Efficient commitment to functional CD34+ progenitor cells from human bone marrow mesenchymal stem-cell-derived induced pluripotent stem cells

PLoS One. 2012;7(4):e34321. doi: 10.1371/journal.pone.0034321. Epub 2012 Apr 9.

Abstract

The efficient commitment of a specialized cell type from induced pluripotent stem cells (iPSCs) without contamination from unknown substances is crucial to their use in clinical applications. Here, we propose that CD34+ progenitor cells, which retain hematopoietic and endothelial cell potential, could be efficiently obtained from iPSCs derived from human bone marrow mesenchymal stem cells (hBMMSC-iPSCs) with defined factors. By treatment with a cocktail containing mesodermal, hematopoietic, and endothelial inducers (BMP4, SCF, and VEGF, respectively) for 5 days, hBMMSC-iPSCs expressed the mesodermal transcription factors Brachyury and GATA-2 at higher levels than untreated groups (P<0.05). After culturing with another hematopoietic and endothelial inducer cocktail, including SCF, Flt3L, VEGF and IL-3, for an additional 7-9 days, CD34+ progenitor cells, which were undetectable in the initial iPSC cultures, reached nearly 20% of the total culture. This was greater than the relative number of progenitor cells produced from human-skin-fibroblast-derived iPSCs (hFib-iPSCs) or from the spontaneous differentiation groups (P<0.05), as assessed by flow cytometry analysis. These induced cells expressed hematopoietic transcription factors TAL-1 and GATA-2 [corrected]. They developed into various hematopoietic colonies when exposed to semisolid media with hematopoietic cytokines such as EPO and G-CSF. Hematopoietic cell lineages were identified by phenotype analysis with Wright-Giemsa staining. The endothelial potential of the cells was also verified by the confirmation of the formation of vascular tube-like structures and the expression of endothelial-specific markers CD31 and VE-CADHERIN. Efficient induction of CD34+ progenitor cells, which retain hematopoietic and endothelial cell potential with defined factors, provides an opportunity to obtain patient-specific cells for iPSC therapy and a useful model for the study of the mechanisms of hematopoiesis and drug screening.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Antigens, CD34 / metabolism*
  • Blotting, Western
  • Bone Marrow / metabolism*
  • Cell Differentiation
  • Cells, Cultured
  • Colony-Forming Units Assay
  • Endothelial Cells / cytology*
  • Endothelial Cells / metabolism
  • Enzyme-Linked Immunosorbent Assay
  • Fibroblasts / cytology*
  • Fibroblasts / metabolism
  • Flow Cytometry
  • Fluorescent Antibody Technique
  • Genes, myc / genetics
  • Granulocyte Colony-Stimulating Factor
  • Growth Substances / pharmacology
  • Hematopoiesis / physiology
  • Hematopoietic Stem Cells / cytology*
  • Hematopoietic Stem Cells / metabolism
  • Humans
  • Induced Pluripotent Stem Cells / cytology*
  • Induced Pluripotent Stem Cells / metabolism
  • Kruppel-Like Factor 4
  • Kruppel-Like Transcription Factors / genetics
  • Kruppel-Like Transcription Factors / metabolism
  • Mesenchymal Stem Cells / cytology*
  • Mesenchymal Stem Cells / metabolism
  • Octamer Transcription Factor-3 / genetics
  • Octamer Transcription Factor-3 / metabolism
  • RNA, Messenger / genetics
  • Real-Time Polymerase Chain Reaction
  • Reverse Transcriptase Polymerase Chain Reaction
  • SOXB1 Transcription Factors / genetics
  • SOXB1 Transcription Factors / metabolism
  • Skin / cytology*
  • Skin / metabolism

Substances

  • Antigens, CD34
  • Growth Substances
  • Kruppel-Like Factor 4
  • Kruppel-Like Transcription Factors
  • Octamer Transcription Factor-3
  • POU5F1 protein, human
  • RNA, Messenger
  • SOX2 protein, human
  • SOXB1 Transcription Factors
  • Granulocyte Colony-Stimulating Factor