Significant Increase In Bone Resorption Research Articles

#1496 High levels of sclerostin promote bone resorption and changes and mineral metabolism

Abstract Background and Aims The physiological function of sclerostin (SOST) remains unknown. It is known that SOST is produced in osteocytes and functions as an inhibitor of the Wnt/β-catenin pathway. Similarly, it is well-established that low levels of SOST lead to bone alterations affecting cortical bone, resulting in osteopetrosis and reduced calciuria. High levels of SOST are associated with osteoporosis. In this study, we aimed to investigate the impact of three elevated doses of SOST on cortical, trabecular, and subchondral bone, as well as its relationship with other parameters of mineral metabolism. Method Bone histomorphometry, mCT, immunohistochemistry, and analysis of mineral metabolism parameters revealed that high doses of SOST over a 14-day period led to a reduction in trabecular bone volume due to a significant increase in bone resorption through the direct activation of osteoclastogenesis. Results Bone resorption, as measured by TRAP activity, was higher in trabecular, cortical, and subchondral bone. Similarly, high doses of SOST increased the number of hypertrophic chondrocytes, consequently expanding the growth plate area. Cortically, positive TRAP staining was observed, suggesting osteocytic osteolysis and trabecularization of cortical bone. The increased bone resorption resulted in a substantial rise in urinary excretion of phosphorus and calcium, accompanied by elevated levels of FGF23 and a significant decrease in PTH. Conclusion The findings suggest that elevated levels of SOST promote bone resorption through the activation of osteoclasts and the generation of osteocytic osteolysis. The increase in calcium and phosphorus levels led to changes in mineral metabolism, indicating a close relationship between SOST and other mineral metabolism parameters.

Androgen Deprivation Therapy Differentially Impacts Bone and Muscle in the Short Term in Physically Active Men With Prostate Cancer.

ABSTRACTAndrogen deprivation therapy (ADT) is a cornerstone of advanced prostate cancer (PCa) therapy. Its use is associated with a loss of bone mineral density (BMD) and a greater risk of falls and osteoporotic fractures. In this prospective cohort study, we examined the impact of ADT on muscle and bone strength in men initiating ADT for PCa. Participants were evaluated at three time points: immediately before (week 0), and 6 and 24 weeks after ADT initiation. Study measures included fasting blood levels (for markers of muscle and bone metabolic activity), MRI and QCT imaging (for muscle fat content, and bone density and architecture), and validated clinical tests of muscle strength and gait. Sixteen men completed all study visits. At baseline and throughout the study, participants exercised a median of four times/week, but still experienced weight gain (+2.0 kg at week 24 versus week 0, p = 0.004). Biochemically, all men sustained dramatic early and persistent reductions in sex hormones post‐ADT, along with a progressive and significant increase in serum C‐telopeptide of type I collagen (CTX, +84% at week 24 versus week 0). There was a trend for rise in serum sclerostin (p = 0.09) and interleukin 6 (IL‐6) (p = 0.08), but no significant change in serum myostatin (p = 0.99). Volumetric BMD by QCT declined significantly at the femoral neck (−3.7% at week 24 versus week 0), particularly at the trabecular compartment. On MRI, there were no significant changes in thigh muscle fat fraction. On physical testing, men developed weaker grip strength, but experienced no worsening in lower extremity and lumbar spine muscle strength, or on functional tests of gait. In conclusion, in physically active men, ADT for 24 weeks results in a significant increase in bone resorption and reduction in BMD, but nonsignificant changes in thigh muscle quality (on imaging) or strength and gait (on functional testing). © 2021 The Authors. JBMR Plus published by Wiley Periodicals LLC on behalf of American Society for Bone and Mineral Research.

Effects of the combined administration of risedronate and menatetrenone on bone loss induced by tacrolimus in rats.

