Examining how host cells affect metabolic behaviors of probiotics is pivotal to better understand the mechanisms underlying the probiotic efficacy in vivo. However, studies to elucidate the interaction between probiotics and host cells, such as intestinal epithelial cells, remain limited. Therefore, in this study, we performed a comprehensive metabolome analysis of a co-culture containing Bifidobacterium breve MCC1274 and induced pluripotent stem cells (iPS)-derived small intestinal-like cells. In the co-culture, we observed a significant increase in several amino acid metabolites, including indole-3-lactic acid (ILA) and phenyllactic acid (PLA). In accordance with the metabolic shift, the expression of genes involved in ILA synthesis, such as transaminase and tryptophan synthesis-related genes, was also elevated in B. breve MCC1274 cells. ILA production was enhanced in the presence of purines, which were possibly produced by intestinal epithelial cells (IECs). These findings suggest a synergistic action of probiotics and IECs, which may represent a molecular basis of host-probiotic interaction in vivo.

Probiotics, particularly those native to the gut microbiota, have a profound influence on the gut environment. In this study, we conducted a randomized placebo-controlled, double-blind, parallel-group comparison trial to investigate the effects of Bifidobacterium longum BB536 (B. longum BB536) on the fecal microbiota and metabolite compositions in healthy individuals. We compared the effects of fermented milk produced solely with Streptococcus thermophiles and Lactobacillus bulgaricus (placebo group) and fermented milk supplemented with B. longum BB536 (BY group). Our findings revealed a significantly greater relative abundance of Faecalibacterium in the BY group than in the placebo group by the 3rd day, a trend that persisted until the end of the trial on the 17th day. Additionally, the BY group presented significantly increased concentrations of tryptophan (Trp), Indole-3-lactic acid, and Indole-3-aldehyde on the 17th day. A significant positive correlation was observed between the relative abundance of Faecalibacterium and the number of viable B. longum BB536 bacteria in the feces. The concentrations of Trp and Indole-3-acetic acid were also significantly correlated with the number of viable B. longum BB536 bacteria in the feces. Our results suggest that B. longum BB536 intake can modulate the gut microbiota and metabolite profiles, which are general indicators for monitoring the gut environment, potentially conferring health benefits to the host.

Bifidobacteria, dominant members of the infant gut microbiota, are essential for immune system development and are commonly added to infant formula as probiotics. However, the preparation of powdered infant formulas involves dissolving the powder in hot water (>70 °C) to inhibit the growth of harmful bacteria, which can also affect probiotic viability. In this study, we assessed the cytokine production capacity of probiotic bifidobacteria exposed to 70 °C for 1 min, replicating the heating conditions for infant formula preparation. We subjected Bifidobacterium longum subsp. longum BB536, B. breve M-16V, and B. longum subsp. infantis M-63 to 70 °C for 1 min. Next, we exposed these strains to dendritic cells in vitro to observe the immunostimulatory activity. We found that heat treatment resulted in reduced cytokine production compared to that of unheated bacterial cells. The reduction in the immunostimulatory effect was less significant in bacterial cells heated at 70 °C for 1 min than in those heated at 90 °C for 15 min-the latter representing the heating condition used to completely kill bacteria during manufacturing. We also discovered that certain heat shock proteins, such as IbpA, which are highly expressed in B. longum BB536, retained their cytokine production capacity, even at 70 °C, but lost this capacity at 90 °C. These findings indicate that incorporating live bifidobacteria into infant formula as immunostimulatory ingredients is beneficial, even after the 70 °C preparation step that renders most cells non-viable.