Inhibition of sphingosine 1-phosphate protects mice against chondrocyte catabolism and osteoarthritis.

OBJECTIVE: Cartilage loss observed in osteoarthritis (OA) is prevented when osteoclasts in the subchondral bone are inhibited in mice. Here, we investigated the role of the osteoclast secretome and of the lipid mediator sphingosine 1-phosphate (S1P) in chondrocyte metabolism and OA. MATERIALS AND METHODS: We used SphK1LysMCre and wild type mice to assess the effect of murine osteoclast secretome in chondrocyte metabolism. Gene and protein expressions of matrix metalloproteinase (Mmp) were quantified in chondrocytes and explants by RT-qPCR and Western blots. SphK1LysMCre mice or wild type mice treated with S1P2 receptor inhibitor JTE013 or anti-S1P neutralizing antibody sphingomab are analyzed by OA score and immunohistochemistry. RESULTS: The osteoclast secretome increased the expression of Mmp3 and Mmp13 in murine chondrocytes and cartilage explants and activated the JNK signaling pathway, which led to matrix degradation. JTE013 reversed the osteoclast-mediated chondrocyte catabolism and protected mice against OA, suggesting that osteoclastic S1P contributes to cartilage damage in OA via S1P/S1P2 signaling. The activity of sphingosine kinase 1 (SphK1) increased with osteoclast differentiation, and its expression was enhanced in subchondral bone of mice with OA. The expression of Mmp3 and Mmp13 in chondrocytes was low upon stimulation with the secretome of Sphk1-lacking osteoclasts. Cartilage damage was significantly reduced in SphK1LysMCre mice, but not the synovial inflammation. Finally, intra-articular administration of sphingomab inhibited the cartilage damage and synovial inflammation. CONCLUSIONS: Lack of S1P in myeloid cells and local S1P neutralization alleviates from osteoarthritis in mice. These data identify S1P as a therapeutic target in OA.

The Matrilin-3 T298M mutation predisposes for post-traumatic osteoarthritis in a knock-in mouse model.

OBJECTIVE: The human matrilin-3 T303M (in mouse T298M) mutation has been proposed to predispose for osteoarthritis, but due to the lack of an appropriate animal model this hypothesis could not be tested. This study was carried out to identify pathogenic mechanisms in a transgenic mouse line by which the mutation might contribute to disease development. METHODS: A mouse line carrying the T298M point mutation in the Matn3 locus was generated and features of skeletal development in ageing animals were characterized by immunohistology, micro computed tomography, transmission electron microscopy and atomic force microscopy. The effect of transgenic matrilin-3 was also studied after surgically induced osteoarthritis. RESULTS: The matrilin-3 T298M mutation influences endochondral ossification and leads to larger cartilage collagen fibril diameters. This in turn leads to an increased compressive stiffness of the articular cartilage, which, upon challenge, aggravates osteoarthritis development. CONCLUSIONS: The mouse matrilin-3 T298M mutation causes a predisposition for post-traumatic osteoarthritis and the corresponding knock-in mouse line therefore represents a valid model for investigating the pathogenic mechanisms involved in osteoarthritis development.

Identification of TGFß signatures in six murine models mimicking different osteoarthritis clinical phenotypes.

