Stimuli-responsive composite biopolymer actuators with selective spatial deformation behavior.

Bioinspired actuators with stimuli-responsive and deformable properties are being pursued in fields such as artificial tissues, medical devices and diagnostics, and intelligent biosensors. These applications require that actuator systems have biocompatibility, controlled deformability, biodegradability, mechanical durability, and stable reversibility. Herein, we report a bionic actuator system consisting of stimuli-responsive genetically engineered silk-elastin-like protein (SELP) hydrogels and wood-derived cellulose nanofibers (CNFs), which respond to temperature and ionic strength underwater by ecofriendly methods. Programmed site-selective actuation can be predicted and folded into three-dimensional (3D) origami-like shapes. The reversible deformation performance of the SELP/CNF actuators was quantified, and complex spatial transformations of multilayer actuators were demonstrated, including a biomimetic flower design with selective petal movements. Such actuators consisting entirely of biocompatible and biodegradable materials will offer an option toward constructing stimuli-responsive systems for in vivo biomedicine soft robotics and bionic research.

Processing-Structure-Properties Relationships of Glycerol-Plasticized Silk Films.

Silk possesses excellent mechanical properties and biocompatibility due to its unique protein sequences and hierarchical structures. Thus, it has been widely used as a biomaterial in a broad spectrum of biomedical applications. In this study, an in-depth investigation of glycerol-plasticized silk films was carried out to understand the processing-structure-properties relationships. A series of glycerol-plasticized silk films with glycerol contents in the range of 0 to 30% (w/w) were prepared. The molecular structures and organizations of silk proteins and the interactions between glycerol and proteins were studied using FTIR, XRD, and DSC. At a low glycerol content (<12%), DSC revealed that the glass transition temperature and thermally induced crystallization temperature decreased as the glycerol content increased, implying that glycerol mainly interacts with silk proteins through hydrogen bonding. As the glycerol content further increased, the chain mobility of the silk proteins was promoted, leading to the formation of ß-sheet structures, water insolubility, and increased crystallinity. In addition, the stretchability and toughness of the films were significantly enhanced. The role of glycerol as a plasticizer in regulating the silk protein structures and determining the properties of the films was thoroughly discussed.

Recent Advances in 3D Printing with Protein-Based Inks.

Three-dimensional (3D) printing is a transformative manufacturing strategy, allowing rapid prototyping, customization, and flexible manipulation of structure-property relationships. Proteins are particularly appealing to formulate inks for 3D printing as they serve as essential structural components of living systems, provide a support presence in and around cells and for tissue functions, and also provide the basis for many essential ex vivo secreted structures in nature. Protein-based inks are beneficial in vivo due to their mechanics, chemical and physical match to the specific tissue, and full degradability, while also to promoting implant-host integration and serving as an interface between technology and biology. Exploiting the biological, chemical, and physical features of protein-based inks can provide key opportunities to meet the needs of tissue engineering and regenerative medicine. Despite these benefits, protein-based inks impose nontrivial challenges to 3D printing such as concentration and rheological features and reconstitution of the structural hierarchy observed in nature that is a source of the robust mechanics and functions of these materials. This review introduces photo-crosslinking mechanisms and rheological principles that underpins a variety of 3D printing techniques. The review also highlights recent advances in the design, development, and biomedical utility of monolithic and composite inks from a range of proteins, including collagen, silk, fibrinogen, and others. One particular focus throughout the review is to introduce unique material characteristics of proteins, including amino acid sequences, molecular assembly, and secondary conformations, which are useful for designing printing inks and for controlling the printed structures. Future perspectives of 3D printing with protein-based inks are also provided to support the promising spectrum of biomedical research accessible to these materials.

Thermoplastic moulding of regenerated silk.

