Receptor Activator Of NF-kappaB Ligand Levels Research Articles

Role of PDT as an adjunct to SRP on whole salivary RANKL and OPG ratio in type-2 diabetic and normoglycemic individuals with chronic periodontitis.

The aim was to assess the effect of scaling and root planing (SRP) with and without adjunct photodynamic therapy (PDT) on the levels of osteoprotegerin (OPG) receptor activator of NF-kappa B ligand (RANKL) in the unstimulated whole saliva (UWS) of type-2 diabetic and normoglycemic individuals with chronic periodontitis (CP). Type-2 diabetic and normoglycemic subjects with CP (Groups 1 and 2, respectively) were divided into test- (SRP + PDT) and control (SRP only) groups. Patient demographics were recorded; and periodontal parameters (marginal bone loss [MBL], probing depth [P.D], plaque index [PI], gingival index [GI], and clinical attachment loss [CAL]) were assessed at baseline and at 3-months-follow-up. Rate of flow of unstimulated whole saliva and levels of RANKL and osteoprotegerin were measured at both time intervals. P < 0.05 was considered statistically significant. Eighty-four persons with CP (42 with and 42 without type-2 DM) were included. At baseline, clinicoradiographic parameters were comparable in all groups. At 3-months of follow-up, there was no significant difference in the clinicoradiographic parameters in all groups. At 3-months of follow-up, there was no significant reduction in whole salivary RANKL and osteoprotegerin levels among individuals in the test and control groups among CP patients with and without CP. The whole salivary RANKL/OPG ratio remains high in patients with poorly-controlled type-2 DM after SRP with or without adjunct PDT.

Comparison of RANKL and osteoprotegerin levels in the gingival crevicular fluid of young cigarette- and waterpipe-smokers and individuals using electronic nicotine delivery systems

The aim of the present observational case-control study was to compare the levels of Receptor activator of NF-kappa B ligand (RANKL) and osteoprotegerin (OPG) in the gingival crevicular fluid (GCF) of cigarette- and waterpipe-smokers and electronic-nicotine-delivery-systems (ENDS)-users. Demographic data was collected using a questionnaire. Clinical periodontal parameters (plaque index [PI], bleeding on probing [BOP], probing depth [PD] and clinical attachment loss [CAL]) were measured; and GCF samples were collected from the deepest periodontal pocket of the mandibular right first molar. The GCF volume was determined and levels of RANKL and OPG were determined. Group comparisons were performed and P < 0.05 was considered statistically significant. One hundred and twenty male individuals (30 cigarette-smokers, 30 waterpipe users, 30 ENDS-users and 30 non-smokers) were included. Scores of PI (P < 0.01) and PD (P < 0.01) were significantly higher among cigarette- and waterpipe smokers, and ENDS-users than non-smokers. The GCF volume was significantly higher among cigarette- (0.92 ± 0.05 μl) (P < 0.01) and waterpipe-smokers (0.66 ± 0.08 μl) (P < 0.001) and ENDS-users (0.62 ± 0.03 μl) (P < 0.01) than non-smokers (0.21 ± 0.007 μl). The GCF RANKL levels were significantly higher among cigarette- (14.9 ± 8.2 pg/ml) (P < 0.001) and waterpipe-smokers (12.6 ± 8.8 pg/ml) (P < 0.01) and ENDS-users (11.5 ± 8.4 pg/ml) (P < 0.01) than non-smokers (3.5 ± 0.7 pg/ml). The GCF OPG levels were significantly higher among cigarette- (95.9 ± 7.2 pg/ml) (P < 0.001) and waterpipe-smokers (86.6 ± 5.8 pg/ml) (P < 0.01) and ENDS-users (77.5 ± 3.4 pg/ml) (P < 0.05) than non-smokers (21.5 ± 10.7 pg/ml). There was no significant difference in GCF RANKL and OPG levels among cigarette- and waterpipe smokers, ENDS users. Cigarette- and waterpipe smoking and ENDS usage is associated with an increased expression of RANKL and OPG in the GCF.

Weekly oral bisphosphonates over 2 years prevent bone loss in cardiac transplant patients.

