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Single cell analysis in the adult aorta after Tie2 deletion

ABSTRACT: Single cell analysis in the adult aorta after Tie2 deletion

PROVIDER: PRJNA680252 | ENA |

REPOSITORIES: ENA

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			Action	DRS
	SRR13109055.fastq.gz	Fastqsanger.gz
	SRR13109056.fastq.gz	Fastqsanger.gz
	SRR13109057.fastq.gz	Fastqsanger.gz
	SRR13109058.fastq.gz	Fastqsanger.gz
	SRR13109059.fastq.gz	Fastqsanger.gz

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Endothelial deletion of EPH receptor A4 alters single-cell profile and Tie2/Akap12 signaling to preserve blood-brain barrier integrity

Project description:Neurobiological consequences of traumatic brain injury (TBI) result from a complex interplay of secondary injury responses and sequela that mediates chronic disability. Endothelial cells are important regulators of the cerebrovascular response to TBI. Our work demonstrates that genetic deletion of endothelial cell (EC)-specific EPH receptor A4 (EphA4) using conditional EphA4f/f/Tie2-Cre and EphA4f/f/VE-Cadherin-CreERT2 knockout (KO) mice, promotes blood-brain barrier (BBB) integrity and tissue protection, which correlates with improved motor function and cerebral blood flow recovery following controlled cortical impact (CCI) injury. scRNAseq of capillary-derived KO ECs showed increased differential gene expression of BBB-related junctional and actin cytoskeletal regulators, namely, A-kinase anchor protein 12, Akap12, whose presence at Tie2 clustering domains is enhanced in KO microvessels. Transcript and protein analysis of CCI-injured whole cortical tissue or cortical-derived ECs suggests EphA4 limits the expression of Cldn5, Akt, and Akap12 and promotes Ang2. Blocking the Tie2 receptor using sTie2-Fc, attenuated BBB and tissue protection and reversed Akap12 mRNA and protein levels in KO compared to WT cortical-derived ECs. Conversely, direct stimulation of Tie2 using Vasculotide, an angiopoietin-1 memetic peptide, phenocopied the neuroprotection. Finally, we report a noteworthy rise in soluble Ang2 in the sera of individuals with acute TBI, highlighting its promising role as a vascular biomarker for early detection of BBB disruption. Overall, we describe a novel contribution of the axon guidance molecule, EphA4, in mediating TBI microvascular dysfunction through negative regulation of Tie2/Akap12 signaling.

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Project description:To identify the miRNA expressing profiles of TIE2 expressing Monocytes (TEMs), we have employed the Agilent Human miRNA 8×60K (Design ID:046064) microarray. Human peripheral blood mononuclear cells (PBMCs) in venous blood from healthy donors were isolated by Lymphoprep (Axis-Shield, Norway). Human monocytes in PBMCs, identified as cells that expressed CD14, were enriched by positive immunomagnetic selection using anti-CD14 MicroBeads (Miltenyi, Germany). TEMs (TIE2+CD14+) and TIE2-Monocytes (Tie2-CD14+) were then isolated by FACS-sorting (Aria II, BD Biosciences) using FITC-conjugated anti-CD14 (BD Biosciences, USA) and APC-conjugated anti-TIE2 (R&D System, USA) antibodies.Three TEMs samples together with their paried TIE2-Monocytes were detected.

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