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 Table of Contents  
BRIEF REPORT
Year : 2018  |  Volume : 13  |  Issue : 3  |  Page : 182-185

Serum vascular endothelial growth factor levels as a marker of skin thickening, digital ischemia, and interstitial lung disease in systemic sclerosis


Institute of Rheumatology, Madras Medical College and RGGGH, Chennai, Tamil Nadu, India

Date of Web Publication21-Aug-2018

Correspondence Address:
Dr. Chinnadurai Saranya
No. 1, 7th Cross Street, Near LIC Colony, Anna Nagar, Pammal, Chennai - 600 070, Tamil Nadu
India
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Source of Support: None, Conflict of Interest: None


DOI: 10.4103/injr.injr_132_17

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  Abstract 


Background: Vascular injury is the initial event in the pathogenesis of systemic sclerosis (SSc) and potent proangiogenic mediators such as vascular endothelial growth factor (VEGF) are overexpressed in the skin and circulation of patients with SSc. The objective of this study was to determine the serum levels of VEGF in patients with SSc and to correlate it with the severity of skin thickening, digital ischemia, and interstitial lung disease (ILD).
Methods: Serum VEGF levels were measured in 55 patients with SSC who fulfilled 2013 ACR/EULAR classification criteria for scleroderma and 30 healthy age- and gender-matched controls by ELISA. All patients underwent detailed clinical examination, baseline blood investigations, chest X-ray, electrocardiogram, and pulmonary function tests. Echocardiography and high-resolution computed tomography scan of lungs were done wherever necessary.
Results: Median serum VEGF in SSc patients was higher than in the controls (675 pg/ml [interquartile range (IQR): 395–920] vs. 180.5 pg/ml [IQR: 155–215], respectively; P = 0.00001). No statistically significant difference was observed between diffuse (662.5 pg/ml [IQR: 441.25–942.5]) and limited SSc (680 pg/ml [IQR: 325–850]) (P = 0.412). Serum VEGF levels correlated significantly with higher modified Rodnan skin scores (r = 0.7168) (P < 0.0001) and lower forced vital capacity (r = −0.6771) (P < 0.0001). Median VEGF levels were higher in patients with digital ischemia (digital pitted scars, digital ulcers, and gangrene) compared to those without ischemic changes (P = 0.001). Patients with ILD (n = 21) had significantly higher median VEGF levels when compared to those without ILD (n = 34) (870 pg/ml [IQR: 592.5–1000] vs. 467.5 pg/ml [IQR: 297.5–760]; P = 0.001). There was no significant difference in serum VEGF levels between early (650 pg/ml [IQR: 375–885]) and late stages (890 pg/ml [IQR: 530–1000]) of disease (P = 0.197).
Conclusion: Serum VEGF levels were elevated in SSc and they correlated with the severity of skin thickening, digital ischemia, and presence of ILD.

Keywords: Digital ischemia, interstitial lung disease, modified Rodnan skin scores, systemic sclerosis


How to cite this article:
Saranya C, Ramesh R, Bhuvanesh M, Balaji C, Balameena S, Rajeswari S. Serum vascular endothelial growth factor levels as a marker of skin thickening, digital ischemia, and interstitial lung disease in systemic sclerosis. Indian J Rheumatol 2018;13:182-5

How to cite this URL:
Saranya C, Ramesh R, Bhuvanesh M, Balaji C, Balameena S, Rajeswari S. Serum vascular endothelial growth factor levels as a marker of skin thickening, digital ischemia, and interstitial lung disease in systemic sclerosis. Indian J Rheumatol [serial online] 2018 [cited 2019 Oct 15];13:182-5. Available from: http://www.indianjrheumatol.com/text.asp?2018/13/3/182/234004




  Introduction Top


Systemic sclerosis (SSc) is an autoimmune disease characterized by microvascular damage and excessive fibrosis in skin and internal organs. Vascular injury is the initial event in the pathogenesis of SSc.[1] Angiogenic factors are likely to be involved in the initiation of endothelial cell dysfunction and vascular injury.[2] Vascular injury and chronic tissue hypoxia induce hypoxia-inducible factor-1 (HIF-1) α-dependent genes such as vascular endothelial growth factor (VEGF) and its receptors.[3] VEGF is an endothelial cell-specific mitogen and a potent angiogenic peptide secreted by a variety of cell types.[4]

VEGF increases blood vessel permeability through two selective endothelial cells' surface receptors, i.e., VEGFR-1 and VEGFR-2. VEGFR-2 mediates most of the known cellular responses to VEGF. VEGFR-1 is thought to modulate VEGFR-2 signaling.[5] VEGF, after receptor binding, stimulates endothelial cell proliferation and plasminogen activation. It also induces α- and β-integrin subunits' expression, changes in collagenase activation resulting in migration of endothelial cells, and tube formation.[6],[7]