Tacrolimus, a calcineurin inhibitor, affects bone metabolism and increases the risk of fracture due to marked bone loss. Bisphosphonates increase the bone mineral density (BMD) in osteoporosis patients. Menatetrenone has less positive effects on BMD but reduces the risk of fracture by improving bone quality. In this study, we investigated the effectiveness of the combined administration of risedronate and menatetrenone against bone loss induced by tacrolimus. Wistar rats were divided into four groups: [1] control, [2] tacrolimus at 1.5 mg/kg, [3] tacrolimus + risedronate at 1.0 mg/kg, and [4] tacrolimus + risedronate + menatetrenone at 20 mg/kg. After the drugs were administered for 4 weeks, bone histomorphometric analysis was performed and bone strength was evaluated using a three point bending method. BMD was measured using quantitative computed tomography. Tacrolimus significantly reduced the BMD and strength properties of the lower limb bones. These tacrolimusinduced decreases were suppressed by risedronate treatment. The combined administration of risedronate and menatetrenone more significantly improved bone strength properties than risedronate alone. Bone histomorphometric analysis revealed a significant increase in bone resorption with tacrolimus. Risedronate alone significantly suppressed the tacrolimus-induced increase in bone resorption but simultaneously reduced bone formation. On the other hand, the combined administration of risedronate and menatetrenone suppressed the tacrolimus-induced increase in bone resorption, in addition to the significant risedronate-induced decrease in bone formation. This study suggests that the combined administration of risedronate and menatetrenone improves bone strength in tacrolimus-treated rats by preventing and ameliorating the risedronate-induced suppression of bone formation.

Osteoporosis senil: una actualización

Knowledge of the mechanisms underlying the development of osteoporosis in the elderly has advanced greatly in the past few years. After an initial sudden loss of bone mineral mass in the peri-menopausal period there follows a more progressive and gradual loss that has also been seen in men. This initial drop in bone mass is due to a significant increase in bone resorption. There is also a significant reduction in bone formation with age that is mainly due to osteoblastogenesis in the bone marrow passing to a second plane, transferring its main role to adipogenesis. In this article, the latest evidence on the pathophysiology of senile osteoporosis is reviewed, highlighting the mechanisms of action of available treatments. Potential future treatments are also considered, which include new therapeutic approaches based on the pathophysiology of osteoporosis in the elderly, mainly on the potential reversibility of the adipogenesis.

Increased fracture risk in patients treated with thiazolidinediones: the role of abnormal bone turnover

Evaluation of: Zinman B, Haffner SM, Herman WH et al. Effect of rosiglitazone, metformin, and glyburide on bone biomarkers in patients with Type 2 diabetes. J. Clin. Endocrinol. Metab. 95(1), 134–142 (2010).Until recently, patients with Type 2 diabetes were thought to be relatively protected against osteoporosis by their high BMI. However, recent epidemiological data suggest that fracture risk is increased in diabetic patients. Additionally, use of thiazolidinedione antidiabetic medications (rosiglitazone and pioglitazone) was unexpectedly found to be associated with an increased risk of fractures. The paper under evaluation in this article examines changes in biochemical markers of bone turnover in subjects within the clinical trial that first raised the issue of fractures. The main finding was a significant increase in bone resorption with rosiglitazone. This conclusion conflicts with other studies examining the effects of thiozolidinediones on bone and, by itself, is unlikely to fully explain the increased fracture risk seen with these drugs.

Paget's Disease of Bone: Histologic Analysis of 754 Patients

Although Paget's disease of bone (PDB) is the second most common metabolic bone disease, to our knowledge, there is only one quantitative analysis on the histological and especially on the histomorphometric level. Therefore, the aim of this study was to analyze, on the basis of the Hamburg Bone Register, PBD in terms of incidence, skeletal distribution, malignant transformation, and histological and histomorphometric characteristics. Bone biopsies and patient files of 754 cases with histologically proven PDB were reviewed in a retrospective study. Quantitative static histomorphometry was performed on a representative subgroup of 247 biopsies derived from patients with manifestation of PDB at the iliac crest and compared with an age- and sex-matched control group. The peak incidence of PDB was between 70 and 80 yr of age. The majority of monostotic skeletal manifestation was localized at the os ilium, followed by the spine and femur. Histomorphometric results showed a high bone turnover with a significant increase in bone resorption and bone formation indices leading to an increased bone volume. Paget sarcoma was diagnosed in 6 of 754 patients, indicating a malignant transformation in 0.8% of the affected patients. Taken together, our study characterizes PDB in Germany on the basis of one of the largest cohorts of patients with histologically proven PDB. Moreover, for the first time, a quantitative histomorphometric approach was taken for >200 cases, where we could show local high bone mass lesions as a result of an increase of both osteoclast and osteoblast indices.

