RESUMO
Hydrogels based on natural polysaccharides have demonstrated efficacy in epithelial recovery from cutaneous burn wounds. Here, we prepared a double-network hydrogel consisting of galactomannan (from Cassia grandis seeds) and κ-carrageenan (commercially sourced), cross-linked with CaCl2, as a matrix for immobilizing lactoferrin and/or Cramoll, aiming at its applicability as dressings for second-degree burn wounds. The formulations obtained [H - hydrogel, HL - hydrogel + lactoferrin, HC - hydrogel + Cramoll and HLC - hydrogel + lactoferrin + Cramoll] were analyzed rheologically as well as in terms of their stability (pH, color, microbial contamination) for 90 days. The burn was created with an aluminum bar (97 ± 3 °C) in the dorsal region of Wistar rats and subsequently treated with hydrogels (H, HL, HC, HLC) and control saline solution (S). The burn was monitored for 3, 7 and 14 days to evaluate the efficacy of the hydrogels in promoting wound healing. The hydrogels did not reveal significant pH or microbiological changes; there was an increase in brightness and a reduction in opacity for H. The rheological analysis confirmed the gel-like viscoelastic signature of the systems without substantial modification of the basic rheological characteristics, however HLC proved to be more rigid, due to rheological synergy when combining protein biomolecules. Macroscopic analyses confirmed centripetal healing with wound contraction: S < H < HC < HL < HLC. Histopathological analyses showed that hydrogel-treated groups reduced inflammation, tissue necrosis and fibrosis, while promoting re-epithelialization with focal acanthosis, especially in HLC due to a positive synergistic effect, indicating its potential as a promising therapy in the repair of burns.
Assuntos
Queimaduras , Carragenina , Galactose , Hidrogéis , Mananas , Ratos Wistar , Cicatrização , Hidrogéis/química , Mananas/química , Mananas/farmacologia , Animais , Queimaduras/terapia , Queimaduras/tratamento farmacológico , Carragenina/química , Cicatrização/efeitos dos fármacos , Ratos , Galactose/análogos & derivados , Galactose/química , Masculino , Lactoferrina/química , ReologiaRESUMO
The production of human recombinant proteins to be used for therapeutic or nutritional purposes must focus on obtaining a molecule that is as close as possible to the native human protein. This biotechnological tool has been documented in various studies published in recent decades, with lactoferrin being one of those that has generated the most interest, being a promising option for recombinant technology. However, stability studies including thermodynamic parameters have not been reported for recombinant lactoferrin (Lf). The objective of this work was to obtain the human recombinant protein using the yeast Komagataella phaffii to study structural changes modifying pH and temperature using circular dichroism spectroscopy (CD). Thermodynamic parameters such as ΔH, ΔS and Tm were calculated and compared with commercial human lactoferrin. We propose the potential use of CD and thermodynamic parameters as a criterion in the production of recombinant proteins to be used in the production of specialized recombinant proteins.
Assuntos
Lactoferrina , Humanos , Lactoferrina/química , Dicroísmo Circular , Proteínas Recombinantes/metabolismo , Temperatura , Concentração de Íons de HidrogênioRESUMO
Liver cancer and leukemia are the fourth and first causes, respectively, of cancer death in children and adults worldwide. Moreover, cancer treatments, although beneficial, remain expensive, invasive, toxic, and affect the patient's quality of life. Therefore, new anticancer agents are needed to improve existing agents. Because bovine lactoferrin (bLF) and its derived peptides have antitumor properties, we investigated the anticancer effect of bLF and LF peptides (LFcin17-30, LFampin265-284 and LFchimera) on liver cancer HepG2 cells and leukemia Jurkat cells. HepG2 and Jurkat cells were incubated with bLF and LF peptides. Cell proliferation was quantified by an MTT assay, and cell morphology and damage were visualized by light microscopy or by phalloidin-TRITC/DAPI staining. The discrimination between apoptosis/necrosis was performed by staining with Annexin V-Alexa Fluor 488 and propidium iodide, and the expression of genes related to apoptosis was analyzed in Jurkat cells. Finally, the synergistic interaction of bLF and LF peptides with cisplatin or etoposide was assessed by an MTT assay and the combination index. The present study demonstrated that bLF and LF peptides inhibited the viability of HepG2 and Jurkat cells, inducing damage to the cell monolayer of HepG2 cells and morphological changes in both cell lines. bLF, LFcin17-30, and LFampin265-284 triggered apoptosis in both cell lines, whereas LFchimera induced necrosis. These results suggested that bLF and LF peptides activate apoptosis by increasing the expression of genes of the intrinsic pathway. Additionally, bLF and LF peptides synergistically interacted with cisplatin and etoposide. In conclusion, bLF and LF peptides display anticancer activity against liver cancer and leukemia cells, representing an alternative or improvement in cancer treatment.
