A human colonic crypt culture system to study regulation of stem cell-driven tissue renewal and physiological function.

摘要: The intestinal epithelium is one of the most rapidly renewing tissues in human body and fulfils vital physiological roles such as barrier function transport nutrients fluid. Investigation gut epithelial physiology health disease has been hampered by lack ex vivo models native epithelium. Recently, remarkable progress made defining stem cells generating organoid cultures. In parallel, we have developed a 3D culture system colonic that recapitulates topological hierarchy cell-driven tissue renewal permits study polarized cells. Here describe methods to establish cultures intact crypts conduct real-time imaging renewal, cellular signalling, function, conjunction with manipulation gene expression lentiviral or adenoviral transduction. Visualization mRNA- protein-expression patterns cultured crypts, cross-validation derived from fixed mucosal biopsies, also described. Alongside studies using organoids, near-native crypt model will help bridge gap exists between investigation colon cancer cell lines and/or animal (tissue) studies, progression clinical trials. To this end, near provides platform aid development novel strategies for prevention inflammatory bowel cancer.