Establishment of Epithelial Inflammatory Injury Model Using Intestinal Organoid Cultures.

Intestinal epithelial dysfunction is critical in the development of inflammatory bowel disease (IBD). However, most cellular experiments related to epithelial barrier studies in IBD have been based on tumor cell line that lack a variety of intestinal epithelial cell types. Thus, intestinal organoids can present the three-dimensional structure and better simulate the physiological structure and function of the intestinal epithelium in vitro. Here, the crypts were isolated from the small intestine of mice; with the participation of major cytokines (EGF, Noggin, and R-Spondin 1 included), the intestinal organoids were established at a density of 100 crypts per well, containing intestinal stem cells (ISC), Paneth cells, goblet cells, and intestinal endocrine cells. We found that tumor necrosis factor-alpha (TNF-α) could induce the inflammatory response of intestinal organoids, and a dose of 10 ng/mL could maintain stable passaging of organoids for dynamic observation. After stimulation with TNF-α, the intestinal organoid cultures showed lower expression of the cell proliferation-related protein identified by monoclonal antibody Ki 67 (Ki67), the ISC marker leucine-rich repeat-containing G protein-coupled receptor 5 (Lgr5), and the intestinal tight junction proteins occludin (Ocln) and claudin-1 (Cldn1) while higher expression of the inflammatory cytokine interleukin- (IL-) 15 and the chemokines C-X-C motif ligand 2 (Cxcl2) and Cxcl10 significantly. In this study, we successfully established an epithelial inflammatory injury model of intestinal organoids, which provides an effective in vitro model for studying the pathogenesis and treatment of IBD.

The Distinguishing Bacterial Features From Active and Remission Stages of Ulcerative Colitis Revealed by Paired Fecal Metagenomes.

Ulcerative colitis (UC) is a serious chronic intestinal inflammatory disease, with an increased incidence in recent years. The intestinal microbiota plays a key role in the pathogenesis of UC. However, there is no unified conclusion on how the intestinal microbiota changes. Most studies focus on the change between UC patients and healthy individuals, rather than the active and remission stage of the same patient. To minimize the influences of genetic differences, environmental and dietary factors, we studied the intestinal microbiota of paired fecal samples from 42 UC patients at the active and remission stages. We identified 175 species of microbes from 11 phyla and found no difference of the alpha and beta diversities between the active and remission stages. Paired t-test analysis revealed differential microbiota at levels of the phyla, class, order, family, genus, and species, including 13 species with differential abundance. For example, CAG-269 sp001916005, Eubacterium F sp003491505, Lachnospira sp000436475, et al. were downregulated in the remission, while the species of Parabacteroides distasonis, Prevotellamassilia sp900540885, CAG-495 sp001917125, et al. were upregulated in the remission. The 13 species can effectively distinguish the active and remission stages. Functional analysis showed that the sporulation and biosynthesis were downregulated, and the hydrogen peroxide catabolic process was upregulated in remission of UC. Our study suggests that the 13 species together may serve as a biomarker panel contributing to identify the active and remission stages of UC, which provides a valuable reference for the treatment of UC patients by FMT or other therapeutic methods.