Fractalkine-induced MFG-E8 leads to enhanced apoptotic cell clearance by macrophages

Clearance of apoptotic cells is crucial to maintain cellular function under normal and pathological conditions. We have recently shown that administration of immature dendritic cell-derived exosomes to septic animals promotes phagocytosis of apoptotic cells and improves survival by providing milk fat globule epidermal growth factor-factor VIII (MFG-E8). MFG-E8 acts as an opsonin for apoptotic cells to be engulfed by phagocytosis. In the present study we investigated whether the CX₃C-chemokine fractalkine (CX ₃CL1) promotes apoptotic cell clearance through the induction of MFG-E8 in peritoneal macrophages. Cultured rat peritoneal macrophages (pMφ) and RAW264.7 macrophages were stimulated with LPS and CX₃CL1. MFG-E8 expression was assessed by Western blot, cytokine secretion was assessed by ELISA, and phagocytosis of apoptotic thymocytes was determined by microscopy. For in vivo studies, cecal ligation and puncture (CLP) was used to induce sepsis in rats and mice. LPS significantly decreased MFG-E8 levels and phagocytosis of apoptotic cells, whereas CX₃CL1 induced MFG-E8 expression in both nonstimulated and LPS-stimulated pMφ, without affecting TNF-α and IL-6 release. Anti-MFG-E8 blocking antibodies completely abrogated the prophagocytic effect of CX₃CL1. Twenty hours after the induction of sepsis in rats via CLP, plasma CX₃CL1 levels as well as MFG-E8 production in peritoneal macrophages decreased by 21% and 56%, respectively. Administration of CX₃CL1 on the other hand induced MFG-E8 and prevented tissue injury. We conclude that CX₃CL1 induces MFG-E8 in vitro and in vivo and enhances clearance of apoptotic cells in an MFG-E8-dependent manner. These findings suggest a possible novel treatment for patients in sepsis.