Host responses to Salmonella typhimurium infection in vitro and in vivo

Thesis (Ph. D.)--University of Washington, 2003Salmonella spp. are facultative intracellular pathogens capable of causing localized and systemic disease of significant morbidity and mortality; disease can be prevented by oral immunization with viable attenuated bacteria. During the Salmonella-host interaction, multiple processes occur that determine the outcome of infection. In this dissertation, I examine several events---bacterial induction of programmed host cell death, innate recognition of bacterial motifs, and adaptive immune responses to microbial antigens.Prior studies observed Salmonella invasion of macrophages induced apoptosis, or non-inflammatory programmed cell death. Results presented here suggest that Salmonella kills macrophages by a unique mechanism distinct from apoptosis and necrosis. Unlike apoptosic cells, Salmonella-infected macrophages displayed diffuse rather than condensed patterns of DNA fragmentation and lacked caspase-3 enzymatic activity, but like necrotic cells, infected macrophages exhibited membrane damage and retained the enzyme PARP in its active uncleaved state. Unique to Salmonella -induced macrophage death was the requirement for caspase-1 activity. These results demonstrate that Salmonella infection of macrophages triggers caspase-1-dependent pro-inflammatory necrosis. During Salmonella infection, CD4+ T cells respond to FliC, the major subunit protein of the flagellar apparatus. Described here is further examination of the CD4+ T cell response to FliC and identification of four discrete FliC epitopes with varying immunodominance in vitro and in vivo . Analysis of CD4+ T cell responses to non-FliC antigens identified unique surface organelles as a rich source of natural antigens. Salmonella antigens directly stimulated Toll-like receptors (TLRs) or were intimately associated with TLR ligands, suggesting that TLR recognition biases T cell responses to specific antigens. Salmonella evaded innate and adaptive immune recognition by modifying or repressing expression of natural antigens during growth in vivo; bacterial regulation of antigen expression was shown to occur at transcriptional and post-transcriptional levels. Experimental dysregulation of FliC expression during in vivo infection profoundly influenced the ensuing mucosal CD4+ T cell response to FliC, indicating that Salmonella preferentially expresses FliC during colonization of the mucosa. These results demonstrate that regulated antigen expression can influence antigen-specific immune responses, and may enable Salmonella to evade immune recognition and continue replication in host tissue