β-榄香烯对K562细胞周期与细胞凋亡的影响及其机制探讨

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

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。

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 G, 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 apoptosisof 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 P21WAFI 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 exoression of Survivin mRNA decrea.~d.