Regulation of vascular endothelial growth factor-C by tumor necrosis factor- in the conjunctiva and pterygium

Vascular endothelial growth factor C (VEGF-C) plays an important role in the development of a pterygium through lymphangiogenesis. We examined the association between VEGF-C and tumor necrosis factor- (TNF-) in the pathogenesis of pterygia. Cultured conjunctival epithelial cells were treated with TNF-, and the gene expression levels of VEGFC were evaluated by quantitative polymerase chain reaction (qPCR) and VEGF-C protein expression levels were measured using an enzyme-linked immunosorbent assay (ELISA). In addition, using ELISA, we evaluated the VEGF-C protein expression in the supernatants of cultured conjunctival epithelial cells, in which we neutralized TNF- using anti-TNF- antibody. The gene expression of tumor necrosis factor receptor superfamily, member 1A (TNFRSF1A), known as TNF receptor 1 (TNFR1), was confirmed using reverse transcription PCR in cultured conjunctival epithelial cells. Immunofluorescence microscopy was used to examine the localization of VEGF-C and TNFR1 in pterygium tissues and TNFR1 expression in cultured conjunctival epithelial cells. Immunohistochemistry was used to examine the localization of TNFR1 in pterygia and normal conjunctival tissues. VEGFC gene expression increased in cultured conjunctival epithelial cells 24 h after the addition of TNF-. The secretion of VEGF-C protein was significantly increased 48 h after the stimulation of cultured conjunctival epithelial cells with TNF-. Increased VEGF-C protein secretion stimulated by TNF- was significantly reduced by anti-TNF- neutralizing antibody treatment. In cultured conjunctival epithelial cells, TNFRSF1A and TNFR1 were expressed. TNFR1 was immunolocalized in normal conjunctival tissues and in human pterygium tissues as well as in VEGF-C-positive epithelial cells from human pterygia. Our data demonstrate that TNF- mediates VEGF-C expression, which plays a critical role in the pathogenesis of pterygia.