雪胆素乙诱导刀豆蛋白A刺激的小鼠淋巴细胞凋亡及其机制的研究

摘要

目的：分析雪胆素乙（CuIIb）对刀豆蛋白A（Con A）刺激的小鼠淋巴细胞体外凋亡的影响，并探讨其作用机制。方法：以Annexin V染色结合流式细胞术分析小鼠淋巴细胞凋亡情况；利用JC-1染色分析淋巴细胞线粒体膜电位变化；Western blotting检测凋亡相关蛋白变化。结果： CuIIb处理后，早期和中期凋亡细胞比例明显增加，淋巴细胞线粒体膜电位降低。同时，CuIIb以剂量依赖方式激活caspase-3凋亡通路并明显降低抗凋亡蛋白survivin的表达。结论：CuIIb诱导淋巴细胞凋亡，其机制可能与其降低线粒体膜电位、激活caspase-3凋亡通路有关。%AIM:To explore the potential anti-inflammatory effect and the underlying mechanism of cucurbita-cin II b (CuIIb) on concanavalin A (Con A)-activated mouse lymphocytes.METHODS:The cell apoptosis was analyzed by flow cytometry with Annexin V staining.The change of mitochondrial membrane potential was detected by JC -1 staining. The levels of caspase-3, phosphorylated H2AX and survivin were detected by Western blotting .RESULTS: Compared with Con A-activated lymphocytes, the significant increases in the proportion of the cells with reduced mitochondrial mem-brane potential or with increased Annexin V staining were observed , reflecting increased apoptosis in CuIIb plus Con A-treated cells.The increased levels of activated caspase-3 and phosphorylated H2AX (a marker of double-stranded DNA breaks) were manifested in Con A-activated cells co-treated with CuIIb.Although only a slight difference of cleavage of po-ly(ADP-ribose) polymerase was found in the cells with or without CuIIb treatment , the expression of survivin, an inhibitor of apoptosis protein, was markedly reduced by CuIIb in Con A-activated cells.CONCLUSION:CuIIb promotes apoptosis in mouse lymphocytes.