Heart transplantation (HTX) is an established therapy for end-stage heart disease. The aim of this study was to determine whether application of oral bisphosphonates is effective in preventing osseous complications after HTX. Thirty-three cardiac transplant recipients were treated with alendronate 70mg/wk or risedronate 35mg/wk in combination with 1000mg calcium and 800IU vitamin D. Markers of bone metabolism and dual-energy X-ray absorptiometry (DXA) were determined directly after HTX and 2years later. Primary endpoints were changes in bone mineral density (BMD), markers of bone metabolism (osteocalcin, crosslaps), serum levels of the cytokines osteoprotegerin (OPG), receptor activator of NF kappa-B ligand (RANKL), and incidence of fractures. Eight patients presented with osteoporosis, and 16 patients with osteopenia by DXA without prevalent fractures. Over 2years, the BMD improved in 2 patients from osteoporosis to osteopenia, and overall BMD remained stable, and fractures did not occur. In addition, the serum levels of OPG increased (P<.0005), and the RANKL levels (P<.001) as well as the RANKL/OPG-ratio decreased significantly (P<.0005). The serum crosslaps showed no significant changes. The BMD showed a significant association with the increased 25-vitamin D levels only in females (P<.001). In heart transplanted patients, weekly oral bisphosphonates in combination with calcium and vitamin D supplementation preserved bone mass, prevented uncoupling of bone resorption/formation and fractures. Bone density should be measured and adequately treated, that is, with regular bisphosphonates.

The impact of serial radon and hyperthermia exposure in a therapeutic adit on pivotal cytokines of bone metabolism in rheumatoid arthritis and osteoarthritis.

Secondary osteoporosis is a frequent complication of rheumatoid arthritis (RA) and the result of an imbalance of catabolic and anabolic mechanisms of bone metabolism. The effects of serial low-dose radon and hyperthermia (LDRnHT) exposure in a therapeutic adit (12 applications in 3weeks) on the serum levels of the cytokines osteoprotegerin (OPG), receptor activator of NF kappa-B ligand (RANKL), tumor necrosis factor-α (TNF-α), and also on the RANKL/OPG ratio were investigated in 25 RA patients and an age-matched control of 24 patients with osteoarthritis (OA). Cytokine measurements were performed at baseline and after completion of LDRnHT. Anti-CCP antibodies (ACPA) were measured in RA patients in parallel. Medication in both groups was limited to non-steroidal anti-inflammatory drugs, and low-dose prednisolone (16 of 24 RA patients) as needed. RA and OA patients showed a significant decrease of TNF-α levels (p < 0.001). Both groups showed significantly decreased levels of RANKL (RA: p < 0.001, OA: p < 0.01). Only the RA patients presented a significant increase of OPG (p < 0.01) and decrease of the RANKL/OPG ratio (p < 0.01), and the ACPA levels (p < 0.001). LDRnHT results in a reduction of osteocatabolic and an increase of osteoanabolic cytokines, which represents the molecular basis for inhibiting osteoclastic activity in secondary osteoporosis and explains in part the effect of LDRnHT this physical therapy modality in a key inflammatory disease. Although reduced ACPA levels were observed under the therapy and although this could potentially contribute to an osteoprotective effect, in this case, it is rather uncertain as the reduction was only minor in magnitude.

Serum levels of M-CSF, RANKL and OPG in rats fed with Kashin-Beck disease-affected diet.

ObjectiveThere were no studies on the macrophage colony-stimulating factor (M-CSF), receptor activator of NF-kappaB ligand (RANKL) and osteoprotegerin (OPG) in the pathogenesis of Kashin-Beck disease (KBD). The objective of the present study was to investigate the serum M-CSF, RANKL and OPG in rats fed with KBD-affected diet.MethodsNinety Wistar rats were divided into five groups. The rats received standard commercial feed with or without T-2 toxin additive, low protein feed with or without or T-2 toxin additive and the KBD-affected feed. The serum bioactivity of M-CSF, RANKL and OPG was tested by enzyme-linked immunosorbent assay.ResultsThe serum levels of M-CSF in E group rats were higher than those in the other groups in the five groups (P < 0.01). The serum levels of RANKL and OPG in E group rats were highest in the five groups and have significant difference compared to the other groups (P < 0.05).ConclusionsThe molecule of M-CSF, RANKL and OPG may be involved in the regulation of epiphyseal plate injury and repair in KBD, and its participation in the pathogenesis of KBD should be studied in the future.

