Loss of Osteopontin Expression Reduces HSV-1-Induced Corneal Opacity

Abstract

Purpose: Corneal opacity and neovascularization (NV) are often described as outcomes of severe herpes simplex virus type 1 (HSV-1) infection. The current study investigated the role of colony-stimulating factor 1 receptor (CSF1R)+ cells and soluble factors in the progression of HSV-1-induced corneal NV and opacity. Methods: MaFIA mice were infected with 500 plaque-forming units of HSV-1 in the cornea following scarification. From day 10 to day 13 post-infection (pi), mice were treated with 40 µg/day of AP20187 (macrophage ablation) or vehicle intraperitoneally. For osteopontin (OPN) neutralization experiments, C57BL/6 mice were infected as above and treated with 2 µg of goat anti-mouse OPN or isotypic control IgG subconjunctivally every 2 days from day 4 to day 12 pi. Mice were euthanized on day 14 pi, and tissue was processed for immunohistochemistry to quantify NV and opacity by confocal microscopy and absorbance or detection of pro- and anti-angiogenic and inflammatory factors and cells by suspension array analysis and flow cytometry, respectively. Results: In the absence of CSF1R+ cells, HSV-1-induced blood and lymphatic vessel growth was muted. These results correlated with a loss in fibroblast growth factor type 2 (FGF-2) and an increase in OPN expression in the infected cornea. However, a reduction in OPN expression in mice did not alter corneal NV but significantly reduced opacity. Conclusions: Our data suggest that CSF1R+ cell depletion results in a significant reduction in HSV-1-induced corneal NV that correlates with the loss of FGF-2 expression. A reduction in OPN expression was aligned with a significant drop in opacity associated with reduced corneal collagen disruption.