过刊浏览

外源性硫化氢预处理通过减轻氧化应激延缓人脐静脉内皮细胞老化

齐洪娜, 崔洁, 刘磊, 卢飞飞, 宋成洁, 石玥, 闫长栋

徐州医学院生理学教研室；心血管研究所；江苏省麻醉学重点实验室，徐州 221002

摘要

本文旨在观察硫化氢(hydrogen sulfide, H₂S)供体硫氢化钠(NaHS)预处理对人脐静脉内皮细胞老化的影响及机制。原代人脐静脉内皮细胞株(HUVECs)传代培养到第12代为老年组；自第4代细胞(年轻组)起，给予NaHS孵育到第12代即为NaHS组。细胞衰老β-半乳糖苷酶(SA β-gal)染色法检测细胞老化程度，DAPI荧光染色法检测细胞凋亡，免疫印迹法测定细胞黄嘌呤氧化酶(XOD)、锰-超氧化物歧化酶(Mn-SOD)和烟酰胺腺嘌呤二核苷酸磷酸(NADPH)氧化酶亚单位蛋白表达，化学比色法检测HUVECs内SOD活性和过氧化氢(H₂O₂)含量。结果显示，与年轻组相比，老年组内皮细胞SA β-gal阳性率和凋亡率显著增加(P < 0.01)，而NaHS预处理能明显减少细胞SA β-gal阳性率和凋亡率(P < 0.01)。与年轻组相比，老年组内皮细胞XOD和NADPH氧化酶亚单位p67phox蛋白表达增加，Mn-SOD蛋白表达减少；NaHS组与老年组相比，XOD和p67phox蛋白表达减少，Mn-SOD蛋白表达增加(P < 0.05)。与年轻组相比，老年组SOD活性降低，H2O2含量增加；NaHS组与老年组相比，SOD活性提高，H2O2含量减少(P < 0.01)。以上结果提示，NaHS预处理能减轻老化HUVECs氧化应激水平，起到延缓HUVECs老化的作用。

关键词： 硫化氢; 老化; 人脐静脉内皮细胞; 预处理; 氧化应激

分类号：R339.3+8

[Exogenous hydrogen sulfide delays the senescence of human umbilical vein endothelial cells by lessening oxidative stress.] [Article in Chinese]

QI Hong-Na, CUI Jie, LIU Lei, LU Fei-Fei, SONG Cheng-Jie, SHI Yue, YAN Chang-Dong

Department of Physiology； Institute of Cardiovascular Disease； Jiangsu Province Key Laboratory of Anesthesiology, Xuzhou Medical College, Xuzhou 221002, China

Abstract

The present study was aimed to investigate the effect of pretreatment with hydrogen sulfide (H2S) on human umbilical vein endothelial cells (HUVECs) senescence and the underlying mechanism. Cultured HUVECs at twelfth and fourth passages were taken as old and young groups, respectively. Sodium hydrosulfide (NaHS, donor of H2S) group was treated with NaHS from fourth to twelfth passage. The cell senescence was determined by senescence-associated β-galactosidase (SA β-gal) staining. DAPI fluorescent dye was used to detect cellular apoptosis. Western blot was used to analyze the expression levels of xanthine oxidase (XOD), manganese-superoxide dismutase (Mn-SOD) and the subunits p67(phox) of nicotinamide adenine dinucleotide phosphate (NADPH) oxidase in the HUVECs. Colorimetric method was used to detect SOD activity and cellular hydrogen peroxide (H2O2) level. The results showed that, compared with young group, the old group exhibited higher SA β-gal positive rate and cellular apoptosis, while NaHS pretreatment decreased SA β-gal positive rate and cellular apoptosis. Compared with the young group, the old group showed increased expression levels of XOD and p67(phox), as well as lower Mn-SOD expression level. With the pretreatment of NaHS, the up-regulations of XOD and p67(phox) levels and down-regulation of Mn-SOD level were inhibited. Compared with the young group, the old group showed lower SOD activity and higher H2O2 level, whereas NaHS pretreatment reversed the changes of SOD activity and H2O2 level. These results suggest that H2S delays senescence of HUVECs through lessening oxidative stress.

Key words: hydrogen sulfide; senescence; human umbilical vein endothelial cells; preconditioning; oxidative stress

收稿日期：2012-03-01　　录用日期：2012-06-01

通讯作者：闫长栋　　E-mail: yancd55@163.com

引用本文：

齐洪娜, 崔洁, 刘磊, 卢飞飞, 宋成洁, 石玥, 闫长栋. 外源性硫化氢预处理通过减轻氧化应激延缓人脐静脉内皮细胞老化[J]. 生理学报 2012; 64 (4): 425-432.

QI Hong-Na, CUI Jie, LIU Lei, LU Fei-Fei, SONG Cheng-Jie, SHI Yue, YAN Chang-Dong. [Exogenous hydrogen sulfide delays the senescence of human umbilical vein endothelial cells by lessening oxidative stress.] [Article in Chinese] . Acta Physiol Sin 2012; 64 (4): 425-432 (in Chinese with English abstract).