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Preparation of Porous Scaffolds from Silk Fibroin Extracted from the Silk Gland of Bombyx mori (B. mori  

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Full Text Available In order to use a simple and ecofriendly method to prepare porous silk scaffolds, aqueous silk fibroin solution (ASF was extracted from silk gland of 7-day-old fifth instar larvae of Bombyx mori (B. mori. SDS-page analysis indicated that the obtained fibroin had a molecular weight higher than 200 kDa. The fabrication of porous scaffolds from ASF was achieved by using the freeze-drying method. The pore of porous scaffolds is homogenous and tends to become smaller with an increase in the concentration of ASF. Conversely, the porosity is decreased. The porous scaffolds show impressive compressive strength which can be as high as 6.9 ± 0.4 MPa. Furthermore, ASF has high cell adhesion and growth activity. It also exhibits high ALP activity. This implies that porous scaffolds prepared from ASF have biocompatibility. Therefore, the porous scaffolds prepared in this study have potential application in tissue engineering due to the impressive compressive strength and biocompatibility.

Liangjun Zhu

2012-06-01

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Macroporous silk fibroin cryogels.  

Science.gov (United States)

Silk fibroin cryogels with remarkable properties were obtained from frozen fibroin solutions (4.2-12.6%) at subzero temperatures between -5 and -22 °C. This was achieved by the addition of ethylene glycol diglycidyl ether (EGDE) into the cryogelation system. EGDE triggers the conformational transition of fibroin from random coil to ?-sheet structure and hence fibroin gelation. One of the unique features of fibroin cryogels is their elasticity that allows them to resist complete compression without any crack development, during which water inside the cryogel is removed. The compressed cryogel immediately swells during unloading to recover its original shape. The scaffolds obtained by freeze-drying of the cryogels consist of regular, interconnected pores of diameters ranging from 50 to 10 ?m that could be regulated by the synthesis parameters. The mechanical compressive strength and the modulus of the scaffolds increase with decreasing pore diameter, that is, with decreasing gelation temperature or, with increasing fibroin or EGDE concentrations in the feed. The scaffolds produced at 12.6% fibroin exhibit a very high compressive modulus (50 MPa) making them good candidates as bone scaffold materials. PMID:23360211

Ak, Fatih; Oztoprak, Zeynep; Karakutuk, Ilknur; Okay, Oguz

2013-03-11

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13C NMR study of silk fibroin synthesis in vitro  

International Nuclear Information System (INIS)

The posterior silkgland of Bombyx mori, silkworm larvae, produces abundant specific silk fibroin at about 80 ?g/cell for one day. In this study, the continuous production of silk fibroin in the posterior silk-gland was monitored by circumfusion cultivation using both C-13 NMR and C-13 labeling techniques in order to quantitatively determined the production of silk fibroin in vitro. The experimental setup is shown. The posterior silkglands of four larvae were collected in a 10 mm NMR sample tube under sterile condition. The C-13 NMR spectra were observed in the culture medium. The time-dependence of the C-13 NMR spectra of the culture medium containing the posterior silkglands are shown. The peak intensity for silk fibroin increased with the lapse of time. Also the peak due to the C? carbon of the serine residue of silk fibroin slightly increased, which indicates that there is a system which converts glycine to serine in the posterior silkglands. In order to discuss this silk production quantitatively, the relative intensity of the C? peaks for silk fibroin and glycine in the cultivation medium was plotted. The ability of the posterior silk-glands to produce silk protein was still high in vitro when the circumfusion cultivation technique was used. (K.I.)

1990-01-01

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Genipin modified silk fibroin nanometric-nets  

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Nanometric silk-fibroin nets were fabricated by electrospinning from regenerated Bombyx mori silk-fibroin (SF)/formic acid solutions with the addition of genipin (GE), 2, 15 and 24 h after the solution preparation. After spinning, the pure fibroin nanofibers were water soluble and needed a further stabilization process, whereas the reaction of fibroin with genipin resulted in water-insoluble fibroin nets due to conformational changes induced in the fibroin by the genipin. SFGE nanofibers pres...

Silva, S. S.; Maniglio, D.; Motta, A.; Mano, J. F.; Reis, R. L.; Migliaresi, C.

2008-01-01

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The nucleotide sequence of two silk gland alanine tRNAs: implications for fibroin synthesis and for initiator tRNA structure.  

Science.gov (United States)

The nucleotide sequences of two major alanine tRNAs from the Bombyx mori posterior silk gland have been determined. One of these tRNAs appears to be specific to the silk gland, where its accumulation is associated with the rapid production of fibroin. Both sequences are identical, with the exception of a single nucleotide in the anticodon stem. A striking feature of both alanine tRNAs is that loop IV contains sequences previously believed to be restricted to initiator tRNA. PMID:884735

Sprague, K U; Hagenbüchle, O; Zuniga, M C

1977-07-01

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Silk Fibroin Based Porous Materials  

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Full Text Available 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.

Shuqin Yan

2009-12-01

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Thermal crystallization mechanism of silk fibroin protein  

Science.gov (United States)

In this thesis, the thermal crystallization mechanism of silk fibroin protein from Bombyx mori silkworm, was treated as a model for the general study of protein based materials, combining theories from both biophysics and polymer physics fields. A systematic and scientific path way to model the dynamic beta-sheet crystallization process of silk fibroin protein was presented in the following sequence: (1) The crystallinity, fractions of secondary structures, and phase compositions in silk fibroin proteins at any transition stage were determined. Two experimental methods, Fourier transform infrared spectroscopy (FTIR) with Fourier self-deconvolution, and specific reversing heat capacity, were used together for the first time for modeling the static structures and phases in the silk fibroin proteins. The protein secondary structure fractions during the crystallization were quantitatively determined. The possibility of existence of a "rigid amorphous phase" in silk protein was also discussed. (2) The function of bound water during the crystallization process of silk fibroin was studied using heat capacity, and used to build a silk-water dynamic crystallization model. The fundamental concepts and thermal properties of silk fibroin with/without bound water were discussed. Results show that intermolecular bound water molecules, acting as a plasticizer, will cause silk to display a water-induced glass transition around 80°C. During heating, water is lost, and the change of the microenvironment in the silk fibroin chains induces a mesophase prior to thermal crystallization. Real time FTIR during heating and isothermal holding above Tg show the tyrosine side chain changes only during the former process, while beta sheet crystallization occurs only during the latter process. Analogy is made between the crystallization of synthetic polymers according to the four-state scheme of Strobl, and the crystallization process of silk fibroin, which includes an intermediate precursor stage before crystallization. (3) The beta-sheet crystallization kinetics in silk fibroin protein were measured using X-ray, FTIR and heat flow, and the structure reveals the formation mechanism of the silk crystal network. Avrami kinetics theories, which were established for studies of synthetic polymer crystal growth, were for the first time extended to investigate protein self-assembly in multiblock silk fibroin samples. The Avrami exponent, n, was close to two for all methods, indicating formation of beta sheet crystals in silk proteins is different from the 3-D spherulitic crystal growth found in most synthetic homopolymers. A microphase separation pattern after chymotrypsin enzyme biodegradation was shown in the protein structures using scanning electron microscopy. A model was then used to explain the crystallization of silk fibroin protein by analogy to block copolymers. (4) The effects of metal ions during the crystallization of silk fibroin was investigated using thermal analysis. Advanced thermal analysis methods were used to analyze the thermal protein-metallic ion interactions in silk fibroin proteins. Results show that K+ and Ca2+ metallic salts play different roles in silk fibroin proteins, which either reduce (K+) or increase (Ca2+ ) the glass transition (Tg) of pure silk protein and affect the thermal stability of this structure.

Hu, Xiao

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Structural analysis and application to biomaterials of the silk fibroins  

International Nuclear Information System (INIS)

Silk fibroin from Bombyx mori silkworm has outstanding mechanical properties despite being spun from aqueous solution. I have clarified two distinct structures in the solid state; silk I and silk II, which mean the structures before and after spinning, by using solid state NMR. Moreover, I have been developing several kinds of biomaterials, such as bone regeneration materials and vascular grafts. In this paper, I present two topics: one is the structural analyses of the silk fibroin in detail, the other is applications of silk fibroins to tissue engineering. In the case of vascular regeneration, I have developed the small diameter vascular grafts made by silk fibroins. The new grafts from silk fibroins have good patency, and these grafts were commonly covered with cells and platelets at 4 weeks after implantation. For bone tissue engineering, I performed structural analyses of a new silk-like peptide, En(AGSGAG)4, in order to consider the molecular design of biomaterials for bone regeneration. (author)

2010-08-01

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Laser Ablation of Silk Protein (Fibroin) Films  

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Fibroin is the main protein component of silk and is expected to have functional applications in bioelectronics and medicine. We investigated nanosecond (ns) pulsed laser ablation of solid fibroin films with/without a dye as a photosensitizer. Laser lights at 248 nm and 532/355/351 nm excited the peptide bond of fibroin and the dye, respectively. The neat film irradiated at 248 nm was scarcely accessible to etching and swelling, and instead, a microscopic pattern (structure) was formed. In contrast, for ablation of the doped film at 532/355/351 nm, we found marked swelling (height ˜500 ?m) and deep etching (depth ˜10 ?m) on the irradiated surfaces. The dye-photosensitized ablation was brought about by a photothermal mechanism, whereas ablation of neat films may be induced by another process, such as a photochemical one. The ablation processes are discussed in terms of the properties of fibroin and the mode of excitation.

Tsuboi, Yasuyuki; Adachi, Hisanori; Yamada, Kazushi; Miyasaka, Hiroshi; Itaya, Akira

2002-07-01

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Green process to prepare silk fibroin/gelatin biomaterial scaffolds.  

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A new all-aqueous and green process is described to form three-dimensional porous silk fibroin matrices with control of structural and morphological features. Silk-based scaffolds are prepared using lyophilization. Gelatin is added to the aqueous silk fibroin solution to change the silk fibroin conformation and silk fibroin-water interactions through adjusting the hydrophilic interactions in silk fibroin-gelatin-water systems to restrain the formation of separate sheet like structures in the material, resulting in a more homogenous structure. Water annealing is used to generate insolubility in the silk fibroin-gelatin scaffold system, thereby avoiding the use of organic solvents such as methanol to lock in the beta-sheet structure. The adjusting of the concentration of gelatin, as well as the concentration of silk fibroin, leads to control of morphological and functional properties of the scaffolds. The scaffolds were homogeneous in terms of interconnected pores, with pore sizes ranging from 100 to 600 microm, depending on the concentration of silk fibroin used in the process. At the same time, the morphology of the scaffolds changed from lamellar sheets to porous structures based on the increase in gelatin content. Compared with salt-leaching aqueous-derived scaffolds and hexafluoroisopropanol (HFIP)-derived scaffolds, these freeze-dried scaffolds had a lower content of beta-sheet, resulting in more hydrophilic features. Most of gelatin was entrapped in the silk fibroin-gelatin scaffolds, without the burst release in PBS solution. During in vitro cell culture, these silk fibroin-gelatin scaffolds had improved cell-compatibility than salt-leaching silk fibroin scaffolds. This new process provides useful silk fibroin-based scaffold systems for use in tissue engineering. Furthermore, the whole process is green, including all-aqueous, room temperature and pressure, and without the use of toxic chemicals or solvents, offering new ways to load bioactive drugs or growth factors into the process. PMID:19924684

Lu, Qiang; Zhang, Xiaohui; Hu, Xiao; Kaplan, David L

2010-03-10

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Rheology of Self-Assembling Silk Fibroin Solutions  

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A robust procedure for preparation of aqueous silk fibroin solutions with a range of concentration up to 25 wt% from domestic Bombyx mori cocoon shells has been established. We have carried out molecular and rheometric characterizations of silk fibroin solutions, and constructed an equilibrium phase diagram. The sharp sol-gel transition can be exploited for rapid solidification of micro-morphological structure. We will discuss the correlations between fluid formulation, rheological properties and processibility of silk fibroin in the talk.

Zhou, Rui; Chen, Song-Bi; Yuan, Xue-Feng

2008-07-01

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Ras1CA overexpression in the posterior silk gland improves silk yield  

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Sericulture has been greatly advanced by applying hybrid breeding techniques to the domesticated silkworm, Bombyx mori, but has reached a plateau during the last decades. For the first time, we report improved silk yield in a GAL4/UAS transgenic silkworm. Overexpression of the Ras1CA oncogene specifically in the posterior silk gland improved fibroin production and silk yield by 60%, while increasing food consumption by only 20%. Ras activation by Ras1CA overexpression in the posterior silk gl...

2011-01-01

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Novel silk fibroin/elastin wound dressings  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Silk fibroin (SF) and elastin (EL) scaffolds were successfully produced for the first time for the treatment of burn wounds. The self-assembly properties of SF, together with the excellent chemical and mechanical stability and biocompatibility, were combined with elastin protein to produce scaffolds with the ability to mimic the extracellular matrix (ECM). Porous scaffolds were obtained by lyophilization and were further crosslinked with genipin (GE). Genipin crosslinking induces the conforma...

Vasconcelos, Andreia; Gomes, Andreia; Paulo, Artur Cavaco

2012-01-01

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The degradation behavior of silk fibroin derived from different ionic liquid solvents  

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Establishing an appropriate degradation rate is critical for tissue engineering scaffolds. In this study, the degradation rate of silk fibroin three-dimensional scaffolds was regulated by changing the molecular weight (MW) of the silk fibroin. The solubility of silk fibroin depends primarily on the ionic ability of the slovent to dissolve silk fibroin, therefore, we regulated the MW of the silk fibroin using LiBr, Ca(NO3)2 and CaCl...

2013-01-01

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Production of silk sericin/silk fibroin blend nanofibers  

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Abstract Silk sericin (SS)/silk fibroin (SF) blend nanofibers have been produced by electrospinning in a binary SS/SF trifluoroacetic acid (TFA) solution system, which was prepared by mixing 20 wt.% SS TFA solution and 10 wt.% SF TFA solution to give different compositions. The diameters of the SS/SF nanofibers ranged from 33 to 837 nm, and they showed a round cross section. The surface of the SS/SF nanofibers was smooth, and the fibers possessed a bead-free structure. The average d...

Zhang Xianhua; Tsukada Masuhiro; Morikawa Hideaki; Aojima Kazuki; Zhang Guangyu; Miura Mikihiko

2011-01-01

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Sonication-Induced Gelation of Silk Fibroin for Cell Encapsulation  

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Purified native silk fibroin forms ?-sheet-rich, physically crosslinked, hydrogels from aqueous solution, in a process influenced by environmental parameters. Previously we reported gelation times of days to weeks for aqueous native silk protein solutions, with high ionic strength and temperature and low pH responsible for increasing gelation kinetics. Here we report a novel method to accelerate the process and control silk fibroin gelation through ultrasonication. Depending on the sonicatio...

2008-01-01

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Controlling silk fibroin particle features for drug delivery  

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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. Se...

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

2010-01-01

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A novel electrospun silk fibroin/hydroxyapatite hybrid nanofibers  

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A novel electrospinning of silk fibroin/hydroxyapatite hybrid nanofibers with different composition ratios was performed with methanoic acid as a spinning solvent. The silk fibroin/hydroxyapatite hybrids containing up to 30% hydroxyapatite nanoparticles could be electrospun into the continuous fibrous structure. The electrospun silk fibroin/hydroxyapatite hybrid nanofibers showed bigger diameter and wider diameter distribution than pure silk fibroin nanofibers, and the average diameter gradually increased from 95 to 582 nm. At the same time, the secondary structure of silk fibroin/hydroxyapatite nanofibers was characterized by X-ray diffraction, Fourier transform infrared analysis, and DSC measurement. Comparing with the pure silk fibroin nanofibers, the crystal structure of silk fibroin was mainly amorphous structure in the hybrid nanofibers. X-ray diffraction results demonstrated the hydroxyapatite crystalline nature remained as evidenced from the diffraction planes (002), (211), (300), and (202) of the hydroxyapatite crystallites, which was also confirmed by Fourier transform infrared analysis. The thermal behavior of hybrid nanofibers exhibited the endothermic peak of moisture evaporation ranging from 86 to 113 Degree-Sign C, and the degradation peak at 286 Degree-Sign C appeared. The SF/HAp nanofibers mats containing 30% HAp nanoparticles showed higher breaking tenacity and extension at break for 1.1688 {+-} 0.0398 MPa and 6.55 {+-} 1.95%, respectively. Therefore, the electrospun silk fibroin/hydroxyapatite hybrid nanofibers should be provided potentially useful options for the fabrication of biomaterial scaffolds for bone tissue engineering. -- Highlights: Black-Right-Pointing-Pointer The novel SF/HAp nanofibers were directly prepared by electrospinning method. Black-Right-Pointing-Pointer The nanofiber diameter had significant related to the content of HAp. Black-Right-Pointing-Pointer The crystal structure of silk fibroin was mainly amorphous structure in the hybrid nanofibers. Black-Right-Pointing-Pointer The HAp crystals existing in the hybrid nanofibers were characterized using XRD and FTIR.

Ming, Jinfa, E-mail: jinfa.ming@gmail.com [National Engineering Laboratory for Modern Silk, Soochow University, Suzhou 215123 (China); College of Textile and Clothing Engineering, Soochow University, Suzhou 215021 (China); Zuo, Baoqi, E-mail: bqzuo@suda.edu.cn [National Engineering Laboratory for Modern Silk, Soochow University, Suzhou 215123 (China); College of Textile and Clothing Engineering, Soochow University, Suzhou 215021 (China)

2012-11-15

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Silk tape nanostructure and silk gland anatomy of trichoptera.  

Science.gov (United States)

Caddisflys (order Trichoptera) construct elaborate protective shelters and food harvesting nets with underwater adhesive silk. The silk fiber resembles a nanostructured tape composed of thousands of nanofibrils (? 120 nm) oriented with the major axis of the fiber, which in turn are composed of spherical subunits. Weaker lateral interactions between nanofibrils allow the fiber to conform to surface topography and increase contact area. Highly phosphorylated (pSX)(4) motifs in H-fibroin blocks of positively charged basic residues are conserved across all three suborders of Trichoptera. Electrostatic interactions between the oppositely charged motifs could drive liquid-liquid phase separation of silk fiber precursors into a complex coacervates mesophase. Accessibility of phosphoserine to an anti-phosphoserine antibody is lower in the lumen of the silk gland storage region compared to the nascent fiber formed in the anterior conducting channel. The phosphorylated motifs may serve as a marker for the structural reorganization of the silk precursor mesophase into strongly refringent fibers. The structural change occurring at the transition into the conducting channel makes this region of special interest. Fiber formation from polyampholytic silk proteins in Trichoptera may suggest a new approach to create synthetic silk analogs from water-soluble precursors. PMID:21953029

Ashton, Nicholas N; Taggart, Daniel S; Stewart, Russell J

2012-06-01

20

In vivo Degradation of Three-Dimensional Silk Fibroin Scaffolds  

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Three-dimensional porous scaffolds prepared from regenerated silk fibroin using either an all aqueous process or a process involving an organic solvent, hexafluoroisopropanol (HFIP) have shown promise in cell culture and tissue engineering applications. However, their biocompatibility and in vivo degradation has not been fully established. The present study was conducted to systematically investigate how processing method (aqueous vs. organic solvent) and processing variables (silk fibroin co...

2008-01-01

 
 
 
 
21

Impact of Protein-Metal Ion Interactions on the Crystallization of Silk Fibroin Protein  

Science.gov (United States)

Proteins can easily form bonds with a variety of metal ions, which provides many unique biological functions for the protein structures, and therefore controls the overall structural transformation of proteins. We use advanced thermal analysis methods such as temperature modulated differential scanning calorimetry and quasi-isothermal TMDSC, combined with Fourier transform infrared spectroscopy, and scanning electron microscopy, to investigate the protein-metallic ion interactions in Bombyx mori silk fibroin proteins. Silk samples were mixed with different metal ions (Ca^2+, K^+, Ma^2+, Na^+, Cu^2+, Mn^2+) with different mass ratios, and compared with the physical conditions in the silkworm gland. Results show that all metallic ions can directly affect the crystallization behavior and glass transition of silk fibroin. However, different ions tend to have different structural impact, including their role as plasticizer or anti-plasticizer. Detailed studies reveal important information allowing us better to understand the natural silk spinning and crystallization process.

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

2009-03-01

22

In vivo NMR analysis of incorporation of [2-13C] glycine into silk fibroin  

International Nuclear Information System (INIS)

The biosynthetic mechanism of silk fibroin in silkworms, Bombyx mori, is unique because this fibrous protein composed mainly of glycine, alanine and serine is produced very rapidly in large quantity in the posterior silk glands. It is very meaningful to investigate into the biosynthesis of silk protein under nondestructive condition by in vivo NMR and C-13 labeling techniques. The sugar metabolism related to the production of silk fibroin was analyzed by monitoring the change in the C-13 labeled peaks in the NMR spectra for silkworms. In this paper, the monitoring of the 2-(C-13) glycine metabolism in Bombyx mori by the C-13 NMR in vivo is reported. In particular, the in vivo transport of glycine from the midgut to the posterior silk gland was measured, and the rate constants were determined with the course of the peak intensity in the C-13 NMR spectra. It is possible to discuss quantitatively the in vivo production of silk fibroin with these rate constants. The experiment and the results are reported. The in vivo C-13 NMR spectra of a 5 day old, 5th instar larva of Bombyx mori after the oral administration of 2-(C-13) glycine are shown. The significant increase of the peak intensity occurred. (K.I.)

1990-01-01

23

Silk protein fibroin from Antheraea mylitta for cardiac tissue engineering.  

Science.gov (United States)

The human heart cannot regenerate after an injury. Lost cardiomyocytes are replaced by scar tissue resulting in reduced cardiac function causing high morbidity and mortality. One possible solution to this problem is cardiac tissue engineering. Here, we have investigated the suitability of non-mulberry silk protein fibroin from Indian tropical tasar Antheraea mylitta as a scaffold for engineering a cardiac patch in vitro. We have tested cell adhesion, cellular metabolic activity, response to extracellular stimuli, cell-to-cell communication and contractility of 3-days postnatal rat cardiomyocytes on silk fibroin. Our data demonstrate that A. mylitta silk fibroin exhibits similar properties as fibronectin, a component of the natural matrix for cardiomyocytes. Comparison to mulberry Bombyx mori silk protein fibroin shows that A. mylitta silk fibroin is superior probably due to its RGD domains. 3D scaffolds can efficiently be loaded with cardiomyocytes resulting in contractile patches. In conclusion, our findings demonstrate that A. mylitta silk fibroin 3D scaffolds are suitable for the engineering of cardiac patches. PMID:22240510

Patra, Chinmoy; Talukdar, Sarmistha; Novoyatleva, Tatyana; Velagala, Siva R; Mühlfeld, Christian; Kundu, Banani; Kundu, Subhas C; Engel, Felix B

2012-03-01

24

Effect of ?-irradiation on the crystalline structure of silk fibroin and silk sericin  

International Nuclear Information System (INIS)

Changes in the crystalline structure of silk sericin and silk fibroin induced by gamma-irradiation in the atmosphere described. The crystalline structure of silk sericin which had been subjected to gamma-irradiation remained unchanged. However the decomposition temperature of the specimen decreased to about 230 deg C, when the total dose of ? rays exceeded 4.6 Mrad. The structure of the silk 1 type crystal of silk fibroin in the solid state, with a low degree of molecular orientation, changed into the silk 2 type crystal, when the total dose of ? rays exceeded 4.6 Mrad. No changes in the crystalline structure were observed in the solid state of the silk 2 type crystal regardless of gamma-irradiation. The decrease in the decomposition temperature of the specimen was attributed to the decrease in the molecular orientation. However, the molecular conformation of silk fibroin with a randomly coiled structure remained unchanged even after gamma-irradiation. (author)

1985-01-01

25

Protein-protein nanoimprinting of silk fibroin films.  

Science.gov (United States)

Protein-protein imprinting of silk fibroin is introduced as a rapid, high-throughput method for the fabrication of nanoscale structures in silk films, through the application of heat and pressure. Imprinting on conformal surfaces is demonstrated with minor adjustments to the system, at resolutions comparable to other currently available nonplanar nanoimprint lithography techniques. PMID:23483712

Brenckle, Mark A; Tao, Hu; Kim, Sunghwan; Paquette, Mark; Kaplan, David L; Omenetto, Fiorenzo G

2013-05-01

26

Microdissection of black widow spider silk-producing glands.  

Science.gov (United States)

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. PMID:21248709

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

2011-01-01

27

Production of silk sericin/silk fibroin blend nanofibers  

Directory of Open Access Journals (Sweden)

Full Text Available Abstract Silk sericin (SS/silk fibroin (SF blend nanofibers have been produced by electrospinning in a binary SS/SF trifluoroacetic acid (TFA solution system, which was prepared by mixing 20 wt.% SS TFA solution and 10 wt.% SF TFA solution to give different compositions. The diameters of the SS/SF nanofibers ranged from 33 to 837 nm, and they showed a round cross section. The surface of the SS/SF nanofibers was smooth, and the fibers possessed a bead-free structure. The average diameters of the SS/SF (75/25, 50/50, and 25/75 blend nanofibers were much thicker than that of SS and SF nanofibers. The SS/SF (100/0, 75/25, and 50/50 blend nanofibers were easily dissolved in water, while the SS/SF (25/75 and 0/100 blend nanofibers could not be completely dissolved in water. The SS/SF blend nanofibers could not be completely dissolved in methanol. The SS/SF blend nanofibers were characterized by Fourier transform infrared (FTIR spectroscopy, differential scanning calorimetry, and differential thermal analysis. FTIR showed that the SS/SF blend nanofibers possessed a random coil conformation and ß-sheet structure.

Zhang Xianhua

2011-01-01

28

Silk fibroin-based scaffolds for bone regeneration.  

Science.gov (United States)

Porous scaffolds were prepared using regenerated Bombyx mori silk fibroin dissolved in water or hexafluoroisopropanol (HFIP). The effects of these two preparations on the formation and growth of new bone on implantation into the rabbit femoral epicondyle was examined. The aqueous-based fibroin (A-F) scaffold exhibited significantly greater osteoconductivity as judged by bone volume, bone mineral content, and bone mineral density at the implant site than the HFIP-based fibroin (HFIP-F) scaffold. Micro-CT analyses showed that the morphology of the newly formed bone differed significantly in the two types of silk fibroin scaffold. After 4 weeks of implantation, new trabecular bone was seen inside the pores of the A-F scaffold implant while the HFIP-F scaffold only contained necrotic cells. No trabecular bone was seen within the pores of the latter scaffolds, although the pores of these did contain giant cells and granulation tissue. PMID:23125151

Kuboyama, Noboru; Kiba, Hideo; Arai, Kiyoshi; Uchida, Ryoichiro; Tanimoto, Yasuhiro; Bhawal, Ujjal K; Abiko, Yoshimitsu; Miyamoto, Sayaka; Knight, David; Asakura, Tetsuo; Nishiyama, Norihiro

2013-02-01

29

Folate conjugated silk fibroin nanocarriers for targeted drug delivery.  

Science.gov (United States)

Disease treatment processes mainly focus on the development of nontoxic, biodegradable, non-immunogenic, biocompatible materials capable of controlled and long-term release of biomolecules. In this work silk protein fibroin from non-mulberry tropical tasar silkworm, Antheraea mylitta, is used to prepare nanoparticles as a drug delivery system. Folate is a vitamin, which is brought into healthy and cancerous cells by folate receptors. The efficiency of silk fibroin-folate nanoparticles loaded with anticancer drug doxorubicin was evaluated by analysing the cell viability, proliferation and endocytosis. Consequently the effects of pro-inflammatory responses by cytokines such as TNF-?, IL-1? and nitric oxide were checked by stimulating the macrophages with folate conjugated silk fibroin nanoparticles. The fibroin-folate nanocarriers are nontoxic, easily taken up by cells and capable of sustained drug release. Nanoparticles loaded with anticancer drug doxorubicin target cancer cells. The results show that silk fibroin-folate nanoparticles may serve as promising nanocarriers for different biomedical and nanotechnology applications in cancer research. PMID:24345855

Subia, Bano; Chandra, Sourov; Talukdar, Sarmistha; Kundu, Subhas C

2014-02-01

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Novel silk fibroin/elastin wound dressings.  

Science.gov (United States)

Silk fibroin (SF) and elastin (EL) scaffolds were successfully produced for the first time for the treatment of burn wounds. The self-assembly properties of SF, together with the excellent chemical and mechanical stability and biocompatibility, were combined with elastin protein to produce scaffolds with the ability to mimic the extracellular matrix (ECM). Porous scaffolds were obtained by lyophilization and were further crosslinked with genipin (GE). Genipin crosslinking induces the conformational transition from random coil to ?-sheet of SF chains, yielding scaffolds with smaller pore size and reduced swelling ratios, degradation and release rates. All results indicated that the composition of the scaffolds had a significant effect on their physical properties, and that can easily be tuned to obtain scaffolds suitable for biological applications. Wound healing was assessed through the use of human full-thickness skin equivalents (EpidermFT). Standardized burn wounds were induced by a cautery and the best re-epithelialization and the fastest wound closure was obtained in wounds treated with 50SF scaffolds; these contain the highest amount of elastin after 6 days of healing in comparison with other dressings and controls. The cytocompatibility demonstrated with human skin fibroblasts together with the healing improvement make these SF/EL scaffolds suitable for wound dressing applications. PMID:22546517

Vasconcelos, Andreia; Gomes, Andreia C; Cavaco-Paulo, Artur

2012-08-01

31

Controlling silk fibroin particle features for drug delivery.  

Science.gov (United States)

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 kosmotropic 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

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

2010-06-01

32

Conductive Au nanowires regulated by silk fibroin nanofibers  

Science.gov (United States)

Conductive Au-biopolymer composites have promising applications in tissue engineering such as nerve tissue regeneration. In this study, silk fibroin nanofibers were formed in aqueous solution by regulating silk self-assembly process and then used as template for Au nanowire fabrication. We performed the synthesis of Au seeds by repeating the seeding cycles for several times in order to increase the density of Au seeds on the nanofibers. After electroless plating, densely decorated Au seeds grew into irregularly shaped particles following silk nanofiber to fill the gaps between particles and finally form uniform continuous nanowires. The conductive property of the Au-silk fibroin nanowires was studied with current-voltage ( I-V) measurement. A typical ohmic behavior was observed, which highlighted their potential applications in nerve tissue regeneration.

Dong, Bo-Ju; Lu, Qiang

2014-03-01

33

Mechanisms of Controlled Release from Silk Fibroin Films  

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The controlled release of fluorescein-iso-thio-cyanate (FITC) labeled dextrans from methanol treated and untreated silk fibroin films was modeled to characterize the release kinetics and mechanisms. Silk films were prepared with FITC-dextrans of various molecular weights (4, 10, 20, 40 kDa). Methanol treatment was used to promote crystallinity. The release data were assessed with two different models, an empirical exponential equation commonly fit to release data and a mechanism based semi-em...

Hines, Daniel J.; Kaplan, David L.

2011-01-01

34

Sonication-induced gelation of silk fibroin for cell encapsulation.  

