Milk fat globule membrane supplementation protects againstβ-lactoglobul-ininduced food allergy in mice via upregulation of regulatory T cells and enhancement of intestinal barrier in a microbiota-derived short-chain fatty acids manner

Milk fat globule membrane(MFGM),which contains abundant glycoproteins and phospholipids,exerts beneficial effects on intestinal health and immunomodulation.The aim of this study was to evaluate the protective effects and possible underlying mechanisms of MFGM on cow’s milk allergy(CMA)in aβ-lactoglobulin(BLG)-induced allergic mice model.MFGM was supplemented to allergic mice induced by BLG at a dose of 400 mg/kg body weight.Results demonstrated that MFGM alleviated food allergy symptoms,decreased serum levels of lipopolysaccharide,pro-inflammatory cytokines,immunoglobulin(Ig)E,Ig G1,and Th2 cytokines including interleukin(IL)-4,while increased serum levels of Th1 cytokines including interferon-γand regulatory T cells(Tregs)cytokines including IL-10 and transforming growth factor-β.MFGM modulated gut microbiota and enhanced intestinal barrier of BLG-allergic mice,as evidenced by decreased relative abundance of Desulfobacterota,Rikenellaceae,Lachnospiraceae,and Desulfovibrionaceae,while increased relative abundance of Bacteroidetes,Lactobacillaceae and Muribaculaceae,and enhanced expressions of tight junction proteins including Occludin,Claudin-1 and zonula occludens-1.Furthermore,MFGM increased fecal short-chain fatty acids(SCFAs)levels,which elevated G protein-coupled receptor(GPR)43 and GPR109A expressions.The increased expressions of GPR43 and GPR109A induced CD103+dendritic cells accumulation and promoted Tregs differentiation in mesenteric lymph node to a certain extent.In summary,MFGM alleviated CMA in a BLG-induced allergic mice model through enhancing intestinal barrier and promoting Tregs differentiation,which may be correlated with SCFAs-mediated activation of GPRs.These findings suggest that MFGM may be useful as a promising functional ingredient against CMA.

Changes in milk fat globule membrane proteins along lactation stage of Laoshan dairy goat

The milk fat globule membrane(MFGM)is a complex structure with numerous functions,and its composition is affected by many factors.There have been few systematic investigations on goat MFGM proteome profiling during lactation.Individual milk samples from 15 healthy dairy goats were obtained at six lactation time points for investigation of the MFGM proteome using both data-independent acquisition(DIA)and data-dependent acquisition(DDA)proteomics techniques combined with multivariate statistical analysis.Using the DIA method,890 variably abundant MFGM proteins were discovered throughout the lactation cycle.From 1 to 240 d,butyrophilin subfamily 1 member A1,lipoprotein lipase,perilipin-2,and adipose triglyceride lipase were upregulated,while APOE,complement C3,clusterin,and IgG were downregulated.Furthermore,from 1 to 90 d,annexin A1,annexin A2,and antithrombin-ll were downregulated,then upregulated by d 240.Albumin had a high degree of connectedness,indicating that it was a key protein,according to protein-protein interaction research.Overall,our findings gave new insights into the biological features of MFGM protein in goat milk throughout lactation,which may aid in the creation of specialized MFGM products and infant formula.

Milk fat globule epidermal growth factor 8 alleviates liver injury in severe acute pancreatitis by restoring autophagy flux and inhibiting ferroptosis in hepatocytes

BACKGROUND Liver injury is common in severe acute pancreatitis(SAP).Excessive autophagy often leads to an imbalance of homeostasis in hepatocytes,which induces lipid peroxidation and mitochondrial iron deposition and ultimately leads to ferroptosis.Our previous study found that milk fat globule epidermal growth factor 8(MFG-E8)alleviates acinar cell damage during SAP via binding toαvβ3/5 integrins.MFG-E8 also seems to mitigate pancreatic fibrosis via inhibiting chaperone-mediated autophagy.AIM To speculate whether MFG-E8 could also alleviate SAP induced liver injury by restoring the abnormal autophagy flux.METHODS SAP was induced in mice by 2 hly intraperitoneal injections of 4.0 g/kg L-arginine or 7 hly injections of 50μg/kg cerulein plus lipopolysaccharide.mfge8-knockout mice were used to study the effect of MFG-E8 deficiency on SAPinduced liver injury.Cilengitide,a specificαvβ3/5 integrin inhibitor,was used to investigate the possible mechanism of MFG-E8.RESULTS The results showed that MFG-E8 deficiency aggravated SAP-induced liver injury in mice,enhanced autophagy flux in hepatocyte,and worsened the degree of ferroptosis.Exogenous MFG-E8 reduced SAP-induced liver injury in a dose-dependent manner.Mechanistically,MFG-E8 mitigated excessive autophagy and inhibited ferroptosis in liver cells.Cilengitide abolished MFG-E8’s beneficial effects in SAP-induced liver injury.CONCLUSION MFG-E8 acts as an endogenous protective mediator in SAP-induced liver injury.MFG-E8 alleviates the excessive autophagy and inhibits ferroptosis in hepatocytes by binding to integrinαVβ3/5.

