RESUMO
Novel sulfonated derivatives of poly(allylamine hydrochloride) (NSPAHs) and N-sulfonated chitosan (NSCH) have been synthesized, and their activity against influenza A and B viruses has been studied and compared with that of a series of carrageenans, marine polysaccharides of well-documented anti-influenza activity. NSPAHs were found to be nontoxic and very soluble in water, in contrast to gel-forming and thus generally poorly soluble carrageenans.In vitroandex vivostudies using susceptible cells (Madin-Darby canine kidney epithelial cells and fully differentiated human airway epithelial cultures) demonstrated the antiviral effectiveness of NSPAHs. The activity of NSPAHs was proportional to the molecular mass of the chain and the degree of substitution of amino groups with sulfonate groups. Mechanistic studies showed that the NSPAHs and carrageenans inhibit influenza A and B virus assembly in the cell.
Assuntos
Antivirais/farmacologia , Quitosana/farmacologia , Vírus da Influenza A Subtipo H1N1/efeitos dos fármacos , Vírus da Influenza A Subtipo H3N2/efeitos dos fármacos , Vírus da Influenza B/efeitos dos fármacos , Poliaminas/farmacologia , Polímeros/farmacologia , Ésteres do Ácido Sulfúrico/farmacologia , Animais , Antivirais/síntese química , Quitosana/síntese química , Cães , Células Epiteliais/efeitos dos fármacos , Células Epiteliais/virologia , Humanos , Vírus da Influenza A Subtipo H1N1/genética , Vírus da Influenza A Subtipo H1N1/crescimento & desenvolvimento , Vírus da Influenza A Subtipo H3N2/genética , Vírus da Influenza A Subtipo H3N2/crescimento & desenvolvimento , Vírus da Influenza B/genética , Vírus da Influenza B/crescimento & desenvolvimento , Concentração Inibidora 50 , Células Madin Darby de Rim Canino , Poliaminas/síntese química , Polieletrólitos , Polímeros/síntese química , RNA Viral/antagonistas & inibidores , RNA Viral/biossíntese , Relação Estrutura-Atividade , Ésteres do Ácido Sulfúrico/síntese química , Montagem de Vírus/efeitos dos fármacos , Ligação Viral/efeitos dos fármacos , Inativação de Vírus/efeitos dos fármacos , Internalização do Vírus/efeitos dos fármacos , Replicação Viral/efeitos dos fármacosRESUMO
In this paper, we present a facile method of synthesis and modification of poly(glycidyl methacrylate) brushes with 6-aminofluorescein (6AF) molecules. Polymer brushes were obtained using surface-grafted atom transfer radical polymerization (SI-ATRP) and functionalized in the presence of triethylamine (TEA) acting both as a reaction catalyst and an agent preventing aggregation of chromophores. Atomic force microscopy (AFM), FTIR, X-ray photoelectron spectroscopy (XPS) were used to study the structure and formation of obtained photoactive platforms. UV-Vis absorption and emission spectroscopy and confocal microscopy were conducted to investigate photoactivity of chromophores within the macromolecular matrix. Owing to the simplicity of fabrication and good ordering of the chromophore in a thin nanometric layer, the proposed method may open new opportunities for obtaining light sensors, photovoltaic devices, or other light-harvesting systems.
RESUMO
This publication presents the synthesis of core-shell nanoparticles, where the core was Ni, and the shell was a Ag-Ni nano alloy. The synthesis was based on the reduction of Ni and Ag ions with sodium borohydride in the presence of trisodium citrate as a stabilizer. In order to determine the phase composition of the obtained nanoparticles, an XRD study was performed, and in order to identify the oxidation states of the nanoparticle components, an XPS spectroscopic study was performed. The composition and shape of the particles were determined using the HR-TEM EDS test. The obtained nanoparticles had a size of 11 nm. The research on catalytic properties was carried out in the model methylene blue reduction system. The investigation of the catalytic activity of colloids was carried out with the use of UV-Vis spectrophotometry. The Ag-Ni alloy was about ten times more active than were pure silver nanoparticles of a similar size.
