Inhibition of Experimental Corneal Neovascularization by the Tight Junction Protein ZO-1

<b><i>Purpose:</i></b> To explore the effects of the tight junction protein zonula occludens 1 (ZO-1) on experimental corneal neovascularization (CNV). <b><i>Methods:</i></b> CNV models were established in the left eyes of BALB/c mice using NaOH. Anti-ZO-1 neutralizing antibody was topically applied to the burnt corneas after modeling thrice a day for 1 week. CD31 expression was analyzed to calculate the ratio of CNV number to area using a corneal whole-mount fluorescent immunohistochemical assay. Messenger ribonucleic acid (mRNA) and protein expression levels of ZO-1, vascular endothelial growth factor (VEGF), interleukin (IL)-1β, IL-6, IL-8, IL-18, monocyte chemoattractant protein-1 (MCP-1), tumor necrosis factor alpha (TNF-α), phosphorylated protein kinase C (pPKC), and clusterin in burned corneas were detected by reverse transcriptase polymerase chain reaction (PCR) and western blot analyses. Infiltration of neutrophils, macrophages, and progenitor cells was examined by flow cytometry. <b><i>Results:</i></b> CNV was obviously greater in 45 s than in 15 s alkali injury group. In another experiment, CNV was obviously greater in the ZO-1 antibody group than in the vehicle-treated group. Corneal mRNA and protein expression levels of VEGF, IL-1β, IL-6, IL-8, IL-18, and MCP-1 were significantly higher in the ZO-1 antibody group than in the control group. Infiltration of neutrophils, macrophages, and progenitor cells was significantly greater in the ZO-1 antibody group than in the control group. TNF-α expression was much higher in 45 s than in 15 s alkali injury group. However, protein expression of pPKC and clusterin was much lower in 45 s than in 15 s alkali injury group. <b><i>Conclusions:</i></b> Anti-ZO-1 neutralizing antibody-treated mice exhibited enhanced alkali-induced CNV through enhanced intracorneal infiltration of progenitor and inflammatory cells.