Exposure of cinnamyl alcohol in co-culture of BEAS-2B and dendritic cells: Interaction between CYP450 and cytokines.

The prevalence of fragrances in various hygiene products contributes to their sensorial allure. However, fragrances can induce sensitization in the skin or respiratory system, and the mechanisms involved in this process are incompletely understood. This study investigated the intricate mechanisms underlying the fragrance's effects on sensitization response, focusing on the interplay between CYP450 enzymes, a class of drug-metabolizing enzymes, and the adaptive immune system. Specifically, we assessed the expression of CYP450 enzymes and cytokine profiles in culture of BEAS-2B and mature dendritic cells (mDC) alone or in co-culture stimulated with 2 mM of a common fragrance, cinnamyl alcohol (CA) for 20 h. CYP1A1, CYP1A2, CYP1B1, CYP2A6, and CYP2A13 were analyzed by RT-PCR and IL-10, IL-12p70, IL-18, IL-33, and thymic stromal lymphopoietin (TSLP) by Cytometric Bead Array (CBA). Through RT-PCR analysis, we observed that CA increased CYP1A2 and CYP1B1 expression in BEAS-2B, with a further increased in BEAS-2B-mDC co-culture. Additionally, exposure to CA increased IL-12p70 levels in mDC rather than in BEAS-2B-mDC co-culture. In regards to IL-18, level was higher in BEAS-2B than in BEAS-2B-mDC co-culture. A positive correlation between the levels of IL-10 and CYP1B1 was found in mDC-CA-exposed and between IL-12p70 and CYP1A1 was found in BEAS-2B after CA exposure. However, IL-12p70 and CYP1A2 as well as IL-18, IL-33, and CYP1A1 levels were negative, correlated mainly in co-culture control. These correlations highlight potential immunomodulatory interactions and complex regulatory relationships. Overall, exposure to CA enhances CYP450 expression, suggesting that CA can influence immune responses by degrading ligands on xenosensitive transcription factors.

Expression patterns of peroxiredoxin genes in bronchial epithelial cells exposed to diesel exhaust particles.

Several mechanisms have been suggested to explain the adverse effects of air pollutants on airway cells. One such explanation is the presence of high concentrations of oxidants and pro-oxidants in environmental pollutants. All animal and plant cells have developed several mechanisms to prevent damage by oxidative molecules. Among these, the peroxiredoxins (PRDXs) are of interest due to a high reactivity with reactive oxygen species (ROS) through the functioning of the thioredoxin/thioredoxin reductase system. This study aimed to verify the gene expression patterns of the PRDX family in bronchial epithelial airway cells (BEAS-2B) cells exposed to diesel exhaust particles (DEPs) at a concentration of 15 µg/mL for 1 or 2 h because this it is a major component of particulate matter in the atmosphere. There was a significant decrease in mRNA fold changes of PRDX2 (0.43 ± 0.34; *p = 0.0220), PRDX5 (0.43 ± 0.34; *p = 0.0220), and PRDX6 (0.33 ± 0.25; *p = 0.0069) after 1 h of exposure to DEPs. The reduction in mRNA levels may consequently lead to a decrease in the levels of PRDX proteins, increasing oxidative stress in bronchial epithelial cells BEAS-2B and thus, negatively affecting cellular functions.

Inhibition of MAPK and STAT3-SOCS3 by Sakuranetin Attenuated Chronic Allergic Airway Inflammation in Mice.