AB0390 Serum RANKL Activity is Suppressed by TNF Alpha Inhibition but not with MTX Alone

BackgroundBone is, together with cartilage a central target of rheumatoid arthritis (RA). Chronic synovitis attacks the juxtaarticular bone, where it causes bone erosions. Formation of bone erosions is a consequence...

Atorvastatin-induced increase in progenitor cell levels is rather caused by enhanced receptor activator of NF-kappaB ligand (RANKL) cell proliferation than by bone marrow mobilization

Background: Statins have been shown to increase the level of circulating progenitor cells in peripheral blood supposedly due to a mobilization of progenitor cells from the bone marrow niche. Osteoclast/osteoblast interaction has been associated with progenitor cell mobilization. Here, we investigated the role of statins on progenitor cell mobilization with a focus on bone metabolism. Methods and results: FGF2−/− and wild type (wt) mice were treated with atorvastatin or placebo. In contrast to wt mice, the number of sca-1/flk-1 positive progenitor cells in peripheral blood (PB) of atorvastatin treated FGF2−/− mice did not increase, and was accompanied by a defective reendothelialization after perielectric injury of the common carotid artery. In wt, but not FGF2−/− mice, statin treatment was associated with increased levels of receptor activator of NF-κB ligand (RANKL) in bone marrow (BM) supernatant. Treatment with recombinant RANKL increased sca-1/flk-1 positive progenitors in FGF2−/− mice. Interestingly, osteoclast activation was not altered. To measure the egress of sca-1/flk-1 positive progenitor cells from the bone marrow, we performed in-situ perfusion experiments of isolated hind limbs. Mobilization was not significantly affected by atorvastatin in both wt and FGF2−/− mice. Furthermore, RANK – the specific receptor to RANKL – is expressed on progenitor cells, and RANKL stimulation increases cell proliferation in vitro and in vivo. Conclusions: Atorvastatin treatment increases RANKL levels with no measurable effect on bone metabolism and mobilization of progenitor cells from BM to PB. RANKL is essential for the statin-mediated increase of progenitor cell levels but predominantly due to a RANKL-induced stimulation of cell proliferation.

Immunomodulatory drugs lenalidomide and pomalidomide inhibit multiple myeloma-induced osteoclast formation and the RANKL/OPG ratio in the myeloma microenvironment targeting the expression of adhesion molecules

Multiple myeloma (MM)-induced osteoclast (OC) formation is mainly due to an imbalance of the receptor activator NF-κB ligand (RANKL)-osteoprotegerin (OPG) ratio in favor of RANKL in the bone microenvironment and to the CCL3 production by MM cells. The purpose of the study was to investigate the effect of the immunomodulatory drugs on RANKL/OPG ratio, the production of pro-osteoclastogenic cytokines, and MM-induced OC formation. We found that invivo concentrations of both lenalidomide (LEN) and pomalidomide (POM) significantly blunted RANKL upregulation normalizing the RANKL/OPG ratio in human osteoprogenitor cells (PreOBs) when co-cultured with MM cells and also inhibited CCL3 production by MM cells. A reduction in CD49d expression, a molecule critically involved in RANKL upregulation in the MM microenvironment, accompanied this effect. Consistently, the pro-osteoclastogenic property of MM cells co-cultured with PreOBs was reduced by both LEN and POM. We further investigated the effect of these drugs on the transcriptional profile of both MM cells and PreOBs by microarray analysis, which showed that adhesion molecules, such as ITGA8 and ICAM2, are significantly downregulated in MM cells. Our data suggest that LEN and POM inhibit MM-induced OC formation through normalization of the RANKL/OPG ratio targeting the expression of adhesion molecules by MM cells.

