osteoprotegerin and Body-Weight

The current study examined the effects of exposure of pregnant dams to fumonisins (FBs; FB1 and FB2), from the seventh day of pregnancy to parturition, on offspring bone metabolism and properties. The rats were randomly divided into three groups intoxicated with FBs at either 0, 60, or 90 mg/kg b.w. Body weight and bone length were affected by fumonisin exposure, irrespective of sex or dose, while the negative and harmful effects of maternal FBs' exposure on bone mechanical resistance were sex and dose dependent. The immunolocalization of osteoprotegerin (OPG) and receptor activator of nuclear factor kappa-Β ligand (RANKL), in bone and articular cartilage, indicated that the observed bone effects resulted from the FB-induced alterations in bone metabolism, which were confirmed by the changes observed in the Western blot expression of OPG and RANKL. It was concluded that the negative effects of prenatal FB exposure on the general growth and morphometry of the offspring bones, as a result of the altered expression of proteins responsible for bone metabolism, were dose and sex dependent.

Topics: Animals; Body Weight; Cancellous Bone; Cartilage, Articular; Dose-Response Relationship, Drug; Female; Fumonisins; Male; Osteoprotegerin; Pregnancy; Prenatal Exposure Delayed Effects; RANK Ligand; Rats; Sex Characteristics

Periodontitis is a lifestyle-related disease that is characterized by chronic inflammation in gingival tissue. Febuxostat, a xanthine oxidase inhibitor, exerts anti-inflammatory and antioxidant effects.. The present study investigated the effects of febuxostat on periodontitis in a rat model.. Male Wistar rats were divided into 3 groups: control, periodontitis, and febuxostat-treated periodontitis groups. Periodontitis was induced by placing a ligature wire around the 2nd maxillary molar and the administration of febuxostat (5 mg/kg/day) was then initiated. After 4 weeks, alveolar bone loss was assessed by micro-computed tomography and methylene blue staining. The expression of osteoprotegerin (OPG), a bone resorption inhibitor, was detected by quantitative RT-PCR and immunological staining, and the number of osteoclasts in gingival tissue was assessed by tartrate-resistant acid phosphatase staining. The mRNA and protein expression levels of the proinflammatory cytokines, tumor necrosis factor-alpha (TNF-α), and interleukin-1 beta (IL-1β), in gingival tissue were measured using quantitative RT-PCR and immunological staining. Oxidative stress in gingival tissue was evaluated by the expression of 4-hydroxy-2-nonenal (4-HNE), and 8-hydroxy-2-deoxyguanosine (8-OHdG). To clarify the systemic effects of periodontitis, blood pressure and glucose tolerance were examined.. In rats with periodontitis, alveolar bone resorption was associated with reductions in OPG and increases in osteoclast numbers. The gingival expression of TNF-α, IL-1β, 4-HNE, and 8-OHdG was up-regulated in rats with periodontitis. Febuxostat significantly reduced alveolar bone loss, proinflammatory cytokine levels, and oxidative stress. It also attenuated periodontitis-induced glucose intolerance and blood pressure elevations.. Febuxostat prevented the progression of periodontitis and associated systemic effects by inhibiting proinflammatory mediators and oxidative stress.

Topics: Alveolar Bone Loss; Animals; Anti-Inflammatory Agents; Antioxidants; Blood Glucose; Blood Pressure; Body Weight; Disease Models, Animal; Febuxostat; Gingiva; Insulin Resistance; Interleukin-1beta; Ligation; Male; Osteoclasts; Osteoprotegerin; Oxidative Stress; Periodontitis; Rats, Wistar; Tumor Necrosis Factor-alpha; X-Ray Microtomography; Xanthine Dehydrogenase

Chronic pediatric inflammatory bowel disease (IBD) leads to lack of bone accrual, bone loss, and increased fractures. Presently there is no cure, and many IBD treatments incur negative side effects. We previously discovered treatment with exogenous irisin resolved inflammatory changes in the colon, gut lymphatics, and bone in a mild IBD rodent model. Here we assess irisin treatment in severe IBD induced via dextran sodium sulfate (DSS). Male Sprague Dawley rats (2-mo-old) were untreated (Con) or given 2% DSS in drinking water. In week two, half of each group (Con + Ir and DSS + Ir) received injections of recombinant irisin (i.p., 2x/wk). After 4 weeks, gut inflammation was associated with declines in bone mineral density and cancellous bone volume. Furthermore, elevated osteocyte TNF-α, interleukin-6, RANKL, OPG, and sclerostin corresponded with higher osteoclast surfaces and lower bone formation rate in DSS animals as well as lower ultimate load. While irisin treatment improved colon inflammation, there were no improvements in bone density or bone mechanical properties; however, irisin elevated bone formation rate, decreased osteoclast surfaces, and reduced osteocyte pro-inflammatory factors. These data highlight the negative impact of chronic gut inflammation on bone as well as the therapeutic potential of irisin as an anti-inflammatory treatment.

Topics: Animals; Biomechanical Phenomena; Body Weight; Bone and Bones; Bone Density; Bone Morphogenetic Proteins; Bone Resorption; Cancellous Bone; Colitis; Colon; Dextran Sulfate; Femur Neck; Fibronectins; Gastrointestinal Tract; Genetic Markers; Inflammation; Intestinal Mucosa; Lymphatic Vessels; Male; Osteocytes; Osteogenesis; Osteoprotegerin; RANK Ligand; Rats, Sprague-Dawley; Tibia; Tomography, X-Ray Computed; Tumor Necrosis Factor-alpha; Weight-Bearing

Topics: Animals; Body Weight; Dipsacaceae; Female; Gene Expression Regulation; Nitric Oxide Synthase Type III; Organ Size; Osteoporosis; Osteoprotegerin; Ovariectomy; Phosphatidylinositol 3-Kinases; Plant Roots; Polysaccharides; Proto-Oncogene Proteins c-akt; RANK Ligand; Rats; Rats, Sprague-Dawley; Receptor Activator of Nuclear Factor-kappa B; Signal Transduction; Uterus; Vascular Endothelial Growth Factor A

In this study, we evaluate the effects of glucagon-like peptide 2 (GLP-2) on bone microarchitecture, bone turnover markers (BTMs) and inflammation markers in ovariectomized (OVX) rats.. In total, 31 Sprague-Dawley rats were divided into the following three groups: sham (control sham-operated with vehicle, n = 7), OV (OVX with vehicle, n = 12), and GLP-2 (OVX with GLP-2, n = 12). Intervention began at the 12th week after surgery and lasted for 4 weeks. The dosage of the GLP-2 was 160 μg/kg/d through subcutaneous injections, and normal saline was used as the vehicle agent. After 4 weeks of treatment, serum BTM and inflammation marker levels were measured by ELISA, and femora samples were analyzed by qRT-PCR, micro-CT, histology and histomorphometry.. After 4 weeks of treatment, serum TRAcP-5b and RANKL levels as well as the CTX-1/P1NP ratio in the GLP-2 group decreased, and ALP activity, P1NP level, and OPG/RANKL ratio increased significantly; qRT-PCR analysis showed that mRNA levels of RANKL decreased, and Runx2, ALP, and Col-1 levels as well as the OPG/RANKL ratio increased significantly in the GLP-2 group compared with the OV group. In bone histology analysis, GLP-2 significantly decreased the AV/MV, Oc.N and Oc.S but increased the Ob.N, BFR and MAR. Analysis with μ-CT showed that the BMD, BV/TV, Tb.N and Conn.D increased significantly in the GLP-2 group compared with the OV group. The levels of serum inflammation markers TNF-α, IL-1β and IL-6 decreased, and TGF-β levels increased in the GLP-2 group compared with the OV group.. GLP-2 may have a positive impact on osteoporosis by promoting bone formation, inhibiting bone resorption and decreasing circulatory inflammation in ovariectomized rats.

Topics: Animals; Body Weight; Bone and Bones; Bone Density; Bone Remodeling; Bone Resorption; Female; Femur; Glucagon-Like Peptide 2; Osteogenesis; Osteoporosis; Osteoprotegerin; Ovariectomy; Random Allocation; Rats; Rats, Sprague-Dawley; X-Ray Microtomography

Osteoporosis is characterized by skeletal fragility and microarchitectural deterioration. The side effects of drugs to treat osteoporosis will negatively affect the health of patients. This study aimed to investigate the therapeutic effects of icariin combined with adipose-derived stem cells on osteoporosis in a postmenopausal osteoporosis model after ovariectomy in rats. After ovariectomy the rats were treated with icariin combined with adipose-derived stem cell transplantation. The levels of alkaline phosphatase, tartrate-resistant acid phosphatase, osteoprotegerin, and bone γ-carboxyglutamate protein in serum were determined by ELISA. The bone mineral density was measured by dual-energy X-ray absorptiometry. The mechanical properties were determined by a three-point bending test. The kidney functions were evaluated by an automatic analyzer and a diagnostic kit. Icariin combined with stem cells significantly reduced body weight gain caused by ovariectomy, significantly decreased alkaline phosphatase, tartrate-resistant acid phosphatase, and bone γ-carboxyglutamate protein content in serum, significantly increased osteoprotegerin content, significantly elevated bone mineral density of the lumbar spine, left femur, and right femur, and enhanced bone biomechanical properties of the femur, including maximum bending load, bending rigidity, and fracture energy, in osteoporotic rats. In addition, icariin combined with stem cells substantially decreased the damage to the liver and kidney in osteoporotic rats. Icariin combined with stem cells can not only ameliorate reduction of bone mass and disruption of the microarchitectural structure of bone tissue caused by osteoporosis in a rat model but can also have a beneficial effect on organ functions, such as those of the liver and kidney.

Topics: Absorptiometry, Photon; Adipose Tissue; Alkaline Phosphatase; Animals; Biomechanical Phenomena; Body Weight; Bone and Bones; Bone Density; Female; Femur; Flavonoids; Humans; Kidney; Liver; Osteocalcin; Osteoporosis, Postmenopausal; Osteoprotegerin; Rats, Sprague-Dawley; Stem Cell Transplantation; Stem Cells; Tartrate-Resistant Acid Phosphatase

Gfi1 is a key molecule in hematopoietic lineage development and mutations in GFI1 cause severe congenital neutropenia (SCN). Neutropenia is associated with low bone mass, but the underlying mechanisms are poorly characterized. Using Gfi1 knock-out mice (Gfi1-ko/ko) as SCN model, we studied the relationship between neutropenia and bone mass upon different pathogen load conditions. Our analysis reveals that Gfi1-ko/ko mice kept under strict specific pathogen free (SPF) conditions demonstrate normal bone mass and survival. However, Gfi1-ko/ko mice with early (nonSPF) or late (SPF+nonSPF) pathogen exposure develop low bone mass. Gfi1-ko/ko mice demonstrate a striking rise of systemic inflammatory markers according to elevated pathogen exposure and reduced bone mass. Elevated inflammatory cytokines include for instance Il-1b, Il-6, and Tnf-alpha that regulate osteoclast development. We conclude that low bone mass, due to low neutrophil counts, is caused by the degree of systemic inflammation promoting osteoclastogenesis.

