A Highly Efficient Human Pluripotent Stem Cell Microglia Model Displays a Neuronal-Co-culture-Specific Expression Profile and Inflammatory Response

Abstract

SummaryMicroglia are increasingly implicated in brain pathology, particularly neurodegenerative disease, with many genes implicated in Alzheimer's, Parkinson's, and motor neuron disease expressed in microglia. There is, therefore, a need for authentic, efficient in vitro models to study human microglial pathological mechanisms. Microglia originate from the yolk sac as MYB-independent macrophages, migrating into the developing brain to complete differentiation. Here, we recapitulate microglial ontogeny by highly efficient differentiation of embryonic MYB-independent iPSC-derived macrophages then co-culture them with iPSC-derived cortical neurons. Co-cultures retain neuronal maturity and functionality for many weeks. Co-culture microglia express key microglia-specific markers and neurodegenerative disease-relevant genes, develop highly dynamic ramifications, and are phagocytic. Upon activation they become more ameboid, releasing multiple microglia-relevant cytokines. Importantly, co-culture microglia downregulate pathogen-response pathways, upregulate homeostatic function pathways, and promote a more anti-inflammatory and pro-remodeling cytokine response than corresponding monocultures, demonstrating that co-cultures are preferable for modeling authentic microglial physiology.

Human Induced Pluripotent Stem Cell–Derived Macrophages for Unraveling Human Macrophage Biology

“…20 We also have performed RNA-sequencing of myeloid progenitors (CD34 + CD43 + CD45 + CD18 + ) harvested during progressive commitment and maturation to IPSDM 8 and showed that the expression profile of those progenitors is highly correlated to that of IPSDM but not CD14 + monocytes (Figure 2). Thus, successful differentiation of circulating monocytes will rely on a more complete understanding of mechanisms regulating definitive hematopoietic specification 24 and identification of factors governing the generation of HSCs from iPSC, 25 knowledge gaps that remain major goals of regenerative medicine.…”

“…Thus, IPSDM may developmentally relate to and be a good model for MYB -independent tissue resident macrophages. Indeed, several iPSC to microglia-like cell differentiation protocols were published this year using cytokines representing the main driver of microglia lineage, 18, 19 co-culture with iPSC-derived neurons 20, 21 and astrocytes, 22 or incubating with cytokines derived from those cell types 23 to recapitulate organ-specific cues. Takata et al has established a protocol that specifically resembles primitive hematopoiesis and YS macrophages for further differentiation into tissue macrophage-like cells by co-culture or engraftment.…”

Advances and Current Challenges Associated with the Use of Human Induced Pluripotent Stem Cells in Modeling Neurodegenerative Disease

“…Activated hiPSC-derived microglia, cocultured with hiPSC-derived cortical neurons, have been shown to exhibit more physiologically correct behaviors than monoculture controls. These behaviors include the downregulation of pathogen response pathways and a more anti-inflammatory cytokine-based response [Haenseler et al, 2017]. Methods for producing myelinating oligodendrocytes from hiPSC are also available [Wang et al, 2013;Ehrlich et al, 2017].…”

“…Although cocultures of hiPSC-derived neurons with microglia have been reported [Haenseler et al, 2017], their ability to faithfully recapitulate inflammatory responses within in vitro models has yet to be demonstrated. Neuroinflammation is a hallmark of AD and neurodegenerative diseases in general [Heneka et al, 2015;Walters et al, 2016].…”

Activated Microglia in the Brain: Mitochondrial and Cell Membrane-Associated Targets for Positron Emission Tomography

“…Investigations of typical microglial cells are limited in vitro, because the source of this cell population in the brain is, as widely accepted, the yolk sac releasing a wave of primitive macrophages as progenitor cells of microglia during the early embryonal development [21] [22] [23] [24]. Microglial population colonizes the whole CNS and maintains itself by local proliferation [23] [24].…”