HSV Latency Research Articles

The significance of reactivation of latent herpes simplex viruses type 1 and 2 in the etiopathogenesis of chronic inflammatory eye diseases and the development of late post-uveal complications

Herpes simplex viruses type 1 and 2 (HSV-1 and HSV-2) cause widespread lifelong infections. These characteristics of herpes simplex virus infections (HSVI) are associated with the presence of two phases in the infectious cycle: the lytic infection phase, which involves the formation of new viral particles, and the latent infection phase, during which the HSV remains in cells in a hidden form that is less accessible to the immune system. The lytic and latent phases of HSVI differ in the form of the viral genome, its localization, number of active viral genes, and expressed viral products. Lytic infection primarily occurs in epithelial cells, while the reservoir of latent virus is the nuclei of sensory neurons of the ganglia innervating the site of lytic infection. “Abortive” infection of Hela transfected cells is considered as experimental analogue of latent neuron infection, in which HSV-1 genomes were detected in the nuclei of surviving cells for up to 5 weeks, retaining the ability to reactivate and induce lytic infection. Frequent subclinical reactivation of HSV with the release of infectious virus has been identified in individuals who have had herpes infections and in healthy people with chronic HSVI. Relapses of herpes disease occur much less frequently. Intercurrent diseases are one of the leading factors in the reactivation of latent HSV. Reactivation of HSV can trigger the exacerbation or development of non-herpetic diseases, complicating their course. Inclusion of specific antiviral agents in the complex therapy of such patients shortened the time till remission onset. HSV reactivation is a prognostically unfavorable factor not only in active eye disease but also in clinical remission. According to our data, subclinical HSV reactivation in patients with remission of uveitis increased the systemic production of pro-inflammatory and angiogenic chemokines, thus contributing to the chronicization of a low-grade inflammatory process and the development of late post-uveal complications. The question of prescribing specific antiviral therapy to such patients remains relevant.

The importance of IFNα2A (Roferon-A) in HSV-1 latency and T cell exhaustion in ocularly infected mice.

Published studies have generated compelling results indicating that type I IFN modulates function of HSV-1 latency-associated transcript (LAT). One member of type I IFN is IFNα2A also called Roferon-A). IFNα2A has been used in monotherapy or in combination therapy with other drugs to treat viral infections and different kinds of cancer in humans. The goal of this study was to determine whether the absence of IFNα2A affects primary and latent infections in ocularly infected mice. Therefore, we generated a mouse strain lacking IFNα2A expression (IFNα2A-/-). Ocular HSV-1 replication, IFN and immune cell expressions on days 3 and 5 post infection (PI), as well as eye disease, survival, latency-reactivation, and T cell exhaustion were evaluated in ocularly infected IFNα2A-/- and wild type (WT) control mice. Absence of IFNα2A did not affect other members of the IFNα family but it affected IFNβ and IFNγ expressions as well as some immune cells on day 5 PI compared to WT mice. Viral replication in the eye, eye disease, and survival amongst ocularly infected IFNα2A-/- mice were similar to that of WT infected mice. The absence of IFNα2A significantly reduced the levels of latency and T cell exhaustion but not time of reactivation compared with control mice. Our results suggest that blocking IFNα2A expression may be a useful tool in reducing latency and the subsequent side effects associated with higher levels of latency.

Presence of CD80 and Absence of LAT in Modulating Cellular Infiltration and HSV-1 Latency.

