The RANKL/RANK/OPG pathway

Understanding of osteoclast formation and activation has advanced considerably since the discovery of the RANKL/RANK/OPG system in the mid 1990s. Osteoblasts and stromal stem cells express receptor activator of NF-jB ligand (RANKL), which binds to its receptor, RANK, on the surface of osteoclasts and their precursors. This regulates the differentiation of precursors into multinucleated osteoclasts and osteoclast activation and survival both normally and in most pathologic conditions associated with increased bone resorption. Osteoprotegerin (OPG) is secreted by osteoblasts and osteogenic stromal stem cells and protects the skeleton from excessive bone resorption by binding to RANKL and preventing it from interacting with RANK. The RANKL/OPG ratio in bone marrow is thus an important determinant of bone mass in normal and disease states. RANKL/RANK signaling also regulates lymph node formation and mammary gland lactational hyperplasia in mice, and OPG protects large arteries of mice from medial calcification. This article reviews the roles of the RANKL/RANK/OPG system in bone and other tissues.     

Osteoprotegerin and its Ligand: A New Paradigm for Regulation of Osteoclastogenesis and Bone Resorption

In just 3 years, striking new advances have been made in understanding the molecular mechanisms that govern the crosstalk between osteoblasts/stromal cells and hematopoietic osteoclast precursor cells that leads to osteoclastogenesis. Led first by the discovery of osteoprotegerin (OPG), a naturally occurring protein with potent osteoclastogenesis inhibitory activity, rapid progress was made to the isolation of RANKL, a transmembrane ligand expressed on osteoblasts/stromal cells that binds to RANK, a transmembrane receptor on hematopoietic osteoclast precursor cells. The interaction of RANK and RANKL initiates a signaling and gene expression cascade that results in differentiation and maturation of osteoclast precursor cells to active osteoclasts capable of resorbing bone. OPG acts as a decoy receptor, binding to RANKL and blocking its interaction with RANK, inhibiting osteoclast development. Many of the calciotropic hormones and cytokines, including 1,25(OH)₂D₃, PTH, PGE₂ and IL-11, appear to act through a dual capacity to inhibit production of OPG and stimulate production of RANKL. Estrogen, on the other hand, appears to inhibit production of RANKL and RANKL-stimulated osteoclastogenesis. Recently, the results of the first clinical trial with OPG supported its potential as a therapeutic agent for diseases such as osteoporosis. The new understanding provided by the RANK/RANKL/OPG paradigm for both differentiation of osteoclasts and their activation has had tremendous impact on the field and opened new avenues for development of possible treatments of diseases characterized by excessive bone resorption.     

骨保护素防治骨质疏松症的研究进展

骨保护素(osteoprotegerin, OPG)、核因子-κB受体活化因子配体(receptor activator of NF-κB ligand, RANKL)、核因子-κB受体活化因子(receptor activator of NF-κB, RANK)是调节骨吸收和骨形成的重要的调控因子.骨质疏松症是多种原因使破骨细胞功能活跃,破坏骨的动态平衡造成的.OPG与RANK竞争性结合RANKL,从而抑制破骨细胞的分化、激活,促进其凋亡.应用重组OPG蛋白和OPG基因治疗的方法 是目前防治骨质疏松症的关注热点.笔者针对这方面的研究现状作简要的综述.     

Novel Aspects on RANK Ligand and Osteoprotegerin in Osteoporosis and Vascular Disease

The clinical coincidence of osteoporosis and vascular disease has long indicated that common mediators may adversely affect bone metabolism and vascular integrity alike. Receptor activator of NF-kappaB ligand (RANKL) is an important cytokine for bone resorption that acts through its osteoclastic receptor, receptor activator of NF-kappaB (RANK), while osteoprotegerin serves as a decoy receptor that binds RANKL and prevents activation of RANK. Skeletal and vascular cells are sources and targets of RANKL and OPG both in vitro and in vivo. Modulation of the RANKL/RANK/OPG system in animals results in a skeletal and vascular phenotype, and administration of OPG may prevent osteoporosis and vascular calcification. Recent studies on OPG serum levels and gene polymorphisms also suggest an important role of this cytokine system in skeletal and vascular diseases. In summary, there is increasing evidence that RANKL and OPG may link the skeletal with the vascular system.     

