UV-assisted Capillary Force Lithography Research Articles

Continuous Tip Widening Technique for Roll-to-Roll Fabrication of Dry Adhesives

In this study, we reported continuous partial curing and tip-shaped modification methods for continuous production of dry adhesive with microscale mushroom-shaped structures. Typical fabrication methods of dry adhesive with mushroom-shaped structures are less productive due to the failure of large tips on pillar during demolding. To solve this problem, a typical pillar structure was fabricated through partial curing, and tip widening was realized through applying the proper pressure. Polyurethane acrylate was used in making the mushroom structure using two-step UV-assisted capillary force lithography (CFL). To make the mushroom structure, partial curing was performed on the micropillar, followed by tip widening. Dry adhesives with properties similar to those of typical mushroom-shaped dry adhesives were fabricated with reasonable adhesion force using the two-step UV-assisted CFL. This production technology was applied to the roll-to-roll process to improve productivity, thereby realizing continuous production without any defects. Such a technology is expected to be applied to various fields by achieving the productivity improvement of dry adhesives, which is essential for various applications.

Efficient myogenic commitment of human mesenchymal stem cells on biomimetic materials replicating myoblast topography

Recent developments in stem cell technologies have demonstrated human mesenchymal stem cells (hMSCs) as a possible cell source for cell-based therapies and regenerative medicine applications. Self-renewal and differentiation abilities of hMSCs have enabled hMSCs to be applied in regeneration of musculoskeletal tissue. hMSCs are able to myogenically differentiate via various approaches; however, the most efficient method has not been developed. Here, we describe the efficient commitment of hMSCs to the myogenic lineage on biomimetic substrates replicating myoblast topography. We have created a tissue culture platform that replicates the micro-and nanoscale topography of fully differentiated skeletal myoblasts. Using UV-assisted capillary force lithography, an optically transparent cellular model of fully differentiated myoblasts was developed using a UV curable poly(urethane acrylate) resin, which was fabricated and employed as a cell-culture substrate for the myogenic pattern of hMSCs. When hMSCs were cultured and differentiated on these biomimetic patterns, cells followed the underlying myoblast pattern and more efficiently committed to myogenic fate. These results demonstrate that myogenic potentials of hMSCs are highly depended on the micro- and nanoscale topographical cues. Furthermore, the described tissue culture platform can be used in larger culture settings with consistent results and easily applied to other lineage of hMSCs.

Advanced micro- and nanofabrication technologies for tissue engineering

The ability of the living body to heal and regenerate itself following trauma is astonishing. Numerous events of repair and regeneration occur during our lifetime, most of which we are never aware of. Unfortunately, in some cases, the injury or defect cannot be adequately repaired solely by nature and medical intervention is required.

Fabrication of poly(ethylene glycol): gelatin methacrylate composite nanostructures with tunable stiffness and degradation for vascular tissue engineering

Although synthetic polymers are desirable in tissue engineering applications for the reproducibility and tunability of their properties, synthetic small diameter vascular grafts lack the capability to endothelialize in vivo. Thus, synthetically fabricated biodegradable tissue scaffolds that reproduce important aspects of the extracellular environment are required to meet the urgent need for improved vascular grafting materials. In this study, we have successfully fabricated well-defined nanopatterned cell culture substrates made of a biodegradable composite hydrogel consisting of poly(ethylene glycol) dimethacrylate (PEGDMA) and gelatin methacrylate (GelMA) by using UV-assisted capillary force lithography. The elasticity and degradation rate of the composite PEG–GelMA nanostructures were tuned by varying the ratios of PEGDMA and GelMA. Human umbilical vein endothelial cells (HUVECs) cultured on nanopatterned PEG–GelMA substrates exhibited enhanced cell attachment compared with those cultured on unpatterned PEG–GelMA substrates. Additionally, HUVECs cultured on nanopatterned PEG-GelM substrates displayed well-aligned, elongated morphology similar to that of native vascular endothelial cells and demonstrated rapid and directionally persistent migration. The ability to alter both substrate stiffness and degradation rate and culture endothelial cells with increased elongation and alignment is a promising next step in recapitulating the properties of native human vascular tissue for tissue engineering applications.

