Effect of Mechanical Instability of Polymer Scaffolds on Cell Adhesion

The adhesion of fibroblast on polymer bilayers composed of a glassy polystyrene (PS) prepared on top of a rubbery polyisoprene (PI) was studied. Since the top PS layer is not build on a glassy, or firm, foundation, the system becomes mechanically unstable with decreasing thickness of the PS layer. When the PS film was thinner than 25 nm, the number of cells adhered to the surface decreased and the cells could not spread well. On a parallel experiment, the same cell adhesion behavior was observed on plasma-treated PS/PI bilayer films, where in this case, the surface was more hydrophilic than that of the intact films. In addition, the fluorescence microscopic observations revealed that the formation of F-actin filaments in fibroblasts attached to the thicker PS/PI bilayer films was greater than those using the thinner PS/PI bilayer films. On the other hand, the thickness dependence of the cell adhesion behavior was not observed for the PS monolayer films. Taking into account that the amount of adsorbed protein molecules evaluated by a quartz crystal microbalance method was independent of the PS layer thickness of the bilayer films, our results indicate that cells, unlike protein molecules, could sense a mechanical instability of the scaffold

Biological Identification of Peptides that Specifically Bind to Poly(phenylene vinylene) Surfaces: Recognition of the Branched or Linear Structure of the Conjugated Polymer

Peptides that bind to poly(phenylene vinylene) (PPV) were identified by the phage display method. Aromatic amino acids were enriched in these peptide sequences, suggesting that a π−π interaction is the key interaction between the peptides and PPV. The surface plasmon resonance (SPR) experiments using chemically synthesized peptides demonstrated that the Hyp01 peptide, with the sequence His-Thr-Asp-Trp-Arg-Leu-Gly-Thr-Trp-His-His-Ser, showed an affinity constant (7.7 × 105 M−1) for the target, hyperbranched PPV (hypPPV) film. This value is 15-fold greater than its affinity for linear PPV (linPPV). In contrast, the peptide screened for linPPV (Lin01) showed the reverse specificity for linPPV. These results suggested that the Hyp01 and Lin01 peptides selectively recognized the linear or branched structure of PPVs. The Ala-scanning experiment, circular dichroism (CD) spectrometry, and molecular modeling of the Hyp01 peptide indicated that adequate location of two Trp residues by forming the polyproline type II (PII) helical conformation allowed the peptide to specifically interact with hypPPV

Polymer-Binding Peptides for the Noncovalent Modification of Polymer Surfaces: Effects of Peptide Density on the Subsequent Immobilization of Functional Proteins

Peptides that specifically bind to polyetherimide (PEI) were selected, characterized, and used for the noncovalent modification of the PEI surface. The peptides were successfully identified from a phage-displayed peptide library. A chemically-synthesized peptide composed of the Thr-Gly-Ala-Asp-Leu-Asn-Thr sequence showed an extremely high binding constant for the PEI films (5.6 × 108 M−1), which was more than three orders of magnitude greater than that for the reference polystyrene films. The peptide was biotinylated and immobilized onto the PEI films to further immobilize streptavidin (SAv). The amount of SAv bound depended on the density of immobilized peptide. It gradually increased with an increasing density of immobilized peptide and achieved a maximum (2.1 pmol cm−2) at a peptide density of 19.8 pmol cm−2. The ratio of peptide used for immobilizing SAv at the maximum value was only 11%, and was partially due to the low accessibility of SAv to the biotin moieties on the PEI films. Moreover, the amount of SAv bound gradually decreased at higher peptide densities, suggesting that the clustering of the peptides also inhibited the binding of SAv. Furthermore, peptides on the PEI films promoted the uniform immobilization of SAv with less structural denaturing. The immobilized SAv was able to further immobilize probe DNA to hybridize with its complementary DNA. These present results suggest that the density of immobilized peptide has a great impact on the surface modifications using polymer-binding peptides

Biological Selection of Peptides for Poly(l-lactide) Substrates

Short peptides that recognize the α form of poly(l-lactide) (PLLA) crystalline films were identified from a phage-displayed peptide library. An enzyme-linked immunosorbent assay (ELISA) revealed that the apparent binding constants of the phage clones for the α form of PLLA were greater than those of the unselected phage library. The specificity index for the α form of PLLA referred to a structurally similar atactic poly(methyl methacrylate) (at-PMMA), supporting the α form of PLLA specific binding of the selected phage. Amino acid residues with proton-donor lateral groups and hydrophobic alkyl groups were relatively enriched in a sequence of heptapeptides on the specific phage clones, thereby suggesting the presence of hydrogen bonding as well as hydrophobic interactions between the α form of PLLA and the peptides. Surface plasmon resonance (SPR) analysis revealed that the binding constant of the freed c22 heptapeptide (Gln-Leu-Met-His-Asp-Tyr-Arg) for the α form of PLLA was greater than those for reference at-PMMA, amorphous PLLA, and the β form of PLLA. It was found that c22 peptide can recognize slight differences in PLLA polymorphs such as a crystalline state and an arrangement of PLLA functional groups

