Peptide aerogels comprising self-assembling nanofibrils

Bioinspired molecular self-assembly is a popular route to novel functional materials for industrial applications. Here we explore, for the first time, the possibility of using organogel and hydrogels of short self-assembling beta-sheet-forming peptides, as starting states for the creation of nanostructured peptide aerogels. The effects of supercritical fluid drying (SCF) and freeze-drying (FD) on the nanofibrillar peptide gel network were investigated. SCF processing was found to cause collapse of peptide organogel networks, presumably because of peptide insolubility in carbon dioxide (CO₂). Freeze-drying of peptide hydrogels proved a more efficient method of removing the solvent without destroying the self-assembled fibrillar network, leading to a microscopic aligned lamellar structure consisting of thousands of stacked peptide nanofibrils. These chiral, nanostructured, low-density aerogels are characterised by chemical versatility and regular display of functional groups on their surface. Appropriately designed peptides can also be triggered to self-assemble in situ inside other porous structures and impart biological-like functionality for catalysis, sensing, separation and filtration applications, or tissue engineering.     

Stimuli-Responsive Peptide Self-Assembly to Construct Hydrogels with Actuation and Shape Memory Behaviors

Hydrogel actuators, capable of generating reversible deformation in response to external stimulus, are widely considered as new emerging intelligent materials for applications in soft robots, smart sensors, artificial muscles, and so on. Peptide self-assembly is widely applied in the construction of intelligent hydrogel materials due to their excellent stimulus response. However, hydrogel actuators based on peptide self-assembly are rarely reported and explored. In this study, a pH-responsive peptide (MA-FIID) is designed and introduced into a poly(N-isopropyl acrylamide) backbone (PNIPAM) to construct bilayer and heterogeneous hydrogel actuators based on the assembly and disassembly of peptide molecules under different pH conditions. These peptide-containing hydrogel actuators can perform controllable bending, bucking, and complex deformation under pH stimulation. Meanwhile, the Hofmeister effect of PNIPAM hydrogels endows these peptide-containing hydrogels with enhanced mechanical strength, ionic stimulus response (CaCl₂), and excellent shape-memory property. This work broadens the application of supramolecular self-assembly in the construction of intelligent hydrogels, and also provides new inspirations for peptide self-assembly to construct smart materials.     

Treatment of Superbug Infection through a Membrane-Disruption and Immune-Regulation Cascade Effect Based on Supramolecular Peptide Hydrogels

Infections from antimicrobial resistant pathogens have been considered as lethal threats to human health if antibiotic treatments become ineffective. Antimicrobial peptides (AMPs) show broad-spectrum antimicrobial activity against resistant strains via a membrane disruptive mechanism without developing further resistance and are promising candidates to combat resistant microbes. However, currently developed small molecular AMPs are limited by weak antimicrobial efficacy and potential cytotoxicity. Herein, an antimicrobial AMP hydrogel based on supramolecular self-assembly is developed. The hydrogel shows strong antimicrobial activity and high potency against antimicrobial resistant microbes. It also possesses high cytocompatibility and immunoregulative activity. The synergistic effects lead to its significant healing of the skin abscess disease induced by the methicillin-resistant Staphylococcus aureus infection. This study not only sheds light on the design strategies and immune-related antimicrobial actions of supramolecular AMP hydrogels, but also advances the development of self-assembly peptide platform to fight against antimicrobial resistant infections.     

Evolution‐Based Design of an Injectable Hydrogel

A new class of simple, linear, amphiphilic peptides are developed that have the ability to undergo triggered self‐assembly into self‐supporting hydrogels. Under non‐gelling aqueous conditions, these peptides exist in a random coil conformation and peptide solutions have the viscosity of water. On the addition of a buffered saline solution, the peptides assemble into a β‐sheet rich network of fibrils, ultimately leading to hydrogelation. A family of nine peptides is prepared to study the influence of peptide length and amino acid composition on the rate of self‐assembly and hydrogel material properties. The amino acid composition is modulated by varying residue hydrophobicity and hydrophilicity on the two opposing faces of the amphiphile. The conformation of peptides in their soluble and gel state is studied by circular dichroism (CD), while the resultant material properties of their gels is investigated using oscillatory sheer rheology. One weight percent gels formed under physiological conditions have storage modulus (G′) values that vary from ≈20 to ≈800 Pa, with sequence length and hydrophobic character playing a dominant roll in defining hydrogel rigidity. Based on the structural and functional data provided by the nine‐peptide family members, an optimal sequence, namely LK13, is evolved. LK13 (LKLKLKLKLKLKL‐NH₂) undergoes triggered self‐assembly, affording the most rigid gel of those studied (G′=797 ± 105). It displays shear thin‐recovery behavior, allowing its delivery by syringe and is cytocompatibile as assessed with murine C3H10t1/2 mesenchymal stem cells.     