OBJECTIVE: TGFß is a key player in cartilage homeostasis and OA pathology. However, few data are available on the role of TGFß signalling in the different OA phenotypes. Here, we analysed the TGFß pathway by transcriptomic analysis in six mouse models of OA. METHOD: We have brought together seven expert laboratories in OA pathophysiology and, used inter-laboratories standard operating procedures and quality controls to increase experimental reproducibility and decrease bias. As none of the available OA models covers the complexity and heterogeneity of the human disease, we used six different murine models of knee OA: from post-traumatic/mechanical models (meniscectomy (MNX), MNX and hypergravity (HG-MNX), MNX and high fat diet (HF-MNX), MNX and seipin knock-out (SP-MNX)) to aging-related OA and inflammatory OA (collagenase-induced OA (CIOA)). Four controls (MNX-sham, young, SP-sham, CIOA-sham) were added. OsteoArthritis Research Society International (OARSI)-based scoring of femoral condyles and ribonucleic acid (RNA) extraction from tibial plateau samples were done by single operators as well as the transcriptomic analysis of the TGFß family pathway by Custom TaqMan® Array Microfluidic Cards. RESULTS: The transcriptomic analysis revealed specific gene signatures in each of the six models; however, no gene was deregulated in all six OA models. Of interest, we found that the combinatorial Gdf5-Cd36-Ltbp4 signature might discriminate distinct subgroups of OA: Cd36 upregulation is a hallmark of MNX-related OA while Gdf5 and Ltbp4 upregulation is related to MNX-induced OA and CIOA. CONCLUSION: These findings stress the OA animal model heterogeneity and the need of caution when extrapolating results from one model to another.

Microcracks in subchondral bone plate is linked to less cartilage damage.

OBJECTIVES: Osteoarthritis (OA) is a disease of the whole joint characterized by cartilage loss and subchondral bone remodeling. The role of microcracks in cartilage integrity and subchondral bone homeostasis is not fully understood. The main goal of this work was to evaluate microcrack density in both calcified cartilage and subchondral bone plate in relation to cartilage damage in humans and to better define the association of microcracks and osteocyte density in subchondral bone. METHODS: We investigated 18 bone cores from cadaveric human knees that were stained with En-Bloc Basic Fuchsin. We quantified microcrack density, osteocyte density, cartilage surfaces and cartilage damage. The presence of microcracks was confirmed for each bone core by scanning electron microscopy. Finally, trabecular subchondral bone parameters were measured by micro-CT. RESULTS: Microcracks were detected in both calcified cartilage and subchondral bone plate. The density of microcracks in both calcified cartilage (CC) and subchondral bone plate (SBP) was negatively correlated with cartilage damage (r = -0.45, p < 0.05). The presence of microcracks in SBP was associated with a lower histological OA score. Osteocytes formed a dendrite network that abruptly stopped at the border of calcified cartilage. Osteocyte density in subchondral bone plate was increased in the presence of microcracks in calcified cartilage. CONCLUSIONS: Subchondral bone plate microcracks might be required for maintaining cartilage homeostasis. Microcracks in calcified cartilage may trigger osteocyte density in subchondral bone plate with subsequent regulation of subchondral bone remodeling to prevent cartilage damage.

Secular trends of hip fractures in France: impact of changing characteristics of the background population.

INTRODUCTION: Hip fractures are a societal burden because of their high morbidity and mortality and the cost they generate. With the aging of the population, worries grow about an increase of the incidence and incidence rate of hip fracture in the future. Controversial data have been provided in relation to the referencepopulation used. The aim of this study was to assess the impact of the choice of the reference population in the incidence rate of hip fracture. METHODS: Data were extracted from the French National Hospital Database related to the hospitalizations for hip fracture in France between 2002 and 2013 in patients over 59 years and were classified by gender and age (59-74, 75-84, over 84 years, over 59 years). The crude incidence rates of hip fracture were calculated by dividing the number of hospitalizations for hip fracture by the corresponding populations. To assess the impact of the choice of the reference population, we then calculated the adjusted incidence rates using direct standardization on age for the 2013 reference population. RESULTS: From 2002 to 2013, the incidence of hip fracture rose by 4.8% in women (from 49,287 to 51,661) and 21.8% in men (from 12,716 to 15,482) aged over 59 years. Meanwhile, French population over 59 years increased more with a rise of 21.3% in women and 28.7% in men, resulting in a decrease in the crude incidence rates of 13.6% in women and 5.4% in men. However, this decrease was larger after direct standardization on the 2013 population of reference as 25.6% in women and 19.2% in men as a result of a difference in age-structure of the population. CONCLUSIONS: The incidence of hip fractures continues to grow despite a reduced incidence rate throughout a 12-year-period.

