Protein Ingredient Quality within Infant Formulas Impacts Plasma Amino Acid Concentrations in Neonatal Minipiglets.

BACKGROUND: Infant formulas (IFs), the only adequate substitute to human milk, are complex matrices that require numerous ingredients and processing steps that may impact protein digestion and subsequent amino acid (AA) absorption. OBJECTIVES: The objective was to understand the impact of the protein ingredient quality within IFs on postprandial plasma AA profiles. METHODS: Four isonitrogenous and isocaloric IFs were produced at a semi-industrial scale using whey proteins from different origins (cheese compared with ideal whey) and denaturation levels (IF-A, -B, -C), and caseins with different supramolecular organizations (IF-C, -D). Ten Yucatan minipiglets (12- to 27-d-old) were used as a human infant model and received each IF for 3 d according to a Williams Latin square followed by a 2-d wash-out period. Jugular plasma was regularly sampled from 10 min preprandial to 4 h postprandial on the third day to measure free AAs, urea, insulin, and glucose concentrations. Data were statistically analyzed using a mixed linear model with diet (IFs), time, and sex as fixed factors and piglet as random factor. RESULTS: IFs made with cheese whey (IF-A and -B) elicited significantly higher plasma total and essential AA concentrations than IFs made with ideal whey (IF-C and -D), regardless of the pre- and postprandial times. Most of the differences observed postprandially were explained by AA homeostasis modifications. IFs based on cheese whey induced an increased plasma concentration of Thr due to both a higher Thr content in these IFs and a Thr-limiting degrading capability in piglets. The use of a nonmicellar casein ingredient led to reduced plasma content of AA catabolism markers (IF-D compared with IF-C). CONCLUSIONS: Overall, our results highlight the importance of the protein ingredient quality (composition and structure) within IFs on neonatal plasma AA profiles, which may further impact infant protein metabolism.

Effect of Docosahexaenoic Acid Encapsulation with Whey Proteins on Rat Growth and Tissue Endocannabinoid Profile.

Modifying the food structure allows a nutrient to be delivered differently, which can modify not only its digestion process but also its subsequent metabolism. In this study, rats received 3 g of omelette daily containing docosahexaenoic acid (DHA) as crude oil or previously encapsulated with whey proteins, whereas a control group received a DHA-free omelette. The results showed that DHA encapsulation markedly induced a different feeding behaviour so animals ate more and grew faster. Then, after four weeks, endocannabinoids and other N-acyl ethanolamides were quantified in plasma, brain, and heart. DHA supplementation strongly reduced endocannabinoid derivatives from omega-6 fatty acids. However, DHA encapsulation had no particular effect, other than a great increase in the content of DHA-derived docosahexaenoyl ethanolamide in the heart. While DHA supplementation has indeed shown an effect on cannabinoid profiles, its physiological effect appears to be mediated more through more efficient digestion of DHA oil droplets in the case of DHA encapsulation. Thus, the greater release of DHA and other dietary cannabinoids present may have activated the cannabinoid system differently, possibly more locally along the gastrointestinal tract. However, further studies are needed to evaluate the synergy between DHA encapsulation, fasting, hormones regulating food intake, and animal growth.

Development of innate immune memory by non-immune cells during Staphylococcus aureus infection depends on reactive oxygen species.

