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Keywords:

  • cell adhesion;
  • material surface;
  • soft lithography;
  • surface chemistry

Abstract

  1. Top of page
  2. Abstract
  3. 1. Introduction
  4. 2. Surface and Cell Adhesion
  5. 3. Manipulation of Cells by Soft Lithography
  6. 4. Conclusions and Future Perspective
  7. Acknowledgements
  8. Biographical Information
  9. Biographical Information
  10. Biographical Information

The adhesion of cells on an extracellular matrix (ECM) (in vivo) or the surfaces of materials (in vitro) is a prerequisite for most cells to survive. The rapid growth of nano/microfabrication and biomaterial technologies has provided new materials with excellent surfaces with specific, desirable biological interactions with their surroundings. On one hand, the chemical and physical properties of material surfaces exert an extensive influence on cell adhesion, proliferation, migration, and differentiation. On the other hand, material surfaces are useful for fundamental cell biology research and tissue engineering. In this Review, an overview will be given of the chemical and physical properties of newly developed material surfaces and their biological effects, as well as soft lithographic techniques and their applications in cell biology research. Recent advances in the manipulation of cell adhesion by the combination of surface chemistry and soft lithography will also be highlighted.


1. Introduction

  1. Top of page
  2. Abstract
  3. 1. Introduction
  4. 2. Surface and Cell Adhesion
  5. 3. Manipulation of Cells by Soft Lithography
  6. 4. Conclusions and Future Perspective
  7. Acknowledgements
  8. Biographical Information
  9. Biographical Information
  10. Biographical Information

In vivo, most mammalian cells are adherent and must attach to and spread on a surface, referred to as the extracellular matrix (ECM), in order to survive, proliferate and function.1 The ECM, an insoluble scaffold largely comprised of proteins (such as fibronectin, laminin, vitronectin, and collagens) as well as other large biomolecules, such as glycosaminoglycans, provide a wide range of biochemical and mechanical cues to cells.1, 2 The attachment of cells to surfaces is a complex process involving cell attachment and spreading, focal adhesion formation, and cytoskeleton organization.2 The initial attachment of a cell to a surface is mediated by ligand-receptor interactions. Integrin is the most important membrane receptor that can bind ECM by recognizing a specific peptide sequence of Arg-Gly-Asp (RGD) present in ECM proteins.3, 4 This binding process is dependent on the nature and conformation of adhesion molecules present at the surface. The main components of tissue structures in vivo are protein fibers such as fibrillar collagen, inducing a heterogeneous network of fiber scaffolds of variable density, orientation, and mechanical strength.5 Furthermore, various chemical cues such as hormones, cytokines, chemokines, ionic strength, pH values are present in ECM, providing multiple chemical stimulations for cells.6 Therefore, cells may encounter matrices with very different physical and chemical properties that profoundly influence cell behavior.

Soft lithography represents a non-photolithographic strategy based on self-assembly and replica molding for carrying out micro- and nanofabrication. It provides a convenient, effective, and low-cost method for the formation and manufacturing of micro- and nanostructures. By employing techniques such as microcontact printing (μCP) and microfluidic printing (μFP), the adhesion of cells on surfaces with various chemical and physical properties could be controlled conveniently. With the emergence of novel materials and the development of soft lithographic techniques, as well as their close combination for biology research, a large number of fundamental biological problems have been addressed. In this Review, we focus on the influence of chemical and physical properties of novel material surfaces on cell adhesion (Section 2.1, Figure 1A, and Section 2.2, Figure 1B), and the control of cell adhesion by the combination of surface chemistry and soft lithography, which includes cells on switchable surface (Section 3.2, Figure 1C), cell pattering (Section 3.3.1, Figure 1D), cell capture (Section 3.3.2, Figure 1E), cell mechanics (Section 3.3.3, Figure 1F), cell co-culture (Section 3.3.4, Figure 1G), and tissue engineering (Section 3.3.5, Figure 1H).

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Figure 1. Schematic representation of the structure of this Review. A) Influence of the chemical properties of surfaces on cells; B) influence of the physical properties of surfaces on cells; C) cells on switchable surface; D) cell pattering; E) cell capture; F) cell mechanics; G) cell co-culture; H) Tissue engineering.

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2. Surface and Cell Adhesion

  1. Top of page
  2. Abstract
  3. 1. Introduction
  4. 2. Surface and Cell Adhesion
  5. 3. Manipulation of Cells by Soft Lithography
  6. 4. Conclusions and Future Perspective
  7. Acknowledgements
  8. Biographical Information
  9. Biographical Information
  10. Biographical Information

Various material surfaces with different chemical and physical cues provide novel insight into how proteins, cells and tissues interact with materials. New materials, and the properties that their surfaces impart, are desirable for the next generation of implants, regenerative medicine and tissue engineering devices, as well as biosensors and drug delivery devices for disease diagnosis and treatment. The study of surface-cell interaction is a key step for the further improvement of biomaterials and their biological applications. Here, we provide an overview of chemical and physical properties of material surfaces developed recently. The biological effects of the surfaces are highlighted.

2.1. Chemical Properties of Surfaces

The ECM provides structural integrity and chemical environments for cells. The chemical components of the ECM, such as ligands, growth factors, cytokines, and ionic strengths all determine the fate of the cell.7 Inspired by key features of the ECM, researchers have developed various model chemical surfaces capable of interacting with biological systems such as proteins, nucleic acid, and cells, among which self-assembled monolayers (SAMs) have become an ideal model substrate mimicking the ECM, because it allows the fine control of the composition of the substrate, tunable length of ligand linkers, and spatial patterning of arbitrary geometry.8 In this Review, according to the adhesive properties, we summarize two classes of SAMs-contributed surfaces: inert and adhesive surfaces and their applications in cell biology research.

