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The osteopetroses and related sclerosing bone dysplasias can have a broad range of manifestations. Especially in the milder forms, sandwich vertebrae are an easily recognizable and reliable radiological hallmark. We report on four patients from three families presenting with sandwich vertebrae and platyspondyly. The long bone phenotypes were discordant with one patient showing modeling defects and patchy osteosclerosis, while the second displayed only metaphyseal sclerotic bands, and the third and fourth had extreme metaphyseal flaring with uniform osteosclerosis. Two of the four patients had experienced pathological fractures, two had developmental delay, but none showed cranial nerve damage, hepatosplenomegaly, or bone marrow failure. According to these clinical features the diagnoses ranged between intermediate autosomal recessive osteopetrosis and dysosteosclerosis. After exclusion of mutations in CLCN7 we performed gene panel and exome sequencing. Two novel mutations in SLC29A3 were found in the first two patients. In the third family a TCIRG1 C-terminal frameshift mutation in combination with a mutation at position +4 in intron 2 were detected. Our study adds two cases to the small group of individuals with SLC29A3 mutations diagnosed with dysosteosclerosis, and expands the phenotypic variability. The finding that intermediate autosomal recessive osteopetrosis due to TCIRG1 splice site mutations can also present with platyspondyly further increases the molecular heterogeneity of dysosteosclerosis-like sclerosing bone dysplasias.

Context Men with congenital hypogonadotropic hypogonadism (CHH) and Kallmann syndrome (KS) have both low circulating testosterone and estradiol levels. Whether bone structure is affected remains unknown. Objective To characterize bone geometry, volumetric density and microarchitecture in CHH/KS. Methods This cross-sectional study, conducted at a single French tertiary academic medical center, included 51 genotyped CHH/KS patients and 40 healthy volunteers. Among CHH/KS men, 98% had received testosterone and/or combined gonadotropins. High-resolution peripheral quantitative computed tomography (HR-pQCT), dual-energy x-ray absorptiometry (DXA), and measurement of serum bone markers were used to determine volumetric bone mineral density (vBMD) and cortical and trabecular microarchitecture. Results CHH and controls did not differ for age, body mass index, and levels of vitamin D and PTH. Despite long-term hormonal treatment (10.8 ± 6.8 years), DXA showed lower areal bone mineral density (aBMD) in CHH/KS at lumbar spine, total hip, femoral neck, and distal radius. Consistent with persistently higher serum bone markers, HR-pQCT revealed lower cortical and trabecular vBMD as well as cortical thickness at the tibia and the radius. CHH/KS men had altered trabecular microarchitecture with a predominant decrease of trabecular thickness. Moreover, CHH/KS men exhibited lower cortical bone area, whereas total and trabecular areas were higher only at the tibia. Earlier treatment onset (before age 19 years) conferred a significant advantage for trabecular bone volume/tissue volume and trabecular vBMD at the tibia. Conclusion Both vBMD and bone microarchitecture remain impaired in CHH/KS men despite long-term hormonal treatment. Treatment initiation during adolescence is associated with enhanced trabecular outcomes, highlighting the importance of early diagnosis.

An association between atherosclerosis and osteoporosis has been reported in several studies. This association could result from local intraosseous atherosclerosis and ischemia, which is shown by limb osteoporosis in patients with peripheral artery disease (PAD), but also could result from bidirectional communication between the skeleton and blood vessels. Systemic bone disorders and PAD are frequent in ESRD. Here, we investigated the possible interaction of these disorders. For 65 prevalent nondiabetic patients on hemodialysis, we measured ankle-brachial pressure index (ABix) and evaluated mineral and bone disorders with bone histomorphometry. In prevalent patients on hemodialysis, PAD (ABix0.9 or1.4/incompressible) was associated with low bone turnover and pronounced osteoblast resistance to parathyroid hormone (PTH), which is indicated by decreased double-labeled surface and osteoblast surface (P0.001). Higher osteoblast resistance to PTH in patients with PAD was characterized by weaker correlation coefficients (slopes) between serum PTH and double-labeled surface (P=0.02) or osteoblast surface (P=0.03). The correlations between osteoclast number or eroded surface and serum mineral parameters, including PTH, did not differ for subjects with normal ABix and PAD. Common vascular risk factors (dyslipidemia, smoking, and sex) were similar for normal, low, and incompressible ABix. Patients with PAD were older and had high C-reactive protein levels and longer hemodialysis vintage. These results indicate that, in prevalent nondiabetic patients with ESRD, PAD associates with low bone turnover and pronounced osteoblast resistance to PTH.

Objectives To determine biochemical, radiological and micro-architectural bone factors related to fragility fractures in idiopathic male osteoporosis (IMO) patients. IMO is a rare disorder characterized by low areal bone mineral density (aBMD) (Z-score Methods We conducted a case–control study in 31 patients with fragility fracture (IMO F +) that had occurred after the age of 40 years and 37 without fracture (IMO F–). We first compared IMO group to 40 age-matched disease-free men. We measured aBMD and bone micro-architectural indices at distal radius and tibia sites with a HR-pQCT scan (XtremeCT) using standard and extended cortical analysis. Urine and blood samples were collected in order to determine the levels of bone-turnover markers and the potential determinant of bone fragility. Models of analysis of covariance, including age, height and weight as adjustment factors, were used to compare the groups. Results Compared to their controls, IMO patients showed marked disturbance of their micro‐architectural parameters at tibia and radius affecting both trabecular and cortical parameters. IMO F + subjects were significantly older than IMO F − subjects (58 ± 8 vs. 53 ± 9 yrs, p = 0.01). BMD Z-score at the total-hip was significantly lower in IMO F + (− 1.3 ± 0.5 vs. − 0.9 ± 0.8 g/cm 2 , p = 0.01). After adjustment, trabecular micro‐architectural parameters, biochemical markers and hormonal parameters were not different in the 2 groups. At distal tibia, cortical v-BMD was significantly lower in IMO F + patients (799 ± 73 vs. 858 ± 60 mg/cm 3 , p = 0.03), while cortical thickness was not different. Conclusion Our results show that patients with IMO display a marked disturbance of trabecular and cortical bone micro-architecture, and that age and low cortical density are determinants of the fracture occurrence.

SOST encodes sclerostin, an inhibitor of bone formation that antagonizes canonical Wnt signaling. Variations of SOST expression have an impact on bone mineral density (BMD) and bone strength. We hypothesized that genetic and epigenetic DNA modifications have an impact on SOST gene expression. By analyzing 43 bone samples from women, we found that rs851054 (G/A) is associated with SOST mRNA expression, donors with one or two G allele(s) displaying higher SOST expression (p

Degenerative scoliosis usually begins at menopause and lateral rotatory olisthesis (LRO) might be a triggering factor in the onset of degenerative scoliosis in postmenopausal women. We set out to evaluate the influence of hormone replacement therapy (HRT) on degenerative scoliosis and on LRO.A cross-sectional study was conducted in 146 postmenopausal women: 75 women had received HRT for more than 1 year (HRT 1) and 71 women had never received HRT or less than 1 year (HRT 1). Scoliotic curve, LRO, sacral slope, lordosis, kyphosis were measured. The excess risk of LRO associated with age, BMI, isometric strength of brachial biceps, bone mineral density, lean mass and HRT was evaluated using a multiple logistic regression model.No difference was found in sacral slope, lumbar lordosis or thoracic kyphosis between both groups or in the presence of scoliosis. The prevalence of LRO was significantly lower in HRT1 than HRT1 (8 vs. 30%) while the risk was dependent on age, HRT and their interaction. LRO increased with age only in HRT1 (11% when aged ≤66 years vs. 39% when aged66 years, p = 0.013), whereas the prevalence of LRO remained stable in HRT1.LRO was significantly lower in women who received HRT. The excess risk of LRO was dependent on both age and HRT status. These findings suggest that HRT might prevent the onset of LRO, and therefore might contribute to the prevention of low back pain.

