Regulation of bone biology by prostaglandin endoperoxide H synthases (PGHS): A rose by any other name...

It is well established that prostaglandins are essential mediators of bone resorption and formation. In the early 1990s, it was discovered that enzymatic reactions producing prostaglandins were regulated by two cyclooxygenase enzymes, one producing prostaglandins constitutively in tissues like the stomach, prostaglandin endoperoxide H synthase-1 (PGHS-1 or COX-1), and another induced by mitogens or inflammatory mediators (PGHS-2 or COX-2). This neat distinction has not been maintained because both enzymes act in different cell systems to provide physiological signaling, constitutively or by induction under certain conditions. For example, the regulation patterns of PGHS-1 and PGHS-2 are distinct, but the evidence shows that PGHS-2 functions constitutively in the skeleton. PGHS-2 hits quickly been established, therefore, as a key regulator of bone biology, capable of rapid and transient expression in bone cells, and mediating osteoclastogenesis, mechanotransduction, bone formation and fracture repair. The goal of this review is to Summarize the current state of our knowledge of PGHS regulation of bone metabolism and to identify some of the key unresolved challenges and questions that require further study. (c) 2006 Elsevier Ltd. All rights reserved.

Transgenic mice that express Cre recombinase in osteoclasts

To study the physiological control of osteoclasts, the bone resorbing cells, we generated transgenic mice carrying the Cre recombinase gene driven by either the tartrate-resistant acid phosphatase (TRAP) or cathepsin K (Ctsk) promoters. TRAP-Cre and Ctsk-Cre transgenic mouse lines were characterized by breeding with LacZ ROSA 26 (R26R) reporter mice and immunohistochemistry for Cre recombinase. The Cre transgene was functional in all lines, with Cre-mediated recombination occurring primarily in the long bones, vertebrae, ribs, and calvaria. Histological analyses of the bones demonstrated that functional Cre protein was present in 1) osteoclasts (Ctsk-Cre); 2) osteoclasts, columnar proliferating, and hypertrophic chondrocytes (TRAP-Cre line 4); and 3) round proliferating chondrocytes (TRAP-Cre line 3). In conclusion, we generated transgenic mouse lines that will enable the deletion of floxed target genes in osteoclasts, which will be valuable tools for studying the regulation of osteoclast function. (C) 2004 Wiley-Liss, Inc.

Orthodontic tooth movement in the prednisolone-treated rat

Adverse effects of corticosteroids on bone metabolism raise concerns as to whether steroid treatment may influence orthodontic movement. This study examined the effect of prednisolone on orthodontic movement using an established rat model. The corticosteroid treated group (N = 6) was administered prednisolone (1 mg/kg) daily, for a 12-day induction period; the control group (N = 6) received equivalent volumes of saline. On day 12, an orthodontic appliance was placed which exerted 30 g of mesial force to the maxillary first molar. Animals were sacrificed on day 24 and tooth movement was measured. Sagittal sections of the molars were stained with haematoxylin and eosin, and for tartrate-resistant acid phosphatase (TRAP) activity. While there were no significant differences in the magnitude of tooth movement between the 2 groups, steroid-treated rats displayed significantly less root resorption on the compression side and fewer TRAP-positive cells within the PDL space on the same side. This suggests steroid treatment suppressed elastic activity.

