Maristela Peres Rangel

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. Subjects 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, Spearmans 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.

Caryocar brasiliense camb protects against genomic and oxidative damage in urethane-induced lung carcinogenesis

The antioxidant effects of Caryocar brasiliense Camb, commonly known as the pequi fruit, have not been evaluated to determine their protective effects against oxidative damage in lung carcinogenesis. In the present study, we evaluated the role of pequi fruit against urethane-induced DNA damage and oxidative stress in forty 8-12 week old male BALB/C mice. An in vivo comet assay was performed to assess DNA damage in lung tissues and changes in lipid peroxidation and redox cycle antioxidants were monitored for oxidative stress. Prior supplementation with pequi oil or its extract (15 µL, 60 days) significantly reduced urethane-induced oxidative stress. A protective effect against DNA damage was associated with the modulation of lipid peroxidation and low protein and gene expression of nitric oxide synthase. These findings suggest that the intake of pequi fruit might protect against in vivo genotoxicity and oxidative stress.

Tissue hyaluronan expression, as reflected in the sputum of lung cancer patients, is an indicator of malignancy

Hyaluronan (HA) shows promise for detecting cancerous change in pleural effusion and urine. However, there is uncertainty about the localization of HA in tumor tissue and its relationship with different histological types and other components of the extracellular matrix, such as angiogenesis. We evaluated the association between HA and degree of malignancy through expression in lung tumor tissue and sputum. Tumoral tissue had significantly increased HA compared to normal tissue. Strong HA staining intensity associated with cancer cells was significant in squamous cell carcinoma compared to adenocarcinoma and large cell carcinoma. A significant direct association was found between tumors with a high percentage of HA and MVD (microvessel density) in tumoral stroma. Similarly significant was the direct association between N1 tumors and high levels of HA in cancer cells. Cox multivariate analysis showed significant association between better survival and low HA. HA increased in sputum from lung cancer patients compared to cancer-free and healthy volunteers and a significant correlation was found between HA in sputum and HA in cancer tissue. Localization of HA in tumor tissue was related to malignancy and reflected in sputum, making this an emerging factor for an important diagnostic procedure in patients suspected to have lung cancer. Further study in additional patients in a randomized prospective trial is required to finalize these results and to validate our quantitative assessment of HA, as well as to couple it to gold standard sputum cytology.

Modeling pulmonary fibrosis by abnormal expression of telomerase/apoptosis/collagen V in experimental usual interstitial pneumonia

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.

Biomolecular analysis of matrix proteoglycans as biomarkers in non small cell lung cancer

Matrix proteoglycans (PGs) have shown promise as biomarker in malignancies. We employed agarose gel eletrophoresis, quantitative real- time reverse transcription-polymerase chain reaction and immunohistochemistry to evaluate the content of sulfated glicosaminoglycans (chondroitin sulfate and heparan sulfate) and expression of PG (biglycan, glypican, perlecan, syndecan e versican) in patient-matched normal and tumor tissues obtained from resected specimens of lung cancer. A significant increase of heparan sulfate (HS) and chondroitin sulfate (CS) concentrations was found in tumor tissue samples when compared to normal lung tissue samples. HS was also significantly increased in adenocarcinomas compared to squamous cell carcinomas. PG gene expression, with exception of syndecan, were significantly decreased in tumor tissue compared to normal lung, coinciding with significant decrease of PG protein levels in tumor cells and stroma compared to normal lung tissue (Kappa coefficient 0.41, 0.42 and 0,28, respectively). Women patients (p = 0.02), non smokers (p = 0.05), T stage (p = 0.009), N stage (p = 0.03) and adenocarcinoma (p = 0.05) were associated with improved overall survival (OS). Patients presenting tumors with low concentration of sulfated GAG and high PGs levels presented better OS compared to patients with high concentration of sulfated GAG and low expression of PGs. Cox regression model controlled by gender, tobacco history and histological type, showed that patients with high perlecan and versican expression in tumor presented respectively high probability of life (β risk 11.64; 1.27 to 15.90) and low risk of death (β risk 0.11; 0.02–0.51). The combined approach suggest matrix (PGs) as biomarkers in lung cancer.

Immunohistochemical and morphometric evaluation of COX 1 and COX-2 in the remodeled lung in idiopathic pulmonary fibrosis and systemic sclerosis

OBJECTIVE: To study the expression of COX-1 and COX-2 in the remodeled lung in systemic sclerosis (SSc) and idiopathic pulmonary fibrosis (IPF) patients, correlating that expression with patient survival. METHODS: We examined open lung biopsy specimens from 24 SSc patients and 30 IPF patients, using normal lung tissue as a control. The histological patterns included fibrotic nonspecific interstitial pneumonia (NSIP) in SSc patients and usual interstitial pneumonia (UIP) in IPF patients. We used immunohistochemistry and histomorphometry to evaluate the expression of COX-1 and COX-2 in alveolar septa, vessels, and bronchioles. We then correlated that expression with pulmonary function test results and evaluated its impact on patient survival. RESULTS: The expression of COX-1 and COX-2 in alveolar septa was significantly higher in IPF-UIP and SSc-NSIP lung tissue than in the control tissue. No difference was found between IPF-UIP and SSc-NSIP tissue regarding COX-1 and COX-2 expression. Multivariate analysis based on the Cox regression model showed that the factors associated with a low risk of death were younger age, high DLCO/alveolar volume, IPF, and high COX-1 expression in alveolar septa, whereas those associated with a high risk of death were advanced age, low DLCO/alveolar volume, SSc (with NSIP), and low COX-1 expression in alveolar septa. CONCLUSIONS: Our findings suggest that strategies aimed at preventing low COX-1 synthesis will have a greater impact on SSc, whereas those aimed at preventing high COX-2 synthesis will have a greater impact on IPF. However, prospective randomized clinical trials are needed in order to confirm that.

