キーワード：生体親和性 細胞接着 バイオ界面 水の構造 医療機器
|田中 賢（たなか まさる）||データ更新日：2018.07.27|
科研費（基盤B,萌芽, 新学術領域）, ERATO, AMED, JST A-STEP, マッチングプランナーなど
2015.04～2016.03, 代表者：田中 賢, 九州大学.
2015.04～2016.03, 代表者：田中 賢, 九州大学.
|1.||Daiki Murakami, Yoko Kitahara, Shingo Kobayashi, Masaru Tanaka, Thermosensitive Polymer Biocompatibility Based on Interfacial Structure at Biointerface, ACS Biomaterials Science and Engineering, 10.1021/acsbiomaterials.8b00081, 4, 5, 1591-1597, 2018.05, [URL], The interfacial structure of a thermosensitive biocompatible polymer, poly[2-(2-methoxyethoxy)ethyl methacrylate] (PMe2MA), at the polymer/phosphate-buffered saline (PBS) interface was investigated by atomic force microscopy. A number of nanometer scale protrusions appeared at 37 °C and disappeared at 22 °C, reversibly. This structural change occurred above the lower critical solution temperature of PMe2MA in PBS (19 °C), indicating that the formation of protrusions was explained by the microphase separation of polymer and water at the interfacial region. The protein adsorption and platelet adhesion onto PMe2MA interface were drastically restrained at 22 °C compared to that at 37 °C. Detachment of NIH3T3 cells accompanied by the dissipation of protrusions on the PMe2MA interface was also demonstrated. These results indicate that the protrusions act as the scaffold for the protein adsorption and cell adhesion..|
|2.||Shingo Kobayashi, Miyuki Wakui, Yukihisa Iwata, Masaru Tanaka, Poly(ω-methoxyalkyl acrylate)s
Nonthrombogenic Polymer Family with Tunable Protein Adsorption, Biomacromolecules, 10.1021/acs.biomac.7b01247, 18, 12, 4214-4223, 2017.12, [URL], A series of polyacrylates with different n-alkyl side chain lengths (1 to 6, and 12 carbons) and a ω-methoxy terminal group (poly(ω-methoxyalkyl acrylate): PMCxA) were prepared to study their nonthrombogenicity using human platelet adhesion, micro bicinchoninic acid (micro BCA) protein assay, and enzyme-linked immunosorbent assay (ELISA) tests. In all cases, human platelet adhesion was suppressed on the PMCxA-coated substrates, and the number of human platelets adhered to the PMC3A (poly(3-methoxypropyl acrylate))-coated surface was comparable to that of commercialized nonthrombogenic coating agent poly(2-methoxyethyl acrylate) (PMEA, equal to PMC2A). The amount of protein adsorbed onto the PMCxA was investigated by micro BCA using bovine serum albumin (BSA) and human fibrinogen (hFbn), revealing that PMC3A exhibited significantly high resistance to nonspecific BSA adsorption. Additionally, the amount of hFbn adsorbed onto the PMC3A was suppressed to the same extent as PMEA. The exposure degree of platelet adhesion sites in adsorbed hFbn was evaluated using an ELISA test, and the degree on the PMCxA with three to six methylene carbons was comparable to the PMEA. The hydration water structure in the hydrated PMCxA was also characterized using differential scanning calorimetry (DSC). The amount of intermediate water, which is the hydration water molecules that moderately interact with the polymer matrix, was maximum in the PMEA with two methylene run lengths, whereas the amount decreased by increasing the number of methyelnes in the side chain. The amount of adsorbed protein increased with a decrease in the amount of intermediate water, suggesting that the protein adsorption amount is tunable by simply changing the number of methylene carbons in the side chain. The present study revealed that poly(ω-methoxyalkyl acrylate)s are useful for blood-contacting medical devices, and PMC3A is the best mode of PMCxA to apply as an antiprotein adsorption coating agent..