Topics: Animals; Body Weight; Bone and Bones; Cell Differentiation; Congenital Bone Marrow Failure Syndromes; Cytokines; DNA-Binding Proteins; Extremities; Genotype; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Neutropenia; Osteoblasts; Osteogenesis; Osteoporosis; Osteoprotegerin; Pasteurellaceae; RANK Ligand; Transcription Factors; Trichomonas

Serum calcium (Ca) is maintained in a narrow range through regulation of Ca metabolism in the intestine, kidney, and bone. Calcium is incorporated and resorbed from bone during bone remodeling via cellular processes as well as by exchange. Both routes contribute to calcium homeostasis. To assess the magnitude of bone turnover contribution to calcium homeostasis we labeled bone with a Ca tracer and measured Ca release following stimulation or suppression of bone resorption. Young growing male rats (n = 162) were dosed with

Topics: Animals; Body Weight; Bone Resorption; Calcium; Growth and Development; Male; Models, Biological; Osteoprotegerin; RANK Ligand; Rats, Sprague-Dawley; Tartrate-Resistant Acid Phosphatase

Drynariae Rhizoma is a kidney-tonifying herb that has a long history in clinical practice for the treatment of bone fractures and joint diseases in China. Flavonoids are considered to be its major active ingredients and are reported to ease bone loss in ovariectomized rats. However, the beneficial effects of the total flavonoids of Drynariae Rhizoma on osteoporosis caused by microgravity or mechanical inactivity remain unknown. This study assessed the effects of total Drynariae Rhizoma flavonoids (DRTF, Qihuang, Beijing, China, national medicine permit No. Z20030007, number of production: 04080081, content of DRTF ≥80%) against bone loss induced by simulated microgravity. A hindlimb unloading tail-suspended rat model was established to determine the effect of DRTF on bone mineral density (BMD), biomechanical strength and trabecular bone microarchitecture. Twenty-eight male Sprague-Dawley rats were divided into four groups: the baseline, control, hindlimb unloading with vehicle (HLU), and hindlimb unloading treated with DRTF (HLU-DRTF, 75 mg/kg/day) groups. Oral DRTF was administered for 4 weeks. The underlying mechanisms of the DRTF actions on disuse-induced osteoporosis are discussed. The results showed that DRTF treatment significantly increased the BMD and mechanical strength of tail-suspended rats. Enhanced bone turnover markers with HLU treatment were attenuated by DRTF administration. Deterioration of trabecular bone induced by HLU was prevented through elevated bone volume/tissue volume (BV/TV), trabecular number (Tb. N), trabecular thickness (Tb. Th) and decreased trabecular separation (Tb. Sp). The present study provides the first evidence that DRTF prevents bone loss induced by HLU treatment, indicating its potential application in the treatment of disuse-induced osteoporosis.

Topics: Animals; beta Catenin; Body Weight; Bone and Bones; Bone Density; Flavonoids; Hindlimb Suspension; Lymphoid Enhancer-Binding Factor 1; Male; Osteoporosis; Osteoprotegerin; Polypodiaceae; RANK Ligand; Rats; Rats, Sprague-Dawley; Rhizome; Spectrometry, Mass, Electrospray Ionization; Wnt3A Protein

Exposure to high-altitude hypoxia causes physiological and metabolic adaptive changes by disturbing homeostasis. Hypoxia-related changes in skeletal muscle affect the closely interconnected energy and regeneration processes. The balance between protein synthesis and degradation in the skeletal muscle is regulated by several molecules such as myostatin, cytokines, vitamin D, and irisin. This study investigates changes in irisin and myostatin levels in male climbers after a 2-week high-altitude expedition, and their association with 25(OH)D and indices of inflammatory processes. The study was performed in 8 men aged between 23 and 31 years, who participated in a 2-week climbing expedition in the Alps. The measurements of body composition and serum concentrations of irisin, myostatin, 25(OH)D, interleukin-6, myoglobin, high-sensitivity C-reactive protein, osteoprotegerin, and high-sensitivity soluble receptor activator of NF-κB ligand (sRANKL) were performed before and after expedition. A 2-week exposure to hypobaric hypoxia caused significant decrease in body mass, body mass index (BMI), free fat mass and irisin, 25-Hydroxyvitamin D levels. On the other hand, significant increase in the levels of myoglobin, high-sensitivity C-reactive protein, interleukin-6, and osteoprotegerin were noted. The observed correlations of irisin with 25(OH)D levels, as well as myostatin levels with inflammatory markers and the OPG/RANKL ratio indicate that these myokines may be involved in the energy-related processes and skeletal muscle regeneration in response to 2-week exposure to hypobaric hypoxia.

Topics: Adult; Altitude; Body Composition; Body Mass Index; Body Weight; C-Reactive Protein; Calcifediol; Fibronectins; Humans; Hypoxia; Interleukin-6; Male; Mountaineering; Myoglobin; Myostatin; Osteoprotegerin; RANK Ligand; Young Adult

Previous studies suggested that oxidative stress is related to the onset and development of osteoporosis. Moreover, it was demonstrated that berberine has a protective effect against oxidative stress-induced injuries. In this study, we aimed to investigate the effect and mechanism of action of berberine on rats with induced osteoporosis. Sixty 8-week-old female Wistar rats were randomly divided into the following 6 groups: control saline-treated, osteoporosis saline-treated, 3 osteoporosis berberine-treated groups (Ber 5, 10, and 20 mg/kg/body weight, respectively), and osteoporosis alendronate-treated (ALD) group. Osteoporosis was induced by bilateral ovariectomy. All treatments were performed for 8 weeks. The bone mineral density (BMD), serum alkaline phosphatase (ALP), osteocalcin, calcium, phosphorus, superoxide dismutase (SOD), methylenedioxyamphetamine (MDA), and glutathione peroxidase (GSH-Px) level was determined in the rat femur tissue. The gene and protein expression of osteoprotegerin (OPG) and receptor activator of nuclear factor kappa-B ligand (RANKL) was analyzed by quantitative reverse transcription PCR and Western blot, respectively. The BMD, SOD and GSH⁃Px levels, and the expression of OPG were significantly lower in osteoporosis compared to control group (all p < 0.05). The serum levels of osteocalcin, ALP, and MDA, and the expression of RANKL were significantly higher in osteoporosis compared to control group (all p < 0.05). Berberine, especially the high doses of berberine, effectively increased SOD, GSH⁃Px, and OPG levels as well as decreased serum osteocalcin, ALP, MDA and RANKL levels in berberine-treated osteoporosis groups (all p < 0.05). To conclude, oxidative stress may promote the development of osteoporosis in rats through the RANK/RANKL/OPG pathway. The antioxidative effect of berberine reduces the development of osteoporosis in rats to some extent.

Topics: Animals; Antioxidants; Berberine; Body Weight; Bone Density; Female; Femur; NF-kappa B; Osteoporosis; Osteoprotegerin; Oxidative Stress; RANK Ligand; Rats; Rats, Wistar; Signal Transduction

Chlormequat Chloride (CCC), a widely used plant growth regulator, could decrease body weight in animals; however, the mechanism has not been well studied. This study was designed to evaluate the skeletal development toxicity of CCC on pubertal male Sprague-Dawley (SD) rats and to investigate whether CCC impacts the development of chondrocyte, osteoblast and osteoclast through growth hormone (GH) and insulin like growth factor 1 (IGF-I). Rats from 23 to 70 on postnatal days were exposed to CCC daily by gavage at doses of 0, 75, 150, and 300mg/kg bw/d. The results showed that the size of femurs and tibias, bone mineral density and biomechanical parameters were significantly decreased in the 300mg/kg bw/d group compared with the control group. The concentration of osteocalcin (OCN) and C-terminal telopeptide of type I collagen (CTX-I) in blood in the 150mg/kg bw/d group was also changed. The mRNA expression ratio of the receptor activator of NF-κB ligand (RANKL) and osteoprotegerin (OPG) in 150 and 300mg/kg bw/d group was increased. Histological analysis of proximal and distal epiphyseal plates of the right femurs showed that both the proliferative zone and hypertrophic zone narrowed in CCC-treated groups. The concentration of IGF-I in blood was reduced with an increase in exposure doses of CCC. The mRNA expression of growth hormone receptor (GHR) in tibia was decreased in the CCC-treated group. The results indicated that CCC might indirectly impact the formation and activation of chondrocytes, osteoblasts and osteoclasts because of the decline of GHR and IGF-I, leading to skeletal development damage.

Topics: Animals; Body Weight; Bone Density; Chlormequat; Chondrocytes; Collagen Type I; Dose-Response Relationship, Drug; Femur; Growth Hormone; Growth Plate; Insulin-Like Growth Factor I; Male; NF-kappa B; Osteoblasts; Osteocalcin; Osteoclasts; Osteoprotegerin; Peptides; RANK Ligand; Rats; Rats, Sprague-Dawley; Receptors, Somatotropin; RNA, Messenger; Skeleton; Tibia

Osteoprotegerin (OPG) has pleiotropic effects on bone metabolism as well as endocrine function. Our aim was to evaluate the relationship between bone mineral density (BMD) and serum OPG concentration in renal transplant recipients.. Fasting blood samples were obtained from 69 renal transplant recipients. BMD was measured in lumbar vertebrae (L2-L4) by dual-energy X-ray absorptiometry. Eight patients (11.6%) had BMD values indicative of osteoporosis, 28 patients (40.6%) had BMD values indicative of osteopenia, and 33 patients had normal BMD values. Increased serum OPG levels (P < .001), decreased body mass index (BMI) (P = .033), and decreased body weight (P = .010) were significantly correlated with low lumbar T-score cut-off points between groups (normal, osteopenia, and osteoporosis).. Women had significantly lower lumbar BMD values than men (P = .013). Menopause (P = .005), use of tacrolimus (P = .020), and use of cyclosporine (P = .046) were associated with lower lumbar BMD in renal transplant recipients. Univariate linear regression analysis revealed that lumbar BMD was positively correlated with height (P = .016), body weight (P = .001), and BMI (P = .015) and negatively correlated with age (P = .039) and log-OPG (P = .001). Multivariate linear regression analysis revealed that log-OPG (β: -0.275, R(2) change = 0.154, P = .014), body weight (β: 0.334, R(2) change = 0.073, P = .004), and age (β: -0.285, R(2) change = 0.079, P = .008) were independent predictors of lumbar BMD values in renal transplant recipients.. Serum OPG concentration correlated negatively with lumbar BMD values in renal transplant recipients.

Topics: Absorptiometry, Photon; Adult; Aged; Body Weight; Bone Density; Female; Humans; Kidney Transplantation; Lumbar Vertebrae; Male; Middle Aged; Osteoporosis; Osteoprotegerin; Transplant Recipients

Osteonecrosis, also termed aseptic necrosis, is the cellular death of bone components due to interruption of the blood supply. Glucocorticoid (GC) therapy is a common non-traumatic cause of osteonecrosis. However, the mechanism by which GCs induce osteonecrosis remains to be elucidated. The aim of the present study was to investigate the effects of GCs on osteoclast and osteoblast differentiation and function in a GC‑induced osteonecrosis mouse model. BALB/c male mice (n=40; 4‑weeks‑old) were treated with dexamethasone and asparaginase for 8 weeks. The control group (n=20) was administered normal saline. The results demonstrated that the GC-treated group had a lower mean weight compared with the control group. Morphologically, 16/37 (43%) mice demonstrated significant osteonecrotic lesions in the GC‑treated group. However, osteonecrotic lesions were not observed in the mice of the control group. Furthermore, immunohistochemistry demonstrated that the GC‑treated group had a higher level of osteoprotegerin compared with the control group, without any change in the expression of receptor activator of nuclear factor‑κB ligand. In addition, tartarate‑resistant acid-phosphatase staining demonstrated significantly decreased osteoclasts in the areas of bone destruction in the GCs-treated group. Furthermore, the present study demonstrated that GCs increased expression levels of osterix and osteocalcin, and decreased expression of matrix metallopeptidase‑9 to regulate the differentiation and function of osteoblasts and osteoclasts. The results of the present study suggested that GCs influence bone remolding resulting in decreased osteoclasts formation/differentiation. Therefore, regulating the differentiation and activity of the osteoclasts may be beneficial to the control and treatment of osteonecrosis.

