Suppression of cyclooxygenase-2 gene transcription by humulon of beer hop extract studied with reference to glucocorticoid.
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In murine osteoblastic MC3T3-E1 cells which produced prostaglandin E2 as a bone resorption factor, the cyclooxygenase-2 induction by tumor necrosis factor alpha (TNFalpha) was suppressed by dexamethasone with an IC(50) of 1 nM. Humulon isolated from hop extract for beer brewing was reported previously as an inhibitor of bone resorption [Tobe, H. et al. (1997) Biosci. Biotech. Biochem. 61, 158-159]. We showed that the compound suppressed the TNFalpha-dependent cyclooxygenase-2 induction with an IC(50) of as low as about 30 nM as demonstrated experimentally by catalytic activity assay, Northern blot analysis and promoter analysis. Reporter gene experiments suggested that humulon blocked the cyclooxygenase-2 expression mediated by NFkappaB and NF-IL6, but the intracellular glucocorticoid receptor was not involved. The catalytic activity of cyclooxygenase-2 was inhibited by humulon with an IC(50) of as high as 1.6 microM. These results showed that humulon suppressed cyclooxygenase-2 induction at the step of transcription. (+info)
The effects of 1-year gluten withdrawal on bone mass, bone metabolism and nutritional status in newly-diagnosed adult coeliac disease patients.
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OBJECTIVES: To evaluate the impact of a 1-year gluten-free diet on bone metabolism and nutritional status in coeliac disease. METHODS: Bone mineral density, serum indices of bone remodelling, clinical and biochemical nutritional assessment were evaluated in 86 consecutive newly-diagnosed, biopsy proven, coeliac disease patients (untreated). A complete reevaluation, including intestinal biopsy, was repeated within 1 year of dietary treatment (treated). RESULTS: Untreated: according to WHO criteria, 34% of patients had a normal bone mineral density, 40% had osteopenia and 26% osteoporosis. Between males and females there were no statistical differences in bone metabolism or in most of the nutritional indices, while, between fertile and postmenopausal women, bone mineral density and several bone metabolism markers were significantly different. Compared to subjects with a normal bone mineral density, osteopenics had higher bone specific alkaline phosphatase (BAP) and Bone-Gla-protein (BGP) values. In patients with a concomitant BAP increase and 25OH vitamin D serum level reduction, bone mineral density and several bone turnover markers were statistically different compared to patients without such a serological pattern. Treated: notwithstanding intestinal biopsy which showed a mucosal recovery in only 57%, gluten-free diet led, even in postmenopausal women, to a significant improvement in bone mineral density, bone metabolism and nutrition, except for folic acid, albumin and pre-albumin serum levels which persisted as abnormal in patients with obdurate mucosal impairment. CONCLUSIONS: Coeliac disease patients are at high risk for developing a low bone mineral density and bone turnover impairment. A gluten-free diet can improve this situation even in postmenopausal women and in patients with incomplete mucosal recovery. (+info)
The cell biology of osteoclast function.
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Osteoclasts are multinucleated cells responsible for bone resorption. They have developed an efficient machinery for dissolving crystalline hydroxyapatite and degrading organic bone matrix rich in collagen fibers. When initiating bone resorption, osteoclasts become polarized, and three distinct membrane domains appear: a ruffled border, a sealing zone and a functional secretory domain. Simultaneously, the cytoskeleton undergoes extensive re-organisation. During this process, the actin cytoskeleton forms an attachment ring at the sealing zone, the membrane domain that anchors the resorbing cell to bone matrix. The ruffled border appears inside the sealing zone, and has several characteristics of late endosomal membrane. Extensive vesicle transport to the ruffled border delivers hydrochloric acid and proteases to an area between the ruffled border and the bone surface called the resorption lacuna. In this extracellular compartment, crystalline hydroxyapatite is dissolved by acid, and a mixture of proteases degrades the organic matrix. The degradation products of collagen and other matrix components are endocytosed, transported through the cell and exocytosed through a functional secretory domain. This transcytotic route allows osteoclasts to remove large amounts of matrix-degradation products without losing their tight attachment to underlying bone. It also facilitates further processing of the degradation products intracellularly during the passage through the cell. (+info)
Collagen fragments in urine derived from bone resorption are highly racemized and isomerized: a biological clock of protein aging with clinical potential.