|3.||Kazuki Fukushima, Yuto Inoue, Yuta Haga, Takayuki Ota, Kota Honda, Chikako Sato, Masaru Tanaka, Monoether-Tagged Biodegradable Polycarbonate Preventing Platelet Adhesion and Demonstrating Vascular Cell Adhesion
A Promising Material for Resorbable Vascular Grafts and Stents, Biomacromolecules, 10.1021/acs.biomac.7b01210, 18, 11, 3834-3843, 2017.11, [URL], We developed a biodegradable polycarbonate that demonstrates antithrombogenicity and vascular cell adhesion via organocatalytic ring-opening polymerization of a trimethylene carbonate (TMC) analogue bearing a methoxy group. The monoether-tagged polycarbonate demonstrates a platelet adhesion property that is 93 and 89% lower than those of poly(ethylene terephthalate) and polyTMC, respectively. In contrast, vascular cell adhesion properties of the polycarbonate are comparable to those controls, indicating a potential for selective cell adhesion properties. This difference in the cell adhesion property is well associated with surface hydration, which affects protein adsorption and denaturation. Fibrinogen is slightly denatured on the monoether-tagged polycarbonate, whereas fibronectin is highly activated to expose the RGD motif for favorable vascular cell adhesion. The surface hydration, mainly induced by the methoxy side chain, also contributes to slowing the enzymatic degradation. Consequently, the polycarbonate exhibits decent blood compatibility, vascular cell adhesion properties, and biodegradability, which is promising for applications in resorbable vascular grafts and stents..
|4.||Kazuhiro Sato, Shingo Kobayashi, Asuka Sekishita, Miyuki Wakui, Masaru Tanaka, Synthesis and Thrombogenicity Evaluation of Poly(3-methoxypropionic acid vinyl ester)
A Candidate for Blood-Compatible Polymers, Biomacromolecules, 10.1021/acs.biomac.7b00221, 18, 5, 1609-1616, 2017.05, [URL], A poly(vinyl acetate) derivative, poly(3-methoxypropionic acid vinyl ester) (PMePVE), was synthesized to develop a new candidate for blood compatible polymers. The monomer MePVE was synthesized by a simple two-step reaction, and then the MePVE was polymerized via free radical polymerization to obtain PMePVE. Human platelet adhesion tests were performed to evaluate the thrombogenicity, and the platelet adhesion was suppressed on the PMePVE-coated substrate. To determine the expression of the nonthrombogenicity of the PMePVE, the plasma protein adsorption and a conformationally altered state of fibrinogen were analyzed by a microBCA assay and enzyme-linked immunosorbent assay. The adsorption and denaturation of the plasma proteins were inhibited on the PMePVE; thus, PMePVE exhibited blood compatibility. A distinctive hydration water structure in the nonthrombogenic polymer, intermediate water (IW), was observed in the hydrated PMePVE by differential scanning calorimetry analysis. The nonthrombogenicity of PMePVE is considered to be brought about by the presence of IW..
