P1339 Extracellular Vesicle-derived msRNA4486 from Faecalibacterium Prausnitzii Attenuates Colitis by Inhibiting MAPK/NF-κB–NLRP3-mediated Intestinal Inflammatory Signaling

Abstract Background The aim of this study was to evaluate the beneficial effect of EVs derived from F. prausnitzii (Fp-EVs) on experimental colitis mice, and further explored the inhibitory effect of Fp-EVs-msRNA on colitis mice and molecular mechanism. Methods The Fp-EVs were separated from the supernatant of Fp culture by ultracentrifugation. The expression profile of small RNA and proteome information of Fp-EVs were obtained by high-throughput sequencing and proteome mass spectrometry. Fp-EVs and Fp-EVs-msRNA mimics were treated with LPS-treated intestinal epithelial cells and DSS/ TNBS-induced colitis mice respectively. RT-PCR, ELISA, WB, FACS, HE, IHC, IF, TUNEL, FITC were used to analyze the molecular expression of inflammation signaling pathway, Treg cell/Th17 cell differentiation, and the degree of colon inflammation and permeability of mice. Results Fp-EVs treatment reduced DSS / TNBS-induced weight loss, hematochezia, disease activity index (DAI) score, and ameliorated colonic epithelial injury and inflammatory cell infiltration in colitis mice. Proteomic analysis showed that a total of 938 proteins were identified in Fp -EVs, including 684 cytoplasmic proteins, 199 extracellular proteins and 175 membrane proteins. According to the msRNA expression profile analysis of Fp -EVs, we screened and verified msRNA1469, msRNA314, msRNA6336, msRNA4486 and msRNA2230 that were highly expressed in Fp and Fp -EVs. We found that the Fp -EVs-msRNA4486-mimic could reduce the activation of MAPKs/NF-κB inflammatory signaling pathway in LPS-stimulated Caco2 cells. Moreover, we also found that Fp -EVs-msRNA4486-agomiR could alleviate DSS-induced colitis and inhibit the activation of the NLRP3- MAPKs/NF-κB inflammatory signaling pathway. Moreover, Fp-EVs-msRNA4486 could suppress the differentiation of Th17 cells. Conclusion Fp-EVs alleviated colon injury and improved intestinal mucosal barrier by regulating intestinal immunity, and inhibiting MAPKs/NF-κB inflammatory signaling pathway. Fp-EVs could carry abundant bacterial components, including msRNAs and protein molecules, among which Fp-EVs-msRNA-4486 could relieve experimental colitis mice by inhibiting the activation of intestinal inflammatory signaling pathway. Conflict of interest: Ye, Lin: No conflict of interest Wang, Xufei: No conflict of interest Xiao, Fangfei: No conflict of interest Li, Xiaolu: No conflict of interest Liu, Gaojie: No conflict of interest Tu, Lewen: No conflict of interest Wang, Yizhong: No conflict of interest Prof. Dr. Zhang, Ting: No conflict of interest