Non-Canonical (RANKL-Independent) Pathways of Osteoclast Differentiation and Their Role in Musculoskeletal Diseases.

Osteoclasts are multinucleated cells derived from mononuclear phagocyte precursors (monocytes, macrophages); in the canonical pathway of osteoclastogenesis, these cells fuse and differentiate to form specialised bone-resorbing osteoclasts in the presence of receptor activator for nuclear factor kappa B ligand (RANKL). Non-canonical pathways of osteoclastogenesis have been described in which several cytokines and growth factors are able to substitute for RANKL. These humoral factors can generally be divided into those which, like RANKL, are tumour necrosis family (TNF) superfamily members and those which are not; the former include TNFα lymphotoxin exhibiting inducible expression and competing with herpes simplex virus glycoprotein D for herpesvirus entry mediator, a receptor expressed by T lymphocytes (LIGHT), a proliferation inducing ligand (APRIL) and B cell activating factor (BAFF); the latter include transforming growth factor beta (TGF-ß), interleukin-6 (IL-6), IL-8, IL-11, nerve growth factor (NGF), insulin-like growth factor-I (IGF-I) and IGF-II. This review summarises the evidence for these RANKL substitutes in inducing osteoclast differentiation from tissue-derived and circulating mononuclear phagocytes. It also assesses the role these factors are likely to play in promoting the pathological bone resorption seen in many inflammatory and neoplastic lesions of bone and joint including rheumatoid arthritis, aseptic implant loosening and primary and secondary tumours of bone.

Use of cost effective and rapid molecular tools for identification of Candida species, opportunistic pathogens.

BACKGROUND AND PURPOSE: Candidiasis is a widespread fungal infection caused by different Candida species. Rapid identification of Candida species in clinical laboratory is becoming increasingly important since the identification and discrimination of ethological agents for early treatment. We aimed at molecular identification of commonly Candida species isolated from clinical samples by using both PCR-RFLP assay and amplification of hwp1 gene. MATERIALS AND METHODS: Clinical samples comprising of vaginal specimens ,cutaneous, sputum, bronchoalveolar lavage(BAL,( and blood cultures were recovered from suspected patients. Candida isolates were initially identified phenotypically and confirmed by molecular approaches based on restriction fragment length polymorphism (PCR-RFLP (with MspI restriction enzyme. Amplification of hwp1 gene was performed for discrimination of C. albicans from C. dubliniensis and C.africana. RESULTS: The most abundant species were C. albicans (n=67; 44.6 %), C. glabrata (n=10; 20 %), C.tropicalis (n=20; 13.3 %), C. krusei (n=12; 8 %), C.parapsilosis (n=11; 7.3 %). Out of 67 C.albicans species, 6 species identified as C. dubliniensis and 4 species identified as C. africana. CONCLUSION: High frequency of non-albicansCandida species and differences in levels of susceptibility to the antifungal agents are important issues in medicine .Therefore, to manage the Candida-related infections properly, molecular diagnostic methods would be fast, reliable and even cost-effective approaches for identification of Candida species.

Sensibilisation of asthmatic patients to extracted antigens from strains of Aspergillus fumigatus, Aspergillus flavus and Aspergillus niger.