Effects of high frequency loading on RANKL and OPG mRNA expression in ST-2 murine stromal cells

BackgroundOscillatory fluid flow (OFF)-induced shear stress leads to positive bone remodeling through pro-formative and anti-resorptive effects on bone cells. In this study, the effects of high frequency OFF on expression of receptor activator of NF-κB ligand (RANKL) and osteoprotegerin (OPG), two important regulators of osteoclast differentiation, were investigated.MethodsCells were exposed to 1 Pa peak shear stress using three loading frequencies (1, 10, and 20 Hz) widely employed in cell, animal, and clinical studies of bone remodeling. Two separate experiments were performed where either the total number of cycles (3600 cycles) or the total loading time (60 min) was kept constant. Real-time RT-PCR was used to quantify mRNA levels of RANKL, OPG.Results3600 cycles of OFF at 1 Hz and 10 Hz loading decreased RANKL/OPG ratio. Interestingly, these results were due to different mechanisms where at 1 Hz the decrease was due to an increase in OPG mRNA, whereas at 10 Hz the decrease was due to a decrease in RANKL mRNA.ConclusionAlthough high frequency OFF does not appear to further enhance the decrease in the RANKL/OPG ratio, these results suggest a potential to differentially control the change in either RANKL or OPG mRNA expression by applying different loading frequencies.

New insights into the pathogenesis of bone loss in recurrent calcium stone formers

Background/aims: Urolithiasis is one of the leading social and economical problems of modern society. It is estimated that 10% of males and 4 % of females between 30 and 50 years of age have urinary tract stone disease. Recurrence rate of urolithiasis is 75% at 15 years with no treatment. Calcium is the major calculus component since 75-80% of renal stones are composed of calcium oxalate, while idiopathic metabolic hypercalciuria is one of the most frequent causes of recurrent calcium urolithiasis. Numerous studies showed that urolithiasis patients have higher rate of bone resorption and lower bone mineral content as well as bone mineral density, which is more evident in idiopathic calcium stone formers. However, mechanisms of action and factors involved in bone loss in these patients are still unknown. Increased production of cytokines such as IL-1, IL-6, TNF as well as osteopontin (OPN), osteoprotegerin (OPG) and RANKL (receptor activator of NF-kappaB ligand) may enhance bone resorption and therefore lower bone mass in patients with idiopathic calcium urolithiasis. Aim of this study is to assess changes of cytokines involved in bone turnover and increased bone resorption, as that might play role in bone loss in recurrent calcium stone formers. Methods: 49 recurrent calcium stone formers were involved in the study and compared to healthy age and sex matched controls. Anthropometrical measurements and metabolic evaluation were performed. Bone mineral density (BMD ; g/cm2) was assessed by dual-energy x-ray absorptiometry (« Lunar Prodigy» ). BMD measurements were performed at the lumbar spine, femoral neck, forearm and total body. Results are presented as bone mineral content (BMC ; g) and bone area (BA ; cm2). Ca2+, Mg2+, P, creatinine, Na+, K+, Cl-, proteins, uric acid, alkaline phosphatase, osteocalcin, calcitonin, PTH, 25-OH D vitamin and 1, 25-OH D vitamin were assessed by standard biochemical serum analysis. IL-1, IL-6, TNF-α , OPG, RANKL and OPN serum levels were determined by quantitive enzyme-linked immunosorbent assay (ELISA) according to manufacture’ s protocol (Quantikine, R&D Systems, Inc., Minneapolis, USA). Results: The frequency of low BMD between male and female recurrent calcium stone formers was 36.4% to 45.8%, respectiveley. Different calcium excretion levels influenced the frequency of low BMD and ranged from 31.6% in normocalciuric patients to 50% in patients with hypercalciuria, respectiveley. The frequency of increased bone turnover markers showed differences between recurent stone formers with and without low BMD as well as between male and female participants. PTH was increased in 54.5% female participants with low BMD and in 25%

Osteogenic induction and 1,25-dihydroxyvitamin D 3 oppositely regulate the proliferation and expression of RANKL and the vitamin D receptor of human periodontal ligament cells

