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Ann Anat 191 (2009) 126—135

0940-9602/$ - sdoi:10.1016/j.

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www.elsevier.de/aanat

Effects of artificial micro- and nano-structuredsurfaces on cell behaviour

E. Martıneza,�, E. Engelb,c, J.A. Planellb,c, J. Samitiera,d

aNanobioengineering Group, Institute for Bioengineering of Catalonia (IBEC), Josep Samitier 1-5,08028 Barcelona, SpainbBio/Non-Bio Interactions for Regenerative Medicine Group, Institut de Bioenginyeria de Catalunya (IBEC),Josep Samitier 1-5, 08028 Barcelona, SpaincBiomaterials, Biomechanics and Tissue Engineering Group, Department of Materials Science and Metallurgy,Universitat Politecnica de Catalunya, Avda. Diagonal 647, 08028 Barcelona, SpaindDepartment of Electronics, University of Barcelona, c/ Martı i Franques 1, 08028 Barcelona, Spain

Received 1 February 2008; accepted 24 May 2008

KEYWORDSMicrostructure;Topography;Cell behaviour;Cell morphology;Cell orientation

ee front matter & 2008aanat.2008.05.006

ing author.ess: emartinez@pcb.ub

SummarySubstrate topography, independently of substrate chemistry, has been reported tohave significant effects on cell behaviour. Based on the use of fabrication techniquesdeveloped by the silicon microtechnology industry, numerous studies can now befound in the literature analyzing cell behaviour as to various micro- and nano-features such as lines, wells, holes and more. Most of these works have been found torelate the micro- and nano-sized topographical features with cell orientation,migration, morphology and proliferation. In recent papers, even the influence ofsubstrate nanotopography on cell gene expression and differentiation has beenpointed out. However, despite the large number of papers published on this topic,significant general trends in cell behaviour are difficult to establish due todifferences in cell type, substrate material, feature aspect-ratio, feature geometryand parameters measured. This paper intends to compile and review the relevantexisting information on the behaviour of cells on micro- and nano-structuredartificial substrates and analyze possible general behavioural trends.& 2008 Elsevier GmbH. All rights reserved.

Elsevier GmbH. All rights rese

.es (E. Martınez).

Introduction

In recent years, the interest in basic knowledgeon cell–substrate interaction has grown increas-ingly as it has now been recognized to play a keyrole in the differences observed in cell behaviour

rved.

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when comparing in vitro and in vivo culturing(Flemming et al., 1999). This thus represents acrucial factor in the fields of tissue engineering,drug development and regenerative medicine.

Cells in their natural environment are surroundedby nanostructures, when contacting with othercells (membranes have nano-size features) or withthe extra-cellular matrix (ECM), formed by biomo-lecules configured in different geometrical ar-rangements (nanopores, nanofibers, nanocrystals).Cell behaviour is determined via intrinsic cellsignals, but also via extrinsic cell signals comingfrom the cell–cell contact and the cell–ECMcomponents. These signals may be chemical(growth factors such as cytokines) or mechanical(tensile forces caused by the cell interacting withmicro- or nano-structured surfaces). For example,mechanical stress has been found to affect thestrength of the integrin–cytoskeleton links and theintegrin receptor distribution and conformation,thus activating intracellular pathways active in celldevelopment and behaviour (Chen et al., 1997;Clark, 1995).

The ability of the substratum to influence cellorientation, migration and cytoskeletal organiza-tion was first noted by Harrison in 1911 when hegrew cells on a spider web and the cells followedthe fibers of the web in a phenomenon calledstereotropism or physical guidance (Harrison,1911). Later, in 1964, it was first proposed thatcells react to the topography or their environment(Curtis and Varde, 1964). Since then, and thanks tothe various micro- and nano-fabrication techniquesdeveloped in the silicon microelectronics industry(Chen and Pepin, 2001), numerous studies haveshown that many cell types react strongly tomicrotopography (Hasirci and Kenar, 2006; Flem-ming et al., 1999; Curtis et al., 2006). Changes incell adhesion (Matsuzaka et al., 2003; Recknoret al., 2004), alignment (Clark et al., 1987, 1990;Recknor et al., 2004), morphology (cytoskeletalorganization) (Wojciak-Stothard et al., 1995; Flem-ming et al., 1999), proliferation (Keselowsky et al.,2007), vitality (Chen et al., 1997) and geneexpression and differentiation (Bruinink and Win-termantel, 2001; Watt et al., 1988) have beenreported.

