Host lysozyme-mediated lysis of Lactococcus lactis facilitates delivery of colitis-attenuating superoxide dismutase to inflamed colons

Beneficial microbes that target molecules and pathways, such as oxidative stress, which can negatively affect both host and microbiota, may hold promise as an inflammatory bowel disease therapy. Prior work showed that a five-strain fermented milk product (FMP) improved colitis in T-bet^−/− Rag2^−/− mice. By varying the number of strains used in the FMP, we found that Lactococcus lactis I-1631 was sufficient to ameliorate colitis. Using comparative genomic analyses, we identified genes unique to L. lactis I-1631 involved in oxygen respiration. Respiration of oxygen results in reactive oxygen species (ROS) generation. Also, ROS are produced at high levels during intestinal inflammation and cause tissue damage. L. lactis I-1631 possesses genes encoding enzymes that detoxify ROS, such as superoxide dismutase (SodA). Thus, we hypothesized that lactococcal SodA played a role in attenuating colitis. Inactivation of the sodA gene abolished L. lactis I-1631’s beneficial effect in the T-bet^−/− Rag2^−/− model. Similar effects were obtained in two additional colonic inflammation models, Il10^−/− mice and dextran sulfate sodium-treated mice. Efforts to understand how a lipophobic superoxide anion (O₂⁻) can be detoxified by cytoplasmic lactoccocal SodA led to the finding that host antimicrobial-mediated lysis is a prerequisite for SodA release and SodA’s extracytoplasmic O₂⁻ scavenging. L. lactis I-1631 may represent a promising vehicle to deliver antioxidant, colitis-attenuating SodA to the inflamed intestinal mucosa, and host antimicrobials may play a critical role in mediating SodA’s bioaccessibility.Inflammatory bowel disease (IBD) pathophysiology is driven by both host genetic mutations and the gut microbiota. Immune dysregulation in IBD can result from deficiencies in acute inflammatory response pathways (1) or impaired counterregulation of immune responsiveness (2). Host production of reactive oxygen species (ROS) is an evolutionarily conserved response to microbes (3). However, chronic and excessive ROS up-regulate host inflammatory pathways (4, 5) and result in oxidative stress. Chronic intestinal inflammation and oxidative stress affect not only the host but also the microbiota. Oxidative stress within the lumen is a fitness challenge for gut anaerobic bacteria. IBD patient fecal microbiomes reflect a pattern of response to oxidative stress with enrichments in genes for sulfate transport and cysteine and glutathione metabolism (6). In IBD, oxidative stress contributes to chronic inflammation and dysbiosis, and modulating oxidative stress may help to restore intestinal homeostasis.Beneficial microbes hold promise for IBD inflammation and dysbiosis (7). However, human clinical trials have shown mixed results (8, 9) because of variations in microbes under study and patient heterogeneity. Preclinical studies that use model systems that recapitulate key features of the human disease are needed to elucidate the mechanism of action of beneficial microbes on hosts and their microbiota. Such information facilitates clinical trial design by identifying patients with the host and microbial features most likely to benefit from the bioactivity of a beneficial microbe. Identifying microbes that target molecules and pathways such as oxidative stress—which negatively affects both host and microbiota—affords opportunities for new IBD therapies.Building on prior studies examining how a five-strain fermented milk product (FMP) affected the gut microbiome in a preclinical model of colitis and human subjects (10–12), herein we focused on how individual bacterial strains in the FMP affected host response in several preclinical colitis models. One of the five strains, Lactococcus lactis subsp. lactis CNCM I-1631 (L. lactis I-1631), reduced gut oxidative stress and attenuated colitis in three mouse models of colonic inflammation: BALB/c T-bet^−/− Rag2^−/− mice, BALB/c Il10^−/− mice, and BALB/c wild-type mice treated with dextran sulfate sodium (DSS), a colitogenic mucosal disruptant. The colitis-attenuating activity of L. lactis I-1631 was dependent on L. lactis I-1631 superoxide dismutase A (SodA), which reduced colonic epithelial ROS. Our data also support that host factors, increased at sites of inflammation, facilitated targeted delivery of L. lactis I-1631 effects. Specifically, lysis of L. lactis I-1631 by the host peptidoglycan hydrolase and antimicrobial lysozyme-1 appeared to mediate L. lactis I-1631 SodA release and reduction in host oxidative stress.     

