Morphological and Immunohistochemical Identification of Villous M Cells in the Small Intestine of Newborn Piglets / Identificación Morfológica e Inmunohistoquímica de los Epiteliocitos Microplegados de las Vellosidades en el Intestino Delgado de Lechones Recién Nacidos

Microfold (M) cells act as antigen-sampling sites for initiating antigen specific mucosal immune responses, but they may also provide a gateway for enteropathogen entry. In this study we demonstrated villous M cells by morphological and immunohistochemical methods to be present in the three regions of the small intestine from newborn piglets. Immunohistochemical analysis, using anti- cytokeratin 18 (CK18) primary antibodies, showed a gradually decreased density of M cells from the lower crypt epithelium to the upper villous epithelium. The proportion of villous M cells was greater in the ileum than in the duodenum or the mid-jejunum. Ultrastructural observation revealed that villous M cells were mainly columnar in shape in the duodenum and the mid-jejunum, and appeared as more pocket-like structure in the ileum. These villous M cells exhibited short and irregular microvilli, rich vesicles and reduced glycocalyx, but lacked the lymphocyte-containing basolateral invagination. Our results support evidence that M cells can develop in the small intestinal villous epithelium of newborn piglets, implying that villous M cells may begin developing in the pig's small intestine during fetal stages, which depends neither on the influence of the mucosal lymphoid tissue nor the antigen from the intestinal lumen stimulation. In addition, the variable morphology and heterogeneity distribution of villous M cells in the three regions of the small intestine may be indicative of its different functional properties. This information extent our understanding of the diversity of M cells and provides important basic knowledge for further research on the actual role of villous M cells in neonate.

Los epiteliocitos microplegados (células M) actúan como receptores de antígeno para iniciar la respuesta inmune específica de las mucosas, pero también pueden proporcionar una puerta de entrada para enteropatógenos. En este estudio, se demostró por métodos morfológicos e inmunohistoquímicos que los epiteliocitos microplegados de las vellosidades están presentes en las tres regiones del intestino delgado de lechones recién nacidos. Se utilizaron anticuerpos primarios de citoqueratina 18 (CK18) para el análisis inmunohistoquímico, el cual mostró una disminución gradual de la densidad de los epiteliocitos microplegados desde el epitelio de las criptas inferiores hasta el epitelio de las vellosidades superiores. La proporción de los epiteliocitos microplegados, fue mayor en el íleon que el duodeno o yeyuno medio. La observación ultraestructural reveló que los epiteliocitos microplegados fueron principalmente de forma columnar en el duodeno y el yeyuno medio. Además, mostraron microvellosidades cortas e irregulares, muchas vesículas y glucocáliz reducidos, pero carecían de invaginaciones basolaterales contenedoras de linfocitos. Nuestros resultados apoyan la evidencia de que los epiteliocitos microplegados pueden desarrollarse en el epitelio de las vellosidades intestinales de los lechones recién nacidos, lo que implica que estas células pueden comenzar a desarrollarse en el intestino delgado del cerdo durante las etapas fetales, y no dependen ni de la influencia del tejido linfoide de las mucosas ni del antígeno para la estimulación del lumen intestinal. Además, la morfología y heterogeneidad de distribución de los epiteliocitos microplegados en las tres regiones del intestino delgado pueden ser indicativas de sus diferentes propiedades funcionales. Esta información mejora nuestra comprensión de la diversidad de los epiteliocitos microplegados y proporciona conocimientos básicos importantes para la investigación sobre el papel de los epiteliocitos microplegados en las vellosidades del neonato.

Evaluation of tee ultrastructure of the small itestine of hiv infected childreen by transmission and scanning electronic microscopy

