RESUMEN
The development of advanced stimuli-responsive systems for medicine, catalysis, or technology requires compartmentalized reaction spaces with triggered activity. Only very few stimuli-responsive systems preserve the compartment architecture, and none allows a triggered activity in situ. We present here a biomimetic strategy to molecular transmembrane transport by engineering synthetic membranes equipped with channel proteins so that they are stimuli-responsive. Nanoreactors with triggered activity were designed by simultaneously encapsulating an enzyme inside polymer compartments, and inserting protein "gates" in the membrane. The outer membrane protein F (OmpF) porin was chemically modified with a pH-responsive molecular cap to serve as "gate" producing pH-driven molecular flow through the membrane and control the in situ enzymatic activity. This strategy provides complex reaction spaces necessary in "smart" medicine and for biomimetic engineering of artificial cells.
Asunto(s)
Materiales Biomiméticos/química , Membrana Celular/química , Porinas/química , Materiales Biomiméticos/farmacología , Membrana Celular/genética , Concentración de Iones de Hidrógeno , Permeabilidad/efectos de los fármacos , Polímeros/químicaRESUMEN
Membrane proteins have been reconstituted on lipid bilayers with zero mean-curvature (cubic phases or vesicles). Here we show that reconstitution of pore-forming membrane proteins can also occur on highly curved lipidic bilayers of reverse hexagonal mesophases, for which the mean-curvature is significantly different from zero. We further show that the membrane protein provides unique topological interconnectivities between the aqueous nanochannels, significantly enhancing mesophase transport properties.
Asunto(s)
Membrana Dobles de Lípidos/química , Proteínas de la Membrana/química , Modelos Biológicos , Porinas/química , Transporte Biológico , Agua/químicaRESUMEN
Lipidic lyotropic liquid crystals are at the frontline of current research for release of target therapeutic molecules due to their unique structural complexity and the possibility of engineering stimuli-triggered release of both hydrophilic and hydrophobic molecules. One of the most suitable lipidic mesophases for the encapsulation and delivery of drugs is the reversed double diamond bicontinuous cubic phase, in which two distinct and parallel networks of â¼4 nm water channels percolate independently through the lipid bilayers, following a Pn3m space group symmetry. In the unperturbed Pn3m structure, the two sets of channels act as autonomous and non-communicating 3D transport pathways. Here, a novel type of bicontinuous cubic phase is introduced, where the presence of OmpF membrane proteins at the bilayers provides unique topological interconnectivities among the two distinct sets of water channels, enabling molecular active gating among them. By a combination of small-angle X-ray scattering, release and ion conductivity experiments, it is shown that, without altering the Pn3m space group symmetry or the water channel diameter, the newly designed perforated bicontinuous cubic phase attains transport properties well beyond those of the standard mesophase, allowing faster, sustained release of bioactive target molecules. By further exploiting the pH-mediated pore-closing response mechanism of the double amino acid half-ring architecture in the membrane protein, the pores of the perforated mesophase can be opened and closed with a pH trigger, enabling a fine modulation of the transport properties by only moderate changes in pH, which could open unexplored opportunities in the targeted delivery of bioactive compounds.
Asunto(s)
Concentración de Iones de Hidrógeno , Membrana Dobles de Lípidos , Porinas/química , Dispersión de Radiación , Dispersión del Ángulo PequeñoRESUMEN
Various domains present the challenges of responding to stimuli in a specific manner, with the desired sensitivity or functionality, and only when required. Stimuli-responsive systems that are appropriately designed can effectively meet these challenges. Here, we introduce nanoreactors that encapsulate photosensitizer-protein conjugates in polymer vesicles as a source of "on demand" reactive oxygen species. Vesicles made of poly(2-methyloxazoline)-poly(dimethylsiloxane)-poly(2-methyloxazoline) successfully encapsulated the photosensitizer Rose Bengal-bovine serum albumin conjugate (RB-BSA) during a self-assembly process, as demonstrated by UV-Vis spectroscopy. A combination of light scattering and transmission electron microscopy indicated that the nanoreactors are stable over time. They serve a dual role: protecting the photosensitizer in the inner cavity and producing in situ reactive oxygen species (ROS) upon irradiation with appropriate electromagnetic radiation. Illumination with appropriate wavelength light allows us to switch on/off and to control the production of ROS. Because of the oxygen-permeable nature of the polymer membrane of vesicles, ROS escape into the environment around vesicles, as established by electron paramagnetic resonance. The light-sensitive nanoreactor is taken up by HeLa cells in a Trojan horse fashion: it is nontoxic and, when irradiated with the appropriate laser light, produces ROS that induce cell death in a precise area corresponding to the irradiation zone. These nanoreactors can be used in theranostic approaches because they can be detected via the fluorescent photosensitizer signal and simultaneously produce ROS efficiently "on demand".
Asunto(s)
Nanotecnología/instrumentación , Fotobiorreactores , Fármacos Fotosensibilizantes/química , Especies Reactivas de Oxígeno/síntesis química , Albúmina Sérica Bovina/química , Albúmina Sérica Bovina/efectos de la radiación , Diseño de Equipo , Análisis de Falla de Equipo , Luz , Fármacos Fotosensibilizantes/efectos de la radiaciónRESUMEN
There is, today, great need for new systems and strategies for therapeutic applications that will lead to improvements in patient conditions and prognoses, especially in complex diseases such as neurodegenerative diseases and cancer. Recently, polymer nanocarriers have been developed to protect and transport active compounds to pathological sites more efficiently than free compounds in terms of stability, amount required, localization and efficacy. There are two strategies to deliver active compounds: conventional drug delivery systems based on transport and release of active compounds in biological compartments and nanoreactors that transport active compounds and permit them to act in situ, behaving like rudimentary artificial organelles. Here, we present both strategies with their advantages and limitations, and indicate how they can contribute to therapeutic improvement. We focus on presenting the design and development of polymer nanocarriers and nanoreactors as an essential stage of conceiving therapeutic approaches. The properties of the polymer carrier and its behavior under biological conditions dramatically influence the efficacy of the active compound, and thus of the treatment scheme. The key contributions that nanocarriers and nanoreactors could make include protecting active compounds from degradation in biological compartments other than those desired, and concentrating such compounds within their assemblage to allow for multiple deliveries in one single polymer assembly. To efficiently cope with the challenges of complex pathological conditions it is necessary to go one step beyond conventional drug delivery systems by designing and developing nanocarriers that mimic organelles, by combining various active molecules in a single carrier, and even by combining therapeutic agents along with agents for detection, as in a theragnostic approach.