Date of Graduation

12-2022

Document Type

Dissertation

Degree Name

Doctor of Philosophy in Cell & Molecular Biology (PhD)

Degree Level

Graduate

Department

Cell & Molecular Biology

Advisor/Mentor

Ceballos, Ruben M.

Committee Member

Durdik, Jeannine M.

Second Committee Member

McNabb, David S.

Third Committee Member

Nakanishi, Nagayasu

Fourth Committee Member

Shew, Woodrow L.

Keywords

Cell tropism; Epilepsy; Human herpesvirus 6; Immunological response; Neural stem cells; Roseolovirus

Abstract

Within the family Herpesviridae, sub-family β-herpesvirinae, and genus Roseolovirus, there areonly three human herpesviruses that have been described: HHV-6A, HHV-6B, and HHV-7. Initially, HHV-6A and HHV-6B were considered as two variants of the same virus (i.e., HHV6). Despite high overall genetic sequence identity (~90%), HHV-6A and HHV-6B are now recognized as two distinct viruses. Sequence divergence (e.g., >30%) in key coding regions and significant differences in physiological and biochemical profiles (e.g., use of different receptors for viral entry) underscores the conclusion that HHV-6A and HHV-6B are distinct viruses of the β-herpesvirinae. Despite these viruses being implicated as causative agents in several nervous system disorders (e.g., multiple sclerosis, epilepsy, and chronic fatigue syndrome), the mechanisms of action and relative contributions of each virus to neurological dysfunction are unclear. Unresolved questions regarding differences in cell tropism, receptor use and binding affinity (i.e., CD46 versus CD134), host neuro-immunological responses, and relative virulence between HHV-6A versus HHV-6B prevent a complete characterization. Although it has been shown that both HHV-6A and HHV-6B can infect glia (and, recently, cerebellar Purkinje cells), cell tropism of HHV-6A versus HHV-6B for different nerve cell types remains vague. In this study, we show that both viruses can infect different nerve cell types (i.e., glia versus neurons) and different neurotransmitter phenotypes derived from differentiated human neural stem cells. As demonstrated by immunofluorescence, HHV-6A and HHV-6B productively infect VGluT1- containing cells (i.e., glutamatergic neurons) and dopamine-containing cells (i.e., dopaminergic neurons). However, neither virus appears to infect GAD67-containing cells (i.e., GABAergic neurons). As determined by qPCR, the expression of immunological factors (e.g., cytokines) in cells infected with HHV-6A versus HHV6-B also differs. These data, along with morphometric and image analyses of infected differentiated neural stem cell cultures, indicate that while HHV6B may have a greater opportunity for transmission, HHV-6A induces more severe cytopathic effects (e.g., syncytia) at the same post-infection end points. Cumulatively, results suggest that HHV-6A is more virulent than HHV-6B in susceptible cells, while neither virus productively infects GABAergic cells. Consistency between these in vitro data and in vivo experiments would provide new insights into potential mechanisms for HHV6-induced epileptogenesis.

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