Low dietary calcium and obesity: a comparative study in genetically obese and normal rats during early growth.

PURPOSE: A low calcium intake (LCaI) may predispose to obesity, and excessive fat mass may be detrimental to bone. The impact of Ca inadequacy would be greater in subjects predisposed to obesity. LCaI effect on obesity development during the rapid growth period was compared in two strains of rats: spontaneously obese IIMb/ß (O) and Wistar (W). Pregnant rats were fed 0.5% (N) or 0.2% (L) of Ca (OLCa, ONCa, WLCa and WNCa). Male pups were fed the maternal diet until day 60. METHODS: Body composition, lipid profile, glucose homeostasis, 25 hydroxyvitamin D, Ca-phosphorus, and bone metabolism were evaluated. RESULTS: BW and body fat were higher, whereas body protein was lower in OLCa versus ONCa (p < 0.05). OLCa presented the highest body fat, glucose, non-HDL and total cholesterol, TGL, insulin levels, and HOMA-IR, liver weight, and adipose perigonadal plus retroperitoneal pads (p < 0.05). WLCa did not exhibit an increase BW and only showed a slight change in body composition with minor biochemical alterations compared to WNCa (p < 0.05). Osteocalcin, CTX, and proximal tibia and lumbar spine BMDs were lower in O than in W rats fed the same Ca diet (p < 0.05). Body ash and Ca content, and total skeleton BMC/BW were lower in OLCa and WLCa versus their corresponding NCa groups (p < 0.05). CONCLUSION: The negative effect of a low Ca diet on fat mass accumulation and lipid profile may be more evident in rats predisposed to obesity. Nevertheless, low CaI interferes with the normal glucose homeostasis leading to an increase in insulin resistance. Low CaI during early growth may be an obesogenic factor that may persist into adult life and may account for the development of obesity and some of its co-morbidities.

Low protein intake magnifies detrimental effects of ovariectomy and vitamin D on bone.

Protein-induced changes in bone and calcium homeostasis could potentially be greater in the elderly and in women at risk for osteoporosis. We hypothesize that a low protein intake would magnify the negative changes in bone metabolism seen in vitamin D (vitD) insufficiency and/or estrogen deficiency. The present study was undertaken to better understand how a low protein diet along with vitD insufficiency could affect bone metabolism using a rodent ovariectomized (OVX) model. Rats (n = 60) underwent ovariectomy (OVX) or sham operation. The first 15 days after surgery, all rats were fed a standard rodent diet. Thereafter, rats (n = 10/group) were fed a low protein diet (LP; 2.5 %) or a control diet (NP; 12.5 %) with 100 IU% vitD (+D; cholecalciferol) or without vitD (-D) for 45 days. The groups were as follows: SHAM + NP + D (control); SHAM + LP + D; SHAM + LP - D; OVX + NP + D; OVX + LP + D; OVX + LP - D. Body weight (BW) of control and OVX + NP + D groups increased while those feeding the LP diet, independently of vitD feedings, decreased (p < 0.05). The OVX + LP - D group presented the lowest serum Ca, phosphorus and osteocalcin levels and the highest CTX levels (p < 0.05). At the end of the study, total skeleton bone mineral content, proximal tibia bone mineral density, bone volume and trabecular number levels decreased as follows: SHAM + NP + D (controls) > SHAM + LP + D > OVX + NP + D > SHAM + LP - D > OVX + LP + D > OVX + LP - D (p < 0.05). A low protein diet negatively affected bone mass and magnified the detrimental effects of vitD and/or estrogen deficiencies.

Polydextrose Enhances Calcium Absorption and Bone Retention in Ovariectomized Rats.

Purpose. To evaluate the effect of polydextrose (PDX) on Ca bioavailability and prevention of loss of bone mass. Methods. Twenty-four two-month-old ovariectomized rats were fed three isocaloric diets only varied in fiber source and content up to 60 days (FOS group, a commercial mixture of short- and long-chain fructooligosaccharide, OVX group fed AIN 93 diet, and PDX group). A SHAM group was included as control. Apparent Ca absorption percentage (%ABS), changes in total skeleton bone mineral content (tsBMC) and bone mineral density (BMD) and femur BMD, % Bone Volume, Ca and organic femur content, caecal weight, and pH were evaluated. Results. %ABS and caecum weight of PDX and FOS were higher, and caecum pH was lower compared to OVX and SHAM. PDX reached a higher pH and lower caecum weight than FOS possibly because PDX is not completely fermented in the colon. Changes in tsBMC and femur BMD in FOS and PDX were significant lower than SHAM but significantly higher than OVX. % Bone Volume and femur % of Ca in PDX were significantly higher than OVX and FOS but lower than SHAM. Conclusions. PDX increased Ca absorption and prevented bone loss in OVX rats.