2.1.1. Inert Surfaces

Surfaces that resist the nonspecific adsorption of biomolecules and cells commonly are called inert surfaces.9, 10 Prime and Whitesides first reported that SAMs tailored with oligo- or poly(ethylene glycol) (OEG or PEG) units were very effective at preventing the nonspecific adsorption of proteins.11 OEG and PEG exclude protein adsorption through mechanisms that are thought to depend on the conformational properties of highly solvated polymer layers. Alkanethiols terminated with tri- or hexa-EG groups are a standard component of SAMs used in biology and biochemistry.8, 9 Apart from OEG or PEG, there are other kinds of SAMs that inhibit the adhesion of proteins and cells effectively including SAMs terminated with oligosarcosines, oligosulfoxides, perfluoroalkyls, or oligo (phosphorylcholine) groups.9, 12–14

2.1.2. Adhesive Surfaces

Cells can adhere to many surfaces through cell-surface interactions. The cell-surface interactions can be divided into two categories: nonspecific and specific interactions. Nonspecific interactions between a cell and a surface do not require a receptor, i.e., only physical attachment. Electrostatic, van der Waals and hydrophobic forces mediate the nonspecific cell–surface adhesion. Proteins typically adsorb to the surface of a biomaterial in a nonspecific way.15 Material surfaces could be modified with functional groups by employing various methods such as SAMs formation, Langmuir–Blodgett deposition, layer-by-layer assembly, and genetically engineered surface-binding peptides among which SAMs are widely implemented in research because the nature and density of functional groups can be easily tuned on SAMs to control nonspecific protein and cell adhesion.16

The nonspecific adsorption of several proteins to SAMs presenting different functional groups such as alkyls, amides, esters, alcohols, and nitriles was studied, demonstrating that the wettability of the SAMs plays a role in protein adsorption efficiency.17, 18 Inspired by the composition of adhesive proteins in mussels, Lee et al. used dopamine self-polymerization to form thin, surface-adherent polydopamine films onto a wide range of inorganic and organic materials.16 Secondary reactions can be used to create a variety of layers, including SAMs through deposition of long-chain molecular building blocks, metal films by electroless metallization, and bioinert and bioactive surfaces via grafting of macromolecules (Figure 2A).16 In a recent study, we studied if polydopamine film can form on OEG SAMs in the same condition as reported for other types of surfaces.19 The result indicates that polydopamine can transfer onto the SAMs with high fidelity and can form flattened sheets on OEG SAMs, suggesting that polydopamine is a soft and sticky material which can be reshaped under compression. The stable adhesion of polydopamine on antifouling surfaces made it suitable for cell patterning. We confined NIH 3T3 fibroblast cells to the polydopamine patterns which were achieved by either μCP or μFP (Figure 2B). Our studies demonstrate that polydopamine is a versatile nonspecific adhesive material for cell adhesion.19

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Figure 2. A) Mussel-inspired surface chemistry. A photograph of a mussel attached to poly(tetrafluoroethylene), schematic illustrations of the interfacial location of Mefp-5, a molecular representation of characteristic amine and catechol groups, the amino acid sequence of Mefp-5, and the structure of dopamine are shown in the figure. Reproduced with permission.16 Copyright 2007, American Association for the Advancement of Science. B) Control cell adhesion on polydopamine patterns. Printed polydopamine on OEG SAM, magnified SEM image of printed polydopamine on OEG SAM, patterned NIH 3T3 cells on polydopamine-printed OEG SAM, and microfluidic-patterned polydopamine on OEG SAM are shown. Reproduced with permission.19 Copyright 2011, America Chemical Society. C) Schematic presentation of surface nanopattern preparation for cell adhesion studies. Micelle nanolithography technique for making ordered (1) and disordered (2) Au nanopatterns on glass surfaces, and fabrication of ligand nanopatterns on PEG passivated glass are also shown. Reproduced with permission.25 Copyright 2009, America Chemical Society.

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Specific cell-material interactions refer to the involvement of specific receptor-ligand bonds in cell adhesion. The expression of receptors on a cell and/or the modulation of receptor affinity or number on a material surface can manipulate cell-material interactions.20, 21 The RGD sequence is an excellent mediator to promote the attachment of cells on surfaces. Its ability to bind a variety of cells through ligand–receptor interactions makes RGD an exceptionally useful sequence for incorporating onto the biomaterial surfaces.22 The RGD modified surfaces can display specificity and binding affinity by changing the conformation and density of the peptides. The surface density of RGD peptides has been demonstrated to elicit different cellular responses and a minimum RGD density of 10 × 10−15 mol/cm2 is needed for cell adhesion.23 The clustering of RGD peptides at the nanoscale level can significantly reduce the average ligand density and cell adhesion.24 Cell adhesion and spreading can be dramatically reduced when the average ligand spacing is greater than ≈67 nm.21 Recently, Ding and colleagues revealed that integrin clustering and such adhesion induced by RGD ligands are dependent on the local order of ligand arrangement on a substrate when the global average ligand spacing is larger than 70 nm, i.e., cell adhesion is “turned off” by RGD nanopattern order and “turned on” by the RGD nanopattern disorder if operating at this range of inter-ligand spacing (Figure 2C).25

2.1.3. Biological Effects of Chemical Surfaces

The interaction between cells and the ECM initiates signaling cascades involved in critical cell functions.4 However, the natural ECM and cell–ECM interactions are complex and a variety of factors can influence the fate of the cells. Many studies have focused on exploring the chemical basis of cell-ECM interactions and showed that chemical functionality and hydrophilicity of the substrate have important roles in cell adhesion and function.26, 27 SAMs with different terminating functional groups on gold, glass, and silicon were used to evaluate the effects of different chemical species on protein adsorption and cell adhesion.28–31 The distribution of ECM proteins varies with protein type and the different terminis of the SAMs.29, 32 The hydrophobicity/hydrophilicity properties of polystyrene surface have been reported to cause different conformations of ECM proteins and different adhered cell numbers, indicating that the conformation of the ECM proteins play a key role in mediating cell adhesion.33, 34 The acute inflammatory response and the adhesion of cells to SAMs with different terminal functional groups were investigated in vivo, and the results showed that the SAMs led to recruitment of inflammatory cells, demonstrating that the chemical nature modulated both the local acute inflammatory reaction and the adhesion of leukocytes.30 Surfaces modified with different chemical groups induced different morphological changes of stem cells and multiple differentiation directions.35, 36 The incorporation of small functional molecules into PEG hydrogels provided a three dimensional (3D) synthetic extracellular environment for stem cells and determined cell differentiation.6 Although the mechanistic details of the chemical moiety-induced biological effects on cells are unknown, the fact that cell-material interactions can be identified as having a crucial role in cell fates.

2.2. Physical Properties of Surfaces

In recent years, it has become increasingly evident that the cellular responses to microenvironmental signals go far beyond the chemical recognition through ligand-receptor interactions, and it encompasses a wide range of physical cues present at the adhesive interface between the cells and the surrounding ECM.37, 38 Cells respond to physical cues at the interfaces, such as the adhesive ligand density and pattern, ECM rigidity, and ECM dimensionality and anisotropy.39–43 In this Review, we introduce the biological effect of the topography and stiffness of the ECM.

