Subbiah Ramesh, Du Ping, Van Se Young, Suhaeri Muhammad, Hwang Mintai P, Lee Kangwon, Park Kwideok
Department of Biomedical Engineering, Korea University of Science and Technology (KUST), Daejon, 305-333, Korea. Center for Biomaterials, Korea Institute of Science and Technology (KIST), Seoul, 136-791, Korea.
Biomed Mater. 2014 Oct 20;9(6):065003. doi: 10.1088/1748-6041/9/6/065003.
An artificial matrix (Fn-Tigra), consisting of graphene oxide (GO) and fibronectin (Fn), is developed on pure titanium (Ti) substrates via an electrodropping technique assisted with a custom-made coaxial needle. The morphology and topography of the resulting artificial matrix is orderly aligned and composed of porous microcavities. In addition, Fn is homogenously distributed and firmly bound onto GO as determined via immunofluorescence and elemental mapping, respectively. The artificial matrix is moderately hydrophobic (63.7°), and exhibits an average roughness of 546 nm and a Young's modulus (E) of approximately 4.8 GPa. The biocompatibility, cellular behavior, and osteogenic potential of preosteoblasts on Fn-Tigra are compared to those of cells cultured on Ti and Ti-GO (Tigra). Cell proliferation and viability are significantly higher on Fn-Tigra and Tigra than that of cells grown on Ti. Focal adhesion molecule (vinculin) expression is highly activated at the central and peripheral area of preosteoblasts when cultured on Fn-Tigra. Furthermore, we demonstrate enhanced in vitro osteogenic differentiation of preosteoblasts cultured on Fn-Tigra over those cultured on bare Ti, as determined via Alizarin red and von Kossa staining, and the analysis of osteocalcin, type I collagen, alkaline phosphatase activity, and calcium contents. Finally, we investigate the biophysical and biomechanical properties of the cells using AFM. While the height and roughness of preosteoblasts increased with time, cell surface area decreased during in vitro osteogenesis over 2 weeks. In addition, the E of cells cultured on Tigra and Fn-Tigra increase in a statistically significant and time-dependent manner by 30%, while those cultured on bare Ti retain a relatively consistent E. In summary, we engineer a biocompatible artificial matrix (Fn-Tigra) capable of osteogenic induction and consequently demonstrate its potential in bone tissue engineering applications.
一种由氧化石墨烯（GO）和纤连蛋白（Fn）组成的人工基质（Fn-Tigra），通过一种借助定制同轴针的电滴技术，在纯钛（Ti）基底上制备而成。所得人工基质的形态和形貌呈有序排列，由多孔微腔组成。此外，通过免疫荧光和元素映射分别测定，Fn均匀分布并牢固地结合在GO上。该人工基质具有适度的疏水性（63.7°），平均粗糙度为546 nm，杨氏模量（E）约为4.8 GPa。将前成骨细胞在Fn-Tigra上的生物相容性、细胞行为和成骨潜力与在Ti和Ti-GO（Tigra）上培养的细胞进行比较。Fn-Tigra和Tigra上的细胞增殖和活力显著高于在Ti上生长的细胞。当在前成骨细胞在Fn-Tigra上培养时，粘着斑分子（纽蛋白）表达在细胞的中央和周边区域高度激活。此外，通过茜素红和冯科萨染色以及骨钙素、I型胶原、碱性磷酸酶活性和钙含量分析，我们证明在Fn-Tigra上培养的前成骨细胞比在裸Ti上培养的细胞具有更强的体外成骨分化能力。最后，我们使用原子力显微镜研究细胞的生物物理和生物力学特性。在前成骨细胞的高度和粗糙度随时间增加的同时，在体外2周的成骨过程中细胞表面积减小。此外，在Tigra和Fn-Tigra上培养的细胞的E以统计学上显著且依赖时间的方式增加了30%，而在裸Ti上培养的细胞的E保持相对一致。总之，我们构建了一种能够诱导成骨的生物相容性人工基质（Fn-Tigra），并因此证明了其在骨组织工程应用中的潜力。
