AB0158 DECREASED AUTOPHAGY IN SALIVARY GLANDS OF PRIMARY SJöGREN'S SYNDROME PATIENTS COULD BE ASSOCIATED WITH AN INCREASED EXPRESSION OF INFLAMMATORY MARKERS. (June 2019) - Barrera, María-José
Abstract : Background: Salivary glands (SG) of primary Sjögren's syndrome (pSS) patients show high levels of IL-6 and endoplasmic reticulum (ER) stress 1 . In response to ER stress salivary epithelial cells trigger the unfolded protein response, seeking alleviate ER stress through several mechanisms, such as autophagy. The autophagy does not only eliminate misfolded protein aggregates, but also decreases the inflammation selectively removing proteins related to TLRs and inflammasomes. Interestingly, the activation of IL-6/JAK/STAT3 signaling pathway inhibits the autophagy through an increase in MCL-1 expression 2 . Given the role of the autophagy as anti-inflammatory mechanism, it is interesting to evaluate if SG from pSS patients show a decrease in autophagy and if this correlates with the increased expression of inflammatory markers. Objectives: To evaluate the expression of autophagy markers in labial SG (LSG) from pSS patients and 3D-acini deficient in autophagy. In addition, the expression of inflammatory marker IL-6, as well as, autophagy inhibitor MCL-1 was determined in 3D-acini control or deficient in autophagy, evaluating the possible participation of IL-6/JAK/STAT3 signaling pathway. Methods: In LSG of 11 anti-Ro/La seropositive pSS patients and 10 control subjects, mRNA levels of ATG5, mTOR and Beclin-1 were measured by qPCR. ATG5, p62 and LC3B protein levels were measured by Western blot in LSG or 3D-acini deficient in autophagy by knocking down ATG5. HSG cells were transduced with lentiviral vectors expressing shRNAs against ATG5 mRNA or a control vector. Later 3D-acini were generated from shATG5 and control cells. Acini were incubated with 10 ng/mL recombinant IL-6 in the presence or absence of 1.5 μM of JAK inhibitor tofacitinib for 24 h. The mRNA levels of IL-6 and MCL-1 were measured by qPCR. Results: A significant decrease in mRNA levels of ATG5, mTOR and Beclin-1 was observed in LSG of SS-patients. In addition, a significant decrease of ATG5 protein levels together with an increase in p62 receptor protein levels was determined in LSG from SS patients. In shATG5 3D-acini an increase in p62 protein levels was observed similar to LSG from SS patients. Also, an increase in mRNA levels of MCL-1 was determined in 3D-acini stimulated with IL-6 and reverted with tofacitinib. Moreover, an increase in mRNA levels of IL-6 was measured in shATG5 3D-acini compared to 3D-acini control. Conclusion: Our results suggest an attenuation of autophagy in salivary epithelial cells from anti-Ro/La seropositive pSS patients, which could be associated with an increased expression of inflammatory markers. We postulate that a plausible via involved in the autophagy attenuation in LSG from pSS patients could be the IL-6/JAK/STAT3 pathway, which induces MCL-1 expression. Further experiments are needed to elucidate this proposal. References: [1] Barrera, MJ., et al. Autoimmun Rev, 2018. 17(8):p.796-808. [2] Qin, B., et al. Sci Rep, 2015. 5: p.15701. Acknowledgement: Fondecyt-Postdoctorado-3170023, Fondecyt-1160015, Fondecyt-Iniciación-11170049. Disclosure of Interests: None declared