circSKA3/微小RNA-532-5p对卵巢癌细胞增殖、迁移及凋亡的影响

Impact of circSKA3/microRNA-532-5p on proliferation, migration and apoptosis of ovarian cancer cells

  • 摘要:
    目的  探讨circSKA3对卵巢癌细胞恶性行为的影响及其可能作用机制。
    方法  体外培养人卵巢癌细胞SK-OV-3, 随机分为si-NC组、si-circSKA3组、微小RNA(miR)-NC组、miR-532-5p组、si-circSKA3+anti-miR-NC组和si-circSKA3+anti-miR-532-5p组。采用实时荧光定量聚合酶链反应(qRT-PCR)检测circSKA3、miR-532-5p的表达量。采用CCK-8法、平板克隆形成实验、流式细胞术、Transwell实验分别检测细胞增殖、凋亡及迁移。采用双荧光素酶报告实验检测circSKA3与miR-532-5p的靶向关系。采用蛋白免疫印迹(Western blot)检测Cleaved-caspase3蛋白表达量。
    结果  卵巢癌组织中的circSKA3表达水平高于癌旁正常卵巢组织, 差异有统计学意义(P < 0.05)。卵巢癌组织中的miR-532-5p表达低于癌旁正常卵巢组织,差异有统计学意义(P < 0.05)。与si-NC组比较, si-circSKA3组的circSKA3表达减少, miR-532-5p表达增加,细胞增殖抑制率、细胞凋亡率升高,细胞克隆形成数和迁移细胞数减少,且Cleaved-caspase3蛋白表达升高,差异有统计学意义(P < 0.05)。circSKA3能够靶向结合miR-532-5p。miR-532-5p模拟物转染后,细胞增殖、细胞迁移抑制,细胞增殖抑制率、细胞凋亡率以及Cleaved-caspase3蛋白水平升高,差异有统计学意义(P < 0.05)。与单独转染circSKA3小干扰RNA比较,同时转染circSKA3小干扰RNA和miR-532-5p抑制剂能够促进细胞增殖和迁移,抑制细胞凋亡率,降低Cleaved-caspase3蛋白表达,差异有统计学意义(P < 0.05)。
    结论  沉默circSKA3可上调miR-532-5p表达,进而抑制卵巢癌细胞的恶性发展。

     

    Abstract:
    Objective  To investigate the effect of circSKA3 on the malignant behaviors of ovarian cancer cells and its potential mechanism.
    Methods  Human ovarian cancer SK-OV-3 cells were cultured in vitro and randomly divided into si-NC group, si-circSKA3 group, miR-NC group, miR-532-5p group and si-circSKA3+anti-miR-NC and si-circSKA3+anti-miR-532-5p group. The expression levels of circSKA3 and miR-532-5p were detected by quantitative real-time polymerase chain reaction (qRT-PCR). Cell proliferation, apoptosis and migration were detected by CCK-8 assay, plate clone formation assay, flow cytometry and Transwell assay. The targeting relationship between circSKA3 and miR-532-5p was evaluated using dual-luciferase reporter assays. Protein expression of Cleaved-caspase3 was measured by Western blotting.
    Results  The expression level of circSKA3 in ovarian cancer tissues was significantly higher than that in adjacent normal ovarian tissues (P < 0.05). The expression of miR-532-5p in the ovarian cancer tissues was significantly lower than that in the adjacent normal ovarian tissues (P < 0.05). Compared with the si-NC group, the si-circSKA3 group showed significantly reduced circSKA3 expression, increased miR-532-5p expression, elevated cell proliferation inhibition rates and apoptosis rates, decreased colony formation numbers and migrating cell counts, and significantly higher Cleaved-caspase3 protein expression (P < 0.05). CircSKA3 was found to target miR-532-5p. After transfection with miR-532-5p mimictors, cell proliferation and cell migration were inhibited, and the inhibition rate of cell proliferation, the apoptosis rate, and the protein level of Cleaved-caspase3 were significantly increased (P < 0.05). Compared with transfection with circSKA3 small interfering RNA alone, co-transfection with circSKA3 small interfering RNA and miR-532-5p inhibitor promoted cell proliferation and migration, inhibited apoptosis rates, and reduced Cleaved-caspase3 protein expression (P < 0.05).
    Conclusion  Silencing circSKA3 upregulates miR-532-5p expression, thereby inhibiting the malignant development of ovarian cancer cells.

     

/

返回文章
返回