Yung Chang

Molecular Simulation Studies of Protein Interactions with Zwitterionic Phosphorylcholine Self-Assembled Monolayers in the Presence of Water

Molecular simulations were performed to study the interactions between a protein (lysozyme, LYZ) and phosphorylcholine-terminated self-assembled monolayers (PC-SAMs) in the presence of explicit water molecules and ions. The results show that the water molecules above the PC-SAM surface create a strong repulsive force on the protein as it approaches the surface. The structural and dynamic properties of the water molecules above the PC-SAM surface were analyzed to provide information regarding the role of hydration in surface resistance to protein adsorption. It can be seen from residence time dynamics that the water molecules immediately above the PC-SAM surface are significantly slowed down as compared to bulk water, suggesting that the PC-SAM surface generates a tightly bound, structured water layer around its head groups. Moreover, the orientational distribution and reorientational dynamics of the interfacial water molecules near the PC-SAM surface were found to have the ionic solvation nature of the PC head groups. These properties were also compared to those obtained previously for an oligo(ethylene glycol) (OEG) SAM system and bulk water.

Zwitterionic Sulfobetaine-Grafted Poly(vinylidene fluoride) Membrane with Highly Effective Blood Compatibility via Atmospheric Plasma-Induced Surface Copolymerization

Development of nonfouling membranes to prevent nonspecific protein adsorption and platelet adhesion is critical for many biomedical applications. It is always a challenge to control the surface graft copolymerization of a highly polar monomer from the highly hydrophobic surface of a fluoropolymer membrane. In this work, the blood compatibility of poly(vinylidene fluoride) (PVDF) membranes with surface-grafted electrically neutral zwitterionic poly(sulfobetaine methacrylate) (PSBMA), from atmospheric plasma-induced surface copolymerization, was studied. The effect of surface composition and graft morphology, electrical neutrality, hydrophilicity and hydration capability on blood compatibility of the membranes were determined. Blood compatibility of the zwitterionic PVDF membranes was systematically evaluated by plasma protein adsorption, platelet adhesion, plasma-clotting time, and blood cell hemolysis. It was found that the nonfouling nature and hydration capability of grafted PSBMA polymers can be effectively controlled by regulating the grafting coverage and charge balance of the PSBMA layer on the PVDF membrane surface. Even a slight charge bias in the grafted zwitterionic PSBMA layer can induce electrostatic interactions between proteins and the membrane surfaces, leading to surface protein adsorption, platelet activation, plasma clotting and blood cell hemolysis. Thus, the optimized PSBMA surface graft layer in overall charge neutrality has a high hydration capability and the best antifouling, anticoagulant, and antihemolytic activities when comes into contact with human blood.

Physical cues of biomaterials guide stem cell differentiation fate.

Akon Higuchi,*,†,‡,§ Qing-Dong Ling, Yung Chang, Shih-Tien Hsu, and Akihiro Umezawa‡ †Department of Chemical and Materials Engineering, National Central University, Jhongli, Taoyuan 32001, Taiwan ‡Department of Reproductive Biology, National Research Institute for Child Health and Development, 2-10-1 Okura, Setagaya-ku, Tokyo 157-8535, Japan Cathay Medical Research Institute, Cathay General Hospital, No. 32, Ln 160, Jian-Cheng Road, Hsi-Chi City, Taipei 221, Taiwan Institute of Systems Biology and Bioinformatics, National Central University, No. 300 Jhongda Rd., Jhongli, Taoyuan 32001, Taiwan Department of Chemical Engineering, R&D Center for Membrane Technology, Chung Yuan Christian University, 200 Chung-Bei Rd., Jhongli, Taoyuan 320, Taiwan Taiwan Landseed Hospital, 77 Kuangtai Road, Pingjen City, Tao-Yuan County 32405, Taiwan

A highly stable nonbiofouling surface with well-packed grafted zwitterionic polysulfobetaine for plasma protein repulsion.

An ideal nonbiofouling surface for biomedical applications requires both high-efficient antifouling characteristics in relation to biological components and long-term material stability from biological systems. In this study we demonstrate the performance and stability of an antifouling surface with grafted zwitterionic sulfobetaine methacrylate (SBMA). The SBMA was grafted from a bromide-covered gold surface via surface-initiated atom transfer radical polymerization to form well-packed polymer brushes. Plasma protein adsorption on poly(sulfobetaine methacrylate) (polySBMA) grafted surfaces was measured with a surface plasmon resonance sensor. It is revealed that an excellent stable nonbiofouling surface with grafted polySBMA can be performed with a cycling test of the adsorption of three model proteins in a wide range of various salt types, buffer compositions, solution pH levels, and temperatures. This work also demonstrates the adsorption of plasma proteins and the adhesion of platelets from human blood plasma on the polySBMA grafted surface. It was found that the polySBMA grafted surface effectively reduces the plasma protein adsorption from platelet-poor plasma solution to a level superior to that of adsorption on a surface terminated with tetra(ethylene glycol). The adhesion and activation of platelets from platelet-rich plasma solution were not observed on the polySBMA grafted surface. This work further concludes that a surface with good hemocompatibility can be achieved by the well-packed surface-grafted polySBMA brushes.

