iR-17在非小细胞肺癌放射抗拒细胞中的表达及作用研究

目的 建立肺癌放射抗拒细胞系H1975R,并探究miR-17在其放射抗拒中的作用。方法 通过梯度剂量连续照射,体外建立肺癌放射抗拒细胞系H1975R。采用克隆形成实验对H1975R和H1975细胞的放射敏感性进行验证。利用RT-PCR检测亲本细胞和放射抗拒细胞的miR-17的表达水平,并通过上调或下调miR-17表达,比较其放射敏感性变化,应用CCK-8法检测细胞增殖,流式细胞术检测细胞凋亡和周期分布。结果 H1975R的放射生物学参数D₀、D_q和SF₂均比亲本H1975大(P＜0.05),显示更强的放射抗拒性。从RT-PCR结果可知,miR-17在H1975R的表达量是H1975的2.67倍(P＜0.01)。上调H1975细胞的miR-17表达后,其放射敏感性降低,而细胞增殖和抗凋亡能力均增强,G₂/M期细胞比例增多(P＜0.05)。下调H1975R细胞的miR-17表达后,其放射敏感性升高,而细胞增殖和抗凋亡能力均降低,G₂/M期细胞比例减少(P＜0.05)。结论 miR-17参与非小细胞肺癌放射抗拒的调控,其可能成为治疗放射抗拒的新靶点。

The expression and role of miR-17 in radioresistant cells of non-small cell lung cancer

Objective The aims of this study were to establish a radioresistant human lung cancer cell line H1975R,and to explore the role of miR-17 in the formation of radioresistance.Methods The lung cancer radioresistant cell line H1975R was established in vitro by continuous irradiation with gradient dose.The radiosensitivity of H1975R and H1975 cells was verified by colony formation assay.We determined the expression levels of miR-17 in parental cells and radioresistant cells by quantitative RT-PCR and The expression of miR-17 was up-regulated or down-regulated,and the changes in radiosensitivity were compared.Cell proliferation was detected by CCK-8 assay.Flow cytometry was performed to determine apoptosis and cycle distribution.Results The radiobiological parameters(D₀,N,D_q and SF₂)of H1975R cells were greater than those in the H1975 cells(P＜0.05),indicating a stronger radioresistance.From the Results of RT-PCR,the expression level of miR-17 in H1975R cells was 2.67 times than that in H1975 cells(P＜0.01).After up-regulating expression of miR-17 in H1975 cells,its radiosensitivity was reduced,but the proliferation,anti-apoptosis and the proportion of cells in the G₂/M phase increased(P＜0.05).Similarly,after down-regulating expression of miR-17 in H1975R cells,the radiosensitivity increased,while the cell proliferation and anti-apoptosis ability decreased,and the proportion of cells in the G₂/M phase decreased(P＜0.05).Conclusion miR-17 is involved in the regulation of radioresistance in non-small cell lung cancer,which may become a new target for the treatment of radioresistance.