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目的:研究microRNA-101(miR-101)对上皮性卵巢癌细胞SKOV3诱导凋亡的作用,并探讨凋亡发生与细胞内Ca~(2+)浓度的关系。方法:采用阳离子脂质体为载体将miR-101 mimics转染SKOV3细胞,随机分为空白对照组、miR-101 NC组和miR-101转染组;实时定量RT-PCR法检测细胞中miR-101表达量;流式细胞术检测细胞凋亡及细胞内Ca~(2+)浓度;Western blot法检测凋亡相关蛋白caspase-3表达。结果:实时定量RT-PCR检测结果显示,miR-101转染组的miR-101表达(5.302±0.069)较空白对照组(1.038±0.036)和miR-101 NC组(1.126±0.062)分别上调了5.10倍和4.70倍(F=5326.276,P=0.000)。流式细胞术检测结果显示,miR-101转染组的细胞凋亡率、细胞内Ca~(2+)浓度分别为(66.36±13.54)%和(228.94±19.51)nmol/L,较空白对照组[(40.60±5.97)%,(62.91±7.73)nmol/L]和miR-101 NC组[(41.54±3.89)%,(59.09±10.66)nmol/L]明显增加,差异有统计学意义(P=0.019);Western blot结果显示,miR-101转染组的细胞凋亡相关蛋白caspase-3表达(1.325±0.052)较空白对照组(0.817±0.073)和miR-101 NC组(0.820±0.062)增加,差异有统计学意义(P=0.000)。结论:miR-101过表达具有诱导上皮性卵巢癌细胞SKOV3凋亡的作用,其诱导凋亡与细胞内Ca~(2+)浓度增加有关。
OBJECTIVE: To study the effect of miR-101 on apoptosis induced by epithelial ovarian cancer cell line SKOV3 and to investigate the relationship between apoptosis and intracellular Ca2 + concentration. Methods: SKOV3 cells were transfected with miR-101 mimics using cationic liposome as a vector and randomly divided into blank control group, miR-101 NC group and miR-101 transfected group. Real-time quantitative RT-PCR was used to detect the expression of miR- The expression of apoptosis-related protein caspase-3 was detected by Western blot. The expression of caspase-3 was detected by flow cytometry. Results: Real-time quantitative RT-PCR results showed that miR-101 expression in miR-101 transfected group was significantly higher than that in control group (1.038 ± 0.036) and miR-101 NC group (1.126 ± 0.062) 5.10-fold and 4.70-fold (F = 5326.276, P = 0.000). The results of flow cytometry showed that the apoptosis rate and the intracellular Ca2 + concentration in the miR-101 transfected group were (66.36 ± 13.54)% and (228.94 ± 19.51) nmol / L, respectively. Compared with the blank control (40.60 ± 5.97)%, (62.91 ± 7.73) nmol / L] and miR-101 NC group (41.54 ± 3.89% vs 59.09 ± 10.66 nmol / L, respectively) P = 0.019) .Western blot analysis showed that the expression of caspase-3 in the miR-101 transfected group was significantly higher than that in the blank control group (0.817 ± 0.073) and the miR-101 NC group (0.820 ± 0.062) ) Increased, the difference was statistically significant (P = 0.000). Conclusion: Overexpression of miR-101 can induce the apoptosis of epithelial ovarian cancer cell line SKOV3. The induction of apoptosis is associated with the increase of intracellular Ca 2+ concentration.