OP0181 Vegf induces vasculopathy and fibrosis in vivo closely resembling human systemic sclerosis
Authors: B. Maurer, A. Akhmetshina, R.E. Gay, G. Schett, B.A. Michel, S. Gay, J.H. Distler, O. Distler
Journal: Annals of the Rheumatic Diseases (2013)

<h3>Background</h3> Levels of VEGF (vascular endothelial growth factor) are significantly elevated in systemic sclerosis (SSc) patients even before the development of skin fibrosis. <h3>Objectives</h3> To analyze the effects of VEGF on vasculopathy and skin fibrosis in different animal models of SSc. <h3>Methods</h3> Density and wall thickness of dermal microvessels as well as (hypo-)dermal thickness were assessed by histology using HE and Masson's trichrome staining and immunohistochemistry using anti-vWF and anti-α-SMA antibodies. In the model of bleomycin-induced skin fibrosis, homo- (+/+) (n=8) and heterozygous (+/–) (n=9) VEGF transgenic (tg) mice and wildtype (wt) mice (n=6/9) were treated with bleomycin and compared to saline treated controls. Additionally, TSK (tight skin)1/VEGF+/+ mice were generated (n=4). For statistical analysis, GraphPad Prism software was employed. Parametric non-related data were expressed as mean±SEM, nonparametric non-related data as median<sub>(Q1,Q3)</sub>. P-values &lt;0.05 were considered statistically significant. <h3>Results</h3> In unchallenged VEGF tg mice, the number of dermal microvessels spontaneously increased, which was most pronounced in VEGF+/– mice (mean±SEM saline treated VEGF+/– 21.5±1.3 vs. VEGF+/+ 15.3±3.1 vs. 5.2±1.1/HPF in wt mice; p&lt;0.05). Similar effects were observed for vessel wall thickness (saline treated VEGF+/– 4.2±0.2 vs. VEGF+/+ 3.5±0.3 vs. 1.9±0.2μm in wt mice; p&lt;0.05), thus suggesting that in vivo, intermediate doses of VEGF are pro-angiogenic, whereas high doses lead to decreased angiogenesis. Strikingly, VEGF+/+, but not VEGF+/– tg mice spontaneously developed skin fibrosis (increase of skin thickness by 1.8±0.2 fold compared to wt mice; p&lt;0.05), indicating a dose-dependent pro-fibrotic effect of VEGF in vivo. Challenge with bleomycin enhanced the effects of VEGF on vasculopathy. VEGF+/– compared to VEGF+/+ tg and wt mice showed the most pronounced increase in both the number of microvessels (31.8±1.6 vs. 19.1±1.4 vs. 10.2±0.9/HPF; p&lt;0.05) and vessel wall thickness (4.0±0.2 vs. 3.3±0.1 vs. 2.3±0.2μm; p≤0.05). Additionally, bleomycin further increased skin thickness in VEGF+/+ and induced skin fibrosis in VEGF+/– tg mice (2.3±0.08 and 2.3±0.07 fold; p&lt;0.05). Bleomycin challenged wt mice showed the same increase in skin thickness (1.7±0.02 fold; p&lt;0.05) as VEGF+/+ tg mice without bleomycin challenge. In conclusion, pro-inflammatory conditions aggravated the effects of VEGF. In the non-inflammatory TSK1 mice, VEGF had similar effects. Whereas TSK1 mice showed a reduced number of dermal microvessels compared to wt mice (median<sub>(Q1,Q3)</sub> 2.0<sub>(1,2)</sub> vs. 5.5<sub>(3,6)</sub>/HPF; p&lt;0.05), in VEGF+/+/TSK1 mice, microvessel density (16.5<sub>(11,25)</sub> vs. 2.0<sub>(1,2)</sub>/HPF in TSK1 mice; p&lt;0.05) and vessel wall thickness increased (4.9<sub>(4,8)</sub> vs. 2.9<sub>(3,4)</sub>μm in TSK1 mice; p&lt;0.05) which was paralleled by an increase in hypodermal thickness (694.5±15.1 vs. 456.0±20.6 in TSK1 vs. 102.9±5.7μm in wt mice; p&lt;0.05). <h3>Conclusions</h3> These in vivo data on VEGF tg mice and two SSc models provide a novel link between the vasculopathy and onset of fibrosis in SSc. High levels of VEGF lead to a decrease of angiogenesis. Notably, these high levels of VEGF have potent profibrotic effects and induce dermal fibrosis without additional stimuli. <h3>Disclosure of Interest</h3> B. Maurer: None Declared, A. Akhmetshina: None Declared, R. Gay: None Declared, G. Schett: None Declared, B. Michel: None Declared, S. Gay: None Declared, J. Distler Shareholder of: stock owner of 4D Science, Grant/Research support from: Boehringer Ingelheim, Celgene, Bayer Pharma, Actelion, Pfizer, Ergonex, BMS, JB Therapeutics, Anaphore, Inc, Sanofi-Aventis, Novartis, Array Biopharma and Active Biotec in the area of potential treatments of scleroderma, Consultant for: Boehringer Ingelheim, Celgene, Bayer Pharma, Actelion, Pfizer, Ergonex, BMS, JB Therapeutics, Anaphore, Inc, Sanofi-Aventis, Novartis, Array Biopharma and Active Biotec in the area of potential treatments of scleroderma, O. Distler Grant/Research support from: Actelion, Pfizer, Ergonex, BMS, Sanofi-Aventis, United BioSource Corporation, medac in the area of potential treatments of scleroderma and its complications. Received lecture honoraria from Actelion, Pfizer, Encysive and Ergonex., Consultant for: Actelion, Pfizer, Ergonex, BMS, Sanofi-Aventis, United BioSource Corporation, medac in the area of potential treatments of scleroderma and its complications. Received lecture honoraria from Actelion, Pfizer, Encysive and Ergonex.