Bernhard F. Gibbs

Human skin mast cells rapidly release preformed and newly generated TNF‐α and IL‐8 following stimulation with anti‐IgE and other secretagogues

Abstract: Several groups have previously reported that rodent or human leukemic mast cells produce inflammatory cytokines such as TNF‐α and IL‐8 as well as the pro‐allergic cytokines IL‐4, IL‐5 and IL‐13. Comparatively little is known, however, regarding the ability of normal human skin mast cells to secrete these factors following either IgE‐dependent or IgE‐independent modes of activation. We therefore investigated whether normal human skin mast cells produce these cytokines following stimulation by a variety of secretagogues. Enriched isolated skin mast cells released both TNF‐α and IL‐8 following activation with either anti‐IgE, SCF, substance P, compound 48/80 or A23187. This release was dose‐ and time‐dependent, with maximal levels being reached within 4 h of stimulation involving, in part, the secretion of preformed stores of both cytokines. In accordance with this, using lysates of highly purified (>90%) skin mast cells, we could demonstrate that both TNF‐α and IL‐8 mRNA and protein were present in both unstimulated as well as stimulated mast cells. In stark contrast to these results, no significant levels of either IL‐4, IL‐5 or IL‐13 were detected, regardless of the secretagogue used or the period of stimulation. These results show that human skin mast cells are capable of rapidly secreting pro‐inflammatory cytokines like TNF‐α and IL‐8 following IgE‐dependent activation and stimulation by the neuropeptide substance P, SCF and the basic polypeptide analogue compound 48/80. In contrast to other types of human mast cells however, human skin mast cells were incapable of secreting IL‐4, IL‐5 or IL‐13 in these settings.

The 21st century renaissance of the basophil? Current insights into its role in allergic responses and innate immunity

Abstract:  Basophils and mast cells express all the three subchains of the high‐affinity immunoglobulin E (IgE) receptor FcɛRI and contain preformed histamine in the cytoplasmic granules. However, it is increasingly clear that these cells play distinct roles in allergic inflammatory disease. Despite their presence throughout much of the animal kingdom, the physiological function of basophils remains obscure. As rodent mast cells are more numerous than basophils, and generate an assortment of inflammatory cytokines, basophils have often been regarded as minor players in allergic inflammation. In humans, however, basophils are the prime early producers of interleukin (IL)‐4 and IL‐13, T helper (Th)2‐type cytokines crucial for initiating and maintaining allergic responses. Basophils also express CD40 ligand which, in combination with IL‐4 and IL‐13, facilitates IgE class switching in B cells. They are the main cellular source for early IL‐4 production, which is vital for the development of Th2 responses. The localization of basophils in various tissues affected by allergic inflammation has now been clearly demonstrated by using specific staining techniques and the new research is shedding light on their selective recruitment to the tissues. Finally, recent studies have shown that basophil activation is not restricted to antigen‐specific IgE crosslinking, but can be caused in non‐sensitized individuals by a growing list of parasitic antigens, lectins and viral superantigens, binding to non‐specific IgE antibodies. This, together with novel IgE‐independent routes of activation, imparts important new insights into the potential role of basophils in both adaptive and innate immunity.

Ambroxol inhibits the release of histamine, leukotrienes and cytokines from human leukocytes and mast cells.