The 'muscle-bone unit' during the pubertal growth spurt

Mechanostat theory postulates that developmental changes in bone strength are secondary to the increasing loads imposed by larger muscle forces. Therefore, the increase in muscle strength should precede the increase in bone strength. We tested this prediction using densitometric surrogate measures of muscle force (lean body mass, LBM) and bone strength (bone mineral content, BMC) in a study on 70 boys and 68 girls who were longitudinally examined during pubertal development. On the level of the total body, the peak in LBM accrual preceded the peak in BMC accretion by an average of 0.51 years in girls and by 0.36 years in boys. In the arms, the maximal increase in LBM was followed by arm peak BMC accrual after an interval of 0.71 years in girls and 0.63 years in boys. In the lower extremities, the maximal increase in LBM was followed by peak BMC accrual after an interval of 0.22 years in girls and 0.48 years in boys. A multiple regression model revealed that total body peak LBM velocity, but not peak height velocity and sex, was independently associated with total body peak BMC velocity (r(2) = 0.50; P < 0.001). Similarly, arm and leg peak LBM velocity, but not peak height velocity and sex, were independently associated with arm and leg peak BMC velocity, respectively (r(2) = 0.61 for arms, r(2) = 0.41 for legs; P < 0.001 in both cases). These results are compatible with the view that bone development is driven by muscle development, although the data do not exclude the hypothesis that the two processes are independently determined by genetic mechanisms. (C) 2004 Elsevier Inc. All rights reserved.

Hearing sensitivity and bone mineral density in older adults: the Health, Aging and Body Composition Study

Bone mineral density (BMD) may be associated with hearing loss in older adults. Demineralization of the cochlear capsule has been associated with hearing loss in those with Paget's disease of the bone and otosclerosis. Osteoporosis may also result in cochlear capsule demineralization. We hypothesized that lower hip BMD and lower heel ultrasound measurements would be associated with hearing loss in a population-based sample of 2,089 older black and white men and women. Bone parameters and hearing function were measured at the fourth clinical follow-up visit. Audiometric threshold testing was used to measure air- and bone-conduction hearing sensitivity. BMD of the hip and its subregions was measured using dual-energy X-ray absorptiometry. Calcaneal bone measurements [broadband ultrasound attenuation (BUA), speed of sound (SOS) and the quantitative ultrasound index (QUI)] were obtained using heel ultrasound. After adjusting for known hearing loss risk factors, no association was found between hearing and any of the bone measurements in whites and black women. In black men, however, lower hip BMD was associated with higher odds of hearing loss; for each standard deviation decrease in total hip BMD, the odds of hearing loss were 1.41 (95% confidence interval 1.08, 1.83), 1.39 (95% CI 1.07, 1.82) for femoral neck BMD and 1.65 (95% CI 1.26, 2.16) for trochanter BMD. Conductive hearing loss was associated with lower heel ultrasound measurements, though only among white men. The results of this study are mixed and inconclusive. Lower BMD of the hip and its subregions was associated with hearing loss among black men, but not among whites or black women. Lower measurements on heel ultrasound were associated with conductive hearing loss, though only among white men. These results suggest that axial and appendicular bone parameters may be modestly associated with hearing loss in older men, but not in women.

Hypothalamic control of bone formation: Distinct actions of leptin and Y2 receptor pathways

Leptin and Y2 receptors on hypothalamic NPY neurons mediate leptin effects on energy homeostasis; however, their interaction in modulating osteoblast activity is not established. Here, direct testing of this possibility indicates distinct mechanisms of action for leptin anti-osteogenic and Y2(-/-) anabolic pathways in modulating bone formation. Introduction: Central enhancement of bone formation by hypothalamic neurons is observed in leptin-deficient oblob and Y2 receptor null mice. Similar elevation in central neuropeptide Y (NPY) expression and effects on osteoblast activity in these two models suggest a shared pathway between leptin and Y2 receptors in the central control of bone physiology. The aim of this study was to test whether the leptin and Y2 receptor pathways regulate bone by the same or distinct mechanisms. Materials and Methods: The interaction of concomitant leptin and Y2 receptor deficiency in controlling bone was examined in Y2(-/-) oblob double mutant mice, to determine whether leptin and Y2 receptor deficiency have additive effects. Interaction between leptin excess and Y2 receptor deletion was examined using recombinant adeno-associated viral vector overproduction of NPY (AAV-NPY) to produce weight gain and thus leptin excess in adult Y2(-/-) mice. Cancellous bone volume and bone cell function were assessed. Results: Osteoblast activity was comparably elevated in oblob, Y2(-/-), and Y2(-/-) oblob mice. However, greater bone resorption in oblob and Y2(-/-) oblob mice reduced cancellous bone volume compared with Y2(-/-). Both wildtype and Y2(-/-) AAV-NPY mice exhibited marked elevation of white adipose tissue accumulation and hence leptin expression, thereby reducing osteoblast activity. Despite this anti-osteogenic leptin effect in the obese AAV-NPY model, osteoblast activity in Y2(-/-) AAV-NPY mice remained significantly greater than in wildtype AAV-NPY mice. Conclusions: This study suggests that NPY is not a key regulator of the leptin-dependent osteoblast activity, because both the leptin-deficient stimulation of bone formation and the excess leptin inhibition of bone formation can occur in the presence of high hypothalamic NPY. The Y2(-/-) pathway acts consistently to stimulate bone formation; in contrast, leptin continues to suppress bone formation as circulating levels increase. As a result, they act increasingly in opposition as obesity becomes more marked. Thus, in the absence of leptin, the cancellous bone response to loss of Y2 receptor and leptin activity can not be distinguished. However, as leptin levels increase to physiological levels, distinct signaling pathways are revealed.