Synergistic Effects of Matrix Nanotopography and Stiffness on Vascular Smooth Muscle Cell Function

Vascular smooth muscle cells (vSMCs) retain the ability to undergo modulation in their phenotypic continuum, ranging from a mature contractile state to a proliferative, secretory state. vSMC differentiation is modulated by a complex array of microenvironmental cues, which include the biochemical milieu of the cells and the architecture and stiffness of the extracellular matrix. In this study, we demonstrate that by using UV-assisted capillary force lithography (CFL) to engineer a polyurethane substratum of defined nanotopography and stiffness, we can facilitate the differentiation of cultured vSMCs, reduce their inflammatory signature, and potentially promote the optimal functioning of the vSMC contractile and cytoskeletal machinery. Specifically, we found that the combination of medial tissue-like stiffness (11 MPa) and anisotropic nanotopography (ridge width_groove width_ridge height of 800_800_600 nm) resulted in significant upregulation of calponin, desmin, and smoothelin, in addition to the downregulation of intercellular adhesion molecule-1, tissue factor, interleukin-6, and monocyte chemoattractant protein-1. Further, our results allude to the mechanistic role of the RhoA/ROCK pathway and caveolin-1 in altered cellular mechanotransduction pathways via differential matrix nanotopography and stiffness. Notably, the nanopatterning of the stiffer substrata (1.1 GPa) resulted in the significant upregulation of RhoA, ROCK1, and ROCK2. This indicates that nanopatterning an 800_800_600 nm pattern on a stiff substratum may trigger the mechanical plasticity of vSMCs resulting in a hypercontractile vSMC phenotype, as observed in diabetes or hypertension. Given that matrix stiffness is an independent risk factor for cardiovascular disease and that CFL can create different matrix nanotopographic patterns with high pattern fidelity, we are poised to create a combinatorial library of arterial test beds, whether they are healthy, diseased, injured, or aged. Such high-throughput testing environments will pave the way for the evolution of the next generation of vascular scaffolds that can effectively crosstalk with the scaffold microenvironment and result in improved clinical outcomes.

Migration of T Cells on Surfaces Containing Complex Nanotopography

T cells navigate complex microenvironments to initiate and modulate antigen-specific immune responses. While recent intravital microscopy study revealed that migration of T cells were guided by various tissue microstructures containing unique nanoscale topographical structures, the effects of complex nanotopographical structures on the migration of T cells have not been systematically studied. In this study, we fabricated surfaces containing nanoscale zigzag structures with various side lengths and turning angles using UV-assisted capillary force lithography and motility of T cells on zigzag patterned surfaces was studied. Motility of T cells was mostly affected by the turning angle, not by the side length, of the zigzag structures. In particular, motility behaviors of T cells near interfaces formed by turning points of zigzag patterns were significantly affected by turning angles. For obtuse turning angles, most of the T cells smoothly crossed the interfaces, but as the turning angle decreased, a substantial fraction of the T cells migrated along the interfaces. When the formation of lamellipodia, thin sheet-like structures typically generated at the leading edges of migrating cells by actin polymerization-driven membrane protrusion, was inhibited by an Arp2/3 inhibitor CK-636, a substantial fraction of T cells on those surfaces containing zigzag patterns with an acute turning angle were trapped at the interfaces formed by the turning points of the zigzag patterns. This result suggests that thin, wide lamellipodia at the leading edges of T cells play critical roles in motility of T cells in complex topographical microenvironments.

Dynamics of T cells on endothelial layers aligned by nanostructured surfaces

In this work, well-aligned endothelial cell (EC) layers were prepared by culturing ECs on surfaces containing nanoscale ridges/grooves fabricated by UV-assisted capillary force lithography. Then, the dynamics of T cells on well-aligned ECs were compared with that on randomly oriented ECs cultured on flat surfaces. With this experimental setting, we demonstrated for the first time that EC alignment is important for the regulation of transendothelial migration (TEM) of T cells, a critical step for leukocyte infiltration; T cells preferentially underwent TEM at the junctions surrounded by more than three ECs only if ECs surrounding those junctions were poorly aligned. As a result, TEM of T cells occurred more quickly and frequently on randomly oriented ECs cultured on flat surfaces than on well-aligned ECs cultured on nanostructured surfaces. This result will suggest a new strategy for the design of synthetic small diameter vascular grafts and extend our current knowledge of leukocyte dynamics on an inflamed endothelium.

Effect of nanogroove geometry on adipogenic differentiation

We present the effect of nanotopographically defined surfaces on adipocyte differentiationusing various nanogroove patterns. Parallel nanogroove arrays with equal inter-groovedistance (400, 550, 800 nm width) and varying distances (550 nm width with three differentspacings of 550, 1100, and 2750 nm) were fabricated by UV-assisted capillary forcelithography (CFL) on 18 mm diameter glass coverslips using biocompatible polyurethane(PU)-based material. After coating with fibronectin and subsequent culture of 3T3-L1preadipocytes, the degree of adipocyte differentiation was determined by Oil Red Ostaining and adipogenic gene expression. We observed that adipocyte differentiation wasslightly but substantially affected by culture on various nanogrooved surfaces. Inparticular, the cell crawling into nanogrooves contributed substantially to anenhanced level of differentiation with higher contact guidance, suggesting thatcell-to-surface interactions would play a role for the adipocyte differentiation.

