Biomaterials, Biomechanics, and Tissue Engineering Group, Department of Materials Science and Metallurgy, Universitat Politècnica de Catalunya, Av. Diagonal 647, 08028 Barcelona, Spain.
Acta Biomater. 2010 Jan;6(1):291-301. doi: 10.1016/j.actbio.2009.07.030. Epub 2009 Jul 25.
We investigated the early events of bone matrix formation, and specifically the role of fibronectin (FN) in the initial osteoblast interaction and the subsequent organization of a provisional FN matrix on different rough titanium (Ti) surfaces. Fluorescein isothiocyanate-labelled FN was preadsorbed on these surfaces and studied for its three-dimensional (3-D) organization by confocal microscopy, while its amount was quantified after NaOH extraction. An irregular pattern of adsorption with a higher amount of protein on topographic peaks than on valleys was observed and attributed to the physicochemical heterogeneity of the rough Ti surfaces. MG63 osteoblast-like cells were further cultured on FN-preadsorbed Ti surfaces and an improved initial cellular interaction was observed with increasing roughness. 3-D reconstruction of the immunofluorescence images after 4 days of incubation revealed that osteoblasts deposit FN fibrils in a specific facet-like pattern that is organized within the secreted total matrix overlying the top of the samples. The thickness of this FN layer increased when the roughness of the underlying topography was increased, but not by more than half of the total maximum peak-to-valley distance, as demonstrated with images showing simultaneous reconstruction of fluorescence and topography after 7 days of cell culture.
我们研究了骨基质形成的早期事件，特别是纤连蛋白 (FN) 在初始成骨细胞相互作用以及随后在不同粗糙钛 (Ti) 表面上临时 FN 基质的组织中的作用。荧光素异硫氰酸酯标记的 FN 预先吸附在这些表面上，并通过共焦显微镜研究其三维 (3-D) 组织，同时在 NaOH 提取后定量其数量。观察到吸附的不规则模式，在地形峰上的蛋白质比在山谷上的蛋白质更多，这归因于粗糙 Ti 表面的物理化学异质性。MG63 成骨样细胞进一步在 FN 预吸附的 Ti 表面上培养，并观察到随着粗糙度的增加初始细胞相互作用得到改善。孵育 4 天后免疫荧光图像的 3-D 重建显示，成骨细胞在分泌的总基质上沉积 FN 原纤维，形成特定的面状图案，该图案位于样本顶部之上。当底层形貌的粗糙度增加时，FN 层的厚度增加，但不会超过总最大峰谷距离的一半以上，这可以通过在细胞培养 7 天后显示同时重建荧光和形貌的图像来证明。
