Objective: To investigate the effect of intermittent hypoxia-a model of obstructive sleep apnea (OSA)-on pancreatic expression of uncoupling protein-2 (UCP2), as well as on glycemic and lipid profiles, in C57BL mice. Methods: For 8 h/day over a 35-day period, male C57BL mice were exposed to intermittent hypoxia (hypoxia group) or to a sham procedure (normoxia group). The intermittent hypoxia condition involved exposing mice to an atmosphere of 92% N and 8% CO2 for 30 s, progressively reducing the fraction of inspired oxygen to 8 ± 1%, after which they were exposed to room air for 30 s and the cycle was repeated (480 cycles over the 8-h experimental period). Pancreases were dissected to isolate the islets. Real-time PCR was performed with TaqMan assays. Results: Expression of UCP2 mRNA in pancreatic islets was 20% higher in the normoxia group than in the hypoxia group (p = 0.11). Fasting serum insulin was higher in the hypoxia group than in the normoxia group (p = 0.01). The homeostasis model assessment of insulin resistance indicated that, in comparison with the control mice, the mice exposed to intermittent hypoxia showed 15% lower insulin resistance (p = 0.09) and 21% higher pancreatic β-cell function (p = 0.01). Immunohistochemical staining of the islets showed no significant differences between the two groups in terms of the area or intensity of α- and β-cell staining for insulin and glucagon. Conclusions: To our knowledge, this is the first report of the effect of intermittent hypoxia on UCP2 expression. Our findings suggest that UCP2 regulates insulin production in OSA. Further study of the role that UCP2 plays in the glycemic control of OSA patients is warranted.

Keywords: Blood glucose; Sleep apnea syndromes; Pancreas; Glucagon-secreting cells.

Obstructive sleep apnea (OSA) has been associated with oxidative stress and various cardiovascular consequences, such as increased cardiovascular disease risk. Quantitative real-time PCR is frequently employed to assess changes in gene expression in experimental models. In this study, we analyzed the effects of chronic intermittent hypoxia (an experimental model of OSA) on housekeeping gene expression in the left cardiac ventricle of rats. Analyses via four different approaches-use of the geNorm, BestKeeper, and NormFinder algorithms; and 2−ΔCt (threshold cycle) data analysis-produced similar results: all genes were found to be suitable for use, glyceraldehyde-3-phosphate dehydrogenase and 18S being classified as the most and the least stable, respectively. The use of more than one housekeeping gene is strongly advised.

Keywords: Cell hypoxia; Reference standards; Sleep apnea, obstructive; Cardiovascular diseases; Models, animal; Polymerase chain reaction.

Objective: Obstructive sleep apnea syndrome is mainly characterized by intermittent hypoxia (IH) during sleep, being associated with several complications. Exposure to IH is the most widely used animal model of sleep apnea, short-term IH exposure resulting in cognitive and neuronal impairment. Pigment epithelium-derived factor (PEDF) is a hypoxia-sensitive factor acting as a neurotrophic, neuroprotective, and antiangiogenic agent. Our study analyzed performance on learning and cognitive tasks, as well as PEDF gene expression and PEDF protein expression in specific brain structures, in rats exposed to long-term IH. Methods: Male Wistar rats were exposed to IH (oxygen concentrations of 21-5%) for 6 weeks-the chronic IH (CIH) group-or normoxia for 6 weeks-the control group. After CIH exposure, a group of rats were allowed to recover under normoxic conditions for 2 weeks (the CIH+N group). All rats underwent the Morris water maze test for learning and memory, PEDF gene expression and PEDF protein expression in the hippocampus, frontal cortex, and temporal cortex being subsequently assessed. Results: The CIH and CIH+N groups showed increased PEDF gene expression in the temporal cortex, PEDF protein expression remaining unaltered. PEDF gene expression and PEDF protein expression remained unaltered in the frontal cortex and hippocampus. Long-term exposure to IH did not affect cognitive function. Conclusions: Long-term exposure to IH selectively increases PEDF gene expression at the transcriptional level, although only in the temporal cortex. This increase is probably a protective mechanism against IH-induced injury.