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Cartilage loss is a common clinical problem, which leads to significant pain, dysfunction, and even disability. As a result, there is growing interest in using small, non-protein molecules to protect or repair cartilage. Kartogenin (KGN), a small hydrophobic molecule, shows chondroprotective and chondrogenic properties. In this study, we embedded KGN in liposomes, and the whole system was stabilized by covering it with n-octadecylated (at two different substitution degrees) chondroitin sulfate (CS) derivatives. We investigated the interactions of empty liposomes and KGN-loaded liposomes with both CS derivatives using various physicochemical techniques, which revealed that hydrophobically modified CSs can interact with both neutral lipid membrane and negatively charged loaded-KGN lipid membrane. The cytotoxicity and chondrogenic properties of the polysaccharides and liposome-CS formulations of KGN were analyzed towards mesenchymal stem cells (MSCs). The results showed that the alkylated CS exhibited cytotoxic properties. The higher substituted CS self-assembles into stable nanoaggregates that can form a corona on the surface of liposomes, eliminating the overall cytotoxicity of this polymer. However, all tested chondrogenic markers' expression levels are enhanced for KGN-loaded liposomes and coated by lower substituted CS. Furthermore, the undesirable hypertrophy effect for this formulation significantly decreased compared to pure polymeric derivative.
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Despite the plethora of research that exists on recombinant human bone morphogenetic protein-2 and -7 (rhBMP-2 and rhBMP-7) and has been clinically approved, there is still a need to gain information that would allow for their more rational use in bone implantology. The clinical application of supra-physiological dosages of these superactive molecules causes many serious adverse effects. At the cellular level, they play a role in osteogenesis and cellular adhesion, migration, and proliferation around the implant. Therefore, in this work, we investigated the role of the covalent binding of rhBMP-2 and rhBMP-7 separately and in combination with ultrathin multilayers composed of heparin and diazoresin in stem cells. In the first step, we optimized the protein deposition conditions via quartz crystal microbalance (QCM). Then, atomic force microscopy (AFM) and enzyme-linked immunosorbent assay (ELISA) were used to analyze protein-substrate interactions. The effect of the protein binding on the initial cell adhesion, migration, and short-term expression of osteogenesis markers was tested. In the presence of both proteins, cell flattening and adhesion became more prominent, resulting in limited motility. However, the early osteogenic marker expression significantly increased compared to the single protein systems. The presence of single proteins resulted in the elongation of cells, which promoted their migration activity.
Assuntos
Heparina , Fator de Crescimento Transformador beta , Humanos , Heparina/farmacologia , Proteína Morfogenética Óssea 2/metabolismo , Compostos Azo/farmacologia , Osteogênese , Proteínas Recombinantes/metabolismo , Diferenciação CelularRESUMO
HYPOTHESIS: Mixed polymer brushes (MPBs) could be synthesized by surface dilution of homopolymer brushes and subsequent grafting of other type of chains in the formed voids. Nanophase separation and dynamics of surface-grafted chains could be tailored by modification of their molecular architecture. Mixed polyelectrolyte and conjugated chains contribute synergistically to tailor properties of the coating. EXPERIMENTS: A new synthetic strategy that allowed spatially controlled grafting of poly(sodium 4-styrenesulfonate) chains (PSSNa) in close neighborhood of poly(3-methylthienyl methacrylate) (PMTM) brushes (precursors of the conjugated chains) using surface-initiated polymerizations was developed. The final mixed conjugated/polyelectrolyte brushes were prepared by template polymerization of pendant thiophene groups in PMTM chains. Surface dynamics and nanophase separation of MPBs were studied by nanoscale resolution IR imaging, SIMS profiling and AFM mapping in selective solvents. FINDINGS: Unconjugated MPBs were shown to undergo vertical, and horizontal nanophase separation, while the size and shape of the nanodomains were dependent on molar ratio of the mixed chains and their relative lengths. Generation of the conjugated chains led to diminishing of nanophase separation thanks to stronger mutual interactions of conjugated PMTM and PSSNa (macromolecular mixing). The obtained systems demonstrated tunable interfacial structure and resistance switching phenomenon desired in construction of smart surfaces or memristive devices.
