Modification of allergic inflammation in murine model of rhinitis by different bacterial ligands: involvement of mast cells and dendritic cells

BACKGROUND: It has been suggested that airway bacterial infections exacerbate allergic disorders, and bacterial components in the air affect allergic inflammation via Toll-like receptors expressed on mast cells and dendritic cells in the airway mucosa. OBJECTIVE: Peptidoglycan (PGN) is a major component of the bacterial cell wall. We investigated the effect of PGN on the effector phase of allergic inflammation, in comparison with the effect of CpG-oligodeoxynucleotides (CpG), which is known to be a Th1 adjuvant. METHODS: Ovalbumin (OVA)-sensitized mice were challenged intranasally with OVA alone or OVA together with PGN or CpG. Nasal allergic symptoms and eosinophilia were scored, and the OVA-specific cytokine response was examined in the cells of cervical lymph nodes and nasal mucosa. Bone marrow-derived mast cells (BMMCs) and dendritic cells (BMDCs) were stimulated with PGN or CpG in vitro, and the expression level of cytokines and chemokines was examined by RT-PCR. In addition, the expression level of chemokines was examined by RT-PCR in mast cells of OVA-sensitized mice challenged with OVA alone or OVA together with PGN or CpG. RESULTS: PGN exposure exacerbated the nasal allergic symptoms and eosinophilia, whereas CpG exposure suppressed them. In addition, PGN exposure increased the OVA-specific IL-4 response in the cells, whereas CpG exposure decreased it. On the other hand, there were no significant differences in the OVA-specific IFN-gamma response. PGN but not CpG induced the expression of thymus and activation-regulated chemokine (TARC) and macrophage/monocyte-derived chemokine (MDC) in both BMMCs and mast cells of mice sensitized and challenged with OVA. CpG but not PGN induced the expression of IFN-beta and interferon-inducible protein-10 (IP-10) in BMDCs, and histamine did not influence this effect. CONCLUSION: These results demonstrate that PGN exposure exacerbates allergic inflammation mainly via mast cells, whereas CpG exposure suppresses allergic inflammation mainly via dendritic cells.     

Effect of topical glucocorticoid treatment on nasal mucosal mast cells in allergic rhinitis

Glococorticoids were previously considered not to affect the immediate allergic reaction. However, in a nasal allergen challenge, an inhibitory effect on the nasal symptoms induced at the challenge has been shown to occur in patients treated with a recently developed Glucocorticoid, budesonide, for 1 week prior to the challenge. This treatment was also found to reduce tissue histamine levels in the nasal mucosa. Mast cells in the mucosa were therefore studied with a view to finding out whether this reduction could be due to a reduction of mast cells. A double-blind study was performed in 14 asymptomatic patients. Nasal biopsies were made before and after I week's treatment with either budesonide or placebo. The number of mast cells was counted in two Epon sections after the specimens has undergone specific staining with toluidine blue. No quantitative or qualitative morphological changes in the mast cells were found as a result of treatment.     

The cellular basis for allergic rhinitis

In both seasonal and perennial rhinitis there is epithelial mast cell accumulation and tissue infiltration by eosinophils. Activation of these cells can be observed by electron microscopy and by elevated levels of tryptase and eosinophil cationic protein in nasal lavage fluid. Furthermore, seasonal increases in the antigen presenting cell (Langerhans'cell) are also evident. Investigations into the mechanisms involved in cell accumulation and activation reveals upregulation of leucocyte endothelial adhesion molecules and an increase in interleukin-4 (IL-4) in naturally occurring rhinitis, while mRNA for IL-4, IL-5 and granulocyte macrophage colony stimulating factor activity and lavage tumour necrosis factor-alpha (TNFα) levels are increased following local allergen challenge. These cytokines may be derived from a variety of sources, including mast cells, eosinophils and T-lymphocytes, and contribute to the underlying inflammatory process in rhinitis.     

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.     

