Doctoral Dissertations

Date of Award

8-1998

Degree Type

Dissertation

Degree Name

Doctor of Philosophy

Major

Comparative and Experimental Medicine

Major Professor

Barry T. Rouse

Committee Members

David Slauson, Philip Bochsler, Albert Ichiki

Abstract

Corneal infection with Herpes simplex virus (HSV) in susceptible mice induces an immunopathological response termed Herpetic Stromal Keratitis (HSK). Studies were initially focused on characterizing early cellular events in the eye and kinetics of viral clearance from the eye following introcular infection with HSV-1. A biphasic influx of neutrophils (PMN) was observed, the initial one lasting for about 48-72 hours post infection and the second surge appearing at around day 8-9. The disappearance of the virus correlated with the disappearance of the PMN. The role of PMN in HSK was evaluated by depleting PMN in CD4-I- T cell reconstituted SCID mice. There was significantly less incidence and severity of HSK in CD4+ T cell reconstituted SCID mice which were depleted of PMN as compared to controls. Such PMN depletion in HSV infected mice however, caused moderate to severe periocular disease, progressive cachexia, and high mortality due to encephalitis. Virus was recovered from the eyes for longer periods and virus titers in the brains of PMN depleted animals were significantly higher than those of the controls. These results suggest that HSV infection of the cornea generates a chemotactic gradient for PMN which subserve an anti-viral role and perhaps serve directly or indirectly as agonists in perpetuating a CD4+ T cell mediated inflammatory reaction.

Secondly, studies on intraocular chemokines and proinflammatory cytokines were performed with reverse transcription polymerase chain reaction (RT-PCR) technique with a view to correlating the observed cellular events with the chemokine profile. Studies were also done to ascertain if the failure of HSV replication to cause clinical HSK was linked to their ability to cause expression of chemokines and proinflammatory cytokines. While productive infection resulted in rapid upregulation and sustained expression of chemokines such as N51/KC, MlPl-α, MIPl-β, MIP-2, MCP-1 and cytokines such as IL-1, Il\L-6, IL-12, and TNF-α, expression of such inflammatory mediators was minimal and transient following unproductive infection. Expression of MlPl-αand lymphotactin along with a biphasic expression of MIP-2, IL-6 and TNF-α were seen only with productive infection and therefore suggests they may play a role in the recruitment of CD4+ T cells and PMN respectively. Productive infection was followed by approximately 50 fold increase in the initial PMN recruitment in the cornea. These data suggest that replication induced proinflammatory milieu in the cornea is crucial for the subsequent progression of HSK.

Finally, the role of antigen recognition in SCID mouse reconstitution experiments was explored by transferring HSV primed and naive CD4-I- T cells from naturally resistant mice into HSV infected SCID mice. While euthymic mice such as C.B-17 and C57B1/6 mice were resistant to HSK, even though their viral clearance rates and immune responses were similar to those of susceptible strains, adoptive transfer of their CD4+ T cells or whole splenocyte populations into SCID mice mediated HSK in the recipients. These data show that the genetically HSK resistant mice possess a CD4+ T cell repertoire capable of causing HSK but yet are kept under 'regulatory control'. The data imply that the HSV infected SCID mouse provides a proinflammatory 'microenvironment' that overrides the regulatory controls and/or cause activation of quiescent cells in a non-specific manner to orchestrate HSK.

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