Electro-Biofabrication. Coupling Electrochemical and Biomolecular Methods to Create Functional Bio-Based Hydrogels.

Twenty years ago, this journal published a review entitled "Biofabrication with Chitosan" based on the observations that (i) chitosan could be electrodeposited using low voltage electrical inputs (typically less than 5 V) and (ii) the enzyme tyrosinase could be used to graft proteins (via accessible tyrosine residues) to chitosan. Here, we provide a progress report on the coupling of electronic inputs with advanced biological methods for the fabrication of biopolymer-based hydrogel films. In many cases, the initial observations of chitosan's electrodeposition have been extended and generalized: mechanisms have been established for the electrodeposition of various other biological polymers (proteins and polysaccharides), and electrodeposition has been shown to allow the precise control of the hydrogel's emergent microstructure. In addition, the use of biotechnological methods to confer function has been extended from tyrosinase conjugation to the use of protein engineering to create genetically fused assembly tags (short sequences of accessible amino acid residues) that facilitate the attachment of function-conferring proteins to electrodeposited films using alternative enzymes (e.g., transglutaminase), metal chelation, and electrochemically induced oxidative mechanisms. Over these 20 years, the contributions from numerous groups have also identified exciting opportunities. First, electrochemistry provides unique capabilities to impose chemical and electrical cues that can induce assembly while controlling the emergent microstructure. Second, it is clear that the detailed mechanisms of biopolymer self-assembly (i.e., chitosan gel formation) are far more complex than anticipated, and this provides a rich opportunity both for fundamental inquiry and for the creation of high performance and sustainable material systems. Third, the mild conditions used for electrodeposition allow cells to be co-deposited for the fabrication of living materials. Finally, the applications have been expanded from biosensing and lab-on-a-chip systems to bioelectronic and medical materials. We suggest that electro-biofabrication is poised to emerge as an enabling additive manufacturing method especially suited for life science applications and to bridge communication between our biological and technological worlds.

Photothermal Fibrous Chitosan/Polydopamine Sponge for Intraoperative Hemostasis and Prevention of Tumor Recurrence in Hepatocellular Carcinoma Resection.

Hepatectomy, a surgical procedure for liver cancer, is often plagued by high recurrence rates worldwide. The recurrence of liver cancer is primarily attributed to microlesions in the liver, changes in the immune microenvironment, and circulating tumor cells in the bloodstream. To address this issue, a novel intervention method that combines intraoperative hemostasis with mild photothermal therapy is proposed, which has the potential to ablate microlesions and improve the immune microenvironment simultaneously. Specifically, the integrated strategy is realized based on the fibrous chitosan/polydopamine sponge (CPDS), which is constructed from shearing-flow-induced oriented hybrid chitosan fibers and subsequent self-assembly of polydopamine. The CPDS demonstrates high elasticity, excellent water absorption, and photothermal conversion performance. The results confirm the efficient hemostatic properties of the fibrous CPDS in various bleeding models. Notably, in subcutaneous and orthotopic postoperative recurrence and metastasis models of hepatocellular carcinoma, the fibrous CPDS significantly inhibits local tumor recurrence and distant metastasis. Moreover, the combination with lenvatinib can substantially enhance the antitumor effect. This comprehensive treatment strategy offers new insights into hepatectomy of liver cancer, representing a promising approach for clinical management.

Cellulose nanofibers embedded chitosan/tannin hydrogel with high antibacterial activity and hemostatic ability for drug-resistant bacterial infected wound healing.

