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Atherosclerosis (AS) is a common pathological basis for many cardiovascular diseases (CVDs) and result in high mortality and immense health and economic burdens worldwide. Early prevention, diagnosis, and treatment are promising approaches for stemming the development and progression of AS. Lactoferrin (Lf) is an iron-binding glycoprotein belonging to the transferrin family. It is widely found in body fluids such as digestive tract fluids, tears, and milk. Lf possesses anti-inflammatory, antibacterial, immunoregulatory, antioxidant and many other physiological functions. The serum Lf level is reportedly associated with the risk of AS and AS-related CVDs. Lf administration is closely involved in several mechanisms, including cholesterol metabolism, foam cell formation, ICAM-1 expression, homocysteine and leptin levels, anti-inflammatory and antioxidant function. Moreover, Lf has also been applied in the sythesis of magnetic resonance imaging (MRI) contrast agents to detect AS. Lf plays an important role in AS and may therefore be used in its diagnosis and treatment. Thus, this article aims to review the association between Lf and the risk of AS and AS-related CVDs, the mechanisms of Lf administration on AS, and its potential application in AS diagnosis. [ABSTRACT FROM AUTHOR]

While endotoxin (lipopolysaccharide) can be harmful and contribute to morbidity and mortality with Gram-negative sepsis or necrotizing enterocolitis in preterm infants, non-toxic amounts are produced as part of the neonatal microbiome and may be present in enteral nutrition and medications administered. The United States Food and Drug Administration has given guidance for endotoxin concentration limits for intravenous medications and fluids of 5 endotoxin units/kg/hour (120 endotoxin units/kg/day), but no guidance for amounts of endotoxin in enteral products. To determine baseline exposure to infants in the neonatal intensive care unit, we examined endotoxin content of enteral formulas and fortification used for preterm infants, as well as bovine lactoferrin products. We also examined endotoxin exposure and outcomes in very low birth weight infants. Endotoxin content was measured using kinetic chromogenic limulus amebocyte lysate analysis. Daily endotoxin exposure from enteral formulas ranged between < 75 to 7110 endotoxin units/kg and from lactoferrin products from 7 to 3720 endotoxin units/kg. In examining neonatal outcomes from a bovine lactoferrin product studied at three different escalating doses (100, 200, and 300 mg/kg/day), we measured endotoxin in the lactoferrin product and daily exposure was 1089 (N = 10), 2178 (N = 10) and 3287 (N = 11) endotoxin units/kg, respectively. There were no cases of necrotizing enterocolitis or mortality and no lactoferrin-related adverse effects in these patients. Enteral endotoxin daily exposures from lactoferrin products are similar to amounts in preterm enteral nutrition and appear safe and not associated with patient harm. Testing enteral products and establishing safety limits may improve care of high risk patients. [ABSTRACT FROM AUTHOR]

Lactoferrin (LF) has in vitro antiviral activity against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). This study aimed to determine the effect of bovine lactoferrin (bLF) in the prevention of SARS-CoV-2 infection in health care personnel. A randomized, double-blinded, placebo-controlled clinical trial was conducted in two tertiary hospitals that provide care to patients with SARS-CoV-2 infection in Lima, Peru. Daily supplementation with 600 mg of enteral bLF versus placebo for 90 days was compared. Participants were weekly screened for symptoms suggestive of SARS-CoV-2 infection and molecular testing was performed on suspected episodes. A serological test was obtained from all participants at the end of the intervention. The main outcome included symptomatic and asymptomatic cases. A sub-analysis explored the time to symptomatic infection. Secondary outcomes were the severity, frequency, and duration of symptomatic infection. The study was prematurely cancelled due to the availability of vaccines against SARS-CoV-2 in Peru. 209 participants were enrolled and randomized, 104 received bLF and 105 placebo. SARS-CoV-2 infection occurred in 11 (10.6%) participants assigned to bLF and in 9 (8.6%) participants assigned to placebo without significant differences (Incidence Rate Ratio = 1.23, 95%CI 0.51–3.06, p-value = 0.64). There was no significant effect of bLF on time to symptomatic infection (Hazard Ratio = 1.61, 95%CI 0.62–4.19, p-value = 0.3). There were no significant differences in secondary outcomes. A significant effect of bLF in preventing SARS-CoV-2 infection was not proven. Further studies are needed to assess the effect of bLF supplementation on SARS-CoV-2 infection. Clinical trial registration ClinicalTrials.gov Identifier: NCT04526821, https://clinicaltrials.gov/ct2/show/NCT04526821?term=LACTOFERRIN&cond=COVID-19&cntry=PE&city=Lima&draw=2&rank=1. [ABSTRACT FROM AUTHOR]

Prevention of preterm birth (PTB) is a global challenge and is one of the most important issues to be addressed in perinatal care. The hypothesis that ascending lower genital infection leads to PTB has been tested in numerous in vitro and in vivo studies. For patients with intractable vaginitis or high-risk patients with successive PTBs, mainly due to intra-uterine infection, the vaginal flora is enhanced to increase systemic immunity and locally propagate Lactobacillus species. It has been shown that the administration of lactoferrin (LF), a prebiotic with minimum side effects, may be effective in suppressing PTB. This hypothesis has been evaluated in this review using various relevant test examples. The findings suggest that LF may play a role in inflammatory protection in pregnant human cervical tissue. The antibacterial and anti-cytokine effects of LF in human-derived mucus-producing cervical cell lines were also demonstrated. It was also clarified that LF suppresses PTB and improves the prognosis of pups in inflammation-induced PTB animal models. Thus, we have identified that LF, a prebiotic contained in breast milk, can be clinically applied to suppress PTB in humans and to prevent PTBs in high-risk pregnancies. [ABSTRACT FROM AUTHOR]

Salivary stones (sialoliths) are calcified structures located in the ductal system of the major salivary glands. Their exact cause is not clear but in general they are characterized by concentric inorganic (hydroxyapatite) layers. The formation is a slow intermittent process which may result in enlargement of the sialolith causing obstruction of saliva secretion resulting in mealtime related pain and swelling of the affected salivary gland. Various studies reported the presence of organic material such as proteins and lipids in the core of sialoliths. In the present study the protein composition of twenty submandibular sialoliths was analyzed. It was found that proteins contributed on average 5% to the dry weight of submandibular stones whereby small salivary stones contained more extractable proteins than large salivary stones. Using a combination of SDS-PAGE gel electrophoresis and Western blotting, we identified α-amylase (in all stones; 100%), lysozyme (95%), lactoferrin (85%), secretory-IgA (75%), MUC7 (60%), complement C4 (60%) and C-reactive protein (35%). The presence, and the combinations, of lactoferrin, lysozyme, s-IgA and α-amylase in sialoliths was confirmed by ELISA. The gradually increasing size of a sialolith might provoke a local inflammatory response in the duct of the submandibular gland whereby the relatively low concentrations of lactoferrin and lysozyme may originate from neutrophils. The interaction of lactoferrin with s-IgA could contribute to the accumulation of lactoferrin in sialoliths. In summary, these results suggest a new pathophysiological role for lactoferrin, in the formation of sialoliths. [ABSTRACT FROM AUTHOR]

