Extracellular calcium and platelet-derived growth factor promote receptor-mediated chemotaxis in osteoblasts through different signaling pathways

Abstract

The discovery of a calcium receptor has stimulated interest in the signaling events underlying extracellular calcium ([Ca2+](o))-induced cell- specific responses. In osteoblasts, elevated levels of extracellular calcium mediate both mitogenesis and chemotaxis. Here we provide evidence that [Ca2+](o)-stimulated chemotaxis of MC3T3-E1 osteoblast-like cells involves a G-protein-linked calcium-sensing receptor. [Ca2+](o) promotes chemotaxis in a concentration-dependent manner. Pertussis toxin blocked almost all of [Ca2+](o)-stimulated chemotaxis but had only a small effect on platelet- derived growth factor (PDGF)-stimulated chemotaxis. Consistent with the signaling model for PDGF-mediated chemotaxis, activation of phospholipase C played a critical role in [Ca2+](o)-initiated chemotaxis: U-73122, an inhibitor of the activation of phospholipase C, blocked approximately 50% of PDGF-stimulated chemotaxis but blocked nearly all of the [Ca2+](o)- stimulated chemotaxis. Down-regulation of protein kinase C also blocked about 50% of PDGF-stimulated chemotaxis but did not block [Ca2+](o)-stimulated chemotaxis. Thus, unlike PDGF-mediated chemotaxis, chemotaxis stimulated by [Ca2+](o) does not appear to require protein kinase C activation. This finding suggests events downstream of inositol 1,4,5-trisphosphate production rather than diacylglycerol production are critical to [Ca2+](o)-promoted chemotaxis of MC3T3-E1 cells. The signal transduction mechanism underlying PDGF-induced chemotaxis involves the activation of phosphoinositide 3- kinase, as judged by the in vivo production of phosphatidylinositol 3,4- diphosphate and 3,4,5-trisphosphate and the partial sensitivity of chemotaxis to wortmannin, an inhibitor of phosphoinositide 3-kinase. In contrast, [Ca2+](o)-stimulated chemotaxis was not blocked by wortmannin and elevations in [Ca2+](o) did not increase the production of lipid products of phosphoinositide 3-kinase. Overall, [Ca2+](o)-promoted chemotaxis of osteoblasts appears to utilize a unique signaling mechanism via a calcium- sensing receptor.