3D Printable Soy/Silk Hybrid Hydrogels for Tissue Engineering Applications.

The development of protein-based 3D printable hydrogel systems with tunable structure and properties is a critical challenge in contemporary biomedicine. Particularly, 3D printing of modular hydrogels comprising different types of protein tertiary structure, such as globular and fibrous, has not yet been achieved. Here we report the extrusion-based 3D printing of hybrid hydrogels photochemically co-cross-linked between globular soy protein isolate (SPI) and fibrous silk fibroin (SF) for the first time. The hierarchical structure and organization of pristine SPI and SF, and 1:3 (SPI/SF) hybrid inks under various shear stress were investigated using in situ rheology combined with small-/ultra-small-angle neutron scattering (Rheo-SANS/USANS). The hybrid ink exhibited an isotropic mass fractal structure that was stable between tested shear rates of 0.1 and 100 s-1 (near printing shear). The kinetics of sol-gel transition during the photo-cross-linking reaction and the micromechanical properties of fabricated hydrogels were investigated using photorheology and atomic force microscopy, where the hybrid hydrogels exhibited tunable storage and Young's moduli in the range of 13-29 and 214-811 kPa, respectively. The cross-link density and printing accuracy of hybrid hydrogels and inks were observed to increase with the increase in SF content. The 3D printed hybrid hydrogels exhibited a micropore size larger than that of solution casted hydrogels; where the 3D printed 1:3 (SPI/SF) hybrid hydrogel showed a pore size about 7.6 times higher than that of the casted hydrogel. Moreover, the fabricated hybrid hydrogels exhibit good mouse fibroblast cell attachment, viability, and proliferation, demonstrating their potential for tissue engineering applications.

Tunable Thermoresponsiveness of Resilin via Coassembly with Rigid Biopolymers.

The ability to tune the thermoresponsiveness of recombinant resilin protein, Rec1-resilin, through a facile coassembly system was investigated in this study. The effects of change in conformation and morphology with time and the responsive behavior of Rec1-resilin in solution were studied in response to the addition of a rigid model polypeptide (poly-l-proline) or a hydrophobic rigid protein (Bombyx mori silk fibroin). It was observed that by inducing more ordered conformations and increasing the hydrophobicity the lower critical solution temperature (LCST) of the system was tuned to lower values. Time and temperature were found to be critical parameters in controlling the coassembly behavior of Rec1-resilin in both the model polypeptide and more complex protein systems. Such unique properties are useful for a wide range of applications, including drug delivery and soft tissue engineering applications.

Engineering interaction between bone marrow derived endothelial cells and electrospun surfaces for artificial vascular graft applications.

The aim of this investigation was to understand and engineer the interactions between endothelial cells and the electrospun (ES) polyvinylidene fluoride-co-hexafluoropropylene (PVDF-HFP) nanofiber surfaces and evaluate their potential for endothelialization. Elastomeric PVDF-HFP samples were electrospun to evaluate their potential use as small diameter artificial vascular graft scaffold (SDAVG) and compared with solvent cast (SC) PVDF-HFP films. We examined the consequences of fibrinogen adsorption onto the ES and SC samples for endothelialisation. Bone marrow derived endothelial cells (BMEC) of human origin were incubated with the test and control samples and their attachment, proliferation, and viability were examined. The nature of interaction of fibrinogen with SC and ES samples was investigated in detail using ELISA, XPS, and FTIR techniques. The pristine SC and ES PVDF-HFP samples displayed hydrophobic and ultrahydrophobic behavior and accordingly, exhibited minimal BMEC growth. Fibrinogen adsorbed SC samples did not significantly enhance endothelial cell binding or proliferation. In contrast, the fibrinogen adsorbed electrospun surfaces showed a clear ability to modulate endothelial cell behavior. This system also represents an ideal model system that enables us to understand the natural interaction between cells and their extracellular environment. The research reported shows potential of ES surfaces for artificial vascular graft applications.

Interaction of platelets with poly(vinylidene fluoride-co-hexafluoropropylene) electrospun surfaces.

