Bovine Milk Lactoferrin Research Articles

The effect of bovine milk lactoferrin-loaded exosomes (exoLF) on human MDA-MB-231 breast cancer cell line

BackgroundCancer is still the most challenging disease and is responsible for many deaths worldwide. Considerable research now focuses on targeted therapy in cancer using natural components to improve anti-tumor efficacy and reduce unfavorable effects. Lactoferrin is an iron-binding glycoprotein found in body fluids. Increasing evidence suggests that lactoferrin is a safe agent capable of inducing anti-cancer effects. Therefore, we conducted a study to evaluate the effects of the exosomal form of bovine milk lactoferrin on a human MDA-MB-231 breast cancer cell line.MethodsThe exosomes were isolated from cancer cells by ultracentrifugation and incorporated with bovine milk lactoferrin through the incubation method. The average size of the purified exosome was determined using SEM imaging and DLS analysis. The maximum percentage of lactoferrin-loaded exosomes (exoLF) was achieved by incubating 1 mg/ml of lactoferrin with 30 µg/ml of MDA-MB-231 cells-derived exosomes. Following treatment of MDA-MB-231 cancer cells and normal cells with 1 mg/ml exoLF MTT assay applied to evaluate the cytotoxicity, PI/ annexin V analysis was carried out to illustrate the apoptotic phenotype, and the real-time PCR was performed to assess the pro-apoptotic protein, Bid, and anti-apoptotic protein, Bcl-2.ResultsThe average size of the purified exosome was about 100 nm. The maximum lactoferrin loading efficiency of exoLF was 29.72%. MTT assay showed that although the 1 mg/ml exoLF treatment of MDA-MB-231 cancer cells induced 50% cell growth inhibition, normal mesenchymal stem cells remained viable. PI/ annexin V analysis revealed that 34% of cancer cells had late apoptotic phenotype after treatment. The real-time PCR showed an elevated expression of pro-apoptotic protein Bid and diminished anti-apoptotic protein Bcl-2 following exoLF treatment.ConclusionThese results suggested that exoLF could induce selective cytotoxicity against cancer cells compared to normal cells. Incorporating lactoferrin into the exosome seems an effective agent for cancer therapy. However, further studies are required to evaluate anti-tumor efficacy and the underlying mechanism of exoLF in various cancer cell lines and animal models.

Improved Neurodevelopmental Outcomes at 5.5 Years of Age in Children Who Received Bovine Milk Fat Globule Membrane and Lactoferrin in Infant Formula Through 12 Months: A Randomized Controlled Trial

To evaluate neurodevelopmental outcomes at 5.5 years of age in children who were previously randomized to cow milk-based infant formula (control) or similar formula (milk fat globule membrane plus lactoferrin [MFGM+LF]) with added sources of bovine MFGM and bovine LF (bLF) through 12 months of age. Children who completed study feeding were invited to participate in follow-up assessments: cognitive development across multiple domains (primary outcome; Wechsler Preschool & Primary Scale of Intelligence™, 4th Edition; WPPSI™-IV), inhibitory control/rule learning (Stroop Task), flexibility/rule learning (Dimensional Change Card Sort; DCCS), and behavior/emotion (Child Behavior Checklist; CBCL). Of 292 eligible participants (Control: 148, MFGM+LF: 144), 116 enrolled and completed assessments (Control: 59, MFGM+LF: 57). There were no group demographic differences except family income (MFGM+LF significantly higher). WPPSI-IV composite scores (mean±SE) for Visual Spatial (100.6±1.7 vs 95.3±1.7; P=0.027), Processing Speed (107.1±1.4 vs 100.0±1.4; P<0.001), and Full Scale IQ (98.7±1.4 vs 93.5±1.5; P=0.012) were significantly higher for MFGM+LF vs Control, even after controlling for demographic/socioeconomic factors. Stroop Task scores were significantly higher in MFGM+LF vs control (P<0.001). Higher DCCS scores (P=0.013) in the border phase (most complex/challenging) were detected and more children passed the border phase (32% vs 12%; P=0.039) for MFGM vs control. No group differences in CBCL score were detected. Children who received infant formula to 12 months of age with added bMFGM and bLF vs standard formula demonstrated improved cognitive outcomes in multiple domains at 5.5 years of age, including measures of intelligence and executive function.