Etelcalcetide: injectable calcimimetic for the treatment of secondary hyperparathyroidism in hemodialysis-dependent patients.

Chronic kidney disease is associated with mineral and bone disorders that are now considered as a syndrome. One of the major complications of this syndrome is secondary hyperparathyroidism (SHPT). SHPT increases bone turnover and the risk of fracture. SHPT is also associated with cardiovascular calcification and high mortality risk. The classical medical therapies of SHPT lack long-term efficacy and have undesirable effects on serum calcium and phosphate levels. Surgical parathyroidectomy is a radical therapeutic solution potentially exposing patients to a permanent state of hypoparathyroidism among other complications. Oral cinacalcet revolutionized the treatment of SHPT because of its great efficacy; however, more than one-third of patients do not respond appropriately to cinacalcet, mostly because of intolerance and lack of compliance. Intravenous etelcalcetide improves medical adherence and reduces pill burden. It is 10-15% superior than cinacalcet in controlling parathyroid hormone, but also leads to more frequent episodes of hypocalcemia.

Multiscale and multimodality computed tomography for cortical bone analysis.

In clinical studies, high resolution peripheral quantitative computed tomography (HR-pQCT) is used to separately evaluate cortical bone and trabecular bone with an isotropic voxel of 82 µm3, and typical cortical parameters are cortical density (D.comp), thickness (Ct.Th), and porosity (Ct.Po). In vitro, micro-computed tomography (micro-CT) is used to explore the internal cortical bone micro-structure with isotropic voxels and high resolution synchrotron radiation (SR); micro-CT is considered the 'gold standard'. In 16 tibias and 8 femurs, HR-pQCT measurements were compared to conventional micro-CT measurements. To test modality effects, conventional micro-CT measurements were compared to SR micro-CT measurements at 7.5 µm3; SR micro-CT measurements were also tested at different voxel sizes for the femurs, specifically, 7.5 µm3 versus 2.8 µm3. D.comp (r = -0.88, p < 10-3) was the parameter best correlated with porosity (Po.V/TV). The correlation was not affected by the removal of pores under 130 µm. Ct.Th was also significantly highly correlated (r = -0.89 p < 10-3), while Ct.Po was correlated with its counterpart Po.V/TV (r = 0.74, p < 10-3). From SR micro-CT and conventional micro-CT at 7.5 µm3 in matching areas, Po.V/TV and pore diameter were underestimated in conventional micro-CT with mean ± standard deviation (SD) biases of -2.5 ± 1.9% and -0.08 ± 0.08 mm, respectively. In contrast, pore number (Po.N) and pore separation (Po.Sp) were overestimated with mean ± SD biases of +0.03 ± 0.04 mm-1 and +0.02 ± 0.04 mm, respectively. The results from the tibia and femur were similar when the results of SR micro-CT at 7.5 µm3 and 2.8 µm3 were compared. Po.V/TV, specific surface of pores (Po.S/Po.V), and Po.N were underestimated with mean biases of -1.7 ± 0.9%, -4.6 ± 4.4 mm-1, and -0.26 ± 0.15 mm-1, respectively. In contrast, pore spacing was overestimated at 7.5 µm3 compared to 2.8 µm3 with mean biases of 0.05 ± 0.03 mm. Cortical bone measurements from HR-pQCT images provided consistent results compared to those obtained using conventional micro-CT at the distal tibia. D.comp was highly correlated to Po.V/TV because it considers both the micro-porosity (Haversian systems) and macro-porosity (resorption lacunae) of cortical bone. The complexity of canal organization, (including shape, connectivity, and surface) are not fully considered in conventional micro-CT in relation to beam hardening and cone beam reconstruction artifacts. With the exception of Po.V/TV measurements, morphological and topological measurements depend on the characteristics of the x-ray beam, and to a lesser extent, on image resolution.

Dementia is a major risk factor for hip fractures in patients with chronic kidney disease.

