RESUMEN
Liver fibrosis occurring as an outcome of non-alcoholic steatohepatitis (NASH) can precede the development of cirrhosis. We investigated the effects of sorafenib in preventing liver fibrosis in a rodent model of NASH. Adult Sprague-Dawley rats were fed a choline-deficient high-fat diet and exposed to diethylnitrosamine for 6 weeks. The NASH group (n=10) received vehicle and the sorafenib group (n=10) received 2.5 mg·kg-1·day-1 by gavage. A control group (n=4) received only standard diet and vehicle. Following treatment, animals were sacrificed and liver tissue was collected for histologic examination, mRNA isolation, and analysis of mitochondrial function. Genes related to fibrosis (MMP9, TIMP1, TIMP2), oxidative stress (HSP60, HSP90, GST), and mitochondrial biogenesis (PGC1α) were evaluated by real-time quantitative polymerase chain reaction (RT-qPCR). Liver mitochondrial oxidation activity was measured by a polarographic method, and cytokines by enzyme-linked immunosorbent assay (ELISA). Sorafenib treatment restored mitochondrial function and reduced collagen deposition by nearly 63% compared to the NASH group. Sorafenib upregulated PGC1α and MMP9 and reduced TIMP1 and TIMP2 mRNA and IL-6 and IL-10 protein expression. There were no differences in HSP60, HSP90 and GST expression. Sorafenib modulated PGC1α expression, improved mitochondrial respiration and prevented collagen deposition. It may, therefore, be useful in the treatment of liver fibrosis in NASH.
Asunto(s)
Adolescente , Adulto , Femenino , Humanos , Masculino , Adulto Joven , Trastorno Depresivo Mayor/terapia , Costos de la Atención en Salud/estadística & datos numéricos , Accesibilidad a los Servicios de Salud/estadística & datos numéricos , Trastornos Relacionados con Sustancias/rehabilitación , Diagnóstico Dual (Psiquiatría) , Trastorno Depresivo Mayor/complicaciones , Trastorno Depresivo Mayor/economía , Encuestas Epidemiológicas , Accesibilidad a los Servicios de Salud/economía , Servicios de Salud Mental/economía , Servicios de Salud Mental/estadística & datos numéricos , Trastornos Relacionados con Sustancias/complicaciones , Trastornos Relacionados con Sustancias/economía , Estados UnidosRESUMEN
Liver fibrosis occurring as an outcome of non-alcoholic steatohepatitis (NASH) can precede the development of cirrhosis. We investigated the effects of sorafenib in preventing liver fibrosis in a rodent model of NASH. Adult Sprague-Dawley rats were fed a choline-deficient high-fat diet and exposed to diethylnitrosamine for 6 weeks. The NASH group (n=10) received vehicle and the sorafenib group (n=10) received 2.5 mg·kg(-1)·day(-1) by gavage. A control group (n=4) received only standard diet and vehicle. Following treatment, animals were sacrificed and liver tissue was collected for histologic examination, mRNA isolation, and analysis of mitochondrial function. Genes related to fibrosis (MMP9, TIMP1, TIMP2), oxidative stress (HSP60, HSP90, GST), and mitochondrial biogenesis (PGC1α) were evaluated by real-time quantitative polymerase chain reaction (RT-qPCR). Liver mitochondrial oxidation activity was measured by a polarographic method, and cytokines by enzyme-linked immunosorbent assay (ELISA). Sorafenib treatment restored mitochondrial function and reduced collagen deposition by nearly 63% compared to the NASH group. Sorafenib upregulated PGC1α and MMP9 and reduced TIMP1 and TIMP2 mRNA and IL-6 and IL-10 protein expression. There were no differences in HSP60, HSP90 and GST expression. Sorafenib modulated PGC1α expression, improved mitochondrial respiration and prevented collagen deposition. It may, therefore, be useful in the treatment of liver fibrosis in NASH.
