Gastric and small intestinal lesions after partial stomach resection with Billroth II or Roux-en-Y reconstruction in the rat

The evolution and phenotypic expression of mucosal lesions of the gastric stump were investigated in male rats submitted to gastric resection with reconstruction by the Billroth II technique (BII with biliopancreatic reflux, BPR) or by the Roux-en-Y procedure (without BPR). Animals were studied at 24, 36, 54 and 64 weeks after surgery and the phenotypic expression of lesions analysed using routine hematoxylin and eosin staining, immunohistochemical staining for pepsinogen isoenzyme 1 and histochemical procedures for mucins (paradoxical concanavalin A, galactose oxidase Schiff (GOS) and sialidase GOS reactions). BPR was found to be responsible for the formation of adenomatous hyperplasia (AH), increasing in incidence and size with time, since the Roux-en-Y procedure failed to induce the gastric stump lesions observed after BII reconstruction. AHs always occurred in the transition of the gastrojejunal junction, a site offering special conditions for BPR influence, and were classified as gastric (G), intestinal (I) and G+I types according to their phenotypic expression. No pure I type AH was diagnosed at any time point. The G and G+I types developed at approximately equal incidences (i.e., G type 7/17, G+I type 10/17 at the 64th week). It was suggested that both gastric and intestinal mucosal elements were stimulated to proliferate by BPR, with the gastric mucosa tending to demonstrate AH. Intestinal type components of AH were found adjacent to the jejunum and not at the stomach margin, indicating an origin from intestinal mucosa. No metaplasia of the gastric mucosa was observed in any animal after partial gastric resection. In 101 rats submitted to the BII procedure, 5 mucinous adenocarcinomas were eventually diagnosed, mostly located in the subserosa of the gastrojejunal junction. All carcinomas expressed the phenotype of cells of the small intestine. Evidence of malignant transformation within the gastric components of AH was not observed even at the 64th week. In conclusion, all lesions induced by BPR in the rat remnant stomach are benign, and the few true cancers that arise in association are derived from the small intestine.

Immunomodulatory activity and control of Salmonella Enteritidis colonization in the intestinal tract of chickens by Lactobacillus based probiotic

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Acute heat stress impairs performance parameters and induces mild intestinal enteritis in broiler chickens: Role of acute hypothalamic-pituitary-adrenal axis activation

Studies on the environmental consequences of stress are relevant for economic and animal welfare reasons. We recently reported that long-term heat stressors (31 +/- 1 degrees C and 36 +/- 1 degrees C for 10 h/d) applied to broiler chickens (Gallus gallus domesticus) from d 35 to 42 of life increased serum corticosterone concentrations, decreased performance variables and the macrophage oxidative burst, and produced mild, multifocal acute enteritis. Being cognizant of the relevance of acute heat stress on tropical and subtropical poultry production, we designed the current experiment to analyze, from a neuroimmune perspective, the effects of an acute heat stress (31 +/- 1 degrees C for 10 h on d 35 of life) on serum corticosterone, performance variables, intestinal histology, and peritoneal macrophage activity in chickens. We demonstrated that the acute heat stress increased serum corticosterone concentrations and mortality and decreased food intake, BW gain, and feed conversion (P < 0.05). We did not find changes in the relative weights of the spleen, thymus, and bursa of Fabricius (P > 0.05). Increases in the basal and the Staphylococcus aureus-induced macrophage oxidative bursts and a decrease in the percentage of macrophages performing phagocytosis were also observed. Finally, mild, multifocal acute enteritis, characterized by the increased presence of lymphocytes and plasmocytes within the lamina propria of the jejunum, was also observed. We found that the stress-induced hypothalamic-pituitary-adrenal axis activation was responsible for the negative effects observed on chicken performance and immune function as well as for the changes in the intestinal mucosa. The data presented here corroborate with those presented in other studies in the field of neuroimmunomodulation and open new avenues for the improvement of broiler chicken welfare and production performance.

