Polysaccharides from Platycodonis Radix ameliorated respiratory syncytial virus-induced epithelial cell apoptosis and inflammation through activation of miR-181a-mediated Hippo and SIRT1 pathways.

Respiratory syncytial virus (RSV) is the leading cause of bronchiolitis in young children, but there are few safe and effective treatments for this disease. Platycodonis Radix is widely used as an antitussive and expectorant drug for preventing various diseases in lower respiratory tract, in which the polysaccharides are one of the main bioactivity constituents. In this study, the protective effects of the P. Radix polysaccharides (PRP) against RSV-induced bronchiolitis in juvenile mice and RSV-induced apoptosis of epithelial HEp-2 cells were investigated. The results showed that PRP obviously decreased the levels of IL-1ß, IL-4, IL-6, TNF-α, IFN-γ and TSLP in lung tissues, and reduced the number of inflammatory cells in bronchoalveolar lavage fluid (BALF) of RSV-infected mice. Furthermore, it reduced the apoptosis of RSV-infected HEp-2 cells and remarkably inhibited the mRNA expressions of RSV L gene, which indicated that PRP affected transcription and replication of RSV in host cells. Compared with that in RSV-infected group, miR-181a-5p in the PRP-treated group presented the highest relative abundance and its expression was violently reduced by approximately 30%. Mechanistically, PRP had the similar effects as miR-181a-5p antagomir on RSV-induced apoptosis and inflammation in HEp-2 cells via upregulating BCL2, MLL3 and SIRT1, which could be reversed by miR-181a-5p mimic. Therefore, it demonstrated that PRP not only protected against RSV-induced lung inflammation in mice but also inhibited apoptosis of RSV-infected HEp-2 cells via suppressing miR-181a-5p and transcriptionally activating Hippo and SIRT1 pathways.

The MIF Antagonist ISO-1 Attenuates Corticosteroid-Insensitive Inflammation and Airways Hyperresponsiveness in an Ozone-Induced Model of COPD.

INTRODUCTION: Macrophage migration inhibitory factor (MIF) is an inflammatory cytokine associated with acute and chronic inflammatory disorders and corticosteroid insensitivity. Its expression in the airways of patients with chronic obstructive pulmonary disease (COPD), a relatively steroid insensitive inflammatory disease is unclear, however. METHODS: Sputum, bronchoalveolar lavage (BAL) macrophages and serum were obtained from non-smokers, smokers and COPD patients. To mimic oxidative stress-induced COPD, mice were exposed to ozone for six-weeks and treated with ISO-1, a MIF inhibitor, and/or dexamethasone before each exposure. BAL fluid and lung tissue were collected after the final exposure. Airway hyperresponsiveness (AHR) and lung function were measured using whole body plethysmography. HIF-1α binding to the Mif promoter was determined by Chromatin Immunoprecipitation assays. RESULTS: MIF levels in sputum and BAL macrophages from COPD patients were higher than those from non-smokers, with healthy smokers having intermediate levels. MIF expression correlated with that of HIF-1α in all patients groups and in ozone-exposed mice. BAL cell counts, cytokine mRNA and protein expression in lungs and BAL, including MIF, were elevated in ozone-exposed mice and had increased AHR. Dexamethasone had no effect on these parameters in the mouse but ISO-1 attenuated cell recruitment, cytokine release and AHR. CONCLUSION: MIF and HIF-1α levels are elevated in COPD BAL macrophages and inhibition of MIF function blocks corticosteroid-insensitive lung inflammation and AHR. Inhibition of MIF may provide a novel anti-inflammatory approach in COPD.

Low Level Laser Therapy Reduces the Development of Lung Inflammation Induced by Formaldehyde Exposure.

