Estudo da permeabilidade de fármacos por meio do modelo de saco intestinal de ratos / Study of drug permeability through the rat gut sac model

Os parâmetros de permeabilidade e solubilidade são fundamentais à absorção oral de fármacos e a partir dessas características, foi criado o Sistema de Classificação Biofarmacêutica, através do qual os fármacos são divididos em quatro classes. Atualmente, para a determinação da solubilidade de um fármaco, existem diversos métodos padronizados por agências regulatórias, no entanto, para a determinação da permeabilidade, os ensaios são passíveis de diversas variações em sua execução, diminuindo a confiabilidade dos resultados obtidos e impossibilitando a comparação dos mesmos quando realizados com técnicas diferentes umas das outras. O objetivo do presente trabalho é avaliar as variáveis experimentais do modelo do saco intestinal que podem influenciar nos resultados de permeabilidade aparente de fármacos e na viabilidade do tecido. O presente estudo foi aprovado pelo Comitê de Ética no Uso de Animais da FCF-USP (109.2018-P574). Foram utilizados 33 Rattus norvegicus da linhagem Wistar, machos, jovens adultos, com peso entre 200 g e 300 g. Para realização do procedimento, cada animal permaneceu em jejum por cerca de quatro horas e após adequada anestesia a porção do jejuno do intestino delgado foi retirada e dividida em seis segmentos de aproximadamente 8,5cm cada. Foram realizados experimentos com e sem inversão do saco intestinal, submetidos a diferentes tempos de banho de gelo após sua ressecção, na presença ou ausência de inibidor da glicoproteína-P (verapamil). Os fármacos naproxeno e famotidina foram empregados como marcadores de alta e baixa permeabilidade, respectivamente. A losartana foi utilizada como substrato da glicoproteína P. Cada um dos sacos intestinais foi colocado em um tubo de ensaio contendo tampão Krebs, a 37°C, saturado com gás carbogênio. Para avaliação da integridade e viabilidade dos segmentos intestinais, observou-se a presença de movimentos peristálticos e coletaram-se amostras do meio de incubação nos tempos 0, 30, 45, 60, 90 e 120 minutos para quantificação dos fármacos e de glicose, uma vez que esta é ativamente transportada para a serosa do intestino delgado. Determinou-se a permeabilidade aparente de cada fármaco e as concentrações de glicose nas diferentes condições experimentais, realizou-se planejamento fatorial multinível e os resultados foram analisados por análise variância (ANOVA), seguida de pós-teste de Tukey. Observou-se que as variáveis experimentais interferiram de forma significativa na viabilidade tecidual e na permeabilidade aparente dos fármacos. Não foram observadas diferenças significativas da permeabilidade de fármacos nos diferentes segmentos do jejuno. A glicose mostrou-se um bom marcador de viabilidade tecidual e foi constatado que a presença ou ausência de movimentos peristálticos não está relacionada diretamente com a viabilidade do tecido. Uma vez que foram constatadas tantas interferências nos resultados, é imprescindível que os procedimentos experimentais sejam padronizados, para que os resultados apresentem menor variabilidade e possam ser comparados entre si

The permeability and solubility parameters are fundamental to the oral absorption of drugs and from these characteristics, the Biopharmaceutical Classification System was created, through which drugs are divided into four classes. Currently, for the determination of the solubility of a drug, there are several methods standardized by regulatory agencies, however, for the determination of permeability, the tests are subject to several variations in their execution, reducing the reliability of the results obtained and making it impossible to compare the results obtained. same when performed with different techniques. The aim of this study is to evaluate if different experimental conditions can influence the results of apparent drug permeability and tissue viability on gut sac model. The present study was approved by the Ethics Committee for the Use of Animals of FCF-USP (109.2018-P574). Thirty-three male, young adult Rattus norvegicus were used, weighing between 200 g and 300 g. To perform the procedure, each animal fasted for about four hours and after adequate anesthesia, the portion of the jejunum of the small intestine was removed and divided into six segments of approximately 8.5 cm each. Experiments were performed with and without inversion of the gut sac, submitted to different times of ice bath after its resection, in the presence or absence of a P-glycoprotein inhibitor (verapamil). The drugs naproxen and famotidine were used as markers of high and low permeability, respectively. Losartan was used as a substrate for P-glycoprotein. Each of the gut sacs was placed in a test tube containing Krebs buffer, at 37°C, saturated with carbogen gas. To evaluate the integrity and viability of the intestinal segments, the presence of peristaltic movements was observed and samples of the incubation medium were collected at 0, 30, 45, 60, 90 and 120 minutes for quantification of drugs and glucose, as it is actively transported to the serosa of the small intestine. The apparent permeability of each drug and the glucose concentrations were determined under different experimental conditions, multilevel factorial design was performed and the results were analyzed by analysis of variance (ANOVA), followed by Tukey's post-test. It was observed that the experimental variables significantly interfered in the tissue viability and in the apparent permeability of the drugs. No significant differences in drug permeability were observed in the different segments of the jejunum. Glucose proved to be a good marker of tissue viability and it was found that the presence or absence of peristaltic movements is not directly related to tissue viability. Since so many interferences were found in the results, it is essential that the experimental procedures be standardized, so that the results show less variability and can be compared between different authors