Asthma allergic disease is caused by airway chronic inflammation. Some intracellular signaling pathways, such as MAPK and STAT3-SOCS3, are involved in the control of airway inflammation in asthma. The flavonoid sakuranetin demonstrated an anti-inflammatory effect in different asthma models. Our aim was to clarify how sakuranetin treatment affects MAPK and STAT3-SOCS3 pathways in a murine experimental asthma model. Mice were submitted to an asthma ovalbumin-induction protocol and were treated with vehicle, sakuranetin, or dexamethasone. We assayed the inflammatory profile, mucus production, and serum antibody, STAT3-SOCS3, and MAPK levels in the lungs. Morphological alterations were also evaluated in the liver. LPS-stimulated RAW 264.7 cells were used to evaluate the effects of sakuranetin on nitric oxide (NO) and cytokine production. In vivo, sakuranetin treatment reduced serum IgE levels, lung inflammation (eosinophils, neutrophils, and Th2/Th17 cytokines), and respiratory epithelial mucus production in ovalbumin-sensitized animals. Considering possible mechanisms, sakuranetin inhibits the activation of ERK1/2, JNK, p38, and STAT3 in the lungs. No alterations were found in the liver for treated animals. Sakuranetin did not modify in vitro cell viability in RAW 264.7 and reduced NO release and gene expression of IL-1ß and IL-6 induced by LPS in these cells. In conclusion, our data showed that the inhibitory effects of sakuranetin on eosinophilic lung inflammation can be due to the inhibition of Th2 and Th17 cytokines and the inhibition of MAPK and STAT3 pathways, reinforcing the idea that sakuranetin can be considered a relevant candidate for the treatment of inflammatory allergic airway disease.

Vesicular acetylcholine transport deficiency potentiates some inflammatory responses induced by diesel exhaust particles.

Endogenous acetylcholine (ACh), which depends of the levels of vesicular ACh transport (VAChT) to be released, is the central mediator of the cholinergic anti-inflammatory system. ACh controls the release of cytokine in different models of inflammation. Diesel exhaust particles (DEP) are one of the major environmental pollutants produced in large quantity by automotive engines in urban center. DEP bind the lung parenchyma and induce inflammation. We evaluated whether cholinergic dysfunction worsens DEP-induced lung inflammation. Male mice with decreased ACh release due to reduced expression of VAChT (VAChT-KD mice) were submitted to DEP exposure for 30 days (3 mg/mL of DEP, once a day, five days a week) or saline. Pulmonary function and inflammation as well as extracellular matrix fiber deposition were evaluated. Additionally, airway and nasal epithelial mucus production were quantified. We found that DEP instillation worsened lung function and increased lung inflammation. Higher levels of mononuclear cells were observed in the peripheral blood of both wild-type (WT) and VAChT-KD mice. Also, both wild-type (WT) and VAChT-KD mice showed an increase in macrophages in bronchoalveolar lavage fluid (BALF) as well as increased expression of IL-4, IL-6, IL-13, TNF-α, and NF-κB in lung cells. The collagen fiber content in alveolar septa was also increased in both genotypes. On the other hand, we observed that granulocytes were increased only in VAChT-KD peripheral blood. Likewise, increased BALF lymphocytes and neutrophils as well as increased elastic fibers in alveolar septa, airway neutral mucus, and nasal epithelia acid mucus were observed only in VAChT-KD mice. The cytokines IL-4 and TNF-α were also higher in VAChT-KD mice compared with WT mice. In conclusion, decreased ability to release ACh exacerbates some of the lung alterations induced by DEP in mice, suggesting that VAChT-KD animals are more vulnerable to the effects of DEP in the lung.

Effects of organic and inorganic compounds of diesel exhaust particles on the mucociliary epithelium: An experimental study on the frog palate preparation.

The toxic actions of acute exposition to different diesel exhaust particles (DEPA) fractions on the mucociliary epithelium are not yet fully understood due to different concentrations of organic and inorganic elements. These chemicals elements produce damage to the respiratory epithelium and exacerbate pre-existent diseases. In our study we showed these differences in two experimental studies. Study I (dose-response curve - DRCS): Forty frog-palates were exposed to the following dilutions: frog ringer, intact DEPA diluted in frog-ringer at 3mg/L, 6mg/L and 12mg/L. Study II (DEPF) (DEPA fractions diluted at 12mg/L): Fifty palates - Frog ringer, intact DEPA, DEPA treated with hexane, nitric acid and methanol. Variables analyzed: relative time of mucociliary transport (MCT), ciliary beating frequency (CBF) and morphometric analysis for mucin profile (neutral/acid) and vacuolization. The Results of DRCS: Group DEPA-12mg/L presented a significant increase in the MCT (p<0.05), proportional volume of acid mucus (p<0.05) and decreased proportional volume of neutral mucus and vacuoles (p<0.05). In relation of DEPF: A significant increase in the MCT associated to a decrease in the proportional volume of neutral mucus was founded in nitric acid group. In addition, a significant increase in the proportional volume of acid mucus was found in methanol group. We concluded that: 1) Increasing concentrations of intact DEPA can progressively increase MCT and promote an acidification of intra-epithelial mucins associated to a depletion of neutral mucus. 2) Intact DEPA seem to act as secretagogue substance, promoting mucus extrusion and consequently reducing epithelial thickness. 3) Organic fraction of low polarity seems to play a pivotal role on the acute toxicity to the mucociliary epithelium, by promoting a significant increase in the MCT associated to changes in the chemical profile of the intracellular mucins.

