Yoshihiro Sasaki

Kyoto University, Kioto, Kyōto, Japan

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Publications (55)156.23 Total impact

  • [Show abstract] [Hide abstract]
    ABSTRACT: Nanogels are nanometer-sized hydrogel nanoparticles with three-dimensional networks of crosslinked polymer chains. We have previously reported a method for constructing protein nanogels using vitamin B6 (pyridoxal)-bearing pullulan (PLPP) as a bio-crosslinker. To validate the potential for using this bio-crosslinker to prepare protein nanogels, we developed an improved method using metal ions to enhance the affinity between the protein and the bio-crosslinker. We investigated the binding of PLPP as a bio-crosslinker to a protein to produce protein nanogels using anionic bovine serum albumin (BSA) as a model protein. Ultraviolet-visible spectroscopic titration of PLPP with BSA showed that the binding of the PLPP to the BSA via the formation of Schiff bases was significantly enhanced in the presence of zinc ions because of the coordination of the Schiff base to metal ions, which shifted the equilibrium of the reaction to the Schiff base formation side. Dynamic light scattering measurements, high-performance liquid chromatography and transmission electron microscopy confirmed that the zinc ions enhanced the ability of PLPP to form nanogels by crosslinking with the anionic BSA despite the strong electrostatic repulsion between the two molecules.
    Polymer Journal 02/2015; 47:201-205. DOI:10.1038/pj.2014.120 · 1.55 Impact Factor
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    ABSTRACT: We developed a new self-assembled amphiphilic nanogel-crosslinked porous (NanoCliP) gel that can trap proteins, liposomes, and cells. The NanoCliP gel was prepared by Michael addition of a self-assembled nanogel of acryloyl group-modified cholesterol-bearing pullulan to pentaerythritol tetra (mercaptoethyl) polyoxyethylene, followed by freezing-induced phase separation. Dynamic rheological analysis revealed that the storage modulus (G′) of the NanoCliP gel was approximately 10 times greater than that of a nonporous nanogel-crosslinked gel. Two-photon excitation deep imaging revealed that the NanoCliP gel comprises interconnected pores of several hundred micrometers in diameter. The NanoCliP gel trapped proteins and liposomes via hydrophobic interactions because its amphiphilic nanogels exhibit chaperone-like activity. Mouse embryonic fibroblasts penetrated the interconnected pores and adhered to the porous surface of fibronectin-complexed NanoCliP gel. In vivo, the NanoCliP gel enhanced cell infiltration, tissue ingrowth, and neovascularization without requiring exogenous growth factors, suggesting that the NanoCliP gel is a promising scaffold for tissue engineering.
    Biomaterials 10/2014; 37. DOI:10.1016/j.biomaterials.2014.10.045 · 8.31 Impact Factor
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    ABSTRACT: Apert syndrome is an autosomal dominantly inherited disorder caused by missense mutations in fibroblast growth factor receptor 2 (FGFR2). Surgical procedures are frequently required to reduce morphological and functional defects in patients with Apert syndrome; therefore, the development of noninvasive procedures to treat Apert syndrome is critical. Here we aimed to clarify the etiological mechanisms of craniosynostosis in mouse models of Apert syndrome and verify the effects of purified soluble FGFR2 harboring the S252W mutation (sFGFR2IIIcS252W) on calvarial sutures in Apert syndrome mice in vitro. We observed increased expression of Fgf10, Esrp1, and Fgfr2IIIb, which are indispensable for epidermal development, in coronal sutures in Apert syndrome mice. Purified sFGFR2IIIcS252W exhibited binding affinity for fibroblast growth factor (Fgf) 2 but also formed heterodimers with FGFR2IIIc, FGFR2IIIcS252W, and FGFR2IIIbS252W. Administration of sFGFR2IIIcS252W also inhibited Fgf2-dependent proliferation, phosphorylation of intracellular signaling molecules, and mineralization of FGFR2S252W-overexpressing MC3T3-E1 osteoblasts. sFGFR2IIIcS252W complexed with nanogels maintained the patency of coronal sutures, whereas synostosis was observed where the nanogel without sFGFR2S252W was applied. Thus, based on our current data, we suggest that increased Fgf10 and Fgfr2IIIb expression may induce the onset of craniosynostosis in patients with Apert syndrome and that the appropriate delivery of purified sFGFR2IIIcS252W could be effective for treating this disorder.