Assuntos
Metacrilatos , Polímeros , Polieletrólitos , Propriedades de Superfície , Polímeros/química , Metacrilatos/químicaRESUMO
The interactions between synthetic polycations and phospholipid bilayers play an important role in some biophysical applications such as gene delivery or antibacterial usage. Despite extensive investigation into the nature of these interactions, their physical and molecular bases remain poorly understood. In this Article, we present the results of our studies on the impact of a hydrophobically modified strong polycation on the properties of a zwitterionic bilayer used as a model of the mammalian cellular membrane. The study was carried out using a set of complementary experimental methods and molecular dynamic (MD) simulations. A new polycation, poly(allyl-N,N-dimethyl-N-hexylammonium chloride) (polymer 3), was synthesized, and its interactions with liposomes composed of 2-oleoyl-1-palmitoyl-sn-glycero-3-phosphocholine (POPC) were examined using dynamic light scattering (DLS), zeta potential measurements, and cryo-transmission electron microscopy (cryo-TEM). Our results have shown that polymer 3 can efficiently associate with and insert into the POPC membrane. However, it does not change its lamellar structure, as was demonstrated by cryo-TEM. The influence of polymer 3 on the membrane functionality was studied by leakage experiments applying a fluorescence dye (calcein) encapsulated in the phospholipid vesicles. The MD simulations of model systems reveal that polymer 3 promotes formation of hydrophilic pores in the membrane, thus increasing considerably its permeability.
Assuntos
Lipídeos/química , Poliaminas/química , Espectroscopia de Ressonância Magnética , Simulação de Dinâmica Molecular , Polieletrólitos , Espectrofotometria InfravermelhoRESUMO
In this study, the protocol of a single-step L-cysteine functionalized silver nanoparticle synthesis was described. Particle size distribution was determined. The crystallinity and chemical properties were investigated using XRD, HR-TEM, and XPS methods. Acute toxicity and irritant properties of obtained nanoparticles were studied using mice and rats as an animal model. The results showed that thanks to the applied protocol, it was possible to synthesize silver nanoparticles with narrow particle size distribution. Moreover, the concentration of final product was extremely high in comparison to other known methods. These nanoparticles showed neither irritant properties nor acute toxicity.
Assuntos
Cisteína/administração & dosagem , Cisteína/síntese química , Portadores de Fármacos/administração & dosagem , Portadores de Fármacos/síntese química , Nanopartículas Metálicas/química , Prata/química , Animais , Feminino , Masculino , Camundongos , Ratos , Ratos Wistar , Pele/efeitos dos fármacos , Pele/metabolismo , Testes de Toxicidade Aguda/métodos , Difração de Raios X/métodosRESUMO
Cationic polymers have shown great potential for the delivery of proteins, nucleic acids forming complexes, called polyplexes. The most important issue in the context of using cationic polymers as carriers is the balance between the high transfection efficiency and low cytotoxicity. In this chapter, we report the preparation of polyallylamine derivatives mainly based on substitution of amino groups by glycidyltrimethylammonium chloride. The resulting polyplexes enhance the transfection of HeLa cell line without cytotoxic effects. Here, we describe methods for preparation and characterization of polyplexes using dynamic light scattering, ζ-potential measurements, gel retardation assay, and atomic force microscopy. Moreover, we provide protocols for the transfection of HeLa cell line by polyplexes, determination of their cytotoxicity, cell uptake, and intracellular trafficking.
