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Hydroxy group containing methacrylamide polymers exhibited excellent permeability into the hepatocyte multicellular tumor spheroids.

Thermoresponsive self-assembled nanogels were conveniently prepared by cholesterol end-capped poly(N-isopropylacrylamide) (PNIPAM) in water. Both cholesterol end-capped PNIPAMs (telelchelic cholesterol PNIPAM, tCH-PNIPAM) formed flower-like nanogels by the self-assembling of four to five polymer chains with multiple domains of cholesterol in water at 20 °C. Meanwhile, one end-group cholesterol-capped PNIPAM (semitelechelic cholesterol PNIPAM, stCH-PNIPAM) was also formed as a nanogel by the self-assembling of 15-20 polymer chains with 3 to 4 cholesterol domains. The hydrophobic cholesterol domains of tCH-PNIPAM nanogels were maintained above the lower critical solution temperature (LCST) of PNIPAM (>32 °C). Differently, the hydrophobic domains of stCH-PNIPAM were disrupted by cholesterol-free PNIPAM chain ends and formed large mesoglobules above the LCST. These transition controls of hydrophilic end-capped smart polymers may open new methodologies to design thermoresponsive nanosystems.

Thermoresponsive polymers that possess both UCST- and LCST-like behaviors have generally been designed using diblock copolymers that are mostly composed of an LCST-like polymer and a UCST polymer. Herein, we prepared an LCST-UCST-type polymer composed of UCST-like thermoresponsive zwitterionic sulfabetaine methacrylate and nonthermoresponsive PEG methacrylate, ZB-PEG, by reversible addition-fragmentation chain transfer (RAFT) copolymerization. By adjusting the PEG composition, ZB-PEG formed a mesoglobule, a microglobule, and the dissociated states in phosphate-buffered saline (PBS). These states were found to be reversible via temperature control. Moreover, this behavior showed high reversibility and succeeded in stabilizing horseradish peroxidase (HRP) in the dilute condition. Such thermoresponsive ZB-PEG can be applied over a wide range of applications in biotechnology and other fields.

Understanding the interactions of soft nanomatters with cell membranes is particularly important for research into nanocarrier-based drug delivery systems, cell engineering, and subcellular imaging. Most nanoparticles, vesicles, micelles, and polymeric aggregates are internalized into endosomes and, eventually, lysosomes in the cytosol because of energy-dependent endocytic processes. Endocytic uptake substantially limits the access to the cytoplasm where a cargo agent acts. Bypassing the endocytic pathways by direct penetration into plasma membrane barriers would enhance the efficacy of nanomedicines. Some zwitterionic polymer nanoaggregates have been shown to permeate into the cell interior in an energy-independent manner. We have elucidated this phenomenon by observing changes in the biomembrane barrier functions against protons as the smallest indicator and have used these results to further design and develop poly(betaines). In this work, we investigated the translocation mechanisms for a series of zwitterionic poly(methacrylamide) and poly(methacrylate) species bearing a pyridinium propane sulfonate moiety in the monomers. Minor differences in the monomer structures and functional groups were observed to have dramatic effects on the interaction with plasma membranes during translocation. The ability to cross the plasma membrane involves a balance among the betaine dipole-dipole interaction, NH-π interaction, π-π interaction, cation-π interaction, and amide hydrogen bonding. We found that the cell-penetrating polysulfobetaines had limited or no detrimental effect on cell proliferation. Our findings enhance the opportunity to design and synthesize soft nanomatters for cell manipulations by passing across biomembrane partitions.

Thermoresponsive self-assembled nanogels were conveniently prepared by cholesterol end-capped poly(

Three-dimensional cell aggregates (spheroids) are becoming a research focus because their construction is similar to that in vivo microenvironments, enabling the acceleration of drug discovery and reducing the need for animal tests, and other advantages. However, the delivery of drugs to the inside of spheroids is time-consuming and has low efficiency. In this study, we selected a sulfobetaine copolymer that translocates to the cell membrane in monolayer cultured cells as a nanocarrier of anticancer drugs. Doxorubicin (Dox) and 17-demethoxy-17-allylamino geldanamycin (17AAG) were modified to the copolymer of sulfobetaine methacrylate and poly(ethylene glycol) methacrylate, P(SB-PEG), and added to glioblastoma A-172 cell spheroids. Dox-P(SB-PEG) showed fast permeation into A-172 spheroids, and the fluorescence in cells was observed in the center area of the spheroids within 1 h of polymer addition. Conversely, only the outer one to two cell layers of spheroids were observed when Dox was added to the spheroids. Dox-P(SB-PEG) in A-172 spheroids was localized in the mitochondria of each cell and exhibited comparable drug efficacy to that of Dox in growth inhibition assays of A-172 spheroids. Moreover, approximately 10-fold higher drug efficacy in growth inhibition and invasion of A-172 spheroids was found using 17AAG-P(SB-PEG). Conjugating anticancer drugs with P(SB-PEG) is a promising strategy to enhance drug permeation and efficacy against spheroid cells.

