Stabilizing Coacervate by Microfluidic Engulfment Induced by Controlled Interfacial Energy.

Low interfacial energy, an intrinsic property of complex coacervate, enables the complex coacervate to easily encapsulate desired cargo substances, making it widely used in encapsulation applications. Despite this advantage, the low interfacial energy of the complex coacervate makes it unstable against mechanical mixing, and changes in pH and salt concentration. Hence, a chemical cross-linker is usually added to enhance the stability of the complex coacervate at the expense of sacrificing all intrinsic properties of the coacervate, including phase transition of the coacervate from liquid to solid. In this study, we observed an abrupt increase in the interfacial energy of the coacervate phase in mineral oil. By controlling the interfacial energy of the coacervate phase using a microfluidic device, we successfully created double engulfed PEG-diacrylate (PEGDA) coacervate microparticles, named DEPOT, in which the coacervate is engulfed in a cross-linked PEGDA shell. The engulfed coacervate remained as a liquid phase, retained its original low interfacial energy property to encapsulate the desired cargo substances, and infiltrated into the target site by a simple solvent exchange from oil to water.

One-step microfluidic synthesis of Janus microhydrogels with anisotropic thermo-responsive behavior and organophilic/hydrophilic loading capability.

We report one-step microfluidic synthesis and characterization of novel Janus microhydrogels composed entirely of the same base material, N-isopropylacrylamide (NIPAAm). The microhydrogels were fabricated by the microfluidic generation of Janus monomer microdroplets based on separation of a supersaturated aqueous NIPAAm solution into NIPAAm-rich and -poor phases followed by UV irradiation. The resulting Janus microhydrogels exhibited tunable anisotropic thermo-responsive behavior and organophilic/hydrophilic loading capability.

A portable pressure pump for microfluidic lab-on-a-chip systems using a porous polydimethylsiloxane (PDMS) sponge.

In this paper, we propose a novel portable and disposable pressure pump using a porous polydimethylsiloxane (PDMS) sponge and demonstrate its application to a microfluidic lab-on-a-chip. The porous PDMS sponge was simply fabricated by a sugar leaching technique based on capillary suction of pre-cured PDMS into lumps of sugar, thereby enabling us to achieve the porous PDMS sponge composed of interconnected micropores. To indicate the characteristics of the porous PDMS sponge and pump, we measured the average porosities of them whose values were 0.64 and 0.34, respectively. A stress-strain relationship of the fabricated portable pressure pump represented a linear behavior in the compressive strain range of 0 to 20%. Within this range, a pumping volume of the pressure pump could be linearly controlled by the compressed strain. Finally, the fabricated porous PDMS pump was successfully demonstrated as a portable pressure pump for a disposable microfluidic lab-on-a-chip for efficient detection of agglutination. The proposed portable pressure pump can be potentially applicable to various disposable microfluidic lab-on-a-chip systems.

Facile Fabrication of Electrospun Nanofiber Membrane-Integrated PDMS Microfluidic Chip via Silver Nanowires-Uncured PDMS Adhesive Layer.

Although direct electrospinning has been frequently utilized to develop a nanofiber membrane-integrated microfluidic chip, the dielectric substrate material retards the deposition of electrospun nanofibers on the substrate, and the rough surface formed by deposited nanofibers hinders the successful sealing. In this study we introduce a facile fabrication process of an electrospun nanofiber membrane-integrated polydimethylsiloxane (PDMS) microfluidic chip, called a NFM-PDMS chip, by applying the functional layer. The functional layer consists of a silver nanowires (AgNWs)-embedded uncured PDMS adhesive layer (SNUP), which not only effectively concentrates the electric field toward the PDMS substrate, but also provides a smooth surface for robust sealing. The AgNWs in the SNUP play a crucial role as a grounded collector and enable approximately 4× faster electrospinning than the conventional method, forming a free-standing nanofiber membrane. The uncured PDMS adhesive layer in the SNUP maintains the smooth surface after electrospinning and allows the rapid and leakage-free bonding of the NFM-PDMS chip using plasma treatment. A practical application of the NFM-PDMS chip is demonstrated by culturing the human keratinocyte cell line, HaCaT cells. The HaCaT cells are well grown on the free-standing nanofiber membrane under dynamic flow conditions, maintaining good viability over 95% for 7 days of culture.

Improved chondrogenic performance with protective tracheal design of Chitosan membrane surrounding 3D-printed trachea.