Investigation of the Hydration of Nonfouling Material Poly(sulfobetaine methacrylate) by Low-Field Nuclear Magnetic Resonance

The strong surface hydration layer of nonfouling materials plays a key role in their resistance to nonspecific protein adsorption. Poly(sulfobetaine methacrylate) (polySBMA) is an effective material that can resist nonspecific protein adsorption and cell adhesion. About eight water molecules are tightly bound with one sulfobetaine (SB) unit, and additional water molecules over 8:1 ratio mainly swell the polySBMA matrix, which is obtained through the measurement of T(2) relaxation time by low-field nuclear magnetic resonance (LF-NMR). This result was also supported by the endothermic behavior of water/polySBMA mixtures measured by differential scanning calorimetry (DSC). Furthermore, by comparing both results of polySBMA and poly(ethylene glycol) (PEG), it is found that (1) the hydrated water molecules on the SB unit are more tightly bound than on the ethylene glycol (EG) unit before saturation, and (2) the additional water molecules after forming the hydration layer in polySBMA solutions show higher freedom than those in PEG. These results might illustrate the reason for higher resistance of zwitterionic materials to nonspecific protein adsorptions compared to that of PEGs.

Hemocompatible Mixed-Charge Copolymer Brushes of Pseudozwitterionic Surfaces Resistant to Nonspecific Plasma Protein Fouling

In this work, the hemocompatibility of a sulfobetaine-like copolymer brush resulting from a mixed-charge copolymerization of the positively charged 11-mercapto-N,N,N-trimethylammonium chloride (TMA) and negatively charged 11-mercaptoundecylsulfonic acid (SA) was studied. Mixed charge distribution in the prepared poly(TMA-co-SA) copolymer brushes was controlled by the regulation of the reaction rate of the surface-initiated atom transfer radical polymerization (ATRP). The adsorption behavior of plasma proteins on a surface grafted with poly(TMA-co-SA) was measured by a surface plasmon resonance (SPR) sensor. The effects of varying temperature, solution pH, and ionic strength on the antifouling characteristics of the mixed-charge copolymer brushes were systematically evaluated, and the protein-fouling resistance was discussed in detail, especially with respect to the effect of ionic strength on the intra- and intermolecular interactions of the poly(TMA-co-SA) with proteins. The adhesion and activation of blood cells on the poly(TMA-co-SA)-grafted surface in contact with human whole blood was also demonstrated. The results suggest that mixed-charge copolymer brushes of poly(TMA-co-SA), which, like zwitterionic homopolymer brushes, have overall charge neutrality, can be used in similar applications for protein-fouling resistance and have excellent hemocompatibility with human whole blood at physiologic temperatures.

Tunable Bioadhesive Copolymer Hydrogels of Thermoresponsive Poly(N-isopropyl acrylamide) Containing Zwitterionic Polysulfobetaine

This work describes a novel tunable bioadhesive hydrogel of thermoresponsive N-isopropylacrylamide (NIPAAm) containing zwitterionic sulfobetaine methacrylate (SBMA). This novel hydrogel highly regulates general bioadhesive foulants through the adsorption of plasma proteins, the adhesion of human platelets and cells, and the attachment of bacteria. In this investigation, nonionic hydrogels of polyNIPAAm, zwitterionic hydrogels of polySBMA, and three copolymeric hydrogels of NIPAAm and SBMA (poly(NIPAAm-co-SBMA)) were prepared. The copolymeric hydrogels exhibited controllable temperature-dependent swelling behaviors and showed stimuli-responsive phase characteristics in the presence of salts. The interactions of these hydrogels with biomolecules and microorganisms were demonstrated by protein adsorption, cell adhesion, and bacterial attachment, which allowed us to evaluate their bioadhesive properties. An enzyme-linked immunosorbent assay (ELISA) with monoclonal antibodies was used to measure different plasma protein adsorptions on the prepared hydrogel surfaces. At a physiological temperature, the high content of the nonionic polyNIPAAm in poly(NIPAAm-co-SBMA) hydrogel exhibits a high protein adsorption due to the interfacial exposure of polyNIPAAm-rich hydrophobic domains. A relatively high content of polySBMA in poly(NIPAAm-co-SBMA) hydrogel exhibits reduced amounts of protein adsorption due to the interfacial hydration of polySBMA-rich hydrophilic segments. The attachment of platelets and the spreading of cells were only observed on polyNIPAAm-rich hydrogel surfaces. Interestingly, the incorporation of zwitterionic SBMA units into the polyNIPAAm gels was found to accelerate the hydration of the cell-cultured surfaces and resulted in more rapid cell detachment. Such copolymer gel surface was shown to be potentially useful for triggered cell detachment. In addition, the interactions of hydrogels with bacteria were also evaluated. The polySBMA-rich hydrogels exhibited evident antimicrobial properties when they were incubated with Gram-positive bacteria ( S. epidermidis ) and Gram-negative bacteria ( E. coli ). This work shows that the bioadhesive properties of poly(NIPAAm-co-SBMA) hydrogels can be effectively controlled via regulated nonionic and zwitterionic molar mass ratios. The tunable-bioadhesive behavior of temperature-sensitive poly(NIPAAm-co-SBMA) makes this biocompatible hydrogel appropriate for biomedical applications.