Lung diseases constitute an important public health problem and its growing level of concern has led to efforts for the development of new therapies, particularly for the control of lung inflammation. Low Level Laser Therapy (LLLT) has been highlighted as a non-invasive therapy with few side effects, but its mechanisms need to be better understood and explored. Considering that pollution causes several harmful effects on human health, including lung inflammation, in this study, we have used formaldehyde (FA), an environmental and occupational pollutant, for the induction of neutrophilic lung inflammation. Our objective was to investigate the local and systemic effects of LLLT after FA exposure. Male Wistar rats were exposed to FA (1%) or vehicle (distillated water) during 3 consecutive days and treated or not with LLLT (1 and 5 hours after each FA exposure). Non-manipulated rats were used as control. 24 h after the last FA exposure, we analyzed the local and systemic effects of LLLT. The treatment with LLLT reduced the development of neutrophilic lung inflammation induced by FA, as observed by the reduced number of leukocytes, mast cells degranulated, and a decreased myeloperoxidase activity in the lung. Moreover, LLLT also reduced the microvascular lung permeability in the parenchyma and the intrapulmonary bronchi. Alterations on the profile of inflammatory cytokines were evidenced by the reduced levels of IL-6 and TNF-α and the elevated levels of IL-10 in the lung. Together, our results showed that LLLT abolishes FA-induced neutrophilic lung inflammation by a reduction of the inflammatory cytokines and mast cell degranulation. This study may provide important information about the mechanisms of LLLT in lung inflammation induced by a pollutant.

Involvement of CD300a Phosphatidylserine Immunoreceptor in Aluminum Salt Adjuvant-Induced Th2 Responses.

Aluminum salt (alum) has been widely used for vaccinations as an adjuvant. Alum not only enhances immunogenicity but also induces Th2 cell immune responses. However, the mechanisms of how alum enhances Th2 cell immune responses have been controversial. In an experimental allergic airway inflammation model, in which alum in conjunction with OVA Ag was i.p. injected for immunization, we found that apoptotic cells and inflammatory dendritic cells (iDC) expressing CD300a, an inhibitory immunoreceptor for phosphatidylserine (PS), significantly increased in number in the peritoneal cavity after the immunization. In contrast, apoptotic cells and iDCs were scarcely observed in the peritoneal cavity after injection of OVA alone. In CD300a-deficient mice, eosinophil infiltration in bronchoalveolar lavage fluid, serum IgE levels, and airway hyperreactivity were significantly decreased after immunization with alum plus OVA compared with wild-type mice. In vitro, iDCs purified from CD300a-deficient mice after the immunization induced significantly less IL-4 production from OT-II naive CD4(+) T cells after coculture with OVA Ag. CD300a expressed on iDCs bound PS on apoptotic cells in the peritoneal cavity after injection of OVA plus alum. Blocking CD300a interaction with PS by injection of a neutralizing anti-CD300a Ab resulted in inhibition of the development of allergic airway inflammation. These results suggest that CD300a is involved in alum-induced Th2 skewing.

The Wnt/ß-catenin pathway attenuates experimental allergic airway disease.

Signaling via the Wnt/ß-catenin pathway plays crucial roles in embryogenesis and homeostasis of adult tissues. In the lung, the canonical Wnt/ß-catenin pathway has been implicated in remodeling processes, development of emphysema, and fibrosis. However, its relevance for the modulation of allergic responses in the lung remains unclear. Using genetically modified mice with lung-specific inducible (doxycycline) Wnt-1 expression (CCSP-rtTA × tetO-Wnt1), the impact of Wnt on the development of allergic airway disease was analyzed. Overexpression of Wnt during the allergen challenge phase attenuated the development of airway inflammation in an acute model, as well as in a more therapeutic model of secondary challenge. These findings were further supported by treatment of allergen-sensitized mice with LiCl during challenge. Similar to Wnt, LiCl prevented the degradation of ß-catenin and, thus, attenuated allergic airway inflammation and hyperresponsiveness. Migration studies revealed that lung-specific expression of Wnt reduced the migration of Ag-loaded dendritic cells (DCs) into the draining lymph nodes following allergen challenge. Administration of in vitro allergen-loaded DCs overcame Wnt-mediated suppression of airway inflammation. Furthermore, in vitro studies confirmed that DC-dependent T cell activation is impaired by blocking ß-catenin degradation. These results demonstrate an important role for the canonical Wnt/ß-catenin pathway in the DC-mediated regulation of allergic responses in the lung.

Engineered silica nanoparticles act as adjuvants to enhance allergic airway disease in mice.

