UBC Theses and Dissertations
Lipopolysaccharide signaling in endothelial cells Dauphinee, Shauna Marie
The endothelium plays a critical role in coordinating the innate immune response through the regulation of vascular tone, leukocyte recruitment and transmigration, and hemostasis. These functions are mediated, in part, by the signaling cascades initiated upon recognition of bacterial and viral products by a family of transmembrane receptors known as Toll-like receptors (TLRs). In endothelial cells, exposure to lipopolysaccharide (LPS), a major cell wall constituent of Gram negative bacteria, results in endothelial activation through TLR4. Recruitment of the adapter protein, MyD88, to the receptor facilitates association of serine threonine kinases of the IL-1 receptor associated kinase (IRAK) family. The IRAKs initiate a phosphorylation cascade through TNFR-associated factor 6 (TRAF6) culminating in activation of proinflammatory signaling pathways including NF-κB and c-Jun NH2-terminal kinase (JNK) pathways. This thesis investigates signaling molecules and pathways downstream of TLR4 in endothelial cells. Specifically, contained herein is a description of the role of heterotrimeric G proteins in endothelial TLR signaling. This thesis identifies for the first time the function of these proteins in multiple TLR signaling pathways. In addition, the work presented here describes the identification and characterization of a novel TLR4 signaling molecule, SAM and SH3 domain containing protein 1 (SASH1). SASH1 promotes LPS-induced NF-κB and JNK, by functioning as a scaffold molecule to bind TRAF6, transforming growth factor-β-activated kinase (TAK1) and IκB-kinase (IKK), thereby increasing proinflammatory cytokine production. The distinct functions of the endothelium in innate immunity highlight the need for an understanding of the signaling cascades initiated by LPS in endothelial cells and will be crucial to our understanding of the pathophysiology of sepsis in the clinic.
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