Milk fat globule epithelial growth factorⅧ(MFG-E8)sustains survival of cancer cells by prompting tumor angiogenesis and suppressing host immunities

Milk fat globule epithelial growth factor VIII(MFG-E8) is a novel adhesion protein mainly produced by macrophages and dendritic cells; it is expressed in most of the human tissues and functions to prompt cancer progression and survival. MFG-E8 contains a signal sequence for secretion, two epidermal growth factor(EGF)-like domains at the NH2 terminus and two discoidin domains with blood-clotting factor V/factor Ⅷ(C1 and C2) at the COOH terminus. The second EGF domain contains an arginine-glycine-aspartic(RGD) integrin-binding motif that engages α_vβ_5 integrins to facilitate cell adhesion and induce integrinmediated signal transduction. Integrin α_vβ_3 associates with VEGF receptor 2, engagement of integrins can promote angiogenesis, which plays key roles in growth, proliferation, and survival of cancer cells. VEGF stimulates the expression of α_vβ_3 and α_vβ_5 integrins on angiogenic vasculature, thereby potentiating effects of VEGF receptor engagement. Mice expressing a mutant form of α_vβ_3 integrin are unable to undergo tyrosine phosphorylation, confirming the important role that this integrin plays in pathological angiogenesis and providing important mechanistic insights. The C-terminus discoidin-like domains promote binding to membrane phospholipids, functioning close to VEGF like angiogenesis. MFG-E8 is an opsonin for apoptotic cells, and it acts as a bridging protein between apoptotic cells and phagocytes. It also influences cell immunities by altering CD4^+ and/or CD8^+ cells. Antibody or small peptide works with MFG-E8 at different functional sites or interacts with EGF-like domains and/or discoidin-like domains may play an important role in anti-angiogenesis or immune restoration. Altering the structures and/or functions of MFG-E8 and/or its domains is promising for development of novel anti-cancer strategies.

Impact of post-ruminally infused macronutrients on bovine mammary gland expression of genes involved in fatty acid synthesis, energy metabolism, and protein synthesis measured in RNA isolated from milk fat

Background: Characterising the regulation of milk component synthesis in response to macronutrient supply is critical for understanding the implications of nutritional interventions on milk production. Gene expression in mammary gland secretory cells was measured using RNA isolated from milk fat globules from 6 Holstein-Friesian cows receiving 5-d abomasal infusions of saline, essential amino acids(AA), or glucose(GG) or palm olein(LG)without(LAA) or with(HAA) essential AA, according to a 6 × 6 Latin square design. RNA was isolated from milk fat samples collected on d 5 of infusion and subjected to real-time quantitative PCR. We hypothesised that m RNA expression of genes involved in de novo milk fatty acid(FA) synthesis would be differently affected by GG and LG,and that expression of genes regulating transfer of tricarboxylic acid cycle intermediates would increase at the HAA level. We also hypothesised that the HAA level would affect genes regulating endoplasmic reticulum(ER)homeostasis but would not affect genes related to the mechanistic target of rapamycin complex 1(m TORC1) or the integrated stress response(ISR) network.Results: Infusion of GG did not affect de novo milk FA yield but decreased expression of FA synthase(FASN).Infusion of LG decreased de novo FA yield and tended to decrease expression of acetyl-Co A carboxylase 1(ACC1).The HAA level increased both de novo FA yield and expression of ACC1, and tended to decrease expression of mitochondrial phosphoenolpyruvate carboxykinase(PCK2). m RNA expression of m TORC1 signaling participants was not affected by GG, LG, or AA level. Expression of the ε subunit of the ISR constituent eukaryotic translation initiation factor 2 B(EIF2 B5) tended to increase at the HAA level, but only in the presence of LG. X-box binding protein 1(XBP1) m RNA was activated in response to LG and the HAA level.Conclusions: Results show that expression of genes involved in de novo FA synthesis responded to glucogenic,lipogenic, and aminogenic substrates, whereas genes regulating intermediate flux through the tricarboxylic acid cycle were not majorly affected. Results also suggest that after 5 d of AA supplementation, milk protein synthesis is supported by enhanced ER biogenesis instead of signaling through the mTORC1 or ISR networks.

