TNF-Tg Mice Research Articles

Du‐Huo‐Ji‐Sheng‐Tang Attenuates Inflammation of TNF‐Tg Mice Related to Promoting Lymphatic Drainage Function

To investigate whether Du-Huo-Ji-Sheng-Tang (DHJST) attenuate inflammation of RA related to lymphatic drainage function in vivo, we treated eight 3-month-old TNF-Tg mice with DHJST (12 g/kg) or the same volume of physiological saline once every day for 12 weeks, and 3-month-old WT littermates were used as negative control. After twelve weeks, we performed NIR-ICG imaging and found that DHJST increased the ICG clearance at the footpad and the pulse of efferent lymphatic vessel between popliteal lymph node and footpad. Histology staining at ankle joints showed that DHJST decreases synovial inflammation, bone erosion, cartilage erosion, and TRAP+ osteoclast area in TNF-Tg mice. Immunohistochemical staining by using anti-Lyve-1 and anti-podoplanin antibody showed that DHJST stimulated lymphangiogenesis in ankle joints of TNF-Tg mice. And zebrafish study suggested that DHJST promoted the formation of lymphatic thoracic duct. In conclusion, DHJST inhibits inflammation severity and promotes lymphangiogenesis and lymphatic drainage function of TNF-Tg mice.

In vivo quantification of lymph viscosity and pressure in lymphatic vessels and draining lymph nodes of arthritic joints in mice

Rheumatoid arthritis (RA) is a chronic inflammatory joint disease with episodic flares. In TNF-Tg mice, a model of inflammatory-erosive arthritis, the popliteal lymph node (PLN) enlarges during the pre-arthritic 'expanding' phase, and then 'collapses' with adjacent knee flare associated with the loss of the intrinsic lymphatic pulse. As the mechanisms responsible are unknown, we developed in vivo methods to quantify lymph viscosity and pressure in mice with wild-type (WT), expanding and collapsed PLN. While no differences in viscosity were detected via multiphoton fluorescence recovery after photobleaching (MP-FRAP) of injected FITC-BSA, a 32.6% decrease in lymph speed was observed in vessels afferent to collapsed PLN (P<0.05). Direct measurement of intra-lymph node pressure (LNP) demonstrated a decrease in expanding PLN versus WT pressure (3.41±0.43 vs. 6.86±0.56cmH2O; P<0.01), which dramatically increased to 9.92±1.79cmH2O in collapsed PLN. Lymphatic pumping pressure (LPP), measured indirectly by slowly releasing a pressurized cuff occluding indocyanine green (ICG), demonstrated an increase in vessels afferent to expanding PLN versus WT (18.76±2.34 vs. 11.04±1.47cmH2O; P<0.01), which dropped to 2.61±0.72cmH2O (P<0.001) after PLN collapse. Herein, we document the first in vivo measurements of murine lymph viscosity and lymphatic pressure, and provide evidence to support the hypothesis that lymphangiogenesis and lymphatic transport are compensatory mechanisms to prevent synovitis via increased drainage of inflamed joints. Furthermore, the decrease in lymphatic flow and loss of LPP during PLN collapse are consistent with decreased drainage from the joint during arthritic flare, and validate these biomarkers of RA progression and possibly other chronic inflammatory conditions.

AB0154 Radon therapy ameliorates disease progression and prolongs survival in TNF-transgenic mice

BackgroundRadon therapy as alternative therapy for rheumatic diseases in curative tunnels such as the Heilstollen Bad Gastein in Austria has been reported to reduce pain and amount of drug usage...