BACKGROUND: With the increase in production and use of engineered nanoparticles (NP; ≤ 100 nm), safety concerns have risen about the potential health effects of occupational or environmental NP exposure. Results of animal toxicology studies suggest that inhalation of NP may cause pulmonary injury with subsequent acute or chronic inflammation. People with chronic respiratory diseases like asthma or allergic rhinitis may be even more susceptible to toxic effects of inhaled NP. Few studies, however, have investigated adverse effects of inhaled NP that may enhance the development of allergic airway disease. METHODS: We investigated the potential of polyethylene glycol coated amorphous silica NP (SNP; 90 nm diameter) to promote allergic airway disease when co-exposed during sensitization with an allergen. BALB/c mice were sensitized by intranasal instillation with 0.02% ovalbumin (OVA; allergen) or saline (control), and co-exposed to 0, 10, 100, or 400 µg of SNP. OVA-sensitized mice were then challenged intranasally with 0.5% OVA 14 and 15 days after sensitization, and all animals were sacrificed a day after the last OVA challenge. Blood and bronchoalveolar lavage fluid (BALF) were collected, and pulmonary tissue was processed for histopathology and biochemical and molecular analyses. RESULTS: Co-exposure to SNP during OVA sensitization caused a dose-dependent enhancement of allergic airway disease upon challenge with OVA alone. This adjuvant-like effect was manifested by significantly greater OVA-specific serum IgE, airway eosinophil infiltration, mucous cell metaplasia, and Th2 and Th17 cytokine gene and protein expression, as compared to mice that were sensitized to OVA without SNP. In saline controls, SNP exposure did cause a moderate increase in airway neutrophils at the highest doses. CONCLUSIONS: These results suggest that airway exposure to engineered SNP could enhance allergen sensitization and foster greater manifestation of allergic airway disease upon secondary allergen exposures. Whereas SNP caused innate immune responses at high doses in non-allergic mice, the adjuvant effects of SNP were found at lower doses in allergic mice and were Th2/Th17 related. In conclusion, these findings in mice suggest that individuals exposed to SNP might be more prone to manifest allergic airway disease, due to adjuvant-like properties of SNP.

Characteristics of patients with allergic rhinitis in an outpatient clinic: a retrospective study.

BACKGROUND: Allergic rhinitis affects a significant proportion of the European population. Few surveys have investigated this disorder in Greek adults. Our objective was to describe the characteristics of patients with allergic rhinitis in an adult outpatient clinic in Thessaloniki, Greece. METHODS: We studied the medical records of adult patients referred to a Clinical Immunology outpatient clinic from 2001 to 2007. The diagnostic procedure was not changed during the whole study period, including the same questionnaire used at the time of diagnosis, skin prick tests, and serum specific IgE. RESULTS: A total of 1851 patient files with diagnosed allergies were analysed and allergic rhinitis was confirmed in 711 subjects (38.4%). According to ARIA classification, persistent allergic rhinitis was more prevalent than intermittent (54.9% vs. 45.1%), while 60.8% of subjects suffered from moderate/severe disease. In multivariable analysis, factors associated with allergic rhinitis were age (for every 10 years increase, OR: 0.84, 95% CI: 0.77-0.91; p<0.001); working in school environment (teachers or students) (OR: 1.46, 95% CI: 1.05-2.02; p=0.023); parental history of respiratory allergy (OR: 2.41, 95% CI: 1.69-3.43; p<0.001); smoking (OR: 0.71, 95% CI: 0.55-0.91; p=0.007); presence of allergic conjunctivitis (OR: 6.16, 95% CI: 4.71-8.06; p<0.001); and asthma (OR: 2.17, 95% CI: 1.57-3.01; p<0.001). Analysis after multiple imputation corroborated the complete case analysis results. CONCLUSIONS: Allergic rhinitis was documented in 38.4% of studied patients and was frequently characterised by significant morbidity. Factors associated with allergic rhinitis provide insight into the epidemiology of this disorder in our region. Further studies on the general population would contribute to evaluating allergic rhinitis more comprehensively.

Down-regulation of 8-oxoguanine DNA glycosylase 1 expression in the airway epithelium ameliorates allergic lung inflammation.

