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How do firm-level collective agreements affect firm performance in a multi-level bargaining system? Using detailed Belgian linked employer-employee panel data, our findings show that firm agreements increase both wage costs and labour productivity (with respect to sector-level agreements). Relying on a recent approach developed by Bartolucci (2014), they also indicate that firm agreements exert a stronger impact on wages than on productivity, so that on average profitability is hampered. However, this rent-sharing effect only holds in sectors where firms are more concentrated. Firm agreements are thus mainly found to raise wages beyond labour productivity when the rents to be shared between workers and firms are relatively big. Overall, this suggests that firm-level agreements benefit both employers and employees – through higher productivity and wages – without being detrimental to firms’ gross profits.

Periostin is a matricellular protein involved in bone formation and bone matrix organization, but it is also produced by other tissues. Its circulating levels have been weakly associated with bone microstructure and prevalent fractures, possibly because periostin measured by the current commercial assays does not specifically reflect bone metabolism. In this context, we developed a new ELISA for a periostin fragment resulting from cathepsin K digestion (K-Postn). We hypothesized that circulating K-Postn levels could be associated with bone fragility. A total of 695 women (age 65.0 ± 1.5 years), enrolled in the Geneva Retirees Cohort (GERICO), were prospectively evaluated over 4.7 ± 1.9 years for the occurrence of low-trauma fractures. At baseline, we measured serum periostin, K-Postn, and bone turnover markers (BTMs), distal radius and tibia microstructure by HR-pQCT, hip and lumbar spine aBMD by DXA, and estimated fracture probability using the Fracture Risk Assessment Tool (FRAX). Sixty-six women sustained a low-trauma clinical fracture during the follow-up. Total periostin was not associated with fractures (HR [95% CI] per SD: 1.19 [0.89 to 1.59], p = 0.24). In contrast, K-Postn was significantly higher in the fracture versus nonfracture group (57.5 ± 36.6 ng/mL versus 42.5 ± 23.4 ng/mL, p

The article explores the link between different institutional features of minimum wage systems and the minimum wage bite. We notably address the striking absence of studies on sectoral-level minima and exploit unique data covering 17 European countries and information from more than 1,100 collective bargaining agreements. Results provide evidence for a neglected trade-off: systems with bargained sectoral-level minima are associated with higher Kaitz indices than systems with statutory floors, but also with more individuals actually paid below prevailing minima. Higher collective bargaining coverage can, to some extent, reduce this trade-off between sharp teeth (high wage floors) and empty mouths (non-compliance/non-coverage).

Periostin (Postn) and transforming growth factor β-induced protein (TGFβIp) are two closely related extracellular matrix (ECM) proteins predominantly distributed in collagen-rich connective tissues submitted to mechanical strain, including bone and more specifically the periosteum. We have investigated the expression of Postn and TGFβIp mRNA by primary osteoblasts isolated from mouse periosteum and calvaria, or by the osteoblast-like MC3T3-E1 cell line, and by osteoclasts from mouse long bones differentiated in vitro. Secretion of Postn was measured with a specific ELISA. Postn and TGFβIp mRNA were concomitantly expressed in all three osteoblast models all along the differentiation process in a time-dependent manner. Both Postn and TGFβIp transcripts appeared early in osteoblast differentiation, and their expression increased 3-10 times in mature osteoblast cells. Expression decreased after differentiation was achieved and when the cultures mineralised. ELISA for secreted Postn showed a similar pattern. When MC3T3-E1 cells were treated with TGF-β, Postn and TGFβIp mRNA expression and secretion were stimulated, whereas 1.25(OH)(2)D(3) had no detectable effect. Osteoclasts also expressed both Postn and TGFβIp during in vitro differentiation. Expression of both Postn and TGFβIp peaks in the early phases of osteoblast differentiation, and decreases later at the start of mineralisation. A novel finding is that Postn and TGFβIp are expressed by osteoclasts in vitro. Therefore Postn and TGFβIp proteins are potential biomarkers of early osteoblast differentiation and new bone formation.

Endogenous morphine and its derivatives (morphine-6-glucuronide [M6G]; morphine-3-glucuronide [M3G]) are formed by mammalian cells from dopamine. Changes in the concentrations of endogenous morphine have been demonstrated in several pathologies (sepsis, Parkinson's disease, etc.), and they might be relevant as pathological markers. While endogenous morphine levels are detectable using enzyme-linked immunosorbant assay (ELISA), mass spectrometry (MS) analysis was, so far, the only approach to detect and quantify M6G. This study describes the preparation of a specific anti-M6G rabbit polyclonal antibody and its validation. The specificity of this antibody was assessed against 30 morphine-related compounds. Then, a M6G-specific ELISA-assay was tested to quantify M6G in the plasma of healthy donors, morphine-treated, and critically ill patients. The antibody raised against M6G displays a strong affinity for M6G, codeine-6-glucuronide, and morphine-3-6-glucuronide, whereas only weak cross-reactivities were observed for the other compounds. Both M6G-ELISA and LC-MS/MS approaches revealed the absence of M6G in the plasma of healthy donors (controls, n = 8). In all positive donors treated with morphine-patch (n = 5), M6G was detected using both M6G-ELISA and LC-MS/MS analysis. Finally, in a study on critically ill patients with circulating endogenous morphine (n = 26), LC-MS/MS analysis revealed that 73% of the positive-patients (19 of 26), corresponding to high M6G-levels in M6G-ELISA, contained M6G. In conclusion, we show that endogenous M6G can be found at higher levels than morphine in the blood of morphine-naive patients. With respect to the interest of measuring endogenous M6G in pathologies, we provide evidences that our ELISA procedure represents a powerful tool as it can easily and specifically detect endogenous M6G levels.

Summary Objective Osteocalcin is a bone-specific protein secreted by osteoblasts and often used as a bone formation biomarker. Rodent studies have reported a hormonal role of osteocalcin on glucose metabolism, increasing insulin secretion and sensitivity and increasing energy expenditure. However, it is unknown whether osteocalcin fulfils the same function in humans. Methods We investigated the relationship between serum osteocalcin and insulin concentrations in 27 prepubertal obese children (9–12 years old) randomly divided into two groups, one of which entered a physical training programme, and 16 nonobese control children. Whole body bone mineral density (WB-BMD), serum osteocalcin, circulating insulin and adiponectin were measured at baseline and after 6 months. Results Trained and untrained obese children had higher WB-BMD than controls at baseline. Trained children also displayed a significant insulin increase and a significant adiponectin decrease while osteocalcin was increased compared to untrained obese children. Significant linear correlations between WB-BMD and adiponectin, delta BMD (variation between baseline and after-training values) and delta adiponectin, insulin and osteocalcin, delta insulin and delta osteocalcin, delta insulin and delta under-carboxylated osteocalcin were found only in trained obese children with no significant relationship in control and untrained obese children. Conclusions In trained obese children, correlations indicate that when BMD is increased, osteocalcin is increased and insulin lowered. This suggests that increased BMD is associated with increased energy metabolism and a decreased level of insulin. We thus report statistically significant relationships between the skeleton (osteocalcin) and energy metabolism (insulin), suggesting a regulatory hormonal loop including osteocalcin and insulin.

Surrogate markers of fracture risk--bone turnover markers (BTMs) and bone mineral density (BMD)--can be used to monitor treatment response. We assessed whether changes in these markers greater than the least significant change (LSC) were associated with fracture risk reduction and greater adherence. This secondary analysis of the Improving Measurements of Persistence on ACtonel Treatment (IMPACT) study--a multinational prospective, open-label, cluster-randomized study of postmenopausal women on oral risedronate 5 mg/d for 52 weeks-assessed adherence by electronic monitors. Urinary N-terminal cross-linked telopeptide of type 1 collagen (uNTX) and serum C-terminal cross-linked telopeptide of type 1 collagen (sCTX) levels were assessed at baseline and weeks 10 and 22, and BMD at baseline and week 52. Fractures were recorded as adverse events. In 2302 women, responses beyond LSC in BTMs (uNTX and sCTX) and BMD (spine only) were associated with a reduced risk of nonvertebral fractures (NVFs) and all fractures. NVF incidence was about 50% lower in patients with 30% or more of uNTX reduction at week 22 (1.6%) than in those with less than 30% reduction (3.2%, p = .015). NVFs also were reduced in patients with more than 3% spine BMD increase at 52 weeks than those with 3% or less. Responses greater than LSC in BTMs and BMD were associated with greater adherence, but there was no association between adherence and fracture outcomes at 52 weeks. Changes greater than the LSC in BTMs and BMD reflect better treatment adherence, were associated with fracture risk reduction, and identify differences in individual responsiveness to risedronate.