Keywords: Sleep apnea, central; Disease models, animal; Cognition; Sleep; Sleep apnea, obstructive.

Objective: To analyze adolescents clinically diagnosed with asthma, in terms of the physiological changes occurring during acute hypoxia and during a maximal stress test. Methods: This was a descriptive, cross-sectional study involving 48 adolescents (12-14 years of age) who were divided into three groups: mild intermittent asthma (MIA, n = 12); mild persistent asthma (MPA, n = 12); and control (n = 24). All subjects were induced to acute hypoxia and were submitted to maximal stress testing. Anthropometric data were collected, and functional variables were assessed before and after the maximal stress test. During acute hypoxia, the time to a decrease in SpO2 and the time to recovery of SpO2 (at rest) were determined. Results: No significant differences were found among the groups regarding the anthropometric variables or regarding the ventilatory variables during the stress test. Significant differences were found in oxygen half-saturation pressure of hemoglobin prior to the test and in PaO2 prior to the test between the MPA and control groups (p = 0.0279 and p = 0.0116, respectively), as was in the oxygen extraction tension prior to the test between the MIA and MPA groups (p = 0.0419). There were no significant differences in terms of the SpO2 times under any of the conditions studied. Oxygen consumption and respiratory efficiency were similar among the groups. The use of a bronchodilator provided no significant benefit during the hypoxia test. No correlations were found between the hypoxia test results and the physiological variables. Conclusions: Our findings suggest that adolescents with mild persistent asthma have a greater capacity to adapt to hypoxia than do those with other types of asthma.

Keywords: Asthma; Adolescent; Cell hypoxia; Exercise test.

Objective: Various studies of emphysema involve long-term exposure of animals to cigarette smoke, focusing on the cell type involved in the protease/antiprotease imbalance and on extracellular matrix degradation. In emphysema, increased expression of metalloproteinases is associated with cytokines, and evidence suggests that the matrix metalloproteinase-12 (MMP-12) plays an important role. Our objective was to investigate tissue inhibitor of metalloproteinase-2 (TIMP-2), tumor necrosis factor-alpha (TNF-α) and interleukin-6 (IL-6) detection by immunohistochemical methods in mouse lung. Methods: Male C57BL/6 mice were exposed 3 times a day to the smoke of 3 cigarettes over a period of 10, 20, 30 or 60 days in an inhalation chamber (groups CS10, CS20, CS30 and CS60, respectively). Controls were exposed to the same conditions in room air. Results: A progressive increase in the number of alveolar macrophages was observed in the bronchoalveolar lavage fluid of the exposed mice. The mean linear intercept, an indicator of alveolar destruction, was greater in all exposed groups when compared to control group. In the CS10, CS20 and CS30 mice, the immunohistochemical index (II) for MMP-12 increased in parallel with a decrease in II for TIMP-2 in the CS10, CS20 and CS30 mice. The II for the cytokines TNF-α and IL-6 was greater in all exposed groups than in the control group. Emphysema, with changes in volume density of collagen and elastic fibers, was observed in the CS60 group. Conclusions: These findings suggest that cigarette smoke induces emphysema with major participation of TNF-α and IL-6 without participation of neutrophils.

Keywords: Macrophages, alveolar; Emphysema; Tissue inhibitor of metalloproteinase-2; Interleukin-6.

