Probiotics prevent necrotizing enterocolitis by modulating enterocyte genes that regulate innate immune-mediated inflammation

Abstract

Necrotizing enterocolitis (NEC), an extensive intestinal inflammatory disease of premature infants, is caused, in part, by an excessive inflammatory response to initial bacterial colonization due to the immature expression of innate immune response genes. In a randomized placebo-controlled clinical trial, supplementation of very low birth weight infants with probiotics significantly reduced the incidence of NEC. The primary goal of this study was to determine whether secreted products of these two clinically effective probiotic strains, Bifidobacterium infantis and Lactobacillus acidophilus, prevented NEC by accelerating the maturation of intestinal innate immune response genes and whether both strains are required for this effect. After exposure to probiotic conditioned media (PCM), immature human enterocytes, immature human intestinal xenografts, and primary enterocyte cultures of NEC tissue (NEC-IEC) were assayed for an IL-8 and IL-6 response to inflammatory stimuli. The latter two models were also assayed for innate immune response gene expression. In the immature xenograft, PCM exposure significantly attenuated LPS and IL-1β-induced IL-8 and IL-6 expression, decreased TLR2 mRNA and TLR4 mRNA, and increased mRNA levels of specific negative regulators of inflammation, SIGIRR and Tollip. In NEC-IEC, PCM decreased TLR2-dependent IL-8 and IL-6 induction and increased SIGIRR and Tollip expression. The attenuated inflammatory response with PCM was reversed with Tollip siRNA-mediated knockdown. The anti-inflammatory secreted factor is a 5- to 10-kDa molecule resistant to DNase, RNase, protease, heat stress, and acid exposure. B. infantis-conditioned media showed superior anti-inflammatory properties to that of L. acidophilus in immature human enterocytes, suggesting a strain specificity to this effect. We conclude that PCM promotes maturation of innate immune response gene expression, potentially explaining the protective effects of probiotics in clinical NEC.

Breast milk, microbiota, and intestinal immune homeostasis

“…In studies from this laboratory (73), we have identified a secreted product from the same B. infantis which on partial characterization is likely a glycan of less than 30 kD size. This factor has an anti-inflammatory effect in a fetal small intestinal cell line (H4) and functions by causing a maturation of the innate inflammatory immune response to an inflammatory stimulus with a reduction in the expression of TLR-4, its signaling molecules and its transcription factor NFκB genes and an increase in negative regulators of this response (TOLLIP, SIGIIR, and A-20) as seen in mature enterocytes (73,74). Finally, a recent study of the molecular response of intestinal bacteria to breast vs. infant formula feeding has shown that the products of breastfed intestinal bacteria have a greater effect on protective genes in enterocytes than products of formula-fed infants (75).…”

<i><b>Bifidobacterium</b></i><b> species lower serum glucose, increase expressions of insulin signaling proteins, and improve adipokine profile in diabetic </b><b>mice </b>

“…After all of the mice had been confirmed to be in a diabetic exerts effects on those pathways remains as yet unknown. The mixture of Bifidobacterium infantis and Lactobacillus acidophilus reportedly decreased the interleukin-6 (IL-6) mRNA level in primary intestinal epithelial cells isolated from resected ileum with necrotizing enterocolitis (11). Bifidobacteria longum significantly down-regulated levels of macrophage chemoattractant protein-1 (MCP-1) and IL-6 in porcine intestinal epithelial cells challenged with heatkilled enterotoxigenic Escherichia coli (34).…”

Case reports: necrotizing enterocolitis

“…There have been in vitro and animal studies as well as human clinical trials investigating the benefits of probiotics [22][23][24] ; however larger trials are necessary prior to recommending routine use in preterm infants [12,25,26] . There are recommendations for early initiation of enteral feeds and advancement of enteral feeds in accordance with evidence based feeding protocols [10,12] , promoting adequate intestinal motility and prevention of carbohydrate malabsorption.…”