Akkermansia muciniphila strain ATCC BAA-835 does not promote short-term intestinal inflammation in gnotobiotic interleukin-10-deficient mice

Akkermansia muciniphila is a common member of the intestinal microbiota of healthy human individuals. Its abundance is negatively associated with inflammatory bowel disease and metabolic disorders and the oral administration of A. muciniphila improves the symptoms of metabolic disease in mice. Therefore, A. muciniphila is a promising candidate for the treatment of type-2 diabetes and obesity. However, some studies using animal models of intestinal inflammation reported that A. muciniphila may exacerbate gut inflammation. Because of these contradictory reports the present study aimed to clarify the role of A. muciniphila in the development of intestinal inflammation and the conditions promoting it. For this purpose, the short-term colitogenic potential of A. muciniphila strain ATCC BAA-835 was investigated in colitis-prone, gnotobiotic IL-10-deficient (Il10^-/-) mice. Il10^-/- mice mono-associated with A. muciniphila showed no signs of intestinal inflammation based on body-weight change, histopathological scoring and inflammatory markers. Additional association of the mice with the colitogenic Escherichia coli strain NC101 led to cecal but not colonic inflammation. However, the severity of the inflammation did not exceed that observed in mice mono-associated with E. coli NC101. Il10^-/- mice colonized with a simplified human intestinal microbiota showed increased histopathology, but no increase in inflammatory markers. Furthermore, co-colonization with A. muciniphila did not modify histopathology. The turnover of intestinal mucus was similar in all groups despite the mucus-degrading property of A. muciniphila. Overall, the data do not support a short-term pro-inflammatory effect of A. muciniphila strain ATCC BAA-835 in the Il10^-/- mouse model for inflammatory bowel disease.     

Dietary iron variably modulates assembly of the intestinal microbiota in colitis-resistant and colitis-susceptible mice

ABSTRACT

Iron deficiency, a common comorbidity of gastrointestinal inflammatory disorders such as inflammatory bowel diseases (IBD), is often treated with oral iron supplementation. However, the safety of oral iron supplementation remains controversial because of its association with exacerbated disease activity in a subset of IBD patients. Because iron modulates bacterial growth and function, one possible mechanism by which iron may exacerbate inflammation in susceptible hosts is by modulating the intestinal microbiota. We, therefore, investigated the impact of dietary iron on the intestinal microbiota, utilizing the conventionalization of germ-free mice as a model of a microbial community in compositional flux to recapitulate the instability of the IBD-associated intestinal microbiota. Our findings demonstrate that altering intestinal iron availability during community assembly modulated the microbiota in non-inflamed wild type (WT) and colitis-susceptible interleukin-10-deficient (Il10^?/-) mice. Depletion of luminal iron availability promoted luminal compositional changes associated with dysbiotic states irrespective of host genotype, including an expansion of Enterobacteriaceae such as Escherichia coli. Mechanistic in vitro growth competitions confirmed that high-affinity iron acquisition systems in E. coli enhance its abundance over other bacteria in iron-restricted conditions, thereby enabling pathobiont iron scavenging during dietary iron restriction. In contrast, distinct luminal community assembly was observed with dietary iron supplementation in WT versus Il10^?/- mice, suggesting that the effects of increased iron on the microbiota differ with host inflammation status. Taken together, shifts in dietary iron intake during community assembly modulate the ecological structure of the intestinal microbiota and is dependent on host genotype and inflammation status.     

Gut microbial diversity is reduced and is associated with colonic inflammation in a piglet model of short bowel syndrome

Background and objectives

Following small bowel resection (SBR), the luminal environment is altered, which contributes to clinical manifestations of short bowel syndrome (SBS) including malabsorption, mucosal inflammation and bacterial overgrowth. However, the impact of SBR on the colon has not been well-defined. The aims of this study were to characterize the colonic microbiota following SBR and to assess the impact of SBR on mucosal inflammation in the colon.

Results

Analysis of the colonic microbiota demonstrated that there was a significant level of dysbiosis both two and six weeks post-SBR, particularly in the phylum Firmicutes, coupled with a decrease in overall bacterial diversity in the colon. This decrease in diversity was associated with an increase in colonic inflammation six weeks post-surgery.

