表达;并且增加了自噬相关蛋白LC3Ⅱ、Beclin-1和线粒体自噬相关蛋白PINK1、Parkin的表达;同时,高糖升高了SIRT3的基因和蛋白质表达(P<0.05)。而SIRT3抑制剂3-TYP使神经元高糖缺氧再灌注损伤加重,同时进一步上调DRP1、LC3Ⅱ和PINK1的蛋白质表达(P<0.05)。结论:高糖可显著加重神经元缺氧再灌注损伤,破坏细胞线粒体功能,激活细胞线粒体自噬;SIRT3可抑制PINK1-Parkin通路介导的线粒体自噬并减轻神经元高糖缺氧再灌注损伤。Objective:To investigate whether high glucose can aggravate oxygen deprivation/reoxygenation(OD/R)injury in neurons through regulating mitophagy and the specific molecular mechanism of regulating mitophagy.Methods:HT22 cells are used to establish a model of cellular oxygen deprivation/reoxygenation to simulate cerebral ischemia/reperfusion injury in vivo.And cells are treated with high glucose medium(HG 50 mmol/L).3-TYP is given to inhibit the expression of SIRT3.CCK8 is used to examine cell viability;flow cytometry is used to detect cell apoptosis,and TMRE fluorescence detection kit is used to detect cell MMP;Western blot is used to detect the expression of mitochondrial division/fusion related protein(DRP1,OPA1,TOM20),autophagy related protein(LC3Ⅱ,Beclin-1),mitophagy related protein(PINK1,Parkin)and SIRT3.Results:Compared with OD groups,the cell density is further reduced,the cell contour is more blurred,the synapses are shortened,the floating cells increase in HG+OD groups,and cells in HG+OD groups have lower survival rate and higher apoptosis rate(P<0.05).Compared with OD groups,cells in HG+OD groups have lower MMP(P<0.05).Compared with OD groups,the expression of P-DRP1 in HG+OD groups is increased,but OPA1 and TOM20 are decreased(P<0.05);the expression of Beclin-1 and LC3Ⅱ as well as PINK1 and Parkin in HG+OD groups are also increased(P<0.05).Furthermore,SIRT3 protein and gene are further increased in HG+OD groups(P<0.05).3-TYP treatment can further aggravate the OD/R injury of neurons in HG groups.And 3-TYP also increases the expression of DRP1,LC3Ⅱ and PINK1.Conclusions:High glucose can aggravate oxygen deprivation/reoxygenation injury in neurons by exacerbation of cell apoptosis,reduction of MMP and activation of mitophagy.In addition,SIRT3 can inhibit PINK1-Parkin pathway-mediated mitophagy and alleviate high glucose aggravated-neuronal OD/R injury.

关 键 词：缺氧再灌注 高糖 线粒体自噬 线粒体融合分裂 SIRT3 

分 类 号：Q2[生物学—细胞生物学]