Reiko Kishikawa

Identification of Italian cypress (Cupressus sempervirens) pollen allergen Cup s 3 using homology and cross-reactivity

BACKGROUND The prevalence of seasonal allergic diseases of the upper airways is increasing in industrialized countries. The Cupressaceae are important causes of pollinosis, particularly in Europe. OBJECTIVE To determine whether the pollen from Cupressus sempervirens (Italian cypress) contains a pathogenesis-related group 5 (PR-5) protein, similar to that found in other allergenic Cupressaceae pollens. METHODS Messenger RNA was purified from Italian cypress pollen, and complementary DNA (cDNA) was synthesized. cDNAs for PR-5 proteins were amplified by polymerase chain reaction and extended by rapid amplification of cDNA ends methods. Recombinant Cup s 3 was expressed in Escherichia coli as a fusion protein. Inhibition enzyme-linked immunosorbent assays were used to test the allergenicity of Cup s 3. RESULTS Three cDNAs were cloned. These clones had approximately 95% identity to Jun a 3 and Cup a 3. Recombinant Cup s 3.0102 maltose-binding protein inhibited the IgE from most patients from binding to an extract of Italian cypress. The extent of inhibition suggested that antibodies to Cup s 3 were a prominent component of the IgE response to Italian cypress pollen. CONCLUSION Cup s 3, an allergen of Italian cypress pollen, was identified based on cross-reactivity and homology with other pollen PR-5 proteins, despite an apparently low level of protein expression. Variations in the content of Cup s 3 in the pollen from different regions or trees should be considered in the choice of extracts for diagnosis and specific immunotherapy for Italian cypress pollen hypersensitivity.

Serum high-sensitivity C-reactive protein can be an airway inflammation predictor in bronchial asthma.

It is controversial that the serum high-sensitivity C-reactive protein (hs-CRP) can be a useful marker of airway inflammation in bronchial asthma, because various factors have been reported to affect hs-CRP levels. We conducted a study in patients with mild bronchial asthma without complications to determine whether hs-CRP is a useful indicator of airway inflammation. The induced sputum cell differentiation, respiratory function tests, bronchial hyperresponsiveness tests, and hs-CRP measurement were performed in the subject population consisted of 40 healthy volunteers and 45 patients with bronchial asthma. The log-transformed (log) serum hs-CRP level was higher in asthmatic patients compared with healthy controls (2.49 ± 0.41 versus 2.21 ± 0.39; p = 0.002). A receiver-operating characteristic (ROC) analysis showed a sensitivity of 0.69 and specificity of 0.70 for a log serum hs-CPR of 2.3 to distinguish asthmatic patents from healthy controls. The log serum hs-CRP level negatively correlated with forced volume in 1 second (FEV1.0)%pred (r = -0.31, p = 0.04), positively correlated with sputum eosinophils (r = 0.34, p = 0.02), negatively correlated with sputum macrophages (r = -0.56, p < 0.001), and did not correlate with log 20% fall in FEV1.0 (PC20) (r = -0.09, p = 0.56). A multiple regression analysis revealed that the log serum hs-CRP concentration significantly correlated with eosinophils (p = 0.019) and neutrophils (p = 0.042) in the sputum, respectively. Serum hs-CRP may be a useful marker of airway inflammation in nonsmoking asthmatic patients without complications, such as heart disease, hypertension, hyperlipidemia, chronic obstructive pulmonary disease, or infection.

The Fractional Exhaled Nitric Oxide and Serum High Sensitivity C-Reactive Protein Levels in Cough Variant Asthma and Typical Bronchial Asthma

