Biological Nanofibers Research Articles

Complex Characterization of Nanofiber Biomaterials Based on Chitosan, Collagen and Fish Gelatine Produced by Electrospinning*

Electrospinning is an increasingly used technique for producing nanofibers from various types of polymers and biopolymers. These fibers are suitable for applications in tissue engineering, wound healing and drug delivery. In this study, we focused on biomaterials produced from raw marine fish collagen, fish gelatin and chitosan. Various collagen/chitosan and fish gelatin/chitosan in several ratios were dissolved in acetic acid. The formed solutions have concentrations in the range from 5 to 90% (g/ml). The effect of different molecular formats (native and denatured collagen) was studied. The prepared solutions were used in electrospinning experiments. The order degree of various phases of raw materials was established from x-ray diffraction (XRD) measurements. The raw materials, bio-polymeric solutions and finite nanofibers biomaterials were characterized using Fourier transform infrared (FT-IR) spectroscopy, scanning electron microscopy (SEM), low-field 1D NMR (nuclear magnetic resonance) relaxometry, 2D NMR T1-T2 COSY (correlation spectroscopy), T1-T2 EXSY (exchange spectroscopy) maps, and high field localized 1H NMR spectroscopy. The T 2-T 2 EXSY experiments showed that the number of components characterized by distinct dynamics is larger than those that can by estimated from 1D T 2-distributions due to molecular exchange. Moreover, the full complexity was also revealed by T 1-T 2 COSY maps. The study provides a better understanding of the nanofiber properties, demonstrates the potential of electrospinning for producing nanofibers, and highlights the importance of using multiple characterization methods for a better understanding of the properties of biological nanofibers.

Mechanical properties of collagen fibrils determined by buckling analysis

The mechanical properties of biological nanofibers such as collagen fibrils are important in many applications, ranging from tissue-engineering to cancer treatment. However, mechanical testing is not straightforward at the nanometer scale. Here, we use the theory of column-buckling to determine the bending properties of individual collagen fibrils. To achieve this, fibrils were deposited on a manually pre-stretched foil, which was then released with the fibrils attached. Atomic Force Microscopy (AFM) imaging was used to determine the tensile modulus by measuring the buckling-wavelength and the radius for each fibril. Comparison with data obtained by AFM nanoindentation and other, more sophisticated methods, shows that our results are in very good agreement. The great advantage of this simple approach is that it can be used to quickly determine mechanical properties without force or stress-strain measurements, which are challenging to obtain accurately and at high throughput at the nanoscale. The method could be applied to any nanofibers, not just collagen fibrils. Statement of significanceCollagen fibrils are the main constituent of the extracellular matrix, and alterations of their mechanical properties can have significant effects on cell adhesion and motility. This has, ultimately, implications in age-related diseases and cancer. Furthermore, tuning the mechanical properties of collagen fibrils could be an important tool in the design of artificial cell scaffolds in tissue-engineering. For these reasons, it is important to have methods that can be used to determine the mechanical properties of fibrils at the single-fibril level and, therefore, at the nanometer scale. The method presented here has the advantage of being easy to use and avoids some of the fundamental issues of more established methods.

Transparent and flexible structurally colored biological nanofiber films for visual gas detection

