Antigen-induced mast cell expansion and bronchoconstriction in a mouse model of asthma

Lung mastocytosis and antigen-induced bronchoconstriction are common features in allergic asthmatics. It is therefore important that animal models of asthma show similar features of mast cell inflammation and reactivity to inhaled allergen. We hypothesized that house dust mite (HDM) would induce mastocytosis in the lung and that inhalation of HDM would trigger bronchoconstriction. Mice were sensitized with intranasal HDM extract, and the acute response to nebulized HDM or the mast cell degranulating compound 48/80 was measured with respiratory input impedance. Using the constant-phase model we calculated Newtonian resistance ( R n) reflecting the conducting airways, tissue dampening ( G), and lung elastance ( H). Bronchoalveolar lavage fluid was analyzed for mouse mast cell protease-1 (mMCP-1). Lung tissue was analyzed for cytokines, histamine, and α-smooth muscle actin (α-SMA), and histological slides were stained for mast cells. HDM significantly increased R n but H and G remained unchanged. HDM significantly expanded mast cells compared with control mice; at the same time mMCP-1, α-SMA, Th2 cytokines, and histamine were significantly increased. Compound 48/80 inhalation caused bronchoconstriction and mMCP-1 elevation similarly to HDM inhalation. Bronchoconstriction was eliminated in mast cell-deficient mice. We found that antigen-induced acute bronchoconstriction has a distinct phenotype in mice. HDM sensitization caused lung mastocytosis, and we conclude that inhalation of HDM caused degranulation of mast cells leading to an acute bronchoconstriction without affecting the lung periphery and that mast cell-derived mediators are responsible for the development of the HDM-induced bronchoconstriction in this model.

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Development of Eosinophilic Airway Inflammation and Airway Hyperresponsiveness in Mast Cell–deficient Mice

Journal of Experimental Medicine ◽

10.1084/jem.186.3.449 ◽

1997 ◽

Vol 186 (3) ◽

pp. 449-454 ◽

Cited By ~ 321

Author(s):

K. Takeda ◽

E. Hamelmann ◽

A. Joetham ◽

L.D. Shultz ◽

G.L. Larsen ◽

...

Keyword(s):

Mast Cell ◽

Mast Cells ◽

Airway Hyperresponsiveness ◽

Cell Activation ◽

Immunoglobulin E ◽

Indirect Evidence ◽

Dynamic Compliance ◽

Lavage Fluid ◽

Mast Cell Activation ◽

Deficient Mice

Mast cells are the main effector cells of immediate hypersensitivity and anaphylaxis. Their role in the development of allergen-induced airway hyperresponsiveness (AHR) is controversial and based on indirect evidence. To address these issues, mast cell–deficient mice (W/W v) and their congenic littermates were sensitized to ovalbumin (OVA) by intraperitoneal injection and subsequently challenged with OVA via the airways. Comparison of OVA-specific immunoglobulin E (IgE) levels in the serum and numbers of eosinophils in bronchoalveolar lavage fluid or lung digests showed no differences between the two groups of mice. Further, measurements of airway resistance and dynamic compliance at baseline and after inhalation of methacholine were similar. These data indicate that mast cells or IgE–mast cell activation is not required for the development of eosinophilic inflammation and AHR in mice sensitized to allergen via the intraperitoneal route and challenged via the airways.

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Biochemical and immunological characterization of multiple glycoforms of mouse mast cell protease 1: comparison with an isolated murine serosal mast cell protease (MMCP-4)

Biochemical Journal ◽

10.1042/bj2940127 ◽

1993 ◽

Vol 294 (1) ◽

pp. 127-135 ◽

Cited By ~ 21

Author(s):

G F J Newlands ◽

D P Knox ◽

S R Pirie-Shepherd ◽

H R P Miller

Keyword(s):

