Interactions between gut-associated lymphoid tissue and colonization levels of indigenous, segmented, filamentous bacteria in the small intestine of mice.

Unlike most other indigenous bacteria, segmented filamentous bacteria (SFB) are potent activators of the mucosal immune system. SFB are strongly anchored to the epithelial cells of the small intestine where they have a preference for mucosal lymphoid epithelium. Since SFB are only present in high numbers shortly after weaning, it was investigated whether an SFB-induced immune reaction results in the removal of these bacteria from the small intestine. A correlation was found between age and colonization levels in the small intestines of SFB monoassociated Swiss mice. Five-week-old athymic BALB/c (nu/nu) mice showed lower colonization levels than their heterozygous littermates, but the opposite was found at the age of 12 weeks. However, SFB inoculation of germfree Swiss mice resulted in higher colonization levels in 5-week-old mice when compared with 4-month-old mice. We conclude that SFB colonization levels in the small intestine are likely influenced by the activity of the mucosal immune system. However, an additional age-dependent factor that modulates SFB colonization levels cannot be excluded.     

Monocyte recruitment, activation, and function in the gut-associated lymphoid tissue during oral Salmonella infection

Neutrophils, monocytes, and dendritic cells (DC) are phenotypically and functionally related phagocytes whose presence in infected tissues is critical to host survival. Their overlapping expression pattern of surface molecules, the differentiation capacity of monocytes, and the presence of monocyte subsets underscores the complexity of understanding the role of these cells during infection. In this study we use five- to seven-color flow cytometry to assess the phenotype and function of monocytes recruited to Peyer's patches (PP) and mesenteric lymph nodes (MLN) after oral Salmonella infection of mice. The data show that CD68(high)Gr-1(int) (intermediate) monocytes, along with CD68(int)Gr-1(high) neutrophils, rapidly accumulate in PP and MLN. The monocytes have increased MHC-II and costimulatory molecule expression and, in contrast to neutrophils and DC, produce inducible NO synthase. Although neutrophils and monocytes from infected mice produce TNF-alpha and IL-1beta upon ex vivo culture, DC do not. In addition, although recruited monocytes internalize Salmonella in vitro and in vivo they did not induce the proliferation of OT-II CD4(+) T cells after coincubation with Salmonella expressing OVA despite their ability to activate OT-II cells when pulsed with the OVA(323-339) peptide. We also show that recruited monocytes enter the PP of infected mice independently of the mucosal address in cell adhesion molecule-1 (MAdCAM-1). Finally, recruited but not resident monocytes increase in the blood of orally infected mice, and MHC-II up-regulation, but not TNF-alpha or iNOS production, occur already in the blood. These studies are the first to describe the accumulation and function of monocyte subsets in the blood and GALT during oral Salmonella infection.     

Morphological alteration of gut-associated lymphoid tissue after long-term total parenteral nutrition in rats

Summary The morphological alteration of gut-associated lymphoid tissue (GALT) induced by long-term absence of dietary stimulation was investigated. Male Wistar rats weighing 230 g were maintained with total parenteral nutrition (TPN). Control rats were allowed to have the same amount of the solution orally. After two weeks, the morphological alteration of GALT was examined. Although no significant difference in weight gain was noted between the two groups, the area comprised by Peyer's patches was decreased in TPN rats. The number of transported lymphocytes and the ratio of helper T (T_h) cells to suppressor/cytotoxic T (T_s/c) cells in intestinal lymph were lowered after TPN treatment. In an immunohistochemical study of the rat ileum, the number of T cells and the T_h/T_s/c ratio were decreased both in the intraepithelial spaces and in the lamina propria of TPN rats. The percentage of interleukin-2 receptor-positive cells and the number of IgA-containing cells in the lamina propria were significantly reduced in TPN rats. These results suggest that dietary stimulation might play a role in the maintenance of GALT function and morphology.     