Bioinspired Self-Assembling Materials for Modulating Enzyme Functions

Enzymes tend to malfunction when they work out of their natural cellular environments. Engineering a favorable microenvironment around enzymes has emerged as an effective strategy to finely tune the enzymatic functions and reshape the biocatalytic activities. Supramolecular self-assembly provides a bottom-up approach for spatial arrangement of functional groups and fabrication of materials with tailorable local properties. In this review, the progress in designing, creating, and tailoring the enzyme microenvironments is discussed, with the bioinspired self-assembling materials as the scaffolds built from molecular building blocks. The relationship between the physicochemical properties and the local environments (pH, substrates, or hydration) of the scaffolds, and the catalytic properties of the scaffolded enzymes are focused upon. The power of the self-assembly to regulate the catalytic systems dynamically is also highlighted. In the end, an outlook on the obstacles, possible solutions, and future directions on the microenvironment engineering of enzymes is provided.     

Achieving Swollen yet Strengthened Hydrogels by Reorganizing Multiphase Network Structure

Many living tissues, such as muscle, become mechanically stronger with growth. Yet, synthetic hydrogels usually exhibit an opposite size-mechanical property relation, that is, swelling-weakening behavior. Herein, a series of swollen yet strengthened polyampholyte (PA) hydrogels are developed via a simple metal-ion solution soaking strategy. In this strategy, a dynamic PA hydrogel (with ionic bonds) is dialyzed in ZrOCl₂ solutions (Step-I) and deionized water (Step-II) successively to obtain equilibrated hydrogels. Due to the specific Zr⁴⁺ ions and PA network structure, Step-I takes several months with sample size and mechanical performance increasing continuously, while Step-II only needs several days. Through this strategy, the resultant hydrogel networks are reorganized and eventually constructed by ionic and metal ligand bonds, enabling the swelling yet strengthening behavior. A systematic study confirms that dialysis time in Step-I and corresponding ZrOCl₂ concentration can significantly affect the multiphase microstructures of the hydrogels, resulting in different mechanical enhancements. The optimized hydrogel possesses 39.2 MPa of Young's modulus and 3.7 MPa of tensile strength, which are 302 and 5.5 times these of the original PA gel, respectively. Despite distinct swelling, these hydrogels still mechanically surpass many existing high-performance hydrogels. This study opens a novel pathway for fabricating swollen yet strengthened hydrogels.     

Triple Enzyme-Regulated Molecular Hydrogels for Carrier-Free Delivery of Lonidamine

Cancerous cells exhibit overexpression of multiple enzymes in various cellular compartments. These enzymes are often undruggable, yet display unique advantages in regulating intracellular self-assembly and dis-assembly of small molecules for cancer targeting. Herein, a self-assembling molecule (LND-1p-ES) for carrier-free delivery of lonidamine specifically to cancerous cells is designed, where LND-1p-ES executes as both a drug and a carrier with combined benefits thereof. Under the precise regulation of phosphatase (ALP) and esterase (CES), LND-1p-ES is capable of self-assembling intracellularly in a spatiotemporal manner, to confer lonidamine-borne nanofibers with enhanced cellular uptake. These nanofibers also facilitate controlled drug release with the aid of cellular proteases. Taking advantages of overexpressing ALP and CES as well as proteases in cancerous cells, the LND-1p-ES formulation demonstrates enhanced potency and selectivity against melanoma cells A375 in vitro and in vivo. In comparison, none/single enzyme responsive compounds fail to show a similar potency or selectivity, further confirming the indispensable roles of these enzymes in the delivery system. Collectively, the research provides a viable strategy to utilize multiple enzymes in cancerous cells for regulation of intracellular self-assembly, which can be expanded to design smart soft materials responsive to multiple biologically relevant biomolecules for enhanced therapeutic efficacy.     

