阿糖胞苷对U937细胞作用与端粒酶相关基因表达水平的研究

A study on the gene expression of telomerase subunits and the cell apoptosis and necrosis in U937 cells induced by cytarabine

目的:检测不同浓度、不同时间阿糖胞苷(cytarabine,Ara-C)作用U937细胞后其凋亡、坏死比例及端粒酶各组分基因mRNA水平的变化。方法:采用不同浓度Ara-C作用相同时间、同一浓度Ara-C作用不同时间处理U937细胞。通过流式细胞仪观察其凋亡和坏死细胞比例;RT-PCR方法检测端粒酶各组分基因人端粒酶催化亚基(hTERT)、人端粒酶RNA成分(hTR)和人端粒酶相关蛋白质(hTP1)mRNA水平变化。结果:体外小剂量(0～0.2μg/ml)Ara-C诱导U937细胞12h,细胞凋亡和坏死比例及端粒酶各组分基因在mRNA水平上无明显变化。而2μg/ml和10μg/mlAra-C组的细胞凋亡和坏死比例明显增加;端粒酶hTERTmRNA水平由0.70±0.08分别下调至0.52±0.06和0.40±0.05,而hTR、hTP1mRNA无变化。10μg/ml的Ara-C作用U937细胞后,细胞凋亡比例随时间延长而增加,12h时达最大值(14.35±2.86)%;细胞坏死比率随时间延长而增加,48h坏死比例最高为(52.28±11.90)%;同时hTERT基因mRNA水平随时间延长而逐渐降低,48h达最低值0.21±0.06;hTR、hTP1mRNA水平无改变。结论:Ara-C能下调hTERT基因的mRNA水平,且有浓度和时间依赖性,其下调该基因水平主要与诱导细胞坏死有关,与细胞凋亡关系甚微。

Objective To investigate the changes of mRNA expression of telomerase subunits in U937 cells induced by cytarabine（Ara-C）, and to observe the cell apoptosis ＆ necrosis. Methods U937 cells cultured were treated with various Ara-C concentration and various treatment duration, the rates of the apoptotic cells and necrotic cells were detected by flow cytometry and the expression of telomerase subunits mRNA was examined by RT-PCR. Results The U937 cells treated with small doses of 0-0.2 μg/ml Ara-C for 12 h in vitro did not influence the transcription of telomerase subunits gene. The rate of apoptotic cells and necrotic cells did not obviously changed. The expression of hTERT was down regulated from 0.70±0.08 to 0.52±0.06 and 0.40±0.05 in the cells treated with 2 μg/ml and 10 μg/ml Ara-C, but hTR and hTP1 expressions had no changes. The rate of apoptotic cells and necrotic cells elevated obviously. With treatment duration, the rate of apoptosis increased in the cells treated with 10 μg/ml Ara-C, and the peak value reached （14.35±2.86）% at the time point of 12 h, The rate of necrosis increased simultaneously with the maximum value of （52.28±11.09）% at the time point of 48 h. With treatment time the mRNA level of hTERT gene decreased with the minimum value of 0.21-±0.06 at the time point of 48 h, but hTR and hTP1 had no similar changes. Conclusions Ara-C can down-regulate the expression of hTERT mRNA in a dose- and time-dependent manner, which may be mainly associated with its inducing potential to cell necrosis rather than cell apoptosis.