Assuntos
Lactoferrina , Neoplasias Hepáticas , Criança , Humanos , Lactoferrina/farmacologia , Lactoferrina/química , Células Jurkat , Células Hep G2 , Cisplatino , Etoposídeo , Qualidade de Vida , Peptídeos/farmacologia , Neoplasias Hepáticas/tratamento farmacológico , NecroseRESUMO
Lactoferrin (L), chitosan (C) and gellan (G) nanoparticles were produced by electrostatic complexation, aiming to enhance the antimicrobial characteristics of these compounds. Binary complexes (lactoferrin-gellan and chitosan-gellan) and ternary complexes (lactoferrin-chitosan-gellan) were studied in eight different proportions of biopolymers. The antimicrobial activity of nanoparticles against S. aureus was compared with that of pure biopolymers and related to nanoparticle size, charge density and morphology. Those with a 4.5L:4.5C:1G ratio presented the highest positive charge density (+57.90 ± 1.50) mV and smaller hydrodynamic diameter (53.53 ± 2.06) nm, which resulted in the highest antimicrobial action (minimum inhibitory concentration of 0.0117 mg/ml). When applied to a coating of fresh strawberries, the nanoparticles were effective in preserving their physicochemical properties, especially in the presence of carboxymethylcellulose that enhanced the adhesion of particles to the fruits. In this study, the antimicrobial action of nanoparticles was better than that of lactoferrin and pure chitosan alone, proving that the antimicrobial properties of such biopolymers were enhanced due to aggregation at the nanoscale. The nanoparticles produced have outstanding application potential as natural food preservers.
Assuntos
Anti-Infecciosos , Quitosana , Fragaria , Nanopartículas , Antibacterianos/farmacologia , Anti-Infecciosos/química , Anti-Infecciosos/farmacologia , Biopolímeros/química , Biopolímeros/farmacologia , Quitosana/química , Quitosana/farmacologia , Lactoferrina/química , Lactoferrina/farmacologia , Nanopartículas/química , Polissacarídeos Bacterianos , Staphylococcus aureusRESUMO
Understanding nonionic surfactant-protein interactions is fundamental from both technological and scientific points of view. However, there is a complete absence of kinetic data for such interactions. We employed surface plasmon resonance (SPR) to determine the kinetic and thermodynamic parameters of bovine lactoferrin-Brij58 interactions at various temperatures under physiological conditions (pH 7.4). The adsorption process was accelerated with increasing temperature, while the desorption rate decreased, resulting in a more thermodynamically stable complex. The kinetic energetic parameters obtained for the formation of the activated complex, [bLF-Brij58], indicated that the potential energy barrier for [bLF-Brij58] formation arises primarily from the reduction in system entropy. [bLF-Brij58]â formation was entropically driven, indicating that hydrophobic interactions play a fundamental role in bLF interactions with Brij58.