|5.||Daiki Murakami, Shingo Kobayashi, Masaru Tanaka, Interfacial Structures and Fibrinogen Adsorption at Blood-Compatible Polymer/Water Interfaces, ACS Biomaterials Science and Engineering, 10.1021/acsbiomaterials.6b00415, 2, 12, 2122-2126, 2016.12, [URL], The interfacial structures of a blood-compatible polymer, poly(2-methoxyethyl acrylate) (PMEA), and several analogues were investigated by atomic force microscopy (AFM). The blood-compatible polymers exhibited nanometer-scale protrusions that spontaneously and homogeneously formed at polymer/water and polymer/phosphate-buffered saline interfaces. AFM observation also revealed that fibrinogen adsorption occurred locally on the protrusions rather than uniformly at the interface, with the regions adjacent to the protrusions apparently preventing the adsorption of fibrinogen. The formation of these interfacial structures may be due to in-plane microphase separation of polymer and water at the interface..|
|6.||Takashi Hoshiba, Eri Nemoto, Kazuhiro Sato, Hiroka Maruyama, Chiho Endo, Masaru Tanaka, Promotion of Adipogenesis of 3T3-L1 Cells on Protein Adsorption-Suppressing Poly(2-methoxyethyl acrylate) Analogs, Biomacromolecules, 10.1021/acs.biomac.6b01340, 17, 11, 3808-3815, 2016.11, [URL], Stem cell differentiation is an important issue in regenerative medicine and tissue engineering. It has been reported that cell shape is one of the factors that determine the lineage commitment of mesenchymal stem cells (MSCs). Therefore, the substrates have been developed to control their shapes. Recently, we found that poly(2-methoxyethyl acrylate) (PMEA) analogs can control tumor cell shape through the alteration of protein adsorption. Here, the adipogenesis of an adipocyte-progenitor cell, 3T3-L1 cells, was attempted; adipogenesis was to be regulated by surfaces coated with PMEA analogs through the control of their shape. The adipogenesis of 3T3-L1 cells was promoted on the surfaces coated with PMEA and its analogs, PMe3A and PMe2A. Evident focal adhesions were hardly observed on these surfaces, suggesting that integrin signal activation was suppressed. Additionally, actin assembly and cell spreading were suppressed on these surfaces. Therefore, the surfaces coated with PMEA analogs are expected to be suitable surfaces to regulate adipogenesis through the suppression of cell spreading. Additionally, we found that protein adsorption correlated with actin assembly and adipogenesis..|
|7.||Kohei Osawa, Shingo Kobayashi, Masaru Tanaka, Synthesis of Sequence-Specific Polymers with Amide Side Chains via Regio-/Stereoselective Ring-Opening Metathesis Polymerization of 3-Substituted cis-Cyclooctene, Macromolecules, 10.1021/acs.macromol.6b01829, 49, 21, 8154-8161, 2016.11, [URL], Highly regio-/stereoregular (trans-head-to-tail) polymers with amide side chains on every eighth backbone carbon were successfully synthesized by ring-opening metathesis polymerization (ROMP) of 3-substituted cis-cyclooctene (3RCOE) using Grubbs second-generation catalyst (G2). Regioregular linear ethylene-acrylamide copolymers were also prepared via hydrogenation of the obtained poly(3RCOE)s. The thermal properties and solubility of the obtained polymers were strongly influenced by the presence of amide hydrogen in the side chains, the presence of unsaturated bonds in the carbon backbone, and the side chain density. The presence of amide hydrogen in the side chains resulted in the formation of crystalline polymers and the lack of solubility of these polymers in common organic solvents. In contrast, the absence of amide hydrogen in the side chains led to the formation of amorphous polymers exhibiting good solubility in common organic solvents, and decreasing values of Tg were observed for amorphous polymers as a result of the saturation of double bonds in the backbone via hydrogenation..|
