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P029. Inflammasome activation P2X7-dependent in Crohn's disease

A. Zelante1, R. Borgoni2, R. D'Incà3, G.C. Sturniolo3, J.F. Blume1, G. Donvito4, S. Falzoni4, F. Di Virgilio4, 1University Hospital Sant'Anna, Medicine, Ferrara, Italy, 2University of Milano-Bicocca, Department of Statistics, Milan, Italy, 3University of Padua, Department of Surgical, Oncological and Gastroenterological Sciences, Padua, Italy, 4University of Ferrara, Department of Experimental and Diagnostic Medicine, Ferrara, Italy

Background

The Inflammasome represents an intracellular multiprotein complex belonging to the innate immune system that identifies molecular damage. The activation of the Inflammasome triggers the maturation and secretion of cytokines IL-1 beta, IL-18, IL-33 which activate on his part inflammatory processes. The most important Inflammasome is NALP3 (NOD-like family) with his components P2X7, NALP3, the adapter ASC and caspase-1. NALP3 and NOD2 polymorphisms are associated with development of Crohn's disease (CD).

Methods

This study analyzed the expression and function of the NALP3-inflammasome by stimulating in vitro PBMCs of CD patients.

We enrolled 62 CD patients: 36 female (58.1%) and 26 male (41.9%) with a mean age of 53.7 years; the control group included 58 subjects (38 female; 20 male; mean age 46 years). The patients have been analyzed with regard to duration, disease activity, smoking habits, comorbidities, type of therapy, previous surgery, familiarity for IBD and CRP values. We isolated PBMCs to extract RNA for rt-PCR and proteins for Western Blotting.

Results

Our results show a significant difference in the expression of the Inflammasome's component in CD patients (Wilcoxon–Mann–Whitney test and multivariate analysis). As expected, there was a correlation between P2X7 and ASC values (cor 0.537; p-value = 0.00003). CD patients had increased expression of IL-1 beta (p = 0.0012) and activation of the P2X7 receptor (p = 0.00001) suggesting a role for the Inflammasome in CD pathogenesis. The increased expression of ASC (p = 0.0166) and NALP3 (p = 0.0082) confirmed the capability of the Inflammasome to maintain disease activity.

Several features impacted on the Inflammasome's expression: disease's location, comorbidities, therapy, smoking habits and elevated CRP values. Increased expression of IL-1 beta (p = 0.0279) and ASC (p = 0.0386) was especially found in ileal CD. Patients receiving immunotherapy showed a different Inflammasome activation compared to patients receiving with mesalazine but disease activity did not modify NALP proteins' expression. Increased CRP predicted increased P2X7 expression independently from other variables.

Conclusion

Inflammasome changed our concept of innate immunity and disclosed new pathogenetic hypothesis of inflammatory diseases. If our results will be confirmed at the mucosal level, P2X7 may be identified as a potential therapeutic target and CD could be classified as an auto-inflammatory diseases.