Tal1促进急性T淋巴白血病Jurkat细胞的增殖

Tal1 promotes proliferation of acute lymphoblastic leukemia Jurkat cells in vitro

目的探讨Tal1对T-ALL细胞增殖及其机制的影响。方法用Tal1慢病毒感染T-ALL细胞株Jurkat,建立稳定的Tal1敲降细胞株(Jurkat-si Tal1)和Tal1过表达细胞株(Jurkat-T1),及si RNA阴性对照细胞(Jurkat-mock1)和过表达阴性对照细胞(Jurkatmock2)。CCK-8检测细胞生长能力;流式细胞术检测细胞周期;Real-time RT-PCR和Western blot检测周期蛋白依赖性激酶抑制因子2(CDKN2A)、周期蛋白依赖性激酶抑制因子1(CDKN2B)m RNA和蛋白表达。结果成功建立Jurkat稳定转染细胞株。CCK8结果表明细胞Jurkat-T1与Jurkat-mock2相比,细胞生长更快,而Jurkat-si Tal1与Jurkat-mock1相比,细胞生长明显减缓。流式细胞术检测细胞周期发现,Jurkat-si Tal1与Jurkat-mock1相比G0/G1期增加,S期减少;而Jurkat-T1与Jurkat-mock2相比,G0/G1期减少,S期增加。Real-time RT-PCR和Western blot结果显示Tal1抑制Jurkat细胞内CDKN2A、CDKN2B的m RNA和蛋白表达。结论 Tal1可以促进T淋巴白血病细胞Jurkat增殖;促进Jurkat细胞由G0/G1期向S期转换,其可能通过Tal1抑制G0/G1和S期负调控蛋白CDKN2A、CDKN2B的表达。

Objective To investigate the role of Tal1 gene, which is aberrantly expressed in 40%- 60% of patients with T lymphocytic leukemia（T-ALL）, in the proliferation of T-ALL cells. Methods We established stable Jurkat-si Tal1 and Jurkat-T1 cell lines by trasnfecting T-ALL Jurkat cells with lentiviral vectors to knock-down or overexpress Tal1. Jurkat cells transfected with negative control si RNAs for Tal1 knock- down（Jurkat- mock1） and over- expression（Jurkat- mock2） served as the control cells. The proliferation of the cells lines was assessed using CCK- 8 assay, and the cell cycle distribution was determined by flow cytometry. The m RNA and protein expressions of cyclin-dependent kinase inhibitor 2（CDKN2A） and cyclin-dependent kinase inhibitor 1（CDKN2B） were measured by real- time RT- PCR and Western blotting, respectively. Results Jurkat- T1 cells showed more active proliferation in vitro than Jurkat-mock2 cells, while Jurkat-si Tal1 cells showed slower growth than Jurkatmock1 cells. In Jurkat-T1 cells, G0/G1 phase cells were decreased and S phase cells increased compared with Jurkat-mock2 cells,and Jurkat-si Tal1 cells showed increased G0/G1 phase cells and decreased S phase cells compared with Jurkat-mock1 cells. Realtime RT- PCR and Western blotting showed that Tal1 inhibited the cellular expression of CDKN2 A and CDKN2 B at both m RNA and protein levels. Conclusion Tal1 promotes the growth and the transition from G0/G1 phase to S phase in T-ALL cells Jurkat by inhibiting the expressions of G0/G1 and S phase negative regulatory proteins CDKN2 A and CDKN2 B.