Study of apoptotic deletion mediated by Bifidobacterium longum with construction of recombinant strains for Serpin encoding gene and phenotypes comparison in a pig cell model

The first part of the research project of the Co-Advisorship Ph.D Thesis was aimed to select the best Bifidobacterium longum strains suitable to set the basis of our study. We were looking for strains with the abilities to colonize the intestinal mucosa and with good adhesion capacities, so that we can test these strains to investigate their ability to induce apoptosis in “damaged” intestinal cells. Adhesion and apoptosis are the two process that we want to study to better understand the role of an adhesion protein that we have previously identified and that have top scores homologies with the recent serpin encoding gene identified in B. longum by Nestlè researchers. Bifidobacterium longum is a probiotic, known for its beneficial effects to the human gut and even for its immunomodulatory and antitumor activities. Recently, many studies have stressed out the intimate relation between probiotic bacteria and the GIT mucosa and their influence on human cellular homeostasis. We focused on the apoptotic deletion of cancer cells induced by B. longum. This has been valued in vitro, performing the incubation of three B.longum strains with enterocyte-like Caco- 2 cells, to evidence DNA fragmentation, a cornerstone of apoptosis. The three strains tested were defined for their adhesion properties using adhesion and autoaggregation assays. These features are considered necessary to select a probiotic strain. The three strains named B12, B18 and B2990 resulted respectively: “strong adherent”, “adherent” and “non adherent”. Then, bacteria were incubated with Caco-2 cells to investigate apoptotic deletion. Cocultures of Caco-2 cells with B. longum resulted positive in DNA fragmentation test, only when adherent strains were used (B12 and B18). These results indicate that the interaction with adherent B. longum can induce apoptotic deletion of Caco-2 cells, suggesting a role in cellular homeostasis of the gastrointestinal tract and in restoring the ecology of damaged colon tissues. These results were used to keep on researching and the strains tested were used as recipient of recombinant techniques aimed to originate new B.longum strains with enhanced capacity of apoptotic induction in “damaged” intestinal cells. To achieve this new goal it was decided to clone the serpin encoding gene of B. longum, so that we can understand its role in adhesion and apoptosis induction. Bifidobacterium longum has immunostimulant activity that in vitro can lead to apoptotic response of Caco-2 cell line. It secretes a hypothetical eukaryotic type serpin protein, which could be involved in this kind of deletion of damaged cells. We had previously characterised a protein that has homologies with the hypothetical serpin of B. longum (DD087853). In order to create Bifidobacterium serpin transformants, a B. longum cosmid library was screened with a PCR protocol using specific primers for serpin gene. After fragment extraction, the insert named S1 was sub-cloned into pRM2, an Escherichia coli - Bifidobacterium shuttle vector, to construct pRM3. Several protocols for B. longum transformation were performed and the best efficiency was obtained using MRS medium and raffinose. Finally bacterial cell supernatants were tested in a dotblot assay to detect antigens presence against anti-antitrypsin polyclonal antibody. The best signal was produced by one starin that has been renamed B. longum BLKS 7. Our research study was aimed to generate transformants able to over express serpin encoding gene, so that we can have the tools for a further study on bacterial apoptotic induction of Caco-2 cell line. After that we have originated new trasformants the next step to do was to test transformants abilities when exposed to an intestinal cell model. In fact, this part of the project was achieved in the Department of Biochemistry of the Medical Faculty of the University of Maribor, guest of the abroad supervisor of the Co-Advisorship Doctoral Thesis: Prof. Avrelija Cencic. In this study we examined the probiotic ability of some bacterial strains using intestinal cells from a 6 years old pig. The use of intestinal mammalian cells is essential to study this symbiosis and a functional cell model mimics a polarised epithelium in which enterocytes are separated by tight junctions. In this list of strains we have included the Bifidobacterium longum BKS7 transformant strain that we have previously originated; in order to compare its abilities. B. longum B12 wild type and B. longum BKS7 transformant and eight Lactobacillus strains of different sources were co-cultured with porcine small intestine epithelial cells (PSI C1) and porcine blood monocytes (PoM2) in Transwell filter inserts. The strains, including Lb. gasseri, Lb. fermentum, Lb. reuterii, Lb. plantarum and unidentified Lactobacillus from kenyan maasai milk and tanzanian coffee, were assayed for activation of cell lines, measuring nitric oxide by Griess reaction, H202 by tetramethylbenzidine reaction and O2 - by cytochrome C reduction. Cytotoxic effect by crystal violet staining and induction on metabolic activity by MTT cell proliferation assay were tested too. Transepithelial electrical resistance (TER) of polarised PSI C1 was measured during 48 hours co-culture. TER, used to observe epithelium permeability, decrease during pathogenesis and tissue becomes permeable to ion passive flow lowering epithelial barrier function. Probiotics can prevent or restore increased permeability. Lastly, dot-blot was achieved against Interleukin-6 of treated cells supernatants. The metabolic activity of PoM2 and PSI C1 increased slightly after co-culture not affecting mitochondrial functions. No strain was cytotoxic over PSI C1 and PoM2 and no cell activation was observed, as measured by the release of NO2, H202 and O2 - by PoM2 and PSI C1. During coculture TER of polarised PSI C1 was two-fold higher comparing with constant TER (~3000 ) of untreated cells. TER raise generated by bacteria maintains a low permeability of the epithelium. During treatment Interleukin-6 was detected in cell supernatants at several time points, confirming immunostimulant activity. All results were obtained using Lactobacillus paracasei Shirota e Carnobacterium divergens as controls. In conclusion we can state that both the list of putative probiotic bacteria and our new transformant strain of B. longum are not harmful when exposed to intestinal cells and could be selected as probiotics, because can strengthen epithelial barrier function and stimulate nonspecific immunity of intestinal cells on a pig cell model. Indeed, we have found out that none of the strains tested that have good adhesion abilities presents citotoxicity to the intestinal cells and that non of the strains tested can induce cell lines to produce high level of ROS, neither NO2. Moreover we have assayed even the capacity of producing certain citokynes that are correlated with immune response. The detection of Interleukin-6 was assayed in all our samples, including B.longum transformant BKS 7 strain, this result indicates that these bacteria can induce a non specific immune response in the intestinal cells. In fact, when we assayed the presence of Interferon-gamma in cells supernatant after bacterial exposure, we have no positive signals, that means that there is no activation of a specific immune response, thus confirming that these bacteria are not recognize as pathogen by the intestinal cells and are certainly not harmful for intestinal cells. The most important result is the measure of Trans Epithelial Electric Resistance that have shown how the intestinal barrier function get strengthen when cells are exposed to bacteria, due to a reduction of the epithelium permeability. We have now a new strain of B. longum that will be used for further studies above the mechanism of apoptotic induction to “damaged cells” and above the process of “restoring ecology”. This strain will be the basis to originate new transformant strains for Serpin encoding gene that must have better performance and shall be used one day even in clinical cases as in “gene therapy” for cancer treatment and prevention.

