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Abstract
Varicella zoster virus (VZV) latency in sensory and autonomic neurons has remained enigmatic and difficult to study, and experimental reactivation has not yet been achieved. We have previously shown that human embryonic stem cell (hESC)-derived neurons are permissive to a productive and spreading VZV infection. We now demonstrate that hESC-derived neurons can also host a persistent non-productive infection lasting for weeks which can subsequently be reactivated by multiple experimental stimuli. Quiescent infections were established by exposing neurons to low titer cell-free VZV either by using acyclovir or by infection of axons in compartmented microfluidic chambers without acyclovir. VZV DNA and low levels of viral transcription were detectable by qPCR for up to seven weeks. Quiescently-infected human neuronal cultures were induced to undergo renewed viral gene and protein expression by growth factor removal or by inhibition of PI3-Kinase activity. Strikingly, incubation of cultures induced to reactivate at a lower temperature (34°C) resulted in enhanced VZV reactivation, resulting in spreading, productive infections. Comparison of VZV genome transcription in quiescently-infected to productively-infected neurons using RNASeq revealed preferential transcription from specific genome regions, especially the duplicated regions. These experiments establish a powerful new system for modeling the VZV latent state, and reveal a potential role for temperature in VZV reactivation and disease.
Highlights
Herpes Zoster, which results from reactivation of latent varicella zoster virus (VZV) is a common and debilitating disease that is frequently complicated by acute pain, diverse neurological sequelae, vision problems and difficult-to-treat chronic pain known as post-herpetic neuralgia
We have previously reported that human embryonic stem cell (hESC)-derived neurons exposed to high MOI cell-free recombinant pOKA-derived Varicella zoster virus (VZV) with fluorescent protein reporters of viral protein expression, results in a spreading, productive infection [21]
In order to obtain non-productive persistent VZV infections in hESC-derived neurons, we exposed them to low PFU (0.001 MOI) of cellfree VZV in the presence of acyclovir (ACV) for 6 days
Summary
Herpes Zoster, which results from reactivation of latent varicella zoster virus (VZV) is a common and debilitating disease that is frequently complicated by acute pain, diverse neurological sequelae, vision problems and difficult-to-treat chronic pain known as post-herpetic neuralgia. The VZV latent state is established in human sensory neurons of ganglia along the entire neuraxis during primary infection and disease, chickenpox. The recent recognition that latent VZV genomes undergo viral transcription in ganglia following post mortem removal raised doubt as to what transcriptional events occur in the latent state [2]. Reports of immunohistochemical detection of VZV proteins in sections from latently-infected ganglia has been confounded by non-specific staining, lipofuschin granules and antibody cross-reactivity with blood group antigens [5]. The most commonly reported transcript in human ganglia is that for ORF63 [7]
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