摘要
蛋白质在表／界面的吸附以及细胞在材料表面的响应行为对材料的生物相容性起着至关重要的作用。当生物材料和机体环境接触后,首先发生的是体液中的蛋白质在表／界面的吸附,吸附的蛋白质层还可进一步诱导后续的细胞黏附、铺展、增殖等行为。由于材料表面拓扑结构是影响蛋白质和细胞行为的一个重要因素,因此,研究表面拓扑结构对蛋白质吸附和细胞响应行为的影响并阐明三者之间的内在联系,这对设计合理的生物材料表面、提高材料的生物相容性具有重要的理论意义和应用价值。
基于此,本研究以聚二甲基硅氧烷和生物陶瓷为基材,利用微／纳加工技术在基材表面构筑特定的拓扑结构,分别研究化学组分均质和异质的表面拓扑结构以及表面拓扑结构和化学组分的协同作用对蛋白质吸附行为和细胞响应行为的影响。具体内容如下：
1.化学组分均质的表面拓扑结构对蛋白质、细胞行为的影响
采用水热法在45S5生物活性玻璃陶瓷表面构筑化学组分均质(羟基磷灰石(HA))的有序纳米棒结构(Ceramic 1)、无规纳米棒结构(Ceramic 2)和纳米片结构(Ceramic 3)。通过蛋白质吸附测试(单一蛋白质吸附体系和血浆蛋白质吸附体系)、蛋白质电泳(SDS-PAGE)和蛋白质印迹(Western-Blot)分析,研究单纯的表面拓扑结构对蛋白质吸附以及细胞黏附行为的影响。结果表明,不论是在单一蛋白质吸附环境还是在人血浆吸附环境下,具有微／纳拓扑结构的表面均有利于纤维蛋白原(Fg)和人血清白蛋白(HSA)的吸附,且样品比表面积越大(Ceramic 1>Ceramic 2>Ceramic 3)蛋白质吸附量也越高。从人骨髓原代细胞培养结果来看,表面纳米棒结构比纳米片结构(Ceramic 3)更有利于细胞的贴壁和铺展,而有序的纳米棒结构(Ceramic 1)又比无规纳米棒结构(Ceramic 2)的效果要好。由上述结果可知,对化学组分均质的表面拓扑结构而言,拓扑结构是影响蛋白质吸附和细胞响应行为的主要因素。
2.化学组分异质的图案化表面拓扑结构对蛋白质、细胞行为的影响
以聚二甲基硅氧烷(PDMS)为基材,通过硅氢加成反应在PDMS表面接枝烯丙基聚乙二醇(PDMS-PEG)。以透射电镜用铜网为光掩膜,采用紫外光刻蚀技术(Xe2 excimer:172 nm)在PDMS-PEG膜片上制备出化学组分异质的图案化表面拓扑结构。而后在此图案化表面上进行荧光素标记蛋白质(Fg-FITC)吸附以及L929细胞培养实验,结果表明,由于曝光区域的PEG被刻蚀除去并暴露出底层的PDMS基材从而促使Fg在该区域吸附,而未曝光部分的PEG则得以保留并能有效的排斥蛋白质的吸附。因此,Fg限定性地吸附于曝光区域并呈现图案化分布。当进行细胞培养时,细胞培养液中的蛋白质(包括与细胞黏附相关的蛋白质如纤粘蛋白(Fn)、玻粘蛋白(Vn)等)也限定性的吸附于曝光区域,而后L929细胞在该蛋白质吸附层的引导下黏附、铺展于曝光区域,从而实现了细胞图案化。从以上结果可知,对化学组分异质的表面拓扑结构而言,化学组分对蛋白质吸附行为和细胞响应行为的影响占主导地位。
3.表面拓扑结构和化学组分的协同作用对蛋白质、细胞行为的影响
以镁黄长石(Akermanite)为基材,将其浸入模拟体液环境中从而在样品表面生成一层类骨磷灰石层(Akermanite-HAp),扫描电子显微镜(SEM)和X射线衍射仪(XRD)测试结果表明新生层的化学组分为羟基磷灰石并具有一定的微／纳米级棒状拓扑结构。而后以未修饰的镁黄长石作为对照组,考察Fg、HSA、Fn、Vn在样品表面的吸附行为以及骨髓间充质干细胞(BMSC)的增殖生长状况。结果表明：由于Akermanite-HAp样品表面生成的类骨磷灰石层具有一定的微／纳拓扑结构,有利于蛋白质的吸附(包括与细胞黏附相关的蛋白质如Fn、Vn等)以及细胞的黏附和附着,同时微／纳米棒之间的缝隙也有利于溶液的流动、营养物质的交换以及氧气的输送等。另外,这种新生成的类骨磷灰石层具有一定的生物活性,自身也能促进成骨细胞的黏附和增殖。在这些因素的共同作用下使得BMSC在Akermanite-HAp样品表面的增殖水平高于对照组。以上结果说明,表面拓扑结构和化学组分是紧密联系、相辅相成的,两者共同决定着蛋白质和细胞在材料表面的行为。
Protein adsorption on interface and cell responsive behaviors such as adhesion, spreading and proliferationon on biomaterial's surface plays a vital role in determining biomaterial's biocompatibility. As is known to us, proteins (from body fluid) adsorption on surface is the first response after biomaterials contacting with biological environment and this preadsorbed protein layer could further mediate subsequent cell behaviors. Since surface topography has evident effects on protein adsorption and cell responsive behaviors; investigating and elucidating the relationship among surface topography, protein and cell is significant for the designing of rational surface of biomaterials and the improvement of their biocompatibility.
