Research Article: Signaling pathways induced by serine proteases to increase intestinal epithelial barrier function

Date Published: July 3, 2017

Publisher: Public Library of Science

Author(s): Kelcie A. Lahey, Natalie J. Ronaghan, Judie Shang, Sébastien P. Dion, Antoine Désilets, Richard Leduc, Wallace K. MacNaughton, Karl X Chai.


Changes in barrier function of the gastrointestinal tract are thought to contribute to the inflammatory bowel diseases Crohn’s disease and ulcerative colitis. Previous work in our lab demonstrated that apical exposure of intestinal epithelial cell lines to serine proteases results in an increase in transepithelial electrical resistance (TER). However, the underlying mechanisms governing this response are unclear. We aimed to determine the requirement for proteolytic activity, epidermal growth factor receptor (EGFR) activation, and downstream intracellular signaling in initiating and maintaining enhanced barrier function following protease treatment using a canine intestinal epithelial cell line (SCBN). We also examined the role of phosphorylation of myosin regulatory light chain on the serine protease-induced increase in TER through. It was found that proteolytic activity of the serine proteases trypsin and matriptase is required to initiate and maintain the protease-mediated increase in TER. We also show that MMP-independent EGFR activation is essential to the sustained phase of the protease response, and that Src kinases may mediate EGFR transactivation. PI3-K and ERK1/2 signaling were important in reaching a maximal increase in TER following protease stimulation; however, their upstream activators are yet to be determined. CK2 inhibition prevented the increase in TER induced by serine proteases. The bradykinin B(2) receptor was not involved in the change in TER in response to serine proteases, and no change in phosphorylation of MLC was observed after trypsin or matriptase treatment. Taken together, our data show a requirement for ongoing proteolytic activity, EGFR transactivation, as well as downstream PI3-K, ERK1/2, and CK2 signaling in protease-mediated barrier enhancement of intestinal epithelial cells. The pathways mediating enhanced barrier function by proteases may be novel therapeutic targets for intestinal disorders characterized by disrupted epithelial barrier function.

Partial Text

The epithelial cells lining the gastrointestinal tract provide a critical barrier to prevent damaging agents from entering the underlying tissue. There are various factors that contribute to barrier function, including Paneth and goblet cell secretions, but it is the tight junction between epithelial cells that provides a final physical barrier against paracellular movement. Decreased barrier function is a characteristic of inflammatory bowel diseases (IBD) such as Crohn’s disease and ulcerative colitis. Over 1 million Americans and 2.5 million Europeans are estimated to have IBD. These diseases incur substantial costs for health care and decrease the quality of life for those affected [1,2]. IBD is a disease of developed nations, and countries that are becoming industrialized show increased incidence of IBD [3].

Recent literature suggests an important role for the endogenous epithelial serine proteases matriptase and prostasin in establishing and maintaining the intestinal epithelial barrier [23,24,39,40]. The apical addition of serine proteases has been shown to significantly increase TER in several colonic epithelial cell lines in an activity-dependent manner [21]. In that study, we showed an important but partial role for PCKζ. Other intracellular mechanisms underlying this response remained unknown. Thus, the main objective of the present study was to determine the downstream signaling events governing protease-mediated barrier enhancement in intestinal epithelial cells.




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