Functional characterization of interleukin 4 and retinoic acid signaling crosstalk during alternative macrophage activation

Retinoic acid possesses potent immunomodulatory properties in various cell types, including macrophages. In this study, we first investigated the effects at the transcriptional and functional levels of exogenous retinoic acid in murine bone marrow-derived macrophages (BMDMs) in the presence or absence of interleukin 4 (IL4), a cytokine with potent anti-inflammatory properties. We examined the effect of IL4 on vitamin A homeostasis in macrophages by quantifying retinoid synthesis and secretion. Our RNAseq data show that exogenous retinoic acid synergizes with IL4 to regulate anti-inflammatory pathways such as oxidative phosphorylation and phagocytosis. Efferocytosis and lysosomal degradation assays validated gene expression changes at the functional level. IL4 treatment altered the expression of several genes involved in vitamin A transport and conversion to retinoic acid. Radiolabeling experiments and mass spectrometry assays revealed that IL4 stimulates retinoic acid production and secretion in a signal transducer and activator of transcription 6 (STAT6)-dependent manner. In summary, our studies highlight the key role of exogenous and endogenous retinoic acid in shaping the anti-inflammatory response of macrophages.