OBJECTIVE: The main purpose of this study sought to evaluate the frequency of sensitivity of Iranian asthmatic patients to three regional Aspergillus species of fumigatus, flavus and niger, by detection of antigen-specific IgE in the patients' sera. PATIENTS AND METHODS: Crude extracts were prepared following the disruption of fungi cell walls by the application of glass beads and their protein fractions were isolated by SDS-PAGE. After electrotransfer of protein bands into the nitrocellulose membrane, IgE-immunoblotting was performed against the sera from 32 asthmatic patients in addition to 20 healthy controls. RESULTS: Our results interestingly showed that all of the studied Iranian asthmatic patients were sensitive to A. fumigatus and A. flavus antigens. This frequency was 65.6% in the case of A. niger, however, all control samples were negative. Age/sex analysis generally indicated higher sensitivities of young patients (<30 years old) to Aspergillus species with a statistical significance in the case of A. niger (P=0.02) and additionally more sensitivity of females. Using Immunoblotting assay, 23 IgE-reactive allergenic components from A. fumigatus, 15 from A. flavus and 13 from A. niger in a broad molecular weight spectrum were identified, among which several fragments were not previously reported. CONCLUSION: Overall, this study found a high frequency of sensitivity of Iranian asthmatic patients to regional isolates of A. fumigatus, A. flavus and A. niger, which suggested the importance of these species in development of asthma. Moreover, we reported allergenic profiles of Iranian isolates in different patterns not previously observed.

Evidence of cis-acting regulatory variation in PTPN22 in patients with rheumatoid arthritis.

OBJECTIVES: To assess whether there are cis-regulatory polymorphisms that regulate protein tyrosine phosphatase, non-receptor type 22 (PTPN22) expression in rheumatoid arthritis (RA). METHODS: RNA was extracted from positively selected CD56+, CD8+, and CD4+ mononuclear cells and the 'residual' cells from 12 RA patients heterozygous for the PTPN22 C1858T single nucleotide polymorphism (SNP) (rs2476601). Relative allelic expression was measured by single base extension (SBE) assay. RESULTS: There was relative differential allelic expression (DAE ≥ 20%) in eight patients (p < 10(-5)); seven patients demonstrated DAE in more than one cell type; four patients had statistically significant differences between these cell populations (p(corrected) < 0.05). CONCLUSIONS: We have demonstrated significant differences in expression of PTPN22 alleles in RA patients, indicating the probable existence of cis-acting regulatory elements.

Osteoclast-like cells in soft tissue leiomyosarcomas.

Giant cell-rich leiomyosarcoma of soft tissues is an unusual variant of malignant smooth muscle tumor characterized by the presence of numerous multinucleated giant cells (MNGCs). The nature of MNGCs and the cellular mechanisms underlying their accumulation in this tumor are poorly understood. Analysis of the expression of osteoclast, macrophage, and smooth muscle markers in two cases of giant cell-rich leiomyosarcoma revealed that the MNGCs in giant cell-rich leiomyosarcoma were negative for smooth muscle markers and that these cells expressed an osteoclast-like phenotype, being positive for CD45, CD68, tartrate-resistant acid phosphatase, and CD51 but negative for CD14 and HLA-DR. Scattered tumor-associated macrophages (TAMs) also expressed this phenotype. Leiomyosarcoma tumor cells strongly reacted for CD51 but were negative for CD14, CD45, and CD68. An analysis of 25 conventional (nongiant cell-containing) leiomyosarcomas found isolated CD68(+) MNGCs in three cases (12%), all of which were grade II/III leiomyosarcomas containing a prominent TAM infiltrate. Leiomyosarcoma-derived TAMs in the presence of receptor activator for nuclear factor kappa B ligand (RANKL) and macrophage colony-stimulating factor were capable of differentiating into osteoclast-like cells capable of resorbing bone. Reverse transcription polymerase chain reaction studies showed that RANKL, osteoprotegerin, and TNF-related apoptosis-inducing ligand were expressed by leiomyosarcoma cells. Our findings indicate that the giant cells found in leiomyosarcomas are osteoclast-like and that they are formed from TAMs by a RANKL-dependent mechanism.

Cobalt, chromium and nickel affect hydroxyapatite crystal growth in vitro.