Objectives To describe HIV children's small intestinal ultrastructural findings. Methods Descriptive, observational study of small intestine biopsies performed between August 1994 and May 1995 at São Paulo, SP, Brazil. This material pertained to 11 HIV infected children and was stored in a laboratory in paraffin blocks. Scanning and transmission electronic microscopy were used to view those intestine samples and ultrastructural findings were described by analyzing digitalized photos of this material. Ethical Committee approval was obtained. Results In most samples scanning microscopy showed various degrees of shortening and decreasing number of microvilli and also completes effacements in some areas. Derangement of the enterocytes was seen frequently and sometimes cells well defined borders limits seemed to be loosened. In some areas a mucous-fibrin like membrane with variable thickness and extension appeared to partially or totally coat the epithelial surface. Fat drops were present in the intestinal lumen in various samples and a bacterium morphologically resembling bacilli was seen in two occasions. Scanning microscopy confirmed transmission microscopy microvilli findings and also showed little “tufts” of those structures. In addition, it showed an increased number of vacuoles and multivesicular bodies inside various enterocytes, an increased presence of intraepithelial lymphocytes, mitochondrial vacuolization and basement membrane enlargement in the majority of samples analyzed. However, some samples exhibited normal aspect. Conclusions Our study showed the common occurrence of various important intestinal ultrastructural alterations with variable degrees among HIV infected children, some of them in our knowledge not described before. .

Objetivos Descrever achados ultra-estruturais do intestino delgado de crianças infectadas pelo HIV. Métodos Estudo descritivo, observacional de biopsias do intestino delgado, realizada entre agosto de 1994 e maio de 1995 em São Paulo - Brasil. Este material pertencia a 11 crianças infectadas pelo HIV e foi armazenado em um laboratório em blocos de parafina. As amostras de intestino delgado foram analisadas por microscopia eletrônica de transmissão e de varredura e achados os achados ultra-estruturais foram descritos por meio da análise de fotos digitalizadas desse material. Foi obtida aprovação pelo Comitê de Ética. Resultados Na maioria das amostras a microscopia de varredura mostrou vários graus de encurtamento e diminuição do número das microvilosidades e até o completo apagamento dessas estruturas em algumas áreas. O desarranjo dos enterócitos foi visto com freqüência e, por vezes, os limites celulares estavam imprecisos. Em algumas áreas uma membrana fibrino-mucosa com espessura e extensão variáveis aparentava revestir parcial ou totalmente a superfície epitelial. Gotas de gordura no lúmen intestinal estavam presentes em várias amostras e bactérias morfologicamente semelhantes a bacilos foram observadas em duas amostras. A microscopia eletrônica de varredura confirmou as observações constatadas nas microvilosidades através da microscopia de transmissão e também mostrou pequenos “tufos” dessas estruturas. Além disso, mostrou aumento do número de vacúolos e de formações multivesiculares dentro de vários enterócitos, aumento da presença de linfócitos intraepiteliais, vacuolização mitocondrial e alargamento da membrana basal na maioria das amostras analisadas. ...

Pre- and postnatal development of rabbit small intestine; a light and scanning electron microscopic study

The small intestine is a highly differentiated organ that accomplishes both digestive and absorptive functions with great efficiency. The small intestine of rabbit was studied from early prenatal stage to full term in 15 fetuses and during the early days of life till maturity in 21 rabbits using light and scanning electron microscopy. In 15 days old fetuses, the epithelium was flat and stratified and the lumen was elliptical in shape. By the E18, the epithelial ridges were formed. The first rudimentary villus formation was formed in 21 days old fetuses and the true villi appeared by 24[th] days of gestation. At 27 days old fetuses, intestinal glands were not appeared yet. By the first day of postnatal life the duodenal glands appeared. The histological maturity of the rabbit small intestine occurred one month after birth. In conclusion, at all stages, the sequential morphologic changes of the rabbit small intestine developed to meet the structural and physiological demands during the fetal stage to be prepared to extra uterine life

Experiencia en el uso de la cápsula endoscópica de intestino delgado en pacientes con sangrado gastrointestinal oscuro / Experience in use of capsule endoscopica of thin intestine in patients with bled dark gastrointestinal

