首页|骨植入材料表面微结构对MC3T3-E1成骨细胞成骨分化的影响

骨植入材料表面微结构对MC3T3-E1成骨细胞成骨分化的影响

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背景:骨植入材料表面微纳结构仿生设计可模拟人体天然骨组织中细胞外环境的结构,从而获得较为理想的骨整合功能,但骨植入材料表面微纳结构调控细胞功能及促进成骨的机制还有待进一研究.目的:分析钛片材料微结构表面对MC3T3-E1成骨细胞成骨分化的影响.方法:①在5 V或20 V恒定电压下,采用酸蚀和阳极氧化技术在钛片表面制备不同管径的纳米管阵列,分别记为R5组、R20组.检测纯钛片(未经酸蚀和阳极氧化处理)、R5组、R20组的表面形貌、粗糙度与亲水性.②将对数生长期的MC3T3-E1成骨细胞分别接种于纯钛片、R5组、R20组钛片表面,成骨诱导培养24 h后,采用RT-PCR检测与免疫荧光染色分析细胞机械敏感通道蛋白1的表达;加入含或不含机械敏感通道蛋白1激活剂Yada1的成骨诱导基,培养7 d后进行碱性磷酸酶染色,培养14 d后进行茜素红染色.结果与结论:①扫描电镜下可见纯钛片表面光滑,R5组、R20组钛片表面可见分布相对均匀且有序的纳米管阵列,平均直径分别约30 nm和100 nm,原子力显微镜进一步验证了扫描电镜观察结果.R5组钛片表面粗糙度大于纯钛片(P<0.05),水接触角小于纯钛片(P<0.05);R20组钛片表面粗糙度大于R5组(P<0.05),水接触角小于R5组(P<0.05).②RT-PCR检测与免疫荧光染色显示,R5组细胞机械敏感通道蛋白1的表达高于纯钛片组(P<0.05),R20组细胞机械敏感通道蛋白1的表达高于R5组(P<0.05).在成骨诱导条件下,与未加入Yada1情况相比,纯钛片组加入Yada1后的细胞碱性磷酸酶活性、钙化结节沉积均无明显变化,R5组、R20组加入Yada1后的细胞碱性磷酸酶活性、钙化结节沉积均显著增加(P<0.05);在加入或未加入Yada1情况下,R5组细胞碱性磷酸酶活性与钙化结节沉积均多于纯钛片组(P<0.05),R20组细胞碱性磷酸酶活性与钙化结节沉积均多于R5组(P<0.05).③结果表明,钛片表面微结构可通过激活机械敏感通道蛋白1促进成骨细胞MC3T3-E1的成骨分化.
Effects of microstructured bone implant material surfaces on osteogenic function of MC3T3-E1 osteoblasts
BACKGROUND:The micro/nanostructured gradient biomimetic surface of implant materials can simulate the structure of the extracellular environment in human bone tissue,thereby achieving perfect bone integration function.However,further research is needed on the mechanisms by which the surface microstructure of bone implant materials regulates cell function and promotes osteogenesis.OBJECTIVE:To analyze the effect of titanium sheet microstructure surface on osteogenic differentiation of MC3T3-E1 osteoblasts.METHODS:(1)At a constant voltage of 5 V or 20 V,nanotube arrays of different diameters were prepared on the surface of titanium sheets by acid etching and anodic oxidation techniques,and were recorded as group R5 and group R20,respectively.The surface morphology,roughness,and hydrophilicity of pure titanium sheet(without acid etching or anodizing treatment)were measured in group R5 and group R20.(2)MC3T3-E1 osteoblasts of logarithmic growth stage were inoculated on the surface of pure titanium sheets,R5 group and R20 group respectively.After 24 hours of osteogenic induction culture,the expression of mechanical sensitive channel protein 1 was analyzed by RT-PCR and immunofluorescence staining.Osteoblast inducible base with or without the mechanosensitive channel protein 1 activator Yada1 was added,and alkaline phosphatase staining was performed after 7 days of culture.Alizarin red staining was performed after 14 days of culture.RESULTS AND CONCLUSION:(1)The surface of pure titanium sheets was smooth under scanning electron microscope.Relatively uniform and orderly nanotube arrays with average diameters of about 30 nm and 100 nm were observed on the surface of titanium sheets of groups R5 and R20,respectively.The results of scanning electron microscope were further verified by atomic force microscopy.The surface roughness of titanium sheet of group R5 was higher than that of pure titanium(P<0.05),and the water contact angle was lower than that of pure titanium(P<0.05).The surface roughness of titanium sheet in group R20 was higher than that in group R5(P<0.05),and the water contact angle was lower than that in group R5(P<0.05).(2)RT-PCR and immunofluorescence staining showed that the expression of mechanosensitive channel protein 1 in group R5 was higher than that in pure titanium group(P<0.05),and the expression of mechanosensitive channel protein 1 in group R20 was higher than that in group R5(P<0.05).Under the osteogenic induction,compared with the condition without Yada1,there were no significant changes in the activity of alkaline phosphatase and the deposition of calcified nodules in pure titanium group after Yada1 addition,while the activity of alkaline phosphatase and the deposition of calcified nodules in groups R5 and R20 after Yada1 addition were significantly increased(P<0.05).With or without Yada1,the alkaline phosphatase activity and calcified nodule deposition in group R5 were higher than those in pure titanium group(P<0.05),and the alkaline phosphatase activity and calcified nodule deposition in group R20 were higher than those in group R5(P<0.05).(3)The results show that the surface microstructure of titanium sheet can promote the osteogenic differentiation of osteoblast MC3T3-E1 by activating mechanosensitive channel protein 1.

bone implant materialtitanium implantmaterial microstructuremechanosensitive channel proteinosteogenic differentiationPIEZO1

黄丽苹、李卉、王新革、王瑞、常蓓、李适廷、兰小蓉、李广文

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西南医科大学附属口腔医院,种植科,口颌面修复重建和再生泸州市重点实验室,四川省泸州市 646000

西南医科大学公共卫生学院流行病与统计教研室,四川省泸州市 646000

军事口腔医学国家重点实验室,国家口腔疾病临床医学研究中心,陕西省口腔医学重点实验室,第四军医大学口腔医院修复科,陕西省西安市 710032

空军军医大学唐都医院军队人员医疗保健中心,陕西省西安市 710032

火箭军特色医学中心口腔科,北京市 100088

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骨植入材料 钛种植体 材料微结构 机械敏感通道蛋白 成骨分化 PIEZO1

四川省科技厅重点研发项目泸州市科技局重点研发计划(面上)西南医科大学自科重点项目西南医科大学附属口腔医院院级重点项目

22YFS06342022-SYF-332022ZD0152022Z01

2025

10.12307/2025.230

中国组织工程研究
中国康复医学会,《中国组织工程研究与临床康复》杂志社

中国组织工程研究

北大核心
影响因子：1.387
ISSN：2095-4344
年,卷(期)：2025.29(10)