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Objective: To test whether inhibition of sclerostin by a targeted monoclonal antibody (Scl-Ab) protects from bone and cartilage damage in inflammatory arthritis. Sclerostin is a potent inhibitor of bone formation and may be responsible for the low level of bone repair in patients with rheumatoid arthritis., Methods: Human tumour necrosis factor transgenic mice (hTNFtg mice) developing inflammatory arthritis and local and bone loss were administered either vehicle, anti-TNF antibody, Scl-Ab, or a combination of both agents. Inflammation, systemic and periarticular bone loss, bone erosion and cartilage damage were evaluated at baseline (week 8) and after 3 weeks of treatment by clinical assessment, micro-CT and histology., Results: Scl-Ab did not affect joint swelling or synovitis. Systemic bone loss in the spine and periarticular bone loss in the proximal tibia were completely blocked and partially reversed by inhibition of sclerostin but not by inhibition of TNF. Moreover, Scl-Ab completely arrested the progression of bone erosion in hTNFtg mice and in combination with TNF inhibition even led to significant regression of cortical bone erosions. Protective effects of Scl-Ab were also observed for the articular cartilage., Conclusions: These data suggest that sclerostin inhibition is a powerful tool to enhance bone repair in inflammatory arthritis.

Objectives: AMG623, also known as A-623, is an antagonist of B-cell activating factor (BAFF). The present study was to evaluate the effects of AMG623 on murine models of autoimmune diseases., Methods: AMG623 was generated through phage library. Inhibitory activities of AMG623 against human and murine BAFF were measured by biacore binding and BAFF-mediated B-cell proliferation assay. Pharmacological effects of AMG623 were studied in BALB/c mice, collagen-induced arthritis model (CIA) and in the NZBxNZW F1 lupus model., Results: AMG623 binds to both soluble and cell surface BAFF. AMG623 blocks both human murine BAFF binding to the receptors. Treatment of AMG623 resulted in B-cell number reduction, and improvement of arthritis and lupus development in mice., Conclusions: AMG623 is a novel modality of BAFF antagonist. AMG623 is a potential therapeutic agent for the treatment of SLE, rheumatoid arthritis, and other B-cell-mediated autoimmune diseases.

Rodent models of immune-mediated arthritis (RMIA) are the conventional approach to evaluating mechanisms of inflammatory joint disease and the comparative efficacy of antiarthritic agents. Rat adjuvant-induced (AIA), collagen-induced (CIA), and streptococcal cell wall-induced (SCW) arthritides are preferred models of the joint pathology that occurs in human rheumatoid arthritis (RA). Lesions of AIA are most severe and consistent; structural and immunological changes of CIA best resemble RA. Lesion extent and severity in RMIA depends on experimental methodology (inciting agent, adjuvant, etc.) and individual physiologic parameters (age, genetics, hormonal status, etc.). The effectiveness of antiarthritic molecules varies with the agent, therapeutic regimen, and choice of RMIA. All RMIA are driven by overactivity of proinflammatory pathways, but the dominant molecules differ among the models. Hence, as with the human clinical experience, the efficacy of various antiarthritic molecules differs among RMIA, especially when the agent is a specific cytokine inhibitor.

Introduction: Bacterial arthritis causes rapidly progressing joint destruction in humans. We have shown that addition of bisphosphonates or corticosteroids to conventional antimicrobial agents decreases the activity of osteoclasts, thereby reducing bone destruction. Here we assess the effect of RANKL-targeted treatments using soluble receptor decoy and osteprotegerin (OPG) on the course and outcome of staphylococcal arthritis., Methods: Treatment was initiated 3 days after Staphylococcus aureus inoculation and included RANK-Fc, OPG-Fc, and OPG-Fc in combination with antibiotics. Control groups were treated with antibiotics, huFc, and PBS. Joints were evaluated for clinical signs of arthritis and histologically for bone and cartilage destruction. Bone mineral density (BMD) was evaluated using a peripheral quantitative computed tomography. Circulating markers of bone metabolism, inflammatory cytokines, and chemokines were analyzed in each group., Results: Mice treated with RANK-Fc or OPG-Fc in combination with antibiotics preserved total BMD and trabecular bone as compared to huFc or antibiotics. Treatment with RANK-Fc or OPG-Fc diminished the levels of bone resorption markers (osteocalcin, CTX-I, and TRACP5b). Neither RANK-Fc nor OPG-Fc influenced significantly the frequency and severity of arthritis., Conclusions: Inhibition of RANKL signalling efficiently prevents bone loss in the mouse model of bacterial arthritis even when started in the overt phase of infection.