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Fragments of the alpha1 C-terminal telopeptide of type I collagen containing the sequence AHDGGR(1209-1214) (CTx) can be measured in urine as an index of bone resorption. We report here that these molecules undergo racemization and isomerization of Asp(1211) in vitro and in vivo, generating a mixture of four isomers: the native peptide form (alphaL), an isomerized form containing a beta-Asp bond (betaL), a racemized form containing a D-Asp residue (alphaD) and an isomerized/racemized form (betaD). To study these reactions at this specific site in collagen, we have employed four immunoassays, each specific for one of the isoforms, and developed HPLC methods for their separation. The kinetics of these reactions were studied in vitro under physiological conditions by incubation of synthetic AHDGGR hexapeptide or mineralized bone collagen. Reactions were found to be strongly shifted towards the beta-Asp forms and slightly in favour of the D-enantiomeric forms. CTx isomers were measured in human urine and in enzymic digests of bovine bone collagen. The results indicated that the extent of racemization and isomerization were correlated with the age and turnover of collagen. The ratios between the native and age-related forms of CTx were elevated in urine from patients with Paget's disease or osteoporosis as compared with that from healthy adults. The alphaL/alphaD CTx ratio had the highest discriminatory power (T-score=23.2; P<0.0001 and T-score=1. 5; P<0.0001 for Paget's disease and osteoporosis respectively). In conclusion, these findings indicate that an assessment of CTx ratios in urine may provide an estimate of bone turnover, aiding in the diagnosis of metabolic bone diseases. (+info)
Matrix metalloproteinase-7-dependent release of tumor necrosis factor-alpha in a model of herniated disc resorption.
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Herniated disc (HD), one of the major causes of low back pain, is often resolved spontaneously without surgical intervention. Resorption is associated with a marked increase in infiltrating macrophages, and the matrix metalloproteinases (MMP) MMP-3 and MMP-7 have been implicated in this phenomenon. We developed a murine organ culture model in which intact intervertebral discs were cocultured with peritoneal macrophages to investigate the role of MMPs in HD resorption. Using macrophages isolated from MMP-null mice, we report that macrophage-produced MMP-7 was required for proteoglycan degradation, loss of wet weight, and macrophage infiltration of cocultured discs. The inability of MMP-7-deficient macrophages to infiltrate discs could not be attributed to a defect in macrophage migration. MMP-7 was required for the release of the cytokine TNF-alpha from peritoneal macrophages. The generation of soluble TNF-alpha was essential for the induction of MMP-3 in disc cocultures, which in turn is required for the generation of a macrophage chemoattractant and subsequent macrophage infiltration. TNF-alpha release from macrophages was necessary but insufficient for disc resorption, which required macrophage infiltration. We conclude that there is extensive communication between macrophages and chondrocytes in HD resorption and that an essential component of this communication is the requirement for MMPs to release soluble bioactive factors. (+info)
Requirement of B7 costimulation for Th1-mediated inflammatory bone resorption in experimental periodontal disease.
