Selective enhancement of endothelial cell VCAM-1 expression by interleukin-10 in the presence of activated leucocytes.

Although initially described as an immunomodulatory cytokine, interleukin-10 (IL-10) has also been proposed to exert proinflammatory effects both in vivo and in vitro. In particular, studies in IL-10 transgenic mice have suggested that IL-10 may activate vascular endothelium to promote leucocyte adhesion and extravasation. In the present study we investigated whether IL-10 activates endothelial cells either directly or indirectly, via signals produced by leucocytes in the endothelial cell environment, using a co-culture of human umbilical vein endothelial cells and peripheral blood mononuclear cells (PBMC). No direct effects of IL-10 on endothelial cell responses were observed. However, in the presence of phytohaemagglutinin-activated PBMC, IL-10 increased the expression on endothelial cells of vascular cell adhesion molecule-1 (VCAM-1) but not of intercellular adhesion molecule-1, E-selectin or major histocompatibility complex (MHC) antigens, an effect mediated by PBMC-derived soluble factors. We also observed that interferon-gamma (IFN-gamma) antagonized VCAM-1 expression on endothelial cells mediated by IL-4 and IL-13. Since IL-10 has previously been documented to down-regulate release of IFN-gamma by PBMC, we propose that the IL-10-mediated reduction of IFN-gamma production by PBMC results in enhanced responsiveness of endothelial cells to PBMC-derived IL-4 and IL-13, and thus increased expression of VCAM-1. Our results suggest that the relative balance of cytokines produced by infiltrating cells in developing inflammatory lesions may differentially modulate endothelial responsiveness in vivo, and that IL-10 might indirectly stabilize VCAM-1 expression on endothelial cells by affecting the balance of leucocyte-derived cytokines in the endothelial environment.     

Treatment with soluble VEGF receptor reduces disease severity in murine collagen-induced arthritis

Maintenance of the invasive pannus in rheumatoid arthritis is an integral part of disease progression. The synovial vasculature plays an important role in the delivery of nutrients, oxygen, and inflammatory cells to the synovium. Vascular endothelial growth factor (VEGF), an endothelial mitogen expressed by cells within the synovial membrane, is thought to contribute to the formation of synovial blood vessels. Our objective in this study was to measure the kinetics of VEGF production in a murine model of collagen-induced arthritis and to determine whether VEGF blockade reduces disease progression. Synovial cells isolated from the knee joints of naive or sham-immunized mice, or from mice immunized with collagen but without arthritis, released little or no detectable VEGF. Onset of arthritis was associated with expression of VEGF mRNA and protein. The levels of VEGF secreted by synovial cells isolated from the joints of mice with severe arthritis were significantly higher than from mice with mild disease. To block VEGF activity, animals were treated after arthritis onset with a soluble form of the Flt-1 VEGF receptor (sFlt), which was polyethylene glycol (PEG)-linked to increase its in vivo half-life. Treatment of arthritic mice with sFlt-PEG significantly reduced both clinical score and paw swelling, compared with untreated or control-treated (heat-denatured sFlt-PEG) animals. There was also significantly less joint inflammation and reduced bone and cartilage destruction in sFlt-PEG-treated animals, as assessed by histology. Our data demonstrate that, in collagen-induced arthritis, expression of the potent angiogenic cytokine VEGF correlates with disease severity. Furthermore, specific blockade of VEGF activity results in attenuation of arthritis in both macroscopic and microscopic parameters. These observations indicate that blood vessel formation is integral to the development of arthritis and that blockade of VEGF activity might be of therapeutic benefit in rheumatoid arthritis.     

VEGF release is associated with reduced oxygen tensions in experimental inflammatory arthritis

OBJECTIVE: The contribution of local VEGF production and subsequent angiogenesis within the synovial membrane to the propagation of arthritis is unclear. The relationship between synovial oxygenation and blood flow in the development of arthritic disease is unknown. We have therefore measured oxygen levels and perfusion rates in the synovial space in a murine model of arthritis. METHODS: Arthritis was induced in DBA/1 mice by immunisation with type II collagen. Oxygen and perfusion levels were measured polarographically using silver needle microelectrodes within the knee joints prior to and 10 days after the onset of arthritis. In addition, synovial cells were isolated from knee joints of naive, pre-arthritic and arthritic mice. RESULTS: Onset of arthritis was associated with a marked reduction in synovial oxygen tensions (pO2). The perfusion rates in naive and arthritic animals were not significantly different: in naive mice, the rate was 0.58 +/- 0.11 ml/min/g and in arthritic joints, 0.64 +/- 0.17 ml/min/g. Furthermore, synovial cells isolated from the knee joints of naive animals did not express mRNA for VEGF, but significant levels were detected in cells from non-arthritic mice immunised with collagen. The onset of arthritis was associated with expression of VEGF mRNA and protein, and correlated negatively with pO2 levels. CONCLUSION: These data demonstrate that decreases in intra-articular pO2 occur in established arthritic conditions and may be the stimulus for local VEGF production. However, perfusion was not increased in arthritic animals and vascular density was unaltered, suggesting that the neovascularisation associated with inflammatory arthritis, is insufficient to restore oxygen homeostasis in the joint.     

