The roles of antigen-specific immune reactions are discussed in different types of autoimmune diseases. Antigen-specific reactions of T cells are carried out at the level of clones. In this regard, we have established a novel method for analyzing T cell clonality. Analyses of several autoimmune disorders suggest the importance of antigen-specific T cell clones. Furthermore, our findings cast doubt on the idea of epitope spreading, which has been proposed recently for T cell activation in autoimmune processes. Our system also appears to work as a new method for the identification of target antigens recognized by such accumulated T cell clones.

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The prevalence of asthma has been increasing in Japan, like in other parts of the world. Asthma management guidelines were developed in Japan in 1993 and 1998. These guidelines have been shown to contribute to the improvement of asthma management in Japan. Most allergists know the guidelines and make use of them. However, only half the non-specialists surveyed knew the guidelines in 1996-97, although the awareness of the guidelines increased over the next 3 years. Further efforts are required to implement the guidelines widely.

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In the present study, neutrophil functions were examined in vitro in 35 patients (15 men and 20 women; mean age 38 years) suffering from perennial allergic rhinitis. Adherence to nylon, serum chemotaxis and phagocytosisbactericidal activity were assayed. A Candida albicans strain was used for the experiments of phagocytosisbactericidal activity. Adherence to patients' neutrophils was found in normal patients (22.6 ± 1.89%) and healthy controls (26.1 ± 1.59%; P > 0.05). In contrast, the chemotactic ability of the patients' sera was inferior to that of healthy control patients (leucotactic index 20.25 ± 0.76 vs 28.04 ± 0.46 µm, respectively; P < 0.0001). Patients' neutrophils phagocytosed 102.77 ± 6.03 microorganisms per 100 cells, while controls phagocytosed 138.05 ± 7.60 microorganisms per 100 cells (P < 0.001). Bactericidal activity was also severely impaired in patients compared with controls (17.19 ± 1.67 vs 33.23 ± 1.96%, respectively; P < 0.0001). The observed host defense disorders provide additional information that helps to explain the increased susceptibility of these patients to bacterial infections.

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Although sulfidopeptide leukotrienes (sLT) are considered to play an important role in the pathogenesis of asthma, their precise action has not been elucidated in asthmatics during exacerbation. In the present study, we examined sputum concentrations of sLT from asthmatic patients in order to determine whether sLT are actively involved in the exacerbation of asthma. We also examined sputum levels of thromboxane (TX) B₂ and histamine because these mediators are considered to be as important as sLT. The induced sputa by 3% hypertonic saline inhalation were treated with high-performance liquid chromatography and levels of sLT, TXB₂ and histamine were measured with enzyme immunoassay kits. These compounds tended to be elevated in asymptomatic asthmatic patients compared with healthy controls, but the differences were not significant. Levels of sLT and TXB₂ showed no difference between atopic and non-atopic patients, but histamine levels were higher in atopic patients than in non-atopic patients. However, sputa during the exacerbation contained significantly higher levels of sLT than those during the asymptomatic state. In contrast, neither histamine nor TXB₂ showed any changes with exacerbation. These results suggest that sLT may be one of the most potent for mounting the exacerbation of asthma.

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The purpose of the present study was to describe patterns in weekly asthma visits to Accident and Emergency (A&E;) facilities in Trinidad during 1997 and to examine associations with climatic conditions. A census of patients with asthma, defined as those who required bronchodilator nebulization, was taken at five A&E; facilities in the Caribbean island of Trinidad from 1 January to 31 December 1997. Data on patients' ages, gender and dates of visits were obtained from the A&E; records. Climate variables, including rainfall, temperature, relative humidity and wind speed, were obtained from the island's meteorological office. There was a total of 27 848 asthma visits to the five facilities during 1997, of which the pediatric population (< 16 years) accounted for 43.7%. There were steady increases in adult and elderly visits from January to December. In the pediatric population, there was a decrease in visits from weeks 536 in the year, followed by a sharp increase during the next 3 weeks. This sharp increase coincided with the start of the academic school year in September after a 2 month break. The results of multiple regression analysis indicated that pediatric visits to the A&E were positively associated with temperature and wind speed, while visits by adults were positively associated with temperature and relative humidity. There were no independent predictors of asthma visits in the elderly. The results show that there is an association between the climate and asthma visits in Trinidad. However, there is a need for further research to explain the increase in pediatric visits at the start of the school year as well as to elucidate the mechanisms for the observed associations between asthma visits and climate variables.

