Propofol has been shown to against intestinal reperfusion injury when treated either before or after ischemia, during which mast cell could be activated. The aim of this study was to evaluate the role of propofol in restoring the intestinal epithelial cells integrity disrupted by mast cell activation or the released tryptase after activation in vitro. We investigated the effect of: (1) tryptase on Caco-2 monolayers in the presence of PAR-2 inhibitor or propofol, (2) mast cell degranulation in a Caco-2/LAD-2 co-culture model in the presence of propofol, and (3) propofol on mast cell degranulation. Epithelial integrity was detected using transepithelial resistance (TER) and permeability to fluorescein isothiocyanate (FITC)-dextran (the apparent permeability coefficient, Papp). The expression of junctional proteins zonula occludens-1 (ZO-1/TJP1) and occludin were determined using western blot analysis and immunofluorescence microscopy. The intracellular levels of reactive oxidative species (ROS) and Ca2+ were measured using flow cytometry. Tryptase directly enhanced intestinal barrier permeability as demonstrated by significant reductions in TER, ZO-1, and occludin protein expression and concomitant increases in Papp. The intestinal barrier integrity was restored by PAR-2 inhibitor but not by propofol. Meanwhile, mast cell degranulation resulted in epithelial integrity disruption in the Caco-2/LAD-2 co-culture model, which was dramatically attenuated by propofol. Mast cell degranulation caused significant increases in intracellular ROS and Ca(2+) levels, which were blocked by propofol and NAC. Propofol pretreatment can inhibit mast cell activation via ROS/Ca(2+) and restore the intestinal barrier integrity induced by mast cell activation, instead of by tryptase.

• MeSH
• Caco-2 buňky MeSH
• degranulace buněk MeSH
• epitelové buňky metabolismus   MeSH
• lidé MeSH
• mastocyty metabolismus   MeSH
• okludin metabolismus   MeSH
• permeabilita MeSH
• propofol * farmakologie   MeSH
• reaktivní formy kyslíku metabolismus   MeSH
• střevní sliznice metabolismus   MeSH
• tryptasy metabolismus   farmakologie   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH

In the current classification of the World Health Organization (WHO), bone marrow mastocytosis (BMM) is a provisional variant of indolent systemic mastocytosis (ISM) defined by bone marrow involvement and absence of skin lesions. However, no additional diagnostic criteria for BMM have been proposed. Within the registry dataset of the European Competence Network on Mastocytosis, we compared characteristics and outcomes of 390 patients with BMM and 1175 patients with typical ISM. BMM patients were significantly older, predominantly male, had lower tryptase and lower burden of neoplastic mast cells, and displayed a higher frequency of allergic reactions, mainly triggered by Hymenoptera, than patients with typical ISM. The estimated 10-year progression-free survival of BMM and typical ISM was 95.9% and 92.6%, respectively. In BMM patients defined by WHO-based criteria, the presence of one B-Finding and tryptase level ≥125 ng/mL were identified as risk factors for progression in multivariate analyses. BMM patients without any of these risk factors were found to have better progression-free survival (p < 0.05) and better overall survival (p < 0.05) than other ISM patients. These data support the proposal to define BMM as a separate SM variant characterized by SM criteria, absence of skin lesions, absence of B-Findings, and tryptase levels <125 ng/mL.

• MeSH
• dospělí MeSH
• kostní dřeň metabolismus   patologie   MeSH
• kožní nemoci patofyziologie   MeSH
• lidé středního věku MeSH
• lidé MeSH
• mastocytóza diagnóza   epidemiologie   metabolismus   MeSH
• mastocyty metabolismus   patologie   MeSH
• míra přežití MeSH
• následné studie MeSH
• prognóza MeSH
• senioři nad 80 let MeSH
• senioři MeSH
• systémová mastocytóza diagnóza   epidemiologie   metabolismus   MeSH
• tryptasy metabolismus   MeSH
• Check Tag
• dospělí MeSH
• lidé středního věku MeSH
• lidé MeSH
• mužské pohlaví MeSH
• senioři nad 80 let MeSH
• senioři MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Geografické názvy
• Evropa MeSH