2.2.1. Topography

In vivo, the basement membrane, composed of ECM components, is a complex network of pores, fibers, ridges, and other features of nanometer sized dimensions.44, 45 Topographical cues generated by the ECM, independent of biochemistry, have direct effects on cell behavior such as adhesion, migration, cytoskeletal arrangements, and differentiation.46–50 Cells are inherently sensitive to local microscale, mesoscale, and nanoscale topographic and molecular patterns in the ECM environment, a phenomenon called “contact guidance”.51–53 The development of microfluidics and micro-/nanofabrication methods to analyze the cellular response to substrate topography has provided new insights into the interactions of cells with their microenvironments.54–57

Grooves and pillars are the most common feature types employed in the study of the effects of surface structures on cells. The influence of groove patterns on the behavior of cell has been extensively investigated by using various cell types such as fibroblast,58 osteoblast,59 epithelial,60 myoblast,61 etc. A large number of studies revealed that cells tend to align to the long axis of the grooves.58, 60, 62 Kaiser et al. defined the role of groove/ridge dimensions on fibroblast cell migration.63 They found that surface structures significantly influenced cell orientation, migration direction, as well as migration speed in the directions parallel and perpendicular to the grooves/ridge in a surface structure dependent way (Figure 3A). Uttayarat et al. investigated the combination of flow shear stress and groove guidance on endothelial cell migration. When flow direction was oriented parallel to microgrooves, the cells migrated along the microgrooves. When microgrooves were oriented perpendicular to the flow, most cells migrated orthogonal to the grooves and downstream with the flow.64 Lee et al. reported that the nanoscale ridge/groove pattern arrays alone can effectively and rapidly induce the differentiation of human embryonic stem cells into a neuronal lineage without the use any differentiation-inducing agents, indicating the significant role of topography in determining cell fates.65 Apart from physical and chemical cues, the cell-cell interactions also influence cell behavior. We employed Madin–Darby canine kidney (MDCK), a cell line with relatively strong intercellular interactions, and NIH 3T3 fibroblast cells, a cell line with relatively weak intercellular interactions, to study the interplay between contact guidance and intercellular connections. The two types of cells were patterned onto polydimethylsiloxane (PDMS) substrates with microgrooves. Although MDCK cells migrate much more slowly than 3T3 cells on flat substrate, the velocity of migration of MDCK cells parallel to the grooves is higher than that of 3T3 cells perpendicular to the grooves and contact and form cell sheet (Figure 3B). Because MDCK cells have distinct group behavior, they contact each other very tightly, and the cell sheet acts as a barrier to prevent further migration of 3T3 cells. The 3T3 cell group could migrate only invade the space between them. After 72 h, MDCK cells had not contacted, but 3T3 cells had migrated around (Figure 3B). This experiment shows, for the first time, that both cell–cell and cell–substrate interactions simultaneously influent cell group behaviors.66

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Figure 3. A) Migration speed of fibroblast cells in the directions parallel and perpendicular to the grooves/ridge in a surface structure dependent way (left: plane surface and right: structured suface). Reproduced with permission.63 Copyright 2006, Elsevier. B) Left panel: the MDCK cell matrix contacts and forms tight epithelial cell sheets, which trap 3T3 cells in the long narrow region. The 3T3 cell group only could migrate along the grooves. Right panel: the MDCK cell matrix did not contact before 3T3 cells drilled through. MDCK cells did not contact perpendicular to the grooves after 72 h. Reproduced with permission.66 C) Scatter graph of BLI values of recipient mice 1 month after transplantation with different numbers of Fluc MuSCs cultured for 7 days on either hydrogel (black) or plastic (red). Representative bioluminescence images of mice transplanted with each culture condition are shown. Reproduced with permission.41 Copyright 2010, American Association for the Advancement of Science. D) Scanning electron micrographs of hMSCs plated on PDMS micropost arrays of the indicated heights. Images at the bottom are magnifications of the boxed regions in the top images. Reproduced with permission.69 Copyright 2010, Nature Publishing Group.

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The influence of pillar patterns on the behavior of cells has also been extensively studied.67, 68 Mesenchymal stem cells (MSCs) preferentially differentiated and osteosarcoma cancer cells increased their malignant transformation due to the micropillar geometry.69, 70 In particular, increase of pillar heights from 1 to 10 μm affected the in vitro adhesion and guide morphology of fibroblasts by laminin expression enhancement.71 Furthermore, the spacing between 5 and 10 μm of pillars was shown to rearrange the actin cytoskeleton and govern fibroblast migration in vitro.72 Nanotopography alone can induce the differentiation of MSCs into neuronal lineage and induced a more significant upregulation of neuronal markers compared to microtopography, highlighting the importance of feature size in topography induced differentiation.73

Other micro-/nanosized features, such as nodes, pits, pores, and so forth have been reported to influence the behavior of cells.74, 75 The topography of the cell substratum plays an important role in regulating cellular behavior, and micro-/nanofabrication techniques provide useful tools for manipulating cells in both fundamental cell biology research and tissue engineering.

2.2.2. Stiffness

In tissues, adherent cells plus the ECM co-contribute to establish a relatively elastic microenvironment.76 Cells ranging from neurons to osteoblasts adhere, contract, and crawl within tissues where the stiffness of the ECM ranges from about 1 Kpa in brain to 100 Kpa in collageous bone.77 The stiffness of the ECM is known to impact on various cell activities from gene transcription, cytoskeleton remodeling, to cell-cell interactions.78–84 Most of the cells not only sense but also respond to the mechanical properties of the ECM by adjusting their focal adhesion structure, cytoskeleton organization, and overall state.83, 85–87 A study on how matrix stiffness couples with ligand density to modulate cellular responses suggested that substrate compliance and ligand density are orthogonal determinants.88 Stem cells that naturally reside in adult tissues exhibit robust regenerative capacity in vivo that is rapidly lost during in vitro culture. Engler and co-workers studied the effect of stiffness on MSC differentiation and indicated that soft matrices mimicking brain are neurogenic, stiffer matrices that mimic muscle are myogenic, and comparatively rigid matrices that mimic collagenous bone prove osteogenic.77 Gilbert et al. reported that muscle stem cells cultured on soft hydrogel substrates that mimic the elasticity of muscle (12 kPa) self-renew in vitro and contribute extensively to muscle regeneration when subsequently transplanted into mice (Figure 3C).41

Many studies used synthetic ECM analogs such as inert polyacrylamide gels in which the concentration of bis-acrylamide crosslinking sets the elasticity over several orders of magnitude, from extremely soft to stiff.89 The methods based on gels, however, have drawbacks, for example, altered cross-linker amount of synthetic gels, impacts not only bulk mechanics but also molecular-scale material properties including porosity, surface chemistry, backbone flexibility and binding properties of immobilized adhesive ligands.69 Fu et al. established a library of micromolded elastomeric micropost arrays to modulate substrate rigidity independently of effects on adhesive and other material surface properties. They demonstrated that micropost rigidity impacts cell morphology, focal adhesions, cytoskeletal contractility and stem cell differentiation (Figure 3D).69 Such studies provide evidence that by recapitulating physiological tissue rigidity, propagation of adult stem cells is possible, enabling future cell-based tissue repair and tissue engineering.