Increased osteoclastic activity in acute Charcot’s osteoarthopathy: the role of receptor activator of nuclear factor-kappaB ligand

Aims/hypothesisOur aims were to compare osteoclastic activity between patients with acute Charcot’s osteoarthropathy and diabetic and healthy controls, and to determine the effect of the receptor activator of nuclear factor-kappaB ligand (RANKL) and its decoy receptor osteoprotegerin (OPG).MethodsPeripheral blood monocytes isolated from nine diabetic Charcot patients, eight diabetic control and eight healthy control participants were cultured in the presence of macrophage-colony stimulating factor (M-CSF) alone, M-CSF and RANKL, and also M-CSF and RANKL with excess concentrations of OPG. Osteoclast formation was assessed by expression of tartrate-resistant acid phosphatase on glass coverslips and resorption on dentine slices.ResultsIn cultures with M-CSF, there was a significant increase in osteoclast formation in Charcot patients compared with healthy and diabetic control participants (p = 0.008). A significant increase in bone resorption was also seen in the former, compared with healthy and diabetic control participants (p < 0.0001). The addition of RANKL to the cultures with M-CSF led to marked increase in osteoclastic resorption in Charcot (from 0.264 ± 0.06% to 41.6 ± 8.1%, p < 0.0001) and diabetic control (0.000 ± 0.00% to 14.2 ± 16.5%, p < 0.0001) patients, and also in healthy control participants (0.004 ± 0.01% to 10.5 ± 1.9%, p < 0.0001). Although the addition of OPG to cultures with M-CSF and RANKL led to a marked reduction of resorption in Charcot patients (41.6 ± 8.1% to 5.9 ± 2.4%, p = 0.001), this suppression was not as complete as in diabetic control patients (14.2 ± 16.5% to 0.45 ± 0.31%, p = 0.001) and in healthy control participants (from 10.5 ± 1.9% to 0.00 ± 0.00%, p < 0.0001).Conclusions/interpretationThese results indicate that RANKL-mediated osteoclastic resorption occurs in acute Charcot’s osteoarthropathy. However, the incomplete inhibition of RANKL after addition of OPG also suggests the existence of a RANKL-independent pathway.

Polymethylmethacrylate particles stimulate bone resorption of mature osteoclasts in vitro

Background Interaction between wear particle debris and the cells at the implant-bone interface is an important contributory factor to periprosthetic bone loss seen in arthroplasties.Method To investigate the effect of this particle-induced response on different stages of osteoclast maturation, polymethylmethacrylate (PMMA) particles were added to a murine osteoclastogenic bone marrow cell culture system at either day 0, day 4, or day 8 of culture, which represented PMMA particle stimulation of precursor osteoclasts, mature osteoclasts, or end-stage osteoclasts, respectively. The number of TRAP-posi-tive multinucleate cells (MNCs) and the degree of bone resorption in culture were measuredResults Treatment of precursor osteoclasts with PMMA particles resulted in a statistically significant increase in TRAP-positive MNCs that persisted for 4 days, but there was no significant increase in bone resorption. Addition of particles to mature osteoclasts resulted in a significant increase in the number of TRAP-positive MNCs that lasted for 8 days, and also a significant increase in bone resorption. Treatment of end-stage osteoclasts with PMMA particles did not result in an increased number of TRAP-positive MNCs and there was no increase in bone resorption.Interpretation Treatment of mature osteoclasts with PMMA particles resulted in an elevated number of TRAP-positive cells. This persisted over a longer period of time than at the other stages of osteoclast development, and there was also a greater increase in bone resorption.

Age-related loss of muscle mass and bone strength in mice is associated with a decline in physical activity and serum leptin

