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目的观察骨肉瘤细胞MG-63在体外诱导逆转过程中DNA甲基转移酶(DNA methyltransferase,Dnmt)的变化,探讨Dnmt与骨肉瘤恶性逆转的相关性及其作用。方法体外用全反式维甲酸(all trans-retinoic acid,ATRA)对MG-63细胞进行诱导分化,观察分化后瘤细胞形态和功能的变化,RT-PCR法检测细胞Dnmt和增殖核抗原(prolifration cell nuclear antigen,PCNA)mRNA的表达；以流式细胞仪检测药物作用前后细胞周期的改变。结果经诱导剂处理后,细胞的生长明显受到抑制,细胞形态和功能分化成熟,碱性磷酸酶的活性明显升高,Dnmt和PCNA mRNA 的表达随着作用时间的延长均降低,细胞周期阻滞于G1期。结论 MG-63细胞在体外恶性逆转诱导过程中,瘤细胞在形态和功能上向成熟分化,PCNA mRNA表达水平和细胞周期的进程与 Dnmt的活性密切相关,抑制Dnmt基因的表达是ATRA诱导骨肉瘤细胞分化的重要机制之一。
Objective To observe the changes of DNA methyltransferase (Dnmt) in osteosarcoma cells MG-63 in vitro and to explore the relationship between Dnmt and malignant reversal of osteosarcoma and its role. Methods The differentiation of MG-63 cells was induced by all-trans retinoic acid (ATRA) in vitro. The morphological and functional changes of the differentiated MG-63 cells were observed. The expression of Dnmt and prolif- eration cell nuclear antigen (PCNA) mRNA expression was detected by flow cytometry before and after the drug effect on cell cycle changes. Results After induced by the inducer, the growth of cells was obviously inhibited, the morphology and function of cells were well differentiated, the activity of alkaline phosphatase was significantly increased, the expression of Dnmt and PCNA mRNA decreased with the prolongation of action time, and cell cycle arrest In G1 phase. Conclusion The MG-63 cells differentiate into mature and differentiated cells in morphology and function during the induction of malignant transformation in vitro. The process of the expression of PCNA mRNA and the cell cycle are closely related to the activity of Dnmt. The inhibition of the expression of Dnmt gene is the result of ATRA-induced osteosarcoma One of the important mechanisms of cell differentiation.