Objectives Treatment for degenerative rotator cuff disease of the shoulder includes physiotherapy. Dynamic humeral centering (DHC) aims at preventing subacromial impingement, which contributes to the disease. The goal of this study was to assess the effectiveness of DHC. Method 69 patients with shoulder pain and impingement syndrome were prospectively included in a single-centre randomised trial with a 12-month follow-up. Patients and assessor were blinded to the study hypothesis and treatment, respectively. DHC and non-specific mobilisation as control were performed for 6 weeks, in 15 supervised individual outpatient sessions, and patients performed daily home exercises. The planned primary outcome was the Constant score including subscores for pain, activity, mobility and strength at 3 months. Secondary outcomes were the Constant score and subscores at 12 months, and medication use for pain at 3 and 12 months. Results The DHC group did not differ from the control group in the total Constant score at 3 months. However, the DHC group showed a higher Constant subscore for pain (12.2 (SD 2.8) vs 9.9 (2.9), least square means difference 2.1, 95% CI 0.7 to 3.5, p=0.004). At 3 months, the DHC group also showed a higher rate of no medication use (96.7% vs 71%, proportional difference 25.7, 95% CI 3.7 to 51.9, p=0.012). There was no other intergroup difference. Conclusions There was no difference in the total Constant score between DHC and controls. However, pain was improved at 3 months after DHC. The differences found in subscores for pain should be explored in future studies. Trial registration clinicaltrials.gov Identifier: NCT 01022775.

Background fractures may have serious implications in an elderly individual, and fracture prevention may include a careful choice of medications. Design the Hypertension in the Very Elderly Trial (HYVET) was a double-blind placebo-controlled trial of a thiazide-like diuretic (indapamide 1.5 mg SR) with the optional addition of the angiotensin-converting enzyme (ACE) inhibitor (perindopril 2-4 mg). Fracture was a secondary end point of the trial. Setting HYVET recruited participants from Eastern and Western Europe, China, Australasia, and Tunisia. Subjects all participants were > or =80 years of age and hypertensive. Methods participants were randomised to receive a thiazide-like diuretic (indapamide 1.5 mg SR) +/- ACE inhibitor (perindopril 2-4 mg) or matching placebos. Incident fractures were validated and analysed based on time to first fracture. Results there were 3,845 participants in HYVET and a total 102 reported fractures (42 in the active and 60 in the placebo group). When taking only validated first fractures, 90 were included in the analyses (38 in the active and 52 in the placebo group). Cox proportional hazard regression, adjusted for key baseline risk factors, resulted in a point estimate of 0.58 (95% CI 0.33-1.00, P = 0.0498). Conclusions despite the lowering of blood pressure, treatment with a thiazide-like diuretic and an ACE inhibitor does not increase and may decrease fracture rate.

Estrogen deficiency in rats is responsible for increased osteoclastic resorption and a subsequent rapid bone loss. TGF-beta, which is known to have acute effects on bone resorption in several in vitro models, has been shown to be secreted by osteoblastic cells in vitro in response to 17 beta-estradiol, but little is known about its in vivo effects on bone resorption. We therefore decided to investigate the short-term effect of TGF-beta 1 on bone resorption in ovariectomized rats. TGF-beta 1 (0.04-20 ng/injection), or vehicle, was injected daily directly into the bone marrow space, through a thin catheter implanted in the distal end of the right femur, during 4 consecutive days, starting 14 days after the ovariectomy. Bone histomorphometry was performed in the secondary spongiosa of the metaphysis of injected femurs and compared with vehicle-injected femurs of sham ovariectomized rats. Ovariectomy was associated with a marked increase in the resorption surface, a 2-fold increase in the number of osteoclasts, and no change in the number of TRAP-positive marrow cells distant from bone surfaces. Bone resorption was significantly lower in the TGF-beta 1-injected bones of ovariectomized rats, as compared with vehicle injected bones: the osteoclast surface and the number of osteoclasts were, respectively, 11.0 +/- 5.1% versus 20.8 +/- 1.3% and 287 +/- 41 versus 505 +/- 53, in bones injected with 0.2 ng of TGF-beta 1 as compared with vehicle-injected bones (mean +/- SE, p < 0.05).(ABSTRACT TRUNCATED AT 250 WORDS)

Type I collagen represents more than 90% of bone matrix. Quantitative analysis of collagen cross-link molecules such as pyridinoline (PYD) provides valuable information on bone resorption rate. We have studied 37 hemodialysis patients who underwent a systematic transiliac bone biopsy for histomorphometry study. Eighteen of them had tetracycline double labeling, allowing to determine dynamic, in addition to static bone parameters. Measurement of serum-free PYD was performed using a new competitive enzyme immunoassay. Serum PYD values were compared with those of three other serum markers of bone metabolism, namely intact PTH (iPTH), bone-specific alkaline phosphatase (bAP), and osteocalcin, for the correlations with bone histomorphometric parameters. Serum PYD levels (mean +/- SD) were significantly higher in dialysis patients than in normal individuals, 90.6 +/- 99.6 nM versus 1.9 +/- 0.4 nM, respectively. Patients with high turnover bone disease had significantly higher serum PYD levels than patients with normal or low bone turnover, 108.8 +/- 108.0 nM versus 34.1 +/- 12.8 nM, respectively. Serum PYD levels were positively correlated with bone resorption parameters including osteoclast surface (r = 0.59, p < 0.0001) and osteoclast number/mm2 (r = 0.61, p < 0.0001), and also with bone formation parameters, osteoblast surface (r = 0.43, p < 0.008), double-labeled surface (r = 0.81, p < 0.001), and BFR (r = 0.91, p < 0.0001). The BFR was better correlated with serum PYD levels than with either serum iPTH or osteocalcin concentrations. However, correlation with serum bAP was comparable.(ABSTRACT TRUNCATED AT 250 WORDS)

The relationship between bone-resorbing cells, assessed by the presence of tartrate-resistant acid phosphatases (TRAP) and morphologic indices of bone resorption, was determined in 29 osteoporotic patients (14 postmenopausal females and 15 males) and 15 dialyzed patients. The number of TRAP-positive cells per unit of cancellous bone area (N.Oc/B.Ar) was higher in dialyzed patients than in those with osteoporosis (16.8 +/- 15.3 versus 4.95 +/- 2.86, p less than 0.05). The amount of bone resorbed at the basic multicellular unit level was estimated by calculating eroded area containing TRAP cells per bone area (E.Ar+/BA). This novel parameter was similar in dialyzed and in osteoporotic patients (41,700 +/- 28,400 versus 32,300 +/- 24,600). In contrast, trabecular spacing (Tb.Sp) was identical in both metabolic bone diseases. Trabecular width (169 +/- 38 versus 127 +/- 32 microns, p less than 0.05) and bone area were higher in dialyzed than in osteoporotic patients. N.Oc/B.Ar was significantly related to E.Ar+/BA in dialyzed (r = 0.76, p less than 0.05) but not in osteoporotic patients. Tb.Sp was significantly correlated to N.Oc/B.Ar and to the number of TRAP-positive cell nuclei per B.Ar (r = 0.44, p less than 0.05) in osteoporotic but not in dialyzed patients. This last result shows that in overt osteoporosis with thin trabeculae, trabecular spacing is related to the number of resorbing cells. In contrast, the spacing of thick trabeculae in dialysis osteodystrophy is not dependent on the number of osteoclasts.

In this study we evaluated whether the fluoride-induced increased bone formation in osteoporosis is mediated by stimulation of bone cell proliferation and/or differentiation. We analyzed the kinetics of DNA synthesis and the phenotypic features of osteoblastic cells isolated from the trabecular bone surface in relationship to histomorphometric indices of bone formation evaluated on the same bone biopsy in 12 osteoporotic patients treated with fluoride. Osteoblastic cells isolated from patients with a higher than normal bone formation rate, increased mean wall thickness of trabecular bone packets, and high trabecular bone volume after fluoride therapy displayed a higher than normal rate of DNA synthesis in vitro. The peak of [3H]thymidine incorporation into DNA, the maximal DNA synthesis, and the area under the growth curve of osteoblastic cells isolated from these patients were higher than the values in normal bone cells obtained from age-matched controls. By contrast, in vitro parameters of osteoblastic cell proliferation were not different from normal in fluoride-treated osteoporosis patients in whom bone formation was not increased, although the duration of treatment and bone fluoride content were not different. Parameters of bone cell proliferation in vitro were increased in correlation with the mean wall thickness, and the latter correlated with the trabecular bone volume, indicating that the augmentation of bone formation and bone volume induced by fluoride was paralleled by an increased proliferation of osteoblastic cells. Basal osteocalcin production (corrected for cell protein) and alkaline phosphatase activity in vitro were comparable, and the response to 1,25-dihydroxyvitamin D3 (10 nmol/liter, 48 h) was not different in normal osteoblastic cells and in cells from fluoride-treated osteoporosis patients whether they had high or normal bone formation. The results show that the fluoride-induced increased bone formation in osteoporotic patients is associated with an increased in vitro proliferative capacity of osteoblastic cells lining the trabecular bone surface, whereas parameters of osteoblast differentiation are not affected. The data also suggest that induction of a higher than normal bone cell proliferation is prerequisite for the stimulation of bone formation by fluoride.

