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      Directed Differentiation of Human Pluripotent Stem Cells to Microglia

      brief-report

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          There is no author summary for this article yet. Authors can add summaries to their articles on ScienceOpen to make them more accessible to a non-specialist audience.

          Summary

          Microglia, the immune cells of the brain, are crucial to proper development and maintenance of the CNS, and their involvement in numerous neurological disorders is increasingly being recognized. To improve our understanding of human microglial biology, we devised a chemically defined protocol to generate human microglia from pluripotent stem cells. Myeloid progenitors expressing CD14/CX3CR1 were generated within 30 days of differentiation from both embryonic and induced pluripotent stem cells (iPSCs). Further differentiation of the progenitors resulted in ramified microglia with highly motile processes, expressing typical microglial markers. Analyses of gene expression and cytokine release showed close similarities between iPSC-derived (iPSC-MG) and human primary microglia as well as clear distinctions from macrophages. iPSC-MG were able to phagocytose and responded to ADP by producing intracellular Ca 2+ transients, whereas macrophages lacked such response. The differentiation protocol was highly reproducible across several pluripotent stem cell lines.

          Highlights

          • •

            Efficient protocol to generate human microglia from PSCs

          • •

            iPSC-derived microglia have ramified morphology and motile processes

          • •

            Expression and cytokine profiles of iPSC-derived microglia resemble primary microglia

          • •

            iPSC-derived microglia can phagocytose beads and respond to ADP

          Abstract

          Douvaras and colleagues generated microglia from pluripotent stem cells using a chemically defined protocol through a myeloid progenitor. iPSC-derived microglia showed highly motile processes, were able to phagocytose and responded to ADP with calcium transients. Microglial identity was further confirmed by gene expression analysis comparing multiple iPSC-derived microglia samples with primary microglia and peripheral blood or other tissue-specific macrophages.

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          The Microglial Sensome Revealed by Direct RNA Sequencing

          Suzanne E Hickman, Nathan D. Kingery, Toshiro K Ohsumi … (2013)
          Microglia, the principal neuroimmune sentinels of the brain, continuously sense changes in their environment and respond to invading pathogens, toxins and cellular debris. Microglia exhibit plasticity and can assume neurotoxic or neuroprotective priming states that determine their responses to danger. We used direct RNA sequencing, without amplification or cDNA synthesis, to determine the quantitative transcriptomes of microglia of healthy adult and aged mice. We validated our findings by fluorescent dual in-situ hybridization, unbiased proteomic analysis and quantitative PCR. We report here that microglia have a distinct transcriptomic signature and express a unique cluster of transcripts encoding proteins for sensing endogenous ligands and microbes that we term the “sensome”. With aging, sensome transcripts for endogenous ligand recognition are downregulated, whereas those involved in microbe recognition and host defense are upregulated. In addition, aging is associated with an overall increase in expression of microglial genes involved in neuroprotection.
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            Local self-renewal can sustain CNS microglia maintenance and function throughout adult life.

            Bahareh Ajami, Jami Bennett, Charles Krieger … (2007)
            Microgliosis is a common response to multiple types of damage in the CNS. However, the origin of the cells involved in this process is still controversial and the relative importance of local expansion versus recruitment of microglia progenitors from the bloodstream is unclear. Here, we investigated the origin of microglia using chimeric animals obtained by parabiosis. We found no evidence of microglia progenitor recruitment from the circulation in denervation or CNS neurodegenerative disease, suggesting that maintenance and local expansion of microglia are solely dependent on the self-renewal of CNS resident cells in these models.
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              The P2Y12 receptor regulates microglial activation by extracellular nucleotides.

              Sharon E Haynes, Gunther Hollopeter, Guang Yang … (2006)
              Microglia are primary immune sentinels of the CNS. Following injury, these cells migrate or extend processes toward sites of tissue damage. CNS injury is accompanied by release of nucleotides, serving as signals for microglial activation or chemotaxis. Microglia express several purinoceptors, including a G(i)-coupled subtype that has been implicated in ATP- and ADP-mediated migration in vitro. Here we show that microglia from mice lacking G(i)-coupled P2Y(12) receptors exhibit normal baseline motility but are unable to polarize, migrate or extend processes toward nucleotides in vitro or in vivo. Microglia in P2ry(12)(-/-) mice show significantly diminished directional branch extension toward sites of cortical damage in the living mouse. Moreover, P2Y(12) expression is robust in the 'resting' state, but dramatically reduced after microglial activation. These results imply that P2Y(12) is a primary site at which nucleotides act to induce microglial chemotaxis at early stages of the response to local CNS injury.
              Article 0 comments Cited 484 times – based on 0 reviews     Review now
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                Author and article information

                Contributors
                Panagiotis Douvaras
                Journal
                Journal ID (nlm-ta): Stem Cell Reports
                Journal ID (iso-abbrev): Stem Cell Reports
                Title: Stem Cell Reports
                Publisher: Elsevier
                ISSN (Electronic): 2213-6711
                Publication date PMC-release: 18 May 2017
                Publication date Collection: 06 June 2017
                Publication date (Electronic): 18 May 2017
                Volume: 8
                Issue: 6
                Pages: 1516-1524
                Affiliations
                [1 ]The New York Stem Cell Foundation Research Institute, New York, NY 10019, USA
                [2 ]Department of Genetics and Genomic Sciences, Icahn Institute for Genomics and Multiscale Biology, Icahn School of Medicine at Mount Sinai, New York, NY 10029, USA
                [3 ]Department of Neurology, Department of Psychiatry, Alzheimer's Disease Research Center, Icahn School of Medicine at Mount Sinai, New York, NY 10029, USA
                [4 ]The Joint Department of Biomedical Engineering, North Carolina State University and University of North Carolina-Chapel Hill, Raleigh, NC 27695, USA
                Author notes
                [∗ ]Corresponding author pdouvaras@ 123456nyscf.org
                [5]

                Senior author

                Article
                Publisher ID: S2213-6711(17)30177-7
                DOI: 10.1016/j.stemcr.2017.04.023
                PMC ID: 5470097
                PubMed ID: 28528700
                SO-VID: 1984afd0-1250-4d5e-a105-cd7682508360
                Copyright © © 2017 The Author(s)
                License:

                This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).

                History
                Date received : 1 September 2016
                Date revision received : 20 April 2017
                Date accepted : 20 April 2017
                Categories
                Subject: Report

                Keywords: human pluripotent stem cells,microglial differentiation,human microglia
                Data availability:
                Keywords: human pluripotent stem cells, microglial differentiation, human microglia

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