Additive bone‐protective effects of anabolic treatment when used in conjunction with RANKL and tumor necrosis factor inhibition in two rat arthritis models

Objective

To investigate whether the bone‐preserving effects of a RANKL antagonist or a tumor necrosis factor (TNF) antagonist could be further improved by the addition of a bone anabolic agent in inflammatory arthritis.

Methods

Lewis rats with either adjuvant‐induced arthritis (AIA) or collagen‐induced arthritis (CIA) were treated for 10 days with PEGylated soluble tumor necrosis factor receptor type I (PEG sTNFRI), interleukin‐1 receptor antagonist (IL‐1Ra), osteoprotegerin (OPG), parathyroid hormone (PTH), or combinations of these agents starting on day 4 after disease onset. Treatment effects were assessed clinically, radiologically, and histologically, and by morphometry for the extent of paw swelling, bone erosive changes, and synovial inflammation.

Results

Paw swelling and synovial inflammation were significantly inhibited by PEG sTNFRI in AIA and CIA, and by IL‐1Ra in CIA. OPG and PTH had no significant effect on these parameters. Analysis of bone erosion revealed a significant bone‐sparing effect of monotherapy with PEG sTNFRI or OPG in both models, whereas IL‐1Ra was only effective in CIA. PTH treatment alone did not show a bone‐protective effect in either model. With the combination of PEG sTNFRI and PTH, erosion scores (−74% in AIA and −61% in CIA versus controls) were significantly lower than those elicited by PEG sTNFRI alone (−41% and −29%, respectively, versus controls). Similar results were also obtained with the combination of OPG and PTH (−88% in AIA and −73% in CIA, compared with −70% and −55%, respectively, with OPG monotherapy). Coadministration of IL‐1Ra and PTH had no synergistic bone‐sparing effect. Morphometric analysis revealed that the addition of PTH to PEG sTNFRI or OPG resulted in higher bone volume and higher osteoblast numbers in both AIA and CIA.

Conclusion

The bone‐protective effects resulting from RANKL or TNF antagonism can be further improved by the addition of a bone anabolic agent.

     

Inhibition of interleukin‐6 receptor directly blocks osteoclast formation in vitro and in vivo

Objective

To investigate the efficacy of a murine anti–interleukin‐6 receptor (anti–IL‐6R) antibody in directly blocking tumor necrosis factor (TNF)– and RANKL‐mediated osteoclastogenesis in vitro and in vivo.

Methods

The efficacy of a murine antibody against IL‐6R in blocking osteoclast differentiation of mononuclear cells stimulated with RANKL was tested. In addition, arthritic human TNFα–transgenic mice were treated with anti–IL‐6R antibody, and osteoclast formation and bone erosion were assessed in arthritic paws.

Results

Blockade of IL‐6R dose dependently reduced osteoclast differentiation and bone resorption in monocyte cultures stimulated with RANKL or RANKL plus TNF. In human TNFα–transgenic mice, IL‐6R blockade did not inhibit joint inflammation, but it strongly reduced osteoclast formation in inflamed joints as well as bone erosion in vivo. Neither the cell influx into joints nor the synovial expression of IL‐6 and RANKL changed with IL‐6R blockade, while the synovial expression of IL‐1 was significantly reduced. In contrast, TNF‐mediated systemic bone loss was not inhibited by IL‐6R blockade.

Conclusion

These data suggest that blockade of IL‐6R directly affects osteoclast formation in vitro and in vivo, suggesting a direct and specific effect of anti–IL‐6R therapy on osteoclasts independently of its antiinflammatory effects. This effect adds significantly to the structure‐sparing potential of pharmacologic blockade of IL‐6R in arthritis.

     

Enhanced Th1 and Th17 responses and arthritis severity in mice with a deficiency of myeloid cell–specific interleukin‐1 receptor antagonist

Objective

The balance between interleukin‐1 (IL‐1) and its specific inhibitor, the IL‐1 receptor antagonist (IL‐1Ra), plays a major role in the development of arthritis. The purpose of this study was to investigate the role of IL‐1Ra produced specifically by myeloid cells in the control of collagen‐induced arthritis (CIA) by using myeloid cell–specific IL‐1Ra–deficient mice (IL‐1Ra^ΔM).

