Abstract This report describes a new surface-treatment technique for cell micropatterning. Cell attachment was selectively controlled on the glass surface using a photochemical reaction. This ...strategy is based on combining 2-methacryloyloxyethyl phosphorylcholine (MPC) polymer, which is known to reduce non-specific adsorption, and a photolabile linker (PL) for selective cell patterning. The MPC polymer was coated directly on the glass surface using a straightforward surface modification method, and was removed by ultraviolet (UV) light illumination. All the surface modification steps were evaluated using static water contact angle measurements, X-ray photoelectron spectroscopy (XPS), atomic force microscopy (AFM), measurements of non-specific protein adsorption, and the cell attachment test. After selective cleavage of the MPC polymer through the photomask, cells attached only to the UV-illuminated region where the MPC polymer was removed, which made the hydrophilic surface relatively hydrophobic. Furthermore, the size of the MC-3T3 E1 cell patterns could be controlled by single cell level. Stability of the cell micropatterns was demonstrated by culturing MC-3T3 E1 cell patterns for 5 weeks on glass slide. The micropatterns were stable during culturing; cell viability also was verified. This method can be a powerful tool for cell patterning research.
In order to provide a protein adsorption resistant surface even when the surface was in contact with a protein solution under completely dry conditions, a new phospholipid copolymer, poly ...(2-methacryloyloxyethyl phosphorylcholine (MPC)-co-2-vinylnaphthalene (vN)) (PMvN), was synthesized. Poly(ethylene terephthalate) (PET) could be readily coated with PMvN by a solvent evaporation method. Dynamic contact angle measurements with water revealed that the surface was wetted very rapidly and had strong hydrophilic characteristics; moreover, molecular mobility at the surface was extremely low. When the surface came in contact with a plasma protein solution containing bovine serum albumin (BSA), the amounts of the plasma protein adsorbed on the dry surface coated with PMvN and that adsorbed on a dry surface coated with poly(MPC-co-n-butyl methacrylate) (PMB) were compared. Substantially lower protein adsorption was observed with PMvN coating. This is due to the rapid hydration behavior of PMvN. We concluded that PMvN can be used as a functional coating material for medical devices without any wetting pretreatment.
Clarifying material–cell interactions after eliminating various effects including cell–cell interactions is a very important issue in tissue and cell engineering. We investigated several cellular ...behaviors dynamically after growing cells individually one by one to eliminate cell–cell contact by direct observation using the quartz crystal microbalance with dissipation (QCM-D) and microscopy. The initial cellular behaviors of several cells (adsorption, attachment and spreading of L929 mouse fibroblasts) on the gold electrode of the QCM-D were classified into three regions according to the slope of the Df plots: I, cell adsorption and desorption; II, attachment and spreading; and III, secretion of microexudates. When the number of cells increased which leads to increase of cell–cell distance, region III disappeared and the slope of the Df plots in region II became steeper. The behavior related to the variation in the number of attached cells was indicated by the adhesion strength between the cells and the gold electrode. Furthermore, the slope of the Df plots was not shown on the (poly(2-methacryloyloxyethyl phosphorylcholine)–co–2-(methacryloyloxy)ethylthiol)-modified gold electrode and almost all of the cells were not attached to the polymer surface. The initial cell attachment behavior, especially the strength of cell adhesion to the material surface, was evaluated quantitatively by determining the slope of the Df plots using the QCM-D system.
There is growing awareness that circadian clocks are closely related to the intracellular redox state across a range of species. As the redox state is determined by the exchange of the redox species, ...electrochemically controlled extracellular electron transfer (EC‐EET), a process in which intracellular electrons are exchanged with extracellular electrodes, is a promising approach for the external regulation of circadian clocks. Herein, we discuss whether the circadian clock can be regulated by EC‐EET using the cyanobacterium Synechococcus elongatus PCC7942 as a model system. In vivo monitoring of chlorophyll fluorescence revealed that the redox state of the plastoquionone pool could be controlled with EC‐EET by simply changing the electrode potential. As a result, the endogenous circadian clock of S. elongatus cells was successfully entrained through periodically modulated EC‐EET by emulating the natural light/dark cycle, even under constant illumination conditions. This is the first example of regulating the biological clock by electrochemistry.
