The mucosal firewalls against commensal intestinal microbes

Mammals coexist with an extremely dense microbiota in the lower intestine. Despite the constant challenge of small numbers of microbes penetrating the intestinal surface epithelium, it is very unusual for these organisms to cause disease. In this review article, we present the different mucosal firewalls that contain and allow mutualism with the intestinal microbiota.

Host responses to intestinal microbial antigens in gluten-sensitive mice

BACKGROUND AND AIMS: Excessive uptake of commensal bacterial antigens through a permeable intestinal barrier may influence host responses to specific antigen in a genetically predisposed host. The aim of this study was to investigate whether intestinal barrier dysfunction induced by indomethacin treatment affects the host response to intestinal microbiota in gluten-sensitized HLA-DQ8/HCD4 mice. METHODOLOGY/PRINCIPAL FINDINGS: HLA-DQ8/HCD4 mice were sensitized with gluten, and gavaged with indomethacin plus gluten. Intestinal permeability was assessed by Ussing chamber; epithelial cell (EC) ultra-structure by electron microscopy; RNA expression of genes coding for junctional proteins by Q-real-time PCR; immune response by in-vitro antigen-specific T-cell proliferation and cytokine analysis by cytometric bead array; intestinal microbiota by fluorescence in situ hybridization and analysis of systemic antibodies against intestinal microbiota by surface staining of live bacteria with serum followed by FACS analysis. Indomethacin led to a more pronounced increase in intestinal permeability in gluten-sensitized mice. These changes were accompanied by severe EC damage, decreased E-cadherin RNA level, elevated IFN-gamma in splenocyte culture supernatant, and production of significant IgM antibody against intestinal microbiota. CONCLUSION: Indomethacin potentiates barrier dysfunction and EC injury induced by gluten, affects systemic IFN-gamma production and the host response to intestinal microbiota antigens in HLA-DQ8/HCD4 mice. The results suggest that environmental factors that alter the intestinal barrier may predispose individuals to an increased susceptibility to gluten through a bystander immune activation to intestinal microbiota.

Production of interferon-gamma by activated T-cell receptor-alphabeta CD8alphabeta intestinal intraepithelial lymphocytes is required and sufficient for disruption of the intestinal barrier integrity

Maintenance of intestinal epithelial barrier function is of vital importance in preventing uncontrolled influx of antigens and the potentially ensuing inflammatory disorders. Intestinal intraepithelial lymphocytes (IEL) are in intimate contact with epithelial cells and may critically regulate the epithelial barrier integrity. While a preserving impact has been ascribed to the T-cell receptor (TCR)-gammadelta subset of IEL, IEL have also been shown to attenuate the barrier function. The present study sought to clarify the effects of IEL by specifically investigating the influence of the TCR-alphabeta CD8alphabeta and TCR-alphabeta CD8alphaalpha subsets of IEL on the intestinal epithelial barrier integrity. To this end, an in vitro coculture system of the murine intestinal crypt-derived cell-line mIC(cl2) and syngeneic ex vivo isolated IEL was employed. Epithelial integrity was assessed by analysis of transepithelial resistance (TER) and paracellular flux of fluorescein isothiocyanate-conjugated (FITC-) dextran. The TCR-alphabeta CD8alphaalpha IEL and resting TCR-alphabeta CD8alphabeta IEL did not affect TER of mIC(cl2) or flux of FITC-dextran. In contrast, activated TCR-alphabeta CD8alphabeta IEL clearly disrupted the integrity of the mIC(cl2) monolayer. No disrupting effect was seen with activated TCR-alphabeta CD8alphabeta IEL from interferon-gamma knockout mice. These findings demonstrate that secretion of interferon-gamma by activated TCR-alphabeta CD8alphabeta IEL is strictly required and also sufficient for disrupting the intestinal epithelial barrier function.

[Tritrichomonas fetus: a new intestinal parasite in Swiss cats]

Recent reports identified Tritrichomonas fetus, the causative agent of bovine trichomonosis, in cats with large-bowel diarrhea in the US. Between July 2007 and August 2008, a total of 105 Swiss cats were tested for T. fetus with the InPouchTM culture system and/or PCR, whereof 27 (26%) yielded positive results. All positive cats were pedigree cats, whereof 22 (81%) were less than 1 year of age (median 5 months). 25 (93%) of these cats lived in multi-cat households, and all but one were kept indoor. The clinical picture was dominated by large bowel diarrhea with increased frequency of defecation and fresh blood and mucus. Furthermore, inflamed anus and fecal incontinence was common. 52% of the T. fetus-positive cats were tested positive for Giardia before, but the treatment with fenbendazole or metronidazole only temporarily alleviated the clinical signs. The treatment with 30 mg/kg of ronidazole q12h p.o. was successful in all but 1 cat with only minor transient adverse effects in 3 cats. In conclusion, T. fetus has to be considered an important causative agent of large bowel diarrhea in cats in Switzerland, especially in young indoor pedigree cats.

