Resolution of herpes simplex virus reactivation in vivo results in neuronal destruction

Abstract

A fundamental question in herpes simplex virus (HSV) pathogenesis is the consequence of viral reactivation to the neuron. Evidence supporting both post-reactivation survival and demise is published. The exceedingly rare nature of this event at the neuronal level in the sensory ganglion has limited direct examination of this important question. In this study, an in-depth in vivo analysis of the resolution of reactivation was undertaken. Latently infected C57BL/6 mice were induced to reactivate in vivo by hyperthermic stress. Infectious virus was detected in a high percentage (60-80%) of the trigeminal ganglia from these mice at 20 hours post-reactivation stimulus, but declined by 48 hours post-stimulus (0-13%). With increasing time post-reactivation stimulus, the percentage of reactivating neurons surrounded by a cellular cuff increased, which correlated with a decrease in detectable infectious virus and number of viral protein positive neurons. Importantly, in addition to intact viral protein positive neurons, fragmented viral protein positive neurons morphologically consistent with apoptotic bodies and containing cleaved caspase-3 were detected. The frequency of this phenotype increased through time post-reactivation. These fragmented neurons were surrounded by Iba1 + cells, consistent with phagocytic removal of dead neurons. Evidence of neuronal destruction post-reactivation prompted re-examination of the previously reported non-cytolytic role of T cells in controlling reactivation. Latently infected mice were treated with anti-CD4/CD8 antibodies prior to induced reactivation. Neither infectious virus titers nor neuronal fragmentation were altered. In contrast, when viral DNA replication was blocked during reactivation, fragmentation was not observed even though viral proteins were expressed. Our data demonstrate that at least a portion of reactivating neurons are destroyed. Although no evidence for direct T cell mediated antigen recognition in this process was apparent, inhibition of viral DNA replication blocked neuronal fragmentation. These unexpected findings raise new questions about the resolution of HSV reactivation in the host nervous system. Herpes simplex virus (HSV)is an endemic human pathogen that establishes latency in neurons and can periodically reactivate over the lifetime of the host. Whether or not neurons survive post-reactivation is controversial and has significant implications for longterm infection. HSV reactivation events can be characterized in mice, which maintain the complexity of host-pathogen interactions, with the goal to provide insight into how the virus behaves in the nervous system. In this report, it is shown that the elimination of infectious virus following reactivation in vivo corresponded with a highly focused cellular response and destruction of the neuron containing HSV proteins. The role of T cells in this response was investigated. Previous work identified T cells as major regulators of HSV reactivation. However, neuronal destruction was still observed when T cell anti...

Neuronal hyperexcitability is a DLK-dependent trigger of herpes simplex virus reactivation that can be induced by IL-1

“…This indicates that reactivation proceeds via a Phase I-wave of gene expression in response to multiple different stimuli. However, we note that there may be differences in the mechanism and kinetics of reactivation with different stimuli and/or strains of HSV-1 as reactivation triggered by axotomy or heat shock following infection with a more pathogenic strain of HSV may bypass Phase I or occur more rapidly, making Phase I difficult to detect ( Cliffe and Wilson, 2017 ; Liang et al, 2009 ; Doll et al, 2020 ). It will be especially interesting to determine in the future whether there are differences in the progression to reactivation with different strains of HSV, further elucidate the underlying progression to reactivation and requirements for Phase I.…”

“…CD8 T cells have been linked to controlling HSV reactivation ( Divito et al, 2006 ), and regulatory T cells have been found to facilitate HSV reactivation in an in vivo model by suppressing CD8 T cells ( Yu et al, 2018 ). However, a recent in vivo study found that Iba+ phagocytic cells play a key role clearing reactivating neurons ( Doll et al, 2020 ), which required viral DNA replication. HSV latency is not fully silent, with detectible lytic gene expression in infected neurons ( Ma et al, 2014 ; Singh and Tscharke, 2020 ).…”

The Potential Role of Herpes Simplex Virus Type 1 and Neuroinflammation in the Pathogenesis of Alzheimer's Disease

“…Several animal models of peripheral HSV1 infection already exist such as the rabbit and mouse eye models and the guinea pig models ( 164 , 165 ). These models have been widely used to study multiple aspects of HSV1 biology, including viral reactivation in the PNS ( 166 – 170 ). However, they have rarely been used to investigate the direct role of herpesvirus-induced neuroinflammatory responses in the PNS in initiating AD in the brain.…”

Neuronal hyperexcitability is a DLK-dependent trigger of HSV-1 reactivation that can be induced by IL-1

“…This indicates that reactivation proceeds via a Phase 1-wave of gene expression in response to multiple different stimuli. However we note that there may be differences in the mechanism and kinetics of reactivation with different stimuli and/or strains of HSV-1 as reactivation triggered by axotomy may bypass Phase I 19,20 and reactivation induced in vivo by heat shock with a more pathogenic strain of HSV triggered more rapid reactivation 71 . It will be especially interesting to determine in the future whether there are differences in the progression to reactivation with different strains of HSV.…”