Iron is an essential element for various lifeforms but is largely insoluble due to the oxygenation of Earth's atmosphere and oceans during the Proterozoic era. Metazoans evolved iron transport glycoproteins, like transferrin (Tf) and lactoferrin (Lf), to keep iron in a non-toxic, usable form, while maintaining a low free iron concentration in the body that is unable to sustain bacterial growth. To survive on the mucosal surfaces of the human respiratory tract where it exclusively resides, the Gram-negative bacterial pathogen Moraxella catarrhalis utilizes surface receptors for acquiring iron directly from human Tf and Lf. The receptors are comprised of a surface lipoprotein to capture iron-loaded Tf or Lf and deliver it to a TonB-dependent transporter (TBDT) for removal of iron and transport across the outer membrane. The subsequent transport of iron into the cell is normally mediated by a periplasmic iron-binding protein and inner membrane transport complex, which has yet to be determined for Moraxella catarrhalis. We identified two potential periplasm to cytoplasm transport systems and performed structural and functional studies with the periplasmic binding proteins (FbpA and AfeA) to evaluate their role. Growth studies with strains deleted in the fbpA or afeA gene demonstrated that FbpA, but not AfeA, was required for growth on human Tf or Lf. The crystal structure of FbpA with bound iron in the open conformation was obtained, identifying three tyrosine ligands that were required for growth on Tf or Lf. Computational modeling of the YfeA homologue, AfeA, revealed conserved residues involved in metal binding. [ABSTRACT FROM AUTHOR]

Lactoferrin (LF) is present in the oviduct, reduces in vitro gamete interaction, and affects sperm capacitation parameters in humans. Our aim was to investigate LF actions on further stages of the reproductive process in the Wistar rat model. Motile sperm were obtained from cauda epididymis to assess LF binding by direct immunofluorescence and LF effect on acrosome reaction (AR) using a Coomassie blue staining. After ovarian hyperstimulation of female rats, oocytes were surgically recovered and coincubated with motile sperm and different doses of LF to estimate the in vitro fertilization (IVF) rate. To evaluate the LF effect on pregnancy and embryo implantation, female rats (80 days old) were placed with males and received daily intraperitoneal injections of LF during one complete estrous cycle (pregnancy experiments) or during the first 8 gestational days (implantation experiments). The number of pregnant females and live born pups was recorded after labor. Moreover, the number of implantation sites was registered during the implantation period. LF was able to bind to the sperm head, midpiece, and tail. 10 and 100 μg/ml LF stimulated the AR but reduced the IVF rate. The administration of 100 and 200 mg/kg LF significantly decreased the number of implantation sites and the litter size, whereas 100 mg/kg LF declined the pregnancy rate. The results suggest that LF might interfere with the reproductive process, possibly interfering with gamete interaction or inducing a premature AR; nevertheless, the mechanisms involved are yet to be elucidated. [ABSTRACT FROM AUTHOR]

The pathogenesis of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection involves dysregulations of iron metabolism, and although the mechanism of this pathology is not yet fully understood, correction of iron metabolism pathways seems a promising pharmacological target. The previously observed effect of inhibiting SARS-CoV-2 infection by ferristatin II, an inducer of transferrin receptor 1 (TfR1) degradation, prompted the study of competition between Spike protein and TfR1 ligands, especially lactoferrin (Lf) and transferrin (Tf). We hypothesized molecular mimicry of Spike protein as cross-reactivity of Spike-specific antibodies with Tf and Lf. Thus, strong positive correlations (R2 > 0.95) were found between the level of Spike-specific IgG antibodies present in serum samples of COVID-19-recovered and Sputnik V-vaccinated individuals and their Tf-binding activity assayed with peroxidase-labeled anti-Tf. In addition, we observed cross-reactivity of Lf-specific murine monoclonal antibody (mAb) towards the SARS-CoV-2 Spike protein. On the other hand, the interaction of mAbs produced to the receptor-binding domain (RBD) of the Spike protein with recombinant RBD protein was disrupted by Tf, Lf, soluble TfR1, anti-TfR1 aptamer, as well as by peptides RGD and GHAIYPRH. Furthermore, direct interaction of RBD protein with Lf, but not Tf, was observed, with affinity of binding estimated by KD to be 23 nM and 16 nM for apo-Lf and holo-Lf, respectively. Treatment of Vero E6 cells with apo-Lf and holo-Lf (1–4 mg/mL) significantly inhibited SARS-CoV-2 replication of both Wuhan and Delta lineages. Protective effects of Lf on different arms of SARS-CoV-2-induced pathogenesis and possible consequences of cross-reactivity of Spike-specific antibodies are discussed. [ABSTRACT FROM AUTHOR]

Milk is a source of proteins with high nutritional value and relevant biological activities. Bioactive milk proteins, like lactoferrin, are important for newborn development and can also be used as ingredients in functional products to improve health. Lactoferrin is essential in infant's diet, since protects against infections and promotes immune system maturation. Bovine lactoferrin is used to supplement formula milk in order to strengthen baby's defences against some pathogenic bacteria. Thus, lactoferrin supplemented formula can be a barrier against emergent pathogens, such as Cronobacter sakazakii, which has caused great concern in the last few years. Milk proteins generate bioactive peptides in the digestion process, and it is known that industrial processing can modify their susceptibility to digestion. Treatments such as heating have been shown to denature whey proteins and make them more easily digestible. Therefore, the aim of this study was to analyze the effect of technological treatments and gastrointestinal digestion on the antibacterial activity against C. sakazakii of proteins present in dairy formulas supplemented with lactoferrin. Commercial bovine lactoferrin has been shown to have antibacterial activity against C. sakazakii, both in the native state and after static in vitro gastrointestinal digestion. In addition, the digests obtained from dairy formulas subjected to technological treatments, either homogenization or pasteurization, have higher antibacterial activity than non-treated formulas. The release of low molecular weight peptides during the in vitro gastric digestion is probably the cause that would explain the enhanced antibacterial activity of the digested dairy formulas. [ABSTRACT FROM AUTHOR]

Iron is by far the most widespread and essential transition metal, possessing crucial biological functions for living systems. Despite chemical advantages, iron biology has forced organisms to face with some issues: ferric iron insolubility and ferrous-driven formation of toxic radicals. For these reasons, acquisition and transport of iron constitutes a formidable challenge for cells and organisms, which need to maintain adequate iron concentrations within a narrow range, allowing biological processes without triggering toxic effects. Higher organisms have evolved extracellular carrier proteins to acquire, transport and manage iron. In recent years, a renewed interest in iron biology has highlighted the role of iron-proteins dysregulation in the onset and/or exacerbation of different pathological conditions. However, to date, no resolutive therapy for iron disorders has been found. In this review, we outline the efficacy of Lactoferrin, a member of the transferrin family mainly secreted by exocrine glands and neutrophils, as a new emerging orchestrator of iron metabolism and homeostasis, able to counteract iron disorders associated to different pathologies, including iron deficiency and anemia of inflammation in blood, Parkinson and Alzheimer diseases in the brain and cystic fibrosis in the lung. [ABSTRACT FROM AUTHOR]

Lactoferrin (LTF) is a natural iron-binding protein with a potential for clinical utility in many human immune disorders, including cancer. A fusion of LTF with the Fc domain of IgG2 (FcLTF) was designed with inherent properties of an extended the half-life in circulation. Furthermore, the effects of LTF and FcLTF were assessed for influence on the activity of natural killer (NK) cells isolated from human peripheral blood, on the NK-92 cell line, and on human monocytes. The NK cytotoxic activity induced by LTF and FcLTF was determined against the human leukemia K562 cell line, and also for monocytes, by measuring TNFα and granzyme B production, and in an assay for Jurkat cell viability. Selected gene expression in NK-92 cells and monocytes, induced by LTF and FcLTF, was performed by Real Time PCR. No significant difference was observed in NK-92 cytotoxicity stimulated by LTF and FcLTF. The effects on NK cells isolated from the human peripheral blood were varied, possibly due to the immunoregulatory nature of LTF sensing the immune status of donors. Furthermore, only the FcLTF group strongly stimulated production of TNFα and granzyme B in isolated monocytes. In addition, only supernatants from the monocyte cultures treated with FcLTF decreased the viability of Jurkat cells. The ability of FcLTF to induce TNFα in monocytes was strongly inhibited by anti-CD32 and moderately inhibited by anti-CD14 antibody. Lastly, it was demonstrated that FcLTF, strongly induced expression of PI3K, with subsequent activation of AKT/mTOR signaling pathway. Overall, it was demonstrated that this novel fusion molecule may be a perferred choice for clinical utility than the wild type LTF. [ABSTRACT FROM AUTHOR]