High-fat diets affect energy and bone metabolism in growing rats.

BACKGROUND: High-fat diets are usually associated with greater weight (W) gain and body fat (BF). However, it is still unclear whether the type and amount of fat consumed influence BF. Additionally, dietary fat intake may also have consequences on skeletal health. OBJECTIVE: To evaluate in healthy growing rats the effects of high-fat diets and type of dietary fat intake (saturated or vegetable oils) on energy and bone metabolism. METHODS: At weaning, male Wistar rats (n = 50) were fed either a control diet (C; fat = 7% w/w) or a high-fat diet (20% w/w) containing either: soybean oil, corn oil (CO), linseed oil (LO), or beef tallow (BT) for 8 weeks. Zoometric parameters, BF, food intake and digestibility, and total and bone alkaline phosphatase (b-AP) were assessed. Total skeleton bone mineral density (BMD) and content (BMC), BMC/W, spine BMD, and bone volume (static-histomorphometry) were measured. RESULTS: Animals fed BT diet achieved lower W versus C. Rats fed high-fat vegetable oil diets showed similar effects on the zoometric parameters but differed in BF. BT showed the lowest lipid digestibility and BMC. In contrast, high vegetable oil diets produced no significant differences in BMC, BMC/W, BMD, spine BMD, and bone volume. Marked differences were observed for LO and BT groups in b-AP and CO and BT groups in bone volume. CONCLUSION: BT diet rich in saturated fatty acids had decreased digestibility and adversely affected energy and bone metabolisms, in growing healthy male rats. There were no changes in zoometric and bone parameters among rats fed high vegetable oil diets.

Salivary bone turnover markers in healthy pre- and postmenopausal women: daily and seasonal rhythm.

No studies had investigated circadian and circannual rhythms of bone biomarkers in whole saliva. We evaluated the salivary daily and seasonal rhythm of carboxy-terminal telopeptide of type I collagen (CTX) and bone alkaline phosphatase (b-ALP). Forty clinical and oral healthy ambulatory pre- and postmenopausal women from two southern Argentine cities: Comodoro Rivadavia (latitude 45º S) and Ushuaia (latitude 54º S) were included in the study. CTX levels were evaluated in serum, urine, and saliva, and b-ALP levels were measured in serum and saliva. In both groups of women, salivary CTX showed a maximum percentage of change early in the morning (80%) and a minimum in the late afternoon (45%), similarly to the pattern observed in urinary samples. No daily rhythm was observed in serum or salivary b-ALP. 25-Hydroxyvitamin D levels decreased in winter vs. summer (p < 0.01) without differences between the two studied groups. Conversely, parathormone reached higher levels in winter (p < 0.05) which induced a slight non-significant increment in salivary CTX and b-ALP levels. The results showed that, as in serum and urinary samples, salivary CTX exhibits daily and a slight seasonal rhythmicity. Whole non-stimulated saliva is a useful tool to detect several oral and systemic diseases because it has important advantages compared to serum and urinary samples. Then, it may also be a promising sample to test changes in bone metabolism contributing to diagnose and to monitor the therapy of several metabolic bone diseases.

Comparative 25-OH-vitamin D level in institutionalized women older than 65 years from two cities in Spain and Argentina having a similar solar radiation index.