UNLABELLED: Chronic kidney disease increases the risk of hip fractures which can be promoted by dementia. We here showed that dementia increased the risk of hip fractures in dialysis patients, but in a similar manner than without dialysis. Attention should be paid to dementia to prevent hip fractures. INTRODUCTION: Hip fractures (HF) are associated with significant morbidity and is further increased in patients with chronic kidney disease (CKD). Dementia, frequent in CKD, might be a risk factor for HF. We here aimed to assess if dementia increased the risk of hip fracture in CKD. METHODS: The study was derived from the French National Database of Hospitalization. Data were obtained over the period 2011-2013. Three populations of subjects >60 years were extracted. Hip fractures, dialysis, and dementia were the main studied factors. The three populations were crossed to estimate the fracture risk based on dementia or dialysis, adjusted for age and gender. The fracture risk was calculated using a multiple logistic regression model. RESULTS: Over this period, 213,180 patients experienced a HF, 660,434 patients were diagnosed for dementia, and 47,430 patients were on dialysis. There was an effect of age and gender on the incidence of HF and dementia. In CKD patients, the risk of HF was significantly higher in demented patients compared to those without dementia: OR 2.0 [95 % CI 1.7-2.4], this being the same for men (OR 2.4 [1.8-3.1]) and women (OR 2.6 [2.0-3.3]) and at any age. However, the adjusted risk for HF in demented patients on dialysis therapy is not different than in demented patients without CKD (OR 1.3 [1.0-1.6]). CONCLUSIONS: Dementia significantly increases the risk of HF in patients on dialysis, but this risk in demented patients is equally high whether receiving dialysis therapy or not. These results highlight dementia as a major risk factor for HF in dialysis and indicate that reduction of fracture risk should include dementia as a risk factor.

Iron overload in a murine model of hereditary hemochromatosis is associated with accelerated progression of osteoarthritis under mechanical stress.

OBJECTIVE: Hereditary hemochromatosis (HH) is a disease caused by mutations in the Hfe gene characterised by systemic iron overload and associated with an increased prevalence of osteoarthritis (OA) but the role of iron overload in the development of OA is still undefined. To further understand the molecular mechanisms involved we have used a murine model of HH and studied the progression of experimental OA under mechanical stress. DESIGN: OA was surgically induced in the knee joints of 10-week-old C57BL6 (wild-type) mice and Hfe-KO mice. OA progression was assessed using histology, micro CT, gene expression and immunohistochemistry at 8 weeks after surgery. RESULTS: Hfe-KO mice showed a systemic iron overload and an increased iron accumulation in the knee synovial membrane following surgery. The histological OA score was significantly higher in the Hfe-KO mice at 8 weeks after surgery. Micro CT study of the proximal tibia revealed increased subchondral bone volume and increased trabecular thickness. Gene expression and immunohistochemical analysis showed a significant increase in the expression of matrix metallopeptidase 3 (MMP-3) in the joints of Hfe-KO mice compared with control mice at 8 weeks after surgery. CONCLUSIONS: HH was associated with an accelerated development of OA in mice. Our findings suggest that synovial iron overload has a definite role in the progression of HH-related OA.

Incidence and risk factors for hip fractures in dialysis patients.

UNLABELLED: This study described the incidence of hip fractures, associated diseases, and related costs generated in dialysis versus non-dialysis patients. INTRODUCTION: Skeletal fractures are a great concern in chronic kidney disease patients and, in particular, hip fractures that enhance the mortality. We aimed to accurately determine the incidence of hip fractures and associated diseases and to calculate the costs generated in dialysis patients. METHODS: We obtained data from the 2010 French National Hospital Database. We first extracted the hospital stays related to hip fractures as a primary diagnosis according to the ICD-10 codes and then the hospitalizations for dialysis. We compared the frequency of comorbidities in both populations. RESULTS: Among the 88,962 patients who suffered from hip fractures, 362 were on dialysis. The incidence was significantly higher in dialysis patients (x4) compared to non-dialysis patients. Women on dialysis experienced hip fractures at an earlier age than non-dialysis women. Dementia was identified as a major risk factor in the dialysis patients (72 vs. 26%, p < 0.0001). Moreover, diabetes and cardiovascular diseases were comorbidities strongly associated with hip fractures in both gender, but hypertension and malnutrition were observed exclusively in men on dialysis. Mortality rate and length of hospital stay were increased (5 days) in both genders. CONCLUSION: The incidence of hip fractures is increased in dialysis patients, affecting a larger percentage of men and women on dialysis than in the non-dialysis population and enhancing the financial burden and mortality. Dementia is a major risk factor for hip fractures in dialysis patients in addition to diabetes and cardiovascular diseases.

