PKC-ERK通路在热损伤诱导单核细胞凋亡中的作用

应用流式细胞检测术、Western印迹、激酶活性测定等技术,检测PKC与ERK在热损伤诱导单核细胞株Raw264.7细胞凋亡中的作用。结果显示热损伤导致PKC短暂激活,PKC激活剂佛波脂(PMA)与热损伤联合作用导致PKC持续活化;并且PKC的持续激活抑制热损伤诱导的Raw264.7细胞凋亡,而PKC的抑制可促进细胞凋亡;ERK活性检测显示热损伤抑制ERK磷酸化,而PMA激活ERK磷酸化活化,并且这种激活作用通过PKC;进一步细胞凋亡检测显示ERK抑制剂PD098059可解除PMA对热损伤诱导Raw264.7细胞凋亡的抑制作用,从而提示PKC通过ERK负调控热损伤诱导的Raw264.7细胞凋亡。

The Regulation of PKC-ERK Signaling Pathway on Heat-induced Monocytic Cell Apoptosis

By using a combined methods including flow cytometric analysis,Western blotting and kinase assay,the effect of PKC-ERK signaling pathway on heat-induced apoptosis in monocytic cell line Raw264.7 was studied. The results demonstrated that PKC was transiently activated 5 minutes after heating and returned to basal level 2 hours later,while PKC was persistently activated by phorbol-12-myristate-13-acetate (PMA). The persistent activation of PKC can inhibit heat-induced Raw264.7 cell apoptosis,while inhibition of PKC promoted heat-induced Raw264.7 cell apoptosis. Furthermore,kinase analysis showed that heat can suppress the activation of ERK,while PMA can induce ERK activation. The inhibitor of PKC,staurosporine,can attenuate the inhibitory effect of PMA on heat-induced Raw264.7 cell apoptosis. Together,the present results indicate that the PKC-ERK signaling pathway can negatively regulate heat-induced Raw264. 7 cell apoptosis.