Chronic exposure of diesel exhaust particles induces alveolar enlargement in mice.

BACKGROUND: Diesel exhaust particles (DEPs) are deposited into the respiratory tract and are thought to be a risk factor for the development of diseases of the respiratory system. In healthy individuals, the timing and mechanisms of respiratory tract injuries caused by chronic exposure to air pollution remain to be clarified. METHODS: We evaluated the effects of chronic exposure to DEP at doses below those found in a typical bus corridor in Sao Paulo (150 µg/m3). Male BALB/c mice were divided into mice receiving a nasal instillation: saline (saline; n = 30) and 30 µg/10 µL of DEP (DEP; n = 30). Nasal instillations were performed five days a week, over a period of 90 days. Bronchoalveolar lavage (BAL) was performed, and the concentrations of interleukin (IL)-4, IL-10, IL-13 and interferon-gamma (INF-γ) were determined by ELISA-immunoassay. Assessment of respiratory mechanics was performed. The gene expression of Muc5ac in lung was evaluated by RT-PCR. The presence of IL-13, MAC2+ macrophages, CD3+, CD4+, CD8+ T cells and CD20+ B cells in tissues was analysed by immunohistochemistry. Bronchial thickness and the collagen/elastic fibers density were evaluated by morphometry. We measured the mean linear intercept (Lm), a measure of alveolar distension, and the mean airspace diameter (D0) and statistical distribution (D2). RESULTS: DEP decreased IFN-γ levels in BAL (p = 0.03), but did not significantly alter IL-4, IL-10 and IL-13 levels. MAC2+ macrophage, CD4+ T cell and CD20+ B cell numbers were not altered; however, numbers of CD3+ T cells (p ≤ 0.001) and CD8+ T cells (p ≤ 0.001) increased in the parenchyma. Although IL-13 (p = 0.008) expression decreased in the bronchiolar epithelium, Muc5ac gene expression was not altered in the lung of DEP-exposed animals. Although respiratory mechanics, elastic and collagen density were not modified, the mean linear intercept (Lm) was increased in the DEP-exposed animals (p ≤ 0.001), and the index D2 was statistically different (p = 0.038) from the control animals. CONCLUSION: Our data suggest that nasal instillation of low doses of DEP over a period of 90 days results in alveolar enlargement in the pulmonary parenchyma of healthy mice.

Organic and inorganic fractions of diesel exhaust particles produce changes in mucin profile of mouse trachea explants.

Diesel exhaust particles (DEP) contain organic and inorganic elements that produce damage to the respiratory epithelium. The aim of this study was to determine the mucus profile of tracheal explants exposed to either crude diesel exhaust particles (DEP) or DEP treated with nitric acid (DEP/NA), with hexane (DEP/HEX), or with methanol (DEP/MET) at concentrations of 50 and 100 µg/ml for 30 and 60 min. Tracheal explants were subjected to morphometric analyses to study acidic (AB+), neutral (PAS+), and mixed (AB+/PAS+) mucus production and vacuolization (V). Incubation with 50 µg/ml crude DEP resulted in a rise in acid mucus production, an increase in vacuolization at 30 min, and reduction in neutral mucus at 30 and 60 min. Tracheas exposed to DEP/MET at 50 µg/ml for 30 or 60 min resulted in a significant decrease in neutral mucus production and an elevation in acid mucus production. DEP/HEX increased vacuolization at both 50 and 100 µg/ml at 30 and 60 min of exposure. Treatment with 50 µg/ml for 30 or 60 min significantly elevated mixed mucus levels. These results suggest that DEP appear to be more toxic when administered in combination with HEX or MET. DEP/MET modified the mucus profile of the epithelium, while DEP/HEX altered mucus extrusion, and these responses might be due to bioavailability of individual elements in DEP fractions.