TNF suppresses acute intestinal inflammation by inducing local glucocorticoid synthesis

Although tumor necrosis factor (alpha) (TNF) exerts proinflammatory activities in a variety of diseases, including inflammatory bowel disease, there is increasing evidence for antiinflammatory actions of TNF. In contrast, glucocorticoids (GCs) are steroid hormones that suppress inflammation, at least in part by regulating the expression and action of TNF. We report that TNF induces extraadrenal production of immunoregulatory GCs in the intestinal mucosa during acute intestinal inflammation. The absence of TNF results in a lack of colonic GC synthesis and exacerbation of dextran sodium sulfate-induced colitis. TNF seems to promote local steroidogenesis by directly inducing steroidogenic enzymes in intestinal epithelial cells. Therapeutic administration of TNF induces GC synthesis in oxazolone-induced colitis and ameliorates intestinal inflammation, whereas inhibition of intestinal GC synthesis abrogates the therapeutic effect of TNF. These data show that TNF suppresses the pathogenesis of acute intestinal inflammation by promoting local steroidogenesis.

Galectin-3 modulates T cell activity and is reduced in the inflamed intestinal epithelium in IBD

BACKGROUND: Galectins are involved at different stages in inflammation. Galectin-3, although mostly described as proinflammatory, can also act as an immunomodulator by inducing apoptosis in T cells. The present study aims to determine galectin-3 expression in the normal and inflamed intestinal mucosa and to define its role in T cell activity. MATERIALS AND METHODS: Galectin-3 was detected by quantitative polymerase chain reaction with total RNA from endoscopic biopsies and by immunohistochemistry. Biopsies and peripheral blood mononuclear cells (PBMC) were stimulated in vitro and were used to assess the functional consequences of inhibition or exogenous addition of galectin-3. RESULTS: Galectin-3 is expressed at comparable levels in controls and inflammatory bowel disease (IBD) patients in remission. In the normal mucosa, galectin-3 protein was mainly observed in differentiated enterocytes, preferentially at the basolateral side. However, galectin-3 was significantly downregulated in inflamed biopsies from IBD patients. Ex vivo stimulation of uninflamed biopsies with tumor necrosis factor led to similar galectin-3 messenger RNA downregulation as in vivo. When peripheral blood mononuclear cells (PBMC) were analyzed, galectin-3 was mainly produced by monocytes. Upon mitogen stimulation, we observed increased proliferation and decreased activation-induced cell death of peripheral blood T cells in the presence of galectin-3-specific small interfering RNA. In contrast, exogenous addition of recombinant galectin-3 led to reduced proliferation of mitogen-stimulated peripheral blood T cells. CONCLUSIONS: Our results suggest that downregulation of epithelial galectin-3 in the inflamed mucosa reflects a normal immunological consequence, whereas under noninflammatory conditions, its constitutive expression may help to prevent inappropriate immune responses against commensal bacteria or food compounds. Therefore, galectin-3 may prove valuable for manipulating disease activity.

