703 - Adaptive variation of miR-141-3p in breast milk confers a preventive and protective effect on neonatal gastrointestinal health by preemptively activating antioxidant signals.

Publication Number: 703.3911

Xu Han, Wuxi Children's Hospital, Wuxi, Jiangsu, China (People's Republic); Le Zhang, Wuxi Children's Hospital, WUXI, Jiangsu, China (People's Republic); Yun Li, Wuxi Children's Hospital, Wuxi, Jiangsu, China (People's Republic)

Background: Necrotizing enterocolitis (NEC) is an acquired disorder of mucosal damage, especially in premature infants. MicroRNAs (miRNAs) have been found to play an important role in the progression of NEC. Using bioinformatics analysis, we found that the level of miR-141-3p was significantly reduced in intestinal tissues from infants with NEC. However, the mechanism of miR-141-3p in the progression of NEC remains unclear.

Objective: Breast milk extracellular vesicles (EVs) containing microRNAs (miRNAs) have been shown to benefit neonatal intestinal health. However, the specific biological function of miRNAs, their underlying mechanism, and the implications of miRNA content variation for gut homeostasis are not fully understood.

Design/Methods: We examined the expression of miR-141-3p in breast milk EVs during different lactation stages of mothers with premature and full-term birth. We also determined the expression of miR-141-3p in the intestine of mice with necrotizing enterocolitis (NEC) and inflammatory bowel disease (IBD). We assessed the effect of miR-141-3p in macrophages by measuring the expressions of pro-inflammatory cytokines. In vivo, neonatal rats were intraperitoneally injected with miR-141-3p agomir, and then a NEC model was built. The intestinal pathology, inflammation, and barrier permeability were evaluated. Further, RNA-seq was conducted, and differentially expressed genes were analyzed. The luciferase reporter assay was employed to verify the direct targeting of miR-141-3p on Keap1 3’UTR.

Results: We found that miR-141-3p was highly expressed in colostrum EVs, particularly in mothers with premature birth. Additionally, miR-141-3p had significantly lower expression in the colon of IBD and NEC mice. The result of FISH analysis demonstrated that miR-141-3p exhibited predominant expression in macrophages and showed a decreased expression level during colitis. We showed that miR-141-3p inhibited the expression of pro-inflammatory cytokines in macrophage challenged with LPS. In the NEC model, miR-141-3p agomir significantly relieved intestine injury and improved the integrity of the intestinal barrier. We found that miR-141-3p reduced the mRNA level of KEAP1 and enhanced the expression of antioxidant genes. The base mutation of the binding site abolished the protective effect of miR-141-3p on the gastrointestinal tract.

Conclusion(s): Our results suggest that miR-141-3p exhibits adaptive variation in breast milk EVs and contributes to pre-activating the KEAP1/NRF2 pathway, which confers protection for newborns against subsequent gastrointestinal challenges.