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The common epidermal growth factor receptor (EGFR) mutations, such as the L858R point mutation in exon 21 and the in-frame deletional mutation in exon 19, have been definitively associated with response to EGFR-tyrosine kinase inhibitors (EGFR-TKI). However, the clinical outcome and response to treatment for many other rarer mutations are still unclear. In this study, we report the results of Brazilian patients in stage IB-IIIA non-small cell lung cancer (NSCLC) following complete resection with minimal residual disease and EGFR mutations treated with adjuvant chemotherapy and/or EGFR-TKIs. The frequency of EGFR mutations was investigated in 70 cases of early stage NSCLC. Mutations in exons 18 and 20, uncommon mutations in exons 19 and 21, as well as in exons 3, 7, 14, 16, 22, 27, and 28, and/or the presence of different mutations in a single tumor (complex mutations) are considered rare. EGFR mutations were detected in 23 tumors (32.9%). Fourteen cases carried rare mutations and were treated with platinum-based chemotherapy and two cases were treated with erlotinib. The clinical outcome is described case by case with references to the literature. Notably, we found two rare EGFR mutations and one of them with an unknown response to chemotherapy and/or EGFR-TKIs. We have provided complementary information concerning the clinical outcome and treatment of patients with early stage NSCLC for several rare EGFR mutations not previously or only rarely reported. Description of cases harboring rare mutations can support the decision-making process in this subset of patients.

The vascular network expansion and functioning are important factors affecting normal intra-uterine fetal development. This study addressed the previously reported antiangiogenic potential of beta-2-glycoprotein I (β2GPI) in vivo in the chick embryo model of angiogenesis. The effects of two naturally occurring β2GPI forms on the development of the chorioallantoic membrane (CAM) vessels and the chicken embryo were investigated. β2GPI monomers and dimers were obtained by fractioned purification and characterized using SDS-PAGE, immunoblot, and ELISA. The egg exposure was performed by injection of small volumes of 2.5 µg/mL solutions of the β2GPI subfractions. Angiogenesis was evaluated through quantitative measurements of vascular architecture parameters in the captured CAM images, using computational analysis of texture contrasts and computer vision techniques. Quantitative information was assigned to the CAM vasculature modifications. In vivo, the β2GPI dimer completely halted the formation of CAM vessels and led to embryo death after 48 h of exposure. The β2GPI monomer allowed the embryo to develop up to the 10th day, despite early changes of CAM vessels. The impaired normal vessel growth proceeded as a self-limited effect. The β2GPI monomer-exposed eggs showed reduced vascularization on the 6th day of incubation, but embryos were viable on the 10th day of incubation, with ingurgitated CAM vessels implying sequelae of the angiogenesis inhibition. Both subfractions impaired CAM vasculature development. The β2GPI dimer proved to be largely more harmful than the β2GPI monomer. β2GPI modification by cleavage or dimerization may play a role in angiogenesis control in vivo.

Background: Variable ventilation improves respiratory function, but it is not known whether the amount of variability in tidal volume (VT) can be reduced in recruited lungs without a deterioration of respiratory system elastance., Methods: Acute lung inflammation was induced by intratracheal instillation of lipopolysaccharide in 35 Wistar rats. Twenty-eight animals were anaesthetized and ventilated in volume-controlled mode. Lungs were recruited by random variation of VT (mean 6 ml kg(-1), coefficient of variation 30%, normal distribution) for 30 min. Animals were randomly assigned to different amounts of VT variability (n=7 for 90 min per group): 30, 15, 7.5, or 0%. Lung function, diffuse alveolar damage, and gene expression of biological markers associated with cell mechanical stress, inflammation, and fibrogenesis were assessed. Seven animals were not ventilated and served as controls for post-mortem analyses., Results: A VT variability of 30%, but not 15, 7.5, or 0%, prevented deterioration of respiratory system elastance [Mean (SD) -7.5 (8.7%), P<0.05; 21.1 (9.6%), P<0.05; 43.3 (25.9), P<0.05; and 41.2 (16.4), P<0.05, respectively]. Diffuse alveolar damage was lower with a VT variability of 30% than with 0% and without ventilation, because of reduced oedema and haemorrhage. A VT variability of 30, 15, or 7.5% reduced the gene expression of amphiregulin, cytokine-induced neutrophil chemoattractant-1, and tumour necrosis factor α compared with a VT variability of 0%., Conclusions: In this model of acute lung inflammation, a VT variability of 30%, compared with 15 and 7.5%, was necessary to avoid deterioration of respiratory system elastance and was not associated with lung histological damage., (© The Author 2016. Published by Oxford University Press on behalf of the British Journal of Anaesthesia. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

We collected a series of 136 lung/bronchial and 56 matched lung parenchyma tissue samples from patients who underwent lung/bronchial biopsies and presented invasive carcinoma after lung surgery. The lung/bronchial samples included basal cell hyperplasia, squamous metaplasia, moderate dysplasia, adenomatous hyperplasia, severe dysplasia, squamous cell carcinoma and adenocarcinoma. Matched lung parenchyma tissue samples included 25 squamous cell carcinomas and 31 adenocarcinomas. Immunohistochemistry was performed to analyze for the distribution of hyaluronidase (Hyal)-1 and -3, and hyaluronan synthases (HAS)-1, -2, and -3. Hyal-1 showed significantly higher expression in basal cell hyperplasia than in moderate dysplasia (P=0.01), atypical adenomatous hyperplasia (P=0.0001), or severe dysplasia (P=0.03). Lower expression of Hyal-3 was found in atypical adenomatous hyperplasia than in basal cell hyperplasia (P=0.01) or moderate dysplasia (P=0.02). HAS-2 was significantly higher in severe dysplasia (P=0.002) and in squamous metaplasia (P=0.04) compared with basal cell hyperplasia. HAS-3 was significantly expressed in basal cell hyperplasia compared with atypical adenomatous hyperplasia (P=0.05) and severe dysplasia (P=0.02). Lower expression of HAS-3 was found in severe dysplasia compared with squamous metaplasia (P=0.01) and moderate dysplasia (P=0.01). Epithelial Hyal-1 and -3 and HAS-1, -2, and -3 expressions were significantly higher in pre-neoplastic lesions than in neoplastic lesions. Comparative Cox multivariate analysis controlled by N stage and histologic tumor type showed that patients with high HAS-3 expression in pre-neoplastic cells obtained by lung/bronchial biopsy presented a significantly higher risk of death (HR=1.19; P=0.04). We concluded that localization of Hyal and HAS in lung/bronchial pre-neoplastic and neoplastic lesions was inversely related to malignancy, which implied that visualizing these factors could be a useful diagnostic procedure for suspected lung cancer. Finalizing this conclusion will require a wider study in a randomized and prospective trial.