In recent tracheal tissue engineering, limitations in cartilage reconstruction, caused by immature delivery of chondrocyte-laden components, have been reported beyond the complete epithelialization and integration of the tracheal substitutes with the host tissue. In an attempt to overcome such limitations, this article introduces a protective design of tissue-engineered trachea (TraCHIM) composed of a chitosan-based nanofiber membrane (CHIM) and a 3D-printed biotracheal construct. The CHIM was created from chitosan and polycaprolactone (PCL) using an electrospinning process. Upon addition of chitosan to PCL, the diameter of electrospun fibers became thinner, allowing them to be stacked more closely, thereby improving its mechanical properties. Chitosan also enhances the hydrophilicity of the membranes, preventing them from slipping and delaminating over the cell-laden bioink of the biotracheal graft, as well as protecting the construct. Two weeks after implantation in Sprague-Dawley male rats, the group with the TraCHIM exhibited a higher number of chondrocytes, with enhanced chondrogenic performance, than the control group without the membrane. This study successfully demonstrates enhanced chondrogenic performance of TraCHIM in vivo. The protective design of TraCHIM opens a new avenue in engineered tissue research, which requires faster tissue formation from 3D biodegradable materials, to achieve complete replacement of diseased tissue.

Rapid harvesting of stem cell sheets by thermoresponsive bulk poly(N-isopropylacrylamide) (PNIPAAm) nanotopography.

To date, cell sheet engineering-based technologies have actualized diverse scaffold-free bio-products to revitalize unintentionally damaged tissues/organs, including cardiomyopathy, corneal defects, and periodontal damage. Although substantial interest is now centered on the practical utilization of these bio-products for patients, the long harvest period of stem cells- or other primary cell-sheets has become a huge hurdle. Here, we dramatically reduce the total harvest period of a cell sheet (from cell layer formation to cell sheet detachment) composed of human bone marrow mesenchymal stem cells (hBMSCs) down to 2 d with the help of bulk thermoresponsive poly(N-isopropylacrylamide) (PNIPAAm) substrate nanotopography, which is not achievable via the previous grafting methods using PNIPAAm. We directly replicated an isotropic 400 nm-nanopore-array pattern on a bulk PNIPAAm substrate through UV polymerization of highly concentrated NIPAAm monomers, which was achieved using a remarkably increased Young's modulus of bulk PNIPAAm that was 1500 times higher than conventional PNIPAAm. The rapid harvesting of the hBMSC sheet on the bulk PNIPAAm substrate nanotopography was not only based on the accelerated formation and maturation of the hBMSC layer, but also the easy detachment of the hBMSC sheet induced by the abrupt change in the surface roughness of the substrate below the lower critical solution temperature (LCST) owing to the enlarged surface area of the substrate. Our findings may contribute to reverse presumptions about the limitations regarding the grafting methods for the cell sheet harvest and could broaden the practical utilization of cell sheets for patients in the near future.

Hydrogel-Assisted Electrospinning for Fabrication of a 3D Complex Tailored Nanofiber Macrostructure.

Electrospinning has shown great potential in tissue engineering and regenerative medicine due to a high surface-area-to-volume ratio and an extracellular matrix-mimicking structure of electrospun nanofibers, but the fabrication of a complex three-dimensional (3D) macroscopic configuration with electrospun nanofibers remains challenging. In the present study, we developed a novel hydrogel-assisted electrospinning process (GelES) to fabricate a 3D nanofiber macrostructure with a 3D complex but tailored configuration by utilizing a 3D hydrogel structure as a grounded collector instead of a metal collector in conventional electrospinning. The 3D hydrogel collector was discovered to effectively concentrate the electric field toward itself similar to the metal collector, thereby depositing electrospun nanofibers directly on its exterior surface. Synergistic advantages of the hydrogel (e.g., biocompatibility and thermally reversible sol-gel transition) and the 3D nanofiber macrostructure (e.g., mechanical robustness and high permeability) provided by the GelES process were demonstrated in a highly permeable tubular tissue graft and a robust drug- or cell-encapsulation construct. GelES is expected to broaden potential applications of electrospinning to not only provide in vivo drug/cell delivery and tissue regeneration but also an in vitro drug testing platform by increasing the degree of freedom in the configuration of the 3D nanofiber macrostructure.

A collagen gel-coated, aligned nanofiber membrane for enhanced endothelial barrier function.

