Abstract
Mucosal biopsies from inflamed colon of inflammatory bowel disease patients exhibit elevated epithelial apoptosis compared with those from healthy individuals, disrupting mucosal homeostasis and perpetuating disease. Therapies that decrease intestinal epithelial apoptosis may, therefore, ameliorate inflammatory bowel disease, but treatments that specifically target apoptotic pathways are lacking. Proteinase-activated receptor-2 (PAR2), a G protein-coupled receptor activated by trypsin-like serine proteinases, is expressed on intestinal epithelial cells and stimulates mitogenic pathways upon activation. We sought to determine whether PAR2 activation and signaling could rescue colonic epithelial (HT-29) cells from apoptosis induced by proapoptotic cytokines that are increased during inflammatory bowel disease. The PAR2 agonists 2-furoyl-LIGRLO (2f-LI), SLIGKV and trypsin all significantly reduced cleavage of caspase-3, -8, and -9, poly(ADP-ribose) polymerase, and the externalization of phosphatidylserine after treatment of cells with IFN-γ and TNF-α. Knockdown of PAR2 with siRNA eliminated the anti-apoptotic effect of 2f-LI and increased the sensitivity of HT-29 cells to cytokine-induced apoptosis. Concurrent inhibition of both MEK1/2 and PI3K was necessary to inhibit PAR2-induced survival. 2f-LI was found to increase phosphorylation and inactivation of pro-apoptotic BAD at Ser(112) and Ser(136) by MEK1/2 and PI3K-dependent signaling, respectively. PAR2 activation also increased the expression of anti-apoptotic MCL-1. Simultaneous knockdown of both BAD and MCL-1 had minimal effects on PAR2-induced survival, whereas single knockdown had no effect. We conclude that PAR2 activation reduces cytokine-induced epithelial apoptosis via concurrent stimulation of MEK1/2 and PI3K but little involvement of MCL-1 and BAD. Our findings represent a novel mechanism whereby serine proteinases facilitate epithelial cell survival and may be important in the context of colonic healing.
Highlights
Inflammatory bowel disease management lacks therapies that heal the epithelial barrier
We sought to determine whether Proteinase-activated receptor-2 (PAR2) activation and signaling could rescue colonic epithelial (HT-29) cells from apoptosis induced by proapoptotic cytokines that are increased during inflammatory bowel disease
Using the HT-29 colonic epithelial cell line, we show that activation of PAR2 decreases TNF-␣- and IFN-␥-induced apoptosis in a MEK1/2- and PI3K-dependent manner
Summary
Inflammatory bowel disease management lacks therapies that heal the epithelial barrier. We sought to determine whether PAR2 activation and signaling could rescue colonic epithelial (HT-29) cells from apoptosis induced by proapoptotic cytokines that are increased during inflammatory bowel disease. We sought to determine whether PAR2 signaling could decrease cytokine-stimulated apoptosis of colonic epithelial cells, based on the contention that even modest changes in the proliferation-apoptosis balance could promote the reestablishment of epithelial homeostasis to drive the resolution of inflammation in IBD. Using the HT-29 colonic epithelial cell line, we show that activation of PAR2 decreases TNF-␣- and IFN-␥-induced apoptosis in a MEK1/2- and PI3K-dependent manner. Phosphorylation and inactivation of the proapoptotic protein, Bcl2-associated death promoter (BAD), as well as up-regulation of anti-apoptotic myeloid cell leukemia 1 (MCL-1), are not fully responsible for the prosurvival effects of PAR2 activation on intestinal epithelial cells
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Free) 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