3. Manipulation of Cells by Soft Lithography

  1. Top of page
  2. Abstract
  3. 1. Introduction
  4. 2. Surface and Cell Adhesion
  5. 3. Manipulation of Cells by Soft Lithography
  6. 4. Conclusions and Future Perspective
  7. Acknowledgements
  8. Biographical Information
  9. Biographical Information
  10. Biographical Information

3.1. Soft Lithography

Whitesides and colleagues have developed a set of techniques for patterning surfaces known as “soft lithography”. The key elements of soft lithography include elastomaric stamps, masks, and prototyping. Soft lithographic techniques, such as SAMs, μCP, and μFP, have been widely used to pattern a variety of different substrates. The details of soft lithography have been extensively reviewed. Here, we only introduce recent progress of the application of soft lithography in biochemistry and biology research.90–92

3.2. Control of Cell Adhesion on Switchable Surface

The structure and function of cells are highly complex. To better regulate cell behavior and resemble complex biological machineries, a variety of methods for the switching of adhesion of cells to surfaces has recently emerged. We focus on electrochemical desorption of SAMs, photoresponsive surfaces, and thermoresponsive surfaces in this Review.

3.2.1. Electrochemical Desorption of SAMs

The stability of alkanethiols on gold surface varies at different range of electrochemical potentials. The potential at which the desorption of the SAMs occurs depends on several factors, such as the length of the alkyl chain, the degree of ordering and the number of intermolecular interactions within the organic film, and the crystallinity of the gold substrate.93 We have developed a method that can release patterned cells for free migration. SAMs of cell adhering and cell-resisting molecules were patterned on gold surface by μCP (Figure 4A). The application of a cathodic voltage released the cells from the confined patterns and caused them to migrate freely. The viability of the cells was not affected by the applied voltage.94 Using this technique, we studied the relationship between the direction of cell migration and its asymmetric shape. We designed teardrop-like asymmetric geometries to first restrict and then release the cells. Cells tend to move toward blunt ends after desorption (Figure 4B).95 The in vivo interactions among cells are complex, thus an in vitro research system that can co-culture multiple types of cells or cells with varying densities is in demand. By electrochemical desorption of SAMs in localized areas defined by a microfluidic system, we patterned multiple types of cells on the same substrate. This technique has the capability to pattern different types of cells with precisely controlled distances while allowing the free exchange of soluble molecules, it also allows these cells to move under the influence of each other.96 Based on our previous work, we applied selective desorption on complex patterns of cells and simulated three types of cell-cell interactions in vivo: 1) those between two types of cells that are both immobilized and confined to isolated areas, such as epithelial cells and polar cells during ovarian development; 2) those between one cell type that is immobile and another that moves freely, such as glial cells and neurons in neurodegenerative disorders; and 3) those between two or more types of cells that are both moving freely, such as hepatocytes and fibroblasts in the liver (Figure 4C). The system is likely to make co-culturing of different types of cells dramatically more accessible to biologists.97

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Figure 4. A) Application of a cathodic voltage pulse released cells from the microislands. The numbers indicate the time (in minutes) elapsed after the voltage pulse. Reproduced with permission.94 Copyright 2003, America Chemical Society. B) Asymmetric patterns polarize immobilized cells and the location of nucleus, Golgi, and centrosome in the tear drop shaped cells. Reproduced with permission.95 Copyright 2005, the National Academy of Sciences. C) Time-lapse phase-contrast and fluorescence micrographs for the three types of cell–cell interactions between 3T6 and NIH3T3 cells. Reproduced with permission.97 D) Demonstration of a dynamic substrate that combines two dynamic properties: (i) the release of RGD ligands and, thus, the release of cells, (ii) the immobilization of RGD ligands and, hence, migration and growth of cells. Reproduced with permission.98 Copyright 2003, America Chemical Society.

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Conventional μCP methods produce static surface stimuli. Once cells are adherent, it is difficult to subsequently alter the matrix environment. Yeo et al. demonstrated an approach using a SAM that incorporates an O-silyl hydroquinone moiety to present a peptide ligand (E*-RGD). The O-silyl hydroquinone ether is electroactive and can be oxidized by electrical potential application and selectively release the peptide and adhered cells from the substrate. The oxidation of O-silyl hydroquinone ether produces benzoquinone group which undergoes a selective immobilization reaction with a diene-tagged peptide (RGD-Cp) by way of a Diels-Alder reaction and, therefore, provides the basis for a second dynamic activity. The benzoquinone is redox active and can be reduced to the hydroquinone, which prevents immobilization of the diene-tagged ligand. The two dynamic properties of the substrate can sequentially release and attach cells (Figure 4D).98 Wittstock and colleagues described a strategy for the real-time local manipulation of the cell-adhesive property of an OEG terminated SAM substrate using ultramicroelectrodes (UMEs). The strategy is based on the cytophobic nature of OEG SAMs which rapidly switch to cell adhesive by exposure to Br2, which can be electrogenerated from Br in aqueous solution. By using this method, they can fabricate cellular micropatterns to direct cell adhesion and growth in situ.99 On-demand immobilization of cells at specific locations in a microfluidic device would advance many types of bioassays. Nishizawa and colleagues reported a strategy to create a patterned surface within a microfluidic channel by electrochemical means. By locally generating hypobromous acid at a microelectrode in the microchannel, the heparin-coated channel surface rapidly switches from antibiofouling to protein-adhering, enabling site-specific immobilization of protein matrices and cells under physiological conditions.100

