β-榄香烯对K562细胞周期与细胞凋亡的影响及其机制探讨

目的探讨β-榄香烯(β-ELE)对人慢性髓性白血病细胞株K562细胞周期及细胞凋亡的影响及其机制。方法K562和不同浓度的β-ELE共同培养后,应用流式细胞仪技术和Hoe-chest33258/PI荧光染色法检测β-ELE对K562细胞周期及凋亡的影响,应用RT-PCR技术测定野生型p53活化片段(P21wild-type p53 activated fregmentl,P21WAF1)、Survivin mRNA水平。结果不同浓度β-ELE能使K562细胞阻滞于G1期,G1期细胞百分比增加,S期细胞百分比明显下降,呈剂量、时间依赖性,且与对照组相比差异均有统计学意义(P<0.01)。β-ELE作用于K562细胞24h,48h后,在G0/G1期前,可见明显的亚二倍体(凋亡峰),其凋亡率呈剂量、时间依赖性,与对照组比较有显著意义(P<0.01)。Hoechest33258/PI免疫荧光技术发现β-ELE能使凋亡细胞数目明显增多(P<0.01)。RT-PCR结果显示β-ELE能下调K562细胞Survivin mRNA表达,上调P21WAF1mRNA的表达(P<0.01)。结论β-ELE能够阻止细胞K562细胞从G1期向S期转换。β-ELE可以诱导K562细胞凋亡,呈剂量、时间依赖性。β-ELE导致细胞周期阻滞可能与上调P21WAF1mRNA的表达有关;促进K562细胞凋亡机制可能与下调Survivin mRNA的表达有关。随着药物浓度的增加,P21WAF1mRNA表达增加,而SurvivinmRNA表达下降。

The influence of β-elemene on cell cycle and apoptosis of K562 Cells and the investigation of its mechanism

Objective To investigate the influence of β-ELE on cell cycle and apoptosis of human chronicity myeloid leukemia cell line K562 and related mechanism.Methods K562 cell line was cultured with 10 μg/ml、20 μg/ml、30 μg /ml、40 μg /ml β-ELE for 24 h or 48 h.Flow cytometry and immunofluorescence technic were employed to measure cell cycle and apoptosis of K562 cell.RT-PCR technique was used to examine the P21WAF1 mRNA and Survivin mRNA level.Results After treated with β-ELE the accumulation of K562 cells was in G1 phase.The percent of cells in G1 phase increased and the percent of cells in S phase decreased in a dose-and-time dependence.Compared with the control group,the difference has statistical significance(P<0.01).The obvious hypodiploid peak was found ahead of G1 phase by flow cytometry technique with the concentration of β-ELE for 24 h or 48 h,and the ratio of apoptosis was increased in a dose-and-time way(P<0.01).Immunofluorescence technic showed that the count of apoptotic cells added significantly with the growth of the concentration of β-ELE(P<0.01).P21WAF1 mRNA level increased and survivin mRNA level decreased when treated with β-ELE(P<0.01).Conclusions β-ELE could inhibit K562 cells to switch G1 to S phase and induce the apoptosis of K562 cells and the apoptosis appeared in a dose-and-time dependence.The mechanism of cell cycle blockage maybe related to the up-regulation expressions of P21WAF1 mRNA in K562 cells.The mechanism of apoptosis maybe related to the down-regulation of Survivin mRNA in K562 cells.The mechanism of apoptosis maybe related to the down-regulation of Survivin mRNA in K562 cells.With the increasing of concentration,the expression of P21WAF1 mRNA increased and the expression of Survivin mRNA decreased.