Objective Human periodontal ligament cells (hPDLCs) may play an important role in osteoclastogenesis in alveolar bone by expressing the receptor activator of NF-KappaB ligand (RANKL) and osteoprotegerin (OPG). The present study aimed to investigate the differences between the effects of osteogenic induction and 1,25-dihydroxyvitamin D 3 (VD 3) on hPDLC proliferation and the expression of RANKL, osteoprotegerin, and the vitamin D receptor (VDR) in hPDLCs. Methods Primary cultures of 11 hPDLC populations from 11 donors were obtained. Three samples of each hPDLC population from passage 3 were, respectively, treated with osteogenic induction medium, 10 −8 M VD 3, or vehicle as a control. Cell proliferation at days 0, 1, 3, 5, and 7 was estimated with the MTT method. At day 6, the mRNA levels of RANKL, OPG and VDR were determined with real-time RT-PCR. Results Osteogenic induction significantly promoted hPDLC proliferation, while VD 3 inhibited proliferation. Osteogenic induction significantly down-regulated the mRNA level of RANKL by 1.61-fold ( P = 0.033) and decreased the level of VDR by 2.13-fold ( P = 0.003), while there was no change in the level of OPG and OPG/RANKL ratio with osteogenic induction. On the contrary, VD 3 significantly up-regulated the level of RANKL by 9.58-fold ( P = 0.001) and increased the level of VDR by 3.15-fold ( P = 0.004), while down-regulating the OPG/RANKL ratio by 7.14-fold ( P = 0.004). Conclusion Osteogenic induction and VD 3 exert opposite effects in regulating hPDLC proliferation and mRNA expression of RANKL and VDR. This may induce hPDLCs to play different roles in alveolar bone metabolism.

New Perspective in Osteoarthritis: The OPG and RANKL System as a Potential Therapeutic Target?

Bone remodelling is tightly regulated by a molecular triad composed of OPG/RANK/RANKL. The receptor activator of NF-kappaB ligand (RANKL) (localized on osteoblasts) enhances osteoclastogenesis via interaction with its receptor RANK (localized on osteoclasts), whereas osteoprotegerin (OPG) (produced by osteoblasts) inhibits this osteoclastogenesis by binding to RANKL. The equilibrium between OPG and RANKL plays a crucial role in the pathophysiology of bone. Although some studies have shown the efficacy of OPG as a therapeutic agent against bone resorption, its bioavailability and mechanism of action after binding to RANKL have only recently been studied. A mechanistic investigation based on what becomes of OPG after binding to cells expressing membranous RANKL demonstrated an internalization process of OPG through the clathrin pathway prior to proteasomal and/or lysosomal degradation. Interestingly, the OPG internalization process reduced the half-life of RANKL. Recent evidence has shown that subchondral bone alterations in osteoarthritis (OA) are intimately involved in cartilage degradation, and that OPG/RANKL may be implicated. Data show that human OA subchondral bone osteoblasts have abnormal OPG and RANKL levels and consequently an altered OPG and RANKL ratio. Further data also reveal the involvement of some osteotropic factors in these altered levels and that some of these factors generally target RANKL with a differential modulation of the RANKL isoforms. Altogether, data suggest that this system could be targeted as a new strategy for the treatment of OA.

Staphylococcus aureusInduces Expression of Receptor Activator of NF-κB Ligand and Prostaglandin E2in Infected Murine Osteoblasts