Early insights into the unique structure and properties of native silk suggested that ß-sheet nanocrystallites in silk would degrade prior to melting when subjected to thermal processing. Since then, canonical approaches for fabricating silk-based materials typically involve solution-derived processing methods, which have inherent limitations with respect to silk protein solubility and stability in solution, and time and cost efficiency. Here we report a thermal processing method for the direct solid-state moulding of regenerated silk into bulk 'parts' or devices with tunable mechanical properties. At elevated temperature and pressure, regenerated amorphous silk nanomaterials with ultralow ß-sheet content undergo thermal fusion via molecular rearrangement and self-assembly assisted by bound water to form a robust bulk material that retains biocompatibility, degradability and machinability. This technique reverses presumptions about the limitations of direct thermal processing of silk into a wide range of new material formats and composite materials with tailored properties and functionalities.

Spinning Regenerated Silk Fibers with Improved Toughness by Plasticizing with Low Molecular Weight Silk.

Low-molecular weight (LMW) silk was utilized as a LMW silk plasticizer for regenerated silk, generating weak physical crosslinks between high-molecular weight (HMW) silk chains in the amorphous regions of a mixed solution of HMW/LMW silk. The plasticization effect of LMW silk was investigated using mechanical testing, Raman spectroscopy, and wide-angle X-ray scattering (WAXS). Small amounts (10%) of LMW silk resulted in a 19.4% enhancement in fiber extensibility and 37.8% increase in toughness. The addition of the LMW silk facilitated the movement of HMW silk chains during drawing, resulting in an increase in molecular chain orientation when compared with silk spun from 100% HMW silk solution. The best regenerated silk fibers produced in this work had an orientation factor of 0.94 and crystallinity of 47.82%, close to the values of natural degummedBombyx mori silk fiber. The approach and mechanism elucidated here can facilitate artificial silk systems with enhanced properties.

Geochemical stability of zero-valent iron modified raw wheat straw innovatively applicated to in situ permeable reactive barrier: N2 selectivity and long-term denitrification.

The zero-valent iron (ZVI) modified wheat straw materials are widely used for treating groundwater by permeable reactive barrier (PRB). We report the performance of a field-scale PRB filled with ZVI modified wheat straw materials for nitrate (NO3-)-contaminated groundwater. In lab-scale PRB filled with ZVI modified wheat straw material, NO3- concentration entering the PRB was varied (27.80-59.86 mg L-1) according to the in situ NO3- contamination. A stable NO3- removal rate of 90% was achieved at a controlled hydraulic retention time of 22 days, together with a proportion of denitrifying bacteria up to 34.37%. The field-scale PRB filled with ZVI modified wheat straw material was successful at removing NO3- from groundwater (removal percentages ≥60%) at a groundwater flow rate of 0.01 m3 d-1. Monitoring of groundwater within this PRB provided evidences that the nitrogen gas (N2) selectivity increased with lower ammonia (NH4+) generated from ZVI reduction of NO3-, and few emission of NO2- present due to denitrification capacity in this PRB. The results are finally compared with the few others reported existing PRBs for nitrate-contaminated groundwater worldwide, and demonstrated that the ZVI modified wheat straw material would be an effective fillings for field PRB to remediate groundwater.

Bi-layered Tubular Microfiber Scaffolds as Functional Templates for Engineering Human Intestinal Smooth Muscle Tissue.

Designing biomimetic scaffolds with in vivo-like microenvironments using biomaterials is an essential component of successful tissue engineering approaches. The intestinal smooth muscle layers exhibit a complex tubular structure consisting of two concentric muscle layers in which the inner circular layer is orthogonally oriented to the outer longitudinal layer. Here, we present a three-dimensional (3D) bi-layered tubular scaffold based on flexible, mechanically robust and well aligned silk protein microfibers to mimic native human intestinal smooth muscle structure. The scaffolds were seeded with primary human intestinal smooth muscle cells to replicate human intestinal muscle tissues in vitro. Characterization of the tissue constructs revealed good biocompatibility and support for cell alignment and elongation in the different scaffold layers to enhance cell differentiation and functions. Furthermore, the engineered smooth muscle constructs supported oriented neurite outgrowth, a requisite step to achieve functional innervation. These results suggested these microfiber scaffolds as functional templates for in vitro regeneration of human intestinal smooth muscle systems. The scaffolding provides a crucial step toward engineering functional human intestinal tissue in vitro, as well as for the engineering of many other types of smooth muscles in terms of their similar phenotypes. Such utility may lead to a better understanding of smooth muscle associated diseases and treatments.

