Adenoviral Gene Delivery from Multilayered Polyelectrolyte Architectures

The alternate layer‐by‐layer (LBL) deposition of polycations and polyanions for the build up of multilayered polyelectrolyte films is an original approach that allows the preparation of tunable, biologically active surfaces. The resulting supramolecular nanoarchitectures can be functionalized with drugs, peptides, and proteins, or DNA molecules that are able to transfect cells in vitro. We monitor, for the first time, the embedding of a bioactive adenoviral (Ad) vector in multilayered polyelectrolyte films. Ad efficiently adsorbs on poly(L ‐lysine)–poly(L ‐glutamic acid) (PLL–PGA), PLL–HA (HA: hyaluronan), poly(allylamin hydrochloride)–poly(sodium‐4‐styrenesulfonate) (PAH–PSS), and CHI–HA (CHI: chitosan) films; it preserves its transduction capacity (which can reach 95 %) for a large number of cell types, and also allows vector uptake into receptor‐deficient cells, thus abrogating the restricted tropism of Ad.     

Build‐up of Polypeptide Multilayer Coatings with Anti‐Inflammatory Properties Based on the Embedding of Piroxicam–Cyclodextrin Complexes

We describe the build‐up of biomaterial coatings based on polypeptide multilayers possessing anti‐inflammatory properties. Poly(L ‐lysine) (PLL) and poly(L ‐glutamic acid) (PGA) are used as polypeptides, and piroxicam (Px) is used as the anti‐inflammatory agent. In order to embed high enough amounts of Px, the drug is incorporated in the films in the form of complexes with a charged 6^A‐carboxymethylthio‐β‐cyclodextrin (cCD). It is shown that this cyclodextrin can solubilize higher amounts of Px than the cyclodextrins used commercially. The anti‐inflammatory properties are evaluated by determining the inhibition of TNFα production by human monocytic THP‐1 cells stimulated with lipopolysaccharide (LPS) bacterial endotoxin. Using Fourier‐transform (FT) Raman spectroscopy, we show that Px is mainly in the neutral form in cCD–Px complexes in solution, and that it remains biologically active under this form, whereas up to now only the zwitterionic form was reported to possess anti‐inflammatory properties. When incorporated in PLL/PGA multilayers, Px in the cCD–Px complexes changes from the neutral to the zwitterionic form. It is shown that these films present anti‐inflammatory properties, which can be delayed, and whose duration can be tuned by changing the film architecture.     

Targeting of Cancer Cells Using Quantum Dot–Polypeptide Hybrid Assemblies That Function as Molecular Imaging Agents and Carrier Systems

A highly tunable quantum dot (QD)–polypeptide hybrid assembly system with potential uses for both molecular imaging and delivery of biomolecular cargo to cancer cells is reported. The tunability of the assembly system, its application for imaging cancer cells, and its ability to carry a biomolecule are demonstrated. The assemblies are formed through the self‐assembly of carboxyl‐functionalized QDs and poly(diethylene glycol‐L ‐lysine)‐poly(L ‐lysine) (PEGLL‐PLL) diblock copolypeptide molecules, and they are modified with peptide ligands containing a cyclic arginine‐glycine‐aspartate [c(RGD)] motif that has affinity for αvβ3 and αvβ5 integrins overexpressed on the tumor vasculature. To illustrate the tunability of the QD‐polypeptide assembly system, it is shown that binding to U87MG glioblastoma cells can be modulated and optimized by changing either the conditions under which the assemblies are formed or the relative lengths of the PEGLL and PLL blocks in the PEGLL‐PLL molecules. The optimized c(RGD)‐modified assemblies bind integrin receptors on U87MG cells and are endocytosed, as demonstrated by flow cytometry and live‐cell imaging. Binding specificity is confirmed by competition with an excess of free c(RGD) peptide. Finally, it is shown that the QD–polypeptide assemblies can be loaded with fluorescently labeled ovalbumin, as a proof‐of‐concept for their potential use in biomolecule delivery.     

