Rigorous Donor Selection for Fecal Microbiota Transplantation in Active Ulcerative Colitis: Key Lessons From a Randomized Controlled Trial Halted for Futility.

BACKGROUND & AIMS: Rigorous donor preselection on microbiota level, strict anaerobic processing, and repeated fecal microbiota transplantation (FMT) administration were hypothesized to improve FMT induction of remission in ulcerative colitis (UC). METHODS: The RESTORE-UC trial was a multi-centric, double-blind, sham-controlled, randomized trial. Patients with moderate to severe UC (defined by total Mayo 4-10) were randomly allocated to receive 4 anaerobic-prepared allogenic or autologous donor FMTs. Allogenic donor material was selected after a rigorous screening based on microbial cell count, enterotype, and the abundance of specific genera. The primary endpoint was steroid-free clinical remission (total Mayo ≤2, no sub-score >1) at week 8. A pre-planned futility analysis was performed after 66% (n = 72) of intended inclusions (n = 108). Quantitative microbiome profiling (n = 44) was performed at weeks 0 and 8. RESULTS: In total, 72 patients were included, of which 66 received at least 1 FMT (allogenic FMT, n = 30 and autologous FMT, n = 36). At week 8, respectively, 3 and 5 patients reached the primary endpoint of steroid-free clinical remission (P = .72), indicating no treatment difference of at least 5% in favor of allogenic FMT. Hence, the study was stopped due to futility. Microbiome analysis showed numerically more enterotype transitions upon allogenic FMT compared with autologous FMT, and more transitions were observed when patients were treated with a different enterotype than their own at baseline (P = .01). Primary response was associated with lower total Mayo scores, lower bacterial cell counts, and higher Bacteroides 2 prevalence at baseline. CONCLUSION: The RESTORE-UC trial did not meet its primary endpoint of increased steroid-free clinical remission at week 8. Further research should additionally consider patient selection, sterilized sham-control, increased frequency, density, and viability of FMT prior to administration. CLINICALTRIALS: gov, Number: NCT03110289.

Enhancing probiotic impact: engineering Saccharomyces boulardii for optimal acetic acid production and gastric passage tolerance.

Saccharomyces boulardii has been a subject of growing interest due to its potential as a probiotic microorganism with applications in gastrointestinal health, but the molecular cause for its probiotic potency has remained elusive. The recent discovery that S. boulardii contains unique mutations causing high acetic acid accumulation and inhibition of bacterial growth provides a possible clue. The natural S. boulardii isolates Sb.P and Sb.A are homozygous for the recessive mutation whi2S270* and accumulate unusually high amounts of acetic acid, which strongly inhibit bacterial growth. However, the homozygous whi2S270* mutation also leads to acetic acid sensitivity and acid sensitivity in general. In the present study, we have constructed a new S. boulardii strain, derived from the widely therapeutically used CMCN I-745 strain (isolated from the pharmaceutical product Enterol), producing even higher levels of acetic acid while keeping the same tolerance toward low pH as the parent Enterol (ENT) strain. This newly engineered strain, named ENT3, has a homozygous deletion of ACH1 and strong overexpression of ALD4. It is also able to accumulate much higher acetic acid concentrations when growing on low glucose levels, in contrast to the ENT wild-type and Sb.P strains. Moreover, we show the antimicrobial capacity of ENT3 against gut pathogens in vitro and observed that higher acetic acid production might correlate with better persistence in the gut in healthy mice. These findings underscore the possible role of the unique acetic acid production and its potential for improvement of the probiotic action of S. boulardii.IMPORTANCESuperior variants of the probiotic yeast Saccharomyces boulardii produce high levels of acetic acid, which inhibit the growth of bacterial pathogens. However, these strains also show increased acid sensitivity, which can compromise the viability of the cells during their passage through the stomach. In this work, we have developed by genetic engineering a variant of Saccharomyces boulardii that produces even higher levels of acetic acid and does not show enhanced acid sensitivity. We also show that the S. boulardii yeasts with higher acetic acid production persist longer in the gut, in agreement with a previous work indicating competition between probiotic yeast and bacteria for residence in the gut.