Introduction: The mechanisms underlying innate immune memory (trained immunity) comprise epigenetic reprogramming of transcriptional pathways associated with alterations of intracellular metabolism. While the mechanisms of innate immune memory carried out by immune cells are well characterized, such processes in non-immune cells, are poorly understood. The opportunistic pathogen, Staphylococcus aureus, is responsible for a multitude of human diseases, including pneumonia, endocarditis and osteomyelitis, as well as animal infections, including chronic cattle mastitis that are extremely difficult to treat. An induction of innate immune memory may be considered as a therapeutic alternative to fight S. aureus infection. Methods: In the current work, we demonstrated the development of innate immune memory in non-immune cells during S. aureus infection employing a combination of techniques including Enzyme-linked immunosorbent assay (ELISA), microscopic analysis, and cytometry. Results: We observed that training of human osteoblast-like MG-63 cells and lung epithelial A549 cells with ß-glucan increased IL-6 and IL-8 production upon a stimulation with S. aureus, concomitant with histones modifications. IL-6 and IL-8 production was positively correlated with an acetylation of histone 3 at lysine 27 (H3K27), thus suggesting epigenetic reprogramming in these cells. An addition of the ROS scavenger N-Acetylcysteine, NAC, prior to ß-glucan pretreatment followed by an exposure to S. aureus, resulted in decreased IL-6 and IL-8 production, thereby supporting the involvement of ROS in the induction of innate immune memory. Exposure of cells to Lactococcus lactis resulted in increased IL-6 and IL-8 production by MG-63 and A549 cells upon a stimulation with S. aureus that was correlated with H3K27 acetylation, suggesting the ability of this beneficial bacterium to induce innate immune memory. Discussion: This work improves our understanding of innate immune memory in non-immune cells in the context of S. aureus infection. In addition to known inducers, probiotics may represent good candidates for the induction of innate immune memory. Our findings may help the development of alternative therapeutic approaches for the prevention of S. aureus infection.

Protein ingredient quality of infant formulas impacts their structure and kinetics of proteolysis under in vitro dynamic digestion.

Infant formula (IF) is a complex matrix requiring numerous ingredients and processing steps. The objective was to understand how the quality of protein ingredients impacts IF structure and, in turn, their kinetics of digestion. Four powdered IFs (A/B/C/D), based on commercial whey protein (WP) ingredients, with different protein denaturation levels and composition (A/B/C), and on caseins with different supramolecular organisations (C/D), were produced at a semi-industrial level after homogenization and spray-drying. Once reconstituted in water (13 %, wt/wt), the IF microstructure was analysed with asymmetrical flow field-flow fractionation coupled with multi-angle light scattering and differential refractometer, transmission electron microscopy and electrophoresis. The rehydrated IFs were subjected to simulated infant in vitro dynamic digestion (DIDGI®). Digesta were regularly sampled to follow structural changes (confocal microscopy, laser-light scattering) and proteolysis (OPA, SDS-PAGE, LC-MS/MS, cation-exchange chromatography). Before digestion, different microstructures were observed among IFs. IF-A, characterized by more denatured WPs, presented star-shaped mixed aggregates, with protein aggregates bounded to casein micelles, themselves adsorbed at the fat droplet interface. Non-micellar caseins, brought by non-micellar casein powder (IF-D) underwent rearrangement and aggregation at the interface of flocculated fat droplets, leading to a largely different microstructure of IF emulsion, with large aggregates of lipids and proteins. During digestion, IF-A more digested (degree of proteolysis + 16 %) at 180 min of intestinal phase than IF-C/D. The modification of the supramolecular organisation of caseins implied different kinetics of peptide release derived from caseins during the gastric phase (more abundant at G80 for IF-D). Bioactive peptide release kinetics were also different during digestion with IF-C presenting a maximal abundance for a large proportion of them. Overall, the present study highlights the importance of the structure and composition of the protein ingredients (WPs and caseins) selected for IF formulation on the final IF structure and, in turn, on proteolysis. Whether it has some physiological consequences remains to be investigated.

Ileal Digestibility of Nitrogen and Amino Acids in Human Milk and an Infant Formula as Determined in Neonatal Minipiglets.

