[目的]探讨miR-152靶向TCEAL5对小鼠垂体瘤细胞AtT-20增殖、凋亡和侵袭的影响.[方法]小鼠垂体瘤AtT-20细胞培养转染后48 h进行实验,实验分5组:mimic NC组、miR-152 mimic组、si-NC组,si-TCEAL5组.用双荧光素酶报告基因检测miR-152与TCEAL5的靶基因关系,MTT法检测各组细胞增殖能力,流式细胞术检测各组细胞凋亡率,Transwell法检测细胞侵袭数目,蛋白免疫印迹法检测各组细胞TCEAL5蛋白表达.[结果]miR-152对TCEAL5有潜在的结合位点,转染miR-152 mimic显著降低了 AtT-20细胞裂解液中荧光素酶的相对活性(1.00±0.31vs0.62±0.11),表明 miR-152 与 TCEAL5 mRNA 存在靶向调节作用.miR-152 mimic 组、si-TCEAL5 组72 h AtT-20 细胞光密度值均显著低于 mimic NC 组和 si-NC 组(1.32±0.09 vs 1.03±0.06;1.22±0.15 vs 0.94±0.01;P＜0.05).转染48h后,miR-152 mimic组、si-TCEAL5组AtT-20细胞凋亡率分别显著高于mimic NC组、si-NC组(3.81±0.23 vs 33.25±6.02;4.16±0.53 vs 25.27±3.61;P＜0.05).与 mimic NC 组比较,miR-152 mimic 组 AtT-20 细胞的侵袭能力显著降低(83.22±10.23 vs 43.35±9.02;P＜0.05);与 si-NC 组比较,si-TCEAL5 组 AtT-20细胞的侵袭能力显著降低(85.16±7.53 vs 55.67±5.65,P＜0.05).与 mimic NC 组比较,miR-152mimic 组 AtT-20细胞 TCEAL5 蛋白表达水平显著降低(0.51±0.03 vs 0.25±0.02,P＜0.05);与 si-NC 组比较,si-TCEAL5 组 AtT-20细胞TCEAL5蛋白表达水平显著降低(0.55±0.03 vs 0.24±0.01,P＜0.05)o[结论]miR-152通过与TCEAL5靶向结合,进而抑制垂体瘤细胞AtT-20增殖、侵袭并促进其凋亡.
Effects of miR-152 targeting TCEAL5 on mouse pituitary tumor cells AtT-20
[Objective]To explore the effect of miR-152 targeting TCEAL5 on proliferation,apoptosis and invasion of mouse pituitary tumor cells AtT-20.[Method]Mouse pituitary tumor AtT-20 cells were cultured and transfected 48 h after transfec-tion.The experiments were carried out in five groups:mimic NC group,miR-152 mimic group,si-NC group,si-TCEAL5 group,and miR-152 mimic+si-TCEAL5 group.Dual luciferase reporter gene was used to detect the relationship between miR-152 and TCEAL5 target genes,MTT was used to detect cell proliferation capacity in each group,and flow cytometry was used to detect cell apoptosis rate in each group.The number of cell invasion was detected by Transwell method,and the protein expression of TCEAL5 was detected by Western Blot.[Result]miR-152 has a potential binding site for TCEAL5.Transfection of miR-152 mimic significantly decreased the relative activity of luciferase in AtT-20 cell lysate(1.00±0.31 vs 0.62±0.11),indicating that miR-152 and TCEAL5 mRNA There was a target-regulating effect.miR-152 mimic group and si-TCEAL5 group 72h AtT-20 cell optical density values were significantly lower than those of mimic NC group and si-NC group(1.32±0.09 vs 1.03±0.06;1.22±0.15 vs 0.94±0.01;P＜0.05).After 48 hours of transfection,the apoptosis rate of AtT-20 cells in miR-152 mimic group and si-TCEAL5 group was significantly higher than that in mimic NC group and si-NC group,respectively(3.81±0.23 vs 33.25±6.02;4.16±0.53 vs 25.27±3.61;P＜0.05).The invasive ability of AtT-20 cells was significantly lower in the miR-152 mimic group compared with the mimic NC group(83.22±10.23 vs 43.35±9.02;P＜0.05);the invasive ability of AtT-20 cells in the si-TCEAL5 group was significantly reduced compared with the si-NC group(85.16±7.53 vs 55.67±5.65;P＜0.05).Compared with the mimic NC group,the protein expression level of AtT-20 cells in the miR-152 mimic group was significantly lower(0.51±0.03 vs 0.25±0.02;P＜0.05);compared with the si-NC group,the TCEAL5 protein expression level was significantly decreased in the si-TCEAL5 group(0.55±0.03 vs 0.24±0.01;P＜0.05).[Conclusion]By targeting to TCEAL5,miR-152 inhibited the proliferation and invasion of pituitary tumor cells AtT-20 and promoted its apoptosis.
NETL
NSTL
万方数据
