Herpesvirus Entry Mediator on Radiation-Resistant Cell Lineages Promotes Ocular Herpes Simplex Virus 1 Pathogenesis in an Entry-Independent Manner

Abstract

Ocular herpes simplex virus 1 (HSV-1) infection leads to a potentially blinding immunoinflammatory syndrome, herpes stromal keratitis (HSK). Herpesvirus entry mediator (HVEM), a widely expressed tumor necrosis factor (TNF) receptor superfamily member with diverse roles in immune signaling, facilitates viral entry through interactions with viral glycoprotein D (gD) and is important for HSV-1 pathogenesis. We subjected mice to corneal infection with an HSV-1 mutant in which HVEM-mediated entry was specifically abolished and found that the HVEM-entry mutant produced clinical disease comparable to that produced by the control virus. HVEM-mediated induction of corneal cytokines, which correlated with an HVEM-dependent increase in levels of corneal immune cell infiltrates, was also gD independent. Given the complexity of HVEM immune signaling, we used hematopoietic chimeric mice to determine which HVEM-expressing cells mediate HSV-1 pathogenesis in the eye. Regardless of whether the donor was a wild-type (WT) or HVEM knockout (KO) strain, HVEM KO recipients were protected from ocular HSV-1, suggesting that HVEM on radiation-resistant cell types, likely resident cells of the cornea, confers wild-type-like susceptibility to disease. Together, these data indicate that HVEM contributes to ocular pathogenesis independently of entry and point to an immunomodulatory role for this protein specifically on radiation-resistant cells.

Murine Corneal Inflammation and Nerve Damage After Infection With HSV-1 Are Promoted by HVEM and Ameliorated by Immune-Modifying Nanoparticle Therapy

“…Plaque assays were performed and virus was propagated on African green monkey kidney cells (Vero) as previously described 18 ; HSV-1 strain 17 was obtained from David Leib (Dartmouth Medical School, Hanover, NH, USA).…”

“…Corneas were inoculated with 2 × 10 6 plaque forming units (PFU) of HSV-1 in 5 μL Dulbecco's modified Eagle's medium (DMEM) after scarification with a 25G needle in a crosshatch pattern as previously described; mock-infected controls instead received a Vero cell lysate after scarification to control for any effects mediated by free DNA and other cellular debris that may be present in the viral preparation. 17,18 …”

“…22 Herpes virus entry mediator is also found on a wide range of leukocytes, including neutrophils, monocytes, dendritic cells, and CD4 + and CD8 + T cells, although the expression of HVEM on these cells once in the cornea has not been assessed. 23,24 We recently have found that HVEM-dependent disease after HSV-1 infection at the corneal surface is mediated by a radiation-resistant cell type or types, 18 leading us to hypothesize that HVEM on corneal epithelial cells, stromal fibroblasts, or resident stromal macrophages, which incompletely turn over after irradiation, 25 may promote pathogenesis in our adoptive transfer model. We investigated the expression of HVEM on a variety of cell lineages before, during, and after acute HSV-1 infection and found that corneal monocytes/macrophages are the first population to express HVEM, followed by polymorphonuclear neutrophils (PMN), double-negative (DN) T cells, and CD4 + T cells.…”

“…16 While HVEM is required for ocular pathogenesis in vivo, 17 the immunomodulatory effects of this receptor are more potent than its entry functions: HVEM increases corneal inflammatory cytokine production and leukocytic infiltration independently of viral entry. 18–20 Herpes virus entry mediator knockout (KO) mice have lower viral loads in eye swabs and subsequent spread to the nervous system, and lessened or absent clinical symptoms, including neurologic disease, periocular lesions, and mortality. 17,18,21 …”

“…18–20 Herpes virus entry mediator knockout (KO) mice have lower viral loads in eye swabs and subsequent spread to the nervous system, and lessened or absent clinical symptoms, including neurologic disease, periocular lesions, and mortality. 17,18,21 …”