Continuous thoracic epidural anesthesia improves gut mucosal microcirculation in rats with sepsis

Microcirculatory dysfunction contributes significantly to tissue hypoxia and multiple organ failure in sepsis. Ischemia of the gut and intestinal hypoxia are especially relevant for the evolution of sepsis because the mucosal barrier function may be impaired, leading to translocation of bacteria and toxins. Because sympathetic blockade enhances intestinal perfusion under physiologic conditions, we hypothesized that thoracic epidural anesthesia (TEA) may attenuate microcirculatory perturbations during sepsis. The present study was designed as a prospective and controlled laboratory experiment to assess the effects of continuous TEA on the mucosal microcirculation in a cecal ligation and perforation model of sepsis in rats. Anesthetized Sprague-Dawley rats underwent laparotomy and cecal ligation and perforation to induce sepsis. Subsequently, either bupivacaine 0.125% (n = 10) or isotonic sodium chloride solution (n = 9) was continuously infused via the thoracic epidural catheter for 24 h. In addition, a sham laparotomy was carried out in eight animals. Intravital videomicroscopy was then performed on six to ten villi of ileum mucosa. The capillary density was measured as areas encircled by perfused capillaries, that is, intercapillary areas. The TEA accomplished recruitment of microcirculatory units in the intestinal mucosa by decreasing total intercapillary areas (1,317 +/- 403 vs. 1,001 +/- 236 microm2) and continuously perfused intercapillary areas (1,937 +/- 512 vs. 1,311 +/- 678 microm2, each P < 0.05). Notably, TEA did not impair systemic hemodynamic variables beyond the changes caused by sepsis itself. Therefore, sympathetic blockade may represent a therapeutic option to treat impaired microcirculation in the gut mucosa resulting from sepsis. Additional studies are warranted to assess the microcirculatory effects of sympathetic blockade on other splanchnic organs in systemic inflammation.

Intestinal macrophages: differentiation and involvement in intestinal immunopathologies

Intestinal macrophages, preferentially located in the subepithelial lamina propria, represent the largest pool of tissue macrophages in humans. As an adaptation to the local antigen- and bacteria-rich environment, intestinal macrophages exhibit several distinct phenotypic and functional characteristics. Notably, microbe-associated molecular pattern receptors, including the lipopolysaccharide (LPS) receptors CD14 and TLR4, and also the Fc receptors for IgA and IgG are absent on most intestinal macrophages under homeostatic conditions. Moreover, while macrophages in the intestinal mucosa are refractory to the induction of proinflammatory cytokine secretion, they still display potent phagocytic activity. These adaptations allow intestinal macrophages to comply with their main task, i.e., the efficient removal of microbes while maintaining local tissue homeostasis. In this paper, we review recent findings on the functional differentiation of monocyte subsets into distinct macrophage populations and on the phenotypic and functional adaptations that have evolved in intestinal macrophages in response to their antigen-rich environment. Furthermore, the involvement of intestinal macrophages in the pathogenesis of celiac disease and inflammatory bowel diseases is discussed.