Hypertonic saline and pentoxifylline enhance survival, reducing apoptosis and oxidative stress in a rat model of strangulated closed loop small bowel obstruction

OBJECTIVES: Intestinal obstruction has a high mortality rate when therapeutic treatment is delayed. Resuscitation in intestinal obstruction requires a large volume of fluid, and fluid combinations have been studied. Therefore, we evaluated the effects of hypertonic saline solution (HS) with pentoxifylline (PTX) on apoptosis, oxidative stress and survival rate. METHODS: Wistar rats were subjected to intestinal obstruction and ischemia through a closed loop ligation of the terminal ileum and its vessels. After 24 hours, the necrotic bowel segment was resected, and the animals were randomized into four groups according to the following resuscitation strategies: Ringer's lactate solution (RL) (RL-32 ml/kg); RL+PTX (25 mg/kg); HS+PTX (HS, 7.5%, 4 ml/kg), and no resuscitation (IO-intestinal obstruction and ischemia). Euthanasia was performed 3 hours after resuscitation to obtain kidney and intestine samples. A malondialdehyde (MDA) assay was performed to evaluate oxidative stress, and histochemical analyses (terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling [TUNEL], Bcl-2 and Bax) were conducted to evaluate kidney apoptosis. Survival was analyzed with another series of animals that were observed for 15 days. RESULTS: PTX in combination with RL or HS reduced the MDA levels (nmol/mg of protein), as follows: kidney IO=0.42; RL=0.49; RL+PTX=0.31; HS+PTX=0.34 (p<0.05); intestine: IO=0.42; RL=0.48; RL+PTX=0.29; HS+PTX=0.26 (p<0.05). The number of labeled cells for TUNEL and Bax was lower in the HS+PTX group than in the other groups (p<0.05). The Bax/Bcl-2 ratio was lower in the HS+PTX group than in the other groups (p<0.05). The survival rate on the 15th day was higher in the HS+PTX group (77%) than in the RL+PTX group (11%). CONCLUSION: PTX in combination with HS enhanced survival and attenuated oxidative stress and apoptosis. However, when combined with RL, PTX did not reduce apoptosis or mortality.

La controvertida participación del GLP-1 en los mecanismos fisiológicos desencadenados tras cirugía bariátrica / The controversial participation of GLP-1 in the physiological mechanisms unchained after bariatric surgery

RESUMEN: Numerosas hipótesis se invocan para explicar el efecto beneficioso sobre el metabolismo glucídico tras la cirugía bariátrica. Algunos autores abogan por la secreción y liberación de distintas sustancias con funciones endocrinas (enterohormonas). Una de las sustancias más señaladas como efector, con efectos contrastados pero datos controvertidos, es el GLP-1. Nuestro estudio se realizó en ratas Wistar macho sanas, para evitar la ausencia de factores de confusión como son la DMT2 y la obesidad. Para conocer el mapa de adaptación a la secreción de GLP-1 tras la cirugía, se designaron 5 grupos: dos grupos control (de ayuno y de estrés quirúrgico); y tres grupos quirúrgicos (gastrectomía vertical, resección del 50 % del intestino medio y el Bypass gástrico con montaje en Y de Roux). Después de tres meses se estudiaron mediante técnicas inmunohistoquímicas el patrón de síntesis de GLP-1 en las distintas porciones del intestino delgado. También se estudió la expresión de los receptores de membrana en las células de los islotes pancreáticos. Se observó la existencia de un significativo aumento del número de células secretoras en íleon, duodeno y yeyuno en los grupos quirúrgicos de técnicas mixtas (RYGB) y malabsortivas (RI50). Igualmente se observó una elevación de los receptores pancreáticos en las mismas técnicas frente a los controles. Nuestros datos indican que la secreción intestinal de GLP-1 y su sensibilidad a nivel pancreáticas están aumentada, como efecto adaptativo a la agresión mecánica del tubo y a la alteración del flujo de nutrientes tras la cirugía.