Platelets are the major contributors in the process of thrombosis and in the failure of biomedical implants. A number of factors influence the platelet interaction with foreign surfaces such as surface morphology, surface chemistry, and adsorbed proteins. This study examined the effect of surface topography and chemistry of pristine and fibrinogen-adsorbed solvent cast (SC) and electrospun (ES) samples of poly(vinylidene fluoride-co-hexafluoropropylene) (PVDF-HFP) on platelet adhesion, activation, and aggregation. Qualitative and quantitative studies of fibrinogen adsorption were performed using time-of-flight secondary ion mass spectrometry (ToF-SIMS), while SEM, aggregometry, and liquid scintillation analyses were performed to evaluate platelet adhesion, aggregation, and serotonin release. While little or no platelet adhesion was observed on pristine ES surfaces, considerable adhesion, and measurable aggregation and serotonin release were observed on pristine SC surfaces. Notably, increased adhesion of platelets was observed following fibrinogen adsorption on SC surface with considerable aggregation and serotonin release compared with ES samples, where limited aggregation and platelet adhesion was observed. A further comparison of platelet adhesion, aggregation, and serotonin release was performed with plasma-adsorbed SC and ES surfaces. SC surfaces showed enhanced platelet adhesion, aggregation, and serotonin release compared to ES surfaces. This study shows that the morphology of samples plays a critical role on the biocompatibility of samples by altering the adsorption and adhesion of biomolecules and cells. The low level of adhesion, low aggregation, and serotonin release of platelets, even in the presence of fibrinogen and plasma-derived proteins, suggested that ES samples have the least thrombogenicity.

Biodegradable and 3D printable lysine functionalized polycaprolactone scaffolds for tissue engineering applications.

Tissue engineering (TE) has sparked interest in creating scaffolds with customizable properties and functional bioactive sites. However, due to limitations in medical practices and manufacturing technologies, it is challenging to replicate complex porous frameworks with appropriate architectures and bioactivity in vitro. To address these challenges, herein, we present a green approach that involves the amino acid (l-lysine) initiated polymerization of ɛ-caprolactone (CL) to produce modified polycaprolactone (PCL) with favorable active sites for TE applications. Further, to better understand the effect of morphology and porosity on cell attachment and proliferation, scaffolds of different geometries with uniform and interconnected pores are designed and fabricated, and their properties are evaluated in comparison with commercial PCL. The scaffold morphology and complex internal micro-architecture are imaged by micro-computed tomography (micro-CT), revealing pore size in the range of ~300-900 µm and porosity ranging from 30 to 70 %, while based on the geometry of scaffolds the compressive strength varied from 143 ± 19 to 214 ± 10 MPa. Additionally, the degradation profiles of fabricated scaffolds are found to be influenced by both the chemical nature and product design, where Lys-PCL-based scaffolds with better porosity and lower crystallinity degraded faster than commercial PCL scaffolds. According to in vitro studies, Lys-PCL scaffolds have produced an environment that is better for cell adhesion and proliferation. Moreover, the scaffold design affects the way cells interact; Lys-PCL with zigzag geometry has demonstrated superior in vitro vitality (>90 %) and proliferation in comparison to other designs. This study emphasizes the importance of enhancing bioactivity while meeting morphology and porosity requirements in the design of scaffolds for tissue engineering applications.

3D-Printable Sustainable Bioplastics from Gluten and Keratin.

Bioplastic films comprising both plant- and animal-derived proteins have the potential to integrate the optimal characteristics inherent to the specific domain, which offers enormous potential to develop polymer alternatives to petroleum-based plastic. Herein, we present a facile strategy to develop hybrid films comprised of both wheat gluten and wool keratin proteins for the first time, employing a ruthenium-based photocrosslinking strategy. This approach addresses the demand for sustainable materials, reducing the environmental impact by using proteins from renewable and biodegradable sources. Gluten film was fabricated from an alcohol-water mixture soluble fraction, largely comprised of gliadin proteins. Co-crosslinking hydrolyzed low-molecular-weight keratin with gluten enhanced its hydrophilic properties and enabled the tuning of its physicochemical properties. Furthermore, the hierarchical structure of the fabricated films was studied using neutron scattering techniques, which revealed the presence of both hydrophobic and hydrophilic nanodomains, gliadin nanoclusters, and interconnected micropores in the matrix. The films exhibited a largely (>40%) ß-sheet secondary structure, with diminishing gliadin aggregate intensity and increasing micropore size (from 1.2 to 2.2 µm) with an increase in keratin content. The hybrid films displayed improved molecular chain mobility, as evidenced by the decrease in the glass-transition temperature from ~179.7 °C to ~173.5 °C. Amongst the fabricated films, the G14K6 hybrid sample showed superior water uptake (6.80% after 30 days) compared to the pristine G20 sample (1.04%). The suitability of the developed system for multilayer 3D printing has also been demonstrated, with the 10-layer 3D-printed film exhibiting >92% accuracy, which has the potential for use in packaging, agricultural, and biomedical applications.