Theoretical Study of the Effect of Multi-Diameter Distribution on the Mie Scattering Characteristics of Milk Fat

To correct the light scattering property parameters of milk fat for improving the detection accuracy,the Mie-theory was used to establish a predictive model for light scattering properties of milk fat globule with multi-diameter distributions, by means of Monte Carlo approach to simulate actual multi-diameter size distribution of milk fat globule in milk fat solution. Scattering coefficient and absorption coefficient of multidiameter distribution milk fat particles were calculated by simulating the particles size distribution in milk fat solution. And the light scattering properties of multi-diameter distribution was compared with that of volume mean diameter,Sauter mean diameter and numerical mean diameter in milk fat solution. Theoretical simulation results indicate that the scattering coefficient and absorption coefficient of milk fat particles are determined by the particle size distribution in milk fat solution. There is a distinct difference in scattering characteristics between the milk fat particles with multi-diameter distribution and that with mean diameters. Compared to that with multi-diameter distribution,the scattering coefficient of the milk fat particles with mean diameter has a maximum mean deviation of 9042 m-1. The particle size distribution is not completely determined by the mean diameters. The dependence of the light scattering properties on the particle size distribution should be considered into the model and simulation. Therefore,it is found that the particle size distribution in milk fat solution is an essential and critical factor to significantly improve the detection accuracy of milk fat content.

Early life administration of milk fat globule membrane promoted SCFA-producing bacteria colonization,intestinal barriers and growth performance of neonatal piglets

Milk fat globule membrane(MFGM)possesses various nutritional and biological benefits for mammals,whereas its effects on neonatal gut microbiota and barrier integrity remained unclear.This study investigated the effects of MFGM administration on microbial compositions and intestinal barrier functions of neonatal piglets.Sixteen newborn piglets were randomly allocated into a CON group or MFGM group,orally administered with saline or MFGM solution(1 g/kg body weight)respectively during the first postnatal week,and all piglets were breastfed during the whole neonatal period.The present study found that the MFGM oral administration during the first postnatal week increased the plasma immunoglobulin(lg)G level,body weight and average daily gain of piglets(P<0.05)on 21 d.Addi-tionally,MFGM administration enriched fecal SCFA-producing bacteria(Ruminococaceae_UCG-002,Ruminococaceae_UCG-010,Ruminococaceae_UCG-004,Ruminococaceae_UCG-014 and[Ruminococcus]_gauvrearuii_group),SCFA concentrations(acetate,propionate and butyrate;P<0.05)and their receptor(G-protein coupled receptor 41,GPR41).Furthermore,MFGM administration promoted intestinal villus morphology(P<0.05)and barrier functions by upregulating genes of tight junctions(E-cadherin,claudin-1,occludin and zonula occludin 1[ZO-1]),mucins(mucin-13 and mucin-20)and interleukin(IL)-22(P<0.05).Positive correlation was found between the beneficial microbes and SCFA levels pairwise with the intestinal barrier genes(P<0.05).In conclusion,orally administrating MFGM during the first postnatal week stimulated SCFA-producing bacteria colonization and SCFA generation,enhanced intes-tinal barrier functions and consequently improved growth performance of neonatal piglets on 21 d.Our findings will provide new insights about MFGM intervention for microbial colonization and intestinal development of neonates during their early life.

Knockout of butyrophilin subfamily 1 member A1(BTN1A1) alters lipid droplet formation and phospholipid composition in bovine mammary epithelial cells

Background: Milk lipids originate from cytoplasmic lipid droplets(LD) that are synthesized and secreted from mammary epithelial cells by a unique membrane-envelopment process. Butyrophilin 1 A1(BTN1 A1) is one of the membrane proteins that surrounds LD, but its role in bovine mammary lipid droplet synthesis and secretion is not well known.Methods: The objective was to knockout BTN1 A1 in bovine mammary epithelial cells(BMEC) via the CRISPR/Cas9 system and evaluate LD formation, abundance of lipogenic enzymes, and content of cell membrane phospholipid(PL) species. Average LD diameter was determined via Oil Red O staining, and profiling of cell membrane phospholipid species via liquid chromatography-tandem mass spectrometry(LC-MS/MS).Results: Lentivirus-mediated infection of the Cas9/sg RNA expression vector into BMEC resulted in production of a homozygous clone BTN1 A1^((-/-)). The LD size and content decreased following BTN1 A1 gene knockout. The m RNA abundance of fatty acid synthase(FASN) and peroxisome proliferator-activated receptor-gamma(PPARG) was downregulated in the BTN1 A1^((-/-))clone. Subcellular analyses indicated that BTN1 A1 and LD were co-localized in the cytoplasm. BTN1 A1 gene knockout increased the percentage of phosphatidylethanolamine(PE) and decreased phosphatidylcholine(PC), which resulted in a lower PC/PE ratio.Conclusions: Results suggest that BTN1 A1 plays an important role in regulating LD synthesis via a mechanism involving membrane phospholipid composition.