Recently, it has also been shown that nanotopo-graphy can also have strong effects on a range ofcell types. Some articles have reported a decreasein cell attachment and focal adhesion complexes innanostructures (Dalby et al., 2004a; Curtis et al.,2004), while cell morphology, proliferation andspreading (checked by cytoskeleton development)are also decreased in these nano-structured sur-faces (Gallagher et al., 2002; Dalby et al., 2004b;

Andersson et al., 2003). The nanostructures havealso been reported to largely alter gene expression(Andersson et al., 2003) and, in some instances,promote or direct cell differentiation (Dalby et al.,2006, 2007; Popat et al., 2007).

Mimicking the random structure of the ECM bynano-fabrication techniques has provided valuableinformation on cell behaviour. The use of multi-walled carbon nanofibers (�100 nm diameter)produced by chemical vapour deposition has re-vealed that osteoblasts seeded on them proliferatemuch better than those grown on flat glass surfaces(Elias et al., 2002). Likewise, alkaline phosphataseactivity, an indicator of osteoblastic bone forma-tion, was also increased on these substrates. Thisindicates that specialized cellular functions may beenhanced when cells grow on a substrate closelymatching the physical topology of the ECM.Additionally, fibroblasts and kidney epithelial cellsseeded onto nanofibers have fewer stress fibers andsmaller focal adhesions than those seeded on glasscoverslips (Schindler et al., 2005).

Despite the accepted idea that the role ofsubstrates is more than that of merely providingmechanical support but that they actually act asintelligent surfaces capable of providing chemicaland topographical signals to guide cell adhesion,spreading, morphology, proliferation and, even-tually, cell differentiation (Wilkinson et al., 2002;Curtis et al., 2006), there is a clear lack ofsystematic studies and just a very few reviewssummarizing these effects (Flemming et al., 1999).The purpose of this paper is, then, to review thetechniques used to fabricate artificial surfaces withmicro- and nano-topographies and the results foundin the literature about the effects of these micro-and nano-features on cell behaviour.

Fabrication of substrates with micro- andnano-topographies

The definition of micro- and nano-structures onthe substrates relies on lithographic methods.Usually, a computer-designed pattern is exposedby means of light, electrons, ions or imprinted.The lithography is carried out on a special lightor electron-sensitive material or imprinted intoa special deformable polymer, which is thenused in subsequent pattern transfer processesas a mask, or, alternatively, used for cell culturingas it is.

The most standard lithography technique is UVlithography, which uses photocurable resists sensi-tive to the UV radiation. A transparent polymer, or

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a metal-coated glass slide, with the desiredfeatures can be used as a mask through which thephotoresist can be irradiated. The photoresist isthen developed and the structures transferred tothe substrate. This patterned resist layer can thenact as a mask for the substrate etching, either bywet etching processes or by dry etching techniques(Madou, 1997; Thompson et al., 1995). The resolu-tion of this method is about 2 mm.

More recent lithographic methods, based onparticle beam, have led to the fabrication ofnano-sized topographical features. The focusedion beam lithography technique uses an energeticion beam (Ga ions for example) that is acceleratedand focused on the sample, thus producing colli-sions with the atoms of the surface and resulting inthe etching of these atoms (Langford, 2002). Theion beam can be controlled to scan the sample inthe shape of the desired patterns with a maximumlateral resolution of about 20 nm. This is a directwriting process that does not require masks.Electron beam lithographic processes consistof using an electron beam to scan the samplesurface, which has been coated with an electron-sensitive polymer (Marrian and Tennant, 2003). Thepolymer is affected by the beam and acts as anetching mask after development. This procedure,then, requires masks and etching procedures, but ithas an excellent lateral resolution of less than10 nm.

Very recently, fast parallel replication techni-ques, such as nano-imprint lithography (Chou et al.,1995), have made it possible to overcome thedifficulties of particle beam techniques (too slowand too expensive), thus allowing the applicationof nano-structured polymeric materials to cellculture (Mills et al., 2005). Materials, such asthermoplastic polymers (polymethylmethacrylate)or UV curable materials (acrylates, epoxies, etc.),can be micro/nanostructured by using these tech-niques, which require the use of a mould with thepattern to be transferred to the polymer. Themould is placed in contact with the polymer surfaceand both are heated and pressed. When thetemperature surpasses the glass transition tem-perature of the polymer (Tg), the polymer startsflowing and the applied pressure forces it to fill thegaps in the mould. The system is finally cooled, andthe mould and the replica can be separated.Imprinting is a quick and easy fabrication methodin which a single mould can be used to produceseveral replicas.