The mechanisms underlying age-related loss of muscle and bone tissue are poorly understood but are thought to involve changes in sex hormone status, physical activity, and circulating levels of inflammatory cytokines. This study attempts to develop an animal model useful for evaluating these mechanisms in vivo. Male C57BL/6 mice were included for study at 3, 6, 12, 18, 24, and 29 months of age. Endocortical mineralizing surface, serum leptin, body weight, and percentage of body fat all increased between 6 and 12 months of age as activity level declined. Serum levels of the inflammatory marker IL-6 increased significantly after 12 months of age, following the observed increase in body weight and percent body fat. Hindlimb muscle mass declined significantly between 18 and 24 months of age, both absolutely and relative to total body mass, with a further decline (∼15%) between 24 and 29 months. Loss of muscle mass after 18 months of age was accompanied by a significant increase in bone resorption, as indicated by serum pyridinoline cross-links, and a significant decrease in fat mass, serum leptin, bone strength, bone mineral density, and vertical cage activity. No significant changes in serum testosterone with aging were detected in the mice, as levels were essentially constant between 6 and 29 months. Our data show that mice lose a significant amount of muscle and bone tissue with age, and this loss of musculoskeletal tissue is accompanied by a drop in serum leptin and preceded by a significant decrease in physical activity.

Mineral density and histomorphometric assessment of bone changes in the proximal tibia early after induction of type II collagen-induced arthritis in growing and mature rats.

Bone changes in both actively growing (6-week-old) and mature (6-month-old) rats with collagen-induced arthritis (CIA) were investigated in order to clarify the mechanisms of osteoporosis near inflamed joints in patients with early rheumatoid arthritis (RA) and juvenile RA. In female Sprague-Dawley rats, the proximal tibiae from the CIA and control groups early after immunization, when any influence of immobilization due to joint pain and swelling is minimal, were studied using dual X-ray absorptiometry and histomorphometry after double-labeling with tetracycline. Arthritis developed within 10-14 days after immunization in both growing and mature rats. Physical activity, growth, and body weight continued to resemble that of the control group for at least 10 days. The bone mineral density in the proximal tibia did not differ significantly between the CIA and control groups. In growing rats, a highly significant increase in bone resorption, and decreases in bone formation and trabecular bone volume became evident histomorphometrically before visible signs of arthritis had developed. In mature rats, bone formation was markedly decreased without an increase in bone resorption. The differences in the reaction between growing and mature rats reflected a difference in the number of remodeling sites (units) and an uncoupling between osteoblasts and osteoclasts. We conclude that osteoporosis near inflamed joints results from an imbalance between bone resorption and formation caused by immune reactions in the CIA rats. Moreover, a decrease in bone formation may, in part, precede the clinical onset of arthritis.

Biochemical markers of bone remodeling and bone sialoprotein in ankylosing spondylitis

The aim of this work was to determine bone mineral density (BMD) in a group of patients with ankylosing spondylitis (AS) and to study alterations in bone remodeling in these patients. Eighteen patients (16 males and two females) with AS, mean age 44.7, range 21–75, and 18 age- and sex-matched healthy controls were studied. BMD was evaluated by dual energy X-ray absorptiometry. The following biochemical markers of bone remodeling were studied: formation – serum amino and carboxyterminal propeptides of procollagen I (PINP and PICP); resorption – urinary total and free deoxypyridinoline and pyridinoline (TDpyr, FDpyr, TPyr and FPyr), crosslinked aminoterminal telopeptides of collagen I (NTX), carboxyterminal telopeptide of collagen I (CTX) and serum bone sialoprotein (BSP). Receiver operating characteristic (ROC) curves of markers were also performed. We found a decrease of bone mass and an increase in TPyr, FPyr, TDpyr, FDpyr, NTX and BSP in AS, but no significant differences were found in PICP, PINP and CTX. FDpyr, FPyr and TPyr showed the highest discrimination between patients and controls according to the results of the ROC curves. TPyr/TDpyr was higher in AS than in controls. We found osteopenia, with a normal formation and a significant increase in bone resorption in AS. FDpyr, FPyr and TPyr seem to present the best sensitivity for the study of alterations of bone resorption in this pathology, although NTX, TDpyr and BSP also show significant differences. The elevation in the ratio TPyr/TDpyr in AS compared to controls indicates that in AS there is a type I-collagen degradation in tissues different from bone.

