Topographical Patterning Research Articles

Topographically and elastically micropatterned PEG-based hydrogels to control cell adhesion and migration

Cell migration is a primary process implicated in several biological processes such as immune response, metastasis or wound healing. Tissue cells respond to, among others, topographic and elastic cues of the extracellular environment. In this work, the migratory behaviour of mouse fibroblasts was investigated on poly(ethylene glycol) hydrogels (PEG) where only one triggering signal was present at a time, i.e. either topographic structures or a stiffness contrast. Despite the intrinsically anti-adhesive properties of PEG polymers, previous experiments in our group have demonstrated that the presence of topographic and mechanical patterns on the surface of those materials do induce fibroblast adhesion. These findings were corroborated in this study. Furthermore, the migration behaviour was investigated. Channel-like structures were prepared by the cast-molding method. We showed that the orientation of the migration tracks with the pattern increased by decreasing the width of the channels; cells migrated randomly inside wide channels, while on narrow ones, they migrated parallel to the direction of the pattern. Besides, biphasic mechanical patterns were created combining polymers with two different degrees of cross-linking with our novel soft-lithographic technique: the Fill-Molding In Capillaries method (FIMIC). In this method, a topographically patterned hydrogel (mold) was filled with the secondary material (filler) via capillary force. This methodology was applied using either pure PEG or a blended material (Blend) of PEG and a block co-polymer containing poly(propylene glycol). Fibroblasts adhered preferentially to the mold lines of the pattern, even on those samples where the mold lines were softer, contradicting the expected durotactic behaviour. Further atomic force microscopy (AFM) analysis of the surface properties of the substrates to explain this phenomenon brought interesting results. The topographic landscape displays convex and concave structures, the former of which being apparently more attractive for the fibroblasts to climb onto and migrate along. On those samples were this topography was detected, the fibroblasts responded to it, while when the sample was practically smooth, they reacted to the elastic pattern. Thus, we demonstrated that the presence of geometric patterns can overrule elastic patterns, helping to elucidate the hierarchy behind the external migratory signals.

Simultaneous topographic and chemical patterning via imprinting defined nano-reactors

A novel, universal strategy to realize simultaneous topographic and chemical patterning via imprinting defined nano-reactors.

Novel soft stamp development for direct micro- and nano-patterning of macroscopic curved surfaces

Surface topographical patterning is a simple way to functionalize surfaces without changing material chemistry. Topographical patterning of nonplanar surfaces has remained a challenge, despite sought after applications in microfluidics, optics, and biomedical technologies. Here the authors develop transparent, reusable soft molds that allow facile micro- and nanopatterning of macroscopically curved surfaces. The authors use bilayer molds with a soft backing and a hard pattern carrying layer to overcome challenges that arise from the opposing need for mold compliance (to allow conformal contact with nonflat substrates) and rigidity (to maintain patterned feature resolution and fidelity). With our approach, high yield curved surface patterning (>98%) over large (2 × 1 cm) area can be effectively achieved. Structure replication down to 80 nm resolution is demonstrated.

Characterization tools for polymer thin films

The nanotechnology has emerged as an effective tool to fabricate next-generation microelectronics, biologically responsive materials, and structured membranes. The self-assembly of nanoscale phases has extensively been studied in thin films because of their potential applications in sub-100 nm structures. The control of the ordering of nanaoscale patterns is critical for various technological applications. A variety of approaches such as topographical and chemical patterning have resulted in an enhancement in long-range orders of nanoscale patterns. The macroscopically large areas of nanoscale domains with single-crystal order in polymer thin films can be utilized to fabricate portable ultra-high density data storages, advanced sensors and ultra-light electronic devices. However, as pattern size decreases below 100 nm, there appear many new challenges such as the cost of patterning and the precise control of the line edge roughness and line width roughness. Precisely controlling nanostructure shapes and placements in material is a continuing challenge. Measurement platform to provide accurate and detailed information about nanostructure orientations and placements is a key to this challenge. In this review, we examine the recent progress of characterization tools in polymer thin films. We highlight our efforts to control surface pattern formations of polymer thin films and our use of statistically-useful scattering techniques and real-space imaging tools to quantify the order of nanoscale patterns. In some technological applications of biological membranes, such as chemical separations, drug delivery and sensors, the orientation distribution of nanostructures is often more important. The real-space imaging methods of characterizing the orientation distribution of nanostructures, such as cross-sectional electron microscopy measurements and depth profiling by alternating etch and surface imaging steps are readily performed on thin polymer films over large areas. However, these real-space imaging techniques are destructive measures of nanostructures in polymer thin films. Also it is challenging to in-situ measure the evolution of orientation of nanoscale patterns during processing by using these destructive real-space imaging techniques. Rotational small-angle neutron scattering (RSANS) and grazing-incidence small-angle x-ray scattering (GISAXS) are effective and non-destructive measurement tools to measure the evolution of orientation distribution of nanoscale patterns during processing. In this rotational small angle neutron scattering method, the sample is rotated in the neuron beam. By accumulating the scattering density at each sample rotation angle, the three-dimensional Fourier space of the internal ordering in the nanostructured film can be mapped. By using this relatively new rotational small angle neutron scattering method and established models for nanoscale patterns, the full three-dimensional orientation distribution of nanoscale patterns can be obtained.

