Axonal transport mediates West Nile virus entry into the central nervous system and induces acute flaccid paralysis

Abstract

West Nile virus (WNV) has emerged as a significant cause of epidemic viral encephalitis and flaccid limb paralysis, yet the mechanism by which it enters the CNS remains uncertain. We used compartmentalized neuron cultures to demonstrate that WNV spreads in both retrograde and anterograde directions via axonal transport. Transneuronal spread of WNV required axonal release of viral particles and was blocked by addition of a therapeutic neutralizing antibody. To test the physiologic significance of axonal transport in vivo, we directly inoculated the sciatic nerve of hamsters with WNV. Intrasciatic infection resulted in paralysis of the hind limb ipsilateral but not contralateral to the injection site. Limb paralysis was blocked either by surgical transection of the sciatic nerve or treatment with the therapeutic neutralizing antibody. Collectively, these studies establish that WNV undergoes bidirectional spread in neurons and that axonal transport promotes viral entry into the CNS and acute limb paralysis. Moreover, antibody therapeutics directly inhibit transneuronal spread of WNV infection and prevent the development of paralysis in vivo.flavivirus ͉ neuron ͉ retrograde ͉ transneuronal spread W est Nile virus (WNV) is a neurotropic member of the Flaviviridae family of RNA viruses and is related to other important arthropod-borne human pathogens. WNV is maintained in an enzootic cycle between mosquitoes and birds and has become an important global cause of epidemic encephalitis. Since its emergence in the United States in 1999, Ϸ26,000 cases of symptomatic WNV infection have been confirmed (www.cdc. gov/ncidod/dvbid/westnile/surv&control.htm#maps), and seroprevalence studies suggest that several million people have been infected (1).Rodent models have provided insight into the mechanisms of WNV spread to the CNS. After s.c. inoculation, WNV-infected dendritic cells traffic to the draining lymph node, resulting in a primary viremia and infection of peripheral tissues. Within 6 days, WNV is cleared from the serum and peripheral organs and enters the CNS and induces neurological disease (reviewed in ref.2). Nonetheless, the specific mechanisms by which WNV or other neurotropic flaviviruses enter into the CNS are largely unknown. CNS infection may occur in part via hematogenous spread, as increased viremia in immunodeficient mice (2) and TNF-␣-mediated changes in blood-brain-barrier permeability correlate with earlier CNS entry (3).Axonal transport from infected peripheral neurons mediates CNS entry and pathogenesis of viruses in the Herpesviridae, Rhabdoviridae, and Picornaviridae families (4-6). Viral spread in neurons is generally mediated by fast axonal transport, a microtubule-associated, anterograde and retrograde transport system. In classical studies, CNS infection of rabies virus or poliovirus was prevented by axonal ligation or degeneration (7,8). Insights into the biology of axonal spread have been facilitated by the development of compartmentalized, or Campenot, chambers for culturing neurons (9). These s...

Differential Replication of Pathogenic and Nonpathogenic Strains of West Nile Virus within Astrocytes

“…Indeed, the nonneuropathogenic phenotype of some strains of WNV has been attributed to an inability to invade the CNS (12,13,27). Increasing evidence suggests that WNV entry into the CNS is a multistep process that can occur through one of several routes (17,20,(31)(32)(33)(34)(35). WNV entry into the CNS has been shown to precede disruption of the BBB and leukocyte infiltration (32,33,36,37), suggesting that WNV utilizes a direct mechanism to initially invade the CNS, such as basolateral secretion of virus particles from infected brain endothelial cells or transcytosis.…”

“…All experiments with HBMVECs and HBCAs were performed on cells passaged no more than 14 times. Primary cortical neurons were prepared from day 15 C57BL/6 mouse embryos as previously described (17,18). Cortical neuron experiments were performed using neurons that were cultured for 3 to 4 days in neurobasal medium containing B27 and L-glutamine (Invitrogen).…”

Encephalitic Arboviruses: Emergence, Clinical Presentation, and Neuropathogenesis

“…Arboviral infections typically occur in highly enervated dermis, which may lead to viral entry into the CNS through peripheral neurons. Both in vitro and in vivo studies have shown that WNV can spread along neurons via retrograde axonal microtube-mediated transport [139,140]. Indeed, intrasciatic inoculation of hamsters with WNV resulted in infection of spinal cord and acute flaccid paralysis.…”

Infection par le virusWest Nilechez l’homme

“…Dans certains cas, le virus peut être inoculé directement dans la circulation sanguine court-circuitant certaines de ces étapes. Le mécanisme d'entrée du virus dans le système nerveux central n'est pas totalement compris mais plusieurs mécanismes ont été envisagés : une perméabilisation de la barrière hémato-encéphalique par les facteurs sécrétés circulants (TNF- [tumor necrosis factor], MMP9 [matrix metalloproteinase]) [10], le transport axonal rétrograde à partir des terminaisons nerveuses des neurones périphériques [11], la transcytose ou l'infection des cellules endothéliales des capillaires du cerveau [12], le transport du virus par les cellules périphériques infectées pénétrant dans le cerveau [10].…”

“…L'examen des tissus cérébraux montra des nodules microgliaux dispersés -notamment dans le tronc cérébral -composés de lymphocytes, histiocytes, avec une infiltration de cellules mononucléées périvasculaires, essentiellement des lymphocytes T CD8 + [13]. L'infection des neurones a été rapportée avec perte de la structure cellulaire et apoptose [11]. Les cellules mononucléées inflammatoires participeraient à l'élimination du virus dans le Les donneurs de sang, dont le sang est testé pour la présence du WNV, constituent d'excellents candidats pour la collecte d'échantillons pré-levés pendant la phase précoce de l'infection -définie par la détection de l'ARN viral en l'absence d'IgM spécifiques -avant le développement potentiel de symptômes [19].…”