Department of Pathology, University of California San Francisco, San Francisco, CA.
Children's Hospital Oakland Research Institute, University of California San Francisco, San Francisco, CA.
Hepatology. 2021 Oct;74(4):2102-2117. doi: 10.1002/hep.31898.
Induced pluripotent stem cells (iPSCs) provide an important tool for the generation of patient-derived cells, including hepatocyte-like cells, by developmental cues through an endoderm intermediate. However, most iPSC lines fail to differentiate into endoderm, with induction resulting in apoptosis.
To address this issue, we built upon published methods to develop an improved protocol. We discovered that doxycycline dramatically enhances the efficiency of iPSCs to endoderm differentiation by inhibiting apoptosis and promoting proliferation through the protein kinase B pathway. We tested this protocol in >70 iPSC lines, 90% of which consistently formed complete sheets of endoderm. Endoderm generated by our method achieves similar transcriptomic profiles, expression of endoderm protein markers, and the ability to be further differentiated to downstream lineages.
Furthermore, this method achieves a 4-fold increase in endoderm cell number and will accelerate studies of human diseases in vitro and facilitate the expansion of iPSC-derived cells for transplantation studies.
诱导多能干细胞（iPSC）通过发育线索在内胚层中间体的作用下为产生包括肝样细胞在内的患者来源细胞提供了一个重要工具。然而，大多数 iPSC 系未能分化为内胚层，诱导后导致细胞凋亡。
为了解决这个问题，我们在已发表的方法的基础上进行了改进，发现通过蛋白激酶 B 通路抑制细胞凋亡和促进增殖，强力霉素可显著提高 iPSC 向内胚层分化的效率。我们在超过 70 条 iPSC 系中测试了该方案，其中 90%的细胞始终形成完整的内胚层片。我们的方法产生的内胚层具有相似的转录组谱、内胚层蛋白标记物的表达以及进一步分化为下游谱系的能力。
此外，该方法使内胚层细胞数量增加了 4 倍，将加速体外人类疾病的研究，并有助于 iPSC 衍生细胞的扩增，以进行移植研究。
