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Updated: Mar 30, 2026
Production and Characterization of Human Macrophages from Pluripotent Stem Cells
Published on: April 16, 2020
Genome-Edited Human Pluripotent Stem Cell-Derived Macrophages as a Model of Reverse Cholesterol Transport--Brief
Rajat M Gupta1, Torsten B Meissner1, Chad A Cowan
1From the Department of Stem Cell and Regenerative Biology, Harvard University, and Harvard Stem Cell Institute, Cambridge, MA (R.M.G., T.B.M., C.A.C., K.M.); and Division of Cardiovascular Medicine, Brigham and Women's Hospital, Boston, MA (R.M.G., K.M.).
Objective:
To create isogenic human pluripotent stem cell-derived macrophages with and without ABCA1 expression as a model for reverse cholesterol transport.
Approach And Results:
The clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated 9 (Cas9) genome-editing system was used to introduce frameshift mutations into the coding sequence of ATP-binding cassette, subfamily A, member 1. Individual human pluripotent stem cell clones with deleterious mutations were identified, expanded, and differentiated into mature macrophages with a cytokine-based, feeder-free differentiation protocol. Wild-type cells demonstrated effective cholesterol efflux to apoAI acceptor, whereas ABCA1(-/-) cells displayed significantly reduced efflux ability and increased expression of proinflammatory cytokines.
Conclusions:
Human pluripotent stem cell-derived macrophages capable of reverse cholesterol transport can be rapidly generated and genetically edited with CRISPR/Cas9. Introduction of homozygous frameshift mutations results in loss of ABCA1 expression in differentiated macrophages and subsequent reduction of cholesterol efflux capability. This facile genome-editing approach and differentiation protocol pave the way for future studies of the molecular determinants of reverse cholesterol transport and other macrophage properties.