BACKGROUND Fractional exhaled nitric oxide (FeNO) is known to be a good marker of airway eosinophilic inflammation in bronchial asthma. Recently, serum high sensitivity C-reactive protein (hs-CRP) has been shown to be also useful to detect the airway inflammation. METHODS Newly diagnosed 90 cough variant asthma and 92 bronchial asthma patients were enrolled. FeNO, serum hs-CRP, pulmonary function tests, bronchial hyperresponsiveness, IgE and sputum eosinophils ratio were compared. Ninety healthy control subjects were set for FeNO and serum hs-CRP normal range reference. We have compared the clinical utilities of FeNO and serum hs-CRP to differentiate bronchial asthma and cough variant asthma. RESULTS FeNO was significantly higher in bronchial asthma (92.6 ± 85.5ppb) than in cough variant asthma (35.6 ± 43.3; p < 0.001) and both were significantly higher than normal range (18.0 ± 6.4, p < 0.001, respectively), and in differentiating between the two groups showed a sensitivity of 0.69 and a specificity of 0.73 at the cutoff value of 28 ppb. Serum hs-CRP did not differ significantly between bronchial asthma (723 ± 1162ng/ml) and cough variant asthma (558 ± 758) even if both were significantly higher than normal range (345 ± 401, p < 0.01 and p < 0.05 respectively). CONCLUSIONS FeNO is more useful than serum hs-CRP in differentiating patients with bronchial asthma from those with cough variant asthma, and healthy persons.

Influence of cigarette smoking on airway inflammation and inhaled corticosteroid treatment in patients with asthma.

BACKGROUND Cigarette smoking induces neutrophilic airway inflammation and relative resistance to inhaled corticosteroids (ICS). OBJECTIVE We evaluated the influence of cigarette smoking on airway inflammation in patients with asthma and also compared the effect of ICS between smoking and nonsmoking in patients with asthma. METHODS Smokers with asthma (n = 81) and nonsmokers with asthma (n = 52) were recruited for the study. We examined lung function, fractional exhaled nitric oxide (FeNO) concentration, induced sputum, and acetylcholine inhalation before and 6 months after inhaling budesonide at 800 μg/day. Thirty-four healthy volunteers were included as controls. RESULTS Smokers with asthma showed a lower forced expiratory volume in 1 second (FEV1) to forced volume capacity (FVC) ratio (p < 0.05), a lower FeNO (p < 0.01), a lower eosinophil proportion (p < 0.05), and a higher neutrophil proportion (p < 0.05) in induced sputum than nonsmokers with asthma. Bronchial hyperresponsiveness (the provocative concentration of acetylcholine [Ach] that produced a 20% fall in FEV1 [PC20-Ach]) was increased in smokers with asthma compared with nonsmokers with asthma (p < 0.05). Both smokers with asthma and nonsmokers with asthma exhibited more prominent airway obstruction, a higher FeNO, and a higher percentage of sputum eosinophils than the controls (p < 0.05 to p < 0.001, each). After 6 months of treatment with inhaled budesonide at 800 μg/day, the improvement in lung function (FEV1 to FVC ratio, flow at 50% forced vital capacity [V50% predicted] and flow at 25% forced vital capacity [V25% predicted]), the eosinophil proportion in induced sputum and PC20-Ach were lower in smokers with asthma than nonsmokers with asthma (p < 0.05). CONCLUSION Smokers with asthma showed neutrophilic airway inflammation in addition to eosinophilic inflammation, and cigarette smoking impaired the efficacy of ICS treatment in mild-to-moderate asthma. These findings have important implications for the management of patients with asthma and who smoke.

Association of matrix metalloproteinase 8 genetic polymorphisms with bronchial asthma in a Japanese population.

Asthma has a strong genetic component. The final disease phenotype results from complex interactions between environment and multiple genes of small-to-modest effects. We investigated whether the polymorphism in genes encoding inflammatory mediators and cytokines is important for solving the onset and progression of asthma. We investigated whether 31 single nucleotide polymorphisms (SNPs) in genes encoding cytokines or monokines (interleukin [IL]-5R, matrix metalloproteinase [MMP] 8, beta2 adrenergic receptor, cytotoxic T-lymphocyte-associated antigen 4, IL-3, C-reactive protein, cytochrome P450 (CYP) 2C9, CYP3A4, a disintegrin and metalloproteinase [ADAM] 33, cysteinyl leukotriene receptor [CysLTR] 1, CysLTR2, eosinophilic cationic protein, glucocorticoid receptor, and leukotriene A 4 hydrolase) are related to asthma development in 206 Japanese bronchial asthma patients and 127 healthy controls. Using multifactor dimensionality reduction (MDR), we identified rs17099451 in MMP8, using a single locus model, with a mean cross-validation of 87.0%. Using a two-locus model, combinations of MMP8 and rs44707 in ADAM33, and MMP8 and rs40401 in IL-3, were identified, with mean cross-validation consistencies reaching 45.0%. Of the SNPs selected by the MDR method, rs17099451 in MMP8 and rs40401 in IL-3 were regarded as the most significant results in a 2 × 2 dominant model analysis. The finding that an MMP8 allele was most strongly related to asthma development indicates that metalloproteinase function is crucial to the airflow limitation process involved in this disease.