•Construct structurally colored film using sustainable biological nanofibers •Multiple advantages are integrated into the one •Achieve ultrafast color response to environment stimuli of less than 1 s •Realize visual gas concentration detection simply upon blowing Structurally colored films hold promise as cheap platforms for use in visual detection. However, achieving accurate stimuli-responsive functions that can be distinguished by the naked eye in a single thin, robust, and practically useful material remains a challenge. Herein, we demonstrate flexible and self-standing structurally colored films fabricated by filtrating sustainable hydrophilic nanofibrillated cellulose through a porous hydrophobic membrane, followed by natural drying. 480–644 nm thick and composed of densely packed, high-aspect-ratio nanofibers entangled in web-like structures, the films are transparent, tailorable into a variety of shapes, and easily foldable. They exhibit iridescence owing to thin-film interference and reversibly change color in seconds when exposed to various moist gases and liquids. We validate the visual gas-detection capabilities of the films by measuring the ethanol concentration in the breath of an inebriated participant and envisage that these materials will find use in automated and broad-spectrum gas-detection devices. Structurally colored films hold promise as cheap platforms for use in visual detection. However, achieving accurate stimuli-responsive functions that can be distinguished by the naked eye in a single thin, robust, and practically useful material remains a challenge. Herein, we demonstrate flexible and self-standing structurally colored films fabricated by filtrating sustainable hydrophilic nanofibrillated cellulose through a porous hydrophobic membrane, followed by natural drying. 480–644 nm thick and composed of densely packed, high-aspect-ratio nanofibers entangled in web-like structures, the films are transparent, tailorable into a variety of shapes, and easily foldable. They exhibit iridescence owing to thin-film interference and reversibly change color in seconds when exposed to various moist gases and liquids. We validate the visual gas-detection capabilities of the films by measuring the ethanol concentration in the breath of an inebriated participant and envisage that these materials will find use in automated and broad-spectrum gas-detection devices.

Electrospun Nanofibrous Scaffolds: Review of Current Progress in the Properties and Manufacturing Process, and Possible Applications for COVID-19.

Over the last twenty years, researchers have focused on the potential applications of electrospinning, especially its scalability and versatility. Specifically, electrospun nanofiber scaffolds are considered an emergent technology and a promising approach that can be applied to biosensing, drug delivery, soft and hard tissue repair and regeneration, and wound healing. Several parameters control the functional scaffolds, such as fiber geometrical characteristics and alignment, architecture, etc. As it is based on nanotechnology, the concept of this approach has shown a strong evolution in terms of the forms of the materials used (aerogels, microspheres, etc.), the incorporated microorganisms used to treat diseases (cells, proteins, nuclei acids, etc.), and the manufacturing process in relation to the control of adhesion, proliferation, and differentiation of the mimetic nanofibers. However, several difficulties are still considered as huge challenges for scientists to overcome in relation to scaffolds design and properties (hydrophilicity, biodegradability, and biocompatibility) but also in relation to transferring biological nanofibers products into practical industrial use by way of a highly efficient bio-solution. In this article, the authors review current progress in the materials and processes used by the electrospinning technique to develop novel fibrous scaffolds with suitable design and that more closely mimic structure. A specific interest will be given to the use of this approach as an emergent technology for the treatment of bacteria and viruses such as COVID-19.

Environmentally friendly biological nanofibers based on waste feather keratin by electrospinning with citric acid vapor modification

AbstractWaste feather keratin (FK)‐based nanofibers by electrospinning and citric acid (CA) vapor modification has been successfully prepared and investigated. FK, poly(vinyl alcohol), and poly(ethylene oxide) have been used as raw materials and CA vapor as cross‐linker. The structural, thermal, hydrophobicity, and mechanical properties of FK‐based nanofibers by CA vapor modification with various cross‐linking time have been completely explored. In order to investigate the effect of H2O vapor on CA vapor modification, H2O vapor modification was performed on the FK‐based nanofibers at the same conditions. The results show that the average diameter of nanofibers increased from 250.83 ± 29.65 nm to 338.79 ± 31.43 nm by CA vapor modification with 15 h. Similarly, the thermal stability and water resistance of FK‐based nanofibers by CA vapor modification have been significantly improved. The tensile strength (σb) and elongation at breakage point (εb) of FK‐based nanofibers after CA vapor modified for 15 h were about 1.5 and 2 times higher than that of nonmodified nanofibers, respectively. By comparison, scanning electron microscopy results suggest that the FK‐based nanofibers modified by H2O vapor cannot maintain the morphology of the nanofibers, resulting in large‐scale adhesion. The thermal properties of FK‐based nanofibers with H2O vapor modification have no obvious change. The hydrophobicity and mechanical properties of FK‐based nanofibers by H2O vapor modification are not as good as that of CA vapor modification. In summary, these results exhibit that nontoxic and natural CA can be used as cross‐linking agent to enhance the comprehensive performance of FK‐based nanofibers. This study provides a new method to modify FK‐based nanofibers and refined the waste feathers, which not only protected the environment, but also gained benefits, which has a broad application prospect.

