Development of a Novel Hierarchically Biofabricated Blood Vessel Mimic Decorated with Three Vascular Cell Populations for the Reconstruction of Small-Diameter Arteries.

The availability of grafts to replace small-diameter arteries remains an unmet clinical need. Here, the validated methodology is reported for a novel hybrid tissue-engineered vascular graft that aims to match the natural structure of small-size arteries. The blood vessel mimic (BVM) comprises an internal conduit of co-electrospun gelatin and polycaprolactone (PCL) nanofibers (corresponding to the tunica intima of an artery), reinforced by an additional layer of PCL aligned fibers (the internal elastic membrane). Endothelial cells are deposited onto the luminal surface using a rotative bioreactor. A bioprinting system extrudes two concentric cell-laden hydrogel layers containing respectively vascular smooth muscle cells and pericytes to create the tunica media and adventitia. The semi-automated cellularization process reduces the production and maturation time to 6 days. After the evaluation of mechanical properties, cellular viability, hemocompatibility, and suturability, the BVM is successfully implanted in the left pulmonary artery of swine. Here, the BVM showed good hemostatic properties, capability to withstand blood pressure, and patency at 5 weeks post-implantation. These promising data open a new avenue to developing an artery-like product for reconstructing small-diameter blood vessels.

Partitioning of an Enzyme-Polymer Surfactant Nanocomplex into Lipid-Rich Cellular Compartments Drives In Situ Hydrolysis of Organophosphates.

Most organophosphates (OPs) are hydrophobic, and after exposure, can sequester into lipophilic regions within the body, such as adipose tissue, resulting in long term chronic effects. Consequently, there is an urgent need for therapeutic agents that can decontaminate OPs in these hydrophobic regions. Accordingly, an enzyme-polymer surfactant nanocomplex is designed and tested comprising chemically supercharged phosphotriesterase (Agrobacterium radiobacter; arPTE) electrostatically conjugated to amphiphilic polymer surfactant chains ([cat.arPTE][S-]). Experimentally-derived structural data are combined with molecular dynamics (MD) simulations to provide atomic level detail on conformational ensembles of the nanocomplex using dielectric constants relevant to aqueous and lipidic microenvironments. These show the formation of a compact admicelle pseudophase surfactant corona under aqueous conditions, which reconfigures to yield an extended conformation at a low dielectric constant, providing insight into the mechanism underpinning cell membrane binding. Significantly, it demonstrated that [cat.arPTE][S-] spontaneously binds to human mesenchymal stem cell membranes (hMSCs), resulting in on-cell OP hydrolysis. Moreover, the nanoconstruct can endocytose and partition into the intracellular fatty vacuoles of adipocytes and hydrolyze sequestered OP.

Correction: Flax fibre reinforced alginate poloxamer hydrogel: assessment of mechanical and 4D printing potential.

Correction for 'Flax fibre reinforced alginate poloxamer hydrogel: assessment of mechanical and 4D printing potential' by Charles de Kergariou et al., Soft Matter, 2024, 20, 4021-4034, https://doi.org/10.1039/D4SM00135D.

Flax fibre reinforced alginate poloxamer hydrogel: assessment of mechanical and 4D printing potential.

The mechanical and printing performance of a new biomaterial, flax fibre-reinforced alginate-poloxamer based hydrogel, for load-bearing and 4D printing biomedical applications is described in this study. The-self suspendable ability of the material was evaluated by optimising the printing parameters and conducting a collapse test. 1% of the flax fibre weight fraction was sufficient to obtain an optimum hydrogel composite from a mechanical perspective. The collapse test showed that the addition of flax fibres allowed a consistent print without support over longer distances (8 and 10 mm) than the unreinforced hydrogel. The addition of 1% of flax fibres increased the viscosity by 39% and 129% at strain rates of 1 rad s-1 and 5 rad s-1, respectively, compared to the unreinforced hydrogel. The distributions of fibre size and orientation inside the material were also evaluated to identify the internal morphology of the material. The difference of coefficients of moisture expansion between the printing direction (1.29 × 10-1) and the transverse direction (6.03 × 10-1) showed potential for hygromorphic actuation in 4D printing. The actuation authority was demonstrated by printing a [0°; 90°] stacking sequence and rosette-like structures, which were then actuated using humidity gradients. Adding fibres to the hydrogel improved the repeatability of the actuation, while lowering the actuation authority from 0.11 mm-1 to 0.08 mm-1. Overall, this study highlighted the structural and actuation-related benefits of adding flax fibres to hydrogels.