In this research, different surface topographical features were prepared on the substrates of poly(dimethylsiloxane) and bioceramic by micro/nano fabrication techniques. Further experiments were conducted to investigate the single effect of either chemical homogeneous or chemical heterogeneous surface topography as well as the synergic effect of surface topography and chemical composition on protein adsorption and cell responsive behaviors. Detail works were concluded as follows:
1. Effect of chemical homogeneous surface topography on protein and cell
Different surface topographical features (ordered hydroxyapatite (HA) nanorod arrays (Ceramic 1), disordered HA nanorod arrays (Ceramic 2) and HA nanosheet arrays (Ceramic 3)) were prepared on 45S5 bioactive glass-ceramic substrate through hydrothermal process. Protein adsorption (single protein adsorption and human plasma adsorption environment), protein electrophoresis (SDS-PAGE), Western-Blot and cell culture experiments were conducted to investigate the effect of surface topography on protein adsorption and cell adhesion. Results indicated that surfaces with micro/nano topography could enhance protein adsorption in both adsorption environment and higher specific surface area (Ceramic 1>Ceramic 2>Ceramic 3) meant a higher protein adsorption amount. Primary human bone marrow cells adhesion and spreading status on the surface of nanorod array samples were better than that on nanosheet array surface, and cell behaviors on ordered nanorod array sample were the best of all. It is concluded that surface topographical feature is the chief factor in mediating protein adsorption and cell responsive behaviors for chemical homogeneous surface topography.
2. Effect of chemical heterogeneous patterned surface topography on protein
adsorption and cell behavior
Allyl-polyethylene glycol (PEG) was grafted onto poly(dimethylsiloxane) surface (PDMS-PEG) using hydrosilylation reaction. Then chemical heterogeneous patterned PDMS-PEG surface topography was fabricated by ultraviolet lithography (Xe2 excimer:172nm) with copper mesh as a photomask. Fluorescence labelled protein (Fg-FITC) adsorption results indicated that, as PEG in UV exposed domains was etched and PDMS substrate was revealed which could promote protein adsorption while reserved PEG in unexposed regions effectively resisted protein adsorption, fibrinogen (Fg) restrictively adsorbed on UV exposed domains and presented a patterned distribution in accord with the patterned substrate. Likewise, proteins in cell culture medium (including cell adhesive proteins, such as fibronectin (Fn) and vitronectin (Vn)) were confined in UV exposed domains, which promoted L929 cell adhesion and spreading, forming a regular cell pattern. Our results informed that chemical composition is the dominant issue in mediating protein adsorption and cell responsive behaviors for chemical heterogeneous surface topography.
3. The synergic effect of surface topography and chemistry on protein and cell
Akermanite bioceramics were immersed in simulated body fluid to obtain bone-like apatite layer coated samples (Akermanite-HAp). Scanning electron microscope (SEM) and X-ray diffraction (XRD) characterizations demonstrated that this apatite layer exhibited specific micro/nano topography with a chemical composition of hydroxyapatite. Investigation of protein adsorption and bone marrow mesenchymal stem cells (BMSC) proliferation were conducted with unmodified bioceramics (Akermanite) as control samples. Results showed that micro/nano-scale rods topography in apatite layer could enhance protein adsorption, which resulted in a higher protein adsorption amount on Akermanite-HAp than that on Akermanite. This preadsorbed protein layer (including cell adhesive proteins, such as Fn and Vn) could favor BMSC adhesion and differentiation on Akermanite-HAp. Besides, microstructure of bone-like apatite layer may increase the capacity of biological liquid adsorption into ceramic substrates, enhance ionic exchanging with body fluids and facilitate the transporting of oxygen and nutrients, which could permit the close contact of cells on surface and provide convenient sites for bone cell colonization. Furthermore, the newly-formed apatite layer could promote the adhesion and proliferation of osteoblast cells due to its bioactivity. This synergic effect of surface topography and chemical composition made Akermanite-HAp more favorable for cell adhesion and proliferation than that of Akermanite. These results suggested that surface topography and chemical composition were closely interconnected and complementary to each other, affecting protein adsorption and cell responsive behaviors on surface together.
引文
[1]颜汉卿,徐国风,生物医学材料学.天津：天津科技翻译出版公司,1993.
[2]Williams D. F., Definitions in Biomaterials. In:Progress in Biomedical Engineering, Amsterdam:Elsevier.1982.
[3]Hench L. L., Polak J. M., Third-generation biomedical materials. Science,2002,295 (5557): 1014-1017.
[4]Hench L. L., Biomaterials. Science,1980,208 (23):826-831.
[5]Hench L. L., Bioactive ceramics. Ann. N.Y.Acad. Sci.,1988,523 (1):54-71.
[6]Hastings G. W., Ducheyne P., Macromolecular biomaterials. CRC,1984.
[7]Hubbell J., Bioactive biomaterials. Curr. Opin. Biotechnol.,1999,10 (2):123-129.
[8]Leeuwenburgh S. C. G., Jansen J. A., Malda J., et al., Trends in biomaterials research:An analysis of the scientific programme of the World Biomaterials Congress 2008. Biomaterials, 2008,29 (21):3047-3052.
[9]沈家骢,超分子层状结构——组装与功能.北京：群学出版社,2004.
[10]Goodman S. B., Barrena E. G., Takagi M., et al., Biocompatibility of total joint replacements: A review. J. Biomed. Mater. Res. A.,2009,90A (2):603-618.
