丙戊酸钠联合粒细胞集落刺激因子诱导髓系白血病细胞分化作用机制的探讨

Study on the mechanism of sodium valproate combined with G-CSF induced myeloid leukemia cell differentiation

目的探讨丙戊酸钠(VPA)和粒细胞集落刺激因子(G-CSF)联合用药对急性髓细胞白血病细胞株U937的诱导分化的作用机制。方法选取对数生长期U937细胞,分别经0.5 mmol/L VPA、50 ng/ml G-CSF单药或两者联合处理后,采用瑞氏染色观察细胞形态,流式细胞仪检测细胞表面抗原及G-CSF受体(G-CSFR)的变化,免疫印迹法检测髓系分化相关转录因子增强子结合蛋白(C/EBPα)及G-CSFR在处理前后的表达。结果 VPA可以诱导U937细胞髓系分化,其表面抗原CD11b表达从(11.49±3.48)%升至(62.48±9.96)%(P<0.05),联合G-CSF后CD11b表达升至(97.65±0.49)%(P<0.05)。流式细胞检测VPA能够使U937细胞表面的G-CSFR表达率从(29.50±6.80)%增至(59.88±9.99)%。VPA能够上调U937细胞C/EBPα的蛋白表达,使细胞G-CSFR蛋白水平明显升高。结论 G-CSF可以加强VPA诱导髓系白血病细胞分化。VPA可能是通过抑制组蛋白去乙酰化酶活性,促进分化转录因子C/EBPα的表达恢复及上调G-CSFR,从而发挥联合诱导分化作用。

Objective To explore the mechanism of sodium valproate （VPA） combined with granulocyte colony-stimulating factor（G-CSF） in the induction of differentiation of U937 leukemic cell line. Methods The U937 cells at logarithmic growth phase were treated with 0. 5 mmol/L VPA or 50 ng/ml G-CSF alone and in combination. Cell differentiation was analyzed by Wright-Giemsa stain cell morphology and flow cytometry. Flow cytometry was performed to detect the expression of G-CSFR in the ceils. The protein level of CCAAT/enhancer-binding proteins C/EBPαand G-CSFR were detected by Western blotting. Results VPA induced myeloid differentiation in U937 cells as demonstrated by up-regulation of C D11b from（11.49±3.48） % to （62.48±9. 96）% （P〈0. 05 ）. When combined with G-CSF, the differentiation was remarkably enhanced to （ 97.65 ±0. 49 ） % （ P 〈 0. 05 ）. The expressions of G-CSFR （C D114） on U937 cells were also up-regulated from （29. 50±6. 80）% to （59. 88±9. 99）% （P〈0. 05 ）. VPA can enhance C/EBPα expression in these ceils. The expression levels of G-CSFR protein were significantly up-regulated. Conclusion Combinations of VPA with G-CSF had an additive or synergistic effect on induction of myeloid differentiationin U937 cells. This effect may be mainly through the increase of C/EBPα and up-regulation of G-CSFR.