Methods

IL‐1Ra^ΔM mice were generated by using the loxP/Cre recombinase system. CIA was induced in IL‐1Ra^ΔM mice and littermate control mice by a single immunization with bovine type II collagen (CII) in Freund's complete adjuvant. Arthritis severity was assessed by clinical and histologic scoring. Draining lymph node (DLN) cell responses were examined ex vivo, and ankle extracts were used in the quantification of cytokines and chemokines.

Results

Clinical and histopathologic evaluations revealed an early disease onset and a severe form of CIA in IL‐1Ra^ΔM mice. This was characterized by increased production of interferon‐γ (IFNγ) and IL‐17 by CII‐stimulated DLN cells. We also observed that the CII‐specific CD4+ T cell response shifted in vivo, from a dominant Th1 response early in the course of the arthritis to the presence of both Th1 and Th17 cytokines later in the disease course. Interestingly, IL‐1Ra levels were higher in the arthritic joints of IL‐1Ra^ΔM mice as compared with the controls, indicating that nonmyeloid cells strongly contribute to the local production of IL‐1Ra. However, this enhanced IL‐1Ra production was not sufficient to limit joint inflammation and tissue damage.

Conclusion

Our results suggest that myeloid cell–derived IL‐1Ra plays a critical role in the control of the development and the severity of CIA by modulating Th1 and Th17 responses in lymphoid organs.

     

CD28‐dependent differentiation into the effector/memory phenotype is essential for induction of arthritis in interleukin‐1 receptor antagonist–deficient mice

Objective

Interleukin‐1 receptor antagonist (IL‐1Ra)–deficient mice on a BALB/c background spontaneously develop a chronic inflammatory polyarthropathy closely resembling that of rheumatoid arthritis in humans. To elucidate the role of CD28 costimulatory signals in the development of this disease, we studied IL‐1Ra/CD28–double‐deficient mice.

Methods

We crossed IL‐1Ra–deficient mice with CD28–deficient mice and observed the incidence and severity of arthritis. To investigate functions of IL‐1Ra/CD28–double‐deficient T cells, cells were stimulated with CD3 monoclonal antibody or allogeneic antigen‐presenting cells (APCs) and their proliferative responses and levels of cytokine production were measured.

Results

Disease severity was lower in IL‐1Ra/CD28–double‐deficient mice than in mice that were deficient only in IL‐1Ra, although incidence of arthritis was not affected by the presence or absence of CD28. When pathogenic IL‐1Ra–KO T cells were transferred into nude mice, severe arthritis developed. Even though T cells from double‐deficient mice showed the same diminished proliferative capacity as was seen in T cells from CD28–single‐deficient animals, nude mice into which double‐deficient T cells were transferred never developed arthritis.

Conclusion

These findings indicate that IL‐1Ra/CD28–double‐deficient T cells can be activated by IL‐1Ra–deficient activated APCs, resulting in induction of arthritis; however, these T cells did not induce the disease under normal conditions, because they did not differentiate into effector/memory phenotype.

     

Dysregulation of interleukin‐10–dependent gene expression in rheumatoid arthritis synovial macrophages

Objective

The inflammatory cytokines tumor necrosis factor α (TNFα) and interleukin‐1 (IL‐1) are produced by activated macrophages, are key mediators of pathogenesis, and are validated therapeutic targets in rheumatoid arthritis (RA) and seronegative spondylarthritis (SpA). IL‐10 is a potent antiinflammatory cytokine that suppresses macrophage TNFα and IL‐1 production, yet is not effective in suppressing inflammatory arthritis. To gain insight into IL‐10 responses in inflammatory arthritis, we used microarray analysis to determine the patterns of IL‐10–inducible gene expression in freshly isolated RA and seronegative SpA synovial macrophages.

