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Diverse requirements for microglial survival, specification, and function revealed by defined-medium cultures

ABSTRACT: Microglia, the resident macrophages of the central nervous system (CNS), engage in various CNS-specific functions that are critical for development and health. To better study the properties that distinguish microglia from other tissue macrophage populations, we have optimized serum-free culture conditions to permit robust survival of highly purified adult microglia under defined-medium conditions. We find that astrocyte-derived factors prevent microglial death ex vivo and that this activity results from three primary components (CSF-1/IL-34, TGF-b2, and cholesterol). Using microglial cultures that have never been exposed to serum, we demonstrate a dramatic change in intrinsic phagocytic capacity after serum exposure. Finally, we find that mature microglia rapidly lose signature gene expression profiles after isolation, and that this loss can be reversed by engrafting cells back into an intact CNS environment. These data indicate that the specialized gene expression profile of mature microglia require continuous instructive signaling from the intact CNS. This RNA-seq dataset describes changes in rat microglial transcriptome in culture with or without serum exposure.

ORGANISM(S): Rattus norvegicus

PROVIDER: GSE96995 | GEO | 2017/05/11

SECONDARY ACCESSION(S): PRJNA380350

REPOSITORIES: GEO

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Project description:Microglia, the resident immune cells of the central nervous system (CNS), have two distinct phenotypes in the developing brain: amoeboid form, known to be amoeboid microglial cells (AMC) and ramified form, known to be ramified microglial cells (RMC) alongside several intermediate forms. The AMC are characterized by being proliferative, phagocytic and migratory whereas the RMC are quiescent and exhibit a slow turnover rate. The AMC transform into RMC with advancing age, and this transformation is indicative of the gradual shift in the microglial functions. Both AMC and RMC respond to CNS inflammation, and they become hypertrophic when they are activated by trauma, infection or neurodegenerative stimuli. The molecular mechanisms and functional significance of morphological transformation of microglia during normal development and in disease conditions is not clear. It is hypothesized that AMC and RMC are functionally regulated by a specific set of genes encoding various signaling molecules and transcription factors. To address this, we carried out cDNA microarray analysis using lectin-labeled AMC and RMC isolated from frozen tissue sections of the corpus callosum of 5-day and 4-week old rat brain respectively, by laser capture microdissection (LCM). The global gene expression profiles of both microglial phenotypes were compared and the differentially expressed genes in AMC and RMC were clustered based on their functional annotations. This genome wide comparative analysis helps in identifying genes that are specific to AMC and RMC. The novel and specific molecules identified in both microglial phenotypes can be targeted for therapeutic purposes in developing and adult brain diseases. We used microarrays to identify the genes specific to amoeboid and ramified microglia. RNA was isolated from the laser-captured amoeboid and ramified microglia from the corpus callosum of 5-day and 4-week old rat brain. The RNA was hybridised onto Affymetrix Rat 230 2.0 array.

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Diverse requirements for microglial survival, specification, and function revealed by defined-medium cultures

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