The threat of avian influenza A (H5N1) infection in humans remains a global health concern. Current influenza vaccines stimulate antibody responses against the surface glycoproteins but are ineffective against strains that have undergone significant antigenic variation. An alternative approach is to stimulate pre-existing memory T cells established by seasonal human influenza A infection that could cross-react with H5N1 by targeting highly conserved internal proteins. To determine how common cross-reactive T cells are, we performed a comprehensive ex vivo analysis of cross-reactive CD4 + and CD8 +

Respiratory syncytial virus (RSV) is an ubiquitous paramyxovirus belonging to the genus Pneumovirus (1). Natural human RSV infections occur in winter outbreaks, usually resulting in common cold syndrome . Infants are especially prone to lower respiratory tract infection, which is the most common single cause of hospitalization of this age group in industrial countries. Vaccination with formaldehyde-inactivated RSV induces neutralizing serum antibodies, but fails to protect against natural infection; moreover, infected vaccinees suffer exacerbated lung disease. Similarly vaccinated cotton rats show enhanced pathological changes in the lungs (2), as do mice primed with recombinant vaccinia viruses expressing single RSV proteins (3). These methods of priming induce both humoral and cellular immune responses to RSV (2-5). Passive transfer of mAbs has not been shown to enhance pathology in RSV infected mice, and some antibodies protect against pulmonary disease (6, 7). Cellmediated immunity may therefore play a role in the pathogenesis of RSV-induced disease. Although polyclonal memory T cells can clear persistent RSV infection in immunodeficient mice (8), the role of T cell subpopulations in immunopathology merits examination.In this study, we use bronchoalveolar lavage (BAL) to monitor pulmonary disease in RSV-infected mice, and show enhanced pathology associated with accelerated clearance of lung virus after intravenous transfer of a cytotoxic RSV-specific T cell line and a CTL clone.

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We used well-gassed hanging drop (20 microliters) cultures with high concentrations of purified T cells from normal BALB/c mice to examine whether dendritic cells (DC) can induce primary antiviral proliferative T cell responses and generate virus-specific CTL. We found that DC exposed to infectious influenza virus in vitro or in vivo in small numbers (0.1-1%) resulted in strong proliferation of responder T cells within 3 d, and this was strongly inhibited by antibodies to class II MHC molecules. In addition, in 5-d cultures, the influenza-treated DC generated CTL specifically able to lyse influenza-infected syngeneic target cells bearing MHC class I antigens. The most potent nucleoprotein (NP) epitope recognized by BALB/c CTL is peptide 147-158 (Arg156-) and influenza-infected DC in vitro stimulated CTL recognizing this peptide, thus mimicking the response in mice primed by intranasal influenza infection. We also induced T cell proliferation and virus-specific CTL in cultures of normal T cells by stimulating with DC pulsed with the natural NP sequence 147-158 or the potent peptide 147-158 (Arg156-). Small numbers of peritoneal exudate cells, after activation with Con A to produce class II MHC expression and after removal of DC with a specific mAb (33DI), did not lead to primary CTL generation but initiated secondary stimulation in vitro. Our results using the hanging drop culture method and DC as APC have implications for studying the T cell repertoire for viral components in humans without the necessity of previous immunization.