Differentiation of Human-Induced Pluripotent Stem Cells into Macrophages

Adult somatic cells can be genetically reprogrammed to obtain induced pluripotent stem cells, iPSCs, that exhibit stem cell-like undifferentiated and proliferative states.

To differentiate human iPSCs into macrophages — white blood cells that phagocytose pathogens and cellular debris — begin with a culture plate with a non-cell adhesive surface.

Add media containing the growth factors bone morphogenetic protein, stem cell factor, and vascular endothelial growth factor. Seed the iPSCs and incubate.

The non-adhesive surface prevents iPSC attachment and maintains the cells in suspension, facilitating aggregation into three-dimensional embryoid bodies, EBs. The growth factors initiate iPSC differentiation within the EBs into mesodermal lineage cells.

Transfer the differentiated EBs to a centrifuge tube. Allow them to settle through gravity. Resuspend the EBs in fresh media containing interleukin-3, IL3, and macrophage colony-stimulating factor, CSF1. Plate the EBs on a gelatin-coated cell culture plate.

Gelatin — an extracellular matrix protein — facilitates EB attachment. IL3 stimulates the EBs to differentiate into hematopoietic stem-progenitor cells, giving rise to myeloid cells. CS