CD80 is the best-known costimulatory molecule for effective T cell functions. Many different reports have summarized the role of CD80 in HSV-1 and its functions in maintaining adaptive immunity, which is the main player in causing herpes stromal keratitis (HSK). To determine the effects of absence or overexpression of CD80 in HSV-1 infection, we infected CD80-/- and WT mice with a recombinant HSV-1 expressing murine CD80 (HSV-CD80) in place of the latency associated transcript (LAT). Parental dLAT2903 virus lacking LAT was used as a control. After infection, critical components of infection like virus replication, eye disease, early cellular infiltrates into the corneas and trigeminal ganglia (TG), latency-reactivation in the infected mice were determined. Our findings reveal that the absence of CD80 in the CD80-/- mice infected with both viruses did not affect the viral titers in the mice eyes or eye disease, but it played a significant role in critical components of HSV-induced immunopathology. The WT mice infected with dLAT2903 virus had significantly higher levels of latency compared with the CD80-/- mice infected with dLAT2903 virus, while levels of latency as determined by gB DNA expression were similar between the WT and CD80-/- mice infected with HSV-CD80 virus. In contrast to the differences in the levels of latency between the infected groups, the absence of CD80 expression in the CD80-/- mice or its overexpression by HSV-CD80 virus did not have any effect on the time of reactivation. Furthermore, the absence of CD80 expression contributed to more inflammation in the CD80-/--infected mice. Overall, this study suggests that in the absence of CD80, inflammation increases, latency is reduced, but reactivation is not affected. Altogether, our study suggests that reduced latency correlated with reduced levels of inflammatory molecules and blocking or reducing expression of CD80 could be used to mitigate the immune responses, therefore controlling HSV-induced infection.

The immunobiology of corneal HSV-1 infection and herpetic stromal keratitis.

SUMMARYHuman alphaherpesvirus 1 (HSV-1) is a highly successful neurotropic pathogen that primarily infects the epithelial cells lining the orofacial mucosa. After primary lytic replication in the oral, ocular, and nasal mucosal epithelial cells, HSV-1 establishes life-long latency in neurons within the trigeminal ganglion. Patients with compromised immune systems experience frequent reactivation of HSV-1 from latency, leading to virus entry in the sensory neurons, followed by anterograde transport and lytic replication at the innervated mucosal epithelial surface. Although recurrent infection of the corneal mucosal surface is rare, it can result in a chronic immuno-inflammatory condition called herpetic stromal keratitis (HSK). HSK leads to gradual vision loss and can cause permanent blindness in severe untreated cases. Currently, there is no cure or successful vaccine to prevent latent or recurrent HSV-1 infections, posing a significant clinical challenge to managing HSK and preventing vision loss. The conventional clinical management of HSK primarily relies on anti-virals to suppress HSV-1 replication, anti-inflammatory drugs (such as corticosteroids) to provide symptomatic relief from pain and inflammation, and surgical interventions in more severe cases to replace damaged cornea. However, each clinical treatment strategy has limitations, such as local and systemic drug toxicities and the emergence of anti-viral-resistant HSV-1 strains. In this review, we summarize the factors and immune cells involved in HSK pathogenesis and highlight alternate therapeutic strategies for successful clinical management of HSK. We also discuss the therapeutic potential of immunoregulatory cytokines and immunometabolism modulators as promising HSK therapies against emerging anti-viral-resistant HSV-1 strains.

APOE4, activation of latent form HSV-1, A beta accumulation and AD progression

The role of APOE4 in Alzheimers disease progression is an area of ongoing research. The core objective of this paper is to investigate the intricate interplay between APOE4, HSV-1, A, and AD progression. Methodology involves assessing A Levels, Spatial Recognition Memory, and Exploratory Behavior in HSV-1-infected Fx5AD Mice with APOE4, alongside monitoring specific biomarkers to confirm HSV-1 activation status. Our expected experimental outcomes suggest that APOE4 accelerates latent HSV-1 activation, leading to heightened A aggregation. Thus, we hypothesize that APOE4 may expedite latent HSV-1 activation, thereby facilitating A accumulation and AD advancement. The investigation of relationship between APOE4, HSV-1, A, and AD progression could give valuable insights into further experiments and treatments of AD.

Gene editing for latent herpes simplex virus infection reduces viral load and shedding in vivo

Anti-HSV therapies are only suppressive because they do not eliminate latent HSV present in ganglionic neurons, the source of recurrent disease. We have developed a potentially curative approach against HSV infection, based on gene editing using HSV-specific meganucleases delivered by adeno-associated virus (AAV) vectors. Gene editing performed with two anti-HSV-1 meganucleases delivered by a combination of AAV9, AAV-Dj/8, and AAV-Rh10 can eliminate 90% or more of latent HSV DNA in mouse models of orofacial infection, and up to 97% of latent HSV DNA in mouse models of genital infection. Using a pharmacological approach to reactivate latent HSV-1, we demonstrate that ganglionic viral load reduction leads to a significant decrease of viral shedding in treated female mice. While therapy is well tolerated, in some instances, we observe hepatotoxicity at high doses and subtle histological evidence of neuronal injury without observable neurological signs or deficits. Simplification of the regimen through use of a single serotype (AAV9) delivering single meganuclease targeting a duplicated region of the HSV genome, dose reduction, and use of a neuron-specific promoter each results in improved tolerability while retaining efficacy. These results reinforce the curative potential of gene editing for HSV disease.