TLR9 is required for the gut-associated lymphoid tissue response following oral infection of Toxoplasma gondii

TLRs expressed by a variety of cells, including epithelial cells, B cells, and dendritic cells, are important initiators of the immune response following stimulation with various microbial products. Several of the TLRs require the adaptor protein, MyD88, which is an important mediator for the immune response following Toxoplasma gondii infection. Previously, TLR9-mediated innate immune responses were predominantly associated with ligation of unmethylated bacterial CpG DNA. In this study, we show that TLR9 is required for the Th1-type inflammatory response that ensues following oral infection with T. gondii. After oral infection with T. gondii, susceptible wild-type (WT; C57BL/6) but not TLR9(-/-) (B6 background) mice develop a Th1-dependent acute lethal ileitis; TLR9(-/-) mice have higher parasite burdens than control WT mice, consistent with depressed IFN-gamma-dependent parasite killing. A reduction in the total T cell and IFN-gamma-producing T cell frequencies was observed in the lamina propria of the TLR9(-/-) parasite-infected mice. TLR9 and type I IFN production was observed by cells from infected intestines in WT mice. TLR9 expression by dendritic cell populations is essential for their expansion in the mesenteric lymph nodes of infected mice. Infection of chimeric mice deleted of TLR9 in either the hemopoietic or nonhemopoietic compartments demonstrated that TLR9 expression by cells from both compartments is important for efficient T cell responses to oral infection. These observations demonstrate that TLR9 mediates the innate response to oral parasite infection and is involved in the development of an effective Th1-type immune response.     

Impacts of high-sucrose diet on circadian rhythms in the small intestine of rats

Excessive sucrose intake, known as fructose toxicity, leads to fatty liver, hyperlipidemia, and metabolic syndrome. Circadian disorders also contribute to metabolic syndrome. Here, we investigated the effect of excessive sucrose intake on circadian rhythms of the small intestine, the main location of sucrose absorption, to elucidate a mechanism of sucrose-induced abnormal lipid metabolism. Male Wistar rats were fed control starch or high-sucrose diets for 4 weeks. High-sucrose diet-induced fatty liver and hypertriglyceridemia in rats. Amplitudes of PER1/2 expression oscillations in the small intestine were reduced by excessive sucrose, while gene expression of GLUT5 and gluconeogenic enzymes was enhanced. These changes would contribute to interfering in lipid homeostasis as well as adaptive responses to control fructose toxicity in rats.     

A comparative study of the gut-associated lymphoid tissue of primates and rodents

Virchows Archiv B Cell Pathology - Previous studies have suggested that the gut-associated lymphoid tissue (GALT) of man is distinct from that of laboratory animals, but it is not clear whether...     

Morphine inhibits mucosal antibody responses and TGF-beta mRNA in gut-associated lymphoid tissue following oral cholera toxin in mice

In this study, we investigated the effect of morphine on the mucosal immune system using fragment cultures of ileal segments, Peyer's patches (PPs), and mesenteric lymph nodes. Mice were implanted s.c. with a morphine slow release pellet. Control groups received a naltrexone slow release pellet, a placebo pellet, or both a morphine and a naltrexone pellet. After 48 h, mice were orally immunized with cholera toxin (CT) and were boosted orally 1 wk later. Animals were sacrificed 1 wk after the booster immunization, and PPs, mesenteric lymph nodes, and ileal segments were cultured in 24-well plates for 12 days. Morphine resulted in a highly significant inhibition of CT-specific IgA and IgG production in fragment culture supernatants of all three tissues compared with placebo. Naltrexone blocked the reduction in Ab levels induced by morphine, indicating that the effect is opioid receptor mediated. Morphine did not significantly alter total IgA levels in any of the tissue culture supernatants. Morphine also inhibited CT-specific IgA and IgG levels in serum. By flow cytometry, morphine did not alter the lymphoid cell composition in PPs compared with placebo. The effect of morphine on TGF-beta, IL-5, and IL-6 mRNA expression in PPs and ileal segments was determined following oral immunization with CT. Morphine significantly decreased TGF-beta mRNA compared with that in the placebo group, and naltrexone blocked this effect. These results indicate that morphine inhibits Ag-specific IgA responses in gut-associated lymphoid tissue at least partially through the inhibition of TGF-beta, a putative IgA switch factor, in the gastrointestinal tract.     

A comparative study of the gut-associated lymphoid tissue of primates and rodents

Previous studies have suggested that the gut-associated lymphoid tissue (GALT) of man is distinct from that of laboratory animals, but it is not clear whether this is due to environmental or true species difference. We have made a comparative study of rats and baboons because, like rats, baboons are herbivorous and relatively unhygienic but they are phylogenetically much more closely related to man. The Peyer's patches of rats, baboons and man are morphologically very similar in all three species but phenotypically those of man and baboons are different to those of rats. Cells with irregular nuclei ("centrocyte-like" cells) surround the mantle zone in all three species. While these cells express surface IgD and IgM in rats, in man and baboons they express surface IgM or IgA. A population of immunoblasts which express cytoplasmic IgA are present in association with the high endothelial venules of rat Peyer's patches. These cells are not present to the same extent in man or baboons. This suggests that the events between the antigenic stimulation of Peyer's patches and the ultimate seeding of the lamina propria with IgA secreting plasma cells may be different in rodents and primates.     

