An All-Recombinant Protein-Based Culture System Specifically Identifies Hematopoietic Stem Cell Maintenance Factors.
An All-Recombinant Protein-Based Culture System Specifically Identifies Hematopoietic Stem Cell Maintenance Factors.
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DOI：
10.1016/j.stemcr.2017.01.015
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发表时间：
2017-03-14
影响因子：
5.9
通讯作者：
Yamazaki S
中科院分区：
文献类型：
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作者：
Ieyasu A;Ishida R;Kimura T;Morita M;Wilkinson AC;Sudo K;Nishimura T;Ohehara J;Tajima Y;Lai CY;Otsu M;Nakamura Y;Ema H;Nakauchi H;Yamazaki S
造血干细胞（HSC）被认为是治疗各种血液疾病的最有前途的治疗靶点之一。然而，由于难以在体外建立稳定的维持和扩增的HSC，它们的供应不足是移植研究的主要限制。为了解决这些问题，我们已经开发了一个完全定义的，全重组蛋白为基础的培养系统。通过这个系统，我们已经确定了血红素结合素（HPX）和白细胞介素-1 α负责HSC的体外维持。随后的分子分析显示，HPX降低了培养的HSC内的细胞内活性氧水平。此外，骨髓免疫染色和3D免疫组化显示，HPX在非髓鞘形成的雪旺细胞中表达，这是已知的HSC龛成分。这些结果突出了这种完全定义的全重组蛋白为基础的培养系统的效用，可重复的体外HSC培养和其潜力，有助于识别的因素，负责在体外维持，扩增和分化的干细胞群。不同的BSA批次改变了HSC对细胞因子的反应方式RSA可以取代BSA以提供具有最小变异性的HSC维持培养。通过比较“好的”和“坏的”BSA的蛋白质谱，鉴定了HPX。和同事证明了BSA批次具有独特的蛋白质谱，并且在它们离体维持小鼠HSC的能力方面差异很大。通过用重组血清白蛋白取代BSA，他们开发了一种标准化的HSC培养平台，可用于鉴定新型维持和扩增因子。
Hematopoietic stem cells (HSCs) are considered one of the most promising therapeutic targets for the treatment of various blood disorders. However, due to difficulties in establishing stable maintenance and expansion of HSCs in vitro, their insufficient supply is a major constraint to transplantation studies. To solve these problems we have developed a fully defined, all-recombinant protein-based culture system. Through this system, we have identified hemopexin (HPX) and interleukin-1α as responsible for HSC maintenance in vitro. Subsequent molecular analysis revealed that HPX reduces intracellular reactive oxygen species levels within cultured HSCs. Furthermore, bone marrow immunostaining and 3D immunohistochemistry revealed that HPX is expressed in non-myelinating Schwann cells, known HSC niche constituents. These results highlight the utility of this fully defined all-recombinant protein-based culture system for reproducible in vitro HSC culture and its potential to contribute to the identification of factors responsible for in vitro maintenance, expansion, and differentiation of stem cell populations. Different BSA lots alter how HSCs respond to cytokines RSA can replace BSA to provide HSC maintenance culture with minimal variability By comparing the protein profiles of “good” and “bad” BSAs, HPX was identified HPX reduces HSC intracellular reactive ROS and is expressed by BM Schwann cells In this article, Yamazaki, Nakauchi, and colleagues demonstrate that BSA batches have unique protein profiles and vary widely in their ability to maintain mouse HSCs ex vivo. By replacing BSA with recombinant serum albumin, they developed a standardized HSC culture platform that can be used to identify novel maintenance and expansion factors.