Effect Of Substrate Topography Research Articles

Effects of substrate topography on the regulation of human fibroblasts and capsule formation via modulating macrophage polarization

The host-material interface is critical in determining the successful integration of medical devices into human tissue. The surface topography can regulate the fibrous capsule formation around implants through macrophage polarization, but the exact mechanism remains unclear. In this study, four types of microgrooves (10 or 50 µm in groove depths and 50 or 200 µm in groove widths) were fabricated on polydimethylsiloxane (PDMS) using lithography. The microgroove surfaces were characterized using the laser scanning confocal microscopy and fourier transform infrared spectroscopy. The effect of surface topography on macrophage phenotypes and conditioned medium (CM) collected from macrophages on human foreskin fibroblast 1 (HFF-1) were investigated. The result revealed that a deeper and narrower microgroove structure means a rougher surface. Macrophages tended to adhere and aggregate on group 50–50 surface (groove depths and widths of 50 µm). THP-1 cell polarized toward both inflammatory M1 and anti-inflammatory M2 macrophages on the surface of each group. Meanwhile, CM from macrophages culture on PDMS differentially up-regulated the proliferation, migration and fibrosis of HFF-1. Among them, the group 50–50 had the strongest promoting effect. In vivo, the inflammatory response and fibrotic capsule around the implants were observed at 1 week and 4 weeks. As time passed, the inflammatory response decreased, while the capsule thickness continued to increase. The rough material surface was more inclined to develop a severe fibrotic encapsulation. In conclusion, this finding further suggested a potential immunomodulatory effect of macrophages in mediating the fibrotic response to implants and facilitated the design of biomaterial interfaces for improving tissue integration.

Effect of substrate topography on the regulation of human corneal stromal cells

Optimal functionality of native corneal stroma depends on a well-ordered arrangement of extracellular matrix (ECM). To develop an in vitro corneal model, replication of the corneal in vivo microenvironment is needed. In this study, the impact of topographic cues on keratocyte phenotype is reported. Photolithography and polymer moulding were used to fabricate microgrooves on polydimethylsiloxane (PDMS) 2–2.5 μm deep and 5 μm, 10 μm, or 20 μm in width. Microgrooves constrained the cells body, compressed nuclei and led to cytoskeletal reorganization. It also influenced the concentration of actin filaments, condensation of chromatin and cell proliferation. Cells became more spread and actin filament concentration decreased as the microgroove width increased. Relationships were also demonstrated between microgroove width and cellular processes such as adhesion, migration and gene expression. Immunocytochemistry and gene expression (RT-PCR) analysis showed that microgroove width upregulated keratocyte specific genes. A microgroove with 5 μm width led to a pronounced alignment of cells along the edges of the microchannels and better supported cell polarization and migration compared with other microgroove widths or planar substrates. These findings provide important fundamental knowledge that could serve as a basis for better-controlled tissue growth and cell-engineering applications for corneal stroma regeneration through topographical patterns.

