Caspase-3在脐血CD34+细胞中的表达及意义

为探讨脐血CD34＾＋细胞在不同细胞因子支持下体外扩增过程中caspase－3表达及意义，采用RT—PCR，Western blot和流式细胞术对脐血CD34＾＋细胞在体外扩增过程中的细胞增殖、细胞凋亡及caspase—3的表达进行了测定。检测结果显示，casPase—3　mRNA在新鲜分离的脐血CD34＾＋细胞中低水平表达，在细胞因子支持下体外培养3天，扩增的CD34＾＋细胞中caspase—3　mRNA和蛋白质水平表达上调，但在这两种细胞中仅能检测到分子量为32kD的非活性酶原形式的casPase－3，随着体外培养时间的延长，在无早期效应细胞因子SCF或FL存在的条件下，casPase-3被激活，可检测到分子量为20kD的裂解片段，细胞凋亡率增加。结论：虽然造血干／祖细胞的凋亡是个复杂的过程，但在脐血CD34＾＋干／祖细胞体外扩增过程中，caspase—3参与了细胞凋亡事件并发挥着重要的作用，早期效应细胞因子SCF和FL可减少造血干／祖细胞的凋亡，对造血干／祖细胞有保护作用。

Expression of Caspase-3 in CD34 + Cord Blood Cells and Its Significance

To evaluate expression of caspase 3 protein in CD34 + cells from cord blood (CB) and its significance during culture in vitro with various growth factors, RT PCR, Western blot and flow cytometry were used to determine cell proliferation, apoptosis and expression of caspase 3 in CD34 + cells during culture in vitro. The results demonstrated that caspase 3 mRNA was constitutively expressed at a low level in freshly isolated CD34 + cells. Expression of caspase 3 mRNA and protein was upregulated when these cells were expanded in suspension culture with growth factors for 3 days. However, only the 32 kD inactive caspase 3 proenzyme was detected in the freshly isolated CD34 + cells and those cells after 3 days expansion with cytokines. Along with the culture time extension (7 days) in vitro, especially without early effective cytokines SCF or FL existence, caspase 3 was activated and a cleavage 20 kD was detected. It is concluded that the caspase 3 is involved in apoptosis of primitive CD34 + cells during expansion in vitro, and early effective cytokines, SCF and FL, could conserve hematopoietic stem cells and suppress apoptosis.