SUMMARY: Numerous hypotheses are invoked to explain the beneficial effect on glucose metabolism after bariatric surgery. Some authors advocate for the secretion and release of various substances with endocrine functions (enterohormones). One of the substances most marked as effector, with contrasting effects but controversial data, is Glucagon-like peptide-1 GLP-1. Our study was performed in healthy male Wistar rats, to avoid the absence of confounding factors such as DMT2 and obesity. In order to know the map of adaptation to GLP-1 secretion after surgery, five groups were designated: Two control groups (fasting and surgical stress); and three surgical groups (vertical sleeve gastrectomy, 50 % midgut resection and Roux-en-Y gastric bypass). After three months, the GLP-1 synthesis pattern was studied by immunohistochemical techniques in the different portions of the small digestive tract. The expression of membrane receptors in pancreatic islet cells was also studied. There was a significant increase in the number of secretory cells in ileum, duodenum and jejunum in mixed surgical (RYGB) and malabsorptive (RI50) groups. An elevation of pancreatic receptors was also observed in the same techniques against controls. Our data indicated that intestinal secretion of GLP1 and its sensitivity to the pancreatic level were increased, both to an adaptive effect to the mechanical aggression of the digestive tube and to the alteration of nutrient flow after surgery.

Absorption mechanism of three curcumin constituents through in situ intestinal perfusion method

This study aimed to investigate the absorption mechanism of three curcumin constituents in rat small intestines. Self-emulsification was used to solubilize the three curcumin constituents, and the rat in situ intestinal perfusion method was used to study factors on drug absorption, including drug mass concentration, absorption site, and the different types and concentrations of absorption inhibitors. Within the scope of experimental concentrations, three curcumin constituents were absorbed in rat small intestines through the active transport mechanism.

Tendencias en la mortalidad por cáncer en México de 1980 a 2011 / Cancer mortality trends in Mexico, 1980-2011

Objetivo. Evaluar las tendencias de mortalidad por cáncer en México entre 1980 y 2011. Material y métodos. Se calcularon las tasas de mortalidad ajustadas por edad y sexo para todos los cánceres y para las 15 localizaciones más frecuentes mediante el método directo y tomando como población estándar la población mundial de 2010. Las tendencias en las tasas de mortalidad y el cambio porcentual anual para cada tipo de cáncer se estimaron a través de un modelo de regresión joinpoint. Resultados. A partir de 2004 y como consecuencia de la reducción de la mortalidad por cáncer de pulmón (-3.2% en hombres y -1.8% en mujeres), estómago (-2.1% en hombres y -2.4% en mujeres) y cérvix (-4.7%), se observó una disminución significativa (~1% anual) en la mortalidad por cáncer en general tanto en el grupo de todas las edades como en el de 35 a 64 años para ambos sexos. La mortalidad por otros cánceres como el de mama y el de ovario, en las mujeres o el de próstata, en los hombres, mostró un aumento sostenido. Conclusiones. Algunas de las reducciones en la mortalidad por cáncer pueden ser parcialmente atribuidas a la efectividad de los programas de prevención establecidos. Sin embargo, se requiere implementar registros adecuados de cáncer con base poblacional para evaluar el impacto real de estos programas, así como diseñar y evaluar intervenciones innovadoras que permitan desarrollar políticas de prevención más costo-efectivas.

Objective. To evaluate trends in cancer mortality in Mexico between 1980-2011. Material and methods. Through direct method and using World Population 2010 as standard population, mortality rates for all cancers and the 15 most frequent locations, adjusted for age and sex were calculated. Trends in mortality rates and annual percentage change for each type of cancer were estimated by joinpoint regression model. Results. As a result of the reduction in mortality from lung cancer (-3.2% -1.8% in men and in women), stomach (-2.1% -2.4% in men and in women) and cervix (-4.7%); since 2004 a significant (~1% per year) decline was observed in cancer mortality in general, in all ages, and in the group of 35-64 years of both sexes. Other cancers such as breast and ovarian cancer in women; as well as for prostate cancer in men, showed a steady increase. Conclusions. Some of the reductions in cancer mortality may be partially attributed to the effectiveness of prevention programs. However, adequate records of population-based cancer are needed to assess the real impact of these programs; as well as designing and evaluating innovative interventions to develop more cost-effective prevention policies.

