Preparation Of Light Responsive Surface And It Mediated Gene Delivery Properties

The physical and chemical properties of the biomaterial surface are significant factors to the interactions between materials and biomolecules or cells.Therefore,it can meet the needs of different processes of life activities.Once constructed,the static characteristics of material surface will remain changeless states during a long period.However,the state of biomolecules and cellular activities are continuous and dynamic processes,so a single surface state has a limited effect on the regulation of biomolecules and cells.In recent years,smart materials have become increasingly prominent in the field of biomedical research.It is possible to precisely regulate the behaviors of biomolecules and cells in a spatio-temporal manner,by constructing an external field responsive material surface and using an external field to regulate the surface properties.The release and cellular uptake of gene/vector complexes are key processes in surface-mediated gene delivery.Precise regulation of these processes will provide solutions to the current shortcomings of surface-mediated gene delivery.Since the complexes and cells are directly interact with material surface,it is feasible to precisely and controllably regulate surface-mediated gene delivery processes by regulating material surface properties using external filed.At the same time,it has great guiding significance for further understanding the interface interaction of material-complexes and material-cells.In this thesis,light responsive TiO₂ nano-structured,silicon with a pn junction,collagen/AuNPs and graphene/Si?Gr/Si?were used as material systems to investigate the light-induced regulation of surface properties,complexes release and cellular uptake.Moreover,the mechanisms of light promoted complexes releases and cellular uptake,as well as the surface-mediated gene delivery on theses surfaces were studied.Based on these works,the functional genes with osteogenic regulation ability were delivered and the performances of osteogenic differentiation were evaluated.The main results are as following:1.Cell detachment and gene delivery on ultraviolet light responsive film based on TiO₂The ZnO/TiO₂ composite nanodot films were prepared by adding zinc acetate into the precursor sol of TiO₂ nanodot,followed by spin-coating and calcination treatment.Comparing to TiO₂ nanodot film,the cell adhesion,proliferation and UV induced cell detachment of the composite nanodot film were improved.Because the integrity of gene was destroyed by UV light,the gene delivery efficiency on ZnO/TiO₂ composite nanodot film was decreased under UV365 illumination.2.Study on the regulation of surface-mediated gene delivery based on photothermal surface of TR/Col/AuNPs composite coatingThe TR/Col/AuNPs composite coating was prepared by hydrothermal and sol-gel method.The composite coating possesses good cell compatibility,photothermal and complex loading capacity.The temperature of the coating surface increased under visible light illumination.Meanwhile,Fourier transform infrared spectroscopy showed that the conformation of collagen changed from order states to disorder states.Such a conformation transitions results in a rapid release of complexes.Moreover,the release of complexes could be precisely controlled in a spatio-temporal manner.In addition,the thermal energy generated by visible light illumination stimulates the rearrangement of the cytoskeleton,which enhances the ability of cellular uptake for LF/GFP.Surface-mediated gene delivery experiment demonstrates that the synergistic effects of release and cellular uptake of complexes promoted surface-mediated gen delivery efficiency.This light-regulated gene delivery method not only works on single cells,but also promotes gene delivery in cell sheets,reflecting a tremendous application prospect in vivo.3.Study on the regulation of surface-mediated gene delivery based on photoelectronic surface of p⁺n/SiThe p⁺n/Si substrates were prepared by alkaline etching and thermal diffusion.The anisotropic etching of single crystal Si in an alkaline solution results in the formation of a pyramidal morphology on silicon surface.Such structure is helpful in reducing front surface reflectance and light trapping.The pn junction is formed by thermal diffusion,which gives the substrate an excellent photoelectric effect.The surface-mediated gene delivery efficiency increased with visible light illumination,and was dependent on light intensity and illumination time.Such light controlled surface-mediated gene delivery method is applicable to a variety of tissue cells and genes.The mechanism of light controlled surface-mediated gene delivery was found as follows:on one hand,photo-generated holes diffuse into p/Si region and accumulate on the surface due to the built-in electric field,increasing the surface potential.Such change causes the rapid release of complexes due to the electrostatic repulsion between the surface and the positively charged complexes.On the other hand,the photovoltage of the substrate stimulates the cytoskeletal rearrangement into an elongated morphology,thereby enhancing the cellular uptake ability.The two processes synergistically regulate the surface-mediated gene delivery based on p⁺n/Si substrate.4.Study on the regulation of surface-mediated gene delivery based on photoelectric and photothermal synergistic surface of Gr/SiGr/Si substrates are prepared by transferring Gr to n/Si.The Raman spectrum displayed that the organic support layer?PLA?on Gr was completely removed,and Gr was monolayer with few defects.The Schottky junction formed between the Gr and Si interface gave the substrate excellent photoelectric performance.The Gr/Si substrate showed good photothermal performance due to the adsorption of near-infrared light?NIR?and photothermal conversion capability of Gr.In addition,the Gr/Si had good biocompatibility and complexes loading ability.Due to the presence of the Schottky junction,the photo-generated holes diffused to Gr and accumulated on surface under visible light illumination,which produced electrostatic repulsion to the positively charged complexes,resulting in the rapid release of that.Due to the photothermal effect of the Gr/Si substrate,the cell membrane permeability was enhanced under NIR illumination,causing a promoted cellular uptake to complexes.The results of surface-mediated gene delivery on Gr/Si demonstrated that the gene delivery efficiency was optimal under the stepwise illumination of visible light and NIR,compared with the single illumination of visible light or NIR.Such results indicated that precise and stepwise controls of the release and cellular uptake of complexes were significant to regulate surface-mediated gene delivery.In this thesis,different light-responsive material surfaces were constructed and the effective regulations of complexes release and cellular uptake were achieved assisted with light.The mechanisms of light enhanced complexes release and cellular uptake were elucidated,and subsequently used in surface-mediated gene delivery.Moreover,their further application in bone tissue repair was explored.These have great guiding significance for stimuli-regulated surface-mediated gene delivery and the construction of a novel tissue repair system.