Modular Bioorthogonal Lipid Nanoparticle Modification Platforms for Cardiac Homing.

Lipid nanoparticles (LNPs) are becoming widely adopted as vectors for the delivery of therapeutic payloads but generally lack intrinsic tissue-homing properties. These extracellular vesicle (EV) mimetics can be targeted toward the liver, lung, or spleen via charge modification of their lipid headgroups. Homing to other tissues has only been achieved via covalent surface modification strategies using small-molecule ligands, peptides, or monoclonal antibodies─methods that are challenging to couple with large-scale manufacturing. Herein, we design a novel modular artificial membrane-binding protein (AMBP) platform for the modification of LNPs postformation. The system is composed of two protein modules that can be readily coupled using bioorthogonal chemistry to yield the AMBP. The first is a membrane anchor module comprising a supercharged green fluorescent protein (scGFP) electrostatically conjugated to a dynamic polymer surfactant corona. The second is a functional module containing a cardiac tissue fibronectin homing sequence from the bacterial adhesin CshA. We demonstrate that LNPs modified using the AMBP exhibit a 20-fold increase in uptake by fibronectin-rich C2C12 cells under static conditions and a 10-fold increase under physiologically relevant shear stresses, with no loss of cell viability. Moreover, we show targeted localization of the AMBP-modified LNPs in zebrafish hearts, highlighting their therapeutic potential as a vector for the treatment of cardiac disease and, more generally, as a smart vector.

Diffusivelike Motions in a Solvent-Free Protein-Polymer Hybrid.

The interaction between proteins and hydration water stabilizes protein structure and promotes functional dynamics, with water translational motions enabling protein flexibility. Engineered solvent-free protein-polymer hybrids have been shown to preserve protein structure, function, and dynamics. Here, we used neutron scattering, protein and polymer perdeuteration, and molecular dynamics simulations to explore how a polymer dynamically replaces water. Even though relaxation rates and vibrational properties are strongly modified in polymer coated compared to hydrated proteins, liquidlike polymer dynamics appear to plasticize the conjugated protein in a qualitatively similar way as do hydration-water translational motions.

Controlling Protein Nanocage Assembly with Hydrostatic Pressure.

Controlling the assembly and disassembly of nanoscale protein cages for the capture and internalization of protein or non-proteinaceous components is fundamentally important to a diverse range of bionanotechnological applications. Here, we study the reversible, pressure-induced dissociation of a natural protein nanocage, E. coli bacterioferritin (Bfr), using synchrotron radiation small-angle X-ray scattering (SAXS) and circular dichroism (CD). We demonstrate that hydrostatic pressures of 450 MPa are sufficient to completely dissociate the Bfr 24-mer into protein dimers, and the reversibility and kinetics of the reassembly process can be controlled by selecting appropriate buffer conditions. We also demonstrate that the heme B prosthetic group present at the subunit dimer interface influences the stability and pressure lability of the cage, despite its location being discrete from the interdimer interface that is key to cage assembly. This indicates a major cage-stabilizing role for heme within this family of ferritins.

Insight into the Structure and Activity of Surface-Engineered Lipase Biofluids.

Despite a successful application of solvent-free liquid protein (biofluids) concept to a number of commercial enzymes, the technical advantages of enzyme biofluids as hyperthermal stable biocatalysts cannot be fully utilized as up to 90-99% of native activities are lost when enzymes were made into biofluids. With a two-step strategy (site-directed mutagenesis and synthesis of variant biofluids) on Bacillus subtilis lipase A (BsLA), we elucidated a strong dependency of structure and activity on the number and distribution of polymer surfactant binding sites on BsLA surface. Here, it is demonstrated that improved BsLA variants can be engineered via site-mutagenesis by a rational design, either with enhanced activity in aqueous solution in native form, or with improved physical property and increased activity in solvent-free system in the form of a protein liquid. This work answered some fundamental questions about the surface characteristics for construction of biofluids, useful for identifying new strategies for developing advantageous biocatalysts.

