Facile modulation of cell adhesion to a poly(ethylene

Facile modulation of cell adhesion to a poly(ethylene glycol)diacrylate film with incorporation of polystyrene nano-spheres

Wenguang Yang1,2 & Haibo Yu1& Gongxin Li1,2 & Yuechao Wang1 & Lianqing Liu1

# Springer Science+Business Media New York 2016

Abstract Poly(ethylene glycol) diacrylate (PEGDA) is a com-mon hydrogel that has been actively investigated for varioustissue engineering applications owing to its biocompatibilityand excellent mechanical properties. However, the nativePEGDA films are known for their bio-inertness which can hin-der cell adhesion, thereby limiting their applications in tissueengineering and biomedicine. Recently, nano composite tech-nology has become a particularly hot topic, and has led to thedevelopment of new methods for delivering desired propertiesto nanomaterials. In this study, we added polystyrene nano-spheres (PS) into a PEGDA solution to synthesize a nano-composite film and evaluated its characteristics. The experi-mental results showed that addition of the nanospheres to thePEGDA film not only resulted in modification of the mechan-ical properties and surface morphology but further improvedthe adhesion of cells on the film. The tensile modulus showedclear dependence on the addition of PS, which enhanced themechanical properties of the PEGDA-PS film. We attribute thehigh stiffness of the hybrid hydrogel to the formation of addi-tional cross-links between polymeric chains and the nano-sphere surface in the network. The effect of PS on cell adhesionand proliferation was evaluated in L929 mouse fibroblast cellsthat were seeded on the surface of various PEGDA-PS films.

Cells density increased with a larger PS concentration, and thecells displayed a spreading morphology on the hybrid films,which promoted cell proliferation. Impressively, cellular stiff-ness could also be modulated simply by tuning the concentra-tion of nano-spheres. Our results indicate that the addition of PScan effectively tailor the physical and biological properties ofPEGDA as well as the mechanical properties of cells, withbenefits for biomedical and biotechnological applications.

Keywords PEGDA . Cell adhesion . Nano composites .

Mechanical properties . Bioengineering

1 Introduction

Despite progress in the field, tissues engineering currentlyfaces a number of challenges such as effectively mimickingthe various extracellular environment and complex tissue ar-chitecture (Gaharwar et al. 2014; Murphy and Atala 2014).Therefore, designing hydrogels with controlled physical,chemical and biological properties will have a positive impacton facilitating the formation of functional tissues. Over the lastdecade, poly(ethylene glycol) diacrylate (PEGDA) hasemerged as one of the most commonly used hydrogels, whichis extensively applied in drug delivery and in the developmentof biosensors, and also shows potential as scaffold for tissueengineering and regenerative medicine (Murphy and Atala2014; Corbin et al. 2013). The main advantages of PEGDAinclude its tunable mechanical properties, tailorable structureand biocompatibility which can closely mimic the physical,chemical and biological properties of most native tissues.Nevertheless, the ability of cells to interact with and attachto the surface of biomaterials and other cells depends on theamount of protein available for adhesion (Gumbiner 1996;Sun et al. 2013; Liu et al. 2013). Moreover, cell adhesion is

* Haibo [email protected]

* Lianqing [email protected]

1 State Key Laboratory of Robotics, Shenyang Institute of Automation,Chinese Academy of Sciences, Shenyang, People’s Republic ofChina

2 University of Chinese Academy of Sciences, Beijing, People’sRepublic of China

Biomed Microdevices (2016) 18:107 DOI 10.1007/s10544-016-0133-4

http://crossmark.crossref.org/dialog/?doi=10.1007/s10544-016-0133-4&domain=pdf

an essential process for monocyte proliferation, migration andfunction (Baumann 2013; Liu and Wang 2014; Wang et al.2016). However, native PEGDA films have shown cell-repellent properties with various types of cells (Schmidtet al. 2012). This means that cell attachment to PEGDA filmsessentially requires the incorporation of cellular componentsto the cell attachment sites. Although this property is normallyused for obtaining a selective pattern of the accumulation ofcells into arbitrary shapes, the bio-inertness of PEGDA canhinder cell adhesion, which has severely hampered its appli-cations in the field of tissue engineering and biomedicine.Therefore, it would be potentially of benefit to prepare a morecell-friendly PEGDA surface with endowed capabilities ofprotein reception and cell adhesion.

