Fractalkine-Induced MFG-E8 Leads to Enhanced Apoptotic Cell Clearance by Macrophages

Michael Miksa,Thanjavur S Ravikumar,Weifeng Dong,Ping Wang,Rongqian Wu,Dhruv Amin

doi:10.2119/2007-00019.miksa

Abstract

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 CX(3)C-chemokine fractalkine (CX(3)CL1) promotes apoptotic cell clearance through the induction of MFG-E8 in peritoneal macrophages. Cultured rat peritoneal macrophages (pMphi) and RAW264.7 macrophages were stimulated with LPS and CX(3)CL1. 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 CX(3)CL1 induced MFG-E8 expression in both nonstimulated and LPS-stimulated pMphi, without affecting TNF-alpha and IL-6 release. Anti-MFG-E8 blocking antibodies completely abrogated the prophagocytic effect of CX(3)CL1. Twenty hours after the induction of sepsis in rats via CLP, plasma CX(3)CL1 levels as well as MFG-E8 production in peritoneal macrophages decreased by 21% and 56%, respectively. Administration of CX(3)CL1 on the other hand induced MFG-E8 and prevented tissue injury. We conclude that CX(3)CL1 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.

Highlights

Fractalkine (CX3CL1), a molecule with the dual function of an adhesion molecule and a chemokine, is mainly derived from nonhematopoietic cells [1,2]
The phagocytosis index of recombinant murine MFG-E8 (rMFG-E8) opsonized apoptotic thymocytes increased on average by 90% compared with nonopsonized thymocytes (Figure 1A and B), showing that murine recombinant milk fat globule protein (MFG)-E8 has a prophagocytic effect on rat peritoneal macrophages
In an experimental sepsis model, we found that plasma CX3CL1 levels significantly decreased, a response that was paralleled by a dramatic decrease in MFG-E8 in peritoneal macrophages and blood

Summary

Introduction

Fractalkine (CX3CL1), a molecule with the dual function of an adhesion molecule and a chemokine, is mainly derived from nonhematopoietic cells [1,2]. It has a unique structure in that the CX3C motifbearing chemokine domain is carried by a mucin-like stalk attached to the cell surface membrane. Activation of endothelial cells by TNF-α, for example, induces CX3CL1 expression on the surface of the cells leading to the adhesion and transmigration of macrophages [8,9]. CX3CL1 is necessary for normal brain development, during which apoptosis naturally occurs [13]

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Conclusion