Other techniques for the fabrication of micro-and nano-structured substrates suitable for cellculture include laser deposition and etching, softlithography and colloidal lithography.

Effects of micro- and nano-topographicalstructures on cell orientation andadhesion

A list of the most important results obtained oncell orientation and adhesion when using micro-and nano-structured surfaces of different materialsis presented in Table 1. It may be noted that a verywide range of cell types such as fibroblasts,osteoblast, nerve cells and mesenchymal stemcells respond profoundly to a groove substrate(Wojciak-Stothard et al., 1995; Wood, 1988; Meyleet al., 1995; Clark et al., 1990; Charest et al.,2004; Miller et al., 2001; Baac et al., 2004; Recknoret al., 2004). Cells seeded onto artificially pro-duced micro- and nano-grooves aligned their shapeand elongated in the direction of the groove(Figure 1b) in almost all cases, although the degreeof morphology depends on cell type, on groovedepth and, to a lesser extent, on groove width(Clark et al., 1990; Dalby et al., 2003; Flemminget al., 1999; Teixeira et al., 2003, 2004). Indeed,for grooves of period greater than 20 mm, no celltype (except red blood cells) has been found thatresponds (Wilkinson et al., 2002). Cell alignment tothe long axis of the groove is accompanied byorganization of actin and other cytoskeletalelements in an orientation parallel to the grooves.Actin and microtubules align along walls and edges,the microtubules being the first element to bealigned, followed by actin (Oakley and Brunette,1993). Orientation often increases with increasingdepth, but decreases with increasing groove width(Clark et al., 1990). It is remarkable, however, thatthe general trend of this parallel alignmenttendency is interrupted in the case of the experi-ments published by Teixeira et al. (2006). In thisstudy, epithelial cells switched from parallel toperpendicular alignment to the grooved sub-strate when features decreased in pitch size from4000 to 400 nm, also changing their focal adhesiondistribution.

Although almost all sorts of cells have been foundto align with respect to grooves, other topographi-cal features as wells, pits or pillars do not showsuch a clear cell alignment for all cell types(Gallagher et al., 2002; Fewster, 1994; Huntet al., 1995). However, studies of Curtis’s groupseem to point out that cells can ‘‘feel’’ symmetriesin the topography (Figure 1a), especially if they arein the nanorange (Curtis et al., 2004).

Regarding cell adhesion, there is not a commontrend for the various cell types or micro/nanos-tructures assayed. To illustrate this fact, Table 1shows that cell adhesion has been reported to be

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Table 1. Summary of the effects of micro- and nano-structured surfaces on cell orientation and adhesion

Feature type Material Widtha/diameter

Depth/height Pitch Cell type Effects on orientation and adhesion Reference

Grooves Quartz 0.5, 5, 10,25mm

0.5, 5 mm 1:1 Murine P388D1macrophage

Orientation Wojciak-Stothardet al. (1995)

Grooves Quartz 1, 4mm 1.1 mm 1:1 Mesenchymalstem cells

Alignment better in the widestgrooves

Wood, (1988)

Grooves Silicon dioxide 0.5 mm 1mm 1:1 Fibroblasts Strong alignment Meyle et al. (1995a)Grooves Silicon dioxide 0.5 mm 1mm 1:1 Keratinocytes No alignment Meyle et al.

(1995b)Grooves PMMA 2, 3, 6, 12mm 0.2, 0.5, 1.1,

1.9 mm1:1 BHK cells Alignment increased with d. and

decreased with w.Clark et al. (1990)

Grooves Photo-responsivePMMA

1mm 250nm 1:1 Humanastrocytes(HAs)

Improved adhesion, strongalignment

Baac et al. (2004)

Grooves PS 1–10 mm 0.5–1.5 mm N/D Rat bone cells Large grooves: focal adhesions allover the surface.

