Co-reporter:Esha Das
Journal of Polymer Science Part A: Polymer Chemistry 2017 Volume 55(Issue 5) pp:876-884
Publication Date(Web):2017/03/01
DOI:10.1002/pola.28440
ABSTRACTHere, we report that carboxylated poly-l-lysine, a polyampholyte, shows lower critical solution temperature (LCST)-type temperature-responsive liquid–liquid phase separation and coacervate formation in aqueous solutions. The phase-separation temperature of polyampholytes is strongly affected by the polymer concentration, balance between the carboxyl and amino groups, hydrophobicity of the side chain, and NaCl concentration in the solution. We concluded that the phase separation was caused by both electrostatic interactions between the carboxyl and amino groups and intermolecular hydrophobic interactions. The addition of NaCl weakened the electrostatic interactions, causing the two phases to remix. The introduction of the hydrophobic moiety decreased the phase-separation temperature by making the molecular interactions stronger. Finally, temperature-responsive hydrogels were prepared from the polyampholytes to explore their applicability as biomaterials and in drug delivery systems. The fine-tuning of the phase-separation temperature of poly-l-lysine-based polyampholytes through molecular design should open new avenues for their use in precisely controlled biomedical applications. © 2016 Wiley Periodicals, Inc. J. Polym. Sci., Part A: Polym. Chem. 2017, 55, 876–884
Co-reporter:Sana Ahmed;Satoshi Fujita
Advanced Healthcare Materials 2017 Volume 6(Issue 14) pp:
Publication Date(Web):2017/07/01
DOI:10.1002/adhm.201700207
Immunotherapy is an exciting new approach to cancer treatment. The development of a novel freeze-concentration method is described that could be applicable in immunotherapy. The method involves freezing cells in the presence of pH-sensitive, polyampholyte-modified liposomes with encapsulated ovalbumin (OVA) as the antigen. In RAW 264.7 cells, compared to unfrozen, freeze-concentration of polyampholyte-modified liposomes encapsulating OVA resulted in efficient OVA uptake and also allowed its delivery to the cytosol. Efficient delivery of OVA to the cytosol was shown to be partly due to the pH-dependence of the polyampholyte-modified liposomes. Cytosolic OVA delivery also resulted in significant up-regulation of the major histocompatibility complex class I pathway through cross-stimulation, as well as an increase in the release of IL-1β, IL-6, and TNF-α. The results demonstrate that the combination of a simple freeze-concentration method and polyampholyte-modified liposomes might be useful in future immunotherapy applications.
Co-reporter:Monika Patel;Tatsuo Kaneko
Journal of Materials Chemistry B 2017 vol. 5(Issue 19) pp:3488-3497
Publication Date(Web):2017/05/17
DOI:10.1039/C7TB00701A
Precise and controlled drug delivery systems are required to facilitate effective therapeutics. To address this need, we devised a micelle–hydrogel composite based on amphiphilic polypeptides as a general carrier model for the switchable and controlled release of dual drugs. Two different di-block polypeptides, poly(L-lysine-b-L-phenylalanine) and poly(L-glutamic acid-b-L-phenylalanine) (PGA–PPA), were synthesized to form distinct self-assembling micellar systems that were loaded with curcumin and amphotericin B, respectively, as model drugs. The drug-loaded micellar mixture was crosslinked utilizing the pendant amino groups of the L-lysine side chains via genipin to yield a micelle–hydrogel composite with PGA–PPA micelles trapped in an interlinked hydrogel system. This composite allowed for controlled multiphasic drug release and could be effectively tuned to moderate the pace and amount of drug release and be easily regulated to switch the drug release kinetics over a range of simple factors such as change in pH, cross-linking density, and composition.
