等离子喷涂与电化学沉积羟基磷灰石涂层形貌对骨髓间充质干细胞的影响  

Influence of plasma spraying and electrochemical deposition of hydroxyapatite coating morphology on bone marrow mesenchymal stem cells

作　　者：孙阳 罗鸣然 郑力 胡维帆 袁峰 Sun Yang;Luo Mingran;Zheng Li;Hu Weifan;Yuan Feng(Graduate School of Xuzhou Medical University,Xuzhou 221000,Jiangsu Province,China;Department of Orthopedics,Affiliated Hospital of Xuzhou Medical University,Xuzhou 221000,Jiangsu Province,China)

机构地区：[1]徐州医科大学研究生院,江苏省徐州市221000 [2]徐州医科大学附属医院骨科,江苏省徐州市221000

出　　处：《中国组织工程研究》2021年第28期4516-4522,共7页Chinese Journal of Tissue Engineering Research

基　　金：江苏省自然科学基金(BE2016647),项目负责人:袁峰。

摘　　要：背景:羟基磷灰石以其优异的生物相容性及骨传导能力在组织工程中作为材料涂层被广泛使用,而其生物性能与制作工艺导致的形貌密切相关。目的:探讨等离子喷涂与电化学沉积羟基磷灰石涂层形貌对骨髓间充质干细增殖、黏附及成骨分化的影响。方法:采用等离子喷涂与电化学沉积法分别在钛支架表面制备羟基磷灰石涂层,以未涂层的钛支架为对照。将大鼠骨髓间充质干细胞与3种支架共培养,共培养1,3,5,7 d时,采用CCK-8法检测细胞增殖;共培养5 d后,采用扫描电镜与激光共聚焦显微镜观察细胞黏附情况;共培养14 d时进行碱性磷酸酶染色,共培养21 d时进行茜素红染色,共培养14,21 d时进行实时定量荧光PCR检测,分析骨髓间充质干细胞的成骨分化。结果与结论:①随着培养时间的延长,各组细胞增殖率增加,等离子喷涂组细胞增殖率高于电化学沉积组、对照组(P<0.05);②扫描电镜显示,3组支架上均可见细胞黏附,其中等离子喷涂组细胞铺展面积最大,对照组细胞铺展面积最小;激光共聚焦显微镜显示,等离子喷涂组肌动蛋白纤维最为致密,细胞伸展状态良好;电化学沉积组细胞肌动蛋白相对致密,细胞伸展状态一般;对照组细胞肌动蛋白相对稀疏,细胞伸展不充分;③等离子喷涂组的碱性磷酸酶活性高于电化学沉积组、对照组,钙结节形成多于电化学沉积组、对照组;④实时定量荧光PCR检测显示,除7 d的骨钙蛋白mRNA与14 d的Runx2 mRNA表达无差异外,等离子喷涂组相同时间下的骨桥蛋白、Ⅰ型胶原蛋白、骨钙蛋白、Runx2 mRNA表达均高于电化学沉积组(P<0.05);⑤结果表明,等离子喷涂羟基磷灰石涂层形貌更有利于骨髓间充质干细胞的增殖、黏附与成骨分化。BACKGROUND:With its excellent biocompatibility and osteoconductivity,hydroxyapatite is widely used as a material coating in tissue engineering,and its biological properties are strongly associated with the morphology caused by the manufacturing process.OBJECTIVE:To investigate the effects of plasma spraying and electrochemical deposition on the stent of hydroxyapatite coating on the proliferation,adhesion and osteogenic differentiation of bone marrow mesenchymal stem cells.METHODS:Hydroxyapatite coating was prepared on titanium stent using plasma spraying and electrochemical deposition.The titanium stent without coating was used as a control.Rat bone marrow mesenchymal stem cells were cocultured with three kinds of stents for 1,3,5,and 7 days.Cell proliferation was detected using CCK-8 assay for 5 days.Cell adhesion was observed using scanning electron microscope and confocal laser microscope.At 14 days,alkaline phosphatase staining was conducted;at 21 days,alizarin red staining was performed.At 14 and 21 days,real-time quantitative fluorescent polymerase chain reaction was conducted to analyze osteogenic differentiation of bone marrow mesenchymal stem cells.RESULTS AND CONCLUSION:(1)With the extension of the culture time,the cell proliferation rate of each group increased.The cell proliferation rate of the plasma spraying group was higher than that of the electrochemical deposition group and the control group(P<0.05).(2)Scanning electron microscopy showed that cell adhesion was seen on the stents in the three groups.The plasma spraying group had the largest cell spreading area,and the control group had the smallest cell spreading area.The laser confocal microscope showed that the actin fibers in the plasma spraying group were the densest and the cells were in good extension;the electrochemical deposition group was relatively dense and the cells were in normal extension.In the control group,cytotactin was relatively sparse and the cells were not fully extended.(3)The alkaline phosphatase activity of the plasma sp

关 键 词：骨 材料 钛支架 羟基磷灰石 涂层 等离子喷涂 电化学沉积 骨髓间充质干细胞 

分 类 号：R459.9[医药卫生—治疗学] R318.08[医药卫生—临床医学]