Rate Of Calcium Deposition Research Articles

Caffeic acid treatment augments the cell proliferation, differentiation, and calcium mineralization in the human osteoblast-like MG-63 cells

Background: Osteoporosis is an imperative health problem that extremely distresses the public that leads to a higher risk to the bones from both spontaneous and accidental bone fractures. Caffeic acid is a polyphenol compound that happens naturally in numerous vegetables such as coffee beans, potatoes, propolis, olives, and carrots with many pharmacological aids. Objectives: The current study was planned to examine the potential of Caffeic acid in proliferation, differentiation, and calcium mineralization of osteoblast-like MG-63 cells. Materials and Methods: The cell viability of Caffeic acid-supplemented MG-63 cells was examined through the 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyltetrazolium bromide (MTT) cytotoxicity test. The mRNA expression of alkaline phosphatase (ALP) and osteocalcin was reviewed through reverse transcription polymerase chain reaction study. The calcium deposition rate in the Caffeic acid-treated MG-63 cells was studied through Alizarin Red S (ARS) staining. Results: The result of the MTT test exposed that the 10 and 20 mg/kg of Caffeic acid supplementation did not show any cytotoxicity to the osteoblast-like MG-63 cells; instead, it helped the viability of MG-63 cells. The expression of ALP is particularly increased in the Caffeic acid-supplemented MG-63 cells, whereas the osteocalcin expression was noticeably diminished. The ARS staining was exhibited that the Caffeic acid treatment was noticeably enhanced the calcium mineralization rate in the osteoblast-like MG-63 cells. Conclusion: Based on the findings of investigation, it was proved that the Caffeic acid treatment was significantly enhanced the cell proliferation, differentiation, and calcium mineralization in the osteoblast-like MG-63 cells. Hence, it was clear that Caffeic acid can be engaged as the potential agent for the purpose of bone regeneration.

Effects of the Whey Protein Hydrolysates of Various Protein Enzymes on the Proliferation and Differentiation of 3T3-E1 Osteoblasts.

In this study, we used various proteinases to investigate the effect of whey protein hydrolysates on proliferation and differentiation of MC3T3-E1 osteoblasts. To confirm hydrolysis of the whey protein hydrolysates, the yield and α-amino acid content were determined. Since osteogenic cell activity is an important factor in osteogenesis, we evaluated the proliferation of osteogenic cells by measuring 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide and alkaline phosphatase (ALP) activity. To analyze bone matrix formation, we identified calcium deposition by staining with Alizaline red-S. The free amino acid content was significantly higher in the whey protein hydrolysates prepared using Protamex, Flavourzyme, and Alcalase than in the control. When cells were treated with 500 μg/mL of whey protein hydrolysates prepared using Protamex and Alcalase, cell proliferation increased by 120% and 130%, respectively, compared with the control group. In addition, ALP activity was significantly higher following treatment with 500 μg/mL of whey protein hydrolysates prepared using Protamex and Alcalase (142.61% and 135.06%, respectively; P<0.05). Furthermore, when treated with 125 μg/mL of the same hydrolysates, the rate of calcium deposition increased significantly to 157.56% compared with the control group (P<0.05). Therefore, our results suggest that whey protein hydrolysates prepared using Protamex and Alcalase may have more beneficial effects on osteoblast proliferation and bone health than those prepared using other proteolytic enzymes.

Effects of nano-engineered surfaces on osteoblast adhesion, growth, differentiation, and apoptosis

Modifying implant surfaces to improve their biocompatibility by enhancing osteoblast activation, growth, differentiation, and induction of greater bone formation with stronger attachments should result in improved outcomes for total joint replacement surgeries. This study tested the hypothesis that nano-structured surfaces, produced by the ion beam-assisted deposition method, enhance osteoblast adhesion, growth, differentiation, bone formation, and maturation. The ion beam-assisted deposition technique was employed to deposit zirconium oxide films on glass substrates. The effects of the ion beam-assisted deposition technique on cellular functions were investigated by comparing adhesion, proliferation, differentiation, and apoptosis of the human osteosarcoma cell line SAOS-2 on coated versus uncoated surfaces. Ion beam-assisted deposition nano-coatings enhanced initial cell adhesion assessed by the number of 4′,6-diamidino-2-phenylindole–stained nuclei on zirconium oxide nano-coated surfaces compared to glass surfaces. This nano-modification also increased cell proliferation as measured by mitochondrial dehydrogenase activity. Moreover, the ion beam-assisted deposition technique improved cell differentiation as determined by the formation of mineralized bone nodules and by the rate of calcium deposition, both of which are in vitro indicators of the successful bone formation. However, programmed cell death assessed by Annexin V staining and flow cytometry was not statistically significantly different between nano-surfaces and glass surfaces. Overall, the results indicate that nano-crystalline zirconium oxide surfaces produced by the ion beam-assisted deposition technique are superior to uncoated surfaces in supporting bone cell adhesion, proliferation, and differentiation. Thus, surface properties altered by the ion beam-assisted deposition technique enhanced bone formation and may increase the biocompatibility of bone cell–associated surfaces.