Bone fragility beyond strength and mineral density: Raman spectroscopy predicts femoral fracture toughness in a murine model of rheumatoid arthritis

Clinical prediction of bone fracture risk primarily relies on measures of bone mineral density (BMD). BMD is strongly correlated with bone strength, but strength is independent of fracture toughness, which refers to the bone's resistance to crack initiation and propagation. In that sense, fracture toughness is more relevant to assessing fragility-related fracture risk, independent of trauma. We hypothesized that bone biochemistry, determined by Raman spectroscopy, predicts bone fracture toughness better than BMD. This hypothesis was tested in tumor necrosis factor-transgenic mice (TNF-tg), which develop inflammatory-erosive arthritis and osteoporosis. The left femurs of TNF-tg and wild type (WT) littermates were measured with Raman spectroscopy and micro-computed tomography. Fracture toughness was assessed by cutting a sharp notch into the anterior surface of the femoral mid-diaphysis and propagating the crack under 3 point bending. Femoral fracture toughness of TNF-tg mice was significantly reduced compared to WT controls (p=0.04). A Raman spectrum-based prediction model of fracture toughness was generated by partial least squares regression (PLSR). Raman spectrum PLSR analysis produced strong predictions of fracture toughness, while BMD was not significantly correlated and produced very weak predictions. Raman spectral components associated with mineralization quality and bone collagen were strongly leveraged in predicting fracture toughness, reiterating the limitations of mineralization density alone.

Mechanisms of bone fragility in a mouse model of glucocorticoid‐treated rheumatoid arthritis: Implications for insufficiency fracture risk

Glucocorticoid (GC) therapy is associated with increased risk of fracture in patients with rheumatoid arthritis (RA). To elucidate the cause of this increased risk, we examined the effects of chronic erosive inflammatory arthritis and GC treatment on bone quality, structure, and biomechanical properties in a murine model. Mice with established arthritis and expressing human tumor necrosis factor α (TNFα) transgene (Tg) and their wild-type (WT) littermates were continually treated with GC (prednisolone 5 mg/kg/day via subcutaneous controlled-release pellet) or placebo for 14, 28, or 42 days. Microstructure, biomechanical properties, chemical composition, and morphology of the tibiae and lumbar vertebral bodies were assessed by micro-computed tomography, biomechanical testing, Raman spectroscopy, and histology, respectively. Serum markers of bone turnover were also determined. TNF-Tg and GC treatment additively decreased mechanical strength and stiffness in both the tibiae and the vertebral bodies. GC treatment in the TNF-Tg mice increased the ductility of tibiae under torsional loading. These changes were associated with significant alterations in the biochemical and structural composition of the mineral and organic components of the bone matrix, a decrease in osteoblast activity and bone formation, and an increase in osteoclast activity. Our findings indicate that the concomitant decrease in bone strength and increase in bone ductility associated with chronic inflammation and GC therapy, coupled with the significant changes in the bone quality and structure, may increase the susceptibility of the bone to failure under low-energy loading. This may explain the mechanism of symptomatic insufficiency fractures in patients with RA receiving GC therapy who do not have radiographic manifestations of fracture.

Validation of 3-dimensional ultrasound versus magnetic resonance imaging quantification of popliteal lymph node volume as a biomarker of erosive inflammatory arthritis in mice

Validation of 3-dimensional ultrasound versus magnetic resonance imaging quantification of popliteal lymph node volume as a biomarker of erosive inflammatory arthritis in mice

Radon therapy ameliorates disease progression and prolongs survival in TNF α tg mice

Backgroundand objectivesRadon therapy as alternative therapy for rheumatic diseases in curative tunnels such as the Heilstollen Bad Gastein in Austria has been reported to reduce pain and amount of drug...

Vascular endothelial growth factor C attenuates joint damage in chronic inflammatory arthritis by accelerating local lymphatic drainage in mice

