Objective: To investigate the regulation of miR-34a on HDAC1 expression and its effect on the apoptosis of acute myeloid leukemia (AML) cells.
Methods: miR-34a mimics, miR-34a inhibitor and miR-34a scramble were transfected into HL-60 cells. The effects of miR-34a expression levels on proliferation and apoptosis of HL-60 cell were detected by CCK8 assay and flow cytometry respectively. The expression of HDAC1 protein was assessed by Western blot after regulating miR-34a expression, the 3'UTR of HDAC1 was cloned and ligated to construct a dual luciferase reporter vector, and then the dual luciferase reporter assay was applied to verify the target of miR-34a, the expression vector pcDNA3.1-HDAC1 was constructed, the interaction of miR-34a and HDAC1 was analyzed by reversion test.
Results: miR-34a over-expression could inhibit the proliferation of HL-60 cells and induce their apoptosis. Bioinformatics analysis indicated that the HDAC1 was a target gene of miR-34a. Western blot indicated that miR-34a overexpression down-regulated the expression of HDAC1. Dual luciferase reporter assay and reversion test showed that miR-34a could act at the 3-UTR of HDAC1 gene to regulate its expression.
Conclusion: miR-34a promotes the apoptosis of HL-60 cells via regulating HDAC1 expression.
题目: miR-34a靶向调控HDAC1基因对急性髓系白血病细胞凋亡的影响.
目的: 探讨急性髓系白血病细胞中miR-34a对HDAC1的调控作用以及对细胞凋亡的影响.
方法: 将miR-34a mimics、miR-34a inhibitor、miR-34a scramble转染到HL-60细胞中，采用CCK8试验和流式细胞术分别检测miR-34a不同表达水平对HL-60细胞增殖和凋亡的影响。应用Western blot检测miR-34a表达改变后HDAC1蛋白表达水平。构建含HDAC1 3′UTR的双荧光素酶报告载体，应用双荧光素酶报告实验验证miR-34a与HDAC1的作用位点。构建不含HDAC1 3′UTR的表达载体，应用回复实验验证miR-34a与HDAC1的相互作用.
结果: miR-34a过表达可导致HL-60细胞的增殖抑制、凋亡率增加。生物信息学分析表明，HDAC1是miR-34a的靶基因；Western blot结果显示，miR-34a高表达可下调HDAC1蛋白表达，双荧光素酶实验和回复实验显示，miR-34a可作用于HDAC1 3′UTR区，调理其表达.
结论: miR-34a促进HL-60细胞凋亡，其作用可能与其对HDAC1表达调控有关.