Hematti Peiman, Tuchman Sascha, Larochelle Andre, Metzger Mark E, Donahue Robert E, Tisdale John F
Hematology Branch, NHLBI, National Institutes of Health, 9000 Rockville Pike, Bethesda, MD 20892, USA.
Stem Cells. 2004;22(6):1062-9. doi: 10.1634/stemcells.22-6-1062.
Hematopoietic stem cells (HSCs) are ideal targets for genetic manipulation in the treatment of several congenital and acquired disorders affecting the hematopoietic compartment. Although G-CSF-mobilized peripheral blood CD34(+) cells are the favored source of hematopoietic stem cells in clinical transplantation, this source of stem cells does not provide meaningful engraftment levels of genetically modified cells compared with G-CSF + stem cell factor (SCF)-mobilized cells in nonhuman primates. Furthermore, the use of G-CSF mobilization can have disastrous consequences in patients with sickle cell disease, a long-held target disorder for HSC-based gene therapy approaches. We therefore conducted a study to compare the levels of genetically modified cells attainable after retroviral transduction of CD34(+) cells collected from a bone marrow (BM) harvest with CD34(+) cells collected from a leukapheresis product after mobilization with G-CSF (n = 3) or G-CSF in combination with SCF (n = 3) in the rhesus macaque autologous transplantation model. Transductions were performed using retroviral vector supernatant on fibronectin-coated plates for 96 hours in the presence of stimulatory cytokines. BM was equal to or better than G-CSF-mobilized peripheral blood as a source of HSCs for retroviral transduction. Although the highest marking observed was derived from G-SCF + SCF-mobilized peripheral blood in two animals, marking in the third originated only from the BM fraction. These results demonstrate that steady-state BM is at least equivalent to G-CSF-mobilized peripheral blood as a source of HSCs for retroviral gene transfer and the only currently available source for patients with sickle cell disease.
造血干细胞（HSCs）是基因操作治疗影响造血系统的多种先天性和获得性疾病的理想靶点。尽管粒细胞集落刺激因子（G-CSF）动员的外周血CD34(+)细胞是临床移植中造血干细胞的首选来源，但与非人灵长类动物中G-CSF加干细胞因子（SCF）动员的细胞相比，这种干细胞来源并不能提供转基因细胞有意义的植入水平。此外，对于镰状细胞病患者（基于造血干细胞的基因治疗方法长期以来的目标疾病），使用G-CSF动员可能会产生灾难性后果。因此，我们进行了一项研究，在恒河猴自体移植模型中，比较从骨髓（BM）采集物中收集的CD34(+)细胞与用G-CSF（n = 3）或G-CSF联合SCF（n = 3）动员后从白细胞分离产物中收集的CD34(+)细胞进行逆转录病毒转导后可获得的转基因细胞水平。在刺激细胞因子存在的情况下，使用逆转录病毒载体上清液在纤连蛋白包被的平板上进行转导96小时。作为逆转录病毒转导的造血干细胞来源，骨髓与G-CSF动员的外周血相当或更好。尽管在两只动物中观察到的最高标记来自G-SCF + SCF动员的外周血，但第三只动物中的标记仅来自骨髓部分。这些结果表明，作为逆转录病毒基因转移的造血干细胞来源，稳态骨髓至少等同于G-CSF动员的外周血，并且是镰状细胞病患者目前唯一可用的来源。