Periostin, a matricellular protein, is overexpressed in the stroma of several cancers. The aim of our study was to investigate more specifically whether periostin expression is associated with bone metastases from breast cancer and to determine its source in the affected bone. Nude mice were inoculated with human MDA-B02 breast cancer cells. Bone metastases-bearing mice were treated with zoledronic acid-an antiresorptive drug-or vehicle. Bone metastases were examined for tumor- and stroma-derived periostin expression by quantitative polymerase chain reaction with human- and mouse-specific primers and immunohistochemistry. Serum periostin and conventional bone turnover markers were also measured. MDA-B02 cells did not express periostin both in vitro and in vivo. However, mouse-derived periostin was markedly overexpressed (eightfold) in metastatic legs compared to noninoculated mice. Serum periostin levels were also markedly increased in metastatic mice and correlated with in situ expression levels. Immunostaining showed that periostin derived from the environing stromal cells of bone metastasis. Bone turnover blockade by zoledronic acid markedly decreased osteolytic lesions but only slightly modulated serum periostin levels. Bone metastases from breast cancer induce overexpression of periostin by surrounding stromal cells. Periostin could be a biochemical marker of the early stromal response associated to breast cancer bone metastasis formation.

Multiple myeloma (MM) is a malignant disorder characterized by the accumulation of neoplastic plasma cells in the bone marrow. Most patients suffer from osteolytic lesions that cause pain and reduced quality of life, and may result in hypercalcemia, fractures, and spinal root or spinal cord compression. At diagnosis 80% of patients already have pathological bone findings when evaluated with conventional radiography (1). MM causes increased bone resorption through osteoclast activation (2–6), and bone resorption is increased even in patients without osteolytic lesions (7). In addition, MM also causes decreased bone formation (8–11). Bone formation is primarily impaired in patients with overt osteolysis, whereas no decrease in bone formation is observed in MM without osteolysis or in monoclonal gammopathy of undetermined significance (MGUS) (7, 12). Likewise, osteoblasts from patients with MM with bone disease are more prone to apoptosis compared to osteoblasts from patients with MM without bone disease (13). Thus, it appears that osteolysis becomes apparent only when increased bone resorption is not compensated for by the formation of new bone (i.e. ‘imbalanced remodeling’). Bisphosphonates, which are the only registered drugs for the treatment of osteolysis in MM (14), mainly target the increased bone resorption caused by osteoclasts (15). The very significant inhibition of osteoclasts obtained by treatment with potent bisphosphonates may in fact inhibit bone formation as some osteoclast activity seems to be important for the stimulation of osteoblasts (16). Indeed, a 50% reduction of osteoblast activity has been observed after initiation of bisphosphonate treatment (17). Accordingly, the protection against osteolysis offered by bisphosphonate treatment is only partial (18). Furthermore, osteoblast function remains impaired when MM has been brought into remission with standard chemotherapy (19), and healing of established bone lesions is rarely observed (20). Evidence suggesting that bortezomib, the first clinically available proteasome inhibitor, could overcome the osteoblast inhibition observed in MM was provided from the APEX trial where serum alkaline phosphatase (AP) was found to be increased in patients responding to bortezomib treatment (21). It was clearly demonstrated that the stimulatory effect on bone formation was induced by bortezomib as no increase in AP was observed in patients responding to the treatment arm lacking bortezomib. Subsequent clinical studies confirmed and extended this observation, showing that the increase in AP was caused by the bone-specific alkaline phosphatase (bALP) (22–24). Cell culture studies on human as well as murine osteoblast cell lines and animal studies have demonstrated that bortezomib stimulates osteoblasts and bone formation (25–30). Furthermore, Giuliani et al. (31) demonstrated that bortezomib treatment increases the number of osteoblasts in the bone marrow of patients with MM. So far, however, the clinical studies have been performed in patients with relapsing MM who have received prior chemotherapy, treatment with bisphosphonates, and frequently have also been treated with glucocorticoids, which are known to inhibit bone formation (32, 33). In this study, we report that bortezomib used alone and in combination with a glucocorticoid as front-line treatment in previously untreated, bisphosphonate-naive patients stimulates bone formation not only as shown by an increase in bALP but also through the use of a novel bone marker, pro-collagen type I N-terminal propeptide (PINP) that is released as collagen is deposited and becomes insoluble during the formation of the organic bone matrix (34) and may, therefore, be a better marker for ongoing bone formation. We also found that the formation of new bone, as reflected by an increase in serum PINP, was inhibited by the addition of a glucocorticoid to the treatment with bortezomib and extended the observations by in vitro studies demonstrating that pulse-treatment with bortezomib induced osteoblast differentiation which was inhibited by glucocorticoid treatment, while osteoblast proliferation was not affected.

Objective To investigate the effects of tocilizumab (TCZ) added to a stable dosage of methotrexate (MTX) on biochemical markers of bone and cartilage metabolism in patients in the multicenter double-blind, placebo-controlled OPTION (Tocilizumab Pivotal Trial in Methotrexate Inadequate Responders) study who have moderate-to-severe rheumatoid arthritis (RA) and an inadequate response to MTX. Methods Included in this study were 416 of the 623 patients with active RA enrolled in the OPTION study. Patients were randomized to receive TCZ (4 mg/kg or 8 mg/kg) or placebo intravenously every 4 weeks, with MTX continued at the stable prestudy doses (10–25 mg for 20 weeks, with a final followup at week 24). Serum biochemical markers of bone formation (osteocalcin, N-terminal propeptide of type I collagen [PINP]), bone resorption (C-terminal crosslinking telopeptide of type I collagen [CTX-I] and C-terminal crosslinking telopeptide of type I collagen generated by matrix metalloproteinases [ICTP]), cartilage metabolism (N-terminal propeptide of type IIA collagen [PIIANP]), collagen helical peptide [HELIX-II]), and matrix metalloproteinase 3 (MMP-3) were measured at baseline and at weeks 4, 16, and 24. Results TCZ induced marked dose-dependent reductions in PIIANP, HELIX-II, and MMP-3 levels at week 4 that were maintained until week 24, an effect associated with increased levels of bone formation markers that were significant as compared with placebo only for PINP and only at 4 weeks (P < 0.01 for both TCZ doses). TCZ induced significant decreases in the bone degradation markers CTX-I and ICTP, providing initial evidence of a beneficial effect on bone turnover. TCZ-treated patients who met the American College of Rheumatology 50% improvement criteria (achieved an ACR50 response) or achieved clinical remission (as determined by a Disease Activity Score in 28 joints