Animal models in OA: a means to explore bone.

Cartilage damage which characterizes osteoarthritis is accompanied with bone lesions. Joint integrity results from the balance in the physiological interactions between bone and cartilage. Several local factors regulate physiological remodeling of cartilage, the disequilibrium of these leading to a higher cartilage catabolism. Several cytokines secreted by bone cells can induce chondrocyte differentiation which suggests their role in the dialogue between both cells. Several animal models of osteoarthritis have been developed in order to assess the mechanism of cartilage loss and chondrocyte functions that encompassed surgical, chemical, or genetic approaches. Indeed, the animal models are helpful to investigate the cartilage changes in relation to changes in bone remodeling. Accumulative in vivo evidence show that increased bone resorption occurs at early stage of the development of osteoarthritis. Inhibition of bone resorbing molecules prevents cartilage damage, confirming the role of bone factors in the cross talk between both tissues. Among these numerous molecules, some participate to the imbalance in cartilage homeostasis and in the pathophysiology of osteoarthritis. These local factors are potential candidates for new drug targets.

Osteoprotegerin inhibits cartilage degradation through an effect on trabecular bone in murine experimental osteoarthritis.

OBJECTIVE: To characterize bone microarchitectural changes and to test the hypothesis that disrupting local cytokine equilibrium could modify cartilage degradation in a murine model of experimental osteoarthritis (OA). METHODS: Ten-week-old male C57BL/6 mice underwent medial meniscectomy of their right knees and a sham operation of their left knees. The mice received intraperitoneal injections of osteoprotegerin (OPG) (10 mg/kg), interleukin-1 receptor antagonist (IL-1Ra) (100 mg/kg), or phosphate buffered saline for 6 weeks. The microarchitecture of the trabecular bone, the OA score, and expression of ADAMTS-4 and ADAMTS-5 were assessed. Proteoglycan release was measured in cartilage explant cultures in the presence of IL-1Ra and OPG. RESULTS: In the meniscectomized knees, bone volume/tissue volume (BV/TV) was lower, whereas trabecular separation, the OA score, and aggrecanase expression were higher than in the sham-operated knees. After treatment with OPG, BV/TV was significantly increased and trabecular separation was reduced in the knees that underwent meniscectomy. The OA score and the number of ADAMTS-positive cells were significantly decreased by treatment with OPG but were not affected by IL-1Ra. Moreover, OPG did not directly reduce the release of proteoglycans from cartilage explant cultures. CONCLUSION: In an experimental model of OA, meniscectomy induced bone loss and cartilage degradation at 6 weeks. Systemic administration of OPG prevented bone and cartilage degradation in vivo but had no effect on cartilage in vitro. These data collectively indicate that bone could be a contributor in the early stages of OA pathogenesis. They further suggest that disruption of RANKL/OPG balance might result in the degradation of cartilage subjected to mechanical loading. Specific targeting of the bone cytokine network might help to prevent OA.

[Recommendations for the management of bone demineralization in cystic fibrosis]. / Recommandations pour la prise en charge de la déminéralisation osseuse dans la mucoviscidose.

A high prevalence of low bone mineralization is documented in adult patients with cystic fibrosis (CF). Osteopenia is present in as much as 85% of adult patients and osteoporosis in 13 to 57% of them. In children, studies are discordant probably because of different control database. Denutrition, inflammation, vitamin D and vitamin K deficiency, altered sex hormone production, glucocorticoid therapy, and physical inactivity are well known risk factors for poor bone health. Puberty is a critical period and requires a careful follow-up for an optimal bone peak mass. This review is a consensus statement established by the national working group of the French Federation of CF Centers to develop practice guidelines for optimizing bone health in patients with CF. Recommendations for screening and for calcium, vitamin D and K supplementation are given. Further work is needed to define indications for treatment with biphosphonates and anabolic agents.