Millions of patients annually suffer life-threatening illnesses caused by bacterial infections of skin wounds. However, the treatment of wounds infected with bacteria is a thorny issue in clinical medicine, especially with drug-resistant bacteria infections. Therefore, there is an increasing interest in developing wound dressings that can efficiently fight against drug-resistant bacterial infections and promote wound healing. In this work, an anti-drug-resistant bacterial chitosan/cellulose nanofiber/tannic acid (CS/CNF/TA) hydrogel with excellent wound management ability was developed by electrospinning and fiber breakage-recombination. The hydrogel exhibited an outstanding antibacterial property exceeding 99.9 %, even for drug-resistant bacteria. This hydrogel could adhere to the tissue surface due to its abundant catechol groups, which avoided the shedding of hydrogel during the movement. Besides, it exhibited extraordinary hemostatic ability during the bleeding phase of the wound and then regulated the wound microenvironment by absorbing water and moisturizing. Moreover, the CS/CNF/TA also promoted the regrowth of vessels and follicles, accelerating the healing of infected wound tissue, with a healing rate exceeding 95 % within a 14-day timeframe. Therefore, the CS/CNF/TA hydrogel opens a new approach for the healing of drug-resistant bacterial infected wounds.

Biological MWCNT/chitosan composite coating with outstanding anti-corrosion property for implants.

Biocompatible coatings that can protect metal implants have great potential in tissue engineering. In this work, MWCNT/chitosan composite coatings with hydrophobic-hydrophilic asymmetric wettability were facilely prepared by one-step in situ electrodeposition. The resultant composite coating exhibits excellent thermal stability and mechanical strength (0.76 MPa), benefiting from the compact internal structure. The thickness of the coating can be controlled precisely by the amounts of transferred charges. The MWCNT/chitosan composite coating demonstrates a lower corrosion rate due to its hydrophobicity and compact internal structure. Compared with exposed 316 L stainless steel, its corrosion rate is reduced by two orders of magnitude from 3.004 × 10-1 mm/yr to 5.361 × 10-3 mm/yr. The content of iron released from 316 L stainless steel into the simulated body fluid drops to 0.1 mg/L under the protection of the composite coating. In addition, the composite coating enables efficient calcium enrichment from simulated body fluids and promotes the formation of bioapatite layers on the coating surface. This study contributes to furthering the practical application of chitosan-based coatings in implant anticorrosion.

Antifatigue Hydration-Induced Polysaccharide Hydrogel Actuators Inspired by Crab Joint Wrinkles.

Joint wrinkles in animals facilitate frequent bending and contribute to the duration of the joint. Inspired by the morphology and function of joint wrinkles, we developed a bionic hydration-induced polymeric actuator with constructed wrinkles at the selected area. Specifically, we adopt electrical writing to create defined single and double cross-linking regions on chitosan (CS) hydrogel. The covalent cross-linking network was constructed by electrical writing-induced covalent cross-linking between CS chains and epichlorohydrin. Subsequent treatment of sodium dodecyl sulfate allows electrostatic cross-linking at the unwritten area with the simultaneous formation of surface wrinkles. The resulting single and double cross-linking hydrogel demonstrates spontaneous deformation behaviors by the influx and efflux of H2O to the electrostatic cross-linking domain under different ion concentrations. Importantly, the wrinkle structure endows the hydrogel with extraordinary antifatigue bending performance. By regulating the surface morphology and spatial cross-linking, we can design novel biomimetic polysaccharide hydrogel actuators with fascinating functions.

Electrodeposition induced covalent cross-linking of chitosan for electrofabrication of hydrogel contact lenses.

To meet the requirement of personalization, there is an urgent need to develop a simple, efficient and versatile manufacturing method for customized contact lens manufabrication. Here, we report a novel electrofabrication methodology (i.e., electrodeposition) for the fabrication of hydrogel contact lenses, which can induce covalent cross-linking between chitosan and epichlorohydrin simultaneously. The transmittance and toughness of hydrogels are improved by electrochemical cross-linking without affecting their oxygen permeability. Furthermore, the geometry of the chitosan based hydrogel contact lenses can be customized simply by the electrode template, and its characteristics can be regulated by electrical signals and electrochemical cross-linking. The electrodeposited hydrogel contact lenses have good optical properties, mechanical properties and biocompatibility, and their anti-adhesion properties to Staphylococcus aureus are close to commercial contact lenses. This work reveals the mechanism of electrochemical cross-linking between chitosan and epichlorohydrin and provides an alternative method for contact lens fabrication.

