PD98059对HL60细胞Ras-MEK1/2-ERK1/2信号转导影响的研究

Impact of PD98059 on the transduction of Ras-MEK1/2-ERK1/2 signaling pathway in HL60 cells

目的探讨MEK1抑制剂PD98059对HL60细胞Ras-MEK1/2-ERK1/2信号转导以及细胞增殖的影响。方法四唑盐比色试验(MTT)法检测PD98059对HL60细胞增殖的抑制作用,流式细胞术观察PD98059对HL60细胞凋亡和G0/G1期阻滞的影响,半定量反转录-聚合酶链反应(RT-PCR)测定PD98059对HL60细胞ERK2、p27、Skp2基因表达的影响,免疫组织化学SP法检测ERK2、p27、Skp2蛋白表达的改变情况。结果 PD98059能够抑制HL60细胞的生长,该抑制作用具有浓度及时间依赖性(P<0.05)。PD98059能够促进HL60细胞凋亡,该作用具有浓度及时间依赖性(P<0.05);PD98059作用HL60细胞48h,随着浓度的增加G0/G1期阻滞增强(P<0.05)。PD98059可使HL60细胞ERK2、Skp2的mRNA及蛋白表达量减少,p27的mR-NA及蛋白表达量增加(P<0.05)。结论 PD98059能够抑制HL60细胞的生长,诱导细胞发生G0/G1期阻滞,促进其凋亡,这可能是通过PD98059阻断Ras-MEK1/2-ERK1/2信号转导,影响了ERK2、Skp2、p27等的表达而实现的。

Objective To explore the suppressive effects of PD98059,the inhibitor of MEKl（extracellular signal-regulated kinase） ,on the Ras-MEK1/2-ERK1/2 signaling pathway＇s transduction and on the proliferation of acute promyelocytic leukemia cell line HL60 cells. Methods The suppressive effects of PD98059 on proliferation of HL60 was measured by MTT assay. Flow cytometry was used to detect the effects of PD98059 on cell apoptosis and cell cycle changes. The mRNA expressions of ERK2, Skp2 and p27 were analyzed by semi-quantitative RT- PCR. Immunohistochemistry was employed to examine the protein expressions of ERK2, Skp2 and p27. Results PD98059 inhibited the growth of HL60 cells, in a dose-and time-dependent manner. PD98059 induced apoptosis of HL60 cells in vitro, and blocked the cells to the Go/G~ phase. PD98059 reduced the mRNA and protein expres- sions of ERK2 and Skp2,in contrast increased the expressions of p27. Conclusion PD98059 has a suppressive effect on HL60 cells growth and induces the apoptosis of HL60 cells in vitro through regulating the process of cell cycle. This effect may be performed by that PD98059 block the Ras-MEK1/2-ERK1/2 signaling pathway in HL60 cells.