Heterogeneity of human basophils and mast cells in response to muscle relaxants

We investigated thein vitro effects of increasing concentrations (10⁻⁵–10⁻³ M) of four muscle relaxants (succinylcholine, d-tubocurarine, vecuronium and atracurium) on histamine release (HR) from human peripheral blood basophils and mast cells isolated from lung parenchyma (HLMC) and skin tissues (HSMC). Basophils released less than 5% of their histamine content when incubated with any one of the muscle relaxants. In contrast, mast cells showed a marked heterogeneity in their response. Succinylcholine did not induce HR from any type of mast cell, and only high concentrations of d-tubocurarine (10⁻³ M) caused HR from HSMC and HLMC. Vecuronium concentration-dependently induced HR from HLMC and HSMC. Atracurium concentration-dependently caused marked HR from HLMC and HSMC up to a maximum of 46.2±15.1% and 30.6±6.0%, respectively. From both HLMC and HSMC HR caused by atracurium and vecuronium was extremely rapid (t_1/2<1 min). The releasing activity of atracurium and vecuronium on HLMC and HSMC was reduced, but not abolished, by lowering the temperature of the incubation buffer to 22°C and 4°C. These results confirm that there are functional differences between human basophils and mast cells and among mast cells isolated from different anatomical sites in response to the muscle relaxants tested.

     

Interleukin-2 receptor expression in human mast cells and basophils

Surgical human thymus, upper respiratory tract, lung and small and large bowel specimens were analyzed for the presence of interleukin 2 receptor (IL2-R)-positive cells. Histochemical (toluidine) and immunologic (anti-IL2-R monoclonal antibody) staining procedures revealed a distinct anti-IL2-R positivity in most of metachromatically staining cells. These positive cells were observed not only in tissues showing strong inflammatory reaction and mast cell hyperplasia, as in Crohn's disease, but also in those not histologically affected by pathologic conditions. This finding suggested that human mast cells, like T blast cells, express the p55 chain of IL2-R on their surface. To see whether IL2-R was being actively synthesized, a cell preparation rich in peripheral blood basophils (PBB), which are cells closely related to mast cells, was obtained. Ultrastructural analysis of PBB after indirect immunogold procedure revealed that the vast majority expressed the IL2-R. Moreover, the presence of intracellular reaction products in the cytoplasm of most membrane-positive PBB was indicative of active antigen synthesis. Furthermore, Northern blot analysis evidenced specific IL2-R mRNA in PBB, while its expression was augmented several times when PBB were cultured in the presence of stimulated T cell supernatant.     

Release of elastase from purified human lung mast cells and basophils

Elastase, a serine protease, is capable of inducing severe lung destruction in experimental animal models. We now report that this proteinase exists preformed in neutrophil-free sonicates of purified human lung mast cells (>98% purity) and in circulating peripheral blood basophils (>97% purity). The elastase levels in both cell types (41–174 ng/10⁶ cells) represents approximately 3–20% of those found in human neutrophils; both cell types released their elastase following anti-IgE and ionophore A23187 challenge. The apparent molecular size of the mast cell enzyme on Sephadex G-100 gel filtration, as well as its inhibition profile, was identical to that of purified human neutrophil elastase. This mast cell elastase is identical to our previously reported mast cell-derived Hageman factor cleaving activity. Mast cell-, basophil-, and neutrophil-derived elastases cleave Hageman factor into fragments of 52,000 and 28.000 Da; cleavage by all three enzymes is inhibited by preincubation with polyclonal antibodies directed against human neutrophil elastase.Supported by grant A120634(ESS)     

Role of mast cells and basophils in pruritus

To protect our body systems, there is a constant interactive conversation between the skin nervous and immune system. Important elements of this conversation in the skin include mast cells, basophils, and sensory nerve fibers. These cells employ a vast array of sensors that detect danger and react accordingly. This reaction, summarized as neurogenic inflammation, manifests at the conscious level as sensations including pain and itch. Here we provide a perspective on the blossoming knowledge that is illuminating connections between mast cells, basophils, and sensory nerve fibers in the mediation of itch. We discuss established mediators and receptors, in particular cytokine and neuropeptide pathways, upstream proteases, and proteinase-activated receptors, and the emerging role of mas-related G-protein-coupled receptors in itch.     

Cytokine production by mast cells and basophils

Mast cells and/or basophils have been implicated in the expression of a wide variety of biological responses, including immediate hypersensitivity reactions, host responses to parasites and neoplasms, angiogenesis, tissue remodeling, and immunologically non-specific inflammatory and fibrotic conditions. Recent findings suggest that an important mechanism by which mast cells influence such biological responses is through the production of a broad panel of multifunctional cytokines. In contrast, the extent to which basophils can produce cytokines is uncertain.     