The inhalation of silica dust is associated with fibrosis and lung cancer, which are triggered by macrophage inflammatory responses; however, how macrophages recognize silica remains largely unknown. Here, we identify by functional expression cloning the class B scavenger receptor SR-B1 as a silica receptor. Through an extracellular α-helix, both mouse and human SR-B1 specifically recognized amorphous and crystalline silica, but not titanium dioxide nanoparticles, latex nanoparticles, or monosodium urate crystals, although all particles exhibited negative surface potentials. Genetic deletion of SR-B1 and masking of SR-B1 by monoclonal antibodies showed that SR-B1-mediated recognition of silica is associated with caspase-1-mediated inflammatory responses in mouse macrophages and human peripheral blood monocytes. Furthermore, SR-B1 was involved in silica-induced pulmonary inflammation in mice. These results indicate that SR-B1 is a silica receptor associated with canonical inflammasome activation.

Investigation of the rotational motion of a fluorescent probe tethered to a protein helps to elucidate the local properties of the solvent and protein near the conjugation site of the probe. In this study, we have developed an instrument for frequency-domain fluorescence (FDF) anisotropy measurements, and studied how the local properties around a protein, actin, can be elucidated from the rotational motion of a dye tethered to actin. Rhodamine 6G (R6G) was attached to Cys-374 using newly-synthesized R6G-maleimide with three different oligo(ethylene glycol) (OEG) linker lengths. The time-resolved anisotropy decay of R6G tethered to G-actin was revealed to be a combination of the two modes of the wobbling motion of R6G and the tumbling motion of G-actin. The rotational diffusion coefficient (RDC) of R6G wobbling was ~0.1 ns(-1) at 20°C and increased with OEG linker length. The use of the three R6G-actin conjugates of different linker lengths was useful to not only figure out the linker length dependence of the rotational motion of R6G but also validate the analyses. In the presence of a cosolvent of glycerol, although the tumbling motion of G-actin was retarded in response to the bulk viscosity, the wobbling motion of R6G tethered to actin exhibited an increase of RDC as glycerol concentration increased. This finding suggests an intricate relationship between the fluid properties of the bulk solvent and the local environment around actin.

Fast intracellular migration and controlled localization of molecules represent significant challenges for future applications of drug discovery and related fields. In this study, thermoresponsive sulfobetaine polymers with pyridinium cations are evaluated as biocompatible and mitochondria-localizing agents. Among the polymers, poly(3-(4-(2-methacrylamido)ethyl pyridinio-1-yl)propane-1-sulfonate), P(E-PySMAAm)14k (Mn = 14 000 g mol-1 ) exhibit thermoresponsiveness with an upper critical solution temperature like behavior in cell culture medium containing serum with minimal cytotoxicity. Upon the addition of P(E-PySMAAm)14k to HeLa cells at temperatures above the clearing point at 37 °C, effective localization is observed in mitochondria. However, increased intensity but nonspecific localization is observed below the clearing point at 4 °C. Doxorubicin is conjugated to the P(E-PySMAAm) and achieves effective mitochondrial delivery while maintaining drug efficacy. Such sulfobetaine polymers represent promising tools for intracellular delivery of molecules.

Mitochondrial targeting and entry, two crucial steps in fighting severe diseases resulting from mitochondria dysfunction, pose important challenges in current nanomedicine. Cell-penetrating peptides or targeting groups, such as Rhodamine-B (Rho), are known to localize in mitochondria, but little is known on how to enhance their effectiveness through structural properties of polymeric carriers. To address this issue, we prepared 8 copolymers of 3-dimethyl(methacryloyloxyethyl)ammonium propane sulfonate and poly(ethyleneglycol) methacrylate, p(DMAPS-ran-PEGMA) (molecular weight, 18.0 M n M n

AC voltage and shear flow are applied to self-assembled, multi-layered microspheres (MLMs) composed of a double hydrophilic block copolymer, poly(ethylene glycol)-block-poly(3-dimethyl(methacryloyloxyethyl) ammoniumpropane sulfonate), (PEG-SB) in order to control dynamic transformations in aqueous solution. Under application of AC voltage, the fusion of MLMs is accelerated by contact and the size grows from ∼1.5 μm to several tens of μm. The rate of growth is controllable by varying the frequency and the polymerization degree of the SB unit. Bursting of the fused MLMs is induced in a closed chamber and the average size of the bursting MLMs is 25 μm. On the other hand, MLMs are also fused under shear flow above 1.5 × 103 s−1. The fused MLMs grow to be a string-like structure with a length of more than 4 cm and the aspect ratio of >4000. The string-like structure splits into MLMs when the shear flow is removed. Such multiple dynamic transformations of MLMs can be applied to versatile droplets as microcargoes, microreactors, and microseparators in fully aqueous droplet microfluidics.