Models of Herpes Simplex Virus Latency

Our current understanding of HSV latency is based on a variety of clinical observations, and in vivo, ex vivo, and in vitro model systems, each with unique advantages and drawbacks. The criteria for authentically modeling HSV latency include the ability to easily manipulate host genetics and biological pathways, as well as mimicking the immune response and viral pathogenesis in human infections. Although realistically modeling HSV latency is necessary when choosing a model, the cost, time requirement, ethical constraints, and reagent availability are also equally important. Presently, there remains a pressing need for in vivo models that more closely recapitulate human HSV infection. While the current in vivo, ex vivo, and in vitro models used to study HSV latency have limitations, they provide further insights that add to our understanding of latency. In vivo models have shed light on natural infection routes and the interplay between the host immune response and the virus during latency, while in vitro models have been invaluable in elucidating molecular pathways involved in latency. Below, we review the relative advantages and disadvantages of current HSV models and highlight insights gained through each.

Role of PD-1 and immune cells in HSV infection and latency.

A large proportion of the world's population is infected with HSV-1. Antiviral CD8+ T cells and CD8α+ dendritic cells are closely related to HSV-1 infection and latency. Latency-associated transcript of HSV-1 and PD-1are involved in the regulation of latency and reactivation of HSV-1. Here, the role of latency-associated transcript, PD-1, CD8+ T cellsand CD8α+ dendritic cells in HSV-1 infection, the inter-relationships betweenthemand how these interactions leadto latency are discussed, possibly providing new ideas for the treatment of HSV-1 infection.

A viral lncRNA tethers HSV-1 genomes at the nuclear periphery to establish viral latency.

Herpes simplex virus 1 (HSV-1) establishes lifelong latency in neuronal cells. Following a stressor, the virus reactivates from latency, virus is shed at the periphery and recurrent disease can occur. During latency, the viral lncRNA termed the latency-associated transcript (LAT) is known to accumulate to high abundance. The LAT is known to impact many aspects of latency though the molecular events involved are not well understood. Here, we utilized a human neuronal cell line model of HSV latency and reactivation (LUHMES) to identify the molecular-binding partners of the LAT during latency. We found that the LAT binds to both the cellular protein, TMEM43, and HSV-1 genomes in LUHMES cells. Additionally, we find that knockdown of TMEM43 prior to infection results in a decreased ability of HSV-1 to establish latency. This work highlights a potential mechanism for how the LAT facilitates the establishment of HSV-1 latency in human neurons.

Intermittent therapy with helicase-primase inhibitor IM-250 efficiently controls recurrent herpes disease and reduces reactivation of latent HSV

Herpes is a contagious life-long infection with persistently high incidence and prevalence, causing significant disease worldwide. Current therapies have efficacy against active HSV infections but no impact on the latent viral reservoir in neurons. Thus, despite treatment, disease recurs from latency and the infectious potential remains unaffected within patients.Here, efficacy of the helicase-primase inhibitor (HPI) IM-250 against chronic neuronal HSV infections utilizing two classic herpes in vivo latency/reactivation animal models (intravaginal guinea pig HSV-2 infection model and ocular mouse HSV-1 infection model) is presented. Intermittent therapy of infected animals with 4–7 cycles of IM-250 during latency silences subsequent recurrences analyzed up to 6 months. In contrast to common experience, our studies show that the latent reservoir is indeed accessible to antiviral therapy altering the latent viral reservoir such that reactivation frequency can be reduced significantly by prior IM-250 treatment.We provide evidence that antiviral treatment during HSV latency can reduce future reactivation from the latent reservoir, supporting a conceptual shift in the antiviral field, and reframing what is achievable with respect to therapy of latent neuronal HSV infections.