The VEGF family includes members VEGF-A (or mentioned commonly as VEGF), B, C, D, and placental growth factor. VEGF-A is important for the formation of blood vessels during development or pathological conditions.[8] Elevated levels of VEGF have been reported in the serum of patients with SSc, rheumatoid arthritis, polymyositis/dermatomyositis, and systemic lupus erythematosus.[9] This study was carried out to determine the serum levels of VEGF-A in SSc patients and to correlate the levels with the severity of skin thickening, digital ischemia, and presence of interstitial lung disease (ILD).


  Methods Top


This was a cross-sectional observational study conducted from October 2015 to March 2016 at our institute. Patients diagnosed as SSc were recruited from the rheumatology outpatient department. All the patients were treatment naïve and fulfilled the 2013 American College of Rheumatology/European League Against Rheumatism (ACR/EULAR) classification criteria for SSc.[10] Treatment naïve was defined as a patient not started on any medications for SSc. We excluded those patients with a history of smoking, systemic hypertension, diabetes mellitus, and valvular and coronary heart disease. Coronary and valvular heart disease was diagnosed based on history, clinical examination, electrocardiogram, and chest X-ray (CXR). Doppler echocardiography was done if any of the above were abnormal. SSc patients who were already on immunosuppressants and other medications were also excluded from the study. Of the total 70 SSc patients, 15 were excluded and 55 treatment-naïve SSc patients were taken as the study group. Thirty healthy age- and gender-matched controls were recruited among the attenders or visitors to the hospital.

All patients underwent detailed clinical examination and laboratory investigations, namely, complete hemogram, erythrocyte sedimentation rate, C-reactive protein, urine routine examination, blood urea, and serum creatinine. Assessment of dyspnea was done using the modified Medical Research Council (MRC) dyspnea score by a single physician. Patients underwent baseline pulmonary function tests (PFTs), which include forced vital capacity (FVC), forced expiratory volume in 1 s (FEV1) and the ratio FEV1/FVC, electrocardiography, and plain chest radiograph. Doppler echocardiography and high-resolution computed tomography scan of the chest were done if there was strong clinical suspicion or PFT abnormality or CXR abnormality suggestive of ILD and/or pulmonary hypertension was present. The degree of skin thickening was evaluated using modified Rodnan skin score (mRSS)[11] by a single physician. Digital ischemic changes were assessed clinically by the presence of digital pitted scars, digital ulcers, and gangrene.

About 5 mL of venous blood was collected by venepuncture from both the patients and the controls into pyrogen-free tubes. Samples were centrifuged at 1500 rpm for 10 min; sera were separated and stored at −70°C for 5 months until further evaluated. Serum level of VEGF-A was determined by immune-enzymatic method using commercially available kit ELISA (Human VEGF-A ELISA kit version 11 by Diaclone, France).

Statistical methods

Mann–Whitney U-test was employed for comparison of two independent values which are not normally distributed. Correlation between numerical values was evaluated by Spearman's rho correlation coefficient. P < 0.05 was considered as statistically significant. Statistical analysis was done using Statistical Package for Social Sciences software version 20 (Armonk, NY, IBM Corp).

Ethical approval

The study was approved by the ethics committee of the institute. Informed written consent was obtained from all patients prior to their enrollment in this study.


  Results Top


A total of 55 treatment-naïve SSc patients were included in the study. The median age of patients and controls was 38 and 39 years, respectively. All the patients and controls were females. Of the 55 SSc patients, 30 were diffuse cutaneous type and 25 were limited cutaneous type. The baseline characteristics of the study group are shown in [Table 1]. The duration of disease is taken from the onset of first non-Raynaud symptom. Early stage of disease was defined as the rapid and severe increasing thickening of the skin (1–3 years) while late stage was defined based on softening of cutaneous induration which occurs after skin induration has reached its peak in extent and severity (>3 years).[12]
Table 1: Baseline characteristics of the study group

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Median serum VEGF in SSc patients was 675 pg/ml (IQR: 395–920) and in controls was 180.5 pg/ml (IQR: 155–215). The difference was statistically significant with P = 0.0001. Median serum VEGF levels in diffuse and limited SSc were 662.5 pg/ml (IQR: 441.25–942.5) and 680 pg/ml (IQR: 325–850), respectively. We found no statistical significant difference between diffuse and limited SSc (P = 0.412). We found positive correlation between serum VEGF levels with mRSS (r = 0.7168) as depicted in [Figure 1].
Figure 1: Correlation of modified Rodnan skin score with serum vascular endothelial growth factor levels

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Similarly, we found that median VEGF levels were higher in patients with digital ischemia (digital pitted scars 690 pg/ml [IQR: 450–846.25] and digital ulcers and gangrene 952.5 pg/ml [IQR: 656.2–1000]) compared to those without ischemic changes 250 pg/ml (IQR: 170–282.5) (P < 0.001).