Micro-scaled topographies direct differentiation of human epidermal stem cells

Human epidermal stem cells initiate terminal differentiation when spreading is restricted on ECM-coated micropatterned islands, soft hydrogels or hydrogel-nanoparticle composites with high nanoparticle spacing. The effect of substrate topography, however, is incompletely understood. To explore this, primary human keratinocytes enriched for stem cells were seeded on a topographical library with over 2000 different topographies in the micrometre range. Twenty-four hours later the proportion of cells expressing the differentiation marker transglutaminase-1 was determined by high content imaging. As predicted, topographies that prevented spreading promoted differentiation. However, we also identified topographies that supported differentiation of highly spread cells. Topographies supporting differentiation of spread cells were more irregular than those supporting differentiation of round cells. Low topography coverage promoted differentiation of spread cells, whereas high coverage promoted differentiation of round cells. Based on these observations we fabricated a topography in 6-well plate format that supported differentiation of spread cells, enabling us to examine cell responses at higher resolution. We found that differentiated spread cells did not assemble significant numbers of hemidesmosomes, focal adhesions, adherens junctions, desmosomes or tight junctions. They did, however, organise the actin cytoskeleton in response to the topographies. Rho kinase inhibition and blebbistatin treatment blocked the differentiation of spread cells, whereas SRF inhibition did not. These observations suggest a potential role for actin polymerization and actomyosin contraction in the topography-induced differentiation of spread cells. Statement of SignificanceThe epidermis is the outer covering of the skin. It is formed by layers of cells called keratinocytes. The basal cell layer contains stem cells, which divide to replace cells in the outermost layers that are lost through a process known as differentiation. In this manuscript we have developed surfaces that promote the differentiation of epidermal stem cells in order to understand the signals that control differentiation. The experimental tools we have developed have the potential to help us to devise new treatments that control diseases such as psoriasis and eczema in which epidermal stem cell proliferation and differentiation are disturbed.

Laser Surface Microstructuring of a Bio-Resorbable Polymer to Anchor Stem Cells, Control Adipocyte Morphology, and Promote Osteogenesis.

New strategies in regenerative medicine include the implantation of stem cells cultured in bio-resorbable polymeric scaffolds to restore the tissue function and be absorbed by the body after wound healing. This requires the development of appropriate micro-technologies for manufacturing of functional scaffolds with controlled surface properties to induce a specific cell behavior. The present report focuses on the effect of substrate topography on the behavior of human mesenchymal stem cells (MSCs) before and after co-differentiation into adipocytes and osteoblasts. Picosecond laser micromachining technology (PLM) was applied on poly (L-lactide) (PLLA), to generate different microstructures (microgrooves and microcavities) for investigating cell shape, orientation, and MSCs co-differentiation. Under certain surface topographical conditions, MSCs modify their shape to anchor at specific groove locations. Upon MSCs differentiation, adipocytes respond to changes in substrate height and depth by adapting the intracellular distribution of their lipid vacuoles to the imposed physical constraints. In addition, topography alone seems to produce a modest, but significant, increase of stem cell differentiation to osteoblasts. These findings show that PLM can be applied as a high-efficient technology to directly and precisely manufacture 3D microstructures that guide cell shape, control adipocyte morphology, and induce osteogenesis without the need of specific biochemical functionalization.

Effect of substrate topography, material wettability and dielectric thickness on reversible electrowetting

Recent experiments by Kavousanakis et al., Langmuir, 2018 [1], showed that reversible electrowetting on superhydrophobic surfaces can be achieved by using a thick solid dielectric layer (e.g. tens of micrometers). It has also been shown, through equilibrium (static) computations, that when the dielectric layer is thick enough the electrostatic pressure is smoothly distributed along the droplet surface, thus the irreversible Cassie to Wenzel wetting transitions can be prevented. In the present work we perform more realistic, dynamic simulations of the electrostatically-induced spreading on superhydrophobic surfaces. To this end, we employ an efficient numerical scheme which enables us to fully take into account the topography of the solid substrate. We investigate in detail the role of the various characteristics of the substrate (i.e. the dielectric thickness, geometry and material wettability) and present relevant flow maps for the resulting wetting states. Through our dynamic simulations, we identify the conditions under which it is possible to achieve reversible electrowetting. We have found that not only the collapse (Cassie-Baxter to Wenzel) transitions but also the contact angle hysteresis of the substrate significantly affects the reversibility.

Effects of substrate stiffness on the tenoinduction of human mesenchymal stem cells.

Although the effects of substrate topography and modulus on MSC differentiation are well known with respect to osteogenesis and adipogenesis, there has been relatively little investigation on the effects of this phenomenon on tenogenesis. Furthermore, a relative role of topographical factors (matrix alignment vs. matrix modulus) in inducing tenogenic differentiation is not well understood. We investigated the effects of modulus and topographical alignment of type I collagen substrate on tendon differentiation. This study showed mimicking the tendon topography, i.e. increasing the substrate modulus as well as alignment increased the tenogenic differentiation. Therefore, the tenoinductive collagen material model developed in this study can be used in the research and development of tissue engineering tendon repair constructs in future.

