Recent advances in extracellular matrix-inspired nanocarriers.

Introduction: ECM-inspired nanocarriers have emerged as a promising platform for drug delivery due to their unique advantages over traditional nanocarrier systems. The ECM is a complex three-dimensional network comprising proteins and polysaccharides that play a key role in maintaining tissue function and homeostasis. Recent advances in the design and synthesis of ECM-inspired nanocarriers have resulted in superior efficacy, targeting, and responsive delivery systems.

Toward understanding biomolecular materials comprising intrinsically disordered proteins simulation and experiment.

Intrinsically disordered proteins (IDPs) yield solutions with tunable phase transition behavior and have been widely applied in designing stimuli-responsive materials. Understanding interactions between amino acid residues of the IDP sequence is critical to designing new IDP-based materials with selective phase behavior, assembly, and mechanical properties. The lack of defined structure for this class of proteins complicates accurate prediction of their molecular-scale behavior.

Architectural control of rod-coil block polypeptide thermoresponsive self-assembly design of coiled-coil orientation.

The architectural control of the self-assembly of a series of block polypeptides comprising a concatenation of an elastin-like peptide and a coiled-coil, bundle-forming peptide (ELP-BFPs), has been demonstrated. Assembly of the polypeptides is controlled by coacervation of the hydrophobic ELP domain, while the type of coiled-coil assembly of the BFP and the specific placement of short histidine tags significantly tunes assembly behavior. Spectrophotometric analysis of self-assembly demonstrated that the transition temperature of assembly can be controlled by the design of the BFP domain and positioning of the His-tags in the constructs.

Growth factors and growth factor gene therapies for treating chronic wounds.

Chronic wounds are an unmet clinical need affecting millions of patients globally, and current standards of care fail to consistently promote complete wound closure and prevent recurrence. Disruptions in growth factor signaling, a hallmark of chronic wounds, have led researchers to pursue growth factor therapies as potential supplements to standards of care. Initial studies delivering growth factors in protein form showed promise, with a few formulations reaching clinical trials and one obtaining clinical approval.

Expanding the molecular language of protein liquid-liquid phase separation.

Understanding the relationship between a polypeptide sequence and its phase separation has important implications for analysing cellular function, treating disease and designing novel biomaterials. Several sequence features have been identified as drivers for protein liquid-liquid phase separation (LLPS), schematized as a 'molecular grammar' for LLPS. Here we further probe how sequence modulates phase separation and the material properties of the resulting condensates, targeting sequence features previously overlooked in the literature.

Controlled Release of Drugs from Extracellular Matrix-Derived Peptide-Based Nanovesicles through Tailored Noncovalent Interactions.

Elastin-collagen nanovesicles (ECnV) have emerged as a promising platform for drug delivery due to their tunable physicochemical properties and biocompatibility. The potential of nine distinct ECnVs to serve as drug-delivery vehicles was investigated in this study, and it was demonstrated that various small-molecule cargo (e.g.

Genetically Fusing Order-Promoting and Thermoresponsive Building Blocks to Design Hybrid Biomaterials.

The unique biophysical and biochemical properties of intrinsically disordered proteins (IDPs) and their recombinant derivatives, intrinsically disordered protein polymers (IDPPs) offer opportunities for producing multistimuli-responsive materials; their sequence-encoded disorder and tendency for phase separation facilitate the development of multifunctional materials. This review highlights the strategies for enhancing the structural diversity of elastin-like polypeptides (ELPs) and resilin-like polypeptides (RLPs), and their self-assembled structures via genetic fusion to ordered motifs such as helical or beta sheet domains. In particular, this review describes approaches that harness the synergistic interplay between order-promoting and thermoresponsive building blocks to design hybrid biomaterials, resulting in well-structured, stimuli-responsive supramolecular materials ordered on the nanoscale.

Genetically Fused Resilin-like Polypeptide-Coiled Coil Bundlemer Conjugates Exhibit Tunable Multistimuli-Responsiveness and Undergo Nanofibrillar Assembly.

Peptide-based materials are diverse candidates for self-assembly into modularly designed and stimuli-responsive nanostructures with precisely tunable compositions. Here, we genetically fused computationally designed coiled coil-forming peptides to the N- and C-termini of compositionally distinct multistimuli-responsive resilin-like polypeptides (RLPs) of various lengths. The successful expression of these hybrid polypeptides in bacterial hosts was confirmed through techniques such as gel electrophoresis, mass spectrometry, and amino acid analysis.

Injectable liposome-containing click hydrogel microparticles for release of macromolecular cargos.

Hydrogel microparticles ranging from 0.1-100 μm, referred to as microgels, are attractive for biological applications afforded by their injectability and modularity, which allows facile delivery of mixed populations for tailored combinations of therapeutics. Significant efforts have been made to broaden methods for microgel production including the materials and chemistries by which they are made.

