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AIM:To investigate the ability of agonists of PAR-2 tostimulate release of tryptase and histamine from human colonmast cells and the potential mechanisms.METHODS:Enzymatically dispersed cells from humancolons were challenged with tc-LIGRLO,tc-OLRGIL,SLIGKV,VKGILS,trypsin,anti-IgE or calcium ionophore A23187,andthe cell supernatants after challenge were collected.Tryptaserelease was determined with a sandwich ELISA procedureand histamine release was measured using a glass fibre-based fluorometric assay.RESULTS:Both PAR-2 agonists tc-LIGRLO-NH_2 and SLIGKV-NH_2 were able to induce dose dependent release of tryptaseand histamine from colon mast cells.More than 2.5 foldincrease in both tryptase and histamine release was provokedby 100μmol/mL tc-LIGRLO-NH_2,in comparison with only2.0 fold increase being stimulated by SLIGKV-NH_2.Thereverse peptides tc-OLRGIL-NH_2 and VKGILS-NH_2 at theconcentrations tested had no effect on the release of thesetwo mediators.The maximum tryptase release elicited bytc-LIGRLO-NH_2 was similar to that induced by anti-IgE(10μg/mL)or calcium ionophore(1μg/mL),though the latterwas a more potent stimulus for histamine release.Bothhistamine and tryptase release in response to tc-LIGRLO-NH_2 were completed within 3 min.Trypsin at concentrationsfrom 1.0 to 100μg/mL was capable of provoking a dosedependent release of tryptase as well as histamine with amaximum of 16 ng/mL tryptase and 14 ng/mL histaminerelease being achieved.An approximately 80% and 70%inhibition of trypsin induced release of tryptase and histaminewere observed with SBTI,respectively.Pretreatment of cellswith metabolic inhibitors or pertussis toxin abolished theactions of tc-LIGRLO-NH_2,SLIGKV-NH_2 and trypsin.CONCLUSION:The agonists of PAR-2 and trypsin arepotent secretagogues of human colon mast cells,which arelikely to contribute to the development of inflammatorydisorders in human gut.
AIM: To investigate the ability of agonists of PAR-2 tostimulate release of tryptase and histamine from human colonmast cells and the potential mechanisms. METHODS: Enzymatically dispersed cells from human colon were challenged with tc-LIGRLO, tc-OLRGIL, SLIGKV, VKGILS, trypsin , anti-IgE or calcium ionophore A23187, and the cell supernatants after challenge were challenged were collected.Tryptase release was determined with a sandwich ELISA procedure and histamine release was measured using a glass fiber-based fluorometric assay .RESULTS: Both PAR-2 agonists tc-LIGRLO-NH_2 and SLIGKV-NH_2 were able to induce dose dependent release of tryptase and histamine from colon mast cells. More than 2.5 fold in crease in both tryptase and histamine release was provokedby 100 μmol / mL tc-LIGRLO-NH_2, in comparison with only 2.0 fold increase being stimulated by SLIGKV-NH_2.Thereverse peptides tc-OLRGIL-NH_2 and VKGILS-NH_2 at theconcentrations tested had no effect on the release of thesetwo mediators.The maximum tryptase release elic ited bytc-LIGRLO-NH_2 was similar to that induced by anti-IgE (10 μg / mL) or calcium ionophore (1 μg / mL) though though latter more a potent stimulus for histamine release. Posthistamine and tryptase release in response to tc-LIGRLO -NH 2 were completed within 3 min. Trypsin at concentrations from 1.0 to 100 μg / mL was capable of provoking a dose dependent release of tryptase as well as histamine with a maximum of 16 ng / mL tryptase and 14 ng / mL histamine release. and 70% inhibition of trypsin induced release of tryptase and histaminewere observed with SBTI, respectively. Treatment of cells with metabolic inhibitors or pertussis toxin abolished theactions of tc-LIGRLO-NH_2, SLIGKV-NH_2 and trypsin.CONCLUSION: The agonists of PAR-2 and trypsin arepotent secretagogues of human colon mast cells, which arelikely to contribute to the development of inflammatory disorders in human gut.