Topics: Animals; Biomarkers; Body Weight; Cell Differentiation; Dexamethasone; Disease Models, Animal; Gene Expression; Glucocorticoids; Male; Matrix Metalloproteinase 9; Mice; Osteoblasts; Osteocalcin; Osteoclasts; Osteogenesis; Osteonecrosis; Osteoprotegerin; RANK Ligand; RNA, Messenger; Tartrate-Resistant Acid Phosphatase

This study was designed to investigate the anti‑osteoporotic activity of polydatin and its possible underlying mechanism. Osteoporosis was induced in mice by ovariectomy (OVX) and the mice were divided into 5 groups: An OVX only group, polydatin groups (10, 20 and 40 mg/kg) and a sham group (n=10/group). After 12 weeks of treatment, body weight, uterine index and the dry weight of thigh‑bones were recorded. In addition, the serum calcium, serum phosphorus, alkaline phosphatase (ALP) and osteoprotegerin (OPG) levels were also determined. Western blot analysis was then conducted to investigate the possible mechanism underlying the effect of polydatin via determining the expression of OPG, receptor activators of nuclear factor‑κB ligand (RANKL) and β‑catenin in the ST2 cell line. The results indicated that intraperitoneal injection of polydatin (10, 20 and 40 mg/kg/day) decreased body weight, and increased uterine index and dry weights of thigh‑bones of ovariectomized mice (P<0.05), and polydatin also significantly increased the serum calcium, phosphorus, ALP and OPG of ovariectomized mice (P<0.05). Results of western blot analysis showed that polydatin upregulated the ratio of OPG/RANKL (P<0.05) and β‑catenin protein in ST2 cells. In conclusion, the results demonstrated that polydatin exhibits anti‑osteoporotic activity via regulating osteoprotegerin, RANKL and β‑catenin.

Topics: Alkaline Phosphatase; Animals; beta Catenin; Biomarkers; Body Weight; Bone and Bones; Calcium; Cell Line; Disease Models, Animal; Female; Gene Expression Regulation; Glucosides; Mice; Osteoporosis; Osteoprotegerin; Ovariectomy; RANK Ligand; Stilbenes

Interleukin (IL)-37 is a newly discovered member of the cytokine IL-1 family. Recent evidence suggests that IL-37, an anti-inflammatory factor, may have a role in atherosclerosis. In this study we used apoE-deficient diabetic mice, an established animal model, to examine the effects of IL-37 on the progression of vascular calcification and atherosclerosis. Compared with the control groups, IL-37-treated (with injection of recombinant protein for 16 weeks) animals had significantly less calcification areas detected by both von Kossa and Alizarin Red staining, and much smaller plaque size of the atherosclerotic lesions and lower plaque vulnerability scores detected by hematoxylin-eosin staining in the aorta root. Our data also showed that IL-37 treatment caused elevated concentrations of osteoprotegerin (OPG) in serum. We detected that the group that received additional anti-OPG antibody reduced the effect of IL-37 treatment. The group that received both IL-37 and anti-OPG had significant larger percentage area of calcified lesion and atherosclerotic plaque size than the IL-37-treated group. Significant changes in disease-relevant cytokines (eg, ALP, BMP-2, TNF-α, IL-18, and IL-10) were also elicited. This is the first report that IL-37 could attenuate not only atherosclerosis, but also vascular calcification. This study may offer a therapeutic potential for the prevention and treatment of calcification and atherosclerotic disease.

Topics: Alkaline Phosphatase; Animals; Apolipoproteins E; Atherosclerosis; Biomarkers; Body Weight; Bone Morphogenetic Protein 2; Cardiotonic Agents; Cell Differentiation; Diabetes Mellitus, Experimental; Immunohistochemistry; Inflammation Mediators; Interleukin-1; Male; Mice, Inbred C57BL; Osteoblasts; Osteoprotegerin; Vascular Calcification

Osteoprotegerin (OPG) is a cytokine that regulates bone resorption by inhibiting osteoclastogenesis, and OPG has been implicated in the process that causes vascular stiffness. An increase in serum OPG level has been associated with the development of arterial stiffness. Kidney transplant (KT) patients are susceptible to aortic stiffness, which is considered to be a predictor of cardiovascular events in this patient population. Carotid-femoral pulse wave velocity (cfPWV) has emerged as a gold standard for non-invasive evaluation of aortic stiffness. The aim of this study was to evaluate the relationship between serum OPG concentration and cfPWV among KT patients. Fasting blood samples were obtained from 57 KT patients and their cfPWV was measured using applanation tonometry. The serum OPG levels were measured using an enzyme-linked immunosorbent assay. Univariable linear regression analysis showed that the cfPWV in KT patients was significantly and positively correlated with age, body weight, waist circumference, body mass index, log-creatinine, systolic blood pressure, diastolic blood pressure, pulse pressure, and the log-OPG concentration. KT patients with metabolic syndrome had higher cfPWV values than those without metabolic syndrome (P = 0.036), which indicates a higher incidence of aortic stiffness in this patient population. Multivariable forward stepwise linear regression analysis of the significant variables showed that the log-OPG (P = 0.001), the log-creatinine (P = 0.004), and the SBP (P = 0.005) remained as independent and positive predictors of cfPWV values. These findings indicate that serum OPG levels are positively associated with cfPWV in KT patients.

Topics: Age Factors; Blood Pressure; Body Mass Index; Body Weight; Creatinine; Enzyme-Linked Immunosorbent Assay; Humans; Kidney Transplantation; Linear Models; Manometry; Metabolic Syndrome; Osteoprotegerin; Pulse Wave Analysis; Vascular Stiffness

Premature osteoporosis and stunted growth are common complications of childhood chronic inflammatory disease. Presently, no treatment regimens are available for these defects in juvenile diseases. We identified the sequential Fc-OPG/hPTH treatment as an experimental therapy that improves the skeletal growth and prevents the bone loss in a mouse model overexpressing IL-6.. Premature osteoporosis and stunted growth are common complications of childhood chronic inflammatory diseases and have a significant impact on patients' quality of life. Presently, no treatment regimens are available for these defects in juvenile diseases. To test a new therapeutic approach, we used growing mice overexpressing the pro-inflammatory cytokine IL-6 (TG), which show a generalized bone loss and stunted growth.. Since TG mice present increased bone resorption and impaired bone formation, we tested a combined therapy with the antiresorptive modified osteoprotegerin, Fc-OPG, and the anabolic PTH. We injected TG mice with Fc-OPG once at the 4th day of life and with hPTH(1-34) everyday from the 16th to the 30th day of age.. A complete prevention of growth and bone defects was observed in treated mice due to normalization of osteoclast and osteoblast parameters. Re-establishment of normal bone turnover was confirmed by RT-PCR analysis and by in vitro experiments that revealed the full rescue of osteoclast and osteoblast functions. The phenotypic recovery of TG mice was due to the sequential treatment, because TG mice treated with Fc-OPG or hPTH alone showed an increase of body weight, tibia length, and bone volume to intermediate levels between those observed in vehicle-treated WT and TG mice.. Our results identified the sequential Fc-OPG/hPTH treatment as an experimental therapy that improves the skeletal growth and prevents the bone loss in IL-6 overexpressing mice, thus providing the proof of principle for a therapeutic approach to correct these defects in juvenile inflammatory diseases.

Topics: Animals; Body Weight; Bone Density Conservation Agents; Cells, Cultured; Disease Models, Animal; Drug Evaluation, Preclinical; Drug Therapy, Combination; Female; Growth Disorders; Interleukin-6; Male; Mice, Transgenic; Osteoclasts; Osteoporosis; Osteoprotegerin; Teriparatide; X-Ray Microtomography

Dietary loading has been reported to have an effect on temporomandibular joint (TMJ) remodeling via periodontal-muscular reflex. We therefore examined whether reducing dietary loading decreased TMJ degradation induced by the unilateral anterior crossbite prosthesis as we recently reported.. Forty 6-week-old female C57BL/6J mice were randomly divided into two experimental and two control groups. One experimental and one control group received small-size diet and the other two groups received large-size diet. Unilateral anterior crossbite prosthesis was created in the two experimental groups. The TMJ samples were collected 3 weeks after experimental operation. Histological changes in condylar cartilage and subchondral bone were assessed by Hematoxylin & Eosin, toluidine blue, Safranin O and tartrate-resistant acid phosphatase staining. Real-time polymerase chain reaction (PCR) and/or immunohistochemistry were performed to evaluate the expression levels of Collagen II, Aggrecan, a disintegrin and metalloproteinase with thrombospondin motifs 5 (ADAMTS-5) and RANKL/RANK/OPG in TMJ condylar cartilage and/or subchondral bone.. Thinner and degraded cartilage, reduced cartilage cellular density, decreased expression levels of Collagen II and Aggrecan, loss of subchondral bone and enhanced osteoclast activity were observed in TMJs of both experimental groups. However, the cartilage degradation phenotype was less severe and cartilage ADAMTS-5 mRNA was lower while OPG/RANKL ratio in cartilage and subchondral bone was higher in the small-size than large-size diet experimental group. No differences of histomorphology and the tested molecules were found between the two control groups.. The current findings suggest that a lower level of functional loading by providing small-size diet could reduce TMJ degradation induced by the biomechanical stimulation from abnormal occlusion.

Topics: ADAM Proteins; ADAMTS5 Protein; Aggrecans; Animals; Body Weight; Cartilage, Articular; Collagen Type II; Dental Prosthesis; Diet; Disease Progression; Female; Malocclusion; Mandibular Condyle; Mastication; Mice; Mice, Inbred C57BL; Osteoarthritis; Osteoclasts; Osteoprotegerin; RANK Ligand; Temporomandibular Joint Disorders

There is only limited data concerning the effect of the newer antiepileptic drugs on bone. The objective of this study was to determine the effect of topiramate (TPM) and lamotrigine (LTG) monotherapy on bone mineral density (BMD), mineral content (BMC), bone markers, body composition and bone mechanical strength in the orchidectomized (ORX) rat model. 24 orchidectomized Wistar rats were divided into control and test groups, 8 rats in each group. The control rats received standard laboratory diet (SLD) while rats in the test group were fed with SLD enriched with LTG or TPM for 12 weeks. Dual energy X-ray absorptiometry was used to measure bone mineral density. The concentrations of bone metabolism markers were assayed in bone homogenate. In addition, both femurs were measured and used for biomechanical testing. Compared to the control group, both test groups had significantly lower weight, fat mass, whole body and femur BMD, BMC and reduced mechanical strength of bone. All of these changes were more pronounced in rats exposed to LTG. In conclusion, both LTG and TPM significantly reduce BMD and body weight and impair mechanical strength of bone. A question arises as to the degree of dependence of the effect on the dose. Further studies are warranted to establish whether LTG and TPM may have a clinically significant effect on BMD exclusively in the model of gonadectomized rats, or whether the effect applies also in the model of gonadally intact animals, and in the respective human models.

Topics: Absorptiometry, Photon; Administration, Oral; Alkaline Phosphatase; Animals; Anticonvulsants; Biomechanical Phenomena; Body Composition; Body Weight; Bone and Bones; Bone Density; Bone Morphogenetic Protein 2; Collagen Type I; Enzyme-Linked Immunosorbent Assay; Fructose; Lamotrigine; Male; Models, Animal; Orchiectomy; Osteoprotegerin; Peptide Fragments; Peptides; Procollagen; Rats; Rats, Wistar; Statistics, Nonparametric; Topiramate; Triazines

The gut microbiota (GM) modulates the hosts metabolism and immune system. Probiotic bacteria are defined as live microorganisms which when administered in adequate amounts confer a health benefit on the host and can alter the composition of the GM. Germ-free mice have increased bone mass associated with reduced bone resorption indicating that the GM also regulates bone mass. Ovariectomy (ovx) results in bone loss associated with altered immune status. The purpose of this study was to determine if probiotic treatment protects mice from ovx-induced bone loss. Mice were treated with either a single Lactobacillus (L) strain, L. paracasei DSM13434 (L. para) or a mixture of three strains, L. paracasei DSM13434, L. plantarum DSM 15312 and DSM 15313 (L. mix) given in the drinking water during 6 weeks, starting two weeks before ovx. Both the L. para and the L. mix treatment protected mice from ovx-induced cortical bone loss and bone resorption. Cortical bone mineral content was higher in both L. para and L. mix treated ovx mice compared to vehicle (veh) treated ovx mice. Serum levels of the resorption marker C-terminal telopeptides and the urinary fractional excretion of calcium were increased by ovx in the veh treated but not in the L. para or the L. mix treated mice. Probiotic treatment reduced the expression of the two inflammatory cytokines, TNFα and IL-1β, and increased the expression of OPG, a potent inhibitor of osteoclastogenesis, in cortical bone of ovx mice. In addition, ovx decreased the frequency of regulatory T cells in bone marrow of veh treated but not probiotic treated mice. In conclusion, treatment with L. para or the L. mix prevents ovx-induced cortical bone loss. Our findings indicate that these probiotic treatments alter the immune status in bone resulting in attenuated bone resorption in ovx mice.