Sequence-Encoded Differences in Phase Separation Enable Formation of Resilin-like Polypeptide-Based Microstructured Hydrogels.

Microstructured hydrogels are promising platforms to mimic structural and compositional heterogeneities of the native extracellular matrix (ECM). The current state-of-the-art soft matter patterning techniques for generating ECM mimics can be limited owing to their reliance on specialized equipment and multiple time- and energy-intensive steps. Here, a photocross-linking methodology that traps various morphologies of phase-separated multicomponent formulations of compositionally distinct resilin-like polypeptides (RLPs) is reported.

Genetic Fusion of Thermoresponsive Polypeptides with UCST-type Behavior Mediates 1D Assembly of Coiled-Coil Bundlemers.

Thermoresponsive resilin-like polypeptides (RLPs) of various lengths were genetically fused to two different computationally designed coiled coil-forming peptides with distinct thermal stability, to develop new strategies to assemble coiled coil peptides via temperature-triggered phase separation of the RLP units. Their successful production in bacterial expression hosts was verified via gel electrophoresis, mass spectrometry, and amino acid analysis. Circular dichroism (CD) spectroscopy, ultraviolet-visible (UV/Vis) turbidimetry, and dynamic light scattering (DLS) measurements confirmed the stability of the coiled coils and showed that the thermosensitive phase behavior of the RLPs was preserved in the genetically fused hybrid polypeptides.

VEGF-Encoding, Gene-Activated Collagen-Based Matrices Promote Blood Vessel Formation and Improved Wound Repair.

Disruption in vascularization during wound repair can severely impair healing. Proangiogenic growth factor therapies have shown great healing potential; however, controlling growth factor activity and cellular behavior over desired healing time scales remains challenging. In this study, we evaluated collagen-mimetic peptide (CMP) tethers for their capacity to control growth factor gene transfer and growth factor activity using our recently developed gene-activated hyaluronic acid-collagen matrix (GAHCM).

Controlled Delivery of Vancomycin from Collagen-tethered Peptide Vehicles for the Treatment of Wound Infections.

Despite the great promise of antibiotic therapy in wound infections, antibiotic resistance stemming from frequent dosing diminishes drug efficacy and contributes to recurrent infection. To identify improvements in antibiotic therapies, new antibiotic delivery systems that maximize pharmacological activity and minimize side effects are needed. In this study, we developed elastin-like peptide and collagen-like peptide nanovesicles (ECnVs) tethered to collagen-containing matrices to control vancomycin delivery and provide extended antibacterial effects against methicillin-resistant (MRSA).

Recombinant protein-based injectable materials for biomedical applications.

Injectable nanocarriers and hydrogels have found widespread use in a variety of biomedical applications such as local and sustained biotherapeutic cargo delivery, and as cell-instructive matrices for tissue engineering. Recent advances in the development and application of recombinant protein-based materials as injectable platforms under physiological conditions have made them useful platforms for the development of nanoparticles and tissue engineering matrices, which are reviewed in this work. Protein-engineered biomaterials are highly customizable, and they provide distinctly tunable rheological properties, encapsulation efficiencies, and delivery profiles.

Modified hyaluronic acid-collagen matrices trigger efficient gene transfer and prohealing behavior in fibroblasts for improved wound repair.

Growth factor therapy has demonstrated great promise for chronic wound repair, but controlling growth factor activity and cell phenotype over desired time frames remains a critical challenge. In this study, we developed a gene-activated hyaluronic acid-collagen matrix (GAHCM) comprising DNA/polyethylenimine (PEI) polyplexes retained on hyaluronic acid (HA)-collagen hydrogels using collagen mimetic peptides (CMPs). We hypothesized that manipulating both the number of CMP-collagen tethers and the ECM composition would provide a powerful strategy to control growth factor gene transfer kinetics while regulating cell behavior, resulting in enhanced growth factor activity for wound repair.

Microgels Formed by Spontaneous Click Chemistries Utilizing Microfluidic Flow Focusing for Cargo Release in Response to Endogenous or Exogenous Stimuli.

Protein therapeutics have become increasingly popular for the treatment of a variety of diseases owing to their specificity to targets of interest. However, challenges associated with them have limited their use for a range of ailments, including the limited options available for local controlled delivery. To address this challenge, degradable hydrogel microparticles, or microgels, loaded with model biocargoes were created with tunable release profiles or triggered burst release using chemistries responsive to endogenous or exogeneous stimuli, respectively.

Therapeutic nanocarriers comprising extracellular matrix-inspired peptides and polysaccharides.

Introduction: The extracellular matrix (ECM) is vital for cell and tissue development. Given its importance, extensive work has been conducted to develop biomaterials and drug delivery vehicles that capture features of ECM structure and function.

Areas Covered: This review highlights recent developments of ECM-inspired nanocarriers and their exploration for drug and gene delivery applications.

A 3-D hydrogel based system for hematopoietic differentiation and its use in modeling down syndrome associated transient myeloproliferative disorder.

