AIMS. To evaluate the anti-inflammatory properties of EVs from Parabacteroides goldsteinii (Pg), a beneficial gut bacterium. The lipopolysaccharide (LPS) of Pg has been associated with anti-inflammatory activity. As EVs carry LPS, we investigated the effects of Pg-EVs in intestinal and immune cells via in vitro models.
METHODS. Pg-EVs were isolated from culture supernatants by ultrafiltration and Size Exclusion Chromatography. EV preparations were characterised by nanoparticle tracking (ZetaView), protein content and TEM. A Caco2 cell model epithelium was grown in transwell inserts for 21 days and the effect of Pg-EVs on epithelium integrity was assessed via Transepithelial Electrical Resistance. The pro/anti-inflammatory nature of Pg-EVs and kit-purified LPS (iNtRON Biotechnology) from Pg-cells and Pg-EVs was analysed in THP-1 Dual cells. Comparisons were made to controls of Escherichia coli O111:B4 LPS and matched EV preparations from uninoculated Pg growth medium and uropathogenic E. coli (UPEC) cultures.
RESULTS. No effect of Pg-EVs on Caco2 epithelium integrity was observed. A range of concentrations of purified Pg-cell-LPS and Pg-EV-LPS did not induce inflammatory Interferon Regulatory Factor (IRF) pathway in THP1-Dual macrophages. Pg-EVs did not induce IRF pathway, while matched UPEC-EVs did. Combination of Pg-cell-LPS and E. coli O111:B4 LPS, but not Pg-EV-LPS nor Pg-EVs, dampened IRF induction compared to E. coli O111:B4 LPS treatment alone. Pg-LPS did not induce the NF-KB pathway, but Pg-EVs were pro-inflammatory and combinations with E. coli LPS were not anti-inflammatory.
CONCLUSIONS. Pg-EVs contain non-inflammatory LPS but EVs have weak proinflammatory effect. Pg-LPS, but not Pg-EVs, show an anti-inflammatory potential.