Regulation of RANKL‐induced osteoclastogenesis by RING finger protein RNF114

Normal bone remodeling is a continuous process orchestrated by bone‐resorbing osteoclasts and bone‐forming osteoblasts, which an imbalance in bone remodeling results in metabolic bone diseases. RANKL, a member of the TNF cytokine family, functions as a key stimulator for osteoclast differentiation and maturation. Here, we report that RNF114, previously identified as a psoriasis susceptibility gene, plays a regulatory role in the RANKL/RANK/TRAF6 signaling pathway that mediates osteoclastogenesis. Our results demonstrated that RNF114 expression was significantly down‐regulated in mouse osteoclast precursor cells undergoing RANKL‐induced osteoclast differentiation. RNF114 knockout did not affect development or viability of the subpopulation of bone marrow macrophages capable of differentiating into osteoclasts in culture. However, in the presence of RANKL, RNF114 knockout bone marrow macrophages exhibited enhanced cell proliferation and augmented osteoclast differentiation, as shown by an increased expression of mature osteoclast markers, increased osteoclastic TRAP activity and bone resorption. Conversely, ectopic expression of RNF114 inhibited CTSK expression, TRAP activity, and bone resorption in RANKL‐treated pre‐osteoclasts. RNF114 also suppressed RANKL‐activated NFATc1 expression and NFAT‐regulated promoter activity. RNF114 suppressed TRAF6‐, but not TAK1/TAB2‐mediated NF‐κB activation downstream of RANKL/RANK. In particular, TRAF6 protein levels were down‐regulated by RNF114, possibly via K48‐mediated proteasome‐dependent degradation. These data suggested that RNF114's inhibitory effect on RANKL‐stimulated osteoclastogenesis was mediated by blocking RANK/TRAF6/NF‐κB signal transduction. © 2017 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 36:159–166, 2018.     

Role for osteoprotegerin in rheumatoid inflammation

Osteoprotegerin (OPG), a member of the TNF-receptor family expressed by osteoblasts, has documented effects on the regulation of bone metabolism. OPG inhibits bone resorption and binds with strong affinity to its ligand RANKL, thereby preventing RANKL from binding to its receptor RANK. This system is regulated by calcium-modifying hormones. OPG may also be pivotal in modulating the immune system. RANKL-deficient mice exhibit both severe immunological abnormalities and osteopetrosis, and activated T cells express RANKL mRNA. RANKL secretion by activated T cells may induce osteoclastogenesis via a mechanism enhanced by several cytokines (TNF-alpha, IL-1, and IL-17) that promote both inflammation and bone resorption. Conversely, this mechanism is inhibited by OPG, IL-4, and IL-10, which have antiinflammatory effects and inhibit osteoclast formation. Activated T cells in the rheumatoid synovium express RANKL. Synoviocytes can differentiate to osteoclast-like cells under specific conditions, particularly when they are cultured with M-CSF and RANKL. Thus, the bony erosions seen in RA may result from RANKL/RANK system activation by activated T cells. This raises the possibility that OPG therapy to block this mechanism might prove beneficial in patients with RA.     

Rôle de l’ostéoprotégérine dans l’inflammation et la polyarthrite rhumatoïde

Osteoprotegerin (OPG), a member of the TNF-receptor family expressed by osteoblasts, has documented effects on the regulation of bone metabolism. OPG inhibits bone resorption and binds with strong affinity to its ligand RANKL, thereby preventing RANKL from binding to its receptor RANK. This system is regulated by calcium-modifying hormones. OPG may also be pivotal in modulating the immune system. RANKL-deficient mice exhibit both severe immunological abnormalities and osteopetrosis, and activated T cells express RANKL mRNA. RANKL secretion by activated T cells may induce osteoclastogenesis via a mechanism enhanced by several cytokines (TNF-α, IL-1, and IL-17) that promote both inflammation and bone resorption. Conversely, this mechanism is inhibited by OPG, IL-4, and IL-10, which have antiinflammatory effects and inhibit osteoclast formation. Activated T cells in the rheumatoid synovium express RANKL. Synoviocytes can differentiate to osteoclast-like cells under specific conditions, particularly when they are cultured with M-CSF and RANKL. Thus, the bony erosions seen in RA may result from RANKL/RANK system activation by activated T cells. This raises the possibility that OPG therapy to block this mechanism might prove beneficial in patients with RA.     

