Charting the transition from in vitro gliogenesis to the in vivo maturation of human glial progenitor cells transplanted into the hypomyelinated mouse brain

Neither rodent models nor in vitro studies of human cells adequately reflect the molecular regulation of human glial progenitor cell (hGPC) expansion, differentiation, and myelination in vivo. To better understand this process, we used scRNA-seq of >60,000 human cells to track the differentiation of hGPCs from pluripotent stem cells (PSCs), both in vitro and in vivo. In vitro, the hGPC pool comprised 4 transcriptionally-distinct subpopulations, the heterogeneity of which was associated with histone modifications of stage-dependent genes. After the neonatal transplant of these progenitor cells into immunodeficient and myelin-deficient shiverer mice, they differentiated further; when extracted back from their host brains, scRNA-Seq revealed that the donor cells were comprised solely of hGPCs, astrocytes and oligodendrocytes. A combination of gene co-expression, motif enrichment, cell-trajectory, and cell-cell interaction analyses revealed that the tissue environment potentiated the further, context-dependent differentiation of hGPCs, via ligand-mediated activation of gene regulatory networks. Together, these data chart the process by which manufactured human GPCs differentiate into mature astrocytes and oligodendrocytes in vivo, while establishing the transcriptional and phenotypic stability of these cells following transplantation.