Cftr Modulates Wnt/β-Catenin Signaling and Stem Cell Proliferation in Murine Intestine

Abstract

Background & AimsCystic fibrosis (CF) patients and CF mouse models have increased risk for gastrointestinal tumors. CF mice exhibit augmented intestinal proliferation of unknown etiology and an altered intestinal environment. We examined the role of Cftr in Wnt/{beta}-catenin signaling, stem cell proliferation and its functional expression in the active intestinal stem cell (ISC) population. Dysregulation of intracellular pH (pHi) in CF ISCs was investigated for facilitation of Wnt/{beta}-catenin signaling.\n\nMethodsCrypt epithelia from wild-type (WT) and CF mice were compared ex vivo and in intestinal organoids (enteroids) for proliferation and Wnt/{beta}-catenin signaling by standard assays. Cftr in ISCs was assessed by immunoblot of sorted Sox9EGFP intestinal epithelia and pHi regulation by confocal microfluorimetry of Lgr5+-EGFP ISCs. Plasma membrane association of the Wnt transducer Disheveled 2 (Dvl2) was assessed by fluorescence imaging of live enteroids from WT and CF mice crossed with Dvl2-EGFP/RosamT/mG mice.\n\nResultsRelative to WT, CF intestinal crypts showed a ~30% increase in epithelial and Lgr5+ ISC proliferation and increased Wnt/{beta}-catenin signaling. Cftr was expressed in Sox9EGFPLo ISCs and loss of Cftr induced an alkaline pHi in Lgr5+-EGFP ISCs. CF crypt-base columnar cells (CBCs) demonstrated a generalized increase in plasma membrane Dvl2-EGFP association as compared to WT. Dvl2-EGFP membrane association was charge- and pH-dependent and increased in WT CBCs by Cftr inhibition.\n\nConclusionsCF intestine exhibits increased ISC proliferation and Wnt/{beta}-catenin signaling. Loss of Cftr increases pHi in ISCs which stabilizes the plasma membrane association of the Wnt transducer Dvl, likely facilitating Wnt/{beta}-catenin signaling. Absence of Cftr-dependent suppression of ISC proliferation in the CF intestine may contribute to increased risk for intestinal tumors.\n\n\n\nO_FIG O_LINKSMALLFIG WIDTH=168 HEIGHT=200 SRC=\"FIGDIR/small/156562_ufig1.gif\" ALT=\"Figure 1\">\nView larger version (27K):\norg.highwire.dtl.DTLVardef@cf807forg.highwire.dtl.DTLVardef@fd24baorg.highwire.dtl.DTLVardef@176497dorg.highwire.dtl.DTLVardef@d2b265_HPS_FORMAT_FIGEXP M_FIG O_FLOATNOGraphical AbstractC_FLOATNO C_FIG