Potato lectin activates basophils and mast cells of atopic subjects by its interaction with core chitobiose of cell-bound non-specific immunoglobulin E

A major factor in non-allergic food hypersensitivity could be the interaction of dietary lectins with mast cells and basophils. Because immunoglobulin E (IgE) contains 10-12% carbohydrates, lectins can activate and degranulate these cells by cross-linking the glycans of cell-bound IgE. The present objective focuses on the effect of potato lectin (Solanum tuberosum agglutinin; STA) for its ability to release histamine from basophils in vitro and mast cells in vivo from non-atopic and atopic subjects. In this study, subjects were selected randomly based on case history and skin prick test responses with food, pollen and house dust mite extracts. Skin prick test (SPT) was performed with STA at 100 microg/ml concentration. Histamine release was performed using leucocytes from non-atopic and atopic subjects and rat peritoneal exudate cells. SPT on 110 atopic subjects using STA showed 39 subjects positive (35%); however, none showed STA-specific IgE; among 20 non-atopic subjects, none were positive by SPT. Maximal histamine release was found to be 65% in atopic subjects (n = 7) compared to 28% in non-atopic subjects (n = 5); the release was inhibited specifically by oligomers of N-acetylglucosamine and correlates well with serum total IgE levels (R(2) = 0.923). Binding of STA to N-linked glycoproteins (horseradish peroxidase, avidin and IgG) was positive by dot blot and binding assay. As potato lectin activates and degranulates both mast cells and basophils by interacting with the chitobiose core of IgE glycans, higher intake of potato may increase the clinical symptoms as a result of non-allergic food hypersensitivity in atopic subjects.     

Relationships between allergic inflammation and nasal airflow in children with persistent allergic rhinitis due to mite sensitization

BACKGROUND: Allergic rhinitis is associated with Th2-dependent inflammation. Nasal obstruction is the most typical symptom in children with mite allergy. OBJECTIVES: The aim of this study was to evaluate the possible relationships among nasal symptoms, allergic inflammation, including inflammatory cells and cytokine pattern, and nasal airflow in children with persistent allergic rhinitis because of mite sensitization. METHODS: Twenty children (13 males and seven females, mean age 13.4 +/- 1.6 years) with persistent rhinitis because of mite allergy were evaluated. All of them had moderate-severe grade of nasal obstruction. Total symptom score (TSS), rhinomanometry, nasal lavage, and nasal scraping were obtained in all subjects. Inflammatory cells were counted by conventional staining; interleukin (IL)-5, and IL-8 were measured by immunoassay on fluids recovered from nasal lavage. RESULTS: Eosinophils were significantly associated with TSS (R = 74.4%, P = 0.0002), with IL-5 (R = 90.6%, P < 0.0001) and with nasal flow (R = -69%, P = 0.0007), but not with IL-8 (R = 0.1%, P = 0.995). Eosinophil levels were shown to independently predict nasal flow (P < 0.001), with flow decreasing linearly for increasing eosinophils, together with a significant effect of neutrophils (P = 0.016, linear increase in flow) and a borderline effect of IL-8 (P = 0.063, linear increase in flow). CONCLUSIONS: This study demonstrates the close association between IL-5 concentration and eosinophil infiltration. In addition, there is clear evidence concerning the relationship between eosinophil infiltration and nasal airflow. Thus, nasal eosinophils can be regarded as the most important predictor of upper airway function. These findings constitute first evidence of the relationship between nasal airflow impairment and Th2-related eosinophilic inflammation in children with persistent allergic rhinitis because of mite sensitization.     

Effect of topical corticosteroids on seasonal increases in epithelial eosinophils and mast cells in allergic rhinitis: a comparison of nasal brush and biopsy methods

BACKGROUND: Nasal brushing and nasal biopsy are well-tolerated sampling techniques. Seasonal grass pollen-induced rhinitis is characterized by epithelial mast cell infiltration and seasonal increases in both epithelial and sub-mucosal eosinophils. OBJECTIVE: To compare the ability of the nasal brush and nasal biopsy techniques to detect natural seasonal increases in eosinophils and mast cells, and to assess the influence of topical corticosteroid. METHODS: Nasal brush samples and nasal biopsies were collected from 46 grass pollen-sensitive seasonal rhinitis patients before the grass pollen season and at the peak of the pollen season following 6 weeks' treatment with either fluticasone propionate aqueous nasal spray (200 microg, twice daily) or placebo nasal spray. RESULTS: Placebo patients showed seasonal increases in epithelial eosinophils both with nasal brushing (P < 0.0001) and biopsy (P < 0.001). Epithelial mast cell numbers also increased during the pollen season as detectable by brushing (P < 0.0001) and biopsy (P < 0.03). Changes in cell numbers measured by nasal brushing correlated with those observed with nasal biopsy, both for eosinophils and mast cells (P < 0.05). Sub-mucosal eosinophils but not mast cells also increased during the pollen season (P < 0.002). Nasal brushing and biopsy revealed that fluticasone treatment inhibited seasonal increases in epithelial eosinophils (P < 0.00001) and epithelial infiltration by mast cells (nasal brushing P < 0.00001 and nasal biopsy P < 0.01). Fluticasone also inhibited seasonal increases in sub-mucosal eosinophils (P < 0.001) and significantly reduced nasal symptoms (P < 0.001). CONCLUSION: Nasal brushing harvests sufficient inflammatory cells from the surface of the nasal mucosa to be used in lieu of nasal biopsies in observation of the effect of drugs on the nasal epithelium.     

