Expressão de fos após pulso de escuro no núcleo pré- geniculado do tálamo do sagui (Callithrix jacchus)

The pregeniculate nucleus (PGN) of the primate s thalamus is an agglomerate neuronal having a cap shaped located dorsomedially to the main relay visual information to the cerebral cortex, the dorsal lateral geniculate nucleus (GLD). Several cytoarchitectonic, neurochemical and retinal projections studies have pointed PGN as a structure homologous to intergeniculate leaflet (IGL) of rodents. The IGL receives retinal terminals and appears to be involved in the integration of photic and non-photic information relaying them, through geniculo-hypothalamic tract (TGH), to the main circadian oscillator in mammals, the suprachiasmatic nucleus (SCN) of the hypothalamus. Thus, the IGL participates in the control of the biological rhythm by modulating the activity of the SCN. Pharmacological and IGL injury studies conclude that it is critical in the processing of non-photic information which is transmitted to the SCN. Other studies have found that especially neurons immunoreactive to neuropeptide Y (NPY) respond to this type of stimulation, determined by its colocation with the FOS protein. Has not been determined if the PGN responds, expressing the FOS protein, to the non-photic stimulus nor the neurochemical nature of these cells. Thus, we apply a dark pulse in the specifics circadian phases and analyze the pattern of expression of FOS protein in PGN of the marmoset (Callithrix jacchus). We found that in all animals analyzed the FOS expression was higher in the experimental than in the control group. There was a higher expression of FOS when the dark pulse was applied during the subjective day between the groups. Still, a subregion of the PGN, known by immunoreactive to NPY, had a greater number of FOS-positive cells in relation to his other just close dorsal region. Our data corroborate the theory that the PGN and IGL are homologous structures that were anatomically modified during the evolutionary process, but kept its main neurochemical and functional characteristics. However, injury and hodological studies are still needed for a more accurate conclusion

Avaliação comportamental e neuroquímica de ratos em dessincronização forçada: possíveis implicações para um modelo animal de oscilações no humor

Bipolar disorder has been growing in several countries. It is a disease with high mortality and has been responsible by the social isolation of the patients. Bipolar patients have alterations in circadian timing system, showing a phase shift in various physiological variables. There are several arguments demonstrating alterations in circadian rhythms may be part of the bipolar disorder pathophysiology. Given the necessity for further elucidation, the goal of this study was to validate the forced desynchronization protocol as an animal model for bipolar disorder. To do this, Wistar rats were submitted to a forced desynchronization protocol which consists in a symmetrical light dark cycle with 22h. Under this protocol, rats dissociate the locomotor activity rhythm into two components: one synchronized to the light / dark cycle with 22h, and another component with period longer than 24 hours following the animal endogenous period. These rhythms with different periods sometimes there is coincidence, which we named CAP (Coincidence Active Phase) and the opposite phase, non-coincidence, called NCAP (Non-Concidence Active Phase). The hypothesis is that in CAP animals present a mania-like behavior and animals in NCAP depressive-like behavior. We found some evidence described in detail throughout this thesis. In sum, the animals under forced desynchronization protocol were more stressed, showed an increase in stereotypic behaviors such as grooming and reduction in other behaviors such as risk assessment and vertical exploration when compared to the control group. The CAP animals showed increased locomotor activity, especially during the dark phase when compared to controls (rats under T24) and less depressive behavior in the forced swim test. The animals in NCAP showed a higher anxiety in elevated plus maze, but they don t have ahnedonia. The animals under dissociation have more labeled 5HT1A cells at the amygdala area, which appoint that they have more amygdala inhibition. Taking these data together, we could partially validated the forced desynchronization protocol as an animal model for mood oscillations

Efeitos da estimulação ambiental sobre os aspectos motores, cognitivos e neuronais em um modelo farmacológico progressivo da doença de Parkinson