Expressao genica associada ao estresse oxidativo no coracao de camundongo apos isquemia intestinal / Gene expression related to oxidative stress in the heart of mice after intestinal ischemia

FUNDAMENTO: O fenômeno da isquemia e reperfusão intestinal é um evento frequente na clínica e está associado a repercussões deletérias em órgãos a distância, em especial ao coração. OBJETIVO: Investigar a expressão gênica do estresse oxidativo e defesa antioxidante no coração de camundongos isogênicos, submetidos a isquemia e reperfusão intestinal (IR). MÉTODOS: Doze camundongos (C57BL/6) foram distribuídos em dois grupos: Grupo IR (GIR) com 60 min de oclusão da artéria mesentérica superior, seguidos de 60 min de reperfusão. Grupo Controle (GC) submetidos a anestesia e a laparotomia sem o procedimento de IR observados por 120 min. As amostras de intestino e coração foram processadas pelo método (RT-qPCR / Reverse transcriptase - quantitative Polymerase Chain Reaction) para determinar a expressão gênica de 84 genes relacionados ao estresse oxidativo ("t" de Student, p < 0,05). RESULTADOS: Observou-se no tecido intestinal (GIR) uma expressão significantemente aumentada em 65 (74,71%) genes em relação ao tecido normal (GC), e 37 (44,04%) genes estiveram hiperexpressos (maior que três vezes o limiar permitido pelo algoritmo). No tocante aos efeitos da I/R intestinal a distância no tecido cardíaco verificou-se a expressão significantemente aumentada de 28 genes (33,33%), mas somente oito genes (9,52%) se hiperexpressaram três vezes acima do limiar. Quatro (7,14%) desses oito genes se expressaram simultaneamente nos tecidos intestinal e cardíaco. No GIR notaram-se cardiomiócitos com núcleos de menor tamanho, picnóticos, ricos em heterocromatina e raros nucléolos, indicando sofrimento cardíaco. CONCLUSÃO: A I/R intestinal promoveu a hiperexpressão estatisticamente significante de oito genes associados ao ...

BACKGROUND: Intestinal ischemia-reperfusion is a frequent clinical event associated to injury in distant organs, especially the heart. OBJECTIVE: To investigate the gene expression of oxidative stress and antioxidant defense in the heart of inbred mice subjected to intestinal ischemia and reperfusion (IR). METHODS: Twelve mice (C57BL / 6) were assigned to: IR Group (GIR) with 60 minutes of superior mesenteric artery occlusion followed by 60 minutes of reperfusion; Control Group (CG) which underwent anesthesia and laparotomy without IR procedure and was observed for 120 minutes. Intestine and heart samples were processed using the RT-qPCR / Reverse transcriptase-quantitative Polymerase Chain Reaction method for the gene expression of 84 genes related to oxidative stress and oxidative defense (Student's "t" test, p < 0.05). RESULTS: The intestinal tissue (GIR) was noted to have an up-regulation of 65 genes (74.71%) in comparison to normal tissue (CG), and 37 genes (44.04%) were hyper-expressed (greater than three times the threshold allowed by the algorithm). Regarding the remote effects of intestinal I/R in cardiac tissue an up-regulation of 28 genes (33.33%) was seen, but only eight genes (9.52%) were hyper-expressed three times above threshold. Four (7.14%) of these eight genes were expressed in both intestinal and cardiac tissues. Cardiomyocytes with smaller and pyknotic nuclei, rich in heterochromatin with rare nucleoli, indicating cardiac distress, were observed in the GIR. CONCLUSION: Intestinal I/R caused a statistically significant over expression of 8 genes associated with oxidative stress in remote myocardial tissue. .

Expression of oxidative stress and antioxidant defense genes in the kidney of inbred mice after intestinal ischemia and reperfusion

PURPOSE: To determine the gene expressions profile related to the oxidative stress and the antioxidant response in the kidneys of mice subjected to intestinal ischemia and reperfusion. METHODS: Twelve inbred mice (C57BL/6) were randomly assigned to one of two groups: the control group (CG) underwent anesthesia and was observed for 120 min and the ischemia/reperfusion group (IRG), animals were anesthetized and subjected to laparotomy and ischemia for 60 minutes followed by 60 minutes of reperfusion. The expressions of 84 genes from the kidney were determined by the Reverse Transcription qualitative Polymerase Chain Reaction (RT-qPCR). All genes that were up regulated by more than threefold using the algorithm [2(ΔΔCt)] were considered statically significant (p<0.05). RESULTS: In the IRG group 29 (34.52%) of 84 genes, were up regulated by more than threefold. The genes that were differentially up regulated in the glutathione peroxidase cluster (10 genes): were Gpx2 and Gpx7. The genes that were up regulated in the peroxidase cluster (16 genes) were following: Duox1, Epx, Lpo, Mpo, Ptgs2, Rag2, Serpinb1b, Tmod1 and Tpo. The genes that up regulated in the reactive oxygen species cluster (16 genes): Il19, Il22, Nos2, Nox1, Noxa1, Noxo1, Recql4 and Sod2. The genes that were up regulated in the oxidative stress cluster (22 genes) were: Mpp4, Nudt15, Upc3 and Xpa. The genes that were up regulated in the oxygen carriers cluster (12 genes) were: Hbq1, Mb, Ngb, Slc38a1 and Xirp1. The peroxiredoxins genes (10) showed no consistent differential regulation. CONCLUSION: The genes related to oxidative stress and antioxidant defense showed increased expression in renal tissue trigged intestinal ischemia and reperfusion.

