摘要
目的:探讨线粒体钙离子单向转运体(mitochondrial calcium uniporter,MCU)在高糖(high glucose,HG)诱导心肌H9c2细胞凋亡中的作用机制。方法:将心肌H9c2细胞随机分为3组:对照(control)组,5.5 mmol/L葡萄糖处理细胞;HG组,25 mmol/L葡萄糖处理细胞;精胺(spermine,Sp)干预(HG+Sp)组,25 mmol/L葡萄糖和5μmol/L Sp共同处理细胞。Western blot检测H9c2细胞MCU、caspase-9和caspase-3蛋白的表达;RT-qPCR检测H9c2细胞MCU的mRNA水平;Rhod-2 AM探针检测线粒体内Ca2+的荧光强度;吸光度法检测丙酮酸脱氢酶(pyruvate dehydrogenase,PDH)的活性;萤火虫萤光素酶检测细胞裂解液ATP的浓度;JC-1染色法检测线粒体膜电位(mitochondrial membrane potential,Δψm);MitoSOXTM染色法检测线粒体活性氧簇(reactive oxygen species,ROS)水平。结果:与control组相比,HG组MCU mRNA和蛋白水平、线粒体内Ca2+浓度、PDH活性、细胞ATP浓度及Δψm降低(P<0.05),而ROS水平及caspase-9和caspase-3凋亡蛋白表达增加(P<0.05);与HG组相比,HG+Sp组线粒体内Ca2+浓度、PDH活性、细胞ATP浓度和Δψm增加(P<0.05),而ROS水平及caspase-9和caspase-3凋亡蛋白表达降低(P<0.05)。结论:高糖通过降低MCU表达导致其活性下降,从而促进心肌H9c2细胞凋亡,其机制可能与线粒体的钙离子稳态失衡、三羧酸循环障碍和线粒体功能损伤有关。
AIM: To investigate the role of mitochondrial calcium uniporter(MCU) in high glucose(HG)-induced apoptosis of cardiac myocytes. METHODS: Cardiac myocytes were exposed to normal glucose(5.5 mmol/L glucose+19.5 mmol/L mannitol), HG(25 mmol/L glucose), or HG combined with 5 μmol/L spermine for 72 h. Mitochondrial free Ca2+ concentration([Ca2+]m), MCU at mRNA and protein levels, pyruvate dehydrogenase(PDH) activity, mitochondrial membrane potential(Δψm), the levels of ATP and reactive oxygen species(ROS), and apoptosis were determined. RESULTS: The [Ca2+]m, the mRNA and protein levels of MCU, PDH activity, ATP levels, and Δψm were reduced(P<0.05), while ROS content and the protein levels of caspase-9 and caspase-3 were increased in HG group(P<0.05). Adding 5 μmol/L spermine returned these parameters toward control levels(P<0.05). Moreover, apoptosis was reduced by adding spermine and HG treatment(P<0.05). CONCLUSION: HG-induced cardiac myocyte apoptosis may be associated with the decreased MCU expression and activity, abnormal mitochondrial Ca2+ handling, deviant mitochon-drial respiratory chain, and mitochondrial dysfunction.
作者
郑文学
荆哲
郭文昀
张涛
陈霞
吴兆琦
崔恒强
杨红宁
张玉秀
惠玲
陈永清
ZHENG Wen-xue;JING Zhe;GUO Wen-yun;ZHANG Tao;CHEN Xia;WU Zhao-qi;CUI Heng-qiang;YANG Hong-ning;ZHANG Yu-xiu;HUI Ling;CHEN Yong-qing(Department of Cardiology,The 940th Hospital of Joint Logistics Support Force of People’s Liberation Army,Lanzhou 730000,China;Department of Clinical Laboratory,The 940th Hospital of Joint Logistics Support Force of People’s Liberation Army,Lanzhou 730000,China)
出处
《中国病理生理杂志》
CAS
CSCD
北大核心
2020年第3期433-438,共6页
Chinese Journal of Pathophysiology
基金
甘肃省自然科学基金项目(No.18JR3RA402)
全军保健专项(No.14BJZ25)。
关键词
糖尿病
心肌细胞
线粒体钙离子单向转运体
活性氧簇
细胞凋亡
Diabetes mellitus
Cardic myocytes
Mitochondrial calcium uniporter
Reactive oxygen species
Apoptosis