Phage nanofibers in nanomedicine: Biopanning for early diagnosis, targeted therapy, and proteomics analysis

Display of a peptide or protein of interest on the filamentous phage (also known as bacteriophage), a biological nanofiber, has opened a new route for disease diagnosis and therapy as well as proteomics. Earlier phage display was widely used in protein-protein or antigen-antibody studies. In recent years, its application in nanomedicine is becoming increasingly popular and encouraging. We aim to review the current status in this research direction. For better understanding, we start with a brief introduction of basic biology and structure of the filamentous phage. We present the principle of phage display and library construction method on the basis of the filamentous phage. We summarize the use of the phage displayed peptide library for selecting peptides with high affinity against cells or tissues. We then review the recent applications of the selected cell or tissue targeting peptides in developing new targeting probes and therapeutics to advance the early diagnosis and targeted therapy of different diseases in nanomedicine. We also discuss the integration of antibody phage display and modern proteomics in discovering new biomarkers or target proteins for disease diagnosis and therapy. Finally, we propose an outlook for further advancing the potential impact of phage display on future nanomedicine. This article is categorized under: Biology-Inspired Nanomaterials > Protein and Virus-Based Structures.

Optical transparency and electrical conductivity of intermediate filaments in Müller cells and single-wall carbon nanotubes

Presently we investigated the electrical conductivity and optical transparency of Müller cell intermediate filaments. For comparison, the same properties were also explored in the model system of single-wall carbon nanotubes. We report the method of separation and purification of porcine (Sus scrofa domestica) intermediate filaments, extracted from the retinal Müller cells. We also report experimental and theoretical methods of measurements and calculations of the resistivity and light transmission yield by the intermediate filaments and single wall carbon nanotubes. The measured resistivity values were (4.7 ± 0.3) × 10−4 and (2.8 ± 0.2) × 10−4 Ω⋅m−1⋅cm2 at 5 °C (278 K), for the intermediate filaments and single wall carbon nanotubes, respectively, being quite close to those of typical metals. We report a method for measuring the light energy transmission by these nanostructures. We found that they efficiently transfer excitation energy along their axis, with the light reemitted at their other end. The measured yields of transferred light energy were 0.50 ± 0.03 and 0.26 ± 0.02 for intermediate filaments and single wall carbon nanotubes, respectively (λexc = 546.1 nm; T = 288 K). The reported results are novel, providing a direct confirmation of the earlier proposed quantum mechanism of light energy transport in the inverted retina of vertebrates. Our data also show that Müller cell intermediate filaments, in addition to their cytoskeletal function, are capable of providing for the light energy transfer within the inverted retina. The data obtained enable a significant progress in our understanding of the high-contrast vision of the vertebrate eyes. The most important conclusion of the current study is the discovery of light energy propagation along natural biological nanofibers (intermediate filaments). This result is completely novel and unique, being reported for the first time.

Synthetic Biogenesis of Bacterial Amyloid Nanomaterials with Tunable Inorganic-Organic Interfaces and Electrical Conductivity.

Amyloids are highly ordered, hierarchal protein nanoassemblies. Functional amyloids in bacterial biofilms, such as Escherichia coli curli fibers, are formed by the polymerization of monomeric proteins secreted into the extracellular space. Curli is synthesized by living cells, is primarily composed of the major curlin subunit CsgA, and forms biological nanofibers with high aspect ratios. Here, we explore the application of curli fibers for nanotechnology by engineering curli to mediate tunable biological interfaces with inorganic materials and to controllably form gold nanoparticles and gold nanowires. Specifically, we used cell-synthesized curli fibers as templates for nucleating and growing gold nanoparticles and showed that nanoparticle size could be modulated as a function of curli fiber gold-binding affinity. Furthermore, we demonstrated that gold nanoparticles can be preseeded onto curli fibers and followed by gold enhancement to form nanowires. Using these two approaches, we created artificial cellular systems that integrate inorganic-organic materials to achieve tunable electrical conductivity. We envision that cell-synthesized amyloid nanofibers will be useful for interfacing abiotic and biotic systems to create living functional materials..

