Microwave Dielectric Response of Bovine Milk as Pregnancy Detection Tool in Dairy Cows.

The most reliable methods for pregnancy diagnosis in dairy herds include rectal palpation, ultrasound examination, and evaluation of plasma progesterone concentrations. However, these methods are expensive, labor-intensive, and invasive. Thus, there is a need to develop a practical, non-invasive, cost-effective method that can be implemented on the farm to detect pregnancy. This study suggests employing microwave dielectric spectroscopy (MDS, 0.5-40 GHz) as a method to evaluate reproduction events in dairy cows. The approach involves the integration of MDS data with information on milk solids to detect pregnancy and identify early embryonic loss in dairy cows. To test the ability to predict pregnancy according to these measurements, milk samples were collected from (i) pregnant and non-pregnant randomly selected cows, (ii) weekly from selected cows (n = 12) before insemination until a positive pregnancy test, and (iii) daily from selected cows (n = 10) prior to insemination until a positive pregnancy test. The results indicated that the dielectric strength of Δε and the relaxation time, τ, exhibited reduced variability in the case of a positive pregnancy diagnosis. Using principal component analysis (PCA), a clear distinction between pregnancy and nonpregnancy status was observed, with improved differentiation upon a higher sampling frequency. Additionally, a neural network machine learning technique was employed to develop a prediction algorithm with an accuracy of 73%. These findings demonstrate that MDS can be used to detect changes in milk upon pregnancy. The developed machine learning provides a broad classification that could be further enhanced with additional data.

The role of milk fat globule size in modulating the composition of postbiotics produced by Bacillus subtilis and their effect on mammary epithelial cells.

Milk lipids are secreted into the milk collecting ducts as milk fat globule (MFG) where they are exposed to microflora of the udder. We hypothesized that MFG size modulates the metabolic fingerprint of B. subtilis. Accordingly, small and large (2.3 and 7.0 µm, respectively) MFG were isolated from cow milk and used as a substrate for B. subtilis. Small MFG enhanced growth, whereas large MFG enhanced biofilm formation. Bacteria incubated with small MFG had increased concentration of metabolites related to energy production whereas metabolome of the bacteria incubated with large MFG had reduced concentrations of metabolites important for biofilm formation. Postbiotics from bacteria grown on large MFG exacerbated the proinflammatory response of MEC to LPS, and changed the expression of key enzymes involved in lipid and protein synthesis. Our results suggest that MFG size modulate growth trajectories and metabolome of B. subtilis, and consequently the stress response of host cells.

Effect of milk fat globules on growth and metabolism in rats fed an unbalanced diet.

We assessed the effects of supplementing milk fat globules (MFG) on the growth and development of the skeleton in rats fed a Western unbalanced diet (UBD). The UBD is high in sugar and fat, low in protein, fiber, and micronutrients, and negatively impacts health. The MFG-a complex lipid-protein assembly secreted into milk-has a unique structure and composition, which differs significantly from isolated and processed dietary ingredients. Rats consuming the UBD exhibited growth retardation and disrupted bone structural and mechanical parameters; these were improved by supplementation with small MFG. The addition of small MFG increased the efficiency of protein utilization for growth, and improved trabecular and cortical bone parameters. Furthermore, consumption of UBD led to a decreased concentration of saturated fatty acids and increased levels of polyunsaturated fatty acids (PUFA), particularly omega-6 PUFA, in the serum, liver, and adipose tissue. The addition of small MFG restored PUFA concentration and the ratio of omega-6 to omega-3 PUFA in bone marrow and adipose tissue. Finally, large but not small MFG supplementation affected the cecal microbiome in rats. Overall, our results suggest that natural structure MFG supplementation can improve metabolism and bone development in rats fed an UBD, with the effects depending on MFG size. Moreover, the benefits of small MFG to bone development and metabolism were not mediated by the microbiome, as the detrimental effects of an UBD on the microbiome were not mitigated by MFG supplementation.

The milk fat globule size governs a physiological switch for biofilm formation by Bacillus subtilis.

Milk lipids are organized in the form of milk fat globules (MFG), ranging in size from 0. 1 to 15 µm. The MFG size is closely associated with the composition of fatty acids, polar lipids, sphingolipids, cholesterol and the content of the MFG membrane (MFGM). Also, the MFGM integral proteins and glycoconjugates differ in composition and structure between different MFG size groups. These compositional differences may modulate the functionality of the MFG and its interaction with microbial cells. We report that small (2.3 µm) MFG facilitates the growth of the Gram-positive bacterium Bacillus subtilis whereas induction of biofilm formation was found in the presence of large (7.0 µm) MFG. Attempting to distinguish between the role played by the size from that played by the composition of the MFG, we compared phospholipid composition between treatments. We found that adjusting the phosphatidylethanolamine (PE) level to the concentration found in the small MFG, increased growth but suppressed biofilm formation in the presence of large MFG. The same normalization protocol for phosphatidylinositol (PI) or sphingomyeline (SM) did not exert a similar effect, suggesting a specific role for PE in regulating bacteria proliferation. We suggest that the content of MFGM, affected by MFG size, governs the ability of B. subtilis to utilize lipids from milk fat. This process might affect the bacterial decision-making toward biofilm formation or growth.

