| Partâ… Preparation of carboxylated graphene oxide/true bone ceramics and a study of its physic-chemical propertyObjective To investigate the preparing method of carboxylated graphene oxide/true bone ceramics, and evaluate its physic-chemical property.Methods The graphene oxide (GO) was prepared by modified Hummers method, and GO was carboxylated to become the carboxylated graphene oxide (CGO). The CGO was identified by FTIR and XPS. The true bone ceramics cube was immersed into0.5mg/ml, lmg/ml and2mg/ml CGO suspending liquid, and the ultrasonic wave was used to combine the CGO on the surface of TBC by different times. Five different kinds of CGO-TBC were prepared. The surface morphology and CGO density were observed by three-dimensional video microscope and scanning electron microscope (SEM), and the best type of CGO-TBC was chosen to study its rate of porosity, pore size, compress and bend intensity. Results Identified by FTIR and XPS, the GO was demonstrated to be carboxylated successfully. The lmg/ml-6h type of CGO-TBC was chosen for the best type by three-dimensional video microscope and SEM. CGO-TBC had the similar pore size and rate of porosity to TBC, and had the higher compress and bend intensity than TBC (P<0.05).Conclusion The1mg/ml-6h type of CGO-TBC showed the best surface morphology and CGO density, remained the similar pore structure, and had the better biomechanics. Part â…¡The study of biocompatibility of CGO-TBC and its controlled release loaded with BMP2-related peptide pBMP2in vitroObjective To evaluate the biocompatibility of CGO-TBC, and evaluate the release kinetics of pBMP2/CGO-TBC in vitro.Methods The leaching liquor of CGO-TBC was prepared. Leaching liquor was used in the experiments of acute toxicity testing, micronucleus test, local skin irritation test, pyrogen test and hemolysis test, and the material was used in the implanting muscle experiment. The MC3T3-E1cells were cultured on the surface of CGO-TBC, and observed under the fluorescent microscope by Calcein/PI staining. The release kinetics of pBMP2/TBC and pBMP2/CGO-TBC was evaluated in vitro.Results The acute toxicity testing, micronucleus test, local skin irritation test and hemolysis test were negative. The pyrogen test is suitable for the request. Fibrous capsule was formed around the material which was implanted in vivo. The survival and shape of MC3T3-E1on the surface of CGO-TBC were normal. The release of pBMP2in CGO-TBC was slower than that in TBC (P<0.05).Conclusion The biocompatibility of CGO-TBC is good, and CGO-TBC had slower release of pBMP2in vitro. It is suitable for CGO-TBC to be a bone repair scaffold and drug carrier. Part â…¢Shape and adhesion of MC3T3-E1cells on the surface of CGO-TBC, and proliferation and osteogenic differentiation of cells on the surface of CGO-TBC loaded with pBMP2Objective To evaluate the effect of CGO-TBC on the shape and adhesion of MC3T3-E1, and to evaluate the effect of pBMP2/CGO-TBC on the proliferation and osteogenic differentiation of cells.Methods The MC3T3-E1cells were cultured on the surface of TBC and CGO-TBC, and the shape and disposition of cells were observed by confocal laser scanning microscope and SEM. The adhesion rate was assessed by precipitation method. The proliferative ability was measured by MTT assay, and the ALP activity was measured by ALP kit.Results The confocal laser scanning microscope and SEM showed the cells on CGO-TBC had a better shape and disposition. The adhesion rate of CGO-TBC group is higher than the TBC group (P<0.05). The results of MTT assay and ALP activity showed that the pBMP2had the similar osteoinductive to BMP2, and CGO-TBC could also promote the proliferation and osteogenic differentiation of MC3T3-E1cells.Conclusion CGO-TBC could be beneficial for the shape and adhesion of MC3T3-E1, and pBMP2/CGO-TBC could promote the proliferation and osteogenic differentiation of MC3T3-E1. The pBMP2could perform the similar osteoinductive to BMP2. Part â…£Repair of rat cranium bone defects using CGO-TBC loaded with pBMP2Objective To evaluate the ability and efficacy of repair of rate cranium bone critical size defects (CSD) using CGO-TBC loaded with pBMP2.Methods24SD rates were randomly divided into six groups:TBC group, CGO-TBC group, pBMP2/TBC group, pBMP2/CGO-TBC group, rhBMP2/TBC group, and rhBMP2/CGO-TBC group. A5mm diameter CSD was made on the cranium bone of rats. Radiographic examination (X-ray and3D-CT) and histological examination (HE staining) were taken to evaluate the bone repair capability of six groups after12weeks.Results The results of radiographic and histological examination suggested that no obvious new bone was found in TBC group. A little new bone was formed around the edge of material in the CGO group. The results of radiographic examination suggested that the bone-material interfaces were unclear for pBMP2/TBC group and pBMP2/CGO-TBC group, and histological examination showed that there were a little woven bone around the edge of pBMP2/TBC, and there were much woven bone around the edge and a little osteoid and woven bone in the center of pBMP2/CGO-TBC. The results of radiographic examination suggested that the bone-material interfaces were also unclear for rhBMP2/TBC group and rhBMP2/CGO-TBC group, and histological examination showed that there were new bone formation in the edge and center of material of rhBMP2/TBC group, and there were more new lamellar bone in the edge and center of material of rhBMP2/CGO-TBC group. The new bone area of TBC group and CGO-TBC group were less than other groups (P<0.05). The rhBMP2/CGO-TBC group had the largest new bone area than other groups (P<0.05). The differences between the pBMP2/TBC group, pBMP2/CGO-TBC group and rhBMP2/TBC group were not significant (P>0.05).Conclusion The CGO-TBC loaded with pBMP2could be a good bone repair complex material, have good osteoinductive activity and deserve further pre-clinical research. |
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