Near superhydrophobic fibrous scaffold for endothelialization: fabrication, characterization and cellular activities.

In this work, we have applied an electrospinning method to control wettability and further hydrophobic modification of a hydrophobic polymer mat of poly(vinylidene fluoride-co-hexafluoropropylene). A correlation between the processing parameters, rheological properties of polymer solutions, and electrospinning ability was made using the polymer's critical entanglement concentration, the boundary between the semidilute unentangled regime and the semidilute entangled regime. The wetting behavior, structural and thermal characteristics of electrospun (ES) mats were evaluated and compared with solvent cast sample using advancing and receding contact angle analyses, differential scanning calorimetry, and small-angle X-ray scattering. To demonstrate the feasibility, the best optimized ES samples were examined for their potential and ability to support bone marrow derived endothelial cell seeding efficiency, adhesion and proliferation. Our studies show that, while different processing techniques can effectively modulate physical and morphological changes such as porosity and hydrophobicity, the cellular adhesion and proliferation are highly time-dependent and controlled by chemical factors. As such, these results suggest that it is the interplay of both physical and chemical factors that determine the endothelialization of porous near superhydrophobic scaffolds. The developed electrospun samples demonstrate their feasibility for endothelialization.

Antifouling and Antimicrobial Study of Nanostructured Mixed-Matrix Membranes for Arsenic Filtration.

Membrane fouling is a major drawback in the membrane filtration industry for water treatment. Mixed-matrix membranes (MMMs) are well known for their enhanced antifouling and antibacterial properties, which could offer potential benefits for membrane filtration processes in the water treatment field. In this work, three electrospun nanofibrous MMMs (P, CP, and MCP, which were, respectively, the pristine polysulfone membrane and mixed-matrix membranes (MMMs) consisting of GO-ZnO and GO-ZnO-iron oxides) were studied for antifouling and antibacterial properties with respect to the arsenic nanofiltration process. The effects of these composites on the antifouling behaviour of the membranes were studied by characterising the bovine serum albumin (BSA) protein adsorption on the membranes and subsequent analysis using microscopic (morphology via scanning electron microscopy) and Brunauer-Emmett-Teller (BET) analyses. The antibacterial properties of these membranes were also studied against Gram-positive Staphylococcus aureus (S. aureus) and Gram-negative Escherichia coli (E. coli). The composite nanoparticle-incorporated membranes showed improved antifouling properties in comparison with the pristine polysulfone (PSF) membrane. The excellent antimicrobial properties of these membranes make them appropriate candidates to contribute to or overcome biofouling issues in water or wastewater treatment applications.

Electrospun Composite Nanofiltration Membranes for Arsenic Removal.

In recent years, significant attention has been paid towards the study and application of mixed matrix nanofibrous membranes for water treatment. The focus of this study is to develop and characterize functional polysulfone (PSf)-based composite nanofiltration (NF) membranes comprising two different oxides, such as graphene oxide (GO) and zinc oxide (ZnO) for arsenic removal from water. PSf/GO- and PSf/ZnO-mixed matrix NF membranes were fabricated using the electrospinning technique, and subsequently examined for their physicochemical properties and evaluated for their performance for arsenite-As(III) and arsenate-As(V) rejection. The effect of GO and ZnO on the morphology, hierarchical structure, and hydrophilicity of fabricated membranes was studied using a scanning electron microscope (SEM), small and ultra-small angle neutron scattering (USANS and SANS), contact angle, zeta potential, and BET (Brunauer, Emmett and Teller) surface area analysis. Fourier transform infrared spectroscopy (FTIR), X-ray diffraction (XRD), and X-ray photoelectron spectroscopy (XPS) were used to study the elemental compositions and polymer-oxide interaction in the membranes. The incorporation of GO and ZnO in PSf matrix reduced the fiber diameter but increased the porosity, hydrophilicity, and surface negative charge of the membranes. Among five membrane systems, PSf with 1% ZnO has the highest water permeability of 13, 13 and 11 L h-1 m-2 bar-1 for pure water, As(III), and As(V)-contaminated water, respectively. The composite NF membranes of PSf and ZnO exhibited enhanced (more than twice) arsenite removal (at 5 bar pressure) of 71% as compared to pristine PSf membranes, at 43%, whereas both membranes showed only a 27% removal for arsenate.