The effect of casein and lactoferrin of bovine's milk on nestin expression of the exposed dental pulp

Background: Pulp tissue can regenerate if damage occurs. The pulp differentiates into new odontoblast-like cells in Progenitor cells. Nestin is a marker of the presence of new dontoblast-like and expressed in pathological conditions. Bovine's milk casein and lactoferrin contain proteins that accelerate cell growth. This study aimed to determine the effect of casein and lactoferrin on the expression of nested cells in the exposed dental pulp. Method: The research method is an experimental laboratory with male 30 Spraque Dawley rats. Rats were grouped as : casein, lactoferrin, casein-lactoferrin, and Ca-(OH)2. Each group consisted of 15 samples of molars which were divided into 5 samples of teeth for observation on the 7, 14, and 21 days. The first molars of rats were prepared until the pulp was exposed. The data of nestin quantity with immunohistochemical (IHC) analysis on histological preparations were analyzed using two-way ANOVA then the least significance difference (LSD) test. Result: The results of the average number of nested cells on 7,14 and 21 days were highest on combination of casein and lactoferrin. Two-way ANOVA statistical proved that there was a significant difference (p&lt;0.05) in each observation period and group and the number of expressions of nestin cells. LSD test showed that there was a significant difference (p&lt;0.05) between groups compared to the control group. Conclusion: Combination casein and lactoferrin extracts affect the expression of the number of nestin cells as a marker of new odontoblast-like cells in the exposed dental pulp.

Modulatory Efficiency of LP/LF Nano-Combination on Neurochemical and Behavioural Retardations in the Brain of Induced-Epileptic Rats.

&lt;b&gt;Background and Objective:&lt;/b&gt; Epilepsy is one of the normal neurological problems that came about because of strange electrical movements and prompt serious and far-reaching cell misfortune in the mind. This study aimed to investigate if a nano-Chitosan formulation loaded with bovine milk lactoperoxidase (LPO) and lactoferrin (LF) could prevent Lithium Chloride/Pilocarpine-induced epilepsy in rats or not. &lt;b&gt;Materials and Methods:&lt;/b&gt; Adult male rats (200-250 g) were partitioned into four groups (8 animals each) as follows: Group (1) Normal rats served as control group and received saline orally, group (2) Normal rats ingested with a daily oral dose of LPO and LF-NPS formulation at 50 mg kg&lt;sup&gt;&lt;/sup&gt;&lt;sup&gt;1&lt;/sup&gt;, group (3) Pilocarpine-induced epileptic rats and group (4) Epilepsy-modeled rats were treated with LPO+LF NPs (50 mg/kg/day, orally) for 6 weeks. &lt;b&gt;Results:&lt;/b&gt; The results revealed that the administration of LPO+LF-NPs markedly improved the induced-epilepsy disorders, this was monitored from the significant reduction in the values of caspase-3, TNF-α, IL-1β, CD4&lt;sup&gt;+&lt;/sup&gt;, MDA and NO coupled with remarkable raise in AchE-ase, dopamine, serotonin, SOD and GPx, CAT and GSH values in both brain regions. &lt;b&gt;Conclusion:&lt;/b&gt; This study supported the anti-epilepsy features of LPO+LF-NPS against Lithium Chloride/Pilocarpine-induced epilepsy in rats through the improvement of the immune response, reduction of inflammation and restoration of the impaired oxidative stress status.

Therapeutic efficacy of Nano-formulation of lactoperoxidase and lactoferrin via promoting immunomodulatory and apoptotic effects

This study identifies promising potential of a novel and safer nanocombination of bovine milk lactoperoxidase (LPO) and lactoferrin (LF) to target breast cancer in vitro and in adult female albino rat model. Favorable selective anticancer effects of the prepared nanocombination were observed, in a dose-dependent manner, against both MCF-7 and MDA cell lines, sparing normal HFB-4 cells. The administration of LPO + LFNPs markedly improved the induced-breast cancer disorders, prolonged survival and reduced the values of serum TNF-α, IL1β, CD4+, ALAT, ASAT, urea, creatinine, cholesterol and triglycerides with remarkable elevation in mammary SOD and GPx activity and GSH level. Moreover, the histopathological findings showed that LPO + LFNPs succeeded in prevention of mammary gland tumorigenesis. Superior efficacy of LPO + LFNPs was observed against pro-inflammatory cytokines through their anti-inflammatory and immunomodulatory properties. The treatment of LPO + LFNPs more significantly modulated the apoptosis and enhanced the expression of cell cycle regulator genes, which demonstrates a successful tumor therapy in vitro and in vivo. Therefore, this study provided evidence that the chemo-preventive feature of LPO + LFNPs may offer a novel alternative therapy for the treatment of breast cancer through enhances apoptosis pathway, improvement of immune response, reduction of inflammation and restoration of the impaired oxidative stress.

