Clinical significance of a basophil activation test for Japanese beekeepers naturally sensitized to honey bee venom.

BACKGROUND: The basophil activation test (BAT) has high sensitivity and specificity for diagnosing Hymenoptera venom allergy and is useful for predicting the clinical sensitivity of bee venom-allergic patients after venom immunotherapy. Patients sensitized to Hymenoptera venom are at risk for systemic reactions (SRs) to subsequent stings. Therefore, a tool that can predict the occurrence of SRs and the severity of Hymenoptera stings is needed. OBJECTIVE: We performed BATs on Japanese beekeepers naturally sensitized to honey bee venom (HBV) and analyzed the positive threshold concentration for the occurrence of SRs following honey bee stings (HBS). METHODS: Sixty-one beekeepers were interviewed and blood samples were taken. Data including history of HBS and the occurrence and severity of SRs to HBS were recorded. Blood samples were exposed to HBV-specific IgE antibodies (sIgE) and BAT was performed. Participants with HBV-sIgE ≥ class 1 were considered sensitized to HBV. The positive threshold for BAT scored as 0.0001, 0.001, 0.01, 0.1, and 1 µg/ml was classified as classes 5, 4, 3, 2, and 1, respectively. Samples negative at 1 µg/ml were classified as class 0. RESULTS: About 40% of beekeepers with a positive BAT threshold ≤ 0.1 µg/ml had SRs after HBS. The mean score of the BAT positivity threshold for beekeepers who developed SRs was significantly lower than that for beekeepers with no history of SRs (2.6 ± 0.8 vs 1.4 ± 1.1, P < 0.01). CONCLUSION: Analysis of the positive threshold of BAT in Japanese beekeepers naturally sensitized to HBV may be a useful tool for predicting the occurrence of SRs.

Differences and similarities in cytokine profiles of macrophage activation syndrome in systemic lupus erythematosus and adult-onset Still's disease.

To clarify the differences and similarities in the cytokine profiles of macrophage activating syndrome (MAS) between systemic lupus erythematosus (SLE) and adult-onset Still's disease (AOSD). The study participants included 9 patients with MAS-SLE, 22 with non-MAS-SLE, 9 with MAS-AOSD, and 13 with non-MAS-AOSD. Serum cytokine levels were measured using a multiplex bead assay. Cytokine levels were compared between patients with SLE and AOSD with/without MAS. Moreover, cytokine patterns were examined using principal component analysis (PCA) and cluster analysis. IL-6, IL-8, IL-18, and TNF-α levels were elevated in patients with SLE and AOSD. IFN-α levels were elevated in SLE, whereas IL-1ß and IL-18 levels were elevated in AOSD. In SLE, IFN-α and IL-10 levels were higher in MAS than in non-MAS and controls. PCA revealed distinctive cytokine patterns in SLE and AOSD, SLE with IFN-α and IP-10, AOSD with IL-1ß, IL-6, and IL-18, and enhanced cytokine production in MAS. PCA and cluster analysis showed no differences in cytokine patterns between the MAS and non-MAS groups. However, serum ferritin levels were correlated with IFN-α levels in SLE. Cytokine profiles differed between SLE and AOSD but not between MAS and non-MAS. MAS is induced by the enhancement of underlying cytokine abnormalities rather than by MAS-specific cytokine profiles. Type I IFN may be involved in MAS development in patients with SLE.

Effect of inhaled corticosteroids on bone mineral density in patients with asthma.