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior

Avaliação do caráter oxidante da violaceína

Violacein is a violet pigment isolated from many gram-negative bacteria, especially from Chromobacterium violaceum, a betaproteobacterium found in the Amazon River in Brazil. It has potential medical applications as an antibacterial, fungicide, anti-tryptanocidal, anti-ulcerogenic and anti-cancer drug, among others. Furthermore, its pro-oxidant activity has been suggested, but only in two specific tumor lineages. Thus, in the present study, the prooxidant effects of violacein were investigated in both normal and tumor cells, seeking to evaluate the cell responses. The evaluation of violacein cytotoxicity using the Trypan blue dye exclusion method indicated that CHO-K1 cells were more resistant than tumor HeLa cells. The oxidative stress induced by violacein was manifested as an increase in intracellular SOD activity in CHO-K1 and MRC-5 cells at a specific concentration range. Nevertheless, a decrease was detected specifically at 6-12 μM in HeLa and MRC-5 cells. Interestingly, the increase in SOD activity was not followed by a concomitant increase in catalase activity. Regarding to oxidative stress biomarkers, increased protein carbonylation and lipid hydroperoxides levels were detected respectively in CHO-K1 and MRC-5 cells treated with violacein at 1.5-3 μM and 3 μM, which may be an evidence that this compound causes oxidative stress specifically in these conditions. Additionally, it is believed that the decline in cell viability observed in MRC-5 cells and HeLa treated with violacein at 6-12 M is due to mechanisms not related to oxidative stress. Moreover, the results suggested that violacein might cause oxidative stress by increasing endogenous levels of O2 -, since the occurrence of an expressive change in SOD activity. In addition, in order to evaluate the antioxidant activity of violacein in the absence of a biological system, the total antioxidant and iron chelating activity were evaluated, so that antioxidant activities were detected at 30 and 60 μM of violacein. Altogether, the results indicate that although oxidative stress is triggered by incubation with violacein, it did not seem to be high enough to cause serious damage to cell biomolecules in HeLa cells and only at specific concentrations in CHOK-1 and MRC-5 cells. Comparing the results obtained in cell culture and the in vitro antioxidant activity evaluation, the results confirmed that violacein presents opposing oxidant features when in presence or absence of a biological system and the antioxidant character only occurs at high concentrations of the pigment.

Xilanas de sabugo de milho como agente antioxidante, citotóxico, anticoagulante e imunomodulador

Prospecting pharmacological active polysaccharides from agricultural byproducts, such as corncobs, is an underexplored practice in the scientific community. Thus, this work aims to expand knowledge about pharmacological activities of polysaccharides extracted from corncobs. From corn cob flour a extract was obtained by ultrasound waves in an alkaline medium, and the end of the process the product was termed PECC (polysaccharidic extract from corncobs). This extract was physicochemical characterized and evaluated by in vitro assays as an antioxidant, cytotoxic, anticoagulant and imunomodulator agent. Results indicated significant activity metal chelating by PECC, and the use of PECC in cell culture cells showed no toxic effects to normal cell lines, but toxic action against HeLa tumor cells due promoting cell death by apoptosis. In addition, other pharmacological effects were observed, the PECC decreased nitric oxide (NO) production by activated macrophages, and prolonged blood clotting time through APTT assay. Then methanolic, ethanolic and ketone fractions were obtained from fractionation of PECC polysaccharides. Five methanolic fractions, six ethanolic fractions and two ketones were obtained; and all fractions were evaluated for antioxidant, cytotoxic, anticoagulant, immunomodulatory activities. E1.4 fraction exhibited significant metal chelating effect, a toxic action to induce apoptosis in HeLa cells, decreased NO production by activated macrophages, and extended blood clotting time. These results showed that the PECC pharmacological active polysaccharides would be present in the fraction E1.4. From fractionation of E1.4 polysaccharide six subfractions with different sizes were obtained: <3; 3-10; 10-30; 30-50; 50-100 and >100 KDa. About 80% of E1.4 polysaccharides had lower size to 10 KDa, and all the subfractions showed over 61% sugar in their chemical compositions. These subfractions exhibited different monosaccharide compositions, but xylose was presented in all of them. The subfractions exhibited distinct pharmacological effects in in vitro assays. Smaller subfractions (<30 KDa) had highest metal chelating activity and greater toxic action in tumor cells. The intermediate fractions (between 30-100 KDa) decreased more NO production of activated macrophages, for other side, the larger size (>100 KDa) modulated a greater number of inflammatory cytokines, and the had greatest anticoagulant effect. Therefore, when analyzing all the results together it is evident that the PECC pharmacological polysaccharides are heteroxylans, and were concentrated in E1.4 fraction, and heteroxilanas pharmacological effects depends on their molecular size. Thus, corncobs could be used as source from molecules with biotechnology potential

