Thermally Reversible Gel-Sol Transition of Hydrogels via Dissociation and Association of an Artificial Protein Nanocage.

Oligomeric protein nanocages often disassemble into their subunits and reassemble by external stimuli. Thus, using these nanocages as cross-linkers for hydrogel network structures is a promising approach to allow hydrogels to undergo stimuli-responsive gel-sol transitions or self-healing. Here, we report hydrogels that show a reversible gel-sol transition resulting from the heat-induced dissociation and reassociation of protein nanocages. The hydrogel contained the 60-mer artificial protein nanocage, TIP60, as a supramolecular cross-linker for polyethylene glycol network structures. The hydrogel showed a gel-to-sol transition upon heating at a temperature above the melting point of TIP60 and immediately returned to a gel state upon cooling to room temperature. During the heating and cooling treatment of the hydrogel, small-angle X-ray scattering analysis suggested the dissociation and reassociation of TIP60. Furthermore, we demonstrated redox-responsive cargo release from TIP60 in the hydrogel. These results showed the potential of TIP60 as a component of multi-stimuli-responsive hydrogels.

Synthesis of Thermoplastic Poly(2-methoxyethyl acrylate)-Based Polyurethane by RAFT and Condensation Polymerization.

Thermoplastic solid poly(2-methoxyethyl acrylate) (PMEA)-based polyurethane (PU) is synthesized through the reversible addition-fragmentation chain transfer (RAFT) polymerization and the condensation polymerization, using hydroxyl-terminated RAFT reagents and diisocyanate, respectively. Neat PMEA is a promising antithrombogenic liquid used in the medical fields. The thermoplastic property of the solid PMEA-based PU due to hydrogen bonding is confirmed by the dynamic mechanical analysis (DMA) at temperature below 72 °C. The antithrombogenic property of PMEA-based PU is also analyzed by the platelet adhesion test. The number of platelets on PMEA-based PU is 17 cells per unit area, which is smaller than that on the fluorinated diamond-like carbon (F-DLC), a well-known highly antithrombogenic material. It is concluded that a newly synthesized PMEA-based PU exhibits thermoplastic characteristics with excellent antithrombogenicity.

Multifunctionality of Iodinated Halogen-Bonded Polymer: Biodegradability, Radiopacity, Elasticity, Ductility, and Self-Healing Ability.

A polymer with high contents of ester bonds and iodine atoms was synthesized, exhibiting sufficient biodegradability and radioactivity for biomedical applications. The iodine moieties of the synthesized polyester can generate halogen bonding between molecules, which may develop additional functional properties through the bonding. In this study, poly(glycerol adipate) (PGA) was selected and synthesized as a polyester, which was then adequately conjugated with three different types of iodine compounds via the hydroxy groups of PGA. It was found that the iodine compounds could effectively work as donors of halogen bonding. The thermal analysis by differential scanning calorimetry (DSC) revealed that the glass transition temperature increased with the increase in the strength of interactions caused by π-π stacking and halogen bonding, eventually reaching 49.6 °C for PGA with triiodobenzoic groups. An elastomeric PGA with monoiodobenzoic groups was also obtained, exhibiting a high self-healing ability at room temperature because of the reconstruction of halogen bonding. Such multifaceted performance of the synthesized polyester with controllable thermal/mechanical properties was realized by halogen bonding, leading to a promising biomaterial with multifunctionality.

Compensation for Orientation Birefringence of PMMA by Blending Bottlebrush Polymers Composed of Well-Controlled Graft Chains.

The birefringence of optical polymers is a great issue in optical devices, inhibiting major applications of polymers to optical lenses and films. In this study, we have synthesized effective bottlebrush polymers with which we could attain almost zero birefringence when mixed with optical poly(methyl methacrylate) (PMMA). In detail, the PMMA bottlebrush polymers (PMMA-BBP) were synthesized by the ring-opening metathesis polymerization (ROMP) of norbornene-terminated PMMA macromonomers prepared via atom transfer radical polymerization (ATRP). Linear PMMA and PMMA-BBP were mixed to fabricate blend-film samples (PMMA/PMMA-BBP), which were uniaxially drawn to introduce molecular orientations. Linear PMMA possessed a negative value for its orientation birefringence, while the value of PMMA/PMMA-BBP increased as the PMMA-BBP content increased, whose orientation birefringence could reach almost zero when the ratio of the linear PMMA to PMMA-BBP became 73:27, regardless of the magnitude of the strain. The results reveal that the orientation birefringence of PMMA can be effectively controlled and removed by blending the appropriate content of PMMA-BBP.

