Gene expression profile at single cell level of peripheral blood mononuclear cells from Cx3cr1-/- mice and wt mice under C57BL/6J background
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ABSTRACT:
ORGANISM(S): Mus musculus
PROVIDER: GSE282569 | GEO | 2025/02/19
REPOSITORIES: GEO
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Project description:In atherosclerosis progression and regression, monocytes or monocyte-derived macrophages are the major immune cells in the plaque. It is important to understand the fate and characteristics of monocyte/macrophage during the plaque progression and regression. To characterize the fate of monocytes/macrophages, we performed single cell RNA sequencing of fate-mapped aortic CX3CR1-derived monocytes/macrophages from Cx3cr1CreERT2-IRES-YFP/+Rosa26floxed-tdTomato/+ mice with AAV-PCSK9 injection and fed a Western Diet. The single cell RNA-seq analyses revealed the heterogeneity of aortic macrophages and identified a stem-like cell cluster in atherosclerotic aorta.
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Project description:Kidney macrophages are comprised of both monocyte-derived and tissue resident populations; however, the heterogeneity of kidney macrophages and factors that regulate their heterogeneity are poorly understood. Herein, we performed single cell RNA sequencing (scRNAseq), fate mapping, and parabiosis to define the cellular heterogeneity of kidney macrophages in healthy mice. Our data indicate that healthy mouse kidneys contain four major subsets of monocytes and two major subsets of kidney resident macrophages (KRM) including a population with enriched Ccr2 expression, suggesting monocyte origin. Surprisingly, fate mapping data using the newly developed Ms4a3Cre Rosa Stopf/f TdT model indicate that less than 50% of Ccr2+ KRM are derived from Ly6chi monocytes. Instead, we find that Ccr2 expression in KRM reflects their spatial distribution as this cell population is almost exclusively found in the kidney cortex. We also identified Cx3cr1 as a gene that governs cortex specific accumulation of Ccr2+ KRM and show that loss of Ccr2+ KRM reduces the severity of cystic kidney disease in a mouse model where cysts are mainly localized to the kidney cortex. Collectively, our data indicate that Cx3cr1 regulates KRM heterogeneity and niche-specific disease progression.
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Project description:Muscle injury triggers inflammation in which infiltrating mononuclear phagocytes are crucial for tissue regeneration. The interaction of the CCL2/CCR2 and CX3CL1/CX3CR1 chemokine axis that guides phagocyte infiltration is incompletely understood. Here, we show that CX3CR1 deficiency promotes muscle repair and rescues Ccl2-/- mice from impaired muscle regeneration as a result of altered macrophage function, not infiltration. Transcriptomic analysis of muscle mononuclear phagocytes reveals that Apolipoprotein E (ApoE) is up-regulated in mice with efficient regeneration. ApoE treatment enhances phagocytosis by mononuclear phagocytes in vitro, and restores phagocytic activity and muscle regeneration in Ccl2-/- mice. Because CX3CR1 deficiency may compensate for defective CCL2-dependant monocyte recruitment by modulating ApoE-dependent macrophage phagocytic activity, targeting CX3CR1 expressed by macrophages might be a powerful therapeutic approach to improve muscle regeneration.
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Project description:Aim of this project was to examine the global gene expression profiles of mononuclear phagocytes recruited from peripheral blood to the alveolar space following alveolar deposition of the TLR2-ligand Pam3CSK4 in transgenic CX3CR1+/GFP mice. Experiment Overall Design: Alveolar macrophages (AM) and peripheral blood monocytes (PBM) were isolated from broncho-alveolar lavages and from blood, respectively, using FACS. Expression profiles from AM and PBM of the same mice were compared on the same slides. Each labeled RNA sample contained RNA pooled from six individuals. Four pairs of RNA from corresponding AM and PBM pools were hybridized, giving a total amount of 24 individual mice analyzed. Two hybridizations were performed with AM labeled with Cy3 and PBM with Cy5, two hybridizations were performed with swaped dyes.