Silicon dioxide (SiO2) nanoparticles (NPs) and titanium dioxide (TiO2) NPs are the most widely used inorganic nanomaterials. Although the individual toxicities of SiO2 and TiO2 NPs have been extensively studied, the combined toxicity of these NPs is much less understood. In this study, we observed unexpected and drastic activation of the caspase-1 inflammasome and production of IL-1β in mouse bone marrow-derived macrophages stimulated simultaneously with SiO2 and TiO2 NPs at concentrations at which these NPs individually do not cause macrophage activation. Consistent with this, marked lung inflammation was observed in mice treated intratracheally with both SiO2 and TiO2 NPs. In macrophages, SiO2 NPs localized in lysosomes and TiO2 NPs did not; while only TiO2 NPs produced ROS, suggesting that these NPs induce distinct cellular damage leading to caspase-1 inflammasome activation. Intriguingly, dynamic light scattering measurements revealed that, although individual SiO2 and TiO2 NPs immediately aggregated to be micrometer size, the mixture of these NPs formed a stable and relatively monodisperse complex with a size of ~250 nm in the presence of divalent cations. Taken together, these results suggest that SiO2 and TiO2 NPs synergistically induce macrophage inflammatory responses and subsequent lung inflammation. Thus, we propose that it is important to assess the synergistic toxicity of various combinations of nanomaterials. Electronic supplementary material The online version of this article (doi:10.1186/s12989-017-0192-6) contains supplementary material, which is available to authorized users.

A double hydrophilic block copolymer, poly(ethylene glycol)-poly(3-dimethyl (methacryloyloxyethyl) ammonium propane sulfonate) (PEG-SB), is synthesized by reversible addition-fragmentation transfer (RAFT) polymerization using PEG methyl ether (4-cyano-4-pentanoate dodecyl trithiocarbonate) as a chain transfer agent. PEG-SB forms multi-layered microspheres with dipole-dipole interactions of the SB side chains as the driving force. The PEG-SB polymers show an upper critical solution temperature (UCST) and the UCST is controllable by the polymerization degree. The PEG-SB microspheres are dissociated above the UCST and then monodispersed microspheres (∼1 μm) are obtained when the solution temperature is decreased below the UCST again. The disassociation/association of the microspheres is also controllable using the concentration of NaCl. These multi-responsive microspheres could be a powerful tool in the field of nano-biotechnology.

We investigate the effects of urea and its derivatives on the ATPase activity and on the in vitro motility of chicken skeletal muscle actomyosin. Mg-ATPase rate of myosin subfragment-1 (S1) is increased by 4-fold by 0.3 M 1,3-diethylurea (DEU), but it is unaffected by urea, thiourea, and 1,3-dimethylurea at ≤ 1 M concentration. Thus, we further examine the effects of DEU in comparison to those of urea as reference. In in vitro motility assay, we find that in the presence of 0.3 M DEU, the sliding speeds of actin filaments driven by myosin and heavy meromyosin (HMM) are significantly decreased to 1/16 and 1/6.6, respectively, compared with the controls. However, the measurement of the actin-activated ATPase activity of HMM shows that the maximal rate, V max , is almost unchanged with DEU. Thus, the myosin-driven sliding motility of actin filaments is significantly impeded in the presence of 0.3 M DEU, whereas the cyclic interaction of myosin with F-actin occurs during the ATP turnover, the rate of which is close to that without DEU. In contrast to DEU, 0.3 M urea exhibits only modest effects on both actin-activated ATPase and sliding motility of actomyosin. Thus, DEU has the effect of uncoupling the sliding motility of actomyosin from its ATP turnover.

In this study, we prepared a new associating polymer, ChMaPLL, by the substitution of the poly(L-lysine) moiety with oligosaccharide amylose primer and cholesterol. ChMaPLL formed positively charged polypeptide nanogels (~50 nm) via self-assembly in water. The nanogels showed a secondary structural transition to an α-helix structure induced by poly(L-lysine) in response to an increase in pH. Oligosaccharides of the nanogels reacted with the phosphorylase a enzyme. Amylose-conjugated nanogels were obtained by enzymatic polymerization. The elongation of the saccharide chain shielded the positive charge of the nanogels. The multiresponsive polysaccharide-polypeptide hybrid nanogels might prove to be useful in the areas of biotechnology and biomedicine.

A self-assembled nanogel, derived from an acid-labile cholesteryl-modified pullulan (acL-CHP), was prepared by grafting vinyl ether-cholesterol substituents onto a 100 kD pullulan main chain polymer backbone. Stable nanogels are formed by acL-CHP self-assemblies at neutral pH. The hydrodynamic radius of the nanogels, observed to be 26.5 ± 5.1 nm at pH 7.0, increased by ~135% upon acidification of the solution to pH 4.0. SEC analysis of the acL-CHP nanogel at pH 4.0 showed that the grafts were nearly 80% degraded after 24 h, whereas little or no degradation was observed over the same time period for a pH stable analog (acS-CHP) at pH 4.0 or the acL-CHP at pH 7.0. Complexation of BSA with the acL-CHP nanogel was observed at pH 7.0 with subsequent release of the protein upon acidification. These findings suggest that stimuli-responsive, self-assembled nanogels can release protein cargo in a manner that is controlled by the degradation rate of the cholesterol-pullulan grafting moiety.