Highly sensitive formaldehyde sensors based on CuO/ZnO composite nanofibrous mats using porous cellulose acetate fibers as templates.

An innovative formaldehyde sensor based on CuO/ZnO composite nanofibrous mats (C-NFMs) coated quartz crystal microbalance (QCM), which is capable of stable determination of formaldehyde gas at ambient temperatures sensitively and selectively, has been successfully fabricated. Triaxial and highly porous C-NFMs with high surface area (126.53 m2 g-1) were synthesized by electrospinning a sol-gel cellulose acetate (CA)/CuAc2/ZnAc2 complex solution and following by calcination process. Benefiting from the unique heterojunction structure, immense pore interconnectivity and large surface area of C-NFMs, the as-developed QCM sensors exhibited an extremely low limit of detection (LOD) down to 26 ppb and a limit of quantification value equals to 87 ppb. Besides, the C-NFMs coated QCM sensors also demonstrated short response times (80s), the long-term stability during 3 weeks as well as good selectivity to formaldehyde over diverse volatile organic compounds. The sorption equilibrium in the adsorption process of QCM coated sensors was well met with the Freundlich model, which certified the heterogeneous adsorption between formaldehyde gas and C-NFMs.

Scalable Production of Biodegradable, Recyclable, Sustainable Cellulose-Mineral Foams via Coordination Interaction Assisted Ambient Drying.

Heavy reliance on petrochemical-based plastic foams in both industry and society has led to severe plastic pollution (the so-called "white pollution"). In this work, we develop a biodegradable, recyclable, and sustainable cellulose/bentonite (Cel/BT) foam material directly from resource-abundant natural materials (i.e., lignocellulosic biomass and minerals) via ambient drying. The strong resistance to the capillary force-driven structural collapse of the preformed three-dimensional (3D) network during the ambient drying process can be ascribed to the purpose-designed cellulose-bentonite coordination interaction, which provides a practical way for the locally scalable production of foam materials with designed shapes without complex processing and intensive energy consumption. Benefiting from the strong cellulose-bentonite coordination interaction, the Cel/BT foam material demonstrates high mechanical strength and outstanding thermal stability, outperforming commercial plastic polystyrene foam. Furthermore, the Cel/BT foam presents environmental impacts much lower than those of petrochemical-based plastic foams as it can be 100% recycled in a closed-loop recycling process and easily biodegraded in the environment (natural cellulose goes back to the carbon cycle, and bentonite minerals return to the geological cycle). This study demonstrates an energy-efficient ambient drying approach for the local and scalable production of an all-natural cellulose/bentonite foam for sustainable packaging, buildings, and beyond, presenting great potential in response to "white pollution" and resource shortage.

A multifunctional dual-shell magnetic nanocomposite with near-infrared light response for synergistic chemo-thermal tumor therapy.

The synergistic tumor therapy in single nanoplatform has always been the goal for high efficacy tumor treatment while still remains great challenge. This paper reports a versatile nanotheranostic platform enlisting magnetic iron oxide nanoparticles, polydopamine (PDA), gold nanocages (Au nanocage) and metal organic framework (MOF, MIL101-NH2 ) in order to achieve synergistic chemothermal tumor therapy both in vitro and in vivo. The prepared magnetic photothermal nanoparticles (MPNPs) exhibit high drug loading capacity (31.34 mg/g), superior photo-thermal capacity (11.5°C enhancement in 180 s), low bio-toxicity, good magnetic resonance with a low dosage of 22 µg/g, as well as high antitumor efficacy in vivo. Such a novel and multifunctional nanoplatform is expected to find promising applications in target tumor synergistic therapy.

TiO2/rectorite-trapped cellulose composite nanofibrous mats for multiple heavy metal adsorption.