Osteoclastogenesis during mouse tooth germ development is mediated by receptor activator of NFΚ-B ligand (RANKL)

To accommodate developing tooth germ in the alveolar bone, active bone resorption and the recruitment of numerous osteoclasts are essential. Recently, the signaling of receptor activator of nuclear factor-B (RANK) and its ligand (RANKL) was reported to play a pivotal role in osteoclast formation and activation. The aim of this study was to examine the expression of RANKL and the contribution of RANK-RANKL signaling to the process of tooth germ and alveolar bone development. In situ hybridization showed RANKL was expressed in dental follicle cells and osteoblasts on the alveolar bone surface surrounding developing tooth germs. To elucidate the function of RANKL, mouse mandibular explants on embryonic day 14 were subjected to organ culture with osteoprotegerin (OPG), an inhibitor of RANK-RANKL signaling as a decoy receptor of RANKL. Many tooth germs were compressed with the surrounding bone tissue in the OPG-treated explants, whereas these abnormalities were not seen in untreated explants. The numbers of tartrate-resistant acid phosphatase (TRAP)-positive osteoclastic cells aligning on the alveolar bone surface were significantly decreased in OPG-treated explants compared with untreated explants. Moreover, TRAP-positive osteoclastic cells were not observed along the alveolar bone surfaces depressing tooth germs. These observations suggest that osteoclastogenesis in the alveolar bone, which is essential for the accommodation of normal tooth development, is mediated by RANK-RANKL signaling.     

Expression profiles of receptor activator of nuclear factor kappaB ligand, receptor activator of nuclear factor kappaB, and osteoprotegerin messenger RNA in aged and ovariectomized rat bones.

The receptor activator of nuclear factor-kappaB ligand (RANKL; also known as tumor necrosis factor-related activation-induced cytokine [TRANCE], osteoprotegerin ligand [OPGL], and osteoclast differentiation factor [ODF]) is a transmembrane ligand expressed in osteoblasts and bone marrow stromal cells. It binds to RANK, which is expressed in osteoclast progenitor cells, and induces osteoclastogenesis. OPG, a decoy receptor for RANKL, also binds to RANKL, and competitive binding of RANKL with RANK or OPG is thought to regulate bone metabolism. To investigate roles of the RANKL/RANK/OPG system in pathophysiological conditions, the expression of RANKL, RANK, and OPG messenger RNA (mRNA) was analyzed in bones of aged and ovariectomized rats by means of in situ hybridization. In the control 8-week-old male and sham-operated female rat bones, the expression of RANKL mRNA was detected in hypertrophic chondrocytes of the growth plate and some periosteal and endosteal mesenchymal cells. The expression of RANK mRNA was detected in osteoclast-like cells and mononuclear cells in contact with the cortical and trabecular bones. The expression of OPG mRNA was detected in proliferating chondrocytes and osteocytes. In the 2.5-year-old rat bones, the expression of RANKL, RANK, and OPG mRNA tended to decrease except for the endosteal region. In the ovariectomized rat bones, the expression of RANKL, RANK, and OPG mRNA increased, and high expression of OPG mRNA was induced in resting chondrocytes and osteocytes. These results suggest that estrogen deficiency stimulates the RANKL/RANK/OPG system and induces OPG in cells that have been thought to be less important for bone metabolism.     

Novel Aspects on RANK Ligand and Osteoprotegerin in Osteoporosis and Vascular Disease

The clinical coincidence of osteoporosis and vascular disease has long indicated that common mediators may adversely affect bone metabolism and vascular integrity alike. Receptor activator of NF-B ligand (RANKL) is an important cytokine for bone resorption that acts through its osteoclastic receptor, receptor activator of NF-B (RANK), while osteoprotegerin serves as a decoy receptor that binds RANKL and prevents activation of RANK. Skeletal and vascular cells are sources and targets of RANKL and OPG both in vitro and in vivo. Modulation of the RANKL/RANK/OPG system in animals results in a skeletal and vascular phenotype, and administration of OPG may prevent osteoporosis and vascular calcification. Recent studies on OPG serum levels and gene polymorphisms also suggest an important role of this cytokine system in skeletal and vascular diseases. In summary, there is increasing evidence that RANKL and OPG may link the skeletal with the vascular system.     