Asunto(s)
Cirrosis Hepática/tratamiento farmacológico , Mitocondrias Hepáticas/efectos de los fármacos , Niacinamida/análogos & derivados , Enfermedad del Hígado Graso no Alcohólico/complicaciones , Compuestos de Fenilurea/uso terapéutico , Inhibidores de Proteínas Quinasas/uso terapéutico , Animales , Chaperonina 60/análisis , Chaperonina 60/genética , Dieta Alta en Grasa/métodos , Dietilnitrosamina , Modelos Animales de Enfermedad , Colágenos Fibrilares/efectos de los fármacos , Glutatión Transferasa/análisis , Glutatión Transferasa/genética , Proteínas HSP90 de Choque Térmico/análisis , Proteínas HSP90 de Choque Térmico/genética , Interleucina-10/análisis , Interleucina-10/genética , Interleucina-6/análisis , Interleucina-6/genética , Cirrosis Hepática/etiología , Cirrosis Hepática/patología , Metaloproteinasa 9 de la Matriz/análisis , Metaloproteinasa 9 de la Matriz/genética , Mitocondrias Hepáticas/metabolismo , Niacinamida/uso terapéutico , Enfermedad del Hígado Graso no Alcohólico/inducido químicamente , Coactivador 1-alfa del Receptor Activado por Proliferadores de Peroxisomas gamma , Polarografía , ARN Mensajero/aislamiento & purificación , Ratas Sprague-Dawley , Sorafenib , Inhibidor Tisular de Metaloproteinasa-1/análisis , Inhibidor Tisular de Metaloproteinasa-1/genética , Inhibidor Tisular de Metaloproteinasa-2/análisis , Inhibidor Tisular de Metaloproteinasa-2/genética , Factores de Transcripción/análisis , Factores de Transcripción/genéticaRESUMEN
In the North of Portugal, a mass vaccination programme of small ruminants was conducted from 2001 to 2004. A study of cost-benefit was carried out for the 2000/2005 period to ascertain the economic benefits of this strategy. In order to estimate the cost of the zoonosis, the compensation costs paid to farmers for culled animals in the Brucellosis Eradication Campaign, data from vaccine Rev. 1 costs, and costs of people internment due to brucellosis were studied. An increase in the cost was observed from 2000 to 2001 (aproximately US$ 110,000), essentially due to compensation for culled animals, but also from vaccination and human internment costs. However, a progressive decrease was observed from 2001 to 2004 (from more than US$1,200,000 in 2001 to US$180,000 in 2004), roughly US$ 1,020,000 less. As the main conclusion, the annual cost of this zoonosis decreased by an average of almost US$600,000 per year, and a total of more than three million dollars were saved from 2000 to 2005. The results of this study suggest that mass vaccination with Rev. 1 reduced overall costs and was effective in reducing animal and human brucellosis costs.
No norte de Portugal, procedeu-se à vacinação massiva contra a brucelose em pequenos ruminantes entre 2001 e 2004. Neste estudo, efetuou-se uma análise de custo benefício entre 2000 e 2005 para apurar os benefícios econômicos dessa estratégia. Para estimar o custo da zoonose, estudaram-se três variáveis: a indenização paga aos proprietários dos animais eliminados durante a campanha de erradicação da brucelose, o custo da vacina Rev. 1 e os custos de internamento das pessoas infectadas por brucelose nas unidades de Serviços de Saúde. Durante os anos em estudo, observou-se um aumento de custo de aproximadamente US$110.000, entre 2000 e 2001, principalmente, devido à indenização dos animais eliminados e à aplicação da vacina e os custos por internamento de pessoas infectadas, seguindo-se uma redução acentuada entre 2001 e 2004 (de US$1.200.000 para US$180.000 em 2004), o que perfez US$1.020.000 poupados. O custo anual da zoonose, em relação às variáveis estudadas, diminuiu em média US$600.000 por ano, sendo poupados mais de US$ 3.000.000 entre 2000 e 2005. Estes resultados sugerem que a vacinação com Rev. 1 reduziu os custos totais da infecção humana e animal relativamente às variáveis estudadas.
Asunto(s)
Animales , Brucelosis/patología , Vacunación , Salud Pública/tendencias , Zoonosis/transmisiónRESUMEN
In the North of Portugal, a mass vaccination programme of small ruminants was conducted from 2001 to 2004. A study of cost-benefit was carried out for the 2000/2005 period to ascertain the economic benefits of this strategy. In order to estimate the cost of the zoonosis, the compensation costs paid to farmers for culled animals in the Brucellosis Eradication Campaign, data from vaccine Rev. 1 costs, and costs of people internment due to brucellosis were studied. An increase in the cost was observed from 2000 to 2001 (aproximately US$ 110,000), essentially due to compensation for culled animals, but also from vaccination and human internment costs. However, a progressive decrease was observed from 2001 to 2004 (from more than US$1,200,000 in 2001 to US$180,000 in 2004), roughly US$ 1,020,000 less. As the main conclusion, the annual cost of this zoonosis decreased by an average of almost US$600,000 per year, and a total of more than three million dollars were saved from 2000 to 2005. The results of this study suggest that mass vaccination with Rev. 1 reduced overall costs and was effective in reducing animal and human brucellosis costs.(AU)
No norte de Portugal, procedeu-se à vacinação massiva contra a brucelose em pequenos ruminantes entre 2001 e 2004. Neste estudo, efetuou-se uma análise de custo benefício entre 2000 e 2005 para apurar os benefícios econômicos dessa estratégia. Para estimar o custo da zoonose, estudaram-se três variáveis: a indenização paga aos proprietários dos animais eliminados durante a campanha de erradicação da brucelose, o custo da vacina Rev. 1 e os custos de internamento das pessoas infectadas por brucelose nas unidades de Serviços de Saúde. Durante os anos em estudo, observou-se um aumento de custo de aproximadamente US$110.000, entre 2000 e 2001, principalmente, devido à indenização dos animais eliminados e à aplicação da vacina e os custos por internamento de pessoas infectadas, seguindo-se uma redução acentuada entre 2001 e 2004 (de US$1.200.000 para US$180.000 em 2004), o que perfez US$1.020.000 poupados. O custo anual da zoonose, em relação às variáveis estudadas, diminuiu em média US$600.000 por ano, sendo poupados mais de US$ 3.000.000 entre 2000 e 2005. Estes resultados sugerem que a vacinação com Rev. 1 reduziu os custos totais da infecção humana e animal relativamente às variáveis estudadas.(AU)
Asunto(s)
Animales , Vacunación , Brucelosis/patología , Salud Pública/tendencias , Zoonosis/transmisiónRESUMEN
AIMS: We performed the first characterization of the microbiota associated with the reef coral Mussismilia braziliensis by means of a culture-independent approach. METHODS AND RESULTS: The main groups were Proteobacteria, Cyanobacteria and unclassified bacteria according to the 16S rDNA libraries. Most of the sequences of the mucus of healthy and diseased M. braziliensis did not find close matches in GenBank (i.e. >97% 16S rDNA similarity). Most of the sequences of seawater and mucus of healthy coral fell into tight clusters (17 and 15 clusters respectively). In contrast, most of the sequences of mucus of diseased coral did not form clusters. The rarefaction curves indicate saturation in the recovery of higher taxa (approximately 40 phyla). However, the number of species in the coral mucus (n = 130-170) and seawater (n = 170) did not reach a plateau. CONCLUSIONS: The coral microbiota encompasses several potentially novel species and higher taxa. The microbiota of M. braziliensis appears to be species-specific. Diseased coral may have provided a suitable place for colonization by opportunistic bacteria, resulting in a greater bacterial diversity. SIGNIFICANCE AND IMPACT OF THE STUDY: The first study on the diversity of the microbiota of the endemic and endangered of extinction coral M. braziliensis.
