南方医科大学学报2016，Vol.36Issue(8)：1140-1143,4.DOI:10.3969/j.issn.1673-4254.2016.08.20
血清饥饿对原代人脐静脉内皮细胞周期同步化的影响
Effects of serum starvation on cell cycle synchronization in primary human umbilical vein endothelial cells
摘要
Abstract
Objective To investigate the optimal starvation conditions of human umbilical vein endothelial cells (HUVECs) and establish a highly efficient and stable method for separating HUVECs. Methods HUVECs harvested from human umbilical cords by digestion with 0.1%collagenase II for 15 min were cultured in endothelial culture medium (ECM) containing 5%fetal bovine serum (FBS), 1%endothelial cell growth factor (ECGS) and 1%penicillin/streptomycin solution(P/S) at 37℃in 5%CO2. The cells were observed for cell morphology under an inverted microscope and identified with immunofluorescence assay. The purity of HUVECs was detected using flow cytometry (FCM). The cell cycles of HUVECs cultured in the presence of 0, 0.1%, 0.5%, and 1%FBS for 0, 6, 12, 18, and 24 h were analyzed with flow cytometry. Results The purity of HUVECs harvested by digestion with 0.1%collagenase II reached 99.67%. The primary HUVECs showed a cobblestone or volute appearance in vitro. Immunocytochemistry showed that HUVECs highly expressed VIII-related antigen. Cell culture in the presence of different concentrations of FBS for 6 h resulted in 70%G0/G1 phase cells, which increased to 80%-90%at 12 h of cell culture, and further to around 95% at 18 and 24 h. Conclusion Digestion with 0.1% collagenase II can obtain high-purity primary HUVECs. Culturing HUVECs in serum-free medium for 12 h can result in a high purity (over 80%) of G0/G1 phase cells.关键词
人脐静脉内皮细胞/细胞原代培养/血清饥饿/细胞周期Key words
human umbilical vein endothelial cells/primary culture/serum starvation/cell cycle引用本文复制引用
覃乙芯,吴卓敏,徐茜,廖文婕,贺帅,岑柏宏,廖路敏,王震,季爱民..血清饥饿对原代人脐静脉内皮细胞周期同步化的影响[J].南方医科大学学报,2016,36(8):1140-1143,4.基金项目
国家自然科学基金（81370449）；广州市科技计划项目产学研协同创新重大专项（201504291720212）Supported by National Natural Science Foundation of China (81370449) （81370449）