Methods

Macrophages from the synovial fluid of 5 patients with RA and 3 with seronegative SpA (2 with psoriatic arthritis and 1 with ankylosing spondylitis) were isolated by positive selection and stimulated ex vivo with IL‐10 or interferon‐γ (IFNγ). Gene expression was analyzed using Affymetrix microarrays and protocols. Real‐time polymerase chain reaction was used to confirm changes in gene expression.

Results

The number of genes induced by IL‐10 in arthritic macrophages was markedly smaller than that induced in control macrophages, and the strength of induction was lower in arthritic macrophages for most genes. The residual response of arthritic macrophages to IL‐10 stimulation was qualitatively altered, such that IL‐10 preferentially increased expression of IFNγ‐inducible genes. In contrast, arthritic macrophages expressed many IFNγ‐inducible genes prior to stimulation, and their response to IFNγ remained mostly intact.

Conclusion

These results demonstrate that IL‐10 responses are dysregulated in RA synovial macrophages. An altered biologic response to IL‐10, with attenuation of its antiinflammatory function and a concomitant retention of IFNγ‐like activating functions, provides a basis for the lack of efficacy of IL‐10 in suppressing inflammatory arthritis.

     

Soluble interleukin‐1 receptor accessory protein ameliorates collagen‐induced arthritis by a different mode of action from that of interleukin‐1 receptor antagonist

Objective

To discern the mode of interleukin‐1 (IL‐1) inhibition of soluble IL‐1 receptor accessory protein (sIL‐1RAcP) by comparison with IL‐1 receptor antagonist (IL‐1Ra) in arthritis.

Methods

Adenoviral vectors encoding either sIL‐1RAcP or IL‐1Ra were administered systemically before onset of collagen‐induced arthritis in DBA/1 mice. Anti–bovine type II collagen IgG and IL‐6 were quantified in serum. Proliferative response of splenic T cells was determined in the presence of sIL‐1RAcP or IL‐1Ra. The effect on IL‐1 inhibition of recombinant sIL‐1RAcP and IL‐1Ra was further examined in vitro, using NF‐κB luciferase reporter cell lines. Quantitative polymerase chain reaction was used to determine the relative messenger RNA expression of the IL‐1 receptors.

Results

Adenoviral overexpression of both sIL‐1RAcP and IL‐1Ra resulted in amelioration of the collagen‐induced arthritis. Both IL‐1 antagonists reduced the circulating levels of antigen‐specific IgG2a antibodies, but only IL‐1Ra was able to inhibit lymphocyte proliferation. By using purified lymphocyte populations derived from NF‐κB reporter mice, we showed that sIL‐1RAcP inhibits IL‐1–induced NF‐κB activity in B cells but not T cells, whereas IL‐1Ra inhibited IL‐1 on both cell types. A study in a panel of NF‐κB luciferase reporter cells showed that the sIL‐1RAcP inhibits IL‐1 signaling on cells expressing either low levels of membrane IL‐1RAcP or high levels of IL‐1RII.

Conclusion

We show that the sIL‐1RAcP ameliorated experimental arthritis without affecting T cell immunity, in contrast to IL‐1Ra. Our results provide data in support of receptor competition by sIL‐1RAcP as an explanation for the different mode of IL‐1 antagonism in comparison with IL‐1Ra.

     

Blockade of NKG2D ameliorates disease in mice with collagen‐induced arthritis: A potential pathogenic role in chronic inflammatory arthritis

Objective

To assess the role of the activating receptor NKG2D in arthritis.

Methods

Levels of NKG2D and its ligands were determined by fluorescence‐activated cell sorting, real‐time polymerase chain reaction, and immunohistochemistry in rheumatoid arthritis (RA) synovial membrane tissue and in paw tissue from arthritic mice. Arthritis was induced in DBA/1 mice by immunization with type II collagen, and mice were treated intraperitoneally with a blocking anti‐NKG2D antibody (CX5) on days 1, 5, and 8 after clinical onset and were monitored for 10 days.