Herein, a collagen gel-coated and aligned nanofiber membrane named Col-ANM is developed, which remarkably improves endothelial barrier function by providing biochemical and topographical cues simultaneously. Col-ANM is fabricated by collagen gel coating process on an aligned polycaprolactone (PCL) nanofiber membrane, which is obtained by a simple electrospinning process adopting a parallel electrode collector. Human umbilical vein endothelial cells (HUVECs) cultured on Col-ANM exhibit remarkably enhanced endothelial barrier function with high expression levels of intercellular junction proteins of ZO-1 and VE-cadherin, a high TEER, and a cellular permeability compared with the artificial porous membranes in commercial cell culture well inserts. The enhanced endothelial barrier function is conjectured to be attributed to the synergistic effects of topographical and biochemical cues provided by the aligned PCL nanofibers and collagen gel in the Col-ANM, respectively. Finally, the reactive oxygen species is applied to the HUVEC monolayer formed on the Col-ANM to destroy the tight junctions between HUVECs. The destruction of the tight junctions is demonstrated by the decreased TEER value over time. Results indicate the potential of Col-ANM in modeling endothelial barrier dysfunction-related diseases.

Bulk poly(N-isopropylacrylamide) (PNIPAAm) thermoresponsive cell culture platform: toward a new horizon in cell sheet engineering.

Although a conventional method of utilizing thermoresponsive grafted poly(N-isopropylacrylamide) (PNIPAAm) enables the harvest of a healthy confluent cell-to-cell junction preserved cell sheet while limiting the use of the trypsin enzyme, the absolute necessity in the delicate control of a sensitive nm-scale PNIPAAm chain length inevitably decelerates the advancement of cell sheet engineering. In this study, we demonstrate, for the first time, a thermoresponsive cell culture platform composed only of a 'bulk' form of a PNIPAAm hydrogel with the Young's modulus being increased up to the MPa scale. The surface roughness of the bulk PNIPAAm hydrogel initially modulated by the cross-linker concentration was altered from the nm- to µm-scale in response to a change in temperature above/below the low critical solution temperature (LCST) of 32 °C. The appropriate control of the surface roughness allowed the stable attachment (above the LCST) and easy detachment (below the LCST) of diverse cells and enabled the harvest of cell sheets composed of cell lines (C2C12 and NIH3T3) or even primary cells (human umbilical vein endothelial cells and keratinocytes). During their incubation at 37 °C, the cell lines were able to be attached on every surface of the prepared PNIPAAm cell culture platforms, whereas the primary cells were found to be only attached on a surface having a roughness below ∼30 nm. Furthermore, in the aspect of cell sheet detachment at the incubation temperature of 20 °C, the cell sheets composed of cell lines were fully detached from the surface of the platform having a roughness of ∼10 µm or higher, while the cell sheets composed of primary cells were entirely detached from the surface with a roughness of ∼19 µm or higher. Based on such behaviors of the diverse cells at a given surface roughness, this study further suggests a universal thermoresponsive cell culture platform which allows the harvest of all types of cells from cell lines to primary cells in a desired shape. Our suggested universal cell culture platform could play a powerful and versatile role in accelerating the advancement of cell sheet engineering.

Surgical removal of embolic material after its unexpected migration through extracranial-intracranial anastomosis in the treatment of Barrow Type D carotid-cavernous fistula: case report.

Endovascular occlusion via the transvenous route is the favored treatment for indirect carotid-cavernous fistulas (CCFs). However, transarterial embolization can be used as an alternative method in patients with an inaccessible venous route. The authors present the case of a 49-year-old woman with a 2-month history of chemosis and proptosis in her right eye. Angiography demonstrated a Barrow Type D CCF. Transarterial Onyx embolization through the accessory meningeal artery was performed after an unsuccessful transvenous approach. Unexpected Onyx migrations to the cerebral arteries were detected while injecting the embolic material. Three hours after failed attempts to retrieve the Onyx cast endovascularly, it was microsurgically removed from the right middle cerebral artery. To the authors' knowledge, this is the first report of the surgical removal of Onyx from a normal cerebral artery.

Ultra-thin, aligned, free-standing nanofiber membranes to recapitulate multi-layered blood vessel/tissue interface for leukocyte infiltration study.