Enzymatic Degradation of Bombyx mori Silk Materials: A Review.

As a biomaterial, silk presents unique features with a combination of excellent mechanical properties, biocompatibility, and biodegradability. The biodegradability aspects of silk biomaterials, especially with options to control the rate from short (days) to long (years) time frames in vivo, make this protein-based biopolymer a good candidate for developing biodegradable devices used for tissue repairs and tissue engineering, as well as medical device implants. Silk materials, including native silk fibers and a broad spectrum of regenerated silk materials, have been investigated in vitro and in vivo to demonstrate degradation by proteolytic enzymes. In this Review, we summarize the findings on these studies on the enzymatic degradation of Bombyx mori (B. mori) silk materials. We also present a discussion on the factors that dictate the degradation properties of silk materials. Finally, in future perspectives, we highlight some key challenges and potential directions toward the future study of the degradation of silk materials.

Probing the binding modes and dynamics of histidine on fumed silica surfaces by solid-state NMR.

Silica nanoparticles can be designed to exhibit a diverse range of morphologies (e.g. non-porous, mesoporous), physical properties (e.g. hydrophobic, hydrophilic) and a wide range of chemical and biomolecular surface functionalizations. In the present work, the adsorption complex of histidine (His) and fumed silica nanoparticles (FSN) is probed using thermal analysis (TGA/DTG) and a battery of solid-state (SS) NMR methods supported by DFT chemical shift calculations. Multinuclear (1H/13C/15N) one- and two-dimensional magic angle spinning (MAS) SSNMR experiments were applied to determine site-specific interactions between His and FSN surfaces as a function of adsorption solution concentration, pH and hydration state. By directly comparing SSNMR observables (linewidth, chemical shift and relaxation parameters) for His-FSN adsorption complexes to various crystalline, amorphous and aqueous His forms, the His structural and dynamic environment on FSN surfaces could be determined at an atomic level. The observed 13C and 15N MAS NMR chemical shifts, linewidths and relaxation parameters show that the His surface layer on FSN has a significant dependence on pH and hydration state. His is highly dynamic on FSN surfaces under acidic conditions (pH 4) as evidenced by sharp resonances with near isotropic chemical shifts regardless of hydration level indicating a non-specific binding arrangement while, a considerably more rigid His environment with defined protonation states is observed at near neutral pH with subtle variations between hydrated and anhydrous complexes. At near neutral pH, less charge repulsion occurs on the FSN surface and His is more tightly bound as evidenced by considerable line broadening likely due to chemical shift heterogeneity and a distribution in hydrogen-bonding strengths on the FSN surface. Multiple His sites exchange with a tightly bound water layer in hydrated samples while, direct interaction with the FSN surface and significant chemical shift perturbations for imidazole ring nitrogen sites and some carbon resonances are observed after drying. The SSNMR data was used to propose an interfacial molecular binding model between His and FSN surfaces under varying conditions setting the stage for future multinuclear, multidimensional SSNMR studies of His-containing peptides on silica nanoparticles and other nanomaterials of interest.

Rapid Soft Tissue Approximation and Repair using Laser-activated Silk Nanosealants.

