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OP03. Induction of autophagy upon DC‑T cell contact is mediated by LKB1–AMPK–mTOR signalling

M.E. Wildenberg1, A.C. Vos2, M. Duijvestein2, A. Verhaar3, G. van den Brink4, D.W. Hommes5

1Academic Medical Center, Tytgat Institute for Intestinal and Liver Research, Amsterdam, Netherlands; 2Leiden University Medical Center, Gastroenterology-Hepatalogy, Leiden, Netherlands; 3Leiden University Medical Center, Leiden, Netherlands; 4Academic Medical Center, Gastroenterology & Hepatology, Amsterdam, Netherlands; 5UCLA, Division of Digestive Diseases, Los Angeles, United States

Background: We have previously shown that upon formation of immunogenic dendritic cell (DC)‑T cell contacts, autophagy is induced. This process then functions in a negative feedback loop by destabilization of the interaction. In Crohn's disease patients carrying risk alleles of ATG16L1, this mechanism is impaired, leading to hyperstable interactions and increased immunogenicity. The correction of autophagy induction after DC‑T cell contact in risk allele carriers may form a therapeutic possibilty resulting in normal interaction stability and decreased T cell activation. We therefore sought to determine the signaling events linking DC‑T cell interactions with the induction of autophagy.

Methods: Interactions were induced between human monocyte derived DC and allogeneic T cells. Activation of signalling pathways was determined by Western Blot analysis. Inihibition of signalling mediators was achieved through siRNA and pharmacological inhibitors.

Results: Upon immunogenic DC‑T cell interactions, phosphorylation of AMPK was increased. AMPK is an inhibitor of mTOR signaling, and indeed mTOR pathway activity was markedly reduced, as shown by decreased phosphorylation of mTOR itself as well as its downstream target S6. It has been shown previously that inhibition of mTOR is a strong inducer of autophagy. Conversely, HMGB1 and tissue transglutaminase 2, which are also known to influence autophagic activity, did not appear to be involved in autophagy induction under these circumstances.

Interestingly, AMPK activation was mediated through activation of LKB1, a molecule which has been shown to be involved in mediating the polarity of intestinal epithelial cells. We show that LKB1 is recruited to the site of DC‑T cell contact, and that inhibition of LKB1 signalling results in impaired induction of autophagy after cell-cell contact. Consequently, cell contacts formed by LKB1 deficient DC were hyperstable and resulted in increased immunogenicity, similar to the effects of autophagy deficiency.

Conclusions: This data shows that upon DC‑T cell interaction, autophagy is induced as a regulatory mechanism and this induction is mediated through the LKB1–AMPK–mTOR signaling pathway. In CD patients carrying the ATG16L1 risk allele this regulatory mechanism is disturbed, leading to hyperstable interactions and excessive immunity. Boosting the mTOR pathway may be an interesting therapeutic option in this subset of CD patients.