多发性骨髓瘤细胞中胰岛素样生长因子结合蛋白7的基因表达及甲基化调控

发布时间：2018-06-04 01:40

本文选题：胰岛素样生长因子结合蛋白 + 多发性骨髓瘤　；参考：《中国实验血液学杂志》2017年03期

【摘要】：目的:研究多发性骨髓瘤细胞株U266中胰岛素样生长因子结合蛋白7(IGFBP7)的表达及甲基转移酶抑制剂5-氮杂-2'-脱氧胞苷(5-aza-dc)对其增殖的影响。方法:体外培养多发性骨髓瘤细胞株U266,并以健康查体者的骨髓单个核细胞(N-BMMNC)作为正常对照,提取RNA,采用实时RT-PCR检测IGFBP7的表达,提取细胞DNA甲基化修饰后采用甲基化特异性PCR(MSP)分析其CpG岛的甲基化状态,不同浓度的甲基转移酶抑制剂5-aza-dc(5、10和20μmol/L)处理U266细胞株,在48 h收集细胞,用实时RT-PCR和蛋白印迹法检测各组IGFBP7 mRNA和蛋白表达,流式细胞术检测药物处理前后各组细胞的细胞凋亡及细胞周期。结果:U266细胞IGFBP7 mRNA表达较正常骨髓细胞降低;MSP检测显示U266 IGFBP7的启动子CpG岛明显甲基化;不同浓度的5-aza-dc处理U266细胞株48 h后,IGFBP7的mRNA表达呈剂量依赖性增高(r=0.952,P0.05),IGFBP7蛋白表达呈剂量依赖性增高(r=0.983,P0.05);随着药物浓度增高,U266在G_0/G_1期细胞逐渐增多(r=0.966),细胞凋亡率逐渐增加(r=0.958)。结论:U266细胞中IGFBP7 mRNA较N-BMMNC低表达,与CpG岛的异常甲基化相关,5-aza-dc可以使U266细胞中IGFBP7mRNA和蛋白表达恢复,并通过阻滞细胞周期和诱导细胞凋亡发挥其对U266细胞的生长抑制作用
[Abstract]:Aim: to investigate the expression of insulin-like growth factor binding protein (IGFBP7) in multiple myeloma cell line U266 and the effect of 5-aza-dc), a methyltransferase inhibitor, on its proliferation. Methods: multiple myeloma cell line U266 was cultured in vitro, and bone marrow mononuclear cells (N-BMMNCs) of healthy examiners were used as normal control. IGFBP7 expression was detected by real-time RT-PCR. DNA methylation modification was used to analyze the methylation status of CpG islands in U266 cells. Different concentrations of methyltransferase inhibitor 5-aza-dccct5 and 20 渭 mol / L were treated to U266 cell lines, and the cells were collected at 48 h. Real-time RT-PCR and Western blot were used to detect the expression of IGFBP7 mRNA and protein in each group. Flow cytometry was used to detect cell apoptosis and cell cycle before and after drug treatment. Results compared with normal bone marrow cells, the IGFBP7 mRNA expression of U266 cells was significantly lower than that of normal bone marrow cells. The promoter CpG island of U266 IGFBP7 was significantly methylated. The expression of IGFBP7 in U266 cells was increased in a dose-dependent manner after 48 h treatment with different concentrations of 5-aza-dc. The expression of IGFBP7 protein in U266 cells was increased in a dose-dependent manner, and the apoptosis rate of U266 cells increased in a dose-dependent manner. Conclusion the expression of IGFBP7 mRNA in U266 cells is lower than that of N-BMMNC, and 5-aza-dc associated with abnormal methylation of CpG island can restore the expression of IGFBP7mRNA and protein in U266 cells, and exert its inhibitory effect on the growth of U266 cells by blocking cell cycle and inducing apoptosis.
【作者单位】：山西医科大学第二医院血液科;
【基金】：山西省卫生计生委科研课题(2015041)
【分类号】：R733.3

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