去除血清诱发细胞凋亡和Bcl—2蛋白的裂解

以过表达原癌基因Bcl-2的HL－60细胞为材料，通过去除血清而诱发凋亡，应用Western blot方法检测细胞凋亡过程中Bcl-2裂解为分子量大约为20kDa的片段，进一步检测Bcl-2的结构变化，结果显示Bcl-2蛋白裂解发生在N端，裂解产物丢失了BH4结构域。通过加入凋亡相关蛋白酶caspase-3抑制剂，可抑制20kDa片段的产生，表明凋亡过程中caspase-3的激活，但细胞生长率测定表明，caspase-3抑制剂并不能阻止去血清诱发的细胞死亡。

INDUCTION OF APOPTOSIS BY WITHDRAWAL SERUM AND CILAEVAGE OF BCL-2 IN HL-60 CELL

Apoptosis were induced by withdrawal serum from culture medium. Bcl-2 protein was cleaved to produce a 20 kDa fragment during apoptosis in HL-60 cell. To assay the change of Bcl-2 protein, two antibody corresponding to peptides 41～54 and 4～21 of Bcl-2 were used. The cleaved Bcl-2 that was cross-immunorective to 41～54 Bcl-2 monoclonal antibody but not 4～21 Bcl-2 polyclonal antibody. This results suggest that Bcl-2 was removed the N-terminal BH-4 region known to be essential for the death-protective activity of Bcl-2. A specific inhibitor of caspase-3, Z-DEVD-FMK, was added to HL-60 cells cultured with serum-free to determine whether caspase-3 could inhibit cleavage of Bcl-2. Bcl-2 cleavage was inhibited completely but no reversed cell growth inhibition by withdrawal serum.