We investigated the effects of acute hypercapnic acidosis and buffered hypercapnia on lung inflammation and apoptosis in experimental acute lung injury (ALI). Twenty-four hours after paraquat injection, 28 Wistar rats were randomized into four groups (n=7/group): (1) normocapnia (NC, PaCO2=35-45 mmHg), ventilated with 0.03%CO2+21%O2+balancedN2; (2) hypercapnic acidosis (HC, PaCO2=60-70 mmHg), ventilated with 5%CO2+21%O2+balancedN2; and (3) buffered hypercapnic acidosis (BHC), ventilated with 5%CO2+21%O2+balancedN2 and treated with sodium bicarbonate (8.4%). The remaining seven animals were not mechanically ventilated (NV). The mRNA expression of interleukin (IL)-6 (p=0.003), IL-1β (p<0.001), and type III procollagen (PCIII) (p=0.001) in lung tissue was more reduced in the HC group in comparison with NC, with no significant differences between HC and BHC. Lung and kidney cell apoptosis was reduced in HC and BHC in comparison with NC and NV. In conclusion, in this experimental ALI model, hypercapnia, regardless of acidosis, reduced lung inflammation and lung and kidney cell apoptosis., (Copyright © 2014 Elsevier B.V. All rights reserved.)

Limitations on tissue proliferation capacity determined by telomerase/apoptosis balance have been implicated in pathogenesis of idiopathic pulmonary fibrosis. In addition, collagen V shows promise as an inductor of apoptosis. We evaluated the quantitative relationship between the telomerase/apoptosis index, collagen V synthesis, and epithelial/fibroblast replication in mice exposed to butylated hydroxytoluene (BHT) at high oxygen concentration. Two groups of mice were analyzed: 20 mice received BHT, and 10 control mice received corn oil. Telomerase expression, apoptosis, collagen I, III, and V fibers, and hydroxyproline were evaluated by immunohistochemistry, in situ detection of apoptosis, electron microscopy, immunofluorescence, and histomorphometry. Electron microscopy confirmed the presence of increased alveolar epithelial cells type 1 (AEC1) in apoptosis. Immunostaining showed increased nuclear expression of telomerase in AEC type 2 (AEC2) between normal and chronic scarring areas of usual interstitial pneumonia (UIP). Control lungs and normal areas from UIP lungs showed weak green birefringence of type I and III collagens in the alveolar wall and type V collagen in the basement membrane of alveolar capillaries. The increase in collagen V was greater than collagens I and III in scarring areas of UIP. A significant direct association was found between collagen V and AEC2 apoptosis. We concluded that telomerase, collagen V fiber density, and apoptosis evaluation in experimental UIP offers the potential to control reepithelization of alveolar septa and fibroblast proliferation. Strategies aimed at preventing high rates of collagen V synthesis, or local responses to high rates of cell apoptosis, may have a significant impact in pulmonary fibrosis.

Idiopathic interstitial pneumonias include complex diseases that have a strong interaction between genetic makeup and environmental factors. However, in many cases, no infectious agent can be demonstrated, and these clinical diseases rapidly progress to death. Theoretically, idiopathic interstitial pneumonias could be caused by the Epstein-Barr virus, cytomegalovirus, adenovirus, hepatitis C virus, respiratory syncytial virus, and herpesvirus, which may be present in such small amounts or such configuration that routine histopathological analysis or viral culture techniques cannot detect them. To test the hypothesis that immunohistochemistry provides more accurate results than the mere histological demonstration of viral inclusions, this method was applied to 37 open lung biopsies obtained from patients with idiopathic interstitial pneumonias. As a result, immunohistochemistry detected measles virus and cytomegalovirus in diffuse alveolar damage-related histological patterns of acute exacerbation of idiopathic pulmonary fibrosis and nonspecific interstitial pneumonia in 38 and 10% of the cases, respectively. Alveolar epithelium infection by cytomegalovirus was observed in 25% of organizing pneumonia patterns. These findings were coincident with nuclear cytopathic effects but without demonstration of cytomegalovirus inclusions. These data indicate that diffuse alveolar damage-related cytomegalovirus or measles virus infections enhance lung injury, and a direct involvement of these viruses in diffuse alveolar damage-related histological patterns is likely. Immunohistochemistry was more sensitive than the histological demonstration of cytomegalovirus or measles virus inclusions. We concluded that all patients with diffuse alveolar damage-related histological patterns should be investigated for cytomegalovirus and measles virus using sensitive immunohistochemistry in conjunction with routine procedures.

Background: Ultra High Dilutions (UHD) are diluted beyond the Avogadro limit with dynamization (dilution with succussion). The process of anuran amphibian metamorphosis is controlled by thyroid hormones, including the resorption of the tadpole tail., Methods: A randomized and blinded study was performed to investigate the influence of triiodothyronine (T3) 5·10(-24)M (10cH) on apoptosis induced by T3 100 nM in Rana catesbeiana tadpoles' tail tips, in vitro. Explants were randomized to three groups: control: no T3 in pharmacological or UHD dose; test: T3 100 nM and challenged with T3 10cH (UHD); positive control: T3 100 nM, treated with unsuccussed ethanol. The apoptotic index and the area of explants of test and control groups at the first and final day of the experiment were compared by t-test., Results: There was no difference in tail tip area between test and control groups, but a significantly higher (p<0.01) index of apoptosis in explants of the test group., Conclusion: This data suggest that T3 10cH modifies the effect of T3 at pharmacological dose, opening new perspectives for further studies and investigation of the dose-effect curve., (Copyright © 2011 The Faculty of Homeopathy. Published by Elsevier Ltd. All rights reserved.)

