褪黑素抑制H_(2)O_(2)诱导的人脐静脉内皮细胞氧化应激及凋亡标志蛋白质的表达  

Melatonin inhibits H_(2)O_(2)-induced oxidative stress and apoptosis marker protein expression in human umbilical vein endothelial cells

作　　者：杨瀚毅 宁佳怡 汪小兰 张益萌 谢婷珂 陈奕玄 韩静 YANG Hanyi;NING Jiayi;WANG Xiaolan;ZHANG Yimeng;XIE Tingke;CHEN Yixuan;HAN Jing(Department of Ophthalmology,Xi′an Medical University,Xi′an 710068;Department of Ophthalmology,the Second Affiliated Hospital of the Air Force Medical University,Xi′an 710038,China)

机构地区：[1]西安医学院眼科学教研室,陕西西安710068 [2]空军军医大学第二附属医院眼科,陕西西安710038

出　　处：《基础医学与临床》2024年第5期645-650,共6页Basic and Clinical Medicine

基　　金：国家自然科学基金(8217111370);陕西省科技攻关重点研发计划(2021SF-158);唐都医院临床重点研究(2021LCYJ019,2018LCYJ008)。

摘　　要：目的探讨褪黑素(MT)对过氧化氢诱导的人脐静脉内皮细胞(HUVECs)氧化应激机制与凋亡标志蛋白质的表达的影响。方法将HUVECs细胞分为空白组和H_(2)O_(2)(100、200、300、400、600和700 mmol/L)组,用CCK8法筛选出造模最佳浓度;为了明确MT对H_(2)O_(2)造成的氧化应激损伤的影响,将HUVECs细胞分为空白组、H_(2)O_(2)组和MT组。Western blot检测细胞中p-p65/p65、SOD2和cleaved-caspase 3的表达;试剂盒检测细胞中超氧化物歧化酶(SOD)活性、丙二醛(MDA)和过氧化氢酶(CAT)浓度;活性氧物质(ROS)染色检测HUVECs细胞内ROS浓度。结果相较于空白组,H_(2)O_(2)组细胞内的p-p65/p65和cleaved-caspase 3表达显著上升(P<0.001),SOD2表达明显下降(P<0.05);SOD活性(P<0.001)和CAT浓度(P<0.01)明显下降,MDA浓度明显上升(P<0.001);ROS含量显著升高(P<0.001)。相较于H_(2)O_(2)组,MT干预组细胞内的p-p65/p65(P<0.001)和cleaved-caspase 3表达(P<0.01)明显下调,SOD2表达明显增高(P<0.001);并且SOD活性(P<0.001)和CAT浓度上升(P<0.05),MDA浓度下降(P<0.001);从ROS染色的结果看来ROS含量有明显下降(P<0.001)。结论MT通过上调SOD2的表达、下调p-p65/p65和cleaved-caspase 3表达,发挥对H_(2)O_(2)介导的HUVECs氧化应激损伤的保护作用。Objective To investigate the effects of melatonin(MT)on oxidative stress and expression of apoptosis marker protein in human umbilical vein endothelial cells(HUVECs)induced by hydrogen peroxide(H_(2)O_(2)).Methods HUVECs were divided into blank group and H_(2)O_(2)(100,200,300,400,600,700 mmol/L)groups.CCK8 method was used to select the best concentration for modeling.To clarify the effect of melatonin on oxidative stress injury caused by H_(2)O_(2),HUVECs were divided into control group,H_(2)O_(2) group and melatonin group.Western blot was used to detect the expression of p-p65/p65,SOD2 and cleaved-caspase 3.The activity of super-oxide dismutase(SOD),the concentration of malondialdehyde(MDA)and catalase(CAT)were measred by commercially available kits.Reactive oxygen species(ROS)staining was used to detect ROS content in HUVECs.Results H_(2)O_(2) increased the expression of p-p65/p65 and cleaved-caspase 3(P<0.001),decreased the expression of SOD2(P<0.05)its biological activity(P<0.001)and the concentration of CAT(P<0.01),elevated the concentration of MDA(P<0.001)and the level of ROS(P<0.001)in HUVECs.Compared with H_(2)O_(2) group,MT treatment decreased the expression of p-p65/p65(P<0.001),SOD2(P<0.001)and cleaved-caspase 3(P<0.01),increased the expression of SOD2(P<0.001)as well as biological activity of SOD(P<0.001)and the concentration of CAT(P<0.05)reduced the concentration of MDA(P<0.001)and the level of ROS(P<0.001)in HUVECs.Conclusions Melatonin plays a protective role in oxidative stress injury of HUVECs induced by H_(2)O_(2) through up-regulating the expression of SOD2 and down-regulating the expression of p-p65/p65 and cleaved-caspase 3.

关 键 词：生物医学工程 褪黑素(MT) 分子生物学 人脐静脉内皮细胞(HUVECs) 氧化应激 

分 类 号：R34[医药卫生—基础医学]