RADIO-OPAQUE AGENTS IN BONE CEMENT INCREASE BONE RESORPTION

A heavy infiltrate of foreign-body macrophages is commonly seen in the fibrous membrane which surrounds an aseptically loose cemented implant. This is in response to particles of polymethylmethacrylate (PMMA) bone cement and other biomaterials. We have previously shown that monocytes and macrophages responding to particles of bone cement are capable of differentiating into osteoclastic cells which resorb bone. To determine whether the radio-opaque additives barium sulphate (BaSO4) and zirconium dioxide (ZrO2) influence this process, particles of PMMA with and without these agents were added to mouse monocytes and cocultured with osteoblast-like cells on bone slices. Osteoclast differentiation, as shown by the presence of the osteoclast-associated enzyme tartrate-resistant acid phosphatase (TRAP) and lacunar bone resorption, was observed in all cocultures. The addition of PMMA alone to these cocultures caused no increase in TRAP expression or bone resorption relative to control cocultures. Adding PMMA particles containing BaSO4 or ZrO2, however, caused an increase in TRAP expression and a highly significant increase in bone resorption. Particles containing BaSO4 were associated with 50% more bone resorption than those containing ZrO2. Our results suggest that radio-opaque agents in bone cement may contribute to the bone resorption of aseptic loosening by enhancing macrophage-osteoclast differentiation, and that PMMA containing is BaSO4 likely to be associated with more osteolysis than that containing ZrO2.

Radio-opaque agents in bone cement increase bone resorption.

A heavy infiltrate of foreign-body macrophages is commonly seen in the fibrous membrane which surrounds an aseptically loose cemented implant. This is in response to particles of polymethylmethacrylate (PMMA) bone cement and other biomaterials. We have previously shown that monocytes and macrophages responding to particles of bone cement are capable of differentiating into osteoclastic cells which resorb bone. To determine whether the radio-opaque additives barium sulphate (BaSO4) and zirconium dioxide (ZrO2) influence this process, particles of PMMA with and without these agents were added to mouse monocytes and cocultured with osteoblast-like cells on bone slices. Osteoclast differentiation, as shown by the presence of the osteoclast-associated enzyme tartrate-resistant acid phosphatase (TRAP) and lacunar bone resorption, was observed in all cocultures. The addition of PMMA alone to these cocultures caused no increase in TRAP expression or bone resorption relative to control cocultures. Adding PMMA particles containing BaSO4 or ZrO2, however, caused an increase in TRAP expression and a highly significant increase in bone resorption. Particles containing BaSO4 were associated with 50% more bone resorption than those containing ZrO2. Our results suggest that radio-opaque agents in bone cement may contribute to the bone resorption of aseptic loosening by enhancing macrophage-osteoclast differentiation, and that PMMA containing BaSO4 is likely to be associated with more osteolysis than that containing ZrO2.

P4. Transcriptional inhibition of the erythropoetin (EPO) gene by raised extracellular calcium: role of the negative calcium response element

The aim of this work was to determine bone mineral density (BMD) in a group of patients with ankylosing spondylitis (AS) and to study alterations in bone remodeling in these patients. Eighteen patients (16 males and two females) with AS, mean age 44.7, range 21–75, and 18 age- and sex-matched healthy controls were studied. BMD was evaluated by dual energy X-ray absorptiometry. The following biochemical markers of bone remodeling were studied: formation – serum amino and carboxyterminal propeptides of procollagen I (PINP and PICP); resorption – urinary total and free deoxypyridinoline and pyridinoline (TDpyr, FDpyr, TPyr and FPyr), crosslinked aminoterminal telopeptides of collagen I (NTX), carboxyterminal telopeptide of collagen I (CTX) and serum bone sialoprotein (BSP). Receiver operating characteristic (ROC) curves of markers were also performed. We found a decrease of bone mass and an increase in TPyr, FPyr, TDpyr, FDpyr, NTX and BSP in AS, but no significant differences were found in PICP, PINP and CTX. FDpyr, FPyr and TPyr showed the highest discrimination between patients and controls according to the results of the ROC curves. TPyr/TDpyr was higher in AS than in controls. We found osteopenia, with a normal formation and a significant increase in bone resorption in AS. FDpyr, FPyr and TPyr seem to present the best sensitivity for the study of alterations of bone resorption in this pathology, although NTX, TDpyr and BSP also show significant differences. The elevation in the ratio TPyr/TDpyr in AS compared to controls indicates that in AS there is a type I-collagen degradation in tissues different from bone.