Results

We demonstrated expression of NKG2D and its ligands on human RA synovial cells and extended this finding to the paws of arthritic mice. Expression of messenger RNA for the NKG2D ligand Rae‐1 was up‐regulated, and NKG2D was present predominantly on natural killer (NK) and CD4+ T cells, in arthritic paw cell isolates. NKG2D was down‐modulated during the progression of collagen‐induced arthritis (CIA). NKG2D expression in arthritic paws was demonstrated by immunohistochemistry. Blockade of NKG2D ameliorated established CIA, with significant reductions in clinical scores and paw swelling. Histologic analysis of arthritic joints from anti‐NKG2D–treated mice demonstrated significant joint protection, compared with control mice. Moreover, anti‐NKG2D treatment significantly reduced both interleukin‐17 production from CD4+ T cells in arthritic paws and splenic NK cell cytotoxic effector functions in vivo and in vitro.

Conclusion

Our findings indicate that blockade of NKG2D in a murine model and in human explants has beneficial therapeutic potential that merits further investigation in RA.

     

Enhanced integrative repair of the porcine meniscus in vitro by inhibition of interleukin‐1 or tumor necrosis factor α

Objective

To examine the hypotheses that increasing concentrations of interleukin‐1 (IL‐1) or tumor necrosis factor α (TNFα) inhibit the integrative repair of the knee meniscus in an in vitro model system, and that inhibitors of these cytokines will enhance repair.

Methods

Explants (8 mm in diameter) were harvested from porcine medial menisci. To simulate a full‐thickness defect, a 4‐mm–diameter core was removed and reinserted. Explants were cultured for 14, 28, or 42 days in the presence of 0–1,000 pg/ml of IL‐1 or TNFα. Explants were also cultured in the presence of IL‐1 or TNFα with IL‐1 receptor antagonist (IL‐1Ra) or TNF monoclonal antibody (mAb). At the end of the culture period, biomechanical testing, cell viability, and histologic analyses were performed to quantify the extent of repair.

Results

Mechanical testing revealed increased repair strength, cell accumulation, and tissue formation at the interface over time under control conditions. Pathophysiologic concentrations of both IL‐1 and TNFα significantly decreased repair strength, cell migration, and tissue formation at the interface. The addition of IL‐1Ra or TNF mAb to explants prevented the effects of IL‐1 or TNFα, respectively.

Conclusion

Our findings document that physiologically relevant concentrations of IL‐1 and TNFα inhibit meniscal repair in vitro and therefore may also inhibit meniscal repair during arthritis or following joint injury. The finding that IL‐1Ra and TNF mAb promoted integrative meniscal repair in an inflammatory microenvironment suggests that intraarticular delivery of IL‐1Ra and/or TNF mAb may be useful clinically to promote meniscal healing following injury.

     

Intraarticular injection of platelet‐rich plasma reduces inflammation in a pig model of rheumatoid arthritis of the knee joint

Objective

Treatment options for rheumatoid arthritis range from symptomatic approaches to modern molecular interventions such as inhibition of inflammatory mediators. Inhibition of inflammation by platelet‐rich plasma (PRP) has been proposed as a treatment for tendinitis and osteoarthritis. The present study was undertaken to investigate the effect of PRP on antigen‐induced arthritis (AIA) of the knee joint in a large animal model.

Methods

Six‐month‐old pigs (n = 10) were systemically immunized by bovine serum albumin (BSA) injection, and arthritis was induced by intraarticular BSA injection. PRP was injected into the knee joints of 5 of the animals after 2 weeks. An additional 5 animals received no systemic immunization (controls). Signs of arthritis were documented by plain histologic analysis, Safranin O staining, and immunohistochemistry analysis for type II collagen (CII), interleukin‐6 (IL‐6), and vascular endothelial growth factor (VEGF). Interleukin‐1β (IL‐1β), IL‐6, tumor necrosis factor α (TNFα), VEGF, and insulin‐like growth factor 1 (IGF‐1) protein content was measured by Luminex assay.