Topics: Animals; Biomarkers; Body Weight; Bone and Bones; Bone Marrow; Bone Resorption; Cytokines; Female; Inflammation Mediators; Lactobacillus; Mice; Mice, Inbred C57BL; Minerals; Organ Size; Osteoprotegerin; Ovariectomy; Probiotics; RANK Ligand; T-Lymphocytes, Regulatory; Transforming Growth Factor beta

To investigate diet and nutrition-related factors associated with bone loss in a group of postmenopausal (PM) women. Nutritional intake, inflammatory markers and body composition (weight, body mass index, fat/lean mass) were analysed for associations with bone mineral density (BMD).. A cross sectional study examining correlations between BMD (Duel-energy X ray absorptiometry; (DXA) and dietary intake (3-day diaries), body composition and plasma bone and inflammatory markers: C-terminal telopeptide of type I collagen (CTX) and procollagen type I N propeptide (P1NP), C- reactive protein (CRP), interleukin 6 and 10 (IL-6, IL-10), tumour necrosis factor (TNF) and osteoprotegerin (OPG).. Community dwelling women from the Auckland, Hawke's Bay and Manawatu regions in New Zealand.. 142 healthy, PM women aged 50-70 years.. OPG (per kilogram fat mass) was increased in women with osteoporosis (p<0.001) compared to groups classified with normal BMD and osteopenia. Protein, vitamin B12, zinc, potassium and dairy intake were all positively correlated with higher BMD while dairy and potassium intakes also inversely correlated with CTX. Body composition (weight, BMI and fat/lean mass) had strong positive associations with BMD. Multiple regression analysis showed body weight, potassium and dairy intake were predictors of increased BMD in PM women and explained 39% (r2=0.39, p< 0.003) of variance.. BMD was negatively correlated with OPG and positively with weight, dairy and potassium intake. This study highlights the importance of maintaining adequate body weight and emphasising dairy and potassium predominantly sourced from fruit/vegetables to reduce bone loss at midlife.

Topics: Absorptiometry, Photon; Aged; Body Composition; Body Mass Index; Body Weight; Bone Density; Bone Diseases, Metabolic; C-Reactive Protein; Collagen Type I; Cross-Sectional Studies; Cytokines; Dairy Products; Diet; Dietary Proteins; Female; Health; Humans; Inflammation; Interleukin-10; Interleukin-6; Middle Aged; New Zealand; Osteoporosis, Postmenopausal; Osteoprotegerin; Peptide Fragments; Peptides; Postmenopause; Potassium; Procollagen; Tumor Necrosis Factor-alpha; Vitamin B 12; Zinc

The aim of this study was to evaluate effect of diosgenin (DG) on rats that had osteoporosis-like features induced by ovariectomy (OVX). Seventy-two six-month-old female Wistar rats were subjected to either ovariectomy (n = 60) or Sham operation (SHAM group, n = 12). Beginning at one week post-ovariectomy, the OVX rats were treated with vehicle (OVX group, n = 12), estradiol valerate (EV group, n = 12), or DG at three doses (DG-L, -M, -H group, n = 12, respectively). After a 12-week treatment, administration of EV or DG-H inhibited OVX-induced weight gain, and administration of EV or DG-H or DG-M had a significantly uterotrophic effect. Bone mineral density (BMD) and indices of bone histomorphometry of tibia were measured. Levels of protein and mRNA expression of osteoprotegerin (OPG) and receptor activator of nuclear factor kappa-B ligand (RANKL) in tibia were evaluated by immunohistochemistry and in situ hybridization. Our results show that DG at a high dose (DG-H) had a significant anti-osteoporotic effect compared to OVX control. DG-H treatment down-regulated expression of RANKL and up-regulated expression of OPG significantly in tibia from OVX rats compared to control, and thus lowered the RANKL/OPG ratio. This suggests that the anti-osteoporotic effect of DG might be associated with modulating the RANKL/OPG ratio and DG had potential to be developed as alternative therapeutic agents of osteoporosis induced by postmenopause.

Topics: Animals; Body Weight; Bone Density; Bone Density Conservation Agents; Diosgenin; Disease Models, Animal; Dose-Response Relationship, Drug; Drug Evaluation, Preclinical; Estradiol; Estrogen Replacement Therapy; Female; Gene Expression Regulation; Humans; Organ Size; Osteoporosis, Postmenopausal; Osteoprotegerin; Ovariectomy; RANK Ligand; Rats; Rats, Wistar; Tibia; Uterus

The aim of this study was to investigate bone tissue and plasma levels of RANKL and OPG in rats with prednisolone-induced osteoporosis and to evaluate the outcomes of physical activity on the skeletal system by treadmill and vibration platform training. Osteoporosis is a disease characterised by low bone mass and structural deterioration of bone tissue leading to bone fragility. Vibration exercise is a new and effective measure to prevent muscular atrophy and osteoporosis. The animals were divided into 5 groups. 1: control rats; 2: rats with osteoporosis receiving prednisolone; 3: rats receiving prednisolone and treadmill training; 4: rats receiving prednisolone and vibration stimulation training; 5: rats receiving prednisolone, treadmill and vibration stimulation training. For bone evaluations we used whole-body scans, histology and histomorphometric analysis. RANKL and OPG expression was evaluated by immunohistochemistry and biochemical analysis. After treatment, our data demonstrated that RANKL expression was significantly increased in groups 2 and 3 and decreased in groups 4 and 5. Conversely, OPG expression was significantly decreased in groups 2 and 3 and increased in groups 4 and 5. In conclusion, our findings suggest that mechanical stimulation inhibits the activity of RANKL. This finding provides new insights into the occurrence and progression of osteoporosis.

Topics: Animals; Body Composition; Body Weight; Bone Density; Disease Progression; Down-Regulation; Glucocorticoids; Immunohistochemistry; Male; Osteoporosis; Osteoprotegerin; Physical Conditioning, Animal; RANK Ligand; Rats; Rats, Wistar; Vibration

Parathyroid hormone (PTH) is the only Food and Drug Administration-approved anabolic agent to treat osteoporosis; however, the cellular targets of PTH action in bone remain controversial. PTH modulates bone turnover by binding to the PTH/PTH-related peptide (PTHrP) type 1 receptor (PPR), a G-protein-coupled receptor highly expressed in bone and kidneys. Osteocytes, the most abundant cells in adult bone, also express PPR. However, the physiological relevance of PPR signaling in osteocytes remains to be elucidated. Toward this goal, we generated mice with PPR deletion in osteocytes (Ocy-PPRKO). Skeletal analysis of these mice revealed a significant increase in bone mineral density and trabecular and cortical bone parameters. Osteoblast activities were reduced in these animals, as demonstrated by decreased collagen type I α1 mRNA and receptor activator of NF-κB ligand (RANKL) expression. Importantly, when subjected to an anabolic or catabolic PTH regimen, Ocy-PPRKO animals demonstrated blunted skeletal responses. PTH failed to suppress SOST/Sclerostin or induce RANKL expression in Ocy-PPRKO animals compared with controls. In vitro, osteoclastogenesis was significantly impaired in Ocy-PPRKO upon PTH administration, indicating that osteocytes control osteoclast formation through a PPR-mediated mechanism. Taken together, these data indicate that PPR signaling in osteocytes is required for bone remodeling, and receptor signaling in osteocytes is needed for anabolic and catabolic skeletal responses.

Topics: Animals; Body Weight; Bone and Bones; Bone Density; Cells, Cultured; Collagen Type I; Collagen Type I, alpha 1 Chain; Female; Gene Expression; Immunohistochemistry; In Situ Hybridization; Male; Mice; Osteoclasts; Osteocytes; Osteoprotegerin; Parathyroid Hormone; RANK Ligand; Receptor, Parathyroid Hormone, Type 1; Reverse Transcriptase Polymerase Chain Reaction; Signal Transduction

Low level of cadmium (Cd) exposure may enhance osteoclasts formation in vitro. The aim of the study was to observe the effects of Cd on osteoclasts formation in vivo. Sprague-Dawley male rats were divided into 4 groups which were given Cd via drinking water at concentrations of 0, 2, 10 and 50 mg/L for 12 weeks. At the 12th week, urine samples were collected from all of the rats. All rats were then sacrificed and the blood was collected for biomarkers assay. Bone tissues were dissected for mineral density determinations, histological investigation, tartrate resistant acid phosphatase staining and immunohistochemical staining. The bone mineral density and bone microstructure index of rats treated with 50mg Cd/L were obviously lower than in control rats. Histochemical investigation showed that Cd could induce osteoclasts formation in a dose-dependent manner. Tartrate resistant acid phosphatase 5b levels in rats treated with Cd were higher than the control. Immunohistochemical investigation showed that Cd could enhance receptor-activated nuclear factor kappa B ligand expression (RANKL) and inhibit osteoprotegerin (OPG) expression. Our study evidences in vivo that excessive bone resorption mediated via osteoclasts is an important way for Cd toxic effects on bone and OPG/RANKL may play an important role.

Topics: Acid Phosphatase; Animals; Biomarkers; Body Weight; Bone and Bones; Bone Resorption; Cadmium; Dose-Response Relationship, Drug; Gene Expression; Isoenzymes; Male; NF-kappa B; Osteoclasts; Osteoprotegerin; RANK Ligand; Rats; Rats, Sprague-Dawley; Tartrate-Resistant Acid Phosphatase

The purpose of this study is to investigate the anti-osteoporotic effects of Radix Dipsaci total saponins (RTS). We showed that RTS was able to improve bone properties by either an increase of osteoblastic activity or a decrease in osteoclastic activity.. Radix Dipsaci has long been used as an anti-osteoporotic drug. The present study investigates the anti-osteoporotic effects of RTS.. Three-month-old female rats were randomly assigned into a sham-operated group (sham) and five ovariectomy (OVX) subgroups, namely, OVX with vehicle (OVX), OVX with 17β-ethinylestradiol (E(2)), and OVX with graded doses of RTS (50, 100, or 200 mg/kg/d). RTS and E(2) were administered orally, daily from 1 week after OVX treatment for 4 months. Bone mass, turnover, and strength were evaluated by dual-energy X-ray absorptiometry, biochemical markers, and the three-point bending test. The trabecular bone microarchitecture was assessed by microCT. In vitro experiments were performed to determine the potential molecular mechanisms of the anti-osteoporotic effect of RTS.. RTS prevented the loss of bone mass induced by OVX. The preventive effect on bone loss was primarily indicated by decreasing levels of bone turnover markers and confirmed by the changes in urinary calcium and phosphorus excretion. The treatment also enhanced the biomechanical strength of bone and prevented the deterioration of trabecular bone microarchitecture. RTS induced MC3T3-E1 and primary osteoblastic cell maturation and differentiation and increased bone formation by increasing BMP-2 synthesis. In addition, RTS inhibited osteoclastogenesis through an increase in osteoprotegrin and a decrease in NF-kB ligand expression in vitro.. RTS treatment can effectively suppress the loss of bone mass induced by OVX and in vitro evidence suggests this could be through actions on both osteoblasts and osteoclasts.

Topics: Animals; Body Weight; Bone Density; Bone Density Conservation Agents; Bone Morphogenetic Protein 2; Calcium; Cell Differentiation; Cell Proliferation; Cells, Cultured; Dipsacaceae; Dose-Response Relationship, Drug; Drug Evaluation, Preclinical; Drugs, Chinese Herbal; Estradiol; Female; Femur; Organ Size; Osteoblasts; Osteoclasts; Osteoporosis; Osteoprotegerin; Ovariectomy; Phosphorus; RANK Ligand; Rats; Rats, Sprague-Dawley; Saponins; Tomography, X-Ray Computed; Up-Regulation; Uterus

Although serum osteoprotegerin (OPG) is significantly increased in diabetic subjects, its potential role in beta cell dysfunction has not been investigated. This study aimed to assess the effect of full-length OPG administered in vivo in mice on pancreatic islet structure and function and its interaction with the renin-angiotensin system (RAS). OPG-treated mice showed increased islet monocyte/macrophage infiltration, fibrosis and apoptosis with reduction of islet function. The remodeling of islet architecture was associated with increased pancreatic expression of components of the RAS, growth factor genes (transforming growth factor β and connective tissue growth factor) and inflammatory molecules (monocyte chemotactic protein-1 and vascular adhesion molecule type 1). Prevention of these changes with improvement of insulin secretion was observed in ramipril treated animals. Our data suggest that OPG might play an important role in promoting beta cell dysfunction and that the upregulation of the local RAS represents one possible mechanism responsible for the OPG-induced beta cell dysfunction.