Science.gov (United States)

Purified native silk fibroin forms beta-sheet-rich, physically cross-linked, hydrogels from aqueous solution, in a process influenced by environmental parameters. Previously we reported gelation times of days to weeks for aqueous native silk protein solutions, with high ionic strength and temperature and low pH responsible for increasing gelation kinetics. Here we report a novel method to accelerate the process and control silk fibroin gelation through ultrasonication. Depending on the sonication parameters, including power output and time, along with silk fibroin concentration, gelation could be controlled from minutes to hours, allowing the post-sonication addition of cells prior to final gel setting. Mechanistically, ultrasonication initiated the formation of beta-sheets by alteration in hydrophobic hydration, thus accelerating the formation of physical cross-links responsible for gel stabilization. K(+) at physiological concentrations and low pH promoted gelation, which was not observed in the presence of Ca(2+). The hydrogels were assessed for mechanical properties and proteolytic degradation; reported values matched or exceeded other cell-encapsulating gel material systems. Human bone marrow derived mesenchymal stem cells (hMSCs) were successfully incorporated into these silk fibroin hydrogels after sonication, followed by rapid gelation and sustained cell function. Sonicated silk fibroin solutions at 4%, 8%, and 12% (w/v), followed by mixing in hMSCs, gelled within 0.5-2 h. The cells grew and proliferated in the 4% gels over 21 days, while survival was lower in the gels with higher protein content. Thus, sonication provides a useful new tool with which to initiate rapid sol-gel transitions, such as for cell encapsulation. PMID:18031805

Wang, Xiaoqin; Kluge, Jonathan A; Leisk, Gary G; Kaplan, David L

2008-03-01

35

Development of a bilayered scaffold based on silk fibroin and silk fibroin/nano-calcium phosphate for osteochondral regeneration  

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Objectives: Osteochondral defect is a common condtion in clinic. Satisfactory outcomes are rarely achieved by traditional methods. Tissue engineering might be a promising strategy for this hinder. The aim of this study is to mimick the stratified structure of osteochondral tissue, by developing a bilayered scaffold for osteochondral regeneration. The developed bilayered scaffold is composed of a porous silk fibroin scaffold as the cartilage-like layer and a porous silk fibro...

Yan, Leping; Oliveira, J. M.; Oliveira, A. L.; Reis, R. L.

2012-01-01

36

The Cytocompatibility of Genipin-Crosslinked Silk Fibroin Films  

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There is an increasing demand for crosslinking methods of silk fibroin (SF) scaffolds in biomedical applications that could maintain the biocompatibility, bioactivity as well as improve the water resistance and mechanical properties of SF materials. In this study, SF was crosslinked effectively with genipin which is a naturally occurring iridoid glucoside and the crosslinking mechanism was investigated through FTIR and amino acid analys...

Lingshuang Wang; Yiyu Wang; Jing Qu; Yongpei Hu; Renchuan You; Mingzhong Li

2013-01-01

37

Gelation of Antheraea pernyi Silk Fibroin Accelerated by Shearing  

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The rapid manufacture of silk fibroin gels in mild conditions is an important subject in the field of silk-based biomaterials. In this study, the gelation of Antheraea pernyi silk fibroin (ASF) aqueous solution was induced by shearing, without chemical cross-link...

2013-01-01

38

A novel growth process of calcium carbonate crystals in silk fibroin hydrogel system  

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We report an interesting finding of calcium carbonate (CaCO3) crystal growth in the silk fibroin (SF) hydrogel with different concentrations by a simple ion diffusion method. The experimental results indicate that the CaCO3 crystals obtained from silk fibroin gels with low and high concentrations are all calcites with unusual morphologies. Time-dependent growth study was carried out to investigate the crystallization process. It is believed that silk fibroin hydrogel plays an important role i...

Ma, Yufei; Feng, Qingling; Bourrat, Xavier

2013-01-01

39

Gelation of Antheraea pernyi Silk Fibroin Accelerated by Shearing  

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Full Text Available The rapid manufacture of silk fibroin gels in mild conditions is an important subject in the field of silk-based biomaterials. In this study, the gelation of Antheraea pernyi silk fibroin (ASF aqueous solution was induced by shearing, without chemical cross-linking agents. Simple shearing controlled and accomplished the steady and rapid conformational transition to ?-sheets with ease. The conformational transformation and rapid gelation mechanisms of ASF induced by shearing were tracked and analyzed by circular dichroism spectrometry, Fourier transform infrared spectroscopy and X-ray diffractometry, then compared with Bombyx mori silk fibroin (BSF. ASF quickly formed hydrogels within 24 - 48 h after shearing under different shearing rates for 30 - 90 min, resulting in sol-gel transformation when the ?-sheet content reached nearly 50%, which is the minimum content needed to maintain a stable hydrogel system in ASF. The gel structures remained stable once formed. The rapid gelation of ASF through shearing compared with BSF was achieved because of ASF’s alternating polyalanine-containing units, which tend to form ?-helix structures spontaneously. Further, the entropic cost during the conformational transition from the ?-helix to the ?-sheet structure is less than the cost of the transition from the random coil structure. This method is a simple, non-chemical cross-linking approach for the promotion of rapid gelation and the protection of the biological properties of ASF, and it may prove useful for application in the field of biomedical materials.

Yu Liu

2013-05-01

40

Determination of multiphoton absorption of silk fibroin using the Z-scan technique.  

Science.gov (United States)

The multiphoton absorption of silk fibroin at 810 nm was determined by open-aperture Z-scan. Three-photon absorption was confirmed at this wavelength and the three-photon cross section of silk fibroin was measured. Silk fibroin of varying molecular weight was tested and the cross section was found to increase exponentially with increasing molecular weight. The confirmation of a relatively large three-photon absorption cross section in silk will help lay the groundwork for future investigation of direct laser writing of three-dimensional structures within a silk matrix. PMID:24514515

Applegate, Matthew B; Marelli, Benedetto; Kaplan, David L; Omenetto, Fiorenzo G

2013-12-01

 
 
 
 
41

Nanolayer Biomaterial Coatings of Silk Fibroin for Controlled Release  

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An all-aqueous, stepwise deposition process with silk fibroin protein for the assembly of nanoscale layered controlled release coatings was exploited. Model compounds, Rhodamine B, Even Blue and Azoalbumin, representing small molecule drugs and therapeutically relevant proteins were incorporated in the nanocoating process and their loading and release behavior was quantified. In addition, the structure and morphology of the coatings were characterized. Release studies in vitro showed that con...

Wang, Xianyan; Hu, Xiao; Daley, Andrea; Rabotyagova, Olena; Cebe, Peggy; Kaplan, David L.

2007-01-01

42

Silk Fibroin Film Loaded Chlorhexidine Diacetate: Interaction and Characteristics  

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This study aimed to prepare Silk Fibroin (SF) films with different weight (0.5, 1 and 1.5%) for loading chlorhexidine diacetate as substrate for study their characteristics including morphology, structure and thermal properties. The morphological observation under scanning electron spectroscopy found that all of films have a smooth surface. With cross-section micrographs, the lowest SF content was smoother texture than other. The secondary structures of various films were determined by F...

Noi, N.; Yaowalak, S.; Yodthong, B.; Prasong, S.

2009-01-01

43

In vivo degradation of three-dimensional silk fibroin scaffolds.  

Science.gov (United States)

Three-dimensional porous scaffolds prepared from regenerated silk fibroin using either an all-aqueous process or a process involving an organic solvent, hexafluoroisopropanol (HFIP), have shown promise in cell culture and tissue engineering applications. However, their biocompatibility and in vivo degradation have not been fully established. The present study was conducted to systematically investigate how processing method (aqueous vs. organic solvent) and processing variables (silk fibroin concentration and pore size) affect the short-term (up to 2 months) and long-term (up to 1 year) in vivo behavior of the protein scaffolds in both nude and Lewis rats. The samples were analyzed by histology for scaffold morphological changes and tissue ingrowth, and by real-time RT-PCR and immunohistochemistry for immune responses. Throughout the period of implantation, all scaffolds were well tolerated by the host animals and immune responses to the implants were mild. Most scaffolds prepared from the all-aqueous process degraded to completion between 2 and 6 months, while those prepared from organic solvent (hexafluoroisopropanol (HFIP)) process persisted beyond 1 year. Due to widespread cellular invasion throughout the scaffold, the degradation of aqueous-derived scaffolds appears to be more homogeneous than that of HFIP-derived scaffolds. In general and especially for the HFIP-derived scaffolds, a higher original silk fibroin concentration (e.g. 17%) and smaller pore size (e.g. 100-200microm) resulted in lower levels of tissue ingrowth and slower degradation. These results demonstrate that the in vivo behavior of the three-dimensional silk fibroin scaffolds is related to the morphological and structural features that resulted from different scaffold preparation processes. The insights gained in this study can serve as a guide for processing scenarios to match desired morphological and structural features and degradation time with tissue-specific applications. PMID:18502501

Wang, Yongzhong; Rudym, Darya D; Walsh, Ashley; Abrahamsen, Lauren; Kim, Hyeon-Joo; Kim, Hyun S; Kirker-Head, Carl; Kaplan, David L

2008-01-01

44

Preparation and Characterization of Nanocomposite and Nanoporous Silk Fibroin Films  

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Nanocomposite and nanoporous silk fibroin (SF) films were prepared by film casting of SF solution containing surfactant-free colloidal nanoparticles of methoxy poly (ethylene glycol)-b-poly (D, L-lactide) diblock copolymer (MPEG-b-PDLL). Self-condensation and nanophase separation of the nanoparticles from SF film matrix during film drying process gave nanopore structures. The colloidal nanoparticles were prepared in SF solution by modified-spontaneous emulsification solven...

Yaowalak Srisuwan; Mangkorn Srisa-ard; Chaiyasit Sittiwet; Yodthong Baimark; Nual- Anong Narkkong; Chirapha Butiman

2008-01-01

45

Design, expression and solid-state NMR characterization of silk-like materials constructed from sequences of spider silk, Samia cynthia ricini and Bombyx mori silk fibroins.  

Science.gov (United States)

Silk has a long history of use in medicine as sutures. To address the requirements of a mechanically robust and biocompatible material, basic research to clarify the role of repeated sequences in silk fibroin in its structures and properties seems important as well as the development of a processing technique suitable for the preparation of fibers with excellent mechanical properties. In this study, three silk-like protein analogs were constructed from two regions selected from among the crystalline region of Bombyx mori silk fibroin, (GAGSGA)(2), the crystalline region of Samia cynthia ricini silk fibroin, (Ala)(12), the crystalline region of spider dragline silk fibroin, (Ala)(6), and the Gly-rich region of spider silk fibroin, (GGA)(4). The silk-like protein analog constructed from the crystalline regions of the spider dragline silk and B. mori silk fibroins, (A(6)SCS)(8), that constructed from the crystalline regions of the S. c.ricini and B. mori silk fibroins, (A(12)SGS)(4), that constructed from and the crystalline region of S. c.ricini silk fibroin and the glycine-rich region of spider dragline silk fibroin, (A(12)SGS)(4),were expressed their molecular weights being about 36.0 kDa, 17.0 kDa and 17.5 kDa, respectively in E. coli by means of genetic engineering technologies. (A(12)SCS)(4) and (A(12)SGS)(4 )undergo a structural transition from alpha-helix to beta-sheet on a change in the solvent treatment from trifluoroacetic acid (TFA) to formic acid (FA). However, (A(6)SCS)(8) takes on the beta-sheet structure predominantly on TFA treatment and FA treatment. Structural analysis was performed on model peptides selected from spider dragline and S. c.ricini silks by means of (13)C CP/MAS NMR. PMID:16002994

Yang, Mingying; Asakura, Tetsuo

2005-06-01

46

The effect of gamma ray on the thermo-mechanical property of silk fibroin  

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Full Text Available This paper studies the effect of gamma ray on the thermo-mechanical properties of silk fibroin experimentally. The experiment reveals that the gamma ray can accelerate biodegradation of silk fibroin by decreasing its thermo-mechanical property.

Jin Huanyu

2013-01-01

47

Differential usage of iso-accepting tRNASer species in silk glands of Bombyx mori.  

Science.gov (United States)

The rapid development of the silk glands of Bombyx mori during the last larval instar shows two phases. During the first 4 days, in both the middle and posterior parts of the silk glands, the ribosomal machinery is assembled and the synthesis of housekeeping proteins starts. During the second phase (the last 4 days), the middle part of the gland synthesis approximately 45 mg of the silk protein sericin (31% serine) and the posterior part of the gland synthesizes approximately 130 mg of the silk protein fibroin (46% glycine, 29% alanine and 12% serine). Silk fibroin and sericin are detectable by the second day and represent 80 and 50% respectively of the total proteins produced at day 8 (refs 1--4). It is known that the tRNA population of the posterior part of the gland is quantitatively adapted to fibroin codon frequency during this period but little is known about the situation in the middle part except for the observation that it contains more tRNASer than does the posterior part. We show here that the two parts contain, and presumably use, different iso-accepting species of tRNASer, the middle part using tRNASer1, which recognizes AGU and AGC codons, and the posterior part using tRNASer2 which recognizes UCA. We also suggest that this differential adaptation of the tRNASer species is under transcriptional control as the two species are accumulated at different rates, but degraded at the same rate. PMID:6782490

Hentzen, D; Chevallier, A; Garel, J P

1981-03-19

48

Cell proliferation and migration in silk fibroin 3D scaffolds.  

Science.gov (United States)

Pore architecture in 3D polymeric scaffolds is known to play a critical role in tissue engineering as it provides the vital framework for the seeded cells to organize into a functioning tissue. In this report, we investigated the effects of different freezing temperature regimes on silk fibroin protein 3D scaffold pore microstructure. The fabricated scaffolds using freeze-dry technique were used as a 3D model to monitor cell proliferation and migration. Pores of 200-250microm diameter were formed by slow cooling at temperatures of -20 and -80 degrees C but were found to be limited in porosity and pore interconnectivity as observed through scanning electron microscopic images. In contrast, highly interconnected pores with 96% porosity were observed when silk solutions were rapidly frozen at -196 degrees C. A detailed study was conducted to assess the affect of pore size, porosity and interconnectivity on human dermal fibroblast cell proliferation and migration on these 3D scaffolds using confocal microscopy. The cells were observed to migrate within the scaffold interconnectivities and were found to reach scaffold periphery within 28 days of culture. Confocal images further confirmed normal cell attachment and alignment of actin filaments within the porous scaffold matrix with well-developed nuclei. This study indicates rapid freeze-drying technique as an alternative method to fabricate highly interconnected porous scaffolds for developing functional 3D silk fibroin matrices for potential tissue engineering, biomedical and biotechnological applications. PMID:19249094

Mandal, Biman B; Kundu, Subhas C

2009-05-01

49

Characteristics of phoxim-exposed gene transcription in the silk gland of silkworms.  

Science.gov (United States)

Silkworm (Bombyx mori), a model Lepidoptera insect, is an important economic insect. Its silk gland is the important organ for silk protein synthesis and secretion. Phoxim exposure causes deficient cocooning of silkworm and has become one of the major negative factors for the silk industry. To study the impact of phoxim exposure on silk gland, using gene chip technology, we examined differentially expressed genes in silk gland after silkworms were exposed to phoxim (4.0?g/mL) for 24h. Functional annotation, classification and KEGG signaling pathway analysis were performed. The results showed that out of 3206 genes detected in silk gland after phoxim exposure, 270 were differentially expressed significantly, including 249 up-regulated genes and 21 down-regulated genes. These differentially expressed genes related to apoptosis, detoxification and protein degradation were selected. Using qRT-PCR, the expression levels of 9 genes involved in apoptosis, detoxification and protein degradation were validated. In addition, the expression profiles of three related fibroin synthesis genes (Fib-H, Fib-L and P25) were analyzed. Our results showed that phoxim exposure induced apoptosis of silk gland cells and inhibition of fibroin synthesis. This may be the cause of deficient silkworm cocooning. PMID:24267702

Ma, L; Xie, Y; Gu, Z Y; Wang, B B; Li, F C; Xu, K Z; Shen, W D; Li, B

2013-11-01

50

Protein-protein nanoimprinting of silk fibroin films  

Science.gov (United States)

Control of the interface between biological tissue and high technology materials is paramount for the development of future applications in biomedicine, especially in the case of implantable integrated devices for signal transduction.[1]-[3] Such work requires careful materials design to develop devices that can efficiently perform technological functions while retaining biocompatibility and biological integration. Silk fibroin protein from the Bombyx mori silkworm has come of considerable interest in this context, owing to its attractive mechanical,[4]-[7] biological, [8][9] and optical properties.[10][11] Recent work has shown adaptation of common micro- and nano-fabrication tools to silk films,[12]-[18] as well as silk protein secondary structure patterning techniques,[19] leading to biocompatible and degradable electronic and photonic devices which can simultaneously act as a carrier and stabilizer for protein pharmaceuticals and other bioactive reagents.[20]-[23] In particular, silk based nanoscale photonic devices face the challenge of sub-wavelength resolution fabrication on a soft polymeric substrate.[15][24] Previous work introduced the possibility of direct, rapid nanoimprinting in silk for the fabrication of photonic structures by leveraging the material properties of this protein.[25

Brenckle, Mark A; Tao, Hu; Kim, Sunghwan; Paquette, Mark; Kaplan, David L; Omenetto, Fiorenzo G

2013-01-01

51

Electrophoretic deposition of tetracycline modified silk fibroin coatings for functionalization of titanium surfaces  

Science.gov (United States)

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.

Zhang, Zhen; Qu, Yinying; Li, Xiaoshuang; Zhang, Sheng; Wei, Qingsong; Shi, Yusheng; Chen, Lili

2014-06-01

52

Swelling and dissolution of silk fibroin (Bombyx mori) in N-methyl morpholine N-oxide.  

Science.gov (United States)

Bombyx mori silk fibers were dissolved in N-methyl morpholine N-oxide (MMNO), an organic cyclic amine oxide used for the solvent spinning of regenerated cellulosic fibers. The commercial MMNO monohydrate used in this study as a solvent for silk is a hygroscopic compound crystalline at room temperature, which becomes an active solvent after melting at 76 degrees C. The degree of hydration of MMNO was checked by DSC measurements. The solvation power of MMNO towards silk fibroin drastically decreased at a water content > or = 20-21% w/w. Dissolution of silk required both thermal and mechanical energy. The optimum temperature was 100 degrees C. At lower temperatures dissolution proceeded very slowly. At higher temperatures, rapid depolymerization of silk fibroin occurred. The value of the Flory-Huggins interaction parameter chi for the MMNO-H2O-silk fibroin system was -8.5, suggesting that dissolution is a thermodynamically favored process. The extent of degradation of silk fibroin was assessed by measuring the intrinsic viscosity and determining the amino acid composition of silk after regeneration with an aqueous methanol solution, which was effective in removing the solvent and coagulating silk. Regenerated silk fibroin membranes were characterized by infrared spectroscopy, differential scanning calorimetry and scanning electron microscopy. The prevailing molecular conformation of silk fibroin chains was the beta-sheet structure, as shown by the intense amide I-III bands at 1704, 1627, 1515, 1260, and 1230 cm(-1). The value of the I1260/I1230 intensity ratio (crystallinity index) was 0.68, comparable to that of the fibers. The DSC thermogram was characteristic of a silk fibroin material with unoriented beta-sheet crystalline structure, with an intense decomposition endotherm at 294 degrees C. The SEM examination of fractured surfaces showed the presence of a dense microstructure with a very fine texture formed by densely packed roundish particles of about 100-200 nm diameter. PMID:10342772

Freddi, G; Pessina, G; Tsukada, M

1999-01-01

53

The degradation behavior of silk fibroin derived from different ionic liquid solvents  

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Full Text Available Establishing an appropriate degradation rate is critical for tissue engineering scaffolds. In this study, the degradation rate of silk fibroin three-dimensional scaffolds was regulated by changing the molecular weight (MW of the silk fibroin. The solubility of silk fibroin depends primarily on the ionic ability of the slovent to dissolve silk fibroin, therefore, we regulated the MW of the silk fibroin using LiBr, Ca(NO32 and CaCl2 to dissolve the silk fibers. SDS-PAGE analysis showed that the MW of the CaCl2-derived silk fibroin was lower than the MW produced using LiBr and Ca(NO32. In vitro and in vivo degradation results showed that the scaffolds prepared by low-MW silk fibroin were more rapidly degraded. Furthermore, FTIR and amino acid analysis suggested that the amorphous regions were preferentially degraded by Collagenase IA, while the SDS-PAGE and amino acid analysis indicated that the scaffolds were degraded into polypeptides (mainly at 10-30 kDa and amino acids. Because the CaCl2-derived scaffolds contained abundant low MW polypeptides, inter-intramolecular entanglement and traversing of molecular chains in the crystallites reduced, which resulted in rapid degradation. The in vivo degradation results suggested that the degradation rate of the CaCl2-derived scaffolds was better matched to dermis regeneration, indicating that the degradation rate of silk fibroin can be effectively regulated by changing the MW to achieve a suitable dermal tissue regeneration rate.

Renchuan You

2013-06-01

54

Structures of Bombyx mori and Samia cynthia ricini silk fibroins studied with solid-state NMR.  

Science.gov (United States)

There are many kinds of silks spun by silkworms and spiders, which are suitable to study the structure-property relationship for molecular design of fibers with high strength and high elasticity. In this review, we mainly focus on the structural determination of two well-known silk fibroin proteins that are from the domesticated silkworm, Bombyx mori, and the wild silkworm, Samia cynthia ricini, respectively. The structures of B. mori silk fibroin before and after spinning were determined by using an appropriate model peptide, (AG)(15), with several solid-state NMR methods; (13)C two-dimensional spin-diffusion solid-state NMR and rotational echo double resonance (REDOR) NMR techniques along with the quantitative use of the conformation-dependent (13)C CP/MAS chemical shifts. The structure of S. c. ricini silk fibroin before spinning was also determined by using a model peptide, GGAGGGYGGDGG(A)(12)GGAGDGYGAG, which is a typical repeated sequence of the silk fibroin, with the solid-state NMR methods. The transition from the structure of B. mori silk fibroin before spinning to the structure after spinning was studied with molecular dynamics calculation by taking into account several external forces applied to the silk fibroin in the silkworm. PMID:15132647

Yao, Juming; Nakazawa, Yasumoto; Asakura, Tetsuo

2004-01-01

55

Controlling microstructure of three-dimensional scaffolds from regenerated silk fibroin by adjusting pH.  

Science.gov (United States)

For tissue engineering, it is very important to design and control the pore architecture of three-dimensional (3D) polymeric scaffolds, which plays an important role in directing tissue formation and function. In this study, 3D porous silk fibroin scaffolds produced using a freeze drying technique were prepared at pHs ranging from 5 to 9. The effects of pH on the pore microstructure of the silk fibroin scaffold were examined by rheometry, FESEM and FTIR. Different pore structures were formed according to the pH of silk fibroin because silk fibroin exhibits water-like behavior under basic conditions and gel-like behavior under acidic conditions. PMID:22524061

Cho, Se Youn; Heo, Semi; Jin, Hyoung-Joon

2012-01-01

56

Crystal growth of calcium carbonate in silk fibroin/sodium alginate hydrogel  

Science.gov (United States)

As known, silk fibroin-like protein plays a pivotal role during the formation of calcium carbonate (CaCO3) crystals in the nacre sheets. Here, we have prepared silk fibroin/sodium alginate nanofiber hydrogels to serve as templates for calcium carbonate mineralization. In this experiment, we report an interesting finding of calcium carbonate crystal growth in the silk fibroin/sodium alginate nanofiber hydrogels by the vapor diffusion method. The experimental results indicate calcium carbonate crystals obtained from nanofiber hydrogels with different proportions of silk fibroin/sodium alginate are mixture of calcite and vaterite with unusual morphologies. Time-dependent growth study was carried out to investigate the crystallization process. It is believed that nanofiber hydrogels play an important role in the process of crystallization. This study would help in understanding the function of organic polymers in natural mineralization, and provide a novel pathway in the design and synthesis of new materials related unique morphology and structure.

Ming, Jinfa; Zuo, Baoqi

2014-01-01

57

Silk Fibroin Conduits: A Cellular and Functional Assessment of Peripheral Nerve Repair  

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Novel silk fibroin conduits were designed with appropriate porosity for peripheral nerve repair. The aim of this work was to utilize these conduits to examine cell inflammatory responses and functional recovery in a sciatic nerve defect model.

Ghaznavi, Amir Mahan; Kokai, Lauren E.; Lovett, Michael L.; Kaplan, David L.; Marra, Kacey G.

2011-01-01

58

Effect of Silk Fibroin Content on the Bionic Mineralization and In Vitro Cellular Compatibility of Silk Fibroin/Hydroxyapaptite Nanocomposites  

Directory of Open Access Journals (Sweden)

Full Text Available Silk fibroin and hydroxyapatite nanocomposites (SF/HA with various SF content were prepared. Effect of Silk fibroin content on the bionic mineralization and cellular compatibility in vitro of HA nanocrystals (n-HA was investigated. The results show that SF content has an obvious effect on the nucleation and growth of n-HA. However, though SF content does not show obvious difference on the nucleation and growth of n-HA, it has obvious effect on the n-HA aggregation. When SF content is less than 20wt%, n-HA orderly disperses in SF matrix. While the SF content is more than 20wt%, the n-HA aggregation becomes disordered. The in vitro cellular compatibility experiments demonstrate that the SF/HA composites exhibit better cell affinity than pure n-HA. However, SF content has no obvious effect on the cell affinity of n-SF/HA 20wt% SF/HA and 30wt% SF/HA show better osteoblast proliferation.

ZHU Yun-Rong, CHEN Yu-Yun, XU Guo-Hua, YE Xiao-Jian, ZHONG Jian, HE Dan-Nong

2012-08-01

59

Study On Degradation Of Silk Fibroin By Irradiation Treatment For Cosmetic And Pharmaceutical Applications  

International Nuclear Information System (INIS)

As a kind of protein, silk fibroin is created with silkworm Bombyx mori in products of silk. The fibroin was irradiated using Co-60 gamma source and its degradability and solubility were investigated with various radiation doses to apply in pharmacy and cosmetic. Addition to the morphological changes of irradiated fibroin fibers shows that its mechanical properties were much influenced by the irradiation. Tensile strength and elongation at break of the silk fibroin significantly decreased with increasing of radiation dose up to 1000 kGy. The tensile strength and elongation at break of the irradiated fibroin at 1000 kGy reduced to 71% and 94% respectively in compared with non-irradiated one. The solubility of silk fibroin in both calcium chloride (CaCl2/C2H5OH/H2O=1:2:8) in mole ratio and distilled water were improved by the irradiation. UV spectrometry revealed the structure of silk fibroin was also changed by irradiation. (author)

2013-08-01

60

Development of artificial dermis using 3D electrospun silk fibroin nanofiber matrix.  

Science.gov (United States)

Several auto- or alloplastic dermal substitutes have been used for full thickness skin defects. However, there is no ideal dermal substitute that is biocompatible and offers adequate mechanical properties and sufficient thickness and volume. Silk fibroin can be considered a possible alternative material for the construction of dermal substitutes. Nanofibers fabricated by electrospinning silk fibroin were considered an excellent candidate due to their desirable biocompatibility. However, it is generally difficult for cells to infiltrate the electrospun silk fibroin due to its small pore size. To overcome this challenge, a key parameter is the control of the pore size of silk fibroin electrospun such that cells can easily infiltrate into the nanofibers and proliferate internally. In this work, a novel approach is introduced by dropping two different size NaCl crystals above the rotating collector, which become incorporated into the nanofibers. The structures and properties of the, resulting electrospun silk fibroin were characterized using scanning electron microscopy (SEM), Fourier transform infrared spectroscopy (FTIR), and its wound healing effect for a full thickness skin defect was compared with Matriderm. Although the dermal regenerative effect of the 3D-SF (3D electrospun silk fibroin nanofiber matrix) was similar to that of Matriderm, the 3D-SF was almost completely degraded and did not induce wound contracture like Matriderm. We suggest that 3D-SF can be applied to the treatment of full thickness skin defects. PMID:24804550

Lee, Ok Joo; Ju, Hyung Woo; Kim, Ji Heui; Lee, Jung Min; Ki, Chang Seok; Kim, Jung-Ho; Moon, Bo Mi; Park, Hyun Jung; Sheikh, Faheem A; Park, Chan Hum

2014-07-01

 
 
 
 
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Artificial crawler model for texture analysis on silk fibroin scaffolds  

Science.gov (United States)

Texture plays an important role in computer vision tasks. Several methods of texture analysis are available. However, these methods are not capable of extracting rich detail in images. This paper presents a novel approach to image texture classification based on the artificial crawler model. Here, we propose a new rule of movement that moves artificial crawler agents not only toward higher intensities but also toward lower ones. This strategy is able of capturing more detail because the agents explore the peaks as well as the valleys. Thus, compared with the state-of-the-art method, this approach shows an increased discriminatory power. Experiments on the most well known benchmark demonstrate the superior performance of our approach. We also tested our approach on silk fibroin scaffold analysis, and results indicate that our method is consistent and can be applied in real-world situations.

Brandoli Machado, Bruno; Nunes Gonçalves, Wesley; Martinez Bruno, Odemir

2014-01-01

62

Intermolecular interactions between natural polysaccharides and silk fibroin protein.  

Science.gov (United States)

Fabricating novel functional and structural materials from natural renewable and degradable materials has attracted much attention. Natural polysaccharides and proteins are the right natural candidates due to their unique structures and properties. The polysaccharide-protein composites or blends were widely investigated, however, there are few systematical studies on the interactions between natural polysaccharides and silk fibroin protein at the molecular level. Among various interactions, hydrogen bonding, electrostatic interactions and covalent bonding play important roles in the structure and properties of the corresponding materials. Therefore, the focus is placed on the three interactions types in this review. A future challenge is to create polysaccharide and protein composites or blends with tailored structure and properties for the wide applications. PMID:23499097

Shang, Songmin; Zhu, Lei; Fan, Jintu

2013-04-01

63

Silk fibroin and sodium alginate blend: Miscibility and physical characteristics.  

Science.gov (United States)

Films of silk fibroin (SF) and sodium alginate (SA) blends were prepared by solution casting technique. The miscibility of SF and SA in those blends was evaluated and scanning electron microscopy (SEM) revealed that SF/SA 25/75wt.% blends underwent microscopic phase separation, resulting in globular structures composed mainly of SF. X-ray diffraction indicated the amorphous nature of these blends, even after a treatment with ethanol that turned them insoluble in water. Thermal analyses of blends showed the peaks of degradation of pristine SF and SA shifted to intermediate temperatures. Water vapor permeability, swelling capacity and tensile strength of SF films could be enhanced by blending with SA. Cell viability remained between 90 and 100%, as indicated by in vitro cytotoxicity test. The SF/SA blend with self-assembled SF globules can be used to modulate structural and mechanical properties of the final material and may be used in designing high performance wound dressing. PMID:24857469

de Moraes, Mariana Agostini; Silva, Mariana Ferreira; Weska, Raquel Farias; Beppu, Marisa Masumi

2014-07-01

64

Antimicrobial Silk Fibroin Hydrogel Instantaneously Induced by Cationic Surfactant  

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Full Text Available Current surgical dressings and woundplast have drawbacks such as poor permeability, airtightness and biocompatibility, thus a new material is obligatory to replace the traditional ones. Due to outstanding biocompatibility of Silk Fibroin (SF and excellent antibacterial properties of Dodecyl Trimethyl Ammonium Bromide (DTAB, a cationic surfactant, the SF/DTAB hydrogel formed instantaneously is recommended. By means of SEM, FTIR and XRD, SF/DTAB hydrogel displayed clustered nanofilaments structure and similar secondary structure and morphology with SF solution mainly consisted of random coils in 1 h, then it tended to be the same as pure SF hydrogel which was composed of porous network structure and primarily contained ?-sheet structure. Excellent elasticity and antibacterial effect were verified by rheological and antibacterial measurements.