Co-reporter:Minkle Jain
Journal of Biomedical Materials Research Part A 2016 Volume 104( Issue 6) pp:1379-1386
Publication Date(Web):
DOI:10.1002/jbm.a.35672
Abstract
Engineered tissues are excellent substitutes for treating organ failure associated with disease, injury, and degeneration. Designing new biomaterials with controlled release profiles, good mechanical properties, and cell adhesion characteristics can be useful for the formation of specific functional tissues. Here, we report the formulation of nanocomposite hydrogels based on carboxylated poly-l-lysine and synthetic clay laponite XLG in which four-arm polyethylene glycol with N-hydroxy succinimide ester (PEG-NHS) was used as the chemical crosslinker. Interestingly, the degradation of this gel could be adjusted from a few days to a few months. Incorporation of laponite XLG resulted in the formation of mechanically tough hydrogels and conferred cytocompatibility. The mechanical properties of the nanocomposite could be modulated by changing the crosslinking density and laponite concentration. The feasibility of using this system for cellular therapies was investigated by evaluating cell adhesion on the nanocomposite surface. Thus, these nanocomposites can serve as scaffolds with tunable mechanical and degradation properties that also provide structural integrity to tissue constructs. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 104A: 1379–1386, 2016.
Co-reporter:Kazuaki Matsumura, Keiko Kawamoto, Masahiro Takeuchi, Shigehiro Yoshimura, Daisuke Tanaka, and Suong-Hyu Hyon
ACS Biomaterials Science & Engineering 2016 Volume 2(Issue 6) pp:1023
Publication Date(Web):April 18, 2016
DOI:10.1021/acsbiomaterials.6b00150
Vitrification is used for cryopreserving oocytes and embryos. Successful vitrification and preservation typically require very rapid cooling. We report a novel slow vitrification method for the cryopreservation of two-dimensional cell constructs using a vitrification solution (VS) of PBS containing 6.5 M ethylene glycol, 0.5 M sucrose, and 10% w/w carboxylated poly-l-lysine (COOH-PLL), a novel polymeric cryoprotectant and stabilizing agent that likely inhibits ice crystallization. Stabilization of the glassy state and inhibition of devitrification were confirmed by thermal analysis. Slow vitrification at rates of 4.9 and 10.8 °C/min using VS with 10% COOH-PLL significantly improved the viability of cultured human mesenchymal stem cell monolayers after freezing and induced less apoptosis than when VS was used without COOH-PLL. Moreover, the cells maintained differentiation capacity. COOH-PLL improved vitrification through the inhibition of devitrification. This novel, simple method for slow vitrification is expected to be widely applicable for the preservation of tissue-engineered constructs and may facilitate the industrialization of regenerative medicine.Keywords: cryopreservation; ice crystallization; polyampholyte; tissue-engineered construct; vitrification
Co-reporter:Wichchulada Chimpibul;Toshio Nagashima;Fumiaki Hayashi;Naoki Nakajima;Suong-Hyu Hyon
Journal of Polymer Science Part A: Polymer Chemistry 2016 Volume 54( Issue 14) pp:2254-2260
Publication Date(Web):
DOI:10.1002/pola.28099
ABSTRACT
Although periodate-oxidized dextran is widely used in biomedical applications, the degradation mechanism of oxidized dextran has not yet been elucidated. Herein, we propose a novel main chain scission mechanism of oxidized dextran triggered by reaction with amine. NMR analysis revealed four hemiacetal substructures during oxidation by periodate. Kinetic analysis showed that the degradation time constant of the C3-removed substructure and increasing time constant of the reducing end protons are consistent with the decrease in molecular weight determined by gel permeation chromatography. A methylene group is generated at the same time constant of degradation, indicating that oxidized dextran degradation proceeds via a Maillard reaction. Oxidized dextran does not degrade in saline solution without reactive amine species. Thus, we conclude that oxidized dextran is degraded in the main chain via Schiff base formation through a Maillard reaction, depending on the oxidation ratio and amino acid concentration. These findings help to elucidate the reaction mechanism of polysaccharide degradation and develop novel biodegradable polysaccharide materials for biomedical applications. © 2016 Wiley Periodicals, Inc. J. Polym. Sci., Part A: Polym. Chem. 2016, 54, 2254–2260
Co-reporter:Robin Rajan, Fumiaki Hayashi, Toshio Nagashima, and Kazuaki Matsumura
Biomacromolecules 2016 Volume 17(Issue 5) pp:
Publication Date(Web):April 14, 2016
DOI:10.1021/acs.biomac.6b00343
