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Mesenchymal stem cells (MSCs) hold immense promise in regenerative medicine due to their unique capabilities, including differentiation into various cell types and paracrine signaling. Among their secretions, extracellular vesicles (EVs) play a pivotal role in mediating therapeutic effects through their bioactive cargo, including microRNAs (miRNAs). These miRNAs are key regulators of gene expression, modulating critical biological processes such as inflammation, immune response, and tissue repair. MSC-derived EVs have demonstrated significant therapeutic potential across diverse diseases, including cardiovascular, neurological, autoimmune, and oncological conditions. This review explores the biogenesis, isolation, and characterization of MSC-EVs, with a focus on the functional roles of miRNAs in disease modulation. The therapeutic prospects of MSC-EV miRNAs as potential biomarkers, drug delivery agents, and modulators of pathophysiological pathways are discussed, providing insights into future translational research and clinical applications.