Bone and joint infections are a rare but serious problem worldwide. Lactoferrin's antimicrobial and antibiofilm activity coupled with its bone-regenerating effects may make it suitable for improving bone and joint infection treatment. However, free lactoferrin (LF) has highly variable oral bioavailability in humans due to potential for degradation in the stomach and small intestine. It also has a short half-life in blood plasma. Therefore, encapsulating LF in nanocarriers may slow degradation in the gastrointestinal tract and enhance LF absorption, stability, permeability and oral bioavailability. This review will summarize the literature on the encapsulation of LF into liposomes, solid lipid nanoparticles, nanostructured lipid carriers, polymeric micro and nanoparticles and hydroxyapatite nanocrystals. The fabrication, characterization, advantages, disadvantages and applications of each system will be discussed and compared. [ABSTRACT FROM AUTHOR]

Lactoferrin (LTF), an iron binding protein, is known to exhibit immune modulatory effects on pulmonary pathology during insult-induced models of primary Mycobacterium tuberculosis (Mtb) infection. The effects of LTF correlate with modulation of the immune related development of the pathology, and altering of the histological nature of the physically compact and dense lung granuloma in mice. Specifically, a recombinant human version of LTF limits immediate progression of granulomatous severity following administration of the Mtb cell wall mycolic acid, trehalose 6,6′-dimycolate (TDM), in part through reduced pro-inflammatory responses known to control these events. This current study investigates a limited course of LTF to modulate not only initiation, but also maintenance and resolution of pathology post development of the granulomatous response in mice. Comparison is made to a fusion of LTF with the Fc domain of IgG2 (FcLTF), which is known to extend LTF half-life in circulation. TDM induced granulomas were examined at extended times post insult (day 7 and 14). Both LTF and the novel FcLTF exerted sustained effects on lung granuloma pathology. Reduction of pulmonary pro-inflammatory cytokines TNF-α and IL-1β occurred, correlating with reduced pathology. Increase in IL-6, known to regulate granuloma maintenance, was also seen with the LTFs. The FcLTF demonstrated greater impact than the recombinant LTF, and was superior in limiting damage to pulmonary tissues while limiting residual inflammatory cytokine production. [ABSTRACT FROM AUTHOR]

Plasmacytoid dendritic cells (pDCs) recognise viral single-stranded RNA (ssRNA) or CpG DNA via Toll-like receptor (TLR)-7 and TLR9, and produce interferon (IFN)-α. Activated pDCs upregulate human leukocyte antigen (HLA)-DR and CD86 expression levels. Ingestion of bovine lactoferrin (LF) activates pDCs, but little is known about its effects. In this study, the effects of LF and its pepsin hydrolysate (LFH) on the production of IFN-α from peripheral blood mononuclear cells (PBMCs) and pDCs were examined. PBMCs were prepared from peripheral blood of healthy adults and incubated with LF, LFH, or lactoferricin (LFcin) in the absence or presence of ssRNA derived from human immunodeficiency virus. The concentration of IFN-α in the supernatant and the expression levels of IFN-α, HLA-DR, and CD86 in pDCs were quantified by enzyme-linked immunosorbent assay and flow cytometry. In the absence of ssRNA, the concentration of IFN-α was negligible and LF had no effect on it. In the presence of ssRNA, IFN-α was detected at a certain level, and LF and LFH significantly increased its concentration. The increase caused by LFH and LFcin were comparable. In addition, LF significantly upregulated the expression levels of IFN-α, HLA-DR, and CD86 in pDCs. LF and its digestive peptides induced IFN-α production and activated pDCs in the presence of ssRNA, suggesting that LF modulates the immune system by promoting pDC activation upon viral recognition. [ABSTRACT FROM AUTHOR]

Lactoferrin (LF) is abundant in human milk and plays an important role in the health of children. Bovine LF (bLF) has high homology with human LF and has been reported to have multiple biological functions. Several clinical studies have been conducted considering these properties, which reported the usefulness of bLF. This review was aimed to provide an overview of the clinical evidence in children. We searched clinical reports investigating the effects of bLF in children and identified 36 studies on the role of bLF in infections, iron metabolism, body growth, cerebral development, and fecal microbiome. Considering the accumulated evidence, bLF may contribute to the child health, particularly by suppressing or alleviating gastrointestinal and respiratory symptoms, and improving the iron status of children with anemia or those at high risk of anemia. The dose of bLF varies depending on the expected effect and target age, but may not necessarily have to be as high as human LF in human milk. Some of the beneficial effects of bLF have not been fully validated due to limited clinical evidence or being observed in the secondary analysis of some studies. Further clinical evidence would add significant value to the use of bLF in child health. [ABSTRACT FROM AUTHOR]

Beyond the absolute and indisputable relevance and efficacy of anti-SARS-CoV-2 vaccines, the rapid transmission, the severity of infection, the absence of the protection on immunocompromised patients, the propagation of variants, the onset of infection and/or disease in vaccinated subjects and the lack of availability of worldwide vaccination require additional antiviral treatments. Since 1987, lactoferrin (Lf) is well-known to possess an antiviral activity related to its physico-chemical properties and to its ability to bind to both heparan sulfate proteoglycans (HSPGs) of host cells and/or surface components of viral particles. In the present review, we summarize in vitro and in vivo studies concerning the efficacy of Lf against DNA, RNA, enveloped and non-enveloped viruses. Recent studies have revealed that the in vitro antiviral activity of Lf is also extendable to SARS-CoV-2. In vivo, Lf oral administration in early stage of SARS-CoV-2 infection counteracts COVID-19 pathogenesis. In particular, the effect of Lf on SARS-CoV-2 entry, inflammatory homeostasis, iron dysregulation, iron-proteins synthesis, reactive oxygen formation, oxidative stress, gut-lung axis regulation as well as on RNA negativization, and coagulation/fibrinolysis balance will be critically reviewed. Moreover, the molecular mechanisms underneath, including the Lf binding to HSPGs and spike glycoprotein, will be disclosed and discussed. Taken together, present data not only support the application of the oral administration of Lf alone in asymptomatic COVID-19 patients or as adjuvant of standard of care practice in symptomatic ones but also constitute the basis for enriching the limited literature on Lf effectiveness for COVID-19 treatment. [ABSTRACT FROM AUTHOR]

Lactoferrin (Lf), a multiple functional natural immune protein, is widely distributed in mammalian milk and glandular secretions (bile, saliva, tears and nasal mucosal secretions, etc.). In the previous study, we found that Lf plays an anti-inflammatory and anti-tumorigenesis role in AOM/DSS (azoxymethane/dextran sulfate sodium) induced mouse colitis-associated colon cancer model. Although we found that Lf has anti-inflammatory effects in chronic inflammation, its specific role and mechanisms in acute inflammation have not been clarified. Here, we reported that the expression levels of Lf were significantly increased when the organism was infected by Gram-negative bacteria. We then explored the role and potential mechanism of Lf in lipopolysaccharide (LPS)-induced acute inflammation. In the LPS-induced acute abdominal inflammation model, Lf deficiency aggravated inflammatory response and promoted macrophage chemotaxis to the inflammation site. Lf inhibited macrophage chemotaxis by suppressing the expression of macrophage-associated chemokines Ccl2 and Ccl5. Highly activated NF-κB signaling in Lf−/− mice was responsible for the high expression of Ccl2 and Ccl5. Our results suggested that the anti-inflammatory effect of Lf offers a new potential treatment for acute inflammatory diseases. [ABSTRACT FROM AUTHOR]

