FFAR4 Functions as a DHA Receptor to Attenuate LPS ‐Induced Inflammation via the cAMP / IκBα Pathway in Large Yellow Croaker ( Larimichthys crocea )

ABSTRACT Docosahexaenoic acid (DHA) exerts anti‐inflammatory effects via free fatty acid receptor 4 (FFAR4) in mammals and is widely used in aquatic feeds for its anti‐inflammatory properties. However, the role of FFAR4 in the anti‐inflammatory effects of DHA in teleosts remains unclear. In this study, FFAR4 overexpression in vivo could significantly suppress LPS‐induced expression of pro‐inflammatory genes ( il‐6 , il‐1β , and tnf‐α ) in the head kidney of large yellow croaker. In contrast, under LPS stimulation, FFAR4 knockdown via exogenous dsRNA significantly aggravated the expression of these pro‐inflammatory genes. Further in vitro experiments demonstrated that FFAR4‐mediated anti‐inflammatory effects were observed only with DHA co‐treatment under LPS stimulation. This regulatory activity was completely abolished in the absence of DHA, indicating that co‐administration of DHA is essential for FFAR4 function. Furthermore, DHA effectively induces receptor internalization in HEK293T cells stably expressing LcFFAR4. DHA exposure rapidly mobilized intracellular calcium ions (Ca 2+ ), elevated cyclic adenosine monophosphate (cAMP) levels, and enhanced phosphorylation of extracellular signal‐regulated kinase 1/2 (ERK1/2). Elevated cAMP levels subsequently triggered cAMP response element‐binding protein (CREB) phosphorylation, which upregulated IκBα transcription and suppressed LPS‐driven inflammatory cascades. In conclusion, this study revealed that FFAR4 mediates the anti‐inflammatory effects of DHA through the cAMP‐CREB‐IκBα signaling pathway, providing novel molecular targets for nutrition‐based anti‐inflammatory strategies in aquaculture.