2009-01-09 | E-GEOD-11978 | biostudies-arrayexpress
Project description:We investigated the frequency of CX3CR1+CD8+ T cells in peripheral blood (PB) before and during immune checkpoint inhibitor (ICI) therapy, and delineated the TCR repertoire in peripheral CX3CR1+CD8+ T-cell subsets and CD8+ tumor-infiltrating lymphocytes (TILs) using preclinical models. To understand the clinical utility of CX3CR1 as a circulating T-cell biomarker, we analyzed longitudinal PB samples from non-small cell lung cancer (NSCLC) patients undergoing anti-PD-1 therapy, and evaluated predictive and prognostic value of changes in the frequency of PB CX3CR1+ CD8+ T cells Methods: Female C57BL/6 mice were inoculated subcutaneously with 5 x 10^5 MC38 tumor cells per mouse on the right flank on day 0. When tumor volume reached approximately 50 mm^3, 200 µg of anti-PD-L1 Ab (clone 10F.9G2, BioXcell) and 100 µg of anti-CTLA-4 Ab (clone 9H10, BioXcell) were administered intraperitoneally every 3 days and every other day, respectively. MC38 tumor-bearing mice were treated for 14 days, and spleens and tumors were harvested. [Tumor cells] Tumors were cut into small pieces of 2-4 mm. Single-cell suspensions were obtained by mechanical dispersion consisting of two 30-min incubations at 37°C, 5% CO2 in 5 ml RPMI 1640 (Gibco) and tumor dissociation kit (Miltenyi Biotec) in C Tubes (Miltenyi Biotec) interspersed with three mechanical dispersions on a GentleMACS dissociator (Miltenyi Biotec). The tumor cell suspensions were then filtered through a cell strainer (70 μm; BD Biosciences). An EasySep Mouse CD8a Positive Selection Kit II (STEMCELL Technologies) was used to isolate murine CD8+ tumor-infiltrating lymphocytes (TILs) for TCR sequencing. [Splenocytes] Spleens were homogenized by forcing the tissue through a cell strainer (70 μm; BD Biosciences). Red blood cells in blood and spleen were lysed using ACK Lysis Buffer (Gibco). For TCR sequencing of murine splenic CD8+ T cells, single cell suspensions from freshly isolated splenocytes were stained. CD45+ CD3+ CD8+ T cells were gated, and CD27lo CX3CR1-, CD27hi CX3CR1-, and CX3CR1+ CD8+ T cells were sorted using BD FACSAria II Cell Sorter. DNA from flow-isolated murine splenic CD8+ T cells and CD8+ TILs, and PB CD8+ T cells was extracted using QIAamp DNA Micro Kit (QIAGEN). DNA was quantified using Qubit dsDNA BR Assay (Invitrogen). Amplification and sequencing of TCRβ CDR3 regions was performed using ImmunoSEQ immune profiling system at the survey level (Adaptive Biotechnologies). Sequencing was performed on an Illumina NextSeq system using 150 cycle mid-output kit (Illumina Inc.). Processed data were uploaded to the ImmunoSEQ Platform (Adaptive Biotechnologies) for bioinformatics analysis. Processed data were downloaded and frequencies/counts for TCR clonotypes and diversity were examined by nucleotide sequences after non-productive reads were filtered out. Results: Clonally expanded TCR repertoires of CD8+ TILs are enriched in the peripheral CX3CR1+ subset during ICI therapy. ICI therapy induces high degree of TCR sequence similarity and clonality between CD8+ TILs and the peripheral CX3CR1+ CD8+ T cells.
2021-01-26 | GSE165383 | GEO
Project description:Distinct subsets of T lymphocytes express CX3CR1 under inflammatory conditions, but little is known about CX3CR1+ CD4+ T cells during Type 2 inflammation in helminth infections. Here, we used a fate-mapping mouse model to characterize CX3CR1+ CD4+ T cells during both acute Nippostrongylus brasiliensis and chronic Schistosoma mansoni helminth infections, revealing CX3CR1+ CD4+ T cells to be an activated tissue homing subset with varying capacity for cytokine production. Tracking these cells over time revealed that maintenance of CX3CR1 itself along with a TH2 phenotype conferred a survival advantage in the inflamed tissue. Single-cell RNA-sequencing analysis of fate-mapped CX3CR1+ CD4+ T cells from both the peripheral tissue and the spleen revealed a considerable level of diversity and identified a distinct population of BCL6+ TCF-1+ PD1+ CD4+ T cells in the spleen during helminth infections. Conditional deletion of BCL6 in CX3CR1+ cells result in fewer CX3CR1+ CD4+ during infection, indicating a role in sustaining CD4+ T cell responses to helminth infections. Overall, our studies revealed the behavior and heterogeneity of CX3CR1+ CD4+ T cells during Type 2 inflammation in helminth infections and identified BCL6 to be important in their maintenance
