The small intestinal epithelium self-renews every four or five days. Intestinal stem cells (Lgr5 crypt base columnar cells (CBCs)) sustain this renewal and reside between terminally differentiated Paneth cells at the bottom of the intestinal crypt. Whereas the signalling requirements for maintaining stem cell function and crypt homeostasis have been well studied, little is known about how metabolism contributes to epithelial homeostasis. Here we show that freshly isolated Lgr5 CBCs and Paneth cells from the mouse small intestine display different metabolic programs. Compared to Paneth cells, Lgr5 CBCs display high mitochondrial activity. Inhibition of mitochondrial activity in Lgr5 CBCs or inhibition of glycolysis in Paneth cells strongly affects stem cell function, as indicated by impaired organoid formation. In addition, Paneth cells support stem cell function by providing lactate to sustain the enhanced mitochondrial oxidative phosphorylation in the Lgr5 CBCs. Mechanistically, we show that oxidative phosphorylation stimulates p38 MAPK activation by mitochondrial reactive oxygen species signalling, thereby establishing the mature crypt phenotype. Together, our results reveal a critical role for the metabolic identity of Lgr5 CBCs and Paneth cells in supporting optimal stem cell function, and we identify mitochondria and reactive oxygen species signalling as a driving force of cellular differentiation.

Trop‐2/EGP‐1/GA733‐1 is a recently identified cell surface glycoprotein highly expressed by human carcinomas. The cytoplasmic tail of Trop‐2 possesses potential serine and tyrosine phosphorylation sites and a phosphatidyl‐inositol binding consensus sequence. Thus, we investigated whether Trop‐2 might be a functional signaling molecule. Using the fluorescent probe Fura‐2, we assayed the cytoplasmic calcium levels in human cancer cells stimulated with anti‐Trop‐2 or control antibodies. Three anti‐Trop‐2 MAbs, Rs7‐7G11, MOv16 and 162‐46.2 specifically induced a transient intracellular calcium level increment in up to 40% of the experiments performed. Polyclonal antisera recognizing recombinant Trop‐2 molecules possessed a much lower stimulation efficiency. The average latency of antibody‐induced Ca2+ rise for OvCa‐432 cells was 64 ± 26 sec. Internal Ca2+ concentrations reached peaks of 190 ± 24 nM vs<0R>. basal levels of 61 ± 4 nM and returned to baseline within 193 ± 37 sec. Similar values were obtained in MCF‐7 cells. For comparison, stimulation of P2‐purinergic receptors on MCF‐7 and OvCa‐432 cells induced a Ca2+ rise in most cases, leading to average internal Ca2+ concentrations of 297 ± 41 and 391 ± 39 nM, respectively. Our findings show that Trop‐2 transduces an intracellular calcium signal, are consistent with the hypothesis that it acts as a cell surface receptor and support a search for a physiological ligand. Int. J. Cancer 76:671–676, 1998.© 1998 Wiley‐Liss, Inc.

IBD syndromes such as Crohn's disease and ulcerative colitis result from the inflammation of specific intestinal segments. Although many studies have reported on the regenerative response of intestinal progenitor and stem cells to tissue injury, very little is known about the response of differentiated lineages to inflammatory cues. Here, we show that acute inflammation of the mouse small intestine is followed by a dramatic loss of Lgr5 stem cells. Instead, Paneth cells re-enter the cell cycle, lose their secretory expression signature, and acquire stem-like properties, thus contributing to the tissue regenerative response to inflammation. Stem cell factor secretion upon inflammation triggers signaling through the c-Kit receptor and a cascade of downstream events culminating in GSK3β inhibition and Wnt activation in Paneth cells. Hence, the plasticity of the intestinal epithelium in response to inflammation goes well beyond stem and progenitor cells and extends to the fully differentiated and post-mitotic Paneth cells.

Adult stem cell niches are often co-inhabited by cycling and quiescent stem cells. In the intestine, lineage tracing has identified Lgr5 + cells as frequently cycling stem cells, whereas Bmi1 +, mTert +, Hopx + and Lrig1 + cells appear to be more quiescent. Here, we have applied a non-mutagenic and cell cycle independent approach to isolate and characterize small intestinal label-retaining cells (LRCs) persisting in the lower third of the crypt of Lieberkühn for up to 100 days. LRCs do not express markers of proliferation and of enterocyte, goblet or enteroendocrine differentiation, but are positive for Paneth cell markers. While during homeostasis, LR/Paneth cells appear to play a supportive role for Lgr5 + stem cells as previously shown, upon tissue injury they switch to a proliferating state and in the process activate Bmi1 expression while silencing Paneth-specific genes. Hence, they are likely to contribute to the regenerative process following tissue insults such as chronic inflammation.