Intestinal Tritrichomonas foetus infection in cats in Switzerland detected by in vitro cultivation and PCR

Tritrichomonas foetus, a parasite well known for its significance as venereally transmitted pathogen in cattle, has recently been identified as a cause of chronic large-bowel diarrhea in domestic cats in the US, UK, and, more recently, also in Norway. In a period of 3 months (October to December 2007), 45 cats of Switzerland suffering from chronic diarrhea were investigated for intestinal infections, including a search for trichomonads. A commercially available in vitro culture system was used to screen for infection, complemented with a PCR and subsequent amplicon sequencing to support speciation. The PCR is based upon amplification of a sequence derived from the internal transcribed spacer region 1 (ITS1) on the ribosomal RNA gene (rRNA) using primers designed to detect a broad range of genera and species belonging to the family of Trichomonadidae. The method was furthermore adapted to the uracil DNA glycosylase (UDG) system in order to prevent carry-over contamination and it included a recombinant internal control to track for inhibitory reactions. Eleven out of the 45 cats were culture-positive, as revealed by microscopic identification of trichomonadid organisms. One of the isolates was subjected to scanning electron microscopy and findings revealed the presence of three flagella, thus placing the isolate into the gender Tritrichomonas sp. PCR and subsequent amplicon sequencing were carried out with ten of the 11 isolates. A total homology with published T. foetus sequences was confirmed in all of the cases. T. foetus therefore appears to range among those organisms that can cause chronic diarrhea in cats in Switzerland.

Small intestinal intussusception in five dogs with acute renal failure and suspected leptospirosis (L. australis)

OBJECTIVES This case series describes 5 dogs with small intestinal intussusception and acute kidney injury due to infection with Leptospira interrogans serovar Australis. CASE SERIES SUMMARY Small intestinal intussusception was observed in 4 dogs diagnosed with acute kidney injury due to leptospirosis presented between 1997 and 2005. Intussusception was diagnosed at initial presentation or later during hospitalization. An additional dog fulfilling our inclusion criteria was presented to a small animal specialty clinic nearby and was included. Upon admission, all dogs were severely azotemic and thrombocytopenic. All 5 dogs showed the strongest microscopic agglutination test serology reaction to L. interrogans serovar Australis. Two dogs survived with no apparent residual renal damage, 1 survived with subsequent mild chronic kidney disease, and 2 dogs were euthanized at the owners' request due to a guarded prognosis. NEW OR UNIQUE INFORMATION PROVIDED Intussusception can occur or may be seen in dogs with leptospirosis due to L. interrogans serovar Australis and patients should be monitored closely for this potential complication. As all 5 dogs described in this case series showed the highest titer for L. interrogans serovar Australis, these precautions may be especially applied in geographic areas where this particular serovar is seen.

Morphometry and allometry of the postnatal marsupial lung development: an ultrastructural study

An utrastructural morphometric study of the postnatally remodelling lungs of the quokka wallaby (Setonix brachyurus) was undertaken. Allometric scaling of the volumes of the parenchymal components against body mass was performed. Most parameters showed a positive correlation with body mass in all the developmental stages, except the volume of type II pneumocytes during the alveolar stage. The interstitial tissue and type II cell volumes increased slightly faster than body mass in the saccular stage, their growth rates declining in the alveolar stage. Conversely, type I pneumocyte volumes increased markedly in both the saccular and alveolar stages. Both capillary and endothelial volumes as well as the capillary and airspace surface areas showed highest rates of increase during the alveolar stage, at which time the rate was notably higher than that of the body mass. The pulmonary diffusion capacity increased gradually, the rate being highest in the alveolar stage and the adult values attained were comparable to those of eutherians.

Clinical parameters, intestinal function, and IGF1 concentrations in colostrum-deprived and colostrum-fed newborn pony foals.