Matsuzaka, et al.(2003)

Narrow grooves: only on the edgesGrooves Polyimide 4mm 5mm 34mm Osteoblasts Strong alignment, no changes in

adhesionCharest et al.(2004)

Grooves PDLA 10mm 3mm 20mm Schwann cells(nerve cells)

Strong alignment Miller et al. (2001)

Grooves PS 10mm 3mm 20mm Rat astrocytes Less adhesion, strong alignment Recknor et al.(2004)

Grooves PS 20–1000 nm 5–530 nm 1:1 Fibroblasts No alignment for depthso35 nm orwidthso100 nm

Loesberg et al.(2007)

Grooves Silicon 330–2100 nm 600nm 400–4000 nm Human cornealepithelial cells

Perpendicular alignment for400–800 nm pitch. Parallel for1600–4000 nm

Teixeira et al.(2006)

Steps PMMA 1–18 mm N/D N/D BHK Alignment at steps Clark et al. (1987)Wells PC 7, 25, 50mm 0.5, 1.5, 2.5 mm 1:1 Fibroblasts No orientation Hunt et al. (1995)Pits PCL 150 nm 80nm 300nm Fibroblasts Less focal contacts and vinculin

patternGallagher et al.(2002)

Pits PCL, PMMA 35, 75 and120 nm

N/D 100, (200,300 nm

Fibroblasts Reduced adhesion, orientation anddistinction of symmetries

Dalby et al.(2004a), Curtis etal. (2001), Curtis etal. (2004)

Random PLGA, PU, PCL 206, 370 nm N/D N/D Bladder smoothmuscle cells

Enhanced adhesion Thapa et al. (2003)

Abbreviation list: N/D: non-determined; PMMA: poly(methylmethacrylate); PDMS: poly-dimethyl siloxane; PC: polycarbonate; PS: polystyrene; PLLA: poly(L-lactide acid); PET:poly(ethylene terephthalate); PBrS: poly(4-bromostyrene); PCL: polycaprolactone; PDLA: poly(D,L-lactic acid); PLGA: polylactic-co-glycolic-acid; PU: poly-ether-urethane.aThe width parameter refers to the groove width.

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Figure 1. Scanning electron microscopic (SEM) images of rat mesenchymal stem cells cultured on (a) non-symmetricPMMA structured surface, feature size 2mm and (b) PMMA surface structured with 2 mm wide lines.

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reduced with nano-structured surfaces for somecell types (fibroblasts in Gallagher et al., 2002),and to be improved for other cell types (musclecells in Thapa et al., 2003 and astrocytes in Baacet al., 2004). Recently, the work of Prof. J.P. Spatzhas demonstrated that cells adhere and spread wellin surfaces with a topography of 58 nm features andthat they do not adhere to nanotopographies of73 nm in size (Cavalcanti-Adam et al., 2007), thusshowing the close relationship between integrinpositioning and nanotopography that could explainthe variations in the results encountered in theliterature.

Effects of micro- and nano-topographicalstructures on cell morphology

In nearly all instances, grooves produce someelongation and nuclear polarization in the morphol-ogy of all sorts of cells (Wojciak-Stothard et al.,1995; Meyle et al., 1993; Matsuzaka et al., 2003;Andersson et al., 2003; Charest et al., 2004) (seesummary in Table 2). The cell membrane conformsto the largest grooves but tends to bridge thenarrowest and deepest grooves (Matsuzaka et al.,2003). Other geometrical features (wells, pits,pillars) with dimensions of less than approximately5 mm, including nano-sized features, seem to leadto smaller, rounded cells with less organizedcytoskeletons. This result is independent of thecell type for the cells assayed (Hunt et al., 1995;Gallagher et al., 2002; Dalby et al., 2002, 2004b;Andersson et al., 2003). An example can be shownin Figure 2, comparing actin filaments developed byrat mesenchymal stem cells cultured on asym-metric 2 mm structures with the well-developed

cytoskeleton developed by the same cells culturedon 50 mm ring structures. This behaviour seems tobe related to the fact that the basement cellmembrane in contact with the nano-structuredsurface will suffer tensile and relaxation mechan-ical forces that will rearrange its components, suchas integrin complexes and thus signalling cellmorphology.

Effects of micro- and nano-topographicalstructures on cell proliferation anddifferentiation

The effects of micro- and nano-structures on cellproliferation are highly diffuse and dependant onthe cell type and structures involved (Dalby et al.,2004c; Miller et al., 2004; Green et al., 1994) (seesummary in Table 3). The reason for this lack ofgeneral trends could be the absence of systematicstudies on this issue, as most of the reports found inthe literature dealing with micro- and nano-structures and cell culture have been written bytechnology groups and are focused on microscopyobservations. As a consequence, cell proliferationis a parameter that is not always measured.Moreover, most of the experiments have beenperformed with very robust and proliferative celllines (fibroblasts, osteoblasts, carcinoma lines) andthis fact could bias a possible extrapolation of thereported results.

