miRNA-19对葡萄膜黑色素瘤细胞增殖、凋亡、迁移和侵袭的影响及其机制研究

目的 探讨miR-19在葡萄膜黑色素瘤(UM)细胞中的表达及其对UM细胞增殖、凋亡、迁移和侵袭的影响及其作用机制.方法 qRT-PCR检测正常视网膜上皮细胞系ARPE-19和UM细胞系 SP6.5、M23中 miR-19的表达.将 M23细胞分为 miR-19 inhibitor 组(转染 miR-19-inhibit...

Effects of miRNA-19 on proliferation,apoptosis, migration and invasion of uveal melanoma cells and its mechanisms

Objective　To investigate the expression of miR-19 in uveal melanoma (UM) and its effects on cell proliferation, apoptosis, migration and invasion and its mechanisms.Methods　Real-time quantitative polymerase chain reaction (qRT-PCR) was used to detect the expression of miR-19 in normal retinal pigment epithelial cell lines ARPE-19 and UM cell lines SP6.5 and M23. The M23 cell line was divided into miR-19 inhibitor group, in which the cells were transfected with miR-19 inhibitor, and miR-19 NC group, in which the cells were transfected with scramble. MTT assay was used to measure cell proliferation. Flow cytometry was used to measure apoptosis. Cell scratch test was used to measure cell migration ability. Transwell assay was used to measure cell invasion. Western blot was used to measure the expression of epithelial growth factor receptor (EGFR)/serine/threonineproteinkinase(AKT) /mammalian target of rapamycin (mTOR) signaling pathway protein.Results　The expression level of miR-19 in normal uveal epithelial cell line ARPE-19 was 1.35±0.13, and 8.35±0.73 and 9.35±0.43 in uveal melanoma cell lines SP6.5 and M23, respectively. The expression level of miR-19 in SP6.5 and M23 was significantly higher than that in ARPE-19 (both P<0.01). After transfection, the expression level of miR-19 in M23 cells of the miR-19 inhibitor group was lower than that in miR-19 NC group (1.05±0.33 vs 8.05±0.64, P<0.01). The results of MTT showed that after 24 hours of transfection, the optical density value was not statistically difference between miR-19 inhibitor group and miR-19 NC group (P>0.05). After transfection for 48 hours, 72 hours, 96 hours, the optical density value of miR-19 inhibitor group was significantly lower than miR-19 NC group (all P<0.05). The apoptosis rate of miR-19 inhibitor group was higher than that of miR-19 NC group ［(15.34±2.35)% vs (8.23±0.72)%, P<0.05］. The scratch healing rate in the miR-19 inhibitor group was lower than that in the miR-19 NC group ［(23.7±2.1)% vs (68.9±5.1)%, P<0.05］. The number of invasive cells in the miR-19 inhibitor group was less than that in the miR-19 NC group (45.1±3.9 vs 115.3±8.9, P<0.05). The expression levels of EGFR protein, AKT protein, and mTOR protein in the miR-19 inhibitor group were lower than those in the miR-19 NC group (0.43±0.03 vs 1.02±0.02, 0.52±0.04 vs 1.12±0.05, and 0.63±0.05 vs 1.41±0.06, respectively, all P<0.05).Conclusion　Knockdown of miR-19 expression can inhibit the proliferation, migration and invasion and promote apoptosis in UM cells. The mechanism may be related to the inhibition of EGFR/AKT/mTOR signaling pathway.