Understanding of the hydration state is an important issue in the chemomechanical energetics of versatile biological functions of polymerized actin (F-actin). In this study, hydration-state differences of F-actin by the bound divalent cations are revealed through precision microwave dielectric relaxation (DR) spectroscopy. G- and F-actin in Ca- and Mg-containing buffer solutions exhibit dual hydration components comprising restrained water with DR frequency f2 (fw: DR frequency of bulk solvent, 17 GHz at 20 °C) and hypermobile water (HMW) with DR frequency f1 (fw). The hydration state of F-actin is strongly dependent on the ionic composition. In every buffer tested, the HMW signal Dhyme (≡ (f1 - fw)δ1/(fwδw)) of F-actin is stronger than that of G-actin, where δw is DR-amplitude of bulk solvent and δ1 is that of HMW in a fixed-volume ellipsoid containing an F-actin and surrounding water in solution. Dhyme value of F-actin in Ca2.0-buffer (containing 2 mM Ca(2+)) is markedly higher than in Mg2.0-buffer (containing 2 mM Mg(2+)). Moreover, in the presence of 2 mM Mg(2+), the hydration state of F-actin is changed by adding a small fraction of Ca(2+) (∼0.1 mM) and becomes closer to that of the Ca-bound form in Ca2.0-buffer. This is consistent with the results of the partial specific volume and the Cotton effect around 290 nm in the CD spectra, indicating a change in the tertiary structure and less apparent change in the secondary structure of actin. The number of restrained water molecules per actin (N2) is estimated to be 1600-2100 for Ca2.0- and F-buffer and ∼2500 for Mg2.0-buffer at 10-15 °C. These numbers are comparable to those estimated from the available F-actin atomic structures as in the first water layer. The number of HMW molecules is roughly explained by the volume between the equipotential surface of -kT/2e and the first water layer of the actin surface by solving the Poisson-Boltzmann equation using UCSF Chimera.

The majority of nanoparticles designed for cellular delivery of drugs and imaging agents enter the cell via endocytotic pathways leading to their entrapment in endosomes that present a robust barrier to further trafficking of the nanoparticles within the cells. A few materials, such as the cell penetrating peptides (CPPs), are known to enter cells not only via endocytosis, but also via translocation through the cell membrane into the cytoplasm, successfully bypassing the endosomes. We report here that random copolymers of 3-dimethyl(methacryloyloxyethyl)ammonium propanesulfonate and poly(ethylene glycol) methacrylate, p(DMAPS-ran-PEGMA), are internalized in cells primarily via translocation through the cell membrane rather than endocytosis. The properties of the polymers and their modes of uptake were investigated systematically by dynamic light scattering, confocal fluorescence microscopy, and flow cytometry. Using specific inhibitors of the cellular uptake machinery in a human cervical carcinoma cell line (HeLa), we show that these nontoxic synthetic polyzwitterions exist in cell media as self-assembled nanospheres that unravel as they adsorb on the plasma membrane and translocate through it. Conjugates of p(DMAPS-ran-PEGMA) with rhodamine B were delivered selectively to the mitochondria, whereas doxorubicin (Dox)-p(DMAPS-ran-PEGMA) conjugates were accumulated in both the nucleus and the mitochondria, effectively inducing apoptosis in HeLa cells. These findings suggest that the noncytotoxic and readily synthesized p(DMAPS-ran-PEGMA) can find applications as bioimaging tools and drug nanocarriers.

This paper reports that water molecules around F-actin, a polymerized form of actin, are more mobile than those around G-actin or in bulk water. A measurement using pulse-field gradient spin-echo (1)H NMR showed that the self-diffusion coefficient of water in aqueous F-actin solution increased with actin concentration by ∼5%, whereas that in G-actin solution was close to that of pure water. This indicates that an F-actin/water interaction is responsible for the high self-diffusion of water. The local viscosity around actin was also investigated by fluorescence measurements of Cy3, a fluorescent dye, conjugated to Cys 374 of actin. The steady-state fluorescence anisotropy of Cy3 attached to F-actin was 0.270, which was lower than that for G-actin, 0.334. Taking into account the fluorescence lifetimes of the Cy3 bound to actin, their rotational correlation times were estimated to be 3.8 and 9.1ns for F- and G-actin, respectively. This indicates that Cy3 bound to F-actin rotates more freely than that bound to G-actin, and therefore the local water viscosity is lower around F-actin than around G-actin.