    PLoS ONE 07/2014; 9(7):e101693. DOI:10.1371/journal.pone.0101693 · 3.53 Impact Factor
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    ABSTRACT: The effects of the supramolecular polyrotaxane (PRX) structure on cellular internalization are investigated by flow cytometry and confocal laser scanning microscopy. AF-545-labeled aminated PRXs (APRXs) containing different numbers of threaded α-cyclodextrins (CDs) and amino groups are synthesized; their cellular uptakes are analyzed using HeLa cells in serum. The APRX threaded CD number is discovered to be a more critical factor for enhancing cellular internalization than the APRX amine content. Additionally, APRXs are demonstrated to be more easily internalized than conventional linear cationic macromolecules. Because increased numbers of threaded CDs are related to increased PRX rigidity, the PRX rigid frame resulting from CD molecules threaded on a poly(ethylene glycol) (PEG) chain is suitable for intracellular tools in therapy and diagnosis.
    Macromolecular Bioscience 03/2014; 14(3). DOI:10.1002/mabi.201300198 · 3.65 Impact Factor
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    ABSTRACT: Enzymatically synthesized glycogen (ESG) bearing hydrophobic groups were prepared to construct a new artificial molecular chaperone system. The radius of the amphiphilic ESG nanogel was approximately 15 nm. The nanogels prevented irreversible aggregation of carbonic anhydrase. Cyclodextrin induced the release of carbonic anhydrase in its active form via a one-step mechanism without dissociation of the nanogels.
    RSC Advances 10/2013; 3(48):25716–25718. DOI:10.1039/C3RA44572K · 3.71 Impact Factor
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    ABSTRACT: The intracellular delivery of enzymes is an essential methodology to extend their therapeutic application. Herein, we have developed dissociable supermolecule-enzyme polyelectrolyte complexes based on reduction-cleavable cationic polyrotaxanes (PRXs) for the reactivation of delivered enzymes. These PRXs are characterized by their supramolecular frameworks of a polymeric chain threading into cyclic molecules, which can form polyelectrolyte complexes with anionic enzymes while retaining their three dimensional structure, although their enzymatic activity is reduced. Upon the addition of a reductant, the PRXs dissociate into their constituent molecules and release the enzymes, resulting in a complete recovery of enzymatic activity. Under the intracellular environment, the PRX-based enzyme complexes showed the highest intracellular enzymatic activity and efficient activation of anticancer prodrugs to induce cytotoxic effects in comparison with the non-dissociable complexes and the commercial cell-penetrating peptide-based reagents. Thus, the intracellularly dissociable supermolecules are an attractive system for delivering therapeutic enzymes into living cells.
    Scientific Reports 07/2013; 3:2252. DOI:10.1038/srep02252 · 5.08 Impact Factor
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    ABSTRACT: Polysaccharide nanogels are one of the most attractive carriers for drug delivery systems. Nanogels encapsulate proteins in their hydrated polymer networks, and minimize the denaturation of proteins. In this study, we demonstrated the cross-linking of acryloyl group-modified polysaccharide nanogels via photopolymerization, which allowed the formation of novel hydrogel particles and macrogels. The mechanical properties of the resultant hydrogels depended on the concentrations of the nanogels and the cross-linkers. The most significant property of the nanogel-cross-linked hydrogel was the ability to encapsulate insulin via hydrophobic interactions. After incubation of the hydrogel containing insulin in water, the hydrogel was degraded by hydrolysis, and insulin was gradually released from the hydrogels over a period of 1 week. According to these results, this nanogel-cross-linked hydrogel prepared via photopolymerization has potential for innovative biomaterials.