Transient reduction in trabecular bone formation after discontinuation of administration of oestradiol-17β to ovariectomized rats

While the osteopenia associated with oestrogen deficiency is thought to arise from a relative defect in bone formation with respect to resorption, oestrogen administration itself leads to a decrease, rather than an increase, in bone formation. This decrease in bone formation, which arises from oestrogen's inhibitory effect on bone turnover, presumably masks any underlying tendency of oestrogen treatment towards stimulation of bone formation. To investigate this further, we have examined the early effect of discontinuing the administration of oestradiol-17 beta (OE2; 40 micrograms/kg) on bone formation indices in ovariectomized 13-week-old rats, before the turnover-induced increase in formation occurs. Histomorphometric indices were assessed at the proximal tibial metaphysis 0, 7, 10, 13 and 16 days following discontinuation of OE2 treatment. Measurements of body weight, uterine weight and longitudinal growth rate confirmed that there were rapid effects of OE2 deficiency on these parameters. We could detect no significant increase in bone resorption, as measured by osteoclast surface and number, until 16 days after ending treatment with OE2; this was coincidental with a reduction in bone volume. Shorter periods of OE2 deficiency were associated with a marked decrease in bone formation, as assessed by dynamic histomorphometric indices. This inhibition of bone formation was largely due to a reduction in double fluorochrome-labeled trabecular surfaces, which were decreased by approximately 70%. We conclude that ending OE2 administration in ovariectomized rats caused a striking decrease in trabecular bone formation, if such indices are assessed prior to the subsequent turnover-induced increase in formation.(ABSTRACT TRUNCATED AT 250 WORDS)

Calcitonin, estrogens and the bone

Estrogen deficiency following natural or surgical menopause, is thought to be the main factor leading to postmenopausal bone loss. Furthermore, after estrogen failure a significant reduction of intestinal calcium absorption and a negativization of calcium balance has been observed. The mechanism of estrogen effect on skeletal tissue is not yet fully elucidated. Recently, specific receptors for estrogens in osteoblastic cells have been described; however their low density does not give a full explanation about their functional role. Therefore estrogens act, at least in part, indirectly through calciotropic hormones. In order to further elucidate this issue, we performed some studies in postmenopausal osteoporotic patients and in fertile oophorectomized women. In the first double blind placebo controlled study, after a 1-yr estrogen treatment period we observed an increase in bone mineral content in the hormone-treated patients. Furthermore, in all treated patients an improvement of intestinal calcium absorption was detected, while 1,25-dihydroxy-vitamin D serum levels did not show significant changes. To further analyse the relationship between estrogens (E) and calcitonin (CT) in postmenopausal osteoporosis, we performed a double blind placebo controlled study to evaluate the effects of 1-yr estro-progestative treatment on CT secretory reserve, evaluated by calcium infusion test. Blood levels of CT showed a progressive increase during the study period in the hormone-treated group, with a significant increase in the CT response to calcium stimulation test, suggesting a modulation of CT secretion by E. Recently, we performed two studies in fertile oophorectomized women. In the first, we followed longitudinally 24 fertile women for 1 yr. In these patients we measured, before and after oophorectomy, biochemical indexes of bone metabolism and bone mass. During the observation period a significant increase in bone resorption and a significant drop in intestinal calcium absorption was observed. In the second study, performed on 14 women before and 6 months after oophorectomy, a treatment with conjugated estrogens allowed the correction of the primary intestinal defect responsible for the reduced calcium absorption. In conclusion, cumulative results of our studies suggest that: (a) estrogen deficiency represents the main pathogenetic factor in postmenopausal osteoporosis; (b) estrogen replacement therapy is effective in the prevention of rapid bone loss that occurs after menopause; (c) estrogens regulate CT secretion in postmenopausal women, so that CT may be considered a mediator of estrogen action on bone; (d) oophorectomy causes and early increase of bone resorption, and a later increase of bone formation; (e) oophorectomy causes a precocious impairment of intestinal calcium absorption, independent of vitamin D metabolism; (f) calcium malabsorption appears to be “resistant” to the action of vitamin D metabolites; and (g) in oophorectomized patients estrogen treatment protects both bone and intestine.

