Overexpression of VEGF165b, an Inhibitory Splice Variant of Vascular Endothelial Growth Factor, Leads to Insufficient Angiogenesis in Patients With Systemic Sclerosis

• Published in: Circulation research Vol. 109; no. 3; pp. e14 - e26
• Main Authors: Manetti, Mirko, Guiducci, Serena, Romano, Eloisa, Ceccarelli, Claudia, Bellando-Randone, Silvia, Conforti, Maria Letizia, Ibba-Manneschi, Lidia, Matucci-Cerinic, Marco
• Format: Journal Article
• Language: English
• Published: United States American Heart Association, Inc 22.07.2011
• Subjects: Alternative Splicing - physiology; Cells, Cultured; Culture Media, Conditioned - pharmacology; Dermis - blood supply; Endothelial Cells - cytology; Endothelial Cells - physiology; Gene Expression - drug effects; Gene Expression - physiology; Humans; Ischemia - genetics; Ischemia - metabolism; Ischemia - physiopathology; Neovascularization, Pathologic - genetics; Neovascularization, Pathologic - metabolism; Neovascularization, Pathologic - physiopathology; Nuclear Proteins - genetics; Nuclear Proteins - metabolism; Phosphoproteins - genetics; Phosphoproteins - metabolism; Phosphorylation - drug effects; Phosphorylation - physiology; RNA-Binding Proteins; Scleroderma, Systemic - genetics; Scleroderma, Systemic - metabolism; Scleroderma, Systemic - physiopathology; Serine-Arginine Splicing Factors; Signal Transduction - drug effects; Signal Transduction - physiology; Transforming Growth Factor beta1 - genetics; Transforming Growth Factor beta1 - metabolism; Transforming Growth Factor beta1 - pharmacology; Vascular Endothelial Growth Factor A - genetics; Vascular Endothelial Growth Factor A - metabolism; Vascular Endothelial Growth Factor A - pharmacology; Vascular Endothelial Growth Factor B - genetics; Vascular Endothelial Growth Factor B - metabolism; Vascular Endothelial Growth Factor B - pharmacology; Vascular Endothelial Growth Factor Receptor-2 - metabolism
• ISSN: 0009-7330; 1524-4571; 1524-4571
• DOI: 10.1161/CIRCRESAHA.111.242057

RATIONALE:Systemic sclerosis (SSc) is characterized by widespread microangiopathy, fibrosis, and autoimmunity. Despite the lack of angiogenesis, the expression of vascular endothelial growth factor A (VEGF) was shown to be upregulated in SSc skin and circulation; however, previous studies did not distinguish between proangiogenic VEGF165 and antiangiogenic VEGF165b isoforms, which are generated by alternative splicing in the terminal exon of VEGF pre-RNA. OBJECTIVE:We investigated whether VEGF isoform expression could be altered in skin and circulation of patients with SSc. METHODS AND RESULTS:Here, we show that the endogenous antiangiogenic VEGF165b splice variant is selectively overexpressed at both the mRNA and protein levels in SSc skin. Elevated VEGF165b expression correlated with increased expression of profibrotic transforming growth factor-β1 and serine/arginine protein 55 splicing factor in keratinocytes, fibroblasts, endothelial cells, and perivascular inflammatory cells. Circulating levels of VEGF165b were significantly higher in patients with SSc than in control subjects. Microvascular endothelial cells (MVECs) isolated from SSc skin expressed and released higher levels of VEGF165b than healthy MVECs. Transforming growth factor-β1 upregulated the expression of VEGF165b and serine/arginine protein 55 in both SSc and healthy MVECs. In SSc MVECs, VEGF receptor-2 was overexpressed, but its phosphorylation was impaired. Recombinant VEGF165b and SSc-MVEC–conditioned medium inhibited VEGF165-mediated VEGF receptor-2 phosphorylation and capillary morphogenesis in healthy MVECs. The addition of anti-VEGF165b blocking antibodies abrogated the antiangiogenic effect of SSc-MVEC–conditioned medium. Capillary morphogenesis was severely impaired in SSc MVECs and could be ameliorated by treatment with recombinant VEGF165 and anti-VEGF165b blocking antibodies. CONCLUSIONS:In SSc, a switch from proangiogenic to antiangiogenic VEGF isoforms may have a crucial role in the insufficient angiogenic response to chronic ischemia.