Decoupling of otolith and somatic growth during anadromous migration in a northern salmonid

We assessed the fish length – otolith length relationship (FL–OL) in Dolly Varden (Salvelinus malma malma) to verify proportional growth. A decoupling was detected during first ocean migration where fish growth was occurring at a greater rate than otolith growth. Because of this decoupling, the application of traditional back-calculation models overestimated the size-at-age in premigratory char. We developed modified back-calculation equations from existing traditional models to account for this decoupling based on discontinuous piecewise regressions. The new biological intercept breakpoint method (BI–BP) provided the most accurate representation of fish size-at-age throughout all life history stages when compared with known size-at-capture values in fish. The decoupling indicates that factors other than somatic growth are important for otolith accretion. Physiological changes during smoltification likely alter calcium uptake and thereby affect calcium deposition rates on otoliths during this short but biologically critical time period of life history. It is probable that species exhibiting similar complex ontogenetic shifts in life history will likely exhibit decoupling to some extent in the FL–OL relationship.

Abstract 799: Tumour micro-environmental factors and the development of mammary calcifications: Recent findings from an in-vitro model

Abstract The presence of microcalcifications on mammographic images represents a highly valuable tool in the early detection of breast cancer, often allowing radiographers to diagnose mammary tumours at an early, in-situ stage. The presence of microcalcifications has also been linked to a number of unfavourable prognostic factors including decreased survival and an increased probability of relapse. Despite their long history of use in the clinic, the precise mechanisms by which microcalcifications are formed remains poorly understood. Pathological soft-tissue calcification is often considered the result of an imbalance between pro- and anti-calcifying factors. However, the possible impact of such an imbalance has not been studied in the context of breast calcification. We have established an in-vitro model of microcalcification formation using the triple-negative adenocarcinoma cell line MDA-MB-231. When cultured in the presence of the osteogenesis-promoting reagents β-glycerophosphate, ascorbic acid and dexamethasone, calcified deposits begin to form by Day 14, as verified by Alizarin Red staining and the quantitative o-cresolphthalein assay. Following the establishment of our model, we set out to identify the underlying molecular triggers initiating the calcification process, in particular the role of micro-environmental factors. Altered magnesium homeostasis has been suggested as an important mediator of tissue calcification. We found that a slight increase in magnesium concentration almost completely blocked calcium deposition in our model. Previous studies in other tissues have shown this protective effect to be dependent on the cation channel TRPM7 which is increased in breast cancer patients with calcifications compared to those without. Unexpectedly, inhibition of the TRPM7 channel by two separate compounds (2-APB and NS-8593) not only failed to reverse the inhibition of microcalcification formation by exogenous magnesium, but actually further decreased calcium deposition by Day 28, suggesting that calcium influx via the TRPM7 channel may be promoting development of mammary calcifications. Finally, we investigated the effect of several tumour associated cytokines on the rate of calcium deposition and found that IL-1β and TNF-α blocked mineralisation whilst IL-6 and BMP2 lead to an increase. Interestingly, co-administration of IL-6 alongside a soluble form of its receptor (sIL-6R) was observed to promote mineralisation even in the absence of dexamethasone, which had previously been essential to the formation of calcifications in our model. To date, our model has yielded a number of important insights into the formation of calcifications in breast cell lines, many of which recapitulate observations from patient studies. It is hoped that this work will contribute to our understanding of the origin of these pre-invasive diagnostic clues. Citation Format: Shane O'Grady, Maria P. Morgan. Tumour micro-environmental factors and the development of mammary calcifications: Recent findings from an in-vitro model [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 799. doi:10.1158/1538-7445.AM2017-799