Mycobacterium avium Subsp. avium infection in four veal calves: differentiation from intestinal tuberculosis

Mycobacterium avium subsp. avium (Maa) is an intracellular pathogen belonging to the Mycobacterium avium-intracellulare complex (MAC). Reservoirs of MAC are the natural environment, wildlife and domestic animals. In adult bovine, MAC infections are typically caused by Mycobacterium avium subsp. paratuberculosis (Map). Maa infections in bovine are rarely reported but may cause clinical disease and pathological lesions similar to those observed in paratuberculosis or those induced by members of the Mycobacterium tuberculosis complex (MTBC). Therefore, differentiation of MAC from MTBC infection should be attempted, especially if unusual mycobacterial lesions are encountered. Four veal calves from a fattening farm dying with clinical signs of otitis media, fever, and weight loss were submitted for necropsy. Samples from affected organs were taken for histologic investigation, bacteriologic culture, and bacterial specification using PCR. Macroscopic thickening of the intestinal mucosa was induced by granulomatous enteritis and colitis. Intracytoplasmic acid-fast bacteria were detected by Ziehl-Neelsen stains and PCR revealed positive results for Mycobacterium avium subsp. avium. Clinical and pathological changes of Maa infection in veal calves had features of Mycobacterium avium subsp. paratuberculosis and the MTBC. Therefore, Mycobacterium tuberculosis complex infection should be considered in cases of granulomatous enteritis in calves.

CD4 T cells are required for both development and maintenance of disease in a new mouse model of reversible colitis

Current therapies to treat inflammatory bowel diseases have limited efficacy, significant side effects, and often wane over time. Little is known about the cellular and molecular mechanisms operative in the process of mucosal healing from colitis. To study such events, we developed a new model of reversible colitis in which adoptive transfer of CD4(+)CD45RB(hi) T cells into Helicobacter typhlonius-colonized lymphopenic mice resulted in a rapid onset of colonic inflammation that was reversible through depletion of colitogenic T cells. Remission was associated with an improved clinical and histopathological score, reduced immune cell infiltration to the intestinal mucosa, altered intestinal gene expression profiles, regeneration of the colonic mucus layer, and the restoration of epithelial barrier integrity. Notably, colitogenic T cells were not only critical for induction of colitis but also for maintenance of disease. Depletion of colitogenic T cells resulted in a rapid drop in tumor necrosis factor α (TNFα) levels associated with reduced infiltration of inflammatory immune cells to sites of inflammation. Although neutralization of TNFα prevented the onset of colitis, anti-TNFα treatment of mice with established disease failed to resolve colonic inflammation. Collectively, this new model of reversible colitis provides an important research tool to study the dynamics of mucosal healing in chronic intestinal remitting-relapsing disorders.Mucosal Immunology advance online publication 16 September 2015; doi:10.1038/mi.2015.93.

Transgenic mice that overexpress the human trefoil peptide pS2 have an increased resistance to intestinal damage.

pS2 is a member of the trefoil peptide family, all of which are overexpressed at sites of gastrointestinal injury. We hypothesized that they are important in stimulating mucosal repair. To test this idea, we have produced a transgenic mice strain that expresses human pS2 (hpS2) specifically within the jejunum and examined the effect of this overexpression on proliferation and susceptibility to indomethacin-induced damage. A transgenic mouse was produced by microinjecting fertilized oocytes with a 1.7-kb construct consisting of rat intestinal fatty acid binding protein promoter (positions -1178 to +28) linked to full-length (490 bp) hpS2 cDNA. Screening for positive animals was by Southern blot analysis. Distribution of hpS2 expression was determined by using Northern and Western blot analyses and immunohistochemical staining. Proliferation of the intestinal mucosa was determined by assessing the crypt cell production rate. Differences in susceptibility to intestinal damage were analyzed in animals that had received indomethacin (85 mg/kg s.c.) 0-30 h previously. Expression of hpS2 was limited to the enterocytes of the villi within the jejunum. In the nondamaged intestine, villus height and crypt cell production rate were similar in transgenic and negative (control) litter mates. However, there was a marked difference in the amount of damage caused by indomethacin in control and transgenic animals in the jejunum (30% reduction in villus height in controls vs. 12% reduction in transgenic animals, P < 0.01) but the damage sustained in the non-hpS2-expressing ileal region was similar in control and transgenic animals. These studies support the hypothesis that trefoil peptides are important in stimulating gastrointestinal repair.

Loss of Apc heterozygosity and abnormal tissue building in nascent intestinal polyps in mice carrying a truncated Apc gene.