Immunogold probe for the light-microscopic phenotyping of human mast cells and basophils

A method is presented whereby light microscopic phenotyping of human basophils and lung mast cells was obtained by simultaneous labeling of surface antigens with specific mouse monoclonal antibodies and an anti-mouse IgG immunogold probe and histochemical staining with alcian blue, a highly specific stain for these cells' granules. The double labeling technique offers a useful immunohistochemical method for the phenotyping of basophils and mast cells in impure cell preparations. The procedure is relatively simple and requires small cell samples while providing accurate information on functional and differentiation markers. Preliminary results of human lung mast cell and peripheral blood basophil phenotyping discloses class I histocompatibility antigens on both cell types and class II antigen HLA DR on 10-20% of mast cells alone. Lung mast cells also unexpectedly exhibit the leukocyte marker HLE-1.     

Forskolin inhibits the release of histamine from human basophils and mast cells

We found that forskolin (10^–7 to 3×10^–5 M) caused dose-related inhibition of antigen-induced histamine release from human basophil leukocytes. The dose-response inhibition curve was paralleled by a forskolin-induced increase in cyclic AMP (cAMP) levels in human leukocyte preparations. The kinetics of inhibition of histamine release and of the increase in leukocyte cAMP were the same.In a second series of experiments we evaluated the effect of forskolin on antigen-induced histamine release from chopped human lung passively sensitized with serum from an allergic patient. Forskolin (10^–7 to 3×10^–5 M) dosedependently inhibited the release of histamine from human lung mast cells. Thus forskolin appears to modulate the release of mediators of the immediate hypersensitivity reaction, presumably through activation of adenylate cyclase in human basophils and mast cells.Supported by Grants from the C.N.R. (83.00430.04 and 84.01756.04) and M.P.I. (Rome, Italy).     

Platelet augmentation of IgE-dependent histamine release from human basophils and mast cells

A factor(s) from human platelets enhances IgE-mediated histamine release from human basophils and mast cells. This effect is directly related to the platelet number; at physiological platelet/leukocyte ratios (40:1), the enhancement was 66 +/- 11%. Platelet stimulation by thrombin more than doubled the enhancement, to 172 +/- 10% at 40:1. Mast cell release was also enhanced by platelets although the magnitude was more limited (86 +/- 13% at 40:1 with thrombin). Direct basophil/platelet contact was unnecessary in that platelet supernatants were fully active; a direct platelet factor/basophil interaction is suggested, however, by the fact that basophils purified 100-fold with respect to other leukocytes were enhanced by the platelet factors. The appearance of platelet-enhancing activity is associated with the release of an alpha-granule marker (PF4) rather than with products of arachidonic acid metabolism (thromboxane B2). The platelet factor(s) responsible for these effects are not dialyzable, are heat stable and do not appear to be identical to PF4 or platelet-derived growth factor (PDGF). Since anti-IgE-stimulated basophils cause PF4 release and this correlates with the release of enhancing factor, we suggest that a pro-inflammatory feed forward relationship exists. Together with our previous data showing that platelets are activated in vivo during antigen challenge of allergic asthmatic subjects, these results suggest that platelets may be important in modulating IgE-mediated allergic reactions in man.     

Roles of basophils and mast cells in cutaneous inflammation

Mast cells and basophils are associated with T helper 2 (Th2) immune responses. Newly developed mast cell-deficient mice have provided evidence that mast cells initiate contact hypersensitivity via activating dendritic cells. Studies using basophil-deficient mice have also revealed that basophils are responsible for cutaneous Th2 skewing to haptens and peptide antigens but not to protein antigens. Recently, several studies reported the existence of innate lymphoid cells (ILCs), which differ from classic T cells in that they lack the T cell receptor. Mast cells and basophils can interact with ILCs and play some roles in the pathogenesis of Th2 responses. Basophil-derived interleukin (IL)-4 enhances the expression of the chemokine CCL11, as well as IL-5, IL-9, and IL-13 in ILC2s, leading to the accumulation of eosinophils in allergic reactions. IL-33-stimulated mast cells can play a regulatory role in the development of ILC2-mediated non-antigen-specific protease-induced acute inflammation. In this review, we discuss the recent advances in our understanding of mast cells and basophils in immunity and inflammation.     