Hydrogel Scaffolds of Amphiphilic and Acidic β‐Sheet Peptides

Amphiphilic and acidic β‐sheet‐forming peptides (AAβPs) having the sequence Pro‐Y‐(Z‐Y)₅‐Pro, Y = Glu or Asp and Z = Phe or Leu may assemble into hydrogel structures at near neutral pH values, several units higher than the intrinsic pKa of their acidic amino acid side chains. The bottom‐to‐top design strategy enables the rationally supported association between the peptides' amino acids composition and bulk pH hydrogelation. Hydrogen bonds between the acidic amino acids side chains in the β‐sheet structure are found to contribute substantially to the stabilization of AAβPs hydrogels. The negatively charged peptides are also found to form gels at lower concentration in presence of calcium ions. Bone forming cells may be cultured on two‐dimensional films of AAβPs hydrogels that form at physiological pH values as well as within three dimensional hydrogel matrices. These acidic‐rich peptides hydrogels may become advantageous in applications related to engineering of mineralized tissues providing controllable, multifunctional calcified scaffolds to affect both the biological activity and the inorganic mineralization.     

Capillary Network‐Like Organization of Endothelial Cells in PEGDA Scaffolds Encoded with Angiogenic Signals via Triple Helical Hybridization

Survival of tissue engineered constructs after implantation depends on proper vascularization. The differentiation of endothelial cells into mature microvasculature requires dynamic interactions between cells, scaffold, and growth factors, which are difficult to recapitulate in artificial systems. Previously, photocrosslinked poly(ethylene glycol) diacrylate (PEGDA) hydrogels displaying collagen mimetic peptides (CMPs), dubbed PEGDA‐CMP, that can be further conjugated with bioactive molecules via CMP‐CMP triple helix hybridization were reported. Here, it is shown that a bifunctional peptide featuring pro‐angiogenic domain mimicking vascular endothelial growth factor (VEGF) and a collagen mimetic domain that can fold into a triple helix conformation can hybridize with CMP side chains of the PEGDA‐CMP hydrogel, which results in presentation of insoluble VEGF‐like signals to endothelial cells. Presentation of VEGF‐like signals on the surface of micropatterned scaffolds in this way transforms cells from a quiescent state to elongated and aligned phenotype suggesting that this system could be used to engineer organized microvasculature. It is also shown that the pro‐angiogenic peptide, when applied topically in combination with modified dextran/PEGDA hydrogels, can enhance neovascularization of burn wounds in mice demonstrating the potential clinical use of CMP‐mediated matrix‐bound bioactive molecules for dermal injuries.     

Surface Decoration of Peptide Nanoparticles Enables Efficient Therapy toward Osteoporosis and Diabetes

A versatile surface decoration strategy to efficiently encapsulate water-soluble peptides is developed. By assembling peptide molecules into nanoparticles, diverse physiochemical properties of these compacted molecules are equalized to the surface properties of nanoparticles. Primarily driven by the generic electrostatic attractions, the surface of as-prepared peptide nanoparticles is decorated with charged amino acids-grafted poly(lactic-co-glycolic acid). This adsorbed polymer layer versatilely blocks the phase transfer of peptide nanoparticles by increasing their affinity to the dispersed phase solvent molecules. Attributed to the ultrahigh encapsulation efficiencies (> 96%), the peptide mass fraction inside the obtained microcomposites is higher than 48%. The plasma calcium level has been efficiently reduced for ≈3 weeks with only one single injection of salmon calcitonin-encapsulated microcomposite in osteoporotic rats. Similarly, one single injection of exenatide-encapsulated microcomposites efficiently controls the glycemic level in type 2 diabetic rats for up to 3 weeks. Overall, the developed versatile surface decoration strategy efficiently encapsulates peptides and improves their pharmacokinetic features, regardless of the molecular structure of peptide cargos.     

