Prednisolone Induces an Increase in Serum Calcium Concentration: Possible Involvement of the Kidney, the Bone, and the Intestine

To evaluate the early effect of glucocorticoids on calcium metabolism, 15 subjects aged 22–58 years (5 males, 10 females) with chronic glomerulonephritis were orally treated with 40 mg daily of prednisolone. Five of these subjects were diagnosed with nephrotic syndrome and none had a serum creatinine concentration of more than 1.4 mg/dl. Serum specimens and 24-hour urine specimens were obtained just before and 24 hours after a single oral dose of prednisolone. Serum calcium, ionized calcium, phosphate, intact parathyroid hormone (PTH), intact osteocalcin and 1,25-dihydroxyvitamin D₃ (1,25(OH)₂D₃), and urinary excretion of calcium, phosphate, and deoxypyridinoline were measured. Both serum calcium and ionized calcium concentrations were significantly increased from 4.39 ± 0.10 to 4.47 ± 0.09 mEq/liter (P= 0.037) and from 2.48 ± 0.04 to 2.55 ± 0.04 mEq/liter (P= 0.002), respectively, 24 hours following a single oral dose of prednisolone. Serum intact PTH concentration slightly decreased, but the difference was not significant by statistical analysis. Serum intact osteocalcin concentration was markedly suppressed. In contrast, no significant changes were observed in urinary excretion of deoxypyridinoline. Serum 1,25(OH)₂D₃ concentration measured in five patients was significantly increased. No significant changes in urinary excretion of calcium was observed in the face of these findings. It thus follows that a single oral dose of prednisolone administration increases serum calcium and ionized calcium concentrations, possibly mediated by suppressed bone formation, increased intestinal absorption of calcium, and impaired urinary excretion of calcium. Received: 19 February 1998 / Accepted: 12 March 1999     

Risedronate, a Highly Effective, Short-Term Oral Treatment for Paget's Disease: A Dose-Response Study

Risedronate is a potent pyridinyl bisphosphonate being developed for bone diseases such as Paget's disease and osteoporosis. In this study, we compared the efficacy, safety, and tolerability of three different doses of oral risedronate in 62 patients with severe Paget's disease of bone [serum alkaline phosphatase (AP) >3 times the upper limit of normal]. Patients were treated at six study centers with either 10, 20, or 30 mg oral risedronate daily for 28 days and followed up to day 85. The primary efficacy parameter was percentage change from baseline in AP excess. The data show that there is a dose-response with risedronate: patients who received 30 mg oral risedronate for 28 days benefited most, with a mean percentage decrease in AP excess of 72.2% (20 mg: 57.9%; 10 mg: 48.0%). Time to response—the first time point when there was a ≥30% reduction from baseline in AP excess and ≥50% reduction from baseline in urinary hydroxyproline (HP)/creatinine–was also significantly shorter (median 29 days) in the 30 mg group compared with the other two groups (20 mg: 43 days and 10 mg: 71 days). Long-term follow-up data up to 33 months from the start of the study indicated that AP remained below baseline levels for all patients. Histologic evaluation of bone formed during risedronate therapy demonstrated that normal lamellar bone was formed as opposed to woven pagetic bone, with no evidence of osteomalacia. Risedronate was well tolerated. Transient decreases in serum calcium and increases in serum intact parathyroid hormone were observed, consistent with the pharmacology of risedronate. In conclusion, risedronate administered at daily doses of 10, 20, and 30 mg for 28 days was effective in reducing the biochemical indices of disease activity in patients with severe Paget's disease of bone. A daily dose of 30 mg was most effective without compromising safety or tolerability. Received: 22 January 1998 / Accepted: 27 July 1998     

Osteoporosis and Bone Mineral Metabolism Disorders in Cirrhotic Patients Referred for Orthotopic Liver Transplantation