Asunto(s)
Antozoos/microbiología , Bacterias/clasificación , Ecosistema , Agua de Mar/microbiología , Animales , Océano Atlántico , Bacterias/genética , Biodiversidad , Brasil , ADN Bacteriano/genética , Biblioteca de Genes , Datos de Secuencia Molecular , Proteobacteria/clasificación , Proteobacteria/genética , ARN Ribosómico 16S/genética , Análisis de Secuencia de ADNRESUMEN
Estudou-se a presença de bacilos álcool-ácido resistentes compatíveis com Mycobacterium avium subsp. paratuberculosis em esfregaços de fezes e tecidos de ovinos. Vinte e seis esfregaços de fezes e 104 de tecidos, pertencentes a 26 animais diagnosticados como paratuberculosos, foram analisados pelo método de Ziehl-Neelsen. Dezesseis (61,5 por cento) esfregaços fecais apresentaram bacilos álcool-ácido resistentes compatíveis no exame microscópico. Vinte animais (76,9 por cento) foram diagnosticados pelo método nos esfregaços de tecidos. Vinte e um animais apresentaram esfregaços positivos nas fezes e nos tecidos, simultaneamente. A sensibilidade de Ziehl-Neelsen para os esfregaços fecais, esfregaços de tecidos e para a combinação de ambos foi de 61,5 por cento, 76,9 por cento e 80,8 por cento, respectivamente.
The presence of acid-fast bacilli compatible with Mycobacterium avium subsp. paratuberculosis in fecal and tissues smears was investigated using the Ziehl-Neelsen staining. A total of 26 fecal smears and 104 tissues smears collected from 26 sheep with confirmed paratuberculosis were analyzed. Sixteen (61.5 percent) fecal smears showed compatible with acid-fast bacilli on microscopic examination after staining. Twenty animals (76.9 percent) were diagnosed based on the positivity of tissues smears. The Ziehl-Neelsen sensitivities to faecal smears, tissues smears, and a combination of both were 61.5 percent, 76.9 percent, and 80.8 percent, respectively.
Asunto(s)
Animales , Heces/microbiología , Mycobacterium avium subsp. paratuberculosis/aislamiento & purificación , OvinosRESUMEN
Estudou-se a presença de bacilos álcool-ácido resistentes compatíveis com Mycobacterium avium subsp. paratuberculosis em esfregaços de fezes e tecidos de ovinos. Vinte e seis esfregaços de fezes e 104 de tecidos, pertencentes a 26 animais diagnosticados como paratuberculosos, foram analisados pelo método de Ziehl-Neelsen. Dezesseis (61,5 por cento) esfregaços fecais apresentaram bacilos álcool-ácido resistentes compatíveis no exame microscópico. Vinte animais (76,9 por cento) foram diagnosticados pelo método nos esfregaços de tecidos. Vinte e um animais apresentaram esfregaços positivos nas fezes e nos tecidos, simultaneamente. A sensibilidade de Ziehl-Neelsen para os esfregaços fecais, esfregaços de tecidos e para a combinação de ambos foi de 61,5 por cento, 76,9 por cento e 80,8 por cento, respectivamente.(AU)
The presence of acid-fast bacilli compatible with Mycobacterium avium subsp. paratuberculosis in fecal and tissues smears was investigated using the Ziehl-Neelsen staining. A total of 26 fecal smears and 104 tissues smears collected from 26 sheep with confirmed paratuberculosis were analyzed. Sixteen (61.5 percent) fecal smears showed compatible with acid-fast bacilli on microscopic examination after staining. Twenty animals (76.9 percent) were diagnosed based on the positivity of tissues smears. The Ziehl-Neelsen sensitivities to faecal smears, tissues smears, and a combination of both were 61.5 percent, 76.9 percent, and 80.8 percent, respectively.(AU)
Asunto(s)
Animales , Mycobacterium avium subsp. paratuberculosis/aislamiento & purificación , Heces/microbiología , OvinosRESUMEN
Despite the efficiency of fludarabine in the induction of clinical responses in B-cell chronic lymphocytic leukemia (B-CLL) patients, resistance to this drug has been documented. The present study tested whether resistance to fludarabine is related to the expression of inhibitor of apoptosis proteins (IAPs) family members. We analyzed the expression of c-IAP1, c-IAP2 and XIAP, by immunocytochemistry, in 30 blood samples from B-CLL patients and correlated protein expression to fludarabine-induced apoptosis estimated by an annexin-V assay. Expression of c-IAP1, c-IAP2 and XIAP were found predominantly in the cytoplasm, and a wide range of staining intensities was observed among distinct samples. No correlation was found between the levels of IAPs expression and prognostic factors such as age, gender, lymphocyte doubling time, white blood cell count or previous treatment. The expression of IAPs also failed to predict the sensitivity to fludarabine-induced apoptosis. Alternative pathways of cell death may explain the independence of fludarabine-induced apoptosis from the high expression of IAPs.
