Quantitative Label-Free Proteomic Analysis of Milk Fat Globule Membrane in Donkey and Human Milk.
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ABSTRACT:
SUBMITTER: Zhang X
PROVIDER: S-EPMC8258387 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Intake of high-fat foods raises postprandial plasma triglycerides and inflammatory markers, which may depend on the type of fat ingested. Dairy products are commonly consumed, but not much is known about the impact of milk fat and the milk fat globule membrane on postprandial inflammation. Here, we aimed to study the effect of milk fat with and without milk fat globule membrane and vegetable blend fat on post-prandial inflammation, with a focus on blood monocyte gene expression. We performed a randomized, double-blind cross-over trial in middle-aged healthy male and female volunteers (BMI 22–27 kg/m2). The participants consumed a meal shake containing 95.5 g of fat consisting of either a vegetable fat blend (VEGE), anhydrous milk fat (AMF, without milk fat globule membrane), or cream (CREAM, containing milk fat globule membrane). Blood monocytes were collected at 0 and 6 hours postprandially and used for bulk RNA sequencing and ex vivo stimulation with LPS. Consumption of all three shakes significantly decreased the percentage of classical monocytes and increased the percentages of intermediate monocytes and non-classical monocytes. No differences in these measures were observed between shakes. Using a threshold of p < 0.01, 787 genes were differentially regulated postprandially between the three shakes. 89 genes were differentially regulated postprandially between AMF and VEGE, 373 genes between AMF and CREAM, and 667 genes between VEGE and CREAM, indicating that the effect of CREAM on monocyte gene expression was distinct from AMF and VEGE. Pathway analyses showed that VEGE significantly increased the expression of genes involved in inflammatory pathways, whereas this was less pronounced after AMF and not observed after CREAM. In addition, CREAM significantly down-regulated the expression of genes involved in energy metabolism-related pathways, such as glycolysis, TCA cycle, and oxidative phosphorylation, as well as HIF1 signaling. Compared to the consumption of an anhydrous milk fat without milk fat globule membrane and a vegetable fat blend, the consumption of cream with milk fat globule membrane downregulated inflammatory pathways in blood monocytes, thus suggesting a potential inflammation inhibitory effect of milk fat globule membrane.
2023-12-09 | GSE249787 | GEO
Project description:Proteomic analysis of membrane proteins is challenged by the proteins solubility and detergent incompatibility with MS analysis. No single perfect protocol can be used to comprehensively characterize the proteome of membrane fraction. Here, we used cow milk fat globule membrane (MFGM) proteome analysis to assess six sample preparation procedures including one in-gel and five in-solution digestion approaches prior to LC-MS/MS analysis. The largest number of MFGM proteins were identified by suspension trapping (S-Trap) and filter-aided sample preparation (FASP) methods, followed by acetone precipitation without clean-up of tryptic peptides method. Protein identifications with highest average coverage was achieved by Chloroform/MeOH, in-gel and S-Trap methods. Most distinct proteins were identified by FASP method, followed by S-Trap. Analyses by Venn diagram, principal-component analysis, hierarchical clustering and the abundance ranking of quantitative proteins highlight differences in the MFGM fraction by the all sample preparation procedures. These results reveal the biased proteins/peptides loss occurred in each protocol. In this study, we found several novel proteins that were not observed previously by in-depth proteomics characterization of MFGM fraction in milk. Thus, a combination of multiple procedures with orthologous properties of sample preparation was demonstrated to improve the protein sequence coverage and expression level accuracy of membrane samples.