Allergic airway inflammation is characterized by increased expression of pro-inflammatory mediators, inflammatory cell infiltration, mucus hypersecretion, and airway hyperresponsiveness, in parallel with oxidative DNA base and strand damage, whose etiological role is not understood. Our goal was to establish the role of 8-oxoguanine (8-oxoG), a common oxidatively damaged base, and its repair by 8-oxoguanine DNA glycosylase 1 (Ogg1) in allergic airway inflammatory processes. Airway inflammation was induced by intranasally administered ragweed (Ambrosia artemisiifolia) pollen grain extract (RWPE) in sensitized BALB/c mice. We utilized siRNA technology to deplete Ogg1 from airway epithelium; 8-oxoG and DNA strand break levels were quantified by Comet assays. Inflammatory cell infiltration and epithelial methaplasia were determined histologically, mucus and cytokines levels biochemically and enhanced pause was used as the main index of airway hyperresponsiveness. Decreased Ogg1 expression and thereby 8-oxoG repair in the airway epithelium conveyed a lower inflammatory response after RWPE challenge of sensitized mice, as determined by expression of Th2 cytokines, eosinophilia, epithelial methaplasia, and airway hyperresponsiveness. In contrast, 8-oxoG repair in Ogg1-proficient airway epithelium was coupled to an increase in DNA single-strand break (SSB) levels and exacerbation of allergen challenge-dependent inflammation. Decreased expression of the Nei-like glycosylases Neil1 and Neil2 that preferentially excise ring-opened purines and 5-hydroxyuracil, respectively, did not alter the above parameters of allergic immune responses to RWPE. These results show that DNA SSBs formed during Ogg1-mediated repair of 8-oxoG augment antigen-driven allergic immune responses. A transient modulation of OGG1 expression/activity in airway epithelial cells could have clinical benefits.

Cyclooxygenase inhibition during allergic sensitization increases STAT6-independent primary and memory Th2 responses.

Immune sensitization and memory generation are required for the development of allergic inflammation. Our previous studies demonstrate that the cyclooxygenase (COX) metabolic pathway is actively involved in allergic responses and COX inhibition increases allergic airway inflammation in a STAT6-independent fashion. To test the hypothesis that COX inhibition augments allergic inflammation by enhancing immune sensitization and memory, we sensitized STAT6 knockout mice with an i.p. injection of OVA with aluminum hydroxide as an adjuvant and treated the mice with the COX inhibitor indomethacin or vehicle for analyses of the primary and memory immune responses. We found that COX inhibition during immune sensitization, but not the allergic challenge phase, was necessary and sufficient to increase allergic inflammation. COX inhibition during sensitization increased the numbers of mature dendritic cells and activated CD4 T cells in the spleen and augmented OVA-specific IL-5 and IL-13 responses of the splenic CD4 T cells at day 5 after sensitization. COX inhibition during sensitization also augmented allergic Th2 response to OVA challenge 90 days after the sensitization. Therefore, COX inhibition during allergic sensitization augments allergic responses by enhancing Th2 cell activation and memory generation and the proallergic effect is STAT6-independent. These findings provide a mechanistic explanation for the increased allergic inflammation previously shown in the mice treated with COX inhibitors and in COX-deficient mice and suggest that use of COX-inhibiting drugs during initial allergen exposure may increase the risk of developing allergic responses.

Anti-inflammatory effects of zinc and alterations in zinc transporter mRNA in mouse models of allergic inflammation.

There is clinical evidence linking asthma with the trace element, zinc (Zn). Using a mouse model of allergic inflammation, we have previously shown that labile Zn decreases in inflamed airway epithelium (Truong-Tran AQ, Ruffin RE, Foster PS, Koskinen AM, Coyle P, Philcox JC, Rofe AM, Zalewski PD. Am J Respir Cell Mol Biol 27: 286-296, 2002). Moreover, mild nutritional Zn deficiency worsens lung function. Recently, a number of proteins belonging to the Solute Carrier Family 39 (ZIP) and Solute Carrier Family 30 (ZnT) have been identified that bind Zn and regulate Zn homeostasis. Mice were sensitized, and subsequently aerochallenged, with ovalbumin to induce acute and chronic airway inflammation. Mice received 0, 54, or 100 microg of Zn intraperitoneally. Tissues were analyzed for Zn content and histopathology. Inflammatory cells were counted in bronchoalveolar lavage fluid. Cytokine and Zn transporter mRNA levels were determined by cDNA gene array and/or real-time PCR. Zn supplementation decreased bronchoalveolar lavage fluid eosinophils by 40 and 80%, and lymphocytes by 55 and 66%, in the acute and chronic models, respectively. Alterations in Zn transporter expression were observed during acute inflammation, including increases in ZIP1 and ZIP14 and decreases in ZIP4 and ZnT4. Zn supplementation normalized ZIP1 and ZIP14, but it did not affect mRNA levels of cytokines or their receptors. Our results indicate that inflammation-induced alterations in Zn transporter gene expression are directed toward increasing Zn uptake. Increases in Zn uptake may be needed to counteract the local loss of Zn in the airway and to meet an increased demand for Zn-dependent proteins. The reduction of inflammatory cells by Zn in the airways provides support for Zn supplementation trials in human asthmatic individuals.