Shan Sun

2013-01-01

65

Self-assembly model, hepatocytes attachment and inflammatory response for silk fibroin/chitosan scaffolds  

International Nuclear Information System (INIS)

Silk fibroin is an attractive natural fibrous protein for biomedical application due to its good biocompatibility and high tensile strength. Silk fibroin is apt to form a sheet-like structure during the freeze-drying process, which is not suitable for the scaffold of tissue engineering. In our former study, the adding of chitosan promoted the self-assembly of silk fibroin/chitosan (SFCS) into a three-dimensional (3D) homogeneous porous structure. In this study, a model of the self-assembly is proposed; furthermore, hepatocytes attachment and inflammatory response for the SFCS scaffold were examined. The rigid chain of chitosan may be used as a template for ?-sheet formation of silk fibroin, and this may break the sheet structure of the silk fibroin scaffold and promote the formation of a 3D porous structure of the SFCS scaffold. Compared with the polylactic glycolic acid scaffold, the SFCS scaffold further facilitates the attachment of hepatocytes. To investigate the inflammatory response, SFCS scaffolds were implanted into the greater omentum of rats. From the results of implantation, we could demonstrate in vivo that the implantation of SFCS scaffolds resulted in only slight inflammation. Keeping the good histocompatibility and combining the advantages of both fibroin and chitosan, the SFCS scaffold could be a prominent candidate for soft tissue engineering, for example, in the liver.

2009-08-01

66

Self-assembly model, hepatocytes attachment and inflammatory response for silk fibroin/chitosan scaffolds  

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Silk fibroin is an attractive natural fibrous protein for biomedical application due to its good biocompatibility and high tensile strength. Silk fibroin is apt to form a sheet-like structure during the freeze-drying process, which is not suitable for the scaffold of tissue engineering. In our former study, the adding of chitosan promoted the self-assembly of silk fibroin/chitosan (SFCS) into a three-dimensional (3D) homogeneous porous structure. In this study, a model of the self-assembly is proposed; furthermore, hepatocytes attachment and inflammatory response for the SFCS scaffold were examined. The rigid chain of chitosan may be used as a template for beta-sheet formation of silk fibroin, and this may break the sheet structure of the silk fibroin scaffold and promote the formation of a 3D porous structure of the SFCS scaffold. Compared with the polylactic glycolic acid scaffold, the SFCS scaffold further facilitates the attachment of hepatocytes. To investigate the inflammatory response, SFCS scaffolds were implanted into the greater omentum of rats. From the results of implantation, we could demonstrate in vivo that the implantation of SFCS scaffolds resulted in only slight inflammation. Keeping the good histocompatibility and combining the advantages of both fibroin and chitosan, the SFCS scaffold could be a prominent candidate for soft tissue engineering, for example, in the liver.

She Zhending; Feng Qingling [State Key Laboratory of New Ceramics and Fine Processing, Department of Materials Science and Engineering, Tsinghua University, Beijing 100084 (China); Liu Weiqiang, E-mail: biomater@mail.tsinghua.edu.c [Center for Advanced Materials and Biotechnology, Research Institute of Tsinghua University in Shenzhen, Shenzhen 518057 (China)

2009-08-15

67

Sodium dodecyl sulfate-induced rapid gelation of silk fibroin.  

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The in situ formation of injectable silk fibroin (SF) hydrogels have potential advantages over various other biomaterials due to the minimal invasiveness during application. Biomaterials need to gel rapidly under physiological conditions after injection. In the current paper, a novel way to accelerate SF gelation using an anionic surfactant, sodium dodecyl sulfate (SDS), as a gelling agent is reported. The mechanism of SDS-induced rapid gelation was determined. At low surfactant concentrations, hydrophobic interactions among the SF chains played a dominant role in the association, leading to decreased gelation time. At higher concentrations of surfactant, electrostatic repulsive forces among micellar aggregates gradually became dominant and gelation was hindered. Gel formation involves the connection of clusters formed by the accumulation of nanoparticles. This process is accompanied by the rapid formation of ?-sheet structures due to hydrophobic and electrostatic interactions. It is expected that the silk hydrogel with short gelation time will be used as an injectable hydrogel in drug delivery or cartilage tissue engineering. PMID:22406911

Wu, Xilong; Hou, Jing; Li, Mingzhong; Wang, Jiangnan; Kaplan, David L; Lu, Shenzhou

2012-07-01

68

Optimization of the silk scaffold sericin removal process for retention of silk fibroin protein structure and mechanical properties.  

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In the process of removing sericin (degumming) from a raw silk scaffold, the fibroin structural integrity is often challenged, leading to mechanical depreciation. This study aims to identify the factors and conditions contributing to fibroin degradation during alkaline degumming and to perform an optimization study of the parameters involved to achieve preservation of fibroin structure and properties. The methodology involves degumming knitted silk scaffolds for various durations (5-90 min) and temperatures (60-100 degrees C). Mechanical agitation and use of the refreshed solution during degumming are included to investigate how these factors contribute to degumming efficiency and fibroin preservation. Characterizations of silk fibroin morphology, mechanical properties and protein components are determined by scanning electron microscopy (SEM), single fiber tensile tests and gel electrophoresis (SDS-PAGE), respectively. Sericin removal is ascertained via SEM imaging and a protein fractionation method involving SDS-PAGE. The results show that fibroin fibrillation, leading to reduced mechanical integrity, is mainly caused by prolonged degumming duration. Through a series of optimization, knitted scaffolds are observed to be optimally degummed and experience negligible mechanical and structural degradation when subjected to alkaline degumming with mechanical agitation for 30 min at 100 degrees C. PMID:20460689

Teh, Thomas K H; Toh, Siew-Lok; Goh, James C H

2010-05-11

69

Optimization of the silk scaffold sericin removal process for retention of silk fibroin protein structure and mechanical properties  

International Nuclear Information System (INIS)

In the process of removing sericin (degumming) from a raw silk scaffold, the fibroin structural integrity is often challenged, leading to mechanical depreciation. This study aims to identify the factors and conditions contributing to fibroin degradation during alkaline degumming and to perform an optimization study of the parameters involved to achieve preservation of fibroin structure and properties. The methodology involves degumming knitted silk scaffolds for various durations (5-90 min) and temperatures (60-100 0C). Mechanical agitation and use of the refreshed solution during degumming are included to investigate how these factors contribute to degumming efficiency and fibroin preservation. Characterizations of silk fibroin morphology, mechanical properties and protein components are determined by scanning electron microscopy (SEM), single fiber tensile tests and gel electrophoresis (SDS-PAGE), respectively. Sericin removal is ascertained via SEM imaging and a protein fractionation method involving SDS-PAGE. The results show that fibroin fibrillation, leading to reduced mechanical integrity, is mainly caused by prolonged degumming duration. Through a series of optimization, knitted scaffolds are observed to be optimally degummed and experience negligible mechanical and structural degradation when subjected to alkaline degumming with mechanical agitation for 30 min at 100 0C.

2010-06-01

70

Preparation of Eri silk fibroin and gelatin blend film loaded chlorhexidine using as model for hydrophilic drug release  

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The objective of this research was to prepare Eri silk fibroin solution for preparing silk film loaded chlorhexidine drug as model for hydrophilic drug release. The Eri silk cocoons were boiled in 0.5%NaCO3 solution at 90?, and then left in air dried at room temperature. The fibroin was dissolved in 9M (Ca(NO3)2) with ethanol (2 by mole) and heated at 70?. The silk fibroin (SF) solution was then dialyzed to exclude salt ...

Yaowalak Srisuwan; Nualchai Kotseang; Komsan Namtaku; Wilaiwan Simchuer; Chirapha Butiman; Prasong Srihanam

2012-01-01

71

Dissolution of Bombyx mori silk fibroin in the calcium nitrate tetrahydrate-methanol system and aspects of wet spinning of fibroin solution.  

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There are still several problems associated with the spinning of dialyzed silk fibroin solutions. In this work some of these problems have been examined. The calcium nitrate tetrahydrate-methanol system was used to dissolve the silk fibroin. A compositional phase diagram was constructed at various concentrations of the solvent system. Regenerated fibroin powders from undialyzed fibroin solution in several coagulants showed different conformations. Regenerated powders from several coagulants except methanol and ethanol were resoluble in water. Atomic absorption analysis revealed that the calcium cations strongly interact with fibroin molecules in dialyzed fibroin solution, which may interfere with the regeneration of a strong fiber. Kinetic studies to determine the diffusion coefficient of methanol into dialyzed and concentrated fibroin solution were reported. The properties of both original and regenerated fibroin such as solubility in water and thermal behaviors using DSC were compared. Regenerated fibroin fiber was spun by the wet spinning method. An X-ray diffractogram showed that the regeneration process decreased the crystallinity of regenerated fibroin fiber. SEM images of the surface and cross section of the regenerated fibroin fibers were discussed. PMID:12741761

Ha, Sung-Won; Park, Young H; Hudson, Samuel M

2003-01-01

72

Determination of molecular weight of silk fibroin by non-gel sieving capillary electrophoresis.  

Science.gov (United States)

A simple non-gel sieving capillary electrophoresis (NGSCE) method was established to determine the MW of silk fibroin using CE. The background electrolyte with a pH of 8.8 was based on three components: polyethylene glycol, tris(hydroxymethyl)aminomethane, and sodium dodecyl sulfate (SDS). NGSCE showed a good linear relationship with satisfactory reproducibility between the migration time and the MW of standard proteins. It was found that the regenerated silk fibroin had an MW around 83 kDa with a wide MW distribution (MWD). This absolute value is lower than the result obtained from SDS-polyacrylamide gel electrophoresis due to the different principles of the methods, but their similar MWD shapes indicated that NGSCE could be a feasible, highly sensitive, rapid method for determination of the MW of silk fibroin. PMID:20922945

Wei, Wei; Zhang, Yaopeng; Shao, Huili; Hu, Xuechao

2010-01-01

73

Effect of freezing methods on the properties of lyophilized porous silk fibroin membranes  

Scientific Electronic Library Online (English)

Full Text Available SciELO Brazil | Language: English Abstract in english Silk fibroin is a fibrous protein that has been extensively studied for application in the biomedical field, and has been used as a scaffold for bone tissue engineering. Biomaterials made of proteins are prone to physical and chemical degradation during storage; lyophilization, a drying method that [...] consists of freezing and drying steps, is known to promote minimal changes in structure and biological activity of biomaterials. This study evaluates the effect of freezing methods on the properties of lyophilized porous silk fibroin membranes. The membranes were obtained from silk fibroin solution, frozen in liquid nitrogen or ultrafreezer, lyophilized, and then characterized by XRD, FTIR, TGA, DSC and SEM. Although the membranes presented similar physical, chemical and microstructural characteristics, quench freezing with liquid nitrogen, followed by lyophilization, promoted collapse of the membranes, while slow cooling performed by ultrafreezer preserved membrane integrity.

Weska, Raquel Farias; Vieira Jr., Wellington Carlos; Nogueira, Grínia Michelle; Beppu, Marisa Masumi.

74

Effect of freezing methods on the properties of lyophilized porous silk fibroin membranes  

Directory of Open Access Journals (Sweden)

Full Text Available Silk fibroin is a fibrous protein that has been extensively studied for application in the biomedical field, and has been used as a scaffold for bone tissue engineering. Biomaterials made of proteins are prone to physical and chemical degradation during storage; lyophilization, a drying method that consists of freezing and drying steps, is known to promote minimal changes in structure and biological activity of biomaterials. This study evaluates the effect of freezing methods on the properties of lyophilized porous silk fibroin membranes. The membranes were obtained from silk fibroin solution, frozen in liquid nitrogen or ultrafreezer, lyophilized, and then characterized by XRD, FTIR, TGA, DSC and SEM. Although the membranes presented similar physical, chemical and microstructural characteristics, quench freezing with liquid nitrogen, followed by lyophilization, promoted collapse of the membranes, while slow cooling performed by ultrafreezer preserved membrane integrity.

Raquel Farias Weska

2009-06-01

75

Engineered silk fibroin protein 3D matrices for in vitro tumor model.  

Science.gov (United States)

3D in vitro model systems that are able to mimic the in vivo microenvironment are now highly sought after in cancer research. Antheraea mylitta silk fibroin protein matrices were investigated as potential biomaterial for in vitro tumor modeling. We compared the characteristics of MDA-MB-231 cells on A. mylitta, Bombyx mori silk matrices, Matrigel, and tissue culture plates. The attachment and morphology of the MDA-MB-231 cell line on A. mylitta silk matrices was found to be better than on B. mori matrices and comparable to Matrigel and tissue culture plates. The cells grown in all 3D cultures showed more MMP-9 activity, indicating a more invasive potential. In comparison to B. mori fibroin, A. mylitta fibroin not only provided better cell adhesion, but also improved cell viability and proliferation. Yield coefficient of glucose consumed to lactate produced by cells on 3D A. mylitta fibroin was found to be similar to that of cancer cells in vivo. LNCaP prostate cancer cells were also cultured on 3D A. mylitta fibroin and they grew as clumps in long term culture. The results indicate that A. mylitta fibroin scaffold can provide an easily manipulated microenvironment system to investigate individual factors such as growth factors and signaling peptides, as well as evaluation of anticancer drugs. PMID:21167597

Talukdar, Sarmistha; Mandal, Mahitosh; Hutmacher, Dietmar W; Russell, Pamela J; Soekmadji, Carolina; Kundu, Subhas C

2011-03-01

76

Silk fibroin porous scaffolds for nucleus pulposus tissue engineering.  

Science.gov (United States)

Intervertebral discs (IVDs) are structurally complex tissue that hold the vertebrae together and provide mobility to spine. The nucleus pulposus (NP) degeneration often results in degenerative IVD disease that is one of the most common causes of back and neck pain. Tissue engineered nucleus pulposus offers an alternative approach to regain the function of the degenerative IVD. The aim of this study is to determine the feasibility of porous silk fibroin (SF) scaffolds fabricated by paraffin-sphere-leaching methods with freeze-drying in the application of nucleus pulposus regeneration. The prepared scaffold possessed high porosity of 92.38±5.12% and pore size of 165.00±8.25?m as well as high pore interconnectivity and appropriate mechanical properties. Rabbit NP cells were seeded and cultured on the SF scaffolds. Scanning electron microscopy, histology, biochemical assays and mechanical tests revealed that the porous scaffolds could provide an appropriate microstructure and environment to support adhesion, proliferation and infiltration of NP cells in vitro as well as the generation of extracellular matrix. The NP cell-scaffold construction could be preliminarily formed after subcutaneously implanted in a nude mice model. In conclusion, The SF porous scaffold offers a potential candidate for tissue engineered NP tissue. PMID:24582244

Zeng, Chao; Yang, Qiang; Zhu, Meifeng; Du, Lilong; Zhang, Jiamin; Ma, Xinlong; Xu, Baoshan; Wang, Lianyong

2014-04-01

77

The Cytocompatibility of Genipin-Crosslinked Silk Fibroin Films  

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Full Text Available There is an increasing demand for crosslinking methods of silk fibroin (SF scaffolds in biomedical applications that could maintain the biocompatibility, bioactivity as well as improve the water resistance and mechanical properties of SF materials. In this study, SF was crosslinked effectively with genipin which is a naturally occurring iridoid glucoside and the crosslinking mechanism was investigated through FTIR and amino acid analysis. The results showed that genipin could react with the -NH2 groups on the side chains of SF macromolecules and to form inter- and intra-molecular covalent bonds, and improved the stability of SF materials significantly. In vitro, the performances of genipin-crosslinked SF films were assessed by seeding L929 cells and compared with ethanol-processed SF films, glutaraldehyde and polyethylene glycol diglycidyl ether crosslinked ones. The genipin-crosslinked SF films showed a similar affinity to cells as ethanol-processed ones, and a higher bioactivity in promoting cell growth and proliferation, inhibition of cell apoptosis, and maintenance of normal cell cycle compared with glutaraldehyde and polyethylene glycol diglycidyl ether crosslinked SF films. These features, combined with the decrease of brittleness of SF films crosslinked with chemical methods, substantiated genipin as an effective and biocompatible agent for the manufacturing of bioactive SF materials which used as tissue engineering scaffolds and drug delivery carriers.

Lingshuang Wang

2013-06-01

78

Investigation of uranium recovery from dilute aqueous solutions using silk fibroin.  

Science.gov (United States)

The uranium uptake ability of silk fibroin was investigated. High ability to uptake uranium from nonsaline water containing 2.500 mg of uranium was observed with the silk fibroin tested. The uranium uptake was very rapid and was dependent on pH, uranium concentration, temperature, and retention time. Almost all uranium taken up is easily eluted with 1 mol/L CH3COONH4. This biomatrix, therefore, appears to have potential for use in a commercial process for uranium recovery from uranium-containing waste water. PMID:7710896

Aslani, M A; Eral, M

1994-01-01

79

Synthesis and characterization of dense membranes of silk fibroin with glycerin  

International Nuclear Information System (INIS)

The addition of plasticizers seeks improvements in mechanical properties of dense membranes of silk fibroin with possible interactions by hydrogen bonds. The aim of the present study was to produce and characterize dense membranes of silk fibroin containing glycerin in two different concentrations. The characterization of the membranes was performed from scanning electron microscopy (SEM), mechanical traction tests, infrared spectroscopy (FTIR-ATR) and X-ray diffraction (XRD). The results indicated that the addition of glycerin allowed obtaining homogeneous and more crystalline membranes and improved their properties of elongation. (author)

2009-10-13

80

In vitro and in vivo release of basic fibroblast growth factor using a silk fibroin scaffold as delivery carrier.  

Science.gov (United States)

Two different solvents were used to prepare two types of silk fibroin scaffolds via the salt-leaching technique, i.e., hexafluoroisopropanol (HFIP) and water. The in vitro release study suggests that the opposite charge between the silk fibroin and basic fibroblast growth factor (bFGF) at physiological pH rendered them to form a complex, and the difference in the solvents used to produce the silk fibroin scaffold did not affect the affinity of silk fibroin to bFGF. However, a higher degradation rate of the aqueous-derived silk fibroin scaffolds provided faster in vitro release kinetics of the bFGF, as compared to the HFIP-derived scaffolds. From the in vivo studies, the use of silk fibroin scaffolds as the carrier matrix enabled the control of the in vivo release of bFGF in a sustained fashion over two weeks, while the majority of the bFGF disappeared within one day after the injection of the bFGF in soluble form. In addition, the in vivo release of bFGF from the silk fibroin scaffolds was not affected by the mode of processing due to their similar degradation behavior in vivo. PMID:20534193

Wongpanit, Panya; Ueda, Hiroki; Tabata, Yasuhiko; Rujiravanit, Ratana

2010-01-01

 
 
 
 
81

In Vitro Evaluation of Electrospun Silk fibroin Scaffolds for Vascular Cell Growth  

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Human aortic endothelial (HAEC) and human coronary artery smooth muscle cell (HCASMC) responses on electrospun silk fibroin scaffolds were studied to evaluate potential for vascular tissue engineering. Cell proliferation studies supported the utility of this biomaterial matrix by both HAECs and HCASMCs. Alignment and elongation of HCASMCs on random nonwoven nanofibrous silk scaffolds was observed within 5 days after seeding based on SEM and confocal microscopy. Short cord-like structures form...

Zhang, Xiaohui; Baughman, Cassandra B.; Kaplan, David L.

2008-01-01

82

Novel genipin cross-linked chitosan-silk fibroin sponges for cartilage engineering strategies  

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The positive interaction of materials with tissues is an important step in regenerative medicine strategies. Hydrogels that are obtained from polysaccharides and proteins are expected to mimic the natural cartilage environment and thus provide an optimum milleu for tissue growth and regeneration. In this work, novel hydrogels composed of blends of chitosan and Bombyx mori silk fibroin were cross-linked with genipin (G) and were freeze dried to obtain chitosan/silk (CSG) sponges. C...

Silva, S. S.; Motta, A.; Rodrigues, Ma?rcia T.; Pinheiro, A. F. M.; Gomes, Manuela E.; Mano, J. F.; Reis, R. L.; Migliaresi, C.

2008-01-01

83

Enzymatic degradation behavior and cytocompatibility of silk fibroin-starch-chitosan conjugate membranes  

Energy Technology Data Exchange (ETDEWEB)

The objective of this study was to investigate the influence of silk fibroin and oxidized starch conjugation on the enzymatic degradation behavior and the cytocompatability of chitosan based biomaterials. The tensile stress of conjugate membranes, which was at 50 Megapascal (MPa) for the lowest fibroin and starch composition (10 weight percent (wt.%)), was decreased significantly with the increased content of fibroin and starch. The weight loss of conjugates in {alpha}-amylase was more notable when the starch concentration was the highest at 30 wt.%. The conjugates were resistant to the degradation by protease and lysozyme except for the conjugates with the lowest starch concentration. After 10 days of cell culture, the proliferation of osteoblast-like cells (SaOS-2) was stimulated significantly by higher fibroin compositions and the DNA synthesis on the conjugate with the highest fibroin (30 wt.%) was about two times more compared to the native chitosan. The light microscopy and the image analysis results showed that the cell area and the lengths were decreased significantly with higher fibroin/chitosan ratio. The study proved that the conjugation of fibroin and starch with the chitosan based biomaterials by the use of non-toxic reductive alkylation crosslinking significantly improved the cytocompatibility and modulated the biodegradation, respectively. - Highlights: Black-Right-Pointing-Pointer Silk fibroin, starch and chitosan conjugates were prepared by reductive alkylation. Black-Right-Pointing-Pointer The enzymatic biodegradation and the cytocompatibility of conjugates were tested. Black-Right-Pointing-Pointer The conjugate with 30% starch composition was degraded by {alpha}-amylase significantly. Black-Right-Pointing-Pointer Higher starch composition in conjugates prevented protease and lysozyme degradation. Black-Right-Pointing-Pointer Fibroin incorporation effectively increased the cell proliferation of conjugates.

Baran, Erkan T., E-mail: erkantur@metu.edu.tr; Tuzlakoglu, Kadriye, E-mail: kadriye@dep.uminho.pt; Mano, Joao F., E-mail: jmano@dep.uminho.pt; Reis, Rui L., E-mail: rgreis@dep.uminho.pt

2012-08-01

84

Enhanced cell adhesion on silk fibroin via lectin surface modification.  

Science.gov (United States)

Various tissue engineering (TE) approaches are based on silk fibroin (SF) as scaffold material because of its superior mechanical and biological properties compared to other materials. The translation of one-step TE approaches to clinical application has generally failed so far due to the requirement of a prolonged cell seeding step before implantation. Here, we propose that the plant lectin WGA (wheat germ agglutinin), covalently bound to SF, will mediate cell adhesion in a time frame acceptable to be part of a one-step surgical intervention. After the establishment of a modification protocol utilizing carbodiimide chemistry, we examined the attachment of cells, with a special focus on adipose-derived stromal cells (ASC), on WGA-SF compared to pure native SF. After a limited time frame of 20min the attachment of ASCs to WGA-SF showed an increase of about 17-fold, as compared to pure native SF. The lectin-mediated cell adhesion further showed an enhanced resistance to trypsin (as a protease model) and to applied fluid shear stress (mechanical stability). Moreover, we could demonstrate that the adhesion of ASCs on the WGA-SF does not negatively influence proliferation or differentiation potential into the osteogenic lineage. To test for in vitro immune response, the proliferation of peripheral blood mononuclear cells in contact with the WGA-SF was determined, showing no alterations compared to plain SF. All these findings suggest that the WGA modification of SF offers important benefits for translation of SF scaffolds into clinical applications. PMID:24530561

Teuschl, Andreas H; Neutsch, Lukas; Monforte, Xavier; Rünzler, Dominik; van Griensven, Martijn; Gabor, Franz; Redl, Heinz

2014-06-01

85

Synthesis of silk fibroin micro- and submicron spheres using a co-flow capillary device.  

Science.gov (United States)

A custom-made co-flow capillary device is used to synthesize monodisperse silk fibroin micro- and submicron-spheres with diameters tunable over a wide range of sizes. A model drug release is examined and control of degradation kinetics is obtained by changing sphere diameter. PMID:24339048

Mitropoulos, Alexander N; Perotto, Giovanni; Kim, Sunghwan; Marelli, Benedetto; Kaplan, David L; Omenetto, Fiorenzo G

2014-02-01

86

Fabrication of Chitosan/Silk Fibroin Composite Nanofibers for Wound-dressing Applications  

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Chitosan, a naturally occurring polysaccharide with abundant resources, has been extensively exploited for various biomedical applications, typically as wound dressings owing to its unique biocompatibility, good biodegradability and excellent antibacterial properties. In this work, composite nanofibrous membranes of chitosan (CS) and silk fibroin (SF) were successfully fabricated by electrospinning. The morphology of electrospun blend nanofibers was observed by scanning electron microscopy (...

Zeng-xiao Cai; Xiu-mei Mo; Kui-hua Zhang; Lin-peng Fan; An-lin Yin; Chuang-long He; Hong-sheng Wang

2010-01-01

87

Preparation and water absorption of cross-linked chitosan/silk fibroin blend films  

International Nuclear Information System (INIS)

Natural polymer blend films composed of chitosan and silk fibroin were prepared by varying the ratio of chitosan to silk fibroin, with and without glutaraldehyde as a crosslinking agent. The effects of the ratio of chitosan to silk fibroin and crosslinking agent on swelling behavior of the blend films were studied. For the swelling behavior, the blend films exhibited a dramatic change in the degree of swelling when immersed in acidic solutions. The degree of swelling of the films increased as the chitosan content increased; the blend film with 80% chitosan content had the maximum degree of swelling. It appeared that crosslinking had occurred in the blend films which helped the films to retain their three dimensional structure. In addition, FTIR spectra of the films showed evidence of hydrogen bonding interaction between chitosan and silk fibroin. For the effect of salt type, the films were immersed in various types of aqueous salt solutions, viz NaCl, LiCl, CaCl2, AlCl3, and FeCl3. The films immersed in AlCl3 and FeCl3 aqueous solutions gave the maximum degree of swelling. The effects of AlCl3 and FeCl3 concentrations on swelling behavior were also investigated. It was found that the maximum degree of swelling of the films occurred at 1.0 x 10-2 M of AlCl3 and FeCl3 aqueous solutions. (author)

2001-03-01

88

Formation of silk fibroin nanoparticles in water-miscible organic solvent and their characterization  

International Nuclear Information System (INIS)

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 ?-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 ?-helix form (Silk I) into anti-parallel ?-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 ?-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

2007-10-01

89

Solution structure of eggcase silk protein and its implications for silk fiber formation  

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Spider silks are renowned for their excellent mechanical properties and biomimetic and industrial potentials. They are formed from the natural refolding of water-soluble fibroins with ?-helical and random coil structures in silk glands into insoluble fibers with mainly ?-structures. The structures of the fibroins at atomic resolution and silk formation mechanism remain largely unknown. Here, we report the 3D structures of individual domains of a ?366-kDa eggcase silk protein that consists...

2009-01-01

90

Preparation and characterization of regenerated Bombyx mori silk fibroin fiber with high strength  

Directory of Open Access Journals (Sweden)

Full Text Available Regenerated Bombyx mori silk fibers were spun from hexafluoro-iso- propanol solution of silk fibroin sponge in methanol used as a coagulant solvent and then elongated in water. The stress-strain curves of the regenerated fibers changed dramatically depending on the draw ratio and the structure was studied by 13C CP/MAS NMR and X-ray diffraction methods. The patterns of 13C CP/MAS NMR spectra of two regenerated fibers with different draw ratios (1× and 3× and native silk fiber are all ?-sheet structure although the fraction of random coil/distorted ?-turn decreases in the order of 1×, 3× and native fiber gradually. On the other hand, azimuthal scans of their X-ray fiber patterns changed remarkably with increasing the draw ratio. This indicates that long-range orientation of the fibroin chain changes remarkably during the drawing process, but the short-range local structure does not change significantly. Regenerated silk fiber with a draw ratio of 3× is a fiber with high strength which is comparable with that of natural silk fiber. The regenerated fiber is also more degradable than natural silk fiber in enzyme solution in vitro.

2008-12-01

91

Freeze-gelled silk fibroin protein scaffolds for potential applications in soft tissue engineering.  

Science.gov (United States)

Recently tissue engineering has escalated much interest in biomedical and biotechnological applications. In this regard, exploration of new and suitable biomaterials is needed. Silk fibroin protein is used as one of the most preferable biomaterials for fabrication of scaffolds and several new techniques are being adopted to fabricate silk scaffolds with greater ease, efficiency and perfection. In this study, a freeze gelation technique is used for fabrication of silk fibroin protein 3D scaffolds, which is both time and energy efficient as compared to the conventional freeze drying technique. The fabricated silk fibroin freeze-gelled scaffolds are evaluated micro structurally for morphology with scanning electron microscopy which reveals relatively homogeneous pore structure and good interconnectivity. The pore sizes and porosity of these scaffolds ranges between 60-110?m and 90-95%, respectively. Mechanical test shows that the compressive strength of the scaffolds is in the range of 20-40kPa. The applicability to cell culture of the freeze gelled scaffolds has been examined with human keratinocytes HaCat cells which show the good cell viability and proliferation of cells after 5 days of culture suggesting the cytocompatibility. The freeze-gelled 3D scaffolds show comparable results with the conventionally prepared freeze dried 3D scaffolds. Thus, this technique may be used as an alternative method for 3D scaffolds preparation and may also be utilized for tissue engineering applications. PMID:21557966

Bhardwaj, Nandana; Chakraborty, Sagar; Kundu, Subhas C

2011-10-01

92

Effects of sterilization methods on the physical, chemical, and biological properties of silk fibroin membranes.  

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Silk fibroin has been widely explored for many biomedical applications, due to its biocompatibility and biodegradability. Sterilization is a fundamental step in biomaterials processing and it must not jeopardize the functionality of medical devices. The aim of this study was to analyze the influence of different sterilization methods in the physical, chemical, and biological characteristics of dense and porous silk fibroin membranes. Silk fibroin membranes were treated by several procedures: immersion in 70% ethanol solution, ultraviolet radiation, autoclave, ethylene oxide, and gamma radiation, and were analyzed by scanning electron microscopy, Fourier-transformed infrared spectroscopy (FTIR), X-ray diffraction, tensile strength and in vitro cytotoxicity to Chinese hamster ovary cells. The results indicated that the sterilization methods did not cause perceivable morphological changes in the membranes and the membranes were not toxic to cells. The sterilization methods that used organic solvent or an increased humidity and/or temperature (70% ethanol, autoclave, and ethylene oxide) increased the silk II content in the membranes: the dense membranes became more brittle, while the porous membranes showed increased strength at break. Membranes that underwent sterilization by UV and gamma radiation presented properties similar to the nonsterilized membranes, mainly for tensile strength and FTIR results. PMID:24259492

de Moraes, Mariana Agostini; Weska, Raquel Farias; Beppu, Marisa Masumi

2014-05-01

93

The effect of gamma irradiation on some morphological and quantitative changes of Bombyx mori L. silk gland  

International Nuclear Information System (INIS)

A study to determine the effect of gamma irradiation of silkworm eggs at doses of 1.00; 2.00 and 3.00 on silk gland weight, length and width is performed. It was found that gamma irradiation of eggs in the stage of embryo most intensive growth in length (B2) at doses 2.00 and 3.00 Gy stimulates increasing of silk glands weight in silkworms on the fifth day instar by 12-25 mg, as well as the silk worm width - by 7-33 ?m, which is of significant importance for the synthesis and secretion of silk proteins (fibroin and sericin). A breed specificity was also observed

1996-01-01

94

Non-mulberry silk fibroin influence osteogenesis and osteoblast-macrophage cross talk on titanium based surface.  

