同源盒基因Nkx2-5促进5-氮胞苷诱导的单层P19细胞表达心肌标志物

目的:探讨同源盒基因Nkx2-5基因在5-氮胞苷(5-aza)诱导单层生长的P19细胞向心肌细胞分化中的作用。方法:以稳定表达Nkx2-5的P19细胞为实验组,P19细胞为对照组。两组细胞均单层培养并以5-aza(1μmol/L)诱导,倒置显微镜下观察细胞的生长状态;在诱导后4、8、12、16 d,以RT-PCR检测转录因子GATA-4、α-肌球蛋白重链(α-MHC)和心房利钠多肽(ANP)基因的表达。结果:对照组经5-aza诱导后ANP没有表达;GATA-4在8、12、16 d有表达;α-MHC在12、16 d有表达。实验组5-aza诱导后,GATA-4在诱导4、8、12、16 d有表达;α-MHC和ANP都是在5-aza诱导后的8、12和16 d表达。两组结果比较显示实验组GATA-4和α-MHC基因的表达时间提前;两组除α-MHC在12 d的表达无差异外,其他取材时间点实验组两个基因的表达量与对照组相比均增高,差异有统计学意义(P<0.05,P<0.01)。结论:Nkx2-5促进5-aza诱导的单层生长的P19细胞向心肌分化。

Nkx2-5 gene promotes expression of cardiac markers in 5-azacytidine-induced monolayer P19 cells

Objective:To study the role of Nkx2-5 gene in 5-azacytidine-induced differentiation of monolayer P19 cells into myocardial cells.Methods:The experiment was divided into two groups:an experimental group and a control group.The cells in the experimental group were P19 cells stably expressing Nkx2-5 gene,and cells in the control group were P19 cells.Under the monolayer-culture condition,the cells of two groups were induced by 5-azacytidine(1μmol/L).The growth of cells were observed by inverted microscope.On the 4~(th)day,8~(th)day,12~(th)day and 16~(th)day after induction,RT-PCR was used to detect the expression of GATA-4,α-MHC and ANP gene.Results:In control group,there was no ANP expression after induction;GATA-4 expression was seen on the 8~(th)day,12~(th)day,and 16~(th)day after induction;and α-MHC expression was found on the 12~(th)day and 16~(th)day.In experimental group,the expression of GATA-4 was detected on the 4~(th)day,8~(th)day,12~(th)day and 16~(th) day after induction:Alpha-MHC and ANP expression was noticed on the 8~(th)day,12~(th)day and 16~(th)day after induction.RT-PCR results showed that the expression of GATA-4 and α-MHC in the experimental group was earlier than that in the control group.And at all time points of observation,the expression of GATA-4 and α-MHC in the experimental group was significantly increased compared with that in the control group(P<0.05,P<0.01),except for α-MHC expression on the 12~(th)day.Conclusion:Nkx2-5 gene can promote 5-azacytidine-induced differentiation of monolayer P19 cells into myocardial cells.