Mucosite e alterações de permeabilidade intestinal em pacientes portadores de câncer colorretal metastático tratados com 5-Fluorouracil (5-FU) e Irinotecano(CPT-11) / Gastrintestinal mucositis and permeability test abnormalities in metastatic colorectal cancer patients treated with 5-Fluoruracil (5-FU) and Irinotecan (CPT-11)

Introdução: A melhora da sobrevida dos pacientes com Câncer Colorretal (CCR)metastático proporciona a exposição por prolongados períodos aos agentes antineoplásicos disponíveis para o seu tratamento. A compreensão dos mecanismos envolvidos na mucosite gastrintestinal provocada por regimes com irinotecano, fluorouracil e ácido folínico (IFL) é necessária para que se trate com eficácia e segurança o maior número possível de doentes. No entanto, o relato das alterações intestinais associadas à mucosite é até hoje baseada apenas em escalas de sintomas(diarréia, dor abdominal, vômitos e náuseas) referidas pelos pacientes. Métodos: Utilizando o teste de permeabilidade intestinal pela recuperação urinária de lactulose e manitol, 25 pacientes de CCR metastático tratados com regime IFL em primeira linha foram submetidos à comparação da permeabilidade intestinal antes do início do tratamento e após 15 e 29 dias do primeiro ciclo do regime. Amostras do soro dos pacientes no momento antecedente ao início do tratamento e após 15 dias do seu início foram colhidas para comparação da dosagem de citocinas potencialmente envolvidas na mediação da mucosite gastrintestinal (TNFα, IL-1β, IL-6, IL-18, IL-18bp e IL-33). Resultados: Em comparação com o controle pré-tratamento, as alterações de permeabilidade intestinal foram significativas (p<0,05) nos dois momentos (D15 e D29) do primeiro ciclo de IFL, guardando correlação significativa com a intensidade de diarréia e dor abdominal reportada pelos pacientes. Não foi possível, entretanto, demonstrar alterações séricas significativas na dosagem dascitocinas testadas em comparação ao controle pré-tratamento. Conclusão: O teste de permeabilidade intestinal, portanto, foi capaz de detectar as alterações funcionais de permeabilidade intestinal secundárias à mucosite gastrintestinal provocada pelo regime IFL em pacientes portadores de CCR metastático.

Introduction: The improving survival of patients with metastatic colorectal cancer (CRC) promotes the increasing exposure of individuals with this disease to the effective systemic antineoplastic agents available. The comprehension of the mechanisms involved in the gastrointestinal mucositis associated with the bolus regimen with irinotecan, fluoruracil and folinic acid (IFL) is necessary to improve its efficacy and safety. Nevertheless, it is difficult to obtain a reliable measurement of the intestinal abnormalities caused by mucositis, as almost all the clinical dataavailable is reported on a patient reported symptoms-based scale analysis. Methods: With the lactulose/ mannitol intestinal permeability test as the experimental platform, 25 metastatic CRC patients treated with first line IFL were compared at the 15th and 29th day of the firs IFL cycle to each one’s pretreatment intestinal permeability performance. Blood samples of the same patients were obtained at the preceding moment of the treatment and at 15th day of the IFL first cycle to compare the serum levels of potentially involved cytokines (TNFα, IL-1β, IL-6, IL-18, IL-18bp e IL-33).Results: In contrast with the pretreatment controls, the permeability test was significantly (p<0,05) altered at the D15 and D29 in patients taking first cycle IFL.This abnormalities also reserve significant (p<0,05) correlation with the diarrhea and abdominal pain symptoms scale reported by the patients. Otherwise, it has not been possible to demonstrate serum levels differences of the addressed cytokines before and after the treatment. Conclusion: The lactulose/ mannitol permeability test is an detects functional intestinal abnormalities secondary to gastrointestinal mucositis caused by IFL to metastatic CRC patients.