    Reactive and Functional Polymers 07/2013; 73(7):958–964. DOI:10.1016/j.reactfunctpolym.2013.02.002 · 2.82 Impact Factor
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    ABSTRACT: Cationic lipoplexes, which consist of cationic lipids and DNA, are widely used in gene transfection. Recently, some reports suggest that the transfection efficiency of lipoplexes is affected by its formulation and physicochemical properties. To improve the lipoplex transfection efficiency, we examined gene transfection using lipoplexes treated with high hydrostatic pressure (HHP) that affects the phase behavior of the phospholipid bilayer. Lipoplexes were prepared with different cationic lipid to DNA ratios and then subjected to hydrostatic pressure at 5,000 and 10,000 atm. Non-pressurized lipoplex and HHP-treated lipoplex (after removing pressure) were evaluated. Dynamic light scattering measurement showed that the size of HHP-treated lipoplex increased compared with non-pressurized lipoplex, suggesting that formulation change and assembly of lipoplexes occurred under high hydrostatic pressure. Lipoplexes with and without HHP treatment were transferred into HeLa and COS-7 cells. The transfection efficiency of HHP-treated lipoplex increased compared with non-pressurized lipoplex, irrespective of the mixture ratio. Also, the transfection efficiency of HHP-treated lipoplex remained higher for several days. These results suggest that high hydrostatic pressure induces structural change in lipoplex, thereby affecting the transfection efficiency.
    01/2013; 2:80-83. DOI:10.14326/abe.2.80
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    ABSTRACT: Cationic liposome (CL) is a promising vector for nucleic acid therapy. In the present study, we investigated the effect of high hydrostatic pressure (HHP) treatment to lipoplex on the lipoplex-based antisense oligodeoxynucleotides (AS-ODNs) delivery in order to improve the transfection efficacy of lipoplex. Cationic liposome consisting of DOTMA and DOPE was used. AS-ODNs were designed to inhibit the expression of firefly luciferase. The complexes of CL and AS-ODN were prepared at various C/A ratios and then pressurized hydrostatically at various atmospheres (∼10,000 atm) for 10 min (HHP treatment). After removal of pressure, the pressurized lipoplexes were used. The lipoplex with and without the HHP treatment was transferred into HeLa cells expressing firefly luciferase transiently. The luciferase activity using the HHP-treated lipoplex was decreased compared to that of the non-pressurized lipoplex. Also, for HEK293 cells expressing luciferase stably, the lipoplex with the HHP treatment could effectively suppress the luciferase expression. In order to elucidate relationship between the structure and the transfection efficiency of the HHP-treated lipoplex, the properties of the HHPtreated lipoplex were examined by various physicochemical analyses. The different physicochemical properties between the lipoplexes with and without HHP treatment were showed, suggesting that the nature of lipoplex was changed by the HHP treatment. We believe that this change of lipoplex properties by the HHP treatment affected the efficiency of gene suppression. This HHP treatment for lipoplex appears to be a promising contribution to gene and oligonucleotide delivery.
    MRS Online Proceeding Library 01/2013; 1498. DOI:10.1557/opl.2013.411
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    Masaru Mukai, Yoshihiro Sasaki, Jun-ichi Kikuchi
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    ABSTRACT: A nanosensory membrane device was constructed for detecting liposome fusion through changes in an enzymatic activity. Inspired by a biological signal transduction system, the device design involved functionalized liposomal membranes prepared by self-assembly of the following molecular components: a synthetic peptide lipid and a phospholipid as matrix membrane components, a Schiff's base of pyridoxal 5'-phosphate with phosphatidylethanolamine as a thermo-responsive artificial receptor, NADH-dependent L-lactate dehydrogenase as a signal amplifier, and Cu(2+) ion as a signal mediator between the receptor and enzyme. The enzymatic activity of the membrane device was adjustable by changing the matrix lipid composition, reflecting the thermotropic phase transition behavior of the lipid membranes, which in turn controlled receptor binding affinity toward the enzyme-inhibiting mediator species. When an effective fusogen anionic polymer was added to these cationic liposomes, membrane fusion occurred, and the functionalized liposomal membranes responded with changes in enzymatic activity, thus serving as an effective nanosensory device for liposome fusion detection.