Polymer composites of magnetic particles are widely used as microwave absorbers. An effective method for obtaining thinner microwave absorbers for device mounting is to increase the volume fraction of magnetic nanoparticles within the composites such that the permeability is enhanced. In this study, composites were prepared using magnetite nanoparticles surface-modified with 4-META (4-methacryloylioxyethyl trimellitate anhydride) and cross-linked with PEG-4SH (pentaerythritol tetra-polyethylene glycol ether with four thiol-modified terminals). These composites have a higher volume fraction of nanoparticles (up to 62 vol%) and higher permeability than conventional epoxy resin composites. In addition, the prepared composites showed good microwave absorption properties (R:L :< � 20 dB) with a smaller matching thickness (d ¼ 8:0 mm) than the epoxy resin composites (d ¼ 9:0 mm). [doi:10.2320/matertrans.MAW201020]

Polymer composites of magnetic particles are widely used as microwave absorbers. An effective method for obtaining thinner microwave absorbers for device design is increasing the volume fraction of magnetic nanoparticles by enhancing the permeability of composites. In this study, composites were prepared using Ni-Zn ferrite nanoparticles surface-modified with 4-META (4-methacryloylioxyethyl trimellitate anhydride) and cross-linked with PEG-4SH (pentaerythritol tetra-polyethylene glycol ether with four thiol-modified terminals). These composites have a high volume fraction of nanoparticles (up to 72 vol%) and permeability (μr max″=5.9). In addition, the prepared composites showed good microwave absorption properties (R.L.

The cholesterol-bearing pullulan (CHP)-nanogels are able to trap hydrophobic drugs or proteins inside the nanogels, which is potential in application to drug delivery system and tissue engineering. On the other hand, prostaglandin E1 (PGE1) plays important roles in wound healing and PGE1 ointment has been clinically used to treat chronic skin ulcers and wounds. The purpose of this study is to evaluate effects of CHP nanogels in combination with prostaglandin E1 on wound healing in full thickness skin defect model. A square skin defect (1 x 1 cm(2)) of full thickness was created on the dorsal of Wistar rats. The wound was treated with CHP nanogels without PGE1 (CHP group) or CHP nanogels containing with PGE1 (CHP/PGE1 group) or PGE1 ointment (PGE1 ointment group). In both CHP/PGE1 and PGE1 ointment groups, approximately 6 microg of PGE1 was applied to each wound. In the control group, the wound was untreated. The wound was evaluated in measuring wound area and histologically. In CHP/PGE1 group, the rate of wound size reduction was significantly higher than the ones of other groups. Histologically, CHP/PGE1 promoted neoepithelialization, neovascularization, and wound closure compared to the other treatments. These results suggest that CHP in combination with PGE1 can promote wound healing, which confirms the efficiency of CHP nanogels-based drug delivery system.

A flexible biosensor using a phospholipid polymer to immobilization of glucose oxidase (GOD) was fabricated and tested. At first, an enzyme membrane formed by immobilizing GOD onto a porous polytetrafluoroethylene (PTFE) membrane using the phospholipid polymer (2-methacryloyloxyethyl phosphorylcholine (MPC) copolymerized with 2-ethylhexylmethacrylate (EHMA):PMEH) was evaluated. According to the result of amperometric measurement, average density of GOD to be immobilized was optimized to 38.9 units cm(-2). Temperature and pH dependences were also investigated. Then, a flexible glucose sensor was fabricated by immobilizing GOD onto a flexible hydrogen peroxide electrode using PMEH. The flexible glucose sensor showed a linear relationship between output currents and glucose concentration in 0.05-1.00 mmol L(-1), with a correlation coefficient of 0.999. The calibration range covered the normal tear glucose level of 0.14-0.23 mmol L(-1). This indicates that the flexible biosensor is considered to be useful for monitoring of glucose in tear fluids.

An enzyme-responsive artificial chaperone system which employs an amphiphilic amylose primer (dodecyl maltopentaose, C12-MP) as a surfactant and phosphorylase b was designed to enable protein refolding. Effective refolding of carbonic anhydrase B after both heat denaturation (70 degrees C for 10min) and guanidine hydrochloride (6M) denaturation was observed by controlled association between the protein molecules and the C12-MP primer micelle through an enzymatic reaction.

Rapidly shrinking poly(N-isopropyl acrylamide) (PNIPAM) hydrogels are prepared by crosslinking with self-assembled nanogels that consist of cholesteryl- and methacryloyl-substituted pullulan (CHPMA). The CHPMA nanogel (R h = 26.4 nm) was used as a crosslinker for a hydrophilic nanodomain. Transmission electron microscopy images of the nanogel-crosslinked PNIPAM hydrogel reveal a well-defined nanoporous structure. The nanogel-crosslinked PNIPAM hydrogel shows rapid shrinking based on its structure. The shrinking half-time was ≈2 min, which is about 3400 times faster than that of a PNIPAM hydrogel crosslinked by methylene(bisacrylamide).