Deactivation of vascular endothelium by monoclonal anti–tumor necrosis factor α antibody in rheumatoid arthritis

Objective. To assess whether monoclonal antibody to tumor necrosis factor α (TNFα) reduces endothelial activation in rheumatoid arthritis (RA). Methods. Levels of serum E-selectin, intercellular adhesion molecule 1 (ICAM-1), and vascular cell adhesion molecule 1 (VCAM-1), and circulating leukocytes (differential counts) were measured in RA patients before and up to 4 weeks after infusion of either placebo or chimeric anti-TNFα antibody cA2 (1 or 10 mg/kg). Results. Treatment with anti-TNFα decreased serum E-selectin and ICAM-1 levels, with the earliest detectable changes observed on days 1–3 after anti-TNFα infusion. No effect on VCAM-1 levels was detected. In parallel, there was a rapid and sustained increase in circulating lymphocytes. The extent of the decrease in serum E-selectin and ICAM-1 levels and the increase in lymphocyte counts was significantly higher (P ≤ 0.05) in patients in whom a clinical benefit of anti-TNFα was observed (≥20% response, by Paulus criteria, at week 4) compared with that in patients who failed to respond to anti-TNFα at this time point. Conclusion. We propose that decreased serum levels of adhesion molecules may reflect diminished activation of endothelial cells in the synovial microvasculature, leading to reduced migration of leukocytes into synovial joints, and thus prolonging the therapeutic effect of anti-TNFα in RA.     

Canonical pathway of nuclear factor kappa B activation selectively regulates proinflammatory and prothrombotic responses in human atherosclerosis

Nuclear factor kappa B (NF-kappa B) activation has been observed in human atherosclerotic plaques and is enhanced in unstable coronary plaques, but whether such activation has a protective or pathophysiological role remains to be determined. We addressed this question by developing a short-term culture system of cells isolated from human atherosclerotic tissue, allowing efficient gene transfer to directly investigate signaling pathways in human atherosclerosis. We found that NF-kappa B is activated in these cells and that this activity involves p65, p50, and c-Rel but not p52 or RelB. This NF-kappa B activation can be blocked by overexpression of I kappa B alpha or dominant-negative I kappa B kinase (IKK)-2 but not dominant-negative IKK-1 or NF-kappa B-inducing kinase, resulting in selective inhibition of inflammatory cytokines (tumor necrosis factor alpha, IL-6, and IL-8), tissue factor, and matrix metalloproteinases without affecting the antiinflammatory cytokine IL-10 or tissue inhibitor of matrix metalloproteinases. Our results demonstrate that the canonical pathway of NF-kappa B activation that involves p65, p50, c-Rel, and IKK-2 is activated in human atherosclerosis and results in selective up-regulation of major proinflammatory and prothrombotic mediators of the disease.     

Immunotherapy of rheumatoid arthritis: past, present and future

Rheumatoid arthritis (RA) is a chronic autoimmune disease, characterized by inflammation of the synovial lining of joints, and the destruction of cartilage and bone. Seminal studies demonstrating that pro-inflammatory cytokines, such as tumor necrosis factor-alpha (TNFalpha), are expressed in RA, has resulted in the approval of anti-TNFalpha biological therapies for its treatment. Although groundbreaking in themselves, these studies have also paved the way for further research to determine whether the targeting of other cytokines and immune pathways might aid in development of the next generation of drugs for the treatment of RA.     

The therapeutic potential of TNF-alpha blockade in rheumatoid arthritis

Rheumatoid arthritis is a chronic autoimmune disease characterised by inflammation of the synovial lining of joints and destruction of cartilage and bone. Many pro-inflammatory cytokines, chemokines and growth factors are expressed in diseased joints, and recognition of the key role of TNF-alpha led to the development of highly effective new therapies. TNF-alpha inhibitors, such as monoclonal anti-TNF-alpha antibody infliximab (Remicade), have demonstrated efficacy in clinical trials. It is now clear that TNF-alpha blockade, in addition to reducing joint inflammation and leukocyte infiltration, also results in decreased formation of new blood vessels in the synovium. Such mechanism of action studies are now paving the way for the development of the next generation of drugs for treatment of rheumatoid arthritis.