Adeno-associated Virus Vectors Efficiently Transduce Mouse and Rabbit Sensory Neurons Coinfected with Herpes Simplex Virus 1 following Peripheral Inoculation

Abstract

Following infection of epithelial tissues, herpes simplex virus 1 (HSV-1) virions travel via axonal transport to sensory ganglia and establish a lifelong latent infection within neurons. Recent studies have revealed that, following intraganglionic or intrathecal injection, recombinant adeno-associated virus (rAAV) vectors can also infect sensory neurons and are capable of stable, longterm transgene expression. We sought to determine if application of rAAV to peripheral nerve termini at the epithelial surface would allow rAAV to traffic to sensory ganglia in a manner similar to that seen with HSV. We hypothesized that footpad or ocular inoculation with rAAV8 would result in transduction of dorsal root ganglia (DRG) or trigeminal ganglia (TG), respectively. To test this, we inoculated the footpads of mice with various amounts of rAAV as well as rAAV capsid mutants. We demonstrated that this method of inoculation can achieve a transduction rate of >90% of the sensory neurons in the DRG that innervate the footpad. Similarly, we showed that corneal inoculation with rAAV vectors in the rabbit efficiently transduced >70% of the TG neurons in the optic tract. Finally, we demonstrated that coinfection of mouse footpads or rabbit eyes with rAAV vectors and HSV-1 resulted in colocalization in nearly all of the HSV-1-positive neurons. These results suggest that rAAV is a useful tool for the study of HSV-1 infection and may provide a means to deliver therapeutic cargos for the treatment of HSV infections or of dysfunctions of sensory ganglia. IMPORTANCEAdeno-associated virus (AAV) has been shown to transduce dorsal root ganglion sensory neurons following direct intraganglionic sciatic nerve injection and intraperitoneal and intravenous injection as well as intrathecal injection. We sought to determine if rAAV vectors would be delivered to the same sensory neurons that herpes simplex virus (HSV-1) infects when applied peripherally at an epithelial surface that had been treated to expose the underlying sensory nerve termini. For this study, we chose two well-established HSV-1 infection models: mouse footpad infection and rabbit ocular infection. The results presented here provide the first description of AAV vectors transducing neurons following delivery at the skin/epithelium/eye. The ability of AAV to cotransduce HSV-1-infected neurons in both the mouse and the rabbit provides an opportunity to experimentally explore and disrupt host and viral proteins that are integral to the establishment of HSV-1 latency, to the maintenance of latency, and to reactivation from latency in vivo. Herpes simplex virus 1 (HSV-1) establishes a lifelong latent infection within the neurons of sensory ganglia. These ganglia are highly specialized structures composed of a diverse assemblage of neuronal and nonneuronal cells. Ganglionic neurons detect a wide variety of sensory inputs, including temperature, touch, and pain, and relay this information into the central nervous system (CNS). Immunohistochemical analyses of infected trigeminal g...

In Vivo Knockdown of the Herpes Simplex Virus 1 Latency-Associated Transcript Reduces Reactivation from Latency

“…(AAV-LATRz-235). We have previously demonstrated that AAV applied to the rabbit eye is efficiently transported to the TG and can efficiently transduce Ͼ70% of the total neurons (22). Seventeen days after treatment with AAV, the rabbit eyes were iontophoresed with epinephrine and swabbed daily for 12 days to collect tears.…”

“…Adeno-associated virus (AAV) vectors were used to deliver a ribozyme to target the HSV-1 LAT being expressed in latently infected sensory neurons. This approach exploits the ability of some serotypes of AAV to be inoculated on the skin or in the cornea, subsequently taken up by nerve termini, and ultimately transported by axonal transport to the neuronal cell bodies, where the HSV-1 latent infection resides (22). We have previously used this approach to deliver small interfering RNAs (siRNAs) to latently infected neurons in the rabbit trigeminal ganglia (TG) to knockdown the cellular insulator protein CTCF (23).…”

Harnessing CRISPR/Cas 9 System for manipulation of DNA virus genome

“…Several viral and nonviral systems have been employed for CRIPR Cas9 delivery. Viral vectors such as lentiviral, adenoviral, and adeno associated virus (AAV) are being widely used as delivery systems with promising results . However, the application of lenti and adeno viral vectors is limited due to low viral titers, risk of internal mutation, long duration of CRISPR/Cas9 activity, and stimulation of the immune system, respectively .…”

“…Viral vectors such as lentiviral, adenoviral, and adeno associated virus (AAV) are being widely used as delivery systems with promising results. [138][139][140][141] However, the application of lenti and adeno viral vectors is limited due to low viral titers, risk of internal mutation, long duration of CRISPR/Cas9 activity, and stimulation of the immune system, respectively. 138,[142][143][144] Given the promising characteristics of AAV such as versatility, less toxicity, minimal immunogenicity and high tropism for neurons, hepatocytes, and epithelial cells, it can be regarded as an attractive delivery system for introduction of anti-HSV, anti-HBV, and anti-HPV to the infected cells.…”

CRISPR/Cas9, a powerful tool to target human herpesviruses

“…As many of these are neurotropic (Salganik et al, ), they are ideal candidates to infect latent HSV and VZV‐infected neurons. Indeed, several studies show efficient delivery of AAV to both the trigeminal and dorsal ganglia by locally infecting the cornea or the footpad of mice, rats, and rabbits (Mason et al, ; Watson et al, ), suggesting that safe and efficient delivery of AAV vectors carrying anti‐herpesviral CRISPR/Cas9 may be possible to cripple or even clear neurotropic alphaherpesviruses from cells. Several AAV serotypes can also infect immune cell types, albeit to low efficiency (Pandya, Ortiz, Ling, Rivers, & Aslanidi, ; Veldwijk et al, ), opening up a way for targeting of latent HCMV‐infected or EBV‐infected cells.…”