[11]李世普,生物医用材料导论.武汉：武汉工业出版社，2000.
[12]Brash J. L., Exploiting the current paradigm of blood-material interactions for the rational design of blood-compatible materials. J. Biomater. Sci., Polym. Ed.,2000,11 (11):1135-1146.
[13]Courtney J. M., Lamba N. M. K., Sundaram S., et al., Biomaterials for blood-contacting applications. Biomaterials,1994,15 (10):737-744.
[14]Schulte V. A., Diez M., Mo□ller M., et al., Surface topography induces fibroblast adhesion on intrinsically nonadhesive poly(ethylene glycol) substrates. Biomacromolecules,2009,10 (10): 2795-2801.
[15]Ratner B. D., The engineering of biomaterials exhibiting recognition and specificity. J. Mol. Recognit.,1996,9 (56):617-625.
[16]Andrade J., Hlady V., Protein adsorption and materials biocompatibility:A tutorial review and suggested hypotheses. Adv. Polym. Sci.,1986,79:1-63.
[17]Hoffman A. S., Blood-biomaterial interactions:An overview. In:Cooper S. L., Peppas N. A., Biomaterials:interfacial phenomena and applications, Washington, DC:American Chemical Society.1982.3-8.
[18]Fang F., Szleifer I., Kinetics and thermodynamics of protein adsorption:A generalized molecular theoretical approach. Biophys. J.2001,80 (6):2568-2589.
[19]高长有,马列,医用高分子材料.北京：化学工业出版社，2006.
[20]Brash J. L., Horbett T. A., Proteins at Interfaces:an overview. In:Brash J. L., Horbett T. A., Proteins at interfaces Ⅱ, Fundamentals and applications, Washington, DC:American Chemical Society.1995.1-23.
[21]Holden M. A., Cremer P. S., Microfluidic tools for studying the specific binding, adsorption, and displacement of proteins at interfaces. Annu. Rev. Phys. Chem.,2005,56:369-387.
[22]Noh H., Vogler E. A., Volumetric interpretation of protein adsorption:Competition from mixtures and the Vroman effect. Biomaterials,2007,28 (3):405-422.
[23]Jung S.-Y., Lim S.-M., Albertorio F., et al., The Vroman effect:□ A molecular level description of Fibrinogen displacement. J. Am. Chem. Soc.,2003,125 (42):12782-12786.
[24]Hunter A. K., Carta G., Protein adsorption on novel acrylamido-based polymeric ion-exchangers-Ⅳ. Effects of protein size on adsorption capacity and rate. J. Chromatogr. A, 2002,971 (1-2):105-116.
[25]Archambault J. G., Brash J. L., Protein repellent polyurethane-urea surfaces by chemical grafting of hydroxyl-terminated poly(ethylene oxide):effects of protein size and charge. Colloids Surf., B,2004,33 (2):111-120.
[26]de Vos W. M., Biesheuvel P. M., de Keizer A., et al., Adsorption of the protein bovine serum albumin in a planar poly(acrylic acid) brush layer as measured by optical reflectometry. Langmuir, 2008,24 (13):6575-6584.
[27]Rechendorff K., Hovgaard M. B., Foss M., et al., Enhancement of protein adsorption induced by surface roughness. Langmuir,2006,22 (26):10885-10888.
[28]Blanco E. M., Horton M. A., Mesquida P., Simultaneous investigation of the influence of topography and charge on protein adsorption using artificial nanopatterns. Langmuir,2008,24 (6): 2284-2287.
[29]Roach P., Farrar D., Perry C. C., Surface tailoring for controlled protein adsorption:□ effect of topography at the nanometer scale and chemistry. J. Am. Chem. Soc.,2006,128 (12): 3939-3945.
[30]Chen H., Yuan L., Song W., et al., Biocompatible polymer materials:role of protein-surface interactions. Prog. Polym. Sci.,2008,33 (11):1059-1087.
[31]Chen H., Zhang Z., Chen Y., et al., Protein repellant silicone surfaces by covalent immobilization of poly(ethylene oxide). Biomaterials,2005,26 (15):2391-2399.
[32]Chen H., Zhang Y. X., Li D., et al., Surfaces having dual fibrinolytic and protein resistant properties by immobilization of lysine on polyurethane through a PEG spacer. J. Biomed. Mater. Res. A.,2009,90A (3):940-946.
[33]Ostuni E., Chapman R. G, Holmlin R. E., et al., A survey of structure-property relationships of surfaces that resist the adsorption of protein. Langmuir,2001,17 (18):5605-5620.
[34]X.D. Zhu, H.S. Fan, X.N. Chen, et al., Contribution of surface charge to bovine serum albumin adsorption on hydroxyapatite ceramic particles. Key Eng. Mater.,2007,330-332: 861-864.
[35]Pasche S., Voros J., Griesser H. J., et al., Effects of ionic strength and surface charge on protein adsorption at PEGylated surfaces. J. Phys. Chem. B,2005,109 (37):17545-17552.
[36]van Kooten T. G, Spijker H. T., Busscher H. J., Plasma-treated polystyrene surfaces:model surfaces for studying cell-biomaterial interactions. Biomaterials,2004,25 (10):1735-1747.
[37]蒋稼欢,生物医学微系统技术及应用.北京：化学工业出版社，2006.
[38]Castner D. G., Ratner B. D., Biomedical surface science:foundations to frontiers. Surf. Sci., 2002,500 (1-3):28-60.