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2011-12-21 | E-GEOD-27859 | biostudies-arrayexpress
Project description:Scavenger receptors Stabilin-1 (Stab1) and Stabilin-2 (Stab2) are preferentially expressed by liver sinusoidal endothelial cells. They mediate the clearance of circulating plasma molecules controlling distant organ homeostasis. Studies suggest that Stab1 and Stab2 may impact atherosclerosis. Although subsets of tissue macrophages also express Stab1, hematopoietic Stab1 deficiency does not modulate atherogenesis. Here, we comprehensively studied how targeting Stab1 and Stab2 affects atherosclerosis. Methods: ApoE-KO mice were interbred with Stab1-KO and Stab2-KO mice and fed a Western diet (WD). For antibody targeting, Ldlr-KO mice were also used. Plasma proteomics were performed in Stabilin-deficient mice and candidates were independently confirmed. Ligand binding studies comprised GST-pull down and endocytosis assays. Plasma proteome effects on monocytes were studied by single cell RNA sequencing in vivo, and by gene expression analyses of Stabilin-ligand-stimulated and plasma-stimulated bone marrow-derived monocytes/macrophages in vitro. Results: Spontaneous and WD-associated atherogenesis was significantly reduced in ApoE-Stab1- and ApoE-Stab2-KO. Similarly, inhibition of Stab1 or Stab2 by monoclonal antibodies significantly reduced WD-associated atherosclerosis in ApoE-KO and Ldlr-KO. While neither plasma lipid levels nor circulating immune cell numbers were decisively altered, plasma proteomics revealed a switch in the plasma proteome, consisting of 231 dysregulated proteins comparing Wildtype with Stab1/2 single and Stab1/2-double KO, and of 43 proteins comparing ApoE-, ApoE-Stab1- and ApoE-Stab2-KO. Among this broad spectrum of common, but also disparate scavenger receptor ligands, Periostin, Reelin and TGFBi, known to modulate atherosclerosis, were independently confirmed as novel circulating ligands of Stab1/2. scRNA-Seq of circulating myeloid cells of ApoE-, ApoE-Stab1- and ApoE-Stab2-KO showed transcriptomic alterations in patrolling (Ccr2-/Cx3cr1++/Ly6Clo) and inflammatory (Ccr2+/Cx3cr1+/Ly6Chi) monocytes including downregulation of pro-atherogenic transcription factor Egr1. Ligand exposure did not alter Egr1 expression in vitro, but exposure of Wildtype bone marrow-derived monocytes/macrophages to plasma from ApoE-Stab1- and ApoE-Stab2-KO mice showed a reverted pro-atherogenic macrophage activation as compared to ApoE-KO plasma including downregulation of Egr1. Conclusions: Inhibition of Stab1/Stab2 mediates an anti-inflammatory switch in the plasma proteome which includes direct Stabilin ligands. The altered plasma proteome suppresses both patrolling and inflammatory macrophages and, thus, systemically protects against atherogenesis. Altogether, anti-Stab1- and anti-Stab2-targeted therapies provide a novel approach for the future treatment of atherosclerosis.
2025-01-21 | PXD029342 | Pride
Project description:We hypothesize that under homeostatic as well as inflammatory conditions circulating monocytes and/or their bone marrow-derived progenitors might contribute to the replenishment of CD103+ and CD103- DC in lymphoid and non-lymphoid compartments. To that end, bone marrow cells from CX3CR1+/gfp C57BL/6 mice were sorted as follows: lineage negative (CD3, CD19, NK1.1, Ter119, Ly6G and CD11c) CX3CR1+c-kit+. Sorted cells were further cultured in vitro under the continuous presence of GM-CSF. lin-CX3CR1+c-kit+ bone marrow cells gave rise to CD103+ and CD103- DC in vitro. To test whether CD103 might be a suitable marker that allows the differentiation of functionally distinct DC subsets generated in vitro, CD11c+CD103+ and CD11c+CD103- DC were sorted from day 5 cultures of lin-CX3CR1+c-kit+ cells and analyzed by whole mouse genome microarrays from Agilent technologies. Compared to CD103+ DC, CD103- DC displayed a strong up-regulation of transcripts for genes involved in innate immunity, whereas those involved in costimulation were down-modulated. Our data suggest distinct functional activity of these two DC subsets. Keywords: Transcriptional profiling-Cell type comparison We compared the transcriptional profile of CD11c+CD103+ and CD11c+CD103- dendritic cells by use of Agilent Whole Mouse Genome Microarrays. Two sets of samples were generated independently as real biological replicates. The microarray analysis was performed as a dual-color experiment, including a dye-swap.
2008-12-17 | E-GEOD-10882 | biostudies-arrayexpress