BACKGROUND: Inhaled corticosteroids (ICS) are a safe treatment for asthma. However, at higher doses, ICS use has been reported to inhibit adrenocortical function. OBJECTIVE: This study aimed to evaluate the effect of ICS on bone mineral density (BMD) in adult patients with asthma. METHODS: Ultrasonic bone densitometry was performed in 40 patients (14 men, 26 women, mean age 61.2 years, mean duration of asthma 6.19 years) who were receiving ICS for asthma, and the whole bone density, thickness of cortical bone, and density of cancellous bone of the radius was measured. The age-matched mean was set as 100%. Lifetime cumulative dose of ICS was calculated using all past prescriptions. RESULTS: No significant correlations were observed between lifetime cumulative ICS dose and whole bone density (r² = 0.011), cortical bone thickness (r² = 0.022), and cancellous bone density (r² = 0.004). No significant differences were observed between lower and higher lifetime cumulative ICS dose among these BMD parameters (104% vs 97%, 103% vs 99%, and 106% vs 91%, respectively). No significant correlations or differences in lifetime cumulative ICS dose were observed by asthma severity, asthma duration, and pulmonary function. Also, serum markers of bone metabolism showed no significant correlations or differences with lifetime cumulative ICS dose. CONCLUSIONS: In the entire study population, long-term ICS use was safe and was not associated with an increased risk of osteoporosis.

Changing patterns of pulmonary abnormalities in rheumatoid arthritis.

BACKGROUND: As a first step in identifying the developmental pathways of pulmonary abnormalities in rheumatoid arthritis (RA), we sought to determine the existing and changing patterns of pulmonary abnormalities. METHODS: We conducted a retrospective cohort study of consecutive patients with RA who underwent high-resolution computed tomography before and during biologic therapy. The presence of 20 pulmonary abnormalities and the changes in those abnormalities were recorded. Patterns of pre-existing and changing abnormalities were examined via cluster analysis, and their relationship was also assessed using the Kaplan-Meier method and log-rank test. RESULTS: A total of 208 subjects were included. Pulmonary abnormalities were observed in 70% of patients: 39% had interstitial lung disease, and 55% had airway disease (AD). Several different pulmonary abnormalities were commonly found to co-exist in several patterns in the same patient. In most patients with pulmonary abnormalities, AD was present alone or in combination with other abnormalities. During the observation period (mean 3.2 years), 172 pulmonary abnormalities had changed in 91 patients: 115 pulmonary abnormalities newly emerged, whereas 42 worsened and 25 demonstrated improvement. Pulmonary abnormalities changed in several patterns. Correlations were observed between pre-existing and new/worsening abnormalities at individual and regional levels, such as new ground-glass opacity (GGO) and pre-existing AD, small nodular patterns, and honeycombing. AD was a possible initial abnormality. CONCLUSIONS: Pulmonary abnormalities occurred and changed in several patterns, which suggests the existence of developmental pathways of pulmonary abnormalities. AD may play an important role in the development of these abnormalities, including GGO.

The role of anti-elastin antibodies in a mouse model of asthma.

BACKGROUND: The role of anti-elastin antibody (Ab) in the lung is unclear, although they may be involved in chronic obstructive pulmonary disease (COPD). Recently, increased anti-elastin Ab levels were reported in asthma. OBJECTIVE: To elucidate the role of anti-elastin Ab in asthma, we created a murine asthma model. Anti-elastin Ab in the airway was neutralized by intratracheal administration of elastin peptide, and the inhibitory effects of anti-elastin Ab on airway remodeling were evaluated. METHODS: BALB/c mice were immunized with ovalbumin (OVA) on days 0 and 14. After immunization, the mice received booster OVA via inhalation twice per week for 9 weeks, and bronchoalveolar lavage fluid (BALF) and lung tissues were evaluated. RESULTS: In lung tissues, airway remodeling occurred after 9 weeks of OVA sensitization. Peak levels of anti-elastin Ab and eosinophils in BALF were detected after 3 weeks of OVA sensitization. Anti-elastin Ab and eosinophil levels in BALF were significantly reduced after 3 weeks by the neutralization of anti-elastin Ab. Peak transforming growth factor-ß1 levels in BALF were detected at 3 weeks after OVA sensitization and were significantly reduced by the neutralization of anti-elastin Ab. Airway remodeling in lung tissues was also significantly inhibited by the neutralization of anti-elastin Ab. CONCLUSIONS: In our murine asthma model, anti-elastin Ab was recruited to the airway by OVA-induced allergic inflammation. Airway remodeling was inhibited by the neutralization of anti-elastin Ab. Anti-elastin Ab may contribute to the progression of airway remodeling.

