VZV latency

As previously described, VZV DNA is able to persist in the host in a dormant state called latency. The cells where the virus stablishes latency are the peripheral neurons, either sensory neurons from the DRG and TG or autonomic neurons.

Latency is a main concern as the immune system cannot detect latently infected cells and therefore the virus cannot be eliminated from the host completely. VZV can remain latent for years, and reactivate later in life causing secondary infection such as herpes zoster and others mentioned above (see section “1.1.2. Varicella zoster virus infection” and Fig. 1.4).

1.1.6.1. Establishment of latency

The factors and mechanisms involved in latency establishment and maintenance are still not fully understood. In contrast to HSV, the lack of suitable animal and in vitro models and the problems to achieve good viral titers in VZV have complicated the research on this virus. Since VZV is highly restricted in non-human cells, the use of human neurons is required to study VZV latency.

During latency, the VZV genome is found as an episome in the nucleus of infected neurons. At this infection stage viral gene expression is dramatically restricted and

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only IE63 has been found to be expressed at early times post-mortem in human TG¹¹⁴. As previously described, IE63 is a tegument protein that has been found to play an important role in inhibiting transcription, IFN-α cell response and apoptosis in vitro^58,60,61. However, it has been also reported that post-mortem time affects viral expression, very probably because of cellular stress. Because of this reason, is difficult to extrapolate results obtained in post-mortem tissues with the in vivo situation¹¹⁴. Some studies also reported the immune detection of IE63 among other proteins in post-mortem human ganglia^115,116. However, two indepdendent studies performed by Zerboni L. and Ouwendijk WJ. in 2012 showed that mouse and rabbit antibodies react to blood type A antigens in neurons causing a false positive result^12,59.

Interestingly, a VZV transcript commonly expressed in latent infected TG neurons has been recently discovered. This transcript, partially antisense to ORF61, has been named VZV latency transcript (VLT). The protein expressed by this transcript can be detected in the nucleus and cytosol with late kinetics and its functions is unknown.

However, the VLT transcript was able to inhibit the expression of IE61, a transactivator for VZV lytic genes, which may suggest a possible role in VZV latency maintanance¹¹⁷.

Despite their limitations some studies performed with rodents have provided valuable information on VZV neuronal infection and latency in vivo. Using the SCID mouse model xenotransplanted with human DRG, the group of Ann M. Arvin showed VZV productive infection during 3-4 weeks⁵³. Then, infection progresses to a non-productive stage where VZV genomes show low transcription of IE63. This suggests that VZV may be able to establish latency in the absence of an adaptive immune response. However, reactivation from this model has not been documented yet.

27 Another research showed that VZV is also able to stablish latency in the enteric nervous system (ENS). Gastrointestinal surgically removed samples from autopsy revealed that VZV DNA and transcripts were present in the ENS¹¹⁸. The use of the guinea pig model showed that the virus can reach the ENS by axonal transport or by direct transport in lymphocytes during viremia¹¹⁸.

There is also a rigorous research in developing realistic and reliable in vitro models as well. These models imply the culture of neurons obtained from fetal sensory ganglia or neurons differentiated from pluripotent stem cells^14,119,120. A recent latency and reactivation model has been published by Markus et al., 2015 using neurons derived from human embryonic stem cells. According to the researchers, latency was induced by infecting the cells in the presence of acyclovir and using very low viral titer of cell-free virus, or by infecting the axons of the neurons. During this stage, transcripts from all VZV genomic regions including IE63 were detected but at much lower degree when compared with the productive infection¹²¹. No infectious virus was obtained unless reactivation was induced. VZV latency from axonal infection using neurons derived from human embryonic stem cells was also confirmed by Tomohiko Sadaoka in 2016¹²².

1.1.6.2. Reactivation

Reactivation is the process by which a latent viral genome is induced to produce infectious viral particles. The factors involved in this process are still poorly understood although it is known that immune suppression, in particular T-cell depletion, plays an important role¹²³. VZV reactivation occurs more often in individuals of advanced age with the consequent decline in immune cells¹²⁴, but also in individuals undergoing immunosuppressive therapy or in HIV positive patients¹²⁵.

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Although very little is known regarding the stimuli that lead to reactivation, the role of growth factors and phosphatidil-inositol kinase 3 (PI3K) has been confirmed.

Therefore, removal of growth factors or inhibition of the Pl3K signaling pathway, a pathway triggered when the receptor TrkA interacts with nerve growth factor (NGF), results in VZV reactivation¹²¹. These pathways are also important for HSV latency and are linked to stress pathways^126,127. Interestingly, using human neurons, Tomohiko Sadaoka showed that the vaccine strain (VOKA) was able to stablish latency, but was less efficient in reactivation¹²². This may be one of the reasons why the vaccine strain is less virulent than its parental strain in vivo.