La cápsula endoscópica es un método diagnóstico que evalúa el tracto gastrointestinal de forma no invasiva. La principal indicación del estudio es la determinación del origen del sangrado gastrointestinal oscuro. Dicha entidad clínica se define, como la hemorragia digestiva de causa desconocida, persistente y recurrente, después de una prueba negativa en el estudio endoscópico convencional. (Colonoscopia – endoscopia digestiva superior). Objetivo: Describir hallazgos de la cápsula endoscópica en pacientes con hemorragias digestivas de origen oscuro. Pacientes y Métodos: Estudio descriptivo de corte trasversal. Se evaluaron 19 pacientes con hemorragia digestiva de origen oscuro que acudieron al Servicio de Gastroenterología del centro de cuarto nivel durante el periodo de marzo 2009 – marzo 2010, excluyéndose a pacientes en estado crítico o terminal, y aquellos con clínica de obstrucción intestinal o fistulas. Utilizando plataforma Given Imaging® SB2. Resultados: Se estudiaron 19 pacientes, 14 (73,68%) sexo masculino, 5 (26,31%) sexo femenino, grupo etáreo comprendido entre 29-60 años edad promedio 39.5 años. Indicación: sangrado digestivo oculto: 6 casos (31.57%). Sangrado digestivo oscuro: 13 caso (68.42 %). Hallazgos en 16 pacientes (84.21%): sangrado activo profuso localizado de causa indeterminada (6.25%), úlceras y erosiones (37.5%), lesión de aspecto tumoral sangrante activo (12,5%), angiodisplasia (37.5%), pólipos (6.25%). Tres casos (15,78%) con evaluación limitada por mala preparación. Conclusión: La cápsula endoscópica se considera en la actualidad una herramienta útil para la detección de lesiones en intestino delgado, siendo la principal indicación en la hemorragia gastrointestinal de origen oscuro. Con un rendimiento diagnóstico de 84% en la evaluación de la Hemorragias Digestiva oscura oculta.

The endoscópica capsule is a method diagnosis that evaluates tracto gastrointestinal of noninvasive form. The main indication of the study is the determination of the origin of bled gastrointestinal dark. This clinical organization is defined, like the digestive hemorrhage of cause unknown, persistent and recurrent, after one proves refusal in the conventional endoscópico study. (Colonoscopia - superior digestive endoscopia). Objective: To describe findings of the endoscópica capsule in patients with digestive hemorrhages of dark origin. Patients and Methods: Descriptive study of trasversal cut. 19 patients with digestive hemorrhage of dark origin evaluated themselves that they went to the Service of Gastroenterology of the center of fourth level during the period of March 2009 - March 2010, excluding itself patients in critical or terminal state, and those with intestinal clinic of obstruction or fistulas. Using platform Given Imaging® SB2. Results: 19 patients studied, 14 (73.68%) masculine sex, 5 (26.31%) feminine sex, etáreo group included/understood between 29-60 years age average 39,5 years. Indication: bled digestive hidden: 6 cases (31,57%). Bled digestive dark: 13 case (68,42%). Findings in 16 patients (84,21%): bled active profuse located of indetermine cause (6,25%), ulcers and erosions (37,5%), injury of tumorlike aspect bleeding assets (12.5%), angiodisplasia (37,5%), polyps (6,25%). Three cases (15.78%) with evaluation limited by bad preparation. Conclusion: The endoscópica capsule considers at the present time a useful tool for the detection of injuries in thin intestine, being the main indication in the gastrointestinal he-morrhage of dark origin. With a yield diagnosis of 84% in the evaluation of the Hemorrhages Digestive dark hidden.

role of probiotics in controlling giardia intestinalis in experimental animals

Efficacy of Lactobacillus acidophilus [Lactospore] for the control Research Institute Warak of Giardia intestianlis infection was evaluated in hamsters. Each El-Hadar Imbaba hamster was infected orally by 10.000 Giardia lamblia cysts. Animal were divided into five groups: Group A: control infected, untreate group. Group B: infected receiving Metronidazole. Group C: infected and receiving Lactospore. Group D: infected and receiving combination of Metronidazole and Lactospore. Group E: receiving Lactospore 7 days before infection acting as a prophylactic group. Groups B, C and D were given the appmpriate drug, three weeks post infection. Two weeks later stool analysis was performed and cysts/gm stool were counted, after which scarification of all groups took place. There was a highly significant difference between control and all treated groups. The highest percentage of reduction [cure rate] was in group D [98.6%]. followed by group B which gave a reduction rate of [93.8 1%]. The effect of the drugs on the vegetative [trophozoite] forms in the small intestine of sacrificed hamsters was studied. Combination of both drugs [group D] revealed a high significant cure rate [99.32%]. On the other hand when metronidazole and lactospore were given alone a reduction rate of 92.22% and 63.4% respectively was observed. The prophylaxis effect of Lactospore was highly noticed with 93.65% parasite reduction. Assessment of cure was also performed by electron microscopic and histopathological examination of the small intestine, peyer's patches and the spleen. Ultrastructural examination of the small intestine revealed remarkable destruction of the intestinal cell projection by Gardia cyst. Partial healing of the destructeci intestinal cell projection by Metronidazole was obvious, while complete healing could be detected in both groups D and E, Combined treatment with metronidazole and lactospore revealed remarkable activation of lymphocytes and macrophages. This easy flow through the endothelial cells lining the sinus in its way to the lumen to enhance their opportunity to overcome the infection could be detected. Histopathological examination revealed complete healing of the intestinal mucosa after the combined treatment while partial healing of the lining epithelium of the intestine was noticed after metronidazole treatment. The present study proves the efficacy of lactospore as a prophylactic agent for Giardiasis when given 7 days pre infection. This might be of considerable interest in eases of travelers diarrhea. In addition, it can strengthen the effect of metronidazole when both drugs are given together