Cryopreservation enables long-term preservation of cells at ultralow temperatures. Current cryoprotective agents (CPAs) have several limitations, making it imperative to develop CPAs with advanced properties. Previously, we developed a novel synthetic polyampholyte-based CPA, copolymer of 2-(dimethylamino)ethyl methacrylate (DMAEMA) and methacrylic acid(MAA) (poly(MAA-DMAEMA)), which showed excellent efficiency and biocompatibility. Introduction of hydrophobicity increased its efficiency significantly. Herein, we investigated the activity of other polyampholytes. We prepared two zwitterionic polymers, poly(sulfobetaine) (SPB) and poly(carboxymethyl betaine) (CMB), and compared their efficiency with poly(MAA-DMAEMA). Poly-SPB showed only intermediate property and poly-CMB showed no cryoprotective property. These data suggested that the polymer structure strongly influences cryoprotection, providing an impetus to elucidate the molecular mechanism of cryopreservation. We investigated the mechanism by studying the interaction of polymers with cell membrane, which allowed us to identify the interactions responsible for imparting different properties. Results unambiguously demonstrated that polyampholytes cryopreserve cells by strongly interacting with cell membrane, with hydrophobicity increasing the affinity for membrane interaction, which enables it to protect the membrane from various freezing-induced damages. Additionally, cryoprotective polymers, especially their hydrophobic derivatives, inhibit the recrystallization of ice, thus averting cell death. Hence, our results provide an important insight into the complex mechanism of cryopreservation, which might facilitate the rational design of polymeric CPAs with improved efficiency.
Co-reporter:Robin Rajan and Kazuaki Matsumura
Journal of Materials Chemistry A 2015 vol. 3(Issue 28) pp:5683-5689
Publication Date(Web):19 Jun 2015
DOI:10.1039/C5TB01021G
We report the novel one-step synthesis of a zwitterionic polymer, polysulfobetaine, via living reversible addition fragmentation chain transfer (RAFT) polymerization. Lysozyme did not aggregate when heated in the presence of this polymer. Amyloid formation, the cause of many diseases, was also suppressed. The zwitterionic polymer was significantly more efficient than previously described inhibitors of protein aggregation. Lysozyme heated in the presence of polysulfobetaine retained its solubility and very high enzymatic efficiency, even after prolonged heating. The secondary structures of lysozyme change with increasing temperature, accompanied by an increase in the β-structure. This change was prevented by mixing the polymer with lysozyme. 1H-NMR before and after aggregation revealed the conformational changes taking place in the lysozyme: during aggregation, lysozyme is transformed into a random coil conformation, thus losing its secondary structure. Presence of the polymer facilitates retention of partial higher order structures and lysozyme solubility at higher temperatures. The high efficiency of the polyampholyte was ascribed to its ability to prevent collisions between aggregating species by acting as a molecular shield.
Co-reporter:Minkle Jain, Robin Rajan, Suong-Hyu Hyon and Kazuaki Matsumura
Biomaterials Science 2014 vol. 2(Issue 3) pp:308-317
Publication Date(Web):25 Nov 2013
DOI:10.1039/C3BM60261C
Hydrogels are promising substrates for tissue engineering applications because of their unique biocompatibility, flexible methods of synthesis, range of constituents, and desirable physical characteristics. Cryopreservation of cell-containing constructs using such hydrogel scaffolds is in high demand in tissue-engineering applications for the production of “off-the-shelf” tissue-engineered products. However, cryopreservation of regenerated tissues including cell sheets and cell constructs is not easy compared to the preservation of cell suspensions, even when cryoprotectants are used. Here, we report a dextran-based polyampholyte hydrogel that itself shows cryoprotective properties, which could be useful for cell encapsulation and tissue engineering applications involving hydrogel formation. Amination was performed by introducing poly-L-lysine onto azide groups conjugated with dextran, and a portion of the amino groups was converted into carboxyl groups. These dextran-based polyampholytes showed good cryoprotective properties for mammalian cells, and the addition of dextran substituted with dibenzylcyclooctyne acid induced in situ hydrogel formation via Cu-free click chemistry with high biocompatibility. Cells encapsulated with such in situ hydrogels can be cryopreserved well without the addition of any cryoprotectants. Thus, these hydrogels can serve as scaffolds with cryoprotective properties that also provide structural integrity to tissue constructs.