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The CD28 costimulation at TCR signaling plays a pivotal role in the regulation of the T cell response. To elucidate the role of T cells in periodontal disease, a system of cell transfer with TCR/CD28-dependent Th1 or Th2 clones was developed in rats. Gingival injection of specific Ag, Actinobacillus actinomycetemcomitans 29-kDa outer membrane protein, and LPS could induce local bone resorption 10 days after the transfer of Ag-specific Th1 clone cells, but not after transfer of Th2 clone cells. Interestingly, the presence of LPS was required not only for the induction of bone resorption but also for Ag-specific IgG2a production. LPS injection elicited the induction of expression of both B7-1 and B7-2 expression on gingival macrophages, which otherwise expressed only MHC class II when animals were injected with Ag alone. The expression of B7 molecules was observed for up to 3 days, which corresponded to the duration of retention of T clone cells in gingival tissues. Either local or systemic administration of CTLA4Ig, a functional antagonist of CD28 binding to B7, could abrogate the bone resorption induced by Th1 clone cells combined with gingival challenge with both Ag and LPS. These results suggest that local Ag-specific activation of Th1-type T cells by B7 costimulation appeared to trigger inflammatory bone resorption, whereas inhibition of B7 expression by CTLA4Ig might be a therapeutic approach for intervention with inflammatory bone resorption. (+info)
Effects of capsaicin-induced sensory denervation on osteoclastic resorption in adult rats.
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Many recent findings suggest that the nervous system has efferent effects on bone. A putative role of the sensory innervation has been assessed by using a synchronised rat model of bone resorption after treating adult animals with the neurotoxin capsaicin. Fourteen days after capsaicin treatment (50 mg kg-1) the right maxillary molars were extracted to activate a wave of resorption along the mandibular cortex. The rats were killed 4 days later (i.e. at the peak of resorption in this model), and their right mandibles were processed for histometric evaluation of resorption along the cortex and of calcitonin gene-related peptide (CGRP)- and substance P (SP)-immunoreactive (IR) fibres in the dental pulp. CGRP-IR and SP-IR fibres were significantly reduced in numbers by the capsaicin treatment (by 58 and 49%, respectively), confirming the success of sensory denervation. The resorption surface was significantly reduced (P < 0.005) versus the sham-treated animals. Although the size of the osteoclast population recruited in the site was not modified, the number of actively resorbing osteoclasts was significantly reduced (P < 0.03). However, the activity of the resorbing cells was not modified. Non-specific esterase-positive osteoclast precursors were also significantly few after capsaicin treatment. These data show that the sensory nervous system is involved in the control of bone resorption at two different levels: (1) in the recruitment of osteoclast precursors, and (2) in regulating the access of recruited cells to the bone surface. (+info)
Recombinant bone morphogenetic protein-7 as an intracorporal bone growth stimulator in unstable thoracolumbar burst fractures in humans: preliminary results.
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The study presented here is a pilot study in five patients with unstable thoracolumbar spine fractures treated with transpedicular OP-1 transplantation, short segment instrumentation and posterolateral fusion. Recombinant bone morphogenetic protein-7 in combination with a collagen carrier, also referred to as OP-1, has demonstrated ability to induce healing in long-bone segmental defects in dogs, rabbits and monkeys and to induce successful posterolateral spinal fusion in dogs without need for autogenous bone graft. Furthermore OP-1 has been demonstrated to be effective as a bone graft substitute when performing the PLIF maneuver in a sheep model. Five patients with single-level unstable burst fracture and no neurological impairment were treated with intracorporal OP-1 transplantation, posterior fixation (USS) and posterolateral fusion. One patient with osteomalacia and an L2 burst fracture had an additional intracorporal transplantation performed proximal to the instrumented segment, i.e. OP-1 into T 12 and autogenous bone into T 11. Follow-up time was 12-18 months. On serial radiographs, Cobb and kyphotic angles, as well as anterior, middle and posterior column heights, were measured. Serial CT scans were performed to determine the bone mineral density at fracture level. In one case, radiographic and CT evaluation after 3 and 6 months showed severe resorption at the site of transplantation, but after 12 months, new bone had started to fill in at the area of resorption. In all cases there was loss of correction with regard to anterior and middle column height and sagittal balance at the latest follow-up. These preliminary results regarding OP-1 as a bone graft substitute and stimulator of new bone formation have been disappointing, as the OP-1 device in this study was not capable of inducing an early sufficient structural bone support. There are indications to suggest that OP-1 application to a fracture site in humans might result in detrimental enhanced bone resorption as a primary event. (+info)