Protective effect of bovine lactoferrin against Cronobacter sakazakii in human intestinal Caco-2/TC7 cells

Milk is a source of bioactive proteins with defensive properties of great value for protecting the newborn. The activity of bovine milk lactoferrin (LF) was investigated as an antibacterial agent in the internalisation of the emergent pathogen Cronobacter sakazakii into Caco-2/TC7 cells, a model of human intestinal epithelium. The effect of LF on oxidative stress and expression of Toll-like receptors (TLR) was also investigated. LF exerted a clear antibacterial activity against C. sakazakii, as well as an inhibitory effect on C. sakazakii adhesion and internalisation into Caco-2/TC7 cells. Incubation with C. sakazakii induced an increase in oxidative stress on the lipid fraction of Caco-2/TC7 cells, which was reversed by LF. Additionally, LF altered the expression of TLR, with a clear decrease in the expression levels of TLR4 at 24 h of incubation. These results suggest that LF has very interesting properties as a potential ingredient for functional foods.

Analysis and comparison of key proteins in Maiwa yak and bovine milk using high-performance liquid chromatography mass spectrometry

Yak milk is an essential and predominant food resource for Tibetan people for subsistence purposes and to combat altitude-induced challenges. Due to its unique qualities, yak milk has recently been gaining broader attention from consumers across China as well in other parts of the world. One of the key characteristics of yak milk is the protein content, which is about 40 to 60% higher than that of native bovine milk. In this work, a sensitive and reproducible high-throughput analytical method was developed employing both ultra high-performance liquid chromatography Orbitrap (Thermo Fisher Scientific) high-resolution accurate mass spectroscopy (UHPLC-HRAM-MS) and UHPLC coupled with triple quadrupole tandem MS (UHPLC-QqQ-MS) to simultaneously analyze 8 milk proteins. A total of 15 Maiwa yak milk samples and 15 bovine milk samples were qualitatively and quantitatively analyzed using targeted proteomics and compared for α-lactalbumin, β-lactoglobulin, αS1-casein, αS2-casein, β-casein, κ-casein, lactoferrin, and osteopontin. Peptides of β-lactoglobulin were used to specifically distinguish yak and bovine milk. The results showed that this novel detection method could quantitatively detect these major and minor milk proteins with >0.99 linear correlation coefficient and a recovery rate between 90 and 120%, with relative standard deviations typically less than 10%. The data revealed that yak milk not only had higher overall milk protein content than bovine milk but higher lactoferrin and osteopontin contents as well. The lactoferrin content of yak milk was about 30% higher than that of bovine milk, and the osteopontin content of yak milk was nearly twice that of bovine milk. The application of this method demonstrates that UHPLC-HRAM-MS and UHPLC-QqQ-MS are suitable for high-throughput qualitative and quantitative analysis of major and minor proteins of yak and bovine milk.

Effect of Bovine Milk Fat Globule Membrane and Lactoferrin in Infant Formula on Gut Microbiome and Metabolome at 4 Months of Age

BackgroundMilk fat globule membrane (MFGM) and lactoferrin (LF) are human-milk bioactive components demonstrated to support gastrointestinal and immune development. Significantly fewer diarrhea and respiratory-associated adverse events through 18 mo of age were previously reported in healthy term infants fed a cow-milk–based infant formula with an added source of bovine MFGM and bovine LF through 12 mo of age.ObjectivesThe aim was to compare microbiota and metabolite profiles in a subset of study participants.MethodsStool samples were collected at baseline (10–14 d of age) and day 120. Bacterial community profiling was performed via 16S rRNA gene sequencing and alpha and beta diversity were analyzed (QIIME 2). Differentially abundant taxa were determined using linear discriminant analysis effect size (LefSE) and visualized (Metacoder). Untargeted stool metabolites were analyzed (HPLC/MS) and expressed as the fold-change between group means (control to MFGM+LF ratio).ResultsAlpha diversity increased significantly in both groups from baseline to 4 mo. Subtle group differences in beta diversity were demonstrated at 4 mo (Jaccard distance; R2 = 0.01, P = 0.042). Specifically, Bacteroides uniformis and Bacteroides plebeius were more abundant in the MFGM+LF group at 4 mo. Metabolite profile differences for MFGM+LF versus control included lower fecal medium-chain fatty acids, deoxycarnitine, and glycochenodeoxycholate, and some higher fecal carbohydrates and steroids (P < 0.05). After applying multiple test correction, the differences in stool metabolomics were not significant.ConclusionsAddition of bovine MFGM and LF in infant formula was associated with subtle differences in stool microbiome and metabolome by 4 mo of age, including increased prevalence of Bacteroides species. Stool metabolite profiles may be consistent with altered microbial metabolism. This trial was registered at https://clinicaltrials.gov as NCT02274883.

