Bifidobacterium Longum Subsp. Infantis EVC001 Metabolites Improve Enterocyte Proliferation and Decrease Pathogen-induced Intestinal Permeability in Premature Infants

Background: Interventions such as C-section delivery, antibiotics, and formula feeding can foster early dysbiosis of the preterm infant gut microbiome, significantly altering the concentrations of bacterial metabolites available for cellular function. This can lead to a cascade of adverse effects to which premature infants are particularly vulnerable. Previous research shows a significant increase in overall total organic acids produced in B. infantis EVC001-fed infants, specifically with an increase in acetate and lactate and a decrease in formate and butyrate. This study investigated the effect of B. infantis EVC001 metabolites on intestinal proliferation and integrity in premature intestinal epithelial cell models.

Methods: Pooled fecal water (FW) from infants colonized with B. infantis EVC001 (EVC001) or those who were not (control), as well as major bacterial metabolites (acetate, lactate, butyrate, and formate) at physiological concentrations were added to Premature intestinal epithelial cells (IECs) and human intestinal epithelial stem cells (hIESCs) and analyzed to assess proliferation, membrane integrity, and cytotoxicity

Results: IECs and hIESCs exposed to EVC001 FW showed significantly increased proliferation shown by ATP expression compared to medium alone and control FW (P < 0.0001). Seahorse mitochondrial assays showed a significant decrease in basal respiration in cells treated with control FW as compared to cells treated with EVC001-fed FW or PBS alone (P = 0.0023, P = 0.0045). Conversely, EVC001-fed FW significantly increased ATP-specific respiration compared to control FW or PBS alone (P = 0.018, P = 0.044). In addition, overall mitochondrial maximal respiratory capacity was significantly increased in hIESCs after exposure to EVC001-fed FW compared to cells treated with control FW (P = 0.015). Significantly decreased lactate dehydrogenase (LDH), an indication of decreased membrane integrity, was detected in enterocytes exposed to EVC001 FW compared to controls FW (P < 0.01). Specific bacterial metabolites, lactate and acetate at median physiological concentrations found in EVC001 infant FW significantly increased Caco-2 intestinal integrity measured by transepithelial electrical resistance compared to medium alone (P < 0.01, P < 0.05, respectively), while control levels did not significantly increase proliferation.

Conclusion: Exposure to EVC001 FW significantly increases premature IEC proliferation by increasing available ATP and mitochondrial capacity while lowering LDH production compared to control FW, which may be explained by significantly greater concentrations of acetate and lactate. These data suggest that metabolites produced by B. infantis EVC001 may promote enterocyte growth and improve intestinal integrity in preterm infants.