Department of Stem Cells and Developmental Biology, Cell Science Research Center, Royan Institute for Stem Cell Biology and Technology, ACECR, P.O. Box, Tehran, 19395-4644, Iran.
Royan Stem Cell Technology Company, Cord Blood Bank, Tehran, Iran.
Stem Cell Res Ther. 2020 Nov 9;11(1):474. doi: 10.1186/s13287-020-01945-8.
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 CD34CD38 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.
小分子化合物在产生、扩增和维持胚胎或成体干细胞方面的潜力已得到广泛认可。本研究旨在寻找一种新的小分子组合，以优化脐带血来源的 CD34 细胞的体外扩增。
考虑到与造血干细胞自我更新最相关的信号通路，CB-CD34 细胞在七种小分子（SB、PD、Chir、Bpv、Pur、Pμ 和 NAM）存在的情况下用细胞因子扩增。采用消除法寻找最佳组合，以有效扩增 CD34 细胞。在每一步中，都研究了扩增细胞的增殖、自我更新和克隆形成能力以及一些造血干细胞相关基因的表达。最后，通过小鼠子宫内移植模型检测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）可使 CD34CD38 细胞数量增加 50 倍。这进一步反映在小分子鸡尾酒扩增细胞的克隆形成能力增加了约 3 倍。这些细胞在 NMRI 模型子宫内移植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。
总之，我们的研究结果介绍了一种新的小分子组合，以提高现有用于扩增造血干细胞的方案的产量。