Thickness and Microdomain Orientation of Asymmetric PS-b-PMMA Block Copolymer Films Inside Periodic Gratings.

The ordering process of asymmetric PS-b-PMMA block copolymers (BCPs) is investigated on flat SiO2 surfaces and on topographically patterned substrates. The topographic patterns consist of periodic gratings of 10 trenches defined by conventional top-down approaches and subsequently neutralized using a P(S-r-MMA) random copolymer (RCP). When the ordering process is accomplished on a flat surface at a temperature ranging between 180 and 230 °C, cylindrical microdomains perpendicularly oriented with respect to the substrate are observed irrespective of annealing temperature. In contrast, when the ordering process occurs on topographically patterned substrates, different phenomena have to be considered. The simultaneous effect of the flow around the gratings and the BCP flux from the zone located between adjacent trenches (mesa) into the inner part of the trenches results in significant thickness variations of the confined BCP film. Therefore, the amount of BCP inside the trenches depends on the width of the mesa region, which acts as a BCP reservoir. Moreover, within each trench group, the BCP thickness progressively decreases from the external to the central trenches composing the periodic grating. The thickness variation of the BCP film within the trenches strongly affects the ordering process, ultimately leading to different orientations of the microdomains within the trenches. In particular, when the annealing temperature is 190 °C a precise confinement of the BCP within the trenches featuring a perpendicular cylinder morphology is observed. At higher temperatures, mixed or parallel orientations of the microdomains are obtained depending on the width of the trenches in the periodic grating.

Neuronal Alignment and Outgrowth on Microwrinkled Conducting Polymer Substrates

ABSTRACTWe report on the results of culturing SH-SY5Y neuron-like cells on PEDOT:PSS wrinkled surfaces fabricated by thermally-induced shrinking of commercial polystyrene sheets. Such smart biointerfaces combine the functional properties of conducting polymers with the topographic patterning at the micro- and sub-microscale, as a result of surface wrinkling. By imposing mechanical constraints during shrinking, anisotropic topographic features are formed, with a spatial periodicity in the range 0.7 - 1.2 μm, tunable by varying the thickness of the PEDOT:PSS thin film. The effectiveness of wrinkled surfaces in enhancing and orientating the outgrowth of neurites is demonstrated by a 42% increase in length and by the 85% of neurites aligned along wrinkles direction (angle 0 < θ < 15°), after 5 days of differentiation. Furthermore, the conductive properties of the PEDOT:PSS film are retained after the surface wrinkling, opening the way for the exploitation of these smart biointerfaces for the electrical stimulation of cells.

Modeling depletion mediated colloidal assembly on topographical patterns

This work reports a model and Monte Carlo simulations of excluded volume mediated interactions between colloids and topographically patterned substrates in the presence of thermosensitive depletants. The model is matched to experiments to yield density, free energy, and potential energy landscapes that quantitatively capture particle microstructures varying from immobilized non-close packed configurations to random fluid states. A numerical model of local excluded volume affects is developed to enable computation of local depletion attraction in the presence of arbitrary geometries. Our findings demonstrate a quantitative modeling method to interpret and predict how surface patterns mediate local depletion interactions, which enables the design of colloidal based materials and devices.