Sequential Electrostatic Assembly of a Polymer Surfactant Corona Increases Activity of the Phosphotriesterase arPTE.

We present a new methodology for the generation of discrete molecularly dispersed enzyme-polymer-surfactant bioconjugates. Significantly, we demonstrate that >3-fold increase in the catalytic efficiency of the diffusion-limited phosphotriesterase arPTE can be achieved through sequential electrostatic addition of cationic and anionic polymer surfactants, respectively. Here, the polymer surfactants assemble on the surface of the enzyme via ion exchange to yield a compact corona. The observed rate enhancement is consistent with a mechanism whereby the polymer-surfactant corona gives rise to a decrease in the dielectric constant in the vicinity of the active site of the enzyme, accelerating the rate-determining product diffusion step. The facile methodology has significant potential for increasing the efficiency of enzymes and could therefore have a substantially positive impact for industrial enzymology.

Biodegradable, Drug-Loaded Nanovectors via Direct Hydration as a New Platform for Cancer Therapeutics.

The stabilization and transport of low-solubility drugs, by encapsulation in nanoscopic delivery vectors (nanovectors), is a key paradigm in nanomedicine. However, the problems of carrier toxicity, specificity, and producibility create a bottleneck in the development of new nanomedical technologies. Copolymeric nanoparticles are an excellent platform for nanovector engineering due to their structural versatility; however, conventional fabrication processes rely upon harmful chemicals that necessitate purification. In engineering a more robust (copolymeric) nanovector platform, it is necessary to reconsider the entire process from copolymer synthesis through self-assembly and functionalization. To this end, a process is developed whereby biodegradable copolymers of poly(ethylene glycol)-block-poly(trimethylene carbonate), synthesized via organocatalyzed ring-opening polymerization, undergo assembly into highly uniform, drug-loaded micelles without the use of harmful solvents or the need for purification. The direct hydration methodology, employing oligo(ethylene glycol) as a nontoxic dispersant, facilitates rapid preparation of pristine, drug-loaded nanovectors that require no further processing. This method is robust, fast, and scalable. Utilizing parthenolide, an exciting candidate for treatment of acute lymphoblastic leukemia (ALL), discrete nanovectors are generated that show strikingly low carrier toxicity and high levels of specific therapeutic efficacy against primary ALL cells (as compared to normal hematopoietic cells).

Functionalized Triblock Copolymer Vectors for the Treatment of Acute Lymphoblastic Leukemia.

The chemotherapeutic Parthenolide is an exciting new candidate for the treatment of acute lymphoblastic leukemia, but like many other small-molecule drugs, it has low aqueous solubility. As a consequence, Parthenolide can only be administered clinically in the presence of harmful cosolvents. Accordingly, we describe the synthesis, characterization, and testing of a range of biocompatible triblock copolymer micelles as particle-based delivery vectors for the hydrophobic drug Parthenolide. The drug-loaded particles are produced via an emulsion-to-micelle transition method, and the effects of introducing anionic and cationic surface charges on stability, drug sequestration, biocompatibility, and efficacy are investigated. Significantly, we demonstrate high levels of efficacy in the organic solvent-free systems against human mesenchymal stem cells and primary T-acute lymphoblastic leukemia patient cells, highlighting the effectiveness of the delivery vectors for the treatment of acute lymphoblastic leukemia.

Effect of Bioconjugation on the Reduction Potential of Heme Proteins.

The modification of protein surfaces employing cationic and anionic species enables the assembly of these biomaterials into highly sophisticated hierarchical structures. Such modifications can allow bioconjugates to retain or amplify their functionalities under conditions in which their native structure would be severely compromised. In this work, we assess the effect of this type of bioconjugation on the redox properties of two model heme proteins, that is, cytochrome c (CytC) and myoglobin (Mb). In particular, the work focuses on the sequential modification by 3-dimethylamino propylamine (DMAPA) and 4-nonylphenyl 3-sulfopropyl ether (S1) anionic surfactant. Bioconjugation with DMAPA and S1 are the initial steps in the generation of pure liquid proteins, which remain active in the absence of water and up to temperatures above 150 °C. Thin-layer spectroelectrochemistry reveals that DMAPA cationization leads to a distribution of bioconjugate structures featuring reduction potentials shifted up to 380 mV more negative than the native proteins. Analysis based on circular dichroism, MALDI-TOF mass spectrometry, and zeta potential measurements suggest that the shift in the reduction potentials are not linked to protein denaturation, but to changes in the spin state of the heme. These alterations of the spin states originate from subtle structural changes induced by DMAPA attachment. Interestingly, electrostatic coupling of anionic surfactant S1 shifts the reduction potential closer to that of the native protein, demonstrating that the modifications of the heme electronic configuration are linked to surface charges.