The anthropogenic release of highly toxic heavy metals into the environment presents a huge challenge for ecosystems and human society. Recoverable and efficient adsorption materials could be obtained by trapping inorganic adsorbents (e.g., TiO2 nanoparticles and rectorite (REC)), in a natural polymer matrix. In this study, a series of cellulose-TiO2/REC composite nanofibrous mats were fabricated via electrospinning. The interactions between inorganic adsorbents and cellulose molecules improved the thermal stability, surface area, tensile strength and adsorption capacity of the mats. We focused on the adsorption of Pb2+, Cu2+ and Cd2+ from acidic solutions onto cellulose-TiO2/REC composite nanofibrous mats in multiple systems because the magnitudes of heavy metal concentrations in wastewater typically varied. The maximum total adsorption capacity of 69.81 mg/g was obtained by Cellulose-TiO2/REC2:1 nanofibrous mats. The composite nanofibrous mats successfully trapped TiO2 nanoparticles, and the obtained cellulose-TiO2/REC nanofibrous mats could be used to remove heavy metals from acidic wastewater.

Electrofabrication of flexible and mechanically strong tubular chitosan implants for peripheral nerve regeneration.

The development of peripheral nerve tissue engineering requires a safe and reliable methodology to construct biodegradable conduits. Herein, a new type of chitosan-based nerve-guide hydrogel conduit (CNHC) with enhanced mechanical flexibility in the wet state was fabricated using a one-step electrofabrication technology. The formation of the chitosan conduit is a physical process which can be conducted in a mild water phase without toxic crosslinks. The current density during electrofabrication has a profound effect on the physical and structural properties of the conduits. Cytocompatibility results indicate that the CNHC can promote cell proliferation and adhesion. Functional and histological tests indicate that the CNHC has the ability to guide the growth of axons through the conduit to reach a distal stump, which is closely similar to the autograft group. Overall, the results of this study demonstrate that the CNHCs from electrofabrication have a great potential in peripheral nerve regeneration.

Chitosan and collagen layer-by-layer assembly modified oriented nanofibers and their biological properties.

Layer-by-layer self-assembly (LBL) is an effective method to prepare potential biomaterial with multilayer coatings, and few reports have focused on the variation of oriented microstructure during LBL process. In this study, polycaprolactone (PCL) and type І collagen (COL) were electrospun to oriented nanofibrous mats, and chitosan (CS) and COL molecules were then deposited on the mats by LBL technique. Zeta potential, FT-IR analysis and XPS measurement indicated the successful fabrication and modification. Changes in surface morphology and increase in surface roughness were observed in LBL process. Additionally, LBL-structured mats exhibited improved mechanical properties with the maximal tensile strength of 35.1 ± 7.0 MPa and the best elongation of 106.0 ± 11.5 %. CCK-8 and live/dead assays illustrated that the cell viability of the mats increased more than 20 % after LBL modification. More importantly, cells seeded onto the mats showed oriented adhesion and growth along the direction of nanofiber arrangement in LBL modified mats, which provided an effective strategy for realizing the controlled growth of cells.

One-step electrochemically induced counterion exchange to construct free-standing carboxylated cellulose nanofiber/metal composite hydrogels.

The fabrication of polymeric composite hydrogel with hierarchical structure in a simple, controllable, and straightforward process poses great importance for manufacturing nanomaterials and subsequent applications. Herein, we report a one-step and template-free counterion exchange method to construct free-standing carboxylated cellulose nanofiber composite hydrogels. Metal ions were electrochemically and locally released from the electrode and chelated with carboxylated cellulose nanofibers, leading to the in-situ formation of composite hydrogels. The properties of composite hydrogels can be easily programmed by the type of electrode, current density, and electrodeposited suspension. Significantly, the composited hydrogels exhibited interconnected nanoporous structure, enhanced thermal degradation, improved mechanical strength and antibacterial activity. The results suggest great potential of anodic electrodeposition to fabricate nanofiber/metal composite hydrogels.

Electrodeposition of Polysaccharide and Protein Hydrogels for Biomedical Applications.