Mast Cell ◽

Amino Acid ◽

Mast Cells ◽

Western Blotting ◽

Trichinella Spiralis ◽

Double Diffusion ◽

Terminal Amino Acid ◽

Mast Cell Protease ◽

Terminal Amino ◽

Mouse Mast Cell

Five highly soluble, chymotrypsin-like, neutral serine proteases, with molecular masses in the range 30-33 kDa, were isolated from Trichinella spiralis-infected mouse small intestine. These enzymes were closely related antigenically on Western blotting and by Ouchterlony double diffusion using a polyclonal, cross-absorbed, sheep antibody raised against mouse mast cell protease-1 (MMCP-1) and on the basis of N-terminal amino acid sequence analysis, were identified as variant forms of MMCP-1. Substrate and inhibitor analysis confirmed that the five variants (MMCP-1 A-E) had similar characteristics, although highly significant (P = 0.025 to P < 0.0001) variations in Km and kcat, were detected. Against human alpha 1-proteinase inhibitor the Ki for MMCP-1C (45 pM) was significantly (P < 0.0001) greater than those for the other proteases (0.76-2.2 pM). The differences in electrophoretic mobility are probably a result of variable glycosylation, since removal of N-linked carbohydrate produced a polypeptide of approx. 28 kDa in each case which was, like the native enzyme, immunoreactive on Western blotting. A much less soluble 28 kDa enzyme was isolated from serosal mast cells and identified as MMCP-4 by N-terminal amino acid sequencing. Like MMCP-1 it has chymotrypsin-like substrate specificities with activity at neutral pH. However, it was antigenically distinct from MMCP-1 and, using sheep anti-MMCP-1, was not detected on Western blotting or by Ouchterlony double diffusion, e.l.i.s.a. or immunohistochemistry. This last technique established that the MMCP-1 variants were uniquely present in enteric mast cells, thereby providing a highly selective means of distinguishing the mucosal and connective tissue mast cell subsets in the mouse.

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The emerging role of mast cell proteases in asthma

European Respiratory Journal ◽

10.1183/13993003.00685-2019 ◽

2019 ◽

Vol 54 (4) ◽

pp. 1900685 ◽

Cited By ~ 4

Author(s):

Gunnar Pejler

Keyword(s):

Mast Cell ◽

Mast Cells ◽

Current Knowledge ◽

Secretory Granules ◽

Cross Linking ◽

Lysosomal Hydrolases ◽

Ige Receptor ◽

Deficient Mice ◽

Lines Of Evidence

It is now well established that mast cells (MCs) play a crucial role in asthma. This is supported by multiple lines of evidence, including both clinical studies and studies on MC-deficient mice. However, there is still only limited knowledge of the exact effector mechanism(s) by which MCs influence asthma pathology. MCs contain large amounts of secretory granules, which are filled with a variety of bioactive compounds including histamine, cytokines, lysosomal hydrolases, serglycin proteoglycans and a number of MC-restricted proteases. When MCs are activated, e.g. in response to IgE receptor cross-linking, the contents of their granules are released to the exterior and can cause a massive inflammatory reaction. The MC-restricted proteases include tryptases, chymases and carboxypeptidase A3, and these are expressed and stored at remarkably high levels. There is now emerging evidence supporting a prominent role of these enzymes in the pathology of asthma. Interestingly, however, the role of the MC-restricted proteases is multifaceted, encompassing both protective and detrimental activities. Here, the current knowledge of how the MC-restricted proteases impact on asthma is reviewed.

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Mouse connective tissue mast cell proteases tryptase and carboxypeptidase A3 play protective roles in itch induced by endothelin-1

Journal of Neuroinflammation ◽

10.1186/s12974-020-01795-4 ◽

2020 ◽

Vol 17 (1) ◽

Author(s):

Elín I. Magnúsdóttir ◽

Mirjana Grujic ◽

Jessica Bergman ◽

Gunnar Pejler ◽

Malin C. Lagerström

Keyword(s):

Mast Cell ◽

Mast Cells ◽

Connective Tissue ◽

Treatment Options ◽

Cell Types ◽

Tissue Type ◽

Human Mast Cell ◽

Knock Out ◽

Endothelin 1 ◽

Deficient Mice

Abstract Background Itch is an unpleasant sensation that can be debilitating, especially if it is chronic and of non-histaminergic origin, as treatment options are limited. Endothelin-1 (ET-1) is a potent endogenous vasoconstrictor that also has the ability to induce a burning, non-histaminergic pruritus when exogenously administered, by activating the endothelin A receptor (ETAR) on primary afferents. ET-1 is released endogenously by several cell-types found in the skin, including macrophages and keratinocytes. Mast cells express ETARs and can thereby be degranulated by ET-1, and mast cell proteases chymase and carboxypeptidase A3 (CPA3) are known to either generate or degrade ET-1, respectively, suggesting a role for mast cell proteases in the regulation of ET-1-induced itch. The mouse mast cell proteases (mMCPs) mMCP4 (chymase), mMCP6 (tryptase), and CPA3 are found in connective tissue type mast cells and are the closest functional homologs to human mast cell proteases, but little is known about their role in endothelin-induced itch. Methods In this study, we evaluated the effects of mast cell protease deficiency on scratching behavior induced by ET-1. To investigate this, mMCP knock-out and transgenic mice were injected intradermally with ET-1 and their scratching behavior was recorded and analyzed. Results CPA3-deficient mice and mice lacking all three proteases demonstrated highly elevated levels of scratching behavior compared with wild-type controls. A modest increase in the number of scratching bouts was also seen in mMCP6-deficient mice, while mMCP4-deficiency did not have any effect. Conclusion Altogether, these findings identify a prominent role for the mast cell proteases, in particular CPA3, in the protection against itch induced by ET-1.

