Abstract Background Crohn’s disease (CD) is a complex disease where the gut microbiome plays an important role. One of the established treatments in CD is Vedolizumab (VDZ), an alpha(4)beta(7) integrin antibody. Finding biomarkers to predict therapy response is still a clinical unmet need, since this treatment has shown endoscopic remission in only a third of CD patients. Ananthakrishnan et al. demonstrated a strong relationship between microbial metagenomics signature and therapy response to VDZ in CD based on the metagenomics composition of baseline fecal samples. While the majority of studies focus on fecal samples, mucosa-adherent bacterial signature could bring forth stable biomarkers. In this study, we sought to identify the signature of the adherent microbiome in intestinal biopies to differentiate responders (R) from non-responders (NR) to VDZ treatment at baseline. Methods We prospectively collected ileal and colonic biopsies from adult CD patients scheduled to start VDZ treatment during baseline- and follow-up (FU) endoscopies. After median 27 weeks of follow-up, patients were classified as either R or NR based on endoscopic response (≥50% reduction in SES-CD score), steroid-free clinical response (≥3 point drop in HBI or HBI ≤4, no systemic steroids) and/or biochemical response (≥50% reduction in C-reactive protein (CRP) and fecal calprotectin or a basal CRP ≤5 g/mL and fecal calprotectin ≤250 µg/g). Microbiome composition of the biopsies was determined using 16S RNA gene V3V4 amplicon sequencing. We measured alpha and beta diversity using Wilcoxon and Adonis metrics. Results In total, 44 CD patients were included in the baseline cohort (28 R and 16 NR) and 53 CD patients were included in the follow up cohort (37 R and 20 NR), for which 21 patients overlap between baseline and follow up. When comparing alpha-diversity between R and NR, we did not find significant differences in ileal (Wilcoxon, p=0.78) and colonic (Wilcoxon, p=0.70) samples at baseline nor at follow up (ileal Wilcoxon, p=0. 27 and colonic Wilcoxon, p=0. 63). Next, comparing the beta-diversity, we demonstrated no significant differences between R and NR in ileal (baseline p=0.96, follow up p=0.11) and colonic (baseline p=0.11, follow up p=0.4). Microbiome profiles did show high inter-individual variation but were highly similar intra-individually both between body site and over time. Conclusion Here, we investigated the microbial signature of VDZ R and NR and demonstrated mucosa-associated microbiome is mostly stable after resolution/non-resolution of inflammation in CD and does not predict response to VDZ therapy. Further analyses on data of infliximab, adalimumab and ustekinumab treated patients, as part of the EPIC Pioneer study, are ongoing.
Abstract Irritable bowel syndrome (IBS) is a common disorder characterized by chronic abdominal pain and changes in bowel movements. Visceral hypersensitivity is thought to be responsible for pain complaints in a subset of patients. In an IBS-like animal model, visceral hypersensitivity was triggered by intestinal fungi, and lower mycobiota α-diversity in IBS patients was accompanied by a shift toward increased presence of Candida albicans and Saccharomyces cerevisiae . Yet, this shift was observed in hypersensitive as well as normosensitive patients and diversity did not differ between IBS subgroups. The latter suggests that, when a patient changes from hyper- to normosensitivity, the relevance of intestinal fungi is not necessarily reflected in compositional mycobiota changes. We now confirmed this notion by performing ITS1 sequencing on an existing longitudinal set of fecal samples. Since ITS1 methodology does not recognize variations within species, we next focused on heterogeneity within cultured healthy volunteer and IBS-derived C. albicans strains. We observed inter- and intra-individual genomic variation and partial clustering of strains from hypersensitive patients. Phenotyping showed differences related to growth, yeast-to-hyphae morphogenesis and gene expression, specifically of the gene encoding fungal toxin candidalysin. Our investigations emphasize the need for strain-specific cause-and-effect studies within the realm of IBS research.
