Effects of hypoxia on maintenance of pluripotency and stability of imprinted gene expression in human embryonic stem cells

Objective To investigate the effect of hypoxia on maintenance of biological characteristics and imprinted genes expression in human embryonic stem cells (hESCs).Methods The FY-hES-7 cells were cultured under hypoxic conditions( 5％ O2 ) followed by identification of features of stem cells by using cytologic fluorescent immunoassaying and reverse transcriptase polymerase chain reaction.Expression of 55 imprinted genes during the early ( 32nd passage) and late phase (52nd passage) was determined via whole genome expression genechip assaying and real-time polymerase chain reaction,with the cells cultured under normoxia condition (21％ O2) for control.Results Both FY-hES-7 cells cultured under hypoxia and normoxia conditions yielded comparable biological characteristics of hESCs and positive expression rates of stem cell surface markers,namely SSEA-3,SSEA-4,TRA-1-60,TRA-1-81 and alkaline phosphatase.Notably,the cells cultured under hypoxia condition were associated with typical cloning morphology and prolonged maintenance of undifferentiated state.Oxygen concentration was found as having more profound influence(P＜0.05) on 55 imprinted genes in FY-hES-7 cells cultured under normoxia condition,suggested by altered expression (21 up-regulated and 18 down-regulated) in 39 genes(70.91％),as compared with those cultured under hypoxia condition evidenced by varied expression(8 up-regulated and 7 down-regulated) in 15 genes(27.27％).Conclusion Hypoxia favors maintenance of the biological features of hESCs,and on the basis of prolonged culture,may result in marked impact on imprintedgene expression,as evidenced by higher stability when compared with those cultured under normoxia condition. Key words: Cell hypoxia; Embryonic stem cells; Imprinted genes; Multipotent stem cells; Stability