A Peptide Motif Recognizing a Polymer Stereoregularity

A combinatorial phage display method was applied to films composed of a stereoregular polymer of methacrylates. The phage clones with selective affinity for isotactic (it) poly(methyl methacrylate) (PMMA) were isolated. Greater amounts of the phage clones bound to it-PMMA, compared to other stereoregular PMMAs. The phage expressing ELWRPTR most strongly bound to the polymer, and the selectivity was also the best. The peptide motif essential for the specific interaction with the stereoregular polymer was revealed

Two-Dimensional Cellular Patterning on a Polymer Film Based on Interfacial Stiffness

The mechanical properties in the outermost region of a polymer film strongly affect various material functions. We here propose a novel and promising strategy for the two-dimensional regulation of the mechanical properties of a polymer film at the water interface based on an inkjet drawing of silica nanoparticles (SNPs) underneath it. A film of poly­(2-hydroxyethyl methacrylate) (PHEMA), which exhibits excellent bioinertness properties at the water interface, was well fabricated on a substrate with a pattern of SNPs. X-ray photoelectron spectroscopy and atomic force microscopy confirmed that the surface of the PHEMA film was flat and chemically homogeneous. However, the film surface was in-plane heterogeneous in stiffness due to the presence of the underlying SNP lines. It was also noted that NIH/3T3 fibroblast cells selectively adhered and formed aggregates on the areas under which an SNP line was drawn

Peptide-Based Switching of Polymer Fluorescence in Aqueous phase

Peptide-Based Switching of Polymer Fluorescence in Aqueous phas

Simple Surface Treatment of Cell-Culture Scaffolds with Ultrafine Bubble Water

We propose a novel method to treat polymeric scaffold surfaces for cell culture with water containing nanobubbles, called ultrafine bubbles (UFBs), with typical diameters less than 1 μm. A thin film of polystyrene (PS) prepared on a solid substrate was exposed to UFB water for 2 days at room temperature. The PS surface was characterized by X-ray photoelectron spectroscopy (XPS), static contact angle measurements in water, and atomic force microscopy (AFM). The surface chemical composition and wettability of PS films remained unchanged after treatment, so that aggregation states of PS at film surfaces remained unaltered by UFB water. On the other hand, after treatment, many UFBs were adsorbed on hydrophobic PS surfaces. To study the effect of UFBs on scaffold properties, the adsorption behavior of fibronectin, which is a typical extracellular matrix protein involved in cell adhesion and proliferation, was examined. While the effect on the adsorption was unclear, the structural denaturation of fibronectin was enhanced after UFB treatment, so that the proliferation of fibroblast cells on PS surfaces was promoted

Water-Induced Crystal Transition and Accelerated Relaxation Process of Polyamide 4 Chains in Microfibers

Microplastics have recently been identified as one of the major contributors to environmental pollution. To design and control the biodegradability of polymer materials, it is crucial to obtain a better understanding of the aggregation states and thermal molecular motion of polymer chains in aqueous environments. Here, we focus on melt-spun microfibers of a promising biodegradable plastic, polyamide 4 (PA4), with a relatively greater number density of hydrolyzable amide groups, which is regarded as an alternative to polyamide 6. Aggregation states and thermal molecular motion of PA4 microfibers without/with a post-heating drawing treatment under dry and wet conditions were examined by attenuated total reflectance-Fourier transform infrared spectroscopy and wide-angle X-ray diffraction analysis in conjunction with dynamic mechanical analysis. Sorbed water molecules in the microfibers induced the crystal transition from a meta-stable γ-form to a thermodynamically stable α-form via activation of the molecular motion of PA4 chains. Also, the post-drawing treatment caused a partial structural change of PA4 chains, from an amorphous phase to a crystalline phase. These findings should be useful for designing PA4-based structural materials applicable for use in marine environments

Mechanical Stabilization of Deoxyribonucleic Acid Solid Films Based on Hydrated Ionic Liquid

Solid films of deoxyribonucleic acid (DNA) containing a hydrated ionic liquid, choline dihydrogen phosphate (CDP), were prepared by a solvent-casting method. Thermal properties, aggregation structure, thermal molecular motion, and tensile properties of CDP-containing DNA films were examined by thermogravimetry (TG), wide-angle X-ray diffraction (WAXD) measurement, dynamic mechanical analysis (DMA), and tensile tests, respectively. The water retentivity of the films at room temperature was much improved with CDP. The packing density of DNA helical chains clearly depended on the amount of CDP in the film. A small amount of CDP contributed to the suppression of the BI → BII conformational transition and the cooperative motion of the DNA duplex in the film. The tensile properties of the film drastically changed in the presence of CDP. When the amount of hydrated CDP in the film increased, the mechanical response of the film changed from glassy-like to rubbery-like via a semicrystalline-like state. The above results make it clear that CDP plays two major roles as a water absorber and plasticizer in the DNA film. Thus, it can be concluded that the use of an ionic liquid as an additive significantly increases the possibility of using a DNA solid film as a structural material