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本研究探讨六亚甲基二乙酰胺(HMBA)对髓系白血病HL-60细胞分化的作用及其分子机制。应用MTT比色法观察不同浓度HMBA在不同时间点对细胞增殖的抑制作用,采用Wright-Giemsa染色观察细胞形态学变化,运用流式细胞仪测定细胞分化抗原CD11b的表达,并进行细胞周期分析,半定量RT-PCR分析c-myc、mad1、p21、p27、hTERT、HDAC1基因mRNA的表达。结果表明:HL-60细胞经不同浓度(0.5、1、2mmol/L)HMBA处理后细胞增殖受到较明显的抑制,并随时间延长、剂量增大抑制作用明显增强。HL-60细胞经2mmol/L HMBA处理后,细胞形态学上趋于分化成熟,细胞停滞于G0/G1期;CD11b表达显著增高;c-myc、hTERT mRNA表达显著下调,mad1、p21、p27mRNA表达显著上调,而HDAC1mRNA表达无明显变化。结论:HMBA能诱导HL-60细胞分化,其分子机制可能是通过调节c-myc/mad1开关引起p21、p27上调,并且下调hTERT mRNA表达,与HDAC1活性抑制无关。
This study was designed to investigate the effect and molecular mechanism of HMBA on myeloid leukemia HL-60 cells. MTT assay was used to observe the inhibitory effect of different concentrations of HMBA on cell proliferation at different time points. The cell morphology was observed by Wright-Giemsa staining. The cell differentiation antigen CD11b expression was analyzed by flow cytometry and cell cycle analysis. Semi-quantitative RT-PCR analysis of c-myc, mad1, p21, p27, hTERT, HDAC1 mRNA expression. The results showed that the proliferation of HL-60 cells was significantly inhibited by HMBA at different concentrations (0.5, 1, 2mmol / L), and prolonged with increasing doses of HMBA. The inhibitory effect was obviously enhanced. The morphology of HL-60 cells treated with 2 mmol / L HMBA tended to differentiate and mature, and the cells arrested in G0 / G1 phase. The expression of CD11b was significantly increased. The expression of c-myc and hTERT mRNA was significantly down-regulated while the expression of mad1, p21 and p27 mRNA Significantly up-regulated, while HDAC1mRNA expression did not change significantly. Conclusion: HMBA can induce the differentiation of HL-60 cells. The molecular mechanism may be upregulation of p21 and p27 by regulating c-myc / mad1 switch and down-regulation of hTERT mRNA expression, which is not related to the inhibition of HDAC1 activity.