Osteomyelitis is an inflammatory disease of the bone that is characterized by the presence of necrotic bone tissue and increased osteoclast activity. Staphylococcus aureus is responsible for approximately 80% of all cases of human osteomyelitis. While the disease is especially difficult to treat, the pathogenesis of S. aureus-induced osteomyelitis is poorly understood. Elucidating the molecular mechanisms by which S. aureus induces osteomyelitis could lead to a better understanding of the disease and its progression and development of new treatments. Osteoblasts can produce several soluble factors that serve to modulate the activity or formation of osteoclasts. Receptor activator of NF-kappaB ligand (RANK-L) and prostaglandin E(2) (PGE(2)) are two such molecules which can promote osteoclastogenesis and stimulate bone resorption. In addition, previous studies in our laboratory have shown that osteoblasts produce inflammatory cytokines, such as interleukin 6, following infection with S. aureus, which could induce COX-2 and in turn PGE(2), further modulating osteoclast recruitment and differentiation. Therefore, we hypothesized that following infection with S. aureus, osteoblasts will express increased levels of RANK-L and PGE(2). The results presented in this study provide evidence for the first time that RANK-L mRNA and protein and PGE(2) expression are upregulated in S. aureus-infected primary osteoblasts. In addition, through the use of the specific COX-2 inhibitor NS 398, we show that when PGE(2) production is inhibited, RANK-L production is decreased. These data suggest a mechanism whereby osteoblasts regulate the production of RANK-L during infection.

Effects of raloxifene therapy on circulating osteoprotegerin and RANK ligand levels in post-menopausal osteoporosis

Previous in vitro studies suggest that the anti-resorptive effect of raloxifene might be mediated by changes in several cytokines involved in the bone remodeling process. In this context, the osteoprotegerin (OPG)- receptor activator of NF kappa B ligand (RANKL) system is considered a key component in the osteoclastogenesis regulation. The aim of this study was to determine the effects of raloxifene treatment on serum concentrations of OPG, receptor RANKL and its relationship with biochemical markers of bone turnover and bone mineral density (BMD) in previously untreated women with post-menopausal osteoporosis. We selected 47 post-menopausal women (mean age 63+/-7 yr) with densitometric criteria of osteoporosis. We determined at baseline, 3, 6, and 12 months anthropometric parameters, biochemical markers of bone turnover, serum levels of 25(OH) D, serum levels of OPG and RANKL. BMD (dual-energy x-ray absorptiometry) in lumbar spine (LS) femoral neck and total hip was measured at baseline and 12 months after raloxifene (60 mg/day) treatment. Serum levels of OPG decreased in the 3rd and 6th month of treatment (p<0.001) and returned to basal levels in the 12th month. There was a significant decrease of RANKL levels and OPG/RANKL ratio after 1 yr of raloxifene treatment. In addition, BMD in LS increased significantly (2.5%) in the 12th month of treatment (p=0.031). Finally, the biochemical markers of bone turnover (total alkaline phosphatase, bone alkaline phosphatase, osteocalcin, tartrate-resistant acid phosphatase, urine cross-linked carboxi-terminal telopeptide of type I collagen) decreased significantly from the 3rd month of treatment. In conclusion, our results support the hypothesis that raloxifene may inhibit osteoclast activity, at least partly modulating the OPG-RANKL system.

Intracellular and surface RANKL are differentially regulated in patients with ankylosing spondylitis

Ankylosing spondylitis (AS) is characterized by ankylosis of axial joints but osteoporosis is also a well-reported feature. T cells have been implicated as a source of receptor activator of NFkappaB ligand (RANKL) in inflammatory bone diseases. Hence, we assessed whether T cells in patients with AS act as a source of RANKL too. Therefore, we investigated the expression of RANKL on T cells from 21 patients with AS by flow cytometry. Bone mineral density (BMD) was evaluated by quantitative computer tomography (QCT) and dual X-ray absorptiometry (DXA) and correlated with serum levels of osteoprotegerin (OPG) and RANKL. BMD was decreased in 45% of all patients when measured with DXA (48% with QCT) and correlated negatively with OPG. Expression of intracellular RANKL was increased on CD4+ (84 vs. 70%) and CD8+ (85.2 vs. 65.3%, P < 0.05) T cells in patients with AS, whereas expression of membrane-bound RANKL was significantly lower (CD4+: 2.2 vs. 8.5% and CD8+: 0.7 vs. 3.2%, P < 0.01). Our results indicate that surface and intracellular RANKL production is differentially regulated on T cells of patients with AS.