To address this challenge, researchers have focused on mod-ifying the surface of PEGDA films to alter the physical-chemical properties for cell attachment. The extracellular ma-trix (ECM) proteins or related peptides such as fibronectin-derived arginine-glycine-aspartic acid-serine (RGDS) sequenceare most commonly used to functionalize the substrate (Penget al. 2011). Nevertheless, the experimental procedures forPEG-RGDS conjugation is tedious and time-consuming.Other researchers have proposed an interesting method to fab-ricate maleic chitosan-PEGDA hybrid hydrogel throughinserting acrylate functional groups on a polysaccharide byuse of the well-known Michael-type reaction between theamines of chitosan and the double bonds of the diacrylate(Saxena et al. 2014). Another PEGDA surface modificationmethod involves the hydroxyl carboxyl reaction. In this meth-od, the carboxyl groups of PEGDA are activated using 1-ethyl-3- [3-dimethylaminopropyl] carbodiimide hydrochloride andN- hydroxysulfosuccinimide chemistry (EDC/NHS) which al-lows for the PEGDA film to bind to the fibronectin that the cellsadhere to (Han et al. 2008). However, the process of PEGDAmodification with a functional group is complicated and thechemical reagents required may be harmful to cells. To over-come these challenges, a simple method to fabricate nanostruc-tures was developed using lithography and microcontact print-ing to enhance the hydrophobicity of PEGDA films and it wasreported that the cells preferred to adhere on the PEGDA sur-face decoratedwith topographicmicropatterns (Kim et al. 2005,2014). Regardless of these benefits, the methods developedthus far only change the surface topography of PEGDA filmsbut are insufficient to regulate cell behaviors.

Recently, nano composite technology has become a partic-ularly hot topic because of its potential to improve materialsproperties (Gaharwar et al. 2013, 2014; Ahadian et al. 2016;Li et al. 2015; Thakur et al. 2016). Carbon nanotubes (CNTs)are typical carbon-based nanomaterials, which have been in-corporated into photo-cross-linked gelatin methacryloylhydrogels. The CNTs embedded hydrogel scaffold shows ex-cellent mechanical integrity and advanced electrophysiologi-cal functions, which was further explored for biosensor and

bio-actuator applications (Shin et al. 2013). In addition, nanocomposite hydrogels were fabricated using various types ofmetallic nanoparticles for biomedical applications includinggold (Au) and silver (Ag) (Thoniyot et al. 2015). For example,gold nanoparticles (AuNPs) possessing superior electrical andthermal conductivity were functionalized with hyaluronic acidfilms for mechanical reinforcement and roughness alteration(Schmidt et al. 2012). Optimized cell adhesion was observedon the films with a more tunable elastic modulus and surfaceroughness. Ag nanoparticles were shown to possess antimi-crobial properties, and have been incorporated into PAAm andNIPAAm, gelatin-based hydrogels for dental filling and burndressings to prevent infections (Garcia-Astrain et al. 2015;Mohan et al. 2010).

In this study, a polystyrene nano-spheres (PS)-PEGDAcomposite was developed for regulating cell adhesion. Byincorporating a gradually increased amount of PS, the hydro-gel showed enhancedmechanical properties and modified sur-face topography. We next investigated the association of celladhesion and proliferation with the surface coverage of the PS.The results suggest that cell adhesion to PEGDA film can beregulated by tailoring according to the concentration of incor-porated PS. Importantly, this method is a relatively convenientapproach to tune the mechanical properties of cells simply bymanipulating the concentration of the nanospheres, and is ex-pected to be widely applicable for specific needs in biotech-nology and medicine.

