Feeding mice in early life a diet containing an experimental infant milk formula (Nuturis®; eIMF), with a lipid structure similar to human milk, transiently lowered body weight (BW) and fat mass gain upon Western-style diet later in life, when compared with mice fed diets based on control IMF (cIMF). We tested the hypothesis that early-life eIMF feeding alters the absorption or the postabsorptive trafficking of dietary lipids in later life. Male C57BL/6JOlaHsd mice were fed eIMF/cIMF from postnatal day 16–42, followed by low- (LFD, American Institute of Nutrition (AIN)-93 G, 7 wt% fat) or high-fat diet (HFD, D12451, 24 wt% fat) until day 63–70. Lipid absorption rate and tissue concentrations were determined after intragastric administration of stable isotope (2H or 13C) labelled lipids in separate groups. Lipid enrichments in plasma and tissues were analysed using GC-MS. The rate of triolein absorption was similar between eIMF and cIMF fed LFD: 3·2 (sd 1·8) and 3·9 (sd 2·1) and HFD: 2·6 (sd 1·7) and 3·8 (sd 3·0) % dose/ml per h. Postabsorptive lipid trafficking, that is, concentrations of absorbed lipids in tissues, was similar in the eIMF and cIMF groups after LFD. Tissue levels of absorbed TAG after HFD feeding were lower in heart (–42 %) and liver (–46 %), and higher in muscle (+81 %, all P < 0·05) in eIMF-fed mice. In conclusion, early-life IMF diet affected postabsorptive trafficking of absorbed lipids after HFD, but not LFD. Changes in postabsorptive lipid trafficking could underlie the observed lower BW and body fat accumulation in later life upon a persistent long-term obesogenic challenge.

The impact of cream processing on milk fat globule membrane (MFGM) was assessed in an industrial setting for the first time. Three creams and their derived MFGM fractions from different stages of the pasteurization procedure at a butter dairy were investigated and compared to a native control as well as a commercial MFGM fraction. The extent of cross-linking of serum proteins to MFGM proteins increased progressively with each consecutive pasteurization step. Unresolved high molecular weight aggregates were found to consist of both indigenous MFGM proteins and β-lactoglobulin as well as αs1- and β-casein. With regards to fat globule stability and in terms of resistance towards coalescence and flocculation after cream washing, single-pasteurized cream exhibited reduced sensitivity to cream washing compared to non- and double-pasteurized creams. Inactivation of the agglutination mechanism and the increased presence of non-MFGM proteins may determine this balance between stable and non-stable fat globules.

Breast-feeding is associated with a lower risk of developing obesity during childhood and adulthood compared with feeding infant milk formula (IMF). Previous studies have shown that an experimental IMF (eIMF; comprising Nuturis®) programmed mouse pups for a lower body weight and fat mass gain in adulthood when challenged with a high-fat diet (HFD) compared with a control IMF (cIMF). Nuturis has a lipid composition and structure more similar to breast milk. Here, the long-term effects were tested of a similar eIMF, but with an adapted lipid composition and a cIMF, on body weight, glucose homoeostasis, liver and adipose tissue. Nutrient composition was similar for the eIMF and cIMF; the lipid fractions comprised approximately 50 % milk fat. C57BL/6JOlaHsd mice were fed cIMF or eIMF from postnatal day (PN) 16–42 followed by an HFD until PN168. Feeding eIMF v. cIMF in early life resulted in a lower body weight (–9 %) and body fat deposition (–14 %) in adulthood (PN105). The effect appeared transient, as from PN126 onwards, after 12 weeks’ HFD, eIMF-fed mice caught up on controls and body and fat weights became comparable between groups. Glucose and energy metabolism were similar between groups. At dissection (PN168), eIMF-fed mice showed larger (+27 %) epididymal fat depots and a lower (–26 %) liver weight without clear morphological aberrations. Our data suggest the size and coating but not the lipid composition of IMF fat globules underlie the programming effect observed. Prolonged exposure to an HFD challenge partly overrules the programming effect of early diet.

In this Research Communication we present a study of the effect of Ca-binding salts on the recovery of milk fat globule membrane (MFGM) from buttermilk. Sodium phosphate buffer was used for the purpose of MFGM recovery from buttermilk for the first time and we showed that 0.1 M buffer at pH 7.2 was the most effective for the recovery of MFGM. The fact of high efficacy of sodium phosphate buffer in recovery of MFGM from buttermilk allowed us to suggest that MFGM in buttermilk is present in association with casein through Ca- bridges formed between phospholipids of MFGM and phosphate groups of casein, primarily with k-casein as the peripheral protein of casein micelles.

