Warner, Benjamin E
(2022)
Novel recombinant varicella-zoster virus to study infection and induction of hypersensitivity in rat models of postherpetic neuralgia.
Doctoral Dissertation, University of Pittsburgh.
(Unpublished)
Abstract
The mechanisms by which VZV causes pain during herpes zoster (HZ) and postherpetic neuralgia (PHN) are not well understood. Part of this stems from the lack of an animal model of reactivated HZ disease and the pain states that follow. However, primary inoculation of the rat foot- or whisker pad with VZV induces prolonged pain behaviors that resemble clinical PHN. To elucidate the infectious processes necessary for the induction of a pain state in this model, we have developed growth conditional VZV in which the turnover of essential proteins can be regulated by insertion of a degron. We report the degron system was effective for generating conditional knockout mutants for VZV proteins from immediate-early genes ORF4 and -63, and the late ORF9 gene. Inoculation of the recombinant viruses into the rat foot- or whisker pad revealed that VZV productive replication was dispensable for induction of hypersensitivity, whereas events early in the infectious process were essential. We then described a newly available transgenic reporter rat that conditionally expresses a tdTomato fluorescent gene which may help clarify VZV infection of rat tissues in vivo. The studies here characterize a recombinant VZV and methods for use in the reporter rat model. Finally, we described additional uses for the growth conditional VZV in two areas of VZV research that have proved difficult: the study of essential gene function and establishment of experimental latency in neuron cultures. The results established that the ORF9 growth conditional VZV may serve as a backbone for additional ORF9 gene mutations to determine essential protein functional domains, though further modifications to the system are required. The ORF4 growth conditional VZV was exploited for the establishment of a latent infection in human embryonic stem cell derived neurons. We showed that infection of neurons with the ORF4 virus was quiescent, resembled experimental latency, and was reactivatable with a chemical stimulus. This work presents an alternative to current latent infection methods that rely on acyclovir which may damage VZV genomes and render them incapable of reactivation.
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Details
| Item Type: |
University of Pittsburgh ETD
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| Status: |
Unpublished |
| Creators/Authors: |
|
| ETD Committee: |
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| Date: |
9 January 2022 |
| Date Type: |
Publication |
| Defense Date: |
8 October 2021 |
| Approval Date: |
9 January 2022 |
| Submission Date: |
8 December 2021 |
| Access Restriction: |
No restriction; Release the ETD for access worldwide immediately. |
| Number of Pages: |
158 |
| Institution: |
University of Pittsburgh |
| Schools and Programs: |
School of Medicine > Microbiology and Immunology |
| Degree: |
PhD - Doctor of Philosophy |
| Thesis Type: |
Doctoral Dissertation |
| Refereed: |
Yes |
| Uncontrolled Keywords: |
varicella-zoster virus, reactivation, herpes zoster, postherpetic neuralgia, degron, pain model |
| Date Deposited: |
10 Jan 2022 03:10 |
| Last Modified: |
10 Jan 2022 03:10 |
| URI: |
http://d-scholarship.pitt.edu/id/eprint/42021 |
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