Infiltrating Cells, Interferon-gamma and Intraocular Spread of HSV-1 after Anterior Chamber Injection

Abstract

Following uniocular anterior chamber (AC) inoculation of HSV-1 (KOS), the anterior segment of the injected eye becomes inflamed and infected; however, virus does not spread from the anterior segment to infect the retina of the injected eye. The overall goal of this study was to identify interferons (IFNs) and early infiltrating cells which may play a role in protecting the retina of the ipsilateral (injected) eye. Female BALB/c, IFNy-/- and macrophage depleted (clodronate, CI2MBP treated) mice were injected in one AC with 3*104 - 6x104 PFU of HSV-1 (KOS). Mice were killed at various time points ranging from 12 to 120 hours post injection (p.i.). The injected eyes were enucleated, snap frozen and frozen sections were stained with antibodies specific for HSV-1, IFNy, Mac-1 (CD11b), Gr-1, CD49b, F4/80, CD4, CD8 and CD11c. The same antibodies were also used to stain freshly isolated single-cell suspensions from the eye or spleen for flow cytometry. Additionally, whole injected eyes were used to determine gene expression levels of IFNs and IFN associated genes. In the anterior segment of the injected eye, the ciliary body and iris were virus infected and inflamed, and infiltrating cells increased during the period of observation. Mac-1 + and F4/80+ cells colocalized with IFNy in the anterior segment and Mac-1 + cells increased in the injected eye beginning at 24 hours p.i. and continuing through 72 hours p.i. Although virus staining was increased in the ciliary body of macrophage depleted mice at 48 and 72 hours p.i., destructive retinitis was not observed in the injected eye of these animals. IFNy gene expression was up regulated in injected eyes of BALB/c mice from 48 to 120 hours p.i., and while HSV-1 infection of IFNy-/- mice resulted in increased virus staining in the ciliary body, destructive retinitis was rarely observed in IFNy-/- mice. Microglia and IFNy play important roles in the immune response to virus infection, but depletion of single cell types or cytokines did not result in early panretinal HSV-1 infection in the injected eye. Taken together, these findings support the idea that the timing and appearance of different cell types and cytokines is critical to protection of the retina of the injected eye from infection due to direct spread of virus; however, it is likely that during the innate immune response in the eye, other cell types and cytokines can compensate for the absence of a single cell type or of a single cytokine.