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Using severe combined immunodeficiency mice engrafted with peripheral blood mononuclear cells from atopic patients, we analyzed the regulatory effects of anti-CD154 antibody on the in vivo induction of human interleukin (IL)-4 and IgE expression. Although anti-CD154 treatment of engrafted mice abrogated mature Cε transcription and IgE production, IL-4 mRNA levels were enhanced by the treatment. In addition, anti-CD154-induced enhancement of intracellular IL-4 synthesis was detectable in both CD4⁺ and CD8⁺ T cell subsets. Furthermore, upregulation of germline Cε transcription could be seen in anti-CD154-treated mice. These results demonstrate that blocking the CD154CD40 interaction with anti-CD154 differentially regulates IL-4 synthesis in T cells and IgE production in B cells. Our data also indicate that antibody ligation of CD154 on T cells causes enhanced synthesis of IL-4, thereby contributing to upregulation of germline Cε transcription in B cells.

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Immunologic mechanisms of allergen immunotherapy are still incompletely understood. Immunotherapy of a higher maximum tolerance dose with reduced risks of systemic reactions, such as anaphylaxis, needs to be developed. Seasonal allergic rhinitis (SAR) is a type I allergic disease characterized by typical seasonal symptoms of rhinitis and an increase in mast cells and T helper (Th) 2-type cells, as well as tissue eosinophilia. The present study was designed to investigate the effect of a modified immunotherapy (IT) with an allergenpullulan conjugate (CS-560) in patients with SAR to Japanese cedar pollen (Japanese cedar pollinosis) using objective parameters, such as analyzing the alteration in the proportion of effector cells like mast cells, eosinophils and T cells, and the Th2 and Th1 cytokine profile. We also analyzed the efficacy of modified IT on the severity of symptoms, using subjective parameters, such as the symptommedication score. Immunotherapy for 15 months duration with the modified drug CS-560 in patients with Japanese cedar pollinosis induced an immune deviation from a Th2- to a Th1-type cytokine profile and inhibited the in-season increase in the proportion of intraepithelial mast cells and eosinophils. These changes were associated with a reduction in the symptommedication score. These data suggest that this modified IT with an allergen pullulan conjugate (CS-560) is an effective mode of treatment of patients with Japanese cedar pollinosis.

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Cross-reactivity in IgE epitopes and T cell epitopes has been reported between major birch pollen allergen (Bet v1) and major apple allergen (Mal d1). To treat people with birch pollinosis complicated by apple hypersensitivity by peptide immunotherapy, a sequential IgE epitope analysis was performed to study IgE epitopes that recognize birch pollen and apple allergens at the level of peptides. Subjects in the present study were three patients who exhibited clinical symptoms indicative of birch pollinosis during the pollen season. Two of the three patients had apple hypersensitivity and the capsulated hydrophobic carrier polymer (CAP)- radioallergosorbent test (RAST) class for apple was at least 2. The IgE epitope assay was performed by peptideCNBr gel binding assay. In two of the three patients, IgE bound strongly to Bet vI 51-70 peptide and also to 11-30, 21-40, 11-120, 111-130 and 141-159 peptides. In the other patient, IgE bound to 11-30, 61-80, 111-130 and 141-159 peptides in a comparable manner. The IgE epitopes of Bet v1 and Mal d1 were found in similar locations. The results of sequential IgE epitope analyses on Bet v1 and Mal d1 revealed epitopes near the proximal regions of these allergens. Thus, the three-dimensional structure of Mal d1 is likely similar to that of Bet v1, which suggests that peptide immunotherapy designed for birch pollen is more than likely to be effective against apple allergy.