Strong Tough Polyampholyte Hydrogels via the Synergistic Effect of Ionic and Metal–Ligand Bonds

Despite existing in biological systems, developing synthetic polyampholyte (PA) hydrogels constructed by both ionic and metal–ligand bonds remains challenging. Herein, a simple secondary equilibrium approach is proposed to fabricate strong and tough PA hydrogels via the synergy of ionic and metal–ligand bonds. The original PA gels (constructed by ionic bonds) are first dialyzed in multivalent metal-ion solutions to reach a swelling equilibrium and then moved to deionized water to dialyze excess free ions to achieve a new equilibrium. Through this approach, the original PA gel network can be optimized and eventually constructed by ionic and metal–ligand bonds, enabling a synergistic reinforcement. By selecting different original PA gel systems and diverse multivalent metal-ions, the proposed approach is proved to be generalizable to fabricate strong and tough PA gels. Additionally, the hydrogels have stable ion-conductivity even at the water-equilibrium state, making them promising as strain sensors. The viscoelastic and elastic contributions to the mechanical properties of the hydrogels by a viscoelastic model are also discussed to further understand the strengthening and toughening mechanisms. The proposed strategy is simple but effective for achieving strong and tough PA-based hydrogels. This study also provides new insights for PA hydrogels in electrolyte environments.     

High‐Strength,Durable All‐Silk Fibroin Hydrogels with Versatile Processability toward Multifunctional Applications

Hydrogels are the focus of extensive research due to their potential use in fields including biomedical, pharmaceutical, biosensors, and cosmetics. However, the general weak mechanical properties of hydrogels limit their utility. Here, pristine silk fibroin (SF) hydrogels with excellent mechanical properties are generated via a binary‐solvent‐induced conformation transition (BSICT) strategy. In this method, the conformational transition of SF is regulated by moderate binary solvent diffusion and SF/solvent interactions. β‐sheet formation serves as the physical crosslinks that connect disparate protein chains to form continuous 3D hydrogel networks, avoiding complex chemical and/or physical treatments. The Young's modulus of these new BSICT–SF hydrogels can reach up to 6.5 ± 0.2 MPa, tens to hundreds of times higher than that of conventional hydrogels (0.01–0.1 MPa). These new materials fill the “empty soft materials' space” in the elastic modulus/strain Ashby plot. More remarkably, the BSICT–SF hydrogels can be processed into different constructions through different polymer and/or metal‐based processing techniques, such as molding, laser cutting, and machining. Thus, these new hydrogel systems exhibit potential utility in many biomedical and engineering fields.     

Tuning the Microenvironment: Click‐Crosslinked Hyaluronic Acid‐Based Hydrogels Provide a Platform for Studying Breast Cancer Cell Invasion

A big challenge in cell culture is the non‐natural environment in which cells are routinely screened, making in vivo phenomena, such as cell invasion, difficult to understand and predict. To study cancer cell invasion, extracellular matrix (ECM) analogs with decoupled mechanical and chemical properties are required. Hyaluronic acid (HA)‐based hydrogels crosslinked with matrix‐metalloproteinase (MMP)‐cleavable peptides are developed to study MDA‐MB‐231 breast cancer cell invasion. Hydrogels are synthesized by reacting furan‐modified HA with bismaleimide peptide crosslinkers in a Diels–Alder click reaction. This new hydrogel takes advantage of the biomimetic properties of HA, which is overexpressed in breast cancer, and eliminates the use of nonadhesive crosslinkers, such as poly(ethylene glycol) (PEG). The crosslink (mechanical) and ligand (chemical) densities are varied independently to evaluate the effects of each parameter on cell migration. Increased crosslink density correlates with decreased MDA‐MB‐231 cell invasion whereas incorporation of MMP‐cleavable sequences within the peptide crosslinker enhances invasion. Increasing the ligand density of pendant GRGDS groups induces cell proliferation, but has no significant impact on invasion. By independently tuning the mechanical and chemical environment of ECM mimetic hydrogels, a platform is provided that recapitulates variable tissue properties and elucidates the role of the microenvironment in cancer cell invasion.     

