Dao M A, Nolta J A
Division of Research Immunology/Bone Marrow Transplantation, Children's Hospital Los Angeles, CA, USA.
Cytokines Cell Mol Ther. 1997 Jun;3(2):81-9.
Interleukin 3 (IL-3) supports the survival of multilineage hematopoietic progenitors, and increases the extent of retrovirally mediated gene transfer into colony-forming cells. However, effects from the supraphysiological levels used in ex-vivo expansion and gene-therapy protocols on subsequent differentiation of human progenitors have not been well defined. In the current studies, the extents of retrovirally mediated transduction and lineage development from CD34+ cells cultured ex vivo in the presence or absence of IL-3 were compared. All transductions were performed in the presence of an irradiated stromal support layer, IL-6 and SCF, with and without inclusion of 10 ng/ml IL-3. Following transduction, colony-forming (CFU) assays were done, and the remaining cells were transplanted into cohorts of sibling beige/nude/xid (bnx) mice. Marrow from the mice was harvested 9-10 months post transplantation. The average extent of human CD45+ cell engraftment in the bone marrow and the human hematopoietic lineages recovered from the mice in the +IL-3 and -IL-3 groups did not vary significantly. No deleterious effects on the extent of engraftment, lineage generation, or survival of clonogenic progenitors was observed with inclusion of IL-3 in the transductions performed on stromal support. The percentage of G418-resistant human progenitors recovered from mice was equivalent. The extent of marking by the neo gene in the marrow of the mice was equal in both groups, and inverse PCR revealed that primitive cells transduced in the absence of IL-3 had generated progeny with slightly better clonal diversity than progenitors transduced in the presence of IL-3. These data show that, while transduction of colony-forming progenitors may not always be apparent, primitive human hematopoietic cells can be transduced to significant levels in the absence of IL-3.
白细胞介素3（IL-3）可维持多谱系造血祖细胞的存活，并增加逆转录病毒介导的基因转移至集落形成细胞的程度。然而，体外扩增和基因治疗方案中使用的超生理水平对人类祖细胞后续分化的影响尚未明确界定。在当前研究中，比较了在有或无IL-3存在的情况下体外培养的CD34+细胞的逆转录病毒介导转导程度和谱系发育情况。所有转导均在有照射的基质支持层、IL-6和SCF存在的情况下进行，添加或不添加10 ng/ml IL-3。转导后，进行集落形成（CFU）测定，其余细胞移植到同基因米色/裸鼠/ xid（bnx）小鼠群体中。移植后9 - 10个月从小鼠采集骨髓。+IL-3组和 -IL-3组小鼠骨髓中人CD45+细胞植入的平均程度以及从小鼠中恢复的人类造血谱系没有显著差异。在基质支持下进行转导时加入IL-3，未观察到对植入程度、谱系生成或克隆性祖细胞存活有有害影响。从小鼠中回收的对G418耐药的人类祖细胞百分比相当。两组小鼠骨髓中neo基因的标记程度相同，反向PCR显示在无IL-3情况下转导的原始细胞产生的子代克隆多样性略优于在有IL-3情况下转导的祖细胞。这些数据表明，虽然集落形成祖细胞的转导可能并不总是明显，但在无IL-3的情况下，原始人类造血细胞可被转导至显著水平。
