Objective: To investigate the effects of miR-23a-3p on proliferation, migration and apoptosis on human acute myeloid leukemia (AML) cells by targeting SMC1A. Methods: Microarray analysis was used to screen differentially expressed microRNAs and mRNAs in human AML cells. Real-time fluorescence quantitative PCR (RT-qRCR) was used to detect the expressions of miR-23a-3p and SMCA in human AML cell line U937. TargetScan database was used to analyze the correlation between miR-23a-3p and SMC1A. Double luciferase reporter gene was used to detect the interaction between miR-23a-3p and SMC1A. The effect of miR-23a-3p expression on the proliferation of U937 cells was detected by clonal assay. The migration, apoptosis, cell cycle and caspase-3 activity of U937 cells regulated by miR-23a-3p were detected by cell scratch assay and flow cytometry, respectively. Western blot was used to detect the expressions of Bax and Bcl-2 in U937 cells. Results: Compared with human normal monocyte SC group (1.00), the expression of miR-23a-3p in U937 cells was up-regulated (2.56±0.78) (P<0.01), while the expression of SMC1A was down-regulated (0.48±0.56, P<0.01). miR-23a-3p specifically bond to SMC1A 3'UTR and regulated the expression activity of SMC1A. Overexpression of miR-23a-3p promoted the proliferation and migration of U937 cells and inhibited the apoptosis of U937 cells, while up-regulation of SMC1A inhibited the proliferation and migration of U937 cells and promoted the apoptosis of U937 cells. The percentages of G(0)/G1 phase, G(2)/M phase and S phase cells in the negative control group were (37.48±0.21)%, (16.78±0.18)% and (45.74±0.15)% respectively, and those in the miR-23a-3p mimics group were (19.96±0.11)%, (41.69±0.24)% and (38.24±0.34)%, respectively. The difference was statistically significant (all P<0.05). The proportions of G(0)/G(1) phase, G(2)/M phase and S phase cells in the group of miR-23a-3p mimics+ pcDNA3.1-SMC1A were (36.88±0.21)%, (30.44±0.33)% and (32.88±0.16)%, respectively, without significant difference when compared with those of the miR-23a-3p mimics group (P>0.05). The relative expression levels of Bax and Bcl-2 protein in the negative control group were 0.55±0.45 and 0.31±0.54, respectively. Overexpression of miR-23a-3p inhibited the expression of Bax protein in U937 cells (0.23±0.13, P<0.001), promoted the expression of Bcl-2 protein (0.50±0.23, P<0.01), while SMC1A increased the expression of Bax protein in U937 cells (0.40±0.11, P<0.01), and inhibited the expression of Bcl-2 protein (0.37±0.15). In the negative control group, caspase-3 activity was (25.82±0.89)%. Overexpression of miR-23a-3p inhibited caspase-3 activity in U937 cells (3.64±0.56)%, P<0.01, while up-regulation of SMC1A promoted caspase-3 activity in U937 cells (15.29±0.85)%, P<0.01. Conclusion: miR-23a-3p can inhibit the proliferation and migration and promote apoptosis of human AML cells by targeting SMC1A.
目的： 探讨miR－23a－3p靶向SMC1A调控人急性髓系白血病(AML)细胞的增殖、迁移和凋亡及其作用机制。 方法： 微阵列分析筛选人AML细胞U937中差异表达的微小RNA和信使RNA，实时荧光定量PCR法检测miR－23a－3p和SMC1A在人AML细胞U937中的表达，TargetScan数据库分析miR－23a－3p与SMC1A的相关性，双荧光素酶报告基因检测miR－23a－3p与SMC1A的相互作用，克隆形成实验检测miR－23a－3p的表达对U937细胞增殖能力的影响，细胞划痕实验检测miR－23a－3p的表达对U937细胞迁移能力的影响，流式细胞术检测miR－23a－3p的表达对U937细胞的凋亡能力、凋亡周期和caspase－3活性的影响，Western blot法检测miR－23a－3p的表达对U937细胞中Bax和Bcl－2蛋白表达的影响。 结果： 与人正常单核细胞SC(1.00)比较，U937细胞中miR－23a－3p的表达(2.56±0.78)上调 (P<0.01)，SMC1A的表达(0.48±0.56)下调(P<0.01)；miR－23a－3p与SMC1A 3′UTR特异性结合，调控SMC1A的表达活性。miR－23a－3p过表达促进U937细胞增殖和迁移，抑制U937细胞的凋亡；而SMC1A上调抑制U937细胞的增殖和迁移，促进了U937细胞的凋亡。阴性对照组中G(0)/G(1)期、G(2)/M期和S期细胞比例分别为(37.48±0.21)%、(16.78±0.18)%和(45.74±0.15)%，miR－23a－3p mimics组分别为(19.96±0.11)%、(41.69±0.24)%和 (38.24±0.34)%，差异均有统计学意义(均P<0.05)。miR－23a－3p mimics＋pcDNA3.1－SMC1A组中G(0)/G(1)期、G(2)/M期和S期细胞比例分别为(36.88±0.21)%、(30.44±0.33)%和(32.88±0.16)%，与miR－23a－3p mimics组比较，差异均无统计学意义(均P>0.05)。阴性对照组中Bax和Bcl－2蛋白的相对表达水平分别为0.55±0.45和0.31±0.54，miR－23a－3p过表达抑制U937细胞中Bax蛋白的表达(0.23±0.13，P<0.001)，促进Bcl－2蛋白的表达(0.50±0.23，P<0.01)，而SMC1A上调促进U937细胞中Bax蛋白的表达(0.40±0.11，P<0.01)，抑制Bcl－2蛋白的表达(0.37±0.15)。阴性对照组中caspase－3的活性为(25.82±0.89)%，miR－23a－3p过表达抑制U937细胞中caspase－3的活性[(3.64±0.56)%，P<0.01]，而SMC1A上调促进U937细胞中caspase－3的活性[(15.29±0.85)%，P<0.01]。 结论： miR－23a－3p通过靶向SMC1A抑制人AML细胞的增殖和迁移，促进其凋亡。.
Keywords: Apoptosis; Human acute myeloid leukemia cells; Migration; Proliferation; SMC1A; miR-23a-3p.