Reengineered graft copolymers as a potential alternative for the bone tissue engineering application by inducing osteogenic markers expression and biocompatibility

Composite scaffolds of nano-hydroxyapatite with demineralized bone matrix were prepared and they were graft copolymerized for better bone regeneration and drug delivery applications. The graft copolymers were characterized for their physiochemical properties using conventional methods like FTIR, TGA, XRD and SEM. The scaffolds were seeded with 3T3 and MG63 cells for studying their biocompatibility and their temporal expression of ALP activity, the rate of calcium deposition and their gene expression of collagen type I (Coll-1), osteopontin (OP), osteonectin (ON), and osteocalcin (OC) were studied. In vivo studies were conducted using sub-cutaneous implantation models in male Wister rats for 6 months. Periodic radiography and post-autopsy histopathology was analysed at 15days, 1, 2, 3, 4, 5, and 6 months. The obtained in vitro results clearly confirm that the bone scaffolds prepared in this study are biocompatible, superior osteoinductivity, capable of supporting growth, maturation of MG 63 osteoblast like cells; the gene expression profile revealed that the material is capable of supporting the in vitro growth and maturation of osteoblast-like cells and maturation. The in vivo results stand a testimony to the in vitro results in proving the biocompatibility and osteoinductivity of the materials.

Calcification in vitro: experimental factors that induce nanocrystal formation (1077.9)

Medial vascular calcification (VC) of arteries is frequently observed in chronic kidney disease and diabetes. Pathogenesis in VC is frequently studied on primary cultures of aortic smooth muscle cells. However, several experimental factors can affect the rate of calcification in vitro, which can be misleading in the research of VC pathogenesis. The aim of this work has been the identification of these factors and to improve the adequacy of the model.Calcification was studied as a function of Pi concentration (1‐3 mM), incubation time and media composition (MEM vs. DMEM). We have determined the rate of calcium deposition, medium pH, nucleation rate, degree of crystallization, cell death and osteogene expression.Calcification was more intense with DMEM than MEM due to increased basicity induced by higher bicarbonate content in DMEM. Cell death (as LDH leakage) correlated and preceded calcification. Particle nucleation depended on pH and Pi concentration, and was already observed after 2h (DMEM) or 5h (MEM) of incubation with the cells. Osteogene expression and crystal structure were also modified by the same experimental conditions.

Combined effects of temperature and ocean acidification on the juvenile individuals of the mussel Mytilus chilensis

Anthropogenic CO2 emissions have led to increasing global mean temperatures (a process called global warming) and ocean acidification. Because both processes are occurring simultaneously, to better understand their consequences on marine species their combined effects must be experimentally evaluated. The aim of this study was to evaluate for the first time the combined effects of ocean acidification and water temperature increase on the total calcification rate, growth rate and survival of juvenile individuals of the mytilid mussel Mytilus chilensis (Hupe). Two temperature levels (12 and 16°C) and three nominal CO2 concentrations (390, 700 and 1000ppm of CO2) were used. We found that the net rate of calcium deposition and total weight were not significantly affected by temperature, but were negatively affected by the levels of CO2. The interactive effects of temperature and CO2 levels affected only the shell dissolution, but this process was not important for the animal's net calcification. These results suggest that individuals of M. chilensis are able to overcome increased temperatures, but not increments of CO2 levels. It is well known that mussels influence their physical and biological surroundings. Therefore, the negative effects of a CO2 increase could have significant ecological consequences, mainly in those habitats where this group is dominant in terms of abundance and biomass. Finally, taking into account that this species inhabit a wide geographic range, with contrasting environmental conditions (e.g., temperature, salinity and, pH), further studies are needed to evaluate the intraspecific variability in the responses of this species to different environmental stressors.

Calcium–bismuth electrodes for large-scale energy storage (liquid metal batteries)

Calcium is an attractive electrode material for use in grid-scale electrochemical energy storage due to its low electronegativity, earth abundance, and low cost. The feasibility of combining a liquid Ca–Bi positive electrode with a molten salt electrolyte for use in liquid metal batteries at 500–700 °C was investigated. Exhibiting excellent reversibility up to current densities of 200 mA cm−2, the calcium–bismuth liquid alloy system is a promising positive electrode candidate for liquid metal batteries. The measurement of low self-discharge current suggests that the solubility of calcium metal in molten salt electrolytes can be sufficiently suppressed to yield high coulombic efficiencies >98%. The mechanisms giving rise to Ca–Bi electrode overpotentials were investigated in terms of associated charge transfer and mass transport resistances. The formation of low density Ca11Bi10 intermetallics at the electrode–electrolyte interface limited the calcium deposition rate capability of the electrodes; however, the co-deposition of barium into bismuth from barium-containing molten salts suppressed Ca–Bi intermetallic formation thereby improving the discharge capacity.