The purpose of this study was to determine the prevalence of osteoporosis, to estimate the bone turnover and hormonal status, and to identify the factors associated with bone disease in patients with end-stage liver disease who were referred for orthotopic liver transplantation. A prospective study was performed on 58 cirrhotic patients (6 with primary biliary cirrhosis, 14 with alcoholic cirrhosis, and 38 with posthepatitic cirrhosis), who were referred for orthotopic liver transplantation. Patients, excluding those with primary biliary cirrhosis, were classified in Child-Pugh groups according to the severity of liver disease (class B [28 patients], class C [24 patients]). Biochemical parameters of bone mineral metabolism and standard liver function tests were measured in all patients. Additionally, serum osteocalcin, urinary hydroxyproline/creatinine ratio, serum intact parathyroid hormone, serum 25-hydroxyvitamin D, serum 1,25-dihydroxyvitamin D, folliclestimulating hormone, and luteinizing hormone levels were determined in patients and controls within the same age range. Plasma testosterone, sex hormone-binding globulin levels, and free testosterone index were obtained for all men included in the study. Bone mass of the lumbar spine and femur were measured by dual X-ray absorptiometry (DPX-L), and were expressed as a standard deviation of mean values (Z-score) from a sex and age-matched control group. Spinal X-rays were obtained to assess vertebral fractures. Osteoporosis was considered as a factor in spinal bone mineral density with a Z-score below 2 or at least one vertebral fracture. Twenty-five patients (43%) had osteoporosis, with lower bone mass measurements in the lumbar spine than in the femoral neck (P < 0.005). Alcoholic and Child-Pugh C patients showed the lowest femoral bone mineral density values. Cirrhotic patients showed lower osteocalcin levels than controls (14.3 ± 5.9 vs. 18.2 ± 8.1 ng/ml; P < 0.05) and showed increased urinary hydroxyproline (125.1 ± 51.5 vs. 107.9 ± 26.6 nM/mg creatinine; P < 0.05). Serum 25-hydroxyvitamin D, 1,25-dihydroxyvitamin D and parathyroid hormone levels were significantly lower in cirrhotic patients than in controls (10.3 ± 9.1 vs. 23.1 ± 26.6 ng/ml; P= 0.000), (12.9 ± 9.1 vs. 48.3 ± 11.5 pg/ml; P= 0.000), (16.6 ± 9.2 vs. 27.9 ± 8.2 pg/ml; P= 0.000), with no differences between Child-Pugh groups. Alcoholic Child-Pugh C patients showed the lowest 25-hydroxyvitamin D serum values (4.5 ± 2.2 ng/ml; P < 0.05). Male patients had lower testosterone levels than controls (302.5 ± 229.4 vs. 556.7 ± 146.5 ng/dl; P= 0.000), with increased sex hormone-binding globulin values. Levels of testosterone and gonadotropin were related to Child-Pugh classification. No correlation was found between bone mass and hormonal values. A significant decrease in bone mass, particularly in the lumbar spine, is seen in end-stage cirrhotic patients. Reduced bone formation and significant disorders of bone mineral metabolism, such as vitamin D deficiency, reduced parathyroid hormone levels, and hypogonadism are involved. Moreover, severity and etiology of the liver disease are the main risk factors for developing bone loss and mineral metabolism disorders in patients referred for orthotopic liver transplantation. Received: 7 March 1996 / Accepted: 24 June 1996     

Short-Term Effect of Vitamin K Administration on Prednisolone-Induced Loss of Bone Mineral Density in Patients with Chronic Glomerulonephritis

Glucocorticoid-induced osteoporosis has been reported to be caused by enhanced bone resorption and suppressed bone formation. To clarify whether administration of vitamin K, which enhances bone formation, prevents prednisolone-induced loss of bone mineral density (BMD), a randomized, prospective, controlled study was conducted on 20 patients with chronic glomerulonephritis scheduled for treatment with prednisolone. All patients were initially treated with 0.8 mg/kg body weight/day of prednisolone (maximum of 40 mg) for 4 weeks, tapering to 20 mg/day over approximately 6 weeks. Ten patients received prednisolone alone (Group 1), and the other 10 patients received prednisolone plus 15 mg of menatetrenone, vitamin K, three times per day (Group 2). BMD of the lumbar spine measured by dual-energy X-ray absorptiometry (DXA) and biochemical markers of bone metabolism in blood and urine were evaluated before and 10 weeks after administration of prednisolone alone or with menatetrenone. In Group 1, treatment with prednisolone significantly reduced BMD of the lumbar spine from 1.14 ± 0.12 to 1.10 ± 0.11 g/cm² (P= 0.0029). Serum intact osteocalcin and procollagen type I C-peptide (PICP) concentrations, biochemical markers of bone formation, were markedly reduced. A biochemical marker of bone resorption, urinary excretion of deoxypyridinoline, was significantly reduced. In Group 2, prednisolone-induced reduction of BMD was prevented by menatetrenone administration (1.09 ± 0.09 to 1.07 ± 0.07 g/cm², P= 0.153). Menatetrenone prevented reduction of PICP concentration by prednisolone but not in serum intact osteocalcin concentration and urinary excretion of deoxypyridinoline. Thus, treatment with prednisolone resulted in loss of BMD of the lumbar spine associated with suppression of both bone formation and bone resorption. Menatetrenone is a useful agent in preventing prednisolone-induced loss of BMD. Received: 7 July 1998 / Accepted: 13 August 1999     