Impaired bone resorption in cathepsin K-deficient mice is partially compensated for by enhanced osteoclastogenesis and increased expression of other proteases via an increased RANKL/OPG ratio

Previous reports indicate that mice deficient for cathepsin K (Ctsk), a key protease in osteoclastic bone resorption, develop osteopetrosis due to their inability to properly degrade organic bone matrix. Some features of the phenotype of Ctsk knockout mice, however, suggest the presence of mechanisms by which Ctsk-deficient mice compensate for the lack of cathepsin K. To study these mechanisms in detail, we generated Ctsk-deficient (Ctsk-/-) mice and analyzed them at the age of 2, 7, and 12 months using peripheral quantitative computed tomography, histomorphometry, resorption marker measurements, osteoclast and osteoblast differentiation cultures, and gene expression analyses. The present study verified the previously published osteopetrotic features of Ctsk-deficient mice. However, these changes did not exacerbate during aging indicating the absence of Ctsk to have its most severe effects during the rapid growth period. Resorption markers ICTP and CTX were decreased in the media of Ctsk-/- osteoclasts cultured on bone slices indicating impaired bone resorption. Ctsk-/- mice exhibited several mechanisms attempting to compensate for Ctsk deficiency. The number of osteoclasts in trabecular bone was significantly increased in Ctsk-/- mice compared to controls, as was the number of osteoclast precursors in bone marrow. The mRNA levels for receptor activator of nuclear factor (kappa)B ligand (RANKL) in Ctsk-/- bones were increased resulting in increased RANKL/OPG ratio favoring osteoclastogenesis. In addition, expression of mRNAs of osteoclastic enzymes (MMP-9, TRACP) and for osteoblastic proteases (MMP-13, MMP-14) were increased in Ctsk-/- mice compared to controls. Impaired osteoclastic bone resorption in Ctsk-/- mice results in activation of osteoblastic cells to produce increased amounts of other proteolytic enzymes and RANKL in vivo. We suggest that increased RANKL expression mediates enhanced osteoclastogenesis and increased protease expression by osteoclasts. These observations underline the important role of osteoblastic cells in regulation of osteoclast activity and bone turnover.     

Mathematical Modeling of Spatio‐Temporal Dynamics of a Single Bone Multicellular Unit

During bone remodeling, bone‐resorbing osteoclasts and bone‐forming osteoblasts are organized in bone multicellular units (BMUs), which travel at a rate of 20–40 μm/d for 6–12 mo, maintaining a cylindrical structure. However, the interplay of local BMU geometry with biochemical regulation is poorly understood. We developed a mathematical model of BMU describing changes in time and space of the concentrations of proresorptive cytokine RANKL and its inhibitor osteoprotegerin (OPG), in osteoclast and osteoblast numbers, and in bone mass. We assumed that osteocytes surrounding a microfracture produce RANKL, which attracted osteoclasts. OPG and RANKL were produced by osteoblasts and diffused through bone, RANKL was eliminated by binding to OPG and RANK. Osteoblasts were coupled to osteoclasts through paracrine factors. The evolution of the BMU arising from this model was studied using numerical simulations. Our model recapitulated the spatio‐temporal dynamics observed in vivo in a cross‐section of bone. In response to a RANKL field, osteoclasts moved as a well‐confined cutting cone. The coupling of osteoclasts to osteoblasts allowed for sufficient recruitment of osteoblasts to the resorbed surfaces. The RANKL field was the highest at the microfracture in front of the BMU, whereas the OPG field peaked at the back of the BMU, resulting in the formation of a RANKL/OPG gradient, which strongly affected the rate of BMU progression and its size. Thus, the spatial organization of a BMU provides important constraints on the roles of RANKL and OPG as well as possibly other regulators in determining the outcome of remodeling in the BMU.     

Inhibition of osteoclastogenesis by RNA interference targeting RANK

ABSTRACT: BACKGROUND: Osteoclasts and osteoblasts regulate bone resorption and formation to allow bone remodeling and homeostasis. The balance between bone resorption and formation is disturbed by abnormal recruitment of osteoclasts. Osteoclast differentiation is dependent on the receptor activator of nuclear factor NF-kappa B (RANK) ligand (RANKL) as well as the macrophage colony-stimulating factor (M-CSF). The RANKL/RANK system and RANK signaling induce osteoclast formation mediated by various cytokines. The RANK/RANKL pathway has been primarily implicated in metabolic, degenerative and neoplastic bone disorders or osteolysis. The central role of RANK/RANKL interaction in osteoclastogenesis makes RANK an attractive target for potential therapies in treatment of osteolysis. The purpose of this study was to assess the effect of inhibition of RANK expression in mouse bone marrow macrophages on osteoclast differentiation and bone resorption. METHODS: Three pairs of short hairpin RNAs (shRNA) targeting RANK were designed and synthesized. The optimal shRNA was selected among three pairs of shRNAs by RANK expression analyzed by Western blot and Real-time PCR. We investigated suppression of osteoclastogenesis of mouse bone marrow macrophages (BMMs) using the optimal shRNA by targeting RANK. RESULTS: Among the three shRANKs examined, shRANK-3 significantly suppressed [88.3%] the RANK expression (p < 0.01). shRANK-3 also brought about a marked inhibition of osteoclast formation and bone resorption as demonstrated by tartrate--resistant acid phosphatase (TRAP) staining and osteoclast resorption assay. The results of our study show that retrovirus-mediated shRANK-3 suppresses osteoclast differentiation and osteolysis of BMMs. CONCLUSIONS: These findings suggest that retrovirus-mediated shRNA targeting RANK inhibits osteoclast differentiation and osteolysis. It may appear an attractive target for preventing osteolysis in humans with a potential clinical application.     