Objective: To compare clinical and pathological staging in patients with non-small cell lung cancer submitted to surgical treatment, as well as to identify the causes of discordance. Methods: Data related to patients treated at the Department of Thoracic Surgery of the Pontifical Catholic University of Rio Grande do Sul São Lucas Hospital were analyzed retrospectively. Sensitivity, specificity, positive predictive value, negative predictive value, and accuracy were calculated for clinical stages IA, IB, and IIB. The kappa index was used to determine the concordance between clinical and pathological staging. Results: Of the 92 patients studied, 33.7% were classified as clinical stage IA, 50% as IB, and 16.3% as IIB. The concordance between clinical and pathological staging was 67.5% for stage IA, 54.3% for IB, and 66.6% for IIB. The accuracy of the clinical staging was greater for stage IA, and a kappa of 0.74, in this case, confirmed a substantial association with pathological staging. The difficulty in evaluating nodal metastatic disease is responsible for the low concordance in patients with clinical stage IB. Conclusions: The concordance between clinical and pathological staging is low, and patients are frequently understaged (in the present study, only one case was overstaged). Strategies are necessary to improve clinical staging and, consequently, the treatment and prognosis of patients with non-small cell lung cancer.

Keywords: Neoplasm staging; Lung neoplasms/diagnosis; Lung neoplasms/surgery; Prognosis.

Objective: To evaluate risk factors associated with asthma symptoms in adolescents in the 13- to 14-year age bracket. Methods: This was a cross-sectional study involving adolescents enrolled in randomly selected public schools in the city of Belo Horizonte, Brazil, and conducted with the use of the International Study of Asthma and Allergies in Childhood (ISAAC) questionnaire and its supplementary module for risk factor assessment. The ISAAC questionnaire was completed by the students themselves, whereas the supplementary questionnaire was completed by their parents or legal guardians. Variables showing p ≤ 0.25 in the univariate analysis were included in the multivariate analysis. Stepwise regression with backward elimination was used for variable selection. Results: We evaluated 375 adolescents, 124 (33.1%) of whom had asthma symptoms. The final multivariate analysis model revealed that asthma symptoms were associated with birth weight < 2,500 g (p < 0.001), day care center or nursery attendance (p < 0.002), maternal history of asthma (p < 0.001), contact with animals during the first year of life (p < 0.027), current contact with animals outside the home (dogs, cats, or farm animals; p < 0.005), and more than 20 cigarettes per day smoked by parents or other household members (p < 0.02). Conclusions: Exposure to animals in and outside the home is associated with asthma symptoms, as is environmental tobacco smoke exposure. Families, health professionals, and administrators of health care facilities should take that into account in order to prevent asthma and reduce asthma morbidity.

Keywords: Asthma; Risk factors; Adolescent.

Objective: To determine whether a short-term protocol using subcutaneous sensitization with ovalbumin, without the use of adjuvants, would induce an eosinophilic response in the lungs of mice similar to that observed in previous, well-established protocols. Methods: Adult female BALB/c mice were randomized and divided into groups according to the number of sensitizations with ovalbumin and the number/dosage of intranasal ovalbumin challenges. The short-term protocol (10 days) consisted of one sensitization with ovalbumin and three ovalbumin challenges (100 µg). Total and differential cell counts in BAL fluid, levels of eosinophil peroxidase in lung tissue, and histopathological examination of the lungs were performed 24 h after the last ovalbumin challenge. Results: No significant differences were found among the groups regarding the variables studied. The short-term protocol, as well as the other protocols studied, induced an eosinophilic response similar to that obtained in the positive control. Conclusions: Subcutaneous sensitization with ovalbumin and without the use of adjuvants resulted in a significant allergic response in the lungs of mice, even in the short-term protocol group. Our findings suggest that this short-term protocol can be used as a first-line pre-clinical test for the study of new medications, reducing the costs and observation periods.

Keywords: Ovalbumin; Mice; Asthma.