Science.gov (United States)

The titanium and its alloys are used as orthopedic dental implants due to their mechanical and bio-inert properties. The bare metal implants are not the ultimate answer for better osteogenesis and implant integration. Physical and chemical modifications are carried out to achieve the goal of improved adhesion and differentiation of the osteoblast. In this work, the silk fibroins from both mulberry and non-mulberry sources are used for surface modification. Silk fibroins are immobilized on titanium surface to facilitate the initial cell adhesion followed by improved cell spreading and better mineralization in order to achieve enhanced osseointegration. The immunological responses along with the effect of cytokines on osteoblast adhesion and function are investigated. The non-mulberry fibroin performs better in the context of the cell adherence and differentiation, which lead to better mineralization. The results indicate that the silk fibroin from non-mulberry source can be used for better osteogenesis on orthopedic implants. PMID:24752225

Naskar, Deboki; Nayak, Sunita; Dey, Tuli; Kundu, Subhas C

2014-01-01

95

Non-mulberry silk fibroin influence osteogenesis and osteoblast-macrophage cross talk on titanium based surface  

Science.gov (United States)

The titanium and its alloys are used as orthopedic dental implants due to their mechanical and bio-inert properties. The bare metal implants are not the ultimate answer for better osteogenesis and implant integration. Physical and chemical modifications are carried out to achieve the goal of improved adhesion and differentiation of the osteoblast. In this work, the silk fibroins from both mulberry and non-mulberry sources are used for surface modification. Silk fibroins are immobilized on titanium surface to facilitate the initial cell adhesion followed by improved cell spreading and better mineralization in order to achieve enhanced osseointegration. The immunological responses along with the effect of cytokines on osteoblast adhesion and function are investigated. The non-mulberry fibroin performs better in the context of the cell adherence and differentiation, which lead to better mineralization. The results indicate that the silk fibroin from non-mulberry source can be used for better osteogenesis on orthopedic implants.

Naskar, Deboki; Nayak, Sunita; Dey, Tuli; Kundu, Subhas C.

2014-01-01

96

Effect of the sterilization method on the properties of Bombyx mori silk fibroin films  

International Nuclear Information System (INIS)

We have compared the effects of different sterilization techniques on the properties of Bombyx mori silk fibroin thin films with the view to subsequent use for corneal tissue engineering. The transparency, tensile properties, corneal epithelial cell attachment and degradation of the films were used to evaluate the suitability of certain sterilization techniques including gamma-irradiation (in air or nitrogen), steam treatment and immersion in aqueous ethanol. The investigations showed that gamma-irradiation, performed either in air or in a nitrogen atmosphere, did not significantly alter the properties of films. The films sterilized by gamma-irradiation or by immersion in ethanol had a transparency greater than 98% and tensile properties comparable to human cornea and amniotic membrane, the materials of choice in the reconstruction of ocular surface. Although steam-sterilization produced stronger, stiffer films, they were less transparent, and cell attachment was affected by the variable topography of these films. It was concluded that gamma-irradiation should be considered to be the most suitable method for the sterilization of silk fibroin films, however, the treatment with ethanol is also an acceptable method. - Highlights: ? The effects of four methods of sterilization on the properties of silk fibroin films were investigated. ? Steam treatment leads to stiffer films but to lower transparency and variable surface topography. ? Degradation of fibroin is enhanced in the films that were gamma-irradiated. ? The effects on mechanical properties are explained through changes in both primary and secondary structure of fibroin. ? Gamma-irradiation and immersion in aqueous ethanol are suggested as preferred methods of sterilization.

2013-03-01

97

Effect of the sterilization method on the properties of Bombyx mori silk fibroin films  

Energy Technology Data Exchange (ETDEWEB)

We have compared the effects of different sterilization techniques on the properties of Bombyx mori silk fibroin thin films with the view to subsequent use for corneal tissue engineering. The transparency, tensile properties, corneal epithelial cell attachment and degradation of the films were used to evaluate the suitability of certain sterilization techniques including gamma-irradiation (in air or nitrogen), steam treatment and immersion in aqueous ethanol. The investigations showed that gamma-irradiation, performed either in air or in a nitrogen atmosphere, did not significantly alter the properties of films. The films sterilized by gamma-irradiation or by immersion in ethanol had a transparency greater than 98% and tensile properties comparable to human cornea and amniotic membrane, the materials of choice in the reconstruction of ocular surface. Although steam-sterilization produced stronger, stiffer films, they were less transparent, and cell attachment was affected by the variable topography of these films. It was concluded that gamma-irradiation should be considered to be the most suitable method for the sterilization of silk fibroin films, however, the treatment with ethanol is also an acceptable method. - Highlights: Black-Right-Pointing-Pointer The effects of four methods of sterilization on the properties of silk fibroin films were investigated. Black-Right-Pointing-Pointer Steam treatment leads to stiffer films but to lower transparency and variable surface topography. Black-Right-Pointing-Pointer Degradation of fibroin is enhanced in the films that were gamma-irradiated. Black-Right-Pointing-Pointer The effects on mechanical properties are explained through changes in both primary and secondary structure of fibroin. Black-Right-Pointing-Pointer Gamma-irradiation and immersion in aqueous ethanol are suggested as preferred methods of sterilization.

George, Karina A. [Queensland Eye Institute, 41 Annerley Road, South Brisbane, Queensland 4101 (Australia); Institute of Health and Biomedical Innovation, Queensland University of Technology, Kelvin Grove, Queensland 4059 (Australia); Shadforth, Audra M.A. [Queensland Eye Institute, 41 Annerley Road, South Brisbane, Queensland 4101 (Australia); Chirila, Traian V., E-mail: traian.chirila@qei.org.au [Queensland Eye Institute, 41 Annerley Road, South Brisbane, Queensland 4101 (Australia); Australian Institute of Bioengineering and Nanotechnology, University of Queensland, St Lucia, 4072 (Australia); Faculty of Health Sciences, University of Queensland, Herston, Queensland 4006 (Australia); Faculty of Science and Engineering, Queensland University of Technology, Brisbane, Queensland 4001 (Australia); Laurent, Matthieu J. [Queensland Eye Institute, 41 Annerley Road, South Brisbane, Queensland 4101 (Australia); Ecole Superieure d' Ingenieurs de Luminy (ESIL), Universite de la Mediterranee Aix-Marseille II, Luminy case 925 13288, Marseille, Cedex 09 (France); Stephenson, Sally-Anne [Institute of Health and Biomedical Innovation, Queensland University of Technology, Kelvin Grove, Queensland 4059 (Australia); Faculty of Health, Queensland University of Technology, Brisbane, Queensland 4001 (Australia); Edwards, Grant A. [Australian Institute of Bioengineering and Nanotechnology, University of Queensland, St Lucia, 4072 (Australia); Madden, Peter W. [Queensland Eye Institute, 41 Annerley Road, South Brisbane, Queensland 4101 (Australia); Faculty of Health Sciences, University of Queensland, Herston, Queensland 4006 (Australia); and others

2013-03-01

98

Drug loading and release on tumor cells using silk fibroin–albumin nanoparticles as carriers  

International Nuclear Information System (INIS)

Polymeric and biodegradable nanoparticles are frequently used in drug delivery systems. In this study silk fibroin–albumin blended nanoparticles were prepared using the desolvation method without any surfactant. These nanoparticles are easily internalized by the cells, reside within perinuclear spaces and act as carriers for delivery of the model drug methotrexate. Methotrexate loaded nanoparticles have better encapsulation efficiency, drug loading ability and less toxicity. The in vitro release behavior of methotrexate from the nanoparticles suggests that about 85% of the drug gets released after 12 days. The encapsulation and loading of a drug would depend on factors such as size, charge and hydrophobicity, which affect drug release. MTT assay and conjugation of particles with FITC demonstrate that the silk fibroin–albumin nanoparticles do not affect the viability and biocompatibility of cells. This blended nanoparticle, therefore, could be a promising nanocarrier for the delivery of drugs and other bioactive molecules. (paper)

2013-01-25

99

Print head design and control for electrohydrodynamic printing of silk fibroin.  

Science.gov (United States)

This study investigates the effect of print head design on the electrohydrodynamic printed resolution of silk fibroin. Needles with large orifices measuring at 800 ?m were used to build five different print heads. The print heads were manufactured, tested, and optimized using four different silk fibroin solution concentrations of 10 wt.%, 15 wt.%, 20 wt.%, and 22 wt.% at applied voltages that ranged from 10 to 20 kV with two different flow rates of 1.5 ?l/min and 2.0 ?l/min. Each print head design behaved in a unique manner in terms of printed line characteristics as the flow rate, voltage and concentration were varied. The highest printed resolution of the order of 1 ?m was achieved using the pinhole reservoir print head. Possible explanations for each of the observed behaviors and design criteria for future print heads are discussed. PMID:23706215

Hashimdeen, S H; Miodownik, M; Edirisinghe, M J

2013-08-01

100

Study on Preparation of n-HA and Silk Fibroin Bio-mineral Material  

Digital Repository Infrastructure Vision for European Research (DRIVER)

A new hydroxyapatite (n-HA)ª²Silk fibroin (SF) bio-mineral material was prepared by using a biomimetic method, with Ca(NO3)2 and Na3PO4 used as the starting reagent to synthesize the inorganic phase. In details, SF was dissolved in Ca(NO3)2 solution without desalting procedure, the compound solution was directly dropped into Na3PO4 solution. In this way, HA was synthesized at 37¡?y ...

Wang Jiang, Zuo Yi

2009-01-01

 
 
 
 
101

Fabrication and Intermolecular Interactions of Silk Fibroin/Hydroxybutyl Chitosan Blended Nanofibers  

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The native extracellular matrix (ECM) is composed of a cross-linked porous network of multifibril collagens and glycosaminoglycans. Nanofibrous scaffolds of silk fibroin (SF) and hydroxybutyl chitosan (HBC) blends were fabricated using 1,1,1,3,3,3-hexafluoro-2-propanol (HFIP) and trifluoroacetic acid (TFA) as solvents to biomimic the native ECM via electrospinning. Scanning electronic microscope (SEM) showed that relatively uniform nanofibers could be obtained when 12% SF was blended with 6% ...

Kui-Hua Zhang; Qiao-Zhen Yu; Xiu-Mei Mo

2011-01-01

102

Preparation and water absorption of cross-linked chitosan/silk fibroin blend films  

Energy Technology Data Exchange (ETDEWEB)

Natural polymer blend films composed of chitosan and silk fibroin were prepared by varying the ratio of chitosan to silk fibroin, with and without glutaraldehyde as a crosslinking agent. The effects of the ratio of chitosan to silk fibroin and crosslinking agent on swelling behavior of the blend films were studied. For the swelling behavior, the blend films exhibited a dramatic change in the degree of swelling when immersed in acidic solutions. The degree of swelling of the films increased as the chitosan content increased; the blend film with 80% chitosan content had the maximum degree of swelling. It appeared that crosslinking had occurred in the blend films which helped the films to retain their three dimensional structure. In addition, FTIR spectra of the films showed evidence of hydrogen bonding interaction between chitosan and silk fibroin. For the effect of salt type, the films were immersed in various types of aqueous salt solutions, viz NaCl, LiCl, CaCl{sub 2}, AlCl{sub 3}, and FeCl{sub 3}. The films immersed in AlCl{sub 3} and FeCl{sub 3} aqueous solutions gave the maximum degree of swelling. The effects of AlCl{sub 3} and FeCl{sub 3} concentrations on swelling behavior were also investigated. It was found that the maximum degree of swelling of the films occurred at 1.0 x 10{sup -2} M of AlCl{sub 3} and FeCl{sub 3} aqueous solutions. (author)

Suesat, Jantip; Rujiravanit, Ratana [Chulalongkorn University, The Petroleum and Petrochemical College, Bangkok (Thailand); Jamieson, Alexander M. [Case Western Reserve Univ., Department of Macromolecular Science, Cleveland (United States); Tokura, Seiichi [Kansai Univ., Faculty of Engineering, Osaka (Japan)

2001-03-01

103

Morphology, Secondary Structure and Thermal Properties of Silk Fibroin/Gelatin Blend Film  

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This study aimed to prepare Silk Fibroin (SF) and Gelatin (G) blend film and study its morphology, secondary structure and thermal properties compared to native SF and G films. The films were prepared from the SF solution by casting on the polystyrene plates. They were investigated their secondary structure by fourier transform-infrared (FTIR) spectroscopy, morphology using Scanning Electron Microscope (SEM). In addition, Thermogravimetric Analysis (TG) and Differential TG (DTG) were used for...

2009-01-01

104

Conformation Transition and Thermal Properties Study of Silk Fibroin and Poly (?-Caprolactone) Blends  

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In this study, influence of intermolecular interactions between Silk Fibroin (SF) and Poly (?-Caprolactone) (PCL) in homogeneous blend form on SF conformation changes and thermal properties of the blends was investigated and discussed. The SF/PCL blends were prepared by solution blending and precipitating method, respectively. Dimethylsulfoxide and isopropanol were used as a solvent and a non-solvent, respectively. The blends were characterized by Fourier Transform Infrared Spectroscop...

Srisa-ard, M.; Baimark, Y.; Srisuwan, Y.

2008-01-01

105

Fabrication and Intermolecular Interactions of Silk Fibroin/Hydroxybutyl Chitosan Blended Nanofibers  

Digital Repository Infrastructure Vision for European Research (DRIVER)

The native extracellular matrix (ECM) is composed of a cross-linked porous network of multifibril collagens and glycosaminoglycans. Nanofibrous scaffolds of silk fibroin (SF) and hydroxybutyl chitosan (HBC) blends were fabricated using 1,1,1,3,3,3-hexa?uoro-2-propanol (HFIP) and trifluoroacetic acid (TFA) as solvents to biomimic the native ECM via electrospinning. Scanning electronic microscope (SEM) showed that relatively uniform nanofibers could be obtained when 12% SF was blended with 6%...

2011-01-01

106

Silk fibroin-based complex particles with bioactive encrustation for bone morphogenetic protein 2 delivery.  

Science.gov (United States)

Application of bone morphogenetic protein 2 (BMP-2) currently faces its challenges, and its efficacy of delivery has to be improved. The proper dosage of the powerful bioactive molecule is still under discussion and needs to be investigated further. In this work, pure silk fibroin particles and particles with calcium carbonate encrustation (complex particles) are designed, developed, and functionalized by BMP-2. These are used to deliver the bioactive molecule to mesenchymal stem cells (MSCs) to induce osteogenic differentiation. Results are compared with those of control groups of BMP-2 carriers under the same condition. Silk fibroin-based particles with size and component variations are prepared by self-assembly, desolvation, and soft template formation to improve BMP-2 loading efficiency. Results show that the particles significantly enhance osteogenic differentiation of MSCs, which is evident in the high ALP enzyme activity as well as the increased level of expression of osteogenic genes. Specifically, the combination of calcium compound and BMP-2 in the silk fibroin-calcium carbonate complex particles synergistically enhances osteogenesis. Release tests and mathematical modeling are applied to describe BMP-2 dissolution profiles, and the release mechanism is based on diffusion and polymer chain relaxation. In summary, the particles show high efficacies of BMP-2 delivery, and introduction of the complex particle can progressively enhance osteogenesis. PMID:24224461

Shi, Pujiang; Abbah, Sunny A; Saran, Kushagra; Zhang, Yong; Li, Jun; Wong, Hee-Kit; Goh, James C H

2013-12-01

107

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

Directory of Open Access Journals (Sweden)

Full Text Available 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.

M. Fazley Elahi

2014-04-01

108

Effects of RGDS sequence genetically interfused in the silk fibroin light chain protein on chondrocyte adhesion and cartilage synthesis.  

Science.gov (United States)

Initial chondrocyte-silk fibroin interactions are implicated in chondrogenesis when using fibroin as a scaffold for chondrocytes. Here, we focused on integrin-mediated cell-scaffold adhesion and prepared cell adhesive fibroin in which a tandem repeat of the Arg-Gly-Asp-Ser (RGDS) sequence was genetically interfused in the fibroin light chain (L-chain) (L-RGDSx2 fibroin). We investigated the effects of the sequence on chondrocyte adhesion and cartilage synthesis, in comparison to the effects of fibronectin. As the physicochemical surface properties (e.g., wettability and zeta potential) of the fibroin substrate were not affected by the modification, specific cell adhesion to the RGDS predominately changed the chondrocyte adhesive state. This suggestion was also supported by the competitive inhibition of chondrocyte attachment to the L-RGDSx2 fibroin substrate with soluble RGD peptides in the medium. Unlike fibronectin, the expression of RGDS in the fibroin L-chain had no effect on chondrocyte spreading area but enhanced mRNA expression levels of integrins alpha5 and beta1, and aggrecan at 12 h after seeding. Although both the sequence and fibronectin increased cell adhesive force, chondrocytes grown on the fibroin substrate exhibited a peak in the force with time in culture. These results suggested that moderate chondrocyte adhesion to fibroin induced by the RGDS sequence was able to maintain the chondrogenic phenotype and, from the histology findings, the sequence could facilitate chondrogenesis. PMID:20643479

Kambe, Yusuke; Yamamoto, Koji; Kojima, Katsura; Tamada, Yasushi; Tomita, Naohide

2010-10-01

109

Evaluation of the properties of silk fibroin films from the non-mulberry silkworm Samia cynthia ricini for biomaterial design.  

Science.gov (United States)

Silk fibroin from a domesticated mulberry silkworm, Bombyx mori, is the most widely used in biomaterial design. We report for the first time the preparation of a relatively smooth (granule free) film of the nonmulberry Samia cynthia ricini fibroin for comparative evaluation of its cell-supporting properties against those of the B. mori fibroin film. The granule formation on the S. c. ricini fibroin film was successfully prevented by facilitating proper rearrangement of the protein molecules, as monitored by FT-IR, by dialysis through a stepwise decrease in the urea concentration in the dialysis media. The lower contact angle of the S. c. ricini fibroin film, compared to the B. mori fibroin film, corresponds well to its lower hydrophobic/hydrophilic amino-acid ratio and grand average of hydropathicity (GRAVY). L929 murine fibroblast cells on the granule-free S. c. ricini fibroin films exhibited greater proliferation and spreading rates than those on the B. mori fibroin films, possibly attributable to its higher content of hydrophilic and positively charged amino acids. It further suggests that fabrication, modification and/or engineering of S. c. ricini fibroin may provide a better biomaterial scaffold design than the more commonly used B. mori fibroin. PMID:21029516

Mai-ngam, Katanchalee; Boonkitpattarakul, Kanhokthorn; Jaipaew, Jirayut; Mai-ngam, Bunpot

2011-01-01

110

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

Science.gov (United States)

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.

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

2009-03-01

111

Multifunctional adhesive silk fibroin with blending of RGD-bioconjugated mussel adhesive protein.  

Science.gov (United States)

Silk has recently been exploited in various fields due to its superior mechanical properties. However, this material's lack of biological functions and relatively poor biodegradation have hindered its wide use in applications related to cells and tissues. Here, we improved the overall characteristics of silkworm silk fibroin (SF) by introduction of RGD peptide-fused recombinant mussel adhesive protein (MAP-RGD). Simple blending of MAP-RGD provided not only bulk-scale adhesive ability but also microscale adhesiveness to cells and various biomolecules. MAP-RGD-blended SF fibers supported enhanced adhesion, proliferation, and spreading of mammalian cells as well as the efficient attachment of biomolecules, including carbohydrate and protein. In addition, the hydrophilicity, swelling, and biodegradability of the MAP-RGD-blended SF material were improved without notable hampering of the original mechanical properties of SF. Therefore, the adhesive silk fibrous scaffold could be successfully used in diverse biomedical engineering applications. PMID:24601579

Yang, Yun Jung; Kwon, Yunkyeoung; Choi, Bong-Hyuk; Jung, Dooyup; Seo, Jeong Hyun; Lee, Ki Hoon; Cha, Hyung Joon

2014-04-14

112

Tubuliform silk protein: A protein with unique molecular characteristics and mechanical properties in the spider silk fibroin family  

Science.gov (United States)

Orb-web weavers can produce up to six different types of silk and a glue for various functions. Tubuliform silk is unique among them due to its distinct amino acid composition, specific time of production, and atypical mechanical properties. To study the protein composing this silk, tubuliform gland cDNA libraries were constructed from three orb-weaving spiders Argiope aurantia, Araneus gemmoides, and Nephila clavipes. Amino acid composition comparison between the predicted tubuliform silk protein sequence (TuSp1) and the corresponding gland protein confirms that TuSp1 is the major component in tubuliform gland in three spiders. Sequence analysis suggests that TuSp1 shares no significant similarity with its paralogues, while it has conserved sequence motifs with the most primitive spider, Euagrus chisoseus silk protein. The presence of large side-chain amino acids in TuSp1 sequence is consistent with the frustrated ?-sheet crystalline structure of tubuliform silk observed in transmission electron microscopy. Repeat unit comparison within species as well as among three spiders exhibits high sequence conservation. Parsimony analysis based on carboxy terminal sequence shows that Argiope and Araneus are more closely related than either is to Nephila which is consistent with phylogenetic analysis based on morphological evidence.

Tian, M.; Lewis, R. V.

2006-02-01

113

Silk fibroin microparticles as carriers for delivery of human recombinant bone morphogenetic protein-2 : in vitro and in vivo bioactivity  

Digital Repository Infrastructure Vision for European Research (DRIVER)

The in vitro and in vivo efficiency of fibroin microparticles as a delivery carrier for bone morphogenetic protein-2 (BMP-2) was evaluated. BMP-2 was encapsulated in silk fibroin particles that were produced by a simple and very mild processing method. The dose-response of BMP-2-loaded fibroin particles was examined in C2C12 cells, after 5 days of culture. The BMP-2 retained most of its activity as observed by the increase in alkaline phosphatase activity, which was much higher when BMP-2 was...

2010-01-01

114

Fabrication of poly(lactic-co-glycolic acid) scaffolds containing silk fibroin scaffolds for tissue engineering applications.  

Science.gov (United States)

The present study deals with the fabrication of poly(lactic-co-glycolic acid) (PLGA) scaffolds modified with silk fibroin for biomedical application. The PLGA solutions were added with salt particles and pressed with high pressures; which were further subjected to salt leaching resulting in the creation of large sized pores in the PLGA scaffolds. To fill up these pores, 2%, 4%, and 8% of silk solutions were added, however, the addition created extra small sized pores. The scaffolds were characterized by various state of techniques; the scanning electronic microscopy revealed the large sized pores in the pristine scaffold can be tailored into smaller architecture by the addition of silk fibroin. The contact angle measurements confirmed the introduction of silk helped to change the hydrophobic nature of PLGA into hydrophilic, which is the main constrain for PLGA. The mechanical properties of scaffold can be easily improved by applying the higher amounts of silk into the scaffolds. The thermal gravimetric analyses and fourier transform infrared spectroscopy confirmed the presence of silk fibroin in scaffolds. The cell viability and cell attachment was checked by culturing the scaffolds with NIH 3T3 fibroblasts and chondrocytes. Furthermore, these results revealed that the introduction of silk had significant impact on the viability of fibroblast also had a good affinity for cell attachment and infiltration of human chondrocytes in scaffolds after culturing the cells for 2 and 5 weeks of time. © 2013 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 102A: 2713-2724, 2014. PMID:24026912

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

2014-08-01

115

Unique molecular architecture of silk fibroin in the waxmoth.  

Czech Academy of Sciences Publication Activity Database

. Ro?. 277, ?. 25 (2002), s. 22639-22647. ISSN 0021-9258Grant CEP: GA ?R GA204/96/1100; GA ?R GA204/00/0019; GA MŠk ME 204Výzkumný zám?r: CEZ:AV0Z5007907Klí?ová slova: silkKód oboru RIV: EG - ZoologieImpakt faktor: 6.696, rok: 2002

Žurovec, Michal; Sehnal, František

116

Rheological properties of native silk fibroins from domestic and wild silkworms, and flow analysis in each spinneret by a finite element method.  

Science.gov (United States)

Silkworms can produce strong and tough fibers at room temperature and from an aqueous solution. Therefore, it seems useful to study the mechanism of fiber formation by silkworms for development of synthetic polymers with excellent mechanical properties. The rheological behaviors of native silk dopes stored in the silk glands of Bombyx mori and Samia cynthia ricini were clarified, and flow simulations of the dopes in each spinneret were performed with a Finite Element Method. Dynamic viscoelastic measurements revealed that silk fibroin stored in silk glands forms a transient network at room temperature, and that the molecular weight for the network node corresponds to the molecular weight of a heterodimer of H-chain and L-chain (B. mori) and a homodimer of H-chains (S. c. ricini), respectively. Also, each dope exhibited zero-shear viscosity and then shear thinning like polymer melts. In addition, shear thickening due to flow-induced crystallization was observed. The critical shear rate for crystallization of B. mori dopes was smaller than that of S. c. ricini dopes. From the flow simulation, it is suggested that domestic and wild silkworms are able to crystallize the dopes in the stiff plate region by controlling shear rate using the same magnitude of extrusion pressure despite differences in rheological properties. PMID:19317399

Moriya, Motoaki; Roschzttardtz, Frederico; Nakahara, Yusuke; Saito, Hitoshi; Masubuchi, Yuichi; Asakura, Tetsuo

2009-04-13

117

Preparing silk fibroin nanofibers through electrospinning: further heparin immobilization toward hemocompatibility improvement.  

Science.gov (United States)

Sodium heparin (HS) was immobilized on the surface of the silk fibroin nanofibers (FS) prepared by electrospinning with the objective of improving the hemocompatibility of the fibers for application as scaffolds in tissue engineering. The nanofiber mats of silk fibroin without (MF-FS) and with (MF-FS/HS) immobilized heparin were characterized through scanning electron microscopy (SEM), Fourier-transform infrared spectroscopy with attenuated total reflectance (FTIR-ATR), thermogravimetric analyses (TGA), energy dispersive spectroscopy (EDS), contact angle, chemical analysis, and biological tests. The formation of hydrogen bonds between the silk fibroin and heparin was discussed based on FTIR-ATR spectra. The amount of immobilized heparin was quantified through papain/N-acetyl-l-cysteine digestion followed by dimethylmethylene blue complexation. Furthermore, the samples with immobilized HS showed higher hydrophilic capability compared to samples without HS due to lower contact angles. It was possible to verify that the capillary end-to-collector distance of 8.5 cm and flow rate of 0.35 mL h(-1) used in the electrospinning process at 20 kV are good conditions for obtaining a small average fiber diameter maintaining the amount of immobilized heparin on MF-FS/HS in ca. 4% w/w. Biological analysis showed that no hemolysis is provoked by MF-FS and MF-FS/HS mat fragments and those such mats are not toxic to Vero cells. However, the MF-FS/HS showed higher cell growth and proliferation than MF-FS, indicating an improvement in the hemocompatibility of the material due to heparin immobilization. PMID:24724905

Cestari, Marília; Muller, Vinícius; Rodrigues, Jean Henrique da Silva; Nakamura, Celso V; Rubira, Adley F; Muniz, Edvani C

2014-05-12

118

Collagen-reinforced electrospun silk fibroin tubular construct as small calibre vascular graft.  

Science.gov (United States)

None of the replacements proposed in the literature for small-calibre blood vessels (SCBV) fully satisfies the stringent requirements that these grafts have to fulfil. Here, an electrospun silk fibroin tubular construct is hybridized with type I collagen gel to produce a biomimetic SCBV graft with physiologically relevant compliance and burst pressure and optimal cytocompatibility. The hybridization of the two polymers results in the formation of a nanofibrillar hydrated matrix, where the collagen gel enhances the mechanical properties of the SF tubular construct and improves the early response of the material to in vitro cell adhesion and proliferation. PMID:23060093

Marelli, Benedetto; Achilli, Matteo; Alessandrino, Antonio; Freddi, Giuliano; Tanzi, Maria Cristina; Farè, Silvia; Mantovani, Diego

2012-11-01

119

Study on the preparation of collagen-modified silk fibroin films and their properties  

International Nuclear Information System (INIS)

Blended films were prepared from a silk fibroin (SF) solution by adding a small amount of type I collagen (<5%). The mechanical properties of the wet films modified by collagen were improved obviously. The elongation at break reached 42%, and the smaller contact angles revealed that modified films had better hydrophilicity. 1% heparin was also added to modify the SF films to further improve the in vitro antithrombogenecity. The internal structure of the modified SF films was investigated with scanning electron microscopy, x-ray diffraction and Fourier transform infrared attenuated total reflection spectroscopy. The result indicates that the addition of a small amount of collagen and heparin did not change their conformation

2006-12-01

120

Preparation and Characterization of Eri (Philosamia ricini) Silk Fibroin Powder  

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Eri (Philosamia ricini) cocoons were dissolved with 9 M Ca(NO3)2 and then dialyzed against distilled water for 3 days. The Silk Fibroin (SF) solution was used as substrate to prepare SF powder by using freeze-drying method. The secondary structure and thermal behavior of SF powder were determined by FT-IR and TGA analyzer, respectively. The SF powder was arranged in micrometer sizes. FT-IR spectra indicated that the SF powder composed of ?-helix and ?-sheet stru...

Yaowalak, S.; Wilaiwan, S.; Prasong, S.

2009-01-01

 
 
 
 
121

The application with tetramethyl pyrazine for antithrombogenicity improvement on silk fibroin surface  

Science.gov (United States)

Chuanxiongqin (tetramethyl pyrazine, TMPZ) is an active ingredient of the Chinese herb and was used to improve the anticoagulant activity of silk fibroin (SF). The side methyl of TMPZ was oxidized, and then linked to polyacrylic acid (PAA) via an ester bond. The prepared conjugate was further mixed with SF solutions at different ratios to make blend films. The resulting products were characterized by FTIR, UV spectrometer and X-ray photoelectron spectroscopy (XPS). The in vitro antithrombogenicity were evaluated by the activated partial thromboplastin time (APTT) and the prothrombin time (PT). It was shown that blend films had longer coagulation time than the pure SF film.

Lian, Xiaojie; Wang, Song; Xu, Guoliang; Lin, Nini; Li, Qian; Zhu, Hesun

2008-11-01

122

The biosynthesis of transfer RNA in insects. I. Increase of amino acid acceptor activity of specific tRNA's utilized for silk protein biosynthesis in the silk gland of Bombyx mori.  

Science.gov (United States)

1) To detect the quantitative changes of amino acid acceptor activity of tRNA's from the posterior and middle silk glands of Bombyx mori at various ages, a relatively simple and rapid method was established using a mixture of radioactive amino acids in Chlorella hydrolysate. 2) The acceptor activities of silk gland tRNA for 15 amino acids tested seemed to be almost on the same level at the end of the 4th moult stage. During the 5th instar, however, characteristic increases were observed in glycine, alanine, and serine acceptor activities in both silk glands. 3) In the posterior silk gland, which produces fibroin, the acceptor activities for glycine and alanine increased more than that for serine. In the middle silk gland, which produces sericine, the acceptor activity for serine increased more than those for glycine and alanine. 4) In the light of observations on the increase of corresponding aminoacyl-tRNA synthetase activities in the silk glands, a functional adaptation of tRNA synthesis in the tissue is discussed. PMID:1228175

Majima, R; Kawakami, M; Shimura, K

1975-08-01

123

Dielectric breakdown strength of regenerated silk fibroin films as a function of protein conformation.  