Methods

Female (4-week old) piglets (5−6/group) received a 75% SBR, a transection (sham) or no surgery. Compositional analysis of the colonic microbiota was performed by high-throughput sequencing, two- and six-weeks post-surgery. The gene expression of the pro-inflammatory cytokines interleukin (IL)-1β, IL-6, IL-8, IL-18 and tumor necrosis factor (TNF)-α in the colonic mucosa was assessed by qRT-PCR and the number of macrophages and percentage inducible nitric oxide synthase (iNOS) staining in the colonic epithelium were quantified by immunohistochemistry.

Conclusions

SBR significantly decreased the diversity of the colonic microbiota and this was associated with an increase in colonic mucosal inflammation. This study supports the hypothesis that SBR has a significant impact on the colon and that this may play an important role in defining clinical outcome.     

Intestinal steroid profiles and microbiota composition in colitic mice

Reduced gut microbiota diversity in conjunction with a bloom of few bacterial species is a common feature in inflammatory bowel disease (IBD) patients. However, the environmental changes caused by inflammation and their possible impact on the microbiota are largely unknown. Since IBD is associated with an impaired intestinal steroid metabolism, we hypothesized that changes in intestinal steroid and particularly bile acid (BA) concentrations affect microbial communities. We used Interleukin-10 deficient (IL-10-/-) mice as a model for chronic gut inflammation. Healthy wild-type mice served as controls. In these animals, intestinal steroid concentrations and gut microbial diversity were analyzed at 24 weeks of age. The IL 10-/- mice developed moderate inflammation in cecum and colon and colorectal tumor formation was observed in 55 % of the animals. Compared to the healthy conditions, gut inflammation was associated with higher intestinal cholesterol and cholic acid concentrations and a reduced microbial diversity. The latter was accompanied by a proliferation of Robinsoniella peoriensis, Clostridium innocuum, Escherichia coli, and Enterococcus gallinarum. All these species proved to be highly bile acid resistant. We concluded that chronic colitis in IL-10-/- mice is associated with changes in intestinal steroid profiles. These changes may be due to alterations in gut microbiota composition or vice versa. Whether the bacterial sterol and bile acid metabolism is implicated in colitis and colorectal carcinoma etiology remains to be clarified.     

Intestinal steroid profiles and microbiota composition in colitic mice

Reduced gut microbiota diversity in conjunction with a bloom of few bacterial species is a common feature in inflammatory bowel disease (IBD) patients. However, the environmental changes caused by inflammation and their possible impact on the microbiota are largely unknown. Since IBD is associated with an impaired intestinal steroid metabolism, we hypothesized that changes in intestinal steroid and particularly bile acid (BA) concentrations affect microbial communities. We used Interleukin-10 deficient (IL-10-/-) mice as a model for chronic gut inflammation. Healthy wild-type mice served as controls. In these animals, intestinal steroid concentrations and gut microbial diversity were analyzed at 24 weeks of age. The IL 10-/- mice developed moderate inflammation in cecum and colon and colorectal tumor formation was observed in 55 % of the animals. Compared to the healthy conditions, gut inflammation was associated with higher intestinal cholesterol and cholic acid concentrations and a reduced microbial diversity. The latter was accompanied by a proliferation of Robinsoniella peoriensis, Clostridium innocuum, Escherichia coli, and Enterococcus gallinarum. All these species proved to be highly bile acid resistant. We concluded that chronic colitis in IL-10-/- mice is associated with changes in intestinal steroid profiles. These changes may be due to alterations in gut microbiota composition or vice versa. Whether the bacterial sterol and bile acid metabolism is implicated in colitis and colorectal carcinoma etiology remains to be clarified.     

Longitudinal analysis of inflammation and microbiota dynamics in a model of mild chronic dextran sulfate sodium-induced colitis in mice