Crowder-directed interactions and conformational dynamics in multistimuli-responsive intrinsically disordered protein.

The consequences of crowding on the dynamic conformational ensembles of intrinsically disordered proteins (IDPs) remain unresolved because of their ultrafast motion. Here, we report crowder-induced interactions and conformational dynamics of a prototypical multistimuli-responsive IDP, Rec1-resilin. The effects of a range of crowders of varying sizes, forms, topologies, and concentrations were examined using spectroscopic, spectrofluorimetric, and contrast-matching small- and ultrasmall-angle neutron scattering investigation. To achieve sufficient neutron contrast against the crowders, deuterium-labeled Rec1-resilin was biosynthesized successfully. Moreover, the ab initio "shape reconstruction" approach was used to obtain three-dimensional models of the conformational assemblies. The IDP revealed crowder-specific systematic extension and compaction with the level of macromolecular crowding. Last, a robust extension-contraction model has been postulated to capture the fundamental phenomena governing the observed behavior of IDPs. The study provides insights and fresh perspectives for understanding the interactions and structural dynamics of IDPs in crowded states.

Mixed-Matrix Membrane Fabrication for Water Treatment.

In recent years, technology for the fabrication of mixed-matrix membranes has received significant research interest due to the widespread use of mixed-matrix membranes (MMMs) for various separation processes, as well as biomedical applications. MMMs possess a wide range of properties, including selectivity, good permeability of desired liquid or gas, antifouling behavior, and desired mechanical strength, which makes them preferable for research nowadays. However, these properties of MMMs are due to their tailored and designed structure, which is possible due to a fabrication process with controlled fabrication parameters and a choice of appropriate materials, such as a polymer matrix with dispersed nanoparticulates based on a typical application. Therefore, several conventional fabrication methods such as a phase-inversion process, interfacial polymerization, co-casting, coating, electrospinning, etc., have been implemented for MMM preparation, and there is a drive for continuous modification of advanced, easy, and economic MMM fabrication technology for industrial-, small-, and bulk-scale production. This review focuses on different MMM fabrication processes and the importance of various parameter controls and membrane efficiency, as well as tackling membrane fouling with the use of nanomaterials in MMMs. Finally, future challenges and outlooks are highlighted.

Microroughness induced biomimetic coating for biodegradation control of magnesium.

Herein we explore a combination of anodization induced micro-roughness and biomimetic coating on pure magnesium (Mg) metal at different applied voltages to control adhesion, biodegradation, and corrosion performance in simulated body fluid solution. The anodic film was fabricated using two different potentials, 3 and 5 V, respectively, to create microroughness on the Mg surface. The microroughened Mg surface was subsequently coated with a biomimetic silk thin film; and the characteristics of the treated Mg-substrates were evaluated using various spectroscopic, microscopic, immersion, and electrochemical techniques. A number of independent measurements, including hydrogen evolution, weight loss and electrochemical methods were employed to assess the corrosion characteristics. The silk-coated anodized samples revealed dramatically reduced degradation rate in terms of volume of hydrogen gas generation and weight loss compared to the respective anodized but uncoated, which revealed that optimized biomimetic silk-coated Mg surface (anodized at 5 V and subsequently biomimetic silk-coated ANMg5V) exhibited the best corrosion performance among all other tested samples. The ANMg5V Silk showed the highest polarization resistance (46.12 kΩ·cm2), protection efficiency (>0.99) and lowest corrosion rate (only 0.017 mm/year) relative to untreated Mg (8.457 mm/year), and anodized Mg (1.039 for anodized at 3 V and 0.986 for anodized at 5 V) surface due to the formation of a pore-free dense biomimetic protective film over Mg surface. The results of the cytotoxicity test confirm that silk-coated samples are significantly less cytotoxic compared to bare and anodized Mg samples. With enhanced corrosion resistance and cytocompatibility, silk-coated Mg could be a potential material for clinical applications.

