Biomedical Applications Of Laponite Or Hydroxyapatite-doped Electrospun Nanofibers

The major advantage of electrospun nanofibers is that the nanofibers posses high specific surface area, high porosity, and three-dimensional (3D) porous nanostructures which can mimic the natural extracellular matrix (ECM) very well. Compared with single-component nanofibers, composite or hybrid nanofibers are more promising due to the unique properties possessed by both host and guest materials. Poly(lactide-co-glycolide acid)(PLGA) is a copolymer widely used in a host of Food and Drug Administration (FDA) approved therapeutic devices oweing to its excellent biocompatibility and biodegradability. γ-poly glutamic acid (y-PGA) is a naturally occurring anionic biopolymer secreted by Bacillus subtilis and is promising for biomedical applications due to its excellent biocompatibility and biodegradability. In this thesis, we used y-PGA and PLGA as base materials to explore the preparation and applications of laponite (LAP) or hydroxyapatite (n-HA)-doped electrospun nanofibers in the fields of drug delivery systems and tissue engineering.With a unique "sandwich-like" structure, the interlayer space of LAP can be used for effective drug encapsulation with high retention capacity. In the second part of this thesis, LAP was used to encapsulate an anticancer drug of doxorubicine (DOX). LAP/DOX composites can be obtained by simply stirring the LAP and DOX mixed solution. The in vitro drug release and anticancer activity of the LAP/DOX were also studied. We show that an exceptionally high loading efficiency of98.3±0.77%can be obtained when the concentration of LAP and DOX is3mg/mL and1mg/mL, respectively. XRD results show that DOX can be encapsulated into the interspace of LAP via the interaction with (001) crystal face. In vitro drug release study demonstrates that much more DOX can be released under acidic pH condition (pH=5.4) than under physiological pH condition. Cell viability assay results reveal that LAP/DOX nanodisks display a significantly higher therapeutic efficacy in inhibiting the growth of a model cancer cell line (human epithelial carcinoma cells, KB cells) than free DOX drug at the same DOX concentrations.The third chapter of the thesis reports the fabrication of uniform electrospun poly(lactic-co-glycolic acid)(PLGA) nanofibers incorporated with LAP nanodisks and the influence of the doped LAP on the surface hydrophilicity, mechanical durability and cytocompatibility was studied. We show that the incorporation of LAP nanodisks can decrease the fiber diameter, porosity and improve the mechanical durability, the surface hydrophilicity and protein adsorption capacity of the nanofibers. The formed PLGA/LAP composite nanofibers possess good compatibility and hemocompatibility. Results form other groups have shown that silicate clay materials have potentials to induce the osteogenic differentiation of human mesenchymal stem cells (hMSC). We used the composite PLGA/LAP nanofibers as scaffolds for osteogenic differentiation of hMSC. The metabolic activity of hMSC cultured onto LAP-doped PLGA nanofibers was analyzed using the resazurin reduction assay. The osteogenic differentiation was evaluated by analyzing the cellular alkaline phosphatase activity and measuring the cellular osteocalcin secretion, as well as by histochemical assay. We show that PLGA/LAP composite nanofibers are able to support the proliferation of hMSC in osteogenic and growth medium. Most strikingly, the doped LAP within the PLGA nanofibers is able to induce the osteoblast differentiation of hMSC in growth medium without any inducing factors while the pure PLGA nanofibers have no such an inducing effect. The fabricated smooth and uniform organic/inorganic LAP-doped PLGA hybrid nanofibers may find many applications in the field of tissue engineering.Based on findings of chapter two and three, in the fourth chapter, LAP nanodisks were first used to encapsulate a model drug of amoxicillin (AMX). Then, the formed LAP/AMX nanodisks were incorporated within PLGA nanofibers via electrospinning to form PLGA/LAP/AMX nanofibers. In vitro drug release behavior, the antimicrobial activity and the cytocompatibility of the composite PLGA/LAP/AMX nanofibers were extensively studied. We show that the AMX-loaded LAP nanodisks with an optimized loading efficiency of9.76±0.57%were able to be incorporated within PLGA nanofibers.The release profile of AMX follows a biphasic mode characterized by a firstly fast and then sustained manner. PLGA/LAP/AMX nanofibers display effective antibacterial activity toward a model bacterium of Staphylococcus aureus and non-compromised cytocompatibility in comparison with pure PLGA nanofibers, therefore may be used in wound dressing applications.The fifth chapter of the thesis discussed the influence of various parameters on the electrospinning as well as the formed nanofibers. By altering the concentration of trifluoroacetic acid under the above optimized parameters, y-PGA nanofibers with controlled morphology can be formed. The formed γ-PGA nanofibers can be rendered with water stability by crosslinking with cystamine as well as a novel reagent of poly(amidoamine) dendrimer of generation two via the EDC coupling. Furthermore, we confirmed the excellent biocompatibility of this water-stable y-PGA nanofiber. The formed y-PGA nanofibers may find extensive applications in tissue engineering.The last part of the thesis mainly concerns the preparation of n-HA-based composite nanofibers through chemical (the aminopropyltriethoxysilane induced surface acetylation or carboxylation modification of n-HA) or physical methods (electrospinning y-PGA/n-HA composite nanofibers from simply mixed solution). Our results reveal that the slight cytotoxicity of the amine-functionalized n-HA-APTS can be eliminated by post-functionalization of the APTS amines with acetyl or carboxyl group. Hemocompatibility assessment demonstrates that the negligible hemolytic activity of the pristine n-HA particles does not appreciably change after the APTS-mediated surface functionalization. The doped n-HA has no obvious influence on the morphology of y-PGA nanofibers and the chemical structure of n-HA can still remain its integrity even if trifluoroacetic acid was used as the solvent. The findings of this thesis will provide novel ideas to design organic or inorganic/organic composite nanofibers for biomedical applications in the fields of tissue engineering and drug delivery systems.