Science.gov (United States)

Derived from Bombyx mori cocoons, regenerated silk fibroin (RSF) exhibits excellent biocompatibility, high toughness, and tailorable biodegradability. Additionally, RSF materials are flexible, optically clear, easily patterned with nanoscale features, and may be doped with a variety bioactive species. This unique combination of properties has led to increased interest in the use of RSF in sustainable and biocompatible electronic devices. In order to explore the applicability of this biopolymer to the development of future bioelectronics, the dielectric breakdown strength (Ebd) of RSF thin films was quantified as a function of protein conformation. The application of processing conditions that increased ?-sheet content (as determined by FTIR analysis) and produced films in the silk II structure resulted in RSF materials with improved Ebd with values reaching up to 400 V/?m. PMID:23987229

Dickerson, Matthew B; Fillery, Scott P; Koerner, Hilmar; Singh, Kristi M; Martinick, Katie; Drummy, Lawrence F; Durstock, Michael F; Vaia, Richard A; Omenetto, Fiorenzo G; Kaplan, David L; Naik, Rajesh R

2013-10-14

124

Silk fibroin protein from mulberry and non-mulberry silkworms: cytotoxicity, biocompatibility and kinetics of L929 murine fibroblast adhesion.  

Science.gov (United States)

Silks fibers and films fabricated from fibroin protein of domesticated mulberry silkworm cocoon have been traditionally utilized as sutures in surgery and recently as biomaterial films respectively. Here, we explore the possibility of application of silk fibroin protein from non-mulberry silkworm cocoon as a potential biomaterial aid. In terms of direct inflammatory potential, fibroin proteins from Antheraea mylitta and Bombyx mori are immunologically inert and invoke minimal immune response. Stimulation of murine peritoneal macrophages and RAW 264.7 murine macrophages by these fibroin proteins both in solution and in the form of films assayed in terms of nitric oxide and TNFalpha production showed comparable stimulation as in collagen. Kinetics of adhesion of L929 murine fibroblasts, for biocompatibility evaluation, monitored every 4 h from seeding and studied over a period of 24 h, reveal A. mylitta fibroin film to be a better substrate in terms of rapid and easier cellularization. Cell viability studies by MTT assay and flow cytometric analyses indicate the ability of fibroin matrices to support cell growth and proliferation comparable to collagen for long-term culture. This matrix may have potential to serve in those injuries where rapid cellularization is essential. PMID:18322779

Acharya, Chitrangada; Ghosh, Sudip K; Kundu, S C

2008-08-01

125

In vitro study on silk fibroin textile structure for anterior cruciate ligament regeneration.  

Science.gov (United States)

A novel hierarchical textile structure made of silk fibroin from Bombyx mori capable of matching the mechanical performance requirements of anterior cruciate ligament (ACL) and in vitro cell ingrowth is described. This sericin-free, Silk Fibroin Knitted Sheath with Braided Core (SF-KSBC) structure was fabricated using available textile technologies. Micro-CT analysis confirmed that the core was highly porous and had a higher degree of interconnectivity than that observed for the sheath. The in vivo cell colonization of the scaffolds is thus expected to penetrate even the internal parts of the structure. Tensile mechanical tests demonstrated a maximum load of 1212.4±56.4 N (under hydrated conditions), confirming the scaffold's suitability for ACL reconstruction. The absence of cytotoxic substances in the extracts of the SF-KSBC structure in culture medium was verified by in vitro tests with L929 fibroblasts. In terms of extracellular matrix production, Human Periodontal Ligament Fibroblasts (HPdLFs) cultured in direct contact with SF-KSBC, compared to control samples, demonstrated an increased secretion of aggrecan (PG) and fibronectin (FBN) at 3 and 7 days of culture, and no change in IL-6 and TNF-? secretion. Altogether, the outcomes of this investigation confirm the significant utility of this novel scaffold for ACL tissue regeneration. PMID:23910255

Farè, Silvia; Torricelli, Paola; Giavaresi, Gianluca; Bertoldi, Serena; Alessandrino, Antonio; Villa, Tomaso; Fini, Milena; Tanzi, Maria Cristina; Freddi, Giuliano

2013-10-01

126

High-affinity integration of hydroxyapatite nanoparticles with chemically modified silk fibroin  

International Nuclear Information System (INIS)

Hydroxyapatite (HA)-based nanocomposites were prepared by a co-precipitation method with silk fibroin (SF) serving as organic matrix. Silk fibroin was chemically modified with an alkali solution or an enzyme attempting to improve the interface between the mineral and the organic matrix. The influences of the alkali and enzyme pretreatments on microstructure and physicochemical properties of HA-SF composite were examined and compared. The results reveal that both the two kinds of pretreatments facilitate the formation of highly ordered three-dimensional porous network throughout the composites, increase the microhardness of the composite, and promote the preferential growth of HA crystallites along c-axis. Among all the as-prepared samples, the composite containing the enzyme pretreated SF shows desirable hierarchical microstructure with higher degree of organization and more uniform pore size distribution. Due to the enzyme pretreatment, HA crystallites undergo obvious changes in morphology from rod-like to whisker-like and in crystal growth towards more apparent epitaxy along c-axis. The alkali pretreatment induces the stronger chemical interactions between HA and SF and thus to strengthen the inorganic-organic interfacial adhesion. The newly developed HA-SF composites are expected to be attractive biomedical materials for bone repair and remodeling

2007-10-01

127

Study on Preparation of n-HA and Silk Fibroin Bio-mineral Material  

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Full Text Available A new hydroxyapatite (n-HAª²Silk fibroin (SF bio-mineral material was prepared by using a biomimetic method, with Ca(NO32 and Na3PO4 used as the starting reagent to synthesize the inorganic phase. In details, SF was dissolved in Ca(NO32 solution without desalting procedure, the compound solution was directly dropped into Na3PO4 solution. In this way, HA was synthesized at 37¡?y controlling Ca/P ratio, meanwhile the minerialized HA interacted with SF to form biomineral material. The morphology and structure of this n-HA-SF biomineral material were characterized by XRD, FTIR, XPS and SEM. The results show that there are strong chemical interaction between n-HA and SF. Moreover, in the biomineral material, the inorganic phase is calciumª²deficient hydroxyapatite, containing a small amount of carbonated HA with an average crystal size of 37.6nm. In addition, The particle size of biomineral materials range from 50nm to 200nm.It can be deduced that silk fibroin chain can regulate the growth and morphology of HA crystals. The compressive strength of the biomineral material is 32.21MPa£¬ which is a good potential material for bone tissue engineering and unloadedª²bearing bone defects repairing.

WANG Jiang,ZUO Yi,YANG Wei-Hu,ZHOU Gang,ZHANG Li,LI Yu-Bao

2009-03-01

128

Bio-inspired capillary dry spinning of regenerated silk fibroin aqueous solution  

International Nuclear Information System (INIS)

To biomimic the spinning process of silkworm or spider, a capillary spinning equipment was applied to spin regenerated silk fibroin (RSF) fibers from RSF aqueous solutions in air. This equipment exhibits a wide processing window for various RSF aqueous solutions. The effects of pH, metal ions, RSF concentration and spinning parameters on the spinnability of the spinning dope and the mechanical properties of the obtained fibers were investigated. As a result, spinning dopes with a pH from 5.2 to 6.9 have good spinnability, especially for the dope with a pH of 6.0 and a Ca2+ concentration of 0.3 M. The RSF concentration of this dope ranges from 44% to 48%. Under optimized conditions of our dry spinning experiments (L/D, 133; take-up speed, 30 mm/s), the obtained as-spun fiber has a breaking strength of 46 MPa, which can be improved up to 359 MPa after a preliminary post-drawing in 80 vol.% ethanol aqueous solution. Highlights: ? Regenerated silk fibroin fibers were prepared by using a dry spinning method. ? Dope compositions affect dope spinnability. ? Spinning parameters affect dope spinnability and fiber properties. ? The breaking stress of the post-treated fiber was up to 359 MPa.

2011-10-10

129

Bio-inspired capillary dry spinning of regenerated silk fibroin aqueous solution  

Energy Technology Data Exchange (ETDEWEB)

To biomimic the spinning process of silkworm or spider, a capillary spinning equipment was applied to spin regenerated silk fibroin (RSF) fibers from RSF aqueous solutions in air. This equipment exhibits a wide processing window for various RSF aqueous solutions. The effects of pH, metal ions, RSF concentration and spinning parameters on the spinnability of the spinning dope and the mechanical properties of the obtained fibers were investigated. As a result, spinning dopes with a pH from 5.2 to 6.9 have good spinnability, especially for the dope with a pH of 6.0 and a Ca{sup 2+} concentration of 0.3 M. The RSF concentration of this dope ranges from 44% to 48%. Under optimized conditions of our dry spinning experiments (L/D, 133; take-up speed, 30 mm/s), the obtained as-spun fiber has a breaking strength of 46 MPa, which can be improved up to 359 MPa after a preliminary post-drawing in 80 vol.% ethanol aqueous solution. Highlights: {yields} Regenerated silk fibroin fibers were prepared by using a dry spinning method. {yields} Dope compositions affect dope spinnability. {yields} Spinning parameters affect dope spinnability and fiber properties. {yields} The breaking stress of the post-treated fiber was up to 359 MPa.

Wei Wei; Zhang Yaopeng, E-mail: zyp@dhu.edu.cn; Zhao Yingmei; Luo Jie; Shao Huili; Hu Xuechao

2011-10-10

130

Contribution to the ultrastructural study of silk-excretion cells and autoradiographic analysis of intracellular fibroin transport in Bombyx mori L  

International Nuclear Information System (INIS)

It is much easier to study the mechanisms involved in the synthesis and exportation of extracellular proteins in the biological material chosen is highly differentiated. The silk-excretion gland of the silkworm is ideal in this respect because during the larva period, especially at the end of the 5th and last stage, the cells at the rear (excreting tube) synthesize and export massive quantities of a single protein: fibroin. These phenomena were explored by a cytological study carried out mainly by electron microscopy and autoradiography. The results obtained are given. They relate first of all to the morphological development of the secretion tube cells from the end of the 4th larva stage to the spinning of the cocoon, and contribute new information on the cell changes during the 4th slough and the end of the 5th age. They also concern intracellular fibroin transport which is proved to take place through the Golgi apparatus, and finally the possible role of the microtubules and microfilaments in fibroin transport and secretion. On this last point the results so far constitute only, a preliminary approach which justifie no final conclusions; they merely suggest that the microfilaments of the apical region are involved in the secretion process

1974-01-01

131

Fabrication of highly interconnected porous silk fibroin scaffolds for potential use as vascular grafts.  

Science.gov (United States)

Silk fibroin (SF) scaffolds have been designed and fabricated for multiple organ engineering owing to SF's remarkable mechanical property, excellent biocompatibility and biodegradability, as well as its low immunogenicity. In this study, an easy-to-adopt and mild approach based on a modified freeze-drying method was developed to fabricate a highly interconnected porous SF scaffold. The physical properties of the SF scaffold, including pore morphology, pore size, porosity and compressive modulus, could be adjusted by the amount of ethanol added, the freezing temperature and the concentration of SF. Fourier transform infrared spectroscopy illustrated that treatment of the lyophilized scaffolds with 90% methanol led to a structure transition of SF from silk I (random coil) to silk II (beta-sheet), which stabilized the SF scaffolds in water. We also incorporated heparin during fabrication to obtain a heparin-loaded scaffold which possessed excellent anticoagulant property. The heparin that was incorporated into the SF scaffolds could be released in a sustain manner for approximately 7days, inhibiting the proliferation of human smooth muscle cells within the scaffold in vitro while promoting neovascularization in vivo. We therefore propose that the SF porous scaffold fabricated here may be an attractive candidate for use as a potential vascular graft for implantation based on its high porosity, excellent blood compatibility and mild fabrication process. PMID:24486642

Zhu, Meifeng; Wang, Kai; Mei, Jingjing; Li, Chen; Zhang, Jiamin; Zheng, Wenting; An, Di; Xiao, Nannan; Zhao, Qiang; Kong, Deling; Wang, Lianyong

2014-05-01

132

Structure and structural changes of the silk fibroin from Samia cynthia ricini using nuclear magnetic resonance spectroscopy.  

Science.gov (United States)

The structure of silk fibroin from a wild silkworm, S. c. ricini, the amino acid sequence of which consists of repeated poly-Ala and Gly-rich regions, was examined by using solution and solid-state NMR methods. The structural transition of the silk fibroin in aqueous solution was monitored by using 13C solution NMR spectroscopy as a function of temperature. The fast exchange with respect to the chemical shift between the helix and coil conformations was observed in the poly-Ala region and the slow conformational change from alpha-helix to random coil was observed for the Gly residue adjacent to the N-terminal Ala residue of the poly-Ala region. The torsion angles of several Ala and Gly residues in the model peptide, GGAGGGYGGDGG(A)12GGA-GDGYGAG, were determined by the conformation-dependent 13C chemical shifts, rotational echo double resonance (REDOR) and 2D spin-diffusion NMR methods. The solid-state NMR analysis leads to the precise silk structure before spinning, where the poly-Ala sequence takes a typical alpha-helix pattern with a tightly winded helical structure at both terminal regions of the poly-Ala sequence. This is expected to stabilize the alpha-helical structure of the poly-Ala region in S. c. ricini silk fibroin from the silkworm. PMID:15468207

Asakura, Tetsuo; Nakazawa, Yasumoto

2004-03-15

133

Preparation of Eri silk fibroin and gelatin blend film loaded chlorhexidine using as model for hydrophilic drug release  

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Full Text Available The objective of this research was to prepare Eri silk fibroin solution for preparing silk film loaded chlorhexidine drug as model for hydrophilic drug release. The Eri silk cocoons were boiled in 0.5%NaCO3 solution at 90?, and then left in air dried at room temperature. The fibroin was dissolved in 9M (Ca(NO32 with ethanol (2 by mole and heated at 70?. The silk fibroin (SF solution was then dialyzed to exclude salt in phosphate buffer. The SF and gelatin (G solutions were mixed for preparation of films in both with and without chlorhexidine. The films were observed their morphology under scanning electron microscope. The results found that all of films were rough of their surfaces, homogeneous texture without phase separation. The native SF film composed of pores throughout the film area but did not observe in native G film. The results also showed that the SF and G can be good interacted to form hydrogen bonds. These were indicated from FTIR spectra and thermal analysis. The chlorhexidine drug has not affect on the changes of film properties. However, the releasing pattern of chlorhexidine from each film was varied. The highest rate of drug releasing was found in the native SF film while the native G film was the lowest. It might be suggested that the drug releasing rate was depended on polarity of each polymer components.

Chirapha Butiman

2012-07-01

134

Conformation Transition and Thermal Properties Study of Silk Fibroin and Poly (?-Caprolactone Blends  

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Full Text Available In this study, influence of intermolecular interactions between Silk Fibroin (SF and Poly (?-Caprolactone (PCL in homogeneous blend form on SF conformation changes and thermal properties of the blends was investigated and discussed. The SF/PCL blends were prepared by solution blending and precipitating method, respectively. Dimethylsulfoxide and isopropanol were used as a solvent and a non-solvent, respectively. The blends were characterized by Fourier Transform Infrared Spectroscopy (FTIR, Differential Scanning Calorimetry (DSC, Thermo Gravimetry (TG and Scanning Electron Microscopy (SEM. Conformation transition of SF component from ?-sheet to random coil forms can be induced by blending with PCL. Melting temperature and heat of melting of the PCL decreased as increasing the SF ratio. Thermal stability of the SF can be enhanced by blending with PCL. FTIR, DSC and TG results suggested that intermolecular hydrogen bonds were formed between SF and PCL molecules in the blends. Homogeneous morphology of blends was illustrated by SEM micrographs.

M. Srisa-Ard

2008-01-01

135

Effect of Methanol Treatment on Regenerated Silk Fibroin Microparticles Prepared by the Emulsification-Diffusion Technique  

Directory of Open Access Journals (Sweden)

Full Text Available Silk Fibroin (SF microparticles containing hollow structure were prepared by a water-in-oil emulsion solvent diffusion method without any surfactants. Aqueous SF solution and ethyl acetate were used as water and oil phases, respectively. Influences of SF concentration and post methanol treatment on microparticle characteristics were investigated. All microparticles contained open hollow structures. Microparticle sizes increased with the SF concentration. Conformation of SF microparticles determined from FTIR spectra changed from predominantly random coil to ?-sheet form after methanol treatment. Particle surfaces of methanol-treated SF microparticles were rougher than those of non-treated. The SF concentrations and methanol treatment did not affect open hollow structure of the SF microparticles. Major advantages of this method are the elimination of surfactants and extractable pore templates. These hollow SF microparticles are expected to be potential used as biodegradable microcarriers of cell and protein drug, because their hollow structures should permit cell attachment and enzyme immobilization.

Yodthong Baimark

2009-01-01

136

Structural characteristics and biological performance of silk fibroin nanofiber containing microalgae spirulina extract.  

Science.gov (United States)

Silk fibroin (SF) nanofiber scaffold containing microalgae Spirulina extract were prepared by electrospinning and the performance and functionality of the scaffold were evaluated. The viscosity and conductivity of the dope solution of Spirulina containing SF were examined for electrospinability and we found that the morphological structure of SF nanofiber is affected by the concentration of Spirulina extract added. The platelet adhesion and coagulation time test confirmed that the Spirulina containing SF nanofiber scaffold had excellent ability to prevent blood clotting or antithrombogenicity that is comparable to heparin. Low cytotoxicity and excellent cell adhesion and proliferation were also observed for Sprulina containing SF nanofiber scaffold by methylthiazolyldiphenyl-tetrazolium bromide assay and confocal fluorescence microscope using fibroblast and human umbilical vein endothelial cells. Based on these results, we believe SF nanofiber scaffold containing Spirulina extract has the potential to be used as tissue engineering scaffold that requires high hemocompatibility. © 2013 Wiley Periodicals, Inc. Biopolymers 101: 307-318, 2014. PMID:23868372

Cha, Bum-Gyu; Kwak, Hyo Won; Park, A Reum; Kim, Shin Hwan; Park, Sook-Young; Kim, Hyun-Jeong; Kim, Ick-Soo; Lee, Ki Hoon; Park, Young Hwan

2014-04-01

137

Deciphering the mechanism of protein interaction with silk fibroin for drug delivery systems.  

Science.gov (United States)

Silk fibroin (SF) is an exceptional drug delivery carrier with respect to stabilizing, protecting, and delivering sensitive biologics. A synopsis of thermodynamic, static light scattering, hydrophobicity probing, and nanoparticle tracking analyses served as a basis to decipher the mechanism of interaction between SF and two model proteins, protamine and polylysine. The impact of salts aiding (chaotropic), not affecting (neutral), or opposing (cosmotropic) SF unfolding was a major determinant, ranging from complete abolishment to maximal interaction efficacy. Evidence is provided, that the underlying mechanism of the remarkable ability to tailor drug/SF interaction throughout such large ranges and by appropriate salt selection is the control of structural breakdown of SF micelles as present in pure SF ad initium. This study provides a mechanistically justified and hypothesis driven blueprint for future experimental designs addressing the controlled interaction of biologics and SF. PMID:24461326

Germershaus, Oliver; Werner, Vera; Kutscher, Marika; Meinel, Lorenz

2014-03-01

138

A Novel Nano composite Particle of Hydroxyapatite and Silk Fibroin: Biomimetic Synthesis and Its Biocompatibility  

International Nuclear Information System (INIS)

A novel bone-like bio material of hydroxyapatite (HAP) and silk fibroin (SF) composite was developed by biomimetic synthesis. The composite was precipitated from drops of Ca(OH)2 suspension and H3PO4 solution with SF. With this method, the HAP nano crystals were obtained by self-assembling on a SF surface whose c-axis was aligned with the long-axis direction of SF in microstructures; this shares the same misconstrues of collagen and HAP with that in the natural bone. The HAP/SF composite then demonstrated that it could promote osteoblast proliferation in vitro and new bone formation in vivo. The novel bio material is a promising material for bone replacement and regeneration.

2010-01-01

139

Characterization of silk fibroin modified surface: a proteomic view of cellular response proteins induced by biomaterials.  

Science.gov (United States)

The purpose of this study was to develop the pathway of silk fibroin (SF) biopolymer surface induced cell membrane protein activation. Fibroblasts were used as an experimental model to evaluate the responses of cellular proteins induced by biopolymer material using a mass spectrometry-based profiling system. The surface was covered by multiwalled carbon nanotubes (CNTs) and SF to increase the surface area, enhance the adhesion of biopolymer, and promote the rate of cell proliferation. The amount of adhered fibroblasts on CNTs/SF electrodes of quartz crystal microbalance (QCM) greatly exceeded those on other surfaces. Moreover, analyzing differential protein expressions of adhered fibroblasts on the biopolymer surface by proteomic approaches indicated that CD44 may be a key protein. Through this study, utilization of mass spectrometry-based proteomics in evaluation of cell adhesion on biopolymer was proposed. PMID:24818131

Yang, Ming-Hui; Yuan, Shyng-Shiou; Chung, Tze-Wen; Jong, Shiang-Bin; Lu, Chi-Yu; Tsai, Wan-Chi; Chen, Wen-Cheng; Lin, Po-Chiao; Chiang, Pei-Wen; Tyan, Yu-Chang

2014-01-01

140

A Novel Nanocomposite Particle of Hydroxyapatite and Silk Fibroin: Biomimetic Synthesis and Its Biocompatibility  

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Full Text Available A novel bone-like biomaterial of hydroxyapatite (HAP and silk fibroin (SF composite was developed by biomimetic synthesis. The composite was precipitated from drops of Ca(OH2 suspension and H3PO4 solution with SF. With this method, the HAP nanocrystals were obtained by self-assembling on a SF surface whose c-axis was aligned with the long-axis direction of SF in microstructures; this shares the same misconstrues of collagen and HAP with that in the natural bone. The HAP/SF composite then demonstrated that it could promote osteoblast proliferation in vitro and new bone formation in vivo. The novel biomaterial is a promising material for bone replacement and regeneration.

Lin Niu

2010-01-01

 
 
 
 
141

Silk fibroin/gelatin blend films crosslinked with enzymes for biomedical applications.  

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Microbial transglutaminase (mTG) and mushroom tyrosinase (MT) are used to crosslink B. mori silk fibroin/gelatin (SF/G) films. Crosslinked and uncrosslinked SF/G films show no phase separation. The thermal behavior and the conformational structure of SF/G films are strongly affected by blending and enzymatic treatment. Formation of high thermally stable crosslinked macromolecular species is observed, suggesting the occurrence of strong intermolecular interactions between the two polymers as confirmed by FT-Raman spectroscopy. Preliminary in vitro tests show that MT-crosslinked blends with G amounts ?40% and mTG-crosslinked SF/G 60/40 films support C2C12 cardiomyocyte adhesion and proliferation. PMID:23956190

Taddei, Paola; Chiono, Valeria; Anghileri, Anna; Vozzi, Giovanni; Freddi, Giuliano; Ciardelli, Gianluca

2013-11-01

142

Microstructural, thermal and antibacterial properties of electron beam irradiated Bombyx mori silk fibroin films  

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The Bombyx mori silk fibroin (SF) films were prepared by solution casting method and the effects of electron beam on structural, thermal and antibacterial responses of the prepared films were studied. The electron irradiation for different doses was carried out using 8 MeV Microtron facility at Mangalore University. The changes in microstructural parameters and thermal stability of the films were investigated using Wide Angle X-ray Scattering (WAXS) and thermogravimetric analysis (TGA) respectively. Both microstructuralline parameters (crystallite size and lattice strain (g in %)) and thermal stability of the irradiated films have increased with radiation dosage. Agar diffusion method demonstrated the antibacterial activity of SF film which was increased after irradiation on both Gram-positive and Gram-negative species.

Asha, S.; Sangappa, Naik, Prashantha; Chandra, K. Sharat; Sanjeev, Ganesh

2014-04-01

143

Modification of silk fibroin using diazonium coupling chemistry and the effects on hMSC proliferation and differentiation.  

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A simple chemical modification method using diazonium coupling chemistry was developed to tailor the structure and hydrophilicity of silk fibroin protein. The extent of modification using several aniline derivatives was characterized using UV-vis and 1H NMR spectroscopies, and the resulting protein structure was analyzed with ATR-FTIR spectroscopy. Introduction of hydrophobic functional groups facilitated rapid conversion of the protein from a random coil to a beta-sheet structure, while addition of hydrophilic groups inhibited this process. hMSCs were grown on these modified silks to assess the biocompatibility of these materials. The hydrophilicity of the silk derivatives was found to affect the growth rate and morphology, but hMSCs were able to attach, proliferate and differentiate into an osteogenic lineage on all of the silk derivatives. PMID:18417206

Murphy, Amanda R; St John, Peter; Kaplan, David L

2008-07-01

144

Study of the effects of different sterilization methods on the properties of dense and porous silk fibroin membranes  

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Silk fibroin has been widely explored for many biomedical applications, due to its biocompatibility and biodegradability. Sterilization is a fundamental step in biomaterials processing, and it must not alter in a negative way the functionality of medical devices. The aim of this study was to analyze the influence of different sterilization methods in the physical and chemical characteristics of dense silk fibroin membranes. Dense fibroin membranes were sterilized by ultraviolet radiation, 70% ethanol, autoclave, ethylene oxide and gamma radiation, and were analyzed by SEM, FTIR-ATR and XRD. The results for sterilization indicated that the methods didn't cause degradation of the membranes, but the methods that used organic solvent, or increase of humidity and/or temperature (70% ethanol, autoclave and ethylene oxide) altered the molecular conformation of fibroin, increasing the proportion of ?-sheet structure, what indicates an increase of crystallinity. This effect may be positive when a slower degradation of the membranes is desired, depending on the application as a bio material. (author)

2009-10-13

145

Modification of Silk Fibroin Using Diazonium Coupling Chemistry and the Effects on hMSC Proliferation and Differentiation  

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A simple chemical modification method using diazonium coupling chemistry was developed to tailor the structure and hydrophilicity of silk fibroin protein. The extent of modification using several aniline derivatives was characterized using UV/vis and 1H NMR spectroscopy, and the resulting protein structure was analyzed with ATR-FTIR spectroscopy. Introduction of hydrophobic functional groups facilitated rapid conversion of the protein from a random coil to a ?-sheet structure, while addition...

2008-01-01

146

Effect of initial cell seeding density on 3D-engineered silk fibroin scaffolds for articular cartilage tissue engineering  

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The repair of articular cartilage defects poses a continuing challenge. Cartilage tissue engineering through the culture of chondrocytes seeded in 3D porous scaffolds has the potential for generating constructs that repair successfully. It also provides a platform to study scaffold-cell and cell-cell interactions. The scaffold affects the growth and morphology of cells growing on it, and concomitantly, cells affect the properties of the resultant tissue construct. Silk fibroin protein from An...

2011-01-01

147

Tightly winding structure of sequential model peptide for repeated helical region in Samia cynthia ricini silk fibroin studied with solid-state NMR  

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There are many kinds of silks from silkworms and spiders with different structures and properties, and thus, silks are suitable to study the structure-property relationship of fibrous proteins. Silk fibroin from a wild silkworm, Samia cynthia ricini, mainly consists of the repeated similar sequences by about 100 times where there are alternative appearances of the polyalanine (Ala)12–13 region and the Gly-rich region. In this paper, a sequential model peptide, GGAGGGYGGDGG(A)12GGAGDGYGAG, w...

2003-01-01

148

An emerging functional natural silk biomaterial from the only domesticated non-mulberry silkworm Samia ricini.  

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Mulberry silk fibroin is a widely used biomaterial and recent work on non-mulberry silk fibroin also suggests it may have similar uses. We expect silk fibroin from the only domesticated non-mulberry eri silkworm, Samia ricini, to possess useful properties as a biomaterial. Eri silk gland fibroin is a heterodimeric protein of approximately 450?kDa. Cytocompatibility evaluation with fibroblasts and osteoblast-like cells shows good cell attachment, viability and proliferation. The matrices, which have high thermal stability and good swellability, are also haemocompatible. Eri silk production is cost effective as no agronomic practices are required for their host plant cultivation. This fibroin provide new opportunities as an alternative natural functional biomaterial in various biomedical applications. PMID:23733347

Pal, Shilpa; Kundu, Joydip; Talukdar, Sarmistha; Thomas, Tintu; Kundu, Subhas C

2013-08-01

149

Formulation and characterization of silk fibroin films as a scaffold for adipose-derived stem cells in skin tissue engineering.  

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Skin substitutes are epidermal, dermal or complete bilayered constructs, composed by natural or synthetic scaffolds and by adherent cells such as fibroblasts, keratinocytes or mesenchymal stem cells. Silk fibroin is a promising polymer to realize scaffolds, since it is biocompatible, biodegradable, and exhibits excellent mechanical properties in terms of tensile strength. Moreover, fibroin can be added of others components in order to modify the biomaterial properties for the purpose. The aim of this work is to prepare silk fibroin films for adipose-derived stem cell (ADSCs) culture as a novel feeder layer for skin tissue engineering. Pectin has been added to promote the protein conformational transition and construct strength, while glycerol as plasticizer, providing biomaterial flexibility. Eighteen formulations were prepared by casting method using fibroin, pectin (range 1-10% w/w), and glycerol (range 0-20% w/w); films were characterized by Fourier transform infrared spectroscopy and differential scanning calorimetry assay, to select the optimal composition. A stable fibroin conformation was obtained using 6% w/w pectin, and the best mechanical properties were obtained using 12% w/w glycerol. Films were sterilized, and human ADSCs were seeded and cultured for 15 days. Cells adhere to the support assuming a fibroblastic-like shape and reaching confluence. The ultrastructural analysis evidences typical active-cell features and adhesion structures that promote cell anchorage to the film, thus developing a multilayered cell structure. This construct could be advantageously employed in cutaneous wound healing or where the use of ADSCs scaffold is indicated either in human or veterinary field. PMID:24046948

Chlapanidas, T; Tosca, M C; Faragò, S; Perteghella, S; Galuzzi, M; Lucconi, G; Antonioli, B; Ciancio, F; Rapisarda, V; Vigo, D; Marazzi, M; Faustini, M; Torre, M L

2013-01-01

150

Silk fibroin microparticles as carriers for delivery of human recombinant bone morphogenetic protein-2: in vitro and in vivo bioactivity.  

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The in vitro and in vivo efficiency of fibroin microparticles as a delivery carrier for bone morphogenetic protein-2 (BMP-2) was evaluated. BMP-2 was encapsulated in silk fibroin particles that were produced by a simple and very mild processing method. The dose-response of BMP-2-loaded fibroin particles was examined in C2C12 cells, after 5 days of culture. The BMP-2 retained most of its activity as observed by the increase in alkaline phosphatase activity, which was much higher when BMP-2 was encapsulated into the particles rather than just surface-adsorbed. After 2 weeks of culture, increased mineralization was observed with BMP-2-loaded particles in comparison to soluble added growth factor. No significant cytotoxicity was detected. When implanted in a rat ectopic model, bone formation was observed by in vivo micro-computed tomography after 2 and 4 weeks postimplantation, with particles loaded with 5 or 12.5?microg BMP-2. An increase in bone density was observed over time. Histology revealed further evidence of ectopic bone formation, observed by strong alizarin red staining and osteocalcin immunostaining. Our findings show that fibroin microparticles may present an interesting option for future clinical applications in the bone tissue engineering field, and therefore, further studies have been planned. PMID:19958078

Bessa, Paulo César; Balmayor, Elizabeth R; Hartinger, Joachim; Zanoni, Gerald; Dopler, Daniela; Meinl, Alexandra; Banerjee, Asmita; Casal, Margarida; Redl, Heinz; Reis, Rui L; van Griensven, Martijn

2010-10-01

151

Injectable silk fibroin/polyurethane composite hydrogel for nucleus pulposus replacement.  