To investigate whether the enhancement of joint lymphangiogenesis by injection of vascular endothelial growth factor C (VEGF-C) adeno-associated virus (AAV) into the affected joints has therapeutic efficacy in chronic inflammatory arthritis in mice. Tumor necrosis factor-transgenic (TNF-Tg) mice were used as a model of chronic inflammatory arthritis. Human VEGF-C was cloned into an AAV expression vector to generate AAV-VEGF-C. The joints of TNF-Tg mice were injected with AAV-VEGF-C or AAV-luciferase (AAV-Luc) as a control. During the 4 months following injection, magnetic resonance imaging of the joints and lymphatic imaging were performed to assess changes in synovial volume and lymph flow from the joint tissues to local draining lymph nodes. Joint inflammation, bone erosion, and cartilage loss were examined by histologic analyses. Lymphatic vessel formation was assessed using immunohistochemistry. Intraarticular administration of AAV-VEGF-C virus significantly attenuated the increase in synovial volume and increased lymphatic vessel number in the joint sections, as compared with that in control AAV-Luc-injected joints, during the 4-month period. This was accompanied by a reduction in the area of inflammation, bone erosion, cartilage loss, and osteoclast numbers. Lymph flow from the joints to local draining lymph nodes was slower in TNF-Tg mice than in wild-type littermates, and was significantly improved with AAV-VEGF-C treatment. Intraarticular injection of AAV-VEGF-C increased lymphangiogenesis and improved lymphatic drainage from the inflamed joints of mice, resulting in attenuation of joint tissue damage. Thus, improvement of joint lymphatic function by local administration of lymphatic growth factors represents a new therapeutic approach for chronic inflammatory arthritis.

CD23+/CD21hi B-cell translocation and ipsilateral lymph node collapse is associated with asymmetric arthritic flare in TNF-Tg mice

IntroductionRheumatoid arthritis (RA) is a chronic autoimmune disease with episodic flares in affected joints. However, how arthritic flare occurs only in select joints during a systemic autoimmune disease remains an enigma. To better understand these observations, we developed longitudinal imaging outcomes of synovitis and lymphatic flow in mouse models of RA, and identified that asymmetric knee flare is associated with ipsilateral popliteal lymph node (PLN) collapse and the translocation of CD23+/CD21hi B-cells (B-in) into the paracortical sinus space of the node. In order to understand the relationship between this B-in translocation and lymph drainage from flaring joints, we tested the hypothesis that asymmetric tumor necrosis factor (TNF)-induced knee arthritis is associated with ipsilateral PLN and iliac lymph node (ILN) collapse, B-in translocation, and decreased afferent lymphatic flow.MethodsTNF transgenic (Tg) mice with asymmetric knee arthritis were identified by contrast-enhanced (CE) magnetic resonance imaging (MRI), and PLN were phenotyped as "expanding" or "collapsed" using LNcap threshold = 30 (Arbitrary Unit (AU)). Inflammatory-erosive arthritis was confirmed by histology. Afferent lymphatic flow to PLN and ILN was quantified by near infrared imaging of injected indocyanine green (NIR-ICG). The B-in population in PLN and ILN was assessed by immunohistochemistry (IHC) and flow cytometry. Linear regression analyses of ipsilateral knee synovial volume and afferent lymphatic flow to PLN and ILN were performed.ResultsAfferent lymph flow to collapsed nodes was significantly lower (P < 0.05) than flow to expanding nodes by NIR-ICG imaging, and this occurred ipsilaterally. While both collapsed and expanding PLN and ILN had a significant increase (P < 0.05) of B-in compared to wild type (WT) and pre-arthritic TNF-Tg nodes, B-in of expanding lymph nodes (LN) resided in follicular areas while B-in of collapsed LN were present within LYVE-1+ lymphatic vessels. A significant correlation (P < 0.002) was noted in afferent lymphatic flow between ipsilateral PLN and ILN during knee synovitis.ConclusionsAsymmetric knee arthritis in TNF-Tg mice occurs simultaneously with ipsilateral PLN and ILN collapse. This is likely due to translocation of the expanded B-in population to the lumen of the lymphatic vessels, resulting in a dramatic decrease in afferent lymphatic flow. PLN collapse phenotype can serve as a new biomarker of knee flare.

Chronic axial compression of the mouse tail segment induces MRI bone marrow edema changes that correlate with increased marrow vasculature and cellularity