[39]Lim J. Y., Donahue H. J., Cell sensing and response to micro- and nanostructured surfaces produced by chemical and topographic patterning. Tissue Eng.,2007,13 (8):1879-1891.
[40]Song W., Chen H., Protein adsorption on materials surfaces with nano-topography. Chin. Sci. Bull.,2007,52 (23):3169-3173.
[41]Curtis A., Wilkinson C., Topographical control of cells. Biomaterials,1997,18 (24): 1573-1583.
[42]Lundqvist M., Sethson I., Jonsson B.-H., Protein adsorption onto silica nanoparticles: conformational changes depend on the particles' curvature and the protein stability. Langmuir, 2004,20(24):10639-10647.
[43]Tsapikouni T. S., Missirlis Y. F., Protein-material interactions:From micro-to-nano scale. Mater. Sci. Eng., B,2008,152 (1-3):2-7.
[44]Koh L. B., Rodriguez I., Venkatraman S. S., Conformational behavior of fibrinogen on topographically modified polymer surfaces. Phys. Chem. Chem. Phys.,2010,12 (35): 10301-10308.
[45]Dufrene Y. F., Marchal T. G., Rouxhet P. G., Influence of substratum surface properties on the organization of adsorbed collagen films:□ in situ characterization by atomic force microscopy. Langmuir,1999,15 (8):2871-2878.
[46]Galli C., Collaud Coen M., Hauert R., et al., Creation of nanostructures to study the topographical dependency of protein adsorption. Colloids Surf., B,2002,26 (3):255-267.
[47]Harrison R. G., The cultivation of tissues in extraneous media as a method of morpho-genetic study. Anat. Rec.,1912,6 (4):181-193.
[48]Zhou F., Yuan L., Huang H., et al., Phenomenon of "contact guidance" on the surface with nano-micro-groove-like pattern and cell physiological effects. Chin. Sci. Bull.,2009,54 (18): 3200-3205.
[49]Charest J. L., Eliason M. T., Garcia A. J., et al., Combined microscale mechanical topography and chemical patterns on polymer cell culture substrates. Biomaterials,2006,27 (11): 2487-2494.
[50]Choi C.-H., Hagvall S. H., Wu B. M., et al., Cell interaction with three-dimensional sharp-tip nanotopography. Biomaterials,2007,28 (9):1672-1679.
[51]Weiss P., Experiments on cell and axon orientation in vitro:The role of colloidal exudates in tissue organization. J. Exp. Zool.,1945,100 (3):353-386.
[52]Rovensky Y. A., Slavnaja I. L., Vasiliev J. M., Behaviour of fibroblast-like cells on grooved surfaces. Exp. Cell. Res.,1971,65 (1):193-201.
[53]Maroudas N. G, Anchorage dependence:Correlation between amount of growth and diameter of bead, for single cells grown on individual glass beads. Exp. Cell. Res.,1972,74 (2): 337-342.
[54]den Braber E. T., de Ruijter J. E., Ginsel L. A., et al., Orientation of ECM protein deposition, fibroblast cytoskeleton, and attachment complex components on silicone microgrooved surfaces. J. Biomed. Mater. Res.,1998,40 (2):291-300.
[55]Von Recum A., Van Kooten T., The influence of micro-topography on cellular response and the implications for silicone implants. J. Biomater. Sci., Polym. Ed.,1996,7 (2):181-198.
[56]den Braber E. T., de Ruijter J. E., Ginsel L. A., et al., Quantitative analysis of fibroblast morphology on microgrooved surfaces with various groove and ridge dimensions. Biomaterials, 1996,17 (21):2037-2044.
[57]Meyle J., Giiltig K., Brich M., et al., Contact guidance of fibroblasts on biomaterial surfaces. J. Mater. Sci.:Mater. Med.,1994,5 (6):463-466-466.
[58]Oakley C., Brunette D. M., Topographic compensation:Guidance and directed locomotion of fibroblasts on grooved micromachined substrata in the absence of microtubules. Cell Motil. Cytoskel.,1995,31 (1):45-58.
[59]Oakley C., Jaeger N. A. F., Brunette D. M., Sensitivity of fibroblasts and their cytoskeletons to substratum topographies:Topographic guidance and topographic compensation by micromachined grooves of different dimensions. Exp. Cell. Res.,1997,234 (2):413-424.
[60]Wood W., Martin P., Structures in focus-filopodia. Int. J. Biochem. Cell Biol.,2002,34 (7): 726-730.
[61]Jiang X., Takayama S., Qian X., et al., Controlling mammalian cell spreading and cytoskeletal arrangement with conveniently fabricated continuous wavy features on poly(dimethylsiloxane). Langmuir,2002,18 (8):3273-3280.
[62]Zhu B., Lu Q., Yin J., et al., Effects of laser-modified polystyrene substrate on CHO cell growth and alignment. J. Biomed. Mater. Res. B.,2004,70B (1):43-48.
[63]Charest J. L., Garcia A. J., King W. P., Myoblast alignment and differentiation on cell culture substrates with microscale topography and model chemistries. Biomaterials,2007,28 (13): 2202-2210.
[64]Loesberg W. A., te Riet J., van Delft F. C. M. J. M., et al., The threshold at which substrate nanogroove dimensions may influence fibroblast alignment and adhesion. Biomaterials,2007,28 (27):3944-3951.