Effect of interleukin-17 on nitric oxide production and osteoclastic bone resorption: is there dependency on nuclear factor-kappaB and receptor activator of nuclear factor kappaB (RANK)/RANK ligand signaling?

Interleukin-17 (IL-17) is a proinflammatory cytokine produced exclusively by activated memory T cells and has recently been found to stimulate osteoclastic resorption. Like other proinflammatory cytokines, IL-17 may affect osteoclastic bone resorption indirectly via osteoblasts, possibly by mechanisms previously reported for chondrocytes that respond in very similarly to osteoblasts. As in chondrocytes, but only in combination with tumor necrosis factor-alpha (TNF-alpha), IL-17 induced nitric oxide (NO) production in osteoblastic cells and fetal mouse metatarsals by a nuclear factor-kappaB (NF-kappaB)-dependent mechanism. This effect was associated with elevated mRNA levels of the NF-kappaB isoforms RelA and p50. In fetal mouse metatarsals, IL-17 stimulated osteoclastic bone resorption only in combination with TNF-alpha. The pathway by which the cytokine combination exerts this effect was examined using inhibitors of NO synthesis and NF-kappaB activation. Although both inhibitors used abolished NO production, they did not prevent the stimulatory effect of the cytokine combination on osteoclastic resorption. In contrast, the inhibitors slightly increased osteoclastic resorption, suggesting a suppressive rather than stimulatory effect of NO on cytokine-induced bone resorption. In addition, we showed that IL-17 + TNF-alpha stimulated osteoclastic resorption independent of NF-kappaB signaling. To further examine the pathway by which osteoclastic resorption was stimulated, we used osteoprotegerin, a specific inhibitor of the receptor activator of NF-kappaB (RANK)/receptor activator of the NF-kappaB ligand (RANKL) pathway. Osteoprotegerin partially inhibited IL-17 + TNF-alpha-stimulated osteoclastic resorption only at the high concentration of 1000 ng/mL, whereas it completely blocked parathyroid hormone-related peptide-stimulated resorption at 300 ng/mL. In conclusion, IL-17 stimulated NO production by an NF-kappaB-dependent pathway in osteoblastic cells and fetal mouse metatarsals only in combination with TNF-alpha. Neither NO production nor NF-kappaB signaling, and only partly the RANK/RANKL pathway, were involved in the stimulatory effect of the cytokine combination on osteoblastic bone resorption in these long bones, suggesting the existence of other pathways by which osteoclastic resorption can be stimulated.     

The ratio of messenger RNA levels of receptor activator of nuclear factor kappaB ligand to osteoprotegerin correlates with bone remodeling indices in normal human cancellous bone but not in osteoarthritis.

The determinants of cancellous bone turnover and trabecular structure are not understood in normal bone or skeletal disease. Bone remodeling is initiated by osteoclastic resorption followed by osteoblastic formation of new bone. Receptor activator of nuclear factor kappaB ligand (RANKL) is a newly described regulator of osteoclast formation and function, the activity of which appears to be a balance between interaction with its receptor RANK and with an antagonist binding protein osteoprotegerin (OPG). Therefore, we have examined the relationship between the expression of RANKL, RANK, and OPG and indices of bone structure and turnover in human cancellous bone from the proximal femur. Bone samples were obtained from individuals with osteoarthritis (OA) at joint replacement surgery and from autopsy controls. Histomorphometric analysis of these samples showed that eroded surface (ES/BS) and osteoid surface (OS/BS) were positively associated in both control (p < 0.001) and OA (p < 0.02), indicating that the processes of bone resorption and bone formation remain coupled in OA, as they are in controls. RANKL, OPG, and RANK messenger RNA (mRNA) were abundant in human cancellous bone, with significant differences between control and OA individuals. In coplotting the molecular and histomorphometric data, strong associations were found between the ratio of RANKL/OPG mRNA and the indices of bone turnover (RANKL/OPG vs. ES/BS: r = 0.93, p < 0.001; RANKL/OPG vs. OS/BS: r = 0.80, p < 0.001). These relationships were not evident in trabecular bone from severe OA, suggesting that bone turnover may be regulated differently in this disease. We propose that the effective concentration of RANKL is related causally to bone turnover.     