SSc patients with ILD (n = 21) had median VEGF levels of 870 pg/ml (IQR: 592.5–1000) when compared to those without ILD (n = 34) 467.5 pg/ml (IQR: 297.5–760). The difference between the two groups was statistically significant (P = 0.001). Higher serum VEGF levels inversely correlated with FVC (r = −0.6771) (P < 0.0001). There was a significant positive correlation between the modified MRC dyspnea score and serum VEGF levels (r = 0.7169) (P < 0.0001). There was no significant difference in serum VEGF levels between early (650 pg/ml [IQR: 375–885]) and late stages (890 pg/ml [IQR: 530–1000]) of disease (P = 0.197). Furthermore, our observation showed no significant difference between VEGF levels in patients with and without telangiectasias (P = 0.076).


  Discussion Top


We found significantly high levels of VEGF levels in our cohort of treatment-naïve SSc patients when compared to the healthy controls. The levels of serum VEGF significantly correlated with the severity of skin thickening (measured by mRSS). A study by Choi et al. showed similar results where the serum VEGF levels correlated well with the extent of skin sclerosis.[13] However, few studies showed no statistically significant correlation between serum VEGF levels with the duration and severity of skin lesion.[14] A study by Distler et al. demonstrated high expression of both VEGF and its receptors, i.e., VEGFR-1 and VEGFR-2 in skin specimens of SSc patients.[3] In an experimental model of VEGF transgenic mice, VEGF dose dependently exerted strong profibrotic effects with increased collagen synthesis by dermal fibroblasts. In addition, VEGF transgenic mice model demonstrated that uncontrolled, chronic, and strong overexpression of VEGF might paradoxically lead to ineffective angiogenesis.[15]

Our data also showed the significantly higher levels of VEGF in patients who had digital ischemic manifestations such as digital pitted scars, digital ulcers, and gangrene than those who had only Raynaud's phenomenon without these features. Similar findings were supported by the study done by Distler et al., demonstrating the higher serum levels of VEGF in patients with fingertip ulcers.[16]

We also found that serum VEGF was significantly elevated in the subset of patients with ILD as evidenced by inverse correlation with FVC. Similar findings were reported by Kikuchi et al., where they studied the correlation of serum VEGF levels with the lung fibrosis.[9] Our study has shown a significant correlation between serum VEGF levels and dyspnea as expressed by the modified MRC dyspnea score. A study by Papaioannou et al. showed similar correlation of serum VEGF levels with the MRC dyspnea score.[17] In this study, we found no statistically significant difference in VEGF levels between early and late stages of SSc. Furthermore, patients with and without telangiectasias showed no significant difference in the serum VEGF levels.

The limitations of this study were that we could not assess the severity of pulmonary hypertension by measuring pulmonary arterial pressure through Doppler echocardiography or right-heart catheterization. Measurement of DLCO (diffusing capacity of the lung for carbon monoxide) could not be done. This study was done by cross-sectional method, evaluating the serum VEGF levels at the initial diagnosis of the patient; hence, correlation of serum VEGF levels with the progressive course of clinical illness and with the treatment response could not be made. Furthermore, mRSS was done by a single observer.

We found that serum VEGF levels were elevated in patients with SSc as compared to healthy controls and significantly correlated with the severity of skin thickening, digital ischemia, and presence of ILD. This may suggest that VEGF is associated with digital ischemia and ILD in SSc. This study may pave way for further research on VEGF as a biomarker in early identification of SSc patients prone to develop fibrosis and vasculopathy complications.

Financial support and sponsorship

Nil.

Conflicts of interest

There are no conflicts of interest.