Zn2+ ions are essential in many physiological processes, including enzyme catalysis, protein structural stabilization, and the regulation of many proteins. The affinities of proteins for Zn2+ ions span several orders of magnitude, with catalytic Zn2+ ions generally held more tightly than structural or regulatory ones. Metal carrier proteins, most of which are not specific for Zn2+, bind these ions with a broad range of affinities that overlap those of catalytic, structural, and regulatory Zn2+ ions and are thought to be responsible for distributing the metal through most cells, tissues, and fluid compartments. While little is known about how many proteins obtain or release these ions, there is now considerable experimental evidence suggesting that metal carrier proteins may be responsible for transferring metals to and from some Zn2+-dependent proteins, thus serving as a major regulatory factor for them. In this review, the biological roles of Zn2+ and structures of Zn2+ binding sites are examined, and experimental evidence demonstrating the direct participation of metal carrier proteins in enzyme regulation is discussed. Mechanisms of metal ion transfer are also offered, and the potential physiological significance of this phenomenon is explored. [ABSTRACT FROM AUTHOR]

We aimed to evaluate the expression of growth differentiation factor-15 (GDF-15) and lactoferrin (Lf) in tumor and their relationship with the body iron-status and overall survival (OS) outcome of patients with breast cancer. A retrospective cohort study of female patients with primary breast cancer was performed. Clinical tumor samples from the Second Affiliated Hospital of Soochow University between December 2008 and June 2014 were collected. The immuno-expression of GDF-15 and Lf was stratified into positive or negative expression. Kaplan–Meier method and Cox proportional hazards regression model were used for data analysis. 74 breast cancer patients with a mean age of 52 years were included into our study. 14 (18.9%) patients were died by the end of August 1, 2019. The serum iron level of patients with GDF-15 (+)/Lf(−) expression was higher than that of patients with other expression patterns (18.2 ± 5.4 vs. 15.5 ± 5.0 μmol/L, P = 0.038), but was not associated with OS. In univariate Cox analyses, GDF-15(+) and GDF-15(+)/Lf(−) were significantly correlated with high mortality risk (HR = 3.75, 95%CI 1.05–13.48, P = 0.025; HR = 5.00, 95%CI 1.56–16.04, P = 0.004, respectively). After adjusted for age, menopause status and primary tumor grade, the association between GDF-15 and OS disappeared. However, the association between GDF-15/Lf and OS still existed in GDF-15(+)/Lf(−) (HR = 4.50, 95%CI 1.31–15.51, P = 0.017). The combined immuno-expression pattern of GDF-15 and Lf was significant associated with high serum iron level. GDF-15/Lf could be a powerful biomarker to predict survival outcome of patients with breast cancer but still needed to be confirmed by future studies. [ABSTRACT FROM AUTHOR]

Bovine milk proteins bind calcium and some bind other metal ions or heme. The examination of heme-binding proteins in colostrum and milk using hemin-agarose beads (HA) showed α-casein, κ-casein and lactoferrin (Lf) to be heme-binding proteins. α-Casein and Lf are iron- and heme-binding proteins, and α- and κ-casein bind to HA, as does Lf. κ-Casein and Lf have higher affinity to zinc ion than does α-casein, and κ-casein and Lf interact with α-casein-immobilized beads (CasB). The addition of α-casein to κ-casein bound to CasB decreased the amount of bound κ-casein compared with in the absence of α-casein, and κ-casein likely increases α-casein self-association. α-Casein binds Lf bound to neither iron nor heme, as shown by experiments with the apo-form. Beads with immobilized poly-l-lysine bind heme but Lf inhibits this binding. These results indicate that α-casein, κ-casein and Lf are both heme- and zinc-binding proteins, and that α-casein interacts with κ-casein and Lf through protein-protein interactions. Additionally, Lf shows higher affinity to hemin than does poly-l-lysine. [ABSTRACT FROM AUTHOR]

Milk derivative bovine Lactoferrin (bLf), a multifunctional glycoprotein available in large quantities and recognized as safe, possesses high homology and identical functions with human Lactoferrin. There are numerous food supplements containing bLf which, however, can vary in its purity, integrity and, consequently, functionality. Here, we report on a clinical trial where bLf (100 mg two times/day) was orally administered before (Arm A) or during meals (Arm B) to pregnant women with hereditary thrombophilia and suffering from anemia of inflammation. A significant increase of the number of red blood cells (RBCs), hemoglobin (Hb), total serum iron (TSI) and serum ferritin (sFtn) levels, along with a significant decrease of interleukin-6 were detected after 30 days in Arm A, but not in Arm B, thus letting us to hypothesize that bLf inefficacy could be related to its degradation by digestive proteases. To verify this hypothesis, bLf was incubated in gastric juice collected before or after meals. An undigested or a digested profile was observed when bLf was incubated in gastric juice sampled before or after meals, respectively. These results can explain the beneficial effect observed when bLf is administered under fasting conditions, i.e. in the absence of active proteases. [ABSTRACT FROM AUTHOR]

Lactoferrin (Lf) has been involved in diverse type of cellular activities and its biochemical properties are species specific. Lf is a bilobal molecule in which each lobe binds with one Fe2+/Fe3+ ion. A lot of physiological effects of Lf are regulated by its iron binding and release properties; however these properties are species-specific. To understand the iron-binding, thermal stability and cytotoxic effect of buffalo Lf (buLf) and contribution of individual N- and C-terminal lobes therein, buLf and the truncated monoferric lobes were expressed in Kluyveromyces lactis or Pichia pastoris yeast expression systems. The iron-uptake/release behavior and thermal stability of recombinant buLf was observed similar to the Lf purified from buffalo milk. Supplementation of recombinant buLf to the buffalo mammary epithelial cells (BuMEC) culture decreased their proliferation and the cell viability in a dose dependent manner. The cell growth decreased by 37% at 1.0 mg/ml Lf. C-lobe decreased the viability of BuMEC by 15% at 1 mg/ml. The C-lobe showed greater cytotoxic effect against BuMEC in comparison to N-lobe. buLf caused a reduced expression of the casein in BuMEC. At 1.0 mg/ml of buLf, CSN2 transcript level was reduced by 74% and 78% in the normal and hormone free media, respectively. The expression of IL-1β gene in BuMEC increased by 4–5 fold in the presence of 1.0 mg/ml of Lf. The effect was similar to that observed in the involutory mammary gland, suggesting the role of elevated level of Lf in remodeling of buffalo mammary tissue during involution. [ABSTRACT FROM AUTHOR]

Probiotics and lactoferrin are currently being used in neonatal intensive care units in the hopes of reducing rates of sepsis and necrotizing enterocolitis (NEC). While studies have shown that these measures can be clinically beneficial to premature babies, and there are ongoing trials to measure their impact on NEC and sepsis rates, little is known about how they may impact microbiota development. We thus employed a newborn piglet model to assess the impact of feeding probiotics or a combination of probiotics and lactoferrin on development of the gastrointestinal microbiota. Healthy full-term piglets were fed either probiotics alone or probiotics and a bovine lactoferrin supplement over the first weeks of life, and their microbiota profiles were compared with unsupplemented controls. We found that both probiotic and probiotic plus lactoferrin treatments impacted the microbial composition within the gastrointestinal tract, with differing impacts on various regions within the gut. In addition, the impact of probiotics was often reversed by the presence of lactoferrin and both feeding interventions altered the microbiota's genetic propensity to use ferric versus ferrous ions. These results suggest that iron availability may be a key factor to consider when designing feeding interventions that target the microbiome. [ABSTRACT FROM AUTHOR]