AIM:To characterize longitudinally the inflammation and the gut microbiota dynamics in a mouse model of dextran sulfate sodium(DSS)-induced colitis.METHODS:In animal models,the most common method used to trigger colitis is based on the oral administration of the sulfated polysaccharides DSS.The murine DSS colitis model has been widely adopted to induce severe acute,chronic or semi-chronic colitis,and has been validated as an important model for the translation of mice data to human inflammatory bowel disease(IBD).However,it is now clear that models characterized by mild intestinal damage are more accurate for studying the effects of therapeutic agents.For this reason,we have developed a murine model of mild colitis to study longitudinally the inflammation and microbiota dynamics during the intestinal repair processes,and to obtain data suitable to support the recovery of gut microbiota-host homeostasis.RESULTS:All plasma cytokines evaluated,except IL-17,began to increase(P<0.05),after 7 d of DSS administration.IL-17 only began to increase 4 d after DSS withdrawal.IL-1βand IL-17 continue to increase during the recovery phase,even when clinical signs of colitis had disappeared.IL-6,IL-10 and IFN-γreached their maxima 4 d after DSS withdrawal and decreased during the late recovery phase.TNFαreached a peak(a three-fold increase,P<0.05),after which it slightly decreased,only to increase again close to the end of the recovery phase.DSS administration induced profound and rapid changes in the mice gut microbiota.After 3 d of DSS administration,we observed a major reduction in Bacteroidetes/Prevotella and a corresponding increase in Bacillaceae,with respect to control mice.In particular,Bacteroidetes/Prevotella decreased from a relative abundance of 59.42%-33.05%,while Bacillaceae showed a concomitant increase from 2.77%to 10.52%.Gut microbiota rapidly shifted toward a healthy profile during the recovery phase and returned normal 4 d after DSS withdrawal.Cyclooxygenase 2 expression started to increase 4 d after DSS withdrawal(P<0.05),when dysbiosis had recovered,and continued to increase during the recovery phase.Taken together,these data indicated that a chronic phase of intestinal inflammation,characterized by the absence of dysbiosis,could be obtained in mice using a single DSS cycle.CONCLUSION:Dysbiosis contributes to the local and systemic inflammation that occurs in the DSS model of colitis;however,chronic bowel inflammation is maintained even after recovery from dysbiosis.     

Cystic fibrosis transmembrane conductance regulator knockout mice exhibit aberrant gastrointestinal microbiota

The composition of the gastrointestinal microbiome is increasingly recognized as a crucial contributor to immune and metabolic homeostasis—deficiencies in which are characteristic of cystic fibrosis (CF) patients. The murine model (CFTR^−/−, CF), has, in previous studies, demonstrated characteristic CF gastrointestinal (GI) manifestations including slowed transit and significant upregulation of genes associated with inflammation. To determine if characteristics of the microbiome are associated with these phenotypes we used a phylogenetic microarray to compare small intestine bacterial communities of wild type and congenic CF mice. Loss of functional CFTR is associated with significant decreases in GI bacterial community richness, evenness and diversity and reduced relative abundance of putative protective species such as Acinetobacter lwoffii and a multitude of Lactobacilliales members. CF mice exhibited significant enrichment of Mycobacteria species and Bacteroides fragilis, previously associated with GI infection and immunomodulation. Antibiotic administration to WT and CF animals resulted in convergence of their microbiome composition and significant increases in community diversity in CF mice. These communities were characterized by enrichment of members of the Lactobacillaceae and Bifidobacteriaceae and reduced abundance of Enterobacteriaceae and Clostridiaceae. These data suggest that Enterobacteria and Clostridia species, long associated with small intestinal overgrowth and inflammatory bowel disease, may suppress both ileal bacterial diversity and the particular species which maintain motility and immune homeostasis in this niche. Thus, these data provide the first indications that GI bacterial colonization is strongly impacted by the loss of functional CFTR and opens up avenues for alternative therapeutic approaches to improve CF disease management.     

Orally administered extract from Prunella vulgaris attenuates spontaneous colitis in mdr1a~(-/-) mice