OBJECTIVE: The present study evaluated and compared vitamin D nutritional status and calcium-phosphorus metabolism in institutionalized women >65 y from two cities that have a similar sun irradiation index (heliophany). METHODS: The study was carried out in women living in similar social-status institutions from geographic cities having a similar solar radiation index (Lleida, Spain, n=49, and suburban Buenos Aires, Argentina [BA], n=48) at the end of summer. RESULTS: Fish consumption was higher in the Lleida group, as was red-meat consumption in the BA group. In both groups mean calcium intake was 800 mg/d. The daily intake of vitamin D was higher in the BA group (P<0.001). A total of 90% in Lleida and 86% in BA had 25-hydroxyvitamin D (25OHD) levels <20 ng/mL. A significant inverse correlation between individual 25OHD and parathyroid hormone (PTH) levels was observed in the two groups of women (r=-0.329, P=0.035). PTH levels >100 pg/mL were found in 24% and 20% of women in Lleida and BA, respectively. There was a marked increase in carboxy-terminal telopeptide cross-links of type I collagen levels and a decrease in 25OHD with an increase in PTH levels (P<0.05). Conversely, bone alkaline phosphatase increased significantly only when the PTH concentration duplicated the reference range. CONCLUSION: Even at the end of summer, vitamin D deficiency/insufficiency was prevalent in the two studied institutionalized elderly women. In the narrow range of the dietary calcium intake (close to 800 mg/d) of both studied groups, secondary hyperparathyroidism was absent when 25OHD levels were >17 ng/dL, indicating changes in the regulation control of serum PTH and consequently the changes in this threshold. As a result, vitamin D deficiency must be reversed to avoid the increment in bone turnover and to ensure the endocrine and paracrine functions of vitamin D for overall health and well-being.

Correlation between salivary and serum markers of bone turnover in osteopenic rats.

BACKGROUND: Previously, we measured bone alkaline phosphatase (b-ALP) and terminal C-telopeptide of collagen type I (CTX) in saliva. The present longitudinal experimental study sought to determine whether salivary concentrations of b-ALP and CTX have the same response as in serum samples under different conditions: normal, increased, and reduced bone remodeling. METHODS: Thirty rats were ovariectomized (OVX) to induce osteopenia 60 days after surgery, and 10 rats were sham operated. Then, the rats were divided into four groups and treated as follows for 45 days: group 1 (G1) = SHAM + vehicle; group 2 (G2) = OVX + 8 microg olpadronate (OPD)/100 g of body weight; group 3 (G3) = OVX + 4 microg OPD/100 g of body weight; and group 4 (G4) = OVX + vehicle. Saliva and serum CTX and b-ALP were determined at 60 days (baseline) and at 75 days (T(75)). Lumbar spine and proximal tibia bone mineral density (BMD) was determined using dual-energy x-ray absorptiometry at baseline and at 105 days. RESULTS: SHAM baseline and T(75) salivary b-ALP and CTX levels correlated with serum concentrations (P <0.01 and P <0.004, respectively). A correlation was observed between saliva and serum concentrations of b-ALP and CTX in OVX at baseline (P <0.0001 and P <0.004, respectively). Baseline salivary b-ALP and CTX levels were lower in SHAM animals compared to OVX groups (P <0.01). After treatment, T(75) saliva and serum CTX remained higher in G4 compared to G1 (P <0.05), was lower in G2 than in G1 (P <0.01) and G3 (P <0.01), and was similar in G1 and G3. Changes in BMD were the result of variations in salivary CTX levels due to OPD treatment (P <0.05). CONCLUSIONS: Saliva determinations may prove to be practical and reliable for the detection of systemic signs of increased bone remodeling, particularly in cases involving pediatric, obese, and elderly patients, and in screening large populations. Moreover, saliva CTX may be one of the best candidate markers to detect the activity and severity of periodontal disease.

[Bone remodeling markers in saliva as compared to serum in rats]. / Marcadores del remodelamiento oseo en saliva y su correlación con los niveles sanguíneos en ratas.

