Estudo da apoptose em pacientes infectados por Helicobacter Pylori portadores de doenças com diferentes riscos de desenvolvimento de câncer gástrico

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No. of bitstreams: 1 Tese Christiane Teixeira Cartelle.pdf: 6577282 bytes, checksum: aae857152af487888ca70cc9cb0808ad (MD5) Previous issue date: 2010-12-28A infecção crônica da mucosa gástrica por H. pylori é a principal causa da gastrite crônica (GC) implicado na patogênese da atrofia e da metaplasia intestinal (MI), condições de risco para o desenvolvimento do carcinoma gástrico (CaG). A infecção na infância determina risco de desenvolvimento do câncer gástrico justificando estudos comparados entre crianças e adultos. O microrganismo modifica o turnover da mucosa gástrica induzindo hiperproliferação e apoptose. No entanto, os mecanismos e vias operantes nos processos proliferativos e apoptóticos não são bem entendidos. Apesar de estudos mostrarem relação entre carcinogenese gástrica e expressão da sintase de óxido nítrico induzida (iNOS), ainda não se sabe qual a correlação entre inflamação, apoptose e ativação desta enzima nas condições de risco para transformação maligna associadas à infecção pelo H. pylori. Neste trabalho investigamos as vias apoptóticas nas gastrites, na mucosa gástrica de crianças e adultos e correlacionamos a intensidade e atividade da inflamação com expressão de iNOS e o índice apoptótico na mucosa gástrica em pacientes adultos com GC e GCA. Utilizamos material retrospectivo e prospectivo de crianças e adultos biopsiados para o esclarecimento de sintomas dispépticos. Amostras negativas e positivas para H. pylori foram classificadas de acordo com as patologias de interesse (UD, GC e GCA). Estudamos por imunohistoquímica a expressão de proteínas pró-e anti-apoptóticas das vias intrínsecas e extrínsecas da apoptose, bem como a proteína iNOS e o marcador M30 CytoDeath, e quantificamos esta expressão por morfometria digital. Em crianças e adultos a via preferencial é intrínseca e há expressão aumentada de proteínas pró-apoptóticas da via intrínseca (Bax e Bak) nos pacientes infectados, além de expressão compensatória de proteínas anti-apoptóticas (Bcl2 e Bclx). Além disso, Bak e Bax tiveram maior expressão nas GCA em relação aos demais diagnósticos levando a hipótese de que a apoptose nos casos de atrofia pode estar relacionada com a susceptibilidade ao câncer. O índice de expressão de iNOS foi maior nos pacientes H. pylori positivos que naqueles negativos. Não houve diferença nos índices apoptóticos entre pacientes H.pylori positivo ou negativo, ou na correlação estatística entre o índice apoptótico e o índice de expressão de iNOS nos pacientes infectados. Houve correlação entre o índice apoptótico e a intensidade da inflamação nos pacientes infectados.Chronic infection of the gastric mucosa by H. pylori is the main cause of chronic gastritis (CG) also implicated in the pathogenesis of atrophy and intestinal metaplasia (IM). These conditions are considered risk for developing gastric cancer (AGC) and it is assumed that infection in childhood increases risk to gastric cancer, so comparative studies between children and adults are justified. The organism modify the turnover of gastric mucosal proliferation and inducing apoptosis. However, the mechanisms and pathways in proliferative and apoptotic processes is not well understood. Although studies showed relationship between gastric carcinogenesis and expression of nitric oxide synthase induced (iNOS) it is still unknown if there is a correlation between inflammation, apoptosis and activation of this enzyme in terms of risk for the malignant transformation associated with infection by H. Pylori. In this study we investigated the apoptotic pathways in gastritis, gastric mucosa of children and adults and correlated the intensity of inflammation, iNOS expression and apoptotic index in gastric mucosa in adult patients with GC and GCA. We used retrospective and prospective material of children and adults biopsied for clarification of dyspeptic symptoms. Negative and positive samples for H. pylori were classified according to the pathologies of interest (UD, GC, and GCA). We studied by immunohistochemistry and quantified by digital morphometry the pro and anti-apoptotic protein expression of intrinsic and extrinsic pathways of apoptosis, as well as iNOS protein and the apoptosis marker M30 CytoDeath expression. In children and adults the preferred route is intrinsic and an increased expression of pro-apoptotic proteins of the intrinsic pathway (Bax and Bak), in addition to compensatory expression of anti-apoptotic proteins (Bcl2 and Bclx). Furthermore, Bax and Bak expression was higher in GCA compared to other diagnoses confirming the hypothesis that apoptosis can be related to susceptibility to cancer in long-term presence of mucosal atrophy. The iNOS expression was higher in H.pylori-negative patients than in the positive ones. There was no difference in apoptotic indices between patients H. Pylori-positive or negative or statistical correlation between the apoptotic index and the iNOS expression in patients H.Pylori-positive. A correlation between apoptotic index and the intensity of inflammation in infected patients was observed for mild and moderate degrees of gastritis

