Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.4.21.4 (trypsin)
42,187 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We investigated the histochemical characteristics of mast cell tryptase in different mouse tissues. By use of peptide substrates, tryptase activity could be demonstrated in unfixed connective tissue mast cells in different tissues, including the stomach. Tryptase activity was better localized after aldehyde fixation and frozen sectioning, and under such conditions was also demonstrated in mucosal mast cells of the stomach but not in those of the gut mucosa. Double staining by enzyme histochemistry followed by toluidine blue indicated that the tryptase activity was present only in mast cells and that all mast cells in the stomach mucosa contained the enzyme. The peptide substrates z-Ala-Ala-Lys-4-methoxy-2-naphthylamide and z-Gly-Pro-Arg-4-methoxy-2-naphthlyamide, which are substrates of choice for demonstrating tryptase in other species, were most effective for demonstrating mouse tryptase. The use of protease inhibitors further indicated that activity present in all mast cells was tryptase. Safranin O did not stain stomach mucosal mast cells, suggesting that the tryptase present in these cells was active in the absence of heparin sulfate proteoglycan.
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PMID:Enzyme histochemistry of tryptase in stomach mucosal mast cells of the mouse. 1021 54

A second protease-activated receptor (PAR-2) that could be activated by trypsin or more physiologically by mast cell tryptase has been recently cloned. Both the structure and activation mechanism of PAR-2 was similar to the functional thrombin receptor (PAR-1). Although many effects of the coagulation protease thrombin on the vascular endothelium could be attributed to PAR-1 activation, very little is known about the physiological and pathophysiological role of PAR-2. We investigated whether stimulation of PAR-2 on endothelial cells induced two cellular responses that play a central role in primary and secondary haemostasis: the release of high molecular weight von Willebrand factor (hmw-VWF) from Weibel-Palade bodies and the de novo synthesis of tissue factor (TF) mRNA and protein. Human umbilical vein endothelial cells (HUVEC) were incubated with agonists for PAR-2 at 37 degrees C. Both trypsin and SLIGKV increased TF mRNA and activity and induced the release of hmw-VWF due to elevated levels of cytosolic Ca2+. Trypsin (10 nm) induced a 6-fold increase of TF mRNA and reduced time until fibrin clot formation to 37%, indicating trebling of the cell surface located TF activity. Stimulation of HUVEC with the PAR-2 agonist peptide SLIGKV induced a dose-dependent increase of TF mRNA up to 6 times and TF activity up to 3 times. Release of hmw-VWF was achieved both after incubation of HUVEC with trypsin and SLIGKV and was directly depending on intracellular Ca2+ mobilization. To make results comparable to the functional thrombin receptor, homologous experiments were carried out using the PAR-1 agonists thrombin and SFLLRN.
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PMID:Endothelial protease-activated receptor-2 induces tissue factor expression and von Willebrand factor release. 1023 35

Myeloperoxidase (MPO) is an important component of the neutrophil response to microbial infection. In this paper we report an additional activity of MPO, the potent and selective inhibition of human mast cell tryptase. MPO inhibits human mast cell tryptase in a time-dependent manner with an IC50 of 16 nM at 1 h. In contrast, MPO does not inhibit trypsin, thrombin, plasmin, factor Xa, elastase, or cathepsin G. It is the native protein conformation of MPO and not its enzyme activity that is responsible for tryptase inhibition. Heparin, at high concentrations, can prevent the inhibition of tryptase by MPO. We have shown by size-exclusion chromatography that MPO promotes the dissociation of active tryptase tetramer to inactive monomer. These data suggest that MPO inhibits tryptase by interfering with the heparin stabilization of tryptase tetramer. We have previously shown that lactoferrin (another neutrophil-associated protein) also inhibits tryptase activity by a similar mechanism. The finding that MPO is a potent inhibitor of tryptase lends further support to the hypothesis that neutrophil proteins, such as MPO and lactoferrin, may play a regulatory role as endogenous suppressers of tryptase enzyme activity.
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PMID:Neutrophil myeloperoxidase is a potent and selective inhibitor of mast cell tryptase. 1033 72

