消退素E1调控高尔基体应激抑制低氧内皮细胞损伤的作用机制
Mechanism of RvE1 inhibiting hypoxic endothelial cell injury by regulating Golgi stress
谢晓秋 1蔡缘缘 1谢和兵 2袁志兵 1龚春香3
作者信息
- 1. 安徽中医药大学药学院,安徽合肥 230012;南通市海门长三角药物高等研究院,江苏南通 226133;西藏神猴药业有限责任公司,江苏南通 226133;广西医科大学睿谷医学检验实验室,广西南宁 530012
- 2. 南通市海门长三角药物高等研究院,江苏南通 226133;西藏神猴药业有限责任公司,江苏南通 226133
- 3. 南京中医药大学附属常州中医医院病理科,江苏常州 213003
- 折叠
摘要
目的 探究高尔基体应激在低氧诱导血管内皮细胞损伤中的作用,并探讨消退素E1(RvE1)对高尔基体应激的调控作用.方法 人脐静脉内皮细胞(HUVEC)分为 20.9％O2、2％O2、2％O2+si-NC、2％O2+si-GRASP65、2％O2+RvE1、2％O2+RvE1+OE-GRASP65-NC、2％O2+RvE1+OE-GRASP65 组.采用透射电镜观察 HUVEC 高尔基体形态变化,CCK-8检测细胞活力,流式细胞术检测活性氧(ROS)水平,ELISA检测细胞培养上清内皮素1(ET-1)、前列环素-2(PGI-2)、RvE1的变化,实时荧光定量PCR(qRT-PCR)及Western blotting检测一氧化氮合成酶(eNOS)、高尔基体外周膜蛋白P65抗体(GRASP65)mRNA和蛋白表达.结果 与20.9％O2组比较,低氧可致血管内皮细胞高尔基体破裂甚至重构,显著抑制HUVEC活力(P＜0.01),显著上调ROS生成(P＜0.01),显著抑制eNOS、PGI-2、RvE1表达(P＜0.01),显著促进ET-1、GRASP65、p-GRASP65表达(P＜0.01);与2％O2组比较,si-GRASP65、RvE1 可以改善低氧HUVEC高尔基体应激反应,显著增强HUVEC活力(P＜0.01),显著下调ROS生成(P＜0.01),显著增加eNOS、PGI-2表达(P＜0.01),显著抑制ET-1、GRASP65、p-GRASP65表达(P＜0.01);OE-GRASP65对RvE1的药效有明显的抵消作用.结论 RvE1具有拮抗内皮细胞低氧损伤作用,其机制可能与其抑制低氧所致高尔基体应激有关.
Abstract
Objective To explore the role of Golgi stress in the injury of hypoxic vascular endothelial cells,and to explore the regulatory effect of fading hormone E1(RvE1)on Golgi stress.Methods Human umbilical vein endothelial cells were divided into 20.9％O2,2％O2,2％O2+si-NC,2％O2+si-GRASP65,2％O2+RvE1,2％O2+RvE1+OE-GRASP65-NC and 2％O2+RvE1+OE-GRASP65 groups.The morphological changes of HUVEC Golgi apparatus were observed by transmission electron microscopy,cell viability was detected by CCK-8,reactive oxygen species(ROS)levels were detected by flow cytometry,and the changes of endothelin-1(ET-1),prostacyclin-2(PGI-2)and RvE1 were detected by ELISA.Real-time quantitative fluorescent PCR(qRT-PCR)and Western blotting were used to detect the mRNA and protein expression of nitric oxide synthetase(eNOS)and Golgi in vitro peripheral membrane protein P65 antibody(GRASP65).Results Compared with 20.9％O2 group,hypoxia could cause rupture or even remodeling of the Golgi apparatus of vascular endothelial cells,significantly inhibit HUVEC activity(P＜0.01),significantly up-regulate ROS production(P＜0.01),and significantly inhibit the expressions of eNOS,PGI-2 and RvE1(P＜0.01).Significantly promoted the expression of ET-1,GRASP65,p-GRASP65(P＜0.01);Compared with 2％O2 group,si-GRASP65 and RvE1 could improve the stress response of hypoxic HUVEC Golgi apparatus,significantly enhance HUVEC activity(P＜0.01),significantly down-regulate ROS production(P＜0.01),and significantly increase the expressions of eNOS and PGI-2(P＜0.01).The expression of ET-1,GRASP65 and p-GRASP65 was significantly inhibited(P＜0.01).OE-GRASP65 has a significant cancelling effect on the efficacy of RvE1.Conclusions RvE1 can antagonize hypoxic injury of endothelial cells possibly by inhibiting hypoxia-induced Golgi stress.
关键词
消退素E1/低氧/内皮细胞/高尔基体应激/活性氧Key words
resolvin E1/hypoxia/endothelial cells/Golgi stress/reactive oxygen species引用本文复制引用
基金项目
国家自然科学基金资助项目(81470247)
西藏自治区科技厅区域科技协同创新专项(QYXTZX-RKZ2022-07)
出版年
2024
NETL
NSTL
万方数据