Influence of Growth Hormone on a New Marker of Cartilage Metabolism (Chondrex)

The aim of this study was to evaluate the usefulness of a major secretory protein of human chondrocytes (chondrex) as a potential serum marker of bone responsiveness to growth hormone (GH). The study included 18 children (10 F, 8 M), aged 10.9 ± 2.3 years, bone age 8.8 ± 2.7 years, height −2.3 ± 0.22 SDS, affected by isolated idiopathic GH deficiency (GHD). Serum samples for evaluation of chondrex, total, and bone alkaline phosphatase were taken before and 3 and 6 months following treatment with rhGH. The basal serum level of chondrex did not differ between patients (37 ± 22 ng/ml) and controls (33 ± 9.8 ng/ml). Following 6 months of treatment with rhGH, a significant increase of height velocity SDS (from −2.8 ± 0.5 to 1.3 ± 0.7), total (from 195 ± 47 to 264 ± 79 U/liter) and bone alkaline phosphatase (from 81 ± 21 to 108 ± 30 U/liter) was observed, while chondrex serum level remained unchanged (from 37 ± 22 to 36 ± 29 ng/ml). It was concluded that chondrex cannot be considered a reliable marker of bone responsiveness to GH in the growing child. Received: 19 March 1999 / Accepted: 3 February 2000     

Increased Serum Levels of N-Telopeptides (NTx) of Bone Collagen in Postmenopausal Nigerian Women

Serum levels of cross-linked N-telopeptides (NTx) of bone collagen, alkaline phosphatase (ALP), and intact parathyroid hormone (PTH) were determined in 64 premenopausal (PRM) and 86 postmenopausal (PSM) women living in northern Nigeria. Serum NTx values were correlated with ALP activity (r = 0.31–0.58, P < 0.01) and PTH (0.32–0.35, P < 0.01)) in all of the subjects studied, and were also related to age (−0.47, P < 0.001) and body mass index (−0.45, P < 0.001) in PRM women. Menopause had the effect of increasing the circulating concentrations of NTx and ALP activity by 15% (P= 0.001) and 11% (P= 0.02), respectively; however, serum levels of PTH were not different between these two groups of women. Compared with Caucasian counterparts matched for age and body mass index, PSM Nigerian women had significantly increased circulating concentrations of NTx (21.7 versus 16.2 nmol BCE/liter, P= 0.01) and demonstrated a trend towards higher ALP activities and PTH levels. These results indicate that (1) discrete reference intervals should be defined for biochemical markers of bone metabolism in African populations, (2) Nigerian women have relatively higher rates of bone turnover, and (3) further investigation of the implications of increased serum NTx should be undertaken using physical methods such as dual X-ray absorptiometry (DXA) and bone ultrasound attenuation. Received: 16 September 1998 / Accepted: 10 January 1999     

A Heterogeneity in Serum Osteocalcin N-Terminal Fragments in Paget's Disease: A Comparison with Other Biochemical Indices in Pre- and Postmenopause

An osteocalcin (OC) nitrogen (N)-terminal sandwich enzyme immunoassay that employs anit-N-20 (amino acids 1–20) polyclonal and monoclonal antibodies has been developed. This assay has demonstrated the existence of a heterogeneity in the OC N-terminal fragments observed in the serum of a patient with Paget's disease and a normal child. The elevation of the serum OC N-terminal value that occurs in Paget's disease was considered to be comparable to the serum alkaline phosphatase (ALP) elevations that also occur. The size of the peaks corresponding to N-terminal OC fragments in Paget's serum decreased 3 months after bisphosphonate treatment. Serum levels of OC N-terminals, and other biochemical indices, were determined for 67 premenopausal and 181 postmenopausal Japanese women. Serum OC N-terminal levels increased significantly (41.2%) in postmenopausal women compared with age-matched premenopausal women. These results strongly suggest that serum OC N-terminal levels reflect bone turnover rates when bone resorption is dominant. Received: 29 April 1997 / Accepted: 8 April 1998     

Fluoride Treatment Increased Serum IGF-1, Bone Turnover,and Bone Mass,but Not Bone Strength,in Rabbits

