ABSTRACT The aim of this review is to show the significant role of HIF-1alpha in inflammatory and infectious diseases. Hypoxia is a physiological characteristic of a wide range of diseases from cancer to infection. Cellular hypoxia is sensed by oxygen-sensitive hydrolase enzymes, which control the protein stability of hypoxia-inducible factor alpha 1 (HIF-1alpha) transcription factors. When stabilized, HIF-1alpha binds with its cofactors to HIF-responsive elements (HREs) in the promoters of target genes to organize a broad ranging transcriptional program in response to the hypoxic environment. HIF-1alpha also plays a regulatory function in response to a diversity of molecular signals of infection and inflammation even under normoxic conditions. HIF-1alpha is stimulated by pro-inflammatory cytokines, growth factors and a wide range of infections. Its induction is a general element of the host response to infection. In this review, we also discuss recent advances in knowledge on HIF-1alpha and inflammatory responses, as well as its direct influence in infectious diseases caused by bacteria, virus, protozoan parasites and fungi.

RESUMO A inflamação causada por Shigella flexneri pode causar danos à mucosa do cólon e morte celular por necrose e apoptose. Esta bactéria pode atingir a corrente sanguínea por esta via e o fígado através da veia porta. A hipóxia é uma condição presente em muitas doenças humanas, podendo induzir a translocação bacteriana a partir do lúmen intestinal. Nós estudamos a capacidade de S. flexneri invadir hepatócitos de rato e células Caco-2 nos microambientes de normóxia e hipóxia, bem como as alterações morfológicas e fisiológicas dessas células após a infecção sob hipóxia. Utilizamos a cultura primária de hepatócitos de ratos como modelo de estudo. Nós analisamos os seguintes parâmetros em condições de normóxia e hipóxia: morfologia, viabilidade celular, recuperação bacteriana e liberação de lactato desidrogenase (LDH). Os resultados mostraram menor quantidade de bactérias dentro das células Caco-2 do que em hepatócitos em condições de normóxia e hipóxia. Nós observamos que quanto maior foi a multiplicidade de infecção (MOI), maior também foi a recuperação bacteriana em hepatócitos. A condição hipóxica foi capaz de diminuir a recuperação de bactérias dos hepatócitos. A citotoxicidade avaliada pela liberação de LDH foi significativamente maior em células submetidas à hipóxia do que normóxia. As células Caco-2 em normóxia produziram 63% mais LDH do que os hepatócitos. O LDH aumentou 164% quando os hepatócitos foram submetidos à hipoxia e apenas 21% quando as células Caco-2 estavam na mesma condição. A apoptose avaliada por TUNEL foi significativamente maior em células submetidas à hipóxia que normóxia. Quando comparamos células hipóxicas houve mais apoptose entre hepatócitos do que nas células Caco-2. Concluindo, nossos resultados contribuem para um melhor conhecimento das interações entre as células estudadas e S. flexneri. Estes dados podem ser úteis no futuro, para definir estratégias de combate a este patógeno virulento.

SUMMARY Inflammation due to Shigella flexneri can cause damage to the colonic mucosa and cell death by necrosis and apoptosis. This bacteria can reach the bloodstream in this way, and the liver through portal veins. Hypoxia is a condition present in many human diseases, and it may induce bacterial translocation from intestinal lumen. We studied the ability of S. flexneri to invade rat hepatocytes and Caco-2 cells both in normoxic and hypoxic microenvironments, as well as morphological and physiological alterations in these cells after infection under hypoxia. We used the primary culture of rat hepatocytes as a model of study. We analyzed the following parameters in normoxic and hypoxic conditions: morphology, cell viability, bacterial recovery and lactate dehydrogenase (LDH) released. The results showed that there were fewer bacteria within the Caco-2 cells than in hepatocytes in normoxic and hypoxic conditions. We observed that the higher the multiplicity of infection (MOI) the greater the bacterial recovery in hepatocytes. The hypoxic condition decreased the bacterial recovery in hepatocytes. The cytotoxicity evaluated by LDH released by cells was significantly higher in cells submitted to hypoxia than normoxia. Caco-2 cells in normoxia released 63% more LDH than hepatocytes. LDH increased 164% when hepatocytes were submitted to hypoxia and just 21% when Caco-2 cells were in the same condition. The apoptosis evaluated by Tunel was significantly higher in cells submitted to hypoxia than normoxia. When comparing hypoxic cells, we obtained more apoptotic hepatocytes than apoptotic Caco-2 cells. Concluding our results contribute to a better knowledge of interactions between studied cells and Shigella flexneri. These data may be useful in the future to define strategies to combat this virulent pathogen.