The effects of nanofiber diameter and orientation on siRNA uptake and gene silencing

While substrate topography influences cell behavior, RNA interference (RNAi) has also emerged as a potent method for understanding and directing cell fate. However, the effects of substrate topography on RNAi remain poorly understood. Here, we report the influence of nanofiber architecture on siRNA-mediated gene-silencing in human somatic and stem cells. The respective model cells, human dermal fibroblasts (HDFs) and mesenchymal stem cells (MSCs), were cultured onto aligned or randomly oriented electrospun poly(ε-caprolactone) fibers of different average diameters (300 nm, 700 nm and 1.3 μm). In HDFs, decreasing fiber diameter from 1.3 μm to 300 nm improved Glyceraldehyde 3-phosphate dehydrogenase (GAPDH) and Collagen-I silencing efficiencies by ∼ 3.8 and ∼4.4 folds respectively (p < 0.05) while the effective siRNA uptake pathway was altered from clathrin-dependent endocytosis to macropinocytosis. In MSCs, aligned fibers generated significantly higher level of gene silencing of RE-1 silencing transcription factor (REST) and green fluorescent protein (GFP) (∼1.6 and ∼1.5 folds respectively, p < 0.05), than randomly-oriented fibers. Aligned fiber topography facilitated functional siRNA uptake through clathrin-mediated endocytosis and membrane fusion. Taken together, our results demonstrated a promising role of three-dimensional fibrous scaffolds in modulating siRNA-mediated gene-silencing and established the critical synergistic role of these substrates in modulating cellular behavior by RNAi.

Effects of substrate topography on current injection and light emission properties of organic light emitting devices

Substrate topography plays a critical role in the function of nano-scale materials and devices. We study small molecular organic light emitting devices (OLEDs) deposited onto non-planar substrates, where the substrate’s radius of curvature in some regions approaches the thickness of the active device layers. As a result, the electric field profile inside the organic charge transport layers is modified, influencing carrier injection, transport, and light emission properties. Experiments and numerical modeling suggest that charge balance and electroluminescence efficiency potentially can be improved in electron injection-limited OLED architectures via substrate geometry. These findings elucidate the optoelectronic behavior (and degradation) of OLEDs on imperfect substrates, and suggest a strategy based on substrate topography for controlling device behavior.

The effect of substrate topography on direct reprogramming of fibroblasts to induced neurons

Cellular reprogramming holds tremendous potential for cell therapy and regenerative medicine. Recently, fibroblasts have been directly converted into induced neurons (iNs) by overexpression of the neuronal transcription factors Ascl1, Brn2 and Myt1L. Hypothesizing that cell–topography interactions could influence the fibroblast-to-neuron reprogramming process, we investigated the effects of various topographies on iNs produced by direct reprogramming. Final iN purity and conversion efficiency were increased on micrograting substrates. Neurite branching was increased on microposts and decreased on microgratings, with a simplified dendritic arbor characterized by the reduction of MAP2+ neurites. Neurite outgrowth increased significantly on various topographies. DNA microarray analysis detected 20 differentially expressed genes in iNs reprogrammed on smooth versus microgratings, and quantitative PCR (qPCR) confirmed the upregulation of Vip and downregulation of Thy1 and Bmp5 on microgratings. Electrophysiology and calcium imaging verified the functionality of these iNs. This study demonstrates the potential of applying topographical cues to optimize cellular reprogramming.

