Bone development and remodeling are dependent on a complex interactive network of humoral and local factors produced by bone cells, bone marrow cells, immunocompetent cells, vascular cells, and precursors for these cell populations. Insights obtained as to which osteoblast factors influence osteoclast (Oc) development and how they have helped define the cellular and molecular processes involved in bone development and remodeling and provide potential new therapeutic strategies to overcome endocrine or local disorders in bone formation and resorption becomes uncoupled. The converse of this avenue of research has to date received only limited attention, namely, how Ocs might regulate osteoblast development, activity, and senescence. There is now a growing awareness based on experimental evidence demonstrating that Ocs synthesize and release cytokines, growth factors, matrix molecules, arachidonic acid metabolites, and other reactive species such as superoxide and nitric oxide. These may influence osteoblast differentiation and action. The objectives outlined in this project re designed to build on our observations that Ocs and Oc-like cells can modulate osteoblast development and activity. During the last period of support we showed that Ocs produce small pro-inflammatory peptides know as chemokines, such as IL-8 and GROalpha, in a regulated fashion and that these chemokines influenced osteoblast development and function. To further define the role of Oc-derived chemokines in normal and pathological bone remodeling we propose: 1) to expand our analysis of chemokines proposed by Ocs and their modulation by hormonal cytokine and matrix-dependent signals, 2) to examine the effects of Oc-derived chemokines, such as IL-8 and GROalpha, on osteoblast development and function, including their influence on osteoblast migration, integrin expression, matrix synthesis or degradation, and 3) to identify and characterize osteoblast chemokine receptors has a function of osteoblast development, physiology, and pathophysiology. Recent signal transduction mechanisms involved in Oc- derived chemokine mediated modulation of osteoblastogenesis and activity will be examined. All of the above studies will use a combination of in vivo and in vitro approaches, and model systems including the rat calvarial injection model for histomorphometric studies, human tissue sections for in situ hybridization and immunohistochemical analysis, isolated human and avian Ocs human Oc-like cells, the mouse Oc-like cell developmental, and cells obtained from an IL-8 receptor knockout mouse. Oc chemokine production, mRNA steady state levels and regulation will be assessed by RT=PCR, RNAse protection assay, chemokine ELISA, and in situ hybridization techniques. Osteoblast development and activity will be evaluated based on multiple biochemical and molecular markers of the osteoblast phenotype, including integrins, matrix proteins, and metalloproteinases. Such studies are anticipated to reveal new aspects of normal bone remodeling mechanisms and have potential to lend insight into skeletal pathologies such as osteoporosis, osteoarthritis and other inflammatory bone disorders.
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