Modelo animal da Doença de Parkinson baseado na expressão de alfa - sinucleína: caracterização comportamental, eletrofisiológica e avaliação dos efei tos da estimulação da medula espinhal

Parkinson disease (PD) is associated with motor symptoms and dopaminergic cell loss in the nigrostriatal pathway. Alpha-synuclein is the major component of the Lewy bodies, the biological hallmarks of disease, and has been associated with familial cases of PD. Recently, the spinal cord stimulation (SCS) showed to be effective to alleviate the Parkinson symptoms in animal models and human patients. In this project, we characterized the motor and electrophysiological effects of alpha-synuclein overexpression in the substantia nigra of rats. We further investigated the effects of spinal electrical stimulation, AMPT and L-dopa administration in this model. Method: Sprague-Dawley rats were injected with empty viral vector or the vector carrying the gene for alpha-synuclein in the substantia nigra, and were tested weekly for 10 weeks in the open field and cylinder tests. A separated group of animals implanted with bilateral electrode arrays in the motor cortex and the striatum were recorded in the open field, during the SCS sessions and the pharmacological experiments. Results: Alpha-synuclein expression resulted in motor asymmetry, observed as the reduction in use of contralateral forepaw in the cylinder test. Animals showed an increase of local field potential activity in beta band three and four weeks after the virus injection, that was not evident after the 5th week. AMPT resulted in a sever parkinsonian state, with reduction in the locomotor activity and significant peak of oscillatory activity in cortex and striatum. SCS was effective to alleviate the motor asymmetry at long term, but did not reduce the corticostriatal low frequency oscillations observed 24 hs after the AMPT administration. These oscillations were attenuated by L-dopa that, even as SCS, was not effective to restore the locomotor activity during the severe dopaminergic depletion period. Discussion: The alpha-synuclein model reproduces the motor impairment and the progressive neurodegenerative process of PD. We demonstrated, by the first time, that this model also presents the increase in low frequency oscillatory activity in the corticostriatal circuit, compatible with parkinsonian condition; and that SCS has a therapeutic effect on motor symptom of this model.

Avaliação imuno-histoquímica das proteínas BMP-4, FGF-8 e Sindecan-1 em tumores odontogênicos

The development and progression of odontogenic tumors have been associated with an imbalance in the activity of growth factors, adhesion molecules, extracellular matrix proteins and their degradation enzymes, angiogenic factors and osteolytic. Some studies have shown that interaction relationships inductive epithelial / mesenchymal determinants of Odontogenesis are mimicked by these tumors. The objective of this research was to investigate the immunolocalization of growth factors (BMP-4 and FGF-8) and Sindecan-1 structural protein in a series of odontogenic tumors presenting different biological behaviors, to contribute to a better understanding of the role of these proteins in tumor development. The sample consisted of 21 of the solid ameloblastoma, odontogenic keratocysts 19 and 14 odontogenic adenomatoid tumors. Increased Sindecan-1 immunostaining was seen in the epithelium of the lesions when compared with mesenchyme. In ameloblastoma and odontogenic keratocysts, this expression was higher than in AOT. Epithelial expression of BMP4 showed quantitatively similar in the three studied lesions; however, when anlisada mesenchymal immunoreactivity, was detected significant higher expression when compared to the ameloblastoma keratocysts. In ameloblastoma, mesenchymal expression was predominantly (p = 0.008), while in keratocyst higher expression in the epithelium was observed (p = 0.046). In all injuries, strong or moderate correlation was observed in the BMP-4 immunoreactivity in the epithelium and mesenchyme. FGF-8, no injury was observed difference between the immunoreactivity in the epithelium or mesenchyme, however in ameloblastoma positive correlation was found (Spearman correlation, rho = 0.857, p <0.001). The results of this study suggest that the three evaluated biomarkers actively involved in the pathogenesis of lesions, especially the expression of ameloblastomas indicating a strong interaction between parenchymal and stromal cells which may contribute to its marked aggressiveness.