2 Materials and methods

2.1 Materials

PEGDA (Mn = 575) and the photoinitiator diphenyl(2,4,6-trimethylbenzoyl)-phosphine oxide (TPO) were obtained fromSigma Aldrich (St. Louis., MO, USA). PS with an average sizeof 100 nm were purchased from BaseLine ChromTechResearch Center. PEGDA and TPO were dissolved in ethylalcohol at a final concentration of 40 % and 0.5 wt%, respec-tively. Then, the PS were added into the PEGDA solution atvarious final concentrations (10%, 20%, 30%, 40%, 50%). Agood dispersion may be achieved by ultrasonic treatment. Allsolutions were magnetically stirred for 30 min and then ultra-sonically treated at 40 kHz for 30 min to ensure that the mate-rials were completely dissolved.

2.2 Fabrication of the PEGDA-PS hydrogel

Glass slides were used as the substrate for preparing the nano-composite hydrogel, as shown in Fig. 1. Prior to ultraviolet(UV) curing, the substrate was washed with alcohol to cleanthe surface and was then dried under nitrogen. The solution(0.5 ml) was pipetted onto the surface of the glass slides. The

107 Page 2 of 9 Biomed Microdevices (2016) 18:107

films containing uniform solution were formed via spin-coating and were then photo cross-linked for 5 min using aUV lamp. The cured films were washed with ethyl alcohol toremove any unreacted solutions and then immersed in aphosphate-buffered saline solution for the cell adhesion test.Furthermore, for the tensile test, the solution was injected intoa rectangular mold (20 mm long, 5 mmwide, and 1 mm thick)and cured under a UV lamp for 10 min.

2.3 Atomic forcemicroscopy (AFM) and scanning electronmicroscopy (SEM)

AFM experiments, including AFM imaging and measurementof cell mechanical properties, were performed using aDimension Icon AFM and Bioscope Catalyst AFM (Bruker,Camarillio, CA, USA). For film surface imaging, the tappingmode of the Dimension Icon AFM was used. The measure-ment of cell mechanical properties was performed with thefluidic contact mode of the Bioscope Catalyst AFM, using atleast 20 force curves of each cell with the same loading rate atdifferent cell positions. The Hertz model was used to calculatethe elastic modulus of cells.

The dried PEGDA-PS hydrogels on the glass slides werecoated with a 10-nm gold film with Sputter Coater, and themorphological variations depending on PS concentrationwereanalyzed using SEM (Zeiss, Germany).

2.4 Cell assay and fluorescence staining

L929 cells (mouse fibroblast cells) were cultivated inDulbecco’s modified Eagle medium with high glucose con-taining 10 % fetal bovine serum and 1 % penicillin-strepto-mycin. Before culturing with the PEGDA-PS films, the cellswere detached from the Petri dish. Subsequently, the cellswere seeded on the films at a concentration of ~104 cells/cm2. The cell medium was changed daily after seeding thecells, and all cells were cultured at 37 °C and 5 % CO2 in anincubator.

An Eclipse Ti microscope (Nikon, Tokyo, Japan) wasemployed to carry out live-cell and fluorescence imaging anal-yses. For cell fluorescence staining, the cells were first fixedwith 4% paraformaldehyde and then treated with 0.1 % TritonX-100 to increase permeability. CytoPainter Phalloidin-iFluor488 Reagent (Abcam, Cambridge, UK) staining solution(100μg/ml) was dropped onto the glass slides containing cellsand kept for 90 min at room temperature. Then, the glassslides were placed on the stage of the Eclipse Ti microscopeand the fluorescence images were acquired.

2.5 Hydrogel mechanical test

To analyze the influence of the nanospheres on the mechanicalproperties of the gels, a tensile test was performed using amechanical test instrument (Bose Enduratec ELF 3200, NewCastle, USA). The specimens (20 mm long, 5 mm wide, and1 mm thick) with different PS concentrations were stretched ata rate of 10 mm/min. Ten samples of each concentration weretested to calculate the Young’s modulus and fracture stress.

2.6 Statistical analysis

Data are presented as the mean ± standard deviation, and sam-ple means were compared using one-way analysis of variancevia OriginPro software. P < 0.05 was considered to be statis-tically significant.