Quantitative Analysis of the Combined Effect of Substrate Rigidity and Topographic Guidance on Cell Morphology

Live cells are exquisitely sensitive to both the substratum rigidity and texture. To explore cell responses to both these types of inputs in a precisely controlled fashion, we analyzed the responses of Chinese hamster ovary (CHO) cells to nanotopographically defined substrata of different rigidities, ranging from 1.8 MPa to 1.1 GPa. Parallel arrays of nanogrooves (800-nm width, 800-nm space, and 800-nm depth) on polyurethane (PU)-based material surfaces were fabricated by UV-assisted capillary force lithography (CFL) over an area of 5 mm × 3 mm. We observed dramatic morphological responses of CHO cells, evident in their elongation and polarization along the nanogrooves direction. The cells were progressively more spread and elongated as the substratum rigidity increased, in an integrin β1 dependent manner. However, the degree of orientation was independent of substratum rigidity, suggesting that the cell shape is primarily determined by the topographical cues.

Synergistically Enhanced Osteogenic Differentiation of Human Mesenchymal Stem Cells by Culture on Nanostructured Surfaces with Induction Media

We have examined the effects of surface nanotopography on in vitro osteogenesis of human mesenchymal stem cells (hMSCs). UV-assisted capillary force lithography was employed to fabricate a scalable (4x5 cm), well-defined nanostructured substrate of a UV curable polyurethane polymer with dots (150, 400, 600 nm diameter) and lines (150, 400, 600 nm width). The influence of osteogenic differentiation of hMSCs was characterized at day 8 by alkaline phosphatase (ALP) assay, RT-PCR, and real-time PCR analysis. We found that hMSCs cultured on the nanostructured surfaces in osteogenic induction media showed significantly higher ALP activity compared to unpatterned PUA surface (control group). In particular, the hMSCs on the 400 nm dot pattern showed the highest level of ALP activity. Further investigation with real-time quantitative RT-PCR analysis demonstrated significantly higher expression of core binding factor 1 (Cbfa1), osteopontin (OP), and osteocalcin (OC) levels in hMSCs cultured on the 400 nm dot pattern in osteogenic induction media. These findings suggest that surface nanotopography can enhance osteogenic differentiation synergistically with biochemical induction substance.

Direct differentiation of human embryonic stem cells into selective neurons on nanoscale ridge/groove pattern arrays

Human embryonic stem cells (hESCs) are pluripotent cells that have the potential to be used for tissue engineering and regenerative medicine. Repairing nerve injury by differentiating hESCs into a neuronal lineage is one important application of hESCs. Biochemical and biological agents are widely used to induce hESC differentiation. However, it would be better if we could induce differentiation of hESCs without such agents because these factors are expensive and it is difficult to control the optimal concentrations for efficient differentiation with reduced side effects. Moreover, the mechanism of differentiation induced by these factors is still not fully understood. In this study, we present evidence that nanoscale ridge/groove pattern arrays alone can effectively and rapidly induce the differentiation of hESCs into a neuronal lineage without the use any differentiation-inducing agents. Using UV-assisted capillary force lithography, we constructed nanoscale ridge/groove pattern arrays with a dimension and alignment that were finely controlled over a large area. Human embryonic stem cells seeded onto the 350-nm ridge/groove pattern arrays differentiated into neuronal lineage after five days, in the absence differentiation-inducing agents. This nanoscale technique could be used for a new neuronal differentiation protocol of hESCs and may also be useful for nanostructured scaffolding for nerve injury repair.

Wettability of nanoengineered dual-roughness surfaces fabricated by UV-assisted capillary force lithography

Micro- and nanoscale combined hierarchical polymer structures were fabricated by UV-assisted capillary force lithography. The method is based on the sequential application of engraved polymer molds with a UV-curable resin of polyurethane acrylate (PUA) followed by surface treatment with a trichloro(1H,1H,2H,2H-perfluorooctyl) silane in vapor phase. Two distinct wetting states were observed on these dual-roughness structures. One is “Cassie–Wenzel state” where a water droplet forms heterogeneous contact with microstructures and homogeneous contact with nanostructures. The other is “Cassie–Cassie state” where a droplet makes heterogeneous contact both with micro- and nanostructures. A simple thermodynamic model was developed to explain static contact angle, hysteresis, and wetting transition on dual-roughness structures.

UV-assisted capillary force lithography for engineering biomimetic multiscale hierarchical structures: From lotus leaf to gecko foot hairs

This feature article provides an overview of the recently developed two-step UV-assisted capillary force lithography and its application to fabricating well-defined micro/nanoscale hierarchical structures. This method utilizes an oxygen inhibition effect in the course of UV irradiation curing and a two-step moulding process, to form multiscale hierarchical or suspended nanobridge structures in a rapid and reproducible manner. After a brief description of the fabrication principles, several examples of the two-step UV-assisted moulding technique are presented. In addition, emerging applications of the multiscale hierarchical structures are briefly described.