Tyrosinase-Mediated Construction of a Silk Fibroin/Elastin Nanofiber Bioscaffold.

Elastin has characteristics of elasticity, biological activity, and mechanical stability. In the present work, tyrosinase-mediated construction of a bioscaffold with silk fibroin and elastin was carried out, aiming at developing a novel medical biomaterial. The efficiency of enzymatic oxidation of silk fibroin and the covalent reaction between fibroin and elastin were examined by spectrophotometry, sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE), and size exclusion chromatography (SEC). The properties of composite air-dried and nanofiber scaffolds were investigated. The results reveal that elastin was successfully bonded to silk fibroins, resulting in an increase in molecular weight of fibroin proteins. ATR-FTIR spectra indicated that tyrosinase treatment impacted the conformational structure of fibroin-based membrane. The thermal behaviors and mechanical properties of the tyrosinase-treated scaffolds were also improved compared with the untreated group. NIH/3T3 cells exhibited optimum densities when grown on the nanofiber scaffold, implying that the nanofiber scaffold has enhanced biocompatibility compared to the air-dried scaffold. A biological nanofiber scaffold constructed from tyrosinase-treated fibroin and elastin could potentially be utilized in biomedical applications.

Supramolecular assembly of asymmetric self-neutralizing amphiphilic peptide wedges.

Mimicking the remarkable dynamic and multifunctional utility of biological nanofibers, such as microtubules, is a challenging and technologically attractive objective in synthetic supramolecular chemistry. Understanding the complex molecular interactions that govern the assembly of synthetic materials, such as peptides, is key to meeting this challenge. Using molecular dynamics simulations to guide molecular design, we explore here the self-assembly of structurally and functionally asymmetric wedge-shaped peptides. Supramolecular assembly into nanofiber gels or multilayered lamellar structures was determined by cooperative influences of hydrogen bonding, amphiphilicity (hydrophilic asymmetry), and the distribution of electrostatic charges on the aqueous self-assembly of asymmetric peptides. Molecular amphiphilicity and β-sheet forming capacity were both identified as necessary, but not independently sufficient, to form supramolecular nanofibers. Imbalances in positive and negative charges prevented nanofiber assembly, while the asymmetric distribution of balanced charges within a peptide is believed to affect peptide conformation and subsequent self-assembly into either nanofibers or lamellar structures. Insights into cooperative molecular interactions and the effects of molecular asymmetry on assembly may aid the development of next-generation supramolecular nanomaterial assemblies.

Hydrodynamic Helical Orientations of Nanofibers in a Vortex

In this review article, I report our recent studies on spectroscopic visualizations of macroscopic helical alignments of nanofibers in vortex flows. Our designed supramolecular nanofibers, formed through self-assemblies of dye molecules, helically align in torsional flows of a vortex generated by mechanical rotary stirring of the sample solutions. The nanofiber, formed through bundling of linear supramolecular polymers, aligns equally in right- and left-handed vortex flows. However, in contrast, a one-handedly twisted nanofiber, formed through helical bundling of the supramolecular polymers, shows unequal helical alignments in these torsional flows. When the helical handedness of the nanofiber matches that of the vortex flow, the nanofiber aligns more efficiently in the flowing fluid. Such phenomena are observed not only with the artificial helical supramolecular nanofibers but also with biological nanofibers such as double-stranded DNA.

Controlled alignment of filamentous supramolecular assemblies of biomolecules into centimeter-scale highly ordered patterns by using nature-inspired magnetic guidance.

Magnetic nanoparticles (MNPs), which can be aligned along a magnetic field, were applied to generate large-scale assemblies of biological nanofibers with the orientation of the constituent nanofibers defined by the applied magnetic field. When decorated with MNPs, these nanofibers could be guided by the external magnetic field to become oriented either horizontally or vertically.