Peptide‐Enhanced Selective Surface Deposition of Polymer‐Based Fragrance Delivery Systems

Surface deposition is a critical step in the application of fragrance‐containing products. This contribution presents a novel strategy to enhance the deposition of polymer‐based fragrance delivery systems onto cotton substrates from the application medium using phage display identified peptides. Following the identification of cotton binding peptide ligands under fabric softening conditions via phage display, the strongest binding peptide ligand is incorporated into two model polymer‐based fragrance delivery systems, viz., polymer profragrances and polymer nanoparticles. The model polymer profragrance used is a linear, water soluble poly(N‐(2‐hydroxypropyl)methacrylamide) conjugate, while poly(styrene‐co‐acrylic acid) (PS‐co‐PAA) nanoparticles prepared via miniemulsion polymerization are chosen as the second model system. The incorporation of the cotton binding peptide ligand into these fragrance delivery systems enhances their surface deposition two‐ to three‐fold, as evidenced by fluorescence intensity measurements. In the case of the fragrance‐containing PS‐co‐PAA nanoparticles, the enhanced surface deposition also translates into an increased fragrance release from the cotton surface according to dynamic headspace sampling measurements.     

Photocurable 3D Printing of High Toughness and Self-Healing Hydrogels for Customized Wearable Flexible Sensors

Currently, most customized hydrogels can only be processed via extrusion-based 3D printing techniques, which is limited by printing efficiency and resolution. Here, a simple strategy for the rapid fabrication of customized hydrogels using a photocurable 3D printing technique is presented. This technique has been rarely used because the presence of water increases the molecular distance between the polymer chains and reduces the monomer polymerization rate, resulting in the failure of rapid solid-liquid separation during printing. Although adding cross-linkers to printing inks can effectively accelerate 3D cross-linked network formation, chemical cross-linking may result in reduced toughness and self-healing ability of the hydrogel. Therefore, an interpenetrated-network hydrogel based on non-covalent interactions is designed to form physical cross-links, affording fast solid-liquid separation. Poly(acrylic acid (AA)-N-vinyl-2-pyrrolidone (NVP)) and carboxymethyl cellulose (CMC) are cross-linked via Zn²⁺-ligand coordination and hydrogen bonding; the resulting mixed AA-NVP/CMC solution is used as the printing ink. The printed poly(AA-NVP/CMC) hydrogel exhibited high tensile toughness (3.38 MJ m⁻³) and superior self-healing ability (healed stress: 81%; healed strain: 91%). Some objects like manipulator are successfully customized by photocurable 3D printing using hydrogels with high toughness and complex structures. This high-performance hydrogel has great potential for application in flexible wearable sensors.     

Peptides for the Biofunctionalization of Silicon for Use in Optical Sensing with Porous Silicon Microcavities

Addressing the surface chemistry of silicon is of fundamental scientific and technical significance due to the wide use of this material in electronics and optics. A novel method of functionalizing silicon (Si) via short peptides with binding specificity for Si is presented. The peptide presenting the highest affinity for Si is identified via phage display technology, and the 12‐mer LLADTTHHRPWT and SPGLSLVSHMQT peptides were found to be specific for the n⁺‐Si and p⁺‐Si surfaces, respectively. In our sensing application, the obtained peptides are used as functionalizing linkers to allow porous silicon microcavities to bind biotin and then capture streptavidin. Molecular detection is monitored via reflectometric interference spectra as shifts in the resonance peaks of the cavity structure. An improved streptavidin sensing (21 times lower detection limit) with peptide‐functionalized porous silicon microcavities is demonstrated, compared to sensing performed with devices functionalized with the commonly used silanization method, suggesting that the modification of Si via Si‐specific peptides provides better interface layers for molecular detection. High‐resolution atomic force microscopy images corroborate this result and reveal the formation of ordered nanometer‐sized molecular layers when peptide‐route functionalization is performed.     

A Delayed Cross-Linking Strategy for Evolvable Hydrogels

Biological tissues grow or evolve through a series of complicated processes of matter and energy internalization, which are highly challenging to mimic in synthetic materials. Herein, a delayed cross-linking strategy is developed to program the reactivity of cross-linking sites and make hydrogels evolvable. The polymer networks are constructed by combining polyvinyl alcohol (PVA) with a polyzwitterion comprising both cationic quaternary ammonium and anionic phenylboronic acid groups (PQBA). Shielding of phenylboronic acid groups in ion pairs and polyzwitterion microdomains delays the cross-linking between PVA and PQBA. Mechanical stimulations unlock the phenylboronic acid groups and dramatically accelerate the cross-linking reaction. A simple stretching treatment makes the hydrogels stronger. Training the hydrogels with five cycles of 200% stretching results in up to ≈13.0- and ≈22.8-fold of enhancements in tensile strength and maximum Young's modulus, respectively. The hydrogels can also self-evolve in a damage-healing process, the fracture strength and maximum Young's modulus of the hydrogels increase by at most ≈7.5- and ≈27.2-fold after five times of repeated tensile rupture and self-healing. The study demonstrates the possibility of designing “living polymeric materials” by programming the cross-linking kinetics of polymer networks.     