    Sensors 12/2012; 12(5):5966-77. DOI:10.3390/s120505966 · 2.05 Impact Factor
  • Polymers 12/2012; 4(4):1056-1064. DOI:10.3390/polym4021056 · 2.51 Impact Factor
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    ABSTRACT: New hybrid poly(ethylene glycol) (PEG) hydrogels crosslinked with both nanogels and nanogel-coated liposome complexes are obtained by Michael addition of the acryloyl group of a cholesterol-bearing pullulan (CHP) nanogel to the thiol group of pentaerythritol tetra(mercaptoethyl) polyoxyethylene. The nanogel-coated liposome complex is stably retained after gelation and the complexes are well dispersed in the hybrid gel. Microrheological measurements show that the strength and gelation time of the hybrid hydrogel can be controlled by changing the liposome:nanogel ratio. The hydrogel is gradually degraded by hydrolysis under physiological conditions. In this process, the nanogel is released first, followed by the nanogel-coated liposomes. Hybrid hydrogels that can incorporate various molecules into the nanogel and liposomes, and release them in a two-step controllable manner, represent a new functional scaffold capable of delivering multiple drugs, proteins or DNA.
    11/2012; 1(6):722-8. DOI:10.1002/adhm.201200175
  • Yoshihiro Sasaki, Kazunari Akiyoshi
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    ABSTRACT: Nanogels are polymer nanoparticles with three-dimensional networks. Recently, various nanogels have been designed, with a particular focus on biomedical applications. In this review, we describe recent progress in the synthesis of functional nanogels by self-assembly of associating polymers and nanogel engineering for advanced biomedical technology including regenerative medicine and drug delivery systems.
    ChemInform 07/2012; 43(31). DOI:10.1002/chin.201231265
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    ABSTRACT: Novel organic–inorganic hybrid nanoparticles consisting of polymer–hydrogel nanoparticles (nanogels) and iron oxide were developed for potential biomedical applications. Hybrid nanoparticles were prepared by a simple procedure using polysaccharide nanogels as a reactive site for iron oxide formation. The hybrid nanoparticles have a narrow size distribution with a diameter of approximately 30 nm and show high colloidal stability. These nanohybrid particles could be used as a contrast medium for magnetic resonance imaging or for magnetic hyperthermia therapy.
    Colloid and Polymer Science 06/2012; 291(6). DOI:10.1007/s00396-012-2868-7 · 2.41 Impact Factor
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    ABSTRACT: We propose a simple method to prepare lipid nanotubes, which can be used to transport biological molecules. By applying shear stress to surface-immobilized liposomes on a solid substrate, we obtained long lipid nanotubes arranged in a well-controlled direction.
    RSC Advances 03/2012; 2(7):2682-2684. DOI:10.1039/C2RA00629D · 3.71 Impact Factor
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    ABSTRACT: A supramolecular system that can activate an enzyme through photo-isomerization was constructed by using a liposomal membrane scaffold. The design of the system was inspired by natural signal transduction systems, in which enzymes amplify external signals to control signal transduction pathways. The liposomal membrane, which provided a scaffold for the system, was prepared by self-assembly of a photoresponsive receptor and a cationic synthetic lipid. NADH-dependent L-lactate dehydrogenase, the signal amplifier, was immobilized on the liposomal surface by electrostatic interactions. Recognition of photonic signals by the membrane-bound receptor induced photo-isomerization, which significantly altered the receptor's metal-binding affinity. The response to the photonic signal was transmitted to the enzyme by Cu(2+) ions. The enzyme amplified the chemical information through a catalytic reaction to generate the intended output signal.
