Fever-range temperature alters continual efferocytosis mediated by mouse pro-inflammatory macrophages

Fever, a cardinal sign of inflammation, has been shown to modulate macrophage functions. Here, we wonder whether fever affects macrophage efferocytosis. This process is critical for the resolution of inflammation and the return to homeostasis with the reprogramming of macrophages toward a pro-resolving phenotype. Using primary mouse bone marrow-derived macrophages (BMDM) stimulated by LPS and IFN-? (M1-like BMDM), we first validated that exposure to febrile-range temperature (39.5°C) induced a heat shock protein response. This was done by RNA sequencing (RNAseq), quantitative RT-PCR (RT-qPCR), and intracellular flow cytometry (FC). Then, we observed that febrile-range temperature decreased pro-inflammatory macrophage efferocytosis assessed by two different FC-based assays and by IncuCyte® live-cell imaging. This reduced efferocytic capacity of macrophages exposed to febrile-range temperature resulted from a decrease capacity to interact with apoptotic cells and to internalise these dying cells. This real-time image-based efferocytosis assay showed also that the heat reduced cell motility of macrophages in response to apoptotic cells. In our RNAseq dataset, we found an upregulation of the Adam17 gene and confirmed this increase by RT-qPCR analysis. Since this gene encodes a protease shedding the efferocytic receptor Mer, we determined Mer expression by FC and soluble Mer in the culture supernatants by ELISA. Febrile-range hyperthermia induced the cleavage of Mer from the cell surface of pro-inflammatory macrophages. Overall, decreased efferocytosis induced by fever-range hyperthermia may result from reduced Mer expression. This suggests that fever, by acting on Mer expression, decreases the efferocytic capacity of macrophages to prolong the pro-inflammatory state.