| In recent years, the surface modification of inorganic nanoparticles with natural biomacromolecules or synthetic polymers, can not only improve the dispersity of nanoparticles in aqueous solution as well as their biocompatibility, but also endow the inorganic nanoparticles with multi-functionalities for specific biomedical applications. In this thesis, the magnetic mesoporous silica nanoparticles (MMSNs) were synthesized and modified with horseradish peroxidase (HRP) and carboxylated polyethylene glycol (PEG5000-COOH) by a conventional N-Hydroxysuccinimide (NHS) / 1-Ethyl-3-(3-dimethyllaminopropyl)carbodiimide hydrochloride (EDC) coupling method, respectively. Subsequently, the fluorescent carbon quantum dots (CDs) were introduced to the MMSNs-HRP and MMSNs-PEG, and then the final products of MMSNs-HRP-CDs and MMSNs-PEG-CDs were obtained, respectively. Their morphology, structure and physicochemical properties have been characterized, and their potential applications for cell biology have also been explored.The main works can be summarized as follows:1. Preparation and Characterization of MMSNs-HRP-CDs for Application of Eliminating Intracellular Reactive Oxygen Species (ROS)(1) Firstly, the magnetite (Fe3O4 nanoparticles were synthesized by a hydrothermal method and capped with oleic acid. The as-synthesized Fe3O4 nanoparticles were transferred to water phase, and then encapsulated with silica shells to form MMSNs. Secondly, the MMSNs were surface-modified with carboxyl group, and then convalently bonded to HRP by a NHS/EDC coupling method. Finally, HRP-immobilized magnetic mesoporous silica nanoparticles (MMSNs-HRP) were conjugated with CDs, and then the MMSNs-HRP-CDs were obtained. The morphology, structure, surface area, and magnetic properties of the samples were characterized by using transmission electron microscopy (TEM), infrared spectroscopy (FT-IR), nitrogen (N2) adsorption - desorption isotherms, ultraviolet-visible (UV-Vis) spectra, fluorescence (PL) spectra, and a physical property measurement system (PPMS) etc. The results have shown that MMSNs are spherical particles in an average size of 60 nm, with a BET specific surface area of 963 m2 g-1 as well as an even pore size of 3.0 nm, and exhibit superparamagnetic at room temperature.(2) Chinese hamster ovary cells (CHO) were selected as a model cell line to study the cytotoxicity of MMSNs-HRP and MMSNs-HRP-CDs by using a dual-wavelength tetrazolium salt (MTT) assay. The results have shown that the cytotoxicity of both samples are very low. A certain amount of hydrogen peroxide was added to induce ROS in CHO cells while the cells were treated with MMSNs-HRP for 20 h, and then the intracellular ROS level was analyzed qualitatively with a fluorescence microscopy and quantatively by a flow cytometry as well. All of the results have demonstrated that MMSNs-HRP can effectively eliminate intracellular ROS.(3) The influence of an applied static magnetic field (-0.3 T) on cellular uptake of the MMSNs-HRP-CDs was studied qualitatively by using a fluorescence microscopy. When the cells were co-incubated with 200 μg mL-1 MMSNs-HRP-CDs for 2 h, the internalization number of MMSNs-HRP-CDs in CHO cells in presence of an applied static magnetic field was much more than that in absence of a magnetic field. This result suggests that the influence of an external static magnetic field on the cellular uptake of nanoparticles can not be neglected under certain circumstances.2. Preparation, Characterization and Biocompatibility of MMSNs-PEG-CDs for Application in Cell Imaging(1) Firstly, MMSNs were synthesized with the same method as above-mentioned, and then surface-modified with amino group. Secondly, PEG5000-COOH was grafted to MMSNs-NH2 by a NHS/EDC coupling method. Finally, the CDs were introduced to MMSNs-PEG to achieve the product of MMSNs-PEG-CDs. The morphology, structure, BET surface area and fluorescence property of MMSNs-PEG-CDs were characterized by using TEM, FT-IR, N2 adsorption-desorption isotherms, UV-Vis and PL spectra. The results have demonstrated that the dispersity of MMSNs-PEG-CDs in aqueous solution becomes better than that of the naked MMSNs.(2) The cervical cancer cells (HeLa) were select as a model cell line to study the cytotoxicity of mesoporous silica nanoparticles (MSNs) and solid silica nanoparticles (SSNs) by using a dual-wavelength MTT assay. The apoptosis rates of HeLa cells treated respectively with MSNs and SSNs were analyzed quantatively by using a flow cytometry. The results have shown that the cytotoxicity of both MSNs and SSNs are low, but the apoptosis rate of HeLa cells induced by MSNs was slightly higher than that induced by SSNs. Furthermore, the cytotoxicity of MMSNs, MMSNs-PEG and MMSNs-PEG-CDs were studied by using the same dual-wavelength MTT assay. The results have demonstrated that the biocompatibility of PEGylated-MMSNs can be improved, so that both cytotoxicity of MMSNs-PEG and MMSNs-PEG-CDs become lower than that of MMSNs.(3) Using HeLa cells as a model cell line, the application of MMSNs-PEG-CDs in cell imaging was investigated by means of a confocal laser scanning microscope. The results indicate that MMSNs-PEG-CDs not only have low cytotoxicity, but also can keep strong and stable fluorescence inside cells, which makes them suitable for long-time cell imaging. |
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