Fabrication of Fibrin Based Electrospun Multiscale Composite Scaffold for Tissue Engineering Applications

Fabricating scaffolds mimicking the native extracellular matrix (ECM) in both structure and function is a key challenge in the field of tissue engineering. Previously we have demonstrated a novel electrospinnig method for the fabrication of fibrin nanofibers using Poly(vinyl alcohol) (PVA) as an 'electrospinning-driving' polymer. Here we demonstrate the fabrication and characterization of a multiscale fibrin based composite scaffold with polycaprolactone (PCL) by sequential electrospinning of PCL microfibers and fibrin nanofibers. This multiscale scaffold has great potential for tissue engineering applications due to the combined benefits of biological nanofibers such as cell attachment and proliferation and that of microfibers such as open structure, larger pore size and adequate mechanical strength. Physico chemical characterization of the electrospun scaffold was done by Scanning Electron Microscopy (SEM), Contact angle analysis, fibrin specific Phosphotungstic acid haematoxyllin (PTAH) staining and evaluation of mechanical properties. SEM data revealed the formation of bead free nanofibers of fibrin with a fiber diameter ranging from 50-500 nm and microfibers of PCL in the size range of 1 microns to 2.5 microns. These dimensions mimic the hierarchical structure of ECM found in native tissues. Cell attachment and viability studies using human mesenchymal stem cells (hMSC) revealed that the scaffold is non toxic and supports cell attachment, spreading and proliferation. In addition, we examined the inflammatory potential of the scaffold to demonstrate its usefulness in tissue engineering applications.

Self-Assembly and Mineralization of Genetically Modifiable Biological Nanofibers Driven by β-Structure Formation

Bioinspired mineralization is an innovative approach to the fabrication of bone biomaterials mimicking the natural bone. Bone mineral hydroxylapatite (HAP) is preferentially oriented with c-axis parallel to collagen fibers in natural bone. However, such orientation control is not easy to achieve in artificial bone biomaterials. To overcome the lack of such orientation control, we fabricated a phage-HAP composite by genetically engineering M13 phage, a nontoxic bionanofiber, with two HAP-nucleating peptides derived from one of the noncollagenous proteins, Dentin Matrix Protein-1 (DMP1). The phage is a biological nanofiber that can be mass produced by infecting bacteria and is nontoxic to human beings. The resultant HAP-nucleating phages are able to self-assemble into bundles by forming β-structure between the peptides displayed on their side walls. The β-structure further promotes the oriented nucleation and growth of HAP crystals within the nanofibrous phage bundles with their c-axis preferentially parallel to the bundles. We proposed that the preferred orientation resulted from the stereochemical matching between the negatively charged amino acid residues within the β-structure and the positively charged calcium ions on the (001) plane of HAP crystals. The self-assembly and mineralization driven by the β-structure formation represent a new route for fabricating mineralized fibers that can serve as building blocks in forming bone repair biomaterials and mimic the basic structure of natural bones.

Controlled growth and differentiation of MSCs on grooved films assembled from monodisperse biological nanofibers with genetically tunable surface chemistries

The search for a cell-supporting scaffold with controlled topography and surface chemistry is a constant topic within tissue engineering. Here we have employed M13 phages, which are genetically modifiable biological nanofibers (∼880 nm long and ∼6.6 nm wide) non-toxic to human beings, to form films for supporting the growth of mesencymal stem cells (MSCs). Films were built from nearly parallel phage bundles separated by grooves. The bundles can guide the elongation and alignment of MSCs along themselves. Phage with peptides displayed on the surface exhibited different control over the fine morphologies and differentiation of the MSCs. When an osteogenic peptide was displayed on the surface of phage, the proliferation and differentiation of MSCs into osteoblasts were significantly accelerated. The use of the grooved phage films allows us to control the proliferation and differentiation of MSCs by simply controlling the concentrations of phages as well as the peptides displayed on the surface of the phages. This work will advance our understanding on the interaction between stem cells and proteins.