An Immortalized Human Dorsal Root Ganglion Cell Line Provides a Novel Context To Study Herpes Simplex Virus 1 Latency and Reactivation

Abstract

A defining characteristic of alphaherpesviruses is the establishment of lifelong latency in host sensory ganglia with occasional reactivation causing recurrent lytic infections. As an alternative to rodent models, we explored the use of an immortalized cell line derived from human dorsal root ganglia. HD10.6 cells proliferate by virtue of a transduced tetracycline-regulated v-myc oncogene. In the presence of doxycycline, HD10.6 cells mature to exhibit neuronal morphology and express sensory neuron-associated markers such as neurotrophin receptors TrkA, TrkB, TrkC, and RET and the sensory neurofilament peripherin. Infection of mature HD10.6 neurons by herpes simplex virus 1 (HSV-1) results in a delayed but productive infection. However, infection at a low multiplicity of infection (MOI) in the presence of acyclovir results in a quiescent infection resembling latency in which viral genomes are retained in a low number of neurons, viral gene expression is minimal, and infectious virus is not released. At least some of the quiescent viral genomes retain the capacity to reactivate, resulting in viral DNA replication and release of infectious virus. Reactivation can be induced by depletion of nerve growth factor; other commonly used reactivation stimuli have no significant effect.IMPORTANCE Infections by herpes simplex viruses (HSV) cause painful cold sores or genital lesions in many people; less often, they affect the eye or even the brain. After the initial infection, the virus remains inactive or latent in nerve cells that sense the region where that infection occurred. To learn how virus maintains and reactivates from latency, studies are done in neurons taken from rodents or in whole animals to preserve the full context of infection. However, some cellular mechanisms involved in HSV infection in rodents are different from those in humans. We describe the use of a human cell line that has the properties of a sensory neuron. HSV infection in these cultured cells shows the properties expected for a latent infection, including reactivation to produce newly infectious virus. Thus, we now have a cell culture model for latency that is derived from the normal host for this virus.KEYWORDS HSV-1, quiescent infection, sensory neurons H erpesviruses are unusual among viruses in that they employ two infection strategies, lytic (replicative or productive) infection and latency (quiescent) infection. Alphaherpesviruses typically replicate in epithelial cells and establish latency in the peripheral nervous system within the sensory neurons of the ganglia that innervate mucosa of the primary infection site (1), such as the trigeminal ganglia (TG) and dorsal root ganglia (DRG). Operationally, latency is defined as "the persistence of a viral genome within tissue where, at any given time, there is a population of cells that lack detectable infectious virus, viral proteins, or viral lytic transcripts that are dormant but have the capability of being reactivated" (2). During latency of herpes simplex viruses

Modeling HSV-1 Latency in Human Embryonic Stem Cell-Derived Neurons

“…Whilst this manuscript was in preparation, Thellman and colleagues described an alternative HSV-1 latency model based on neurons derived from an immortalized human DRG cell line designated HD10.6 [79]. Proliferation of the cell line is driven by the expression of a v-myc oncogene.…”

Herpes Simplex Virus 1 Infection Promotes the Growth of a Subpopulation of Tumor Cells in Three-Dimensional Uveal Melanoma Cultures

“…Following the viral inoculation or mock infection, the cultures were incubated and maintained with culture medium, with refreshing of the medium every 2 days. Some viruses and mock-infected 3D cultures were exposed to and maintained in 100 μM acyclovir (an antiviral agent that inhibits HSV-1 DNA replication) following the virus or mock-inoculation procedures ( 38 , 39 ). All cultures were observed daily using an inverted fluorescence microscope for cell growth and GFP expression.…”

Herpes Simplex Virus Establishment, Maintenance, and Reactivation: In Vitro Modeling of Latency

“…For example, the HD10.6 cell line was derived from human DRG and proliferates via a tetracycline-regulated v- myc oncogene [ 105 ]. In the presence of doxycycline, cellular proliferation is suppressed and cells mature to exhibit a sensory neuron-associated phenotype (SNAP cells) [ 105 , 106 ]. These cells were immortalized using the same technique as the Lund human mesencephalic (LUHMES) cell line, which was isolated from human mesencephalon and is widely used in neurodegenerative disease studies [ 107 , 108 ].…”