脱氧佛波醇乙酸酯通过激活PKC/ERK通路诱导急性髓系白血病细胞分化

12-Deoxyphorbol 13-acetate inducing acute myeloid leukemia cell differentiation through PKC/ERK pathway

目的研究脱氧佛波醇乙酸酯(12-deoxyphorbol 13-acetate,Prostratin,DPA)对人急性髓系白血病(AML)细胞株HL-60、NB4和U937细胞分化的影响,并证明其通过激活蛋白激酶C/胞外信号调节激酶(PKC/ERK)通路诱导细胞分化。方法 1μmol/L DPA作用HL-60细胞3 d后观察细胞形态的改变;不同剂量的DPA作用HL-60、NB4、U937细胞24 h后,流式细胞仪检测细胞分化表面标志变化;Western印迹检测ERK蛋白磷酸化的改变;流式细胞仪检测丝裂原活化蛋白激酶激酶(MEK)抑制剂和PKC抑制剂对细胞分化表面标志的影响;Western印迹检测MEK抑制剂和PKC抑制剂对ERK蛋白磷酸化的影响。结果 DPA可诱导HL-60细胞出现白血病细胞分化的形态。DPA剂量依赖性地诱导AML细胞CD11b表达。DPA在诱导分化剂量范围内可剂量依赖性地引起ERK磷酸化。MEK化学小分子抑制剂U0126可完全抑制DPA引起的ERK磷酸化和CD11b表达;PKC非选择性抑制剂GFX可完全抑制DPA引起的ERK磷酸化和CD11b表达。结论 DPA其通过激活PKC/ERK通路能够明显诱导白血病细胞分化。

ObjectiveTo explore the effect of 12-deoxyphorbol 13-acetate（Prostratin,DPA） on cell differentiation in acute myeloid leukemia（AML） cell lines,and to prove that the mechanism of action is through PKC/ERK pathway.Methods HL-60 cells were cultured with 1 μmol/L DPA,and cell morphology was observed by microscope after 3 days.AML cells were cultured with varying concentrations of DPA.After 24 h of treatment,the CD11b expression level was determined by flow cytometry.The protein expression of phospho-ERK was detected by Western blot.The effect of U0126 and GFX on HL-60 cells＇ CD11b expression was measured by flow cytometry.The effect of U0126 and GFX on AML cells＇ phospho-ERK expression was measured by Western blot.ResultsDPA could induce HL-60 cells differentiation and exhibit a typical cell morphology.DPA up-regulated the expression of CD11b in AML cells in a concentration-dependent manner,and exhibited a dose dependent up-regulatory effect on phospho-ERK expression in the same varying concentrations.Both U0126 and GFX inhibited the expression of CD11b and phospho-ERK induced by DPA.ConclusionDPA can induce significant cell differentiation of AML cells.The underlying mechanisms may be through activating PKC/ERK pathway.