Pulmonary remodeling is an important feature of asthma physiopathology that can contribute to irreversible changes in lung function. Although neurokinins influence lung inflammation, their exact role in the extracellular matrix (ECM) remodeling remains to be determined. Our objective was to investigate whether inactivation of capsaicin-sensitive nerves modulates pulmonary ECM remodeling in animals with chronic lung inflammation. After 14 days of capsaicin (50 mg/kg, sc) or vehicle administration, male Hartley guinea pigs weighing 250-300 g were submitted to seven inhalations of increasing doses of ovalbumin (1, 2.5, and 5 mg/mL) or saline for 4 weeks. Seventy-two hours after the seventh inhalation, animals were anesthetized and mechanically ventilated and the lung mechanics and collagen and elastic fiber content in the airways, vessels and lung parenchyma were evaluated. Ovalbumin-exposed animals presented increasing collagen and elastic fiber content, respectively, in the airways (9.2 ± 0.9; 13.8 ± 1.2), vessels (19.8 ± 0.8; 13.4 ± 0.5) and lung parenchyma (9.2 ± 0.9; 13.8 ± 1.2) compared to control (P < 0.05). Capsaicin treatment reduced collagen and elastic fibers, respectively, in airways (1.7 ± 1.1; 7.9 ± 1.5), vessels (2.8 ± 1.1; 4.4 ± 1.1) and lung tissue (2.8 ± 1.1; 4.4 ± 1.1) of ovalbumin-exposed animals (P < 0.05). These findings were positively correlated with lung mechanical responses to antigenic challenge (P < 0.05). In conclusion, inactivation of capsaicin-sensitive nerve fibers reduces pulmonary remodeling, particularly collagen and elastic fibers, which contributes to the attenuation of pulmonary functional parameters.

Setting: A tertiary care research centre in São Paolo, Brazil., Objective: To quantify interleukin (IL) 8, tumour necrosis factor alpha (TNF-alpha), vascular endothelial growth factor (VEGF) and transforming growth factor beta(1) (TGF-beta(1)) in pleural fluid from tuberculous patients, correlating its values with the histopathological patterns in pleural biopsies., Design: Cytokines were quantified in patients with transudates secondary to congestive heart failure (n = 8) and exudates secondary to tuberculosis (TB; n = 39). In parietal pleural biopsies from TB patients, the histological patterns of the inflammatory response were quantified by morphometric analysis (stereological point-counting method)., Results: IL-8, TNF-alpha, VEGF and TGF-beta(1) levels were higher in TB than in transudates. A positive correlation existed between components of the fibrinoid exudative phase with pleural fluid IL-8 (R = 0.52, P = 0.004) and VEGF (R = 0.42, P = 0.0021) levels. A negative correlation existed between pleural fluid IL-8 (R = -0.37, P = 0.048) and VEGF (R = -0.44, P = 0.0015) levels with tissue components of fibroproliferation., Conclusion: The high pleural levels of TNF-alpha, IL-8, VEGF and TGF-beta(1) suggest the involvement of these cytokines in the TB immunological response. The positive correlation between pleural fluid IL-8 and VEGF with the components of the acute exudative phase and the negative correlation between these cytokines with the fibroproliferative components suggest a temporary inflammatory response in the pleural space.

Intrapleural instillation of talc has been used in the treatment of recurrent pleural effusions but can, in rare instances, result in respiratory failure. Side-effects seem to be related to composition, size and inflammatory power of talc particles. The aim of this study was to evaluate the inflammatory response to intrapleural injection of talc containing small particles (ST) or talc containing particles of mixed size (MT). 100 rabbits received intrapleural talc, 50 with ST (median 6.41 mum) and 50 with MT (median 21.15 mum); the control group was composed of 35 rabbits. Cells, lactate dehydrogenase, C-reactive protein (CRP), interleukin (IL)-8 and vascular endothelial growth factor were evaluated in serum and bronchoalveolar lavage at 6, 24, 48, 72 and 96 h. Lung histology and the presence of talc were also analysed. Statistics were performed using ANOVA and an unpaired t-test. Most of the parameters showed greater levels in the animals injected with talc than in the controls, suggesting a systemic and pulmonary response. Higher serum levels of CRP and IL-8 were observed in the animals injected with ST. Talc particles were observed in both lungs with no differences between groups. Lung cell infiltrate was more evident in the ST group. In conclusion, talc with larger particles should be the preferred choice in clinical practice in order to induce safer pleurodesis.

The present study compared the effects of early short-term with prolonged low-dose corticosteroid therapy in acute lung injury (ALI). In total, 120 BALB/c mice were randomly divided into five groups. In the control group, saline was intratracheally (i.t.) instilled. In the ALI group, mice received Escherichia coli lipopolysaccharide (10 microg i.t.). ALI animals were further randomised into four subgroups to receive saline (0.1 mL i.v.) or methylprednisolone (2 mg x kg(-1) i.v.) at 6 h, 24 h or daily (for 7 days, beginning at day 1). At 1, 3 and 8 weeks, in vivo and in vitro lung mechanics and histology (light and electron microscopy), collagen and elastic fibre content, cytokines in bronchoalveolar lavage fluid and the expression of matrix metalloproteinase (MMP)-9 and -2 were measured. In vivo (static elastance and viscoelastic pressure) and in vitro (tissue elastance and resistance) lung mechanics, alveolar collapse, cell infiltration, collagen and elastic fibre content and the expression of MMP-9 and MMP-2 were increased in ALI at 1 week. Methylprednisolone led to a complete resolution of lung mechanics, avoided fibroelastogenesis and the increase in the expression of MMP-9 and MMP-2 independent of steroid treatment design. Thus, early short-term, low-dose methylprednisolone is as effective as prolonged therapy in acute lung injury.

