Hypoxia stress has been demonstrated to impede animal embryonic development, spermatogenesis, and lactation, leading to decreased animal production performance. However, the impact of hypoxia-induced activation of hypoxia inducible factor-1 (HIF-1) signaling on milk protein and fat synthesis remains unclear. L-leucine, a branched-chain amino acid, is known to modulate milk protein and fat synthesis. Therefore, our study aimed to evaluate the effect of L-leucine on milk protein and fat synthesis under hypoxic conditions and shed light on the molecular mechanism using an in vitro model. The results indicated that hypoxia treatment significantly decreased the synthesis of α-casein and β-casein, as well as inhibited factors related to milk fat synthesis in bovine mammary epithelial cells (MAC-T). Additionally, hypoxia stress suppressed the activities of the mammalian target of rapamycin (mTOR) and protein kinase B (AKT). Interfering with HIF-1α significantly reversed the expression of AKT, mTOR and factors related to milk synthesis. Importantly, supplementation with L-leucine activated AKT/mTOR signaling, thereby enhancing milk protein and fat synthesis in MAC-T cells to some extent. In conclusion, these findings suggest that HIF-1 signaling plays an important role in milk synthesis and that L-leucine may stimulate the synthesis of milk protein and fat by activating the AKT/mTOR signaling pathway under hypoxic conditions, making it a potential additive for promoting milk synthesis inhibited by hypoxia.

Intracellular amino acids (AA) regulate milk protein synthesis within the mammary glands by modifying mammary plasma flow (MPF) and AA transporter activity. Amino acid transporters catalyze translocation using Na+-gradient, substrate gradient (uniporters), and exchange mechanisms; further, they exhibit specificity for individual AA or groups of AA with similar side-chain properties within each transport system. Non-essential AA are actively transported through Na+-dependent transporters and, thus, are often utilized as intracellular currencies for EAA transport through exchange transporters. Therefore, it was hypothesized that individual EAA supplementation would compete with other EAA for shared transporters, and supplementation with Ala, Gln, and Gly would stimulate EAA transport through exchange transporters. Ten primiparous lactating dairy cows were divided into 2 groups based on milk production and were randomly assigned to treatment sequences within 2 balanced 5 × 5 Latin Squares by group. Period length was 14 d. Treatments were 9-d jugular infusions of 1) saline; 2) 34.5 g Val/d; 3) 32.7 g Ala/d: 40 g Gln/d: 26.7 g Gly/d (AQG); 4) 43 g Lys/d; or 5) 33.5 g Ile/d. All cows were fed a common base diet formulated to contain 15.0% CP. Ile, Lys, or AQG infusions did not affect milk protein or milk production; however, Val infusion decreased both. The effects of Val infusion on milk protein production appeared to be partially driven by decreased DMI. The decline in milk protein percentage indicated that milk lactose production was also affected. Additionally, Val infusion increased MPF efficiency (MPF/Milk; L/L) by approximately 44%. Val infusion tended to decrease or decreased mammary net uptakes of Lys, Leu, Met, and total AA. Ile infusion tended to increase its mammary net uptakes but did not affect any other AA. Lys and AQG infusions did not affect any mammary net uptakes. Val infusion tended to decrease Phe and total NEAA mammary clearance rates. AQG infusion stimulated Tyr clearance rates and tended to decline System N mammary clearance rates. Mammary uptake to milk protein output ratios (U:O) of BCAA did not differ from 1 for Val-infused cows, which indicated that little intramammary catabolism was occurring. Additionally, the average NEAA U:O in response to all treatments except Val was 0.70, but Val-infused cows had NEAA U:O that averaged 0.09 indicating increased synthesis within the glands. The effects of Val on mammary net clearance rates of multiple EAA support the incorporation of AA limitations in ration optimizers to prevent AA imbalances. It is possible that over-supplementation of EAA other than Val may also decrease DMI and mammary activity. Identifying efficiency apexes for each of the EAA will allow more precise diet formulation and supplementation, leading to improved production efficiency.

Smoking during lactation harmfully affects the amount and constituents of breast milk. Infants who consume breast milk containing miR-210-5p may have a higher risk of brain-related diseases. We investigated whether smoking during lactation decreases β-casein concentrations in milk and whether miR-210-5p expression is involved in smoking-induced β-casein suppression. During lactation, maternal CD1 mice were exposed to cigarette smoke (1.7 mg of tar and 14 mg of nicotine) in a smoke chamber for 1 h twice/day for five consecutive days. Control mice were placed in an air-filled chamber equivalent in size to the smoke chamber, with maternal separation times identical to those of the smoked mice. Maternal exposure to smoke during lactation significantly decreased β-casein expression in the mammary epithelia of smoked mice compared to that of the control mice. Signal transducer and activator transcription 5 (STAT5) and phosphorylated STAT5 (pSTAT5) are transcription factors involved in β-casein expression. In the mammary epithelia of smoked mice, the pSTAT5 and STAT5 levels were significantly lower, and miR-210-5p expression was significantly higher than that of the control mice. The β-casein, pSTAT5, and STAT5 protein levels of miR-210-5p mimic-transfected human mammary epithelial MCF-12A cells were significantly lower than those of control siRNA-transfected cells. These results indicate that smoke exposure led to an increase in miR-210-5p expression in mammary epithelium and a decrease in pSTAT5 and β-casein protein levels through the inhibition of STAT5 expression. Moreover, nicotine treatment decreased β-casein protein levels and increased miR-210-5p expression in non-malignant human mammary epithelial MCF-12A cells in a concentration-dependent manner, demonstrating that nicotine significantly affects the β-casein and miR-210-5p levels of breast milk. These results highlight the adverse effects of smoking on breast milk, providing essential information for healthcare professionals and general citizens.