 
  References Top

1.
Varga J, Lafyatis R. Etiology and pathogenesis of systemic sclerosis. In: Hochberg MC, Silman AJ, Smolen JS, Weinblatt ME, Weisman MH (eds). Rheumatology, 6th ed. Philadelphia PA: Mosby; 2015. p. 1177.  Back to cited text no. 1
    
2.
Voelkel NF, Cool C, Taraceviene-Stewart L, Geraci MW, Yeager M, Bull T, et al. Janus face of vascular endothelial growth factor: The obligatory survival factor for lung vascular endothelium controls precapillary artery remodeling in severe pulmonary hypertension. Crit Care Med 2002;30:S251-6.  Back to cited text no. 2
    
3.
Distler O, Distler JH, Scheid A, Acker T, Hirth A, Rethage J, et al. Uncontrolled expression of vascular endothelial growth factor and its receptors leads to insufficient skin angiogenesis in patients with systemic sclerosis. Circ Res 2004;95:109-16.  Back to cited text no. 3
    
4.
Ferrara N, Gerber HP, LeCouter J. The biology of VEGF and its receptors. Nat Med 2003;9:669-76.  Back to cited text no. 4
    
5.
Shibuya M. Vascular endothelial growth factor (VEGF) and its receptor (VEGFR) signaling in angiogenesis: A crucial target for anti- and pro-angiogenic therapies. Genes Cancer 2011;2:1097-105.  Back to cited text no. 5
    
6.
Dvorak HF, Detmar M, Claffey KP, Nagy JA, van de Water L, Senger DR, et al. Vascular permeability factor/vascular endothelial growth factor: An important mediator of angiogenesis in malignancy and inflammation. Int Arch Allergy Immunol 1995;107:233-5.  Back to cited text no. 6
    
7.
Distler JH, Wenger RH, Gassmann M, Kurowska M, Hirth A, Gay S, et al. Physiologic responses to hypoxia and implications for hypoxia-inducible factors in the pathogenesis of rheumatoid arthritis. Arthritis Rheum 2004;50:10-23.  Back to cited text no. 7
    
8.
Zhang F, Tang Z, Hou X, Lennartsson J, Li Y, Koch AW, et al. VEGF-B is dispensable for blood vessel growth but critical for their survival, and VEGF-B targeting inhibits pathological angiogenesis. Proc Natl Acad Sci U S A 2009;106:6152-7.  Back to cited text no. 8
    
9.
Kikuchi K, Kubo M, Kadono T, Yazawa N, IHN H, Tamaki K, et al. Serum concentrations of vascular endothelial growth factor in collagen diseases. Br J Dermatol 1998;139:1049-51.  Back to cited text no. 9
    
10.
van den Hoogen F, Khanna D, Fransen J, Johnson SR, Baron M, Tyndall A, et al. 2013 classification criteria for systemic sclerosis: An American college of rheumatology/European league against rheumatism collaborative initiative. Ann Rheum Dis 2013;72:1747-55.  Back to cited text no. 10
    
11.
Clements PJ, Lachenbruch PA, Seibold JR, Zee B, Steen VD, Brennan P, et al. Skin thickness score in systemic sclerosis: An assessment of interobserver variability in 3 independent studies. J Rheumatol 1993;20:1892-6.  Back to cited text no. 11
    
12.
Clements P, Medsger TA, Feghali C. Cutaneous involvement in systemic sclerosis. In: Clements P, Furst DE, editors. Sytemic Sclerosis. 2nd ed. Philadelphia: Lippincott Williams and Wilkins; 2004. p. 129-50.  Back to cited text no. 12
    
13.
Choi JJ, Min DJ, Cho ML, Min SY, Kim SJ, Lee SS, et al. Elevated vascular endothelial growth factor in systemic sclerosis. J Rheumatol 2003;30:1529-33.  Back to cited text no. 13
    
14.
Dziankowska-Bartkowiak B, Waszczykowska E, Zalewska A, Sysa-Jêdrzejowska A. Evaluation of serum vascular endothelial growth factor and endostatin in systemic sclerosis patients – Correlation with lung and cardio-vascular system involvement. Centr Eur J Immunol 2004;29:15-22.  Back to cited text no. 14
    
15.
Maurer B, Distler A, Suliman YA, Gay RE, Michel BA, Gay S, et al. Vascular endothelial growth factor aggravates fibrosis and vasculopathy in experimental models of systemic sclerosis. Ann Rheum Dis 2014;73:1880-7.  Back to cited text no. 15
    
16.
Distler O, Del Rosso A, Giacomelli R, Cipriani P, Conforti ML, Guiducci S, et al. Angiogenic and angiostatic factors in systemic sclerosis: Increased levels of vascular endothelial growth factor are a feature of the earliest disease stages and are associated with the absence of fingertip ulcers. Arthritis Res 2002;4:R11.  Back to cited text no. 16
    
17.
Papaioannou AI, Zakynthinos E, Kostikas K, Kiropoulos T, Koutsokera A, Ziogas A, et al. Serum VEGF levels are related to the presence of pulmonary arterial hypertension in systemic sclerosis. BMC Pulm Med 2009;9:18.  Back to cited text no. 17
    


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