纳米钛酸钙涂层的钛合金的制备及体外生物相容性的评价
发布时间：2018-11-19 21:13
【摘要】：目的：通过体外细胞实验,对比纯钛、纳米钛酸钙、羟基磷灰石三者的生物相容性,为钛酸钙涂层钛合金材料作为新型体内植入物的可行性提供一定的实验依据。 方法：1.MC3T3-E1细胞的传代培养,待细胞培养稳定后,将实验材料与细胞一起复合培养。 2.将实验材料分成三组,A组(无涂层钛板TI);B组(羟基磷灰石涂层钛板HA-TI):C组(钛酸钙涂层钛板CTO-TI)各60片灭菌备用。 3.采用成骨细胞Calcein-AM+PI+DAPI的荧光染色、MTT法成骨细胞活性测定,来评价成骨细胞粘附与增殖。 4.采用成骨细胞肌动蛋白(actin)+β微管蛋白+DAPI的免疫荧光染色、成骨细胞碱性磷酸酶(ALP)活性测定,来评价细胞的分化与成熟。 5.采用成骨细胞Ⅰ型胶原蛋白mRNA表达、材料表面细胞生长的扫描电镜观察,来评价成骨细胞的成骨钙化能力。 6.数据用x±s表示,采用SPSS17.0统计分析软件对实验数据进行分析,各组间比较采用单因素方差分析(One-way ANOVE),多个样本均数间的多重比较采用LSD-t检验。P0.05有统计学差异。 结果：1.成骨细胞Calcein-AM+PI+DAPI的荧光染色、MTT法成骨细胞活性测定提示：随着时间的延长,B、C组材料比A组更加有利于细胞的增值,有统计学意义(P0.05)；B组与C组材料比较,两者对细胞毒性相差不大,没有统计学意义(P0.05)。 2.采用成骨细胞肌动蛋白(actin)+β微管蛋白+DAPI的免疫荧光、成骨细胞碱性磷酸酶(ALP)活性测定提示：随着时间的延长,B、C组材料比A组更加有利于细胞的分化与成熟(P0.05)；B组与C组材料两者之间相差不大(P0.05)。 3.成骨细胞Ⅰ型胶原蛋白mRNA表达、材料表面细胞生长的扫描电镜观察提示：B、C组材料比A组更加具有成骨矿化能力,促使骨细胞成骨钙化,有统计学意义(P0.05)；B组与C组材料比较,两者成骨能力差别不大,没有统计学意义(P0.05)。 结论：钛合金表面纳米级钛酸钙涂层后的材料无明显细胞毒副作用,具有良好的生物相容性,在促进成骨细胞增殖、成骨等方面与羟基磷灰石相近,为其作为生物材料植入体内提供了一定的实验依据。
[Abstract]:Objective: to compare the biocompatibility of pure titanium, nanometer calcium titanate and hydroxyapatite in vitro, and to provide some experimental basis for the feasibility of calcium titanate coated titanium alloy as a new type of implant in vivo. Methods: 1.MC3T3-E1 cells were subcultured and co-cultured with cells after stable cell culture. 2. The experimental materials were divided into three groups: group A (uncoated titanium plate TI); B group) (hydroxyapatite coated titanium plate HA-TI): C group (calcium titanate coated titanium plate CTO-TI) 60 slices each). 3. Osteoblast adhesion and proliferation were evaluated by fluorescence staining of osteoblast Calcein-AM PI DAPI and assay of osteoblast activity by MTT method. 4. The differentiation and maturation of osteoblasts were evaluated by immunofluorescence staining of actin (actin) 尾 tubulin DAPI and (ALP) activity of alkaline phosphatase in osteoblasts. 5. The expression of type I collagen mRNA in osteoblasts and the growth of osteoblasts were observed by scanning electron microscope to evaluate the osteogenic calcification ability of osteoblasts. 6. The data were expressed by x 卤s, and the experimental data were analyzed by SPSS17.0 statistical analysis software. The data were compared by single factor variance analysis (One-way ANOVE),). LSD-t test was used to compare the multiple mean of multiple samples. P05 had statistical difference. Results: 1. The fluorescence staining of osteoblast Calcein-AM PI DAPI and the assay of osteoblast activity by MTT method showed that: with the extension of time, the material of group C was more favorable than group A, which had statistical significance (P0.05). Group B and group C materials, the cytotoxicity between the two groups, there was no statistical significance (P0.05). 2. Using immunofluorescence of osteoblast actin (actin) 尾 tubulin DAPI, the activity of alkaline phosphatase (ALP) in osteoblasts was determined. Group C was more conducive to cell differentiation and maturation than group A (P0.05). There was no significant difference between group B and group C (P0.05). 3. The expression of type I collagen mRNA in osteoblasts and the growth of osteoblasts on the surface of osteoblasts were observed by scanning electron microscope (SEM). The results showed that the material in group B _ (C) had more osteogenic mineralization ability than group A, and promoted osteocalcification of osteocytes (P0.05). Group B and C group of materials, the osteogenic ability of the difference between the two groups, there was no statistical significance (P0.05). Conclusion: the materials coated with nanometer calcium titanate on titanium alloy have no obvious cytotoxic side effects and have good biocompatibility. They are similar to hydroxyapatite in promoting osteoblast proliferation and osteogenesis. It provides a certain experimental basis for its implantation as a biomaterial in vivo.