Assuntos
Cetomacrogol/metabolismo , Lactoferrina/metabolismo , Tensoativos/metabolismo , Temperatura , Adsorção , Cetomacrogol/química , Entropia , Concentração de Íons de Hidrogênio , Interações Hidrofóbicas e Hidrofílicas , Cinética , Lactoferrina/química , Ligação Proteica , Ressonância de Plasmônio de Superfície , Tensoativos/químicaRESUMO
Chimeric peptides containing short sequences derived from bovine Lactoferricin (LfcinB) and Buforin II (BFII) were synthetized using solid-phase peptide synthesis (SPPS) and characterized via reversed-phase liquid chromatography and mass spectrometry. The chimeras were obtained with high purity, demonstrating their synthetic viability. The chimeras' antibacterial activity against Gram-positive and Gram-negative strains was evaluated. Our results showed that all the chimeras exhibited greater antibacterial activity against the evaluated strains than the individual sequences, suggesting that chemical binding of short sequences derived from AMPs significantly increased the antibacterial activity. For each strain, the chimera with the best antibacterial activity exerted a bacteriostatic and/or bactericidal effect, which was dependent on the concentration. It was found that: (i) the antibacterial activity of a chimera is mainly influenced by the linked sequences, the palindromic motif RLLRRLLR being the most relevant one; (ii) the inclusion of a spacer between the short sequences did not significantly affect the chimera's synthesis process; however, it enhanced its antibacterial activity against Gram-negative and Gram-positive strains; on the other hand, (iii) the replacement of Arg with Lys in the LfcinB or BFII sequences improved the chimeras' synthesis process without significantly affecting their antibacterial activity. These results illustrate the great importance of the synthesis of chimeric peptides for the generation of promising antibacterial peptides.
Assuntos
Antibacterianos/química , Lactoferrina/química , Fragmentos de Peptídeos/química , Proteínas/química , Animais , Antibacterianos/síntese química , Antibacterianos/farmacologia , Bactérias/efeitos dos fármacos , Infecções Bacterianas/tratamento farmacológico , Bovinos , Humanos , Lactoferrina/síntese química , Lactoferrina/farmacologia , Fragmentos de Peptídeos/síntese química , Fragmentos de Peptídeos/farmacologia , Proteínas/síntese química , Proteínas/farmacologia , Técnicas de Síntese em Fase SólidaRESUMO
Due to the emergence of multidrug-resistant pathogens, it is necessary to develop options to fight infections caused by these agents. Lactoferrin (Lf) is a cationic nonheme multifunctional glycoprotein of the innate immune system of mammals that provides numerous benefits. Lf is bacteriostatic and/or bactericidal, can stimulate cell proliferation and differentiation, facilitate iron absorption, improve neural development and cognition, promote bone growth, prevent cancer and exert anti-inflammatory and immunoregulatory effects. Lactoferrin is present in colostrum and milk and is also produced by the secondary granules of polymorphonuclear leukocytes, which store this glycoprotein and release it at sites of infection. Lf is also present in many fluids and exocrine secretions, on the surfaces of the digestive, respiratory and reproductive systems that are commonly exposed to pathogens. Apo-Lf (an iron-free molecule) can be microbiostatic due to its ability to capture ferric iron, blocking the availability of host iron to pathogens. However, apo-Lf is mostly microbicidal via its interaction with the microbial surface, causing membrane damage and altering its permeability function. Lf can inhibit viral entry by binding to cell receptors or viral particles. Lf is also able to counter different important mechanisms evolved by microbial pathogens to infect and invade the host, such as adherence, colonization, invasion, production of biofilms and production of virulence factors such as proteases and toxins. Lf can also cause mitochondrial and caspase-dependent regulated cell death and apoptosis-like in pathogenic yeasts. All of these mechanisms are important targets for treatment with Lf. Holo-Lf (the iron-saturated molecule) can contain up to two ferric ions and can also be microbicidal against some pathogens. On the other hand, lactoferricins (Lfcins) are peptides derived from the N-terminus of Lf that are produced by proteolysis with pepsin under acidic conditions, and they cause similar effects on pathogens to those caused by the parental Lf. Synthetic analog peptides comprising the N-terminus Lf region similarly exhibit potent antimicrobial properties. Importantly, there are no reported pathogens that are resistant to Lf and Lfcins; in addition, Lf and Lfcins have shown a synergistic effect with antimicrobial and antiviral drugs. Due to the Lf properties being microbiostatic, microbicidal, anti-inflammatory and an immune modulator, it represents an excellent natural alternative either alone or as adjuvant in the combat to antibiotic multidrug-resistant bacteria and other pathogens. This review aimed to evaluate the data that appeared in the literature about the effects of Lf and its derived peptides on pathogenic bacteria, protozoa, fungi and viruses and how Lf and Lfcins inhibit the mechanisms developed by these pathogens to cause disease.
