biostudies-arrayexpressCarlos RelañoMus musculushttps://www.ebi.ac.uk/biostudies/studies/E-MTAB-14540We dissected the changes in the infiltrating T cell population in the aorta of cDC1-impaired mice compared with control mice in atheroprone Ldlr-/- mice by scRNAseq. To this aim, lethally irradiated Ldlr-/- CD45.1 mice were grafted with cDC1-impaired Xcr1Cre-DTA or control Xcr1Cre bone marrow (BM) and fed high cholesterol diet (HCD) for 8 weeks. At the endpoint, 10 mice per group were pooled and aortic CD45+ CD3+ CD11c- I-A/I-E– F4/80- Ly6C- Ly6G- NK1.1- were sorted and sequenced. A number of 6259 CD3+ T cells were sequenced in Xcr1Cre control chimeric mice, while 4072 cells were sequenced in cDC1-impaired Xcr1Cre-DTA chimeric mice. Nine different clusters were identified based on the following gene expression profile.biostudies-arrayexpressNucleic Acid Extraction - Cells were counted, and their viability was checked using the Countess III cell counter (Thermofisher). Each cell suspension was loaded into one port of a Chromium Next GEM Chip G (10x Genomics). Single cells were encapsulated into emulsion droplets using the Chromium Controller (10x Genomics).Sample Collection - Chimeric Ldr-/- CD45.1 mice were grafted with BM from Xcr1Cre-DTA or control Xcr1Cre mice. After BM reconstitution, these chimeric mice were fed HCD for 8 weeks and aortae were collected. The aortas were kept in cold Dulbecco's Modified Eagle Medium (DMEM, Gibco) before being cut into smaller pieces that were incubated in digestion buffer (Collagenase A (25 mg/ml; Sigma-Aldrich), Dispase II (25 mg/ml; Sigma-Aldrich), DNase I (250 μg/ml), elastase (25 μg/ml), and Liberase TL (0.2 units/mL; Wünsch) for 30 min at 37ºC in a water bath. The digested tissue was mixed by pipetting, filtered through a 70 μm strainer and centrifuged at 400g for 5 min at 4ºC.To achieve sufficient cell numbers, 10 aortae per genotype were pooled and CD3+ T cells were sorted. Briefly, the pool of aortae was pre-purified using MACS® Cell Separation CD45 MicroBeads (Miltenyi Biotec). The CD45+ fraction was collected, and cells were stained in FACS Buffer with anti-CD16/32 and antibody cocktail containing CD45-BV421 (Biolegend #103133), CD11c-FITC (Biolegend #117305), NK1.1-FITC (Biolegend #108705), I-A/I-E-FITC, Ly6C-FITC, Ly6G-FITC, F480-FITC, B220-FITC (Biolegend #103205), CD3-PECy7 (Biolegend #100219) and DAPI as a live/dead marker. CD45+ CD3+ CD11c- I-A/I-E–F480-Ly6C-Ly6G-NK1.1- cells were sorted using FACSAria™ Fusion Cell Sorter (Becton Dickinson).Sequencing - Individual libraries were diluted to 10 nM and pooled for sequencing. Library pool was loaded at 700 pM onto a P3 flow cell (100 cycles) of the NextSeq 2000 (Illumina) in paired-end configuration (28bp Read1, 10bp Index1, 10pb Index 2 and 90bp Read2). FastQ files for each sample were obtained using cell ranger mkfastq pipeline (10x Genomics).Library Construction - Sequencing libraries were prepared using the Chromium Next GEM Single Cell 3' Kit v3.1 (10x Genomics) following the manufacturer instructions. The average size of each library was then calculated using a High sensitivity DNA chip on a 2100 Bioanalyzer (Agilent Technologies) and the concentration was determined using the Qubit fluorometer (Thermofisher)OrganizationMINSEQE ScoreAssays and DataProcessed DataMAGE-TAB FilesData Transformation - Single-cell filtering and clustering were performed using the Scater and Seurat (v4) R packages. Data was normalized using Seurat's NormalizeData function on default options.MetabolomicsUnknownTranscriptomicsGenomicsProteomicsNextSeq 2000RNA-seq of coding RNA from single cellsMus musculusMiguel GalánCarlos RelañofalseSingle-cell RNA-seq of aortic CD3+ T cellsWe dissected the changes in the infiltrating T cell population in the aorta of cDC1-impaired mice compared with control mice in atheroprone Ldlr-/- mice by scRNAseq. To this aim, lethally irradiated Ldlr-/- CD45.1 mice were grafted with cDC1-impaired Xcr1Cre-DTA or control Xcr1Cre bone marrow (BM) and fed high cholesterol diet (HCD) for 8 weeks. At the endpoint, 10 mice per group were pooled and aortic CD45+ CD3+ CD11c- I-A/I-E– F4/80- Ly6C- Ly6G- NK1.1- were sorted and sequenced. A number of 6259 CD3+ T cells were sequenced in Xcr1Cre control chimeric mice, while 4072 cells were sequenced in cDC1-impaired Xcr1Cre-DTA chimeric mice. Nine different clusters were identified based on the following gene expression profile.2025-06-18T00:00:00Z2025-06-18T11:01:04.677Z2024-10-15T13:31:30.865ZE-MTAB-14540ERP165153EFO_0002944EFO_0004170EFO_0005684EFO_0005518EFO_0003816EFO_0004184