Objective: In this study, we investigated the effect of silencing JunB on the differentiation of C3H10T1/2 cells to brown adipocytes and thermogenic capacity. Methods: C3H10T1/2 cells were cultured in vitro and transiently transfected with siRNA to establish a JunB silencing model; the best JunB-siRNA group was selected by CCK-8 and Western blot; differentiation was induced by a cocktail method, and the groups were divided into blank control group (Control), induced differentiation model group (Differentiation Model, DM), and induced differentiation model combined JunB silencing group (DM+JunB-siRNA); the formation of lipid droplets was analysed by Oil Red O staining. Western blot and immunofluorescence staining to detect UCP1 and PGC-1α protein expression, and JC-1 kit to detect mitochondrial membrane potential. Results: JunB-siRNA had no significant effect on cell viability, and JunB-siRNA I silencing had the best effect; silencing of JunB increased the accumulation of lipid droplets in brown adipocytes, the expression of UCP1 and PGC-1α was up-regulated, and the mitochondrial membrane potential was enhanced. Conclusion: JunB silencing increased the lipid droplet content and thermogenesis of brown adipocytes, which may be achieved by improving mitochondrial function.