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Low c-kit expression of cultured mast cells of mi/mi genotype may be involved in their defective responses to fibroblasts that express the ligand for c-kit

Blood ◽

10.1182/blood.v80.6.1454.1454 ◽

1992 ◽

Vol 80 (6) ◽

pp. 1454-1462 ◽

Cited By ~ 6

Author(s):

Y Ebi ◽

Y Kanakura ◽

T Jippo-Kanemoto ◽

T Tsujimura ◽

T Furitsu ◽

...

Keyword(s):

Mast Cell ◽

Mast Cells ◽

Tyrosine Phosphorylation ◽

Messenger Rna ◽

Tissue Type ◽

Phenotypic Change ◽

3T3 Fibroblasts ◽

Mouse Mast Cell ◽

And Function

Abstract Mutant mice of mi/mi genotype are osteopetrotic and deficient in tissue mast cells due to a defect in osteoclasts and mast cells. In an effort to further understand the mechanisms behind why mi/mi mouse-derived cultured mast cells (mi/mi-CMC) responded to interleukin-3 (IL-3), but not to the proliferative stimuli presented by fibroblasts, mi/mi-CMC and congenic normal (+/+) mouse-derived CMC (+/+-CMC), both of which expressed the phenotypic characteristics of immature mast cells, were cocultured with Swiss albino/3T3 fibroblasts in a medium containing IL- 3. In the in vitro CMC/fibroblast coculture, mi/mi-CMC did not acquire the phenotypes of connective tissue-type mast cells (CTMC), while +/+- CMC did. In addition, attachment of mi/mi-CMC to the fibroblasts was found to be significantly lower than that of +/+-CMC. Because the interaction of c-kit product with its ligand (stem cell factor [SCF]) is known to play an important role not only in proliferation and differentiation of mast cells but also in attachment of CMC to fibroblasts, the expression and function of c-kit were investigated in mi/mi-CMC and +/+-CMC. Recombinant rat SCF (rrSCF164) induced a dose- dependent proliferation of +/+-CMC. Also, rrSCF164 induced +/+-CMC to acquire the phenotypes of CTMC in the medium containing IL-3. By contrast, rrSCF164 did not stimulate the proliferation of mi/mi-CMC nor induce a phenotypic change of the cells from immature mast cells to mature, CTMC-like mast cells. Immunoblotting with antiphosphotyrosine antibody showed that rrSCF164 induced considerable tyrosine phosphorylation of 145- to 165-Kd protein, the product of c-kit, in +/+- CMC, whereas tyrosine phosphorylation of the protein was barely detectable in mi/mi-CMC. Northern blot and flow cytometry analyses showed that mi/mi-CMC expressed much less c-kit at both protein and message levels than +/+-CMC. Further, mi/mi-CMC were found to differ from +/+-CMC in the expression of mouse mast cell protease-6 (MMCP-6) and MMCP-2 messenger RNA transcripts. These results suggest that the gene product of the mi locus may be important in regulating the expression of gene products such as c-kit, and that mast cell deficiency of mi/mi mice appears to be due, at least in part, to impaired signaling through the c-kit receptor because of the low c-kit expression.

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Evidence for the interaction of mast cell-degranulating peptide with pertussis toxin-sensitive G proteins in mast cells

European Journal of Pharmacology Molecular Pharmacology ◽

10.1016/0922-4106(91)90037-i ◽

1991 ◽

Vol 207 (3) ◽

pp. 249-255 ◽

Cited By ~ 19

Author(s):

Mouafak Mousli ◽

Christian Bronner ◽

Jean-Luc Bueb ◽

Yves Landry

Keyword(s):

Mast Cell ◽

Mast Cells ◽

G Proteins ◽

Pertussis Toxin ◽

Mast Cell Degranulating

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IgE Enhances Mouse Mast Cell FcεRI Expression In Vitro and In Vivo: Evidence for a Novel Amplification Mechanism in IgE-dependent Reactions

Journal of Experimental Medicine ◽

10.1084/jem.185.4.663 ◽

1997 ◽

Vol 185 (4) ◽

pp. 663-672 ◽

Cited By ~ 295

Author(s):

Masao Yamaguchi ◽

Chris S. Lantz ◽

Hans C. Oettgen ◽

Ildy M. Katona ◽

Tony Fleming ◽

...

