Effects of serum starvation on cell cycle synchronization in primary human umbilical vein endothelial cells
QIN Yixin
WU Zhuomin
XU Qian
LIAO Wenjie
HE Shuai
CEN Bohong
LIAO Lumin
WANG Zhen
JI Aimin
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.
Keywords：human umbilical vein endothelial cellsprimary cultureserum starvationcell cycle
Publication Date：2016-01-01
Online Publishing Date：2025-08-15(First online date of this platform, not the publication date of the document)
Pages：4( 1140-1143 )