【摘要】 目的研究孕激素拮抗剂米非司酮对白血病多药耐药细胞 K562/A02的逆转作用及其机制。方法 MTT 法检测米非司酮作用72 h 后 K562/A02细胞增殖及其对阿霉素杀伤敏感性的变化;流式细胞术检测米非司酮作用前后 K562/A02细胞表面 P 糖蛋白的表达和细胞内柔红霉素的浓度;免疫组化法观察米非司酮作用前后 K562/A02细胞凋亡相关蛋白 bcl-2、Bax、caspase-3的表达;RT-PCR 检测米非司酮作用后对 K562/A02细胞内葡萄糖神经酰胺合成酶(GcS)mRNA 表达的影响。结果2.5、5.0和10.0μmol/L 米非司酮不抑制 K562/A02细胞的增殖,但上述浓度的米非司酮作用后K562/A02细胞对阿霉素的敏感性较前分别增强了1.68、4.17和10.71倍。K562/A02细胞表面 P 糖蛋白的表达为(49.03±5.32)%,10 μmol/L 米非司酮作用72 h 后降低到(28.60±2.13)%(P<0.01);K562/A02细胞内柔红霉素的浓度为(61.07±8.61)%,而10 μmol/L米非司酮作用后升高到(92.72±3.48)%(P<0.01)。经10 μmol/L 米非司酮作用后,bcl-2蛋白表达由(56±9)%降低到(37±6)%(P<0.05);Bax 蛋白由(40±5)%升高到(87±10)%(P<0.01);caspase-3蛋白则由(36±7)%升高到(89±6)%(P<0.01)。RT-PCR 结果显示 K562/A02细胞 GcS mRNA 的表达较K562细胞明显升高,10 μmol/L 米非司酮能明显降低 K562/A02细胞内 GcS mRNA 的表达。结论米非司酮可逆转白血病 K562/A02细胞的多药耐药,且具有剂量依赖性。10 μmol/L 米非司酮能明显逆转白血病 K562/A02细胞的多药耐药,其机制与降低 P 糖蛋白的水平,调节凋亡相关蛋白 bcl-2、Bax、caspase-3的表达,降低 GcS mRNA 有关。更多还原

【Abstract】 Objective To study whether progestogen antagnnist mifepristone could reverse multidrug resistance of K562/A02 cells and its mechanisms.Methods MTT was used to study the proliferation of K562/A02 cells and sensitivity of K562/A02 cells to ADM after 72 hours treatment with mifepristone.Flow cytometry was used to assay the expression of P-glycoprotein and the mean fluorescent intensity of intracellular daunorubicin.The expressions of apoptosis related proteins(bcl-2,Bax and caspase-3)were assayed by im- munohistochemistry and the glucosylceramide synthase mRNA expression by RT-PCR before and after mife- pristone treatment.Results MTT assay revealed that 2.5,5.0 and 10 μmol/L mefepristone did not affect the proliferation of K562/A02 cells,but enhanced the sensitivity of K562/A02 cells to ADM,by 1.68-, 4.17-and 10.71-fold increase,respectively.Expression of P-gp in K562/A02 cells was(49.03±5.32)%, and was decreased to(28.60±2.13)%(P<0.01)after 10 μmol/L mifepristone treatment for 72 hours. and intracelluar DNR accumulation in K562/A02 was(61.07±8.61)%,and was increased to(92.72± 3.48)%(P<0.01).After 10 μmol/L mifepristone treatment,the expression of bcl-2 protein was decreased from(56±9)% to(37±6)%(P<0.05),Bax and caspase-3 proteins was increased from(40±5)% to (87±10)%(P<0.01),and from(36±7)% to(89±6)%(P<0.01)respectively.RT-PCR analysis revealed that expression of glucosylceramide synthase mRNA was higher in K562/A02 than in K562 cells, whereas 10 μmol/L mifepristone significantly down-regulated its expression in K562/A02 cells.Conclusion Mifepristone at 10 μmol/L could dose-dependently reverse the multidrug resistance of K562/A02 cells.The possible mechanisms are related with decreasing the expression of P-gp,regulating the expression of apoptosis related proteins and decreasing the expression of glucosylceramide synthase.更多还原