Silkworm silk-based materials and devices generated using bio-nanotechnology.

PubMed

Huang, Wenwen; Ling, Shengjie; Li, Chunmei; Omenetto, Fiorenzo G; Kaplan, David L

2018-06-25

Silks are natural fibrous protein polymers that are spun by silkworms and spiders. Among silk variants, there has been increasing interest devoted to the silkworm silk of B. mori, due to its availability in large quantities along with its unique material properties. Silk fibroin can be extracted from the cocoons of the B. mori silkworm and combined synergistically with other biomaterials to form biopolymer composites. With the development of recombinant DNA technology, silks can also be rationally designed and synthesized via genetic control. Silk proteins can be processed in aqueous environments into various material formats including films, sponges, electrospun mats and hydrogels. The versatility and sustainability of silk-based materials provides an impressive toolbox for tailoring materials to meet specific applications via eco-friendly approaches. Historically, silkworm silk has been used by the textile industry for thousands of years due to its excellent physical properties, such as lightweight, high mechanical strength, flexibility, and luster. Recently, due to these properties, along with its biocompatibility, biodegradability and non-immunogenicity, silkworm silk has become a candidate for biomedical utility. Further, the FDA has approved silk medical devices for sutures and as a support structure during reconstructive surgery. With increasing needs for implantable and degradable devices, silkworm silk has attracted interest for electronics, photonics for implantable yet degradable medical devices, along with a broader range of utility in different device applications. This Tutorial review summarizes and highlights recent advances in the use of silk-based materials in bio-nanotechnology, with a focus on the fabrication and functionalization methods for in vitro and in vivo applications in the field of tissue engineering, degradable devices and controlled release systems.

Smart assembly of polymer fibers: lessons from major ampullate spider silk

NASA Astrophysics Data System (ADS)

Viney, Christopher

1996-02-01

Studies of major ampullate silk (MAS), especially the secretions and fibers produced by the spider Nephila clavipes (golden orb weaver), have yielded several results of potential value to the materials scientist/engineer. There are lessons to be learned about synthesis, processing and microstructural design of high-tensile polymer fibers. The 'smart' aspect of silk production in nature concerns the ability of the spider to rapidly process a concentrated, viscous aqueous solution of silk protein (stored in the gland) into water-insoluble fiber on demand. This process centers on the assembly of a shear-sensitive supramolecular liquid crystalline phase by aggregation of the solubilized globular protein molecules.

Self-assembly of silk-elastinlike protein polymers into three-dimensional scaffolds for biomedical applications

NASA Astrophysics Data System (ADS)

Zeng, Like

Production of brand new protein-based materials with precise control over the amino acid sequences at single residue level has been made possible by genetic engineering, through which artificial genes can be developed that encode protein-based materials with desired features. As an example, silk-elastinlike protein polymers (SELPs), composed of tandem repeats of amino acid sequence motifs from Bombyx mori (silkworm) silk and mammalian elastin, have been produced in this approach. SELPs have been studied extensively in the past two decades, however, the fundamental mechanism governing the self-assembly process to date still remains largely unresolved. Further, regardless of the unprecedented success when exploited in areas including drug delivery, gene therapy, and tissue augmentation, SELPs scaffolds as a three-dimensional cell culture model system are complicated by the inability of SELPs to provide the embedded tissue cells with appropriate biochemical stimuli essential for cell survival and function. In this dissertation, it is reported that the self-assembly of silk-elastinlike protein polymers (SELPs) into nanofibers in aqueous solutions can be modulated by tuning the curing temperature, the size of the silk blocks, and the charge of the elastin blocks. A core-sheath model was proposed for nanofiber formation, with the silk blocks in the cores and the hydrated elastin blocks in the sheaths. The folding of the silk blocks into stable cores -- affected by the size of the silk blocks and the charge of the elastin blocks -- plays a critical role in the assembly of silk-elastin nanofibers. The assembled nanofibers further form nanofiber clusters on the microscale, and the nanofiber clusters then coalesce into nanofiber micro-assemblies, interconnection of which eventually leads to the formation of three-dimensional scaffolds with distinct nanoscale and microscale features. SELP-Collagen hybrid scaffolds were also fabricated to enable independent control over the scaffolds' biochemical input and matrix stiffness. It is reported herein that in the hybrid scaffolds, collagen provides essential biochemical cues needed to promote cell attachment and function while SELP imparts matrix stiffness tunability. To obtain tissue-specificity in matrix stiffness that spans over several orders of magnitude covering from soft brain to stiff cartilage, the hybrid SELP-Collagen scaffolds were crosslinked by transglutaminase at physiological conditions compatible for simultaneous cell encapsulation. The effect of the increase in matrix stiffness induced by such enzymatic crosslinking on cellular viability and proliferation was also evaluated using in vitro cell assays.

Silk Fibroin Aqueous-Based Adhesives Inspired by Mussel Adhesive Proteins.

PubMed

Burke, Kelly A; Roberts, Dane C; Kaplan, David L

2016-01-11

Silk fibroin from the domesticated silkworm Bombyx mori is a naturally occurring biopolymer with charged hydrophilic terminal regions that end-cap a hydrophobic core consisting of repeating sequences of glycine, alanine, and serine residues. Taking inspiration from mussels that produce proteins rich in L-3,4-dihydroxyphenylalanine (DOPA) to adhere to a variety of organic and inorganic surfaces, the silk fibroin was functionalized with catechol groups. Silk fibroin was selected for its high molecular weight, tunable mechanical and degradation properties, aqueous processability, and wide availability. The synthesis of catechol-functionalized silk fibroin polymers containing varying amounts of hydrophilic polyethylene glycol (PEG, 5000 g/mol) side chains was carried out to balance silk hydrophobicity with PEG hydrophilicity. The efficiency of the catechol functionalization reaction did not vary with PEG conjugation over the range studied, although tuning the amount of PEG conjugated was essential for aqueous solubility. Adhesive bonding and cell compatibility of the resulting materials were investigated, where it was found that incorporating as little as 6 wt % PEG prior to catechol functionalization resulted in complete aqueous solubility of the catechol conjugates and increased adhesive strength compared with silk lacking catechol functionalization. Furthermore, PEG-silk fibroin conjugates maintained their ability to form β-sheet secondary structures, which can be exploited to reduce swelling. Human mesenchymal stem cells (hMSCs) proliferated on the silks, regardless of PEG and catechol conjugation. These materials represent a protein-based approach to catechol-based adhesives, which we envision may find applicability as biodegradable adhesives and sealants.

Synthetic Engineering of Spider Silk Fiber as Implantable Optical Waveguides for Low-Loss Light Guiding.

PubMed

Qiao, Xin; Qian, Zhigang; Li, Junjie; Sun, Hongji; Han, Yao; Xia, Xiaoxia; Zhou, Jin; Wang, Chunlan; Wang, Yan; Wang, Changyong

2017-05-03

A variety of devices used for biomedical engineering have been fabricated using protein polymer because of their excellent properties, such as strength, toughness, biocompatibility, and biodegradability. In this study, we fabricated an optical waveguide using genetically engineered spider silk protein. This method has two significant advantages: (1) recombinant spider silk optical waveguide exhibits excellent optical and biological properties and (2) biosynthesis of spider silk protein can overcome the limitation to the research on spider silk optical waveguide due to the low yield of natural spider silk. In detail, two kinds of protein-based optical waveguides made from recombinant spider silk protein and regenerative silkworm silk protein were successfully prepared. Results suggested that the recombinant spider silk optical waveguide showed a smoother surface and a higher refractive index when compared with regenerative silkworm silk protein. The optical loss of recombinant spider silk optical waveguide was 0.8 ± 0.1 dB/cm in air and 1.9 ± 0.3 dB/cm in mouse muscles, which were significantly lower than those of regenerative silkworm silk optical waveguide. Moreover, recombinant spider silk optical waveguide can meet the demand to guide and efficiently deliver light through biological tissue. In addition, recombinant spider silk optical waveguide showed low toxicity to cells in vitro and low-level inflammatory reaction with surrounding tissue in vivo. Therefore, recombinant spider silk optical waveguide is a promising implantable device to guide and deliver light with low loss.

Hybrid scaffolds based on PLGA and silk for bone tissue engineering.

PubMed

Sheikh, Faheem A; Ju, Hyung Woo; Moon, Bo Mi; Lee, Ok Joo; Kim, Jung-Ho; Park, Hyun Jung; Kim, Dong Wook; Kim, Dong-Kyu; Jang, Ji Eun; Khang, Gilson; Park, Chan Hum

2016-03-01

Porous silk scaffolds, which are considered to be natural polymers, cannot be used alone because they have a long degradation rate, which makes it difficult for them to be replaced by the surrounding tissue. Scaffolds composed of synthetic polymers, such as PLGA, have a short degradation rate, lack hydrophilicity and their release of toxic by-products makes them difficult to use. The present investigations aimed to study hybrid scaffolds fabricated from PLGA, silk and hydroxyapatite nanoparticles (Hap NPs) for optimized bone tissue engineering. The results from variable-pressure field emission scanning electron microscopy (VP-FE-SEM), equipped with EDS, confirmed that the fabricated scaffolds had a porous architecture, and the location of each component present in the scaffolds was examined. Contact angle measurements confirmed that the introduction of silk and HAp NPs helped to change the hydrophobic nature of PLGA to hydrophilic, which is the main constraint for PLGA used as a biomaterial. Thermo-gravimetric analysis (TGA) and FT-IR spectroscopy confirmed thermal decomposition and different vibrations caused in functional groups of compounds used to fabricate the scaffolds, which reflected improvement in their mechanical properties. After culturing osteoblasts for 1, 7 and 14 days in the presence of scaffolds, their viability was checked by MTT assay. The fluorescent microscopy results revealed that the introduction of silk and HAp NPs had a favourable impact on the infiltration of osteoblasts. In vivo experiments were conducted by implanting scaffolds in rat calvariae for 4 weeks. Histological examinations and micro-CT scans from these experiments revealed beneficial attributes offered by silk fibroin and HAp NPs to PLGA-based scaffolds for bone induction. Copyright © 2015 John Wiley & Sons, Ltd.

Phase separation and mechanical properties of an elastomeric biomaterial from spider wrapping silk and elastin block copolymers.

PubMed

Muiznieks, Lisa D; Keeley, Fred W

2016-10-01

Elastin and silk spidroins are fibrous, structural proteins with elastomeric properties of extension and recoil. While elastin is highly extensible and has excellent recovery of elastic energy, silks are particularly strong and tough. This study describes the biophysical characterization of recombinant polypeptides designed by combining spider wrapping silk and elastin-like sequences as a strategy to rationally increase the strength of elastin-based materials while maintaining extensibility. We demonstrate a thermo-responsive phase separation and spontaneous colloid-like droplet formation from silk-elastin block copolymers, and from a 34 residue disordered region of Argiope trifasciata wrapping silk alone, and measure a comprehensive suite of tensile mechanical properties from cross-linked materials. Silk-elastin materials exhibited significantly increased strength, toughness, and stiffness compared to an elastin-only material, while retaining high failure strains and low energy loss upon recoil. These data demonstrate the mechanical tunability of protein polymer biomaterials through modular, chimeric recombination, and provide structural insights into mechanical design. © 2016 Wiley Periodicals, Inc. Biopolymers 105: 693-703, 2016. © 2016 Wiley Periodicals, Inc.

Functionalized hybrid nanofibers to mimic native ECM for tissue engineering applications

NASA Astrophysics Data System (ADS)

Karuppuswamy, Priyadharsini; Venugopal, Jayarama Reddy; Navaneethan, Balchandar; Laiva, Ashang Luwang; Sridhar, Sreepathy; Ramakrishna, Seeram

2014-12-01

Nanotechnology being one of the most promising technologies today shows an extremely huge potential in the field of tissue engineering to mimic the porous topography of natural extracellular matrix (ECM). Natural polymers are incorporated into the synthetic polymers to fabricate functionalized hybrid nanofibrous scaffolds, which improve cell and tissue compatibility. The present study identified the biopolymers - aloe vera, silk fibroin and curcumin incorporated into polycaprolactone (PCL) as suitable substrates for tissue engineering. Different combinations of PCL with natural polymers - PCL/aloe vera, PCL/silk fibroin, PCL/aloe vera/silk fibroin, PCL/aloe vera/silk fibroin/curcumin were electrospun into nanofibrous scaffolds. The fabricated two dimensional nanofibrous scaffolds showed high surface area, appropriate mechanical properties, hydrophilicity and porosity, required for the regeneration of diseased tissues. The nanofibrous scaffolds were characterized by Scanning electron microscope (SEM), porometry, Instron tensile tester, VCA optima contact angle measurement and FTIR to analyze the fiber diameter and morphology, porosity and pore size distribution, mechanical strength, wettability, chemical bonds and functional groups, respectively. The average fiber diameter of obtained fibers ranged from 250 nm to 350 nm and the tensile strength of PCL scaffolds at 4.49 MPa increased upto 8.3 MPa for PCL/silk fibroin scaffolds. Hydrophobicity of PCL decreased with the incorporation of natural polymers, especially for PCL/aloe vera scaffolds. The properties of as-spun nanofiber scaffolds showed their potential as promising scaffold materials in tissue engineering applications.

Hydrophobic drug-triggered self-assembly of nanoparticles from silk-elastin-like protein polymers for drug delivery.

PubMed

Xia, Xiao-Xia; Wang, Ming; Lin, Yinan; Xu, Qiaobing; Kaplan, David L

2014-03-10

Silk-elastin-like protein polymers (SELPs) combine the mechanical and biological properties of silk and elastin. These properties have led to the development of various SELP-based materials for drug delivery. However, SELPs have rarely been developed into nanoparticles, partially due to the complicated fabrication procedures, nor assessed for potential as an anticancer drug delivery system. We have recently constructed a series of SELPs (SE8Y, S2E8Y, and S4E8Y) with various ratios of silk to elastin blocks and described their capacity to form micellar-like nanoparticles upon thermal triggering. In this study, we demonstrate that doxorubicin, a hydrophobic antitumor drug, can efficiently trigger the self-assembly of SE8Y (SELPs with silk to elastin ratio of 1:8) into uniform micellar-like nanoparticles. The drug can be loaded in the SE8Y nanoparticles with an efficiency around 6.5% (65 ng doxorubicin/μg SE8Y), S2E8Y with 6%, and S4E8Y with 4%, respectively. In vitro studies with HeLa cell lines demonstrate that the protein polymers are not cytotoxic (IC50 > 200 μg/mL), while the doxorubicin-loaded SE8Y nanoparticles showed a 1.8-fold higher cytotoxicity than the free drug. Confocal laser scanning microscopy (CLSM) and flow cytometry indicate significant uptake of the SE8Y nanoparticles by the cells and suggest internalization of the nanoparticles through endocytosis. This study provides an all-aqueous, facile method to prepare nanoscale, drug-loaded SELPs packages with potential for tumor cell treatments.

Silk fibroin/gold nanocrystals: a new example of biopolymer-based nanocomposites

NASA Astrophysics Data System (ADS)

Noinville, S.; Garnier, A.; Courty, A.

2017-05-01

The dispersion of nanoparticles in ordered polymer nanostructures can provide control over particle location and orientation, and pave the way for tailored nanomaterials that have enhanced mechanical, electrical, or optical properties. Here we used silk fibroin, a natural biopolymer, to embed gold nanocrystals (NCs), so as to obtain well-ordered structures such as nanowires and self-assembled triangular nanocomposites. Monodisperse gold NCs synthesized in organic media are mixed to silk fibroin and the obtained nanocomposites are characterized by UV-visible spectroscopy, transmission electron microscopy (TEM), scanning electron microscopy (FE-SEM), atomic force microscopy (AFM) and Infrared spectroscopy. The optical properties study of gold NCs and silk-gold nanocomposites shows that the Surface Plasmon band is blue shifted compared to gold NCs. The size and shape of NCs gold superlattices can be well controlled by the presence of silk fibroin giving nanowires and also self-assembled triangular nanocomposites as characterized by TEM, FE-SEM and AFM. The strong interaction between gold NCs and silk fibroin is also revealed by the conformation change of silk protein in presence of gold NCs, as shown by FTIR analysis. The formation of such ordered nanocomposites (gold NCs/silk fibroin) will provide new nanoplasmonic devices.

Silk scaffolds connected with different naturally occurring biomaterials for prostate cancer cell cultivation in 3D.

PubMed

Bäcker, Anne; Erhardt, Olga; Wietbrock, Lukas; Schel, Natalia; Göppert, Bettina; Dirschka, Marian; Abaffy, Paul; Sollich, Thomas; Cecilia, Angelica; Gruhl, Friederike J

2017-02-01

In the present work, different biopolymer blend scaffolds based on the silk protein fibroin from Bombyx mori (BM) were prepared via freeze-drying method. The chemical, structural, and mechanical properties of the three dimensional (3D) porous silk fibroin (SF) composite scaffolds of gelatin, collagen, and chitosan as well as SF from Antheraea pernyi (AP) and the recombinant spider silk protein spidroin (SSP1) have been systematically investigated, followed by cell culture experiments with epithelial prostate cancer cells (LNCaP) up to 14 days. Compared to the pure SF scaffold of BM, the blend scaffolds differ in porous morphology, elasticity, swelling behavior, and biochemical composition. The new composite scaffold with SSP1 showed an increased swelling degree and soft tissue like elastic properties. Whereas, in vitro cultivation of LNCaP cells demonstrated an increased growth behavior and spheroid formation within chitosan blended scaffolds based on its remarkable porosity, which supports nutrient supply matrix. Results of this study suggest that silk fibroin matrices are sufficient and certain SF composite scaffolds even improve 3D cell cultivation for prostate cancer research compared to matrices based on pure biomaterials or synthetic polymers. © 2016 Wiley Periodicals, Inc.

Effects of Fiber Reinforcement on Clay Aerogel Composites

PubMed Central

Finlay, Katherine A.; Gawryla, Matthew D.; Schiraldi, David A.

2015-01-01

Novel, low density structures which combine biologically-based fibers with clay aerogels are produced in an environmentally benign manner using water as solvent, and no additional processing chemicals. Three different reinforcing fibers, silk, soy silk, and hemp, are evaluated in combination with poly(vinyl alcohol) matrix polymer combined with montmorillonite clay. The mechanical properties of the aerogels are demonstrated to increase with reinforcing fiber length, in each case limited by a critical fiber length, beyond which mechanical properties decline due to maldistribution of filler, and disruption of the aerogel structure. Rather than the classical model for reinforced composite properties, the chemical compatibility of reinforcing fibers with the polymer/clay matrix dominated mechanical performance, along with the tendencies of the fibers to kink under compression. PMID:28793515

Osteoinductive recombinant silk fusion proteins for bone regeneration.

PubMed

Dinjaski, Nina; Plowright, Robyn; Zhou, Shun; Belton, David J; Perry, Carole C; Kaplan, David L

2017-02-01

Protein polymers provide a unique opportunity for tunable designs of material systems due to the genetic basis of sequence control. To address the challenge of biomineralization interfaces with protein based materials, we genetically engineered spider silks to design organic-inorganic hybrid systems. The spider silk inspired domain (SGRGGLGGQG AGAAAAAGGA GQGGYGGLGSQGT) 15 served as an organic scaffold to control material stability and to allow multiple modes of processing, whereas the hydroxyapatite binding domain VTKHLNQISQSY (VTK), provided control over osteogenesis. The VTK domain was fused either to the N-, C- or both terminals of the spider silk domain to understand the effect of position on material properties and mineralization. The addition of the VTK domain to silk did not affect the physical properties of the silk recombinant constructs, but it had a critical role in the induction of biomineralization. When the VTK domain was placed on both the C- and N-termini the formation of crystalline hydroxyapatite was significantly increased. In addition, all of the recombinant proteins in film format supported the growth and proliferation of human mesenchymal stem cells (hMSCs). Importantly, the presence of the VTK domain enhanced osteoinductive properties up to 3-fold compared to the control (silk alone without VTK). Therefore, silk-VTK fusion proteins have been shown suitable for mineralization and functionalization for specific biomedical applications. Organic-inorganic interfaces are integral to biomaterial functions in many areas of repair and regeneration. Several protein polymers have been investigated for this purpose. Despite their success the limited options to fine-tune their material properties, degradation patterns and functionalize them for each specific biomedical application limits their application. Various studies have shown that the biological performance of such proteins can be improved by genetic engineering. The present study provides data relating protein design parameters and functional outcome quantified by biomineralization and human mesenchymal stem cell differentiation. As such, it helps the design of osteoinductive recombinant biomaterials for bone regeneration. Copyright © 2016 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

Processing, Properties and Morphology of Optical Limiting Silk Membranes

DTIC Science & Technology

2006-02-28

Christopher Brewer, Donald Ott, Polymer, 45 8451 (2004). 2. Electrospun Bombyx mori Gland Silk, S. Putthanarat, R.K. Eby, W. Kataphinan, S. Jones, R...Science, The University of Akron, Akron, OH. 3. American Physical Society (APS) meeting, March 2005, "Electrospun liquid silk from the gland of Bombyx Mori ...provide a good compatible matrix. They have previously demonstrated that GFP molecules can be incorporated into cast membranes of silk from B. mori

Integration of Natural Polymers and Synthetic Nanostructures

DTIC Science & Technology

2014-11-20

interphase reinforcement. Reconstituted silk fibroin from Bombyx Mori silkworm silks is an mass- production available source of the excellent biopolymeric ...we replaced traditional synthetic binders in GO papers with universal biopolymer “binder”-silk fibroin. The fabrication of GO-SF microfilms using ...selectively patterning its secondary structure using capillary transfer lithography (CTL) and solvent-assisted micro -contact molding (SAMIM) (Figure

Structure to function: Spider silk and human collagen

NASA Astrophysics Data System (ADS)

Rabotyagova, Olena S.

Nature has the ability to assemble a variety of simple molecules into complex functional structures with diverse properties. Collagens, silks and muscles fibers are some examples of fibrous proteins with self-assembling properties. One of the great challenges facing Science is to mimic these designs in Nature to find a way to construct molecules that are capable of organizing into functional supra-structures by self-assembly. In order to do so, a construction kit consisting of molecular building blocks along with a complete understanding on how to form functional materials is required. In this current research, the focus is on spider silk and collagen as fibrous protein-based biopolymers that can shed light on how to generate nanostructures through the complex process of self-assembly. Spider silk in fiber form offers a unique combination of high elasticity, toughness, and mechanical strength, along with biological compatibility and biodegrability. Spider silk is an example of a natural block copolymer, in which hydrophobic and hydrophilic blocks are linked together generating polymers that organize into functional materials with extraordinary properties. Since silks resemble synthetic block copolymer systems, we adopted the principles of block copolymer design from the synthetic polymer literature to build block copolymers based on spider silk sequences. Moreover, we consider spider silk to be an important model with which to study the relationships between structure and properties in our system. Thus, the first part of this work was dedicated to a novel family of spider silk block copolymers, where we generated a new family of functional spider silk-like block copolymers through recombinant DNA technology. To provide fundamental insight into relationships between peptide primary sequence, block composition, and block length and observed morphological and structural features, we used these bioengineered spider silk block copolymers to study secondary structure, morphological features and assembly. Aside from fundamental perspectives, we anticipate that these results will provide a blueprint for the design of precise materials for a range of potential applications such as controlled release devices, functional coatings, components of tissue regeneration materials and environmentally friendly polymers in future studies. In the second part of this work, human collagen type I was studied as another representative of the family of fibrous proteins. Collagen type I is the most abundant extracellular matrix protein in the human body, providing the basis for tissue structure and directing cellular functions. Collagen has a complex structural hierarchy, organized at different length scales, including the characteristic triple helical feature. In the present study we assessed the relationship between collagen structure (native vs. denatured) and sensitivity to UV radiation with a focus on changes in the primary structure, conformation, microstructure and material properties. Free radical reactions are involved in collagen degradation and a mechanism for UV-induced collagen degradation related to structure was proposed. The results from this study demonstrated the role of collagen supramolecular organization (triple helix) in the context of the effects of electromagnetic radiation on extracellular matrices. Owing to the fact that both silks and collagens are proteins that have found widespread interest for biomaterial related needs, we anticipate that the current studies will serve as a foundation for future biomaterial designs with controlled properties. Furthermore, fundamental insight into self-assembly and environmentally-2mediated degradation, will build a foundation for fundamental understanding of the remodeling and functions of these types of fibrous proteins in vivo and in vitro. This type of insight is essential for many areas of scientific inquiry, from drug delivery, to scaffolds for tissue engineering, and to the stability of materials in space.

Amorphous Silk Fibroin Membranes for Separation of CO2

NASA Technical Reports Server (NTRS)

Aberg, Christopher M.; Patel, Anand K.; Gil, Eun Seok; Spontak, Richard J.; Hagg, May-Britt

2009-01-01

Amorphous silk fibroin has shown promise as a polymeric material derivable from natural sources for making membranes for use in removing CO2 from mixed-gas streams. For most applications of silk fibroin, for purposes other than gas separation, this material is used in its highly crystalline, nearly natural form because this form has uncommonly high tensile strength. However, the crystalline phase of silk fibroin is impermeable, making it necessary to convert the material to amorphous form to obtain the high permeability needed for gas separation. Accordingly, one aspect of the present development is a process for generating amorphous silk fibroin by treating native silk fibroin in an aqueous methanol/salt solution. The resulting material remains self-standing and can be prepared as thin film suitable for permeation testing. The permeability of this material by pure CO2 has been found to be highly improved, and its mixed-gas permeability has been found to exceed the mixed-gas permeabilities of several ultrahigh-CO2-permeable synthetic polymers. Only one of the synthetic polymers poly(trimethylsilylpropyne) [PTMSP] may be more highly permeable by CO2. PTMSP becomes unstable with time, whereas amorphous silk should not, although at the time of this reporting this has not been conclusively proven.

Silk fibroin/chitosan thin film promotes osteogenic and adipogenic differentiation of rat bone marrow-derived mesenchymal stem cells.

PubMed

Li, Da-Wei; He, Jin; He, Feng-Li; Liu, Ya-Li; Liu, Yang-Yang; Ye, Ya-Jing; Deng, Xudong; Yin, Da-Chuan

2018-04-01

As a biodegradable polymer thin film, silk fibroin/chitosan composite film overcomes the defects of pure silk fibroin and chitosan films, respectively, and shows remarkable biocompatibility, appropriate hydrophilicity and mechanical properties. Silk fibroin/chitosan thin film can be used not only as metal implant coating for bone injury repair, but also as tissue engineering scaffold for skin, cornea, adipose, and other soft tissue injury repair. However, the biocompatibility of silk fibroin/chitosan thin film for mesenchymal stem cells, a kind of important seed cell of tissue engineering and regenerative medicine, is rarely reported. In this study, silk fibroin/chitosan film was prepared by solvent casting method, and the rat bone marrow-derived mesenchymal stem cells were cultured on the silk fibroin/chitosan thin film. Osteogenic and adipogenic differentiation of rat bone marrow-derived mesenchymal stem cells were induced, respectively. The proliferation ability, osteogenic and adipogenic differentiation abilities of rat bone marrow-derived mesenchymal stem cells were systematically compared between silk fibroin/chitosan thin film and polystyrene tissue culture plates. The results showed that silk fibroin/chitosan thin film not only provided a comparable environment for the growth and proliferation of rat bone marrow-derived mesenchymal stem cells but also promoted their osteogenic and adipogenic differentiation. This work provided information of rat bone marrow-derived mesenchymal stem cells behavior on silk fibroin/chitosan thin film and extended the application of silk fibroin/chitosan thin film. Based on the results, we suggested that the silk fibroin/chitosan thin film could be a promising material for tissue engineering of bone, cartilage, adipose, and skin.

Biomimetic Nanofibrillation in Two-Component Biopolymer Blends with Structural Analogs to Spider Silk

NASA Astrophysics Data System (ADS)

Xie, Lan; Xu, Huan; Li, Liang-Bin; Hsiao, Benjamin S.; Zhong, Gan-Ji; Li, Zhong-Ming

2016-10-01

Despite the enormous potential in bioinspired fabrication of high-strength structure by mimicking the spinning process of spider silk, currently accessible routes (e.g., microfluidic and electrospinning approaches) still have substantial function gaps in providing precision control over the nanofibrillar superstructure, crystalline morphology or molecular orientation. Here the concept of biomimetic nanofibrillation, by copying the spiders’ spinning principles, was conceived to build silk-mimicking hierarchies in two-phase biodegradable blends, strategically involving the stepwise integration of elongational shear and high-pressure shear. Phase separation confined on nanoscale, together with deformation of discrete phases and pre-alignment of polymer chains, was triggered in the elongational shear, conferring the readiness for direct nanofibrillation in the latter shearing stage. The orderly aligned nanofibrils, featuring an ultralow diameter of around 100 nm and the “rigid-soft” system crosslinked by nanocrystal domains like silk protein dopes, were secreted by fine nanochannels. The incorporation of multiscale silk-mimicking structures afforded exceptional combination of strength, ductility and toughness for the nanofibrillar polymer composites. The proposed spider spinning-mimicking strategy, offering the biomimetic function integration unattainable with current approaches, may prompt materials scientists to pursue biopolymer mimics of silk with high performance yet light weight.

Biomimetic Nanofibrillation in Two-Component Biopolymer Blends with Structural Analogs to Spider Silk.

PubMed

Xie, Lan; Xu, Huan; Li, Liang-Bin; Hsiao, Benjamin S; Zhong, Gan-Ji; Li, Zhong-Ming

2016-10-03

Despite the enormous potential in bioinspired fabrication of high-strength structure by mimicking the spinning process of spider silk, currently accessible routes (e.g., microfluidic and electrospinning approaches) still have substantial function gaps in providing precision control over the nanofibrillar superstructure, crystalline morphology or molecular orientation. Here the concept of biomimetic nanofibrillation, by copying the spiders' spinning principles, was conceived to build silk-mimicking hierarchies in two-phase biodegradable blends, strategically involving the stepwise integration of elongational shear and high-pressure shear. Phase separation confined on nanoscale, together with deformation of discrete phases and pre-alignment of polymer chains, was triggered in the elongational shear, conferring the readiness for direct nanofibrillation in the latter shearing stage. The orderly aligned nanofibrils, featuring an ultralow diameter of around 100 nm and the "rigid-soft" system crosslinked by nanocrystal domains like silk protein dopes, were secreted by fine nanochannels. The incorporation of multiscale silk-mimicking structures afforded exceptional combination of strength, ductility and toughness for the nanofibrillar polymer composites. The proposed spider spinning-mimicking strategy, offering the biomimetic function integration unattainable with current approaches, may prompt materials scientists to pursue biopolymer mimics of silk with high performance yet light weight.

Biomimetic Nanofibrillation in Two-Component Biopolymer Blends with Structural Analogs to Spider Silk

PubMed Central

Xie, Lan; Xu, Huan; Li, Liang-Bin; Hsiao, Benjamin S.; Zhong, Gan-Ji; Li, Zhong-Ming

2016-01-01

Despite the enormous potential in bioinspired fabrication of high-strength structure by mimicking the spinning process of spider silk, currently accessible routes (e.g., microfluidic and electrospinning approaches) still have substantial function gaps in providing precision control over the nanofibrillar superstructure, crystalline morphology or molecular orientation. Here the concept of biomimetic nanofibrillation, by copying the spiders’ spinning principles, was conceived to build silk-mimicking hierarchies in two-phase biodegradable blends, strategically involving the stepwise integration of elongational shear and high-pressure shear. Phase separation confined on nanoscale, together with deformation of discrete phases and pre-alignment of polymer chains, was triggered in the elongational shear, conferring the readiness for direct nanofibrillation in the latter shearing stage. The orderly aligned nanofibrils, featuring an ultralow diameter of around 100 nm and the “rigid−soft” system crosslinked by nanocrystal domains like silk protein dopes, were secreted by fine nanochannels. The incorporation of multiscale silk-mimicking structures afforded exceptional combination of strength, ductility and toughness for the nanofibrillar polymer composites. The proposed spider spinning-mimicking strategy, offering the biomimetic function integration unattainable with current approaches, may prompt materials scientists to pursue biopolymer mimics of silk with high performance yet light weight. PMID:27694989

Structure and applications of a temperature responsive recombinant protein hydrogel based on silk- and elastin-like amino acid motifs

NASA Astrophysics Data System (ADS)

Drummy, Lawrence; Tomczak, Melanie; Macauliffe, Joseph; Vaia, Richard; Naik, Rajesh

2008-03-01

Proteins form the main components of many natural materials, and they can be designed to offer tailored functionality and material properties. Silk elastin-like proteins (SELP)s come from a family of repeat sequence protein polymers based on Bombyx mori silk and mammalian elastin that are recombinantly expressed in E. coli. SELP gels are formed by heating the protein solutions in order to induce physical crosslinking of the silk β-sheet regions, they contain approximately 80-90% water by weight and they can be used for encapsulation of enzymes or nanoparticles. For example, horseradish peroxidase demonstrates added resistance to drying and heat treatment when encapsulated in the gel matrix. During gel formation, small angle X-ray scattering shows intensity increases in two distinct regions of reciprocal space, one reversible with temperature and one irreversible. By fitting the scattering data to a unified power-law/Gunier model, morphological parameters are extracted. The thermally reversible intensity changes are attributed to a hydrophilic/hydrophobic transition in the elastin segments, while the irreversible intensity change is due to the crystalline regions formed by the silk blocks.

Enhancing analysis of cells and proteins by fluorescence imaging on silk-based biomaterials: modulating the autofluorescence of silk.

PubMed

Neo, Puay Yong; Tan, Daryl Jian-An; Shi, Pujiang; Toh, Siew Lok; Goh, James Cho-Hong

2015-02-01

Silk is a versatile and established biomaterial for various tissue engineering purposes. However, it also exhibits strong autofluorescence signals-thereby hindering fluorescence imaging analysis of cells and proteins on silk-derived biomaterials. Sudan Black B (SB) is a lysochrome dye commonly used to stain lipids in histology. It has also been reported to be able to quench autofluorescence of tissues in histology and has been tested on artificial biomedical polymers in recent years. It was hypothesized that SB would exert similar quenching effects on silk, modulating the autofluorescence signals, and thereby enabling improved imaging analysis of cells and molecules of interests. The quenching effect of SB on the intrinsic fluorescence properties of silk and on commercial fluorescent dyes were first investigated in this study. SB was then incorporated into typical fluorescence-based staining protocols to study its effectiveness in improving fluorescence-based imaging of the cells and proteins residing with the silk-based biomaterials. Silk processed into various forms of biomaterials (e.g., films, sponges, fibers, and electrospun mats) was seeded with cells and cultured in vitro. At sacrificial time points, specimens were harvested, fixed, and prepared for fluorescence staining. SB, available commercially as a powder, was dissolved in 70% ethanol (0.3% [w/v]) to form staining solutions. SB treatment was introduced at the last step of typical immunofluorescence staining protocols for 15-120 min. For actin staining protocols by phalloidin toxin, SB staining solutions were added before and after permeabilization with Triton-X for 15-30 min. Results showed that ideal SB treatment duration is about 15 min. Apart from being able to suppress the autofluorescence of silk, this treatment duration was also not too long to adversely affect the fluorescent labeling probes used. The relative improvement brought about by SB treatment was most evident in the blue and green emission wavelengths compared with the red emission wavelength. This study has showed that the use of SB is a cost and time effective approach to enhance fluorescence-based imaging analyses of cell-seeded silk biomaterials, which otherwise would have been hindered by the unmodulated autofluorescence signals.

Controlled Release from Recombinant Polymers

PubMed Central

Price, Robert; Poursaid, Azadeh; Ghandehari, Hamidreza

2014-01-01

Recombinant polymers provide a high degree of molecular definition for correlating structure with function in controlled release. The wide array of amino acids available as building blocks for these materials lend many advantages including biorecognition, biodegradability, potential biocompatibility, and control over mechanical properties among other attributes. Genetic engineering and DNA manipulation techniques enable the optimization of structure for precise control over spatial and temporal release. Unlike the majority of chemical synthetic strategies used, recombinant DNA technology has allowed for the production of monodisperse polymers with specifically defined sequences. Several classes of recombinant polymers have been used for controlled drug delivery. These include, but are not limited to, elastin-like, silk-like, and silk-elastinlike proteins, as well as emerging cationic polymers for gene delivery. In this article, progress and prospects of recombinant polymers used in controlled release will be reviewed. PMID:24956486

The elaborate structure of spider silk

PubMed Central

Römer, Lin

2008-01-01

Biomaterials, having evolved over millions of years, often exceed man-made materials in their properties. Spider silk is one outstanding fibrous biomaterial which consists almost entirely of large proteins. Silk fibers have tensile strengths comparable to steel and some silks are nearly as elastic as rubber on a weight to weight basis. In combining these two properties, silks reveal a toughness that is two to three times that of synthetic fibers like Nylon or Kevlar. Spider silk is also antimicrobial, hypoallergenic and completely biodegradable. This article focuses on the structure-function relationship of the characterized highly repetitive spider silk spidroins and their conformational conversion from solution into fibers. Such knowedge is of crucial importance to understanding the intrinsic properties of spider silk and to get insight into the sophisticated assembly processes of silk proteins. This review further outlines recent progress in recombinant production of spider silk proteins and their assembly into distinct polymer materials as a basis for novel products. PMID:19221522

Structure and morphology of regenerated silk nano-fibers produced by electrospinning

NASA Astrophysics Data System (ADS)

Zarkoob, Shahrzad

The impressive physical and mechanical properties of natural silk fiberssp1 and the possibility of producing these proteins using biotechnology,sp2 have provided the impetus for recent efforts in both the biosynthesissp{3,4} and the spinning of these protein based biopolymers.sp{5,6,7} The question still remains: whether fibers spun from solutions with similar chemical makeup can produce fibers with similar structures and therefore with the possibility of improved properties. Since genetically engineered silk solutions were not readily available, the first objective of this project was to completely dissolve the Bombyx mori cocoon and the Nephila clavipes dragline silk while maintaining the molecular weight integrity of the polymer. The second objective was to develop a system for re-spinning from very small amount of the resulting silk solutions by the process of electrospinning. The third objective was, to produce regenerated silk fibers with diameters that are several orders of magnitude smaller than the original fibers, suitable for direct observation and analysis by transmission electron microscopy and electron diffraction. And finally, to compare these results to structural information obtained from natural (as spun by the organism) fibers to see if the regenerated solutions are able to form the same structure as the original fibers. Both types of silk fibers were successfully dissolved while maintaining the polymer integrity. Small quantities (25-50 mul) of these solutions were used to electrospin fibers with diameters ranging from 8nm-200nm. The fibers were observed by optical, scanning electron, and transmission electron microscopy. These nano fibers showed optical retardation, appeared to have a circular cross-section, and were dimensionally stable at temperatures above 280sp°C. Electron diffraction patterns of annealed electrospun fibers of B. mori and N. clavipes showed reflections, demonstrating orientational and semicrystalline order in the material comparable to natural silk. In addition, electron diffraction was also obtained form extended microtomed single dragline fibers of N. clavipes, and the d-spacings agreed well with thoes obtained from WAXD of dragline fiber bundles.

Bioconjugation of silk fibroin nanoparticles with enzyme and Peptide and their characterization.

PubMed

Wang, Fei; Zhang, Yu-Qing

2015-01-01

Bombyx mori silk fibroin is a type of protein-based polymer with unique characteristics that is widely used in the research and development of medical biomaterials. The degummed filament of silk fibroin can be dissolved in a highly concentrated salt solution. After desalination, the regenerated liquid silk fibroin (LSF) solution could be made into various forms of silk biomaterials, such as powder, fiber, film, porous matrix, 3D scaffold, and hydrogel, depending on its application. In this study, we mixed the liquid silk solution with enzymes, including oxidase and hydrolase, and rapidly injected the mixture into an excess of acetone. The enzyme retained most of its enzymatic activity and was also captured in silk fibroin nanoparticles (SFNs), which instantly formed via a configuration transition of the regenerated silk protein from a random coil and α-helix to a β-sheet. The resulting enzyme-captured SFNs displayed a fine crystal structure with a high activity recovery and good thermal stability. Moreover, the affinities of these modified enzymes to their substrate did not evidently suffer from the capture. When only the liquid silk solution was rapidly injected into acetone, the resulting globular SFNs with the same crystallinity were also a good carrier that was covalently conjugated to enzymes and insulin. Thus, silk protein nanoparticles are of potential value as an enzyme or peptide delivery system for the research and development of medical biomaterials. In this report, the bioconjugation of SFNs with glucose oxidase, superoxidase, β-glucosidase, L-asparaginase, neutral protease, and insulin and their characterization are described in detail. © 2015 Elsevier Inc. All rights reserved.

Control of silk microsphere formation using polyethylene glycol (PEG).

PubMed

Wu, Jianbing; Zheng, Zhaozhu; Li, Gang; Kaplan, David L; Wang, Xiaoqin

2016-07-15

A one step, rapid method to prepare silk microspheres was developed, with particle size controlled by the addition of polyethylene glycol (PEG). PEG molecular weight (4.0K-20.0KDa) and concentration (20-50wt%), as well as silk concentration (5-20wt%), were key factors that determined particle sizes varying in a range of 1-100μm. Addition of methanol to the PEG-silk combinations increased the content of crystalline β-sheet in the silk microspheres. To track the distribution and degradation of silk microspheres in vivo, 3-mercaptopropionic acid (MPA)-coated CdTe quantum dots (QDs) were physically entrapped in the silk microspheres. QDs tightly bound to the β-sheet domains of silk via hydrophobic interactions, with over 96% of the loaded QDs remaining in the silk microspheres after exhaustive extraction. The fluorescence of QDs-incorporated silk microspheres less stable in cell culture medium than in phosphate buffer solution (PBS) and water. After subcutaneous injection in mice, microspheres prepared from 20% silk (approx. 30μm diameter particles) still fluoresced at 24h, while those prepared from 8% silk (approx. 4μm diameter particles) and free QDs were not detectable, reflecting the QDs quenching and particle size effect on microsphere clearance in vivo. The larger microspheres were more resistant to cell internalization and degradation. Since PEG is an FDA-approved polymer, and silk is FDA approved for some medical devices, the methods developed in the present study will be useful in a variety of biomedical applications where simple, rapid and scalable preparation of silk microspheres is required. The work is of significance to the biomaterial and controlled release society because it provides a new option for fabricating silk microspheres in one simple step of mixing silk and polyethylene glycol (PEG), with the size and properties of microspheres controllable by PEG molecular weight as well as PEG and silk concentrations. Although fabrication of silk microspheres have been reported previously using spray-drying, liposome-templating, polyvinyl alcohol (PVA) emulsification, etc., applications were hindered due to harsh conditions (temperature, solvents, etc.) and complicated procedures used as well as low yield and less controllable particle size (usually <10μm). Since PEG is an FDA-approved polymer, and silk is FDA approved for some medical devices, the methods developed in the present study will be useful in a variety of biomedical applications where simple, rapid and scalable preparation of silk microspheres is required. Copyright © 2016 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

Electrospun Silk Biomaterial Scaffolds for Regenerative Medicine

PubMed Central

Zhang, Xiaohui; Reagan, Michaela R; Kaplan, David L.

2009-01-01

Electrospinning is a versatile technique that enables the development of nanofiber-based biomaterial scaffolds. Scaffolds can be generated that are useful for tissue engineering and regenerative medicine since they mimic the nanoscale properties of certain fibrous components of the native extracellular matrix in tissues. Silk is a natural protein with excellent biocompatibility, remarkable mechanical properties as well as tailorable degradability. Integrating these protein polymer advantages with electrospinning results in scaffolds with combined biochemical, topographical and mechanical cues with versatility for a range of biomaterial, cell and tissue studies and applications. This review covers research related to electrospinning of silk, including process parameters, post treatment of the spun fibers, functionalization of nanofibers, and the potential applications for these material systems in regenerative medicine. Research challenges and future trends are also discussed. PMID:19643154

The development of a novel wound healing material, silk-elastin sponge.

PubMed

Kawabata, Shingo; Kawai, Katsuya; Somamoto, Satoshi; Noda, Kazuo; Matsuura, Yoshitaka; Nakamura, Yoko; Suzuki, Shigehiko

2017-12-01

Silk-elastin is a recombinant protein polymer with repeating units of silk and elastin blocks. This novel wound healing promoting material has the ability to self-assemble from a liquid to a gel. We have already reported that an aqueous solution of silk-elastin has the potential to accelerate wound healing; however, there are several problems in applying silk-elastin in the clinical setting. To solve these problems, we developed a silk-elastin sponge that is easy to use in the clinical setting. In the present study, we examined whether the wound healing effect of the silk-elastin sponge is equal to the aqueous solution of silk-elastin in vivo. The granulation tissue formation promoting effect of the silk-elastin sponge was equal to that of the aqueous solution the silk-elastin, as after application to the wound surface, the sponge was absorbed and dissolved by the exudate. At body temperature the silk-elastin then formed temperature gel. The silk-elastin gel that was obtained contained abundant cytokines from the exudate. We believe that silk-elastin sponge can be applied to various wounds that are difficult to treat with the aqueous solution.

Biocompatible silk-conducting polymer composite trilayer actuators

NASA Astrophysics Data System (ADS)

Fengel, Carly V.; Bradshaw, Nathan P.; Severt, Sean Y.; Murphy, Amanda R.; Leger, Janelle M.

2017-05-01

Biocompatible materials capable of controlled actuation are in high demand for use in biomedical applications such as dynamic tissue scaffolding, valves, and steerable surgical tools. Conducting polymer actuators are of interest because they operate in aqueous electrolytes at low voltages and can generate stresses similar to natural muscle. Recently, our group has demonstrated a composite material of silk and poly(pyrrole) (PPy) that is mechanically robust, made from biocompatible materials, and bends under an applied voltage when incorporated into a simple bilayer device architecture and actuated using a biologically relevant electrolyte. Here we present trilayer devices composed of two silk-PPy composite layers separated by an insulating silk layer. The trilayer architecture allows one side to expand while the other contracts, resulting in improved performance over bilayer devices. Specifically, this configuration shows a larger angle of deflection per volt applied than the analogous bilayer system, while maintaining a consistent current response throughout cycling. In addition, the overall motion of the trilayer devices is more symmetric than that of the bilayer analogs, allowing for fully reversible operation.

Bioprospecting Finds the Toughest Biological Material: Extraordinary Silk from a Giant Riverine Orb Spider

PubMed Central

Agnarsson, Ingi; Kuntner, Matjaž; Blackledge, Todd A.

2010-01-01

Background Combining high strength and elasticity, spider silks are exceptionally tough, i.e., able to absorb massive kinetic energy before breaking. Spider silk is therefore a model polymer for development of high performance biomimetic fibers. There are over 41.000 described species of spiders, most spinning multiple types of silk. Thus we have available some 200.000+ unique silks that may cover an amazing breadth of material properties. To date, however, silks from only a few tens of species have been characterized, most chosen haphazardly as model organisms (Nephila) or simply from researchers' backyards. Are we limited to ‘blindly fishing’ in efforts to discover extraordinary silks? Or, could scientists use ecology to predict which species are likely to spin silks exhibiting exceptional performance properties? Methodology We examined the biomechanical properties of silk produced by the remarkable Malagasy ‘Darwin's bark spider’ (Caerostris darwini), which we predicted would produce exceptional silk based upon its amazing web. The spider constructs its giant orb web (up to 2.8 m2) suspended above streams, rivers, and lakes. It attaches the web to substrates on each riverbank by anchor threads as long as 25 meters. Dragline silk from both Caerostris webs and forcibly pulled silk, exhibits an extraordinary combination of high tensile strength and elasticity previously unknown for spider silk. The toughness of forcibly silked fibers averages 350 MJ/m3, with some samples reaching 520 MJ/m3. Thus, C. darwini silk is more than twice tougher than any previously described silk, and over 10 times better than Kevlar®. Caerostris capture spiral silk is similarly exceptionally tough. Conclusions Caerostris darwini produces the toughest known biomaterial. We hypothesize that this extraordinary toughness coevolved with the unusual ecology and web architecture of these spiders, decreasing the likelihood of bridgelines breaking and collapsing the web into the river. This hypothesis predicts that rapid change in material properties of silk co-occurred with ecological shifts within the genus, and can thus be tested by combining material science, behavioral observations, and phylogenetics. Our findings highlight the potential benefits of natural history–informed bioprospecting to discover silks, as well as other materials, with novel and exceptional properties to serve as models in biomimicry. PMID:20856804

Materials Fabrication from Bombyx mori Silk Fibroin

PubMed Central

Rockwood, Danielle N.; Preda, Rucsanda C.; Yücel, Tuna; Wang, Xiaoqin; Lovett, Michael L.; Kaplan, David L.

2013-01-01

Silk fibroin, derived from Bombyx mori cocoons, is a widely used and studied protein polymer for biomaterial applications. Silk fibroin has remarkable mechanical properties when formed into different materials, demonstrates biocompatibility, has controllable degradation rates from hours to years, and it can be chemically modified to alter surface properties or to immobilize growth factors. A variety of aqueous or organic solvent processing methods can be used to generate silk biomaterials for a range of applications. In this protocol we include methods to extract silk from B. mori cocoons in order to fabricate hydrogels, tubes, sponges, composites, fibers, microspheres and thin films. These materials can be used directly as biomaterials for implants, as scaffolding in tissue engineering and in vitro disease models, and for drug delivery. PMID:21959241

Silk sericin loaded alginate nanoparticles: Preparation and anti-inflammatory efficacy.

PubMed

Khampieng, Thitikan; Aramwit, Pornanong; Supaphol, Pitt

2015-09-01

In this study, silk sericin loaded alginate nanoparticles were prepared by the emulsification method followed by internal crosslinking. The effects of various silk sericin loading concentration on particle size, shape, thermal properties, and release characteristics were investigated. The initial silk sericin loadings of 20, 40, and 80% w/w to polymer were incorporated into these alginate nanoparticles. SEM images showed a spherical shape and small particles of about 71.30-89.50 nm. TGA analysis showed that thermal stability slightly increased with increasing silk sericin loadings. FTIR analysis suggested interactions between alginate and silk sericin in the nanoparticles. The release study was performed in acetate buffer at normal skin conditions (pH 5.5; 32 °C). The release profiles of silk sericin exhibited initial rapid release, consequently with sustained release. These silk sericin loaded alginate nanoparticles were further incorporated into topical hydrogel and their anti-inflammatory properties were studied using carrageenan-induced paw edema assay. The current study confirms the hypothesis that the application of silk sericin loaded alginate nanoparticle gel can inhibit inflammation induced by carrageenan. Copyright © 2015 Elsevier B.V. All rights reserved.

Structure-property relationships of Thai silk-microcrystalline cellulose biocomposite materials fabricated from ionic liquid.

PubMed

DeFrates, Kelsey; Markiewicz, Theodore; Callaway, Kayla; Xue, Ye; Stanton, John; Salas-de la Cruz, David; Hu, Xiao

2017-11-01

Biomaterials made from natural proteins and polysaccharides have become increasingly popular in the biomedical field due to their good biocompatibility and tunable biodegradability. However, the low miscibility of polysaccharides with proteins presents challenges in the creation of protein-polysaccharide composite materials. In this study, neat 1-allyl-3-methylimidazolium chloride (AMIMCl) ionic liquid was used to regenerate Thailand gold Bombyx mori silk and microcrystalline cellulose blended films. This solvent was found to not only effectively dissolve both natural polymers, but also preserve the structure and integrity of the polymers. A single glass transition temperature for each blend was found in DSC curves, indicating good miscibility between the Thai silk and cellulose molecules. The structural composition as well as the morphology and thermal stability of blend films were then determined using FTIR, SEM and TGA. It was found that by varying the ratio of Thai silk to cellulose, the thermal and physical properties of the material could be tuned. Blended films tended to be more thermally stable which could be due to the presence of hydrophobic-hydrophobic or electrostatic interactions between the silk and cellulose. These studies offered a new pathway to understand the tunable properties of protein-polysaccharide composite biomaterials with controllable physical and biological properties. Copyright © 2017 Elsevier B.V. All rights reserved.

Effects of the amino acid sequence on thermal conduction through β-sheet crystals of natural silk protein.

PubMed

Zhang, Lin; Bai, Zhitong; Ban, Heng; Liu, Ling

2015-11-21

Recent experiments have discovered very different thermal conductivities between the spider silk and the silkworm silk. Decoding the molecular mechanisms underpinning the distinct thermal properties may guide the rational design of synthetic silk materials and other biomaterials for multifunctionality and tunable properties. However, such an understanding is lacking, mainly due to the complex structure and phonon physics associated with the silk materials. Here, using non-equilibrium molecular dynamics, we demonstrate that the amino acid sequence plays a key role in the thermal conduction process through β-sheets, essential building blocks of natural silks and a variety of other biomaterials. Three representative β-sheet types, i.e. poly-A, poly-(GA), and poly-G, are shown to have distinct structural features and phonon dynamics leading to different thermal conductivities. A fundamental understanding of the sequence effects may stimulate the design and engineering of polymers and biopolymers for desired thermal properties.

The complexity of silk under the spotlight of synthetic biology.

PubMed

Vollrath, Fritz

2016-08-15

For centuries silkworm filaments have been the focus of R&D innovation centred on textile manufacture with high added value. Most recently, silk research has focused on more fundamental issues concerning bio-polymer structure-property-function relationships. This essay outlines the complexity and fundamentals of silk spinning, and presents arguments for establishing this substance as an interesting and important subject at the interface of systems biology (discovery) and synthetic biology (translation). It is argued that silk is a generic class of materials where each type of silk presents a different embodiment of emergent properties that combine genetically determined (anticipatory) and environmentally responsive components. In spiders' webs the various silks have evolved to form the interactive components of an intricate fabric that provides an extended phenotype to the spider's body morphology. © 2016 The Author(s). published by Portland Press Limited on behalf of the Biochemical Society.

Comparative evaluation of in vivo biocompatibility and biodegradability of regenerated silk scaffolds reinforced with/without natural silk fibers.

PubMed

Mobini, Sahba; Taghizadeh-Jahed, Masoud; Khanmohammadi, Manijeh; Moshiri, Ali; Naderi, Mohammad-Mehdi; Heidari-Vala, Hamed; Ashrafi Helan, Javad; Khanjani, Sayeh; Springer, Armin; Akhondi, Mohammad-Mehdi; Kazemnejad, Somaieh

2016-01-01

Nowadays, exceptional advantages of silk fibroin over synthetic and natural polymers have impelled the scientists to application of this biomaterial for tissue engineering purposes. Recently, we showed that embedding natural degummed silk fibers in regenerated Bombyx mori silk-based scaffold significantly increases the mechanical stiffness, while the porosity of the scaffolds remains the same. In the present study, we evaluated degradation rate, biocompatibility and regenerative properties of the regenerated 2% and 4% wt silk-based composite scaffolds with or without embedded natural degummed silk fibers within 90 days in both athymic nude and wild-type C57BL/6 mice through subcutaneous implantation. In all scaffolds, a suitable interconnected porous structure for cell penetration was seen under scanning electron microscopy. Compressive tests revealed a functional relationship between fiber reinforcement and compressive modulus. In addition, the fiber/fibroin composite scaffolds support cell attachment and proliferation. On days 30 to 90 after subcutaneous implantation, the retrieved tissues were examined via gross morphology, histopathology, immunofluorescence staining and reverse transcription-polymerase chain reaction as shown in Figure 1. Results showed that embedding the silk fibers within the matrix enhances the biodegradability of the matrix resulting in replacement of the composite scaffolds with the fresh connective tissue. Fortification of the composites with degummed fibers not only regulates the degradation profile but also increases the mechanical performance of the scaffolds. This report also confirmed that pore size and structure play an important role in the degradation rate. In conclusion, the findings of the present study narrate key role of additional surface area in improving in vitro and in vivo biological properties of the scaffolds and suggest the potential ability of these fabricated composite scaffolds for connective tissue regeneration. spjba;30/6/793/FIG10885328215601925F1fig1-0885328215601925Figure 1.Illustrative summary of the main methods and findings.RS: regenerated silk; RSF: regenerated fibroin/ silk fiber composite scaffolds; H&E: Hematoxylin and eosin; COX-1: Cyclooxygenase. © The Author(s) 2015.

Silk sericin: A versatile material for tissue engineering and drug delivery.

PubMed

Lamboni, Lallepak; Gauthier, Mario; Yang, Guang; Wang, Qun

2015-12-01

Sericin is an inexpensive glycoprotein obtained as a by-product in the silk industry. Its variable amino acid composition and diverse functional groups confer upon it attractive bioactive properties, which are particularly interesting for biomedical applications. Because of its antioxidant character, moisturizing ability, and mitogenic effect on mammalian cells, sericin is useful in cell culture and tissue engineering. Its positive effects on keratinocytes and fibroblasts have led to the development of sericin-based biomaterials for skin tissue repair, mainly as wound dressings. Additionally, sericin can be used for bone tissue engineering owing to its ability to induce nucleation of bone-like hydroxyapatite. Stable silk sericin biomaterials, such as films, sponges, and hydrogels, are prepared by cross-linking, ethanol precipitation, or blending with other polymers. Sericin may also be employed for drug delivery because its chemical reactivity and pH-responsiveness facilitate the fabrication of nano- and microparticles, hydrogels, and conjugated molecules, improving the bioactivity of drugs. Here, we summarized the recent advancements in the study of silk sericin for application in tissue engineering and drug delivery. Copyright © 2015 Elsevier Inc. All rights reserved.

Biomimetic Hybridization of Kevlar into Silk Fibroin: Nanofibrous Strategy for Improved Mechanic Properties of Flexible Composites and Filtration Membranes.

PubMed

Lv, Lili; Han, Xiangsheng; Zong, Lu; Li, Mingjie; You, Jun; Wu, Xiaochen; Li, Chaoxu

2017-08-22

Silk, one of the strongest natural biopolymers, was hybridized with Kevlar, one of the strongest synthetic polymers, through a biomimetic nanofibrous strategy. Regenerated silk materials have outstanding properties in transparency, biocompatibility, biodegradability and sustainability, and promising applications as diverse as in pharmaceutics, electronics, photonic devices and membranes. To compete with super mechanic properties of their natural counterpart, regenerated silk materials have been hybridized with inorganic fillers such as graphene and carbon nanotubes, but frequently lose essential mechanic flexibility. Inspired by the nanofibrous strategy of natural biomaterials (e.g., silk fibers, hemp and byssal threads of mussels) for fantastic mechanic properties, Kevlar was integrated in regenerated silk materials by combining nanometric fibrillation with proper hydrothermal treatments. The resultant hybrid films showed an ultimate stress and Young's modulus two times as high as those of pure regenerated SF films. This is not only because of the reinforcing effect of Kevlar nanofibrils, but also because of the increasing content of silk β-sheets. When introducing Kevlar nanofibrils into the membranes of silk nanofibrils assembled by regenerated silk fibroin, the improved mechanic properties further enabled potential applications as pressure-driven nanofiltration membranes and flexible substrates of electronic devices.

The effect of ageing on the mechanical properties of the silk of the bridge spider Larinioides cornutus (Clerck, 1757)

NASA Astrophysics Data System (ADS)

Lepore, Emiliano; Isaia, Marco; Mammola, Stefano; Pugno, Nicola

2016-05-01

Spider silk is regarded as one of the best natural polymer fibers especially in terms of low density, high tensile strength and high elongation until breaking. Since only a few bio-engineering studies have been focused on spider silk ageing, we conducted nano-tensile tests on the vertical naturally spun silk fibers of the bridge spider Larinioides cornutus (Clerck, 1757) (Arachnida, Araneae) to evaluate changes in the mechanical properties of the silk (ultimate stress and strain, Young’s modulus, toughness) over time. We studied the natural process of silk ageing at different time intervals from spinning (20 seconds up to one month), comparing silk fibers spun from adult spiders collected in the field. Data were analyzed using Linear Mixed Models. We detected a positive trend versus time for the Young’s modulus, indicating that aged silks are stiffer and possibly less effective in catching prey. Moreover, we observed a negative trend for the ultimate strain versus time, attesting a general decrement of the resistance force. These trends are interpreted as being due to the drying of the silk protein chains and the reorientation among the fibers.

Functionalized Silk Materials

DTIC Science & Technology

2010-06-10

properties, such as toughness, biocompatibility and biodegrability. Trends in spider silk-like block copolymer secondary structure and assembly behavior...to construct transistors on ultrathin sheets of polyimide . Briefly, the doped silicon nanomembranes were transfer printed onto a film of polyimide ...layer of polyimide was used to encapsulate the active devices. Dry etching the polymer layers completed the fabrication of an array of isolated

Silk scaffolds in bone tissue engineering: An overview.

PubMed

Bhattacharjee, Promita; Kundu, Banani; Naskar, Deboki; Kim, Hae-Won; Maiti, Tapas K; Bhattacharya, Debasis; Kundu, Subhas C

2017-11-01

Bone tissue plays multiple roles in our day-to-day functionality. The frequency of accidental bone damage and disorder is increasing worldwide. Moreover, as the world population continues to grow, the percentage of the elderly population continues to grow, which results in an increased number of bone degenerative diseases. This increased elderly population pushes the need for artificial bone implants that specifically employ biocompatible materials. A vast body of literature is available on the use of silk in bone tissue engineering. The current work presents an overview of this literature from materials and fabrication perspective. As silk is an easy-to-process biopolymer; this allows silk-based biomaterials to be molded into diverse forms and architectures, which further affects the degradability. This makes silk-based scaffolds suitable for treating a variety of bone reconstruction and regeneration objectives. Silk surfaces offer active sites that aid the mineralization and/or bonding of bioactive molecules that facilitate bone regeneration. Silk has also been blended with a variety of polymers and minerals to enhance its advantageous properties or introduce new ones. Several successful works, both in vitro and in vivo, have been reported using silk-based scaffolds to regenerate bone tissues or other parts of the skeletal system such as cartilage and ligament. A growing trend is observed toward the use of mineralized and nanofibrous scaffolds along with the development of technology that allows to control scaffold architecture, its biodegradability and the sustained releasing property of scaffolds. Further development of silk-based scaffolds for bone tissue engineering, taking them up to and beyond the stage of human trials, is hoped to be achieved in the near future through a cross-disciplinary coalition of tissue engineers, material scientists and manufacturing engineers. The state-of-art of silk biomaterials in bone tissue engineering, covering their wide applications as cell scaffolding matrices to micro-nano carriers for delivering bone growth factors and therapeutic molecules to diseased or damaged sites to facilitate bone regeneration, is emphasized here. The review rationalizes that the choice of silk protein as a biomaterial is not only because of its natural polymeric nature, mechanical robustness, flexibility and wide range of cell compatibility but also because of its ability to template the growth of hydroxyapatite, the chief inorganic component of bone mineral matrix, resulting in improved osteointegration. The discussion extends to the role of inorganic ions such as Si and Ca as matrix components in combination with silk to influence bone regrowth. The effect of ions or growth factor-loaded vehicle incorporation into regenerative matrix, nanotopography is also considered. Copyright © 2017 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

Molecular and macro-scale analysis of enzyme-crosslinked silk hydrogels for rational biomaterial design.

PubMed

McGill, Meghan; Coburn, Jeannine M; Partlow, Benjamin P; Mu, Xuan; Kaplan, David L

2017-11-01

Silk fibroin-based hydrogels have exciting applications in tissue engineering and therapeutic molecule delivery; however, their utility is dependent on their diffusive properties. The present study describes a molecular and macro-scale investigation of enzymatically-crosslinked silk fibroin hydrogels, and demonstrates that these systems have tunable crosslink density and diffusivity. We developed a liquid chromatography tandem mass spectroscopy (LC-MS/MS) method to assess the quantity and order of covalent tyrosine crosslinks in the hydrogels. This analysis revealed between 28 and 56% conversion of tyrosine to dityrosine, which was dependent on the silk concentration and reactant concentration. The crosslink density was then correlated with storage modulus, revealing that both crosslinking and protein concentration influenced the mechanical properties of the hydrogels. The diffusive properties of the bulk material were studied by fluorescence recovery after photobleaching (FRAP), which revealed a non-linear relationship between silk concentration and diffusivity. As a result of this work, a model for synthesizing hydrogels with known crosslink densities and diffusive properties has been established, enabling the rational design of silk hydrogels for biomedical applications. Hydrogels from naturally-derived silk polymers offer versitile opportunities in the biomedical field, however, their design has largely been an empirical process. We present a fundamental study of the crosslink density, storage modulus, and diffusion behavior of enzymatically-crosslinked silk hydrogels to better inform scaffold design. These studies revealed unexpected non-linear trends in the crosslink density and diffusivity of silk hydrogels with respect to protein concentration and crosslink reagent concentration. This work demonstrates the tunable diffusivity and crosslinking in silk fibroin hydrogels, and enables the rational design of biomaterials. Further, the characterization methods presented have applications for other materials with dityrosine crosslinks, which are found in nature as post-translational modificaitons, as well as in engineered matrices such as tyramine-substituted hyaluronic acid and recombinant resilin. Copyright © 2017 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

Nature's Mechanisms for Tough, Self-healing Polymers and Polymer Adhesives

NASA Astrophysics Data System (ADS)

Hansma, Paul

2007-03-01

Spider silk^2 and the natural polymer adhesives in abalone shells^3 and bone^4,5 can give us insights into nature's mechanisms for tough, self-healing polymers and polymer adhesives. The natural polymer adhesives in biomaterials have been optimized by evolution. An optimized polymer adhesive has five characteristics. 1) It holds together the strong elements of the composite. 2) It yields just before the strong elements would otherwise break. 3) It dissipates large amounts of energy as it yields. 4) It self heals after it yields. 5) It takes just a few percent by weight. Both natural polymer adhesives and silk rely on sacrificial bonds and hidden length for toughness and self-healing.^6 A relatively large energy, of order 100eV, is required to stretch a polymer molecule after a weak bond, a sacrificial bond, breaks and liberates hidden length, which was previously hidden, typically in a loop or folded domain, from whatever was stretching the polymer. The bond is called sacrificial if it breaks at forces well below the forces that could otherwise break the polymer backbone, typically greater than 1nN. In many biological cases, the breaking of sacrificial bonds has been found to be reversible, thereby also providing a ``self-healing'' property to the material.^2-4 Individual polymer adhesive molecules based on sacrificial bonds and hidden length can supply forces of order 300pN over distances of 100s of nanometers. Model calculations show that a few percent by weight of adhesives based on these principles could be optimized adhesives for high performance composite materials including nanotube and graphene sheet composites. ^2N. Becker, E. Oroudjev, S. Mutz et al., Nature Materials 2 (4), 278 (2003). ^3B. L. Smith, T. E. Schaffer, M. Viani et al., Nature 399 (6738), 761 (1999). ^4J. B. Thompson, J. H. Kindt, B. Drake et al., Nature 414 (6865), 773 (2001). ^5G. E. Fantner, T. Hassenkam, J. H. Kindt et al., Nature Materials 4, 612 (2005). ^6G. E. Fantner, E. Oroudjev, G. Schitter et al., Biophysical Journal 90 (4), 1411 (2006).

Microstructural Parameters in 8 MeV Electron-Irradiated BOMBYX MORI Silk Fibers by Wide-ANGLE X-Ray Scattering Studies (waxs)

NASA Astrophysics Data System (ADS)

Sangappa, Asha, S.; Sanjeev, Ganesh; Subramanya, G.; Parameswara, P.; Somashekar, R.

2010-01-01

The present work looks into the microstructural modification in electron irradiated Bombyx mori P31 silk fibers. The irradiation process was performed in air at room temperature using 8 MeV electron accelerator at different doses: 0, 25, 50 and 100 kGy. Irradiation of polymer is used to cross-link or degrade the desired component or to fix the polymer morphology. The changes in microstructural parameters in these natural polymer fibers have been computed using wide angle X-ray scattering (WAXS) data and employing line profile analysis (LPA) using Fourier transform technique of Warren. Exponential, Lognormal and Reinhold functions for the column length distributions have been used for the determination of crystal size, lattice strain and enthalpy parameters.

Influence of Layer-by-Layer Polyelectrolyte Deposition and EDC/NHS Activated Heparin Immobilization onto Silk Fibroin Fabric

PubMed Central

Elahi, M. Fazley; Guan, Guoping; Wang, Lu; King, Martin W.

2014-01-01

To enhance the hemocompatibility of silk fibroin fabric as biomedical material, polyelectrolytes architectures have been assembled through the layer-by-layer (LbL) technique on silk fibroin fabric (SFF). In particular, 1.5 and 2.5 bilayer of oppositely charged polyelectrolytes were assembled onto SFF using poly(allylamine hydrochloride) (PAH) as polycationic polymer and poly(acrylic acid) (PAA) as polyanionic polymer with PAH topmost. Low molecular weight heparin (LMWH) activated with 1-ethyl-3-(dimethylaminopropyl) carbodiimide hydrochloride (EDC) and N-hydroxysuccinimide (NHS) was then immobilized on its surface. Alcian Blue staining, toluidine blue assay and X-ray photoelectron spectroscopy (XPS) confirmed the presence of heparin on modified SFF surfaces. The surface morphology of the modified silk fibroin fabric surfaces was characterized by scanning electron microscopy (SEM) and atomic force microscopy (AFM), and obtained increased roughness. Negligible hemolytic effect and a higher concentration of free hemoglobin by a kinetic clotting time test ensured the improved biological performance of the modified fibroin fabric. Overall, the deposition of 2.5 bilayer was found effective in terms of biological and surface properties of the modified fibroin fabric compared to 1.5 bilayer self-assembly technique. Therefore, this novel approach to surface modification may demonstrate long term patency in future in vivo animal trials of small diameter silk fibroin vascular grafts. PMID:28788601

Silk and PEG as means to stiffen a parylene probe for insertion in the brain: toward a double time-scale tool for local drug delivery

NASA Astrophysics Data System (ADS)

Lecomte, A.; Castagnola, V.; Descamps, E.; Dahan, L.; Blatché, M. C.; Dinis, T. M.; Leclerc, E.; Egles, C.; Bergaud, C.

2015-12-01

The use of soft materials as substrate for neural probes aims at achieving better compliance with the surrounding neurons while maintaining minimal rejection. Many strategies have emerged to enable such probes to penetrate the cortex, among which the use of resorbable polymers. We performed several tests involving two resorbable polymers considered most promising: polyethylene glycol (PEG) and silk fibroin (SF) from Bombyx Mori silkworms. Our coating method provides a repeatable, uniform structure optimized for a stress-reduced insertion of a parylene-C neural probe. Standard compression tests as well as in vitro and in vivo insertion assessments show that both SF and PEG-coated probes are stiff enough to avoid the buckling effect during insertion in the cortex. However, with a buckling force of 300 mN and a mechanical holding in vitro of tens of minutes, we assess silk fibroin to be more reliable for practical handling. In vivo first try-outs in mouse brain showed neither buckling issues of the probe nor undesired alteration of the signal recording. Moreover, we evidenced two distinct time scales in the bioresorption of our polymer coatings: silk fibroin degrades itself in a matter of weeks and PEG dissolves itself within seconds in the presence of water. We then present a hybrid PEG and SF coating that could be used as a drug delivery system with different time scales to reduce both the acute and the chronic body reaction.

Electromechanical response of silk fibroin hydrogel and conductive polycarbazole/silk fibroin hydrogel composites as actuator material.

PubMed

Srisawasdi, Thanida; Petcharoen, Karat; Sirivat, Anuvat; Jamieson, Alexander M

2015-11-01

Pure silk fibroin (SF) hydrogel and polycarbazole/silk fibroin (SF/PCZ) hydrogels were fabricated by solvent casting technique to evaluate electromechanical responses, dielectric properties, and cantilever deflection properties as functions of electric field strength, SF concentration, glutaraldehyde concentration, and PCZ concentration in the blends. Electromechanical properties were characterized in oscillatory shear mode at electric field strengths ranging from 0 to 600V/mm and at a temperature of 27°C. For both the pristine SF and SF/PCZ hydrogels, the storage modulus response (ΔG') and the storage modulus sensitivity (ΔG'/G'0) increased dramatically with increasing electric field strength. The pristine hydrogel possessed the highest storage modulus sensitivity value of 5.87, a relatively high value when compared with other previously studied electroactive polymers. With the addition of conductive PCZ in SF hydrogel, the storage modulus sensitivity and the relative dielectric constant decreased; the conductive polymer thus provided the softening effect under electric field. In the deflection response, the dielectrophoresis force and deflection distance increased monotonically with electric field strength, where the pure SF hydrogel showed the highest deflection distance and dielectrophoresis force. Copyright © 2015 Elsevier B.V. All rights reserved.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lewis, Randolph

Spider silks have the potential to provide new bio-inspired materials for numerous applications in bioenergetics and products ranging from protective clothing to artificial ligaments and tendons. A number of spider silk genes have been cloned and sequenced by the Lewis laboratory revealing the basis for understanding the key elements of spider silk proteins with respect to their materials performance. In particular, specific amino acid motifs have been identified which have been conserved for over 125 million years in all spiders that use their silk to physically trap prey. The key element in taking the next step toward generating bio-based materialsmore » from spider silks will be to move from the current descriptive data to predictive knowledge. Current efforts are focused on mimicking spider silk through synthetic proteins. In developing synthetic silk fibers, we first need to understand the complete secondary and tertiary structure of natural silk so that we can compare synthetic constructs to the natural material. Being able to compare the structure on a single fiber level is critical to the future of molecular directed mimic development because we can vary mechanical properties by different spinning methods. The new generation of synchrotron x-ray diffraction and neutron beamlines will allow, for the first time, determination of the molecular structure of silk fibers and synthetic mimics. We propose an exciting new collaborative research team working jointly between Argonne National Laboratory, Arizona State U. and the University of Wyoming to address the ?characterization of synthetic and natural spider silk fibers using x-ray and neutron diffraction.? Thus these new methodologies will provide understanding of current fibers and determine changes needed to produce fibers with specific properties. The following specific aims are proposed: ? Synthesize spider silk fibers with molecular structures mimicking that of natural silks. Test the mechanic properties of these materials and compare them to natural silk fibers. ? Develop x-ray and neutron diffraction techniques to better determine the structure in amorphous and semicrystalline biopolymers, such as spider silk fibers. ? Combine mechanical testing and structural x-ray and neutron diffraction data to develop a molecular understanding of the structure-function relationship in spider silk materials. ? Elucidate the role water plays in spider silk fiber formation and structure. Emphasis will be placed on combined neutron and NMR studies. ? Use solid-state Nuclear Magnetic Resonance (NMR) to characterize synthetic and natural spider silk materials that show potential as a biomimetic material or bio-inspired polymer architecture. ? Develop EPSCoR student and postdoctoral training and exposure to national laboratory facilities. ? Further develop scientific outreach and chemical education programs and research.« less

In situ synthesis of silver nanoparticles uniformly distributed on polydopamine-coated silk fibers for antibacterial application.

PubMed

Lu, Zhisong; Xiao, Jing; Wang, Ying; Meng, Mei

2015-08-15

Fabrication of silver nanoparticles (AgNPs)-modified silk for antibacterial application is one of the hottest topics in the textile material research. However, the utilization of a polymer as both 3-dimensional matrix and reductant for the in-situ synthesis of AgNPs on silk fibers has not been realized. In this work, a facile, efficient and green approach was developed to in-situ grow AgNPs on the polydopamine (PDA)-functionalized silk. AgNPs with the size of 30-90 nm were uniformly deposited on the silk fiber surface with the PDA coating layer as a reduction reagent. The AgNPs exhibit excellent face-centered cubic crystalline structures. The bacterial growth curve and inhibition zone assays clearly demonstrate the antibacterial properties of the functionalized silk. Both high Ag(+) release level and long-time release profile were observed for the as-prepared AgNPs-PDA-coated silk, indicating the high-density loading of AgNPs and the possible long-term antibacterial effects. This work may provide a new method for the preparation of AgNPs-functionalized silk with antibacterial activity for the clothing and textile industry. Copyright © 2015 Elsevier Inc. All rights reserved.

Introducing biomimetic shear and ion gradients to microfluidic spinning improves silk fiber strength.

PubMed

Li, David; Jacobsen, Matthew M; Gyune Rim, Nae; Backman, Daniel; Kaplan, David L; Wong, Joyce Y

2017-05-31

Silkworm silk is an attractive biopolymer for biomedical applications due to its high mechanical strength and biocompatibility; as a result, there is increasing interest in scalable devices to spin silk and recombinant silk so as to improve and customize their properties for diverse biomedical purposes (Vepari and Kaplan 2007 Prog. Polym. Sci. 32 ). While artificial spinning of regenerated silk fibroins adds tunability to properties such as degradation rate and surface functionalization, the resulting fibers do not yet approach the mechanical strength of native silkworm silk. These drawbacks reduce the applicability and attractiveness of artificial silk (Kinahan et al 2011 Biomacromolecules 12 ). Here, we used computational fluid dynamic simulations to incorporate shear in tandem with biomimetic ion gradients by coupling a modular novel glass microfluidic device to our previous co-axial flow device. Fibers spun with this combined apparatus demonstrated a significant increase in mechanical strength compared to fibers spun with the basic apparatus alone, with a three-fold increase in Young's modulus and extensibility and a twelve-fold increase in toughness. These results thus demonstrate the critical importance of ionic milieu and shear stress in spinning strong fibers from solubilized silk fibroin.

Mechanical properties of silk of the Australian golden orb weavers Nephila pilipes and Nephilaplumipes.

PubMed

Kerr, Genevieve G; Nahrung, Helen F; Wiegand, Aaron; Kristoffersen, Joanna; Killen, Peter; Brown, Cameron; Macdonald, Joanne

2018-02-22

Silks from orb-weaving spiders are exceptionally tough, producing a model polymer for biomimetic fibre development. The mechanical properties of naturally spun silk threads from two species of Australian orb-weavers, Nephila pilipes and Nephila plumipes , were examined here in relation to overall thread diameter, the size and number of fibres within threads, and spider size. N. pilipes , the larger of the two species, had significantly tougher silk with higher strain capacity than its smaller congener, producing threads with average toughness of 150 MJ m -3 , despite thread diameter, mean fibre diameter and number of fibres per thread not differing significantly between the two species. Within N. pilipes , smaller silk fibres were produced by larger spiders, yielding tougher threads. In contrast, while spider size was correlated with thread diameter in N. plumipes , there were no clear patterns relating to silk toughness, which suggests that the differences in properties between the silk of the two species arise through differing molecular structure. Our results support previous studies that found that the mechanical properties of silk differ between distantly related spider species, and extends on that work to show that the mechanical and physical properties of silk from more closely related species can also differ remarkably. © 2018. Published by The Company of Biologists Ltd.

Silk-polypyrrole biocompatible actuator performance under biologically relevant conditions

NASA Astrophysics Data System (ADS)

Hagler, Jo'elen; Peterson, Ben; Murphy, Amanda; Leger, Janelle

Biocompatible actuators that are capable of controlled movement and can function under biologically relevant conditions are of significant interest in biomedical fields. Previously, we have demonstrated that a composite material of silk biopolymer and the conducting polymer polypyrrole (PPy) can be formed into a bilayer device that can bend under applied voltage. Further, these silk-PPy composites can generate forces comparable to human muscle (>0.1 MPa) making them ideal candidates for interfacing with biological tissues. Here silk-PPy composite films are tested for performance under biologically relevant conditions including exposure to a complex protein serum and biologically relevant temperatures. Free-end bending actuation performance, current response, force generation and, mass degradation were investigated . Preliminary results show that when exposed to proteins and biologically relevant temperatures, these silk-PPy composites show minimal degradation and are able to generate forces and conduct currents comparable to devices tested under standard conditions. NSF.

Native Silk Feedstock as a Model Biopolymer: A Rheological Perspective.

PubMed

Laity, Peter R; Holland, Chris

2016-08-08

Variability in silk's rheology is often regarded as an impediment to understanding or successfully copying the natural spinning process. We have previously reported such variability in unspun native silk extracted straight from the gland of the domesticated silkworm Bombyx mori and discounted classical explanations such as differences in molecular weight and concentration. We now report that variability in oscillatory measurements can be reduced onto a simple master-curve through normalizing with respect to the crossover. This remarkable result suggests that differences between silk feedstocks are rheologically simple and not as complex as originally thought. By comparison, solutions of poly(ethylene-oxide) and hydroxypropyl-methyl-cellulose showed similar normalization behavior; however, the resulting curves were broader than for silk, suggesting greater polydispersity in the (semi)synthetic materials. Thus, we conclude Nature may in fact produce polymer feedstocks that are more consistent than typical man-made counterparts as a model for future rheological investigations.

In Vivo Coating of Bacterial Magnetic Nanoparticles by Magnetosome Expression of Spider Silk-Inspired Peptides.

