Identification of miRNA Reference Genes in Extracellular Vesicles from Adipose Derived Mesenchymal Stem Cells for Studying Osteoarthritis.

摘要: Osteoarthritis (OA) leads to chronic pain and disability, traditional conservative treatments are not effective in the long term. The intra-articular injection of mesenchymal stem cells (MSCs) is considered a novel therapy for OA whose efficacy mainly relies on adaptive release paracrine molecules which either soluble or extracellular vesicles (EVs) embedded. correct quantification EV-miRNAs using reliable reference genes (RGs) crucial step optimizing this future therapeutic cell-free approach. purpose study rate stabilities literature-selected proposed RGs adipose derived-MSCs (ASCs). EVs were isolated by ultracentrifugation from ASCs cultured with without inflammatory priming mimicking synovial fluid condition. Expression putative (let-7a-5p, miR-16-5p, miR-23a-3p, miR-26a-5p, miR-101-3p, miR-103a-3p, miR-221-3p, miR-423-5p, miR-425-5p, U6 snRNA) was scored algorithms geNorm, NormFinder, BestKeeper ΔCt method. miR-16a-5p/miR-23a-3p yielded most stable RGs, whereas let-7a-5p/miR-425-5p performed poorly. Outcomes validated qRT-PCR miR-146a-5p, reported be ASC-EVs enriched involved OA. Incorrect RG selection affected evaluation miR-146a-5p abundance modulation inflammation, both values resulting strongly donor-dependent. Our findings demonstrated that an integrated approach multiple necessary identify reliable, miRNAs evaluation. A would increase accuracy embedded molecule assessments aimed develop strategies treatment based EVs.