ABSTRACT
Many epithelial stem cell populations follow a pattern of stochastic stem cell divisions called 'neutral drift'. It is hypothesised that neutral competition between stem cells protects against the acquisition of deleterious mutations. Here we use a Porcupine inhibitor to reduce Wnt secretion at a dose where intestinal homoeostasis is maintained despite a reduction of Lgr5+ stem cells. Functionally, there is a marked acceleration in monoclonal conversion, so that crypts become rapidly derived from a single stem cell. Stem cells located further from the base are lost and the pool of competing stem cells is reduced. We tested whether this loss of stem cell competition would modify tumorigenesis. Reduction of Wnt ligand secretion accelerates fixation of Apc-deficient cells within the crypt leading to accelerated tumorigenesis. Therefore, ligand-based Wnt signalling influences the number of stem cells, fixation speed of Apc mutations and the speed and likelihood of adenoma formation.
Subject(s)
Carcinogenesis/metabolism , Cell Transformation, Neoplastic/metabolism , Intestinal Mucosa/cytology , Intestinal Mucosa/metabolism , Stem Cells/cytology , Stem Cells/metabolism , Wnt Signaling Pathway , Acyltransferases/antagonists & inhibitors , Adenoma/etiology , Adenoma/metabolism , Adenoma/pathology , Adenomatous Polyposis Coli Protein/deficiency , Adenomatous Polyposis Coli Protein/genetics , Adenomatous Polyposis Coli Protein/metabolism , Animals , Carcinogenesis/drug effects , Cell Transformation, Neoplastic/drug effects , Colorectal Neoplasms/etiology , Colorectal Neoplasms/metabolism , Colorectal Neoplasms/pathology , Enzyme Inhibitors/pharmacology , Intestinal Mucosa/drug effects , Ligands , Membrane Proteins/antagonists & inhibitors , Mice , Mice, Inbred C57BL , Mice, Transgenic , Pyrazines/pharmacology , Pyridines/pharmacology , Stem Cells/drug effects , Wnt Signaling Pathway/drug effectsABSTRACT
Gene expression-based classification systems have identified an aggressive colon cancer subtype with mesenchymal features, possibly reflecting epithelial-to-mesenchymal transition (EMT) of tumor cells. However, stromal fibroblasts contribute extensively to the mesenchymal phenotype of aggressive colon tumors, challenging the notion of tumor EMT. To separately study the neoplastic and stromal compartments of colon tumors, we have generated a stroma gene filter (SGF). Comparative analysis of stromahigh and stromalow tumors shows that the neoplastic cells in stromahigh tumors express specific EMT drivers (ZEB2, TWIST1, TWIST2) and that 98% of differentially expressed genes are strongly correlated with them. Analysis of differential gene expression between mesenchymal and epithelial cancer cell lines revealed that hepatocyte nuclear factor 4α (HNF4α), a transcriptional activator of intestinal (epithelial) differentiation, and its target genes are highly expressed in epithelial cancer cell lines. However, mesenchymal-type cancer cell lines expressed only part of the mesenchymal genes expressed by tumor-derived neoplastic cells, suggesting that external cues were lacking. We found that collagen-I dominates the extracellular matrix in aggressive colon cancer. Mimicking the tumor microenvironment by replacing laminin-rich Matrigel with collagen-I was sufficient to induce tumor-specific mesenchymal gene expression, suppression of HNF4α and its target genes, and collective tumor cell invasion of patient-derived colon tumor organoids. The data connect collagen-rich stroma to mesenchymal gene expression in neoplastic cells and to collective tumor cell invasion. Targeting the tumor-collagen interface may therefore be explored as a novel strategy in the treatment of aggressive colon cancer.
Subject(s)
Colonic Neoplasms/genetics , Epithelial-Mesenchymal Transition/genetics , Hepatocyte Nuclear Factor 4/genetics , Tumor Microenvironment/genetics , Cell Differentiation/genetics , Cell Line, Tumor , Cell Movement/genetics , Cell Proliferation/genetics , Collagen/genetics , Collagen/metabolism , Colonic Neoplasms/pathology , Gene Expression Regulation, Neoplastic , Humans , Stromal Cells/metabolism , Stromal Cells/pathologyABSTRACT
The intestinal epithelium and the hair follicle represent examples of rapidly self-renewing tissue in adult mammals. We have recently identified a novel stem cell gene Lgr5 expressed in multiple adult tissues. At the bottoms of crypts in small intestine and colon as well as in hair follicles, Lgr5 marks cycling cells with stem cell properties (Barker et al. 2007; Jaks et al. 2008). Using an inducible Lgr5-Cre knockin allele in conjunction with the Rosa26-LacZ Cre reporter strain, long-term lineage-tracing experiments were performed in adult mice. The Lgr5(+ve) crypt-based cell generated all epithelial lineages during a 14-month period, implying that it represents the stem cell of the small intestine and colon. Similarly, lineage tracing during a 14-month period revealed that Lgr5(+ve) cells located in the bulge of the hair follicle sustained multiple rounds of hair growth. These observations support the counterintuitive notion that Lgr5(+ve) cells are actively cycling, yet represent long-term stem cells of these adult, self-renewing tissues.