Objective: To investigate the kinetics of bony spur formation and the relationship of bony spur formation to synovial inflammation and bone erosion in 2 rat arthritis models, and to address whether bony spur formation depends on the expression of tumor necrosis factor alpha (TNFalpha) or RANKL., Methods: Analysis of the kinetics of synovial inflammation, bone erosion, osteoclast formation, and growth of bony spurs was performed in rat collagen-induced arthritis (CIA) and adjuvant-induced arthritis (AIA). In addition, inhibition experiments were performed to assess whether inhibition of TNFalpha and RANKL by pegylated soluble TNF receptor type I (pegTNFRI) and osteoprotegerin (OPG), respectively, affected bony spur formation., Results: Bony spurs emerged from the periosteal surface close to joints, and initial proliferation of mesenchymal cells was noted as early as 3 days and 5 days after onset of CIA and AIA, respectively. Initiation of bony spur formation occurred shortly after the onset of inflammation and bone erosion. Neither pegTNFRI nor OPG could significantly halt the osteophytic responses in CIA and AIA., Conclusion: These results suggest that bony spur formation is triggered by inflammation and initial structural damage in these rat models of inflammatory arthritis. Moreover, emergence of bony spurs depends on periosteal proliferation and is not affected by inhibition of either TNFalpha or RANKL. Bony spur formation can thus be considered a process that occurs independent of TNFalpha and RANKL and is triggered by destructive arthritis.

Introduction: Rats with adjuvant-induced arthritis (AIA) were necropsied on 14 occasions during preclinical, acute clinical and chronic clinical stages of AIA progression to characterize local (joint protein extracts) and systemic (serum) levels of mediators regulating inflammation and bone erosion in conjunction with lymphoid tissue-specific leukocyte kinetics., Results: Systemic increases in alpha1 acid glycoprotein, tumor necrosis factor-alpha (TNFalpha), interleukin (IL)-17, transforming growth factor beta (TGFbeta), and chemokine (C-C motif) ligand 2 (CCL2) together with local IL-1alpha/beta and TGFbeta enrichment and local lymphoid hyperplasia preceded the onset of clinical disease and joint damage. Systemic upregulation of TNFalpha, IL-6, IL-17, TGFbeta, IL-18, CCL2, receptor activator of nuclear factor-kappabeta ligand (RANKL), and prostaglandin E(2) during acute and/or chronic AIA coincided with systemic leukocytosis and CD4+ T cell increase in blood and spleen. In contrast, progression of joint erosions during clinical AIA was associated with intra-articular increases in IL-1alpha/beta, IL-6, RANKL, IL-17, TGFbeta, CCL2, and KC/GRO and also a dramatic decline in osteoprotegerin., Conclusion: These data indicate that systemic and local events in inflammatory arthritis are discrete processes, driven by multiple cellular and humoral mediators with distinct kinetic profiles.

