RED BLOOD CELL GENERATION FROM HUMAN INDUCED PLURIPOTENT STEM CELLS OF DIFFERENT DONOR CELL TYPE ORIGIN

Abstract

Manufacturing red blood cells (RBC) from human induced pluripotent stem cells (iPS cells) offers the potential to produce large quantities of patients' specific RBC for transfusions purposes. Epigenetic memory in iPS cells in regard to their donor cell type of origin might lead to variations in their differentiation capacities. We have generated iPS cells from human cord blood CD34+ hematopoietic stem cells (HSC) (CD34-iPS) and neural stem cells (NSC-iPS) and evaluated their differentiation potential into hematopoietic precursor and mature RBC. For hematopoietic induction, iPS cells were allowed to form embryoid bodies (EBs) under cytokine stimulation for 21 days. Thereafter, dissociated single cells were applied to a three-step protocol for human erythropoiesis for additional 18-25 days. We have found a similar hematopoietic induction potential among our cell lines. After EB dissociation on day 21, hematopoietic commitment, measured by CD43 expression, was about 20% for all cell lines. Colony-forming unit assays demonstrate a similar distribution of myeloid (CFU-M/CFU-GM), erythroid (BFU-E/CFU-E) and mixed (CFU-GEMM) colonies. Hematopoietic cells further developed into erythroid precursors as determined by >90% expression of glycophorin A, followed by maturation into normoblasts and partially enucleated RBC. All human iPS derived erythrocytes predominantly present fetal hemoglobin (>85%), some embryonic and only a minor amount of adult hemoglobin. In summary, we were able to recapitulate the development of RBC from all human iPS cell lines evaluated. In addition, our data hint at a similar erythrocyte induction potential of iPS cells, independent of their donor cell type origin.