Allergic inflammatory response to short ragweed allergenic extract in HLA-DQ transgenic mice lacking CD4 gene.

To investigate the role of HLA-DQ molecules and/or CD4(+) T cells in the pathogenesis of allergic asthma, we generated HLA-DQ6 and HLA-DQ8 transgenic mice lacking endogenous class II (Abeta(null)) and CD4 genes and challenged them intranasally with short ragweed allergenic extract (SRW). We found that DQ6/CD4(null) mice developed a strong eosinophilic infiltration into the bronchoalveolar lavage and lung tissue, while DQ8/CD4(null) mice were normal. However, neither cytokines nor eosinophil peroxidase in the bronchoalveolar lavage of DQ6/CD4(null) mice was found. In addition, the airway reactivity to methacholine was elevated moderately in DQ6/CD4(null) mice compared with the high response in DQ/CD4(+) counterparts and was only partially augmented by CD4(+) T cell transfer. The DQ6/CD4(null) mice showed Th1/Th2-type cytokines and SRW-specific Abs in the immune sera in contrast to a direct Th2 response observed in DQ6/CD4(+) mice. The proliferative response of spleen mononuclear cells and peribronchial lymph node cells demonstrated that the response to SRW in DQ6/CD4(null) mice was mediated by HLA-DQ-restricted CD4(-)CD8(-)NK1.1(-) T cells. FACS analysis of PBMC and spleen mononuclear cells demonstrated an expansion of double-negative (DN) CD4(-)CD8(-)TCRalphabeta(+) T cells in SRW-treated DQ6/CD4(null) mice. These cells produced IL-4, IL-5, IL-13, and IFN-gamma when stimulated with immobilized anti-CD3. IL-5 ELISPOT assay revealed that DN T cells were the cellular origin of IL-5 in allergen-challenged DQ6/CD4(null) mice. Our study shows a role for HLA-DQ-restricted CD4(+) and DN T cells in the allergic response.

Sensitization to neumoallergens influence in mothers of asthmatic children.

Allergy diseases in our province have the characteristic of early appearance of asthma and predominant monosensitization to domestic dust mites due to the climatic condition. In this work we study the influence in allergic children. We selected 1028 mothers that brought their children to hospital for the first time. We divided them in two groups. One was comprised of 459 mothers of atopic children (MA) and another of 569 mothers of non atopic children (MC). All mothers were skin-prick tested (Phazet). The papula area, the mothers with aeroallergen sensitization, were 180 (17.5%) between MA 126 (27.4%) and MC 54 (9.5%). The diagnosis of allergic mothers was higher in MA than in MC. Asthma 8.39%, Rhinitis 4.75% and dermatitis 2%. 53.31% lived on the coast. 51.45% had house moulds. 31% had animals. 34.9% were smokers. There were more adverse drug reactions in MA than in MC. 180 mothers presented sensitization. D. pteronyssinus 66.3%, D. farinae 65.5% Lolium P. 14.55%, Cats 11.6% (MA > MC), Artemisia Vulgaris 6.25% and Cladosporium Herbarum 0.6%. There was no correlation between skin sensitization and domestic animals. There was no influence between children's IgE and mothers with skin sensitization. Skin sensitization to grass pollen correlated with the mother who was living in another country. The mothers who have been living more than five years in our province presented more skin sensitization to aero-allergens.