地西他滨联合丙戊酸增强HL-60细胞hPer3基因的表达

目的探讨地西他滨(DCA)和丙戊酸(VPA)联用对白血病细胞株HL-60的作用及对human peri-od3基因(hPer3)的表达调控的影响。方法 DCA 1.0及4.0μmol·L-1,VPA 2.0 mmol·L-1,VPA 2.0 mmol·L-1+DCA 1.0μmol·L-1,VPA 2.0 mmo.L-1+DCA 4.0μmol·L-1作用HL-60细胞48 h。MTT法检测细胞存活,FITC-AnnexinⅤ/PI检测细胞凋亡,甲基化聚合酶链反应和实时荧光定量PCR检测hPer3基因启动子甲基化状态和mRNA表达,流式细胞术检测CD14表达率。结果 VPA 2.0+DCA 1.0联合用药组生长抑制率为(49.6±5.2)%,VPA 2.0+DCA 4.0联合用药组为(66.3±7.3)%,均高于其相应单药组,差异具有统计学意义(P<0.01)。VPA 2.0+DCA 1.0联合用药组〔早期:(167±3)%,晚期:(32±4)%〕和VPA 2.0+DCA 4.0联合用药组〔早期:(37±5)%,晚期:(36±5)%〕凋亡率均高于其相应单药组,差异具有统计学意义(P<0.01)。VPA 2.0+DCA 1.0联合用药组和VPA 2.0+DCA 4.0联合用药组hPer3启动子甲基化状态均明显低于其相应单药组。VPA2.0+DCA 1.0联合用药组和VPA 2.0+DCA 4.0联合用药组hPer3 mRNA表达分别为1.75±0.33和3.02±0.36,均明显高于其相应单药组,差异具有显著统计学意义(P<0.01)。VPA 2.0+DCA 1.0联合用药组和VPA 2.0+DCA 4.0联合用药组CD14表达率分别为(16.39±1.68)%和(14.82±0.94)%,均明显高于其相应单药组,差异具有显著统计学意义(P<0.01)。结论 DCA联合VPA能显著加强抗白血病效应,作用机制可能与上调hPer3基因的表达相关。

Synergistic effect of decitabine and valproic acid on human period3 gene expression in leukemia HL-60 cells

OBJECTIVE To investigate the synergistic effect of decitabine(DCA) and valproic acid(VPA) on human period3(hPer3) gene expression in leukemia HL-60 cells.METHODSCells were treated for 48 h by DCA 1.0,4.0 μmol·L-1,VPA 2.0 mmol·L-1,VPA 2.0 mmol·L-1+DCA 1.0 μmol·L-1,and VPA 2.0 mmo·L-1+DCA 4.0 μmol·L-1.Then inhibition rates of cell proliferation were detected by methyl thiazolyl tetrazolium(MTT).The early apoptosis rate was detected by staining with Annexin Ⅴ and PI.The promoter methylation status of hPer3 gene was detected by MS-PCR.The hPer3 mRNA expressions level was detected by real-time PCR.The CD14 expression was detected by flow cytometry.RESULTS Compared with corresponding VPA or DCA single group,the inhibition rate of VPA 2.0+DCA 1.0 group and VPA 2.0+DCA 4.0 was significantly higher(P<0.01),the apoptosis rate of VPA 2.0+DCA 1.0 group(early period:(16.98±2.85)%,late period:(32.44±4.13)%)and VPA 2.0+DCA 4.0(early period:(36.87±4.99)%,late period:(35.61±5.22)%)was significantly higher(P<0.01),the promoter methylation status of hPer3 in VPA 2.0+DCA 1.0 group and VPA 2.0+DCA 4.0 group was significantly higher(P<0.01),the hPer3 mRNA expression in VPA 2.0+DCA 1.0 group(1.75±0.33) and VPA 2.0+DCA 4.0 group(3.02±0.36) was significantly lower(P<0.01),and the CD14 expression in VPA 2.0+DCA 1.0 group((14.82±0.94)%) and VPA 2.0+DCA 4.0 group((16.39±1.68)%) was significantly lower(P<0.01).CONCLUSION VPA enhances the antileukemia activity of DCA possibly by inducing the expression of hPer3 mRNA.