Induced pluripotent stem cells (iPSCs) provide an extraordinary tool for disease modeling owing to their potential to differentiate into the desired cell type. The differentiation of iPSCs is typically performed on 2-dimensional monolayers of stromal cell or animal tissue derived extracellular matrices. Recent advancements in disease modeling have utilized iPSCs in 3-dimensional (3D) cultures to study diseases such as muscular dystrophy, cardiomyopathy, and pulmonary fibrosis.

Application of Thermoresponsive Intrinsically Disordered Protein Polymers in Nanostructured and Microstructured Materials.

Modulation of inter- and intramolecular interactions between bioinspired designer molecules can be harnessed for developing functional structures that mimic the complex hierarchical organization of multicomponent assemblies observed in nature. Furthermore, such multistimuli-responsive molecules offer orthogonal tunability for generating versatile multifunctional platforms via independent biochemical and biophysical cues. In this review, the remarkable physicochemical and mechanical properties of genetically engineered protein polymers derived from intrinsically disordered proteins, specifically elastin and resilin, are discussed.

Placement of Tyrosine Residues as a Design Element for Tuning the Phase Transition of Elastin-peptide-containing Conjugates: Experiments and Simulations.

Elastin-like polypeptides (ELP) have been widely used in the biomaterials community due to their controllable, thermoresponsive properties and biocompatibility. Motivated by our previous work on the effect of tryptophan (W) substitutions on the LCST-like transitions of short ELPs, we studied a series of short ELPs containing tyrosine (Y) and/or phenylalanine (F) guest residues with only 5 or 6 pentapeptide repeat units. A combination of experiments and molecular dynamics (MD) simulations illustrated that the substitution of F with Y guest residues impacted the transition temperature (T) of short ELPs when conjugated to collagen-like-peptides (CLP), with a reduction in the transition temperature observed only after substitution of at least two residues.

Alteration of Microstructure in Biopolymeric Hydrogels Compositional Modification of Resilin-Like Polypeptides.

Heterogeneities in hydrogel scaffolds are known to impact the performance of cells in cell-laden materials constructs, and we have employed the phase separation of resilin-like polypeptides (RLPs) as a means to generate such materials. Here, we study the compositional features of resilin-like polypeptides (RLPs) that further enable our control of their liquid-liquid phase separation (LLPS) and how such control impacts the formation of microstructured hydrogels. The evaluation of the phase separation of RLPs in solutions of ammonium sulfate offers insights into the sequence-dependent LLPS of the RLP solutions, and atomistic simulations, along with 2D-nuclear Overhauser effect spectroscopy (NOESY) and correlated spectroscopy (COSY) H NMR, suggest specific amino acid interactions that may mediate this phase behavior.

Regulation of neovasculogenesis in co-cultures of aortic adventitial fibroblasts and microvascular endothelial cells by cell-cell interactions and TGF-β/ALK5 signaling.

Adventitial fibroblasts (AFs) are critical mediators of vascular remodeling. However, the contributions of AFs towards development of vasculature and the specific mechanisms by which these cells regulate physiological expansion of the vasa vasorum, the specialized microvasculature that supplies nutrients to the vascular wall, are not well understood. To determine the regulatory role of AFs in microvascular endothelial cell (MVEC) neovasculogenesis and to investigate the regulatory pathways utilized for communication between the two cell types, AFs and MVECs were cultured together in poly(ethylene glycol)-based hydrogels.

Substrate stiffness directs the phenotype and polarization state of cord blood derived macrophages.

Cord blood (CB) mononuclear cell populations have demonstrated significant promise in biomaterials-based regenerative therapies; however, the contributions of monocyte and macrophage subpopulations towards proper tissue healing and regeneration are not well understood, and the phenotypic responses of macrophage to microenvironmental cues have not been well-studied. In this work, we evaluated the effects of cytokine stimulation and altered substrate stiffness. Macrophage derived from CB CD14 monocytes adopted distinct inflammatory (M1) and anti-inflammatory (M2a and M2c) phenotypes in response to cytokine stimulation (M1: lipopolysaccharide (LPS) and interferon (IFN-γ); M2a: interleukin (IL)-4 and IL-13; M2c: IL-10) as determined through expression of relevant cell surface markers and growth factors.

Multi-stimuli-responsive, liposome-crosslinked poly(ethylene glycol) hydrogels for drug delivery.

The development of hybrid hydrogels has been of great interest over recent decades, especially in the field of biomaterials. Such hydrogels provide various opportunities in tissue engineering, drug delivery, and regenerative medicine due to their ability to mimic cellular environments, sequester and release therapeutic agents, and respond to stimuli. Herein we report the synthesis and characterization of an injectable poly(ethylene glycol) hydrogel crosslinked thiol-maleimide reactions and containing both chemically crosslinked temperature-sensitive liposomes (TSLs) and matrix metalloproteinase-sensitive peptide crosslinks.