The effect of substrate topography on hFOB cell behavior and initial cell adhesion evaluated by a cytodetacher

This study examined human fetal osteoblast (hFOB) cell morphology, adhesion force, and proliferation on a titanium-coated grooved surface. V-shaped grooves with a depth of 2.4 μm (T1) or 4.8 μm (T2) were produced in silicon wafers using photolithography and wet etching techniques. The grooved substrates were coated with a 200-nm-thick layer of titanium using a sputtering system. Smooth Ti-coated Si wafers were used as control surfaces. Analysis of the scanning electron microscopy observations shows that the cells responded to the micropattern by spreading out and becoming elongated. The MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay indicated that the grooved specimens had a significantly larger number of cells than did the control group after 5- and 15-day cultures. The cytocompatibility of specimens was quantitatively evaluated by a cytodetacher, which directly measures the detachment shear force of an individual cell to the substrate. After 30-min culture, the cell adhesion forces were 48.4, 136.6, and 103.3 nN for the smooth specimen, the T1 specimen, and the T2 specimen, respectively. The cell adhesion strengths were 294, 501, and 590 Pa for the smooth specimen, the T1 specimen, and the T2 specimen, respectively. The cell adhesion force and cell adhesion strength indicate the quality of cell adhesion, explaining the largest number of cells on grooved specimens. The experimental results suggest that the grooved patterns affect the cell shape and cytoskeletal structure, and thus influence the cell proliferation and cell adhesion force. The cytodetachment test with nanonewton resolution is a sensitive method for studying cell-biomaterial interaction.

Effect of substrate topography for YBa2 Cu3 O7-x coated conductors on the nucleation mechanism of buffer layer

The surface condition of substrate tape is an important factor to obtain epitaxial buffer layer on biaxially textured Ni tape for YBa2Cu3O7-x coated conductors. We prepare ceria films on Ni single crystal, biaxially textured Ni tape and sulfured Ni substrates by direct current magnetron sputtering. The results show that the ceria films prepared on Ni single crystal and sulfured Ni substrates each have a poor-textured grain structure. However, the ceria film fabricated on rolling assisted biaxially textured substrate (RABiTS) exhibits a good c-axis texture and desirable surface morphology. Reflection high-energy electron diffraction analysis indicates that the c(22) superstructure on the RABiTS Ni surface has a dramatic effect on the heteroepitaxial growth of oxide buffer layer.

INFLUENCE OF SUBSTRATE TOPOGRAPHY ON THE NUCLEATION AND GROWTH OF 321 STAINLESS STEEL THIN FILMS PRODUCED BY MAGNETRON SPUTTERING

We have investigated the effects of substrate topography on the nucleation and growth behavior of 321 stainless steel (ss) films, introducing textured surfaces into mica substrates by the abrasion treatment. In this study, two groups of samples were prepared at three different sputtering time within 12 s using radio frequency (r.f.) magnetron sputtering and characterized by atomic force microscopy. Good nucleation uniformity has been obtained on the unabraded mica substrates due to the statistical distribution of nucleation sites. Especially, we have already observed an interesting unusual nucleation phenomenon, the island–rim structure on the abraded mica substrates after 4 s, where the island is fractal-like and the rim around the island was only occupied by few grains for nucleation. These ramified islands were located at the wide grooves introduced as predominate nucleation sites. The island–rim structure formed in initial nucleation process is closely associated with x, y velocity components of surface atoms and the distribution of active sites (related intimately to the surface free energy σ and strain energy ε) for nucleation. With the increasing of the sputtering time, voids and overlarge grains have been observed in the island–rim region after the sputtering time of 8 s and 12 s, respectively.

