Cells have the capacity to modulate the microenvironment through the secretion of molecules (cytokines, chemokines, matrix proteins…) and vesicles, which vary according to the pathology involved, such as cancer or the natural process such as senescence. Cellular senescence is a process that enhance with ageing and its association with the onset of age-related diseases. The process is characterized by less proliferation, increased β-galactosidase activity and specific secretory phenotype known as SASP. SASP lead the microenvironment to a more pro-inflammatory one and has the capacity to induce paracrine senescence in neighbouring cells. Regulation of small extracellular vesicles (sEV), which are part of SASP, have a high potential to develop drugs that modulate the senescence transmission. Our aim is to find a proteomic signature of the SASP mediated by sEV to reveal pathways associated with the senescence transmission. In this study we knock-down in mesenchymal stem cells RELA and RAB27A, genes implicated in the paracrine senescence and sEV biogenesis respectively, using CRISPR-Cas9 methodology. After their treatment with senescent or non-senescent sEV, we evaluated the senescence phenotype (proliferation and β-galactosidase activity) in the knock-down cells achieving the inhibition of senescence transmission. Finally, we performed a shotgun study by Tandem Mass Tag (TMT) using 10-plex to identify, quantify and compare the proteome of senescent-sEV recipient cells with or without the knock-down genes. This quantitative and comparative proteomic analysis identified 4099 proteins and quantify 4044 which of 28 were differentially regulated by the SASP mediated by sEV. Further bioinformatic analysis of the dysregulated proteins show the implication of the Golgi traffic and network which evidencing that is involved in the SASP mediated by sEV. This data will be useful to design new therapeutic strategies or support the actual ones against age-related diseases

IX Congress of the Spanish Proteomics Society and 4th Joint Meeting of Spanish, Portuguese and French Proteomics Societies, Córdoba, 6-9 February 2024

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