Asunto(s)
Antineoplásicos/farmacología , Apoptosis/efectos de los fármacos , Proteínas Inhibidoras de la Apoptosis/metabolismo , Leucemia Linfocítica Crónica de Células B/tratamiento farmacológico , Vidarabina/análogos & derivados , Adulto , Anciano , Anexina A5/metabolismo , Células Cultivadas , Pruebas Inmunológicas de Citotoxicidad , Femenino , Humanos , Inmunohistoquímica , Proteínas Inhibidoras de la Apoptosis/genética , Leucemia Linfocítica Crónica de Células B/patología , Masculino , Persona de Mediana Edad , Vidarabina/farmacologíaRESUMEN
Oxidative stress and hepatic mitochondria play a role in the pathogenesis of nonalcoholic fatty liver disease. The aim of the present study was to evaluate the role of hepatic mitochondrial dysfunction and oxidative stress in the pathogenesis of the disease. Fatty liver was induced in Wistar rats with a choline-deficient diet (CD; N = 7) or a high-fat diet enriched with PUFAs-omega-3 (H; N = 7) for 4 weeks. The control group (N = 7) was fed a standard diet. Liver mitochondrial oxidation and phosphorylation were measured polarographically and oxidative stress was estimated on the basis of malondialdehyde and glutathione concentrations. Moderate macrovacuolar liver steatosis was observed in the CD group and mild liver steatosis was observed in the periportal area in the H group. There was an increase in the oxygen consumption rate by liver mitochondria in respiratory state 4 (S4) and a decrease in respiratory control rate (RCR) in the CD group (S4: 32.70 +/- 3.35; RCR: 2.55 +/- 0.15 ng atoms of O2 min-1 mg protein-1) when compared to the H and control groups (S4: 23.09 +/- 1.53, 17.04 +/- 2.03, RCR: 3.15 +/- 0.15, 3.68 +/- 0.15 ng atoms of O2 min-1 mg protein-1, respectively), P < 0.05. Hepatic lipoperoxide concentrations were significantly increased and the concentration of reduced glutathione was significantly reduced in the CD group. A choline-deficient diet causes moderate steatosis with disruption of liver mitochondrial function and increased oxidative stress. These data suggest that lipid peroxidation products can impair the flow of electrons along the respiratory chain, causing overreduction of respiratory chain components and enhanced mitochondrial reactive oxygen species. These findings are important in the pathogenesis of nonalcoholic fatty liver disease.
Asunto(s)
Hígado Graso/etiología , Mitocondrias Hepáticas/fisiología , Enfermedades Mitocondriales/complicaciones , Estrés Oxidativo/fisiología , Animales , Deficiencia de Colina/complicaciones , Modelos Animales de Enfermedad , Ácidos Grasos Omega-3/administración & dosificación , Hígado Graso/metabolismo , Masculino , Mitocondrias Hepáticas/metabolismo , Fosforilación , Ratas , Ratas Wistar , Especies Reactivas de Oxígeno , Índice de Severidad de la EnfermedadRESUMEN
Oxidative stress and hepatic mitochondria play a role in the pathogenesis of nonalcoholic fatty liver disease. The aim of the present study was to evaluate the role of hepatic mitochondrial dysfunction and oxidative stress in the pathogenesis of the disease. Fatty liver was induced in Wistar rats with a choline-deficient diet (CD; N = 7) or a high-fat diet enriched with PUFAs-omega-3 (H; N = 7) for 4 weeks. The control group (N = 7) was fed a standard diet. Liver mitochondrial oxidation and phosphorylation were measured polarographically and oxidative stress was estimated on the basis of malondialdehyde and glutathione concentrations. Moderate macrovacuolar liver steatosis was observed in the CD group and mild liver steatosis was observed in the periportal area in the H group. There was an increase in the oxygen consumption rate by liver mitochondria in respiratory state 4 (S4) and a decrease in respiratory control rate (RCR) in the CD group (S4: 32.70 ± 3.35; RCR: 2.55 ± 0.15 ng atoms of O2 min-1 mg protein-1) when compared to the H and control groups (S4: 23.09 ± 1.53, 17.04 ± 2.03, RCR: 3.15 ± 0.15, 3.68 ± 0.15 ng atoms of O2 min-1 mg protein-1, respectively), P < 0.05. Hepatic lipoperoxide concentrations were significantly increased and the concentration of reduced glutathione was significantly reduced in the CD group. A choline-deficient diet causes moderate steatosis with disruption of liver mitochondrial function and increased oxidative stress. These data suggest that lipid peroxidation products can impair the flow of electrons along the respiratory chain, causing overreduction of respiratory chain components and enhanced mitochondrial reactive oxygen species. These findings are important in the pathogenesis of nonalcoholic fatty liver disease.
