Normal bone remodeling requires the coordinated activities of different bone cells, and is influenced by a variety of hormones and growth factors. Bone cells communicate via intercellular aqueous pores called gap junctions, and express three different gap junction proteins: connexin43 (Cx43), connexin45 (Cx45) and connexin46 (Cx46). Cx43 and Cx45 are found on the plasma membrane of osteoblasts, where they interact to form junctions with different molecular permeabilities. The relative abundance of Cx43 and Cx45 is a critical determinant of the expression of important bone proteins by these cells. In contrast, Cx46 has been found only in an intracellular pool in osteoblastic cells, suggesting that this gap junction protein is functional only in certain specialized circumstances. Bone cells also express P2U purinergic receptor, which mediate cell-cell communication by propagating calcium signaling among cells. Some of the hormones and growth factors that influence bone remodeling alter cell-cell communication. For instance, parathyroid hormone (PTH) and prostaglandin E2 (PGE2) affect bone protein production and also increase the expression and function of Cx43. This proposal seeks to test the hypothesis that intercellular communication is regulated spatially and temporally in bone cells in a manner that allows specific forms of intercellular communication. Using molecular and fluorescence imaging techniques, these studies seek to (1) define the expression of gap junction proteins and P2U receptors in vivo, by immunohistochemistry, and in vitro, in two physiologically relevant bone models, the MC3T3-E1 cell line and rat calvarial osteoblasts, under conditions where mineralization does and does not occur; (2) examine the function of gap junction proteins and P2U receptors under mineralizing and non-mineralizing conditions; and (3) examine the role of hormones, and other factors that influence bone turnover, on cell-cell communication in these models and explore the mechanisms by which observed effects occur. It is suggested (by the applicant) that these studies will define the ways in which physiologically relevant communication among bone cells is regulated, and suggest new approaches to the treatment of metabolic bone diseases such as osteoporosis.
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