AIM: To investigate the ability of a Prunella vulgaris(P. vulgaris) ethanolic extract to attenuate spontaneous typhlocolitis in mdr1a-/- mice. METHODS: Vehicle(5% ethanol) or P. vulgaris ethanolic extract(2.4 mg/d) were administered daily by oral gavage to mdr1a-/- or wild type FVBWT mice from 6 wk of age up to 20 wk of age. Clinical signs of disease were noted by monitoring weight loss. Mice experiencingweight loss in excess of 15% were removed from the study. At the time mice were removed from the study, blood and colon tissue were collected for analyses that included histological evaluation of lesions, inflammatory cytokine levels, and myeloperoxidase activity. RESULTS: Administration of P. vulgaris extracts to mdr1a-/- mice delayed onset of colitis and reduced severity of mucosal inflammation when compared to vehicle-treated mdr1a-/- mice. Oral administration of the P. vulgaris extract resulted in reduced(P 0.05) serum levels of IL-10(4.6 ± 2 vs 19.4 ± 4), CXCL9(1319.0 ± 277 vs 3901.0 ± 858), and TNFα(9.9 ± 3 vs 14.8 ± 1) as well as reduced gene expression by more than two-fold for Ccl2, Ccl20, Cxcl1, Cxcl9, IL-1 α, Mmp10, VCAM-1, ICAM, IL-2, and TNFα in the colonic mucosa of mdr1a-/- mice compared to vehicle-treated mdr1a-/-mice. Histologically, several microscopic parameters were reduced(P 0.05) in P. vulgaris-treated mdr1a-/-mice, as was myeloperoxidase activity in the colon(2.49 ± 0.16 vs 3.36 ± 0.06, P 0.05). The numbers of CD4+ T cells(2031.9 ± 412.1 vs 5054.5 ± 809.5) and germinal center B cells(2749.6 ± 473.7 vs 4934.0 ± 645.9) observed in the cecal tonsils of P. vulgaris-treated mdr1a-/- were significantly reduced(P 0.05) from vehicle-treated mdr1a-/- mice. Vehicle-treated mdr1a-/- mice were found to produce serum antibodies to antigens derived from members of the intestinal microbiota, indicative of severe colitis and a loss of adaptive tolerance to the members of the microbiota. These serum antibodies were greatly reduced or absent in P. vulgaris-treated mdr1a-/- mice. CONCLUSION: The anti-inflammatory activity of P. vulgaris ethanolic extract effectively attenuated the severity of intestinal inflammation in mdr1a-/- mice.     

Intestinal microbiota: A source of novel biomarkers in inflammatory bowel diseases?

The human intestine harbours a complex microbial ecosystem that performs manifold functions important to the nutrition and health of its host. Extensive study has revealed that the composition of the intestinal microbiota is altered in individuals with inflammatory bowel disease (IBD). The IBD associated intestinal microbiota generally has reduced species richness and diversity, lower temporal stability, and disruption of the secreted mucus layer structure. Multiple studies have identified certain bacterial taxa that are enriched or depleted in IBD including Enterobacteriaceae, Ruminococcus gnavus, and Desulfovibrio (enriched) and Faecalibacterium prausnitzii, Lachnospiraceae, and Akkermansia (depleted). Additionally, the relative abundance of some taxa appears to correlate with established markers of disease activity such as Enterobacteriaceae (enriched) and Lachnospiraceae (depleted). Signature shifts in fecal microbial community composition may therefore prove to be valuable as diagnostic biomarkers, particularly for longitudinal monitoring of disease activity and response to treatments.     

Fungi and inflammatory bowel diseases: Alterations of composition and diversity

Objective. Altered bacterial diversity of the intestinal mucosa-associated microbiota may reflect the net influence of lifestyle factors associated with the development of chronic inflammatory bowel diseases (IBD). While a reduced bacterial diversity has been reported in IBD, little is known about the fungal microbiota. The aim of this study was to carry out a systematic analysis of intestinal fungal microbiota in IBD. Material and methods. The mucosa-associated fungal microbiota of 104 colonic biopsy tissues from 47 controls and 57 IBD patients was investigated using metagenomic 18S rDNA-based denaturing gradient gel electrophoresis (DGGE), clone libraries, sequencing, and in situ hybridization techniques. Results. Fungi-specific 18S rDNA signatures could be detected in all 104 patients, accounting for only a small proportion of the intestinal microbiota (0.02% of the mucosal and 0.03% of the fecal microbiota). An overall fungal biodiversity of 43 different operational taxonomic units (OTUs) was found in the clone libraries. The qualitative composition of fungal microbiota was different between patients with IBD and controls. The DGGE profiles showed a higher mean fungal diversity in patients with Crohn's disease (CD) in comparison with controls (10.8±3.1 versus 6.2±2.4 for CD, p ≤ 0.001). No disease-specific fungal species were found in the CD and ulcerative colitis (UC) group. Conclusions. Diverse fungal species are part of the normal enteric microbiota, but diversity is increased and composition of the fungal communities varies in IBD. Further work is needed to investigate whether the alteration of the fungal flora in IBD is secondary to an imbalanced bacterial microbiota or an independent etiologic factor.     

