In this work we have established a robust protocol that allows the derivation of microglia-like cells from human pluripotent stem cells. These cells are derived in a defined medium that mimics the serum-free environment of the CNS interstitial milieu, supporting electrophysiological maturation of neurons, proliferation and maturation of astrocytes, and differentiation and maintenance of oligodendrocytes. We demonstrated that pMGLs are highly phagocytic and, based on their phenotypic and gene expression profiles, resemble primary fetal and adult human microglia. In particular, they bypass a colony-forming monocytic stage, are initially amoeboid and capable of proliferation, extensive migration and phagocytosis. They eventually settle into a highly ramified morphology, enhanced by co-culture with differentiated neurons and glia. These cells express many of the markers expected to be characteristic of microglial expression phenotypes, such as P2RY12/13, HEXB, GPR34 and TMEM119. Transcriptome sequence analysis shows that they resemble human primary fetal and adult microglia, unlike other macrophages.