Application of instrumental methods for quality control of lactoferrin and raw materials for its production

Bovine cattle milk lactoferrin is industrially produced from cow’s milk and used in a variety of functional products, including baby food, nutritional supplements, dairy products, and veterinary drugs. The possibility of obtaining lactoferrin from lactic acid milk wheyrises a particular interest. The purpose of this study is to assess the possibility of using and adapting existing spectrophotometric methods for determining the concentration of total protein and an electrophoretic method for analyzing the protein composition of whey fractions and lactoferrin substances. A comparative study of two methods for determining the total protein in whey fractions and lactoferrin preparations biuret and Bradford method was carried out. The validation parameters of both methods were defined. An analytical range from 30 μg / ml to 1000 μg / ml characterizes the linearity of the Bradford method. The electrophoretic profile of milk, milk whey, whey fractions and substances of lactoferrinwere analyzed. The results obtained indicate a high information content of the electrophoretic method for characterizing the purification efficiency of whey fractions.

Oral Bovine Milk Lactoferrin Administration Suppressed Myopia Development through Matrix Metalloproteinase 2 in a Mouse Model.

Recent studies have reported an association between myopia development and local ocular inflammation. Lactoferrin (LF) is an iron-binding protein present in saliva, tears, and mother’s milk. Furthermore, sequestering iron by LF can cause its antibacterial property. Moreover, LF has an anti-inflammatory effect. We aimed to determine the suppressive effect of LF against the development and progress of myopia using a murine lens-induced myopia (LIM) model. We divided male C57BL/6J mice (3 weeks old) into two groups. While the experimental group was orally administered LF (1600 mg/kg/day, from 3-weeks-old to 7-weeks-old), a similar volume of Ringer’s solution was administered to the control group. We subjected the 4-week-old mice to −30 diopter lenses and no lenses on the right and left eyes, respectively. We measured the refraction and the axial length at baseline and 3 weeks after using a refractometer and a spectral domain optical coherence tomography (SD-OCT) system in both eyes. Furthermore, we determined the matrix metalloproteinase-2 (MMP-2) activity, and the amount of interleukin-6 (IL-6), MMP-2, and collagen 1A1 in the choroid or sclera. The eyes with a minus lens showed a refractive error shift and an axial length elongation in the control group, thus indicating the successful induction of myopia. However, there were no significant differences in the aforementioned parameters in the LF group. While LIM increased IL-6 expression and MMP-2 activity, it decreased collagen 1A1 content. However, orally administered LF reversed these effects. Thus, oral administration of LF suppressed lens-induced myopia development by modifying the extracellular matrix remodeling through the IL-6–MMP-2 axis in mice.

Detection of lactoferrin in bovine and goat milk by enzyme-linked immunosorbent assay

Lactoferrin (LF) has been proposed as a multifaceted functional ingredient of food. A competitive enzyme-linked immunosorbent assay (ELISA) was established to determine the LF in milk by using bovine LF antiserum and bovine LF-Biotin conjugate. The LF concentration in cow milk samples could be determined by this ELISA. However, the LF concentration in goat milk could not be measured using this method. Thus, the goat LF ELISA was established using goat LF-Biotin conjugate to measure the LF concentration in goat milk. Milk samples were collected for measurement of their LF concentrations. The mean LF level for the individual cow milk samples with the somatic cell count (SCC) below 1 × 105, 1 to 2.5 × 105 or 2.5 × 105 to 5 × 105 cells/mL was found to be about 176, 466 or 742 μg/mL, respectively. On the other hand, the LF level was about 200 μg/mL in goat bulk milks with the methylene blue reduction test (MBRT) time more than 5 hr. In conclusion, both LF ELISA methods developed in this study could be successfully used to determine the LF concentration in goat and cow milk.