Mutations in the APC (adenomatous polyposis coli) gene appear to be responsible for not only familial adenomatous polyposis but also many sporadic cases of gastrointestinal cancers. Using homologous recombination in mouse embryonic stem cells, we constructed mice that contained a mutant gene encoding a product truncated at a 716 (Apc delta 716). Mendelian transmission of the gene caused most homozygous mice to die in utero before day 8 of gestation. The heterozygotes developed multiple polyps throughout the intestinal tract, mostly in the small intestine. The earliest polyps arose multifocally during the third week after birth, and new polyps continued to appear thereafter. Surprisingly, every nascent polyp consisted of a microadenoma covered with a layer of the normal villous epithelium. These microadenomas originated from single crypts by forming abnormal outpockets into the inner (lacteal) side of the neighboring villi. We carefully dissected such microadenomas from nascent polyps by peeling off the normal epithelium and determined their genotype by PCR: all microadenomas had already lost the wild-type Apc allele, whereas the mutant allele remained unchanged. These results indicate that loss of heterozygosity followed by formation of intravillous microadenomas is responsible for polyposis in Apc delta 716 intestinal mucosa. It is therefore unlikely that the truncated product interacts directly with the wild-type protein and causes the microadenomas by a dominant negative mechanism.

Avaliação da permeabilidade de fármacos antirretrovirais por meio de modelo ex vivo com emprego de segmentos da mucosa bucal de suínos

A via de administração oral é a forma favorita de administração de fármacos em função das vantagens que apresenta, dentre elas destacam-se: a adesão do paciente, conveniência e praticidade. Em função disto, a maioria dos medicamentos comercializados encontra-se disponível na forma farmacêutica de administração oral, entretanto, o sucesso de um tratamento medicamentoso por esta via requer que a absorção gastrointestinal do fármaco seja suficiente para assegurar a sua disponibilidade no local de ação (VOLPE, 2010). No entanto, a absorção do fármaco no trato gastrointestinal é complexa e pode ser influenciada por vários fatores, os quais têm impacto sobre a dissolução, solubilidade e permeabilidade do fármaco. Com o intuito de aumentar a biodisponibilidade de fármacos, que possuem absorção dificultada pela via oral, a via de administração pela mucosa bucal vem sendo uma alternativa na atualidade farmacêutica. Esta mucosa é um tecido não queratinizado, altamente vascularizado e apresenta poucas enzimas metabolizadoras. Tais características possibilitam boa absorção de fármacos sem que ocorra a metabolização pré-sistêmica, ou efeito de primeira passagem, somando-se ao fato desta apresentar fácil acessibilidade para a administração de fármacos (VRIES, M. E et al., 1991; NIELSEN, H. M &RASSING, M. R, 1999). Nesse sentido, o presente trabalho teve como objetivo avaliar a permeabilidade dos fármacos antirretrovirais (lamivudina e estavudina) por meio de modelo ex vivo em segmentos da mucosa bucal de suínos, com emprego de câmaras de difusão do tipo células de Franz. Para avaliação da permeabilidade bucal dos fármacos antirretrovirais, lamivudina e estavudina, e dos marcadores para transporte transcelular (metoprolol) e paracelular (fluoresceína sódica), empregou-se método ex-vivo, em células de Franz, com segmento de mucosa bucal de suíno ( a 37ºC, meio Ringer- Krebs- HEPES, pH 7,4), e Franz posterior análise das concentrações das substâncias permeadas (fármacos e marcadores) por cromatografia líquida de alta eficiência. Os resultados obtidos, por meio do protocolo desenvolvido, demonstram que o transporte através da via paracelular (marcador fluoresceína) foi mais expressivo que o transporte transcelular (marcador metoprolol), o que provavelmente se deve ao fato dos espaços intercelulares da mucosa bucal serem mais frouxos do que aqueles observados na mucosa intestinal (junções íntimas). Quanto à lamivudina e estavudina, os resultados de permeabilidade indicaram que estes fármacos permearam por mecanismo semelhante ao do metoprolol, isto é, por via transcelular.