Asunto(s)
Animales , Masculino , Ratas , Hígado Graso/etiología , Mitocondrias Hepáticas/fisiología , Enfermedades Mitocondriales/complicaciones , Estrés Oxidativo/fisiología , Deficiencia de Colina/complicaciones , Modelos Animales de Enfermedad , /administración & dosificación , Hígado Graso/metabolismo , Mitocondrias Hepáticas/metabolismo , Fosforilación , Ratas Wistar , Especies Reactivas de Oxígeno , Índice de Severidad de la EnfermedadRESUMEN
OBJECTIVE: The "red complex," composed of Bacteroides forsythus, Porphyromonas gingivalis, and Treponema denticola, is implicated in severe forms of periodontal diseases. The purpose of this study was to assess the occurrence of the red complex in root canal infections through the use of a sensitive technique-the 16S rDNA-directed polymerase chain reaction (PCR). STUDY DESIGN: Samples were obtained from 50 necrotic pulps with periradicular pathosis. Ten cases were diagnosed as acute periradicular abscesses. DNA was extracted from the samples and analyzed with a PCR-based identification assay. RESULTS: At least 1 member of the red complex was found in 33 of 50 cases. T denticola, P gingivalis, and B forsythus were detected in 44%, 30%, and 26% of the cases, respectively. The red complex was found in 4 of 50 cases. No particular signs or symptoms were associated with the presence of these bacterial species. CONCLUSIONS: Despite what is indicated in reports with respect to marginal periodontitis, red complex bacteria-either singularly or collectively-was not associated with any particular pattern of clinical symptoms. However, because the bacterial species from the red complex are recognized oral pathogens, their occurrence in root canal infections suggests that they may play a role in the pathogenesis of periradicular diseases.
Asunto(s)
Bacteroides/patogenicidad , Absceso Periapical/microbiología , Porphyromonas gingivalis/patogenicidad , Treponema/patogenicidad , Adolescente , Adulto , Infecciones Bacterianas/genética , Bacteroides/aislamiento & purificación , Distribución de Chi-Cuadrado , Sondas de ADN , ADN Bacteriano/análisis , ADN Ribosómico/análisis , Humanos , Persona de Mediana Edad , Reacción en Cadena de la Polimerasa , Porphyromonas gingivalis/aislamiento & purificación , Treponema/aislamiento & purificaciónRESUMEN
BACKGROUND: The pathogenesis of non-steroidal anti-inflammatory drug (NSAID) enteropathy is complex. It involves uncoupling of mitochondrial oxidative phosphorylation which alters the intercellular junction and increases intestinal permeability with consequent intestinal damage. Metronidazole diminishes the inflammation induced by indomethacin but the mechanisms remain speculative. A direct effect on luminal bacteria has traditionally been thought to account for the protective effect of metronidazole. However, a protective effect of metronidazole on mitochondrial oxidative phosphorylation has never been tested. AIMS: To assess the protective effect of metronidazole on mitochondrial uncoupling induced by indomethacin and also on the increased intestinal permeability and macroscopic damage. MATERIAL AND METHODS: The protective effect of metronidazole was evaluated in rats given indomethacin; a macroscopic score was devised to quantify intestinal lesions, and intestinal permeability was measured by means of (51)Cr-ethylenediaminetetraacetic acid. The protective effect of metronidazole against mitochondrial uncoupling induced by indomethacin was assessed using isolated coupled rat liver mitochondria obtained from rats pretreated with metronidazole or saline. RESULTS: Metronidazole significantly reduced the macroscopic intestinal damage and increase in intestinal permeability induced by indomethacin; furthermore, at the mitochondrial level, it significantly reduced the increase in oxygen consumption in state 4 induced by indomethacin and caused less reduction of the respiratory control rate. CONCLUSION: Our study confirmed the beneficial effects of metronidazole on intestinal damage and intestinal permeability, and demonstrated, for the first time, a direct protective effect of metronidazole on uncoupling of mitochondrial oxidative phosphorylation caused by NSAIDs.