Emerging bioadhesives: from traditional bioactive and bioinert to a new biomimetic protein-based approach.

Bioadhesives have reached significant milestones over the past two decades. Research has shown not only to produce adhesives capable of adhering to dry tissue but recently wet tissue as well. However, most bioadhesives developed have exhibited high adhesion strength yet lack other properties required for versatility in application, such as elasticity, biocompatibility and biodegradability. Adapting from limitations met from early bioadhesives and meeting the current demand allows novel bioadhesives to reach new milestones for the future. In this review, we overview the progression and variations of bioadhesives, current trends, characterisation techniques and conclude with future perspectives for bioadhesives for tissue engineering applications.

Resilin-mimetics as a smart biomaterial platform for biomedical applications.

Intrinsically disordered proteins have dramatically changed the structure-function paradigm of proteins in the 21st century. Resilin is a native elastic insect protein, which features intrinsically disordered structure, unusual multi-stimuli responsiveness and outstanding resilience. Advances in computational techniques, polypeptide synthesis methods and modular protein engineering routines have led to the development of novel resilin-like polypeptides (RLPs) including modular RLPs, expanding their applications in tissue engineering, drug delivery, bioimaging, biosensors, catalysis and bioelectronics. However, how the responsive behaviour of RLPs is encoded in the amino acid sequence level remains elusive. This review summarises the milestones of RLPs, and discusses the development of modular RLP-based biomaterials, their current applications, challenges and future perspectives. A perspective of future research is that sequence and responsiveness profiling of RLPs can provide a new platform for the design and development of new modular RLP-based biomaterials with programmable structure, properties and functions.

Polyelectrolyte Gels: Fundamentals, Fabrication and Applications.

Polyelectrolyte gels are an important class of polymer gels and a versatile platform with charged polymer networks with ionisable groups. They have drawn significant recent attention as a class of smart material and have demonstrated potential for a variety of applications. This review begins with the fundamentals of polyelectrolyte gels, which encompass various classifications (i.e., origin, charge, shape) and crucial aspects (ionic conductivity and stimuli responsiveness). It further centralises recent developments of polyelectrolyte gels, emphasising their synthesis, structure-property relationships and responsive properties. Sequentially, this review demonstrates how polyelectrolyte gels' flourishing properties create attractiveness to a range of applications including tissue engineering, drug delivery, actuators and bioelectronics. Finally, the review outlines the indisputable appeal, further improvements and emerging trends in polyelectrolyte gels.

Additive Manufacturing of Polymer Materials: Progress, Promise and Challenges.

The use of additive manufacturing (AM) has moved well beyond prototyping and has been established as a highly versatile manufacturing method with demonstrated potential to completely transform traditional manufacturing in the future. In this paper, a comprehensive review and critical analyses of the recent advances and achievements in the field of different AM processes for polymers, their composites and nanocomposites, elastomers and multi materials, shape memory polymers and thermo-responsive materials are presented. Moreover, their applications in different fields such as bio-medical, electronics, textiles, and aerospace industries are also discussed. We conclude the article with an account of further research needs and future perspectives of AM process with polymeric materials.

Engineering a Bioactive Hybrid Coating for In Vitro Corrosion Control of Magnesium and Its Alloy.

