Kirby S L, Cook D N, Walton W, Smithies O
Department of Medicine, University of North Carolina, Chapel Hill 27599, USA.
Proc Natl Acad Sci U S A. 1996 Sep 3;93(18):9402-7. doi: 10.1073/pnas.93.18.9402.
The long-term efficacy of gene therapy using bone marrow transplantation requires the engraftment of genetically altered totipotent hematopoietic stem cells (THSCs). Ex vivo expansion of corrected THSCs is one way to increase the efficiency of the procedure. Similarly, selective in vivo expansion of the therapeutic THSCs rather than the endogenous THSCs could favor the transplant. To test whether a conferred proliferative advantage gene can facilitate the in vitro and in vivo expansion of hematopoietic stem cells, we have generated transgenic mice expressing a truncated receptor for the growth factor erythropoietin. These mice are phenotypically normal, but when treated in vivo with exogenous erythropoietin they exhibit a marked increase in multipotent, clonogenic hematopoietic cells [colony-forming units in the spleen (CFU-S) and CFUs that give rise to granulocytes, erythroid cells, macrophages, and megakaryocytes within the same colony (CFU-GEMM)] in comparison with the wild-type mice. In addition, long-term in vitro culture of tEpoR transgenic bone marrow in the presence of erythropoietin induces exponential expansion of trilineage hematopoietic stem cells not seen with wild-type bone marrow. Thus, the truncated erythropoietin receptor gene shows promise as a means for obtaining cytokine-inducible hematopoietic stem cell proliferation to facilitate the direct targeting of THSCs and to provide a competitive repopulation advantage for transplanted therapeutic stem cells.
使用骨髓移植进行基因治疗的长期疗效需要植入基因改变的全能造血干细胞（THSCs）。体外扩增校正后的THSCs是提高该程序效率的一种方法。同样，治疗性THSCs而非内源性THSCs的选择性体内扩增可能有利于移植。为了测试赋予增殖优势的基因是否能促进造血干细胞的体外和体内扩增，我们构建了表达截短型促红细胞生成素生长因子受体的转基因小鼠。这些小鼠表型正常，但在体内用外源性促红细胞生成素治疗时，与野生型小鼠相比，它们的多能、克隆性造血细胞[脾集落形成单位（CFU-S）以及能在同一集落中产生粒细胞、红细胞、巨噬细胞和巨核细胞的集落形成单位（CFU-GEMM）]显著增加。此外，在促红细胞生成素存在的情况下，对tEpoR转基因骨髓进行长期体外培养可诱导三系造血干细胞呈指数扩增，而野生型骨髓则未出现这种情况。因此，截短型促红细胞生成素受体基因有望作为一种手段，实现细胞因子诱导的造血干细胞增殖，以促进THSCs的直接靶向，并为移植的治疗性干细胞提供竞争性的再填充优势。