CVD and associated metabolic diseases are linked to chronic inflammation, which can be modified by diet. The objective of the present study was to determine whether there is a difference in inflammatory markers, blood metabolic and lipid panels and lymphocyte gene expression in response to a high-fat dairy food challenge with or without milk fat globule membrane (MFGM). Participants consumed a dairy product-based meal containing whipping cream (WC) high in saturated fat with or without the addition of MFGM, following a 12 h fasting blood draw. Inflammatory markers including IL-6 and C-reactive protein, lipid and metabolic panels and lymphocyte gene expression fold changes were measured using multiplex assays, clinical laboratory services and TaqMan real-time RT-PCR, respectively. Fold changes in gene expression were determined using the Pfaffl method. Response variables were converted into incremental AUC, tested for differences, and corrected for multiple comparisons. The postprandial insulin response was significantly lower following the meal containing MFGM (P < 0·01). The gene encoding soluble epoxide hydrolase (EPHX2) was shown to be more up-regulated in the absence of MFGM (P = 0·009). Secondary analyses showed that participants with higher baseline cholesterol:HDL-cholesterol ratio (Chol:HDL) had a greater reduction in gene expression of cluster of differentiation 14 (CD14) and lymphotoxin β receptor (LTBR) with the WC+MFGM meal. The protein and lipid composition of MFGM is thought to be anti-inflammatory. These exploratory analyses suggest that addition of MFGM to a high-saturated fat meal modifies postprandial insulin response and offers a protective role for those individuals with higher baseline Chol:HDL.

The process of agglutination causes firm cream layers in bovine milk, and a functioning agglutination mechanism is paramount to the quality of non-homogenized milks. The phenomenon is not well-described, but it is believed to occur due to interactions between immunoglobulins (Ig) and milk fat globules. For the first time, this paper demonstrates how the process of agglutination can be visualized using confocal laser scanning microscopy, rhodamine red and a fluoresceinisothiocynat-conjugated immunoglobulin M antibody. The method was used to illustrate the effect on agglutination of storage temperature and pasteurization temperature. Storage at 5 °C resulted in clearly visible agglutination which, however, was markedly reduced at 15 °C. Increasing storage temperature to 20 or 37 °C cancelled any detectable interaction between IgM and milk fat globules, whereby the occurrence of cold agglutination was documented. Increasing 20 s pasteurization temperatures from 69 °C to 71 °C and further to 73 °C lead to progressively higher inactivation of IgM and, hence, reduction of agglutination. Furthermore, 2-dimensional sodium dodecyl sulfate polyacrylamide gel electrophoresis showed that changes in storage temperature caused a redistribution of Ig-related proteins in milk fat globule membrane isolates. Poly-immunoglobulin G receptor was present in milk fat globule preparations stored at cold (4 °C) conditions, but absent at storage at higher temperature (25 °C). The findings provide valuable knowledge to dairy producers of non-homogenized milk in deciding the right pasteurization temperature to retain the crucial agglutination mechanism.

We conducted a randomised, double-blind, placebo-controlled trial to elucidate the effects of dietary milk fat globule membrane (MFGM) on the physical performance of community-dwelling Japanese adults. For this 24-week study, 115 middle-aged subjects (range 50–70 years old) were invited, of whom 113 (seventy-two women, forty-one men) completed the trial. Participants were then divided into either the placebo control or MFGM group. Measurements of physical performance (without undertaking any mandatory exercise) examining muscle strength, agility and balance were tested every 6 weeks until 24 weeks. Analyses were performed using the intention-to-treat method for all participants. Although the effects of MFGM on muscle strength and agility were not significant, we noted that the parameter for balance (such as the ability to stand on one leg with eyes closed for longer durations) increased in the MFGM group (mean 10·1 (95 % CI 8·25, 12·4) s) compared with the placebo (mean 7·53 (95 % CI 6·11, 9·30) s) (P = 0·046). Similarly, application of the mixed-effect model for repeated measures under unstructured covariance also revealed that the effect of MFGM was significant when compared with the placebo (10·2 (95 % CI 8·33, 12·4) v. 7·61 (95 % CI 6·17, 9·30) s) (P = 0·045). In conclusion, we demonstrated that MFGM had an effect on the physical performance of community-dwelling Japanese adults despite mandatory exercise. However, studies using larger cohorts of individuals from different demographic backgrounds are required to further elucidate the mechanisms underlying these effects and to extend the application of MFGM.