Changes of bone turnover with aging are responsible for bone loss and play a major role in osteoporosis. Although an increase of bone turnover has been documented at the time of menopause, the subsequent abnormalities of bone resorption and formation and their potential role in determining bone mass in the elderly have not been investigated. To address this issue, we have measured a battery of new sensitive and specific markers of bone turnover in a population-based study of 653 healthy women analyzed cross-sectionally, including 432 women postmenopausal from 1 to 40 years, and the data were correlated with bone mineral density (BMD) measured by dual-energy X-ray absorptiometry (DXA) at different skeletal sites. Bone formation was assessed by serum osteocalcin (OC), serum bone-specific alkaline phosphatase (B-ALP), serum C-propeptide of type I collagen (PICP), and bone resorption by the urinary excretion of two pyridinoline cross-linked peptides (NTX and CTX). Bone turnover increased in perimenopausal women with both irregular menses and elevated serum follicle stimulating hormone (FSH). Menopause induced a 37-52% and 79-97% increase in the bone formation and bone resorption marker levels, respectively (p < 0.0001 except for PICP). In postmenopausal women, bone formation markers did not decrease with age. When resorption markers were corrected by whole body bone mineral content (BMC), the fraction of bone resorbed per day was not correlated with age in postmenopausal women and remained elevated for up to 40 years after menopause. In premenopausal women, the bone turnover rate accounted for only 0-10% of the variation in whole body BMC, total hip, distal radius, and lumbar spine BMD. With increasing time after menopause, the importance of the bone turnover rate as a determinant of bone mass increased at all sites and accounted for up to 52% of the BMD variance in elderly women. Thus, in women 20 years or more postmenopause, bone turnover was higher in those in the lowest quartile than in those in the highest quartile of BMD. In elderly women, 20 years since menopause and over, but not in younger ones, serum PTH was negatively correlated with serum 25-hydroxyvitamin D (r = -0.22, p < 0.05) and explained only 5-8% of the bone turnover variance (p < 0.01-0.001). These data indicate that the overall rates of both bone formation and bone resorption remain high in elderly women. The rate of bone turnover appears to play an increasing role as a determinant of bone mass with increasing time since menopause with a high bone turnover rate being associated with a low bone mass. Thus assessing bone marker levels may be useful in the evaluation of osteoporosis risk. In elderly women, secondary hyperparathyroidism caused in part by reduced serum 25-hydroxyvitamin D appears to be a marginal determinant of an increased bone turnover rate.

We developed a sensitive and specific two-site radioimmunoassay (IRMA) for human osteocalcin using human osteocalcin as a standard and two monoclonal antibodies raised against human osteocalcin purified from human cortical bone, a solid-phase anti-25–37 region and a tracer anti-5–13 sequence of the molecule. A wide range of osteocalcin levels (up to 300 ng/ml) can be measured with a sensitivity of 0.4 ng/ml. The intra- and interassay coefficients of variation are less than 4 and 6%, respectively. The recovery of human osteocalcin from serum samples ranges from 96 to 103%. IRMA was linear for serial sample dilutions in a wide range of serum osteocalcin levels, even in patients with chronic renal failure on hemodialysis. Depletion of serum in intact osteocalcin demonstrated that IRMA detects, in addition to the intact peptide, a large N-terminal midregion fragment that represents about 50% of total osteocalcin levels in normals and patients with Paget's disease and up to 75% in patients with chronic renal failure. This large fragment, previously unrecognized because it cannot be distinguished from intact osteocalcin with gel filtration chromatography, is not generated in vitro by incubation of the serum up to 26 h. We measured osteocalcin in the serum of 309 healthy adults (180 men and 129 women, age range 20–95 years), 36 patients with Paget's disease, 12 patients with primary hyperparathyroidism, 70 patients with chronic renal failure on hemodialysis, and 10 patients on corticosteroid therapy, simultaneously with human IRMA and with a conventional radioimmunoassay (RIA) based on bovine reagents. A tight correlation (r = 0.889) was observed between the two assays in the normal population, but the values obtained with IRMA were about threefold higher (mean 23.3 ± 10.5 versus 7.5 ± 3.4 ng/ml) than those obtained with RIA. Reported as Z scores, that is, number of standard deviations from the predicted normal mean adjusted for sex and age, these two assays (IRMA and RIA) gave concordant results in patients with Paget's disease (4.05 ± 6.21 versus 2.41 ± 2.53), primary hyperparathyroidism (4.14 ± 7.17 versus 2.13 ± 2.28), chronic renal failure (25.32 ± 24.49 versus 6.93 ± 5.48), and glucocorticoid treatment (-1.48 ± 0.78 versus −1.11 ± 0.57). However, IRMA was more discriminant from controls for all these metabolic bone diseases because the absolute values of mean Z scores with IRMA were significantly higher than those obtained with the RIA (p < 0.05–0.0001). We conclude that this new human-specific IRMA of osteocalcin may be more sensitive than bovine RIA for the clinical investigation of metabolic bone diseases.

Increased bone turnover has been suggested as a potential risk factor for osteoporotic fractures. We investigated this hypothesis in a prospective cohort study performed on 7598 healthy women more than 75 years of age. One hundred and twenty-six women (mean years 82.5) who sustained a hip fracture during a mean 22-month follow-up were age-matched with three controls who did not fracture. Baseline samples were collected prior to fracture for the measurement of two markers of bone formation and three urinary markers of bone resorption: type I collagen cross-linked N- (NTX) or C-telopeptide (CTX) and free deoxypyridinoline (free D-Pyr). Elderly women had increased bone formation and resorption compared with healthy premenopausal women. Urinary excretion of CTX and free D-Pyr, but not other markers, was higher in patients with hip fracture than in age-matched controls (p = 0.02 and 0.005, respectively). CTX and free D-Pyr excretion above the upper limit of the premenopausal range was associated with an increased hip fracture risk with an odds ratio (95% confidence interval) of 2.2 (1.3-3.6) and 1.9 (1.1-3.2), respectively, while markers of formation were not. Increased bone resorption predicted hip fracture independently of bone mass, i.e., after adjustment for femoral neck bone mineral density (BMD) and independently of mobility status assessed by the gait speed. Women with both a femoral BMD value of 2.5 SD or more below the mean of young adults and either high CTX or high free D-Pyr levels were at greater risk of hip fracture, with an odds ratio of 4.8 and 4.1, respectively, than those with only low BMD or high bone resorption. Elderly women are characterized by increased bone turnover, and some markers of bone resorption predict the subsequent risk of hip fracture independently of hip BMD. Combining the measurement of BMD and bone resorption may be useful to improve the assessment of the risk of hip fracture in elderly women.

We have measured the free and peptide-bound type I collagen cross-link excretions in normal women and in patients with metabolic bone disease using the HPLC technique and immunoassays recognizing specifically the free or peptide-bound forms of pyridinoline (Pyr). After menopause, free deoxypyridinoline (free D-Pyr) excretion measured by HPLC without urine hydrolysis and expressed as a fraction of the total excretion was lower than in premenopausal women (45 ± 15% vs. 59 ± 12%, p > 0.005), whereas the fraction of free Pyr was not changed. In normal pre- and postmenopausal women (n = 43), the fraction of free D-Pyr was negatively correlated with bone turnover rate as assessed by the total urinary excretion of Pyr (r = –0.64, p > 0.001). In patients with a variety of metabolic bone diseases characterized by increased bone turnover (osteoporosis, Paget's disease, and hyperthyroidism), the fractions of free Pyr and free D-Pyr were significantly lower than in premenopausal controls (p > 0.001 for all diseases). After 3 days of intravenous (iv) treatment with the bisphosphonate pamidronate in patients with Paget's disease and osteoporosis, the urinary excretion of cross-linked peptides measured by high performance liquid chromatography (HPLC) or enzyme-linked immunoassay (ELISA) (NTX and CrossLaps) was markedly decreased (–52% and –85% for NTX, –71% and –93% for CrossLaps™ in Paget's disease and osteoporosis, respectively). The excretion of total cross-links was decreased to a lesser extent after treatment (–25% and –25% for total Pyr, –37% and –45% for total D-Pyr) and, surprisingly, free cross-links measured either by HPLC without urine hydrolysis or with an ELISA specific for free D-Pyr were unchanged after treatment. In contrast to bisphosphonate therapy, estrogen treatment of postmenopausal women decreased not only total and cross-linked peptides but also the free cross-link excretion that was reduced by about 30–40%. The different effects of bisphosphonate and estrogen therapy on the excretion of peptide-bound and free cross-link excretion were confirmed by gel filtration of the urine on Sephadex G 25 before and after treatment. In conclusion, we have shown that increased bone turnover in patients with metabolic bone disease could result in a larger increase of the urinary excretion of cross-linked peptides over the increase of free cross-links. Bisphosphonate therapy decreased markedly cross-linked peptides without significant change in free cross-link excretion contrasting with a decrease of both free and peptide-bound cross-links after estrogen therapy. These data suggest that these two antiresorptive therapies may affect differently the pattern of bone collagen degradation, an intriguing possibility that should be further investigated in vitro.

