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P030 SUCNR1 mediates inflammasome activation: Relevance in Ulcerative Colitis

Bauset, C.(1);Lis-López, L.(1);Ortiz-Masia, D.(1);Gisbert-Ferrándiz, L.(1);Coll, S.(1);Calatayud, S.(1);Barrachina, M.D.(1);Cosín-Roger, J.(2);

(1)Faculty of Medicine- University of Valencia, Department of Pharmacology, Valencia, Spain;(2)Hospital Dr. Peset, Department of Endocrinology and Metabolism, Valencia, Spain

Background

Ulcerative colitis (UC) is characterized by a diffuse, continuous, and chronic inflammation of mucosa and submucosa layers in the colon1. Inflammasome complex is involved in the intestinal homeostasis regulation, but its role in UC has not been established yet. We have recently reported that SUCNR1 mediates intestinal inflammation and fibrosis2. We aim to analyze the role of SUCNR1 in inflammasome activation and UC.

Methods

Intestinal resections from UC and non-IBD patients were obtained. HT29 cells were treated with succinate 1mM and an inflammasome activator cocktail (TNF-α 25ng/ml, IFN-γ 20 ng/ml and LPS 1µg/ml) for 24 hours and transfected with SUCNR1 siRNA. Chronic DSS-colitis was induced in wild-type (WT) and SUNCR1-/- (KO) mice with 4 cycles of DSS. Gene expression and protein levels of SUCNR1 and inflammasome markers were analyzed by qPCR, WesternBlot and ELISA. Histology of murine tissue was analyzed by Hematoxylin-Eosin staining. Results were expressed as fold induction (mean±SEM, n≥5). Statistical analysis was performed with one-way ANOVA followed by Newman-Keuls test. Correlations were analyzed with the Spearman coefficient.

Results

In UC patients, gene expression of SUCNR1 (4.47±1.70), Nlrp3 (1.97±0.38), Caspase-1 (2.08±0.33) and IL-1β (6.90±2.02) were significantly increased vs non-IBD. SUCNR1 positively correlates with the expression of Caspase-1 (r=0.46) and ASC (r=0.45). Protein levels of SUCNR1 (151.00±8.29), Nlrp3 (215.20±53.35) and Caspase-1 (518.30±231.50) were increased in UC vs non-IBD. HT29 cells treated with succinate and inflammasome cocktail showed an increased gene expression of SUCNR1 (4.45±1.17), Nlrp3 (3.13±0.29), Caspase-1 (70.40±16.14), ASC (2.38±0.37), IL-18 (2.52±0.31) and IL-1β (2.32±0.18). Of interest, siSUCNR1 cells treated with the cocktail and succinate showed a significant reduction in the expression of Nlrp3 (1.00±0.15), IL-1β (0.98±0.21), and ASC (1.19±0.14). In parallel, protein levels of Caspase-1 in siSUCNR1 cells treated with cocktail and succinate were significantly reduced (609.00±116.20) vs non-transfected cells (1095.00±148.30). IL-1β levels were also significantly reduced in siSUCNR1 cells (148.60±16.13) vs non-transfected cells (744.70±94.06). Finally, WT-DSS mice exhibited a worse colon histology and an increased protein expression of Caspase-1 (170.60±32.20), compared with KO-DSS (61.39±4.59).

Conclusion

SUCNR1 is increased and positively correlates with the expression of inflammasome components in UC patients. Moreover, SUCNR1 mediates inflammasome activation and its absence ameliorates chronic DSS-colitis. Hence, SUCNR1 might be a potential pharmacological target for UC treatment.

Ref: 1. Lancet. 2018 Dec 23;390(10114):2769-2778. 2. Mucosal Immunol. 2019 Jan;12(1):178-187

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