Atividade toxicológica do extrato das folhas de Kalanchoe brasiliensis em camundongos Swiss

The species of the genus Kalanchoe (Crassulaceae) are widely used in Brazil as a medicinal product to treat cough, boils, gastritis and other diseases. In this scenario stands out K. brasiliensis, popularly known as coirama or saião. This paper aims at the oral exposure of mice to a hydroalcoholic extract of leaves of Kalanchoe brasiliensis. The animals (total of 100) were divided into 12 groups (6 males and 6 females) for the acute assessment; and groups of 10 for subchronic evaluation. Test groups were treated with doses of 250 mg / kg, 500 mg / kg, 1000 mg / kg and 2000 mg / kg and the control group received 0.9% saline. The animals were observed for 14 days for acute evaluation and 30 days for subchronic evaluation, and in that period analyzed the appearance of clinical signs, changes in weight and consumption of water and food. After the observation period, histopathological analysis of the organs, biochemical serum and haematological parameters for the assessment of the subchronic groups were processed. Differences and changes in body weight were not observed among the groups, nor consumption of water and food, there were no deaths among the groups in the two types of assays. Histopathological analysis showed some alterations compatible with low acute hepatic toxicity. The results of blood glucose, triglycerides, ALT, urea and creatinine showed differences between the control group and the test concentrations studied (p <0.05), but these differences do not show relevant change in the clinical picture of animals. The results showed that the extract K. brasiliensis has low acute toxicity at the doses used and no toxicity when administered for 30 days. This highlights the importance of this species as future and promising candidate for phytotherapics, so has its pharmacological trials completed.

Spondias tuberosa Arr. (UMBU): estudo fitoquímico e avaliação do potencial anti-inflamatório

Spondias tuberosa Arruda (Anacardiaceae) is a fruitful tree popularly known as umbuzeiro, tapereba or umbu. It is a native and endemic species from Brazil, widespread in Brazilian Northeast. The species is important in folk medicine of the semi-arid Northeast, where it is mainly used to treat various inflammatory conditions, digestive problems as well as viral and bacterial infections. However, despite the common use in folk medicine, there are scarce pharmacological and phytochemicals studies that afford scientific evidence to its popular use. Therefore, this study aimed to characterize the chemical markers in S. tuberosa leaves extract, obtained by maceration ethanol:water (70:30, [v/v]), and evaluate its anti-inflammatory potential in vivo. The phytochemical profile in TLC analysis suggested the occurence of the flavonoids rutin and isoquercitrin. HPLC analysis enabled us to confirm the presence of flavonoids and also, were detected the phenolic acids, chlorogenic acid and caffeic acid. In addition was developed and validated a HPLC method to evaluate the content of the identified compounds in S. tuberosa leaves extract according to RDC 899/2003 of ANVISA and ICH Guidelines 2005. In order to evaluate the anti-inflammatory potential of S. tuberosa leaves extract, the peritonitis and paw edema models induced by carrageenan were used, administration i.p. in mice. The results highlighted the anti-inflammatory property in vivo at 125, 250 and 500 mg/kg since a decrease in leukocyte influx to the site of inflammation, diameter of the edema and the level of myeloperoxidase were observed when compared to the drug control dexamethasone (2 mg/kg, i.p. route). Taken together, the results pointed out S. tuberosa as a potential species for developing phytotherapic derivatives in according to its popular use. With regard to the characterization markers, chlorogenic acid, caffeic acid, rutin and isoquercitrin were identified and quantified in Spondias tuberosa leaves extract so they could be used in quality control analyses of the raw material and extracts of this species.

