肿瘤坏死因子α致HUVECs内皮型一氧化氮合酶降解  

Degradation of e NOS induced by TNF-α in human umbilical vein endothelial cells

作　　者：徐秀丹 夏勇[1,2] 阎江洪[3] 何岸[1] 龙洋[1] 罗素新[1] 

机构地区：[1]重庆医科大学附属第一医院心血管内科,重庆400016 [2]美国俄亥俄州立大学心肺研究所,美国哥伦布43210 [3]重庆医科大学生命科学研究院,重庆400016

出　　处：《基础医学与临床》2017年第8期1067-1071,共5页Basic and Clinical Medicine

基　　金：国家重点基础研究发展计划(973计划)(2014CB542402);国家自然科学基金(81270210);重庆市科委(cstc2015shmszx0064)

摘　　要：目的研究肿瘤坏死因子-α(TNF-α)是否可以通过细胞内蛋白质降解途径引起人脐静脉内皮细胞(HUVECs)内皮型一氧化氮合酶(eNOS)的蛋白量减少。方法建立原代HUVECs培养,选取TNF-α不同浓度(0.01、0.1、1和10 ng/m L)、不同时间(24、48和72 h)处理HUVECs;溶酶体抑制剂氯化铵(NH4Cl)、caspase抑制剂(caspase inhibitor)和泛素-蛋白酶体抑制剂(MG-132)预处理HUVECs 1.5 h后加入TNF-α(1 ng/m L)共处理24 h,Western blot方法检测HUVECs中eNOS蛋白表达。结果与对照组比较,1 ng/m L TNF-α处理细胞24 h,eNOS蛋白量明显减少(P<0.01);MG-132与TNF-α共处理组,eNOS蛋白量明显增加(P<0.01)。结论 TNF-α可以通过泛素-蛋白酶体途径引起eNOS蛋白降解。Objective To investigate does intracellular protein degradation pathway play an important role in de- crease of endothelial nitric oxide synthase （eNOS） in human umbilical vein endothelial cells （HUVECs）. Methods To establish a primary HUVECs culture methods, the HUVECs were incubated with concentration gradient group of TNF-α（0.01, 0. 1, 1 and 10 ng/mL） in different time periods （24, 48 and 72 h）. The HUVECs were pretreated with NH4Cl or treated with caspase inhibitor or MG- 132 1.5 h prior to incubation for an additional 24 h with TNF-α. The expression of eNOS was detected via Western blot assay. Results Treatment of the HUVECs with TNF-α（0. 01-10 ng/mL） led to a dose-dependent reduction of eNOS expression. And treatment with TNF-α （1 ng/mL） reduced the eNOS expression in a time-depended manner. Compared with the TNF-αgroup, the protein expression level of eNOS was obviously increased in the co-working group of MG- 123 and TNF-α. Conclusions TNF-α induces degradation of eNOS through a ubiquitin-proteasome pathway.

关 键 词：HUVECS 肿瘤坏死因子-Α 内皮型一氧化氮合酶 蛋白质降解途径 MG-132 

分 类 号：R54[医药卫生—心血管疾病]