Prevalence of type 2 diabetes mellitus (T2DM) is rapidly increasingly and is a key risk for CVD development, now recognised as the leading cause of death globally. Dietary strategies to reduce CVD development include reduction of saturated fat intake. Milk and dairy products are the largest contributors to dietary saturated fats in the UK and reduced consumption is often recommended as a strategy for risk reduction. However, overall evidence from prospective cohort studies does not confirm a detrimental association between dairy product consumption and CVD risk. The present review critically evaluates the current evidence on the association between milk and dairy products and risk of CVD, T2DM and the metabolic syndrome (collectively, cardiometabolic disease). The effects of total and individual dairy foods on cardiometabolic risk factors and new information on the effects of the food matrix on reducing fat digestion are also reviewed. It is concluded that a policy to lower SFA intake by reducing dairy food consumption to reduce cardiometabolic disease risk is likely to have limited or possibly negative effects. There remain many uncertainties, including differential effects of different dairy products and those of differing fat content. Focused and suitably designed and powered studies are needed to provide clearer evidence not only of the mechanisms involved, but how they may be beneficially influenced during milk production and processing.

The present study examined the effect of milk phospholipids (milk-PL) on lipid metabolism and on other risk factors for CVD, in comparison with milk fat (control) or soya phospholipids (soya-PL), respectively. Two double-blind parallel-group intervention trials were conducted in overweight or obese male subjects. In the first trial (trial 1), sixty-two men consumed milk enriched with either 2 g milk-PL or 2 g milk fat (control) for 8 weeks. In trial 2, fifty-seven men consumed milk enriched with either 3 g milk-PL or 2·8 g soya-PL for 7 weeks. In trial 1, milk-PL as compared with control reduced waist circumference but did not affect plasma lipids (total, HDL- and LDL-cholesterol, total cholesterol:HDL-cholesterol ratio, TAG, phospholipids), apoB, apoA1, glucose, insulin, insulin sensitivity index, C-reactive protein, IL-6, soluble intracellular adhesion molecule and total homocysteine (tHcy). Serum activities of alanine transaminase and aspartate transaminase were not changed. Activity of γ-glutamyl transferase (GGT), a marker of fatty liver, increased in the control but not in the milk-PL group, with a significant intervention effect. In trial 2, milk-PL as compared with soya-PL did not affect the above-mentioned parameters, but decreased GGT. Subjects with the methylenetetrahydrofolate reductase mutations CT and TT had 11 % (P < 0·05) higher baseline tHcy concentrations than those with the wild-type CC. However, genotype did not modulate the phospholipid intervention effect on tHcy. In conclusion, supplementation with milk-PL as compared with control fat reduced waist circumference and, as compared with both control fat and soya-PL, GGT activity.

Epidemiological studies have demonstrated protective effects of breast-feeding on childhood obesity. Differences between human milk and infant milk formula (IMF) in dietary lipid structure may contribute to this effect. In our mouse model, feeding a diet containing large lipid droplets coated with phospholipids (PL) (Nuturis®; PL of milk fat globule membrane (MFGM) fraction origin) in early life protected against excessive body fat accumulation following a diet challenge in adult life. We now set out to determine the relevance of increased droplet size and/or MFGM lipid droplet coating to the observed anti-obesogenic effects in adult life. From day 16 to 42, male mouse pups were exposed to diets with small (S) or large (L) lipid droplets (0·3 v. 2·9 µm average mode diameter, respectively), either without MFGM or with MFGM coating around the lipid droplet, resulting in four groups: S (control diet), L, Scoating and Lcoating (Nuturis® IMF diet). Mice were subsequently challenged with a Western-style diet until dissection at postnatal day 98. A non-challenged group served as reference (REF). We repeatedly determined body composition between postnatal day 42 and 98. At day 98 plasma and gene expression measurements were performed. Only the Nuturis® IMF diet (Lcoating) in early life containing MFGM-coated large lipid droplets reduced body fat mass to a level comparable with the REF group. These data support the notion that the structural aspects of lipids in human milk, for example, both lipid droplet size as well as the MFGM coating, may contribute to its reported protective effect against obesity in later life.

Meals high in SFA, particularly palmitate, are associated with postprandial inflammation and insulin resistance. Milk fat globule membrane (MFGM) has anti-inflammatory properties that may attenuate the negative effects of SFA-rich meals. Our objective was to examine the postprandial metabolic and inflammatory response to a high-fat meal composed of palm oil (PO) compared with PO with an added dairy fraction rich in MFGM (PO+MFGM) in overweight and obese men and women (n 36) in a randomised, double-blinded, cross-over trial. Participants consumed two isoenergetic high-fat meals composed of a smoothie enriched with PO with v. without a cream-derived complex milk lipid fraction ( dairy fraction rich in MFGM) separated by a washout of 1–2 weeks. Serum cytokines, adhesion molecules, cortisol and markers of inflammation were measured at fasting, and at 1, 3 and 6 h postprandially. Glucose, insulin and lipid profiles were analysed in plasma. Consumption of the PO + MFGM v. PO meal resulted in lower total cholesterol (P = 0·021), LDL-cholesterol (P = 0·046), soluble intracellular adhesion molecule (P = 0·005) and insulin (P = 0·005) incremental AUC, and increased IL-10 (P = 0·013). Individuals with high baseline C-reactive protein (CRP) concentrations (≥3 mg/l, n 17) had higher (P = 0·030) insulin at 1 h after the PO meal than individuals with CRP concentrations <3 mg/l (n 19). The addition of MFGM attenuated this difference between CRP groups. The addition of a dairy fraction rich in MFGM attenuated the negative effects of a high-SFA meal by reducing postprandial cholesterol, inflammatory markers and insulin response in overweight and obese individuals, particularly in those with elevated CRP.

