Expression of Vascular Adhesion Protein-1 in Ocular Tissues of Endotoxin-Induced Uveitis

Abstract

Purpose:: Vascular adhesion protein-1 (VAP-1), also known as the cell-surface enzyme semicarbazide-sensitive amine oxidase (SSAO), is an endothelial cell adhesion molecule involved in leukocyte recruitment. The vascular endothelium of a number of tissues express VAP-1 under normal and inflamed conditions, but the expression and localization of VAP-1 in ocular tissues has not been explored. Here we investigate the VAP-1 expression in inflamed ocular tissues using the Endotoxin-Induced Uveitis (EIU) model of ocular inflammation.

Methods:: EIU was induced in male Lewis rats by injection of 100µg LPS into one hind footpad. The expression level of VAP-1 and other adhesion molecules, P-selectin, intercellular adhesion molecule-1 (ICAM-1), and vascular cell adhesion molecule-1 (VCAM-1) was assessed in the retinas of EIU animals using reverse transcriptional polymerase chain reaction (RT-PCR). To further evaluate VAP-1 expression in the EIU retina, real time PCR was performed. SSAO enzymatic activity was measured in retinal and uveal tissue extracts and in the plasma of normal and EIU animals. Immunofluorescent staining was performed to examine the presence and localization of VAP-1 protein in ocular tissues.

Results:: RT-PCR and measurements of SSAO enzymatic activity revealed constitutive expression of VAP-1 in retinal and choroidal tissues under normal conditions. In line with previous reports, P-selectin and ICAM-1 were upregulated in retinal tissues after LPS stimulation. Real time PCR showed a 1.5-fold increase in the VAP-1 mRNA level in the retina 24 hours after LPS stimulation (P<0.05). Immunofluorescent staining showed VAP-1 expression on the vascular endothelial cells in sclera, choroid and retina. Furthermore, immunofluorescent staining revealed the translocation of VAP-1 from the cytosol to the luminal surface of vascular endothelial cells in the EIU retina.

Conclusions:: The current results demonstrate up-regulation of retinal VAP-1 mRNA and VAP-1 re-localization to the retinal endothelial surface during EIU. These data suggest that VAP-1 may play a role in leukocyte transmigration in the retina and in the pathogenesis of uveitis.

Keywords: uveitis-clinical/animal model • inflammation