Elucidation of apoptotic pathway of human B-cell lymphoma and its therapeutic application

我们建立了一个新的人类伯基特淋巴瘤细胞线（HBL-9），展示了一个独特的特征，在体外文化中的自发性凋亡。在孵化和快速进展后，36小时的先天性凋亡开始。我们分析了Bcl-2家族和caspase主要使用西方印迹方法，以利用HBL-9细胞线的自发性凋亡的信号路径。免疫性化学蛋白和Western blot分析揭示了Bcl-2家族（bcl-2、bcl-X-D2 L-D2、Bax和坏蛋白）、Apaf-1和细胞色素C在每个收获细胞样本（0 h、12 h、24 h、36 h、48 h）上的表达。Western blot分析揭示的透明蛋白片段caspase-2、caspase-3、caspase-8在36小时和48小时后孵化但没有透明蛋白片段caspase-1、caspase-4、caspase-6、caspase-9、caspase-10。作为在自发性凋亡、PARP、DF 45/ICAD和拉明B的执行过程中发现的蛋白质基底，但不是拉明A。Caspase抑制剂Z-VAD-FMK和Asp-CHI-D22-DCB抑制剂D2-DCB的两种凋亡和处理caspase-2、caspase-3、caspase-7、caspase-8、caspase-2抑制剂Ac-VDVD-CHO和caspase-3/7抑制剂Ac-DEVD-CHO抑制剂处理caspase-2和caspase-3的凋亡，这些数据索引了HBL-9细胞的凋亡途径;（1）信号路径的可能性，Bcl-2家族→ Apaf-1 →细胞色素C → caspase-9，不是一家公司在HBL-9细胞中自发凋亡的执行阶段。（2）最少四个caspase（caspase-2、caspase-3、caspase-7和caspase-8）的激活要求。（3）caspase-2、caspase-3、caspase-7和caspase-8的关键效应蛋白上游的可能性。

We established a new human Burkitt's lymphoma cell line (HBL-9) which shows a unique feature of spontaneous apoptosis in vitro culture. The spontaneous apoptosis began at 36h after incubation and progressed rapidly. We analyzed bcl-2 family and caspases mainly using Western blot method to elucidate the signaling pathway for execution of spontaneous apoptosis of the HBL-9 cell line. Immunohistochemical stein and Western blot analysis revealed no expression of bcl-2 family (bcl-2, bcl-XィイD2LィエD2, bax and bad protein), Apaf-1, and cytochrome C on each harvested cell sample at the indicated times (0h, 12h, 24h, 36h, 48h). Western blot analysis revealed cleaved proteolytic fragments of caspase-2, caspase-3, caspase-8 at 36h and 48h after incubation but no cleaved proteolytic fragments of caspase-1, caspase-4, caspase-6, caspase-9, caspase-10. As protein substrates by caspase during the execution of spontaneous apoptosis, PARP, DF45/ICAD and lamin B were found but not lamin A. Caspase inhibitors Z-VAD-FMK and Asp-CHィイD22ィエD2-DCB inhibited both apoptosis and the processing caspase-2, caspase-3, caspase-7, caspase8, Caspase-2 inhibitors Ac-VDVAD-CHO and caspase-3/7 inhibitors Ac-DEVD-CHO inhibited the processing of caspase-2 and caspase-3 respectively but not apoptosis.These data indicates the following on the apoptotic pathway of HBL-9 cells;(1) The possibility that a signaling pathway, bcl-2 family→Apaf-1→cytochromeC→caspase-9, does not a company the execution phase of spontaneous apoptosis of HBL-9 cells.(2) The requirements for activations of at least four caspases (caspase-2, caspase-3, caspase-7 and caspase-8) for spontaneous apoptosis.(3) The possibility that a key effector protease upstream of caspase-2, caspase-3, caspase-7 and caspase-8.