The first international Rome consensus conference on gut microbiota and faecal microbiota transplantation in inflammatory bowel disease.

BACKGROUND: Several randomised clinical trials (RCTs) performing faecal microbiota transplantation (FMT) for the management of inflammatory bowel disease (IBD), particularly for ulcerative colitis, have recently been published, but with major variations in study design. These include differences in administered dose, route and frequency of delivery, type of placebo and evaluated endpoints. Although the overall outcomes appear to be promising, they are highly dependent on both donor and recipient factors. OBJECTIVE: To develop concensus-based statements and recommendations for the evaluation, management and potential treatment of IBD using FMT in order to move towards standardised practices. DESIGN: An international panel of experts convened several times to generate evidence-based guidelines by performing a deep evaluation of currently available and/or published data. Twenty-five experts in IBD, immunology and microbiology collaborated in different working groups to provide statements on the following key issues related to FMT in IBD: (A) pathogenesis and rationale, (B) donor selection and biobanking, (C) FMT practices and (D) consideration of future studies and perspectives. Statements were evaluated and voted on by all members using an electronic Delphi process, culminating in a plenary consensus conference and generation of proposed guidelines. RESULTS AND CONCLUSIONS: Our group has provided specific statements and recommendations, based on best available evidence, with the end goal of providing guidance and general criteria required to promote FMT as a recognised strategy for the treatment of IBD.

High Acetate Concentration Protects Intestinal Barrier and Exerts Anti-Inflammatory Effects in Organoid-Derived Epithelial Monolayer Cultures from Patients with Ulcerative Colitis.

Short-chain fatty acids as well as their bacterial producers are of increasing interest in inflammatory bowel diseases. Although less studied compared to butyrate, acetate might also be of interest as it may be less toxic to epithelial cells, stimulate butyrate-producing bacteria by cross-feeding, and have anti-inflammatory and barrier-protective properties. Moreover, one of the causative factors of the probiotic potency of Saccharomyces cerevisae var. boulardii is thought to be its high acetate production. Therefore, the objective was to preclinically assess the effects of high acetate concentrations on inflammation and barrier integrity in organoid-based monolayer cultures from ulcerative colitis patients. Confluent organoid-derived colonic epithelial monolayers (n = 10) were exposed to basolateral inflammatory stimulation or control medium. After 24 h, high acetate or control medium was administered apically for an additional 48 h. Changes in TEER were measured after 48 h. Expression levels of barrier genes and inflammatory markers were determined by qPCR. Pro-inflammatory proteins in the supernatant were quantified using the MSD platform. Increased epithelial resistance was observed with high acetate administration in both inflamed and non-inflamed conditions, together with decreased expression levels of IL8 and TNFα and CLDN1. Upon high acetate administration to inflamed monolayers, upregulation of HIF1α, MUC2, and MKI67, and a decrease of the majority of pro-inflammatory cytokines was observed. In our patient-derived human epithelial cell culture model, a protective effect of high acetate administration on epithelial resistance, barrier gene expression, and inflammatory protein production was observed. These findings open up new possibilities for acetate-mediated management of barrier defects and inflammation in IBD.

Eosinophil Depletion as a Potential Therapeutic Strategy in Acute and Chronic Intestinal Inflammation Based on a Dextran Sulfate Sodium Colitis Model.