Bone markers are useful tools to measure bone remodeling; currently they are assessed in serum and urinary samples; however there is little information concerning their measurement in saliva. The present experimental study evaluates the possibility to measure collagen type I carboxiterminal telopeptide (CTX) and bone alkaline phosphatase (b-AP) in saliva, its correlation with serum samples in normal conditions and in the increase of the bone remodeling due to estrogen deficiency. Twenty four normal adult Wistar rats (300 +/- 20 g) [12 SHAM and 12 rats after 1 week of bilateral ovariectomy (OVX)] were studied. Fasting serum and total saliva after stimulation with pilocarpine were collected. In both samples were measured: CTX (ng/ml) by ELISA (RatLabs, Osteometer Bio Tech, Denmark) and b-AP (IU/L) (Wiener, colorimetrically). Both CTX and b-AL in serum samples were significantly higher in OVX than in SHAM rats (15.3 +/- 4.0 vs. 21.8 +/- 6.4, p < 0.05 y 71 +/- 29 vs. 104 +/- 23; p < 0.01, respectively). Saliva presented the same behaviour (3.6 +/- 0.5 vs. 6.4 +/- 2.9; p < 0.02 y 73 +/- 29 vs. 90 +/- 8; p < 0.003, respectively). When saliva CTX and b-AP were plotted against serum concentration significant positive correlations were obtained: r = 0.58, p < 0.05 and r = 0.59; p < 0.05, respectively. In conclusion, the present results are promisory in the sense of the potential use of a salivary-based test for evaluating bone remodeling. However, the use of this methodology for clinical practice needs extensive additional investigations.

Vitamin D insufficiency reduces the protective effect of bisphosphonate on ovariectomy-induced bone loss in rats.

The present study was carried out to obtain an experimental model of vitamin D (vit D) insufficiency and established osteopenia (experiment 1) to then investigate whether vit D status, i.e. normal or insufficient, interferes with bone mass recovery resulting from bisphosphonate therapy (experiment 2). Rats (n = 40) underwent OVX (n = 32) or a sham operation (n = 8). The first 15 days post-surgery, all groups were kept under fluorescent tube lighting and fed a diet containing 200 IU% vit D (+D). They were then assigned during an additional 45 days to receive either +D or a diet lacking vit D (-D) and kept under 12 h light/dark cycles using fluorescent or red lighting. Serum 25HOD was significantly lower in -D rats (P < 0.0001). The type of lighting did not induce differences in 25OHD, calcium (sCa), phosphorus (sP), bone alkaline phosphatase (b-AL), CTX, bone density or histology. No osteoid was observed in undecalcified bone sections. Experiment 2 (105 days): rats were fed either +D or -D according to experiment 1 and were treated with either placebo or 16 mug olpadronate (OPD)/100 g rat/week during the last 45 days. Whereas 25HOD was significantly lower (P < 0.0001) in -D/OPD than in +D/OPD rats, no significant differences in sCa, sP, b-AL or CTX were observed. OPD prevented the loss of lumbar spine (LS) and proximal tibia (PT) BMD and the decrease in bone volume (BV/TV) (P < 0.05) and in the number of trabeculae observed in untreated rats. However, +D/OPD animals presented significantly higher values of LS BMD, PT BMD and BV/TV than -D/OPD rats (P < 0.05). No osteoid was observed in undecalcified sections of bone. In summary, this is the first experimental study to provide evidence that differences in vit D status may affect the anticatabolic response to bisphosphonate treatment. However, the molecular mechanism through which vit D insufficiency reduces the effect of the aminobisphosphonate remains to be defined.

Marcadores del remodelamiento oseo en saliva y su correlacion con los niveles sanguineos en ratas / Bone remodeling markers in saliva as compared to serum in rats

Si bien es conocida la utilidad de marcadores óseos en suero u orina para determinar cambios en el remodelamiento óseo, la misma no ha sido totalmente estudiada en saliva. Este trabajo evalúa la correlación entre dos marcadores del recambio óseo: la fosfatasa alcalina ósea (isoforma ósea, FAO) y el telopéptido C-terminal del colágeno tipo I (CTX), medidos simultáneamente en suero y saliva de ratas Wistar (250 a 300 g), SHAM (n=12) y ovariectomizadas (OVX) (n=12). Luego de una semana de la cirugía se extrajo sangre en ayunas y saliva total estimulada donde se evaluó CTX (ELISA, RatLabs, Osteometer Bio Tech, Dinamarca) y FAO (Wiener, colorimetría). En el suero, tanto CTX (ng/ml) como FAO (UI/l) en ratas OVX fueron significativamente mayores que en ratas SHAM (15.3±4.0 vs. 21.8±6.4, p<0.05 y 71±29 vs. 104±23; p<0.01, respectivamente). En saliva se observó el mismo comportamiento (3.6±0.5 vs. 6.4±2.9; p<0.02 y 73±29 vs.90±8; p<0.003, respectivamente). Las concentraciones de CTX y FAO en saliva se correlacionaron positivamente con los respectivos niveles sanguíneos (r= 0.58, p<0.05 y r= 0.59, p<0.05, respectivamente). Los resultados del presente estudio preliminar y la sencillez en la obtención de la saliva total ofrecerían una alternativa no invasiva al suero para la medición del remodelamiento óseo. Sin embargo, la aplicación de esta metodología a la clínica humana requiere extensas investigaciones adicionales.