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Project description:Gradient-polyacrylamide-gel electrophoresis of human milk serum separated three high-Mr glycoprotein bands. The properties of the components corresponding to the three bands (tentatively termed 'Components C, B and A' in their order of migration) were compared by staining with four monoclonal antibodies and lectins. Components B and C both reacted with the four antibodies, but Component A did not. Components B and C were stained with peanut (Arachis hypogaea) agglutinin (PNA) and wheat (Triticum)-germ agglutinin (WGA), Component A being stained with soya-bean (Glycine max) agglutinin as well as PNA and WGA. These results suggest that Components B and C were related molecules, whereas Component A was markedly different from them. The reactivities of Components B and C were the same as those of PAS-0, a high-Mr periodate/Schiff (PAS)-positive glycoprotein previously isolated from human milk fat-globule membrane (MFGM). Component C, whose electrophoretic mobility was the same as PAS-0, could have been a soluble form of PAS-0. A high-Mr glycoprotein having the same properties as Component A was also observed in MFGM. The amino acid composition of the isolated Component A was significantly different from that of Component C and PAS-0, high threonine and serine contents being characteristic of Component A. The carbohydrate content of Component A was 65-80%, and was much higher than that of Component C and PAS-0. Fucose, galactose, N-acetylglucosamine, N-acetylgalactosamine and sialic acid were each detected as constituent sugars of Component A. Component A represents, therefore, a new high-Mr glycoprotein species in human milk serum and MFGM. Since these glycoproteins were high-Mr mucin-like glycoproteins, the names 'HM glycoprotein-A' and 'HM glycoprotein-C' were proposed for Component A and Component C (PAS-O) respectively.
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Project description:A major periodate--Schiff-positive component from milk-fat-globule membrane of human breast milk has been purified by selectively extracting the membrane glycoproteins, followed by lectin affinity chromatography and gel filtration on Sephadex G-200 in the presence of protein-dissociating agents. The purified glycoprotein, termed epithelial membrane glycoprotein (EMGP-70), has an estimated mol.wt. of 70 000 and yields a single band under reducing conditions on sodium dodecyl sulphate/polyacrylamide-gel electrophoresis. The glycoprotein contains 13.5% carbohydrate by weight, with fucose, mannose, galactose, N-acetylglucosamine and sialic acid 17.2, 17.0, 21.1, 7.9 and 36.6% respectively of the carbohydrate moiety. Aspartic and glutamic acid and serine are the major amino acid residues.
| S-EPMC1162574 | biostudies-other
Project description:The aim of this study was to investigate if milk fat globule membrane (MFGM) enclosing the dairy fat influence peripheral blood mononuclear cells (PBMC) gene expression. This study was a 8-week single-blind, randomized, controlled isocaloric trial with two parallel groups including overweight (mean BMI: 28) adult women (n=30). All subjects consumed 40 g dairy fat per day either as cream (MFGM diet) or as butter oil (control diet). MFGM diet and control diet showed differential responses in plasma cholesterol. Absolute differences in gene expression were calculated for each gene in each subject, comparing after with before the intervention. These absolute differences in gene expression were compared between intervention groups with T-test and with Significance Analysis of Microarrays (SAM). 19 genes were significantly different regulated by the two diets with a FDR of 23%. Most of thechanges in gene expression correlated with changes in blood lipids. Patients were examined in the morning after 12 h's fasting. Whole blood was obtained before (W0) and after completion (W8) of the dietary intervention for isolation of PBMCs and RNA extraction. From total RNA we prepared and hybridised biotinylated complementary RNA to GeneChip Human Gene 1.1 ST Arrays (Affymetrix Inc., Santa Clara, CA), and then washed, stained and scanned the slides using standardised protocols (Affymetrix Inc.). Signicance analysis of microarrays (SAM) was use to compare the difference in gene expression between groups.
2015-08-01 | E-GEOD-60148 | biostudies-arrayexpress
Project description:The human milk fat globule membrane (MFGM) contains important lipids for growing infants. Anthropometric measurements, milk samples, and infant milk intake were collected in a cohort of eleven healthy mother-infant dyads during exclusive breastfeeding from birth to six months. One hundred and sixty-six MFGM lipids were analysed using liquid chromatography-mass spectrometry, and the infant intake was calculated. The concentrations and intake were compared and associations between infant intake and growth characteristics explored. The lipid concentrations and infant intake varied widely between mother-infant dyads and between months one and three. The infant intake for many species displayed positive correlations with infant growth, particularly phospholipid species. The high variation in lipid intake is likely an important factor in infant growth, with strong correlations identified between the intake of many MFGM lipids and infant head circumference and weight. This study highlights the need for intake measurements and inclusion in cohort studies to elucidate the role of the human milk lipidome in infant growth and development.