Results

In the pigs with AIA, plain histologic analysis revealed severe arthritic changes in the synovium. Safranin O and CII staining showed decreased proteoglycan and CII content in cartilage. Immunohistochemistry analysis revealed increased levels of IL‐6 and VEGF in synovium and cartilage, and protein concentrations of IL‐6, VEGF, IL‐1β, and IGF‐1 in synovium and cartilage were elevated as well; in addition, TNFα protein was increased in cartilage. Treatment with PRP led to attenuation of these arthritic changes in the synovium and cartilage.

Conclusion

We have described a porcine model of AIA. Experiments using this model demonstrated that PRP can attenuate arthritic changes as assessed histologically and based on protein synthesis of typical inflammatory mediators in the synovial membrane and cartilage.

     

Single and combined inhibition of tumor necrosis factor,interleukin‐1, and RANKL pathways in tumor necrosis factor–induced arthritis: Effects on synovial inflammation,bone erosion,and cartilage destruction

Objective

To investigate the efficacy of single and combined blockade of tumor necrosis factor (TNF), interleukin‐1 (IL‐1), and RANKL pathways on synovial inflammation, bone erosion, and cartilage destruction in a TNF‐driven arthritis model.

Methods

Human TNF–transgenic (hTNFtg) mice were treated with anti‐TNF (infliximab), IL‐1 receptor antagonist (IL‐1Ra; anakinra), or osteoprotegerin (OPG; an OPG‐Fc fusion protein), either alone or in combinations of 2 agents or all 3 agents. Synovial inflammation, bone erosion, and cartilage damage were evaluated histologically.

Results

Synovial inflammation was inhibited by anti‐TNF (−51%), but not by IL‐1Ra or OPG monotherapy. The combination of anti‐TNF with either IL‐1Ra (−91%) or OPG (−81%) was additive and almost completely blocked inflammation. Bone erosion was effectively blocked by anti‐TNF (−79%) and OPG (−60%), but not by IL‐1Ra monotherapy. The combination of anti‐TNF with IL‐1Ra, however, completely blocked bone erosion (−98%). Inhibition of bone erosion was accompanied by a reduction of osteoclast numbers in synovial tissue. Cartilage destruction was inhibited by anti‐TNF (−43%) and was weakly, but not significantly, inhibited by IL‐1Ra, but was not inhibited by OPG monotherapy. The combination of anti‐TNF with IL‐1Ra was the most effective double combination therapy in preventing cartilage destruction (−80%). In all analyses, the triple combination of anti‐TNF, IL‐1Ra, and OPG was not superior to the double combination of anti‐TNF and IL‐1Ra.

Conclusion

Articular changes caused by chronic overexpression of TNF are not completely blockable by monotherapies that target TNF, IL‐1, or RANKL. However, combined approaches, especially the combined blockade of TNF and IL‐1 and, to a lesser extent, TNF and RANKL, lead to almost complete remission of disease. Differences in abilities to block synovial inflammation, bone erosion, and cartilage destruction further strengthen the rationale for using combined blockade of more than one proinflammatory pathway.

     

Lack of response to anakinra in rheumatoid arthritis following failure of tumor necrosis factor α blockade

Objective.

In patients with rheumatoid arthritis (RA) treated with tumor necrosis factor α (TNFα)– blocking therapy, there is heterogeneity of response. This raises the possibility that in certain circumstances, cytokines such as interleukin‐1 (IL‐1) may dominate the drive toward joint inflammation. This study was undertaken to investigate whether blocking the action of IL‐1 with an IL‐1 receptor antagonist (IL‐1Ra) is efficacious in patients with disease that did not respond to TNFα blockade.

Methods.

We identified 26 RA patients whose disease had failed to respond to TNFα‐blocking therapy, defined as failure to achieve or sustain a 20% improvement in disease activity according to the criteria of the American College of Rheumatology (ACR20 response). These patients were then treated with anakinra (100 mg/day subcutaneously) for 12 weeks, and their levels of response were assessed.

