Glucagon-Like Peptide-2 Stimulates S-Phase Entry of Intestinal Lgr5+ Stem Cells

BACKGROUND & AIMS: Leucine-rich repeat-containing G-protein–coupled receptor-5 (Lgr5)+/olfactomedin-4 (Olfm4)+ intestinal stem cells (ISCs) in the crypt base are crucial for homeostatic maintenance of the epithelium. The gut hormone, glucagon-like peptide-2(1–33) (GLP-2), stimulates intestinal proliferation and growth; however, the actions of GLP-2 on the Lgr5+ ISCs remain unclear. The aim of this study was to determine whether and how GLP-2 regulates Lgr5+ ISC cell-cycle dynamics and numbers. METHODS: Lgr5-Enhanced green-fluorescent protein - internal ribosome entry site – Cre recombinase – estrogen receptor T2 (eGFP-IRES-creERT2) mice were acutely administered human Glycine(2) (Gly2)-GLP-2, or the GLP-2–receptor antagonist, GLP-2(3–33). Intestinal epithelial insulin-like growth factor-1–receptor knockout and control mice were treated chronically with human Gly2 (hGly2)–GLP-2. Cell-cycle parameters were determined by 5-Ethynyl-2'-deoxyuridine (EdU), bromodeoxyuridine, antibody #Ki67, and phospho-histone 3 labeling and cell-cycle gene expression. RESULTS: Acute hGly2–GLP-2 treatment increased the proportion of eGFP+EdU+/OLFM4+EdU+ cells by 11% to 22% (P < .05), without affecting other cell-cycle markers. hGly2–GLP-2 treatment also increased the ratio of eGFP+ cells in early to late S-phase by 97% (P < .001), and increased the proportion of eGFP+ cells entering S-phase by 218% (P < .001). hGly2–GLP-2 treatment induced jejunal expression of genes involved in cell-cycle regulation (P < .05), and increased expression of Mcm3 in the Lgr5-expressing cells by 122% (P < .05). Conversely, GLP-2(3–33) reduced the proportion of eGFP+EdU+ cells by 27% (P < .05), as well as the expression of jejunal cell-cycle genes (P < .05). Finally, chronic hGly2–GLP-2 treatment increased the number of OLFM4+ cells/crypt (P < .05), in an intestinal epithelial insulin-like growth factor-1–receptor–dependent manner. CONCLUSIONS: These findings expand the actions of GLP-2 to encompass acute stimulation of Lgr5+ ISC S-phase entry through the GLP-2R, and chronic induction of Lgr5+ ISC expansion through downstream intestinal insulin-like growth factor-1 signaling.