Elsevier

Immunology Letters

Volume 103, Issue 2, 15 March 2006, Pages 159-166
Immunology Letters

Interleukin-18 induces the production of vascular endothelial growth factor (VEGF) in rheumatoid arthritis synovial fibroblasts via AP-1-dependent pathways

https://doi.org/10.1016/j.imlet.2005.10.020Get rights and content

Abstract

Interleukin-18 (IL-18) is a novel pro-inflammatory cytokine which has been implicated to play a pathogenic role in rheumatoid arthritis (RA). Vascular endothelial growth factor (VEGF) plays an important role in angiogenesis in rheumatoid synoviocytes. In present study, we examined the effect of IL-18 on VEGF production in fibroblast-like synoviocytes (FLS) isolated from the patients with RA. FLS were prepared from the synovial tissues of patients with RA and osteoarthritis (OA) and cultured in the presence of IL-18. The production of VEGF from FLS was measured in culture supernatants by enzyme-linked immunosorbent assay (ELISA). The VEGF messenger RNA (mRNA) expression and AP-1 binding activity of VEGF transcript were determined by reverse transcription-polymerase chain reaction (RT-PCR) and electrophoretic mobility shift assay (EMSA). IL-18 and VEGF levels of sera and synovial fluids (SF) of RA patients (n = 30) were significantly higher than those of OA patients (n = 20). IL-18 dose-dependently increased the production of VEGF. The effect of IL-18 on VEGF production appeared to be as potent as IL-1β, whereas tumor necrosis factor (TNF)-α and interferon (IFN)-γ showed little effects on VEGF production. AP-1-specific inhibitor Curcumin dose-dependently abrogated the effect of IL-18 on VEGF production. The VEGF enhancement of IL-18 was associated with increased AP-1 binding activity to the VEGF promoter site. These findings suggest IL-18 as an angiogenic factor in RA and down-regulation of IL-18 activity or AP-1 signal pathway can be potential therapeutic targets for RA.

Introduction

Rheumatoid arthritis (RA) is a systemic autoimmune disease with chronic inflammation characterized by hyperplasia of synovial cells and angiogenesis in affected joints, which ultimately leads to the destruction of cartilage and bone [1]. Neovascularization, the formation of new blood vessels, can maintain the chronic inflammatory status by transporting the inflammatory cells to the site of synovitis as well as supplying nutrients and oxygen to pannus. A number of angiogenic factors have been investigated for their significances of neovascularization in RA joints. These include acidic and basic fibroblast growth factors, platelet-derived endothelial cell growth factor and transforming growth factor (TGF)-α, TGF-β, angiogenin, and vascular endothelial growth factor (VEGF) [2], [3].

VEGF is a heparin binding, dimeric glycoprotein that induces the proliferation and migration of endothelial cells to form new vessels, and increases the penetration and extravagation of plasma macromolecules [4], [5]. VEGF has shown to play an important role in wound healing [6], embryonic development [7], growth of certain solid tumors, and ascites formation [8]. Recently several reports demonstrated that VEGF was also implicated in the pathogenesis of RA. VEGF level was significantly increased in synovial fluids of RA patients than in those in osteoarthritis or other form of arthritis [9], [10]. VEGF polypeptide and messenger RNA (mRNA) were expressed on subsynovial macrophages and synovial lining cells in the synovial tissues of RA patients [9], [11]. Cultured synovial cells are also known to express VEGF under hypoxic conditions [12], [13], [14] or when stimulated with interleukin (IL)-1, IL-6 [15], [16], prostaglandin (PG) [15], TGF-β [13], [17], and CD40 ligation [18]. Furthermore, treatment with a soluble form of the Flt-1 VEGF receptor significantly attenuated disease severity in murine collagen-induced arthritis (CIA) [19].

IL-18 is a novel immunoregulatory cytokine with pleiotropic activities, which is known to involve in the development of type I helper T cell (Th1) responses [20]. This cytokine is structurally similar to IL-1β, and considered as a member of IL-1 family [21]. IL-18 up-regulates the expression of IL-2Rα, and promotes interferon (IFN)-γ, tumor necrosis factor (TNF)-α, and graulocyte macrophage colony-stimulating factor (GM-CSF) production by activation of Th1 clones [22], [23]. IL-18 was detected in RA synovial membrane in macrophages together with lining layer of fibroblasts, and the level of IL-18 was increased in the joint and in the serum of RA patients [24]. IL-18 promoted production of IFN-γ, TNF-α, GM-CSF, and CXC chemokines from fibroblast-like synoviocytes (FLS) [24].

While IL-18 has been implicated in RA pathogenesis, its spectrum of action that promotes inflammation and angiogenesis in the synovial tissue has not been completely elucidated. Park et al. suggested IL-18 as a novel angiogenic mediator in RA [25]. On the contrary, IL-18 has been shown to inhibit angiogenesis and tumor growth in other animal models [26], [27]. Recent studies have indicated that IL-18 might act as either as angiogenic or angiostatic factor, but the true function of this protein in angiogenesis is unclear.

In this study, we investigated the role of IL-18 on the VEGF production in RA synovial fibroblasts. Our study suggests that IL-18 could be an angiogenic mediator with triggering effect of VEGF production in patients with RA.

Section snippets

Patients and controls

Informed consent was obtained from 30 patients (6 men and 24 women) with RA, who fulfilled the 1987 revised criteria of the American College of Rheumatology [28]. The mean age of the RA patients was 50 ± 8 years (range 23–71 years). All medications were stopped 48 h before study entry. Comparisons were made with 20 patients with osteoarthritis (OA) (4 men and 16 women) and with 20 healthy controls (4 men and 16 women) who had no rheumatic diseases. The mean age of the OA patients and healthy

Increase of IL-18 and VEGF level in synovial fluids of RA patients

VEGF and IL-18 in serum and SF were examined from the patients with RA (n = 30), and compared with osteoarthritis (OA, n = 20) patients and in normal controls (n = 20) (Fig. 1). We measured the levels of VEGF and IL-18 in serum and synovial fluid simultaneously. VEGF (RA 1216 ± 211 pg/ml, OA 434 ± 131 pg/ml, normal 335 ± 146 pg/ml) and IL-18 (RA 442 ± 106 pg/ml, OA 201 ± 51 pg/ml, normal 135 ± 63 pg/ml) titers in serum of RA were higher than those of OA and normal controls (Fig. 1A and B). The mean levels of VEGF in

Discussions

In this study, we examined the potential angiogenic role of IL-18 in FLS of RA patients. We found evidence that IL-18 directly increased the production of VEGF by RA FLS, which is a novel function of IL-18. VEGF have been reported to be a participant in synovitis [9], [10], [11], by promoting new vessel formation and vascular permeability [4]. Provided that angiogenesis is a key process of synovial inflammation of RA, factors that regulate VEGF production by synovial cells are of great

Acknowledgement

This work was supported by Grant (R11-2002-098-05001-0) from the Korea Science and Engineering Foundation through the Rheumatism Research Center at Catholic University of Korea.

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