Newton, Suzanne Elisabeth (2011) Characterization of the molecular and pharmacological roles of the human novel endokinin peptides. (PhD thesis), Kingston University, .
Abstract
The tachykinins are the largest known peptide family containing the highly conserved carboxyl terminal motif FXGLM-NH[sub]2, in which the mammalian tachykinin family consists of Substance P (SP), Neurokinin A (NKA), NKB and Endokinin B (EKB). The tachykinins mediate their effects through three G protein-coupled receptors, neurokinin (NK) 1, 2 and 3 in mammals. Each of the tachykinins can ligand bind with each of the receptors though with varying affinities, and both SP and EKB appear to exhibit a similar pattern of potency, with preference to the NK[sub]1 receptor. Many studies have reported EKB to elicit similar effects to SP; this raises the questions as to why there are two peptides with apparently similar functions. The aim of the present study was to establish whether there are any distinct roles between SP and EKB in the model cell line U251 MG, looking at their effects on growth, cell signalling and their expression patterns after stress-related conditions. In addition their expression patterns and promoter regions were analysed using bioinformatics tools. SP and EKB affected growth of U251 MG cells through the NK[sub]1 receptor. Additionally both peptides elicited activation of the PKC-cRaf-MEK1/2-ERK1/2- p90RSK pathway. Activation of this pathway triggered mRNA expression of MMP- 9. ICAM-1 expression was affected by SP through the NK[sub]1 receptor, though EKB would appear to reduce signalling similar to NK[sub]1 receptor antagonism suggesting EKB may block ICAM-1 signalling pathway. Bioinformatic analysis of expression patterns and promoter regions revealed the tachykinin gene (TAC) 1 may play a role in development and immune system processes more than other peptide genes. TAC4 had a high proportion of stress-related transcription factors, and was the only gene whose expression is predicted not to be restricted to the CNS, which may explain its peripheral expression. Additionally the tachykinin receptor gene (T ACR) 1 has been implicated in cell cycle control, which is in agreement with NK[sub]1 receptors' ability to affect proliferation in U251 MG cells as described above. Expression analysis of the tachykinin peptide and receptor genes in U251 MG cells under stress-related conditions (serum starvation, hypoxia and oxidative stress) revealed TAC4 to be involved with responses to oxidative stress, along with short isoform of NK[sub]1. TACR2 and TACR3 expression were up-regulated during serum starvation and hypoxic conditions, implicating them in adapting to these conditions. This study has revealed many differences between SP and novel endokinin peptides, and has revealed potential distinct roles for TAC4 from TAC1, with TAC4 implicated in a major role during oxidative stress. These tachykinins may play a major role in both cancer progression and strokes or traumatic brain injury (TBI). Targeting these tachykinins may well provide a novel treatment option for both of these conditions.
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