Annexin 5 overexpression increased articular chondrocyte apoptosis induced by basic calcium phosphate crystals.

OBJECTIVES: Basic calcium phosphate (BCP) crystals (octacalcium phosphate (OCP), carbapatite (CA) and hydroxyapatite (HA)) are associated with severe forms of osteoarthritis. In advanced osteoarthritis, cartilage shows chondrocyte apoptosis, overexpression of annexin 5 (A5) and BCP crystal deposition within matrix vesicles. We assessed in vitro whether BCP crystals and overexpression of A5 increased chondrocyte apoptosis. METHODS: Apoptosis was induced by BCP crystals, tumour necrosis factor (TNF)-alpha (20 ng/ml) and Fas ligand (20 ng/ml) in normal articular chondrocytes (control) and in A5 overexpressed chondrocytes, performed by adenovirus infection. Apoptosis was assessed by caspase 3 (Cas3) activity, and DNA fragmentation. RESULTS: All BCP crystals, TNF-alpha and Fas ligand induced chondrocyte apoptosis as demonstrated by decreased cell viability and increased Cas3 activity and DNA fragmentation. TUNEL (terminal deoxyribonucleotide transferase-mediated dUTP nick end-labelling)-positive staining chondrocytes were increased by OCP (12.4 (5.2)%), CA (9.6 (2.6)%) and HA (9.2 (3.0)%) crystals and TNF-alpha (9.6 (2.4)%) stimulation compared with control (3.1 (1.9)%). BCP crystals increased Cas3 activity in a dose-dependent fashion. BCP-crystal-induced chondrocyte apoptosis was independent from TNF-alpha and interleukin-1beta pathways but required cell-crystal contact and intralysosomal crystal dissolution. Indeed, preincubation with ammonium chloride, a lysosomal inhibitor of BCP crystal dissolution, significantly decreased BCP-crystal-induced Cas3 activity. Finally, overexpression of A5 enhanced BCP crystal- and TNF-alpha-induced chondrocyte apoptosis. CONCLUSIONS: Overexpression of A5 and the presence of BCP crystals observed in advanced osteoarthritis contributed to chondrocyte apoptosis. Our results suggest a new pathophysiological mechanism for calcium-containing crystal arthropathies.

Complex segregation analysis accounting for GxE of bone mineral density in European pedigrees selected through a male proband with low BMD.

Osteoporosis is a common multifactorial disorder characterized by low bone mass (BMD) and high susceptibility to low-trauma fractures. Family and twin studies have found a strong genetic component in the determination of BMD, but the mode of inheritance of this trait is not yet fully understood. BMD is a complex trait whose expression is confounded by environmental influences and polygenic inheritance. Detection of potential gene-environment interactions is of great interest in the determination of bone health status. Here we have conducted segregation analyses, using the regressive class D models, in a sample of 100 European pedigrees (NEMO) with 713 subjects (524 measured for phenotypes) identified via a male with low BMD values at either the Lumbar Spine or the Femoral Neck. Segregation analyses were conducted on the residuals of LS-BMD and FN-BMD adjusted for gender, age and BMI. We tested for gene-covariate (GxE) interactions, and investigated the impact of significant GxE interactions on segregation results. Without GxE a major effect was found to be marginally significant in LS-BMD and highly significant in FN-BMD. For both traits the Mendelian hypothesis was rejected. Significant Age x gene and BMI x gene interactions were revealed. Accounting for GxE increased statistical evidence for a major factor in LS-BMD, and improved the fit of the data to the Mendelian transmission model for both traits. The best fitting models suggested a codominant major gene accounting for 45% (LS-BMD) and 44% (FN-BMD) of the adjusted BMDs. However, substantial residual correlations were also found, and these remained highly significant after accounting for the major gene.