RANKL acts directly on RANK‐expressing prostate tumor cells and mediates migration and expression of tumor metastasis genes

Metastases to bone are a frequent complication of human prostate cancer and result in the development of osteoblastic lesions that include an underlying osteoclastic component. Previous studies in rodent models of breast and prostate cancer have established that receptor activator of NF-kappaB ligand (RANKL) inhibition decreases bone lesion development and tumor growth in bone. RANK is essential for osteoclast differentiation, activation, and survival via its expression on osteoclasts and their precursors. RANK expression has also been observed in some tumor cell types such as breast and colon, suggesting that RANKL may play a direct role on tumor cells. Male CB17 severe combined immunodeficient (SCID) mice were injected with PC3 cells intratibially and treated with either PBS or human osteprotegerin (OPG)-Fc, a RANKL antagonist. The formation of osteolytic lesions was analyzed by X-ray, and local and systemic levels of RANKL and OPG were analyzed. RANK mRNA and protein expression were assessed on multiple prostate cancer cell lines, and events downstream of RANK activation were studied in PC3 cells in vitro. OPG-Fc treatment of PC3 tumor-bearing mice decreased lesion formation and tumor burden. Systemic and local levels of RANKL expression were increased in PC3 tumor bearing mice. PC3 cells responded to RANKL by activating multiple signaling pathways which resulted in significant changes in expression of genes involved in osteolysis and migration. RANK activation via RANKL resulted in increased invasion of PC3 cells through a collagen matrix. These data demonstrate that host stromal RANKL is induced systemically and locally as a result of PC3 prostate tumor growth within the skeleton. RANK is expressed on prostate cancer cells and promotes invasion in a RANKL-dependent manner.

Diabetes modulates gene expression in the gingival tissues of patients with chronic periodontitis

This study evaluated whether diabetes modulates gene expression [interleukin (IL)-1beta, IL-1ra, IL-6, IL-8, IL-10; tumor necrosis factor (TNF)-alpha; interferon (IFN)-gamma, receptor activator of NF-kappaB ligand (RANKL) and osteoprotegerin (OPG)] in sites with periodontitis. Gingival biopsies were harvested and divided into three groups--Control group: systemically and periodontally healthy subjects (n = 10); Periodontitis group: systemically healthy subjects diagnosed with chronic periodontitis (n = 20); Diabetes group: type 1 diabetic subjects, diagnosed with chronic periodontitis (n = 20). Total RNA was obtained and analyzed by quantitative polymerase chain reaction. Data analysis demonstrated that, except for OPG, mRNA levels for all factors were increased by inflammation (P < 0.001). Interleukin-1beta, IL-1ra, IL-6, IL-8, IFN-gamma, and RANKL mRNA levels were higher in the diabetic group when compared with the control non-periodontitis group (P < 0.05), whereas IL-10 and OPG were lower (P < 0.05). No difference was observed for TNF-alpha between diabetic and control groups (P > 0.05). Diabetes lowered IL-1beta, IL-8, IL-10, TNF-alpha, RANKL, and OPG mRNA levels in sites with comparable type of periodontitis (P < 0.001). Moreover, increased RANKL:OPG and IL-6:IL-10 ratios were found. Taken together, these data suggest that decreased levels of IL-10 and OPG may play an important role in the periodontal breakdown in diabetic patients.

Gingival crevicular fluid levels of RANKL and OPG in periodontal diseases: implications of their relative ratio

Receptor activator of NF-kappaB ligand (RANKL) and osteoprotegerin (OPG) are a system of molecules that regulate bone resorption. This study aims to compare the levels of RANKL, OPG and their relative ratio in gingival crevicular fluid (GCF) of healthy and periodontal disease subjects. GCF was obtained from healthy (n=21), gingivitis (n=22), chronic periodontitis (n=28), generalized aggressive periodontitis (n=25) and chronic periodontitis subjects under immunosuppressant therapy (n=11). RANKL and OPG concentrations in GCF were measured by enzyme-linked immunosorbent assays. RANKL levels were low in health and gingivitis groups, but increased in all three forms of periodontitis. OPG levels were higher in health than all three periodontitis, or gingivitis groups. There were no differences in RANKL and OPG levels between chronic and generalized aggressive periodontitis groups, whereas these were lower in the immunosuppressed chronic periodontitis group. The RANKL/OPG ratio was significantly elevated in all three periodontitis forms, compared with health or gingivitis, and positively correlated to probing pocket depth and clinical attachment level. GCF RANKL and OPG levels were oppositely regulated in periodontitis, but not gingivitis, resulting in an enhanced RANKL/OPG ratio. This ratio was similar in all three periodontitis groups and may therefore predict disease occurrence.