From the Cover: A human IL10 BAC transgene reveals tissue-specific control of IL-10 expression and alters disease outcome

Interleukin (IL)-10 is an immunoregulatory cytokine that is produced by diverse cell populations. Studies in mice suggest that the cellular source of IL-10 is a key determinant in various disease pathologies, yet little is known regarding the control of tissue-specific human IL-10 expression. To assess cell type-specific human IL-10 regulation, we created a human IL-10 transgenic mouse with a bacterial artificial chromosome (hIL10BAC) in which the IL10 gene is positioned centrally. Since human IL-10 is biologically active in the mouse, we could examine the in vivo capacity of tissue-specific human IL-10 expression to recapitulate IL-10-dependent phenotypes by reconstituting Il10^−/− mice (Il10^−/−/hIL10BAC). In response to LPS, Il10^−/−/hIL10BAC mice proficiently regulate IL-10-target genes and normalize sensitivity to LPS toxicity via faithful human IL-10 expression from macrophages and dendritic cells. However, in the Leishmania donovani model of pathogen persistence, Il10^−/−/hIL10BAC mice did not develop the characteristic IL-10⁺IFN-γ⁺CD4 T cell subset thought to mediate persistence and, like Il10^−/− mice, cleared the parasites. Furthermore, the IL-10-promoting cytokine IL-27 failed to regulate transgenic human IL-10 production in CD4⁺ T cells in vitro which together suggests that the hIL10BAC encodes for weak T cell-specific IL-10 expression. Thus, the hIL10BAC mouse is a model of human gene structure and function revealing tissue-specific regulatory requirements for IL-10 expression which impacts disease outcomes.     

Dismicrobism in inflammatory bowel disease and colorectal cancer: Changes in response of colocytes

Patients with inflammatory bowel disease (IBD) have an increased risk of 10%-15% developing colorectal cancer (CRC) that is a common disease of high economic costs in developed countries. The CRC has been increasing in recent years and its mortality rates are very high. Multiple biological and biochemical factors are responsible for the onset and progression of this pathology. Moreover, it appears absolutely necessary to investigate the environmental factors favoring the onset of CRC and the promotion of colonic health. The gut microflora, or microbiota, has an extensive diversity both quantitatively and qualitatively. In utero, the intestine of the mammalian fetus is sterile. At birth, the intestinal microbiota is acquired by ingesting maternal anal or vaginal organisms, ultimately developing into a stable community, with marked variations in microbial composition between individuals. The development of IBD is often associated with qualitative and quantitative disorders of the intestinal microbial flora (dysbiosis). The healthy human gut harbours about 10 different bacterial species distributed in colony forming units which colonize the gastrointestinal tract. The intestinal microbiota plays a fundamental role in health and in the progression of diseases such as IBD and CRC. In healthy subjects, the main control of intestinal bacterial colonization occurs through gastric acidity but other factors such as endoluminal temperature, competition between different bacterial strains, peristalsis and drugs can influence the intestinal microenvironment. The microbiota exerts diverse physiological functions to include: growth inhibition of pathogenic microorganisms, synthesis of compounds useful for the trophism of colonic mucosa, regulation of intestinal lymphoid tissue and synthesis of amino acids. Furthermore, mucus seems to play an important role in protecting the intestinal mucosa and maintaining its integrity. Changes in the microbiota composition are mainly influenced by diet and age, as well as genetic factors. Increasing evidence indicates that dysbiosis favors the production of genotoxins and metabolites associated with carcinogenesis and induces dysregulation of the immune response which promotes and sustains inflammation in IBD leading to carcinogenesis. A disequilibrium in gut microflora composition leads to the specific activation of gut associated lymphoid tissue. The associated chronic inflammatory process associated increases the risk of developing CRC. Ulcerative colitis and Crohn’s disease are the two major IBDs characterized by an early onset and extraintestinal manifestations, such as rheumatoid arthritis. The pathogenesis of both diseases is complex and not yet fully known. However, it is widely accepted that an inappropriate immune response to microbial flora can play a pivotal role in IBD pathogenesis.     

Location-specific effect of microbiota and MyD88-dependent signaling on Wnt/β-catenin pathway and intestinal stem cells

Intestinal homeostasis depends on the proper activity of the intestinal stem cells (ISCs) and an appropriate host response to the normal resident microbiota. The question on the effect of microbiota on ISCs behavior has not been addressed yet. Canonical Wnt pathway and ISC gene expression signature was compared in germfree vs. conventional and MyD88^−/− vs. Myd88^+/+ mice based on publicly available gene expression data sets. Microbiota and MyD88-dependent signaling have distinct effects on the Wnt pathway and ISC at gene expression level. In addition, the effect of microbiota and MyD88-dependent signaling on Wnt pathway and ISCs show regional variation. The net effect of microbiota on Wnt pathway and ISCs cannot be inferred from the available data. Nonetheless, the data are suggestive of a potential regulatory mechanism of the Wnt pathway by the microbiota and plausibly by any alteration in the microbiota composition.     

