Gorbahn Miriam, Klein Marcus O, Lehnert Michael, Ziebart Thomas, Brüllmann Dan, Köper Ingo, Wagner Wilfried, Al-Nawas Bilal, Veith Michael
Department of Applied Natural Science, University of Applied Sciences Gelsenkirchen, Recklinghausen, Germany.
J Oral Maxillofac Surg. 2012 Aug;70(8):1827-34. doi: 10.1016/j.joms.2012.04.004.
Despite the undeniable potential of cell adhesion molecules such as fibronectin to support osteogenic cell responses and consecutive dental implant healing, the most beneficial mode of application onto titanium implant surfaces still requires investigation. Unspecific fibronectin adsorption on titanium dioxide (TiO(2)) surfaces can result in low-loading, high-desorption rates and protein-metal interactions with impaired biologic activity. The aim of the present study was to monitor the osteogenic cell responses (cell adhesion, proliferation, and differentiation) specifically to fibronectin biofunctionalized TiO(2).
An innovative biomimetic streptavidin-biotin layer system allows flexible, but stable, specific binding of biotinylated biomolecules such as fibronectin on TiO(2) surfaces. Transparent glass disks were sputtered with TiO(2). The biomimetic layer system was immobilized by self-assembly and consisted of silane, biotin-derivate, streptavidin, and biotinylated fibronectin (bFN). For the control group, unbiotinylated fibronectin was directly coated onto TiO(2). Early cell adhesion dynamics were quantified using automated processing of light microscopy images within the first 24 hours. Relative mRNA expression of integrin-β1, cyclin D1, runt-related gene 2, alkaline phosphatase, and osteocalcin was obtained using quantitative real-time polymerase chain reactions 3 and 7 days after incubation.
Although untreated TiO(2) preserved a rather immature osteogenic phenotype, both unbiotinylated fibronectin and bFN promoted osteogenic cell adhesion and cell differentiation. In particular, runt-related gene 2 expression was significantly promoted by bFN after 3 days. In contrast, cyclin D1 expression was decreased for unbiotinylated fibronectin and bFN after 7 days.
The introduced biomimetic layer system contributes a coherent immobilization approach of adhesion molecules with promotion of osteogenic cell response in vitro.
尽管诸如纤连蛋白等细胞粘附分子在支持成骨细胞反应及后续牙种植体愈合方面具有不可否认的潜力，但将其应用于钛种植体表面的最有益方式仍有待研究。纤连蛋白在二氧化钛（TiO₂）表面的非特异性吸附可能导致低负载、高解吸率以及蛋白质与金属的相互作用，进而损害生物活性。本研究的目的是监测成骨细胞对纤连蛋白生物功能化TiO₂的特异性反应（细胞粘附、增殖和分化）。
一种创新的仿生链霉亲和素 - 生物素层系统允许生物素化的生物分子（如纤连蛋白）在TiO₂表面灵活但稳定地特异性结合。透明玻璃圆盘溅射有TiO₂。仿生层系统通过自组装固定，由硅烷、生物素衍生物、链霉亲和素和生物素化纤连蛋白（bFN）组成。对于对照组，将未生物素化的纤连蛋白直接涂覆在TiO₂上。使用光学显微镜图像的自动处理在最初24小时内对早期细胞粘附动力学进行定量。在培养3天和7天后，使用定量实时聚合酶链反应获得整合素 - β1、细胞周期蛋白D1、 runt相关基因2、碱性磷酸酶和骨钙素的相对mRNA表达。
尽管未处理的TiO₂保留了相当不成熟的成骨表型，但未生物素化的纤连蛋白和bFN均促进了成骨细胞粘附和细胞分化。特别是，bFN在3天后显著促进了runt相关基因2的表达。相比之下，未生物素化的纤连蛋白和bFN在7天后细胞周期蛋白D1的表达降低。
引入的仿生层系统提供了一种连贯的粘附分子固定方法，可促进体外成骨细胞反应。