Keyword(s):

Mast Cell ◽

Mast Cells ◽

Cell Activation ◽

Immunoglobulin E ◽

Specific Antigen ◽

Mast Cell Activation ◽

Proinflammatory Mediators ◽

Mouse Mast Cell

The binding of immunoglobulin E (IgE) to high affinity IgE receptors (FcεRI) expressed on the surface of mast cells primes these cells to secrete, upon subsequent exposure to specific antigen, a panel of proinflammatory mediators, which includes cytokines that can also have immunoregulatory activities. This IgE- and antigen-specific mast cell activation and mediator production is thought to be critical to the pathogenesis of allergic disorders, such as anaphylaxis and asthma, and also contributes to host defense against parasites. We now report that exposure to IgE results in a striking (up to 32-fold) upregulation of surface expression of FcεRI on mouse mast cells in vitro or in vivo. Moreover, baseline levels of FcεRI expression on peritoneal mast cells from genetically IgE-deficient (IgE −/−) mice are dramatically reduced (by ∼83%) compared with those on cells from the corresponding normal mice. In vitro studies indicate that the IgE-dependent upregulation of mouse mast cell FcεRI expression has two components: an early cycloheximide-insensitive phase, followed by a later and more sustained component that is highly sensitive to inhibition by cycloheximide. In turn, IgE-dependent upregulation of FcεRI expression significantly enhances the ability of mouse mast cells to release serotonin, interleukin-6 (IL-6), and IL-4 in response to challenge with IgE and specific antigen. The demonstration that IgE-dependent enhancement of mast cell FcεRI expression permits mast cells to respond to antigen challenge with increased production of proinflammatory and immunoregulatory mediators provides new insights into both the pathogenesis of allergic diseases and the regulation of protective host responses to parasites.

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Fate of two mast cell tryptases in V3 mastocytosis and normal BALB/c mice undergoing passive systemic anaphylaxis: prolonged retention of exocytosed mMCP-6 in connective tissues, and rapid accumulation of enzymatically active mMCP-7 in the blood.

Journal of Experimental Medicine ◽

10.1084/jem.184.3.1061 ◽

1996 ◽

Vol 184 (3) ◽

pp. 1061-1073 ◽

Cited By ~ 58

Author(s):

N Ghildyal ◽

D S Friend ◽

R L Stevens ◽

K F Austen ◽

C Huang ◽

...

Keyword(s):

Extracellular Matrix ◽

Mast Cell ◽

Mast Cells ◽

Chromosome 17 ◽

Macromolecular Complex ◽

Connective Tissues ◽

Systemic Anaphylaxis ◽

Rich Domain ◽

Passive Systemic Anaphylaxis ◽

Mouse Mast Cell

The mouse mast cell protease granule tryptases designated mMCP-6 and mMCP-7 are encoded by highly homologous genes that reside on chromosome 17. Because these proteases are released when mast cells are activated, we sought a basis for distinctive functions by examining their fates in mice undergoing passive systemic anaphylaxis. 10 min-1 h after antigen (Ag) was administered to immunoglobulin (Ig)E-sensitized mice, numerous protease/proteoglycan macromolecular complexes appeared in the extracellular matrix adjacent to most tongue and heart mast cells of normal BALB/c mice and most spleen and liver mast cells of V3 mastocytosis mice. These complexes could be intensively stained by anti-mMCP-6 Ig but not by anti-mMCP-7 Ig. Shortly after Ag challenge of V3 mastocytosis mice, large amounts of properly folded, enzymatically active mMCP-7 were detected in the plasma. This plasma-localized tryptase was approximately 150 kD in its multimeric state and approximately 32 kD in its monomeric state, possessed an NH2 terminus identical to that of mature mMCP-7, and was not covalently bound to any protease inhibitor. Comparative protein modeling and electrostatic calculations disclosed that mMCP-6 contains a prominent Lys/Arg-rich domain on its surface, distant from the active site. The absence of this domain in mMCP-7 provides an explanation for its selective dissociation from the exocytosed macromolecular complex. The retention of exocytosed mMCP-6 in the extracellular matrix around activated tissue mast cells suggests a local action. In contrast, the rapid dissipation of mMCP-7 from granule cores and its inability to be inactivated by circulating protease inhibitors suggests that this tryptase cleaves proteins located at more distal sites.

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