Topics: Animals; Apoptosis; Blood Glucose; Blood Pressure; Body Weight; Cell Lineage; Cell Movement; Chemokine CCL2; Connective Tissue Growth Factor; Fibrosis; Gene Expression Regulation; Humans; Insulin; Islets of Langerhans; Macrophages; Mice; Monocytes; Organ Size; Osteoprotegerin; Peptidyl-Dipeptidase A; Receptor, Angiotensin, Type 1; Systole; Transforming Growth Factor beta; Vascular Cell Adhesion Molecule-1

Clinical evidence indicates that fat is inversely proportional to bone mass in elderly obese women. However, it remains unclear whether obesity accelerates bone loss. In this report we present evidence that increased visceral fat leads to inflammation and subsequent bone loss in 12-month-old C57BL/6J mice that were fed 10% corn oil (CO)-based diet and a control lab chow (LC) for 6 months. As expected from our previous work, CO-fed mice demonstrated increased visceral fat and enhanced total body fat mass compared to LC. The adipocyte-specific PPARγ and bone marrow (BM) adiposity were increased in CO-fed mice. In correlation with those modifications, inflammatory cytokines (IL-1β, IL-6, TNF-α) were significantly elevated in CO-fed mice compared to LC-fed mice. This inflammatory BM microenvironment resulted in increased superoxide production in osteoclasts and undifferentiated BM cells. In CO-fed mice, the increased number of osteoclasts per trabecular bone length and the increased osteoclastogenesis assessed ex-vivo suggest that CO diet induces bone resorption. Additionally, the up-regulation of osteoclast-specific cathepsin k and RANKL expression and down-regulation of osteoblast-specific RUNX2/Cbfa1 supports this bone resorption in CO-fed mice. Also, CO-fed mice exhibited lower trabecular bone volume in the distal femoral metaphysis and had reduced OPG expression. Collectively, our results suggest that increased bone resorption in mice fed a CO-enriched diet is possibly due to increased inflammation mediated by the accumulation of adipocytes in the BM microenvironment. This inflammation may consequently increase osteoclastogenesis, while reducing osteoblast development in CO-fed mice.

Topics: Adipocytes; Animals; Biomarkers; Body Weight; Bone Marrow; Bone Resorption; Corn Oil; Dietary Fats; Female; Femur; Gene Expression; Intra-Abdominal Fat; Mice; Mice, Inbred C57BL; Obesity; Osteoblasts; Osteoclasts; Osteoprotegerin; PPAR gamma; RANK Ligand

Osteoblasts play a critical role in the maintenance of bone mass through bone formation and regulation of bone resorption. Targeted expression of a constitutively active engineered G(i)-coupled G protein-coupled receptor (GPCR) to osteoblasts in vivo leads to severe osteopenia. However, little is known about the role of endogenous receptor-mediated G(i) signaling in regulating osteoblast function. In this study, we investigated the skeletal effects of blocking G(i)-coupled signaling in osteoblasts in vivo. This was accomplished by transgenic expression of the catalytic subunit of pertussis toxin (PTX) under control of the collagen Iα 2.3-kb promoter. These mice, designated Col1(2.3)(+)/PTX(+), showed increased cortical thickness at the femoral midshaft at 12 weeks of age. This correlated with increased periosteal bone formation associated with expanded mineralizing surface observed in 8-week-old mice of both genders. The cancellous bone phenotype of the Col1(2.3)(+)/PTX(+) mice was sexually dimorphic, with increases in fractional bone volume at the distal femur seen only in females. Similarly, while cancellous bone-formation rates were unchanged in males, they could not be quantified for female Col1(2.3)(+)/PTX(+) mice owing to the disorganized nature of the labeling pattern, which was consistent with rapid formation of woven bone. Alterations in osteoclast activity did not appear to participate in the phenotype. These data demonstrate that G(i)-coupled signaling by GPCRs endogenous to osteoblasts plays a complex role in the regulation of bone formation in a manner that is dependent on both gender and the anatomic site within bone.

Topics: Age Factors; Animals; Body Weight; Calcification, Physiologic; Collagen Type I; Cyclins; Doxycycline; Female; Femur; Fibrillar Collagens; Gene Expression; GTP-Binding Protein alpha Subunits, Gi-Go; Male; Mice; Mice, Inbred Strains; Mice, Transgenic; Osteoblasts; Osteoclasts; Osteogenesis; Osteoprotegerin; Peptide Fragments; Peptides; Pertussis Toxin; Procollagen; Promoter Regions, Genetic; RANK Ligand; Receptors, Calcitonin; Receptors, G-Protein-Coupled; Sex Characteristics; Signal Transduction; X-Ray Microtomography

Both physical activity and body mass affect bone properties. In this study we examined how diet-induced obesity combined with voluntary physical activity affects bone properties. Forty 7-week-old male C57BL/6J mice were assigned to four groups evenly: control diet (C), control diet + running (CR), high-fat diet (HF, 60% energy from fat), and high-fat diet + running (HFR). After 21-week intervention, all mice were killed and the left femur was dissected for pQCT and mechanical measurements. Body mass increased 80% in HF and 62% in HFR, with increased epididymal fat pad weight and impaired insulin sensitivity. Except for total and trabecular volumetric bone mineral density (BMD), bone traits correlated positively with body mass, fat pad, leptin, and osteoprotegerin. Obesity induced by a high-fat diet resulted in increased femoral bone cross-sectional area, mineral content (BMC), polar moment of inertia, and mechanical parameters. Of the mice accessing the running wheel, those fed the control diet had thinner cortex and less total metaphyseal BMC and BMD, with enlarged metaphyseal marrow cavity, whereas mice fed the high-fat diet had significantly higher trabecular BMD and smaller marrow cavity. However, the runners had a weaker femoral neck as indicated by decreased maximum flexure load. These results suggest that voluntary running exercise affects bone properties in a site-specific manner and that there is a complex interaction between physical activity and obesity. Thus, both diet and exercise should be considered when optimizing the effects on body composition and bone, even though the underlying mechanisms remain partly unknown.

Topics: Adipose Tissue; Animals; Biomarkers; Body Weight; Bone Density; Dietary Fats; Femur; Glucose Tolerance Test; Insulin; Insulin Secretion; Leptin; Male; Mice; Mice, Inbred C57BL; Motor Activity; Obesity; Osteocalcin; Osteoprotegerin; Physical Conditioning, Animal; Stress, Mechanical; Tomography, X-Ray Computed

Cyclooxygenase-2 (COX-2) knockout (KO) mice in inbred strains can have renal dysfunction with secondary hyperparathyroidism (HPTH), making direct effects of COX-2 KO on bone difficult to assess. COX-2 KO mice in an outbred CD-1 background did not have renal dysfunction but still had two-fold elevated PTH compared to wild type (WT) mice. Compared to WT mice, KO mice had increased serum markers of bone turnover, decreased femoral bone mineral density (BMD) and cortical bone thickness, but no differences in trabecular bone volume by microCT or dynamic histomorphometry. Because PTH is a potent inducer of COX-2 and prostaglandin (PG) production, we examined the effects of COX-2 KO on bone responses after 3 weeks of intermittent PTH. Intermittent PTH increased femoral BMD and cortical bone area more in KO mice than in WT mice and increased trabecular bone volume in the distal femur in both WT and KO mice. Although not statistically significant, PTH-stimulated increases in trabecular bone tended to be greater in KO mice than in WT mice. PTH increased serum markers of bone formation and resorption more in KO than in WT mice but increased the ratio of osteoblastic surface-to-osteoclastic surface only in KO mice. PTH also increased femoral mineral apposition rates and bone formation rates in KO mice more than in WT mice. Acute mRNA responses to PTH of genes that might mediate some anabolic and catabolic effects of PTH tended to be greater in KO than WT mice. We conclude that (1) the basal bone phenotype in male COX-2 KO mice might reflect HPTH, COX-2 deficiency or both, and (2) increased responses to intermittent PTH in COX-2 KO mice, despite the presence of chronic HPTH, suggest that absence of COX-2 increased sensitivity to PTH. It is possible that manipulation of endogenous PGs could have important clinical implications for anabolic therapy with PTH.

Topics: Animals; Biomarkers; Body Weight; Bone Remodeling; Cyclooxygenase 1; Cyclooxygenase 2; Femur; Gene Expression Profiling; Gene Expression Regulation; Health; Humans; Hyperthyroidism; Kidney; Male; Membrane Proteins; Mice; Mice, Knockout; Organ Size; Osteoprotegerin; Parathyroid Hormone; Phenotype; Radiography; RANK Ligand; RNA, Messenger

Sustained parathyroid hormone (PTH) elevation stimulates bone remodeling (ie, both resorption and formation). The former results from increased RANKL synthesis, but the cause of the latter has not been established. Current hypotheses include release of osteoblastogenic factors from osteoclasts or from the bone matrix during resorption, modulation of the production and activity of osteoblastogenic factors from cells of the osteoblast lineage, and increased angiogenesis. To dissect the contribution of these mechanisms, 6-month-old Swiss-Webster mice were infused for 5 days with 470 ng/h PTH(1-84) or 525 ng/h soluble RANKL (sRANKL). Both agents increased osteoclasts and osteoblasts in vertebral cancellous bone, but the ratio of osteoblasts to osteoclasts and the increase in bone formation was greater in PTH-treated mice. Cancellous bone mass was maintained in mice receiving PTH but lost in mice receiving sRANKL, indicating that maintenance of balanced remodeling requires osteoblastogenic effects beyond those mediated by osteoclasts. Consistent with this contention, PTH, but not sRANKL, decreased the level of the Wnt antagonist sclerostin and increased the expression of the Wnt target genes Nkd2, Wisp1, and Twist1. Furthermore, PTH, but not sRANKL, increased the number of blood vessels in the bone marrow. Weekly injections of the RANKL antagonist osteoprotegerin at 10 µg/g for 2 weeks prior to PTH infusion eliminated osteoclasts and osteoblasts and prevented the PTH-induced increase in osteoclasts, osteoblasts, and blood vessels. These results indicate that PTH stimulates osteoclast-dependent as well as osteoclast-independent (Wnt signaling) pro-osteoblastogenic pathways, both of which are required for balanced focal bone remodeling in cancellous bone.

Topics: Adaptor Proteins, Signal Transducing; Animals; Body Weight; Bone Marrow; Bone Morphogenetic Proteins; Bone Remodeling; Calcium; Cell Count; Female; Genetic Markers; Glycoproteins; Humans; Intercellular Signaling Peptides and Proteins; Mice; Models, Biological; Osteoblasts; Osteoclasts; Osteoprotegerin; Parathyroid Hormone; RANK Ligand; Signal Transduction; Solubility; Spine; Wnt Proteins

Vitamin E is found to reverse the effects of nicotine on bone and this study aimed to determine its mechanism. Male Sprague Dawley rats were divided into four groups and treated for 3 months: Group 1 was the control group (RC). Groups 2 (N), 3 (N+TT) and 4 (N+ATF) received nicotine 7 mg/kg throughout the treatment period. In addition, groups 3 and 4 received tocotrienol 60 mg/kg and alpha-tocopherol 60 mg/kg respectively during months 2 and 3. Parameters measured were serum osteoprotegerin (OPG), serum receptor activator of nuclear factor kappa B ligand (RANKL), femoral and lumbar bone calcium content and body weight. Nicotine did not affect OPG or RANKL levels but reduced bone calcium content suggesting the calcium loss is not due to increase osteoclastogenesis. OPG was increased in N+ATF while RANKL was slightly increased in N+TT. Both vitamin E supplements restored bone calcium loss induced by nicotine. Nicotine impaired weight gain in all treatment groups starting week 4 however, N+TT group was comparable to RC from week 6 onwards. Bone protective effects of ATF, but not TT, may be partly due to inhibition of osteoclastogenesis.