BACKGROUND: Infant formula (IF) has to provide at least the same amount of amino acids (AAs) as human milk (HM). AA digestibility in HM and IF was not studied extensively, with no data available for tryptophan digestibility. OBJECTIVES: The present study aimed to measure the true ileal digestibility (TID) of total nitrogen and AAs in HM and IF to estimate AA bioavailability using Yucatan mini-piglets as an infant model. METHODS: Twenty-four 19-day-old piglets (males and females) received either HM or IF for 6 days or a protein-free diet for 3 days, with cobalt-EDTA as an indigestible marker. Diets were fed hourly over 6 h before euthanasia and digesta collection. Total N, AA, and marker contents in diets and digesta were measured to determine the TID. Unidimensional statistical analyses were conducted. RESULTS: Dietary N content was not different between HM and IF, while true protein was lower in HM (-4 g/L) due to a 7-fold higher non-protein N content in HM. The TID of total N was lower (P < 0.001) for HM (91.3 ± 1.24%) than for IF (98.0 ± 0.810%), while the TID of amino acid nitrogen (AAN) was not different (average of 97.4 ± 0.655%, P = 0.272). HM and IF had similar (P > 0.05) TID for most of the AAs including tryptophan (96.7 ± 0.950%, P = 0.079), except for some AAs (lysine, phenylalanine, threonine, valine, alanine, proline, and serine), with small significant difference (P < 0.05). The first limiting AA was the aromatic AAs, and the digestible indispensable AA score (DIAAS) was higher for HM (DIAASHM = 101) than for IF (DIAASIF = 83). CONCLUSION: HM, compared to IF, had a lower TID for total N only, whereas the TID of AAN and most AAs, including Trp, was high and similar. A larger proportion of non-protein N is transferred to the microbiota with HM, which is of physiological relevance, although this fraction is poorly considered for IF manufacturing.

Peptidomic Characterization and Amino Acid Availability after Intake of Casein vs. a Casein Hydrolysate in a Pig Model.

It is known that casein hydrolysis accelerates gastrointestinal transit in comparison to intact casein, although the effect of the protein hydrolysis on the composition of the digests is not fully understood. The aim of this work is to characterize, at the peptidome level, duodenal digests from pigs, as a model of human digestion, fed with micellar casein and a previously described casein hydrolysate. In addition, in parallel experiments, plasma amino acid levels were quantified. A slower transit of nitrogen to the duodenum was found when the animals received micellar casein. Duodenal digests from casein contained a wider range of peptide sizes and a higher number of peptides above five amino acids long in comparison with the digests from the hydrolysate. The peptide profile was markedly different, and although ß-casomorphin-7 precursors were also found in hydrolysate samples, other opioid sequences were more abundant in the casein digests. Within the same substrate, the evolution of the peptide pattern at different time points showed minimal changes, suggesting that the protein degradation rate relies more on the gastrointestinal location than on digestion time. Higher plasma concentrations of methionine, valine, lysine and amino acid metabolites were found in animals fed with the hydrolysate at short times (<200 min). The duodenal peptide profiles were evaluated with discriminant analysis tools specific for peptidomics to identify sequence differences between both substrates that can be used for future human physiological and metabolic studies.

Encapsulation of DHA oil with heat-denatured whey protein in Pickering emulsion improves its bioaccessibility.

This study compared the bioaccessibility of docosahexaenoic acid (DHA) provided encapsulated or unencapsulated within a food matrix. DHA oil was composed of DHA-enriched triacylglycerols prepared as Pickering emulsion by encapsulation with heat-denatured whey protein isolate particles and then incorporated into homogenized liquid egg to get omelets. The effect of encapsulation was analyzed by using a static in vitro digestion model of the adult, which digestive fluid enzymes have also been characterized by proteomics. First, the size of lipid droplets was shown to be smaller and uniformly dispersed in omelets with encapsulated-DHA oil compared to non-encapsulated-DHA oil. Distribution of droplets was more regular with encapsulated-DHA oil as well. As a consequence, we showed that encapsulating DHA oil promoted the hydrolysis by pancreatic lipase during the intestinal phase. A larger proportion of DHA enriched-triacylglycerols was hydrolyzed after two hours of digestion, leading to a greater release in free DHA. Thus, only 32% of DHA remained esterified in the triacylglycerols with encapsulated-DHA oil, compared to 43% with non-encapsulated-DHA oil. The DHA in free form ultimately represented 52% of the total DHA with encapsulated-DHA oil, compared to 40% with non-encapsulated-DHA oil. Finally, our results showed that as much DHA was released after one hour of intestinal digestion when the DHA oil was encapsulated as after two hours when the DHA oil was not encapsulated. Therefore, DHA bioaccessibility was significantly improved by encapsulation of DHA oil in omelets.

