Research Article: Three-Dimensional Normal Human Neural Progenitor Tissue-Like Assemblies: A Model of Persistent Varicella-Zoster Virus Infection

Date Published: August 1, 2013

Publisher: Public Library of Science

Author(s): Thomas J. Goodwin, Maureen McCarthy, Nikolaus Osterrieder, Randall J. Cohrs, Benedikt B. Kaufer, Roger D. Everett.


Varicella-zoster virus (VZV) is a neurotropic human alphaherpesvirus that causes varicella upon primary infection, establishes latency in multiple ganglionic neurons, and can reactivate to cause zoster. Live attenuated VZV vaccines are available; however, they can also establish latent infections and reactivate. Studies of VZV latency have been limited to the analyses of human ganglia removed at autopsy, as the virus is strictly a human pathogen. Recently, terminally differentiated human neurons have received much attention as a means to study the interaction between VZV and human neurons; however, the short life-span of these cells in culture has limited their application. Herein, we describe the construction of a model of normal human neural progenitor cells (NHNP) in tissue-like assemblies (TLAs), which can be successfully maintained for at least 180 days in three-dimensional (3D) culture, and exhibit an expression profile similar to that of human trigeminal ganglia. Infection of NHNP TLAs with cell-free VZV resulted in a persistent infection that was maintained for three months, during which the virus genome remained stable. Immediate-early, early and late VZV genes were transcribed, and low-levels of infectious VZV were recurrently detected in the culture supernatant. Our data suggest that NHNP TLAs are an effective system to investigate long-term interactions of VZV with complex assemblies of human neuronal cells.

Partial Text

Varicella-zoster virus (VZV) is a ubiquitous human herpesvirus, as evidenced by a seroprevalence of more than 95% worldwide [1]. Among human herpesviruses, VZV has the smallest genome of approximately 125 kbp, which contains at least 70 open reading frames (ORFs) and consists of two unique regions, unique long (UL) and unique short (US), each flanked by inverted repeat regions (TRL, IRL, TRS, IRS) [2], [3]. Primary VZV infection typically causes childhood varicella (chickenpox). During primary infection the virus gains access to and establishes latency in multiple cranial, dorsal root and autonomic ganglia. Varicella vaccination programs have successfully reduced the incidence of clinical disease in children in the USA by about 80% [4], [5]; however, the attenuated vaccine, like wild-type VZV, is still able to establish latency in the peripheral nervous system [6]. Both wild-type and vaccine virus can reactivate from latency, particularly in elderly and immunocompromised individuals. Reactivation is associated with a declining VZV specific T-cell immunity and can result in herpes zoster (shingles) especially in the elderly, which is characterized by severe pain and often followed by postherpetic neuralgia [7], [8]. Reactivation can also lead to progressive outer retinal necrosis and stroke by ischemic vasculopathy [9], [10]. With respect to disease severity, duration and quality-of-life impairment, VZV reactivation in adulthood can be more serious than primary childhood infection.

Establishment of latent infection and subsequent reactivation is integral to the alphaherpesvirus life cycle and ensures that virus is maintained in the population by intermittent virus production and transmission [22]. However, VZV appears to be unique among alphaherpesviruses with respect to establishment of and reactivation from the persistent/latent state [23]–[26], emphasizing the need for accurate nomenclature. VZV-host interactions are classified as acute, “rapid production of infectious virions followed by rapid resolution and elimination of infection”, or persistent, “virus particles or products continue to be produced for long periods in which virions are continuously or intermittently produced” [27]. In most human cells in culture, VZV infection is acute and cells succumb to virus infection within 3–5 days most likely through apoptosis [28]–[31]. The low ratio (1∶40,000) of infectious to defective VZV particles indicates that production of complete virions is extremely inefficient [32]. Human neurons are known to be latently infected by VZV, but the lack of a suitable animal model has hindered investigations into this exceptional relationship between VZV and a host cell or organ. Our common understanding is that VZV infection of neurons results in latency. Latency is an extreme variant of persistent infection where, as exemplified by herpes simplex virus type 1 (HSV-1), “infectious virions can no longer be isolated” [27]. During latency, most HSV-1 genes are silenced through epigenetic modification of resident histones or by virus-specific miRNAs [33]–[35]. Therefore, HSV-1 gene transcription in infected neurons is restricted to the latency-associated transcript (LAT) at the cellular level, while no viral proteins and infectious virions are produced [36]–[38].




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