UC Davis

OBJECTIVE: To characterize effects of inflammatory cytokine priming on viability, mitochondrial morphology, and extracellular vesicle (EV) cargo in feline placenta-derived mesenchymal stromal cells (MSCs). METHODS: Feline placenta-derived MSCs from 7 healthy cats were cultured under control, serum-deprived, or cytokine-stimulated (tumor necrosis factor-α, interferon-γ, IL-1, IL-6) conditions in vitro. Viability/apoptosis were assessed by flow cytometry and mitochondrial morphology by confocal microscopy with MicroP/MATLAB; EVs were characterized by surface markers, RNA sequencing, and microRNA (miR) profiling. RESULTS: Viability exceeded 96% across all conditions, with no significant intergroup differences. Kiel 67 proliferation marker and metabolic activity were significantly reduced under serum-free and cytokine-stimulated conditions versus controls (both P < .001). Cytokine priming increased branching and simple tubular mitochondria, whereas serum deprivation increased fragmented forms. Extracellular vesicle transcriptomics identified 329 differentially expressed genes, including upregulated immunosuppressive receptors C-C motif chemokine receptor 8 and poliovirus receptor-related immunoglobulin domain-containing protein and downregulated proinflammatory histone transcripts. microRNA (miR) profiling revealed 23 differentially expressed miRs, with enrichment of anti-inflammatory miR-204, let-7g, and miR-181b-2 and depletion of proinflammatory miR-34a and miR-320a. CONCLUSIONS: Cytokine priming induces mitochondrial elongation and shifts EV cargo toward an immunosuppressive phenotype without compromising viability, supporting inflammatory priming as a strategy to optimize MSC-based therapies for T-cell-mediated inflammatory diseases. CLINICAL RELEVANCE: Inflammatory priming prior to MSC administration may enhance therapeutic efficacy in cats, supporting investigation of cytokine licensing as a translatable approach to improving MSC therapy outcomes in veterinary medicine.