Whole genome characterization of in vitro generated mouse colonic monolayers
Ontology highlight
ABSTRACT:
PROVIDER: PRJNA478953 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR7465030.fastq.gz | Fastqsanger.gz | |||
| SRR7465031.fastq.gz | Fastqsanger.gz | |||
| SRR7465032.fastq.gz | Fastqsanger.gz | |||
| SRR7465033.fastq.gz | Fastqsanger.gz | |||
| SRR7465034.fastq.gz | Fastqsanger.gz |
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Project description:Dietary fibre usually comes associated with phenolic compounds either trapped in a network formed by indigestible polysaccharides or, in some cases, bound to this matrix by covalent bonds. This natural association protects some phenolic compounds during their journey through the gastrointestinal tract, reaching the colon, where most polysaccharides undergo fermentation, weakening the matrix and promoting the release of these compounds to exert benefits by modulating intestinal function. Unfortunately, there are still difficulties in investigating the impact of the human microbiota on the release of phenolic compounds from the indigestible fraction and on their effects on the intestinal epithelium. In this study, grape pomace skin was subjected to in vitro digestion to obtain an indigestible fraction, which was further incorporated into SHIME bioreactors with human colonic microbiota. Throughout these stages, we monitored the release of phenolic compounds using HPLC-DAD. We determined the impact of a phenolic-rich indigestible fraction on the fermentative capacity of human microbiota by measuring the short-chain fatty acids (SCFA) levels and the effect of the resulting colonic ferments on the barrier properties of differentiated monolayers of Caco-2 cells by monitoring the transepithelial electrical resistance and the transcriptomic profile by RNA sequencing.
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Project description:Background: The Inflammatory Bowel Disease (IBD) ulcerative colitis (UC) is characterized by colonic mucosal inflammation and barrier dysfunction. We hypothesize that UC causes persistent defects in mucosal homeostasis, evident even in the absence of active inflammation, contributing to disease chronicity. Methods: To test this, we grew patient biopsy-derived sigmoid colonoids into air-liquid interface (ALI) monolayers, characterizing them through microscopy, proteomics, bulk RNAseq, and their susceptibility to UC patient-isolated Escherichia coli pathobiont p19A. Results: Non-IBD ALI monolayers formed uniform crypt-like structures and a thick mucus layer containing all epithelial-derived proteins previously identified in human colonic mucus. In contrast, ALI monolayers from UC patients displayed a range of impairments, classified as a mild phenotype with distorted architecture and a thinner, more permeable mucus layer to a severe phenotype with defects in cellular differentiation and an inability to produce a mucus layer. Transcriptome analysis identified activated pathways associated with extracellular matrix formation and cell signaling, including numerous cancer-associated genes in UC ALI monolayers, which also proved significantly more susceptible to E. coli p19A. Conclusions: Taken together, the culturing of patient biopsies into ALI colonoid monolayers provides a powerful model to assess human colonic mucosal development, healing, homeostasis, and mucus barrier function, revealing that UC-derived colonoid monolayers display a range of developmental and functional defects that persist in the absence of inflammation.
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