Bovine milk-derived exosomes attenuate NLRP3 inflammasome and NF-κB signaling in the lung during neonatal necrotizing enterocolitis.

PURPOSE: Necrotizing enterocolitis (NEC), an inflammatory intestinal disease common in premature infants, has been associated with the development of lung damage. Toll-like receptor 4 has been shown to regulate inflammation in the NEC lungs, however, other important inflammatory mechanisms have not been thoroughly investigated. In addition, we reported that milk-derived exosomes were able to attenuate intestinal injury and inflammation in experimental NEC. This study aims to (i) investigate the role of the NLRP3 inflammasome and NF-κB pathway in regulating lung damage during experimental NEC; and (ii) evaluate the therapeutic potential of bovine milk exosomes in reducing lung inflammation and injury during NEC. METHODS: NEC was induced by gavage feeding of hyperosmolar formula, hypoxia, and lipopolysaccharide administration in neonatal mice from postnatal days 5-9. Exosomes were obtained by ultracentrifugation of bovine milk and administered during each formula feed. RESULTS: The lung of NEC pups showed increased inflammation, tissue damage, NLRP3 inflammasome expression, and NF-κB pathway activation, which were attenuated upon exosome administration. CONCLUSION: Our findings suggest that the lung undergoes significant inflammation and injury following experimental NEC which are attenuated by bovine milk-derived exosomes. This emphasizes the therapeutic potential of exosomes not just on the intestine but also on the lung.

Structure-Function Relationships of Human Milk Oligosaccharides on the Intestinal Epithelial Transcriptome in Caco-2 Cells and a Murine Model of Necrotizing Enterocolitis.

SCOPE: Necrotizing enterocolitis (NEC) is a devastating gastrointestinal emergency affecting preterm infants. Breastmilk protects against NEC, partly due to human milk oligosaccharides (HMOs). HMO compositions are highly diverse, and it is unclear if anti-NEC properties are specific to carbohydrate motifs. Here, this study compares intestinal epithelial transcriptomes of five synthetic HMOs (sHMOs) and examines structure-function relationships of HMOs on intestinal signaling. METHODS AND RESULTS: This study interrogates the transcriptome of Caco-2Bbe1 cells in response to five synthetic HMOs (sHMOs) using RNA sequencing: 2'-fucosyllactose (2'-FL), 3-fucosyllactose (3FL), 6'-siallyllactose (6'-SL), lacto-N-tetraose (LNT), lacto-N-neotetraose (LNnT). Protection against intestinal barrier dysfunction and inflammation occurred in an HMO-dependent manner. Each sHMO exerts a unique set of host transcriptome changes and modulated unique signaling pathways. There is clustering between HMOs bearing similar side chains, with little overlap in gene regulation which is shared by all sHMOs. Interestingly, most sHMOs protect pups against NEC, exerting divergent mechanisms on intestinal cell morphology and inflammation. CONCLUSIONS: These results demonstrate that while structurally distinct HMOs impact intestinal physiology, their mechanisms of action differ. This finding establishes the first structure-function relationship of HMOs in the context of intestinal cell signaling responses and offers a functional framework by which to screen and design HMO-like compounds.

Amniotic fluid and breast milk: a rationale for breast milk stem cell therapy in neonatal diseases.

Amniotic fluid and breast milk play important roles in structural development throughout fetal growth and infancy. Given their significance in physical maturation, many studies have investigated the therapeutic and protective roles of amniotic fluid and breast milk in neonatal diseases. Of particular interest to researchers are stem cells found in the two fluids. These stem cells have been investigated due to their ability to self-replicate, differentiate, reduce tissue damage, and their expression of pluripotent markers. While amniotic fluid stem cells have received some attention regarding their ability to treat neonatal diseases, breast milk stem cells have not been investigated to the same extent given the recency of their discovery. The purpose of this review is to compare the functions of amniotic fluid, breast milk, and their stem cells to provide a rationale for the use of breast milk stem cells as a therapy for neonatal diseases. Breast milk stem cells present as an important tool for treating neonatal diseases given their ability to reduce inflammation and tissue damage, as well as their multilineage differentiation potential, easy accessibility, and ability to be used in disease modelling.

Protective effects of vitamin D against injury in intestinal epithelium.

BACKGROUND: Vitamin D deficiency is associated with intestinal barrier dysfunction, which contributes to pathogenesis of acute intestinal injury in children. We aim to investigate the effects of vitamin D on intestinal injury in intestinal epithelial cells and organoids. METHODS: Lipopolysaccharide (LPS) was used to induce injury in intestinal epithelial cells (IEC-18) and organoids, and the effect of vitamin D was assessed. Cell viability was measured and inflammation cytokines TNFα and IL-8 were quantified. FITC-dextran 4 kDa (FD4) permeability was measured using Transwell while tight junction markers were assessed by immunofluorescence staining in IEC-18 and intestinal organoids. Data were compared using one-way ANOVA with Bonferroni post-test. RESULTS: IEC-18 viability was decreased by LPS treatment, but was prevented by vitamin D. The upregulation of inflammation was inhibited by vitamin D, which also decreased epithelium permeability. Vitamin D restored tight junction ZO-1 and claudin 2. In addition, vitamin D decreased TNFα expression and prevented the disruption of ZO-1 in injured organoids. CONCLUSIONS: Vitamin D rescued epithelial barrier function by improving permeability and restoring tight junctions, leading to decrease inflammation. This study confirms the protective effects of vitamin D, which could be used as a treatment strategy for infants at risk of developing intestinal injury.

Necrotizing Enterocolitis: State of the Art in Translating Experimental Research to the Bedside.

Necrotizing enterocolitis (NEC) is a devastating intestinal disease that continues to have high morbidity and mortality among preterm neonates, despite medical advancements in neonatology and neonatal care. To investigate the pathogenesis of the disease and explore novel form of treatment, a variety of experimental models of NEC have been developed and used by various investigators. These experimental models range from in vitro evaluation of intestinal epithelial cells and intestinal organoids to in vivo models of the disease in neonatal mice, rats, and piglets. Most recently, human-derived intestinal organoids have also been developed and investigated. In this review, we will briefly discuss these experimental models and the contributions that they have made to our understanding of NEC. We will also point to the ischemia/reperfusion (I/R) model of intestinal injury which has been used as an indirect model of NEC by some investigators. Advancements in laboratory research into this devastating disease have continued to expand our knowledge on the pathogenesis and prevention of NEC as well as the effectiveness of therapeutic options for management of this severe disease.