Serum vitamin D and colonic vitamin D receptor in inflammatory bowel disease

AIM: To determine serum vitamin D levels and colonic vitamin D receptor(VDR) expression in inflammatory bowel disease(IBD) and non-IBD patients and correlate these with histopathology.METHODS: Puerto Rican IBD(n = 10) and non-IBD(n = 10) patients ≥ 21 years old scheduled for colonoscopy were recruited. Each patient completed a questionnaire and provided a serum sample and a colonic biopsy of normal-appearing mucosa. For IBD patients, an additional biopsy was collected from visually diseased mucosa. Serum vitamin D levels were measured by ultra-performance liquid chromatography and mass spectrometry. Hematoxylin and eosin stained tissue sections from colonic biopsies were classified histologically as normal or colitis(active/inactive), and scored for the degree of inflammation present(0-3, inactive/absent to severe). Tissue sections from colonic biopsies were also stained by immunohistochemistry for VDR, for which representative diagnostic areas were photographed and scored for staining intensity using a 4-point scale.RESULTS: The IBD cohort was significantly younger(40.40 ± 5.27, P 0.05) than the non-IBD cohort(56.70 ± 1.64) with a higher prevalence of vitamin D deficiency(40% vs 20%, respectively) and insufficiency(70% vs 50%, respectively). Histologic inflammation was significantly higher in visually diseased mucosa from IBD patients(1.95 ± 0.25) than in normalappearing mucosa from control patients(0.25 ± 0.08, P 0.01) and from IBD patients(0.65 ± 0.36, P 0.05) and correlated inversely with VDR expression in visually diseased colonic tissue from IBD patients(r =-0.44, P 0.05) and from IBD patients with Crohn's disease(r =-0.69, P 0.05), but not in normal-appearing colonic tissue from control patients or IBD patients. Control and IBD patient serum vitamin D levels correlated positively with VDR expression in normal colon from control and IBD patients(r = 0.38, P 0.05) and with patient age(r = 0.54, P 0.01). CONCLUSION: Levels of serum vitamin D correlate positively with colonic VDR expression in visually normal mucosa whereas inflammation correlates negatively with colonic VDR expression in visually diseased mucosa in Puerto Rican patients.     

A decrease in anaerobic bacteria promotes <Emphasis Type="Italic">Candida glabrata</Emphasis> overgrowth while β-glucan treatment restores the gut microbiota and attenuates colitis

Background

The intestinal microbiota plays a crucial role in the maintenance of gut homeostasis. Changes in crosstalk between the intestinal epithelial cells, immune cells and the microbiota are critically involved in the development of inflammatory bowel disease. In the experimental mouse model, the development of colitis induced by dextran sulfate sodium (DSS) promotes overgrowth of the opportunistic yeast pathogen Candida glabrata. Conversely, fungal colonization aggravates inflammatory parameters. In the present study, we explored the effect of C. glabrata colonization on the diversity of the gut microbiota in a DSS-induced colitis model, and determined the impact of soluble β-glucans on C. glabrata-host interactions.

Results

Mice were administered a single inoculum of C. glabrata and were exposed to DSS treatment for 2 weeks in order to induce acute colitis. For β-glucan treatment, mice were administered with soluble β-glucans purified from C. glabrata (3?mg per mouse), orally and daily, for 5 days, starting on day 1. The number of C. glabrata colonies and changes in microbiota diversity were assessed in freshly collected stool samples from each tagged mouse, using traditional culture methods based on agar plates. An increase in Escherichia coli and Enterococcus faecalis populations and a reduction in Lactobacillus johnsonii and Bacteroides thetaiotaomicron were observed during colitis development. This decrease in L. johnsonii was significantly accentuated by C. glabrata overgrowth. Oral administration of β-glucans to mice decreased the overgrowth of aerobic bacteria and IL-1β expression while L. johnsonii and B. thetaiotaomicron populations increased significantly. β-glucan treatment increased IL-10 production via PPARγ sensing, promoting the attenuation of colitis and C. glabrata elimination.

Conclusions

This study shows that the colonic inflammation alters the microbial balance, while β-glucan treatment increases the anaerobic bacteria and promotes colitis attenuation and C. glabrata elimination.