Lipidome changes, with a focus on phospholipids, due to feeding systems and processing in goat milk.

We evaluated the effects of processing - pasteurization and yoghurt manufacturing - on some health-promoting lipidome components in milk from two feeding treatments - brushland grazing or hay-feeding in confinement - in dairy goats. The contents of fat and protein were higher, and of urea, lower, in grazing goats. Fatty acid composition - at the exception of saturated fatty acids - was affected by dietary management and milk processing. Phospholipid contents was lower in confined goats, with little effect for processing. The phospholipid-to-triglyceride ratio was decreased by pasteurization. Sensitivity to pasteurization of phospholipid composition differed between feeding treatments. The percentage of sphingomyelin increased following pasteurization, with no response for fermentation to yoghurt. These results can be exploited to modulate health-promoting fats in dairy products.

Progesterone Regulation of Milk Fat Globule Size Is VLDL Dependent.

Progesterone plays a pivotal role during mammogenesis and serves as an inhibitor of the secretory activation of mammary cells in the last days of gestation. However, its role during lactogenesis, in particular its involvement in lipid metabolism, and milk fat content and composition, is unknown. Here, we provide new evidence of progesterone's involvement in the regulation of milk fat globule (MFG) synthesis and secretion. Findings from both in vivo and in vitro studies indicated that the concentration and the direction (increase vs. decrease) of progesterone concentration to which the mammary epithelial cells (MECs) are exposed affect MFG size. This was found to be very-low-density lipoprotein (VLDL) dependent: in the presence of VLDL, the proportion of MEC with small lipid droplets (<1 µm) increased 2.4-fold, and the proportion of large lipid droplets (>1 µm) increased 4-fold; in the absence of VLDL, no differences were found. The findings add to our understanding of the mechanism underlying the regulation of MFG size and provide new evidence for progesterone's role in lipid metabolism in the mammary gland during lactogenesis. The fact that the size, synthesis, and composition of MFG are affected by the cyclic pattern of progesterone concentration in the circulation might have physiologically relevant consequences, in particular on milk as a nutritional source.

Lipid Droplet Fusion in Mammary Epithelial Cells is Regulated by Phosphatidylethanolamine Metabolism.

Mammary epithelial cells (MEC) secrete fat in the form of milk fat globules (MFG) which are found in milk in diverse sizes. MFG originate from intracellular lipid droplets, and the mechanism underlying their size regulation is still elusive. Two main mechanisms have been suggested to control lipid droplet size. The first is a well-documented pathway, which involves regulation of cellular triglyceride content. The second is the fusion pathway, which is less-documented, especially in mammalian cells, and its importance in the regulation of droplet size is still unclear. Using biochemical and molecular inhibitors, we provide evidence that in MEC, lipid droplet size is determined by fusion, independent of cellular triglyceride content. The extent of fusion is determined by the cell membrane's phospholipid composition. In particular, increasing phosphatidylethanolamine (PE) content enhances fusion between lipid droplets and hence increases lipid droplet size. We further identified the underlying biochemical mechanism that controls this content as the mitochondrial enzyme phosphatidylserine decarboxylase; siRNA knockdown of this enzyme reduced the number of large lipid droplets threefold. Further, inhibition of phosphatidylserine transfer to the mitochondria, where its conversion to PE occurs, diminished the large lipid droplet phenotype in these cells. These results reveal, for the first time to our knowledge in mammalian cells and specifically in mammary epithelium, the missing biochemical link between the metabolism of cellular complex lipids and lipid-droplet fusion, which ultimately defines lipid droplet size.

Interplay between protein glycosylation pathways in Alzheimer's disease.

Deviations from the normal nucleoplasmic protein O-GlcNAcylation, as well as from normal protein sialylation and N-glycosylation in the secretory pathway, have been reported in Alzheimer's disease (AD). However, the interplay between the cytoplasmic protein O-GlcNAcylation and the secretory N-/O-glycosylation in AD has not been described. We present a comprehensive analysis of the N-, O-, and O-GlcNAc-glycomes in AD-affected brain regions as well as in AD patient serum. We detected marked differences in levels of glycan involved in both protein O-GlcNAcylation and N-/O-glycosylation between patients and healthy individuals and revealed brain region-specific glycosylation-related pathology in patients. These alterations are not general for other neurodegenerative conditions, such as frontotemporal dementia and corticobasal degeneration. The alterations in the AD glycome in the serum could potentially lead to novel glyco-based biomarkers for AD progression. Strikingly, negative interrelationship was found between the pathways of protein O-GlcNAcylation and N-/O-glycosylation, suggesting a novel intracellular cross-talk.