Yersinia pestis is the causative agent of plague. As adequate antibiotic treatment falls short and currently no effective vaccine is available, alternative therapeutic strategies are needed. In order to contribute to solving this problem we investigated the therapeutic potential of the peptide construct LFchimera against the safer-to-handle Y. pestis simulants Yersinia enterocolitica and Yersinia pseudotuberculosis in vitro. LFchimera is a heterodimeric peptide construct mimicking two antimicrobial domains of bovine lactoferrin, i.e. lactoferrampin and lactoferricin. LFchimera has been shown to be a potent antimicrobial peptide against a variety of bacteria in vitro and in vivo. Also Y. enterocolitica and Y. pseudotuberculosis have been shown to be susceptible for LFchimera in vitro. As Yersiniae spp. adhere to and invade host cells upon infection, we here investigated the effects of LFchimera on these processes. It was found that LFchimera has the capacity to inhibit host-cell invasion by Yersiniae spp. in vitro. This effect appeared to be host-cell mediated, not bacteria-mediated. Furthermore it was found that exposure of human HeLa epithelial cells to both LFchimera and the bacterial strains evoked a pro-inflammatory cytokine release from the cells in vitro. [ABSTRACT FROM AUTHOR]

Among the properties of lactoferrin (LF) are bactericidal, antianemic, immunomodulatory, antitumour, antiphlogistic effects. Previously we demonstrated its capacity to stabilize in vivo HIF-1-alpha and HIF-2-alpha, which are redox-sensitive multiaimed transcription factors. Various tissues of animals receiving recombinant human LF (rhLF) responded by expressing the HIF-1-alpha target genes, hence such proteins as erythropoietin (EPO), ceruloplasmin, etc. were synthesized in noticeable amounts. Among organs in which EPO synthesis occurred were brain, heart, spleen, liver, kidneys and lungs. Other researchers showed that EPO can act as a protectant against severe brain injury and status epilepticus in rats. Therefore, we tried rhLF as a protector against the severe neurologic disorders developed in rats, such as the rotenone-induced model of Parkinson’s disease and experimental autoimmune encephalomyelitis as a model of multiple sclerosis, and observed its capacity to mitigate the grave symptoms. Moreover, an intraperitoneal injection of rhLF into mice 1 h after occlusion of the medial cerebral artery significantly diminished the necrosis area measured on the third day in the ischaemic brain. During this period EPO was synthesized in various murine tissues. It was known that EPO induces nuclear translocation of Nrf2, which, like HIF-1-alpha, is a transcription factor. In view that under conditions of hypoxia both factors demonstrate a synergistic protective effect, we suggested that LF activates the Keap1/Nrf2 signaling pathway, an important link in proliferation and differentiation of normal and malignant cells. J774 macrophages were cultured for 3 days without or in the presence of ferric and ferrous ions (RPMI-1640 and DMEM/F12, respectively). Then cells were incubated with rhLF or Deferiprone. Confocal microscopy revealed nuclear translocation of Nrf2 (the key event in Keap1/Nrf2 signaling) induced by apo-rhLF (iron-free, RPMI-1640). The reference compound Deferiprone (iron chelator) had the similar effect. Upon iron binding (in DMEM/F12) rhLF did not activate the Keap1/Nrf2 pathway. Added to J774, apo-rhLF enhanced transcription of Nrf2-dependent genes coding for glutathione S-transferase P and heme oxygenase-1. Western blotting revealed presence of Nrf2 in mice brain after 6 days of oral administration of apo-rhLF, but not Fe-rhLF or equivalent amount of PBS. Hence, apo-LF, but not holo-LF, induces the translocation of Nrf2 from cytoplasm to the nucleus, probably due to its capacity to induce EPO synthesis. [ABSTRACT FROM AUTHOR]

Herein we review commercial bovine lactoferrin quality issues by describing an example of industrial production, the current status of global quality standardization, and quality-activity concerns for further discussion. Morinaga Milk Industry has been industrially producing bovine lactoferrin in Milei GmbH, Germany, since 1989. We delineate its production and quality as an example of safe and high-quality manufacturing. Currently, global standardization in the quality of bovine lactoferrin is progressing through Novel Food and GRAS in the EU and USA, respectively. Novel Food was applied or notified to seven lactoferrin manufacturers and GRAS was notified to three manufacturers, two of which are for infant use and one is for adult use, by the end of 2017. The specifications of these regulations are relatively high, including more than 95% lactoferrin purity in protein, which means that such companies can supply relatively high-grade lactoferrin. There appear to be several concerns regarding lactoferrin quality affecting activities, including contamination of lipopolysaccharide (LPS) and angiogenin, purity, and degradation of lactoferrin sample. Although LPS is immunologically toxic when invading the body, it is distributed normally in foods and the gut. However, an industrial lactoferrin sample may contain LPS at a maximum LPS/lactoferrin molecule ratio = 1/1724, which means 99.9% of the lactoferrin molecule is LPS-free. It is difficult to speculate that LPS contained in a lactoferrin sample affects its activities. Finally in order to achieve good and reproducible results, we make proposals to researchers a use of high-grade lactoferrin, careful storage, and indication the manufacturers’ names and specifications in the paper. [ABSTRACT FROM AUTHOR]

Human lactoferrin is an iron-binding glycoprotein present at high concentrations in breast milk and colostrum. It is produced by many exocrine glands and widely distributed in a variety of body fluids. This protein has antimicrobial, immunomodulatory, antioxidant, and anticancer properties. Two important hLf receptors have been identified: LDL receptor related protein (LRP1), a low specificity receptor, and intelectin-1 (ITLN1), a high specificity receptor. No data are present on the role of hLf on the biliary epithelium. Our aims have been to evaluate the expression of Lf and its receptors in human and murine cholangiocytes and its effect on proliferation. Immunohistochemistry and immunofluorescence (IF) were conducted on human healthy and primary biliary cholangitis (PBC) liver samples as well as on liver samples obtained from normal and bile duct ligated (BDL) mice to evaluate the expression of Lf, LRP1 and ITLN1. Cell proliferation in vitro studies were performed on human cholangiocyte cell lines via 3-(4,5-dimetiltiazol-2-il)-2,5-diphenyltetrazolium assay as well as IF to evaluate proliferating cell nuclear antigen (PCNA) expression. Our results show that mouse and human cholangiocytes express Lf, LRP1 and ITLN1, at higher extent in cholangiocytes from BDL and PBC samples. Furthermore, the in vitro addition of bovine Lf (bLf) has a proliferative effect on human cholangiocyte cell line. The results support a proliferative role of hLf on the biliary epithelium; this pro-proliferative effect of hLf and bLf on cholangiocytes could be particularly relevant in human cholangiopathies such as PBC, characterized by cholangiocyte death and ductopenia. [ABSTRACT FROM AUTHOR]

Lactoferrin (LF) is a multifunctional protein in mammalian milk. We previously reported that enteric-coated bovine LF reduced the visceral fat in a double-blind clinical study. We further demonstrated that bovine LF (bLF) inhibited adipogenesis and promoted lipolysis in white adipocytes, but the effect of bLF on brown adipocytes has not been clarified. In this study, we investigated the effects of bLF on energy expenditure and cyclic adenosine monophosphate (cAMP)-protein kinase A (PKA) signaling pathway using human reprogrammed brown adipocytes generated by gene transduction. bLF at concentrations of ≥ 100 μg/mL significantly increased uncoupling protein 1 (UCP1) mRNA levels, with the maximum value observed 4 h after bLF addition. At the same time point, bLF stimulation also significantly increased oxygen consumption. Signaling pathway analysis revealed rapid increases of intracellular cAMP and cAMP response element-binding protein (CREB) phosphorylation levels beginning 5 min after bLF addition. The mRNA levels of peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC1α) were also significantly increased after 1 h of bLF stimulation. H-89, a specific PKA inhibitor, abrogated bLF-induced UCP1 gene expression. Moreover, receptor-associated protein (Rap), an antagonist of low-density lipoprotein receptor-related protein 1 (LRP1), significantly reduced bLF-induced UCP1 gene expression in a dose-dependent manner. These results suggest that bLF promotes UCP1 gene expression in brown adipocytes through the cAMP-PKA signaling pathway via the LRP1 receptor, leading to increased energy expenditure. [ABSTRACT FROM AUTHOR]