BACKGROUND: A role for eosinophils in intestinal inflammation and fibrosis in the context of inflammatory bowel disease has been suggested, yet the precise nature, whether causal or secondary remains debated. Hence, it remains unclear whether targeting eosinophils should be further explored as a treatment option in inflammatory bowel disease. METHODS: Acute and chronic dextran sulfate sodium colitis was induced in wild-type C57BL/6 mice. Eosinophils were depleted by anti-CCR3 injections before colitis induction in a chronic model and after colitis onset in an acute model in order to investigate the impact of eosinophil depletion on pre-existing colitis. Inflammation was assessed using the disease activity index, macroscopic damage, and histological disease activity score. In the chronic model, fibrosis was assessed by examining colon weight/length ratio, collagen deposition through Martius Scarlet Blue staining, hydroxyproline assay, and COL1A1 expression. Protein and gene expression were assessed using the Meso Scale Discovery platform and real-time quantitative polymerase chain reaction. RESULTS: In the acute and chronic colitis model, eosinophil depletion resulted in reduced disease activity and faster recovery, as observed via the total area under the curve of the disease activity index (P = .004 and P = .02, respectively), macroscopic damage score (P = .009 and P = .08, respectively), and histological disease activity score (P = .09 and P = .002, respectively). In the acute model, the accelerated recovery was accompanied by an increase in interleukin (IL)-10 (P = .03) and a decrease in IL-4 (P = .03) and IL-6 (P = .009). Colon weight/length ratio and collagen deposition were not affected by eosinophil depletion. CONCLUSIONS: Eosinophil depletion prevents and decreases intestinal inflammation in a preclinical dextran sulfate sodium model without affecting fibrosis. These results pave the way for exploring eosinophil depletion as a novel treatment modality in addressing intestinal inflammation.

This study investigated the effect of eosinophil depletion on intestinal inflammation and fibrosis in a dextran sulfate sodium colitis model. Our findings suggest a proinflammatory role for eosinophils and therefore warrant further investigation into eosinophil targeting as a treatment option for inflammatory bowel disease patients.

Effect of Mutant and Engineered High-Acetate-Producing Saccharomyces cerevisiae var. boulardii Strains in Dextran Sodium Sulphate-Induced Colitis.

Acetate-producing Saccharomyces cerevisiae var. boulardii strains could exert improved effects on ulcerative colitis, which here, was preclinically evaluated in an acute dextran sodium sulphate induced model of colitis. Nine-week-old female mice were divided into 12 groups, receiving either drinking water or 2.75% dextran sodium sulphate for 7 days, combined with a daily gavage of various treatments with different levels of acetate accumulation: sham control (phosphate buffered saline, no acetate), non-probiotic control (Baker's yeast, no acetate), probiotic control (Enterol®, transient acetate), and additionally several Saccharomyces cerevisiae var. boulardii strains with respectively no, high, and extra-high acetate accumulation. Disease activity was monitored daily, and feces samples were collected at different timepoints. On day 14, the mice were sacrificed, upon which blood and colonic tissue were collected for analysis. Disease activity in inflamed mice was lower when treated with the high-acetate-producing strain compared to sham and non-probiotic controls. The non-acetate-producing strain showed higher disease activity compared to the acetate-producing strains. Accordingly, higher histologic inflammation was observed in non- or transient-acetate-producing strains compared to the sham control, whereas this increase was not observed for high- and extra-high-acetate-producing strains upon induction of inflammation. These anti-inflammatory findings were confirmed by transcriptomic analysis of differentially expressed genes. Moreover, only the strain with the highest acetate production was superior in maintaining a stable gut microbial alpha-diversity upon inflammation. These findings support new possibilities for acetate-mediated management of inflammation in inflammatory bowel disease by administrating high-acetate-producing Saccharomyces cerevisae var. boulardii strains.

Short chain fatty acids and its producing organisms: An overlooked therapy for IBD?

The gut microbiome and the intestinal immune system are driving contributors to inflammatory bowel diseases (IBD). Both have an important signalling factor in common: short-chain fatty acids (SCFAs). SCFAs (acetate, propionate and butyrate) are produced by bacterial fermentation in the gut and exert several effects on host metabolism and immune system. This review provides an overview of the current knowledge of these effects, with specific focus on energy metabolism, intestinal barrier, immune system, and disease activity in IBD. To conclude, more research is needed on the cross-feeding mechanisms in the gut microbiome, as well as on the therapeutic potential of SCFAs on different disease models. Also randomized controlled trials and prospective cohort studies should investigate the clinical impact of SCFA administration.