Active Transport of Bile Acids Decreases Mucin 2 in Neonatal Ileum: Implications for Development of Necrotizing Enterocolitis

Abstract

Necrotizing enterocolitis (NEC) is the most common gastrointestinal emergency of premature infants, but its etiology remains unclear. We have previously shown that mucin 2 (Muc2) positive goblet cells are significantly decreased in NEC. We have also shown that ileal bile acids (BAs) are significantly increased during the development of this disease. Because BAs can affect mucins, we hypothesized that elevated ileal BAs contribute to decreased Muc2 in experimental NEC. The role of Muc2 in NEC was evaluated in Winnie +/+ mice, a strain that produces aberrant Muc2. Muc2 and trefoil factor 3 (Tff3) were assessed in neonatal rats subjected to the NEC protocol when bile acids were removed, and in ileal explants from newborn and older rats cultured with and without BAs. Further, the role of active transport of BAs was determined using neonatal rats given the apical sodium dependent bile acid transporter (Asbt) inhibitor SC-435 and in neonatal Asbt knockout mice subjected to the NEC protocol. Mice with aberrant Muc2 had significantly greater incidence and severity of NEC. Using both in vivo and ex vivo techniques, we determined that BAs decrease Muc2 positive cells in neonatal but not older ileum. However, Tff3 positive cells are not decreased by BAs. In addition, active transport of BAs is required for BAs to decrease Muc2 in immature ileum. These data show that functional Muc2 plays a critical role in the prevention of NEC and BAs can potentiate the decreased Muc2 in disease development. Further, BAs have a more profound effect on Muc2 in immature versus older ileum, which may explain at least in part why NEC occurs almost exclusively in premature infants.

Fecal Bile Salts and the Development of Necrotizing Enterocolitis in Preterm Infants

“…In agreement with the postulated role of increased BS uptake in the distal small intestine, ASBT expression (mRNA and immunohistochemistry) was increased in intestinal samples of preterm infants with NEC in comparison to ileal samples taken during surgery on preterm infants with other diseases [6]. Furthermore, BSs have been shown to decrease ileal mucin production more profoundly in immature than in older ileums, and alter the intestinal mucus layer which might compromise the intestinal barrier function [7]. …”

The role of intestinal epithelial barrier function in the development of NEC

“…In both humans 87,88 and rats 76 with NEC, the number of mucin 2 and trefoil factor 3 goblet cells is significantly reduced and mice with genetically aberrant mucin 2 develop more severe disease than those with normal mucin. 89 In addition, ileal BAs, increased in NEC and associated with disease severity, decrease ileal mucin levels in neonatal but not adult ileum, 89 data which indicate a possible mechanism for development of NEC in premature infants. Zhang et.…”

Toll-like receptor regulation of intestinal development and inflammation in the pathogenesis of necrotizing enterocolitis

“…It has been suggested that increased luminal bile acids [165], abnormal bile acid metabolism [147] and dysregulated bile acid transporters [166, 167] contribute to NEC via dampening goblet cell differentiation and decreasing mucin Muc2 expression in premature ileum [148]. We further showed that bile acids activated Notch signaling plays a role in the development of NEC [66].…”

“…In seeking to understand the mechanisms by which TLR4 regulates enterocyte differentiation, TLR4 deletion was found to suppress Notch signaling in the small intestinal epithelium, which resulted in increased goblet cell differentiation and decreased NEC severity [66]. Interestingly, bile acids, which are significantly increased during the development of NEC [147] and are associated with reduced goblet cells and decreased mucin Muc2 expression in premature ileum [148], are significantly reduced in TLR4 ΔIEC mice and TLR4 knockout mice [66]. …”