We used data from the Osteoporotic Fractures in Men (MrOS) study to test the hypothesis that men with higher levels of bone turnover would have accelerated bone loss and an elevated risk of fracture. MrOS enrolled 5995 subjects65 yr; hip BMD was measured at baseline and after a mean follow-up of 4.6 yr. Nonspine fractures were documented during a mean follow-up of 5.0 yr. Using fasting serum collected at baseline and stored at -190 degrees C, bone turnover measurements (type I collagen N-propeptide [PINP]; beta C-terminal cross-linked telopeptide of type I collagen [betaCTX]; and TRACP5b) were obtained on 384 men with nonspine fracture (including 72 hip fractures) and 947 men selected at random. Among randomly selected men, total hip bone loss was 0.5%/yr among those in the highest quartile of PINP (44.3 ng/ml) and 0.3%/yr among those in the lower three quartiles (p = 0.01). Fracture risk was elevated among men in the highest quartile of PINP (hip fracture relative hazard = 2.13; 95% CI: 1.23, 3.68; nonspine relative hazard = 1.57, 95% CI: 1.21, 2.05) or betaCTX (hip fracture relative hazard = 1.76, 95 CI: 1.04, 2.98; nonspine relative hazard = 1.29, 95% CI: 0.99, 1.69) but not TRACP5b. Further adjustment for baseline hip BMD eliminated all associations between bone turnover and fracture. We conclude that higher levels of bone turnover are associated with greater hip bone loss in older men, but increased turnover is not independently associated with the risk of hip or nonspine fracture.

Udgivelsesdato: Jul-31 Several studies have evaluated the impact of low dose statin (20-80 mg/day) on bone metabolism with inconclusive results despite promising data of preclinical studies. In this study, we investigated the effect of high dose simvastatin (HD-Sim) on biochemical markers of bone turnover and disease activity in six heavily pretreated patients with multiple myeloma (MM). These patients were treated with simvastatin (15 mg/kg/day) for 7 days followed by a rest period of 21 days in two 4-week cycles. Endpoints were changes in the level of biochemical markers of (i) osteoclast activity (tartrate resistant acid phosphatase, TRACP); (ii) bone resorption (collagen fragments CTX and NTX); (iii) bone formation (osteocalcin and aminoterminal propeptide of type I collagen PINP); (iv) cholesterol; (v) regulators of bone metabolism [osteoprotegerin (OPG) and Dickkopf-1 (DKK-1)] and (vi) disease activity (monoclonal proteins or free light chains in serum). TRACP activity in serum and levels of collagen fragments (NTX) in urine increased for all patients temporarily during the 7 days of treatment with HD-Sim indicating that osteoclasts may have been stimulated rather than inhibited. The other markers of bone metabolism showed no change. None of the patients showed any reduction in free monoclonal light chains or monoclonal proteins in serum during treatment with HD-Sim. In spite of the fact that bone turn over effects of HD-Sim may have been blunted by concomitant treatment of patients with other drugs we observed a transient increase in markers of osteoclast activity. This sign of a transient stimulation of osteoclast activity suggests that HD-Sim may be harmful rather than beneficial for MM patients. For this reason and because of gastro-intestinal side effects the study was stopped prematurely. Copyright (c) 2008 John Wiley & Sons, Ltd.

BACKGROUND. For patients with bone metastases, high N-telopeptide of type I collagen (NTX) levels correlate with increased risks of skeletal-related events and death. However, the relation between NTX decreases and clinical benefits is unclear. METHODS. Correlations between NTX normalization during treatment and clinical outcome were retrospectively analyzed in 3 large, phase 3 trials. Urinary NTX levels were measured at baseline and at Month 3 in patients with bone metastases from breast cancer (BC; n = 578), hormone-refractory prostate cancer (HRPC; n = 472), or nonsmall-cell lung cancer and other solid tumors (NSCLC/OST; n = 291) who received zoledronic acid or control (pamidronate for BC; placebo for HRPC and NSCLC/OST) for up to 24 months. NTX levels were characterized as normal (N

Fracture efficacy of PTH and alendronate (ALN) is only partly explained by changes in BMD, and bone collagen properties have been suggested to play a role. We analyzed the effects of PTH(1-84) and ALN on urinary alphaalpha/betabeta CTX ratio, a marker of type I collagen isomerization and maturation in postmenopausal women with osteoporosis. In the first year of the previously published PaTH study, postmenopausal women with osteoporosis were assigned to PTH(1-84) (100 microg/d; n = 119), ALN (10 mg/d; n = 60), or PTH and ALN together (n = 59). We analyzed patients on ALN alone (n = 60) and a similar number of patients assigned to PTH alone (n = 63). During the second year, women on PTH in the first year were reallocated to placebo (n = 31) or ALN (n = 32) and women with ALN continued on ALN. During the first year, there was no significant change in alphaalpha/betabeta CTX ratio with PTH or ALN. At 24 mo, there was a marked increase of the alphaalpha/betabeta CTX ratio in women who had received PTH during the first year, followed by a second year of placebo (median: +45.5, p < 0.001) or ALN (+55.2%, p < 0.001). Conversely, the alphaalpha/betabeta CTX ratio only slightly increased (+16%, p < 0.05) after 2 yr of continued ALN. In conclusion, treatment with PTH(1-84) for 1 yr followed by 1 yr of placebo or ALN may be associated with decreased type I collagen isomerization. The influence of these biochemical changes of type I collagen on bone fracture resistance remains to be studied.

In this longitudinal, prospective, and population-based study (n = 1044), seven BTMs were assessed before and after trauma in 113 elderly women (85 with fractures). Markers were not altered in the immediate postfracture period but were clearly elevated during fracture repair. Recent fracture should thus be taken into account when markers are used in clinical practice.Fracture may influence the levels of bone turnover markers (BTM) and have implications for their use in clinical practice. In this longitudinal, prospective, and population-based study, we assessed prefracture levels of BTMs and compared them with postfracture levels of the same individuals immediately after fracture and during fracture repair. This is the first study in which the effect of fracture on bone markers has been evaluated with prefracture samples available.Serum and urine were collected at the emergency unit from 85 women (77.9 +/- 1.8 yr) who sustained a fracture after low-energy trauma and 28 controls (77.8 +/- 2.0 yr) with similar trauma but no fracture. All were participants of the Malmö OPRA study (n = 1044), and pretrauma samples were collected 1.05 +/- 0.85 yr before. Bone turnover was assessed by seven different BTMs reflecting different stages of bone metabolism {C-terminal cross-linked telopeptides of type I collagen [S-CTX], S-TRACP5b, N-terminal propeptides of type I collagen [S-PINP], serum osteocalcin (S-OC[1-49] and S-TotalOC), urinary deoxypyridinoline [U-DPD], and urinary osteocalcin [U-OC]}.BTMs sampled within a few hours after fracture were not altered from preinjury levels. Both bone formation and bone resorption markers were, however, significantly increased 4 mo after fracture. The elevation was most pronounced after hip fracture. Bone turnover remained elevated up to 12 mo after fracture.We believe this study extends our knowledge on the skeletal postfracture metabolic processes. In addition, it may provide a basis for future means to monitor pharmacological intervention promoting fracture healing.

