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目的探索Per2在CCAAT增强子结合蛋白α(C/EBPα)参与调控的慢性粒细胞白血病(CML)发生、发展中的作用。方法通过脂质体介导将真核表达载体pGenesil-3-SiPer2和空载体pGenesil-3-HK转染到pEGFP-C/EBPα-K562细胞,利用新霉素筛选稳定干扰Per2的pEGFP-C/EBPα-K562单克隆细胞株。利用RT-PCR和Western blot分别检测转染组、空载组、对照组细胞p53、c-myc、cyclinB1的mRNA及蛋白表达水平的改变。结果成功构建稳定干扰Per2表达的pEGFP-C/EBPα-K562细胞株。与对照组和空载组细胞相比,转染组pGenesil-3-SiPer2-K562细胞的p53mRNA及蛋白表达水平明显降低,差异有统计学意义(P<0.01),而cyclinB1和c-myc基因的mRNA及蛋白表达水平则显著升高,差异有统计学意义(P<0.01)。结论 Per2基因在C/EBPα参与调控的CML发生发展中起重要作用,该通路的发现对于研究CML的发生及调控机制具有重要意义。
Objective To explore the role of Per2 in the development and progression of chronic myelogenous leukemia (CML) which is involved in the regulation of CCAAT enhancer binding protein α (C / EBPα). Methods The eukaryotic expression vector pGenesil-3-SiPer2 and empty vector pGenesil-3-HK were transfected into pEGFP-C / EBPα-K562 cells by liposome. The recombinant plasmid pEGFP-C / EBPα-K562 monoclonal cell line. The mRNA and protein expression levels of p53, c-myc and cyclinB1 were detected by RT-PCR and Western blot in transfected, empty and control groups respectively. Results The pEGFP-C / EBPα-K562 cell line stably interfered with Per2 expression was successfully constructed. The expression of p53 mRNA and protein in transfected pGenesil-3-SiPer2-K562 cells was significantly lower than that in the control and blank control groups (P <0.01), while the expression of cyclinB1 and c-myc mRNA and protein expression levels were significantly increased, the difference was statistically significant (P <0.01). Conclusion Per2 gene plays an important role in the development of CML regulated by C / EBPα. The discovery of this pathway is of great significance for the study of the occurrence and regulation of CML.