VEGF‐Functionalized Polyelectrolyte Multilayers as Proangiogenic Prosthetic Coatings

Stimulation of transprosthetic vascularization represents an interesting strategy in implantology to allow rapid tissue integration and finally to avoid prosthetic rejection. To achieve this goal, we modified the surface of porous titanium implants with polyelectrolyte multilayer (PEM) films functionalized with vascular endothelial growth factor (VEGF). Among the two PEM systems investigated, poly(L‐lysine)/poly(L‐glutamic acid) (PLL/PGA) and poly(allylamine hydrochloride)/poly(sodium 4‐styrenesulfonate) (PAH/PSS), the (PAH/PSS)₄ architecture was selected to functionalize porous titanium, both for its high efficiency to adsorb VEGF and for its biocompatibility toward endothelial cells. In an original way, we unambiguously demonstrated that VEGF adsorbed on (PAH/PSS)₄ maintains its bioactivity in vitro and stimulates endothelial cells proliferation. This effect was correlated with specific activation of intracellular signaling pathways induced by successive phosphorylation of the endothelial VEGF receptor VEGFR2 and mitogen‐activated protein kinases (MAPK) ERK1/2. By clearly demonstrating the proangiogenic activity of the VEGF‐PEM coating in vitro, the present study constitutes a first step toward in vivo application.     

Biofunctional Polyelectrolyte Multilayers and Microcapsules: Control of Non‐Specific and Bio‐Specific Protein Adsorption

Layers of the polyelectrolytes poly(allylamine hydrochloride) (PAH, polycationic) and poly(styrene sulfonate) (PSS, polyanionic) are consecutively adsorbed on flat silicon oxide surfaces, forming stable, ultrathin multilayer films. Subsequently, a final monolayer of the polycationic copolymer poly(L ‐lysine)‐graft‐poly(ethylene glycol) (PLL‐g‐PEG) is adsorbed onto the PSS‐terminated multilayer in order to impart protein resistance to the surface. The growth of each of the polyelectrolyte layers and the protein resistance of the resulting [PAH/PPS]_n(PLL‐g‐PEG) multilayer (n = 1–4) are followed quantitatively ex situ using X‐ray photoelectron spectroscopy and in situ using real‐time optical‐waveguide lightmode spectroscopy. In a second approach, the same type of [PAH/PSS]_n(PLL‐g‐PEG) multilayer coatings are successfully formed on the surface of colloidal particles in order to produce surface‐functionalized, hollow microcapsules after dissolution of the core materials (melamine formaldehyde (MF) and poly(lactic acid) (PLA; colloid diameters: 1.2–20 μm). Microelectrophoresis and confocal laser scanning microscopy are used to study multilayer formation on the colloids and protein resistance of the final capsule. The quality of the PLL‐g‐PEG layer on the microcapsules depends on both the type of core material and the dissolution protocols used. The greatest protein resistance is achieved using PLA cores and coating the polyelectrolyte microcapsules with PLL‐g‐PEG after dissolution of the cores. Protein adsorption from full serum on [PAH/PPS]_n(PLL‐g‐PEG) multilayers (on both flat substrates and microcapsules) decreases by three orders of magnitude in comparison to the standard [PAH/PPS]_n layer. Finally, biofunctional capsules of the type [PAH/PPS]_n(PLL‐g‐PEG/PEG‐biotin) (top copolymer layer with a fraction of the PEG chains end‐functionalized with biotin) are produced which allow for specific recognition and immobilization of controlled amounts of streptavidin at the surface of the capsules. Biofunctional multilayer films and capsules are believed to have a potential for future applications as novel platforms for biotechnological applications such as biosensors and carriers for targeted drug delivery.     

Polyelectrolyte multilayer films of controlled stiffness modulate myoblast cells differentiation

Beside chemical properties and topographical features, mechanical properties of gels have been recently demonstrated to play an important role in various cellular processes, including cell attachment, proliferation, and differentiation. In this work, we used multilayer films made of poly(L-lysine)/Hyaluronan (PLL/HA) of controlled stiffness to investigate the effects of mechanical properties of thin films on skeletal muscle cells (C2C12 cells) differentiation. Prior to differentiation, cells need to adhere and proliferate in growth medium. Stiff films (E(0) > 320 kPa) promoted formation of focal adhesions and organization of the cytoskeleton as well as an enhanced proliferation, whereas soft films were not favorable for cell anchoring, spreading or proliferation. Then C2C12 cells were switched to a low serum containing medium to induce cell differentiation, which was also greatly dependent on film stiffness. Although myogenin and troponin T expressions were only moderately affected by film stiffness, the morphology of the myotubes exhibited striking stiffness-dependent differences. Soft films allowed differentiation only for few days and the myotubes were very short and thick. Cell clumping followed by aggregates detachment could be observed after ~2 to 4 days. On stiffer films, significantly more elongated and thinner myotubes were observed for up to ~ 2 weeks. Myotube striation was also observed but only for the stiffer films. These results demonstrate that film stiffness modulates deeply adhesion, proliferation and differentiation, each of these processes having its own stiffness requirement.     

