Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: UNIPROT:P15088 (mast cell)
 14,925 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Tissue histamine is stored in mast cell granules, presumably as a histamine-heparin complex. Heparin is a polyelectrolyte, with a fraction of its anionic charge neutralized by condensed counterions. The interaction of heparin with histamine in aqueous solution was quantitatively characterized by 1H nuclear magnetic resonance (NMR) spectroscopy. Binding constants were determined from chemical shift-pH titration data for the C2H proton of the imidazolium ring for a wide range of histamine, heparin, and Na+ concentrations. The results indicate a binding stoichiometry of 1 histamine per heparin disaccharide repeat unit. The binding is electrostatic, as indicated by the strong dependence of the binding constant on Na+ concentration. From an analysis of the binding constants using the counterion condensation theory of polyelectrolytes, it was determined that the binding of H2A2+ results in displacement of 1.72 Na+ ions from the counterion condensation volume of heparin and that H2A2+ makes 2 ionic interactions with heparin. The displacement of Na+ from the counterion condensation volume of heparin by H2A2+ was also studied by 23Na NMR. From 23Na spin-lattice relaxation time data, it was determined directly that 1.78 Na+ ions are displaced per H2A2+ bound by heparin. The results are discussed in terms of the ion exchange process which takes place when histamine is released by mast cells.
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PMID:Quantitative characterization of the binding of histamine by heparin. 976 Feb 48

The addition of rat mast cell granules to confluent bovine pulmonary artery endothelial cell monolayers resulted in the formation of numerous lacunae in the cultures. Several lines of evidence identified heparin proteoglycan as the component of the granule matrix responsible for the effect: presence of the activity in the proteoglycan fraction after chromatography of granule extracts, inhibition of granule activity by digestion with heparinase I, the failure of proteolysis of the proteoglycan fraction with proteinase K to significantly diminish its activity, and the failure of chymase and carboxypeptidase inhibitors to inhibit granule activity. The onset of hole formation was delayed for several hours after granule addition to the culture, and maximal hole formation occurred between 8 and 16 hours and was sustained as long as 24 hours. The lacunae formed by the separation of motile endothelial cells within the monolayer and was not attributable to cell contractile activity or cell loss. Time-lapse video recording showed that the holes were dynamic, individual holes expanding and regressing over a period of hours. Formation of lacunae occurred on gelatin and fibronectin surfaces alike. The presence of active chymase in the granules prevented the action of the proteoglycan. Heparin glycosaminoglycan as distinct from the proteoglycan did not similarly affect the endothelial monolayers but did block the action of granules added subsequently, indicating the likelihood of a heparin-reactive receptor or binding site.
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PMID:Mast cell granule heparin proteoglycan induces lacunae in confluent endothelial cell monolayers. 1032 11

Different subpopulations of mast cells are characterized by their abundant contents of either tryptase or in addition chymase. These two neutral proteases are found in mast cells and may thus hold a key to the understanding of mast cell dependent reactions. Such studies are however hampered by the lack of readily available supplies of chymase. We have therefore studied the simultaneous purification of both proteases from hairless moro hr/hr mouse skin, using a sequence of salt extractions and chromatographic separations. After three steps of extraction, a 13-fold purification with an 82% yield was obtained for tryptase and a 15-fold purification with a 90% yield for chymase. Further one step purification on conventional sephadex, sephacryl and octyl sepharose columns was unsatisfactory because of further protein contamination of the fractions. Heparin affinity chromatography caused a high loss of tryptase and residual protein contamination. Gradient elution on a benzamidine sepharose 6B column resulted however in a single, low yield (17.9%) tryptase peak and a broader, high yield (>90%) chymase peak, and a 34% yield high purity fraction. The proteases thus purified exhibited their typical inhibitor profile. On Western blot analysis and on autoradiography in the presence of the serine protease inhibitor diisopropylfluorophosphate (DFP), only one 28 kD molecule with chymase activity was identified, whereas a broad 32-38 kD band of tryptase monomers was noted. Taken together, these data show that, after salt extraction and a single benzamidine affinity chromatography step, both mast cell chymase and tryptase can be separated and in case of chymase also highly purified, allowing thus for the study of biological activities of this molecule.
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PMID:Purification of mast cell proteases from murine skin. 1053 69

Mouse mast cell protease (mMCP) 6 and mMCP-7 are homologous tryptases stored in granules as macromolecular complexes with heparin and/or chondroitin sulfate E containing serglycin proteoglycans. When pro-mMCP-7 and pseudozymogen forms of this tryptase and mMCP-6 were separately expressed in insect cells, all three recombinant proteins were secreted into the conditioned medium as properly folded, enzymatically inactive 33-kDa monomers. However, when their propeptides were removed, mMCP-6 and mMCP-7 became enzymatically active and spontaneously assumed an approximately 150-kDa tetramer structure. Heparin was not required for this structural change. When incubated at 37 degrees C, recombinant mMCP-7 progressively lost its enzymatic activity in a time-dependent manner. Its N-linked glycans helped regulate the thermal stability of mMCP-7. However, the ability of this tryptase to form the enzymatically active tetramer was more dependent on a highly conserved Trp-rich domain on its surface. Although recombinant mMCP-6 and mMCP-7 preferred to form homotypic tetramers, these tryptases readily formed heterotypic tetramers in vitro. This latter finding indicates that the tetramer structural unit is a novel way the mast cell uses to assemble varied combinations of tryptases.
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PMID:Formation of enzymatically active, homotypic, and heterotypic tetramers of mouse mast cell tryptases. Dependence on a conserved Trp-rich domain on the surface. 1061 25