Topics: Animals; Body Weight; Bone and Bones; Calcium; Male; Nicotine; Osteoprotegerin; RANK Ligand; Rats; Rats, Sprague-Dawley; Vitamin E

Several animal studies have showed that gelatin may be effective for minimizing bone loss in OVX rats with established osteopenia. To gain insight into how cod bone gelatin administration affects bone loss after ovariectomy, studies were carried out focusing on bone quality and the molecular mechanisms. Eighty-four female rats were ovariectomized, 12 sham-operated, divided into six groups of 12 each and treated one week after ovariectomy either with vehicle or cod bone gelatin (0.375, 0.75, 1.5, 3, 6 mg/kg body weight) for 90 days. Bone densitometry, microCT analysis, real-time PCR analysis and biochemical analysis were used at the end of the study. After 90 days, BMD of proximal tibia and femoral neck decreased in OVX rats, whereas the loss of BMD in those regions was prevented at 3 g/kg (P<0.05). However, the BMD of midshaft femurs showed no significant differences. BV/TV, Tb.N. and Tb.Th. in the 3 g/kg group were, respectively, 30.4% (P<0.05), 145.5% (P<0.05) and 81.5% (P<0.05) higher than in the OVX group. A significant decrease was detected in urine CTX, NTX and DPD, suggesting decreased bone resorption. Treatment with 3 g/kg and 6 g/kg cod bone gelatin attenuated the increase in serum IL-1beta, IL-6 and TNF-alpha observed in the OVX group. Real-time PCR showed significantly decreased levels of mRNA expression for RANKL at the dosage of 6 g/kg and the RANKL/OPG mRNA ratio in the 3 g/kg and 6 g/kg group significantly decreased compared to the OVX group (P<0.05). In conclusion, our data confirmed that the cod bone gelatin treatment at 3 g/kg is effective in the prevention of estrogen deficient bone loss by modulating the expression of RANKL and OPG and suppressing the release of proinflammatory cytokines.

Topics: Alkaline Phosphatase; Amino Acids; Animals; Body Weight; Bone and Bones; Bone Density; Collagen Type I; Estradiol; Female; Femur; Gelatin; Gene Expression; Interleukin-1beta; Interleukin-6; Organ Size; Osteocalcin; Osteoprotegerin; Ovariectomy; Polymerase Chain Reaction; RANK Ligand; Rats; Rats, Sprague-Dawley; Tibia; Tumor Necrosis Factor-alpha; Uterus

Osteoprotegerin (OPG) is a soluble tumour necrosis factor-receptor-like molecule present in connective tissues, especially bone and vasculature. It is known to accumulate in the arterial wall in diabetes. As its synthesis in vascular cells is decreased by insulin, we wanted to elucidate the acute effects of insulin on plasma OPG concentrations in individuals with type 2 diabetes and obese individuals compared with lean controls.. The study population consisted of ten type 2 diabetic, ten obese subjects, and ten lean subjects with no family history of diabetes.. All subjects underwent a 4-h euglycemic-hyperinsulinemic clamp. Plasma OPG, insulin, lactate, HbA1c, cholesterol, triglycerides, free fatty acids (FFA), and glucose disposal rate were measured before and at the end of the clamp.. Baseline OPG concentrations did not differ significantly between groups. Insulin infusion decreased plasma OPG concentrations in all groups (P<0.01); however, the fall in OPG was 50% less in obese and type 2 diabetic individuals (P=0.007). Baseline OPG correlated with fasting insulin, baseline lactate, and low density lipoprotein cholesterol in the diabetic group, and with baseline FFA in the lean group. The relative change of OPG in response to insulin correlated inversely with HbA1c and baseline FFA in the lean group.. Acute hyperinsulinemia decreases plasma OPG, but with diminished effect in individuals with type 2 diabetes and obesity. Increased levels of OPG in arteries and plasma in diabetes together with the capability of plasma OPG as a cardiovascular risk predictor may be related to the described effects of insulin.

Topics: Acute Disease; Body Weight; Cardiovascular Diseases; Diabetes Mellitus, Type 2; Female; Glucose Clamp Technique; Humans; Hyperinsulinism; Insulin; Male; Middle Aged; Obesity; Osteoprotegerin; Risk Factors

The present study was undertaken to investigate the protective effects of ethanol extract from the root of Morinda Officinalis (RMO) on ovariectomy-induced bone loss. Administration of RMO extract increased trabecular bone mineral content and bone mineral density of tibia, improved the levels of phosphorus (P), calcium (Ca) and OPG, decreased the levels of DPD/Cr, TRAP, ACTH and corticosterone, but did not reverse the levels of ALP, TNF-alpha and IL-6 in serum of ovariectomized rats. These findings demonstrated that RMO extract reduced bone loss in ovariectomized rats, probably via the inhibition of bone resorption, but was not involved with bone formation. Anthraquinones and polysaccharides from Morinda officinals could be responsible for their antiosteoporotic activity, and the action mechanism of these constituents needs to be further studied. Therefore, RMO has the potential to develop a clinically useful antiosteoporotic agent.

Topics: Acid Phosphatase; Adrenocorticotropic Hormone; Animals; Body Weight; Bone Density; Calcium; Chromatography, High Pressure Liquid; Corticosterone; Female; Interleukin-6; Isoenzymes; Molecular Structure; Morinda; Organ Size; Osteoprotegerin; Ovariectomy; Phosphorus; Plant Extracts; Rats; Rats, Sprague-Dawley; Tartrate-Resistant Acid Phosphatase; Tumor Necrosis Factor-alpha

We analyzed the relation of osteoprotegerin (OPG) with insulin sensitivity, lipid profile, serum glutamic pyruvic transaminase (SGPT), adipocytokines, and C-reactive protein (CRP) in obese and non-obese subjects.. In the study, 170 subjects (106 obese and 64 non-obese, sex ratio female/male=2.03) were included. Thirty-two obese subjects were reevaluated 6 months after the weight loss induced by bariatric surgery.. OPG did not differ between obese and non-obese subjects (respective mean values 5.17 and 4.96 pmol/l) or according to gender, but was positively correlated with age (P<0.0001 for both groups). OPG was statistically higher in 18 obese diabetic subjects compared with non-diabetics (P=0.03). After adjustment for age, no significant correlation was found between OPG and body mass index (BMI), waist, systolic and diastolic blood pressure, cholesterol, triglycerides, high density lipoprotein (HDL) cholesterol, leptin, and adiponectin in both the obese and non-obese subjects. However, OPG was positively correlated with homeostasis model assessment (HOMA) index and SGPT levels in obese subjects at baseline (r=0.295, r=0.20, P<0.05) and after adjustment for age (r=0.28, r=0.20, P<0.05). OPG was also significantly correlated with CRP; this correlation persisted after adjustment for age in obese subjects (r=0.30, P<0.01). In a multivariate analysis in the obese group, HOMA index and CRP were independent predictors of OPG while SGPT was not. Six months post-surgery, OPG did not change, despite a significant reduction in glucose, SGPT, cholesterol, triglycerides, CRP, and leptin values (P=0.02, P=0.006, P=0.007, P<0.001, P<0.001, P<0.001 respectively) and a significant increase in adiponectin and HDL values (P<0.001 for both variables).. Our results show that in obese subjects, OPG is not related to BMI. However, we describe new relationships between OPG and both HOMA index and CRP.

Topics: Adipokines; Adult; Alanine Transaminase; Bariatric Surgery; Body Weight; C-Reactive Protein; Cardiovascular Diseases; Cholesterol, HDL; Cross-Sectional Studies; Diabetes Mellitus, Type 2; Female; Homeostasis; Humans; Insulin Resistance; Male; Middle Aged; Obesity; Osteoprotegerin; Regression Analysis; Risk Factors; Triglycerides; Weight Loss

We performed this study to clarify whether celecoxib, a selective cyclooxygenase-2 (COX-2) inhibitor, prevents trabecular bone mass reduction by suppressing arthritis-related increase of bone resorption, and to discriminate differences in actions on bone among celecoxib, SC-58560 (a selective COX-1 inhibitor), and indomethacin. Eight-week-old DBA/1J male mice were divided into six groups as follows. Control untreated (Normal) and collagen-induced arthritic (CIA) mice were compared with four treatment groups: celecoxib was orally administered to CIA mice at doses of 0 (Vehicle), 16 (COX2L), and 75 (COX2H) mg/kg, in addition to two groups of mice treated with SC-58560 (COX1) or indomethacin (IND). Histomorphometry showed a significant decrease in tibial trabecular bone volume in arthritic mice, which was corrected by COX2H. The increased osteoclast surface and number in the Vehicle group were suppressed by COX2L, COX2H, and IND. The decreased bone formation rate in Vehicle was elevated by COX2H without statistical significance. A high ratio of mRNA expression of receptor activator of NF-kappaB ligand (RANKL)/osteoprotegerin (OPG) in Vehicle synovial tissue was suppressed by COX2L and COX2H. The increased expression of interleukin (IL)-6 mRNA in Vehicle was suppressed by COX2L, COX2H, and IND, although no difference in this expression was observed in bone marrow cells among all groups. In conclusion, in CIA mice, celecoxib suppresses arthritis-related increase in bone resorption at low and high doses and prevents trabecular bone mass reduction at high doses in association with suppression of osteoclast development in bone marrow through inhibition of RANKL/OPG ratio and IL-6 mRNA expression in inflammatory synovial tissue.

Topics: Amino Acids; Animals; Arthritis, Experimental; Biomarkers; Body Weight; Bone and Bones; Bone Marrow Cells; Cells, Cultured; Cyclooxygenase 2 Inhibitors; Gene Expression Regulation; Interleukin-6; Male; Mice; Organ Size; Osteocalcin; Osteoprotegerin; RANK Ligand; RNA, Messenger; Synovial Membrane; Tibia

Using osteoprotegerin (OPG)-knockout mice, we demonstrated that in vivo the effects of both genistein and 17beta-estradiol (E2) on bone metabolism were completely abolished. In contrast, zoledronic acid could effectively suppress bone resorption and prevent bone loss.. The anti-resorptive effects of E2 on bone metabolism are considered to be mediated via modulation of the osteoblast-derived paracrine factor OPG. Recently, the phytoestrogen genistein was found to suppress bone resorption by enhancing osteoblastic production of OPG. However, the mechanism underlying the in vivo effects of E2 and genistein on bone is not entirely understood, and a central question in this regard is whether E2 regulates bone metabolism via an OPG-dependent pathway.. After mating heterozygous (OPG+/-) mice, homozygous (OPG-/-) and wild-type (WT) with a mixed C57BL/6J x 129/SV background were obtained. The study involved 6-week-old female OPG-/- (n=40) and WT mice (n=8). The OPG-/- mice were randomly divided into 5 groups (n=8 per group) as follows: (1) genistein-treated mice (Gen) that were subcutaneously injected with genistein at a maximal dose (0.8 mg/day); (2) E2-treated mice (E2) that were subcutaneously injected with E2 at a dose (0.03 microg/day); (3) DMSO control mice (DMSO) that were subcutaneously injected with a mixture of dimethylsulfoxide (DMSO) and polyethyleneglycol-300; (4) zoledronic acid-treated mice (Zol) that were subcutaneously injected with zoledronic acid at a dose of (150 microg/kg) twice per week; and (5) H2O control mice that were subcutaneously injected with sterilized water twice per week. The doses of genistein, estrogen and zoledronic acid were selected based on the results of dose-response effect of agents on bone versus uterus in OPG-/- mice. The mice were sacrificed 6 weeks after this intervention. The microarchitecture of the trabecular and cortical bone was assessed by performing microcomputed tomography (micro-CT) for the right proximal tibia. The bone mineral density (BMD) of the left femur was measured by dual-energy X-ray absorptiometry (DXA). The biomechanical parameters of the right femur were determined by a three-point bend testing. Serum levels of bone alkaline phosphatase (B-ALP), tartarate-resistant acid phosphatase-5b (TRACP-5b), and receptor activator of nuclear factor kappaB ligand (RANKL) were determined by performing ELISA.. DXA analysis revealed that the total BMD of the femur was not significantly altered in the Gen, E2, H2O, and DMSO groups. The three-point bending test revealed no significant differences in the biomechanical parameters, including ultimate loading, ultimate stress, stiff index, and elastic modulus, and micro-CT analysis revealed that the microarchitectural parameters of the trabecular bone (vBMD, tBMD, BVF, BSF, SMI, Tb.N, Conn.D, Tb.Sp, and Tb.Th) and cortical bone (Ct.Th, Mm, In.Pm, Ot.Pm, Ma.Ar, Ct.Ar, Tt.Ar, Ct.BMD, and Ct.BMC) did not differ among the groups. Genistein and E2 treatment did not alter the serum TRACP-5b, B-ALP, or RANKL levels. However, in addition to increasing the bone mass, zoledronic acid could effectively improve biomechanical parameters and could completely prevent deterioration of the bone architecture in the OPG-/- mice.. The effects of genistein and E2 on bone metabolism in vivo were lost completely in OPG-deficient mice, suggesting that the effect of these agents on bone metabolism seems to be entirely dependent on OPG. In contrast, zoledronic acid could effectively suppress bone resorption and completely prevent the bone loss in the OPG-/- mice--an effect that is likely to be independent of the OPG pathway.