Afffinity measurement of lactoferrin (LF)-anti-LF immunoglobulin in Yolk (IgY) complexes by competitive indirect enzyme-linked immunosorbent assay (CI-ELISA)

Competitive indirect enzyme-linked immunosorbent assay (CI-ELISA) was employed to perform the affinity measurement of dissociation constant (Kd) and affinity constants (Ka) for bovine milk lactoferrin (LF) and IgY (immunoglobulin in yolk) specific against LF using antisera from rabbit and hen as references. In liquid phase equilibrium measurements by CI-ELISA, the elevation in antigen level in antigen-antibody mixtures decreased the ELISA values, suggesting the increased competition of free LF in solution with that coated on plate for LF-specific IgY purified by immunoaffinity chromatography (purified IgY). From the Klotz plots of the binding of LF to purified IgY, Kd and Ka were determined to be about 2.6 × 10-8 M and 0.5 × 108 M-1, respectively. The Kd values of 1500-fold diluted crude IgY, diluted sera from hen and rabbit were determined to be very close to that of purified IgY, revealing that CI-ELISA under liquid phase equilibrium by CI-ELISA was appropriate for the affinity measurement of IgY samples.

Evaluation of Bioactivities of the Bovine Milk Lactoferrin-Osteopontin Complex in Infant Formulas.

Human milk contains several bioactive proteins, including lactoferrin (LF) and osteopontin (OPN). These two proteins have been shown to form a complex, which shows increased bioactivities. Bovine LF and OPN can also form such a complex. We assessed bioactivities of the bovine LF-OPN complex (at molar ratios of LF:OPN = 3:1, 5:1, or 8:1) in a formula protein matrix, including LF, OPN, bovine whey protein hydrolysate, and α-lactalbumin. Our results show that the bovine LF-OPN complex together with formula proteins is resistant to in vitro digestion, stimulates intestinal cell proliferation (by 15-50%) and differentiation (by 30-50%), increases antibacterial activity (by 25-50%), and enhances intestinal immunity. The 3:1 ratio of LF to OPN exhibits the most potent effects, as compared with the other two ratios. In conclusion, adding bovine LF and OPN to infant formulas may result in increased stability of the two components and enhanced bioactivities, possibly improving outcomes in formula-fed infants.

Binding analysis of bovine milk proteins, especially casein interactions and the interaction between α-casein and lactoferrin, using beads immobilised with zinc ion, poly-l-lysine or α-casein

Binding analyses of milk proteins (α-, β- and κ-casein, lactoferrin, α-lactalbumin, and β-lactoglobulin) were performed using beads immobilised with zinc ion (ZnB), poly-l-lysine (PLB) or α-casein (CasB). The amount of κ-casein bound to ZnB and CasB was decreased by the presence of α-casein, but the amount of α-casein increased in the absence of κ-casein. PLB strongly bound casein family proteins and β-lactoglobulin, but not lactoferrin or α-lactalbumin. CasB directly bound with apo-lactoferrin (lacking iron and haem). Lactoferrin in bovine milk was separated by binding with CasB after treating the milk with PLB. These results suggest that interactions between κ-casein and α-casein resulted in a lower affinity of κ-casein for zinc ion and α-casein, that κ-casein enhanced α-casein self-association, and that interactions between α-casein and lactoferrin are protein–protein interactions not due to haem-mediated binding. The combination of PLB and CasB is a simple method for obtaining lactoferrin from bovine milk.

Kinetic modelling of the heat stability of bovine lactoferrin in raw whole milk

The thermal denaturation of the naturally occurring lactoferrin in raw bovine whole milk has been measured over a range of temperatures from 65 to 121 °C for holding times from 2 up to 300 s designed to mimic heating processes used in the food manufacturing industry. Data were generated by heat treatments of raw whole milk from two different sources, New Zealand (NZ) and China, and with lactoferrin at two different levels of naturally occurring iron saturation. Denaturation rates for lactoferrin in the Chinese-sourced milk samples were at least two-fold greater than lactoferrin in the NZ milk over the temperature range 65–80 °C. At 85 °C rates of denaturation were the same although more than 80-fold greater than at 65 °C whereas, at 95 °C, lactoferrin in the NZ milk denatured at twice the rate of that in the Chinese milk. The differences in denaturation rates up to 80 °C likely reflected the greater heat stability of the more iron-saturated NZ milk samples. From these data, a combined kinetic and Arrhenius-based model has been developed and validated that allows calculation of native lactoferrin retained after thermal processing at any temperature-time combination in the experimental range (65–95 °C). This model should have applicability in the evaluation of existing processes and for the design of processes to improve the retention of lactoferrin during food manufacture.