INTRODUCTION previous studies have suggested that smoking, living alone and having a high body mass index may increase risk of developing dementia whereas a normal body mass index, having received education and moderate alcohol consumption may decrease risk. Dementia risk also increases with age and is thought to be higher in hypertensives. METHOD we used data collected in the Hypertension in the Very Elderly Trial (HYVET), and cognitive function was assessed using the Mini-Mental State Examination (MMSE) at baseline and annually. Participants with a fall in MMSE to

In addition to the numerous roles of histamine in both the immune and nervous systems, previous studies have suggested that this bioamine might also be involved in bone metabolism. Following our observations of impaired bone resorption in ovariectomized rats after histamine receptor antagonist treatment, we focused in this study on osteoclasts and osteoclast precursors. We looked for a direct action of histamine on these cells using both in vivo and in vitro approaches. In vivo, we triggered a remodeling sequence in rat mandibular bone and treated the animals with either histamine or histamine receptor antagonists. Histamine was shown to increase the number of osteoclasts and osteoclast precursors whereas antagonists of histamine receptor-1 and -2 decreased both osteoclast recruitment and resorption. In vitro, spleen cells from histamine-deficient mice were treated with receptor activator for nuclear factor kappa B ligand and macrophage colony stimulating factor, giving rise to both reduced numbers of osteoclasts and decreased resorption on dentin slices. Histamine enhanced resorption in these cultures in a dose-dependent manner. In addition, we identified osteoclast precursors as a source of histamine. In contrast, histamine increased the receptor activator for nuclear factor kappa B ligand/osteoprotegerin ratio in primary osteoblasts that did not secrete histamine. We observed a differential expression of histamine receptor-1 and -2 mRNAs in both primary osteoclasts and osteoblasts, confirming their functional roles with selective antagonists. Thus, histamine acts directly on osteoclasts, osteoclast precursors, and osteoblasts, promoting osteoclastogenesis through autocrine/paracrine mechanisms.

Sex steroids are major determinants of bone mass, and hormonal contraceptives may affect bone mineral density (BMD) in women. Combination contraceptives probably have no impact on BMD, except perhaps when started within 3 years after the menarche. Progestogen-only contraceptives are being increasingly used. Injectable medroxyprogesterone acetate, a potent inhibitor of gonadotropin release, can induce bone loss, most notably in young women. Other progestogens are used in lower doses that have weaker antigonadotropin effects. Levonorgestrel and etonorgestrel implants have unclear effects on BMD but are probably safe. The impact of high- and low-dose oral progestogens on BMD has not been investigated, although no adverse effects would be expected.

Bone mass is under strong genetic control, with heritability estimates greater than 50% and is likely determined by complex interactions between genetic and environmental factors.The objective of the study was to localize genes contributing to bone mineral density (BMD) variation.An autosomal genome-wide scan for BMD at the lumbar spine and femoral neck was conducted with variance components linkage methods.A total of 103 pedigrees (Network in Europe on Male Osteoporosis Family Study) ascertained through a male relative with low (Z-scoreor = -2) BMD values at either lumbar spine or femoral neck.Nonparametric multipoint logarithm of the odds ratio scores for lumbar spine and femoral neck BMD values adjusted for age, gender, and body mass index.We identified a total of eight chromosomal regions with logarithm of the odds ratio score of 1.5 or greater (Por = 5 x 10(-3)): on 1q42-43, 11q12-13, 12q23-24, 17q21-23, 21q22, and 22q11 for lumbar spine and on 5q31-33 and 13q12-14 for femoral neck BMD.Four of our detected quantitative trait loci (QTL) reached the genome-wide criteria for significant (17q,21-23, Por = 2 x 10(-5)) or suggestive (11q12-13, 22q11, and 13q12-14, Por = 7 x 10(-4)) linkage. Apart from 22q11, which is a novel QTL, all other loci provide consistent replication for previously reported QTLs for BMD and other bone-related traits. Finally, several of our specific-linkage areas encompass prominent candidate genes: type 1 collagen (COL1A1) and the sclerosteosis/van Buchem disease (SOST) genes on 17q21-23; the low-density lipoprotein receptor-related protein 5 (LRP5) gene on 11q12-13; and the rank ligand gene on 13q12-14. Further analysis of these positive regions by fine linkage disequilibrium mapping is thus warranted.

Sclerosing bone disorders are a diagnostic challenge. However, hereditary sclerosing disorders often have characteristic radiological features that allow their diagnosis. Osteocondensation can result from decreased bone resorption; malignant recessive osteopetroses have been related to mutations in several genes necessary for osteoclast function and also, more recently, to osteoclast differentiation (RANK-L). Albers-Schonberg disease or autosomal-dominant osteopetrosis type II has the characteristic 'sandwich vertebrae' and 'bone within bone' radiological features. It has been related to mutation in chloride channel 7, which is necessary for osteoclast acidification. Osteocondensation can also be related to increased bone formation. Camurati-Engelman dysplasia is a disabilitating disorder with leg pain and weakness, and thickening of the diaphysis of long bones on x ray. It is due to activating mutations in the gene encoding TGF-beta, a growth factor that increases bone formation. Other less common recessive or dominant sclerosing disorders, such as endosteal hyperostosis, sclerostosis, van Buchen disease and high bone mass syndrome, are due to mutations in two genes (LRP5 and SOST) of the Wnt pathway that induce increased osteoblast activity. Recent elucidation of the molecular mechanism responsible for several hereditary diseases with osteocondensation has improved our comprehension of bone remodelling. It has allowed the discovery of new targets for the treatment of postmenopausal osteoporosis, some of which are already being investigated in clinical trials. Molecular mechanism for some hereditary osteocondensation remains to be discovered.

International audience; Bone development is a complex process in which several cell types interact, proliferate, differentiate, and die to give rise to skeletal structures. These processes are highly integrated and require continuous and coordinated regulation by soluble molecular signals and transcription factors to assure harmonious bone development and morphogenesis. In the bone, transcription factors often have multiple functions and control the differentiation of more than one skeletal cellular component. In particular, Distal-less (Dlx) homeobox transcription factors play a central role in regulating the proliferation and differentiation of the three major cell types that constitute bone: chondrocytes, osteoblasts, and osteoclasts. The aim of this review is to summarize what is known about the role of Dlx genes in osteogenesis and to emphasize their role as coordinators at different levels of skeletal development. The elucidation of these unifying roles could improve our understanding not only of bone development but also of adult bone anabolism and catabolism resulting in bone homeostasis and might thus help to further our understanding of the genetic factors responsible for predisposition to multifactorial conditions such as osteoporosis.

The LDL receptor-related protein 5 (LRP5) is a member of the LDL receptor family, which also includes the VLDL receptor and the apolipoprotein E receptor 2. The LRP5 is a co-receptor of Wnt located on the osteoblast membrane between two other receptors, Frizzled and Kremen. Frizzled and LRP5 bind to Wnt, thereby stabilizing beta-catenin and activating bone formation. When the dickkopf protein (Dkk) binds to Kremen and LRP5, this last undergoes internalization and therefore becomes unable to bind Wnt; this leads to degradation of beta-catenin and to inhibition of bone formation. In humans, loss of LRP5 function causes osteoporosis-pseudoglioma syndrome, which is characterized by congenital blindness and extremely severe childhood-onset osteoporosis (lumbar spine Z-score often < -4) with fractures. The G171V mutation prevents Dkk from binding to LRP5, thereby increasing LRP5 function; the result is high bone mass due to uncoupling of bone formation and resorption. The Z-scores in this condition can exceed +6 at the hip and spine. The LRP5 and Wnt/beta-catenin reflect the level of bone formation and play a central role in bone mass accrual and normal distribution. Furthermore, LRP5 may contribute to mediate mechanical loads within bone tissue. Identification of the Wnt/beta-catenin pathway is a breakthrough in the elucidation of pathophysiological mechanisms affecting bone tissue and suggests new treatment targets for patients with osteoporosis or specific malignant conditions such as myeloma and sclerotic bone metastases.