Magnetic resonance imaging (MRI) of bone marrow edema (BME) has been found to be helpful in the diagnosis of back pain attributed to degenerative disk disease (DDD) and spondyloarthropathy (SA), but its interpretation is limited by a lack of knowledge of its nature and natural history. We assessed effects of compressive forces to mouse tail segments of WT and TNF-Tg mice with SA, via contrast enhanced-MRI and histology. Normalized marrow contrast enhancement (NMCE) of uninstrumented WT vertebrae significantly decrease, threefold (p < 0.01) from 8 to 12 weeks of age, while the NMCE of TNF-Tg vertebrae remained elevated. Compressive loading (6x body weight) increased NMCE twofold (p < 0.02) within 2 weeks in WT tails, which was equal to 6x loaded TNF-Tg tails within 4 weeks. Histology confirmed degenerative changes and that load-induced NMCE corresponded to increased vascular sinus tissue (35 +/- 3% vs. 19 +/- 3%; p < 0.01) and cellularity (4,235 +/- 886 vs.1,468 +/- 320 cells/mm(2); p < 0.01) for the loaded versus unloaded WT, respectively. However, micro-computed tomography (CT) analyses failed to detect significant load-induced changes to bone. While the bone marrow of loaded WT and TNF-Tg vertebrae were similar, histology demonstrated mild cellular infiltrate and increased osteoclastic resorption in the WT tails versus severe inflammatory-erosive arthritis in TNF-Tg joints. Significant (p < 0.05) decreases in cortical and trabecular bone volume in uninstrumented TNF-Tg versus WT vertebrae were confirmed by micro-CT. Thus, chronic load-induced DDD causes BME signals in vertebrae similar to those observed from SA, and both DDD and SA signals correlate with a conversion from yellow to red marrow, with increased vascularity.

Expanded CD23+/CD21hi B Cells in Inflamed Lymph Nodes Are Associated with the Onset of Inflammatory-Erosive Arthritis in TNF-Transgenic Mice and Are Targets of Anti-CD20 Therapy

Anti-CD20 B cell depletion therapy (BCDT) is very effective for some patients with rheumatoid arthritis (RA); however the pathogenic role of B lymphocytes in RA and the primary targets of BCDT are unknown. The human TNF transgenic (hTNF-Tg) mouse model of RA displays a chronic, progressive disease that spreads from distal to proximal joints and is generally considered to be adaptive immune system independent. We have previously reported that knee arthritis in hTNF-Tg mice is accompanied by structural and functional changes of the adjoining popliteal lymph node (PLN), detectable by contrast-enhanced magnetic resonance imaging. To better understand these changes, in this paper we show that onset of knee synovitis and focal erosions are paralleled by PLN contraction and accumulation of large numbers of B cells in the lymphatic sinus spaces within the node. Flow cytometry from TNF-Tg mice 2, 4-5, and 8-12 mo old demonstrated that B cell accumulation in the PLN follows ankle arthritis, but commences before knee disease, and involves early expansion of CD21(hi), CD23(+), IgM(hi), CD1d(+), activation marker-negative, polyclonal B cells that are found to be specifically restricted to lymph nodes draining inflamed, arthritic joints. The same B cell population also accumulates in PLNs of K/BxN mice with autoantigen-dependent arthritis. Strikingly, we show that BCDT ameliorates hTNF-Tg disease and clears follicular and CD21(hi), CD23(+) B cells from the PLNs. On the basis of these findings, we propose a model whereby B cells contribute to arthritis in mice, and possibly RA, by directly affecting the structure, composition, and function of joint-draining lymph nodes.

NF-κB p100 limits TNF-induced bone resorption in mice by a TRAF3-dependent mechanism

TNF and RANKL mediate bone destruction in common bone diseases, including osteoarthritis and RA. They activate NF-kappaB canonical signaling directly in osteoclast precursors (OCPs) to induce osteoclast formation in vitro. However, unlike RANKL, TNF does not activate the alternative NF-kappaB pathway efficiently to process the IkappaB protein NF-kappaB p100 to NF-kappaB p52, nor does it appear to induce osteoclast formation in vivo in the absence of RANKL. Here, we show that TNF limits RANKL- and TNF-induced osteoclast formation in vitro and in vivo by increasing NF-kappaB p100 protein accumulation in OCPs. In contrast, TNF induced robust osteoclast formation in vivo in mice lacking RANKL or RANK when the mice also lacked NF-kappaB p100, and TNF-Tg mice lacking NF-kappaB p100 had more severe joint erosion and inflammation than did TNF-Tg littermates. TNF, but not RANKL, increased OCP expression of TNF receptor-associated factor 3 (TRAF3), an adapter protein that regulates NF-kappaB p100 levels in B cells. TRAF3 siRNA prevented TNF-induced NF-kappaB p100 accumulation and inhibition of osteoclastogenesis. These findings suggest that upregulation of TRAF3 or NF-kappaB p100 expression or inhibition of NF-kappaB p100 degradation in OCPs could limit bone destruction and inflammation-induced bone loss in common bone diseases.