Infant nutrition affects the microbiota-gut-brain axis: Comparison of human milk vs. infant formula feeding in the piglet model.

Early nutrition plays a dominant role in infant development and health. It is now understood that the infant diet impacts the gut microbiota and its relationship with gut function and brain development. However, its impact on the microbiota-gut-brain axis has not been studied in an integrative way. The objective here was to evaluate the effects of human milk (HM) or cow's milk based infant formula (IF) on the relationships between gut microbiota and the collective host intestinal-brain axis. Eighteen 10-day-old Yucatan mini-piglets were fed with HM or IF. Intestinal and fecal microbiota composition, intestinal phenotypic parameters, and the expression of genes involved in several gut and brain functions were determined. Unidimensional analyses were performed, followed by multifactorial analyses to evaluate the relationships among all the variables across the microbiota-gut-brain axis. Compared to IF, HM decreased the α-diversity of colonic and fecal microbiota and modified their composition. Piglets fed HM had a significantly higher ileal and colonic paracellular permeability assessed by ex vivo analysis, a lower expression of genes encoding tight junction proteins, and a higher expression of genes encoding pro-inflammatory and anti-inflammatory immune activity. In addition, the expression of genes involved in endocrine function, tryptophan metabolism and nutrient transport was modified mostly in the colon. These diet-induced intestinal modifications were associated with changes in the brain tissue expression of genes encoding the blood-brain barrier, endocrine function and short chain fatty acid receptors, mostly in hypothalamic and striatal areas. The integrative approach underlined specific groups of bacteria (Veillonellaceae, Enterobacteriaceae, Lachnospiraceae, Rikenellaceae, and Prevotellaceae) associated with changes in the gut-brain axis. There is a clear influence of the infant diet, even over a short dietary intervention period, on establishment of the microbiota-gut-brain axis.

Spatial-temporal mapping of the intra-gastric pepsin concentration and proteolysis in pigs fed egg white gels.

While there is a consensus that food structure affects food digestion, the underlying mechanisms remain poorly understood. A previous experiment in pigs fed egg white gels of same composition but different structures evidenced such effect on food gastric disintegration. In this study, we detailed the consequences on intra-gastric pH, pepsin concentration and proteolysis by sampling throughout the stomach over 6 h digestion. Subsequent amino acid absorption was investigated as well by blood sampling. While acidification was almost homogeneous after 6 h digestion regardless of the gel, pepsin distribution never became uniform. Pepsin started to accumulate in the pylorus/antrum region before concentrating in the body stomach beyond 4 h, time from which proteolysis really started. Interestingly, the more acidic and soft gel resulted in a soon (60 min) increase in proteolysis, an earlier and more intense peak of plasmatic amino acids, and a final pepsin concentration three times higher than with the other gels.

Encapsulation of Docosahexaenoic Acid Oil Substantially Improves the Oxylipin Profile of Rat Tissues.

Docosahexaenoic acid (DHA) is a major n-3 polyunsaturated fatty acid (PUFA) particularly involved in cognitive and cardiovascular functions. Due to the high unsaturation index, its dietary intake form has been considered to improve oxidation status and to favor bioaccessibility and bioavailability as well. This study aimed at investigating the effect of DHA encapsulated with natural whey protein. DHA was dietary provided as triacylglycerols to achieve 2.3% over total fatty acids. It was daily supplied to weanling rats for four weeks in omelet as food matrix, consecutively to a 6-hour fasting. First, when DHA oil was encapsulated, consumption of chow diet was enhanced leading to promote animal growth. Second, the brain exhibited a high accretion of 22.8% DHA, which was not improved by dietary supplementation of DHA. Encapsulation of DHA oil did not greatly affect the fatty acid proportions in tissues, but remarkably modified the profile of oxidized metabolites of fatty acids in plasma, heart, and even brain. Specific oxylipins derived from DHA were upgraded, such as Protectin Dx in heart and 14-HDoHE in brain, whereas those generated from n-6 PUFAs were mainly mitigated. This effect did not result from oxylipins measured in DHA oil since DHA and EPA derivatives were undetected after food processing. Collectively, these data suggested that dietary encapsulation of DHA oil triggered a more efficient absorption of DHA, the metabolism of which was enhanced more than its own accretion in our experimental conditions. Incorporating DHA oil in functional food may finally improve the global health status by generating precursors of protectins and maresins.