Elevation of anti-elastin antibody in patients with asthma.

BACKGROUND: It is often difficult to differentiate between asthma and chronic obstructive pulmonary disease (COPD), and useful biomarkers are needed for accurate diagnosis. OBJECTIVE: We evaluated anti-elastin antibody to identify useful biomarkers for differentiating between a diagnosis of asthma and COPD. METHODS: Patients with asthma (male to female ratio = 10/13; mean age, 67.3 years), COPD (16/0; 74.8 years) and controls (8/4; 72.3 years) were enrolled. Samples from sputum and serum were collected and levels of anti-elastin Ab were measured. RESULTS: The levels of anti-elastin Ab in sputum were significantly higher in asthma (11.4 ± 7.16 µg/mL) than in COPD (5.82 ± 5.16 µg/mL; P < 0.01), and serum levels in asthma (67.4 ± 29.7 µg/mL) were also significantly higher than in COPD or controls (45.0 ± 12.8 µg/mL; P < 0.05, 38.6 ± 10.4 µg/mL; P < 0.01, respectively). Anti-elastin Ab in sputum showed a positive correlation with smoking in asthma (r2 = 0.218, P < 0.05). However, no significant differences were observed in the levels of anti-elastin Ab and eosinophils, asthma phenotypes, inhaled corticosteroids, or severity in patients with asthma. Elastin was strongly expressed under the airway basement membrane in asthma compared with COPD or the healthy control. CONCLUSIONS: Anti-elastin Ab in sputum could be a useful biomarker for COPD and asthma in ever-smokers. In asthma, anti-elastin Ab was recruited to the airways by both airway allergic inflammation and smoking, and it may contribute to the progression of airway remodeling via autoimmune inflammation, but not emphysema, in COPD.

Emergency Treatment of Anaphylaxis in Japanese Beekeepers.

Objectives: Honeybee stings often lead to anaphylactic shock. We surveyed Japanese beekeepers to examine whether adrenaline auto-injectors are properly used after honeybee stings.Methods: We contacted representatives of the Japanese Beekeeping Association in all 47 prefectures for assistance distributing allergist-developed questionnaires. Representatives in 33 prefectures distributed questionnaires to their members and we received valid responses from 826 beekeepers.Results: Adrenaline auto-injectors had been prescribed to only 46 of the 826 participants (5.6%) to prevent systemic reaction (SR) to honeybee stings. Of the 33 beekeepers who experienced a honeybee sting after adrenaline auto-injector prescription, 16 (48.5%) developed SRs; 9 of these 16 (56.3%) were treated with an adrenaline auto-injector.Conclusions: Japanese beekeeping organizations should consider encouraging medical institutions to prescribe adrenaline auto-injectors. Furthermore, physicians and other health care workers should better educate beekeepers and others who have been prescribed an adrenaline auto-injector in order to improve compliance and raise awareness of the risk posed by SRs.

Th2 cell differentiation from naive CD4+ T cells is enhanced by autocrine CC chemokines in atopic diseases.