Leukocyte infiltration plays critical roles in tissue inflammation for pathogen clearance and tumor eradication. This process is regulated by complex microenvironments in blood vessels, including inflamed endothelium, blood flow, and perivascular components. The role of perivascular components in leukocyte infiltration has not been systematically investigated until recently mostly due to lack of technology. In this work, we developed a three-dimensional multi-layered blood vessel/tissue model with a nanofiber membrane, enabling real-time visualization of dynamic leukocyte infiltration and subsequent interaction with perivascular macrophages. We directly fabricated a highly aligned, free-standing nanofiber membrane with an ultra-thin thickness of ∼1 µm in microfluidic systems. Coating the nanofiber membrane with matrigel showed synergetic topographical and biochemical effects on the reconstitution of a well-aligned endothelial monolayer on the membrane. Our 3D multi-layered blood vessel/tissue model will offer a powerful and versatile tool for investigating the mechanism of leukocyte tissue infiltration and subsequent immune responses.

Decellularized corneal lenticule embedded compressed collagen: toward a suturable collagenous construct for limbal reconstruction.

Recently, compressed collagen has attracted much attention as a potential alternative for a limbal epithelial stem cell (LESC) carrier to treat limbal stem cell deficiency (LSCD), in that it can provide mechanically improved collagen fibrillar structures compared to conventional collagen hydrogel. However, its clinical efficacy as an LESC carrier has not yet been studied through in vivo transplantation due to limited mechanical strength that cannot withstand a force induced by surgical suturing and low resistance to enzymatic degradation. This study firstly presents a suturable LESC carrier based on compressed collagen in the form of a biocomposite. The biocomposite was achieved by integrating a decellularized corneal lenticule, which is a decellularized stromal tissue obtained from corneal refractive surgery, inside a compressed collagen to form a sandwich structure. A suture retention test verified that the biocomposite has a much higher suture retention strength (0.56 ± 0.12 N) compared to the compressed collagen (0.02 ± 0.01 N). The biocomposite also exhibited more than 3 times higher resistance to enzymatic degradation, indicating long-term stability after transplantation. In vitro cell culture results revealed that the biocomposite effectively supported the expansion and stratification of the LESCs with expressions of putative stem cell and differentiated corneal epithelial cell markers. Finally, the biocomposite verified its clinical efficacy by stably delivering the LESCs onto an eye of a rabbit model of LSCD and effectively reconstructing the ocular surface.

Synthesis of Poly(N-isopropylacrylamide) Janus Microhydrogels for Anisotropic Thermo-responsiveness and Organophilic/Hydrophilic Loading Capability.

Janus microparticles are compartmentalized particles with differing molecular structures and/or functionality on each of their two sides. Because of this unique property, Janus microparticles have been recognized as a new class of materials, thereby attracting a great deal of attention from various research fields. The versatility of these microparticles has been exemplified through their uses as building blocks for self-assembly, electrically responsive actuators, emulsifiers for painting and cosmetics, and carriers for drug delivery. This study introduces a detailed protocol that explicitly describes a synthetic method for designing novel Janus microhydrogels composed of a single base material, poly(N-isopropylacrylamide) (PNIPAAm). Janus microdroplets are firstly generated via a hydrodynamic focusing microfluidic device (HFMD) based on the separation of a supersaturated aqueous NIPAAm monomer solution and subsequently polymerized through exposure to UV irradiation. The resulting Janus microhydrogels were found to be entirely composed of the same base material, featured an easily identifiable compartmentalized morphology, and exhibited anisotropic thermo-responsiveness and organophilic/hydrophilic loading capability. We believe that the proposed method introduces a novel hydrogel platform with the potential for advanced synthesis of multi-functional Janus microhydrogels.

Nanotopography Promotes Pancreatic Differentiation of Human Embryonic Stem Cells and Induced Pluripotent Stem Cells.

Although previous studies suggest that nanotopographical features influence properties and behaviors of stem cells, only a few studies have attempted to derive clinically useful somatic cells from human pluripotent stem cells using nanopatterned surfaces. In the present study, we report that polystyrene nanopore-patterned surfaces significantly promote the pancreatic differentiation of human embryonic and induced pluripotent stem cells. We compared different diameters of nanopores and showed that 200 nm nanopore-patterned surfaces highly upregulated the expression of PDX1, a critical transcription factor for pancreatic development, leading to an approximately 3-fold increase in the percentage of differentiating PDX1(+) pancreatic progenitors compared with control flat surfaces. Furthermore, in the presence of biochemical factors, 200 nm nanopore-patterned surfaces profoundly enhanced the derivation of pancreatic endocrine cells producing insulin, glucagon, or somatostatin. We also demonstrate that nanopore-patterned surface-induced upregulation of PDX1 is associated with downregulation of TAZ, suggesting the potential role of TAZ in nanopore-patterned surface-mediated mechanotransduction. Our study suggests that appropriate cytokine treatments combined with nanotopographical stimulation could be a powerful tool for deriving a high purity of desired cells from human pluripotent stem cells.