PubMed

Mickoleit, Frank; Borkner, Christian B; Toro-Nahuelpan, Mauricio; Herold, Heike M; Maier, Denis S; Plitzko, Jürgen M; Scheibel, Thomas; Schüler, Dirk

2018-03-12

Magnetosomes are natural magnetic nanoparticles with exceptional properties that are synthesized in magnetotactic bacteria by a highly regulated biomineralization process. Their usability in many applications could be further improved by encapsulation in biocompatible polymers. In this study, we explored the production of spider silk-inspired peptides on magnetosomes of the alphaproteobacterium Magnetospirillum gryphiswaldense. Genetic fusion of different silk sequence-like variants to abundant magnetosome membrane proteins enhanced magnetite biomineralization and caused the formation of a proteinaceous capsule, which increased the colloidal stability of isolated particles. Furthermore, we show that spider silk peptides fused to a magnetosome membrane protein can be used as seeds for silk fibril growth on the magnetosome surface. In summary, we demonstrate that the combination of two different biogenic materials generates a genetically encoded hybrid composite with engineerable new properties and enhanced potential for various applications.

Beating the Heat - Fast Scanning Melts Silk Beta Sheet Crystals

NASA Astrophysics Data System (ADS)

Cebe, Peggy; Hu, Xiao; Kaplan, David L.; Zhuravlev, Evgeny; Wurm, Andreas; Arbeiter, Daniela; Schick, Christoph

2013-01-01

Beta-pleated-sheet crystals are among the most stable of protein secondary structures, and are responsible for the remarkable physical properties of many fibrous proteins, such as silk, or proteins forming plaques as in Alzheimer's disease. Previous thinking, and the accepted paradigm, was that beta-pleated-sheet crystals in the dry solid state were so stable they would not melt upon input of heat energy alone. Here we overturn that assumption and demonstrate that beta-pleated-sheet crystals melt directly from the solid state to become random coils, helices, and turns. We use fast scanning chip calorimetry at 2,000 K/s and report the first reversible thermal melting of protein beta-pleated-sheet crystals, exemplified by silk fibroin. The similarity between thermal melting behavior of lamellar crystals of synthetic polymers and beta-pleated-sheet crystals is confirmed. Significance for controlling beta-pleated-sheet content during thermal processing of biomaterials, as well as towards disease therapies, is envisioned based on these new findings.

Electroresponsive Aqueous Silk Protein As “Smart” Mechanical Damping Fluid

PubMed Central

2015-01-01

Here we demonstrate the effectiveness of an electroresponsive aqueous silk protein polymer as a smart mechanical damping fluid. The aqueous polymer solution is liquid under ambient conditions, but is reversibly converted into a gel once subjected to an electric current, thereby increasing or decreasing in viscosity. This nontoxic, biodegradable, reversible, edible fluid also bonds to device surfaces and is demonstrated to reduce friction and provide striking wear protection. The friction and mechanical damping coefficients are shown to modulate with electric field exposure time and/or intensity. Damping coefficient can be modulated electrically, and then preserved without continued power for longer time scales than conventional “smart” fluid dampers. PMID:24750065

The mechanical design of spider silks: from fibroin sequence to mechanical function.

PubMed

Gosline, J M; Guerette, P A; Ortlepp, C S; Savage, K N

1999-12-01

Spiders produce a variety of silks, and the cloning of genes for silk fibroins reveals a clear link between protein sequence and structure-property relationships. The fibroins produced in the spider's major ampullate (MA) gland, which forms the dragline and web frame, contain multiple repeats of motifs that include an 8-10 residue long poly-alanine block and a 24-35 residue long glycine-rich block. When fibroins are spun into fibres, the poly-alanine blocks form (&bgr;)-sheet crystals that crosslink the fibroins into a polymer network with great stiffness, strength and toughness. As illustrated by a comparison of MA silks from Araneus diadematus and Nephila clavipes, variation in fibroin sequence and properties between spider species provides the opportunity to investigate the design of these remarkable biomaterials.

Heat Capacity of Spider Silk-like Block Copolymers

PubMed Central

Huang, Wenwen; Krishnaji, Sreevidhya; Hu, Xiao; Kaplan, David; Cebe, Peggy

2012-01-01

We synthesized and characterized a new family of di-block copolymers based on the amino acid sequences of Nephila clavipes major ampulate dragline spider silk, having the form HABn and HBAn (n=1–3), comprising an alanine-rich hydrophobic block, A, a glycine-rich hydrophilic block, B, and a histidine tag, H. The reversing heat capacities, Cp(T), for temperatures below and above the glass transition, Tg, were measured by temperature modulated differential scanning calorimetry. For the solid state, we then calculated the heat capacities of our novel block copolymers based on the vibrational motions of the constituent poly(amino acid)s, whose heat capacities are known or can be estimated from the ATHAS Data Bank. For the liquid state, the heat capacity was estimated by using the rotational and translational motions in the polymer chain. Excellent agreement was found between the measured and calculated values of the heat capacity, showing that this method can serve as a standard by which to assess the Cp for other biologically inspired block copolymers. The fraction of beta sheet crystallinity of spider silk block copolymers was also determined by using the predicted Cp, and was verified by wide angle X-ray diffraction and Fourier transform infrared spectroscopy. The glass transition temperatures of spider silk block copolymer were fitted by Kwei’s equation and the results indicate that attractive interaction exists between the A-block and B-block. PMID:23869111

Physical characterization of functionalized spider silk: electronic and sensing properties

PubMed Central

Steven, Eden; Park, Jin Gyu; Paravastu, Anant; Lopes, Elsa Branco; Brooks, James S; Englander, Ongi; Siegrist, Theo; Kaner, Papatya; Alamo, Rufina G

2011-01-01

This work explores functional, fundamental and applied aspects of naturally harvested spider silk fibers. Natural silk is a protein polymer where different amino acids control the physical properties of fibroin bundles, producing, for example, combinations of β-sheet (crystalline) and amorphous (helical) structural regions. This complexity presents opportunities for functional modification to obtain new types of material properties. Electrical conductivity is the starting point of this investigation, where the insulating nature of neat silk under ambient conditions is described first. Modification of the conductivity by humidity, exposure to polar solvents, iodine doping, pyrolization and deposition of a thin metallic film are explored next. The conductivity increases exponentially with relative humidity and/or solvent, whereas only an incremental increase occurs after iodine doping. In contrast, iodine doping, optimal at 70 °C, has a strong effect on the morphology of silk bundles (increasing their size), on the process of pyrolization (suppressing mass loss rates) and on the resulting carbonized fiber structure (that becomes more robust against bending and strain). The effects of iodine doping and other functional parameters (vacuum and thin film coating) motivated an investigation with magic angle spinning nuclear magnetic resonance (MAS-NMR) to monitor doping-induced changes in the amino acid-protein backbone signature. MAS-NMR revealed a moderate effect of iodine on the helical and β-sheet structures, and a lesser effect of gold sputtering. The effects of iodine doping were further probed by Fourier transform infrared (FTIR) spectroscopy, revealing a partial transformation of β-sheet-to-amorphous constituency. A model is proposed, based on the findings from the MAS-NMR and FTIR, which involves iodine-induced changes in the silk fibroin bundle environment that can account for the altered physical properties. Finally, proof-of-concept applications of functionalized spider silk are presented for thermoelectric (Seebeck) effects and incandescence in iodine-doped pyrolized silk fibers, and metallic conductivity and flexibility of micron-sized gold-sputtered silk fibers. In the latter case, we demonstrate the application of gold-sputtered neat spider silk to make four-terminal, flexible, ohmic contacts to organic superconductor samples. PMID:27877440

Tuning chemical and physical cross-links in silk electrogels for morphological analysis and mechanical reinforcement.

PubMed

Lin, Yinan; Xia, Xiaoxia; Shang, Ke; Elia, Roberto; Huang, Wenwen; Cebe, Peggy; Leisk, Gary; Omenetto, Fiorenzo; Kaplan, David L

2013-08-12

Electrochemically controlled, reversible assembly of biopolymers into hydrogel structures is a promising technique for on-demand cell or drug encapsulation and release systems. An electrochemically sol-gel transition has been demonstrated in regenerated Bombyx mori silk fibroin, offering a controllable way to generate biocompatible and reversible adhesives and other biomedical materials. Despite the involvement of an electrochemically triggered electrophoretic migration of the silk molecules, the mechanism of the reversible electrogelation remains unclear. It is, however, known that the freshly prepared silk electrogels (e-gels) adopt a predominantly random coil conformation, indicating a lack of cross-linking as well as thermal, mechanical, and morphological stabilities. In the present work, the tuning of covalent and physical β-sheet cross-links in silk hydrogels was studied for programming the structural properties. Scanning electron microscopy (SEM) revealed delicate morphology, including locally aligned fibrillar structures, in silk e-gels, preserved by combining glutaraldehyde-cross-linking and ethanol dehydration. Fourier transform infrared (FTIR) spectroscopic analysis of either electrogelled, vortex-induced or spontaneously formed silk hydrogels showed that the secondary structure of silk e-gels was tunable between non-β-sheet-dominated and β-sheet-dominated states. Dynamic oscillatory rheology confirmed the mechanical reinforcement of silk e-gels provided by controlled chemical and physical cross-links. The selective incorporation of either chemical or physical or both cross-links into the electrochemically responsive, originally unstructured silk e-gel should help in the design for electrochemically responsive protein polymers.

Microphase Separation Controlled Beta Sheet Crystallization Kinetics in Silk Fibroin Protein.

NASA Astrophysics Data System (ADS)

Hu, Xiao; Lu, Qiang; Kaplan, David; Cebe, Peggy

2009-03-01

We investigate the mechanism of isothermal crystallization kinetics of beta-sheet crystals in silk multiblock fibrous proteins. The Avrami analysis kinetic theory, for studies of synthetic polymer crystal growth, is for the first time extended to investigate protein self-assembly in beta-sheet rich Bombyx mori silk fibroin samples, using time-resolved Fourier transform infrared spectroscopy, differential scanning calorimetry and synchrotron real-time wide-angle X-ray scattering. Results indicate formation of beta sheet crystals in silk proteins is different from the 3-D spherulitic crystal growth found in synthetic homopolymers. Observations by scanning electron microscopy support the view that the protein structures vary during the different stages of crystal growth, and show a microphase separation pattern after chymotrypsin enzyme biodegradation. We present a model to explain the crystallization of the multiblock silk fibroin protein, by analogy to synthetic block copolymers. This model could be widely applicable in other proteins with multiblock (i.e., crystallizable and non-crystallizable) domains.

Structural Properties of Silk Electro-Gels

NASA Astrophysics Data System (ADS)

Tabatabai, A. P.; Urbach, J. S.; Blair, D. L.; Kaplan, D. L.

2013-03-01

The interest in Bombyx Mori silk emerges from its biocompatibility and its structural superiority to synthetic polymers. Our particular interest lies in understanding the capabilities of silk electro-gels because of their reversibility and tunable adhesion. We create an electro-gel by applying a DC electric potential across a reconstituted silk fibroin solution derived directly from Bombyx Mori cocoons. This process leads to the intermolecular self-assembly of fibroin proteins into a weak gel. In this talk we will present our results on the effects of applied shear on electro-gels. We quantify the structural properties while dynamically imaging shear induced fiber formation; known as fibrillogenesis. It is observed that the mechanical properties and microstructure of these materials are highly dependent on shear history. We will also discuss the role of surface modification, through micro-patterning, on the observed gel structure. Our results provide an understanding of both the viscoelastiticity and microstucture of reconstituted silks that are being utilized as tissue scaffolds. This work is supported by a grant from the AFOSR FA9550-07-1-0130.

Integration of amorphous tantalum silicon nitride (TaSiN) films as diffusion barriers in a Cu/SiLK(TM) metallization scheme

NASA Astrophysics Data System (ADS)

Padiyar, Sumant Devdas

2003-09-01

Current and future performance requirements for high- speed integrated circuit (IC) devices have placed great emphasis on the introduction of novel materials, deposition techniques and improved metrology techniques. The introduction of copper interconnects and more currently low-k dielectric materials in IC fabrication are two such examples. This introduction necessitates research on the compatibility of these materials and process techniques with adjacent diffusion barrier materials. One candidate, which has attracted significant attention is tantalum-silicon-nitride (TaSiN) on account of its superior diffusion barrier performance and high recrystallization temperature1. The subject of this dissertation is an investigation of the integration compatibility and performance of TaSiN barrier layers with a low-k dielectric polymer (SiLK ®2). A plasma- enhanced chemical vapor deposition (PECVD) approach is taken for growth of TaSiN films in this work due to potential advantages in conformal film coverage compared to more conventional physical vapor deposition methods. A Design of Experiment (DOE) methodology was introduced for PECVD of TaSiN on SiLK to optimize film properties such as film composition, resistivity, growth rate and film roughness with respect to the predictors viz. substrate temperature, precursor gas flow and plasma power. The first pass study determined the response window for optimized TaSiN film composition, growth rate and low halide contamination and the compatibility of the process with an organic polymer substrate, i.e. SiLK. Second-pass studies were carried out to deposit ultra- thin (10nm) films on: (a)blanket SiLK to investigate the performance of TaSiN films against copper diffusion, and (b)patterned SiLK to evaluate step coverage and conformality. All TaSiN depositions were carried out on SiO2 substrates for baseline comparisons. A second purpose of the diffusion barrier in IC processing is to improve interfacial adhesion between the barrier and the adjacent dielectric material; especially important for an organic polymer like SiLK. Hence, a detailed study was undertaken to evaluate the interfacial adhesion of TaSiN with SiLK and SiO2 and study the dependence of the adhesion with the film composition. The results of diffusion barrier performance studies, conformality studies, and interfacial adhesion studies of TaSiN films are discussed in relation to the elemental compositions of the films. 1J. S. Reid, M. Nicolet, J. Appl. Phys. 79 (2) p. 1109 (1996). 2SiLK is a low-k dielectric candidate registered by Dow Chemical Company, MI.

Biomimetic Composite Scaffold for Breast Reconstruction Following Tumor Resection

DTIC Science & Technology

2005-09-01

developing an innovative biomimetic scaffold materials by combining two natural polymers: silk fibroin (from Bombyx mori silk worm) and chitosan (from...M.D. Anderson Cancer Center (UTMDACC) is a component of the University of Texas System. It is one of the nation’s original three Comprehensive Cancer...follow protocols • Properly collect, measure, and document data from the lab, clinic, and/or animal facilities 22 Table 2. Origin of Country and

Processing and surface modification of novel natural-origin architectures aimed for biomedical applications

NASA Astrophysics Data System (ADS)

Silva, Simone dos Santos

In the last decades, tissue engineering has emerged as a potential therapeutical tool aimed at developing substitutes that are able to restore proper function of the damaged organs/tissues. Nature-inspired routes involving natural origin polymer-based systems represent an attractive alternative to produce novel materials by mimicking the tissue environment required for tissue regeneration. Moreover, further modifications of these systems allow the adjustment of their properties in accordance with the requirements for successful biomedical applications. The main goal of the present thesis is to develop and modify natural origin polymer-based systems using simple methodologies such as sol-gel, surface modification by means of plasma treatment and blending of chitosan with proteins (soy protein isolate and silk fibroin). A sol-gel method was used to improve the bulk properties of chitosan by the incorporation of an inorganic component at the sub-nanometric level. Chitosan/siloxane hybrid materials were synthesised, where essentially urea bridges covalently bond the chitosan to the polysiloxane network. These bifunctional materials exhibit interesting photoluminescence features and a bioactive behaviour. In most situations in the biomedical field, the surface of a biomaterial is in direct contact with living tissues. Therefore, the surface characteristics play a fundamental role on the implant biocompatibility. In this thesis, nitrogen and argon plasma treatment was applied on chitosan membranes in order to improve their surface properties. The applied modifications promoted differences on surface chemistry, wettability and roughness, which reflected in a significant improvement of fibroblast adhesion and proliferation onto chitosan membranes. Besides the surface modification, blending of chitosan with proteins such as soy protein isolate and silk fibroin was also used to modify the bulk properties of chitosan. In situ cross-linking with glutaraldehyde solutions was used to enhance the interaction between the components of the blend. Hence, membranes with different morphologies, water absorption and degradability were obtained. The biological assays suggested that the cross-linking with lower glutaraldehyde concentration promotes better cell adhesion on the membranes. The morphological characterization showed that both surface roughness surface and surface energy were dependent on soy protein content. Structural investigations by FTIR and NMR indicated that the blends are not completely miscible due to a weak polysaccharide-protein interaction. In another related work, novel hydrogels were produced combining Bombyx mori silk fibroin and chitosan. In this case, these systems were cross-linked with genipin. These hydrogels were freeze dried to obtain cross-linked chitosan/silk sponges. Rheological and mechanical properties, structural aspects and morphological features of the porous structures were evaluated. The results revealed stable and ordered structures, similar porosities, and swelling capability that depended on the pH. The cytotoxicity assay indicated that cellular viability was about 100% in all sponges and for all time points studied (1, 3, 7 and 14 days), demonstrating the extremely low cytotoxicity levels of the materials. Cell studies using chondrocytes-like cells seeded onto sponges, including cell viability (MTS assay), proliferation (DNA test), morphology (SEM analysis) and matrix production (GAGs quantification), showed a significant high adhesion, proliferation and matrix production with the time of culture. The findings in this work suggested that the properties of the sponges can be manipulated by either change chitosan/silk fibroin ratio or through genipin cross-linking. Parallel to this study, the possibility of obtaining modified silk nanometric nets using electrospinning processing from regenerated silk fibroin/formic acid with addition of genipin was explored. Modified silk nanofibers with diameters ranging from 140 nm to 590 nm were developed. The changes on the secondary structure of nanofibers, induced by the reaction of silk fibroin with genipin, promoted a higher integrity of these modified nanofibers in water. In summary, the findings from these works demonstrated the potential and versatility of the proposed strategies in obtaining different structures (e.g. membranes, hydrogels) using mixtures of chitosan with proteins or with inorganic agents for improving the performance of natural origin polymer-based materials to be used in biomedical applications.

Water-based preparation of spider silk films as drug delivery matrices.

PubMed

Agostini, Elisa; Winter, Gerhard; Engert, Julia

2015-09-10

The main focus of this work was to obtain a drug delivery matrix characterized by biocompatibility, water insolubility and good mechanical properties. Moreover the preparation process has to be compatible with protein encapsulation and the obtained matrix should be able to sustain release a model protein. Spider silk proteins represent exceptional natural polymers due to their mechanical properties in combination with biocompatibility. As both hydrophobic and slowly biodegrading biopolymers, recombinant spider silk proteins fulfill the required properties for a drug delivery system. In this work, we present the preparation of eADF4(C16) films as drug delivery matrices without the use of any organic solvent. Water-based spider silk films were characterized in terms of protein secondary structure, thermal stability, zeta-potential, solubility, mechanical properties, and water absorption and desorption. Additionally, this study includes an evaluation of their application as a drug delivery system for both small molecular weight drugs and high molecular weight molecules such as proteins. Our investigation focused on possible improvements in the film's mechanical properties including plasticizers in the film matrix. Furthermore, different film designs were prepared, such as: monolayer, coated monolayer, multilayer (sandwich), and coated multilayer. The release of the model protein BSA from these new systems was studied. Results indicated that spider silk films are a promising protein drug delivery matrix, capable of releasing the model protein over 90 days with a release profile close to zero order kinetic. Such films could be used for several pharmaceutical and medical purposes, especially when mechanical strength of a drug eluting matrix is of high importance. Copyright © 2015 Elsevier B.V. All rights reserved.

Phase Behaviour and Miscibility Studies of Collagen/Silk Fibroin Macromolecular System in Dilute Solutions and Solid State.

PubMed

Ghaeli, Ima; de Moraes, Mariana A; Beppu, Marisa M; Lewandowska, Katarzyna; Sionkowska, Alina; Ferreira-da-Silva, Frederico; Ferraz, Maria P; Monteiro, Fernando J

2017-08-18

Miscibility is an important issue in biopolymer blends for analysis of the behavior of polymer pairs through the detection of phase separation and improvement of the mechanical and physical properties of the blend. This study presents the formulation of a stable and one-phase mixture of collagen and regenerated silk fibroin (RSF), with the highest miscibility ratio between these two macromolecules, through inducing electrostatic interactions, using salt ions. For this aim, a ternary phase diagram was experimentally built for the mixtures, based on observations of phase behavior of blend solutions with various ratios. The miscibility behavior of the blend solutions in the miscible zones of the phase diagram was confirmed quantitatively by viscosimetric measurements. Assessing the effects of biopolymer mixing ratio and salt ions, before and after dialysis of blend solutions, revealed the importance of ion-specific interactions in the formation of coacervate-based materials containing collagen and RSF blends that can be used in pharmaceutical, drug delivery, and biomedical applications. Moreover, the conformational change of silk fibroin from random coil to beta sheet, in solution and in the final solid films, was detected by circular dichroism (CD) and Fourier transform infrared spectroscopy (FTIR), respectively. Scanning electron microscopy (SEM) exhibited alterations of surface morphology for the biocomposite films with different ratios. Surface contact angle measurement illustrated different hydrophobic properties for the blended film surfaces. Differential scanning calorimetry (DSC) showed that the formation of the beta sheet structure of silk fibroin enhances the thermal stability of the final blend films. Therefore, the novel method presented in this study resulted in the formation of biocomposite films whose physico-chemical properties can be tuned by silk fibroin conformational changes by applying different component mixing ratios.

Exploring the Properties of Genetically Engineered Silk-Elastin-Like Protein Films.

PubMed

Machado, Raul; da Costa, André; Sencadas, Vitor; Pereira, Ana Margarida; Collins, Tony; Rodríguez-Cabello, José Carlos; Lanceros-Méndez, Senentxu; Casal, Margarida

2015-12-01

Free standing films of a genetically engineered silk-elastin-like protein (SELP) were prepared using water and formic acid as solvents. Exposure to methanol-saturated air promoted the formation of aggregated β-strands rendering aqueous insolubility and improved the mechanical properties leading to a 10-fold increase in strain-to-failure. The films were optically clear with resistivity values similar to natural rubber and thermally stable up to 180 °C. Addition of glycerol showed to enhance the flexibility of SELP/glycerol films by interacting with SELP molecules through hydrogen bonding, interpenetrating between the polymer chains and granting more conformational freedom. This detailed characterization provides cues for future and unique applications using SELP based biopolymers. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Development and characterization of silk fibroin coated quantum dots

NASA Astrophysics Data System (ADS)

Nathwani, B. B.; Needham, C.; Mathur, A. B.; Meissner, K. E.

2008-02-01

Recent progress in the field of semiconductor nanocrystals or Quantum Dots (QDs) has seen them find wider acceptance as a tool in biomedical research labs. As produced, high quality QDs, synthesized by high temperature organometallic synthesis, are coated with a hydrophobic ligand. Therefore, they must be further processed to be soluble in water and to be made biocompatible. To accomplish this, the QDs are generally coated with a synthetic polymer (eg. block copolymers) or the hydrophobic surface ligands exchanged with hydrophilic material (eg. thiols). Advances in this area have enabled the QDs to experience a smooth transition from being simple inorganic fluorophores to being smart sensors, which can identify specific cell marker proteins and help in diagnosis of diseases such as cancer. In order to improve the biocompatibility and utility of the QDs, we report the development of a procedure to coat QDs with silk fibroin, a fibrous crystalline protein extracted from Bombyx Mori silkworm. Following the coating process, we characterize the size, quantum yield and two-photon absorption cross section of the silk coated QDs. Additionally, the results of biocompatibility studies carried out to compare the properties of these QD-silks with conventional QDs are presented. These natural polymer coatings on QDs could enhance the intracellular delivery and enable the use of these nanocrystals as an imaging tool for studying subcellular machinery at the molecular level.

Invited review liquid crystal models of biological materials and silk spinning.

PubMed

Rey, Alejandro D; Herrera-Valencia, Edtson E

2012-06-01

A review of thermodynamic, materials science, and rheological liquid crystal models is presented and applied to a wide range of biological liquid crystals, including helicoidal plywoods, biopolymer solutions, and in vivo liquid crystals. The distinguishing characteristics of liquid crystals (self-assembly, packing, defects, functionalities, processability) are discussed in relation to biological materials and the strong correspondence between different synthetic and biological materials is established. Biological polymer processing based on liquid crystalline precursors includes viscoelastic flow to form and shape fibers. Viscoelastic models for nematic and chiral nematics are reviewed and discussed in terms of key parameters that facilitate understanding and quantitative information from optical textures and rheometers. It is shown that viscoelastic modeling the silk spinning process using liquid crystal theories sheds light on textural transitions in the duct of spiders and silk worms as well as on tactoidal drops and interfacial structures. The range and consistency of the predictions demonstrates that the use of mesoscopic liquid crystal models is another tool to develop the science and biomimetic applications of mesogenic biological soft matter. Copyright © 2011 Wiley Periodicals, Inc.

Atomistic modeling of interphases in spider silk fibers

NASA Astrophysics Data System (ADS)

Fossey, Stephen Andrew

The objective of this work is to create an atomistic model to account for the unusual physical properties of silk fibers. Silk fibers have exceptional mechanical toughness, which makes them of interest as high performance fibers. In order to explain the toughness, a model for the molecular structure based on simple geometric reasoning was formulated. The model consists of very small crystallites, on the order of 5 nm, connected by a noncrystalline interphase. The interphase is a region between the crystalline phase and the amorphous phase, which is defined by the geometry of the system. The interphase is modeled as a very thin (<5 nm) film of noncrystalline polymer constructed using a Monte Carlo, rotational isomeric states approach followed by simulated annealing in order to achieve equilibrium chain configurations and density. No additional assumptions are made about density, orientation, or packing. The mechanical properties of the interphase are calculated using the method of Theodoreau and Suter. Finally, observable properties such as wide angle X-ray scattering and methyl rotation rates are calculated and compared with experimental data available in the literature.

Protein-based flexible whispering gallery mode resonators

NASA Astrophysics Data System (ADS)

Yilmaz, Huzeyfe; Pena-Francesch, Abdon; Xu, Linhua; Shreiner, Robert; Jung, Huihun; Huang, Steven H.; Özdemir, Sahin K.; Demirel, Melik C.; Yang, Lan

2016-02-01

The idea of creating photonics tools for sensing, imaging and material characterization has long been pursued and many achievements have been made. Approaching the level of solutions provided by nature however is hindered by routine choice of materials. To this end recent years have witnessed a great effort to engineer mechanically flexible photonic devices using polymer substrates. On the other hand, biodegradability and biocompatibility still remains to be incorporated. Hence biomimetics holds the key to overcome the limitations of traditional materials in photonics design. Natural proteins such as sucker ring teeth (SRT) and silk for instance have remarkable mechanical and optical properties that exceed the endeavors of most synthetic and natural polymers. Here we demonstrate for the first time, toroidal whispering gallery mode resonators (WGMR) fabricated entirely from protein structures such as SRT of Loligo vulgaris (European squid) and silk from Bombyx mori. We provide here complete optical and material characterization of proteinaceous WGMRs, revealing high quality factors in microscale and enhancement of Raman signatures by a microcavity. We also present a most simple application of a WGMR as a natural protein add-drop filter, made of SRT protein. Our work shows that with protein-based materials, optical, mechanical and thermal properties can be devised at the molecular level and it lays the groundwork for future eco-friendly, flexible photonics device design.

Biocompatible and biodegradable dual-drug release system based on silk hydrogel containing silk nanoparticles.

PubMed

Numata, Keiji; Yamazaki, Shoya; Naga, Naofumi

2012-05-14

We developed a facile and quick ethanol-based method for preparing silk nanoparticles and then fabricated a biodegradable and biocompatible dual-drug release system based on silk nanoparticles and the molecular networks of silk hydrogels. Model drugs incorporated in the silk nanoparticles and silk hydrogels showed fast and constant release, respectively, indicating successful dual-drug release from silk hydrogel containing silk nanoparticles. The release behaviors achieved by this dual-drug release system suggest to be regulated by physical properties (e.g., β-sheet contents and size of the silk nanoparticles and network size of the silk hydrogels), which is an important advantage for biomedical applications. The present silk-based system for dual-drug release also demonstrated no significant cytotoxicity against human mesenchymal stem cells (hMSCs), and thus, this silk-based dual-drug release system has potential as a versatile and useful new platform of polymeric materials for various types of dual delivery of bioactive molecules.

Silk-fibronectin protein alloy fibres support cell adhesion and viability as a high strength, matrix fibre analogue

NASA Astrophysics Data System (ADS)

Jacobsen, Matthew M.; Li, David; Gyune Rim, Nae; Backman, Daniel; Smith, Michael L.; Wong, Joyce Y.

2017-04-01

Silk is a natural polymer with broad utility in biomedical applications because it exhibits general biocompatibility and high tensile material properties. While mechanical integrity is important for most biomaterial applications, proper function and integration also requires biomaterial incorporation into complex surrounding tissues for many physiologically relevant processes such as wound healing. In this study, we spin silk fibroin into a protein alloy fibre with whole fibronectin using wet spinning approaches in order to synergize their respective strength and cell interaction capabilities. Results demonstrate that silk fibroin alone is a poor adhesive surface for fibroblasts, endothelial cells, and vascular smooth muscle cells in the absence of serum. However, significantly improved cell attachment is observed to silk-fibronectin alloy fibres without serum present while not compromising the fibres’ mechanical integrity. Additionally, cell viability is improved up to six fold on alloy fibres when serum is present while migration and spreading generally increase as well. These findings demonstrate the utility of composite protein alloys as inexpensive and effective means to create durable, biologically active biomaterials.

Advancing the frontiers of silk fibroin protein-based materials for futuristic electronics and clinical wound-healing (Invited review).

PubMed

Koh, Leng-Duei; Yeo, Jingjie; Lee, Yeong Yuh; Ong, Qunya; Han, Mingyong; Tee, Benjamin C-K

2018-05-01

The present review will introduce the basic concepts of silk-based electronics/optoelectronics including the latest technological advances on the use of silk fibroin in combination with other functional components, with an emphasis on improving the performance of next-generation silk-based materials. It also highlights the patterning of silk fibroin to produce micro/nano-scale features, as well as the functionalization of silk fibroin to impart antimicrobial (i.e. antibacterial) properties. Silk-based bioelectronics have great potential for advanced or futuristic bio-applications including e-skins, e-bandages, biosensors, wearable displays, implantable devices, artificial muscles, etc. Notably, silk-based organic field-effect transistors have highly promising applications in e-skins and biosensors; silk-based electrodes/antennas are used for in vivo bioanalysis or sensing purpose (e.g., measurement of neurotransmitter such as dopamine) in addition to their use as food sensors; silk-based diodes can be applied as light sources for wound healing or tissue engineering, e.g., in cutaneous wound closure or induction of photothrombosis of corneal neovascularization; silk-based actuators have promising applications as artificial muscles; whereas silk-based memristors have exciting applications as logic or synaptic network for realizing e-skins or bionic brains. Copyright © 2018 Elsevier B.V. All rights reserved.

Gel spinning of silk tubes for tissue engineering

PubMed Central

Lovett, Michael; Cannizzaro, Christopher; Vunjak-Novakovic, Gordana; Kaplan, David L.

2011-01-01

Tubular vessels for tissue engineering are typically fabricated using a molding, dipping, or electrospinning technique. While these techniques provide some control over inner and outer diameters of the tube, they lack the ability to align the polymers or fibers of interest throughout the tube. This is an important aspect of biomaterial composite structure and function for mechanical and biological impact of tissue outcomes. We present a novel aqueous process system to spin tubes from biopolymers and proteins such as silk fibroin. Using silk as an example, this method of winding an aqueous solution around a reciprocating rotating mandrel offers substantial improvement in the control of the tube properties, specifically with regard to winding pattern, tube porosity, and composite features. Silk tube properties are further controlled via different post-spinning processing mechanisms such as methanol-treatment, air-drying, and lyophilization. This approach to tubular scaffold manufacture offers numerous tissue engineering applications such as complex composite biomaterial matrices, blood vessel grafts and nerve guides, among others. PMID:18801570

Enhanced stiffness of silk-like fibers by loop formation in the corona leads to stronger gels.

PubMed

Rombouts, Wolf H; Domeradzka, Natalia E; Werten, Marc W T; Leermakers, Frans A M; de Vries, Renko J; de Wolf, Frits A; van der Gucht, Jasper

2016-11-01

We study the self-assembly of protein polymers consisting of a silk-like block flanked by two hydrophilic blocks, with a cysteine residue attached to the C-terminal end. The silk blocks self-assemble to form fibers while the hydrophilic blocks form a stabilizing corona. Entanglement of the fibers leads to the formation of hydrogels. Under oxidizing conditions the cysteine residues form disulfide bridges, effectively connecting two corona chains at their ends to form a loop. We find that this leads to a significant increase in the elastic modulus of the gels. Using atomic force microscopy, we show that this stiffening is due to an increase of the persistence length of the fibers. Self-consistent-field calculations indicate a slight decrease of the lateral pressure in the corona upon loop formation. We argue that this small decrease in the repulsive interactions affects the stacking of the silk-like blocks in the core, resulting in a more rigid fiber. © 2016 Wiley Periodicals, Inc.

Silk constructs for delivery of muskuloskeletal therapeutics

PubMed Central

Meinel, Lorenz; Kaplan, David L.

2012-01-01

Silk fibroin (SF) is a biopolymer with distinguishing features from many other bio- as well as synthetic polymers. From a biomechanical and drug delivery perspective, SF combines remarkable versatility for scaffolding (solid implants, hydrogels, threads, solutions), with advanced mechanical properties and good stabilization and controlled delivery of entrapped protein and small molecule drugs, respectively. It is this combination of mechanical and pharmaceutical features which render SF so exciting for biomedical applications. his pattern along with the versatility of this biopolymer have been translated into progress for musculoskeletal applications. We review the use and potential of silk fibroin for systemic and localized delivery of therapeutics in diseases affecting the musculoskeletal system. We also present future directions for this biopolymer as well as the necessary research and development steps for their achievement. PMID:22522139

Nonlinear control of high-frequency phonons in spider silk

NASA Astrophysics Data System (ADS)

Schneider, Dirk; Gomopoulos, Nikolaos; Koh, Cheong Y.; Papadopoulos, Periklis; Kremer, Friedrich; Thomas, Edwin L.; Fytas, George

2016-10-01

Spider dragline silk possesses superior mechanical properties compared with synthetic polymers with similar chemical structure due to its hierarchical structure comprised of partially crystalline oriented nanofibrils. To date, silk’s dynamic mechanical properties have been largely unexplored. Here we report an indirect hypersonic phononic bandgap and an anomalous dispersion of the acoustic-like branch from inelastic (Brillouin) light scattering experiments under varying applied elastic strains. We show the mechanical nonlinearity of the silk structure generates a unique region of negative group velocity, that together with the global (mechanical) anisotropy provides novel symmetry conditions for gap formation. The phononic bandgap and dispersion show strong nonlinear strain-dependent behaviour. Exploiting material nonlinearity along with tailored structural anisotropy could be a new design paradigm to access new types of dynamic behaviour.

Antimicrobial Silk Ocular Drug Delivery Implant for Chronic Posterior Segment Diseases

DTIC Science & Technology

2016-06-01

antibody bevacizumab. The device was covalently coated with the antibacterial polymer HMPE1 to protect against biofilm formation and the transmission of harmful bacterial to the interior of the ocular chamber.

Enzymatically crosslinked silk-hyaluronic acid hydrogels.

PubMed

Raia, Nicole R; Partlow, Benjamin P; McGill, Meghan; Kimmerling, Erica Palma; Ghezzi, Chiara E; Kaplan, David L

2017-07-01

In this study, silk fibroin and hyaluronic acid (HA) were enzymatically crosslinked to form biocompatible composite hydrogels with tunable mechanical properties similar to that of native tissues. The formation of di-tyrosine crosslinks between silk fibroin proteins via horseradish peroxidase has resulted in a highly elastic hydrogel but exhibits time-dependent stiffening related to silk self-assembly and crystallization. Utilizing the same method of crosslinking, tyramine-substituted HA forms hydrophilic and bioactive hydrogels that tend to have limited mechanics and degrade rapidly. To address the limitations of these singular component scaffolds, HA was covalently crosslinked with silk, forming a composite hydrogel that exhibited both mechanical integrity and hydrophilicity. The composite hydrogels were assessed using unconfined compression and infrared spectroscopy to reveal of the physical properties over time in relation to polymer concentration. In addition, the hydrogels were characterized by enzymatic degradation and for cytotoxicity. Results showed that increasing HA concentration, decreased gelation time, increased degradation rate, and reduced changes that were observed over time in mechanics, water retention, and crystallization. These hydrogel composites provide a biologically relevant system with controllable temporal stiffening and elasticity, thus offering enhanced tunable scaffolds for short or long term applications in tissue engineering. Copyright © 2017 Elsevier Ltd. All rights reserved.

Microfabrication of a spider-silk analogue through the liquid rope coiling instability

NASA Astrophysics Data System (ADS)

Gosselin, Frederick P.; Therriault, Daniel; Levesque, Martin

2012-02-01

Spider capture silk outperforms most synthetic materials in terms of specific toughness. We developed a technique to fabricate tough microstructured fibers inspired by the molecular structure of the spider silk protein. To fabricate microfibers (with diameter ˜30μm) with various mechanical properties, we yield the control of their exact geometry to the liquid rope coiling instability. This instability causes a thread of honey to wiggle as it buckles when hitting a surface. Similarly, we flow a filament of viscous polymer solution towards a substrate moving perpendicularly at a slower velocity than the filament flows. The filament buckles repetitively giving rise to periodic meanders and stitch patterns. As the solvent evaporates, the filament solidifies into a fiber with a geometry bestowed by the instability. Microtraction tests performed on fibers show interesting links between the mechanical properties and the instability patterns. Some coiling patterns give rise to high toughness due to the sacrificial bonds created when the viscous filament loops over itself and fuse. The sacrificial bonds in the microstructured fiber play an analogous role to that of the hydrogen bonds present in the molecular structure of the silk protein which give its toughness to spider silk.

Silk fibroin nanoparticles prepared by electrospray as controlled release carriers of cisplatin.

PubMed

Qu, Jing; Liu, Yu; Yu, Yanni; Li, Jing; Luo, Jingwan; Li, Mingzhong

2014-11-01

To maintain the anti-tumor activity of cis-dichlorodiamminoplatinum (CDDP) while avoiding its cytotoxicity and negative influence on normal tissue, CDDP-loaded silk fibroin nanoparticles approximately 59 nm in diameter were successfully prepared by electrospray without using organic solvent. CDDP was incorporated into nanoparticles through metal-polymer coordination bond exchange. In vitro release tests showed that the cisplatin in the nanoparticles could be slowly and sustainably released for more than 15 days. In vitro anti-cancer experiments and intracellular Pt content testing indicated that CDDP-loaded silk fibroin nanoparticles were easily internalized by A549 lung cancer cells, transferring CDDP into cancer cells and then triggering their apoptosis. In contrast, the particles were not easily internalized by L929 mouse fibroblast cells and hence showed weaker cell growth inhibition. The CDDP-loaded silk fibroin nanoparticles showed sustained and efficient killing of tumor cells but weaker inhibition of normal cells. In general, this study provides not only a novel method for preparing CDDP-loaded silk fibroin nanoparticles but also a new carrier system for clinical therapeutic drugs against lung cancers and other tumors. Copyright © 2014 Elsevier B.V. All rights reserved.

In vivo bioresponses to silk proteins.

PubMed

Thurber, Amy E; Omenetto, Fiorenzo G; Kaplan, David L

2015-12-01

Silks are appealing materials for numerous biomedical applications involving drug delivery, tissue engineering, or implantable devices, because of their tunable mechanical properties and wide range of physical structures. In addition to the functionalities needed for specific clinical applications, a key factor necessary for clinical success for any implanted material is appropriate interactions with the body in vivo. This review summarizes our current understanding of the in vivo biological responses to silks, including degradation, the immune and inflammatory response, and tissue remodeling with particular attention to vascularization. While we focus in this review on silkworm silk fibroin protein due to the large quantity of in vivo data thanks to its widespread use in medical materials and consumer products, spider silk information is also included if available. Silk proteins are degraded in the body on a time course that is dependent on the method of silk fabrication and can range from hours to years. Silk protein typically induces a mild inflammatory response that decreases within a few weeks of implantation. The response involves recruitment and activation of macrophages and may include activation of a mild foreign body response with the formation of multinuclear giant cells, depending on the material format and location of implantation. The number of immune cells present decreases with time and granulation tissue, if formed, is replaced by endogenous, not fibrous, tissue. Importantly, silk materials have not been demonstrated to induce mineralization, except when used in calcified tissues. Due to its ability to be degraded, silk can be remodeled in the body allowing for vascularization and tissue ingrowth with eventual complete replacement by native tissue. The degree of remodeling, tissue ingrowth, or other specific cell behaviors can be modulated with addition of growth or other signaling factors. Silk can also be combined with numerous other materials including proteins, synthetic polymers, and ceramics to enhance its characteristics for a particular function. Overall, the diverse array of silk materials shows excellent bioresponses in vivo with low immunogenicity and the ability to be remodeled and replaced by native tissue making it suitable for numerous clinical applications. Copyright © 2015 Elsevier Ltd. All rights reserved.

A Comparison of Electrospun Polymers Reveals Poly(3-Hydroxybutyrate) Fiber as a Superior Scaffold for Cardiac Repair

PubMed Central

Castellano, Delia; Blanes, María; Marco, Bruno; Cerrada, Inmaculada; Ruiz-Saurí, Amparo; Pelacho, Beatriz; Araña, Miriam; Montero, Jose A.; Cambra, Vicente; Prosper, Felipe

2014-01-01

The development of biomaterials for myocardial tissue engineering requires a careful assessment of their performance with regards to functionality and biocompatibility, including the immune response. Poly(3-hydroxybutyrate) (PHB), poly(e-caprolactone) (PCL), silk, poly-lactic acid (PLA), and polyamide (PA) scaffolds were generated by electrospinning, and cell compatibility in vitro, and immune response and cardiac function in vitro and in vivo were compared with a noncrosslinked collagen membrane (Col) control material. Results showed that cell adhesion and growth of mesenchymal stem cells, cardiomyocytes, and cardiac fibroblasts in vitro was dependent on the polymer substrate, with PHB and PCL polymers permitting the greatest adhesion/growth of cells. Additionally, polymer substrates triggered unique expression profiles of anti- and pro-inflammatory cytokines in human peripheral blood mononuclear cells. Implantation of PCL, silk, PLA, and PA patches on the epicardial surface of healthy rats induced a classical foreign body reaction pattern, with encapsulation of polymer fibers and induction of the nonspecific immune response, whereas Col and PHB patches were progressively degraded. When implanted on infarcted rat heart, Col, PCL, and PHB reduced negative remodeling, but only PHB induced significant angiogenesis. Importantly, Col and PHB modified the inflammatory response to an M2 macrophage phenotype in cardiac tissue, indicating a more beneficial reparative process and remodeling. Collectively, these results identify PHB as a superior substrate for cardiac repair. PMID:24564648

Silk Materials Functionalized via Genetic Engineering for Biomedical Applications.

PubMed

Deptuch, Tomasz; Dams-Kozlowska, Hanna

2017-12-12

The great mechanical properties, biocompatibility and biodegradability of silk-based materials make them applicable to the biomedical field. Genetic engineering enables the construction of synthetic equivalents of natural silks. Knowledge about the relationship between the structure and function of silk proteins enables the design of bioengineered silks that can serve as the foundation of new biomaterials. Furthermore, in order to better address the needs of modern biomedicine, genetic engineering can be used to obtain silk-based materials with new functionalities. Sequences encoding new peptides or domains can be added to the sequences encoding the silk proteins. The expression of one cDNA fragment indicates that each silk molecule is related to a functional fragment. This review summarizes the proposed genetic functionalization of silk-based materials that can be potentially useful for biomedical applications.

Dynamic Camouflage Materials Based on Silk-Reflectin Chimeras

DTIC Science & Technology

2012-08-01

Dynamic Camouflage Materials Based on Silk -Reflectin Chimeras Final Performance Report for FA9550-09-1-0513 (Program Manager: Hugh DeLong...efforts to bioengineer silk -reflectin chimeric proteins, with the silk component serving as one of the organizing elements for material functions and...Further contributions may also come from the silk due to its novel light guiding properties and diffractive optics. Variants in silk block sizes

Silk Materials Functionalized via Genetic Engineering for Biomedical Applications

PubMed Central

Deptuch, Tomasz

2017-01-01

The great mechanical properties, biocompatibility and biodegradability of silk-based materials make them applicable to the biomedical field. Genetic engineering enables the construction of synthetic equivalents of natural silks. Knowledge about the relationship between the structure and function of silk proteins enables the design of bioengineered silks that can serve as the foundation of new biomaterials. Furthermore, in order to better address the needs of modern biomedicine, genetic engineering can be used to obtain silk-based materials with new functionalities. Sequences encoding new peptides or domains can be added to the sequences encoding the silk proteins. The expression of one cDNA fragment indicates that each silk molecule is related to a functional fragment. This review summarizes the proposed genetic functionalization of silk-based materials that can be potentially useful for biomedical applications. PMID:29231863

Structural study of Bombyx mori silk fibroin during processing for regeneration

NASA Astrophysics Data System (ADS)

Ha, Sung-Won

Bombyx mori silk fibroin has excellent mechanical properties combined with flexibility, tissue compatibility, and high oxygen permeability in the wet condition. This important material should be dissolved and regenerated to be utilized as useful forms such as gel, film, fiber, powder, or non-woven. However, it has long been a problem that the regenerated fibroin materials show poor mechanical properties and brittleness. These problems were technically solved by improving a fiber processing method reported here. The regenerated fibroin fibers showed much better mechanical properties compared to the original silk fibers. This improved technique for the fiber processing of Bombyx mori silk fibroin may be used as a model system for other semi-crystalline fiber forming proteins, becoming available through biotechnology. The physical and chemical properties of the regenerated fibers were characterized by SinTechRTM tensile testing, X-ray diffraction, solid state 13C NMR spectroscopy, and SEM. Unlike synthetic polymers, the molecular weight distribution of Bombyx mori silk fibroin is mono-disperse because silk fibroin is synthesized from DNA template. Genetic studies have revealed the entire amino acid sequence of Bombyx mori silk fibroin. It is known that the crystalline silk II structure is composed of hexa-amino acid sequences, GAGAGS. However, in the amino acid sequence of Bombyx mori silk fibroin heavy chain, there are present 11 chemically irregular but evolutionarily conserved sequences with about 31 amino acid residues (irregular GT˜GT sequences). The structure and role of these irregular sequences have remained unknown. One of the most frequently appearing irregular sequences was synthesized by a peptide synthesizer. The three-dimensional structure of this irregular silk peptide was studied by the high resolution two-dimensional NMR technique. The three-dimensional structure of this peptide shows that it makes a turn or loop structure (distorted O shape), which means the proceeding backbone direction is changed 180° by this sequence. This may facilitate the beta-sheet formation of the crystal forming building blocks, GAGAGS/GY˜GY sequences, in fibroin heavy chain. It may also facilitate the solubilization of the fibroin heavy chain within the silk gland.

Electrospinning of Chitosan-Based Solutions for Tissue Engineering and Regenerative Medicine

PubMed Central

Qasim, Saad B.; Khurshid, Zohaib; Shah, Altaf H.; Husain, Shehriar; Rehman, Ihtesham Ur

2018-01-01

Electrospinning has been used for decades to generate nano-fibres via an electrically charged jet of polymer solution. This process is established on a spinning technique, using electrostatic forces to produce fine fibres from polymer solutions. Amongst, the electrospinning of available biopolymers (silk, cellulose, collagen, gelatine and hyaluronic acid), chitosan (CH) has shown a favourable outcome for tissue regeneration applications. The aim of the current review is to assess the current literature about electrospinning chitosan and its composite formulations for creating fibres in combination with other natural polymers to be employed in tissue engineering. In addition, various polymers blended with chitosan for electrospinning have been discussed in terms of their potential biomedical applications. The review shows that evidence exists in support of the favourable properties and biocompatibility of chitosan electrospun composite biomaterials for a range of applications. However, further research and in vivo studies are required to translate these materials from the laboratory to clinical applications. PMID:29385727

Comparative Study of Ultrasonication-Induced and Naturally Self-Assembled Silk Fibroin-Wool Keratin Hydrogel Biomaterials

PubMed Central

Vu, Trang; Xue, Ye; Vuong, Trinh; Erbe, Matthew; Bennet, Christopher; Palazzo, Ben; Popielski, Lucas; Rodriguez, Nelson; Hu, Xiao

2016-01-01

This study reports the formation of biocompatible hydrogels using protein polymers from natural silk cocoon fibroins and sheep wool keratins. Silk fibroin protein contains β-sheet secondary structures, allowing for the formation of physical cross-linkers in the hydrogels. Comparative studies were performed on two groups of samples. In the first group, ultrasonication was used to induce a quick gelation of a protein aqueous solution, enhancing the ability of Bombyx mori silk fibroin chains to quickly entrap the wool keratin protein molecules homogenously. In the second group, silk/keratin mixtures were left at room temperature for days, resulting in naturally-assembled gelled solutions. It was found that silk/wool blended solutions can form hydrogels at different mixing ratios, with perfectly interconnected gel structure when the wool content was less than 30 weight percent (wt %) for the first group (ultrasonication), and 10 wt % for the second group (natural gel). Differential scanning calorimetry (DSC) and temperature modulated DSC (TMDSC) were used to confirm that the fibroin/keratin hydrogel system was well-blended without phase separation. Fourier transform infrared spectroscopy (FTIR) was used to investigate the secondary structures of blended protein gels. It was found that intermolecular β-sheet contents significantly increase as the system contains more silk for both groups of samples, resulting in stable crystalline cross-linkers in the blended hydrogel structures. Scanning electron microscopy (SEM) and atomic force microscopy (AFM) were used to analyze the samples’ characteristic morphology on both micro- and nanoscales, which showed that ultrasonic waves can significantly enhance the cross-linker formation and avoid phase separation between silk and keratin molecules in the blended systems. With the ability to form cross-linkages non-chemically, these silk/wool hydrogels may be economically useful for various biomedical applications, thanks to the good biocompatibility of protein molecules and the various characteristics of hydrogel systems. PMID:27618011

Silk-Based Biomaterials for Sustained Drug Delivery

PubMed Central

Yucel, Tuna; Lovett, Michael L.; Kaplan, David L.

2014-01-01

Silk presents a rare combination of desirable properties for sustained drug delivery, including aqueous-based purification and processing options without chemical cross-linkers, compatibility with common sterilization methods, controllable and surface-mediated biodegradation into non-inflammatory by-products, biocompatibility, utility in drug stabilization, and robust mechanical properties. A versatile silk-based toolkit is currently available for sustained drug delivery formulations of small molecule through macromolecular drugs, with a promise to mitigate several drawbacks associated with other degradable sustained delivery technologies in the market. Silk-based formulations utilize silk’s well-defined nano- through microscale structural hierarchy, stimuli-responsive self-assembly pathways and crystal polymorphism, as well as sequence and genetic modification options towards targeted pharmaceutical outcomes. Furthermore, by manipulating the interactions between silk and drug molecules, near-zero order sustained release may be achieved through diffusion- and degradation-based release mechanisms. Because of these desirable properties, there has been increasing industrial interest in silk-based drug delivery systems currently at various stages of the developmental pipeline from pre-clinical to FDA-approved products. Here, we discuss the unique aspects of silk technology as a sustained drug delivery platform and highlight the current state of the art in silk-based drug delivery. We also offer a potential early development pathway for silk-based sustained delivery products. PMID:24910193

Machine-Washable PEDOT:PSS Dyed Silk Yarns for Electronic Textiles.

PubMed

Ryan, Jason D; Mengistie, Desalegn Alemu; Gabrielsson, Roger; Lund, Anja; Müller, Christian

2017-03-15

Durable, electrically conducting yarns are a critical component of electronic textiles (e-textiles). Here, such yarns with exceptional wear and wash resistance are realized through dyeing silk from the silkworm Bombyx mori with the conjugated polymer:polyelectrolyte complex PEDOT:PSS. A high Young's modulus of approximately 2 GPa combined with a robust and scalable dyeing process results in up to 40 m long yarns that maintain their bulk electrical conductivity of approximately 14 S cm -1 when experiencing repeated bending stress as well as mechanical wear during sewing. Moreover, a high degree of ambient stability is paired with the ability to withstand both machine washing and dry cleaning. For the potential use for e-textile applications to be illustrated, an in-plane thermoelectric module that comprises 26 p-type legs is demonstrated by embroidery of dyed silk yarns onto a piece of felted wool fabric.

Machine-Washable PEDOT:PSS Dyed Silk Yarns for Electronic Textiles

PubMed Central

2017-01-01

Durable, electrically conducting yarns are a critical component of electronic textiles (e-textiles). Here, such yarns with exceptional wear and wash resistance are realized through dyeing silk from the silkworm Bombyx mori with the conjugated polymer:polyelectrolyte complex PEDOT:PSS. A high Young’s modulus of approximately 2 GPa combined with a robust and scalable dyeing process results in up to 40 m long yarns that maintain their bulk electrical conductivity of approximately 14 S cm–1 when experiencing repeated bending stress as well as mechanical wear during sewing. Moreover, a high degree of ambient stability is paired with the ability to withstand both machine washing and dry cleaning. For the potential use for e-textile applications to be illustrated, an in-plane thermoelectric module that comprises 26 p-type legs is demonstrated by embroidery of dyed silk yarns onto a piece of felted wool fabric. PMID:28245105

Preliminary investigation of airgap electrospun silk-fibroin-based structures for ligament analogue engineering.

PubMed

Sell, S A; McClure, M J; Ayres, C E; Simpson, D G; Bowlin, G L

2011-01-01

The process of electrospinning has proven to be highly beneficial for use in a number of tissue-engineering applications due to its ease of use, flexibility and tailorable properties. There have been many publications on the creation of aligned fibrous structures created through various forms of electrospinning, most involving the use of a metal target rotating at high speeds. This work focuses on the use of a variation known as airgap electrospinning, which does not use a metal collecting target but rather a pair of grounded electrodes equidistant from the charged polymer solution to create highly aligned 3D structures. This study involved a preliminary investigation and comparison of traditionally and airgap electrospun silk-fibroin-based ligament constructs. Structures were characterized with SEM and alignment FFT, and underwent porosity, permeability, and mechanical anisotropy evaluation. Preliminary cell culture with human dermal fibroblasts was performed to determine the degree of cellular orientation and penetration. Results showed airgap electrospun structures to be anisotropic with significantly increased porosity and cellular penetration compared to their traditionally electrospun counterparts.

Exploring the effects of electrospinning processing protocols on fiber surface morphology and polymer chain conformation

NASA Astrophysics Data System (ADS)

Stephens, Jean S.

Electrospinning is a fiber formation technique that uses electrostatic forces to create continuous, nanometer diameter fibers. The work presented here focuses on the continuing efforts to build a stronger fundamental understanding of electrospinning by exploring structure/property/process relationships by investigating the effects of process protocols on fiber surface morphology and polymer chain conformation. By varying the processing parameters it has been possible to produce fibers with unique surface features, microtextured/nanoporous fibers and nanowebs. In the microtextured/nanoporous fiber studies, changing the solution concentration, solvent volatility, and relative humidity was found to alter the size, shape, and distribution of pores on the fiber surface. The mechanisms that can explain the pore formation and texturing on the surface of the fibers are phase separation (aggregation into polymer rich and polymer lean regions) and breath figures (evaporative cooling and vapor condensation). Through a judicious choice of the electrospinning processing parameters we have also been able to create "web" like structures of nanofibers (5--25 nm) from collagen, dragline silk analog, nylon, and denatured collagen. Electrostatic repulsion and thin film dewetting are thought to be responsible for the formation of the nanowebs. These unique structures were characterized using FESEM, TEM, OM, and AFM. Raman spectroscopy, initially developed as a "real time" characterization technique to study electrospun fiber formation, has also been used to investigate the effect of electrospinning on the chain conformation of bioinspired polymers. Comparing the spectrum of the bulk material to that of the electrospun material identified conformational changes in nylon 6 and dragline silk analog. The conformational change in nylon 6 (alpha-form to gamma-form) results from the stresses induced on the electrospinning jet during fiber formation, whereas the conformational change in the silk analog (beta-sheet to alpha-helical) result from electric field assembling of the charged a-helical segments of the protein polymer in solution. The investigations described here have allowed us to build a virtual database of the processing conditions needed to create materials for tissue engineering constructs. Electrospun collagen membranes have been used in preliminary cell attachment studies. From the trials it was observed that the cells migrated into the membranes indicating that the membranes are suitable for tissue engineering scaffolds.

Silk-elastin-like protein polymer matrix for intraoperative delivery of an oncolytic vaccinia virus.

PubMed

Price, Daniel L; Li, Pingdong; Chen, Chun-Hao; Wong, Danni; Yu, Zhenkun; Chen, Nanhai G; Yu, Yong A; Szalay, Aladar A; Cappello, Joseph; Fong, Yuman; Wong, Richard J

2016-02-01

Oncolytic viral efficacy may be limited by the penetration of the virus into tumors. This may be enhanced by intraoperative application of virus immediately after surgical resection. Oncolytic vaccinia virus GLV-1h68 was delivered in silk-elastin-like protein polymer (SELP) in vitro and in vivo in anaplastic thyroid carcinoma cell line 8505c in nude mice. GLV-1h68 in SELP infected and lysed anaplastic thyroid cancer cells in vitro equally as effectively as in phosphate-buffered saline (PBS), and at 1 week retains a thousand fold greater infectious plaque-forming units. In surgical resection models of residual tumor, GLV-1h68 in SELP improves tumor control and shows increased viral β-galactosidase expression as compared to PBS. The use of SELP matrix for intraoperative oncolytic viral delivery protects infectious viral particles from degradation, facilitates sustained viral delivery and transgene expression, and improves tumor control. Such optimization of methods of oncolytic viral delivery may enhance therapeutic outcomes. © 2014 Wiley Periodicals, Inc.

Silk-elastin-like protein polymer matrix for intraoperative delivery of an oncolytic vaccinia virus

PubMed Central

Price, Daniel L.; Li, Pingdong; Chen, Chun-Hao; Wong, Danni; Yu, Zhenkun; Chen, Nanhai G.; Yu, Yong A.; Szalay, Aladar A.; Cappello, Joseph; Fong, Yuman; Wong, Richard J.

2016-01-01

Background Oncolytic viral efficacy may be limited by the penetration of the virus into tumors. This may be enhanced by intraoperative application of virus immediately after surgical resection. Methods Oncolytic vaccinia virus GLV-1h68 was delivered in silk-elastin-like protein polymer (SELP) in vitro and in vivo in anaplastic thyroid carcinoma cell line 8505c in nude mice. Results GLV-1h68 in SELP infected and lysed anaplastic thyroid cancer cells in vitro equally as effectively as in phosphate-buffered saline (PBS), and at 1 week retains a thousand fold greater infectious plaque-forming units. In surgical resection models of residual tumor, GLV-1h68 in SELP improves tumor control and shows increased viral β-galactosidase expression as compared to PBS. Conclusion The use of SELP matrix for intraoperative oncolytic viral delivery protects infectious viral particles from degradation, facilitates sustained viral delivery and transgene expression, and improves tumor control. Such optimization of methods of oncolytic viral delivery may enhance therapeutic outcomes. PMID:25244076

Formulation of Biologically-Inspired Silk-Based Drug Carriers for Pulmonary Delivery Targeted for Lung Cancer

PubMed Central

Kim, Sally Yunsun; Naskar, Deboki; Kundu, Subhas C.; Bishop, David P.; Doble, Philip A.; Boddy, Alan V.; Chan, Hak-Kim; Wall, Ivan B.; Chrzanowski, Wojciech

2015-01-01

The benefits of using silk fibroin, a major protein in silk, are widely established in many biomedical applications including tissue regeneration, bioactive coating and in vitro tissue models. The properties of silk such as biocompatibility and controlled degradation are utilized in this study to formulate for the first time as carriers for pulmonary drug delivery. Silk fibroin particles are spray dried or spray-freeze-dried to enable the delivery to the airways via dry powder inhalers. The addition of excipients such as mannitol is optimized for both the stabilization of protein during the spray-freezing process as well as for efficient dispersion using an in vitro aerosolisation impactor. Cisplatin is incorporated into the silk-based formulations with or without cross-linking, which show different release profiles. The particles show high aerosolisation performance through the measurement of in vitro lung deposition, which is at the level of commercially available dry powder inhalers. The silk-based particles are shown to be cytocompatible with A549 human lung epithelial cell line. The cytotoxicity of cisplatin is demonstrated to be enhanced when delivered using the cross-linked silk-based particles. These novel inhalable silk-based drug carriers have the potential to be used as anti-cancer drug delivery systems targeted for the lungs. PMID:26234773

GC/MS-based metabolomic studies reveal key roles of glycine in regulating silk synthesis in silkworm, Bombyx mori.

PubMed

Chen, Quanmei; Liu, Xinyu; Zhao, Ping; Sun, Yanhui; Zhao, Xinjie; Xiong, Ying; Xu, Guowang; Xia, Qingyou

2015-02-01

Metabolic profiling of silkworm, especially the factors that affect silk synthesis at the metabolic level, is little known. Herein, metabolomic method based on gas chromatography-mass spectrometry was applied to identify key metabolic changes in silk synthesis deficient silkworms. Forty-six differential metabolites were identified in Nd group with the defect of silk synthesis. Significant changes in the levels of glycine and uric acid (up-regulation), carbohydrates and free fatty acids (down-regulation) were observed. The further metabolomics of silk synthesis deficient silkworms by decreasing silk proteins synthesis using knocking out fibroin heavy chain gene or extirpating silk glands operation showed that the changes of the metabolites were almost consistent with those of the Nd group. Furthermore, the increased silk yields by supplying more glycine or its related metabolite confirmed that glycine is a key metabolite to regulate silk synthesis. These findings provide important insights into the regulation between metabolic profiling and silk synthesis. Copyright © 2014 Elsevier Ltd. All rights reserved.

Buyid Silk and the Tale of Bibi Shahrbanu: Identification of Biomarkers of Artificial Aging (Forgery) of Silk.

PubMed

Moini, Mehdi; Rollman, Christopher M

2017-10-03

Buyid silk forgery is one of the most famous silk forgeries in the world. In 1924-1925, excavation of the Bibi Shahrbanu site in Iran unearthed several silk textiles. The silks were thought to be of the Buyid period (934-1062 BCE) of the Persian Empire and have since been known as the "Buyid silks". In the 1930s, more silk appeared and was reported as being from the Buyid period as well. Controversy over the authenticity of these silks escalated after the purchase of the silks by museums throughout the world. Extensive investigations of several of these silks have been conducted over the years with respect to iconography, weaving patterns, dyes/mordant, style, and even radiocarbon dating. It was found that most of the silks are not from Buyid period. To test the authenticity of these silk fabrics, the recently developed silk dating technique using amino acid racemization (AAR) in conjunction with capillary electrophoresis mass spectrometry was applied to 13 Buyid silk specimens from the Textile Museum collections. Among these silk specimens, the AAR ratios of only one specimen were consistent with authentic silk fabrics collected from various museums. In addition, the aspartic acid racemization ratio of this specimen was also consistent with its 14 C dating. The other "Buyid silks" showed excessive levels of amino acid racemization not only for aspartic acid, but also for phenylalanine and tyrosine, inconsistent with racemization rates of these amino acids in authentic historical silk fabrics. Treatment of modern silk with a base at different pH and temperature reproduced the AAR pattern of the Buyid silks, implying that chemical treatment with a base at relatively high temperatures was perhaps the method used to artificially age these fabrics. The results imply that the racemization ratios of aspartic acid, phenylalanine, and tyrosine can be used as biomarkers for identification of naturally versus artificially aged silk.

Enhanced mechanical properties of thermosensitive chitosan hydrogel by silk fibers for cartilage tissue engineering.

PubMed

Mirahmadi, Fereshteh; Tafazzoli-Shadpour, Mohammad; Shokrgozar, Mohammad Ali; Bonakdar, Shahin

2013-12-01

Articular cartilage has limited repair capability following traumatic injuries and current methods of treatment remain inefficient. Reconstructing cartilage provides a new way for cartilage repair and natural polymers are often used as scaffold because of their biocompatibility and biofunctionality. In this study, we added degummed chopped silk fibers and electrospun silk fibers to the thermosensitive chitosan/glycerophosphate hydrogels to reinforce two hydrogel constructs which were used as scaffold for hyaline cartilage regeneration. The gelation temperature and gelation time of hydrogel were analyzed by the rheometer and vial tilting method. Mechanical characterization was measured by uniaxial compression, indentation and dynamic mechanical analysis assay. Chondrocytes were then harvested from the knee joint of the New Zealand white rabbits and cultured in constructs. The cell proliferation, viability, production of glycosaminoglycans and collagen type II were assessed. The results showed that mechanical properties of the hydrogel were significantly enhanced when a hybrid with two layers of electrospun silk fibers was made. The results of GAG and collagen type II in cell-seeded scaffolds indicate support of the chondrogenic phenotype for chondrocytes with a significant increase in degummed silk fiber-hydrogel composite for GAG content and in two-layer electrospun fiber-hydrogel composite for Col II. It was concluded that these two modified scaffolds could be employed for cartilage tissue engineering. © 2013.

Functional regeneration of ligament-bone interface using a triphasic silk-based graft.

PubMed

Li, Hongguo; Fan, Jiabing; Sun, Liguo; Liu, Xincheng; Cheng, Pengzhen; Fan, Hongbin

2016-11-01

The biodegradable silk-based scaffold with unique mechanical property and biocompatibility represents a favorable ligamentous graft for tissue-engineering anterior cruciate ligament (ACL) reconstruction. However, the low efficiency of ligament-bone interface restoration barriers the isotropic silk graft to common ACL therapeutics. To enhance the regeneration of the silk-mediated interface, we developed a specialized stratification approach implementing a sequential modification on isotropic silk to constitute a triphasic silk-based graft in which three regions respectively referring to ligament, cartilage and bone layers of interface were divided, followed by respective biomaterial coating. Furthermore, three types of cells including bone marrow mesenchymal stem cells (BMSCs), chondrocytes and osteoblasts were respectively seeded on the ligament, cartilage and bone region of the triphasic silk graft, and the cell/scaffold complex was rolled up as a multilayered graft mimicking the stratified structure of native ligament-bone interface. In vitro, the trilineage cells loaded on the triphasic silk scaffold revealed a high proliferative capacity as well as enhanced differentiation ability into their corresponding cell lineage. 24 weeks postoperatively after the construct was implanted to repair the ACL defect in rabbit model, the silk-based ligamentous graft exhibited the enhancement of osseointegration detected by a robust pullout force and formation of three-layered structure along with conspicuously corresponding matrix deposition via micro-CT and histological analysis. These findings potentially broaden the application of silk-based ligamentous graft for ACL reconstruction and further large animal study. Copyright © 2016 Elsevier Ltd. All rights reserved.

Applications of micro-SAXS/WAXS to study polymer fibers

NASA Astrophysics Data System (ADS)

Riekel, C.

2003-01-01

Instrumentation and selected applications for X-ray microdiffraction experiments on polymer and biopolymer fibers at the European Synchrotron Radiation Facility (ESRF) microfocus beamline are reviewed. Combined SAXS/WAXS experiments can be performed on single fibers with a beam size down to about 5 μm. WAXS experiments can be performed down to about 2 μm and in exceptional cases down to 0.1 μm beam size. The instrumental possibilities are demonstrated for the production line of spider silk.

Glycopolymer functionalization of engineered spider silk protein-based materials for improved cell adhesion.

PubMed

Hardy, John G; Pfaff, André; Leal-Egaña, Aldo; Müller, Axel H E; Scheibel, Thomas R

2014-07-01

Silk protein-based materials are promising biomaterials for application as tissue scaffolds, due to their processability, biocompatibility, and biodegradability. The preparation of films composed of an engineered spider silk protein (eADF4(C16)) and their functionalization with glycopolymers are described. The glycopolymers bind proteins found in the extracellular matrix, providing a biomimetic coating on the films that improves cell adhesion to the surfaces of engineered spider silk films. Such silk-based materials have potential as coatings for degradable implantable devices. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Proteins--The Basis of Life

ERIC Educational Resources Information Center

Wrigley, Colin

2012-01-01

Proteins are a diverse class of biochemical macromolecules, including substances as (apparently) unrelated as silk and sinew, hair and horn, feathers and flagella, enzymes and epidermis, gelatine (jelly) and gluten and gore, spider web, meat and fish muscle. Yet they are unified by being polymers of amino acids. Discovery of the nature of proteins…

Flexible and wearable electronic silk fabrics for human physiological monitoring

NASA Astrophysics Data System (ADS)

Mao, Cuiping; Zhang, Huihui; Lu, Zhisong

2017-09-01

The development of textile-based devices for human physiological monitoring has attracted tremendous interest in recent years. However, flexible physiological sensing elements based on silk fabrics have not been realized. In this paper, ZnO nanorod arrays are grown in situ on reduced graphene oxide-coated silk fabrics via a facile electro-deposition method for the fabrication of silk-fabric-based mechanical sensing devices. The data show that well-aligned ZnO nanorods with hexagonal wurtzite crystalline structures are synthesized on the conductive silk fabric surface. After magnetron sputtering of gold electrodes, silk-fabric-based devices are produced and applied to detect periodic bending and twisting. Based on the electric signals, the deformation and release processes can be easily differentiated. Human arterial pulse and respiration can also be real-time monitored to calculate the pulse rate and respiration frequency, respectively. Throat vibrations during coughing and singing are detected to demonstrate the voice recognition capability. This work may not only help develop silk-fabric-based mechanical sensing elements for potential applications in clinical diagnosis, daily healthcare monitoring and voice recognition, but also provide a versatile method for fabricating textile-based flexible electronic devices.

PEGylated Silk Nanoparticles for Anticancer Drug Delivery.

PubMed

Wongpinyochit, Thidarat; Uhlmann, Petra; Urquhart, Andrew J; Seib, F Philipp

2015-11-09

Silk has a robust clinical track record and is emerging as a promising biopolymer for drug delivery, including its use as nanomedicine. However, silk-based nanomedicines still require further refinements for full exploitation of their potential; the application of "stealth" design principals is especially necessary to support their evolution. The aim of this study was to develop and examine the potential of PEGylated silk nanoparticles as an anticancer drug delivery system. We first generated B. mori derived silk nanoparticles by driving β-sheet assembly (size 104 ± 1.7 nm, zeta potential -56 ± 5.6 mV) using nanoprecipitation. We then surface grafted polyethylene glycol (PEG) to the fabricated silk nanoparticles and verified the aqueous stability and morphology of the resulting PEGylated silk nanoparticles. We assessed the drug loading and release behavior of these nanoparticles using clinically established and emerging anticancer drugs. Overall, PEGylated silk nanoparticles showed high encapsulation efficiency (>93%) and a pH-dependent release over 14 days. Finally, we demonstrated significant cytotoxicity of drug loaded silk nanoparticles applied as single and combination nanomedicines to human breast cancer cells. In conclusion, these results, taken together with prior silk nanoparticle data, support a viable future for silk-based nanomedicines.

Heat Capacity of B. Mori Silk Fibroin Based on the Vibrational-Motion of Poly(amino acid)s.

NASA Astrophysics Data System (ADS)

Pyda, Marek; Hu, Xiao; Cebe, Peggy

2009-03-01

Bombyx mori silk fibroin heat capacities with and without water have been determined based on the vibrational motions of poly(amino acid)s and water, using the Advanced Thermal Analysis System (ATHAS) Data Bank. The heat capacities, Cp, of dry silk and silk-water were linked to their vibrational spectra based on the group and skeletal vibration contributions. For dry silk, the experimental and calculated Cp agree to better than ±3% between 200 K and 435 K. The heat capacity of the solid silk-water system, below the glass transition, was estimated from a sum of linear combinations of the molar fractions of the vibrational heat capacities of dry silk and glassy water. Calculations are compared to experimental data obtained from calorimetric methods, using hermetic and non-hermetic pans. The approach presented allows one to predict the low temperature vibrational heat capacity for dry silk and for the silk-water system down to zero kelvin, and, together with an extension to higher temperatures, above the glass transition. This can be used as a reference baseline for quantitative thermal analysis of this biomaterial..

The processing and heterostructuring of silk with light

NASA Astrophysics Data System (ADS)

Sidhu, Mehra S.; Kumar, Bhupesh; Singh, Kamal P.

2017-09-01

Spider silk is a tough, elastic and lightweight biomaterial, although there is a lack of tools available for non-invasive processing of silk structures. Here we show that nonlinear multiphoton interactions of silk with few-cycle femtosecond pulses allow the processing and heterostructuring of the material in ambient air. Two qualitatively different responses, bulging by multiphoton absorption and plasma-assisted ablation, are observed for low- and high-peak intensities, respectively. Plasma ablation allows us to make localized nanocuts, microrods, nanotips and periodic patterns with minimal damage while preserving molecular structure. The bulging regime facilitates confined bending and microwelding of silk with materials such as metal, glass and Kevlar with strengths comparable to pristine silk. Moreover, analysis of Raman bands of microwelded joints reveals that the polypeptide backbone remains intact while perturbing its weak hydrogen bonds. Using this approach, we fabricate silk-based functional topological microstructures, such as Mobiüs strips, chiral helices and silk-based sensors.

The processing and heterostructuring of silk with light.

PubMed

Sidhu, Mehra S; Kumar, Bhupesh; Singh, Kamal P

2017-09-01

Spider silk is a tough, elastic and lightweight biomaterial, although there is a lack of tools available for non-invasive processing of silk structures. Here we show that nonlinear multiphoton interactions of silk with few-cycle femtosecond pulses allow the processing and heterostructuring of the material in ambient air. Two qualitatively different responses, bulging by multiphoton absorption and plasma-assisted ablation, are observed for low- and high-peak intensities, respectively. Plasma ablation allows us to make localized nanocuts, microrods, nanotips and periodic patterns with minimal damage while preserving molecular structure. The bulging regime facilitates confined bending and microwelding of silk with materials such as metal, glass and Kevlar with strengths comparable to pristine silk. Moreover, analysis of Raman bands of microwelded joints reveals that the polypeptide backbone remains intact while perturbing its weak hydrogen bonds. Using this approach, we fabricate silk-based functional topological microstructures, such as Mobiüs strips, chiral helices and silk-based sensors.

Silk-elastin-like protein biomaterials for the controlled delivery of therapeutics.

PubMed

Huang, Wenwen; Rollett, Alexandra; Kaplan, David L

2015-05-01

Genetically engineered biomaterials are useful for controlled delivery owing to their rational design, tunable structure-function, biocompatibility, degradability and target specificity. Silk-elastin-like proteins (SELPs), a family of genetically engineered recombinant protein polymers, possess these properties. Additionally, given the benefits of combining semi-crystalline silk-blocks and elastomeric elastin-blocks, SELPs possess multi-stimuli-responsive properties and tunability, thereby becoming promising candidates for targeted cancer therapeutics delivery and controlled gene release. An overview of SELP biomaterials for drug delivery and gene release is provided. Biosynthetic strategies used for SELP production, fundamental physicochemical properties and self-assembly mechanisms are discussed. The review focuses on sequence-structure-function relationships, stimuli-responsive features and current and potential drug delivery applications. The tunable material properties allow SELPs to be pursued as promising biomaterials for nanocarriers and injectable drug release systems. Current applications of SELPs have focused on thermally-triggered biomaterial formats for the delivery of therapeutics, based on local hyperthermia in tumors or infections. Other prominent controlled release applications of SELPs as injectable hydrogels for gene release have also been pursued. Further biomedical applications that utilize other stimuli to trigger the reversible material responses of SELPs for targeted delivery, including pH, ionic strength, redox, enzymatic stimuli and electric field, are in progress. Exploiting these additional stimuli-responsive features will provide a broader range of functional biomaterials for controlled therapeutics release and tissue regeneration.

Silks produced by insect labial glands

PubMed Central

Sutherland, Tara

2008-01-01

Insect silks are secreted from diverse gland types; this chapter deals with the silks produced by labial glands of Holometabola (insects with pupa in their life cycle). Labial silk glands are composed of a few tens or hundreds of large polyploid cells that secrete polymerizing proteins which are stored in the gland lumen as a semi-liquid gel. Polymerization is based on weak molecular interactions between repetitive amino acid motifs present in one or more silk proteins; cross-linking by disulfide bonds may be important in the silks spun under water. The mechanism of long-term storage of the silk dope inside the glands and its conversion into the silk fiber during spinning is not fully understood. The conversion occurs within seconds at ambient temperature and pressure, under minimal drawing force and in some cases under water. The silk filament is largely built of proteins called fibroins and in Lepidoptera and Trichoptera coated by glue-type proteins known as sericins. Silks often contain small amounts of additional proteins of poorly known function. The silk components controlling dope storage and filament formation seem to be conserved at the level of orders, while the nature of polymerizing motifs in the fibroins, which determine the physical properties of silk, differ at the level of family and even genus. Most silks are based on fibroin β-sheets interrupted with other structures such as α-helices but the silk proteins of certain sawflies have predominantly a collagen-like or polyglycine II arrangement and the silks of social Hymenoptera are formed from proteins in a coiled coil arrangement. PMID:19221523

Potential applications of silk sericin, a natural protein from textile industry by-products.

PubMed

Aramwit, Pornanong; Siritientong, Tippawan; Srichana, Teerapol

2012-03-01

Silk is composed of two major proteins, fibroin (fibrous protein) and sericin (globular, gumming protein). Fibroin has been used in textile manufacturing and for several biomaterial applications, whereas sericin is considered a waste material in the textile industry. Sericin has recently been found to activate the proliferation of several cell-lines and has also shown various biological activities. Sericin can form a gel by itself; however, after mixing with other polymers and cross-linking it can form a film or a scaffold with good characteristics that can be used in the cosmetic and pharmaceutical industries. Sericin is proven to cause no immunological responses, which has resulted in a more acceptable material for biological applications.

A model for structural changes of reconstituted fibroin gels during deformation

NASA Astrophysics Data System (ADS)

Jin, Peiran; Olmsted, Peter; Georgetown University, Physics Department Team

Silk from silkworms has been used in the textile industry for thousands of years. Recently, a physical electrogel(e-gel) was made by reconstituting Bombyx mori silk into stable aqueous solutions and then applying small DC electric field. The e-gels exhibit distinctive strain hardening and are partially recoverable from strain. To explain these phenomena, we build a coarse grained model of fibroin protein polymers, which comprise crystallizable domains and amorphous domains. We find that the kinetics of unfolding and folding of crystalline domains changes the number and functionality of crosslinks in the physical network, and thus contributes to the strain hardening of the gel and the non-recoverable strain. Georgetown University and the Ives Foundation.

Phononic band gap and mechanical anisotropy in spider silk

NASA Astrophysics Data System (ADS)

Papadopoulos, Periklis; Gomopoulos, Nikos; Kremer, Friedrich; Fytas, George

2010-03-01

Spider dragline silk is a semi-crystalline biopolymer exhibiting superior properties compared to synthetic polymers with similar chemical structure, such as polyamides. This is ascribed to the hierarchical nanostructure that is created in the spinning duct. During this process the aqueous solution of the two protein constituents of dragline silk is crystallized, while the macromolecules maintain their high orientation, leading to a high value of the Young's modulus (in the order of 10 GPa) along the fiber. We employed spontaneous Brillouin light scattering to measure the longitudinal modulus (M//,,M) along the two symmetry directions of the native fiber with increased (decreased) pre-strain created by stretching (supercontracting after hydration). A strong mechanical anisotropy is found; at about 18% strain M///M˜5. Most important, an unexpected finding is the first observation of a unidirectional hypersonic phononic band gap in biological structures. This relates to the existence of a strain-dependent correlation length of the mechanical modulus in the submicron range along the fiber axis.

Novel fabrication of fluorescent silk utilized in biotechnological and medical applications.

PubMed

Kim, Dong Wook; Lee, Ok Joo; Kim, Seong-Wan; Ki, Chang Seok; Chao, Janet Ren; Yoo, Hyojong; Yoon, Sung-Il; Lee, Jeong Eun; Park, Ye Ri; Kweon, HaeYong; Lee, Kwang Gill; Kaplan, David L; Park, Chan Hum

2015-11-01

Silk fibroin (SF) is a natural polymer widely used and studied for diverse applications in the biomedical field. Recently, genetically modified silks, particularly fluorescent SF fibers, were reported to have been produced from transgenic silkworms. However, they are currently limited to textile manufacturing. To expand the use of transgenic silkworms for biomedical applications, a solution form of fluorescent SF needed to be developed. Here, we describe a novel method of preparing a fluorescent SF solution and demonstrate long-term fluorescent function up to one year after subcutaneous insertion. We also show that fluorescent SF labeled p53 antibodies clearly identify HeLa cells, indicating the applicability of fluorescent SF to cancer detection and bio-imaging. Furthermore, we demonstrate the intraoperative use of fluorescent SF in an animal model to detect a small esophageal perforation (0.5 mm). This study suggests how fluorescent SF biomaterials can be applied in biotechnology and clinical medicine. Copyright © 2015 Elsevier Ltd. All rights reserved.

