Hematopoietic and Endothelial Differentiation of Human Induced Pluripotent Stem Cells

TLDR

Induced pluripotent stem cells enable in‑vitro disease modeling and regenerative therapy with immunologically compatible cells, yet clinical application of iPSC‑derived blood cells remains unresolved, though they are already useful for studying blood disease mechanisms and identifying corrective molecules. This study aimed to characterize and compare the hematopoietic and endothelial differentiation potential of seven human iPSC lines reprogrammed from fetal, neonatal, and adult fibroblasts with five human embryonic stem cell lines using an OP9 co‑culture system. The authors employed OP9 stromal co‑culture to induce hematopoietic and endothelial differentiation of the iPSC and hESC lines, followed by semisolid media with hematopoietic growth factors to assess colony‑forming ability and phenotypic subsets. All seven iPSC lines produced CD34⁺CD43⁺ hematopoietic progenitors and CD31⁺CD43⁻ endothelial cells in OP9 coculture, formed all hematopoietic colony types including GEMM in semisolid media, and yielded phenotypically distinct primitive hematopoietic subsets, with differentiation efficiency varying but overall patterns mirroring those of hESCs.

Abstract

Abstract Induced pluripotent stem cells (iPSCs) provide an unprecedented opportunity for modeling of human diseases in vitro, as well as for developing novel approaches for regenerative therapy based on immunologically compatible cells. In this study, we employed an OP9 differentiation system to characterize the hematopoietic and endothelial differentiation potential of seven human iPSC lines obtained from human fetal, neonatal, and adult fibroblasts through reprogramming with POU5F1, SOX2, NANOG, and LIN28 and compared it with the differentiation potential of five human embryonic stem cell lines (hESC, H1, H7, H9, H13, and H14). Similar to hESCs, all iPSCs generated CD34+CD43+ hematopoietic progenitors and CD31+CD43− endothelial cells in coculture with OP9. When cultured in semisolid media in the presence of hematopoietic growth factors, iPSC-derived primitive blood cells formed all types of hematopoietic colonies, including GEMM colony-forming cells. Human induced pluripotent cells (hiPSCs)-derived CD43+ cells could be separated into the following phenotypically defined subsets of primitive hematopoietic cells: CD43+CD235a+CD41a± (erythro-megakaryopoietic), lin−CD34+CD43+CD45− (multipotent), and lin−CD34+CD43+CD45+ (myeloid-skewed) cells. Although we observed some variations in the efficiency of hematopoietic differentiation between different hiPSCs, the pattern of differentiation was very similar in all seven tested lines obtained through reprogramming of human fetal, neonatal, or adult fibroblasts with three or four genes. Although several issues remain to be resolved before iPSC-derived blood cells can be administered to humans for therapeutic purposes, patient-specific iPSCs can already be used for characterization of mechanisms of blood diseases and for identification of molecules that can correct affected genetic networks.