Madlambayan Gerard J, Rogers Ian, Purpura Kelly A, Ito Caryn, Yu Mei, Kirouac Daniel, Casper Robert F, Zandstra Peter W
Institute of Biomaterials and Biomedical Engineering, University of Toronto, Toronto, Ontario, Canada.
Biol Blood Marrow Transplant. 2006 Oct;12(10):1020-30. doi: 10.1016/j.bbmt.2006.07.005.
The clinical potential of umbilical cord blood-derived stem and progenitor cells has been demonstrated in various animal and human transplantation studies. However, the need for increased numbers of appropriate umbilical cord blood-derived cells continues to limit the development and success of these therapies. Ex vivo expansion has been widely studied as a method to overcome this limitation. We describe the use of a clinically relevant single-use, closed-system bioprocess capable of generating greater numbers of hematopoietic stem and progenitor cells that maintain in vivo and in vitro developmental potential. In addition to expanded numbers of CD34+ cells, CD34(+)CD38(-) cells, colony-forming cells, and long-term culture-initiating cells, the bioprocess generated > or =3.3-fold more long-term nonobese diabetic/severe combined immunodeficient repopulating cells (quantitatively determined using limiting dilution analysis) than present at input. Interestingly, these cells were also capable of multilineage engraftment and were shown to maintain their engraftment potency on a per long-term nonobese diabetic/severe combined immunodeficient repopulating cell basis compared with input noncultured cells. The developmental capacity of bioprocess-generated cells was further demonstrated by their ability to repopulate secondary nonobese diabetic/severe combined immunodeficient recipients. In vitro lineage analysis confirmed that bioprocess-generated cells could differentiate into myeloid and natural killer, B, and T cell lymphoid lineages. This in-depth analysis describes a bioprocess that generates human hematopoietic stem and progenitor cells with conserved hematopoietic activity, establishes analysis criteria for in vitro hematopoietic stem cell expansion studies, and serves as a foundation to test the therapeutic utility of cultured hematopoietic stem cells in large animals and humans.
脐带血来源的干细胞和祖细胞的临床潜力已在各种动物和人体移植研究中得到证实。然而，对增加合适数量的脐带血来源细胞的需求仍然限制了这些疗法的发展和成功。体外扩增作为一种克服这一限制的方法已得到广泛研究。我们描述了一种临床相关的一次性、封闭系统生物工艺的应用，该工艺能够产生更多数量的造血干细胞和祖细胞，这些细胞在体内和体外均保持发育潜力。除了扩增的CD34+细胞、CD34(+)CD38(-)细胞、集落形成细胞和长期培养起始细胞外，该生物工艺产生的长期非肥胖糖尿病/重症联合免疫缺陷再填充细胞（使用极限稀释分析定量测定）比输入时多3.3倍以上。有趣的是，这些细胞还能够进行多谱系植入，并且与输入的未培养细胞相比，在每个长期非肥胖糖尿病/重症联合免疫缺陷再填充细胞的基础上显示出保持其植入能力。生物工艺产生的细胞的发育能力通过其在二次非肥胖糖尿病/重症联合免疫缺陷受体中再填充的能力进一步得到证明。体外谱系分析证实，生物工艺产生的细胞可以分化为髓系和自然杀伤细胞、B细胞和T细胞淋巴谱系。这一深入分析描述了一种产生具有保守造血活性的人类造血干细胞和祖细胞的生物工艺，建立了体外造血干细胞扩增研究的分析标准，并为测试培养的造血干细胞在大型动物和人类中的治疗效用奠定了基础。
