抑制GSK-3β活性促进NF-κB诱导的儿童急性淋巴细胞白血病细胞凋亡

Inhibition of Glycogen Synthase Kinase-3β Promotes NF-κB Induced-apoptosis in Pediatric Acute Lymphocyte Leukemia Cells

核转录因子-κB(NF-κB)是维持急性淋巴细胞白血病(ALL)细胞生存的关键因子.近年来发现,糖原合成酶激酶-3β(GSK-3β)可以正性调控NF-κB的活性.本研究通过抑制GSK-3β活性初步探讨ALL细胞中GSK-3β在NF-κB诱导细胞凋亡中的作用机制.收集ALL患儿骨髓单个核细胞,采用免疫荧光细胞化学方法检测到ALL细胞核内GSK-3β有明显聚集.体外培养ALL细胞后经GSK-3β抑制剂氯化锂(LiCl)和SB216763处理,采用Western印迹和EMSA检测发现,ALL细胞核内GSK-3β表达下降,而NF-κBP65蛋白无明显变化,但是其活性明显降低.同时RT-PCR结果显示,NF-κB下游抗凋亡基因存活素(survivin)的表达随之下降,AnnexinV-PE/7-AAD双染流式细胞仪检测结果证实,ALL细胞凋亡明显增加(P<0.01).该结果表明,抑制GSK-3β活性可以下调NF-κB的转录活性,并通过下调抗凋亡基因存活素的表达而促进ALL细胞的凋亡.

Nuclear factor-κB （NF-κB） is a key factor in acute lymphoblastic leukemia （ALL） cell survival,.Glycogen synthase kinase-3β （GSK-3β） has recently been found to positively regulate the activity of NF-κB. Here, we have explored the effect of GSK-3β inhibition on NF-κB-induced apoptosis in ALL cells. Mononuclear cells were isolated from the heparinized bone marrow of children with ALL by density gradient centrifugation. GSK-3β significantly accumulates in nuclei of ALL cells by immunofluorescence staining method. ALL cells were treated with GSK-3β inhibitors, lithium chloride （LiC1） or SB216763 in vitro, leading to decrease of its nuclear pool, and suppression NF-κB transcriptional activity by Western blot and EMSA analyses. Furthermore, we observed that GSK-3β inhibition resulted in a marked reduction in the expression of the NF-κB antiapoptotic target gene survivin by RT-PCR analysis, and a dose-dependent increase in the number of apoptotie cells by Annexin V-PE/ 7-AAD double staining flow cytometry. These results suggest that GSK-3β inhibition could down-regulate NF-κB transcriptional activity and the gene expression of survivin, holding the potential to enhance apoptosis in pediatric ALL ceils.