An effective intracellular protein delivery system with self-assembled cationic nanogels is reported. Interaction of proteins with self-assembled nanogels of cationic cholesteryl group-bearing pullulans (CHPNH 2) was investigated by dynamic light scattering (DLS), transmission electron micrograph (TEM), fluorescence resonance energy transfer (FRET), and fluorescence correlation spectroscopy (FCS). The cationic nanogels strongly interacted with bovine serum albumin (BSA) and formed monodispersed nanoparticels (

Polymerizable nanogels were prepared by self-assembly of cholesteryl group-bearing pullulan (CHP) with methacryloyl groups (CHPMA). The CHPMA nanogel was polymerized with 2-methacryloyloxyethyl phosphorylcholine (MPC) by radical polymerization in dilute aqueous solution. The solution properties of the polymers in water were investigated by TEM, SEC-MALS, and fluorescence quenching technique. Monodispersed hybrid nanogels of CHPMA-MPC (CM nanogels) (25-30 nm in radius of gyration) were obtained by using CHPMA nanogel as a seed-nanogel. CM nanogels have a dual cross-linking structure that is physically cross-linked with the cholesteryl groups and chemically cross-linked with the MPC polymer chains. CM nanogels trap heat-denatured carbonic anhydrase B (CAB) and prevent their aggregations. The nanogels maintained the ability of trapping and releasing enzymes by host-guest interaction of cholesteryl group and cyclodextrin.

To obtain a hydrogel-like elastic membrane, we prepared semi-interpenetrating polymer networks (IPNs) by the radical polymerization of methacrylates such as 2-methacryloyloxyethyl phosphorylcholine (MPC), 2-hydroxyethylmethacrylate, and triethyleneglycol dimethacrylate diffused into segmented polyurethane (SPU) membranes swollen with a monomer mixture. The values of Young's modulus for the hydrated semi-IPN membranes were less than that for an SPU membrane because of higher hydration, but they were much higher than that for a hydrated MPC polymer gel (non-SPU). According to a thermal analysis, the MPC polymer influenced the segment association of SPU. The diffusion coefficient of 8-anilino-1-naphthalenesulfonic acid sodium salt from the semi-IPN membrane could be controlled with different MPC unit concentrations in the membrane, and it was about 7 × 102 times higher than that of the SPU membrane. Fibroblast cell adhesion on the semi-IPN membrane was effectively reduced by the MPC units. We concluded that semi-IPNs composed of the MPC polymer and SPU may be novel polymer materials possessing attractive mechanical, diffusive-release, and nonbiofouling properties. © 2002 Wiley Periodicals, Inc. J Polym Sci Part A: Polym Chem 41: 68–75, 2003

Segmented polyurethanes, (SPU)s, are widely used in the biomedical fields because of their excellent mechanical property. However, when blood is in contact with the SPU, non-specific biofouling on the SPU occurs which reduces its mechanical property. To obtain novel blood compatible elastomers, the surface of the SPU was modified with 2-methacryloyloxyethyl phosphorylcholine (MPC) by forming a semi-interpenetrating polymer network (semi-IPN). The SPU film modified by MPC polymer with the semi-IPN (MS-IPN film) was prepared by visible light irradiation of the SPU film in which the monomers were diffused. X-ray photoelectron spectroscopy confirmed that the MPC units were exposed on the MS-IPN film surface. The mechanical properties of the MS-IPN film characterized by tensile testing were similar to those of the SPU film. Platelet adhesion on MS-IPN films was also investigated before and after stress loading to determine the effects of the surface modification on the blood compatibility. Many platelets did adhere on the SPU film before and after stress loading. On the other hand, the MS-IPN film prevented platelet adhesion even after repeated stress loading.

Polymer membranes are widely used in biomedical applications such as hemodialysis, membrane oxygenator, etc. When the membranes come in contact with blood or body fluids, protein adsorption and cell adhesion occur rapidly. Nonspecific protein adsorption and cell adhesion on the membranes induce not only various bio-rejections but also a decrease in their performance. We hypothesized that a blood compatible gas-permeable membrane could be prepared from polyethylene (PE) porous membranes modified with phospholipid polymers. In this study, poly[(2-methacryloyloxyethyl phosphorylcholine) (MPC)-co-dodecyl methacrylate] (PMD) skin film adhered to a PE porous membrane (PMD/PE porous membrane) was prepared. Elution of PMD was not detected meaning that the PMD film did not detach from the PE porous membrane even after soaking in water for more than 6 months. The permeation coefficient of oxygen gas through the PE membrane with the adhered PMD containing more than 0.20 mole fraction of the MPC unit, was the same as that of the original PE porous membrane. The PMD surface effectively reduced biofouling. We concluded that the PMD/PE porous membrane is useful as a novel membrane oxygenator due to its excellent gas-permeability and blood compatibility.