Topics: Acid Phosphatase; Alkaline Phosphatase; Animals; Biomechanical Phenomena; Body Weight; Bone Density; Bone Remodeling; Bone Resorption; Diphosphonates; Dose-Response Relationship, Drug; Endometrium; Estradiol; Female; Femur; Genistein; Genotype; Imidazoles; Isoenzymes; Liver; Mice; Mice, Inbred C57BL; Mice, Inbred Strains; Mice, Knockout; Osteoprotegerin; RANK Ligand; Tartrate-Resistant Acid Phosphatase; Tibia; Uterus; Zoledronic Acid

Maxillary expansion (ME) is a common practice in orthodontics that aims to increase the constricted maxillary arch width. Relapse often occurs, however, and better treatment strategies are needed. In order to develop a more effective method, this study was designed to further examine the process of tissue remodeling during ME, to identify the changes in expression of several proteins of interest, and to clarify the molecular mechanism responsible for tissue remodeling.. Male Wistar rats were randomly divided into control and ME groups. The rats were euthanized at various intervals over 11 days, and the dissected palates were prepared for histological examination. The structure of the midpalatal sutures changed little during the first three days. Proteins from samples in the ground midpalatal tissues obtained on the third day were subjected to two-dimensional polyacrylamide gel electrophoresis (2-DE) and matrix assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF MS) analysis. Validation of protein expression was performed by Western blot analyses.. From day 5, chondrocytes in the inner layer of suture cartilage and osteoblasts at the end of the suture cartilage began to proliferate, and the skeletal matrix increased later adjacent to the cartilage in the ME group. Comparative proteomic analysis showed increases in 22 protein spots present in the ME group. The changes in three proteins closely related to osteogenesis (parathyroid hormone, osteoprotegerin and vimentin) were confirmed by Western blotting.. Many proteins are over-expressed during ME, and they may play an important role in the remodeling process.

Topics: Animals; Body Weight; Cartilage; Cell Proliferation; Collagen Type I; Male; Neovascularization, Physiologic; Osteoblasts; Osteogenesis; Osteoprotegerin; Palatal Expansion Technique; Palate; Parathyroid Hormone; Proliferating Cell Nuclear Antigen; Random Allocation; Rats; Rats, Wistar; Up-Regulation; Vimentin

Dickkopf-1 (DKK1) has been found to act as a potent Wnt signaling-inhibitory factor for regulating skeletal disorders. We investigated whether modulation of DKK1 expression by end-capped phosphorothioate DKK1 antisense oligonucleotide could alter estrogen loss-induced bone loss. Ovariectomized or sham-operated rats were given 20 microg/kg/day DKK1 sense or antisense oligonucleotide or vehicle for 28 days. Femurs and tibiae were dissected to assess bone mass, biomechanical strength, immunohistochemistry and ex vivo osteoclast formation. We found that DKK1 antisense oligonucleotide significantly abrogated the suppressing effect of ovariectomy on weight, mineral content, mineral density and peak load of femurs. DKK1 antisense oligonucleotide treatment reduced ovariectomy promotion of ex vivo osteoclast differentiation of primary M-CSF-dependent bone marrow macrophages. Histomorphometric observation demonstrated that DKK1 antisense oligonucleotide treatment increased osteoblast number and impaired ovariectomy-promoted trabecular bone loss and osteoclast number in bone tissue. Osteoblastic cells adjacent to endosteum of trabecular bone and chondrocytes at calcified cartilage expressed intensive DKK1 and RANKL and weak OPG immunostaining in ovariectomized rat bone microenvironments. Osteogenic cells and chondral cells displayed weak DKK1, RANKL and OPG expression of bone tissue after DKK1 antisense oligonucleotide treatment. Taken together, attenuation of DKK1 expression in ovariectomized rat bone tissue alleviated loss of bone mass and biomechanical property. The regulatory action of DKK1 antisense oligonucleotide treatment on bone tissue appeared to suppress the promoting effect of estrogen deficiency on osteoclastogenesis-stimulatory factor RANKL expression and osteoclast differentiation. Control of DKK1 signaling can be used in the future as an alternative strategy for protecting estrogen deficiency induction of bone loss.

Topics: Animals; Biomechanical Phenomena; Body Weight; Bone Resorption; Cell Differentiation; Chondrocytes; Estrogens; Female; Femur; Immunohistochemistry; Intercellular Signaling Peptides and Proteins; Oligonucleotides, Antisense; Osteoblasts; Osteoclasts; Osteoprotegerin; Ovariectomy; RANK Ligand; Rats; Rats, Sprague-Dawley; Tibia

We have previously demonstrated that parathyroid hormone-related protein (PTHrP) is a cachexia inducer, but it is still not known what PTHrP effects on target tissues induce the cachexia. Therefore, we examined the effects of anti-PTHrP antibody and osteoprotegerin (OPG) on PTHrP-producing tumor-induced cachexia. Nude mice bearing PTHrP-producing human lung cancer cells (HARA-B) exhibited cachexia with hypercalcemia 3-4 weeks after inoculation, accompanied by losses in body, adipose tissue, and muscle weight. OPG ameliorated hypercalcemia, as did neutralization of PTHrP with antibody; and it increased both body and adipose tissue weights. These increases in body and adipose tissue weight, however, were significantly less than those in mice treated with anti-PTHrP antibody. Simultaneous administration of OPG and anti-PTHrP antibody caused significant increases in body, adipose tissue, and muscle weight, along with an immediate decrease in blood ionized calcium levels. The increase in body weight was similar to that observed in mice treated with anti-PTHrP antibody alone, and the decrease in the blood ionized calcium levels was significantly greater than that in mice treated with OPG or anti-PTHrP antibody alone. These results suggest that an effect of PTHrP on target tissues other than hypercalcemia is involved in the development of cachexia. Expression of cachexia-inducing proinflammatory cytokines (interleukin-6 and leukemia inhibitory factor) is stimulated by PTHrP. This might be a mechanism by which PTHrP produces tumor-induced cachexia. It is also suggested that OPG and anti-PTHrP antibody synergistically act to ameliorate hypercalcemia, although the mechanism responsible for this is unclear.

Topics: Animals; Antibodies, Monoclonal; Body Weight; Cachexia; Disease Models, Animal; Glycoproteins; Humans; Hypercalcemia; Lung Neoplasms; Mice; Mice, Nude; Neoplasm Transplantation; Osteoprotegerin; Parathyroid Hormone-Related Protein; Receptors, Cytoplasmic and Nuclear; Receptors, Tumor Necrosis Factor; Tumor Cells, Cultured

This study examined the mechanisms of osteoclast-mediated bone invasion in a model of oral squamous cell carcinoma (OSCC). C3H/HeN mice were inoculated with SCC VII cells into the masseter region to establish an animal model of mandibular invasion by OSCC.. The mice were divided into three groups: a control group, given daily s.c. injections of saline; group 1, given 2 microg per mouse per day of the bisphosphonate YM529; and group 2, given 10 microg per mouse per day of YM529. After 3 weeks of treatment, the lesions were studied by micro-computed tomography. After tartrate-resistant acid phosphatase (TRAP) staining, the osteoclasts were easily identified, and the percentages of the area occupied by osteoclasts were calculated by computer for each sample. The tumors were analyzed by RT-PCR to determine the mRNA expression of interleukin-6 (IL-6), parathyroid hormone-related protein (PTHrP), tumor necrosis factor-alpha (TNF-alpha), receptor activator of nuclear factor-kappaB (RANK), RANK ligand (RANKL), and osteoprotegerin.. SCC VII cells rapidly multiplied in the masseter muscle of the mice. Bone invasion was evident only in the control group on micro-computed tomography. On TRAP-stained slices, the percentages of osteoclasts in groups 1 and 2 were significantly lower than that in the control group. The mRNA expressions of IL-6, PTHrP, THF-alpha, and RANK decreased as the concentration of YM529 increased.. We conclude that various cancer-derived cytokines play important roles in the invasion of bone by OSCC. YM529, a third-generation bisphosphonate, can suppress osteoclast-mediated bone invasion by OSCC. The mechanism of this effect might involve inhibition of cytokines such as IL-6, PTHrP, TNF-alpha, and RANK by YM529.

Topics: Animals; Body Weight; Bone Resorption; Carcinoma, Squamous Cell; Carrier Proteins; Cell Line, Tumor; Diphosphonates; Disease Models, Animal; Gene Expression; Glycoproteins; Imaging, Three-Dimensional; Imidazoles; Interleukin-6; Male; Mandible; Membrane Glycoproteins; Mice; Mice, Inbred C3H; Mouth Neoplasms; Neoplasm Invasiveness; Osteoclasts; Osteoprotegerin; Parathyroid Hormone-Related Protein; RANK Ligand; Receptor Activator of Nuclear Factor-kappa B; Receptors, Cytoplasmic and Nuclear; Receptors, Tumor Necrosis Factor; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Tomography, X-Ray Computed; Tumor Necrosis Factor-alpha

Osteoprotegerin (OPG) is a naturally occurring protein, which prevents bone resorption by inhibition of osteoclastogenesis, function, and survival. Therefore, recombinant OPG may be an attractive drug in the treatment of chronic bone resorptive diseases such as osteoporosis. Gene therapy has the potential to achieve long-term treatment by delivering genes of anti-resorptive proteins to the recipient. The effects of OPG gene therapy on fracture healing have not been described previously. The influence of OPG gene therapy on callus formation, callus tissue structural strength, apparent material properties, and histology of tibia fractures in rats was investigated after 3 weeks and 8 weeks of healing. Intramuscular administration of adeno-associated virus (AAV) vector-mediated OPG resulted in increased levels of OPG in serum of approximately 100 ng/ml throughout the study period. Control animals with fractures received transduction with an AAV reporter gene construct (AAV-enhanced green fluorescent protein (eGFP)), and in this group serum OPG levels remained at baseline (<10 ng/ml). After 3 weeks of healing, AAV-OPG treatment reduced the number of osteoclasts in the callus tissue (33%, P < 0.001). However, AAV-OPG treatment did not influence callus dimensions, callus bone mineral content (BMC), fracture structural strength, or apparent callus tissue material properties. After 8 weeks of healing, AAV-OPG treatment reduced the number of osteoclasts in the callus tissue (31%, P < 0.001) compared with AAV-eGFP fractures. Furthermore, deposition of new woven bone at the fracture line of the original cortical bone was hampered (new woven bone present: in all AAV-eGFP animals, in 41% of AAV-OPG-treated animals, P < 0.001). AAV-OPG treatment also increased callus BMC (18%, P = 0.023) compared with AAV-eGFP fractures. AAV-OPG did not influence callus dimensions, structural strength of the fractures, or ultimate stress, whereas elastic modulus was reduced in the AAV-OPG groups (37%, P = 0.039). The experiment demonstrates that AAV-OPG gene therapy decreases the fracture remodeling, but this does not influence the structural strength of healing fractures.

