Horii Mariko, Bui Tony, Touma Ojeni, Cho Hee Young, Parast Mana M
Department of Pathology, University of California San Diego, La Jolla, California.
Sanford Consortium for Regenerative Medicine, University of California San Diego, La Jolla, California.
Curr Protoc Stem Cell Biol. 2019 Sep;50(1):e96. doi: 10.1002/cpsc.96.
We previously established a two-step protocol for differentiation of human pluripotent stem cells (hPSCs) into trophoblasts, using a StemPro-based minimal medium (EMIM) with bone morphogenetic protein-4 (BMP4). This protocol was suboptimal, resulting in induction of mixed mesoderm and trophoblast markers. Furthermore, adapting hPSCs to StemPro has proven difficult, and prolonged culture in this medium has been shown to promote genomic instability. Therefore, we moved on to the use of new media, including E8, and most recently, StemFlex, for rapid adaptation from feeder to non-feeder conditions. In the new protocol, we have incorporated the WNT inhibitor IWP2 into the first step, resulting in uniform differentiation of hPSCs into cytotrophoblast (CTB)-like cells, without induction of the mesoderm lineage. We also show that, at the end of the second step, there are distinct populations of terminally differentiated multinucleated human chorionic gonadotropin (hCG)-producing syncytiotrophoblast (STB) and HLAG extravillous trophoblast (EVT)-like cells. © 2019 by John Wiley & Sons, Inc.
我们之前建立了一种将人多能干细胞（hPSC）分化为滋养层细胞的两步法方案，使用基于StemPro的基础培养基（EMIM）并添加骨形态发生蛋白-4（BMP4）。该方案并不理想，会诱导中胚层和滋养层标志物的混合表达。此外，事实证明让hPSC适应StemPro很困难，并且在这种培养基中长时间培养已显示会促进基因组不稳定。因此，我们转而使用包括E8以及最近的StemFlex在内的新培养基，以便从饲养层条件快速适应无饲养层条件。在新方案中，我们在第一步中加入了WNT抑制剂IWP2，从而使hPSC均匀分化为细胞滋养层（CTB）样细胞，而不会诱导中胚层谱系。我们还表明，在第二步结束时，存在终末分化的产生人绒毛膜促性腺激素（hCG）的多核合体滋养层（STB）和HLA-G绒毛外滋养层（EVT）样细胞的不同群体。©2019约翰威立国际出版公司。