    Chemistry - A European Journal 03/2012; 18(11):3258-63. DOI:10.1002/chem.201103552 · 5.70 Impact Factor
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    ABSTRACT: Molecular chaperones selectively trap heat-denatured proteins or their intermediates, primarily by hydrophobic interactions, to prevent irreversible aggregation resulting from macromolecular host (molecular chaperone)-guest (protein) interactions. The molecular chaperone function is an important concept that is expected to lead to breakthroughs in drug delivery systems, especially for protein or peptide delivery in regenerative medicine, such as bone regeneration. We have reported that polysaccharide nanogels act as artificial molecular chaperones. To further clarify the molecular chaperone function of nanogels as protein carriers, the elucidation of nanogel-protein interactions are especially important. Here, we investigated the interaction of a protein with a polysaccharide nanogel using fluorescence correlation spectroscopy at variable temperatures, using fluorescence-labeled bovine serum albumin (BSA) as a model protein. In particular, thermodynamic parameters of the heat-induced complexation of protein with CHP nanogels were evaluated using the van't Hoff plot. The plot shows that the CHP nanogels strongly complexed with heat-denatured BSA. The increased hydrophobicity of the denatured, unfolded protein may prefer complexation with amphiphilic hydrogel nanoparticles over complexation with the completely folded native protein. Thermodynamic parameters suggest that the complexation is entropically driven, rather than enthalpically, under the conditions studied.
    Current Drug Discovery Technologies 06/2011; 8(4):308-13. DOI:10.2174/157016311798109371
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    ABSTRACT: Cell-free protein synthesis is a promising technique for the rapid production of proteins. However, the application of the cell-free systems requires the development of an artificial chaperone that prevents aggregation of the protein and supports its correct folding. Here, nanogel-based artificial chaperones are introduced that improve the folding efficiency of rhodanese produced in cell-free systems. Although rhodanese suffers from rapid aggregation, rhodanese was successfully expressed in the presence of the nanogel and folded to the enzymatically active form after addition of cyclodextrin. To validate the general applicability, the cell-free synthesis of ten water-soluble proteins was examined. It is concluded that the nanogel enables efficient expression of proteins with strong aggregation tendency.
    Macromolecular Bioscience 06/2011; 11(6):814-20. DOI:10.1002/mabi.201000457 · 3.65 Impact Factor
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    ABSTRACT: Protein-crosslinked nanogels were prepared by introducing vitamin B6 (pyridoxal) to hydrophilic polysaccharide. The pH dependence of Schiff base formation was utilized to generate a pH-sensitive nanogel system. This approach represents a new method to prepare hybrid nanogels crosslinked with biomolecules.
    Polymer Chemistry 04/2011; 2(6):1267-1270. DOI:10.1039/C1PY00100K · 5.37 Impact Factor
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    ABSTRACT: Molecular communication is a novel communication paradigm inspired by information processing in biological systems using molecules as information medium. In this study, we achieved the selective propagation of transport vesicles which is an essential technique to establish a molecular communication system using artificial cells. The hybridisation of DNA was employed to conduct a signal-induced delivery of transport vesicles from a sender to a receiver vesicle formed with phospholipids. Oligonucleotide lipids each with a different single-stranded DNA, which acts as recognition tags, were embedded in the sender, receiver and transport vesicles. The addition of a DNA signal with a complementary sequence to connect two oligonucleotide lipids induced the assembly of two types of large unilamellar vesicles as well as recombination of the vesicles. In addition, the selective delivery of transport vesicles between sender and receiver giant vesicles is discussed, based on the results of fluorescence microscopic observations.
    Supramolecular Chemistry 03/2011; 23(3-4-3–4):218-225. DOI:10.1080/10610278.2010.521835 · 2.13 Impact Factor

Publication Stats

521 Citations
156.23 Total Impact Points

Institutions

  • 2012–2014
    • Kyoto University
      • • Department of Polymer Chemistry
      • • Department of Synthetic Chemistry and Biological Chemistry
      Kioto, Kyōto, Japan
  • 2008–2014
    • Tokyo Medical and Dental University
      • Institute of Biomaterials and Bioengineering
      Edo, Tōkyō, Japan
  • 2000–2010
    • Nara Institute of Science and Technology
      • Graduate School of Materials Science
      Ikuma, Nara, Japan
  • 2005
    • Saitama Institute of Technology
      Saitama, Saitama, Japan