A invasão de hepatócitos por Listeria monocytogenes (LM) e Salmonella Typhimurium (ST) pode estimular a liberação do Fator de Necrose Tumoral (TNF-α) e induzir a apoptose celular. Neste estudo comparamos o comportamento de hepatócitos invadidos por três sorotipos de L. monocytogenes (LM-4a, LM-4b e LM-1/2a) e por ST para entender qual bacteria é mais efetiva no processo infeccioso. Nós quantificamos a liberação de TNF-α pelos hepatócitos por ELISA e as taxas de apoptose pelas técnicas de anexina V (apoptose precoce) e TUNEL (apoptose tardia). A morfologia das células foi estudada também. A taxa de liberação de TNF-α foi mais alta em hepatócitos invadidos por ST. ST e LM-1/2a induziram as maiores taxas de apoptose pelo método TUNEL, enquanto LM-4b produziu as maiores taxas de apoptose por anexina V. Os hepatócitos invadidos apresentaram várias alterações morfológicas. Na análise do conjunto de dados, os sorotipos LM-4b e LM-1/2a provaram ser os mais eficientes na invasão celular, enquanto que ST adaptou-se mais rápido ao meio e induziu a liberação precoce de TNF-α pelos hepatócitos.

Invasion of hepatocytes by Listeria monocytogenes (LM) and Salmonella Typhimurium (ST) can stimulate tumor necrosis factor alpha (TNF-α) release and induce apoptosis. In this study, we compared the behavior of hepatocytes invaded by three L. monocytogenes serotypes (LM-4a, LM-4b and LM-1/2a) and by ST to understand which bacterium is more effective in the infectious process. We quantified TNF-α release by ELISA, apoptosis rates by annexin V (early apoptosis) and TUNEL (late apoptosis) techniques. The cell morphology was studied too. TNF-α release rate was highest in ST-invaded hepatocytes. ST and LM-1/2a induced the highest apoptosis production rates evaluated by TUNEL. LM-4b produced the highest apoptosis rate measured by annexin. Invaded hepatocytes presented various morphological alterations. Overall, LM-4b and LM-1/2a proved to be the most efficient at cell invasion, although ST adapted faster to the environment and induced earlier hepatocyte TNF-α release.

A ascite é o acúmulo de líquido livre de origem patológica na cavidade abdominal, fenômeno presente em várias doenças da prática clínica. A doença mais associada com ascite é a cirrose hepática. Na sua fisiopatologia destacam-se três teorias que ocorrem sempre em determinado paciente, porém em momentos diferentes de sua doença: vasodilatação, "overflow" e "underfill". O conceito mais moderno sugere que as três teorias estão presentes no mesmo paciente com cirrose, dependendo do tempo de evolução de sua doença. A teoria da vasodilatação estaria presente desde a fase pré-ascítica até a ascite de longa data. A teoria do overflow seria predominante nos primeiros meses de ascite e a teoria underfill explicaria a maioria dos achados em pacientes com ascite por longo tempo. Neste artigo são comentadas em detalhes as várias doenças que produzem ascite, os métodos diagnósticos empregados na pesquisa clínica da ascite, as complicações da ascite e as opções terapêuticas disponíveis. Em cada item é mostrado o grau de evidência (A até C) presente na literatura médica.

