Exosomal circRNAs contribute to intestinal development via the VEGF signalling pathway in human term and preterm colostrum

Abstract

Human breast milk (HBM) provides essential nutrients for newborn growth and development, and contains a variety of biologically active ingredients that can affect gastrointestinal tract and immune system development in breastfed infants. HBM also contains mRNAs, microRNAs and lncRNAs, most of which are encapsulated in milk-derived exosomes and exhibit various important infant development related biological functions. While previous studies have shown that exosomal circRNAs are involved in the intestinal epithelial cells’ proliferation and repair. However, the effect of HBM exosomal circRNAs on intestinal development is not clear. In this study, we identified 6756 circRNAs both in preterm colostrum (PC) and term colostrum (TC), of which 66 were upregulated, and 42 were downregulated (|fold change>2|, p < 0.05) in PC. Pathway analysis showed that the VEGF signalling pathway was involved, and network analysis revealed that the differentially expressed circRNAs bound various miRNAs. Further analyses showed that has_circRNA_405708 and has_circRNA_104707 were involved in the VEGF signalling pathway, and that they all bound various mirRNAs. Exosomes found in preterm colostrum (PC) and term colostrum (TC) promoted VEGF protein expression and induced the proliferation and migration of small intestinal epithelial cells (FHCs). Exosomal circRNAs found in human colostrum (HC) binding to related miRNAs may regulate VEGF signalling, and intestinal development.

Non-Coding RNAs in Human Breast Milk: A Systematic Review

“…Our initial search conducted in March 2019 returned 1565 entries, of which 20 were ultimately included in this review (Figure 1). Examination of the reference lists of included articles and an update of the literature search in September 2020 returned a further seven articles (15,36,42,47,49,51,53), and a further three studies were published and integrated into our summary as we were finalising this manuscript (54)(55)(56).…”

“…However, prior to library preparation, Bioanalyzer results indicate significant quantities of RNAs up to 500 nt and possibly 1000 nt (41,43,48). Detailed examination of the origins and function of longer ncRNAs is limited to two articles which have investigated lncRNAs (41) and circRNAs (54). Fifty-five of the 87 developmentally related miRNAs investigated in Karlsson et al (41) were expressed in EVs from at least one of the 30 breast milk samples analyzed.…”

“…Five of these lncRNAs were consistently expressed in 90-100% of the samples, including CRNDE, DANCR, GAS5, SRA1 and ZFAS1 which may be involved in metabolism, adipogenesis and immune cell regulation. More recently, Zhou et al (54) expanded the exploration of human milk ncRNAs and identified 6756 circRNAs associated with milk EVs using a microarray based platform. The overall circRNA profile was not presented in this study, instead they identified differentially expressed miRNAs between term and preterm milk and investigated potential functions of the differentially expressed circRNAs as described below (54).…”

“…In order to further support proposed physiological effects of breast milk miRNAs, experiments to clarify whether milk vesicles are taken up by cells were conducted in four studies, from two research groups. Additionally, the uptake of vesicles and potential function of circRNAs was investigated by Zhou et al (54). Each of these studies investigated if milk vesicles, most commonly referred to as exosomes, were internalized in cell cultures of different kinds.…”

Exosomes: Potential Disease Biomarkers and New Therapeutic Targets

“…Exosomes are also known to be involved in maintaining the health of infants, being their main function to transport proteins and RNA in the mother's milk to the infant's intestinal system, protecting them from acidity and degradation, thus allowing them to remain intact and to be absorbed [85,86]. These molecules significantly promote the viability of intestinal and gingival epithelial cells, increasing their proliferation and stimulating their activity [87]. For example, some exosomal circular RNAs (circRNAs), found in colostrum, play important biological roles through binding to their respective miRNAs, which promote vascular endothelial growth factor expression and induce proliferation and migration of small intestinal epithelial cells.…”