Tiwari Abhilasha, Wong Cynthia S, Nekkanti Lakshmi P, Deane James A, McDonald Courtney, Li Jingang, Pham Yen, Sutherland Amy E, Jenkin Graham, Kirkland Mark A
The Ritchie Centre, Hudson Institute of Medical Research, Clayton, Victoria, Australia.
Institute for Frontier Materials, Deakin University, Geelong, Victoria, Australia.
Curr Protoc Stem Cell Biol. 2018 Feb 28;44:2A.11.1-2A.11.13. doi: 10.1002/cpsc.42.
Clinical hematopoietic stem/progenitor cell (HSPC) transplantation outcomes are strongly correlated with the number of cells infused. Hence, to generate sufficient HSPCs for transplantation, the best culture parameters for expansion are critical. It is generally assumed that the defined oxygen (O ) set for the incubator reflects the pericellular O to which cells are being exposed. Studies have shown that low O tension maintains an undifferentiated state, but the expansion rate may be constrained because of limited diffusion in a static culture system. A combination of low ambient O and dynamic culture conditions has been developed to increase the reconstituting capacity of human HSPCs. In this unit, the protocols for serum-free expansion of HSPCs at 5% and 20% O in static and dynamic nutrient flow mode are described. Finally, the impact of O tension on HSPC expansion in vitro by flow cytometry and colony forming assays and in vivo through engraftment using a murine model is assessed. © 2018 by John Wiley & Sons, Inc.
临床造血干细胞/祖细胞（HSPC）移植结果与输注的细胞数量密切相关。因此，为了产生足够用于移植的HSPC，最佳的扩增培养参数至关重要。通常认为，培养箱设定的特定氧气（O）水平反映了细胞所接触的细胞周围O水平。研究表明，低氧张力可维持细胞的未分化状态，但在静态培养系统中，由于扩散受限，扩增速率可能会受到限制。已开发出低环境氧与动态培养条件相结合的方法来提高人HSPC的重建能力。在本单元中，将描述在静态和动态营养流模式下，在5%和20% O条件下HSPC无血清扩增的方案。最后，通过流式细胞术和集落形成试验评估氧张力对体外HSPC扩增的影响，并通过小鼠模型体内植入评估其影响。© 2018约翰威立父子公司。