Objective Type II collagen, which consists of a large helical domain and telopeptides at each end, is the most abundant protein of cartilage matrix. The aim of this study was to develop a biochemical marker reflecting the degradation of the helical region of type II collagen and to evaluate its clinical performance in patients with osteoarthritis (OA) and rheumatoid arthritis (RA). Methods We developed a competitive polyclonal antibody–based enzyme-linked immunosorbent assay (ELISA) using the 622–632 peptide derived from the sequence of the α1 chain of human type II collagen (HELIX-II) as immunogen and standard. We measured urinary levels of HELIX-II peptide and C-terminal crosslinking telopeptide of type II collagen (CTX-II) in 90 patients with knee OA (73% women; mean ± SD age 63.0 ± 8.0 years, mean ± SD disease duration 6.1 ± 6.8 years), 89 patients with early RA (disease duration ≤3 years) (79% women; mean ± SD age 48.7 ± 11.6 years), 25 patients with Paget's disease of bone (HELIX-II only), and 162 healthy controls. In RA patients, we investigated the relationships between baseline urinary HELIX-II and CTX-II levels and the progression of joint destruction as measured by the changes in the total Sharp score (average from 2 independent readers) over 1 year. Results The intraassay and interassay variations of the HELIX-II ELISA were lower than 13% and 15%, respectively. The HELIX-II ELISA showed no significant cross-reactivity with human intact or denatured type II collagen, with the homologous peptides from human type I or type III collagens, or with HELIX-II peptides elongated (by 1 amino acid) or shortened (by 1 or 2 amino acids) at the C-terminal end, indicating that the HELIX-II ELISA recognized a neoepitope from the α1 chain of type II collagen. Median urinary HELIX-II levels were increased in patients with knee OA (by 56%; P < 0.0001) or early RA (by 123%; P < 0.0001) compared with those in age- and sex-matched healthy controls. Baseline urinary HELIX-II levels in the highest tertile were associated with an increased risk of radiographic progression in RA (increase in the total Sharp score ≥0.5 units/year), with an odds ratio (OR) of 5.9 (95% confidence interval [95% CI] 2.0–17.2) after adjustment for serum C-reactive protein (CRP) levels and baseline joint damage. Patients with increased levels of both urinary HELIX-II and CTX-II had the highest risk of progression (OR 17.5 [95% CI 3.1–99]). Conclusion The HELIX-II ELISA is specific for type II collagen degradation, has adequate technical performance, and can distinguish patients with knee OA or RA from healthy controls. Elevated HELIX-II levels are associated with increased risk of radiographic progression in RA independently of CRP levels, baseline joint damage, and urinary CTX-II levels. The HELIX-II ELISA should be useful for the clinical investigation of patients with arthritis and for identifying RA patients at higher risk of progression.

We used data from the Fracture Intervention Trial to assess the relationship change in bone turnover after 1 year of alendronate or placebo treatment and subsequent hip, non-spine, and spine fracture risk among 6186 postmenopausal women. In the alendronate group (n = 3105), greater reductions in one or more biochemical marker were associated with a lower risk of fracture. Introduction: There are few data on the relationship between short-term change in biochemical markers of bone turnover and non-spine fracture risk among bisphosphonate-treated women, and the clinical use of such measurements is unknown. Materials and Methods: We measured biochemical markers of bone turnover (bone-specific alkaline phosphatase [bone ALP], intact N-terminal propeptide of type I collagen, and C-terminal crosslinked telopeptide of type 1 collagen) and BMD of the spine and hip at baseline and after 1 year of alendronate or placebo. During a mean follow-up of 3.6 years, 72 hip, 786 non-spine, and 336 vertebral fractures were documented. Results and Conclusions: Each 1 SD reduction in 1-year change in bone ALP was associated with fewer spine (odds ratio = 0.74; CI: 0.63, 0.87), non-spine (relative hazard [RH] = 0.89; CI: 0.78, 1.00; p < 0.050), and hip fractures (RH = 0.61; CI: 0.46, 0.78). Alendronate-treated women with at least a 30% reduction in bone ALP had a lower risk of non-spine (RH = 0.72; CI: 0.55, 0.92) and hip fractures (RH = 0.26; CI: 0.08, 0.83) relative to those with reductions

Objective. To investigate in a randomized clinical trial setting with an aggressive combination-therapy arm and a mild-monotherapy arm, whether therapy-induced changes in urinary C-terminal crosslinking telopeptide of type I collagen (CTX-I) and type II collagen (CTX-II) predict 5-year radiographic progression in patients with rheumatoid arthritis (RA). Methods. Patients had participated in the COBRA (Combinatietherapie Bij Reumatoide Artritis) trial comparing aggressive step-down combination therapy (the COBRA regimen, including temporary high-dose prednisolone, temporary low-dose methotrexate, and sulfasalazine [SSZ]) and mild monotherapy (SSZ). Urinary CTX-I and CTX-II levels were measured at baseline and 3, 6, 9, and 12 months after initiation of treatment. Radiographs were scored according to the modified Sharp/van der Heijde method (mean of 2 independent readers who were aware of the sequence). Individual long-term radiographic progression was estimated, using baseline radiographs and all radiographs obtained during the followup period, by simple linear regression analysis (curve fitting). Results. Both COBRA therapy and SSZ monotherapy produced a significant decrease in urinary CTX-I and CTX-II levels at 3 months, and this decrease was amplified at 6 months. COBRA therapy suppressed CTX-II (change from baseline levels -36% and -43% at 3 and 6 months, respectively), but not CTX-I, significantly better than did SSZ (-17% and -21% at 3 and 6 months, respectively) at 3 and 6 months. The magnitude of the decrease in urinary CTX-II levels at 3 months significantly predicted long-term (5-year) radiographic progression (β = 0.48 [95% confidence interval (95% CI) 0.13, 0.83]). This effect was independent of the change in disease activity and inflammation indices at 3 months. Patients whose CTX-II levels were normalized (

Bone marker variability has raised concern for its use in individual patients. Serum osteocalcin (formation) and CTX (resorption) were measured every year for 4 years in 268 postmenopausal women. Seventy percent to 80% of women classified as having high bone turnover at baseline were similarly classified by the same methods 4 years later. Introduction: High bone marker levels are a risk factor for osteoporosis in postmenopausal women, but variability of measurements has raised doubts about their clinical use in an individual patient. Methods: We studied 268 untreated postmenopausal women (50–81 years of age) belonging to a population-based prospective cohort. We collected fasting morning blood samples every year for 4 years to measure serum intact osteocalcin (OC) and serum C-terminal cross-linked telopeptide of type I collagen (CTX) as bone formation and resorption markers, respectively. Results: Serum OC and CTX remained stable during follow-up (+1.2%/year, p = 0.003 and −0.13%/year, p = 0.70 for OC and S-CTX, respectively). At baseline, women were classified as having low (tertile 1), intermediate (tertile 2), or high (tertile 3) bone turnover. Agreement of classification between baseline and 4-year measurements was moderate (κ [95% CI]: 0.51 [0.43–0.59] and 0.52 [0.44–0.60] for OC and S-CTX, respectively). Less than 10% of women in tertile 1 or 3 of either marker at baseline were found in the opposite tertile 4 years later. When the two markers were combined, only 2% of women at high turnover at baseline—defined as OC and/or S-CTX in tertile 3—were classified at low turnover 4 years later. Among women classified at high bone turnover at baseline (tertile 3), 70–80 % were also found at high turnover 4 years later. Among women in tertile 2, only 51% and 43% for OC and CTX, respectively, remained in the same tertile at the second measurement. Conclusions: Serum levels of bone formation and resorption markers are stable over 4 years in postmenopausal women, on average. The majority of women classified as having high bone turnover were similarly classified by the same methods 4 years later. However, 20–30 % of these women at risk for fracture would be incorrectly classified, suggesting that further investigation would be required to reduce the number of patients who would be treated unnecessarily if the decision was made on bone marker measurement. For women with intermediate levels, classification may be improved by a second measurement or by combining two markers.