Binding of herpesvirus entry mediator (HVEM) and HSV-1 gD affect reactivation but not latency levels.

Previously we reported that the HSV-1 latency associated transcript (LAT) specifically upregulates the cellular herpesvirus entry mediator (HVEM) but no other known HSV-1 receptors. HSV-1 glycoprotein D (gD) binds to HVEM but the effect of this interaction on latency-reactivation is not known. We found that the levels of latent viral genomes were not affected by the absence of gD binding to HVEM. However, reactivation of latent virus in trigeminal ganglia explant cultures was blocked in the absence of gD binding to HVEM. Neither differential HSV-1 replication and spread in the eye nor levels of latency influenced reactivation. Despite similar levels of latency, reactivation in the absence of gD binding to HVEM correlated with reduced T cell exhaustion. Our results indicate that HVEM-gD signaling plays a significant role in HSV-1 reactivation but not in ocular virus replication or levels of latency. The results presented here identify gD binding to HVEM as an important target that influences reactivation and survival of ganglion resident T cells but not levels of latency. This concept may also apply to other herpesviruses that engages HVEM.

Palmer Station, Antarctica: A ground-based spaceflight analog suitable for validation of biomedical countermeasures for deep space missions

Astronauts are known to exhibit a variety of immunological alterations during spaceflight including changes in leukocyte distribution and plasma cytokine concentrations, a reduction in T-cell function, and subclinical reactivation of latent herpesviruses. These alterations are most likely due to mission-associated stressors including circadian misalignment, microgravity, isolation, altered nutrition, and increased exposure to cosmic radiation. Some of these stressors may also occur in terrestrial situations. This study sought to determine if crewmembers performing winterover deployment at Palmer Station, Antarctica, displayed similar immune alterations. The larger goal was to validate a ground analog suitable for the evaluation of countermeasures designed to protect astronauts during future deep space missions. For this pilot study, plasma, saliva, hair, and health surveys were collected from Palmer Station, Antarctica, winterover participants at baseline, and at five winterover timepoints. Twenty-six subjects consented to participate over the course of two seasons. Initial sample processing was performed at Palmer, and eventually stabilized samples were returned to the Johnson Space Center for analysis. A white blood cell differential was performed (real time) using a fingerstick blood sample to determine alterations in basic leukocyte subsets throughout the winterover. Plasma and saliva samples were analyzed for 30 and 13 cytokines, respectively. Saliva was analyzed for cortisol concentration and three latent herpesviruses (DNA by qPCR), EBV, HSV1, and VZV. Voluntary surveys related to general health and adverse clinical events were distributed to participants. It is noteworthy that due to logistical constraints caused by COVID-19, the baseline samples for each season were collected in Punta Arenas, Chile, after long international travel and during isolation. Therefore, the Palmer pre-mission samples may not reflect a true normal ‘baseline’. Minimal alterations were observed in leukocyte distribution during winterover. The mean percentage of monocyte concentration elevated at one timepoint. Plasma G-CSF, IL1RA, MCP-1, MIP-1β, TNFα, and VEGF were decreased during at least one winterover timepoint, whereas RANTES was significantly increased. No statistically significant changes were observed in mean saliva cytokine concentrations. Salivary cortisol was substantially elevated throughout the entire winterover compared to baseline. Compared to shedding levels observed in healthy controls (23%), the percentage of participants who shed EBV was higher throughout all winterover timepoints (52–60%). Five subjects shed HSV1 during at least one timepoint throughout the season compared to no subjects shedding during pre-deployment. Finally, VZV reactivation, common in astronauts but exceptionally rare in ground-based stress analogs, was observed in one subject during pre-deployment and a different subject at WO2 and WO3. These pilot data, somewhat influenced by the COVID-19 pandemic, do suggest that participants at Palmer Station undergo immunological alterations similar to, but likely in reduced magnitude, as those observed in astronauts. We suggest that winterover at Palmer Station may be a suitable test analog for spaceflight biomedical countermeasures designed to mitigate clinical risks for deep space missions.

Role of IL-27 in HSV-1-Induced Herpetic Stromal Keratitis.