The Role of Cyclooxygenase-2 (COX-2) in Inflammatory Bone Resorption

Prostaglandin E2 (PGE(2)) is an important inflammatory mediator that plays an essential role in the development and progression of periradicular diseases. Cyclooxygenase-2 (COX-2) is the inducible enzyme responsible for increased PGE(2) levels during inflammation and other pathologic processes. The purpose of this study was to determine the role of COX-2-mediated PGE(2) synthesis in osteoclast formation in response to endodontic pathogens and materials. Primary osteoblast cultures and osteoclast cultures were prepared from COX-2 knockout (K/O) and wild-type (WT) littermates. These cultured cells were exposed to lipopolysaccharide (LPS) or root canal obturation materials including gutta-percha (GP), Resilon (RS), mineral trioxide aggregates (MTAs), and AH Plus (AH+). Osteoclast formation was evaluated using tartrate-resistant acid phosphatase (TRAP) staining. The expression of receptor activator of NF-kappaB ligand (RANKL) and osteoprotegerin (OPG) was determined by real-time polymerase chain reaction (PCR) analysis. It was found that in both WT and K/O cultures, treatment with LPS led to a marked increase in osteoclast formation. The number of osteoclasts formed was significantly lower in K/O cultures compared to WT cultures. Exposure to endodontic materials did not lead to any significant osteoclast formation. LPS and endodontic materials caused a decrease in both RANKL and OPG expression in WT cells. In K/O cells, the baseline levels of RANKL and OPG expression were dramatically decreased compared to the WT cells. In conclusion, COX-2-mediated PGE(2) expression is required for LPS-induced inflammatory bone resorption and maintaining the baseline level of RANKL and OPG expression. LPS-induced osteoclast formation may be independent of the RANKL pathway.

Osteoprotegerin (OPG) binds with Tumor Necrosis Factor-Related Apoptosis-Inducing Ligand (TRAIL): Suppression of TRAIL-induced apoptosis in ameloblastomas

Osteoprotegerin (OPG) is a useful receptor in inhibiting Receptor Activator of NFkappaB Ligand (RANKL) in inducing osteoclastogenesis. Tumor Necrosis Factor (TNF)-Related Apoptosis-Inducing Ligand (TRAIL) is a potent apoptosis-inducing ligand in ameloblastomas. Since OPG has been reported to bind to TRAIL as well, the effect of OPG in TRAIL's function in inducing apoptosis should also be investigated. To investigate on the expression of OPG in ameloblastomas, immuhistochemistry, immunofluorescence and Western blot were performed. From the immunohistochemistry results, we found that OPG was expressed in ameloblastoma tissues. Expression of OPG was clearly seen in AM-1 cells by immunofluorescence as well. Additionally, Western blot analysis confirmed OPG expression in ameloblastoma tissues and AM-1 cells. To investigate on the potential of TNFalpha, TRAIL and RANKL in inducing apoptosis, we performed an apoptosis assay. From the apoptosis assay, we found that TRAIL had the highest potential in inducing apoptosis compared to TNFalpha and RANKL. To investigate the binding of OPG with RANKL and TRAIL, we performed a binding assay. We noticed that OPG preferably bind with RANKL than TRAIL. However, at low levels of RANKL, marked binding of OPG with TRAIL was seen. As we suspected that the binding of OPG and TRAIL might cause the effect of TRAIL in inducing apoptosis in ameloblastomas, we combined the treatment of OPG and TRAIL in AM-1 cells. From the apoptosis assay, we found that under treatment of OPG, TRAIL's function in inducing apoptosis was suppressed. These data suggest that by binding with TRAIL, OPG suppressed TRAIL's function in inducing apoptosis in ameloblastomas.