Induction of tryptase and histamine release from human colon mast cells by lgE dependent or independent mechanisms

AIM: To investigate the tryptase and histamine release ability of human colon mast cells upon IgE dependent or independent activation and the potential mechanisms.METHODS: Enzymatically dispersed cells from human colons were challenged with anti-IgE or calcium ionophore A23187, and the cell supernatants after challenge were collected. Both concentration dependent and time course studies with anti-IgE or calcium ionophore A23187 were performed. Tryptase release was determined with a sandwich ELISA procedure and histamine release was measured using a glass fibre-based fiuorometric assay.RESULTS:Both anti-IgE and calcium ionophore were able to induce dose dependent release of histamine from colon mast cells with up to approximately 60% and 25% net histamine release being achieved with 1μg/mL calcium ionophore and 10μg/mL anti-IgE, respectively. Dose dependent release of tryptase was also observed with up to approximately 19ng/mL and 21ng/mL release of tryptase being achieved with 10μg/mL anti-IgE and 1μg/mL calcium ionophore, respectively. Time course study revealed that both tryptase and histamine release from colon mast cells stimulated by anti-IgE initiated within 10 sec and reached their maximum release at 6 min following challenge. Pretreatment of cells with metabolic inhibitors abolished the actions of anti-IgE as well as calcium ionophore. Tryptase and histamine release, particularly that induced by calcium ionophore was inhibited by pretreatment of cells with pertussis toxin.CONCLUSION: Both anti-IgE and calcium ionophore are able to induce significant release of tryptase and histamine from colon mast cells, indicating that this cell type is likely to contribute to the pathogenesis of colitis and other mast cell associated intestinal diseases.

Activation of human colon mast cells through proteinase activated receptor-2

AIM:To investigate the ability of agonists of PAR-2 to stimulate release of tryptase and histamine from human colon mast cells and the potential mechanisms.METHODS:Enzymatically dispersed cells from human colons were challenged with tc-LIGRLO, tc-OLRGIL, SLIGKV,VKGILS, trypsin, anti-IgE or calcium ionophore A23187,and the cell supematants after challenge were collected. Tryptase release was determined with a sandwich ELISA procedure and histamine release was measured using a glass fibrebased fluorometric assay.RESULTS: Both PAR-2 agonists tc-LIGRLO-NH2 and SLIGKVNH2 were able to induce dose dependent release of tryptase and histamine from colon mast cells. More than 2.5 fold increase in both tryptase and histamine release was provoked by 100μmol/mL tc-LIGRLO-NH2, in comparison with only 2.0 fold increase being stimulated by SLIGKV-NH2，The reverse peptides tc-OLRGIL-NH2 and VKGILS-NH2 at the concentrations tested had no effect on the release of these two mediators.The maximum tryptase release elicited by tc-LIGRLO-NH2 was similar to that induced by anti-IgE(10μg/mL) or calcium ionophore (1μg/mL), though the latter was a more potent stimulus for histamine release.Both histamine and tryptase release in response to tc-LIGRLONH2 were completed within 3 rain. Trypsin at concentrations from 1.0 to 100μg/mL was capable of provoking a dose dependent release of tryptase as well as histamine with a maximum of 16ng/mL tryptase and 14ng/mL histamine release being achieved. An approximately 80% and 70% inhibition of trypsin induced release of tryptase and histamine were observed with SBTI, respectively. Pretreatment of cells with metabolic inhibitors or pertussis toxin abolished the actions of tc-LIGRLO-NH2, SLIGKV-NH2 and trypsin.CONCLUSION: The agonists of PAR-2 and trypsin are potent secretagogues of human colon mast cells, which are likely to contribute to the development of inflammatory disorders in human gut.