Malignancy of pulmonary large cell carcinomas (LCC) increases from classic LCC through LCC with neuroendocrine morphology (LCCNM) to large cell neuroendocrine carcinomas (LCNEC). However, the histological classification has sometimes proved to be difficult. Because the malignancy of LCC is highly dependent on proteins with functions in the cell cycle, DNA repair, and apoptosis, p53 has been targeted as a potentially useful biological marker. p53 mutations in lung cancers have been shown to result in expression and protein expression also occurs in the absence of mutations. To validate the importance of both p53 protein expression (by immunostaining) and p53 gene mutations in lung LCC (by PCR-single strand conformational polymorphism analysis of exons 5, 6, 7, and 8) and to study their relationships with clinical factors and sub-classification we investigated the correlation of p53 abnormalities in 15 patients with LCC (5 classic LCC, 5 LCNEC, and 5 LCCNM) who had undergone resection with curative intent. Of these patients, 5/15 expressed p53 and none had mutant p53 sequences. There was a negative survival correlation with positive p53 immunostaining (P = 0.05). After adjustment for stage, age, gender, chemotherapy, radiotherapy, and histological subtypes by multivariate analysis, p53 expression had an independent impact on survival. The present study indicates that p53 assessment may provide an objective marker for the prognosis of LCC irrespective of morphological variants and suggests that p53 expression is important for outcome prediction in patients with the early stages of LCC. The results reported here should be considered to be initial results because tumors from only 15 patients were studied: 5 each from LCC, LCNEC and LCCNM. This was due to the rarity of these specific diseases.

One strand of research on the scientific basis of homeopathy is based on inversion effects of dilutions and the biophysical properties of information transfer. A model developed by Endler, was the basis for the study of the influence of high-diluted solution (1:1026 part by weight) of thyroid glands on the rate of metamorphosis of the frog Rana catesbeiana from the no legged to four-legged stage. The glands were obtained from tadpoles and prepared according by (dilution and succussion). Similar pure hydroalcoholic solution (unsuccussed) was used as control. In order to identify significant differences in the frequencies of four-legged tadpoles, in homeopathic and control group, we used a chi-square goodness-of-fit test (P<0.01) and the cumulative risk for metamorphosis by Cox's Proportional Hazards model (P<0.05). The number of animals that reached the four-legged stage is generally smaller in the treated group, than in the hydroalcoholic control group. It was postulated that thyroid hormones transmitted information' specific to the molecules used to prepare the solution, even though the molarity was beyond Avogadro's number.

The aim of this study was to determine whether an intrapleural injection of barium sulphate would produce pleurodesis in rats. Additionally, respiratory mechanics and pleural remodelling were analysed. Single intrapleural injection of barium sulphate (100%) or saline was given to Wistar rats. Respiratory system, lung, and chest wall elastic, resistive and viscoelastic/inhomogeneous pressures were measured by the end-inflation occlusion method at 2 and 30 days after injection. The pleura were examined for gross and histopathological evidence of pleural inflammation and fibrosis, and the underlying lungs were also studied by morphometry. All pulmonary mechanical parameters increased at day 2, but were not different from control at 30 days after injection. Chest wall mechanical parameters did not change. Macroscopic evaluation demonstrated pleural adherence without haemothorax. Histopathologic analysis showed pleural inflammation and fibrosis. There was no alveolar inflammation or fibrosis in both groups. In conclusion, barium sulphate induced pleurodesis with either no changes in respiratory mechanics or lung lesion at day 30.

Rounded atelectasis (RA) is a benign and unusual form of subpleural lung collapse that has been described mostly in asbestos-exposed workers. This form of atelectasis manifests as a lung nodule and can be confused with bronchogenic carcinoma upon conventional radiologic examination. The objective of the present study was to evaluate the variation in contrast uptake in computed tomography for the identification of asbestos-related RA in Brazil. Between January 1998 and December 2000, high-resolution computed tomography (HRCT) was performed in 1658 asbestos-exposed workers. The diagnosis was made in nine patients based on a history of prior asbestos exposure, the presence of characteristic (HRCT) findings and lesions unchanged in size over 2 years or more. In three of them the diagnosis was confirmed during surgery. The dynamic contrast enhancement study was modified to evaluate nodules and pulmonary masses. All nine patients with RA received iodide contrast according to weight. The average enhancement after iodide contrast was infused, reported as Hounsfield units (HU), increased from 62.5+/-9.7 to 125.4+/-20.7 (P < 0.05), with a mean enhancement of 62.5+/-19.7 (range 40 to 89) and with a uniform dense opacification. In conclusion, in this study all patients with RA showed contrast enhancement with uniform dense opacification. The main clinical implication of this finding is that this procedure does not permit differentiation between RA and malignant pulmonary neoplasm.

Acute respiratory distress syndrome (ARDS) can be derived from two pathogenetic pathways: a direct insult on lung cells (pulmonary ARDS (ARDSp)) or indirectly (extrapulmonary ARDS (ARDSexp)). This review reports and discusses differences in biochemical activation, histology, morphological aspects, respiratory mechanics and response to different ventilatory strategies between ARDSp and ARDSexp. In ARDSp the direct insult primarily affects the alveolar epithelium with a local alveolar inflammatory response while in ARDSexp the indirect insult affects the vascular endothelium by inflammatory mediators through the bloodstream. Radiological pattern in ARDSp is characterised by a prevalent alveolar consolidation while the ARDSexp by a prevalent ground-glass opacification. In ARDSp the lung elastance, while in ARDSexp the chest wall and intra-abdominal chest elastance are increased. The effects of positive end-expiratory pressure, recruitment manoeuvres and prone position are clearly greater in ARDSexp. Although these two types of acute respiratory distress syndrome have different pathogenic pathways, morphological aspects, respiratory mechanics, and different response to ventilatory strategies, at the present, is still not clear, if this distinction can really ameliorate the outcome.