Topics: Animals; Biomechanical Phenomena; Body Weight; Bone Remodeling; Bony Callus; Female; Fractures, Bone; Genetic Therapy; Glycoproteins; Humans; Osteoprotegerin; Radiography; Rats; Rats, Wistar; Receptors, Cytoplasmic and Nuclear; Receptors, Tumor Necrosis Factor; Tibia

Herbal Sambucus williamsii HANCE (SWH) is a folk medicine with a long history of safe use for treatment of bone fractures and joint diseases in China. The present study was designed to investigate if SWH extract could be used for treatment of postmenopausal osteoporosis. SWH extracts (30 or 60 mg/100 g body weight/d) were orally administrated to four-months-old ovariectomized (OVX) rats for 3 months. SWH extracts did not alter weight gain and uterus weight in OVX rats. SWH extracts significantly increased serum Ca levels (p<0.05, vs. OVX control group) as well as decreased urinary Ca excretion (p<0.01, vs. OVX control group) in OVX rats. The upregulation of serum alkaline phosphatase, serum osteocalcin as well as urinary deoxypyridinoline levels by OVX was suppressed by treatment with SWH extracts in rats (p<0.05, vs. OVX control group). SWH extract increased the stiffness of femur at both dosage (p<0.05, vs. OVX control group) and increased tibial bone mineral density at 60 mg/100 g body weight/d (p<0.05, vs. OVX control group) in OVX rats. Our results indicate that orally administrated SWH extracts can decrease urinary calcium excretion and bone turnover rate in OVX rats, resulting in positive effects on biomechanical strength of bone and bone mineral density. This study is the first to report that SWH could be considered as a potential candidate for management of postmenopausal osteoporosis. Then in vitro experiments were performed to determine the potential molecular mechanism of the anti-osteoporotic effect of SWH. Results suggested that chloroform fraction and ethyl acetate fraction of SWH can inhibit osteoclastogenesis osteoclast by modulating the expression of osteoprotegrin (OPG) and receptor activator of NF-kappaB ligand (RANKL) mRNA in osteoblastic UMR 106 cells. Both of them increased OPG mRNA and decreased RANKL mRNA expression, resulting in a dose-dependent increase in OPG/RANKL mRNA ratio (p<0.01, vs. vehicle-treated). Taken together, SWH treatment can effectively suppress the OVX-induced increase in bone turnover and its effects might be mediated by a decrease in osteoclastogenesis.

Topics: Animals; Base Sequence; Biomechanical Phenomena; Body Weight; Bone and Bones; Carrier Proteins; Cell Line; DNA Primers; Female; Glycoproteins; Membrane Glycoproteins; Organ Size; Osteoporosis; Osteoprotegerin; Ovariectomy; RANK Ligand; Rats; Rats, Sprague-Dawley; Receptors, Cytoplasmic and Nuclear; Receptors, Tumor Necrosis Factor; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Uterus

Osteoporosis is a disease characterized by low bone mass and structural deterioration of bone tissue, leading to bone fragility and increased susceptibility to fractures. The microgravity of space creates an extreme environment that provides a model for osteoporosis in humans. This greatly accelerated form of osteopenia results in a 0.5-2% loss of bone mass per month. Rat models for this osteoporosis have been examined on many occasions, but STS-108 was the first Space Shuttle flight to use mice. Data reported to date indicate that spaceflight experiments with mice hold promise in predicting some spaceflight effects on humans. Due to the cost and infrequency of flights, ground-based models have been developed to mimic the deleterious effects of the microgravity environment. Hindlimb suspension is one such localized model. This model removes gravitational loading from the hindlimbs by suspending the animal by its tail to a guy wire that runs lengthwise across the cage. Because mice had not flown before STS-108, a direct comparison of this model's ability to predict spaceflight results has not been examined. The objective of this research is to closely repeat the STS-108 profile, with hindlimb suspension replacing spaceflight. This includes examining the ability of the protein osteoprotegerin, an osteoclast-inhibiting therapeutic, to mitigate the deleterious effects of skeletal unloading. It is expected that the results will lead to better understanding of the mechanisms of mineralization and bone remodeling to aid in development of countermeasures to prevent spaceflight induced osteoporosis and aid the treatment of osteoporosis here on earth.

Topics: Animals; Body Weight; Bone Density; Femur; Glycoproteins; Hindlimb; Hindlimb Suspension; Mice; Mice, Inbred C57BL; Muscular Disorders, Atrophic; Osteoprotegerin; Receptors, Cytoplasmic and Nuclear; Receptors, Tumor Necrosis Factor; Space Flight; Weightlessness; Weightlessness Simulation

Osteoblasts regulate the recruitment and activity of osteoclasts through expression of RANKL and osteoprotegerin (OPG). To determine whether expression of RANKL and OPG change with age and how these changes relate to the bone loss of aging, we measured bone mass and cancellous volume, and expression of RANKL, OPG, alkaline phosphatase (AP), osteocalcin (OC), and alpha I collagen (COLL) in whole bone and osteoblast-like cells in culture using 6-week- (young), 6-month- (adult), and 24-month-old (old) mice. Cancellous volume decreased by 20% from young to adult and by 52% from adult to old. RANKL mRNA levels in whole bone were 2.1-fold and 4.4-fold higher in adult and old mice, respectively, compared with young mice, whereas OPG mRNA levels decreased with age slightly. RANKL expression was negatively (r = -0.99) and OPG was positively (r = 0.92) correlated with cancellous bone volume. Expression of RANKL was higher and OPG lower in cells from older animals early in culture (day 7). With cell maturation, RANKL mRNA levels in cells from young and adult mice increased, whereas levels in cells from old animals decreased. By 21 and 28 days of culture, no differences were found in RANKL mRNA in osteoblast-like cells among different age groups. We conclude that expression of RANKL and OPG change with age in whole bone and in cultured osteoblast-like cells. These changes favor increased osteoclast over osteoblast activity, and may explain, in part, the imbalance in bone formation and resorption associated with aging.

Topics: Aging; Animals; Blotting, Western; Body Weight; Bone and Bones; Carrier Proteins; Cells, Cultured; Glycoproteins; Male; Membrane Glycoproteins; Mice; Mice, Inbred C57BL; Osteoblasts; Osteoporosis; Osteoprotegerin; RANK Ligand; Receptor Activator of Nuclear Factor-kappa B; Receptors, Cytoplasmic and Nuclear; Receptors, Tumor Necrosis Factor; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Time Factors

Rat models of immobilization-induced osteopenia are characterized by uncoupling of bone metabolism, i.e., increased bone resorption and decreased bone formation in trabecular bone. Using such a rat model, the efficacy of osteoclastogenesis inhibitory factor (OCIF)/osteoprotegerin, a novel secreted protein that inhibits osteoclastogenesis, in reducing bone loss was investigated. Male Fischer rats were neurectomized and injected intramuscularly with either OCIF (0.2, 1.0, or 5.0 mg/kg body weight) or vehicle once daily for 7 days. On the eighth day after sciatic neurectomy, significant bone loss was observed in the vehicle-injected rats. OCIF ameliorated the decrease in bone mineral density (BMD) of both the proximal and distal femur in a dose-dependent manner. OCIF also ameliorated the decrease in bone strength of the femoral neck at the highest dose. A high correlation (r = 0.805) was detected between the BMD of the distal femur and the bone strength of the femoral neck. When OCIF was administered intermittently to the immobilized rats twice weekly (on days 1 and 4) after immobilization, it also ameliorated the decrease in BMD of the distal femur. These results suggest that OCIF has therapeutic potential for the treatment of immobilization-induced osteopenia.

Topics: Animals; Body Weight; Bone Density; Bone Resorption; Dose-Response Relationship, Drug; Femur; Glycoproteins; Injections, Intramuscular; Male; Osteoprotegerin; Rats; Rats, Inbred F344; Receptors, Cytoplasmic and Nuclear; Receptors, Tumor Necrosis Factor; Restraint, Physical; Sciatic Nerve

Low bone mineral density (BMD) is a major risk factor for the development of osteoporosis and there is strong evidence to suggest that the procurement and preservation of peak BMD is genetically determined. In an effort to identify factors responsible for susceptibility to low BMD in the Irish population, we investigated its possible association with polymorphisms in the Osteoprotegerin (OPG) gene, Type I collagen alpha 1 (COLIA1) Sp1 binding site and vitamin D receptor (VDR) start codon. Following a systematic screening of the regulatory and coding regions of the OPG gene, we identified a novel G1181C polymorphism in exon 1 and a T950C polymorphism in the promoter region of the OPG gene. Participants were recruited from the Bone Densitometry Unit of Cork University Hospital, including 381 postmenopausal women aged 61.26 +/- 8.50 (mean +/- SD) and 130 premenopausal women aged 46.30 +/- 6.50 (mean +/- SD). Following association analysis using both the premenopausal and postmenopausal cohorts we found that postmenopausal women carrying one or more C alleles of the G1181C polymorphism had 14.8% lower BMD (P = 0.05) at the lumbar spine and 14.4% lower BMD (P = 0.04) at the FN. However, both were nonsignificant when the Bonferroni correction factor (0.01 significance level) was applied to correct for multiple hypothesis testing. We found no association between alleles of the T950C OPG polymorphism and BMD. Similarly, we have found a lack of association between the VDR (fok1) polymorphism or COLIA1 Sp1 polymorphism and low BMD in either postmenopausal or premenopausal women in this population.

Topics: Base Sequence; Body Height; Body Weight; Bone Density; Codon, Initiator; Collagen Type I; DNA Primers; Female; Glycoproteins; Humans; Ireland; Mass Screening; Menopause; Middle Aged; Osteoprotegerin; Polymorphism, Genetic; Polymorphism, Single Nucleotide; Postmenopause; Premenopause; Promoter Regions, Genetic; Receptors, Calcitriol; Receptors, Cytoplasmic and Nuclear; Receptors, Tumor Necrosis Factor; Retrospective Studies

A requirement for tooth eruption is the resorption of alveolar bone. Because bone resorption is stimulated by dexamethasone both in vivo and in vitro, dexamethasone 21-phosphate, a soluble form of dexamethasone, was injected into rats to determine its effect on tooth eruption. Such dexamethasone injections accelerate the time of intra-osseous eruption in rat incisors but do not accelerate the eruption time of rat molars when injected into rats. The injections of dexamethasone 21-phosphate also accelerate the time of eyelid opening in the postnatal rats, as well as retarding growth, as measured by body weight. These effects of dexamethasone 21-phosphate parallel the effects of epidermal growth factor injections, including the absence of an effect on molar eruption. This suggests that the molecular signals for the initiation of tooth eruption (i.e., onset of bone resorption) differ between rat incisors and molars. Given that rat incisors are teeth of continuous eruption whereas rat molars are teeth of limited eruption, as are human teeth, care must be taken in extrapolating results derived from rat incisors to human dentition. In vitro, dexamethasone has no effect on the gene expression of either osteoprotegerin or epidermal growth factor in dental follicle cells derived from molars. Because osteoprotegerin expression during normal tooth eruption is transitorily inhibited early postnatally in the molar dental follicle to allow osteoclast formation, the absence of inhibition of its expression by dexamethasone could explain why dexamethasone does not accelerate eruption in molars.

Topics: Animals; Animals, Newborn; Body Weight; Dexamethasone; DNA Primers; Epidermal Growth Factor; Eyelids; Gene Expression; Glucocorticoids; Glycoproteins; Incisor; Injections, Subcutaneous; Molar; Osteoprotegerin; Rats; Rats, Sprague-Dawley; Receptors, Cytoplasmic and Nuclear; Receptors, Tumor Necrosis Factor; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Tooth Eruption