Metals are widely used in orthopaedics and recent studies have reported that patients with metal implants have a significant increase of metal levels in serum and synovial fluid. Femoral neck fracture occurred in some patients with metal-on-metal implants for unknown reasons. Recently, bone quality has emerged as an important factor of bone strength and few studies have investigated the effects of metal ions on hydroxyapatite properties. In the present study, we investigated the effects of Co(2+), Cr(3+) and Ni(2+) on hydroxyapatite (HA) growth in vitro, using carboxymethylated poly(2-hydroxyethyl methacrylate) (pHEMA) as a biomaterial for calcification. We have demonstrated that metal ions reduced the quantity of mineral formed at the surface of the polymer and decreased the ratio Ca/P by 1.12-, 1.05- and 1.08-fold for Cr(2+), Cr(3+) and Ni(2+) respectively. Furthermore, the size of calcospherites was significantly increased in the metal-doped HA compared to the controls, indicating a possible effect of metal ions on the crystal lattice. Indeed, the presence of metal ions increased the crystal size as well as the crystallinity of HA and reduce the lattice parameter c of the HA framework. The information obtained from this work suggests that the quality of the mineral around metallic implants could be altered. However, further investigation should be conducted to further elucidate the effects of metal incorporation on bone mineral and the functional consequences.

Increased osteoclastic activity in acute Charcot's osteoarthropathy: the role of receptor activator of nuclear factor-kappaB ligand.

AIMS/HYPOTHESIS: Our 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). METHODS: Peripheral 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. RESULTS: In 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/INTERPRETATION: These 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.

Cellular and humoral mechanisms of osteoclast formation in Ewing's sarcoma.

Cellular mechanisms that account for tumour osteolysis associated with Ewing's sarcoma are uncertain. Osteoclasts are marrow-derived multinucleated cells (MNCs) that effect tumour osteolysis. Osteoclasts are known to form from macrophages by both receptor activator for nuclear factor-kappaB (RANK) ligand (RANKL)-dependent and -independent mechanisms. In this study, our aim has been to determine whether tumour-associated macrophages (TAMs) isolated from Ewing's sarcoma are capable of differentiating into osteoclasts and to characterise the cellular and humoral mechanisms whereby this occurs. Tumour-associated macrophages were isolated from two Ewing's sarcomas and cultured on both coverslips and dentine slices for up to 21 days with soluble RANKL and macrophage colony stimulating factor (M-CSF). Osteoclast formation from TAMs (CD14+) was evidenced by the formation of tartrate-resistant acid phosphatase (TRAP) and vitronectin receptor (VNR)-positive MNCs, which were capable of carrying out lacunar resorption. This osteoclast formation was inhibited by the addition of bisphosphonates. Both Ewing's sarcoma-derived fibroblasts and some bone stromal cells expressed RANKL and supported osteoclast formation by a contact-dependent mechanism. We also found that osteoclast differentiation occurred when Ewing's TAMs were cultured with tumour necrosis factor (TNF)-alpha in the presence of M-CSF and that TC71 Ewing's sarcoma cells stimulated osteoclast formation through the release of a soluble factor, the action of which was abolished by an antibody to TNF-alpha. These results indicate that TAMs in Ewing's sarcoma are capable of osteoclast differentiation by both RANKL-dependent and TNF-alpha-dependent mechanisms and that Ewing's sarcoma cells produce osteoclastogenic factor(s). Our findings suggest that anti-resorptive and anti-osteoclastogenic therapies may be useful in inhibiting the osteolysis of Ewing's sarcoma.

RANKL-dependent and RANKL-independent mechanisms of macrophage-osteoclast differentiation in breast cancer.

