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P081. ER stress leads to epithelial to mesenchymal transition (EMT) in intestinal epithelial cells in a JNK- and Src-dependent manner

C. Stanzel, A. Terhalle, M. Scharl, G. Rogler, University Hospital Zurich, Gastroenterology & Hepatology, Zurich, Switzerland

Background

Fistulae are a frequent complication in Crohn's disease patients. Epithelial to mesenchymal transition (EMT) has been identified as an important driving force of their formation. On the other hand, the importance of ER stress and related pathways in intestinal inflammation has been demonstrated by recent evidence from animal models, the phenotype of human IBD, as well as genetic data. We showed previously, that ER stress leads to the onset of EMT in intestinal epithelial cells. Here, we investigated candidate signaling pathways and therapeutic substances.

Methods

HT-29 cells were stimulated with the ER stress inducer tunicamycin (TM). In order to identify candidate pathways, we used specific inhibitors targeting JNK, Src, Smad or siRNA targeting Ets1 transcription factor prior to stimulation with TM. Polyamines, butyrates and bile acids were tested for their potential therapeutic effect by pre-treating the cells with the substances followed by addition of TM. The extent of ER stress and EMT was determined by measuring typical markers by Western blot and quantitative real time PCR.

Results

Upon treatment with TM induction of the transcription factor Ets1 was found, which is known to be induced upon ER stress and to play a role in induction of EMT. However, knockdown of Ets1 by siRNA did not influence mRNA levels of the typical fibroblast markers vimentin, beta-6-integrin or the ER stress marker gp96. Similar, inhibiting Smad only lead to slightly reduced levels of beta-6-integrin. On the other hand, stimulation with JNK inhibitor II revealed that induction of Ets, beta-6-integrin and gp96 was JNK-dependent. In addition, the Src kinase inhibitor PP2 reduced the expression of Ets1 and gp96, while induction of beta-6-integrin was found to be Src-independent. Analysing potential therapeutic substances, neither spermidine nor sodium phenylbutyrate could counteract the effect of TM. However, pre-incubation of the cells with 10 mM of the bile acid tauroursodeoxycholic acid decreased significantly the levels of vimentin, beta-6-integrin and gp96.

Conclusion

Our data suggest that the JNK and Src pathway are involved in the regulation of ER stress-mediated EMT during fistula formation in Crohn's disease. The bile acid tauroursodeoxycholic acid represents a therapeutic treatment option.