Inhibitors of cyclo-oxygenase-2 and secretory phospholipase A2 preserve bone architecture following ovariectomy in adult rats

Epidemiological evidence and in vitro data suggest that COX-2 is a key regulator of accelerated remodeling. Accelerated states of osteoblast and osteoclast activity are regulated by prostaglandins in vitro, but experimental evidence for specific roles of cyclooxygenase-2 (COX-2) and secretory phospholipase A(2) (sPLA(2)) in activated states of remodeling in vivo is lacking. The aim of this study was to determine the effect of specific inhibitors of sPLA(2)-IIa and COX-2 on bone remodeling activated by estrogen deficiency in adult female rats. One hundred and twenty-four adult female Wistar rats were ovariectomized (OVX) or sham-operated. Rats commenced treatment 14 days after surgery with either vehicle, a COX-2 inhibitor (DFU at 0.02 mg/kg/day and 2.0 mg/kg/day) or a sPLA(2)-group-IIa inhibitor (KH064 at 0.4 mg/kg/day and 4.0 mg/kg/day). Treatment continued daily until rats were sacrificed at 70 days or 98 days post-OVX. The right tibiae were harvested, fixed and embedded in methylmethacrylate for structural histomorphometric bone analysis at the proximal tibial metaphysis. The specific COX-2 or sPLA(2) inhibitors prevented ovariectomy-induced (OVX-induced) decreases in trabecular connectivity (P < 0.05); suppressed the acceleration of bone resorption; and maintained bone turnover at SHAM levels following OVX in the rat. The sPLA2 inhibitor significantly suppressed increases in osteoclast surface induced by OVX (P < 0.05), while the effect of COX-2 inhibition was less marked. These findings demonstrate that inhibitors of COX-2 and sPLA(2)-IIa can effectively suppress OVX-induced bone loss in the adult rat by conserving trabecular bone mass and architecture through reduced bone remodeling and decreased resorptive activity. Moreover, we report an important role of sPLA(2)-IIa in osteoclastogenesis that may be independent of the COX-2 metabolic pathway in the OVX rat in vivo. (c) 2006 Elsevier Inc. All rights reserved.

Relationship between bicarbonate retention and bone characteristics in broiler chickens

Determination of the bicarbonate retention factor (BRF) is an important step during development of the indicator amino acid oxidation technique for use in a new model. A series of 4-h oxidation experiments were performed to determine the BRF of broilers aged 7, 14, 21, 28, 35, and 42 d using 4 birds per age group. A priming dose of 1.2 mu Ci of (NaHCO3)-C-14, followed by eight half-hourly doses of 1 mu Ci of (NaHCO3)-C-14 were given orally to each of 4 birds per age. The percentage of 14 C dose expired by the bird at a steady state was measured. These birds, as well as 12 additional birds matched for age and BW, were killed, and femur bone mineral density was measured by quantitative computed tomography to determine the relationship between bone development and bicarbonate retention at each age. There was a correlation (r = 0.50; P < 0.05) between total cross-sectional femur bone mineral density and bicarbonate retention at each age. A prediction equation (Y = 6.95 x 10(-2) X - 3.51 x 10(5)X(2) + 27.58; P < 0.0001, R-2 = 0.79) where Y = bicarbonate retention and X = BW was generated to predict Y as a function of X. Bicarbonate retention values peaked at 28 d, during the stage of the most rapid bone deposition and the highest growth rate. A constant BRF was found from 1,900 to 2,700 g of BW of 35.15 +/- 1.095% (mean SEM). This retention factor will allow the accurate correction of oxidation of C-14-labeled substrates in broilers of different ages and BW in future indicator amino acid oxidation studies.