Recombinant DNA production of spider silk proteins

PubMed Central

Tokareva, Olena; Michalczechen-Lacerda, Valquíria A; Rech, Elíbio L; Kaplan, David L

2013-01-01

Spider dragline silk is considered to be the toughest biopolymer on Earth due to an extraordinary combination of strength and elasticity. Moreover, silks are biocompatible and biodegradable protein-based materials. Recent advances in genetic engineering make it possible to produce recombinant silks in heterologous hosts, opening up opportunities for large-scale production of recombinant silks for various biomedical and material science applications. We review the current strategies to produce recombinant spider silks. PMID:24119078

Protein Bricks: 2D and 3D Bio-Nanostructures with Shape and Function on Demand.

PubMed

Jiang, Jianjuan; Zhang, Shaoqing; Qian, Zhigang; Qin, Nan; Song, Wenwen; Sun, Long; Zhou, Zhitao; Shi, Zhifeng; Chen, Liang; Li, Xinxin; Mao, Ying; Kaplan, David L; Gilbert Corder, Stephanie N; Chen, Xinzhong; Liu, Mengkun; Omenetto, Fiorenzo G; Xia, Xiaoxia; Tao, Tiger H

2018-05-01

Precise patterning of polymer-based biomaterials for functional bio-nanostructures has extensive applications including biosensing, tissue engineering, and regenerative medicine. Remarkable progress is made in both top-down (based on lithographic methods) and bottom-up (via self-assembly) approaches with natural and synthetic biopolymers. However, most methods only yield 2D and pseudo-3D structures with restricted geometries and functionalities. Here, it is reported that precise nanostructuring on genetically engineered spider silk by accurately directing ion and electron beam interactions with the protein's matrix at the nanoscale to create well-defined 2D bionanopatterns and further assemble 3D bionanoarchitectures with shape and function on demand, termed "Protein Bricks." The added control over protein sequence and molecular weight of recombinant spider silk via genetic engineering provides unprecedented lithographic resolution (approaching the molecular limit), sharpness, and biological functions compared to natural proteins. This approach provides a facile method for patterning and immobilizing functional molecules within nanoscopic, hierarchical protein structures, which sheds light on a wide range of biomedical applications such as structure-enhanced fluorescence and biomimetic microenvironments for controlling cell fate. © 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Artificial Polymeric Scaffolds as Extracellular Matrix Substitutes for Autologous Conjunctival Goblet Cell Expansion

PubMed Central

He, Min; Storr-Paulsen, Thomas; Wang, Annie L.; Ghezzi, Chiara E.; Wang, Siran; Fullana, Matthew; Karamichos, Dimitrios; Utheim, Tor P.; Islam, Rakibul; Griffith, May; Islam, M. Mirazul; Hodges, Robin R.; Wnek, Gary E.; Kaplan, David L.; Dartt, Darlene A.

2016-01-01

Purpose We fabricated and investigated polymeric scaffolds that can substitute for the conjunctival extracellular matrix to provide a substrate for autologous expansion of human conjunctival goblet cells in culture. Methods We fabricated two hydrogels and two silk films: (1) recombinant human collagen (RHC) hydrogel, (2) recombinant human collagen 2-methacryloylxyethyl phosphorylcholine (RHC-MPC) hydrogel, (3) arginine-glycine-aspartic acid (RGD) modified silk, and (4) poly-D-lysine (PDL) coated silk, and four electrospun scaffolds: (1) collagen, (2) poly(acrylic acid) (PAA), (3) poly(caprolactone) (PCL), and (4) poly(vinyl alcohol) (PVA). Coverslips and polyethylene terephthalate (PET) were used for comparison. Human conjunctival explants were cultured on scaffolds for 9 to 15 days. Cell viability, outgrowth area, and the percentage of cells expressing markers for stratified squamous epithelial cells (cytokeratin 4) and goblet cells (cytokeratin 7) were determined. Results Most of cells grown on all scaffolds were viable except for PCL in which only 3.6 ± 2.2% of the cells were viable. No cells attached to PVA scaffold. The outgrowth was greatest on PDL-silk and PET. Outgrowth was smallest on PCL. All cells were CK7-positive on RHC-MPC while 84.7 ± 6.9% of cells expressed CK7 on PDL-silk. For PCL, 87.10 ± 3.17% of cells were CK7-positive compared to PET where 67.10 ± 12.08% of cells were CK7-positive cells. Conclusions Biopolymer substrates in the form of hydrogels and silk films provided for better adherence, proliferation, and differentiation than the electrospun scaffolds and could be used for conjunctival goblet cell expansion for eventual transplantation once undifferentiated and stratified squamous cells are included. Useful polymer scaffold design characteristics have emerged from this study. PMID:27832279

Artificial Polymeric Scaffolds as Extracellular Matrix Substitutes for Autologous Conjunctival Goblet Cell Expansion.

PubMed

He, Min; Storr-Paulsen, Thomas; Wang, Annie L; Ghezzi, Chiara E; Wang, Siran; Fullana, Matthew; Karamichos, Dimitrios; Utheim, Tor P; Islam, Rakibul; Griffith, May; Islam, M Mirazul; Hodges, Robin R; Wnek, Gary E; Kaplan, David L; Dartt, Darlene A

2016-11-01

We fabricated and investigated polymeric scaffolds that can substitute for the conjunctival extracellular matrix to provide a substrate for autologous expansion of human conjunctival goblet cells in culture. We fabricated two hydrogels and two silk films: (1) recombinant human collagen (RHC) hydrogel, (2) recombinant human collagen 2-methacryloylxyethyl phosphorylcholine (RHC-MPC) hydrogel, (3) arginine-glycine-aspartic acid (RGD) modified silk, and (4) poly-D-lysine (PDL) coated silk, and four electrospun scaffolds: (1) collagen, (2) poly(acrylic acid) (PAA), (3) poly(caprolactone) (PCL), and (4) poly(vinyl alcohol) (PVA). Coverslips and polyethylene terephthalate (PET) were used for comparison. Human conjunctival explants were cultured on scaffolds for 9 to 15 days. Cell viability, outgrowth area, and the percentage of cells expressing markers for stratified squamous epithelial cells (cytokeratin 4) and goblet cells (cytokeratin 7) were determined. Most of cells grown on all scaffolds were viable except for PCL in which only 3.6 ± 2.2% of the cells were viable. No cells attached to PVA scaffold. The outgrowth was greatest on PDL-silk and PET. Outgrowth was smallest on PCL. All cells were CK7-positive on RHC-MPC while 84.7 ± 6.9% of cells expressed CK7 on PDL-silk. For PCL, 87.10 ± 3.17% of cells were CK7-positive compared to PET where 67.10 ± 12.08% of cells were CK7-positive cells. Biopolymer substrates in the form of hydrogels and silk films provided for better adherence, proliferation, and differentiation than the electrospun scaffolds and could be used for conjunctival goblet cell expansion for eventual transplantation once undifferentiated and stratified squamous cells are included. Useful polymer scaffold design characteristics have emerged from this study.

Genome sequence of the Japanese oak silk moth, Antheraea yamamai: the first draft genome in the family Saturniidae

PubMed Central

Kim, Seong-Ryul; Kwak, Woori; Kim, Hyaekang; Kim, Kee-Young; Kim, Su-Bae; Choi, Kwang-Ho; Kim, Seong-Wan; Hwang, Jae-Sam; Kim, Minjee; Kim, Iksoo; Goo, Tae-Won

2018-01-01

Abstract Background Antheraea yamamai, also known as the Japanese oak silk moth, is a wild species of silk moth. Silk produced by A. yamamai, referred to as tensan silk, shows different characteristics such as thickness, compressive elasticity, and chemical resistance compared with common silk produced from the domesticated silkworm, Bombyx mori. Its unique characteristics have led to its use in many research fields including biotechnology and medical science, and the scientific as well as economic importance of the wild silk moth continues to gradually increase. However, no genomic information for the wild silk moth, including A. yamamai, is currently available. Findings In order to construct the A. yamamai genome, a total of 147G base pairs using Illumina and Pacbio sequencing platforms were generated, providing 210-fold coverage based on the 700-Mb estimated genome size of A. yamamai. The assembled genome of A. yamamai was 656 Mb (>2 kb) with 3675 scaffolds, and the N50 length of assembly was 739 Kb with a 34.07% GC ratio. Identified repeat elements covered 37.33% of the total genome, and the completeness of the constructed genome assembly was estimated to be 96.7% by Benchmarking Universal Single-Copy Orthologs v2 analysis. A total of 15 481 genes were identified using Evidence Modeler based on the gene prediction results obtained from 3 different methods (ab initio, RNA-seq-based, known-gene-based) and manual curation. Conclusions Here we present the genome sequence of A. yamamai, the first genome sequence of the wild silk moth. These results provide valuable genomic information, which will help enrich our understanding of the molecular mechanisms relating to not only specific phenotypes such as wild silk itself but also the genomic evolution of Saturniidae. PMID:29186418

Silk as a biocohesive sacrificial binder in the fabrication of hydroxyapatite load bearing scaffolds.

PubMed

McNamara, Stephanie L; Rnjak-Kovacina, Jelena; Schmidt, Daniel F; Lo, Tim J; Kaplan, David L

2014-08-01

Limitations of current clinical methods for bone repair continue to fuel the demand for a high strength, bioactive bone replacement material. Recent attempts to produce porous scaffolds for bone regeneration have been limited by the intrinsic weakness associated with high porosity materials. In this study, ceramic scaffold fabrication techniques for potential use in load-bearing bone repairs have been developed using naturally derived silk from Bombyx mori. Silk was first employed for ceramic grain consolidation during green body formation, and later as a sacrificial polymer to impart porosity during sintering. These techniques allowed preparation of hydroxyapatite (HA) scaffolds that exhibited a wide range of mechanical and porosity profiles, with some displaying unusually high compressive strength up to 152.4 ± 9.1 MPa. Results showed that the scaffolds exhibited a wide range of compressive strengths and moduli (8.7 ± 2.7 MPa to 152.4 ± 9.1 MPa and 0.3 ± 0.1 GPa to 8.6 ± 0.3 GPa) with total porosities of up to 62.9 ± 2.7% depending on the parameters used for fabrication. Moreover, HA-silk scaffolds could be molded into large, complex shapes, and further machined post-sinter to generate specific three-dimensional geometries. Scaffolds supported bone marrow-derived mesenchymal stem cell attachment and proliferation, with no signs of cytotoxicity. Therefore, silk-fabricated HA scaffolds show promise for load bearing bone repair and regeneration needs. Copyright © 2014 Elsevier Ltd. All rights reserved.

Silk as a biocohesive sacrificial binder in the fabrication of hydroxyapatite load bearing scaffolds

PubMed Central

McNamara, Stephanie L.; Rnjak-Kovacina, Jelena; Schmidt, Daniel; Lo, Tim J.; Kaplan, David L.

2014-01-01

Limitations of current clinical methods for bone repair continue to fuel the demand for a high strength, bioactive bone replacement material. Recent attempts to produce porous scaffolds for bone regeneration have been limited by the intrinsic weakness associated with high porosity materials. In this study, ceramic scaffold fabrication techniques for potential use in load-bearing bone repairs have been developed using naturally derived silk from Bombyx mori. Silk was first employed for ceramic grain consolidation during green body formation, and later as a sacrificial polymer to impart porosity during sintering. These techniques allowed preparation of hydroxyapatite (HA) scaffolds that exhibited a wide range of mechanical and porosity profiles, with some displaying unusually high compressive strength up to 152.4 ± 9.1 MPa. Results showed that the scaffolds exhibited a wide range of compressive strengths and moduli (8.7 ± 2.7 MPa to 152.4 ± 9.1 MPa and 0.3 ± 0.1 GPa to 8.6 ± 0.3 GPa) with total porosities of up to 62.9 ± 2.7% depending on the parameters used for fabrication. Moreover, HA-silk scaffolds could be molded into large, complex shapes, and further machined post-sinter to generate specific three-dimensional geometries. Scaffolds supported bone marrow-derived mesenchymal stem cell attachment and proliferation, with no signs of cytotoxicity. Therefore, silk-fabricated HA scaffolds show promise for load bearing bone repair and regeneration needs. PMID:24881027

A silk-based encapsulation platform for pancreatic islet transplantation improves islet function in vivo.

PubMed

Hamilton, Diana C; Shih, Hank H; Schubert, Richard A; Michie, Sara A; Staats, Paul N; Kaplan, David L; Fontaine, Magali J

2017-03-01

The success of pancreatic islet (PI) transplantation is challenged by PI functional damage during the peritransplantation period. A silk-based encapsulation platform including mesenchymal stromal cells (MSCs) was evaluated for islet cell delivery in vivo. Islet equivalents (IEQs) were transplanted into the epididymal fat pads of mice with streptozotocin-induced diabetes. Three PI combinations were tested: (A) co-encapsulated in silk with MSCs; (b) encapsulated in silk alone; or (c) pelleted. Blood glucose levels were monitored and intraperitoneal glucose tolerance test (IPGTT) was performed upon return to euglycaemia. Grafts were removed for histology and cytokine content analysis. Mice with PI grafts in silk showed a prompt return to euglycaemia. IPGTT was significantly improved with PI in silk with MSCs, compared to PI in silk alone or pelleted. Both Th 1 and Th 2 cytokines were increased in PI grafts in silk, but Th 1 cytokines were decreased significantly with PI and MSC co-encapsulation. Histological analysis showed osteogenesis and chondrogenesis in the silk grafts containing MSCs. Future studies will evaluate MSC stability and function in vivo and improve silk biocompatibility for applications in islet transplantation. Copyright © 2015 John Wiley & Sons, Ltd. Copyright © 2015 John Wiley & Sons, Ltd.

Silk Nanospheres and Microspheres from Silk/PVA Blend Films for Drug Delivery

PubMed Central

Wang, Xiaoqin; Yucel, Tuna; Lu, Qiang; Hu, Xiao; Kaplan, David L.

2009-01-01

Silk fibroin protein-based micro- and nanospheres provide new options for drug delivery due to their biocompatibility, biodegradability and their tunable drug loading and release properties. In the present study, we report a new aqueous-based preparation method for silk spheres with controllable sphere size and shape. The preparation was based on phase separation between silk fibroin and polyvinyl alcohol (PVA) at a weight ratio of 1/1 and 1/4. Water-insoluble silk spheres were easily obtained from the blend in a three step process: (1) air-drying the blend solution into a film, (2) film dissolution in water and (3) removal of residual PVA by subsequent centrifugation. In both cases, the spheres had approximately 30% beta-sheet content and less than 5% residual PVA. Spindle-shaped silk particles, as opposed to the spherical particles formed above, were obtained by stretching the blend films before dissolving in water. Compared to the 1/1 ratio sample, the silk spheres prepared from the 1/4 ratio sample showed a more homogeneous size distribution ranging from 300 nm up to 20 μm. Further studies showed that sphere size and polydispersity could be controlled either by changing the concentration of silk and PVA or by applying ultrasonication on the blend solution. Drug loading was achieved by mixing model drugs in the original silk solution. The distribution and loading efficiency of the drug molecules in silk spheres depended on their hydrophobicity and charge, resulting in different drug release profiles. The entire fabrication procedure could be completed within one day. The only chemical used in the preparation except water was PVA, an FDA-approved ingredient in drug formulations. Silk micro- and nanospheres reported have potential as drug delivery carriers in a variety of biomedical applications. PMID:19945157

Silk Self-Assembly Mechanisms and Control-From Thermodynamics to Kinetics

PubMed Central

Lu, Qiang; Zhu, Hesun; Zhang, Cencen; Zhang, Feng; Zhang, Bing; Kaplan, David L.

2012-01-01

Silkworms and spiders generate fibres that exhibit high strength and extensibility. The underlying mechanisms involved in processing silk proteins into fiber form remain incompletely understood, resulting in the failure to fully recapitulate the remarkable properties of native fibers in vitro from regenerated silk solutions. In the present study, the extensibility and high strength of regenerated silks were achieved by mimicking the natural spinning process. Conformational transitions inside micelles, followed by aggregation of micelles and their stabilization as they relate to the metastable structure of silk are described. Subsequently, the mechanisms to control the formation of nanofibrous structures were elucidated. The results clarify that the self-assembly of silk in aqueous solution is a thermodynamically driven process where kinetics also play a key role. Four key factors, molecular mobility, charge, hydrophilic interactions and concentration underlie the process. Adjusting these factors can balance nanostructure and conformational composition, and be used to achieve silk-based materials with properties comparable to native fibers. These mechanisms suggest new directions to design silk-based multifunctional materials. PMID:22320432

Recombinant protein blends: silk beyond natural design.

PubMed

Dinjaski, Nina; Kaplan, David L

2016-06-01

Recombinant DNA technology and new material concepts are shaping future directions in biomaterial science for the design and production of the next-generation biomaterial platforms. Aside from conventionally used synthetic polymers, numerous natural biopolymers (e.g., silk, elastin, collagen, gelatin, alginate, cellulose, keratin, chitin, polyhydroxyalkanoates) have been investigated for properties and manipulation via bioengineering. Genetic engineering provides a path to increase structural and functional complexity of these biopolymers, and thereby expand the catalog of available biomaterials beyond that which exists in nature. In addition, the integration of experimental approaches with computational modeling to analyze sequence-structure-function relationships is starting to have an impact in the field by establishing predictive frameworks for determining material properties. Herein, we review advances in recombinant DNA-mediated protein production and functionalization approaches, with a focus on hybrids or combinations of proteins; recombinant protein blends or 'recombinamers'. We highlight the potential biomedical applications of fibrous protein recombinamers, such as Silk-Elastin Like Polypeptides (SELPs) and Silk-Bacterial Collagens (SBCs). We also discuss the possibility for the rationale design of fibrous proteins to build smart, stimuli-responsive biomaterials for diverse applications. We underline current limitations with production systems for these proteins and discuss the main trends in systems/synthetic biology that may improve recombinant fibrous protein design and production. Copyright © 2016. Published by Elsevier Ltd.

Tissue response and biodegradation of composite scaffolds prepared from Thai silk fibroin, gelatin and hydroxyapatite.

PubMed

Tungtasana, Hathairat; Shuangshoti, Somruetai; Shuangshoti, Shanop; Kanokpanont, Sorada; Kaplan, David L; Bunaprasert, Tanom; Damrongsakkul, Siriporn

2010-12-01

This work aimed to investigate tissue responses and biodegradation, both in vitro and in vivo, of four types of Bombyx mori Thai silk fibroin based-scaffolds. Thai silk fibroin (SF), conjugated gelatin/Thai silk fibroin (CGSF), hydroxyapatite/Thai silk fibroin (SF4), and hydroxyapatite/conjugated gelatin/Thai silk fibroin (CGSF4) scaffolds were fabricated using salt-porogen leaching, dehydrothermal/chemical crosslinking and an alternate soaking technique for mineralization. In vitro biodegradation in collagenase showed that CGSF scaffolds had the slowest biodegradability, due to the double crosslinking by dehydrothermal and chemical treatments. The hydroxyapatite deposited from alternate soaking separated from the surface of the protein scaffolds when immersed in collagenase. From in vivo biodegradation studies, all scaffolds could still be observed after 12 weeks of implantation in subcutaneous tissue of Wistar rats and also following ISO10993-6: Biological evaluation of medical devices. At 2 and 4 weeks of implantation the four types of Thai silk fibroin based-scaffolds were classified as "non-irritant" to "slight-irritant", compared to Gelfoam(®) (control samples). These natural Thai silk fibroin-based scaffolds may provide suitable biomaterials for clinical applications.

Silk-based delivery systems of bioactive molecules

PubMed Central

Numata, Keiji; Kaplan, David L

2010-01-01

Silks are biodegradable, biocompatible, self-assemblying proteins that can also be tailored via genetic engineering to contain specific chemical features, offering utility for drug and gene delivery. Silkworm silk has been used in biomedical sutures for decades and has recently achieved Food and Drug Administration approval for expanded biomaterials device utility. With the diversity and control of size, structure and chemistry, modified or recombinant silk proteins can be designed and utilized in various biomedical application, such as for the delivery of bioactive molecules. This review focuses on the biosynthesis and applications of silk-based multi-block copolymer systems and related silk protein drug delivery systems. The utility of these systems for the delivery of small molecule drugs, proteins and genes are reviewed. PMID:20298729

Self-Assembly of a Modular Polypeptide Based on Blocks of Silk-Mimetic and Elastin-Mimetic Sequences

DTIC Science & Technology

2002-04-01

Silk -Mimetic and Elastin-Mimetic Sequences DISTRIBUTION: Approved for public release, distribution unlimited This paper is part of the following...724 © 2002 Materials Research Society N3.8 Self-Assembly of a Modular Polypeptide based on Blocks of Silk -Mimetic and Elastin- Mimetic Sequences...Chrystelle S. Cazalis, and Vincent P. Conticello* Department of Chemistry, Emory University, Atlanta, GA 30322 ABSTRACT Spider dragline silk fiber displays

Controlling silk fibroin particle features for drug delivery

PubMed Central

Lammel, Andreas; Hu, Xiao; Park, Sang-Hyug; Kaplan, David L.; Scheibel, Thomas

2010-01-01

Silk proteins are a promising material for drug delivery due to their aqueous processability, biocompatibility, and biodegradability. A simple aqueous preparation method for silk fibroin particles with controllable size, secondary structure and zeta potential is reported. The particles were produced by salting out a silk fibroin solution with potassium phosphate. The effect of ionic strength and pH of potassium phosphate solution on the yield and morphology of the particles was determined. Secondary structure and zeta potential of the silk particles could be controlled by pH. Particles produced by salting out with 1.25 M potassium phosphate pH 6 showed a dominating silk II (crystalline) structure whereas particles produced at pH 9 were mainly composed of silk I (less crystalline). The results show that silk I rich particles possess chemical and physical stability and secondary structure which remained unchanged during post treatments even upon exposure to 100% ethanol or methanol. A model is presented to explain the process of particle formation based on intra- and intermolecular interactions of the silk domains, influenced by pH and kosmotrope salts. The reported silk fibroin particles can be loaded with small molecule model drugs, such as alcian blue, rhodamine B, and crystal violet, by simple absorption based on electrostatic interactions. In vitro release of these compounds from the silk particles depends on charge – charge interactions between the compounds and the silk. With crystal violet we demonstrated that the release kinetics are dependent on the secondary structure of the particles. PMID:20219241

Fabrication and Biocompatibility of Electrospun Silk Biocomposites

PubMed Central

Wei, Kai; Kim, Byoung-Suhk; Kim, Ick-Soo

2011-01-01

Silk fibroin has attracted great interest in tissue engineering because of its outstanding biocompatibility, biodegradability and minimal inflammatory reaction. In this study, two kinds of biocomposites based on regenerated silk fibroin are fabricated by electrospinning and post-treatment processes, respectively. Firstly, regenerated silk fibroin/tetramethoxysilane (TMOS) hybrid nanofibers with high hydrophilicity are prepared, which is superior for fibroblast attachment. The electrospinning process causes adjacent fibers to ‘weld’ at contact points, which can be proved by scanning electron microscope (SEM). The water contact angle of silk/tetramethoxysilane (TMOS) composites shows a sharper decrease than pure regenerated silk fibroin nanofiber, which has a great effect on the early stage of cell attachment behavior. Secondly, a novel tissue engineering scaffold material based on electrospun silk fibroin/nano-hydroxyapatite (nHA) biocomposites is prepared by means of an effective calcium and phosphate (Ca–P) alternate soaking method. nHA is successfully produced on regenerated silk fibroin nanofiber within several min without any pre-treatments. The osteoblastic activities of this novel nanofibrous biocomposites are also investigated by employing osteoblastic-like MC3T3-E1 cell line. The cell functionality such as alkaline phosphatase (ALP) activity is ameliorated on mineralized silk nanofibers. All these results indicate that this silk/nHA biocomposite scaffold material may be a promising biomaterial for bone tissue engineering. PMID:24957869

Silk as an innovative biomaterial for cancer therapy.

PubMed

Jastrzebska, Katarzyna; Kucharczyk, Kamil; Florczak, Anna; Dondajewska, Ewelina; Mackiewicz, Andrzej; Dams-Kozlowska, Hanna

2015-01-01

Silk has been used for centuries in the textile industry and as surgical sutures. In addition to its unique mechanical properties, silk possesses other properties, such as biocompatibility, biodegradability and ability to self-assemble, which make it an interesting material for biomedical applications. Although silk forms only fibers in nature, synthetic techniques can be used to control the processing of silk into different morphologies, such as scaffolds, films, hydrogels, microcapsules, and micro- and nanospheres. Moreover, the biotechnological production of silk proteins broadens the potential applications of silk. Synthetic silk genes have been designed. Genetic engineering enables modification of silk properties or the construction of a hybrid silk. Bioengineered hybrid silks consist of a silk sequence that self-assembles into the desired morphological structure and the sequence of a polypeptide that confers a function to the silk biomaterial. The functional domains can comprise binding sites for receptors, enzymes, drugs, metals or sugars, among others. Here, we review the current status of potential applications of silk biomaterials in the field of oncology with a focus on the generation of implantable, injectable and targeted drug delivery systems and the three-dimensional cancer models based on silk scaffolds for cancer research. However, the systems described could be applied in many biomedical fields.

Recombinant DNA production of spider silk proteins.

PubMed

Tokareva, Olena; Michalczechen-Lacerda, Valquíria A; Rech, Elíbio L; Kaplan, David L

2013-11-01

Spider dragline silk is considered to be the toughest biopolymer on Earth due to an extraordinary combination of strength and elasticity. Moreover, silks are biocompatible and biodegradable protein-based materials. Recent advances in genetic engineering make it possible to produce recombinant silks in heterologous hosts, opening up opportunities for large-scale production of recombinant silks for various biomedical and material science applications. We review the current strategies to produce recombinant spider silks. © 2013 The Authors. Microbial Biotechnology published by John Wiley & Sons Ltd and Society for Applied Microbiology.

Silk-based delivery systems of bioactive molecules.

PubMed

Numata, Keiji; Kaplan, David L

2010-12-30

Silks are biodegradable, biocompatible, self-assembling proteins that can also be tailored via genetic engineering to contain specific chemical features, offering utility for drug and gene delivery. Silkworm silk has been used in biomedical sutures for decades and has recently achieved Food and Drug Administration approval for expanded biomaterials device utility. With the diversity and control of size, structure and chemistry, modified or recombinant silk proteins can be designed and utilized in various biomedical application, such as for the delivery of bioactive molecules. This review focuses on the biosynthesis and applications of silk-based multi-block copolymer systems and related silk protein drug delivery systems. The utility of these systems for the delivery of small molecule drugs, proteins and genes is reviewed. Copyright © 2010 Elsevier B.V. All rights reserved.

Production of novel biopolymers in plants: recent technological advances and future prospects.

PubMed

Snell, Kristi D; Singh, Vijay; Brumbley, Stevens M

2015-04-01

The production of novel biopolymers in plants has the potential to provide renewable sources of industrial materials through agriculture. In this review we will highlight recent progress with plant-based production of polyhydroxyalkanoates (PHAs), silk, elastin, collagen, and cyanophycin with an emphasis on the synthesis of poly[(R)-3-hydroxybutyrate] (PHB), a renewable biodegradable PHA polymer with potential commercial applications in plastics, chemicals, and feed markets. Improved production of PHB has required manipulation of promoters driving expression of transgenes, reduction in activity of endogenous enzymes in competing metabolic pathways, insertion of genes to increase carbon flow to polymer, and basic plant biochemistry to understand metabolic limitations. These experiments have increased our understanding of carbon availability and partitioning in different plant organelles, cell types, and organs, information that is useful for the production of other novel molecules in plants. Copyright © 2014 Elsevier Ltd. All rights reserved.

Regulation of Silk Genes by Hox and Homeodomain Proteins in the Terminal Differentiated Silk Gland of the Silkworm Bombyx mori

PubMed Central

Takiya, Shigeharu; Tsubota, Takuya; Kimoto, Mai

2016-01-01

The silk gland of the silkworm Bombyx mori is a long tubular organ that is divided into several subparts along its anteroposterior (AP) axis. As a trait of terminal differentiation of the silk gland, several silk protein genes are expressed with unique regional specificities. Most of the Hox and some of the homeobox genes are also expressed in the differentiated silk gland with regional specificities. The expression patterns of Hox genes in the silk gland roughly correspond to those in embryogenesis showing “colinearity”. The central Hox class protein Antennapedia (Antp) directly regulates the expression of several middle silk gland–specific silk genes, whereas the Lin-1/Isl-1/Mec3 (LIM)-homeodomain transcriptional factor Arrowhead (Awh) regulates the expression of posterior silk gland–specific genes for silk fiber proteins. We summarize our results and discuss the usefulness of the silk gland of Bombyx mori for analyzing the function of Hox genes. Further analyses of the regulatory mechanisms underlying the region-specific expression of silk genes will provide novel insights into the molecular bases for target-gene selection and regulation by Hox and homeodomain proteins. PMID:29615585

Preparation and mechanical properties of layers made of recombinant spider silk proteins and silk from silk worm

NASA Astrophysics Data System (ADS)

Junghans, F.; Morawietz, M.; Conrad, U.; Scheibel, T.; Heilmann, A.; Spohn, U.

2006-02-01

Layers of recombinant spider silks and native silks from silk worms were prepared by spin-coating and casting of various solutions. FT-IR spectra were recorded to investigate the influence of the different mechanical stress occurring during the preparation of the silk layers. The solubility of the recombinant spider silk proteins SO1-ELP, C16, AQ24NR3, and of the silk fibroin from Bombyx mori were investigated in hexafluorisopropanol, ionic liquids and concentrated salt solutions. The morphology and thickness of the layers were determined by Atomic Force Microscopy (AFM) or with a profilometer. The mechanical behaviour was investigated by acoustic impedance analysis by using a quartz crystal microbalance (QCMB) as well as by microindentation. The density of silk layers (d<300 nm) was determined based on AFM and QCMB measurements. At silk layers thicker than 300 nm significant changes of the half-band-half width can be correlated with increasing energy dissipation. Microhardness measurements demonstrate that recombinant spider silk and sericine-free Bombyx mori silk layers achieve higher elastic penetration modules EEP and Martens hardness values HM than those of polyethylenterephthalate (PET) and polyetherimide (PEI) foils.

Silk Electrogel Based Gastroretentive Drug Delivery System

NASA Astrophysics Data System (ADS)

Wang, Qianrui

Gastric cancer has become a global pandemic and there is imperative to develop efficient therapies. Oral dosing strategy is the preferred route to deliver drugs for treating the disease. Recent studies suggested silk electro hydrogel, which is pH sensitive and reversible, has potential as a vehicle to deliver the drug in the stomach environment. The aim of this study is to establish in vitro electrogelation e-gel based silk gel as a gastroretentive drug delivery system. We successfully extended the duration of silk e-gel in artificial gastric juice by mixing silk solution with glycerol at different ratios before the electrogelation. Structural analysis indicated the extended duration was due to the change of beta sheet content. The glycerol mixed silk e-gel had good doxorubicin loading capability and could release doxorubicin in a sustained-release profile. Doxorubicin loaded silk e-gels were applied to human gastric cancer cells. Significant cell viability decrease was observed. We believe that with further characterization as well as functional analysis, the silk e-gel system has the potential to become an effective vehicle for gastric drug delivery applications.

Fire Safety Aspects of Polymeric Materials. Volume 1. Materials. State of the Art

DTIC Science & Technology

1977-01-01

polymers (e.g. silk, collagen) has not been investigated ( Crawshaw et a!., 1973). 4.3 Synthetic Fibers The production of man-made fibers and natural...Thornton, A. M., Modern Plastics 43(2), 154 (1964). Coskren, R. J., SAMPE Quart., 4, No. 4, July (1973). Crawshaw , G. H., Delman, A. D., and Mehta, P

Surface modification of silk fibroin fabric using layer-by-layer polyelectrolyte deposition and heparin immobilization for small-diameter vascular prostheses.

PubMed

Elahi, M Fazley; Guan, Guoping; Wang, Lu; Zhao, Xinzhe; Wang, Fujun; King, Martin W

2015-03-03

There is an urgent need to develop a biologically active implantable small-diameter vascular prosthesis with long-term patency. Silk-fibroin-based small-diameter vascular prosthesis is a promising candidate having higher patency rate; however, the surface modification is indeed required to improve its further hemocompatibility. In this study, silk fibroin fabric was modified by a two-stage process. First, the surface of silk fibroin fabric was coated using a layer-by-layer polyelectrolyte deposition technique by stepwise dipping the silk fibroin fabric into a solution of cationic poly(allylamine hydrochloride) (PAH) and anionic poly(acrylic acid) (PAA) solution. The dipping procedure was repeated to obtain the PAH/PAA multilayers deposited on the silk fibroin fabrics. Second, the polyelectrolyte-deposited silk fibroin fabrics were treated in EDC/NHS-activated low-molecular-weight heparin (LMWH) solution at 4 °C for 24 h, resulting in immobilization of LMWH on the silk fibroin fabrics surface. Scanning electron microscopy, atomic force microscopy, and energy-dispersive X-ray data revealed the accomplishment of LMWH immobilization on the polyelectrolyte-deposited silk fibroin fabric surface. The higher the number of PAH/PAA coating layers on the silk fibroin fabric, the more surface hydrophilicity could be obtained, resulting in a higher fetal bovine serum protein and platelets adhesion resistance properties when tested in vitro. In addition, compared with untreated sample, the surface-modified silk fibroin fabrics showed negligible loss of bursting strength and thus reveal the acceptability of polyelectrolytes deposition and heparin immobilization approach for silk-fibroin-based small-diameter vascular prostheses modification.

pH-Dependent anticancer drug release from silk nanoparticles

PubMed Central

Seib, F. Philipp; Jones, Gregory T.; Rnjak-Kovacina, Jelena; Lin, Yinan; Kaplan, David L.

2013-01-01

Silk has traditionally been used as a suture material because of its excellent mechanical properties and biocompatibility. These properties have led to the development of different silk-based material formats for tissue engineering and regenerative medicine. Although there have been a small number of studies about the use of silk particles for drug delivery, none of these studies have assessed the potential of silk to act as a stimulus-responsive anticancer nanomedicine. This report demonstrates that an acetone precipitation of silk allowed the formation of uniform silk nanoparticles (98 nm diameter, polydispersity index 0.109), with an overall negative surface charge (-33.6 ±5.8 mV), in a single step. Silk nanoparticles were readily loaded with doxorubicin (40 ng doxorubicin/μg silk) and showed pH-dependent release (pH 4.5>> 6.0 > 7.4). In vitro studies with human breast cancer cell lines demonstrated that the silk nanoparticles were not cytotoxic (IC50 >120/μ/ml) and that doxorubicin-loaded silk nanoparticles were able to overcome drug resistance mechanisms. Live cell fluorescence microscopy studies showed endocytic uptake and lysosomal accumulation of silk nanoparticles. In summary, the pH-dependent drug release and lysosomal accumulation of silk nanoparticles demonstrated the ability of drug-loaded silk nanoparticles to serve as a lysosomotropic anticancer nanomedicine. PMID:23625825

Self-assembly of silk fibroin under osmotic stress

NASA Astrophysics Data System (ADS)

Sohn, Sungkyun

The supramolecular self-assembly behavior of silk fibroin was investigated using osmotic stress technique. In Chapter 2, a ternary phase diagram of water-silk-LiBr was constructed based on X-ray results on the osmotically stressed regenerated silk fibroin of Bombyx mori silkworm. Microscopic data indicated that silk I is a hydrated structure and a rough estimate of the number of water molecules lost by the structure upon converting from silk I to silk II has been made, and found to be about 2.2 per [GAGAGS] hexapeptide. In Chapter 3, wet-spinning of osmotically stressed, regenerated silk fibroin was performed, based on the prediction that the enhanced control over structure and phase behavior using osmotic stress method helps improve the physical properties of wet-spun regenerated silk fibroin fibers. The osmotic stress was applied in order to pre-structure the regenerated silk fibroin molecule from its original random coil state to more oriented state, manipulating the phase of the silk solution in the phase diagram before the start of spinning. Monofilament fiber with a diameter of 20 microm was produced. In Chapter 4, we investigated if there is a noticeable synergistic osmotic pressure increase between co-existing polymeric osmolyte and salt when extremely highly concentrated salt molecules are present both at sample subphase and stressing subphase, as is the case of silk fibroin self-assembly. The equilibration method that measures osmotic pressure relative to a reference with known osmotic pressure was introduced. Osmotic pressure of aqueous LiBr solution up to 2.75M was measured and it was found that the synergistic effect was insignificant up to this salt concentration. Solution parameters of stressing solutions and Arrhenius kinetics based on time-temperature relationship for the equilibration process were derived as well. In Chapter 5, self-assembly behavior of natural silk fibroin within the gland of Bombyx mori silkworm was investigated using osmotic stress technique. Microscopic and thermodynamic details of this self-assembly process along the spinline have been assessed. Formation of a needle-shaped molecular lath under appropriate osmotic stress was found. Silk I degree of hydration of silk gland was quantitatively estimated by image analysis of optical micrographs and the numbers varied from 2.2 to 2.7 depending on the region in the gland. Osmotic pressure in the gland was also estimated by equilibration method.

Functionalized bioengineered spider silk spheres improve nuclease resistance and activity of oligonucleotide therapeutics providing a strategy for cancer treatment.

PubMed

Kozlowska, Anna Karolina; Florczak, Anna; Smialek, Maciej; Dondajewska, Ewelina; Mackiewicz, Andrzej; Kortylewski, Marcin; Dams-Kozlowska, Hanna

2017-09-01

Cell-selective delivery and sensitivity to serum nucleases remain major hurdles to the clinical application of RNA-based oligonucleotide therapeutics, such as siRNA. Spider silk shows great potential as a biomaterial due to its biocompatibility and biodegradability. Self-assembling properties of silk proteins allow for processing into several different morphologies such as fibers, scaffolds, films, hydrogels, capsules and spheres. Moreover, bioengineering of spider silk protein sequences can functionalize silk by adding peptide moieties with specific features including binding or cell recognition domains. We demonstrated that modification of silk protein by adding the nucleic acid binding domain enabled the development of a novel oligonucleotide delivery system that can be utilized to improve pharmacokinetics of RNA-based therapeutics, such as CpG-siRNA. The MS2 bioengineered silk was functionalized with poly-lysine domain (KN) to generate hybrid silk MS2KN. CpG-siRNA efficiently bound to MS2KN in contrary to control MS2. Both MS2KN complexes and spheres protected CpG-siRNA from degradation by serum nucleases. CpG-siRNA molecules encapsulated into MS2KN spheres were efficiently internalized and processed by TLR9-positive macrophages. Importantly, CpG-STAT3siRNA loaded in silk spheres showed delayed and extended target gene silencing compared to naked oligonucleotides. The prolonged Stat3 silencing resulted in the more pronounced downregulation of interleukin 6 (IL-6), a proinflammatory cytokine and upstream activator of STAT3, which limits the efficacy of TLR9 immunostimulation. Our results demonstrate the feasibility of using spider silk spheres as a carrier of therapeutic nucleic acids. Moreover, the modified kinetic and activity of the CpG-STAT3siRNA embedded into silk spheres is likely to improve immunotherapeutic effects in vivo. We demonstrated that modification of silk protein by adding the nucleic acid binding domain enabled the development of a novel oligonucleotide delivery system that can be utilized to improve pharmacokinetics of RNA-based therapeutics. Although, the siRNA constructs have already given very promising results in the cancer therapy, the in vivo application of RNA-based oligonucleotide therapeutics still is limited due to their sensitivity to serum nucleases and some toxicity. We propose a carrier for RNA-based therapeutics that is made of bioengineered spider silk. We showed that functionalized bioengineered spider silk spheres not only protected RNA-based therapeutics from degradation by serum nucleases, but what is more important the embedding of siRNA into silk spheres delayed and extended target gene silencing compared with naked oligonucleotides. Moreover, we showed that plain silk spheres did not have unspecific effect on target gene levels proving not only to be non-cytotoxic but also very neutral vehicles in terms of TLR9/STAT3 activation in macrophages. We demonstrated advantages of novel delivery technology in safety and efficacy comparing with delivery of naked CpG-STAT3siRNA therapeutics. Copyright © 2017 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

Uncovering the structure-function relationship in spider silk

NASA Astrophysics Data System (ADS)

Yarger, Jeffery L.; Cherry, Brian R.; van der Vaart, Arjan

2018-03-01

All spiders produce protein-based biopolymer fibres that we call silk. The most studied of these silks is spider dragline silk, which is very tough and relatively abundant compared with other types of spider silks. Considerable research has been devoted to understanding the relationship between the molecular structure and mechanical properties of spider dragline silks. In this Review, we overview experimental and computational studies that have provided a wealth of detail at the molecular level on the highly conserved repetitive core and terminal regions of spider dragline silk. We also discuss the role of the nanocrystalline β-sheets and amorphous regions in determining the properties of spider silk fibres, endowing them with strength and elasticity. Additionally, we outline imaging techniques and modelling studies that elucidate the importance of the hierarchical structure of silk fibres at the molecular level. These insights into structure-function relationships can guide the reverse engineering of spider silk to enable the production of superior synthetic fibres.

Effect of Processing on Silk-Based Biomaterials: Reproducibility and Biocompatibility

PubMed Central

Wray, Lindsay S.; Hu, Xiao; Gallego, Jabier; Georgakoudi, Irene; Omenetto, Fiorenzo G.; Schmidt, Daniel; Kaplan, David L.

2012-01-01

Silk fibroin has been successfully used as a biomaterial for tissue regeneration. In order to prepare silk fibroin biomaterials for human implantation a series of processing steps are required to purify the protein. Degumming to remove inflammatory sericin is a crucial step related to biocompatibility and variability in the material. Detailed characterization of silk fibroin degumming is reported. The degumming conditions significantly affected cell viability on the silk fibroin material and the ability to form three-dimensional porous scaffolds from the silk fibroin, but did not affect macrophage activation or β-sheet content in the materials formed. Methods are also provided to determine the content of residual sericin in silk fibroin solutions and to assess changes in silk fibroin molecular weight. Amino acid composition analysis was used to detect sericin residuals in silk solutions with a detection limit between 1.0% and 10% wt/wt, while fluorescence spectroscopy was used to reproducibly distinguish between silk samples with different molecular weights. Both methods are simple and require minimal sample volume, providing useful quality control tools for silk fibroin preparation processes. PMID:21695778

Spider Silk Fibers Spun from Soluble Recombinant Silk Produced in Mammalian Cells

NASA Astrophysics Data System (ADS)

Lazaris, Anthoula; Arcidiacono, Steven; Huang, Yue; Zhou, Jiang-Feng; Duguay, François; Chretien, Nathalie; Welsh, Elizabeth A.; Soares, Jason W.; Karatzas, Costas N.

2002-01-01

Spider silks are protein-based ``biopolymer'' filaments or threads secreted by specialized epithelial cells as concentrated soluble precursors of highly repetitive primary sequences. Spider dragline silk is a flexible, lightweight fiber of extraordinary strength and toughness comparable to that of synthetic high-performance fibers. We sought to ``biomimic'' the process of spider silk production by expressing in mammalian cells the dragline silk genes (ADF-3/MaSpII and MaSpI) of two spider species. We produced soluble recombinant (rc)-dragline silk proteins with molecular masses of 60 to 140 kilodaltons. We demonstrated the wet spinning of silk monofilaments spun from a concentrated aqueous solution of soluble rc-spider silk protein (ADF-3; 60 kilodaltons) under modest shear and coagulation conditions. The spun fibers were water insoluble with a fine diameter (10 to 40 micrometers) and exhibited toughness and modulus values comparable to those of native dragline silks but with lower tenacity. Dope solutions with rc-silk protein concentrations >20% and postspinning draw were necessary to achieve improved mechanical properties of the spun fibers. Fiber properties correlated with finer fiber diameter and increased birefringence.

Fabrication and characterization of biomaterial film from gland silk of muga and eri silkworms.

PubMed

Dutta, Saranga; Talukdar, Bijit; Bharali, Rupjyoti; Rajkhowa, Rangam; Devi, Dipali

2013-05-01

This study discusses the possibilities of liquid silk (Silk gland silk) of Muga and Eri silk, the indigenous non mulberry silkworms of North Eastern region of India, as potential biomaterials. Silk protein fibroin of Bombyx mori, commonly known as mulberry silkworm, has been extensively studied as a versatile biomaterial. As properties of different silk-based biomaterials vary significantly, it is important to characterize the non mulberry silkworms also in this aspect. Fibroin was extracted from the posterior silk gland of full grown fifth instars larvae, and 2D film was fabricated using standard methods. The films were characterized using SEM, Dynamic contact angle test, FTIR, XRD, DSC, and TGA and compared with respective silk fibers. SEM images of films reveal presence of some globules and filamentous structure. Films of both the silkworms were found to be amorphous with random coil conformation, hydrophobic in nature, and resistant to organic solvents. Non mulberry silk films had higher thermal resistance than mulberry silk. Fibers were thermally more stable than the films. This study provides insight into the new arena of research in application of liquid silk of non mulberry silkworms as biomaterials. Copyright © 2012 Wiley Periodicals, Inc.

[Engineered spider silk: the intelligent biomaterial of the future. Part I].

PubMed

Florczak, Anna; Piekoś, Konrad; Kaźmierska, Katarzyna; Mackiewicz, Andrzej; Dams-Kozłowska, Hanna

2011-06-17

The unique properties of spider silk such as strength, extensibility, toughness, biocompatibility and biodegradability are the reasons for the recent development in silk biomaterial technology. For a long time scientific progress was impeded by limited access to spider silk. However, the development of the molecular biology strategy was a breaking point in synthetic spider silk protein design. The sequences of engineered spider silk are based on the consensus motives of the corresponding natural equivalents. Moreover, the engineered silk proteins may be modified in order to gain a new function. The strategy of the hybrid proteins constructed on the DNA level combines the sequence of engineered silk, which is responsible for the biomaterial structure, with the sequence of polypeptide which allows functionalization of the silk biomaterial. The functional domains may comprise receptor binding sites, enzymes, metal or sugar binding sites and others. Currently, advanced research is being conducted, which on the one hand focuses on establishing the particular silk structure and understanding the process of silk thread formation in nature. On the other hand, there are attempts to improve methods of engineered spider silk protein production. Due to acquired knowledge and recent progress in synthetic protein technology, the engineered silk will turn into intelligent biomaterial of the future, while its industrial production scale will trigger a biotechnological revolution.

The advances and perspectives of recombinant protein production in the silk gland of silkworm Bombyx mori.

PubMed

Xu, Hanfu

2014-10-01

The silk gland of silkworm Bombyx mori, is one of the most important organs that has been fully studied and utilized so far. It contributes finest silk fibers to humankind. The silk gland has excellent ability of synthesizing silk proteins and is a kind tool to produce some useful recombinant proteins, which can be widely used in the biological, biotechnical and pharmaceutical application fields. It's a very active area to express recombinant proteins using the silk gland as a bioreactor, and great progress has been achieved recently. This review recapitulates the progress of producing recombinant proteins and silk-based biomaterials in the silk gland of silkworm in addition to the construction of expression systems. Current challenges and future trends in the production of valuable recombinant proteins using transgenic silkworms are also discussed.

Regenerated silk materials for functionalized silk orthopedic devices by mimicking natural processing

PubMed Central

Li, Chunmei; Hotz, Blake; Ling, Shengjie; Guo, Jin; Haas, Dylan S.; Marelli, Benedetto; Omenetto, Fiorenzo; Lin, Samuel J.; Kaplan, David L.

2016-01-01

Silk fibers spun by silkworms and spiders exhibit exceptional mechanical properties with a unique combination of strength, extensibility and toughness. In contrast, the mechanical properties of regenerated silk materials can be tuned through control of the fabrication process. Here we introduce a biomimetic, all-aqueous process, to obtain bulk regenerated silk-based materials for the fabrication of functionalized orthopedic devices. The silk materials generated in the process replicate the nano-scale structure of natural silk fibers and possess excellent mechanical properties. The biomimetic materials demonstrated excellent machinability, providing a path towards the fabrication of a new family of resorbable orthopedic devices where organic solvents are avoided, thus allowing functionalization with bioactive molecules to promote bone remodeling and integration. PMID:27697669

Characterizing the Effects of Washing by Different Detergents on the Wavelength-Scale Microstructures of Silk Samples Using Mueller Matrix Polarimetry.

PubMed

Dong, Yang; He, Honghui; He, Chao; Zhou, Jialing; Zeng, Nan; Ma, Hui

2016-08-10

Silk fibers suffer from microstructural changes due to various external environmental conditions including daily washings. In this paper, we take the backscattering Mueller matrix images of silk samples for non-destructive and real-time quantitative characterization of the wavelength-scale microstructure and examination of the effects of washing by different detergents. The 2D images of the 16 Mueller matrix elements are reduced to the frequency distribution histograms (FDHs) whose central moments reveal the dominant structural features of the silk fibers. A group of new parameters are also proposed to characterize the wavelength-scale microstructural changes of the silk samples during the washing processes. Monte Carlo (MC) simulations are carried out to better understand how the Mueller matrix parameters are related to the wavelength-scale microstructure of silk fibers. The good agreement between experiments and simulations indicates that the Mueller matrix polarimetry and FDH based parameters can be used to quantitatively detect the wavelength-scale microstructural features of silk fibers. Mueller matrix polarimetry may be used as a powerful tool for non-destructive and in situ characterization of the wavelength-scale microstructures of silk based materials.

Characterizing the Effects of Washing by Different Detergents on the Wavelength-Scale Microstructures of Silk Samples Using Mueller Matrix Polarimetry

PubMed Central

Dong, Yang; He, Honghui; He, Chao; Zhou, Jialing; Zeng, Nan; Ma, Hui

2016-01-01

Silk fibers suffer from microstructural changes due to various external environmental conditions including daily washings. In this paper, we take the backscattering Mueller matrix images of silk samples for non-destructive and real-time quantitative characterization of the wavelength-scale microstructure and examination of the effects of washing by different detergents. The 2D images of the 16 Mueller matrix elements are reduced to the frequency distribution histograms (FDHs) whose central moments reveal the dominant structural features of the silk fibers. A group of new parameters are also proposed to characterize the wavelength-scale microstructural changes of the silk samples during the washing processes. Monte Carlo (MC) simulations are carried out to better understand how the Mueller matrix parameters are related to the wavelength-scale microstructure of silk fibers. The good agreement between experiments and simulations indicates that the Mueller matrix polarimetry and FDH based parameters can be used to quantitatively detect the wavelength-scale microstructural features of silk fibers. Mueller matrix polarimetry may be used as a powerful tool for non-destructive and in situ characterization of the wavelength-scale microstructures of silk based materials. PMID:27517919

Flexible, highly sensitive pressure sensor with a wide range based on graphene-silk network structure

NASA Astrophysics Data System (ADS)

Liu, Ying; Tao, Lu-Qi; Wang, Dan-Yang; Zhang, Tian-Yu; Yang, Yi; Ren, Tian-Ling

2017-03-01

In this paper, a flexible, simple-preparation, and low-cost graphene-silk pressure sensor based on soft silk substrate through thermal reduction was demonstrated. Taking silk as the support body, the device had formed a three-dimensional structure with ordered multi-layer structure. Through a simple and low-cost process technology, graphene-silk pressure sensor can achieve the sensitivity value of 0.4 kPa - 1 , and the measurement range can be as high as 140 kPa. Besides, pressure sensor can have a good combination with knitted clothing and textile product. The signal had good reproducibility in response to different pressures. Furthermore, graphene-silk pressure sensor can not only detect pressure higher than 100 kPa, but also can measure weak body signals. The characteristics of high-sensitivity, good repeatability, flexibility, and comfort for skin provide the high possibility to fit on various wearable electronics.

Discerning Silk Produced by Bombyx mori from Those Produced by Wild Species Using an Enzyme-Linked Immunosorbent Assay Combined with Conventional Methods.

PubMed

You, Qiushi; Li, Qingqing; Zheng, Hailing; Hu, Zhiwen; Zhou, Yang; Wang, Bing

2017-09-06

Recently, much interest has been paid to the separation of silk produced by Bombyx mori from silk produced by other species and tracing the beginnings of silk cultivation from wild silk exploitation. In this paper, significant differences between silks from Bombyx mori and other species were found by microscopy and spectroscopy, such as morphology, secondary structure, and amino acid composition. For further accurate identification, a diagnostic antibody was designed by comparing the peptide sequences of silks produced by Bombyx mori and other species. The results of the noncompetitive indirect enzyme-linked immunosorbent assay (ELISA) indicated that the antibody that showed good sensitivity and high specificity can definitely discern silk produced by Bombyx mori from silk produced by wild species. Thus, the antibody-based immunoassay has the potential to be a powerful tool for tracing the beginnings of silk cultivation. In addition, combining the sensitive, specific, and convenient ELISA technology with other conventional methods can provide more in-depth and accurate information for species identification.

Development of an enzyme-linked-immunosorbent-assay technique for accurate identification of poorly preserved silks unearthed in ancient tombs.

PubMed

Zheng, Qin; Wu, Xiaofeng; Zheng, Hailing; Zhou, Yang

2015-05-01

We report the preparation of a specific fibroin antibody and its use for the identification of unearthed ancient silk relics. Based on the 12-amino-acid repeat sequence "GAGAGSGAGAGS", which is found in fibroin of the silkworm Bombyx mori, a specific antibody against fibroin was prepared in rabbits through peptide synthesis and carrier-protein coupling. This antibody was highly specific for fibroin found in silk. Using this antibody we have successfully identified four silk samples from different time periods. Our results reveal, for the first time, a method capable of detecting silk from a few milligrams of archaeological fabric that has been buried for thousands of years, confirming that the ancient practice of wearing silk products while praying for rebirth dated back to at least 400 BCE. This method also complements current approaches in silk detection, especially for the characterization of poorly preserved silks, promoting the investigation of silk origins and of ancient clothing cultures.

Evaluating drug trafficking on the Tor Network: Silk Road 2, the sequel.

PubMed

Dolliver, Diana S

2015-11-01

Housing an illicit, online drug retail market generating sales in the millions of USD, the Silk Road was a profitable marketplace with a growing and loyal consumer base. Following its FBI-forced shut down in October 2013, the Silk Road enjoyed newfound fame that contributed to an increase in new users downloading and accessing the Tor Network; however, with this particular marketplace out of order, Silk Road 2 was launched to fill the void. The goals of this study were to (1) compare the metrics of Silk Road 2 to the original site, and to (2) determine if there were any indications of the presence of more sophisticated drug trafficking operations. Data were collected from Silk Road 2 during the months of August and September 2014 using webcrawling software. Silk Road 2 was a much smaller marketplace than the original Silk Road. Of the 1834 unique items for sale, 348 were drug items sold by 145 distinct vendors shipping from 19 countries. Of the drug items advertised, most were stimulants and hallucinogens. The United States is both the number one country of origin for drug sales on Silk Road 2 and the number one destination country. Interestingly, 73% of all vendor accounts on Silk Road 2 advertised drug items, even though drugs only constituted 19% of all items advertised. This study was the first to research Silk Road 2, the replacement illicit marketplace to the original virtual Silk Road. This study was also the first to examine indications of the presence of more coordinated drug trafficking efforts in an online setting. The findings indicated that while Silk Road 2 was not primarily a drug market, there were indications that some vendor accounts may have connections reaching beyond a base retail market. Copyright © 2015 Elsevier B.V. All rights reserved.

Improving the mechanical properties of collagen-based membranes using silk fibroin for corneal tissue engineering.

PubMed

Long, Kai; Liu, Yang; Li, Weichang; Wang, Lin; Liu, Sa; Wang, Yingjun; Wang, Zhichong; Ren, Li

2015-03-01

Although collagen with outstanding biocompatibility has promising application in corneal tissue engineering, the mechanical properties of collagen-based scaffolds, especially suture retention strength, must be further improved to satisfy the requirements of clinical applications. This article describes a toughness reinforced collagen-based membrane using silk fibroin. The collagen-silk fibroin membranes based on collagen [silk fibroin (w/w) ratios of 100:5, 100:10, and 100:20] were prepared by using silk fibroin and cross-linking by 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide. These membranes were analyzed by scanning electron microscopy and their optical property, and NaCl and tryptophan diffusivity had been tested. The water content was found to be dependent on the content of silk fibroin, and CS10 membrane (loading 10 wt % of silk fibroin) performed the optimal mechanical properties. Also the suture experiments have proved CS10 has high suture retention strength, which can be sutured in rabbit eyes integrally. Moreover, the composite membrane proved good biocompatibility for the proliferation of human corneal epithelial cells in vitro. Lamellar keratoplasty shows that CS10 membrane promoted complete epithelialization in 35 ± 5 days, and their transparency is restored quickly in the first month. Corneal rejection reaction, neovascularization, and keratoconus are not observed. The composite films show potential for use in the field of corneal tissue engineering. © 2014 Wiley Periodicals, Inc.

Curcumin-functionalized silk materials for enhancing adipogenic differentiation of bone marrow-derived human mesenchymal stem cells

PubMed Central

Li, Chunmei; Luo, Tingting; Zheng, Zhaozhu; Murphy, Amanda R.; Wang, Xiaoqin; Kaplan, David L.

2014-01-01

Curcumin, a natural phenolic compound derived from the plant Curcuma longa, was physically entrapped and stabilized in silk hydrogel films and its influence on human bone marrow-derived mesenchymal stem cells (hBMSCs) was assessed related to adipogenic differentiation. The presence of curcumin significantly reduced silk gelation time and changed the porous morphology of gel matrix, but did not change the formation of silk beta-sheet structure. Based on spectrofluorimetric analysis, curcumin likely interacted with hydrophobic residues in silk, interacting with the beta-sheet domains formed in the hydrogels. The antioxidant activity of silk film-associated curcumin remained functional over at least one month in both the dry and hydrated state. Negligible curcumin was released from silk hydrogel films over 48 hours incubation in aqueous solution. For hBMSCs cultured on silk films containing more than 0.25 mg/mL curcumin, cell proliferation was inhibited while adipogenesis was significantly promoted based on transcripts as well as oil red O staining. When hBMSCs were cultured in media containing free curcumin, both proliferation and adipogenesis of hBMSCs were inhibited when curcumin concentrations exceeded 5 μM, which is more than 1,000-times higher than the level of curcumin released from the films in aqueous solution. Thus, silk film-associated curcumin exhibited different effects on hBMSC proliferation and differentiation when compared to curcumin in solution. PMID:25132274

Thromboelastometric and platelet responses to silk biomaterials.

PubMed

Kundu, Banani; Schlimp, Christoph J; Nürnberger, Sylvia; Redl, Heinz; Kundu, S C

2014-05-13

Silkworm's silk is natural biopolymer with unique properties including mechanical robustness, all aqueous base processing and ease in fabrication into different multifunctional templates. Additionally, the nonmulberry silks have cell adhesion promoting tri-peptide (RGD) sequences, which make it an immensely potential platform for regenerative medicine. The compatibility of nonmulberry silk with human blood is still elusive; thereby, restricts its further application as implants. The present study, therefore, evaluate the haematocompatibility of silk biomaterials in terms of platelet interaction after exposure to nonmulberry silk of Antheraea mylitta using thromboelastometry (ROTEM). The mulberry silk of Bombyx mori and clinically used Uni-Graft W biomaterial serve as references. Shortened clotting time, clot formation times as well as enhanced clot strength indicate the platelet mediated activation of blood coagulation cascade by tested biomaterials; which is comparable to controls.

Regenerated silk materials for functionalized silk orthopedic devices by mimicking natural processing.

PubMed

Li, Chunmei; Hotz, Blake; Ling, Shengjie; Guo, Jin; Haas, Dylan S; Marelli, Benedetto; Omenetto, Fiorenzo; Lin, Samuel J; Kaplan, David L

2016-12-01

Silk fibers spun by silkworms and spiders exhibit exceptional mechanical properties with a unique combination of strength, extensibility and toughness. In contrast, the mechanical properties of regenerated silk materials can be tuned through control of the fabrication process. Here we introduce a biomimetic, all-aqueous process, to obtain bulk regenerated silk-based materials for the fabrication of functionalized orthopedic devices. The silk materials generated in the process replicate the nano-scale structure of natural silk fibers and possess excellent mechanical properties. The biomimetic materials demonstrate excellent machinability, providing a path towards the fabrication of a new family of resorbable orthopedic devices where organic solvents are avoided, thus allowing functionalization with bioactive molecules to promote bone remodeling and integration. Copyright © 2016 Elsevier Ltd. All rights reserved.

Bio-functionalized silk hydrogel microfluidic systems.

PubMed

Zhao, Siwei; Chen, Ying; Partlow, Benjamin P; Golding, Anne S; Tseng, Peter; Coburn, Jeannine; Applegate, Matthew B; Moreau, Jodie E; Omenetto, Fiorenzo G; Kaplan, David L

2016-07-01

Bio-functionalized microfluidic systems were developed based on a silk protein hydrogel elastomeric materials. A facile multilayer fabrication method using gelatin sacrificial molding and layer-by-layer assembly was implemented to construct interconnected, three dimensional (3D) microchannel networks in silk hydrogels at 100 μm minimum feature resolution. Mechanically activated valves were implemented to demonstrate pneumatic control of microflow. The silk hydrogel microfluidics exhibit controllable mechanical properties, long-term stability in various environmental conditions, tunable in vitro and in vivo degradability in addition to optical transparency, providing unique features for cell/tissue-related applications than conventional polydimethylsiloxane (PDMS) and existing hydrogel-based microfluidic options. As demonstrated in the work here, the all aqueous-based fabrication process at ambient conditions enabled the incorporation of active biological substances in the bulk phase of these new silk microfluidic systems during device fabrication, including enzymes and living cells, which are able to interact with the fluid flow in the microchannels. These silk hydrogel-based microfluidic systems offer new opportunities in engineering active diagnostic devices, tissues and organs that could be integrated in vivo, and for on-chip cell sensing systems. Copyright © 2016 Elsevier Ltd. All rights reserved.

Silk Fibroin as Edible Coating for Perishable Food Preservation

NASA Astrophysics Data System (ADS)

Marelli, B.; Brenckle, M. A.; Kaplan, D. L.; Omenetto, F. G.

2016-05-01

The regeneration of structural biopolymers into micelles or nanoparticles suspended in water has enabled the design of new materials with unique and compelling properties that can serve at the interface between the biotic and the abiotic worlds. In this study, we leveraged silk fibroin quintessential properties (i.e. polymorphism, conformability and hydrophobicity) to design a water-based protein suspension that self-assembles on the surface of food upon dip coating. The water-based post-processing control of the protein polymorphism enables the modulation of the diffusion of gases through the silk fibroin thin membranes (e.g. O2 and CO2 diffusion, water vapour permeability), which is a key parameter to manage food freshness. In particular, an increased beta-sheet content corresponds to a reduction in oxygen diffusion through silk fibroin thin films. By using the dip coating of strawberries and bananas as proof of principle, we have shown that the formation of micrometre-thin silk fibroin membranes around the fruits helps the management of postharvest physiology of the fruits. Thus, silk fibroin coatings enhance fruits’ shelf life at room conditions by reducing cell respiration rate and water evaporation. The water-based processing and edible nature of silk fibroin makes this approach a promising alternative for food preservation with a naturally derived material.

Silk Fibroin as Edible Coating for Perishable Food Preservation

PubMed Central

Marelli, B.; Brenckle, M. A.; Kaplan, D. L.; Omenetto, F. G.

2016-01-01

The regeneration of structural biopolymers into micelles or nanoparticles suspended in water has enabled the design of new materials with unique and compelling properties that can serve at the interface between the biotic and the abiotic worlds. In this study, we leveraged silk fibroin quintessential properties (i.e. polymorphism, conformability and hydrophobicity) to design a water-based protein suspension that self-assembles on the surface of food upon dip coating. The water-based post-processing control of the protein polymorphism enables the modulation of the diffusion of gases through the silk fibroin thin membranes (e.g. O2 and CO2 diffusion, water vapour permeability), which is a key parameter to manage food freshness. In particular, an increased beta-sheet content corresponds to a reduction in oxygen diffusion through silk fibroin thin films. By using the dip coating of strawberries and bananas as proof of principle, we have shown that the formation of micrometre-thin silk fibroin membranes around the fruits helps the management of postharvest physiology of the fruits. Thus, silk fibroin coatings enhance fruits’ shelf life at room conditions by reducing cell respiration rate and water evaporation. The water-based processing and edible nature of silk fibroin makes this approach a promising alternative for food preservation with a naturally derived material. PMID:27151492

Genetically engineered silk-collagen-like copolymer for biomedical applications: production, characterization and evaluation of cellular response.

PubMed

Włodarczyk-Biegun, Małgorzata K; Werten, Marc W T; de Wolf, Frits A; van den Beucken, Jeroen J J P; Leeuwenburgh, Sander C G; Kamperman, Marleen; Cohen Stuart, Martien A

2014-08-01

Genetically engineered protein polymers (GEPP) are a class of multifunctional materials with precisely controlled molecular structure and property profile. Representing a promising alternative for currently used materials in biomedical applications, GEPP offer multiple benefits over natural and chemically synthesized polymers. However, producing them in sufficient quantities for preclinical research remains challenging. Here, we present results from an in vitro cellular response study of a recombinant protein polymer that is soluble at low pH but self-organizes into supramolecular fibers and physical hydrogels at neutral pH. It has a triblock structure denoted as C2S(H)48C2, which consists of hydrophilic collagen-inspired and histidine-rich silk-inspired blocks. The protein was successfully produced by the yeast Pichia pastoris in laboratory-scale bioreactors, and it was purified by selective precipitation. This efficient and inexpensive production method provided material of sufficient quantities, purity and sterility for cell culture study. Rheology and erosion studies showed that it forms hydrogels exhibiting long-term stability, self-healing behavior and tunable mechanical properties. Primary rat bone marrow cells cultured in direct contact with these hydrogels remained fully viable; however, proliferation and mineralization were relatively low compared to collagen hydrogel controls, probably because of the absence of cell-adhesive motifs. As biofunctional factors can be readily incorporated to improve material performance, our approach provides a promising route towards biomedical applications. Copyright © 2014 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

New Materials Based on Spider Silk

DTIC Science & Technology

2007-11-06

encoding the honey bee secretion signal was (genetically) fused to the spider silk protein. A respective recombinant virus was used to infect high...2007). Role of individual spider silk domains during assembly. Poster at the conference on "Protein Assembly in Materials, Biology and Medicine

Microdissection of black widow spider silk-producing glands.

PubMed

Jeffery, Felicia; La Mattina, Coby; Tuton-Blasingame, Tiffany; Hsia, Yang; Gnesa, Eric; Zhao, Liang; Franz, Andreas; Vierra, Craig

2011-01-11

Modern spiders spin high-performance silk fibers with a broad range of biological functions, including locomotion, prey capture and protection of developing offspring. Spiders accomplish these tasks by spinning several distinct fiber types that have diverse mechanical properties. Such specialization of fiber types has occurred through the evolution of different silk-producing glands, which function as small biofactories. These biofactories manufacture and store large quantities of silk proteins for fiber production. Through a complex series of biochemical events, these silk proteins are converted from a liquid into a solid material upon extrusion. Mechanical studies have demonstrated that spider silks are stronger than high-tensile steel. Analyses to understand the relationship between the structure and function of spider silk threads have revealed that spider silk consists largely of proteins, or fibroins, that have block repeats within their protein sequences. Common molecular signatures that contribute to the incredible tensile strength and extensibility of spider silks are being unraveled through the analyses of translated silk cDNAs. Given the extraordinary material properties of spider silks, research labs across the globe are racing to understand and mimic the spinning process to produce synthetic silk fibers for commercial, military and industrial applications. One of the main challenges to spinning artificial spider silk in the research lab involves a complete understanding of the biochemical processes that occur during extrusion of the fibers from the silk-producing glands. Here we present a method for the isolation of the seven different silk-producing glands from the cobweaving black widow spider, which includes the major and minor ampullate glands [manufactures dragline and scaffolding silk], tubuliform [synthesizes egg case silk], flagelliform [unknown function in cob-weavers], aggregate [makes glue silk], aciniform [synthesizes prey wrapping and egg case threads] and pyriform [produces attachment disc silk]. This approach is based upon anesthetizing the spider with carbon dioxide gas, subsequent separation of the cephalothorax from the abdomen, and microdissection of the abdomen to obtain the silk-producing glands. Following the separation of the different silk-producing glands, these tissues can be used to retrieve different macromolecules for distinct biochemical analyses, including quantitative real-time PCR, northern- and western blotting, mass spectrometry (MS or MS/MS) analyses to identify new silk protein sequences, search for proteins that participate in the silk assembly pathway, or use the intact tissue for cell culture or histological experiments.

Silk Fibroin Based Porous Materials

PubMed Central

Zhang, Qiang; Yan, Shuqin; Li, Mingzhong

2009-01-01

Silk from the Bombyx mori silkworm is a protein-based fiber. Bombyx mori silk fibroin (SF) is one of the most important candidates for biomedical porous material based on its superior machinability, biocompatibility, biodegradation, bioresorbability, and so on. In this paper, we have reviewed the key features of SF. Moreover we have focused on the morphous, technical processing, and biocompatibility of SF porous materials, followed by the application research. Finally, we provide a perspective the potential and problems of SF porous materials.

Clay-Enriched Silk Biomaterials for Bone Formation

PubMed Central

Mieszawska, Aneta J.; Llamas, Jabier Gallego; Vaiana, Christopher A.; Kadakia, Madhavi P.; Naik, Rajesh R.; Kaplan, David L.

2011-01-01

The formation of silk protein/clay composite biomaterials for bone tissue formation is described. Silk fibroin serves as an organic scaffolding material offering mechanical stability suitable for bone specific uses. Clay montmorillonite (Cloisite ® Na+) and sodium silicate are sources of osteoinductive silica-rich inorganic species, analogous to bioactive bioglass-like bone repair biomaterial systems. Different clay particle-silk composite biomaterial films were compared to silk films doped with sodium silicate as controls for support of human bone marrow derived mesenchymal stem cells (hMSCs) in osteogenic culture. The cells adhered and proliferated on the silk/clay composites over two weeks. Quantitative real-time RT-PCR analysis revealed increased transcript levels for alkaline phosphatase (ALP), bone sialoprotein (BSP), and collagen type 1 (Col I) osteogenic markers in the cells cultured on the silk/clay films in comparison to the controls. Early evidence for bone formation based on collagen deposition at the cell-biomaterial interface was also found, with more collagen observed for the silk films with higher contents of clay particles. The data suggest that the silk/clay composite systems may be useful for further study toward bone regenerative needs. PMID:21549864

Inhibitory effect of corn silk on skin pigmentation.

PubMed

Choi, Sang Yoon; Lee, Yeonmi; Kim, Sung Soo; Ju, Hyun Min; Baek, Ji Hwoon; Park, Chul-Soo; Lee, Dong-Hyuk

2014-03-03

In this study, the inhibitory effect of corn silk on melanin production was evaluated. This study was performed to investigate the inhibitory effect of corn silk on melanin production in Melan-A cells by measuring melanin production and protein expression. The corn silk extract applied on Melan-A cells at a concentration of 100 ppm decreased melanin production by 37.2% without cytotoxicity. This was a better result than arbutin, a positive whitening agent, which exhibited a 26.8% melanin production inhibitory effect at the same concentration. The corn silk extract did not suppress tyrosinase activity but greatly reduced the expression of tyrosinase in Melan-A cells. In addition, corn silk extract was applied to the human face with hyperpigmentation, and skin color was measured to examine the degree of skin pigment reduction. The application of corn silk extract on faces with hyperpigmentation significantly reduced skin pigmentation without abnormal reactions. Based on the results above, corn silk has good prospects for use as a material for suppressing skin pigmentation.

Silk-microfluidics for advanced biotechnological applications: A progressive review.

PubMed

Konwarh, Rocktotpal; Gupta, Prerak; Mandal, Biman B

2016-01-01

Silk based biomaterials have not only carved a unique niche in the domain of regenerative medicine but new avenues are also being explored for lab-on-a-chip applications. It is pertinent to note that biospinning of silk represents nature's signature microfluidic-maneuver. Elucidation of non-Newtonian flow of silk in the glands of spiders and silkworms has inspired researchers to fabricate devices for continuous extrusion and concentration of silk. Microfluidic channel networks within porous silk scaffolds ensure optimal nutrient and oxygen supply apart from serving as precursors for vascularization in tissue engineering applications. On the other hand, unique topographical features and surface wettability of natural silk fibers have inspired development of a number of simple and cost-effective devices for applications like blood typing and chemical sensing. This review mirrors the recent progress and challenges in the domain of silk-microfluidics for prospective avant-garde applications in the realm of biotechnology. Copyright © 2016 Elsevier Inc. All rights reserved.

Regeneration of high-quality silk fibroin fiber by wet spinning from CaCl2-formic acid solvent.

PubMed

Zhang, Feng; Lu, Qiang; Yue, Xiaoxiao; Zuo, Baoqi; Qin, Mingde; Li, Fang; Kaplan, David L; Zhang, Xueguang

2015-01-01

Silks spun by silkworms and spiders feature outstanding mechanical properties despite being spun under benign conditions. The superior physical properties of silk are closely related to its complicated hierarchical structures constructed from nanoscale building blocks, such as nanocrystals and nanofibrils. Here, we report a novel silk dissolution behavior, which preserved nanofibrils in CaCl2-formic acid solution, that enables spinning of high-quality fibers with a hierarchical structure. This process is characterized by simplicity, high efficiency, low cost, environmental compatibility and large-scale industrialization potential, as well as having utility and potential for the recycling of silk waste and the production of silk-based functional materials. Copyright © 2014. Published by Elsevier Ltd.

Human stem cell neuronal differentiation on silk-carbon nanotube composite

NASA Astrophysics Data System (ADS)

Chen, Chi-Shuo; Soni, Sushant; Le, Catherine; Biasca, Matthew; Farr, Erik; Chen, Eric Y.-T.; Chin, Wei-Chun

2012-02-01

Human embryonic stem cells [hESCs] are able to differentiate into specific lineages corresponding to regulated spatial and temporal signals. This unique attribute holds great promise for regenerative medicine and cell-based therapy for many human diseases such as spinal cord injury [SCI] and multiple sclerosis [MS]. Carbon nanotubes [CNTs] have been successfully used to promote neuronal differentiation, and silk has been widely applied in tissue engineering. This study aims to build silk-CNT composite scaffolds for improved neuron differentiation efficiency from hESCs. Two neuronal markers (β-III tubulin and nestin) were utilized to determine the hESC neuronal lineage differentiation. In addition, axonal lengths were measured to evaluate the progress of neuronal development. The results demonstrated that cells on silk-CNT scaffolds have a higher β-III tubulin and nestin expression, suggesting augmented neuronal differentiation. In addition, longer axons with higher density were found to associate with silk-CNT scaffolds. Our silk-CNT-based composite scaffolds can promote neuronal differentiation of hESCs. The silk-CNT composite scaffolds developed here can serve as efficient supporting matrices for stem cell-derived neuronal transplants, offering a promising opportunity for nerve repair treatments for SCI and MS patients.

Beating the Heat: Fast Scanning Melts Beta Sheet Crystals

NASA Astrophysics Data System (ADS)

Cebe, Peggy; Hu, Xiao; Kaplan, David; Zhuravlev, Evgeny; Wurm, Andreas; Arbeiter, Daniella; Schick, Christoph

2014-03-01

Beta-pleated-sheet crystals are among the most stable of protein secondary structures, and are responsible for the remarkable physical properties of many fibrous proteins, such as silk. Previous thinking was that beta-pleated-sheet crystals in the dry solid state would not melt upon input of heat energy alone. Indeed, at conventional heating rates (~1-50 °C/min), silk exhibits its glass transition (~175 °C), followed by cold crystallization, and then by immediate thermal degradation beginning at about 225 °C. Here we demonstrate that beta-pleated-sheet crystals can melt directly from the solid state to become random coils, helices, and turns. We use fast scanning chip calorimetry at 2,000 K/s to avoid thermal degradation, and report the first reversible thermal melting of protein beta-pleated-sheet crystals, exemplified by silk fibroin. The similarity between thermal melting behavior of lamellar crystals of synthetic polymers and beta-pleated-sheet crystals is confirmed. The authors acknowledge support from the National Science Foundation and German Academic Exchange Service DAAD; EZ acknowledges a European Union funded Marie Curie EST fellowship (ADVATEC); XH and DK acknowledge NIH P41 Tissue Engineering Resource Center.

Facile preparation of paclitaxel loaded silk fibroin nanoparticles for enhanced antitumor efficacy by locoregional drug delivery.

PubMed

Wu, Puyuan; Liu, Qin; Li, Rutian; Wang, Jing; Zhen, Xu; Yue, Guofeng; Wang, Huiyu; Cui, Fangbo; Wu, Fenglei; Yang, Mi; Qian, Xiaoping; Yu, Lixia; Jiang, Xiqun; Liu, Baorui

2013-12-11

Non-toxic, safe materials and preparation methods are among the most important factors when designing nanoparticles (NPs) for future clinical application. Here we report a novel and facile method encapsulating anticancer drug paclitaxel (PTX) into silk fibroin (SF), a biocompatible and biodegradable natural polymer, without adding any toxic organic solvents, surfactants or other toxic agents. The paclitaxel loaded silk fibroin nanoparticles (PTX-SF-NPs) with a diameter of 130 nm were formed in an aqueous solution at room temperature by self-assembling of SF protein, which demonstrated mainly silk I conformation in the NPs. In cellular uptake experiments, coumarin-6 loaded SF NPs were taken up efficiently by two human gastric cancer cell lines BGC-823 and SGC-7901. In vitro cytotoxicity studies demonstrated that PTX kept its pharmacological activity when incorporating into PTX-SF-NPs, while SF showed no cytotoxicity to cells. The in vivo antitumor effects of PTX-SF-NPs were evaluated on gastric cancer nude mice exnograft model. We found that locoregional delivery of PTX-SF-NPs demonstrated superior antitumor efficacy by delaying tumor growth and reducing tumor weights compared with systemic administration. Furthermore, the organs of mice in NP treated groups didn't show obvious toxicity, indicating the in vivo safety of SF NPs. These results suggest that SF NPs are promising drug delivery carriers, and locoregional delivery of SF NPs could be a potential future clinical cancer treatment regimen.

Hierarchical charge distribution controls self-assembly process of silk in vitro

NASA Astrophysics Data System (ADS)

Zhang, Yi; Zhang, Cencen; Liu, Lijie; Kaplan, David L.; Zhu, Hesun; Lu, Qiang

2015-12-01

Silk materials with different nanostructures have been developed without the understanding of the inherent transformation mechanism. Here we attempt to reveal the conversion road of the various nanostructures and determine the critical regulating factors. The regulating conversion processes influenced by a hierarchical charge distribution were investigated, showing different transformations between molecules, nanoparticles and nanofibers. Various repulsion and compressive forces existed among silk fibroin molecules and aggregates due to the exterior and interior distribution of charge, which further controlled their aggregating and deaggregating behaviors and finally formed nanofibers with different sizes. Synergistic action derived from molecular mobility and concentrations could also tune the assembly process and final nanostructures. It is suggested that the complicated silk fibroin assembly processes comply a same rule based on charge distribution, offering a promising way to develop silk-based materials with designed nanostructures.

Silk micrococoons for protein stabilisation and molecular encapsulation

NASA Astrophysics Data System (ADS)

Shimanovich, Ulyana; Ruggeri, Francesco S.; de Genst, Erwin; Adamcik, Jozef; Barros, Teresa P.; Porter, David; Müller, Thomas; Mezzenga, Raffaele; Dobson, Christopher M.; Vollrath, Fritz; Holland, Chris; Knowles, Tuomas P. J.

2017-07-01

Naturally spun silks generate fibres with unique properties, including strength, elasticity and biocompatibility. Here we describe a microfluidics-based strategy to spin liquid native silk, obtained directly from the silk gland of Bombyx mori silkworms, into micron-scale capsules with controllable geometry and variable levels of intermolecular β-sheet content in their protein shells. We demonstrate that such micrococoons can store internally the otherwise highly unstable liquid native silk for several months and without apparent effect on its functionality. We further demonstrate that these native silk micrococoons enable the effective encapsulation, storage and release of other aggregation-prone proteins, such as functional antibodies. These results show that native silk micrococoons are capable of preserving the full activity of sensitive cargo proteins that can aggregate and lose function under conditions of bulk storage, and thus represent an attractive class of materials for the storage and release of active biomolecules.

Tunable green graphene-silk biomaterials: Mechanism of protein-based nanocomposites.

