Mo1805 Ablation of DCLK1 in Intestinal Epithelium Exacerbates Colonic Epithelial Barrier Damage in Response to DSS Treatment

Abstract

Background: During normal homeostasis the intestinal epithelium maintains a dynamic architecture with rapid renewal every 3-5 days. Cytotoxic and genotoxic insults cause excessive epithelial damage and crypt injury. Intestinal stem cells (ISCs) are responsible for homeostasis and crypt regeneration. It is believed that the quiescent ISCs may represent a population that can be called upon to rescue the crypt following severe injury. Doublecortinlike kinase 1 (Dclk1) is a putative quiescent ISC and tuft cell marker. It is predominantly expressed in relatively quiescent cells near the traditional +4 location within the intestinal crypt. Dclk1+ cells sorted from murine small intestine form spheroids in suspension culture, and when transplanted into nude mice develop into nodules demonstrating early intestinal epithelial secretory lineage commitment. These data suggest that Dclk1 is a putative ISC marker. The role of Dclk1 in epithelium regeneration in response to injury is not clear. In this study, we investigate the role of Dclk1 in colonic epithelial restitution following injury. Aim: To determine whether Dclk1 regulates colonic epithelial barrier function following DSS-induced colitis. Methods: Age and gender matched control (Dclk1flox/flox) and Dclk1 KO (Villin-Cre;Dclk1flox/flox) mice were fed 3% Dextran Sulfate Sodium (DSS) in drinking water for 7 days, followed by water for 2 days. Body weight and occult blood were monitored daily. To assess epithelial barrier permeability, both Dclk1 KO and control mice were gavaged with FITC-dextran. Blood was collected 4 hours later and the serum concentration of FITC-dextran was measured. Corresponding colon tissues were subjected to histological examination, gene expression, and protein expression analysis. Results: Dclk1 KO mice lost significantly more weight than control mice following DSS treatment. The serum concentration of FITC-dextran in Dclk1 KO mice was approximately 2 fold higher than in control mice. The gene expression levels of ISC markers, Dclk1, Lgr5, Bmi1, mTert, and Msi-1, were significantly downregulated in the colon of Dclk1 KO mice compared to control mice following DSS treatment. Protein levels of Ecadherin, Claudin-1, and Claudin-7 were similar in untreated control and Dclk1 KO mice, but decreased dramatically in Dclk1 KO mice following DSS treatment. H&E staining of colon from DSS-treated mice indicated increased inflammation in Dclk1 KO mice relative to controls. Conclusions: In summary, these results show that loss of Dclk1 in colonic epithelial cells results in exacerbated damage to barrier function following DSS treatment. These data also demonstrate a role for Dclk1 in the regulation of other stem cell markers following DSS-induced injury. Taken together, this study suggests that Dclk1 plays an important role in colonic epithelial restitution in response to severe injury.