We hypothesized that fluoride partly acts by changing the levels of circulating calcium-regulating hormones and skeletal growth factors. The effects of oral fluoride on 24 female, Dutch-Belted, young adult rabbits were studied. The rabbits were divided into two study groups, one control and the other receiving about 16 mg fluoride/rabbit/day in their drinking water. After 6 months of fluoride dosing, all rabbits were euthanized and bone and blood samples were taken for analyses. Fluoride treatment increased serum and bone fluoride levels by over an order of magnitude (P < 0.001), but did not affect body weight or the following serum biochemical variables: urea, creatinine, phosphorus, total protein, albumin, bilirubin, SGOT, or total alkaline phosphatase. No skeletal fluorosis or osteomalacia was observed histologically, nor did fluoride affect serum PTH or Vitamin D metabolites (P > 0.4). BAP was increased 37% (P < 0.05) by fluoride; serum TRAP was increased 42% (P < 0.05); serum IGF-1 was increased 40% (P < 0.05). Fluoride increased the vertebral BV/TV by 35% (P < 0.05) and tibial ash weight by 10% (P < 0.05). However, the increases in bone mass and bone formation were not reflected in improved bone strength. Fluoride decreased bone strength by about 19% in the L5 vertebra (P < 0.01) and 25% in the femoral neck (P < 0.05). X-ray diffraction showed altered mineral crystal thickness in fluoride-treated bones (P < 0.001), and there was a negative association between crystal width and fracture stress of the femur (P < 0.02). In conclusion, fluoride's effects on bone mass and bone turnover were not mediated by PTH. IGF-1 was increased by fluoride and was associated with increased bone turnover, but was not correlated with bone formation markers. High-dose fluoride treatment did not improve, but decreased, bone strength in rabbits, even in the absence of impaired mineralization. Received: 5 November 1996 / Accepted: 3 January 1997     

Bone Turnover and Insulin-like Growth Factor I Levels Increase After Improved Glycemic Control in Noninsulin-dependent Diabetes Mellitus

It is unclear whether both bone resorption and formation are affected by glycemic control, and contribute to diabetic osteopenia. In this study, 20 patients with noninsulin-dependent diabetes mellitus (12 men and 8 postmenopausal women) and 20 healthy control subjects (10 men and 10 postmenopausal women) were examined at baseline and 2 months. The diabetic patients showed an improvement of glycemic control (decreased HbA_1c) at the second measurement. Analysis of variance showed that there was no effect of gender on the variables that increased with improved glycemic control, and therefore results are presented for both male and female subjects. Baseline values of serum osteocalcin, a marker of formation, were significantly lower in diabetic patients compared with healthy subjects (2.5 ± 1.3 versus 4.4 ± 1.4 ng/ml; P= 0.0006), but markers of bone resorption [urinary pyridinoline (PYD), deoxypyridinoline (DPD)] did not differ. Improved glycemic control in diabetic patients resulted in increased values of PYD (P= 0.012), DPD (P= 0.049), serum osteocalcin (P= 0.001), and serum insulin-like growth factor I (IGF-I, P= 0.003), but no change in serum parathyroid hormone or 25-hydroxyvitamin D. In diabetic patients there were inverse correlations for the percent change from baseline to improved glycemic control for osteocalcin and HbA_1c (r =−0.53; P= 0.016) and glucose (r =−0.46; P= 0.050). These data suggest that improved glycemic control is accompanied by an increase in bone turnover for male and female diabetic patients, possibly mediated by increased levels of circulating IGF-I. Received: 8 August 1997 / Accepted: 20 January 1998     

Serum Levels of Pyridinium Crosslinks in Postmenopausal Women and in Paget's Disease of Bone

The purpose of this study was to measure pyridinium crosslinks in serum by high performance liquid chromatography (HPLC) and to correlate levels with urinary excretion in patients with different osteometabolic conditions. Blood and spot urine samples were collected between 9 and 11 A.M. in 92 early postmenopausal, untreated women (age 52.3 ± 2.6 years, months since menopause 20.4 ± 9.6), 17 patients with active Paget's disease (10 males, aged 65.1 ± 12.6) and 24 healthy premenopausal women (aged 28.4 ± 4.2). Urinary excretion of the total fraction (free + peptide bound) of pyridinolines (Pyr, Dpyr) was measured by HPLC. Before HPLC analysis, serum samples were submitted to a clean-up procedure by ultrafiltration. In 42 postmenopausal women, bone loss was calculated from two bone mass measurements (L2–L4, DXA), performed at study entry and after 12 months. Statistical analysis was performed by Student's t test for independent samples and linear regression analysis. In pagetic patients' serum levels of Pyr and Dpyr were more than threefold increased over the mean observed in healthy controls and were closely correlated with total alkaline phosphatase levels (Pyr: r = 0.73; Dpyr: r = 0.72, P < 0.0005). Compared with controls, postmenopausal women had significantly increased levels of both urinary and serum Pyr and Dpyr (P < 0.003). In pagetic patients and postmenopausal women, crosslinks serum levels were correlated with their urinary excretion with r values ranging from 0.46 to 0.84. In postmenopausal women, only serum Dpyr correlated with the rate of bone loss (r =−0.36, P= 0.02). The data suggest that serum levels of pyridinium cross-links are correlated with urinary excretion in patients with different osteometabolic conditions. The determination of serum levels prevents limitations related to urinary specimen collection and may be a more practical method for routine application, avoiding corrections for urinary creatinine which could be misleading. Received: 13 August 1996 / Accepted: 25 April 1997     