Differentiation of MC3T3-E1 on poly(4-styrenesulfonic acid-co-maleic acid)sodium salt-coated films

Polyelectrolyte multilayer (PEM) film can modify the surface properties of materials to improve cellular responses. In this study poly(diallyldimethylammonium chloride) (PDADMAC), poly(sodium 4-styrene sulfonate) (PSS) and poly(4-styrenesulfonic acid-co-maleic acid) sodium salt (PSS-co-MA) were assembled into PEM {(PDADMAC/PSS)(4)/PDADMAC+PSS-co-MA} film on glass surfaces and its ability on affecting osteoblast functions were examined. PSS-co-MA film showed an increase roughness and more wettable surface as compared to glass. When the osteoblast cell line, MC3T3-E1, was seeded on the surfaces, no differences were observed in cell attachment or spreading on either PSS-co-MA film or glass at 4-16 hours. However, increases in alkaline phosphatase activity (day-5 and 7) and the expression of osteocalcin mRNA/protein at day-13 were observed. Cells cultured on PSS-co-MA film developed a faster rate of calcium deposition at day-15 compared to control. In conclusion, PSS-co-MA film enhanced osteoblast differentiation and could be used to promote mineralization and improve osseointegration for dental implants.

Osteogenic Effects of Rest Inserted and Continuous Cyclic Tensile Strain on hASC Lines with Disparate Osteodifferentiation Capabilities

We investigated the effects of two types of cyclic tensile strain, continuous and rest inserted, on osteogenic differentiation of human adipose-derived adult stem cells (hASCs). The influence of these mechanical strains was tested on two hASC lines having different mineral deposition potential, with one cell line depositing approximately nine times as much calcium as the other hASC line after 14 days of culture in osteogenic medium on tissue culture plastic. Results showed that both continuous (10% strain, 1 Hz) and rest inserted cyclic tensile strain (10% strain, 1 Hz, 10 s rest after each cycle) regimens increased the amount and rate of calcium deposition for both high and low calcium depositing hASC lines as compared to unstrained controls. The response was similar for both types of tensile strain for a given cell line, however, cyclic tensile strain had a much stronger osteogenic effect on the high calcium depositing hASC line, suggesting that mechanical loading has a greater effect on cell lines that already have an innate ability to produce bone as compared to cell lines that do not. This is the first study to investigate the osteodifferentiation effects of cyclic tensile strain on hASCs and the first to show that both continuous (10%, 1 Hz) and rest inserted (10%, 1 Hz, 10 s rest) cyclic tensile strain accelerate hASC osteodifferentiation and increase calcium accretion.

Dietary calcium and health

Dietary calcium and health

Calcium kinetics are altered in clinically stable girls with cystic fibrosis.

Reduced bone mass in individuals with cystic fibrosis (CF) may result from alterations in calcium metabolism. Bone calcium deposition and resorption rates, calcium balance, and markers of bone turnover were assessed using stable isotopes of calcium in 22 prepubertal and pubertal girls with CF. Bone calcium deposition was associated with the availability of dietary calcium, total serum osteocalcin, and leptin concentrations. Reduced bone mass in individuals with CF may result from inadequate bone calcium (Ca) deposition, and excessive resorption, although these parameters have not been directly assessed in children with CF. We used stable Ca isotopes to measure rates of bone Ca deposition (Vo+), resorption, and retention in 22 clinically stable girls with CF (aged 7-18 yr). Rates of bone Ca deposition were determined by mathematically modeling the disappearance of iv Ca stable isotope ((42)Ca) for 6 d post dosing. Indirect markers of bone turnover and hormones associated with pubertal development were also assessed. Rates of bone Ca deposition and retention were highest during early puberty (Tanner stages 2 and 3). Calcium deposition rates in prepubertal (Tanner 1) and postmenarchal girls (Tanner stages 4 and 5) did not support substantial bone Ca retention. Net absorption of dietary Ca and serum osteocalcin and leptin concentrations were positively associated with Vo+. Time post menarche and serum leptin concentrations explained 91% of the variability in Vo+ (P = 0.0007). Serum total osteocalcin was low (10.9 +/- 5.4 ng/ml), and a substantial percentage of osteocalcin was undercarboxylated (54.3 +/- 11.8%). We concluded that increased calcium absorption and serum leptin concentrations were significantly associated with rates of bone Ca deposition, demonstrating an impact of nutritional status on this process. Rates of bone Ca deposition were lower than typically reported in healthy children, as were indirect markers of bone formation. These alterations in bone turnover contribute to reduced bone mass in girls with CF.