Neuropeptide Y immunoreactivity in the cat claustrum: A light- and electron-microscopic investigation

The claustrum is a telencephalic nucleus located ventrolateral to the basal ganglia in the mammalian brain. It has an extensive reciprocal connectivity with most if not all of the cerebral cortex, in particular, primary sensory areas. However, despite renewed and growing interest amongst investigators, there remains a paucity of data concerning its peptidergic profile. The aim of the present study was to examine the presence, morphology, distribution and ultrastructure of neuropeptide Y-immunoreactive (NPY-ir) neurons and fibers in the claustrum of the cat. Ten adult healthy cats from both sexes were used. All animals received human and ethical treatment in accordance with the Principles of Laboratory Animal Care. Subjects were irreversibly anesthetized and transcardially perfused with fixative solution containing glutaraldehyde and paraformaldehyde. Brains were promptly removed, postfixed and sectioned. Slices were incubated with polyclonal anti-NPY antibodies according to the standard avidin–biotin–peroxidase complex method adopted by our Department of Anatomy, Histology and Embryology. NPY-ir neurons and fibers were found to be diffusely distributed throughout the claustrum, with no obvious topographic or functional patterning other than larger numbers in its central/broadest part (stereotaxic planes A12–A16). Neurons were generally classified by diameter into three sizes: small (under 17μm), medium (17–25μm) and large (over 25μm). Staining density is varied with some neurons appearing darker than others. At the electron-microscopic level NPY immunoproduct was observed within neurons, dendrites and terminal boutons, each differing relative to their ultrastructural attributes. Two types of NPY-ir synaptic boutons were found. Lastly, it is of interest to note that gender-specific differences were not observed.

Enigmatic cranial superstructures among Chamorro ancestors from the Mariana Islands: gross anatomy and microanatomy.

This study focuses on the gross anatomy, anatomic relations, microanatomy, and the meaning of three enigmatic, geographically patterned, and quasi-continuous superstructures of the posterior cranium. Collectively known as occipital superstructures (OSSs), these traits are the occipital torus tubercle (TOT), retromastoid process (PR), and posterior supramastoid tubercle (TSP). When present, TOT, PR, and TSP develop at posterior cranial attachment sites of the upper trapezius, superior oblique, and sternocleidomastoid muscles, respectively. Marked expression and co-occurrence of these OSSs are virtually circumscribed within Oceania and reach highest recorded frequencies in protohistoric Chamorros (CHamoru) of the Mariana Islands. Prior to undertaking scanning electron microscopy (SEM) work, our working multifactorial model for OSS development was that early-onset, long-term, and chronic activity-related microtrauma at enthesis sites led to exuberant reactive or reparative responses in a substantial minority of genetically predisposed (and mostly male) individuals. SEM imaging, however, reveals topographic patterning that questions, but does not negate, activity induction of these superstructures. Although OSSs appear macroscopically as relatively large and discrete phenomena, SEM findings reveal a unique, widespread, and seemingly systemic distribution of structures over the occipital surface that have the appearance of OSS microforms. Nevertheless, apparent genetic underpinnings, anatomic relationships with muscle entheses, and positive correlation of OSS development with humeral robusticity continue to suggest that these superstructures have potential to at once bear witness to Chamorro population history and inform osteobiographical constructions of chronic activity patterns in individuals bearing them. Further work is outlined that would illuminate the proximate and ultimate meanings of OSS.

Generation of hierarchical nano- and microwrinkle structure for smooth muscle cell alignment

On the contrary to the complicated and high-cost photolithography based topographical patterning, the non-lithographical strain responsive wrinkling method can generate a variety of wrinkle structures with ease, high uniformity, and cost-effectiveness. The strain responsive wrinkling approach relied on the modulus difference between a thin and stiff film and a soft substrate, resulting in a periodic out-of-plane buckling deformation upon the release of the compressive stress. While the previous reports were dedicated to the micropattern generation, we, in this study, investigated the effect of the UV/O exposure time (10, 20, 30, 40, 50, and 60 min) and the stretching rate (10, 20, 30, and 40%) of the PDMS substrate on the wavelength and the amplitude of the wrinkle patterns in details. Sole nanowrinkle as well as hierarchical nano/microwrinkle patterns could be fabricated through the fine control of those factors. Furthermore, we examined the human aortic smooth muscle cell (HASMCs) alignment on the topographical patterned surface, and found that more than 80% of the HASMCs were cultured and aligned well along with the hierarchical nano/microwrinkle rather than the nanowrinkle pattern.