Experimental and epidemiological evidence suggests that lycopene, a carotenoid present in tomatoes, tomato products, and several fruits and vegetables, may play a role in preventing certain cancers in humans. We have investigated the effect of lycopene pretreatment on lipid peroxidation, oxidative damage to DNA, and histopathological changes in liver of animals subjected to intraperitoneal (ip) ferric nitrilotriacetate (Fe-NTA) administration. Compared with control rats, liver of Fe-NTA-treated animals showed a significant increase in the 8-oxo-7,8-dihydro-2'-deoxyguanosine level and a 75% increase in malondialdehyde accumulation concomitant with histopathological changes. Five days of lycopene pretreatment (10 mg/kg body weight, ip) almost completely prevented liver biomolecule oxidative damage and protected the tissue against the observed histological alterations.

This work demonstrates that glycerol-preserved acellular allodermis can be used as support for the proliferation of human keratinocytes and that the characteristics of this bioengineered tissue suggest its possible use as a permanent skin substitute for therapeutic challenges such as extensive burns as well as its possible use as an in vitro model for pharmacological studies. The removal of all basal membrane components during preparation of the dermal support also provides an original in vitro situation that allows observation of the reorganization of the dermal-epidermal junction. The tissue composite obtained is constituted of dermis covered by a well attached, multistratified epithelium with morphological characteristics that resemble human epidermis as evidenced by light and transmission electron microscopy, including the neoformation, albeit incomplete, of the dermal-epidermal junction. Assessment of involucrin and cytokeratin 14 expression by immunohistochemical assays established differentiation patterns. Both immerse and air-liquid interface culture systems were tested.

Unlabelled: This study was undertaken to test whether there is structural remodeling of lung parenchyma that could lead to tissue mechanical changes at an early phase of varying degrees of acute lung injury (ALI). Tissue resistance (R), dynamic elastance (E), and hysteresivity (eta) were analyzed during sinusoidal oscillations of rat lung parenchymal strips 24 h after intraperitoneal injection of saline (C) or paraquat (P [10, 15, 25, and 30 mg/kg]). These strips were also stained in order to quantify the amount of collagen and of three types of elastic fibers (elaunin, oxytalan, and fully developed elastic fibers) in the alveolar septa. E augmented progressively from C to P25, but the data from the P25 and P30 groups were not different (p < 0.0001). R and eta increased from C to P10 and from P15 to P25 (p < 0.001). Collagen fiber content increased exponentially with the severity of the injury. Elaunin and fully developed elastic fibers remained unchanged in the five groups, while oxytalan fibers increased only in the P25 and P30 groups. In conclusion, the pronounced mechanical changes at the tissue level and fibroelastogenesis happened at an early phase of the disease and even in mildly abnormal lung parenchyma., Keywords: elastance; collagen fibers; elastin; paraquat

Asphyxia is a name given to different kinds of lesions that can produce similar histologic findings. Thus, because of the varied nature of the different kinds of lesions, as well as the incidence of similar qualitative histologic findings with different causes, the aim of this work was to study special kinds of injuries with particular subsequent impairment. These include some diagnostic problems of sudden death of natural causes, including aspiration, suffocation, drowning, and strangulation. Ranking was made of 167 victims based on the diagnosis as having: aspiration (n = 35), suffocation (n = 88), drowning (n = 27), and strangulation (n = 17). Stepwise discriminant analysis of the resulting data showed that lung necropsies from victims of these four events could be distinguished from one another. Statistical differences among the four groups were observed for eight morphologic parameters. A robust discriminant function permitted an adequate classification of the four groups of disease in 85.03% of the cases. Lung autopsies with congestion, septal hemorrhage, and foreign body showed a specificity of 100% for victims of aspiration, whereas ductal overinsufflation, interstitial edema, and bronchiolar constriction showed a specificity of 81.8% in victims of suffocation. Intraalveolar edema and dilatation of the alveolar spaces with secondary compression of the septal capillaries characterized drowning. Victims of strangulation showed a strong alveolar hemorrhage, with alveolar collapse and overinsufflation, associated with bronchiolar dilatation. It is concluded that semiquantitative analysis of lung autopsies might be a useful supplementary histologic criterion to support the diagnosis of asphyxia.

The dynamic mechanical properties of lung tissue and its contents of collagen and elastic fibers were studied in strips prepared from mice instilled intratracheally with saline (C) or silica [15 (S15) and 30 days (S30) after instillation]. Resistance, elastance, and hysteresivity were studied during oscillations at different frequencies on S15 and S30. Elastance increased from C to silica groups but was similar between S15 and S30. Resistance was augmented from C to S15 and S30 and was greater in S30 than in S15 at higher frequencies. Hysteresivity was higher in S30 than in C and S15. Silica groups presented a greater amount of collagen than did C. Elastic fiber content increased progressively along time. This increment was related to the higher amount of oxytalan fibers at 15 and 30 days, whereas elaunin and fully developed elastic fibers were augmented only at 30 days. Silicosis led not only to pulmonary fibrosis but also to fibroelastosis, thus assigning a major role to the elastic system in the silicotic lung.

The present study was undertaken in order to describe the morphological differences between pulmonary lesions in acute respiratory distress syndrome originating from direct pulmonary injury (ARDSp) and those originating from extrapulmonary injury (ARDSexp). We investigated a total of 38 ARDS-patients (27 males) ranging in age from 19 to 75 years, classified according to underlying disease in pulmonary (ARDSp) and extrapulmonary disease (ARDSexp). The extent of acute diffuse alveolar damage was assessed morphometrically on histologic gross sections in the upper and lower lobes of one lung. The lesions showed quantitative differences in extent and distribution according to underlying disease (primary pulmonary or secondary involvement). In pulmonary ARDS, a predominance of alveolar collapse (16.6%+/-12.3% versus 10.3%+/-11.9%, p = 0,03), fibrinous exudate (1.7%+/-3.2% versus 0.4%+/-1.1%, p = 0.01) and alveolar wall edema (11.2%+/-7.4% versus 6.6%+/-4.4%, p = 0,05) were found compared to extrapulmonary ARDS. We conclude that the morphology of acute diffuse alveolar damage (DAD) is mainly determined by underlying disease (pulmonary ARDS or extrapulmonary ARDS) differing in quantitative terms within the lung. Physiological, radiographic and respiratory system mechanics differences described in ARDSp and ARDSexp may therefore be due to morphometric differences in pulmonary lesions.