Construction of a xylose-metabolizing yeast by genome integration of xylose isomerase gene and investigation of the effect of xylitol on fermentation.

A yeast with the xylose isomerase (XI) pathway was constructed by the multicopy integration of XI overexpression cassettes into the genome of the Saccharomyces cerevisiae MT8-1 strain. The resulting yeast strain successfully produced ethanol from both xylose as the sole carbon source and a mixed sugar, consisting of xylose and glucose, without any adaptation procedure. Ethanol yields in the fermentation from xylose and mixed sugar were 61.9% and 62.2% of the theoretical carbon recovery, respectively. Knockout of GRE3, a gene encoding nonspecific aldose reductase, of the host yeast strain improved the fermentation profile. Not only specific ethanol production rates but also xylose consumption rates was improved more than twice that of xylose-metabolizing yeast with the XI pathway using GRE3 active yeast as the host strain. In addition, it was demonstrated that xylitol in the medium exhibits a concentration-dependent inhibition effect on the ethanol production from xylose with the yeast harboring the XI-based xylose metabolic pathway. From our findings, the combination of XI-pathway integration and GRE3 knockout could be result in a consolidated xylose assimilation pathway and increased ethanol productivity.

Micropatterning of a 2-methacryloyloxyethyl phosphorylcholine polymer surface by hydrogenated amorphous carbon thin films for endothelialization and antithrombogenicity.

The existing first-generation drug-eluting stent (DES) has caused late and very late stent thrombosis related to incomplete stent endothelialization. Hence, biomaterials that possess sufficient anti-thrombogenicity and endothelialization with the controlled drug release system have been highly required. In this work, we have developed a newly designed drug-release platform composed of 2-methacryloyloxyethyl phosphorylcholine (MPC) polymer, a non-thrombogenic polymer, and micropatterned hydrogenated amorphous carbon (a-C:H), a cell-compatible thin film. The platelet adhesion and the endothelial cell adhesion behavior on the micropatterned substrates were investigated in vitro. The results indicated that the micropatterned a-C:H/MPC polymer substrates effectively supported the human umbilical vein endothelial cell (HUVEC) proliferation, while suppressing the platelet adhesion. Interestingly, the HUVEC exhibited different shape and behavior by changing the island size of the micropatterned a-C:H. By introducing both a non-thrombogenic polymer and cell-compatible thin films through a simple patterning method, we demonstrated that the platform had the potential to be utilized as a base material for DES with cell controllability. STATEMENT OF SIGNIFICANCE: The current first-generation drug-eluting stents (DES) would cause late and very late stent thrombosis due to the incomplete endothelialization of the metal stent material. In this work, we have developed a new DES platform composed of a 2-methacryloyloxyethyl phosphorylcholine (MPC) polymer micropatterned by hydrogenated amorphous carbon (a-C:H). Two types of differently micropatterned a-C:H stent surface were made. Our studies revealed that the micropatterned a-C:H/MPC polymer substrates could effectively enhance the endothelial cell (EC) proliferation, simultaneously suppressing the platelet adhesion, becoming a highly biocompatible material especially for indwelling devices including a drug-release device. The new drug-release platform could be utilized as a base material for cell-controllable coating on DES.

Nanocomposite injectable gels capable of self-replenishing regenerative extracellular microenvironments for in vivo tissue engineering.

Injectable hydrogels are biomaterials that have the potential to provide scaffolds to cells for in situ tissue regeneration with a minimally invasive implantation procedure. The success of in vivo tissue engineering utilizing injectable gels depends on providing cells with appropriate scaffolds that present an instructive extracellular microenvironment, which strongly influences the survival, proliferation, organization, and function of cells encapsulated within gels. One of the most important abilities of injectable gels to achieve this function is to adsorb and retain a wide variety of requisite bioactive molecules including nutrients, extracellular matrices, and growth/differentiation factors within gels. Previously, we developed nanocomposite injectable gels fabricated by simple combination of common biodegradable copolymers, poly(lactide-co-glycolide)-b-poly(ethylene glycol)-b-poly(lactide-co-glycolide) (PLGA-PEG-PLGA), and synthetic clay nanoparticles (LAPONITE®). We revealed that the nanocomposite injectable gels strongly adsorb ECM molecules including collagen and heparin within gels and retain them due to the ability of LAPONITE® in synchronization with the degradation of PLGA-PEG-PLGA and subsequent release of the degradation products. Human dermal fibroblast cells cultured on the nanocomposite gels showed enough high cell viability and proliferation for at least a week. Moreover, various kinds of human cells encapsulated within the nanocomposite gels exhibited significantly higher survival, proliferation, and three-dimensional organization in comparison with the PLGA-PEG-PLGA gel, LAPONITE® gel, and Matrigel. Furthermore, transplantation of mouse myoblast cells with the nanocomposite gels in model mice of skeletal muscle injury dramatically enhanced tissue regeneration and functional recovery, whereas cell transplantation with the PLGA-PEG-PLGA gel did not. Thus, the nanocomposite injectable gels possess unique abilities to self-replenish the regenerative extracellular microenvironment within the gels in the body, demonstrating the potential utility of the nanocomposite injectable gels for in vivo tissue engineering.