Changes in the level of biochemical markers of bone resorption with risedronate treatment for osteoporosis were examined as a surrogate for the decrease in fracture risk. Greater decreases in bone resorption markers were associated with greater decreases in vertebral (and nonvertebral) fractures. Antifracture efficacy of antiresorptive therapies is only partially explained by increases in bone mineral density. Early decreases in bone resorption may also play a role. We tested this hypothesis by measuring two bone resorption markers, the C-telopeptide of type I collagen (CTX) and the N-telopeptide of type I collagen (NTX), in osteoporotic patients in risedronate vertebral fracture trials. We studied 693 women with at least one vertebral deformity (mean age, 69 ± 7 years) who received calcium (and vitamin D if required) and placebo or risedronate 5 mg daily for 3 years. The reductions in urinary CTX (median, 60%) and NTX (51%) at 3-6 months with risedronate therapy were significantly associated (p < 0.05) with the reduction in vertebral fracture risk (75% over 1 year and 50% over 3 years). The changes in both CTX and NTX accounted for approximately one-half (CTX, 55%; NTX, 49%) of risedronate's effect in reducing the risk of vertebral fractures in the first year and approximately two-thirds (CTX, 67%; NTX, 66%) over 3 years compared with placebo. The changes in CTX and NTX accounted for 77% and 54%, respectively, of risedronate's effect in reducing the risk of nonvertebral fractures over 3 years compared with placebo. The relationships between vertebral fracture risk and changes from baseline in CTX and NTX were not linear (p < 0.05). There was little further improvement in fracture benefit below a decrease of 55-60% for CTX and 35-40% for NTX. The decrease in bone resorption in patients taking risedronate accounts for a large proportion of the reduction in fracture risk. There may be a level of bone resorption reduction below which there is no further fracture benefit.

Bone resorption may generate collagen fragments such as ICTP and CTX, which can be quantified in serum and/or urine by using specific immunoassays, and which are used as clinical markers. However, the relative abundance of ICTP and CTX varies according to the type of bone pathology, suggesting that these two fragments are generated through distinct collagenolytic pathways. In this study, we analyzed the release of ICTP and CTX from bone collagen by the proteinases reported to play a role in the solubilization of bone matrix. Cathepsin K released large amounts of CTX, but did not allow a detectable release of ICTP. Conversely, the matrix metalloproteinases (MMPs) MMP-2, -9, -13, or -14 released ICTP, but did not allow a detectable release of CTX. Next we analyzed the release of ICTP and CTX from bone explants cultured in the presence of well-established inhibitors of these proteinases and of matrix solubilization. An inhibitor of cysteine proteinases including cathepsin K, inhibited the release of CTX, but not the release of ICTP. MMP inhibitors inhibited the release of ICTP, but also that of CTX, in agreement with the putative role of MMPs in the initiation of bone resorption in addition to matrix solubilization. Similarly the treatment of mice bearing bone metastasis with an MMP inhibitor led to a significant reduction of serum ICTP and CTX, and osteolytic lesions. We conclude that the generation of ICTP and CTX depends on different collagenolytic pathways. This finding may explain why these two markers may discriminate between different bone pathologies.

In addition to its presence in blood samples from morphine-treated patients and heroin addicts, morphine is endogenously synthesized from dopamine in mammals [1, 2]. However, the physiological relevance of endogenous morphine (eM) remains an open question. Several studies report an increase in eM in blood and urine samples in pathological conditions such as infection, inflammation, bulimia, anorexia and Parkinson's disease [2, 3]. For example, eM concentrations reached on average 2.5 ng ml−1 in the serum of patients suffering from systemic infections (sepsis), whereas healthy donors and control patients displayed no or very low eM concentrations [4]. Therefore, quantification of endogenous and exogenous morphine is of great interest for both clinical and forensic analyses. As both endogenous and exogenous morphine are present at low concentrations, the quantification method implemented must be very sensitive. Besides, no consensus yet exists regarding blood sample preparation for morphine measurement using ELISA. As lithium-heparinate (plasma) and serum samples are both routinely used in clinical practice, we compared the morphine concentrations using these two sampling methods. This study was approved by our institutional review board for human experimentation (CPP no. 08/62-protocol no. IVD1 EUDRACT no. 2008-A01345-50). After informed consent, samples were collected from patients requiring clinical laboratory analyses. Blood from 80 patients was collected in two types of sampling tubes, as follows: (i) for serum analysis, blood was collected in dry tubes (BD, Plymouth, UK; SSTII 377615), and serum was obtained after centrifugation (4°C, 1500g, 10 min); and (ii) for plasma analysis, blood was collected in lithium-heparinate tubes (BD; LH-PSTII 367378, 17 i.u. ml−1) and plasma was recovered after centrifugation (4°C, 1500g, 10 min). Collected serum and plasma were immediately stored at −80°C (for 3 weeks to 3 months). Samples were thawed only once, 15 min prior to ELISA testing. In these conditions, no effect of freezing on morphine concentrations has been noticed. Both plasma and serum morphine concentrations were quantified using a morphine-specific ELISA kit (ref. 213-0480; Immunalysis, Pomona, CA, USA) [4]. No cross-reactivity has been observed for morphine metabolites (morphine-6-glucuronide, morphine-3-glucuronide and codeine) and 75 other compounds (related to eM biosynthesis pathway or not; see also kit's datasheet). The plasma or serum samples were tested in duplicates (40 µl) at 21°C. The coefficient of variation values were between 0% and 8%. All samples with a higher coefficient of variation value were retested in order to obtain a coefficient of variation below or equal to 8%. Morphine standards (0–25 ng ml−1) were diluted in phosphate-buffered saline containing 1% bovine serum albumin (PBS-BSA; pH 7.2; w/v) for serum samples and in PBS-BSA 1% containing lithium-heparinate (17 i.u. ml−1) for plasma samples. The detection limit for morphine was 0.1 ng ml−1. In a set of control experiments, known concentrations of morphine (0.28 and 11 ng ml−1) were diluted in PBS, PBS–lithium-heparinate (17 i.u. ml−1, pH 7.2), PBS–BSA (1%; w/v) or PBS–lithium-heparinate–BSA buffer. Using the ELISA kit, no statistical differences in morphine concentrations were observed (n = 8, Mann–Whitney U-test, P > 0.3) when 0.28 ng ml−1 of morphine was diluted in PBS (0.26 ± 0.03 ng ml−1), PBS–lithium-heparinate [0.26 ± 0.02 ng ml−1; 95% confidence interval (CI) of difference with PBS −0.03 to 0.03], PBS–BSA (0.27 ± 0.02 ng ml−1; 95% CI of difference with PBS −0.04 to 0.02) or PBS–lithium-heparinate–BSA (0.28 ± 0.04 ng ml−1; 95% CI of difference with PBS −0.04 to 0.0). Likewise, no statistical difference (n = 6, Mann–Whitney U-test, P > 0.2) was observed for 11 ng ml−1 of morphine diluted in PBS (11.06 ± 0.58 ng ml−1), PBS–lithium-heparinate (11.42 ± 0.36 ng ml−1; 95% CI of difference with PBS −0.63 to 0.10), PBS–BSA (11.01 ± 0.5 ng ml−1; 95% CI of difference with PBS −0.25 to 0.55) or PBS–lithium-heparinate–BSA (11.36 ± 0.55 ng ml−1; 95% CI of difference with PBS −0.94 to 0.52). This experiment clearly demonstrates that lithium-heparinate and BSA do not introduce a bias in the detection of morphine using this particular ELISA kit. However, when serum and lithium-heparinate tubes were spiked with a known concentration of morphine prior to fresh blood collection (healthy donors, n = 6; 10 ng ml−1), 86 ± 7% (95% CI 79.8–92.2%) of the initial morphine concentration was recovered in plasma samples, but only 42 ± 3% (95% CI 37.9–46.4%) in serum samples (Mann–Whitney U-test, P < 0.01). Finally, 80 patients distributed in the following three groups were tested: (i) healthy donors (n = 16); (ii) morphine-treated patients (morphine skin patch 12–48 h before blood collection, 20 mg day−1, n = 8); and (iii) critically ill patients (n = 56, naive for morphine), including patients with no infection and no inflammation (n = 18), systemic inflammatory response syndromes (n = 8), severe symstemic inflammatory response syndrome (n = 16), sepsis (n = 4), severe sepsis (n = 2) and septic shock (n = 8). In healthy donors, eM was undetectable in both serum and plasma samples. In morphine-treated patients (Figure 1), the mean morphine concentration in serum samples ranged from 3.2 to 31 ng ml−1 (median 8.1 ng ml−1; 95% CI 5.3–20.1 ng ml−1), whereas plasma values ranged from 25.7 to 114.8 ng ml−1 (median 54.8 ng ml−1; 95% CI 41.9–85.0 ng ml−1). On average, morphine concentrations in serum were 78 ± 15% (95% CI 68–88%) lower than in plasma (Wilcoxon signed-rank test, P < 0.05). In critically ill patients, eM concentrations ranged between 0 (not detectable) and 23.6 ng ml−1 in serum samples (median 1.1 ng ml−1; 95% CI 1.9–4.8 ng ml−1) and between 0.15 and 76.3 ng ml−1 (median 2.8 ng ml−1; 95% CI 6.7–15.5 ng ml−1) in plasma samples. Again, serum concentrations of eM were lower (from +13 to −98%; mean 65 ± 32%; 95% CI 57–73%) compared with plasma (Wilcoxon signed-rank test, P < 0.01). The presence of morphine was confirmed by liquid chromatography coupled to tandem mass spectrometry analysis performed on plasma and serum from eight patients not treated with morphine but positive for eM, whereas morphine was not detected in patients negative for eM in ELISA (n = 4) [4]. As expected, no morphine was detected in samples from four healthy donors. Figure 1 Concentrations of morphine in serum expressed as a function of the corresponding morphine concentrations in plasma in critically ill patients (n = 56, triangles) and in morphine-treated patients (n = 8, crosses). Inset shows a higher magnification of ... Our results revealed that morphine concentrations were systematically higher in plasma compared with serum for 61 of 64 patients and thus indicate that lithium-heparinate samples give a more accurate quantification of morphine blood concentrations. The cause of such a difference in morphine quantification between serum and plasma using this specific ELISA kit is currently under investigation by our group and might involve morphine binding to blood proteins [5] other than serum albumin. In line with this possibility, morphine detection in the blood has been shown to be highly dependent on urea concentrations (chaotropic agent), suggesting a link between morphine-binding proteins and morphine bioavailability [5]. Likewise, lithium is also a chaotropic/denaturing agent; therefore, it might explain why this ELISA quantification method for morphine is more efficient in the presence of lithium. The present results clearly demonstrate that to obtain accurate ELISA determinations of exogenous and endogenous morphine concentrations in the blood, the analysis should be carried out on lithium-heparinate plasma samples rather than serum samples.