Migration speed and directionality switch of normal epithelial cells after TGF-β1-induced EMT (tEMT) on micro-structured polydimethylsiloxane (PDMS) substrates with variations in stiffness and topographic patterning

The epithelial to mesenchymal transition (EMT) involves several physiological and pathological phenomena and endows cells with invasive and migratory properties. However, the effects of substrate stiffness and topography on the migration of cells before or after transforming growth factor-β1 (TGF-β1)-induced EMT (tEMT) are unknown. Herein, we seed control or tEMT NMuMG cells on the 2D patterns consisted of 1 μm or 5 μm line-widths and groove or cone patterns on either 2 MPa (1.96 ± 0.48 MPa) or 4 MPa (3.70 ± 0.74 MPa) polydimethylsiloxane (PDMS) substrates. After tEMT, the increased expression of α-SMA with vinculin in focal adhesion (FA) sites led to an acceleration of tEMT cell motility. On the 2 MPa substrate, the most influenced substrate was the 1 μm, cone-patterned substrate, where the tEMT cells' motility decelerated by 0.13 μm/min (36% slower than the cells on groove pattern). However, on the 5 μm, groove-patterned substrate, where the tEMT cells demonstrated the most rapid motility relative to the control cells, with an increment of 0.18 μm/min (100%). Among the different physical cues from substrate, the cone pattern could impede the migration speed of tEMT cells. Furthermore, we recommend the groove-patterned with a 5 μm line-width substrate as a useful tool to differentiate control and tEMT cells by migration speed.

Ephrin‐B2 reverse signaling is required for topography but not pattern formation of lateral superior olivary inputs to the inferior colliculus

Graded and modular expressions of Eph-ephrins are known to provide positional information for the formation of topographic maps and patterning in the developing nervous system. Previously we have shown that ephrin-B2 is expressed in a continuous gradient across the tonotopic axis of the central nucleus of the inferior colliculus (CNIC), whereas patterns are discontinuous and modular in the lateral cortex of the IC (LCIC). The present study explores the involvement of ephrin-B2 signaling in the development of projections to the CNIC and LCIC arising from the lateral superior olivary nuclei (LSO) prior to hearing onset. Anterograde and retrograde fluorescent tracing methods in neonatal fixed tissue preparations were used to compare topographic mapping and the establishment of LSO layers/modules in wild-type and ephrin-B2(lacZ/+) mice (severely compromised reverse signaling). At birth, pioneer LSO axons occupy the ipsilateral IC in both groups but are delayed contralaterally in ephrin-B2(lacZ/+) mutants. By the onset of hearing, both wild-type and mutant projections form discernible layers bilaterally in the CNIC and modular arrangements within the ipsilateral LCIC. In contrast, ephrin-B2(lacZ/+) mice lack a reliable topography in LSO-IC projections, suggesting that fully functional ephrin-B2 reverse signaling is required for normal projection mapping. Taken together, these ephrin-B2 findings paired with known coexpression of EphA4 suggest the importance of these signaling proteins in establishing functional auditory circuits prior to experience.

Urokinase receptor counteracts vascular smooth muscle cell functional changes induced by surface topography.

Current treatments for human coronary artery disease necessitate the development of the next generations of vascular bioimplants. Recent reports provide evidence that controlling cell orientation and morphology through topographical patterning might be beneficial for bioimplants and tissue engineering scaffolds. However, a concise understanding of cellular events underlying cell-biomaterial interaction remains missing. In this study, applying methods of laser material processing, we aimed to obtain useful markers to guide in the choice of better vascular biomaterials. Our data show that topographically treated human primary vascular smooth muscle cells (VSMC) have a distinct differentiation profile. In particular, cultivation of VSMC on the microgrooved biocompatible polymer E-shell induces VSMC modulation from synthetic to contractile phenotype and directs formation and maintaining of cell-cell communication and adhesion structures. We show that the urokinase receptor (uPAR) interferes with VSMC behavior on microstructured surfaces and serves as a critical regulator of VSMC functional fate. Our findings suggest that microtopography of the E-shell polymer could be important in determining VSMC phenotype and cytoskeleton organization. They further suggest uPAR as a useful target in the development of predictive models for clinical VSMC phenotyping on functional advanced biomaterials.