Magnesium (Mg) and its alloys are promising biodegradable metallic implant materials. However, their clinical applications are limited by their fast corrosion rate in the biological environment. In this work, with an outlook to improve the in vitro corrosion resistance of Mg and WE43 Mg alloy, a layer-by-layer interfacially engineered anticorrosive and bioactive coating consisting of a natural oxide lower layer, hydroxyapatite (HA) middle layer, and silk fibroin (SF) top layer was fabricated and investigated. Anodization was used to create natural oxide layer induced microroughness on substrates. The electrochemically deposited HA layer improved the surface microroughness and microhardness but significantly decreased Mg ion release, hydrogen gas evolution, and weight loss in simulated body fluid. The spin-coated SF layer further decreased hydrophilicity, in vitro degradation, and corrosion rate. The nonspecific and specific intermolecular interactions between fabricated layers along with their mechanical interlocking interface contributed to improved adhesion strength and integrity of the coating. The SF+HA-coated samples showed enhanced degradation and corrosion resistance due to a synergistic effect of the underlying HA layer, hindering the ingress of aggressive ions and the top hydrophobic SF layer, preventing the ingress of corrosive solution. The SF+HA-coated Mg and WE43 Mg alloy samples exhibited 50 and 26 times decreased corrosion rate, respectively, compared to uncoated samples. Moreover, in vitro cytotoxicity and cell culture studies using a mouse fibroblast cell showed that the SF+HA hybrid coating improved the cell viability, attachment, and proliferation, with cells exhibiting elongated morphology on coated samples as compared to a round shape on uncoated samples.

Nanostructure evolution in high-temperature perfluorosulfonic acid ionomer membrane by small-angle X-ray scattering.

The high-temperature morphology of supported liquid membranes (SLMs) prepared from perfluorinated membranes such as Nafion and Hyflon and hydrophobic ionic liquid 1-butyl-3-methylimidazolium bis(trifluoromethylsulfonyl)imide (BMI-TFSI) has been investigated by small-angle X-ray scattering (SAXS). Proton conductivity results of SLMs before and after leaching show an increase in conductivity with temperature up to 160 °C in an anhydrous environment. DSC results show that crystallites within perfluorinated membranes are thermally stable up to 196 °C. High-temperature SAXS results have been used to correlate structure and morphology of supported liquid membranes with high-temperature conductivity data. The ionic liquid essentially acts as a proton solvent in a similar way to water in hydrated Nafion membranes and increases size of clusters, which allow percolation to be achieved more easily. The cation of the ionic liquid interacts with sulfonate groups within ionic domains through electrostatic interactions and displaces protons. Protons can associate with free anions of the ionic liquid, which are loosely associated with cations and can transport by hopping from anion sites within the membrane. The ionic liquid contributes to proton conductivity at high temperature through achievement of long-range ordering and subsequent percolation.

Silk fibroins in multiscale dimensions for diverse applications.

Silk biomaterials in different forms such as particles, coatings and their assemblies, represent unique type of materials in multiple scales and dimensions. Herein, we provide an overview of multi-scale silk fibroin materials including silk particles, silk coatings and silk assemblies, each of which represents a unique type of material with wide range of applications. They feature tunable structures and mechanical properties with excellent biocompatibility, which are essentially required for various biomedical and drug delivery applications. The review focuses on bringing a new perspective on the utilization of regenerated silk fibroins in modern biomedicine by beginning with the fabrication of silk in multiscale dimensions and their state-of-the-art applications in various biomedical and bioelectronic fields. It covers the fundamentals of processing silk fibroins in multi-dimensions (sizes and shapes) with a specific emphasis on its structural tunability at various length scales (nano-micro) by using the latest fabrication methods/mechanisms and advanced fabrication technologies, followed by their recent applications in diverse fields of biomedicine.

Bioprintable tough hydrogels for tissue engineering applications.

Bioprinting is an advanced fabrication approach to engineer complex living structures as the conventional fabrication methods are incapable of integrating structural and biological complexities. It offers the versatility of printing different cell incorporated hydrogels (bioink) layer by layer; offering control over spatial resolution and cell distribution to mimic native tissue architectures. However, the bioprinting of tough hydrogels involve additional complexities, such as employing complex crosslinking or reinforcing mechanisms during printing and pre/post printing cellular activities. Solving this complexity requires attention from engineering, material science and cell biology perspectives. In this review, we discuss different types of bioprinting techniques with focus on current state-of-the-art in bioink formulations and pivotal characteristics of bioinks for tough hydrogel printing. We discuss the scope of transition from 3D to 4D bioprinting and some of the advanced characterization techniques for in-depth understanding of the 3D printing process from the microstructural perspective, along with few specific applications and conclude with the future perspectives in biofabrication of hydrogels for tissue engineering applications.