Sinusoidal wavy surfaces for curvature-guided migration of T lymphocytes.

Micro/nanofabricated surfaces have been widely used for the study of topography-guided migration of cells. While the current studies mostly utilized micro/nanostructures containing sharp edges, internal tissues guiding migration of cells such as blood and lymphatic vessels, bone cavities, perivascular tracks have smooth microscale topographical structures. To overcome these limitations, we fabricated sinusoidal wavy surfaces with various wavelengths by deep X-ray lithography enabling precise and simultaneous control of amplitudes and wavelengths. Using these surfaces, we systematically studied curvature-guided migration of T cells. The majority of T cells migrated along the concave surfaces of sinusoidal wavy structures and as wavelength increased (or curvature decreased), preference to concave surfaces decreased. Integrin-mediated adhesion augmented the tendency of T cells crawling along grooves of highly curved wavy surfaces. To understand mechanisms of curvature-guided migration of T cells, T cells were treated with small molecule drugs such as blebbistatin and CK636, inhibiting myosin II activity and lamellipodia formation, respectively. While lamellipodia-inhibited T cells frequently crossed ridges, myosin II-inhibited T cells were mostly confined within concave surfaces. These results suggest that lamellipodia regulate local actin polymerization in response to surface curvature to maintain T cells within concave surfaces while myosin II-mediated contractile forces push T cells out of concave surfaces to make T cells less sensitive to surface curvature.

Microfluidic-assisted fabrication of flexible and location traceable organo-motor.

In this paper, we fabricate a flexible and location traceable micromotor, called organo-motor, assisted by microfluidic devices and with high throughput. The organo-motors are composed of organic hydrogel material, poly (ethylene glycol) diacrylate (PEGDA), which can provide the flexibility of their structure. For spatial and temporal traceability of the organo-motors under magnetic resonance imaging (MRI), superparamagnetic iron oxide nanoparticles (SPION; Fe3O4) were incorporated into the PEGDA microhydrogels. Furthermore, a thin layer of platinum (Pt) was deposited onto one side of the SPION-PEGDA microhydrogels providing geometrical asymmetry and catalytic propulsion in aqueous fluids containing hydrogen peroxide solution, H2O2. Furthermore, the motion of the organo-motor was controlled by a small external magnet enabled by the presence of SPION in the motor architecture.

Conservative condylectomy alone for the correction of mandibular asymmetry caused by osteochondroma of the mandibular condyle: a report of five cases.

We describe our experience with conservative condylectomy for the correction of facial asymmetry in five patients with osteochondroma of the mandibular condyle. All five patients presented with malocclusion and facial asymmetry, which are common clinical findings of osteochondroma involving the mandibular condyle. We performed conservative condylectomy without additional orthognathic surgery for all five patients, preserving the vertical height of the condylar process as much as possible. Following surgery, intermaxillary traction using a skeletal anchorage system with rubber elastics was performed on all patients to improve occlusion, and, when necessary, additional minimal orthodontic treatment was performed. The mean follow-up period was 42 months. At the last follow-up visit, all patients exhibited satisfactory facial symmetry and remodeling of the remaining condyle, with stable health and no signs of recurrence. In conclusion, conservative condylectomy alone, without subsequent orthognathic surgery, is adequate for the restoration of facial symmetry and the preservation of vertical condylar height in select patients with condylar osteochondroma.

Nanoengineered Polystyrene Surfaces with Nanopore Array Pattern Alters Cytoskeleton Organization and Enhances Induction of Neural Differentiation of Human Adipose-Derived Stem Cells.