Herpetic stromal keratitis (HSK) is a painful and vision-impairing disease caused by recurrent HSV-1 infection of the cornea. The virus replication in the corneal epithelium and associated inflammation play a dominant role in HSK progression. Current HSK treatments targeting inflammation or virus replication are partially effective and promote HSV-1 latency, and long-term use can cause side effects. Thus, understanding molecular and cellular events that control HSV-1 replication and inflammation is crucial for developing novel HSK therapies. In this study, we report that ocular HSV-1 infection induces the expression of IL-27, a pleiotropic immunoregulatory cytokine. Our data indicate that HSV-1 infection stimulates IL-27 production by macrophages. Using a primary corneal HSV-1 infection mouse model and IL-27 receptor knockout mice, we show that IL-27 plays a critical role in controlling HSV-1 shedding from the cornea, the optimum induction of effector CD4+ T cell responses, and limiting HSK progression. Using invitro bone marrow-derived macrophages, we show that IL-27 plays an antiviral role by regulating macrophage-mediated HSV-1 killing, IFN-β production, and IFN-stimulated gene expression after HSV-1 infection. Furthermore, we report that IL-27 is critical for macrophage survival, Ag uptake, and the expression of costimulatory molecules involved in the optimum induction of effector T cell responses. Our results indicate that IL-27 promotes endogenous antiviral and anti-inflammatory responses and represents a promising target for suppressing HSK progression.

CLEAR Strategy Inhibited HSV Proliferation Using Viral Vectors Delivered CRISPR-Cas9.

Herpes simplex virus type 1 (HSV-1) is a leading cause of encephalitis and infectious blindness. The commonly used clinical therapeutic drugs are nucleoside analogues such as acyclovir. However, current drugs for HSV cannot eliminate the latent virus or viral reactivation. Therefore, the development of new treatment strategies against latent HSV has become an urgent need. To comprehensively suppress the proliferation of HSV, we designed the CLEAR strategy (coordinated lifecycle elimination against viral replication). VP16, ICP27, ICP4, and gD-which are crucial genes that perform significant functions in different stages of the HSV infection lifecycle-were selected as targeting sites based on CRISPR-Cas9 editing system. In vitro and in vivo investigations revealed that genome editing by VP16, ICP27, ICP4 or gD single gene targeting could effectively inhibit HSV replication. Moreover, the combined administration method (termed "Cocktail") showed superior effects compared to single gene editing, which resulted in the greatest decrease in viral proliferation. Lentivirus-delivered CRISPR-Cas9/gRNA editing could effectively block HSV replication. The CLEAR strategy may provide new insights into the potential treatment of refractory HSV-1-associated diseases, particularly when conventional approaches have encountered resistance.

HSV-1 LAT Promoter Deletion Viruses Exhibit Strain-Specific and LAT-Dependent Epigenetic Regulation of Latent Viral Genomes in Human Neurons.

Herpes simplex virus 1 (HSV-1) establishes latency in neurons and expresses long noncoding RNAs termed the latency-associated transcripts (LATs). Two previous studies using HSV-1 recombinants containing deletions in the LAT promoter revealed opposing effects of the promoter deletion regarding the heterochromatinization of latent viral genomes in mice ganglia. Confounding variables in these studies include viral strains utilized (17syn+ versus KOS), anatomical infection site (footpad versus eye) and infectious virus dose (500 versus 1 × 105 PFU), and to date the basis for the differences between the two studies remains unresolved. We recently reported that 17syn+ and KOS display distinct differences in heterochromatin levels during latency in human neurons. This raised the possibility that the discrepancy seen between the two previous studies could be explained by strain-specific differences within the LAT region. Here, we examine two recombinants containing orthologous 202 bp LAT promoter deletions, 17ΔPst and KOSΔPst, in a human neuronal model of latency and reactivation (LUHMES). We found that LUHMES neurons recapitulate previous observations in mice where deletion of the LAT promoter results in an increase in H3K27me3 deposition on the viral genome compared to the parental strain 17syn+ but a decrease compared to the parental strain KOS. We also found distinct strain-specific differences in the production of viral transcripts and proteins during latency. These results indicate that the function and/or regulation of the LATs differs between HSV-1 strains and may shed light on some discrepancies found in the literature when examining the function of the LATs. IMPORTANCE Herpes simplex virus 1 (HSV-1) establishes a lifelong infection in neuronal cells. Periodically, the virus reactivates from this latent state and causes recurrent disease. Mechanisms that control entry into and maintenance of latency are not well understood, though epigenetic posttranslational modification of histones associated with the viral genome are known to play an important role. During latency, the latency-associated transcript (LAT) is known to impact epigenetic marks, but the ultimate effect has been a point of controversy. Here, we utilize a human neuronal cell line model of HSV latency and reactivation (LUHMES) to characterize latency for two HSV-1 wild-type strains and their respective LAT promoter deletion viruses. We find that the LAT acts in a strain-specific manner to influence levels of chromatin marks, viral transcription, and viral protein production. This work highlights the need to account for strain-specific differences when characterizing the LAT's function and the dynamics of reactivation.