Bone markers are useful tools to measure bone remodeling; currently they are assessed in serum and urinary samples; however there is little information concerning their measurement in saliva. The present experimental study evaluates the possibility to measure collagen type I carboxiterminal telopeptide (CTX) and bone alkaline hosphatase (b-AP) in saliva, its correlation with serum samples in normal conditions and in the increase of the bone remodeling due to estrogen deficiency. Twenty four normal adult Wistar rats (300±20 g) [12 SHAM and 12 rats after 1 week of bilateral ovariectomy (OVX)] were studied. Fasting serum and total saliva after stimulation with pilocarpine were collected. In both samples were measured: CTX (ng/ml) by ELISA (RatLabs, Osteometer Bio Tech, Denmark) and b-AP (IU/L) (Wiener, colorimetrically). Both CTX and b-AL in serum samples were significantly higher in OVX than in SHAM rats (15.3±4.0 vs. 21.8±6.4, p<0.05 y 71±29 vs. 104±23; p<0.01, respectively). Saliva presented the same behaviour (3.6±0.5 vs. 6.4±2.9; p<0.02 y 73±29 vs. 90±8; p<0.003, respectively). When saliva CTX and b-AP were plotted against serum concentration significant positive correlations were obtained: r=0.58, p<0.05 and r=0.59; p<0.05, respectively. In conclusion, the present results are promisory in the sense of the potential use of a salivary-based test for evaluating bone remodeling. However, the use of this methodology for clinical practice needs extensive additional investigations.

Marcadores del remodelamiento oseo en saliva y su correlacion con los niveles sanguineos en ratas / Bone remodeling markers in saliva as compared to serum in rats

Si bien es conocida la utilidad de marcadores óseos en suero u orina para determinar cambios en el remodelamiento óseo, la misma no ha sido totalmente estudiada en saliva. Este trabajo evalúa la correlación entre dos marcadores del recambio óseo: la fosfatasa alcalina ósea (isoforma ósea, FAO) y el telopéptido C-terminal del colágeno tipo I (CTX), medidos simultáneamente en suero y saliva de ratas Wistar (250 a 300 g), SHAM (n=12) y ovariectomizadas (OVX) (n=12). Luego de una semana de la cirugía se extrajo sangre en ayunas y saliva total estimulada donde se evaluó CTX (ELISA, RatLabs, Osteometer Bio Tech, Dinamarca) y FAO (Wiener, colorimetría). En el suero, tanto CTX (ng/ml) como FAO (UI/l) en ratas OVX fueron significativamente mayores que en ratas SHAM (15.3±4.0 vs. 21.8±6.4, p<0.05 y 71±29 vs. 104±23; p<0.01, respectivamente). En saliva se observó el mismo comportamiento (3.6±0.5 vs. 6.4±2.9; p<0.02 y 73±29 vs.90±8; p<0.003, respectivamente). Las concentraciones de CTX y FAO en saliva se correlacionaron positivamente con los respectivos niveles sanguíneos (r= 0.58, p<0.05 y r= 0.59, p<0.05, respectivamente). Los resultados del presente estudio preliminar y la sencillez en la obtención de la saliva total ofrecerían una alternativa no invasiva al suero para la medición del remodelamiento óseo. Sin embargo, la aplicación de esta metodología a la clínica humana requiere extensas investigaciones adicionales.(AU)