Depressed Neuronal Growth Associated Protein (GAP)-43 Expression in the Small Intestines of Mice Experimentally Infected with Neodiplostomum seoulense

Neodiplostomum seoulense (Digenea: Neodiplostomidae) is an intestinal trematode that can cause severe mucosal pathology in the small intestines of mice and even mortality of the infected mice within 28 days after infection. We observed neuronal growth associated protein-43 (GAP-43) expression in the myenteric plexus of the small intestinal wall of N. seoulense-infected mice until day 35 post-infection (PI). BALB/c mice were infected with 200 or 500 N. seoulense metacercariae isolated from naturally infected snakes and were killed every 7 days for immunohistochemical demonstration of GAP-43 in the small intestines. N. seoulense-infected mice showed remarkable dilatation of intestinal loops compared with control mice through days 7-28 PI. Conversely, GAP-43 expression in the mucosal myenteric plexus was markedly (P<0.05) reduced in the small intestines of N. seoulense-infected mice during days 7-28 PI and was slightly normalized at day 35 PI. From this study, it is evident that neuronal damage occurs in the intestinal mucosa of N. seoulense-infected mice. However, the correlation between intestinal pathology, including the loop dilatation, and depressed GAP-43 expression remains to be elucidated.

Aspectos funcionais, microbiológicos e morfológicos intestinais em crianças infectadas pelo vírus da imunodeficiência humana / Functional, microbiological and morphological intestinal findings among human immunodeficiency virus infected children

RACIONAL: O trato gastrointestinal é freqüentemente acometido nas crianças infectadas pelo vírus da imunodeficiência humana, com importantes repercussões no seu estado nutricional e sobrevida. A maioria dos estudos relacionados a esse tema foi desenvolvida com adultos, sendo menos investigado o problema nas crianças OBJETIVOS: Estudar aspectos digestivo-absortivos, microbiológicos e morfológicos intestinais em crianças infectadas pelo vírus da imunodeficiência humana MATERIAL E MÉTODOS: Onze crianças infectadas pelo vírus da imunodeficiência humana, menores de 13 anos, pertencentes às categorias clínicas A, B ou C, divididas em dois grupos: cinco pacientes com relato atual ou recente de diarréia e seis pacientes sem diarréia nos 30 dias que antecederam à inclusão no estudo. Investigação proposta: biopsia de intestino delgado e reto para análise morfológica e microbiológica, coprocultura, protoparasitológico de fezes, pesquisa de rotavírus, micobactérias e Cryptosporidium; teste da D-xilose RESULTADOS: Todos os pacientes testados (9/11) apresentavam má absorção da D-xilose (8,4-24,4 mg/dL). Os achados histopatológicos de intestino delgado foram inespecíficos, representados em sua maioria, por enteropatia grau I a II (6/10). Em todos os casos foi constatado aumento do infiltrado celular do córion. As alterações histopatológicas do reto também foram inespecíficas, com presença de aumento do infiltrado celular do córion. A pesquisa de microorganismos enteropatogênicos só foi positiva em dois casos, sendo identificado Mycobacterium avium intracellulare e Cryptosporidium nas fezes CONCLUSÕES: Demonstrou-se alta prevalência (100 por cento) de má absorção intestinal em crianças infectadas pelo vírus da imunodeficiência humana, com ou sem diarréia. Não foi possível estabelecer correlações quanto à presença de agentes enteropatogênicos, má absorção intestinal, alterações morfológicas intestinais e ocorrência ou não de diarréia. Não houve correlação...

BACKGROUD: Gastrointestinal tract disorders are frequent among human immunodeficiency virus infected children, with important repercussions on nutrition and survival. Most studies related to this subject were restricted to adults, being less investigated the problem in the children. AIMS: To study intestinal digestion, absorption, microbiological and morphological findings among human immunodeficiency virus infected children. MATERIAL AND METHODS: Eleven human immunodeficiency virus infected children under 13 years old, belonging to clinical categories A, B or C, separated in two groups: five patients with current or recent episode of diarrhea and six patients without diarrhea in the last 30 days preceding entering in study. Investigation proposed: microbiological and morphological analysis of small intestine and rectum biopsy; stool exams for bacterium, parasite, rotavirus, Mycobacterium species and Cryptosporidium; D-xylose test RESULTS: All tested subjects (9/11) had low D-xylose absorption (8,4 _ 24,4 mg d/L). Small intestinal mucosa histology findings were nonspecific, represented, in majority, of grade I/II enteropathy (6/10). Increased cellular infiltration of the chorion was observed in all specimens. Rectum histology alterations were also nonspecific, with chorion increased cellular infiltration. Mycobacterim avium intracellulare and Cryptosporidium were the solely microorganisms founded, both in stool CONCLUSIONS: Our study demonstrated high prevalence (100 percent) of intestinal malabsorption among human immunodeficiency virus infected children, despite the occurrence or not of diarrhea. It was not possible to establish relationships between the presence of microorganisms, intestinal malabsorption, intestinal morphologic findings and the occurrence or not of diarrhea. There was no correlation between D-xylose and intensity of villous atrophy.