Stacking the odds: Multiple sites for HSV-1 latency.

A hallmark of herpes simplex virus (HSV) infection is the establishment of latent virus in peripheral sensory ganglia of the latently infected host. We and others originally reported that the latency-associated transcript (LAT) is the only abundantly expressed viral gene in neurons within trigeminal ganglia (TG) of a latently infected host. Here, we investigated the possible contribution of various cells [i.e., B cells, dendritic cells (DCs), fibroblasts, glial cells, innate lymphoid cells (ILCs), macrophages, microglia, monocytes, natural killer cells, neurons, neutrophils, and T cells] isolated from TG of latently infected mice. Our results demonstrated that all of these cell types contain LAT, with DCs, neurons, and ILCs having the most LAT+ cells. These results suggest that HSV-1 can establish a quiescent/latent infection in a subset of nonneuronal cells, which enhances the chances that the virus will survive in its host.

Deletion of the Transcriptional Coactivator HCF-1 In Vivo Impairs the Removal of Repressive Heterochromatin from Latent HSV Genomes and Suppresses the Initiation of Viral Reactivation.

Transcription of herpes simplex virus 1 (HSV-1) immediate early (IE) genes is controlled at multiple levels by the cellular transcriptional coactivator, HCF-1. HCF-1 is complexed with epigenetic factors that prevent silencing of the viral genome upon infection, transcription factors that drive initiation of IE gene expression, and transcription elongation factors required to circumvent RNAPII pausing at IE genes and promote productive IE mRNA synthesis. Significantly, the coactivator is also implicated in the control of viral reactivation from latency in sensory neurons based on studies that demonstrate that HCF-1-associated epigenetic and transcriptional elongation complexes are critical to initiate IE expression and viral reactivation. Here, an HCF-1 conditional knockout mouse model (HCF-1cKO) was derived to probe the role and significance of HCF-1 in the regulation of HSV-1 latency/reactivation in vivo. Upon deletion of HCF-1 in sensory neurons, there is a striking reduction in the number of latently infected neurons that initiate viral reactivation. Importantly, this correlated with a defect in the removal of repressive chromatin associated with latent viral genomes. These data demonstrate that HCF-1 is a critical regulatory factor that governs the initiation of HSV reactivation, in part, by promoting the transition of latent viral genomes from a repressed heterochromatic state. IMPORTANCE Herpes simplex virus is responsible for a substantial worldwide disease burden. An initial infection leads to the establishment of a lifelong persistent infection in sensory neurons. Periodic reactivation can result in recurrent oral and genital lesions to more significant ocular disease. Despite the significance of this pathogen, many of the regulatory factors and molecular mechanisms that govern the viral latency-reactivation cycles have yet to be elucidated. Initiation of both lytic infection and reactivation are dependent on the expression of the viral immediate early genes. In vivo deletion of a central component of the IE regulatory paradigm, the cellular transcriptional coactivator HCF-1, reduces the epigenetic transition of latent viral genomes, thus suppressing HSV reactivation. These observations define HCF-1 as a critical regulator that controls the initiation of HSV reactivation from latency in vivo and contribute to understanding of the molecular mechanisms that govern viral reactivation.