Human and bovine lactoferrin (hLf and bLf) are multifunctional iron-binding glycoprotein constitutively synthesized and secreted by glandular epithelial cells and by neutrophils following induction. HLf and bLf possess very high similarity of sequence. Therefore, most of the in vitro and in vivo studies are carried out with commercial bLf (cbLf), available in large quantities and recognized by Food and Drug Administration (FDA, USA) as a safe substance. Physico-chemical heterogeneity of different cbLf preparations influences their effectiveness. CbLf iron-saturation affects thermal stability and resistance to proteolysis. Moreover, other metal ions such as Al(III), Cu(II), Mg(II), Mn(II), Zn(II) are chelated by cbLf, even if at lower affinity than Fe(III). Ca(II) is also sequestered by the carboxylate groups of sialic acid present on glycan chains of cbLf thus provoking the release of LPS, contributing to bactericidal activity. Similarly to more than 50% of eukaryotic proteins, cbLf possesses five N-glycosylation sites, also contributing to the resistance to proteolysis and, putatively, to the protection of intestinal mucosa from pathogens. CbLfs possess several functions as anti-microbial, anti-biofilm, anti-adhesive, anti-invasive and anti-inflammatory activities. They are also relevant modulators of iron and inflammatory homeostasis. However, the efficacy of cbLfs in exerting several functions can be erratic mainly depending from integrity, degree of iron and other metal ions saturation, N-glycosylation sites and chains, desialylated forms, Ca(II) sequestration, presence of contaminants and finally the ability to enter inside nucleus. [ABSTRACT FROM AUTHOR]

In the last 20 years, several new genes and proteins involved in iron metabolism in eukaryotes, particularly related to pathological states both in animal models and in humans have been identified, and we are now starting to unveil at the molecular level the mechanisms of iron absorption, the regulation of iron transport and the homeostatic balancing processes. In this review, we will briefly outline the general scheme of iron metabolism in humans and then focus our attention on the cellular iron export system formed by the permease ferroportin and the ferroxidase ceruloplasmin. We will finally summarize data on the role of the iron binding protein lactoferrin on the regulation of the ferroportin/ceruloplasmin couple and of other proteins involved in iron homeostasis in inflamed human macrophages. [ABSTRACT FROM AUTHOR]

Rotaviral gastroenteritis causes a high rate of infant mortality and severe healthcare implications worldwide. Several studies have pointed out that human milk and dairy fractions, such as whey and buttermilk, possess antirotaviral activity. This activity has been mainly associated with glycoproteins, among them lactoferrin (LF). Thermal treatments are necessary to provide microbiological safety and extend the shelf life of milk products, though they may diminish their biological value. High hydrostatic pressure (HHP) treatment is a non-thermal method that causes lower degradation of food components than other treatments. Thus, the main objective of this study was to prove the antirotaviral activity of LFs from different origin and to evaluate the effect of several thermal and HHP treatments on that activity. LF exerted a high antirotaviral activity, regardless of its origin. Native LFs from bovine, ovine, swine and camel milk, and the human recombinant forms, at 1 mg/mL, showed neutralizing values in the range 87.5-98.6%, while human LF neutralized 58.2%. Iron saturation of bovine LF did not modify its antirotaviral activity. Results revealed interspecies differences in LFs heat susceptibility. Thus, pasteurization at 63 °C for 30 min led to a decrease of 60.1, 44.5, 87.1, 3.8 and 8% of neutralizing activity for human, bovine, swine, ovine and camel LFs, respectively. Pasteurization at 75 °C for 20 s was less harmful to the activity of LFs, with losses ranging from 0 to 13.8%. HHP treatment at 600 MPa for 15 min did not cause any significant decrease in the neutralizing activity of LFs. [ABSTRACT FROM AUTHOR]

Osteonecrosis of the jaws is an emerging pathological condition characterized by un-exposure or exposure of the necrotic bone, independently from the etiology. This term is usually referred to medication-related osteonecrosis of the jaws due to severe adverse reaction to certain medicines, as bisphosphonates, used for the treatment of cancer and osteoporosis. The management of patients with Bisphosphonate-Related Osteonecrosis of the Jaws (BRONJ) remains challenging because surgical and medical interventions may not eradicate this pathology. The goal of treatment of patients at risk of developing BRONJ or of those who have active disease is the preservation of quality of life by controlling pain, managing infection, and preventing the development of new areas of necrosis. The treatment of osteonecrosis consists in the surgical removal of necrotic bone followed by antibiotic therapy and application of sterile greasy gauze until the wound closure. The classical medical treatment has been compared with the innovative one consisting in the application of sterile greasy gauze soaked with bovine lactoferrin (bLf) after surgery. Here, for the first time, bLf efficacy on wound repair in subjects suffering from BRONJ with the progressive destruction of bone in the mandible or maxilla has been demonstrated. The positive results consist in a significant shorter time of wound closure (1 or 2 weeks) compared to that observed with classical surgical treatment (2-3 months). These promising results are an interesting tool for the innovative treatment of this pathology and for increasing the quality of life of these patients. [ABSTRACT FROM AUTHOR]

Lactoferrin (Lf) and transferrin (Tf) are iron-binding proteins that can bind various metal ions. This study demonstrates the heme-binding activity of bovine Lf and Tf using biotinylated hemin. When both proteins were coated on separate plate wells, each directly bound biotinylated hemin. On the other hand, when biotinylated hemin was immobilized on an avidin-coated plate, soluble native Lf bound to the immobilized biotinylated hemin whereas native Tf did not, suggesting that a conformational change triggered by coating on the plate allows the binding of denatured Tf with hemin. Incubation of Lf with hemin-agarose resulted in negligible binding of Lf with biotinylated hemin. Lf in bovine milk also bound to immobilized biotinylated hemin. These results demonstrate that bovine Lf has specific heme-binding activity, which is different from Tf, suggesting that either Tf lost heme-binding activity during its evolution or that Lf evolved heme-binding activity from its Tf ancestral gene. Additionally, Lf in bovine milk may bind heme directly, but may also bind heme indirectly by interaction with other milk iron- and/or heme-binding proteins. [ABSTRACT FROM AUTHOR]

Neutrophil gelatinase-associated lipocalin (NGAL) and lactoferrin (Lf) are among the key components of the innate immune system due to their ability to bind iron with high affinity and thus control inflammation. The aim of this study was to test the use of NGAL and LF measurements in meconium for the assessment of the intrauterine homeostasis. NGAL and Lf concentrations were measured using ELISA kits in all serial meconium portions (n = 81) collected from 20 healthy neonates. Mean ± SD meconium concentration of Lf was 45.07 ± 78.53 µg/g and more than 1000-fold higher compared with that of NGAL at 1.93 ± 2.46 ng/g. The correlation between the two proteins (r = 0.83, p < 0.0001) was found only for portions with Lf concentrations > 25 μg/g. High variability of NGAL and Lf concentrations in meconium and their correlations prove their key role as biomarkers of the fetal condition in utero. NGAL and Lf measured in meconium are candidate biomarkers for fetal iron status. [ABSTRACT FROM AUTHOR]