Nerve growth factor localization in the nasal mucosa of patients with persistent allergic rhinitis

Background and objectives:  Nerve growth factor (NGF) and NGF receptors have been shown to be expressed by structural and infiltrating inflammatory cells in the human allergic bronchial mucosa and conjunctiva. In the nose, a positive immunostaining for NGF was recently reported in biopsies of subjects undergoing surgery for refractory nasal obstruction. This study was aimed at studying by immunohistochemistry NGF expression and localization in the nasal mucosa from subjects with moderate/severe persistent allergic rhinitis and natural allergen exposure.
Methods:  Immunostaining for NGF, tryptase and eosinophil cationic protein was performed in human nasal turbinate sections of 25 patients affected by persistent allergic rhinitis and sensitization to Dermatophagoides pteronyssinus .
Results:  NGF was consistently expressed in the epithelium and in the submucosa of allergic rhinitic subjects, preferentially localized in eosinophils and mast cells. A strong NGF immunostaining was found in mucous cells of the epithelial lining and in the submucosal glands.
Conclusions:  As previously shown for allergic asthma and allergic conjunctivitis, NGF is also detectable in the nasal mucosa of patients with persistent allergic rhinitis. The preferential NGF localization in mucous cells of the epithelial lining and in the submucosal glands suggests a possible role for NGF in modulating secretion in allergic rhinitis and possibly other allergic diseases.     

The history of mast cell and basophil research – some lessons learnt from the last century

This year (2013) marks the 50th anniversary of death of Otto Carl Willy Prausnitz (1876–1963) and Heinz Küstner (1897–1963). The two physicians, when working at the Hygiene Institute at the University of Breslau, Germany (Prausnitz was the Head of the Institute), described in 1921 what is still called today the Prausnitz–Küstner or PK reaction showing that allergy could be transferred from the allergic person by transferring serum to a healthy person. Their discovery ended the belief that an anaphylactic/allergic reaction was caused by poisons, but to the contrary showed that the presence of the hypersensitivity factor could be transferred to other people. We know now that this factor is immunoglobulin E (IgE), sensitizing mast cells and basophils to respond to an allergic stimulus. We take this occasion to retrace some of the important discoveries and lessons learnt from the last century relating to the function of these two cell types as effectors of the IgE system and the mediators they produce.     

Cellular infiltration and cytokine mRNA expression in perennial allergic rhinitis

BACKGROUND: Allergen challenge in allergic rhinitis patients leads to local eosinophilia and Th2-type cytokine expression. Natural exposure to grass pollen is additionally characterized by epithelial mast-cell infiltration. We hypothesized that perennial allergic rhinitis is also associated with T-cell and eosinophil infiltration of the nasal mucosa, local Th2-type cytokine expression, and increased numbers of nasal epithelial mast cells. METHODS: Nasal biopsies from perennial allergic rhinitis patients and controls were analysed by immunocytochemistry for different cell populations and in situ hybridization for cytokine mRNA-expressing cells. RESULTS: Perennial allergic rhinitis was associated with increased numbers of submucosal CD3+ T cells (P=0.05), EG2+ activated eosinophils (P=0.01), and CD68+ macrophages (P=0.01) compared to controls. Epithelial, but not submucosal, tryptase-positive mast cells were also elevated in rhinitics compared to controls (P=0.01). The numbers of cells expressing interleukin (IL)-5 were higher (P=0.01) and the numbers of cells expressing IL-2 were lower (P=0.04) in rhinitic patients than controls. There were no significant differences for either IL-4 or interferon-gamma between the groups. CONCLUSIONS: Perennial allergic rhinitis is characterized by mast-cell migration into the epithelium; submucosal infiltration by T cells, eosinophils, and macrophages; and an imbalance in local T-cell cytokine production in favour of enhanced IL-5 and reduced IL-2 expression.     

