He S, Gaça M D, Walls A F
Immunopharmacology Group, University of Southampton, Southampton General Hospital, United Kingdom.
J Pharmacol Exp Ther. 1998 Jul;286(1):289-97.
Tryptase, the most abundant protein product of human mast cells is emerging as an important mediator and target for therapeutic intervention in allergic disease. We have investigated the potential of tryptase and inhibitors of tryptase to modulate histamine release from human mast cells. Addition of purified human tryptase in concentrations ranging from 1 to 100 mU/ml stimulated a concentration-dependent release of histamine from cells dispersed from tonsil, although not from skin tissue. The reaction dependent on an intact catalytic site being inhibited by heat inactivation of the enzyme, or by preincubating with the tryptase inhibitors APC366 or leupeptin or the tryptic substrate N-benzoyl-DL-arginine-p-nitroanilide (BAPNA). Tryptase-induced histamine release took approximately 6 min to reach completion, appeared to require exogenous calcium and magnesium, and on the basis of inhibition by antimycin A and 2-deoxy-D-glucose, seemed to be a noncytotoxic process. Pre-incubation of cells with tryptase at concentrations that were suboptimal for histamine release had little effect on their responsiveness to anti-immunoglobulin (Ig) E or to calcium ionophore A23187, but at higher concentrations their subsequent activation was inhibited. APC366 significantly inhibited histamine release induced by anti-IgE or calcium ionophore from both tonsil and skin cells, with up to 90% inhibition being observed at a concentration of 100 microM with skin. IgE-dependent histamine release was inhibited also by leupeptin, benzamidine and BAPNA. Tryptase may act as an amplification signal for mast cell activation, and this could account at least partly for the potent mast cell stabilizing properties of tryptase inhibitors.
类胰蛋白酶是人类肥大细胞中含量最丰富的蛋白质产物，正逐渐成为过敏性疾病治疗干预的重要介质和靶点。我们研究了类胰蛋白酶及其抑制剂调节人肥大细胞组胺释放的潜力。添加浓度范围为1至100 mU/ml的纯化人源类胰蛋白酶，可刺激从扁桃体分散的细胞组胺浓度依赖性释放，但皮肤组织细胞无此现象。该反应依赖完整的催化位点，可被酶的热失活、与类胰蛋白酶抑制剂APC366或亮抑蛋白酶肽预孵育、或与胰蛋白酶底物N-苯甲酰-DL-精氨酸对硝基苯胺（BAPNA）预孵育所抑制。类胰蛋白酶诱导的组胺释放约6分钟完成，似乎需要外源性钙和镁，基于抗霉素A和2-脱氧-D-葡萄糖的抑制作用，这似乎是一个非细胞毒性过程。用对组胺释放次优的浓度的类胰蛋白酶预孵育细胞，对其对抗免疫球蛋白（Ig）E或钙离子载体A23187的反应性影响不大，但在较高浓度下其随后的激活受到抑制。APC366显著抑制抗IgE或钙离子载体诱导的扁桃体和皮肤细胞组胺释放，在皮肤细胞中浓度为100 microM时观察到高达90%的抑制。亮抑蛋白酶肽、苯甲脒和BAPNA也抑制IgE依赖性组胺释放。类胰蛋白酶可能作为肥大细胞激活的放大信号，这至少可以部分解释类胰蛋白酶抑制剂强大的肥大细胞稳定特性。