PubMed

Wang, Fang; Jyothirmayee Aravind, S S; Wu, Hao; Forys, Joseph; Venkataraman, Venkat; Ramanujachary, Kandalam; Hu, Xiao

2017-10-01

Green graphene materials prepared by photoreduction of graphite oxide were first time blended with aqueous-based silk fibroin proteins to improve the mechanical and thermal properties of silk biomaterials, and their nanocomposite interaction mechanism was illustrated. Powder X-ray diffraction (XRD) analysis confirmed the complete exfoliation of graphite oxide to graphene in presence of focused pulses of solar radiation. By varying the concentration of graphene (0.1wt% to 10wt%), a series of free standing graphene-silk films were prepared and were systematically characterized by Fourier transform infrared spectroscopy (FTIR), scanning electron microscopy (SEM) and nanoindentation measurements. The homogeneity of graphene in silk as well as the thermal stability of the composite films was demonstrated by thermal gravimetric analysis (TGA) and temperature-modulated differential scanning calorimetry (TMDSC). Surprisingly, silk composite film containing only 0.5wt% of graphene gives the highest Young's modulus of 1.65GPa (about 5.8 times higher than the pure silk's modulus), indicating a nano-composite to micro-composite transition of silk-graphene structure occurred around this mixing ratio. This finding provided an easy approach to improve the elastic modulus and other physical properties of silk materials by adding a tiny amount of graphene sheets. Fibroblast cells studies also proved that these graphene-silk materials can significantly improve cell adhesion, growth and proliferation. This protein nanocomposite study provided a useful model to understand how to manipulate the hydrophobic-hydrophobic and polar-polar interactions between high-surface-area inorganic nanomaterials and amphiphilic protein materials, which has many emerging applications in the material science and engineering, such as bio-device fabrication, drug storage and release, and tissue regeneration. Copyright © 2017 Elsevier B.V. All rights reserved.

In planta production of ELPylated spidroin-based proteins results in non-cytotoxic biopolymers.

PubMed

Hauptmann, Valeska; Menzel, Matthias; Weichert, Nicola; Reimers, Kerstin; Spohn, Uwe; Conrad, Udo

2015-02-19

Spider silk is a tear-resistant and elastic biopolymer that has outstanding mechanical properties. Additionally, exiguous immunogenicity is anticipated for spider silks. Therefore, spider silk represents a potential ideal biomaterial for medical applications. All known spider silk proteins, so-called spidroins, reveal a composite nature of silk-specific units, allowing the recombinant production of individual and combined segments. In this report, a miniaturized spidroin gene, named VSO1 that contains repetitive motifs of MaSp1 has been synthesized and combined to form multimers of distinct lengths, which were heterologously expressed as elastin-like peptide (ELP) fusion proteins in tobacco. The elastic penetration moduli of layered proteins were analyzed for different spidroin-based biopolymers. Moreover, we present the first immunological analysis of synthetic spidroin-based biopolymers. Characterization of the binding behavior of the sera after immunization by competitive ELISA suggested that the humoral immune response is mainly directed against the fusion partner ELP. In addition, cytocompatibility studies with murine embryonic fibroblasts indicated that recombinant spidroin-based biopolymers, in solution or as coated proteins, are well tolerated. The results show that spidroin-based biopolymers can induce humoral immune responses that are dependent on the fusion partner and the overall protein structure. Furthermore, cytocompatibility assays gave no indication of spidroin-derived cytotoxicity, suggesting that recombinant produced biopolymers composed of spider silk-like repetitive elements are suitable for biomedical applications.

Microdissection of Black Widow Spider Silk-producing Glands

PubMed Central

Hsia, Yang; Gnesa, Eric; Zhao, Liang; Franz, Andreas; Vierra, Craig

2011-01-01

Modern spiders spin high-performance silk fibers with a broad range of biological functions, including locomotion, prey capture and protection of developing offspring 1,2. Spiders accomplish these tasks by spinning several distinct fiber types that have diverse mechanical properties. Such specialization of fiber types has occurred through the evolution of different silk-producing glands, which function as small biofactories. These biofactories manufacture and store large quantities of silk proteins for fiber production. Through a complex series of biochemical events, these silk proteins are converted from a liquid into a solid material upon extrusion. Mechanical studies have demonstrated that spider silks are stronger than high-tensile steel 3. Analyses to understand the relationship between the structure and function of spider silk threads have revealed that spider silk consists largely of proteins, or fibroins, that have block repeats within their protein sequences 4. Common molecular signatures that contribute to the incredible tensile strength and extensibility of spider silks are being unraveled through the analyses of translated silk cDNAs. Given the extraordinary material properties of spider silks, research labs across the globe are racing to understand and mimic the spinning process to produce synthetic silk fibers for commercial, military and industrial applications. One of the main challenges to spinning artificial spider silk in the research lab involves a complete understanding of the biochemical processes that occur during extrusion of the fibers from the silk-producing glands. Here we present a method for the isolation of the seven different silk-producing glands from the cobweaving black widow spider, which includes the major and minor ampullate glands [manufactures dragline and scaffolding silk] 5,6, tubuliform [synthesizes egg case silk] 7,8, flagelliform [unknown function in cob-weavers], aggregate [makes glue silk], aciniform [synthesizes prey wrapping and egg case threads] 9 and pyriform [produces attachment disc silk] 10. This approach is based upon anesthetizing the spider with carbon dioxide gas, subsequent separation of the cephalothorax from the abdomen, and microdissection of the abdomen to obtain the silk-producing glands. Following the separation of the different silk-producing glands, these tissues can be used to retrieve different macromolecules for distinct biochemical analyses, including quantitative real-time PCR, northern- and western blotting, mass spectrometry (MS or MS/MS) analyses to identify new silk protein sequences, search for proteins that participate in the silk assembly pathway, or use the intact tissue for cell culture or histological experiments. PMID:21248709

Multiscale mechanisms of nutritionally induced property variation in spider silks.

PubMed

Blamires, Sean J; Nobbs, Madeleine; Martens, Penny J; Tso, I-Min; Chuang, Wei-Tsung; Chang, Chung-Kai; Sheu, Hwo-Shuenn

2018-01-01

Variability in spider major ampullate (MA) silk properties at different scales has proven difficult to determine and remains an obstacle to the development of synthetic fibers mimicking MA silk performance. A multitude of techniques may be used to measure multiscale aspects of silk properties. Here we fed five species of Araneoid spider solutions that either contained protein or were protein deprived and performed silk tensile tests, small and wide-angle X-ray scattering (SAXS/WAXS), amino acid composition analyses, and silk gene expression analyses, to resolve persistent questions about how nutrient deprivation induces variations in MA silk mechanical properties across scales. Our analyses found that the properties of each spider's silk varied differently in response to variations in their protein intake. We found changes in the crystalline and non-crystalline nanostructures to play specific roles in inducing the property variations we found. Across treatment MaSp expression patterns differed in each of the five species. We found that in most species MaSp expression and amino acid composition variations did not conform with our predictions based on a traditional MaSp expression model. In general, changes to the silk's alanine and proline compositions influenced the alignment of the proteins within the silk's amorphous region, which influenced silk extensibility and toughness. Variations in structural alignment in the crystalline and non-crystalline regions influenced ultimate strength independent of genetic expression. Our study provides the deepest insights thus far into the mechanisms of how MA silk properties vary from gene expression to nanostructure formations to fiber mechanics. Such knowledge is imperative for promoting the production of synthetic silk fibers.

Production And Characterization Of Synthetic Spider Silks Based On Nephila Clavipes Major Ampullate Silk Proteins

NASA Astrophysics Data System (ADS)

An, Bo

The extraordinary mechanical properties of orb-weaving spider silks have served spiders for over 400 million years. However, only in the late 20th century did we start to understand the molecular nature of spider silk that contributes to its incredible properties as biomaterials. Among all seven types of spider silks, major ampullate silk from typical orb-weaving spiders is the toughest of all, it consists of primarily two proteins: MaSp1 and MaSp2. Variable ratios and conserved motifs of these two proteins in all the native spider silks demonstrate the significant role of MaSp1 and MaSp2 in controlling the mechanical properties of the fiber. The amino acid sequences of the orb weaving spider silk proteins have remained almost unchanged for more than 100 million years. Interestingly, MaSp1 and MaSp2 are the only two components in all studied dragline silk fibers from these spiders. The mechanical properties of native dragline silk vary slightly between species, which are believed to relate to the ratio of MaSp1 to MaSp2 in the silk. Both of these facts clearly indicate the importance of these two proteins to the mechanical properties of the fiber. Various types of synthetic spider silk fibers have been produced and studied in an effort to mass-produce man-made fibers with qualities comparable to native spider silk. To investigate the roles of MaSp1 and MaSp2 in silk fiber, synthetic MaSp1 (major abundant protein in Nephila clavipes major ampullate silks) only fibers, MaSp1/MaSp2 protein mixture fibers and chimeric protein fibers with both MaSp1 and MaSp2 sequence features have been produced and tested for mechanical properties. Solid-State Nuclear Magnetic Resonance was used to characterize the structure of silk fibers and reveal the relation between fiber spatial structure and mechanical properties.

Impact of processing parameters on the haemocompatibility of Bombyx mori silk films.

PubMed

Seib, F Philipp; Maitz, Manfred F; Hu, Xiao; Werner, Carsten; Kaplan, David L

2012-02-01

Silk has traditionally been used for surgical sutures due to its lasting strength and durability; however, the use of purified silk proteins as a scaffold material for vascular tissue engineering goes beyond traditional use and requires application-orientated biocompatibility testing. For this study, a library of Bombyx mori silk films was generated and exposed to various solvents and treatment conditions to reflect current silk processing techniques. The films, along with clinically relevant reference materials, were exposed to human whole blood to determine silk blood compatibility. All substrates showed an initial inflammatory response comparable to polylactide-co-glycolide (PLGA), and a low to moderate haemostasis response similar to polytetrafluoroethylene (PTFE) substrates. In particular, samples that were water annealed at 25 °C for 6 h demonstrated the best blood compatibility based on haemostasis parameters (e.g. platelet decay, thrombin-antithrombin complex, platelet factor 4, granulocytes-platelet conjugates) and inflammatory parameters (e.g. C3b, C5a, CD11b, surface-associated leukocytes). Multiple factors such as treatment temperature and solvent influenced the biological response, though no single physical parameter such as β-sheet content, isoelectric point or contact angle accurately predicted blood compatibility. These findings, when combined with prior in vivo data on silk, support a viable future for silk-based vascular grafts. Copyright © 2011 Elsevier Ltd. All rights reserved.

The influence of specific binding of collagen-silk chimeras to silk biomaterials on hMSC behavior

PubMed Central

An, Bo; DesRochers, Teresa M.; Qin, Guokui; Xia, Xiaoxia; Thiagarajan, Geetha; Brodsky, Barbara; Kaplan, David

2012-01-01

Collagen-like proteins in the bacteria Streptococcus pyogenes adopt a triple-helix structure with a thermal stability similar to that of animal collagens, can be expressed in high yield in E. coli and can be easily modified through molecular biology techniques. However, potential applications for such recombinant collagens are limited by their lack of higher order structure to achieve the physical properties needed for most biomaterials. To overcome this problem, the S. pyrogenes collagen domain was fused to a repetitive Bombyx mori silk consensus sequence, as a strategy to direct specific non-covalent binding onto solid silk materials whose superior stability, mechanical and material properties have been previously established. This approach resulted in the successful binding of these new collagen-silk chimeric proteins to silk films and porous scaffolds, and the binding affinity could be controlled by varying the number of repeats in the silk sequence. To explore the potential of collagen-silk chimera for regulating biological activity, integrin (Int) and fibronectin (Fn) binding sequences from mammalian collagens were introduced into the bacterial collagen domain. The attachment of bioactive collagen-silk chimeras to solid silk biomaterials promoted hMSC spreading and proliferation substantially in comparison to the controls. The ability to combine the biomaterial features of silk with the biological activities of collagen allowed more rapid cell interactions with silk-based biomaterials, improved regulation of stem cell growth and differentiation, as well as the formation of artificial extracellular matrices useful for tissue engineering applications. PMID:23088839

Nanostructure and molecular mechanics of spider dragline silk protein assemblies

PubMed Central

Keten, Sinan; Buehler, Markus J.

2010-01-01

Spider silk is a self-assembling biopolymer that outperforms most known materials in terms of its mechanical performance, despite its underlying weak chemical bonding based on H-bonds. While experimental studies have shown that the molecular structure of silk proteins has a direct influence on the stiffness, toughness and failure strength of silk, no molecular-level analysis of the nanostructure and associated mechanical properties of silk assemblies have been reported. Here, we report atomic-level structures of MaSp1 and MaSp2 proteins from the Nephila clavipes spider dragline silk sequence, obtained using replica exchange molecular dynamics, and subject these structures to mechanical loading for a detailed nanomechanical analysis. The structural analysis reveals that poly-alanine regions in silk predominantly form distinct and orderly beta-sheet crystal domains, while disorderly regions are formed by glycine-rich repeats that consist of 31-helix type structures and beta-turns. Our structural predictions are validated against experimental data based on dihedral angle pair calculations presented in Ramachandran plots, alpha-carbon atomic distances, as well as secondary structure content. Mechanical shearing simulations on selected structures illustrate that the nanoscale behaviour of silk protein assemblies is controlled by the distinctly different secondary structure content and hydrogen bonding in the crystalline and semi-amorphous regions. Both structural and mechanical characterization results show excellent agreement with available experimental evidence. Our findings set the stage for extensive atomistic investigations of silk, which may contribute towards an improved understanding of the source of the strength and toughness of this biological superfibre. PMID:20519206

Nanostructure and molecular mechanics of spider dragline silk protein assemblies.

PubMed

Keten, Sinan; Buehler, Markus J

2010-12-06

Spider silk is a self-assembling biopolymer that outperforms most known materials in terms of its mechanical performance, despite its underlying weak chemical bonding based on H-bonds. While experimental studies have shown that the molecular structure of silk proteins has a direct influence on the stiffness, toughness and failure strength of silk, no molecular-level analysis of the nanostructure and associated mechanical properties of silk assemblies have been reported. Here, we report atomic-level structures of MaSp1 and MaSp2 proteins from the Nephila clavipes spider dragline silk sequence, obtained using replica exchange molecular dynamics, and subject these structures to mechanical loading for a detailed nanomechanical analysis. The structural analysis reveals that poly-alanine regions in silk predominantly form distinct and orderly beta-sheet crystal domains, while disorderly regions are formed by glycine-rich repeats that consist of 3₁-helix type structures and beta-turns. Our structural predictions are validated against experimental data based on dihedral angle pair calculations presented in Ramachandran plots, alpha-carbon atomic distances, as well as secondary structure content. Mechanical shearing simulations on selected structures illustrate that the nanoscale behaviour of silk protein assemblies is controlled by the distinctly different secondary structure content and hydrogen bonding in the crystalline and semi-amorphous regions. Both structural and mechanical characterization results show excellent agreement with available experimental evidence. Our findings set the stage for extensive atomistic investigations of silk, which may contribute towards an improved understanding of the source of the strength and toughness of this biological superfibre.

Silk-based biomaterials functionalized with fibronectin type II promotes cell adhesion.

PubMed

Pereira, Ana Margarida; Machado, Raul; da Costa, André; Ribeiro, Artur; Collins, Tony; Gomes, Andreia C; Leonor, Isabel B; Kaplan, David L; Reis, Rui L; Casal, Margarida

2017-01-01

The objective of this work was to exploit the fibronectin type II (FNII) module from human matrix metalloproteinase-2 as a functional domain for the development of silk-based biopolymer blends that display enhanced cell adhesion properties. The DNA sequence of spider dragline silk protein (6mer) was genetically fused with the FNII coding sequence and expressed in Escherichia coli. The chimeric protein 6mer+FNII was purified by non-chromatographic methods. Films prepared from 6mer+FNII by solvent casting promoted only limited cell adhesion of human skin fibroblasts. However, the performance of the material in terms of cell adhesion was significantly improved when 6mer+FNII was combined with a silk-elastin-like protein in a concentration-dependent behavior. With this work we describe a novel class of biopolymer that promote cell adhesion and potentially useful as biomaterials for tissue engineering and regenerative medicine. This work reports the development of biocompatible silk-based composites with enhanced cell adhesion properties suitable for biomedical applications in regenerative medicine. The biocomposites were produced by combining a genetically engineered silk-elastin-like protein with a genetically engineered spider-silk-based polypeptide carrying the three domains of the fibronectin type II module from human metalloproteinase-2. These composites were processed into free-standing films by solvent casting and characterized for their biological behavior. To our knowledge this is the first report of the exploitation of all three FNII domains as a functional domain for the development of bioinspired materials with improved biological performance. The present study highlights the potential of using genetically engineered protein-based composites as a platform for the development of new bioinspired biomaterials. Copyright © 2016 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

Sequence of Spider Aciniform and Piriform Silks

DTIC Science & Technology

2001-09-19

7/98nd subtan-6/01 4. TITLE AND SUBTITLE Sequence of Spider Aciniform and Piriform Silks 5. FUNDING NUMBERS DAAD19-01-1-0569 6...aciniform glands from Argiope trifasciata were used to construct a cDNA library. The library was probed with various DNA probes based on known spider silk ...sequence in a number of other spider silks . The 5’end of the clone still appears to be repetitive sequence and thus it is unlikely to be a full-length

Scale-up of water-based spider silk film casting using a film applicator.

PubMed

Agostini, Elisa; Winter, Gerhard; Engert, Julia

2017-10-30

Spider silk proteins for applications in drug delivery have attracted an increased interest during the past years. Some possible future medical applications for this biocompatible and biodegradable material are scaffolds for tissue engineering, implantable drug delivery systems and coatings for implants. Recently, we reported on the preparation of water-based spider silk films for drug delivery applications. In the current study, we describe the development of a manufacturing technique for casting larger spider silk films from aqueous solution employing a film applicator. Films were characterized in terms of morphology, water solubility, protein secondary structure, thermal stability, and mechanical properties. Different post-treatments were evaluated (phosphate ions, ethanol, steam sterilization and water vapor) to increase the content of β-sheets thereby achieving water insolubility of the films. Finally, the mechanical properties of the spider silk films were improved by incorporating 2-pyrrolidone as plasticizer. Copyright © 2017 Elsevier B.V. All rights reserved.

Acid extraction and purification of recombinant spider silk proteins.

PubMed

Mello, Charlene M; Soares, Jason W; Arcidiacono, Steven; Butler, Michelle M

2004-01-01

A procedure has been developed for the isolation of recombinant spider silk proteins based upon their unique stability and solubilization characteristics. Three recombinant silk proteins, (SpI)7, NcDS, and [(SpI)4/(SpII)1]4, were purified by extraction with organic acids followed by affinity or ion exchange chromatography resulting in 90-95% pure silk solutions. The protein yield of NcDS (15 mg/L culture) and (SpI)7 (35 mg/L) increased 4- and 5-fold, respectively, from previously reported values presumably due to a more complete solubilization of the expressed recombinant protein. [(SpI)4/(SpII)1]4, a hybrid protein based on the repeat sequences of spidroin I and spidroin II, had a yield of 12.4 mg/L. This method is an effective, reproducible technique that has broad applicability for a variety of silk proteins as well as other acid stable biopolymers.

Antibiotic free selection for the high level biosynthesis of a silk-elastin-like protein

PubMed Central

Barroca, Mário; Rodrigues, Paulo; Sobral, Rómulo; Costa, M. Manuela R.; Chaves, Susana R.; Machado, Raul; Casal, Margarida; Collins, Tony

2016-01-01

Silk-elastin-like proteins (SELPs) are a family of genetically engineered recombinant protein polymers exhibiting mechanical and biological properties suited for a wide range of applications in the biomedicine and materials fields. They are being explored as the next generation of biomaterials but low productivities and use of antibiotics during production undermine their economic viability and safety. We have developed an industrially relevant, scalable, fed-batch process for the high level production of a novel SELP in E. coli in which the commonly used antibiotic selection marker of the expression vector is exchanged for a post segregational suicide system, the separate-component-stabilisation system (SCS). SCS significantly augments SELP productivity but also enhances the product safety profile and reduces process costs by eliminating the use of antibiotics. Plasmid content increased following induction but no significant differences in plasmid levels were discerned when using SCS or the antibiotic selection markers under the controlled fed-batch conditions employed. It is suggested that the absence of competing plasmid-free cells improves host cell viability and enables increased productivity with SCS. With the process developed, 12.8 g L−1 purified SELP was obtained, this is the highest SELP productivity reported to date and clearly demonstrates the commercial viability of these promising polymers. PMID:27982135

Dual-functional composite with anticoagulant and antibacterial properties based on heparinized silk fibroin and chitosan.

PubMed

Wang, Jianglin; Hu, Wei; Liu, Qun; Zhang, Shengmin

2011-07-01

Heparinized biomaterials exhibit great anticoagulant properties. However, they promote proliferation of Staphylococcus aureus (S. aureus) and therefore cause infection within the bloodstream upon implantation in vivo. In the present study, an interesting dual-functional composite with anticoagulant and antibacterial properties based on heparinized silk fibroin and chitosan was synthesized. First, heparin was grafted onto the silk fibroin by covalent immobilization with N-(3-dimethylaminopropyl)-N'-ethylcarbodiimide (EDC) and N-hydroxysuccinimide (NHS). All data gathered from Fourier transform infrared spectroscopy (FTIR), scanning electron microscopy (SEM) and elemental analysis (EA) indicated that the heparin was successfully immobilized onto the silk fibroin. The dual-functional composite of heparinized silk fibroin and chitosan was then fabricated by a blending method. The anticoagulant activity of the heparinized materials was evaluated using the prothrombin time (PT), activated partial thromboplastin time (APTT) and thrombin time (TT). The results showed that both heparinized silk fibroin and the composite material exhibited better hemocompatibility in comparison with single silk fibroin or chitosan. The antibacterial property of the materials was investigated by the pour-plate method. Results further suggested that the composite antibacterial property with respect to S. aureus was significantly enhanced. The dual-functionality of the composite material may supply a potential choice in blood contact devices. Copyright © 2011 Elsevier B.V. All rights reserved.

The influence of specific binding of collagen-silk chimeras to silk biomaterials on hMSC behavior.

PubMed

An, Bo; DesRochers, Teresa M; Qin, Guokui; Xia, Xiaoxia; Thiagarajan, Geetha; Brodsky, Barbara; Kaplan, David L

2013-01-01

Collagen-like proteins in the bacteria Streptococcus pyogenes adopt a triple-helix structure with a thermal stability similar to that of animal collagens, can be expressed in high yield in Escherichia coli and can be easily modified through molecular biology techniques. However, potential applications for such recombinant collagens are limited by their lack of higher order structure to achieve the physical properties needed for most biomaterials. To overcome this problem, the S. pyogenes collagen domain was fused to a repetitive Bombyx mori silk consensus sequence, as a strategy to direct specific non-covalent binding onto solid silk materials whose superior stability, mechanical and material properties have been previously established. This approach resulted in the successful binding of these new collagen-silk chimeric proteins to silk films and porous scaffolds, and the binding affinity could be controlled by varying the number of repeats in the silk sequence. To explore the potential of collagen-silk chimera for regulating biological activity, integrin (Int) and fibronectin (Fn) binding sequences from mammalian collagens were introduced into the bacterial collagen domain. The attachment of bioactive collagen-silk chimeras to solid silk biomaterials promoted hMSC spreading and proliferation substantially in comparison to the controls. The ability to combine the biomaterial features of silk with the biological activities of collagen allowed more rapid cell interactions with silk-based biomaterials, improved regulation of stem cell growth and differentiation, as well as the formation of artificial extracellular matrices useful for tissue engineering applications. Copyright © 2012 Elsevier Ltd. All rights reserved.

Transcript Profile Analyses of Maize Silks Reveal Effective Activation of Genes Involved in Microtubule-Based Movement, Ubiquitin-Dependent Protein Degradation, and Transport in the Pollination Process

PubMed Central

Chen, Hao; Sun, Wei; Zhang, Xian Sheng

2013-01-01

Pollination is the first crucial step of sexual reproduction in flowering plants, and it requires communication and coordination between the pollen and the stigma. Maize (Zea mays) is a model monocot with extraordinarily long silks, and a fully sequenced genome, but little is known about the mechanism of its pollen–stigma interactions. In this study, the dynamic gene expression of silks at four different stages before and after pollination was analyzed. The expression profiles of immature silks (IMS), mature silks (MS), and silks at 20 minutes and 3 hours after pollination (20MAP and 3HAP, respectively) were compared. In total, we identified 6,337 differentially expressed genes in silks (SDEG) at the four stages. Among them, the expression of 172 genes were induced upon pollination, most of which participated in RNA binding, processing and transcription, signal transduction, and lipid metabolism processes. Genes in the SDEG dataset could be divided into 12 time-course clusters according to their expression patterns. Gene Ontology (GO) enrichment analysis revealed that many genes involved in microtubule-based movement, ubiquitin-mediated protein degradation, and transport were predominantly expressed at specific stages, indicating that they might play important roles in the pollination process of maize. These results add to current knowledge about the pollination process of grasses and provide a foundation for future studies on key genes involved in the pollen–silk interaction in maize. PMID:23301084

Heterogeneous nucleation of hydroxyapatite on protein: structural effect of silk sericin

PubMed Central

Takeuchi, Akari; Ohtsuki, Chikara; Miyazaki, Toshiki; Kamitakahara, Masanobu; Ogata, Shin-ichi; Yamazaki, Masao; Furutani, Yoshiaki; Kinoshita, Hisao; Tanihara, Masao

2005-01-01

Acidic proteins play an important role during mineral formation in biological systems, but the mechanism of mineral formation is far from understood. In this paper, we report on the relationship between the structure of a protein and hydroxyapatite deposition under biomimetic conditions. Sericin, a type of silk protein, was adopted as a suitable protein for studying structural effect on hydroxyapatite deposition, since it forms a hydroxyapatite layer on its surface in a metastable calcium phosphate solution, and its structure has been reported. Sericin effectively induced hydroxyapatite nucleation when it has high molecular weight and a β sheet structure. This indicates that the specific structure of a protein can effectively induce heterogeneous nucleation of hydroxyapatite in a biomimetic solution, i.e. a metastable calcium phosphate solution. This finding is useful in understanding biomineralization, as well as for the design of organic polymers that can effectively induce hydroxyapatite nucleation. PMID:16849195

Electrodeposited silk coatings for bone implants.

PubMed

Elia, Roberto; Michelson, Courtney D; Perera, Austin L; Brunner, Teresa F; Harsono, Masly; Leisk, Gray G; Kugel, Gerard; Kaplan, David L

2015-11-01

The aim of this study was to characterize the mechanical properties and drug elution features of silk protein-based electrodeposited dental implant coatings. Silk processing conditions were modified to obtain coatings with a range of mechanical properties on titanium studs. These coatings were assessed for adhesive strength and dissolution, with properties tuned using water vapor annealing or glycerol incorporation to modulate crystalline content. Coating reproducibility was demonstrated over a range of silk concentrations from 1% to 10%. Surface roughness of titanium substrates was altered using industry relevant acid etching and grit blasting, and the effect of surface topography on silk coating adhesion was assessed. Florescent compounds were incorporated into the silk coatings, which were modulated for crystalline content, to achieve four days of sustained release of the compounds. This silk electrogelation technique offers a safe and relatively simple approach to generate mechanically robust, biocompatible, and degradable implant coatings that can also be functionalized with bioactive compounds to modulate the local regenerative tissue environment. © 2014 Wiley Periodicals, Inc.

Electrodeposited silk coatings for bone implants

PubMed Central

Elia, Roberto; Michelson, Courtney D.; Perera, Austin L.; Brunner, Teresa F.; Harsono, Masly; Leisk, Gray G.; Kugel, Gerard; Kaplan, David L.

2014-01-01

The aim of this study was to characterize the mechanical properties and drug elution features of silk protein-based electrodeposited dental implant coatings. Silk processing conditions were modified to obtain coatings with a range of mechanical properties on titanium studs. These coatings were assessed for adhesive strength and dissolution, with properties tuned using water vapor annealing or glycerol incorporation to modulate crystalline content. Coating reproducibility was demonstrated over a range of silk concentrations from 1 to 10%. Surface roughness of titanium substrates was altered using industry relevant acid etching and grit blasting, and the effect of surface topography on silk coating adhesion was assessed. Florescent compounds were incorporated into the silk coatings, which were modulated for crystalline content, to achieve four days of sustained release of the compounds. This silk electrogelation technique offers a safe and relatively simple approach to generate mechanically robust, biocompatible and degradable implant coatings that can also be functionalized with bioactive compounds to modulate the local regenerative tissue environment. PMID:25545462

Novel silk fibroin films prepared by formic acid/hydroxyapatite dissolution method.

PubMed

Ming, Jinfa; Liu, Zhi; Bie, Shiyu; Zhang, Feng; Zuo, Baoqi

2014-04-01

Bombyx mori silk fibroin from the silkworm was firstly found to be soluble in formic acid/hydroxyapatite system. The rheological behavior of silk fibroin solution was significantly influenced by HAp contents in dissolved solution. At the same time, silk fibroin nanofibers were observed in dissolved solution with 103.6±20.4nm in diameter. Moreover, the structure behavior of SF films prepared by formic acid/hydroxyapatite dissolution method was examined. The secondary structure of silk fibroin films was attributed to silk II structure (β-sheet), indicating that the hydroxyapatite contents in dissolved solution were not significantly affected by the structure of silk fibroin. The X-ray diffraction results exhibited obviously hydroxyapatite crystalline nature existing in silk fibroin films; however, when the hydroxyapatite content was 5.0wt.% in dissolved solution, some hydroxyapatite crystals were converted to calcium hydrogen phosphate dehydrate in silk fibroin dissolution process. This result was also confirmed by Fourier transform infrared analysis and DSC measurement. In addition, silk fibroin films prepared by this dissolution method had higher breaking strength and extension at break. Based on these analyses, an understanding of novel SF dissolution method may provide an additional tool for designing and synthesizing advanced materials with more complex structures, which should be helpful in different fields, including biomaterial applications. Copyright © 2014 Elsevier B.V. All rights reserved.

Dating silk by capillary electrophoresis mass spectrometry.

PubMed

Moini, Mehdi; Klauenberg, Kathryn; Ballard, Mary

2011-10-01

A new capillary electrophoresis mass spectrometry (CE-MS) technique is introduced for age estimation of silk textiles based on amino acid racemization rates. With an L to D conversion half-life of ~2500 years for silk (B. mori) aspartic acid, the technique is capable of dating silk textiles ranging in age from several decades to a few-thousand-years-old. Analysis required only ~100 μg or less of silk fiber. Except for a 2 h acid hydrolysis at 110 °C, no other sample preparation is required. The CE-MS analysis takes ~20 min, consumes only nanoliters of the amino acid mixture, and provides both amino acid composition profiles and D/L ratios for ~11 amino acids.

Genetically Programmable Thermoresponsive Plasmonic Gold/Silk-Elastin Protein Core/Shell Nanoparticles

PubMed Central

2015-01-01

The design and development of future molecular photonic/electronic systems pose the challenge of integrating functional molecular building blocks in a controlled, tunable, and reproducible manner. The modular nature and fidelity of the biosynthesis method provides a unique chemistry approach to one-pot synthesis of environmental factor-responsive chimeric proteins capable of energy conversion between the desired forms. In this work, facile tuning of dynamic thermal response in plasmonic nanoparticles was facilitated by genetic engineering of the structure, size, and self-assembly of the shell silk-elastin-like protein polymers (SELPs). Recombinant DNA techniques were implemented to synthesize a new family of SELPs, S4E8Gs, with amino acid repeats of [(GVGVP)4(GGGVP)(GVGVP)3(GAGAGS)4] and tunable molecular weight. The temperature-reversible conformational switching between the hydrophilic random coils and the hydrophobic β-turns in the elastin blocks were programmed to between 50 and 60 °C by site-specific glycine mutation, as confirmed by variable-temperature proton NMR and circular dichroism (CD) spectroscopy, to trigger the nanoparticle aggregation. The dynamic self-aggregation/disaggregation of the Au-SELPs nanoparticles was regulated in size and pattern by the β-sheet-forming, thermally stable silk blocks, as revealed by transmission electron microscopy (TEM) and dynamic light scattering (DLS). The thermally reversible, shell dimension dependent, interparticle plasmon coupling was investigated by both variable-temperature UV–vis spectroscopy and finite-difference time-domain (FDTD)-based simulations. Good agreement between the calculated and measured spectra sheds light on design and synthesis of responsive plasmonic nanostructures by independently tuning the refractive index and size of the SELPs through genetic engineering. PMID:24712906

Bombyx mori silk: From mechanical properties to functionalities

NASA Astrophysics Data System (ADS)

Koh, Leng Duei

Bombyx mori silkworms are the main producer of silk worldwide. It has been used as high-end textile fibers and as surgical sutures, and is being further developed for various emerging biomedical applications including drug delivery, tissue engineering, sensing, and imaging. The silk fibroin features a hierarchical architecture consisting of beta-sheet crystallites embedded in a less ordered amorphous matrix, which accounts for its unique combination of lustre appearance, soft-to-touch texture, and impressive mechanical properties. Notably, many applications of silk take advantage of its impressive mechanical properties, which by nature surpass many natural and synthetic materials. Interestingly, both the silkworm silk and spider dragline silk share similar hierarchical architecture but possess great disparity in mechanical properties. Inspired by spider dragline silk with much superior strength and toughness, there is an ever growing interest to enhance the mechanical properties of Bombyx mori silk. Here, we design a green and facile feeding method to modulate the structures of silk fibroin at the nanoscale using citric acid (CA), and achieved greatly enhanced mechanical properties. The silk obtained (i.e., CA silk) emerges to be the intrinsically toughest silkworm silk, with mechanical properties that exceed those of the previously reported natural and enhanced silkworm silk, and compare well with those of naturally produced spider silk (including those from spiders Araneus diadematus, Nephila clavipes, etc.).The underlying interactions of CA with fibroin structures are revealed by both advanced characterizations and simulations. It is found that CA interacts with fibroin, resulted in remarkably shorter crystallites, and thus giving the outstanding strength and toughness of the CA silk. The greatly enhanced mechanical properties are expected to lead to better functionalities and wider applications of the Bombyx mori silkworm silk. Silkworms usually produce white silk with normal feed containing no xenobiotics. Here, through introducing fluorescent xenobiotics into silkworm's diet and monitoring the resulting color and fluorescence in the silkworm's body, we established an understanding on the in vivo uptake of xenobiotics in silkworms that leads to direct production of intrinsically colored and/or luminescent silk by the silkworms. The molecular properties-directed absorption, distribution and excretion of xenobiotics were investigated using a series of fluorescent molecules as model compounds in a silkworm model. The efficient uptake of xenobiotics into silk is further studied through quantitative analysis of the intrinsically colored and highly luminescent silk secreted by silkworm. Criteria for effective uptake have been established based on the relationship between the structure-dependent hydrophobicity of various dyes vs. the amount selectively absorbed into the silk. The biological incorporation of dyes into silk, in particular its fibroin is a greener method of producing the functional silk because it eliminates the need of an external dyeing process, along with the resources (water, energy and additional chemicals) associated with conventional dyeing of silk. Beyond the absorption of dyes to produce color and luminescence in the silk, this feeding concept can also be expanded to incorporate other functional molecules (e.g., drugs, antibacterial agents, perfumes and nutrients) into silk with therapeutic or nutritional value.

Efficacy of Several Pesticide Products on Brown Widow Spider (Araneae: Theridiidae) Egg Sacs and Their Penetration Through the Egg Sac Silk.

PubMed

Vetter, Richard S; Tarango, Jacob; Campbell, Kathleen A; Tham, Christine; Hayashi, Cheryl Y; Choe, Dong-Hwan

2016-02-01

Information on pesticide effects on spiders is less common than for insects; similar information for spider egg sacs is scarcer in the open literature. Spider egg sacs are typically covered with a protective silk layer. When pesticides are directly applied to egg sacs, the silk might prevent active ingredients from reaching the eggs, blocking their insecticidal effect. We investigated the impact of six water-based pesticide sprays and four oil-based aerosol products against egg sacs of brown widow spiders, Latrodectus geometricus C. L. Koch. All water-based spray products except one failed to provide significant mortality to egg sacs, resulting in successful spiderling emergence from treated egg sacs at a similar rate to untreated egg sacs. In contrast to water-based sprays, oil-based aerosols provided almost complete control, with 94-100% prevention of spiderling emergence. Penetration studies using colored pesticide products indicated that oil-based aerosols were significantly more effective in penetrating egg sac silk than were the water-based sprays, delivering the active ingredients on most (>99%) of the eggs inside the sac. The ability of pesticides to penetrate spider egg sac silk and deliver lethal doses of active ingredients to the eggs is discussed in relation to the chemical nature of egg sac silk proteins. Our study suggests that pest management procedures primarily relying on perimeter application of water-based sprays might not provide satisfactory control of brown widow spider eggs. Determination of the most effective active ingredients and carrier characteristics warrant further research to provide more effective control options for spider egg sacs. © The Authors 2015. Published by Oxford University Press on behalf of Entomological Society of America. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

Silk-based biomaterials.

PubMed

Altman, Gregory H; Diaz, Frank; Jakuba, Caroline; Calabro, Tara; Horan, Rebecca L; Chen, Jingsong; Lu, Helen; Richmond, John; Kaplan, David L

2003-02-01

Silk from the silkworm, Bombyx mori, has been used as biomedical suture material for centuries. The unique mechanical properties of these fibers provided important clinical repair options for many applications. During the past 20 years, some biocompatibility problems have been reported for silkworm silk; however, contamination from residual sericin (glue-like proteins) was the likely cause. More recent studies with well-defined silkworm silk fibers and films suggest that the core silk fibroin fibers exhibit comparable biocompatibility in vitro and in vivo with other commonly used biomaterials such as polylactic acid and collagen. Furthermore, the unique mechanical properties of the silk fibers, the diversity of side chain chemistries for 'decoration' with growth and adhesion factors, and the ability to genetically tailor the protein provide additional rationale for the exploration of this family of fibrous proteins for biomaterial applications. For example, in designing scaffolds for tissue engineering these properties are particularly relevant and recent results with bone and ligament formation in vitro support the potential role for this biomaterial in future applications. To date, studies with silks to address biomaterial and matrix scaffold needs have focused on silkworm silk. With the diversity of silk-like fibrous proteins from spiders and insects, a range of native or bioengineered variants can be expected for application to a diverse set of clinical needs.

Natural polymers for the microencapsulation of cells

PubMed Central

Gasperini, Luca; Mano, João F.; Reis, Rui L.

2014-01-01

The encapsulation of living mammalian cells within a semi-permeable hydrogel matrix is an attractive procedure for many biomedical and biotechnological applications, such as xenotransplantation, maintenance of stem cell phenotype and bioprinting of three-dimensional scaffolds for tissue engineering and regenerative medicine. In this review, we focus on naturally derived polymers that can form hydrogels under mild conditions and that are thus capable of entrapping cells within controlled volumes. Our emphasis will be on polysaccharides and proteins, including agarose, alginate, carrageenan, chitosan, gellan gum, hyaluronic acid, collagen, elastin, gelatin, fibrin and silk fibroin. We also discuss the technologies commonly employed to encapsulate cells in these hydrogels, with particular attention on microencapsulation. PMID:25232055

Nanoparticle self-assembly by a highly stable recombinant spider wrapping silk protein subunit.

PubMed

Xu, Lingling; Tremblay, Marie-Laurence; Orrell, Kathleen E; Leclerc, Jérémie; Meng, Qing; Liu, Xiang-Qin; Rainey, Jan K

2013-10-01

Artificial spider silk proteins may form fibers with exceptional strength and elasticity. Wrapping silk, or aciniform silk, is the toughest of the spider silks, and has a very different protein composition than other spider silks. Here, we present the characterization of an aciniform protein (AcSp1) subunit named W1, consisting of one AcSp1 199 residue repeat unit from Argiope trifasciata. The structural integrity of recombinant W1 is demonstrated in a variety of buffer conditions and time points. Furthermore, we show that W1 has a high thermal stability with reversible denaturation at ∼71°C and forms self-assembled nanoparticle in near-physiological conditions. W1 therefore represents a highly stable and structurally robust module for protein-based nanoparticle formation. Copyright © 2013 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.

Controlling the cell adhesion property of silk films by graft polymerization.

PubMed

Dhyani, Vartika; Singh, Neetu

2014-04-09

We report here a graft polymerization method to improve the cell adhesion property of Bombyx mori silk fibroin films. B. mori silk has evolved as a promising material for tissue engineering because of its biocompatibility and biodegradability. However, silk's hydrophobic character makes cell adhesion and proliferation difficult. Also, the lack of sufficient reactive amino acid residues makes biofunctionalization via chemical modification challenging. Our study describes a simple method that provides increased chemical handles for tuning of the surface chemistry of regenerated silk films (SFs), thus allowing manipulation of their bioactivity. By grafting pAAc and pHEMA via plasma etching, we have increased carboxylic acid and hydroxyl groups on silk, respectively. These modifications allowed us to tune the hydrophilicity of SFs and provide functional groups for bioconjugation. Our strategy also allowed us to develop silk-based surface coatings, where spatial control over cell adhesion can be achieved. This control over cell adhesion in a particular region of the SFs is difficult to obtain via existing methods of modifying the silk fibroin instead of the SF surface. Thus, our strategy will be a valuable addition to the toolkit of biofunctionalization for enhancing SFs' tissue engineering applications.

Spinning of Fibers from Aqueous Solutions

DTIC Science & Technology

2003-08-01

recombinant silk product BioSteel . Publications, patents and presentations 1. Arcidiacono, S., et al., Purification and characterization of recombinant...ABSTRACT Previous funding supporting this research focused primarily on development of the aqueous-based method for processing silk into spin solutions. Much...of this effort consisted of production of recombinant silk protein in bacterial and yeast expression systems. In spite of the small quantities

Synthetic Adhesive Attachment Discs based on Spider Pyriform Silk Architecture

NASA Astrophysics Data System (ADS)

Jain, Dharamdeep; Sahni, Vasav; Dhinojwala, Ali

2014-03-01

Among the variety of silks produced by spiders, pyriform silk is used in conjunction with the dragline silk to attach webs to different surfaces. Cob weaver spiders employ different architectural patterns to utilize the pyriform silk and form attachment joints with each pattern having a characteristic adhesive performance. The staple pin architecture is a one of the strongest attachment designs employed by spiders to attach their webs. Here we use a synthetic approach to create the a similar patterned architecture attachment discs on aluminum substrate using thermoplastic polyurethane. Measurable pull off forces are generated when the synthetic discs are peeled off a surface. This innovative adhesive strategy can be a source of design in various biomedical applications. Financial Support from National Science Foundation.

Electrodeposited gels prepared from protein alloys

PubMed Central

Lin, Yinan; Wang, Siran; Chen, Ying; Wang, Qianrui; Burke, Kelly A; Spedden, Elise M; Staii, Cristian; Weiss, Anthony S; Kaplan, David L

2015-01-01

Aim Silk-tropoelastin alloys, composed of recombinant human tropoelastin and regenerated Bombyx mori silk fibroin, are an emerging, versatile class of biomaterials endowed with tunable combinations of physical and biological properties. Electrodeposition of these alloys provides a programmable means to assemble functional gels with both spatial and temporal controllability. Materials & methods Tropoelastin-modified silk was prepared by enzymatic coupling between tyrosine residues. Hydrogel coatings were electrodeposited using two wire electrodes. Results & discussion Mechanical characterization and in vitro cell culture revealed enhanced adhesive capability and cellular response of these alloy gels as compared with electrogelled silk alone. Conclusion These electro-depositable silk-tropoelastin alloys constitute a suitable coating material for nanoparticle-based drug carriers and offer a novel opportunity for on-demand encapsulation/release of nanomedicine. PMID:25816881

The potential of turmeric (Curcuma xanthorrhiza) in agroforestry system based on silk tree (Albizia chinensis)

NASA Astrophysics Data System (ADS)

Purnomo, D.; Budiastuti, M. S.; Sakya, A. T.; Cholid, M. I.

2018-03-01

Turmeric (Curcuma xanthorrhiza Roxb.) is a traditional medicinal plant. In Indonesia, it is generally cultivated in village home gardens. Famers conducted very simpple cultivation method of turmeric, without specific maintenance and below varies tree. The experiment was conducted by cultivating turmeric below silk trees as in agroforetry system. The experiment was arranged split plot design, the first factor was three level of irradiation (turmeric monoculture/full irradiation, turmeric below silktree with pruning canopy, and turmeric below silk tree no pruning). The second factor was fertilizer NPK 15-15-15 with three levels of doses (100, 150, and 200 kg ha-1). Cultivating turmeric in agroforestry system based on silk tree which were one year old and not yet needed pruning, application of NPK 15-15-15 fertilizer 100 kg ha-1 was enough. The rhizome yield of turmeric 3 months age reaches 139 g per plant (fresh weight). Litter fall from a silk tree one year old in one year is 30 kg per tree per year.

Mechanical and Physical Properties of Recombinant Spider Silk Films Using Organic and Aqueous Solvents

PubMed Central

2015-01-01

Spider silk has exceptional mechanical and biocompatibility properties. The goal of this study was optimization of the mechanical properties of synthetic spider silk thin films made from synthetic forms of MaSp1 and MaSp2, which compose the dragline silk of Nephila clavipes. We increased the mechanical stress of MaSp1 and 2 films solubilized in both HFIP and water by adding glutaraldehyde and then stretching them in an alcohol based stretch bath. This resulted in stresses as high as 206 MPa and elongations up to 35%, which is 4× higher than the as-poured controls. Films were analyzed using NMR, XRD, and Raman, which showed that the secondary structure after solubilization and film formation in as-poured films is mainly a helical conformation. After the post-pour stretch in a methanol/water bath, the MaSp proteins in both the HFIP and water-based films formed aligned β-sheets similar to those in spider silk fibers. PMID:25030809

An amperometric new methylene blue N-mediating sensor for hydrogen peroxide based on regenerated silk fibroin as an immobilization matrix for peroxidase.

PubMed

Qian, J; Liu, Y; Liu, H; Yu, T; Deng, J

1996-05-01

A simple and effective procedure was described for the immobilization of peroxidase in regenerated silk fibroin membrane prepared from waste silk. The membranes of regenerated silk fibroin with or without peroxidase, before or after the ethanol treatment, were characterized by ir spectra. An amperometric H202 sensor, based on the immobilized peroxidase in regenerated silk fibroin membrane, in the use of new methylene blue N as an electron transfer mediator, was fabricated. The characteristics of the sensor with respect to linearity, response time, effect of pH and temperature, stability, and reproducibility were investigated. Dependences of Michaelis-Menten constant KMapp on the concentration of the mediator, and the applied potential were also studied and the results were presented. The sensor was highly sensitive to H2O2 with a detection limit of 1.0 x 10(-7)M and with response time of less than 40 s.

Construction of a functional silk-based biomaterial complex with immortalized chondrocytes in vivo.

PubMed

Ni, Yusu; Jiang, Yi; Wen, Jianchuan; Shao, Zhenzhong; Chen, Xin; Sun, Shan; Yu, Huiqian; Li, Wen

2014-04-01

To explore the feasibility of constructing a functional biomaterial complex with regenerated silk fibroin membrane and immortalized chondrocytes in vivo. Rat auricular chondrocytes (RACs) were transfected with the lentivirus vector pGC-FU-hTERT-3FLAG or pGC-FU-GFP-3FLAG, encoding the human telomerase reverse transcriptase (hTERT) or GFP gene. The effects of regenerated silk fibroin film on the adhesion, growth of immortalized chondrocytes and expression of collagen II in vitro were analyzed with immunofluorescent histochemistry. Immortalized RACs were transformed. Induction by nutrient medium promoted higher expression levels of collagen II in transformed chondrocytes. The regenerated silk fibroin film was not cytotoxic to immortalized chondrocytes and had no adverse influence on their adhesion. Collagen II expression was good in the immortalized chondrocytes in vivo. The construction of a silk-based biomaterial complex with immortalized chondrocytes may provide a feasible kind of functional biomaterial for the repair of cartilage defects in clinical applications. Copyright © 2013 Wiley Periodicals, Inc.

Review structure of silk by raman spectromicroscopy: from the spinning glands to the fibers.

PubMed

Lefèvre, Thierry; Paquet-Mercier, François; Rioux-Dubé, Jean-François; Pézolet, Michel

2012-06-01

Raman spectroscopy has long been proved to be a useful tool to study the conformation of protein-based materials such as silk. Thanks to recent developments, linearly polarized Raman spectromicroscopy has appeared very efficient to characterize the molecular structure of native single silk fibers and spinning dopes because it can provide information relative to the protein secondary structure, molecular orientation, and amino acid composition. This review will describe recent advances in the study of the structure of silk by Raman spectromicroscopy. A particular emphasis is put on the spider dragline and silkworm cocoon threads, other fibers spun by orb-weaving spiders, the spinning dope contained in their silk glands and the effect of mechanical deformation. Taken together, the results of the literature show that Raman spectromicroscopy is particularly efficient to investigate all aspects of silk structure and production. The data provided can lead to a better understanding of the structure of the silk dope, transformations occurring during the spinning process, and structure and mechanical properties of native fibers. Copyright © 2011 Wiley Periodicals, Inc.

Role of chondroitin sulphate tethered silk scaffold in cartilaginous disc tissue regeneration.

PubMed

Bhattacharjee, Maumita; Chawla, Shikha; Chameettachal, Shibu; Murab, Sumit; Bhavesh, Neel Sarovar; Ghosh, Sourabh

2016-04-12

Strategies for tissue engineering focus on scaffolds with tunable structure and morphology as well as optimum surface chemistry to simulate the anatomy and functionality of the target tissue. Silk fibroin has demonstrated its potential in supporting cartilaginous tissue formation both in vitro and in vivo. In this study, we investigate the role of controlled lamellar organization and chemical composition of biofunctionalized silk scaffolds in replicating the structural properties of the annulus region of an intervertebral disc using articular chondrocytes. Covalent attachment of chondroitin sulfate (CS) to silk is characterized. CS-conjugated silk constructs demonstrate enhanced cellular metabolic activity and chondrogenic redifferentiation potential with significantly improved mechanical properties over silk-only constructs. A matrix-assisted laser desorption ionization-time of flight analysis and protein-protein interaction studies help to generate insights into how CS conjugation can facilitate the production of disc associated matrix proteins, compared to a silk-only based construct. An in-depth understanding of the interplay between such extra cellular matrix associated proteins should help in designing more rational scaffolds for cartilaginous disc regeneration needs.

3D Bioprinting of Self-Standing Silk-Based Bioink.

PubMed

Zheng, Zhaozhu; Wu, Jianbing; Liu, Meng; Wang, Heng; Li, Chunmei; Rodriguez, María J; Li, Gang; Wang, Xiaoqin; Kaplan, David L

2018-03-01

Silk/polyethylene glycol (PEG) hydrogels are studied as self-standing bioinks for 3D printing for tissue engineering. The two components of the bioink, silk fibroin protein (silk) and PEG, are both Food and Drug Administration approved materials in drug and medical device products. Mixing PEG with silk induces silk β-sheet structure formation and thus gelation and water insolubility due to physical crosslinking. A variety of constructs with high resolution, high shape fidelity, and homogeneous gel matrices are printed. When human bone marrow mesenchymal stem cells are premixed with the silk solution prior to printing and the constructs are cultured in this medium, the cell-loaded constructs maintain their shape over at least 12 weeks. Interestingly, the cells grow faster in the higher silk concentration (10%, w/v) gel than in lower ones (7.5 and 5%, w/v), likely due to the difference in material stiffness and the amount of residual PEG remaining in the gel related to material hydrophobicity. Subcutaneous implantation of 7.5% (w/v) bioink gels with and without printed fibroblast cells in mice reveals that the cells survive and proliferate in the gel matrix for at least 6 week postimplantation. The results suggest that these silk/PEG bioink gels may provide suitable scaffold environments for cell printing and function. © 2018 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Natural Occurring Silks and Their Analogues as Materials for Nerve Conduits.

PubMed

Radtke, Christine

2016-10-20

Spider silk and its synthetic derivatives have a light weight in combination with good strength and elasticity. Their high cytocompatibility and low immunogenicity make them well suited for biomaterial products such as nerve conduits. Silk proteins slowly degrade enzymatically in vivo, thus allowing for an initial therapeutic effect such as in nerve scaffolding to facilitate endogenous repair processes, and then are removed. Silks are biopolymers naturally produced by many species of arthropods including spiders, caterpillars and mites. The silk fibers are secreted by the labial gland of the larvae of some orders of Holometabola (insects with pupa) or the spinnerets of spiders. The majority of studies using silks for biomedical applications use materials from silkworms or spiders, mostly of the genus Nephila clavipes . Silk is one of the most promising biomaterials with effects not only in nerve regeneration, but in a number of regenerative applications. The development of silks for human biomedical applications is of high scientific and clinical interest. Biomaterials in use for biomedical applications have to meet a number of requirements such as biocompatibility and elicitation of no more than a minor inflammatory response, biodegradability in a reasonable time and specific structural properties. Here we present the current status in the field of silk-based conduit development for nerve repair and discuss current advances with regard to potential clinical transfer of an implantable nerve conduit for enhancement of nerve regeneration.

Protein composition correlates with the mechanical properties of spider ( Argiope trifasciata ) dragline silk.

PubMed

Marhabaie, Mohammad; Leeper, Thomas C; Blackledge, Todd A

2014-01-13

We investigated the natural variation in silk composition and mechanical performance of the orb-weaving spider Argiope trifasciata at multiple spatial and temporal scales in order to assess how protein composition contributes to the remarkable material properties of spider dragline silk. Major ampullate silk in orb-weaving spiders consists predominantly of two proteins (MaSp1 and MaSp2) with divergent amino acid compositions and functionally different microstructures. Adjusting the expression of these two proteins therefore provides spiders with a simple mechanism to alter the material properties of their silk. We first assessed the reliability and precision of the Waters AccQ-Tag amino acid composition analysis kit for determining the amino acid composition of small quantities of spider silk. We then tested how protein composition varied within single draglines, across draglines spun by the same spider on different days, and finally between spiders. Then, we correlated chemical composition with the material properties of dragline silk. Overall, we found that the chemical composition of major ampullate silk was in general homogeneous among individuals of the same population. Variation in chemical composition was not detectable within silk spun by a single spider on a single day. However, we found that variation within a single spider's silk across different days could, in rare instances, be greater than variation among individual spiders. Most of the variation in silk composition in our investigation resulted from a small number of outliers (three out of sixteen individuals) with a recent history of stress, suggesting stress affects silk production process in orb web spiders. Based on reported sequences for MaSp genes, we developed a gene expression model showing the covariation of the most abundant amino acids in major ampullate silk. Our gene expression model supports that dragline silk composition was mostly determined by the relative abundance of MaSp1 and MaSp2. Finally, we showed that silk composition (especially proline content) strongly correlated with some measures of mechanical performance, particularly how much fibers shrunk during supercontraction as well as their breaking strains. Our findings suggest that spiders are able to change the relative expression rates of different MaSp genes to produce silk fibers with different chemical compositions, and hence, different material properties.

Atomistic model of the spider silk nanostructure

NASA Astrophysics Data System (ADS)

Keten, Sinan; Buehler, Markus J.

2010-04-01

Spider silk is an ultrastrong and extensible self-assembling biopolymer that outperforms the mechanical characteristics of many synthetic materials including steel. Here we report atomic-level structures that represent aggregates of MaSp1 proteins from the N. Clavipes silk sequence based on a bottom-up computational approach using replica exchange molecular dynamics. We discover that poly-alanine regions predominantly form distinct and orderly beta-sheet crystal domains while disorderly structures are formed by poly-glycine repeats, resembling 31-helices. These could be the molecular source of the large semicrystalline fraction observed in silks, and also form the basis of the so-called "prestretched" molecular configuration. Our structures are validated against experimental data based on dihedral angle pair calculations presented in Ramachandran plots, alpha-carbon atomic distances, as well as secondary structure content.

Sequence-structure correlations in silk: Poly-Ala repeat of N. clavipes MaSp1 is naturally optimized at a critical length scale.

PubMed

Bratzel, Graham; Buehler, Markus J

2012-03-01

Spider silk is a self-assembling biopolymer that outperforms many known materials in terms of its mechanical performance despite being constructed from simple and inferior building blocks. While experimental studies have shown that the molecular structure of silk has a direct influence on the stiffness, toughness, and failure strength of silk, few molecular-level analyses of the nanostructure of silk assemblies in particular under variations of genetic sequences have been reported. Here we report atomistic-level structures of the MaSp1 protein from the Nephila Clavipes spider dragline silk sequence, obtained using an in silico approach based on replica exchange molecular dynamics (REMD) and explicit water molecular dynamics. We apply this method to study the effects of a systematic variation of the poly-alanine repeat lengths, a parameter controlled by the genetic makeup of silk, on the resulting molecular structure of silk at the nanoscale. Confirming earlier experimental and computational work, a structural analysis reveals that poly-alanine regions in silk predominantly form distinct and orderly β-sheet crystal domains while disorderly regions are formed by glycine-rich repeats that consist of 3(10)-helix type structures and β-turns. Our predictions are directly validated against experimental data based on dihedral angle pair calculations presented in Ramachandran plots combined with an analysis of the secondary structure content. The key result of our study is our finding of a strong dependence of the resulting silk nanostructure depending on the poly-alanine length. We observe that the wildtype poly-alanine repeat length of six residues defines a critical minimum length that consistently results in clearly defined β-sheet nanocrystals. For poly-alanine lengths below six, the β-sheet nanocrystals are not well-defined or not visible at all, while for poly-alanine lengths at and above six, the characteristic nanocomposite structure of silk emerges with no significant improvement of the quality of the β-sheet nanocrystal geometry. We present a simple biophysical model that explains these computational observations based on the mechanistic insight gained from the molecular simulations. Our findings set the stage for understanding how variations in the spidroin sequence can be used to engineer the structure and thereby functional properties of this biological superfiber, and present a design strategy for the genetic optimization of spidroins for enhanced mechanical properties. The approach used here may also find application in the design of other self-assembled molecular structures and fibers and in particular biologically inspired or completely synthetic systems. Copyright © 2011 Elsevier Ltd. All rights reserved.

Conferring biological activity to native spider silk: A biofunctionalized protein-based microfiber.

PubMed

Wu, Hsuan-Chen; Quan, David N; Tsao, Chen-Yu; Liu, Yi; Terrell, Jessica L; Luo, Xiaolong; Yang, Jen-Chang; Payne, Gregory F; Bentley, William E

2017-01-01

Spider silk is an extraordinary material with physical properties comparable to the best scaffolding/structural materials, and as a fiber it can be manipulated with ease into a variety of configurations. Our work here demonstrates that natural spider silk fibers can also be used to organize biological components on and in devices through rapid and simple means. Micron scale spider silk fibers (5-10 μm in diameter) were surface modified with a variety of biological entities engineered with pentaglutamine tags via microbial transglutaminase (mTG). Enzymes, enzyme pathways, antibodies, and fluorescent proteins were all assembled onto spider silk fibers using this biomolecular engineering/biofabrication process. Additionally, arrangement of biofunctionalized fiber should in of itself generate a secondary level of biomolecular organization. Toward this end, as proofs of principle, spatially defined arrangement of biofunctionalized spider silk fiber was shown to generate effects specific to silk position in two cases. In one instance, arrangement perpendicular to a flow produced selective head and neck carcinoma cell capture on silk with antibodies complexed to conjugated protein G. In a second scenario, asymmetric bacterial chemotaxis arose from asymmetric conjugation of enzymes to arranged silk. Overall, the biofabrication processes used here were rapid, required no complex chemistries, were biologically benign, and also the resulting engineered silk microfibers were flexible, readily manipulated and functionally active. Deployed here in microfluidic environments, biofunctional spider silk fiber provides a means to convey complex biological functions over a range of scales, further extending its potential as a biomaterial in biotechnological settings. Biotechnol. Bioeng. 2017;114: 83-95. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

Multiscale mechanisms of nutritionally induced property variation in spider silks

PubMed Central

Nobbs, Madeleine; Martens, Penny J.; Tso, I-Min; Chuang, Wei-Tsung; Chang, Chung-Kai; Sheu, Hwo-Shuenn

2018-01-01

Variability in spider major ampullate (MA) silk properties at different scales has proven difficult to determine and remains an obstacle to the development of synthetic fibers mimicking MA silk performance. A multitude of techniques may be used to measure multiscale aspects of silk properties. Here we fed five species of Araneoid spider solutions that either contained protein or were protein deprived and performed silk tensile tests, small and wide-angle X-ray scattering (SAXS/WAXS), amino acid composition analyses, and silk gene expression analyses, to resolve persistent questions about how nutrient deprivation induces variations in MA silk mechanical properties across scales. Our analyses found that the properties of each spider’s silk varied differently in response to variations in their protein intake. We found changes in the crystalline and non-crystalline nanostructures to play specific roles in inducing the property variations we found. Across treatment MaSp expression patterns differed in each of the five species. We found that in most species MaSp expression and amino acid composition variations did not conform with our predictions based on a traditional MaSp expression model. In general, changes to the silk’s alanine and proline compositions influenced the alignment of the proteins within the silk’s amorphous region, which influenced silk extensibility and toughness. Variations in structural alignment in the crystalline and non-crystalline regions influenced ultimate strength independent of genetic expression. Our study provides the deepest insights thus far into the mechanisms of how MA silk properties vary from gene expression to nanostructure formations to fiber mechanics. Such knowledge is imperative for promoting the production of synthetic silk fibers. PMID:29390013

The role of 3D structure and protein conformation on the innate and adaptive immune responses to silk-based biomaterials.

PubMed

Bhattacharjee, Maumita; Schultz-Thater, Elke; Trella, Emanuele; Miot, Sylvie; Das, Sanskrita; Loparic, Marko; Ray, Alok R; Martin, Ivan; Spagnoli, Giulio C; Ghosh, Sourabh

2013-11-01

We have investigated monocyte and T cell responsiveness to silk based biomaterials of different physico-chemical characteristics. Here we report that untransformed CD14+ human monocytes respond to overnight exposure to silk fibroin-based biomaterials in tridimensional form by IL-1β and IL-6, but not IL-10 gene expression and protein production. In contrast, fibroin based materials in bidimensional form are unable to stimulate monocyte responsiveness. The elicitation of these effects critically requires contact between biomaterials and responding cells, is not sustained and becomes undetectable in longer term cultures. We also observed that NF-κβ and p38 MAP kinase play key roles in monocyte activation by silk-based biomaterials. On the other hand, fibroin based materials, irrespective of their physico-chemical characteristics appeared to be unable to induce the activation of peripheral blood T cells from healthy donors, as evaluated by the expression of activation markers and IFN-γ gene. Crown Copyright © 2013. Published by Elsevier Ltd. All rights reserved.

Subtle charge balance controls surface-nucleated self-assembly of designed biopolymers.

PubMed

Charbonneau, Céline; Kleijn, J Mieke; Cohen Stuart, Martien A

2014-03-25

We report the surface-nucleated self-assembly into fibrils of a biosynthetic amino acid polymer synthesized by the yeast Pichia pastoris. This polymer has a block-like architecture, with a central silk-like block labeled SH, responsible for the self-assembly into fibrils, and two collagen-like random coil end blocks (C) that colloidally stabilize the fibers in aqueous solution. The silk-like block contains histidine residues (pKa≈6) that are positively charged in the low pH region, which hinders self-assembly. In aqueous solution, CSHC self-assembles into fibers above a pH-dependent critical nucleation concentration Ccb. Below Ccb, where no self-assembly occurs in solution, fibril formation can be induced by a negatively charged surface (silica) in the pH range of 3.5-7. The density of the fibers at the surface and their length are controlled by a subtle balance in charge between the protein polymer and the silica surface, which is evidenced from the dependence on pH. With increasing number density of the fibers at the surface, their average length decreases. The results can be explained on the basis of a nucleation-and-growth mechanism: the surface density of fibers depends on the rate of nucleation, while their growth rate is limited by transport of proteins from solution. Screening of the charges on the surface and histidine units by adding NaCl influences the nucleation-and-growth process in a complicated fashion: at low pH, the growth is improved, while at high pH, the nucleation is limited. Under conditions where nucleation in the bulk solution is not possible, growth of the surface-nucleated fibers into the solution--away from the surface--can still occur.

Genome editing of BmFib-H gene provides an empty Bombyx mori silk gland for a highly efficient bioreactor

PubMed Central

Ma, Sanyuan; Shi, Run; Wang, Xiaogang; Liu, Yuanyuan; Chang, Jiasong; Gao, Jie; Lu, Wei; Zhang, Jianduo; Zhao, Ping; Xia, Qingyou

2014-01-01

Evolution has produced some remarkable creatures, of which silk gland is a fascinating organ that exists in a variety of insects and almost half of the 34,000 spider species. The impressive ability to secrete huge amount of pure silk protein, and to store proteins at an extremely high concentration (up to 25%) make the silk gland of Bombyx mori hold great promise to be a cost-effective platform for production of recombinant proteins. However, the extremely low production yields of the numerous reported expression systems greatly hindered the exploration and application of silk gland bioreactors. Using customized zinc finger nucleases (ZFN), we successfully performed genome editing of Bmfib-H gene, which encodes the largest and most abundant silk protein, in B. mori with efficiency higher than any previously reported. The resulted Bmfib-H knocked-out B. mori showed a smaller and empty silk gland, abnormally developed posterior silk gland cells, an extremely thin cocoon that contain only sericin proteins, and a slightly heavier pupae. We also showed that removal of endogenous Bmfib-H protein could significantly increase the expression level of exogenous protein. Furthermore, we demonstrated that the bioreactor is suitable for large scale production of protein-based materials. PMID:25359576

The effects of corn silk on glycaemic metabolism.

PubMed

Guo, Jianyou; Liu, Tongjun; Han, Linna; Liu, Yongmei

2009-11-23

Corn silk contains proteins, vitamins, carbohydrates, Ca, K, Mg and Na salts, fixed and volatile oils, steroids such as sitosterol and stigmasterol, alkaloids, saponins, tannins, and flavonoids. Base on folk remedies, corn silk has been used as an oral antidiabetic agent in China for decades. However, the hypoglycemic activity of it has not yet been understood in terms of modern pharmacological concepts. The purpose of this study is to investigate the effects of corn silk on glycaemic metabolism. Alloxan and adrenalin induced hyperglycemic mice were used in the study. The effects of corn silk on blood glucose, glycohemoglobin (HbA1c), insulin secretion, damaged pancreatic beta-cells, hepatic glycogen and gluconeogenesis in hyperglycemic mice were studied respectively. After the mice were orally administered with corn silk extract, the blood glucose and the HbA1c were significantly decreased in alloxan-induced hyperglycemic mice (p < 0.05, p < 0.01, respectively), while the level of insulin secretionn was markedly elevated in alloxa-induced hyperglycemic mice (p < 0.05). The alloxan-damaged pancreatic beta-cells of the mice were partly recovered gradually after the mice were administered with corn silk extract 15 days later. Also, the body weight of the alloxan-induced hyperglycemic mice was increased gradually. However, ascension of blood glucose induced by adrenalin and gluconeogenesis induced by L-alanine were not inhibited by corn silk extract treatment (p > 0.05). Although corn silk extract increased the level of hepatic glycogen in the alloxan-induced hyperglycemic mice, there was no significant difference between them and that of the control group(p > 0.05). Corn silk extract markedly reduced hyperglycemia in alloxan-induced diabetic mice. The action of corn silk extract on glycaemic metabolism is not via increasing glycogen and inhibiting gluconeogenesis but through increasing insulin level as well as recovering the injured beta-cells. The results suggest that corn silk extract may be used as a hypoglycemic food or medicine for hyperglycemic people in terms of this modern pharmacological study.

DNA preservation in silk.

PubMed

Liu, Yawen; Zheng, Zhaozhu; Gong, He; Liu, Meng; Guo, Shaozhe; Li, Gang; Wang, Xiaoqin; Kaplan, David L

2017-06-27

The structure of DNA is susceptible to alterations at high temperature and on changing pH, irradiation and exposure to DNase. Options to protect and preserve DNA during storage are important for applications in genetic diagnosis, identity authentication, drug development and bioresearch. In the present study, the stability of total DNA purified from human dermal fibroblast cells, as well as that of plasmid DNA, was studied in silk protein materials. The DNA/silk mixtures were stabilized on filter paper (silk/DNA + filter) or filter paper pre-coated with silk and treated with methanol (silk/DNA + PT-filter) as a route to practical utility. After air-drying and water extraction, 50-70% of the DNA and silk could be retrieved and showed a single band on electrophoretic gels. 6% silk/DNA + PT-filter samples provided improved stability in comparison with 3% silk/DNA + filter samples and DNA + filter samples for DNA preservation, with ∼40% of the band intensity remaining at 37 °C after 40 days and ∼10% after exposure to UV light for 10 hours. Quantitative analysis using the PicoGreen assay confirmed the results. The use of Tris/borate/EDTA (TBE) buffer enhanced the preservation and/or extraction of the DNA. The DNA extracted after storage maintained integrity and function based on serving as a functional template for PCR amplification of the gene for zinc finger protein 750 (ZNF750) and for transgene expression of red fluorescence protein (dsRed) in HEK293 cells. The high molecular weight and high content of a crystalline beta-sheet structure formed on the coated surfaces likely accounted for the preservation effects observed for the silk/DNA + PT-filter samples. Although similar preservation effects were also obtained for lyophilized silk/DNA samples, the rapid and simple processing available with the silk-DNA-filter membrane system makes it appealing for future applications.

Bone Tissue Engineering with Premineralized Silk Scaffolds

PubMed Central

Kim, Hyeon Joo; Kim, Ung-Jin; Kim, Hyun Suk; Li, Chunmei; Wada, Masahisa; Leisk, Gary G.; Kaplan, David L.

2009-01-01

Silks fibroin biomaterials are being explored as novel protein-based systems for cell and tissue culture. In the present study, biomimetic growth of calcium phosphate on porous silk fibroin polymeric scaffolds was explored to generate organic/inorganic composites as scaffolds for bone tissue engineering. Aqueous-derived silk fibroin scaffolds were prepared with the addition of polyaspartic acid during processing, followed by the controlled deposition of calcium phosphate by exposure to CaCl2 and Na2HPO4. These mineralized protein-composite scaffolds were subsequently seeded with human bone marrow stem cells (hMSC) and cultured in vitro for 6 weeks under osteogenic conditions with or without BMP-2. The extent of osteoconductivity was assessed by cell numbers, alkaline phosphatase and calcium deposition, along with immunohistochemistry for bone related outcomes. The results suggest increased osteoconductive outcomes with an increase in initial content of apatite and BMP-2 in the silk fibroin porous scaffolds. The premineralization of these highly porous silk fibroin protein scaffolds provided enhanced outcomes for the bone tissue engineering. PMID:18387349

Silk/nano-material hybrid: properties and functions

NASA Astrophysics Data System (ADS)

Steven, Eden; Lebedev, Victor; Laukhina, Elena; Laukhin, Vladimir; Alamo, Rufina G.; Rovira, Concepcio; Veciana, Jaume; Brooks, James S.

2014-03-01

Silk continues to emerge as a material of interest in electronics. In this work, the interaction between silk and conducting nano-materials are investigated. Simple fabrication methods, physical, electronic, thermal, and actuation properties are reported for spider silk / carbon nanotube (CNT-SS) and Bombyx mori / (BEDT-TTF)-based organic molecular conductor hybrids (ET-S). The CNT-SS fibers are produced via water and shear assisted method, resulting in fibers that are tough, custom-shapeable, flexible, and electrically conducting. For ET-S bilayer films, a layer transfer technique is developed to deposit linked crystallites of (BEDT-TTF)2I3 molecular conductor onto silk films, generating highly piezoresistive semi-transparent films. In both cases, the hybridization allows us to gain additional functions by harnessing the water-dependent properties of silk materials, for example, as humidity sensor and electrical current- or water-driven actuators. SEM, TEM, FT-IR, and resistance measurements under varying temperature, strain, and relative humidity reveal the synergistic interactions between the bio- and nano-materials. E.S. is supported by NSF-DMR 1005293.

Natural printed silk substrate circuit fabricated via surface modification using one step thermal transfer and reduction graphene oxide

NASA Astrophysics Data System (ADS)

Cao, Jiliang; Huang, Zhan; Wang, Chaoxia

2018-05-01

Graphene conductive silk substrate is a preferred material because of its biocompatibility, flexibility and comfort. A flexible natural printed silk substrate circuit was fabricated by one step transfer of graphene oxide (GO) paste from transfer paper to the surface of silk fabric and reduction of the GO to reduced graphene oxide (RGO) using a simple hot press treatment. The GO paste was obtained through ultrasonic stirring exfoliation under low temperature, and presented excellent printing rheological properties at high concentration. The silk fabric was obtained a surface electric resistance as low as 12.15 KΩ cm-1, in the concentration of GO 50 g L-1 and hot press at 220 °C for 120 s. Though the whiteness and strength decreased with the increasing of hot press temperature and time slowly, the electric conductivity of RGO surface modification silk substrate improved obviously. The surface electric resistance of RGO/silk fabrics increased from 12.15 KΩ cm-1 to 18.05 KΩ cm-1, 28.54 KΩ cm-1 and 32.53 KΩ cm-1 after 10, 20 and 30 washing cycles, respectively. The results showed that the printed silk substrate circuit has excellent washability. This process requires no chemical reductant, and the reduction efficiency and reduction degree of GO is high. This time-effective and environmentally-friendly one step thermal transfer and reduction graphene oxide onto natural silk substrate method can be easily used to production of reduced graphene oxide (RGO) based flexible printed circuit.

Fabrication of 3D porous SF/β-TCP hybrid scaffolds for bone tissue reconstruction.

PubMed

Park, Hyun Jung; Min, Kyung Dan; Lee, Min Chae; Kim, Soo Hyeon; Lee, Ok Joo; Ju, Hyung Woo; Moon, Bo Mi; Lee, Jung Min; Park, Ye Ri; Kim, Dong Wook; Jeong, Ju Yeon; Park, Chan Hum

2016-07-01

Bio-ceramic is a biomaterial actively studied in the field of bone tissue engineering. But, only certain ceramic materials can resolve the corrosion problem and possess the biological affinity of conventional metal biomaterials. Therefore, the recent development of composites of hybrid composites and polymers has been widely studied. In this study, we aimed to select the best scaffold of silk fibroin and β-TCP hybrid for bone tissue engineering. We fabricated three groups of scaffold such as SF (silk fibroin scaffold), GS (silk fibroin/small granule size of β-TCP scaffold) and GM (silk fibroin/medium granule size of β-TCP scaffold), and we compared the characteristics of each group. During characterization of the scaffold, we used scanning electron microscopy (SEM) and a Fourier transform infrared spectroscopy (FTIR) for structural analysis. We compared the physiological properties of the scaffold regarding the swelling ratio, water uptake and porosity. To evaluate the mechanical properties, we examined the compressive strength of the scaffold. During in vitro testing, we evaluated cell attachment and cell proliferation (CCK-8). Finally, we confirmed in vivo new bone regeneration from the implanted scaffolds using histological staining and micro-CT. From these evaluations, the fabricated scaffold demonstrated high porosity with good inter-pore connectivity, showed good biocompatibility and high compressive strength and modulus. In particular, the present study indicates that the GM scaffold using β-TCP accelerates new bone regeneration of implanted scaffolds. Accordingly, our scaffold is expected to act a useful application in the field of bone tissue engineering. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 104A: 1779-1787, 2016. © 2016 Wiley Periodicals, Inc.

Towards a new generation of fibre optic chemical sensors based on spider silk threads

NASA Astrophysics Data System (ADS)

Hey Tow, Kenny; Chow, Desmond M.; Vollrath, Fritz; Dicaire, Isabelle; Gheysens, Tom; Thévenaz, Luc

2017-04-01

A spider uses up to seven different types of silk, all having specific functions, to build its web. For scientists, native silk - directly extracted from spiders - is a tough, biodegradable and biocompatible thread used mainly for tissue engineering and textile applications. Blessed with outstanding optical properties, this protein strand can also be used as an optical fibre and is, moreover, intrinsically sensitive to chemical compounds. In this communication, a pioneering proof-of-concept experiment using spider silk, in its pristine condition, as a new type of fibre-optic relative humidity sensor will be demonstrated and its potential for future applications discussed.

Spider Silk Processing for Spidroin Recovery from Crossopriza Lyoni Web

NASA Astrophysics Data System (ADS)

Mohtar, J. A.; Ooi, W. L.; Yusuf, F.

2018-03-01

Spider silk is a potential biomaterial that can be used in various applications for its outstanding physicomechanical properties attributed by the spidroin composition. Efforts for commercializing spider silks have been mainly focused on the characterization of spidroins from the Entelegyne spiders for exceptional fibre construction. Hence, studies on silk proteins from the Haplogyne species remain neglected. The aim of this study is to isolate spidroin from Crossopriza lyoni web. Silk processing involved the pretreatment of fibres for the shell layer removal from the surface. A screening study was conducted to analyze the effect of temperature, incubation time and agitation speed on spidroin extraction using Ajisawa’s reagent by OFAT analysis followed by statistical optimization of the extraction process via RSM for maximal protein recovery. All parameters exerted significant effect on spidroin recovery (p<0.05) in which the maximum protein concentration (451.78 ± 0.110 µg/ml) was obtained at optimal condition of 70°C, 350 rpm and 1.25 hours. The discovery of spidroin from this study provides a basic platform for engineering spider silk to meet the demand for a variety of silk-based products in the near future.

Spider Silk-CBD-Cellulose Nanocrystal Composites: Mechanism of Assembly

PubMed Central

Meirovitch, Sigal; Shtein, Zvi; Ben-Shalom, Tal; Lapidot, Shaul; Tamburu, Carmen; Hu, Xiao; Kluge, Jonathan A.; Raviv, Uri; Kaplan, David L.; Shoseyov, Oded

2016-01-01

The fabrication of cellulose-spider silk bio-nanocomposites comprised of cellulose nanocrystals (CNCs) and recombinant spider silk protein fused to a cellulose binding domain (CBD) is described. Silk-CBD successfully binds cellulose, and unlike recombinant silk alone, silk-CBD self-assembles into microfibrils even in the absence of CNCs. Silk-CBD-CNC composite sponges and films show changes in internal structure and CNC alignment related to the addition of silk-CBD. The silk-CBD sponges exhibit improved thermal and structural characteristics in comparison to control recombinant spider silk sponges. The glass transition temperature (Tg) of the silk-CBD sponge was higher than the control silk sponge and similar to native dragline spider silk fibers. Gel filtration analysis, dynamic light scattering (DLS), small angle X-ray scattering (SAXS) and cryo-transmission electron microscopy (TEM) indicated that silk-CBD, but not the recombinant silk control, formed a nematic liquid crystalline phase similar to that observed in native spider silk during the silk spinning process. Silk-CBD microfibrils spontaneously formed in solution upon ultrasonication. We suggest a model for silk-CBD assembly that implicates CBD in the central role of driving the dimerization of spider silk monomers, a process essential to the molecular assembly of spider-silk nanofibers and silk-CNC composites. PMID:27649169

Spider Silk-CBD-Cellulose Nanocrystal Composites: Mechanism of Assembly.

PubMed

Meirovitch, Sigal; Shtein, Zvi; Ben-Shalom, Tal; Lapidot, Shaul; Tamburu, Carmen; Hu, Xiao; Kluge, Jonathan A; Raviv, Uri; Kaplan, David L; Shoseyov, Oded

2016-09-18

The fabrication of cellulose-spider silk bio-nanocomposites comprised of cellulose nanocrystals (CNCs) and recombinant spider silk protein fused to a cellulose binding domain (CBD) is described. Silk-CBD successfully binds cellulose, and unlike recombinant silk alone, silk-CBD self-assembles into microfibrils even in the absence of CNCs. Silk-CBD-CNC composite sponges and films show changes in internal structure and CNC alignment related to the addition of silk-CBD. The silk-CBD sponges exhibit improved thermal and structural characteristics in comparison to control recombinant spider silk sponges. The glass transition temperature (Tg) of the silk-CBD sponge was higher than the control silk sponge and similar to native dragline spider silk fibers. Gel filtration analysis, dynamic light scattering (DLS), small angle X-ray scattering (SAXS) and cryo-transmission electron microscopy (TEM) indicated that silk-CBD, but not the recombinant silk control, formed a nematic liquid crystalline phase similar to that observed in native spider silk during the silk spinning process. Silk-CBD microfibrils spontaneously formed in solution upon ultrasonication. We suggest a model for silk-CBD assembly that implicates CBD in the central role of driving the dimerization of spider silk monomers, a process essential to the molecular assembly of spider-silk nanofibers and silk-CNC composites.

Regulation of Silk Material Structure by Temperature-Controlled Water Vapor Annealing

PubMed Central

Hu, Xiao; Shmelev, Karen; Sun, Lin; Gil, Eun-Seok; Park, Sang-Hyug; Cebe, Peggy; Kaplan, David L.