Lung sound analysis helps localize airway inflammation in patients with bronchial asthma

Purpose Airway inflammation can be detected by lung sound analysis (LSA) at a single point in the posterior lower lung field. We performed LSA at 7 points to examine whether the technique could identify the location of airway inflammation in patients with asthma. Patients and methods Breath sounds were recorded at 7 points on the body surface of 22 asthmatic subjects. Inspiration sound pressure level (ISPL), expiration sound pressure level (ESPL), and the expiration-to-inspiration sound pressure ratio (E/I) were calculated in 6 frequency bands. The data were analyzed for potential correlation with spirometry, airway hyperresponsiveness (PC20), and fractional exhaled nitric oxide (FeNO). Results The E/I data in the frequency range of 100–400 Hz (E/I low frequency [LF], E/I mid frequency [MF]) were better correlated with the spirometry, PC20, and FeNO values than were the ISPL or ESPL data. The left anterior chest and left posterior lower recording positions were associated with the best correlations (forced expiratory volume in 1 second/forced vital capacity: r=−0.55 and r=−0.58; logPC20: r=−0.46 and r=−0.45; and FeNO: r=0.42 and r=0.46, respectively). The majority of asthmatic subjects with FeNO ≥70 ppb exhibited high E/I MF levels in all lung fields (excluding the trachea) and V50%pred <80%, suggesting inflammation throughout the airway. Asthmatic subjects with FeNO <70 ppb showed high or low E/I MF levels depending on the recording position, indicating uneven airway inflammation. Conclusion E/I LF and E/I MF are more useful LSA parameters for evaluating airway inflammation in bronchial asthma; 7-point lung sound recordings could be used to identify sites of local airway inflammation.

Trigger of bronchial hyperresponsiveness development may not always need eosinophilic airway inflammation in very early stage of asthma.

Background Cough variant asthma (CVA), a suggested precursor of standard bronchial asthma (SBA), is characterized by positive bronchial hyperresponsiveness (BHR) and a chronic cough response to bronchodilator that persists for >8 weeks. Objective Airway inflammation, BHR, and airway obstructive damage were analyzed to assess whether CVA represents early or mild-stage SBA. Methods Patients with newly diagnosed CVA (n = 72) and SBA (n = 84) naive to oral or inhaled corticosteroids and without exacerbated asthma were subjected to spirometry, impulse oscillometry, BHR tests, sputum induction, and fractional exhaled nitric oxide measurements. Results In the patients with CVA, spirometry demonstrated higher forced expiratory volume in 1 second (FEV1) to forced vital capacity ratio, FEV1 percent predicted, flow volume at 50% of vital capacity % predicted, and flow volume at 25% of vital capacity % predicted values, and impulse oscillometry demonstrated lower R5–Z20, AX, and Fres, and higher X5 values. In addition, the fractional exhaled nitric oxide and sputum eosinophil numbers were lower and the PC20 was higher than in patients with moderate SBA. However, these factors were similar in the patients with CVA and in the patients with intermittent mild SBA. A significantly smaller proportion of the patients with CVA had increased sputum eosinophils than the patients with intermittent mild SBA (p < 0.0001). However, interestingly, among the patients with CVA, no significant differences in the PC20 values were found between the patients with and those without increased sputum eosinophils. Conclusions All measures of central and peripheral airway obstruction, eosinophilic inflammation, and airway hyperresponsiveness in patients with CVA were milder than in patients with moderate SBA but were similar to those of patients with intermittent mild SBA. In CVA, the BHR was not affected by airway eosinophilic inflammation, which indicated that the very early development of BHR may not always need airway eosinophilic inflammation.