3.2.2. Photoresponsive Surfaces

Photoresponsive polymers are photosensitive materials whose physical and chemical properties, such as conformation, shape, surface wettability, membrane potential, membrane permeability, pH, solubility, sol-gel transition temperature, and phase separation temperature of polymer, can be changed reversibly by photoirradiation.101, 102 There are numerous functional groups that can render polymers photoresponsive, such as azo groups, merocyanines, fulgides, etc.102 Azo dyes are particularly popular as photoresponsive groups and, consequently, many examples of photoresponsive polymer surfaces incorporating azo dyes can be found. Wan et al. used surface-tethered azo for the immobilization of partially cyclodextrin-modified poly (acrylic acid) via the photocontrolled host-guest interaction of the azo dye and the cyclodextrin. This surface was capable of binding and releasing cytochrome C.103 Pearson et al. reported a method to incorporate azo groups to interacting motifs. The azo capable of binding to chymotrypsin in the Z-form; rendered the system to be photoswitchable. The extent of binding of a-chymotrypsin to the azo-modified surface could be modulated by irradiation with either UV or visible light. Upon UV irradiation, the maximum binding capacity of the surface was activated, after irradiation with visible light, the binding capacity was reduced to approximately 60% of the maximum (Figure 5A).104 Auernheimer et al. synthesized a set of RGD peptides containing azo as spacer between the acrylamide anchor and the RGD peptide to control cell adhesion, the result showed that the distance and orientation of the RGD peptides at the surface can be changed and the poly(methyl methacrylate) surfaces allows some level of control of cell adhesion.105 Many previous studies have shown that presenting RGD peptide in a background of PEG terminated SAMs allows specific interactions between cells and surfaces, and prevents nonspecific adhesion.106 We generated a surface that allows the azo unit to reversibly present a ligand that contains RGD peptide on SAMs. The E isomer of azo can present the RGD peptide for cell adhesion, while the Z isomer of azo can mask the RGD peptide in PEG terminated SAMs to prevent cell adhesion (Figure 5B). The interconversion between E and Z can be achieved with two wavelengths of light (UV light, 340–380 nm, for the E to Z conversion, and visible light, 450–490 nm, for the Z to E conversion) generated by the mercury lamp of a standard fluorescence microscope. Because the E-to-Z isomerization is completely reversible, this method provides the only means we know to control cell adhesion reversibly on a molecularly well-defined surface.107

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Figure 5. A) Schematic of surface photoswitching showing surface-attached (E)-1 and R-chymotrypsin-bound (Z)-1. Reproduced with permission.104 Copyright 2007, America Chemical Society. B) The azo moiety can be converted photochemically between the E and Z conformations to either present or mask the RGD ligand and hence modulate biospecific cell adhesion. Reproduced with permission.107 C) Phase-contrast microscopy images of L929 mouse fibroblasts on poly(OEGMA-co-MEO2MA)-modified gold substrates after 44 h of incubation at 37 °C and 30 min after cooling the sample to 25 °C. The top panel shows a schematic view of the polymer coatings at 37 and 25 °C. Reproduced with permission.117

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3.2.3. Thermoresponsive Surfaces

Temperature is a very convenient stimulus for the manipulation of cell adhesion by controlling “on-off” switching of thermoresponsive surfaces prepared using thermoresponsive polymers.108, 109 Thermoresponsive polymers generally exhibit a lower critical solution temperature (LCST), below which they are soluble, and above which they dehydrate and aggregate.109 The surfaces made of these stimuli-responsive polymers switch from hydrophilic to hydrophobic states in response to temperature changes. Because of a LCST around 32 °C in water, poly(N-isopropylacrylamide) (PIPAAm) has been proven to be an excellent thermosensitive material for controlling cell adhesion.110 PIPAAm-grafted cell culture substrates exhibit a thermoresponse during cell attachment/detachment.111 Cultured cells are harvested when the temperature is decreased from 37 to 20 °C without the use of digestive enzymes or chelating agents. A confluent cultured cell monolayer, which is detached from the PIPAAm-grafted cell culture substrate by decreasing the temperature, is used in regenerative medicine.112 By controlling the thickness of the grafted thermoresponsive polymeric layer on cell culture dishes, the recovery of confluent cells could be achieved.113, 114 The recovered cell sheets retaining the cell-cell junctions were easily transferred onto other materials to construct 3D tissues from cell sheets using a thermoresponsive cell culture dish.112, 115 Apart from PIPAAm, several other thermally responsive systems can also be cited, such as poly- or oligo (ethylene glycol) methacrylate (OEGMA) derivatives.116 Wischerhoff and co-workers showed that copolymers of 2-(2-methoxyethoxy) ethyl methacrylate (MEO2MA) and OEGMA exhibited a LCST in water that can be adjusted to physiological temperature by varying the composition of the two monomers. Polymer brushes have been prepared on flat gold surfaces demonstrating an ability to control cell adhesion.117 At 37 °C cells adhered to the surface and when the temperature of the medium was decreased to 25 °C, a cell rounding was observed which allowed their easy detachment from the surface without trypsinization (Figure 5C).117 Collectively, the cell culture method using the thermoresponsive cell culture dish is considered a powerful tool for investigating the molecular machinery involved in cell-surface detachment.

3.3. Fundamental Biological Research using Soft Lithography

3.3.1. Cell Patterning

The ability to position cells on a substrate with control over their size and spatial arrangement has facilitated fundamental studies in cellular research.118 Micropatterned cell cultures are ideal to address fundamental issues such as cell-cell interaction and cell-substrate interaction.119, 120 Cell adhesive regions of varying shape and size can be controlled by using microfabrication techniques. When cells adhere to the substrate, the shape of the cells will closely match the size and shape of the adhesive patterns.121 In this way, many fundamental cell problems have been addressed and it has been shown that the shape of cells and the changes in the patterns of cell contact dramatically affect cell viability,39 division,122 proliferation, and differentiation.123 Studies addressing the relationship between cell shape and cytoskeleton demonstrated that concave features promote the assembly of contractile stress filaments, convex features promote the assembly of lamellipodia, and punctuate features promote the assembly of strong focal adhesions.124–126 Substrate patterning provides a useful tool for studying neuronal behavior.127 ECM proteins and cell-cell adhesion molecules play important roles in the development and differentiation of neurons. Recently, we fabricated laminin stripes on a background of poly-L-lysine as substrates for the growth of rat hippocampal neurons, and found that a sharp change of the concentration of laminin guides the growth of neurites by leading the growth cones in a time- and space-dependent manner. The percentage of neurites that grow along the edge of laminin stripes (where there is a sharp change of concentration) decreases as a function of the concentration of laminin under a threshold value. The actin cytoskeleton plays an important role in the process of growth cone's response to the sharp change of concentration of laminin on micropatterns.128 Neurons are dynamically coupled with each other through neurite-mediated adhesion during development. Understanding the collective behavior of neurons in circuits is important for understanding neural development. We established a two-dimensional model for studying collective neuronal migration of a circuit, with hippocampal neurons on Matrigel-coated SAMs. When the neural circuit is subject to geometric constraints of a critical scale, we found that the collective behavior of neuronal migration is spatiotemporally coordinated. Neuronal somata that are evenly distributed upon adhesion tend to aggregate at the geometric center of the circuit, forming monoclusters. Clustering formation is geometry-dependent. Finally, somata clustering is neuron-type specific, and glutamatergic and GABAergic neurons tend to aggregate homophilically. The discovery of geometry-dependent collective neuronal migration and the formation of somata clustering in vitro shed light on neural development in vivo.129 The behavior of neurons on patterned substrates may aid in the design of scaffoldings and nerve guides tailored for regeneration and repair of the nervous system. Recently, we reported a method for replica molding electrospun fibers on the surface of PDMS and its application in culturing and guiding of neurons. With this method, microgrooves and microstructures composed of microgrooves can be obtained. PDMS is integrated into the microfluidic chip as a substrate to successfully pattern and guide neurites on the PDMS surface with microgrooves.130 Conclusively, the studies on behavior of patterned neurons can help us to understand how neurons develop and organize into functional circuits and networks in vivo. Furthermore, it would be possible to develop information technologies, such as neuron-based biosensors and artificial neuronal networks.