A novel trypsin-type serine proteinase, which processes the precursors of the envelope fusion glycoproteins of pneumotropic Sendai and human influenza A viruses, was purified to homogeneity from pig lungs. On SDS/PAGE, the purified enzyme gave a protein band corresponding to about 32 kDa, and has an apparent molecular mass of 120 kDa, as determined by gel permeation chromatography. Immunohistochemical staining with antibodies against this enzyme revealed that the enzyme is located in pig lung mast cells. The N-terminal 44-amino-acid sequence of the enzyme exhibits about 80% identity with those of mast cell tryptases from other species. Of the inhibitors tested, di-isopropyl fluorophosphate, antipain, leupeptin, benzamidine and a few proteinaceous inhibitors, such as mucus protease inhibitor and aprotinin, inhibited this enzyme activity. Heparin stabilized the enzyme, but high-ionic-strength conditions did not, unlike for human mast cell tryptase. The purified enzyme efficiently processed the fusion glycoprotein precursor of Sendai virus and slowly processed hemagglutinin of human influenza A virus, and triggered the infectivity of Sendai virus in a dose-dependent manner, although human mast cell tryptase beta and rat mast cell tryptase (rat MCP-7) from lungs did not process these fusion glycoproteins at all. These results suggest that mast cell tryptase in pig lungs is the possible trigger of the pneumotropic virus infections.
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PMID:Mast cell tryptase from pig lungs triggers infection by pneumotropic Sendai and influenza A viruses. Purification and characterization. 1082 3

Heparin is a glycosaminoglican used in prophylactic and treatment of thrombosis. Heparin possesses also non-anticoagulant properties, including modulation of various proteases, anticomplement activity, and anti-inflammatory actions. Inhaled heparin has been shown to reduce early phase of asthmatic reaction and suppress allergen induced rise in bronchial hyperreactivity. Heparin inhibits the acute cutaneous reaction due to allergens. Moreover, inhaled heparin prevents exercise-induced asthma. The exact mechanism of heparin action in bronchial asthma remains obscure. It has been observed that heparin acts as a specific blocker of IP3 receptors and inhibits IP3-mediated calcium release in various cell types, including vascular smooth muscle and airway smooth muscle. In this mechanism heparin inhibits allergen induced mast cell degranulation and prevents subsequent development of reaction cascade leading to inflammation, bronchial hyperreactivity and asthma. It also modulates migration of proinflammatory cells, eosinophils and neutrophils, into the site of allergic reaction. Furthermore, heparin inhibits the increased vascular permeability induced by a wide range of agonists acting via specific receptors located on the vascular endothelial cells. The cationic peroxidases, such as major basic protein and eosinophil peroxidase, are neutralized by the highly anionic heparin; thus heparin inhibits the epithelial damage induced by some of these cationic proteins. The mechanism involved in the control of bronchial hyperreactivity by heparin has been studied little and is yet poorly understood. Heparin deserves further investigations in large number of subjects to provide further insight into the pathophysiology of asthma. Heparin may also be of clinical importance and may form the basis of novel therapeutic approaches.
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PMID:[The role of heparin in allergic inflammation]. 1094 59

Heparin is an intracellular product of vertebrate mast cell currently used as exogenous anticoagulant. Despite of the potent biological activities of exogenous heparin, its physiological function has not been clearly established yet. Here, a heparin with similar structure and anticoagulant properties to the mammalian counterpart was shown to occur as the intracellular product of test cells, a cell monolayer that surrounds egg of the invertebrate Styela plicata (Chordata-Tunicata). As in the case of mammalian mast cells, heparin from the ascidian test cells is removed from the intracellular granules after incubation with compound 48/80. Following fertilization, the test cells surrounding the developing larva still retain heparin as metachromatic granulation. In the adult invertebrate, heparin occurs as intracellular granules at the apical tip of epithelial cells surrounding the lumen of both intestine and pharynx, in close contact with the external environment. This is the first description of the presence of heparin in cytoplasmic granules of epithelial-like cells around the lumen of sites exposed to external agents. This arrangement may reflect the participation of heparin in defense mechanisms in this invertebrate.
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PMID:Occurrence of heparin in the invertebrate styela plicata (Tunicata) is restricted to cell layers facing the outside environment. An ancient role in defense? 1095 56

