Regular aticle
The role of COX-2 in angiogenesis and rheumatoid arthritis☆

https://doi.org/10.1016/S0014-4800(03)00019-4Get rights and content

Abstract

Recent evidence suggests that cyclooxygenase (COX)-2 is a mediator of angiogenesis, and COX-2 activity is known to be upregulated in the rheumatoid arthritis (RA) synovium. We examined whether mediation of angiogenesis by COX-2 was occuring in cells of the RA synovium and in microvascular endothelial cells (ECs) that are similar to those found in the RA synovium. We demonstrate that rofecoxib, a selective COX-2 inhibitor, acts directly on human dermal microvascular ECs (HMVECs) to inhibit their chemotactic and tube forming ability. Likewise, pretreatment of HMVECs with rofecoxib significantly inhibited their ability to form tubes induced by conditioned media (CM) of activated RA synovial fibroblasts. When RA synovial fibroblasts were pretreated with rofecoxib for 16 h and then stimulated with interleukin (IL)-1β, their CM induced significantly less HMVEC tube formation when compared with CM from vehicle-treated RA synovial fibroblasts. ELISAs performed on activated RA fibroblast CM for known proangiogenic factors demonstrated a significant reduction in bFGF, in addition to the expected decrease in PGE2. Our studies suggest that COX-2-induced angiogenic activity is an active mechanism within diseased synovium and may provide an additional rationale for the use of COX-2 inhibitors in RA.

Introduction

Neovascularization is a hallmark of rheumatoid arthritis (RA) synovial tissue (ST), likely driven by the increased demand for nutrients and oxygen required by the invasive and proliferative synovial pannus Folkman 1995, Koch 1998, Walsh 1999. Early RA synovium is characterized by an increase in sublining capillary and postcapillary venule density. The quantity of synovial blood vessels correlates with synovial cell hyperplasia, mononuclear cell number, and the Ritchie articular index of joint tenderness (Rooney et al., 1988). Studies in animal models provide optimism about the use of angiogenesis inhibitors for RA Brooks et al 1994, Oliver et al 1995, Storgard et al 1999, Kim et al 2002, Yin et al 2002. In short, angiogenesis appears to be a key event in the initiation and persistence of rheumatoid disease and can be strategically inhibited to reduce inflammation in animal models of RA.

Prior to the advent of distinct probes for COX isoforms, many joint cell types were shown to positively immunostain for COX (Sano et al., 1992). Joints from RA patients exhibited abundant COX expression, to higher levels than found in ST from osteoarthritis patients or nonarthritic ST (Sano et al., 1992). Later studies with human and animal tissues used specific COX antibodies and demonstrated COX-2 immunostaining of synovial blood vessels, synovial lining cells, chondrocytes, lymphoid aggregates, and subsynovial fibroblast-like cells Crofford et al 1994, Kang et al 1996, Siegle et al 1998. The advancement of specific COX-2 inhibitors to the clinic represents a major step forward in the pharmacologic approach to the treatment of arthritis. These specific inhibitors have been shown in clinical trials to relieve pain and inflammation associated with RA as effectively as nonsteroidal anti-inflammatory drugs with significantly less adverse side effects (Schnitzer and Hochberg, 2002).

Evidence from a variety of models demonstrates that COX-1 and/or COX-2 regulate angiogenesis or various facets of the angiogenic response Majima et al 1997, Katori et al 1998, Tsujii et al 1998, Daniel et al 1999, Hull et al 1999, Jones et al 1999, Sawaoka et al 1999, Yamada et al 1999, Dicker et al 2001, Hernandez et al 2001. The mechanism through which COX-2 promotes an angiogenic phenotype is likely related to the proangiogenic properties of select eicosanoids, including PGE2, PGE1, and TxA2 Form and Auerbach 1983, Diaz-Flores et al 1994, Nie et al 2000. The RA ST has been examined for eicosanoid production and the principal products of COX activity are PGE2, PGF, TxA2, and PGI2 (Goldstein, 1988). While evidence suggests that COX-2 modulates angiogenesis and COX-2 is upregulated in RA ST, it is not known whether the proangiogenic function of COX-2 contributes to the persistent vascularization in inflamed synovium. In addition, at the onset of these studies, it was not clear whether endothelial COX-2 directly played a role in in vitro angiogenesis. Therefore, we examined the role of COX-2 in RA synovial fibroblasts and microvascular ECs. Our results represent the first demonstration that COX-2 induced angiogenic activity is likely an active mechanism within diseased synovium and provides an additional rationale for the use of COX-2 inhibitors in RA.