The authors have found only a single report(Gerecht et al., 2007) dealing with the effectsof nanotopography on human embryonic stemcells. Results have shown that human ESCs dorespond to nano-scale substrate topography in amanner similar to other terminally differentiated

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Table 2. Summary of the effects of micro- and nano-structured surfaces on cell morphology

Feature type Material Widtha/diameter

Depth/height Pitch Cell type Effects on morphology Reference

Grooves Quartz 0.5, 5, 10,25 mm

0.5, 5mm 1:1 Murine P388D1macrophage

Elongation, more in widergrooves

Wojciak-Stothard et al.(1995)

Grooves Epoxy 0.5 mm 1 mm 1:1 Human fibroblasts Cytoskeleton oriented withgrooves

Meyle et al.(1993)

Grooves PS 1–10 mm 0.5–1.5 mm N/D Rat bone cells W 4 5mm: cells followed thesurface. Narrow grooves: cellsbridge

Matsuzaka, et al.(2003)

Grooves Polyimide 4 mm 5 mm 34 mm Osteoblasts Elongation Charest et al.(2004)

Grooves Ti-coated Si 15 mm 200 nm No symmetry T24, human bladdercarcinoma

Nanopillars: less round andmore stellate, smaller

Andersson et al.(2003)

Pillars 110 nm 1:1 Grooves: elongationWells PC 7, 25, 50 mm 0.5, 1.5,

2.5 mm1:1 Fibroblasts No elongation Hunt et al.

(1995)Pits PCL 150 nm 80 nm 300 nm Fibroblasts F-actin cytoskeleton less

developedGallagher et al.(2002)

Pillars PMMA 100 nm 160 nm N/D Fibroblasts Smaller, less organized actincytoskeleton

Dalby et al.(2004b)

Random PS and PBrS 13, 35, 95 nm N/D N/D Human endothelial Round cells on PS, Arcuatemorphology largest for the13 nm islands

Dalby et al.(2002)

Abbreviation list: N/D: non-determined; PMMA: poly(methylmethacrylate); PDMS: poly-dimethyl siloxane; PC: polycarbonate; PS: polystyrene; PLLA: poly(L-lactide acid); PET: poly(ethyleneterephthalate); PBrS: poly(4-bromostyrene); PCL: polycaprolactone; PDLA: poly(D,L-lactic acid); PLGA: polylactic-co-glycolic-acid; PU: poly-ether-urethane.aThe width parameter refers to the groove width.

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Figure 2. Confocal microscopic images of rat mesenchymal stem cells cultured on PMMA microstructured substrates (a)non-symmetric 2 mm structures and (b) 50 mm rings. Cell nuclei are inmunostained in blue and actin fibers in red.

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mammalian cells, thus demonstrating cell elonga-tion and alignment, changes in the organization andpolarization of the cytoskeleton proteins andreduced proliferation.

Although a decrease in cell proliferation is themost frequent result of the few papers found in theliterature, also works claiming a high increase incell proliferation with the nanostructured datacould be found (Popat et al., 2007).

Similarly, changes in gene expression have beenobserved (Dalby et al., 2003; Andersson et al.,2003; Wojciak-Stothard et al., 1996; Matsuzakaet al., 1999) but no systematic study has beencarried out until now. In fact, there are only a fewreports studying the effect of controlled topogra-phy on stem cells and even fewer presenting resultson the effects on cell differentiation. However, thepublished data show an effect of nanotopographyon cell commitment. Dalby et al. (2006, 2007) hasshown that random circular nanostructures pro-mote and direct osteoblast differentiation ofmesenchymal stem cells without the need ofosteogenic promoter cell culture medium andPopat et al. (2007) has reported an increase inthe alkaline phosphatase activity and matrixproduction of marrow stromal cells when theyare cultured on nanoporous alumina substrates.In fact, it has also been published that cellshape regulates the switch in the lineage commit-ment of human mesenchymal stem cells (hMSC)by modulating endogenous RhoA activity. WhenhMSC are allowed to adhere, flatten and spread,they undergo osteogenesis, whereas unspread,round cells become adipocytes (McBeath et al.,2004). Apparently, changes in cell spreadingalter RhoA-mediated cytoskeletal contractility,

focal adhesion assembly and downstream integrinsignalling.

