THESIS
Complete Title of Thesis:
"Reovirus
Myelitis: development and analysis of
mechanisms of spinal cord injury in a novel model of viral myelitis."
Prepared under the direction of: Kenneth L Tyler, MD
SUMMARY
Virus infection of
the central nervous system (CNS) including myelitis and encephalitis are
important causes of morbidity and mortality throughout the world. Current
treatment options for these devastating illnesses are limited and often only
marginally effective. We developed an experimental model of viral myelitis utilizing reovirus
infection, a classic experimental system that has been extensively utilized to
study viral pathogenesis. Two neurotropic strains of reovirus, type 3 Abney and
Dearing, induce a progressive paralysis in 90% of infected mice. Investigation
of mechanisms of injury found apoptosis of reovirus-infected neurons by
co-localization of active caspase 3 and reovirus antigen in lumbar spinal cord
(SC) sections. Western blot analysis found activation of the protease Calpain; as
well as increases in inducible nitric oxide synthase (iNOS) and heme-oxygenase 1,
markers of oxidative stress. Laser scanning confocal microscopy of infected SC
illustrated dramatic activation and proliferation of microglia and astrocytes
(“gliosis”) in SCs of infected animals, indicating an innate immune response of
these uninfected cells to reovirus infection of neurons. Further evaluation of
the innate immune response identified up-regulation of interferon stimulated
genes including genes known to sensitize cells to apoptosis when
over-expressed. Up-regulation of inflammatory cytokines and chemokines were
found in microglia-enriched fractions of SC cellular suspensions. Even though
the neonatal mice used in our studies have immature immune systems, a T-cell
response was detected in paralyzed animals. Ex vivo SC slice cultures
established that CNS trafficking of hematopoietically derived inflammatory
cells was not required for reovirus induced CNS tissue destruction or
up-regulation of inflammatory cytokines and chemokines. The cardinal clinical,
pathological and immunological features of reovirus myelitis, closely parallel
those seen in important types of human viral myelitis, thereby establishing
this as a relevant experimental model system. New model systems are required to
study the basis of virus-induced spinal cord injury and to test potential
therapeutic interventions. Future studies utilizing knock-out mice with defined
immune deficits in both the SC ex vivo slice system and in in vivo
infection will enable us to evaluate the timing and role of specific
inflammatory cell types and their mediators in the
pathogenesis of viral myelitis.