• We are sorry, but NCBI web applications do not support your browser and may not function properly. More information
Logo of clinexpimmunolLink to Publisher's site
Clin Exp Immunol. Apr 2003; 132(1): 158–162.
PMCID: PMC1808664

High serum vascular endothelial growth factor correlates with disease activity of spondylarthropathies

Abstract

Angiogenesis is involved in chronic inflammatory joint diseases such as rheumatoid arthritis (RA). Vascular endothelial growth factor (VEGF) plays a crucial role in angiogenesis. The spondylarthropathies (SpA) are characterized by enthesitis and synovitis, in which blood vessels participate. The objective of this study was to investigate serum VEGF levels and their potential associations with disease activity markers for SpA. Sera were collected from 105 patients with SpA (72 with ankylosing spondylitis (AS), four with psoriatic arthritis (PsA), six with reactive arthritis (ReA), eight with enteropathic arthropathy and 15 with undifferentiated SpA), 50 patients with rheumatoid arthritis (RA) and 64 healthy controls. Disease activity in SpA patients was assessed using the Bath Ankylosing Spondylitis Disease Activity Index (BASDAI) and laboratory parameters of inflammation [erythrocyte sedimentation rate (ESR) and C-reactive protein level (CRP)]. Serum VEGF levels were significantly higher in SpA patients (316·4 ± 215·6 pg/ml) and RA patients (405·2 ± 366·5) than in controls (217·3 ± 145·2) (P = 0·003). In SpA patients, serum VEGF levels correlated with disease activity indices (BASDAI: r = 0·22, P = 0·04; ESR: r = 0·3, P = 0·003; and CRP: r = 0·23, P = 0·02). Serum VEGF levels were not associated with presence of extra-articular manifestations or syndesmophytes or with the grade of sacroiliitis. These results suggest that VEGF and therefore angiogenesis may play a role in SpA pathogenesis and may serve as a disease activity marker in SpAs.

Keywords: angiogenesis bone formation spondylarthropathies VEGF

Introduction

The spondylarthropathies (SpA) include several disorders with common clinical and radiological characteristics. Ankylosing spondylitis (AS) is considered the most typical pattern of SpA. Other patterns consist of reactive arthritis (ReA), psoriatic arthritis (PsA) and enteropathic arthropathy (EA) [1].

The rheumatic manifestations of SpA include spinal symptoms, peripheral arthritis and enthesitis. Most patients present with low back pain and sacroiliitis and/or progressive spinal ankylosis and deformity. Enthesitis, defined as inflammation at sites of ligament, tendon and capsule attachment to bone, is a hallmark of SpA. Conversely, synovitis is not a prominent feature of SpA.

The pathological processes underlying the structural lesions and clinical manifestations have been studied extensively in AS. Examination of the sacroiliac joints has shown a compact inflammatory infiltrate containing CD4+ and CD8+ T cells, macrophages and large amounts of tumour necrosis factor (TNF)-α messenger RNA [2]. Similarly, studies of the entheses have shown inflammation characterized by oedema and inflammatory infiltrates composed mainly of CD8+ T cells [3]. In addition, the peripheral joint manifestations of SpA are related to hyperplasia of the synovial membrane, with lymphocyte infiltration, fibrosis and vascular lesions [4].

Angiogenesis is defined by the growth of new capillary blood vessels from the pre-existing vasculature [5]. This process is involved in disease states such as metastatic dissemination and chronic inflammatory joint disease [5,6]. Angiogenesis is a key event in the development and persistence of rheumatoid synovitis [710].

Angiogenesis is regulated by pro-angiogenic factors and angiogenesis inhibitors [8, 11, 12]. Among the pro-angiogenic factors, vascular endothelial growth factor (VEGF) plays a pivotal role in human RA and animal models of arthritis [12]. VEGF has been detected in RA synovial fluids, and serum VEGF levels have been shown to correlate with disease activity [13]. Angiogenesis within the cartilage is also crucial to enchondral bone formation and has been implicated in the development of osteophytes in osteoarthritic joints [11].

The vasculature is also involved in the pathogenesis of SpA. Thus, increased vascularity is a prominent feature of AS synovitis [4]. Sacroiliitis and enthesitis are characterized by fibrous lesions and bone formation, which require active participation of the blood vessels. The role of VEGF in the pathogenesis of SpA has not been examined previously. The objective of this study was to evaluate serum VEGF levels in a cohort of SpA patients and to look for associations with markers of disease activity.

