In vitro expansion of single Lgr5^sup +^ liver stem cells induced by Wnt-driven regeneration

• Published in: Nature (London) 2013-02, Vol.494 (7436), p.247
• Main Authors: Huch, Meritxell, Dorrell, Craig, Boj, Sylvia F, van Es, Johan H, Li, Vivian SW, van de Wetering, Marc, Sato, Toshiro, Hamer, Karien, Sasaki, Nobuo, Finegold, Milton J, Haft, Annelise, Vries, Robert G, Grompe, Markus, Clevers, Hans
• Format: Article
• Language: English
• Subjects: Liver; Proteins; Rodents; Stem cells
• ISSN: 0028-0836; 1476-4687

The Wnt target gene Lgr5 (leucine-rich-repeat-containing G-protein-coupled receptor 5) marks actively dividing stem cells in Wnt-driven, self-renewing tissues such as small intestine and colon1, stomach2 and hair follicles3. A three-dimensional culture system allows long-term clonal expansion of single Lgr5^sup +^ stem cells into transplantable organoids (budding cysts) that retain many characteristics of the original epithelial architecture2,4,5. A crucial component of the culture medium is the Wnt agonist RSPO16, the recently discovered ligand of LGR57,8. Here we show that Lgr5-lacZ is not expressed in healthy adult liver, however, small Lgr5-LacZ^sup +^ cells appear near bile ducts upon damage, coinciding with robust activation of Wnt signalling. As shown by mouse lineage tracing using a new Lgr5-IRES-creERT2 knock-in allele, damage-induced Lgr5^sup +^ cells generate hepatocytes and bile ducts in vivo. Single Lgr5^sup +^ cells from damaged mouse liver can be clonally expanded as organoids in Rspo1-based culture medium over several months. Such clonal organoids can be induced to differentiate in vitro and to generate functional hepatocytes upon transplantation into Fah^sup -/-^ mice. These findings indicate that previous observations concerning Lgr5^sup +^ stem cells in actively self-renewing tissues can also be extended to damage-induced stem cells in a tissue with a low rate of spontaneous proliferation. [PUBLICATION ABSTRACT]