Asunto(s)
Antibacterianos/farmacología , Antiinflamatorios no Esteroideos/farmacología , Indometacina/farmacología , Metronidazol/farmacología , Mitocondrias/efectos de los fármacos , Fosforilación Oxidativa/efectos de los fármacos , Animales , Interacciones Farmacológicas , Mucosa Intestinal/efectos de los fármacos , Mucosa Intestinal/metabolismo , Masculino , Consumo de Oxígeno/efectos de los fármacos , Permeabilidad/efectos de los fármacos , Ratas , Ratas Wistar , Análisis de Regresión , Estadísticas no Paramétricas , Desacopladores/farmacologíaRESUMEN
Mutations at codons 12, 13, or 61 of the H-ras, K-ras, and N-ras have been detected in human neoplasias by a variety of techniques. Some of these techniques are very sensitive and can detect K-ras mutation in 90% of the cases of pancreatic adenocarcinomas. We analyzed 11 samples of pancreatic adenocarcinoma, three samples of pancreatic mucinous cystadenoma, and two samples without tumors in formalin-fixed paraffin embedded tissue sections. K-ras mutations at codon 12 were detected by a two-step PCR-enriched technique in all the samples of pancreatic adenocarcinoma, but not in cystadenoma or control samples. This technique may be useful for early detection of pancreatic cancer.
Asunto(s)
Codón/análisis , Genes ras/genética , Mutación/genética , Neoplasias Pancreáticas/genética , ADN de Neoplasias/análisis , ADN de Neoplasias/aislamiento & purificación , Humanos , Sensibilidad y EspecificidadRESUMEN
Multidrug resistance in leukemic cells is associated with decreased drug accumulation. A resistant cell line and cells from 11 patients with chronic lymphoid leukemia B were used for the evaluation of intracellular accumulation of daunorubicin (DNR), idarubicin (IDA), epirubicin (EPI) and rhodamine-123 (Rh-123). Cyclosporin A (CSA) and verapamil were used to test their modulatory effects on anthracyclines and the fluorescent dye. In leukemic samples there was a tendency for a lower accumulation index in samples tested with Rh-123 as compared to anthracyclines. IDA was a poorer substrate to P-glycoprotein (Pgp) than two of its analogues, e.g. DNR and EPI. A good correlation (80%) was found between Rh-123 accumulation and Pgp expression by phosphatase-anti-alkaline phosphatase. A strict correlation (100%) was found between modulation by CSA of Rh-123 accumulation and immunoreactivity to Pgp. Two discordant results were seen suggesting that other mechanisms of resistance could be present. The Rh-123 accumulation test seems to give a better indication than anthracyclines, however, it is not selective and may allow the detection of other drug-transport pumps.
Asunto(s)
Antibióticos Antineoplásicos/metabolismo , Ciclosporina/farmacología , Colorantes Fluorescentes/metabolismo , Leucemia Linfocítica Crónica de Células B/metabolismo , Rodaminas/metabolismo , Verapamilo/farmacología , Miembro 1 de la Subfamilia B de Casetes de Unión a ATP/metabolismo , Adulto , Anciano , Anciano de 80 o más Años , Resistencia a Múltiples Medicamentos , Femenino , Citometría de Flujo , Humanos , Inmunohistoquímica , Leucemia Linfocítica Crónica de Células B/tratamiento farmacológico , Masculino , Persona de Mediana Edad , Rodamina 123 , Células Tumorales Cultivadas/efectos de los fármacos , Células Tumorales Cultivadas/metabolismoRESUMEN
The N2-Mercaptopropionylglycine (N2-MPG) is a potent antioxidant by inhibiting the abnormal production of xantina-oxidase. The aim of this research is to analyze the antioxidant capacity of this tiol compound by offering some protection to pancreatic tissue in the acute pancreatitis (AP). The induction of AP was obtained through two methods: a) supramaximal dose of cerulein; b) infusion of 2.5% sodium taurocholate into the biliopancreatic duct of the rat. Thirty-six male Wistar rats (220-270 g) were divided into four groups. AP with cerulein (Two parenteral doses of 20 micrograms/kg; one hour interval): in two groups: GI: nineteen rats previously treated with N2-MPG (100 mg/kg) ten minutes before AP. GII (control): seventeen animals which received saline 0.9%. AP with taurocholate (0.5 ml into the main biliopancreatic duct): in other two groups: GIII: eleven rats previously treated with N2-MPG (100 mg/kg) ten minutes before AP. GIV (control): fifteen animals which received saline 0.9%. The albumin leakage into the cell interstice as an inflammatory parameter was measured through Evans-Blue (EB) colorimetry, that links totally with serum albumin after injection into the pancreatic tissue, immediately before induction of AP. The rats were sacrificed one hour after. Water tissue content was also measured. There was a relevant reduction of EB leakage in GI (344 +/- 27 micrograms/gtissue) when compared to GII (729 +/- 84 micrograms/gtissue), p < 0.01, and in GIII (386 +/- 52 micrograms/gtissue) when compared to GIV (543 +/- 53 micrograms/gtissue), p < 0.05. There was no difference in tissue water content between GI (88.2 +/- 0.6%) and GII (87.4 +/- 0.9%), but certainly between GIII (77.7 +/- 2.1%) and GIV (82.8 +/- 1.2%), p < 0.05. The amilase levels didn't show any difference among the four groups. These results suggest that the use of the antioxidant N2-MPG offers a protective action, at least in rats, reducing the severity of AP induced by supramaximal dose of cerulein, and even in a more severe AP such as produced by sodium taurocholate at 2.5%, although apparently not interfering with its pathogenesis. It also strengthens the actual participation of free radicals of oxygen in the physiopathology of acute pancreatitis.