Abstract.Objectives and Design: The effects of the mucolytic agents ambroxol and N-acetylcystein (NAC) were studied on the release of histamine, leukotrienes, cytokines and superoxide anions from a variety of cells involved in the pathogenesis of allergic inflammation.¶Subjects: Mast cells were isolated from human adenoids and skin (n = 5–6). Basophils, monocytes and granulocytes were obtained from Buffy-coat blood obtained from healthy blood donors (n = 4–7) and enriched by density centrifugation.¶Treatment and Methods: Ambroxol or NAC were added to the cells for different periods before stimulation with various immunological and non-immunological secretagogues. Histamine release from mast cells, basophils and monocytes was assayed either by radioimmunoassay or spectrofluorometrically. LTC4 (basophils), LTB4 (neutrophil/eosinophil granulocytes or monocytes), IL-4 and IL-13 (basophils) were measured by ELISA.¶Results: Ambroxol inhibited histamine release by more than 50% from human adenoidal mast cells (1000 μM ambroxol) and skin mast cells (100 μM ambroxol) stimulated by Con A and compound 48/80, respectively. Ambroxol (100 μM) strikingly inhibited anti-IgE induced release of both histamine, LTC4, IL-4 and IL-13 from basophils and reduced both histamine and LTB4 release induced by C5a or Zymosan in monocytes. The drug also reduced LTB4 and superoxide anion production in granulocytes stimulated by zymosan or fMLP. In all cell types studied, ambroxol was more efficacious following a short preincubation (5–15 min) of the drug with the cells before stimulation. In contrast, NAC produced no clear effects on any of the different cell types studied, regardless of the preincubation period, the concentration or the stimulus employed.¶Conclusions: Unlike NAC, ambroxol is able to not only inhibit acute mediator release from mast cells and leukocytes but also reduce immunomodulatory cytokine generation from basophils and may have beneficial effects in the treatment of allergic respiratory diseases.

Do basophils play a role in immunity against parasites

The link between parasites and eosinophilia has been known for more than a century, although the role of eosinophils in host protection is still an open issue. Much less appreciated, however, is the concurrent systemic induction of a related cell type, the basophil, in parasitized hosts. To date, little is known about the role of basophils in immunity against parasites, but recent evidence points to a possible crucial role in the initiation of T-helper type 2 responses in the host. In this article, we review the current understanding of parasitic infections and basophils and discuss their putative role in immunity.

Regulation of mediator secretion in human basophils by p38 mitogen-activated protein kinase: phosphorylation is sensitive to the effects of phosphatidylinositol 3-kinase inhibitors and calcium mobilization

Although human basophils modulate allergic diseases by secreting histamine, leukotriene C4, interleukin (IL)‐4, and IL‐13, the intermediary signals controlling the release of these mediators are poorly understood. Here, we show that p38 mitogen‐activated protein kinase (MAPK) crucially affects basophil activation following stimulation with various secretagogues. Phosphorylation of p38 MAPK occurred within 5 min following anti‐immunoglobulin (Ig)E stimulation, but was more rapidly activated in basophils stimulated with formyl‐Met‐Leu‐Phe or A23187. Additionally, activation of p38 MAPK to the above stimuli was dependent on extracellular influx and intracellular mobilization of calcium. SB 203580, a specific p38 MAPK inhibitor, blocked anti‐IgE‐induced secretion of all basophil mediators and reduced not only p38 MAPK, but also extracellular signal‐regulated kinases 1 and 2 activity, whereas the MAPK antagonist, PD 098059, did not affect p38 MAPK. IgE‐dependent activation of p38 MAPK and MKK3/6 was affected by LY 294002 and wortmannin, suggesting that these kinases are targets for phosphatidylinositol 3 kinase (PI 3‐K). We conclude that p38 MAPK is a pivotal regulator of basophil function downstream of PI 3‐K activation and calcium mobilization.

Inhibitors of PI 3-kinase and MEK kinase differentially affect mediator secretion from immunologically activated human basophils.

Effects of inhibitors of PI 3‐kinase and MEK kinases were investigated on histamine, leukotriene C4 (LTC4), and cytokine release from human basophils stimulated with anti‐IgE. The PI 3‐kinase antagonists wortmannin (> 10 nM) and LY 294002 (> 1 μM) strongly inhibited anti‐IgE‐induced release of all mediators by 40–100%. This was contrasted by the effects of the MEK kinase inhibitor PD 098059, which weakly inhibited histamine, interleukin (IL)‐4, and IL‐13 release but was substantially more efficacious at blocking LTC4 production (> 70% at 10 μM). Previous studies have shown that arachidonic acid synthesis is controlled by MEK kinases. We observed that wortmannin, LY 294002, and PD 098059 reduce basophil ERK‐1,2 activation, thus implying that, with regard to arachidonic acid metabolism, MEK kinases are a downstream target for PI‐3‐kinase. Our results demonstrate a universal regulatory role played by PI 3‐kinases in basophil mediator production and release, whereas MEK kinase signaling is largely limited to controlling arachidonic acid metabolism. J. Leukoc. Biol. 65: 883–890; 1999.

