An overlooked subset of Cx3cr1wt/wt microglia in the Cx3cr1CreER-Eyfp/wt mouse has a repopulation advantage over Cx3cr1CreER-Eyfp/wt microglia following microglial depletion
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ABSTRACT:
PROVIDER: PRJNA775631 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR16599306.fastq.gz | Fastqsanger.gz | |||
| SRR16599307.fastq.gz | Fastqsanger.gz | |||
| SRR16599308.fastq.gz | Fastqsanger.gz | |||
| SRR16599309.fastq.gz | Fastqsanger.gz | |||
| SRR16599310.fastq.gz | Fastqsanger.gz |
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Project description:Cx3cr1CreER-Eyfp/wt mice contain a subset of microglia lacking Cre and EYFP expression. These microglial escape Cre-mediated recombination and gain a repopulation advantage following Cre-driven DTA-mediated microglial depletion.
2022-02-01 | GSE186700 | GEO
Project description:BackgroundFluorescent reporter labeling and promoter-driven Cre-recombinant technologies have facilitated cellular investigations of physiological and pathological processes, including the widespread use of the Cx3cr1CreER-Eyfp/wt mouse strain for studies of microglia.MethodsImmunohistochemistry, Flow Cytometry, RNA sequencing and whole-genome sequencing were used to identify the subpopulation of microglia in Cx3cr1CreER-Eyfp/wt mouse brains. Genetically mediated microglia depletion using Cx3cr1CreER-Eyfp/wtRosa26DTA/wt mice and CSF1 receptor inhibitor PLX3397 were used to deplete microglia. Primary microglia proliferation and migration assay were used for in vitro studies.ResultsWe unexpectedly identified a subpopulation of microglia devoid of genetic modification, exhibiting higher Cx3cr1 and CX3CR1 expression than Cx3cr1CreER-Eyfp/wtCre+Eyfp+ microglia in Cx3cr1CreER-Eyfp/wt mouse brains, thus termed Cx3cr1highCre-Eyfp- microglia. This subpopulation constituted less than 1% of all microglia under homeostatic conditions, but after Cre-driven DTA-mediated microglial depletion, Cx3cr1highCre-Eyfp- microglia escaped depletion and proliferated extensively, eventually occupying one-third of the total microglial pool. We further demonstrated that the Cx3cr1highCre-Eyfp- microglia had lost their genetic heterozygosity and become homozygous for wild-type Cx3cr1. Therefore, Cx3cr1highCre-Eyfp- microglia are Cx3cr1wt/wtCre-Eyfp-. Finally, we demonstrated that CX3CL1-CX3CR1 signaling regulates microglial repopulation both in vivo and in vitro.ConclusionsOur results raise a cautionary note regarding the use of Cx3cr1CreER-Eyfp/wt mouse strains, particularly when interpreting the results of fate mapping, and microglial depletion and repopulation studies.