A new class of artificial molecular transport system is constructed by polymeric microspheres. The microspheres are prepared by self-assembly of poly(ethylene glycol)-block-poly(3-dimethyl(methacryloyloxyethyl)ammonium propane sulfonate), PEG-b-PDMAPS, by intermolecular dipole-dipole interaction of sulfobetaine side chains in water. Below the upper critical solution temperature (UCST) of PEG-b-PDMAPS, the microspheres (∼1μm) interact with other microspheres by partial and transit fusion. In order to apply the interaction between microspheres, a 3'-TAMRA-labeled single-stranded DNA oligomer (ssDNA) is encapsulated into a PEG-b-PDMAPS microsphere by thermal treatment. The exchange of ssDNA between microspheres is confirmed by fluorescence resonance energy transfer (FRET) quenching derived from double-stranded formation with complementary 5'-BHQ-2-labeled ssDNA encapsulated in PEG-b-PDMAPS microspheres. The exchange rate of ssDNA is controllable by tuning the composition of the polymer. The contact-dependent transport of molecules can be applied in the areas of microreactors, sensor devices, etc.

Heat capacities of optically isotropic mesogens, BABH(8), ANBC(16) and ANBC(18), have been measured by adiabatic calorimetry. The existence of solid-solid phase transitions with large entropy chang...

2-Methacryloyloxyethyl phosphorylcholine (MPC) polymers, which have excellent biocompatibility, have been receiving increasing attention in biomedical and bioengineering fields; however, the mechanical strength of the hydrated MPC polymers is not sufficient for use in these fields as a bulk material. Therefore, we hypothesized that a novel material might be realized by reinforcing the MPC polymer network with segmented polyurethane (SPU). Semi-interpenetrating polymer networks (IPNs) composed of crosslinked MPC polymer and SPU were prepared. The mechanical properties of the IPN membrane were significantly improved compared with those of the MPC polymer membrane. Three-dimensional polymer networks of the MPC polymer in the IPNs were observed after solvent extraction of SPU. An X-ray photoelectron spectrum analysis revealed that the MPC units were exposed on the IPN surface. When the IPN was alternately soaked in water and ethanol, the swelling ratio was found to be completely reversible and no disintegration of the network structure was observed. The permeation coefficient of 1,4-di(2-hydroxyethoxy)benzene through the IPN membrane was 1.11 × 10−7 cm−2s−1. The amount of adsorbed protein and the number of adherent platelets on the IPN membrane were effectively reduced compared with those on SPU. We concluded that IPNs composed of the MPC polymer and SPU are a new bulk biomaterial, which possesses both blood compatibility and good mechanical properties. © 2000 John Wiley & Sons, Inc. J Biomed Mater Res, 52, 701–708, 2000.

Themolar heat capacity of the thermotropic cubic mesogen 1,2-bis(4- n -alkoxybenzoyl)hydrazine, BABH(8) for short, with a purity of 99.43 mol% has been precisely measured with an adiabatic calorimeter at temperatures between 14 and 480 K. The enthalpy and entropy gained at each phase transition across the phase sequence of \[crystal(2) crystal(1) cubic mesophase SmC isotropic liquid] have been determined. The existence of a solid-to-solid phase transition with a fairly large entropy change seems to be necessary for the alkyl moieties attached to both sides of the molecule to play the role of 'solvent' in the cubic mesophase. On the basis of curvature elasticity considerations, the small energy difference between the cubic and SmC phases is favourably accounted for in terms of the jointed-rod micelles model. The reason for the immiscibility of BABH(8) with the cubic D mesogen, 4- n -hexadecyloxy-3- nitrobiphenyl-4-carboxylic acid is discussed in terms of the large difference in their molecular size and of ...

Tumor necrosis factor (TNF)-α exerts its biological function via TNF type 1 and type 2 receptors (TNFR1 and TNFR2). We have previously reported that bone resorption induced by lipopolysaccharide (LPS) in TNFR2-deficient mice is accelerated compared to that in wild-type (WT) mice. Although these results suggested that TNFR2 might have a protective role in bone resorption, we could not exclude the possibility that TNFR2 has no role in bone resorption. To clarify the role of TNFR2, we developed a TNF-α-induced bone resorption model using cholesterol-bearing pullulan nanogel as a TNF-α carrier to minimize the influence of inflammatory cytokines other than TNF-α. Injections of human TNF-α (hTNF), an agonist of mouse TNFR1, stimulated bone resorption lacunae on the calvariae in WT mice, but mouse TNF-α (mTNF), an agonist of both mouse TNFR1 and TNFR2, could not. To eliminate the possibility that the TNFR1 agonistic effects of hTNF were stronger than those of mTNF, we used the same model in TNFR2-deficient mice. Injection of mTNF resulted in clear bone resorption lacunae to the same extent observed after using hTNF in the TNFR2-deficient mice. Histomorphometric analysis of osteoclast number supported the observed changes in bone resorption lacunae. These data suggest that TNFR2 has a protective role in TNF-α-induced bone resorption.

