Eun Chul Kim, M.D., Man Soo Kim, M.D. Department of Ophthalmology & Visual Science, College of Medicine, Catholic University of Korea, Seoul, Korea The authors have no financial interest in the subject matter of this poster.

 potent growth factor that stimulates the proliferation of blood vessels  Injured states, VEGF expression upregulated and corneal neovascularization  important player in nerve growth  In vitro :VEGF and its receptor are expressed by neurons and stimulate neurogenic, neuroprotective, and neurotrophic activities  VEGF  proliferation of cortical neurons,  protection of central and peripheral neurons from hypoxia-induced death  promotion of axonal outgrowth in peripheral neurons  Systemic inhibition of VEGF with bevacizumab may cause apoptosis in retinal neurons (Invest Ophthalmol Vis Sci. 2007;48:1773–1781.)

 Nerve growth factor promotes corneal healing: structural, biochemical, and molecular analyses of rat and human corneas. Invest Ophthalmol Vis Sci Apr;41(5):  The Use of Nerve Growth Factor in Surgical Wound Healing of the Cornea Ophthalmic Res 2006;38:177–181  Use of nerve growth factor to treat congenital neurotrophic corneal ulceration. Cornea Apr;25(3):352-5

 Reciprocal angiogenic and neurotrophic effects on blood vessels and neurons (Curr Pharm Des 2006;12:2609–2622.)  Capillary sprouting was promoted by NGF via the release of VEGF. (Proc Natl Acad Sci U S A 2001;98:4160–4165.)  Overexpression of NGF in brown adipose tissue of NGF-transgenic mice : elevated both mRNA and protein levels of VEGF and VEGFRs. (Histochem Cell Biol. 2006;125: )  Activated VEGF stimulated angiogenesis : blocked by NGF neutralizing antibodies. (Histochem Cell Biol. 2006;125: )  NGF stimulated the expression of VEGF isoforms in epithelial ovarian cancer and the effect was dose-dependent and inhibited by an NFG antibody and by a TrkA inhibitor. (Brain Res 2002;953:12–16.)

 To investigate whether anti–VEGF antibodies such as bevacizumab eyedrops have inhibitory effects on corneal epithelial wound healing and NGF expression in rats.  Necessity  Studies have indicated that VEGF plays a role in mediating corneal nerve repair 11 and that anti–VEGF delays corneal epithelial wound healing in rabbits. (Invest Ophthalmol Vis Sci. 2009;50: )  However, implications for anti–VEGF therapy for corneal wound healing by VEGF inhibition leading to NGF expression have not been explored.

 120 Sprague-Dawley male rats (250–300 g) were divided into two groups of 60 rats each..  Bevacizumab Group: instilled anti-VEGF(5 mg/ml) and antibiotics (Cravit ® : 0.5% levofloxacin) four times daily  Control Group : antibiotics eyedrops only four times daily  Corneal wound healing was evaluated by fluorescein staining at 0, 24, 48, and 72 hours after 5 mm-sized epithelial debridement.  Nerve growth factor (NGF) and vascular endothelial growth factor (VEGF) protein were measured in rat corneas by ELISA.  Immunofluorescent staining for NGF and VEGF were performed in rat corneas.  NGF mRNA and VEGF mRNA were measured in rat corneas by real time PCR.

 Wound healing in the bevacizumab group was lower than that in the control group at 24, 48, and 72 hours after debridement, respectively (*P < 0.05; n = 20 per group; Mann- Whitney U test). Bevacizumab Control 0 Hour 24 Hours 48 Hours72 Hours

Bevacizumab Control 0 Hour 24 Hours48 Hours 72 Hours Photographs in the bevacizumab group show subepithelial haze at 24 hours, subepithelial fibrosis at 48 hours, and slight stromal thinning at 72 hours after debridement. In the control group, however, epithelial defects were healed without any haze.

 The cornea was trephined with a 4.0-mm-diameter trephine before epithelial debridement and 24, 48, and 72 hours after wounding Corneal VEGF and NGF concentrations in the bevacizumab group were decreased compared to that in the control group at 24, 48, and 72 hours after debridement, respectively (*P < 0.05; n = 20 per group; Mann-Whitney U test).

 At 48 hours, immunofluorescent stains of VEGF and NGF were most strong in control corneas(DAPI counter stain ) NGF VEGF BevacizumabControl DAPI was used as nuclear staining (blue). Original magnification, 200X. At 48 hours, immunofluorescent stains of VEGF and NGF were most strong in control cornea, but no stain in bevacizumab cornea.

The control VEGF and NGF is regarded as a standard control (RQ = 1) VEGF and NGF mRNA expressions in bevacizumab group were significantly decreased compared with the control group at 24, 48, and 72 hours after debridement, respectively (*P < 0.05; n = 20 per group; Mann-Whitney U test)

 Corneal epithelial healing was delayed in Anti-VEGF group  VEGF and NGF was expressed after corneal damage.  VEGF and NGF was less expressed in Anti-VEGF group  VEGF and NGF was more expressed in control group  Anti-VEGF (Bevacizumab) may negatively affect corneal wound healing.  Anti-VEGF (Bevacizumab) may downregulate NGF expression.  Caution should be applied in damaged and thin cornea.