Recently, the first clinical trials on Bioartificial Livers (BALs) loaded with a proliferative human hepatocyte cell source have started.There are two cell lines that are currently in an advanced state of BAL development; HepaRG and HepG2/C3A.In this study we aimed to compare both cell lines on applicability in BALs and to identify possible strategies for further improvement.We tested both cell lines in monolayer-and BAL cultures on growth characteristics, hepatic differentiation, nitrogen-, carbohydrate-, amino acid-and xenobiotic metabolism.Interestingly, both cell lines adapted the hepatocyte phenotype more closely when cultured in BALs; e.g.monolayer cultures produced lactate, while BAL cultures showed diminished lactate production (C3A) or conversion to elimination (HepaRG), and urea cycle activity increased upon BAL culturing in both cell lines.HepaRG-BALs outperformed C3A-BALs on xenobiotic metabolism, ammonia elimination and lactate elimination, while protein synthesis was comparable.In BAL cultures of both cell lines ammonia elimination correlated positively with glutamine production and glutamate consumption, suggesting ammonia elimination was mainly driven by the balance between glutaminase and glutamine synthetase activity.Both cell lines lacked significant urea cycle activity and both required multiple culture weeks before reaching optimal differentiation in BALs.In conclusion, culturing in BALs enhanced hepatic functionality of both cell lines and from these, the HepaRG cells are the most promising proliferative cell source for BAL application.
View this article online at wileyonlinelibrary.com. Potential conflict of interest: Nothing to report. We thank Qvartskhava c.s. for drawing our attention to the different mouse lines used in their and our study and their potential effects on the resulting phenotypes. We opted for the Alfp‐Cre line with the three Afp enhancers1 to assure complete excision in all pericentral hepatocytes. Qvartskhava c.s. cite Pruneau et al.,2 who reported that Alfp‐Cre mice ectopically express human growth hormone in their pituitary and hypothalamus and suffer, as a result, from peripheral growth‐hormone deficiency, growth retardation, and a fatty liver. In their reply to Pruneau et al., Mueller and Moriggl argue that they did not observe growth retardation in their Stat5fl/+/Nr3c1fl/+/AlfpCre+/− mice and that hepatic steatosis is also observed in Stat5fl/fl/AlbCre+/− mice.3 Similarly, we did not observe growth retardation in Glul5fl/fl/AlfpCre+/− (our Fig. 1). Given that growth retardation only occurs if growth hormone, its receptor, or Stat5b are homozygously deficient (Teglund et al.4 and references therein), Pruneau c.s.2 probably studied AlfpCre+/+ rather than the AlfpCre+/− mice that are usually used for gene excision. Furthermore, we did not observe the pericentral macrovesicular fat deposition in Stat5–/– mice5 that we observed in Glul5fl/fl/AlfpCre+/− mice. Another important difference between the study of Qvartskhava c.s. and our study is that they used C57BL/6J mice, whereas we used FVB/N mice. Although circulating ammonia concentrations attributed to hepatic GS deficiency increased ∼1.7‐fold in both studies, absolute concentrations were ∼5‐fold higher in C57BL/6J than in FVB/N mice because of a deletion in the Nnt gene of C57BL/6J mice. It will be interesting to establish to what extent these substantial strain‐specific differences in background ammonia concentrations affect the adverse effects of hyperammonemia.
'%3e%3cpath fill='%23012169' d='M0 0v30h60V0z'/%3e%3cpath stroke='%23fff' stroke-width='6' d='m0 0 60 30m0-30L0 30'/%3e%3cpath stroke='%23C8102E' stroke-width='4' d='m0 0 60 30m0-30L0 30' clip-path='url(%23b)'/%3e%3cpath stroke='%23fff' stroke-width='10' d='M30 0v30M0 15h60'/%3e%3cpath stroke='%23C8102E' stroke-width='6' d='M30 0v30M0 15h60'/%3e%3c/g%3e%3c/svg%3e)