Osteopontin modulates CD44‐dependent chemotaxis of peritoneal macrophages through G‐protein‐coupled receptors: evidence of a role for an intracellular form of …

B Zhu, K Suzuki, HA Goldberg… - Journal of cellular …, 2004 - Wiley Online Library
B Zhu, K Suzuki, HA Goldberg, SR Rittling, DT Denhardt, CAG McCulloch, J Sodek
Journal of cellular physiology, 2004Wiley Online Library
Expression of osteopontin (OPN) by activated T‐cells and macrophages is required for the
development of cell‐mediated inflammatory responses. Acting through integrin αvβ3 and
CD44 receptors, OPN can promote chemoattraction and pro‐inflammatory cytokine
expression by macrophages. In this study, we have used periotoneal macrophages from
OPN−/, CD44−/−, and WT mice to study the relationship between OPN and CD44 in
macrophage migration. Using confocal microscopy, we show that OPN co‐distributes with …
Abstract
Expression of osteopontin (OPN) by activated T‐cells and macrophages is required for the development of cell‐mediated inflammatory responses. Acting through integrin αvβ3 and CD44 receptors, OPN can promote chemoattraction and pro‐inflammatory cytokine expression by macrophages. In this study, we have used periotoneal macrophages from OPN−/, CD44−/−, and WT mice to study the relationship between OPN and CD44 in macrophage migration. Using confocal microscopy, we show that OPN co‐distributes with CD44 inside macrophages at cell edges and in cell processes in a mutually dependent manner. The existence of an intracellular form of OPN is supported by pulse‐chase studies in which a thrombin‐sensitive, phosphorylated protein immunoprecipitated with OPN antibodies is retained inside macrophages. In OPN−/− and CD44−/− macrophages, the absence of CD44 and OPN, respectively, is associated with the formation of fewer cell processes, reduced cell fusion required to form functional multinucleated osteoclasts in the presence of CSF‐1 and RANKL, and impaired chemotaxis. Whereas the chemotaxis of CD44−/− cells to various chemoattractants is almost completely abrogated, a differential effect is seen with the OPN−/− cells. Thus, OPN−/− cells migrate normally towards CSF‐1 but not towards fMLP and MCP‐1, which signal through G‐protein coupled receptors (GPCRs). That the GPCR‐mediated migration is dependent upon the level of cell‐surface CD44 is indicated by the reduced cell‐surface expression of CD44 in OPN−/− cells and a comparable impairment in the chemotaxis of CD44+/− cells. Although chemotaxis of OPN−/− cells could be rescued by an OPN substratum, or by addition of high levels of OPN in solution, no response is evident with physiological levels of OPN, indicating a requirement for the CD44‐associated intracellular OPN in CD44 cell‐surface expression. These studies indicate, therefore, that the level of cell surface CD44 is critical for GPCR‐mediated chemotaxis by peritoneal macrophages and suggest that a novel intracellular form of OPN may modulate CD44 activities involved in these processes. J. Cell. Physiol. 198: 155–167, 2004. © 2003 Wiley‐Liss, Inc.
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