Eicosapentaenoic Acid Potentiates Brown Thermogenesis through FFAR4-dependent Up-regulation of miR-30b and miR-378

Emerging evidence suggests that n-3 polyunsaturated fatty acids (PUFA) promote brown adipose tissue thermogenesis. However, the underlying mechanisms remain elusive. Here, we hypothesize that n-3 PUFA promotes brown adipogenesis by modulating miRNAs. To test this hypothesis, murine brown preadipocytes were induced to differentiate the fatty acids of palmitic, oleate, or eicosapentaenoic acid (EPA). The increases of brown-specific signature genes and oxygen consumption rate by EPA were concurrent with up-regulation of miR-30b and 378 but not by oleate or palmitic acid. Next, we hypothesize that free fatty acid receptor 4 (Ffar4), a functional receptor for n-3 PUFA, modulates miR-30b and 378. Treatment of Ffar4 agonist (GW9508) recapitulated the thermogenic activation of EPA by increasing oxygen consumption rate, brown-specific marker genes, and miR-30b and 378, which were abrogated in Ffar4-silenced cells. Intriguingly, addition of the miR-30b mimic was unable to restore EPA-induced Ucp1 expression in Ffar4-depleted cells, implicating that Ffar4 signaling activity is required for up-regulating the brown adipogenic program. Moreover, blockage of miR-30b or 378 by locked nucleic acid inhibitors significantly attenuated Ffar4 as well as brown-specific signature gene expression, suggesting the signaling interplay between Ffar4 and miR-30b/378. The association between miR-30b/378 and brown thermogenesis was also confirmed in fish oil-fed C57/BL6 mice. Interestingly, the Ffar4 agonism-mediated signaling axis of Ffar4-miR-30b/378-Ucp1 was linked with an elevation of cAMP in brown adipocytes, similar to cold-exposed or fish oil-fed brown fat. Taken together, our work identifies a novel function of Ffar4 in modulating brown adipogenesis partly through a mechanism involving cAMP activation and up-regulation of miR-30b and miR-378.