2011-01-01

We present a simple and effective method to obtain refined control of the molecular structure of silk biomaterials through physical temperature-controlled water vapor annealing (TCWVA). The silk materials can be prepared with control of crystallinity, from a low content using conditions at 4°C (alpha-helix dominated silk I structure), to highest content of ~60% crystallinity at 100°C (beta-sheet dominated silk II structure). This new physical approach covers the range of structures previously reported to govern crystallization during the fabrication of silk materials, yet offers a simpler, green chemistry, approach with tight control of reproducibility. The transition kinetics, thermal, mechanical, and biodegradation properties of the silk films prepared at different temperatures were investigated and compared by Fourier transform infrared spectroscopy (FTIR), differential scanning calorimetry (DSC), uniaxial tensile studies, and enzymatic degradation studies. The results revealed that this new physical processing method accurately controls structure, in turn providing control of mechanical properties, thermal stability, enzyme degradation rate, and human mesenchymal stem cell interactions. The mechanistic basis for the control is through the temperature controlled regulation of water vapor, to control crystallization. Control of silk structure via TCWVA represents a significant improvement in the fabrication of silk-based biomaterials, where control of structure-property relationships is key to regulating material properties. This new approach to control crystallization also provides an entirely new green approach, avoiding common methods which use organic solvents (methanol, ethanol) or organic acids. The method described here for silk proteins would also be universal for many other structural proteins (and likely other biopolymers), where water controls chain interactions related to material properties. PMID:21425769

Sequence Identification, Recombinant Production, and Analysis of the Self-Assembly of Egg Stalk Silk Proteins from Lacewing Chrysoperla carnea.

PubMed

Neuenfeldt, Martin; Scheibel, Thomas

2017-06-13

Egg stalk silks of the common green lacewing Chrysoperla carnea likely comprise at least three different silk proteins. Based on the natural spinning process, it was hypothesized that these proteins self-assemble without shear stress, as adult lacewings do not use a spinneret. To examine this, the first sequence identification and determination of the gene expression profile of several silk proteins and various transcript variants thereof was conducted, and then the three major proteins were recombinantly produced in Escherichia coli encoded by their native complementary DNA (cDNA) sequences. Circular dichroism measurements indicated that the silk proteins in aqueous solutions had a mainly intrinsically disordered structure. The largest silk protein, which we named ChryC1, exhibited a lower critical solution temperature (LCST) behavior and self-assembled into fibers or film morphologies, depending on the conditions used. The second silk protein, ChryC2, self-assembled into nanofibrils and subsequently formed hydrogels. Circular dichroism and Fourier transform infrared spectroscopy confirmed conformational changes of both proteins into beta sheet rich structures upon assembly. ChryC3 did not self-assemble into any morphology under the tested conditions. Thereby, through this work, it could be shown that recombinant lacewing silk proteins can be produced and further used for studying the fiber formation of lacewing egg stalks.

Rate-Dependent Behavior of the Amorphous Phase of Spider Dragline Silk

PubMed Central

Patil, Sandeep P.; Markert, Bernd; Gräter, Frauke

2014-01-01

The time-dependent stress-strain behavior of spider dragline silk was already observed decades ago, and has been attributed to the disordered sequences in silk proteins, which compose the soft amorphous matrix. However, the actual molecular origin and magnitude of internal friction within the amorphous matrix has remained inaccessible, because experimentally decomposing the mechanical response of the amorphous matrix from the embedded crystalline units is challenging. Here, we used atomistic molecular dynamics simulations to obtain friction forces for the relative sliding of peptide chains of Araneus diadematus spider silk within bundles of these chains as a representative unit of the amorphous matrix in silk fibers. We computed the friction coefficient and coefficient of viscosity of the amorphous phase to be in the order of 10−6 Ns/m and 104 Ns/m2, respectively, by extrapolating our simulation data to the viscous limit. Finally, we used a finite element method for the amorphous phase, solely based on parameters derived from molecular dynamics simulations including the newly determined coefficient of viscosity. With this model the time scales of stress relaxation, creep, and hysteresis were assessed, and found to be in line with the macroscopic time-dependent response of silk fibers. Our results suggest the amorphous phase to be the primary source of viscosity in silk and open up the avenue for finite element method studies of silk fiber mechanics including viscous effects. PMID:24896131

Silk-Elastinlike Copolymers for Breast Cancer Gene Therapy

DTIC Science & Technology

2005-05-01

linearized expression vector were mixed hydrolysis of the sample in 6 N HCI at 110 ’C for 20 h. at a high monomer-to-vector molar ratio in the presence of...measurements were taken for each sample GAGTA(GA(GTGC(GGTGTA(GAGTTCCTGCGATrT’GC( for calculation of q. TAcCAC(;AGTA(;(;CGTACCGCGAGG’AGGAGTGCC(;,GGTC The...P (Pro), proline ; Q (Gln), steps are to examine the influence of polymer molecular glutamine; Q, weight equilibrium swelling ratios of hydrogels

Genetically engineered nanocarriers for drug delivery.

PubMed

Shi, Pu; Gustafson, Joshua A; MacKay, J Andrew

2014-01-01

Cytotoxicity, low water solubility, rapid clearance from circulation, and off-target side-effects are common drawbacks of conventional small-molecule drugs. To overcome these shortcomings, many multifunctional nanocarriers have been proposed to enhance drug delivery. In concept, multifunctional nanoparticles might carry multiple agents, control release rate, biodegrade, and utilize target-mediated drug delivery; however, the design of these particles presents many challenges at the stage of pharmaceutical development. An emerging solution to improve control over these particles is to turn to genetic engineering. Genetically engineered nanocarriers are precisely controlled in size and structure and can provide specific control over sites for chemical attachment of drugs. Genetically engineered drug carriers that assemble nanostructures including nanoparticles and nanofibers can be polymeric or non-polymeric. This review summarizes the recent development of applications in drug and gene delivery utilizing nanostructures of polymeric genetically engineered drug carriers such as elastin-like polypeptides, silk-like polypeptides, and silk-elastin-like protein polymers, and non-polymeric genetically engineered drug carriers such as vault proteins and viral proteins.

Genetically engineered nanocarriers for drug delivery

PubMed Central

Shi, Pu; Gustafson, Joshua A; MacKay, J Andrew

2014-01-01

Cytotoxicity, low water solubility, rapid clearance from circulation, and off-target side-effects are common drawbacks of conventional small-molecule drugs. To overcome these shortcomings, many multifunctional nanocarriers have been proposed to enhance drug delivery. In concept, multifunctional nanoparticles might carry multiple agents, control release rate, biodegrade, and utilize target-mediated drug delivery; however, the design of these particles presents many challenges at the stage of pharmaceutical development. An emerging solution to improve control over these particles is to turn to genetic engineering. Genetically engineered nanocarriers are precisely controlled in size and structure and can provide specific control over sites for chemical attachment of drugs. Genetically engineered drug carriers that assemble nanostructures including nanoparticles and nanofibers can be polymeric or non-polymeric. This review summarizes the recent development of applications in drug and gene delivery utilizing nanostructures of polymeric genetically engineered drug carriers such as elastin-like polypeptides, silk-like polypeptides, and silk-elastin-like protein polymers, and non-polymeric genetically engineered drug carriers such as vault proteins and viral proteins. PMID:24741309

Physical properties of organic and biomaterials: Fundamentals and applications

NASA Astrophysics Data System (ADS)

Steven, Eden

Silk materials are natural protein-based materials with an exceptional toughness. In addition to their toughness, silk materials also possess complex physical properties and functions resulting from a particular set of amino-acid arrangement that produces structures with crystalline beta-sheets connected by amorphous chains. Extensive studies have been performed to study their structure-function relationship leading to recent advancements in bio-integrated devices. Applications to fields other than textiles and biomedicine, however, have been scarce. In this dissertation, an investigation of the electronic properties, functionalization, and role of silk materials (spider silk and Bombyx mori cocoon silk) in the field of organic materials research is presented. The investigation is conducted from an experimental physics point of view where correlations with charge transport mechanisms in disordered, semiconducting, and insulating materials are made when appropriate. First, I present the electronic properties of spider silk fibers under ambient, humidified, iodized, polar solvent exposure, and pyrolized conditions. The conductivity is exponentially dependent on relative humidity changes and the solvent polarity. Iodine doping increases the conductivity only slightly but has pronounced effects on the pyrolization process, increasing the yield and flexibility of the pyrolized silk fibers. The iodized samples were further studied using magic angle spinning nuclear magnetic resonance (MAS-NMR) and Fourier transform infrared spectroscopy (FTIR) revealing non-homogenous iodine doping and I2 induced hydrogenation that are responsible for the minimal conductivity improvement and the pyrolization effects, respectively. Next, I present the investigation of silk fiber functionalization with gold and its role in electrical measurements. The gold functionalized silk fiber (Au-SS) is metallic down to cryogenic temperatures, has a certain amount of flexibility, and possesses magnetic field independent conductivity at low temperatures. This allows their use as micro-wires and flexible electrodes for transport measurements of small organic samples. I also found that neat spider silk fiber can be used as the mask for lithographic processes, providing a simple route of fabricating adhesive stamp electrodes for measuring transport properties of supra-micron samples in the lateral range of 15 mum--100 mum and thickness > 1 mum at low temperatures and high magnetic fields. The current-voltage characteristic of the insulating channel in tape adhesive electrodes revealed Fowler-Nordheim tunneling mechanism. For electronic sensing and actuating device applications, I have developed a simple method for silk functionalization with carbon nanotubes (CNT) facilitated by polar attraction and supercontraction, a phenomenon where silk is softened when exposed to water. Uniform CNT coating and CNT penetration into the silk fiber surface are evident from the SEM and cross-sectional TEM studies. The conductivity of the carbon nanotube functionalized silk fiber (CNT-SS) follows variable range hopping behavior with activation energy similar to that observed in buckpaper. In addition to being electrically conducting, the CNT-SS is custom-shapeable, flexible, and sensitive to humidity, allowing its use as a heart-pulse and humidity resistive sensors, as well as for current-driven actuators. Finally, I present the investigation of the processed Bombyx mori silk thin film. The silk thin film exhibits actuating and self-healing properties similar to those of a biological muscle. Proof-of-concept silk-based bio-mimetic muscle and water-based memory device are demonstrated. The silk thin film is also used as the dielectric layer of a diF-TESADT organic field effect transistor (FET) where I observed a lower operating voltage and an enhancement in the mobility of the device compared with the FET using SiO 2 dielectric layer, accompanied with an anomalous source-drain current-voltage characteristics. This dissertation aims to demonstrate the different aspects of exercising experimental physics to an inter-disciplinary research subject. The fundamental characterization and instrumentation developed in this work are intended to stimulate future discoveries by providing new experimental tools to study electronic transport properties of new materials. In addition, the device fabrication principles will be valuable for development of more environmental-friendly electronics. The treatment presented in this dissertation should serve as a roadmap for future studies of natural materials from an experimental physics point of view.

Sticky Situation: An Investigation of Robust Aqueous-Based Recombinant Spider Silk Protein Coatings and Adhesives.

PubMed

Harris, Thomas I; Gaztambide, Danielle A; Day, Breton A; Brock, Cameron L; Ruben, Ashley L; Jones, Justin A; Lewis, Randolph V

2016-11-14

The mechanical properties and biocompatibility of spider silks have made them one of the most sought after and studied natural biomaterials. A biomimetic process has been developed that uses water to solvate purified recombinant spider silk proteins (rSSps) prior to material formation. The absence of harsh organic solvents increases cost effectiveness, safety, and decreases the environmental impact of these materials. This development allows for the investigation of aqueous-based rSSps as coatings and adhesives and their potential applications. In these studies it was determined that fiber-based rSSps in nonfiber formations have the capability to coat and adhere numerous substrates, whether rough, smooth, hydrophobic, or hydrophilic. Further, these materials can be functionalized for a variety of processes. Drug-eluting coatings have been made with the capacity to release a variety of compounds in addition to their inherent ability to prevent blood clotting and biofouling. Additionally, spider silk protein adhesives are strong enough to outperform some conventional glues and still display favorable tissue implantation properties. The physical properties, corresponding capabilities, and potential applications of these nonfibrous materials were characterized in this study. Mechanical properties, ease of manufacturing, biodegradability, biocompatibility, and functionality are the hallmarks of these revolutionary spider silk protein materials.

An antibacterial and absorbable silk-based fixation material with impressive mechanical properties and biocompatibility

NASA Astrophysics Data System (ADS)

Shi, Chenglong; Pu, Xiaobing; Zheng, Guan; Feng, Xinglong; Yang, Xuan; Zhang, Baoliang; Zhang, Yu; Yin, Qingshui; Xia, Hong

2016-11-01

Implant-associated infections and non-absorbing materials are two important reasons for a second surgical procedure to remove internal fixation devices after an orthopedic internal fixation surgery. The objective of this study was to produce an antibacterial and absorbable fixation screw by adding gentamicin to silk-based materials. The antibacterial activity was assessed against Staphylococcus aureus (S. aureus) and Escherichia coli (E. coli) in vitro by plate cultivation and scanning electron microscopy (SEM). We also investigated the properties, such as the mechanical features, swelling properties, biocompatibility and degradation, of gentamicin-loaded silk-based screws (GSS) in vitro. The GSS showed significant bactericidal effects against S. aureus and E. coli. The antibacterial activity remained high even after 4 weeks of immersion in protease solution. In addition, the GSS maintained the remarkable mechanical properties and excellent biocompatibility of pure silk-based screws (PSS). Interestingly, after gentamicin incorporation, the degradation rate and water-absorbing capacity increased and decreased, respectively. These GSS provide both impressive material properties and antibacterial activity and have great potential for use in orthopedic implants to reduce the incidence of second surgeries.

Phenol red-silk tyrosine cross-linked hydrogels.

PubMed

Sundarakrishnan, Aswin; Herrero Acero, Enrique; Coburn, Jeannine; Chwalek, Karolina; Partlow, Benjamin; Kaplan, David L

2016-09-15

Phenol red is a cytocompatible pH sensing dye that is commonly added to cell culture media, but removed from some media formulations due to its structural mimicry of estrogen. Phenol red free media is also used during live cell imaging, to avoid absorbance and fluorescence quenching of fluorophores. To overcome these complications, we developed cytocompatible and degradable phenol red-silk tyrosine cross-linked hydrogels using horseradish peroxidase (HRP) enzyme and hydrogen peroxide (H2O2). Phenol red added to silk during tyrosine crosslinking accelerated di-tyrosine formation in a concentration-dependent reaction. Phenol red diffusion studies and UV-Vis spectra of phenol red-silk tyrosine hydrogels at different pHs showed altered absorption bands, confirming entrapment of dye within the hydrogel network. LC-MS of HRP-reacted phenol red and N-acetyl-l-tyrosine reaction products confirmed covalent bonds between the phenolic hydroxyl group of phenol red and tyrosine on the silk. At lower phenol red concentrations, leak-proof hydrogels which did not release phenol red were fabricated and found to be cytocompatible based on live-dead staining and alamar blue assessments of encapsulated fibroblasts. Due to the spectral overlap between phenol red absorbance at 415nm and di-tyrosine fluorescence at 417nm, phenol red-silk hydrogels provide both absorbance and fluorescence-based pH sensing. With an average pKa of 6.8 and good cytocompatibiltiy, phenol red-silk hydrogels are useful for pH sensing in phenol red free systems, cellular microenvironments and bioreactors. Phenol red entrapped within hydrogels facilitates pH sensing in phenol red free environments. Leak-proof phenol red based pH sensors require covalent binding techniques, but are complicated due to the lack of amino or carboxyl groups on phenol red. Currently, there is no simple, reliable technique to covalently link phenol red to hydrogel matrices, for real-time pH sensing in cell culture environments. Herein, we take advantage of phenolic groups for covalent linkage of phenol red to silk tyrosine in the presence of HRP and H2O2. The novelty of the current system stems from its simplicity and the use of silk protein to create a cytocompatible, degradable sensor capable of real-time pH sensing in cell culture microenvironments. Copyright © 2016 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

The other prey-capture silk: Fibres made by glow-worms (Diptera: Keroplatidae) comprise cross-β-sheet crystallites in an abundant amorphous fraction.

PubMed

Walker, Andrew A; Weisman, Sarah; Trueman, Holly E; Merritt, David J; Sutherland, Tara D

2015-09-01

Glow-worms (larvae of dipteran genus Arachnocampa) are restricted to moist habitats where they capture flying prey using snares composed of highly extensible silk fibres and sticky mucus droplets. Little is known about the composition or structure of glow-worm snares, or the extent of possible convergence between glow-worm and arachnid capture silks. We characterised Arachnocampa richardsae silk and mucus using X-ray scattering, Fourier transform infrared spectroscopy and amino acid analysis. Silk but not mucus contained crystallites of the cross-β-sheet type, which occur in unrelated insect silks but have not been reported previously in fibres used for prey capture. Mucus proteins were rich in Gly (28.5%) and existed in predominantly a random coil structure, typical of many adhesive proteins. In contrast, the silk fibres were unusually rich in charged and polar residues, particularly Lys (18.1%), which we propose is related to their use in a highly hydrated state. Comparison of X-ray scattering, infrared spectroscopy and amino acid analysis data suggests that silk fibres contain a high fraction of disordered protein. We suggest that in the native hydrated state, silk fibres are capable of extension via deformation of both disordered regions and cross-β-sheet crystallites, and that high extensibility is an adaptation promoting successful prey capture. This study illustrates the rich variety of protein motifs that are available for recruitment into biopolymers, and how convergently evolved materials can nevertheless be based on fundamentally different protein structures. Crown Copyright © 2015. Published by Elsevier Inc. All rights reserved.

Nanoscale probing of electron-regulated structural transitions in silk proteins by near-field IR imaging and nano-spectroscopy

PubMed Central

Qin, Nan; Zhang, Shaoqing; Jiang, Jianjuan; Corder, Stephanie Gilbert; Qian, Zhigang; Zhou, Zhitao; Lee, Woonsoo; Liu, Keyin; Wang, Xiaohan; Li, Xinxin; Shi, Zhifeng; Mao, Ying; Bechtel, Hans A.; Martin, Michael C.; Xia, Xiaoxia; Marelli, Benedetto; Kaplan, David L.; Omenetto, Fiorenzo G.; Liu, Mengkun; Tao, Tiger H.

2016-01-01

Silk protein fibres produced by silkworms and spiders are renowned for their unparalleled mechanical strength and extensibility arising from their high-β-sheet crystal contents as natural materials. Investigation of β-sheet-oriented conformational transitions in silk proteins at the nanoscale remains a challenge using conventional imaging techniques given their limitations in chemical sensitivity or limited spatial resolution. Here, we report on electron-regulated nanoscale polymorphic transitions in silk proteins revealed by near-field infrared imaging and nano-spectroscopy at resolutions approaching the molecular level. The ability to locally probe nanoscale protein structural transitions combined with nanometre-precision electron-beam lithography offers us the capability to finely control the structure of silk proteins in two and three dimensions. Our work paves the way for unlocking essential nanoscopic protein structures and critical conditions for electron-induced conformational transitions, offering new rules to design protein-based nanoarchitectures. PMID:27713412

Unravelling the biodiversity of nanoscale signatures of spider silk fibres

NASA Astrophysics Data System (ADS)

Silva, Luciano P.; Rech, Elibio L.

2013-12-01

Living organisms are masters at designing outstanding self-assembled nanostructures through a hierarchical organization of modular proteins. Protein-based biopolymers improved and selected by the driving forces of molecular evolution are among the most impressive archetypes of nanomaterials. One of these biomacromolecules is the myriad of compound fibroins of spider silks, which combine surprisingly high tensile strength with great elasticity. However, no consensus on the nano-organization of spider silk fibres has been reached. Here we explore the biodiversity of spider silk fibres, focusing on nanoscale characterization with high-resolution atomic force microscopy. Our results reveal an evolution of the nanoroughness, nanostiffness, nanoviscoelastic, nanotribological and nanoelectric organization of microfibres, even when they share similar sizes and shapes. These features are related to unique aspects of their molecular structures. The results show that combined nanoscale analyses of spider silks may enable the screening of appropriate motifs for bioengineering synthetic fibres from recombinant proteins.

Silk: a potential medium for tissue engineering.

PubMed

Sobajo, Cassandra; Behzad, Farhad; Yuan, Xue-Feng; Bayat, Ardeshir

2008-01-01

Human skin is a complex bilayered organ that serves as a protective barrier against the environment. The loss of integrity of skin by traumatic experiences such as burns and ulcers may result in considerable disability or ultimately death. Therefore, in skin injuries, adequate dermal substitutes are among primary care targets, aimed at replacing the structural and functional properties of native skin. To date, there are very few single application tissue-engineered dermal constructs fulfilling this criterion. Silk produced by the domestic silkworm, Bombyx mori, has a long history of use in medicine. It has recently been increasingly investigated as a promising biomaterial for dermal constructs. Silk contains 2 fibrous proteins, sericin and fibroin. Each one exhibits unique mechanical and biological properties. Comprehensive review of randomized-controlled trials investigating current dermal constructs and the structures and properties of silk-based constructs on wound healing. This review revealed that silk-fibroin is regarded as the most promising biomaterial, providing options for the construction of tissue-engineered skin. The research available indicates that silk fibroin is a suitable biomaterial scaffold for the provision of adequate dermal constructs.

Sericin removal from raw Bombyx mori silk scaffolds of high hierarchical order.

PubMed

Teuschl, Andreas Herbert; van Griensven, Martijn; Redl, Heinz

2014-05-01

Silk fibroin has previously been described as a promising candidate for ligament tissue engineering (TE) approaches. For biocompatibility reasons, silkworm silk requires removal of sericin, which can elicit adverse immune responses in the human body. One disadvantage of the required degumming process is the alteration of the silk fiber structural properties, which can hinder textile engineering of high order hierarchical structures. Therefore, the aim of this study was to find a way to remove sericin from a compact and highly ordered raw silk fiber matrix. The wire rope design of the test model scaffold comprises several levels of geometric hierarchy. Commonly used degumming solutions fail in removing sericin in this wire rope design. Weight loss measurements, picric acid and carmine staining as well as scanning electron microscopy demonstrated that the removal of sericin from the model scaffold of a wire rope design can be achieved through a borate buffer-based system. Furthermore, the borate buffer degummed silks were shown to be nontoxic and did not alter cell proliferation behavior. The possibility to remove sericin after the textile engineering process has taken place eases the production of highly ordered scaffold structures and may expand the use of silk as scaffold material in further TE and regenerative medicine applications.

Enhanced Cellular Adhesion on Titanium by Silk Functionalized with titanium binding and RGD peptides

PubMed Central

Vidal, Guillaume; Blanchi, Thomas; Mieszawska, Aneta J.; Calabrese, Rossella; Rossi, Claire; Vigneron, Pascale; Duval, Jean-Luc; Kaplan, David L.; Egles, Christophe

2012-01-01

Soft tissue adhesion on titanium represents a challenge for implantable materials. In order to improve adhesion at the cell/material interface we used a new approach based on the molecular recognition of titanium by specific peptides. Silk fibroin protein was chemically grafted with titanium binding peptide (TiBP) to increase adsorption of these chimeric proteins to the metal surface. Quartz Crystal Microbalance was used to quantify the specific adsorption of TiBP-functionalized silk and an increase in protein deposition by more than 35% was demonstrated due to the presence of the binding peptide. A silk protein grafted with TiBP and fibronectin-derived RGD peptide was then prepared. The adherence of fibroblasts on the titanium surface modified with the multifunctional silk coating demonstrated an increase in the number of adhering cells by 60%. The improved adhesion was demonstrated by Scanning Electron Microscopy and immunocytochemical staining of focal contact points. Chick embryo organotypic culture also revealed strong adhesion of endothelial cells expanding on the multifunctional silk-peptide coating. These results demonstrated that silk functionalized with TiBP and RGD represents a promising approach to modify cell-biomaterial interfaces, opening new perspectives for implantable medical devices, especially when reendothelialization is required. PMID:22975628

Spider Silks-Biomimetics Beyond Silk Fibers: Hydrogels, films & Adhesives from Aqueous Recombinant Spider Silk dopes: A Synchrotron X-Ray Nano-Structural Study

NASA Astrophysics Data System (ADS)

Sampath, Sujatha; Jones, Justin; Harris, Thomas; Lewis, Randolph

2015-03-01

With a combination of high strength and extensibility, spider silk's (SS) mechanical properties surpass those of any man made fiber. The superior properties are due to the primary protein composition and the complex hierarchical structural organization from nanoscale to macroscopic length scales. Considerable progress has been made to synthetically mimic the production of fibers based on SS proteins. We present synchrotron x-ray micro diffraction (SyXRD) results on new fibers and gels (hydrogels, lyogels) from recombinant SS protein water-soluble dopes. Novelty in these materials is two-fold: water based rather than widely used HFIP acid synthesis, makes them safe in medical applications (replacement for tendons & ligaments). Secondly, hydrogels morphology render them as excellent carriers for targeted drug delivery biomedical applications. SyXRD results reveal semi-crystalline structure with ordered beta-sheets and relatively high degree of axial orientation in the fibers, making them the closest yet to natural spider silks. SyXRD on the gels elucidate the structural transformations during the self-recovery process through mechanical removal and addition of water. Studies correlating the observed structural changes to mechanical properties are underway.

Reactive Inkjet Printing of Biocompatible Enzyme Powered Silk Micro-Rockets.

PubMed

Gregory, David A; Zhang, Yu; Smith, Patrick J; Zhao, Xiubo; Ebbens, Stephen J

2016-08-01

Inkjet-printed enzyme-powered silk-based micro-rockets are able to undergo autonomous motion in a vast variety of fluidic environments including complex media such as human serum. By means of digital inkjet printing it is possible to alter the catalyst distribution simply and generate varying trajectory behavior of these micro-rockets. Made of silk scaffolds containing enzymes these micro-rockets are highly biocompatible and non-biofouling. © 2016 The Authors. Published by WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Gold nanoparticle-embedded silk protein-ZnO nanorod hybrids for flexible bio-photonic devices

NASA Astrophysics Data System (ADS)

Gogurla, Narendar; Kundu, Subhas C.; Ray, Samit K.

2017-04-01

Silk protein has been used as a biopolymer substrate for flexible photonic devices. Here, we demonstrate ZnO nanorod array hybrid photodetectors on Au nanoparticle-embedded silk protein for flexible optoelectronics. Hybrid samples exhibit optical absorption at the band edge of ZnO as well as plasmonic energy due to Au nanoparticles, making them attractive for selective UV and visible wavelength detection. The device prepared on Au-silk protein shows a much lower dark current and a higher photo to dark-current ratio of ∼105 as compared to the control sample without Au nanoparticles. The hybrid device also exhibits a higher specific detectivity due to higher responsivity arising from the photo-generated hole trapping by Au nanoparticles. Sharp pulses in the transient photocurrent have been observed in devices prepared on glass and Au-silk protein substrates due to the light induced pyroelectric effect of ZnO, enabling the demonstration of self-powered photodetectors at zero bias. Flexible hybrid detectors have been demonstrated on Au-silk/polyethylene terephthalate substrates, exhibiting characteristics similar to those fabricated on rigid glass substrates. A study of the performance of photodetectors with different bending angles indicates very good mechanical stability of silk protein based flexible devices. This novel concept of ZnO nanorod array photodetectors on a natural silk protein platform provides an opportunity to realize integrated flexible and self-powered bio-photonic devices for medical applications in near future.

Rate-dependent behavior of the amorphous phase of spider dragline silk.

PubMed

Patil, Sandeep P; Markert, Bernd; Gräter, Frauke

2014-06-03

The time-dependent stress-strain behavior of spider dragline silk was already observed decades ago, and has been attributed to the disordered sequences in silk proteins, which compose the soft amorphous matrix. However, the actual molecular origin and magnitude of internal friction within the amorphous matrix has remained inaccessible, because experimentally decomposing the mechanical response of the amorphous matrix from the embedded crystalline units is challenging. Here, we used atomistic molecular dynamics simulations to obtain friction forces for the relative sliding of peptide chains of Araneus diadematus spider silk within bundles of these chains as a representative unit of the amorphous matrix in silk fibers. We computed the friction coefficient and coefficient of viscosity of the amorphous phase to be in the order of 10(-6) Ns/m and 10(4) Ns/m(2), respectively, by extrapolating our simulation data to the viscous limit. Finally, we used a finite element method for the amorphous phase, solely based on parameters derived from molecular dynamics simulations including the newly determined coefficient of viscosity. With this model the time scales of stress relaxation, creep, and hysteresis were assessed, and found to be in line with the macroscopic time-dependent response of silk fibers. Our results suggest the amorphous phase to be the primary source of viscosity in silk and open up the avenue for finite element method studies of silk fiber mechanics including viscous effects. Copyright © 2014 Biophysical Society. Published by Elsevier Inc. All rights reserved.

Understanding the Mechanical Properties and Structure Transition of Antheraea pernyi Silk Fiber Induced by Its Contraction.

PubMed

Wang, Yu; Wen, Jianchuan; Peng, Bo; Hu, Bingwen; Chen, Xin; Shao, Zhengzhong

2018-02-23

Like most major ampullate silks of spider, the length of Antheraea pernyi silkworm silk can shrink to a certain degree when the fiber is in contact with water. However, what happens in terms of molecule chain level and how it correlates to the mechanical properties of the silk during its contraction is not yet fully understood. Here, we investigate the water-induced mechanical property changes as well as the structure transition of two kinds of A. pernyi silk fiber, which are forcibly reeled from two different individuals (silkworm a and silkworm b; the silk fiber from either one represents the lower and upper limit of the distribution of mechanical properties, respectively). The tensile test results present that most of the mechanical parameters except the post-yield modulus and breaking strain for both silk fibers have the same variation trend before and after their water contraction. Synchrotron FTIR and Raman spectra show that the native filament from silkworm a contains more α-helix structures than that in silkworm b filament, and these α-helices are partially converted to β-sheet structures after the contraction of the fibers, while the order of both β-sheet and α-helix slightly increase. On the other side, the content and orientation of both secondary structural components in silkworm b fiber keep unchanged, no matter if it is native or contracted. 13 C CP/MAS NMR results further indicate that the α-helix/random coil to β-sheet conformational transition that occurred in the silk of silkworm a corresponds the Ala residues. Based upon these results, the detailed structure transition models of both as-reeled A. pernyi silk fibers during water contraction are proposed finally to interpret their properties transformation.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lu, Q.; Hu, X; Wang, X

Water-insoluble regenerated silk materials are normally produced by increasing the {beta}-sheet content (silk II). In the present study water-insoluble silk films were prepared by controlling the very slow drying of Bombyx mori silk solutions, resulting in the formation of stable films with a predominant silk I instead of silk II structure. Wide angle X-ray scattering indicated that the silk films stabilized by slow drying were mainly composed of silk I rather than silk II, while water- and methanol-annealed silk films had a higher silk II content. The silk films prepared by slow drying had a globule-like structure at the coremore » surrounded by nano-filaments. The core region was composed of silk I and silk II, surrounded by hydrophilic nano-filaments containing random turns and {alpha}-helix secondary structures. The insoluble silk films prepared by slow drying had unique thermal, mechanical and degradative properties. Differential scanning calorimetry results revealed that silk I crystals had stable thermal properties up to 250 C, without crystallization above the T{sub g}, but degraded at lower temperatures than silk II structure. Compared with water- and methanol-annealed films the films prepared by slow drying had better mechanical ductility and were more rapidly enzymatically degraded, reflecting the differences in secondary structure achieved via differences in post processing of the cast silk films. Importantly, the silk I structure, a key intermediate secondary structure for the formation of mechanically robust natural silk fibers, was successfully generated by the present approach of very slow drying, mimicking the natural process. The results also point to a new mode of generating new types of silk biomaterials with enhanced mechanical properties and increased degradation rates, while maintaining water insolubility, along with a low {beta}-sheet content.« less

Punctuated evolution of viscid silk in spider orb webs supported by mechanical behavior of wet cribellate silk

NASA Astrophysics Data System (ADS)

Piorkowski, Dakota; Blackledge, Todd A.

2017-08-01

The origin of viscid capture silk in orb webs, from cribellate silk-spinning ancestors, is a key innovation correlated with significant diversification of web-building spiders. Ancestral cribellate silk consists of dry nanofibrils surrounding a stiff, axial fiber that adheres to prey through van der Waals interactions, capillary forces, and physical entanglement. In contrast, viscid silk uses chemically adhesive aqueous glue coated onto a highly compliant and extensible flagelliform core silk. The extensibility of the flagelliform fiber accounts for half of the total work of adhesion for viscid silk and is enabled by water in the aqueous coating. Recent cDNA libraries revealed the expression of flagelliform silk proteins in cribellate orb-weaving spiders. We hypothesized that the presence of flagelliform proteins in cribellate silk could have allowed for a gradual shift in mechanical performance of cribellate axial silk, whose effect was masked by the dry nature of its adhesive. We measured supercontraction and mechanical performance of cribellate axial silk, in wet and dry states, for two species of cribellate orb web-weaving spiders to see if water enabled flagelliform silk-like performance. We found that compliance and extensibility of wet cribellate silk increased compared to dry state as expected. However, when compared to other silk types, the response to water was more similar to other web silks, like major and minor ampullate silk, than to viscid silk. These findings support the punctuated evolution of viscid silk mechanical performance.

Applicability of biotechnologically produced insect silks.

PubMed

Herold, Heike M; Scheibel, Thomas

2017-09-26

Silks are structural proteins produced by arthropods. Besides the well-known cocoon silk, which is produced by larvae of the silk moth Bombyx mori to undergo metamorphosis inside their silken shelter (and which is also used for textile production by men since millennia), numerous further less known silk-producing animals exist. The ability to produce silk evolved multiple independent times during evolution, and the fact that silk was subject to convergent evolution gave rise to an abundant natural diversity of silk proteins. Silks are used in air, under water, or like honey bee silk in the hydrophobic, waxen environment of the bee hive. The good mechanical properties of insect silk fibres together with their non-toxic, biocompatible, and biodegradable nature renders these materials appealing for both technical and biomedical applications. Although nature provides a great diversity of material properties, the variation in quality inherent in materials from natural sources together with low availability (except from silkworm silk) impeded the development of applications of silks. To overcome these two drawbacks, in recent years, recombinant silks gained more and more interest, as the biotechnological production of silk proteins allows for a scalable production at constant quality. This review summarises recent developments in recombinant silk production as well as technical procedures to process recombinant silk proteins into fibres, films, and hydrogels.

Microwave assisted synthesis of luminescent carbonaceous nanoparticles from silk fibroin for bioimaging.

PubMed

Gao, Hongzhi; Teng, Choon Peng; Huang, Donghong; Xu, Wanqing; Zheng, Chaohui; Chen, Yisong; Liu, Minghuan; Yang, Da-Peng; Lin, Ming; Li, Zibiao; Ye, Enyi

2017-11-01

Bombyx mori silk as a natural protein based biopolymer with high nitrogen content, is abundant and sustainable because of its mass product all over the world per year. In this study, we developed a facile and fast microwave-assisted synthesis of luminescent carbonaceous nanoparticles using Bombyx mori silk fibroin and silk solution as the precursors. As a result, the obtained carbonaceous nanoparticles exhibit a photoluminescence quantum yield of ~20%, high stability, low cytotoxicity, high biocompatibility. Most importantly, we successfully demonstrated bioimaging using these luminescent carbonaceous nanoparticles with excitation dependent luminescence. In addition, the microwave-assisted hydrothermal method can be extended to convert other biomass into functional nanomaterials. Copyright © 2017 Elsevier B.V. All rights reserved.

Plasticity in Major Ampullate Silk Production in Relation to Spider Phylogeny and Ecology

PubMed Central

Boutry, Cecilia; Řezáč, Milan; Blackledge, Todd Alan

2011-01-01

Spider major ampullate silk is a high-performance biomaterial that has received much attention. However, most studies ignore plasticity in silk properties. A better understanding of silk plasticity could clarify the relative importance of chemical composition versus processing of silk dope for silk properties. It could also provide insight into how control of silk properties relates to spider ecology and silk uses. We compared silk plasticity (defined as variation in the properties of silk spun by a spider under different conditions) between three spider clades in relation to their anatomy and silk biochemistry. We found that silk plasticity exists in RTA clade and orbicularian spiders, two clades that differ in their silk biochemistry. Orbiculariae seem less dependent on external spinning conditions. They probably use a valve in their spinning duct to control friction forces and speed during spinning. Our results suggest that plasticity results from different processing of the silk dope in the spinning duct. Orbicularian spiders seem to display better control of silk properties, perhaps in relation to their more complex spinning duct valve. PMID:21818328

Engineered hybrid spider silk particles as delivery system for peptide vaccines.

PubMed

Lucke, Matthias; Mottas, Inès; Herbst, Tina; Hotz, Christian; Römer, Lin; Schierling, Martina; Herold, Heike M; Slotta, Ute; Spinetti, Thibaud; Scheibel, Thomas; Winter, Gerhard; Bourquin, Carole; Engert, Julia

2018-07-01

The generation of strong T-cell immunity is one of the main challenges for the development of successful vaccines against cancer and major infectious diseases. Here we have engineered spider silk particles as delivery system for a peptide-based vaccination that leads to effective priming of cytotoxic T-cells. The recombinant spider silk protein eADF4(C16) was fused to the antigenic peptide from ovalbumin, either without linker or with a cathepsin cleavable peptide linker. Particles prepared from the hybrid proteins were taken up by dendritic cells, which are essential for T-cell priming, and successfully activated cytotoxic T-cells, without signs of immunotoxicity or unspecific immunostimulatory activity. Upon subcutaneous injection in mice, the particles were taken up by dendritic cells and accumulated in the lymph nodes, where immune responses are generated. Particles from hybrid proteins containing a cathepsin-cleavable linker induced a strong antigen-specific proliferation of cytotoxic T-cells in vivo, even in the absence of a vaccine adjuvant. We thus demonstrate the efficacy of a new vaccine strategy using a protein-based all-in-one vaccination system, where spider silk particles serve as carriers with an incorporated peptide antigen. Our study further suggests that engineered spider silk-based vaccines are extremely stable, easy to manufacture, and readily customizable. Copyright © 2018 Elsevier Ltd. All rights reserved.

Precise Protein Photolithography (P3): High Performance Biopatterning Using Silk Fibroin Light Chain as the Resist

PubMed Central

Liu, Wanpeng; Zhou, Zhitao; Zhang, Shaoqing; Shi, Zhifeng; Tabarini, Justin; Lee, Woonsoo; Zhang, Yeshun; Gilbert Corder, S. N.; Li, Xinxin; Dong, Fei; Cheng, Liang; Liu, Mengkun; Kaplan, David L.; Omenetto, Fiorenzo G.

2017-01-01

Precise patterning of biomaterials has widespread applications, including drug release, degradable implants, tissue engineering, and regenerative medicine. Patterning of protein‐based microstructures using UV‐photolithography has been demonstrated using protein as the resist material. The Achilles heel of existing protein‐based biophotoresists is the inevitable wide molecular weight distribution during the protein extraction/regeneration process, hindering their practical uses in the semiconductor industry where reliability and repeatability are paramount. A wafer‐scale high resolution patterning of bio‐microstructures using well‐defined silk fibroin light chain as the resist material is presented showing unprecedent performances. The lithographic and etching performance of silk fibroin light chain resists are evaluated systematically and the underlying mechanisms are thoroughly discussed. The micropatterned silk structures are tested as cellular substrates for the successful spatial guidance of fetal neural stems cells seeded on the patterned substrates. The enhanced patterning resolution, the improved etch resistance, and the inherent biocompatibility of such protein‐based photoresist provide new opportunities in fabricating large scale biocompatible functional microstructures. PMID:28932678

[Application of silk-based tissue engineering scaffold for tendon / ligament regeneration].

PubMed

Hu, Yejun; Le, Huihui; Jin, Zhangchu; Chen, Xiao; Yin, Zi; Shen, Weiliang; Ouyang, Hongwei

2016-03-01

Tendon/ligament injury is one of the most common impairments in sports medicine. The traditional treatments of damaged tissue repair are unsatisfactory, especially for athletes, due to lack of donor and immune rejection. The strategy of tissue engineering may break through these limitations, and bring new hopes to tendon/ligament repair, even regeneration. Silk is a kind of natural biomaterials, which has good biocompatibility, wide range of mechanical properties and tunable physical structures; so it could be applied as tendon/ligament tissue engineering scaffolds. The silk-based scaffold has robust mechanical properties; combined with other biological ingredients, it could increase the surface area, promote more cell adhesion and improve the biocompatibility. The potential clinical application of silk-based scaffold has been confirmed by in vivo studies on tendon/ligament repairing, such as anterior cruciate ligament, medial collateral ligament, achilles tendon and rotator cuff. To develop novel biomechanically stable and host integrated tissue engineered tendon/ligament needs more further micro and macro studies, combined with product development and clinical application, which will give new hope to patients with tendon/ligament injury.

Synergistic toughening of composite fibres by self-alignment of reduced graphene oxide and carbon nanotubes.

PubMed

Shin, Min Kyoon; Lee, Bommy; Kim, Shi Hyeong; Lee, Jae Ah; Spinks, Geoffrey M; Gambhir, Sanjeev; Wallace, Gordon G; Kozlov, Mikhail E; Baughman, Ray H; Kim, Seon Jeong

2012-01-31

The extraordinary properties of graphene and carbon nanotubes motivate the development of methods for their use in producing continuous, strong, tough fibres. Previous work has shown that the toughness of the carbon nanotube-reinforced polymer fibres exceeds that of previously known materials. Here we show that further increased toughness results from combining carbon nanotubes and reduced graphene oxide flakes in solution-spun polymer fibres. The gravimetric toughness approaches 1,000 J g(-1), far exceeding spider dragline silk (165 J g(-1)) and Kevlar (78 J g(-1)). This toughness enhancement is consistent with the observed formation of an interconnected network of partially aligned reduced graphene oxide flakes and carbon nanotubes during solution spinning, which act to deflect cracks and allow energy-consuming polymer deformation. Toughness is sensitive to the volume ratio of the reduced graphene oxide flakes to the carbon nanotubes in the spinning solution and the degree of graphene oxidation. The hybrid fibres were sewable and weavable, and could be shaped into high-modulus helical springs.

Building Interfaces: Mechanisms, fabrication, and applications at the biotic/abiotic interface for silk fibroin based bioelectronic and biooptical devices

NASA Astrophysics Data System (ADS)

Brenckle, Mark

Recent efforts in bioelectronics and biooptics have led to a shift in the materials and form factors used to make medical devices, including high performance, implantable, and wearable sensors. In this context, biopolymer-based devices must be processed to interface the soft, curvilinear biological world with the rigid, inorganic world of traditional electronics and optics. This poses new material-specific fabrication challenges in designing such devices, which in turn requires further understanding of the fundamental physical behaviors of the materials in question. As a biopolymer, silk fibroin protein has remarkable promise in this space, due to its bioresorbability, mechanical strength, optical clarity, ability to be reshaped on the micro- and nano-scale, and ability to stabilize labile compounds. Application of this material to devices at the biotic/abiotic interface will require the development of fabrication techniques for nano-patterning, lithography, multilayer adhesion, and transfer printing in silk materials. In this work, we address this need through fundamental study of the thermal and diffusional properties of silk protein as it relates to these fabrication strategies. We then leverage these properties to fabricate devices well suited to the biotic/abiotic interface in three areas: shelf-ready sensing, implantable transient electronics, and wearable biosensing. These example devices will illustrate the advantages of silk in this class of bioelectronic and biooptical devices, from fundamentals through application, and contribute to a silk platform for the development of future devices that combine biology with high technology.

Growth factor-functionalized silk membranes support wound healing in vitro.

PubMed

Bienert, M; Hoss, M; Bartneck, M; Weinandy, S; Böbel, M; Jockenhövel, S; Knüchel, R; Pottbacker, K; Wöltje, M; Jahnen-Dechent, W; Neuss, S

2017-08-16

Chronic wounds represent a serious problem in daily medical routine requiring improved wound care. Silk of the domesticated silkworm (Bombyx mori) has been used to form a variety of biomaterials for medical applications. We genetically engineered B. mori to produce silk functionalized with growth factors to promote wound healing in vitro. In this study FGF-, EGF-, KGF-, PDGF- or VEGF-functionalized silk membranes were compared to native B. mori silk membranes without growth factors for their ability to support wound healing in vitro. All silk membranes were cytocompatible and supported macrophage secretion of neutrophil recruiting factor CXCL1 and monocyte chemoattractant protein 1 (MCP-1). VEGF-functionalized silk significantly outperformed other growth factor-functionalized silk membranes, but not native silk in angiogenesis assays. In addition, EGF- and VEGF-functionalized silk membranes slightly enhanced macrophage adhesion compared to silk without growth factors. In wound healing assays in vitro (reduction of wound lesion), dermal equivalents showed a higher wound healing capacity when covered with EGF-, FGF- or VEGF-functionalized silk membranes compared to native, KGF- or PDGF-functionalized silk membranes. Keratinocyte migration and growth is overstimulated by KGF- and VEGF-functionalized silk membranes. In conclusion, growth factor-functionalized silk membranes prepared from genetically engineered silk worm glands are promising wound dressings for future wound healing therapies.

Processing and characterization of silk sericin from Bombyx mori and its application in biomaterials and biomedicines.

PubMed

Cao, Ting-Ting; Zhang, Yu-Qing

2016-04-01

Bombyx mori silk is composed of 60-80% fibroin, 15-35% sericin and 1-5% non-sericin component including wax, pigments, sugars and other impurities. For two decades, the protein-based silk fibroin was extensively used in the research and development of medical biomaterials and biomedicines. Sericin is frequently ignored and abandoned as a byproduct or waste in the processing of traditional silk fabrics, silk floss or modern silk biomaterials. However, similar to fibroin, sericin is not only a highly useful biological material, but also a lot of biological activity. Moreover, the non-sericin component present with sericin in the cocoon shell also has a strong biological activity. In this review, the extraction and recovery methods of sericin and the non-sericin component from the cocoon layer are reported, and their composition, properties and biological activity are described to produce a comprehensive report on biomedical materials and biological drugs. In addition, related problems or concerns present in the research and development of sericin are discussed, and a potential application of sericin in sustainable development is also presented. Copyright © 2015 Elsevier B.V. All rights reserved.

Natural Non-Mulberry Silk Nanoparticles for Potential-Controlled Drug Release

PubMed Central

Wang, Juan; Yin, Zhuping; Xue, Xiang; Kundu, Subhas C.; Mo, Xiumei; Lu, Shenzhou

2016-01-01

Natural silk protein nanoparticles are a promising biomaterial for drug delivery due to their pleiotropic properties, including biocompatibility, high bioavailability, and biodegradability. Chinese oak tasar Antheraea pernyi silk fibroin (ApF) nanoparticles are easily obtained using cations as reagents under mild conditions. The mild conditions are potentially advantageous for the encapsulation of sensitive drugs and therapeutic molecules. In the present study, silk fibroin protein nanoparticles are loaded with differently-charged small-molecule drugs, such as doxorubicin hydrochloride, ibuprofen, and ibuprofen-Na, by simple absorption based on electrostatic interactions. The structure, morphology and biocompatibility of the silk nanoparticles in vitro are investigated. In vitro release of the drugs from the nanoparticles depends on charge-charge interactions between the drugs and the nanoparticles. The release behavior of the compounds from the nanoparticles demonstrates that positively-charged molecules are released in a more prolonged or sustained manner. Cell viability studies with L929 demonstrated that the ApF nanoparticles significantly promoted cell growth. The results suggest that Chinese oak tasar Antheraea pernyi silk fibroin nanoparticles can be used as an alternative matrix for drug carrying and controlled release in diverse biomedical applications. PMID:27916946

Viability and neuronal differentiation of neural stem cells encapsulated in silk fibroin hydrogel functionalized with an IKVAV peptide.

PubMed

Sun, Wei; Incitti, Tania; Migliaresi, Claudio; Quattrone, Alessandro; Casarosa, Simona; Motta, Antonella

2017-05-01

Three-dimensional (3D) porous scaffolds combined with therapeutic stem cells play vital roles in tissue engineering. The adult brain has very limited regeneration ability after injuries such as trauma and stroke. In this study, injectable 3D silk fibroin-based hydrogel scaffolds with encapsulated neural stem cells were developed, aiming at supporting brain regeneration. To improve the function of the hydrogel towards neural stem cells, silk fibroin was modified by an IKVAV peptide through covalent binding. Both unmodified and modified silk fibroin hydrogels were obtained, through sonication, with mechanical stiffness comparable to that of brain tissue. Human neural stem cells were encapsulated in both hydrogels and the effects of IKVAV peptide conjugation on cell viability and neural differentiation were assessed. The silk fibroin hydrogel modified by IKVAV peptide showed increased cell viability and an enhanced neuronal differentiation capability, which contributed to understanding the effects of IKVAV peptide on the behaviour of neural stem cells. For these reasons, IKVAV-modified silk fibroin is a promising material for brain tissue engineering. Copyright © 2015 John Wiley & Sons, Ltd. Copyright © 2015 John Wiley & Sons, Ltd.

Unraveling the Molecular Requirements for Macroscopic Silk Supercontraction.

PubMed

Giesa, Tristan; Schuetz, Roman; Fratzl, Peter; Buehler, Markus J; Masic, Admir

2017-10-24

Spider dragline silk is a protein material that has evolved over millions of years to achieve finely tuned mechanical properties. A less known feature of some dragline silk fibers is that they shrink along the main axis by up to 50% when exposed to high humidity, a phenomenon called supercontraction. This contrasts the typical behavior of many other materials that swell when exposed to humidity. Molecular level details and mechanisms of the supercontraction effect are heavily debated. Here we report a molecular dynamics analysis of supercontraction in Nephila clavipes silk combined with in situ mechanical testing and Raman spectroscopy linking the reorganization of the nanostructure to the polar and charged amino acids in the sequence. We further show in our in silico approach that point mutations of these groups not only suppress the supercontraction effect, but even reverse it, while maintaining the exceptional mechanical properties of the silk material. This work has imminent impact on the design of biomimetic equivalents and recombinant silks for which supercontraction may or may not be a desirable feature. The approach applied is appropriate to explore the effect of point mutations on the overall physical properties of protein based materials.

Genomic perspectives of spider silk genes through target capture sequencing: Conservation of stabilization mechanisms and homology-based structural models of spidroin terminal regions.

PubMed

Collin, Matthew A; Clarke, Thomas H; Ayoub, Nadia A; Hayashi, Cheryl Y

2018-07-01

A powerful system for studying protein aggregation, particularly rapid self-assembly, is spider silk. Spider silks are proteinaceous and silk proteins are synthesized and stored within silk glands as liquid dope. As needed, liquid dope is near-instantaneously transformed into solid fibers or viscous adhesives. The dominant constituents of silks are spidroins (spider fibroins) and their terminal domains are vital for the tight control of silk self-assembly. To better understand spidroin termini, we used target capture and deep sequencing to identify spidroin gene sequences from six species representing the araneoid families of Araneidae, Nephilidae, and Theridiidae. We obtained 145 terminal regions, of which 103 are newly annotated here, as well as novel variants within nine diverse spidroin types. Our comparative analyses demonstrated the conservation of acidic, basic, and cysteine amino acid residues across spidroin types that had been proposed to be important for monomer stability, dimer formation, and self-assembly from a limited sampling of spidroins. Computational, protein homology modeling revealed areas of spidroin terminal regions that are highly conserved in three-dimensions despite sequence divergence across spidroin types. Analyses of our dense sampling of terminal regions suggest that most spidroins share stabilization mechanisms, dimer formation, and tertiary structure, despite producing functionally distinct materials. Copyright © 2018 The Authors. Published by Elsevier B.V. All rights reserved.

Identification and classification of silks using infrared spectroscopy

PubMed Central

Boulet-Audet, Maxime; Vollrath, Fritz; Holland, Chris

2015-01-01

ABSTRACT Lepidopteran silks number in the thousands and display a vast diversity of structures, properties and industrial potential. To map this remarkable biochemical diversity, we present an identification and screening method based on the infrared spectra of native silk feedstock and cocoons. Multivariate analysis of over 1214 infrared spectra obtained from 35 species allowed us to group silks into distinct hierarchies and a classification that agrees well with current phylogenetic data and taxonomies. This approach also provides information on the relative content of sericin, calcium oxalate, phenolic compounds, poly-alanine and poly(alanine-glycine) β-sheets. It emerged that the domesticated mulberry silkmoth Bombyx mori represents an outlier compared with other silkmoth taxa in terms of spectral properties. Interestingly, Epiphora bauhiniae was found to contain the highest amount of β-sheets reported to date for any wild silkmoth. We conclude that our approach provides a new route to determine cocoon chemical composition and in turn a novel, biological as well as material, classification of silks. PMID:26347557

Inducing β-Sheets Formation in Synthetic Spider Silk Fibers by Aqueous Post-Spin Stretching

PubMed Central

Hinman, Michael B.; Holland, Gregory P.; Yarger, Jeffery L.; Lewis, Randolph V.

2012-01-01

As a promising biomaterial with numerous potential applications, various types of synthetic spider silk fibers have been produced and studied in an effort to produce manmade fibers with mechanical and physical properties comparable to those of native spider silk. In this study, two recombinant proteins based on Nephila clavipes Major ampullate Spidroin 1 (MaSp1) consensus repeat sequence were expressed and spun into fibers. Mechanical test results showed that fiber spun from the higher molecular weight protein had better overall mechanical properties (70 KD versus 46 KD), whereas postspin stretch treatment in water helped increase fiber tensile strength significantly. Carbon-13 solid-state NMR studies of those fibers further revealed that the postspin stretch in water promoted protein molecule rearrangement and the formation of β-sheets in the polyalanine region of the silk. The rearrangement correlated with improved fiber mechanical properties and indicated that postspin stretch is key to helping the spider silk proteins in the fiber form correct secondary structures, leading to better quality fibers. PMID:21574576

Modifying the mechanical properties of silk nanofiber scaffold by knitted orientation for regenerative medicine applications.

PubMed

Dodel, M; Hemmati Nejad, N; Bahrami, S H; Soleimani, M; Hanaee-Ahvaz, H

2016-08-31

Tissue reconstruction is among the increasing applications of polymer nanofibers. Fibrous scaffolds (mats) can be easily produced using the electrospinning method with structure and biomechanical properties similar to those of a cellular matrix. Electrospinning is widely used in the production of nanofibers and the GAP-method electrospinning is one of the means of producing fully aligned nanofibers. In this research, using the GAP-method, knitted fibrous scaffolds were made of silk fibroin, which is a biocompatible and biodegradable polymer. To extract fibroin from cocoons, the sodium chloride solution as well as dialysis and freeze-drying techniques were employed. The molecular weight of the extracted fibroin was measured with the SDS-Page electrophoresis technique. Moreover, the pure fibroin structure was examined using the ATR-FTIR method, and the viscosity of the solution used for electrospinning was measured with the Brookfield rotational viscometer. The scaffolds were prepared through electrospinning of the silk fibroin in pure formic acid solution. The following three structures were electrospun: 1) a random structure; 2) a knitted structure with an interstitial angle of 60 degrees; 3) a knitted structure with an interstitial angle of 90 degrees. Morphology of the resulting fibers was studied with a SEM (scanning electron microscope). Fibroin scaffolds are degradable in water. Therefore, they were fixated through immersion in methanol to be prepared for assays. The mechanical properties of the scaffolds were also studied using a tensile strength test device. The effect of methanol on the strength properties of the samples was also assessed. The hydrophilic potential of the samples was measured via a contact angle test. To increase the hydrophilicity of the scaffold surfaces, the cold oxygen plasma technique was employed. Finally, the biocompatibility and cell adhesion of the resulting scaffolds were examined through a HEK 293 cell culture, and the results were analyzed through the MTT, DAPI staining, and SEM imaging techniques. Results revealed that the oriented knitted structure contributed to the increase in Young's modulus and the maximum strength of scaffolds as compared to the random samples. Moreover, this structure can also be a suitable alternative to the typical chemical means of increasing strength.

Nuclear Magnetic Resonance (NMR) Spectroscopic Characterization of Nanomaterials and Biopolymers

NASA Astrophysics Data System (ADS)

Guo, Chengchen

Nanomaterials have attracted considerable attention in recent research due to their wide applications in various fields such as material science, physical science, electrical engineering, and biomedical engineering. Researchers have developed many methods for synthesizing different types of nanostructures and have further applied them in various applications. However, in many cases, a molecular level understanding of nanoparticles and their associated surface chemistry is lacking investigation. Understanding the surface chemistry of nanomaterials is of great significance for obtaining a better understanding of the properties and functions of the nanomaterials. Nuclear magnetic resonance (NMR) spectroscopy can provide a familiar means of looking at the molecular structure of molecules bound to surfaces of nanomaterials as well as a method to determine the size of nanoparticles in solution. Here, a combination of NMR spectroscopic techniques including one- and two-dimensional NMR spectroscopies was used to investigate the surface chemistry and physical properties of some common nanomaterials, including for example, thiol-protected gold nanostructures and biomolecule-capped silica nanoparticles. Silk is a natural protein fiber that features unique properties such as excellent mechanical properties, biocompatibility, and non-linear optical properties. These appealing physical properties originate from the silk structure, and therefore, the structural analysis of silk is of great importance for revealing the mystery of these impressive properties and developing novel silk-based biomaterials as well. Here, solid-state NMR spectroscopy was used to elucidate the secondary structure of silk proteins in N. clavipes spider dragline silk and B. mori silkworm silk. It is found that the Gly-Gly-X (X=Leu, Tyr, Gln) motif in spider dragline silk is not in a beta-sheet or alpha-helix structure and is very likely to be present in a disordered structure with evidence for 31-helix confirmation. In addition, the conformations of the Ala, Ser, and Tyr residues in silk fibroin of B. mori were investigated and it indicates that the Ala, Ser, and Tyr residues are all present in disordered structures in silk I (before spinning), while show different conformations in silk II (after spinning). Specifically, in silk II, the Ala and Tyr residues are present in both disordered structures and beta-sheet structures, and the Ser residues are present primarily in beta-sheet structures.

Silk protein hydrolysate increases glucose uptake through up-regulation of GLUT 4 and reduces the expression of leptin in 3T3-L1 fibroblast.

PubMed

Lee, Hyun-Sun; Lee, Hyun Jung; Suh, Hyung Joo

2011-12-01

The purpose of our research was to test the hypothesis that silk protein hydrolysate increases glucose uptake in cultured murine embryonic fibroblasts. Insulin sensitizing activity was observed in a cell-based glucose uptake assay using 3T3-L1 embryonic fibroblasts. The treatment of 1 mg/mL of silk peptide E5K6 plus 0.2 nM insulin was associated with a significant increase in glucose uptake (124.0% ± 2.5%) compared to treatment with 0.2 nM insulin alone. When the 3T3-L1 cells were induced to differentiate into fibroblasts, fat droplets formed inside the cells. Silk peptide E5K6 reduced the formation of fat droplets at the 1-mg/mL dosage (86.1% ± 2.5%) when compared to the control (100.0% ± 5.8%). A 1 mg/mL dose of silk peptide E5K6 significantly increased GLUT 4 expression (131.5% ± 4.0%). The treatment of 1 mg/mL of silk peptide E5K6 did not present any changes for adipogenic expressed genes, but leptin expression was significantly increased by silk peptide E5K6 supplementation (175.9% ± 11.1%). From these results, silk peptide E5K6 increased glucose uptake via up-regulation of GLUT 4 and decreased fat accumulation via the up-regulation of leptin. Copyright © 2011 Elsevier Inc. All rights reserved.

Transgenic Silk Moths to Produce Spider Silk

DTIC Science & Technology

2008-01-24

concentrated protein solutions, have failed or are inefficient. Currently, silk is produced from the cocoon of the silk moth Bombyx mori ; however, this silk...repetitive domains of spider dragline silk with the N- and C- terminal domains of the Bombyx mori silk gene, Fibroin-H (Fib-H). Various SpF genes have been...transgenic insects and the biosynthetic capacity of the domesticated silkmoth, Bombyx mori . The elasticity and strength of spider silk make it ideal for

DOE Office of Scientific and Technical Information (OSTI.GOV)

Bradshaw, Nathan P.; Severt, Sean Y.; Wang, Zhaoying

Biocompatible materials capable of controlled actuation under biologically relevant conditions are in high demand for use in a number of biomedical applications. Recently, we demonstrated that a composite material composed of silk biopolymer and the conducting polymer poly(pyrrole) can bend under an applied voltage using a simple bilayer device. Here we present further characterization of these bilayer actuators using time of flight secondary ion mass spectrometry, and provide clarification on the mechanism of actuation and factors affecting device performance and stability. We will discuss the results of this study in the context of strategies for optimization of device performance.

The Structure and Properties of Parachute Cloths

NASA Technical Reports Server (NTRS)

Mcnicholas, H J; Hedrick, F

1930-01-01

The requisite properties of a parachute cloth are discussed and the methods for measuring these properties described. In addition to the structural analysis of the cloths, the properties measured were weight, breaking strength, tear resistance, elasticity, and air permeability. Thirty-six silk cloths of domestic manufacture, not previously used in parachute construction are compared with some silk cloths of foreign manufacture. These foreign cloths were ones proven by trial and extended use to be suitable materials for parachute construction. Contrary to the belief that domestic woven cloths were not suitable materials for parachute construction, it is shown that many domestic silk cloths are satisfactory and in some respects superior to the foreign products. Based on a comparative study of all the cloths, specifications are drawn for the manufacture of silk parachute cloth.

Manufacture and Drug Delivery Applications of Silk Nanoparticles.

PubMed

Wongpinyochit, Thidarat; Johnston, Blair F; Seib, F Philipp

2016-10-08

Silk is a promising biopolymer for biomedical and pharmaceutical applications due to its outstanding mechanical properties, biocompatibility and biodegradability, as well its ability to protect and subsequently release its payload in response to a trigger. While silk can be formulated into various material formats, silk nanoparticles are emerging as promising drug delivery systems. Therefore, this article covers the procedures for reverse engineering silk cocoons to yield a regenerated silk solution that can be used to generate stable silk nanoparticles. These nanoparticles are subsequently characterized, drug loaded and explored as a potential anticancer drug delivery system. Briefly, silk cocoons are reverse engineered first by degumming the cocoons, followed by silk dissolution and clean up, to yield an aqueous silk solution. Next, the regenerated silk solution is subjected to nanoprecipitation to yield silk nanoparticles - a simple but powerful method that generates uniform nanoparticles. The silk nanoparticles are characterized according to their size, zeta potential, morphology and stability in aqueous media, as well as their ability to entrap a chemotherapeutic payload and kill human breast cancer cells. Overall, the described methodology yields uniform silk nanoparticles that can be readily explored for a myriad of applications, including their use as a potential nanomedicine.

Effect of Silk Protein Processing on Drug Delivery from Silk Films

PubMed Central

Pritchard, Eleanor M.; Hu, Xiao; Finley, Violet; Kuo, Catherine K.; Kaplan, David L.

2013-01-01

Sericin removal from the core fibroin protein of silkworm silk is a critical first step in the use of silk for biomaterial-related applications, but degumming can affect silk biomaterial properties, including molecular weight, viscosity, diffusivity and degradation behavior. Increasing the degumming time (10, 30, 60 and 90 min) decreases the average molecular weight of silk protein in solution, silk solution viscosity, and silk film glass transition temperature, and increases the rate of degradation of silk film by protease. Model compounds spanning a range of physical-chemical properties generally showed an inverse relationship between degumming time and release rate through a varied degumming time silk coating. Degumming provides a useful control point to manipulate silk’s material properties. PMID:23349062

New concept of 3D printed bone clip (polylactic acid/hydroxyapatite/silk composite) for internal fixation of bone fractures.

PubMed

Yeon, Yeung Kyu; Park, Hae Sang; Lee, Jung Min; Lee, Ji Seung; Lee, Young Jin; Sultan, Md Tipu; Seo, Ye Bin; Lee, Ok Joo; Kim, Soon Hee; Park, Chan Hum

Open reduction with internal fixation is commonly used for the treatment of bone fractures. However, postoperative infection associated with internal fixation devices (intramedullary nails, plates, and screws) remains a significant complication, and it is technically difficult to fix multiple fragmented bony fractures using internal fixation devices. In addition, drilling in the bone to install devices can lead to secondary fracture, bone necrosis associated with postoperative infection. In this study, we developed bone clip type internal fixation device using three- dimensional (3D) printing technology. Standard 3D model of the bone clip was generated based on computed tomography (CT) scan of the femur in the rat. Polylacticacid (PLA), hydroxyapatite (HA), and silk were used for bone clip material. The purpose of this study was to characterize 3D printed PLA, PLA/HA, and PLA/HA/Silk composite bone clip and evaluate the feasibility of these bone clips as an internal fixation device. Based on the results, PLA/HA/Silk composite bone clip showed similar mechanical property, and superior biocompatibility compared to other types of the bone clip. PLA/HA/Silk composite bone clip demonstrated excellent alignment of the bony segments across the femur fracture site with well-positioned bone clip in an animal study. Our 3D printed bone clips have several advantages: (1) relatively noninvasive (drilling in the bone is not necessary), (2) patient-specific design (3) mechanically stable device, and (4) it provides high biocompatibility. Therefore, we suggest that our 3D printed PLA/HA/Silk composite bone clip is a possible internal fixation device.

Modifying the Mechanical Properties of Silk Fiber by Genetically Disrupting the Ionic Environment for Silk Formation.

PubMed

Wang, Xin; Zhao, Ping; Li, Yi; Yi, Qiying; Ma, Sanyuan; Xie, Kang; Chen, Huifang; Xia, Qingyou

2015-10-12

Silks are widely used biomaterials, but there are still weaknesses in their mechanical properties. Here we report a method for improving the silk fiber mechanical properties by genetic disruption of the ionic environment for silk fiber formation. An anterior silk gland (ASG) specific promoter was identified and used for overexpressing ion-transporting protein in the ASG of silkworm. After isolation of the transgenic silkworms, we found that the metal ion content, conformation and mechanical properties of transgenic silk fibers changed accordingly. Notably, overexpressing endoplasmic reticulum Ca2+-ATPase in ASG decreased the calcium content of silks. As a consequence, silk fibers had more α-helix and β-sheet conformations, and their tenacity and extension increased significantly. These findings represent the in vivo demonstration of a correlation between metal ion content in the spinning duct and the mechanical properties of silk fibers, thus providing a novel method for modifying silk fiber properties.

In vivo effects of metal ions on conformation and mechanical performance of silkworm silks.

PubMed

Wang, Xin; Li, Yi; Liu, Qingsong; Chen, Quanmei; Xia, Qingyou; Zhao, Ping

2017-03-01

The mechanism of silk fiber formation is of particular interest. Although in vitro evidence has shown that metal ions affect conformational transitions of silks, the in vivo effects of metal ions on silk conformations and mechanical performance are still unclear. This study explored the effects of metal ions on silk conformations and mechanical properties of silk fibers by adding K + and Cu 2+ into the silk fibroin solutions or injecting them into the silkworms. Aimed by CD analysis, FTIR analysis, and mechanical testing, the conformational and mechanical changes of the silks were estimated. By using BION Web Server, the interactions of K + and N-terminal of silk fibroin were also simulated. We presented that K + and Cu 2+ induced the conformational transitions of silk fibroin by forming β-sheet structures. Moreover, the mechanical parameters of silk fibers, such as strength, toughness and Young's modulus, were also improved after K + or Cu 2+ injection. Using BION Web Server, we found that potassium ions may have strong electrostatic interactions with the negatively charged residues. We suggest that K + and Cu 2+ play crucial roles in the conformation and mechanical performances of silks and they are involved in the silk fiber formation in vivo. Our results are helpful for clarifying the mechanism of silk fiber formation, and provide insights for modifying the mechanical properties of silk fibers. Copyright © 2016 Elsevier B.V. All rights reserved.

Osteoinductive-nanoscaled silk/HA composite scaffolds for bone tissue engineering application.

PubMed

Huang, Xiaowei; Bai, Shumeng; Lu, Qiang; Liu, Xi; Liu, Shanshan; Zhu, Hesun

2015-10-01

Osteoinductive silk/hydroxyapatite (HA) composite scaffolds for bone regeneration were prepared by combining silk with HA/silk core-shell nanoparticles. The HA/silk nanoparticles were directly dispersed in silk solution to form uniform silk/HA blend and then composite scaffolds after a freeze-drying process. The HA/silk nanoparticles uniformly distributed in silk scaffolds at nanometer scale at varying HA content up to 40%, and substantially improved the compressive strength of the scaffolds produced. Rat bone mesenchymal stem cells (rBMSCs) were cultured in these scaffolds and cell proliferation was analyzed by confocal microscopy and DNA assay. Gene expression and biochemical assays were employed to study the influence of increasing HA/silk nanoparticles on in vitro osteogenic differentiation of rBMSCs. Increasing HA/silk nanoparticles inside silk scaffolds improved the growth and osteogenic capability of rBMSCs in the absence of osteogenic growth factors, and also significantly increased the calcium and collagen I deposition. In addition, compared to silk/HA composite scaffolds containing HA aggregates, the scaffolds loaded with HA/silk nanoparticles showed remarkably higher stiffness and better osteogenic property at same HA content, implying a preferable microenvironment for rBMSCs. These results suggest that the osteogenic property as well as mechanical property of silk/HA scaffolds could be further improved through fabricating their structure and topography at nanometer scale, providing more suitable systems for bone regeneration. © 2014 Wiley Periodicals, Inc.

Conservation of a pH-sensitive structure in the C-terminal region of spider silk extends across the entire silk gene family.

PubMed

Strickland, Michelle; Tudorica, Victor; Řezáč, Milan; Thomas, Neil R; Goodacre, Sara L

2018-06-01

Spiders produce multiple silks with different physical properties that allow them to occupy a diverse range of ecological niches, including the underwater environment. Despite this functional diversity, past molecular analyses show a high degree of amino acid sequence similarity between C-terminal regions of silk genes that appear to be independent of the physical properties of the resulting silks; instead, this domain is crucial to the formation of silk fibers. Here, we present an analysis of the C-terminal domain of all known types of spider silk and include silk sequences from the spider Argyroneta aquatica, which spins the majority of its silk underwater. Our work indicates that spiders have retained a highly conserved mechanism of silk assembly, despite the extraordinary diversification of species, silk types and applications of silk over 350 million years. Sequence analysis of the silk C-terminal domain across the entire gene family shows the conservation of two uncommon amino acids that are implicated in the formation of a salt bridge, a functional bond essential to protein assembly. This conservation extends to the novel sequences isolated from A. aquatica. This finding is relevant to research regarding the artificial synthesis of spider silk, suggesting that synthesis of all silk types will be possible using a single process.

Rheology and dynamic light scattering of silk fibroin solution extracted from the middle division of Bombyx mori silkworm.

PubMed

Ochi, Akie; Hossain, Khandker S; Magoshi, Jun; Nemoto, Norio

2002-01-01

Dynamic light scattering (DLS) and rheological measurements were performed on aqueous silk fibroin solutions extracted from the middle division of Bombyx mori silkworm over a wide range of polymer concentration C from 0.08 to 27.5 wt %. DLS results obtained in the dilute region of C less than 1 wt % are consistent with a model that an elementary unit is a large protein complex consisting of silk fibroin and P25 with a 6:1 molar ratio. Rheological measurements in the dilute C region reveal that those units (or clusters) with the hydrodynamic radius of about 100 nm form a network extending over the whole sample volume with small pseudoplateau modulus mainly by ionic bonding between COO(-) ions of the fibroin molecules and divalent metallic ions such as Ca(2+) or Mg(2+) ions present in the sample and also that, after a yield stress is reached, steady plastic flow is induced with viscosity much lower than the zero-shear viscosity estimated from creep and creep recovery measurements by 4-6 orders of magnitude. Angular frequency omega dependencies of the storage and the loss shear moduli, G'(omega) and G' '(omega), measured in the linear viscoelastic region, indicate that all solutions possess the pseudoplateau modulus in the low omega region and samples become highly viscoleastic for C greater, similar 4.2 wt %. Above C = 11.2 wt % another plateau appears at the high omega end accompanied by a distinct maximum of G' ' in the intermediate omega region. The relaxation motion with tau = 0.5 s corresponding to the maximum of G' ' is one of characteristic properties of the fibroin solutions in the high C region. Thermorheological behaviors of the solution with C = 27.5 wt % show that the network structure formed in the MM part of the silk gland is susceptible to temperature and a more stable homogeneous network is realized by raising the temperature up to T = 65 degrees C.

Atomic force microscopy of orb-spider-web-silks to measure surface nanostructuring and evaluate silk fibers per strand

NASA Astrophysics Data System (ADS)

Kane, D. M.; Naidoo, N.; Staib, G. R.

2010-10-01

Atomic force microscopy (AFM) study is used to measure the surface topology and roughness of radial and capture spider silks on the micro- and nanoscale. This is done for silks of the orb weaver spider Argiope keyserlingi. Capture silk has a surface roughness that is five times less than that for radial silk. The capture silk has an equivalent flatness of λ /100 (5-6 nm deep surface features) as an optical surface. This is equivalent to a very highly polished optical surface. AFM does show the number of silk fibers that make up a silk thread but geometric distortion occurs during sample preparation. This prevented AFM from accurately measuring the silk topology on the microscale in this study.

'Silk Road', the virtual drug marketplace: a single case study of user experiences.

PubMed

Van Hout, Marie Claire; Bingham, Tim

2013-09-01

The online promotion of 'drug shopping' and user information networks is of increasing public health and law enforcement concern. An online drug marketplace called 'Silk Road' has been operating on the 'Deep Web' since February 2011 and was designed to revolutionise contemporary drug consumerism. A single case study approach explored a 'Silk Road' user's motives for online drug purchasing, experiences of accessing and using the website, drug information sourcing, decision making and purchasing, outcomes and settings for use, and perspectives around security. The participant was recruited following a lengthy relationship building phase on the 'Silk Road' chat forum. The male participant described his motives, experiences of purchasing processes and drugs used from 'Silk Road'. Consumer experiences on 'Silk Road' were described as 'euphoric' due to the wide choice of drugs available, relatively easy once navigating the Tor Browser (encryption software) and using 'Bitcoins' for transactions, and perceived as safer than negotiating illicit drug markets. Online researching of drug outcomes, particularly for new psychoactive substances was reported. Relationships between vendors and consumers were described as based on cyber levels of trust and professionalism, and supported by 'stealth modes', user feedback and resolution modes. The reality of his drug use was described as covert and solitary with psychonautic characteristics, which contrasted with his membership, participation and feelings of safety within the 'Silk Road' community. 'Silk Road' as online drug marketplace presents an interesting displacement away from 'traditional' online and street sources of drug supply. Member support and harm reduction ethos within this virtual community maximises consumer decision-making and positive drug experiences, and minimises potential harms and consumer perceived risks. Future research is necessary to explore experiences and backgrounds of other users. Copyright © 2013 Elsevier B.V. All rights reserved.

Biomimetic Nucleation of Hydroxyapatite Crystals Mediated by Antheraea pernyi Silk Sericin Promotes Osteogenic Differentiation of Human Bone Marrow Derived Mesenchymal Stem Cells

PubMed Central

2015-01-01

Biomacromolecules have been used as templates to grow hydroxyapatite crystals (HAps) by biomineralization to fabricate mineralized materials for potential application in bone tissue engineering. Silk sericin is a protein with features desirable as a biomaterial, such as increased hydrophilicity and biodegradation. Mineralization of the silk sericin from Antheraea pernyi (A. pernyi) silkworm has rarely been reported. Here, for the first time, nucleation of HAps on A. pernyi silk sericin (AS) was attempted through a wet precipitation method and consequently the cell viability and osteogenic differentiation of BMSCs on mineralized AS were investigated. It was found that AS mediated the nucleation of HAps in the form of nanoneedles while self-assembling into β-sheet conformation, leading to the formation of a biomineralized protein based biomaterial. The cell viability assay of BMSCs showed that the mineralization of AS stimulated cell adhesion and proliferation, showing that the resultant AS biomaterial is biocompatible. The differentiation assay confirmed that the mineralized AS significantly promoted the osteogenic differentiation of BMSCs when compared to nonmineralized AS as well as other types of sericin (B. mori sericin), suggesting that the resultant mineralized AS biomaterial has potential in promoting bone formation. This result represented the first work proving the osteogenic differentiation of BMSCs directed by silk sericin. Therefore, the biomineralization of A. pernyi silk sericin coupled with seeding BMSCs on the resultant mineralized biomaterials is a useful strategy to develop the potential application of this unexplored silk sericin in the field of bone tissue engineering. This study lays the foundation for the use of A. pernyi silk sericin as a potential scaffold for tissue engineering. PMID:24666022

Biomimetic nucleation of hydroxyapatite crystals mediated by Antheraea pernyi silk sericin promotes osteogenic differentiation of human bone marrow derived mesenchymal stem cells.

PubMed

Yang, Mingying; Shuai, Yajun; Zhang, Can; Chen, Yuyin; Zhu, Liangjun; Mao, Chuanbin; OuYang, Hongwei

2014-04-14

Biomacromolecules have been used as templates to grow hydroxyapatite crystals (HAps) by biomineralization to fabricate mineralized materials for potential application in bone tissue engineering. Silk sericin is a protein with features desirable as a biomaterial, such as increased hydrophilicity and biodegradation. Mineralization of the silk sericin from Antheraea pernyi (A. pernyi) silkworm has rarely been reported. Here, for the first time, nucleation of HAps on A. pernyi silk sericin (AS) was attempted through a wet precipitation method and consequently the cell viability and osteogenic differentiation of BMSCs on mineralized AS were investigated. It was found that AS mediated the nucleation of HAps in the form of nanoneedles while self-assembling into β-sheet conformation, leading to the formation of a biomineralized protein based biomaterial. The cell viability assay of BMSCs showed that the mineralization of AS stimulated cell adhesion and proliferation, showing that the resultant AS biomaterial is biocompatible. The differentiation assay confirmed that the mineralized AS significantly promoted the osteogenic differentiation of BMSCs when compared to nonmineralized AS as well as other types of sericin (B. mori sericin), suggesting that the resultant mineralized AS biomaterial has potential in promoting bone formation. This result represented the first work proving the osteogenic differentiation of BMSCs directed by silk sericin. Therefore, the biomineralization of A. pernyi silk sericin coupled with seeding BMSCs on the resultant mineralized biomaterials is a useful strategy to develop the potential application of this unexplored silk sericin in the field of bone tissue engineering. This study lays the foundation for the use of A. pernyi silk sericin as a potential scaffold for tissue engineering.

Molecular mechanics of silk nanostructures under varied mechanical loading.

PubMed

Bratzel, Graham; Buehler, Markus J

2012-06-01

Spider dragline silk is a self-assembling tunable protein composite fiber that rivals many engineering fibers in tensile strength, extensibility, and toughness, making it one of the most versatile biocompatible materials and most inviting for synthetic mimicry. While experimental studies have shown that the peptide sequence and molecular structure of silk have a direct influence on the stiffness, toughness, and failure strength of silk, few molecular-level analyses of the nanostructure of silk assemblies, in particular, under variations of genetic sequences have been reported. In this study, atomistic-level structures of wildtype as well as modified MaSp1 protein from the Nephila clavipes spider dragline silk sequences, obtained using an in silico approach based on replica exchange molecular dynamics and explicit water molecular dynamics, are subjected to simulated nanomechanical testing using different force-control loading conditions including stretch, pull-out, and peel. The authors have explored the effects of the poly-alanine length of the N. clavipes MaSp1 peptide sequence and identify differences in nanomechanical loading conditions on the behavior of a unit cell of 15 strands with 840-990 total residues used to represent a cross-linking β-sheet crystal node in the network within a fibril of the dragline silk thread. The specific loading condition used, representing concepts derived from the protein network connectivity at larger scales, have a significant effect on the mechanical behavior. Our analysis incorporates stretching, pull-out, and peel testing to connect biochemical features to mechanical behavior. The method used in this study could find broad applications in de novo design of silk-like tunable materials for an array of applications. Copyright © 2011 Wiley Periodicals, Inc.

'Surfing the Silk Road': a study of users' experiences.

PubMed

Van Hout, Marie Claire; Bingham, Tim

2013-11-01

The online drug marketplace called 'Silk Road' has operated anonymously on the 'Deep Web' since 2011. It is accessible through computer encrypting software (Tor) and is supported by online transactions using peer to peer anonymous and untraceable crypto-currency (Bit Coins). The study aimed to describe user motives and realities of accessing, navigating and purchasing on the 'Silk Road' marketplace. Systematic online observations, monitoring of discussion threads on the site during four months of fieldwork and analysis of anonymous online interviews (n=20) with a convenience sample of adult 'Silk Road' users was conducted. The majority of participants were male, in professional employment or in tertiary education. Drug trajectories ranged from 18 months to 25 years, with favourite drugs including MDMA, 2C-B, mephedrone, nitrous oxide, ketamine, cannabis and cocaine. Few reported prior experience of online drug sourcing. Reasons for utilizing 'Silk Road' included curiosity, concerns for street drug quality and personal safety, variety of products, anonymous transactioning, and ease of product delivery. Vendor selection appeared to be based on trust, speed of transaction, stealth modes and quality of product. Forums on the site provided user advice, trip reports, product and transaction reviews. Some users reported solitary drug use for psychonautic and introspective purposes. A minority reported customs seizures, and in general a displacement away from traditional drug sourcing (street and closed markets) was described. Several reported intentions to commence vending on the site. The study provides an insight into 'Silk Road' purchasing motives and processes, the interplay between traditional and 'Silk Road' drug markets, the 'Silk Road' online community and its communication networks. Copyright © 2013 Elsevier B.V. All rights reserved.