Chronic thiazide treatment is associated with high BMD. We report that patients and mice with null mutations in the thiazide-sensitive NaCl cotransporter (NCC) have higher renal tubular Ca reabsorption, higher BMD, and lower bone remodeling than controls, as well as abnormalities in Ca metabolism, mainly caused by Mg depletion. Introduction: Chronic thiazide treatment decreases urinary Ca excretion (UVCa) and increases BMD. To understand the underlying mechanisms, Ca and bone metabolism were studied in two models of genetic inactivation of the thiazide-sensitive NaCl cotransporter (NCC): patients with Gitelman syndrome (GS) and Ncc knockout (Ncc−/−) mice. Materials and Methods: Ca metabolism was analyzed in GS patients and Ncc−/− mice under conditions of low dietary Ca. BMD was measured by DXA in patients and mice, and bone histomorphometry was analyzed in mice. Results: GS patients had low plasma Mg. They exhibited reduced UVCa, but similar serum Ca and GFR as control subjects, suggesting increased renal Ca reabsorption. Blood PTH was lower despite lower serum ionized Ca, and Mg repletion almost corrected both relative hypoparathyroidism and low UVCa. BMD was significantly increased in GS patients at both lumbar (+7%) and femoral (+16%) sites, and osteocalcin was reduced. In Ncc−/− mice, serum Ca and GFR were unchanged, but UVCa was reduced and PTH was elevated; Mg repletion largely corrected both abnormalities. Trabecular and cortical BMD were higher than in Ncc+/+ mice (+4% and +5%, respectively), and despite elevated PTH, were associated with higher cortical thickness and lower endosteal osteoclastic surface. Conclusions: Higher BMD is observed in GS patients and Ncc−/− mice. Relative hypoparathyroidism (human) and bone resistance to PTH (mice), mainly caused by Mg depletion, can explain the low bone remodeling and normal/low serum Ca despite increased renal Ca reabsorption.

Background. End-stage renal disease is often associated with altered bone metabolism. Methods. In order to investigate the determinant factors of bone mineral density (BMD) and the risk factors of fractures, we studied 70 patients; 26 women (23 post-menopausal) and 44 men, (mean ± SD) aged 60.5 ± 14.3 years, treated by standard haemodialysis (HD) for 6.4 ± 6.8 years. Main circulating bone biochemical markers were assessed and BMD was measured with a Lunar DPX densitometer at five sites. BMD results are expressed as a function of age and gender (Z-score). Results. Mean Z-score was markedly decreased at the mid-radius (-2.75 ± 1.23) whereas it was normal at the femoral neck (-0.42 ± 1.13) and lumbar spine (0.02 ± 2.13), and total body (-0.62 ± 1.53). Time on HD was negatively correlated to the Z-score at the mid-radius and total body but not at the other sites. Serum intact parathyroid hormone (iPTH), whole PTH or cyclase activating PTH (CAP) and bone-specific alkaline phosphatase concentrations were negatively correlated with Z-scores at all sites. Twenty-one out of 70 patients had sustained a total of 27 fractures since the beginning of dialysis therapy (seven ribs, seven ankles, six vertebrae, three humerus, two wrists and two hips). They had a total body Z-score significantly lower than that of patients without fractures,-1.34 ± 1.54 vs -0.37 ± 1.46, respectively (P< 0.031); however, their Z-scores at the other sites were not different. They were on HD for longer time, 10.4 ± 9.5 vs 5.0 ± 5.1, respectively (P

Background. Osteomalacia is now a rare disease in dialysis patients in developed countries since the withdrawal of aluminium overload. The involvement of fluoride and strontium in the pathogenesis of the disease has been suggested. The aim of this study was to investigate a possible association between osteomalacia in dialysis patients and the fluoride or strontium contents of bone. Methods. Of 271 bone biopsies from chronic haemodialysis patients referred to our centre, we studied the nine biopsies from patients with osteomalacia. They were compared with 23 biopsies from patients with hyperparathyroidism and 24 biopsies from patients with adynamic bone disease. Histomorphometric static and dynamic indices were measured. Bone fluoride and strontium contents were measured in biopsies from haemodialysis patients, and were compared with those of control patients. Results. In the nine patients with osteomalacia, we found an absence of double labelled surfaces and increased osteoid thickness. Mild aluminium overload was observed in two of the nine patients. The bone strontium content of the entire dialysis population studied was not significantly different from control values (0.023 ± 0.001 vs 0.019 ± 0.002% mol/mol, P = 0.15). However, bone strontium level was slightly but significantly increased in patients with osteomalacia (0.030 ± 0.005%), compared with both controls (0.019 ± 0.002%, P < 0.05) and the other bone diseases (0.021 ± 0.002%, P < 0.05). Bone fluoride content was significantly higher in the entire dialysis population than in the controls (0.33 ± 0.04 vs 0.13 ± 0.018% (g/g ash weight), P = 0.04). It was increased in osteomalacic patients compared with controls and with patients having hyperparathyroidism or adynamic bone disease. There was no correlation between formation indices (OV/BV, OS BS, Ob.S/BS) and bone fluoride or strontium content. Conclusions. We found a prevalence of osteomalacia of 3.3% in our biopsy series for chronic dialysis patients. However, although bone strontium and fluoride contents were slightly increased, no causal relationship with these individual metals and osteomalacia could be firmly established in this small number of patients. The hypothesis of strontium- or fluoride-induced osteomalacia in renal patients merits further investigation.

Hepatitis C-associated osteosclerosis (HCAO) is characterized by increased bone mass following hepatitis C infection. We report here a case of HCAO that lasted 8 years before the patient received antiviral hepatitis treatment. Seven years after the antiviral treatment, the evolution of radiographs and densitometry showed skeletal recovery of osteosclerosis. This case strengthens the relationship between viral infection and osteosclerosis.

International audience; High resolution-peripheral quantitative computed tomography (HR-pQCT) measurements are carried out in clinical research protocols to analyze cortical bone. Micro-computed tomography (micro-CT) is a standard tool for ex vivo examination of bone in 3D. The aim of this work was to evaluate cortical measurements derived from HR-pQCT images compared to those from synchrotron radiation (SR) micro-CT in a distal position (4.2 cm from the distal pilon). Twenty-nine tibia specimens were scanned with HR-pQCT using protocols provided by the manufacturer. The standard measured outcomes included volumetric bone density (g HA/cm3) of the cortical region (Dcomp), and the cortical thickness (Ct.Th, mm). New features, such as cortical porosity (Ct.Po) and mean pore diameter (Ct.Po.Dm), were measured by an auto-contouring process. All tibias were harvested from the posterior region and imaged with SR micro-CT (voxel size = 7.5μm). The cortical thickness, (Ct.Thmicro-CT), porosity (PoV/TV), pore diameter, pore spacing, pore number, and degree of mineralization of bone (DMB) were obtained for SR micro-CT images. For standard measurements on HR-pQCT images, site matched analyses with micro-CT were completed to obtain Dcomplocal and Ct.Thlocal. Dcomp was highly correlated to PoV/TV (r = − 0.84,p < 10− 4) but not to DMB. Dcomplocal was correlated to PoV/TV (r = − 0.72, p < 10− 4) and to DMB (r = 0.40, p > 0.05). Ct.Thlocal and Ct.Thmicro-CT were moderately correlated (r = 0.53,p < 0.01). Ct.Th and Ct.Po results from the autocontouring process are influenced by the level of trabecularization of the cortical bone and need manual correction of the endosteal contour. Distal tibia is a reliable region to study cortical bone with Dcomp as the best parameter because it reflects both the micro-porosity (Havers canals) and macro-porosity (resorption lacunae) of the cortical bone.

In chronic kidney disease the excessive production of parathyroid hormone increases the bone resorption rate and leads to histologic bone signs of secondary hyperparathyroidism. However, in other situations, the initial increase in parathyroid hormone and bone remodeling may be slowed down excessively by a multitude of factors including age, ethnic origin, sex, and treatments such as vitamin D, calcium salts, calcimimetics, steroids, and so forth, leading to low bone turnover or adynamic bone disease. Both high and low bone turnover diseases actually are observed equally in chronic kidney disease patients treated by dialysis, and all types of renal osteodystrophy are associated with an increased risk of skeletal fractures, reduced quality of life, and poor clinical outcomes. Unfortunately, the diagnosis of these bone abnormalities cannot be obtained correctly by current clinical, biochemical, and imaging methods. Therefore, bone biopsy has been, and still remains, the gold standard analysis for assessing the exact type of renal osteodystrophy. It is also the unique way to assess the mechanisms of action, safety, and efficacy of new bone-targeting therapies.

