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Hemokinin-1 (HK-1) is a mammalian tachykinin that originates primarily from non-neuronal cells. HK-1 and substance P (SP) signal through the neurokinin 1 (NK1) receptor, and exhibit overlapping immune and inflammatory properties. The first aim of this thesis was to examine the inflammatory role of HK-1 in the human colonic mucosa, compared with SP. Functional studies demonstrated that HK-1 evoked the release of prostaglandin E2 (PGE2) in a time- and concentration-dependent manner, and these actions required the activation of both NK1 and NK2 receptors as well as a cyclooxygenase-2 (COX-2) -dependent mechanism. Molecular studies showed that HK-1 not only stimulated COX-2 expression but also suppressed 15-hydroxyprostaglandin dehydrogenase (15-PGDH) expression at both gene and protein levels, suggesting an additional mechanism for HK-1-evoked PGE2 release. Furthermore, functional and molecular studies demonstrated that HK-1 increased the generation of multiple proinflammatory cytokines and chemokines, at both gene and protein levels. These effects were mediated by both NK1 and NK2 receptors. In parallel studies, SP had very similar responses in stimulating PGE2, proinflammatory cytokines and chemokines, but no effect on the expression of monocyte chemoattractant protein-1 (MCP-1) and regulated on activation, normal T cell expressed and secreted (RANTES). In summary, these studies demonstrated HK-1 had very similar actions with SP in inducing inflammatory mediators from the human colonic mucosa. The second aim of this thesis was to characterise the expression of PGE2-related enzymes in the human colonic mucosa from control and disease patients. Disease colonic tissues were obtained from the marginal resection specimens of patients with ulcerative colitis (UC), Crohn’s disease (CD) and acute diverticular disease (DD). Tissues from UC and CD appeared histologically abnormal; however, those from acute DD appeared relatively normal. In UC and CD, COX-2 expression was increased, and 15-PGDH expression was reduced. COX-1 was unchanged. In contrast, the decrease of COX-2 and the increase of 15-PGDH was seen in acute DD. A trend towards a decrease in COX-1 was also noted. In summary, these studies demonstrated the inverse mucosal expression of PGE2-related enzymes in acute DD compared to IBD, and suggests the implication of low PGE2 level in acute DD.

My research involves biochemical analysis of venom from a fascinating parasitoid jewel wasp Ampulex compressa. Most parasitoid wasps envenomate the host by stinging into the body cavity to cause paralysis and developmental arrest, prior to deposition of eggs externally or within the body cavity. A. compressa instead uses a different subjugation strategy by injecting venom directly into the central nervous system, eliciting a behavioral sequence culminating in hypokinesia, a 7-10 day lethargy advantageous to wasp reproduction. Hypokinesia is a specific, venom-induced behavioral state characterized by suppression of the escape response and reduced spontaneous walking, leaving other motor functions unaffected. This specificity of action is particularly unique among venoms and interestingly, effects of the venom on the escape response are reversible as the cockroach may recover after 7-10 days if not consumed by the wasp larvae. Venom-induced hypokinesia raises an interesting biological question: How can such a potent biochemical cocktail cause such long-lasting, specific, yet reversible effects on behavior? I approached this question in two ways: objective one - bioinformatic analysis of the venom and venom gland tissue to determine what the venom is made of, and objective two - functional analysis of key venom components to determine how the venom works. To address objective 1, I used advanced bioinformatics techniques to generate transcriptomes of the venom tissue and proteomes of the venom and venom tissue. Next generation sequencing of venom gland RNA has yielded full-length coding sequences and quantification of venom transcript levels, while mass spectroscopy based protein analysis has validated the presence of venom proteins. These analyses will allow construction of a comprehensive A. compressa “venome” that will help inform functional analyses of the venom and its role in hypokinesia induction. For objective two, I focused on the characterization of the most abundant peptide in the venom, tentatively named Ampulexin 1, and pharmacological analysis of an interesting venom peptide neurotransmitter, called tachykinin. Analysis of venom tachykinin action on cockroach brain receptors may reveal an interesting case of the evolution of a neurotransmitter from one animal to target the nervous system of another.