The cellular and humoral mechanisms accounting for tumour osteolysis in metastatic breast cancer are uncertain. Osteoclasts, the specialised multinucleated cells responsible for tumour osteolysis, are derived from monocyte/macrophage precursors. Breast cancer-derived tumour-associated macrophages (TAMs) are capable of osteoclast differentiation but the cellular and humoral mechanisms controlling this activity are uncertain. In this study, TAMs were isolated from primary breast cancers and cultured in the presence and absence of cytokines/growth factors influencing osteoclastogenesis. Extensive TAM-osteoclast differentiation occurred only in the presence of RANKL and M-CSF; this process was inhibited by OPG and RANK:Fc, decoy receptors for RANKL. Breast cancer-derived fibroblasts and human bone stromal cells expressed mRNA for RANKL, OPG and TRAIL, and co-culture of these fibroblasts with human monocytes stimulated osteoclast formation by a RANKL-dependent mechanism. Osteoclast formation and lacunar resorption also occurred by a RANKL-independent mechanism when the conditioned medium from breast cancer cells, MDA-MB-231 and MCF-7, was added (with M-CSF) to monocyte cultures. Our findings indicate that TAMs in breast cancer are capable of osteoclast differentiation and that breast cancer-derived fibroblasts and breast cancer cells contribute to this process by producing soluble factors that influence osteoclast formation by RANKL-dependent and RANKL-independent mechanisms respectively.

LIGHT (TNFSF14), a novel mediator of bone resorption, is elevated in rheumatoid arthritis.

OBJECTIVE: Human osteoclast formation from mononuclear phagocyte precursors involves interactions between tumor necrosis factor (TNF) ligand superfamily members and their receptors. LIGHT is a transmembrane protein expressed and shed from the surface of activated T cells. Since activated T cells have been implicated in osteoclastogenesis in rheumatoid arthritis (RA), this study sought to determine whether LIGHT can regulate RANKL/cytokine-induced osteoclast formation, to identify the mechanism by which LIGHT influences osteoclastogenesis, and to investigate the presence of LIGHT in the serum of RA patients. METHODS: The effect of LIGHT on human and murine osteoclast formation was assessed in the presence and absence of neutralizing reagents to known osteoclastogenic factors. Serum levels of LIGHT in RA patients were measured by enzyme-linked immunosorbent assay. RESULTS: In the presence and absence of RANKL, LIGHT induced osteoclast formation from both human peripheral blood mononuclear cells and murine macrophage precursors, in a dose-dependent manner, whereas no inhibition was observed by adding osteoprotegerin, RANK:Fc, TNFalpha, or interleukin-8 or by blocking the LIGHT receptors herpesvirus entry mediator or lymphotoxin beta receptor. However, formation of osteoclasts was significantly decreased by the soluble decoy receptor for LIGHT, DcR3, and by blocking antibodies to the p75 component of the TNF receptor. A significant increase in LIGHT levels in the serum of RA patients compared with normal controls was also noted. CONCLUSION: Our results indicate that LIGHT promotes RANKL-mediated osteoclastogenesis and that it can induce osteoclast formation by a mechanism independent of RANKL. The increased concentration of LIGHT in patients with RA raises the possibility that LIGHT may play a role in immunopathogenic conditions that are associated with localized or systemic bone loss.

Malignant melanoma and bone resorption.

The cellular and humoral mechanisms accounting for osteolysis in skeletal metastases of malignant melanoma are uncertain. Osteoclasts, the specialised multinucleated cells that carry out bone resorption, are derived from monocyte/macrophage precursors. We isolated tumour-associated macrophages (TAMs) from metastatic (lymph node/skin) melanomas and cultured them in the presence and absence of osteoclastogenic cytokines and growth factors. The effect of tumour-derived fibroblasts and melanoma cells on osteoclast formation and resorption was also analysed. Melanoma TAMs (CD14+/CD51-) differentiated into osteoclasts (CD14-/CD51+) in the presence of receptor activator for nuclear factor kappaB ligand (RANKL) and macrophage-colony stimulating factor. Tumour-associated macrophage-osteoclast differentiation also occurred via a RANKL-independent pathway when TAMs were cultured with tumour necrosis factor-alpha and interleukin (IL)-1alpha. RT-PCR showed that fibroblasts isolated from metastatic melanomas expressed RANKL messenger RNA and the conditioned medium of cultured melanoma fibroblasts was found to be capable of inducing osteoclast formation in the absence of RANKL; this effect was inhibited by the addition of osteoprotegerin (OPG). We also found that cultured human SK-Mel-29 melanoma cells produce a soluble factor that induces osteoclast differentiation; this effect was not inhibited by OPG. Our findings indicate that TAMs in metastatic melanomas can differentiate into osteoclasts and that melanoma fibroblasts and melanoma tumour cells can induce osteoclast formation by RANKL-dependent and RANKL-independent mechanisms, respectively.

