SUMMARY Inflammatory cytokines have been proposed to regulate epithelial homeostasis during intestinal inflammation. We report here that interferon-γ (IFN-γ) regulates the crucial homeostatic functions of cell proliferation and apoptosis through serine-threonine protein kinase AKT-β-catenin and Wingless-Int (Wnt)-β-catenin signaling pathways. Short-term exposure of intestinal epithelial cells to IFN-γ resulted in activation of β-catenin through AKT, followed by induction of the secreted Wnt inhibitor Dkk1. Consequently, we observed an increase in Dkk1-mediated apoptosis upon extended IFN-γ treatment, and reduced proliferation through depletion of the Wnt co-receptor LRP6. These effects were enhanced by tumor necrosis factor-α (TNF)-α, suggesting synergism between the two cytokines. Consistent with these results, colitis in vivo was associated with decreased β-catenin-T-cell factor (TCF) signaling, loss of plasma membrane-associated LRP6, and reduced epithelial cell proliferation. Proliferation was partially restored in IFN-γ - deficient mice. Thus, we propose that IFN-γ regulates intestinal epithelial homeostasis by sequential regulation of converging β-catenin signaling pathways.

Desmosomal cadherins mediate cell–cell adhesion in epithelial tissues and have been known to be altered in cancer. We have previously shown that one of the two intestinal epithelial desmosomal cadherins, desmocollin-2 (Dsc2) loss promotes colonic epithelial carcinoma cell proliferation and tumor formation. In this study we show that loss of the other intestinal desmosomal cadherin, desmoglein-2 (Dsg2) that pairs with Dsc2, results in decreased epithelial cell proliferation and suppressed xenograft tumor growth in mice. Dsg2-deficient cells demonstrated a compensatory increase in Dsc2 expression, and small interfering RNA-mediated loss of Dsc2 restored proliferation in Dsg2-deficient cells. Dsg2 downregulation inhibited epidermal growth factor receptor (EGFR) signaling and cell proliferation through altered phosphorylation of EGFR and downstream extracellular signal-regulated kinase activation in parallel with inhibited EGFR receptor internalization. Additionally, we demonstrated a central role of Dsc2 in controlling EGFR signaling and cell proliferation in intestinal epithelial cells. Consistent with these findings, analyses of human colon cancers demonstrated increased Dsg2 protein expression. Taken together, these data demonstrate that partner desmosomal cadherins Dsg2 and Dsc2 play opposing roles in controlling colonic carcinoma cell proliferation through differential effects on EGFR signaling.

This study provides evidence that decreased expression of the desmosomal cadherin desmocollin-2 enhances intestinal epithelial cell proliferation and promotes tumor formation via an Akt/β-catenin pathway.

Summary The migration of polymorphonuclear leukocytes (PMN) across the intestinal epithelium is a histopathological hallmark of many mucosal inflammatory diseases including inflammatory bowel disease (IBD). The terminal transmigration step is the detachment of PMN from the apical surface of the epithelium and their subsequent release into the intestinal lumen. The current study sought to identify epithelial proteins involved in the regulation of PMN migration across intestinal epithelium at the stage where PMN reach the apical epithelial surface. A panel of antibodies reactive with IFNγ-stimulated T84 intestinal epithelial cells was generated. Screening efforts identified one mAb, GM35, which prevented PMN detachment from the apical epithelial surface. Microsequencing studies identified the GM35 antigen as human CD44. Transfection studies confirmed this result by demonstrating the loss of the functional activity of the GM35 mAb following attenuation of epithelial CD44 protein expression. Immunoblotting and immunofluoresence revealed the GM35 antigen to be an apically expressed v6 variant-exon containing form of CD44 (CD44v6). ELISA analysis demonstrated the release of soluble CD44v6 by T84 cells during PMN transepithelial migration (TEM). In addition, the observed release of CD44v6 was blocked by GM35 treatment supporting a connection between CD44v6 release and PMN detachment. Increased expression of CD44v6 and the GM35 antigen was detected in inflamed ulcerative colitis tissue. This study demonstrates for the first time that epithelial expressed CD44v6 plays a role in PMN clearance during inflammatory episodes through regulation of the terminal detachment of PMNs from the apical epithelial surface into the lumen of the intestine.

Expression of the tight junction protein junctional adhesion molecule‐A (JAM‐A) has been linked to proliferation and tumour progression. However, a direct role for JAM‐A in regulating proliferative processes has not been shown. By using complementary in vivo and in vitro approaches, we demonstrate that JAM‐A restricts intestinal epithelial cell (IEC) proliferation in a dimerization‐dependent manner, by inhibiting Akt‐dependent β‐catenin activation. Furthermore, IECs from transgenic JAM‐A−/−/β‐catenin/T‐cell factor reporter mice showed enhanced β‐catenin‐dependent transcription. Finally, inhibition of Akt reversed colonic crypt hyperproliferation in JAM‐A‐deficient mice. These data establish a new link between JAM‐A and IEC homeostasis.