Tissue approximation and repair have been conventionally performed with sutures and staples, but these means are inherently traumatic. Tissue approximation using laser-responsive nanomaterials can lead to rapid tissue sealing and repair, and is an attractive alternative to existing clinical methods. Here, we demonstrate the use of laser-activated nanosealants (LANS) with gold nanorods (GNRs) embedded in silk fibroin polypeptide matrices. The adaptability of LANS for sealing soft tissues is demonstrated using two different modalities: insoluble thin films for internal, intestinal tissue repair, and semi-soluble pastes for external repair, shown by skin repair in live mice. Laser repaired intestinal tissue held over seven times more fluid pressure than sutured intestine and also prevented bacterial leakage. Skin incisions in mice closed using LANS' showed indication of increased mechanical strength and faster repair compared to suturing. Laser-activated silk-GNR nanosealants rapidly seal soft-tissue tears and show high promise for tissue approximation and repair in trauma and routine surgery.

Structural Comparison of Various Silkworm Silks: An Insight into the Structure-Property Relationship.

Silkworm silk has attracted considerable attention in recent years due to its excellent mechanical properties, biocompatibility, and promising applications in biomedical sector. However, a clear understanding of the molecular structure and the relationship between the excellent mechanical properties and the silk protein sequences are still lacking. This study carries out a thorough comparative structural analysis of silk fibers of four silkworm species ( Bombyx mori, Antheraea pernyi, Samia cynthia ricini, and Antheraea assamensis). A combination of characterization techniques including scanning electron microscopy, mechanical test, synchrotron X-ray diffraction, Fourier transform infrared spectroscopy (FTIR), and NMR spectroscopy was applied to investigate the morphologies, mechanical properties, amino acid compositions, nanoscale organizations, and molecular structures of various silkworm silks. Furthermore, the structure-property relationship is discussed by correlating the molecular structural features of silks with their mechanical properties. The results show that a high content of ß-sheet structures and a high crystallinity would result in a high Young's modulus for silkworm silk fibers. Additionally, a low content of ß-sheet structures would result in a high extensibility.

Characterizing gold nanoparticles by NMR spectroscopy.

Gold nanoparticles have attracted considerable attention in recent research because of their wide applications in various fields such as material science, electrical engineering, physical science, and biomedical engineering. Researchers have developed many methods for synthesizing different kinds of gold nanoparticles, where the sizes and surface chemistry of the nanoparticles are considered to be the two key factors. Traditionally, the sizes of nanoparticles are determined by electron microscopy whereas the surface chemistry is characterized by optical spectroscopies such as infrared spectroscopy and Raman spectroscopy. Compared with that, nuclear magnetic resonance (NMR) spectroscopy provides a more advanced and convenient way for size determination and surface chemistry investigations by combining one- and multiple-dimensional NMR spectroscopy and diffusion-order NMR spectroscopy. Here, we show a thorough study that NMR spectroscopy can be applied to characterize small thiol-protected gold nanoparticles, including size determination, surface chemistry investigation, and structural study. The results show that the nanoparticles' sizes determined by NMR agree well with transmission electron microscopy results. Furthermore, the ligand densities of nanoparticles were determined by quantitative NMR spectroscopy, and the structures of ligands capped on the surfaces were studied thoroughly by one- and multiple-dimensional NMR spectroscopy. In this work, we establish a general method for researchers to characterize nanostructures by using NMR spectroscopy.

Comparative Study of Strain-Dependent Structural Changes of Silkworm Silks: Insight into the Structural Origin of Strain-Stiffening.

Structure-property relationships of silk is an intriguing topic for silk-based biomaterials research since these features are related to biomimicking the processing in natural silk fiber formation which results in excellent mechanical properties. Strain-stiffening is common for spider silks and nonmulberry silkworm silks. However, the structural origin of strain-stiffening remains unclear. In this paper, the strain-dependent structural change of Antheraea pernyi silkworm silk is studied by X-ray fiber diffraction and Fourier transform infrared spectroscopy under stretching. Based on a combination of mechanical and structural analysis, the molecular origins of strain-stiffening in A. pernyi silk were determined. The relatively high content of the ß-sheets within the amorphous domains in A. pernyi silk is responsible for strain-stiffening, where "molecular spindles" enhance the extensibility and toughness of the fiber.

Understanding iridium oxide nanoparticle surface sites by their interaction with catechol.