3.3.2. Cell Capture

Circulating tumor cells (CTCs) are rare in the blood of patients with metastatic cancer, hence their isolation presents a tremendous technical challenge.131, 132 Microfluidic lab-on-a-chip devices provide unique opportunities for cell sorting and rare-cell detection.133 Nagrath and co-workers developed a microfluidic device, which consists of an array of microposts that are made chemically functional with anti-epithelial-cell adhesion-molecule antibodies, to efficiently and reproducibly isolate CTCs from the blood of patients with common epithelial tumors.133 Wang et al. used a 3D nanostructured substrate, namely, a silicon-nanopillar array coated with epithelial-cell adhesion-molecule antibody, to enhance local topographic interactions between nanoscale cell-surface components and the substrates surface, resulting in enhanced cell-capture efficiency.134 They further integrated an antibody-coated silicon-nanopillar substrate with an overlaid PDMS microfluidic chaotic mixer and significantly improved sensitivity in detecting rare CTCs from whole blood.135 The disposal of maximum non-target species is just as important as retention of maximum target species, aiming at this, Gleghorn et al. presented a theoretical framework for the use of staggered obstacle arrays to create size-dependent particle trajectories that maximize prostate cancer CTCs capture while minimizing the capture of other blood cells.136 Zheng et al. proposed a fluidic scheme to promote maximum target-cell attachment by using a slow flow field, following by a faster flow field for maximum detachment of non-target cells.137

3.3.3. Cell Mechanics

Mechanical stimuli play important roles in the development and maintenance of many tissues.138, 139 Cells sense mechanical microenvironment and initiate signaling pathways by activating mechanosensitive molecules and triggering signaling cascades that eventually alter patterns of gene expression.138, 140 There are many methods that can mimic the in vivo mechanical microenvironment of cells, such as optical tweezers,141 flow chambers,142 and micropipettes.143 A straightforward method to provide mechanical stimuli to cells is to culture cells on elastic membranes where cells are stretched, and fixed for immunostaining or biochemical analysis.144 Existing devices do not allow real-time viewing of live cells at high resolutions due to large distances between cells and objectives and vertical shifts of elastic membranes during stretching. We present a device for stretching cells adhering to elastic membranes in equiaxial or uniaxial mode, meanwhile allowing real-time imaging of molecular dynamics of live cells at high resolution on a microscope during the entire process of the stretch. We obtained high-resolution images of stress fibers at each stage of the stretch, and for the first time, captured real-time images of the process of stress fiber disassembly and reassembly during stretching.145 Based on elastic membrane and microfluidic technique, Douville et al. developed a microsystem to mimic the mechanical environment in alveolus and demonstrated significant morphological differences between alveolar epithelial cells exposed to combination of mechanical and surface-tension stresses compared to cell populations exposed solely to the cyclic stretch. This “alveoli-on-a-chip” research describes new tools for studying the combined effects of two kinds of mechanical stress on cells (Figure 6A).146 Cells normally exist in complex organ systems that are fed by blood vessels and affected by environmental changes, such as the expansion and contraction of lung tissues during inspiration and expiration. These conditions cannot be replicated in an ordinary Petri dish. Ingber and co-workers developed a bioinspired “lung-on-a-chip” that reconstitutes the functional alveolar capillary interface of the human lung (Figure 6B). The device reproduces complex, integrated organ-level responses to bacteria, inflammatory cytokines, and nanoparticles introduced into the alveolar space.147 Based on a similar device, Ingber's group presented an in vitro living cell-based model of the intestine that mimics the mechanical, structural, absorptive, transport and pathophysiological properties of the human gut, i.e., a biomimetic “human gut-on-a-chip” (Figure 6C).148 Mechanically active “organ-on-a-chip” microdevices that reconstitute tissue/tissue interfaces critical to organ function have expanded the capabilities of in vitro cell culture models. To solve the limitations of conventional methods in cardiovascular diseases research, Gunther and colleagues developed a microfluidic device to facilitate the assessment of resistance artery structure and function under physiological conditions. The platform, also termed as “artery–on-a-chip” allows for on-chip fixation, long-term culture and fully automated acquisition of up to ten dose-response sequences of intact mouse mesenteric artery segments in a well-defined microenvironment (Figure 6D).149 Recently, Grosberg et al. reported a ‘‘heart-on-a-chip’’ device that exploits muscular thin film technology biohybrid constructs of an engineered, anisotropic ventricular myocardium on an elastomeric thin film to measure contractility, combined with a quantification of action potential propagation, and cytoskeletal architecture in multiple tissues in the same experiment (Figure 6E). The chip can replicate the hierarchical tissue architectures of laminar cardiac muscle.150

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Figure 6. Biologically inspired design of human “organ-on-a-chip” microdevices. A) A cross-sectional view of an ‘‘alveoli-on-a-chip’’ device. By withdrawing fluid from the ‘‘actuation channel’’, the membrane can be forced to deform and relax stretching cells and propagating the meniscus over a specified cell region. Reproduced with permission.146 Copyright 2011, The Royal Society of Chemistry. B) A “lung-on-a-chip” microdevice recreates physiological breathing movements by applying vacuum to the side chambers and causing mechanical stretching of the PDMS membrane forming the alveolar-capillary barrier. Reproduced with permission.147 Copyright 2010, American Association for the Advancement of Science. C) A schematic of a “gut-on-a-chip” device showing the flexible porous membrane lined by gut epithelial cells cross horizontally through the central microchannel, and vacuum chambers on both sides. Reproduced with permission.148 Copyright 2012, The Royal Society of Chemistry. D) An “artery-on-a-chip” model consisting of endothelial cells and smooth muscle cells and the reversible procedures for artery segment loading, fixation and inspection. Reproduced with permission.149 Copyright 2010, The Royal Society of Chemistry. E) ‘‘Heart-on-a-chip’’ assembly and the contractility experiment on it: (i) films attached to the substrate, (ii) films bend up at diastole and peak systole, and (iii) the length of films (blue) and x-projection (red) overlaid on ‘‘heart on-a-chip’’ images. Reproduced with permission.150 Copyright 2011, The Royal Society of Chemistry.