Thermally Stable and Solvent Resistant Mesoporous Honeycomb Films from a Crosslinkable Polymer

The preparation of mesoporous honeycomb films, also known as breath figure arrays (BFAs), from poly(styrene‐co‐maleic anhydride) is reported. Films containing regular arrays of micrometer‐sized air‐holes were prepared by evaporation of a chloroform solution of a mixture of the above polymer including 10 % of an amphiphilic polyion complex under high humidity that leads to the formation of a hexagonally packed monolayer of water droplets in the polymer film. The porous films were characterized by optical and scanning electron microscopy. Crosslinking was achieved by immersion in an ethanol solution of an α, ω alkyldiamine and the chemical reaction was monitored by infrared spectroscopy. The non‐crosslinked films are hydrophobic with a water contact angle of more than 90°, whereas the crosslinked films became hydrophilic, so that a water drop penetrated into the films. After crosslinking, the honeycomb structure was stable to up to 350 °C, an increase of more than 150 K as compared to the non‐crosslinked films.     

Rigidity‐Patterned Polyelectrolyte Films to Control Myoblast Cell Adhesion and Spatial Organization

In vivo, cells are sensitive to the stiffness of their microenvironment and to the spatial organization of the stiffness. In vitro studies of this phenomenon can help to better understand the mechanisms of the cell response to spatial variations of the matrix stiffness. Here, polelyelectrolyte multilayer films made of poly(L ‐lysine) and a photoreactive hyaluronan derivative are designed. These films can be photo‐crosslinked through a photomask to create spatial patterns of rigidity. Quartz substrates incorporating a chromium mask are prepared to expose selectively the film to UV light (in a physiological buffer), without any direct contact between the photomask and the soft film. It is shown that these micropatterns are chemically homogeneous and flat, without any preferential adsorption of adhesive proteins. Three groups of pattern geometries differing by their shape (circles or lines), size (from 2 to 100 μm), or interspacing distance between the motifs are used to study the adhesion and spatial organization of myoblast cells. The results pave the way for the study of the different steps of myoblast fusion in response to matrix rigidity in well‐defined geometrical conditions.     

Multifunctional Graphene–PEDOT Microelectrodes for On‐Chip Manipulation of Human Mesenchymal Stem Cells

All‐solution‐processed multifunctional organic bioelectronics composed of reduced graphene oxide (rGO) and dexamethasone 21‐phosphate disodium salt (DEX)‐loaded poly(3,4‐ethylenedioxythiophene) (PEDOT) microelectrode arrays on indium tin oxide glass are reported. They can be used to manipulate the differentiation of human mesenchymal stem cells (hMSCs). In the devices, the rGO material functions as an adhesive coating to promote the adhesion and alignment of hMSC cells and to accelerate their osteogenic differentiation. The poly(L ‐lysine‐graft‐ethylene glycol) (PLL‐g‐PEG)‐coated PEDOT electrodes serve as electroactive drug‐releasing electrodes. In addition, the corresponding three‐zone parallel devices operate as efficient drug‐releasing components through spatial‐temporal control of the release of the drug DEX from the PEDOT matrix. Such devices can be used for long‐term cell culturing and controlled differentiation of hMSCs through electrical stimulation.     

Charge‐Reversal Drug Conjugate for Targeted Cancer Cell Nuclear Drug Delivery

DNA‐toxin anticancer drugs target nuclear DNA or its associated enzymes to elicit their pharmaceutical effects, but cancer cells have not only membrane‐associated but also many intracellular drug‐resistance mechanisms that limit their nuclear localization. Thus, delivering such drugs directly to the nucleus would bypass the drug‐resistance barriers. The cationic polymer poly(L ‐lysine) (PLL) is capable of nuclear localization and may be used as a drug carrier for nuclear drug delivery, but its cationic charges make it toxic and cause problems in in‐vivo applications. Herein, PLL is used to demonstrate a pH‐triggered charge‐reversal carrier to solve this problem. PLL's primary amines are amidized as acid‐labile β‐carboxylic amides (PLL/amide). The negatively charged PLL/amide has a very low toxicity and low interaction with cells and, therefore, may be used in vivo. But once in cancer cells' acidic lysosomes, the acid‐labile amides hydrolyze into primary amines. The regenerated PLL escapes from the lysosomes and traverses into the nucleus. A cancer‐cell targeted nuclear‐localization polymer–drug conjugate has, thereby, been developed by introducing folic‐acid targeting groups and an anticancer drug camptothecin (CPT) to PLL/amide (FA‐PLL/amide‐CPT). The conjugate efficiently enters folate‐receptor overexpressing cancer cells and traverses to their nuclei. The CPT conjugated to the carrier by intracellular cleavable disulfide bonds shows much improved cytotoxicity.     

