Exosomes derived from mesenchymal stem cells (MSCs) show therapeutic potential despite limited yield. In contrast, the yield of exosome-mimetic vesicles (EMVs), which share a similar structure and size with exosomes is high. Previous studies have compared their proteomic profiles, and the microRNA (miRNA) expression signatures remain unexplored. EMVs from human bone marrow MSCs were isolated through continuous extrusion and exosomes were isolated from the supernatant via differential ultracentrifugation. miRNA sequencing was performed using high-throughput sequencing, and the miRNA expression profiles of MSC-EMVs, MSC-exosomes, and MSCs were compared. Following the comparison of differentially expressed miRNAs in MSC-EMVs and MSC-exosomes, target gene prediction and functional enrichment analyses were performed. Furthermore, a trend analysis was conducted on the miRNA expression levels in the three groups to further explore the relationship between miRNA expression levels. Our study confirmed that EMVs could be stably produced and that their yield was approximately 100-fold higher than that of exosomes. A total of 763 known miRNAs were identified through comparison using the miRBase library. The miRNAs in EMVs and exosomes overlapped with those in MSCs; however, EMVs shared more miRNAs with the parent cells. Comparative analysis identified 21 upregulated and 17 downregulated miRNAs in EMVs versus exosomes, while trend analysis revealed 108 miRNAs preferentially expressed in MSCs and EMVs. Functional enrichment of differentially expressed miRNAs provides new insights for EMV-based therapies. Importantly, we demonstrated that both MSC-EMVs and MSC-exosomes significantly attenuated LPS-induced inflammation in THP-1 macrophages by modulating cytokine secretion (ELISA), suppressing iNOS expression (immunofluorescence), and inhibiting NF-κB activation (western blot). In an lipopolysaccharide (LPS)-induced acute kidney injury model, both vesicle types effectively reduced renal inflammation and tissue damage (histopathology and protein analysis). Our findings not only present the first comprehensive miRNA profiling comparison between MSC-derived EMVs and exosomes but also validate their comparable anti-inflammatory efficacy, supporting EMVs as a viable high-yield alternative for cell-free therapies.