Alterações hepáticas na expressão gênica e atividade da catalase e superóxido dimutase em ratos diabéticos induzidos por estreptozootocina

The correlation between the type 1 diabetes mellitus and oxidative stress have been described in several studies, however its underlying mechanisms are not fully elucidated. The present work aimed to evaluate the effects of four weeks of streptozootocin-induced (STZ) diabetes in the redox homeostasis of rat hepatocytes. Thus, the liver of male Wistar rats from control and diabetic groups were collected and the activity and expression of antioxidant enzymes, as well the main markers of oxidative stress and content of H2O2 in these tissues were measured. The diabetes induced the activity of superoxide dismutase (SOD) and the gene expression of its mitochondrial isoform, SOD2. However, the expression of SOD1, the cytoplasmic isoform, was reduced by this disease. The activity and expression of catalase (CAT), as well the expression of glutathione peroxidase 1 (GPX1) and peroxiredoxin 4 (PRX4) were drastically reduced in the hepatocytes of diabetics rats. Even with this debility in the peroxidases mRNA expression, the content of H2O2 was reduced in the liver of diabetics rats when compared to the control group. The diabetes caused an increase of lipid peroxidation and a decrease of protein thiol content, showing that this disease causes distinct oxidative effects in different cell biomolecules. Our results indicate that four week of diabetes induced by STZ is already enough to compromise the enzymatic antioxidant systems of the hepatocytes.

Correlação da imunoexpressão do fator de choque térmico 1 (hsf1) com aspectos clinicopatológicos em carcinomas de células escamosas de língua oral

Squamous cell carcinoma of oral tongue shows high rates of morbidity and mortality in the population, therefore, great efforts are being made to classify morphological changes and identify biomarkers that have prognostic value and that are able to group patients in individualized therapeutic options. From this perspective, there is the heat shock factor 1 (HSF1), which is a heat shock factor transcription protein (HSPs) that allows the cancer to deal with stressors associated with malignancy, acting differently in tumor progression. This research aimed to perform a clinico-pathological analysis of 70 cases of oral tongue squamous cell carcinoma (OTSCC) and immunohistochemical study of the expression of HSF1 protein in OTSCC, comparing it with 30 specimens of normal oral mucosa (NOM), and correlating this immunostaining with clinico-pathological aspects of OTSCC. To analyze the association between immunoexpression of HSF1 and clinicophatoloical aspects, the cases were categorized in minor and major overexpression, based in the median immunostaining score. Regarding the cases of OTSCC, 57.1% showed clinical stage III or IV, 82.9% were graded as high grade according to Bryne (1998) and 47.1% as high risk of malignancy according to Brandwein-Gensler et al., (2005). A disease free survival rate of 47.84% and overall survival rate of 68.20% was observed in the analyzed cases, and the high degree of malignancy according to Bryne’s system (1998) (p=0.05), tumor size T3 or T4 (p=0.04), local recurrence (p=0.02), and perineural invasion (p=0.02) determined negative impacts in survival time. We observed also a statistically significant result (p<0.01) when comparing the immunoreactivity of HSF1 between NOM and OTSCC. This significantly increased expression of HSF1 in cases of OTSCC suggests that this protein acts, indeed, in the pathogenesis of this disease. However, there were no statistically significant associations between this overexpression and the clinico-pathological parameters analyzed. This finding may reflect the influence of epigenetic events on HSF1 gene or a possible stability of this protein expression throughout disease progression.

Correlação da imunoexpressão do fator de choque térmico 1 (hsf1) com aspectos clinicopatológicos em carcinomas de células escamosas de língua oral

Squamous cell carcinoma of oral tongue shows high rates of morbidity and mortality in the population, therefore, great efforts are being made to classify morphological changes and identify biomarkers that have prognostic value and that are able to group patients in individualized therapeutic options. From this perspective, there is the heat shock factor 1 (HSF1), which is a heat shock factor transcription protein (HSPs) that allows the cancer to deal with stressors associated with malignancy, acting differently in tumor progression. This research aimed to perform a clinico-pathological analysis of 70 cases of oral tongue squamous cell carcinoma (OTSCC) and immunohistochemical study of the expression of HSF1 protein in OTSCC, comparing it with 30 specimens of normal oral mucosa (NOM), and correlating this immunostaining with clinico-pathological aspects of OTSCC. To analyze the association between immunoexpression of HSF1 and clinicophatoloical aspects, the cases were categorized in minor and major overexpression, based in the median immunostaining score. Regarding the cases of OTSCC, 57.1% showed clinical stage III or IV, 82.9% were graded as high grade according to Bryne (1998) and 47.1% as high risk of malignancy according to Brandwein-Gensler et al., (2005). A disease free survival rate of 47.84% and overall survival rate of 68.20% was observed in the analyzed cases, and the high degree of malignancy according to Bryne’s system (1998) (p=0.05), tumor size T3 or T4 (p=0.04), local recurrence (p=0.02), and perineural invasion (p=0.02) determined negative impacts in survival time. We observed also a statistically significant result (p<0.01) when comparing the immunoreactivity of HSF1 between NOM and OTSCC. This significantly increased expression of HSF1 in cases of OTSCC suggests that this protein acts, indeed, in the pathogenesis of this disease. However, there were no statistically significant associations between this overexpression and the clinico-pathological parameters analyzed. This finding may reflect the influence of epigenetic events on HSF1 gene or a possible stability of this protein expression throughout disease progression.