Oriented astroglial cell growth on micropatterned polystyrene substrates

In an effort to develop a permissive environment for neural stem cell differentiation, directional growth of astrocytes has been achieved on polymer substrates in vitro. Manipulating a combination of physical and chemical cues, astrocyte adhesion and alignment in vitro were examined. To provide physical guidance, micropatterned polymer substrates of polystyrene (PS) were fabricated. Laminin was selectively adsorbed onto the grooves of the patterned surface. Rat type-1 astrocytes were seeded onto the micropatterned PS substrates, and the effects of substrate topography and the adsorption of laminin to the PS substrates on the behavior and morphology of the astrocytes were explored. The astrocytes were found to align parallel to the micropatterned grooves at initial seeding densities of approximately 7500, 13,000, and 20,000 cells/cm 2 due to the effects of the physical and chemical guidance mechanisms. Adsorbing laminin in the microgrooves of the micropatterned PS substrates improved cell adhesion and spreading of cytoskeletal filaments significantly. At these initial seeding densities, over 85% astrocyte alignment in the direction of the grooves was achieved on the micropatterned PS substrates with laminin adsorbed in the grooves. This combination of guidance cues has the potential to provide a permissive substrate for in vivo regeneration within the central nervous system.

Templated Self‐Assembly of Block Copolymers: Effect of Substrate Topography

Topographical confinement is used to template the formation of nanoscale domains in a self‐assembled block copolymer film. The topographical template controls the row spacings and feature dimensions of the copolymer and can deliberately introduce defects in the arrays (see Figure). For example, a sharp edge feature leads to a missing domain and pins the lateral position of the polymer array. (See also inside cover.)

Evolution of microstructure and texture during casting of AISI 304 stainless steel strip

The solidification behavior of AISI 304 stainless steel strip was studied using a melt/substrate contact apparatus, whereby a copper substrate embedded in a moving paddle is rapidly immersed into a steel melt to produce thin (∼1-mm gage) as-cast coupons. For cases where other casting conditions were kept constant, the effect of substrate topography and melt superheat on the development of microstructure and texture during solidification was studied using electron backscatter diffraction (EBSD) and optical microscopy. It was found that nucleation and growth of grains during solidification were influenced both by substrate topography and melt superheat. A ridged substrate produced a high density of randomly oriented grains at the chill surface with the preferred growth of 〈001〉-oriented grains perpendicular to the substrate wall producing a coarse columnar grain structure exhibiting a strong 〈001〉 fiber texture at the strip center. In contrast, a smooth substrate resulted in a lower nucleation density to produce a very coarse-grained columnar microstructure with moderate and essentially constant 〈001〉 fiber texture throughout the strip thickness. By the manipulation of casting parameters, it is possible to produce strip-cast austenitic stainless steel with a particular microstructure and texture.

Modeling 3D effects of substrate topography on step coverage and film morphology of thin metal films

This paper presents the use of three-dimensional (3D)-films to investigate 3D effects of microstructure and topography on the deposition of metal films over integrated circuit topography. 3D-films is a fully 3D thin film growth simulator which models both the evolution of the surface profile and the internal microstructure of the film. The simulator produces a detailed depiction of the film, including grain and columnar structure, and is capable of producing density and porosity information. This paper first presents the application of this simulator to specific problems posed by the sputter deposition of refractory metals as barriers and the filling of contacts and vias by electroplating of copper and CVD of tungsten. Specific problems addressed are step coverage over a variety of dual damascene topographies, the formation of notches and poor step coverage using directed sputtering and conformal filling of features. The program is used to address the ramifications of using geometric optimization of the via/contact geometry and the application of resputtering fluxes. Finally, issues concerning incomplete filling of dual damascene structures using CVD and plating are addressed.

The effect of substrate topography on the local electronic structure of WS 2 nanotubes

We explore the possibility of controlling electronic properties along an inorganic nanotube (INT) through the influence of nanometer-scale features in the underlying substrate. We examined single multi-walled WS 2 INTs using scanning tunneling microscopy (STM) in high vacuum. As long as the INTs lie flat on MoS 2 (0 0 0 1) or graphite (0 0 0 1) surfaces, they appear semimetallic. However, when the INT is suspended above the surface due to crossing steps or other nanotubes, a band gap opens up. We discuss this observation in terms of either a potential drop under the INT, or a change in its electronic properties due to its distortion when it lies flat on a surface.