The accumulation of free fluid of pathological origin in the peritoneal cavity is named ascites, and, in clinical practice this phenomenon is present in several diseases. The most common cause of ascites is liver cirrhosis. In the pathophysiology of ascites three theories are noteworthy: vasodilation, overflow and underfill. The modern concept suggest that these three theories are present in the same patient with cirrhosis depending on the disease evolution time. The vasodilation theory would be important in the pre-ascitic phase as well as during all the ascites evolution time. The overflow theory would be important in the first months of development of ascites in cirrhosis, and the underfill theory would explain most of the findings in patients with ascites for a long time. This article comments in detailed, several diseases that produce ascites, the diagnostic methods employed in clinical investigation of ascites' complication and therapeutics options available. In each item the evidence grade (A to C) found in medical literature is shown.

This study verified the correlation between the serum levels of TNF alpha and different clinical forms of tuberculosis. We described a group of 24 patients presenting several clinical forms of tuberculosis and a control group of 13 healthy individuals. The levels of TNF alpha were measured by bioassay method. The levels of TNF-alpha had significant differences between the tuberculosis and control groups. The patients with abnormal chest X-Ray findings had higher TNF alpha levels (15328.48 ± 4602.19 pg/mL) when compared to patients with normal X-Rays (3353.18 ± 1495.29 pg/mL) (p<0.05). Patients that lost weight had higher TNF alpha levels (15468.54 ± 4580.54 pg/mL) than those that didn't loose weight (2904.98 ± 1367.89) (p<0.05). The levels of TNF alpha were higher in patients with a positive PPD skin test than in those with a negative PPD test (p<0.05). There was a positive correlation between patients' clinical severity and the serum levels of TNF alpha. In patients with successive measurements of TNF alpha, we observed that there was a drop in cytokine levels, and also a clinical improvement concomitantly. We concluded that there was a correlation between serum TNF alpha levels and chest X-Ray alterations, loss of weight, positive PPD skin test and clinical severity in patients with tuberculosis. There was evidence of a worse clinical outcome in patients with tuberculosis that presented higher TNF alpha serum levels.

The type of immune response induced by tuberculosis (Th1 or Th2) and its correlation with the clinical outcome is unclear. We studied 13 patients with active tuberculosis (TBC). The peripheral blood mononuclear cells producing IFN-gamma (PBMC-IG) were measured by enzyme-linked immunospot (ELISPOT) technique. The control group had ten healthy individuals vaccinated against tuberculosis. We collected blood samples of each patient in two moments: a) in the hospital admission without treatment (TBC1); b) after seven to 20 days of treatment (TBC2). The comparison of the spots forming units of PBMC-IG between TBC group and controls showed that there was a significant difference between TBC1 and control group (p < 0.001) and between TBC2 and control group (p < 0.005), but there was no difference between TBC1 and TBC2 (p > 0.05). A positive correlation was found between PBMC-IG and hemoglobin value, as well as between PBMC-IG and weight loss. There was no correlation between PBMC-IG and other variables [age, erythrocyte sedimentation rate (ESR), C-reactive protein (CRP)]. We conclude that tuberculosis activates Th1 immune response due to increase of PBMC producing IFN-gamma. There was no difference between the first sample (TBC1) and the second sample (TBC2) of PBMC-IG. This result can have occurred due to treatment influence, or can indicate that the immune response reachs a plateau. The positive correlation among PBMC-IG and both hemoglobin level and weight loss indicates that may exist a link between patient's clinical status and the immune response intensity.