In the last few decades, polysaccharide and protein hydrogels have attracted significant attentions and been applied in various engineering fields. Polysaccharide and protein hydrogels with appealing physical and biological features have been produced to meet different biomedical applications for their excellent properties related to biodegradability, biocompatibility, nontoxicity, and stimuli responsiveness. Numerous methods, such as chemical crosslinking, photo crosslinking, graft polymerization, hydrophobic interaction, polyelectrolyte complexation and electrodeposition have been employed to prepare polysaccharide and protein hydrogels. Electrodeposition is a facile way to produce different polysaccharide and protein hydrogels with the advantages of temporal and spatial controllability. This paper reviews the recent progress in the electrodeposition of different polysaccharide and protein hydrogels. The strategies of pH induced assembly, Ca2+ crosslinking, metal ions induced assembly, oxidation induced assembly derived from electrochemical methods were discussed. Pure, binary blend and ternary blend polysaccharide and protein hydrogels with multiple functionalities prepared by electrodeposition were summarized. In addition, we have reviewed the applications of these hydrogels in drug delivery, tissue engineering and wound dressing.

Applications of chitin and chitosan nanofibers in bone regenerative engineering.

Promoting bone regeneration and repairing defects are urgent and critical challenges in orthopedic clinical practice. Research on bone substitute biomaterials is essential for improving the treatment strategies for bone regeneration. Chitin and its derivative, chitosan, are among the most abundant natural biomaterials and widely found in the shells of crustaceans. Chitin and chitosan are non-toxic, antibacterial, biocompatible, degradable, and have attracted significant attention in bone substitute biomaterials. Chitin/chitosan nanofibers and nanostructured scaffolds have large surface area to volume ratios and high porosities. These scaffolds can be fabricated by electrospinning, thermally induced phase separation and self-assembly, and are widely used in biomedical applications such as biological scaffolds, drug delivery, bacterial inhibition, and wound dressing. Recently, some chitin/chitosan-based nanofibrous scaffolds have been found structurally similar to bone's extracellular matrix and can assist in bone regeneration. This review outlines the biomedical applications and biological properties of chitin/chitosan-based nanofibrous scaffolds in bone tissue engineering.

Incorporating chitin derived glucosamine sulfate into nanofibers via coaxial electrospinning for cartilage regeneration.

Chitin is the second abundant natural polysaccharide, and the development of chitin and its derivatives have received more and more attention. Glucosamine sulfate (GAS) obtained by the hydrolysis of chitin can promote the growth of chondrocytes. The coaxial electrospinning technology had been utilized to encapsulated GAS into the core of polycaprolactone (PCL) nanofibers. It could protect the GAS from the environment and allow it to release sustainably over time. From the results of scanning electron microscopy (SEM), PCL/GAS nanofibers performed a typical fiber scaffold surface. Transmission electron microscopy (TEM) and energy dispersive X-ray spectroscopy (EDX) results confirmed the feasibility to use coaxial electrospinning to load GAS. Tensile tests exhibited that PCL/GAS mats could be stretched more than twice its own length. Besides, the cell experiments illustrated that PCL/GAS had a significant effect on the proliferation and growth of rat articular chondrocytes, attesting its prospect for cartilage regeneration.

Chitosan/tannic acid bilayers layer-by-layer deposited cellulose nanofibrous mats for antibacterial application.

The research and development of environmentally friendly and nontoxic biomass products has become an important topic of worldwide concern. In this study, natural materials were used for producing a kind of antibacterial mats. Cellulose acetate (CA) mats prepared by electrospinning technology were converted to cellulose mats via alkali hydrolysis. Chitosan (CS) and tannic acid (TA) were used to fabricate the composite mats by using layer-by-layer (LBL) self-assembly technology. The cellulose mats exhibited good fibrous structure, three-dimensional network and small average fiber diameter ranging from 300 to 400 nm. Besides, the results of mechanical properties testing and water contact angle measurements of these LBL-structured mats demonstrated that the LBL technology was able to improve their surface characteristics, hydrophilicity and mechanical properties. The analysis of antibacterial activity of the mats revealed over 86% antibacterial activity against Escherichia coli and up to 99% antibacterial activity against Staphylococcus aureus. Hence, the LBL-structured cellulose mats have excellent antibacterial activity and mechanical properties. Therefore, these nano-cellulose mats can be expected to have considerable development prospects for food packaging or wound dressing.