Multifunctional surface modification of silk fabric via graphene oxide repeatedly coating and chemical reduction method

NASA Astrophysics Data System (ADS)

Cao, Jiliang; Wang, Chaoxia

2017-05-01

Multifunctional silk fabrics with electrical conductive, anti-ultraviolet and water repellent were successfully prepared by surface modification with graphene oxide (GO). The yellow-brown GO deposited on the surface of silk fabric was converted into graphitic black reduced graphene (RGO) by sodium hydrosulfite. The surface properties of silk fabrics were changed by repeatedly RGO coating process, which have been proved by SEM and XPS. The SEM results showed that the RGO sheets were successive form a continuously thin film on the surface of silk fabrics, and the deposition of GO or RGO also can be proved by XPS. The electrical conductivity was tested by electrical surface resistance value of the silk fabric, the surface resistance decreased with increasing of RGO surface modification times, and a low surface resistance value reached to 3.24 KΩ cm-1 after 9 times of modification, indicating the silk obtained excellent conductivity. The UPF value of one time GO modification silk fabric (silk-1RGO) was enhanced significantly to 24.45 in comparison to 10.40 of original silk. The contact angle of RGO coating silk samples was all above of 120°. The durability of RGO coated silk fabrics was tested by laundering. The electrical surface resistance of silk-4RGO (65.74 KΩ cm-1), silk-6RGO (15.54 KΩ cm-1) and silk-8RGO (3.86 KΩ cm-1) fabrics was up to 86.82, 22.30 and 6.57 KΩ cm-1 after 10 times of standard washing, respectively. The UPF value, contact angle and color differences of RGO modified silk fabric slightly changed before and after 10 times of standard washing. Therefore, the washing fastness of electric conduction, anti-ultraviolet and water repellent multifunctional silk fabrics was excellent.

Osiris9a is a major component of silk fiber in lepidopteran insects.

PubMed

Liu, Chun; Hu, Wenbo; Cheng, Tingcai; Peng, Zhangchuan; Mita, Kazuei; Xia, Qingyou

2017-10-01

In a previous high-throughput proteomics study, it was found that the silkworm cocoon contains hundreds of complex proteins, many of which have unknown functions, in addition to fibroins, sericins, and some protease inhibitors. Osiris was one of the proteins with no known function. In this study, we identified the Osiris gene family members and constructed a phylogenetic tree based on the sequences from different species. Our results indicate that the Osiris9 gene subfamily contains six members; it is specifically expressed in lepidopteran insects and has evolved by gene duplication. An Osiris gene family member from Bombyx mori was designated as BmOsiris9a (BmOsi9a) on the basis of its homology to Drosophila melanogaster Osiris9. The expression pattern of BmOsi9a showed that it was highly expressed only in the middle silk gland of silkworm larvae, similar to Sericin1 (Ser1). BmOsi9a was visualized as two bands in western blot analysis, implying that it probably undergoes post-translational modifications. Immunohistochemistry analysis revealed that BmOsi9a was synthesized and secreted into the lumen of the middle silk gland, and was localized in the sericin layer in the silk fiber. BmOsi9a was found in the silk fibers of not only three Bombycidae species, viz. B. mori, B. mandarina, and B. huttoni, but also in the fibers collected from Saturniidae species, including Antheraea assama, Antheraea mylitta, and Samia cynthia. Although the exact biological function of Osi9a in the silk fibers is unknown, our results are important because they demonstrate that Osi9a is a common structural component of silk fiber and is expressed widely among the silk-producing Bombycidae and Saturniidae insects. Our results should help in understanding the role of Osi9a in silk fibers. Copyright © 2017 Elsevier Ltd. All rights reserved.

Phosphoproteomic analysis of the posterior silk gland of Bombyx mori provides novel insight into phosphorylation regulating the silk production.

PubMed

Song, Jia; Che, Jiaqian; You, Zhengying; Ye, Lupeng; Li, Jisheng; Zhang, Yuyu; Qian, Qiujie; Zhong, Boxiong

2016-10-04

To understand phosphorylation event regulating silk synthesis in the posterior silk gland of Bombyx mori, phosphoproteome was profiled in a pair of near-isogenic lines, a normally cocooning strain (IC) and a nakedly pupated strain (IN) that the silk production is much lower than IC. In the posterior silk gland of the IC and IN, 714 and 658 phosphosites resided on 554 and 507 phosphopeptides from 431 and 383 phosphoproteins, were identified, respectively. Of all the phosphosites, the single phosphosite was the dominate phosphorylation form, comprising>60% of all the phosphosites in two phenotypic of silk production. All these phosphosites were classified as acidophilic and proline-directed kinase classes, and three motifs were uniquely identified in the IC. The motif S-P-P might be important for regulating phosphorylation network of silk protein synthesis. The dynamically phosphorylated proteins participated in ribosome, protein transport and energy metabolism suggest that phosphorylation may play key roles in regulating silk protein synthesis and secretion. Furthermore, fibroin heavy chain, an important component of silk protein, was specifically phosphorylated in the IC strain, suggesting its role to ensure the normal formation of silk structure and silk secretion. The data gain new understanding of the regulatory processes of silk protein synthesis and offer as starting point for further research on the silk production at phosphoproteome level. Despite the knowledge on regulation of silk protein synthesis in the posterior silk gland has gained at the gene or protein levels, how phosphorylation event influences the silk yield is largely unknown. To this end, we constructed a pair of silkworm near-isogenic lines that showed different cocooning phenotypes, and the phosphoproteome of the posterior silk gland of two isolines was compared. Here, we reported the first phosphoproteome data on the silkworm and found several key pathways related protein synthesis are regulated by phosphorylation, thereby influencing the silk production. The data provide valuable resources for further functional assay of targeted protein phosphorylation that regulates the silk synthesis in silkworm. Copyright © 2016 Elsevier B.V. All rights reserved.

Optically probing torsional superelasticity in spider silks

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kumar, Bhupesh; Thakur, Ashish; Panda, Biswajit

2013-11-11

We investigate torsion mechanics of various spider silks using a sensitive optical technique. We find that spider silks are torsionally superelastic in that they can reversibly withstand great torsion strains of over 10{sup 2−3} rotations per cm before failure. Among various silks from a spider, we find the failure twist-strain is greatest in the sticky capture silk followed by dragline and egg-case silk. Our in situ laser-diffraction measurements reveal that torsional strains on the silks induce a nano-scale transverse compression in its diameter that is linear and reversible. These unique torsional properties of the silks could find applications in silk-basedmore » materials and devices.« less

Subchronic toxicity study of corn silk with rats.

PubMed

Wang, Cuina; Zhang, Tiehua; Liu, Jun; Lu, Shuang; Zhang, Cheng; Wang, Erlei; Wang, Zuozhao; Zhang, Yan; Liu, Jingbo

2011-09-01

Corn silk is a traditional herbal medicine in China, which has been used in many parts of the world for the treatment of edema as well as for cystitis, gout, kidney stones, nephritis, prostatitis and similar ailments. However, there is little scientific evidence about its safety. As a part of its safety assessment, a subchronic toxicity was performed in this paper. The subchronic toxicity was investigated in male and female Wistar rats by dietary administration at concentrations of 0.5%, 2.0% and 8.0% (w/w) for 90 days. Overall health, body weight, food consumption, hematology, blood chemistry, organ weights, gross and microscopic appearance of tissues were compared between test and control groups. A number of significant differences were seen between groups, but none of them was considered to be adverse. Based on the present study, the no-observed-adverse-effect level (NOAEL) of corn silk is at least 8.0% which corresponds to a mean daily corn silk intake of approximately 9.354 and 10.308 g/day/kg body weight for males and females, respectively. The results obtained in the present study suggest that consumption of corn silk has no adverse effects and support the safety of corn silk for humans. Copyright © 2011 Elsevier Ireland Ltd. All rights reserved.

Silk-ionomer and silk-tropoelastin hydrogels as charged three-dimensional culture platforms for the regulation of hMSC response.

PubMed

Calabrese, Rossella; Raia, Nicole; Huang, Wenwen; Ghezzi, Chiara E; Simon, Marc; Staii, Cristian; Weiss, Anthony S; Kaplan, David L

2017-09-01

The response of human bone marrow-derived mesenchymal stem cells (hMSCs) encapsulated in three-dimensional (3D) charged protein hydrogels was studied. Combining silk fibroin (S) with recombinant human tropoelastin (E) or silk ionomers (I) provided protein composite alloys with tunable physicochemical and biological features for regulating the bioactivity of encapsulated hMSCs. The effects of the biomaterial charges on hMSC viability, proliferation and chondrogenic or osteogenic differentiation were assessed. The silk-tropoelastin or silk-ionomers hydrogels supported hMSC viability, proliferation and differentiation. Gene expression of markers for chondrogenesis and osteogenesis, as well as biochemical and histological analysis, showed that hydrogels with different S/E and S/I ratios had different effects on cell fate. The negatively charged hydrogels upregulated hMSC chondrogenesis or osteogenesis, with or without specific differentiation media, and hydrogels with higher tropoelastin content inhibited the differentiation potential even in the presence of the differentiation media. The results provide insight on charge-tunable features of protein-based biomaterials to control hMSC differentiation in 3D hydrogels, as well as providing a new set of hydrogels for the compatible encapsulation and utility for cell functions. Copyright © 2016 John Wiley & Sons, Ltd. Copyright © 2016 John Wiley & Sons, Ltd.

3D Printing of Hierarchical Silk Fibroin Structures.

PubMed

Sommer, Marianne R; Schaffner, Manuel; Carnelli, Davide; Studart, André R

2016-12-21

Like many other natural materials, silk is hierarchically structured from the amino acid level up to the cocoon or spider web macroscopic structures. Despite being used industrially in a number of applications, hierarchically structured silk fibroin objects with a similar degree of architectural control as in natural structures have not been produced yet due to limitations in fabrication processes. In a combined top-down and bottom-up approach, we exploit the freedom in macroscopic design offered by 3D printing and the template-guided assembly of ink building blocks at the meso- and nanolevel to fabricate hierarchical silk porous materials with unprecedented structural control. Pores with tunable sizes in the range 40-350 μm are generated by adding sacrificial organic microparticles as templates to a silk fibroin-based ink. Commercially available wax particles or monodisperse polycaprolactone made by microfluidics can be used as microparticle templates. Since closed pores are generated after template removal, an ultrasonication treatment can optionally be used to achieve open porosity. Such pore templating particles can be further modified with nanoparticles to create a hierarchical template that results in porous structures with a defined nanotopography on the pore walls. The hierarchically porous silk structures obtained with this processing technique can potentially be utilized in various application fields from structural materials to thermal insulation to tissue engineering scaffolds.

Glycerol-plasticised silk membranes made using formic acid are ductile, transparent and degradation-resistant.

PubMed

Allardyce, Benjamin J; Rajkhowa, Rangam; Dilley, Rodney J; Redmond, Sharon L; Atlas, Marcus D; Wang, Xungai

2017-11-01

Regenerated silk fibroin membranes tend to be brittle when dry. The use of plasticisers such as glycerol improve membrane ductility, but, when combined with aqueous processing, can lead to a higher degradation rate than solvent-annealed membranes. This study investigated the use of formic acid as the solvent with glycerol to make deformable yet degradation-resistant silk membranes. Here we show that membranes cast using formic acid had low light scattering, with a diffuse transmittance of less than 5% over the visible wavelengths, significantly lower than the 20% transmittance of aqueous derived silk/glycerol membranes. They had 64% β-sheet content and lost just 30% of the initial silk weight over 6h when tested with an accelerated enzymatic degradation assay, in comparison the aqueous membranes completely degraded within this timeframe. The addition of glycerol also improved the maximum elongation of formic acid derived membranes from under 3% to over 100%. They also showed good cytocompatibility and supported the adhesion and migration of human tympanic membrane keratinocytes. Formic acid based, silk/glycerol membranes may be of great use in medical applications such as repair of tympanic membrane perforation or ocular applications where transparency and resistance to enzymatic degradation are important. Copyright © 2017 Elsevier B.V. All rights reserved.

Design and Optimization of Resorbable Silk Internal Fixation Devices

NASA Astrophysics Data System (ADS)

Haas, Dylan S.

Limitations of current material options for internal fracture fixation devices have resulted in a large gap between user needs and hardware function. Metal systems offer robust mechanical strength and ease of implantation but require secondary surgery for removal and/or result in long-term complications (infection, palpability, sensitivity, etc.). Current resorbable devices eliminate the need for second surgery and long-term complications but are still associated with negative host response as well as limited functionality and more difficult implantation. There is a definitive need for orthopedic hardware that is mechanically capable of immediate fracture stabilization and fracture fixation during healing, can safely biodegrade while allowing complete bone remodeling, can be resterilized for reuse, and is easily implantable (self-tapping). Previous work investigated the use of silk protein to produce resorbable orthopedic hardware for non- load bearing fracture fixation. In this study, silk orthopedic hardware was further investigated and optimized in order to better understand the ability of silk as a fracture fixation system and more closely meet the unfulfilled market needs. Solvent-based and aqueous-based silk processing formulations were cross-linked with methanol to induce beta sheet structure, dried, autoclaved and then machined to the desired device/geometry. Silk hardware was evaluated for dry, hydrated and fatigued (cyclic) mechanical properties, in vitro degradation, resterilization, functionalization with osteoinductive molecules and implantation technique for fracture fixation. Mechanical strength showed minor improvements from previous results, but remains comparable to current resorbable fixation systems with the advantages of self-tapping ability for ease of implantation, full degradation in 10 months, ability to be resterilized and reused, and ability to release molecules for osteoinudction. In vivo assessment confirmed biocompatibility, showed improved bone deposition and remodeling with functionalization and showed promising feasibility of fracture fixations with minor adjustments to geometry. The proposed silk orthopedic hardware exhibits high potential as a resorbable fixation system that can bridge the gap between the current materials for internal fixation devices.

Silkworms transformed with chimeric silkworm/spider silk genes spin composite silk fibers with improved mechanical properties

USDA-ARS?s Scientific Manuscript database

The development of a spider silk manufacturing process is of great interest. piggyBac vectors were used to create transgenic silkworms encoding chimeric silkworm/spider silk proteins. The silk fibers produced by these animals were composite materials that included chimeric silkworm/spider silk prote...

Mechanical features of various silkworm crystalline considering hydration effect via molecular dynamics simulations.

PubMed

Kim, Yoonjung; Lee, Myeongsang; Choi, Hyunsung; Baek, Inchul; Kim, Jae In; Na, Sungsoo

2018-04-01

Silk materials are receiving significant attention as base materials for various functional nanomaterials and nanodevices, due to its exceptionally high mechanical properties, biocompatibility, and degradable characteristics. Although crystalline silk regions are composed of various repetitive motifs with differing amino acid sequences, how the effect of humidity works differently on each of the motifs and their structural characteristics remains unclear. We report molecular dynamics (MD) simulations on various silkworm fibroins composed of major motifs (i.e. (GAGAGS) n , (GAGAGA) n , and (GAGAGY) n ) at varying degrees of hydration, and reveal how each major motifs of silk fibroins change at each degrees of hydration using MD simulations and their structural properties in mechanical perspective via steered molecular dynamics simulations. Our results explain what effects humidity can have on nanoscale materials and devices consisting of crystalline silk materials.

Improved strength of silk fibers in Bombyx mori trimolters induced by an anti-juvenile hormone compound.

PubMed

Guo, Kaiyu; Dong, Zhaoming; Zhang, Yan; Wang, Dandan; Tang, Muya; Zhang, Xiaolu; Xia, Qingyou; Zhao, Ping

2018-05-01

Bombyx mori silk fibers with thin diameters have advantages of lightness and crease-resistance. Many studies have used anti-juvenile hormones to induce trimolters in order to generate thin silk; however, there has been comparatively little analysis of the morphology, structure and mechanical properties of trimolter silk. This study induced two kinds of trimolters by appling topically anti-juvenile hormones and obtained thin diameter silk. Scanning electron microscope (SEM), FTIR analysis, tensile mechanical testing, chitin staining were used to reveal that the morphology, conformation and mechanical property of the trimolter silk. Cocoon of trimolters were highly densely packed by thinner fibers and thus had small apertures. We found that the conformation of trimolter silk fibroin changed and formed more β-sheet structures. In addition, analysis of mechanical parameters yielded a higher Young's modulus and strength in trimolter silk than in the control. By chitin staining of silk gland, we postulated that the mechanical properties of trimolters' silk was enhanced greatly during to the structural changes of silk gland. We induced trimolters by anti-juvenile hormones and the resulting cocoons were more closely packed and had smaller silk fiber diameters. We found that the conformation of trimolters silk fibroin had a higher content of β-sheet structures and better mechanical properties. Our study revealed the structures and mechanical properties of trimolter silk, and provided a valuable reference to improve silk quality by influencing molting in silkworms. Copyright © 2018 Elsevier B.V. All rights reserved.

Fabrication and Optimization of Stable, Optically Transparent, and Reusable pH-Responsive Silk Membranes

PubMed Central

Toytziaridis, Andreas; Dicko, Cedric

2016-01-01

The fabrication of silk-based membranes that are stable, optically transparent and reusable is yet to be achieved. To address this bottleneck we have developed a method to produce transparent chromogenic silk patches that are optically responsive to pH. The patches were produced by blending regenerated silk fibroin (RSF), Laponite RD (nano clay) and the organic dyes neutral red and Thionine acetate. The Laponite RD played a central role in the patch mechanical integrity and prevention of dye leaching. The process was optimized using a factorial design to maximize the patch response to pH by UV absorbance and fluorescence emission. New patches of the optimized protocol, made from solutions containing 125 μM neutral red or 250 μM of Thionine and 15 mg/mL silk, were further tested for operational stability over several cycles of pH altering. Stability, performance, and reusability were achieved over the tested cycles. The approach could be extended to other reporting molecules or enzymes able to bind to Laponite. PMID:27854303

A high selective methanol gas sensor based on molecular imprinted Ag-LaFeO3 fibers.

PubMed

Rong, Qian; Zhang, Yumin; Wang, Chao; Zhu, Zhongqi; Zhang, Jin; Liu, Qingju

2017-09-21

Ag-LaFeO 3 molecularly imprinted polymers (ALMIPs) were fabricated, which provided special recognition sites to methanol. Then ALMIPs fiber 1, fiber 2 and fiber 3 were prepared using filter paper, silk and carbon fibers template, respectively. Based on the observation of X-ray diffraction (XRD), scanning electron microscope (SEM), transmission electron microscope (TEM), and Nitrogen adsorption surface area analyzer (BET), the structure, morphology and surface area of the fibers were characterized. The ALMIPs fibers (fiber 1, fiber 2 and fiber 3) show excellent selectivity and good response to methanol. The responses to 5 ppm methanol and the optimal operating temperature of ALMIPs fibers are 23.5 and 175 °C (fiber 1), 19.67 and 125 °C (fiber 2), 17.59 and 125 °C (fiber 3), and a lower response (≤10, 3, 2) to other test gases including formaldehyde, acetone, ethanol, ammonia, gasoline and benzene was measured, respectively.

Post-secretion processing influences spider silk performance

PubMed Central

Blamires, Sean J.; Wu, Chung-Lin; Blackledge, Todd A.; Tso, I-Min

2012-01-01

Phenotypic variation facilitates adaptations to novel environments. Silk is an example of a highly variable biomaterial. The two-spidroin (MaSp) model suggests that spider major ampullate (MA) silk is composed of two proteins—MaSp1 predominately contains alanine and glycine and forms strength enhancing β-sheet crystals, while MaSp2 contains proline and forms elastic spirals. Nonetheless, mechanical properties can vary in spider silks without congruent amino acid compositional changes. We predicted that post-secretion processing causes variation in the mechanical performance of wild MA silk independent of protein composition or spinning speed across 10 species of spider. We used supercontraction to remove post-secretion effects and compared the mechanics of silk in this ‘ground state’ with wild native silks. Native silk mechanics varied less among species compared with ‘ground state’ silks. Variability in the mechanics of ‘ground state’ silks was associated with proline composition. However, variability in native silks did not. We attribute interspecific similarities in the mechanical properties of native silks, regardless of amino acid compositions, to glandular processes altering molecular alignment of the proteins prior to extrusion. Such post-secretion processing may enable MA silk to maintain functionality across environments, facilitating its function as a component of an insect-catching web. PMID:22628213

Silk from crickets: a new twist on spinning.

PubMed

Walker, Andrew A; Weisman, Sarah; Church, Jeffrey S; Merritt, David J; Mudie, Stephen T; Sutherland, Tara D

2012-01-01

Raspy crickets (Orthoptera: Gryllacrididae) are unique among the orthopterans in producing silk, which is used to build shelters. This work studied the material composition and the fabrication of cricket silk for the first time. We examined silk-webs produced in captivity, which comprised cylindrical fibers and flat films. Spectra obtained from micro-Raman experiments indicated that the silk is composed of protein, primarily in a beta-sheet conformation, and that fibers and films are almost identical in terms of amino acid composition and secondary structure. The primary sequences of four silk proteins were identified through a mass spectrometry/cDNA library approach. The most abundant silk protein was large in size (300 and 220 kDa variants), rich in alanine, glycine and serine, and contained repetitive sequence motifs; these are features which are shared with several known beta-sheet forming silk proteins. Convergent evolution at the molecular level contrasts with development by crickets of a novel mechanism for silk fabrication. After secretion of cricket silk proteins by the labial glands they are fabricated into mature silk by the labium-hypopharynx, which is modified to allow the controlled formation of either fibers or films. Protein folding into beta-sheet structure during silk fabrication is not driven by shear forces, as is reported for other silks.

Study on Mobile Object Positioning and Alarming System Based on the “Map World” in the Core Area of the Silk Road Economic Belt

NASA Astrophysics Data System (ADS)

Mu, Kai

2017-02-01

The established “Map World” on the National Geographic Information Public Service Platform offers free access to many geographic information in the Core Area of the Silk Road Economic Belt. Considering the special security situation and severe splittism and anti-splittism struggles in the Core Area of the Silk Road Economic Belt, a set of moving target positioning and alarming platform based on J2EE platform and B/S structure was designed and realized by combining the “Map World” data and global navigation satellite system. This platform solves various problems, such as effective combination of Global Navigation Satellite System (GNSS) and “Map World” resources, moving target alarming setting, inquiry of historical routes, system management, etc.

Multifunctional silk-heparin biomaterials for vascular tissue engineering applications

PubMed Central

Seib, F. Philipp; Herklotz, Manuela; Burke, Kelly A.; Maitz, Manfred F.; Werner, Carsten; Kaplan, David L.

2013-01-01

Over the past 30 years, silk has been proposed for numerous biomedical applications that go beyond its traditional use as a suture material. Silk sutures are well tolerated in humans, but the use of silk for vascular engineering applications still requires extensive biocompatibility testing. Some studies have indicated a need to modify silk to yield a hemocompatible surface. This study examined the potential of low molecular weight heparin as a material for refining silk properties by acting as a carrier for vascular endothelial growth factor (VEGF) and improving silk hemocompatibility. Heparinized silk showed a controlled VEGF release over 6 days; the released VEGF was bioactive and supported the growth of human endothelial cells. Silk samples were then assessed using a humanized hemocompatibility system that employs whole blood and endothelial cells. The overall thrombogenic response for silk was very low and similar to the clinical reference material polytetrafluoroethylene. Despite an initial inflammatory response to silk, apparent as complement and leukocyte activation, the endothelium was maintained in a resting, anticoagulant state. The low thrombogenic response and the ability to control VEGF release support the further development of silk for vascular applications. PMID:24099708

Single Honeybee Silk Protein Mimics Properties of Multi-Protein Silk

PubMed Central

Sutherland, Tara D.; Church, Jeffrey S.; Hu, Xiao; Huson, Mickey G.; Kaplan, David L.; Weisman, Sarah

2011-01-01

Honeybee silk is composed of four fibrous proteins that, unlike other silks, are readily synthesized at full-length and high yield. The four silk genes have been conserved for over 150 million years in all investigated bee, ant and hornet species, implying a distinct functional role for each protein. However, the amino acid composition and molecular architecture of the proteins are similar, suggesting functional redundancy. In this study we compare materials generated from a single honeybee silk protein to materials containing all four recombinant proteins or to natural honeybee silk. We analyse solution conformation by dynamic light scattering and circular dichroism, solid state structure by Fourier Transform Infrared spectroscopy and Raman spectroscopy, and fiber tensile properties by stress-strain analysis. The results demonstrate that fibers artificially generated from a single recombinant silk protein can reproduce the structural and mechanical properties of the natural silk. The importance of the four protein complex found in natural silk may lie in biological silk storage or hierarchical self-assembly. The finding that the functional properties of the mature material can be achieved with a single protein greatly simplifies the route to production for artificial honeybee silk. PMID:21311767

Evaluations of candidate encapsulation designs and materials for low-cost silicon photovoltaic arrays

NASA Technical Reports Server (NTRS)

Gaines, G. B.; Carmichael, D. C.; Sliemers, F. A.; Brockway, M. C.; Bunk, A. R.; Nance, G. P.

1978-01-01

Three encapsulation designs for silicon photovoltaic arrays based on cells with silk-screened Ag metallization have been evaluated: transparent polymeric coatings over cells laminated between two films or sheets of polymeric materials; cells adhesively bonded to a glass cover with a polymer pottant and a glass or other substrate component. Silicone and acrylic coatings were assessed, together with acrylic sheet, 0.635 mm fiberglass-reinforced polyester sheet, 0.102 mm polycarbonate/acrylic dual-layer film, 0.127 mm fluorocarbon film, soda-lime glass, borosilicate glass, low-iron glass, and several adhesives. The encapsulation materials were characterized by light transmittance measurements, determination of moisture barrier properties and bond strengths, and by the performance of cells before and after encapsulation. Silicon and acrylic coatings provided inadequate protection. Acrylic and fluorocarbon films displayed good weatherability and acceptable optical transmittance. Borosilicate, low-iron and soda-lime-float glasses were found to be acceptable candidate encapsulants for most environments.

Injectable silk-based biomaterials for cervical tissue augmentation: an in vitro study.

PubMed

Brown, Joseph E; Partlow, Benjamin P; Berman, Alison M; House, Michael D; Kaplan, David L

2016-01-01

Cerclage therapy is an important treatment option for preterm birth prevention. Several patient populations benefit from cerclage therapy including patients with a classic history of cervical insufficiency; patients who present with advanced cervical dilation prior to viability; and patients with a history of preterm birth and cervical shortening. Although cerclage is an effective treatment option in some patients, it can be associated with limited efficacy and procedure complications. Development of an alternative to cerclage therapy would be an important clinical development. Here we report on an injectable, silk protein-based biomaterial for cervical tissue augmentation. The rationale for the development of an injectable biomaterial is to restore the native properties of cervical tissue. While cerclage provides support to the tissue, it does not address excessive tissue softening, which is a central feature of the pathogenesis of cervical insufficiency. Silk protein-based hydrogels, which are biocompatible and naturally degrade in vivo, are suggested as a platform for restoring the native properties of cervical tissue and improving cervical function. We sought to study the properties of an injectable, silk-based biomaterial for potential use as an alternative treatment for cervical insufficiency. These biomaterials were evaluated for mechanical tunability, biocompatibility, facile injection, and in vitro degradation. Silk protein solutions were cross-linked by an enzyme catalyzed reaction to form elastic biomaterials. Biomaterials were formulated to match the native physical properties of cervical tissue during pregnancy. The cell compatibility of the materials was assessed in vitro using cervical fibroblasts, and biodegradation was evaluated using concentrated protease solution. Tissue augmentation or bulking was demonstrated using human cervical tissue from nonpregnant hysterectomy specimens. Mechanical compression tests measured the tissue stiffness as a function of the volume of injected biomaterial. Silk protein concentration, molecular weight, and concentration of cross-linking agent were varied to generate biomaterials that functioned from hard gels to viscous fluids. Biomaterials that matched the mechanical features of cervical tissues were chosen for further study. Cervical fibroblasts cultured on these biomaterials were proliferative and metabolically active over 6 days. Biomaterials were degraded in protease solution, with rate of mass loss dependent on silk protein molecular weight. Injection of cervical tissue samples with 100 μL of the biomaterial resulted in a significant volume increase (22.6% ± 8.8%, P < .001) with no significant change in tissue stiffness. Cytocompatible, enzyme cross-linked silk protein biomaterials show promise as a tissue bulking agent. The biomaterials were formulated to match the native mechanical properties of human cervical tissue. These biomaterials should be explored further as a possible alternative to cerclage for providing support to the cervix during pregnancy. Copyright © 2016 Elsevier Inc. All rights reserved.

Analysis of proteome dynamics inside the silk gland lumen of Bombyx mori.

PubMed

Dong, Zhaoming; Zhao, Ping; Zhang, Yan; Song, Qianru; Zhang, Xiaolu; Guo, Pengchao; Wang, Dandan; Xia, Qingyou

2016-04-22

The silk gland is the only organ where silk proteins are synthesized and secreted in the silkworm, Bombyx mori. Silk proteins are stored in the lumen of the silk gland for around eight days during the fifth instar. Determining their dynamic changes is helpful for clarifying the secretion mechanism of silk proteins. Here, we identified the proteome in the silk gland lumen using liquid chromatography-tandem mass spectrometry, and demonstrated its changes during two key stages. From day 5 of the fifth instar to day 1 of wandering, the abundances of fibroins, sericins, seroins, and proteins of unknown functions increased significantly in different compartments of the silk gland lumen. As a result, these accumulated proteins constituted the major cocoon components. In contrast, the abundances of enzymes and extracellular matrix proteins decreased in the silk gland lumen, suggesting that they were not the structural constituents of silk. Twenty-five enzymes may be involved in the regulation of hormone metabolism for proper silk gland function. In addition, the metabolism of other non-proteinous components such as chitin and pigment were also discussed in this study.

Analysis of proteome dynamics inside the silk gland lumen of Bombyx mori

PubMed Central

Dong, Zhaoming; Zhao, Ping; Zhang, Yan; Song, Qianru; Zhang, Xiaolu; Guo, Pengchao; Wang, Dandan; Xia, Qingyou

2016-01-01

The silk gland is the only organ where silk proteins are synthesized and secreted in the silkworm, Bombyx mori. Silk proteins are stored in the lumen of the silk gland for around eight days during the fifth instar. Determining their dynamic changes is helpful for clarifying the secretion mechanism of silk proteins. Here, we identified the proteome in the silk gland lumen using liquid chromatography–tandem mass spectrometry, and demonstrated its changes during two key stages. From day 5 of the fifth instar to day 1 of wandering, the abundances of fibroins, sericins, seroins, and proteins of unknown functions increased significantly in different compartments of the silk gland lumen. As a result, these accumulated proteins constituted the major cocoon components. In contrast, the abundances of enzymes and extracellular matrix proteins decreased in the silk gland lumen, suggesting that they were not the structural constituents of silk. Twenty-five enzymes may be involved in the regulation of hormone metabolism for proper silk gland function. In addition, the metabolism of other non-proteinous components such as chitin and pigment were also discussed in this study. PMID:27102218

High-Toughness Silk Produced by a Transgenic Silkworm Expressing Spider (Araneus ventricosus) Dragline Silk Protein

PubMed Central

Kuwana, Yoshihiko; Sezutsu, Hideki; Nakajima, Ken-ichi; Tamada, Yasushi; Kojima, Katsura

2014-01-01

Spider dragline silk is a natural fiber that has excellent tensile properties; however, it is difficult to produce artificially as a long, strong fiber. Here, the spider (Araneus ventricosus) dragline protein gene was cloned and a transgenic silkworm was generated, that expressed the fusion protein of the fibroin heavy chain and spider dragline protein in cocoon silk. The spider silk protein content ranged from 0.37 to 0.61% w/w (1.4–2.4 mol%) native silkworm fibroin. Using a good silk-producing strain, C515, as the transgenic silkworm can make the raw silk from its cocoons for the first time. The tensile characteristics (toughness) of the raw silk improved by 53% after the introduction of spider dragline silk protein; the improvement depended on the quantity of the expressed spider dragline protein. To demonstrate the commercial feasibility for machine reeling, weaving, and sewing, we used the transgenic spider silk to weave a vest and scarf; this was the first application of spider silk fibers from transgenic silkworms. PMID:25162624

High-toughness silk produced by a transgenic silkworm expressing spider (Araneus ventricosus) dragline silk protein.

PubMed

Kuwana, Yoshihiko; Sezutsu, Hideki; Nakajima, Ken-ichi; Tamada, Yasushi; Kojima, Katsura

2014-01-01

Spider dragline silk is a natural fiber that has excellent tensile properties; however, it is difficult to produce artificially as a long, strong fiber. Here, the spider (Araneus ventricosus) dragline protein gene was cloned and a transgenic silkworm was generated, that expressed the fusion protein of the fibroin heavy chain and spider dragline protein in cocoon silk. The spider silk protein content ranged from 0.37 to 0.61% w/w (1.4-2.4 mol%) native silkworm fibroin. Using a good silk-producing strain, C515, as the transgenic silkworm can make the raw silk from its cocoons for the first time. The tensile characteristics (toughness) of the raw silk improved by 53% after the introduction of spider dragline silk protein; the improvement depended on the quantity of the expressed spider dragline protein. To demonstrate the commercial feasibility for machine reeling, weaving, and sewing, we used the transgenic spider silk to weave a vest and scarf; this was the first application of spider silk fibers from transgenic silkworms.

The role of capture spiral silk properties in the diversification of orb webs.

PubMed

Tarakanova, Anna; Buehler, Markus J

2012-12-07

Among a myriad of spider web geometries, the orb web presents a fascinating, exquisite example in architecture and evolution. Orb webs can be divided into two categories according to the capture silk used in construction: cribellate orb webs (composed of pseudoflagelliform silk) coated with dry cribellate threads and ecribellate orb webs (composed of flagelliform silk fibres) coated by adhesive glue droplets. Cribellate capture silk is generally stronger but less-extensible than viscid capture silk, and a body of phylogenic evidence suggests that cribellate capture silk is more closely related to the ancestral form of capture spiral silk. Here, we use a coarse-grained web model to investigate how the mechanical properties of spiral capture silk affect the behaviour of the whole web, illustrating that more elastic capture spiral silk yields a decrease in web system energy absorption, suggesting that the function of the capture spiral shifted from prey capture to other structural roles. Additionally, we observe that in webs with more extensible capture silk, the effect of thread strength on web performance is reduced, indicating that thread elasticity is a dominant driving factor in web diversification.

Stem cell-extracellular vesicles as drug delivery systems: New frontiers for silk/curcumin nanoparticles.

PubMed

Perteghella, Sara; Crivelli, Barbara; Catenacci, Laura; Sorrenti, Milena; Bruni, Giovanna; Necchi, Vittorio; Vigani, Barbara; Sorlini, Marzio; Torre, Maria Luisa; Chlapanidas, Theodora

2017-03-30

The aim of this work was to develop a novel carrier-in-carrier system based on stem cell-extracellular vesicles loaded of silk/curcumin nanoparticles by endogenous technique. Silk nanoparticles were produced by desolvation method and curcumin has been selected as drug model because of its limited water solubility and poor bioavailability. Nanoparticles were stable, with spherical geometry, 100nm in average diameter and the drug content reached about 30%. Cellular uptake studies, performed on mesenchymal stem cells (MSCs), showed the accumulation of nanoparticles in the cytosol around the nuclear membrane, without cytotoxic effects. Finally, MSCs were able to release extracellular vesicles entrapping silk/curcumin nanoparticles. This combined biological-technological approach represents a novel class of nanosystems, combining beneficial effects of both regenerative cell therapies and pharmaceutical nanomedicine, avoiding the use of viable replicating stem cells. Copyright © 2017 Elsevier B.V. All rights reserved.

Liquid crystalline spinning of spider silk

NASA Astrophysics Data System (ADS)

Vollrath, Fritz; Knight, David P.

2001-03-01

Spider silk has outstanding mechanical properties despite being spun at close to ambient temperatures and pressures using water as the solvent. The spider achieves this feat of benign fibre processing by judiciously controlling the folding and crystallization of the main protein constituents, and by adding auxiliary compounds, to create a composite material of defined hierarchical structure. Because the `spinning dope' (the material from which silk is spun) is liquid crystalline, spiders can draw it during extrusion into a hardened fibre using minimal forces. This process involves an unusual internal drawdown within the spider's spinneret that is not seen in industrial fibre processing, followed by a conventional external drawdown after the dope has left the spinneret. Successful copying of the spider's internal processing and precise control over protein folding, combined with knowledge of the gene sequences of its spinning dopes, could permit industrial production of silk-based fibres with unique properties under benign conditions.

Functional hepatocyte clusters on bioactive blend silk matrices towards generating bioartificial liver constructs.

PubMed

Janani, G; Nandi, Samit K; Mandal, Biman B

2018-02-01

The creation of in vitro functional hepatic tissue simulating micro-environmental niche of native liver is a keen area of research due to its demand in bioartificial liver (BAL) and cell-based tissue engineering. Here, we investigated the potential of novel blend (BA) silk scaffold fabricated by blending mulberry (Bombyx mori, BM) silk fibroin with cell adhesion motif (RGD) rich non-mulberry (Antheraea assamensis, AA) silk fibroin, in generating a functional liver construct. Three-dimensional (3D) porous silk scaffolds (BM, AA and BA) were physico-chemically characterized and functionally evaluated using human hepatocarcinoma cells (HepG2) and primary neonatal rat hepatocytes. The growth and distribution of hepatocytes within the scaffolds were tracked by FESEM, alamar blue proliferation assay and live/dead staining. Hemocompatible BA scaffolds supported the formation of high density hepatocyte clusters, facilitating cell-matrix and cell-cell interactions. Blend scaffolds evinced enhanced liver-specific functions of cultured hepatocytes in terms of albumin synthesis, urea synthesis and cytochrome P450 enzyme activity over 21 days. Subcutaneous implantation of scaffolds demonstrated minimal macrophage infiltration in blend scaffolds. These findings substantiate that the integral property of blend (BA) scaffold offers a befitting environment by influencing spheroidal growth of hepatocytes with enhanced biological activity. Collectively, the present study provides a new 3D bio-matrix niche for growing functional liver cells that would have future prospects in BAL as well as regenerative medicine. An end stage liver disease called cirrhosis perturbs the self-healing ability and physiological functions of liver. Due to the scarcity of healthy donors, a functional in vitro hepatic construct retaining the liver-specific functions is in great demand for its prospects in bioartificial liver (BAL) and cell-based tissue engineering. Physicochemical attributes of a matrix influence the behavior of cultured hepatocytes in terms of attachment, morphology and functionality. Mulberry and non-mulberry silk fibroin presents unique amino acid sequence with difference in hydrophobicity and crystallinity. Considering this, the present study focuses on the development of a suitable three-dimensional (3D) bioactive matrix incorporating both mulberry silk fibroin and cell adhesion motif (RGD) rich non-mulberry silk fibroin. Porous silk blend scaffolds facilitated the formation of hepatocyte clusters with enhanced liver-specific functions emphasizing both cell-cell and cell-matrix interactions. Hemocompatibility and integral property of blend scaffolds offers a biological niche for seeding functional liver cells that would have future prospects in biohybrid devices. Copyright © 2017 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

Synergistic toughening of composite fibres by self-alignment of reduced graphene oxide and carbon nanotubes

PubMed Central

Shin, Min Kyoon; Lee, Bommy; Kim, Shi Hyeong; Lee, Jae Ah; Spinks, Geoffrey M.; Gambhir, Sanjeev; Wallace, Gordon G.; Kozlov, Mikhail E.; Baughman, Ray H.; Kim, Seon Jeong

2012-01-01

The extraordinary properties of graphene and carbon nanotubes motivate the development of methods for their use in producing continuous, strong, tough fibres. Previous work has shown that the toughness of the carbon nanotube-reinforced polymer fibres exceeds that of previously known materials. Here we show that further increased toughness results from combining carbon nanotubes and reduced graphene oxide flakes in solution-spun polymer fibres. The gravimetric toughness approaches 1,000 J g−1, far exceeding spider dragline silk (165 J g−1) and Kevlar (78 J g−1). This toughness enhancement is consistent with the observed formation of an interconnected network of partially aligned reduced graphene oxide flakes and carbon nanotubes during solution spinning, which act to deflect cracks and allow energy-consuming polymer deformation. Toughness is sensitive to the volume ratio of the reduced graphene oxide flakes to the carbon nanotubes in the spinning solution and the degree of graphene oxidation. The hybrid fibres were sewable and weavable, and could be shaped into high-modulus helical springs. PMID:22337128

Autonomous self-healing structural composites with bio-inspired design

PubMed Central

D’Elia, Eleonora; Eslava, Salvador; Miranda, Miriam; Georgiou, Theoni K.; Saiz, Eduardo

2016-01-01

Strong and tough natural composites such as bone, silk or nacre are often built from stiff blocks bound together using thin interfacial soft layers that can also provide sacrificial bonds for self-repair. Here we show that it is possible exploit this design in order to create self-healing structural composites by using thin supramolecular polymer interfaces between ceramic blocks. We have built model brick-and-mortar structures with ceramic contents above 95 vol% that exhibit strengths of the order of MPa (three orders of magnitude higher than the interfacial polymer) and fracture energies that are two orders of magnitude higher than those of the glass bricks. More importantly, these properties can be fully recovered after fracture without using external stimuli or delivering healing agents. This approach demonstrates a very promising route towards the design of strong, ideal self-healing materials able to self-repair repeatedly without degradation or external stimuli. PMID:27146382

Autonomous self-healing structural composites with bio-inspired design.

PubMed

D'Elia, Eleonora; Eslava, Salvador; Miranda, Miriam; Georgiou, Theoni K; Saiz, Eduardo

2016-05-05

Strong and tough natural composites such as bone, silk or nacre are often built from stiff blocks bound together using thin interfacial soft layers that can also provide sacrificial bonds for self-repair. Here we show that it is possible exploit this design in order to create self-healing structural composites by using thin supramolecular polymer interfaces between ceramic blocks. We have built model brick-and-mortar structures with ceramic contents above 95 vol% that exhibit strengths of the order of MPa (three orders of magnitude higher than the interfacial polymer) and fracture energies that are two orders of magnitude higher than those of the glass bricks. More importantly, these properties can be fully recovered after fracture without using external stimuli or delivering healing agents. This approach demonstrates a very promising route towards the design of strong, ideal self-healing materials able to self-repair repeatedly without degradation or external stimuli.

Autonomous self-healing structural composites with bio-inspired design

NASA Astrophysics Data System (ADS)

D'Elia, Eleonora; Eslava, Salvador; Miranda, Miriam; Georgiou, Theoni K.; Saiz, Eduardo

2016-05-01

Strong and tough natural composites such as bone, silk or nacre are often built from stiff blocks bound together using thin interfacial soft layers that can also provide sacrificial bonds for self-repair. Here we show that it is possible exploit this design in order to create self-healing structural composites by using thin supramolecular polymer interfaces between ceramic blocks. We have built model brick-and-mortar structures with ceramic contents above 95 vol% that exhibit strengths of the order of MPa (three orders of magnitude higher than the interfacial polymer) and fracture energies that are two orders of magnitude higher than those of the glass bricks. More importantly, these properties can be fully recovered after fracture without using external stimuli or delivering healing agents. This approach demonstrates a very promising route towards the design of strong, ideal self-healing materials able to self-repair repeatedly without degradation or external stimuli.

Characterization and assembly of a GFP-tagged cylindriform silk into hexameric complexes.

PubMed

Öster, Carl; Svensson Bonde, Johan; Bülow, Leif; Dicko, Cedric

2014-04-01

Spider silk has been studied extensively for its attractive mechanical properties and potential applications in medicine and industry. The production of spider silk, however, has been lagging behind for lack of suitable systems. Our approach focuses on solving the production of spider silk by designing, expressing, purifying and characterizing the silk from cylindriform glands. We show that the cylindriform silk protein, in contrast to the commonly used dragline silk protein, is fully folded and stable in solution. With the help of GFP as a fusion tag we enhanced the expression of the silk protein in Escherichia coli and could optimize the downstream processing. Secondary structures analysis by circular dichroism and FTIR shows that the GFP-silk fusion protein is predominantly α-helical, and that pH can trigger a α- to β-transition resulting in aggregation. Structural analysis by small angle X-ray scattering suggests that the GFP-Silk exists in the form of a hexamer in solution. Copyright © 2013 Wiley Periodicals, Inc.

Elastic, silk-cardiac extracellular matrix hydrogels exhibit time-dependent stiffening that modulates cardiac fibroblast response

PubMed Central

Stoppel, Whitney L.; Gao, Albert E.; Greaney, Allison M.; Partlow, Benjamin P.; Bretherton, Ross C.; Kaplan, David L.; Black, Lauren D.

2018-01-01

Heart failure is the leading cause of death in the United States and rapidly becoming the leading cause of death worldwide. While pharmacological treatments can reduce progression to heart failure following myocardial infarction, there still exists a need for new therapies that promote better healing post injury for a more functional cardiac repair and methods to understand how the changes to tissue mechanical properties influence cell phenotype and function following injury. To address this need, we have optimized a silk-based hydrogel platform containing cardiac tissue-derived extracellular matrix (cECM). These silk-cECM hydrogels have tunable mechanical properties, as well as rate-controllable hydrogel stiffening over time. In vitro, silk-cECM scaffolds led to enhanced cardiac fibroblast (CF) cell growth and viability with culture time. cECM incorporation improved expression of integrin an focal adhesion proteins, suggesting that CFs were able to interact with the cECM in the hydrogel. Subcutaneous injection of silk hydrogels in rats demonstrated that addition of the cECM led to endogenous cell infiltration and promoted endothelial cell ingrowth after 4 weeks in vivo. This naturally derived silk fibroin platform is applicable to the development of more physiologically relevant constructs that replicate healthy and diseased tissue in vitro and has the potential to be used as an injectable therapeutic for cardiac repair. PMID:27480328

Spider Transcriptomes Identify Ancient Large-Scale Gene Duplication Event Potentially Important in Silk Gland Evolution

PubMed Central

Clarke, Thomas H.; Garb, Jessica E.; Hayashi, Cheryl Y.; Arensburger, Peter; Ayoub, Nadia A.

2015-01-01

The evolution of specialized tissues with novel functions, such as the silk synthesizing glands in spiders, is likely an influential driver of adaptive success. Large-scale gene duplication events and subsequent paralog divergence are thought to be required for generating evolutionary novelty. Such an event has been proposed for spiders, but not tested. We de novo assembled transcriptomes from three cobweb weaving spider species. Based on phylogenetic analyses of gene families with representatives from each of the three species, we found numerous duplication events indicative of a whole genome or segmental duplication. We estimated the age of the gene duplications relative to several speciation events within spiders and arachnids and found that the duplications likely occurred after the divergence of scorpions (order Scorpionida) and spiders (order Araneae), but before the divergence of the spider suborders Mygalomorphae and Araneomorphae, near the evolutionary origin of spider silk glands. Transcripts that are expressed exclusively or primarily within black widow silk glands are more likely to have a paralog descended from the ancient duplication event and have elevated amino acid replacement rates compared with other transcripts. Thus, an ancient large-scale gene duplication event within the spider lineage was likely an important source of molecular novelty during the evolution of silk gland-specific expression. This duplication event may have provided genetic material for subsequent silk gland diversification in the true spiders (Araneomorphae). PMID:26058392

Improvement of Physical and Wound Adhesion Properties of Silk Sericin and Polyvinyl Alcohol Dressing Using Glycerin.

PubMed

Aramwit, Pornanong; Ratanavaraporn, Juthamas; Siritientong, Tippawan

2015-08-01

This study aimed to use glycerin to improve physical and wound adhesion properties of a wound dressing made of silk sericin and polyvinyl alcohol (PVA). Glycerin of a natural-derived plasticizer was used to modify the properties of silk sericin/PVA scaffolds. Various concentrations of glycerin were mixed with silk sericin and PVA and then fabricated into the scaffolds by a freeze-drying technique. The control study was performed to examine the properties of the silk sericin/PVA scaffolds with and without glycerin. Physical, mechanical, wound adhesion properties, the release profile of silk sericin, and in vivo safety of the silk sericin/PVA scaffolds with and without glycerin were investigated. The silk sericin/PVA scaffolds with glycerin exhibited more homogenous structure, less compressive modulus, higher Young modulus and elongation percentage, and a higher degree of crosslinking compared with the scaffold without glycerin. The silk sericin/PVA scaffold with 2% wt/vol glycerin showed more controlled release of silk sericin than the other scaffolds. The sustained release of silk sericin from the scaffold with glycerin would be advantageous for long-term healing of wounds. The silk sericin/PVA scaffold with 2% (wt/vol) glycerin was less adhesive to the wound compared with the scaffold without glycerin. Furthermore, the implantation of silk sericin/PVA scaffolds with 2% (wt/vol) glycerin did not cause any irritation to the tissue. The silk sericin/PVA scaffolds with glycerin were introduced as a biocompatible, more flexible, and less adhesive wound dressing than the scaffold without glycerin.

Silk gene expression of theridiid spiders: implications for male-specific silk use.

PubMed

Correa-Garhwal, Sandra M; Chaw, R Crystal; Clarke, Thomas H; Ayoub, Nadia A; Hayashi, Cheryl Y

2017-06-01

Spiders (order Araneae) rely on their silks for essential tasks, such as dispersal, prey capture, and reproduction. Spider silks are largely composed of spidroins, members of a protein family that are synthesized in silk glands. As needed, silk stored in silk glands is extruded through spigots on the spinnerets. Nearly all studies of spider silks have been conducted on females; thus, little is known about male silk biology. To shed light on silk use by males, we compared silk gene expression profiles of mature males to those of females from three cob-web weaving species (Theridiidae). We de novo assembled species-specific male transcriptomes from Latrodectus hesperus, Latrodectus geometricus, and Steatoda grossa followed by differential gene expression analyses. Consistent with their complement of silk spigots, male theridiid spiders express appreciable amounts of aciniform, major ampullate, minor ampullate, and pyriform spidroin genes but not tubuliform spidroin genes. The relative expression levels of particular spidroin genes varied between sexes and species. Because mature males desert their prey-capture webs and become cursorial in their search for mates, we anticipated that major ampullate (dragline) spidroin genes would be the silk genes most highly expressed by males. Indeed, major ampullate spidroin genes had the highest expression in S. grossa males. However, minor ampullate spidroin genes were the most highly expressed spidroin genes in L. geometricus and L. hesperus males. Our expression profiling results suggest species-specific adaptive divergence of silk use by male theridiids. Copyright © 2017 The Authors. Published by Elsevier GmbH.. All rights reserved.

Lithium-free processing of silk fibroin.

PubMed

Zheng, Zhaozhu; Guo, Shaozhe; Liu, Yawen; Wu, Jianbing; Li, Gang; Liu, Meng; Wang, Xiaoqin; Kaplan, David

2016-09-01

Silk fibroin protein was purified from Bombyx mori silkworm cocoons using a novel dialysis strategy to avoid fibroin aggregation and pre-mature formation of β-sheets. The degummed silk fibers were dissolved in Ajisawa's reagent, a mixture of CaCl2-EtOH-H2O, that is less expensive than lithium bromide. The dissolved solutions were dialyzed against either water or urea solution with a stepwise decrease in concentration. When the steps of 4 M-2 M-1 M-0 M urea (referred to as silk-TS-4210) were adopted, the purified silk fibroin had smaller aggregates (<10 nm), similar average molecular weight (225 kDa) and a lower content of β-sheet (∼15%) compared to the sample processing methods (silk-TS-210, 10, 0) studied here. This outcome was close to the fibroin purified by the lithium bromide (silk-Li-0) method. Polyvinyl alcohol-emulsified silk microspheres generated using the purified solution had a similar size distribution and morphology when compared to lithium bromide dissolved solutions, while glycerol-blended silk films showed different mechanical properties. The silk-Li-0 generated films with the highest breaking strength (5.7 MPa ± 0.3) while the silk-TS-4210 had the highest extension at break (215.1% ± 12.5). The films prepared from silk-TS-4210 were cytocompatible to support the adhesion and proliferation of human mesenchymal stem cells, with improvements compared to the other samples likely due to the porous morphology of these films. © The Author(s) 2016.

Synthesis of gold nanoparticles using silk fibroin and their characterization

NASA Astrophysics Data System (ADS)

Gowda, Mahadeva; Harisha, K. S.; Ranjana, T.; Harish, K. V.; Narayana, B.; Byrappa, K.; Sangappa, Y.

2018-05-01

The synthesis of metal nanoparticales by environmentally friendly processes is an important aspect of nanotechnology today. One such approach that shows immense potential is based on the in situ synthesis of gold nanoparticles (AuNPs) using naturally available materials such as aqueous silk fibroin (SF) obtained from Bombyx mori silk. The UV-visible absorption study revealed the formation of AuNPs by showing characteristic surface plasmon resonance (SPR) band at 525 nm. The X-ray diffraction (XRD) analysis study suggests the synthesized gold nanoparticles are FCC crystal structure. The transmission electron microscopy (TEM) images showed that the formed AuNPs are spherical in shape with smooth edges.

Biotechnology and Composite Materials

DTIC Science & Technology

1993-04-01

protein fibroin which are glued together by the protein sericin . Many other insects produce silk, most notably spiders. Spider silks Zre proteins with...silk is boiled to remove the soluble sticky sericin protein, and the remaining fibroin portion of the silk is then unwound and used as silk fiber. Orb

Synthesis of hierarchical three-dimensional copper oxide nanostructures through a biomineralization-inspired approach

NASA Astrophysics Data System (ADS)

Fei, Xiang; Shao, Zhengzhong; Chen, Xin

2013-08-01

Three-dimensional (3D) copper oxide (CuO) nanostructures were synthesized in a regenerated Bombyx mori silk fibroin aqueous solution at room temperature. In the synthesis process, silk fibroin served as the template and helped to form the hierarchical CuO nanostructures by self-assembly. Cu(OH)2 nanowires were formed initially, and then they transformed into almond-like CuO nanostructures with branched edges and a compact middle. The size of the final CuO nanostructures can be tuned by varying the concentration of silk fibroin in the reaction system. A possible mechanism has been proposed based on various characterization techniques, such as scanning and transmission electron microscopy, X-ray diffraction, and thermogravimetric analysis. The synthesized CuO nanostructured material has been evaluated as an anode material for lithium ion batteries, and the result showed that they had a good electrochemical performance. The straightforward energy-saving method developed in this research may provide a useful preparation strategy for other functional inorganic materials through an environmentally friendly process.Three-dimensional (3D) copper oxide (CuO) nanostructures were synthesized in a regenerated Bombyx mori silk fibroin aqueous solution at room temperature. In the synthesis process, silk fibroin served as the template and helped to form the hierarchical CuO nanostructures by self-assembly. Cu(OH)2 nanowires were formed initially, and then they transformed into almond-like CuO nanostructures with branched edges and a compact middle. The size of the final CuO nanostructures can be tuned by varying the concentration of silk fibroin in the reaction system. A possible mechanism has been proposed based on various characterization techniques, such as scanning and transmission electron microscopy, X-ray diffraction, and thermogravimetric analysis. The synthesized CuO nanostructured material has been evaluated as an anode material for lithium ion batteries, and the result showed that they had a good electrochemical performance. The straightforward energy-saving method developed in this research may provide a useful preparation strategy for other functional inorganic materials through an environmentally friendly process. Electronic supplementary information (ESI) available: SEM image of the synthesized CuO without silk fibroin addition, and TEM images of the synthesized CuO with different silk fibroin concentrations. See DOI: 10.1039/c3nr01872e

Spider Webs and Silks.

ERIC Educational Resources Information Center

Vollrath, Fritz

1992-01-01

Compares the attributes of the silk from spiders with those of the commercially harvested silk from silkworms. Discusses the evolution, design, and effectiveness of spider webs; the functional mechanics of the varieties of silk that can be produced by the same spider; and the composite, as well as molecular, structure of spider silk thread. (JJK)

Mechanics and Morphology of Silk Drawn from Anesthetized Spiders

NASA Astrophysics Data System (ADS)

Madsen, B.; Vollrath, F.

CO2 and N2 anesthetized Nephila spiders produced dragline silk with mechanical properties that differed from control silk as a function of time under anesthesia. Silk from CO2 spiders had a significantly lower breaking strain and breaking energy, significantly higher initial modulus, and marginally lower breaking stress. At the onset of anesthesia the silk diameter became highly variable. During deep anesthesia silk either became thinner or retained cross-section but fibrillated.

Facts and myths of antibacterial properties of silk.

PubMed

Kaur, Jasjeet; Rajkhowa, Rangam; Afrin, Tarannum; Tsuzuki, Takuya; Wang, Xungai

2014-03-01

Silk cocoons provide protection to silkworm from biotic and abiotic hazards during the immobile pupal phase of the lifecycle of silkworms. Protection is particularly important for the wild silk cocoons reared in an open and harsh environment. To understand whether some of the cocoon components resist growth of microorganisms, in vitro studies were performed using gram negative bacteria Escherichia coli (E. coli) to investigate antibacterial properties of silk fiber, silk gum, and calcium oxalate crystals embedded inside some cocoons. The results show that the previously reported antibacterial properties of silk cocoons are actually due to residues of chemicals used to isolate/purify cocoon elements, and properly isolated silk fiber, gum, and embedded crystals free from such residues do not have inherent resistance to E. coli. This study removes the uncertainty created by previous studies over the presence of antibacterial properties of silk cocoons, particularly the silk gum and sericin. Copyright © 2013 Wiley Periodicals, Inc.

The role of capture spiral silk properties in the diversification of orb webs

PubMed Central

Tarakanova, Anna; Buehler, Markus J.

2012-01-01

Among a myriad of spider web geometries, the orb web presents a fascinating, exquisite example in architecture and evolution. Orb webs can be divided into two categories according to the capture silk used in construction: cribellate orb webs (composed of pseudoflagelliform silk) coated with dry cribellate threads and ecribellate orb webs (composed of flagelliform silk fibres) coated by adhesive glue droplets. Cribellate capture silk is generally stronger but less-extensible than viscid capture silk, and a body of phylogenic evidence suggests that cribellate capture silk is more closely related to the ancestral form of capture spiral silk. Here, we use a coarse-grained web model to investigate how the mechanical properties of spiral capture silk affect the behaviour of the whole web, illustrating that more elastic capture spiral silk yields a decrease in web system energy absorption, suggesting that the function of the capture spiral shifted from prey capture to other structural roles. Additionally, we observe that in webs with more extensible capture silk, the effect of thread strength on web performance is reduced, indicating that thread elasticity is a dominant driving factor in web diversification. PMID:22896566

High-performance spider webs: integrating biomechanics, ecology and behaviour

PubMed Central

Harmer, Aaron M. T.; Blackledge, Todd A.; Madin, Joshua S.; Herberstein, Marie E.

2011-01-01

Spider silks exhibit remarkable properties, surpassing most natural and synthetic materials in both strength and toughness. Orb-web spider dragline silk is the focus of intense research by material scientists attempting to mimic these naturally produced fibres. However, biomechanical research on spider silks is often removed from the context of web ecology and spider foraging behaviour. Similarly, evolutionary and ecological research on spiders rarely considers the significance of silk properties. Here, we highlight the critical need to integrate biomechanical and ecological perspectives on spider silks to generate a better understanding of (i) how silk biomechanics and web architectures interacted to influence spider web evolution along different structural pathways, and (ii) how silks function in an ecological context, which may identify novel silk applications. An integrative, mechanistic approach to understanding silk and web function, as well as the selective pressures driving their evolution, will help uncover the potential impacts of environmental change and species invasions (of both spiders and prey) on spider success. Integrating these fields will also allow us to take advantage of the remarkable properties of spider silks, expanding the range of possible silk applications from single threads to two- and three-dimensional thread networks. PMID:21036911

Effects of Japanese beetle (Coleoptera: Scarabaeidae) and silk clipping in field corn.

PubMed

Steckel, Sandy; Stewart, S D; Tindall, K V

2013-10-01

Japanese beetle (Popillia japonica Newman) is an emerging silk-feeding insect found in fields in the lower Corn Belt and Midsouthern United States. Studies were conducted in 2010 and 2011 to evaluate how silk clipping in corn affects pollination and yield parameters. Manually clipping silks once daily had modest effects on yield parameters. Sustained clipping by either manually clipping silks three times per day or by caging Japanese beetles onto ears affected total kernel weight if it occurred during early silking (R1 growth stage). Manually clipping silks three times per day for the first 5 d of silking affected the number of kernels per ear, total kernel weight, and the weight of individual kernels. Caged beetles fed on silks and, depending on the number of beetles caged per ear, reduced the number of kernels per ear. Caging eight beetles per ear significantly reduced total kernel weight compared with noninfested ears. Drought stress before anthesis appeared to magnify the impact of silk clipping by Japanese beetles. There was evidence of some compensation for reduced pollination by increasing the size of pollinated kernels within the ear. Our results showed that it requires sustained silk clipping during the first week of silking to have substantial impacts on pollination and yield parameters, at least under good growing conditions. Some states recommend treating for Japanese beetle when three Japanese beetles per ear are found, silks are clipped to < 13 mm, and pollination is < 50% complete, and that recommendation appears to be adequate.

Differential scanning fluorimetry illuminates silk feedstock stability and processability.

PubMed

Dicko, C; Kasoju, N; Hawkins, N; Vollrath, F

2016-01-07

The ability to design and implement silk feedstock formulations for tailored spinning has so far eluded the bioengineers. Recently, the high throughput screening technique of differential scanning fluorimetry (DSF) demonstrated the link between the instability transition temperature (Ti) and the processability of the silk feedstock. Using DSF we screened a large set of chemicals known to affect solvent quality. A multivariate analysis of the results shows that, regardless of the diversity of chemicals, three groupings are significantly distinguishable: G1 = similar to native silk; G2 = largely dominated by electrostatic interactions; and G3 = dominated by chelating interactions. We propose a thermodynamic analysis based on a pre- and post-transition fit to estimate the van't Hoff enthalpies (ΔHv) and the instability temperature (Ti). Our analysis shows that the ΔTi and ΔHv values were distinct: G1 (ΔTi = 0.23 ± 0.2; ΔHv = -159.1 ± 5.6 kcal mol(-1)), G2 (ΔTi = -7.3 ± 0.7; ΔHv = -191.4 ± 5.5 kcal mol(-1)), and G3 (ΔTi = -19.9 ± 3.3; ΔHv = -68.8 ± 6.0 kcal mol(-1)). Our analysis further combined the ΔTi value and the ΔHv value using stability ΔΔG to find that G1 only marginally stabilizes native silks (ΔΔG = -0.15 ± 0.04 kcal mol(-1)), whereas G2 and G3 destabilize native silk (ΔΔG = 3.8 ± 0.11 and ΔΔG = 3.8 ± 0.3 kcal mol(-1), respectively). Here our analysis shows that native silk has a complex multistep transition that is possibly non-cooperative. However, all three groupings also show a direct and cooperative transition with varied stabilization effects. This analysis suggests that native silks are able to sample multiple substates prior to undergoing (or to delay) the final transition. We conclude by hypothesizing that the observed energetic plasticity may be mediated by a fragile packaging of the silk tertiary structure that is readily lost when the solvent quality changes.

Regeneration of Bombyx mori silk nanofibers and nanocomposite fibrils by the electrospinning process

NASA Astrophysics Data System (ADS)

Ayutsede, Jonathan Eyitouyo

In recent years, there has been significant interest in the utilization of natural materials for novel nanoproducts such as tissue engineered scaffolds. Silkworm silk fibers represent one of the strongest natural fibers known. Silkworm silk, a protein-based natural biopolymer, has received renewed interest in recent years due to its unique properties (strength, toughness) and potential applications such as smart textiles, protective clothing and tissue engineering. The traditional 10--20 mum diameter, triangular-shaped Bombyx mori fibers have remained unchanged over the years. However, in our study, we examine the scientific implication and potential applications of reducing the diameter to the nanoscale, changing the triangular shape of the fiber and adding nanofillers in the form of single wall carbon nanotubes (SWNT) by the electrospinning process. The electrospinning process preserves the natural conformation of the silk (random and beta-sheet). The feasibility of changing the properties of the electrospun nanofibers by post processing treatments (annealing and chemical treatment) was investigated. B. mori silk fibroin solution (formic acid) was successfully electrospun to produce uniform nanofibers (as small as 12 nm). Response Surface Methodology (RSM) was applied for the first time to experimental results of electrospinning, to develop a processing window that can reproduce regenerated silk nanofibers of a predictable size (d < 100nm). SWNT-silk multifunctional nanocomposite fibers were fabricated for the first time with anticipated properties (mechanical, thermal and electrically conductive) that may have scientific applications (nerve regeneration, stimulation of cell-scaffold interaction). In order to realize these applications, the following areas need to be addressed: a systematic investigation of the dispersion of the nanotubes in the silk matrix, a determination of new methodologies for characterizing the nanofiber properties and establishing the nature of the silk-SWNT interactions. A new visualization system was developed to characterize the transport properties of the nanofibrous assemblies. The morphological, chemical, structural and mechanical properties of the nanofibers were determined by field emission environmental scanning microscopy, Fourier transform infrared and Raman spectroscopy, wide angle x-ray diffraction and microtensile tester respectively.

Silk fibroin-based scaffolds for tissue engineering

NASA Astrophysics Data System (ADS)

Li, Zi-Heng; Ji, Shi-Chen; Wang, Ya-Zhen; Shen, Xing-Can; Liang, Hong

2013-09-01

Silk fibroin (SF) from the Bombyx mori silkworm exhibits attractive potential applications as biomechanical materials, due to its unique mechanical and biological properties. This review outlines the structure and properties of SF, including of its biocompatibility and biodegradability. It highlights recent researches on the fabrication of various SF-based composites scaffolds that are promising for tissue engineering applications, and discusses synthetic methods of various SF-based composites scaffolds and valuable approaches for controlling cell behaviors to promote the tissue repair. The function of extracellular matrices and their interaction with cells are also reviewed here.

Genetic engineered color silk: fabrication of a photonics material through a bioassisted technology.

PubMed

Shimizu, Katsuhiko

2018-05-15

Silk produced by the silkworm Bombyx mori is an attractive material because of its luster, smooth and soft texture, conspicuous mechanical strength, good biocompatibility, slow biodegradation, and carbon neutral synthesis. Silkworms have been domesticated and bred for production of better quality and quantity of silk, resulting in the development of sericulture and the textile industry. Silk is generally white, so dyeing is required to obtain colored fiber. However, the dyeing process involves harsh conditions and generates a large volume of waste water, which have environmentally and economically negative impacts. Although some strains produce cocoons that contain pigments derived from the mulberry leaves that they eat, the pigments are distributed in the sericin layer and are lost during gumming. In trials for production of colored silk by feeding silkworms on diets containing dyes, only limited species of dye molecules were incorporated into the silk threads. A method for the generation of transgenic silkworm was established in conjunction with the discovery of green fluorescent protein (GFP), and silkworms carrying the GFP gene spun silk threads that formed cocoons that glowed bright green and still retained the original properties of silk. A wide range of color variation of silk threads has been obtained by replacing the GFP gene with the genes of other fluorescent proteins chosen from the fluorescent protein palette. The genetically modified silk with photonic properties can be processed to form various products including linear threads, 2D fabrics, and 3D materials. The transgenic colored silk could be economically advantageous due to addition of a new value to silk and reduction of cost for water waste, and environmentally preferable for saving water. Here, I review the literature regarding the production methods of fluorescent silk from transgenic silkworms and present examples of genetically modified color silk.

Curcumin-functionalized silk biomaterials for anti-aging utility.

PubMed

Yang, Lei; Zheng, Zhaozhu; Qian, Cheng; Wu, Jianbing; Liu, Yawen; Guo, Shaozhe; Li, Gang; Liu, Meng; Wang, Xiaoqin; Kaplan, David L

2017-06-15

Curcumin is a natural antioxidant that is isolated from turmeric (Curcuma longa) and exhibits strong free radical scavenging activity, thus functional for anti-aging. However, poor stability and low solubility of curcumin in aqueous conditions limit its biomedical applications. Previous studies have shown that the anti-oxidation activity of curcumin embedded in silk fibroin films could be well preserved, resulting in the promoted adipogenesis from human mesenchymal stem cells (hMSCs) cultured on the surface of the films. In the present study, curcumin was encapsulated in both silk fibroin films (silk/cur films) and nanoparticles (silk/cur NPs), and their anti-aging effects were compared with free curcumin in solution, with an aim to elucidate the mechanism of anti-aging of silk-associated curcumin and to better serve biomedical applications in the future. The morphology and structure of silk/cur film and silk/cur NP were characterized using SEM, FTIR and DSC, indicating characteristic stable beta-sheet structure formation in the materials. Strong binding of curcumin molecules to the beta-sheet domains of silk fibroin resulted in the slow release of curcumin with well-preserved activity from the materials. For cell aging studies, rat bone marrow mesenchymal stem cells (rBMSCs) were cultured in the presence of free curcumin (FC), silk/cur film and silk/cur NP, and cell proliferation and markers of aging (P53, P16, HSP70 gene expression and β-Galactosidase activity) were examined. The results indicated that cell aging was retarded in all FC, silk/cur NP and silk/cur film samples, with the silk-associated curcumin superior to the FC. Copyright © 2017 Elsevier Inc. All rights reserved.

DOE Office of Scientific and Technical Information (OSTI.GOV)

An, Bo; Jenkins, Janelle E; Sampath, Sujatha

Dragline silk from orb-weaving spiders is a copolymer of two large proteins, major ampullate spidroin 1 (MaSp1) and 2 (MaSp2). The ratio of these proteins is known to have a large variation across different species of orb-weaving spiders. NMR results from gland material of two different species of spiders, N. clavipes and A. aurantia, indicates that MaSp1 proteins are more easily formed into β-sheet nanostructures, while MaSp2 proteins form random coil and helical structures. To test if this behavior of natural silk proteins could be reproduced by recombinantly produced spider silk mimic protein, recombinant MaSp1/MaSp2 mixed fibers as well asmore » chimeric silk fibers from MaSp1 and MaSp2 sequences in a single protein were produced based on the variable ratio and conserved motifs of MaSp1 and MaSp2 in native silk fiber. Mechanical properties, solid-state NMR, and XRD results of tested synthetic fibers indicate the differing roles of MaSp1 and MaSp2 in the fiber and verify the importance of postspin stretching treatment in helping the fiber to form the proper spatial structure.« less

A comparative study of the refractive index of silk protein thin films towards biomaterial based optical devices

NASA Astrophysics Data System (ADS)

Bucciarelli, A.; Mulloni, V.; Maniglio, D.; Pal, R. K.; Yadavalli, V. K.; Motta, A.; Quaranta, A.

2018-04-01

Over the last two decades, silk fibroin has been exploited as a versatile optical material in biological applications due to a combination of unique properties. Recently, protocols have been developed to produce a silk fibroin negative tone resist that is UV crosslinkable, thereby allowing micro and nanoscale patterning of the protein using traditional photolithographic tools. The same protocol has been applied to the silk protein sericin to develop a sericin resist. Despite the immense potential of these biomaterials to develop micro optical patterns on silicon and glass surfaces, as well as self-standing components, their refractive indexes are not well characterized. In this work, optimizing a method to obtain extremely smooth, thin films, the refractive index (RI) of fibroin and sericin proteins and resists were characterized using ellipsometry. The parameters of the Sellmeier and Cauchy dispersion laws have been determined to obtain the RI over a large wavelength range. A complete morphological study of the films has been conducted. In addition, the effect of solvent on the optical properties of silk fibroin and sericin thin films are reported, with differences in values explained by examining the change in the protein secondary structure.

Silk fibroin nanostructured materials for biomedical applications

NASA Astrophysics Data System (ADS)

Mitropoulos, Alexander N.

Nanostructured biopolymers have proven to be promising to develop novel biomedical applications where forming structures at the nanoscale normally occurs by self-assembly. However, synthesizing these structures can also occur by inducing materials to transition into other forms by adding chemical cross-linkers, changing pH, or changing ionic composition. Understanding the generation of nanostructures in fluid environments, such as liquid organic solvents or supercritical fluids, has not been thoroughly examined, particularly those that are based on protein-based block-copolymers. Here, we examine the transformation of reconstituted silk fibroin, which has emerged as a promising biopolymer due to its biocompatibility, biodegradability, and ease of functionalization, into submicron spheres and gel networks which offer applications in tissue engineering and advanced sensors. Two types of gel networks, hydrogels and aerogels, have small pores and large surface areas that are defined by their structure. We design and analyze silk nanoparticle formation using a microfluidic device while offering an application for drug delivery. Additionally, we provide a model and characterize hydrogel formation from micelles to nanoparticles, while investigating cellular response to the hydrogel in an in vitro cell culture model. Lastly, we provide a second model of nanofiber formation during near-critical and supercritical drying and characterize the silk fibroin properties at different drying pressures which, when acting as a stabilizing matrix, shows to improve the activity of entrapped enzymes dried at different pressures. This work has created new nanostructured silk fibroin forms to benefit biomedical applications that could be applied to other fibrous proteins.

Environmentally friendly surface modification of silk fiber: Chitosan grafting and dyeing

NASA Astrophysics Data System (ADS)

Davarpanah, Saideh; Mahmoodi, Niyaz Mohammad; Arami, Mokhtar; Bahrami, Hajir; Mazaheri, Firoozmehr

2009-01-01

In this paper, the surface modification of silk fiber using anhydrides to graft the polysaccharide chitosan and dyeing ability of the grafted silk were studied. Silk fiber was degummed and acylated with two anhydrides, succinic anhydride (SA) and phthalic anhydride (PA), in different solvents (dimethyl sulfoxide (DMSO) and N, N-dimethyl formamide (DMF)). The effects of anhydrides, solvents, anhydride concentration, liquor ratio (L:R) and reaction time on acylation of silk were studied. The polysaccharide chitosan was grafted to the acylated silk fiber and dyed by acid dye (Acid Black NB.B). The effects of pH, chitosan concentration, and reaction time on chitosan grafting of acylated silk were investigated. The physical properties show sensible changes regardless of weight gain. Scanning electron microscopy (SEM) analysis showed the presence of foreign materials firmly attached to the surface of silk. FTIR spectroscopy provided evidence that chitosan was grafted onto the acylated silk through the formation of new covalent bonds. The dyeing of the chitosan grafted-acylated silk fiber indicated the higher dye ability in comparison to the acylated and degummed silk samples. The mechanism of chitosan grafting over degummed silk through anhydride linkage was proposed. The findings of this research support the potential production of new environmentally friendly textile fibers. It is worthwhile to mention that the grafted samples have antibacterial potential due to the antibacterial property of chitosan molecules.

Cell proliferation by silk gut incorporating FGF-2 protein microcrystals.

PubMed

Kotani, Eiji; Yamamoto, Naoto; Kobayashi, Isao; Uchino, Keiro; Muto, Sayaka; Ijiri, Hiroshi; Shimabukuro, Junji; Tamura, Toshiki; Sezutsu, Hideki; Mori, Hajime

2015-06-08

Silk gut processed from the silk glands of the silkworm could be an ideal biodegradable carrier for cell growth factors. We previously demonstrated that polyhedra, microcrystals of Cypovirus 1 polyhedrin, can serve as versatile carrier proteins. Here, we report the generation of a transgenic silkworm that expresses polyhedrin together with human basic fibroblast growth factor (FGF-2) in its posterior silk glands to utilize silk gut as a proteinaceous carrier to protect and slowly release active cell growth factors. In the posterior silk glands, polyhedrin formed polyhedral microcrystals, and FGF-2 became encapsulated within the polyhedra due to a polyhedron-immobilization signal. Silk gut powder prepared from posterior silk glands containing polyhedron-encapsulated FGF-2 stimulated the phosphorylation of p44/p42 MAP kinase and induced the proliferation of serum-starved NIH3T3 cells by releasing bioactive FGF-2. Even after a one-week incubation at 25 °C, significantly higher biological activity of FGF-2 was observed for silk gut powder incorporating polyhedron-encapsulated FGF-2 relative to silk gut powder with non-encapsulated FGF-2. Our results demonstrate that posterior silk glands incorporating polyhedron-encapsulated FGF-2 are applicable to the preparation of biodegradable silk gut, which can protect and release FGF-2 that is produced in a virus- and serum-free expression system with significant application potential.

A novel marine silk.

PubMed

Kronenberger, Katrin; Dicko, Cedric; Vollrath, Fritz

2012-01-01

The discovery of a novel silk production system in a marine amphipod provides insights into the wider potential of natural silks. The tube-building corophioid amphipod Crassicorophium bonellii produces from its legs fibrous, adhesive underwater threads that combine barnacle cement biology with aspects of spider silk thread extrusion spinning. We characterised the filamentous silk as a mixture of mucopolysaccharides and protein deriving from glands representing two distinct types. The carbohydrate and protein silk secretion is dominated by complex β-sheet structures and a high content of charged amino acid residues. The filamentous secretion product exits the gland through a pore near the tip of the secretory leg after having moved through a duct, which subdivides into several small ductules all terminating in a spindle-shaped chamber. This chamber communicates with the exterior and may be considered the silk reservoir and processing/mixing space, in which the silk is mechanically and potentially chemically altered and becomes fibrous. We assert that further study of this probably independently evolved, marine arthropod silk processing and secretion system can provide not only important insights into the more complex arachnid and insect silks but also into crustacean adhesion cements.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Okada, Shoko; Weisman, Sarah; Trueman, Holly E.

Aposthonia gurneyi, an Australian webspinner species, is a primitive insect that constructs and lives in a silken tunnel which screens it from the attentions of predators. The insect spins silk threads from many tiny spines on its forelegs to weave a filmy sheet. We found that the webspinner silk fibers have a mean diameter of only 65 nm, an order of magnitude smaller than any previously reported insect silk. The purpose of such fine silk may be to reduce the metabolic cost of building the extensive tunnels. At the molecular level, the A. gurneyi silk has a predominantly beta-sheet proteinmore » structure. The most abundant clone in a cDNA library produced from the webspinner silk glands encoded a protein with extensive glycine-serine repeat regions. The GSGSGS repeat motif of the A. gurneyi silk protein is similar to the well-known GAGAGS repeat motif found in the heavy fibroin of silkworm silk, which also has beta-sheet structure. As the webspinner silk gene is unrelated to the silk gene of the phylogenetically distant silkworm, this is a striking example of convergent evolution.« less

Exploring the mechanism of microarteriogenesis in porous silk fibroin film.

PubMed

Bai, Lun; Wang, Guangqian; Tan, Xiaoyan; Xu, Jianmei

2012-01-01

Purpose. Based on the experiment of the microarteriogenesis that is associated with angiogenesis during tissue repair process in porous silk fibroin films (PSFFs), we investigate the characteristics of micro-arteriogenesis and explore its mechanism. Methods. After the porous silk fibroin materials are implanted into the back hypodermal tissue of SD rats, the arteriole development and the morphogenesis of smooth muscle cell are histologically monitored and the micro-arteriogenesis is quantitatively analyzed. Results. 10 days after implantation, the arteriole density reaches the highest level in the junction of silk fibroin materials with tissues. Three weeks later, the arteriolar density in the materials reaches the maximum, and the arterioles in the junction of materials with tissues appear to be in a mature and upgrading state. Modeling of Microarteriogenesis. The arterioles in materials are generated after capillary angiogenesis. It is inferred that arteriolar development does not start until the network of the capillaries is formed. At first, the arterioles grow in the conjunct area of precapillaries with arterioles. Then with the extension of the arterioles, the upgrade of arterioles in connecting area is observed at a later stage. Based on the observation, the conditions and the mechanism of microarterializations as well as the upgrade of arterioles are analyzed.

DOE Office of Scientific and Technical Information (OSTI.GOV)

Cebe, Peggy; Partlow, Benjamin P.; Kaplan, David L.