Effect of hydrolysis and microwave treatment on the antibacterial activity of native bovine milk lactoferrin against Cronobacter sakazakii

Bovine lactoferrin (bLF) is an iron-binding glycoprotein used in functional and therapeutic products due to its biological properties, the most important being its antimicrobial activity. In this study, hydrolysates of bovine lactoferrin (bLFH) obtained with pepsin, chymosin and microbial rennet were assayed against Cronobacter sakazakii (104 CFU/mL) in different media: phosphate buffered saline (PBS), bovine skim milk and whey, and reconstituted powdered infant formula (PIFM). The results obtained have shown that hydrolysis of bLF enhances its antibacterial activity against C. sakazakii. The three types of bLFH dissolved in PBS reduced C. sakazakii growth from a concentration of 0.1 mg/mL and inhibited it completely above 0.5 mg/mL, after 4 and 8 h of incubation at 37 °C. The three bLFH (1 and 2 mg/mL) did not show any antibacterial activity in skim milk, whey and reconstituted PIFM after 8 h of incubation at 37 °C. However, C. sakazakii growth was completely inhibited in whey when pepsin and chymosin bLFH (2 mg/mL) were combined with undigested bLF (2 mg/mL), after 8 h of incubation at 37 °C. On the other hand, the combination of any of the three hydrolysates with bLF showed very low activity in skim milk and practically no activity in reconstituted PIFM. Furthermore, the effect of temperature after reconstitution (4, 23 and 37 °C), on the antibacterial activity of bLF (2.5 and 5 mg/mL) in reconstituted PIFM contaminated with C. sakazakii (10–102 CFU/mL) was also investigated. bLF at 5 mg/mL significantly reduced (p < .05) the proliferation of C. sakazakii in reconstituted PIFM at 37 °C until 2 h. C. sakazakii did not grow at 4 °C for 6 days in reconstituted PIFM with or without bLF. The effect of microwave heating (450, 550 and 650 W for 5, 10 and 15 s) on the antibacterial activity and stability of bLF (2.5 mg/mL) in reconstituted PIFM contaminated with C. sakazakii (10–102 CFU/mL) was also studied. The antibacterial activity of bLF was maintained after treatments at 450 and 550 W for 5 s, which kept 94 and 89% of bLF immunoreactivity, respectively. Moreover, microwave treatments of reconstituted PIFM with or without bLF, at 650 W for 5 s, and at 450, 550 and 650 W for 10 and 15 s, completely inactivated C. sakazakii.

The bovine Lactoferrin-Osteopontin complex increases proliferation of human intestinal epithelial cells by activating the PI3K/Akt signaling pathway

Lactoferrin (LF) and osteopontin (OPN), multifunctional proteins involved in cell proliferation, can form a complex. LF binds iron, whereas OPN binds calcium. We investigated whether iron- and calcium-binding influences complex formation and the pro-proliferation property of the LF-OPN complex, and the mechanism behind this effect. LF-OPN complexes were prepared using bovine milk LF and OPN, and effects on proliferation of human intestinal epithelial cells (HIECs) were evaluated using a BrdU proliferation assay. Of the four complexes formed by apo- and holo-LF/OPN, the apo-Lf&holo-OPN complex (AH) exhibited the strongest pro-proliferative effect on HIECs, and we therefore focused on AH. AH was resistant to in vitro gastrointestinal digestion, co-localized with both LF and OPN receptors as revealed by confocal microscopy, and stimulated proliferation of HIECs by activating PI3K/Akt signaling. In conclusion, forming a LF-OPN complex may help both proteins to resist digestion and increase the capacity to promote intestinal development in infants.

