Daxx抑制K562细胞向巨核细胞分化

发布时间：2019-03-13 11:46

【摘要】：目的:观察过表达死亡结构域相关蛋白(Daxx)对K562细胞活力和向巨核细胞分化的影响。方法:建立稳定过表达Daxx的K562细胞,荧光显微镜观察、实时荧光定量PCR和Western blot检测Daxx的过表达效果,CCK-8法检测过表达后细胞活力的变化;佛波酯(PMA)诱导K562细胞向巨核细胞系分化,Western blot检测在K562细胞向巨核细胞分化过程中Daxx和p-ERK的表达变化,流式细胞术检测CD41和CD61的表达变化;PMA处理过表达Daxx的K562细胞,NBT还原实验检测细胞分化情况,流式细胞术检测过表达Daxx后CD41和CD61的表达变化,Western blot检测p-ERK的蛋白水平。结果:建立了稳定的过表达Daxx的K562细胞,过表达Daxx抑制K562细胞的活力。PMA诱导K562细胞向巨核细胞分化,CD41和CD61表达水平增高,同时p-ERK的蛋白水平升高,Daxx表达水平下降。过表达Daxx可以抑制K562细胞向巨核细胞分化,CD41和CD61表达降低,同时p-ERK的蛋白水平降低。结论:过表达Daxx可以抑制K562细胞生长及向巨核细胞分化,同时抑制ERK的磷酸化。
[Abstract]:Aim: to observe the effect of overexpression of death domain associated protein (Daxx) on K562 cell viability and differentiation into megakaryocyte. Methods: K562 cells with stable over-expression of Daxx were established. The over-expression of Daxx was detected by real-time fluorescence quantitative PCR and Western blot, and the changes of cell viability after over-expression were detected by CCK-8 method. The differentiation of K562 cells into megakaryocytes was induced by phorbol ester (PMA)., Western blot was used to detect the expression of Daxx and p-ERK during the differentiation of K562 cells into megakaryocytes, and the expression of CD41 and CD61 was detected by flow cytometry. K562 cells overexpressing Daxx were treated with PMA. The differentiation of K562 cells was detected by NBT reduction assay. The expression of CD41 and CD61 after Daxx was detected by flow cytometry. The expression of CD41 and CD61 was detected by, Western blot to detect the protein level of p-ERK. Results: a stable Daxx-overexpressed K562 cell line was established. Overexpression of Daxx inhibited K562 cell viability. PMA induced the differentiation of K562 cells into megakaryocytes, increased the expression of CD41 and CD61, increased the protein level of p-ERK and decreased the expression level of Daxx. Overexpression of Daxx inhibited the differentiation of K562 cells into megakaryocytes, decreased the expression of CD41 and CD61, and decreased the protein level of p-ERK. Conclusion: overexpression of Daxx can inhibit the growth and differentiation of K562 cells to megakaryocytes, and inhibit the phosphorylation of ERK.
【作者单位】：嘉兴学院医学院;
【基金】：“十二五”浙江省高校药理学重点学科资助项目嘉兴市科技局项目(No.2015AY23063) 嘉兴学院重点SRT项目(No.851715065)
【分类号】：R392

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