Blockade of Dickkopf (DKK)-1 induces fusion of sacroiliac joints

ObjectiveTo study whether Dickkopf (DKK)-1, an inhibitor of wingless (Wnt) signalling, is involved in the fusion of sacroiliac joints.MethodsMice transgenic for tumour necrosis factor (TNFtg mice), which develop bilateral sacroiliitis,...

MRI and Quantification of Draining Lymph Node Function in Inflammatory Arthritis

While erosion and tissue necrosis are the end-stage result of inflammatory arthritis, factors that can predict their initiation and severity are unknown. In an effort to identify these prognostic factors we developed contrast-enhanced (CE)-magnetic resonance imaging (MRI) for the mouse knee to assess the pathogenesis of inflammatory arthritis. Using this approach to study synovitis and draining lymph node (LN) function we first demonstrated that the LNs of TNF-Tg mice at 5 months are significantly larger and have greater enhancement in comparison to wild-type (WT) mice. This difference correlated with the abundance of dilated LYVE-1+ sinuses in the draining LNs. Dynamic CE-MRI further demonstrated differences between TNF-Tg and WT mice in the kinetics of LN enhancement. We established an LN capacity (LNcap) measurement that is a function of both volume and CE. We demonstrated that TNF-Tg mice have a 15-fold increase over WT levels at 5 months age (P < 0.001). Amelioration of arthritis with anti-TNF therapy resulted in a significant decrease in LNcap (P < 0.0001) that approached WT levels within 4 weeks. Interestingly, this functional decrease was not associated with a reduction of lymphatic vessels, which persist after therapy in both LNs and synovium. To assess the relationship between draining LN function and synovitis, a regression analysis was performed that demonstrated a significant negative correlation (R(2) = 0.63, P = 0.01) between LNcap and synovial volume. TNF-Tg mice with a lower LNcap display an accelerated progression of arthritis. These results indicate a protective function of enhanced lymphatic drainage in inflammatory arthritis.

Increased lymphangiogenesis in joints of mice with inflammatory arthritis

Angiogenesis is involved in the pathogenesis of inflammatory arthritis, but little is known about the role of lymphangiogenesis in this setting. Here, we examined whether tumor necrosis factor (TNF) stimulates osteoclast precursors (OCPs) to produce the lymphatic growth factor, vascular endothelial growth factor-C (VEGF-C), and induce lymphangiogenesis. We used TNF-transgenic (Tg) mice and mice with serum-induced arthritis. OCPs were purified by fluorescence-activated cell sorting of CD11b+/Gr-1-/lo blood or bone marrow cells and subjected to microarray analysis or were generated from spleen or joint cells and treated with TNF. Expression of VEGFs was analyzed and examined by real-time reverse transcription-polymerase chain reaction and Western blotting. Immunostaining and magnetic resonance imaging were used to quantify lymphatic vessels and volumes of synovium and draining lymph nodes. TNF stimulated VEGF-C expression by OCPs and increased nuclear factor-kappa B (NF-κB) binding to an NF-κB sequence in the VEGF-C promoter. OCPs from joints of TNF-Tg mice express high levels of VEGF-C. Lymphatic vessel numbers and size were markedly increased in joint sections of TNF-Tg mice and mice with serum-induced arthritis. The severity of synovitis correlated with draining lymph node size. In summary, TNF induces OCPs to produce VEGF-C through NF-κB, leading to significantly increased lymphangiogenesis in joints of arthritic mice. The lymphatic system may play an important role in the pathogenesis of inflammatory arthritis.