Listeria monocytogenes, agente etiológico de infecção grave de origem alimentar, utiliza mecanismos sofisticados de entrada no citoplasma do hospedeiro e manipulação do citoesqueleto, resultando em morte celular. As interações entre células do hospedeiro e bactérias podem resultar em produção de citocinas como o Fator de Necrose Tumoral alfa (TNF-alfa). Hepatócitos têm potencial de produzir citocinas pro-inflamatórias como TNF-alfa, quando invadidos por bactérias. No presente trabalho demonstramos o comportamento dos hepatócitos invadidos por L. monocytogenes pela análise microscópica, determinação da produção de TNF-alfa por bioensaio e análise da apoptose pela técnica TUNEL. A presença da bactéria, na razão que variou de 5 a 50.000 bactérias por célula, induziu ruptura das monocamadas celulares. Observamos presença de bactérias internalizadas na 1ª hora de incubação por microscopia eletrônica. Os níveis de TNF-alfa aumentaram da 1ª hora de incubação até a 6ª hora, variando de 0 a 3749 pg/mL. Nas 7ª e 8ª horas de incubação, ocorreram quedas não significativas dos níveis de TNF-alfa, indicando possível saturação dos receptores celulares. A quantidade de TNF-alfa produzido por hepatócitos foi dependente do tempo de incubação, assim como da proporção entre bactérias e células. A taxa de apoptose aumentou diretamente com o tempo de incubação (1 h a 8 + 24 h), variando de 0 a 43%, assim como com a razão bactérias : células. Estes resultados mostram a habilidade de L. monocytogenes não-hemolítica em invadir os hepatócitos, e as principais conseqüências deste fenômeno são: liberação de TNF-alfa e indução de apoptose. Assim, podemos especular que hepatócitos usam apoptose induzida por TNF-alfa para liberar bactérias de seu interior, facilitando a destruição destas pelo sistema imune.

Listeria monocytogenes, etiological agent of severe human foodborne infection, uses sophisticated mechanisms of entry into host cytoplasm and manipulation of the cellular cytoskeleton, resulting in cell death. The host cells and bacteria interaction may result in cytokine production as Tumor Necrosis Factor (TNF) alpha. Hepatocytes have potential to produce pro-inflammatory cytokines as TNF-alpha when invaded by bacteria. In the present work we showed the behavior of hepatocytes invaded by L. monocytogenes by microscopic analysis, determination of TNF-alpha production by bioassay and analysis of the apoptosis through TUNEL technique. The presence of bacterium, in ratios that ranged from 5 to 50,000 bacteria per cell, induced the rupture of cellular monolayers. We observed the presence of internalized bacteria in the first hour of incubation by electronic microscopy. The levels of TNF-alpha increased from first hour of incubation to sixth hour, ranging from 0 to 3749 pg/mL. After seven and eight hours of incubation non-significant TNF-alpha levels decrease occurred, indicating possible saturation of cellular receptors. Thus, the quantity of TNF-alpha produced by hepatocytes was dependent of the incubation time, as well as of the proportion between bacteria and cells. The apoptosis rate increased in direct form with the incubation time (1 h to 8 + 24 h), ranging from 0 to 43%, as well as with the bacteria : cells ratio. These results show the ability of hepatocyte invasion by non-hemolytic L. monocytogenes, and the main consequences of this phenomenon were the release of TNF-alpha by hepatocytes and the induction of apoptosis. We speculate that hepatocytes use apoptosis induced by TNF-alpha for release bacteria to extracellular medium. This phenomenon may facilitate the bacteria destruction by the immune system.

Os radicais livres de oxigênio são moléculas que apresentam elétrons não pareados em sua órbita externa, capazes de transformar outras moléculas com as quais se encontram, como proteínas, carbohidratos, lípides e o ácido desoxirribonucleico. Essas moléculas são geradas em situações clínicas onde microambientes de hipóxia são seguidos por microambientes de reoxigenação. Nesse grupo estão o choque hemodinâmico, a septicemia, a resposta inflamatória sistêmica, as hepatites fulminantes, o transplante de órgãos, e a insuficiência respiratória, entre outras condições. Neste trabalho discutimos os principais conceitos sobre os radicais livres de oxigênio: os principais tipos, sua formação e a forma como atuam sobre todas as estruturas celulares provocando lesão tecidual significativa. Os principais sistemas de defesa antioxidante existentes para combater o estresse oxidativo são comentados, com destaque para a glutationa, superóxido dismutase, catalase, glutationa peroxidase e N-acetilcisteína. A influência dos radicais livres de oxigênio sobre as principais doenças pulmonares também é discutida, com ênfase nos produtos do cigarro, doença pulmonar obstrutiva crônica, asma, apnéia obstrutiva do sono e síndrome do desconforto respiratório agudo.