Egg source natural proteins LBL modified cellulose nanofibrous mats and their cellular compatibility.

Natural-based nanocomposites are competitive and promising materials for biomedical applications due to their biocompatibility. Herein, a novel natural-based composite was fabricated by alternately depositing lysozyme (LY) and albumin egg (AE) on electrospun cellulose nanofibrous mats via layer-by-layer self-assembly (LBL) technology. To indicate the successful deposition process and investigate the variations of the mats during LBL process, the surface morphology, physical property, chemical composition, wetting behavior and thermal stability were systematically studied. The results showed that the surface morphology and composition of the mats were significantly influenced by LBL process, which further resulted in the variation of wetting behavior. Besides, the mechanical properties were enhanced after LBL modification. In addition, the LBL structured nanofibrous mats exhibited antibacterial activity and excellent biocompatibility with L929 fibroblasts. In brief, LY and AE coated LBL structured cellulose nanofibrous mats, especially the 15 bilayers coated mats, have considerably potential applications in the biomedical field.

Construction of cellulose nanofibers/quaternized chitin/organic rectorite composites and their application as wound dressing materials.

Traumatic injury is a major cause of mortality, and poor wound healing affects millions of people. Thus, the development of effective wound dressings is essential for speeding up wound healing and decreasing mortality. In this study, a suspension of carboxylated brown algae cellulose nanofibers (BACNFs) with a high aspect ratio was freeze dried to prepare a sponge. The sponge showed high porosity and water absorption capacity; thus, it can absorb wound exudates when used as a wound dressing. In addition, quaternized ß-chitin (QC) with antibacterial properties was intercalated into the interlayer space of the organic rectorite (OREC) via electrostatic interactions to obtain composite suspensions (QCRs) with improved antimicrobial activity compared to that of QC alone. Subsequently, the BACNF sponge was soaked in the QCR suspension to absorb QCRs via electrostatic interactions and hydrogen bonding from which cellulose nanofiber/quaternized chitin/organic rectorite composite (BACNF/QCR) sponges were constructed via freeze-drying. The in vivo animal tests demonstrated that the BACNF/QCR sponges rapidly induced hemostasis in a rat tail amputation test, making them superior to the traditional hemostatic materials. Furthermore, BACNFs/QCRs could substantially promote collagen synthesis and neovascularization, thereby accelerating wound healing 3 days earlier than gauze. This multi-functional biomedical material, fabricated using natural substances, shows great potential to be used for wound healing.

A simple mechanical agitation method to fabricate chitin nanogels directly from chitin solution and subsequent surface modification.

The development of physical approaches and green technologies to construct novel chitin materials is essential for the exquisite utilization of the renewable and valuable resource of chitin. In the present study, chitin nanogels were simply fabricated from a chitin solution dissolved in 8% NaOH/4% urea aqueous solvent by high speed stirring. The mechanical stirring generated in situ heat that induced the regeneration of chitin chains and ensured good dispersion of the nanogels. The prepared nanogels were composed of spherical nanoparticles of size 20 to 30 nm with some aggregates. The formation of chitin nanogels was confirmed to be a physical process without using organic solvent or chemical crosslinking. Rheological tests revealed a shear thinning behavior of the nanogels and injectable hydrogels were developed accordingly. The chitin nanogels showed no toxicity to L929 cells and cell attachment on the surface of the nanogel was observed. Further, monodispersed cationic nanogels and anionic nanogels were facilely obtained by deacetylating and TEMPO-mediated oxidizing chitin nanogels, and demonstrated different antibacterial properties.