Transfer Printing of Cell Layers with an Anisotropic Extracellular Matrix Assembly using Cell‐Interactive and Thermosensitive Hydrogels

The structure of tissue plays a critical role in its function and therefore a great deal of attention has been focused on engineering native tissue‐like constructs for tissue engineering applications. Transfer printing of cell layers is a new technology that allows controlled transfer of cell layers cultured on smart substrates with defined shape and size onto tissue‐specific defect sites. Here, the temperature‐responsive swelling‐deswelling of the hydrogels with groove patterns and their versatile and simple use as a template to harvest cell layers with anisotropic extracellular matrix assembly is reported. The hydrogels with a cell‐interactive peptide and anisotropic groove patterns are obtained via enzymatic polymerization. The results show that the cell layer with patterns can be easily transferred to new substrates by lowering the temperature. In addition, multiple cell layers are stacked on the new substrate in a hierarchical manner and the cell layer is easily transplanted onto a subcutaneous region. These results indicate that the evaluated hydrogel can be used as a novel substrate for transfer printing of artificial tissue constructs with controlled structural integrity, which may hold potential to engineer tissue that can closely mimic native tissue architecture.     

Dual Stimuli‐Responsive Supramolecular Polypeptide‐Based Hydrogel and Reverse Micellar Hydrogel Mediated by Host–Guest Chemistry

Versatile strategies are currently being discovered for the fabrication of synthetic polypeptide‐based hybrid hydrogels, which have potential applications in polymer therapeutics and regenerative medicine. Herein, a new concept—the reverse micellar hydrogel—is introduced, and a versatile strategy is provided for fabricating supramolecular polypeptide‐based normal micellar hydrogel and reverse micellar hydrogels from the same polypeptide‐based copolymer via the cooperation of host–guest chemistry and hydrogen‐bonding interactions. The supramolecular hydrogels are thoroughly characterized, and a mechanism for their self‐assembly is proposed. These hydrogels can respond to dual stimuli—temperature and pH—and their mechanical and controlled drug‐release properties can be tuned by the copolymer topology and the polypeptide composition. The reverse micellar hydrogel can load 10% of the anticancer drug doxorubicin hydrochloride (DOX) and sustain DOX release for 45 days, indicating that it could be useful as an injectable drug delivery system.     

Recapitulation of In Situ Endochondral Ossification Using an Injectable Hypoxia-Mimetic Hydrogel

Due to the limited ability for perfusion, traditional intramembranous ossification (IMO) often fails to recapitulate the natural regeneration process of most long bones and craniofacial bones. Alternatively, endochondral ossification (ECO) strategy has emerged and has been evidenced to circumvent the drawbacks in the routine application of IMO. Here, an injectable, poly(glycerol sebacate)-co-poly (ethylene glycol)/polyacrylic acid (PEGS/PAA) hydrogels are successfully developed to induce a hypoxia-mimicking environment and subsequently recapitulate ECO via in situ iron chelation. With the incorporation of PAA, these hydrogels present remarkable viscoelasticity and high efficacy of iron ion-chelating after injection, giving rise to the activation of HIF-1α signaling pathway and suppression of inflammatory responses, and thereby improving chondrogenic differentiation in the early stage and facilitating vascularization in the later stage, which consequently trigger typical ECO. More importantly, through sustained and stable expression of HIF-1α regulated by PEGS/PAA hydrogels throughout the regeneration, a harmonious chondrogenic/osteogenic balance can be struck and thereby accelerating the progress of ECO compared to the PEGS. The findings provide an efficient strategy to achieve in situ ECO via biomaterial-based iron ion-chelating and ensuing hypoxia-mimicking, representing a novel and promising concept for future application in bone regeneration.