Changes in enzymatic activity of small intestine and kidney of rats by a methionine deficient diet

The effect of methionine dietary deficiency on food intake, weight gain, liver and kidney weight, feed conversion rate, protein efficiency ratio, maltase and leucineaminopeptidase (LAPase) activities of the intestinal mucosa as well as renal LAPase activity was studied. Three groups of female Wistar rats, weighing between 40-60 g, were fed for 25 days on either Diet A (casein supplemented with 0.6% DL-methionine), Diet B (amino acid mixture simulating casein also supplemented with 0.6% methionine) or Diet C (amino acid mixture with 0.67% methionine deficiency with respect to Diet A). The results show no significant differences in either growth or enzymatic activity between the rats fed on Diet A and those on Diet B. The animals fed on Diet C show an increase in intestinal (P less than 0.01, vs Diet B) and renal (P less than 0.005, vs Diet A) LAPase activity, although intestinal maltase activity remained unchanged. Food intake, weight gain, organ weight and nutritional parameters obtained in rats fed on Diet C showed no statistically significant changes, with the exception of kidney weight which decreased (P less than 0.005) when compared to those fed on Diet B.     

Body weight and tissue composition in rats made obese by a cafeteria diet. Effect of 24 hours starvation

Rat body size and tissue composition changes from pre-weaning to three months age resulted from voluntary hyperphagia triggered by offering a cafeteria diet. The effects of a 24 hour starvation period in both cafeteria and chow fed controls were compared. Obesity develops earlier in females than in males. This difference is related to the growth patterns in both sexes. Obesity occurs at the stages of development when growth rate decreases. Cafeteria fed female rats attained a 32% greater weight than their controls, with lumbar adipose cords that were 4 times heavier and brown interscapular adipose tissue 2 times heavier than controls. The overall cafeteria fed versus chow fed rat differences in the effects of a 24 hour starvation, were minor but less liver glycogen and much more skeletal muscle lipids were mobilized in the cafeteria fed rats than in controls.     

Catecholaminergic, cholinergic and peptidergic innervation of gut-associated lymphoid tissue in porcine jejunum and ileum

With its abundance of neurons and immunocytes, the gut is a potentially important site for the study of the interaction between the nervous and immune systems. Using immunohistochemical techniques, we tested the hypothesis that gut-associated lymphoid tissue in the porcine small intestine might receive catecholaminergic, cholinergic and peptidergic innervation. Antibodies against protein gene product (PGP) 9.5 were employed to detect neuronal membranes; antibodies against tyrosine hydroxylase (TH), type 2 vesicular monoamine transporter (VMAT-2) and choline acetyltransferase (ChAT) were used to detect catecholaminergic and cholinergic neurons; and antibodies to neuromedin U-8 (NMU-8), substance P (SP) and vasoactive intestinal peptide (VIP) were also used. PGP9.5-immunoreactive nerve fibers were observed between jejunal Peyer's patch (PP) follicles and in submucosal ganglia localized at the base of continuous ileal PP. Many ChAT-positive and a few TH-/VMAT-2-immunoreactive neurons or axons adjacent to jejunal and ileal PP were observed. Neurons and fibers from ganglia situated between or at the base of PP follicles manifested robust immunoreactivities to VIP and NMU-8; relatively less SP immunoreactivity was observed at these locations. All neuromedin-U 8-positive neurons observed exhibited immunoreactivity to ChAT as did some VIP-positive neurons. The specific chemical coding of enteric neurons in close apposition to jejunal and ileal PP and the differential localization of neuropeptides within the jejunal and ileal PP are indicative of neuroimmunomodulation at these sites.     

Effect of prostaglandin E2 on the small intestine of indomethacin-treated rats

In the present study, the protective effect of PGE2 on intestinal damage in indomethacin-treated adult rats was investigated. Ileal integrity was evaluated making use of different biochemical and histological parameters: activities of sucrase, maltase and diamine oxidase; concentrations of DNA, putrescine, spermidine and spermine; incorporation of 3H-thymidine into DNA; mitotic index and mucosal thickness. Results expressed per g of mucosal weight, showed that: maltase and diamine oxidase activities as well as DNA, spermidine and spermine concentrations decreased markedly in indomethacin-treated rats when compared to control rats; the decrease of maltase activity as well as DNA, spermidine and spermine concentration was less pronounced in PGE2-treated rats when compared to indomethacin-treated rats; 3H-thymidine incorporation into DNA and mitotic index values showed no significant variation in the course of different treatments; mucosal thickness increased strongly, in PGE2-protected rats. We suggest that PGE2 could protect the rat's intestinal mucosa against the effects of indomethacin through a trophic action on intestinal villi.     

