Brief Report: Efficient Generation of Hematopoietic Precursors and Progenitors from Human Pluripotent Stem Cell Lines

Abstract

By mimicking embryonic development of the hematopoietic system, we have developed an optimized in vitro differentiation protocol for the generation of precursors of hematopoietic lineages and primitive hematopoietic cells from human embryonic stem cells (ESC) and induced pluripotent stem cells (iPSCs). Factors such as cytokines, extra cellular matrix components, and small molecules as well as the temporal association and concentration of these factors were tested on seven different human ESC and iPSC lines. We report the differentiation of up to 84% human CD45+ cells (average 41% ± 16%, from seven pluripotent lines) from the differentiation culture, including significant numbers of primitive CD45+/CD341 and CD45+/CD341/CD38− hematopoietic progenitors. Moreover, the numbers of hematopoietic progenitor cells generated, as measured by colony forming unit assays, were comparable to numbers obtained from fresh umbilical cord blood mononuclear cell isolates on a per CD45+ cell basis. Our approach demonstrates highly efficient generation of multipotent hematopoietic progenitors with among the highest efficiencies reported to date (CD45+/CD341) using a single standardized differentiation protocol on several human ESC and iPSC lines. Our data add to the cumulating evidence for the existence of an in vitro derived precursor to the hematopoietic stem cell (HSC) with limited engrafting ability in transplanted mice but with multipotent hematopoietic potential. Because this protocol efficiently expands the preblood precursors and hematopoietic progenitors, it is ideal for testing novel factors for the generation and expansion of definitive HSCs with long-term repopulating ability.

Searching for the Key to Expand Hematopoietic Stem Cells

“…At present, multipotent hematopoietic progenitors (short-term HSC) with limited engrafting ability in transplanted mice were obtained (Woods et al, 2011). Other groups reported efficient generation of cells that mostly produce the myeloid lineage following long term engraftment or produce CD34+ hematopoietic precursors that have phenotype similar to adult HSC but might best correspond to the embryonic stage of yolk-sac, aortogonadalmesonephros (AGM), and/or fetal liver stage of hematopoiesis (Melichar et al, 2011;Narayan et al, 2006 andfor review : Tian andKaufman, 2008).…”

“…Differentiation of iPSC into hematopoietic lineage have been achieved using a combination of specific cytokines and growth factors (Sakamoto et al, 2010) and have already demonstrated from both mouse and human iPSC (Lengerke et al, 2009;Niwa et al, 2009;Woods et al, 2011). However, the number of cells obtained in view of therapeutic use is still insufficient and small molecules that might expand the production of hematopoietic cells have yet to be found.…”

The immunogenicity of cells derived from induced pluripotent stem cells

“…While this approach is promising, it remains a challenge to differentiate hESCs into functional hematopoietic stem cells that can efficiently reconstitute the human immune system in vivo. [7][8][9] …”

MicroRNAs Contribute to Induced Pluripotent Stem Cell Somatic Donor Memory

“…Herein we study miRNA donor memory, using as a model the reprogramming of hematopoietic progenitor cells (HPCs). Given that blood progenitors are suitable for reprogramming (15)(16)(17) and that successful differentiation from iPSCs to long term repopulating HPCs has not yet been reported (18,19), we propose that the miRNAs with conserved expression levels between HPCs and their derived LP-iPSCs may play a relevant biological role in determining the ability of these cells to revert toward HPCs in vitro. Applying bioinformatic comparison algorithms on microarray generated data, we detected a set of miRNAs highly expressed in HPCs as well as in the resulting LP-iPSCs, but lost in HP-iPSCs.…”