• MeSH
• aktivace komplementu * imunologie   MeSH
• anafylaxe * imunologie   metabolismus   MeSH
• bazofily imunologie   MeSH
• biologické markery * metabolismus   MeSH
• bradykinin imunologie   metabolismus   MeSH
• chymasy imunologie   metabolismus   MeSH
• cytokiny * imunologie   metabolismus   MeSH
• faktor aktivující destičky imunologie   metabolismus   MeSH
• fenotyp * MeSH
• histamin imunologie   metabolismus   MeSH
• imunoglobulin E imunologie   MeSH
• imunoglobulin G imunologie   MeSH
• individualizovaná medicína MeSH
• interleukin-6 imunologie   metabolismus   MeSH
• karboxypeptidasy A imunologie   metabolismus   MeSH
• lidé MeSH
• mastocyty imunologie   MeSH
• tryptasy imunologie   metabolismus   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• přehledy MeSH

BACKGROUND: A salivary proteome-transcriptome project on the hard tick Ixodes scapularis revealed that Kunitz peptides are the most abundant salivary proteins. Ticks use Kunitz peptides (among other salivary proteins) to combat host defense mechanisms and to obtain a blood meal. Most of these Kunitz peptides, however, remain functionally uncharacterized, thus limiting our knowledge about their biochemical interactions. RESULTS: We discovered an unusual cysteine motif in a Kunitz peptide. This peptide inhibits several serine proteases with high affinity and was named tryptogalinin due to its high affinity for β-tryptase. Compared with other functionally described peptides from the Acari subclass, we showed that tryptogalinin is phylogenetically related to a Kunitz peptide from Rhipicephalus appendiculatus, also reported to have a high affinity for β-tryptase. Using homology-based modeling (and other protein prediction programs) we were able to model and explain the multifaceted function of tryptogalinin. The N-terminus of the modeled tryptogalinin is detached from the rest of the peptide and exhibits intrinsic disorder allowing an increased flexibility for its high affinity with its inhibiting partners (i.e., serine proteases). CONCLUSIONS: By incorporating experimental and computational methods our data not only describes the function of a Kunitz peptide from Ixodes scapularis, but also allows us to hypothesize about the molecular basis of this function at the atomic level.

• MeSH
• aminokyselinové motivy MeSH
• cystein chemie   genetika   MeSH
• fylogeneze MeSH
• inhibitory serinových proteinas chemie   klasifikace   genetika   metabolismus   MeSH
• klíště chemie   genetika   metabolismus   MeSH
• lidé MeSH
• molekulární modely MeSH
• molekulární sekvence - údaje MeSH
• proteiny členovců chemie   klasifikace   genetika   metabolismus   MeSH
• rekombinantní proteiny chemie   klasifikace   genetika   metabolismus   MeSH
• Rhipicephalus chemie   genetika   metabolismus   MeSH
• sekvenční homologie aminokyselin MeSH
• slinné proteiny a peptidy chemie   klasifikace   genetika   metabolismus   MeSH
• terciární struktura proteinů MeSH
• tryptasy antagonisté a inhibitory   chemie   metabolismus   MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

Matrix metalloproteinases (MMPs) is a family of proteolytic enzymes involved in remodeling of extracellular matrix. Although proteolytic enzymes are produced by many cell types, mast cells seem to be more important than other types in remodeling of pulmonary arteries during hypoxia. Therefore, we tested in vitro production of MMPs and serine proteases in four cell types (mast cells, fibroblasts, vascular smooth muscle cells and endothelial cells) cultivated for 48 h under normoxic or hypoxic (3 % O2) conditions. MMP-13 was visualized by immunohistochemistry, MMP-2 and MMP-9 were detected by zymography in cell lysates. Enzymatic activities (MMPs, tryptase and chymase) were estimated in the cultivation media. Hypoxia had a minimal effect on total MMP activity in the cultivation media of all types of cells, but immunofluorescence revealed higher intensity of MMP-13 in the cells exposed to hypoxia except of fibroblasts. Tryptase activity was three times higher and chymase activity twice higher in mast cells cultivated in hypoxia than in those cultured in normoxia. Among all cell types studied here, mast cells are the most abundant source of proteolytic enzymes under normoxic and hypoxic conditions. Moreover, in these cells hypoxia increases the production of both specific serine proteases tryptase and chymase, which can act as MMPs activators.