Objective The known risk factors for radiologic progression in rheumatoid arthritis (RA) are not optimally discriminative in patients with early disease who do not have evidence of radiologic damage. We sought to determine whether urinary C-terminal crosslinking telopeptide of type I (CTX-I) and type II (CTX-II) collagen (markers of bone and cartilage destruction, respectively) are associated with long-term radiologic progression in patients with early RA. Methods This was a prospective study of 110 patients with early RA who were participating in the COBRA (Combinatietherapie Bij Reumatoide Artritis) clinical trial and followup study, a randomized controlled trial comparing the efficacy of oral pulse prednisolone, methotrexate, plus sulfasalazine with sulfasalazine alone. We investigated the relationship between baseline levels of urinary CTX-I and CTX-II and the mean annual progression of joint destruction over a median of 4 years, as measured by changes in the modified Sharp score (average of 2 independent readers). Results In multivariate logistic regression analysis, baseline urinary CTX-I and CTX-II levels in the highest tertile were the strongest predictors of radiologic progression (Sharp score increase >2 units/year; odds ratio 7.9 and 11.2, respectively), independently of treatment group, erythrocyte sedimentation rate (ESR), Disease Activity Score in 28 joints, rheumatoid factor (RF), and baseline joint damage (Sharp score). The likelihood ratios for a positive test were 3.8 and 8.0 for CTX-I and CTX-II, respectively, which compared favorably with the likelihood ratios for the ESR (3.0), baseline joint damage (1.6), and RF (1.8). When patients were grouped according to the presence (Sharp score ≥4, n = 49) and absence (Sharp score

Objective The hallmark of osteoarthritis (OA) is the loss of articular cartilage. This loss arises from an imbalance between cartilage synthesis and cartilage degradation over a variable period of time. The aims of this study were to investigate the rates of these processes in patients with knee OA using two new molecular markers and to investigate whether the combined use of these markers could predict the progression of joint damage evaluated by both radiography and arthroscopy of the joints during a period of 1 year. Methods Seventy-five patients with medial knee OA (51 women, 24 men; mean ± SD age 63 ± 8 years, mean ± SD disease duration 4.8 ± 5.2 years) were studied prospectively. At baseline, we measured serum levels of N-propeptide of type IIA procollagen (PIIANP) and urinary excretion of C-terminal crosslinking telopeptide of type II collagen (CTX-II) as markers of type II collagen synthesis and degradation, respectively. Joint space width (JSW) on radiography and medial chondropathy at arthroscopy (assessed using a 100-mm visual analog scale [VAS]) were measured in all patients at baseline and in 52 patients at 1 year. Progression of joint destruction was defined as a decrease of ≥0.5 mm in JSW on radiography and as increased chondropathy (an increase in the VAS score of >8.0 units) between the baseline and 1-year evaluations. Results At baseline, compared with 58 healthy age- and sex-matched controls, patients with knee OA had decreased serum levels of PIIANP (20 ng/ml versus 29 ng/ml; P < 0.001) and increased urinary excretion of CTX-II (618 ng/mmole creatinine [Cr] versus 367 ng/mmole Cr; P < 0.001). The highest discrimination between OA patients and controls was obtained by combining PIIANP and CTX-II in an uncoupling index (Z score CTX-II − Z score PIIANP), which yielded a mean Z score of 2.9 (P < 0.0001). Increased baseline values in the uncoupling index were associated with greater progression of joint damage evaluated either by changes in JSW (r = −0.46, P = 0.0016) or by VAS score (r = 0.36, P = 0.014). Patients with both low levels of PIIANP (less than or equal to the mean − 1 SD in controls) and high levels of CTX-II (greater than or equal to the mean + 1 SD in controls) had an 8-fold more rapid progression of joint damage than other patients (P = 0.012 and P < 0.0001 as assessed by radiography and arthroscopy, respectively) and had relative risks of progression of 2.9 (95% confidence interval [95% CI] 0.80–11.1) and 9.3 (95% CI 2.2–39) by radiography and arthroscopy, respectively. Conclusion Patients with knee OA are characterized by an uncoupling of type II collagen synthesis and degradation which can be detected by assays for serum PIIANP and urinary CTX-II. The combination of these two new markers could be useful for identifying knee OA patients at high risk for rapid progression of joint damage.

