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Basic Research of hematopoietic stem cell transplantation utilizing CXCR4 gene transfer

利用CXCR4基因转移的造血干细胞移植的基础研究

基本信息

批准号：
16390293
负责人：
OTSU Makoto
金额：
$ 9.28万
依托单位：
University of Tsukuba
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
2004
资助国家：
日本
起止时间：
2004 至 2005
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-16390293/
关键词：
CXCR4 CXCL12 hematopietic stem cell transplantation graft failure retroviral vector WHIM syndrome 血液幹細胞骨髄移植レトロウイルベクター WHIM

项目摘要

Objective : Studies have suggested that the stromal derived factor-1(SDF-1) and its receptor CXCR4 play an important role in HSC (hematopoietic stem cell) homing and engraftment upon transplantation. Recently, C-terminus truncation of CXCR4 has been identified as causative mutations in WHIM syndrome, an immune deficiency disorder characterized by Warts, Hypogammaglobulinemia, Infections and Myelokathexis. We have tested if forced expression of truncated CXCR4 (WHIM mutants) could confer enhanced responsiveness to SDF-1 on murine HSC using a retroviral vector system.Methods : We constructed retroviral vectors that express wild type murine CXCR4 or WHIM mutants using the pGCsam-backbone, then generated highly concentrated retroviral particles pseudotyped with the vesicular stomatitis virus glycoprotein (VSV-G).Erythroleukemia K562 cells were transduced with the above retroviral vectors to test functionality of the mutant CXCR4 molecules. We also transduced HSC-rich marrow populations to test responsiveness to SDF-1 of murine HSCs expressing WHIM mutants in both in vitro and in vivo experimental systems.Results : Both K562 and murine c-Kit positive/lineage-marker negative BM cells were successfully transduced with all the retrovi Future directions : Establishment of the reliable in vivo homing assay must be achieved for future experiments.

目的：研究表明，基质衍生因子-1（SDF-1）及其受体CXCR 4在造血干细胞（hematopoietic stem cell，HSC）移植后的归巢和植入中起重要作用。最近，CXCR 4的C-末端截短已被鉴定为WHIM综合征的致病突变，WHIM综合征是一种以疣、低丙种球蛋白血症、感染和骨髓增生异常为特征的免疫缺陷疾病。我们已经测试了截短的CXCR 4的强制表达（WHIM突变体）可以使用逆转录病毒载体系统赋予小鼠HSC对SDF-1增强的应答性。我们使用pGCsam骨架构建了表达野生型鼠CXCR 4或WHIM突变体的逆转录病毒载体，然后用水泡性口炎病毒糖蛋白（VSV-G）产生高浓度的假型逆转录病毒颗粒，用上述逆转录病毒载体转导红白血病K562细胞以测试突变CXCR 4分子的功能性。我们还转导了富含HSC的骨髓群体，以测试在体外和体内实验systems.Results中表达WHIM突变体的鼠HSC对SDF-1的反应性：K562和鼠c-Kit阳性/谱系标记阴性的BM细胞都成功地转导了所有的逆转录病毒。未来的方向：必须为未来的实验建立可靠的体内归巢测定。