Studies of the small bowel surface by scanning electron microscopy in infants with persistent diarrhea

We describe the ultrastructural abnormalities of the small bowel surface in 16 infants with persistent diarrhea. The age range of the patients was 2 to 10 months, mean 4.8 months. All patients had diarrhea lasting 14 or more days. Bacterial overgrowth of the colonic microflora in the jejunal secretion, at concentrations above 10(4) colonies/ml, was present in 11 (68.7 percent) patients. The stool culture was positive for an enteropathogenic agent in 8 (50.0 percent) patients: for EPEC O111 in 2, EPEC O119 in 1, EAEC in 1, and Shigella flexneri in 1; mixed infections due to EPEC O111 and EAEC in 1 patient, EPEC O119 and EAEC in 1 and EPEC O55, EPEC O111, EAEC and Shigella sonnei in 1. Morphological abnormalities in the small bowel mucosa were observed in all 16 patients, varying in intensity from moderate 9 (56.3 percent) to severe 7 (43.7 percent). The scanning electron microscopic study of small bowel biopsies from these subjects showed several surface abnormalities. At low magnification (100X) most of the villi showed mild to moderate stunting, but on several occasions there was subtotal villus atrophy. At higher magnification (7,500X) photomicrographs showed derangement of the enterocytes; on several occasions the cell borders were not clearly defined and very often microvilli were decreased in number and height; in some areas there was a total disappearance of the microvilli. In half of the patients a mucus-fibrinoid pseudomembrane was seen partially coating the enterocytes, a finding that provides additional information on the pathophysiology of persistent diarrhea

Microscopia eletrônica de varredura do intestino delgado na diarréia persistente / Scanning electronic microscopy of the small intestine in the persistent diarrhea

Foram estudadas 16 crianças menores de um ano, de ambos os sexos, hospitalizadas por diarréia persistente, com o objetivo de analisar a estrutura do epitélio do intestino delgado através das microscopias de luz e eletrônica de varredura. Foram pesquisados enteropatógenos nas fezes e no suco jejunal. Os pacientes foram submetidos a intubaçäo naso-jejunal com cápsula de Watson para obtençäo de secreçäo jejunal e biopsia de intestino delgado. Os fragmentos de duodeno foram fixados em formol 10 por cento para microscopia de luz, em cortes semi-finos, e em soluçäo de Karnowsky modificada para a varredura, sendo pós-incluídos em tetróxido de ósmio 1 por cento, desidratados numa série gradual de etanol, secados pelo método do ponto crítico, com CO2, cobertos com prata, e analisados através de um JEOL JSM 5300 Scanning Microscope. As coproculturas foram positivas em 11 (68,7 por cento) pacientes: EPEC 0111 em quatro; EPEC 0119 em um; EAggEC em cinco; Shigella flexneri em dois e Shigella sonnei em um; infecçöes mistas por EAggEC, associadas a EPEC 0111 foram caracterizadas em dois pacientes. A microscopia de luz, 56,2 por cento dos pacientes apresentaram enteropatia grau II e, em todos, foram observadas alteraçöes epiteliais como esfacelamento de microvilosidades, vacuolizaçäo citoplasmática, corpos muultivesiculares, e infiltrado linfocitário e eosinofílico, este inclusive no córion. A microscopia eletrónica de varredura, observaram-se, em todos os casos, vilosidades e células epiteliais achatadas e disformes, com ausência ou diminuiçäo das microvilosidades, e em metade dos casos a presença de um material mucoso recobrindo parte do epitélio. A aderência firme entre este material e o epitélio deu-lhe o aspecto de uma psedomembrana. Observou-se um infiltrado de linfócitos sobre a superfície apical dos enterócitos de uma espécime, ao lado de gotículas de gordura. Concluiu-se que as alteraçöes encontradas podem ser devidas a uma associaçäo genérica entre o agente enteropatogênico deflagador da diarréia e o surgimento de intolerância alimentar que a perpetua.

