{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,7,24]],"date-time":"2026-07-24T06:02:45Z","timestamp":1784872965719,"version":"3.55.0"},"reference-count":34,"publisher":"Frontiers Media SA","license":[{"start":{"date-parts":[[2026,7,24]],"date-time":"2026-07-24T00:00:00Z","timestamp":1784851200000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/100032850","name":"EU4Health","doi-asserted-by":"publisher","award":["101102733"],"award-info":[{"award-number":["101102733"]}],"id":[{"id":"10.13039\/100032850","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/100019180","name":"HORIZON EUROPE European Research Council","doi-asserted-by":"publisher","award":["PE00000007, INF-ACT"],"award-info":[{"award-number":["PE00000007, INF-ACT"]}],"id":[{"id":"10.13039\/100019180","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/501100004008","name":"Istituto Superiore di Sanit\u00e1","doi-asserted-by":"publisher","id":[{"id":"10.13039\/501100004008","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":["frontiersin.org"],"crossmark-restriction":true},"short-container-title":["Front. Immunol."],"abstract":"<jats:sec>\n                    <jats:title>Introduction<\/jats:title>\n                    <jats:p>The dialogue between the immune and nervous systems in the central nervous system (CNS) is a fundamental challenge in neuroscience and neuroimmunology. Microglia, the brain's resident immune cells, continuously communicate with neurons to maintain brain homeostasis, support development, and orchestrate responses to injury.<\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Methods<\/jats:title>\n                    <jats:p>We adapted the CEBPA\/SPI1 transcription factor overexpression approach to generate induced microglia-like cells (iMG) from human embryonic stem cells (hESCs), establishing a reproducible and standardized baseline framework for neuron-microglia co-culture. iMG characterization included assessment of key microglial markers (GPR34, MMP9, TMEM119, IBA1, P2Y12), phagocytic capability benchmarked against the human microglial cell line HMC3, and high-resolution scanning electron microscopy (SEM). iMGs were then co-cultured with hESC-derived NGN2-induced neurons (iNeu), with systematic optimization of seeding ratios and temporal parameters.<\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Results<\/jats:title>\n                    <jats:p>iMGs expressed key microglial markers and demonstrated phagocytic capabilities comparable to HMC3 cells. In co-culture, iMGs exhibited remarkable phenotypic plasticity, with upregulation of both homeostatic and activation-associated microglial genes. Reciprocally, neurons showed increased expression of maturation and synaptic markers, indicating a bidirectional crosstalk. SEM revealed spatially variable iMG morphology \u2014 transitioning from ramified to amoeboid forms depending on proximity to neuronal structures.<\/jats:p>\n                  <\/jats:sec>\n                  <jats:sec>\n                    <jats:title>Discussion<\/jats:title>\n                    <jats:p>\n                      This contact-based\n                      <jats:italic>in vitro<\/jats:italic>\n                      system provides a versatile platform for investigating neuroimmune interactions, designed to be further integrated with additional CNS cell types, and applicable to the study of microglial activation, neuron-microglia signalling, and the impact of infections on CNS homeostasis.\n                    <\/jats:p>\n                  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