Mast cell heterogeneity

Increasing evidence for the existence of inter- and intra-species mast cell heterogeneity has expanded the potential biological role of this cell. Early studies suggesting that mast cells at mucosal sites differ morphologically and histochemically from connective tissue mast cells have been confirmed using isolated intestinal mucosal mast cells in the rat and more recently in man. These studies also established that mucosal mast cells are functionally distinct from connective tissue mast cells. Thus, mucosal and connective tissue mast cells differ in their responsiveness to a variety of mast cell secretagogues and antiallergic agents. Speculation about the therapeutic use of antiallergic drugs in disorders involving intestinal mast cells cannot, therefore, be based on extrapolation from studies of their effects on mast cells from other sites. Regulatory mechanisms for mast cell secretion may also be heterogeneous since mucosal mast cells differ from connective tissue mast cells in their response to a variety of physiologically occurring regulatory peptides. The development of techniques to purify isolated mast cell subpopulations will facilitate future analysis of the biochemical basis of the functional heterogeneity of mast cells.     

Mast cell activation and tissue cell proliferation

Summary The effect of mast cell activation and degranulation on the proliferation in the intact mesentery was studied in Sprague-Dawley rats. Mast cell activation was achieved by a single intraperitoneal injection of Compound 48/80.The proliferation was studied using three independent methods for estimation of cell production and DNA synthesis: 1. the mitotic index, 2. the relative number of cells having a DNA content in the S and G2 regions, by Feulgen photometric measurement in individual cells, and 3. the specific DNA activity, employing a method which combines a liquid scintillation technique after an intravenous injection of ³H-thymidine and Feulgen photometric determination of the DNA content per membrane preparation.It was found that the proliferation of the normal mesenchymal cells adjacent to the activated and degranulated mast cells in the mesentery was significantly increased within 24 and 32 h, the maximum increase being more than 20-fold compared to untreated controls. The results suggest that the common type of mast cell may have a pathophysiological function related to stimulation of local cell proliferation.Supported by grants from the Swedish Medical Research Council (Project 12X-2235) and from the Medical Faculty, University of LinköpingWe thank Brita Söderlund, Margareta Odenö and Iréne Svensson for skilful technical assistance, and Erik Leander, Ph. D., for help with statistical methodsPart of this work was presented at the 7th Meeting of the European Study Group for Cell Proliferation, 5–9 May 1975, in Amsterdam, The Netherlands     

Mast cell activation and autism

Autism spectrum disorders (ASD) are neurodevelopmental disorders characterized by varying degrees of dysfunctional communication and social interactions, repetitive and stereotypic behaviors, as well as learning and sensory deficits. Despite the impressive rise in the prevalence of autism during the last two decades, there are few if any clues for its pathogenesis, early detection or treatment. Increasing evidence indicates high brain expression of pro-inflammatory cytokines and the presence of circulating antibodies against brain proteins. A number of papers, mostly based on parental reporting on their children's health problems, suggest that ASD children may present with “allergic-like” problems in the absence of elevated serum IgE and chronic urticaria. These findings suggest non-allergic mast cell activation, probably in response to environmental and stress triggers that could contribute to inflammation. In utero inflammation can lead to preterm labor and has itself been strongly associated with adverse neurodevelopmental outcomes. Premature babies have about four times higher risk of developing ASD and are also more vulnerable to infections, while delayed development of their gut-blood-brain barriers makes exposure to potential neurotoxins likely. Perinatal mast cell activation by infectious, stress-related, environmental or allergic triggers can lead to release of pro-inflammatory and neurotoxic molecules, thus contributing to brain inflammation and ASD pathogenesis, at least in a subgroup of ASD patients. This article is part of a Special Issue entitled: Mast cells in inflammation.     

Mast cell tryptase and asthma

Recent physiological and pharmacological studies have indicated the potential importance of tryptase, the major protein component in mast cells, in inflammatory diseases (especially asthma). Being released at inflammatory sites after the activation of mast cells, tryptase is capable of causing bronchohyperresponsiveness and infiltration of eosinophils, neutrophils, etc. in animal airways. The mechanisms by which tryptase causes bronchoconstriction involve probably the potentiation of other chemical mediators such as histamine, production of bradykinin via the hydrolysis of kininogen, and cleavage of the bronchodilating peptides VIP (vasoactive intestinal peptide) and PHM (peptide histidine-methionine). Tryptase has also been found to be a potent mitogen in vitro for airway smooth muscle cells and epithelial cells, implying its role in the hyperplasia of the asthmatic airways. The experimental data providing evidence for the above roles of tryptase are summarized in the present review, as well as the effects of tryptase inhibition in animal asthma models. The potential strategies for the development of anti-asthmatic agents based on the inhibition of tryptase are discussed.     