Assuntos
Anti-Infecciosos/química , Anti-Infecciosos/farmacologia , Lactoferrina/química , Lactoferrina/farmacologia , Peptídeos/química , Peptídeos/farmacologia , Animais , Anti-Infecciosos/síntese química , Antiparasitários/síntese química , Antiparasitários/química , Antiparasitários/farmacologia , Bactérias/efeitos dos fármacos , Aderência Bacteriana/efeitos dos fármacos , Parede Celular/efeitos dos fármacos , Técnicas de Química Sintética , Fungos/efeitos dos fármacos , Interações Hospedeiro-Patógeno , Humanos , Peptídeos/síntese química , Proteólise/efeitos dos fármacos , Relação Estrutura-Atividade , Virulência/efeitos dos fármacos , Fatores de Virulência , Vírus/efeitos dos fármacosRESUMO
Naringenin (NG) is a flavonoid with many bioactive properties, however, its bitterness limits its use in foods. It is known that complex formation with proteins can mask this undesirable sensory property. Therefore, a trained panel evaluated the effect of bovine lactoferrin (LF) on NG bitterness using time-intensity analysis. LF reduced the maximum bitterness intensity and overall bitterness perception for NG by 27% and 33%, respectively. Isothermal titration nanocalorimetry (ITC), molecular docking (DC), and molecular dynamics (MD) were used to characterize NG-LF binding. These techniques provided similar values of ΔG° for binding ( [Formula: see text] = -33.42 kJ mol-1; [Formula: see text] = -32.22 kJ mol-1; [Formula: see text] = -31.84 kJ mol-1). ITC showed that the complex formation is primarily entropy driven and DC suggested that NG binds at a hydrophobic site in LF. Here are presented strategic tools for promoting NG incorporation in food and health products.
Assuntos
Flavanonas/metabolismo , Flavanonas/farmacologia , Lactoferrina/química , Lactoferrina/metabolismo , Paladar , Adulto , Animais , Calorimetria/métodos , Bovinos , Entropia , Feminino , Flavanonas/química , Humanos , Interações Hidrofóbicas e Hidrofílicas , Masculino , Simulação de Acoplamento Molecular , Simulação de Dinâmica Molecular , TermodinâmicaRESUMO
Black pepper essential oil (EO) was encapsulated by complex coacervation with lactoferrin and sodium alginate using transglutaminase as a cross-linking agent. The encapsulation efficiency varied from 31.66 to 84.48%. Chemical and morphological characteristics suggest that the EO was encapsulated in a lactoferrin/sodium alginate shell. The chemical composition of the encapsulated EO was identified by gas chromatography (GC) and nuclear magnetic resonance (NMR). The GC and NMR analyses indicated good core protection with the materials used. The stability of the black pepper EO capsules under in vitro digestion was evaluated. Theses capsules demonstrated the low release of the EO during gastric digestion and higher release in intestinal digestion. These results suggest that these capsules can be used to transport active ingredients and that they are resistant to oral and gastric conditions that were tested in vitro.
Assuntos
Alginatos/química , Lactoferrina/química , Óleos Voláteis/química , Piper nigrum/química , Cápsulas/química , Cromatografia GasosaRESUMO
Lactoferrin (LF) is a glycoprotein that serves as a potential vehicle for small bioactive molecules in food. In an effort to improve this functionality, the kinetic and thermodynamic interaction of LF with naringin (NR) was studied by surface plasmon resonance (SPR). The results demonstrated that the association rate constant between LF and NR was 5.00â¯×â¯104â¯M-1â¯s-1, while the dissociation rate of the complex was 0.36â¯s-1, at 25⯰C. The stable complex predominated over free molecules (ΔG25°C0=-29.35â¯kJâ¯mol-1), and the binding constant was 1.39â¯×â¯105â¯M-1, at 25⯰C. The association of LF and NR to form an intermediate complex occurred in multi-steps. Nevertheless, the intermediate complex formation from the dissociation of the stable complex occurred in a single step with the activation energy independent of temperature. This study provides an important basis to explore LF as a vehicle for bioactive molecules.