Co-reporter:Sana Ahmed, Fumiaki Hayashi, Toshio Nagashima, Kazuaki Matsumura
Biomaterials 2014 35(24) pp: 6508-6518
Publication Date(Web):
DOI:10.1016/j.biomaterials.2014.04.030
Co-reporter:Robin Rajan and Kazuaki Matsumura
Journal of Materials Chemistry A 2015 - vol. 3(Issue 28) pp:NaN5689-5689
Publication Date(Web):2015/06/19
DOI:10.1039/C5TB01021G
We report the novel one-step synthesis of a zwitterionic polymer, polysulfobetaine, via living reversible addition fragmentation chain transfer (RAFT) polymerization. Lysozyme did not aggregate when heated in the presence of this polymer. Amyloid formation, the cause of many diseases, was also suppressed. The zwitterionic polymer was significantly more efficient than previously described inhibitors of protein aggregation. Lysozyme heated in the presence of polysulfobetaine retained its solubility and very high enzymatic efficiency, even after prolonged heating. The secondary structures of lysozyme change with increasing temperature, accompanied by an increase in the β-structure. This change was prevented by mixing the polymer with lysozyme. 1H-NMR before and after aggregation revealed the conformational changes taking place in the lysozyme: during aggregation, lysozyme is transformed into a random coil conformation, thus losing its secondary structure. Presence of the polymer facilitates retention of partial higher order structures and lysozyme solubility at higher temperatures. The high efficiency of the polyampholyte was ascribed to its ability to prevent collisions between aggregating species by acting as a molecular shield.
Co-reporter:Minkle Jain, Robin Rajan, Suong-Hyu Hyon and Kazuaki Matsumura
Biomaterials Science (2013-Present) 2014 - vol. 2(Issue 3) pp:NaN317-317
Publication Date(Web):2013/11/25
DOI:10.1039/C3BM60261C
Hydrogels are promising substrates for tissue engineering applications because of their unique biocompatibility, flexible methods of synthesis, range of constituents, and desirable physical characteristics. Cryopreservation of cell-containing constructs using such hydrogel scaffolds is in high demand in tissue-engineering applications for the production of “off-the-shelf” tissue-engineered products. However, cryopreservation of regenerated tissues including cell sheets and cell constructs is not easy compared to the preservation of cell suspensions, even when cryoprotectants are used. Here, we report a dextran-based polyampholyte hydrogel that itself shows cryoprotective properties, which could be useful for cell encapsulation and tissue engineering applications involving hydrogel formation. Amination was performed by introducing poly-L-lysine onto azide groups conjugated with dextran, and a portion of the amino groups was converted into carboxyl groups. These dextran-based polyampholytes showed good cryoprotective properties for mammalian cells, and the addition of dextran substituted with dibenzylcyclooctyne acid induced in situ hydrogel formation via Cu-free click chemistry with high biocompatibility. Cells encapsulated with such in situ hydrogels can be cryopreserved well without the addition of any cryoprotectants. Thus, these hydrogels can serve as scaffolds with cryoprotective properties that also provide structural integrity to tissue constructs.
Co-reporter:Monika Patel, Tatsuo Kaneko and Kazuaki Matsumura
Journal of Materials Chemistry A 2017 - vol. 5(Issue 19) pp:NaN3497-3497
Publication Date(Web):2017/04/12
DOI:10.1039/C7TB00701A
Precise and controlled drug delivery systems are required to facilitate effective therapeutics. To address this need, we devised a micelle–hydrogel composite based on amphiphilic polypeptides as a general carrier model for the switchable and controlled release of dual drugs. Two different di-block polypeptides, poly(L-lysine-b-L-phenylalanine) and poly(L-glutamic acid-b-L-phenylalanine) (PGA–PPA), were synthesized to form distinct self-assembling micellar systems that were loaded with curcumin and amphotericin B, respectively, as model drugs. The drug-loaded micellar mixture was crosslinked utilizing the pendant amino groups of the L-lysine side chains via genipin to yield a micelle–hydrogel composite with PGA–PPA micelles trapped in an interlinked hydrogel system. This composite allowed for controlled multiphasic drug release and could be effectively tuned to moderate the pace and amount of drug release and be easily regulated to switch the drug release kinetics over a range of simple factors such as change in pH, cross-linking density, and composition.