Gastrointestinal digestion enhances the endothelium-dependent vasodilation of a whey hydrolysate in rat aortic rings.

Whey proteins present encrypted biofunctional peptides that need to be released from the native protein to exert their biological activity. Antihypertensive whey peptides are the most studied ones, which can be explained by high prevalence of this chronic degenerative disease. The present study investigated whether the molecular changes occurred during the gastrointestinal digestion of a whey protein hydrolysate could modulate its vasorelaxant potential in rat aortic rings. Spectrophotometric data and SDS-PAGE gel showed a small degree of hydrolysis during the gastric phase and intense intestinal proteolysis. RP-HPLC revealed the formation of a large peptide profile. During the simulated digestion, 198 peptides were generated and identified and, left-shifted the concentration-response curve of the endothelium-dependent vasorelaxation, as recorded for the digested hydrolysates. In conclusion, gastrointestinal digestion of the whey hydrolysate leads to the generation of bioactive peptides with enhanced vasodilatory potency, reinforcing the relevance of whey-derived products in blood pressure regulation.

Whey-based cheese provides more postprandial plasma leucine than casein-based cheese: A pig study.

With a long-term nutrition goal for healthy aging, the aim of this study was to compare the bioavailability of amino acids, in particular the leucine, after the ingestion of two solid and isocaloric dairy products (cheese) based either on whey or on caseins, by using pig as an in vivo digestion model. The whey-based cheese contained 25% more leucine than Mozzarella, however its digestion by pigs resulted in a concentration of postprandial plasma leucine between 2 h and 5 h30 twice higher than that produced during the digestion of Mozzarella. Noting that the dry matter of the duodenal effluents were similar after each of the two cheese meals, differences in gastric emptying would not explain the difference in leucine bioavailability. These results suggest the possibility of stimulating more efficiently the muscle synthesis in elderly people with cheese based on whey proteins rather than those based on caseins.

Gastric Emptying and Dynamic In Vitro Digestion of Drinkable Yogurts: Effect of Viscosity and Composition.

Gastric emptying of food is mainly driven by the caloric concentration, the rheological properties of the chyme, and the physical state (liquid/solid) of food once in the stomach. The present work investigated: (1) The effect of the composition and the viscosity of drinkable yogurts on gastric emptying in pigs, and (2) the behavior of yogurts during dynamic in vitro digestion. Three isocaloric liquid yogurts were manufactured: Two enriched in protein and fiber showing either a low (LV) or high (HV) viscosity, one control enriched in sugar and starch (CT). They were labelled with 99mTc-sulfur colloid and given to pigs (n = 11) to determine gastric emptying pattern by gamma scintigraphy. Then dynamic in vitro digestion of the yogurts was done using the parameters of gastric emptying determined in vivo. Gastric emptying half-times were significantly longer for LV than CT, whereas HV exhibited an intermediate behavior. In vitro gastric digestion showed a quick hydrolysis of caseins, whereas whey proteins were more resistant in the stomach particularly for LV and HV. During the intestinal phase, both whey proteins and caseins were almost fully hydrolyzed. Viscosity was shown to affect the behavior of yogurt in the small intestine.

Impact of homogenization of pasteurized human milk on gastric digestion in the preterm infant: A randomized controlled trial.