Patients And Methods

Patients

We recruited 105 consecutive patients who met European Spondyloarthropathy Study Group (ESSG) [14] criteria for SpA and were receiving follow-up at our department. The following information was recorded: age, sex and disease duration; symptoms: back pain, spinal stiffness, peripheral arthritis, enthesitis and extra-articular manifestations (uveitis, cardiac involvement) at the time of the study; radiological changes (sacroiliitis, with the grade on the New York scale [15] and syndesmophytes on postero-anterior and lateral plain radiographs of the thoracic and lumbar spine); the Bath Ankylosing Spondylitis Disease Activity Index (BASDAI), which evaluates disease activity [16]; the Westergren erythrocyte sedimentation rate (ESR) and the serum C-reactive protein (CRP) level; whether HLA-B27 was present; and the platelet count.

Fifty patients with RA meeting 1987 American College of Rheumatology criteria were included for purposes of comparison [17]. For each of these patients, the following were recorded: age, sex, and disease duration; extra-articular manifestations (subcutaneous nodules, vasculitis, sicca syndrome, pulmonary or cardiac involvement); tender and swollen joint counts; the Health Assessment Questionnaire (HAQ) score; Larsen's radiological erosions score [18]; the ESR and CRP level; the platelet count; and whether tests were positive for rheumatoid factors.

Controls

The controls were 64 healthy Caucasians with no history of inflammatory joint disease or osteoarthritis. They were chosen from the hospital staff. The controls were matched to the SpA patients on age and sex.

This study was approved by our ethics committee, and all study participants gave their informed consent.

Methods

Serum VEGF levels were determined using a sensitive sandwich ELISA (R&D Systems, Minneapolis, MN, USA). Briefly, this test uses microwell plates precoated with monoclonal antibody and an enzyme-linked polyclonal antibody that is specific for human VEGF. For each study participant, 100 µl of serum was used. All analyses were performed in duplicate. Concentrations were expressed in pg/ml after optical density determination at 450 nm. The detection limit of the VEGF assay was 9·0 pg/ml.

Statistical analysis

Values are expressed as means ± s.d. (medians are also given). The Kruskal–Wallis test was used to compare age and VEGF levels in the three groups (SpA, RA and controls) and to evaluate differences in VEGF measurements between the SpA subgroups. When a significant difference was found, the exact P-values were calculated using the Mann–Whitney U-test. This test was also used to compare variables (disease duration, ESR, CRP and platelets) between SpA and RA patients and to compare VEGF values between SpA subgroups (SpA with and without extra-articular manifestations, syndesmophytes, HLA-B27 or sacroiliitis; and SpA in males and females). Qualitative data (sex) were analysed using the χ2 test. Spearman's r-test was used to calculate correlations between VEGF and indices of disease activity of SpA or RA. Values of P smaller than 0·05 were considered significant.

Results

The demographic, clinical and radiological characteristics of the SpA patients, RA patients and controls are reported in Table 1. Of the 105 patients with SpA, 72 had AS, six had ReA, four had PsA, eight had enteropathic arthropathy (ulcerative colitis or Crohn's disease) and 15 had undifferentiated SpA. Radiological sacroiliitis was found in 80% of SpA patients (grade 0 = 18; grade 1 = 18; grade 2 = 42; grade 3 = 23; ND = 2), whereas syndesmophytes were visible on plain radiographs of the thoracic and lumbar spine in only 21% of SpA patients. Extra-articular manifestations were present in 36% of SpA patients and peripheral arthritis in 36% at the time of evaluation. The HLA-B27 antigen was detected in 83% of SpA patients. Among our RA patients, 42% had extra-articular organ involvement.

Table 1
Clinical, radiological and laboratory features in patients with spondyloarthropathy or rheumatoid arthritis and in healthy controls

The three groups differed regarding age (Kruskal–Wallis, P < 0·0001) and pairwise tests showed that the patients with RA were older, compared to the patients with SpA (Mann–Whitney: P < 0·0001) and to the controls (Mann–Whitney: P < 0·0001). However, age was similar in the SpA group and the control group (Mann–Whitney: P = 0·9). The proportion of female patients was higher in the RA group than in the SpA and control groups (P = 0·0005 and P = 0·007, respectively), whereas the sex ratio was even in the SpA and control groups (P = 0·1). No differences were found between SpA and RA for ESR, CRP, platelet count, or disease duration (all P-values> 0·05).

Serum VEGF levels were significantly higher in the SpA group (316·4 ± 215·6 pg/ml; median, 244·2) and RA group (405·2 ± 366·5 pg/ml; median, 290·4), compared to the control group (217·3 ± 145·2 pg/ml; median, 187·5) (Kruskal–Wallis: P = 0·003). Both SpA and RA patients had higher VEGF values compared to controls (Mann–Whitney P = 0·005 and P = 0·002, respectively), whereas VEGF levels were similar in the RA and SpA groups (P = 0·3) (Fig. 1).