| S-EPMC8783445 | biostudies-literature
Project description:ER71 mutant embryos are nonviable and lack hematopoietic and endothelial lineages. To further define the functional role for ER71 in cell lineage decisions, we generated genetically modified mouse models. We engineered an ER71-EYFP transgenic mouse model by fusing the 3.9 kb ER71 promoter to the EYFP reporter gene. Using FACS and transcriptional profiling, we examined the EYFP+ populations of cells in ER71 mutant and wildtype littermates. In the absence of ER71, we observed an increase in the number of EYFP expressing cells, increased expression of the cardiac molecular program and decreased expression of the hemato-endothelial program compared to the wildtype littermate controls. We have also generated a novel ER71-Cre transgenic mouse model using the same 3.9 kb ER71 promoter. Genetic fate mapping studies revealed that the ER71 expressing cells daughter hematopoietic and endothelial lineages in the wildtype background. In the absence of ER71, these cell populations contributed to alternative mesodermal lineages including the cardiac lineage. To extend these analyses, we used an inducible ES/EB system and observed that ER71 overexpression repressed cardiogenesis. Together, these studies identify ER71 as a critical regulator of mesodermal fate decisions, acting to specify the hematopoietic and endothelial lineages at the expense of cardiac lineages. This enhances our understanding of the mechanisms that govern mesodermal fate decisions early during embryogenesis. 12samples were analyzed, including triplicates of WT; EYFP positive, WT EYFP negative, ER71 MT; EYFP positive and ER71 MT; EYFP negative cells
2011-11-02 | E-GEOD-32223 | biostudies-arrayexpress
Project description:Although most tissue macrophages are embryonically derived it is evident that circulating monocytes can compete for virtually any macrophage niche. Monocytes can thus become long-lived replacements of tissue macrophages that are indistinguishable from their embryonic counterparts, but the factors regulating this process are incompletely understood. To study niche competition in the CNS we depleted microglia with >95% efficiency using CX3CR1CreER/+R26DTA/+ mice and monitored long-term repopulation. The microglial niche was repopulated within weeks by a combination of local microglia proliferation giving rise to CX3CR1+F4/80lowClec12a– microglia, as well as infiltration of CX3CR1+F4/80hiClec12a+ macrophages. Adoptive transfer experiments demonstrated that peripherally-derived macrophages arose directly from Ly6Chi monocytes without contribution from hematopoietic progenitors and was independent of BBB breakdown. After repopulation we sorted microglia and monocyte-derived macrophages and performed transcriptional, epigenetic (DNA methylation) and ex vivo functional profiling. This revealed that Ly6Chi monocytes upregulated microglia gene expression and adopted microglia DNA methylation signatures. However, in contrast to proliferating microglia, which rapidly regained their homeostatic gene signature, monocyte-derived macrophages retained a distinct gene signature associated with antigen presentation, interferon signaling and chemotaxis. This translated into functional changes in monocyte-derived macrophages, as demonstrated by differences in surface marker expression, phagocytosis and cytokine production. Our results demonstrate that monocytes are imprinted by the CNS microenvironment but remain transcriptionally, epigenetically and functionally distinct. This may have implications for neuroinflammatory disease states and direct design of novel therapies.
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Project description:In this study, we aimed to deplete the aging microglia and investigate the impact of the newly-born microglia in Alzheimer's pathology using the 3xTg mouse model. Specifically, >24 mo male 3xTg mice were given chow containing 1200mg/kg PLX5622 (AIN-76A-D1001i, Research Diests, NJ, USA) for 2 weeks to deplete their microglia. Control diet with the same base formula were given to control group and to experimental groups during a 4-week repopulation phase immediately following PLX treatment. At the end of the repopulation, we pooled sorted, viable microglia (CD45int CD11b) from 3-5 mice/group and ran single-cell RNAseq analysis to determine the similarities and differences between the aged and repopulated cells in the context of Alzheimer's Disease. We also included non-transgenic controls as a separate group.
2022-02-01 | GSE190607 | GEO
Project description:During early embryogenesis microglia arise from yolk sac progenitors populating the developing CNS, where they are maintained as tissue-resident macrophages throughout the organism’s lifespan. Here, we describe an experimental system that allows the specific conditional ablation of microglia in vivo. Strikingly, we found that the microglia compartment was reconstituted within one week following depletion. Microglia repopulation relied entirely on a CNS-resident, internal pool and was independent from bone marrow-derived precursors. Newly formed microglia were found in highly proliferative, organized micro-clusters that dissolve once steady state was achieved. Gene expression profiling revealed prominent expression of Interleukin-1 (IL-1) receptor in proliferating microglia. During the repopulation phase, IL-1 signaling was neutralized by treatment with IL-1 receptor antagonist that impaired microglia proliferation. Hence, microglia harbor a highly efficient potential to restore themselves without contribution of peripheral myeloid cells. IL-1 signaling significantly participates in this restorative proliferation process and is involved in stabilizing microglia maintenance. bone marrow macrophages, wild type microglia, and repopulating microglia
2015-07-15 | E-GEOD-68376 | biostudies-arrayexpress
Project description:Over the past decade, genetic evidence has demonstrated that microglial dysregulation is likely to play a central role in the development of Alzheimer's disease (AD). As resident immune cells in the brain, microglia become dystrophic and senescent during the chronic progression of AD. To explore whether replenishing the brain with new microglia is beneficial to AD, we employed a CSF1R inhibitor PLX3397 to deplete microglia and induce repopulation after the inhibitor withdrawal in 5xFAD transgenic mice. We observed that microglial repopulation ameliorates AD-associated cognitive deficits, accompanied by elevation of synaptic proteins and hippocampal long-term potentiation (LTP). In addition, microglial morphology is restored after microglial self-renewal, and amyloid pathology is reduced with long-term repopulation but not short-term. Transcriptome analysis showed that repopulating microglia in 5xFAD mice recovers a gene expression profile that is highly similar to microglia from WT mice. Notably, the neurotrophic signaling pathway and hippocampal neurogenesis dysregulated in the AD brain are restored after microglial replenishment. At last, we confirmed that microglial repopulation rescues brain-derived neurotrophic factor (BDNF) expression to contribute to synaptic plasticity. Together, we conclude that microglial self-renewal benefits AD brain by restoring the BDNF neurotrophic signaling pathway. Thus, the proper replenishment of microglia may be an effective and novel therapeutic strategy for ameliorating cognition impairment in AD.