Section snippets

Cells and generation of conditioned medium (CM)

Human dermal microvascular endothelial cells (HMVECs) were purchased from Cambrex Bio Science Walkersville (Walkersville, MD) and grown in endothelial cell growth medium (EGM, Cambrex Bio Science, Walkersville, MD) with 10% FBS. Cell assays were performed in endothelial basal medium (EBM, Cambrex Bio Science) supplemented as described below. Fibroblasts were isolated from fresh RA STs. Patients met the criteria established by the American College of Rheumatology (Arnett et al., 1988). All

Rofecoxib inhibits PMA-induced HMVEC chemotaxis

Coculture findings from an early study using human umbilical vein ECs (HUVECs) suggested that COX-1 but not COX-2 was important in regulating facets of angiogenesis (Tsujii et al., 1998). Results from a subsequent study performed using rat primary aortic ECs and HMVECs, however, suggested that EC COX-2 was of importance (Jones et al., 1999). In light of this discrepancy, we set out to directly analyze the effects of a specific COX-2 inhibitor on ECs involved in chemotaxis and tube formation. We

Acknowledgements

This work was supported by a grant from Merck & Company, Inc. (J.M.W. and A.E.K.), NIH grants AI-40987 and HL-58695 (A.E.K.), the Veterans’ Administration Research Service (A.E.K.), and the Gallagher Professorship for Arthritis Research (A.E.K.).

References (48)

  • B Berse et al.

    Hypoxia augments cytokine (transforming growth factor-beta (TGF-beta) and IL-1-induced vascular endothelial growth factor secretion by human synovial fibroblasts

    Clin. Exp. Immunol.

    (1999)
  • A Blanco et al.

    Involvement of tyrosine kinases in the induction of cyclo-oxygenase-2 in human endothelial cells

    Biochem. J.

    (1995)
  • P.C Brooks et al.

    Requirement of vascular integrin alpha v beta 3 for angiogenesis

    Science

    (1994)
  • R Bucala et al.

    Constitutive production of inflammatory and mitogenic cytokines by rheumatoid synovial fibroblasts

    J. Exp. Med.

    (1991)
  • C.C Chan et al.

    Rofecoxib [Vioxx, MK-0966; 4-(4′-methylsulfonylphenyl)-3-phenyl-2-(5H)-furanone]a potent and orally active cyclooxygenase-2 inhibitor. Pharmacological and biochemical profiles

    J. Pharmacol. Exp. Ther.

    (1999)
  • L.J Crofford et al.

    Involvement of nuclear factor kappa B in the regulation of cyclooxygenase-2 expression by interleukin-1 in rheumatoid synoviocytes

    Arthritis Rheum.

    (1997)
  • L.J Crofford et al.

    Basic biology and clinical application of specific cyclooxygenase-2 inhibitors

    Arthritis Rheum.

    (2000)
  • L.J Crofford et al.

    Cyclooxygenase-1 and -2 expression in rheumatoid synovial tissueseffects of interleukin-1 beta, phorbol ester, and corticosteroids

    J. Clin. Invest.

    (1994)
  • T.O Daniel et al.

    Thromboxane A2 is a mediator of cyclooxygenase-2-dependent endothelial migration and angiogenesis

    Cancer Res.

    (1999)
  • L Diaz-Flores et al.

    Intense vascular sprouting from rat femoral vein induced by prostaglandins E1 and E2

    Anat. Rec.

    (1994)
  • A.P Dicker et al.

    Targeting angiogenic processes by combination rofecoxib and ionizing radiation

    Am. J. Clin. Oncol.

    (2001)
  • J Folkman

    Angiogenesis in cancer, vascular, rheumatoid and other disease

    Nat. Med.

    (1995)
  • D.M Form et al.

    PGE2 and angiogenesis

    Proc. Soc. Exp. Biol. Med.

    (1983)
  • I.M Goldstein

    Agents that interfere with arachidonic acid metabolism

  • Cited by (64)

    • Inhibition of regulator of G protein signaling 10, aggravates rheumatoid arthritis progression by promoting NF-κB signaling pathway

      2021, Molecular Immunology
      Citation Excerpt :

      This activation may cause an overexpression in COX-2 levels and pro-inflammatory factor release. COX-2 is expressed via NF-κB activation during inflammatory responses in chondrocytes and could directly lead to angiogenesis and support the progression of RA (Ulivi et al., 2008; Woods et al., 2003). It can also lead to cartilage resorption by inducing high levels of prostaglandin E2 (PGE2), which decreases the proliferation of chondrocytes, enhances MMP-activity, and inhibits aggrecan synthesis in chondrocytes (Nédélec et al., 2001).

    • Comparative study of anti-VEGF Ranibizumab and Interleukin-6 receptor antagonist Tocilizumab in Adjuvant-induced Arthritis

      2018, Toxicology and Applied Pharmacology
      Citation Excerpt :

      Although the precise etiology of RA remains obscure, heightened immune response is thought to play a vital role in provoking joint inflammation and bone erosion (Garnero et al., 2010; Kremer et al., 2011). In this context, infiltrating synovial tissue/joints with immune cells and synoviocytes triggers the production of pro-inflammatory cytokines such as tumor necrosis factor-alpha (TNF-α) and interleukins (ILs) including IL-1b, IL-6, IL-17 and IL-18 that enhance synovial cell proliferation (Darwish et al., 2013) and up-regulate cyclooxygenase-II (COX-II) expression especially by TNF-α and IL-1b (Lee et al., 2000; Woods et al., 2003). Angiogenesis was also found to be a fundamental contributor to RA development and progression (Maruotti et al., 2006; Szekanecz et al., 2009a).

    View all citing articles on Scopus

    Preliminary results were presented as part of the 66th National Meeting of the American College of Rheumatology.

    View full text