Leprosy classification methods: a comparative study in a referral center in Brazil

Objectives: Different methods for the classification of leprosy have been proposed since the 1930s. The aim of this study was to compare the current methods at a referral center in Brazil. Methods: The World Health Organization (WHO) operational classification was compared to the Ridley and Jopling classification, the Madrid classification, and a classification based on the number of body areas affected by skin and/or neural lesions (NBAA). The correlation between the clinical and histopathological components of the Ridley and Jopling classification was assessed. Results: The agreement between the WHO operational classification and the Ridley and Jopling classification was 77.6% (kappa = 0.53). The WHO operational classification tended to overestimate the number of multibacillary patients. The WHO operational classification showed its best agreement with the NBAA. There was perfect agreement between the clinical and histopathological Ridley and Jopling classification in 46.9% of the patients. Conclusions: The agreement between the WHO operational classification and the Ridley and Jopling classification was better than any other purely clinical classification, reinforcing the importance and simplicity of the operational method. Although major disagreement between the clinical and histopathological Ridley and Jopling classification was uncommon, perfect agreement occurred in less than half of the cases, and was even lower for the borderline lepromatous and tuberculoid forms. Possible reasons for the differences are discussed; these showed that there may be room for improvement in the Ridley and Jopling classification histopathological criteria

High-Fat Diet Increases HMGB1 Expression and Promotes Lung Inflammation in Mice Subjected to Mechanical Ventilation

This study aims to evaluate the effects of a high-fat diet and mechanical ventilation on the pulmonary and systemic inflammatory response in C57BL/6 mice. Male C57BL/6 mice were divided into two groups: one received a standard diet, and the other received a high-fat diet. After 10 weeks, the groups were further divided into two groups each: control group (CG), mechanical ventilation group (MVG), diet group (DG), and diet mechanical ventilation group (DMVG). MVG and DMVG underwent mechanical ventilation for 60 minutes. All animals were euthanized for subsequent analysis. Animals receiving a high-fat diet presented higher body mass, adipose index, and greater adipocyte area. In the lung, the expression of HMGB1 was greater in DG and DMVG than in CG and MVG. CCL2 and IL-22 levels in MVG and DMVG were increased compared to those in CG and DG, whereas IL-10 and IL-17 were decreased. Superoxide dismutase activity was higher in MVG and DMVG than in CG. Catalase activity was lower in DG than in CG, and in MV groups, it was lower than that in CG and DG. MV and obesity promote inflammation and pulmonary oxidative stress in adult C57BL/6 mice

Correction: Enteric Neuronal Damage, Intramuscular Denervation and Smooth Muscle Phenotype Changes as Mechanisms of Chagasic Megacolon: Evidence from a Long-Term Murine Model of Tripanosoma cruzi Infection.

[This corrects the article DOI: 10.1371/journal.pone.0153038.]

Enteric neuronal damage, intramuscular denervation and smooth muscle phenotype changes as mechanisms of chagasic megacolon : evidence from a long - term murine model of Tripanosoma cruzi infection.

We developed a novel murine model of long-term infection with Trypanosoma cruzi with the aim to elucidate the pathogenesis of megacolon and the associated adaptive and neuromuscular intestinal disorders. Our intent was to produce a chronic stage of the disease since the early treatment should avoid 100% mortality of untreated animals at acute phase. Treatment allowed animals to be kept infected and alive in order to develop the chronic phase of infection with low parasitism as in human disease. A group of Swiss mice was infected with the Y strain of T. cruzi. At the 11th day after infection, a sub-group was euthanized (acute-phase group) and another sub-group was treated with benznidazole and euthanized 15 months after infection (chronic-phase group). Whole colon samples were harvested and used for studying the histopathology of the intestinal smooth muscle and the plasticity of the enteric nerves. In the acute phase, all animals presented inflammatory lesions associated with intense and diffuse parasitism of the muscular and submucosa layers, which were enlarged when compared with the controls. The occurrence of intense degenerative inflammatory changes and increased reticular fibers suggests inflammatoryinduced necrosis of muscle cells. In the chronic phase, parasitism was insignificant; however, he architecture of Aüerbach plexuses was focally affected in the inflamed areas, and a significant decrease in the number of neurons and in the density of intramuscular nerve bundles was detected. Other changes observed included increased thickness of the colon wall, diffuse muscle cell hypertrophy, and increased collagen deposition, indicating early fibrosis in the damaged areas. Mast cell count significantly increased in the muscular layers. We propose a model for studying the long-term (15 months) pathogenesis of Chagasic megacolon in mice that mimics the human disease, which persists for several years and has not been fully elucidated. We hypothesize that the long-term inflammatory process mediates neuronal damage and intramuscular and intramural denervation, leading to phenotypic changes in smooth muscle cells associated with fibrosis. These long-term structural changes may represent the basic mechanism for the formation of the Chagasic megacolon

High-Fat diet increases HMGB1 expression and promotes lung inflammation in mice subjected to mechanical ventilation.