3 Results and discussions

3.1 PS affect film morphology

The nanostructure of the surface is the most important factorcontrolling the adhesion between cells and a substrate.Accordingly, we studied the effect of the addition of nano-spheres on the film’s morphology. PEGDA hydrogel solutionswith different concentrations of PSwere cross-linked with UV

Fig. 1 The PEGDA-PS film wasphoto cross-linked on glass slidesvia UVexposure. Thenanospheres were uniformlydistributed on the PEGDA films.Immersed spheres provided asurface to cross-link withpolymeric chains, and the otherscreated a raised structure on thesurface of the films

Biomed Microdevices (2016) 18:107 Page 3 of 9 107

light, and the surface profile of the fabricated PEGDA hydro-gel was analyzed by SEM (Fig. 2). A representative SEMimage showing a typical cured pure PEGDA hydrogel thatcan prevent cell adherence is presented in Fig. 2f, indicatingthat the surface of the film is relatively flat. However, nano-structures were formed when PS were added to the PEGDAsolution. Some spheres were immersed into the film’s interiorwhile others were fixed on the surface, as shown in Fig. 1. The

spheres increased with increasing PS concentration in thePEGDA solution (Fig. 2b–e), eventually covering the wholefilm (Fig. 2a).

Using the combined AFM images of PEGDA film (Fig. 3),we can calculated the coverage ratio of PS as follows:

Coverageratio %ð Þ¼ Areaof spheresAreaof film

ð1Þ

Fig. 3 AFM images of PEGDAfilm surface morphology withdifferent levels of coverage of thepolystyrene spheres

Fig. 2 SEM analysis of differentconcentrations of polystyrenesphere on the PEGDA film. a-fPS concentrations range from 0%to 50 %, respectively. Scar barsare 1 μm


The size of the whole area of each image is 1 μm× 1 μm.The coverage ratio of PS reached 80 % with a 50 % PS con-centration, as shown in Figs. 3a and 4a, indicating that nano-structures formed by a layer of bare spheres were successfullyfabricated. The calculated values for the coverage ratio ofdifferent concentrations are shown in Fig. 4a, which indicatedthat with the increase in PS concentration, the coverage area ofthe spheres became lager. Detailed information of the modi-fying surface could be obtained from the three-dimensionalAFM image of the surface, as shown in Fig. 4b. The sphereson the PEGDA surface formed nanostructures resembling aseries of mountain peaks. Importantly, the surface morpholo-gy could be controlled by modulating the density of the nano-spheres. Compared with the complexity of previous methodsfor fabricating nanostructures on a PEGDA hydrogel, the fab-ricating process presented herein does not require expensiveequipment or time-consuming steps.

3.2 Effect of PS on mechanical properties of hydrogels

In addition to the observed variation in morphology, we ex-pected that the nanocomposites would induce changes in themechanical properties of the hydrogels. To quantify thechanges in mechanical properties from the addition of PS,we measured the elastic modulus of the films (sample dimen-sion: 20 mm× 5 mm× ∼1 mm) with a tensile test instrument(Fig. 5a). Stress–strain curves of hydrogels with different con-centrations of PS were obtained from the results of the uniax-ial tensile test. As shown in Fig. 5b and c, the hydrogel withPS became significantly stiffer than the pure hydrogel, indi-cating that the addition of nanoparticles reinforced the cross-linked network of the hydrogel. However, greater deformationof the pure hydrogel was observed during fracture stressing,whereas no significant difference was observed among thesamples with various PS concentrations. Therefore, as expect-ed, the mechanical properties were changed by the addition ofnanoparticles. These results also suggest that the reinforce-ment of the hydrogels from PS incorporation is due to theformation of additional cross-links in the network. Testing of

the PEGDA-PS hydrogels also revealed a significant increasein the elastic modulus, of almost 100 % (Fig. 5e).Furthermore, the nanoparticles restricted chain movements,which led to a small deformation when stretched to fracture.The improved physical properties of the hybrid PEGDA hy-drogel is mainly attributed to the enhanced interactions be-tween the polymer chains and nanospheres. During the fabri-cation process, the nanospheres distributed along the PEGDAfilms uniformly, and the immersed spheres provided the sur-face for cross-linking with polymeric chains to further en-hance these interactions.

