The two primary human inflammatory bowel diseases, Crohn's disease (CD) and ulcerative colitis (UC), are idiopathic relapsing disorders characterized by chronic inflammation of the intestinal tract. Although several lines of reasoning suggest that gastrointestinal (GI) microbes influence inflammatory bowel disease (IBD) pathogenesis, the types of microbes involved have not been adequately described. Here we report the results of a culture-independent rRNA sequence analysis of GI tissue samples obtained from CD and UC patients, as well as non-IBD controls. Specimens were obtained through surgery from a variety of intestinal sites and included both pathologically normal and abnormal states. Our results provide comprehensive molecular-based analysis of the microbiota of the human small intestine. Comparison of clone libraries reveals statistically significant differences between the microbiotas of CD and UC patients and those of non-IBD controls. Significantly, our results indicate that a subset of CD and UC samples contained abnormal GI microbiotas, characterized by depletion of commensal bacteria, notably members of the phyla Firmicutes and Bacteroidetes. Patient stratification by GI microbiota provides further evidence that CD represents a spectrum of disease states and suggests that treatment of some forms of IBD may be facilitated by redress of the detected microbiological imbalances.

https://www.pnas.org/content/pnas/104/34/13780.full.pdf

Molecular-phylogenetic characterization of microbial community imbalances in human inflammatory bowel diseases

A decrease in the abundance and biodiversity of intestinal bacteria within the dominant phylum Firmicutes has been observed repeatedly in Crohn disease (CD) patients. In this study, we determined the composition of the mucosa-associated microbiota of CD patients at the time of surgical resection and 6 months later using FISH analysis. We found that a reduction of a major member of Firmicutes, Faecalibacterium prausnitzii, is associated with a higher risk of postoperative recurrence of ileal CD. A lower proportion of F. prausnitzii on resected ileal Crohn mucosa also was associated with endoscopic recurrence at 6 months. To evaluate the immunomodulatory properties of F. prausnitzii we analyzed the anti-inflammatory effects of F. prausnitzii in both in vitro (cellular models) and in vivo [2,4,6-trinitrobenzenesulphonic acid (TNBS)-induced] colitis in mice. In Caco-2 cells transfected with a reporter gene for NF-kappaB activity, F. prausnitzii had no effect on IL-1beta-induced NF-kappaB activity, whereas the supernatant abolished it. In vitro peripheral blood mononuclear cell stimulation by F. prausnitzii led to significantly lower IL-12 and IFN-gamma production levels and higher secretion of IL-10. Oral administration of either live F. prausnitzii or its supernatant markedly reduced the severity of TNBS colitis and tended to correct the dysbiosis associated with TNBS colitis, as demonstrated by real-time quantitative PCR (qPCR) analysis. F. prausnitzii exhibits anti-inflammatory effects on cellular and TNBS colitis models, partly due to secreted metabolites able to block NF-kappaB activation and IL-8 production. These results suggest that counterbalancing dysbiosis using F. prausnitzii as a probiotic is a promising strategy in CD treatment.

/pdf/faecalibacterium-prausnitzii-is-an-anti-inflammatory-389ys91ar1.pdf

Faecalibacterium prausnitzii is an anti-inflammatory commensal bacterium identified by gut microbiota analysis of Crohn disease patients

We are facing a global metabolic health crisis provoked by an obesity epidemic. Here we report the human gut microbial composition in a population sample of 123 non-obese and 169 obese Danish individuals. We find two groups of individuals that differ by the number of gut microbial genes and thus gut bacterial richness. They contain known and previously unknown bacterial species at different proportions; individuals with a low bacterial richness (23% of the population) are characterized by more marked overall adiposity, insulin resistance and dyslipidaemia and a more pronounced inflammatory phenotype when compared with high bacterial richness individuals. The obese individuals among the lower bacterial richness group also gain more weight over time. Only a few bacterial species are sufficient to distinguish between individuals with high and low bacterial richness, and even between lean and obese participants. Our classifications based on variation in the gut microbiome identify subsets of individuals in the general white adult population who may be at increased risk of progressing to adiposity-associated co-morbidities.