Evidence for Multiple Bone Resorption-Stimulating Factors Produced by Normal Human Keratinocytes in Culture*

Conditioned medium from cultured normal human foreskin keratinocytes enhanced the release of calcium from neonatal mouse calvaria in organ culture. Unfractionated keratinocyte-conditioned medium (KCM) stimulated bone resorption in a dose-dependent manner, but it did not increase the concentration of prostaglandin E2 (PGE2) in the bone culture medium until a maximal dose of KCM for resorption was used. Furthermore, inhibitors of PGE2 synthesis, indomethacin, ibuprofen, and piroxicam, did not inhibit KCM-induced calcium release. High concentrations of KCM increased cAMP production by calvaria in the presence of isobutylmethylxanthine, but the increase was small compared with that produced by a dose of bovine PTH that caused a similar level of bone resorption. The bone resorption-stimulating activity of KCM was not lost after incubation at 56 C for 60 min, but it was lost after heating at 100 C for 10 min. Fractionation of KCM by gel filtration chromatography revealed two distinct peaks of bone resorption-stimulating activity. One peak, KCMI, caused a significant increase in bone resorption at 2 micrograms protein/ml. KCMI did not increase medium PGE2, and inhibition of PGE2 synthesis in bone had no effect on KCMI-induced bone resorption. KCMI failed to increase cAMP production by human osteosarcoma SaOS-2 cells. Another peak, KCMII, caused a dose-dependent increase in bone resorption, and a significant increase in medium calcium was noted at a 20-fold lower concentration (0.1 microgram protein/ml) than with KCMI. In contrast to KCMI, the increase in bone resorption stimulated by KCMII was accompanied by a parallel increase in the production of PGE2, and inhibition of PGE2 synthesis completely inhibited the bone resorption-stimulating activity of KCMII. KCMII also caused an increase in cAMP production by SaOS-2 cells. We conclude that KCM contains at least two distinct bone resorption-stimulating factors, one of which acts via a PG-mediated mechanism and the other by a PG-independent pathway.

Endotoxin-stimulated mononuclear phagocytes secrete soluble factors with bone-resorbing activity.

An in vitro co‐culture system composed of bone marrow‐derived mononuclear phagocytes (obtained from endotoxin‐responsive mice) and 45Ca‐labeled calvaria (obtained from endotoxin‐hyporesponsive mice) was established in order to examine the direct effect of bacterial endotoxin on bone resorption mediated by mononuclear phagocytes. Co‐cultures treated with endotoxin exhibited a significant increase in bone resorption assessed in terms of the %45Ca released. This increase was observed in the absence of direct contact between the mononuclear phagocyte population and bone, indicating that enhanced resorption occurred as a function of soluble factors secreted by mononuclear phagocytes in response to endotoxin. Co‐cultures which contained devitalized 45Ca‐calvaria failed to exhibit an increase in the %45Ca released following endotoxin treatment. Consequently, the soluble factors secreted by endotoxin‐stimulated mononuclear phagocytes appeared to modulate the activity of endogenous bone‐resorbing cells. Two soluble factors with bone‐resorbing activity were distinguished upon subsequent analysis. These results suggest that the interaction between plaque‐associated bacterial endotoxin and mononuclear phagocytes may have important consequences relevant to the resorption of alveolar bone characteristic of chronic periodontal disease.

Effects of calcium and (inorganic) phosphate on bone resorption produced by vitamin D3 metabolites in vitro.

The effects of changing the concentrations of calcium (Ca2+) and inorganic phosphate (Pi) on 45Ca release from prelabeled fetal rat bones caused by parathyroid hormone (PTH), 25-hydroxyvitamin D3 (25-OH-D3) and 1, 25-dihydroxyvitamin D3 (1,25-(OH)2-D3) were tested, in vitro. Increasing total (Ca2+) from 1.25 mM to 2.5 mM caused a significant increase in bone resorption caused by submaximal concentrations of these materials. Further increase in total (Ca2+) to 3.5 mM abolished this potentiating effect of bone resorption. Increasing the (Pi) from 1 mM to 2-3 mM caused significant inhibition of bone resorption caused by these resorbing materials. In addition, a (Pi) of 3 mM caused significant inhibition of higher concentrations of PTH and 25-OH-D3. (Pi) of 2 and 3 mM also caused significant inhibition of 45Ca release from control bones. It is concluded that a moderate increase in medium (Ca2+) mimics the effect of these bone resorbing materials on bone. Phosphate appears to be a direct inhibitor of bone resorption. Therefore, changing the (Ca2+) and (Pi) in the medium can affect the bone resorbing effect of vitamin D3 metabolites, similar to their effect on PTH.