Polymorphic eruption of pregnancy (PEP) and herpes gestationis (HG) are pregnancy-related dermatoses of unknown aetiology with eosinophil infiltration which, at early stages, may show similar clinical and histopathological features. To determine the relative contributions of eosinophils, neutrophils and mast cells to the pathogenesis of PEP and HG through deposition of granule proteins, we studied tissue and serum from 15 patients with PEP and 10 with HG. Using indirect immunofluorescence with antibodies to human eosinophil granule major basic protein (MBP), eosinophil-derived neurotoxin (EDN), eosinophil cationic protein (ECP), neutrophil elastase and mast cell tryptase, we determined and compared cellular and extracellular staining patterns in lesional skin biopsy specimens and, using immunoassay, measured MBP, EDN, and ECP in patients' sera. Eosinophil infiltration and extracellular protein deposition of all three eosinophil granule proteins were present in both PEP and HG indicating a pathogenic role for eosinophils in both diseases. Staining for eosinophil granule proteins was especially prominent in urticarial lesions and around blisters in HG. EDN and ECP serum levels in PEP and ECP serum levels in HG were significantly increased compared with those in normal pregnant and normal nonpregnant serum. Neutrophils were more prominent in HG specimens than in PEP specimens; extracellular neutrophil elastase was minimally present and similar in both diseases. Mast cell numbers and extracellular tryptase deposition did not differ between the two diseases and did not differ from mast cell counts in skin of normal pregnant women. This study shows that eosinophil granule proteins are deposited extracellularly in tissue and are increased in serum in both PEP and HG. Moreover, eosinophil involvement in the two diseases is more consistent than neutrophil and mast cell involvement. Comparatively, tissue eosinophil infiltration and extracellular protein deposition is more extensive in HG than in PEP, suggesting that eosinophil involvement is greater in the pathogenesis of HG than PEP and similar to that found in bullous pemphigoid.
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PMID:Polymorphic eruption of pregnancy and herpes gestationis: comparison of granulated cell proteins in tissue and serum. 1035 84

1. Proteases regulate cells by cleaving proteinase-activated receptors (PARs). Thrombin and trypsin cleave PAR-1 and PAR-2 on neurons and astrocytes of the brain to regulate morphology, growth and survival. We hypothesized that thrombin and mast cell tryptase, which are generated and released during trauma and inflammation, regulate enteric neurons by cleaving PAR-1 and PAR-2. 2. We detected immunoreactive PAR-1 and PAR-2 in > 60 % of neurons from the myenteric plexus of guinea-pig small intestine in primary culture. A large proportion of neurons that expressed substance P, vasoactive intestinal peptide or nitric oxide synthase also expressed PAR-1 and PAR-2. We confirmed expression of PAR-1 and PAR-2 in the myenteric plexus by RT-PCR using primers based on sequences of cloned guinea-pig receptors. 3. Thrombin, trypsin, tryptase, a filtrate from degranulated mast cells, and peptides corresponding to the tethered ligand domains of PAR-1 and PAR-2 increased [Ca2+]i in > 50 % of cultured myenteric neurons. Approximately 60 % of neurons that responded to PAR-1 agonists responded to PAR-2 agonists, and > 90 % of PAR-1 and PAR-2 responsive neurons responded to ATP. 4. These results indicate that a large proportion of myenteric neurons that express excitatory and inhibitory neurotransmitters and purinoceptors also express PAR-1 and PAR-2. Thrombin and tryptase may excite myenteric neurons during trauma and inflammation when prothrombin is activated and mast cells degranulate. This novel action of serine proteases probably contributes to abnormal neurotransmission and motility in the inflamed intestine.
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PMID:Thrombin and mast cell tryptase regulate guinea-pig myenteric neurons through proteinase-activated receptors-1 and -2. 1035 15

Proteolytically activated receptors define a new subclass among the G-protein coupled receptors. Proteinase activated receptor-2 (PAR-2), the second member to be identified of this growing receptor subclass, can be activated by trypsin and trypsin-like serine proteases such as mast cell tryptase. PAR-2 is expressed in endothelial cells. Here we have studied if activation of PAR-2 changes the coagulation properties of cultured human umbilical vein endothelial cells. We show that activation of PAR-2 induces rapid and transient formation of tissue factor mRNA with a maximum level 1 hour after receptor stimulation. The increased mRNA level was accompanied by an increased tissue factor activity at the endothelial cell surface, shortening coagulation time in a standard clotting assay. The level of tissue factor activity after PAR-2 activation was comparable with the effects of thrombin receptor (PAR-1) activation although neither of the two protease receptors were as strong inducers of tissue factor as tumor necrosis factor-alpha.
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PMID:Stimulation of proteinase activated receptor-2 causes endothelial cells to promote blood coagulation in vitro. 1040 79

Proteinase-activated receptor-2 (PAR-2) is a G-protein coupled receptor. Tryptic proteases cleave PAR-2 exposing a tethered ligand (SLIGKV), which binds and activates the receptor. Although PAR-2 is highly expressed by cultured keratinocytes and is an inflammatory mediator, its precise localization in the normal and inflamed human skin is unknown, and the proteases that activate PAR-2 in the skin have not been identified. We localized PAR-2 in human skin by immunohistochemistry, examined PAR-2 expression by RT-PCR and RNA blotting, and investigated PAR-2 activation by mast cell tryptase. PAR-2 was localized to keratinocytes, especially in the granular layer, to endothelial cells, hair follicles, myoepithelial cells of sweat glands, and dermal dendritic-like cells. PAR-2 was also highly expressed in keratinocytes and endothelial cells of inflamed skin. PAR-2 mRNA was detected in normal human skin by RT-PCR, and in cultured human keratinocytes and dermal microvascular endothelial cells by Northern hybridization. Trypsin, tryptase and a peptide corresponding to the tethered ligand (SLIGKVNH2) increased [Ca2+]i in keratinocytes, measured using Fura-2/AM. Although tryptase-containing mast cells were sparsely scattered in the normal dermis, they were numerous in the dermis in atopic dermatitis, and in the dermis, dermal-epidermal border, and occasionally within the lower epidermis in psoriasis. Tryptase may activate PAR-2 on keratinocytes and endothelial cells during inflammation.
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PMID:Proteinase-activated receptor-2 in human skin: tissue distribution and activation of keratinocytes by mast cell tryptase. 1043 26

