Student Research Committee, Tabriz University of Medical Sciences, Tabriz, Iran.
Department of Immunology, Division of Hematology, Faculty of Medicine, Tabriz University of Medical Sciences, Tabriz, Iran.
Asian Pac J Cancer Prev. 2021 Nov 1;22(11):3723-3734. doi: 10.31557/APJCP.2021.22.11.3723.
DNA methylation is a reversible biochemical process determinant of gene expression that is frequently observed in acute lymphoblastic leukemia (ALL). This is believed to arise from aberrant DNA methyltransferase activity establishing abnormal levels of DNA methylation in tumor cells. DNA methyltransferase inhibitor, 5-azacytidine (5-AZA), is a clinically used epigenetic drug which induces promoter demethylation and gene re-expression in human cancers. In this study, we investigated the cytotoxicity of on MOLT4 and Jurkat leukemic cell line in vitro and characterized the underlying molecular mechanisms of cell death and motility. MOLT4 and Jurkat cells were treated with 5-AZA for 12, 24 and 48 hours. The effect of the 5-AZA treatment on cell viability (MTT assay), apoptosis (annexin V/PI staining), microRNA (miRNA) and mRNA expression (real-time PCR) was measured. The results showed that 5-AZA could induce MOLT4 and Jurkat apoptotic cell death in vitro in a time-dependent manner and probably via apoptotic mechanisms. We found that treatment with 5-AZA could increase the expression of epigenetically silenced miRNAs, miR-34a, miR-34b and miR-124-1 in treated cells. In addition, mRNA analyses demonstrated that MOLT4 and jurkat cells, expressed p53 gene more than 10-fold higher compared with untreated cells in three independent experiments while the cells suppressed the expression of a subset of functionally related genes including MYC, BCL2, APEX, SIRT1, SNAIL1 and vimentin to some extent, following 5-AZA treatment. We found that a miRNAs expression level in treated cell lines was closely correlated to the expression of their target genes. Together, these findings suggest that 5-AZA may affect the viability of MOLT4 and jurkat cells, at least in part, by regulating the transcription of genes that are associated with cellular apoptotic response.
DNA 甲基化是一种决定基因表达的可逆生化过程，在急性淋巴细胞白血病 (ALL) 中经常观察到。据信，这是由于异常的 DNA 甲基转移酶活性导致肿瘤细胞中异常水平的 DNA 甲基化。DNA 甲基转移酶抑制剂 5-氮杂胞苷 (5-AZA) 是一种临床应用的表观遗传药物，可诱导人类癌症中的启动子去甲基化和基因再表达。在这项研究中，我们研究了 5-AZA 在体外对 MOLT4 和 Jurkat 白血病细胞系的细胞毒性，并对细胞死亡和运动的潜在分子机制进行了表征。将 MOLT4 和 Jurkat 细胞用 5-AZA 处理 12、24 和 48 小时。用 MTT 测定法测定 5-AZA 处理对细胞活力的影响、凋亡（ Annexin V/PI 染色）、微小 RNA（miRNA）和 mRNA 表达（实时 PCR）。结果表明，5-AZA 可以以时间依赖的方式诱导 MOLT4 和 Jurkat 细胞体外凋亡性细胞死亡，可能通过凋亡机制。我们发现，用 5-AZA 处理可增加经处理细胞中表观遗传沉默的 miRNA、miR-34a、miR-34b 和 miR-124-1 的表达。此外，mRNA 分析表明，与未处理细胞相比，MOLT4 和 jurkat 细胞在三个独立实验中 p53 基因的表达高出 10 倍以上，而细胞在一定程度上抑制了包括 MYC、BCL2、APEX、SIRT1、SNAIL1 和波形蛋白在内的一组功能相关基因的表达，在 5-AZA 处理后。我们发现，经处理的细胞系中 miRNA 的表达水平与靶基因的表达密切相关。总之，这些发现表明，5-AZA 可能通过调节与细胞凋亡反应相关的基因的转录，至少在一定程度上影响 MOLT4 和 jurkat 细胞的活力。