Introduction: Rat adjuvant-induced arthritis (AIA) and collagen-induced arthritis (CIA) feature bone loss and systemic increases in TNFalpha, IL-1beta, and receptor activator of NF-kappaB ligand (RANKL). Anti-IL-1 or anti-TNFalpha therapies consistently reduce inflammation in these models, but systemic bone loss often persists. RANKL inhibition consistently prevents bone loss in both models without reducing joint inflammation. Effects of these therapies on systemic markers of bone turnover and inflammation have not been directly compared., Methods: Lewis rats with established AIA or CIA were treated for 10 days (from day 4 post onset) with either PBS (Veh), TNFalpha inhibitor (pegsunercept), IL-1 inhibitor (anakinra), or RANKL inhibitor (osteoprotegerin (OPG)-Fc). Local inflammation was evaluated by monitoring hind paw swelling. Bone mineral density (BMD) of paws and lumbar vertebrae was assessed by dual X-ray absorptiometry. Markers and mediators of bone resorption (RANKL, tartrate-resistant acid phosphatase 5b (TRACP 5B)) and inflammation (prostaglandin E2 (PGE2), acute-phase protein alpha-1-acid glycoprotein (alpha1AGP), multiple cytokines) were measured in serum (day 14 post onset)., Results: Arthritis progression significantly increased paw swelling and ankle and vertebral BMD loss. Anti-TNFalpha reduced paw swelling in both models, and reduced ankle BMD loss in AIA rats. Anti-IL-1 decreased paw swelling in CIA rats, and reduced ankle BMD loss in both models. Anti-TNFalpha and anti-IL-1 failed to prevent vertebral BMD loss in either model. OPG-Fc reduced BMD loss in ankles and vertebrae in both models, but had no effect on paw swelling. Serum RANKL was elevated in AIA-Veh and CIA-Veh rats. While antiTNFalpha and anti-IL-1 partially normalized serum RANKL without any changes in serum TRACP 5B, OPG-Fc treatment reduced serum TRACP 5B by over 90% in both CIA and AIA rats. CIA-Veh and AIA-Veh rats had increased serum alpha1AGP, IL-1beta, IL-8 and chemokine (C-C motif) ligand 2 (CCL2), and AIA-Veh rats also had significantly greater serum PGE2, TNFalpha and IL-17. Anti-TNFalpha reduced systemic alpha1AGP, CCL2 and PGE2 in AIA rats, while anti-IL-1 decreased systemic alpha1AGP, IL-8 and PGE2. In contrast, RANKL inhibition by OPG-Fc did not lessen systemic cytokine levels in either model., Conclusions: Anti-TNFalpha or anti-IL-1 therapy inhibited parameters of local and systemic inflammation, and partially reduced local but not systemic bone loss in AIA and CIA rats. RANKL inhibition prevented local and systemic bone loss without significantly inhibiting local or systemic inflammatory parameters.

Rats with collagen-induced arthritis (CIA) were necropsied on 14 occasions from 4 days after induction to 27 days after disease onset to evaluate the kinetics of local (joint protein extracts) and systemic (serum) levels of inflammatory and pro-erosive factors. Systemic increases in alpha1 acid glycoprotein and KC/GRO together with systemic and local enrichment of interleukin (IL)-1beta, IL-6, CCL2, transforming growth factor (TGF)-beta and elevated IL-1alpha and IL-18 in joint extracts preceded the onset of clinical disease. Systemic upregulation of IL-1beta, IL-6, TGF-beta CCL2, RANKL and prostaglandin E(2) (PGE(2)) during acute and/or chronic CIA coincided with systemic leukocytosis and a CD4+ T-cell increase in blood and spleen. In contrast, progression of joint erosions during clinical CIA was associated with intra-articular increases in IL-1alpha/beta, IL-6, IL-18, CCL2, KC/GRO and RANKL, and a dramatic decline in osteoprotegerin (OPG). These data indicate that systemic and local events in inflammatory arthritis can be discrete processes, driven by multiple cellular and humoral mediators with distinct temporospatial profiles.

As anti-inflammatory treatments used in rheumatoid arthritis, such as glucocorticoids, often result in secondary detrimental effects on bone health, the objective of this study was to investigate the effects of oestrogen therapy (ET) on the development and activity of collagen-induced arthritis (CIA) in rats, with a focus on assessment of chondroprotective effects using biomarkers of type II collagen degradation. Forty female Lewis rats were allocated into four intervention groups: (i) control + vehicle; (ii) CIA + vehicle; (iii) CIA + ET; and (iv) CIA + prednisolone. During the 28-day intervention period we monitored body weight, time-point of disease onset, incidence of manifest disease and paw volume. Levels of the type II collagen degradation marker (CTX-II) were measured in serum. At euthanasia, hind paws were isolated, extracted for proteins and measured for the concentration of CTX-II. Matrix metalloproteinase (MMP) activity was evaluated using gelatinase zymography. Oestrogen treatment delayed the time-point of disease onset and reduced the incidence and degree of manifest immunoarthritis significantly, assessed by macroscopic evaluation of hind paw inflammation and paw volume. Measures of serum or tissue levels of CTX-II showed significantly reduced type II collagen degradation elicited by oestrogen treatment. In alignment, a decreased activity of MMP-2 and MMP-9 was found in the paw protein extracts. We have demonstrated that the anti-inflammatory effect of ET is linked to chondroprotective effects in an animal model of systemic immunoarthritis. As ET has positive rather than negative effects on bone health in contrast to prednisolone, these observations may be important for potential combination therapy.