Autoimmunity and Bone

Focal erosions of cartilage and bone, which occur in the joints of patients with autoimmune inflammatory arthritis (i.e., rheumatoid arthritis (RA) and psoriatic arthritis [PsA]), represent the most debilitating and irreversible components of the disease. Over the last decade, seminal breakthroughs in our understanding of the cells and signal transduction pathways central to this process have been elucidated. From this information an established paradigm has been developed to explain focal erosions in which osteoclasts responsible for erosions are derived from bone marrow-derived myeloid precursors. Using the tumor necrosis factor (TNF) transgenic mouse model of erosive arthritis and anti-TNF clinical trials with PsA patients, we have demonstrated that systemic TNF induces the migration of CD11b+ osteoclast precursors (OCP) from the bone marrow into peripheral blood. These OCP can then enter the joints in blood vessels, translocate across the receptor activator of NF-kappaB ligand (RANKL) rich inflamed synovium, and differentiate into active osteoclasts. In direct contrast to this, systemic lupus erythematosus (SLE) patients appear to have an innate resistance to bone resorption. Our hypothesis to explain this phenomenon is that systemic interferon-alpha (IFN-alpha) diverts the bone marrow-derived myeloid precursors away from the osteoclast lineage and stimulates their differentiation into dendritic cells (DC). In support of this model, several labs have used microarray analyses to define the IFN-induced transcriptome in peripheral blood mononuclear cells (PBMC) from SLE patients. Here we propose the hypothesis that systemic TNF induces osteoclastic differentiation of PBMC in PsA patients that correlates with their erosive disease, and that the innate immune TNF/IFN axis in patients with autoimmune disease dictates their erosive phenotype. To demonstrate this, we injected wild-type C57B/6 and TNF-Tg mice with poly I:C, which is known to induce systemic IFN responses, and show its dominant effects on increasing the number of circulating CD11b+/CD11c+ precursor dendritic cells (pDC), concomitant with a dramatic reduction in CD11b+/CD11c- OCP. Thus, systemic factors produced by autoimmunity have a dramatic impact on active myelopoiesis and bone homeostasis.

Tumor Necrosis Factor Promotes Runx2 Degradation through Up-regulation of Smurf1 and Smurf2 in Osteoblasts

Tumor necrosis factor (TNF) plays an important role in the pathogenesis of inflammatory bone loss through stimulation of osteoclastic bone resorption and inhibition of osteoblastic bone formation. Compared with the well established role of TNF in osteoclastogenesis, mechanisms by which TNF inhibits osteoblast function have not been fully determined. Runx2 is an osteoblast-specific transcription factor whose steady-state protein levels are regulated by proteasomal degradation, mediated by the E3 ubiquitin ligases, Smurf1 and Smurf2. We hypothesized that TNF inhibits osteoblast function through Smurf-mediated Runx2 degradation. We treated C2C12 and 2T3 osteoblast precursor cell lines and primary osteoblasts with TNF and found that TNF, but not interleukin-1, significantly increased Smurf1 and Smurf2 expression. TNF increased the degradation of endogenous or transfected Runx2 protein, which was blocked by treating cells with a proteasomal inhibitor or by infecting cells with small interfering (si)RNA against Smurf1 or Smurf2. TNF inhibited the expression of bone morphogenetic protein and transforming growth factor-beta signaling reporter constructs, and the inhibition of each was blocked by Smurf1 siRNA and Smurf2 siRNA, respectively. Overexpression of Smurf1 and/or Smurf2 siRNAs prevented the inhibitory effect of TNF on Runx2 reporter. Consistent with these in vitro findings, bones from TNF transgenic mice or TNF-injected wild type mice had increased Smurf1 and decreased Runx2 protein levels. We propose that one of the mechanisms by which TNF inhibits bone formation in inflammatory bone disorders is by promoting Runx2 proteasomal degradation through up-regulation of Smurf1 and Smurf2 expression.

RANK signaling is not required for TNFalpha-mediated increase in CD11(hi) osteoclast precursors but is essential for mature osteoclast formation in TNFalpha-mediated inflammatory arthritis.