Using fast scanning calorimetry (FSC), we investigated the glass transition and crystal melting of samples of B. mori silk fibroin containing Silk I and/or Silk II crystals. Due to the very short residence times at high temperatures during such measurements, thermal decomposition of silk protein can be significantly suppressed. FSC was performed at 2000 K/s using the Mettler Flash DSC1 on fibroin films with masses around 130–270 ng. Films were prepared with different crystalline fractions (ranging from 0.26 to 0.50) and with different crystal structures (Silk I, Silk II, or mixed) by varying the processing conditions. These included water annealingmore » at different temperatures, exposure to 50% MeOH in water, or autoclaving. The resulting crystal structure was examined using wide angle X-ray scattering. Degree of crystallinity was evaluated from Fourier transform infrared (FTIR) spectroscopy and from analysis of the heat capacity increment at the glass transition temperature. Silk fibroin films prepared by water annealing at 25 °C were the least crystalline and had Silk I structure. FTIR and FSC studies showed that films prepared by autoclaving or 50% MeOH exposure were the most crystalline and had Silk II structure. Intermediate crystalline fraction and mixed Silk I/Silk II structures were found in films prepared by water annealing at 37 °C. FSC results indicate that Silk II crystals exhibit endotherms of narrower width and have higher mean melting temperature Tm(II) = 351 ± 2.6 °C, compared to Silk I crystals which melt at Tm(I) = 292 ± 3.8 °C. Films containing mixed Silk I/Silk II structure showed two clearly separated endothermic peaks. Evidence suggests that the two types of crystals melt separately and do not thermally interconvert on the extremely short time scale (0.065 s between onset and end of melting) of the FSC experiment.« less

Variation in Protein Intake Induces Variation in Spider Silk Expression

PubMed Central

Blamires, Sean J.; Wu, Chun-Lin; Tso, I-Min

2012-01-01

Background It is energetically expensive to synthesize certain amino acids. The proteins (spidroins) of spider major ampullate (MA) silk, MaSp1 and MaSp2, differ in amino acid composition. Glutamine and proline are prevalent in MaSp2 and are expensive to synthesize. Since most orb web spiders express high proline silk they might preferentially attain the amino acids needed for silk from food and shift toward expressing more MaSp1 in their MA silk when starved. Methodology/Principal Findings We fed three spiders; Argiope aetherea, Cyrtophora moluccensis and Leucauge blanda, high protein, low protein or no protein solutions. A. aetherea and L. blanda MA silks are high in proline, while C. moluccesnsis MA silks are low in proline. After 10 days of feeding we determined the amino acid compositions and mechanical properties of each species' MA silk and compared them between species and treatments with pre-treatment samples, accounting for ancestry. We found that the proline and glutamine of A. aetherea and L. blanda silks were affected by protein intake; significantly decreasing under the low and no protein intake treatments. Glutmaine composition in C. moluccensis silk was likewise affected by protein intake. However, the composition of proline in their MA silk was not significantly affected by protein intake. Conclusions Our results suggest that protein limitation induces a shift toward different silk proteins with lower glutamine and/or proline content. Contradictions to the MaSp model lie in the findings that C. moluccensis MA silks did not experience a significant reduction in proline and A. aetherea did not experience a significant reduction in serine on low/no protein. The mechanical properties of the silks could not be explained by a MaSp1 expressional shift. Factors other than MaSp expression, such as the expression of spidroin-like orthologues, may impact on silk amino acid composition and spinning and glandular processes may impact mechanics. PMID:22363691

Design and Research of Service Platform for Protection and Dissemination of Cultural Heritage Resources of The Silk Road in the Territory of China

NASA Astrophysics Data System (ADS)

Zhang, L.; Zhang, W.; Zeng, S. J.; Na, W.; Yang, H.; Huang, J.; Tan, X. D.; Sun, Z. J.

2015-08-01

The Silk Road, a major traffic route across the Eurasia continent, has been a convergence for the exchange, communication and dissemination of various cultures such as nations, materials, religions and arts for more than two thousand years. And the cultural heritage along the long and complicate route has been also attractive. In recent years, the Silk Road - the Road Network along the Chang'an-Tianshan Mountain has been listed in the Directory of World Cultural Heritage. The rare and rich cultural resources along the Silk Road, especially those in the territory of China, have attracted attentions of the world. This article describes the research ideas, methods, processes and results of the planning design on the internet-based dissemination services platform system for cultural heritage resources. First of all, it has defined the targeting for dissemination services and the research methods applied for the Silk Road heritage resources, based on scientific and objective spatial measurement and research on history and geography, to carry on the excavation of values of cultural resource for the target users. Then, with the front-end art exhibit by means of innovative IT, time and space maps of cultural heritage resources, interactive graphics display, panoramic three-dimensional virtual tour, and the Silk Road topics as the main features, a comprehensive and multi-angle cultural resources dissemination services platform is built. The research core of the platform is a demand-oriented system design on the basis of cultural resources and features as the fundamental, the value of contemporary manifestation as the foundation, and cultural dissemination and service as a starting point. This platform has achieved, temporal context generalization, interest profiles extension, online and offline adaptation, and other prominent innovations. On the basis of routes heritage resource protection and dissemination services with complex relationship between time and space, and the Silk Road as the representative, practice and research of the platform in the internet context help to provide an application reference and theoretical basis.

Effect of polyamines on mechanical and structural properties of Bombyx mori silk.

PubMed

Yerra, Aparna; Mysarla, Danti Kumari; Siripurapu, Prasanthi; Jha, Anjali; Valluri, Satyavathi V; Mamillapalli, Anitha

2017-01-01

Silkworm, Bombyx mori (B. mori) belongs to the Lepidoptera family. The silk produced from this insect, mulberry silk, gained lot of importance as a fabric. Silk is being exploited as a biomaterial due to its surprising strength and biocompatibility. Polyamines (PA) are important cell growth regulators. In the present work the effect of treatment of polyamines, putrescine (Put), spermidine (Spd), and spermine (Spm) on the quantity and quality of silk produced was assessed. Results showed that exogenous feeding of Spd at a concentration of 50 µM increased fiber length significantly. Analysis by Fourier transform infrared (FTIR) on the properties of silk obtained from Spd treated silkworms revealed an increase in percentage of absorption with no difference in peak positions of amide I and amide III groups. Scanning electron microscopy (SEM) revealed an increase in diameter of silk. Further, analysis at molecular level showed an increase in fibroin expression in Spd treated silk glands. However, the Spd treatment showed no significant difference with respect to fibroin to sericin ratio per unit weight of cocoon, silk tenacity, and percent elongation. Thus, the present results show that polyamine treatment would influence silk quality at structural, mechanical, and molecular level in the Bombyx mori, which can be exploited in silk biomaterial production. © 2016 Wiley Periodicals, Inc.

Formation of silk fibroin nanoparticles in water-miscible organic solvent and their characterization

NASA Astrophysics Data System (ADS)

Zhang, Yu-Qing; Shen, Wei-De; Xiang, Ru-Li; Zhuge, Lan-Jian; Gao, Wei-Jian; Wang, Wen-Bao

2007-10-01

When Silk fibre derived from Bombyx mori, a native biopolymer, was dissolved in highly concentrated neutral salts such as CaCl2, the regenerated liquid silk, a gradually degraded peptide mixture of silk fibroin, could be obtained. The silk fibroin nanoparticles were prepared rapidly from the liquid silk by using water-miscible protonic and polar aprotonic organic solvents. The nanoparticles are insoluble but well dispersed and stable in aqueous solution and are globular particles with a range of 35-125 nm in diameter by means of TEM, SEM, AFM and laser sizer. Over one half of the ɛ-amino groups exist around the protein nanoparticles by using a trinitrobenzenesulfonic acid (TNBS) method. Raman spectra shows the tyrosine residues on the surface of the globules are more exposed than those on native silk fibers. The crystalline polymorph and conformation transition of the silk nanoparticles from random-coil and α-helix form (Silk I) into anti-parallel β-sheet form (Silk II) are investigated in detail by using infrared, fluorescence and Raman spectroscopy, DSC, 13C CP-MAS NMR and electron diffraction. X-ray diffraction of the silk nanoparticles shows that the nanoparticles crystallinity is about four fifths of the native fiber. Our results indicate that the degraded peptide chains of the regenerated silk is gathered homogeneously or heterogeneously to form a looser globular structure in aqueous solution. When introduced into excessive organic solvent, the looser globules of the liquid silk are rapidly dispersed and simultaneously dehydrated internally and externally, resulting in the further chain-chain contact, arrangement of those hydrophobic domains inside the globules and final formation of crystalline silk nanoparticles with β-sheet configuration. The morphology and size of the nanoparticles are relative to the kinds, properties and even molecular structures of organic solvents, and more significantly to the looser globular substructure of the degraded silk fibroin in aqueous solution. It is possible that the silk protein nanoparticles are potentially useful in biomaterials such as cosmetics, anti-UV skincare products, industrial materials and surface improving materials, especially in enzyme/drug delivery system as vehicle.

Effect of silk protein surfactant on silk degumming and its properties.

PubMed

Wang, Fei; Cao, Ting-Ting; Zhang, Yu-Qing

2015-10-01

The silk protein surfactant (SPS) first used as a silk degumming agent in this study is an amino acid-type anionic surfactant that was synthesized using silk fibroin amino acids and lauroyl chloride. We studied it systematically in comparison with the traditional degumming methods such as sodium carbonate (Na2CO3) and neutral soap (NS). The experimental results showed that the sericin can be completely removed from the silk fibroin fiber after boiling the fibers three times for 30 min and using a bath ratio of 1:80 (g/mL) and a concentration of 0.2% SPS in an aqueous solution. The results of the tensile properties, thermal analysis, and SEM all show that SPS is similar to the NS, far superior to Na2CO3. In short, SPS may be used as an environmentally friendly silk degumming/refining agent in the silk textile industry and in the manufacture of silk floss quilts. Copyright © 2015 Elsevier B.V. All rights reserved.

Silk protein aggregation kinetics revealed by Rheo-IR.

PubMed

Boulet-Audet, Maxime; Terry, Ann E; Vollrath, Fritz; Holland, Chris

2014-02-01

The remarkable mechanical properties of silk fibres stem from a multi-scale hierarchical structure created when an aqueous protein "melt" is converted to an insoluble solid via flow. To directly relate a silk protein's structure and function in response to flow, we present the first application of a Rheo-IR platform, which couples cone and plate rheology with attenuated total reflectance infrared spectroscopy. This technique provides a new window into silk processing by linking shear thinning to an increase in molecular alignment, with shear thickening affecting changes in the silk protein's secondary structure. Additionally, compared to other static characterization methods for silk, Rheo-IR proved particularly useful at revealing the intrinsic difference between natural (native) and reconstituted silk feedstocks. Hence Rheo-IR offers important novel insights into natural silk processing. This has intrinsic academic merit, but it might also be useful when designing reconstituted silk analogues alongside other polymeric systems, whether natural or synthetic. Copyright © 2013 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

Expression of a maize Myb transcription factor driven by a putative silk-specific promoter significantly enhances resistance to Helicoverpa zea in transgenic maize.

PubMed

Johnson, Eric T; Berhow, Mark A; Dowd, Patrick F

2007-04-18

Hi II maize (Zea mays) plants were engineered to express maize p1 cDNA, a Myb transcription factor, controlled by a putative silk specific promoter, for secondary metabolite production and corn earworm resistance. Transgene expression did not enhance silk color, but about half of the transformed plant silks displayed browning when cut, which indicated the presence of p1-produced secondary metabolites. Levels of maysin, a secondary metabolite with insect toxicity, were highest in newly emerged browning silks. The insect resistance of transgenic silks was also highest at emergence, regardless of maysin levels, which suggests that other unidentified p1-induced molecules likely contributed to larval mortality. Mean survivor weights of corn earworm larvae fed mature browning transgenic silks were significantly lower than weights of those fed mature nonbrowning transgenic silks. Some transgenic pericarps browned with drying and contained similar molecules found in pericarps expressing a dominant p1 allele, suggesting that the promoter may not be silk-specific.

Invited review the coiled coil silk of bees, ants, and hornets.

PubMed

Sutherland, Tara D; Weisman, Sarah; Walker, Andrew A; Mudie, Stephen T

2012-06-01

In this article, we review current knowledge about the silk produced by the larvae of bees, ants, and hornets [Apoidea and Vespoidea: Hymenoptera]. Different species use the silk either alone or in composites for a variety of purposes including mechanical reinforcement, thermal regulation, or humidification. The characteristic molecular structure of this silk is α-helical proteins assembled into tetrameric coiled coils. Gene sequences from seven species are available, and each species possesses a copy of each of four related silk genes that encode proteins predicted to form coiled coils. The proteins are ordered at multiple length scales within the labial gland of the final larval instar before spinning. The insects control the morphology of the silk during spinning to produce either fibers or sheets. The silk proteins are small and non repetitive and have been produced artificially at high levels by fermentation in E. coli. The artificial silk proteins can be fabricated into materials with structural and mechanical properties similar to those of native silks. Copyright © 2011 Wiley Periodicals, Inc.

Highly water-absorbing silk yarn with interpenetrating network via in situ polymerization.

PubMed

Lee, Ka I; Wang, Xiaowen; Guo, Xia; Yung, Ka-Fu; Fei, Bin

2017-02-01

Silk was modified via in situ polymerization of two monomers acrylamide and sodium acrylate by swelling in an effective LiBr dissolution system. Swelling of natural silks in LiBr solutions of low concentration was clearly observed under optical microscope, and their conformational changes were revealed by X-ray diffraction (XRD) and Fourier transform infrared (FTIR) spectroscopy. Dissolution tests and FTIR spectra of these modified silks suggested the presence of interpenetrating network of polyacrylamide and poly(sodium acrylate) in the silk yarns. These modified silks exhibited superior water absorption to that of raw silk and greatly improved mechanical properties in both dry and wet states. These novel modified silks also showed low cytotoxicity towards skin keratinocytes, having potential applications in biomedical textiles. This modification method by in situ polymerization after swelling in LiBr provides a new route to highly enhance the properties and performance of silk for various applications. Copyright © 2016 Elsevier B.V. All rights reserved.

Comparative transcriptome analyses on silk glands of six silkmoths imply the genetic basis of silk structure and coloration.

PubMed

Dong, Yang; Dai, Fangyin; Ren, Yandong; Liu, Hui; Chen, Lei; Yang, Pengcheng; Liu, Yanqun; Li, Xin; Wang, Wen; Xiang, Hui

2015-03-17

Silk has numerous unique properties that make it a staple of textile manufacturing for several thousand years. However, wider applications of silk in modern have been stalled due to limitations of traditional silk produced by Bombyx mori. While silk is commonly produced by B. mori, several wild non-mulberry silkmoths--especially members of family Saturniidae--produce silk with superior properties that may be useful for wider applications. Further utilization of such silks is hampered by the non-domestication status or limited culturing population of wild silkworms. To date there is insufficient basic genomic or transcriptomic data on these organisms or their silk production. We sequenced and compared the transcriptomes of silk glands of six Saturniidae wild silkmoth species through next-generation sequencing technology, identifying 37758 ~ 51734 silkmoth unigenes, at least 36.3% of which are annotated with an e-value less than 10(-5). Sequence analyses of these unigenes identified a batch of genes specific to Saturniidae that are enriched in growth and development. Analyses of silk proteins including fibroin and sericin indicate intra-genus conservation and inter-genus diversification of silk protein features among the wild silkmoths, e.g., isoelectric points, hydrophilicity profile and amino acid composition in motifs of silk H-fibroin. Interestingly, we identified p25 in two of the silkmoths, which were previously predicted to be absent in Saturniidae. There are rapid evolutionary changes in sericin proteins, which might account for the highly heterogeneity of sericin in Saturniidae silkmoths. Within the six sikmoths, both colored-cocoon silkmoth specific transcripts and differentially expressed genes between the colored-cocoon and non-colored-cocoon silkmoths are significantly enriched in catalytic activity, especially transferase activity, suggesting potentially viable targets for future gene mining or genetic manipulation. Our results characterize novel and potentially valuable gene resources of saturniid silkmoths that may facilitate future genetic improvement and modification of mulberry silkworms. Our results suggest that the disparate features of silk--coloration, retention, strength, etc. --are likely not only due to silk proteins, but also to the environment of silk assembly, and more specifically, that stable silk coloration exhibited by some Saturniidae silkmoths may be attributable to active catalytic progress in pigmentation.

Lyophilized Silk Fibroin Hydrogels for the Sustained Local Delivery of Therapeutic Monoclonal Antibodies

PubMed Central

Guziewicz, Nicholas; Best, Annie; Perez-Ramirez, Bernardo; Kaplan, David L.

2011-01-01

The development of sustained delivery systems compatible with protein therapeutics continues to be a significant unmet need. A lyophilized silk fibroin hydrogel matrix (lyogel) for the sustained release of pharmaceutically relevant monoclonal antibodies is described. Sonication of silk fibroin prior to antibody incorporation avoids exposing the antibody to the sol-gel transition inducing shear stress. Fourier Transform Infrared (FTIR) analysis showed no change in silk structural composition between hydrogel and lyogel or with increasing silk fibroin concentration. Antibody release from hydrogels occurred rapidly over 10 days regardless of silk concentration. Upon lyophilization, sustained antibody release was observed over 38 days from lyogels containing 6.2% (w/w) silk fibroin and above. In 3.2% (w/w) silk lyogels, antibody release was comparable to hydrogels. Swelling properties of lyogels followed a similar threshold behavior. Lyogels at 3.2% (w/w) silk recovered approximately 90% of their fluid mass upon rehydration, while approximately 50% fluid recovery was observed at 6.2% (w/w) silk and above. Antibody release was primarily governed by hydrophobic/hydrophilic silk-antibody interactions and secondarily altered by the hydration resistance of the lyogel. Hydration resistance was controlled by altering β-sheet (crystalline) density of the matrix. The antibody released from lyogels maintained biological activity. Silk lyogels offer an advantage as a delivery matrix over other hydrogel materials for the slow release of the loaded protein, making lyogels suitable for long-term sustained release applications. PMID:21216004

Spider Transcriptomes Identify Ancient Large-Scale Gene Duplication Event Potentially Important in Silk Gland Evolution.

PubMed

Clarke, Thomas H; Garb, Jessica E; Hayashi, Cheryl Y; Arensburger, Peter; Ayoub, Nadia A

2015-06-08

The evolution of specialized tissues with novel functions, such as the silk synthesizing glands in spiders, is likely an influential driver of adaptive success. Large-scale gene duplication events and subsequent paralog divergence are thought to be required for generating evolutionary novelty. Such an event has been proposed for spiders, but not tested. We de novo assembled transcriptomes from three cobweb weaving spider species. Based on phylogenetic analyses of gene families with representatives from each of the three species, we found numerous duplication events indicative of a whole genome or segmental duplication. We estimated the age of the gene duplications relative to several speciation events within spiders and arachnids and found that the duplications likely occurred after the divergence of scorpions (order Scorpionida) and spiders (order Araneae), but before the divergence of the spider suborders Mygalomorphae and Araneomorphae, near the evolutionary origin of spider silk glands. Transcripts that are expressed exclusively or primarily within black widow silk glands are more likely to have a paralog descended from the ancient duplication event and have elevated amino acid replacement rates compared with other transcripts. Thus, an ancient large-scale gene duplication event within the spider lineage was likely an important source of molecular novelty during the evolution of silk gland-specific expression. This duplication event may have provided genetic material for subsequent silk gland diversification in the true spiders (Araneomorphae). © The Author(s) 2015. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.

Aqueous multiphoton lithography with multifunctional silk-centred bio-resists.

PubMed

Sun, Yun-Lu; Li, Qi; Sun, Si-Ming; Huang, Jing-Chun; Zheng, Bo-Yuan; Chen, Qi-Dai; Shao, Zheng-Zhong; Sun, Hong-Bo

2015-10-16

Silk and silk fibroin, the biomaterial from nature, nowadays are being widely utilized in many cutting-edge micro/nanodevices/systems via advanced micro/nanofabrication techniques. Herein, for the first time to our knowledge, we report aqueous multiphoton lithography of diversiform-regenerated-silk-fibroin-centric inks using noncontact and maskless femtosecond laser direct writing (FsLDW). Initially, silk fibroin was FsLDW-crosslinked into arbitrary two/three-dimensional micro/nanostructures with good elastic properties merely using proper photosensitizers. More interestingly, silk/metal composite micro/nanodevices with multidimension-controllable metal content can be FsLDW-customized through laser-induced simultaneous fibroin oxidation/crosslinking and metal photoreduction using the simplest silk/Ag(+) or silk/[AuCl4](-) aqueous resists. Noticeably, during FsLDW, fibroin functions as biological reductant and matrix, while metal ions act as the oxidant. A FsLDW-fabricated prototyping silk/Ag microelectrode exhibited 10(4)-Ω(-1 ) m(-1)-scale adjustable electric conductivity. This work not only provides a powerful development to silk micro/nanoprocessing techniques but also creates a novel way to fabricate multifunctional metal/biomacromolecule complex micro/nanodevices for applications such as micro/nanoscale mechanical and electrical bioengineering and biosystems.

Comparative proteomics reveal diverse functions and dynamic changes of Bombyx mori silk proteins spun from different development stages.

PubMed

Dong, Zhaoming; Zhao, Ping; Wang, Chen; Zhang, Yan; Chen, Jianping; Wang, Xin; Lin, Ying; Xia, Qingyou

2013-11-01

Silkworms (Bombyx mori) produce massive amounts of silk proteins to make cocoons during the final stages of larval development. Although the major components, fibroin and sericin, have been the focus for a long time, few researchers have realized the complexity of the silk proteome. We collected seven kinds of silk fibers spun by silkworm larvae at different developmental stages: the silks spun by new hatched larvae, second instar day 0 larvae, third instar day 0 larvae, fourth instar day 0 larvae, and fourth instar molting larvae, the scaffold silk used to attach the cocoon to the substrate and the cocoon silk. Analysis by liquid chromatography-tandem mass spectrometry identified 500 proteins from the seven silks. In addition to the expected fibroins, sericins, and some known protease inhibitors, we also identified further protease inhibitors, enzymes, proteins of unknown function, and other proteins. Unsurprisingly, our quantitative results showed fibroins and sericins were the most abundant proteins in all seven silks. Except for fibroins and sericins, protease inhibitors, enzymes, and proteins of unknown function were more abundant than other proteins. We found significant change in silk protein compositions through development, being consistent with their different biological functions and complicated formation.

Aqueous multiphoton lithography with multifunctional silk-centred bio-resists

PubMed Central

Sun, Yun-Lu; Li, Qi; Sun, Si-Ming; Huang, Jing-Chun; Zheng, Bo-Yuan; Chen, Qi-Dai; Shao, Zheng-Zhong; Sun, Hong-Bo

2015-01-01

Silk and silk fibroin, the biomaterial from nature, nowadays are being widely utilized in many cutting-edge micro/nanodevices/systems via advanced micro/nanofabrication techniques. Herein, for the first time to our knowledge, we report aqueous multiphoton lithography of diversiform-regenerated-silk-fibroin-centric inks using noncontact and maskless femtosecond laser direct writing (FsLDW). Initially, silk fibroin was FsLDW-crosslinked into arbitrary two/three-dimensional micro/nanostructures with good elastic properties merely using proper photosensitizers. More interestingly, silk/metal composite micro/nanodevices with multidimension-controllable metal content can be FsLDW-customized through laser-induced simultaneous fibroin oxidation/crosslinking and metal photoreduction using the simplest silk/Ag+ or silk/[AuCl4]− aqueous resists. Noticeably, during FsLDW, fibroin functions as biological reductant and matrix, while metal ions act as the oxidant. A FsLDW-fabricated prototyping silk/Ag microelectrode exhibited 104-Ω−1 m−1-scale adjustable electric conductivity. This work not only provides a powerful development to silk micro/nanoprocessing techniques but also creates a novel way to fabricate multifunctional metal/biomacromolecule complex micro/nanodevices for applications such as micro/nanoscale mechanical and electrical bioengineering and biosystems. PMID:26472600

Metal nanoparticles triggered persistent negative photoconductivity in silk protein hydrogels

NASA Astrophysics Data System (ADS)

Gogurla, Narendar; Sinha, Arun K.; Naskar, Deboki; Kundu, Subhas C.; Ray, Samit K.

2016-03-01

Silk protein is a natural biopolymer with intriguing properties, which are attractive for next generation bio-integrated electronic and photonic devices. Here, we demonstrate the negative photoconductive response of Bombyx mori silk protein fibroin hydrogels, triggered by Au nanoparticles. The room temperature electrical conductivity of Au-silk hydrogels is found to be enhanced with the incorporation of Au nanoparticles over the control sample, due to the increased charge transporting networks within the hydrogel. Au-silk lateral photoconductor devices show a unique negative photoconductive response under an illumination of 325 nm, with excitation energy higher than the characteristic metal plasmon resonance band. The enhanced photoconductance yield in the hydrogels over the silk protein is attributed to the photo-oxidation of amino groups in the β-pleated sheets of the silk around the Au nanoparticles followed by the breaking of charge transport networks. The Au-silk nanocomposite does not show any photoresponse under visible illumination because of the localization of excited charges in Au nanoparticles. The negative photoconductive response of hybrid Au-silk under UV illumination may pave the way towards the utilization of silk for future bio-photonic devices using metal nanoparticle platforms.

Aqueous multiphoton lithography with multifunctional silk-centred bio-resists

NASA Astrophysics Data System (ADS)

Sun, Yun-Lu; Li, Qi; Sun, Si-Ming; Huang, Jing-Chun; Zheng, Bo-Yuan; Chen, Qi-Dai; Shao, Zheng-Zhong; Sun, Hong-Bo

2015-10-01

Silk and silk fibroin, the biomaterial from nature, nowadays are being widely utilized in many cutting-edge micro/nanodevices/systems via advanced micro/nanofabrication techniques. Herein, for the first time to our knowledge, we report aqueous multiphoton lithography of diversiform-regenerated-silk-fibroin-centric inks using noncontact and maskless femtosecond laser direct writing (FsLDW). Initially, silk fibroin was FsLDW-crosslinked into arbitrary two/three-dimensional micro/nanostructures with good elastic properties merely using proper photosensitizers. More interestingly, silk/metal composite micro/nanodevices with multidimension-controllable metal content can be FsLDW-customized through laser-induced simultaneous fibroin oxidation/crosslinking and metal photoreduction using the simplest silk/Ag+ or silk/[AuCl4]- aqueous resists. Noticeably, during FsLDW, fibroin functions as biological reductant and matrix, while metal ions act as the oxidant. A FsLDW-fabricated prototyping silk/Ag microelectrode exhibited 104-Ω-1 m-1-scale adjustable electric conductivity. This work not only provides a powerful development to silk micro/nanoprocessing techniques but also creates a novel way to fabricate multifunctional metal/biomacromolecule complex micro/nanodevices for applications such as micro/nanoscale mechanical and electrical bioengineering and biosystems.

The effect of sterilization methods on the physical properties of silk sericin scaffolds.

PubMed

Siritientong, Tippawan; Srichana, Teerapol; Aramwit, Pornanong

2011-06-01

Protein-based biomaterials respond differently to sterilization methods. Since protein is a complex structure, heat, or irradiation may result in the loss of its physical or biological properties. Recent investigations have shown that sericin, a degumming silk protein, can be successfully formed into a 3-D scaffolds after mixing with other polymers which can be applied in skin tissue engineering. The objective of this study was to investigate the effectiveness of ethanol, ethylene oxide (EtO) and gamma irradiation on the sterilization of sericin scaffolds. The influence of these sterilization methods on the physical properties such as pore size, scaffold dimensions, swelling and mechanical properties, as well as the amount of sericin released from sericin/polyvinyl alcohol/glycerin scaffolds, were also investigated. Ethanol treatment was ineffective for sericin scaffold sterilization whereas gamma irradiation was the most effective technique for scaffold sterilization. Moreover, ethanol also caused significant changes in pore size resulting from shrinkage of the scaffold. Gamma-irradiated samples exhibited the highest swelling property, but they also lost the greatest amount of weight after immersion for 24 h compared with scaffolds obtained from other sterilization methods. The results of the maximum stress test and Young's modulus showed that gamma-irradiated and ethanol-treated scaffolds are more flexible than the EtO-treated and untreated scaffolds. The amount of sericin released, which was related to its collagen promoting effect, was highest from the gamma-irradiated scaffold. The results of this study indicate that gamma irradiation should have the greatest potential for sterilizing sericin scaffolds for skin tissue engineering.

MicroRNA profile of silk gland reveals different silk yields of three silkworm strains.

PubMed

Qin, Sheng; Danso, Blessing; Zhang, Jing; Li, Juan; Liu, Na; Sun, Xia; Hou, Chengxiang; Luo, Heng; Chen, Keping; Zhang, Guozheng; Li, Muwang

2018-05-05

Silk proteins are synthesized and secreted by the silk gland. The differential gene expression in it leads to different silk yield among various silkworm strains. As crucial factors, microRNAs (miRNAs) regulate protein synthesis at post-transcriptional level in silk gland. MiRNAs expression level in the silk gland of three silkworm strains (Jingsong, Lan10 and Dazao) was analyzed and 33 differentially expressed miRNAs (DEMs) were discovered between JingSong (JS) and Lan10 (L10), 60 DEMs between JS and Dazao, 54 DEMs between L10 and Dazao respectively. The DEMs target genes were predicted combing with two different methods and their functions were annotated according to gene ontology. Our previous studies showed that a batch of genes related to silk yield were identified in JS and L10 strains by comparative transcriptome and quantitative trait loci (QTL) method. Thirteen DEMs whose target genes are related to protein biosynthesis processes were screened by combining with these researches. Twelve DEMs potentially regulate nineteen genes which exist in our QTL results. Six common DEMs potentially regulate the genes in both of previous results. Finally, five DEMs were selected to verify their expression levels between JS and L10 by qRT-PCR, which showed similar difference as the results of small RNA-sequencing. MiRNAs in the silk gland may directly affect silk protein biosynthesis in different silkworm strains. In current work, we identified a batch of DEMs which potentially regulate the genes related to silk yield. Further functionally study of these miRNAs will contribute to improve varieties and boost the silk yield. Our research provides a basis for studying these miRNAs and their functions in silk production. Copyright © 2018 Elsevier B.V. All rights reserved.

Programming function into mechanical forms by directed assembly of silk bulk materials

PubMed Central

Patel, Nereus; Duggan, Thomas; Perotto, Giovanni; Shirman, Elijah; Li, Chunmei; Kaplan, David L.; Omenetto, Fiorenzo G.

2017-01-01

We report simple, water-based fabrication methods based on protein self-assembly to generate 3D silk fibroin bulk materials that can be easily hybridized with water-soluble molecules to obtain multiple solid formats with predesigned functions. Controlling self-assembly leads to robust, machinable formats that exhibit thermoplastic behavior consenting material reshaping at the nanoscale, microscale, and macroscale. We illustrate the versatility of the approach by realizing demonstrator devices where large silk monoliths can be generated, polished, and reshaped into functional mechanical components that can be nanopatterned, embed optical function, heated on demand in response to infrared light, or can visualize mechanical failure through colorimetric chemistries embedded in the assembled (bulk) protein matrix. Finally, we show an enzyme-loaded solid mechanical part, illustrating the ability to incorporate biological function within the bulk material with possible utility for sustained release in robust, programmably shapeable mechanical formats. PMID:28028213

Bioengineered silk scaffolds in 3D tissue modeling with focus on mammary tissues.

PubMed

Maghdouri-White, Yas; Bowlin, Gary L; Lemmon, Christopher A; Dréau, Didier

2016-02-01

In vitro generation of three-dimensional (3D) biological tissues and organ-like structures is a promising strategy to study and closely model complex aspects of the molecular, cellular, and physiological interactions of tissue. In particular, in vitro 3D tissue modeling holds promises to further our understanding of breast development. Indeed, biologically relevant 3D structures that combine mammary cells and engineered matrices have improved our knowledge of mammary tissue growth, organization, and differentiation. Several polymeric biomaterials have been used as scaffolds to engineer 3D mammary tissues. Among those, silk fibroin-based biomaterials have many biologically relevant properties and have been successfully used in multiple medical applications. Here, we review the recent advances in engineered scaffolds with an emphasis on breast-like tissue generation and the benefits of modified silk-based scaffolds. Copyright © 2015 Elsevier B.V. All rights reserved.

Silk Gland Gene Expression during Larval-Pupal Transition in the Cotton Leaf Roller Sylepta derogata (Lepidoptera: Pyralidae)

PubMed Central

Su, Honghua; Cheng, Yuming; Wang, Zhongyang; Li, Zhong; Stanley, David; Yang, Yizhong

2015-01-01

The cotton leaf roller, Sylepta derogata, is a silk-producing insect pest. While young larvae feed on the underside of leaves, the older ones roll cotton leaves and feed on the leaf edges, which defoliates cotton plants. The larvae produce silk to stabilize the rolled leaf and to balloon from used to new leaves. Despite the significance of silk in the biology of pest insect species, there is virtually no information on the genes involved in their silk production. This is a substantial knowledge gap because some of these genes may be valuable targets for developing molecular pest management technologies. We addressed the gap by posing the hypothesis that silk gland gene expression changes during the transition from larvae to pupae. We tested our hypothesis using RNA-seq to investigate changes in silk gland gene expression at three developmental stages, 5th instar larvae (silk producing; 15,445,926 clean reads), prepupae (reduced silk producing; 13,758,154) and pupae (beyond silk producing; 16,787,792). We recorded 60,298 unigenes and mapped 50,158 (larvae), 48,415 (prepupae) and 46,623 (pupae) of them to the NCBI database. Most differentially expressed genes in the 5th instar larvae/prepupae libraries were relevant to nucleotide synthesis and maintenance of silk gland function. We identified down-regulated transcriptional factors and several genes involved in silk formation in the three libraries and verified the expression pattern of eight genes by qPCR. The developmental- and tissue-specific expression patterns of the fibroin light chain gene showed it was highly expressed during the larval silk-producing stage. We recorded highest expression of this gene in the larval silk gland, compared to other tissues, including midgut, hindgut, epidermis, Malpighian tubes, hemolymph and fat body. These data are a genetic resource to guide selection of key genes that may be targeted for in planta and other gene-silencing technologies for sustainable cotton agriculture. PMID:26352931

Silk Gland Gene Expression during Larval-Pupal Transition in the Cotton Leaf Roller Sylepta derogata (Lepidoptera: Pyralidae).

PubMed

Su, Honghua; Cheng, Yuming; Wang, Zhongyang; Li, Zhong; Stanley, David; Yang, Yizhong

2015-01-01

The cotton leaf roller, Sylepta derogata, is a silk-producing insect pest. While young larvae feed on the underside of leaves, the older ones roll cotton leaves and feed on the leaf edges, which defoliates cotton plants. The larvae produce silk to stabilize the rolled leaf and to balloon from used to new leaves. Despite the significance of silk in the biology of pest insect species, there is virtually no information on the genes involved in their silk production. This is a substantial knowledge gap because some of these genes may be valuable targets for developing molecular pest management technologies. We addressed the gap by posing the hypothesis that silk gland gene expression changes during the transition from larvae to pupae. We tested our hypothesis using RNA-seq to investigate changes in silk gland gene expression at three developmental stages, 5th instar larvae (silk producing; 15,445,926 clean reads), prepupae (reduced silk producing; 13,758,154) and pupae (beyond silk producing; 16,787,792). We recorded 60,298 unigenes and mapped 50,158 (larvae), 48,415 (prepupae) and 46,623 (pupae) of them to the NCBI database. Most differentially expressed genes in the 5th instar larvae/prepupae libraries were relevant to nucleotide synthesis and maintenance of silk gland function. We identified down-regulated transcriptional factors and several genes involved in silk formation in the three libraries and verified the expression pattern of eight genes by qPCR. The developmental- and tissue-specific expression patterns of the fibroin light chain gene showed it was highly expressed during the larval silk-producing stage. We recorded highest expression of this gene in the larval silk gland, compared to other tissues, including midgut, hindgut, epidermis, Malpighian tubes, hemolymph and fat body. These data are a genetic resource to guide selection of key genes that may be targeted for in planta and other gene-silencing technologies for sustainable cotton agriculture.

Duplication and concerted evolution of MiSp-encoding genes underlie the material properties of minor ampullate silks of cobweb weaving spiders.

PubMed

Vienneau-Hathaway, Jannelle M; Brassfield, Elizabeth R; Lane, Amanda Kelly; Collin, Matthew A; Correa-Garhwal, Sandra M; Clarke, Thomas H; Schwager, Evelyn E; Garb, Jessica E; Hayashi, Cheryl Y; Ayoub, Nadia A

2017-03-14

Orb-web weaving spiders and their relatives use multiple types of task-specific silks. The majority of spider silk studies have focused on the ultra-tough dragline silk synthesized in major ampullate glands, but other silk types have impressive material properties. For instance, minor ampullate silks of orb-web weaving spiders are as tough as draglines, due to their higher extensibility despite lower strength. Differences in material properties between silk types result from differences in their component proteins, particularly members of the spidroin (spider fibroin) gene family. However, the extent to which variation in material properties within a single silk type can be explained by variation in spidroin sequences is unknown. Here, we compare the minor ampullate spidroins (MiSp) of orb-weavers and cobweb weavers. Orb-web weavers use minor ampullate silk to form the auxiliary spiral of the orb-web while cobweb weavers use it to wrap prey, suggesting that selection pressures on minor ampullate spidroins (MiSp) may differ between the two groups. We report complete or nearly complete MiSp sequences from five cobweb weaving spider species and measure material properties of minor ampullate silks in a subset of these species. We also compare MiSp sequences and silk properties of our cobweb weavers to published data for orb-web weavers. We demonstrate that all our cobweb weavers possess multiple MiSp loci and that one locus is more highly expressed in at least two species. We also find that the proportion of β-spiral-forming amino acid motifs in MiSp positively correlates with minor ampullate silk extensibility across orb-web and cobweb weavers. MiSp sequences vary dramatically within and among spider species, and have likely been subject to multiple rounds of gene duplication and concerted evolution, which have contributed to the diverse material properties of minor ampullate silks. Our sequences also provide templates for recombinant silk proteins with tailored properties.

Interactive Story Development for the Unit of Turks on the Silk Road in Social Sciences Course

ERIC Educational Resources Information Center

Karamete, Aysen; Topraklioglu, Kivanç

2017-01-01

With this study, creating interactive story that includes interaction factors was purposed in order to support teaching of the unit of Turks on the Silk Road in Social Sciences course of 6th grades. The research method was defined as Design and Development Research and ADDIE pattern that is one of the teaching design pattern was based while…

Strontium isotope evidence for a highly mobile population on the Pamir Plateau 2500 years ago

NASA Astrophysics Data System (ADS)

Wang, Xueye; Tang, Zihua; Wu, Jing; Wu, Xinhua; Wu, Yiqun; Zhou, Xinying

2016-10-01

Archeological researches have proposed arguments for human mobility and long-distance trading over the Eurasia before the Silk Roads. Here we utilize biologically available strontium isotope analysis to assess the extent of pre-Silk Road population movements and cultural communications across the Asian interior. From an early Iron Age cemetery (ca. 2500 yr B.P.) on the eastern Pamir Plateau, mean 87Sr/86Sr ratios from 34 individuals display considerable isotopic variability, and 10 individuals are distinguished as migrants based on the local strontium isotope range of 0.710296-0.710572 defined by 12 ovicaprine bones. Comparison of the proportion (10/34) with the regional census data completed in 1909 A.D. (3% non-locals) suggests a highly migratory behavior on the plateau 2500 years ago. Furthermore, exotic mortuary objects, such as silk fabrics from eastern China and angular harp originated from the Near East, clearly demonstrate an interaction between different cultures on the plateau before the establishment of the Silk Road.

Effect of gamma radiation on the mechanical properties of natural silk fiber and synthetic E-glass fiber reinforced polypropylene composites: A comparative study

NASA Astrophysics Data System (ADS)

Shubhra, Quazi T. H.; Alam, A. K. M. M.

2011-11-01

Silk is a strong natural proteinous fiber and E-glass is a very strong synthetic fiber. Compression molding method was used to fabricate B. mori silk fiber reinforced polypropylene (PP) matrix composites. The tensile strength (TS), tensile modulus (TM), bending strength (BS), bending modulus (BM) and impact strength (IS) of prepared composites were 55.1 MPa, 780 MPa, 56.3 MPa, 3450 MPa and 17 kJ/m 2, respectively. Synthetic E-glass fiber reinforced PP based composites were fabricated in the same way and TS, TM, BS, BM, IS of E-glass fiber reinforced polypropylene composites were found to be 128.7 MPa, 4350 MPa, 141.6 MPa, 6300 MPa and 19 kJ/m 2, respectively. Gamma radiation is high energy ionizing radiation and was applied to increase the mechanical properties of the composites. Application of gamma ray increases the mechanical properties of silk/PP composites to a greater extent than that of E-glass/PP composites.

Silk-based blood stabilization for diagnostics.

PubMed

Kluge, Jonathan A; Li, Adrian B; Kahn, Brooke T; Michaud, Dominique S; Omenetto, Fiorenzo G; Kaplan, David L

2016-05-24

Advanced personalized medical diagnostics depend on the availability of high-quality biological samples. These are typically biofluids, such as blood, saliva, or urine; and their collection and storage is critical to obtain reliable results. Without proper temperature regulation, protein biomarkers in particular can degrade rapidly in blood samples, an effect that ultimately compromises the quality and reliability of laboratory tests. Here, we present the use of silk fibroin as a solid matrix to encapsulate blood analytes, protecting them from thermally induced damage that could be encountered during nonrefrigerated transportation or freeze-thaw cycles. Blood samples are recovered by simple dissolution of the silk matrix in water. This process is demonstrated to be compatible with a number of immunoassays and provides enhanced sample preservation in comparison with traditional air-drying paper approaches. Additional processing can remediate interactions with conformational structures of the silk protein to further enhance blood stabilization and recovery. This approach can provide expanded utility for remote collection of blood and other biospecimens empowering new modalities of temperature-independent remote diagnostics.

Activation of the Ubiquitin Proteasome Pathway by Silk Fibroin Modified Chitosan Nanoparticles in Hepatic Cancer Cells

PubMed Central

Yang, Ming-Hui; Chung, Tze-Wen; Lu, Yi-Shan; Chen, Yi-Ling; Tsai, Wan-Chi; Jong, Shiang-Bin; Yuan, Shyng-Shiou; Liao, Pao-Chi; Lin, Po-Chiao; Tyan, Yu-Chang

2015-01-01

Silk fibroin (SF) is a protein with bulky hydrophobic domains and can be easily purified as sericin-free silk-based biomaterial. Silk fibroin modified chitosan nanoparticle (SF-CSNP), a biocompatible material, has been widely used as a potential drug delivery system. Our current investigation studied the bio-effects of the SF-CSNP uptake by liver cells. In this experiment, the characterizations of SF-CSNPs were measured by particle size analysis and protein assay. The average size of the SF-CSNP was 311.9 ± 10.7 nm, and the average zeta potential was +13.33 ± 0.3 mV. The SF coating on the SF-CSNP was 6.27 ± 0.17 μg/mL. Moreover, using proteomic approaches, several proteins involved in the ubiquitin proteasome pathway were identified by analysis of differential protein expressions of HepG2 cell uptake the SF-CSNP. Our experimental results have demonstrated that the SF-CSNP may be involved in liver cancer cell survival and proliferation. PMID:25588218

Use of the Stable Nitrogen Isotope to Reveal the Source-Sink Regulation of Nitrogen Uptake and Remobilization during Grain Filling Phase in Maize

PubMed Central

Yang, Lan; Guo, Song; Chen, Qinwu; Chen, Fanjun; Yuan, Lixing; Mi, Guohua

2016-01-01

Although the remobilization of vegetative nitrogen (N) and post-silking N both contribute to grain N in maize (Zea mays L.), their regulation by grain sink strength is poorly understood. Here we use 15N labeling to analyze the dynamic behaviors of both pre- and post-silking N in relation to source and sink manipulation in maize plants. The results showed that the remobilization of pre-silking N started immediately after silking and the remobilized pre-silking N had a greater contribution to grain N during early grain filling, with post-silking N importance increasing during the later filling stage. The amount of post-silking N uptake was largely driven by post-silking dry matter accumulation in both grain as well as vegetative organs. Prevention of pollination during silking had less effect on post-silking N uptake, as a consequence of compensatory growth of stems, husk + cob and roots. Also, leaves continuously export N even though grain sink was removed. The remobilization efficiency of N in the leaf and stem increased with increasing grain yield (hence N requirement). It is suggested that the remobilization of N in the leaf is controlled by sink strength but not the leaf per se. Enhancing post-silking N uptake rather than N remobilization is more likely to increase grain N accumulation. PMID:27606628

Lyophilized Silk Sponges: A Versatile Biomaterial Platform for Soft Tissue Engineering

PubMed Central

2015-01-01

We present a silk biomaterial platform with highly tunable mechanical and degradation properties for engineering and regeneration of soft tissues such as, skin, adipose, and neural tissue, with elasticity properties in the kilopascal range. Lyophilized silk sponges were prepared under different process conditions and the effect of silk molecular weight, concentration and crystallinity on 3D scaffold formation, structural integrity, morphology, mechanical and degradation properties, and cell interactions in vitro and in vivo were studied. Tuning the molecular weight distribution (via degumming time) of silk allowed the formation of stable, highly porous, 3D scaffolds that held form with silk concentrations as low as 0.5% wt/v. Mechanical properties were a function of silk concentration and scaffold degradation was driven by beta-sheet content. Lyophilized silk sponges supported the adhesion of mesenchymal stem cells throughout 3D scaffolds, cell proliferation in vitro, and cell infiltration and scaffold remodeling when implanted subcutaneously in vivo. PMID:25984573

Mechanical Properties of Transgenic Silkworm Silk Under High Strain Rate Tensile Loading

NASA Astrophysics Data System (ADS)

Chu, J.-M.; Claus, B.; Chen, W.

2017-12-01

Studies have shown that transgenic silkworm silk may be capable of having similar properties of spider silk while being mass-producible. In this research, the tensile stress-strain response of transgenic silkworm silk fiber is systematically characterized using a quasi-static load frame and a tension Kolsky bar over a range of strain-rates between 10^{-3} and 700/s. The results show that transgenic silkworm silk tends to have higher overall ultimate stress and failure strain at high strain rate (700/s) compared to quasi-static strain rates, indicating rate sensitivity of the material. The failure strain at the high strain rate is higher than that of spider silk. However, the stress levels are significantly below that of spider silk, and far below that of high-performance fiber. Failure surfaces are examined via scanning electron microscopy and reveal that the failure modes are similar to those of spider silk.

Rapid Characterization of Spider Silk Genes via Exon Capture

DTIC Science & Technology

2015-03-28

SECURITY CLASSIFICATION OF: Spider silks are high-performance materials with an array of potential military and civilian applications. As such, there...is persistent demand for the mass production of silks, which requires knowledge of the underlying silk gene sequences. Spidroins ( spider fibroins...2015 1-May-2014 31-Jan-2015 Approved for Public Release; Distribution Unlimited Final Report: Rapid Characterization of Spider Silk Genes via Exon

Web building and silk properties functionally covary among species of wolf spider.

PubMed

Lacava, Mariángeles; Camargo, Arley; Garcia, Luis F; Benamú, Marco A; Santana, Martin; Fang, Jian; Wang, Xungai; Blamires, Sean J

2018-04-15

Although phylogenetic studies have shown covariation between the properties of spider major ampullate (MA) silk and web building, both spider webs and silks are highly plastic so we cannot be sure whether these traits functionally covary or just vary across environments that the spiders occupy. As MaSp2-like proteins provide MA silk with greater extensibility, their presence is considered necessary for spider webs to effectively capture prey. Wolf spiders (Lycosidae) are predominantly non-web building, but a select few species build webs. We accordingly collected MA silk from two web-building and six non-web-building species found in semirural ecosystems in Uruguay to test whether the presence of MaSp2-like proteins (indicated by amino acid composition, silk mechanical properties and silk nanostructures) was associated with web building across the group. The web-building and non-web-building species were from disparate subfamilies so we estimated a genetic phylogeny to perform appropriate comparisons. For all of the properties measured, we found differences between web-building and non-web-building species. A phylogenetic regression model confirmed that web building and not phylogenetic inertia influences silk properties. Our study definitively showed an ecological influence over spider silk properties. We expect that the presence of the MaSp2-like proteins and the subsequent nanostructures improves the mechanical performance of silks within the webs. Our study furthers our understanding of spider web and silk co-evolution and the ecological implications of spider silk properties. © 2018 European Society For Evolutionary Biology. Journal of Evolutionary Biology © 2018 European Society For Evolutionary Biology.

Concentration state dependence of the rheological and structural properties of reconstituted silk.

PubMed

Mo, Chunli; Holland, Chris; Porter, David; Shao, Zhengzhong; Vollrath, Fritz

2009-10-12

The ability to control the processing of artificial silk is key to the successful application of this important and high performance biopolymer. Understanding where our current reconstitution process can be improved will not only aid us in the creation of better materials, but will also provide insight into the natural material along the way. This study aims to understand what proportion of reconstituted silk contributes to its rheological properties and what conformational state the silk proteins are in. It shows, for the first time, that a change in rheological properties can be related to a change in silk structures present in solution and reveals a low concentration gel state for silk that may have important implications for future successful artificial processing of silk.

The protective ability of Camellia meal extract on the silk protein

NASA Astrophysics Data System (ADS)

Weng, JZ; Cai, C.; Zhang, DY; Dai, BK

2018-02-01

With the enhancement of living standards, people pay more and more attention to the health. The edible oil become more and more popular, but also produced a large amount of Camellia meal which can not fully put into utilization. In this study, the extracting liquid of Camellia meal was used on the process of silk degumming. Firstly, tussah silk was treated by degumming in the Na2CO3 solution, and the preliminary condition of tussah silk degumming was obtained by orthogonal experiment: the concentration Na2CO3 was 0.1%, the degumming time was 1 hour, and the ratio of silk/water was 40:1. Then the extract of Camellia meal (GCJSY) was added before the bleaching process of tussah silk to investigate the protective ability of GCJSY on the silk protein basry on the residual ratio of the silk. While the concentration of GYJSY was 0.08%, the residual ratio of silk after degumming in the Na2CO3 solution and bleaching in the 2% H2O2 solution was 87.2%.

Structuring of Functional Spider Silk Wires, Coatings, and Sheets by Self-Assembly on Superhydrophobic Pillar Surfaces.

PubMed

Gustafsson, Linnea; Jansson, Ronnie; Hedhammar, My; van der Wijngaart, Wouter

2018-01-01

Spider silk has recently become a material of high interest for a large number of biomedical applications. Previous work on structuring of silk has resulted in particles (0D), fibers (1D), films (2D), and foams, gels, capsules, or microspheres (3D). However, the manufacturing process of these structures is complex and involves posttreatment of chemicals unsuitable for biological applications. In this work, the self-assembly of recombinant spider silk on micropatterned superhydrophobic surfaces is studied. For the first time, structuring of recombinant spider silk is achieved using superhydrophobic surfaces under conditions that retain the bioactivity of the functionalized silk. By tuning the superhydrophobic surface geometry and the silk solution handling parameters, this approach allows controlled generation of silk coatings, nanowires, and sheets. The underlying mechanisms and governing parameters are discussed. It is believed that the results of this work pave the way for fabrication of silk formations for applications including vehicles for drug delivery, optical sensing, antimicrobial coatings, and cell culture scaffolds. © 2017 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

Biocompatibility of a Sonicated Silk Gel for Cervical Injection During Pregnancy

PubMed Central

Critchfield, Agatha S.; Mccabe, Reid; Klebanov, Nikolai; Richey, Lauren; Socrate, Simona; Norwitz, Errol R.; Kaplan, David L.; House, Michael

2014-01-01

Objective: To evaluate the biocompatibility of silk gel for cervical injection. Study Design: Silk gel was injected into the cervix of pregnant rats on day 13 (n = 11) and harvested at day 17. Histology of silk gel was compared with suture controls. Also, human cervical fibroblasts were cultured on silk gel and tissue culture plastic (TCP) in vitro. Cell viability, proliferation, metabolic activity, gene expression (COL1A1, COL3A1, and COX2), and release of proinflammatory mediators (interleukin [IL] 6 and IL-8) were evaluated. Results: In vivo, a mild foreign body response was seen surrounding the silk gel and suture controls. In vitro, cervical fibroblasts were viable, metabolically active, and proliferating at 72 hours. Release of IL-6 and IL-8 was similar on silk gel and TCP. Collagen and COX2 gene expression was similar or slightly decreased compared with TCP. Conclusions: Silk gel was well tolerated in vivo and in vitro, which supports continuing efforts to develop silk gels as an alternative to cervical cerclage. PMID:24520079

Silk fibroin-chondroitin sulfate scaffold with immuno-inhibition property for articular cartilage repair.

PubMed

Zhou, Feifei; Zhang, Xianzhu; Cai, Dandan; Li, Jun; Mu, Qin; Zhang, Wei; Zhu, Shouan; Jiang, Yangzi; Shen, Weiliang; Zhang, Shufang; Ouyang, Hong Wei

2017-11-01

The demand of favorable scaffolds has increased for the emerging cartilage tissue engineering. Chondroitin sulfate (CS) and silk fibroin have been investigated and reported with safety and excellent biocompatibility as tissue engineering scaffolds. However, the rapid degradation rate of pure CS scaffolds presents a challenge to effectively recreate neo-tissue similar to natural articular cartilage. Meanwhile the silk fibroin is well used as a structural constituent material because its remarkable mechanical properties, long-lasting in vivo stability and hypoimmunity. The application of composite silk fibroin and CS scaffolds for joint cartilage repair has not been well studied. Here we report that the combination of silk fibroin and CS could synergistically promote articular cartilage defect repair. The silk fibroin (silk) and silk fibroin/CS (silk-CS) scaffolds were fabricated with salt-leaching, freeze-drying and crosslinking methodologies. The biocompatibility of the scaffolds was investigated in vitro by cell adhesion, proliferation and migration with human articular chondrocytes. We found that silk-CS scaffold maintained better chondrocyte phenotype than silk scaffold; moreover, the silk-CS scaffolds reduced chondrocyte inflammatory response that was induced by interleukin (IL)-1β, which is in consistent with the well-documented anti-inflammatory activities of CS. The in vivo cartilage repair was evaluated with a rabbit osteochondral defect model. Silk-CS scaffold induced more neo-tissue formation and better structural restoration than silk scaffold after 6 and 12weeks of implantation in ICRS histological evaluations. In conclusion, we have developed a silk fibroin/ chondroitin sulfate scaffold for cartilage tissue engineering that exhibits immuno-inhibition property and can improve the self-repair capacity of cartilage. Severe cartilage defect such as osteoarthritis (OA) is difficult to self-repair because of its avascular, aneural and alymphatic nature. Current scaffolds often focus on providing sufficient mechanical support or bio-mimetic structure to promote cartilage repair. Thus, silk has been adopted and investigated broadly. However, inflammation is one of the most important factors in OA. But few scaffolds for cartilage repair reported anti-inflammation property. Meanwhile, chondroitin sulfate (CS) is a glycosaminoglycan present in the natural cartilage ECM, and has exhibited a number of useful biological properties including anti-inflammatory activity. Thus, we designed this silk-CS scaffold and proved that this scaffold exhibited good anti-inflammatory effects both in vitro and in vivo, promoted the repair of articular cartilage defect in animal model. Copyright © 2017 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

A New Preparation Method for Anisotropic Silk Fibroin Nerve Guidance Conduits and Its Evaluation In Vitro and in a Rat Sciatic Nerve Defect Model

PubMed Central

Schuh, Christina; Halbweis, Robert; Pajer, Krisztián; Márton, Gábor; Hopf, Rudolf; Mosia, Shorena; Rünzler, Dominik; Redl, Heinz; Nógrádi, Antal; Hausner, Thomas

2015-01-01

Over the past decade, silk fibroin (SF) has been emergently used in peripheral nerve tissue engineering. Current approaches aiming at producing SF-based nerve guidance conduits (SF-NGCs) used dissolved silk based on either aqueous solutions or organic solvents. In this study, we describe a novel procedure to produce SF-NGCs: A braided tubular structure of raw Bombyx mori silk is subsequently processed with the ternary solvent CaCl2/H2O/ethanol, formic acid, and methanol to improve its mechanical and topographical characteristics. Topographically, the combination of the treatments results in a fusion of the outer single silk fibers to a closed layer with a thickness ranging from about 40 to 75 μm. In contrast to the outer wall, the inner lumen (not treated with processing solvents) still represents the braided structure of single fibers. Mechanical stability, elasticity, and kink characteristics were evaluated with a custom-made test system. The modification procedure described here drastically improved the elastic properties of our tubular raw scaffold, favoring its use as a NGC. A cell migration assay with NIH/3T3-fibroblasts revealed the impermeability of the SF-NGC wall for possible invading and scar-forming cells. Moreover, the potential of the SF-NGC to serve as a substratum for Schwann cells has been demonstrated by cytotoxicity tests and live-dead stainings of Schwann cells grown on the inner surface of the SF-NGC. In vivo, the SF-NGC was tested in a rat sciatic nerve injury model. In short-term in vivo studies, it was proved that SF-NGCs are not triggering host inflammatory reactions. After 12 weeks, we could demonstrate morphological and functional reinnervation of the distal targets. Filled with collagen, a higher number of axons could be found in the distal to the graft (1678±303), compared with the empty SF-NGC (1274±146). The novel SF-NGC presented here shows promising results for the treatment of peripheral nerve injuries. The modification of braided structures to adapt their mechanical and topographical characteristics may support the translation of SF-based scaffolds into the clinical setting. However, further improvements and the use of extracellular matrix molecules and Schwann cells are suggested to enable silk tube based conduits to bridge long-distance nerve gaps. PMID:25819471

Nano/micro hybrid scaffold of PCL or P3HB nanofibers combined with silk fibroin for tendon and ligament tissue engineering.

PubMed

Naghashzargar, Elham; Farè, Silvia; Catto, Valentina; Bertoldi, Serena; Semnani, Dariush; Karbasi, Saeed; Tanzi, Maria Cristina

2015-07-04

A novel biodegradable nano/micro hybrid structure was obtained by electrospinning P3HB or PCL nanofibers onto a twisted silk fibroin (SF) structure, with the aim of fabricating a suitable scaffold for tendon and ligament tissue engineering. The electrospinning (ES) processing parameters for P3HB and PCL were optimized on 2D samples, and applied to produce two different nano/micro hybrid constructs (SF/ES-PCL and SF/ES-P3HB).Morphological, chemico-physical and mechanical properties of the novel hybrid scaffolds were evaluated by SEM, ATR FT-IR, DSC, tensile and thermodynamic mechanical tests. The results demonstrated that the nanofibers were tightly wrapped around the silk filaments, and the crystallinity of the SF twisted yarns was not influenced by the presence of the electrospun polymers. The slightly higher mechanical properties of the hybrid constructs confirmed an increase of internal forces due to the interaction between nano and micro components. Cell culture tests with L929 fibroblasts, in the presence of the sample eluates or in direct contact with the hybrid structures, showed no cytotoxic effects and a good level of cytocompatibility of the nano/micro hybrid structures in term of cell viability, particularly at day 1. Cell viability onto the nano/micro hybrid structures decreased from the first to the third day of culture when compared with the control culture plastic, but appeared to be higher when compared with the uncoated SF yarns. Although additional in vitro and in vivo tests are needed, the original fabrication method here described appears promising for scaffolds suitable for tendon and ligament tissue engineering.

Silk elasticity as a potential constraint on spider body size.

PubMed

Rodríguez-Gironés, Miguel A; Corcobado, Guadalupe; Moya-Laraño, Jordi

2010-10-07

Silk is known for its strength and extensibility and has played a key role in the radiation of spiders. Individual spiders use different glands to produce silk types with unique sets of proteins. Most research has studied the properties of major ampullate and capture spiral silks and their ecological implications, while little is known about minor ampullate silk, the type used by those spider species studied to date for bridging displacements. A biomechanical model parameterised with available data shows that the minimum radius of silk filaments required for efficient bridging grows with the square root of the spider's body mass, faster than the radius of minor ampullate silk filaments actually produced by spiders. Because the morphology of spiders adapted to walking along or under silk threads is ill suited for moving on a solid surface, for these species there is a negative relationship between body mass and displacement ability. As it stands, the model suggests that spiders that use silk for their displacements are prevented from attaining a large body size if they must track their resources in space. In particular, silk elasticity would favour sexual size dimorphism because males that must use bridging lines to search for females cannot grow large. 2010 Elsevier Ltd. All rights reserved.

Investigating the transverse optical structure of spider silk micro-fibers using quantitative optical microscopy

NASA Astrophysics Data System (ADS)

Little, Douglas J.; Kane, Deb M.

2017-01-01

The transverse optical structure of two orb-weaver (family Araneidae) spider dragline silks was investigated using a variant of the inverse-scattering technique. Immersing the silks in a closely refractive index-matched liquid, the minimum achievable image contrast was greater than expected for an optically homogeneous silk, given what is currently known about the optical absorption of these silks. This "excess contrast" indicated the presence of transverse optical structure within the spider silk. Applying electromagnetic scattering theory to a transparent double cylinder, the minimum achievable irradiance contrast for the Plebs eburnus and Argiope keyserlingi dragline silks was determined to be consistent with step index refractive index contrasts of 1-4×10-4 and 6-7×10-4, respectively, supposing outer-layer thicknesses consistent with previous TEM studies (50 nm and 100 nm, respectively). The possibility of graded index refractive index contrasts within the spider silks is also discussed. This is the strongest evidence, to date, that there is a refractive index contrast associated with the layered morphology of spider silks and/or variation of proportion of nanocrystalline components within the spider silk structure. The method is more generally applicable to optical micro-fibers, including those with refractive index variations on a sub-wavelength scale.

Effects of Microwave Radiation on Selected Mechanical Properties of Silk

NASA Astrophysics Data System (ADS)

Reed, Emily Jane

Impressive mechanical properties have served to peak interest in silk as an engineering material. In addition, the ease with which silk can be altered through processing has led to its use in various biomaterial applications. As the uses of silk branch into new territory, it is imperative (and inevitable) to discover the boundary conditions beyond which silk no longer performs as expected. These boundary conditions include factors as familiar as temperature and humidity, but may also include other less familiar contributions, such as exposure to different types of radiation. The inherent variations in mechanical properties of silk, as well as its sensitivity to moisture, suggest that in an engineering context silk is best suited for use in composite materials; that way, silk can be shielded from ambient moisture fluctuations, and the surrounding matrix allows efficient load transfer from weaker fibers to stronger ones. One such application is to use silk as a reinforcing fiber in epoxy composites. When used in this way, there are several instances in which exposure to microwave radiation is likely (for example, as a means of speeding epoxy cure rates), the effects of which remain mostly unstudied. It will be the purpose of this dissertation to determine whether selected mechanical properties of B. mori cocoon silk are affected by exposure to microwave radiation, under specified temperature and humidity conditions. Results of our analyses are directly applicable wherever exposure of silk to microwave radiation is possible, including in fiber reinforced epoxy composites (the entire composite may be microwaved to speed epoxy cure time), or when silk is used as a component in the material used to construct the radome of an aircraft (RADAR units use frequencies in the microwave range of the electromagnetic spectrum), or when microwave energy is used to sterilize biomaterials (such as cell scaffolds) made of silk. In general, we find that microwave exposure does not detract from the average mechanical properties of silk, but that it may increase the spread of data points around that average. Along the way, we come to a number of useful conclusions, summarized here: Regarding silk in general: • Storage conditions can have a significant and enduring effect on tensile properties of degummed B. mori silk. Samples stored in a sealed container with desiccant (silica gel) have a lower yield stress and yield strain than samples stored without desiccant and they also relax more rapidly in stress relaxation tests. The ability of this silk to resist plastic deformation is optimized at intermediate hydration levels. Sensitivity to the humidity levels encountered by samples prior to testing complicates the interpretation of results, and makes inter-laboratory comparisons challenging. Silk storage conditions should therefore be reported---and, ideally, standardized---to enable useful comparison between studies. (Abstract shortened by UMI.)

The impact of environment change on culture evolution in east Ancient Silk Road.

NASA Astrophysics Data System (ADS)

Dong, G.; Liu, F.; Li, G.; Zhang, D. D.; Lee, H. F.; Chen, F.

2017-12-01

Ancient Silk Road played an important role in culture communication between west and east parts of the Eurasia during Bronze Age and historical period. Tens of archaic civilizations rise and fall in east parts of the Ancient Silk Road, climate change is attributed as one of the most important driving forces, while the process and mechanism for the impact of environmental change on culture evolution in the area has not been well-understood. Here we report new paleoclimate data based on multi-proxy analysis from two well-dated aeolian deposit sequences in the Hexi Corridor and Qaidam basin, where locate at the throat position of the Ancient Silk Road. Comparing with high-resolution tree rings from Qilian Mountain nearby, and archaeological evidence and historical documents, we proposed that two desertification events occurred in west Hexi Corridor between 3400-3100 BP and post 1450 AD, which induced two cultural discontinuity in that area. Climate was dry between 3400-2900 BP and wet between 2900-2000 BP in lowlands of east Qaidam basin, mismatching with the development of Nuomuhong Bronze culture in the area during 3400-2450 BP. We propose culture evolution in east Ancient Silk Road was mainly influenced by precipitation change of highlands in mountain areas,which was further influenced by large-scale vapor transport.

Ion Electrodiffusion Governs Silk Electrogelation.

PubMed

Kojic, Nikola; Panzer, Matthew J; Leisk, Gary G; Raja, Waseem K; Kojic, Milos; Kaplan, David L

2012-07-14

Silk electrogelation involves the transition of an aqueous silk fibroin solution to a gel state (E-gel) in the presence of an electric current. The process is based on local pH changes as a result of water electrolysis - generating H(+) and OH(-) ions at the (+) and (-) electrodes, respectively. Silk fibroin has a pI=4.2 and when local pH

Dissolution and regeneration of non-mulberry Eriogyna Pyretorum silk fibroin

NASA Astrophysics Data System (ADS)

Guo, Yuhang; Li, Xiufang; Zhang, Qiang; Yan, Shuqin; You, Renchuan

2017-10-01

Protein-based materials have been actively pursued as biomaterials because of their nontoxicity, biocompatibility and biodegradability. In this work, we demonstrated the potential of Eriogyna pyretorum silk fibroin (ESF), a non-mulberry silk protein, as biomaterials. The degummed ESF fibers could be dissolved completely by Ca(NO3)2/H2O/C2H5OH solution to produce regenerated ESF. The solubility was strongly dependent on the addition of C2H5OH, heating temperature and dissolving time. α-helix and random coil are main molecular conformation in aqueous ESF solution. The sol-gel transition behavior of regenerated ESF was also studied, indicating that the conformational transition of regenerated ESF from random coil/α-helix to β-sheet during gelation. Especially, ESF showed more rapid gelation than mulberry silk fibroin (BSF). Consequently, the gelation rate of BSF could be controlled ranging from tens of minutes to days by changing the ESF ratio, providing useful options for the fabrication of silk hydrogels. Water-stable regenerated ESF film could be achieved by using aqueous ethanol to induce structural transition. Tensile tests showed that the ESF films have a dry strength of approximate 31.0 MPa and a wet strength of approximate 3.3 MPa. This study provides new opportunities as an alternative natural protein material for biomedical applications.

Recursive Directional Ligation Approach for Cloning Recombinant Spider Silks.

PubMed

Dinjaski, Nina; Huang, Wenwen; Kaplan, David L

2018-01-01

Recent advances in genetic engineering have provided a route to produce various types of recombinant spider silks. Different cloning strategies have been applied to achieve this goal (e.g., concatemerization, step-by-step ligation, recursive directional ligation). Here we describe recursive directional ligation as an approach that allows for facile modularity and control over the size of the genetic cassettes. This approach is based on sequential ligation of genetic cassettes (monomers) where the junctions between them are formed without interrupting key gene sequences with additional base pairs.

Invited review current progress and limitations of spider silk for biomedical applications.

PubMed

Widhe, Mona; Johansson, Jan; Hedhammar, My; Rising, Anna

2012-06-01

Spider silk is a fascinating material combining remarkable mechanical properties with low density and biodegradability. Because of these properties and historical descriptions of medical applications, spider silk has been proposed to be the ideal biomaterial. However, overcoming the obstacles to produce spider silk in sufficient quantities and in a manner that meets regulatory demands has proven to be a difficult task. Also, there are relatively few studies of spider silk in biomedical applications available, and the methods and materials used vary a lot. Herein we summarize cell culture- and in vivo implantation studies of natural and synthetic spider silk, and also review the current status and future challenges in the quest for a large scale production of spider silk for medical applications. Copyright © 2011 Wiley Periodicals, Inc.

The effect of native silk fibroin powder on the physical properties and biocompatibility of biomedical polyurethane membrane.

PubMed

Zhuang, Yan; Zhang, Qian; Feng, Jinqi; Wang, Na; Xu, Weilin; Yang, Hongjun

2017-04-01

Naturally derived fibers such as silk fibroin can potentially enhance the biocompatibility of currently used biomaterials. This study investigated the physical properties of native silk fibroin powder and its effect on the biocompatibility of biomedical polyurethane. Native silk fibroin powder with an average diameter of 3 µm was prepared on a purpose-built machine. A simple method of phase inversion was used to produce biomedical polyurethane/native silk fibroin powder hybrid membranes at different blend ratios by immersing a biomedical polyurethane/native silk fibroin powder solution in deionized water at room temperature. The physical properties of the membranes including morphology, hydrophilicity, roughness, porosity, and compressive modulus were characterized, and in vitro biocompatibility was evaluated by seeding the human umbilical vein endothelial cells on the top surface. Native silk fibroin powder had a concentration-dependent effect on the number and morphology of human umbilical vein endothelial cells growing on the membranes; cell number increased as native silk fibroin powder content in the biomedical polyurethane/native silk fibroin powder hybrid membrane was increased from 0% to 50%, and cell morphology changed from spindle-shaped to cobblestone-like as the native silk fibroin powder content was increased from 0% to 70%. The latter change was related to the physical characteristics of the membrane, including hydrophilicity, roughness, and mechanical properties. The in vivo biocompatibility of the native silk fibroin powder-modified biomedical polyurethane membrane was evaluated in a rat model; the histological analysis revealed no systemic toxicity. These results indicate that the biomedical polyurethane/native silk fibroin powder hybrid membrane has superior in vitro and in vivo biocompatibility relative to 100% biomedical polyurethane membranes and thus has potential applications in the fabrication of small-diameter vascular grafts and in tissue engineering.

Exploring the Mechanism of Microarteriogenesis in Porous Silk Fibroin Film

PubMed Central

Bai, Lun; Wang, Guangqian; Tan, Xiaoyan; Xu, Jianmei

2012-01-01

Purpose. Based on the experiment of the microarteriogenesis that is associated with angiogenesis during tissue repair process in porous silk fibroin films (PSFFs), we investigate the characteristics of micro-arteriogenesis and explore its mechanism. Methods. After the porous silk fibroin materials are implanted into the back hypodermal tissue of SD rats, the arteriole development and the morphogenesis of smooth muscle cell are histologically monitored and the micro-arteriogenesis is quantitatively analyzed. Results. 10 days after implantation, the arteriole density reaches the highest level in the junction of silk fibroin materials with tissues. Three weeks later, the arteriolar density in the materials reaches the maximum, and the arterioles in the junction of materials with tissues appear to be in a mature and upgrading state. Modeling of Microarteriogenesis. The arterioles in materials are generated after capillary angiogenesis. It is inferred that arteriolar development does not start until the network of the capillaries is formed. At first, the arterioles grow in the conjunct area of precapillaries with arterioles. Then with the extension of the arterioles, the upgrade of arterioles in connecting area is observed at a later stage. Based on the observation, the conditions and the mechanism of microarterializations as well as the upgrade of arterioles are analyzed. PMID:23304150

Traveling the Silk Road: A Measurement of a Large Anonymous Online Marketplace

DTIC Science & Technology

2012-11-28

Silk Road, an anonymous, international online marketplace that operates as a Tor hidden service and uses Bitcoin as its exchange currency. We gather...analysis of Silk Road, an anonymous, international on- line marketplace that operates as a Tor hidden service and uses Bitcoin as its exchange currency. We...anonymity, Silk Road needs to also preserve payment anonymity. To that effect, Silk Road only supports Bitcoin (BTC, [30]) as a trading currency

An Unlikely Silk: The Composite Material of Green Lacewing Cocoons

DOE Office of Scientific and Technical Information (OSTI.GOV)

Weisman, Sarah; Trueman, Holly E.; Mudie, Stephen T.

2009-01-15

Spiders routinely produce multiple types of silk; however, common wisdom has held that insect species produce one type of silk each. This work reports that the green lacewing (Mallada signata, Neuroptera) produces two distinct classes of silk. We identified and sequenced the gene that encodes the major protein component of the larval lacewing cocoon silk and demonstrated that it is unrelated to the adult lacewing egg-stalk silk. The cocoon silk protein is 49 kDa in size and is alanine rich (>40%), and it contains an {alpha}-helical secondary structure. The final instar lacewing larvae spin protein fibers of {approx}2 {mu}m diametermore » to construct a loosely woven cocoon. In a second stage of cocoon construction, the insects lay down an inner wall of lipids that uses the fibers as a scaffold. We propose that the silk protein fibers provide the mechanical strength of the composite lacewing cocoon whereas the lipid layer provides a barrier to water loss during pupation.« less

De novo design of recombinant spider silk proteins for material applications.

PubMed

Zheng, Ke; Ling, Shengjie

2018-05-21

Spider silks are well known for their superior mechanical properties that are stronger and tougher than steel despite being assembled at close to ambient conditions and using water as the solvent. However, it is a significant challenge to utilize spider silks for practical applications due to their limited sources. Fortunately, genetic engineering techniques offer a promising approach to produce useable amounts of spider silk variants. Starting from these recombinant spider silk proteins, a series of experiments and simulations strategies were developed to improve the recombinant spider silk proteins (RSSP) material design and fabrication with the aim of biomimicking the structure-property-function relationships of spider silks. Accordingly, in this review, we first introduce the structure-property-function relationship of spider silks. Then, we discuss the recent progress in the genetic synthesis of RSSPs and summarize their related multiscale self-assembly behaviors. Finally, we outline works utilizing multiscale modeling to assist RSSP material design. This article is protected by copyright. All rights reserved.

DNA replication events during larval silk gland development in the silkworm, Bombyx mori.

PubMed

Zhang, Chun-Dong; Li, Fang-Fang; Chen, Xiang-Yun; Huang, Mao-Hua; Zhang, Jun; Cui, Hongjuan; Pan, Min-Hui; Lu, Cheng

2012-07-01

The silk gland is an important organ in silkworm as it synthesizes silk proteins and is critical to spinning. The genomic DNA content of silk gland cells dramatically increases 200-400 thousand times for the larval life span through the process of endomitosis. Using in vitro culture, DNA synthesis was measured using BrdU labeling during the larval molt and intermolt periods. We found that the cell cycle of endomitosis was activated during the intermolt and was inhibited during the molt phase. The anterior silk gland, middle silk gland, and posterior silk gland cells asynchronously exit the endomitotic cycle after day 6 in 5th instar larvae, which correlated with the reduced expression of the cell cycle-related cdt1, pcna, cyclin E, cdk2 and cdk1 mRNAs in the wandering phase. Additional starvation had no effect on the initiation of silk gland DNA synthesis of the freshly ecdysed larvae. Copyright © 2012 Elsevier Ltd. All rights reserved.

Effect of microgravity on primordial germ cells (PGCs) in silk chicken offspring ( Gallus gallus domesticus)

NASA Astrophysics Data System (ADS)

Zhou, Zhenming; Li, Zandong

2011-08-01

Primordial germ cells (PGCs), precursors of germline cells, display a variety of antigens during their migration to target gonads. Here, we used silk chicken offspring ( Gallus gallus domesticus) embryos subjected to space microgravity to investigate the influence of microgravity on PGCs. The ShenZhou-3 unmanned spaceship carried nine fertilized silk chicken eggs, named the flight group, returned to Earth after 7 days space flight. And the control group has the same clan with the flight group. PGCs from flight and control group silk chicken offspring embryos were examined during migration by using two antibodies (2C9 and anti-SSEA-1), in combination with the horseradish peroxidase detection system, and using periodic acid-Schiff's solution (PAS) reaction. After incubation for about 30 h, SSEA-1 and 2C9 positive cells were detected in the germinal crescent of flight and control group silk chicken offspring embryos. After incubation of eggs for 2-2.5 days, SSEA-1 and 2C9 positive cells were detected in embryonic blood vessels of flight and control group silk chicken offspring embryos. After incubation of eggs for 5.5 days, PGCs in the dorsal mesentery and gonad could also be identified in flight and control group silk chicken offspring embryos by using SSEA-1 and 2C9 antibodies. Based on location and PAS staining, these cells were identified as PGCs. Meanwhile, at the stage of PGCs migration and then becoming established in the germinal ridges, no difference in SSEA-1 or 2C9 staining was detected between female and male PGCs in flight and control group silk chicken offspring embryos. Although there were differences in the profiles of PGC concentration between male and female embryos during the special circulating stage, changing profile of PGCs concentration was similar in same sex between flight and control group offspring embryos. We concluded that there is little effect on PGCs in offspring embryos of microgravity-treated chicken and that PGC development appears to be normal.

Biopolymer-Based Nanoparticles for Drug/Gene Delivery and Tissue Engineering

PubMed Central

Nitta, Sachiko Kaihara; Numata, Keiji

2013-01-01

There has been a great interest in application of nanoparticles as biomaterials for delivery of therapeutic molecules such as drugs and genes, and for tissue engineering. In particular, biopolymers are suitable materials as nanoparticles for clinical application due to their versatile traits, including biocompatibility, biodegradability and low immunogenicity. Biopolymers are polymers that are produced from living organisms, which are classified in three groups: polysaccharides, proteins and nucleic acids. It is important to control particle size, charge, morphology of surface and release rate of loaded molecules to use biopolymer-based nanoparticles as drug/gene delivery carriers. To obtain a nano-carrier for therapeutic purposes, a variety of materials and preparation process has been attempted. This review focuses on fabrication of biocompatible nanoparticles consisting of biopolymers such as protein (silk, collagen, gelatin, β-casein, zein and albumin), protein-mimicked polypeptides and polysaccharides (chitosan, alginate, pullulan, starch and heparin). The effects of the nature of the materials and the fabrication process on the characteristics of the nanoparticles are described. In addition, their application as delivery carriers of therapeutic drugs and genes and biomaterials for tissue engineering are also reviewed. PMID:23344060

Biomaterial-based regional chemotherapy: Local anticancer drug delivery to enhance chemotherapy and minimize its side-effects.

PubMed

Krukiewicz, Katarzyna; Zak, Jerzy K

2016-05-01

Since the majority of anticancer pharmacological agents affect not only cancer tissue but also normal cells, chemotherapy is usually accompanied with severe side effects. Regional chemotherapy, as the alternative version of conventional treatment, leads to the enhancement of the therapeutic efficiency of anticancer drugs and, simultaneously, reduction of toxic effects to healthy tissues. This paper provides an insight into different approaches of local delivery of chemotherapeutics, such as the injection of anticancer agents directly into tumor tissue, the use of injectable in situ forming drug carriers or injectable platforms in a form of implants. The wide range of biomaterials used as reservoirs of anticancer drugs is described, i.e. poly(ethylene glycol) and its copolymers, polyurethanes, poly(lactic acid) and its copolymers, poly(ɛ-caprolactone), polyanhydrides, chitosan, cellulose, cyclodextrins, silk, conducting polymers, modified titanium surfaces, calcium phosphate based biomaterials, silicone and silica implants, as well as carbon nanotubes and graphene. To emphasize the applicability of regional chemotherapy in cancer treatment, the commercially available products approved by the relevant health agencies are presented. Copyright © 2016 Elsevier B.V. All rights reserved.

Biomedical Applications of Mulberry Silk and its Proteins: A Review

NASA Astrophysics Data System (ADS)

Nivedita, S.; Sivaprasad, V.

2014-04-01

Silk is a natural fibre used mainly for aesthetic purposes. It has also been used for making surgical sutures for centuries. The recent rediscovery of silk's biological properties have led to new areas of research and utilization in cosmetic, health and medical fields. The silk proteins, fibroin and sericin are processed into biomaterials because of bio-compatibility, bio-degradability, excellent mechanical properties, thermo tolerance and UV protective properties. Silk proteins could be obtained as pure liquids and regenerated in different forms suitable for tissue engineering applications. This paper presents some of the biomedical products and biomaterials made from native, degraded and regenerated silk and their fabrication techniques.

Controllable in situ synthesis of silver nanoparticles on multilayered film-coated silk fibers for antibacterial application.

PubMed

Meng, Mei; He, Huawei; Xiao, Jing; Zhao, Ping; Xie, Jiale; Lu, Zhisong

2016-01-01

Layer-by-layer (LbL) assembly is a versatile technique for the preparation of multilayered polymeric films. However, fabrication of LbL polymetic film on silk for the in situ growth of high-density silver nanoparticles (AgNPs) has not been realized. Herein poly(acrylic acid) (PAA)/poly(dimethyldiallylammonium chloride) (PDDA) multilayers are constructed on silk via the LbL approach, subsequently serving as a 3-dimensional matrix for in situ synthesis of AgNPs. After 8 rounds of LbL assembly, the silk is fully covered with a layer of polymeric film. AgNPs with good crystalline structures could be in-situ generated in the silk-coated multilayers and their amount could be tailored by adjusting the bilayer numbers. The as-prepared silk could effectively kill the existing bacteria and inhibit the bacterial growth, demonstrating the antimicrobial activity. Moreover, the release of Ag(+) from the modified silk can last for 120 h, rendering the modified silk sustainable antimicrobial activity. This work may provide a novel method to prepare AgNPs-functionalized antimicrobial silk for potential applications in textile industry. Copyright © 2015 Elsevier Inc. All rights reserved.