Results.

After 3 months of anakinra therapy, only 2 of 26 patients (8%) achieved an ACR20 response; none achieved an ACR50 or ACR70 response. A rise in the mean C‐reactive protein level and an increase in the mean swollen joint count were noted during the study period.

Conclusion.

This study demonstrates that patients with disease that fails to respond to TNFα blockade also do not respond to IL‐1Ra. These data do not provide evidence of a dominant role for IL‐1 in patients who do not respond to TNFα blockade, but they do not exclude a role for other proinflammatory mediators.

     

Inhibition of toll‐like receptor 4 breaks the inflammatory loop in autoimmune destructive arthritis

Objective

Degeneration of extracellular matrix of cartilage leads to the production of molecules capable of activating the immune system via Toll‐like receptor 4 (TLR‐4). The objective of this study was to investigate the involvement of TLR‐4 activation in the development and progression of autoimmune destructive arthritis.

Methods

A naturally occurring TLR‐4 antagonist, highly purified lipopolysaccharide (LPS) from Bartonella quintana, was first characterized using mouse macrophages and human dendritic cells (DCs). Mice with collagen‐induced arthritis (CIA) and mice with spontaneous arthritis caused by interleukin‐1 receptor antagonist (IL‐1Ra) gene deficiency were treated with TLR‐4 antagonist. The clinical score for joint inflammation, histologic characteristics of arthritis, and local expression of IL‐1 in joints were evaluated after treatment.

Results

The TLR‐4 antagonist inhibited DC maturation induced by Escherichia coli LPS and cytokine production induced by both exogenous and endogenous TLR‐4 ligands, while having no effect on these parameters by itself. Treatment of CIA using TLR‐4 antagonist substantially suppressed both clinical and histologic characteristics of arthritis without influencing the adaptive anti–type II collagen immunity crucial for this model. Treatment with TLR‐4 antagonist strongly reduced IL‐1β expression in articular chondrocytes and synovial tissue. Furthermore, such treatment inhibited IL‐1–mediated autoimmune arthritis in IL‐1Ra^−/− mice and protected the mice against cartilage and bone pathology.

Conclusion

In the present study, we demonstrate for the first time that inhibition of TLR‐4 suppresses the severity of experimental arthritis and results in lower IL‐1 expression in arthritic joints. Our data suggest that TLR‐4 might be a novel target in the treatment of rheumatoid arthritis.

     

Arthritis in mice that are deficient in interleukin‐1 receptor antagonist is dependent on genetic background

Objective

To determine the effect of deletion of interleukin‐1 receptor antagonist (IL‐1Ra) protein in an animal model of rheumatoid arthritis.

Methods

BALB/c mice deficient in IL‐1Ra (IL‐1Ra^−/−) were bred with collagen‐induced arthritis (CIA)–susceptible DBA/1 mice and B10 mice transgenic for HLA–DRB1*0101 (B10.DR1). After generation of IL‐1Ra^−/− mice on the DBA/1 and B10.DR1 backgrounds, the mice were observed for the development of spontaneous arthritis and immunized for induction of CIA.

Results

We found that although BALB/c mice deficient in IL‐1Ra (BALB/c^−/−) spontaneously developed chronic inflammatory arthritis, DBA/1 IL‐1Ra–deficient (DBA/1^−/−) and B10.DR1 IL‐1Ra–deficient (B10.DR1^−/−) mice did not. Splenocytes from BALB/c^−/− mice produced elevated levels of IL‐2, IL‐4, IL‐6, IL‐10, IL‐17, and granulocyte–macrophage colony‐stimulating factor in response to anti‐CD3 stimulation. After immunization with type II collagen (CII), DBA/1^−/− and B10.DR1^−/− mice had a significantly earlier onset of CIA, and with increased severity compared with IL‐1Ra^+/+ mice. Immunization of BALB/c^−/− mice with CII did not aggravate spontaneous arthritis. All of the immunized mice developed antibodies to CII that correlated with arthritis severity. Levels of antibody to CII in the BALB/c^−/− strain were relatively low.

