B. infantis administration preserves the distribution of occludin and claudin 2 and 4 at tight junctions in a neonatal mouse model of necrotizing enterocolitis

We previously found that, in experimental NEC, claudins 2, 4 and 7 are disrupted at tight junctions (TJ) and intestinal permeability is increased, which is prevented by B. infantis. Whether B. infantis stabilizes claudins at TJ remains unknown. To determine this, claudins 2, 4 and 7 and occludin were examined by immunofluorescence and claudin protein distribution by western blot of subcellular fractions in the intestine of newborn mice submitted to the NEC protocol (stressed) for 24 h or dam-fed controls. Secondly, TJ were examined in stressed pups treated with B. infantis (3x107 CFU) or vehicle only. We found that: 1) in dam-fed pups, claudins 2, 4 and occludin were concentrated at TJ structures of enterocytes, with only minimal cytoplasmic detection, while claudin 7 was detected along cellular membranes. In stressed pups, claudin 2 and 4, and occludin association with TJ was decreased and claudins 4 and 7 were noted throughout the cytoplasm; 2) In dam fed controls, claudins 2, 4 and 7 co-fractionate with caveolin 1 in membrane-rich fractions, while in stressed pups, these predominate in high-density fractions; 3) in B. infantis-treated mice, claudins 2 and 4, and occludin concentration at TJ was preserved, and co-fractionation of claudins 2, 4 and 7 with caveolin 1 in low-density fractions was preserved. In conclusion, B. infantis preserved the distribution of occludin and claudin 2 and 4 at TJ in a neonatal mouse NEC model.