Short‐Time Tuning of the Biological Activity of Functionalized Polyelectrolyte Multilayers

This article demonstrates the tuning of the biological activity of a surface functionalized by a polyelectrolyte multilayer. The interaction of protein A with macrophages is used as the model system. The film consists of two polypeptides, poly(lysine) and poly(glutamic acid); each “build‐up” solution is a mixture of the respective D ‐ and L ‐enantiomers (d and l enantiomers). Cells are deposited on top of the film, and they produce tumor necrosis factor alpha (TNF‐α) as they come into contact with the protein. Depending upon the d/l‐enantiomer ratio of the polyelectrolyte solutions used for the film build‐up, and the embedding depth of the protein, the production of TNF‐α commences after a varying induction time and displays a transition from no‐production to full‐production, which takes place over a period of time that depends on the film's composition and embedding depth. Thus, it is shown that by changing these two parameters the timing of the protein's activity can be accurately tuned.     

A Facile In Situ Approach to Polypyrrole Functionalization Through Bioinspired Catechols

A template‐free benign approach to modify polypyrrole (PPy) with bioinspired catechol derivatives dopamine (DA), 1,2‐dihydroxybenzene or catechol (CA), and l ‐3,4‐dihydroxyphenylalanine (DOPA) is reported. It is found that PPy functionalized with these catechol derivatives (DA, CA, and DOPA) exhibited fibrous structure, smaller particle size, good water dispersibility, and enhanced film adhesion. Surprisingly, it is found that adding a small amount of catechols can also improve PPy's electrical conductivity. This rapid, one‐step, in situ, template‐free method provided an alternative strategy to the facile production of PPy fibers. Among these three catechols, functionalized PPy and DA‐PPy exhibits the smallest particle size and best performance in both adhesion and electrical conductivity. In contrast, the control phenylethlamine (PA) modification had almost negligible influence on the PPy properties, which provides strong evidence that instead of amine functional group or coexistence of both catechol and amine moieties, catechol itself is responsible for the successful functionalization of PPy and overall performance improvement. Furthermore, catechol‐PPy nanofibers are blended into polyvinyl alcohol (PVA) aqueous solution and casted to form thin films; as‐synthesized conductive films are found able to bond strongly onto the surface and may find broad applications in manufacturing biosensors and electronic devices.     

Pyridylamino‐β‐cyclodextrin as a Molecular Chaperone for Lipopolysaccharide Embedded in a Multilayered Polyelectrolyte Architecture

Layer‐by‐layer self‐assembled polyelectrolyte films containing a charged cyclodextrin and lipopolysaccharide (LPS) are developed for the first time as a potential model for local endotoxin antagonist delivery. We have examined the biological activity of a lipopolysaccharide from E. coli incorporated into multilayered architectures made of poly‐(L ‐lysine) and poly‐(L ‐glutamic acid). Used in such build‐ups, a polycationic cyclodextrin, heptakis(6‐deoxy‐6‐pyridylamino)‐β‐cyclodextrin showed molecular chaperone properties by enabling restoration of the LPS biological activity whenever lost upon interaction with poly‐(L ‐lysine).     