In vitro mucus transportability, cytogenotoxicity, and hematological changes as non-destructive physiological biomarkers in fish chronically exposed to metals.

The biomonitoring of fish using biomarkers represents a useful tool for the assessment of aquatic pollution. This study evaluated the sublethal toxic effects of aquatic pollution on fish collected from a site contaminated by metals. Water and fish (Oreochromis niloticus) samples were collected from a pond in the Parque Ecológico do Tietê (PET) that lies along the Tietê River (São Paulo, Brazil), and from a control site (an experimental fish farm). The metal content of the water was evaluated, and fish were used to examine the properties of gill mucus and blood. The PET fish were evaluated for alterations in the in vitro transportability of mucus and changes in blood properties (e.g., cell volume, hemoglobin concentration, red blood cells, and white blood cell count). The results of the water analyzes indicated metal levels above the legal standards for Fe (0.71 mg/L), Ni (0.06 mg/L), Mn (0.11 mg/L), and Pb (0.48 mg/L). Compared to the controls, the hematologic parameter analyzes of PET fish revealed significantly higher numbers of erythrocytes (RBC), leukocytes (WBC), lymphocytes, erythroblasts, and Mean Corpuscular Volume (MCV); however, the hemoglobin content and Mean Corpuscular Hemoglobin Concentration (MCHC) values were significantly lower. The frequencies of nuclear abnormalities and micronuclei were significantly higher and the mucociliary transport was significantly lower in PET fish than in the controls. These results suggest that fish from the contaminated site exhibit a series of physiological responses, which probably indicate health disturbances. Furthermore, the results suggest that blood and mucus are promising, non-destructive targets for use in the monitoring of pollution.

Diesel exhaust particulates affect cell signaling, mucin profiles, and apoptosis in trachea explants of Balb/C mice.

Particulate matter from diesel exhaust (DEP) has toxic properties and can activate intracellular signaling pathways and induce metabolic changes. This study was conducted to evaluate the activation of extracellular signal-regulated kinase (ERK) and c-Jun N-terminal kinase (JNK) and to analyze the mucin profile (acid (AB(+) ), neutral (PAS(+) ), or mixed (AB/PAS(+) ) mucus) and vacuolization (V) of tracheal explants after treatment with 50 or 100 µg/mL DEP for 30 or 60 min. Western blot analyses showed small increases in ERK1/2 and JNK phosphorylation after 30 min of 100 µg/mL DEP treatment compared with the control. An increase in JNK phosphorylation was observed after 60 min of treatment with 50 µg/mL DEP compared with the control. We did not observe any change in the level of ERK1/2 phosphorylation after treatment with 50 µg/mL DEP. Other groups of tracheas were subjected to histological sectioning and stained with periodic acid-Schiff (PAS) reagent and Alcian Blue (AB). The stained tissue sections were then subjected to morphometric analysis. The results obtained were compared using ANOVA. Treatment with 50 µg/mL DEP for 30 min or 60 min showed a significant increase (p < 0.001) in the amount of acid mucus, a reduction in neutral mucus, a significant reduction in mixed mucus, and greater vacuolization. Our results suggest that compounds found in DEPs are able to activate acid mucus production and enhance vacuolization and cell signaling pathways, which can lead to airway diseases.

Effects of cigarette smoking intensity on the mucociliary clearance of active smokers.

BACKGROUND: Smoking impairs mucociliary clearance and increases respiratory infection frequency and severity in subjects with and without smoking-related chronic lung diseases. OBJECTIVE: This study evaluated the effects of smoking intensity on mucociliary clearance in active smokers. METHODS: Seventy-five active smokers were grouped into light (1-10 cigarettes/day; n = 14), moderate (11-20 cigarettes/day; n = 34) and heavy smokers (≥21 cigarettes/day; n = 27) before starting a smoking cessation programme. Smoking behaviour, nicotine dependence, pulmonary function, carbon monoxide in exhaled air (exCO), carboxyhaemoglobin (COHb) and mucociliary clearance measured by the saccharin transit time (STT) test were all evaluated. An age-matched non-smoker group (n = 24) was assessed using the same tests. RESULTS: Moderate (49 ± 7 years) and heavy smokers (46 ± 8 years) had higher STT (p = 0.0001), exCO (p < 0.0001) and COHb (p < 0.0001) levels compared with light smokers (51 ± 15 years) and non-smokers (50 ± 11 years). A positive correlation was observed between STT and exCO (r = 0.4; p < 0.0001), STT and cigarettes/day (r = 0.3, p = 0.02) and exCO and cigarettes/day (r = 0.3, p < 0.01). CONCLUSION: Smoking impairs mucociliary clearance and is associated with cigarette smoking intensity.