Hydration properties of adenine nucleotides and orthophosphate (Pi) in aqueous solutions adjusted to pH=8 with NaOH were studied by high-resolution microwave dielectric relaxation (DR) spectroscopy at 20 °C. The dielectric spectra were analyzed using a mixture theory combined with a least-squares Debye decomposition method. Solutions of Pi and adenine nucleotides showed qualitatively similar dielectric properties described by two Debye components. One component was characterized by a relaxation frequency (f(c)=18.8-19.7 GHz) significantly higher than that of bulk water (17 GHz) and the other by a much lower f(c) (6.4-7.6 GHz), which are referred to here as hyper-mobile water and constrained water, respectively. By contrast, a hydration shell of only the latter type was found for adenosine (f(c)~6.7 GHz). The present results indicate that phosphoryl groups are mostly responsible for affecting the structure of the water surrounding the adenine nucleotides by forming one constrained water layer and an additional three or four layers of hyper-mobile water.

The Catalytic properties of ISI-1, 3, 4, 5 and 6, synthesized in mixtures of water and organic solvent, have been studied. Although having acidic properties similar to a sample of ZSM-5 zeolite comprising small and aggregated crystallites, the five zeolites showed low selectivities for C 5 + product and fraction of aromatics in the C 5 + product in syngas-to-gasoline and methanol-to-gasoline reactions. Under methanol-to-gasoline reaction conditions, ISI-3, 4, 5 and 6 showed a high production of p -xylene in the xylene product. From the results of physicochemical measurements, the unique Catalytic properties of each ISI zeolite can be attributed to its crystal structure, pore size and large crystallite size.

Functional nanogels have been designed by the self-assembly of various associating polymers. In particular, cholesterol-bearing polysaccharides form physically crosslinked nanogels by self-assembly in water. The nanogels trap proteins mainly by hydrophobic interaction and show chaperon-like activity. They are useful as polymeric nanocarriers especially in protein delivery. Macrogels with well-defined nanostructures were obtained by self-assembly and chemical crosslinking of these nanogels as building blocks.

The Catalytic properties of ISI-1, 3, 4, 5 and 6, synthesized in mixtures of water and organic solvent, have been studied. Although having acidic properties similar to a sample of ZSM-5 zeolite comprising small and aggregated crystallites, the five zeolites showed low selectivities for C 5 + product and fraction of aromatics in the C 5 + product in syngas-to-gasoline and methanol-to-gasoline reactions. Under methanol-to-gasoline reaction conditions, ISI-3, 4, 5 and 6 showed a high production of p -xylene in the xylene product. From the results of physicochemical measurements, the unique Catalytic properties of each ISI zeolite can be attributed to its crystal structure, pore size and large crystallite size.

Novel hybrid hyaluronan (HA) hydrogel encapsulating nanogels was designed for sustained delivery of protein. HA modified with 2-aminoethyl methacrylate was cross-linked via Michael addition in the presence of cholesteryl group-bearing pullulan (CHP) nanogels. The nanogels were physically entrapped and well dispersed in a three-dimensional network of chemically cross-linked HA (HA gel). Therapeutic peptides and proteins, such as glucagon-like peptide-1, insulin and erythropoietin, were spontaneously trapped in the CHP nanogels in the HA gel just by immersing hybrid hydrogels into the drug solutions. CHP/protein complex nanogels were released from the hybrid hydrogels in a sustained manner both in vitro and in vivo. The release was controlled by the cross-linking density and the degradability of the HA gel, modulated by the initial gelation condition. The synergy between the CHP nanogel as a drug reservoir and the HA gel as a nanogel-releasing matrix of the hybrid hydrogel system simultaneously achieved both simple drug loading and controlled release with no denaturation of the protein drugs. This is a new method of fabricating biodegradable controlled release matrix with molecular chaperone-like activity for therapeutic proteins.

Highly branched cyclic dextrin derivatives (CH-CDex) that are partly substituted with cholesterol groups have been synthesized. The CH-CDex forms monodisperse and stable nanogels with a hydrodynamic radii of approximately 10 nm by the self-assembly of 4-6 CH-CDex macromolecules in water. The CH-CDex nanogels spontaneously trap 10-16 molecules of fluorescein isothiocyanate-labeled insulin (FITC-Ins). The complex shows high colloidal stability: no dissociation of trapped insulin is observed after at least 1 month in phosphate buffer (0.1 M, pH 8.0). In the presence of bovine serum albumin (BSA, 50 mg . mL(-1)), which is a model blood system, the FITC-Ins trapped in the nanogels is continuously released ( approximately 20% at 12 h) without burst release. The high-density nanogel structure derived from the highly branched CDex significantly affects the stability of the nanogel-protein complex.