The aim of this study was to evaluate the effect of a chemical sclerosing agent, aluminum hydroxide, on pleural remodeling and on respiratory mechanics in rats. Saline (2 mL) or aluminum hydroxide [2 mL (0.15 g/mL)] was instilled intrapleurally in anesthetized male rats. The animals were studied 7 or 30 days after the instillation. Respiratory system, lung, and chest wall elastic, resistive, and viscoelastic/inhomogeneous pressures were measured by the end-inflation occlusion method. We studied the pleural remodeling process by means of semiquantitative analysis of the induced inflammation and quantitative analysis of the collagen extracellular matrix component. The effects on the underlying lung were analyzed morphometrically. Chest wall elastic and viscoelastic pressures increased after aluminum hydroxide instillation independent of time after instillation. Pleural inflammation was observed 7 days after instillation, while pleural adherence with a marked increase in the type I/type III collagen ratio was present 30 days after instillation. Histological examination demonstrated no differences in lung parenchyma among the groups. In conclusion, the present model describes the establishment of pleurodesis by aluminum hydroxide, which thwarts the normal chest wall mechanical profile without inducing any changes in the underlying lungs. The results were disclosed by both mechanical and morphological evaluation of the pleural remodeling.

Context: Oncocytomas are generally small and present slow growth. Finding of the tumor usually occurs incidentally. Their incidence is higher among male patients. Oncocytomas in mucous bronchial glands are extremely rare., Case Report: A 35-year-old male who presented bronchial oncocytoma. The tumor was found after bronchoscopy that investigated an atelectasis of the upper left lobe. Histological examination with optical microscopy revealed a mature neoplasm formed by ovoid cells with thin, granular, eosinophilic cytoplasm and small nuclei similar to oncocytes. Electron microscopy showed mitochondrial hyperplasia. A three-year follow-up after thoracotomy followed by lobectomy and removal of the bronchial tumor was uneventful.

Aminoguanidine is a drug known for more than a century, which has been attracting increasing interest in recent years due to the discovery of new pharmacological properties. This study investigated the effects of aminoguanidine on the fibrotic response induced by intratracheal administration of bleomycin to rats. Three groups of animals were studied: Group A (n = 19) corresponded to the control group. Group B (n = 20) received 10 IU/kg bleomycin intratracheal, and Group C (n = 12) received the same amount of bleomycin as Group B followed by 50 mg/kg/day aminoguanidine bicarbonate for 4 weeks. Aminoguanidine led to significant reductions in total hydroxyproline content of the lungs in Group C compared to Group B (Group A: 1.83+/-0.14 mg x Group B: 3.46+/-0.36 mg x Group C: 2.09+/-0.22 mg). Morphometric collagen studies carried out on histological sections stained with Sirius red F3BA showed that aminoguanidine promoted a significant reduction of the area occupied by collagen in the axial and septal zones of the lungs (Axial region = Group A: 4.29+/-1.31% x Group B: 19.30+/-4.86% x Group C: 8.52+/-1.96%; Septal region = Group A: 0.15+/-0.06% x Group B: 0.61+/-0.21% x Group C: 0.15+/-0.06%). These results suggest that aminoguanidine is a potential therapeutic agent for the treatment and prevention of pulmonary fibrosis which is associated with different clinical conditions.

Background: Tumor stage and its histological subtype remain the most important predictors of clinical behavior in current pulmonary practice of lung cancer. However, many investigators agree that these parameters are not sufficient to predict which tumor will recur, even after radical curative surgery. Therefore, it is necessary to evaluate the significance of other morphological, biological and molecular parameters beyond TNM classification., Methods: Pathological specimens were collected from 45 patients after resection for stage IA (five), stage IB (10), stage IIB (10), stage IIIA (14) and stage IV (six) lung adenocarcinomas. A panel of two morphological (proportion of stroma within the tumor and degree of tumor differentiation), two biological [DNA ploidy and argyrophilic nucleolar organizer region (AgNOR)] and three molecular (immunohistochemical expression of Ki-67, p53 and bcl-2) markers was chosen for analysis of the primary tumor. Life Tables for Survival were used to analyze the individual impact of each variable on survival. Cox proportional hazards model analysis was used to construct an independent tumor status model for cancer recurrence and death. Chi-squared analyses were used to determine the statistically significant relationship among all the variables present in the study., Results: Multivariate analysis demonstrated statistically significant risk for the following markers: AgNOR, p53 and bcl-2, controlled for stages and surgical resection., Conclusions: The immunohistochemical expression of p53 and bcl-2 oncogenes and the expression of AgNOR cell proliferation index are critical values in the progression of lung adenocarcinomas. They can express the biological tumor status and indicate a more accurate prognosis.

Data from 64 patients who underwent surgical resection of lung adenocarcinomas were studied to identify clinicopathologic markers that might provide prognostic information on the clinical behavior of this neoplasia Patient staging was performed in accordance with the tumor-node-metastasis system as follows: Stage I (n = 29), Stage II (n = 11), Stage IIIA (n = 21), and Stage IIIB (n = 3). Overall follow-up time corresponded to the follow-up time for patients who were alive and to the survival time for patients who had died, all of them expressed in months. Data included age, staging, histologic type, morphometric assessment of histologic features related to tumor (stroma and vascularization), and immunohistochemical detection of proliferation cell markers (Ki-67 protein and proliferating cell nuclear antigen) and p53 protein. The morphometric assessment was made by the point-counting procedure. Data analysis included Life Tables for Survival and Cox Regression models. Overall follow-up analysis showed that significant univariate predictors (P < .05) were T stage; N stage; tumor stromal proportion; and immunohistochemical indexes of proliferating cell nuclear antigen, Ki-67, and p53 proteins. Variables that presented independent predictive value for overall follow-up with the multivariate model (P < .05) were sex, T stage, N stage, tumor stromal proportion, and immunohistochemical detection of p53 protein. We conclude that tumor stromal proportion and immunohistochemical detection of p53 protein, controlled for sex, T stage, and N stage, may be of critical value in the evaluation of recurrence of lung adenocarcinoma, serving as indicators for a more accurate prognosis.