Echinococcus multilocularis metacestode extract triggers human basophils to release interleukin-4

Infections with parasitic helminths are associated with a T helper 2 (Th2) immune response and IgE production. The underlying mechanism, however, is only partially understood. Recently we have isolated a protein from extracts of Schistosoma mansoni eggs that triggers human basophils from non‐sensitized donors to release interleukin‐4 (IL‐4), the key cytokine of a Th2 response. We called this protein IPSE (for IL‐4‐inducing principle from Schistosoma mansoni eggs). Supposing that IPSE‐like IL‐4‐inducing activities might be a general principle shared among different helminth species, we investigated extracts from the cestode E. multilocularis for its effect on human basophils. Our results showed that extracts from metacestodes of E. multilocularis cause basophil degranulation, as well as the secretion of histamine, IL‐4 and IL‐13, in a dose‐dependent manner. IgE stripping and resensitization of basophils indicated that the mechanism of IL‐4 induction requires the presence of IgE on the cells. Since analogous properties have been demonstrated earlier for IPSE, we think that S. mansoni and E. multilocularis may induce a Th2 response in their hosts via a related mechanism, namely, by the induction of IL‐4 release from basophils.

IgE autoantibodies against the intracellular domain of BP180

Background  Bullous pemphigoid (BP) is an autoimmune subepidermal blistering disease characterized by autoantibodies against the hemidesmosomal proteins BP180 (type XVII collagen) and BP230. BP not only involves IgG‐mediated neutrophil activation, leading to blistering, but also IgE‐dependent activation of mast cells and basophils. While IgG and IgE autoantibodies target the extracellular noncollagenous (NC) 16A domain of BP180, little is known whether other BP180 regions are targeted by these antibody classes.

Lack of protease activated receptor (PAR) expression in purified human basophils

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In vitro investigations with the histamine H1 receptor antagonist, epinastine (WAL 801 CL), on isolated human allergic effector cells.

Abstract.Objective and Design: Skin mast cells, basophils and eosinophils are effector cells of acute and subacute allergic responses due to their capacity to produce a large number of (pro)inflammatory mediators. Histamine H1 receptor antagonists, such as epinastine (WAL 801 CL), have been described to partially exert antiallergic and anti-inflammatory effects both in vivo and in vitro in addition to their antihistaminergic properties. The aim of the present study was to investigate whether epinastine could influence the in vitro activation of isolated human skin mast cells, basophils and eosinophils induced by different secretagogues.¶Methods: Cells were isolated from healthy women following plastic surgery and healthy blood donors, respectively. Mast cells were isolated by enzymatic digestion of the skin. Blood cells were isolated by gradient centrifugation and negative selection with magnetic beads.¶Results: A wide range of concentrations of the drug (1 nmol/l to 100 mmol/l) did not significantly inhibit histamine release from basophils induced by immunologic (anti-IgE, concanavalin A, priming factors interleukin-3 and interleukin-5) and non immunologic (A23187, ionomycin, 12-o-tetradecanoyl-phorbol-13-acetate, C5a, formyl-methionyl-leucyl-phenylalanine) stimuli. Furthermore, the drug had no effect on A23187-induced release of eosinophil cationic protein from eosinophils. However, at a concentration >0.1nmol/l, IgE-mediated LTC4 production from basophils was significantly suppressed. Histamine release from skin mast cells due to anti-IgE or A23187 was inhibited by epinastine in a dose-dependent fashion, whereas substance P-induced activation as well as stem cell factor priming were not. Epinastine did not inhibit isolated protein kinase C from rat brain.¶Conclusion: The results confirm previous in vivo and in vitro observations obtained from animal models that epinastine exerts antiallergic and antiinflammatory effects. Whether the observed effects are due to non specific membrane interactions or by influencing intracellular signal transduction elements has to be further elucidated.