Regulation of cell volume by glycosaminoglycans

Cell volume is regulated by a delicate balance between ion distribution across the plasma membrane and the osmotic properties of intra‐ and extracellular components. Using a fluorescent calcein indicator, we analysed the effects of glycosaminoglycans on the cell volume of hyaluronan producing fibroblasts and hyaluronan deficient HEK cells over a time period of 30 h. Exogenous glycosaminoglycans induced cell blebbing after 2 min and swelling of fibroblasts to about 110% of untreated cell volume at low concentrations which decreased at higher concentrations. HEK cells did not show cell blebbing and responded by shrinking to 65% of untreated cell volume. Heparin induced swelling of both fibroblasts and HEK cells. Hyaluronidase treatment or inhibition of hyaluronan export led to cell shrinkage indicating that the hyaluronan coat maintained fibroblasts in a swollen state. These observations were explained by the combined action of the Donnan effect and molecular crowding. J. Cell. Biochem. 113: 340–348, 2012. © 2011 Wiley Periodicals, Inc.     

Mast cell responses to helminth infection

Mast cells have been suggested to be major effector cells in the immune response to infection with helminths. It is now clear, however, that mast cells are heterogeneous and have a diversity of important functions. In this review, Timothy Lee, Mark Swieter and Dean Befus point out that much of the confusion about the role of mast cells in immunity stems from methods and interpretations which are inadequate for the diversity of roles played by these cells in host responses to parasites. Classical histochemistry may fail to reveal active mast cells, and studies using chemical antagonists are difficult to interpret until we know more about the action of the drugs. The authors show that current research is extending our knowledge of mast cell heterogeneity, and helping to define the powerful array of mediators that they can use to orchestrate the immune response to helminth infections.     

Mast cell heterogeneity in normal man

Thermographic analysis of the skin of the forearm of normal men submitted to nasal instillation of 1 ml of a solution of 48/80, 1 X 10(-2), demonstrates that the skin vessels undergo local vasodilatation. Erythema and wheals sometimes appear, due to the stimulation of the dermal mast cells. Mast cells of the nasal mucosa are never stimulated by such instillation. The differences between dermal and nasal mast cell reactions are in accordance with the concept of mast cell heterogeneity.     

Effect of glycosaminoglycans on fibronectin-mediated cell attachment

In this study, the role of glycosaminoglycans in fibronectin-mediated cell attachment to collagen has been investigated. While it has been established that fibronectin possesses binding sites for several glycosaminoglycans, it was found that only dextran sulfate and macromolecular heparin could decrease the initial rate of cell attachment to collagen. Low molecular weight heparin was inactive. This study suggests that the glycosaminoglycan binding site of fibronectin plays a role in the mechanism of cell attachment.     

Mast cell transcription factors--regulators of cell fate and phenotype

The direct interaction of the antibiotic primycin with the plasma membrane was investigated by employing the well-characterized ergosterol-producing, amphotericin B-sensitive parental Candida albicans strain 33erg(+) and its ergosterol-less amphotericin B-resistant plasma membrane mutant erg-2. The growth inhibition concentration in shaken liquid medium was 64 μgml(-1) for 33erg(+) and 128 μgml(-1) for erg-2, suggesting that the plasma membrane composition influences the mode of action of primycin. To determine the primycin-induced changes in the plasma membrane dynamic, electron paramagnetic resonance (EPR) spectroscopy methods were used, the spin-labeled fatty acid 5-(4,4-dimethyloxazolidine-N-oxyl)stearic acid) being applied for the in vivo measurements. The phase transition temperatures of untreated strain 33erg(+) and its mutant erg-2 were 12.5°C and 11°C, respectively. After 128 μgml(-1) primycin treatment, these values increased to 17.5°C and 16°C, revealing a significant reduction in the phospholipid flexibility. Saturation transfer EPR measurements demonstrated that, the rotational correlation times of the spin label molecule for the control samples of 33erg(+) and erg-2 were 60 ns and 100 ns. These correlation times gradually decreased on the addition of increasing primycin concentrations, reaching 8 μs and 1 μs. The results indicate the plasma membrane "rigidizing" effect of primycin, a feature that may stem from its ability to undergo complex formation with membrane constituent fatty acid molecules, causing alterations in the structures of phospholipids in the hydrophobic surface near the fatty acid chain region.     

Mast cell-associated TNF promotes dendritic cell migration

Mast cells represent a potential source of TNF, a mediator which can enhance dendritic cell (DC) migration. Although the importance of mast cell-associated TNF in regulating DC migration in vivo is not clear, mast cells and mast cell-derived TNF can contribute to the expression of certain models of contact hypersensitivity (CHS). We found that CHS to FITC was significantly impaired in mast cell-deficient Kit(W-sh/W-sh) or TNF(-/)(-) mice. The reduced expression of CHS in Kit(W-sh/W-sh) mice was fully repaired by local transfer of wild-type bone marrow-derived cultured mast cells (BMCMCs), but was only partially repaired by transfer of TNF(-/)(-) BMCMCs. Thus, mast cells, and mast cell-derived TNF, were required for optimal expression of CHS to FITC. We found that the migration of FITC-bearing skin DCs into draining lymph nodes (LNs) 24 h after epicutaneous administration of FITC in naive mice was significantly reduced in mast cell-deficient or TNF(-/)(-) mice, but levels of DC migration in these mutant mice increased to greater than wild-type levels by 48 h after FITC sensitization. Mast cell-deficient or TNF(-/)(-) mice also exhibited significantly reduced migration of airway DCs to local LNs at 24 h after intranasal challenge with FITC-OVA. Migration of FITC-bearing DCs to LNs draining the skin or airways 24 h after sensitization was repaired in Kit(W-sh/W-sh) mice which had been engrafted with wild-type but not TNF(-/)(-) BMCMCs. Our findings indicate that mast cell-associated TNF can contribute significantly to the initial stages of FITC-induced migration of cutaneous or airway DCs.     