Oxygen free radicals are molecules that present unpaired electrons in their outer orbit and can transform other molecules such as proteins, carbohydrates, lipids and deoxyribonucleic acid. Oxygen free radicals are produced in various clinical conditions in which hypoxic microenvironments are generated and reoxygenation follows. Such situations include clinical shock, septicemia, systemic inflammatory response, fulminant hepatitis, organ transplant and respiratory failure. In this review, we discuss the main concepts related to oxygen free radicals: the principal types and their formation, as well as the way in which they affect cellular structures and cause significant tissue damage. We present also the main antioxidants that guard against oxidative stress, including glutathione, glutathione peroxidase, superoxide dismutase, catalase, and N-acetylcysteine. The influence of oxygen free radicals on the principal pulmonary diseases are also discussed, with special emphasis given to oxygen free radicals in cigarette smoke, chronic obstructive pulmonary disease, asthma, sleep apnea syndrome and acute respiratory distress syndrome.

Os mecanismos que determinam o clearance ou a persistência da infecção viral nas hepatites virais crônicas não estão ainda bem identificados. O progresso no conhecimento sobre as ferramentas genéticas moleculares tem permitido detectar variações na resposta imune, que freqüentemente são associadas com polimorfismos do genoma humano. As diferenças na susceptibilidade do hospedeiro para as doenças infecciosas e a intensidade das doenças não podem ser atribuídas apenas à virulência do agente microbiano. Neste artigo são discutidos vários avanços recentes no conhecimento sobre a influência dos genes humanos nas hepatites crônicas B e C, a saber: a) As associações entre os polimorfismos HLA e a susceptibilidade ou resistência às doenças hepáticas virais; b) Alelos protetores influenciando as hepatites virais B (HVB) e C (HVC); c) Alelos prejudiciais influenciando HVB e HVC; d) Genes candidatos associados com a evolução clínica de HVB e HVC (genes que influenciam as células estreladas do fígado, a produção de TGF-beta1 e TNF-alfa, os depósitos de ferro hepáticos, a produção de angiotensina II, entre outros). O conhecimento das associações genéticas com as hepatites virais crônicas pode fornecer indícios para o pleno entendimento de como se desenvolvem as suas complicações terminais, como a cirrose e o carcinoma hepatocelular. Em futuro próximo, a análise do genoma humano será capaz de elucidar o curso natural de uma hepatite viral, bem como a sua resposta à terapêutica.

The mechanisms that determine viral clearance or viral persistence in chronic viral hepatitis have yet to be identified. Recent advances in molecular genetics have permitted the detection of variations in immune response, often associated with polymorphism in the human genome. Differences in host susceptibility to infectious disease and disease severity cannot be attributed solely to the virulence of microbial agents. Several recent advances concerning the influence of human genes in chronic viral hepatitis B and C are discussed in this article: a) the associations between human leukocyte antigen polymorphism and viral hepatic disease susceptibility or resistance; b) protective alleles influencing hepatitis B virus (HBV) and hepatitis C virus (HCV) evolution; c) prejudicial alleles influencing HBV and HCV; d) candidate genes associated with HBV and HCV evolution; d) other genetic factors that may contribute to chronic hepatitis C evolution (genes influencing hepatic stellate cells, TGF-beta1 and TNF-alpha production, hepatic iron deposits and angiotensin II production, among others). Recent discoveries regarding genetic associations with chronic viral hepatitis may provide clues to understanding the development of end-stage complications such as cirrhosis or hepatocellular carcinoma. In the near future, analysis of the human genome will allow the elucidation of both the natural course of viral hepatitis and its response to therapy.