International audience; Autosomal dominant osteopetrosis type II (ADO II) is a rare, heritable bone disorder characterized by a high bone mass and insufficient osteoclast activity. Mutations in the CLCN7 gene have been reported to cause ADO II. To gain novel insights into the pathways dysregulated in ADOII osteoclasts, we identified changes in gene expression in osteoclasts from patients with a heterozygous mutation of CLCN7. To do this, we carried out a transcriptomic study comparing gene expression in the osteoclasts of patients with ADO II and healthy donors. Our data show that, according to our selection criteria, 182 genes were differentially expressed in osteoclasts from patients and controls. From the 18 displaying the highest change in microarray, we confirmed differential expression for seven by qPCR. Although two of them have previously been found to be expressed in osteoclasts (ITGB5 and SERPINE2), the other five (CES1 (carboxyl esterase 1), UCHL1 (ubiquitin carboxy-terminal esterase L1, also known as ubiquitin thiolesterase), WARS (tryptophanyl-tRNA syn-thetase), GBP4 (guanylate-binding protein 4), and PRF1) are not yet known to have a role in this cell type. At the protein level, we confirmed elevated expression of ITGB5 and reduced expression of WARS, PRF1, and SERPINE2. Transfection of ClC-7 harboring the G215R mutation into osteoclasts resulted in an increased ITGB5 and reduced PRF1 expression of borderline significance. Finally, we observed that the ADO II patients presented a normal or increased serum level of bone formation markers, demonstrating a coupling between dysfunctional osteoclasts and osteoblasts. Sphingosine kinase 1 mRNA was expressed at the same level in ADO II and control osteoclasts. In conclusion, these data suggest that in addition to an acidification dysfunction caused by the CLCN7 mutation, a change in ITGB5, PRF1, WARS, and SERPINE2 expression could be part of the osteoclastic phenotype of ADO II.

We analyzed the use of different promoters and the splicing patterns of the exons encoding 5'- and 3'-untranslated sequence amounts of parathyroid hormone-related protein (PTHrP) gene products in breast cancers. Tumor samples from 74 cases of primary breast cancer that had been followed from 1 to 14 years were selected retrospectively according to the occurrence of metastasis: 18 patients developed no metastasis (NM), 56 developed metastases (M), 22 of whom developed metastases in soft tissues (MB-) and 34 of whom developed bone metastases (MB+). The amount of the 1-139 isoform mRNA was much higher in the tumors of patients who later developed metastases (M: 0.29 +/- 0.03) than in those of patients who developed no metastases (NM, 0.13 +/- 0.03; p0.01). This isoform mRNA was also more abundant in breast tumors from patients who developed bone metastases (MB+, 0.39 +/- 0.04) than in those of patients who developed metastases in soft tissues (MB-, 0.15 +/- 0.03; p0. 0001). By contrast, the amounts of the 1-141 isoform mRNA in these three groups of tumors were similar, but its concentration was higher in the tumors of premenopausal women than in those of postmenopausal women (p0.05). Analysis with 5' untranslated regions-specific primers showed transcription from all three putative transcription start sites of PTHrP (P1, P2, and P3). The P3-initiated transcripts were more abundant in patients who developed metastases (M, 0.31 +/- 0.03) than in the nonmetastatic tumors (NM, 0.13 +/- 0.03; p0.01). The amount of P3 element did not differ with the site of metastasis (BM+, 0.32 +/- 0.05; BM-, 0. 28 +/- 0.05; NS). The same trend was observed for the P2 element. However, the use of P2-initiated messages was strongly associated with the absence of estrogen receptors from the breast tumors (p0. 01). We thus find a close association between the pattern of PTHrP gene expression and the outcome of breast cancer. The P3-initiated start site and the presence of PTHrP 139 mRNA could help identify patients at risk of developing metastases.

Studying consanguineous families with Ghosal hematodiaphyseal dysplasia syndrome (GHDD), a disorder of increased bone density, we identified mutations in TBXAS1, which encodes thromboxane synthase (TXAS). TXAS, an enzyme of the arachidonic acid cascade, produces thromboxane A(2) (TXA(2)). Platelets from subjects with GHDD showed a specific deficit in arachidonic acid-produced aggregation. We also found that TXAS and TXA(2) modulated expression of TNFSF11 and TNFRSF11B (encoding RANKL and osteoprotegerin (OPG), respectively) in primary cultured osteoblasts.

Diastrophic dysplasia (DTD) is a chondrodysplasia caused by mutations in the SLC26A2 gene, leading to reduced intracellular sulfate pool in chondrocytes, osteoblasts and fibroblasts. Hence, proteoglycans are undersulfated in the cartilage and bone of DTD patients. To characterize the bone phenotype of this skeletal dysplasia we used the Slc26a2 knock-in mouse (dtd mouse), that was previously validated as an animal model of DTD in humans. X-rays, bone densitometry, static and dynamic histomorphometry, and in vitro studies revealed a primary bone defect in the dtd mouse model. We showed in vivo that this primary bone defect in dtd mice is due to decreased bone accrual associated with a decreased trabecular and periosteal appositional rate at the cell level in one month-old mice. Although the osteoclast number evaluated by histomorphometry was not different in dtd compared to wild-type mice, urine analysis of deoxypyridinoline cross-links and serum levels of type I collagen C-terminal telopeptides showed a higher resorption rate in dtd mice compared to wild-type littermates. Electron microscopy studies showed that collagen fibrils in bone were thinner and less organized in dtd compared to wild-type mice. These data suggest that the low bone mass observed in mutant mice could possibly be linked to the different bone matrix compositions/organizations in dtd mice triggering changes in osteoblast and osteoclast activities. Overall, these results suggest that proteoglycan undersulfation not only affects the properties of hyaline cartilage, but can also lead to unbalanced bone modeling and remodeling activities, demonstrating the importance of proteoglycan sulfation in bone homeostasis., Highlights ► The osteopenic phenotype in a mouse model of proteoglycan undersulfation has been characterized. ► In vivo and in vitro studies revealed a primary bone defect in the dtd mouse model. ► Low bone mass in mutant mice is linked to bone matrix alterations triggering changes in osteoblast and osteoclast activities. ► Electron microscopy showed that collagen fibrils were thinner and less organized in mutant compared to wild-type mice. ► Results demonstrate that the SLC26A2 gene not only affects chondrogenesis, but also leads to unbalanced bone modeling and remodeling activities.

Osteoclasts (OCs), which form by fusion of hematopoietic precursor cells, are typically present in large numbers in giant cell tumors of bone (GCTBs). These tumors may, therefore, contain cells which secrete factors that stimulate recruitment and differentiation of OC precursors. Multinucleated cells resembling OCs also form in cultures of human cord blood monocytes (CBMs) stimulated by 1.25 dihydroxyvitamin D3, but these cells lack the ability to form bone resorption pits, the defining functional characteristic of mature OCs. CBMs may thus require additional stimulation to form OCs; we therefore investigated whether GCTBs are a source of such a stimulus. CBMs were stimulated in long term (21 day) culture by medium conditioned by explants of GCTBs; media collected within 15 days of explant (early-CM) and after 15 days (late-CM) were employed. We also cocultured CBMs with primary GCTB-derived stromal cells as well as immortalized bone marrow stroma-derived cells. CBMs stimulated by early-CM formed resorption pits on cortical bone slices; however, stimulation by late-CM resulted in virtually no resorption. Both early-CM and late-CM increased CBM proliferation, but not the proportion of vitronectin receptor positive or multinucleated cells. Coculture of CBMs with stromal cells of GCTBs or bone marrow did not result in bone resorption, although these stromal cells (most expressing alkaline phosphatase but progressively losing parathyroid hormone receptor expression) expressed mRNA for cytokines involved in OC differentiation, including macrophage-CSF, granulocyte-macrophage-CSF, IL-11, IL-6, and stem cell factor. Our results indicate that CBMs are capable of terminal OC differentiation in vitro, a process requiring 1,25 dihydroxyvitamin D3 as well as diffusible factor(s) which can be derived from GCTB. Stromal cells of GCTB may produce such factors in vivo, but do not support OC differentiation in vitro, possibly through phenotypic instability in culture.