Objective To evaluate whether measurements of urinary glucosyl-galactosyl-pyridinoline (Glc-Gal-PYD) and urinary C-terminal crosslinking telopeptide of type II collagen (CTX-II), 2 new markers of destruction of the synovium and cartilage collagen breakdown, respectively, are associated with the progression of joint damage in patients with early rheumatoid arthritis (RA), and to compare this association with that with serum matrix metalloproteinase 3 (MMP-3), a proteinase expressed by synovial tissue and chondrocytes, and that with serum C-reactive protein (CRP), an index of systemic inflammation. Methods The prospective study cohort comprised 116 patients with early RA who were part of a large, double-blind, randomized study comparing the efficacy of etanercept and methotrexate. The relationship between baseline levels of urinary Glc-Gal-PYD, urinary CTX-II, and serum MMP-3 and the progression of joint destruction, as measured by changes in the modified Sharp score (average findings of 2 independent readers) over 1 year, was investigated. Results Levels of urinary Glc-Gal-PYD (+70%), urinary CTX-II (+104%), and serum MMP-3 (+219%) were elevated compared with the levels in 76 healthy controls. The baseline levels of Glc-Gal-PYD (r = 0.30), CTX-II (r = 0.25), and MMP-3 (r = 0.29) correlated with the changes over 1 year in the total Sharp score (joint space narrowing and bone erosion). Patients with baseline levels of Glc-Gal-PYD, CTX-II, and MMP-3 that were higher than the mean + 2 SD in healthy controls had a significantly greater progression of joint damage, with an increase in the total Sharp score over 1 year that was from 3- to 8-fold higher than that in patients with low baseline levels of these markers. Moreover, patients with these higher levels of Glc-Gal-PYD, CTX-II, and MMP-3 had a higher risk of progression of the disease (increase in total Sharp score ≥0.5 units) than did the other patients (relative risks and 95% confidence intervals [95% CI] 3.3 [95% CI 1.5–7.4], 2.5 [95% CI 1.1–5.7], and 2.5 [95% CI 1.1–5.6], respectively). The baseline serum level of CRP was not significantly associated with the progression of joint damage. Adjustment of the levels of Glc-Gal-PYD, CTX-II, and MMP-3 according to radiologic damage at baseline did not alter their association with progression. After adjustment for serum CRP, the relative risk slightly decreased, but remained significant, for Glc-Gal-PYD (2.6 [95% CI 1.1–6.3]). Patients with both increased levels of the molecular markers and radiologic damage at baseline had a higher risk of progression of joint damage than did those with either high molecular marker levels or radiologic damage. Conclusion High baseline levels of Glc-Gal-PYD, CTX-II, and MMP-3 are associated with increased risk of progression of joint destruction over 1 year in early RA. The association between baseline levels of urinary Glc-Gal-PYD and progression of joint erosion was independent of the severity of radiologic damage and inflammation at baseline. Combining the measurements of these molecular markers with radiologic assessment of joint damage may be useful for identifying patients with RA who are at high risk of rapid progression and for whom early aggressive treatment would be beneficial.

Dystonia has a wide clinical spectrum from early-onset generalized to late-onset sporadic, task-specific forms. The genetic origin of the former has been clearly established. A critical role of repetitive skilled motor tasks has been put forward for the latter, while underlying vulnerability traits are still being searched for. Using magnetoencephalography, we looked for structural abnormalities reflecting a preexisting dysfunction. We studied finger representations of both hands in the primary sensory cortex, as compared in 23 patients with unilateral task-specific dystonia and 20 control subjects. A dramatic disorganization of the nondystonic hand representation was found in all patients, and its amount paralleled the severity of the dystonic limb motor impairment. Abnormalities were also observed in the cortex coding the dystonic limb representation, but they were important only in the most severely affected patients. The abnormal cortical finger representations from the nondystonic limb appear to be endophenotypic traits of dystonia. That finger representations from the dystonic limb were almost normal for the less severely affected patients may be due to intrinsic beneficial remapping in reaction against the primary disorder.

There are relatively few data concerning age-related changes of bone turnover in men. The aim of the study was to evaluate age-related changes of the levels of serum and urinary biochemical markers of bone metabolism in a large cohort of 934 men aged 19-85 years and to investigate their association with bone mineral density (BMD). Bone formation was evaluated using serum levels of osteocalcin (OC), bone alkaline phosphatase (BAP), and N-terminal extension propeptide of type I collagen (PINP). Bone resorption was evaluated by measurement of urinary excretion of beta-isomerized C-terminal telopeptide of collagen type I beta-CTX) of free deoxypyridinoline (fDpyr) and total Dpyr (tDPyr) and of the serum level of beta-CTX. Levels of biochemical bone markers were very high in young men and decreased rapidly until the age of 40 years and then more slowly until 60 years of age. After the age of 60 years, markers of bone formation remained stable while resorption markers showed a moderate and variable increase with aging. Serum and urinary beta-CTX levels were elevated only in about 5% of elderly men. The age-related increase of urinary excretion of tDpyr and of its free and peptide-bound fractions was related to the presence of elevated levels in a subgroup of about 15% of elderly men. Before 60 years of age, levels of biochemical bone markers were not correlated with BMD, whereas after 60 years of age, they were correlated negatively with BMD. After adjustment for age and body weight, BMD in men with the highest levels of biochemical bone markers (i.e., in the upper quartile) was 1.8-12.5% (i.e., 0.25-0.89 SD) lower than in men with levels of biochemical bone markers in the lowest quartile. In conclusion, bone turnover in men is high in young adults and decreases to reach a nadir at 55-60 years of age. After the age of 60 years, bone resorption markers--but not bone formation markers--increase in some men and are associated with lower BMD, suggesting that this imbalance is responsible for increasing bone loss in elderly men.

The aims of this study performed in ewes were: (1) to confirm in this animal model the effects on bone of ovariectomy (OVX) alone or associated with Lentaron (L), a potent peripheral aromatase inhibitor, used to amplify the effects of OVX and (2) to evaluate the effects of a new selective estrogen receptor modulator (SERM; MDL 103,323) on bone remodeling. Thirty-nine old ewes were divided into five groups: sham (n = 7); OVX (n = 8); OVX + L (n = 8); OVX + L + MDL; 0.1 mg/kg per day (n = 8); and OVX + L + MDL 1 mg/kg per day (n = 8). The animals were treated for 6 months. Biochemical markers of bone turnover (urinary excretion of type 1 collagen C-telopeptide [CTX], serum osteocalcin [OC], and bone alkaline phosphatase [BAP]) were measured each month. Bone biopsy specimens were taken at the beginning and after death at the end of the experiment. Bone mineral density (BMD) was measured by dual-energy X-ray absorptiometry (DXA) on the lumbar spine and femur. OVX induced a significant increase in biochemical markers. This effect was the highest after 3 months for CTX (+156% vs. sham) and after 4 months for OC and BAP (+74% and +53% vs. sham, respectively). L tended to amplify the effect of OVX on OC and BAP. OVX induced significant increases in the porosity, eroded, and osteoid surfaces in cortical bone but no effect was observed in cancellous bone. MDL treatment reduced the bone turnover as assessed by bone markers, which returned to sham levels as well as histomorphometry both in cortical and in cancellous bone. Cancellous osteoid thickness decreased by 27% (p < 0.05), mineralizing perimeter by 81% (p < 0.05), and activation frequency by 84% (p < 0.02) versus OVX + L. Femoral and spinal BMD were increased by MDL and tended to return to the sham values. The effects of OVX on bone turnover were different on cortical and cancellous bone. These effects on cortical bone were reflected by changes in biochemical markers. MDL markedly reduces bone turnover and increases BMD suggesting that this new agent may prevent postmenopausal bone loss.

Objective To examine functional interleukin-6 (IL-6) −174 GC allelic variants in relation to bone turnover and bone mineral density (BMD) in postmenopausal women. Methods Four hundred thirty-four healthy women living in the community (mean ± SD age 71.7 ± 5.7 years) were genotyped for the IL-6 −174 GC polymorphism. Serum levels of C-telopeptide of type I collagen (CTx), a marker of bone resorption, and osteocalcin (OC), a marker of bone formation, were determined. BMD at the hip and forearm was measured by dual-energy x-ray absorptiometry. Results CTx levels differed significantly (P = 0.006) among IL-6 genotypes (mean ± SEM 0.275 ± 0.02 ng/ml, 0.325 ± 0.01 ng/ml, and 0.356 ± 0.02 ng/ml in women with the CC genotype [n = 68], the GC genotype [n = 204], and the GG genotype [n = 162], respectively). Compared with the GG group, age-adjusted odds ratios for high bone resorption were 0.65 (95% confidence interval [95% CI] 0.41–1.0, P = 0.06) and 0.37 (95% CI 0.18–0.73, P = 0.0047) in GC and CC subjects, respectively. In contrast, OC levels did not differ by genotype. BMD at the hip and forearm was 1.5–5% higher in CC subjects compared with GG subjects (P not significant). When the cohort was divided according to the median age (70.5 years), BMD was significantly decreased in older compared with younger postmenopausal women with the GG and GC genotypes (−9.6% on average; P < 0.01), but not in those with the CC genotype (−5.1% on average; P not significant). Conclusion Compared with the GC and GG IL-6 −174 GC genotypes, the CC genotype is associated with lower bone resorption and lesser decrease in bone mass in older postmenopausal women. These results suggest that IL-6 −174 GC alleles may be significant determinants of the risk for osteoporosis in elderly subjects.