New roles of osteoblasts involved in osteoclast differentiation

Bone-resorbing osteoclasts are formed from a monocyte/macrophage lineage under the strict control of bone-forming osteoblasts. So far, macrophage colony-stimulating factor (M-CSF), receptor activator of nuclear factor-κB ligand (RANKL), and osteoprotegerin (OPG) produced by osteoblasts play major roles in the regulation of osteoclast differentiation. Recent studies have shown that osteoblasts regulate osteoclastogenesis through several mechanisms independent of M-CSF, RANKL, and OPG production. Identification of osteoclast-committed precursors in vivo demonstrated that osteoblasts are involved in the distribution of osteoclast precursors in bone. Interleukin 34 (IL-34), a novel ligand for c-Fms, plays a pivotal role in maintaining the splenic reservoir of osteoclast-committed precursors in M-CSF deficient mice. IL-34 is also able to act as a substitute for osteoblast-producing M-CSF in osteoclastogenesis. Wnt5a, produced by osteoblasts, enhances osteoclast differentiation by upregulating RANK expression through activation of the non-canonical Wnt pathway. Semaphorin 3A produced by osteoblasts inhibits RANKL-induced osteoclast differentiation through the suppression of immunoreceptor tyrosine-based activation motif signals. Thus, recent findings show that osteoclast differentiation is tightly regulated by osteoblasts through several different mechanisms. These newly identified molecules are expected to be promising targets of therapeutic agents in bone-related diseases.     

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

BACKGROUND: 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. METHODS: 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. RESULTS: 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. CONCLUSION: 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.     

Involvement of receptor activator of NFkappaB ligand and tumor necrosis factor-alpha in bone destruction in rheumatoid arthritis

Bone loss represents a major unsolved problem in rheumatoid arthritis (RA). The skeletal complications of RA consist of focal bone erosions and periarticular osteoporosis at sites of active inflammation, and generalized bone loss with reduced bone mass. New evidence indicates that osteoclasts are key mediators of all forms of bone loss in RA. TNF-alpha is one of the most potent osteoclastogenic cytokines produced in inflammation and is pivotal in the pathogenesis of RA. Production of tumor necrosis factor-alpha (TNF-alpha) and other proinflammatory cytokines in RA is largely CD4(+) T-cell dependent and mostly a result of interferon-gamma (IFN-gamma) secretion. Synovial T cells contribute to synovitis by secreting IFN-gamma and interleukin (IL)-17 as well as directly interacting with macrophages and fibroblasts through cell-to-cell contact mechanisms. Activated synovial T cells express both membrane-bound and soluble forms of receptor activator of NF-kappaB ligand (RANKL). In rheumatoid synovium, fibroblasts also provide an abundant source of RANKL. Furthermore, TNF-alpha and IL-1 target stromal-osteoblastic cells to increase IL-6, IL-11, and parathyroid hormone-related protein (PTHrP) production as well as expression of RANKL. In the presence of permissive levels of RANKL, TNF-alpha acts directly to stimulate osteoclast differentiation of macrophages and myeloid progenitor cells. In addition, TNF-alpha induces IL-1 release by synovial fibroblasts and macrophages, and IL-1, together with RANKL, is a major survival and activation signal for nascent osteoclasts. Consequently, TNF-alpha and IL-1, acting in concert with RANKL, can powerfully promote osteoclast recruitment, activation, and osteolysis in RA. The most convincing support for this hypothesis has come from in vivo studies of animal models. Protection of bone in the presence of continued inflammation in arthritic rats treated with osteoprotegerin (OPG) supports the concept that osteoclasts mediate bone loss, providing further evidence that OPG protects bone integrity by downregulating osteoclastogenesis and promoting osteoclast apoptosis. Modulation of the RANKL/OPG equilibrium in arthritis may provide additional skeletal benefits, such as chondroprotection. The nexus between T-cell activation, TNF-alpha overproduction, and the RANKL/OPG/RANK ligand-receptor system points to a unifying paradigm for the entire spectrum of skeletal pathology in RA. Strategies that address osteoclastic bone resorption will represent an important new facet of therapy for RA.