Iridium oxide (IrOx) is one of the best water splitting electrocatalysts, but its active site details are not well known. As with all heterogeneous catalysts, a strategy for counting the number of active sites is not clear, and understanding their nature and structure is remarkably difficult. In this work, we performed a combined study using optical spectroscopy, magnetic resonance and electrochemistry to characterize the interaction of IrOx nanoparticles (NPs) with a probe molecule, catechol. The catalyst is heterogeneous given that the substrate is in a different phase, but behaves as a homogeneous catalyst from the point of view of electrochemistry since it remains in colloidal suspension. We find two types of binding sites: centers A which bind catechol irreversibly making up 21% of the surface, and centers B which bind catechol reversibly making up 79% of the surface. UV-vis absorption spectroscopy shows that the A sites are responsible for the characteristic blue color of the NPs. Electrochemical experiments indicate that the B sites are catalytically active and we give the number of active sites per nanoparticle. We conclude by performing a survey of ligands used in solar cell architectures and show which ones bind well to the surface and which ones inhibit the catalytic activity when doing so, presenting quantitative guidelines for the correct handling of IrOx nanoparticles during their incorporation into multifunctional solar energy harvesting architectures. We suggest ligands binding on the surface oxygen atoms allow for large bound ligand densities with no detrimental effect on the catalytic activity.

Secondary Structure Adopted by the Gly-Gly-X Repetitive Regions of Dragline Spider Silk.

Solid-state NMR and molecular dynamics (MD) simulations are presented to help elucidate the molecular secondary structure of poly(Gly-Gly-X), which is one of the most common structural repetitive motifs found in orb-weaving dragline spider silk proteins. The combination of NMR and computational experiments provides insight into the molecular secondary structure of poly(Gly-Gly-X) segments and provides further support that these regions are disordered and primarily non-ß-sheet. Furthermore, the combination of NMR and MD simulations illustrate the possibility for several secondary structural elements in the poly(Gly-Gly-X) regions of dragline silks, including ß-turns, 310-helicies, and coil structures with a negligible population of α-helix observed.

Probing the Impact of Acidification on Spider Silk Assembly Kinetics.

Spiders utilize fine adjustment of the physicochemical conditions within its silk spinning system to regulate spidroin assembly into solid silk fibers with outstanding mechanical properties. However, the exact mechanism about which this occurs remains elusive and is still hotly debated. In this study, the effect of acidification on spider silk assembly was investigated on native spidroins from the major ampullate (MA) gland fluid excised from Latrodectus hesperus (Black Widow) spiders. Incubating the protein-rich MA silk gland fluid at acidic pH conditions results in the formation of silk fibers that are 10-100 µm in length and ∼2 µm in diameter as judged by optical and electron microscope methods. The in vitro spider silk assembly kinetics were monitored as a function of pH with a (13)C solid-state MAS NMR approach. The results confirm the importance of acidic pH in the spider silk self-assembly process with observation of a sigmoidal nucleation-elongation kinetic profile. The rates of nucleation and elongation as well as the percentage of ß-sheet structure in the grown fibers depend on the pH. These results confirm the importance of an acidic pH gradient along the spinning duct for spider silk formation and provide a powerful spectroscopic approach to probe the kinetics of spider silk formation under various biochemical conditions.

Characteristics of zero-valent iron surface oxide films under the catalytic interface reactions by assisting ligands in nitrate-contaminated groundwater.