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An increased airway muscle content is one of the characteristic features of airway remodeling. Proinflammatory substances, growth factors and mechanical strain promote airway muscle cell proliferation and extracellular signal-regulated kinase (Erk) plays an important role in proinflammatory substance- and growth factor-induced airway smooth muscle (ASM) cell proliferation; however, little is known about intracellular signals that regulate mechanical strain- induced ASM cell proliferation. Therefore, in the present study we examined the role of Erk in mechanical strain-induced DNA synthesis and cell proliferation in human ASM cells. We examined the effect of mechanical strain on the threonine and tyrosine phosphorylation of Erk and activation of Erk and the effect of PD98059 on Erk activity, DNA synthesis and cell proliferation in mechanical strain-loaded human ASM cells. The results showed that mechanical strain-induced phosphorylation and activation of Erk in ASM cells and attenuation of Erk activation by PD98059 resulted in the inhibition of mechanical strain-induced Erk activity and increases in DNA synthesis and cell proliferation in ASM cells. These results indicate that Erk plays an important role in mechanical strain-induced increases in DNA synthesis and cell proliferation in human ASM cells.

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Although the clinical effectiveness of long-term low-dose administration of macrolides for diffuse pan- bronchiolitis, sinobronchial syndrome, chronic sinusitis and otitis media with effusion has been well documented, the mechanism of action remains to be determined. To clarify the effect of macrolides on the initiation of immune responses, we investigated changes in the expression levels of human leukocyte antigen (HLA)-DR, a major histocompatibility complex class II antigen, and of costimulatory molecules, such as CD54, CD80 and CD86, on monocytes following administration of macrolides. The expression of HLA-DR, CD54, CD80 and CD86 on cultured human peripheral mononuclear cells following stimulation by interferon (IFN)-γ and lipopolysaccharides (LPS) was analyzed using flow cytometry in the presence and absence of macrolides. The macrolides tested inhibited the expression of CD54, CD80 and CD86 on cultured monocytes following stimulation by IFN-γ and LPS, while the expression of HLA-DR on monocytes was not affected. Suppression of CD80 expression was the most significant among the costimulatory molecules tested and occurred in a dose-dependent manner. These results suggest that macrolides may downregulate the expression levels of costimulatory molecules, such as CD80, and may normalize the hyperimmune responses that may be responsible for the chronic intractable inflammation.

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We wanted to define allergen-specific antibodies that change due to specific immunotherapy. We conducted a study with grass pollen-allergic patients and compared allergen-specific IgE and IgG4 before and 5 months after the onset of immunotherapy. Twenty-seven patients were treated with a mixture of two grass species: Phleum pratense and Dactilis glomerata. Sera of patients were tested for IgE and IgG4 against four recombinant allergens (RA): rPhl p 1, 2, 5 and rBet v 2. Specific IgE and IgG4 to timothy and olive pollen were also evaluated. No change in total and specific IgE levels to RA was seen, except to rPhl p 5. We found a decrease in specific IgE levels to olive after immunotherapy. Ten of 10 patients with specific IgE against a single recombinant allergen or two RA showed the same pattern of sensitization before and after 5 months of immunotherapy and the administration of 4000 U/mL allergen extract. Interestingly, we found a significant increase in specific IgG4 to rBet v 2 and olive after grass immunotherapy. These results indicate that application of two grass species in immunotherapy may be sufficient to induce an IgE and IgG4 response to RA, grass and olive extracts. The observations in the present study indicate that monitoring of antibodies against RA is necessary to evaluate patients' pattern of sensitization and emphasize the need of component-resolved immunotherapy.

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Bronchial hyperresponsiveness (BHR) is a characteristic feature of bronchial asthma, yet its precise mechanism remains obscure. Human studies have demonstrated that T helper 2 cytokines, including interleukin (IL)-4 and IL-5, are involved in the development of airway inflammation and BHR. In the present study, we examined the role of IL-5 in the onset and aggravation of bronchial responsiveness to acetylcholine in two strains of IL-5 transgenic (Tg) mice, derived from the C3H/HeN and BALB/c strains. Three inhalations of antigen (ovalbumin) caused an increase in the number of eosinophils in bronchoalveolar lavage fluid (BALF) and a significant elevation in serum IgE in wild-type mice. In contrast with wild-type animals, systemic overproduction of IL-5 resulted in massive airway eosinophilia, especially around the peribronchi and perivascular regions of the tissues, after repeated antigen provocation. In C3H/HeN background IL-5 Tg mice, repeated antigen provocation did not induce BHR similar to that of wild-type mice. In contrast, antigen-induced BHR was observed in BALB/c- background mice, but there were no significant differences between the magnitude of BHR in wild-type and IL-5 Tg mice. Furthermore, antigen-induced IL-5 production in BALF was detected in both C3H/HeN and BALB/c mice and was elevated to a similar degree in both wild-type and IL-5 Tg mice. These findings suggest that systemic overproduction of IL-5 or airway eosinophilia is not, by itself, important in the development or aggravation of antigen-induced BHR in mice.

