Metabolic acidosis, a common clinical disorder, produces bone mineral dissolution. The proposed studies are designed to examine the effects of alterations in proton concentration ([H+]) on the elemental composition of bone mineral. We will study the effects of H+ on both physicochemical mineral dissolution and cell-mediated resorption in cultured neonatal mouse calvariae and in synthetic ceramic carbonated apatite (CAP) disks using a high resolution scanning ion microprobe in conjunction with measurements of ion flux. With the microprobe we obtain sensitive, high resolution, mass resolved images and spectra of the ionic composition of bone and CAP disks. We propose to use the high resolution scanning ion microprobe to test the following hypotheses: 1) That cells and/or proteins contribute to measured bone surface ion composition by comparing ion concentration and localization in cultured calvariae a) treatment with hydrazine (to deproteinate the bone) and b) in areas of high and low ratios of C-N bonds to Ca).2) Compare the effects on bone elemental composition of chronic culture of mouse calvariae in physiologically acidic medium with chronic metabolic acidosis in vivo by comparing the changes in mineral ion concentration and localization in calvariae after in vitro culture in acid medium with calvariae from mice with a comparable level of systemic acidosis. 3) That ionic flux from CAP disks cultured in physiologically acidic medium is similar to that from calvariae by comparing Ca and P efflux, H influx and ion composition of calvariae and CAP disks during culture in medium simulating metabolic and respiratory acidosis. 4) That H+ mediated bone Ca release involves dissolution of bone Ca with CO3= as well as with P by determining the change in the ratio of Ca/P on the bone and CAP disks after incubation in acidic medium. In addition we will compare the base of the "resorption pits: to the surrounding mineral when isolated osteoclasts are incubated on cortical bone and CAP disks in acidic medium. 5) That physicochemical mineral dissolution results in a similar relative release of K and Ca while cell-mediated resorption results in greater relative release of Ca than K, by prelabeling calvariae and CAP disks with 44Ca and 41K and determining the change in ion ratios after cell and physicochemical Ca release (calvariae in acidic medium), cell-mediated resorption (calvariae + 1,25(OH)2D3) and physicochemical dissolution (calvariae in acidic medium + calcitonin and CAP disks in acidic medium. 6) That medium [HCO3- ] regulates cell-mediated Ca release by comparing the quantity of Ca released from calvariae during compensated metabolic and respiratory acidosis. 7) That metabolic alkalosis will stimulate bone formation in vivo and will cause mineralization in vitro by comparing the ratio of 44Ca/40Ca in calvariae of alkalotic mice injected with 44Ca with calvariae incubated in alkalotic 44Ca medium. 8) That the first Ca deposited in bone is also the first Ca released by prelabeling calvariae with 44Ca in vivo and determining the change in the ratio of 44Ca/40Ca during in vitro incubation in acidic medium. These studies will allow us to obtain a better understanding of how changes in pH affect the bone mineral in order to construct a model of H+-mediated bone Ca release.
NIH ExPorter
NIH ExPorter