THU0229 HSA-MIR-513C-3P OVEREXPRESSION DECREASES XBP-1S CORRELATING WITH INCREASED INFLAMMATION AND AUTOANTIBODIES IN SALIVARY GLANDS FROM SJÖGREN’S SYNDROME PATIENTS
Authors: P. Carvajal, S. Aguilera, B. Heathcote, I. Castro, D. Jara, M. J. Barrera, V. Maracaja, S. González, V. Aliaga, C. Molina, M. J. González
Journal: Annals of the Rheumatic Diseases (2020)

Background: Endoplasmic reticulum (ER) stress and the Unfolded Protein Response (UPR) are linked to inflammation in a variety of human pathologies including autoimmune diseases. Salivary glands (SG) from Sjögren’s syndrome (SS) patients have high levels of IFN-gamma among other cytokines, which trigger or exacerbate protein unfolding or misfolding, inducing ER stress. In this study, we focused on the IRE1α/XBP-1 pathway of the UPR, whose transcription factor is XBP-1s, which induces genes linked to regulation of the secretory pathway. We recently found that SG of SS patients have reduced levels of XBP-1s transcripts associated with an increased DNA methylation of XBP-1 promoter [1]. We postulate that other epigenetic mechanisms, such as miRNAs, could coexist with promoter DNA hyper-methylation to regulate XBP-1s expression. Objectives: Due to hsa-miR-513c-3p overexpression has been reported in SG from SS patients [2] and that XBP-1 is a predicted target of hsa-miR-513c-3p, the aim of this study was to evaluate whether mRNA levels of XBP-1s is modulated by hsa-miR-513c-3p and also if IFN-gamma modify the expression hsa-miR-513c-3p and XBP-1s. Methods: SG biopsies from 16 SS-patients with low and high focus score and 5 controls were analyzed. hsa-miR-513c-3p levels were measured by Taqman miRNA assays, whereas XBP-1s mRNA levels were determined by qRT-PCR. Additionally, in vitro assays using IFN-gamma and chemically synthesized hsa-miR-513c-3p mimics and inhibitors were performed to study its effect on XBP-1s expression. Results: By Taqman assays we validated the overexpression of hsa-miR-513c-3p in SG from 8 SS patients with low (p=0.03) and 8 SS-patients with high (p=0.003) focus score, compared with SG from 5 controls. In the same samples, a decrease of XBP-1s transcript levels was observed in SG from SS-patients with low (p=0.002) and high (p=0.026) focus score. XBP1s transcript levels were negatively correlated with hsa-miR-513c-3p (r=-0.47, p=0.014), Ro (r=-0.73, p=0.0009), ANA (r=-0.7, p=0.0033) and focus score (r=-0.72, p=0.001). Stimulation of 3D-acini with 1 ng/mL IFN-gamma increase the hsa-miR-513c-3p levels (p=0.014) and decrease the XBP-1s transcript levels (p=0.027). A negative correlation was found between hsa-miR-513c-3p and XBP-1s transcript levels in 3D-acini stimulated with IFN-gamma (r=-0.87, p=0.0001). The XBP-1s transcript levels were decreased in HSG cells transfected with hsa-miR-513c-3p mimic and increased in HSG cells transfected with the miRNA inhibitor. Conclusion: IFN-gamma-induced upregulation of hsa-miR-513c-3p is consistent with the presence of STAT1-binding elements in its promoter region. Our findings suggest that the combined action of miRNAs and DNA methylation modulated by IFN-gamma could explain the altered expression of XBP-1s, a key transcription factor involved in cellular proteostasis, affecting secretory function in LSG from SS-patients. Our results confirm previous correlations found between XBP-1s protein levels and clinical parameters of SS-patients, suggesting an association of XBP-1s with inflammation and impaired SG function. References: [1]D. Sepúlveda, et al , Rheumatology (Oxford), 2018;57:1021-32. [2]I. Alevizos, et al ,. Arthritis Rheum, 2011;63:535-44. Acknowledgments : Fondecyt 1160015, Fondecyt Iniciación 11170049, Fondecyt Postdoctorado 3170023, CONICYT fellowship (PC, DJ). Disclosure of Interests: : None declared