【学位授予单位】：中南大学
【学位级别】：硕士
【学位授予年份】：2012
【分类号】：R318.08
本文编号：2343440
[Abstract]:Objective: to compare the biocompatibility of pure titanium, nanometer calcium titanate and hydroxyapatite in vitro, and to provide some experimental basis for the feasibility of calcium titanate coated titanium alloy as a new type of implant in vivo. Methods: 1.MC3T3-E1 cells were subcultured and co-cultured with cells after stable cell culture. 2. The experimental materials were divided into three groups: group A (uncoated titanium plate TI); B group) (hydroxyapatite coated titanium plate HA-TI): C group (calcium titanate coated titanium plate CTO-TI) 60 slices each). 3. Osteoblast adhesion and proliferation were evaluated by fluorescence staining of osteoblast Calcein-AM PI DAPI and assay of osteoblast activity by MTT method. 4. The differentiation and maturation of osteoblasts were evaluated by immunofluorescence staining of actin (actin) 尾 tubulin DAPI and (ALP) activity of alkaline phosphatase in osteoblasts. 5. The expression of type I collagen mRNA in osteoblasts and the growth of osteoblasts were observed by scanning electron microscope to evaluate the osteogenic calcification ability of osteoblasts. 6. The data were expressed by x 卤s, and the experimental data were analyzed by SPSS17.0 statistical analysis software. The data were compared by single factor variance analysis (One-way ANOVE),). LSD-t test was used to compare the multiple mean of multiple samples. P05 had statistical difference. Results: 1. The fluorescence staining of osteoblast Calcein-AM PI DAPI and the assay of osteoblast activity by MTT method showed that: with the extension of time, the material of group C was more favorable than group A, which had statistical significance (P0.05). Group B and group C materials, the cytotoxicity between the two groups, there was no statistical significance (P0.05). 2. Using immunofluorescence of osteoblast actin (actin) 尾 tubulin DAPI, the activity of alkaline phosphatase (ALP) in osteoblasts was determined. Group C was more conducive to cell differentiation and maturation than group A (P0.05). There was no significant difference between group B and group C (P0.05). 3. The expression of type I collagen mRNA in osteoblasts and the growth of osteoblasts on the surface of osteoblasts were observed by scanning electron microscope (SEM). The results showed that the material in group B _ (C) had more osteogenic mineralization ability than group A, and promoted osteocalcification of osteocytes (P0.05). Group B and C group of materials, the osteogenic ability of the difference between the two groups, there was no statistical significance (P0.05). Conclusion: the materials coated with nanometer calcium titanate on titanium alloy have no obvious cytotoxic side effects and have good biocompatibility. They are similar to hydroxyapatite in promoting osteoblast proliferation and osteogenesis. It provides a certain experimental basis for its implantation as a biomaterial in vivo.
【学位授予单位】：中南大学
【学位级别】：硕士
【学位授予年份】：2012
【分类号】：R318.08
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