This study aims to evaluate the effects of a high-fat diet and mechanical ventilation on the pulmonary and systemic inflammatory response in C57BL/6 mice. Male C57BL/6 mice were divided into two groups: one received a standard diet, and the other received a high-fat diet. After 10 weeks, the groups were further divided into two groups each: control group (CG), mechanical ventilation group (MVG), diet group (DG), and diet mechanical ventilation group (DMVG). MVG and DMVG underwent mechanical ventilation for 60 minutes. All animals were euthanized for subsequent analysis. Animals receiving a high-fat diet presented higher body mass, adipose index, and greater adipocyte area. In the lung, the expression of HMGB1 was greater in DG and DMVG than in CG and MVG. CCL2 and IL-22 levels in MVG and DMVG were increased compared to those in CG and DG, whereas IL-10 and IL-17 were decreased. Superoxide dismutase activity was higher in MVG and DMVG than in CG. Catalase activity was lower in DG than in CG, and in MV groups, it was lower than that in CG and DG. MV and obesity promote inflammation and pulmonary oxidative stress in adult C57BL/6 mice

Exogenous surfactant prevents hyperoxia-induced lung injury in adult mice

Abstract Background In addition to the risk of developing ventilator-induced lung injury, patients with ARDS are at risk of developing hyperoxic injury due the supra-physiological oxygen supplementation clinically required to reverse hypoxemia. Alterations of endogenous surfactant system participate in the pulmonary dysfunction observed in ARDS. Administration of exogenous surfactant could have protective effects during hyperoxia. Methods Male BALB/c mice (8–10 weeks), a strain highly sensitive to hyperoxia, received the exogenous surfactant-containing protein SP-B and SP-C by intranasal instillation 12 h before starting 24 h of exposure to hyperoxia in an inhalation chamber and were compared to mice receiving hyperoxia alone and to controls subjected to normoxia. Results Compared to the hyperoxia group, the administration of exogenous surfactant was able to reduce lung inflammation through a reduction in the influx of neutrophils and inflammatory biomarkers such as TNF, IL-17, and HMGB1 expression. The antioxidant activity prevented oxidative damage by reducing lipid peroxidation and protein carbonylation and increasing superoxide dismutase activity when compared to the hyperoxia group. Conclusion Our results offer new perspectives on the effects and the mechanism of exogenous surfactant in protecting the airway and lungs, in oxygen-rich lung microenvironment, against oxidative damage and aggravation of acute inflammation induced by hyperoxia

Histopathological features and evidence of parasitism in the colons of Swiss mice infected with 50,000 trypomastigotes of the Y strain of <i>T</i>. <i>cruzi</i>.

<p>A, C, E, G, and H represent acute-phase aspects (11 d.a.i.). B, D, F, and I represent chronic-phase aspects (15 m.a.i.). (A) The colons of the control group of paired animals in the acute phase present normal cellularity and thickness. (B) In the chronic phase, no alterations were observed in the control group. (C) In acute-phase infected animals, a mononuclear inflammatory infiltrate was observed in the submucosal and muscular layers (arrow). In the myenteric plexus (arrowhead) and in the inner muscular layer, there is evidence of muscle fiber necrosis (thick arrow). This transmural pattern is merged with non-inflamed areas (not shown). (D) In contrast, in the chronic-phase infected group, mononuclear infiltrates are focal and more intense in the outer muscular layer in the periganglionar and perivascular areas (arrow). (E) Intense parasitism (inset,arrow) is associated with inflammatory infiltrates in the acute phase. (F) In the chronic phase, parasites are scarce and not associated with inflammatory foci (inset). (G) The acute-phase infected group showed strong iNOS positivity associated with inflammatory and degenerative changes in the myenteric plexus (arrowheads) compared (I) with the weak staining in inflammatory cells (arrow, and inset) in the chronic-phase infected group. A possible glial cell of ganglia (arrowhead) is also stained. (H) In addition, a greater amount of reticular fibers with thickened areas around the ganglia (arrow) were present in the acute phase. The results represent two independent experiments. A, B, C, D, I, H Magnification at 20x. Scale bar corresponds to 20 μm. E, F Magnification at 4x. Scale Bar corresponds to 100 μm. G Magnification at 40x. Scale bar corresponds to 10 μm. HE staining in A, B, C and D. Immunohistochemistry with anti-<i>T</i>. <i>cruzi</i> antibody in E, and F, and insets of E and F. Immunohistochemistry with anti-iNOS antibody in G and I. Silver Gomori staining in H.</p