Mast cell heterogeneity in the human uvea

To identify chymase- and tryptase-positive mast cells in the human uvea, and to study their associations with different types of resident uveal cells, uveal specimens from 24 human donor eyes were cryosectioned in sagittal and tangential planes. Enzyme histochemical staining of chymase was combined with immunohistochemical staining for tryptase, detected with the APAAP method. Fluorescence immunohistochemistry was performed with antibodies against c-kit, alpha smooth muscle actin, protein gene product (PGP) 9.5, CD45, and HLA-DR. In different uveal compartments, the total amounts of mast cells were calculated and the distributions of chymase and tryptase were quantified. All uveal mast cells were c-kit and CD45 positive and HLA-DR negative. No association existed between mast cells and actin-containing cells. Only a few mast cells were in close association with PGP 9.5-labeled nerve fibers. In the choroid, most mast cells were located in the inner central part (mean density = 48.9/mm²), and contained both chymase and tryptase (96%). The ciliary muscle contained numerous mast cells (mean density = 33.7/mm²), many of them tryptase positive but chymase negative (63%). In the pars plana, a high number of chymase-positive, tryptase-negative mast cells were found (20%). In the iris only a few mast cells were present. Although the choroid contains the most common subtype of mast cells, a unique situation concerning the distribution of chymase and tryptase is present in the anterior uveal tissues. A possible role for these cells in the special immunological situation of the anterior eye chamber merits further investigation. Accepted: 16 September 1999     

Mast cell chymase regulates dermal mast cell number in mice

Chymase inhibitor reduced the increase in the number of dermal mast cells in 2,4-dinitrofluorobenzene-induced dermatitis in a dose-dependent manner. Intradermal injection of human chymase to mouse ear significantly increased histamine content, the marker for mast cell number in the skin. These results suggest that chymase released by mast cells may participate in local mast cell accumulation in a positive feedback fashion. Immunohistochemical analysis revealed that the intradermal injection of chymase reduces expression of stem cell factor (SCF) on surface of the skin keratinocytes. In addition, incubation of human keratinocytes with chymase in vitro resulted in release of SCF into the culture medium. Since soluble SCF is thought to regulate mast cell number, the chymase-induced mast cell accumulation may occur via the ability of chymase to process membrane-bound SCF on the epidermal keratinocytes.     

Mast cell population in rats with experimental atherosclerosis

Mast cell population was studied in rats with experimental atherosclerosis. It has been established that animals kept for 8 months on atherogenic diet revealed marked changes in mast cell population. Predominance of light cells and cell defects were noted. Heparin saturation index was reduced (0.35), as compared to the control (3.9). Stimulation of anticoagulation system by DIP-alpha-thrombin in such animals revealed no heparin in the blood. Mast cell subpopulation was characterized by light cell predominance and low heparin saturation index. The nature of cell degradation remained unchanged. The data obtained indicate the defects in mast cell pool in animals with experimental atherosclerosis.     

Mast cell degranulation requires N-ethylmaleimide-sensitive factor-mediated SNARE disassembly

Mast cells possess specialized granules that, upon stimulation of surface FcR with IgE, fuse with the plasma membrane, thereby releasing inflammatory mediators. A family of membrane fusion proteins called SNAREs, which are present on both the granule and the plasma membrane, plays a role in the fusion of these granules with the plasma membrane of mast cells. In addition to the SNAREs themselves, it is likely that the SNARE accessory protein, N-ethylmaleimide-sensitive factor (NSF), affects the composition and structure of the SNARE complex. NSF is a cytoplasmic ATPase that disassembles the SNARE complexes. To investigate the role of NSF in mast cell degranulation, we developed an assay to measure secretion from transiently transfected RBL (rat basophilic leukemia)-2H3 mast cells (a tumor analog of mucosal mast cells). RBL-2H3 cells were cotransfected with a plasmid encoding a human growth hormone secretion reporter along with either wild-type NSF or an NSF mutant that lacks ATPase activity. Human growth hormone was targeted to and released from secretory granules in RBL-2H3 cells, and coexpression with mutant NSF dramatically inhibited regulated exocytosis from the transfected cells. Biochemical analysis of SNARE complexes in these cells revealed that overexpression of the NSF mutant decreased disassembly and resulted in an accumulation of SNARE complexes. These data reveal a role for NSF in mast cell exocytosis and highlight the importance of SNARE disassembly, or priming, in regulated exocytosis from mast cells.