Bone stromal cells in pagetic bone and Paget's sarcoma express RANKL and support human osteoclast formation.

Paget's disease is a focal disorder of bone remodelling, in which there is an increase in osteoclast formation. A rare complication of Paget's disease is the development of a sarcoma, most commonly an osteosarcoma. Osteoclast formation occurs in the presence of macrophage-colony stimulating factor and receptor activator for nuclear factor-kappaB ligand (RANKL), and it has been shown that bone stromal cells in Paget's disease can influence osteoclast formation by modulating the expression of RANKL and its decoy receptor, osteoprotegerin (OPG). In this study we show that pagetic bone stromal cells express RANKL and that these cells promote osteoclast formation by a RANKL-dependent mechanism. Osteoclast formation in co-cultures of monocytes and either pagetic bone stromal cells or Paget's sarcoma stromal cells was not only induced by a contact-dependent mechanism but also occurred via the release of a soluble factor. In contrast to bone stromal cells isolated from normal controls, stromal cells isolated from morphologically normal bone in one patient with Paget's disease also stimulated osteoclast formation in this way; this osteoclastogenesis was inhibited by OPG. Our results indicate that Paget's bone stromal cells support osteoclast formation by a RANKL-dependent process which involves not only cell-cell contact but also secretion of soluble RANKL.

Synovial fluid macrophages are capable of osteoclast formation and resorption.

To determine whether synovial fluid (SF) macrophages isolated from the SF of osteoarthritis (OA), rheumatoid arthritis (RA) and pyrophosphate arthropathy (PPA) joints are capable of osteoclast formation, and to investigate the cellular and humoral factors required for this to occur, SF macrophages (CD14+) were isolated from the knee joint SF from patients with OA, RA and PPA and cultured for up to 14 days with macrophage-colony stimulating factor (M-CSF) and soluble receptor activator for nuclear factor-kappaB ligand (RANKL) or tumour-necrosis factor-alpha (TNFalpha) and interleukin-1alpha (IL-1alpha). Osteoclast differentiation was assessed by expression of tartrate-resistant acid phosphatase (TRAP) and vitronectin receptor (VNR), F-actin ring formation and lacunar resorption. Osteoclast formation and lacunar resorption was seen in RANKL-treated cultures of SF macrophages isolated from OA, RA and PPA joints with the largest amount of resorption noted in RA and PPA SF macrophage cultures. In TNFalpha/IL-1alpha-treated RA and PPA SF macrophage cultures, osteoclasts capable of lacunar resorption were also formed. Lacunar resorption was more extensive in RANKL than TNFalpha/IL-1alpha-treated cultures. These findings indicate that SF macrophages are capable of differentiating into mature osteoclasts capable of lacunar resorption. M-CSF in combination with RANKL or TNFalpha/IL-1alpha was required for osteoclast formation. As inflammatory synovial fluids contain an increase in the number of macrophages and an increase in the amounts of RANKL, TNFalpha and IL-1alpha, these findings suggest that one means whereby bone erosions may form in rheumatoid or crystal arthritis is by differentiation of synovial fluid macrophages into osteoclasts.

Arthroplasty membrane-derived fibroblasts directly induce osteoclast formation and osteolysis in aseptic loosening.