Molecular dynamics simulations reveal a dielectric-responsive coronal structure in protein-polymer surfactant hybrid nanoconstructs.

Solvent-free liquid proteins are a new class of thermally stable hybrid bionanomaterials that are produced by extensive lyophilization of aqueous solutions of protein-polymer surfactant nanoconjugates followed by thermal annealing. The hybrid constructs, which consist of a globular protein core surrounded by a monolayer of electrostatically coupled polymer surfactant molecules, exhibit nativelike structure, function, and backbone dynamics over a large temperature range. Despite the key importance of the polymer surfactant shell, very little is known about the atomistic structure of the corona and how it influences the phase behavior and properties of these novel nanoscale objects. Here we present molecular dynamics simulations of protein-polymer surfactant nanoconjugates consisting of globular cores of myoglobin or lysozyme and demonstrate that the derived structural parameters are highly consistent with experimental values. We show that the coronal layer structure is responsive to the dielectric constant of the medium and that the mobility of the polymer surfactant molecules is significantly hindered in the solvent-free state, providing a basis for the origins of retained protein dynamics in these novel biofluids. Taken together, our results suggest that the extension of molecular dynamics simulations to hybrid nanoscale objects could be of generic value in diverse areas of soft matter chemistry, bioinspired engineering, and biomolecular nanotechnology.

Hydrogel-Based Artificial Synapses for Sustainable Neuromorphic Electronics.

Hydrogels find widespread applications in biomedicine because of their outstanding biocompatibility, biodegradability, and tunable material properties. Hydrogels can be chemically functionalized or reinforced to respond to physical or chemical stimulation, which opens up new possibilities in the emerging field of intelligent bioelectronics. Here, the state-of-the-art in functional hydrogel-based transistors and memristors is reviewed as potential artificial synapses. Within these systems, hydrogels can serve as semisolid dielectric electrolytes in transistors and as switching layers in memristors. These synaptic devices with volatile and non-volatile resistive switching show good adaptability to external stimuli for short-term and long-term synaptic memory effects, some of which are integrated into synaptic arrays as artificial neurons; although, there are discrepancies in switching performance and efficacy. By comparing different hydrogels and their respective properties, an outlook is provided on a new range of biocompatible, environment-friendly, and sustainable neuromorphic hardware. How potential energy-efficient information storage and processing can be achieved using artificial neural networks with brain-inspired architecture for neuromorphic computing is described. The development of hydrogel-based artificial synapses can significantly impact the fields of neuromorphic bionics, biometrics, and biosensing.

A 3D In-vitro model of the human dentine interface shows long-range osteoinduction from the dentine surface.

Emerging regenerative cell therapies for alveolar bone loss have begun to explore the use of cell laden hydrogels for minimally invasive surgery to treat small and spatially complex maxilla-oral defects. However, the oral cavity presents a unique and challenging environment for in vivo bone tissue engineering, exhibiting both hard and soft periodontal tissue as well as acting as key biocenosis for many distinct microbial communities that interact with both the external environment and internal body systems, which will impact on cell fate and subsequent treatment efficacy. Herein, we design and bioprint a facile 3D in vitro model of a human dentine interface to probe the effect of the dentine surface on human mesenchymal stem cells (hMSCs) encapsulated in a microporous hydrogel bioink. We demonstrate that the dentine substrate induces osteogenic differentiation of encapsulated hMSCs, and that both dentine and ß-tricalcium phosphate substrates stimulate extracellular matrix production and maturation at the gel-media interface, which is distal to the gel-substrate interface. Our findings demonstrate the potential for long-range effects on stem cells by mineralized surfaces during bone tissue engineering and provide a framework for the rapid development of 3D dentine-bone interface models.

Generalisation of the yield stress measurement in three point bending collapse tests: application to 3D printed flax fibre reinforced hydrogels.

