CALCIUM TRANSPORT, BONE CELL FUNCTION, AND AGING

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Bone is a dynamic tissue maintained in a constant state of flux. During much of adult life overall bone mass remains constant due to a net balance between osteoblastic mediated bone formation and osteoclastic controlled bone resorption. This "balance" results from the phenomenon of "coupling" which describes new bone formation in response to bone resorption and vice versa. It has been postulated that osteoporosis and the age-related alterations in bone mass result from defective osteoblastic function and/or acquired changes in the "coupling" process. Using "putative" rodent osteoblastic models, the complexities which characterize the control of osteoblastic function, growth and differentiation are gradually being unraveled. Factors controlling osteoclastic bone resorption such as the cell's ability to secrete large amounts of hydrogen ion at bone attachment sites have also been identified. A number of published observations also support the hypothesis that osteoblasts regulate the number and activity of osteoclasts by producing short range paracrine factors; conversely, the modulation of osteoblastic controlled bone formation by osteoclastic activity has yet to be examined in detail. Consistent with our on-going research interests in age-related alterations in bone mass and calcium transport, and the past accomplishments of our Program Project endeavors, the experimental protocols described in this application will address: (1) the effect of calcitrophic hormones on young and aged rodent and human osteoblasts with emphasis on the regulation of intracellular calcium control, calcium transport and cell growth and the production of neutral metalloproteinases and proteinase inhibitors; (2) the critical role of alkaline phosphatase as an index of osteoblastic activity and as a modulator of bone mineralization; (3) the interrelationship between the control of cell differentiation by 1,25(OH)2D3 and its recently- discovered control of cell Ca2+ homeostasis; (4) the sequence of events leading to amplified expression and polarization of proton pumps during osteoclastic differentiation; and (5) the modulation of bone formation as a result of osteoclastic activity with emphasis on age-related changes in the "coupling signals." To accomplish this goal, we plan to utilize rat and human organs and osteoblastic cell cultures, renal tubular cell cultures, immunocytochemistry and in situ hybridization techniques, a variety of fluorescent probes and spectro-fluorometric systems with video image analysis designed to quantitate intracellular calcium and pH transients in single cell preparations.