PURPOSE: Both macrophages and fibroblasts are the main cell types found in periprosthetic tissues surrounding failed joint arthroplasties. These fibroblasts are known to express RANKL and to produce TNFalpha, factors which promote osteoclast formation and bone resorption. In this study we have analysed the role that arthroplasty membrane-derived fibroblasts (AFb) play in inducing the generation of bone resorbing osteoclasts. METHODS: Fibroblasts were isolated from periprosthetic tissues and co-cultured with human monocytes in an osteoclast differentiation assay in the presence or absence of M-CSF and inhibitors of RANKL (OPG) and/or TNFalpha. RANKL expression by AFbs was determined by RT-PCR and the extent of osteoclast differentiation by the expression of TRAP, VNR and evidence of lacunar resorption. RESULTS: In the presence of M-CSF, large numbers of TRAP(+) and VNR(+) multinucleated cells capable of lacunar resorption, were noted in co-cultures of monocytes and RANKL-expressing AFbs. Cell-cell contact was required for osteoclast formation. The addition of OPG and anti-TNFalpha alone significantly reduced but did not abolish the extent of osteoclast formation, whereas the addition of both together abolished osteoclast formation and lacunar resorption. CONCLUSION: Our results indicate that fibroblasts in periprosthetic tissues are capable of inducing the differentiation of normal human peripheral blood mononuclear cells to mature osteoclasts by a mechanism that involves both RANKL and TNFalpha. Suppression of both RANKL and inflammatory cytokines is likely to be required to control periprosthetic osteolysis.

In vitro and in vivo biological responses to a novel radiopacifying agent for bone cement.

Iodixanol (IDX) and iohexol (IHX) have been investigated as possible radiopacification agents for polymethylmethacrylate (PMMA) bone cement, to replace the currently used barium sulphate and zirconia. IDX and IHX are both water-soluble iodine-based contrast media and for the last 20 years have been used extensively in clinical diagnostic procedures such as contrast media enhanced computed tomography, angiography and urography. One of the major reasons to remove the current radiopacifying agents is their well-documented cytotoxicity and their potential to increase bone resorption. Using in vitro bone resorption assays, the effect of PMMA particles plus IDX or IHX to induce osteoclast formation and lacunar resorption on dentine slices has been investigated. These responses have been compared with the in vitro response to PMMA particles containing the conventional radiopacifying agents, that is, barium sulphate and zirconia. In parallel, the in vivo reaction, in terms of new bone formation, to particles of these materials has been tested using a bone harvest chamber in rabbit tibiae. In vitro cell culture showed that PMMA containing IHX resulted in significantly less bone resorption than PMMA containing the conventional opacifiers. In vivo testing, however, showed no significant differences between the amounts of new bone formed around cement samples containing the two iodine-based opacifying agents in particulate form, although both led to fewer inflammatory cells than particles of PMMA containing zirconia. Our results suggest that a non-ionic radiopacifier could be considered as an alternative to the conventional radiopacifying agents used in biomaterials in orthopaedic surgery.

Transforming growth factor-beta induces osteoclast formation in the absence of RANKL.

Transforming growth factor beta (TGFbeta) is a multifunctional growth factor that is produced by many cells in bone and is abundant in the bone matrix. TGFbeta is known to regulate RANKL-induced osteoclast formation and bone resorbing activity. In this study we sought to determine whether TGFbeta could directly induce osteoclast formation by a RANKL-independent mechanism. We found that the addition of TGFbeta to cultures of human monocytes and RAW 264.7 cells (in the presence of M-CSF and the absence of RANKL, TNFalpha or IL-6/IL-11) was sufficient to induce the formation of TRAP+ and VNR+ cells, which formed actin rings and were capable of extensive lacunar resorption. The addition of osteoprotegerin or antibodies to TNFalpha and its receptors, as well as antibodies to gp130, did not inhibit lacunar resorption, indicating that TGFbeta did not act by stimulating RANKL, TNF or IL-6 production by monocytes. TGFbeta-induced osteoclast formation was qualitatively different from that induced by RANKL with numerous TRAP+/VNR+ mononuclear and small multinucleated cells being formed; these cells produced many small resorption lacunae. Our results indicate that TGFbeta, which is abundant in the bone matrix, can, in the presence of M-CSF, directly induce mononuclear phagocyte osteoclast precursors to differentiate into osteoclastic cells capable of lacunar resorption.