We report on a family affected by Camurati-Engelmann disease, characterized by radiological signs limited to the tibia, and associated with overweight or obesity, which is not a known feature of this disorder. The affected patients were heterozygous for a c.466C > T mutation (which predicts p.Arg156Cys) in the latency associated protein (LAP)-coding domain of the TGFB1 gene. This mutation had previously been reported once in another family with a similar, atypical phenotype, which suggests a possible phenotype/genotype relationship.

Objective Subchondral bone modifications occur early in the development of osteoarthritis (OA). The level of bone resorption might impact cartilage remodeling. We therefore assessed the in vivo and in vitro effects of targeting bone resorption in OA and cartilage metabolism. Methods OA was induced by meniscectomy (MNX) in ovariectomized osteopenic mice (OP) treated with estradiol (E2), pamidronate (PAM), or phosphate buffered saline (PBS) for 6 weeks. We assessed the subchondral bone and cartilage structure and the expression of cartilage matrix proteases. To assess the involvement of bone soluble factors in cartilage metabolism, supernatant of human bone explants pre-treated with E2 or PAM were transferred to cartilage explants to assess proteoglycan release and aggrecan cleavage. OPG/RANKL mRNA expression was assessed in bone explants by real-time quantitative PCR. The role of osteoprotegerin (OPG) in the bone-cartilage crosstalk was tested using an OPG neutralizing antibody. Results Bone mineral density of OP mice and osteoclast number were restored by E2 and PAM (p

International audience; Objectives: Spousal concordance has already been observed for several other diseases but it has not yet been studied for bone loss. Our aim was to determine whether or not bone changes were the same within spouses who have shared the same environment for many decades.Methods: We conducted a prospective study on an average of 5±3 years in a single center. We evaluated in the 104 spouses who came at least twice and had more than 1 year of follow-up the changes in bone mineral density (BMD) at femoral neck adjusted on weight and height, and on hormone replacement therapy use and duration for women. Lifestyle characteristics and biological parameters were evaluated at baseline.Results: The mean (±SD) baseline age of wives was 63±5 years and that of their husbands was 66±5 years. They had been living together for 40±8 years. Most of the environmental baseline factors were correlated within partners: BMI (r=0.26, P

Sclerosing bone disorders can be subdivided according to their clinical presentation, the primarily affected cell type, and the cellular pathways. Osteoclast-rich osteopetrosis and related disorders have been related in most cases to mutations in genes required for osteoclast function. More recently, osteoclast-poor forms of osteopetrosis have been described as being connected to factors that govern osteoclast differentiation. However, increased bone formation can also cause osteosclerosis. Camurati-Engelman disease and osteopoikilosis are both related transforming growth factor-beta signaling. Rare recessive or dominant sclerosing disorders, such as endosteal hyperostosis, sclerosteosis, van Buchem disease, high bone-mass syndrome, and osteopathia striata, are caused by mutations in genes involved in the Wnt pathway, which regulates osteoblast differentiation. Finally, a third entity, including Ghosal syndrome and pachydermoperiostosis, is related to mutations in genes of the eicosanoid pathway. Clinical aspects and the consequences for our understanding of bone biology are discussed.

Osteopathia striata with cranial sclerosis (OSCS) is an X-linked dominant condition marked by linear striations mainly affecting the metaphyseal region of the long bones and pelvis in combination with cranial sclerosis. Recently, the disease-causing gene was identified as the WTX gene (FAM123B), an inhibitor of WNT signaling. A correlation was suggested between the position of the mutation and male lethality. We performed genotype and phenotype studies using 18 patients from eight families with possible WTX gene defects and expanded the clinical spectrum of the affected females. All investigated families diagnosed with OSCS had WTX gene defects. One family had a WTX gene deletion; three of four point mutations were novel. The earlier reported WTX c.1072C>T was detected in four sporadic patients and appears to be a hotspot for mutations. Based on the nature of the mutation present in a surviving male patient, our data do not support the hypothesis raised by Jenkins et al. (2009) regarding a genotype-phenotype correlation for male lethality. The finding of a gene involved in WNT signaling as the cause of this sclerosing bone phenotype is not unexpected, but further functional studies are needed to explain the specific features. The WTX gene is mutated in different types of cancer, and it remains to be explained why osteopathia striata patients appear not to have an increased risk of cancer.

Transforming growth factor-beta (TGF-beta) is a multifunctional polypeptide, abundant in bone, that regulates both proliferation and differentiation of a wide variety of cells, but its role in osteoclast differentiation remains controversial. We have recently shown that long-term cultures of human cord blood monocytes, in the presence of 1,25 dihydroxycholecalciferol (1,25-(OH)2D3), give rise to cells that express two markers of the osteoclast phenotype, namely, the vitronectin receptor (VNR) and the calcitonin receptor (CTR). TGF-beta enhanced the proportion of cells expressing the VNR. In the present study, we investigated the effect of TGF-beta on the expression of CTR in cord blood monocytes cultured during 3 weeks in the presence of 1,25-(OH)2D3. When added within the first 2 weeks of culture, TGF-beta (500 pg/ml) significantly decreased the cell protein content. TGF-beta alone did not stimulate basal cAMP production. The 10 nM-sCT-stimulated cAMP production was enhanced by increasing TGF-beta concentrations from 50 pg/ml to 1,000 pg/ml: for 500 pg/ml TGF-beta, it was 294 +/- 28% vs. 140 +/- 25% for control cultures (p less than 0.01). The sCT dose-response curves showed a higher cAMP production from 10(-9) M to 10(-7) M of sCT in the presence of 500 pg/ml TGF-beta than in control cultures. The increase was 325 +/- 36% in the presence of TGF-beta and 195 +/- 13% in the absence of TGF-beta, for 10(-7) M sCT (p less than 0.01). This effect of TGF-beta on cAMP production was not observed either when it was added to monocyte cultures the last day or 2 hours before the end of the culture or in MCF7, a human breast cancer cell line that expresses CTR. [125I]-sCT binding studies performed on confluent cells showed similar Kd in control and TGF-beta-treated cells. By contrast, the CTR number was significantly increased in the presence of TGF-beta: 6.1 +/- 2 x 10(4) receptors per cell in control cultures and 28.8 +/- 8.1 x 10(4) receptors per cell in TGF-beta-treated cultures (p less than 0.05). It is thus suggested that TGF-beta increases the number of CTR of these cells that have other features of preosteoclasts. The role of this cytokine on the process of osteoclast differentiation and in bone resorption is thus emphasized.

Objective To evaluate the efficacy of a functional restoration program for patients with chronic low back pain, using overall disability and work ability as the primary evaluation criteria. Patients and methods We prospectively studied patients aged 18 years or older who had been on sick leave because of nonspecific low back pain for at least 3 months and whose job position was still open. The program was delivered on a day-hospital basis 5 days a week for 5 weeks. Patients were followed up for 1 year. Results We included 39 patients, 11 females and 28 males with a mean (± SD) age of 43 ± 8 years and a mean sick-leave duration of 10 ± 7 months. After 1 year, 26 (67%) patients reported improvements and 25 (64%) had returned to work. Compared to the year before the program, the number of sick leave days was decreased by 51% (120 ± 140 vs. 244 ± 114, P

International audience; In men, vertebral fractures are poorly associated with bone density, and both cortical and trabecular micro-architectural changes could contribute to bone fragility. Bone histomorphometry makes it possible to investigate both the thickness and porosity of cortical bone, which has been reported to have a major impact on the biomechanical properties of bone. We therefore conducted a cross sectional study using iliac crest biopsies to investigate the trabecular and cortical bone structure in men with or without vertebral fractures.We selected 93 bone biopsies from men with idiopathic osteoporosis (defined as a T-score