• MeSH
• arteria pulmonalis cytologie   MeSH
• buněčné linie MeSH
• chymasy metabolismus   MeSH
• endoteliální buňky cytologie   enzymologie   MeSH
• fibroblasty cytologie   enzymologie   MeSH
• financování organizované MeSH
• hypoxie metabolismus   MeSH
• krysa rodu rattus MeSH
• kyslík farmakologie   MeSH
• mastocytom MeSH
• mastocyty cytologie   enzymologie   MeSH
• matrixová metaloproteinasa 13 metabolismus   MeSH
• matrixová metaloproteinasa 2 metabolismus   MeSH
• matrixová metaloproteinasa 9 metabolismus   MeSH
• myocyty hladké svaloviny cytologie   enzymologie   MeSH
• myši MeSH
• potkani Wistar MeSH
• proteasy metabolismus   MeSH
• skot MeSH
• svaly hladké cévní cytologie   MeSH
• tryptasy metabolismus   MeSH
• zvířata MeSH
• Check Tag
• krysa rodu rattus MeSH
• mužské pohlaví MeSH
• myši MeSH
• skot MeSH
• zvířata MeSH

haracterized by fibrotization and muscularization of the walls of peripheral pulmonary arteries. This vessel remodeling is accompanied by an increase in the amount of lung mast cells (LMC) and the presence of small collagen cleavage products in the vessel walls. We hypothesize that hypoxia activates LMC, which release matrix metalloproteinases (MMPs) cleaving collagen and starting increased turnover of connective tissue proteins. This study was designed to determine whether in vitro hypoxia stimulates production of MMPs in rat LMC and increases their collagenolytic activity. The LMC were separated on the Percoll gradient and then were divided into two groups and cultivated for 24 h in 21 % O2+ 5 % CO2 or in 10 % O2 + 5 % CO2. Presence of the rat interstitial tissue collagenase (MMP-13) in LMC was visualized by immunohistological staining and confirmed by Western blot analysis. Total MMPs activity and tryptase activity were measured in both cultivation media and cellular extracts. Exposure to hypoxia in vitro increased the amount of cells positively labeled by anti-MMP-13 antibody as well as activities of all measured enzymes. The results therefore support the concept that LMC are an important source of increased collagenolytic activity in chronic hypoxia.

• Klíčová slova
• Hypoxia in vitro, Rat lung mast cells, Tissue metalloproteinases, Tryptase, Remodeling of pulmonary vessels,
• MeSH
• financování organizované MeSH
• hypoxie buňky MeSH
• imunohistochemie MeSH
• kolagen metabolismus   MeSH
• krysa rodu rattus MeSH
• kultivační média metabolismus   MeSH
• kultivované buňky MeSH
• mastocyty enzymologie   MeSH
• matrixová metaloproteinasa 13 metabolismus   MeSH
• matrixová metaloproteinasa 2 metabolismus   MeSH
• matrixová metaloproteinasa 9 metabolismus   MeSH
• matrixové metaloproteinasy metabolismus   MeSH
• plíce cytologie   enzymologie   MeSH
• potkani Wistar MeSH
• separace buněk MeSH
• tryptasy metabolismus   MeSH
• upregulace MeSH
• western blotting MeSH
• zvířata MeSH
• Check Tag
• krysa rodu rattus MeSH
• mužské pohlaví MeSH
• zvířata MeSH