O conhecimento sobre a patogênese da febre tifóide vem crescendo nos últimos anos, principalmente sobre os fenômenos celulares e moleculares, responsáveis pelas manifestações clínicas desta doença. Neste artigo são discutidas várias descobertas recentes, a saber: a) O sistema bacteriano de secreção protéica tipo III; b) Os cinco genes de virulência da Salmonella spp. que codificam os Sips (Salmonella invasion protein) A, B, C, D e E, capazes de induzir apoptosis em macrófagos; c) A função dos receptores Toll R2 e Toll R4 presentes na superfície de macrófagos (descobertos na Drosophila). Os receptores da família Toll são críticos na sinalização mediada por LPS nos macrófagos, em associação com LBP e CD14; d) As linhas de defesa imune entre o lúmen intestinal e os órgãos internos; e) O papel fundamental das células endoteliais no desvio inflamatório, da corrente sanguínea para os tecidos infectados pela bactéria. Além destes aspectos, os autores comentam a correlação entre as características clínicas da febre tifóide e os fenômenos celulares e moleculares da doença, bem como as conseqüências terapêuticas deste conhecimento.

The knowledge about typhoid fever pathogenesis is growing in the last years, mainly about the cellular and molecular phenomena that are responsible by clinical manifestations of this disease. In this article are discussed several recent discoveries, as follows: a) Bacterial type III protein secretion system; b) The five virulence genes of Salmonella spp. that encoding Sips (Salmonella invasion protein) A, B, C, D and E, which are capable of induce apoptosis in macrophages; c) The function of Toll R2 and Toll R4 receptors present in the macrophage surface (discovered in the Drosophila). The Toll family receptors are critical in the signalizing mediated by LPS in macrophages in association with LBP and CD14; d) The lines of immune defense between intestinal lumen and internal organs; e) The fundamental role of the endothelial cells in the inflammatory deviation from bloodstream into infected tissues by bacteria. In addition to above subjects, the authors comment the correlation between the clinical features of typhoid fever and the cellular and molecular phenomena of this disease, as well as the therapeutic consequences of this knowledge.

Descrito caso de mulher de 25 anos, negra, procedente da cidade de São Paulo - Brasil, que desenvolveu quadro agudo de colangite obstrutiva por Ascaris lumbricoides, com septicemia e múltiplos abscessos hepáticos. Era portadora de traço falciforme e tinha tido um parto normal há 3 meses. Foi estabelecido o diagnóstico de infestação maciça por Ascaris lumbricoides em vias biliares com ultrasonografia abdominal e colangiografia retrógrada endoscópica. Sessenta vermes foram retirados do colédoco e drenados os abscessos hepáticos por cirurgia. O quadro infeccioso era polimicrobiano. Houve recuperação total da paciente após longa evolução, com antibioticoterapia de amplo espectro e novas cirurgias com extração de vermes residuais. Os autores apresentam os métodos diagnósticos empregados, o quadro clínico-bioquímico e a abordagem terapêutica. A fisiopatologia da ascaridíase biliar é discutida.

This is a report of a 25 years old black woman from the city of São Paulo, Brazil, who developed acute obstructive cholangitis of Ascaris lumbricoides with septicemia and multiple hepatic abscesses. The patient had sickle cell trait and normal delivery 3 months ago. Massive infestation of the biliary tract by Ascaris lumbricoides was diagnosed by abdominal ultrasonography and endoscopic retrograde cholangiography. Sixty worms were removed from the common bile duct and hepatic abscesses were drained by surgery. The infectious process was polymicrobial. The patient's recovery was complete after a long evolution with a wide spectrum antibiotic therapy. New surgeries were needed to remove residual worms in the biliary tract. The diagnostic methods, clinical-biochemical features and also the clinical and surgical management are presented. The biliary ascariasis pathophysiology is commented.