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In degenerative disc disease, an injectable hydrogel can fill a degenerate area completely, reduce the risk of implant migration and subsequent loss of height of the intervertebral disc, and minimise surgical defects. Here, we propose a method of preparing an injectable silk fibroin/polyurethane (SF/PU) composite hydrogel by chemical cross-linking under physiological conditions. Mechanical testing was used to determine the mechanical strength of the hydrogel. The impact of hydrogel height on the biomechanical properties was discussed to estimate the working capacity of the hydrogel for further clinical application. Rheological properties were also examined to assess the practical ability of the hydrogel for clinical application. Hydrogel injection and cell assessment is also of interest for clinical application. An SF/PU composite hydrogel can be injected through a small incision. A cell proliferation assay using bone marrow stromal cells showed positive cell viability and increased proliferation over a seven-day period in culture. Importantly, the hydrogel can be monitored in real-time using X-ray fluoroscopy during and after surgery according to the results of X-ray fluoroscopy examination, and shows good visibility based on X-ray assays. In particular, the hydrogel offers the clinically important advantage of visibility in CT and T2-weighted magnetic resonance imaging. Based on the results of the current study, the SF/AU composite hydrogel may offer several advantages for future application in nucleus pulposus replacement. PMID:22231270

Hu, Jingen; Chen, Bin; Guo, Fang; Du, Jingyu; Gu, Pengcheng; Lin, Xiangjin; Yang, Weiping; Zhang, Hailong; Lu, Min; Huang, Yiping; Xu, Gewen

2012-03-01

152

Mesenchymal stem cell-seeded multilayered dense collagen-silk fibroin hybrid for tissue engineering applications.  

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Tissue engineering of multilayered constructs that model complex tissues poses a significant challenge for regenerative medicine. In this study, a three-layered scaffold consisting of an electrospun silk fibroin (SF) mat sandwiched between two dense collagen (DC) layers was designed and characterized. It was hypothesized that the SF layer would endow the DC-SF-DC construct with enhanced mechanical properties (e.g., apparent modulus, tensile strength, and toughness), while the surrounding DC layers provide an extracellular matrix-like environment for mesenchymal stem cell (MSC) growth. MSC-seeded DC-SF-DC hybrids were produced using the plastic compression technique and characterized morphologically, chemically, and mechanically. Moreover, MSC viability was assessed for up to 1 wk in culture. Scaffold analyses confirmed compaction and integration of the meso-scaled multilayered DC-SF-DC hybrid, which was reflected in a significantly higher toughness value when compared to DC and SF alone. MSCs directly incorporated into the DC layers remained viable for up to day 7. The ease of multilayered construct fabrication, enhanced biomechanical properties, along with uniformity of cell distribution confirmed the possibility for the incorporation and segregation of different cell types within distinct layers for the regeneration of complex tissues, such as skin, or central nervous system dura mater. PMID:21751393

Ghezzi, Chiara E; Marelli, Benedetto; Muja, Naser; Hirota, Nobuaki; Martin, James G; Barralet, Jake E; Alessandrino, Antonio; Freddi, Giuliano; Nazhat, Showan N

2011-10-01

153

Morphology and thermal stability of silk fibroin/starch blended microparticles  

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Full Text Available In the present study biodegradable microparticles of silk fibroin (SF/starch blends were prepared by a simple water-in-oil emulsion solvent diffusion technique. SF/starch blended solution and ethyl acetate were used as water and oil phases, respectively. The influence of SF/starch ratios on characteristics of the blended microparticles was investigated. The SF conformation of microparticle matrices from FTIR analysis was changed from random coil to ?-sheet form by blending with starch. The blended microparticles had lower dissolution in water than those of SF and starch microparticles. The 1/3 (w/w SF/starch blended microparticles exhibited the lowest dissolution. The SF and starch microparticles showed irregular and deflated shapes, respectively. The blended microparticles were nearly spherical in shapes and smaller sizes. Thermal stability of the blended microparticles slightly increased with the starch blended ratio. The results suggested that SF conformational transition, thermal stability, morphology and dissolution of the blended microparticles can be adjusted by varying the blended ratio.

2010-12-01

154

Mechanically-reinforced electrospun composite silk fibroin nanofibers containing hydroxyapatite nanoparticles.  

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Electrospun silk fibroin (SF) scaffolds provide large surface area, high porosity, and interconnection for cell adhesion and proliferation and they may replace collagen for many tissue engineering applications. Despite such advantages, electrospun SF scaffolds are still limited as bone tissue replacement due to their low mechanical strengths. While enhancement of mechanical strengths by incorporating inorganic ceramics into polymers has been demonstrated, electrospinning of a mixture of SF and inorganic ceramics such as hydroxyapatite is challenging and less studied due to the aggregation of ceramic particles within SF. In this study, we aimed to enhance the mechanical properties of electrospun SF scaffolds by uniformly dispersing hydroxyapatite (HAp) nanoparticles within SF nanofibers. HAp nanoaprticles were modified by ?-glycidoxypropyltrimethoxysilane (GPTMS) for uniform dispersion and enhanced interfacial bonding between HAp and SF fibers. Optimal conditions for electrospinning of SF and GPTMS-modified HAp nanoparticles were identified to achieve beadless nanofibers without any aggregation of HAp nanoparticles. The MTT and SEM analysis of the osteoblasts-cultured scaffolds confirmed the biocompatibility of the composite scaffolds. The mechanical properties of the composite scaffolds were analyzed by tensile tests for the scaffolds with varying contents of HAp within SF fibers. The mechanical testing showed the peak strengths at the HAp content of 20 wt.%. The increase of HAp content up to 20 wt.% increased the mechanical properties of the composite scaffolds, while further increase above 20 wt.% disrupted the polymer chain networks within SF nanofibers and weakened the mechanical strengths. PMID:24857500

Kim, Hyunryung; Che, Lihua; Ha, Yoon; Ryu, WonHyoung

2014-07-01

155

Elimination of large particulate units from silk fibroin PLD films by post-treatments  

International Nuclear Information System (INIS)

After depositing silk fibroin (SF) thin films by pulsed IR-laser deposition, extraordinarily large particulate units up to several micrometers were observed. They include debris from the target and severely agglomerated protein units. Occurrence of those large particles was found to be minimum on the vertical substrate. We tried to eliminate large particulate units by two post-treatment operations, i.e. dry gaseous blow-off (GBO) and rinsing in water under simultaneous ultrasonication (WSU). Change in the surface structure by these post-treatments was observed by optical and electron microscopes with varying area from 1mm square down to 1?m square. GBO turned out to be suitable to eliminate the lightly attached particulates of 1-10?m, mostly those pulled out from the target while preserving morphological and chemical properties of smallest units underneath. WSU, on the other hand, pelt off more strongly attached surface irregularities. However, morphological change with an increase in the surface roughness in the range of 1nm was also observed after WSU. The latter might be associated with possible sonochemical effects

2007-04-01

156

Study of synthesis of nano-hydroxyapatite using a silk fibroin template  

International Nuclear Information System (INIS)

Nano-hydroxyapatite (HA) was directly synthesized on a silk fibroin (SF) template using the property of SF being soluable in a concentrated CaCl2 solution as a HA source of calcium at pH 7.4 and room temperature. The microstructure and bonding state were investigated by x-ray diffraction (XRD), Fourier transform infrared spectroscopy (FTIR), differential scanning calorimetry-thermogravimetry analysis (DSC-TG) and transmission electron microscopy (TEM). The results indicated that the HA crystals were poorly crystallized with a rod-like shape of 20-60 nm length and 10-20 nm diameter. Strong molecular interactions and chemical bonds might be present between SF and HA. There were other nucleation sites such as carbonyl (-C-O) and amine (-N-H-) groups on SF molecules besides the carboxyl (-COOH) and hydroxyl (-OH) groups previously reported. During the formation of HA, the coordination action between specific functional groups on SF and calcium ions (Ca2+) played an important role. The crystallinity of HA was improved and had an orientation growth along (0 0 2) at the presence of SF, resulting in a structure similar to natural bone. It was concluded that SF could regulate the structure and morphology of HA effectively. (communication)

2010-08-01

157

Tip-induced micropatterning of silk fibroin protein using in situ solution atomic force microscopy.  

Science.gov (United States)

Silk fibroin (SF) is a promising candidate for a variety of application in the fields of tissue engineering, drug delivery, and biomedical optics. Recent research has already begun to explore the construction of nano- and micropatterned SF films under ambient environment. The structure and biocompatibility of SF are dependent on SF state (solution or solid) and the method of drying the SF solution to prepare various biomaterials such as films, sponges, and fibers. Therefore, it is important to explore the construction of SF nano- and micropatterns under aqueous solution. This paper reports a novel application of atomic force microscopy (AFM) under liquid for direct deposition of the relatively hydrophobic protein SF onto hydrophilic mica. We demonstrate that the AFM tip, in either the contact or the tapping mode, can fabricate SF micropatterns on mica with controlled surface topography. We show that the deposition process requires a mechanical force-induced SF sol-gel transition followed by a transfer to the mica surface at the tip-surface contact, and the efficiency of this process depends on not only AFM operation mode but also the SF bulk concentration, the SF amount on mica, and the AFM tip spring constant. PMID:23276203

Zhong, Jian; Ma, Mengjia; Zhou, Juan; Wei, Daixu; Yan, Zhiqiang; He, Dannong

2013-02-01

158

Preparation and in vitro characterization of biomorphic silk fibroin scaffolds for bone tissue engineering.  

Science.gov (United States)

In bone tissue engineering, a qualified scaffold should provide a three-dimensional porous structure mimicking the extracellular matrix of bone except good biological properties. In this study, biomorphic silk fibroin (SF) scaffolds were fabricated from cane by an innovative biotemplating-negative mold process. The physicochemical properties, in vitro enzymatic degradation behavior and biocompatibility of the biomorphic SF scaffolds were investigated. The results showed that the scaffolds well inherited the original porous morphology of cane, and possessed good mechanical stability. The scaffolds had a compressive modulus of 1.56?±?0.08 MPa and a porosity of 82.73%, and exhibited a bimodal pore size distribution (15 and 172 ?m). The degradation ratio of the SF scaffolds increased with prolonging degradation time and reached 29% within 21 days when exposed to 1.0 U/mL collagenase IA. The in vitro cytocompatibility evaluation indicated that the scaffolds could support cell attachment, proliferation, and osteogenic differentiation of osteoblast-like MC3T3-E1 cells, as assessed by SEM, fluorescent staining, MTT, and ALP activity assays. The results indicated the potential of biomorphic SF scaffolds for bone tissue engineering. © 2013 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 102A: 2961-2971, 2014. PMID:24123779

Qian, Junmin; Suo, Aili; Jin, Xinxia; Xu, Weijun; Xu, Minghui

2014-09-01

159

Fabrication and Intermolecular Interactions of Silk Fibroin/Hydroxybutyl Chitosan Blended Nanofibers  

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Full Text Available The native extracellular matrix (ECM is composed of a cross-linked porous network of multifibril collagens and glycosaminoglycans. Nanofibrous scaffolds of silk fibroin (SF and hydroxybutyl chitosan (HBC blends were fabricated using 1,1,1,3,3,3-hexa?uoro-2-propanol (HFIP and trifluoroacetic acid (TFA as solvents to biomimic the native ECM via electrospinning. Scanning electronic microscope (SEM showed that relatively uniform nanofibers could be obtained when 12% SF was blended with 6% HBC at the weight ratio of 50:50. Meanwhile, the average nano?brous diameter increased when the content of HBC in SF/HBC blends was raised from 20% to 100%. Fourier transform infrared spectra (FTIR and 13C nuclear magnetic resonance (NMR showed SF and HBC molecules existed in hydrogen bonding interactions but HBC did not induce conformation of SF transforming from random coil form to ?-sheet structure. X-ray diffraction (XRD confirmed the different structure of SF/HBC blended nanofibers from both SF and HBC. Thermogravimetry-Differential thermogravimetry (TG-DTG results demonstrated that the thermal stability of SF/HBC blend nanofibrous scaffolds was improved. The results indicated that the rearrangement of HBC and SF molecular chain formed a new structure due to stronger hydrogen bonding between SF and HBC. These electrospun SF/HBC blended nanofibers may provide an ideal tissue engineering scaffold and wound dressing.

Xiu-Mei Mo

2011-03-01

160

In vitro and in vivo Evaluation of Hemocompatibility of Silk Fibroin Based Artificial Vascular Grafts  

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Full Text Available Artificial vascular grafts with low thrombogenicity are generally required to avoid blood platelet adhesion and to minimize intimal hyperplasia, thus retaining vascular patency. In this study, we aimed to determine the acute and subacute hemocompatibility of silk fibroin (SF grafts by in vitro and in vivo evaluation. Blood contact reaction with SF grafts was examined by thrombin-anti-thrombin III complex (TAT formation, platelet activation level by beta-thromboglobulin (beta-TG, complement system response (C3a and SC5b-9, platelet and fibrin deposition and compared with commercially available polyethylene terephthalate (PET artificial grafts in vitro. The biocompatibility and coagulation-inducing effect of coating materials were evaluated by in vivo implantation in rats. Two weeks after implantation, SF grafts showed low subacute coagulation. All blood parameters evaluated for animals implanted with SF-coated grafts showed almost the same values as those for sham-operated animals. Our results support the suggestion that SF will be a suitable material for vascular regeneration in future.

Derya Aytemiz

2014-03-01

 
 
 
 
161

Antheraea pernyi silk fibroin for targeted gene delivery of VEGF165-Ang-1 with PEI.  

Science.gov (United States)

Vascularization is a crucial challenge in tissue engineering. One solution for this problem is to implant scaffolds that contain functional genes that promote vascularization by providing angiogenic growth factors via a gene delivery carrier. Poly(ethylenimine) (PEI) is a gene delivery carrier with high transfection efficiency but with cytotoxicity. To solve this problem, we utilized Antheraea pernyi silk fibroin (ASF), which has favorable cytocompatibility and biodegradability, RGD sequences and a negative charge, in conjunction with PEI, as the delivery vector for vascular endothelial growth factor (VEGF) 165-angiopoietin-1 (Ang-1) dual gene simultaneous expression plasmid, creating an ASF/PEI/pDNA complex. The results suggested that the zeta potential of the ASF/PEI/pDNA complex was significantly lower than that of the PEI/pDNA complex. Decreased nitrogen and increased oxygen on the surface of the complex demonstrated that the ASF had successfully combined with the surface of the PEI/pDNA. Furthermore, the complexes resisted digestion by nucleic acid enzymes and degradation by serum. L929 cells were cultured and transfected in vitro and improved cytotoxicity was found when the cells were transfected with ASF/PEI/pDNA compared with PEI/pDNA. In addition, the transfection efficiency and VEGF secretion increased. In general, this study provides a novel method for decreasing the cytotoxicity of PEI gene delivery vectors and increasing transfection efficiency of angiogenesis-related genes. PMID:24867887

Ma, Caili; Lv, Linlin; Liu, Yu; Yu, Yanni; You, Renchuan; Yang, Jicheng; Li, Mingzhong

2014-06-01

162

Effect of polyaspartic acid on hydroxyapatite deposition in silk fibroin blend films  

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Full Text Available Polyaspartic acid/silk fibroin/hydroxyapatite (PASP/SF-HA composites have been synthesized by biomimetic processing. SF solution was mixed with different contents of PASP to prepare the PASP/SF blend membranes. After ethanol treatment and premineralization process, the blend membranes were immersed into 1.5 simulated body fluid (1.5 SBF for 24 h to induce apatite deposition at 37±0.5°C. Fourier transform infrared spectroscopy (FTIR and X-ray diffraction (XRD results revealed that a conformation transition of SF occurred after the addition of PASP and ethanol treatment. The FTIR and XRD results also confirmed that the main component of apatite deposition was HA. Scanning electron microscopy (SEM showed that the content of HA increased with increasing PASP concentration .Inductively Coupled Plasma-Atomic Emission Spectrometry (ICP results revealed that the Ca/P molar ratio could reach 1.45, which was close to the Ca/P ratio of apatite. It was appropriate to conclude that the increasing content of PASP had a distinct effect on HA deposition in the blend films.

2010-05-01

163

Cartilage Tissue Engineering with Silk Fibroin Scaffolds Fabricated by Indirect Additive Manufacturing Technology  

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Full Text Available Advanced tissue engineering (TE technology based on additive manufacturing (AM can fabricate scaffolds with a three-dimensional (3D environment suitable for cartilage regeneration. Specifically, AM technology may allow the incorporation of complex architectural features. The present study involves the fabrication of 3D TE scaffolds by an indirect AM approach using silk fibroin (SF. From scanning electron microscopic observations, the presence of micro-pores and interconnected channels within the scaffold could be verified, resulting in a TE scaffold with both micro- and macro-structural features. The intrinsic properties, such as the chemical structure and thermal characteristics of SF, were preserved after the indirect AM manufacturing process. In vitro cell culture within the SF scaffold using porcine articular chondrocytes showed a steady increase in cell numbers up to Day 14. The specific production (per cell basis of the cartilage-specific extracellular matrix component (collagen Type II was enhanced with culture time up to 12 weeks, indicating the re-differentiation of chondrocytes within the scaffold. Subcutaneous implantation of the scaffold-chondrocyte constructs in nude mice also confirmed the formation of ectopic cartilage by histological examination and immunostaining.

Chih-Hao Chen

2014-03-01

164

Vitamin E-loaded silk fibroin nanofibrous mats fabricated by green process for skin care application.  

Science.gov (United States)

In the present study, we reported fabrication and skin benefit of a novel vitamin E (VE)-loaded silk fibroin (SF) nanofibrous mats. RRR-?-Tocopherol polyethylene glycol 1000 succinate (VE TPGS), a water-soluble derivative of VE, was incorporated into SF nanofiber successfully by aqua solution electrospinning for the first time. Morphology of the composite nanofibers changed with the different amount of VE TPGS: a ribbon-like shape for lower loading dose of VE TPGS, while a round shape for higher loading dose (more than 4% (wt/wt) based on the weight of SF). After treated with 75% (v/v) ethanol vapor, the composite nanofibrous mats showed an excellent water-resistant ability. In vitro study disclosed a sustained release behavior of VE TPGS disassociated from the nanofibrous mats. The mouse skin fibroblasts (L929 cells) cultured on the VE-loaded SF nanofibrous mats spread and proliferated much better than on cover slips. Moreover, the incorporation of VE TPGS was found strengthening the ability of SF nanofibrous mats on protecting the cells against oxidation stress induced by tert-butyl hydroperoxide. Our data presented impressive skin benefits of this VE-loaded SF nanofibrous mats, suggesting a promising applicative potential of this novel product on personal skin care, tissue regeneration and other related area. PMID:23396066

Sheng, Xiaoyue; Fan, Linpeng; He, Chuanglong; Zhang, Kuihua; Mo, Xiumei; Wang, Hongsheng

2013-05-01

165

Direct electrochemistry and electrocatalysis of heme-proteins in regenerated silk fibroin film  

International Nuclear Information System (INIS)

A biocompatible silk fibroin (SF) film provided a feasible microenvironment for heme-proteins to direct electron transfer on graphite electrodes (GE). Myoglobin (Mb), hemoglobin (Hb), horseradish peroxidase (HRP), and catalase (Cat) incorporated in SF films exhibited a pair of well-defined, nearly reversible cyclic voltammetric peaks, corresponding to the reaction of hemeFe (III) + e ? hemeFe (II). The formal potential (E 0), the apparent coverage (?) and the electron transfer rate constant (k s) of four proteins in SF films were evaluated by analyzing the cyclic voltammograms (CVs) of heme-proteins. The formal potential was pH dependent, suggesting that proton ion was involved in the reaction. Ultraviolet visible (UV-vis) spectra and reflectance absorbance infrared (RAIR) spectra indicated that heme-proteins in SF films were not grossly denatured. The structure of heme-proteins-SF films was investigated using scanning electron microscopy (SEM) and RAIR. It indicated that there existed intermolecular interaction between heme-proteins and SF and this governed their different morphology in SF films. Hydrogen peroxide and nitric oxide were catalytically reduced by the heme-proteins in SF films, showing the potential applicability of the heme-proteins-SF films as the new type of biosensors based on the protein film voltammetry

2006-02-03

166

Morphology, Secondary Structure and Thermal Properties of Silk Fibroin/Gelatin Blend Film  

Directory of Open Access Journals (Sweden)

Full Text Available This study aimed to prepare Silk Fibroin (SF and Gelatin (G blend film and study its morphology, secondary structure and thermal properties compared to native SF and G films. The films were prepared from the SF solution by casting on the polystyrene plates. They were investigated their secondary structure by fourier transform-infrared (FTIR spectroscopy, morphology using Scanning Electron Microscope (SEM. In addition, Thermogravimetric Analysis (TG and Differential TG (DTG were used for thermal properties investigation. The results found that the SF/G blend film composed of both ?-helix and ?-sheet structures which were similar characteristics of the native SF and G. This result was similar to the TG and DTG analysis according to blending between SF and G is not enhancing thermal stability of the film. However, changes in some absorption bands and temperatures were also observed from the blend film. The result suggested that chemical interaction and hydrogen bonding between SF and G could be formed. The formation could be affected to the uniform of the surface throughout the film under SEM.

Ong-chiari Watcharin

2009-01-01

167

Silk fibroin layer-by-layer microcapsules for localized gene delivery.  

Science.gov (United States)

Herein, we describe the delivery of plasmid DNA (pDNA) using silk fibroin (SF) layer-by-layer assembled microcapsules. Deposition of fluorescently labeled SF onto polystyrene (PS) template particles resulted in increasing fluorescence intensity and decreasing surface charge in correlation to SF layer number. After removal of the PS core, hollow, monodisperse, and structurally stable SF microcapsules of variable size and shell thickness were obtained. Plasmid DNA encoding for enhanced green fluorescent protein (eGFP) was loaded onto 1 or 4 ?m capsules, either by incorporation of pDNA within the innermost layer of the shell or by adsorption to the microcapsules surface, and in vitro pDNA release, cytotoxicty and eGFP expression were studied. Sustained pDNA release over 3 days was observed using both loading techniques, being accelerated in the presence of protease. DNA loaded SF microcapsules resulted in efficient cell transfection along with low cytotoxicity after 3 days incubation compared to treatment with pDNA/branched polyethylenimine complexes. Among the tested conditions highest transfection efficiencies were achieved using 1 ?m capsules where pDNA was adsorbed to the capsule surface. Our results suggest that SF microcapsules are suitable for the localized delivery of pDNA, combining low cytotoxicity and high transfection efficiency. PMID:24930849

Li, Linhao; Puhl, Sebastian; Meinel, Lorenz; Germershaus, Oliver

2014-09-01

168

Genipin-crosslinked silk fibroin/hydroxybutyl chitosan nanofibrous scaffolds for tissue-engineering application.  

Science.gov (United States)

To improve water-resistant ability and mechanical properties of silk fibroin (SF)/hydroxybutyl chitosan (HBC) nanofibrous scaffolds for tissue-engineering applications, genipin, glutaraldehyde (GTA), and ethanol were used to crosslink electrospun nanofibers, respectively. The mechanical properties of nanofibrous scaffolds were obviously improved after 24 h of crosslinking with genipin and were superior to those crosslinked with GTA and ethanol for 24 h. SEM indicated that crosslinked nanofibers with genipin and GTA vapor had good water-resistant ability. Characterization of the microstructure (porosity and pore structure) demonstrated crosslinked nanofibrous scaffolds with genipin and GTA vapor had lager porosities and mean diameters than those with ethanol. Characterization of FTIR-ATR and (13)C NMR clarified both genipin and GTA acted as crosslinking agents for SF and HBC. Furthermore, genipin could induce SF conformation from random coil or ?-helix to ?-sheet. Although GTA could also successfully crosslink SF/HBC nanofibrous scaffolds, in long run, genipin maybe a better method due to lower cytotoxicity than GTA. Cell viability studies and wound-healing test in rats clarified that the genipin-crosslinked SF/HBC nanofibrous scaffolds had a good biocompatibility both in vitro and in vivo. These results suggested that genipin-crosslinked SF/HBC nanofibrous scaffolds might be potential candidates for wound dressing and tissue-engineering scaffolds. PMID:20824649

Zhang, Kuihua; Qian, Yongfang; Wang, Hongsheng; Fan, Linpeng; Huang, Chen; Yin, Anlin; Mo, Xiumei

2010-12-01

169

Synthesis and characterization of dense membranes of silk fibroin with glycerin;Sintese e caracterizacao de membranas densas de fibroina de seda com glicerina  

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The addition of plasticizers seeks improvements in mechanical properties of dense membranes of silk fibroin with possible interactions by hydrogen bonds. The aim of the present study was to produce and characterize dense membranes of silk fibroin containing glycerin in two different concentrations. The characterization of the membranes was performed from scanning electron microscopy (SEM), mechanical traction tests, infrared spectroscopy (FTIR-ATR) and X-ray diffraction (XRD). The results indicated that the addition of glycerin allowed obtaining homogeneous and more crystalline membranes and improved their properties of elongation. (author)

Silva, Mariana F.; Moraes, Mariana A. de; Weska, Raquel F.; Nogueira, Grinia M.; Beppu, Marisa M., E-mail: beppu@feq.unicamp.b [Universidade Estadual de Campinas (FEQ/UNICAMP), SP (Brazil). Faculdade de Engenharia Quimica

2009-07-01

170

Effect of Strongly Alkaline Electrolyzed Water on Silk Degumming and the Physical Properties of the Fibroin Fiber  

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Strongly alkaline electrolyzed water (SAEW) was prepared by electrolysis of tap water in a laboratory-made water electrolyzer. The pH of stored SAEW was stable for more than one month. The hardness of the electrolyzed water was 30% lower and the Na+ concentration was 18% higher than those of the tap water. Silkworm cocoon shells were boiled in pH 11.50 SAEW at a ratio of 1?40?80 (W/V) for 20 min and the sericin layers around the silk fibroin fibers were removed completely. The tensile pro...

2013-01-01

171

Production and characterization of a silk-like hybrid protein, based on the polyalanine region of Samia cynthia ricini silk fibroin and a cell adhesive region derived from fibronectin.  

Science.gov (United States)

There are a variety of silkworms and silk fibroins produced by them. Silks have many inherent suitable properties for biomaterials. In this paper, a novel silk-like hybrid protein, [DGG(A)(12)GGAASTGRGDSPAAS](5), which consists of polyalanine region of silk fibroin from a wild silkworm, Samia cynthia ricini, and cell adhesive region including Arg-Gly-Asp (RGD) sequence, derived from fibronectin, was designed and produced. The genes encoding the hybrid protein were constructed and expressed in Escherichia coli. The main conformation of the polyalanine region, that is, either alpha-helix or beta-sheet, could be easily controlled by treatment with different acidic solvents, trifluoroacetic acid or formic acid, respectively. This structural change was monitored with 13C CP/MAS NMR. Higher cell adhesive and growth activities of the hybrid protein compared with those of collagen were obtained. PMID:14607499

Asakura, Tetsuo; Tanaka, Chikako; Yang, Mingying; Yao, Juming; Kurokawa, Masato

2004-02-01

172

Immobilization of acetylcholinesterase via biocompatible interface of silk fibroin for detection of organophosphate and carbamate pesticides  

Energy Technology Data Exchange (ETDEWEB)

An amperometric biosensor for the detection of organophosphate and carbamate pesticides was developed based on the immobilization of acetylcholinesterase (AChE) on regenerated silk fibroin (SF) matrix by non-covalent adsorption. SF and AChE were coated sequentially on the surface of the glassy carbon electrode (GCE) which was modified with multiwall carbon nanotube (MWNTs). The obtained biosensor was denoted as AChE-SF/MWNTs/GCE. The atomic force microscopy images showed that the SF matrix provided a more homogeneous interface for the AChE immobilization. The aggregation of immobilizing AChE was therefore avoided. The cyclic voltammogram of thiocholine at this biosensor exhibited a well defined oxidation peak at 0.667 V (vs. SCE). The inhibition rate of methyl parathion to the immobilized AChE was proportional to the logarithm of the concentration of methyl parathion over the range of the concentration of methyl parathion from 3.5 Multiplication-Sign 10{sup -6} to 2.0 Multiplication-Sign 10{sup -3} M with a detection limit of 5.0 Multiplication-Sign 10{sup -7} M. Similarly, the linearly response range of carbaryl was from 1.0 Multiplication-Sign 10{sup -7} to 3.0 Multiplication-Sign 10{sup -5} M with a detection limit of 6.0 Multiplication-Sign 10{sup -8} M. The experimental results indicate that AChE not only can be immobilized steadily on the SF matrix, but also the bioactivity of immobilizing AChE can be preserved effectively.

Xue Rui [College of Environmental and Energy Engineering, Beijing University of Technology, Beijing 100124 (China); Kang Tianfang, E-mail: kangtf@yahoo.cn [College of Environmental and Energy Engineering, Beijing University of Technology, Beijing 100124 (China); Lu Liping; Cheng Shuiyuan [College of Environmental and Energy Engineering, Beijing University of Technology, Beijing 100124 (China)

2012-06-01

173

In vitro biocompatibility evaluation of silk-fibroin/polyurethane membrane with cultivation of HUVECs  

Science.gov (United States)

In order to investigate the in vitro biocompatibility of a novel polyurethane (PU) membrane modified by incorporation of superfine silk-fibroin powder (SFP), which was prepared for small-diameter vascular grafts, with the cultivation of human umbilical vein endothelial cells (HUVECs), PU and SFP were mixed with the ratios of 9:1, 7:3, 5:5, 3:7 (PU:SFP) to make four composite materials. Unmodified PU and polytetrafluoroethylene (PTFE) were added as control groups. CCK-8 assay was used to evaluate the cytotoxicity of these biomaterials. Data were processed using SPSS, and P < 0.05 was considered to be statistically significant. Adherence and spreading of HUVECs on the surface of specimens was observed using direct contact cultivation. The toxicity ratings of the novel composites were grade 0-1, which is in the acceptable range. In all the experimental groups except control, SFP/PU with ratio of 1:9 had the least cytotoxicity property, and more content of SFP in the composite showed no improvement of the biocompatibility. HUVECs strongly attached to and grew on the surface of the biomaterials, and proliferated rapidly. The proliferation ability increased with increased proportion of SFP; however the cell quantity on the surface of the materials decreased when the proportion of SFP was equal to or larger than that of PU in the composite. It is concluded that this novel material has excellent cellular affinity with no cytotoxicity to HUVECs. Adding SFP gives PU better biocompatibility, while further research on optimum blend ratios is still needed.

Zhou, Mei; Wang, Wei-Ci; Liao, Yong-Gui; Liu, Wen-Qi; Yu, Miao; Ouyang, Chen-Xi

2014-03-01

174

Fabrication and characterization of silk fibroin/bioactive glass composite films  

International Nuclear Information System (INIS)

Composite films of silk fibroin (SF) with nano bioactive glass (NBG) were prepared by the solvent casting method, and the structures and properties of the composite films were characterized. Fourier transform infrared (FT-IR) spectroscopy analysis shows that the random coil and ?-sheet structure co-exist in the SF films. Results of field emission scanning electron microscope (FESEM) indicate that the NBG particles are uniformly dispersed in the SF films. The measurements of the water contact angles suggest that the incorporation of NBG into SF can improve the hydrophilicity of the composites. The bioactivity of the composite films was evaluated by soaking in 1.5 times simulated body fluid (1.5 × SBF), and formation of a hydroxycarbonate apatite (HCA) layer was determined by XRD and FESEM. The results show that the SF/NBG composite film is bioactive as it induces the formation of HCA on the surface of the composite film after soaking in 1.5 × SBF for 7 days. In vitro osteoblasts attachment and proliferation tests show that the composite film is a good matrix for the growth of osteoblasts. Consequently, the incorporation of NBG into the SF film can enhance both the bioactivity and biocompatibility of the film, which suggests that the SF/NBG composite film may be a potential biomaterial for bone tissue engineering. - Highlights: ? The incorporation of NBG into SF can improve the hydrophilicity of the SF/NBG composite films. ? The SF/NBG composite films show the better bioactivity than the pure SF film. ? The SF/NBG composite films facilitate cell growth and promote cell proliferation and differentiation.