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Mast cells in the allergic nasal mucosa are reported to have higher concentrations of high affinity type I Fcε receptors (FcεRI) than mast cells in the non-allergic mucosa. IgE binds to FcεRI on mast cells. However, it is not clear whether different zones within the nasal mucosa harbor different quantities of IgE-bound mast cells. We stained IgE and mast cells from the nasal mucosa of patients with allergic and non-allergic rhinitis using a double-labeling technique with anti-IgE and antitryptase antibodies and compared the intensities of IgE staining of mast cells. In the allergic nasal mucosa, mast cells displayed more IgE staining in the lamina propria than in the epithelial layer. Mast cells displayed more IgE-staining in the allergic nasal mucosa than in the non-allergic mucosa. We speculate that the quantity of IgE that binds to mast cells depends on the quantity of IgE in nasal mucosal tissue.

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Milk allergy in a 1-year and 8-month-old boy is reported. At 1 year and 1 month of age, the patient presented with anaphylaxis, including erythema, which was initially localized to the contact site of the anterior chest, and wheezing accompanied by dyspnea, 5 min after contact with milk allergen through his atopic skin. These symptoms continued for 50 min. Seventy minutes after the disappearance of the initial erythema, the patient developed subsequent erythematous lesions distributed throughout the neck and head area that persisted for as long as 24 h. On another occasion, he also exhibited a pale face and generalized erythema immediately after an accidental oral ingestion of milk at the age of 1 year and 8 months. He had been unsettled for several hours when an intravenous steroid was administered. His serum IgE was 590 IU/mL and the radioallergosorbent test (RAST) scores against milk, α-lactoalbumin, β-lactoglobulin, casein and cheese were 5, 2, 3, 5 and 5, respectively. This is a rare case of a patient with milk allergy who fell into anaphylaxis following both cutaneous contact with and oral ingestion of the offending milk protein. Care should be taken with patients with food allergies because cutaneous contact with the offending food may cause adverse reactions, including anaphylaxis.

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A 63-year-old man was referred to our hospital because of a dry cough. His chest roentgenogram revealed ground-glass opacities and honeycomb formations bilaterally in the lower lung fields. Pulmonary function tests showed a depleted lung volume and decreased arterial oxygen tension. He was clinically diagnosed as having idiopathic interstitial pneumonia (IIP). A β₂-adrenoceptor agonist was administrated because the patient's symptoms improved after its inhalation. Following treatment with an oral β₂-adrenoceptor agonist, the dry cough disappeared, lung function tests remained unchanged and an improvement in arterial oxygen tension was observed. Although β₂-adrenoceptor agonist therapy does not improve disease activity or progression in patients with IIP, its use may mitigate symptoms associated with the disease.

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Olopatadine hydrochloride (olopatadine; KW-4679), (Z)-11-[(3-dimethylamino)propylidene]-6,11-dihydrodibenz[b,e]oxepin-2-acetic acid monohydrochloride, is an antiallergic drug with selective and potent histamine H₁ receptor antagonist activity. In the present study, we investigated the effect of olopatadine on the release of peptide leukotrienes (P-LT), potent inflammatory mediators, from human eosinophils. Human eosinophils were purified from venous blood of healthy donors by negative selection using the anti-CD16 antibody. When human eosinophils were stimu- lated with the calcium ionophore A23187 (1 µmol/L), the amount of P-LT release was approximately 1200 pg/ 10⁵ cells, while thromboxane (TX) B₂ release was under the detection limit (< 20 pg/10⁵ cells). Olopatadine inhibited the A23187-induced P-LT release from human eosinophils with an IC₅₀ of 4.5 µmol/L. Ketotifen also inhibited this reaction with an IC₅₀ of 39.4 µmol/L. The inhibitory effect of olopatadine on the P-LT release may contribute to the antiallergic efficacy of this drug.

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