EFFECT OF TOPOGRAPHICALLY PATTERNED POLY(DIMETHYLSILOXANE) SURFACES ON Pseudomonas aeruginosa ADHESION AND BIOFILM FORMATION

Bacterial pathogens, such as Pseudomonas aeruginosa, readily form biofilms on surfaces, limiting the efficacy of antimicrobial and antibiotic treatments. To mitigate biofilm formation, surfaces are often treated with antimicrobial agents, which have limited lifetime and efficacy. Recent studies have shown that well-ordered topographic patterns can limit bacterial attachment to surfaces and limit biofilm formation. In this study, nano and microscale patterned poly(dimethylsiloxane) surfaces were evaluated for their ability to affect adhesion and biofilm formation by Pseudomonas aeruginosa. Feature size and spacing were varied from 500 nm to 2 μm and included repeating arrays of square pillars, holes, lines and biomimetc Sharklet™ patterns. Bacterial surface adhesion and biofilm formation was assessed in microfluidic flow devices and under static conditions. Attachment profiles under static and fluid flow varied within topography types, sizes and spacing. Pillar structures of all sizes yielded lower surface attachment than line-based patterns and arrays of holes. This trend was also observed for biomimetic Sharklet™ patterns, with reduced bacterial attachment to "raised" features as compared to "recessed" features. Notably, none of the topographically patterned surfaces outperformed smooth surfaces (without topography) for resisting cell adhesion. Initial surface attachment patterns were indicative of subsequent biofilm formation and coverage, suggesting a direct role of surface topography in biofilm-based biofouling.

Nuclear and cellular alignment of primary corneal epithelial cells on topography

The basement membrane of the corneal epithelium presents biophysical cues in the form of topography and compliance that can modulate cytoskeletal dynamics, which, in turn, can result in altering cellular and nuclear morphology and alignment. In this study, the effect of topographic patterns of alternating ridges and grooves on nuclear and cellular shape and alignment was determined. Primary corneal epithelial cells were cultured on either planar or topographically patterned (400-4000 nm pitch) substrates. Alignment of individual cell body was correlated with respective nucleus for the analysis of orientation and elongation. A biphasic response in alignment was observed. Cell bodies preferentially aligned perpendicular to the 800 nm pitch; and with increasing pitch, cells increasingly aligned parallel to the substratum. Nuclear orientation largely followed this trend with the exception of those on 400 nm. On this biomimetic size scale, some nuclei oriented perpendicular to the topography while their cytoskeleton elements aligned parallel. Both nuclei and cell bodies were elongated on topography compared to those on flat surfaces. Our data demonstrate that nuclear orientation and shape are differentially altered by topographic features that are not mandated by alignment of the cell body. This novel finding suggests that nuanced differences in alignment of the nucleus versus the cell body exist and that these differences could have consequences on gene and protein regulation that ultimately regulate cell behaviors. A full understanding of these mechanisms could disclose novel pathways that would better inform evolving strategies in cell, stem cell, and tissue engineering as well as the design and fabrication of improved prosthetic devices.

Electrical nano-imprint lithography

We present a novel technique called electrical nano-imprint lithography (e-NIL) for topographic and electrostatic patterning of thermoplastic electret films at the nanometer scale. This versatile parallel process consists of simultaneously transferring micro- or nano-patterns from a conductive mold into a thermoplastic electret film and injecting positive or negative electrical charges into the bottom of the imprinted patterns. As proof of concept, we used this e-NIL process to fabricate arrays of 5 μm and 300 nm wide topographic charged patterns into polymethylmethacrylate (PMMA) thin films coated on silicon wafers. We demonstrated that these patterned PMMA films, exhibiting thousands of topographically confined and electrostatically active sites, can be used for high-throughput directed assembly of colloidal nanoparticles.

Abstract 269: Collagen Topographical Patterning Modulates Endothelial Cell Morphology, Gene Expression and Function