Let's do the time warp again: Electrochemically controlled extracellular electron transfer can regulate the circadian clock of the cyanobacterium Synechococcus elongatus under constant light intensity through periodic modification of the intracellular redox state. This method could potentially allow for direct or indirect regulation of the biological clock across a range of species.
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► Redox phospholipid polymer microparticles as doubly functional polymer was prepared successfully. ► The polymer functions both biocompatible platform and electron transfer mediator. ...► Enzyme activity was maintained at least one month by the immobilization at 25°C. ► Electrons generated by the enzymatic reaction could transfer to the gold substrate through the redox polymer layer effectively. ► The microparticles are useful platform for understanding interfacial phenomena and, furthermore, making oxidase-based biodevices.
We prepared redox phospholipid polymer microparticles for immobilizing an enzyme in order to maintain activity for a long time and obtain highly effective electron transfer to a gold substrate as an electrode. To achieve these double functions, an amphiphilic redox phospholipid polymer, poly(2-methacryloyloxyethyl phosphorylcholine-co-n-butyl methacrylate-co-p-nitrophenyloxycarbonyl oligo(ethylene glycol) methacrylate (MEONP)-co-vinylferrocene (VFc)) (PMBNF) was synthesized. The polystyrene (PS) microparticles were modified by employing a simple solution dip-coating technique to form the PMBNF layer on the surface. As one of the model enzyme oxidases, a glucose oxidase (GOx) was immobilized on the PMBNF/PS microparticles by the reaction between the MEONP units in the PMBNF layer and the amino group in the GOx. The activity of immobilized GOx is maintained well; for example, activity of more than 80% of the initial activity was observed even after storage at both 4°C and 25°C (ionic strength: 0.10mol/L, phosphate buffer solution, pH 7.0) for at least one month. The GOx/PMBNF/PS microparticles were arrayed on a gold substrate in a monolayer, and then, crosslinked to each other with a polymeric diamine compound. The PMBNF/PS microparticles demonstrated an efficient electron transfer from immobilized GOx to the gold surface. From these results, we concluded that the PMBNF layer on the PS microparticles possessed double functions such as stable enzyme immobilization ability and efficient electron transfer ability.
To obtain a stable and highly sensitive bioimaging fluorescence probe, polymer nanoparticles with embedded quantum dots were covered with an artificial cell membrane. These nanoparticles were ...designed by assembling phospholipid polar groups as a platform, and oligopeptide was immobilized as a bioaffinity moiety on the surface of the nanoparticles. The polymer nanoparticles showed resistance to cellular uptake from HeLa cells owing to the nature of the phosphorylcholine groups. When arginine octapeptide was immobilized at the surface of the nanoparticles, they were able to penetrate the membrane of HeLa cells effectively. Cytotoxicty of the nanoparticles was not observed even after immobilization of oligopeptide. Thus, we obtained stable fluorescent polymer nanoparticles covered with an artificial cell membrane, which are useful as an excellent bioimaging probe and as a novel evaluation tool for oligopeptide functions in the target cells.
Abstract We previously reported that modification of GALA peptide on the surface of liposomes enhanced fusion with endosomal membrane, and cytoplasmic release of encapsulated macromolecules. We ...report herein that an additional coating of GALA-modified liposomes with 2-methacryloyloxyethyl phosphorylcholine (MPC) polymer resulted in a two order of magnitude enhancement in the transfection activity of encapsulating plasmid DNA (pDNA). Quantification of the delivered gene copies in whole cells and isolated nuclei revealed that the increase of transfection activity can be attributed to improved efficiencies in cellular uptake and post-nuclear delivery processes. Imaging studies revealed that the intracellular dissociation of pDNA from the lipid envelope is enhanced by GALA modification and further coating with MPC polymer in a stepwise manner. The MPC polymer-coating decreased the ζ-potential of GALA-modified liposomes, suggesting that it assisted in the functional display of negatively charged GALA on the cationic liposomes by providing shielding from mutual electrostatic interactions. Collectively, these data indicate that MPC polymer-coating induced the fusogenic activity of the GALA-modified envelope with endosomes, leading to a more effective cytoplasmic release pDNA. The extensive fusion of the lipid envelope may also reduce electrostatic interactions between mRNA and cationic lipid components, thereby resulting in an enhancement in the translation process.