The pathogenic mechanisms of lipopolysaccharide (LPS)-induced lung injury have not been classified. This study examined the physiological changes after endotoxin inhalation and related those to features of pulmonary inflammation in mice. Pulmonary mechanics, histopathology, and bronchoalveolar lavage fluid (BALF) from BALB/c mice were analysed at different occasions (3, 24, 48 and 72 h) after inhalation of saline or LPS from Escherichia coli (0.3 (L0.3) or 10 mg x mL(-1) (L10)). Mice were sedated, anaesthetized, and ventilated. After chest wall resection static (Est) and dynamic (Edyn) elastances, deltaE (Edyn-Est), resistive (deltaP1) and viscoelastic/inhomogeneous pressures (deltaP2), and deltaP1+deltaP2 (deltaPtot) were obtained by end-inflation occlusion method. Lungs were prepared for histopathology. In parallel groups, tumour necrosis factor (TNF)-alpha, neutrophils, and protein were evaluated in the BALF. L0.3 and L10 showed a time-dependent production of TNF-alpha preceding a massive neutrophil infiltration. In L10 BALF there was an increase in protein level at 24 and 48 h. Est and Edyn increased early in L0.3 (65%, 63%) and L10 (41%, 51%). In L10 deltaE, deltaP2, and deltaPtot showed a gradual rise. At 72 h all groups were similar. L0.3 showed an early increase in cellularity, which returned to normal at 72 h. L10 presented the same pattern with the cell count remaining elevated until 72 h. In conclusion, lipopolysaccharide inhalation led to elastic and viscoelastic pulmonary changes together with tumour necrosis factor-alpha production and neutrophil infiltration in mouse lung.

Respiratory mechanics, chest wall configuration, and lung morphometry were determined in rats before and at 30 (PTX.30) and 60 (PTX.60) min after pneumothorax induction (intrathoracic injection of 8 ml of room air; 50% collapse). Pneumothorax increased respiratory system and lung elastances and viscoelastic/inhomogeneous pressures in both groups, but respiratory system and lung resistive pressures increased only in PTX.60 group. Antero-posterior diameters at the third intercostal space and xiphoid levels, circumference at xiphoid level, and thoracic cephalo-caudal diameter increased significantly after pneumothorax induction independently of temporal evolution. In both groups lung collapse, hyperinflation, and interstitial and alveolar edema were present. Additionally, in PTX.60 group the central airways calibre diminished in relation to PTX.30. In conclusion pneumothorax yields changes in respiratory system and lung elastic and viscoelastic parameters, which are related to alveolar collapse and edema, respectively. Temporal evolution of pneumothorax also leads to changes in lung resistive pressure, probably because of airway narrowing.

This paper investigates the effects of air pollution in urethane-induced lung tumours in mice by means of histological, morphometrical, and DNA ploidy. The experimental exposure was done in locations with different air pollution profiles: a polluted area (downtown São Paulo) and a "clean" environment. Swiss mice were employed and urethane (3 g/kg) was used as a carcinogenic substance. All the animals, whether exposed or not to air pollution, were sacrificed after 6 months, and the lung lesions were analysed. The results showed a significant effect of air pollution on tumour progression, observed by changes in the phenotype of the tumour cells as demonstrated by morphometry and DNA ploidy. We observed more atypical adenomas in the air pollution-exposed group (p = 0.02). Coherently, morphometric differences were also detected between the two groups. Neoplasms of exposed mice exhibited an increase in the nuclear fraction (p = 0.002) and in the nucleus/cytoplasm ratio (p = 0.011), as a decrease in the stromal fraction (p < 0.001). There was a higher risk of aneuploidy in the 6-months-of-air-pollution-exposure group (relative risk: 1.58; 95% of confidence interval: 1.007 to 2.403). These results indicate that urban air pollution accelerates the process of progression towards malignancy.

Study Objectives: Many reports have shown the efficacy of talc to induce an effective pleurodesis. However, there is little information about the side effects related to this sclerosing agent. The objective of this experimental study is to recognize the systemic distribution of talc after its instillation into the pleural space of rats., Design: Forty animals were assigned to receive talc through a catheter placed in a left minimal thoracotomy. They were randomly divided in two groups: group 1 received 20 mg of talc and group 2 received 10 mg in the same total volume of 1 mL of saline solution. Half of the animals in each group were killed 24 h and the other half 48 h after the procedure. BAL was collected and histologic sections of both lungs, chest wall, liver, kidneys, spleen, heart, and brain were examined. Crystals were tracked using polarized light and we have used a "birefringent particles index of deposition" in an attempt to quantify the amount or talc encountered in different organs., Results: Talc crystals were found in every organ of all animals studied (100%). There was no statistical difference either on the dose of talc used or in the time of death. The amount of talc was statistically different in the organs, which made us divagate about a route of absorption., Conclusions: We conclude that there is a progressive deposition of talc particles in the organs examined after its administration into the pleural space of normal rats. This report suggests that there is a rapid absorption of talc through the pleural surface and that the systemic distribution thereafter is not dose related. Further studies are necessary to assess the amount of crystals and the clinical correlation to these findings.