Effects of intrauterine exposure to concentrated ambient particles on allergic sensitization in juvenile mice.

Intrauterine exposure to particulate matter (PM) has been associated with an increased risk of asthma development, which may differ by the age of asthma onset, sex, and pollutant concentration. To investigate the pulmonary effects of in utero exposure to concentrated urban ambient particles (CAPs) in response to house dust mite (HDM) sensitization in juvenile mice. Mice were exposed to CAPs (600 µg/m3 PM2.5) during the gestational period. Twenty-two-day postnatal mice were sensitized with HDM (100 µg, intranasally, 3 times per week). Airway responsiveness (AHR), serum immunoglobulin, and lung inflammation were assessed after 43 days of the postnatal period. Female (n = 47) and male (n = 43) mice were divided into four groups as follows: (1) FA: not exposed to CAPs; (2) CAPs: exposed to CAPs; (3) HDM: sensitized to HDM; and (4) CAPs+HDM: exposed to CAPs and HDM-sensitized. PM2.5 exposure did not worsen lung hyperresponsiveness or allergic inflammation in sensitized animals. The levels of the lung cytokines IL-4, TNF-α, and IL-2 were differentially altered in male and female animals. Males presented hyporesponsiveness and increased lung macrophagic inflammation. There were no epigenetic changes in the IL-4 gene. In conclusion, intrauterine exposure ambient PM2.5 did not worsened allergic pulmonary susceptibility but affected the pulmonary immune profile and lung function, which differed by sex.

Low-dose chlorine exposure impairs lung function, inflammation and oxidative stress in mice.

AIM: To explore the different consequences of acute and chronic exposure to chlorine gas (Cl2) on the functional and histological parameters of health mice. MAIN METHODS: Firstly, male BALB/c mice were acute exposed to 3.3 or 33.3 or 70.5 mg/m3 Cl2. We analyzed the lung function, the inflammatory cells in the bronchoalveolar lavage, cell influx in the peribrochoalveolar space and mucus production. In a second phase, mice were chronic exposed to 70.5 mg/m3 Cl2. Besides the first phase analyses, we also evaluated the epithelial cells thickness, collagen deposition in the airways, immunohistochemistry stain for IL-1ß, iNOS, IL-17 and ROCK-2 and the levels of IL-5, IL-13, IL-17, IL-1ß and TNF-α in lung homogenate. KEY FINDINGS: Acute exposure to chlorine impaired the lung function, increased the number of inflammatory cells in the BALF and in the airways, also increased the mucus production. Furthermore, when chlorine was exposed chronically, increased the airway remodeling with collagen deposition and epithelial cells thickness, positive cells for IL-1ß, iNOS, IL-17 in the airways and in the alveolar walls and ROCK-2 in the alveolar walls, lung inflammation with increased levels of IL-5, IL-13, IL-1ß and TNF-α in the lung homogenate, and also, induced the acid mucus production by the nasal epithelium. SIGNIFICANCE: Acute and chronic exposure to low dose of chlorine gas worsens lung function, induces oxidative stress activation and mucus production and contributes to augmenting inflammation in health mice.

Effectiveness of the 40-Minute Handmade Manikin Program to Teach Hands-on Cardiopulmonary Resuscitation at School Communities.