Asunto(s)
Antioxidantes/uso terapéutico , Glicina/análogos & derivados , Pancreatitis/metabolismo , Compuestos de Sulfhidrilo/farmacología , Enfermedad Aguda , Animales , Antioxidantes/metabolismo , Antioxidantes/farmacología , Modelos Animales de Enfermedad , Glicina/metabolismo , Glicina/farmacología , Glicina/uso terapéutico , Masculino , Pancreatitis/inducido químicamente , Pancreatitis/tratamiento farmacológico , Pancreatitis/patología , Ratas , Ratas Wistar , Neoplasias del Recto , Compuestos de Sulfhidrilo/metabolismo , Compuestos de Sulfhidrilo/uso terapéutico , Xantina Deshidrogenasa/metabolismo , Xantina Oxidasa/metabolismoRESUMEN
Acute Pancreatitis (AP) is known to produce morphologic and functional changes in liver. Administration of low doses of caerulein significant decreases the content of pancreatic enzymes, leading to reduced mortality of animals submitted to AP. The present study was designed to assess the effect of acute reduction of pancreatic enzymatic content in the hepatic mitochondrial function. Wistar male rats, submitted to AP by injection of Na thaurocholate into the pancreatic duct, with and without previous i.v. injection of 0.133 microgram Kg-1h-1 of caerulein for three hours, were divided in four groups: Group I: No caerulein infusion and AP; Group II: Previous caerulein infusion and with AP; Group III: Caerulein infusion without AP; Group IV (control): No caerulein infusion and without AP. After 2 hours of induction of AP the livers were removed and prepared to the mitochondrial oxidative and phosphorylative activities, measured polarographically with determination of oxygen consumption without ADP (Basal respiration-State 4) and in the presence of ADP (Activated respiration-State 3). Ascitic fluid contents of amylase, trypsin and total protein were routinely determined. After 2 hours of AP there was a significant increase in state 4 respiration (41%) and a decrease in respiratory control ratio and in ADP/O ratio (p < 0.05) in animals of Group I (AP without caerulein) when compared to Group II (AP with caerulein) (Table 1). Ascitic fluids contents of amylase (A) and trypsin (T) showed decrease in animals of Group II with AP that received previous caerulein infusion (A = 80 +/- 10 U/ml, T = 9.75 +/- 1.25 U/ml), when compared to animals of Group I that did not receive caerulein (A = 231 +/- 24, T = 40.32 +/- 5.19) (p < 0.001). Caerulein infusion by itself (Group III) did not have any influence on mitochondrial liver dysfunction. Reduction of pancreatic enzyme content through caerulein infusion attenuates the damage of mitochondrial respiration, demonstrated by uncoupling phase on mitochondrial function in experimental AP. Further studies are needed to elucidate this phenomena, but it is probably related to the decreased of the pathogenic effects of pancreatic activated enzymes that reach the systemic circulation in reduced amounts.
Asunto(s)
Hígado/patología , Páncreas/enzimología , Pancreatitis/patología , Enfermedad Aguda , Animales , Líquido Ascítico/química , Ceruletida , Fármacos Gastrointestinales , Hígado/metabolismo , Masculino , Mitocondrias/metabolismo , Pancreatitis/sangre , Pancreatitis/inducido químicamente , Ratas , Ratas WistarRESUMEN
We studied the alterations in the metabolism of liver mitochondria in rats with acute pancreatitis. Male Wistar rats were allocated to a control group (group I) and to five other groups corresponding to 2, 4, 12, 24 and 48 h after the induction of acute pancreatitis by the injection of 5% sodium taurocholate into the pancreatic duct. Sham-operated animals were submitted to the same surgical steps except for the induction of acute pancreatitis. Mitochondrial oxidation and phosphorylation were measured polarographically by determining oxygen consumption without ADP (basal respiration, state 4) and in the presence of ADP (activated respiration, state 3). Serum amylase, transaminases (ALT and AST) and protein were also determined. Ascitic fluid, contents of amylase, trypsin and total protein were also determined and arterial blood pressure was measured in all groups. In ascitic fluid, trypsin and amylase increased reaching a maximum at 2 and 4 h, respectively. Serum amylase increased at 2 h reaching a maximum at 4 h. Serum transaminase levels increased at 12 and 24 h. After 2 h (and also 4 h) there was an increase in state 4 respiration (45.65 +/- 1.79 vs 28.96 +/- 1.50) and a decrease in respiration control rate (3.53 +/- 0.09 vs 4.45 +/- 0.08) and in the ADP/O ratio (1.77 +/- 0.02 vs 1.91 +/- 0.01) compared to controls (P < 0.05). These results indicate a disruption of mitochondrial function, which recovered after 12 h. In the 48-h groups there was mitochondrial damage similar to that occurring in ischemic lesion. Beat-to-beat analysis (30 min) showed that arterial blood pressure remained normal up to 24 h (111 +/- 3 mmHg) while a significant decrease occurred in the 48-h group (91 +/- 4 mmHg). These data suggest biphasic damage in mitochondrial function in acute pancreatitis: an initial uncoupled phase, possibly secondary to enzyme activity, followed by a temporary recovery and then a late and final dysfunction, associated with arterial hypotension, possibly related to ischemic damage.