|8.||Shingo Kobayashi, Kousaku Fukuda, Maiko Kataoka, Masaru Tanaka, Regioselective Ring-Opening Metathesis Polymerization of 3-Substituted Cyclooctenes with Ether Side Chains., Macromolecules, 10.1021/acs.macromol.6b00273, 49, 7, 2493-2501, 2016.04, [URL], Allyl-substituted cyclooctenes with ether side-chains [methoxy, methoxy-terminated oligo(ethylene glycol)s, and tetrahydrofurfuryloxy group] were prepared as monomers and polymerized by ring-opening metathesis polymerization (ROMP) using Grubbs second-generation catalyst. In all cases, the ROMP of allyl-substituted monomers proceeded in a regio- and stereoselective manner to afford polymers with remarkably high head-to-tail regioregularity with high trans-stereoregularity. The regio- and stereoregularity of polymers were affected by the bulkiness of the substituent, and the ROMP of tetrahydrofurfuryloxy-substituted cyclooctene exhibited nearly perfect regio- (head-to-tail = 99%) and stereoselectivity (trans-double bond = 99%). Chemical hydrogenation of obtained polymers afforded model poly(ethylene-co-vinyl ether)s with precisely placed ether branches on every eighth backbone carbon. Differential scanning calorimetry (DSC) was used to study the thermal properties, and the hydrophilicity of polymers was evaluated by water contact angle measurement. The surface hydrophilicity of unsaturated polymers was effectively tuned by changing the side-chain length of oligo(ethylene glycol) groups while maintaining the hydrophobic character unchanged for saturated versions..|
|9.||Takashi Hoshiba, Takayuki Otaki, Eri Nemoto, Hiroka Maruyama, Masaru Tanaka, Blood-Compatible Polymer for Hepatocyte Culture with High Hepatocyte-Specific Functions toward Bioartificial Liver Development, ACS applied materials & interfaces, 10.1021/acsami.5b05210, 7, 32, 18096-18103, 2015.08, [URL], The development of bioartificial liver (BAL) is expected because of the shortage of donor liver for transplantation. The substrates for BAL require the following criteria: (a) blood compatibility, (b) hepatocyte adhesiveness, and (c) the ability to maintain hepatocyte-specific functions. Here, we examined blood-compatible poly(2-methoxyethyl acrylate) (PMEA) and poly(tetrahydrofurfuryl acrylate) (PTHFA) (PTHFA) as the substrates for BAL. HepG2, a human hepatocyte model, could adhere on PMEA and PTHFA substrates. The spreading of HepG2 cells was suppressed on PMEA substrates because integrin contribution to cell adhesion on PMEA substrate was low and integrin signaling was not sufficiently activated. Hepatocyte-specific gene expression in HepG2 cells increased on PMEA substrate, whereas the expression decreased on PTHFA substrates due to the nuclear localization of Yes-associated protein (YAP). These results indicate that blood-compatible PMEA is suitable for BAL substrate. Also, PMEA is expected to be used to regulate cell functions for blood-contacting tissue engineering..|
|10.||Taito Sekine, Yusaku Tanaka, Chikako Sato, Masaru Tanaka, Tomohiro Hayashi, Evaluation of Factors To Determine Platelet Compatibility by Using Self-Assembled Monolayers with a Chemical Gradient, Langmuir, 10.1021/acs.langmuir.5b01216, 31, 25, 7100-7105, 2015.06, [URL], Intercorrelation among surface chemical composition, packing structure of molecules, water contact angles, amounts and structures of adsorbed proteins, and blood compatibility was systematically investigated with self-assembled monolayers (SAMs) with continuous chemical composition gradients. The SAMs were mixtures of two thiols: n-hexanethiol (hydrophobic and protein-adsorbing) and hydroxyl-tri(ethylene glycol)-terminated alkanethiol (hydrophilic and protein-resistant) with continuously changing mixing ratios. From the systematic analyses, we found that protein adsorption is governed both by sizes of proteins and hydrophobic domains of the substrate. Furthermore, we found a clear correlation between adsorption of fibrinogen and adhesion of platelets. Combined with the results of surface force measurements, we found that the interfacial behavior of water molecules is profoundly correlated with protein resistance and antiplatelet adhesion. On the basis of these results, we conclude that the structuring of water at the SAM-water interface is a critical factor in this context..|
