Navegando por Palavras-chave "Magnetofection"

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    Desenvolvimento de nanopartículas magnéticas de sílica mesoporosa como vetores não virais em magnetofecção
    (Universidade Federal de São Paulo (UNIFESP), 2020-05-15) Francisco, Giorgio Fernando De Avelar [UNIFESP]; Bizeto, Marcos Augusto [UNIFESP]; Universidade Federal de São Paulo
    Gene therapy is based on introduction of exogenous genetic material into the cells aiming the treatment of genetic diseases from the inactivation/substitutions of defective genes or by the introduction of genes which can express therapeutic proteins. Incompetent recombination virus based vectors are the most utilized at this transfer, but instead of the high efficiency, there are many risks associated to the possibility of the expression of remaining viral genes, promoting intense immunologic response and generation of competent replication virus by recombination which can promote oncogenesis. Thence, the development of non-viral vectors, mainly based on inorganic nanoparticles has become an interesting technological and scientific topic. The transfection process is complex, and many cell barriers must be transposed by the vector. The cell entry, generally, occurs by endocytosis, which requires ideals charge and size. After passing through cell membrane, the vector is imprisoned in cell compartments which interior has acidic medium and rich in nucleases that degrade the genetic material. Nucleic acids released at cytosol in this stage can be transferred to the cell nucleus, passing through the nucleus membrane where it will be transcript. In this work, capacity (or efficiency) of magnetofection was evaluated in order to improve the rate of cell uptake by endocytosis of vectors based in mesoporous silica nanoparticles. The magnetofection process consists in cell transfection with magnetic nanoparticles by applying an external magnetic field. Initially, it was synthesized different types of mesoporous silica nanoparticles combined with superparamagnetic nucleus of magnetite (Fe3O4), which were chemically modified with propyldiethylenotriamine to enable the conjugation with molecules of plasmid DNA responsible to promote the expression of green fluorescent protein (GFP) in HeLa cells. Among the as-prepared magnetic vectors, just one of them presented propitious characteristics to transfection study. This vector was prepared by hydrolysis of silica inorganic precursor using sodium hydroxide as catalyst around magnetic nanoparticles in the presence of micelle aggregates of surfactant hexadecyltrimethylammonium bromide, to shape the formation of silica wall pores. The as-prepared nanomaterial infrared spectrum presented the main bands of the vibrational modes of magnetite, silica and propyldiethylenotriamine group. The N2 physissortion isotherms showed an improvement on the surface area of 14 m2 /g to 254 m2 /g after the covering, with an isotherm of type IV, which is characteristic of mesoporous material. The X ray diffractometry identified the presence of magnetite as a crystal at face centered cubic form and the amorphous silica. The transmission electron microscopy showed an agglomerate of spherical particles with a well defined core-shell structure. The colloidal stability of the propyldiethylenotriamine modified nanoparticles was evaluated in dispersion made in sodium chloride solution by zeta potential (26 ± 1mV) and by determination of hydrodynamic size (516 ± 74nm) by dynamic light scattering. Finally, the magnetofection of HeLa cells with the as-prepared magnetic nanoparticles showed a higher transfection rate (5.4 ± 3.2%) when compared to the control groups presenting a good perspective of its use in gene therapy.