Regulation of hepcidin/iron-signalling pathway interactions by commensal bifidobateria plays an important role for the inhibition of metaflammationrelated biomarkers

Increased concentration of ferrous iron in the gastrointestinal tract increases the number of various pathogens
and induces inflammation. LPS and/or high-fat diet-associated metaflammation is mediated through a quaternary receptor signaling complex containing iron-regulated pathway, IL-6/STAT inflammatory signaling
pathway, hepcidin regulatory pathway, and common TLR4/NF-κB signaling pathway. We, therefore, investigated whether bifidobacteria directly or indirectly ameliorate LPS- and/or high-fat diet-associated metaflammation by reduction of intestinal iron concentration and/or the above-mentioned pathways.
Material & methods: We used a triple co-culture model of HT-29/B6, HMDM and HepG2 cells with apically added
Bifidobacterium pseudolongum (DSMZ 20099), in the absence or presence of iron, LPS or oleate. Expressions of the
biomarkers of interest were determined after 24 h incubation by TaqMan qRT-PCR, cell-based ELISA or Western
blot.
Results: Bifidobacteria inhibited LPS- and oleate-induced protein expression of inflammatory cytokines (IL-6,
TNF-α) concomitantly with decreases in cellular TG and iron concentration. Exposure of co-cultured cells to
bifidobacteria blocked NF-kB activity through inhibition of IκBα, p38 MAPK, and phosphorylation of NF-kB 65
subunit. TaqMan qRT-PCR and Western blot analysis revealed that bifidobacteria downregulated mRNA and
protein expression of BMP6, DMT1, hepcidin, L-ferritin, ferroportin, IL-6, TfR1, Stat3, and TLR4 following exposure to excessive extracellular LPS, oleate and iron. However, the patterns of TLR2 mRNA and protein expression were quite the opposite of those of TLR4.
Conclusion: Commensal bifidobacteria ameliorate metaflammation/inflammatory responses to excessive extracellular LPS, oleate and iron through at least two molecular/signaling mechanisms: i. modulation of interactions
of the hepcidin- and iron-signaling pathways via reduction of excess iron; ii. reduction of pro-inflammatory
cytokines and hepcidin production through inhibition of the TLR4/NF-kB pathway. This may be a molecular