Bystander training in cardiopulmonary resuscitation (CPR) is crucial to improve the victims' survival and quality of life after sudden cardiac arrest. This observational study aimed to determine the success rate of 2 different programs of CPR training for children, adolescents, and adults in school communities. We assessed the development and acquisition of the following CPR skills: checking local safety, assessing victim's responsiveness, calling for help, assessing victim's breathing, and performing chest compression (hands and straight arms placement on the chest, compression velocity, depth, and chest release) using a 40-minute program with handmade manikins or the 120-minute program using intermediate-fidelity manikins. There were 1,630 learners (mean age 16 years, 38% male) in the 40-minute program, and 347 learners (mean age 27 years, 32% male) in the 120-minute program. The lowest successful pass rate of learners that developed CPR skills was 89.4% in the 40-minute program and 84.5% in the 120-minute program. The chances of success increased with age in the same program (compression rate and depth). The success rate also increased with the more extended and intermediate-cost program at the same age (assessing victim's responsiveness, calling for help, and assessing the victim's respiration). In conclusion, a 40-minute and cheaper (low-cost handmade manikin) CPR program was adequate to develop and acquire the overall CPR skills for ≥89% at school communities, independently of gender. However, some individual CPR skills can be further improved with increasing age and using the longer and intermediate-cost program.

The effects of chronic exposure to traffic derived air pollution on the ocular surface.

OBJECTIVES: The purpose of this study was to explore the clinical relevance of chronic exposure to ambient levels of traffic derived air pollution on the ocular surface. METHODS: A panel study involving 55 volunteers was carried out in São Paulo, Brazil. We measured the mean individual levels of nitrogen dioxide (NO(2)) exposure for 7 days. All subjects answered the Ocular Symptom Disease Index (OSDI) and a symptoms inventory. Subsequently, subjects underwent Schirmer I test, biomicroscopy, vital staining and tear breakup time (TBUT) assessment. Subject's mean daily exposure to NO(2) was categorized in quartiles. Statistical analysis was performed using one-way ANOVA, Tukey HSD and Chi-Square tests. RESULTS: A dose-response pattern was detected between OSDI scores and NO(2) quartiles (p<0.05). There was a significant association between NO(2) quartiles and reported ocular irritation (Chi(2)=9.2, p<0.05) and a significant negative association between TBUT and NO(2) exposure (p<0.05, R=-0.316, Spearman's correlation). There was a significant increase in the frequency of meibomitis in subjects exposed to higher levels of NO(2) (p<0.05). CONCLUSIONS: Subjects exposed to higher levels of traffic derived air pollution reported more ocular discomfort symptoms and presented greater tear film instability, suggesting that the ocular discomfort symptoms and tear breakup time could be used as convenient bioindicators of the adverse health effects of traffic derived air pollution exposure.

Smoking load reduction is insufficient to downregulate miR-301b, a lung cancer promoter.

Several circulating miRNAs identified in the plasma of smokers have been implicated as promoters of nasopharyngeal and lung carcinoma. To investigate the plasma profile of miRNAs in subjects who reduces the number of smoked cigarettes and who quit after six months. We accompanied 28 individuals enrolled in a Smoking Cessation Program over 6 months. At Baseline, clinical characteristics, co-morbidities, and smoking history were similar among subjects. After 6 months, two groups were defined: who successfully quitted smoking (named "quitters", n = 18, mean age 57 years, 11 male) and who reduced the number of cigarettes smoked (20-90%) but failed to quit smoking (named "smokers", n = 10, mean age 52 years, 3 male). No significant clinical changes were observed between groups at baseline and after a 6-month period, however, quitters showed significant downregulations in seven miRNAs at baseline: miR-17 (- 2.90-fold, p = 0.029), miR-20a (- 3.80-fold, p = 0.021); miR-20b (- 4.71-fold, p = 0.027); miR-30a (- 3.95-fold, p = 0.024); miR-93 (- 3.63-fold, p = 0.022); miR-125a (- 1.70-fold, p = 0.038); and miR-195 (- 5.37-fold, p = 0.002), and after a 6-month period in 6 miRNAs: miR-17 (- 5.30-fold, p = 0.012), miR-20a (- 2.04-fold, p = 0.017), miR-20b (- 5.44-fold, p = 0.017), miR-93 (- 4.00-fold, p = 0.041), miR-101 (- 4.82-fold, p = 0.047) and miR-125b (- 3.65-fold, p = 0.025). Using time comparisons, only quitters had significant downregulation in miR-301b (- 2.29-fold, p = 0.038) after 6-month. Reductions in the number of smoked cigarettes was insufficient to change the plasma profile of miRNA after 6 months. Only quitting smoking (100% reduction) significantly downregulated miR-301b related to hypoxic conditions, promotion of cell proliferation, decreases in apoptosis, cancer development, and progression as increases in radiotherapy and chemotherapy resistance.