| S-EPMC8471100 | biostudies-literature
Project description:(1) Background: Milk fat globule membrane (MFGM), composing fat droplets responsible for lipid transport in breast milk, has been shown to possess immunological and antimicrobial effects. Standard formulas (SF) are devoid of MFGMs during the production process. The study's aim was to evaluate the safety and benefits of MFGMs supplementation in children. (2) Methods: We searched four databases for randomized controlled trials evaluating the supplementation of MFGMs in children. Growth parameters were chosen as the primary outcome. (3) Results: Twenty-four publications of seventeen studies were included. Meta-analyses assessing the primary outcomes at the age of 4 months included four studies (814 children) comparing the MFGM-supplemented formulas and SF, and two trials (549 children) comparing the MFGM-supplemented formulas and breastfeeding. The primary outcomes were non-inferior in all the experimental MFGM formulas compared to SF, or even represented more similar results to breastfed infants. The promising effects, including a lower incidence of acute otitis media and improved cognitive development, cannot be firmly confirmed due to the small amount of existing evidence. No significant adverse effects were reported in any of the assessed products. (4) Conclusions: The available data signaled beneficial effects and a good safety profile, requiring future research with well-designed trials.
| S-EPMC7996302 | biostudies-literature
Project description:The aim of this study was to investigate if milk fat globule membrane (MFGM) enclosing the dairy fat influence peripheral blood mononuclear cells (PBMC) gene expression. This study was a 8-week single-blind, randomized, controlled isocaloric trial with two parallel groups including overweight (mean BMI: 28) adult women (n=30). All subjects consumed 40 g dairy fat per day either as cream (MFGM diet) or as butter oil (control diet). MFGM diet and control diet showed differential responses in plasma cholesterol. Absolute differences in gene expression were calculated for each gene in each subject, comparing after with before the intervention. These absolute differences in gene expression were compared between intervention groups with T-test and with Significance Analysis of Microarrays (SAM). 19 genes were significantly different regulated by the two diets with a FDR of 23%. Most of thechanges in gene expression correlated with changes in blood lipids.
2015-08-01 | GSE60148 | GEO
Project description:The molecular mechanism underlying milk fat globule secretion in mammary epithelial cells ostensibly involves the formation of complexes between plasma membrane butyrophilin and cytosolic xanthine oxidoreductase. These complexes bind adipophilin in the phospholipid monolayer of milk secretory granules, the precursors of milk fat globules, enveloping the nascent fat globules in a layer of plasma membrane and pinching them off the cell. However, using freeze-fracture immunocytochemistry, we find these proteins in locations other than those previously inferred. Significantly, butyrophilin in the residual plasma membrane of the fat globule envelope is concentrated in a network of ridges that are tightly apposed to the monolayer derived from the secretory granule, and the ridges coincide with butyrophilin labeling in the globule monolayer. Therefore, we propose that milk fat globule secretion is controlled by interactions between plasma membrane butyrophilin and butyrophilin in the secretory granule phospholipid monolayer rather than binding of butyrophilin-xanthine oxidoreductase complexes to secretory granule adipophilin.
| S-EPMC1502467 | biostudies-literature
Project description:During lactation, mammary epithelial cells secrete huge amounts of milk from their apical side. The current view is that caseins are secreted by exocytosis, whereas milk fat globules are released by budding, enwrapped by the plasma membrane. Owing to the number and large size of milk fat globules, the membrane surface needed for their release might exceed that of the apical plasma membrane. A large-scale proteomics analysis of both cytoplasmic lipid droplets and secreted milk fat globule membranes was used to decipher the cellular origins of the milk fat globule membrane. Surprisingly, differential analysis of protein profiles of these two organelles strongly suggest that, in addition to the plasma membrane, the endoplasmic reticulum and the secretory vesicles contribute to the milk fat globule membrane. Analysis of membrane-associated and raft microdomain proteins reinforces this possibility and also points to a role for lipid rafts in milk product secretion. Our results provide evidence for a significant contribution of the endoplasmic reticulum to the milk fat globule membrane and a role for SNAREs in membrane dynamics during milk secretion. These novel aspects point to a more complex model for milk secretion than currently envisioned.
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