Endothelial cells (ECs) exposed to laminar shear stress align in the direction of blood flow and express low levels of adhesion molecules. However, it is unknown whether these biological responses result from shear stress per se, or if they can be recapitulated using topography cues from the underlying extracellular matrix (ECM). We hypothesized that collagen topographical patterning can regulate EC assembly and function in the absence of shear stress. We fabricated collagen-coated polydimethylsiloxane microchannels (30μm wide) to induce EC alignment. When grown on microchannels, the ECs underwent striking re-organization of their actin cytoskeleton (aligned within 10 degrees of the microchannel direction) and their focal adhesions (Fig A). Mimicking the reported EC responses to laminar shear stress, aligned ECs were more quiescent (reduced Ki67 expression), and less adhesive for monocytes (with 50% reductions in the expression of intercellular adhesion molecule 1, and in monocyte adhesion in a functional binding assay). DNA microarrays revealed a transcriptional signature of 600 genes that were differentially expressed by ECs cultured on the patterned substrates (Fig B), including genes previously not associated with EC function such as histones (HIST1H2AH, HIST2H2BF) and heat shock proteins (HSPB7, HSPB9). These results demonstrate that topographical cues from the underlying ECM are potent regulators of EC morphology and function and mimic the effects of laminar shear-stress. This work highlights the importance of cell-ECM interactions in maintenance of EC phenotype and has implications in the design of vascular conduits to minimize atherogenesis.

Silk Film Culture System for <em>in vitro</em> Analysis and Biomaterial Design

Silk films are promising protein-based biomaterials that can be fabricated with high fidelity and economically within a research laboratory environment 1,2 . These materials are desirable because they possess highly controllable dimensional and material characteristics, are biocompatible and promote cell adhesion, can be modified through topographic patterning or by chemically altering the surface, and can be used as a depot for biologically active molecules for drug delivery related applications 3-8 . In addition, silk films are relatively straightforward to custom design, can be designed to dissolve within minutes or degrade over years in vitro or in vivo, and are produce with the added benefit of being transparent in nature and therefore highly suitable for imaging applications 9-13. The culture system methodology presented here represents a scalable approach for rapid assessments of cell-silk film surface interactions. Of particular interest is the use of surface patterned silk films to study differences in cell proliferation and responses of cells for alignment 12,14 . The seeded cultures were cultured on both micro-patterned and flat silk film substrates, and then assessed through time-lapse phase-contrast imaging, scanning electron microscopy, and biochemical assessment of metabolic activity and nucleic acid content. In summary, the silk film in vitro culture system offers a customizable experimental setup suitable to the study of cell-surface interactions on a biomaterial substrate, which can then be optimized and then translated to in vivo models. Observations using the culture system presented here are currently being used to aid in applications ranging from basic cell interactions to medical device design, and thus are relevant to a broad range of biomedical fields.

Silk Film Culture System for <em>in vitro</em> Analysis and Biomaterial Design

Silk films are promising protein-based biomaterials that can be fabricated with high fidelity and economically within a research laboratory environment 1,2 . These materials are desirable because they possess highly controllable dimensional and material characteristics, are biocompatible and promote cell adhesion, can be modified through topographic patterning or by chemically altering the surface, and can be used as a depot for biologically active molecules for drug delivery related applications 3-8 . In addition, silk films are relatively straightforward to custom design, can be designed to dissolve within minutes or degrade over years in vitro or in vivo, and are produce with the added benefit of being transparent in nature and therefore highly suitable for imaging applications 9-13. The culture system methodology presented here represents a scalable approach for rapid assessments of cell-silk film surface interactions. Of particular interest is the use of surface patterned silk films to study differences in cell proliferation and responses of cells for alignment 12,14 . The seeded cultures were cultured on both micro-patterned and flat silk film substrates, and then assessed through time-lapse phase-contrast imaging, scanning electron microscopy, and biochemical assessment of metabolic activity and nucleic acid content. In summary, the silk film in vitro culture system offers a customizable experimental setup suitable to the study of cell-surface interactions on a biomaterial substrate, which can then be optimized and then translated to in vivo models. Observations using the culture system presented here are currently being used to aid in applications ranging from basic cell interactions to medical device design, and thus are relevant to a broad range of biomedical fields.

Directing self-assembly of nanostructures kinetically: Patterning and the Ehrlich-Schwoebel barrier

We present the results of kinetic Monte Carlo simulations of homoepitaxial growth on a patterned substrate in the presence of an extra barrier to a diffusing adatom crossing of steps from above (Ehrlich-Schwoebel barrier) on topographically patterned surfaces. Our results indicate that over a wide range of Ehrlich-Schwoebel barrier heights, incident atom fluxes, and temperatures that multilayer islands or ``growth mounds'' grow in arrangements which are directed by the topographical pattern. Our simulations indicate that a series of arrangements should form as the temperature is changed due to a competition between the temperature-dependent mound size and the pattern period. We compare these predictions with experimental observation of directed mound assembly on nanopatterned GaAs(001).