Abstract 1604: Treatment with Pequi (Caryocar brasiliense Camb) fruit pulp oil decrease the DNA damage caused by chemical carcinogenesis in experimental model of lung cancer.

Proceedings: AACR 103rd Annual Meeting 2012‐‐ Mar 31‐Apr 4, 2012; Chicago, IL Background: It is well known that a daily consumption of foods that are rich in antioxidants prevents the oxidative damage caused by reactive species (ROS), including DNA damage, and can reduce the risk of cancer, atherosclerosis and other degenerative diseases. The pulp of Caryocar brasiliense Camb, most known as pequi, is a Brazilian fruit that has high levels of antoxidants properties, such as vitamin C, carotenoids, phenolic compounds like flavonoids, saponins and tannins, and essential oils. The aim of this study was to evaluate the action of the pequi oil in the DNA damage in experimental model of lung cancer induced by the urethane. Material and Methods: Male BALB/c mice (n=14) received by gavage 0,5μL/mg/day of pequi oil (CBC oil) during 75 days. After 15 days of the beginning of the gavage, 10 of these animals received two doses of 1,5g/kg intraperitoneal of urethane (Urethane + CBC oil). Urethane animals (n=4) were only submitted to two doses of 1,5g/kg intraperitoneal of urethane (urethane group). After these 75 days, lung tissues were submitted to histological and biochemical analysis by comet assay method to evaluated the DNA damage. The DNA damage was quantificate by image analysis. Results: The lung parenchyma from the urethane groups without oil and with oil showed neoplasic formations induced by the chemical carcinogenesis in contrast with oil control group. The image analysis of the comet assay showed a statistical significant diminish of the DNA damage cells in the urethane oil group when compared with urethane control group (p=0.001). The decreased DNA damage was very similar that we obtained in the oil control group. Conclusion: We conclude that the different antioxidant components found in the pequi oil diminish the DNA damage in the mouse lung after chemical carcinogenesis, suggesting that this type of strategies may have a greater impact in lung cancer treatment. Financial Support: FAPESP, CNPq Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 1604. doi:1538-7445.AM2012-1604

Abstract 427: Impact of glycosaminoglycans on lung cancer diagnosis and prognosis of patients

Introduction: Treatment of lung cancer patients depends on histological type, performance status and clinical stage. Because of limitations of these parameters in predicting disease behavior, different markers have been investigated to better discriminate lung cancer evolution. Glycosaminoglycans(GAGs) are important molecules of the extracellular matrix hydrating and maintaining the osmotic balance of tissues. Many have reported that GAGs have different behaviors when in the presence of malignant tissues. In this study, we sought to examine the sulfated GAGs and hyaluronan (HA) concentrations and their impact on diagnosis and/or prognosis of patients with non small cell lung cancer. Methods: Sulfated GAGs and HA were examined in tumoral and non-tumoral tissues from 45 patients who underwent lobectomy for lung cancer. Their preoperative clinical stages were T 1-3 N 0-1 M 0 and the mean follow-up was 19.3 months. Their histologic types were adenocarcinomas (n=23), squamous cells carcinomas (SqCC) (n=16) and large cell carcinoma (n=6). Tissue samples were dehydrated with acetone and incubated with a proteolytic enzyme. The HA chains were diluted (1:100) in blocking buffer [0.05M Tris-HCl, pH 7.4, 1% BSA]. The levels of HA were measured by a noncompetitive enzyme-linked immunosorbent assay (ELISA)-like fluorometric assay. The sulfated GAG chains (heparan, dermatan and chondroitin sulfate – HS, DS and CS), were precipitated with ethanol and the pellet was dried and dissolved in DNAse(5μl/mg).The different types of sulfated GAGs and their concentration were identified after gel electrophoresis in diaminopropane buffer. Results: A distinct profile of sulfated GAGs and HA was observed between non-tumoral and tumoral areas. HS and HA showed significantly higher concentration in tumoral than in non-tumoral areas (p=0.02 and p=0.0001, respectively). The adenocarcinomas had higher amounts of HS than SqCC specimens(p=0.02). One hundred % of tumoral areas presented CS while the normal almost never (p Conclusion: The presence of CS and higher concentrations of HS and HA in patients with lung cancer suggest a possible role of those molecules in this disease, but more importantly provide potential biochemical markers for differentiating normal from lung cancer patients. The correlation between the histologic types and the amounts of HS provide a possible role of this GAG on the development of tumor aggressiveness considering that one of its functions is to bind itself to growth factors and regulate their action. However, further studies are needed to determine whether or not these concentrations are able to be diagnostic/prognostic markers of lung cancer. Financial support: FAPESP Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 427. doi:10.1158/1538-7445.AM2011-427