Fig. 1
Serum concentration of vascular endothelial growth factor in patients with spondyloarthropathies or rheumatoid arthritis and in healthy controls. Box plots represent the lower quartile, the median and the upper quartile. The ends of the lines indicate ...

In the SpA group, serum VEGF levels were positively correlated with ESR, CRP and the BASDAI score (r = 0·30, P = 0·003; r = 0·23, P = 0·02; and r = 0·22, P = 0·04, respectively) but were not correlated with disease duration or age (P > 0·05). Serum VEGF was positively correlated with the platelet count (r = 0·31, P = 0·003). VEGF levels were not influenced by gender, presence of extra-articular disease or syndesmophytes or grade of sacroiliitis (all P-values> 0·05). Compared to controls, serum VEGF levels were higher both in the SpA patients with peripheral disease (P = 0·05) and in those with axial disease (P = 0·006). The 17 HLA-B27-negative patients had higher serum VEGF levels (419·6 ± 243·2; median, 448·1) than did the 85 HLA-B27-positive patients (297·2 ± 208·3; median, 230·2] pg/ml; P = 0·04). However, after exclusion of the HLA-B27-negative patients, VEGF remained higher in the SpA group compared to the control group (297·2 ± 208·3 pg/ml; median, 230·2 pg/ml; versus 217·3 ± 145·2 pg/ml; median, 187·5 pg/ml; P = 0·03). Serum VEGF levels were similar in the subgroups defined by the type of SpA (Kruskal–Wallis: P = 0·5) (Fig. 2).

Fig. 2
Serum vascular endothelial growth factor concentration in the spondyloarthropathy subgroups. AS: ankylosing spondylitis; PsA: psoriatic arthritis; ReA: reactive arthritis; EA: enteropathic arthropathy: USpA: undifferentiated spondyloarthropathy.

In the RA group, serum VEGF was not correlated with ESR, CRP or platelet count (all P-values > 0·05).

Discussion

Angiogenesis is an active process necessary for physiological functions such as wound healing and the female reproductive cycle. Angiogenesis also plays an important role in abnormal phenomena such as tumour growth and pannus formation in RA [5,10]. VEGF is considered a pivotal factor in angiogenesis and has been detected in rheumatoid synovial fluid [8, 9, 13]. VEGF is produced mainly by macrophages and synovial lining cells and acts specifically on endothelial cells. Previous studies found higher VEGF levels in RA patients compared to healthy subjects, patients with osteoarthritis and patients with other collagen diseases [13,1921]. A correlation between VEGF and markers of inflammation, in particular CRP, has been reported [13,19]. The results in our RA group agree in part with these observations.

Angiogenesis also plays a role in bone formation. During formation of bone from cartilage or enchondral ossification, invasion of the cartilage by new blood vessels precedes osteoblastic transformation and ossification. Chondrocytes and osteoblasts can produce angiogenic factors such as VEGF [11]. Thus, angiogenesis within cartilage is important for enchondral ossification and contributes to the development of osteophytes. Patients with osteoarthritis had higher serum VEGF levels than healthy controls but lower levels than RA patients [11]. These data indicate that angiogenesis plays a central role in diseases characterized by bone formation.

The SpAs are characterized both by bone formation, resulting in progressive fusion of the sacroiliac joints and ossification of entheses, and by synovitis. Additional arguments in favour of vascular involvement include the presence of nail fold capillaroscopy abnormalities in patients with AS and the occurrence of extra-articular manifestations in SpAs [22]. These observations strongly support a role for angiogenesis in the pathogenesis of SpA and warrant evaluations of angiogenic factors in these diseases.

In our study, serum VEGF levels were significantly higher in the patients with SpA and RA than in the controls. Furthermore, serum VEGF levels were correlated strongly with clinical and laboratory indices of disease activity of SpA. These results support a role for angiogenesis in the pathogenesis of SpA, particularly the development of inflammatory lesions. Serum VEGF levels were significantly elevated in both patients with axial SpA and patients with peripheral SpA, compared to controls, suggesting that angiogenesis contributes to both enthesitis and synovitis. Surprisingly, serum VEGF levels were not correlated with the presence of syndesmophytes or the grade of sacroiliitis. One explanation may be that serum VEGF was measured at a single point in time, whereas syndesmophytes and sacroiliitis develop over many years. Another unexpected finding, for which we have no explanation, is the lower VEGF level in patients with than without the HLA-B27 antigen.

