Establishment of Epithelial Inflammatory Injury Model Using Intestinal Organoid Cultures

Abstract

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.

Exploring the Potential Links Between Gut Bacteria-Related Metabolites and the Risk of Cardiovascular Diseases

“…Epidemiological data has established a link between the consumption of high-calorie, low-fiber, high-monosaccharide, and high-fat diets and the prevalence of several chronic diseases with high morbidity and mortality rates in modern Western 2 societies, including type II diabetes, IBD, and colorectal cancer. Notably, there is accumulating evidence suggesting that IBD in canines and humans share analogous clinical and molecular features (Xing et al, 2023). Consequently, preclinical studies involving dogs with naturally occurring IBD present a valuable opportunity to deepen our understanding of disease pathogenesis and pioneer new therapeutic strategies.…”

A Strainer-Based Platform for the Collection and Immunolabeling of Porcine Epidemic Diarrhea Virus-Infected Porcine Intestinal Organoid

“…Many studies have indicated that intestinal organoids are a potential model of inflammatory bowel disease, intestinal cancer, and other digestive diseases [15][16][17]. Intestinal organoids have more advantages than other intestinal disease models (cell lines or laboratory animals), and these advantages include a more complete cell system of the gut (simulating the physiological structure and function of the enteric canal) than single-cell lines, a shorter growth cycle, and smaller individual differences than laboratory animals [18,19]. The experimental techniques for intestinal organoids have been gradually systematized, providing a new platform for studying intestinal diseases.…”