BACKGROUND & AIMS: It has been suggested that homogenization of Holder-pasteurized human milk (PHM) could improve fat absorption and weight gain in preterm infants, but the impact on the PHM digestive kinetics has never been studied. Our objective was to determine the impact of PHM homogenization on gastric digestion in preterm infants. METHODS: In a randomized controlled trial, eight hospitalized tube-fed preterm infants were their own control to compare the gastric digestion of PHM and of homogenized PHM (PHHM). PHM was obtained from donors and, for half of it, was homogenized by ultrasonication. Over a six-day sequence, gastric aspirates were collected twice a day, before and 35, 60 or 90 min after the start of PHM or PHHM ingestion. The impact of homogenization on PHM digestive kinetics and disintegration was tested using a general linear mixed model. Results were expressed as means ± SD. RESULTS: Homogenization leaded to a six-fold increase in the specific surface (P < 0.01) of lipid droplets. The types of aggregates formed during digestion were different between PHM and PHHM, but the lipid fraction kept its initial structure all over the gastric digestion (native globules in PHM vs. blend of droplets in PHHM). Homogenization increased the gastric lipolysis level (P < 0.01), particularly at 35 and 60 min (22 and 24% higher for PHHM, respectively). Homogenization enhanced the proteolysis of serum albumin (P < 0.05) and reduced the meal emptying rate (P < 0.001, half-time estimated at 30 min for PHM and 38 min for PHHM). The postprandial gastric pH was not affected (4.7 ± 0.9 at 90 min). CONCLUSIONS: Homogenization of PHM increased the gastric lipolysis level. This could be a potential strategy to improve fat absorption, and thus growth and development in infants fed with PHM; however, its gastrointestinal tolerance needs to be investigated further. This trial was registered at clinicaltrials.gov as NCT02112331.

Impact of human milk pasteurization on gastric digestion in preterm infants: a randomized controlled trial.

BACKGROUND: Holder pasteurization has been reported to modify human milk composition and structure by inactivating bile salt-stimulated lipase (BSSL) and partially denaturing some of its proteins, potentially affecting its subsequent digestion. OBJECTIVE: We sought to determine the impact of human milk pasteurization on gastric digestion (particularly for proteins and lipids) in preterm infants who were fed their mothers' own milk either raw or pasteurized. DESIGN: In a randomized controlled trial, 12 hospitalized tube-fed preterm infants were their own control group in comparing the gastric digestion of raw human milk (RHM) with pasteurized human milk (PHM). Over a 6-d sequence, gastric aspirates were collected 2 times/d before and after RHM or PHM ingestion. The impact of milk pasteurization digestive kinetics and disintegration was tested with the use of a general linear mixed model. RESULTS: Despite inactivating BSSL, instantaneous lipolysis was not affected by pasteurization (mean ± SD at 90 min: 12.6% ± 4.7%; P > 0.05). Lipolysis occurred in milk before digestion and was higher for PHM than for RHM (mean ± SD: 3.2% ± 0.6% and 2.2% ± 0.8%, respectively; P < 0.001). Pasteurization enhanced the proteolysis of lactoferrin (P < 0.01) and reduced that of α-lactalbumin (only at 90 min) (P < 0.05). Strong emulsion destabilization was observed, with smaller aggregates and a higher specific surface for PHM (P < 0.05). Pasteurization did not affect gastric emptying (∼30-min half time) or pH (mean ± SD: 4.4 ± 0.8) at 90 min. CONCLUSIONS: Overall, pasteurization had no impact on the gastric digestion of lipids and some proteins from human milk but did affect lactoferrin and α-lactalbumin proteolysis and emulsion disintegration. Freeze-thawing and pasteurization increased the milk lipolysis before digestion but did not affect gastric lipolysis. Possible consequences on intestinal digestion and associated nutritional outcomes were not considered in this study. This trial was registered at clinicaltrials.gov as NCT02112331.

Acid and rennet gels exhibit strong differences in the kinetics of milk protein digestion and amino acid bioavailability.