Normal Acquisition and Loss of Bone Mass

The natural patterns of bone mass accumulation and loss with age represent the templates of individual life cycle periods that are distinguished by marked, physiologically and genetically identifiable, changes in bone mass. During the third trimester of pregnancy, maternal calcium absorption increases and the fetus accumulates about two-thirds of the total bone mass of the term infant. In early infancy, human milk calcium is derived primarily from maternal bone stores, which incur substantial bone losses that are quickly replenished during and after weaning. At puberty, a marked increase in bone mass occurs in conjunction with the initial physical and hormonal changes that characterize this stage. Calcium absorption and bone calcium deposition rates peak in females shortly before menarche. At that time, the bone calcium deposition rate is approximately five times that of adulthood. Skeletal bone mass reaches over 90% of its maximum by age 18 (earlier in females) but does not peak until age 25–30. At some point in mid-life, women experience perimenopause, the 3- to 5-year period prior to menopause during which estrogen levels begin to drop and there are marked increases in bone resorption and loss. Throughout adulthood, calcium absorption efficiency from the diet gradually declines.

Calcium absorption, bone mass accumulation, and kinetics increase during early pubertal development in girls.

To evaluate the changes in calcium and bone mineral metabolism associated with early pubertal development, we performed longitudinal measurements of calcium absorption, calcium kinetics, bone mineral content, and hormonal markers related to puberty in a multiethnic group of girls beginning when they were 7 or 8 yr old. Girls were Tanner stage 1 (breast) at the start of the study. They were placed on a 1200 mg/day dietary calcium intake and studied at approximately 6-month intervals until they reached Tanner stage 2 (breast). Results at that time point (PUB) were compared to values obtained approximately 1 yr earlier (LatePRE) and those 1 yr before that (EarlyPRE). We found an increase in calcium absorption comparing PUB to LatePRE (n = 34; 36.6 +/- 8.7% vs. 30.7 +/- 9.9%; P = 0.002). Using whole body, dual energy, x-ray absorptiometry scanning, we found an increase in calcium gain during the LatePRE to PUB period compared with that during the EarlyPRE to LatePRE period (135 +/- 53 vs. 110 +/- 45 mg/day; P = 0.04). Calcium kinetic studies showed a significant increase in the bone calcium deposition rate (Vo+) during the PUB compared to the LatePRE period. Hormonal and biochemical markers of bone development were also significantly increased at PUB compared to LatePRE. Hormonal activity, as evidenced by the unstimulated LH level, was significantly correlated with calcium gain between the LatePRE and PUB studies and the bone calcium deposition rate in the PUB study. These data demonstrate, using multiple independent methods, an increase in calcium utilization associated with the earliest physical signs of puberty.

Vitamin D receptor gene Fok1 polymorphism predicts calcium absorption and bone mineral density in children.

The vitamin D receptor (VDR) gene has been implicated as one of the major genetic components of osteoporosis. We evaluated the relationship between markers of mineral status and restriction fragment length polymorphisms of the VDR gene in 72 healthy children age 7-12 years. Using stable isotope techniques and dual-energy X-ray absorptiometry, we measured dietary calcium absorption, bone calcium deposition rates, and total body bone mineral density (BMD). The Fok1 polymorphism at the VDR translation initiation site was significantly associated with BMD (p = 0.02) and calcium absorption (p = 0.04). Children who were FF homozygotes had a mean calcium absorption that was 41.5% greater than those who were ff homozygotes and 17% greater absorption than Ff heterozygotes. BMD was 8.2% greater in the FF genotype than the ff genotype and 4.8% higher than the Ff genotype. These results suggest a substantial relationship between the VDR gene and bone metabolism at one or more levels, including dietary absorption of calcium and BMD in growing children.

