Use of calcium isotopes for assessment of bone turnover: Methodology and botanical intervention studies

Osteoporosis affects 9 million Americans and is responsible for 2 million fractures per year. Current osteoporosis drug therapies reduce fracture risk but have been linked to severe adverse events. New osteoporosis prevention and treatment therapies are needed, but traditional methods for evaluating therapies are costly and time consuming. Bone-seeking label turnover is an alternative approach that allows for rapid screening of potential osteoporosis therapies. This dissertation focuses on the development and application of calcium isotope tracers for evaluating bone turnover. The first study presented in this dissertation was conducted to validate the ability of this methodology to screen therapies acting through anabolic mechanisms. We treated 45Ca-labeled rats with teriparatide, an anabolic osteoporosis drug, and found that urinary tracer excretion decreased and predicted changes in bone mineral content, confirming this method is suitable for anabolic therapies. In the second study, we evaluated the extent to which bone calcium is able to contribute to serum calcium homeostasis independently of osteoclast-mediated bone resorption. 45Ca-labeled rats were treated with OPG to inhibit bone resorption, and changes in tracer appearance in the serum and urine were monitored. In OPG treated rats, serum 45Ca decreased by 81%, indicating that bone resorption is the primary mechanism for removal of calcium from bone. The third and fourth studies in this dissertation focus on evaluating potential botanical therapies for osteoporosis. In study three, the efficacy of dietary soy isoflavones as a co-therapy with risedronate, an antiresorptive drug, was evaluated. 45Ca-labeled rats were treated with isoflavones alone or in combination with two different doses of risedronate in a crossover design. Though dietary isoflavones reduced bone turnover, they did not further increase the efficacy of risedronate when used in combination therapy. However, dietary isoflavones combined with a reduced dose of risedronate was as effective as a standard dose of risedronate alone, suggesting that drug dose reduction may be possible when isoflavones are used as a cotherapy. In study four, the bone-sparing effects of dietary dried grape were evaluated. 45Ca-labeled ovariectomized rats were randomized to a diet containing 25% dried grape powder or control diet for 8 weeks. Urinary 45Ca excretion was monitored throughout treatment, and Ca balance and kinetic studies were performed. At the end of 8 weeks, bones were collected and evaluated for density, microarchitecture, and strength. Rats fed the dried grape diet had increased bone Ca retention and improved femoral structure and strength compared to control rats. The work presented in this dissertation further advances bone-seeking label turnover methodology and provides pre-clinical groundwork for evaluating potential botanical therapies for osteoporosis