Bacterial translocation in rats nonfunctioning diverted distal colon

To investigate whether the alterations of the diverted colon segment mucosa, evidenced in fecal colitis, would be able to alter Bacterial Translocation (BT). Methods: Sixty-two Wistar male rats ranging from 220 to 320 grams of weight, were divided in two groups: A (Colostomy) and B (Control), with 31 animals each one. In group A, all animals underwent end colostomy, one stoma, in ascending colon; and in the 70th POD was injected in five rats, by rectal route – diverted segment - 2ml of a 0.9% saline solution in animals (A1 subgroup); in eight it was inoculated, by rectal route, 2ml of a solution containing Escherichia coli ATCC 25922 (American Type Culture Collection), in a concentration of 108 Colony Forming Unit for milliliters (CFU/ml) - A2 Subgroup; in ten animals the same solution of E. coli was inoculated, in a concentration of 1011 CFU/ml (A3 Subgroup); and in eight it was collected part of the mucus found in the diverted distal colonic segment for neutral sugars and total proteins dosage (A4 subgroup). The animals from the group B underwent the same procedures of group A, but with differences in the colostomy confection. In rats from subgroups A1, A2, A3, B1, B2, and B3 2ml of blood were aspirated from the heart, and fragments from mesenteric lymphatic nodule, liver, spleen, lung and kidney taken for microbiological analysis, after their death. This analysis consisted of evidencing the presence of E. coli ATCC 25922 CFU. Mann-Whitney and ANOVA Tests were applied as analytic techniques for association of variables. Results: The occurrence of BT was evidenced only in those animals in which inoculated concentration of E. coli ATCC 25922, reached levels of 1011CFU/ml, i.e. in Subgroups A3 and B3, although, being significantly greater (80%) in those animals without colostomy (subgroup B3) when compared to the ones with colostomy (20%) from the subgroup A3 (P <0.05). Lung, liver and mesenteric lymphatic nodules were the tissues with larger percentile of bacterial recovery, so much in subgroup A3, as in B3. Blood culture was considered positive in 60% of the animals from subgroup B3 and in 10% of those from subgroup A3 (p <0.05). There was greater concentration of neutral sugars, in subgroup A4 - mean 27.3mg/ml -, than in subgroup B4 - mean 8.4mg/ml - (P <0.05). Conclusion: The modifications in the architecture of intestinal mucosa in colitis following fecal diversion can cause alterations in the intestinal barrier, but it does not necessarily lead to an increased frequency of BT

Bacterial translocation in rats nonfunctioning diverted distal colon

To investigate whether the alterations of the diverted colon segment mucosa, evidenced in fecal colitis, would be able to alter Bacterial Translocation (BT). Methods: Sixty-two Wistar male rats ranging from 220 to 320 grams of weight, were divided in two groups: A (Colostomy) and B (Control), with 31 animals each one. In group A, all animals underwent end colostomy, one stoma, in ascending colon; and in the 70th POD was injected in five rats, by rectal route – diverted segment - 2ml of a 0.9% saline solution in animals (A1 subgroup); in eight it was inoculated, by rectal route, 2ml of a solution containing Escherichia coli ATCC 25922 (American Type Culture Collection), in a concentration of 108 Colony Forming Unit for milliliters (CFU/ml) - A2 Subgroup; in ten animals the same solution of E. coli was inoculated, in a concentration of 1011 CFU/ml (A3 Subgroup); and in eight it was collected part of the mucus found in the diverted distal colonic segment for neutral sugars and total proteins dosage (A4 subgroup). The animals from the group B underwent the same procedures of group A, but with differences in the colostomy confection. In rats from subgroups A1, A2, A3, B1, B2, and B3 2ml of blood were aspirated from the heart, and fragments from mesenteric lymphatic nodule, liver, spleen, lung and kidney taken for microbiological analysis, after their death. This analysis consisted of evidencing the presence of E. coli ATCC 25922 CFU. Mann-Whitney and ANOVA Tests were applied as analytic techniques for association of variables. Results: The occurrence of BT was evidenced only in those animals in which inoculated concentration of E. coli ATCC 25922, reached levels of 1011CFU/ml, i.e. in Subgroups A3 and B3, although, being significantly greater (80%) in those animals without colostomy (subgroup B3) when compared to the ones with colostomy (20%) from the subgroup A3 (P <0.05). Lung, liver and mesenteric lymphatic nodules were the tissues with larger percentile of bacterial recovery, so much in subgroup A3, as in B3. Blood culture was considered positive in 60% of the animals from subgroup B3 and in 10% of those from subgroup A3 (p <0.05). There was greater concentration of neutral sugars, in subgroup A4 - mean 27.3mg/ml -, than in subgroup B4 - mean 8.4mg/ml - (P <0.05). Conclusion: The modifications in the architecture of intestinal mucosa in colitis following fecal diversion can cause alterations in the intestinal barrier, but it does not necessarily lead to an increased frequency of BT