BACKGROUND: Chemokines are involved not only in regulating leucocyte recruitment, but also in other activities. However, functions other than cell recruitment remain poorly understood. We have already shown that the production of CC chemokine ligand (CCL)17 and CCL22 by antigen-stimulated naïve CD4+  T cells was higher in asthmatic patients than in healthy controls. However, the role of these chemokines in stimulated naïve CD4+ T cells remains unclear. OBJECTIVE: To clarify the biological function of CCL17 and CCL22 on naïve CD4+ T, we examined effects of these two chemokines on naïve CD4+ T cells expressing CC chemokine receptor (CCR)4 (a receptor for CCL17 and CCL22) during differentiation of Th2 cells in asthmatic patients as allergic subjects. METHODS: Naïve CD4+ T cells were prepared from healthy controls and patients with asthma. We analysed effect of CCL17 and CCL22, and blocking their receptor on differentiation of Th2 cells. RESULTS: Production of CCL17 and CCL22 by activated naive CD4+ T cells under Th2 condition was much more in asthmatic patients than in healthy controls. Proliferation and survival of the Th2 differentiating cells and restimulation-induced IL-4 production were much greater in asthmatic patients than in healthy controls. These cell biological phenomena were inhibited by blockade of CCR4. The biological effects of exogenous CCL17 and CCL22 were apparently observed in both healthy controls and asthmatic patients. The effectiveness of these chemokines on naïve CD4+ T cells from healthy controls was stronger than those from asthmatic patients. We found that thymic stromal lymphopoietin (TSLP), a Th2 promoting chemokine, is involved in the activation of CD4+ naïve T cells via production of CCL17 and CCL22. CONCLUSIONS AND CLINICAL RELEVANCE: These data suggest that CCL17 and CCL22 produced by TSLP-primed naïve CD4+ T cells in asthma might contribute to an increase in Th2 cells via autocrine loops.

Tofacitinib for refractory interstitial lung diseases in anti-melanoma differentiation-associated 5 gene antibody-positive dermatomyositis.

Objective: We aimed to determine the outcome of combination therapy with tofacitinib (TOF) in a case series of refractory rapidly progressive interstitial lung disease (ILD) associated with anti-melanoma differentiation-associated 5 gene (MDA5) antibody-positive (Ab+) DM. Patients who had poor prognostic factors and failed to respond to immunosuppressive therapy were selected for TOF treatment. Methods: Five patients with anti-MDA5 Ab+ DM-ILD who failed to respond to triple therapy with high dose glucocorticoids, CSA and CYC were given additional TOF (10 mg/day). To identify the poor prognostic factors, data from 15 consecutive patients (seven survived and eight died) with anti-MDA5 Ab+ DM-ILD before induction of TOF were analysed. Results: Three poor prognostic factors were identified: serum ferritin level >1000 ng/ml before therapy; ground-glass opacities in all six lung fields before therapy; and worsening of pulmonary infiltrates during therapy. All six patients who had all of the three factors and received triple therapy died before TOF therapy. There were five patients who had all of the three prognostic factors and failed to respond to triple therapy, but were able to receive the combination therapy with TOF; among them, three survived and two died. The survival rate of patients who received TOF was significantly better than that of the historical controls with immunosuppressive therapy before TOF. The patients who received TOF experienced complicated adverse events, particularly viral infection. Conclusion: Combination therapy with TOF might have the potential to control refractory anti-MDA5 Ab+ DM-ILD.

Effect of adjusting the combination of budesonide/formoterol on the alleviation of asthma symptoms.

BACKGROUND: The combination of budesonide + formoterol (BFC) offers the advantages of dose adjustment in a single inhaler according to asthma symptoms. We analyzed the relationship between asthma symptoms in terms of peak expiratory flow (PEF) and dose adjustment by the patient. METHODS: Twenty-eight patients with asthma who used BFC for alleviation of their symptoms (12 men, 16 women; 60 years old) were instructed that the inhaled BFC dose could be increased to a maximum of 8 inhalations per day according to symptom severity. Patients measured and recorded PEF every morning and evening in their asthma diary along with their symptoms and the dose of drugs taken. RESULTS: Sixteen of the 28 patients increased their dose for asthma symptoms. The time to recovery from the asthma symptoms was significantly shorter when cough was the only symptom present compared with dyspnea or wheeze (1.4 vs. 5.3 or 6.6 days, p < 0.05) and when they had only one symptom compared with two or three symptoms (1.3 vs. 5.7 or 10.5, p < 0.01). The relationship between PEF (% of personal best) when the dose was increased (Y) and the days for the increased dose to achieve a PEF greater than PEF in the symptom-free state (X) was determined to be Y = - 0.591X + 89.2 (r2 = 0.299, p < 0.001). CONCLUSION: As a guide for increasing the BFC dose when patients with mild asthma have asthma symptoms, the dose should be increased when cough is present or PEF is decreased to 88.9% (i.e., X = 0.5).