In vitro basic fibroblast growth factor (bFGF) delivery using an antithrombogenic 2-methacryloyloxyethyl phosphorylcholine (MPC) polymer coated with a micropatterned diamond-like carbon (DLC) film.

In this study, a newly designed drug-release platform composed of an antithrombogenic 2-methacryloyloxyethyl phosphorylcholine (MPC) polymer was introduced, which was impregnated with basic fibroblast growth factor (bFGF) (bFGF/MPC polymer) to enhance the endothelial cell activation. The platform was also coated with an ultrathin micropatterned diamond-like carbon (DLC) film (DLC/bFGF/MPC polymer) to precisely control the drug release rate and the cell compatibility. The resulting DLC/bFGF/MPC polymer could effectively prolong the bFGF release rate by depositing the micropatterned DLC. The number of adherent platelets on the DLC/bFGF/MPC polymer was significantly lower (about 1/14) than that on a currently used stent made of stainless steel (SUS316L), indicating the enhanced antithrombogenicity in the DLC/bFGF/MPC polymer. The proliferation of endothelial cells on the DLC/bFGF/MPC polymer and the DLC/MPC polymer (without bFGF) were also examined. It was found that the optical density of HUVEC on the DLC/bFGF/MPC polymer determined by WST-8 assay was higher by 25%than that on the DLC/MPC polymer (without bFGF) measured after 72 h of incubation. Our results suggest that the released bFGF that contributes to the expression of other growth factors results in the early proliferation of the HUVEC on the DLC/bFGF/MPC polymer. © 2017 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 105A: 3384-3391, 2017.

Solution parameters for the fabrication of thinner silicone fibers by electrospinning.

Silicone fibers were synthesized by electrospinning, where 17 solvents with different chemical properties (boiling point, conductivity, viscosity, dielectric constant, and solubility parameter) were used to dissolve the silicone polymer for the formation of fibers through electrospinning. Previous reports on the miniaturization of fibers of polymers dissolved in a solvent suggested that the low viscosity and the high conductivity of the polymer solution were the key parameters to form thinner fibers when using a single solvent. Here we have found a powerful way to search for good solvents to reduce the fiber diameters as well as to dissolve the polymers. By considering different types of solvents, it was found that the solubility parameters conclusively determined the smallest fiber diameters of the silicone polymers. The solubility parameter of the silicone polymer should be lower than those of the solvents to make thinner fibers. The results have revealed the strong relationship between the diameters of the fibers and the solubility parameters of the solvents, and they indicate that the solubility parameter could be a good indicative parameter in selecting solvents during the fabrication of thinner fibers by electrospinning, especially for siloxane polymers.

Laminin active peptide/agarose matrices as multifunctional biomaterials for tissue engineering.

Cell adhesive peptides derived from extracellular matrix components are potential candidates to afford bio-adhesiveness to cell culture scaffolds for tissue engineering. Previously, we covalently conjugated bioactive laminin peptides to polysaccharides, such as chitosan and alginate, and demonstrated their advantages as biomaterials. Here, we prepared functional polysaccharide matrices by mixing laminin active peptides and agarose gel. Several laminin peptide/agarose matrices showed cell attachment activity. In particular, peptide AG73 (RKRLQVQLSIRT)/agarose matrices promoted strong cell attachment and the cell behavior depended on the stiffness of agarose matrices. Fibroblasts formed spheroid structures on the soft AG73/agarose matrices while the cells formed a monolayer with elongated morphologies on the stiff matrices. On the stiff AG73/agarose matrices, neuronal cells extended neuritic processes and endothelial cells formed capillary-like networks. In addition, salivary gland cells formed acini-like structures on the soft matrices. These results suggest that the peptide/agarose matrices are useful for both two- and three-dimensional cell culture systems as a multifunctional biomaterial for tissue engineering.