Sindbis virus (SINV), the prototype alphavirus, causes encephalomyelitis in mice and provides a model for studying the pathogenesis of arboviral encephalomyelitis. Our previous studies with SINV have shown that survival is dependent on host and viral factors that include age and genetic background, virulence of the virus and the host immune response. Neuroadapted SINV (NSV) is virulent for adult C57BL/6 (B6) mice and is a model for virus-induced fatal encephalomyelitis in mature animals. NSV-infected mice develop weakness that progresses to paralysis and death within 7-10 days. Virus clearance from the brain and spinal cord begins 5-6 days after infection and is coincident with the onset of neurological disease. Although the antiviral immune response is essential for virus clearance, it can also contribute to fatal disease. We have shown that survival after NSV infection is improved in T cell-deficient mice and in mice with pharmacologic inhibition of the inflammatory response. Therefore, virus-specific immune cells entering the central nervous system (CNS) contribute to neuronal damage, but the mechanisms and immune factors that cause neuronal death during fatal encephalomyelitis are not known. In preliminary studies, we have determined that deficiency of the important regulatory cytokine interleukin (IL)-10 accelerates the onset of fatal NSV-induced paralytic disease. Accelerated disease is associated with an early increase in the CNS of CD4+ T cells producing IL-17 (Th17 cells) and a delay in virus clearance. Determination of the role of Th17 cells in NSV-induced immunopathology and identification of the mechanism(s) by which they influence outcome is important for developing interventions and for identifying host determinants of susceptibility to severe disease.
The specific aims of the proposal are: (1) Determine whether Th17 cells entering the CNS during fatal viral encephalomyelitis have or evolve a "pathogenic" phenotype. Th17 cells in IL-10-/- and wild type (WT) B6 mice will be characterized for expression of transcription factors, cytokines and surface receptors using multicolor flow cytometry. The CNS environment for Th17 cells will be assessed using qRT-PCR and immunoassays. The transcriptome of virus- induced CNS Th17 cells will be determined. (2) Determine whether Th17 cells are responsible for virus- induced immunopathologic disease. Th17 cells and outcome will be studied in mice deficient in factors required for Th1 and Th17 differentiation, genetically resistant BALB/c mice, mice infected with an avirulent strain of SINV and mice rescued from fatal encephalitis by antiviral antibody. (3) Determine the mechanism of CD4+ T cell-mediated neuronal damage and test candidate therapeutics. Effectors will be identified using genetically deficient mice, neutralizing antibodies, immunohistochemistry and CNS slice cultures followed by identification of therapies that prevent immunopathologic CNS damage. (4) Determine the mechanism for delayed virus clearance. Production of IFN-? and E2 antibody in the CNS and the effect of IL-17 and other Th17 effector molecules on the antiviral effects of IFN-? and anti-E2 antibody will be measured.

Public Health Relevance

Infection of the brain and spinal cord by mosquito-transmitted viruses can lead to death or long-term disability in those that survive. We have shown that the immune response is responsible for much of the neurologic damage during infection. This research will determine the mechanisms by which the immune system damages virus-infected neurons and will help to identify interventions that prevent this damage.

Agency
National Institute of Health (NIH)
Type
Research Project (R01)
Project #
1R01NS087539-01
Application #
8690404
Study Section
Clinical Neuroimmunology and Brain Tumors Study Section (CNBT)
Program Officer
Wong, May
Project Start
Project End
Budget Start
Budget End
Support Year
1
Fiscal Year
2014
Total Cost
Indirect Cost
Name
Johns Hopkins University
Department
Microbiology/Immun/Virology
Type
Schools of Public Health
DUNS #
City
Baltimore
State
MD
Country
United States
Zip Code
21218
Kulcsar, Kirsten A; Baxter, Victoria K; Greene, Ivorlyne P et al. (2014) Interleukin 10 modulation of pathogenic Th17 cells during fatal alphavirus encephalomyelitis. Proc Natl Acad Sci U S A 111:16053-8