Cancer is one of the leading causes of death worldwide, with over 10 million fatalities annually. While tumors can be surgically removed and treated with chemotherapy, radiotherapy, immunotherapy, hormonal therapy, or combined therapies, current treatments often result in toxic side effects in normal tissue. Therefore, researchers are actively seeking ways to selectively eliminate cancerous cells, minimizing the toxic side effects in normal tissue. Caseins and its derivatives have shown promising anti-cancer potential, demonstrating antitumor and cytotoxic effects on cells from various tumor types without causing harm to normal cells. Collectively, these data reveals advancements in the study of caseins and their derivative peptides, particularly providing a comprehensive understanding of the molecular mechanism of action in cancer therapy. These mechanisms occur through various signaling pathways, including (i) the increase of interferon-associated STAT1 signaling, (ii) the suppression of stemness-related markers such as CD44, (iii) the attenuation of the STAT3/HIF1-α signaling, (iv) the down-expression of uPAR and PAI-1, (v) the loss of mitochondrial membrane potential and reduced intracellular ATP production, (vi) the increase of caspase-3 activity, and (vii) the suppression of TLR4/NF-кB signaling. Therefore, we conclude that casein could be an effective adjuvant for cancer treatment.

In this present study, a three-factor Box-Behnken, response surface methodology (RSM) design was employed to optimize the skimmed milk powder (SMP)/whey protein concentrate (WPC) ratio (0.25-0.75%w/v) as a source of milk protein, inulin (1-2%w/v), and honey (4-6%w/v) for production of high-quality goat milk yoghurt (GMY). The resulting ANOVA and response surface equations revealed the significant effect (p < 0.05) of these variables on the various attributes such as total solid (%), pH, titratable acidity [(LA) % by weight], syneresis (%), DPPH (% inhibition), viscosity (m.Pa⋅s), whiteness index (WI), and overall acceptability (OA). The coefficient of determination (R2) for all response variables ranged from 0.88 to 0.99. Lack-of-fit tests resulted in non-significant F-values. The optimal conditions were determined as SMP/WPC at 0.36%w/v, inulin at 1.00%w/v, and honey at 6.00%w/v. The optimum values for total solid, pH, titratable acidity, syneresis, DPPH, viscosity, WI, and OA were 22.03, 4.46, 0.77, 6.34, 25.20, 182.30, 76.29 and 8.37, respectively with desirability value of 0.95.

This chapter provides an overarching view of the multifaceted aspects of milk β-casein, focusing on its genetic variants A1 and A2. The work examines the current landscape of A1-free milk versus regular milk, delving into health considerations, protein detection methods, technological impacts on dairy production, non-bovine protein, and potential avenues for future research. Firstly, it discussed ongoing debates surrounding categorizing milk based on A1 and A2 β-casein variants, highlighting challenges in establishing clear regulatory standards and quality control methods. The chapter also addressed the molecular distinction between A1 and A2 variants at position 67 of the amino acid chain. This trait affects protein conformation, casein micelle properties, and enzymatic susceptibility. Variations in β-casein across animal species are acknowledged, casting doubt on non-bovine claims of \"A2-like\" milk due to terminology and genetic differences. Lastly, this work explores the burgeoning field of biotechnology in milk production.

