HOME
Return to Table of Contents
IDENTIFICATION AND FUNCTIONAL ROLE OF A PROSTAGLANDIN TRANSPORTER IN PGE2 INDUCED BARRIER ALTERATIONS
M Lejeune, P Leung, K Chadee
Gastrointestinal Research Group, University of Calgary, Alberta
Prostaglandin E2 (PGE2) is an important pro-inflammatory lipid mediator that is produced in excess in inflammatory bowel diseases (IBD). Indeed, high output PGE2 is associated with various barrier defective enteric pathologies. However, the role and the mechanism whereby excessive PGE2 alter colonic epithelial barrier is not clearly known. We have already shown in vitro that PGE2 signals through E-Prostanoid (EP) receptor 4 and dose-dependently decrease colonic Trans Epithelial Resistance (TER) indicating a loss of epithelial barrier function. Herein, we investigated the significance and the role of prostaglandin transporter (PGT) in PGE2 induced colonic barrier disruption. The impetus for this study was prompted by the observation that there was an apparent differential expression of EP4 receptor in the colonic epithelium during IBD. Notably, EP4 receptors were over-expressed throughout the colonic epithelial cells from patients with IBD whereas; in control healthy colons EP4 receptors were localized only to the epithelial apical plasma membranes. We theorize that PGT maybe involved in the initiation of colonic inflammation by actively transporting PGE2 from lamina propria towards the site of apical EP4 receptors. Thus, to determine if PGE2 are actively transported across colonic cells towards the apical receptors we probed for the presence of a PGT expression in T84 cells by Real Time PCR and Western blot. Surprisingly, PGT transcript and protein were constitutively expressed in T84 cells. More importantly, the expression of PGT in confluent T84 colonic epithelial monolayer was increased 2-fold following stimulation with TNF-alpha. This indicates the potential of pro-inflammatory cytokines in regulating PGT expression. Moreover, to establish that PGE2 is actively transported across T84 epithelial monolayer we traced for the basolateral to apical translocation of 3H-labeled PGE2 as well as an inert 14C-labeled Ethanolamine. A gradual increase in the apical translocation was observed only with 3H-PGE2 with a peak occurring at 24hr. Remarkably, TNF-alpha pretreatment of T84 monolayer significantly accelerated the translocation of 3H-PGE2 as early as 12hr. Thus, it is clear that PGT can play a major role in initiating colonic inflammation by transporting PGE2 towards their appropriate site of action. Taken together, these novel findings unravel a complex cellular mechanism whereby PGE2 plays a critical role in altering the colonic epithelial barrier.
Funded by CCFC