Assuntos
DNA/genética , Poliaminas/química , Poliaminas/farmacologia , Animais , Linhagem Celular , Difusão Dinâmica da Luz , Células HeLa , Humanos , Lipossomos , Camundongos , Microscopia de Força Atômica , Poliaminas/síntese química , TransfecçãoRESUMO
Polyelectrolyte multilayers (PEMs) have found application in modifying material surfaces to make them adhesive or non-adhesive for animal cells. However, PEMs made of strong polyelectrolytes are not fully recognized in the literature. This study focuses on the interplay between the properties of PEM assembled from strong polyelectrolytes and cell adhesion and motility. Strong polycations (with quaternary ammonium groups) and a polyanion (with sulfonate groups) were obtained by modification of poly(allylamine hydrochloride) (PAH). Two types of multilayer films were assembled from these PAH derivatives and used to investigate the behavior of human skin fibroblasts (HSFs). The effect of surface charge, hydrophobicity, and film thickness on adhesion of HSFs in a serum-containing medium was studied with immunofluorescence microscopy. The results showed that adhesion of HSFs was strongly depended on the chemical functions of the terminal layer, whereas the wettability was not important. The surface of PEM can be strongly cytophobic (the quaternary ammonium terminal groups) or strongly cytophilic (the sulfonate terminal groups). Finally, the motile activity of HSFs seeded on glass coated with a varying number of polymer layers was investigated. It was demonstrated using an in vitro model that coating the substrate with only two polymer layers can considerably increase the average speed of HSFs movement and stimulate cell migration into the wound.
Assuntos
Movimento Celular , Eletrólitos/química , Fibroblastos/citologia , Polímeros/química , Pele/citologia , Adesão Celular/efeitos dos fármacos , Movimento Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Relação Dose-Resposta a Droga , Fibroblastos/efeitos dos fármacos , Humanos , Polímeros/farmacologia , Pele/efeitos dos fármacos , Relação Estrutura-Atividade , Propriedades de SuperfícieRESUMO
Non-viral gene carriers for safe and efficient gene transfection have become of particular interest among researchers of different disciplines ranging from physical chemistry to biotechnology. Recently polymeric vectors have been extensively studied as potentially new gene transfer agents. Until now most of the research efforts were made to optimize the gene-to-polymer weight ratio of polyplexes for safe and efficient gene transfection. In this work, we report on the development of novel poly(allylamine) derivatives with different balance of the primary, secondary, tertiary, and quaternary amino groups. All derivatives were able to complex pDNA into polyplexes at low gene-to-polymer weight ratios i.e., 1:1 or 1:2. Moreover, the examined polyplexes were less cytotoxic and showed better transfection efficiency when compared to linear poly(ethyleneimine). These results indicate that the presence of quaternary ammonium groups is important in the formation of stable polyplexes. Polymers with all types of amino groups showed large potential for gene delivery. Furthermore, polyplexes with such derivatives were well internalized by cells and ended up into acidic late endosomes.
Assuntos
DNA/administração & dosagem , Técnicas de Transferência de Genes , Poliaminas/administração & dosagem , Transporte Biológico , Sobrevivência Celular/efeitos dos fármacos , DNA/química , Endossomos/metabolismo , Células HeLa , Humanos , Luciferases/genética , Luciferases/metabolismo , Plasmídeos , Poliaminas/químicaRESUMO
Liposomes are used for in vitro or in vivo vectorization of drugs, proteins, or nucleic acids. However, the main problem with the application of liposomes for this purpose is their low stability in contact with blood serum. In this article, interactions between the whole serum and anionic liposomes, both bare and covered with strong polycations, were studied. The polycations of different chemical structures were prepared by the modification of poly(allylamine hydrochloride) (PAH). Dynamic light scattering (DLS), zeta potential and transmission cryo-electron microscopy (cryo-TEM) measurements showed that the adsorption of the polycations on the anionic liposomes induced a reversible aggregation of vesicles. The stable isolated polyelectrolyte-covered vesicles were obtained after the addition of sufficient amounts of the polycations. The effect of full serum on the morphology and stability of the polycation-coated liposomes was studied using cryo-TEM and a fluorescence method. The cryo-TEM analysis revealed that the introduction of serum caused the osmotic-driven destabilization of the bare liposomes or formation of twinned vesicles. Due to these processes the liposomes lost most of their content immediately after serum addition. The polycation-covered liposomes showed improved stability in the presence of serum. Partial deflation of the vesicles was observed, however, the loss of the content was significantly limited. The effect of the polymer structure, especially the position of the charged groups with respect to the main polymer backbone, on the stabilization of the polycation-covered liposomes in the presence of serum was discussed.
