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Lgr5 +  intestinal stem cells (ISCs) drive the fast renewal of intestinal epithelium. Several signaling pathways have been shown to regulate ISC fates. However, it is unclear what are the essential signals to sustain the ISC self-renewal. Here we show that coordination between Wnt and BMP signaling activity is necessary and sufficient to maintain Lgr5 +  ISCs self-renewal. The key function of R-spondin1 is to achieve a high activity of Wnt signaling in the organoid culture. Using the GSK3 inhibitor CHIR-99021 and the BMP type I receptor inhibitor LDN-193189, we can maintain Lgr5 +  ISCs without growth factors in vitro. Our results define the basic signaling pathways sustaining Lgr5 +  ISCs and set up a convenient and economical culture system for their in vitro expansion. This work also set up an example for growth factor-free culture of other adult stem cells.

A growth factor-free culture system underscores the coordination between Wnt and BMP signaling in Lgr5+ intestinal stem cell maintenance