Relationships Between Bone Mass Measurements and Lifetime Physical Activity in a Swedish Population

Lifetime occupational and leisure time activities were assessed by a questionnaire in order to evaluate their relationship to bone mass measurements and biochemical markers of bone metabolism in a population of 61 women and 61 men, randomly selected from a Swedish population register, to represent ages between 22 and 85 years. We also considered possible confounders by using questions about smoking habits, milk consumption, hormone replacement therapy (HRT), and menopausal age. Bone mineral density (BMD) and bone mineral content (bone mass, BMC) of the total body, lumbar spine, and proximal femur (neck, trochanter, Ward's triangle) were measured by dual energy X-ray absorptiometry (DXA), and BMD of the forearm with single energy X-ray absorptiometry (SXA). In addition, both DXA and SXA provided information on bone area. Quantitative ultrasound measurements (QUS) at the heel were performed to assess the speed of sound (SOS) and broadband ultrasound attenuation (BUA). Fasting blood samples were analyzed for biochemical markers of bone metabolism as well as parathyroid hormone (PTH) and total serum calcium. After adjustment for confounding factors, neither BMD nor QUS measurements were consistently related to lifetime leisure time or occupational activities; nor were there any consistent patterns relating biochemical markers of bone metabolism to bone mass measurements. However, physical activity seemed to influence bone mass, area, and width more than density. In men, high levels of leisure time activity were associated with raised values for lumbar spine area (6.2%) and width (3.3%) as well as for femoral neck area (5.5%) compared with their low activity counterpart. Men exposed to high levels of occupational activity demonstrated lower lumbar spine BMD (10.9%) and area (5.3%) than men with low activity levels. Within an unselected Swedish population, estimation of lifetime occupational and sport activities as well as bedrest, using a questionnaire, demonstrated no major effects on bone density. However, the association between high levels of lifetime activity and raised values for bone mass, area, and width indicate that geometrical changes in bone may provide better estimations of mechanically induced bone strength than bone density, at least in men. Received: 20 May 1997 / Accepted: 15 October 1997     

Risk Factors for the Development of Vertebral and Total Skeleton Osteoporosis in Patients with Primary Biliary Cirrhosis

The objectives of this work was to (1) study the bone mineral density (BMD) of the lumbar spine, total skeleton, and body composition in patients with primary biliary cirrhosis (PBC) and (2) evaluate the risk factors (premature menopause, stages of the disease, hyperbilirubinemia) and bone and liver biochemical parameters for the development of osteoporosis. We studied 23 women with a compatible diagnosis of PBC. The BMD and body composition were evaluated by X-ray absorptiometry (Lunar DPX-L). The average age of the population was 56.7 ± 10.2 years. The BMD of the lumbar spine and of the total skeleton was 1.3 SDs below the normal population matched for sex and age. In the total skeleton, the legs were the most severely affected area (Z score −1.5). The body composition showed no significant difference compared with the normal population. The BMD of 56% of the patients was less than −2.5 SDs from the average normal young values. Patients with a history of vertebral fractures had diminished mineral density of the lumbar spine, as did those who had had no fractures. Of the risk factors studied, patients with premature menopause had a lower bone mass compared with patients with normal menopausal age (Z score of the total skeleton was −2.1 ± 1.8 versus −1.1 ± 1.0) but the difference did not reach statistical significance. The bone mass was not affected in patients with regular menstrual cycles. There were no statistically significant differences in high levels of bilirubin, advanced stages of the disease, or the biochemical variables studied. It is concluded that patients with primary biliary cirrhosis present diminished cortical and trabecular bone mass, whereas body composition was unaffected. Premature hormone deficit, possibly triggered by the chronic hepatic pathology, is a contributing factor to the osteoporosis in this population. Received: 21 October 1997 / Accepted: 5 March 1998     

Does Leptin Have an Effect on Bone in Adult Women?