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The “organ-on-a-chip” platforms that mimic normal complex interactions between living tissues within an organ, as well as the physical cues that cells normally experience in the body, allow us to address new fundamental biological questions, replace a manually demanding procedure with a scalable approach, and provide low-cost alternatives to animal and clinical studies for drug screening and toxicology applications.151, 152

3.3.4. Cell Co-Culture

The cells in vivo are not closed units, they contact their neighbors and the ECM. These contacts can not only connect cells mechanically but also take part in metabolism and signal transduction.153 When the connection between integrin and the ECM changes, the cytoskeleton will rearrange and the cells can spread and migrate. Cell-ECM and cell-cell interactions, therefore, play important roles in vivo.154 For example, contacts between cell-cell and cell-ECM play significant roles in tissue morphogenesis in the process of embryonic development. Tight connections between vascular endothelial cells comprise the structural foundation of the blood-brain barrier. When neurons lose their contacts with each other, transmission of excitation will break off to result in degenerative pathology. Traditional co-culture systems do not allow different types of cells to adhere on the same substrate with high levels of control, which hampers study of the cell-cell interactions. Therefore, the development of new tools capable of controlling the temporal and spatial arrangement of different kinds of cells is meaningful for cell biology research. Micropatterned co-cultures provide a platform for the study of cell-cell interactions between two or more types of cells in a functional tissue model.

Tumor metastasis involves a multistep process which depends not only on cell-adhesion molecules that control direct cell-cell interactions, but also on cell-secreted molecules. Furthermore, direct cell-cell interactions and cytokine release are not sufficient for intra- and extravasation of tumor cells, but also the events entail changes in cell motility.155 Kaji and colleagues investigated the interactions between HeLa cells and human umbilical vein endothelial cells (HUVECs) by monitoring their movements in a controllable co-culture system. Two complementary, detachable, cell substrates were fabricated by replica molding. Co-culturing was started by mechanically assembling two complementary substrates attached with two types of cells. Using this co-culture system as a tumor/endothelium model, they researched the migration behavior of the two types of cells. The system, capable of characterizing the direction and speed of cell movements, is a convenient tool for investigating the mechanism of tumor cell intra- and extravasation.156

Wound healing is an important process involving complex cell-cell interactions and some pathological events such as pulmonary fibrosis.157 In conventional wound healing research models, the cell-free area is generated by mechanical cell removal or trypsin digestion which detaches and damages the cells, leading to the release of the intracellular content and cellular debris into the nutrition medium. Recently, a microfabricated, soft elastic microstencil consisting of an array of long rectangular trenches, in which kidney epithelial cells were grown to confluence, was employed to replace the traditional scratch tests used in standard wound-healing assays. This study showed that the technique did not traumatize the cells located at the edge of the wound and in vitro wound-healing processes are not necessarily dependent on cell injury and can even be induced in the absence of cell damage.158 Felder et al. created wounds in microchannels by introducing a laminar flow of trypsin focused between two adjacent flows of the cell culture medium.157 Using this system, they analyzed the effects of hepatocyte growth factor (HGF) on alveolar epithelial regeneration. In vivo, the cell-cell distance and the migration of interacting cells influence the wound healing process. How to precisely control cell positions and their migration temporally and spatially is of significance. By combining electrochemistry and microfluidics techniques, we can control not only cell adhesion spatially and temporally, but also the distances between several types of cells on the same surface. By using this method, we developed a microchip that achieves co-culture and makes a ‘‘wound’’ without mechanical tension on adjacent normal cells. We studied the response of MDCK cell collective migration to NIH 3T3 fibroblasts, and lysed the MDCK cell monolayer to mimic the ‘‘wound’’. We observed the phenomenon of contact inhibition of locomotion, where a cell ceases to continue moving in the same direction after contacting with another cell.159

3.3.5. Tissue Engineering

Tissue engineering is a difficult task where living cells must be organized into tissues with structural and physiological features resembling actual structures in the body.160 The ultimate goal of tissue engineering as a medical treatment concept is to replace or restore the anatomic structure and function of damaged, injured, or missing tissue, ultimately providing doctors the ability to replace entire organs.161 But barriers still exist, for example, most efforts in cell micropatterning use microfabrication techniques that are based on silicon, gold, or glass substrates. It is difficult to detach patterned cells from these structures and maintain their arrangements at the same time. Creating cellular patterns on biocompatible and biodegradable biomaterials might solve this problem. Co et al. patterned human microvascular endothelial cells and fibroblast cells on chitosan and gelatin films and fabricated capillary tube-like structures via the assembly of the two kinds of cells.162 Multiple cell types in normal tissues are highly organized and, with their associated vasculature, are often no more than 200 μm from an active capillary supply in order to receive essential metabolic components.163 Tissue fabrication techniques rely largely on scaffold based approaches, however, the insufficient nutrient supply and confined cell migration with the scaffold seriously affect the viability of cells within the scaffold. To solve this problem, Papenburg et al. developed 3D scaffolds consisting of stacked multi-layered porous sheets featuring microchannels to culture cells. The inner-porosity of the sheets allows diffusion of nutrients and signalling molecules between the layers, whereas the microchannels facilitate nutrient supply on all layers. They seeded different types of cells in individual sheets to achieve distribution of the cells and maintained long-time viability of the cells.163 Although the porous scaffolds partially meet the requirement of nutrients for cultured cells, the limited capability to generate microscale vascularization for mass transport, and different rates of cell proliferation compared to scaffold degradation remain problems. Tsuda et al. reported a method to fabricate prevascularized tissue equivalents using multilayered cultures combining micropatterned endothelial cells as vascular precursors with fibroblast monolayer sheets as tissue matrix. Their cell culture substrates were covalently grafted with different thermo-responsive polymers to permit spatial switching of cell adhesion and detachment using applied small temperature changes. The multilayer tissues self-organized into microvascular-like networks after a 5-day tissue culture.164 This technique holds great potential for application in the study of cell-cell communication and angiogenesis in reconstructed 3D environments, as well as for the fabrication of tissues with complex, multicellular architecture. To maintain stable nutrient supply and waste elimination, a bioreactor is a good in vitro model for recapitulating functions of various tissues or organs. Domansky et al. developed a bioreactor that fosters maintenance of 3D liver tissue cultures under constant perfusion and they have integrated multiple bioreactors into an array in a multiwell plate format. The cells in the 3D cultures remained functionally viable after seven days of culture (Figure 7A).165 2D cell-culture systems do not accurately replicate the structure, function, or physiology of living tissues, and systems for 3D cultures exist do not replicate the spatial distributions of oxygen, metabolites, and signaling molecules in tissues. To tackle this problem, Whitesides and colleagues recently reported that stacking and destacking layers of paper impregnated with suspensions of cells in a ECM hydrogel makes it possible to control oxygen and nutrient gradients in 3D and to analyze molecular and genetic responses. Stacking assembles the ‘‘tissue’’, whereas destacking disassembles it, and allows its analysis (Figure 7B). Their study offers a uniquely flexible approach to study cell responses to 3D molecular gradients and to mimic tissue- and organ-level functions.166