Assuntos
Microscopia Crioeletrônica , Eletrólitos/química , Lipossomos/química , Lipossomos/ultraestrutura , Microscopia Eletrônica de Transmissão , Soro/química , Animais , Bovinos , Fluoresceínas/metabolismo , Hidrodinâmica , Fosfatidilcolinas/química , Poliaminas/química , Polieletrólitos , Polímeros/química , Pressão , Eletricidade EstáticaRESUMO
Poly(allylamine hydrochloride) (PAH) has found many applications both in biotechnology and biomedical fields. However, its high toxicity toward various mammalian cells significantly limits its effective usage. This study focuses on improving the biological properties of PAH by its modification to strong polyelectrolytes. The strong polycations were prepared by the direct quaternization of PAH amino groups or by the attachment of glycidyltrimethylammonium chloride to these groups. The biological properties, such as cytotoxicity toward human skin fibroblasts (HSFs), proliferation and migration of the cells on a polymeric surface, and antibacterial activities against two pathogenic bacteria, Staphylococcus aureus and Escherichia coli, were determined. All the modified polyelectrolytes are considerably less toxic to HSFs as compared to PAH. Moreover, the directly quaternized polycations are stronger biocides against S. aureus than the parent polymer. Contrary to PAH, thin films of the modified polyelectrolytes improve or do not affect HSFs proliferation and can stimulate cell migration into the wound, as was demonstrated using an in vitro model. The relationship between the structure of the modified polymers (amount and localization of the quaternary ammonium groups) and the biological activity is discussed. Due to the improved biological properties, the obtained polycations may be potentially useful for a variety of biotechnological and biomedical applications.
Assuntos
Antibacterianos/química , Materiais Biocompatíveis/química , Poliaminas/química , Adulto , Antibacterianos/síntese química , Antibacterianos/farmacologia , Materiais Biocompatíveis/síntese química , Materiais Biocompatíveis/farmacologia , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Células Cultivadas , Escherichia coli/efeitos dos fármacos , Infecções por Escherichia coli/prevenção & controle , Fibroblastos/citologia , Fibroblastos/efeitos dos fármacos , Humanos , Poliaminas/síntese química , Poliaminas/farmacologia , Polieletrólitos , Infecções Estafilocócicas/prevenção & controle , Staphylococcus aureus/efeitos dos fármacosRESUMO
Molecular dynamics simulation has been used to study the specific interactions between poly(ethylene glycol) (PEG) and three drug molecules for which PEG is used to aid delivery: paclitaxel and piroxicam, where PEG is a carrier agent, and hematoporphyrin, where PEG is covalently attached to form a "stealth shield". Simulating at physiological salt concentration, we found no evidence of any specific interaction between paclitaxel or piroxicam with PEG, but found a strong interaction for the case of hematoporphyrin. This interaction is lipophilic in nature, between the nonpolar (CH(2))(2) groups of the PEG and the porphin ring of the hematoporphyrin. This interaction was found to be strong enough that the PEG aggregated to the hematoporphyrin, independent of whether or not it was covalently bound. Interestingly, when the simulation was repeated in absence of salt we found evidence of this interaction being weakened. This led us to hypothesize a previously unforeseen mechanism: interaction with salt cations cause the PEG to coil around the salt ions, each ion binding to many PEG oxygens, increasing the exposure of the nonpolar ethylene groups, thus increasing the effective hydrophobicity of PEG. The Hydrophobic ethylene groups of the PEG chains adhere strongly to the hydrophobic porphin ring. Experiments involving absorption spectra measurements were conducted, and these results also indicated that presence of salt at physiological level increases the effective attractive interaction between PEG and hematoporphyrin. Taken together, our results demonstrate that while PEG, due to its solubility in both polar and nonpolar solvents, may act as a dissolution aid for paclitaxel and piroxicam, of the three drug molecules studied it will only have a protective role for the case of the hematoporphyrin.