The possible involvement of mast cell tryptase and chymase in subepidermal bullous diseases was studied enzyme-histochemically in specimens from erythematous and vesicular skin and from non-involved skin of patients with dermatitis herpetiformis, bullous pemphigoid, erythema multiforme, infective bullous eruption and linear IgA dermatosis. Patients with pemphigus were biopsied for comparison. The immunoreactivity of chymase inhibitors, alpha1-proteinase inhibitor (alpha1-PI) and alpha1-antichymotrypsin (alpha1-AC), in mast cells was demonstrated using the sequential double staining method. Tryptase-positive mast cells were unchanged or only slightly increased in number in erythematous lesions and slightly decreased in blistering skin compared with healthy-looking skin. Only occasionally were mast cells seen in apparent contact with the basement membrane zone. Chymase-positive mast cells and the chymase/tryptase ratio steadily decreased during the development of the lesions in each subepidermal bullous disease. The percentage of alpha1-PI+ and/or alpha1-AC+ mast cells increased simultaneously, which could explain the disappearance of chymase activity. Similar results were obtained regardless of the bullous disease. The results were also similar in pemphigus, which is an intraepidermal bullous disease. In conclusion, these results show significant alterations in mast cell chymase and protease inhibitors in a range of different bullous diseases, suggesting mast cell involvement. The apparent inactivation of chymase could be due to the action of chymase inhibitors detected in numerous mast cells. However, these alterations probably reflect general inflammation rather than a specific reaction in a certain bullous disease.
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PMID:Mast cells in developing subepidermal bullous diseases: emphasis on tryptase, chymase and protease inhibitors. 1049 9

Emerging evidence suggests that mast cell tryptase is a therapeutic target for the treatment of asthma. The effects of this serine protease are associated with both pathophysiologic pulmonary responses and pathologic changes of the asthmatic airway. In this study, the tryptase inhibitor 1,5-bis-[4-[(3-carbamimidoyl-benzenesulfonylamino)-methyl]-p henoxy]-pentane (AMG-126737) was evaluated for its pharmacologic effects against allergen-induced airway responses. AMG-126737 is a potent inhibitor of human lung mast cell tryptase (Ki = 90 nM), with greater than 10- to 200-fold selectivity versus other serine proteases. Intratracheal administration of AMG-126737 inhibited the development of airway hyperresponsiveness in allergen-challenged guinea pigs with an ED50 of 0.015 mg/kg. In addition, the compound exhibited oral activity in the guinea pig model. The in vivo activity of AMG-126737 was confirmed in a sheep model of allergen-induced airway responses, where the compound inhibited early and late phase bronchoconstriction responses and the development of airway hyperresponsiveness. These results support the proposed role of tryptase in the pathology of asthma and suggest that AMG-126737 has potential therapeutic utility in this pulmonary disorder.
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PMID:Inhibition of allergen-induced pulmonary responses by the selective tryptase inhibitor 1,5-bis-[4-[(3-carbamimidoyl-benzenesulfonylamino)-methyl]-phenoxy]-pen tane (AMG-126737). 1059 Nov 55

A markedly elevated serum level of mast cell tryptase (77.6 microg/L; 95th percentile in normals 13.5 microg/L) was detected in a patient treated for 5 years with wasp venom immunotherapy because of severe anaphylaxis following a wasp sting. Retrospective analysis of stored serum samples taken during the course of immunotherapy revealed that the tryptase level had been elevated for at least 3 years. Despite several dermatological examinations, skin changes of mastocytosis had been missed. Re-examination of the patient revealed sparse macules on the thorax and thighs; Darier's sign was negative. Histologically, mast cell accumulation in these lesions was demonstrable. No signs of systemic mastocytosis were detected. The most appropriate diagnosis was telangiectasia macularis eruptiva perstans. Even in patients with highly elevated tryptase levels, mastocytosis may go undiagnosed. As mastocytosis predisposes to severe anaphylaxis, the condition should be looked for in patients with such reactions by clinical examination and measurement of serum tryptase levels.
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PMID:Mastocytosis associated with severe wasp sting anaphylaxis detected by elevated serum mast cell tryptase levels. 1060 62


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