Efecto de la diarrea sobre la utilización de nutrientes en ratas con desnutrición proteico-calórica o proteica / Effect of diarrhea on nutrient utilization in protein deficient or protein-calorie deficient rats

La diarrea magnifica los efectos de la desnutrición. En consecuencia, aquí se estudió el efecto de la diarrea sobre dos tipos de desnutrición (proteica y proteico-calórica). El experimento incluyó 42 ratas jóvenes de la cepa Sprague Dawley que se distribuyeron en tres grupos (14 ratas/grupo). Durante los primeros 16 días del experimento, el primer grupo recibió una dieta control ad-libitum, el segundo recibió la misma dieta pero su consumo se redujo en un 50% y el tercer grupo recibió una dieta deficiente en proteína. Al final de este período había ratas bien nutridas (controles) y con desnutrición proteica y calórico-proteica. Luego, a la mitad de estas ratas en cada grupo, se les produjo diarrea con lactosa y todas las ratas continuaron con su dieta y el régimen de alimentación preasignado durante una semana. Así, durante este período había ratas controles así como con deficiencia proteica o calórico-proteica que tenían diarrea y grupos idénticos que no tenían diarrea. Los resultados mostraron que la diarrea causó una disminución del consumo y del crecimiento en las ratas del grupo control y deficiente en proteína. Sin embargo, el grupo con deficiencia calórico-proteica no redujo su consumo ni disminuyó su crecimiento en respuesta a la diarrea. La consecuencia de esto fue que la diarrea produjo desnutrición en el grupo control y aumentó la desnutrición en el grupo deficiente en proteína, pero no tuvo un efecto adicional en el grupo con deficiencia calórico-proteica. Además, la reducción en la absorción aparente del nitrógeno y de la grasa asociada con la diarrea, fue mayor en las ratas deficientes en proteína. Este grupo también presentó las actividades más bajas de disacaridasas intestinales. Esto resultados muestran que la diarrea tiene un efecto negativo mayor en ratas con deficiencia proteica que con deficiencia calórico-proteica.

Diarrhea increases the effects of malnutrition. Accordingly, the effect of diarrhea on two types of malnutrition (protein deficiency and protein-calorie deficiency) was studied. The experiment included 42 young Sprague Dawley rats. The rats were distributed into three groups with 14 rats per group. During the first 16 of the experiment, the first group was fed a control diet ad libitum, the second received the same diet but with food intake reduced in 50% whereas the third group was offered a protein deficient diet. Thus, at the end of this period there were well-fed rats (control), as well as protein and protein-calorie malnourished rats. Then one half of the rats in each group were given lactose to produce diarrhea and all rats continued with their previously assigned diet and feeding regime during one more week. Therefore, during this period there were control rats, protein deficient rats and protein-calorie deficient rats with and without diarrhea. The results showed that diarrhea caused a substantial reduction in food intake and growth in the well-fed rats and also in the group fed the protein deficient diet. However, the protein-calorie deficient group did not reduce its intake nor its growth rate. As a result, diarrhea caused malnutrition in the control group and increased malnutrition in the protein deficient but it did not have an additional effect in the protein-calorie deficient rats. The apparent absorption of lipids and nitrogen measured in these rats showed that the absorption reduction caused by diarrhea was more pronounced in the protein deficient group. This group also had the lowest activities of intestinal disaccharidases. These results showed that diarrhea had a more detrimental effect in protein deficient than in protein-calorie deficient rats.

alpha-Lipoic acid ameliorates altered colonic contractility and intestinal transit in STZ-diabetic rats.

alpha-Lipoic acid treatment (100 mg/kg/day for 2 weeks after 6 weeks of untreated diabetes) of streptozotocin diabetic rats partially but significantly reversed both reduced contractile response of distal colon to acetylcholine and delayed transit of charcoal meal in small intestine compared to diabetic control. These effects of alpha-Lipoic acid were associated with complete reversal of diabetes induced increased plasma lipid peroxidation level. alpha-Lipoic acid had no effect on any of the parameters measured in non-diabetic rats. These findings demonstrate contribution of oxidative stress in the development of physiological changes of gut in diabetes.

