FRACTALKINE INCREASES APOPTOTIC CELL CLEARANCE BY UPREGULATING MFG-E8 IN MACROPHAGES

Abstract

Clearance of apoptotic cells is crucial in sepsis. We have previously shown that administration of immature dendritic cellderived exosomes containing milk fat globule epidermal growth factor-factor VIII (MFG-E8) to septic animals promoted phagocytosis of apoptotic cells and improved survival. MFG-E8 is an opsonin for apoptotic cells and acts as a bridging protein between phosphatidylserine on apoptotic cells and integrins on phagocytes. Fractalkine (Fr) is a CX3C-chemokine (both a chemoatractant and an adhesion molecule) that is upregulated by proinflammatory cytokines and has been proposed to play a role under inflammatory conditions. A recent study has shown that Fr increased MFG-E8 gene transcription in microglia. We hypothezised that Fr upregulates MFG-E8 in peritoneal macrophages and promotes apoptotic cell clearance without inducing inflammation. To study this, rat peritoneal macrophages (pMφ) were isolated and cultured. MFG-E8 expression was assessed by Western blot after incubation with either LPS or Fr (without LPS). The ability of macrophages to engulf apoptotic thymocytes (A0TC) was determined by TUNEL. Cytokine secretion was assessed by ELISA in cultured cell supernatant. Our results have shown that recombinant MFG-E8 (1 μg/ml) doubles phagocytosis of A0TC by pMφ. Further results are shown in the table below (PI = phagocytosis index): We found similar results in the murine RAW 264.7 cell line, indicating that fractalkine is an important prophagocytic factor. We conclude that fractalkine enhances clearance of apoptotic cells by induction of MFG-E8 in macrophages. Since it does not induce a proinflammatory response in vitro by itself, fractalkine may be safe for the use in animal models of sepsis (NIH R01 GM057468).Table