Enforced Expression of HOXB4 in Human Embryonic Stem Cells Enhances the Production of Hematopoietic Progenitors but Has No Effect on the Maturation of Red Blood Cells.
Enforced Expression of HOXB4 in Human Embryonic Stem Cells Enhances the Production of Hematopoietic Progenitors but Has No Effect on the Maturation of Red Blood Cells.
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DOI：
10.5966/sctm.2015-0324
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发表时间：
2016-08
影响因子：
6
通讯作者：
Forrester LM
中科院分区：
文献类型：
--
作者：
Jackson M;Ma R;Taylor AH;Axton RA;Easterbrook J;Kydonaki M;Olivier E;Marenah L;Stanley EG;Elefanty AG;Mountford JC;Forrester LM
能够从人类多能干细胞产生可扩展数量的红细胞 (RBC) 的稳健分化方案向临床的转化受到了影响，因为产生的红细胞尚未完全成熟，表达胚胎和胎儿球蛋白，并且不能有效地去核。 HOXB4 编程增加了造血祖细胞和未成熟红细胞的产生，但无法解决与成熟成人样去核红细胞产生相关的固有挑战。我们开发了一种强大的、符合良好生产规范的分化方案，能够从人类多能干细胞 (hPSC) 中产生可扩展数量的红细胞 (RBC)。然而，由于产生的红细胞尚未完全成熟，该方案向临床的转化受到了影响；因此，它们表达胚胎和胎儿的球蛋白，而不是成年球蛋白，并且它们不能有效地去核。根据之前的研究，我们预测外源 HOXB4 的激活会增加 hPSC 的造血祖细胞 (HPC) 的产生，并假设它还可能促进更成熟、确定的红细胞的产生。使用他莫昔芬诱导的 HOXB4-ERT2 表达系统，我们首先证明 HOXB4 的激活确实增加了 hPSC 的 HPC 产量，这一点通过集落形成单位培养物活性和 CD43+CD34+ 祖细胞的存在来确定。 HOXB4 的激活导致未成熟 CD235a+/CD71+ 红系细胞比例适度但显着增加。然而，这并没有导致更成熟的 CD235a+/CD71− 细胞的显着增加。在 HOXB4 活性增强的情况下产生的红细胞表达胚胎 (ε) 和胎儿 (γ) 球蛋白，但不表达成人 (β) 球蛋白，并且去核细胞的比例与对照培养物相当。我们得出的结论是，用转录因子 HOXB4 进行编程可以增加造血祖细胞和未成熟红系细胞的产生，但不能解决与成熟成人样去核红细胞产生相关的固有挑战。随着全球献血量的减少和可输血传播感染的增加，人们对从多能干细胞等替代来源体外提取红细胞 (RBC) 产生了浓厚的兴趣。开发了可翻译的协议来生成红细胞；然而，这些红细胞具有不成熟的表型。据推测，转录因子 HOXB4 可以增强它们的产生和成熟。尽管 HOXB4 增加了红系祖细胞的产生，但它并没有促进它们的成熟。尽管仍然存在挑战，但已经建立了一个强大的系统来测试其他候选人并补充该领域的知识库。
Translation of a robust differentiation protocol capable of producing scalable quantities of red blood cells (RBCs) from human pluripotent stem cells to the clinic was compromised because the RBCs produced were not fully mature, expressed embryonic and fetal globins, and did not enucleate efficiently. Programming with HOXB4 increased hematopoietic progenitor and immature erythroid cell production but could not resolve the inherent challenges associated with mature adult-like enucleated RBC production. We have developed a robust, Good Manufacturing Practice-compatible differentiation protocol capable of producing scalable quantities of red blood cells (RBCs) from human pluripotent stem cells (hPSCs). However, translation of this protocol to the clinic has been compromised because the RBCs produced are not fully mature; thus, they express embryonic and fetal, rather than adult globins, and they do not enucleate efficiently. Based on previous studies, we predicted that activation of exogenous HOXB4 would increase the production of hematopoietic progenitor cells (HPCs) from hPSCs and hypothesized that it might also promote the production of more mature, definitive RBCs. Using a tamoxifen-inducible HOXB4-ERT2 expression system, we first demonstrated that activation of HOXB4 does increase the production of HPCs from hPSCs as determined by colony-forming unit culture activity and the presence of CD43+CD34+ progenitors. Activation of HOXB4 caused a modest, but significant, increase in the proportion of immature CD235a+/CD71+ erythroid cells. However, this did not result in a significant increase in more mature CD235a+/CD71− cells. RBCs produced in the presence of enhanced HOXB4 activity expressed embryonic (ε) and fetal (γ) but not adult (β) globins, and the proportion of enucleated cells was comparable to that of the control cultures. We conclude that programming with the transcription factor HOXB4 increases the production of hematopoietic progenitors and immature erythroid cells but does not resolve the inherent challenges associated with the production of mature adult-like enucleated RBCs. As worldwide blood donations decrease and transfusable transmitted infections increase, intense interest has ensued in deriving red blood cells (RBCs) in vitro from alternative sources such as pluripotent stem cells. A translatable protocol was developed to generate RBCs; however, these RBCs have an immature phenotype. It was hypothesized that the transcription factor HOXB4 could enhance their production and maturation. Although HOXB4 increased the production of erythroid progenitors, it did not promote their maturation. Despite the remaining challenges, a robust system has been established to test other candidates and add to the knowledge base in this field.