RESUMO
Supramolecular nanogels are an emerging class of polymer nanocarriers for intracellular delivery, due to their straightforward preparation, biocompatibility, and capability to spontaneously encapsulate biologically active components such as DNA. A completely biodegradable three-component cationic supramolecular nanogel was designed exploiting the multivalent host-guest interaction of cyclodextrin and adamantane attached to a polypeptide backbone. While cyclodextrin was conjugated to linear poly-L-lysine, adamantane was grafted to linear as well as star shaped poly-L-lysine. Size control of nanogels was obtained with the increase in the length of the host and guest polymer. Moreover, smaller nanogels were obtained using the star shaped polymers because of the compact nature of star polymers compared to linear polymers. Nanogels were loaded with anionic model cargoes, pyranine and carboxyfluorescein, and their enzyme responsive release was studied using protease trypsin. Confocal microscopy revealed successful transfection of mammalian HeLa cells and intracellular release of pyranine and plasmid DNA, as quantified using a luciferase assay, showing that supramolecular polypeptide nanogels have significant potential in gene therapy applications.
Assuntos
Peptídeos , Polímeros , Animais , DNA , Células HeLa , Humanos , NanogéisRESUMO
In previous studies it was shown that polysorbate 80(PS80)-coated poly(n-butylcyano-acrylate) nanoparticles (PBCA-NP) are able to cross the blood-brain barrier (BBB) in vitro and in vivo. In order to explore and extend the potential applications of PBCA-NP as drug carriers, it is important to ascertain their effect on the BBB. The objective of the present study was to determine the effect of PS80-coated PBCA-NP on the BBB integrity of a porcine in vitro model. This has been investigated by monitoring the development of the transendothelial electrical resistance (TEER) after the addition of PBCA-NP employing impedance spectroscopy. Additionally, the integrity of the BBB in vitro was verified by measuring the passage of the reference substances (14)C-sucrose and FITC-BSA after addition of PBCA-NP. In this study we will show that the application of PS80-coated PBCA-NP leads to a reversible disruption of the barrier after 4h. The observed disruption of the barrier could also be confirmed by (14)C-sucrose and FITC-BSA permeability studies. Comparing the TEER and permeability studies the lowest resistances and maximal values for permeabilities were both observed after 4h. These results indicate that PS80-coated PBCA-NP might be suitable for the use as drug carriers. The reversible disruption also offers the possibility to use these particles as specific opener of the BBB. Instead of incorporating the therapeutic agents into the NP, the drugs may cross the BBB after being applied simultaneously with the PBCA-NP.
Assuntos
Barreira Hematoencefálica/metabolismo , Portadores de Fármacos/química , Portadores de Fármacos/metabolismo , Embucrilato/química , Nanopartículas/química , Animais , Transporte Biológico , Radioisótopos de Carbono/metabolismo , Fluoresceína-5-Isotiocianato/metabolismo , Polissorbatos/química , Albumina Sérica/metabolismo , Sacarose/metabolismo , SuínosRESUMO
A brain delivery probe was prepared by covalently conjugating lactoferrin (Lf) to a poly(ethylene glycol) (PEG)-coated Fe(3)O(4) nanoparticle in order to facilitate the transport of the nanoparticles across the blood-brain barrier (BBB) by receptor-mediated transcytosis via the Lf receptor present on cerebral endothelial cells. The efficacy of the Fe(3)O(4)-Lf conjugate to cross the BBB was evaluated in vitro using a cell culture model for the blood-brain barrier as well as in vivo in SD rats. For an in vitro experiment, a well-established porcine BBB model was used based on the primary culture of cerebral capillary endothelial cells grown on filter supports, thus allowing one to follow the transfer of nanoparticles from the apical (blood) to the basolateral (brain) side. For in vivo experiments, SD rats were used as animal model to detect the passage of the nanoparticles through the BBB by MRI techniques. The results of both in vitro and in vivo experiments revealed that the Fe(3)O(4)-Lf probe exhibited an enhanced ability to cross the BBB in comparison to the PEG-coated Fe(3)O(4) nanoparticles and further suggested that the Lf-receptor-mediated transcytosis was an effective measure for delivering the nanoparticles across the BBB.