Effect of honeycomb-patterned surface topography on the adhesion and signal transduction of porcine aortic endothelial cells
Effect of honeycomb-patterned surface topography on the adhesion and signal transduction of porcine aortic endothelial cells
复制标题
DOI：
10.1021/la7003326
复制
发表时间：
2007-07-17
期刊：
影响因子：
3.9
通讯作者：
Shimomura, M.
中科院分区：
文献类型：
--
作者：
Yamamoto, S.;Tanaka, M.;Shimomura, M.
表面形貌在细胞反应中具有重要作用。在这里，调查背后的细胞响应表面形貌的机制，我们准备了蜂窝状（HC）图案化的膜，从聚（ε-己内酯）（PCL）与微图案化的表面形貌通过铸造水不混溶溶剂的聚合物溶液在高湿度下。我们采用原子力显微镜（AFM）和激光共聚焦扫描显微镜（CLSM）表征了纤维连接蛋白（Fn）在膜上的吸附。用免疫荧光标记法观察了猪主动脉内皮细胞（PAECs）对吸附在HC-图案化膜上的Fn分子的反应。用免疫沉淀法测定培养1 h时酪氨酸残基自磷酸化的黏着斑激酶（pFAK）的表达。Fn选择性地吸附在孔边缘周围，形成环状聚集体。免疫染色结果显示，PAECs粘附到HC图案化的膜上，位于孔周边的焦点接触点。这些点对应于Fn的吸附位点。在HC图案化膜上1小时后，pFAK的表达是相应平膜上的3倍，表明由Fn与整联蛋白受体之间的结合介导的信号传导在HC图案化膜上更高度活化。这些结果表明，细胞对HC图案化膜的反应（例如，粘附模式和FAK的磷酸化）来源于由膜的孔结构确定的Fn的规则排列的吸附模式。
Surface topography has vital roles in cellular response. Here, to investigate the mechanism behind cellular response to surface topography, we prepared honeycomb (HC)-patterned films from poly(epsilon-caprolactone) (PCL) with micropatterned surface topography by casting a polymer solution of water-immiscible solvent under high humidity. We characterized the adsorption of fibronectin (Fn) on the film using atomic force microscopy (AFM) and confocal laser scanning microscopy (CLSM). The response of porcine aortic endothelial cells (PAECs) to adsorbed Fn molecules onto HC-patterned films was observed by immunofluorescence labeling of vinculin and the actin fiber of PAECs cultured for 1 and 72 h in serum-free medium. The expression of focal adhesion kinase autophosphorylated at the tyrosine residue (pFAK) at 1 h culture was determined using an immunoprecipitation method. Fn adsorbed selectively around the pore edges to form ring-shaped aggregates. The immunostaining results revealed that PAECs adhered to the HC-patterned films at focal contact points localized around pore peripheries. These points correspond to adsorption sites of Fn. The expression of pFAK after 1 h on the HC-patterned film was 3 times higher than that on a corresponding flat film, indicating that the signaling mediated by the binding between Fn and the integrin receptor was more highly activated on the HC-patterned film. These results suggest that the cellular response to HC-patterned films (e.g., adhesion pattern and phosphorylation of FAK) originates from the regularly aligned adsorption pattern of Fn determined by the pore structure of the film.