Development and Regulation of Calcium Metabolism in Healthy Girls

The major components of calcium metabolism, as evaluated by a dual-tracer stable isotope method, were determined in 100 studies of 68 healthy girls, aged 5–18 y and analyzed from a developmental and regulatory viewpoint. Bone calcium deposition and removal rates were closely correlated with the size of the exchangeable bone calcium compartment. All three quantities, as well as intestinal calcium absorption, peaked at or near menarche. Both bone calcium deposition and removal rates were positively and linearly correlated with calcium absorption. However, in this correlation, because bone calcium deposition increased 70% faster than calcium absorption, most of the increase in the bone calcium compartment and its turnover must have occurred in response to something other than intestinal calcium input; presumably this occurred in response to developmental signals. Nevertheless, the constancy of the serum calcium in the face of a large intestinal calcium input and the modest way in which excretion overcame the calcium load in this population point to the importance of the exchangeable bone calcium compartment, in dynamic equilibrium with the bone mineral, as the site at which most of the load is taken up. In this population of girls, as in older women, this increase in the skeletal calcium balance resulted from a decrease in the bone calcium removal rate that was greater than the corresponding increase in the bone calcium deposition rate.

Regulation of calcium and strontium deposition on the otoliths of juvenile tilapia, Oreochromis niloticus

Radioisotopes of calcium and strontium were used to test for a relationship between the environmental concentrations of each element and their respective deposition rates on the otolith. On the basis of 45Ca and 89Sr assays, the rate of strontium deposition on the otolith was influenced by strontium concentrations in the water, while the rate of calcium deposition was affected neither by food nor water calcium concentrations. Thus, the deposition rate of strontium on the otolith at least partially reflects environmental availability, while that of calcium does not. Sources in the water contributed 75% of calcium and 88% of strontium to the sagittae of Nile tilapia, Oreochromis niloticus, with the remainder being provided by the diet.

Promotion of bone repair in the cortical bone of the tibia in rats by low energy laser (He-Ne) irradiation.

The effect of low energy laser (He-Ne) irradiation on bone repair in the cortical part of the tibia of the rat was investigated using biochemical and radioactive labeling methods. A fixed round hole was created in the lateral aspect of the tibia and the newly formed tissue was collected from the gap in the cortical bone. Alkaline phosphatase (ALP) and calcium progressively accumulated at the site of injury, peaking at 9 and 13 days postinjury, respectively. Direct irradiation of the hole injury with He-Ne laser on days 5 and 6 postinjury altered osteoblastic activity at the injured site as reflected by alkaline phosphatase activity. The laser irradiation also caused a significant increase ( approximately 2-fold) in calcium accumulation at the site of injury for 9-18 days postinjury. The rate of calcium deposition, measured by radioactive calcium, was significantly higher (approximately 2-fold) in the laser-irradiated rats as compared with controls. It is concluded that the process of bone repair in a hole created in the rat tibia is markedly enhanced by direct He-Ne laser irradiation of the injured site at the optimal energy level and time postinjury.

Variables related to urinary calcium excretion in young girls.

The relations among dietary and calcium kinetic factors and 24-h urinary mineral excretion were evaluated in a group of 89 healthy girls (51 white and 38 black) aged 4.9-16.7 years. Nutrient intakes were calculated for each participant using a weighed intake of all food and beverage on the day of the 24-h urine collection study and two subsequent 24-h food records. A significant relation was noted between urinary calcium and sodium excretion (r = 0.55; p < 0.0001). No significant relations were found between urinary calcium and (a) calcium intake (r = 0.08), (b) protein intake (r = 0.14), or (c) phosphorus intake (r = 0.11). Urinary calcium was not significantly related to fractional calcium absorption (r = 0.03) or net calcium absorption (r = 0.11), but was significantly associated with the bone calcium deposition rate (r = 0.24; p < 0.03). Using a multiple regression model, both urinary sodium and the bone calcium deposition rate were independent predictors of urinary calcium excretion in this population (r = 0.57; p = 0.0001). A substantial number of the children in this population had urinary calcium excretion > 4 mg/kg/day (12%). The incidence of hypercalciuria differed between the racial groups and was markedly higher in the white than in the black children (17.6 vs. 5.3%). Over a range of usual calcium intakes, during the rapid-bone-growth period in childhood and early adolescence, urinary calcium appears relatively unaffected by calcium intake and is most strongly associated with urinary sodium levels.