3.3 Cell adhesion, mechanical properties,and proliferation are associated with surface coverageof the PS

As one kind of the most common hydrogels, PEGDA hasbeen widely investigated for various tissue engineering appli-cations owing to its biocompatibility and excellent mechanicalproperties, however, native PEGDA hydrogels also have cellrepellent properties. By exploiting these properties, we previ-ously fabricated arbitrary shaped PEGDA microstructures topattern cells (Yang et al. 2015, 2016). However, to be consid-ered for biomedical applications, a PEGDA scaffold mustsupport cell adhesion and promote cell proliferation, andtherefore the cell-repellant property must be overcome. Wehypothesized that the addition of nanospheres may enhancethe surface interactions between the nanoparticles and thecells.

Two different hydrogels (pure PEGDA and PEGDA-PS)were cultured with L929 cells. Figure 6 shows that the cellscould grow on the different substrates after culturing for3 days. The pure PEGDA films could not support cell attach-ment, as shown in Fig. 6a and e. In addition, compared withcells growing on a polystyrene tissue culture plate (Fig. 6d andh), those growing on the PEGDA surface displayed a roundshape, indicating poor spreading properties and no stable cell-matrix interactions. However, the PEGDA-PS hydrogelallowed for better cell adhesion compared to the pure

Fig. 4 a Coverage ratio ofpolystyrene nanospheres onPEGDA films. b AFM three-dimensional image of a PEGDAfilm with 30 % polystyrenenanospheres


PEGDA, and a portion of the cells showed a good spreadingshape, as expected. Better adhesion to the substrates shouldhave a positive impact on cell growth and proliferation. This is

because without adhesion to the surface, many cells will stopproliferating and can undergo programmed cell death. Indeed,after 3 days of culture, there was no proliferation of cells

Fig. 5 Effect of polystyrene nanospheres (PS) on the mechanical prop-erties of hydrogels. aHydrogel specimens were tested using a mechanicaltest instrument. b Stress–strain curves of hydrogels with different concen-trations of PS. cWith an increasing PS concentration, the fracture energy

increased. d Changes in the fracture strain with the addition of PS weresmaller relative to those of the pure hydrogel. e The Young modulus ofdifferent hydrogels is calculated. Statistically different pairs (P < 0.05) areindicated by horizontal lines and * or #

Fig. 6 Optical microscopy of cells growing on different films afterculturing for 3 days. a, e Cells growing on pure PEGDA film showinga round shape. b, f Cells growing on the PEGDA film composed of 30 %polystyrene spheres, c, g cells growing on the PEGDA film composed of

50 % polystyrene spheres indicating that a portion of the cells couldadhere to the PEGDA-PS film. d, h Cells adhered on the polystyrenetissue culture plate (TCPS). Scale bars are 100 μm


growing on the pure PEGDA, whereas the cell density in-creased significantly due to the addition of PS.

These results clearly suggest that the addition of PS influ-enced the biocompatibility of the PEGDA hydrogel. To direct-ly evaluate the effect of PS on cell adhesion and proliferation,L929mouse fibroblast cells were seeded on the surfaces of thevarious PEGDA-PS films, and the effect of the addition of PSon cell adhesion was determined by analysis of fluorescenceimages. Cells were cultured with different films for two days

and were stained using CytoPainter Phalloidin-iFluor 488Reagent. As shown in Fig. 7, no cells adhered to the substratewhen grown on the pure PEGDA film, and the addition of PSsignificantly enhanced cell adhesion.

In general, integrins serve as the bridges for cell-cell andcell-ECM interactions, and couple the ECM to the cytoskele-ton. Together with signals arising from receptors for solublegrowth factors, integrins determine the biological activity of acell in a given context, and thus play an important role inmany

Fig. 7 Fluorescence imagesshowing the effect of the additionof polystyrene nanospheres toPEGDA films on cell adhesion. aThe pure PEGDA film. b–fPEGDA films containingpolystyrene nanospheres atconcentrations ranging from 10 to50 %; the addition of PSenhanced cell adhesion to thesubstrate. Scale bar is 100 μm

