The Chemokine Receptor CCR2 Mediates the Binding and Internalization of Monocyte Chemoattractant Protein-1 along Brain Microvessels

Abstract

Previous results from this laboratory revealed the presence of high-affinity saturable binding sites for monocyte chemoattractant protein-1 (MCP-1) along human brain microvessels (Andjelkovic et al., 1999; Andjelkovic and Pachter, 2000), which suggested that CC chemokine receptor 2 (CCR2), the recognized receptor for this chemokine, was expressed by the brain microvascular endothelium. To test the role of CCR2 directly in mediating MCP-1 interactions with the brain microvasculature, we assessed MCP-1 binding activity in murine brain microvessels isolated from wild-type mice and from CCR2 (-/-) mice engineered to lack this receptor. Results demonstrate that MCP-1 binding is greatly attenuated in microvessels prepared from CCR2 (-/-) mice compared with wild-type controls. Moreover, microvessels from wild-type mice exhibited MCP-1-induced downmodulation in MCP-1 binding and a recovery of binding activity that was not dependent on de novo protein synthesis. Furthermore, MCP-1 was shown to be internalized within wild-type microvessels, but not within microvessels obtained from CCR2 (-/-) mice, additionally demonstrating that CCR2 is obligatory for MCP-1 endocytosis. Last, internalization of MCP-1, but not transferrin, was observed to be inhibited by disruption of caveolae. Internalized MCP-1 also colocalized at some sites with caveolin-1, a major protein of caveolae, implying that this chemokine is endocytosed, in part, via nonclathrin-coated vesicles. These results prompt consideration that MCP-1 signals may be relayed across the blood-brain barrier by highly specialized interactions of this chemokine with its cognate receptor, CCR2, along brain microvascular endothelial cells.