2023-07-24 | GSE223156 | GEO
Project description:RNA-Seq transcriptome comparison of the following cell populations (n=3-4 independent samples per cell population): a) CD11c-eYFP+ cells FACS sorted from brain of female adult mice 4 days after cerebral ischemia, b) CD11c-eYFP+ cells FACS sorted from brain of female parabiotic mice 4 days after cerebral ischemia c) CX3CR1+ microglia sorted from the ischemic brain of female CX3CR1CreERT2-ROSA26 tdTomato mice. Purpose: The goal of this study is to compare the transcriptome profile (RNA-Seq) of infiltrating cD11c-eYFP+ cells and microglia, both collected from ischemic brains of mice. Methods: RNA samples were obtained from FACS sorted eYFP+ cells of the ipsilateral brain hemisphere of CD11c-eYFP mice 4 days post-ischemia, the ipsilateral brain hemisphere of CD11c-eYFP/WT parabiotic mice 4 days post-ischemia, and from microglial cells sorted from the ipsilateral brain hemisphere of Cx3cr1CreERT2:ROSA26dTomato mice 4 days post-ischemia. NGS was performed (RNA-Seq) to compare the transcriptome of these populations. Results: the populations we compared clearly separated the differentially expressed genes in an unsupervised cluster analysis. 1509 genes were overrepresented in microglia and 1183 genes were overrepresented in CD11c-eYFP+ cells in the ischemic brain. Conclusions: Our study is the first comparative analysis of the transcriptomes of microglia and the infiltrating CD11c-eYFP+ cells derived from the ischemic brain of mice. The results show that the infiltrating CD11c-eYFP cell population in the ischemic brain tissue of parabiotic mice displays overrepresentation of genes typical of dendritic cells, immune functions, and ClassII antigen presentation, amongst others, that are not found represented in microglia.
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Project description:To determine the transcriptomic changes underlying the MIA microglia phenotype and their reversal by PLX, we performed RNA-sequencing of magnetic CD11B beads-isolated microglia from the whole brain in Saline and MIA male and female offspring at E17, P7, P20, and P60, as well as from MG-REP male and female offspring at P60. Microglial transcriptome reveals novel MIA and repopulation modules and overlap of MIA microglial genes with ASD gene network. 14,225 microglial genes from RNA-seq data revealed distinct transcriptional signatures of immature microglia (IM module, 4681 transcripts, enriched in E17 and P7 Saline), MIA immature microglia (MIA-IM module, 2816 transcripts, enriched in E17 and P7 MIA), Juvenile microglia (JM module, 2318 transcripts, enriched in P20 Saline and MIA), Adult microglia (AM module, 3276 transcripts, enriched in P60 Saline + CTRL, P60 MIA + CTRL and P60 MIA + MG-REP), and repopulated adult microglia (REP-AM module, 1,134 transcripts, enriched in P60 Saline + MG-REP)
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