Timing of autophagy and apoptosis during posterior silk gland degeneration in Bombyx mori.

PubMed

Montali, Aurora; Romanelli, Davide; Cappellozza, Silvia; Grimaldi, Annalisa; de Eguileor, Magda; Tettamanti, Gianluca

2017-07-01

Over the years, the silkworm, Bombyx mori, has been manipulated by means of chemical and genetic approaches to improve silk production both quantitatively and qualitatively. The silk is produced by the silk gland, which degenerates quickly once the larva has finished spinning the cocoon. Thus, interfering with this degeneration process could help develop new technologies aimed at ameliorating silk yield. To this end, in this work we studied the cell death processes that lead to the demise of the posterior silk gland of B. mori, directing in particular our attention to autophagy and apoptosis. We focused on this portion of the gland because it produces fibroin, the main component of the silk thread. By using multiple markers, we provide a morphological, biochemical and molecular characterization of the apoptotic and autophagic processes and define their timing in this biological setting. Our data demonstrate that the activation of both autophagy and apoptosis is preceded by a transcriptional rise in key regulatory genes. Moreover, while autophagy is maintained active for several days and progressively digests silk gland cells, apoptosis is only switched on at a very late stage of silk gland demise. Copyright © 2017 Elsevier Ltd. All rights reserved.

Proteomic Analysis of Silk Viability in Maize Inbred Lines and Their Corresponding Hybrids

PubMed Central

Wang, Yafei; Zhao, Xiaofeng; Zhang, Fangfang; Tang, Jihua; Fu, Zhiyuan

2015-01-01

A long period of silk viability is critical for a good seed setting rate in maize (Zea mays L.), especially for inbred lines and hybrids with a long interval between anthesis and silking. To explore the molecular mechanism of silk viability and its heterosis, three inbred lines with different silk viability characteristics (Xun928, Lx9801, and Zong3) and their two hybrids (Xun928×Zong3 and Lx9801×Zong3) were analyzed at different developmental stages by a proteomic method. The differentially accumulated proteins were identified by mass spectrometry and classified into metabolism, protein biosynthesis and folding, signal transduction and hormone homeostasis, stress and defense responses, and cellular processes. Proteins involved in nutrient (methionine) and energy (ATP) supply, which support the pollen tube growth in the silk, were important for silk viability and its heterosis. The additive and dominant effects at a single locus, as well as complex epistatic interactions at two or more loci in metabolic pathways, were the primary contributors for mid-parent heterosis of silk viability. Additionally, the proteins involved in the metabolism of anthocyanins, which indirectly negatively regulate local hormone accumulation, were also important for the mid-parent heterosis of silk viability. These results also might imply the developmental dependence of heterosis, because many of the differentially accumulated proteins made distinct contributions to the heterosis of silk viability at specific developmental stages. PMID:26630375

Proteomic Analysis of Silk Viability in Maize Inbred Lines and Their Corresponding Hybrids.

PubMed

Ma, Zhihui; Qin, Yongtian; Wang, Yafei; Zhao, Xiaofeng; Zhang, Fangfang; Tang, Jihua; Fu, Zhiyuan

2015-01-01

A long period of silk viability is critical for a good seed setting rate in maize (Zea mays L.), especially for inbred lines and hybrids with a long interval between anthesis and silking. To explore the molecular mechanism of silk viability and its heterosis, three inbred lines with different silk viability characteristics (Xun928, Lx9801, and Zong3) and their two hybrids (Xun928×Zong3 and Lx9801×Zong3) were analyzed at different developmental stages by a proteomic method. The differentially accumulated proteins were identified by mass spectrometry and classified into metabolism, protein biosynthesis and folding, signal transduction and hormone homeostasis, stress and defense responses, and cellular processes. Proteins involved in nutrient (methionine) and energy (ATP) supply, which support the pollen tube growth in the silk, were important for silk viability and its heterosis. The additive and dominant effects at a single locus, as well as complex epistatic interactions at two or more loci in metabolic pathways, were the primary contributors for mid-parent heterosis of silk viability. Additionally, the proteins involved in the metabolism of anthocyanins, which indirectly negatively regulate local hormone accumulation, were also important for the mid-parent heterosis of silk viability. These results also might imply the developmental dependence of heterosis, because many of the differentially accumulated proteins made distinct contributions to the heterosis of silk viability at specific developmental stages.

Influence of spider silk on refugia preferences of the recluse spiders Loxosceles reclusa and Loxosceles laeta (Araneae: Sicariidae).

PubMed

Vetter, Richard S; Rust, Michael K

2010-06-01

In a previous experimental study, recluse spiders Loxosceles reclusa Gertsch and Mulaik and Loxosceles laeta (Nicolet) (Araneae: Sicariidae) preferred small cardboard refugia covered with conspecific silk compared with never-occupied refugia. Herein, we investigated some factors that might be responsible for this preference using similar cardboard refugia. When the two Loxosceles species were given choices between refugia previously occupied by their own and by the congeneric species, neither showed a species-specific preference; however, each chose refugia coated with conspecific silk rather than those previously inhabited by a distantly related cribellate spider, Metaltella simoni (Keyserling). When L. laeta spiders were offered refugia that were freshly removed from silk donors compared with heated, aged refugia from the same silk donor, older refugia were preferred. Solvent extracts of L. laeta silk were chosen approximately as often as control refugia when a range of solvents (methylene chloride:methanol, water, and hexane) were used. However, when acetone was used on similar silk, there was a statistical preference for the control, indicating that there might be a mildly repellent aspect to acetone-washed silk. Considering the inability to show attraction to chemical aspects of fresh silk, it seems that physical attributes may be more important for selection and that there might be repellency to silk of a recently vacated spider. These findings are discussed in regard to pest management strategies to control recluse spiders.

Preparation and characterization of silk fibroin as a biomaterial with potential for drug delivery

PubMed Central

2012-01-01

Background Degummed silk fibroin from Bombyx mori (silkworm) has potential carrier capabilities for drug delivery in humans; however, the processing methods have yet to be comparatively analyzed to determine the differential effects on the silk protein properties, including crystalline structure and activity. Methods In this study, we treated degummed silk with four kinds of calcium-alcohol solutions, and performed secondary structure measurements and enzyme activity test to distinguish the differences between the regenerated fibroins and degummed silk fibroin. Results Gel electrophoresis analysis revealed that Ca(NO3)2-methanol, Ca(NO3)2-ethanol, or CaCl2-methanol treatments produced more lower molecular weights of silk fibroin than CaCl2-ethanol. X-ray diffraction and Fourier-transform infrared spectroscopy showed that CaCl2-ethanol produced a crystalline structure with more silk I (α-form, type II β-turn), while the other treatments produced more silk II (β-form, anti-parallel β-pleated sheet). Solid-State 13C cross polarization and magic angle spinning-nuclear magnetic resonance measurements suggested that regenerated fibroins from CaCl2-ethanol were nearly identical to degummed silk fibroin, while the other treatments produced fibroins with significantly different chemical shifts. Finally, enzyme activity test indicated that silk fibroins from CaCl2-ethanol had higher activity when linked to a known chemotherapeutic drug, L-asparaginase, than the fibroins from other treatments. Conclusions Collectively, these results suggest that the CaCl2-ethanol processing method produces silk fibroin with biomaterial properties that are appropriate for drug delivery. PMID:22676291

The Widespread Prevalence and Functional Significance of Silk-Like Structural Proteins in Metazoan Biological Materials

PubMed Central

McDougall, Carmel; Woodcroft, Ben J.

2016-01-01

In nature, numerous mechanisms have evolved by which organisms fabricate biological structures with an impressive array of physical characteristics. Some examples of metazoan biological materials include the highly elastic byssal threads by which bivalves attach themselves to rocks, biomineralized structures that form the skeletons of various animals, and spider silks that are renowned for their exceptional strength and elasticity. The remarkable properties of silks, which are perhaps the best studied biological materials, are the result of the highly repetitive, modular, and biased amino acid composition of the proteins that compose them. Interestingly, similar levels of modularity/repetitiveness and similar bias in amino acid compositions have been reported in proteins that are components of structural materials in other organisms, however the exact nature and extent of this similarity, and its functional and evolutionary relevance, is unknown. Here, we investigate this similarity and use sequence features common to silks and other known structural proteins to develop a bioinformatics-based method to identify similar proteins from large-scale transcriptome and whole-genome datasets. We show that a large number of proteins identified using this method have roles in biological material formation throughout the animal kingdom. Despite the similarity in sequence characteristics, most of the silk-like structural proteins (SLSPs) identified in this study appear to have evolved independently and are restricted to a particular animal lineage. Although the exact function of many of these SLSPs is unknown, the apparent independent evolution of proteins with similar sequence characteristics in divergent lineages suggests that these features are important for the assembly of biological materials. The identification of these characteristics enable the generation of testable hypotheses regarding the mechanisms by which these proteins assemble and direct the construction of biological materials with diverse morphologies. The SilkSlider predictor software developed here is available at https://github.com/wwood/SilkSlider. PMID:27415783

Tough silk fibers prepared in air using a biomimetic microfluidic chip.

PubMed

Luo, Jie; Zhang, Lele; Peng, Qingfa; Sun, Mengjie; Zhang, Yaopeng; Shao, Huili; Hu, Xuechao

2014-05-01

Microfluidic chips with single channel were built to mimic the shear and elongation conditions in the spinning apparatus of spider and silkworm. Silk fibers dry-spun from regenerated silk fibroin (RSF) aqueous solution using the chip could be tougher than degummed natural silk. The artificial silk exhibited a breaking strength up to 614 MPa, a breaking elongation up to 27% and a breaking energy of 101 kJ/kg. Copyright © 2014 Elsevier B.V. All rights reserved.

Solubilization of spider silk proteins and its structural analysis using Fourier transform infrared spectroscopy

NASA Astrophysics Data System (ADS)

Osbin, K.; Jayan, Manuel; Bhadrakumari, S.; Predeep, P.

2017-06-01

This study investigates the presence of various amide bands present in different spider silk species, which provides extraordinary physical properties. Three different spider silks were collected from Western Ghats region. The collected spider silks samples belonging to the spider Heteropoda venatoria (species 1), Hersilia savignyi (species 2) and Pholcus phalangioides (species 3). Fourier transform infrared (FTIR) spectra reveals the protein peaks in the amide I, II, and III regions in all the three types of spider silk species.

Development of new smart materials and spinning systems inspired by natural silks and their applications

NASA Astrophysics Data System (ADS)

Cheng, Jie; Lee, Sang-Hoon

2015-12-01

Silks produced by spiders and silkworms are charming natural biological materials with highly optimized hierarchical structures and outstanding physicomechanical properties. The superior performance of silks relies on the integration of a unique protein sequence, a distinctive spinning process, and complex hierarchical structures. Silks have been prepared to form a variety of morphologies and are widely used in diverse applications, for example, in the textile industry, as drug delivery vehicles, and as tissue engineering scaffolds. This review presents an overview of the organization of natural silks, in which chemical and physical functions are optimized, as well as a range of new materials inspired by the desire to mimic natural silk structure and synthesis.

3D Functional Corneal Stromal Tissue Equivalent Based on Corneal Stromal Stem Cells and Multi-Layered Silk Film Architecture.

PubMed

Ghezzi, Chiara E; Marelli, Benedetto; Omenetto, Fiorenzo G; Funderburgh, James L; Kaplan, David L

2017-01-01

The worldwide need for human cornea equivalents continues to grow. Few clinical options are limited to allogenic and synthetic material replacements. We hypothesized that tissue engineered human cornea systems based on mechanically robust, patterned, porous, thin, optically clear silk protein films, in combination with human corneal stromal stem cells (hCSSCs), would generate 3D functional corneal stroma tissue equivalents, in comparison to previously developed 2D approaches. Silk film contact guidance was used to control the alignment and distribution of hCSSCs on RGD-treated single porous silk films, which were then stacked in an orthogonally, multi-layered architecture and cultured for 9 weeks. These systems were compared similar systems generated with human corneal fibroblasts (hCFs). Both cell types were viable and preferentially aligned along the biomaterial patterns for up to 9 weeks in culture. H&E histological sections showed that the systems seeded with the hCSSCs displayed ECM production throughout the entire thickness of the constructs. In addition, the ECM proteins tested positive for keratocyte-specific tissue markers, including keratan sulfate, lumican, and keratocan. The quantification of hCSSC gene expression of keratocyte-tissue markers, including keratocan, lumican, human aldehyde dehydrogenase 3A1 (ALDH3A1), prostaglandin D2 synthase (PTDGS), and pyruvate dehydrogenase kinase, isozyme 4 (PDK4), within the 3D tissue systems demonstrated upregulation when compared to 2D single silk films and to the systems generated with the hCFs. Furthermore, the production of ECM from the hCSSC seeded systems and subsequent remodeling of the initial matrix significantly improved cohesiveness and mechanical performance of the constructs, while maintaining transparency after 9 weeks.

Effect of residual sericin on the structural characteristics and properties of regenerated silk films.

PubMed

Lee, Ji Hye; Song, Dae Woong; Park, Young Hwan; Um, In Chul

2016-08-01

Regenerated silk film has been increasingly attracting the research community's attention for biomedical applications due to its good biocompatibility and excellent cyto-compatibility. However, some limitations regarding its mechanical properties, such as brittleness, have restricted the use of silk films for industrial biomedical applications. In this study, regenerated silk films with different residual sericin content were prepared applying controlled degumming conditions to evaluate the effect of sericin content on the structure and properties of the films generated. When the residual sericin content increased to 0.6%, crystallinity index and breaking strength of silk films were increased. Above this value, these parameters then decreased. A 1.5 fold increase of silk film elongation properties was obtained when incorporating 16% sericin. Regardless of sericin content, all regenerated silk films showed excellent cyto-compatibility, comparable to the one obtained with tissue culture plates. Copyright © 2016 Elsevier B.V. All rights reserved.

Protective effect of silk lutein on ultraviolet B-irradiated human keratinocytes.

PubMed

Pongcharoen, Sutatip; Warnnissorn, Prateep; Leŗtkajornsin, Ongart; Limpeanchob, Nanteetip; Sutheerawattananonda, Manote

2013-01-01

Carotenoids are efficient antioxidants that are of great importance for human health. Lutein and zeaxanthin are carotinoids present in high concentrations in the human retina which are involved in the photoprotection of the human eye. Lutein may also protect the skin from ultraviolet (UV)-induced damage. The present study investigated the protective effect of lutein extracted from yellow silk cocoons of Bombyx mori on human keratinocytes against UVB irradiation. A human keratinocyte cell line and primary human keratinocytes were used to investigate the UVB protection effects of silk lutein and plant lutein. Silk lutein showed no cytotoxicity to keratinocytes. Treatment with silk lutein prior to UVB irradiation enhanced cell viability and cell proliferation, and reduced cell apoptosis. The protective effects of silk lutein may be superior to those of plant lutein. Silk lutein may have a benefit for protection of keratinocytes against UVB-irradiation.

Spider genomes provide insight into composition and evolution of venom and silk

PubMed Central

Sanggaard, Kristian W.; Bechsgaard, Jesper S.; Fang, Xiaodong; Duan, Jinjie; Dyrlund, Thomas F.; Gupta, Vikas; Jiang, Xuanting; Cheng, Ling; Fan, Dingding; Feng, Yue; Han, Lijuan; Huang, Zhiyong; Wu, Zongze; Liao, Li; Settepani, Virginia; Thøgersen, Ida B.; Vanthournout, Bram; Wang, Tobias; Zhu, Yabing; Funch, Peter; Enghild, Jan J.; Schauser, Leif; Andersen, Stig U.; Villesen, Palle; Schierup, Mikkel H; Bilde, Trine; Wang, Jun

2014-01-01

Spiders are ecologically important predators with complex venom and extraordinarily tough silk that enables capture of large prey. Here we present the assembled genome of the social velvet spider and a draft assembly of the tarantula genome that represent two major taxonomic groups of spiders. The spider genomes are large with short exons and long introns, reminiscent of mammalian genomes. Phylogenetic analyses place spiders and ticks as sister groups supporting polyphyly of the Acari. Complex sets of venom and silk genes/proteins are identified. We find that venom genes evolved by sequential duplication, and that the toxic effect of venom is most likely activated by proteases present in the venom. The set of silk genes reveals a highly dynamic gene evolution, new types of silk genes and proteins, and a novel use of aciniform silk. These insights create new opportunities for pharmacological applications of venom and biomaterial applications of silk. PMID:24801114

Adhesion modulation using glue droplet spreading in spider capture silk

PubMed Central

Zhang, Ci; Blackledge, Todd A.

2017-01-01

Orb web spiders use sticky capture spiral silk to retain prey in webs. Capture spiral silk is composed of an axial fibre of flagelliform silk covered with glue droplets that are arranged in a beads-on-a-string morphology that allows multiple droplets to simultaneously extend and resist pull off. Previous studies showed that the adhesion of capture silk is responsive to environmental humidity, increasing up to an optimum humidity that varied among different spider species. The maximum adhesion was hypothesized to occur when the viscoelasticity of the glue optimized contributions from glue spreading and bulk cohesion. In this study, we show how glue droplet shape during peeling contributes significantly to capture silk adhesion. Both overspreading and underspreading of glue droplets reduces adhesion through changes in crack propagation and failure regime. Understanding the mechanism of stimuli-responsive adhesion of spider capture silk will lead to new designs for smarter adhesives. PMID:28490605

Uptake of atmospheric carbon dioxide into silk fiber by silkworms.

PubMed

Magoshi, Jun; Tanaka, Toshihisa; Sasaki, Haruto; Kobayashi, Masatoshi; Magoshi, Yoshiko; Tsuda, Hidetoshi; Becker, Mary A; Inoue, Shun-ichi; Ishimaru, Ken

2003-01-01

The relation between the uptake of atmospheric CO(2) and insect's production of silk fiber has not yet been reported. Here, we provide the first quantitative demonstrations that four species of silkworms (Bombyx mori, Samia cynthia ricini, Antheraea pernyi, and Antheraea yamamai) and a silk-producing spider (Nephila clavata) incorporate atmospheric CO(2) into their silk fibers. The abundance of (13)C incorporated from the environment was determined by mass spectrometry and (13)C NMR measurements. Atmospheric CO(2) was incorporated into the silk fibers in the carbonyl groups of alanine, aspartic acid, serine, and glycine and the C(gamma) of aspartic acid. We show a simple model for the uptake of atmospheric CO(2) by silkworms. These results will demonstrate that silkworm has incorporated atmospheric CO(2) into silk fiber via the TCA cycle; however, the magnitude of uptake into the silk fibers is smaller than that consumed by the photosynthesis in trees and coral reefs.

Synergistic adhesion mechanisms of spider capture silk.

PubMed

Guo, Yang; Chang, Zheng; Guo, Hao-Yuan; Fang, Wei; Li, Qunyang; Zhao, Hong-Ping; Feng, Xi-Qiao; Gao, Huajian

2018-03-01

It is well known that capture silk, the main sticky component of the orb web of a spider, plays an important role in the spider's ability to capture prey via adhesion. However, the detailed mechanism with which the spider achieves its unparalleled high-adhesion performance remains elusive. In this work, we combine experiments and theoretical analysis to investigate the adhesion mechanisms of spider silk. In addition to the widely recognized adhesion effect of the sticky glue, we reveal a synergistic enhancement mechanism due to the elasticity of silk fibres. A balance between silk stiffness, strength and glue stickiness is crucial to endow the silk with superior adhesion, as well as outstanding energy absorption capacity and structural robustness. The revealed mechanisms deepen our understanding of the working principles of spider silk and suggest guidelines for biomimetic designs of spider-inspired adhesion and capture devices. © 2018 The Author(s).

Adhesion modulation using glue droplet spreading in spider capture silk.

PubMed

Amarpuri, Gaurav; Zhang, Ci; Blackledge, Todd A; Dhinojwala, Ali

2017-05-01

Orb web spiders use sticky capture spiral silk to retain prey in webs. Capture spiral silk is composed of an axial fibre of flagelliform silk covered with glue droplets that are arranged in a beads-on-a-string morphology that allows multiple droplets to simultaneously extend and resist pull off. Previous studies showed that the adhesion of capture silk is responsive to environmental humidity, increasing up to an optimum humidity that varied among different spider species. The maximum adhesion was hypothesized to occur when the viscoelasticity of the glue optimized contributions from glue spreading and bulk cohesion. In this study, we show how glue droplet shape during peeling contributes significantly to capture silk adhesion. Both overspreading and underspreading of glue droplets reduces adhesion through changes in crack propagation and failure regime. Understanding the mechanism of stimuli-responsive adhesion of spider capture silk will lead to new designs for smarter adhesives. © 2017 The Author(s).

Size exclusion chromatography for analyses of fibroin in silk: optimization of sampling and separation conditions

NASA Astrophysics Data System (ADS)

Pawcenis, Dominika; Koperska, Monika A.; Milczarek, Jakub M.; Łojewski, Tomasz; Łojewska, Joanna

2014-02-01

A direct goal of this paper was to improve the methods of sample preparation and separation for analyses of fibroin polypeptide with the use of size exclusion chromatography (SEC). The motivation for the study arises from our interest in natural polymers included in historic textile and paper artifacts, and is a logical response to the urgent need for developing rationale-based methods for materials conservation. The first step is to develop a reliable analytical tool which would give insight into fibroin structure and its changes caused by both natural and artificial ageing. To investigate the influence of preparation conditions, two sets of artificially aged samples were prepared (with and without NaCl in sample solution) and measured by the means of SEC with multi angle laser light scattering detector. It was shown that dialysis of fibroin dissolved in LiBr solution allows removal of the salt which destroys stacks chromatographic columns and prevents reproducible analyses. Salt rich (NaCl) water solutions of fibroin improved the quality of chromatograms.

Universal composition-structure-property maps for natural and biomimetic platelet-matrix composites and stacked heterostructures.

PubMed

Sakhavand, Navid; Shahsavari, Rouzbeh

2015-03-16

Many natural and biomimetic platelet-matrix composites--such as nacre, silk, and clay-polymer-exhibit a remarkable balance of strength, toughness and/or stiffness, which call for a universal measure to quantify this outstanding feature given the structure and material characteristics of the constituents. Analogously, there is an urgent need to quantify the mechanics of emerging electronic and photonic systems such as stacked heterostructures. Here we report the development of a unified framework to construct universal composition-structure-property diagrams that decode the interplay between various geometries and inherent material features in both platelet-matrix composites and stacked heterostructures. We study the effects of elastic and elastic-perfectly plastic matrices, overlap offset ratio and the competing mechanisms of platelet versus matrix failures. Validated by several 3D-printed specimens and a wide range of natural and synthetic materials across scales, the proposed universally valid diagrams have important implications for science-based engineering of numerous platelet-matrix composites and stacked heterostructures.

Silk scaffolds with tunable mechanical capability for cell differentiation

PubMed Central

Bai, Shumeng; Han, Hongyan; Huang, Xiaowei; Xu, Weian; Kaplan, David L.; Zhu, Hesun; Lu, Qiang

2015-01-01

Bombyx mori silk fibroin is a promising biomaterial for tissue regeneration and is usually considered an “inert” material with respect to actively regulating cell differentiation due to few specific cell signaling peptide domains in the primary sequence and the generally stiffer mechanical properties due to crystalline content formed in processing. In the present study, silk fibroin porous 3D scaffolds with nanostructures and tunable stiffness were generated via a silk fibroin nanofiber-assisted lyophilization process. The silk fibroin nanofibers with high β-sheet content were added into the silk fibroin solutions to modulate the self-assembly, and to directly induce water-insoluble scaffold formation after lyophilization. Unlike previously reported silk fibroin scaffold formation processes, these new scaffolds had lower overall β-sheet content and softer mechanical properties for improved cell compatibility. The scaffold stiffness could be further tuned to match soft tissue mechanical properties, which resulted in different differentiation outcomes with rat bone marrow-derived mesenchymal stem cells towards myogenic and endothelial cells, respectively. Therefore, these silk fibroin scaffolds regulate cell differentiation outcomes due to their mechanical features. PMID:25858557

Electrophoretic deposition of tetracycline modified silk fibroin coatings for functionalization of titanium surfaces

NASA Astrophysics Data System (ADS)

Zhang, Zhen; Qu, Yinying; Li, Xiaoshuang; Zhang, Sheng; Wei, Qingsong; Shi, Yusheng; Chen, Lili

2014-06-01

Electrophoretic deposition has been widely used for the fabrication of functional coatings onto metal implant. A characteristic feature of this process is that positively charged materials migrate toward the cathode and can deposit on it. In this study, silk fibroin was decorated with tetracycline in aqueous solution to impart positive charge, and then deposited on negatively titanium cathode under certain electric field. The characterization of the obtained coatings indicated that the intermolecular hydrogen bonds formed between the backbone of silk fibroin and tetracycline molecular. In vitro biological tests demonstrated that osteoblast-like cells achieved acceptable cell affinity on the tetracycline cross-linked silk fibroin coatings, although greater cell viability was seen on pure silk fibroin coatings. The cationic silk fibroin coatings showed remarkable antibacterial activity against gram-positive (Staphylococcus aureus) and gram-negative (Escherichia coli) bacteria. Therefore, we concluded that electrophoretic deposition was an effective and efficient technique to prepare cationic silk fibroin coatings on the titanium surface and that cationic silk fibroin coatings with acceptable biocompatibility and antibacterial property were promising candidates for further loading of functional agents.

Effects of silk fibroin in murine dry eye

NASA Astrophysics Data System (ADS)

Kim, Chae Eun; Lee, Ji Hyun; Yeon, Yeung Kyu; Park, Chan Hum; Yang, Jaewook

2017-03-01

The study aimed to investigate the effects of silk fibroin in a mouse model of dry eye. The experimental dry eye mouse model was developed using more than twelve-weeks-old NOD.B10.H2b mice exposing them to 30-40% ambient humidity and injecting them with scopolamine hydrobromide for 10 days. Tear production and corneal irregularity score were measured by the instillation of phosphate buffered saline or silk fibroin. Corneal detachment and conjunctival goblet cell density were observed by hematoxylin and eosin or periodic acid Schiff staining in the cornea or conjunctiva. The expression of inflammatory markers was detected by immunohistochemistry in the lacrimal gland. The silk group tear production was increased, and corneal smoothness was improved. The corneal epithelial cells and conjunctival goblet cells were recovered in the silk groups. The expression of inflammatory factors was inhibited in the lacrimal gland of the silk group. These results show that silk fibroin improved the cornea, conjunctiva, and lacrimal gland in the mouse model of dry eye. These findings suggest that silk fibroin has anti-inflammatory effects in the experimental models of dry eye.

Infrared and Raman Study of the Recluse Spider Silk

NASA Astrophysics Data System (ADS)

Wang, S. L.; Wang, Qijue; Xing, Zhen; Schniepp, H. C.; Qazilbash, M. M.

Spider silk exhibits remarkable mechanical properties, such as high tensile strength and toughness. We want to gain insight into the composition and structure of spider silk to discover the origin of these properties. We are especially interested in the organization of the crystalline beta sheets that are expected to contribute to the high strength of the silk from the recluse spider, Loxosceles laeta. The recluse spider produces a silk that has a unique geometry amongst arachnids. We measure the silk's optical properties, particularly the infrared-active and Raman-active vibrations. Broadband infrared transmission spectra were collected in the spectral range between 600 cm-1 and 4000 cm-1, with light polarized parallel and perpendicular to the long axis of the silk. Raman micro-spectroscopy was performed in the spectral range 500 cm-1 and 4000 cm- 1 with a 514 nm laser. The infrared and Raman vibrational modes are fit with Lorentzian and pseudo-Voigt functions. The vibrational modes are assigned to specific structures and electronic bonds in the silk. This work was supported by NASA/ Virginia Space Grant Consortium.

Characteristics of platelet gels combined with silk

PubMed Central

Pallotta, Isabella; Kluge, Jonathan A.; Moreau, Jodie; Calabrese, Rossella

2014-01-01

Platelet gel, a fibrin network containing activated platelets, is widely used in regenerative medicine due the capacity of platelet-derived growth factors to accelerate and direct healing processes. However, limitations to this approach include poor mechanical properties, relatively rapid degradation, and the lack of control of release of growth factors at the site of injection. These issues compromise the ability of platelet gels for sustained function in regenerative medicine. In the present study, a combination of platelet gels with silk fibroin gel was studied to address the above limitations. Mixing sonicated silk gels with platelet gels extended the release of growth factors without inhibiting gel forming ability. The released growth factors were biologically active and their delivery was modified further by manipulation of the charge of the silk protein. Moreover, the silk gel augmented both the rheological properties and compressive stiffness of the platelet gel, tuned by the silk concentration and/or silk/platelet gel ratio. Silk-platelet gel injections in nude rats supported enhanced cell infiltration and blood vessel formation representing a step towards new platelet gel formulations with enhanced therapeutic impact. PMID:24480538

Optimized extraction of polysaccharides from corn silk by pulsed electric field and response surface quadratic design.

PubMed

Zhao, Wenzhu; Yu, Zhipeng; Liu, Jingbo; Yu, Yiding; Yin, Yongguang; Lin, Songyi; Chen, Feng

2011-09-01

Corn silk is a traditional Chinese herbal medicine, which has been widely used for treatment of some diseases. In this study the effects of pulsed electric field on the extraction of polysaccharides from corn silk were investigated. Polysaccharides in corn silk were extracted by pulsed electric field and optimized by response surface methodology (RSM), based on a Box-Behnken design (BBD). Three independent variables, including electric field intensity (kV cm(-1) ), ratio of liquid to raw material and pulse duration (µs), were investigated. The experimental data were fitted to a second-order polynomial equation and also profiled into the corresponding 3-D contour plots. Optimal extraction conditions were as follows: electric field intensity 30 kV cm(-1) , ratio of liquid to raw material 50, and pulse duration 6 µs. Under these condition, the experimental yield of extracted polysaccharides was 7.31% ± 0.15%, matching well with the predicted value. The results showed that a pulsed electric field could be applied to extract value-added products from foods and/or agricultural matrix. Copyright © 2011 Society of Chemical Industry.

Determination of active ingredients in corn silk, leaf, and kernel by capillary electrophoresis with electrochemicaI detection.

PubMed

Lin, Miao; Chu, Qing-Cui; Tian, Xiu-Hui; Ye, Jian-Nong

2007-01-01

Corn has been known for its accumulation of flavones and phenolic acids. However, many parts of corn, except kernel, have not drawn much attention. In this work, a method based on capillary zone electrophoresis with electrochemical detection has been used for the separation and determination of epicatechin, rutin, ascorbic acid (Vc), kaempferol, chlorogenic acid, and quercetin in corn silk, leaf, and kernel. The distribution comparison of the ingredients among silk, leaf, and kernel is discussed. Several important factors--including running buffer acidity, separation voltage, and working electrode potential--were evaluated to acquire the optimum analysis conditions. Under the optimum conditions, the analytes could be well separated within 19 min in a 40-mmol/L borate buffer (pH 9.2). The response was linear over three orders of magnitude with detection limits (S/N = 3) ranging from 4.97 x 10(-8) to 9.75 x 10(-8) g/mL. The method has been successfully applied for the analysis of corn silk, leaf, and kernel with satisfactory results.

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

PubMed

Hong, Yanqing; Zhu, Xueke; Wang, Ping; Fu, Haitian; Deng, Chao; Cui, Li; Wang, Qiang; Fan, Xuerong

2016-04-01

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.

Behaviour of human mesenchymal stem cells on a polyelectrolyte-modified HEMA hydrogel for silk-based ligament tissue engineering.

PubMed

Bosetti, M; Boccafoschi, F; Calarco, A; Leigheb, M; Gatti, S; Piffanelli, V; Peluso, G; Cannas, M

2008-01-01

The aim of this study was to design a functional bio-engineered material to be used as scaffold for autologous mesenchymal stem cells in ligament tissue engineering. Polyelectrolyte modified HEMA hydrogel (HEMA-co-METAC), applied as coating on silk fibroin fibres, has been formulated in order to take advantage of the biocompatibility of the polyelectrolyte by increasing its mechanical properties with silk fibres. Human bone marrow mesenchymal stem cells behaviour on such reinforced polyelectrolyte has been studied by evaluating cell morphology, cell number, attachment, spreading and proliferation together with collagen matrix production and its mRNA expression. Silk fibroin fibres matrices with HEMA-co-METAC coating exhibited acceptable mechanical behaviour compared to the natural ligament, good human mesenchymal stem cell adhesion and with mRNA expression studies higher levels of collagen types I and III expression when compared to control cells on polystyrene. These data indicate high expression of mRNA for proteins responsible for the functional characteristics of the ligaments and suggest a potential for use of this biomaterial in ligament tissue-engineering applications.

Milled non-mulberry silk fibroin microparticles as biomaterial for biomedical applications.

PubMed

Bhardwaj, Nandana; Rajkhowa, Rangam; Wang, Xungai; Devi, Dipali

2015-11-01

Silk fibroin has been widely employed in various forms as biomaterials for biomedical applications due to its superb biocompatibility and tunable degradation and mechanical properties. Herein, silk fibroin microparticles of non-mulberry silkworm species (Antheraea assamensis, Antheraea mylitta and Philosamia ricini) were fabricated via a top-down approach using a combination of wet-milling and spray drying techniques. Microparticles of mulberry silkworm (Bombyx mori) were also utilized for comparative studies. The fabricated microparticles were physico-chemically characterized for size, stability, morphology, chemical composition and thermal properties. The silk fibroin microparticles of all species were porous (∼5μm in size) and showed nearly spherical morphology with rough surface as revealed from dynamic light scattering and microscopic studies. Non-mulberry silk microparticles maintained the typical silk-II structure with β-sheet secondary conformation with higher thermal stability. Additionally, non-mulberry silk fibroin microparticles supported enhanced cell adhesion, spreading and viability of mouse fibroblasts than mulberry silk fibroin microparticles (p<0.001) as evidenced from fluorescence microscopy and cytotoxicity studies. Furthermore, in vitro drug release from the microparticles showed a significantly sustained release over 3 weeks. Taken together, this study demonstrates promising attributes of non-mulberry silk fibroin microparticles as a potential drug delivery vehicle/micro carrier for diverse biomedical applications. Copyright © 2015 Elsevier B.V. All rights reserved.

Influence of pH, temperature, and concentration on stabilization of aqueous hornet silk solution and fabrication of salt-free materials.

PubMed

Kameda, Tsunenori

2015-01-01

We found that an aqueous solution of silk from cocoons produced by hornet larvae (hornet silk) can be obtained when the solution is adjusted to basic conditions of pH > 9.2. It is known that native hornet cocoons can be dissolved in concentrated aqueous solution of salts, such as lithium bromide (LiBr) and calcium chloride (CaCl2). Upon the removal of these salts from solution by dialysis, solidification, gelation, or sedimentation of hornet silk is known to occur. In the present study, under basic conditions, however, no such solidification occurred, even after salt removal. In this study, ammonia was used for alkalization of solution because it is volatilized during the casting process and pure hornet silk materials can be obtained after drying. The effects of the concentrations of hornet silk and ammonia, as well as dialysis temperature, on preventing gelation during dialysis were investigated. Dialysis conditions that limit the degradation of hornet silk by hydrolysis in alkali solution were identified. Moreover, casting conditions to prepare flexible and transparent hornet silk film from aqueous ammonia solution were optimized. Molecular structural analysis of hornet silk in aqueous ammonia solution and cast film indicated the formation of α-helix conformations. © 2014 Wiley Periodicals, Inc.

Distinct spinning patterns gain differentiated loading tolerance of silk thread anchorages in spiders with different ecology.

PubMed

Wolff, Jonas O; van der Meijden, Arie; Herberstein, Marie E

2017-07-26

Building behaviour in animals extends biological functions beyond bodies. Many studies have emphasized the role of behavioural programmes, physiology and extrinsic factors for the structure and function of buildings. Structure attachments associated with animal constructions offer yet unrealized research opportunities. Spiders build a variety of one- to three-dimensional structures from silk fibres. The evolution of economic web shapes as a key for ecological success in spiders has been related to the emergence of high performance silks and thread coating glues. However, the role of thread anchorages has been widely neglected in those models. Here, we show that orb-web (Araneidae) and hunting spiders (Sparassidae) use different silk application patterns that determine the structure and robustness of the joint in silk thread anchorages. Silk anchorages of orb-web spiders show a greater robustness against different loading situations, whereas the silk anchorages of hunting spiders have their highest pull-off resistance when loaded parallel to the substrate along the direction of dragline spinning. This suggests that the behavioural 'printing' of silk into attachment discs along with spinneret morphology was a prerequisite for the evolution of extended silk use in a three-dimensional space. This highlights the ecological role of attachments in the evolution of animal architectures. © 2017 The Author(s).

More than a safety line: jump-stabilizing silk of salticids.

PubMed

Chen, Yung-Kang; Liao, Chen-Pan; Tsai, Feng-Yueh; Chi, Kai-Jung

2013-10-06

Salticids are diurnal hunters known for acute vision, remarkable predatory strategies and jumping ability. Like other jumpers, they strive for stability and smooth landings. Instead of using inertia from swinging appendages or aerodynamic forces by flapping wings as in other organisms, we show that salticids use a different mechanism for in-air stability by using dragline silk, which was previously believed to function solely as a safety line. Analyses from high-speed images of jumps by the salticid Hasarius adansoni demonstrate that despite being subject to rearward pitch at take-off, spiders with dragline silk can change body orientation in the air. Instantaneous drag and silk forces calculated from kinematic data further suggest a comparable contribution to deceleration and energy dissipation, and reveal that adjustments by the spider to the silk force can reverse its body pitch for a predictable and optimal landing. Without silk, upright-landing spiders would slip or even tumble, deferring completion of landing. Thus, for salticids, dragline silk is critical for dynamic stability and prey-capture efficiency. The dynamic functioning of dragline silk revealed in this study can advance the understanding of silk's physiological control over material properties and its significance to spider ecology and evolution, and also provide inspiration for future manoeuvrable robot designs.

Enhancing the Mechanical Toughness of Epoxy-Resin Composites Using Natural Silk Reinforcements

DOE PAGES

Yang, Kang; Wu, Sujun; Guan, Juan; ...

2017-09-20

Strong and tough epoxy composites are developed using a less-studied fibre reinforcement, that of natural silk. Two common but structurally distinct silks from the domestic B. mori/Bm and the wild A. pernyi/Ap silkworms are selected in fabric forms. We show that the toughening effects on silk-epoxy composites or SFRPs are dependent on the silk species and the volume fraction of silk. Both silks enhance the room-temperature tensile and flexural mechanical properties of the composite, whereas the more resilient Ap silk shows a more pronounced toughening effect and a lower critical reinforcement volume for the brittle-ductile transition. Specifically, our 60 vol.%more » Ap-SFRP displays a three-fold elevation in tensile and flexural strength, as compared to pure epoxy resin, with an order of magnitude higher breaking energy via a distinct, ductile failure mode. Importantly, the 60 vol.% Ap-SFRP remains ductile with 7% flexural elongation at lower temperatures (-50 °C). Under impact, these SFRPs show significantly improved energy absorption, and the 60 vol.% Ap-SFRP has an impact strength some eight times that of pure epoxy resin. Lastly, the findings demonstrate both marked toughening and strengthening effects for epoxy composites from natural silk reinforcements, which presents opportunities for mechanically superior and "green" structural composites.« less

Accelerated In Vitro Degradation of Optically Clear Low β-Sheet Silk Films by Enzyme-Mediated Pretreatment

PubMed Central

Shang, Ke; Rnjak-Kovacina, Jelena; Lin, Yinan; Hayden, Rebecca S.; Tao, Hu; Kaplan, David L.

2013-01-01

Purpose: To design patterned, transparent silk films with fast degradation rates for the purpose of tissue engineering corneal stroma. Methods: β-sheet (crystalline) content of silk films was decreased significantly by using a short water annealing time. Additionally, a protocol combining short water annealing time with enzymatic pretreatment of silk films with protease XIV was developed. Results: Low β-sheet content (17%–18%) and enzymatic pretreatment provided film stability in aqueous environments and accelerated degradation of the silk films in the presence of human corneal fibroblasts in vitro. The results demonstrate a direct relationship between reduced β-sheet content and enzymatic pretreatment, and overall degradation rate of the protein films. Conclusions: The novel protocol developed here provides new approaches to modulate the regeneration rate of silk biomaterials for corneal tissue regeneration needs. Translational Relevance: Patterned silk protein films possess desirable characteristics for corneal tissue engineering, including optical transparency, biocompatibility, cell alignment, and tunable mechanical properties, but current fabrication protocols do not provide adequate degradation rates to match the regeneration properties of the human cornea. This novel processing protocol makes silk films more suitable for the construction of human corneal stroma tissue and a promising way to tune silk film degradation properties to match corneal tissue regeneration. PMID:24049717

Accelerated in vitro Degradation of Optically Clear Low β-sheet Silk Films by Enzyme-Mediated Pretreatment

PubMed Central

Shang, Ke; Rnjak-Kovacina, Jelena; Lin, Yinan; Hayden, Rebecca S.; Hu, Tao; Kaplan, David L.

2013-01-01

Purpose To design patterned, transparent silk films with fast degradation rates for the purpose of tissue engineering corneal stroma, Methods β-sheet (crystalline) content of silk films was decreased significantly by using a short water annealing time. Additionally, a protocol combining short water annealing time with enzymatic pretreatment of silk films with protease XIV was developed. Results Low β-sheet content (17–18%) and enzymatic pre-treatment provided film stability in aqueous environments and accelerated degradation of the silk films in the presence of human corneal fibroblasts in vitro. The results demonstrate a direct relationship between reduced β-sheet content and enzymatic pre-treatment and overall degradation rate of the protein films. Conclusions The novel protocol developed here provides new approaches to modulate the regeneration rate of silk biomaterials for corneal tissue regeneration needs. Translational relevance Patterned silk protein films possess desirable characteristics for corneal tissue engineering, including optical transparency, biocompatibility, cell alignment and tunable mechanical properties, but current fabrication protocols do not provide adequate degradation rates to match the regeneration properties of the human cornea. This novel processing protocol makes silk films more suitable for the construction of human corneal stroma tissue and a promising way to tune silk film degradation properties to match corneal tissue regeneration. PMID:23579493

More than a safety line: jump-stabilizing silk of salticids

PubMed Central

Chen, Yung-Kang; Liao, Chen-Pan; Tsai, Feng-Yueh; Chi, Kai-Jung

2013-01-01

Salticids are diurnal hunters known for acute vision, remarkable predatory strategies and jumping ability. Like other jumpers, they strive for stability and smooth landings. Instead of using inertia from swinging appendages or aerodynamic forces by flapping wings as in other organisms, we show that salticids use a different mechanism for in-air stability by using dragline silk, which was previously believed to function solely as a safety line. Analyses from high-speed images of jumps by the salticid Hasarius adansoni demonstrate that despite being subject to rearward pitch at take-off, spiders with dragline silk can change body orientation in the air. Instantaneous drag and silk forces calculated from kinematic data further suggest a comparable contribution to deceleration and energy dissipation, and reveal that adjustments by the spider to the silk force can reverse its body pitch for a predictable and optimal landing. Without silk, upright-landing spiders would slip or even tumble, deferring completion of landing. Thus, for salticids, dragline silk is critical for dynamic stability and prey-capture efficiency. The dynamic functioning of dragline silk revealed in this study can advance the understanding of silk's physiological control over material properties and its significance to spider ecology and evolution, and also provide inspiration for future manoeuvrable robot designs. PMID:23925983

Morus planting and silk production in Oxus civilization (2000 BC), Transoxiana

NASA Astrophysics Data System (ADS)

Zhou, X.; Li, X.; Wang, J.

2017-12-01

Silk is the most important luxury in ancient trade, which promoted communications between east and west civilizations that facilitated the development of them. Currently, probably more than 99% of the silk in commerce in the world today comes from a single species called the silkworm or mulberry silk moth, Bombyx mori (L.), of the family Bombycidae. At present, a list of occurrences of unearthed early silk thread, textile and terracotta figures of China suggests that wild silk has been widely used for weave in Yangtze river and Yellow River basin at least 5000 cal yr BP. On the other hand, the earliest silk fabric out China is from the earliest civilization Harappa site (4500-4000 cal a BP) that confirmed the early use of wild silkworms in South Asia.. In addition, in the Egyptian Deir al Medina relics site, the unearthed silk also as old as nearly 3000 cal a BP, and a serials of pre-han silk also unearthed 3000-2500 cal a BP in Europe. But, the appearance of these out-China silk is usually considered to be the result of the early trade or wild silkmoth products. Here we present a synthesis study of chronology, entomology, protein group, the paleoethnobotany form three Bronze Age site (Sapalli, Jarzgudan, Molleli) concerning on the paleo environment, agriculture gardening, and the possible mulberry silkworm utilization in Bronze Age Tensoxiana 4000 cal a BP. Together with archaeobotany and chronology data from Xinjiang, Hexi corridor and the Loess Plateau in China, we analysis the status of the mulberry silk factory in Bronze Age in Transoxiana and it possible connection to ancient China.

Reversible Hydrogel–Solution System of Silk with High Beta-Sheet Content

PubMed Central

2015-01-01

Silkworm silk has been widely used as a textile fiber, as biomaterials and in optically functional materials due to its extraordinary properties. The β-sheet-rich natural nanofiber units of about 10–50 nm in diameter are often considered the origin of these properties, yet it remains unclear how silk self-assembles into these hierarchical structures. A new system composed of β-sheet-rich silk nanofibers about 10–20 nm in diameter is reported here, where these nanofibers formed into “flowing hydrogels” at 0.5–2% solutions and could be transformed back into the solution state at lower concentrations, even with a high β-sheet content. This is in contrast with other silk processed materials, where significant β-sheet content negates reversibility between solution and solid states. These fibers are formed by regulating the self-assembly process of silk in aqueous solution, which changes the distribution of negative charges while still supporting β-sheet formation in the structures. Mechanistically, there appears to be a shift toward negative charges along the outside of the silk nanofibers in our present study, resulting in a higher zeta potential (above −50 mV) than previous silk materials which tend to be below −30 mV. The higher negative charge on silk nanofibers resulted in electrostatic repulsion strong enough to negate further assembly of the nanofibers. Changing silk concentration changed the balance between hydrophobic interactions and electrostatic repulsion of β-sheet-rich silk nanofibers, resulting in reversible hydrogel–solution transitions. Furthermore, the silk nanofibers could be disassembled into shorter fibers and even nanoparticles upon ultrasonic treatment following the transition from hydrogel to solution due to the increased dispersion of hydrophobic smaller particles, without the loss of β-sheet content, and with retention of the ability to transition between hydrogel and solution states through reversion to longer nanofibers during self-assembly. These reversible solution-hydrogel transitions were tunable with ultrasonic intensity, time, or temperature. PMID:25056606

Reversible hydrogel-solution system of silk with high beta-sheet content.

PubMed

Bai, Shumeng; Zhang, Xiuli; Lu, Qiang; Sheng, Weiqin; Liu, Lijie; Dong, Boju; Kaplan, David L; Zhu, Hesun

2014-08-11

Silkworm silk has been widely used as a textile fiber, as biomaterials and in optically functional materials due to its extraordinary properties. The β-sheet-rich natural nanofiber units of about 10-50 nm in diameter are often considered the origin of these properties, yet it remains unclear how silk self-assembles into these hierarchical structures. A new system composed of β-sheet-rich silk nanofibers about 10-20 nm in diameter is reported here, where these nanofibers formed into "flowing hydrogels" at 0.5-2% solutions and could be transformed back into the solution state at lower concentrations, even with a high β-sheet content. This is in contrast with other silk processed materials, where significant β-sheet content negates reversibility between solution and solid states. These fibers are formed by regulating the self-assembly process of silk in aqueous solution, which changes the distribution of negative charges while still supporting β-sheet formation in the structures. Mechanistically, there appears to be a shift toward negative charges along the outside of the silk nanofibers in our present study, resulting in a higher zeta potential (above -50 mV) than previous silk materials which tend to be below -30 mV. The higher negative charge on silk nanofibers resulted in electrostatic repulsion strong enough to negate further assembly of the nanofibers. Changing silk concentration changed the balance between hydrophobic interactions and electrostatic repulsion of β-sheet-rich silk nanofibers, resulting in reversible hydrogel-solution transitions. Furthermore, the silk nanofibers could be disassembled into shorter fibers and even nanoparticles upon ultrasonic treatment following the transition from hydrogel to solution due to the increased dispersion of hydrophobic smaller particles, without the loss of β-sheet content, and with retention of the ability to transition between hydrogel and solution states through reversion to longer nanofibers during self-assembly. These reversible solution-hydrogel transitions were tunable with ultrasonic intensity, time, or temperature.

Silk film biomaterials for ocular surface repair

NASA Astrophysics Data System (ADS)

Lawrence, Brian David

Current biomaterial approaches for repairing the cornea's ocular surface upon injury are partially effective due to inherent material limitations. As a result there is a need to expand the biomaterial options available for use in the eye, which in turn will help to expand new clinical innovations and technology development. The studies illustrated here are a collection of work to further characterize silk film biomaterials for use on the ocular surface. Silk films were produced from regenerated fibroin protein solution derived from the Bombyx mori silkworm cocoon. Methods of silk film processing and production were developed to produce consistent biomaterials for in vitro and in vivo evaluation. A wide range of experiments was undertaken that spanned from in vitro silk film material characterization to in vivo evaluation. It was found that a variety of silk film properties could be controlled through a water-annealing process. Silk films were then generated that could be use in vitro to produce stratified corneal epithelial cell sheets comparable to tissue grown on the clinical standard substrate of amniotic membrane. This understanding was translated to produce a silk film design that enhanced corneal healing in vivo on a rabbit injury model. Further work produced silk films with varying surface topographies that were used as a simplified analog to the corneal basement membrane surface in vitro. These studies demonstrated that silk film surface topography is capable of directing corneal epithelial cell attachment, growth, and migration response. Most notably epithelial tissue development was controllably directed by the presence of the silk surface topography through increasing cell sheet migration efficiency at the individual cellular level. Taken together, the presented findings represent a comprehensive characterization of silk film biomaterials for use in ocular surface reconstruction, and indicate their utility as a potential material choice in the development of innovative procedures and technologies for corneal repair.

An All-Silk-Derived Dual-Mode E-skin for Simultaneous Temperature-Pressure Detection.

PubMed

Wang, Chunya; Xia, Kailun; Zhang, Mingchao; Jian, Muqiang; Zhang, Yingying

2017-11-15

Flexible skin-mimicking electronics are highly desired for development of smart human-machine interfaces and wearable human-health monitors. Human skins are able to simultaneously detect different information, such as touch, friction, temperature, and humidity. However, due to the mutual interferences of sensors with different functions, it is still a big challenge to fabricate multifunctional electronic skins (E-skins). Herein, a combo temperature-pressure E-skin is reported through assembling a temperature sensor and a strain sensor in both of which flexible and transparent silk-nanofiber-derived carbon fiber membranes (SilkCFM) are used as the active material. The temperature sensor presents high temperature sensitivity of 0.81% per centigrade. The strain sensor shows an extremely high sensitivity with a gauge factor of ∼8350 at 50% strain, enabling the detection of subtle pressure stimuli that induce local strain. Importantly, the structure of the SilkCFM in each sensor is designed to be passive to other stimuli, enabling the integrated E-skin to precisely detect temperature and pressure at the same time. It is demonstrated that the E-skin can detect and distinguish exhaling, finger pressing, and spatial distribution of temperature and pressure, which cannot be realized using single mode sensors. The remarkable performance of the silk-based combo temperature-pressure sensor, together with its green and large-scalable fabrication process, promising its applications in human-machine interfaces and soft electronics.

Response of human corneal fibroblasts on silk film surface patterns.

PubMed

Gil, Eun Seok; Park, Sang-Hyug; Marchant, Jeff; Omenetto, Fiorenzo; Kaplan, David L

2010-06-11

Transparent, biodegradable, mechanically robust, and surface-patterned silk films were evaluated for the effect of surface morphology on human corneal fibroblast (hCF) cell proliferation, orientation, and ECM deposition and alignment. A series of dimensionally different surface groove patterns were prepared from optically graded glass substrates followed by casting poly(dimethylsiloxane) (PDMS) replica molds. The features on the patterned silk films showed an array of asymmetric triangles and displayed 37-342 nm depths and 445-3 582 nm widths. hCF DNA content on all patterned films were not significantly different from that on flat silk films after 4 d in culture. However, the depth and width of the grooves influenced cell alignment, while the depth differences affected cell orientation; overall, deeper and narrower grooves induced more hCF orientation. Over 14 d in culture, cell layers and actin filament organization demonstrated that confluent hCFs and their cytoskeletal filaments were oriented along the direction of the silk film patterned groove axis. Collagen type V and proteoglycans (decorin and biglycan), important markers of corneal stromal tissue, were highly expressed with alignment. Understanding corneal stromal fibroblast responses to surface features on a protein-based biomaterial applicable in vivo for corneal repair potential suggests options to improve corneal tissue mimics. Further, the approaches provide fundamental biomaterial designs useful for bioengineering oriented tissue layers, an endemic feature in most biological tissue structures that lead to critical tissue functions.

Economic feasibility and environmental impact of synthetic spider silk production from escherichia coli.

PubMed

Edlund, Alan M; Jones, Justin; Lewis, Randolph; Quinn, Jason C

2018-05-25

Major ampullate spider silk represents a promising protein-based biomaterial with diverse commercial potential ranging from textiles to medical devices due to its excellent physical and thermal properties. Recent advancements in synthetic biology have facilitated the development of recombinant spider silk proteins from Escherichia coli (E. coli). This study specifically investigates the economic feasibility and environmental impact of synthetic spider silk manufacturing. Pilot scale data was used to validate an engineering process model that includes all of the required sub-processing steps for synthetic fiber manufacture: production, harvesting, purification, drying, and spinning. Modeling was constructed modularly to support assessment of alternative downstream processing technologies. The techno-economic analysis indicates a minimum sale price from pioneer and optimized E. coli plants of $761 kg -1 and $23 kg -1 with greenhouse gas emissions of 572 kg CO 2-eq. kg -1 and 55 kg CO 2-eq. kg -1 , respectively. Elevated costs and emissions from the pioneer plant can be directly tied to the high material consumption and low protein yield. Decreased production costs associated with the optimized plant includes improved protein yield, process optimization, and an N th plant assumption. Discussion focuses on the commercial potential of spider silk, the production performance requirements for commercialization, and the impact of alternative technologies on the system. Copyright © 2018 Elsevier B.V. All rights reserved.

The use of injectable sonication-induced silk hydrogel for VEGF165 and BMP-2 delivery for elevation of the maxillary sinus floor

PubMed Central

Zhang, Wenjie; Wang, Xiuli; Wang, Shaoyi; Zhao, Jun; Xu, Lianyi; Zhu, Chao; Zeng, Deliang; Chen, Jake; Zhang, Zhiyuan; Kaplan, David L.; Jiang, Xinquan

2011-01-01

Sonication-induced silk hydrogels were previously prepared as an injectable bone replacement biomaterial, with a need to improve osteogenic features. Vascular endothelial growth factor (VEGF165) and bone morphogenic protein-2 (BMP-2) are key regulators of angiogenesis and osteogenesis, respectively, during bone regeneration. Therefore, the present study aimed at evaluating in situ forming silk hydrogels as a vehicle to encapsulate dual factors for rabbit maxillary sinus floor augmentation. Sonication-induced silk hydrogels were prepared in vitro and the slow release of VEGF165 and BMP-2 from these silk gels was evaluated by ELISA. For in vivo studies for each time point (4 and 12 weeks), 24 sinus floors elevation surgeries were made bilaterally in 12 rabbits for the following four treatment groups: silk gel (group Silk gel), silk gel/VEGF165 (group VEGF), silk gel/BMP-2 (group BMP-2), silk gel/VEGF165/BMP-2 (group V+B) (n=6 per group). Sequential florescent labeling and radiographic observations were used to record new bone formation and mineralization, along with histological and histomorphometric analysis. At week 4, VEGF165 promoted more tissue infiltration into the gel and accelerated the degradation of the gel material. At this time point, the bone area in group V+B was significantly larger than those in the other three groups. At week 12, elevated sinus floor heights of groups BMP-2 and V+B were larger than those of the Silk gel and VEGF groups, and the V+B group had the largest new bone area among all groups. In addition, a larger blood vessel area formed in the remaining gel areas in groups VEGF and V+B. In conclusion, VEGF165 and BMP-2 released from injectable and biodegradable silk gels promoted angiogenesis and new bone formation, with the two factors demonstrating an additive effect on bone regeneration. These results indicate that silk hydrogels can be used as an injectable vehicle to deliver multiple growth factors in a minimally invasive approach to regenerate irregular bony cavities. PMID:21889205

Surface processing and ageing behavior of silk fabrics treated with atmospheric-pressure plasma for pigment-based ink-jet printing

NASA Astrophysics Data System (ADS)

Zhang, Chunming; Wang, Libing; Yu, Miao; Qu, Lijun; Men, Yajing; Zhang, Xiangwu

2018-03-01

Pigment inkjet printing has highlighted the advantages of cost-effective, short production cycle and environment-friendly. However, patterns directly printed with pigment inks usually have low color yields and blurry images which are caused by bleeding phenomenon. This work presents an atmospheric-pressure plasma method for improving the pigment-based ink-jet printing performance of silk fabrics. The effects of surface changes induced are discussed, with data derived from morphological study by atomic force microscopy (AFM), chemical analysis using X-ray photoelectron spectroscopy (XPS) and contact angle measurement. Ink-jet printing experiments were conducted to study the influence of measured changes on anti-bleeding property and color strength of treated and original samples. The ageing experiment indicates that the modified silk fabrics should be printed within 24 h after plasma processing for maximum color yields. This study explores an effective approach for the atmospheric-pressure plasma, which can provide its significant use in improving the surface properties and ink-jet printing performance of fabrics.

Effect of UV-light on the uniaxial tensile properties and structure of uncoated and TiO2 coated Bombyx mori silk fibers

NASA Astrophysics Data System (ADS)

Aksakal, Baki; Koç, Kenan; Yargı, Önder; Tsobkallo, Katherina

2016-01-01

The effect of UV-light on the uniaxial tensile properties and the structure of uncoated and TiO2 coated silk fibers in the bave form by using sol-gel method was investigated with tensile testing and FT-IR/ATR spectroscopy methods after the silk filaments were exposed to UV-light with high intensity of 760 W/m2 for different times from 0.5 h to 1 day. It was clearly observed that TiO2 coating considerably increased the Young's modulus of the uncoated silk single filament by around 17% before the UV-irradiation. The yield point and the post yield region disappeared on the stress-strain curves of both uncoated and TiO2 coated silk filaments after UV-irradiation time higher than 1 h. Except for the Young's modulus, most of the tensile characteristics of both uncoated and TiO2 coated silk filaments decreased remarkably with increasing UV-irradiation time, e.g., after 1 h irradiation, although the Young's modulus slightly changed and ultimate tensile strength decreased by only around 18% and 23%, for the uncoated and TiO2 coated silk filaments, respectively; breaking extension decreased dramatically by 67% and 72%, respectively, for uncoated and TiO2 coated silk filaments. Only the Young's modulus of TiO2 coated silk filaments which can be considered as a more stable tensile characteristic became significantly higher than that of the uncoated silk filaments with increasing UV-irradiation time. After 1 day irradiation, even though the uncoated silk filaments could not be tested and completely lost of their fiber properties, the TiO2 coated silk filaments showed a stress-strain curve in initial elastic region with Young's modulus of ∼13 GPa which indicates considerable protective effect of TiO2 on the silk fiber structure, especially on the β-sheet microcrystals against UV-radiation. The FT-IR/ATR spectral results showed that significant photodegradation took place in not only crystalline but also amorphous regions which were deduced from the decrease in the absorbance ratios of the bands assigned to CH3 rocking, Cα-Cβ, Cα-C stretching vibrations in β-sheet crystalline regions as well as the Amide I, II, and III bands for both crystalline and amorphous regions. Even though the ratio of crystalline to amorphous regions in uncoated silk filaments decreased significantly, the ratio in TiO2 coated silk filaments became almost constant with increasing UV-irradiation time which may indicate more stable β-sheet microcrystals against photodegradation.

The construction of an EST database for Bombyx mori and its application

PubMed Central

Mita, Kazuei; Morimyo, Mitsuoki; Okano, Kazuhiro; Koike, Yoshiko; Nohata, Junko; Kawasaki, Hideki; Kadono-Okuda, Keiko; Yamamoto, Kimiko; Suzuki, Masataka G.; Shimada, Toru; Goldsmith, Marian R.; Maeda, Susumu

2003-01-01

To build a foundation for the complete genome analysis of Bombyx mori, we have constructed an EST database. Because gene expression patterns deeply depend on tissues as well as developmental stages, we analyzed many cDNA libraries prepared from various tissues and different developmental stages to cover the entire set of Bombyx genes. So far, the Bombyx EST database contains 35,000 ESTs from 36 cDNA libraries, which are grouped into ≈11,000 nonredundant ESTs with the average length of 1.25 kb. The comparison with FlyBase suggests that the present EST database, SilkBase, covers >55% of all genes of Bombyx. The fraction of library-specific ESTs in each cDNA library indicates that we have not yet reached saturation, showing the validity of our strategy for constructing an EST database to cover all genes. To tackle the coming saturation problem, we have checked two methods, subtraction and normalization, to increase coverage and decrease the number of housekeeping genes, resulting in a 5–11% increase of library-specific ESTs. The identification of a number of genes and comprehensive cloning of gene families have already emerged from the SilkBase search. Direct links of SilkBase with FlyBase and WormBase provide ready identification of candidate Lepidoptera-specific genes. PMID:14614147

The Study of Importance of the Balance Space Food -Storage Method -

NASA Astrophysics Data System (ADS)

Katayama, Naomi; Yamashita, Masamichi; Hashimoto, Hirofumi; Space Agriculture Task Force, J.

Providing foods to space crew is the important requirements to support long term manned space exploration. Foods fill not only physiological requirements to sustain life, but psychological needs for refreshment and joy during the long and hard mission to extraterrestrial planets. We designed joyful and healthy recipe with materials, which can be produced by the bio-regenerative agricultural system operated at limited resources available in Mars base, Moon base and spaceship. We need to think about how to use the storage food when we have the time of emergency. The pupa of the silkworm becomes the important nourishment source as protein and lipid. The silk thread uses it as clothing and cosmetics and medical supplies. However, we can use the silk thread as food as protein. The silk thread is mad of sericin and fibroin. The sericin is used for cosmetics mainly, but can make sheet food by mixing it with rice flour. We can make Japanese rolled sushi with this product. In addition, we can make spring roll and gyoza and shao-mai. As for the fibroin which is the subject of the silk thread, is to extract it high pressure heat; of the protein can powder it, and can use it as food. Even if there is the silk thread in this way after having made it clothes once, we can do it to food again. We can reuse the cotton thread as carbohydrates equally, too. We can use the wood as carbohydrates, also. Based upon the foregoing, we use the pupa of the silkworm as protein and lipid, and the silk thread as protein, and the cotton thread and wood as carbohydrates. It is recommended as healthy meal balance; Protein: Lipid: Carbohydrate ratio equal 15-20We succeeded to develop joyful and nutritious space recipe at the end. Since energy consumption for physical exercise activities under micro-or sub-gravity is less than the terrestrial case, choice of our space foods is essencial to suppress blood sugar level, and prevent the metabolic syndrome. Because of less need of agricultural resources at choosing ecological members from the lower ladder of the food chain, our space recipe could be a proposal to solve the food problem on Earth.

Silk Roads or Steppe Roads? The Silk Roads in World History.

ERIC Educational Resources Information Center

Christian, David

2000-01-01

Explores the prehistory of the Silk Roads, reexamines their structure and history in the classical era, and explores shifts in their geography in the last one thousand years. Explains that a revised understanding of the Silk Roads demonstrates how the Afro-Eurasian land mass has been linked by networks of exchange since the Bronze Age. (CMK)

Derivation of a variational principle for plane strain elastic-plastic silk biopolymers

NASA Astrophysics Data System (ADS)

He, J. H.; Liu, F. J.; Cao, J. H.; Zhang, L.

2014-01-01

Silk biopolymers, such as spider silk and Bombyx mori silk, behave always elastic-plastically. An elastic-plastic model is adopted and a variational principle for the small strain, rate plasticity problem is established by semi-inverse method. A trial Lagrangian is constructed where an unknown function is included which can be identified step by step.

Structural and thermal properties of silk fibroin - Silver nanoparticles composite films

NASA Astrophysics Data System (ADS)

Shivananda, C. S.; Rao B, B. Lakshmeesha; Shetty, G. Rajesh; Sangappa, Y.

2018-05-01

In this work, silk fibroin-silver nanoparticles (SF-AgNPs) composite films have been prepared by simple solution casting method. The composite films were examined for structural and thermal properties using X-ray diffraction (XRD), thermogravimatric (TGA) and differential scanning calorimetry (DSC) analysis. The XRD results showed that with the introduction of AgNPs in the silk fibroin matrix the amorphous nature of the silk fibroin decreases with increasing nanoparticles concentration. The silk fibroin films possess good thermal stability with the presence of AgNPs.

Silk fabric dyed with extract of sophora flower bud.

PubMed

Yan, Su; Pan, Shanshan; Ji, Junling

2018-02-01

This study analysed the use of sophora flower bud extract for dyeing and the resulting colour character and fastness of dyed silk fabric. The pigment composition on the silk fabric and recycling of this extract were also studied. The results indicated that the dyed silk fabric possessed good washing, rubbing and perspiration fastness, and the pigment composition on the silk fabric was mainly rutin and quercetin. The average recovery rate of the dye was 55.00%. These results demonstrate that the sophora flower bud extract is an effective natural dye.

Structure-Function-Property-Design Interplay in Biopolymers: Spider Silk

PubMed Central

Tokareva, Olena; Jacobsen, Matthew; Buehler, Markus; Wong, Joyce; Kaplan, David L.

2013-01-01

Spider silks have been a focus of research for almost two decades due to their outstanding mechanical and biophysical properties. Recent advances in genetic engineering have led to the synthesis of recombinant spider silks, thus helping to unravel a fundamental understanding of structure-function-property relationships. The relationships between molecular composition, secondary structures, and mechanical properties found in different types of spider silks are described, along with a discussion of artificial spinning of these proteins and their bioapplications, including the role of silks in biomineralization and fabrication of biomaterials with controlled properties. PMID:23962644

3D freeform printing of silk fibroin.

PubMed

Rodriguez, Maria J; Dixon, Thomas A; Cohen, Eliad; Huang, Wenwen; Omenetto, Fiorenzo G; Kaplan, David L

2018-04-15

Freeform fabrication has emerged as a key direction in printing biologically-relevant materials and structures. With this emerging technology, complex structures with microscale resolution can be created in arbitrary geometries and without the limitations found in traditional bottom-up or top-down additive manufacturing methods. Recent advances in freeform printing have used the physical properties of microparticle-based granular gels as a medium for the submerged extrusion of bioinks. However, most of these techniques require post-processing or crosslinking for the removal of the printed structures (Miller et al., 2015; Jin et al., 2016) [1,2]. In this communication, we introduce a novel method for the one-step gelation of silk fibroin within a suspension of synthetic nanoclay (Laponite) and polyethylene glycol (PEG). Silk fibroin has been used as a biopolymer for bioprinting in several contexts, but chemical or enzymatic additives or bulking agents are needed to stabilize 3D structures. Our method requires no post-processing of printed structures and allows for in situ physical crosslinking of pure aqueous silk fibroin into arbitrary geometries produced through freeform 3D printing. 3D bioprinting has emerged as a technology that can produce biologically relevant structures in defined geometries with microscale resolution. Techniques for fabrication of free-standing structures by printing into granular gel media has been demonstrated previously, however, these methods require crosslinking agents and post-processing steps on printed structures. Our method utilizes one-step gelation of silk fibroin within a suspension of synthetic nanoclay (Laponite), with no need for additional crosslinking compounds or post processing of the material. This new method allows for in situ physical crosslinking of pure aqueous silk fibroin into defined geometries produced through freeform 3D printing. Copyright © 2018 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

Into the groove: instructive silk-polypyrrole films with topographical guidance cues direct DRG neurite outgrowth.

PubMed

Hardy, John G; Khaing, Zin Z; Xin, Shangjing; Tien, Lee W; Ghezzi, Chiara E; Mouser, David J; Sukhavasi, Rushi C; Preda, Rucsanda C; Gil, Eun S; Kaplan, David L; Schmidt, Christine E

2015-01-01

Instructive biomaterials capable of controlling the behaviour of the cells are particularly interesting scaffolds for tissue engineering and regenerative medicine. Novel biomaterials are particularly important in societies with rapidly aging populations, where demand for organ/tissue donations is greater than their supply. Herein we describe the preparation of electrically conductive silk film-based nerve tissue scaffolds that are manufactured using all aqueous processing. Aqueous solutions of Bombyx mori silk were cast on flexible polydimethylsiloxane substrates with micrometer-scale grooves on their surfaces, allowed to dry, and annealed to impart β-sheets to the silk which assures that the materials are stable for further processing in water. The silk films were rendered conductive by generating an interpenetrating network of polypyrrole and polystyrenesulfonate in the silk matrix. Films were incubated in an aqueous solution of pyrrole (monomer), polystyrenesulfonate (dopant) and iron chloride (initiator), after which they were thoroughly washed to remove low molecular weight components (monomers, initiators, and oligomers) and dried, yielding conductive films with sheet resistances of 124 ± 23 kΩ square(-1). The micrometer-scale grooves that are present on the surface of the films are analogous to the natural topography in the extracellular matrix of various tissues (bone, muscle, nerve, skin) to which cells respond. Dorsal root ganglions (DRG) adhere to the films and the grooves in the surface of the films instruct the aligned growth of processes extending from the DRG. Such materials potentially enable the electrical stimulation (ES) of cells cultured on them, and future in vitro studies will focus on understanding the interplay between electrical and topographical cues on the behaviour of cells cultured on them.

The performance of silk scaffolds in a rat model of augmentation cystoplasty.

PubMed

Seth, Abhishek; Chung, Yeun Goo; Gil, Eun Seok; Tu, Duong; Franck, Debra; Di Vizio, Dolores; Adam, Rosalyn M; Kaplan, David L; Estrada, Carlos R; Mauney, Joshua R

2013-07-01

The diverse processing plasticity of silk-based biomaterials offers a versatile platform for understanding the impact of structural and mechanical matrix properties on bladder regenerative processes. Three distinct groups of 3-D matrices were fabricated from aqueous solutions of Bombyx mori silk fibroin either by a gel spinning technique (GS1 and GS2 groups) or a solvent-casting/salt-leaching method in combination with silk film casting (FF group). SEM analyses revealed that GS1 matrices consisted of smooth, compact multi-laminates of parallel-oriented silk fibers while GS2 scaffolds were composed of porous (pore size range, 5-50 μm) lamellar-like sheets buttressed by a dense outer layer. Bi-layer FF scaffolds were comprised of porous foams (pore size, ~400 μm) fused on their external face with a homogenous, nonporous silk film. Silk groups and small intestinal submucosa (SIS) matrices were evaluated in a rat model of augmentation cystoplasty for 10 weeks of implantation and compared to cystotomy controls. Gross tissue evaluations revealed the presence of intra-luminal stones in all experimental groups. The incidence and size of urinary calculi was the highest in animals implanted with gel spun silk matrices and SIS with frequencies ≥57% and stone diameters of 3-4 mm. In contrast, rats augmented with FF scaffolds displayed substantially lower rates (20%) and stone size (2 mm), similar to the levels observed in controls (13%, 2 mm). Histological (hematoxylin and eosin, Masson's trichrome) and immunohistochemical (IHC) analyses showed comparable extents of smooth muscle regeneration and contractile protein (α-smooth muscle actin and SM22α) expression within defect sites supported by all matrix groups similar to controls. Parallel evaluations demonstrated the formation of a transitional, multi-layered urothelium with prominent uroplakin and p63 protein expression in all experimental groups. De novo innervation and vascularization processes were evident in all regenerated tissues indicated by Fox3-positive neuronal cells and vessels lined with CD31 expressing endothelial cells. In comparison to other biomaterial groups, cystometric analyses at 10 weeks post-op revealed that animals implanted with the FF matrix configuration displayed superior urodynamic characteristics including compliance, functional capacity, as well as spontaneous non voiding contractions consistent with control levels. Our data demonstrate that variations in scaffold processing techniques can influence the in vivo functional performance of silk matrices in bladder reconstructive procedures. Copyright © 2013 Elsevier Ltd. All rights reserved.

Nanosized CaP-silk fibroin-PCL-PEG-PCL/PCL based bilayer membranes for guided bone regeneration.

PubMed

Türkkan, Sibel; Pazarçeviren, A Engin; Keskin, Dilek; Machin, Nesrin E; Duygulu, Özgür; Tezcaner, Ayşen

2017-11-01

Guided bone regeneration (GBR) concept has been developed to prevent the formation of non-functional scar tissue layer on defect site by undertaking barrier role. In this study, a new bilayer membrane which consisted of one layer of electrospun silk fibroin/PCL-PEG-PCL incorporating nanocalcium phosphate (SPCA) 1 and one layer of PCL membrane was developed for GBR. To improve the osteoconductivity of membranes, nanosized calcium phosphate particles synthesized by Flame Spray Pyrolysis method were incorporated into membranes at 10% (wt) (SPCA10) and 20% (wt) (SPCA20) of the polymer content. The structural and chemical analyses revealed the well-integrated two layers of membranes with a total thickness of ca 100μm. In the regenerative layer, the highly porous mesh structure had a thickness of 12.6μm with randomly oriented fibers having diameters around 760nm, and nanoparticles dispersed homogenously. The mechanical test results showed remarkable improvement on the tensile strength of membranes with incorporation of nanoparticles. Higher water affinity of nanoCaP included membranes was proved by lower contact angle values and higher percent water uptake capacity. Biomineralization assay revealed that nucleation and growth of apatites around fibers of SPCA10 and SPCA20 were apparent while on SPCA0 apatite minerals were barely detected after 10days. Human dental pulp stem cells (DPSC) were seeded on electrospun layer of the bilayer membranes for biocompatibility and osteo-compatibility study. Increasing nanoCaP amount resulted in higher cell adhesion, proliferation, ALP activity and calcium deposition on membranes. These overall results confirmed the biocompatibility and potential applicability of proposed membranes for GBR treatments. Copyright © 2017 Elsevier B.V. All rights reserved.

Carbon nanoscroll-silk crystallite hybrid structures with controllable hydration and mechanical properties.

PubMed

Cheng, Yuan; Koh, Leng-Duei; Wang, Fan; Li, Dechang; Ji, Baohua; Yeo, Jingjie; Guan, Guijian; Han, Ming-Yong; Zhang, Yong-Wei

2017-07-06

Hybrid structures of nanomaterials (e.g. tubes, scrolls, threads, cages) and biomaterials (e.g. proteins) hold tremendous potential for applications as drug carriers, biosensors, tissue scaffolds, cancer therapeutic agents, etc. However, in many cases, the interacting forces at the nano-bio interfaces and their roles in controlling the structures and dynamics of nano-bio-hybrid systems are very complicated but poorly understood. In this study, we investigate the structure and mechanical behavior of a protein-based hybrid structure, i.e., a carbon nanoscroll (CNS)-silk crystallite with a hydration level controllable by an interlayer interaction in CNS. Our findings demonstrate that CNS with a reduced core size not only shields the crystallite from a weakening effect of water, but also markedly strengthens the crystallite. Besides water shielding, the enhanced strength arises from an enhanced interaction between the crystallite and CNS due to the enhanced interlayer interaction in CNS. In addition, the interfacial strength for pulling the crystallite out of the CNS-silk structure is found to be dependent on both the interlayer interaction energy in CNS as well as the sequence of protein at the CNS-silk interface. The present study is of significant value in designing drugs or protein delivery vehicles for biomedical applications, and serves as a general guide in designing novel devices based on rolled-up configurations of two-dimensional (2D) materials.

Silkworm Sericin: Properties and Biomedical Applications.

PubMed

Kunz, Regina Inês; Brancalhão, Rose Meire Costa; Ribeiro, Lucinéia de Fátima Chasko; Natali, Maria Raquel Marçal

2016-01-01

Silk sericin is a natural polymer produced by silkworm, Bombyx mori , which surrounds and keeps together two fibroin filaments in silk thread used in the cocoon. The recovery and reuse of sericin usually discarded by the textile industry not only minimizes environmental issues but also has a high scientific and commercial value. The physicochemical properties of the molecule are responsible for numerous applications in biomedicine and are influenced by the extraction method and silkworm lineage, which can lead to variations in molecular weight and amino acid concentration of sericin. The presence of highly hydrophobic amino acids and its antioxidant potential make it possible for sericin to be applied in the food and cosmetic industry. The moisturizing power allows indications as a therapeutic agent for wound healing, stimulating cell proliferation, protection against ultraviolet radiation, and formulating creams and shampoos. The antioxidant activity associated with low digestibility of sericin that expands the application in the medical field, such as antitumour, antimicrobial and anti-inflammatory agent, anticoagulant, acts in colon health, improving constipation and protects the body from obesity through improved plasma lipid profile. In addition, the properties of sericin allow its application as a culture medium and cryopreservation, in tissue engineering and for drug delivery, demonstrating its effective use, as an important biomaterial.

Silkworm Sericin: Properties and Biomedical Applications

PubMed Central

Ribeiro, Lucinéia de Fátima Chasko

2016-01-01

Silk sericin is a natural polymer produced by silkworm, Bombyx mori, which surrounds and keeps together two fibroin filaments in silk thread used in the cocoon. The recovery and reuse of sericin usually discarded by the textile industry not only minimizes environmental issues but also has a high scientific and commercial value. The physicochemical properties of the molecule are responsible for numerous applications in biomedicine and are influenced by the extraction method and silkworm lineage, which can lead to variations in molecular weight and amino acid concentration of sericin. The presence of highly hydrophobic amino acids and its antioxidant potential make it possible for sericin to be applied in the food and cosmetic industry. The moisturizing power allows indications as a therapeutic agent for wound healing, stimulating cell proliferation, protection against ultraviolet radiation, and formulating creams and shampoos. The antioxidant activity associated with low digestibility of sericin that expands the application in the medical field, such as antitumour, antimicrobial and anti-inflammatory agent, anticoagulant, acts in colon health, improving constipation and protects the body from obesity through improved plasma lipid profile. In addition, the properties of sericin allow its application as a culture medium and cryopreservation, in tissue engineering and for drug delivery, demonstrating its effective use, as an important biomaterial. PMID:27965981

Composite biodegradable biopolymer coatings of silk fibroin - Poly(3-hydroxybutyric-acid-co-3-hydroxyvaleric-acid) for biomedical applications

NASA Astrophysics Data System (ADS)

Miroiu, Floralice Marimona; Stefan, Nicolaie; Visan, Anita Ioana; Nita, Cristina; Luculescu, Catalin Romeo; Rasoga, Oana; Socol, Marcela; Zgura, Irina; Cristescu, Rodica; Craciun, Doina; Socol, Gabriel

2015-11-01

Composite silk fibroin-poly(3-hydroxybutyric-acid-co-3-hydroxyvaleric-acid) (SF-PHBV) biodegradable coatings were grown by Matrix Assisted Pulsed Laser Evaporation on titanium substrates. Their physico-chemical properties and particularly the degradation behavior in simulated body fluid at 37 °C were studied as first step of applicability in local controlled release for tissue regeneration applications. SF and PHBV, natural biopolymers with excellent biocompatibility, but different biodegradability and tensile strength properties, were combined in a composite to improve their properties as coatings for biomedical uses. FTIR analyses showed the stoichiometric transfer from targets to coatings by the presence in the spectra of the main absorption maxima characteristic of both polymers. XRD investigations confirmed the FTIR results showing differences in crystallization behavior with respect to the SF and PHBV content. Contact angle values obtained through wettability measurements indicated the MAPLE deposited coatings were highly hydrophilic; surfaces turning hydrophobic with the increase of the PHBV component. Degradation assays proved that higher PHBV contents resulted in enhanced resistance and a slower degradation rate of composite coatings in SBF. Distinct drug-release schemes could be obtained by adjusting the SF:PHBV ratio to controllably tuning the coatings degradation rate, from rapid-release formulas, where SF predominates, to prolonged sustained ones, for larger PHBV content.