The surface passivation layer coating on zero-valent iron (ZVI) particles impedes the electron transfer from ZVI to nitrate. To enhance the efficiency of nitrate reduction by Fe(0), we tested the chemical process and the thickness of the iron oxide film on the surface of Fe(0) particles, utilizing Fe2+aq in aqueous solution and wheat straw as ligands. A novel principal surface catalyzing reaction was formulated as follows: [Formula: see text] . When Fe2+aq concentration increased from 0 - 200 mg·L-1, the NO3- removal rate increased from 6.95% to 82.6% respectively during 12 h and it was 48%, 72%, 79% and 94% respectively in Fe0/WS ratio of 0, 0.25, 0.5 and 1 system. Uniform surface iron oxide films formed around the Fe(0) particles within 12 h after the adding Fe2+aq or wheat straw to the Fe(0) system. The composition and thickness of these films were dependent on the quantity of added materials. X-ray diffraction (XRD) analysis revealed that surface oxide iron mainly consisted of Fe2+ or Fe3+ oxides, with Fe3O4 being predominant. The X-ray photoelectron spectroscopy (XPS) etching indicated that the addition of Fe(0)/straw at mass ratios of 1 or system with 20 mg·L-1 Fe2+aq resulted in the thinnest surface iron oxide layer. The study demonstrated that reducing the oxide layer's thickness was achieved through partial catalysis and enhanced complexation capacity. This reduction was facilitated by the introduction of Fe2+aq or wheat straw into the Fe(0) system, potentially improving proton dissociation and promoting the ligand-assisted dissolution of Fe3+ oxides.

Injectable cell-laden silk acid hydrogel.

This study presents an injectable cell-laden hydrogel system based on silk acid, a carboxylated derivative of natural silk fibroin, which exhibits promising applications in biomedicine. The hydrogel is produced under physiological conditions (37 °C and pH 7.4) via physical crosslinking. Notably, the hydrogel demonstrates remarkable cytocompatibility, enabling efficient cell encapsulation, and exhibits good injectability. These promising results strongly indicate the potential of silk acid hydrogel for transformative applications, including 3D cell culture, targeted cell delivery, and tissue engineering.

Fabrication and Evaluation of Hyaluronidase-Responsive Scaffolds by Electrospinning with Antibacterial Properties for Tympanic Membrane Repair.

Tympanic membrane perforation (TMP) is prevalent in clinical settings. Patients with TMPs often suffer from infections caused by Staphylococcus aureus and Pseudomonas aeruginosa, leading to middle ear and external ear canal infections, which hinder eardrum healing. The objective of this study is to fabricate an enzyme-responsive antibacterial electrospun scaffold using poly(lactic-co-glycolic acid) and hyaluronic acid for the treatment of infected TMPs. The properties of the scaffold were characterized, including morphology, wettability, mechanical properties, degradation properties, antimicrobial properties, and biocompatibility. The results indicated that the fabricated scaffold had a core-shell structure and exhibited excellent mechanical properties, hydrophobicity, degradability, and cytocompatibility. Furthermore, in vitro bacterial tests and ex vivo investigations on eardrum infections suggested that this scaffold possesses hyaluronidase-responsive antibacterial properties. It may rapidly release antibiotics when exposed to the enzyme released by S. aureus and P. aeruginosa. These findings suggest that the scaffold has great potential for repairing TMPs with infections.

Rapid UV Photo-Cross-Linking of α-Lactalbumin Hydrogel Biomaterial To Enable Wound Healing.

Although both the function and biocompatibility of protein-based biomaterials are better than those of synthetic materials, their usage as medical material is currently limited by their high costs, low yield, and low batch-to-batch reproducibility. In this article, we show how α-lactalbumin (α-LA), rich in tryptophan, was used to produce a novel type of naturally occurring, protein-based biomaterial suitable for wound dressing. To create a photo-cross-linkable polymer, α-LA was methacrylated at a 100-g batch scale with >95% conversion and 90% yield. α-LAMA was further processed using photo-cross-linking-based advanced processing techniques such as microfluidics and 3D printing to create injectable hydrogels, monodispersed microspheres, and patterned scaffolds. The obtained α-LAMA hydrogels show promising biocompatibility and degradability during in vivo testing. Additionally, the α-LAMA hydrogel can accelerate post-traumatic wound healing and promote new tissue regeneration. In conclusion, cheap and safe α-LAMA-based biomaterials could be produced, and they have a beneficial effect on wound healing. As a result, there may arise a potential partnership between the dairy industry and the development of pharmaceuticals.