To address the controversy of whether TNFalpha can compensate for RANKL in osteoclastogenesis in vivo, we used a TNFalpha-induced animal model of inflammatory arthritis and blocked RANKL/RANK signaling. TNFalpha increased osteoclast precursors available for RANK-dependent osteoclastogenesis. RANK signaling is not required for the TNFalpha-stimulated increase in CD11b(hi) osteoclast precursors but is essential for mature osteoclast formation. Although critical roles of TNFalpha in inflammatory arthritis and RANKL in bone resorption have been firmly established, a central controversy remains about the extent to which TNFalpha can compensate for RANKL during osteoclastogenesis and the stage at which RANK signaling is required for osteoclastogenesis. Here, we used the human TNFalpha transgenic mouse model (TNF-Tg) of erosive arthritis to determine if there are both RANK-dependent and -independent stages of osteoclastogenesis in TNFalpha-induced erosive arthritis. Osteoclastogenesis and osteoclast precursor (OCP) frequency were analyzed using histology, fluorescence-activated cell sorting (FACS), and cell culture from (1) TNF-Tg mice treated with the RANKL antagonist, RANK:Fc, or (2) TNF-Tg X RANK -/- mice generated by crossing TNF-Tg mice with RANK-/- mice. Treatment of TNF-Tg mice, which have increased OCPs in their spleens, with RANK:Fc dramatically reduced osteoclast numbers on the surface of their arthritic joints and within their bones, but did not decrease CD11b(hi) OCP numbers in their spleens. Long-term RANK:Fc administration alleviated joint erosion. Furthermore, TNF-Tg x RANK -/- mice had severe osteopetrosis, no osteoclasts, and no joint erosion, but increased CD11b(hi) precursor numbers that failed to form mature osteoclasts in vitro. RANK signaling is essential for mature osteoclast formation in TNFalpha-mediated inflammatory arthritis but not for the TNFalpha-induced increase in CD11b(hi) OCP that subsequently can differentiate into osteoclasts in inflamed joints.

The TNF-alpha transgenic mouse model of inflammatory arthritis.

Rheumatoid arthritis is a chronic inflammatory disorder that affects multiple peripheral joints. It is the most common form of inflammatory arthritis and is characterized by synovial hyperplasia, immune cell infiltration, cartilage destruction, and bone erosion. To gain insight into the etiology of the disease, a variety of animal models have been established. Twelve years ago George Kollias' laboratory generated a transgenic (Tg) mouse that over-expresses human TNF-alpha, and develops an erosive polyarthritis with many characteristics observed in rheumatoid arthritis patients. The phenotype of this mouse model validated the theory that TNF-alpha is at the apex of the pro-inflammatory cascade in rheumatoid arthritis, and foreshadowed the remarkable success of anti-TNF-alpha therapy that has transformed the effective management of this disease. As such, the TNF-Tg mice are very useful tools for dissecting the molecular mechanisms of the pathogenic process and evaluating the efficacy of novel therapeutic strategies for rheumatoid arthritis. In this review we (1) provide a brief summary of TNF-alpha biology and the role of this dominant cytokine in rheumatoid arthritis, (2) describe the various TNF-Tg models and their phenotypes, and (3) give examples of how this model has been used experimentally.

Endostatin gene transfer inhibits joint angiogenesis and pannus formation in inflammatory arthritis.

Rheumatoid arthritis is a prevalent example of an inflammatory angiogenic disease, which is mediated by pro-inflammatory and pro-angiogenic cytokines such as tumor necrosis factor (TNF). To evaluate the effect of the potent anti-angiogenic factor endostatin on TNF-induced inflammatory arthritis, we injected an endostatin-expressing lentiviral vector directly into the joints of human TNF-transgenic mice before the onset of disease. Histological analysis of the injected joints 8 weeks later revealed that endostatin reduced blood vessel density within the synovial tissues and an overall mean arthritis index. In vitro and in vivo examination of the potential mechanism by which endostatin inhibited the arthritis revealed that endostatin blocks TNF-induced activation of JNK and JNK-dependent pro-angiogenic gene expression. These data suggest a novel mechanism by which endostatin inhibits angiogenesis, and demonstrates the potential utility of anti-angiogenic gene therapy for treatment of inflammatory arthritis.