Bifidobacterium longum subsp. longum K5 alleviates inflammatory response and prevents intestinal barrier injury induced by LPS in vitro based on comparative genomics

Abstract

• K5 inhibited TNF-α, IL-1β, IL-6, ROS and NO level better than K15 in the LPS-induced RAW 264.7 cell model. • K5 increased TEER value and decreased paracellular permeability in LPS-induced Caco-2 cell monolayers model. • Comparative genomics showed K5 had a specific gene encoding fatty acid synthase associated with the immune system. • Comparative genomics showed K5 and K15 had four genes with only an amino acid mutation. Excessive immune responses and intestinal barrier injury are leading causes of inflammatory bowel diseases (IBD). Previously, we demonstrated that B. longum subsp . longum K5 alleviated DSS-stimulated colitis in mice, while K15 could not. Here, we investigated both strains' underlying mechanisms regarding their anti-inflammatory and intestinal barrier protective capacities. We assessed the potential characters of two strains, anti-inflammation activities in the LPS-induced RAW 264.7 cell model, and intestinal epithelial barrier functions in the LPS-induced Caco-2 cell monolayers model. Results showed that B. longum subsp . longum K5 exhibited high inhibition against the adhesion of E. coli ATCC 25922 to HT-29 and acetic acid production. B. longum subsp. longum K5 (MOI 1:100) significantly decreased the pro-inflammatory cytokines (TNF-α, IL-1β, IL-6) and inflammatory mediator ROS, NO level in LPS (1 μg/mL) -induced RAW 264.7 cells. It also significantly increased TEER value and decreased paracellular permeability of Caco-2 cells stimulated by LPS. B. longum subsp. longum K5 (MOI 1:100) up-regulated ZO-1 , occlaudin and claudin-1 mRNA expression, down-regulated the mRNA expression level of TLR4 and pro-inflammatory cytokines ( TNF-α, IL-1β, IL-6 ) in LPS (100 μg/mL) -stimulated Caco-2 cells. While B. longum subsp. longum K15 did not do as well as B. longum subsp. longum K5 on these assays. Furthermore, the whole-genome sequencing results revealed a gene encoding fatty acid synthase associated with the immune system in K5 specific genes, LacI family related with carbohydrate metabolism, and serine/threonine-protein kinase involved in signal transduction with an amino acid mutation. Our findings provided the theoretical basis for B. longum subsp . longum K5 to be used as a candidate for IBD therapy.