Background: Several lung diseases are characterized by a predominantly neutrophilic inflammation. A better understanding of the mechanisms of action of some drugs on the airway inflammation of such diseases may bring advances to the treatment. Objective: To develop a method to induce pulmonary neutrophilic response in mice, without active infection. Methods: Eight adult Swiss mice were used. The study group (n = 4) received an intranasal challenge with 1 x 1012 CFU/ml of Pseudomonas aeruginosa (Psa), frozen to death. The control group (n = 4) received an intranasal challenge with saline solution. Two days after the intranasal challenge, a bronchoalveolar lavage (BAL) was performed with total cell and differential cellularity counts. Results: The total cell count was significantly higher in the group with Psa, as compared to the control group (median of 1.17 x 106 and 0.08 x 106, respectively, p = 0.029). In addition to this, an absolute predominance of neutrophils was found in the differential cellularity of the mice that had received the Psa challenge. Conclusion: The model of inducing a neutrophilic pulmonary disease using frost-dead bacteria was successfully developed. This neutrophilic inflammatory response induction model in Swiss mice lungs may be an important tool for testing the anti-inflammatory effect of some antimicrobial drugs on the inflammation of the lower airways.

Keywords: Animal experimentation. Lung disesases. Inflammation. Neutrophils. Pseudomonas.

Objective: To compare the prevalence rates of drug-resistant Mycobacterium tuberculosis in patients under intermittent treatment with those observed in patients under daily treatment. Methods: We used World Health Organization data regarding 5,138 patients with active pulmonary TB in Brazil, separated into two groups: patients in the Federal District of Brasília, treated with a one-month daily regimen followed by an intermittent thrice-weekly regimen; and patients in other parts of Brazil, treated with a daily regimen only. The resistance pattern was categorized as primary or acquired, based on the history of previous treatment. Multidrug resistance was defined as resistance to at least isoniazid and rifampin, whereas monoresistance was defined as resistance to only one drug. Results: The prevalence of primary resistance in the Federal District of Brasília and in the other parts of Brazil, respectively, was as follows: overall, 9.2% and 9.3% (p = 0.94); monoresistance, 6.6% and 6.9% (p = 0.89); and multidrug resistance, 1.0% and 1.2% (p = 0.85). The prevalence of acquired resistance in the Federal District of Brasília and in the other parts of Brazil, respectively, was as follows: overall, 15.8% and 26.8% (p = 0.39); monoresistance, 5.3% and 13.7% (p = 0.33); and multidrug resistance, 0.0% and 10.2% (p = 0.16). Conclusions: No significant differences were found between patients treated with an intermittent regimen and those treated with a daily regimen in term of resistance rates.

Keywords: Tuberculosis, pulmonary; Drug resistance; Drug administration schedule; Tuberculosis, multidrug-resistant.

Objective: To investigate the diagnostic value of α-enolase (ENO1) and serum ENO1 autoantibody levels in lung cancer. Methods: Immunohistochemistry staining and ELISA were performed to detect ENO1 expression in lung tissue and serum ENO1 autoantibody levels, respectively. Results: The expression of ENO1 was higher in lung cancer tissues than in benign lung disease tissues (p < 0.001). The proportion of lung cancer samples expressing ENO1 was not significantly different among the various pathological classification groups. The proportion of samples expressing ENO1 was higher in lung cancer patients in stages I/II than in those in stages III/IV (χ2 = 5.445; p = 0.018). The expression of ENO1 in lung cancer tissues was not associated with age, gender, or smoking history. Serum ENO1 antibody levels were significantly higher in the lung cancer group than in the benign lung disease and control groups (p < 0.001). The differences among the pathological classification groups were not statistically significant. Serum ENO1 antibody levels were also in lung cancer patients in stages I/II than in those in stages III/IV (p < 0.01). Serum ENO1 antibody levels were not associated with age, gender, or smoking history in lung cancer patients. The ROC curve representing the diagnosis of lung cancer based on ENO1 antibody levels had an area under the curve of 0.806. Conclusions: Our results suggest that high levels of ENO1 are associated with the clinical stage of lung cancer and that ENO1 expression and its serum autoantibody levels show diagnostic value in lung cancer.

Keywords: Phosphopyruvate hydratase/analysis; Enzyme-linked immunosorbent assay; Immunohistochemistry; Lung neoplasms.

