Proceedings of The Physiological Society
Newcastle University (2009) Proc Physiol Soc 16, C17
Epidermal Growth Factor chronically regulates chloride secretion across colonic epithelial cells.
M. S. Mroz1, F. Toumi1, F. O'Mahony1, S. J. Keely1
1. RCSI, Dublin, Ireland.
Cl- secretion, the primary driving force for intestinal fluid secretion, is dysregulated in many intestinal disorders, leading to the clinical manisfestation of diarrhoea. Growth factors, such as epidermal growth factor (EGF), are important regulators of epithelial Cl- secretion and previous work from our laboratory has shown that EGF chronically enhances colonic epithelial secretory function. Aim: To investigate molecular mechanisms underlying EGF potentiation of colonic epithelial secretory capacity. Monolayers of T84 cells, were mounted in Ussing chambers and Cl- secretion measured as changes in short-circuit current (Isc). Transport protein expression was measured by RT-PCR. Results are expressed as mean±sem for a series of n experiments. Statistical analyses were made by paired Student t-test or one way ANOVA amd Newman-Keuls post-test. p values<0.05 were considered to be significant. Results: Acute treatment with EGF (100 ng/ml for 15 min) induced increases in mRNA expression of key components of the Cl- secretory pathway, including CFTR (270±20% of control; n=3; p<0.01), NKCC1 (338±90% of control; n=3; p<0.001), KCNN4 (150±20% of control; n=5; p<0.05) and the Na/K-ATPase β1 subunit (370±100% of control; n=4; p<0.01). EGF also increased mRNA levels of transmembrane protein 16A (TMEM16A) (256±45% of control; n=6; p<0.01) a recently identified protein associated with Ca2+-dependent Cl- channel activity. To determine signaling pathways involved in mediating the effects of EGF, we examined the effects of PKC (GF 109203X), PI3-K (LY294002), ERK (PD98059) and p38 MAPK (SB203580) inhibitors on the ability of the growth factor to enhance Isc responses to either the Ca2+ or cAMP-dependent secretagogues, carbachol(CCh) and forskolin(FSK) (Table 1). EGF chronically increases colonic epithelial secretory function through upregulation of key transport proteins that comprise the Cl- secretory pathway, including TMEM16A, a novel Ca2+-dependent Cl- conductance that has not previously been shown to be expressed in colonic epithelia. Signaling pathways underlying EGF regulation of transport protein expression are complex and involve multiple effector proteins such as MAPKs, PKC, and PI3-kinase. Our data suggest that modulation of EGFR-dependent signaling pathways may prove a useful approach in therapy of intestinal disorders associated with dysregulated epithelial transport.
Where applicable, experiments conform with Society ethical requirements