Allergic TH2 Response Governed by B-Cell Lymphoma 6 Function in Naturally Occurring Memory Phenotype CD4+ T Cells.

Transcriptional repressor B-cell lymphoma 6 (Bcl6) appears to regulate TH2 immune responses in allergies, but its precise role is unclear. We previously reported that Bcl6 suppressed IL-4 production in naïve CD4+ T cell-derived memory TH2 cells. To investigate Bcl6 function in allergic responses in naturally occurring memory phenotype CD4+ T (MPT) cells and their derived TH2 (MPTH2) cells, Bcl6-manipulated mice, highly conserved intron enhancer (hcIE)-deficient mice, and reporter mice for conserved noncoding sequence 2 (CNS2) 3' distal enhancer region were used to elucidate Bcl6 function in MPT cells. The molecular mechanisms of Bcl6-mediated TH2 cytokine gene regulation were elucidated using cellular and molecular approaches. Bcl6 function in MPT cells was determined using adoptive transfer to naïve mice, which were assessed for allergic airway inflammation. Bcl6 suppressed IL-4 production in MPT and MPTH2 cells by suppressing CNS2 enhancer activity. Bcl6 downregulated Il4 expression in MPTH2 cells, but not MPT cells, by suppressing hcIE activity. The inhibitory functions of Bcl6 in MPT and MPTH2 cells attenuated allergic responses. Bcl6 is a critical regulator of IL-4 production by MPT and MPTH2 cells in TH2 immune responses related to the pathogenesis of allergies.

Molecular biological analysis in a patient with multiple lung adenocarcinomas.

The utility of molecular biological analysis in lung adenocarcinoma has been demonstrated. Herein we report a rare case presenting as multiple lung adenocarcinomas with four different EGFR gene mutations detected in three lung tumors. After opacification was detected by routine chest X-ray, the patient, a 64-year-old woman, underwent chest computed tomography which revealed a right lung segment S4 ground-glass nodule (GGN). Follow-up computed tomography revealed a 42 mm GGN nodule with a 26 mm nodule (S6) and a 20 mm GGN (S10). Histopathology of resected specimens from the right middle and lower lobes revealed all three nodules were adenocarcinomas. Four EGFR mutations were detected; no three tumors had the same mutations. Molecular biological analysis is a promising tool for the diagnosis of primary tumors in patients with multiple lung carcinomas of the same histotype, enabling appropriate treatment.

Development of chronic allergic responses by dampening Bcl6-mediated suppressor activity in memory T helper 2 cells.

Mice deficient in the transcriptional repressor B-cell CLL/lymphoma 6 (Bcl6) exhibit similar T helper 2 (TH2) immune responses as patients with allergic diseases. However, the molecular mechanisms underlying Bcl6-directed regulation of TH2 cytokine genes remain unclear. We identified multiple Bcl6/STAT binding sites (BSs) in TH2 cytokine gene loci. We found that Bcl6 is modestly associated with the BSs, and it had no significant effect on cytokine production in newly differentiated TH2 cells. Contrarily, in memory TH2 (mTH2) cells derived from adaptively transferred TH2 effectors, Bcl6 outcompeted STAT5 for binding to TH2 cytokine gene loci, particularly Interleukin4 (Il4) loci, and attenuated GATA binding protein 3 (GATA3) binding to highly conserved intron enhancer regions in mTH2 cells. Bcl6 suppressed cytokine production epigenetically in mTH2 cells to negatively tune histone acetylation at TH2 cytokine gene loci, including Il4 loci. In addition, IL-33, a pro-TH2 cytokine, diminished Bcl6's association with loci to which GATA3 recruitment was inversely augmented, resulting in altered IL-4, but not IL-5 and IL-13, production in mTH2 cells but no altered production in newly differentiated TH2 cells. Use of a murine asthma model that generates high levels of pro-TH2 cytokines, such as IL-33, suggested that the suppressive function of Bcl6 in mTH2 cells is abolished in severe asthma. These findings indicate a role of the interaction between TH2-promoting factors and Bcl6 in promoting appropriate IL-4 production in mTH2 cells and suggest that chronic allergic diseases involve the TH2-promoting factor-mediated functional breakdown of Bcl6, resulting in allergy exacerbation.