Fig. 8 Effect of polystyrene nanospheres on the mechanical propertiesand proliferation of cells. a A schematic diagram of cells adhering on thePEGDA-PS film surfaces via attachment of integrins to the polystyrenespheres. b Cell growth improved as the PS concentration increased. cAFM mechanical measurements of cells growing on different films,

indicating that the addition of PS could enhance the cellular stiffness. dThe viability of cells adhering on the different nano composites.Statistically different pairs (P < 0.05) are indicated by horizontal linesand *


cellular biological processes (Seguin et al. 2015). Therefore,on the one hand, the PEGDA hydrogel surface roughnesschanged significantly due to the high concentration of PS asshown in Figs. 3 and 4, which improved the cell adhesion tothe hydrogel, and the large number of integrins promoted cellgrowth in turn (Fig. 8a). On the other hand, recent studies havesuggested that cell adhesion and spread are mainly controlledby the mechanical property of the substrate (i.e., the elasticmodulus) (Nemir et al. 2010; Jaiswal et al. 2015; Rehfeldtet al. 2007). As shown in Fig. 5b, the addition of nanospheresimproved the mechanical properties of the hybrid film. Thesetwo factors can explain the enhanced growth with higher con-centrations of PS on the film. As shown in Fig. 8b, afterculturing for 3 days, the cell density on the 50 % PEGDA-PS film was significantly higher than that on all other films.

Furthermore, the cellular stiffness, which reflects the intra-cellular tension acting along stress fibers, decreased or in-creased instantly in response to the elongating or compressingstimulus, respectively. Cells adhere to a substrate through theirintegrins, which link with the cytoskeleton to regulate the me-chanical properties of cells (Snijder et al. 2013; Mierke 2013;Schoen et al. 2013; Yamahashi et al. 2015). The results of theAFM measurements on cells growing on different films indi-cated that the average Young’s modulus of cells growing on the10 % PEGDA-PS hydrogel was 0.62 ± 0.02 kPa, while that ofcells growing on the 50 % PEGDA-PS hydrogel was higher, at1.01 ± 0.05 kPa (Fig. 8c), which was similar to the Young’smodulus of cells growing on polystyrene tissue culture plates,as a control group (1.04 ± 0.04 kPa). As shown in Fig. 2a, whenthe concentration of PS reached 50 %, the spheres covered anarea of roughly 80 % on the PEGDA film. This suggests thatboth the cell adhesive density and the mechanical properties ofcells could be regulated by tuning the concentration of PS.

4 Conclusions

In summary, we demonstrated that the addition of PS intophoto cross-linked PEGDA resulted in a hybrid film(PEGDA-PS) with a significantly altered surface morphologyand mechanical properties. Comparative studies of surfacetopography via SEM and AFMwere conducted to gain insightinto the changes caused by the addition of the nanospheres.The tensile modulus showed clear dependence on the presenceof PS, and the mechanical properties of the PEGDA-PS filmwere enhanced by PS treatment. Furthermore, L929 mousefibroblast cells seeded on the surface with PS indicated thatthe physical addition of the nanospheres not only improvedthe physical properties of the polymeric networks but couldalso modulate the biocompatibility of the film. Therefore, aseries of hybrid films was fabricated to examine the relation-ships between PS concentration and cell adhesion. Cell den-sity increased with greater PS concentrations, and the cells

displayed a spreading morphology on the hybrid films, whichin turn promoted cell proliferation. In addition, cellular stiff-ness was modulated simply by tuning the PS concentration.As PEGDA is frequently used in a broad range of applicationsin biomaterials science, this study demonstrates a feasiblemethod to obtain control over the mechanical properties andto tailor cell adhesion according to the requirements for tissueengineering.

Acknowledgments This research work was partially supported by theNational Natural Science Foundation of China (Project No. 61302003,61475183 and 61503258), the NSFC/RGC Joint Research Scheme(Project No. 51461165501) and the CAS FEA International PartnershipProgram for Creative Research Teams and the Instrument DevelopingProject of the Chinese Academy of Sciences (Grant No. yz201339).