Conclusion

Substrate micro- and nano-topography, indepen-dently of substrate biochemistry, seem to havesignificant effects on cell behaviour. As reported inthe studies summarized here, substrate topography(with micro- or nano-features) has direct effects oncell orientation, morphology and cytoskeletonarrangements. As a general rule, the effects aremore pronounced for smaller, nano-sized surfacefeatures, although there is a lack of knowledge onthe basic mechanisms the cell uses to detect andrespond to this nanotopography. Effects of sub-strate topography on cell proliferation and differ-entiation cannot yet be established, due to thepaucity of studies, sometimes with contradictoryresults, found in the literature. Recent publicationsseem to reveal that the effect of the surfacegeometry is not just due to the changes in theamount of adsorbed proteins (nanotopographygreatly enhance the surface area) (Curtis et al.,2006). On the contrary, the basement cell mem-brane in contact with the nano-structured surfacewill suffer tensile and relaxation mechanical forcesthat will rearrange its components and/or open ionchannels that will trigger cell behaviour. Furtherstudies focused in the deep insight between thesubstrate topography and the cell signalling that itgenerates would be extremely helpful in contribut-ing to the development of cellular environmentscommitting desired cell responses.

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Table 3. Summary of the effects of micro- and nano-structured surfaces on cell proliferation and/or differentiation

Featuretype

Material Widtha/diameter

Depth/ height Pitch Cell type Effects on proliferation and/or differentiation

Reference

Grooves PS Micro (manual) N/D N/D Rat bone marrow cells.Osteblasts MC3T3

RBMC influenced by grooves(osteoblast differentiation).MC3T4 not influenced

Bruinink and Wintermantel(2001)

Grooves Quartz 12.5 mm 5 mm 1:1 Fibroblasts Gene expression largelychanged

Dalby et al. (2003)

Grooves Quartz 2–10 mm 30–280 nm N/D Murine macrophages Higher phagocytotic activitywhen topographysize ¼ collagen fiber

Wojciak-Stothard et al. (1996)

Grooves PLLA, PS 1, 2, 5, 10 mm 0.5, 1, 1.5mm 1:1 Rat bone cells Better mineralization with d.1mm and w. 1–2mm

Matsuzaka, et al. (1999)

Grooves PDMS 600 nm 600 nm 1:1 Human embryonic stem cells Reduced proliferation Gerecht et al. (2007)Grooves Ti-coated Si 15 mm 200 nm 1:1 Asymmetry T24, human bladder

carcinomaLess production of cytokinesin nanopillar surfaces

Andersson et al. (2003)

Pillars 110 nmPores Alumina 72 nm N/D N/D Mouse marrow stromal cells Proliferation increased 45%,

increased osteoblastdifferentiation

Popat et al. (2007)

Wells PDMS 2, 5, 10 mm N/D 1:1 Human fibroblasts 2 and 5mm betterproliferation. 10 mm no effect

Green et al. (1994)

Wells PC 7, 25, 50 mm 0.5, 1.5,2.5 mm

1:1 Fibroblasts No effects Hunt et al. (1995)

Wells,random

PMMA 120 nm 100 nm 300 nm Human mesenchymal stemcells

Stimulated differentiationand production of bonemineral in vitro

Dalby et al. (2007)

Pillars PMMA 100 nm 160 nm N/D Fibroblasts Less spreading Dalby et al. (2004c)Beads Silica on PEI-

coated siliconGradientconcentration

73 nm 73 nm 73 nm Rat calvarial osteoblasts Particles (nanotopography)reduced cell proliferation

Kunzler et al. (2007)

Random PLGA ? nano-range Nano-range N/D Rat aortic smooth musclecells, rat aortic endothelialcells

Improve on cell densities withthe nanostructure

Miller et al. (2004)

Random PMMA 2mm–100 nm 3–10 nm N/D Bone marrow cells (stemcells)

Differentiation to osteoblastspromoted

Dalby et al. (2006)

Circles nonordered

Abbreviation list: N/D: non-determined; PMMA: poly(methylmethacrylate); PDMS: poly-dimethyl siloxane; PC: polycarbonate; PS: polystyrene; PLLA: poly(L-lactide acid); PET: poly(ethyleneterephthalate); PBrS: poly(4-bromostyrene); PCL: polycaprolactone; PDLA: poly(D,L-lactic acid); PLGA: polylactic-co-glycolic-acid; PU: poly-ether-urethane.aThe width parameter refers to the groove width.

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Acknowledgements

This paper and the work it concerns weregenerated in the context of the CellPROM project,funded by the European Community as Contract no.NMP4-CT-2004-500039 and it reflects only theauthors’ views. The authors are also grateful tothe Spanish Ministry of Science and Education (MEC)for support through project TEC2004-06514-C03,and for the provision of grants through the Ramon yCajal (EM).

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