Volumetric and areal bone mineral density measures are associated with cardiovascular disease in older men and women: The health, aging, and body composition study

The associations of volumetric (vBMD) and areal (aBMD) bone mineral density measures with prevalent cardiovascular disease (CVD) and subclinical peripheral arterial disease (PAD) were investigated in a cohort of older men and women enrolled in the Health, Aging, and Body Composition Study. Participants were 3,075 well-functioning white and black men and women (42% black, 51% women), aged 68-80 years. Total hip, femoral neck, and trochanter aBMD were measured using dual-energy X-ray absorptiometry. Quantitative computed tomography was used to evaluate spine trabecular, integral, and cortical vBMD measures in a subgroup (n = 1,489). Logistic regression was performed to examine associations of BMD measures with CVD and PAD. The prevalence of CVD (defined by coronary heart disease, PAD, cerebrovascular disease, or congestive heart failure) was 29.8%. Among participants without CVD, 10% had subclinical PAD (defined as ankle-arm index < 0.9). Spine vBMD measures were inversely associated with CVD in men (odds ratio of integral [ORintegral] = 1.34, 95% confidence interval [CI] 1.10-1.63; ORtrabecular = 1.25, 95% CI 1.02-1.53; ORcortical = 1.36, 95% CI 1.11-1.65). In women, for each standard deviation decrease in integral vBMD, cortical vBMD, or trochanter aBMD, the odds of CVD were significantly increased by 28%, 27%, and 22%, respectively. Total hip aBMD was associated with subclinical PAD in men (OR = 1.39, 95% CI 1.03-1.84) but not in women. All associations were independent of age and shared risk factors between BMD and CVD and were not influenced by inflammatory cytokines (interleukin-6 and tumor necrosis factors-alpha). In conclusion, our results provide further evidence for an inverse association between BMD and CVD in men and women. Future research should investigate common pathophysiological links for osteoporosis and CVD.

Population pharmacokinetics of itraconazole and its active metabolite hydroxy-itraconazole in paediatric cystic fibrosis and bone marrow transplant patients