Udgivelsesdato: 2008-Aug We collected population-based young normal hip and spine BMD data from 17 centres across Europe to assess between centre differences and to compare reference values with the US NHANES-III data. There was strong evidence of between country heterogeneity, but not between centres within countries. Hip BMD mean values were lower in European women, but SD's differed little from the NHANES-III USA results in both sexes. It may be necessary to adjust NHANES-III based T-scores by adding/subtracting a country-specific adjustment factor. INTRODUCTION: It remains unclear whether young normal BMD reference values specific to an American population can be validly used for T-score calculation in Europeans. METHODS: We collected population based BMD data from 1163 men and 329 women aged 19-29 years from 17 centres across Europe to compare mean and SD values with the NHANES-III study USA results. BMD(g/cm(2)) was measured at the hip and spine using DXA densitometers cross-calibrated with the European Spine Phantom (ESP). The only exclusions were for technically inadequate scans. A linear regression model was used to derive reference values. To allow for direct comparison with published NHANES III study data, the cross-calibrated BMD values were converted using the ESP equations to Hologic QDR 1000 units. RESULTS: In men, the overall mean(SD) BMD values expressed in Hologic-QDR1000 units of measurement, were: femoral neck 0.912(0.132); trochanter 0.793(0.124); and L2-L4 spine 1.027(0.123). The respective estimates in women were: 0.826(0.115); 0.670(0.093); and 0.983(0.107). However the I(2) statistic for heterogeneity indicated moderate to strong evidence of between-centre heterogeneity. There was, however, no significant heterogeneity observed between centres within countries, suggesting that this variation arose from national differences. Compared to the NHANES III population-based US data, the mean values in women were significantly lower at both sites due to some lower national European means. However, at all sites and in both sexes the SD's were very similar between the US and Europe. There was some evidence that recruiting volunteers resulted in biased values in women. CONCLUSION: Our T-score normal values for the lumbar spine (L2-L4) should be more reliable for spine-specific risk assessment than some non-representative normal ranges, and should be evaluated for that purpose in Europe. If T-scores are to be used to compare individual data with ranges seen in normal young subjects of the same nationality, it may be necessary to adjust femoral NHANES III-based T-scores by adding (or subtracting) a country-specific adjustment factor. In risk assessment it is probably sufficient to use NHANES III-based hip T-scores, as supplied for the hip by densitometer manufacturers, interpreting them in light of recent international meta-analysis data on the relationship between BMD and fracture risk.

An inverse relationship between arterial calcifications and bone activity has been documented in patients with ESRD. Calcium overload is associated with arterial calcification, which is associated with arterial stiffening. Whether bone activity interacts with calcium load, aortic stiffness, or arterial calcification is unknown. This study assessed the impact of bone activity on the relationships between the dosage of calcium-containing phosphate binders and aortic stiffness (measured by pulse wave velocity) or abdominal aorta calcification score. Aortic stiffness and calcification were both positively associated with calcium load and negatively associated with bone activity. A significant interaction was found between dosage of calcium-containing phosphate binders and bone activity such that calcium load had a significantly greater influence on aortic calcifications and stiffening in the presence of adynamic bone disease. Independent of any other factor, including dosage of calcium-containing phosphate binders, adynamic bone was associated with greater aortic stiffening, suggesting cross-talk between the bone and arterial walls.

For the treatment of secondary hyperparathyroidism (HPTH-II) in dialysis patients and hypercalcemia in patients with parathyroid carcinoma. Calcimimetics are a new class of drugs approved in the European Community and the United States by the Food and Drug Administration that were designed to suppress parathyroid hormone (PTH) levels with a simultaneous reduction in serum calcium and phosphorus levels, and calcium phosphorus product (Ca x P). Hypocalcemia is a frequent finding during the correction phase of the HPTH-II with calcimimetics. By contrast, the appearance of a hypercalcemia has yet to be described. In this paper, we report a case of severe hypercalcemia of immobilization in a 40-year-old hemodialyzed woman treated by cinacalcet HCl for a severe HPTH-II (PTH>1,000 pg/mL). A kidney transplantation recipient 1983 to 1995, she was diagnosed with Charcot-Marie Tooth disease in 1991. She had multiple orthopedic interventions for kidney-related osteoarticular problems probably favored by the kidney graft and the immunosuppressive treatment. While she was receiving the maximum dose of 180 mg/day of cinacalcet HCl and PTH at 443 pg/mL, she needed to be hospitalized for a right hip prothesis. Two weeks after the intervention she developed a symptomatic hypercalcemia of 3.57 mmol/L which was resistant to several measures including lowering the calcium concentration in the dialysate, withdrawing all vitamin D and calcium supplementation and the administration of calcitonin. Her serum calcium level was finally stabilized in the 2.37-2.95 mmol/L by administration of a single intravenous dose of pamidronate. This observation illustrates that the pharmacological activation of the parathyroid CaR and other putative CaR on bone cells by calcimimetics did not protect against the occurrence of hypercalcemia of immobilization favored by a severe HPTH-II in a hemodialysis patient.

Osteoporosis is one of the most common degenerative diseases. It is characterized by reduced bone mineral density (BMD) with an increased risk for bone fractures. There is a substantial genetic contribution to BMD, although the genetic factors involved in the pathogenesis of human osteoporosis are largely unknown. Mice with a targeted deletion of either the cannabinoid receptor type 1 (Cnr1) or type 2 (Cnr2) gene show an alteration of bone mass, and pharmacological modification of both receptors can regulate osteoclast activity and BMD. We therefore analyzed both genes in a systematic genetic association study in a human sample of postmenopausal osteoporosis patients and matched female controls. We found a significant association of single polymorphisms (P = 0.0014) and haplotypes (P = 0.0001) encompassing the CNR2 gene on human chromosome 1p36, whereas we found no convincing association for CNR1. These results demonstrate a role for the peripherally expressed CB2 receptor in the etiology of osteoporosis and provide an interesting novel therapeutical target for this severe and common disease.

Arterial calcification (AC) is a common complication of end-stage renal disease (ESRD). The mechanisms responsible are complex, including disturbances of mineral metabolism and active expression of various mineral-regulating proteins. An inverse relationship between AC and bone density has been documented in uremic patients. In the study presented here, which included 58 patients with ESRD on hemodialysis (HD), bone-histomorphometry characteristics were compared with the AC scores (0 to 4) determined according to the number of arterial sites with calcifications. Patients with AC scores of 0 (no calcifications), or 1 or 2 (mild calcifications) had similar serum parathyroid hormone levels and bone histomorphometry, with larger osteoclast resorption, higher osteoclast numbers, and larger osteoblastic and double tertracycline-labeled surfaces. In contrast, patients with high AC scores (3 and 4) were characterized by lower serum parathyroid hormone, low osteoclast numbers and osteoblastic surfaces, smaller or absent double tetracycline-labeled surfaces, and high percentages of aluminum-stained surfaces. According to multivariate analysis, AC score was positively associated with age (P < 0.0001), daily dose of calcium-containing phosphate binders (P = 0.009), and bone aluminum-stained surfaces (P = 0.037), and an inverse correlation was observed with osteoblastic surfaces (P = 0.001). A high AC score is associated with bone histomorphometry suggestive of low bone activity and adynamic bone disease. These findings suggest that therapeutic interventions associated with excessive lowering of parathyroid activity (parathyroidectomy, excessive calcium or aluminum load) favor lower bone turnover and adynamic bone disease, which could influence the development and progression of AC.

The physiological maintenance of bone mass is ensured by bone tissue renewal, allowing old bone tissue to be replaced by an equivalent mass of bone matrix. After mechanical or hormonal stress activation, a phase of resorption by osteoclasts occurs, followed by a phase of bone formation by osteoblasts. Among the multiple factors involved in osteoblastic differentiation are the following: Cbfa1 (transcription factor); low density lipoprotein receptor-related protein-5 (LRP-5), a membrane lipoprotein receptor and protein Wnt co-receptor, which plays an important role during development. It is possible that osteoblastic proliferation and differentiation are regulated by distinct pathways. Osteoclastic differentiation is also regulated by numerous factors: osteoprotegerin (OPG); RANK-L (receptor activator of nuclear factor kappa B ligand, a transmembrane protein related to tumour necrosis factor [TNF], the binding of which to its RANK receptor induces osteoclastic differentiation); and soluble TNF receptors. Local regulation of the OPG/RANK-L ratio could explain postmenopausal loss of bone mass. OPG could play a major role in myeloma and Paget disease. Osteolysis together with bone metastasis could also be related to the local production of RANK-L. OPG decreases osteolysis and offers an interesting therapeutic perspective for the treatment of osteoporosis and other diseases associated with bone hyper-resorption.