Mast cells are suggested to participate in regenerative processes, but their influence on epithelialization and wound healing has not been well studied. Since mast cells can be found in contact with epidermis in chronic inflammatory skin diseases and venous ulcers, the effect of mast cells on keratinocyte growth was studied. Keratinocytes were cultured in serum-free conditions with (complete medium) or without (basal medium) epidermal growth factor (EGF) and bovine pituitary extract (BPE) to reach subconfluence in a 24-well plate, and the cells were treated with different mast cell mediators histamine, heparin and tryptase, or lysate from HMC-1 cells, a human leukemic mast cell line. Whole skin cultures were used as a model for in vitro wounds to study the effect of mast cells on epithelial outgrowth from skin specimens. Histamine inhibited 3H-thymidine incorporation of keratinocytes dose-dependently by 29% at 1 mM, and 89% at 5 mM histamine. In whole skin culture, histamine inhibited epithelial outgrowth dose-dependently by 64% already at 0.1 mM histamine and maximally (91%) at 1 mM histamine. Heparin inhibited 3H-thymidine incorporation dose-dependently by up to 33% at 2 microg/ml in the absence, but not in the presence, of EGF/BPE. In contrast, in whole skin culture, heparin first inhibited the epithelial outgrowth by up to 27% at 2 microg/ml, but then reversed the inhibition to 30% stimulation at 200 microg/ml. Skin tryptase (0.0285 to 2.85 microg/ml) with or without heparin (0.5 to 20 microg/ml) did not affect thymidine incorporation in keratinocytes. Lysate from HMC-1 cells, but not that from control, neuroblastoma cells, inhibited 3H-thymidine incorporation in keratinocytes dose-dependently, and maximal (47%) inhibition was reached with 16,700 lysed HMC-1 cells/ml. In whole skin culture, HMC-1 lysate inhibited the epithelial outgrowth by up to 36% at 67,000 lysed cells/ml. The results show that mast cells and their mediators are inhibitory to keratinocyte 3H-thymidine incorporation and epithelial outgrowth in vitro, although, the inhibitory effect of histamine was seen at high concentrations suggesting a requirement for close morphologic vicinity of mast cells to keratinocytes. Thus, mast cells are assumed to control epidermal regeneration and to impair epithelialization of chronic ulcers.
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PMID:Inhibition of keratinocyte growth in cell culture and whole skin culture by mast cell mediators. 1138 Jun 14

Heparin-deficient mice, generated by gene targeting of N-deacetylase/N-sulfotransferase-2 (NDST-2), display severe mast cell defects, including an absence of stored mast cell proteases. However, the mechanism behind these observations is not clear. Here we show that NDST-2+/+ bone marrow-derived mast cells cultured in the presence of IL-3 synthesise, in addition to highly sulphated chondroitin sulphate (CS), small amounts of equally highly sulphated heparin-like polysaccharide. The corresponding NDST-2-/- cells produced highly sulphated CS only. Carboxypeptidase A (CPA) activity was detected in NDST+/+ cells but was almost absent in the NDST-/- cells, whereas tryptase (mouse mast cell protease 6; mMCP-6) activity and antigen was detected in both cell types. Antigen for the chymase mMCP-5 was detected in NDST-2+/+ cells but not in the heparin-deficient cells. Northern blot analysis revealed mRNA expression of CPA, mMCP-5 and mMCP-6 in both wild-type and NDST-2-/- cells. A approximately 36 kDa CPA band, corresponding to proteolytically processed active CPA, as well as a approximately 50 kDa pro-CPA band was present in NDST-2+/+ cells. The NDST-2-/- mast cells contained similar levels of pro-CPA as the wild-type mast cells, but the approximately 36 kDa band was totally absent. This indicates that the processing of pro-CPA to its active form may require the presence of heparin and provides the first insight into a mechanism by which the absence of heparin may cause disturbed secretory granule organisation in mast cells.
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PMID:Altered storage of proteases in mast cells from mice lacking heparin: a possible role for heparin in carboxypeptidase A processing. 1210 44

Heparin is a glycosaminoglycan (GAG) that is extracted primarily from porcine intestinal tissues and is widely used as a clinical anticoagulant. It is biosynthesized as a proteoglycan and stored exclusively in mast cells and is partially degraded to peptidoglycan and GAG on immunologically activated mast cell degranulation. In contrast, the structurally related heparan sulfate, is the polysaccharide portion of a ubiquitous proteoglycan, localized on cell surface and in the extracellular matrix of all animal tissues. Heparin and heparan sulfate are made in the Golgi through a similar biosynthetic pathway. The current study was undertaken in a search for alternative, non-mammalian, sources of anticoagulant heparin. The heparin/heparan sulfate family of GAGs, prepared and purified from turkey intestine, were assayed for anticoagulant activity and structurally characterized. The resulting GAGs displayed a very low anticoagulant activity when compared to those obtained from porcine intestine using an identical procedure. Structural characterization studies clearly demonstrate that heparan sulfate is the major GAG in the turkey intestine. This observation is rationalized based on differences in the mammalian and avian coagulation and immune systems.
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PMID:Turkey intestine as a commercial source of heparin? Comparative structural studies of intestinal avian and mammalian glycosaminoglycans. 1252 47


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