Improved Neurodevelopmental Outcomes Associated with Bovine Milk Fat Globule Membrane and Lactoferrin in Infant Formula: A Randomized, Controlled Trial

To evaluate neurodevelopment, growth, and health outcomes in infants receiving bovine milk fat globule membrane (MFGM) and lactoferrin in infant formula. Healthy term infants were randomized to a cow's milk-based infant formula or MFGM+LF (a similar infant formula, with an added source of bovine milk fat globule membrane [bMFGM; whey protein-lipid concentrate, 5g/L] and bovine lactoferrin [0.6g/L]) through 365days of age. The Bayley Scales of Infant Development, 3rd edition cognitive composite score at day 365 was the primary outcome. Secondary outcomes included tolerance measures through day 365, additional neurodevelopmental and language outcomes, growth, and medically confirmed adverse events through day545. Of 451 infants enrolled (control, 228; MFGM+LF, 223), 291 completed study feeding and Bayley-III testing at day 365 (control, 148; MFGM+LF, 143). The mean cognitive (+8.7), language (+12.3), and motor (+12.6) scores were higher (P< .001) for the MFGM+LF group; no differences were observed at day 545. Global development scores from day 120 to day 275 and attention at day 365 were significantly improved. Few group differences in day 545 neurodevelopmental outcomes were detected, however scores of some subcategories of the MacArthur-Bates Communicative Development Inventories were higher (P< .05) in the MFGM+LF group. The overall incidence of respiratory-associated adverse events and diarrhea were significantly lower for the MFGM+LF group through day545. Infants receiving formula with added bovine MFGM and bovine lactoferrin had an accelerated neurodevelopmental profile at day 365 and improved language subcategories at day 545. Formulas were associated with age-appropriate growth and significantly fewer diarrhea and respiratory-associated adverse events through 545days of age. TRIAL REGISTRATION CLINICALTRIALS.GOV:: NCT02274883.

Lactoferrin translocates to the nucleus of bovine rectal epithelial cells in the presence of Escherichia coli O157:H7

Enterohemorrhagic Escherichia coli (EHEC) O157:H7 is a foodborne pathogen which causes illness in humans. Ruminants are the main reservoirs and EHEC predominantly colonizes the epithelium of the recto-anal junction of cattle. Immunosuppression by EHEC promotes re-infection of cattle. However, bovine lactoferrin (bLF) apparently can overrule the immunosuppression by inducing EHEC-specific IgA responses at the mucosal site. The IgA responses are significantly correlated with reduced EHEC shedding and the absence of colonization at the rectal mucosa following re-infection. Therefore, to examine the interaction between bLF and bovine rectal epithelial cells, we first developed a method to establish a primary cell culture of epithelial cells of the rectum of cattle. Furthermore, we used LC–MS/MS to demonstrate the presence of secreted lactoferrin in bovine milk and the absence of a “delta” isoform which is known to translocate to the nucleus of cells. Nevertheless, lactoferrin derived from bovine milk was internalized by rectal epithelial cells and translocated to the nuclei. Moreover, nuclear translocation of bLF was significantly enhanced when the epithelial cells were inoculated with EHEC, as demonstrated by confocal fluorescence microscopy and confirmed by Raman microscopy and 3D imaging.

Bovine Milk Lactoferrin Selectively Kills Highly Metastatic Prostate Cancer PC-3 and Osteosarcoma MG-63 Cells In Vitro.

Prostate cancer and osteosarcoma are the second most common type of cancer affecting men and the fifth most common malignancy among adolescents, respectively. The use of non-toxic natural or natural-derived products has been one of the current strategies for cancer therapy, owing to the reduced risks of induced-chemoresistance development and the absence of secondary effects. In this perspective, lactoferrin (Lf), a natural protein derived from milk, emerges as a promising anticancer agent due to its well-recognized cytotoxicity and anti-metastatic activity. Here, we aimed to ascertain the potential activity of bovine Lf (bLf) against highly metastatic cancer cells. The bLf effect on prostate PC-3 and osteosarcoma MG-63 cell lines, both displaying plasmalemmal V-ATPase, was studied and compared with the breast cancer MDA-MB-231 and the non-tumorigenic BJ-5ta cell lines. Cell proliferation, cell death, intracellular pH, lysosomal acidification, and extracellular acidification rate were evaluated. Results show that bLf inhibits proliferation, induces apoptosis, intracellular acidification, and perturbs lysosomal acidification only in highly metastatic cancer cell lines. By contrast, BJ-5ta cells are insensitive to bLf. Overall, our results establish a common mechanism of action of bLf against highly metastatic cancer cells exhibiting plasmalemmal V-ATPase. This study opens promising perspectives for further research on the anticancer role of Lf, which ultimately will contribute to its safer and more rational application in the human therapy of these life-threatening cancers.