Nrf2 positively regulates autophagy antioxidant response in human bronchial epithelial cells exposed to diesel exhaust particles.

Diesel exhaust particles (DEP) are known to generate reactive oxygen species in the respiratory system, triggering cells to activate antioxidant defence mechanisms, such as Keap1-Nrf2 signalling and autophagy. The aim of this study was to investigate the relationship between the Keap1-Nrf2 signalling and autophagy pathways after DEP exposure. BEAS-2B cells were transfected with silencing RNA (siRNA) specific to Nrf2 and exposed to DEP. The relative levels of mRNA for Nrf2, NQO1, HO-1, LC3B, p62 and Atg5 were determined using RT-PCR, while the levels of LCB3, Nrf2, and p62 protein were determined using Western blotting. The autophagy inhibitor bafilomycin caused a significant decrease in the production of Nrf2, HO-1 and NQO1 compared to DEPs treatment, whereas the Nrf2 activator sulforaphane increased the LC3B (p = 0.020) levels. BEAS-2B cells exposed to DEP at a concentration of 50 µg/mL for 2 h showed a significant increase in the expression of LC3B (p = 0.001), p62 (p = 0.008), Nrf2 (p = 0.003), HO-1 (p = 0.001) and NQO1 (p = 0.015) genes compared to control. In siRNA-transfected cells, the LC3B (p < 0.001), p62 (p = 0.001) and Atg5 (p = 0.024) mRNA levels and the p62 and LC3II protein levels were decreased, indicating that Nrf2 modulated the expression of autophagy markers (R < 1). These results imply that, in bronchial cells exposed to DEP, the Nrf2 system positively regulates autophagy to maintain cellular homeostasis.

Severe pulmonary disease in an adult primary ciliary dyskinesia population in Brazil.

Primary Ciliary Dyskinesia (PCD) is underdiagnosed in Brazil. We enrolled patients from an adult service of Bronchiectasis over a two-year period in a cross-sectional study. The inclusion criteria were laterality disorders (LD), cough with recurrent infections and the exclusion of other causes of bronchiectasis. Patients underwent at least two of the following tests: nasal nitric oxide, ciliary movement and analysis of ciliary immunofluorescence, and genetic tests (31 PCD genes + CFTR gene). The clinical characterization included the PICADAR and bronchiectasis scores, pulmonary function, chronic Pseudomonas aeruginosa (cPA) colonization, exhaled breath condensate (EBC) and mucus rheology (MR). Forty-nine of the 500 patients were diagnosed with definite (42/49), probable (5/49), and clinical (2/49) PCD. Twenty-four patients (24/47) presented bi-allelic pathogenic variants in a total of 31 screened PCD genes. A PICADAR score > 5 was found in 37/49 patients, consanguinity in 27/49, LD in 28/49, and eight PCD sibling groups. FACED diagnosed 23/49 patients with moderate or severe bronchiectasis; FEV1 ≤ 50% in 25/49 patients, eight patients had undergone lung transplantation, four had been lobectomized and cPA+ was determined in 20/49. The EBC and MR were altered in all patients. This adult PCD population was characterized by consanguinity, severe lung impairment, genetic variability, altered EBC and MR.

Th17/Treg imbalance in COPD progression: A temporal analysis using a CS-induced model.