Bovine lactoferrin (bLf) is a natural iron-binding protein and it has been suggested to be a prebiotic agent, but this finding remains inconclusive. This study explores the prebiotic potential of bLf in 14 probiotics. Initially, bLf (1-32 mg/mL) treatment showed occasional and slight prebiotic activity in several probiotics only during the late experimental period (48, 78 h) at 37 °C. We subsequently supposed that bLf exerts stronger prebiotic effects when probiotic growth has been temperately retarded. Therefore, we incubated the probiotics at different temperatures, namely 37 °C, 28 °C, room temperature (approximately 22-24 °C), and 22 °C, to retard or inhibit their growth. As expected, bLf showed more favorable prebiotic activity in several probiotics when their growth was partially retarded at room temperature. Furthermore, at 22 °C, the growth of Bifidobacterium breve, Lactobacillus coryniformis, L. delbrueckii, L. acidophilus, B. angulatum, B. catenulatum, and L. paraplantarum were completely blocked. Notably, these probiotics started regrowing in the presence of bLf (1-32 mg/mL) in a significant and dose-dependent manner. Accordingly, bLf significantly increased the growth of Pediococcus pentosaceus, L. rhamnosus, and L. paracasei (BCRC 17483; a locally isolated strain) when their growth was retarded by incubation at 22 °C. In conclusion, bLf showed inconsistent prebiotic activity in the 14 probiotics at 37 °C, but revealed strong prebiotic activity in 10 probiotic strains at 22 °C. Therefore, this study enables determining additional roles of Lf in probiotic strains, which can facilitate developing novel combinational approaches by simultaneously using Lf and specific probiotics. [ABSTRACT FROM AUTHOR]

Translocation of bacteria, primarily Gram-negative pathogenic flora, from the intestinal lumen into the circulatory system leads to sepsis. In newborns, and especially very low birth weight infants, sepsis is a major cause of morbidity and mortality. The results of recently conducted clinical trials suggest that lactoferrin, an iron-binding protein that is abundant in mammalian colostrum and milk, may be an effective agent in preventing sepsis in newborns. However, despite numerous basic studies on lactoferrin, very little is known about how metal saturation of this protein affects a host's health. Therefore, the main objective of this study was to elucidate how iron-depleted, iron-saturated, and manganese-saturated forms of lactoferrin regulate intestinal barrier function via interactions with epithelial cells and macrophages. For these studies, a human intestinal epithelial cell line, Caco-2, was used. In this model, none of the tested lactoferrin forms induced higher levels of apoptosis or necrosis. There was also no change in the production of tight junction proteins regardless of lactoferrin metal saturation status. None of the tested forms induced a pro-inflammatory response in Caco-2 cells or in macrophages either. However, the various lactoferrin forms did effectively inhibit the pro-inflammatory response in macrophages that were activated with lipopolysaccharide with the most potent effect observed for apolactoferrin. Lactoferrin that was not bound to its cognate receptor was able to bind and neutralize lipopolysaccharide. Lactoferrin was also able to neutralize microbial-derived antigens, thereby potentially reducing their pro-inflammatory effect. Therefore, we hypothesize that lactoferrin supplementation is a relevant strategy for preventing sepsis. [ABSTRACT FROM AUTHOR]

Strongly pronounced argyrosis caused by adding AgCl to the feed of laboratory rats efficiently mimics the deficiency of ceruloplasmin (CP) ferroxidase activity. Bringing the concentration of AgCl in the feedstuff of lactating rats to 250 mg % and keeping their progeny (Ag-rats) for 3 months on the same silver-containing feed provided the serum iron content 1.4 times lower than that in the control group. Besides, the ferroxidase activity of CP dropped to zero. In CP purified from sera of Ag-rats two copper ions were substituted with two silver ions. Using rat models of both post-hemorrhagic and hemolytic anemia we showed that the deficiency of CP ferroxidase activity in Ag-rats affects the iron content in serum, though does not prevent the recovery of hemoglobin level accompanied by exhaustion of iron caches in liver and spleen. When apo-lactoferrin (apo-LF) was administered to Ag-rats suffering from either post-hemorrhagic or hemolytic anemia, both hemoglobin and serum iron were restored more rapidly than in the control animals. In independent experiments Ag-rats were compared with those fed on regular diet and the former displayed a prolonged 3-day stabilization of hypoxia-inducible factors 1 and 2 alpha (HIF-1a and HIF-2a) along with an increased serum concentration of erythropoietin. Introduction to Ag-rats of active CP separately or together with apo-LF reduced that effect to 1 day only. It is concluded that saturation of apo-LF with iron, provided by active CP, can strongly affect its protective capacity. [ABSTRACT FROM AUTHOR]

Lactoferrin (Lf), present in colostrum and milk is a member of the transferrin family of iron-binding glyco-proteins, with stronger binding capacity to ferric iron than hemoglobin, myoglobin or transferrin. Unlike hemoglobin and myoglobin, iron-bound Lf is reasonably stable to gastric and duodenal digestive conditions. Unlike ferrous iron, ferric iron is not directly reactive with oxygen supporting the capacity of Lf capture of heme iron to suppress reactive oxygen species (ROS) production. We therefore hypothesized that bovine Lf could capture and thereby terminate the cycle of ROS production by heme iron. The transfer of heme iron from either intact or digested forms of hemoglobin and myoglobin and from intact ferritin was demonstrated by in vitro methods, monitoring Fe-saturation status of Lf by changes in absorptivity at 465 nm. The results are discussed in the context of new proposed opportunities for orally administered Lf to regulate oxidative damage associated with heme iron. In addition to potentially suppressing oxidative heme-iron-mediated tissue damage in the lumen, Lf is expected to also reverse the overload of ferritin-bound iron, that accompanies chronic inflammation and aging. These new proposed uses of Lf are additional to known host defense functions that include anti-microbial, anti-viral properties, immune and cancer cell growth regulation. The findings and interpretations presented require clinical substantiation and may support important additional protective and therapeutic uses for Lf in the future. [ABSTRACT FROM AUTHOR]

Copper-containing plasma protein ceruloplasmin (Cp) forms a complex with lactoferrin (Lf), an iron-binding protein, and with the heme-containing myeloperoxidase (Mpo). In case of inflammation, Lf and Mpo are secreted from neutrophil granules. Among the plasma proteins, Cp seems to be the preferential partner of Lf and Mpo. After an intraperitoneal injection of Lf to rodents, the 'Cp-Lf' complex has been shown to appear in their bloodstream. Cp prevents the interaction of Lf with protoplasts of Micrococcus luteus. Upon immunoprecipitation of Cp, the blood plasma becomes depleted of Lf and in a dose-dependent manner loses the capacity to inhibit the peroxidase activity of Mpo, but not the Mpo-catalyzed oxidation of thiocyanate in the (pseudo)halogenating cycle. Antimicrobial effect against E. coli displayed by a synergistic system that includes Lf and Mpo-HO-chloride, but not thiocyanate, as the substrate for Mpo is abrogated when Cp is added. Hence, Cp can be regarded as an anti-inflammatory factor that restrains the halogenating cycle and redirects the synergistic system Mpo-HO-chloride/thiocyanate to production of hypothiocyanate, which is relatively harmless for the human organism. Structure and functions of the '2Cp-2Lf-Mpo' complex and binary complexes Cp-Lf and 2Cp-Mpo in inflammation are discussed. [ABSTRACT FROM AUTHOR]

Lactoferrin (LF) is an important antimicrobial and immune regulatory protein present in neutrophils and most exocrine secretions of mammals. The antimicrobial activity of LF has been related to the presence of an antimicrobial peptide sequence, called lactoferricin (LFcin), located in the N-terminal region of the protein. The antimicrobial activity of bovine LFcin is considerably stronger than the human version. In this work, chimera peptides combining segments of bovine and human LFcin were generated in order to study their antimicrobial activity and mechanism of action. In addition, the relevance of the conserved disulfide bridge and the resulting cyclic structure of both LFcins were analyzed by using 'click chemistry' and sortase A-catalyzed cyclization of the peptides. The N-terminal region of bovine LFcin (residues 17-25 of bovine LF) proved to be very important for the antimicrobial activity of the chimera peptides against E. coli, when combined with the C-terminal region of human LFcin. Similarly the cyclic bovine LFcin analogs generated by 'click chemistry' and sortase A preserved the antimicrobial activity of the original peptide, showing the significance of these two techniques in the design of cyclic antimicrobial peptides. The mechanism of action of bovine LFcin and its active derived peptides was strongly correlated with membrane leakage in E. coli and up to some extent with the ability to induce vesicle aggregation. This mechanism was also preserved under conditions of high ionic strength (150 mM NaCl) illustrating the importance of these peptides in a more physiologically relevant system. [ABSTRACT FROM AUTHOR]

