Establishment and Optimization of Culture Technique for Enteroids / 胃肠病学

ABSTRACT

Background:Enteroids are considered to be the best tool for studies on intestinal epithelium in vitro and have a widely application prospects,however,there are no associated reports in China. Aims:To establish and optimize the culture technique for enteroids and provide a fantastic platform for the basic research of small intestinal epithelial cells in China. Methods:L-WRN cells were cultured routinely and the conditioned medium with different concentrations of fetal bovine serum(FBS)was collected. Six to eight weeks old C57BL/ 6 mice were sacrificed and 15 cm small intestine from the terminal ileum was removed and cut longitudinally. Crypts were digested with EDTA and then collected and embedded in Matrigel? Matrix;after polymerization of Matrigel? Matrix,L-WRN conditioned medium at different concentration gradient was added. The budding ratio and length of buds were measured dynamically under microscope. The enteroids were re-embedded for subculture when certain length of buds was reached. Results:Compared with L-WRN conditioned medium containing 20% FBS,the conditioned medium containing 10% FBS was more favorable for enteroids culture in vitro. When conditioned medium accounted for 10% ,15% ,20% ,25% or 30% of the mixed medium,they all promoted the growth of enteroids and the 15% one seemed to yield better result. Conclusions:An enteroids culture technique was successfully established for the first time in China. When the L-WRN conditioned medium containing 10% FBS accounts for 15% of the mixed medium,it might promote budding better than the others.