A comparison of exosomes derived from different periods breast milk on protecting against intestinal organoid injury

Aim of the study Human breast milk reduces the risk and severity of necrotizing enterocolitis (NEC). Exosomes are extracellular vesicles (EVs) found in high concentrations in milk, and they mediate intercellular communication and immune responses. The aim of this study is to compare the protective effects of exosomes that are derived from different time periods of breast milk production against intestinal injury using an ex vivo intestinal organoid model. Methods Colostrum, transitional and mature breast milk samples from healthy lactating mothers were collected. Exosomes were isolated using serial ultracentrifugation and filtration. Exosomes’ presence was confirmed using transmission electron microscopy (TEM) and western blot. To form the intestinal organoids, terminal ileum was harvested from neonatal mice pups at postnatal day 9, crypts were isolated and organoids were cultured in matrigel. Organoids were either cultured with exposure to lipopolysaccharide (LPS), or in treatment groups where both LPS and exosomes were added in the culturing medium. Inflammatory markers and organoids viability were evaluated. Main results Human milk-derived exosomes were successfully isolated and characterized. LPS administration reduced the size of intestinal organoids, induced inflammation through increasing TNFα and TLR4 expression, and stimulated intestinal regeneration. Colostrum, transitional and mature human milk-derived exosome treatment all prevented inflammatory injury, while exosomes derived from colostrum were most effective at reducing inflammatory cytokine. Conclusions Human breast milk-derived exosomes were able to protect intestine organoids against epithelial injury induced by LPS. Colostrum exosomes offer the best protective effect among the breast-milk derived exosomes. Human milk exosomes can be protective against the development of intestinal injury such as that seen in NEC.