Effect of somatostatin on D-galactose transport across the small intestine of rats

Somatostatin was found to diminish control and theophylline-treated tissue sugar accumulation as well as control and also to diminish theophylline mucosal to serosal D-galactose fluxes. When Na+ was removed from the bath solution, sugar transport was unaltered by the presence of somatostatin. The same results were obtained with phlorizin in the medium. These results seem to suggest that the action of somatostatin is restricted to the Na+-dependent sugar carrier located on the brush border of the intestinal epithelium.     

Effect of distal small bowel resection on ACAT activity and microsomal lipid composition in rat small intestine

1. The acyl-CoA:cholesterol acyltransferase (ACAT) activity and lipid composition of intestinal microsomal membrane were investigated 6 weeks after both 50 and 75% distal small bowel resection (DSBR). 2. No changes in both microsomal ACAT activity and cholesteryl ester levels were found, while microsomal non-esterified cholesterol content was increased after the surgical operation. 3. The total phospholipid content of the microsomes did not change as a result of DSBR. 4. The microsomal phospholipid fatty acid composition showed a significant increase in saturated fatty acids together with no changes in both total monounsaturated and total polyunsaturated fatty acids after resection. 5. An increase in the levels of linoleic acid accompanied by a decrease in arachidonic acid was found in remnant intestine of resected rats.     

Co-localization of vimentin and cytokeratins in M-cells of rabbit gut-associated lymphoid tissue (GALT)

Summary The occurrence of cytokeratins, vimentin, and desmin in the dome epithelia and adjacent non-dome epithelia in four locations of gut-associated lymphoid tissues (GALT) of adult and newborn rabbits (Peyer's patches, sacculus rotundus, caecal lymphoid patches and appendix) was studied with monoclonal antibodies, using the indirect immunoperoxidase technique. In all locations investigated in adult animals, antibodies specific for vimentin labelled (1) M-cells, which engulf intraepithelial lymphocytes, (2) columnar epithelial cells at the base of the domes lacking an apparent contact with lymphocytes (immature M-cells), and (3) flat cells, which lie in the lamina propria under the dome epithelium, and which line the basal lamina with thin cytoplasmic processes. In newborn rabbits, columnar epithelial cells resembling the immature M-cells of adults were selectively stained with vimentin antibodies. In M-cells, the strongest immunoreactivity was present in the perinuclear region and close to the pocket membrane, whereas the most apical and most basal parts of the cytoplasm showed no vimentin-immunoreactivity. Enterocytes in the dome epithelium and in the non-dome epithelium were vimentin-negative. M-cells and enterocytes bound antibodies against cytokeratin peptides 18 and 19 in adults and newborn animals. Compared with enterocytes, M-cells showed less intense staining for cytokeratins. Dome epithelia and no-dome epithelia did not contain desmin-immunoreactive cells. The results suggest that vimentin is a sensitive marker for M-cells in rabbit GALT.     

Changes in the small intestine of Schistosoma mansoni-infected mice fed a high-fat diet

The consumption of a high-fat diet modifies both the morphology of the small intestine and experimentally tested effects of schistosomiasis mansoni. However, whether a schistosomiasis infection associated with a high-fat diet causes injury to the small intestine has never been investigated. Mice were fed either a high-fat or a standard-fat diet for 6 months and were then infected with Schistosoma mansoni cercariae. Physical characteristics of the intestinal tissue (mucosal thickness, small intestinal villi length and height, and abundance of goblet cells and enterocytes on the villous surface) and the distribution of granulomas along the intestinal segments and their developmental stage were measured at the time of sacrifice (9 or 17 weeks post-infection). The group fed a high-fat diet exhibited different granuloma stages, whereas the control group possessed only exudative granulomas. The chronically infected mice fed a high-fat diet exhibited higher granuloma and egg numbers than the acutely infected group. Exudative, exudative/exudative-productive and exudative-productive granulomas were present irrespective of diet. Computer-aided morphometric analysis confirmed that villus length, villus width, muscular height and submucosal height of the duodenal and jejunal segments were affected by diet and infection. In conclusion, a high-fat diet and infection had a significant impact on the small intestine morphology and morphometry among the animals tested.