Colostrum (COL) contains cytokines and growth factors that may enhance intestinal development in neonates. The hypothesis of this study was that besides providing immunoglobulins, COL is important for intestinal function and meconium release in foals. Newborn foals were either fed COL (n = 5) or an equal amount of milk replacer (MR, n = 7) during the first 24 hours of life. To ensure passive immunity, all foals received 1 L plasma. Postnatal development, meconium release, intestinal motility, white blood cell count, insulin-like growth factor 1, and intestinal absorptive function (xylose absorption test) were evaluated. Clinical findings and meconium release were not affected by feeding of COL or MR. Ultrasonography revealed a slightly larger jejunum and stomach in group COL versus MR (P < 0.05). The percentage of polymorphonuclear leucocytes was higher in foals of group MR versus group COL (P < 0.05) and the percentage of lymphocytes was lower in MR compared with COL foals (P < 0.05). Plasma insulin-like growth factor 1 concentration increased during the first 14 days after birth in both groups. A xylose absorption test on Day 5 revealed similar increases in plasma xylose concentrations after oral intake. In conclusion, feeding of COL versus MR was without effect on meconium release and intestinal absorptive function. Differences between foals fed COL and MR with regard to intestinal function are apparently without clinical relevance. In foals that have not received maternal COL, there is no major risk of intestinal problems if they are fed MR and provided with immunoglobulins by transfusion of plasma.

Lack of Host Gut Microbiota Alters Immune Responses and Intestinal Granuloma Formation during Schistosomiasis

Fatalities from schistosome infections arise due to granulomatous, immune-mediated responses to eggs that become trapped in host tissues. Schistosome-specific immune responses are characterized by initial Th1 responses and our previous studies demonstrated that Myd88-deficient mice failed to initiate such responses in vivo. Paradoxically, schistosomal antigens fail to stimulate innate cells to release pro-inflammatory cytokines in vitro. Since S. mansoni infection is an intestinal disease, we hypothesized that commensal bacteria could act as bystander activators of the intestinal innate immune system to instigate Th1 responses. Using a broad spectrum of orally-administered antibiotics and antimycotics we analyzed schistosome-infected mice that were simultaneously depleted of gut bacteria. After depletion there was significantly less inflammation in the intestine which was accompanied by decreased intestinal granuloma development. In contrast, liver pathology remained unaltered. In addition, schistosome-specific immune responses were skewed and fecal egg excretion was diminished. This study demonstrates that host microbiota can act as a third partner in instigating helminth-specific immune responses.

Interferon-γ Induces Expression of MHC Class II on Intestinal Epithelial Cells and Protects Mice from Colitis

Immune responses against intestinal microbiota contribute to the pathogenesis of inflammatory bowel diseases (IBD) and involve CD4(+) T cells, which are activated by major histocompatibility complex class II (MHCII) molecules on antigen-presenting cells (APCs). However, it is largely unexplored how inflammation-induced MHCII expression by intestinal epithelial cells (IEC) affects CD4(+) T cell-mediated immunity or tolerance induction in vivo. Here, we investigated how epithelial MHCII expression is induced and how a deficiency in inducible epithelial MHCII expression alters susceptibility to colitis and the outcome of colon-specific immune responses. Colitis was induced in mice that lacked inducible expression of MHCII molecules on all nonhematopoietic cells, or specifically on IECs, by continuous infection with Helicobacter hepaticus and administration of interleukin (IL)-10 receptor-blocking antibodies (anti-IL10R mAb). To assess the role of interferon (IFN)-γ in inducing epithelial MHCII expression, the T cell adoptive transfer model of colitis was used. Abrogation of MHCII expression by nonhematopoietic cells or IECs induces colitis associated with increased colonic frequencies of innate immune cells and expression of proinflammatory cytokines. CD4(+) T-helper type (Th)1 cells - but not group 3 innate lymphoid cells (ILCs) or Th17 cells - are elevated, resulting in an unfavourably altered ratio between CD4(+) T cells and forkhead box P3 (FoxP3)(+) regulatory T (Treg) cells. IFN-γ produced mainly by CD4(+) T cells is required to upregulate MHCII expression by IECs. These results suggest that, in addition to its proinflammatory roles, IFN-γ exerts a critical anti-inflammatory function in the intestine which protects against colitis by inducing MHCII expression on IECs. This may explain the failure of anti-IFN-γ treatment to induce remission in IBD patients, despite the association of elevated IFN-γ and IBD.