白藜芦醇抑制叔丁基过氧化物诱导的外周血内皮祖细胞损伤  

Resveratrol attenuates apoptosis of endothelial progenitor cells induced by tert-butyl hydroperoxide

作　　者：郑浩[1,2] 李占鲁 沈啸华 傅国胜[1] 

机构地区：[1]浙江大学医学院附属邵逸夫医院心内科,浙江杭州310016 [2]杭州市下沙医院心内科,浙江杭州310058

出　　处：《中国病理生理杂志》2017年第6期1073-1079,共7页Chinese Journal of Pathophysiology

基　　金：国家自然科学青年基金资助项目(No.30900615);浙江省自然科学基金资助项目(No.Y2090173)

摘　　要：目的:观察白藜芦醇对叔丁基过氧化物诱导的人外周血内皮祖细胞(EPC)凋亡的影响,探讨其可能机制。方法:密度梯度离心法获取人外周血单个核细胞,培养4 d后,收集贴壁细胞。实验分为对照组、叔丁基过氧化物诱导组和白藜芦醇+叔丁基过氧化物组。采用二苯基四氮唑溴盐(MTT)比色法和5-溴脱氧尿嘧啶核苷(Brd U)ELISA法检测EPC增殖能力;采用改良Boyden小室法检测EPC增殖及迁移能力;Annexin V-FITC/PI染色及流式细胞术检测EPC凋亡率;比色法检测caspase-3活性;荧光探针H_2DCF-DA法检测细胞内活性氧簇(ROS)水平;Western blot检测cleaved caspase-3、Bax及Bcl-2蛋白的表达。结果:白藜芦醇能抑制叔丁基过氧化物诱导的EPC凋亡,并增加EPC增殖及迁移能力;白藜芦醇降低EPC细胞内ROS水平,抑制caspase-3的活性及cleaved caspase-3的蛋白水平,同时能促进Bcl-2及抑制Bax蛋白表达。结论:白藜芦醇对叔丁基过氧化物诱导的EPC凋亡有抑制作用,其机制可能与抗氧化应激有关。AIM： To investigate the effect of resveratrol on the apoptosis of endothelial progenitor cells （EPC） induced by tert-butyl hydroperoxide （tBHP） and the underlying mechanisms. METHODS ： Total mononuclear cells were isolated from peripheral blood by density gradient centrifugation, and the cells were cultured on fibronectin-coated culture dishes. After 4 d of culture, attached cells were divided into control group, tBHP group, and resveratrol plus tBHP group （pretreated with resveratrol for 24 h and then cultured with 100 jjimol/L tBHP for 6 h ） . The proliferation and migration a-bilities of the EPC were assessed by MTT assay, BrdU incorporation assay and modified Boyden chamber assay. The pro-portion of apoptotic EPC was determined by flow cytometry after staining with fluorescein isothiocyanate -conjugated Annexin V and propodium iodide. The level of reactive oxygen species （ROS） was determined by H2DCF-DA method. Caspase-3 activity was assay using a caspase-3 colorimetric assay kit. The protein levels of cleaved caspase-3, Bcl-2 and Bax were de-termined by Western blot. RESULTS ： Resveratrol decreased EPC apoptosis induced by tBHP in a dose-dependent man-ner. Moreover, resveratrol increased the proliferation and migration abilities of the EPC. Resveratrol decreased intracellular ROS level, caspase-3 activity and the protein level of cleaved caspase-3. Resveratrol also decreased protein expression of Bax and increased protein expression of Bcl-2. CONCLUSION ： Resveratrol attenuates EPC apoptosis induced by oxidative stress, and its mechanisms may be related to protecting the mitochondrial function.

关 键 词：白藜芦醇 内皮祖细胞 细胞凋亡 氧化应激 

分 类 号：R363.21[医药卫生—病理学] R965[医药卫生—基础医学]