Osteoclast formation from circulating precursors in osteoporosis.

OBJECTIVE: An imbalance between bone formation and bone resorption is thought to underlie the pathogenesis of reduced bone mass in osteoporosis. Bone resorption is carried out by osteoclasts. which are formed from marrow-derived cells that circulate in the monocyte fraction. The aim of this study was to determine the role of osteoclast formation in the pathogenesis of bone loss in osteoporosis. METHODS: The proportion of circulating osteoclast precursors and their relative sensitivity to the osteoclastogenic effects of M-CSF. 1,25(OH)2D3 and RANKL were assessed in primary osteoporosis patients and normal controls. RESULTS: Although there was no difference in the number of circulating osteoclast precursors in osteoporosis patients and normal controls. osteoclasts formed from osteoporosis patients exhibited substantially increased resorptive activity relative to normal controls. Although no increased sensitivity to the osteoclastogenic effects of 1,25(OH)D3 or M-CSF was noted, increased bone resorption was found in osteoporosis peripheral blood mononuclear cell (PBMC) cultures to which these factors were added. CONCLUSION: Our findings suggest that osteoclast functional activity rather than formation is increased in primary involutional osteoporosis and that dexamethasone acts to increase osteoclast formation.

Cellular mechanisms of osteoclast formation and lacunar resorption in giant cell granuloma of the jaw.

BACKGROUND: Giant cell granuloma (GCG) is an osteolytic tumour of the jaw which is characterised by the presence of both mononuclear and multinucleated (osteoclast-like) giant cell components. The nature of these component cells and the pathogenesis of the extensive osteolysis associated with this lesion is uncertain. METHODS: Using cell culture techniques and immunohistochemistry, we defined the phenotypic characteristics of the mononuclear and multinucleated cells present in four cases of GCG of the jaw. We also analysed the cellular and humoral factors associated with osteoclast formation and osteolysis in these tumours and determined whether GCG stromal cells are capable of supporting osteoclast formation. RESULTS: GCG-derived giant cells expressed the phenotypic characteristics of osteoclasts (TRAP+, VNR+, and calcitonin responsive) and were capable of lacunar resorption. In addition to macrophages, the mononuclear cell population contained numerous spindle-shaped stromal cells which proliferated in culture and expressed RANKL; these GCG-stromal cells were capable of supporting human osteoclast formation from circulating monocyte precursors. CONCLUSION: Our findings indicate that the giant cells in GCG of the jaw are osteoclast-like and formed from monocyte/macrophage precursors which differentiate into osteoclasts under the influence of RANKL-expressing mononuclear stromal cells found in this lesion.

Interleukin-6 and interleukin-11 support human osteoclast formation by a RANKL-independent mechanism.

Interleukin-6 (IL-6) and interleukin-11 (IL-11) are known to influence osteoclast formation and bone resorption. In order to determine whether IL-6 and IL-11 could independently support human osteoclast formation, these factors were added to cultures of human peripheral blood mononuclear cells of the monocyte (CD14(+)) fraction in the presence of macrophage colony-stimulating factor (M-CSF). Under these conditions, IL-6 and IL-11 induced the formation of multinucleated cells which were positive for TRAP, VNR, and calcitonin receptor and capable of lacunar resorption. Osteoclastogenesis induced by IL-6 and IL-11 was inhibited by the addition of an anti-gp130 antibody but not by osteoprotegerin. These results indicate that IL-6 and IL-11, which are thought to play a role in several osteolytic bone disorders, are directly capable of inducing osteoclast formation by a RANKL-independent mechanism.