[65]Biggs M. J. P., Richards R. G., McFarlane S., et al., Adhesion formation of primary human osteoblasts and the functional response of mesenchymal stem cells to 330□nm deep microgrooves. J. R. Soc. Interface,2008,5 (27):1231-1242.
[66]Teixeira A. I., Abrams G. A., Bertics P. J., et al., Epithelial contact guidance on well-defined micro- and nanostructured substrates. J Cell Sci,2003,116 (10):1881-1892.
[67]Yim E. K. F., Pang S. W., Leong K. W., Synthetic nanostructures inducing differentiation of human mesenchymal stem cells into neuronal lineage. Exp. Cell. Res.,2007,313 (9):1820-1829.
[68]Gomez N., Lu Y., Chen S. C., et al., Immobilized nerve growth factor and microtopography have distinct effects on polarization versus axon elongation in hippocampal cells in culture. Biomaterials,2007,28 (2):271-284.
[69]Pietak A., McGregor A., Gauthier S., et al., Are micropatterned substrates for directed cell organization an effective method to create ordered 3D tissue constructs? J. Tissue Eng. Regen. Med.,2008,2 (7):450-453.
[70]Seidlits S. K., Lee J. Y., Schmidt C. E., Nanostructured scaffolds for neural applications. Nanomedicine,2008,3 (2):183-199.
[71]Almarza A. J., Yang G. G., Woo S. L. Y., et al., Positive changes in bone marrow-derived cells in response to culture on an aligned bioscaffold. Tissue Eng. Part A,2008,14 (9):1489-1495.
[72]Gates B., Xu Q., Stewart M., et al., New approaches to nanofabrication:molding, printing, and other techniques. Chem. Rev.,2005,105 (4):1171-1196.
[73]Xia D., Li D., Ku Z., et al., Top-down approaches to the formation of silica nanoparticle patterns. Langmuir,2007,23 (10):5377-5385.
[74]Villagomez C. J., Sasaki T., Tour J. M., et al., Bottom-up assembly of molecular wagons on a surface. J. Am. Chem. Soc.,2010,132 (47):16848-16854.
[75]Wu C.-D., Fang T.-H., Lin J.-F., Formation mechanism and mechanics of dip-pen nanolithography using molecular dynamics. Langmuir,2009,26 (5):3237-3241.
[76]Greene M. E., Kinser C. R., Kramer D. E., et al., Application of scanning probe microscopy to the characterization and fabrication of hybrid nanomaterials. Microsc. Res. Tech.,2004,64 (5-6):415-434.
[77]Kim J. H., Seo M., Kim S. Y., Lithographically patterned breath figure of photoresponsive small molecules:dual-patterned honeycomb lines from a combination of bottom-up and top-down lithography.Adv. Mater.,2009,21 (41):4130-4133.
[78]Falconnet D., Koenig A., Assi T., et al., A combined photolithographic and molecular-assembly approach to produce functional micropatterns for applications in the biosciences. Adv. Funct. Mater.,2004,14 (8):749-756.
[79]Bratton D., Yang D., Dai J. Y., et al., Recent progress in high resolution lithography. Polym. Adv. Technol.,2006,17 (2):94-103.
[80]Xia Y., Whitesides G. M., Soft Lithography. Angew. Chem. Int. Ed.,1998,37 (5):550-575.
[81]Saito N., Hayashi K., Sugimura H., et al., The decomposition mechanism of p-chloromethylphenyltrimethoxysiloxane self-assembled monolayers on vacuum ultraviolet irradiation. J. Mater. Chem.,2002,12 (9):2684-2687.
[82]Tanaka S., Naganuma Y., Kato C., et al., Surface modification of vinyl polymers by vacuum ultraviolet light irradiation. J. Photopolym. Sci. Technol.,2003,16 (2):165-170.
[83]Hozumi A., Masuda T., Hayashi K., et al., Spatially defined surface modification of poly(methyl methacrylate) using 172 nm vacuum ultraviolet light. Langmuir,2002,18 (23): 9022-9027.
[84]Juan Lopez Gejo, Manoj N., Sumalekshmy S., et al., Vacuum-ultraviolet photochemically initiated modification of polystyrene surfaces:morphological changes and mechanistic investigations. Photochem. Photobiol. Sci.,2006,5 (10):948-954.
[85]Asakura S., Hozumi A., Yamaguchi T., et al., A simple lithographic method employing 172 nm vacuum ultraviolet light to prepare positive- and negative-tone poly(methyl methacrylate) patterns. Thin Solid Films,2006,500 (1-2):237-240.
[86]Matsuzawa M., Potember R. S., Stenger D. A., et al., Containment and growth of neuroblastoma cells on chemically patterned substrates.J. Neurosci. Methods,1993,50 (2): 253-260.
[87]Hayashi K., Saito N., Sugimura H., et al., Regulation of the surface potential of silicon substrates in micrometer scale with organosilane self-assembled monolayers. Langmuir,2002,18 (20):7469-7472.
[88]Dyer P. E., Excimer laser polymer ablation:twenty years on. Appl. Phys. A- Mater.,2003,77 (2):167-173.
[89]Kurella A., Dahotre N. B., Review paper:surface modification for bioimplants:the role of laser surface engineering. J. Biomater. Appl.,2005,20 (1):5-50.
[90]Lippert T., Interaction of photons with polymers:From surface modification to ablation. Plasma Processes Polym.,2005,2 (7):525-546.
[91]Callewaert K., Martele Y., Breban L., et al., Excimer laser induced patterning of polymeric surfaces. Appl. Surf. Sci.,2003,208:218-225.
