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Study on Optimization and Biocompatibility Evaluation of Multi-component Nano-hydroxyapatite-based Biomaterials

Author: YaoYuLi
Tutor: SunHong
School: Hebei Union University
Course: Pathology and Pathophysiology
Keywords: Hydroxyapatite Composite Scaffolds MC3T3-E1 cells Cytocompatibility Histocompatibility
CLC: R318.08
Type: Master's thesis
Year: 2011
Downloads: 21
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Abstract

Objective To prepare five different proportions of the new multi-component two-dimensional nano-hydroxyapatite-based composites, the MC3T3-E1 cells with different proportions of these five groups of materials outside the Community culture by inverted phase contrast microscope, scanning electron microscopy (SEM), tetrazolium salt (MTT) method, F-DA staining preferably a cell adhesion, proliferation and growth of a group of the best materials, material composition in this group on the basis of three-dimensional composite scaffolds constructed by cell compatibility and tissue compatibility evaluation as a cell scaffold for bone tissue engineering feasibility of the design and preparation of biological material to provide a theoretical basis for the clinical application of biological materials to broaden provide some technical support. Method 1 The preferred dimensional composite: MC3T3-E1 cells were cultured in vitro, taking the well growing cells were passaged experiments. Experimental groups: one (material 1 cells); 2 (material 2 cells); 3 (material 3 cells); 4 (material 4 cells)); 5 (material 5 cells); 6 control group (cell culture medium). By inverted phase contrast microscope, scanning electron microscopy (SEM), tetrazolium salt (MTT) method, F-DA fluorescence staining cells on the material adhesion, proliferation and growth. (2) Three-dimensional composite scaffolds Cytocompatibility experimental groups: material group (cell culture materials) and control group (cell culture medium). By inverted phase contrast microscope, HE staining, tetrazolium salt (MTT), scanning electron microscopy (SEM), F-DA and Hoechst33258 fluorescence staining cells in three-dimensional composite scaffold adhesion, proliferation and growth . 3-D scaffolds histocompatibility complex experimental groups: the three-dimensional composite scaffolds SD rats were implanted subcutaneously material group (material SD rats) control group (SD rats). By HE staining dimensional composite scaffolds in SD rats inflammation and vascularization within the material situation. Results 1. MC3T3-E1 cells were cultured in vitro inverted phase contrast microscope cell culture flasks: just seeded in culture flasks passaged cells are spherical, suspension, 2h ~ 3h adherent cells began, 24h fusiform cells, polygonal or fusiform, projections extending protrusions are connected with neighboring cells, 48h contact with cells gradually into a sheet, 3 ~ 5d cells grow into a dense monolayer, ill-defined. 2-dimensional group 1) inverted phase contrast microscope observation: opaque material, can not see the growth of cells on the material circumstances, only by observing the color of the medium material is contaminated. 2) Scanning electron microscopy revealed cell morphology changes over time, develop 14d, the third group of cell surface very full. 3) MTT results shows a third group of materials with the best adhesive properties of the cell material in the third group was significantly higher than the growth rate of the material of the fifth group of the proliferation rate (P lt; 0.05), and is similar to the control group (P gt; 0.05). 4) F-DA fluorescence staining the surface of the third group of materials best state of cell growth, cell aggregation growth, with formation of nodules trend. 3-D materials Cytocompatibility 1) HE staining seen MC3T3-E1 cell morphology clear, fusiform, spindle or polygonal, abundant cytoplasm, nucleus large and clear, round or oval. 2) SME cell morphology seen very full, fusiform or polygonal, and a plurality of filaments extending projections, forming a anchor-like structure, tightly adhered to the material, and has a good stretch condition. 3) F-DA staining of cell growth seen well clear fusiform or polygonal shape, some areas of the cell aggregation growth, there is a trend to form nodules. 4) Hoechst33258 staining, the cells grow not only on the material surface, and the growth in the hole, a multilayer growth. 5) MTT assay materials and control groups showed two cell activity, 7d, 14d and 21d of the absorbance value (OD value) between the two groups was not statistically significant (P gt; 0.05). 4-D material group histocompatibility: SD-rats subcutaneous HE staining seen SD-rats subcutaneously implanted material within the formation of blood vessels and inflammation lighter. Conclusion: In a preferred prepared on the basis of multi-component composite nano-hydroxyapatite-based three-dimensional scaffolds have good biocompatibility in vitro can promote MC3T3-E1 cell adhesion and proliferation, inflammation implanted light, and the formation of blood vessels, is a promising material for bone tissue engineering.

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