References

S. Ahadian, S. Yamada, J. Ramón-Azcón, M. Estili, X. Liang, K.Nakajima, H. Shiku, A. Khademhosseini, T. Matsue, J. Ramón-Azcón, M. Estili, X. Liang, K. Nakajima, H. Shiku, A.Khademhosseini, T. Matsue, Hybrid hydrogel-aligned carbon nano-tube scaffolds to enhance cardiac differentiation of embryoid bodies.Acta Biomater. 31, 134–143 (2016)

K. Baumann, Cell adhesion: extracellular bonds. Nat. Rev. Mol. CellBiol. 14, 404 (2013)

E.A. Corbin, L.J. Millet, J.H. Pikul, C.L. Johnson, J.G. Georgiadis, W.P.King, R. Bashir, Micromechanical properties of hydrogels measuredwith MEMS resonant sensors. Biomed. Microdevices 15, 311–319(2013)

A.K. Gaharwar, C. Rivera, C.-J. Wu, B.K. Chan, G. Schmidt,Photocrosslinked nanocomposite hydrogels from PEG and silicananospheres: structural, mechanical and cell adhesion characteris-tics. Mater. Sci. Eng. C 33, 1800–1807 (2013)

A.K. Gaharwar, N.A. Peppas, A. Khademhosseini, Nanocompositehydrogels for biomedical applications. Biotechnol. Bioeng. 111,441–453 (2014)

C. Garcia-Astrain, C. Chen, M. Buron, T. Palomares, A. Eceiza, L. Fruk,M.A. Corcuera, N. Gabilondo, Biocompatible hydrogel nanocom-posite with covalently embedded silver nanoparticles.Biomacromolecules 16, 1301–1310 (2015)

B.M. Gumbiner, Cell adhesion: the molecular basis of tissue architectureand morphogenesis. Cell 84, 345–357 (1996)

L. Han, G. Mapili, S. Chen, K. Roy, Freeform fabrication of biologicalscaffolds by projection photopolymerization. J. Manuf. Sci. Eng.130, 021005 (2008)

M.K. Jaiswal, J.R. Xavier, J.K. Carrow, P. Desai, D. Alge, A.K.Gaharwar, Mechanically stiff nanocomposite hydrogels at ultralownanoparticle content. ACS Nano 10, 246–256 (2015)

P. Kim, D.H. KIM, B. KIM, S.K. Choi, S.H. Lee, A. Khademhosseini, R.Langer, K.Y. Suh, Fabrication of nanostructures of polyethyleneglycol for applications to protein adsorption and cell adhesion.Nanotechnology 16, 2420–2426 (2005)

P. Kim, A. Yuan, K.-H. Nam, A. Jiao, D.-H. Kim, Fabrication of poly(ethylene glycol): gelatin methacrylate composite nanostructureswith tunable stiffness and degradation for vascular tissue engineer-ing. Biofabrication 6, 024112 (2014)

J. LI, A.T.-L. Lam, J.P.W. Toh, S. Reuveny, S.K.-W. Oh, W.R. Birch,Fabrication of uniform-sized poly-ɛ-caprolactone microspheres andtheir applications in human embryonic stem cell culture. Biomed.Microdevices 17, 1–12 (2015)


X. Liu, S. Wang, Three-dimensional nano-biointerface as a new platformfor guiding cell fate. Chem. Soc. Rev. 43, 2385–2401 (2014)

X. Liu, L. Chen, H. Liu, G. Yang, P. Zhang, D. Han, S. Wang, L. Jiang,Bio-inspired soft polystyrene nanotube substrate for rapid and high-ly efficient breast cancer-cell capture. NPGAsiaMater 5, e63 (2013)

C.T.Mierke, The role of focal adhesion kinase in the regulation of cellularmechanical properties. Phys. Biol. 10, 065005 (2013)

Y.M. Mohan, K. Vimala, V. Thomas, K. Varaprasad, B. Sreedhar, S.K.Bajpai, K.M. Raju, Controlling of silver nanoparticles structure byhydrogel networks. J. Colloid Interface Sci. 342, 73–82 (2010)

S.V. Murphy, A. Atala, 3D bioprinting of tissues and organs. Nat.Biotechnol. 32, 773–785 (2014)

