吉西他滨诱导人肺癌细胞株SPC-A-1凋亡及对HMGA1基因表达的影响

Effects of Gemcitabine on Apoptosis and HMGA1 Expression in Human Lung Cancer Cell Line SPC-A-1

[目的]探讨吉西他滨对人肺癌细胞株SPC-A-1增殖、凋亡及HMGA1基因表达的影响。[方法]采用四甲基偶氮唑蓝(MTT)比色法观察吉西他滨对人肺癌细胞株SPC-A-1增殖的抑制效应;利用DNA琼脂糖凝胶电泳法、细胞AO/EB荧光染色法及流式细胞术检测吉西他滨诱导肿瘤细胞凋亡的作用;采用逆转录—多聚酶链反应技术(RT-PCR)观察吉西他滨对人肺癌细胞株SPC-A-1凋亡相关基因HMGA1mRNA的影响。[结果]吉西他滨对人肺癌SPC-A-1细胞株增殖有抑制作用,其抑制效应具有时间和剂量依赖性;0.1μmol/L、1μmol/L和10μmol/L浓度吉西他滨处理细胞72h后,AO/EB荧光染色法计算细胞凋亡百分比分别是18.56%±1.04%、39.45%±0.83%、48.48%±0.85%(P<0.05);细胞经1μmol/L的吉西他滨处理72h后,DNA琼脂糖凝胶电泳法显示有特异性的DNA梯状条带;0.1μmol/L、1μmol/L和10μmol/L吉西他滨处理细胞72h后,凋亡率分别为20.65%±0.83%、38.53%±0.97%、51.48%±1.04%,对照组为5.19%±0.89%(P<0.05);随着吉西他滨浓度的增加,HM-GA1基因表达下调,各组与β-actin条带灰度比值分别为0.856±0.030、0.696±0.007、0.543±0.037,对照组为0.907±0.007(P<0.05)。[结论]吉西他滨可抑制人肺癌SPC-A-1细胞株的生长,促进凋亡,可能的机制和下调HMGA1基因表达有关。

[Purpose ] To explore the effects of gemcitabine on proliferation, apoptosis and HMGA1 expression in human lung cancer cell line SPC-A-1. [Methods ] The inhibitive effect of gemcitabine on proliferation of SPC-A-1 cells was detected by MTT assay; apoptosis-induced, by DNA gel electrophoresis, acridine orange（AO）/ethidium bromide （EB） fluorescent staining and flow eytometry; HMGA1mRNA expression, by retrotranscriptase polymerase chain reaction （RT-PCR）. [Results] Gemcitabine inhibited the proliferation of human lung cancer cell line SPC-A-1 in a dose-, time-dependent manner. The apoptosis rates of SPC-A-1 cells after cultured with 0.1 μmol/L, 1 μmol/L and 10μ mol/L gemcitahine for 72h by AO/EB fluorescent staining were 18.56%±1.04%, 39.45%±0.83%, 48.48%±0.85% respectively（P〈0.05）. The typical DNA ladder of apoptosis appeared on DNA electrophoresis when cultured with 1 μmol/L gemcitabine for 72h. The apoptosis rates of SPC-A-1 cells cultured with 0.1μmol/L,1μmol/L and 10μmol/L gemcitabine for 72h were 20.65%±0.83%, 38.53%±0.97%, 51.48%±1.04% respectively. Those compared to the control group （5.19%±0.89%） were significantly different（P〈0.05）. HMGA1 expression decreased in SPC-A-1 cells with the concentration of gemcitabine increased. The ratio of gray values to β-actin were 0.856±0.030, 0.696±0.007, 0.543±0.037 and the control group was 0.907±0.007 （P〈0.05）. [Conclusion] Gemcitabine may inhibit growth of human lung cancer cell line SPC-A-1 and induce apoptosis. The mechanism may be associated with down-regulation of HMGA1 expression.