Recent studies have implicated leptin in the modulation of bone mass during skeletal development. Whether leptin also exerts an influence on bone after growth has stopped is unknown at present. In this cross-sectional study on 94 women (60 premenopausal, 34 postmenopausal) aged 40–60 years, we analyzed the relationship between serum leptin and bone density and bone cortex geometry and bone metabolism. Total and trabecular bone density as well as total and cortical bone area were determined by quantitative computed tomography (QCT) at the distal radius. Bone metabolism was assessed by measuring bone-specific alkaline phosphatase, osteocalcin, procollagen type I C-terminal propeptide (PICP) and collagen type I C-terminal telopeptide in serum, and deoxypyridinoline in urine samples. None of the indices of bone density or geometry was significantly related to leptin serum concentrations (P > 0.05) before or after adjustment for body mass index (BMI). PICP was associated with serum leptin in the postmenopausal group only (r =−0.40 after adjustment for BMI; P= 0.009). Yet, as none of the other markers of bone metabolism exhibited a significant correlation with serum leptin in any of the menopausal groups, this association is likely to be due to the influence of extraskeletal factors on PICP serum levels. Thus, it appears that leptin has less influence on the mature than on the growing skeleton. Received: 5 January 1998 / Accepted: 12 May 1998     

The PTH-Calcium Relationship During a Range of Infused PTH Doses in the Parathyroidectomized Rat

To establish the PTH dosage that maintains normal mineral homeostasis in the PTX rat, a series of doses of rat 1-34 PTH were infused via a subcutaneously implanted miniosmotic pump. The doses were 0, 0.011, 0.022, 0.044, and 0.11 μg/100 g/hour. After 48 hours, serum calcium ranged from 5.56 ± 0.02 to 16.29 ± 0.25 mg/dl, ANOVA P < 0.001, and serum phosphorus from 12.49 ± 0.03 to 5.33 ± 0.34 mg/dl, ANOVA P < 0.001. By post hoc test, the serum calcium level was different (P < 0.05) at every PTH dose; the serum phosphorus level was different (P < 0.05) at every PTH dose except between the two highest doses. The PTH dosage that produced a normal serum calcium (10.09 ± 0.10 mg/dl) and phosphorus (6.90 ± 0.18 mg/dl) was 0.022 μg/100 g/hour. The relationship between increasing doses of PTH and both serum calcium and phosphorus was curvilinear and the calcium-phosphorus product was remarkably constant from a serum calcium of 7–13 mg/dl. The increase in serum calcium and the decrease in serum phosphorus were more rapid at lower than at higher PTH doses so that for both, an asymptote was reached. At the highest serum calcium values, the calcium-phosphorus product increased and in individual rats, an increase in serum phosphorus was associated with a decrease in serum calcium. In summary, this study shows that (1) for rat 1-34 PTH, the normal replacement dose in the PTX rat with normal renal function on a normal diet is 0.022 μg/100 g/hour; (2) the relationship between PTH and both serum calcium and phosphorus is curvilinear, and an asymptote is reached for both; and (3) the calcium-phosphorus product is remarkably constant as the serum calcium increases from 7 to 13 mg/dl and only increased during marked hypercalcemia when serum phosphorus did not decrease further or even tended to increase. Received: 30 May 1997 / Accepted: 15 October 1997     

Changes in Bone Turnover During the Menstrual Cycle in Cynomolgus Monkeys

It is well established that estrogen deficiency at menopause results in increased bone turnover, which is reflected in increased concentrations of markers of bone formation and bone resorption in serum and urine. Since serum 17β-estradiol concentrations vary markedly throughout the menstrual cycle, one would expect to see changes in bone turnover as well. Studies in humans have not yielded consistent results, perhaps because of differences in diet and activity throughout the test period. Therefore, we examined changes in bone biomarkers throughout the menstrual cycle in cynomolgus macaques. Seven intact female cynomolgus macaques (Macaca fascicularis) were evaluated. Vaginal swabs for menstrual blood were performed 3 times/week to determine the stage of the reproductive cycle. Blood and urine were collected at weekly or biweekly intervals for a total of eight samples per monkey for analysis of serum 17β-estradiol, progesterone, parathyroid hormone (PTH), osteocalcin, bone-specific alkaline phosphatase (BSAP), and urinary CrossLaps™. Cycle lengths were determined, and collection days were adjusted to a standardized length of 28 days for all monkeys. Values for bone biomakers were evaluated as % mean for each monkey cycle. By fitting the data to a sine wave (cosinor analysis), serum osteocalcin, BSAP, and urinary CrossLaps demonstrated significant cycles with peaks at days 2.6, 7.3, and 27.8, respectively. Serum osteocalcin and urinary CrossLaps were inversely correlated to serum 17β-estradiol. Urinary CrossLaps were significantly lower in the week just prior to and during ovulation when estradiol was elevated. No rhythm was detected in serum PTH. A peak in bone resorption when serum 17β-estradiol is at its nadir is consistent with the hypothesis that estrogen decreases bone turnover. Received: 19 March 1999 / Accepted: 30 September 1999     