This study aimed at determining the kinetics of milk protein digestion and amino acid absorption after ingestion by six multi-canulated mini-pigs of two gelled dairy matrices having the same composition, similar rheological and structural properties, but differing by their mode of coagulation (acidification/renneting). Duodenal, mid-jejunal effluents and plasma samples were collected at different times during 7h after meal ingestion. Ingestion of the acid gel induced a peak of caseins and ß-lactoglobulin in duodenal effluents after 20min of digestion and a peak of amino acids in the plasma after 60min. The rennet gel induced lower levels of both proteins in the duodenum (with no defined peak) as well as much lower levels of amino acids in the plasma than the acid gel. Plasma ghrelin concentrations suggested a potentially more satiating effect of the rennet gel compared to the acid gel. This study clearly evidences that the gelation process can significantly impact on the nutritive value of dairy products.

The heat treatment and the gelation are strong determinants of the kinetics of milk proteins digestion and of the peripheral availability of amino acids.

This study aimed to determine the kinetics of milk protein digestion and amino acid absorption after ingestion of four dairy matrices by six minipigs: unheated or heated skim milk and corresponding rennet gels. Digestive contents and plasma samples were collected over a 7 h-period after meal ingestion. Gelation of milk slowed down the outflow of the meal from the stomach and the subsequent absorption of amino acids, and decreased their bioavailability in peripheral blood. The gelled rennet matrices also led to low levels of milk proteins at the duodenum. Caseins and ß-lactoglobulin, respectively, were sensitive and resistant to hydrolysis in the stomach with the unheated matrices, but showed similar digestion with the heated matrices, with a heat-induced susceptibility to hydrolysis for ß-lactoglobulin. These results suggest a significant influence of the meal microstructure (resulting from heat treatment) and macrostructure (resulting from gelation process) on the different steps of milk proteins digestion.

Nisin quantification by ELISA allows the modeling of its apparent diffusion coefficient in model cheeses.

The diffusion of small solutes in cheese is of key importance for most enzymatic reactions involved in the ripening process. However, only a limited amount of data is available on salt diffusion and practically none on peptide diffusion. Nisin, a bacteriocin peptide, migrated in model cheeses made from ultrafiltered (UF) retentate. A profile concentration device and an enzyme-linked immunosorbent assay (ELISA), specifically developed for nisin quantification in cheese, were used to model the apparent diffusion coefficients for nisin according to Fick's law. This average coefficient was 49.5 µm(2)/s in UF cheese (n = 2). When 10% gelatin was added to the retentate, this value decreased to 34.4 µm(2)/s (n = 2). The two cheeses differed in their macrostructure (rheology) and microstructure (confocal microscopy). This study provides the first apparent diffusion coefficients for a peptide in cheese and supports the hypothesis that composition and structure influence the diffusion of small solutes such as peptides.

Localisation of caveolin in mammary tissue depends on cell type.

Caveolins, components of caveolae, are expressed in mammary tissue. In order to determine whether caveolins are present in different mammary cell types and whether their localisation depends on the physiological stage or species, cav-1 and cav-2 were characterised by immunoblotting in mammary tissues from the mouse, ewe and rabbit and localised, by immunofluorescence and electron microscopy, in mammary tissues from the mouse and ewe. At all the physiological stages studied, cav-1 and cav-2 were present in endothelial and myoepithelial cells in which flask-shaped caveolae were abundant. However, labelling of cav-1 and cav-2 associated with small vesiculo-tubular structures (including those close to lipid droplets) was low in epithelial cells. To study the possible association of cav-1 with lipid droplets, lactating ewe mammary fragments were treated in vitro with brefeldin A. This treatment did not modify the association of cav-1-labelled structures with lipid droplets. Finally, HC11 and MCF-10A mammary cell lines were treated with oleic acid. The total quantity of cav-1 was little affected by the treatment, although the lipid droplet labelling of cav-1 was amplified in MCF-10A cells. Thus, the synthesis and localisation of caveolins are mostly dependent upon the cell types of mammary tissue and upon their state of differentiation.