[92]Fogarty B. A., Heppert K. E., Cory T. J., et al., Rapid fabrication of poly(dimethylsiloxane)-based microchip capillary electrophoresis devices using CO2 laser ablation. Analyst,2005,130 (6):924-930.
[93]Liu G.-Y., Xu S., Qian Y., Nanofabrication of self-assembled monolayers using scanning probe lithography. Acc. Chem. Res.,2000,33 (7):457-466.
[94]Zhou, Bruckbauer A., Ying, et al., Building Three-Dimensional Surface Biological Assemblies on the Nanometer Scale.Nano Lett.,2003,3 (11):1517-1520.
[95]Geissler M., Xia Y. N., Patterning:Principles and some new developments. Adv. Mater.,2004, 16 (15):1249-1269.
[96]Xia Y, Rogers J. A., Paul K. E., et al., Unconventional methods for fabricating and patterning nanostructures. Chem. Rev.,1999,99 (7):1823-1848.
[97]Offenhausser A., Bocker-Meffert S., Decker T., et al., Microcontact printing of proteins for neuronal cell guidance. Soft Matter,2007,3 (3):290-298.
[98]Kane R. S., Takayama S., Ostuni E., et al., Patterning proteins and cells using soft lithography. Biomaterials,1999,20 (23-24):2363-2376.
[99]Lee E. J., Chan E. W. L., Yousaf M. N., Spatio-Temporal Control of Cell Coculture Interactions on Surfaces. ChemBioChem,2009,10 (10):1648-1653.
[100]Ma H. W., Hyun J. H., Stiller P., et al., "Non-fouling" oligo(ethylene glycol)-functionalized polymer brushes synthesized by surface-initiated atom transfer radical polymerization. Adv. Mater.,2004,16 (4):338-341.
[101]Ma H. W., Hyun J., Zhang Z. P., et al., Fabrication of biofunctionalized quasi-three-dimensional microstructures of a nonfouling comb polymer using soft lithography. Adv. Funct. Mater.,2005,15 (4):529-540.
[102]Whitesides G. M., Grzybowski B., Self-assembly at all scales. Science,2002,295 (5564): 2418-2421.
[103]Bishop A. R., Nuzzo R. G., Self-assembled monolayers:recent developments and applications. Curr. Opin. Colloid Interface Sci.,1996,1 (1):127-136.
[104]Schreiber F., Structure and growth of self-assembling monolayers. Prog. Surf. Sci.,2000,65 (5-8):151-256.
[105]Gooding J. J., Mearns F., Yang W., et al., Self-assembled monolayers into the 21st Century: recent advances and applications. Electroanalysis,2003,15 (2):81-96.
[106]Senaratne W., Andruzzi L., Ober C. K., Self-assembled monolayers and polymer brushes in biotechnology:current applications and future perspectives. Biomacromolecules,2005,6 (5): 2427-2448.
[107]Kumar A., Biebuyck H. A., Whitesides G. M., Patterning self-assembled monolayers: applications in materials science. Langmuir,1994,10 (5):1498-1511.
[108]Mayoral R., Requena J., Moya J. S., et al.,3D Long-range ordering in an SiO2 submicrometer-sphere sintered superstructure. Adv. Mater.,1997,9 (3):257-260.
[109]Larserr A., Grier D., Like-charge attractions in metastable colloidal crystallites. Nature, 1997,385 (16):230-233.
[110]Denkov N., Velev O., Kralchevski P., et al., Mechanism of formation of two-dimensional crystals from latex particles on substrates. Langmuir,1992,8 (12):3183-3190.
[111]Zhang J. H., Li Y. F., Zhang X. M., et al., Colloidal self-assembly meets nanofabrication: from two-dimensional colloidal crystals to nanostructure arrays. Adv. Mater.,2010,22 (38): 4249-4269.
[112]Zhang J. H., Yang B., Patterning colloidal crystals and nanostructure arrays by soft lithography. Adv. Funct. Mater.,2010,20 (20):3411-3424.
[113]Holtz J. H., Asher S. A., Polymerized colloidal crystal hydrogel films as intelligent chemical sensing materials. Nature,1997,389 (6653):829-832.
[114]Yap F. L., Zhang Y, Assembly of polystyrene microspheres and its application in cell micropatterning. Biomaterials,2007,28 (14):2328-2338.
[115]郇春艳,胡平,组织工程用生物材料的表面修饰技术.化工进展,2003,1：13-17.
[116]Lee K.-B., Park S.-J., Mirkin C. A., et al., Protein nanoarrays generated by dip-pen nanolithography. Science,2002,295 (5560):1702-1705.
[117]Hoff J. D., Cheng L.-J., Meyhofer E., et al., Nanoscale protein patterning by imprint lithography. Nano Lett.,2004,4 (5):853-857.
[118]Gerlach A., Knebel G, Guber A. E., et al., Microfabrication of single-use plastic microfluidic devices for high-throughput screening and DNA analysis. Microsyst. Technol.,2002, 7 (5):265-268.
[119]Chen H., Hu X. Y., Zhang Y. X., et al., Effect of chain density and conformation on protein adsorption at PEG-grafted polyurethane surfaces. Colloids Surf., B,2008,61 (2):237-243.
[120]Zhang Z., Chao T., Chen S. F., et al., Superlow fouling sulfobetaine and carboxybetaine polymers on glass slides. Langmuir,2006,22 (24):10072-10077.