Transcriptional repression of p27 is essential for murine embryonic development.

The Nczf gene has been identified as one of Ncx target genes and encodes a novel KRAB zinc-finger protein, which functions as a sequence specific transcriptional repressor. In order to elucidate Nczf functions, we generated Nczf knockout (Nczf-/-) mice. Nczf-/- mice died around embryonic day 8.5 (E8.5) with small body size and impairment of axial rotation. Histopathological analysis revealed that the cell number decreased and pyknotic cells were occasionally observed. We examined the expression of cell cycle related genes in Nczf-/- mice. p27 expression was increased in E8.0 Nczf-/- mice compared to that of wild type mice. Nczf knockdown by siRNA resulted in increased expression of p27 in mouse embryonic fibroblasts (MEFs). Furthermore, p27 promoter luciferase reporter gene analysis confirmed the regulation of p27 mRNA expression by Nczf. Nczf-/-; p27-/- double knockout mice survived until E11.5 and the defect of axial rotation was restored. These data suggest that p27 repression by Nczf is essential in the developing embryo.

Improved sensitivity to venom specific-immunoglobulin E by spiking with the allergen component in Japanese patients suspected of Hymenoptera venom allergy.

BACKGROUND: Ves v 5 and Pol d 5, which constitute antigen 5, are recognized as the major, most potent allergens of family Vespidae. Several studies have reported the diagnostic sensitivity of the novel recombinant (r)Ves v 5 and rPol d 5 allergens in routine clinical laboratory settings by analyzing a group of Vespula and Polistes venom-allergic patients. In this study, we analyzed the sensitivity to venom specific (s)IgE by spiking with rVes v 5 and rPol d 5 in Japanese patients suspected of Hymenoptera venom allergy. METHODS: Subjects were 41 patients who had experienced systemic reactions to hornet and/or paper wasp stings. Levels of serum sIgE against hornet and paper wasp venom by spiking with rVes v 5 and rPold d 5, respectively, as improvement testing, compared with hornet and paper wasp venom, as conventional testing, were measured by ImmunoCAP. RESULTS: Of the 41 patients, 33 (80.5%) were positive (≥0.35 UA/ml) for hornet and/or paper wasp venom in conventional sIgE testing. sIgE levels correlated significantly (P < 0.01) between hornet (R = 0.92) or paper wasp venom (R = 0.78) in improvement testing and conventional testing. To determine specificity, 20 volunteers who had never experienced a Hymenoptera sting were all negative for sIgE against these venoms in both improvement and conventional testing. Improved sensitivity was seen in 8 patients negative for sIgE against both venoms in conventional testing, while improvement testing revealed sIgE against hornet or paper wasp venom in 5 (total 38 (92.7%)) patients. CONCLUSIONS: The measurement of sIgE following spiking of rVes v 5 and rPol d 5 by conventional testing in Japanese subjects with sIgE against hornet and paper wasp venom, respectively, improved the sensitivity for detecting Hymenoptera venom allergy. Improvement testing for measuring sIgE levels against hornet and paper wasp venom has potential for serologically elucidating Hymenoptera allergy in Japan.