Estradiol inhibits adhesion and promotes apoptosis in murine osteoclasts in vitro.

Osteoporosis caused by estrogen deficiency is characterized by enhanced bone resorption mediated by osteoclasts. Adhesion to bone matrix and survival of differentiated osteoclasts is necessary to resorb bone. The aim of our study was to investigate the in vitro effects of estradiol on murine osteoclasts. RAW 264.7 cells treated with 30 ng/ml RANK-L were used as a model for osteoclastogenesis. Estradiol (10(-8)M) for 5 days induced an inhibition of osteoclast differentiation and beta3 expression. Estradiol inhibited significantly the adhesion of mature osteoclasts by 30%. Furthermore estradiol-induced apoptosis shown by with nuclear condensation and Bax/Bcl2 ratio. In addition, estradiol enhanced caspase-3, -8 and -9 activities. This effect completely disappeared using specific caspase-8 inhibitor. However, increased caspase-3 activity by estradiol was observed in the presence of caspase-9 inhibitor, indicating the preferential involvement of caspase-8 pathway. Fas and FasL mRNA expression was not regulated by estradiol. However, estradiol enhanced caspase-3 activity in Fas-induced apoptosis on mature osteoclasts, suggesting that this might interact with the Fas-signaling pathway. These data suggest that estradiol decreases bone resorption by several mechanisms including adhesion and apoptosis of osteoclasts.

LRP5 gene polymorphisms and idiopathic osteoporosis in men.

Mutations in the low-density lipoprotein receptor-related protein 5 gene (LRP5) have demonstrated the role of LRP5 in bone mass acquisition. LRP5 variants were recently reported to contribute to the population-based variance in vertebral bone mass and size in males. To investigate whether LRP5 variants are implicated in idiopathic male osteoporosis, we studied 78 men with low BMD (<2.5 T score or < -2 Z score) aged less than 70 years (mean +/- SD: 50 +/- 16 years) in whom secondary causes of osteoporosis had been excluded and 86 controls (51 +/- 10 years). Genotypes and haplotypes were based on LRP5 missense substitutions in exons 9 (c.2047G > A, p.V667M) and 18 (c.4037C > T, p.A1330V), and their association with osteoporosis evaluated after adjustment for multiple clinical and environmental variables using logistic regression. The presence of osteoporosis was significantly associated with LRP5 haplotypes (P = 0.0036) independent of age (P = 0.006), weight (P = 0.004), calcium intake (P = 0.002), alcohol (P = 0.005) and tobacco (P = 0.004) consumption. Accordingly, the odds ratio for osteoporosis was 3.78 (95% CI 1.27-11.26, P < 0.001) in male carriers of haplotype 3 (c.2047A-4037T, n = 20 cases and 12 controls) versus homozygous carriers of haplotype 1 (c.2047G-4037C, n = 42 cases and 61 controls). In conclusion, these data indicate beyond a significant role for environmental factors, an association between LRP5 variants and idiopathic osteoporosis in males, pointing to a role of LRP5 in this disease.

TNF-alpha antibodies and osteoprotegerin decrease systemic bone loss associated with inflammation through distinct mechanisms in collagen-induced arthritis.