Human adipose-derived stem cells (hADSCs) can differentiate into various cell types depending on chemical and topographical cues. One topographical cue recently noted to be successful in inducing differentiation is the nanoengineered polystyrene surface containing nanopore array-patterned substrate (NP substrate), which is designed to mimic the nanoscale topographical features of the extracellular matrix. In this study, efficacies of NP and flat substrates in inducing neural differentiation of hADSCs were examined by comparing their substrate-cell adhesion rates, filopodia growth, nuclei elongation, and expression of neural-specific markers. The polystyrene nano Petri dishes containing NP substrates were fabricated by a nano injection molding process using a nickel electroformed nano-mold insert (Diameter: 200 nm. Depth of pore: 500 nm. Center-to-center distance: 500 nm). Cytoskeleton and filopodia structures were observed by scanning electron microscopy and F-actin staining, while cell adhesion was tested by vinculin staining after 24 and 48 h of seeding. Expression of neural specific markers was examined by real-time quantitative polymerase chain reaction and immunocytochemistry. Results showed that NP substrates lead to greater substrate-cell adhesion, filopodia growth, nuclei elongation, and expression of neural specific markers compared to flat substrates. These results not only show the advantages of NP substrates, but they also suggest that further study into cell-substrate interactions may yield great benefits for biomaterial engineering.

Enhanced osteogenic fate and function of MC3T3-E1 cells on nanoengineered polystyrene surfaces with nanopillar and nanopore arrays.

During in vitro culture, cell fate and function, including cell adhesion, morphology, proliferation and differentiation, are affected by surface characteristics, such as geometry, wettability, hardness, chemistry and charge. This study replicated two different types of nanoengineered polystyrene surfaces (NPS) containing nanopillar (NPS-Pi) or nanopore (NPS-Po) arrays by hot embossing and investigated their topographical effects on cell behavior using osteoblast-like MC3T3-E1 cells. To mass-replicate NPS, rigid metal nano-stamps were manufactured by nickel electroforming onto two different nano-templates: (1) a nanopore-arrayed anodic aluminum oxide nano-template using two-step electrochemical oxidation and (2) a nanopillar-arrayed polymer using hot embossing process. The physical and mechanical properties of the NPS, including geometry, wettability, hardness and elastic modulus, were evaluated with the help of field emission-scanning electron microscopy, a contact angle meter, and a nanoindenter. The nanotopography maintained the bulk property, while drastically changing the surface properties. In vitro the NPS had significant effects on MC3T3-E1 cell morphology, attachment, proliferation and osteogenic differentiation compared to a flat substrate due to the altered physical and mechanical surface properties of the nanoengineered surface. Interestingly, the NPS-Po was more effective at enhancing cell proliferation and osteogenesis differentiation. One potential explanation for these results may be that the subcellular binding sites induced by the nanostructures changed the cell morphology and promoted contractile cytoskeletons, thereby enhancing osteogenic differentiation. This, which allows for the cost-effective replication of NPS and the control of cell behavior, has various applications with respect to biomedical and cell surface interaction studies, in addition to enhanced osteogenic cell fate and function.

Micropattern array with gradient size (µPAGS) plastic surfaces fabricated by PDMS (polydimethylsiloxane) mold-based hot embossing technique for investigation of cell-surface interaction.

Recently, it was found that the variations of physical environment significantly affect cell behaviors including cell proliferation, migration and differentiation. Through a plastic surface with controlled mechanical properties such as stiffness, one can change the orientation and migration of cells in a particular direction, thereby determining cell behaviors. In this study, we demonstrate a polydimethylsiloxane (PDMS) mold-based hot embossing technique for rapid, simple and low-cost replication of polystyrene (PS) surfaces having micropatterns. The PDMS mold was fabricated by UV-photolithography followed by PDMS casting; the elastomeric properties of PDMS enabled us to obtain conformal contact of the PDMS mold to a PS surface and to create high transcription quality of micropatterns on the PS surface. Two different types of circular micropillar and microwell arrays were successfully replicated on the PS surfaces based on the suggested technique. The micropatterns were designed to have various diameters (2-150 µm), spacings (2-160 µm) and heights (1.4, 2.4, 8.2 and 14.9 µm), so as to generate the gradient of physical properties on the surface. Experimental parametric studies indicated that (1) the embossing temperature became a critical processing parameter as the aspect ratio of micropattern increased and (2) the PDMS mold-based hot embossing could successfully replicate micropatterns, even having an aspect ratio of 2.7 for micropattern diameter of 6 µm, with an optimal processing condition (embossing pressure and temperature of 0.4 MPa and 130 °C, respectively) in this study. We carried out cell experiments with adipose-derived stem cells on the replicated PS surface with the height of 1.4 µm to investigate cellular behaviors in response to the micropattern array with gradient size. Cellular experiment results showed that the micropillar-arrayed surface improved cell proliferation as compared with the microwell-arrayed surface. We could also estimate the ranges of pattern sizes having the desired effects on the cellular behaviors.