Objective: The objective of the study was to characterise the population pharmacokinetic properties of itraconazole and its active metabolite hydroxyitraconazole in a representative paediatric population of cystic fibrosis and bone marrow transplant (BMT) patients and to identify patient characteristics influencing the pharmacokinetics of itraconazole. The ultimate goals were to determine the relative bioavailability between the two oral formulations (capsules vs oral solution) and to optimise dosing regimens in these patients. Methods: All paediatric patients with cystic fibrosis or patients undergoing BMT at The Royal Children's Hospital, Brisbane, QLD, Australia, who were prescribed oral itraconazole for the treatment of allergic bronchopulmonary aspergillosis (cystic fibrosis patients) or for prophylaxis of any fungal infection (BMT patients) were eligible for the study. Blood samples were taken from the recruited patients as per an empirical sampling design either during hospitalisation or during outpatient clinic visits. ltraconazole and hydroxy-itraconazole plasma concentrations were determined by a validated high-performance liquid chromatography assay with fluorometric detection. A nonlinear mixed-effect modelling approach using the NONMEM software to simultaneously describe the pharmacokinetics of itraconazole and its metabolite. Results: A one-compartment model with first-order absorption described the itraconazole data, and the metabolism of the parent drug to hydroxy-itraconazole was described by a first-order rate constant. The metabolite data also showed one-compartment characteristics with linear elimination. For itraconazole the apparent clearance (CLitraconazole) was 35.5 L/hour, the apparent volume of distribution (V-d(itraconazole)) was 672L, the absorption rate constant for the capsule formulation was 0.0901 h(-1) and for the oral solution formulation was 0.96 h-1. The lag time was estimated to be 19.1 minutes and the relative bioavailability between capsules and oral solution (F-rel) was 0.55. For the metabolite, volume of distribution, V-m/(F (.) f(m)), and clearance, CL/(F (.) fm), were 10.6L and 5.28 L/h, respectively. The influence of total bodyweight was significant, added as a covariate on CLitraconazoie/F and V-d(itraconazole)/F (standardised to a 70kg person) using allometric three-quarter power scaling on CLitraconazole/F, which therefore reflected adult values. The unexplained between-subject variability (coefficient of variation %) was 68.7%, 75.8%, 73.4% and 61.1% for CLitraconazoie/F, Vd(itraconazole)/F, CLm/(F (.) fm) and F-rel, respectively. The correlation between random effects of CLitraconazole and Vd((itraconazole)) was 0.69. Conclusion: The developed population pharmacokinetic model adequately described the pharmacokinetics of itraconazole and its active metabolite, hydroxy-itraconazole, in paediatric patients with either cystic fibrosis or undergoing BMT. More appropriate dosing schedules have been developed for the oral solution and the capsules to secure a minimum therapeutic trough plasma concentration of 0.5 mg/L for these patients.

Does high intensity resistance training maintain bone mass during moderate weight loss in older overweight adults with type 2 diabetes

The aim was to investigate whether the addition of supervised high intensity progressive resistance training to a moderate weight loss program (RT+WLoss) could maintain bone mineral density (BMD) and lean mass compared to moderate weight loss (WLoss) alone in older overweight adults with type 2 diabetes. We also investigated whether any benefits derived from a supervised RT program could be sustained through an additional home-based program. This was a 12-month trial in which 36 sedentary, overweight adults aged 60 to 80 years with type 2 diabetes were randomized to either a supervised gymnasium-based RT+WLoss or WLoss program for 6 months (phase 1). Thereafter, all participants completed an additional 6-month home-based training without further dietary modification (phase 2). Total body and regional BMD and bone mineral content (BMC), fat mass (FM) and lean mass (LM) were assessed by DXA every 6 months. Diet, muscle strength (1-RM) and serum total testosterone, estradiol, SHBG, insulin and IGF-1 were measured every 3 months. No between group differences were detected for changes in any of the hormonal parameters at any measurement point. In phase 1, after 6 months of gymnasium-based training, weight and FM decreased similarly in both groups (P < 0.01), but LM tended to increase in the RT+WLoss (n=16) relative to the WLoss (n = 13) group [net difference (95% CI), 1.8% (0.2, 3.5), P < 0.05]. Total body BMD and BMC remained unchanged in the RT+WLoss group, but decreased by 0.9 and 1.5%, respectively, in the WLoss group (interaction, P < 0.05). Similar, though non-significant, changes were detected at the femoral neck and lumbar spine (L2-L4). In phase 2, after a further 6 months of home-based training, weight and FM increased significantly in both the RT+WLoss (n = 14) and WLoss (n = 12) group, but there were no significant changes in LM or total body or regional BMD or BMC in either group from 6 to 12 months. These results indicate that in older, overweight adults with type 2 diabetes, dietary modification should be combined with progressive resistance training to optimize the effects on body composition without having a negative effect on bone health.

Hypothalamic regulation of cortical bone mass: Opposing activity of Y2 receptor and leptin pathways

NeuropeptideY-, Y2 receptor (Y2)-, and leptin-deficient mice show similar anabolic action in cancellous bone but have not been assessed in cortical bone. Cortical bone mass is elevated in Y2(-/-) mice through greater osteoblast activity. In contrast, leptin deficiency results in reduced bone mass. We show opposing central regulation of cortical bone.