The Study Of Copper-Doped Borate Bioactive Glass Scaffolds And Microfibers For Bone And Skin Defects Repair

Part 1 THE STUDY OF COPPER-DOPED BORATE BIOACTIVE GLASS SCAFFOLDS FOR BONE DEFECTS REPAIROBJECTIVE: There is growing interest in the use of synthetic biomaterials to deliver inorganic ions(such as copper ions) that are known to stimulate osteogenesis and angiogenesis in vivo. In the present study, we fabricated copper-doped borosilicate bioactive glass(BG-Cu) scaffolds and investigated the ion release and hydroxyapatite(HA) conversion of bioactive glass scaffolds in simulated body fluid(SBF). This study also explored the effects of varying amounts of copper in the scaffolds on the response of human bone marrow-derived mesenchymal stem cells(BMSCs) in vitro and on blood vessel formation and bone regeneration in rat calvarial defects in vivo.METHODS: Porous scaffolds of a borosilicate bioactive glass doped with 0.5, 1.0 and 3.0 wt.% Cu O were created using a foam replication method. The porosity and compressive strength of the scaffolds were measured. The microstructure of the scaffolds was examined in a field-emission scanning electron microscope(SEM). The as-fabricated scaffolds were ground to form a powder and analyzed using X-ray diffraction(XRD) to check for the presence of any crystalline phases. The concentration of Cu ions released from the bioactive glass into the SBF was determined using inductively coupled plasma atomic emission spectroscopy(ICP-AES). After seeding the BMSCs onto the scaffolds, we used SEM, CCK-8, ALP activity and q RT-PCR methods to study cell compatibility, cell adhesion, proliferation, and differentiation. In vivo bone regenetaion and new blood vessels formation in SD rat calvarial defects were evaluated by sequential fluorescent labeling, Microfil perfusion, Micro-CT and histological examination.RESULTS: There was no observable difference in surface morphology, porosity and mechanical strength among the four groups of as-fabricated scaffolds(BG, BG-0.5Cu, BG-1Cu and BG-3Cu). In SBF, BG-Cu scaffolds released copper ions and converted to HA. In vitro, the Cu in the glass was not toxic to the BMSCs at the concentrations used and BG-Cu scaffolds supported BMSCs adhesion and proliferation. Compared to BG scaffolds, BG-Cu scaffolds significantly promoted the ALP activity and osteogenic(RUNX2, BMP-2 and OPN) / angiogenic(VEGF and b FGF) gene expression of BMSCs. The bone regeneration and blood vessel formation in BG-3Cu implantation group were significantly higher than BG implantation group and the control group.CONCLUTIONS: 1. BG-Cu scaffolds obtained by a foam replication method had the microstructure and mechanical strength similar to human cancellous bone, and showed excellent in vitro bioactivity; In SBF, BG-Cu scaffolds released angiogenic copper ions and converted to HA. 2. Compared with undoped BG scaffolds, BG-Cu scaffolds promoted cell adhesion, proliferation, and osteogenic/angiogenic differentiation of BMSCs. 3. BG-3Cu scaffolds in vivo effectively promoted bone tissue and blood vessel regeneration and repair.Part 2 THE STUDY OF COPPER-DOPED BORATE BIOACTIVE GLASS MICROFIBERS FOR SKIN DEFECTS REPAIROBJECTIVE: Angiogenesis is a prerequisite process for wound tissue repair and regeneration. Cu elements play a key role in the angiogenic process. In this study, we fabricated copper-doped borate bioactive glass(Cu-BG) microfibers and investigated the release of Cu2+ ions from the Cu-doped microfibers and the conversion of the microfibers to HA in SBF. We systematically investigated the effects of the soluble ionic product of the microfibers on the response of cells(HUVECs and fibroblasts) in vitro and the capacity of the microfibers to stimulate angiogenesis and to heal full-thickness skin defects in vivo.METHODS: Microfibers of a borate bioactive glass doped with 0.5, 1.0 and 3.0 wt.% Cu O were created and their physiochemical properties were characterized. The concentration of Cu ions released from the bioactive glass into the SBF was determined using inductively coupled plasma atomic emission spectroscopy(ICP-AES). We prepared soluble ionic product of the microfibers and explored the cytotoxic effects of the soluble ionic product of the different microfibers on the response of HUVECs and fibroblasts using CCK-8 assay. Transwell, tubule formation activity and ELISA assays were used to investigate the soluble ionic product of the different microfibers on migration, capillary-like structure formation and VEGF secretion capacities of HUVECs. q RT-PCR assay was used to investigate the soluble ionic product of the different microfibers on angiogenesis-related gene expression of fibroblasts. The capacity of the microfibers to stimulate angiogenesis and to heal full-thickness skin defects in a rodent model were assessed by gross photos, Microfil perfusion(Micro-CT scan) and histological examination.RESULTS: The as-prepared microfibers had the feel and appearance of cotton wool. There was no observable difference in the surface morphology of the four groups of as-prepared microfibers(BG, 0.5Cu-BG, 1Cu-BG and 3Cu-BG). Each microfiber had a uniform diameter at the microscale level but within each group, the diameters were in the range 0.4 to 1.2 μm, with an average value of 0.85 μm. In SBF, Cu-BG microfibers released copper ions and converted to HA. The soluble ionic product of the Cu-BG microfibers significantly stimulated migration, capillary-like structure formation and VEGF secretion capacities of HUVECs and promoted angiogenesis-related gene expression of fibroblasts. The implantation of microfibers in full-thickness skin defects of SD rats confirmed that 3Cu-BG microfiber could significantly promote angiogenesis and accelerate wound healing.CONCLUTIONS: The as-prepared microfibers had the feel and appearance of cotton wool and showed excellent in vitro bioactivity; In SBF, Cu-BG microfibers released angiogenic copper ions and converted to HA. 2. Compared with the soluble ionic product of undoped BG microfibers, the soluble ionic product of the Cu-BG microfibers stimulated HUVECs migration, tube formation and VEGF secretion of HUVECs and promoted angiogenesis-related gene expression of fibroblasts. 3. 3Cu-BG microfibers in vivo effectively promoted blood vessel formation and accelerated wound healing.