INTRODUCTION: Rheumatoid arthritis (RA) is associated with focal and systemic bone loss involving cytokines such as RANKL and TNF-alpha. RANK-L promotes focal and systemic osteoporosis, whereas osteoprotegerin (OPG) inhibits bone resorption. Although anti-TNF-alpha antibodies (anti-TNF-alpha Ab) decrease joint inflammation and bone erosions, their effects on bone loss are unknown. The aim of this study was to evaluate the effects of OPG and anti-TNF-alpha Ab, separately or in combination, on inflammation and bone remodeling in collagen-induced arthritis (CIA), a model of RA. METHODS: DBA/1 mice (n=28) were immunized with bovine type II collagen and treated with OPG-Fc or anti-TNF-alpha Ab or both, or saline. One group of mice (n=7) was not immunized (naive group). Urinary deoxypyridinoline (D-pyr) and whole-body bone mineral density (BMD) were measured at baseline and at sacrifice. Histomorphometric parameters were evaluated at the femoral metaphysis. RESULTS: Anti-TNF-alpha Ab, but not OPG, decreased the clinical arthritis score (P<0.02 vs. saline) and the histological score of inflammation. The BMD change from baseline to sacrifice (DeltaBMD) was significantly smaller in CIA mice than naive mice. OPG and anti-TNF-alpha Ab significantly increased DeltaBMD versus saline, and the effect was greater with OPG (P<0.003). DeltaD-pyr decreased by 65% with OPG and 13% with anti-TNF-alpha Ab. Compared with saline, OPG increased trabecular bone volume (BV/TV) (P<0.02), decreased trabecular separation (P<0.02), and decreased the bone formation rate (BFR) (P<0.01). Anti-TNF-alpha Ab produced no significant changes in bone volume or trabecular separation but increased trabecular thickness (P<0.02 vs. saline) to a value close to that in naive mice, suggesting preservation of bone formation. No additive effects of OPG and anti-TNF-alpha Ab were found. CONCLUSIONS: Systemic OPG and anti-TNF-alpha Ab therapy prevented bone loss in CIA mice through distinct mechanisms involving decreased bone resorption and preserved bone formation. Combining these two agents might help to prevent bone loss in inflammatory diseases.

17beta-estradiol downregulates beta3-integrin expression in differentiating and mature human osteoclasts.

The increased bone resorption observed after estrogen withdrawal is responsible for bone loss and may lead to osteoporosis. The mechanism by which estradiol inhibits bone resorption is known to involve decreased osteoclastogenesis, however, the effect on osteoclast adhesion remains unclear. We examined the in vitro effect of estradiol and raloxifene on human osteoclast differentiation and function. Human peripheral blood mononuclear cells were cultured with M-CSF/RANK-L for 18 days, and we evaluated bone resorption, the expression of the protein and mRNA of the integrins, c-jun and c-fos in the presence or absence of estradiol. In this human model, beta3-integrin expression increased at the mRNA and protein levels during osteoclast differentiation, whereas that of beta5-integrin did not. We found that estradiol and raloxifene directly inhibited bone resorption on bone slices by 50%, and decreased the expression of beta3-integrin mRNA (60%) and protein (20%) in a time-dependent manner. Moreover, the mRNAs of c-fos and c-jun were both diminished by estradiol and raloxifene, particularly in early osteoclasts, but also to a lesser extent in mature cells. These findings suggest that the direct inhibitory action of estradiol on bone resorption may affect human osteoclast differentiation through downregulation of c-fos and c-jun and adhesion through modulation of beta3-integrin.

[Genetics of osteoporosis]. / Génétique de l'ostéoporose.

Osteoporosis is a multifactorial disease involving genetic component and several environmental factors. Some rare diseases that are associated with osteoporosis such as Lobstein disease or the "pseudoglial osteoporosis" syndrom are monogenetic. Nevertheless common osteoporosis is a polygenic affection resulting from the interaction between the polymorphism of different genes and the environmental factors. The genetic component of osteoporosis encompasses roughly 60 to 70% of bone mineral density, whereas the effect on fracture risk seems lower because of the importance of other environmental factors as falls. Many polymorphisms of candidate genes involved in the regulation of bone mass have been correlated to bone density. It is likely that many genes participate to the regulation of bone density although the existence of a major gene is highly suspected. Moreover linkage analysis after genome-wide search in populations with severe osteoporosis has focused on some regions of interest (QTL) on the chromosomes. This will allow to localize one or more specific genes. The current genetic studies on different populations affected by osteoporosis or not will be useful in order to better predict the fracture risk in association with bone density and biochemical markers of bone turnover. Moreover, this will lead to the development of new treatments of osfeoporosis and will help to adapt the therapy for individual patients.