Asunto(s)
Modelos Animales de Enfermedad , Hepatopatías/etiología , Pancreatitis/complicaciones , Animales , Masculino , Ratas , Ratas WistarRESUMEN
We studied the alterations in the metabolism of liver mitochondria in rats with acute pancreatitis. Male Wistar rats were allocated to a control group (group I) and to five other groups corresponding to 2,4, 12,24 and 48 h after the induction of acute pancreatitis by the injection of 5 per cent sodium taurocholate into the pancreatic duct. Sham-operated animals were submitted to the same surgical steps except for the induction of acute pancreatitis. Mitochondrial oxidation and phosphorylation were measured polarographically by determining oxygen consumption without ADP (basal respiration, state 4) and in the presence of ADP (activated respiration, state 3). Serum amylase, transaminases (ALT and AST) and protein were also determined. Ascitic fluid, contents of amylase, trypsin and total protein were also determined and arterial blood pressure was measured in all groups. In ascitic fluid, trypsin and amylase increased reaching a maximum at 2 and 4h, respectively. Serum amylase increased at 2 h reaching a maximum at 4 h. Serum transaminase levels increased at 12 and 24 h. After 2 h (and also 4 h) there was an increase in state 4 respiration (45.65 + 1.79 vs 28.96 + 1.50) and a decrease in respiration control rate (3.53 + 0.09 vs 4.45 + 0.08) and in the ADP/O ratio (1.77 + 0.02 vs 1.91 + 0.01) compared to controls (P<0.05). These results indicate a disruption of mitochondrial function, which recovered after 12 h. In the 48-h groups there was mitochondrial damage similar to that occurring in ischemic lesion. Beat-to-beat analysis (30 min) showed that arterial blood pressure remained normal up to 24 h (111 + 3 mmHg) while a significant decrease occurred in the 48-h group (91 + 4 mmHg). These data suggest biphasic damage in mitochondrial function in acute pancreatitis: an inital uncoupled phase, possibly secondary to enzyme activity, followed by a temporary recovery and then a late and final dysfunction, associated with arterial hypotension, possibly related to ischemic damage.
Asunto(s)
Ratas , Animales , Masculino , Modelos Animales de Enfermedad , Hígado/fisiopatología , Pancreatitis/fisiopatología , Ratas WistarRESUMEN
Previously the heat-stable enterotoxin in Vibrio cholerae and V. mimicus has been detected by suckling mouse assay, a non-specific approach, and by DNA probes, a time-consuming method. This report describes a polymerase chain reaction (PCR) procedure for the detection of the stn (NAG-ST) and sto (O1-ST) gene sequences that is rapid and specific, allowing toxin gene molecular characterisation. A total of 34 V. cholerae and V. mimicus isolates was examined for ST and CT genes. The NAG-ST gene sequence was amplified in 13 of 22 non-O1/non-O139 V. cholerae and three of five V. mimicus strains. A new enterotoxin gene sequence pattern was found with MseI and TaqI restriction endonuclease PCR fragment digestion of two V. cholerae isolates, in addition to the pattern anticipated from the Genbank sequence, and found with the other ST+. These results show that ST-PCR detection is useful for the characterisation of V. cholerae and V. mimicus.
Asunto(s)
Enterotoxinas/genética , Vibrio cholerae/genética , Vibrio/genética , Animales , ADN Bacteriano/análisis , Electroforesis en Gel de Poliacrilamida , Genes Bacterianos , Humanos , Reacción en Cadena de la Polimerasa , Polimorfismo de Longitud del Fragmento de Restricción , Sensibilidad y Especificidad , Vibrio/clasificación , Vibrio cholerae/clasificaciónRESUMEN
Alternative therapy for refractory leukemic patients is being increasingly adopted. Circumvention of multidrug resistance represents a strategy that has been taken into account when conventional chemotherapy failed. In this work a group of 15 refractory, heavily pretreated, patients was enrolled in a circumvention protocol including etoposide (ETO) and cyclosporin A (CSA). All patients received etoposide prior to this schedule. Toxicity to circumvention protocol was acceptable and only one serious side-effect was observed. Two hematological clinical responses were seen, both of which were positive to P-glycoprotein immunostaining and exhibited in vitro modulation by CSA in cultures using the thymidine incorporation assay. Three out of four patients negative for P-glycoprotein achieved a minor response. Three out of six clinical failures were also negative for Pgp immunostaining one of which exhibited sinergistic effect between ETO and CSA. Our study suggests that hematological response to ETO and CSA association can be obtained in intensely pretreated leukemic patients. Several factors may affect the response such as clinical status before this therapy. Additionally, it also suggests that not all CSA effects on the combination ETO-CSA can be attributed to Pgp modulation.