2012-05-01

175

Effect of initial cell seeding density on 3D-engineered silk fibroin scaffolds for articular cartilage tissue engineering.  

Science.gov (United States)

The repair of articular cartilage defects poses a continuing challenge. Cartilage tissue engineering through the culture of chondrocytes seeded in 3D porous scaffolds has the potential for generating constructs that repair successfully. It also provides a platform to study scaffold-cell and cell-cell interactions. The scaffold affects the growth and morphology of cells growing on it, and concomitantly, cells affect the properties of the resultant tissue construct. Silk fibroin protein from Antheraea mylitta, a non-mulberry Indian tropical tasar silkworm, is a potential biomaterial for diverse applications due to its widespread versatility as a mechanically robust, biocompatible, tissue engineering material. Analysis of silk fibroin scaffolds seeded with varying initial densities (25, 50 and 100 million cells/ml) and cultured for 2 weeks showed that thickness and wet weight increased by 60-70% for the highest cell density, and DNA, GAG and collagen content of the cartilaginous constructs increased with increasing cell density. Mechanical characterization of the constructs elucidated that the highest density constructs had compressive stiffness and modulus 4-5 times that of cell-free scaffolds. The present results indicate the importance of cell seeding density in the rapid formation of a functional cartilaginous tissue. PMID:21906805

Talukdar, Sarmistha; Nguyen, Quynhhoa T; Chen, Albert C; Sah, Robert L; Kundu, Subhas C

2011-12-01

176

3D silk fibroin scaffold incorporating titanium dioxide (TiO2) nanoparticle (NPs) for tissue engineering.  

Science.gov (United States)

The present study deals with fabrication of scaffolds composing of silk fibroin and TiO2 NPs fabricated using a salt-leaching process. At first instance, the TiO2 NPs were prepared by using sol-gel synthesis, affording to have average diameter of 77±21?m. Furthermore, the aqueous solutions of silk fibroin were mixed with 0.2%, 2.0% and 4.0% of TiO2 NPs and salt-leaching process was introduced which resulted in creation of porous scaffolds modified with TiO2 NPs. The presence of TiO2 NPs in scaffolds was confirmed by VP-FE-SEM-EDS, TGA and XRD. The presence of TiO2 NPs influenced in decrease in pore size and swelling behavior of composite scaffolds. The resultant mechanical property of scaffolds was improved upon the introduction of TiO2 NPs. Moreover, cell cytotoxicity results for 1, 3 and 7 days; revealed no toxic behavior to osteoblasts. However, a mild toxicity to NIH 3T3 fibroblasts was observed with the scaffolds containing 4.0% TiO2 NPs. The cell fixation results from 1 and 7 days of incubation indicated the attachment, spreading and subsequent proliferation of fibroblasts. However, these findings were independent to the amount of TiO2 NPs in scaffolds. PMID:24794196

Kim, Jung-Ho; Sheikh, Faheem A; Ju, Hyung Woo; Park, Hyun Jung; Moon, Bo Mi; Lee, Ok Joo; Park, Chan Hum

2014-07-01

177

Silk Fibroin/Nano-CaP Bilayered scaffolds for osteochondral tissue engineering  

Digital Repository Infrastructure Vision for European Research (DRIVER)

In this study, bilayered silk and silk/nano-CaP scaffolds were developed for osteochondral (OC) tissue engineering. Aqueous silk solution (16 wt.%) was used for preparation of the cartilage-like layer and, for generation of the silk/nano-CaP suspension and the bottom layer (CaP/Silk: 16 wt.%). The scaffolds were formed by using salt-leaching/lyophilization approach. The scanning electron microscopy revealed that the both layers presented porous structure and integrated well. Mi...

Yan, Leping; Oliveira, J. M.; Oliveira, A. L.; Reis, R. L.

2014-01-01

178

Incorporation of Exogenous RGD Peptide and Inter-Species Blending as Strategies for Enhancing Human Corneal Limbal Epithelial Cell Growth on Bombyx mori Silk Fibroin Membranes  

Directory of Open Access Journals (Sweden)

Full Text Available While fibroin isolated from the cocoons of domesticated silkworm Bombyx mori supports growth of human corneal limbal epithelial (HLE cells, the mechanism of cell attachment remains unclear. In the present study we sought to enhance the attachment of HLE cells to membranes of Bombyx mori silk fibroin (BMSF through surface functionalization with an arginine-glycine-aspartic acid (RGD-containing peptide. Moreover, we have examined the response of HLE cells to BMSF when blended with the fibroin produced by a wild silkworm, Antheraea pernyi, which is known to contain RGD sequences within its primary structure. A procedure to isolate A. pernyi silk fibroin (APSF from the cocoons was established, and blends of the two fibroins were prepared at five different BMSF/APSF ratios. In another experiment, BMSF surface was modified by binding chemically the GRGDSPC peptide using a water-soluble carbodiimide. Primary HLE were grown in the absence of serum on membranes made of BMSF, APSF, and their blends, as well as on RGD-modified BMSF. There was no statistically significant enhancing effect on the cell attachment due to the RGD presence. This suggests that the adhesion through RGD ligands may have a complex mechanism, and the investigated strategies are of limited value unless the factors contributing to this mechanism become better known.

Traian V. Chirila

2013-05-01

179

Preparation and characterization of biomimetic silk fibroin/chitosan composite nanofibers by electrospinning for osteoblasts culture  

Science.gov (United States)

In this study, we have successfully fabricated electrospun bead-free silk fibroin [SF]/chitosan [CS] composite nanofibers [NFs] covering the whole range of CS content (0%, 25%, 50%, 75%, and 100%). SF/CS spinning solutions were prepared in a mixed solvent system of trifluoroacetic acid [TFA] and dichloromethane. The morphology of the NFs was observed by scanning electron microscope, and the average fiber diameter ranges from 215 to 478 nm. Confocal laser scanning microscopy confirms the uniform distribution of SF and CS within the composite NFs. To increase biocompatibility and preserve nanostructure when seeded with cells in culture medium, NFs were treated with an ethanol/ammonia aqueous solution to remove residual TFA and to change SF protein conformation. After the chemical treatment, SF/CS NFs could maintain the original structure for up to 54 days in culture medium. Properties of pristine and chemically treated SF/CS NFs were investigated by Fourier transform infrared spectroscopy [FT-IR], X-ray diffraction [XRD], and thermogravimetry/differential scanning calorimetry [TG/DSC]. Shift of absorption peaks in FT-IR spectra confirms the conformation change of SF from random coil to ?-sheet by the action of ethanol, which is also consistent with the SF crystalline diffraction patterns measured by XRD. From TG/DSC analysis, the decomposition temperature peaks due to salt formation from TFA and protonated amines disappeared after chemical treatment, indicating complete removal of TFA by binding with ammonium ions during the treatment. This was also confirmed with the disappearance of F1s peak in X-ray photoelectron spectroscopy spectra and disappearance of TFA salt peaks in FT-IR spectra. The composite NFs could support the growth and osteogenic differentiation of human fetal osteoblastic [hFOB] cells, but each component in the composite NF shows distinct effect on cell behavior. SF promotes hFOB proliferation while CS enhances hFOB differentiation. The composite SF/CS NFs will be suitable for bone tissue engineering applications by choosing a suitable blend composition. PACS: 87.85.jf; 87.85.Rs; 68.37.Hk.

Chen, Jyh-Ping; Chen, Shih-Hsien; Lai, Guo-Jyun

2012-03-01

180

Electrospun composites of PHBV, silk fibroin and nano-hydroxyapatite for bone tissue engineering.  

Science.gov (United States)

Electrospinning of fibrous scaffolds containing nano-hydroxyapatite (nHAp) embedded in a matrix of functional biomacromolecules offers an attractive route to mimicking the natural bone tissue architecture. Functional fibrous substrates will support cell attachment, proliferation and differentiation, while the role of HAp is to induce cells to secrete extracellular matrix (ECM) for mineralization to form bone. Electrospinning of biomaterials composed of polyhydroxybutyrate-co-(3-hydroxyvalerate) with 2% valerate fraction (PHBV), nano-hydroxyapatite (nHAp), and Bombyx mori silk fibroin essence (SF), Mw=90KDa, has been achieved for nHAp and SF solution concentrations of 2 (w/vol) % each and 5 (w/vol) % each. The structure and properties of the nanocomposite fibrous membranes were investigated by means of Scanning Electron Microscopy in combination with Energy Dispersive X-Ray Analysis (SEM/EDX), Fourier Transformed Infrared Spectroscopy (FT-IR), uniaxial tensile and compressive mechanical testing, degradation tests and in vitro bioactivity tests. SEM images showed smooth, uniform and continuous fibre deposition with no bead formation, and fibre diameters of between 10 and 15 ?m. EDX and FT-IR confirmed the presence of nHAp and SF. After one month in deionised water, tests showed less than 2% weight loss with the samples retaining their fibrous morphology, confirming that this material biodegrades slowly. After 28 days of immersion in Simulated Body Fluid (SBF) an apatite layer was visible on the surface of the fibres, proving their bioactivity. Preliminary in vitro biological assessment showed that after 1 and 3 days in culture, cells were attached to the fibres, retaining their morphology while presenting a flattened appearance and elongated shape on the surface of fibres. Young's modulus was found to increase from 0.7 kPa (±0.33 kPa) for electrospun samples of PHBV only to 1.4 kPa (±0.54 kPa) for samples with 2 (w/vol) % each of nHAp and SF. Samples prepared with 5 (w/vol) % each of nHAp and SF did not show a similar improvement. PMID:24094204

Pa?cu, Elena I; Stokes, Joseph; McGuinness, Garrett B

2013-12-01

 
 
 
 
181

The nucleotide sequence adjacent to poly(A) in silk fibroin messenger RNA.  

Science.gov (United States)

The 3'-terminal nucleotide sequence of fibroin mRNA has been determined. A cDNA transcript primed by oligo(dT) hybridized to the poly(A) segment of fibroin mRNA was used to determine the sequence immediately adjacent to poly(A). A specific primer complementary to this sequence then allowed synthesis of 5'-terminally homogeneous transcripts amenable to analysis by rapid sequencing methods. The 3' terminus of fibroin mRNA contains an unusually long oligo(U) stretch, as well as the sequence A2UA3 recently found in 3' non-coding regions of other eukaryotic mRNAs. It lacks the oligonucleotides characteristic of the repetitive coding region of fibroin mRNA. PMID:457673

Hagenbüchle, O; Krikeles, M S; Sprague, K U

1979-08-10

182

Electrospun liquid silk from the gland of Bombyx mori silk/ Green Fluorescent Proteins (GFP)/ poly(ethylene oxide)  

Science.gov (United States)

We have previously demonstrated that Green Fluorescent Protein (GFP) molecules can be incorporated into silk films and maintain their nonlinear optical properties as well as resist to damage at fluence of 0.1-0.2 J/cm^2 [1]. In the present study we report the incorporation of GFP into electrsopun fibers of liquid silk from the gland of B. mori. PEO was added to the silk/GFP solution to improve the processability. The silk/GFP/PEO solutions were successfully elctrospun and the morphology of fibers was characterized using optical microscopy, and scanning electron microscopy. The resulting fibers exhibit fluorescent under the UV microscope in the reflection mode (epifluorescence) indicating incorporation of the GFP. The fiber diameters are less than 500 nm. Other characterization techniques are being applied. [1]. S. Putthanarat, et.al., ``Nonlinear Optical Transmission of Silk/Green Fluorescent Protein (GFP) Films,'' Polymer 2004;45:8451.

Putthanarat, Sirina; Jones, Sharon; Farmer, Barry

2005-03-01

183

Composite Scaffolds Containing Silk Fibroin, Gelatin, and Hydroxyapatite for Bone Tissue Regeneration and 3D Cell Culturing.  

Science.gov (United States)

Three-dimensional (3D) silk fibroin scaffolds were modified with one of the major bone tissue derivatives (nano-hydroxyapatite) and/or a collagen derivative (gelatin). Adhesion and proliferation of mouse embryonic fibroblasts (MEF) within the scaffold were increased after modification with either nano-hydroxyapatite or gelatin. However, a significant increase in MEF adhesion and proliferation was observed when both additives were introduced into the scaffold. Such modified composite scaffolds provide a new and better platform to study wound healing, bone and other tissue regeneration, as well as artificial organ bioengineering. This system can further be applied to establish experimental models to study cell-substrate interactions, cell migration and other complex processes, which may be difficult to address using the conventional two-dimensional culture systems. PMID:24772332

Moisenovich, M M; Arkhipova, A Yu; Orlova, A A; Drutskaya, M S; Volkova, S V; Zacharov, S E; Agapov, I I; Kirpichnikov, M P

2014-01-01

184

Three-Dimensional Porous Network Structure Developed in Hydroxyapatite-Based Nanocomposites Containing Enzyme Pretreated Silk Fibroin  

International Nuclear Information System (INIS)

Chemically modified silk fibroin (SF) with an enzyme, Proteinase K, has been incorporated into hydroxyapatite (HAp)-based nanocomposite attempting to strengthen the interfacial bonding between the mineral phase and the organic matrix. Particular emphasis is laid on the microstructure and microhardness of the composite along with the crystallographic properties of HAp. The whisker-like HAp crystallites of nanometer size show the preferential self-assembly and anisotropic crystal growth along c-axis. There appears porous microstructure with 70% of open porosity and pore size distribution of 10-115 um in the composite. Attributed to the enzyme modification, the crosslinkage between HAp clusters and SF matrix is improved to form an enhanced three-dimensional network extending throughout the composites and an increase of 35% in microhardness of the composite is achieved as well

2004-02-01

185

Composite chitosan/silk fibroin nanofibers for modulation of osteogenic differentiation and proliferation of human mesenchymal stem cells.  

Science.gov (United States)

Nanofibrous membrane scaffolds of chitosan (CS), silk fibroin (SF) and CS/SF blend were prepared by electrospinning and studied for growth and osteogenic differentiation of human bone marrow mesenchymal stem cells (hMSCs). The morphology and physico-chemical properties of all membrane scaffolds were compared. The influence of CS and SF on cell proliferation was assessed by the MTS assay, whereas osteogenic differentiation was determined from the Alizarin Red staining, alkaline phosphatase activity and expression of osteogenic marker genes. The osteogenic differentiation and proliferation of hMSCs were enhanced by CS and SF nanofibers, respectively. Blending CS with SF retained the osteogenesis nature of CS without negatively influencing the cell proliferative effect of SF. By taking advantage of the differentiation/proliferation cues from individual components, the electrospun CS/SF composite nanofibrous membrane scaffold is suitable for bone tissue engineering. PMID:25037354

Lai, Guo-Jyun; Shalumon, K T; Chen, Shih-Hsien; Chen, Jyh-Ping

2014-10-13

186

Composite biocompatible hydroxyapatite-silk fibroin coatings for medical implants obtained by Matrix Assisted Pulsed Laser Evaporation  

Energy Technology Data Exchange (ETDEWEB)

The aim of this study was to obtain biomimetic inorganic-organic thin films as coatings for metallic medical implants. These contain hydroxyapatite, the inorganic component of the bony tissues, and a natural biopolymer - silk fibroin - added in view to induce the surface functionalization. Hydroxyapatite (HA), silk fibroin (FIB) and composite HA-FIB films were obtained by Matrix Assisted Pulsed Laser Evaporation (MAPLE) in order to compare their physical and biological performances as coatings on metallic prostheses. We used an excimer laser source (KrF*, {lambda} = 248 nm, {tau} = 25 ns) operated at 10 Hz repetition rate. Coatings were deposited on quartz, Si and Ti substrates and then subjected to physical (FTIR, XRD, AFM, SEM) analyses, correlated with the results of the cytocompatibility in vitro tests. The hybrid films were synthesized from frozen targets of aqueous suspensions with 3:2 or 3:4 weight ratio of HA:FIB. An appropriate stoichiometric and functional transfer was obtained for 0.4-0.5 J/cm{sup 2} laser fluence. FTIR spectra of FIB and HA-FIB films exhibited distinctive absorption maxima, in specific positions of FIB random coil form: 1540 cm{sup -1} amide II, 1654 cm{sup -1} amide I, 1243 cm{sup -1} amide III, while the peak from 1027 cm{sup -1} appeared only for HA and composite films. Osteosarcoma SaOs2 cells cultured 72 h on FIB and HA-FIB films showed increased viability, good spreading and normal cell morphology. The well-elongated, flattened cells are a sign of an appropriate interaction with the MAPLE FIB and composite HA-FIB coatings.

Miroiu, F.M., E-mail: marimona.miroiu@inflpr.ro [National Institute for Lasers, Plasma, and Radiation Physics, 409 Atomistilor Street, RO-77125, MG-36, Magurele-Ilfov (Romania); Socol, G.; Visan, A.; Stefan, N.; Craciun, D.; Craciun, V.; Dorcioman, G.; Mihailescu, I.N. [National Institute for Lasers, Plasma, and Radiation Physics, 409 Atomistilor Street, RO-77125, MG-36, Magurele-Ilfov (Romania); Sima, L.E.; Petrescu, S.M. [Institute of Biochemistry, Romanian Academy, 296 Splaiul Independentei, 060031 Bucharest (Romania); Andronie, A.; Stamatin, I. [3Nano-SAE Alternative Energy Sources-University of Bucharest, Faculty of Physics, 409 Atomistilor Street, RO-77125, Magurele-Ilfov (Romania); Moga, S.; Ducu, C. [University of Pitesti, Targul din Vale Str, no. 1, 110040 Pitesti (Romania)

2010-05-25

187

Nanofibrous architecture of silk fibroin scaffolds prepared with a mild self-assembly process  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Besides excellent biocompatibility and biodegradability, a useful tissue engineering scaffold should provide suitable macropores and nanofibrous structure, similar to extracellular matrix (ECM), to induce desired cellular activities and to guide tissue regeneration. In the present study, a mild process to prepare porous and nanofibrous silk based scaffolds from aqueous solution is described. Using collagen to control the self-assembly of silk, nanofibrous silk scaffolds were firstly achieved ...

Lu, Qiang; Wang, Xiuli; Lu, Shenzhou; Li, Mingzhong; Kaplan, David L.; Zhu, Hesun

2011-01-01

188

Regenerated silk fibroin scaffold and infrapatellar adipose stromal vascular fraction as feeder-layer: a new product for cartilage advanced therapy.  

Science.gov (United States)

Articular cartilage has limited repair and regeneration potential, and the scarcity of treatment modalities has motivated attempts to engineer cartilage tissue constructs. The use of chondrocytes in cartilage tissue engineering has been restricted by the limited availability of these cells, their intrinsic tendency to lose their phenotype during the expansion, as well as the difficulties during the first cell adhesion to the scaffold. Aim of this work was to evaluate the intra-articular adipose stromal vascular fraction attachment on silk fibroin scaffold to promote chondrocytes adhesion and proliferation. Physicochemical characterization has demonstrated that three-dimensionally organized silk fibroin scaffold is an ideal biopolymer for cartilage tissue engineering; it allows cell attachment, scaffold colonization, and physically cell holding in the area that must be repaired; the use of adipose-derived stem cells is a promising strategy to promote adhesion and proliferation of chondrocytes to the scaffold as an autologous human feeder layer. PMID:21338265

Chlapanidas, Theodora; Faragò, Silvio; Mingotto, Federica; Crovato, Francesca; Tosca, Marta Cecilia; Antonioli, Barbara; Bucco, Massimo; Lucconi, Giulia; Scalise, Alessandro; Vigo, Daniele; Faustini, Massimo; Marazzi, Mario; Torre, Maria Luisa

2011-07-01

189

Isolation of a Clone Encoding a Second Dragline Silk Fibroin. (Reannouncement with New Availability Information).  

Science.gov (United States)

Spider dragline silk is a unique protein fiber possessing both high tensile strength and high elasticity. A partial cDNA clone for one dragline silk protein (Spidroin 1) was previously isolated. However, the predicted amino acid sequence could not account...

M. B. Hinnman R. V. Lewis

1992-01-01

190

The rapid anastomosis between prevascularized networks on silk fibroin scaffolds generated in vitro with cocultures of human microvascular endothelial and osteoblast cells and the host vasculature.  

Science.gov (United States)

The survival and functioning of a bone biomaterial upon implantation requires a rapidly forming and stably functioning vascularization that connects the implant to the recipient. We have previously shown that human microcapillary endothelial cells (HDMEC) and primary human osteoblast cells (HOS) in coculture on various 3-D bone biomaterial scaffolds rapidly distribute and self-assemble into a morphological structure resembling bone tissue. Endothelial cells form microcapillary-like structures containing a lumen and these were intertwined between the osteoblast cells and the biomaterial. This tissue-like self-assembly occurred in the absence of exogenously added angiogenic stimuli or artificial matrices. The purpose of this study was to determine whether this in vitro pre-formed microvasculature persists and functions in vivo and to determine how the host responds to the cell-containing scaffolds. The scaffolds with cocultures were implanted into immune-deficient mice and compared to scaffolds without cells or with HDMEC alone. Histological evaluation and immunohistochemical staining with human-specific antibodies of materials removed 14 days after implantation demonstrated that the in vitro pre-formed microcapillary structures were present on the silk fibroin scaffolds and showed a perfused lumen that contained red blood cells. This proved anastomosis with the host vasculature. Chimeric vessels in which HDMEC were integrated with the host's ingrowing (murine) capillaries were also observed. No HDMEC-derived microvessel structures or chimeric vessels were observed on implanted silk fibroin when precultured with HDMEC alone. In addition, there was migration of the host (murine) vasculature into the silk fibroin scaffolds implanted with cocultures, whereas silk fibroin alone or silk fibroin precultured only with HDMEC were nearly devoid of ingrowing host microcapillaries. Therefore, not only do the in vitro pre-formed microcapillaries in a coculture survive and anastomose with the host vasculature to become functioning microcapillaries after implantation, the coculture also stimulates the host capillaries to rapidly grow into the scaffold to vascularize the implanted material. Thus, this coculture-based pre-vascularization of a biomaterial implant may have great potential in the clinical setting to treat large bone defects. PMID:20619788

Unger, Ronald E; Ghanaati, Shahram; Orth, Carina; Sartoris, Anne; Barbeck, Mike; Halstenberg, Sven; Motta, Antonella; Migliaresi, Claudio; Kirkpatrick, C James

2010-09-01

191

The design of silk fiber composition in moths has been conserved for more than 150 million years.  

Science.gov (United States)

The silk of caterpillars is secreted in the labial glands, stored as a gel in their lumen, and converted into a solid filament during spinning. Heavy chain fibroin (H-fibroin), light chain fibroin (L-fibroin), and P25 protein constitute the filament core in a few species that have been analyzed. Identification of these proteins in Yponomeuta evonymella, a moth from a family which diverged from the rest of Lepidoptera about 150 million years ago, reveals that the mode of filament construction is highly conserved. It is proposed that association of the three proteins is suited for long storage of hydrated silk dope and its rapid conversion to filament. Interactions underlying these processes depend on conserved spacing of critical amino acid residues that are dispersed through the L-fibroin and P25 and assembled in the short ends of the H-fibroin molecule. Strength, elasticity, and other physical properties of the filament are determined by simple amino acid motifs arranged in repetitive modules that build up most of the H-fibroin. H-Fibroin synergy with L-fibroin and P25 does not interfere with motif diversification by which the filament acquires new properties. Several types of motifs in complex repeats occur in the silks used for larval cobwebs and pupal cocoons. Restriction of silk use to cocoon construction in some lepidopteran families has been accompanied by simplification of H-fibroin repeats. An extreme deviation of the silk structure occurs in the Saturniidae silkmoths, which possess modified H-fibroin and lack L-fibroin and P25. PMID:16755355

Yonemura, Naoyuki; Sehnal, Frantisek

2006-07-01

192

Wound healing effect of silk fibroin/alginate-blended sponge in full thickness skin defect of rat.  

Science.gov (United States)

Silk fibroin (SF) and alginate (AA) have been proved to be invaluable natural materials in the field of biomedical engineering. This study was designed to compare the wound healing effect of SF, AA and SF/AA-blended sponge (SF/AA) with clinically used Nu Gauze(TM) (CONT) in a rat full thickness wound model. Two circular skin wounds on the back of rat were covered with either of CONT, SF, AA or SF/AA. On the postoperative days of 3, 7, 10 and 14, residual wound area was calculated, and skin wound tissues were biopsied to measure the area of regenerated epithelium and collagen deposition as well as the number of proliferating cell nuclear antigen (PCNA)-immunoreactive cells. Half healing time (HT(50)) of SF/AA was dramatically reduced as compared with that of SF, AA or CONT. Furthermore, SF/AA significantly increased the size of re-epithelialization and the number of PCNA positive cells, whereas the effect of SF/AA on collagen deposition was not significantly different as compared with that of SF or AA. These results demonstrate that the wound healing effect of SF/AA is the best among other treatments including SF and AA, and this synergic effect is mediated by re-epithelialization via rapid proliferation of epithelial cell. PMID:16691353

Roh, Dae-Hyun; Kang, Seuk-Yun; Kim, Jeom-Yong; Kwon, Young-Bae; Young Kweon, Hae; Lee, Kwang-Gill; Park, Young-Hwan; Baek, Rong-Min; Heo, Chan-Yeong; Choe, Joon; Lee, Jang-Hern

2006-06-01

193

Therapeutic efficacy of antibiotic-loaded gelatin microsphere/silk fibroin scaffolds in infected full-thickness burns.  

Science.gov (United States)

Despite advances in burn treatment, burn infection remains a major cause of morbidity and mortality. In this study, an antibacterial silk fibroin (SF) scaffold for burn treatment was designed; gelatin microspheres (GMs) were impregnated with the antibiotic gentamycin sulfate (GS), and the GS-impregnated GMs were then embedded in a SF matrix to fabricate GS/GM/SF scaffolds. The developed GS/GM/SF scaffolds could serve as a dermal regeneration template in full-thickness burns. The average pore size and porosity of the GS/GM/SF scaffolds were 40-80?m and 85%, respectively. Furthermore, the drug release rate of the scaffolds was significantly slower than that of either GS/GM or GS/SF scaffolds. And the composite scaffold exhibited stronger antimicrobial activities against Escherichia coli, Staphylococcus aureus and Pseudomonas aeruginosa. Hence, we evaluated the wound-healing effects and antibacterial properties of the GS/GM/SF scaffolds in a rat full-thickness burn infection model. Over 21days, the GS/GM/SF scaffolds not only significantly reduced burn infection by P. aeruginosa but also accelerated the regeneration of the dermis and exhibited higher epithelialization rates than did GS/SF and SF scaffolds. Thus, GS/GM/SF scaffolds are potentially effective for treatment of full-thickness infected burns, and GS/GM/SF scaffolds are a promising therapeutic tool for severely burned patients. PMID:24704698

Lan, Yong; Li, Weichang; Jiao, Yanpeng; Guo, Rui; Zhang, Yi; Xue, Wei; Zhang, Yuanming

2014-07-01

194

Mass-spectrometric determination of the amino acid sequences in peptides isolated from protein silk fibroin of Bombyx mori  

Science.gov (United States)

Several peptides were isolated from the protein silk fibroin of Bombyx mori by means of ion-exchange chromatography of a chymotryptic digest. The sequences of three of the peptides, Gly-Ala-Gly-Tyr, Gly-Val-Gly-Tyr and Gly-Ala-Gly-Ala-Gly-Ala-Gly-Tyr, were known from previous chemical work, but the sequence of the fourth, Gly-Ala-Gly-Val-Gly-Ala-Gly-Tyr, was previously only partially known. The necessary volatility for mass-spectrometric examination of the peptides was achieved by permethylation of the N-acetyl-peptide methyl ester derivatives. From the mass spectra it was possible to confirm the known sequences and to establish that of the partially known one. In one instance it was possible to deduce from the same mass spectrum the sequence of a main peptide component and that of a small amount of contaminating peptide. These results demonstrate for the first time the use of mass spectrometry in the determination of the amino acid sequences in peptides from a protein hydrolysate.

Geddes, A. J.; Graham, G. N.; Morris, H. R.; Lucas, F.; Barber, M.; Wolstenholme, W. A.

1969-01-01

195

Development of micro-structural units in the silk fibroin thin films prepared by near-infrared pulsed laser deposition  

International Nuclear Information System (INIS)

Development of micro-structural units in a silk fibroin (SF) thin film deposited on Si (100) by using 1064 nm pulsed laser beam was examined. The smallest units ranging ca 10-20 nm were always observed in more or less uniformly dispersed states, which we coined as smallest protein units. At the same time, we frequently observed much larger chunks. Formation of such chunks was suppressed by choosing smaller fluence, shorter deposition time or by sputtering intermittently. When the laser fluence does not appreciably exceed the empirically determined threshold of 1.7 J/cm2, the basic secondary structure of SF, i.e. anti-parallel ?-sheet, was well preserved, as confirmed by Fourier transform infrared spectroscopy. Since the target material exhibits only very small extent of absorption of 1064 nm beam, we suspected that formation of radicals and free electrons and subsequent optical breakdown are mainly responsible for the deposition process, although a number of mechanistic questions remain open. The present near-infrared pulsed laser deposition technique seems attractive since it is free from appreciable damage of the protein secondary structure and solubility of the protein species, and a compact, relatively inexpensive laser source suffices

2006-12-05

196

The anticoagulant ability of ferulic acid and its applications for improving the blood compatibility of silk fibroin  

International Nuclear Information System (INIS)

The hemocompatibility of silk fibroin (SF) was improved with ferulic acid (FA) by graft polymerization. Ferulic acid is an active ingredient of many Chinese herbal medicines, such as Chuanxiong (Rhizoma ligustici wallichii), Danggui (Angelica sinensis) and Awei (Asafoetida giantfennel), which have been used to treat cardiovascular diseases by Chinese physicians for thousands of years. The inhibitory functions of FA on blood coagulation and erythrocyte agglutination were first characterized by a Lee-White test tube method and a micropipette technique, respectively. Then, FA was immobilized on SF by graft polymerization and the surface composition of modified SF was characterized by attenuated total reflectance Fourier-transform infrared (ATR-FTIR), x-ray photoelectron spectroscopy (XPS) and optical microscopy. The anticoagulant activity of modified SF was assessed, respectively, by in vitro clotting time measurements on a photo-optical clot detection instrument and with the Lee-White test tube method. The test results indicated that in comparison to untreated SF, the anticoagulant activity of modified SF has been improved significantly. Moreover, the SF surface composition is altered by FA but its ?-sheet conformation is not disturbed.

2008-12-01

197

Cytotoxicity and endothelial cell adhesion of lyophilized and irradiated bovine pericardium modified with silk fibroin and chitosan.  

