A small molecule esculetin accelerates postprandial lipid circulation by activating C/EBPb and augmenting CD36-mediated phagocytosis in adipose tissue macrophages
Rationale: Exogenous lipid metabolism, influenced by dietary intake, contributes to postprandial dyslipidemia. Adipose tissue macrophages (ATMs) mediate the phagocytosis of postprandial lipids from the exogenous diet, generating high-density lipoprotein (HDL) particles that facilitate lipid circulation and excretion. However, the underlying mechanisms remain poorly understood. This study investigates the effects of esculetin, a coumarin compound, on postprandial cholesterol circulation and excretion following a high-fat meal. Methods: Mice were fed a lipid-rich meal for three days to assess the effects of esculetin on postprandial lipid circulation, using serum lipid profiling and metabolomics analysis. Epididymal white adipose tissue (eWAT) removal and flow cytometry were performed to analyze ATMs and confirm their role in mediating esculetin's effects on postprandial lipemia. Epigenetic profiling, transcriptome analysis, chromatin immunoprecipitation, and Terahertz chemical microscopy were employed to elucidate the molecular targets and mechanisms of esculetin. Results: Esculetin significantly elevates postprandial HDL cholesterol levels to values comparable to pitavastatin and modifies serum metabolites involved in bile-mediated cholesterol excretion, leading to increased bile acid concentrations in the bile. This effect is mediated by an increased ratio and phagocytic activity of a subset of ATMs expressing the scavenger receptor CD36, as eWAT removal and CD36 blockade inhibit this response. Furthermore, esculetin enhances the uptake of oxidized LDL via CD36, as demonstrated in cultured macrophages, and induces epigenetic changes controlled by the key transcription factor C/EBPß, accompanied by increased C/EBPß binding to the Cd36 promoter. A direct interaction between esculetin and C/EBPß was observed using Terahertz chemical microscopy. Additionally, the activation of C/EBPß by esculetin in ATMs was confirmed in vivo. Conclusion: Esculetin accelerates postprandial lipid circulation by binding to C/EBPß and enhancing CD36-dependent phagocytosis in ATMs. Overall design: RAW 264.6 cells were transfected with the LentiCRISPR v2 plasmid containing gRNA against Cebpb or empty vector using the Gene Pulser Xcell Eukaryotic System (165-2661, Bio-Rad, CA, USA). Cells were stimulated with 25 µg/mL DIL-oxLDL and either 0 or 40 µg/mL esculetin for 24 hr