Hemocompatibility of poly(vinylidene fluoride) membrane grafted with network-like and brush-like antifouling layer controlled via plasma-induced surface PEGylation.

In this work, the hemocompatibility of PEGylated poly(vinylidene fluoride) (PVDF) microporous membranes with varying grafting coverage and structures via plasma-induced surface PEGylation was studied. Network-like and brush-like PEGylated layers on PVDF membrane surfaces were achieved by low-pressure and atmospheric plasma treatment. The chemical composition, physical morphology, grafting structure, surface hydrophilicity, and hydration capability of prepared membranes were determined to illustrate the correlations between grafting qualities and hemocompatibility of PEGylated PVDF membranes in contact with human blood. Plasma protein adsorption onto different PEGylated PVDF membranes from single-protein solutions and the complex medium of 100% human plasma were measured by enzyme-linked immunosorbent assay (ELISA) with monoclonal antibodies. Hemocompatibility of the PEGylated membranes was evaluated by the antifouling property of platelet adhesion observed by scanning electron microscopy (SEM) and the anticoagulant activity of the blood coagulant determined by testing plasma-clotting time. The control of grafting structures of PEGylated layers highly regulates the PVDF membrane to resist the adsorption of plasma proteins, the adhesion of platelets, and the coagulation of human plasma. It was found that PVDF membranes grafted with brush-like PEGylated layers presented higher hydration capability with binding water molecules than with network-like PEGylated layers to improve the hemocompatible character of plasma protein and blood platelet resistance in human blood. This work suggests that the hemocompatible nature of grafted PEGylated polymers by controlling grafting structures gives them great potential in the molecular design of antithrombogenic membranes for use in human blood.

Dual-Thermoresponsive Phase Behavior of Blood Compatible Zwitterionic Copolymers Containing Nonionic Poly(N-isopropyl acrylamide)

Thermoresponsive statistical copolymers of zwitterionic sulfobetaine methacrylate (SBMA) and nonionic N-isopropylacrylamide (NIPAAm) were prepared with an average molecular weight of about 6.0 kDa via homogeneous free radical copolymerization. The aqueous solution properties of poly(SBMA-co-NIPAAm) were measured using a UV--visible spectrophotometer. The copolymers exhibited controllable lower and upper critical solution temperatures in aqueous solution and showed stimuli-responsive phase transition in the presence of salts. Regulated zwitterionic and nonionic molar mass ratios led to poly(SBMA-co-NIPAAm) copolymers having double-critical solution temperatures, where the water-insoluble polymer microdomains are generated by the zwitterionic copolymer region of polySBMA or nonionic copolymer region of polyNIPAAm depending on temperature. A high content of the nonionic polyNIPAAm in poly(SBMA-co-NIPAAm) exhibits nonionic aggregation at high temperatures due to the desolvation of polyNIPAAm, whereas relatively low content of polyNIPAAm in poly(SBMA-co-NIPAAm) exhibits zwitterionic aggregation at low temperatures due to the desolvation of polySBMA. Plasma protein adsorption on the surface coated with poly(SBMA-co-NIPAAm) was measured with a surface plasmon resonance (SPR) sensor. The copolymers containing polySBMA above 29 mol % showed extremely low protein adsorption and high anticoagulant activity in human blood plasma. The tunable and switchable thermoresponsive phase behavior of poly(SBMA-co-NIPAAm), as well as its high plasma protein adsorption resistance and anticoagulant activity, suggests a potential for blood-contacting applications.

Biomaterials for the Feeder-Free Culture of Human Embryonic Stem Cells and Induced Pluripotent Stem Cells

Biomaterials for the Feeder-Free Culture of Human Embryonic Stem Cells and Induced Pluripotent Stem Cells Akon Higuchi,* Qing-Dong Ling, Yi-An Ko, Yung Chang, and Akihiro Umezawa Department of Chemical and Materials Engineering, National Central University, Jhongli, Taoyuan, 32001 Taiwan Department of Reproduction, National Research Institute for Child Health and Development, 2-10-1 Okura, Setagaya-ku, Tokyo 157-8535, Japan Cathay Medical Research Institute, Cathay General Hospital, No. 32, Ln 160, Jian-Cheng Road, Xi-Zhi Dist., New Taipei City, Taiwan Institute of Systems Biology and Bioinformatics, National Central University, No. 300, Jhongda RD., Jhongli, Taoyuan, 32001 Taiwan Department of Chemical Engineering, R&DCenter forMembrane Technology, Chung Yuan Christian University, 200 Chung-Bei Rd., Jhongli, Taoyuan 320, Taiwan