Determination of Roxithromycin by Liquid Chromatography/Mass Spectrometry after Multiple-Dose Oral Administration in Broilers

A highly sensitive and specific method for the determination of roxithromycin in broiler tissues by LC/MS was developed and validated. A dichloromethane extract of the sample was separated on C18 reversed-phase column with acetonitrile-50 mM ammonium acetate (80:20, v/v) as the mobile phase and analyzed by LC/MS via atmospheric pressure ionization/electrospray ionization interface. The limit of detection and limit of quantitation were 1 ng/g and 5 ng/g. The method has been successfully applied to determine for roxithromycin in various tissues of broilers. Residue concentrations were associated with administered dose. At the termination of treatment, roxithromycin was found in all collected samples for both dose groups. Liver was detected to have the highest residual concentration of roxithromycin. Residue concentrations of roxithromycin were lower than its LOQ in all tissues from both dose groups 10 days after the treatment of roxithromycin mixed with drinking water at a dose rate of 15 mg/L or 60 mg/L to each broiler for 7 days.

Topology of Scavenger Receptor Class B Type I (SR-BI) on Brush Border Membrane

Both hydropathy plot and in vitro translation results predict the topology of SR-BI; the receptor is an integral membrane protein of 509 amino acids, consisting of a short cytoplasmic N-terminus of 9 amino acids followed by a first transmembrane domain of 22 amino acids, the extracellular domain of 408 amino acids, the second transmembrane domain of 22 amino acids, and the cytoplasmic C-terminus of 47 amino acids. The immunoblot of rBBMV in the presence or absence of pAb589 peptide antigen (the C-terminal 22 amino acid residues of SR-BI) confirmed that the bands at apparent molecular weight of 140 and 210 kDa are SR-BI related protein which might be multimeric forms of SR-BI. 125I apo A-I overlay analysis showed that SR-BI can bind to its ligand, apo A-I, only when it is thoroughly matured - glycosylated and dimerized. The antibody which was generated against extracellular domain of SR-BI (pAb230) not only prevented 125I-labeled apo A-I from binding to 140 kDa band but also inhibited the esterified cholesterol uptake of rabbit BBMV with its IC50 value of 40 microgram/ml of IgG. In contrast, the antibody generated against the C-terminal domain of SR-BI (pAb589) did not show any effect either on cholesterol uptake of rabbit BBMV or 125I-labeled apo A-I binding to 140 kDa band. Overall results show that the ligand binding site of SR-BI in rabbit BBMV is located in extracellular domain, and SR-BI is only functional when it is part of dimeric forms which rationalize the previously found cooperative nature of the binding interaction and maybe a fundamental finding towards the so far poorly understood mechanism of SR-BI function.

Celiac disease: clinical features and diagnostic criteria.

Celiac disease (CD) classically presents between 9-18 months of age but in India there is a significant delay in the onset of symptoms (3-4 years) and the age of presentation (6.8-9.2 years). Failure to thrive, anemia, and muscle wasting are more common in children. Symptomatic CD represents the tip of the iceberg. Clinically there may be overt, silent, latent and potential CD. Modified ESPGAN criteria for diagnosis of CD is based on demonstration of characteristic histological changes in small intestinal biopsy while on gluten, and unequivocal clinical improvement when on gluten free diet. However, in developing countries there are several conditions which can give rise to villous atrophy and using it as a diagnostic criteria may lead to overdiagnosis of CD. IgA antigliadin antibody has a high sensitivity (98%) and specificity (90%) for CD and hence, may be an ideal adjunct to modified ESPGAN criteria in the diagnosis of CD in India.

Ontogeny of the small intestine

During development the gastrointestinal tract undergoes marked changes in many physiological and anatomic properties. The remarkable degree of coordination between the development of the gastrointestinal function suggests that the processes may be signalled by some factors, such as weaning, nutrient intake, growth and hormones. The interactions between nutrition and intestinal development begin when fetuses start swallowing amniotic fluid and extend past weaning. Hormonal control plays a major role in the ontogeny of the small intestine. There are late effects of early nutrition, and the normal progress of ontogeny may be important to ensure that the intestine is capable of adaptation in later life.

A microtiter plate assay for superoxide using MTT reduction method.

A simple, rapid and sensitive microtiter plate assay for superoxide using the reduction of tetrazolium dye MTT to its coloured formazan has been developed. The colour formed can be measured using a microtiter plate reader and the extent of reduction of MTT indicates the amount of superoxide generation. A comparison of the sensitivities of different procedures for the quantitation of superoxide generated by X-XO system has been made. The MTT reduction due to superoxide was confirmed by inhibiting the reduction using purified superoxide dismutase. Using this method superoxide generation by mitochondria and microsomes was demonstrated and this procedure is suitable for detection of intracellularly generated superoxide. The proposed method is inexpensive and is suitable for a routine analysis of large number of samples.