Expressão imuno-histoquímica do ativador de plasminogênio do tipo uroquinase e seu receptor em carcinoma epidermoide de língua oral e sua relação com parâmetros clínico-patológicos

Squamous cell carcinoma (SCC ) is the most common malignancy of the oral cavity (OSCC), with a high mortality rate. Due to this, the discovery of biomarkers that facilitate the understanding of the biological behavior of the tumor and improve treatment is necessary. Urokinase type plasminogen activator (uPA) and its receptor, uPAR, are responsible for the proteolysis of structures of the basement membrana and extracellular matrix, facilitating tumor invasion. This study aims to assess the immuno expression of these proteins in 46 cases of squamous cell carcinoma of the oral tongue (OTSCC). These results were related to the presence of metastasis, clinical TNM staging, locoregional recurrence, outcome of the lesion and histological grading. Immunostaining of each case was evaluated semiquantitatively, in the front of invasion and center of the tumor, in which scores were assigned: 0 (0% of positive cells), 1 (1-10% of positive cells), 2 (11 -50% positive cells) and 3 (more than 50% positive cells). The expression of uPA was observed in 93.5% (n=43) of the cases in the front of invasion, with predominance of score 2 (n=16; 34.8%) and in 67.9% (n=31) of the cases in the center of the tumor, with predominance of score 1 (n=15; 32.6%). Overall, the immunoexpression of uPA was not associated with clinical parameters. Regarding the malignant histological grading, a higher expression of uPA was observed in cases of high-grade malignancy comp ared to low-grade malignancy (p=0.05). Regarding the morphological parameters, increased expression of uPA was observed in the worst mode of invasion (p=0.03 ). The expression of uPAR was observed in 73.9% of cases in the front of invasion, with a predominance of score 1 (n=21; 45.6 %), and in 47.5% (n=21) of the cases in the center of the tumor, with a predominance of score 0 (n=25; 54.4%). Although no statistical differences were observed in relation to lymph node metastasis, clinical TNM staging, outcome, and histological grading, there was a higher expression of uPAR in cases with locoregional recurrence (p=0.04). Regarding the tumor intra -localization, it was observed an increased expression of uPA and uPAR at the front of invasion in relation to the center of the tumor (p<0.001). Regarding the correlation between uPA and uPAR, there was no statistical sign ificance. Based on these results, it is suggested that uPA and uPAR are involved in the progression of CELO, mainly in the deeper region of the tumor.

Expressão imuno-histoquímica do ativador de plasminogênio do tipo uroquinase e seu receptor em carcinoma epidermoide de língua oral e sua relação com parâmetros clínico-patológicos