Longitudinal Investigation on Allergenic Conifer Pollen in Japan for Successful Prevention and Treatment against Japanese Cedar Pollinosis

Longitudinal Investigation on Allergenic Conifer Pollen in Japan for Successful Prevention and Treatment against Japanese Cedar Pollinosis In Japan allergic people has been recognized increasing according to the increase of Japanese cedar, Cryptomeria japonica (JC) pollinosis since first reported in 1964. We have detected allergenic airborne pollen when Japanese government has begun to investigate the background and treatment on JC pollinosis as a serious social problem since July1986. Urgently the gravitational method with Durham’s sampler has been adopted to know huge number of JC and cypress family, Cupressaceae family (Cypress) pollen grains as causative agent. The area of national artificial forest of over 31 years old has become about 7 times from1970. We have estimated annual fluctuation of Japanese important allergenic pollen, with climatic factor during about 30 years for successful treatment of pollinosis in near future. There are institutions in fifteen locations monitoring airborne pollen by Durham’s sampler in Japan between the latitude of 30 to 40 degrees north. Airborne pollen samples were collected at each institute and were sent to our hospital on a daily basis. We counted pollen grains per cm2 through microscope, classifying and summarizing them. From 1986 to 2014 we have referred to the change in meteorological condition at the close to 9 of the pollen monitoring locations and estimated statically. JC and Cypress pollen counts have shown huge but remarkable annual fluctuation and they have been increasing gradually according to over 31 y-old conifer planting area expanding. The mean temperature in summer time, JC male flower formation, has a significant correlation with next year pollen counts. The pollination season has started earlier in north and later in central Japan gradually through a changing on mean temperature. In Japan allergenic conifer pollen counts have been increasing because of over 31 y-old conifer plantations area increase through the climate change about 30 years. Moreover, it has changed the start of pollination season.

The Relationship between Symptom Flare of Atopic Dermatitis and Airborne Japanese Cedar and Cypress Pollen Counts: A Self-Scoring Diary Study

Background. With an increase in Japanese cedar and cypress (JC) pollinosis, the relationship between JC pollen and atopic dermatitis (AD) has been studied. Some reports suggest that JC pollen can be one exacerbating factor for AD, but there has been no report that discusses JC pollen counts relating to AD symptom flare although actual airborne JC pollen counts can widely fluctuate throughout the pollen season. Objective. The relationship between symptom flare of AD and airborne JC pollen counts was examined. Methods. We monitored JC pollen counts in real time and divided the counts into low and high level. We then analyzed self-scored “itch intensity” recorded by 14 AD patients through a self-scoring diary. Results. Among the 14 patients, 7 had significantly higher itch intensity while the pollen counts were high. Conclusion. Even during the pollen season, actual airborne pollen counts can widely fluctuate. Our study suggested that symptom flare of AD could be influenced by the actual pollen counts.

Lung Sound Analysis and the Respiratory Cycle Dependence of Impulse Oscillometry in Asthma Patients

Objective A lung sound analysis (LSA) is useful for detecting airway inflammation and obstruction in patients with asthma. To elucidate the mechanism of LSA, we investigated the relationship between the exhalation-to-inhalation sound pressure ratio in the low frequency range between 100 and 195 Hz (E/I LF) and the respiratory cycle dependence of impulse oscillometry (IOS) parameters. Methods Asthma patients underwent IOS [resistance of the respiratory system at 5 Hz (R5) and 20 Hz (R20), the reactance area (AX), resonant frequency of reactance (Fres), and reactance of the respiratory system at 5 Hz (X5)], spirography, and an LSA. The correlation between the LSA-derived E/I LF values and the respiratory cycle dependence of the IOS parameters was analyzed. Patients Thirty-four patients with mild to moderate bronchial asthma, who had not received oral or inhaled corticosteroids and who had no episodes of rumbling or wheezing were examined. Results The E/I LF value was significantly correlated with the differences of the R5 and R5-R20 values between exhalation and inhalation (p=0.035 and p=0.050) in a multivariate analysis. Conclusion E/I LF appears to be an index that expresses the respiratory cycle dependence of asthma as well as IOS.