This paper describes the extrusion pressure's effect on composite hydrogel inks' filaments subjected to three point bending collapse tests. The composite considered in this work consists of an alginate-poloxamer hydrogel reinforced with flax fibres. Increased extrusion pressure resulted in more asymmetrical filaments between the support pillars. Furthermore, the material and printing conditions used in the present study led to the production of curved specimens. These two characteristics implicitly limit the validity of the yield stress equations commonly used in open literature. Therefore, a new system of equations was derived for the case of asymmetrical and curved filaments. A post-processing method was also created to obtain the properties required to evaluate this yield stress. This new equation was then implemented to identify the strength of failed hydrogels without flax fibre reinforcement. A statistical analysis showed this new equation's significance, which yielded statistically higher (i.e. 1.15 times larger) strength values compared to the numbers obtained with the open literature equations. At larger extrusion pressures, longer periods were needed for the material to converge towards its final shape. Larger extrusion pressure values led to lower yield stresses within the composite hydrogel filament: a 5 kPa increase in extrusion pressure lowered the yield stress by 19%. In comparison, a 15 kPa increase led to a 29% decrease in the yield stress. Overall this study provides guidelines to standardize three point bending collapse tests and analysis comparison between different materials.

Tissue adhesives based on chitosan for biomedical applications.

Chitosan bio-adhesives bond strongly with various biological tissues, such as skin, mucosa, and internal organs. Their adhesive ability arises from amino acid and hydroxyl groups in chitosan, facilitating interactions with tissue surfaces through chemical (ionic, covalent, and hydrogen) and physical (chain entanglement) bonding. As non-toxic, biodegradable, and biocompatible materials, chitosan bio-adhesives are a safe option for medical therapies. They are particularly suitable for drug delivery, wound healing, and tissue regeneration. In this review, we address chitosan-based bio-adhesives and the mechanisms associated with them. We also discuss different chitosan composite-based bio-adhesives and their biomedical applications in wound healing, drug delivery, hemostasis, and tissue regeneration. Finally, challenges and future perspectives for the clinical use of chitosan-based bio-adhesives are discussed.

Protocol to decellularize porcine right ventricular outflow tracts using a 3D printed flow chamber.

Surgical treatment of pediatric congenital heart disease with tissue grafts is a lifesaving intervention. Decellularization to reduce immunogenicity of tissue grafts is an increasingly popular alternative to glutaraldehyde fixation. Here, we present a protocol to decellularize porcine right ventricular outflow tracts using a 3D printed flow chamber. We describe steps for 3D printing the flow rig, preparing porcine tissue, and using the flow rig to utilize shear forces for decellularization. We then detail procedures for characterizing the acellular scaffold. For complete details on the use and execution of this protocol, please refer to Vafaee et al.1.

Oxidative stress modulating nanomaterials and their biochemical roles in nanomedicine.

Many pathological conditions are predominantly associated with oxidative stress, arising from reactive oxygen species (ROS); therefore, the modulation of redox activities has been a key strategy to restore normal tissue functions. Current approaches involve establishing a favorable cellular redox environment through the administration of therapeutic drugs and redox-active nanomaterials (RANs). In particular, RANs not only provide a stable and reliable means of therapeutic delivery but also possess the capacity to finely tune various interconnected components, including radicals, enzymes, proteins, transcription factors, and metabolites. Here, we discuss the roles that engineered RANs play in a spectrum of pathological conditions, such as cancer, neurodegenerative diseases, infections, and inflammation. We visualize the dual functions of RANs as both generator and scavenger of ROS, emphasizing their profound impact on diverse cellular functions. The focus of this review is solely on inorganic redox-active nanomaterials (inorganic RANs). Additionally, we deliberate on the challenges associated with current RANs-based approaches and propose potential research directions for their future clinical translation.

Redox transitions in an electrolyte-free myoglobin fluid.

Redox responses associated with the heme prosthetic group in a myoglobin-polymer surfactant solvent-free liquid are investigated for the first time in the absence of an electrolyte solution. Cyclic voltammograms from the biofluid exhibit responses that are consistent with planar diffusion of mobile charges in the melt. Temperature-dependent dynamic electrochemical and rheological responses are rationalized in terms of the effective electron hopping rate between heme centers and the transport of intrinsic ionic species in the viscous protein liquid.