BACKGROUND: The imbalance between pro- and anti-inflammatory immune responses plays a pivotal role in chronic obstructive pulmonary disease (COPD) development and progression. To clarify the pathophysiological mechanisms of this disease, we performed a temporal analysis of immune response-mediated inflammatory progression in a cigarette smoke (CS)-induced mouse model with a focus on the balance between Th17 and Treg responses. METHODS: C57BL/6 mice were exposed to CS for 1, 3 or 6 months to induce COPD, and the control groups were maintained under filtered air conditions for the same time intervals. We then performed functional (respiratory mechanics) and structural (alveolar enlargement) analyses. We also quantified the NF-κB, TNF-α, CD4, CD8, CD20, IL-17, IL-6, FOXP3, IL-10, or TGF-ß positive cells in peribronchovascular areas and assessed FOXP3 and IL-10 expression through double-label immunofluorescence. Additionally, we evaluated the gene expression of NF-κB and TNF in bronchiolar epithelial cells. RESULTS: Our CS-induced COPD model exhibited an increased proinflammatory immune response (increased expression of the NF-κB, TNF-α, CD4, CD8, CD20, IL-17, and IL-6 markers) with a concomitantly decreased anti-inflammatory immune response (FOXP3, IL-10, and TGF-ß markers) compared with the control mice. These changes in the immune responses were associated with increased alveolar enlargement and impaired lung function starting on the first month and third month of CS exposure, respectively, compared with the control mice. CONCLUSION: Our results showed that the microenvironmental stimuli produced by the release of cytokines during COPD progression lead to a Th17/Treg imbalance.

Composition of diesel particles influences acute pulmonary toxicity: an experimental study in mice.

Ambient particles have been consistently associated with adverse health effects, yielding mainly high cardiorespiratory morbidity and mortality. Diesel engines represent a major source of particles in the urban scenario. We aimed to modify the composition of diesel particles, by means of different extraction procedures, to relate changes in chemical profile to corresponding indicators of respiratory toxicity. Male BALB/c mice were nasally instilled with saline, or with diesel particles, treated or not, and assigned to five groups: saline (SHAM), intact diesel particles (DEP), and diesel particles previously treated with methanol (METH), hexane (HEX), or nitric acid (NA). Elemental composition and organic compounds were analyzed. Twenty-four hours after nasal instillation, respiratory parameters were measured and lung tissue was collected for histological analysis. Static elastance was significantly increased in groups DEP and MET in relation to the other groups. HEX and NA were different from DEP but not significantly different from SHAM and METH groups. The difference between dynamic and static elastance was increased in DEP, METH, and NA treatments; HEX was not statistically different from SHAM. DEP and METH groups presented significantly increased upper airways resistance, while DEP, METH, and NA showed higher peripheral airways resistance values. All groups had a higher total resistance than SHAM. DEP, METH, and NA showed significant increased infiltration of polymorphonuclear cells. In conclusion, diesel particles treated with hexane (HEX) resulted in a respiratory-system profile very similar to that in SHAM group, indicating that hexane treatment attenuates pulmonary inflammation elicited by diesel particles.

Nasal mucus transportability in children with cleft palate.

OBJECTIVES: Children with cleft palate (CP) have a high prevalence of sinusitis. Considering that nasal mucus properties play a pivotal role in the upper airway defense mechanism, the aim of the study was to evaluate nasal mucus transportability and physical properties from children with CP. SETTING: Hospital for Rehabilitation of Craniofacial Anomalies, School of Dentistry, University of São Paulo, Bauru, SP, Brazil and Laboratory of Experimental Air Pollution, School of Medicine, University of São Paulo, São Paulo, SP, Brazil. METHODS: Nasal mucus samples were collected by nasal aspiration from children with CP and without CP (non-CP). Sneeze clearance (SC) was evaluated by the simulated sneeze machine. In vitro mucus transportability (MCT) by cilia was evaluated by the frog palate preparation. Mucus physical surface properties were assessed by measuring the contact angle (CA). Mucus rheology was determined by means of a magnetic rheometer, and the results were expressed as log G* (vectorial sum of viscosity and elasticity) and tan delta (relationship between viscosity and elasticity) measured at 1 and 100 rad/s. RESULTS: Mucus samples from children with CP had a higher SC than non-CP children (67+/-30 and 41+/-24 mm, respectively, p<0.05). Mucus samples from children with CP had a lower CA (24+/-16 degrees and 35+/-11 degrees , p<0.05) and a higher tan delta 100 (0.79+/-0.24 and 0.51+/-0.12, p<0.05) than non-CP children. There were no significant differences in mucus MCT, log G* 1, tan delta 1 and log G* 100 obtained for CP and non-CP children. CONCLUSIONS: Nasal mucus physical properties from children with CP are associated with higher sneeze transportability. The high prevalence of sinusitis in children with CP cannot be explained by changes in mucus physical properties and transportability.