Heterogeneity of calcium channels in mast cells and basophils and the possible relevance to pathophysiology of lung diseases: A review

Calcium plays a critical role in the formation and secretion of a wide variety of chemical mediators. Calcium slow-channel blockers,e.g. nifedipine and verapamil, have been shown to inhibit the synthesis of SRS (SRS-A, leukotrienes) in human and guinea pig lung tissue, thromboxane A₂ formation in rat lung and platelet activating factor in human neutrophils. Verapamil and nifedipine also prevent the release of lysosomal enzymes from rabbit and human polymorphonuclear neutrophils. Calcium-channel blockers produce variable inhibitory effects on allergic and nonallergic histamine secretion. Ca⁺⁺-entry blockers also inhibit the Ca⁺⁺ uptake (influx) into mast cells. Many of these inhibitory effects of Ca⁺⁺ antagonists are antagonized by an increased extracellular Ca⁺⁺ ion concentration. The magnitude of the inhibitory influences of Ca⁺⁺-channel blockers on allergic and nonallergic release of chemical mediators appears to depend on the cell source, species, nature and the concentration of the secretory stimuli as well as on the composition and pH of buffers and the concentration of Ca⁺⁺-entry blockers used. The data summarized in this review suggest the existence of a functional heterogeneity of Ca⁺⁺ channels in leukocytes, mast cells and basophils. Interference with the Ca⁺⁺-dependent steps involved in the formation and/or release of chemical mediators appears to be the primary mode of action for Ca⁺⁺-channel blockers in these cells.The differential effects of Ca⁺⁺ antagonists on Ca⁺⁺-dependent activation of phospholipase A₂, 5-lipoxygenase, and calmodulin (or other intracellular Ca⁺⁺-binding proteins) in different cell types (mast cells, basophils, leukocytes, lung tissue, etc.) may explain the variation of their effectiveness in inhibiting the synthesis/release of chemical mediators and antagonizing bronchoconstriction in response to diverse stimuli.During the process of hypersensitization and in immediate hypersensitivity diseases, Ca⁺⁺ homeostasis (uptake, mobilization, distribution, relocation, etc.) may be altered in leukocytes (mast cells, basophils) and lung tissues. The altered Ca⁺⁺ homeostasis could be responsible for the induction of airway hyperreactivity in asthmatics and for hyperreleasability of chemical mediators from leukocytes, mast cells and other cell types.The development of drugs (Ca⁺⁺-channel blockers, antiallergic agents) that are capable of selectively altering Ca⁺⁺-dependent functions in leukocytes (mast cells, basophils, macrophages) and lung tissue in disease staes would offer an attractive alternative and an effective therapeutic approach for obstructive respiratory diseases,e.g. allergic asthma, exercise-induced asthma and a variety of other mediator-dependent allergic disorders.     

The reactivity of dispersed human lung mast cells and peripheral blood basophils to acetylcholine]

To clarify the relation between human lung mast cells and parasympathetic nerve function as well as IgE mediated allergic reactions, highly purified dispersed human lung mast cells were obtained by using the techniques of scissors dispersion, enzymatic treatment, percoll centrifugation and exclusion of adherent cells. The reactivity to acetylcholine was examined by observing the histamine release of purified mast cells. Moreover, peripheral blood basophils, which have many functional similarities with mast cells, were also examined in the same manner. The following results were obtained; 1) Histamine was significantly released from dispersed human lung mast cells at a final concentration of 10(-5) M acetylcholine (p less than 0.05); the peak of histamine release was 10(-4) M of acetylcholine. Acetylcholine had the additional effect of releasing histamine in response to anti-IgE. Histamine release was partially inhibited by 10(-5) M atropine. 2) Basophils had no response to acetylcholine. These results suggest that human lung mast cells play an important role in the defensive mechanism as an effector cell of acetylcholine-mediated autonomic nerve system as well as IgE-mediated allergic reaction.     

The role of mast cells and basophils in inflammation

Mast cells are positioned in the asthmatic airways so that they are able to respond to the inhaled environment. During active disease, the cells are primed to secrete an array of preformed and newly generated inflammatory mediators including histamine, neutral proteases and heparin sulphate, prostaglandins and cysteinyl leukotrienes as well as an array of cytokines and chemokines that are involved in leucocyte recruitment and activation. These cells are a potent source of mediators in both allergen- and exercise-induced asthma and possibly in asthma provoked by other stimuli such as adenosine and inhaled air pollutants. The important role played by mast cells in maintaining airway dysfunction in asthma is underpinned by the efficacy of mediator inhibitors, such as those interfering with the release or action of the leukotrienes, agents that inhibit mast cell activation such as sodium cromoglycate and the recently studied E-20 humanized monoclonal antibody that binds to and removes IgE. The recent discovery of novel inhibitory pathways involving inhibitory motifs (ITIMS) on critical cell surface signalling molecules has opened up new possibilities for preventing mast cell activation. Future research will focus on more effective ways for inhibiting the mast cell's contribution to asthma and understanding what role this unique cell has in the pathogenesis of airway wall remodelling.