Small molecule compounds have been well recognized for their promising power in the generation, expansion, and maintenance of embryonic or adult stem cells. The aim of this study was to identify a novel combination of small molecules in order to optimize the ex vivo expansion of umbilical cord blood-derived CD34+ cells. Considering the most important signaling pathways involved in the self-renewal of hematopoietic stem cells, CB-CD34+ cells were expanded with cytokines in the presence of seven small molecules including SB, PD, Chir, Bpv, Pur, Pμ, and NAM. The eliminativism approach was used to find the best combination of selected small molecules for effective ex vivo expansion of CD34+ cell. In each step, proliferation, self-renewal, and clonogenic potential of the expanded cells as well as expression of some hematopoietic stem cell-related genes were studied. Finally, the engraftment potential of expanded cells was also examined by the mouse intra-uterine transplantation model. Our data shows that the simultaneous use of SB431542 (TGF-β inhibitor), Chir9901 (GSK3 inhibitor), and Bpv (PTEN inhibitor) resulted in a 50-fold increase in the number of CD34+CD38− cells. This was further reflected in approximately 3 times the increase in the clonogenic potential of the small molecule cocktail-expanded cells. These cells, also, showed a 1.5-fold higher engraftment potential in the peripheral blood of the NMRI model of in utero transplantation. These results are in total conformity with the upregulation of HOXB4, GATA2, and CD34 marker gene as well as the CXCR4 homing gene. Taken together, our findings introduce a novel combination of small molecules to improve the yield of existing protocols used in the expansion of hematopoietic stem cells.

中文翻译：

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SB431542，Chir9901和Bpv的组合作为脐血造血干细胞培养的新型补充剂

小分子化合物因其在胚胎或成体干细胞的产生，扩增和维持中的潜力而广为人知。这项研究的目的是确定一种新的小分子组合，以优化脐带血来源的CD34 +细胞的离体扩增。考虑到造血干细胞自我更新中涉及的最重要的信号通路，在七个小分子（包括SB，PD，Chir，Bpv，Pur，Pμ和NAM）存在下，CB-CD34 +细胞被细胞因子扩增。进化论方法用于找到所选小分子的最佳组合，以有效地离体扩增CD34 +细胞。在每一步中，扩散，自我更新，研究了扩增细胞的克隆形成潜能以及一些造血干细胞相关基因的表达。最后，还通过小鼠子宫内移植模型检查了扩增细胞的植入潜力。我们的数据表明，同时使用SB431542（TGF-β抑制剂），Chir9901（GSK3抑制剂）和Bpv（PTEN抑制剂）可导致CD34 + CD38-细胞数量增加50倍。这进一步反映在小分子混合物扩张的细胞的克隆形成潜能增加约3倍。这些细胞在子宫内移植的NMRI模型的外周血中也显示出1.5倍的更高植入潜力。这些结果与HOXB4，GATA2和CD34标记基因以及CXCR4归巢基因的上调完全一致。