Conclusion

These data indicate that the spontaneous arthritis of IL‐1Ra deficiency is highly dependent on non–major histocompatibility complex genes and that autoimmunity to CII is not the major disease‐inducing event. Class II immune response genes are more important for the regulation of CIA, and although these 2 models of arthritis share many pathogenic mechanisms, they also have significant differences.

     

Gene therapy targeting the Tie2 function ameliorates collagen‐induced arthritis and protects against bone destruction

Objective

In a previous study, we demonstrated that Tie2 regulates angiogenesis in arthritis. The current study was performed to determine whether systemic delivery of a soluble Tie2 receptor (ExTek) using an adenoviral vector (AdExTek) as a Tie2 inhibitor affects arthritis development and progression in an animal model.

Methods

We used a collagen‐induced arthritis (CIA) mouse model to study the outcome of treatment with either AdExTek or a control vector. The onset, incidence, and severity of arthritis were quantified. Immunohistologic analysis of endothelium obtained from the paws was performed. Bone destruction in paws was analyzed using phase‐contrast radiography.

Results

The data showed that systemic delivery of ExTek before disease development significantly inhibited the onset, incidence, and severity of arthritis. When AdExTek was given after disease onset, the severity of disease in mice treated with AdExTek was significantly lower than that in the control group at 35 days postimmunization, which correlated with significantly diminished angiogenesis in mouse paws. Strikingly, AdExTek treatment protected bone from erosion in the CIA model and reduced levels of RANKL. No differences in collagen‐specific antibodies were detected between these 2 groups.

Conclusion

We demonstrated that blocking Tie2 receptor activation inhibits angiogenesis and arthritis development and protects against bone destruction in a CIA mouse model. These findings identify Tie2 as a therapeutic target for arthritis treatment and imply that interventions designed to target the Tie2 pathway could be clinically beneficial.

     

Immunolocalization of interleukin‐1 receptors in the sarcolemma and nuclei of skeletal muscle in patients with idiopathic inflammatory myopathies

Objective

Interleukin‐1 (IL‐1) acts via its receptors to induce gene expression that mediates protein synthesis involved in inflammation. Increased expression of IL‐1α and IL‐1β in muscle tissue from patients with polymyositis and dermatomyositis has been demonstrated. It is not known whether the reciprocal IL‐1 receptors are expressed in human muscle tissue. The purpose of this study was to investigate the expression of IL‐1 receptors and their ligands in muscle tissue from patients with myositis and from healthy controls.

Methods

Muscle biopsy tissues from 10 patients with polymyositis or dermatomyositis and 7 healthy control subjects were investigated by immunohistochemistry using antibodies against IL‐1 receptor type I (IL‐1RI), IL‐1RII, IL‐1α, IL‐1β, and IL‐1 receptor antagonist (IL‐1Ra). Quantification was performed by computerized image analysis, and localization of expression was determined by double staining using immunofluorescence and confocal microscopy.

Results

In tissue samples from the patients, IL‐1RI and IL‐1RII were expressed in muscle fibers, inflammatory cells, and endothelial cells. Expression in muscle fibers was localized to the sarcolemma and nuclei. IL‐1α was expressed in endothelial cells and inflammatory cells, whereas IL‐1β and IL‐1Ra were expressed only in inflammatory cells. Expression of the two IL‐1 receptors and their ligands was significantly higher in patients than in controls. IL‐1 receptor expression on muscle fibers was most pronounced in the vicinity of cells expressing IL‐1α and IL‐1β.

Conclusion

The increased expression of IL‐1 receptor and the colocalization with reciprocal ligands in patients with myositis but not in healthy controls support the hypothesis of a crucial role of IL‐1 in the pathogenesis of polymyositis and dermatomyositis.