Objetivo: Avaliar o dano oxidativo (oxidação lipídica, oxidação proteica, thiobarbituric acid-reactive substances [TBARS, substâncias reativas ao ácido tiobarbitúrico], e carbonilação) e inflamação (expressão de phosphorylated AMP-activated protein kinase e de phosphorylated mammalian target of rapamycin (p-AMPK e p-mTOR, respectivamente) em tecido pulmonar e músculos do diafragma em camundongos C57BL/6 machos expostos à fumaça de cigarro (FC) por 7, 15, 30, 45 ou 60 dias. Métodos: Trinta e seis camundongos machos da espécie C57BL/6 foram divididos em seis grupos (n = 6/grupo): grupo controle e 5 grupos expostos a FC por 7, 15, 30, 45 e 60 dias, respectivamente. Resultados: Comparados aos camundongos controle, os camundongos expostos à FC apresentaram menor peso corporal em 30 dias. Nos camundongos expostos à FC (comparados aos controle) as maiores diferenças (aumentos) nos níveis de TBARS foram observados no dia 7 no músculo diafragma, comparado ao dia 45 em tecido pulmonar; as maiores diferenças (aumentos) nos níveis de carbonilas foram observados no dia 7 em ambos os tipos de tecido; e os níveis de sulfidrilas foram menores, nos dois tipos de tecidos, em todos os tempos. No tecido pulmonar e no músculo diafragma, a expressão de p-AMPK exibiu um comportamento semelhante ao dos níveis de TBARS. A expressão de p-mTOR foi maior que o valor controle nos dias 7 e 15 no tecido pulmonar, assim como no dia 45 no músculo diafragma. Conclusões: Nossos dados demonstram que a exposição à FC produz dano oxidativo tanto no tecido pulmonar quanto (primariamente) no tecido muscular, tendo um efeito adicional no músculo respiratório, como é frequentemente observado em fumantes com DPOC.

Keywords: Oxidative stress; Mice; Respiratory system; Smoking; Inflammation.

Objective: EBUS-TBNA cytological sampling is routinely performed for pathological diagnosis, mediastinal staging, and molecular testing in lung cancer patients. EBUS-TBNA samples are not formally accepted for testing programmed death-ligand 1 (PD-L1) expression. The objective of the study was to compare the feasibility, reproducibility, and accuracy of PD-L1 expression assessment in cytological specimens and histological samples. Methods: We prospectively collected histological (transbronchial forceps biopsy) and cytological (EBUS-TBNA) samples from peribronchial neoplastic lesions during an endoscopic procedure at the same target lesion for the pathological diagnosis and molecular assessment of stage IV non-small cell lung cancer (NSCLC). Results: Fifteen patients underwent the procedure. Adequate cytological samples (at least 100 neoplastic cells) were obtained in 12 cases (92.3%). Assessment of PD-L1 expression was similar between histological and cytological samples (agreement rate = 92%). Sensitivity and diagnostic accuracy of EBUS-TBNA cytological specimens were 88.9% and 100%, respectively. Conclusions: The evaluation of PD-L1 expression in EBUS-TBNA cytological specimens is feasible and presents good reproducibility when compared with routine histological samples. EBUS-TBNA cytological samples could be used for the assessment of PD-L1 expression in patients with NSCLC as a minimally invasive approach in stage IV NSCLC cancer patients.

Keywords: Ultrasonography; Biopsy, needle; Lung neoplasms; Molecular targeted therapy.

Objective: To assess children and adolescents diagnosed with mild intermittent or mild persistent asthma, in terms of their aerobic capacity between attacks. Methods: We included 33 children and adolescents recently diagnosed with asthma (mild intermittent or mild persistent) and 36 healthy children and adolescents. Those with asthma were evaluated between attacks. All participants underwent clinical evaluation; assessment of baseline physical activity level; pre- and post-bronchodilator spirometry; and a maximal exercise test, including determination of maximal voluntary ventilation, maximal oxygen uptake, respiratory quotient, maximal minute ventilation, ventilatory equivalent, ventilatory reserve, maximal HR, SpO2, and serum lactate. Results: No significant differences were found among the groups (intermittent asthma, persistent asthma, and control) regarding anthropometric or spirometric variables. There were no significant differences among the groups regarding the variables studied during the maximal exercise test. Conclusions: A diagnosis of mild intermittent/persistent asthma has no effect on the aerobic capacity of children and adolescents between asthma attacks.