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Figure 7. A) A simulation of oxygen transport in the multiwells and cell viability assay of rat liver cells cultured for seven days in the perfused multiwell. Reproduced with permission.165 Copyright 2010, The Royal Society of Chemistry. B) Generation of 3D cultures of defined physical dimensions in the paper-supported hydrogels. Reproduced with permission.166 Copyright 2009, the National Academy of Sciences. C) Three types of cells on a SIRM before and after rolling. Each color indicates a different type of cell. Red are endothelial cells; green: smooth muscle cells; blue: fibroblasts. Reproduced with permission.167

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At the core of tissue engineering is the construction of 3D scaffolds out of biomaterials to provide mechanical support and guide cell growth into new tissues or organs. Recently, we reported a new fabrication strategy that results in stable tubular tissue with a high structural similarity to many biological tubular tissues. Using a stress-induced rolling membrane (SIRM) technique, we used two fabrication steps for their tubular structures. First, different types of cells were delivered and patterned on a two dimensional SIRM using microfluidic channels. Then, the SIRM was released to roll up into a 3D tube. The tubes had different types of cells at specific locations, i.e., different parts of the tube wall were made up of different cells (Figure 7C). Mimicking structural and functional features is a prerequisite for fabricating functional tubular tissues in vitro, and the realization of structural-tissue mimicry may have wide applications in simulation of many tubular tissues and enriches the toolbox for 3D micro-/nanofabrication by initially patterning in 2D and transforming it into 3D.167

4. Conclusions and Future Perspective

  1. Top of page
  2. Abstract
  3. 1. Introduction
  4. 2. Surface and Cell Adhesion
  5. 3. Manipulation of Cells by Soft Lithography
  6. 4. Conclusions and Future Perspective
  7. Acknowledgements
  8. Biographical Information
  9. Biographical Information
  10. Biographical Information

In this Review, we introduced basic principles of cell adhesion on surfaces, chemical and physical properties of novel surfaces and their effects on cell behavior, as well as advances in soft lithography and its use in cell adhesion research. Surfaces with functional chemical groups, such as inert SAMs and adhesive SAMs, and biomimetic physical structures, such as nanopillars and microgrooves, allow us to address basic biological questions and explore novel approaches for manipulating cells. The extensive development of soft lithographic techniques provides us with useful tools for studying cell patterning, cell mechanics, cell capture, cell co-culture, and tissue engineering. The close combination of surface functioning and soft lithographic techniques has already seen success and holds considerable promise for their role in cell biology research. Despite the advances mentioned above, we still face some problems, such as the less robust performance of functionalized surfaces in complex microenvironments, insufficient in vitro condition for optimizing cell function, and still limited approach in precise manipulating cell behavior, all of which will be addressed in our future studies. A further key goal is to engineer highly defined surfaces that can mimic the ECM environment and provide methods to manipulate cell behavior. Achieving this goal will facilitate not only the progress in the field of fundamental cell biology research, but also the development of advanced materials for tissue engineering.

Acknowledgements

  1. Top of page
  2. Abstract
  3. 1. Introduction
  4. 2. Surface and Cell Adhesion
  5. 3. Manipulation of Cells by Soft Lithography
  6. 4. Conclusions and Future Perspective
  7. Acknowledgements
  8. Biographical Information
  9. Biographical Information
  10. Biographical Information

This research was supported by the Ministry of Science and Technology (grant no. 2009CB930001 and 2011CB933201), the National Science Foundation of China (grant no. 31170905, 90813032, 20890020, and 50902025), and the Chinese Academy of Sciences (grant no. KJCX2-YW-M15).

Biographical Information

  1. Top of page
  2. Abstract
  3. 1. Introduction
  4. 2. Surface and Cell Adhesion
  5. 3. Manipulation of Cells by Soft Lithography
  6. 4. Conclusions and Future Perspective
  7. Acknowledgements
  8. Biographical Information
  9. Biographical Information
  10. Biographical Information
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Xingyu Jiang obtained his B.S. from the University of Chicago (1999), followed by an A.M. (2001) and a Ph.D. (2004) from Harvard University (Chemistry), working with Prof. George Whitesides on microfluidics and cell patterning. After a postdoctoral fellowship with Prof. Whitesides, he joined the National Center for Nanoscience and Technology of China (NCNST) in 2005 where he has remained since. Xingyu's research interests include surface chemistry, microfluidics, micro-/nanofabrication, cell biology, immunoassays, and nanomedicine.

Biographical Information

  1. Top of page
  2. Abstract
  3. 1. Introduction
  4. 2. Surface and Cell Adhesion
  5. 3. Manipulation of Cells by Soft Lithography
  6. 4. Conclusions and Future Perspective
  7. Acknowledgements
  8. Biographical Information
  9. Biographical Information
  10. Biographical Information
Thumbnail image of

Wenfu Zheng received his Ph.D. in Biophysics from Peking University (China) in 2008, after 2 years of postdoctoral research in the National Center for Nanoscience and Technology of China (NCNST), he worked as an assistant professor in NCNST. His research focuses on microfluidics, biomechanics, and physiopathology of atherosclerosis.

Biographical Information

  1. Top of page
  2. Abstract
  3. 1. Introduction
  4. 2. Surface and Cell Adhesion
  5. 3. Manipulation of Cells by Soft Lithography
  6. 4. Conclusions and Future Perspective
  7. Acknowledgements
  8. Biographical Information
  9. Biographical Information
  10. Biographical Information
Thumbnail image of

Wei Zhang received his B.S. (1999) and Ph.D. (2003) in Material Science and Engineering from Tsinghua University (China). After 3 years of postdoctoral research in the Department of Chemistry, State University of New York, he worked at Institute of Tsinghua University in Shenzhen. He joined the National Center for Nanoscience and Technology of China (NCNST) as an associate professor in 2008. His research interests include tissue engineering and materials for drug delivery.