Copper‐Assisted Weak Polyelectrolyte Multilayer Formation on Microspheres and Subsequent Film Crosslinking

The formation of weak polyelectrolyte films on planar and spherical supports has recently evoked major interest, as such coatings allow novel material properties to be tunable by pH and salt adjustment of the polyelectrolyte deposition conditions. We report on the build up of multilayers of the weak polyelectrolytes poly(acrylic acid) (PAA) and poly(allylamine hydrochloride) (PAH) on submicrometer‐sized polystyrene (PS) and silica colloid spheres (～ 500 nm) with the aid of copper ion templating. The copper ions complex to the carboxylate groups of PAA, facilitating the formation of PAA/PAH multilayers on the particles. Regular growth of the layers on the colloid spheres with each polyelectrolyte deposition step was confirmed by microelectrophoresis, single‐particle light scattering (SPLS), and transmission electron microscopy (TEM), with an average bilayer thickness of ～ 3 nm. The polyelectrolyte multilayer‐coated particles formed stable colloidal dispersions, with ζ‐potentials ranging from 30 mV (PAH outer layer) and –50 mV (PAA outer layer). Complementary quartz‐crystal microbalance and UV‐vis spectrophotometry studies on PAA/PAH multilayers formed on planar supports were performed to examine the film formation and the role of copper ion binding to the layers. PAA/PAH multilayers formed on colloid particles were also chemically crosslinked by using the activator 1‐ethyl‐3‐(3‐dimethylaminopropyl) carbodiimide (EDC). The degree of film crosslinking could be readily controlled by varying the concentration of EDC employed. Following solvent decomposition of the template particles coated with crosslinked PAA/PAH multilayers, intact hollow polymer capsules were obtained. These capsules were found to be impenetrable to polystyrene.     

Creation of Superhydrophobic Electrospun Nonwovens Fabricated from Naturally Occurring Poly(Amino Acid) Derivatives

Creation of superhydrophobic materials bio‐inspired by nature fascinates many scientists. One of the most intriguing challenges in this field is the fabrication of these materials using biopolymers from the viewpoint of green chemistry and environmental chemistry. Here, superhydrophobic and biodegradable nonwovens are constructed by electrospinning from a naturally occurring poly(amino acid), poly(γ‐glutamic acid) (γ‐PGA), modified with a hydrophobic α‐amino acid, l ‐phenylalanine. The contact angle of a water droplet on the materials is 154°, and the droplet remains stuck to the material surface even if it is inverted, clearly indicating a petal‐type superhydrophobic property. Biodegradability and post‐functionalization of the nonwovens as well as cell adhesion on the superhydrophobic materials are also evaluated. As far as we know, this is the first report on biodegradable materials exhibiting a petal‐type superhydrophobicity. The material design and processing shown here can be applied to various bioresources and such functional materials will become a new class of functional materials satisfying some of the requirements in green science.     

On the Origin of Green Emission in Polyfluorene Polymers: The Roles of Thermal Oxidation Degradation and Crosslinking

The green emission of poly(9,9′′‐dioctylfluorenyl‐2,7′′‐diyl), end‐capped by polyhedral oligomeric silsequioxanes, (PFO‐POSS) has been investigated by photoluminescence (PL) and photoexcitation (PE), gel permeation chromatography (GPC), and transmission Fourier transform infrared (FTIR) spectroscopy. The green emission is closely correlated with thermal oxidation degradation and crosslinking of the polymer and is enhanced by annealing at elevated temperatures. The green‐to‐blue emission intensity ratio, used to assess the emission properties of thin (90 nm) films, was 3.70, 4.35, and 1.54 for an air‐annealed film, its insoluble residue (crosslinked), and a film cast from its soluble portion, respectively. For thick (5–6 μm) film, the ratios are 13.33, 13.33, and 0.79, respectively. However, FTIR spectroscopy of thick films leads to the conclusion that the carbonyl‐to‐aromatic ring concentration ratio are 0.018, 0.015, and 0.032, respectively. Focusing on the recast films, the green emission is relatively low while the carbonyl concentration is relatively high. This suggests that the energy traps at crosslinked chains play an important role in green emission. It is likely that the crosslinking enhances the excitation energy migration and energy transfer to the defects by hindering chain segment twisting.     

A Reusable Interface Constructed by 3‐Aminophenylboronic Acid‐Functionalized Multiwalled Carbon Nanotubes for Cell Capture,Release, and Cytosensing

A newly developed electrochemical cell sensor for the determination of K562 leukemia cells using 3‐aminophenylboronic acid (APBA)‐functionalized multiwalled carbon nanotubes (MWCNTs) films is demonstrated. The films are generated by the covalent coupling between the ? NH₂ groups in APBA and the ? COOH group in the acid‐oxidized MWCNTs. As a result of the sugar‐specific affinity interactions, the K562 leukemia cells are firmly bound to the APBA‐functionalized MWCNTs film via boronic acid groups. Compared to electropolymerized APBA films, the presence of MWCNTs not only provides abundant boronic acid domains for cell capture, their high electrical conductivity also makes the film suitable for electrochemical sensing applications. The resulting modified electrodes are tested as cell detection sensors. This work presents a promising platform for effective cell capture and constructing reusable cytosensors.     