Osteogenesis Imperfecta: Bone Turnover, Bone Density, and Ultrasound Parameters

We studied 21 patients (11 men and 10 women) with osteogenesis imperfecta (OI) and 21 age- and sex-matched controls. In all patients we measured serum levels of total alkaline phosphatase (ALP), type I procollagen carboxy-terminal propeptide (PICP), osteocalcin (BGP), urinary excretion of hydroxyproline (HOP/Cr), and pyridinoline crosslinks (Pyr/Cr). Bone mineral density was measured at the distal radius (BMD-R) and at the lumbar spine (BMD-LS) by dual X-ray absorptiometry (DXA). Ultrasound parameters were also performed at the calcaneous with the Achilles device and at the phalanxes with DBM Sonic 1200. A significant reduction (P < 0.001) in BMD and in ultrasound parameters was found in OI patients compared with normals. PICP was significantly reduced in the OI patients compared with controls (P < 0.001); other markers of bone turnover were higher in OI than in controls, but the difference did not reach the statistical significance. A significant correlation (P < 0.05) was found between PICP and BMD at the lumbar spine and between PICP and ultrasound parameters at the calcaneous. On the basis of our data, we conclude that patients with OI show low values of BMD and ultrasound parameters; therefore in these patients, not only is bone mass disturbed but also bone quality. The reduced levels of PICP in OI patients confirm that most OI patients have defects in collagen I biosynthesis. These defects may contribute to the fragility of OI bone by interfering with complete mineralization and/or normal tissue structure. PICP may be considered a useful marker in the clinical management of OI. Received: 26 March 1998 / Accepted: 15 January 1999     

Geographic Differences in Bone Turnover: Data from a Multinational Study in Healthy Postmenopausal Women

Biochemical markers of bone metabolism (bone markers) are used increasingly to monitor response to therapy and may be predictors of bone loss and fractures. The relationship between fracture rates, which differ between countries, and the rate of bone turnover has not been examined. Therefore, we explored the geographic variability of bone turnover in a selected, healthy study population of 619 postmenopausal women, ages 40–61, participating in a clinical trial of raloxifene hydrochloride for osteoporosis prevention. The subjects were distributed among 38 investigative sites in 10 countries (9–211 subjects/country) on four continents (North America, n = 277, Europe, n = 168, Australia, n = 125, and Africa, n = 49). Specimens for serum osteocalcin (OC), bone-specific alkaline phosphatase (BSAP), and urine type I collagen fragment/urinary creatinine ratio (CTX) were handled in a uniform fashion and assayed in a central laboratory. Mean levels of OC (P < 0.001), BSAP (P= 0.006), and CTX (P < 0.001) varied significantly by country (ANOVA), with the lowest values typically in German and Spanish subjects and the highest in American and Canadian subjects. The consistent pattern and wide ranges of mean bone marker values (OC 1.6-fold, BSAP 1.7-fold, CTX 3.1-fold) between countries suggest clinically significant differences in bone turnover. Geographic differences in bone markers were not explained by the determined potential confounders of age, years posthysterectomy, total serum cholesterol, and serum follicle stimulating hormone (FSH). We conclude that bone marker values vary substantially by country in this selected study population, suggesting systematic geographic differences in bone metabolism that potentially relate to osteoporotic fracture rates. Received: 28 November 1997 / Accepted: 23 March 1998     

Effects of Indomethacin Administration on Bone Turnover and Bone Mass in Adjuvant-Induced Arthritis in Rats

We examined the bone turnover and bone mass in adjuvant-induced arthritis in rats and assessed the effects of indomethacin in this model. One hundred ten SD rats, 6 weeks of age, were assigned to 11 groups and injected with adjuvant or solvent in the right foot. Adjuvant-injected rats were orally administered indomethacin at doses of 0 (vehicle), 0.1 (low), 0.5 (medium), and 1.5 (high) mg/kg body weight from the start (day 0). Animals were sacrificed on days 0, 14 (acute phase), and 28 (chronic phase). In the arthritic-control group, serum osteocalcin level and bone mineral content of the fourth lumbar body (L4) and the femur were significantly reduced on day 14. Serum alkaline-phosphatase was increased on day 28. Trabecular bone volume of L4 was decreased on day 14, and the value was further decreased on day 28. Bone formation rate (BFR/BS) was significantly reduced on day 14, and then osteoclast number (Oc.N/BS) increased on day 28. Indomethacin treatment dose-dependently prevented increases in paw volume and osteoclast number. In the high dose group, these indices were maintained at the same level with those in the normal group. However, indomethacin treatments were not able to maintain the parameters of bone formation such as serum osteocalcin and BFR/BS values, and the trabecular bone mass decrease was only partially prevented. These data clearly indicated both reduced bone formation and increased bone resorption as the causes of bone loss in adjuvant-induced arthritis in rats. Increased bone resorption seemed to be due to the increased activity of prostaglandins, but bone formation defect would be related to other factors in this animal model. Received: 13 January 1996 / Accepted: 3 May 1996     

