MicroRNA Was Involved In The Molecular Mechanism Of 2'-fucosyllactose In The Remission Of Cow's Milk Allergy

Cow's milk allergy is one of the most common food allergy,which seriously endangers the health of infants and young children.Currently,most studies believed that the TLR4/NF-?B pathway plays a central role in the pathogenesis of allergies.And microRNA(miRNA),as a newly discovered host gene regulator,has been proved to involved in TLR4-mediated inflammatory signal transduction and production of inflammatory factors.Human milk is the best food source for infants and young children,which can effectively maintain the growth and development of infants and their immune tolerance.An important reason of the immune function of human milk may lie in its high content-human milk oligosaccharides(HMOs),but the specific mechanism of its allergy relief is still unknown.Therefore,this research intends to the most abundant monocase in the HMO,2'-fucosyllactose(2'-FL),as the research object by milk allergy of cell and animal models,application of ELISA and real-time quantitative-PCR and Western blot,cell transfection,were used to analyse effect of 2'-FL on inhibiting TLR4/NF-?B signaling pathway and relieving allergy.This study provides new research ideas and strategies for further revealing the relationship between HMOs and intestinal health and allergy prevention or treatment.It is of great significance for the rational development and application of 2'-FL products to improve the health of infants and young children.1.Regulation of 2'-FL in the ?-lactoglobulin(?-Lg)-induced allergy in vivoTo establish a model of mice induced by ?-Lg and to analyze the effect of 2'-FL intervention on anaphylaxis induced by ?-Lg and its possible mechanism.They were divided into the normal group,the ?-Lg allergy group(10 mg/kg BW)and the HMO intervention group(gavaged 400mg/kg BW).Low dose 2'-FL intervention group(gavaged 200mg/kg BW);medium dose 2'-FL intervention group(gavaged 400mg/kg BW);high dose 2'-FL intervention group(gavaged 600mg/kg BW).The results showed that mice with type-lg allergy showed allergic symptoms,significant weight loss,increased mast cells,increased serum specific IgE level and other pathological characteristics,which were relieved by HMO and 2'-FL intervention.Compared with the normal group,the expression levels of TLR4/NF-?B pathway mRNA and mir-146a were up-regulated in the allergy group,and the HMO,medium and high dose 2'-FL intervention significantly inhibited the hyperexpression of TLR4/NF-?B pathway and miR-146a induced by allergy.Serum levels of IL-1?,IL-6 and TNF-? in mice with ?-Lg allergy were significantly increased compared with those in the normal group,and the expressions of HMO,medium and high-dose 2'-FL intervention group were lower than those in the allergy group.2.Regulation of 2'-FL in the ?-Lg-induced allergy in vitroThe model of ?-Lg-induced macrophage allergy was established to analyze the effect of 2'-FL intervention on ?-Lg-induced allergic reaction and its possible mechanism.They were divided into normal group,LPS control group(1 ?g/mL),?-Lg allergy group(1 mg/mL),and HMO intervention group(200 ?g/mL).Low dose 2'-FL intervention group(100 ?g/mL);medium dose 2'-FL intervention group(200 ?g/mL);high dose 2'-FL intervention group(400 ?g/mL).The results showed that the levels of NO release,iNOS mRNA expression and proinflammatory cytokines in the ?-Lg allergy group were significantly increased,and there was NO significant difference between the two groups and the LPS positive control group,suggesting that the allergy caused macrophage activation,and the TLR4/NF-?B pathway was significantly increased in the allergy group.2'-FL and HMO intervention effectively reduced the level of macrophage activation and cytokine expression,and 2'-FL and HMO significantly inhibited the up-regulation of TLR4/NF-?B pathway caused by ?-Lg allergy.3.Role of miR-146a in the mechanism of 2'-FL relieving ?-Lg allergyThe model of induced macrophage allergy was established,and it was found that the expression of miR-146a increased in macrophage cells with ?-Lg allergy,and the expression of miR-146a decreased with the remission of allergy after 2'-FL and HMO intervention.MiR-146a inhibitor transfection was performed on the allergic cells,and the effect of miR-146a on the mechanism of ?-Lg allergy was analyzed with HMO intervention.The results showed that after transfection with miR-146a inhibitor,the expression level of miR-146a in macrophages treated with ?-Lg allergy decreased by about 6 times compared with the negative control group,indicating the efficacy of miR-146a inhibitor transfection.NO release,cytokine levels and mRNA expression levels of IRAK1,TRAF6 and NF-?B were significantly higher in the treatment group than those in the treatment group.In addition,inhibitor transfection partially eliminated the inhibition of 2'-FL on macrophage activation and cytokine production.The above results indicate that 2'-FL can effectively alleviate the inflammatory response caused by the ?-Lg allergy,and may play an anti-allergy role by inhibiting the TLR4/NF-?B pathway and regulating the expression of miR-146a.In addition,the anti-allergic effect of 2'-FL at the same dose is better than that of HMO,which indicates that 2'-FL has the application prospect as the active substance to relieve milk allergy,and provides a new idea for the prevention and treatment of milk allergy.