[121]Henzler-Wildman K., Kern D., Dynamic personalities of proteins. Nature,2007,450 (7172): 964-972.
[122]Feng W., Zhu S., Ishihara K., et al., Adsorption of fibrinogen and lysozyme on silicon grafted with Poly(2-methacryloyloxyethyl Phosphorylcholine) via surface-initiated atom transfer radical polymerization. Langmuir,2005,21 (13):5980-5987.
[123]Galli C., Coen M. C., Hauert R., et al., Protein adsorption on topographically nanostructured titanium. Surf. Sci.,2001,474:L180-L184.
[124]Sutherland D. S., Broberg M., Nygren H., et al., Influence of nanoscale surface topography and chemistry on the functional behaviour of an adsorbed model macromolecule. Macromol. Biosci.,2001,1 (6):270-273.
[125]Angeles Contreras M., Bale W. F., Spar I. L., Iodine monochloride (IC1) iodination techniques. Methods Enzymol.,1983,92:277-292.
[126]Wagner M. S., Horbett T. A., Castner D. G, Characterizing multicomponent adsorbed protein films using electron spectroscopy for chemical analysis, time-of-flight secondary ion mass spectrometry, and radiolabeling:capabilities and limitations. Biomaterials,2003,24 (11): 1897-1908.
[127]Protein-Data, Data from Protein Data Bank:www.pdb.org.
[128]Yu Q., Zhang Y, Chen H., et al., Protein adsorption on poly(N-isopropylacrylamide)-modified silicon surfaces:Effects of grafted layer thickness and protein size. Colloids Surf., B,2010,76 (2):468-474.
[129]Chen H., Brook M. A., Chen Y, et al., Surface properties of PEO-silicone composites: reducing protein adsorption. J. Biomater. Sci., Polym. Ed.,2005,16 (4):531-548.
[130]Yamada K. M., Hahn L. H., Olden K., Structure and function of the fibronectins. Prog. Clin. Biol. Res.,1980,41:797-819.
[131]Preissner K. T., Structure and biological role of vitronectin. Annu. Rev. Cell Biol.,2003,7 (1):275-310.
[132]Anselme K., Osteoblast adhesion on biomaterials. Biomaterials,2000,21 (7):667-681.
[133]Bignon A., Chouteau J., Chevalier J., et al., Effect of micro- and macroporosity of bone substitutes on their mechanical properties and cellular response. J. Mater. Sci.:Mater. Med.,2003, 14(12):1089-1097.
[134]LeGeros R. Z., Lin S., Rohanizadeh R., et al., Biphasic calcium phosphate bioceramics: preparation, properties and applications. J. Mater. Sci.:Mater. Med.,2003,14 (3):201-209.
[135]Rogers K. R., Recent advances in biosensor techniques for environmental monitoring. Anal. Chim. Acta,2006,568 (1-2):222-231.
[136]Luong J. H. T., Male K. B., Glennon J. D., Biosensor technology:Technology push versus market pull. Biotechnol. Adv.,2008,26 (5):492-500.
[137]Gooding J. J., Biosensor technology for detecting biological warfare agents:Recent progress and future trends. Anal. Chim. Acta,2006,559 (2):137-151.
[138]Veiseh M., Zareie M. H., Zhang M. Q., Highly selective protein patterning on gold-silicon substrates for biosensor applications. Langmuir,2002,18 (17):6671-6678.
[139]Iwata R., Suk-In P., Hoven V. P., et al., Control of nanobiointerfaces generated from well-defined biomimetic polymer brushes for protein and cell manipulations. Biomacromolecules, 2004,5 (6):2308-2314.
[140]Suh K. Y., Seong J., Khademhosseini A., et al., A simple soft lithographic route to fabrication of poly(ethylene glycol) microstructures for protein and cell patterning. Biomaterials, 2004,25 (3):557-563.
[141]Gleiche M., Chi L. F., Fuchs H., Nanoscopic channel lattices with controlled anisotropic wetting. Nature,2000,403 (6766):173-175.
[142]Iwanaga S., Akiyama Y., Kikuchi A., et al., Fabrication of a cell array on ultrathin hydrophilic polymer gels utilising electron beam irradiation and UV excimer laser ablation. Biomaterials,2005,26 (26):5395-5404.
[143]Lee J. Y, Shah S. S., Zimmer C. C., et al., Use of photolithography to encode cell adhesive domains into protein microarrays. Langmuir,2008,24 (5):2232-2239.
[144]Zhang S., Yan L., Altman M., et al., Biological surface engineering:a simple system for cell pattern formation. Biomaterials,1999,20 (13):1213-1220.
[145]Rozkiewicz D. I., Kraan Y, Werten M. W. T., et al., Covalent microcontact printing of proteins for cell patterning. Chem. Eur. J.,2006,12 (24):6290-6297.
[146]Magnani A., Priamo A., Pasqui D., et al., Cell behaviour on chemically microstructured surfaces. Mater. Sci. Eng., C-Bio. S.,2003,23 (3):315-328.
[147]Sugiura S., Edahiro J.-i., Sumaru K., et al., Surface modification of polydimethylsiloxane with photo-grafted poly(ethylene glycol) for micropatterned protein adsorption and cell adhesion. Colloids Surf., B,2008,63 (2):301-305.
[148]Hou S., Yang K., Qin M., et al., Patterning of cells on functionalized poly(dimethylsiloxane) surface prepared by hydrophobin and collagen modification. Biosens. Bioelectron.,2008,24 (4): 912-916.