Comparison of olfactory function in patients with seasonal and perennial allergic rhinitis

Hyposmia is a common symptom in allergic rhinitis. However, little is known about differences in the olfactory function of patients with seasonal or perennial allergy. A prospective controlled study was performed on 28 patients with allergic rhinitis to grass pollen and on 47 patients with allergic rhinitis to mites. Sixty-six healthy volunteers served as a control. Olfactory function was evaluated by a modified Connecticut Chemosensory Clinical Research Center testing procedure for threshold, identification, and discrimination. The grass pollen-allergic patients were tested prcseasonally and after 3 weeks of intraseasonal grass pollen exposure: the mite-allergic patients and the volunteers were tested once. In the mite allergies, olfactory threshold, identification, and discrimination tests were significantly worse than in the volunteers (all P<0.0001). In the grass pollen allergies, the results in olfactory identification and discrimination tests were not different from the controls if tested out of the season (both F>0.05). However, in threshold testing (P=0.0139), the results were worse. Intraseasonally, the grass pollen allergies showed a significant decrease in threshold, identification (both P<0.001), and discrimination testing (P=0.0()29). If the intraseasonal pollen allergies were compared to the mite allergies, they showed better results in identification (F=0.(X)87) and threshold (F<0.001) tests, but worse results m discrimination testing (P=0.(XM)2). Therefore, the different kind of allergen exposure seems to result in a different pattern of allergic olfactory dysfunction.     

Evidence for an inflammatory pathophysiology in idiopathic rhinitis

BACKGROUND: The pathophysiology of idiopathic rhinitis is unknown but the disease is classified as being non-allergic on the basis of negative serum IgE radioallergosorbent assay (RAST) and skin prick tests. In contrast, allergic rhinitis has a Th2 type inflammatory pathology mediated by IgE and mast cells. OBJECTIVE: To test the null hypothesis that there would be no difference in the cellular infiltrate for key Th2-associated inflammatory cells between allergic and idiopathic rhinitis. METHODS: We applied strict selection criteria in the recruitment of allergic and idiopathic rhinitis cases. In contrast to previous studies which used cytology or small biopsies, we studied all layers of the mucosa by using whole, full-thickness nasal turbinate specimens with an average length of 2.5 cm. Immunohistochemistry and in situ hybridization techniques were used to compare the distribution and cell populations of mast cells, IgE positive (IgE+) cells, eosinophils and plasma cells in perennial allergic (n = 11) and idiopathic (n = 17) rhinitis, and control nasal mucosal tissue (n = 9). RESULTS: Mast cells and IgE+ cells were significantly increased within the epithelium of allergic and idiopathic mucosa compared to normal mucosa (P < 0.05). More IgE+ cells were present in the allergic group compared to the idiopathic group with the majority of IgE+ cells being mast cells. Both rhinitic groups showed increased eosinophilia localized to the superficial submucosa compared to normal mucosa (P < 0.05). More plasma cells were present in the allergic rhinitic tissue. CONCLUSION: Idiopathic and allergic rhinitic mucosa show similarities in their inflammatory infiltrate suggesting that both groups share a highly localized Th2, IgE-mediated cellular immunopathology.     

Unstimulated basophils in atopic and nonatopic subjects express intracellular interleukin-4: detection by flow cytometry

Background IgE-stimulated cultured basophils from atopic subjects are capable of secreting interleukin-4 (IL-4). We describe a flow-cytometric technique which identified intracellular IL-4 in unstimulated basophils unseparated from peripheral blood mononuclear cells (PBMC) in both atopic (AT) and nonatopic (NC) volunteers.
Methods Freshly isolated PBMC were fixed in 4% paraformaldehyde (PFA). Surface staining with 22E7, a noncompetitive anti-Fc-a antibody, allowed identification of basophils. Permeabilization by 0.1 % saponin allowed staining of intracellular cytokines with specific monoclonal antibodies (mAbs). Two series of experiments utilizing different protocols and anticytokine mAbs were performed. The first protocol required a two-stage fluorochrome staining technique. The availability of fluorochrome-conjugated mAbs allowed a simpler, one-stage labelling procedure for the second protocol.
Results With the first protocol, IL-4 (but not IFN-y), immunoreactivity was detectable in a majority (median 77%) of peripheral blood basophils from both AT and NC subjects (N = 8). Basophil IL-4 immunoreactivity was again evident in experiment 2 but did not differ significantly between AT and NC subjects - either evaluated as percentage of IL-4+ basophils (AT median=66%, NC median=38.4%, P=0.41) or lL-4-specific mean fluorescence (AT median=0.85, NC median=0.3, P=0.07).
Conclusions This simple technique allowed the study of intracellular cytokine expression in unstimulated blood basophils. It demonstrated constitutive basophil expression of lL-4 (but not IFN-y) in all subjects, with no significant increases in atopies.