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Updated: Nov 1, 2025
Synaptic Microcircuit Modeling with 3D Cocultures of Astrocytes and Neurons from Human Pluripotent Stem Cells
Published on: August 16, 2018
Enhanced HSC-like cell generation from mouse pluripotent stem cells in a 3D induction system cocultured with stromal
Wei Shan1,2,3,4, Qin Yu5, Yan Long1,2,3,4
1Bone Marrow Transplantation Center, The First Affiliated Hospital, School of Medicine, Zhejiang University, No.79 Qingchun Road, Hangzhou, Zhejiang, PR China.
Background:
Decades of efforts have attempted to differentiate the pluripotent stem cells (PSCs) into truly functional hematopoietic stem cells (HSCs), yet the problems of low differentiation efficiency in vitro and poor hematopoiesis reconstitution in vivo still exist, mainly attributing to the lack of solid, reproduced, or pursued differentiation system.
Methods:
In this study, we established an in vitro differentiation system yielding in vivo hematopoietic reconstitution hematopoietic cells from mouse PSCs through a 3D induction system followed by coculture with OP9 stromal cells. The in vivo hematopoietic reconstitution potential of c-kit+ cells derived from the mouse PSCs was evaluated via m-NSG transplantation assay. Flow cytometry analysis, RNA-seq, and cell cycle analysis were used to detect the in vitro hematopoietic ability of endothelial protein C receptor (EPCR, CD201) cells generated in our induction system.
Results:
The c-kit+ cells from 3D self-assembling peptide induction system followed by the OP9 coculture system possessed apparently superiority in terms of in vivo repopulating activity than that of 3D induction system followed by the 0.1% gelatin culture. We interestingly found that our 3D+OP9 system enriched a higher percentage of CD201+c-kit+cells that showed more similar HSC-like features such as transcriptome level and CFU formation ability than CD201-c-kit+cells, which have not been reported in the field of mouse PSCs hematopoietic differentiation. Moreover, CD201+ hematopoietic cells remained in a relatively slow cycling state, consistent with high expression levels of P57 and Ccng2. Further, we innovatively demonstrated that notch signaling pathway is responsible for in vitro CD201+ hematopoietic cell induction from mouse PSCs.
Conclusions:
Altogether, our findings lay a foundation for improving the efficiency of hematopoietic differentiation and generating in vivo functional HSC-like cells from mouse PSCs for clinical application.