Milk bioactivity refers to the specific health effects of milk components beyond nutrition. The science of milk bioactivity involves the systematic study of these components and their health effects, as verified by empirical data, controlled experiments, and logical arguments. Conversely, \'faith in milk bioactivity\' can be defined as personal opinion, meaning, value, trust, and hope for health effects that are beyond investigation by natural, social, or human sciences. Faith can be strictly secular, but also influenced by spirituality or religion. The aim of this paper is to show that scientific knowledge is frequently supplemented with faith convictions to establish personal and public understanding of milk bioactivity. Mammalian milk is an immensely complex fluid containing myriad proteins, carbohydrates, lipids, and micronutrients with multiple functions across species, genetics, ages, environments, and cultures. Human health includes not only physical health, but also social, mental, and spiritual health, requiring widely different fields of science to prove the relevance, safety, and efficacy of milk interventions. These complex relationships between milk feeding and health outcomes prevent firm conclusions based on science and logic alone. Current beliefs in and understanding of the value of breast milk, colostrum, infant formula, or isolated milk proteins (e.g., immunoglobulins, α-lactalbumin, lactoferrin, and growth factors) show that both science and faith contribute to understand, stimulate, or restrict the use of milk bioactivity. The benefits of breastfeeding for infants are beyond doubt, but the strong beliefs in its health effects rely not only on science, and mechanisms are unclear. Likewise, fear of, or trust in, infant formula may rely on both science and faith. Knowledge from science safeguards individuals and society against \'milk bioactivity superstition\'. Conversely, wisdom from faith-based convictions may protect science from unrealistic \'milk bioactivity scientism\'. Honesty and transparency about the potentials and limitations of both scientific knowledge and faith convictions are important when informing individuals and society about the nutritious and bioactive qualities of milk.

BACKGROUND: Dairy consumption is associated with many health benefits. However, to our knowledge, no clinical trials examined the effects of milk protein concentrate (MPC) on metabolic health in overweight and obese adults. This study investigated the effect of supplementation with MPC on glycaemic status, lipid profile, biomarkers of inflammation, and anthropometric measurements in women with obesity under a weight loss diet.
METHODS: This is a single-blind, open-labelled, parallel-group, randomized trial. Forty-four healthy women with obesity were randomized into a control (n = 22) or MPC (n = 22) group. Participants in the MPC group were supplemented with 30 g of MPC per day for 8 weeks. Both groups were on a calorie-restricted diet plan with 800 Kcal lower intakes than their needs. Blood samples, dietary intake, and body composition were assessed before and after the intervention.
RESULTS: MPC group had a significantly lower body mass index (P = 0.009), waist circumference (P = 0.013), fat mass (P = 0.021), appetite score (P = 0.002), fasting blood sugar (P < 0.001), insulin (P = 0.027), low-density lipoprotein cholesterol (P = 0.025), and leptin (P = 0.014) levels and higher high-density lipoprotein cholesterol (P = 0.001) and adiponectin (P = 0.032) compared to the control group after supplementation. Lean body mass, total cholesterol, and triglyceride did not differ significantly (P > 0.05).
CONCLUSIONS: Daily intake of 30 g of MPC for 8 weeks may improve several anthropometric and metabolic markers in women with obesity under a hypocaloric diet.

Methionine dipeptide (Met-Met) could improve milk protein synthesis in bovine epithelia mammary cells and lactating mice, while the effects of Met-Met on lactation performance, rumen fermentation and microbiota profile in lactating dairy cows have not been explored. For this reason, 60 Chinese lactating Holstein cows were allocated into three treatment groups: control group (CON), 6 g/d methionine dipeptide group (MM), and 6.12 g/d rumen-protected methionine dipeptide group (RPMM). The experiment lasted for 10 weeks to monitor lactation performance, plasma amino acid profile and rumen fermentation parameters and microbiota profile. Results showed that MM increased the energy-corrected milk (ECM), and RPMM increased both milk yield and ECM (p < 0.05). The milk protein concentration and yield were increased by MM and RPMM (p < 0.05). The rumen fermentation showed that RPMM increased total volatile fatty acids, acetate and valerate concentrations (p < 0.05). The relative abundance of Firmicutes, including Succiniclasticum, Selenomonas and Clostridium_XlVa, were enriched and the Prevotella was decreased by RPMM (p < 0.05). In summary, daily supplementing with 6 g of MM or RPMM in lactating dairy cows could improve milk yield and both percentage and yield of milk protein, and RPMM benefited the rumen fermentation and altered the bacterial composition. These results provided the first evidence that Met-Met supplementation can improve lactation performance of dairy cows.

Previous research has demonstrated that in pregnant mice deficient in l-methionine (Met), the mixture of the dipeptide l-methionyl-l-methionine (Met-Met) with Met was more effective than Met alone in promoting mammogenesis and lactogenesis. This study aimed to investigate the role of a novel long noncoding RNA (lncRNA), named mammary gland proliferation-associated lncRNA (MGPNCR), in these processes. Transcriptomic analysis of mammary tissues from Met-deficient mice, supplemented either with a Met-Met/Met mixture or with Met alone, revealed significantly higher MGPNCR expression in the Met group compared to the mixture group, a finding recapitulated in a mammary epithelial cell model. Our findings suggested that MGPNCR hindered mammogenesis and milk protein synthesis by binding to eukaryotic initiation factor 4B (eIF4B). This interaction promoted the dephosphorylation of eIF4B at serine-422 by enhancing its association with protein phosphatase 2A (PP2A). Our study sheds light on the regulatory mechanisms of lncRNA-mediated dipeptide effects on mammary cell proliferation and milk protein synthesis. These insights underscore the potential benefits of utilizing dipeptides to improve milk protein in animals and potentially in humans.