|11.||Toyoaki Hirata, Hisao Matsuno, Daisuke Kawaguchi, Tomoyasu Hirai, Norifumi L. Yamada, Masaru Tanaka, Keiji Tanaka, Effect of local chain dynamics on a bioinert interface, Langmuir, 10.1021/acs.langmuir.5b00258, 31, 12, 3661-3667, 2015.03, [URL], Although many kinds of synthetic polymers have been investigated to construct blood-compatible materials, only a few have achieved success. To establish molecular designs for blood-compatible polymers, the chain structure and dynamics at the water interface must be understood using solid evidence as the first bench mark. Here we show that polymer dynamics at the water interface impacts on structure of the interfacial water, resulting in a change in protein adsorption and of platelet adhesion. As a particular material, a blend composed of poly(2-methoxyethyl acrylate) (PMEA) and poly(methyl methacrylate) was used. PMEA was segregated to the water interface. While the local conformation of PMEA at the water interface was insensitive to its molecular weight, the local dynamics became faster with decreasing molecular weight, resulting in a disturbance of the network structure of waters at the interface. This leads to the extreme suppression of protein adsorption and platelet adhesion..|
|12.||Shigeaki Morita, Masaru Tanaka, Effect of sodium chloride on hydration structures of PMEA and P(MPC- R -BMA), Langmuir, 10.1021/la502550d, 30, 35, 10698-10703, 2014.09, [URL], The hydration structures of two different types of biomaterials, i.e., poly(2-methoxyethyl acrylate) (PMEA) and a random copolymer of 2-methacryloyloxyethyl phosphorylcholine and n-butyl methacrylate (P(MPC-r-BMA)), were investigated by means of attenuated total reflection infrared (ATR-IR) spectroscopy. The effects of the addition of sodium chloride to liquid water in contact with the surfaces of the polymer films were examined. The neutral polymer of PMEA was easily dehydrated by NaCl addition, whereas the zwitterionic polymer of P(MPC-r-BMA) was hardly dehydrated. More specifically, nonfreezing water having a strong interaction with the PMEA chain and freezing bound water having an intermediate interaction were hardly dehydrated by contacting with normal saline solution, whereas freezing water having a weak interaction with the PMEA chain was readily dehydrated. In contrast, freezing water in P(MPC-r-BMA) is exchanged for the saline solution contacting with the material surface without dehydration..|
|13.||Takashi Hoshiba, Mayo Nikaido, Masaru Tanaka, Characterization of the attachment mechanisms of tissue-derived cell lines to blood-compatible polymers, Advanced Healthcare Materials, 10.1002/adhm.201300309, 3, 5, 775-784, 2014.06, [URL], Recent advances in biomedical engineering require the development of new types of blood-compatible polymers that also allow non-blood cell attachment for the isolation of stem cells and circulating tumor cells (CTCs) from blood and for the development of artificial organs for use under blood-contact conditions. Poly(2-methoxyethyl acrylate) (PMEA) and poly(tetrafurfuryl acrylate) (PTHFA) were previously identified as blood-compatible polymers. Here, it is demonstrated that cancer cells can attach to the PMEA and PTHFA substrates, and the differences in the attachment mechanisms to the PMEA and PTHFA substrates between cancer cells and platelets are investigated. It is also found that the adsorption-induced deformation of fibrinogen, which is required for the attachment and activation of platelets, does not occur on the PMEA and PTHFA substrates. In contrast, fibronectin is deformed on the PMEA and PTHFA substrates. Therefore, it is concluded that cancer cells and not platelets can attach to the PMEA and PTHFA substrates based on this protein-deformation difference between these substrates. Moreover, it is observed that cancer cells attach to the PMEA substrate via both integrin-dependent and -independent mechanisms and attach to the PTHFA substrate only through an integrin-dependent mechanism. It is expected that PMEA and PTHFA will prove useful for blood-contact biomedical applications..|
主要総説, 論評, 解説, 書評, 報告書等
|1.||田中 賢, 生体適合性材料に関する発表多数, 高分子学会.|
|2.||田中 賢, 生体適合性材料に関する発表多数, 高分子学会.|
2015.10～2015.10.20, 高分子学会, 座長（Chairmanship）.
2015.10～2015.10.20, 高分子学会, 座長（Chairmanship）.
2015.10～2015.10.20, 日本化学会化学フェスタ, 企画委員.
2015.10～2015.10.20, 高分子学会, 特別シンポジウム企画.
2015.10～2015.10, 生体適合性制御と要求特性掌握から実践する高分子バイオマテリアルの設計・開発戦略－モノマー（いち）からデザインするバイオインターフェイスと上市までの道筋－, 国内, 編集委員長.
Max Planck Institute for Intelligent Systems, Technical University of Denmark, Cambridge University, Germany, Denmark, UnitedKingdom, 2015.10～2015.10.
International Association of Advanced Materials Scientists Award for the year 2016 (IAAM SA-2016), International Association of Advanced Materials, 2016.03.
高分子学会旭化成賞, 高分子学会, 2011.10.