Antigen‐specific cytotoxic T lymphocytes protect against lethal West Nile virus encephalitis

Abstract

Infection with West Nile virus (WNV) causes fatal encephalitis in immunocompromised animals. Previous studies in mice have established that T cell protection is required for clearance of WNV infection from tissues and preventing viral persistence. The current study assessed whether specific WNV peptide epitopes could elicit a cytotoxic T lymphocyte (CTL) response capable of protecting against virus infection. Hidden . Of the 35 peptides predicted to bind MHC class I molecules, one immunodominant CTL recognition peptide was identified in each of the envelope and non-structural protein 4B genes. Addition of these but not control peptides to CD8 + T cells from WNV-infected mice induced IFN-c production. CTL clones that were generated ex vivo lysed peptide-pulsed or WNV-infected target cells in an antigen-specific manner. Finally, adoptive transfer of a mixture of envelope-and nonstructural protein 4B-specific CTL to recipient mice protected against lethal WNV challenge. Based on this, we conclude that CTL responses against immundominant WNV epitopes confer protective immunity and thus should be targets for inclusion in new vaccines.

The relative contribution of antibody and CD8+ T cells to vaccine immunity against West Nile encephalitis virus

“…Intracellular IFN-γ staining was performed on splenocytes at indicated days after infection or vaccination as previously described [37,38]. Briefly, splenocytes from infected or vaccinated mice were stimulated with 1μg/ml of the WNV NS4B peptide (#33, D b -restricted, SSVWNATTAI) or WNV E peptide (#3, K b -restricted, IALTFLAV) and WNV E peptide (#28, D b -restricted, TVWRNRETL) [38,39] for four hours at 37ºC with the addition of Golgi plug ™ (1μl/ml, BD Biosciences), washed and stained with fluorescein isothiocyanate (FITC)-conjugated mAbs to CD8 or a FITC-conjugated isotype control mAb (BD Biosciences) at 4ºC for 30 minutes.…”

“…Briefly, splenocytes from infected or vaccinated mice were stimulated with 1μg/ml of the WNV NS4B peptide (#33, D b -restricted, SSVWNATTAI) or WNV E peptide (#3, K b -restricted, IALTFLAV) and WNV E peptide (#28, D b -restricted, TVWRNRETL) [38,39] for four hours at 37ºC with the addition of Golgi plug ™ (1μl/ml, BD Biosciences), washed and stained with fluorescein isothiocyanate (FITC)-conjugated mAbs to CD8 or a FITC-conjugated isotype control mAb (BD Biosciences) at 4ºC for 30 minutes. After washing and fixing with 1% PFA in PBS, cells were permeabilized with saponin and stained with an allophycocyanin (APC)-conjugated IFN-γ antibody or an isotype control (BD Biosciences) at 4ºC for 30 minutes, washed and analyzed by flow cytometry.…”

“…CD8 + T cells that are generated during WNV infection protect mice against challenge [38,39,43,44]. Prior immunization studies with live attenuated, DNA plasmid, or protein subunit vaccines against WNV have demonstrated the induction of memory T cell responses [21,25,32], although their role in protection against challenge was not evaluated.…”

“…Prior immunization studies with live attenuated, DNA plasmid, or protein subunit vaccines against WNV have demonstrated the induction of memory T cell responses [21,25,32], although their role in protection against challenge was not evaluated. To begin to address this question, we first assessed the efficiency of induction of WNV-specific CD8 + T cells after immunzation by analyzing intracellular IFN-γ production ex vivo after antigen-specific restimulation with D b and K b -restricted WNV NS4B and E peptides [37,38]. Splenocytes were harvested from mice on day 14 or 60 after primary immunization or infection or on day 70, 10 days after boosting and the level of CD8 + T cell activation was analyzed by flow cytometry (Fig 3A).…”

Infection par le virusWest Nilechez l’homme

“…La réponse des lymphocytes T est donc importante pour combattre l'infection [9]. Chez la souris, il a été montré que les lymphocytes T CD4 + et CD8 + participent à l'élimination du virus en périphérie et dans le système nerveux central [28,29]. Le transfert de lymphocytes T CD8 + chez des souris défi-cientes en lymphocytes permet de contrôler l'infection par le WNV [30].…”