While the Complement protein system in human milk is well characterised, there is little information on its presence and activity in bovine milk. Complement forms part of the innate immune system, hence the importance of its contribution during milk ingestion to the overall defences of the neonate. A bactericidal sequestration assay, featuring a Complement sensitive strain, Escherichia coli 0111, originally used to characterise Complement activity in human milk was successfully applied to freshly drawn bovine milk samples, thus, providing an opportunity to compare Complement activities in both human and bovine milks. Although not identical in response, the levels of Complement activity in bovine milk were found to be closely comparable with that of human milk. Differential counts of Esch. coli 0111 after 2 h incubation were 6·20 and 6·06 log CFU/ml, for raw bovine and human milks, respectively – the lower value representing a stronger Complement response. Exposing bovine milk to a range of thermal treatments e.g. 42, 45, 65, 72, 85 or 95 °C for 10 min, progressively inhibited Complement activity by increasing temperature, thus confirming the heat labile nature of this immune protein system. Low level Complement activity was found, however, in 65 and 72 °C heat treated samples and in retailed pasteurised milk which highlights the outer limit to which high temperature, short time (HTST) industrial thermal processes should be applied if retention of activity is a priority. Concentration of Complement in the fat phase was evident following cream separation, and this was also reflected in the further loss of activity recorded in low fat variants of retailed pasteurised milk. Laboratory-based churning of the cream during simulated buttermaking generated an aqueous (buttermilk) phase with higher levels of Complement activity than the fat phase, thus pointing to a likely association with the milk fat globule membrane (MFGM) layer.

The bovine milk fat globule membrane (MFGM) contains several antimicrobial components with proven efficacy in vitro, but in vivo evidence is scarce. The present study was performed to determine the efficacy of the bovine MFGM in vivo. Rats were fed diets based on bovine skimmed milk powder (low in MFGM) or bovine sweet buttermilk powder (high in MFGM). After dietary adaptation, rats were orally infected with Salmonella enteritidis or Listeria monocytogenes. Whereas sweet buttermilk powder did not protect rats against infection with S. enteritidis, it protected against L. monocytogenes, as shown by a lower colonisation and translocation of this pathogen. Protection coincided with higher listericidal capacity of gastric and caecal contents. The digestion products of phosphoglycerides and sphingomyelin are bactericidal in vitro. To study their role, rats were fed diets containing either 0·1 % phosphatidylcholine or sphingomyelin, or a control diet. After dietary adaptation, rats were infected with L. monocytogenes. Since Listeria colonisation was not affected by these diets, phosphoglycerides and sphingomyelin are not involved in the protective effect of sweet buttermilk. Additional in vitro experiments were performed to further explore the mechanism of the beneficial effects of sweet buttermilk. Inhibition of the adherence of L. monocytogenes to the intestinal mucosa is the most likely explanation, since sweet buttermilk powder inhibited the binding of L. monocytogenes in both a haemagglutination assay and a Caco-2 cell adherence assay. In conclusion, sweet buttermilk powder, which is rich in MFGM, protects against L. monocytogenes infection in rats, probably by preventing adherence of this pathogen to the intestinal mucosa.

Milk fat globule membranes (MFGM) were isolated from the milk of mid-lactation Holstein cows. The purified MFGM were fractionated using 1-dimensional SDS gels. Tryptic peptides from gel slices were further fractionated on a micro-capillary high performance liquid chromatograph connected to a nanospray-tandem mass spectrometer. Analysis of the data resulted in 120 proteins being identified by two or more unique peptide sequences. Of these 120 proteins, 71% are membrane associated proteins with the remainder being cytoplasmic or secreted proteins. Only 15 of the proteins identified in the cow MFGM were the same as proteins identified in previous mouse or human MFGM proteomic studies. Thus, the bulk of the proteins identified are new for bovine MFGM proteomics. The proteins identified were associated with membrane/protein trafficking (23%), cell signalling (23%), unknown functions (21%), fat transport/metabolism (11%), transport (9%), protein synthesis/folding (7%), immune proteins (4%) and milk proteins (2%). The proteins associated with cell signalling or membrane/protein trafficking may provide insights into MFGM secretion mechanisms. The finding of CD14, toll like receptor (TLR2), and TLR4 on MFGM suggests a direct role for the mammary gland in detecting an infection.