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目的探讨Survivin基因在白血病糖皮质激素(GC)敏感型(CEM)和GC耐药型(Jurkat)细胞株中的表达差异及其在GC耐药中的临床意义。方法实验分为未暴露组和暴露组,并设阴性对照及空白对照。应用四甲基偶氮唑蓝(MTT)法检测细胞增殖情况,流式细胞术分析细胞凋亡率,反转录PCR(RT-PCR)检测Survivin基因mRNA水平,Western blot法分析蛋白水平。结果 1.MTT结果显示:地塞米松磷酸钠(DEX)以剂量、时间依赖方式抑制CEM和Jurkat细胞株增殖。CEM和Jurkat的24h抑制细胞增殖50%的药物浓度(IC50)分别为442μmol.L-1和13μmol.L-1,其差异有统计学意义(P<0.01);CEM和Jurkat的48hIC50分别为273μmol.L-1和3μmol.L-1,其差异有统计学意义(P<0.01)。DEX对Jurkat的抑制增殖效应远较CEM差。2.流式细胞仪分析结果证实:DEX对Jurkat的诱导凋亡作用显著低于CEM(P<0.01)。3.mRNA和蛋白水平结果均显示:未暴露组Jurkat细胞株Sur-vivin表达水平较CEM显著升高(P<0.05)。暴露组CEM细胞株Survivin表达变化较Jurkat细胞株显著(P<0.01)。结论 Sur-vivin基因在GC敏感型和GC耐药型白血病细胞株中的表达差异可能与细胞株对GC的敏感性相关。其在GC耐药型白血病细胞株中的持续高表达可能是GC耐药发生的机制之一。
Objective To investigate the difference of Survivin gene expression in GC-sensitive (CEM) and GC-resistant (Jurkat) cell lines and its clinical significance in GC drug resistance. Methods The experiment was divided into unexposed group and exposed group, with negative control and blank control. The cell proliferation was detected by MTT assay. The apoptosis rate was analyzed by flow cytometry. The mRNA level of Survivin was detected by reverse transcription-polymerase chain reaction (RT-PCR). The protein level was analyzed by Western blot. Results 1. MTT results showed that dexamethasone sodium phosphate (DEX) inhibited the proliferation of CEM and Jurkat cell lines in a dose-and time-dependent manner. The drug concentration (IC50) of CEM and Jurkat for 50% inhibition of cell proliferation for 24h was 442μmol.L-1 and 13μmol.L-1, respectively, with significant difference (P <0.01); 48hIC50 of CEM and Jurkat were 273μmol .L-1 and 3μmol.L-1, the difference was statistically significant (P <0.01). The inhibitory effect of DEX on Jurkat proliferation is far worse than that of CEM. Flow cytometry analysis confirmed that the apoptosis induced by DEX on Jurkat was significantly lower than that of CEM (P <0.01). The results of mRNA and protein levels showed that Survivin expression in Jurkat cells was significantly higher than that in CEM (P <0.05). Survivin expression in exposed CEM cell line was significantly higher than that in Jurkat cell line (P <0.01). Conclusion The difference of Sur-vivin gene expression in GC-sensitive and GC-resistant leukemia cell lines may be related to the sensitivity of cell lines to GC. Its continued high expression in GC-resistant leukemia cell lines may be one of the mechanisms of GC resistance.