Comparison of Assay of Total and Bone-Specific Alkaline Phosphatase in the Assessment of Osteoblast Activity in Patients with Metastatic Bone Disease

The evaluation of response of osseous metastases to systemic treatments is often low as a consequence of the different radiologic appearances that make objective assessment not only difficult but sometimes impossible. Radiographic evidence of recalcification, the UICC criterion of response, is often evident for 6 months and sometimes may be delayed even more. This accounts for lower response rates in bone with respect to other metastatic sites in clinical trials. A transient rise in bone formation indices may provide an early indication of bone healing and, along with measurement of symptomatic changes, could ameliorate the response evaluation. Among the biochemical markers of bone formation, total alkaline phosphatase (TALP) is widely employed, but it lacks specificity. Estimation of bone isoenzyme (E-BALP) by electrophoretic techniques is time consuming and semiquantitative. The immunoradiometric assay (I-BALP) seems to overcome these limitations. In this study, we compared the two methods of bone isoenzyme estimation with each other and with the levels of bone gla protein (BGP) and carboxyterminal propeptide of type I procollagen (PICP) in a group of 136 cancer patients with bone metastases stratified as having lytic or mixed and blastic lesions at X-ray, and in 62 cancer patients without apparent bone involvement. The same indices were also evaluated prospectively in a patient subset submitted to chemotherapy associated with pamidronate. The aims of the study were to evaluate whether I-BALP is superior to E-BALP and whether both methods of bone isoenzyme estimation are more advantageous than TALP, BGP, and PICP in the assessment of osteoblast activity either in baseline conditions or in response to treatment. In bone metastatic patients with lytic appearances, values above the cut-off limit were observed in 32.1%, 23.3%, 48.9%, 32.9%, and 14% for, TALP, E-BALP, I-BALP, PICP, and BGP, while the corresponding percentages in those with blastic/mixed appearances were 74.0%, 84.8%, 76.9%, 51.9%, and 43.8%, respectively. In the patients without bone involvement, values within the normal range were 90.2%, 98.2%, 89.6%, 71.7%, and 90.2%, respectively. Levels of TALP, E-BALP, and I-BALP were reciprocally correlated in the three groups examined. In bone metastatic patients, however, the degree of correlation of the enzymes with PICP and BGP was weak. Liver isoenzyme of alkaline phosphatase (LALP) was found to correlate with E-BALP, but not with I-BALP, in patients with mixed/blastic lesions. Thirty-eight patients were submitted to pamidronate therapy (60 mg every 3 weeks, administered 4 times) in association with cytotoxic treatment. Osteoblastic markers were determined before any administration. Serum TALP, E-BALP, and I-BALP showed a transient rise in 9 cases, a progressive reduction in 12, no change in 2, and a progressive increase in 6. Changes in E-BALP and I-BALP from baseline were greater than those of TALP. A divergent pattern between TALP and both I-BALP and E-BALP was found in 9 patients, whereas a divergent temporal profile between the two methods of bone isoenzyme estimation was recorded in only 3 patients. Eight out of 38 cases obtained a partial recalcification of lytic and mixed lesions. Seven of them showed the concomitant early increase in TALP, E-BALP, and I-BALP followed by a gradual decline (osteoblastic flare), whereas 1 patient demonstrated the flare of E-BALP and I-BALP but not of TALP. No relationship was found between response and temporal changes in in BGP and PICP serum levels. We conclude that I-BALP is a useful marker for detecting excess osteoblastic activity in patients who have at imaging ``pure' lytic bone metastases. In the longitudinal evaluation of patients receiving multiple pamidronate infusions plus chemotherapy, TALP, E-BALP, and I-BALP, but not BGP and PICP, appeared to be useful to identify responders in bone. A slight advantage of measurements of serum bone isoenzyme (by both techniques) over TALP is apparent, but this study fails to demonstrate a clear superiority of I-BALP over E-BALP. Received: 12 March 1996 / Accepted: 24 March 1997