biostudies-arrayexpressJan KubovčiakMus musculus10x cellranger v6.1.1https://www.ebi.ac.uk/biostudies/studies/E-MTAB-14958Ischemic stroke is a severe medical condition that leads to neurological impairments, including the loss of sensory, motor, and cognitive functions. Focal cerebral ischemia (FCI) occurs when blood supply to a specific brain region is interrupted, resulting in cell death. Cells expressing neural-glial antigen 2 (NG2) include glial cells, primarily oligodendrocyte precursors, and perivascular cells. In both cell types, the canonical Wnt signaling pathway is active, with its activity increasing after ischemic injury, influencing the tissue’s response to damage. Following ischemia, NG2 glia rapidly proliferate, migrate to the injury site, contribute to glial scar formation alongside astrocytes, and play a role in brain tissue regeneration. Their potential to differentiate into cell types beyond oligodendrocytes has been documented. Perivascular cells, which regulate vascular contraction, serve as an informational bridge between vascular endothelial cells and glial cells, particularly astrocytes. The plasticity of pericytes following brain injury has also been suggested. Previously, we analyzed NG2-expressing cells in Rosa26-tdTomato/Cspg4-CreERT2 mice three days after middle cerebral artery occlusion (MCAO) or sham surgery (CTRL) without Wnt pathway modulation (ArrayExpress accession number: E-MTAB-11967). In this study, we extended our analysis by constitutively activating the Wnt/β-catenin signaling pathway, which is crucial for stem cell maintenance, proliferation, and neuronal differentiation. To investigate these processes, we isolated NG2 cells and their derivatives labeled with the red fluorescent protein tdTomato from the cortex of Rosa26-tdTomato/Cspg4-CreERT2 mice three days after MCAO. Mice subjected to sham surgery served as healthy controls. Wnt pathway activation was achieved using an allele that enables constitutive stabilization of β-catenin (Ctnnb1 delEx3). Our aim was to determine how the Wnt pathway overactivation affects the distribution of NG2 glial and perivascular cell subtypes, identify their derivatives in the cortex of healthy and ischemic mice, and characterize their abundance and gene expression profiles.biostudies-arrayexpressLibrary Construction - Single-cell RNA-seq libraries were prepared using Chromium controller instrument and Chromium Next Gem single-cell 3’ reagent kit version 3.1 (both 10X Genomics, Pleasanton, CA, USA) according to the manufacturer’s protocol targeting 4000 cells per sample.Nucleic Acid Extraction - cDNA was extracted from cell suspension during library preparation routine using Chromium next gem single-cell 3′ reagent kit (version 3.1).Sequencing - The libraries were sequenced using NextSeq 500 instrument (Illumina, San Diego, CA, USA).Sample Collection - In adult Rosa26-tdTomato/Cspg4-CreERT2 male mice carrying the Ctnnb1 delEx3 allele, middle cerebral artery occlusion (MCAO) was performed to induce focal cerebral ischemia. Three days after surgery, the cerebral cortex adjacent to the injury site was isolated. For healthy tissue (CTRL), sham-operated mice were used as controls. Fluorescence-activated cell sorting (FACS) was employed to isolate viable cells that were CD31/45-negative and CellTrace-calcein green-positive, expressing tdTomato/NG2 red fluorescence. For each sample, we used tdTomato/NG2 cell isolated from 8 biological replicates.OrganizationMINSEQE ScoreAssays and DataProcessed DataMAGE-TAB FilesData Transformation - 10x cellranger software version 6.1.1 was used to quantify expression per cell. Raw feature-barcode matrices are provided as processed data files.Sequence Alignment - Fastq files were processed using 10x cellranger software version 6.1.1 using GRCm39 assembly with Ensembl annotation version 104. Raw feature-barcode matrices are provided as processed data files.MetabolomicsUnknownTranscriptomicsGenomicsProteomicsNextSeq 500RNA-seq of coding RNA from single cellsMus musculusMiroslava AnderovaLucie JaneckovaJan KubovčiakJan KriskaVladimir KorinekMichal KolarTomáš KnotekfalseSingle cell expression profiling of NG2 cell lineage upon constitutive activation of Wnt/β-catenin signaling in adult mice cortex three days after focal celebral ischemiaIschemic stroke is a severe medical condition that leads to neurological impairments, including the loss of sensory, motor, and cognitive functions. Focal cerebral ischemia (FCI) occurs when blood supply to a specific brain region is interrupted, resulting in cell death. Cells expressing neural-glial antigen 2 (NG2) include glial cells, primarily oligodendrocyte precursors, and perivascular cells. In both cell types, the canonical Wnt signaling pathway is active, with its activity increasing after ischemic injury, influencing the tissue’s response to damage. Following ischemia, NG2 glia rapidly proliferate, migrate to the injury site, contribute to glial scar formation alongside astrocytes, and play a role in brain tissue regeneration. Their potential to differentiate into cell types beyond oligodendrocytes has been documented. Perivascular cells, which regulate vascular contraction, serve as an informational bridge between vascular endothelial cells and glial cells, particularly astrocytes. The plasticity of pericytes following brain injury has also been suggested. Previously, we analyzed NG2-expressing cells in Rosa26-tdTomato/Cspg4-CreERT2 mice three days after middle cerebral artery occlusion (MCAO) or sham surgery (CTRL) without Wnt pathway modulation (ArrayExpress accession number: E-MTAB-11967). In this study, we extended our analysis by constitutively activating the Wnt/β-catenin signaling pathway, which is crucial for stem cell maintenance, proliferation, and neuronal differentiation. To investigate these processes, we isolated NG2 cells and their derivatives labeled with the red fluorescent protein tdTomato from the cortex of Rosa26-tdTomato/Cspg4-CreERT2 mice three days after MCAO. Mice subjected to sham surgery served as healthy controls. Wnt pathway activation was achieved using an allele that enables constitutive stabilization of β-catenin (Ctnnb1 delEx3). Our aim was to determine how the Wnt pathway overactivation affects the distribution of NG2 glial and perivascular cell subtypes, identify their derivatives in the cortex of healthy and ischemic mice, and characterize their abundance and gene expression profiles.2026-08-05T00:00:00Z2026-08-05T08:01:37.782Z2025-04-08T12:05:05.447ZE-MTAB-14958ERP171353E-MTAB-11967EFO_0002944EFO_0004170EFO_0005684EFO_0004917EFO_0005518EFO_0003816EFO_0004184