Squamous cell carcinoma (SCC ) is the most common malignancy of the oral cavity (OSCC), with a high mortality rate. Due to this, the discovery of biomarkers that facilitate the understanding of the biological behavior of the tumor and improve treatment is necessary. Urokinase type plasminogen activator (uPA) and its receptor, uPAR, are responsible for the proteolysis of structures of the basement membrana and extracellular matrix, facilitating tumor invasion. This study aims to assess the immuno expression of these proteins in 46 cases of squamous cell carcinoma of the oral tongue (OTSCC). These results were related to the presence of metastasis, clinical TNM staging, locoregional recurrence, outcome of the lesion and histological grading. Immunostaining of each case was evaluated semiquantitatively, in the front of invasion and center of the tumor, in which scores were assigned: 0 (0% of positive cells), 1 (1-10% of positive cells), 2 (11 -50% positive cells) and 3 (more than 50% positive cells). The expression of uPA was observed in 93.5% (n=43) of the cases in the front of invasion, with predominance of score 2 (n=16; 34.8%) and in 67.9% (n=31) of the cases in the center of the tumor, with predominance of score 1 (n=15; 32.6%). Overall, the immunoexpression of uPA was not associated with clinical parameters. Regarding the malignant histological grading, a higher expression of uPA was observed in cases of high-grade malignancy comp ared to low-grade malignancy (p=0.05). Regarding the morphological parameters, increased expression of uPA was observed in the worst mode of invasion (p=0.03 ). The expression of uPAR was observed in 73.9% of cases in the front of invasion, with a predominance of score 1 (n=21; 45.6 %), and in 47.5% (n=21) of the cases in the center of the tumor, with a predominance of score 0 (n=25; 54.4%). Although no statistical differences were observed in relation to lymph node metastasis, clinical TNM staging, outcome, and histological grading, there was a higher expression of uPAR in cases with locoregional recurrence (p=0.04). Regarding the tumor intra -localization, it was observed an increased expression of uPA and uPAR at the front of invasion in relation to the center of the tumor (p<0.001). Regarding the correlation between uPA and uPAR, there was no statistical sign ificance. Based on these results, it is suggested that uPA and uPAR are involved in the progression of CELO, mainly in the deeper region of the tumor.

Caracterização do material particulado e avaliação do risco ocupacional e mecanismos moleculares associados à queima artesanal da castanha de caju

Brazil is among the largest cashew nut producers of the world. However, the roasting process is still carried out artisanally, especially in the Brazilian semiarid region. In face of this occupational problem, the aim of this study was to perform a physical-chemical characterization of the particulate matter (PM) emitted by the roasting of cashew nuts, as well as to determine the occupational risk and molecular mechanisms associated. The most evident PM characteristics were the prevalence of fine particles, typical biomass burning morphologies such as tar ball and the presence of the elements K, Cl, S, Ca and Fe. In addition, atmospheric modeling analyses suggest that these particles can reach neighboring regions of the emission source. Polycyclic aromatic hydrocarbons (PAHs) with carcinogenic potential, such as benzo[a]pyrene, dibenz[a,h]anthracene, benzo[a]anthracene, benzo[b]fluoranthene, chrysene, benzo[k]fluoranthene, indeno[1,2,3-c,d]pyrene and benzo[j]fluoranthene were the most abundant PAHs found in the two air monitoring campaigns. Among the identified oxy-PAH the benzanthrone (7H-benz[d,e]anthracen-7-one) had the highest concentration and the evaluation of lifetime cancer risk showed an increase of 12 to 37 cases of cancer for every 10,000 exposed people. Chemical analysis of roasted cashew nuts identified the PAHs: phenanthrene, benzo[g,h,i]perylene, pyrene and benzo[a]pyrene, besides the 3-pentadecilfenol allergen (urushiol analogue) as prevalent. Occupational exposure to PAHs was confirmed by the increase of urinary 1-hydroxypyrene levels and genotoxic effects were evidenced by the increase on micronuclei and nuclear bud frequency in exfoliated buccal mucosa cells among the exposed workers. Other biomarkers of effects such as karyorrhexis, pyknotic, karyolytic, condensed chromatin and binucleated cells also have their frequencies increased when compared to an unexposed control group. The investigation of the molecular mechanisms associated with the PM organic extract showed cytotoxicity in human lung cell lines (A549) at concentrations ≥ 4 nM BaPeq. Using non-cytotoxic doses the extract was able to activate proteins involved in the DNA damage response pathway (Chk1 and p53). Moreover, the specific contribution of the four most representative PAHs in the cashew nut roasting sample showed that benzo[a]pyrene was the most efficient to activate Chk1 and p53. Finally, the organic extract was able to increase persistently the mRNA expression involved in the PAHs metabolism (CYP1A1 and CYP1B1), inflammatory response (IL-8 and TNF-α) and cell cycle arrest (CDKN1A) for DNA repair (DDB2). The high PM concentrations and its biological effects associated warn of the serious harmful effects of artisanal cashew nut roasting and urgent actions should be taken to the sustainable development of this activity.