Bone markers are useful tools to measure bone remodeling; currently they are assessed in serum and urinary samples; however there is little information concerning their measurement in saliva. The present experimental study evaluates the possibility to measure collagen type I carboxiterminal telopeptide (CTX) and bone alkaline hosphatase (b-AP) in saliva, its correlation with serum samples in normal conditions and in the increase of the bone remodeling due to estrogen deficiency. Twenty four normal adult Wistar rats (300±20 g) [12 SHAM and 12 rats after 1 week of bilateral ovariectomy (OVX)] were studied. Fasting serum and total saliva after stimulation with pilocarpine were collected. In both samples were measured: CTX (ng/ml) by ELISA (RatLabs, Osteometer Bio Tech, Denmark) and b-AP (IU/L) (Wiener, colorimetrically). Both CTX and b-AL in serum samples were significantly higher in OVX than in SHAM rats (15.3±4.0 vs. 21.8±6.4, p<0.05 y 71±29 vs. 104±23; p<0.01, respectively). Saliva presented the same behaviour (3.6±0.5 vs. 6.4±2.9; p<0.02 y 73±29 vs. 90±8; p<0.003, respectively). When saliva CTX and b-AP were plotted against serum concentration significant positive correlations were obtained: r=0.58, p<0.05 and r=0.59; p<0.05, respectively. In conclusion, the present results are promisory in the sense of the potential use of a salivary-based test for evaluating bone remodeling. However, the use of this methodology for clinical practice needs extensive additional investigations. (AU)

Marcadores del remodelamiento oseo en saliva y su correlacion con los niveles sanguineos en ratas / Bone remodeling markers in saliva as compared to serum in rats

Si bien es conocida la utilidad de marcadores óseos en suero u orina para determinar cambios en el remodelamiento óseo, la misma no ha sido totalmente estudiada en saliva. Este trabajo evalúa la correlación entre dos marcadores del recambio óseo: la fosfatasa alcalina ósea (isoforma ósea, FAO) y el telopéptido C-terminal del colágeno tipo I (CTX), medidos simultáneamente en suero y saliva de ratas Wistar (250 a 300 g), SHAM (n=12) y ovariectomizadas (OVX) (n=12). Luego de una semana de la cirugía se extrajo sangre en ayunas y saliva total estimulada donde se evaluó CTX (ELISA, RatLabs, Osteometer Bio Tech, Dinamarca) y FAO (Wiener, colorimetría). En el suero, tanto CTX (ng/ml) como FAO (UI/l) en ratas OVX fueron significativamente mayores que en ratas SHAM (15.3±4.0 vs. 21.8±6.4, p<0.05 y 71±29 vs. 104±23; p<0.01, respectivamente). En saliva se observó el mismo comportamiento (3.6±0.5 vs. 6.4±2.9; p<0.02 y 73±29 vs.90±8; p<0.003, respectivamente). Las concentraciones de CTX y FAO en saliva se correlacionaron positivamente con los respectivos niveles sanguíneos (r= 0.58, p<0.05 y r= 0.59, p<0.05, respectivamente). Los resultados del presente estudio preliminar y la sencillez en la obtención de la saliva total ofrecerían una alternativa no invasiva al suero para la medición del remodelamiento óseo. Sin embargo, la aplicación de esta metodología a la clínica humana requiere extensas investigaciones adicionales.(AU)

Bone markers are useful tools to measure bone remodeling; currently they are assessed in serum and urinary samples; however there is little information concerning their measurement in saliva. The present experimental study evaluates the possibility to measure collagen type I carboxiterminal telopeptide (CTX) and bone alkaline hosphatase (b-AP) in saliva, its correlation with serum samples in normal conditions and in the increase of the bone remodeling due to estrogen deficiency. Twenty four normal adult Wistar rats (300±20 g) [12 SHAM and 12 rats after 1 week of bilateral ovariectomy (OVX)] were studied. Fasting serum and total saliva after stimulation with pilocarpine were collected. In both samples were measured: CTX (ng/ml) by ELISA (RatLabs, Osteometer Bio Tech, Denmark) and b-AP (IU/L) (Wiener, colorimetrically). Both CTX and b-AL in serum samples were significantly higher in OVX than in SHAM rats (15.3±4.0 vs. 21.8±6.4, p<0.05 y 71±29 vs. 104±23; p<0.01, respectively). Saliva presented the same behaviour (3.6±0.5 vs. 6.4±2.9; p<0.02 y 73±29 vs. 90±8; p<0.003, respectively). When saliva CTX and b-AP were plotted against serum concentration significant positive correlations were obtained: r=0.58, p<0.05 and r=0.59; p<0.05, respectively. In conclusion, the present results are promisory in the sense of the potential use of a salivary-based test for evaluating bone remodeling. However, the use of this methodology for clinical practice needs extensive additional investigations. (AU)