Sorotipos de Escherichia coli enteropatogênica clássica (EPEC) causadores de alteraçöes da ultra-estrutura do intestino delgado e invasäo de células Hela / Serotypes of classical enteropathogic that cause changes in the small bowel ultrastructure and invasion of Hela cells

Escherichia coli enteropatogênica clássica (EPEC) é a principal causa de diarréia em lactentes menores de um ano de idade nos países subdesenvolvidos. In vitro, cepas de EPEC sao capazes de aderir a células HeLa ou Hep-2, obedecendo um padrao característico denominado adesao localizada. Estudos in vivo demonstram que as cepas de EPEC provocam profundas alteraçoes nas micro-vilosidades do enterócito, causando lesoes em pedestal. Objetivos. Neste trabalho, os autores relatam as alteraçoes da ultra-estrutura do intestino delgado causada pelos sorotipos 0111ab:H2, 0119:H6 e 018ab:H14, em lactentes portadores de diarréia aguda, e a capacidade destas bactérias de penetrar e se multiplicar no interior de células HeLa. Pacientes e Métodos. Estes sorotipos de EPEC foram isolados das fezes de três pacientes portadores de diarréia aguda que apresentaram intolerância alimentar e agravo nutricional. Alteraçoes ultra-estruturais do intestino delagado foram observadas nos espécimes estudados com formaçao de pedestal e penetraçao das bactérias no interior do citoplasma dos enterócitos. Estas bactérias também penetraram e se multiplicaram no interior das células HeLa. Conclusao. As alteraçoes ultra-estruturaisdescritas em pacientes com diarréria aguda devem ser levadas em conta quanto ao seu potencial efeito de provocar prolongamento da diarréia, seja por um fenômeno secretor ou por má absorçao dos nutrientes.

Ultrastructural study of enteropathogenic Escherichia coli Olllab: H2 infection in an infant with acute diarrhea

Escherichia coli enteropatogênica clássica é o mais importante agente etiológico como causa de diarréia aguda nos países subdesenvolvidos, especialmente nos lactentes menores de uma ano de idade. As cepas de Escherichia coli enteropatogênica sao capazes de provocar profundas alteraçoes no citoesqueleto do enterócito, as quais sao denominadas aderência e apagamento das microvilosidades do enterócito, as quais sao denominadas aderência e apagamento das microvilosidades, associadas à formaçao de pedestais. Descreve-se uma cepa de Escherichia coli enteropatogênica 0111ab:H2 isolada das fezes de um lactente com diarréia aguda, no 11§ dia de doença, a qual provocou lesoes de aderência e apagamento das microvilosidades do enterócito e inclusive penetrou no interior do citoplasma do mesmo, assim como invadiu células HeLa em cultura de tecido in vitro e da mesma forma no teste da alça ileal ligada de coelho in vivo, todas as lesoes detectadas em estudo ultra-estrutural. Esta observaçao demonstra que lesoes graves da mucosa do intestino delgado causadas por uma cepa de Escherichia coli enteropatogênica 0111ab:H2 podem ocorrer ainda em um estágio precoce da infecçao entérica.

Transit time, bacterial growth and electron microscope study of the small intestine in schistosomal hepatic fibrosis with portal pressure chances

Transit time, bacterial growth together with both light and electron microscopy of small intestine were studied in relation to different portal pressure levels in 23 patients with schistosomal hepatic fibrosis Schistosomal patients were divided into two groups: the first included those without ascites or elevated portal pressure [100-150 mm H[2]0] while the second group comprised patients with ascites and a portal pressure varying between 180 and 400 mm H[2]0. Although all findings were within normal in the first group patients, yet those of the second group showed delayed intestinal transit time[80%], small bowel bacterial overgrowth whether aerobic or anaerobic [80%] along with mild, moderate or marked light and electron microscopic pathological changes which seemed to depend on the extent of rise of the portal pressure. In spite of the fact that all studied parameters seem to be of importance, yet further attempts should be made in order to clarify the role and sequence of event, each of these factors play, in induction of altered intestinal functions in hepatic schistosomiasis patients