S. Nemir, H.N. Hayenga, J.L. West, PEGDA hydrogels with patternedelasticity: novel tools for the study of cell response to substraterigidity. Biotechnol. Bioeng. 105, 636–644 (2010)

R. Peng, X. Yao, J. Ding, Effect of cell anisotropy on differentiation ofstem cells on micropatterned surfaces through the controlled singlecell adhesion. Biomaterials 32, 8048–8057 (2011)

F. Rehfeldt, A.J. Engler, A. Eckhardt, F. Ahmed, D.E. Discher, Cell re-sponses to the mechanochemical microenvironment—implicationsfor regenerative medicine and drug delivery. Adv. Drug Deliv. Rev.59, 1329–1339 (2007)

S. Saxena, M.W. Spears Jr., H. Yoshida, J.C. Gaulding, A.J. Garca, L.A.Lyon, Microgel film dynamics modulate cell adhesion behavior.Soft Matter 10, 1356–1364 (2014)

S. Schmidt, N. Madaboosi, K. Uhlig, D. Köhler, A. Skirtach, C. Duschl,H.Möhwald, D.V. Volodkin, Control of cell adhesion bymechanicalreinforcement of soft polyelectrolyte films with nanoparticles.Langmuir 28, 7249–7257 (2012)

I. Schoen, B.L. Pruitt, V. Vogel, The Yin-Yang of rigidity sensing: howforces and mechanical properties regulate the cellular response tomaterials. Annu. Rev. Mater. Res. 43, 589–618 (2013)

L. Seguin, J.S. Desgrosellier, S.M. Weis, D.A. Cheresh, Integrins andcancer: regulators of cancer stemness, metastasis, and drug resis-tance. Trends Cell Biol. 25, 234–240 (2015)

S.R. Shin, S.M. Jung, M. Zalabany, K. Kim, P. Zorlutuna, S.B. Kim, M.Nikkhah, M. Khabiry, M. Azize, J. Kong, K.T. Wan, T. Palacios,M.R. Dokmeci, H. Bae, X.W. Tang, A. Khademhosseini, Carbon-nanotube-embedded hydrogel sheets for engineering cardiac con-structs and bioactuators. ACS Nano 7, 2369–2380 (2013)

J. Snijder, V.S. Reddy, E.R.May,W.H. Roos, G.R. Nemerow, G.J. Wuite,Integrin and defensin modulate the mechanical properties of adeno-virus. J. Virol. 87, 2756–2766 (2013)

K. Sun, H. Liu, S. Wang, L. Jiang, Cytophilic/cytophobic design ofnanomaterials at biointerfaces. Small 9, 1444–1448 (2013)

Thakur, T., Xavier, J. R., Cross, L., Jaiswal, M. K., Mondragon, E.,Kaunas, R., Gaharwar, A. K. (2016) Photocrosslinkable and elasto-meric hydrogels for bone regeneration. J Biomed Mater Res Part A.

P. Thoniyot, M.J. Tan, A.A. Karim, D.J. Young, X.J. Loh, Nanoparticle-hydrogel composites: concept, design, and applications of thesepromising, multi-functional materials. Adv Sci 2, 1–13 (2015)

L. Wang, H. Liu, F. Zhang, G. Li, S. Wang, Smart thin hydrogel coatingsharnessing hydrophobicity and topography to capture and releasecancer cells. Small 12, 4697–4701 (2016)

Y. Yamahashi, P.J. Cavnar, L.E. Hind, E. Berthier, D.A. Bennin, D.Beebe, A. Huttenlocher, Integrin associated proteins differentiallyregulate neutrophil polarity and directed migration in 2D and 3D.Biomed. Microdevices 17, 1–9 (2015)

W. Yang, H. Yu, F. Wei, G. Li, Y. Wang, L. Liu, Selective pattern ofcancer cell accumulation and growth using UV modulating printingof hydrogels. Biomed. Microdevices 17, 1–8 (2015)

W. Yang, H. Yu, G. Li, B. Wang, Y. Wang, L. Liu, Regulation of breastcancer cell behaviours by the physical microenvironment construct-ed via projection microstereolithography. Biomater Sci 4, 863–870(2016)


Documents

Facile modulation of cell adhesion to a poly(ethylene