Unlabelled: It has been postulated that regional anesthesia, when feasible, is the best anesthetic approach in asthmatic patients. However, there are reports of severe bronchospasm during regional anesthesia. In the present study, we developed an experimental model of spinal (subarachnoid) anesthesia in guinea pigs and studied respiratory system responsiveness to aerosolized methacholine. The animals received sodium pentobarbital (50 mg/kg intraperitoneally), a tracheotomy, and mechanical ventilation. Four groups of animals were studied: guinea pigs that received spinal anesthesia with lidocaine (500 microL of 2% solution) (n = 7); guinea pigs that received spinal administration of isotonic sodium chloride solution (500 microL) (n = 7); guinea pigs that received an intraperitoneal injection of lidocaine (500 microL of 2% solution) (n = 6); and control guinea pigs (n = 7). The concentration of methacholine chloride that resulted in 50% of the maximal value of respiratory system elastance was lower in guinea pigs that received spinal anesthesia compared with the other three groups (P < 0.005 for control group, P < 0.01 for spinal saline group, and P < 0.05 for intraperitoneal lidocaine group). Our results suggest that spinal anesthesia results in an increase in pulmonary responsiveness to bronchoconstrictive stimuli., Implications: Regional anesthesia has been considered the best anesthetic approach in asthmatic patients, although there are reports of severe bronchospasm. We developed an experimental model of spinal anesthesia with lidocaine in guinea pigs and studied respiratory responsiveness to methacholine, a bronchoactive agonist. Spinal anesthesia resulted in an increase in respiratory responsiveness.

Study Objectives: To evaluate the potential associations between long-term exposure to air pollution and histopathologic evidence of damage to the lungs in humans., Design: Lung tissue samples were collected during necropsies of individuals who died due to violent causes, selected on the basis of their exposure background., Patients: The exposed group was composed of individuals who lived in Guarulhos, an area with high mean levels of inhalable particles. The control group was composed of individuals who lived in two cities with economies based on agricultural activities: Ribeirão Preto and Ourinhos., Interventions: Information about cigarette smoking and occupational exposure was obtained from family members., Measurements and Results: Morphometric evaluation of the main bronchus was conducted to determine the volume ratio of submucosal glands. Histopathologic alterations of the bronchioli were evaluated by scoring the presence of inflammatory reaction, wall thickening, and secretory hyperplasia. The number of spots of carbon deposition was counted along the regions of lymphatic drainage (visceral pleura and axial connective tissue around bronchi and blood vessels). Statistical analysis was done by means of regression models controlled for age, smoking, and occupational exposure. Lungs collected from the high pollution area presented evidence of more histopathologic damage in comparison to those from the clean environments. These effects were observed even after controlling for individual differences in age, sex, and cigarette smoking levels., Conclusions: These results suggest that long-term exposure to air pollution may contribute to the pathogenesis of airway disease, and that urban levels of air pollution have adverse effects on the respiratory tract.

The biologic behavior of tumoral cells plays a significant role in the progression of the neoplasia, because 30 to 35% of patients with Stage I squamous cell carcinoma relapse. The present study was designed to determine whether age, pathologic parameters, DNA ploidy, and a cell proliferation index (the area of nucleolar organizer regions, AgNOR), could be used to predict survival in patients who undergo resection for limited squamous cell carcinoma of the lung. For histopathologic analysis, the parameters of histologic grading, pleural involvement, vascular invasion, and residual disease were considered. The cell proliferation index was evaluated by mitotic index, AgNOR quantification, and DNA ploidy by means of digital image analysis. Fifty-two patients (median age, 60 yr +/- 8.6 yr) were staged according to the TNM staging system. Cox univariate analysis showed that stage, residual disease, vascular invasion, histologic grading, DNA ploidy, and AgNOR were significant predictors of survival. Many of the univariate predictors of cancer death, however were eliminated when Cox multivariate models were computed. The variable that exhibited the most robust predictive value for overall survival was AgNOR. We conclude that measurement of cell proliferation might serve as a prognostic marker in squamous cell carcinoma of the lung.

The expression of the nucleolar organizer regions (NORs) was quantified in paraffin sections of tumors and lymph node metastasis, by means of digital image analysis, in 75 patients with resected non-small cell lung cancer (NSCLC). Patients were divided in two groups: early stage (stages I and II) and advanced stage (stages IIIa, IIIb and IV). The prognostic significance of AgNOR expression was tested by Cox regression analysis in models controlled for age, sex, vital status, stage and histological type. Tumors at early stages had a lower expression of AgNOR than those at more advanced diseases. The mean values obtained for NORs in advanced disease were almost the same as those in the primary tumors when compared with the corresponding lymph node metastasis (r = 0.90; p < 0.01; linear regression). The prognostic role of AgNOR was significant only for tumors at stages I and II and not for advanced neoplasms (stages IIIa, IIIb and IV). These results encourage the inclusion of AgNOR quantitation in routine material, especially in early lung cancer.

Background: Malignant fibrous histiocytoma (MFH) arising primarily in the lungs is rare, and a preoperative diagnosis, as well as a surgical planning, is very important because of the tumor's propensity for vascular invasion and its low incidence of lymph node metastasis. The correct preoperative diagnosis of thoracic MFH is not easy to establish because the small fragments from needle and transbronchial biopsies are often inadequate for a conclusive histologic analysis. A preoperative bronchial brushing cytology suggestion of the diagnosis of primary MFH of the lungs may be helpful in such cases., Case: A 37-year-old male presented with a large, irregular mass in the inferior and middle lobes on chest roentgenography as well as on computed tomography. Two bronchoscopies were performed, with the diagnosis of undifferentiated large cell carcinoma. After surgical resection a more sophisticated pathologic analysis, including immunohistochemical and ultrastructural studies, revealed a primary MFH of the lungs. Revision of the bronchial brushing cytology disclosed many spindle-shaped cells with a "comet" configuration, strongly suggestive of MFH., Conclusion: The bronchial brushing cytology features of spindle-shaped and bizarre, multinucleated giant cells with a comet appearance may be the key to the cytomorphologic diagnosis of MFH.

Nucleolar organizer regions identified by means of an argyrophilic technique (AgNOR) were quantified by digital image analysis in 81 patients with squamous cell carcinoma of the lung. Survival rate was modeled as function of AgNOR expression by Cox regression models controlled for staging, histologic grade, age, and sex. Our results indicate that AgNOR expression has a significant prognostic role. Tumors at lower stages have a lower expression of AgNOR than those with more advanced disease. Tumors with high histopathologic grade have a higher expression of AgNOR. Patients with low AgNOR expression presented a higher survival rate than those with higher amounts of AgNOR. The foregoing results strongly encourage the development of prospective trials to assess the real role of AgNOR in determining prognosis in squamous cell carcinoma of the lung.