/pdf/richness-of-human-gut-microbiome-correlates-with-metabolic-1i65keoatj.pdf

Richness of human gut microbiome correlates with metabolic markers

We normally live in symbiosis with approximately 10(13) bacteria present in the colon. Among the several mechanisms maintaining the bacteria/host balance, there is limited understanding of the structure, function, and properties of intestinal mucus. We now demonstrate that the mouse colonic mucus consists of two layers extending 150 mum above the epithelial cells. Proteomics revealed that both of these layers have similar protein composition, with the large gel-forming mucin Muc2 as the major structural component. The inner layer is densely packed, firmly attached to the epithelium, and devoid of bacteria. In contrast, the outer layer is movable, has an expanded volume due to proteolytic cleavages of the Muc2 mucin, and is colonized by bacteria. Muc2(-/-) mice have bacteria in direct contact with the epithelial cells and far down in the crypts, explaining the inflammation and cancer development observed in these animals. These findings show that the Muc2 mucin can build a mucus barrier that separates bacteria from the colon epithelia and suggest that defects in this mucus can cause colon inflammation.

https://www.pnas.org/content/pnas/105/39/15064.full.pdf

The inner of the two Muc2 mucin-dependent mucus layers in colon is devoid of bacteria

Microbial Influences in Inflammatory Bowel Diseases

The composition and spatial organization of the mucosal flora in biopsy specimens from patients with inflammatory bowel disease (IBD; either Crohn's disease or ulcerative colitis), self-limiting colitis, irritable-bowel syndrome (IBS), and healthy controls were investigated by using a broad range of fluorescent bacterial group-specific rRNA-targeted oligonucleotide probes. Each group included 20 subjects. Ten patients who had IBD and who were being treated with antibiotics were also studied. Use of nonaqueous Carnoy fixative to preserve the mucus layer was crucial for detection of bacteria adherent to the mucosal surface (mucosal bacteria). No biofilm was detectable in formalin-fixed biopsy specimens. Mucosal bacteria were found at concentrations greater than 109/ml in 90 to 95% of IBD patients, 95% of patients with self-limiting colitis, 65% of IBS patients, and 35% of healthy controls. The mean density of the mucosal biofilm was 2 powers higher in IBD patients than in patients with IBS or controls, and bacteria were mostly adherent. Bacteroides fragilis was responsible for >60% of the biofilm mass in patients with IBD but for only 30% of the biofilm mass in patients with self-limiting colitis and 40% of the biofilm in IBS patients but for <15% of the biofilm in IBD patients. In patients treated with (5-ASA) or antibiotics, the biofilm could be detected with 4,6-diamidino-2-phenylindole but did not hybridize with fluorescence in situ hybridization probes. A Bacteroides fragilis biofilm is the main feature of IBD. This was not previously recognized due to a lack of appropriate tissue fixation. Both 5-ASA and antibiotics suppress but do not eliminate the adherent biofilm.

Spatial organization and composition of the mucosal flora in patients with inflammatory bowel disease.

Background: Wireless capsule endoscopy (WCE) offers endoscopic access to the small bowel and may therefore change diagnostic and therapeutic strategies in small bowel diseases. Aim: The aim of this prospective study was to validate the gain in information and therapeutic impact of WCE in patients with Crohn’s disease. Methods: Fifty six consecutive patients with Crohn’s disease underwent computed tomography (CT) enteroclysis, and if stenoses Results: In 15 patients (27%), WCE could not be performed due to strictures detected by CT enteroclysis. From the other 41 patients, jejunal or ileal lesions were found in 25 patients by WCE compared with 12 by CT enteroclysis (p = 0.004). This gain in information was mainly due to detection of small mucosal lesions such as villous denudation, aphthoid ulcerations, or erosions. Both methods were not significantly different in the detection of lesions in the terminal/neoterminal ileum (WCE 24 patients, CT enteroclysis 20 patients). Therapy was changed due to WCE findings in 10 patients. Consecutively, all of them improved clinically. Conclusions: Capsule endoscopy improves the diagnosis of small bowel Crohn’s disease. This may have significant therapeutic impact.

https://gut.bmj.com/content/gutjnl/54/3/369.full.pdf

Small bowel involvement in Crohn’s disease: a prospective comparison of wireless capsule endoscopy and computed tomography enteroclysis

