毛蕊异黄酮对宫颈癌HeLa细胞增殖及凋亡的影响

Effects of calycosin on proliferation and apoptosis of HeLa cells of cervical cancer

  • 摘要:
    目的 探讨毛蕊异黄酮对宫颈癌HeLa细胞增殖及凋亡的影响及其可能作用机制。
    方法 体外培养人宫颈癌细胞HeLa, 设为对照组(未经毛蕊异黄酮处理)、毛蕊异黄酮-低组(25 μmol/L)、毛蕊异黄酮-中组(50 μmol/L)、毛蕊异黄酮-高组(100 μmol/L); 经100 μmol/L毛蕊异黄酮培养24 h, 细胞分别设为微小RNA(miR)-NC组、miR-4766-5p组、毛蕊异黄酮+anti-miR-NC组以及毛蕊异黄酮+anti-miR-4766-5p组。采用CCK-8法、平板克隆形成实验与流式细胞术分别检测细胞增殖、克隆形成及凋亡; 采用实时荧光定量聚合酶链反应(qRT-PCR) 和蛋白质免疫印迹(Western blot)分别检测miR-4766-5p和凋亡相关蛋白表达量。
    结果 与对照组比较, 毛蕊异黄酮-低组、毛蕊异黄酮-中组、毛蕊异黄酮-高组的细胞增殖受到抑制,凋亡蛋白含量、miR-4766-5p表达量升高,且呈剂量依赖性,差异有统计学意义(P < 0.05)。与癌旁组织比较,宫颈癌组织中miR-4766-5p的表达量降低,差异有统计学意义(P < 0.05)。与miR-NC组比较, miR-4766-5p组细胞增殖能力降低,凋亡蛋白含量升高,差异有统计学意义(P < 0.05)。毛蕊异黄酮+anti-miR-4766-5p组的细胞增殖抑制率、凋亡率和cleaved-caspase3、cleaved-caspase9蛋白水平低于毛蕊异黄酮+anti-miR-NC组,细胞克隆形成能力高于毛蕊异黄酮+anti-miR-NC组,差异有统计学意义(P < 0.05)。
    结论 毛蕊异黄酮可通过上调miR-4766-5p表达抑制宫颈癌细胞增殖,诱导细胞凋亡。

     

    Abstract:
    Objective To investigate the effect of calycosin on proliferation and apoptosis of HeLa cells of cervical cancer and its possible mechanism.
    Methods Human cervical cancer HeLa cells were cultured in vitro and divided into control group (without treatment with calycosin), calycosin-low group (25 μmol/L), calycosin-medium group (50 μmol/L) and calycosin-high group (100 μmol/L); the cells were cultured with 100 μmol/L of calycosin for 24 h, and were divided into microRNA (miR)-NC group, miR-4766-5p group, calycosin+anti-miR-NC group, and calycosin+anti-miR-4766-5p group, respectively. Cell proliferation, clonal formation and apoptosis were detected by CCK-8 assay, plate cloning assay and flow cytometry; real-time quantitative fluorescent polymerase chain reaction (qRT-PCR) and Western blot were used to detect the expression levels of miR-4766-5p and apoptosis-related proteins, respectively.
    Results Compared with the control group, cell proliferation was significantly inhibited, apoptosis-protein content and miR-4766-5p expression were significantly increased in a dose-dependent manner in the calycosin-low group, calycosin-medium group and calycosin-high group (P < 0.05). The expression level of miR-4766-5p in cervical cancer tissues was significantly decreased compared with paracancer tissues (P < 0.05). Compared with the miR-NC group, the cell proliferation capacity of the miR-4766-5p group was significantly decreased, and the apoptotic protein content was significantly increased (P < 0.05). The cell proliferation inhibition rate, apoptosis rate and cleaved-caspase3 and cleaved-caspase9 protein levels in the calycosin+anti-miR-4766-5p group were significantly lower than those in the calycosin+anti-miR-NC group, the increase of cell clonogenesis ability was significantly higher than that of the calycosin+anti-miR-NC group (P < 0.05).
    Conclusion Calycosin can inhibit the proliferation of cervical cancer cells and induce cell apoptosis by up-regulating the expression of miR-4766-5p.

     

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