Objective: To analyze the kinetics of osteoclastogenesis in 2 models of chronic immune-mediated arthritis and 1 model of acute arthritis., Methods: Adjuvant-induced arthritis (AIA) and collagen-induced arthritis (CIA) in Lewis rats were used as models of chronic arthritis. Acute arthritis was induced in Lewis rats by injecting carrageenan into the hind paw. Osteoclasts were identified by cathepsin K immunohistochemistry at various time points after the onset of arthritis. The location, size, and nucleation of osteoclasts were also analyzed., Results: In both AIA and CIA, multinucleated and cathepsin K-positive osteoclasts first were observed on the day of disease onset. Initially, osteoclasts were localized at the periosteum next to the synovial membrane and in subchondral bone channels. The number, size, and nucleation of osteoclasts rapidly increased, leading to severe bone loss within days after disease onset. In addition, numerous mononucleated cathepsin K-positive osteoclast precursor cells emerged in the synovial membrane. All osteoclasts (cathepsin K-positive, multinucleated, attached to bone) and osteoclast precursors (cathepsin K-positive, mononucleated or multinucleated, within synovial tissue) were also positive for a macrophage-specific marker. Upon induction of acute arthritis with carrageenan, osteoclasts formed transiently in subchondral bone, but regressed 7 days after disease onset., Conclusion: Functional osteoclasts are generated at the earliest stage of arthritis, and new precursors are continuously formed in the synovial membrane to replenish the osteoclast pool. These data indicate that anti-resorptive therapies may provide the most effective bone protection, when treatment is started soon after the onset of arthritis.

Unlabelled: RANKL is an essential mediator of bone erosions, but the role of RANKL in systemic bone loss had not been studied in arthritis. RANKL protein was increased in rat joint extracts and serum at the earliest stages of arthritis. Osteoprotegerin (OPG) treatment reversed local and systemic bone loss, suggesting that RANKL is both a marker and mediator of bone loss in arthritis., Introduction: RANKL is well established as an essential mediator of bone erosions in inflammatory arthritis, but the role of RANKL in systemic bone loss in arthritis had not been studied. We hypothesized that serum RANKL could serve as both a mediator and as a novel biomarker for local and systemic bone loss in arthritis. We challenged this hypothesis in two established rat models of inflammatory arthritis. We sought to determine whether serum RANKL was elevated early in disease progression and whether RANKL suppression could prevent both local and systemic bone loss in these models., Materials and Methods: Detailed time-course studies were conducted in animals with collagen-induced (CIA) or adjuvant-induced (AIA) arthritis to evaluate the onset and progression of inflammation (paw swelling), bone erosions, osteoclast numbers, and RANKL protein levels in arthritic joints and in serum. Additional CIA and AIA rats (n=8/group) received placebo (PBS) or recombinant OPG (3 mg/kg three times weekly) for 10 days beginning 4 days after disease onset (first macroscopic evidence of hind paw erythema and edema) to assess the role of RANKL in local and systemic bone loss., Results: RANKL protein was significantly elevated in the joints and serum of CIA and AIA rats within 1-2 days of disease onset. Increased RANKL levels were associated with local (hind paw) and systemic (vertebral) osteopenia in both models. The RANKL inhibitor OPG prevented local and systemic osteopenia in both models of established disease., Conclusions: RANKL protein is significantly increased both locally and systemically during the earliest stages of inflammatory arthritis in rats, suggesting that serum RANKL might have prognostic value for bone erosions and systemic osteopenia in this condition. RANKL inhibition through OPG prevented local and systemic bone loss in these arthritis models, suggesting that RANKL inhibition is a promising new approach for treating bone loss in arthritis.