Background: Migration is an important virulence factor for intestinal bacteria. However, the role of bacterial mobility in the penetration of viscous mucus and their spatial organization within the colon is relatively unknown. Methods: Movements of fecal bacteria were assessed in gels of varying agarose concentrations and were compared with patterns of bacterial distribution observed in colons from conventional and Enterobacter cloacae-monoassociated mice. Bacteria were visualized using fluorescence in situ hybridization. Results: Long curly bacteria moved best in moderate viscosity gels, short rods and cocci preferred a low viscous environment, whereas high viscosity immobilized all bacterial groups. The spatial distribution of bacteria in the murine colon was also shape- and not taxonomy-dependent, indicating the existence of vertical (surface to lumen) and longitudinal (proximal to distal colon) viscosity gradients within the mucus layer. Our results suggest that mucus viscosity is low in goblet cells, at the crypt basis and close to the intestinal lumen, whereas sites adjacent to the columnar epithelium have a high mucus viscosity. The mucus viscosity increased progressively toward the distal colon, separating bacteria selectively in the proximal colon and completely in the distal colon. Conclusions: The site-specific regulation of mucus secretion and dehydration make the mucus layer firm and impenetrable for bacteria in regions close to the intestinal mucosa but loose and lubricating in regions adjacent to the luminal contents. Selective control of mucus secretion and dehydration may prove to be a key factor in the management of chronic diseases in which intestinal pathogens are involved. (Inflamm Bowel Dis 2007;13:963–970)

Viscosity gradient within the mucus layer determines the mucosal barrier function and the spatial organization of the intestinal microbiota

Background: Thalidomide improves clinical symptoms in patients with therapy refractory Crohn's disease, as shown in two recent studies. The mechanism of this effect however is still unknown. Suppression of tumour necrosis factor α (TNF-α) by thalidomide has been suggested as a possible mechanism. However, effects on other cytokines have not been adequately investigated.

Aim: The aim of our study was to investigate the effects of thalidomide on cytokine production in patients with inflammatory bowel disease (IBD).

Methods: Ten patients with therapy refractory IBD (nine Crohn's disease, one ulcerative colitis) received thalidomide 300 mg daily in a 12 week open label study. Production of TNF-α, interleukin (IL)-1β, IL-6, and IL-12 was investigated in short term cultures of stimulated colonic lamina propria mononuclear cells (LPMC) and peripheral blood monocytes (PBMC) before and after 12 weeks of treatment. LPMC were also cultured with graded doses of thalidomide.

Results: Three patients discontinued treatment because of sedative side effects. In the other patients, disease activity decreased significantly, with four patients achieving remission. Production of TNF-α and IL-12 decreased during treatment with thalidomide: LPMC (TNF-α: 42.3 (8.3) pg/ml v 16.4 (6.3); IL-12: 9.7 (3.3) v 5.0 (2.5); p<0.04) and PBMC (TNF-α: 62.8 (14.6) v 22.5 (9.2); p<0.02). Production of IL-1β and IL-6 did not change significantly. Culturing of LPMC with thalidomide showed a dose dependent decrease in TNF-α and IL-12 production.

Conclusion: The clinical effects of thalidomide in Crohn's disease may be mediated by reduction of both TNF-α and IL-12.

https://gut.bmj.com/content/gutjnl/50/2/196.full.pdf

Thalidomide reduces tumour necrosis factor alpha and interleukin 12 production in patients with chronic active Crohn's disease.

Apart from its anti-inflammatory activity, which has been used for the treatment of active Crohn’s disease, thalidomide is also a potent inhibitor of angiogenesis. We therefore studied the effect of thalidomide in six patients with severe recurrent intestinal bleeding refractory to standard treatment (three patients with Crohn’s disease (CD), three patients with obscure intestinal bleeding; mean of 56 blood transfusions within the last 24 months). Bleeding stopped within two weeks after the start of thalidomide in all patients. Haemoglobin normalised without further transfusions for the whole observation period (mean follow up 33 months) while patients needed a mean of 2.2 (CD) and 3.1 (obscure bleeding) blood units/month in the 12 months before treatment. After three months of thalidomide therapy, serum levels of vascular endothelial growth factor were strongly suppressed compared with pretreatment levels. (CD 818 (82) v 129 (86) pg/ml; obscure bleeding 264 (68) v 50 (25) pg/ml). All six patients reported transient fatigue. Peripheral neuropathy was observed in one patient with CD after nine months and was reversible after lowering the dose to 100 mg daily. These results indicate that thalidomide might be useful for patients with otherwise refractory intestinal bleeding.

https://gut.bmj.com/content/gutjnl/53/4/609.full.pdf

H. Lochs

Papers

Spatial organization and composition of the mucosal flora in patients with inflammatory bowel disease.

Small bowel involvement in Crohn’s disease: a prospective comparison of wireless capsule endoscopy and computed tomography enteroclysis

Viscosity gradient within the mucus layer determines the mucosal barrier function and the spatial organization of the intestinal microbiota

Thalidomide reduces tumour necrosis factor alpha and interleukin 12 production in patients with chronic active Crohn's disease.

Thalidomide for treatment of severe intestinal bleeding