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本研究主要探讨低氧微环境对慢性髓系白血病细胞系K562分化的影响及钠氢交换蛋白1(NHE1)在该过程中的作用。利用低氧模拟物CoCl2或于低氧(2%O2、5%CO2和93%N2)环境中培养K562细胞,应用激光共聚焦显微镜测定细胞内pH值(intracellular pH,pHi),瑞氏染色观察细胞形态,实时定量RT-PCR检测基因表达,Western blot技术检测MAPK磷酸化水平的改变。结果表明:与对照组相比,低氧培养的K562细胞表现出成熟相关的形态学改变,并伴随CCAAT/增强子结合蛋白(C/EBPα)的mRNA水平上调;特异性NHE1抑制剂Cariporide预处理能够促进低氧诱导的K562细胞分化,Cariporide处理后K562细胞的P38和ERK5蛋白激酶磷酸化水平显著增强。结论 :低氧及低氧模拟物能够诱导K562细胞系分化,抑制NHE1活性协同促进低氧诱导的K562细胞分化,其机制可能是通过增强细胞内MAPK蛋白激酶磷酸化水平促进分化。
The aim of this study was to investigate the effect of hypoxic microenvironment on the differentiation of chronic myeloid leukemia cell line K562 and the role of sodium hydrogen exchange protein 1 (NHE1) in this process. K562 cells were cultured in hypoxia mimic CoCl2 or in hypoxia (2% O2, 5% CO2 and 93% N2), and the intracellular pH (pHi) was measured by confocal laser scanning microscopy Cell morphology, real-time quantitative RT-PCR detection of gene expression, Western blot detection of MAPK phosphorylation level changes. The results showed that hypoxia-cultured K562 cells showed maturation-related morphological changes compared with the control group, accompanied by up-regulation of CCAAT / enhancer binding protein (C / EBPα) mRNA levels; Cariporide-specific NHE1 inhibitor Can promote hypoxia-induced K562 cell differentiation, Cariporide treatment of K562 cells P38 and ERK5 protein kinase phosphorylation levels were significantly enhanced. CONCLUSION: Hypoxia and hypoxia mimics can induce the differentiation of K562 cell line and inhibit the activity of NHE1 synergistically to promote hypoxia-induced K562 cell differentiation. The mechanism may be through promoting the phosphorylation of MAPK protein kinase in cells.