丁酸钠增强U937细胞凋亡敏感性的分子机制研究

Molecular mechanism of enhanced apoptotic response in U937 cells mediated by sod ium butyrate

目的 探讨丁酸钠 (NaBu)对细胞周期检测点效应及对U937细胞凋亡的敏感性。方法以U937 ASPI3K(ATM阴性 )、U937 pZeosv2 (+) (野生型ATM基因 )两种U937的变异细胞系作为细胞模型。用免疫沉淀及激酶活性测定p38MAPK、ERK1的激酶活性。用免疫印迹分析Bad磷酸化灭活。结果 经NaBu预处理的U937 pZeosv2 (+)细胞经13 7Cs照射后 ,细胞凋亡敏感性呈NaBu剂量依赖性增强。这种增强效应可被p38MAPK阻滞剂OLM阻断 ,但不能被p34cdc2激酶的特异性抑制剂ALP及CDK2阻滞剂CDK2 Ⅰ阻断。经NaBu预处理的U937 ASPI3K细胞经13 7Cs照射后 ,细胞凋亡敏感性进一步增强 ,这种增强效应可被OLM阻断。放射线可显著增强p38MAPK激酶活性 ,抑制ERK1激酶活性 ;NaBu预处理与放射线联用后 ,对p38MAPK激酶活性的增强有极其显著的协同效应。放射线诱导U937 ASPI3K细胞Bad蛋白磷酸化灭活 ,在NaBu协同作用下 ,Bad蛋白磷酸化灭活效应进一步增强。结论 NaBu通过p38MAPK激酶活性 ,增强细胞凋亡敏感性 ,该效应与ATM基因是否缺失无关 。

Objective To study the effects of sodium butyrate (NaBu) on cell cycle checkpoint and the apoptosis sensitivity in U937 cells. Methods Two mutant U937 cell lines, U937 ASPI3K (ATM negative) and U937 pZeosv2(+) (AT M wild type), were used as the cell model system. Immunoprecipitation and kina se assay were used to examine the p38 MAPK and ERK1 kinase activities. Western b lot was used to analyze the phosphorylation of Bad protein. Results U937 pZeosv2(+) pretreated with NaBu exhibited enhanced apoptotic response in a NaBu dose dependent fashion upon 137 Cs irradiation, which could be abol is hed by olomoucine (OLM), a p38 MAPK specific inhibitor. On the other hand, Cycli n dependent kinase 2 (CDK2) specific inhibitor CDK2 Ⅰ and p34cdc2/cyclinB inhi bitor alsterpaullone (ALP) failed to block the effects of NaBu. Similar results were also observed in U937 ASPI3K. The effect of irradiation on p38 MAPK and ER K1 was strikingly potentiated by NaBu. Furthermore, inactivation of irradiated B ad protein via phosphorylation on serine 136 was also enhanced. Conclusion NaBu is able to enhance the apoptotic response in U937 cells, which is mediated by p38 MAPK activation but not ATM status. [WT5'HZ