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Comparative proteomic analysis identifies specific signature proteins in small extracellular vesicles from granulocytes and CD4+ T cell subpopulations

ABSTRACT: Proteomic analysis of small extracellular vesicles (sEV) is a powerful tool for the characterization of cell-to-cell signalling. While several studies describe the proteome profile of different immune cell types, none have analysed the content of their delivered sEV. The aim of the present study was to compare the protein signature of sEV delivered from cell subpopulations with importance for chronic inflammatory diseases: granulocytes (i.e., neutrophils and eosinophils) and CD4+ T cells (i.e., TH1, TH2 and TH17). Qualitative (DDA) and quantitative (DIA-SWATH) proteomic analysis of in vitro produced sEV resulted in the identification of proteomic differences depending on the originating cell source. The main differences were found between granulocytes and TH cell-derived sEV, with higher abundance of antimicrobial proteins (e.g., LCN2, LTF, MPO) in sEV from granulocytes and an enrichment of ribosomal proteins (RPL and RPS proteins) in the TH ones. Additionally, we found differentially abundant proteins between neutrophil and eosinophil sEV (e.g., ILF2, LTF, LCN2) and among TH subsets (e.g., ISG15, ITGA4, ITGB2 or NAMPT). These results highlight the potential of sEV proteins as a source of biomarkers for diseases where the different immune cell subsets studied participate, particularly chronic inflammatory pathologies such as asthma or COPD.

INSTRUMENT(S): TripleTOF 6600

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Leukocyte, Extracellular Vesicle

DISEASE(S): Chronic Inflammation

SUBMITTER: Susana Bravo

LAB HEAD: Susana Belén Bravo López

PROVIDER: PXD046108 | Pride | 2025-02-18

REPOSITORIES: pride

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Project description:In patients with severe kidney disease AVF are surgically placed in order to create good access for hemodialysis. In these dialysis patients the failure rates of AVF can be as high as 24% within 6 months after surgery, causing ineffective dialysis and necessitating additional clinical interventions. The pathological processes known to lead to AVF failure are beginning to be unravelled include the formation of venous neointimal hyperplasia (VNIH), thrombosis (Chang et al. PMID 16105066), and venous stenosis (Kanterman et al. PMID7892454, Tang et al. 1998), resulting in a reduced blood flow through the fistula. We established a rat model for AVF failure in human kidney dialysis patients. The characterization of this model has been previously described (Globerman et al. 2011, PubmedID:22002501). In this model the AVFs are surgically constructed in the right leg by connecting the superficial epigastric vein SEV to the common femoral artery (CFA), resulting in exposure of the SEV to arterial pressure with pulsatile and low resistant flow patterns (Globerman et al. 2011, PMID22002501). In the present study we utilized this AVF model in order to assess the effects of arterialized flow, with consequent pathological changes of the vessel wall due to surgical AVF instalment, on the transcriptome of endothelium from the SEV. Within the SEV these pathologies of the vessel wall include the formation of NIH in the main branch, and stenosis in the side branches. By employing this rat model we assessed the changes of the endothelial transcriptome in relation to these pathologies in order to gain mechanistic understanding of the potential roles of venous endothelium in AVF failure, as well as to identify potential biomarkers preceding AVF failure. AVF surgery was performed on N=6 rats, and 13-14 days after surgery the rats were sacrificed. AVFs were extracted from the rats, and microarray transcriptome analyses were performed on luminal endothelial cells that isolated from four different sites of the AVF by employing Laser Capture Microdissection (LCM). These sites included (i) N=5 AVF sections of the superficial epigastric vein (SEV) with neointimal hyperplasia (NIH), (ii) N=3 AVF sections of SEV side branches with luminal stenosis, (iii), N=6 sections of the SEV located distally from a ligation thereby omitting exposure to arterialized flow and consequently preventing the development of NIH or stenosis, and (iv) N=3 sections of the AVF common femoral artery without signs of NIH or stenosis.

			Action	DRS
	SWATH_library__FDR.xlsx	Xlsx
	TH17_DDA1.wiff	Wiff
	TH17_DDA1.wiff.scan	Wiff
	TH17_DDA1__FDR.xlsx	Xlsx
	TH17_DDA2.wiff	Wiff

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Comparative proteomic analysis identifies specific signature proteins in small extracellular vesicles from granulocytes and CD4+ T cell subpopulations

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