Impact of extensively hydrolyzed infant formula on the probiotic and postbiotic properties of <i>Lactobacillus fermentum</i> in an <i>in vitro</i> co-culture model

Early gut homeostasis is a balance between dietary antigen exposure, gut barrier function, microbiome establishment and orchestration of innate and adaptive immune responses. Imbalances during this early time of development can lead to increased susceptibility to immune reactions like allergy. Especially for infants with a predisposition to allergies and who cannot be exclusively breastfed, there are different human milk substitutes, including hydrolyzed infant formula, which are supposed to prevent allergy-associated mechanisms. The physiologic mechanism beyond the destruction of cow's milk allergenic structures in those formulas are currently not fully understood. Therefore, our aim was to elucidate the impact of hydrolyzed infant formula on intestinal homeostasis and presumed mechanisms behind the beneficial effects. In this study, we used a triple co-culture <i>in vitro</i> model of gut inflammation and homeostasis, including enterocyte-, goblet- and macrophage-like cells in a transwell setup, to assess the effect of extensively hydrolyzed (eHF) infant formula compared to standard cow's milk-based infant formula with intact protein (iPF). These formulas were combined or not with heat-inactivated <i>Limosilactobacillus fermentum</i> CECT 5716 (<i>Lf</i>) to test the effect of probiotic compounds in combination with different types of infant formula (<i>i.e.</i> eHF and iPF) on the intestinal barrier and cytokine production. Under LPS-inflammatory trigger, eHF and eHF + <i>Lf</i> increased mucus production and MUC2 mRNA expression, restored epithelial barrier integrity and increased secretion of regulatory TGFβ, compared to respective controls. These results suggest a beneficial role for eHF, and especially eHF + <i>Lf</i>, in restoring intestinal homeostasis and attenuating pro-inflammatory responses.