Doping of Conjugated Polythiophenes with Alkyl Silanes

A strong modification of the electronic properties of solution‐processable conjugated polythiophenes by self‐assembled silane molecules is reported. Upon bulk doping with hydrolized fluoroalkyl trichlorosilane, the electrical conductivity of ultrathin polythiophene films increases by up to six orders of magnitude, reaching record values for polythiophenes: (1.1 ± 0.1) × 10³ S cm^?1 for poly(2,5‐bis(3‐tetradecylthiophen ‐2‐yl)thieno[3,2‐b]thiophene) (PBTTT) and 50 ± 20 S cm^?1 for poly(3‐hexyl)thiophene (P3HT). Interband optical absorption of the polymers in the doped state is drastically reduced, making these highly conductive films transparent in the visible range. The dopants within the porous polymer matrix are partially crosslinked via a silane self‐polymerization mechanism that makes the samples very stable in vacuum and nonpolar environments. The mechanism of SAM‐induced conductivity is believed to be based on protonic doping by the free silanol groups available within the partially crosslinked SAM network incorporated in the polythiophene structure. The SAM‐doped polythiophenes exhibit an intrinsic sensing effect: a drastic and reversible change in conductivity in response to ambient polar molecules, which is believed to be due to the interaction of the silanol groups with polar analytes. The reported electronic effects point to a new attractive route for doping conjugated polymers with potential applications in transparent conductors and molecular sensors.     

Self‐Defensive Biomaterial Coating Against Bacteria and Yeasts: Polysaccharide Multilayer Film with Embedded Antimicrobial Peptide

Prevention of pathogen colonization of medical implants is a major medical and financial issue since infection by microorganisms constitutes one of the most serious complications after surgery or critical care. Immobilization of antimicrobial molecules on biomaterials surfaces is an efficient approach to prevent biofilm formation. Herein, the first self‐defensive coating against both bacteria and yeasts is reported, where the release of the antimicrobial peptide is triggered by enzymatic degradation of the film due to the pathogens themselves. Biocompatible and biodegradable polysaccharide multilayer films based on functionalized hyaluronic acid by cateslytin (CTL), an endogenous host‐defensive antimicrobial peptide, and chitosan (HA‐CTL‐C/CHI) are deposited on a planar surface with the aim of designing both antibacterial and antifungal coating. After 24 h of incubation, HA‐CTL‐C/CHI films fully inhibit the development of Gram‐positive Staphylococcus aureus bacteria and Candida albicans yeasts, which are common and virulent pathogens agents encountered in care‐associated diseases. Hyaluronidase, secreted by the pathogens, leads to the film degradation and the antimicrobial action of the peptide. Furthermore, the limited fibroblasts adhesion, without cytotoxicity, on HA‐CTL‐C/CHI films highlights a medically relevant application to prevent infections on catheters or tracheal tubes where fibrous tissue encapsulation is undesirable.     

Mimicking Biological Phenol Reaction Cascades to Confer Mechanical Function

Phenol reaction cascades are commonly used in nature to create crosslinked materials that perform mechanical functions. These processes are mimicked by electrochemically initiating a reaction cascade to examine if the mechanical properties of a biopolymer film can be predictably altered. Specifically, thin films (≈ 30–45 μm) of the polysaccharide chitosan are cast onto gold‐coated silicon wafers, the chitosan‐coated wafers are immersed in catechol‐containing solutions, and the phenol is anodically oxidized. The product of this oxidation is highly reactive and undergoes reaction with chitosan chains adjacent to the anode. After reaction, the flexible chitosan film can be peeled from the wafer. Chemical and physical evidence support the conclusion that electrochemically initiated reactions crosslink chitosan. When gold is patterned onto the wafer, the electrochemical crosslinking reactions are spatially localized and impart anisotropic mechanical properties to the chitosan film. Further, deswelling of chitosan films can reversibly transduce environmental stimuli into contractile forces. Films patterned to have spatial variations in crosslinking respond to such environmental stimuli by undergoing reversible changes in shape. These results suggest the potential to enlist electrochemically initiated reaction cascades to engineer chitosan films for actuator functions.