Bovine lactoferrin (bLf) is a natural glycoprotein, and it shows broad-spectrum antimicrobial activity. However, reports on the influences of bLf on probiotic bacteria have been mixed. We examined the effects of apo-bLf (between 0.25 and 128 mg/mL) on both aerobic and anaerobic cultures of probiotics. We found that bLf had similar effects on the growth of probiotics under aerobic or anaerobic conditions, and that it actively and significantly (at concentrations of >0.25 mg/mL) retarded the growth rate of Bifidobacterium bifidum (ATCC 29521), B. longum (ATCC 15707), B. lactis (BCRC 17394), B. infantis (ATCC 15697), Lactobacillus reuteri (ATCC 23272), L. rhamnosus (ATCC 53103), and L. coryniformis (ATCC 25602) in a dose-dependent manner. Otherwise, minimal inhibitory concentrations (MICs) were 128 or >128 mg/mL against B. bifidum, B. longum, B. lactis, L. reuteri, and L. rhamnosus (ATCC 53103). With regard to MICs, bLf showed at least four-fold lower inhibitory effect on probiotics than on pathogens. Intriguingly, bLf (>0.25 mg/mL) significantly enhanced the growth of Rhamnosus (ATCC 7469) and L. acidophilus (BCRC 14065) by approximately 40-200 %, during their late periods of growth. Supernatants produced from aerobic but not anaerobic cultures of L. acidophilus reduced the growth of Escherichia coli by about 20 %. Thus, bLf displayed a dose-dependent inhibitory effect on the growth of most probiotic strains under either aerobic or anaerobic conditions. An antibacterial supernatant prepared from the aerobic cultures may have significant practical use. [ABSTRACT FROM AUTHOR]

Preterm neonates are at risk to acquire infections. In addition to the high mortality associated with sepsis, these patients are at risk for long-term disabilities, particularly neurodevelopment impairment. Several interventions have been evaluated to reduce rates of infections in neonates but have not proven efficacy. Lactoferrin (LF), a milk glycoprotein with anti-inflammatory, immunomodulatory and anti-microbial properties, has the potential to prevent infections in young children. We performed a review of current and ongoing clinical trials of LF for prevention of neonatal sepsis, and found eleven registered clinical trials that include more than 6,000 subjects. Few of these trials have finished; despite their small sample size, the preliminary results show a trend towards a positive protective effect of LF on neonatal infections. Larger trials are underway to confirm the findings of these initial studies. This information will help to define LF's role in clinical settings and, if proven effective, would profoundly affect the treatment of low birth weight neonates as a cost-effective intervention worldwide. [ABSTRACT FROM AUTHOR]

Conflicting data are reported on pro- or anti-inflammatory activity of bovine lactoferrin (bLf) in different cell models as phagocytes or epithelial cell lines infected by bacteria. Here we evaluated the bLf effect on epithelial models mimicking two human pathologies characterized by inflammation and infection with specific bacterial species. Primary bronchial epithelium from a cystic fibrosis (CF) patient and differentiated intestinal epithelial cells were infected with Pseudomonas aeruginosa LESB58 isolated from a CF patient and Adherent-Invasive Escherichia coli LF82 isolated from a Crohn's disease patient. Surprisingly, bLf significantly reduced the intracellular bacterial survival, but differently modulated the inflammatory response. These data lead us to hypothesize that bLf differentially acts depending on the epithelial model and infecting pathogen. To verify this hypothesis, we explored whether bLf could modulate ferroportin (Fpn), the only known cellular iron exporter from cells, that, by lowering the intracellular iron level, determines a non permissive environment for intracellular pathogens. Here, for the first time, we describe the bLf ability to up-regulate Fpn protein in infected epithelial models. Our data suggest that the mechanism underlying the bLf modulating activity on inflammatory response in epithelial cells is complex and the bLf involvement in modulating cellular iron homeostasis should be taken into account. [ABSTRACT FROM AUTHOR]

We herein summarized the effects of lactoferrin (LF) on bifidobacteria. Many in vitro studies previously reported the growth-promoting (bifidogenic) effects of LF on bifidobacteria. The involvement of bound iron, sugar chains, and LF peptides has been proposed in this bifidogenic mechanism. Peptides in the LF pepsin hydrolysate (LFH) showed stronger bifidogenic activity than natural LF; therefore, we speculated that peptides may be the bifidogenic active principle of LF. LF or its peptides may be recognized by LF-binding proteins on the surface of bifidobacterial cells, and the cationic nature or disulfide bonds of LF or its peptides may play a crucial role in its recognition by these proteins. Of the bifidobacterial species so far identified, human LF and peptides in human LFH were more likely to show bifidogenic activity especially to Bifidobacterium bifidum, and bovine LF (bLF) and peptides in bovine LFH (bLFH) to B. breve and B. infantis. In animal studies, the administration of LF to mice or piglets increased bifidobacteria levels in the intestine. In human trials, the administration of LF-containing formula to infants increased bifidobacteria levels in the feces; however, human milk achieved better results than LF-containing formula. In the case of breast-fed infants, LF may show bifidogenic activity synergistically with other milk components such as human milk oligosaccharides. As bLFH showed stronger bifidogenic activity than natural bLF, especially to B. breve and B. infantis in vitro, and these species are known to be infant-specific species, bLFH may be a beneficial ingredient in formula. [ABSTRACT FROM AUTHOR]

Iron balance is tightly linked to inflammation and it has been demonstrated that many proteins involved in cellular iron management are up- or down-regulated by inflammatory stimuli, ultimately leading to iron retention in the reticuloendothelial system. Ferroportin is a key player in maintenance of correct iron homeostasis, because it is the only known mammalian cellular iron exporter. In this work we show that incubation of THP-1 monocytes/macrophages with lactoferrin prevents the LPS-induced decrease of ferroportin by reducing secretion of IL-6. [ABSTRACT FROM AUTHOR]

Delta-lactoferrin (∆Lf) is a transcription factor belonging to the lactoferrin family, the expression of which inhibits cell proliferation and leads to Skp1 and DcpS gene transactivation. In this study, we showed that ∆Lf expression also induces cell death via apoptosis in HEK 293 and MCF7 cells using a cell viability assay and DNA fragmentation. Western blot analyses showed that apoptosis was caspase-9, 7 and 8 dependent. Proteolytic cleavage of the endonuclease PARP was significantly increased. The levels of expression of Bcl family members were detected by immunochemistry and showed that the Bcl-xl/Bax and Bcl-2/Bax protein ratios were decreased. We determined that the pro-apoptotic effects of ∆Lf are mainly mediated by the activation of the mitochondria-dependent death-signaling pathway. Apoptosis induction by ∆Lf is concomitant with increased cellular levels of Bax protein. Analysis of the Bax promoter region detected a ∆Lf response element located at −155 bp from the transcription start site. Both luciferase reporter gene and chromatin immunoprecipitation assays confirmed that ∆Lf interacts in vitro and in vivo specifically with this sequence. Its deletion, realized using directed mutagenesis, totally abolished ∆Lf transcriptional activity, identifying it as a ∆Lf-responsive element. These results indicate that the Bax gene is a novel ∆Lf target. Moreover we also showed that the O-GlcNAc/P interplay, which controls ∆Lf transcriptional activity, modulates Bax transactivation. [ABSTRACT FROM AUTHOR]