HUMAN MAST-CELL TRYPTASE - A STIMULUS OF MICROVASCULAR LEAKAGE AND MAST-CELL ACTIVATION

We have investigated the potential of tryptase to stimulate an increas e in microvascular permeability following injection into the skin of g uinea pigs. Tryptase was isolated from high salt extracts of human lun g tissue by octyl-agarose and heparin-agarose chromatography. Injectio n of purified tryptase (2.5 ng-2.5 mu g/site) into the skin of guinea pigs which had been injected intravenously with Evans blue dye provoke d a dose-dependent increase in microvascular permeability. The skin re actions elicited by tryptase were apparent up to 80 min following inje ction, while histamine-induced microvascular leakage resolved complete ly by 40 min. Heat-inactivation of tryptase, or preincubating the prot einase with certain proteinase inhibitors, significantly reduced the e xtent of microvascular leakage, suggesting dependency on an intact cat alytic site. No evidence was found or a synergistic or antagonistic in teraction between tryptase (2.5 ng-2.5 mu g/site) and histamine (1-10 mu g/site) when these mast cell products were injected together. Addit ion of heparin to tryptase (10:1; w/w) prior to injection was without effect on tryptase-induced microvascular leakage. Pretreatment of guin ea pigs with a combination of the histamine H-1 receptor antagonist py rilamine and the histamine H-2 receptor antagonist cimetidine (both 10 mg/kg), partially abolished tryptase-induced microvascular leakage as well as attenuating the reaction to histamine. Reasoning that the mic rovascular leakage induced by tryptase is likely to involve the releas e of histamine, we investigated the ability of tryptase to stimulate h istamine release from dispersed guinea-pig skin and lung cells in vitr o. Tryptase was found to induce concentration-dependent histamine rele ase from both sources of tissue. Mast cell activation stimulated by tr yptase in vitro was inhibited by heat treating the enzyme or by additi on of proteinase inhibitors, suggesting a requirement for an intact ca talytic site. Histamine release was inhibited also by preincubating ce lls with the metabolic inhibitors antimycin A and 2-deoxy-D-glucose in dicating that the mechanism was energy-requiring and non-cytotoxic. We conclude that human mast cell tryptase may be a potent stimulus of mi crovascular leakage. The activation of mast cells by this proteinase m ay represent an amplification process in allergic inflammation.