[149]McDonald J. C., Whitesides G. M., Poly(dimethylsiloxane) as a material for fabricating microfluidic devices. Acc. Chem. Res.,2002,35 (7):491-499.
[150]Piruska A., Nikcevic I., Lee S. H., et al., The autofluorescence of plastic materials and chips measured under laser irradiation. Lab Chip,2005,5 (12):1348-1354.
[151]Chen H., Wang L., Zhang Y. X., et al., Fibrinolytic poly(dimethyl siloxane) surfaces. Macromol. Biosci.,2008,8 (9):863-870.
[152]Wang H., Djurisic A. B., Chan W. K., et al., Factors affecting phase and height contrast of diblock copolymer PS-b-PEO thin films in dynamic force mode atomic force microscopy. Appl. Surf. Sci.,2005,252 (4):1092-1100.
[153]Linder V., Verpoorte E., Thormann W., et al., Surface Biopassivation of Replicated Poly(dimethylsiloxane) Microfluidic Channels and Application to Heterogeneous Immunoreaction with On-Chip Fluorescence Detection. Anal. Chem.,2001,73 (17):4181-4189.
[154]江雷,冯琳,仿生智能纳米界面材料.北京：化学工业出版社，2007.
[155]Sun T., Feng L., Gao X., et al., Bioinspired surfaces with special wettability. Acc. Chem. Res.,2005,38:644-652.
[156]Neinhuis C., Barthlott W., Characterization and Distribution of Water-repellent, Self-cleaning Plant Surfaces. Ann Bot,1997,79 (6):667-677.
[157]Denis F. A., Hanarp P., Sutherland D. S., et al., Protein adsorption on model surfaces with controlled nanotopography and chemistry. Langmuir,2002,18 (3):819-828.
[158]Singhvi R., Kumar A., Lopez G. P., et al., Engineering cell shape and function. Science, 1994,264 (5159):696-698.
[159]Vijayaraj M., Gadiou R., Anselme K., et al., The influence of surface chemistry and pore size on the adsorption of proteins on nanostructured carbon materials. Adv. Funct. Mater.,2010, 20 (15):2489-2499.
[160]Kokubo T., Takadama H., How useful is SBF in predicting in vivo bone bioactivity? Biomaterials,2006,27 (15):2907-2915.
[161]Wu C. T., Chang J., Synthesis and apatite-formation ability of akermanite. Mater. Lett., 2004,58 (19):2415-2417.
[162]Wu C. T., Chang J., A novel akermanite bioceramic:preparation and characteristics. J. Biomater. Appl.,2006,21 (2):119-129.
[163]Kokubo T., Surface chemistry of bioactive glass-ceramics. J. Non-Cryst. Solids,1990,120 (1-3):138-151.
[164]Maniatopoulos C., Sodek J., Melcher A. H., Bone formation in vitro by stromal cells obtained from bone marrow of young adult rats. Cell Tissue Res.,1988,254 (2):317-330.
[165]Li D., Chen H., McClung W. G., et al., Lysine-PEG-modified polyurethane as a fibrinolytic surface:Effect of PEG chain length on protein interactions, platelet interactions and clot lysis. Acta Biomater.,2009,5 (6):1864-1871.
[166]Van Der Plas A., Nijweide P. J., Cell-cell interactions in the osteogenic compartment of bone.Bone,1988,9(2):107-111.
[167]Le Nihouannen D., Daculsi G, Saffarzadeh A., et al., Ectopic bone formation by microporous calcium phosphate ceramic particles in sheep muscles. Bone,2005,36 (6): 1086-1093.
[168]Habibovic P., Yuan H. P., van der Valk C. M., et al.,3D microenvironment as essential element for osteoinduction by biomaterials. Biomaterials,2005,26 (17):3565-3575.
[169]Rouahi M., Gallet O., Champion E., et al., Influence of hydroxyapatite microstructure on human bone cell response. J. Biomed. Mater. Res. A.,2006,78A (2):222-235.
[170]Ducheyne P., Qiu Q., Bioactive ceramics:the effect of surface reactivity on bone formation and bone cell function. Biomaterials,1999,20 (23-24):2287-2303.
[171]Oonishi H., Hench L. L., Wilson J., et al., Quantitative comparison of bone growth behavior in granules of Bioglass(?), A-W glass-ceramic, and hydroxyapatite. J. Biomed. Mater. Res.,2000, 51(1):37-46.
[172]Olmo N., Martin A. I., Salinas A. J., et al., Bioactive sol-gel glasses with and without a hydroxycarbonate apatite layer as substrates for osteoblast cell adhesion and proliferation. Biomaterials,2003,24 (20):3383-3393.
[173]Hulshoff J. E. G, van Dijk K., de Ruijter J. E., et al., Interfacial phenomena:an in vitro study of the effect of calcium phosphate (Ca-P) ceramic on bone formation. J. Biomed. Mater. Res.,1998,40 (3):464-474.
[174]Loty C., Sautier J.-M., Boulekbache H., et al., In vitro bone formation on a bone-like apatite layer prepared by a biomimetic process on a bioactive glass-ceramic. J. Biomed. Mater. Res., 2000,49 (4):423-434.
[175]Hench L. L., Bioceramics:from concept to clinic. J. Am. Ceram. Soc.,1991,74 (7): 1487-1510.
[176]Webster T. J., Ergun C., Doremus R. H., et al., Specific proteins mediate enhanced osteoblast adhesion on nanophase ceramics. J. Biomed. Mater. Res. A.,2000,51 (3):475-483.