Keywords: : Asthma; Exercise; Respiratory function tests.

Objective: To study the effect of perillyl alcohol on the gene expression of human pulmonary adenocarcinoma cells. Methods: Pulmonary adenocarcinoma cells were incubated with perillyl alcohol in dilutions ranging from 0.03% to 0.0003% for 48 hours. Alterations were observed in the cell morphology, and cell viability was quantified using [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] assays. Protein synthesis of samples previously targeted with S35 was analyzed using electrophoresis on a polyacrylamide gel. Expression of the proteins p53 and p44/42 was determined using the Western blot method. Results: After 48 hours of incubation, greater numbers of morphological alterations were observed in cells treated with the 0.03% perillyl alcohol dilution than in those treated with perillyl alcohol diluted to 0.003% or further. Treatment with perillyl alcohol dilutions of 0.03%, 0.003% and 0.0003% inhibited cellular viability by 60.17% (p < 0.001), 15.62% (p < 0.001) and 11.53% (p < 0.05), respectively. The results show that 28-kDa, 42-kDa and 110-kDa proteins were induced. No statistically significant effect on p53 expression was observed. In comparison with the expression of a-tubulin, the 0.003% perillyl alcohol dilution induced an increase in p42 phosphorylation and a marked decrease in p44 phosphorylation. Conclusion: The results suggest that there are other, previously undescribed, metabolic pathways for perillyl alcohol effects in human pulmonary adenocarcinoma cells.

Keywords: Adenocarcinoma; Lung neoplasms; Monoterpenes; Cell culture; Lung/cytology

Silica is one of the main occupational agents associated with lung cancer. It is classified by the International Agency for Research on Cancer (IARC) as a Group 1 substance, i.e., carcinogenic for humans, since 1996. The objective of this paper is to report two cases of workers previously exposed to silica who developed lung cancer, seen in an out-patient clinic, focusing on important points of the current literature on the subject.

Objective: To assess the incidence of necrotizing pneumonia (NP) in children submitted to thoracoscopy, comparing patients with and without NP in terms of the presentation and clinical evolution. Methods: A retrospective study of children with pleural empyema submitted to thoracoscopy. Thoracoscopy was performed in patients not previously submitted to thoracic drainage and in whom there was evidence of loculated effusion or pneumothorax, as well as in those previously submitted to thoracic drainage and in whom there was persistent pneumothorax or fever with purulent discharge. On the basis of the thoracoscopy findings, patients were distributed into two groups: those with NP (NP group) and those without (no-NP group). Results: The study sample comprised 52 patients. Of the 24 patients with NP, 19 (79%) had undergone thoracic drainage prior to thoracoscopy, 11 (46%) presented with pneumothorax, and 16 (67%) developed bronchopleural fistula. In the NP group, the median drainage time and the median length of hospital stay were 18 and 19 days, respectively. Of the 28 patients without NP, 10 (36%) had undergone thoracic drainage prior to thoracoscopy, 9 (32%) presented pneumothorax, and 5 (18%) developed bronchopleural fistula. In the no-NP group, the median drainage time and the median length of hospital stay were 6 and 10 days, respectively. Conclusions: Pneumothorax should raise the suspicion of NP. Early thoracoscopy can be a valuable treatment option for NP in children because it hastens recovery in comparison with the medical treatment alone and avoids extensive late thoracotomy lung resections.

Keywords: Keywords: Empyema, pleural; Thoracoscopy; Pneumonia; Child.