In this study, we evaluated VEGF in the serum, not the plasma. Serum collection causes activation of platelets, which may release VEGF. However, the few available data on plasma VEGF have produced conflicting results [23]. One study in cancer patients found undetectable or very low VEGF levels in plasma contrasting with very high levels in serum [24]. In our study, a correlation between serum VEGF and platelet count was found in the SpA group but not in the RA group.

Angiogenesis has seldom been investigated in AS and other SpAs. The only angiogenic factor that has been evaluated in AS is endothelial cell-stimulating angiogenesis factor, or ESAF, a poorly characterized endothelial growth factor. This pro-angiogenic factor is involved in osteoarthritis but not in RA [11]. Taylor et al. reported that serum ESAF in patients with AS was elevated as compared to controls and correlated with an enthesitis index. Interestingly, levels of ESAF were higher in patients with more advanced sacroiliitis but were not correlated with laboratory parameters [25,26].

VEGF is secreted by various cell types, including macrophages, which are present in the synovial membrane and entheses in patients with SpA [2,3]. VEGF is inducible by various factors such as hypoxia and inflammatory cytokines including TNF-α [58]. TNF-α seems to be a key cytokine in the pathophysiology of AS: a large amount of TNF-α messenger RNA has been found in sacroiliac joints, and anti-TNF-α agents have recently been found effective in AS and SpA [2,27]. Moreover, histological examination of synovium from patients with AS treated with an anti-TNF-α agent showed reductions in synovial lining thickness, vascularity and infiltration with neutrophils and macrophages [27]. Thus, it would be relevant to assess the effect of anti-TNF-α agents on angiogenic factors in SpA.

At the time of the study, all our patients were receiving non-steroidal anti-inflammatory drugs, and some were also taking sulphasalazine or methotrexate. These drugs can reduce endothelial cell proliferation or inhibit angiogenesis, which may have influenced our results [8,11]. Conversely, if angiogenesis is involved in the pathophysiology of SpA, then inhibition of angiogenesis may contribute to the efficacy of these drugs.

Our data support a role for angiogenesis in the pathophysiology and activity SpA, because levels of the potent angiogenic factor VEGF were elevated in our SpA patients and correlated with disease activity. As angiogenesis plays a role in synovial membrane vascularity and excessive bone formation, it may contribute to the genesis of synovitis and enthesitis in SpA. Further studies are required to understand better the role of VEGF and other angiogenic factors in SpA, with special attention to VEGF expression in affected sacroiliac joints and peripheral joints. Furthermore, agents that interfere with angiogenesis may hold potential for the treatment of SpA.

Acknowledgments

This work was supported by a grant from the non-profit organization ‘Association Franc-Comtoise pour la Recherche et l’Enseignement en Rhumatologie’.

