| The term prostanoids collectively describes prostaglandins, prostacyclin and thromboxanes. These compounds are products of arachidonic acid metabolism by the cyclooxygenase pathway. Prostanoids mediate various physiological and pathophysiological effects through their interaction with membrane-bound receptors. In this research, a thorough characterization of the recombinant human (h) PGD2 receptor (DP) was performed, with respect to its radioligand binding and signal transduction properties using prostanoids and prostanoid analogues. The recombinant hDP receptor was then used along with other recombinant human prostanoid receptors to identify a novel specific agonist, L-644,698. This compound exhibits high affinity and potency at the hDP receptor. Moreover. L-644,698 demonstrates at least 300-fold higher selectivity for the hDP receptor than for any of the other seven recombinant human prostanoid receptors tested. Thus. L-644,698 is one of the most selective DP-specific agonists as yet described. Subsequently, the rat (r) DP receptor was cloned and functionally expressed. Pharmacological characterization using L-644,698 and other DP-specific ligands confirmed the identity of this protein as a homologue of human DP, and validated the use of the cDNA corresponding to rDP as a template from which to make rDP-specific riboprobes for in situ hybridization studies. mRNA corresponding to rDP was localized to the CNS and GI tract by the in situ hybridization technique. Within the GI tract. rDP-specific signals were observed repeatedly in the mucous-secreting goblet cells and, less often, in the adjacent epithelium of the stomach, duodenum, ileum, and colon. These observations corroborate prior data demonstrating an abundance of both hDP- and mDP-specific mRNA in G1 tract tissues (especially in small intestine), and suggest a novel biological role for the DP receptor, namely the regulation of mucin secretion. DP-specific mRNA was then localized to the mucous-secreting goblet cells of the human colon, justifying the use of an established in vitro cell model of human origin for the study of mucin secretion, the LS174T colonic adenocarcinoma cell line. The endogenous expression of the hDP receptor on LS174T cells was confirmed pharmacologically using L-644,698 and other DP-specific ligands. Subsequently, actuation of hDP was shown to stimulate mucin secretion in this cell line, through the use of DP-specific agonists (PGD2, L-644,698) and a DP-specific antagonist (BW A868C). |