APOE-ε4 genes may accelerate the activation of the latent form of HSV-1 which would lead to a faster progression of AD

This study investigates the impact of APOE alleles and latent Herpes Simplex Type 1 virus (HSV-1) activation on Alzheimer’s disease (AD) progression using the 5xFAD mouse model. APOE ε4 is recognized as a substantial genetic risk factor for sporadic AD, while HSV-1 has been linked to AD pathogenesis through inflammation and plaque formation. The experimental approach involves the introduction of human neurons carrying latent HSV-1 into 5xFAD mice harboring various APOE alleles (APOE2, APOE3, APOE4), along with stress induction and pharmacological interventions. The study aims to elucidate the combined impact of these variables on AD progression and the formation of Aβ plaques. Our anticipated results suggest that APOE ε4 may accelerate AD development, especially in conjunction with HSV-1 activation, while APOE ε2 may exert a mitigating influence. These findings have the potential to advance our understanding of the intricate mechanisms underpinning AD and provide insights into potential therapeutic approaches. Further exploration of these interactions could offer critical insights into the pursuit of effective AD treatments.

Establishing a Herpesvirus Quiescent Infection in Differentiated Human Dorsal Root Ganglion Neuronal Cell Line Mediated by Micro-RNA Overexpression.

HSV-1 is a neurotropic pathogen associated with severe encephalitis, excruciating orofacial sensation, and other chronic neuropathic complications. After the acute infection, the virus may establish a lifelong latency in the neurons of trigeminal ganglia (TG) and other sensory and autonomic ganglia, including the dorsal root ganglia (DRG), etc. The reactivation occurred periodically by a variety of physical or emotional stressors. We have been developing a human DRG neuronal cell-culture model HD10.6, which mimics the mature neurons for latency and reactivation with robust neuronal physiology. We found that miR124 overexpression without acyclovir (ACV) could maintain the virus in a quiescent infection, with the accumulation of latency-associate transcript (LAT). The immediate-early (IE) gene ICP0, on the other hand, was very low and the latent viruses could be reactivated by trichostatin A (TSA) treatment. Together, these observations suggested a putative role of microRNA in promoting HSV-1 latency in human neurons.

Ex Vivo Herpes Simplex Virus Reactivation Involves a Dual Leucine Zipper Kinase-Dependent Wave of Lytic Gene Expression That Is Independent of Histone Demethylase Activity and Viral Genome Synthesis.

Herpes simplex virus 1 (HSV-1) maintains a lifelong latent infection in neurons and periodically reactivates, resulting in the production of infectious virus. The exact cellular pathways that induce reactivation are not understood. In primary neuronal models of HSV latency, the cellular protein dual leucine zipper kinase (DLK) has been found to initiate a wave of viral gene expression known as phase I. Phase I occurs independently of both viral DNA replication and the activities of histone demethylase enzymes required to remove repressive heterochromatin modifications associated with the viral genome. In this study, we investigated whether phase I-like gene expression occurs in ganglia reactivated from infected mice. Using the combined trigger of explant-induced axotomy and inhibition of phosphatidylinositide 3-kinase (PI3K) signaling, we found that HSV lytic gene expression was induced rapidly from both sensory and sympathetic neurons. Ex vivo reactivation involved a wave of viral late gene expression that occurred independently of viral genome synthesis and histone demethylase activity and preceded the detection of infectious virus. Importantly, we found that DLK was required for the initial induction of lytic gene expression. These data confirm the essential role of DLK in inducing HSV-1 gene expression from the heterochromatin-associated genome and further demonstrate that HSV-1 gene expression during reactivation occurs via mechanisms that are distinct from lytic replication. IMPORTANCE Reactivation of herpes simplex virus from a latent infection is associated with clinical disease. To develop new therapeutics that prevent reactivation, it is important to understand how viral gene expression initiates following a reactivation stimulus. Dual leucine zipper kinase (DLK) is a cellular protein that has previously been found to be required for HSV reactivation from sympathetic neurons in vitro. Here, we show that DLK is essential for reactivation from sensory ganglia isolated from infected mice. Furthermore, we show that DLK-dependent gene expression ex vivo occurs via mechanisms that are distinct from production replication, namely, lytic gene expression that is independent of viral DNA replication and histone demethylase activity. The identification of a DLK-dependent wave of lytic gene expression from sensory ganglia will ultimately permit the development of novel therapeutics that target lytic gene expression and prevent the earliest stage of reactivation.