References

1. Calin A. Terminology, introduction, diagnostic criteria, and overview. In: Calin A, Taurog JD, editors. The spondylarthritides. Oxford: Oxford University Press; 1998. pp. 1–15.
2. Braun J, Bollow M, Neure L, et al. Use of immunohistological and in situ hybridization techniques in the examination of sacroiliac joint biopsy specimens from patients with ankylosing spondylitis. Arthritis Rheum. 1995;38:499–505. [PubMed]
3. Laloux L, Voisin MC, Allain J, et al. Immunohistological study of entheses in spondylarthropathies: comparison in rheumatoid arthritis and osteoarthritis. Ann Rheum Dis. 2001;60:316–21. [PMC free article] [PubMed]
4. Kidd BL, Moore K, Walters MT, Smith JL, Cawley MID. Immunohistological features of synovitis in ankylosing spondylitis: a comparison with rheumatoid arthritis. Ann Rheum Dis. 1989;48:92–8. [PMC free article] [PubMed]
5. Folkman J. Angiogenesis in cancer, vascular, rheumatoid arthritis and other disease. Nature Med. 1995;1:27–31. [PubMed]
6. Weber AJ, De Bandt M. Angiogenesis: general mechanisms and implications for rheumatoid arthritis. Joint Bone Spine. 2000;67:366–83. [PubMed]
7. Paleolog EM. Angiogenesis: a critical process in the pathogenesis of RA − a role for VEGF? Br J Rheumatol. 1996;35:917–20. [PubMed]
8. Koch AE. Angiogenesis. Implications for rheumatoid arthritis. Arthritis Rheum. 1998;41:951–62. [PubMed]
9. Walsh DA. Angiogenesis and arthritis. Rheumatology. 1999;38:103–12. [PubMed]
10. Walsh DA, Pearson CI. Angiogenesis in the pathogenesis of inflammatory joint and lung diseases. Arthritis Res. 2001;3:147–53. [PMC free article] [PubMed]
11. Ballara SC, Miotla JM, Paleolog EM. New vessels, new approaches: angiogenesis as a therapeutic target in musculoskeletal disorders. Int J Exp Pathol. 1999;80:235–50. [PMC free article] [PubMed]
12. Amoroso A, Del Porto F, Di Monaco C, Manfredini P, Afeltra A. Vascular endothelial growth factor: a key mediator of neoangiogenesis. A review. Eur Rev Med Pharmacol Sci. 1997;1:17–25. [PubMed]
13. Lee S, Joo YS, Kim WU, et al. Vascular endothelial growth factor levels in the serum and synovial fluid of patients with rheumatoid arthritis. Clin Exp Rheumatol. 2001;19:321–4. [PubMed]
14. Dougados M, Van der Linden S, Juhlin R, et al. The European Spondylarthropathy Study Group preliminary criteria for the classification of spondylarthropathy. Arthritis Rheum. 1991;34:1218–27. [PubMed]
15. Van der Linden S, Valkenburg HA, Cats A. Evaluation of diagnosis criteria for ankylosing spondylitis. A proposal for modification of the New York criteria. Arthritis Rheum. 1984;27:361–8. [PubMed]
16. Garrett P, Jenkinsson T, Kennedy LG, Whitelock H, Gaisford P, Calin A. A new approach to defining disease status in ankylosing spondylitis: the Bath Ankylosing Spondylitis Disease Activity Index. J Rheumatol. 1994;21:2286–91. [PubMed]
17. Arnett FC, Edworthy SM, Bloch DA, et al. The American Rheumatism Association 1987 revised criteria for the classification of rheumatoid arthritis. Arthritis Rheum. 1988;31:315–24. [PubMed]
18. Larsen A. How to apply Larsen score in evaluating radiographs of rheumatoid arthritis in long term studies? J Rheumatol. 1995;22:1974–5. [PubMed]
19. Harada M, Mitsuyama K, Yoshida H, et al. Vascular endothelial growth factor in patients with rheumatoid arthritis. Scand J Rheumatol. 1998;27:377–80. [PubMed]
20. Pinheiro GRC, Andrade CAF, Gayer CR, Coelho MS, Freire SM, Scheinberg MA. Serum vascular endothelial growth factor in late rheumatoid arthritis. Clin Exp Rheumatol. 2001;19:721–3. [PubMed]
21. Kikuchi K, Kubo M, Kadono T, Yazawa N, Ihn H, Tamaki K. Serum concentrations of vascular endothelial growth factor in collagen diseases. Br J Dermatol. 1998;139:1049–51. [PubMed]
22. Wendling D, Risold JC. Microcirculation in ankylosing spondylitis. Ann Rheum Dis. 1994;53:284. [PMC free article] [PubMed]
23. George ML, Eccles SA, Tutton MG, Abulafi AM, Swift RI. Correlation of plasma and serum vascular endothelial growth factor levels with platelet count in colorectal cancer: clinical evidence of platelet scavenging? Clin Cancer Res. 2000;6:3147–52. [PubMed]
24. Salven P, Orpana A, Joensuu H. Leukocytes and platelets of patients with cancer contain high levels of vascular endothelial growth factor. Clin Cancer Res. 1999;5:487–91. [PubMed]
25. Taylor HG, Weiss JB, McLaughlin B, Dawes PT. Raised endothelial cell stimulating angiogenesis factor in ankylosing spondylitis. Clin Exp Rheumatol. 1993;11:537–9. [PubMed]
26. Jones PBB, Makki RJ, Weiss JB. Endothelial cell stimulating angiogenesis factor − a new biological marker for disease activity in ankylosing spondylitis? Br J Rheumatol. 1994;33:332–5. [PubMed]
27. Baeten D, Kruithof E, Van den Bosch Demetter P, et al. Immunomodulatory effects of anti-tumor necrosis factor α therapy on synovium in spondylarthropathy. Histologic findings in eight patients from an open-label pilot study. Arthritis Rheum. 2001;44:186–95. [PubMed]

Articles from Clinical and Experimental Immunology are provided here courtesy of British Society for Immunology

Formats:

Related citations in PubMed

See reviews...See all...

Cited by other articles in PMC

See all...

Links

  • MedGen
    MedGen
    Related information in MedGen
  • PubMed
    PubMed
    PubMed citations for these articles
  • Substance
    Substance
    PubChem Substance links

Recent Activity

Your browsing activity is empty.

Activity recording is turned off.

Turn recording back on

See more...