The molecular characterization of the human PLC beta1 gene was just reported by Peruzzi et al. [Biochim. Biophys. Acta 1582 (2002) 46]. This prompted us to investigate the effects of dexamethasone on PLC beta1 expression in two types of human vascular smooth muscle cells--coronary artery smooth muscle cells (hCASMC) and aortic smooth muscle cells (hAoSMC), since glucocorticoids are known to affect the signaling pathways of Gprotein coupled receptors. Semi-quantitative RT-PCR was used to analyze mRNA expression and Western-blot for protein expression. Dexamethasone treatment in the two types of cells studied decreased (mRNA and protein) PLC beta1 isozyme expression. A rapid (2 h) fall in mRNA occurred in hCASMC after treatment, and hCASMC were more sensitive to dexamethasone (1 nM versus 100 nM) than hAoSMC. The major reduction (80%) was observed after 48 h of exposure in both VSMC. Treatment with mifeprisone, an antagonist of glucocorticoid receptors, blunted the dexamethasone effect on PLC beta1 mRNA and showed that this effect was mediated by glucocorticoids receptors.

Parathyroid hormone-related protein (PTHrP) is expressed in the mammary gland and appears to be critical to the morphogenesis of this structure. PTHrP production in the breast is generally attributed to epithelial cells. Because the stromal component of the breast produces factors implicated in proliferation and differentiation of mammary epithelial tissue and tumors, the aim of this study was to investigate the PTHrP expression by mammary fibroblasts from breast cancer tumors and normal breast. PTHrP antibodies labeled intralobular fibroblasts in normal breast and stromal fibroblasts that surround tumor cells. PTHrP was constitutively produced by the cultured mammary fibroblasts, independent of serum stimulation. Normal (15.83 +/- 1.72 fmol/10(6) cells) and pathological breast fibroblasts (19.87 +/- 5.76) secreted similar amounts of PTHrP. PTH/PTHrP receptor mRNA was detected by RT-PCR in all the samples tested. Fibroblasts from normal breast were both PTH and PTHrP-cAMP responsive (453 +/- 133% and 513 +/- 133%, respectively, from basal stimulation), whereas pathological breast fibroblasts were minimally PTHrP-cAMP responsive (183 +/- 36%). The production of other fibroblastic factors implicated in tumor growth and invasiveness was also examined. Interleukin-6 (IL-6), tumor necrosis factor-alpha (INF-alpha), and pro-matrix metalloproteinase (MMP)-1 were not affected by the status of the tissue. In contrast, increased levels of pro-MMP-2 were produced in fibroblasts that originated from pathological (290 +/- 62 ng/10(6) cells) samples compared with those from normal donors (125 +/- 41 ng/10(6) cells). PTHrP production was correlated with TNF-alpha and pro-MMP-2 production. However, inhibition with specific neutralizing antibodies against TNF-alpha or PTHrP, or with a PTHrP antagonist, showed that these factors did not regulate each other. In conclusion, breast fibroblasts are constitutive PTHrP-producing cells with the potential for autocrine signaling through the PTH/PTHrP receptor.

Estrogen (E2) deficiency is responsible for increased bone turnover in the postmenopausal period, and it can be prevented by estrogen replacement therapy. The way estrogen acts on bone cells is not fully understood. Human bone marrow cell cultures may be a reliable model for studying the action of steroids on osteoclastogenesis in vitro. We examine the effects of estradiol and Raloxifene, a selective estrogen receptor modulator, on human primary bone marrow cells cultured for 15 days. 17beta-estradiol and Raloxifene significantly decreased the number of tartrate-resistant acid phosphatase multinucleate cells from osteoclast precursors on day 15. Estrogen receptor alpha (ER-alpha) mRNA was present in bone marrow mononuclear cells cultured for 5 days, but there was no estrogen receptor beta (ER-beta) mRNA, suggesting that this effect was mediated by ER-alpha. 15-day cultures no longer contained ER-alpha mRNA, suggesting that estrogen acts on early events of osteoclast differentiation. Finally, 10-8 M 17beta-estradiol has no effect on the release of IL-6 and IL-6-sr into the medium of marrow mononuclear cells cultured for 5 or 15 days. Osteoclast apoptosis was not affected by estradiol or Raloxifene after 15 days of culture under our conditions. In conclusion, we have shown that both estradiol and Raloxifene inhibit osteoclast differentiation in human bone marrow mononuclear cultures. The biological effect that can mimic in vivo differentiation could be mediated through ER-alpha.

Circulating biochemical markers of bone remodeling in uremic patients. Chronic renal failure is often associated with bone disorders, including secondary hyperparathyroidism, aluminum-related low-turnover bone disease, osteomalacia, adynamic osteopathy, osteoporosis, and skeletal β2-microglobulin amyloid deposits. In spite of the enormous progress made during the last few years in the search of noninvasive methods to assess bone metabolism, the distinction between high- and low-turnover bone diseases in these patients still frequently requires invasive and/or costly procedures such as bone biopsy after double tetracycline labeling, scintigraphic-scan studies, computed tomography, and densitometry. This review is focused on the diagnostic value of several new serum markers of bone metabolism, including bone-specific alkaline phosphatase (bAP), procollagen type I carboxy-terminal extension peptide (PICP), procollagen type I cross-linked carboxy-terminal telopeptide (ICTP), pyridinoline (PYD), osteocalcin, and tartrate-resistant acid phosphatase (TRAP) in patients with chronic renal failure. Most of the observations made by several groups converge to the conclusion that serum bAP is the most sensitive and specific marker to evaluate the degree of bone remodeling in uremic patients. Nonetheless, PYD and osteocalcin, in spite of their retention and accumulation in the serum of renal insufficient patients, are also excellent markers of bone turnover. The future generalized use of these markers, individually or in combination with other methods, will undoubtedly improve the diagnosis and the treatment of the complex renal osteodystrophy.

The effect of prostaglandin E2 (PGE2) on osteoclast (OC) differentiation is unclear, either stimulator or inhibitor, depending on the in vitro system used. This probably reflects indirect mechanisms through intermediate cells. We have investigated the direct effect of PGE2 on human OC differentiation from cord blood monocytes (CBMs) in the absence of stromal cells. Macrophages and multinucleated cells (MNCs) resembling OCs form in cultures of CBMs stimulated by 1,25-dihydroxyvitamin D3. In the present study, CBMs were cultured for 3 weeks, as previously described, in the presence or absence of PGE2. The number of MNCs was significantly reduced in the presence of PGE2 as was the proliferation of cultured CBMs, assessed on day 7. Immunohistochemistry was performed to evaluate macrophage markers (CD11b and CD14) and OC marker (beta3-chain). PGE2 significantly increased the numbers of CD11b-positive and CD14-positive cells, whereas the number of beta3-chain-positive cells was significantly decreased. beta3-Chain, c-fos, and human calcitonin receptor (h-CTR) messenger RNA (mRNA) expressions were evaluated by reverse transcription-PCR with RNA extracted from cultured CBMs. In the presence of PGE2, expression of beta3-chain and c-fos mRNA was reduced from the first week of culture. h-CTR mRNA expression was also reduced, and only the h-CTR1 isoform was detected in the presence of PGE2. In addition, when PGE2 was added only during the last week of culture, when no CBM proliferation occurred, the number of CD11b- and beta3-positive cells was unchanged compared to that in the control culture, as were the proportion of MNCs, the fusion index, and the expression of c-fos mRNA. In conclusion, our results suggest that PGE2 has an inhibitory effect on human OC differentiation from CBMs, possibly by reducing precursor proliferation in these cultures. We also hypothesize that PGE2 may reduce OC differentiation by increasing the proportion of precursor cells that differentiate into macrophages. In addition, this may be the result of inhibition of the c-fos expression in CBMs.

Cytokines and growth factors are important in bone tissue as mediators of cell-to-cell and matrix-to-cell communication. Cytokines also locally mediate the effects of several hormones on bone cells. Indeed, calciotropic hormones modulate the bone-cell production rate of these factors and, conversely, can change the number of receptors for these hormones on bone cells. Most cytokines are active in bone, but the existence of a bone-specific cytokine is still questioned. Recent work has searched for estradiol modulation of osteoblastic cytokine acting on osteoclast differentiation. In mice, increased interleukin-6 production by osteoblasts is responsible for increased bone resorption occurring after ovariectomy. Interleukin-6 could also be an autocrine or paracrine factor in the pathogenesis of increased resorption occurring in giant cell tumor or in Paget's disease. During osteoporosis and age-related bone changes, modifications of the production of insulin-like growth factor I or of one of its binding proteins could be responsible for low bone formation. Interrelationships between cytokines and hormones can affect the action of parathyroid hormone-related peptide on bone cells. The field of implication of cytokines in metabolic bone disease is growing.