Derived EVs when compared with regular hepatocyte-derived EV controls, which includes let-7 members of the family. Remedy of human HSCs with TGF-/LPS (20 ng/ ml) for 72 h induced a important decrease of let-7a and let-7b in each activated and control states. Transfection of let-7a and let-7b precursors in human HSCs markedly induced the expression of cellular senescence markers p16 and CCl2, and blunted the enhanced expression of -SMA, collagen a1, MMP-2 and MMP9 (key genes involved inside the activation of HHSCs) by TGF-/LPS treatment. Therapy with MSC/LSC derived EVs (30 g/ml, 72 h) phenocopied the senescence/anti-fibrosis effects of let-7 overexpression in activated HHSCs by TGF-/LPS. A complementary mass spectrometry-based proteomics strategy with luciferase reporter assay identified TLR4, the key LPS receptor, as putative let-7 cluster target. Furthermore, the expressions of senescent hepatic stellate markersIntroduction: MSC-based cell therapy has received fantastic interest within the previous years, especially in regenerative medicine and tissue repair. The concept of priming consists in preconditioning the cells for the duration of the culture phase (often with cytokines or hypoxia) to improve their effects. The literature shows that MSC EVs can PKCθ MedChemExpress recapitulate a substantial element from the valuable effects of the cells they originate from, and that miRNAs are key players in EVs action. Hence, in the present perform, our aim was to establish if IFN or hypoxia priming of MSC could modify their EVs miRNA content material. Strategies: Human bone marrow MSC from 5 healthful donors have been isolated and cultured at 20 of O2 in MEM-alpha/FBS medium till 600 confluence, then with (IFN) or without (CONT) interferongamma (25ng/ml, 48 h) or in hypoxia (three O2 throughout the duration on the culture method). Then the cells were rinced with PBS and placed in serum no cost MEM for 48 h. The conditioned media was collected and EV have been isolated by ultracentrifugation (100 000g for 1h10). Total RNA was isolated and reverse transcribed. Pools of CONT, IFN and HYP cDNA have been ready, miRNA profiling was performed utilizing Exiqon miRnome PCR panel I and II. Then, selected miRNAs have been measured on each sample. Final results: A set of 89 miRNAs was detected (quantification cycle 35) in at the very least one of the pools of MSC EVs. They were measured on every individual sample. 41 miRNAs had been measured in all samples; results wereJOURNAL OF EXTRACELLULAR VESICLESnormalized with 5 endogenous miRNAs. Hypoxia induced no important modification of EVs miRNA content. IFN priming induced a important increase in hsa-miR-106a-5p, 25-3p, 126-3p, 451a and 665. Their S1PR3 Formulation validated targets have been determined with miRTarBase and also the proteins have been analysed with Panther classification method. Among probably the most cited pathways, we discovered p53, inflammation, Wnt signalling, Apoptosis signalling and Angiogenesis.Summary/conclusion: MSC priming can modify the miRNA landscape of their EVs. IFN priming modifies MSCs EVs miRNA involved in biological pathways relevant to tissue repair. Functional analysis of those EVs with chosen miRNAs inhibition is needed to evaluate the biological effects of such an approach. Funding: This function has been funded by the french Path G ale de l’Armement, Biomedef PDH-1SMO-1ISEV2019 ABSTRACT BOOKIndustry Poster Session Thursday 25 April 2019 Place: Level three, Hall AIP.01 IP.Standardizing F-NTA measurements: evaluation of four-wavelengths nanoparticle tracking analysis with cell-line derived EVs Clemens Helmbrechta and Pao.