Science.gov (United States)

Grafts of biological tissues have been used since the 1960s as an alternative to the mechanical heart prostheses. Nowadays, the most consolidated treatment to bovine pericardial (BP) bioprostheses is the crosslinking with glutaraldehyde (GA), although GA may induce calcification in vivo. In previous work, our group demonstrated that electron beam irradiation applied to lyophilized BP in the absence of oxygen promoted crosslinks among collagen fibers of BP tissue. In this work, the incorporation of silk fibroin (SF) and chitosan (CHIT) in the BP not treated with GA was studied. The samples were irradiated and then analyzed for their cytotoxicity and the ability of adhesion and growth of endothelial cells. Initially, all samples showed cytotoxicity. However, after a few washing cycles, the cytotoxicity due to acetic acid and ethanol residues was removed from the biomaterial making it suitable for the biofunctional test. The samples modified with SF/CHIT and electron beam irradiated favored the adhesion and growth of endothelial cells throughout the tissue. PMID:21595719

Rodas, Andrea C D; Polak, Roberta; Hara, Priscila H; Lee, Emily I; Pitombo, Ronaldo N M; Higa, Olga Z

2011-05-01

198

The anticoagulant ability of ferulic acid and its applications for improving the blood compatibility of silk fibroin  

Energy Technology Data Exchange (ETDEWEB)

The hemocompatibility of silk fibroin (SF) was improved with ferulic acid (FA) by graft polymerization. Ferulic acid is an active ingredient of many Chinese herbal medicines, such as Chuanxiong (Rhizoma ligustici wallichii), Danggui (Angelica sinensis) and Awei (Asafoetida giantfennel), which have been used to treat cardiovascular diseases by Chinese physicians for thousands of years. The inhibitory functions of FA on blood coagulation and erythrocyte agglutination were first characterized by a Lee-White test tube method and a micropipette technique, respectively. Then, FA was immobilized on SF by graft polymerization and the surface composition of modified SF was characterized by attenuated total reflectance Fourier-transform infrared (ATR-FTIR), x-ray photoelectron spectroscopy (XPS) and optical microscopy. The anticoagulant activity of modified SF was assessed, respectively, by in vitro clotting time measurements on a photo-optical clot detection instrument and with the Lee-White test tube method. The test results indicated that in comparison to untreated SF, the anticoagulant activity of modified SF has been improved significantly. Moreover, the SF surface composition is altered by FA but its {beta}-sheet conformation is not disturbed.

Wang Song; Gao Zhen; Chen Xiaomeng; Lian Xiaojie; Zhu Hesun [School of Material Science and Engineering, Beijing Institute of Technology, Beijing 100081 (China); Zheng Jun; Sun Lizhong [Department of Cardiac Surgery, Cardiovascular Institute and Fu Wai Hospital, CAMS and PUMC, Beijing 100037 (China)], E-mail: wangsongbit@hotmail.com

2008-12-15

199

Transgenic protein production in silkworm silk glands requires cathepsin and chitinase of Autographa californica multicapsid nucleopolyhedrovirus.  

Science.gov (United States)

The silkworm Bombyx mori represents an established in vivo system for the production of recombinant proteins. Baculoviruses have been extensively investigated and optimised for the expression of high protein levels inside the haemolymph of larvae and pupae of this lepidopteran insect. Current technology includes deletion of genes responsible for the activity of virus-borne proteases, which in wild-type viruses, cause liquefaction of the host insect and enhance horizontal transmission of newly synthesised virus particles. Besides the haemolymph, the silk gland of B. mori provides an additional expression system for recombinant proteins. In this paper, we investigated how silk gland can be efficiently infected by a Autographa californica multicapsid nuclear polyhedrosis virus (AcMNPV). We demonstrated that the viral chitinase and the cysteine protease cathepsin are necessary to permit viral entry into the silk gland cells of intrahaemocoelically infected B. mori larvae. Moreover, for the first time, we showed AcMNPV crossing the basal lamina of silk glands in B. mori larvae, and we assessed a new path of infection of silk gland cells that can be exploited for protein production. PMID:24477386

Wöltje, Michael; Böbel, Melanie; Rheinnecker, Michael; Tettamanti, Gianluca; Franzetti, Eleonora; Saviane, Alessio; Cappellozza, Silvia

2014-05-01

200

High-performance, low-operating voltage, and solution-processable organic field-effect transistor with silk fibroin as the gate dielectric  

Science.gov (United States)

We report the use of silk fibroin as the gate dielectric material in solution-processed organic field-effect transistors (OFETs) with poly(3-hexylthiophene) (P3HT) as the semiconducting layer. Such OFETs exhibit a low threshold of -0.77 V and a low-operating voltage (0 to -3 V) compatible with the voltage level commonly-used in current electronic industry. The carrier mobility of such OFETs is as high as 0.21 cm2 V-1 s-1 in the saturation regime, comparable to the best value of P3HT-based OFETs with dielectric layer that is not solution-processed. The high-performance of this kind of OFET is related with the high content of ? strands in fibroin dielectric which leads to an array of fibers in a highly ordered structure, thus reducing the trapping sites at the semiconductor/dielectric interface.

Shi, Leilei; Xu, Xinjun; Ma, Mingchao; Li, Lidong

2014-01-01

 
 
 
 
201

Immobilized Lentivirus Vector on Chondroitin Sulfate-Hyaluronate Acid-Silk Fibroin Hybrid Scaffold for Tissue-Engineered Ligament-Bone Junction  

Science.gov (United States)

The lack of a fibrocartilage layer between graft and bone remains the leading cause of graft failure after anterior cruciate ligament (ACL) reconstruction. The objective of this study was to develop a gene-modified silk cable-reinforced chondroitin sulfate-hyaluronate acid-silk fibroin (CHS) hybrid scaffold for reconstructing the fibrocartilage layer. The scaffold was fabricated by lyophilizing the CHS mixture with braided silk cables. The scanning electronic microscopy (SEM) showed that microporous CHS sponges were formed around silk cables. Each end of scaffold was modified with lentiviral-mediated transforming growth factor-?3 (TGF-?3) gene. The cells on scaffold were transfected by bonded lentivirus. In vitro culture demonstrated that mesenchymal stem cells (MSCs) on scaffolds proliferated vigorously and produced abundant collagen. The transcription levels of cartilage-specific genes also increased with culture time. After 2 weeks, the MSCs were distributed uniformly throughout scaffold. Deposited collagen was also found to increase. The chondral differentiation of MSCs was verified by expressions of collagen II and TGF-?3 genes in mRNA and protein level. Histology also confirmed the production of cartilage extracellular matrix (ECM) components. The results demonstrated that gene-modified silk cable-reinforced CHS scaffold was capable of supporting cell proliferation and differentiation to reconstruct the cartilage layer of interface.

Sun, Liguo; Li, Hongguo; Qu, Ling; Zhu, Rui; Fan, Xiangli; Xue, Yingsen; Xie, Zhenghong; Fan, Hongbin

2014-01-01

202

Comparative architecture of silks, fibrous proteins and their encoding genes in insects and spiders.  

Science.gov (United States)

The known silk fibroins and fibrous glues are thought to be encoded by members of the same gene family. All silk fibroins sequenced to date contain regions of long-range order (crystalline regions) and/or short-range order (non-crystalline regions). All of the sequenced fibroin silks (Flag or silk from flagelliform gland in spiders; Fhc or heavy chain fibroin silks produced by Lepidoptera larvae) are made up of hierarchically organized, repetitive arrays of amino acids. Fhc fibroin genes are characterized by a similar molecular genetic architecture of two exons and one intron, but the organization and size of these units differs. The Flag, Ser (sericin gene) and BR (Balbiani ring genes; both fibrous proteins) genes are made up of multiple exons and introns. Sequences coding for crystalline and non-crystalline protein domains are integrated in the repetitive regions of Fhc and MA exons, but not in the protein glues Ser1 and BR-1. Genetic 'hot-spots' promote recombination errors in Fhc, MA, and Flag. Codon bias, structural constraint, point mutations, and shortened coding arrays may be alternative means of stabilizing precursor mRNA transcripts. Differential regulation of gene expression and selective splicing of the mRNA transcript may allow rapid adaptation of silk functional properties to different physical environments. PMID:12470814

Craig, Catherine L; Riekel, Christian

2002-12-01

203

Tightly winding structure of sequential model peptide for repeated helical region in Samia cynthia ricini silk fibroin studied with solid-state NMR.  

Science.gov (United States)

There are many kinds of silks from silkworms and spiders with different structures and properties, and thus, silks are suitable to study the structure-property relationship of fibrous proteins. Silk fibroin from a wild silkworm, Samia cynthia ricini, mainly consists of the repeated similar sequences by about 100 times where there are alternative appearances of the polyalanine (Ala)(12-13) region and the Gly-rich region. In this paper, a sequential model peptide, GGAGGGYGGDGG(A)(12)GGAGDGYGAG, which is a typical sequence of the silk fibroin, was synthesized, and the atomic-level conformations of Gly residues at the N- and C-terminal ends of the polyalanine region were determined as well as that of the central Ala residue using (13)C 2D spin diffusion solid-state nuclear magnetic resonance (NMR) under off-magic angle spinning. In the model peptide with alpha-helical conformation, the torsion angle of the central Ala residue, the 19th Ala, was determined to be (phi, psi) = (-60 degrees, -50 degrees ), which was a typical alpha-helical structure, but the torsion angles of two Gly residues, the 12th and 25th Gly residues, which are located at the N- and C-terminal ends of the polyalanine region, were determined to be (phi, psi) = (-70 degrees, -30 degrees ) and (phi, psi) = (-70 degrees, -20 degrees ), respectively. Thus, it was observed that the turns at both ends of polyalanine with alpha-helix conformation in the model peptide are tightly wound. PMID:12649423

Nakazawa, Yasumoto; Bamba, Mie; Nishio, Satoko; Asakura, Tetsuo

2003-04-01

204

Dynamic processes involved in the pre-vascularization of silk fibroin constructs for bone regeneration using outgrowth endothelial cells.  

Science.gov (United States)

For successful bone regeneration tissue engineered bone constructs combining both aspects, namely a high osteogenic potential and a rapid connection to the vascular network are needed. In this study we assessed the formation of pre-vascular structures by human outgrowth endothelial cells (OEC) from progenitors in the peripheral blood and the osteogenic differentiation of primary human osteoblasts (pOB) on micrometric silk fibroin scaffolds. The rational was to gain more insight into the dynamic processes involved in the differentiation and functionality of both cell types depending on culture time in vitro. Vascular tube formation by OEC was assessed quantitatively at one and 4 weeks of culture. In parallel, we assessed the temporal changes in cell ratios by flow cytometry and in the marker profiles of endothelial and osteogenic markers by quantitative real-time PCR. In terms of OEC, we observed an increase in tube length, tube area, number of nodes and number of vascular meshes within a culture period of 4 weeks, but a decrease in endothelial markers in real-time PCR. At the same time early osteogenic markers were downregulated, while marker expression associated with progressing mineralized matrix was upregulated in later stages of the culture. In addition, deposition of matrix components, such as collagen type I, known as a pro-angiogenic substrate for endothelial cells, appeared to increase with time indicated by immunohistochemistry. In summary, the study suggests a progressing maturation of the tissue construct with culture time which seems to be not effected by culture conditions mainly designed for outgrowth endothelial cells. PMID:19091396

Fuchs, Sabine; Jiang, Xin; Schmidt, Harald; Dohle, Eva; Ghanaati, Shahram; Orth, Carina; Hofmann, Alexander; Motta, Antonella; Migliaresi, Claudio; Kirkpatrick, Charles J

2009-03-01

205

Fine-tuning scaffolds for tissue regeneration: effects of formic acid processing on tissue reaction to silk fibroin.  

Science.gov (United States)

Formic acid (FA) plays a key role in the preparation of silk fibroin (SF) scaffolds from cocoons of Bombyx mori and is used for fibre distribution. In this study, we used a subcutaneous implantation model in Wistar rats to examine SF scaffolds prepared by treating the degummed cocoon with FA for either 30 or 60 min. The tissue reaction and inflammatory response to SF was assessed by qualitative histology at intervals from 3 to 180 days. Additionally, dynamic biomaterial-induced vascularization and biomaterial degradation were quantified using a technique for analysing an image of the entire implanted biomaterial. Varying the FA treatment time led to different scaffold morphologies and resulted in two distinct peri-implant tissue reactions. The 30 min-treated scaffold was integrated into the surrounding tissue beginning at day 3 after implantation and vascularization increased 10-fold from 15 to 180 days, while the scaffold was continuously degraded throughout the first 90 days. In contrast, the 60 min-treated SF scaffold appeared as bulk for the first 90 days after implantation, after which a rapid degradation and vascularization process began. After 180 days, the tissue response was similar for both scaffolds, with eventual formation of a well vascularized connective tissue integrating the SF fibres. This study indicates that by modifying the FA treatment time, the tissue reaction to SF scaffolds can be tailored for different tissue-engineering applications. The tunability and biocompatibility of SF make it an attractive scaffold for exploration in regenerative medicine and clinical tissue engineering. PMID:20112273

Ghanaati, Shahram; Orth, Carina; Unger, Ronald E; Barbeck, Mike; Webber, Matthew J; Motta, Antonella; Migliaresi, Claudio; James Kirkpatrick, C

2010-08-01

206

Enhancement of anti-inflammatory activity of PEP-1-FK506 binding protein by silk fibroin peptide.  

Science.gov (United States)

Silk fibroin (SF) peptide has been traditionally used as a treatment for flatulence, spasms, and phlegm. In this study, we examined whether SF peptide enhanced the antiinflammatory effect of PEP-1-FK506 binding protein (PEP-1-FK506BP) through comparing the anti-inflammatory activities of SF peptide and/or PEP-1-FK506BP. In the presence or absence of SF peptide, transduction levels of PEP-1-FK506BP into HaCaT cells and mice skin and anti-inflammatory activities of PEP-1-FK506BP were identified by Western blot and histological analyses. SF peptide alone effectively reduced both mice ear edema and the elevated levels of cyclooxygenase-2, interleukin-6 and -1beta, and tumor necrosis factor-alpha, showing similar anti-inflammatory effect to that of PEP-1-FK506BP. Furthermore, co-treatment with SF peptide and PEP-1- FK506BP exhibited more enhanced anti-inflammatory effects than the samples treated with SF peptides or PEP- 1-FK506BP alone, suggesting the possibility that SF peptide and PEP-1-FK506BP might interact with each other. Moreover, the transduction data demonstrated that SF peptide did not affect the transduction of PEP-1- FK506BP into HaCaT cells and mice skin, indicating that the improvement of anti-inflammatory effect of PEP-1- FK506BP was not caused by enhanced transduction of PEP-1-FK506BP. Thus, these results suggest the possibility that co-treatment with SF peptide and PEP-1-FK506BP may be exploited as a useful therapy for various inflammationrelated diseases. PMID:22534296

Kim, Dae Won; Hwang, Hyun Sook; Kim, Duk-Soo; Sheen, Seung Hoon; Heo, Dong Hwa; Hwang, Gyojun; Kang, Suk Hyung; Kweon, Haeyong; Jo, You-Young; Kang, Seok Woo; Lee, Kwang-Gill; Park, Jinseu; Eum, Won Sik; Cho, Yong-Jun; Choi, Soo Young

2012-04-01

207

The development of injectable gelatin/silk fibroin microspheres for the dual delivery of curcumin and piperine.  

Science.gov (United States)

The objective of this study was to develop the microspheres from gelatin (G) and silk fibroin (SF) aimed to be applied for the controlled release of curcumin and piperine. The glutaraldehyde-crosslinked G/SF microspheres at various weight blending ratios (100/0, 70/30, 50/50, and 30/70) were successfully fabricated by water in oil emulsion technique. The microspheres prepared from all compositions were in a round shape with homogeneous size distribution both in the dried (194-217 ?m) and swollen states (297-367 ?m). When subjected in collagenase solution at physiological condition, the G microspheres gradually degraded within 14 days while the blended G/SF microspheres, particularly at 50/50 and 30/70, were not degraded. For the release application, the microspheres were loaded with curcumin and/or piperine. It was found that the microspheres composed of SF tended to entrap curcumin and piperine with the high entrapment and loading efficiencies, possibly due to their hydrophobic interactions. The G/SF microspheres, particularly at the ratios of 50/50 and 30/70, released curcumin and piperine in a sustained manner both for the single and dual release systems. The controlled dual release of curcumin and piperine from the G/SF microspheres would prolong their half-life, provide the optimal concentrations for therapeutic effects at a target site, and improve the bioavailability of curcumin. These novel injectable microspheres dually releasing curcumin and piperine would be introduced for the treatment of diseases without the need of operation. PMID:24186150

Ratanavaraporn, Juthamas; Kanokpanont, Sorada; Damrongsakkul, Siriporn

2014-02-01

208

New application of silk protein  

Energy Technology Data Exchange (ETDEWEB)

Gunma prefecture is famous for sericulture and silk textile industry district in Japan. In Gunma prefecture, some kinds of new generation silk as high performance and high quality silk were developed. These silk are used not only for the new textile materials but also for new industrial materials. New application of silk protein, fibroin and sericin, is considered. (author)

Kamiishi, Youichi [Textile Research Institute of Gunma, Kiryu, Gunma (Japan)

2000-03-01

209

DNA Synthesis in the Giant Nuclei of Insects - Control Machinery and Structures Observed in the Silk-Producing Gland of Bombyx Mori  

International Nuclear Information System (INIS)

The existence in many insect organs of giant nuclei without visible chromosomes raises the question of possible homologies between the chromatin structures of these nuclei and those of polytene nuclei or common euploid cells. Studies have been made of the nuclei in the silk-producing gland of Bombyx mori. The DNA synthesis is cyclic. During the third stage there are three successive synthesis cycles, which appear to be relatively autonomous in the individual nuclei. For more than 24 hours after moulting, however, synthesis is greatly reduced; moulting factors thus cause synchronization of all the nuclei. This leads to the conclusion that the triggering of a synthesis cycle is controlled by general factors external to the cell. At the end of larval development, DNA synthesis is suspended at the moment when large-scale secretion of silk begins. Evaluation of the pool of endogenic precursors of DNA shows that it is considerably reduced at the end of the DNA synthesis period. The hypothesis proposed is that large-scale synthesis of fibroin requires polarization of the metabolism, hence the depletion of the nucleotide pool and the end of DNA synthesis. DNA synthesis within a single nucleus is to some extent asynchronic. In particular, a well-defined, delayed-synthesis structure visible only in the female seems to be a possible homologue of a sex chromosome. Other asynchronisms are also apparent, though less clearly. Functional studies thus allow the supposition that in the giant nucleus replication units retain an individuality comparable to that of a polytene chromosome. These observations together lead to the conclusion that a nucleus in the silk-producing gland has physiological and structural characteristics similar to those of a polytene nucleus, differing from it essentially in the lesser degree of condensation of its structures. (author)

1968-06-01

210

Characterization of Tetracycline-loaded Thai Silk Fibroin/Gelatin Blend Films  

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Full Text Available This study was aimed to prepare SF/G blend films loaded tetracycline by a solvent evaporation method. Firstly silk cocoons were degummed and then dissolved by CaCl2:Ethanol:H2O (1:2:8 by mole. On another way, Gelatin (G was prepared from gelatin powder. The mixture solution of SF/G ratios with 3:0, 2:1, 1:1, 1:2 and 0:3 were prepared before casting on the 5 cm polystyrene plates. All of films were investigated their morphology, chemical structures, thermal properties and percent transparency by using Scanning Electron Microscope (SEM, Fourier transform infrared (FTIR spectroscopy, Thermogravimetric Analyzer (TA and UV-Vis spectrometer, respectively. The results found that the surfaces of blend films were gradually smooth without phase separation when the SF component decreased. The FTIR results of SF/G films showed strong regions for amide I, amide II and amide III which were the mixture characteristics of SF and G. The blend films rapidly decomposed in maximum rate after 300°C. The rate of weight lost depending on the content G meanwhile rapidly increased of weight lost when the G content was increased. The heat flow curves indicated that the blend films composed of multiple peaks of maximum decomposition temperatures as well as endo/exo-thermic. Finally, tetracycline could be interacted with G in excellent profile affected to the lowest of percent transparency.

Srihanam Prasong

2010-01-01

211

Silk Coatings on PLGA and Alginate Microspheres for Protein Delivery  

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Bombyx mori silk fibroin self-assembles on surfaces to form ultrathin nanoscale coatings based on our prior studies using layer-by-layer deposition techniques driven by hydrophobic interactions between silk fibroin protein molecules. In the present study, polylactic-co-glycolic acid (PLGA) and alginate microspheres were used as substrates and coated with silk fibroin. The coatings were visualized by confocal laser scanning microscopy using fluorescein-labeled silk fibroin. On PLGA microsphere...

Wang, Xiaoqin; Wenk, Esther; Hu, Xiao; Castro, Guillermo R.; Meinel, Lorenz; Wang, Xianyan; Li, Chunmei; Merkle, Hans; Kaplan, David L.

2007-01-01

212

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

Science.gov (United States)

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. PMID:21882171

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

2012-06-01

213

Morphology and composition of the spider major ampullate gland and dragline silk.  

Science.gov (United States)

Spider silk is made of unique proteins-spidroins-secreted and stored as a protein solution (dope) in specialized glands. The major ampullate gland, source of the dragline silk, is composed of a tail, a sac and an elongated duct. For this gland, several different types of epithelial cells and granules have been described, but it is largely unknown how they correlate with spidroin production. It is also not settled what parts of the large spidroins end up in the final silk, and it has been suggested that the N-terminal domain (NT) is lacking. Here we show that NT is present in the dope and throughout dragline silk fibers, including the skin layer, and that the major ampullate tail and sac consist of three different and sharply demarcated zones (A-C), each with a distinct epithelial cell type. Finally, we show that spidroins are produced in the A and B zone epithelia, while the C zone granules lack spidroins. PMID:23837699

Andersson, Marlene; Holm, Lena; Ridderstråle, Yvonne; Johansson, Jan; Rising, Anna

2013-08-12

214

In vitro and in vivo studies on the cytotoxicity of irradiated silk fibroin against mouse melanoma tumor cell  

International Nuclear Information System (INIS)

The physicochemical properties of proteins can be altered by irradiation. But, it is rarely that the researches on the functional properties of irradiated proteins have been reported. Fibroin is a fibrous protein derived from silkworm Bombyx mori and has been suggested as a biomaterial for biomedical application. Therefore, fibroin was selected as a model protein and was examined with the irradiation effects on the cytotoxicity of fibroin on tumor cell. The cytotoxicity of fibroin against mouse melanoma cell (B16BL6) showed a significant increase dependent upon the increase of irradiation dose. And also, the splenocyte proliferation activities of fibroin were increased by gamma irradiation. In addition, the oral administration of irradiated fibroin significantly increased the inhibition rate of tumor growth in tumor-bearing mouse model. The reason might be due to the change of protein structure by gamma irradiation and is being studied. From these result, it could be concluded that the irradiated fibroin might be a potential candidate as a valuable product in food and medical industry.

2008-09-21

215

Fabrication and Biocompatibility of Electrospun Silk Biocomposites  

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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 ...

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

2011-01-01

216

One-pot, green, rapid synthesis of flowerlike gold nanoparticles/reduced graphene oxide composite with regenerated silk fibroin as efficient oxygen reduction electrocatalysts.  

Science.gov (United States)

Flowerlike gold nanoparticles (Au NPs)/reduced graphene oxide (RGO) composites were fabricated by a facile, one-pot, environmentally friendly method in the presence of regenerated silk fibroin (RSF). The influences of reaction time, temperature, and HAuCl(4): RGO ratio on the morphology of Au NPs loaded on RGO sheets were discussed and a tentative mechanism for the formation of flowerlike Au NPs/RGO composite was proposed. In addition, the flowerlike Au NPs/RGO composite showed superior catalytic performance for oxygen reduction reaction (ORR) to Au/RGO composites with other morphologies. Our work provides an alternative facile and green approach to synthesize functional metal/RGO composites. PMID:23323590

Xu, Shengjie; Yong, Liu; Wu, Peiyi

2013-02-01

217

Purification and some properties of G-factor from the silk gland of silkworm  

International Nuclear Information System (INIS)

Purification of G-factor from the silk gland of silkworms and some of its properties have been studied in comparison with the G-factor from E. coli and rat liver transferase II. In the experiment, GTP-?-32P was prepared from GDP and 32Pi by photophosphorylation with spinach chloroplasts, and purified by Dowex-1 chromatography. The purified G-factor was homogeneous on disc gel electrophoresis. Molecular weight of 80,000 was estimated from a calibrated Sephadex G-200 column. The purified G-factor exhibited uncoupled ribosome-dependent GTPase activity, and was inhibited by fusidic acid, SH inhibitor, and diphtheria toxin plus NAD. The G-factor from E. coli and that from the silk gland were not interchangeable in GTPase reaction. (Yamanoto, Y.)

1972-01-01

218

A silk fibroin based hepatocarcinoma model and the assessment of the drug response in hyaluronan-binding protein 1 overexpressed HepG2 cells.  

Science.gov (United States)

Microenvironment around tumor cells plays an important role in its malignancy or invasiveness. Hyaluronan (HA), a major component of extracellular matrix is found to be elevated in most of cancerous niche/microenvironment and performs regulatory role in the progression of tumors and metastasis. Overexpression of the hyaladherin, hyaluronan-binding protein 1 (HABP1) in the hepatocarcinoma cells (HepG2) termed as HepR21 leads to enhanced cell proliferation with increased HA 'pool' associated with HA 'cables' indicating elevated tumorous potential under 2D culture conditions. For in vitro experimentation, scaffold based three dimensional niche modeling may have greater acceptance than conventional 2D culture condition. Thus, we have examined the influence of intrinsic properties of non-mulberry tropical tasar silk fibroin on the HepR21 cells in order to develop a 3D hepatocarcinoma construction to act as model. The scaffold of tasar silk fibroin of Antheraea mylitta when efficiently loaded with transformed hepatocarcinoma cells, HepR21; exhibits enhanced adhesiveness, viability, metabolic activity, proliferation and enlarged cellular morphology in 3D compared to its parent cell line HepG2, supporting the earlier observation made in 2D system. In addition, formation of multicellular aggregates, the indicator of tumor progression is also revealed in silk based 3D culture conditions. Further, the use of 4-MU (a hyaluronan synthase inhibitor) on HepR21 cells reduces the HA level and downregulates the expression of growth promoting factors like pAKT and PKC; while upregulating the expression of the tumor suppressor p53. Thus, 4-MU efficiently reduces the tumor potency associated with increased HA pool as well as HA cables and the effect of 4-MU doubling up as an anticancer agent in 2D and 3D are also comparable. The in vitro 3D multicellular model demonstrates the insight of hepatocarcinoma progression and offers the predictability of cellular response to transfection efficacy, drug treatment and therapeutic intervention. PMID:24016853

Kundu, Banani; Saha, Paramita; Datta, Kasturi; Kundu, Subhas C

2013-12-01

219

Bone Tissue Engineering with Premineralized Silk Scaffolds  

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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 ...

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

2008-01-01

220

Solution structure of eggcase silk protein and its implications for silk fiber formation  

Science.gov (United States)

Spider silks are renowned for their excellent mechanical properties and biomimetic and industrial potentials. They are formed from the natural refolding of water-soluble fibroins with ?-helical and random coil structures in silk glands into insoluble fibers with mainly ?-structures. The structures of the fibroins at atomic resolution and silk formation mechanism remain largely unknown. Here, we report the 3D structures of individual domains of a ?366-kDa eggcase silk protein that consists of 20 identical type 1 repetitive domains, one type 2 repetitive domain, and conserved nonrepetitive N- and C-terminal domains. The structures of the individual domains in solution were determined by using NMR techniques. The domain interactions were investigated by NMR and dynamic light-scattering techniques. The formation of micelles and macroscopic fibers from the domains was examined by electron microscopy. We find that either of the terminal domains covalently linked with at least one repetitive domain spontaneously forms micelle-like structures and can be further transformed into fibers at ?37 °C and a protein concentration of >0.1 wt%. Our biophysical and biochemical experiments indicate that the less hydrophilic terminal domains initiate the assembly of the proteins and form the outer layer of the micelles whereas the more hydrophilic repetitive domains are embedded inside to ensure the formation of the micelle-like structures that are the essential intermediates in silk formation. Our results establish the roles of individual silk protein domains in fiber formation and provide the basis for designing miniature fibroins for producing artificial silks.

Lin, Zhi; Huang, Weidong; Zhang, Jingfeng; Fan, Jing-Song; Yang, Daiwen

2009-01-01

 
 
 
 
221

Engineering adipose-like tissue in vitro and in vivo utilizing human bone marrow and adipose-derived mesenchymal stem cells with silk fibroin 3D scaffolds.  

Science.gov (United States)

Biomaterials derived from silk fibroin prepared by aqueous (AB) and organic (HFIP) solvent-based processes, along with collagen (COL) and poly-lactic acid (PLA)-based scaffolds were studied in vitro and in vivo for their utility in adipose tissue engineering strategies. For in vitro studies, human bone marrow and adipose-derived mesenchymal stem cells (hMSCs and hASCs) were seeded on the various biomaterials and cultured for 21 days in the presence of adipogenic stimulants (AD) or maintained as noninduced controls. Alamar Blue analysis revealed each biomaterial supported initial attachment of hMSCs and hASCs to similar levels for all matrices except COL in which higher levels were observed. hASCs and hMSCs cultured on all biomaterials in the presence of AD showed significant upregulation of adipogenic mRNA transcript levels (LPL, GLUT4, FABP4, PPARgamma, adipsin, ACS) to similar extents when compared to noninduced controls. Similarly Oil-Red O analysis of hASC or hMSC-seeded scaffolds displayed substantial amounts of lipid accumulating adipocytes following cultivation with AD. The data revealed AB and HFIP scaffolds supported similar extents of lipid accumulating cells while PLA and COL scaffolds qualitatively displayed lower and higher extents by comparison, respectively. Following a 4-week implantation period in a rat muscle pouch defect model, both AB and HFIP scaffolds supported in vivo adipogenesis either alone or seeded with hASCs or hMSCs as assessed by Oil-Red O analysis, however the presence of exogenous cell sources substantially increased the extent and frequency of adipogenesis observed. In contrast, COL and PLA scaffolds underwent rapid scaffold degradation and were irretrievable following the implantation period. The results suggest that macroporous 3D AB and HFIP silk fibroin scaffolds offer an important platform for cell-based adipose tissue engineering applications, and in particular, provide longer-term structural integrity to promote the maintenance of soft tissue in vivo. PMID:17765303

Mauney, Joshua R; Nguyen, Trang; Gillen, Kelly; Kirker-Head, Carl; Gimble, Jeffrey M; Kaplan, David L

2007-12-01

222

The role of physiological mechanical cues on mesenchymal stem cell differentiation in an airway tract-like dense collagen-silk fibroin construct.  

Science.gov (United States)

Airway tracts serve as a conduit of transport in the respiratory system. Architecturally, these are composed of cartilage rings that offer flexibility and prevent collapse during normal breathing. To this end, the successful regeneration of an airway tract requires the presence of differentiated chondrocytes and airway smooth muscle cells. This study investigated the role of physiological dynamic mechanical stimulation, in vitro, on the differentiation of mesenchymal stem cells (MSCs), three-dimensionally seeded within a tubular dense collagen matrix construct-reinforced with rings of electrospun silk fibroin mat (TDC-SFC). In particular, the role of either shear stress supplied by laminar fluid flow or cyclic shear stress in combination with circumferential strain, provided by pulsatile flow, on the chondrogenic differentiation, and contractile lineage of MSCs, and their effects on TDC-SFC morphology and mechanical properties were analysed. Chondrogenic differentiation of MSCs was observed in the presence of chondrogenic supplements under both static and laminar flow cultures. In contrast, physiological pulsatile flow resulted in preferential cellular orientation within TDC-SFC, as dictated by dynamic circumferential strain, and induced MSC contractile phenotype expression. In addition, pulsatile flow decreased MSC-mediated collagen matrix remodelling and increased construct circumferential strength. Therefore, TDC-SFC demonstrated the central role of a matrix in the delivery of mechanical stimuli over chemical factors, by providing an in vitro niche to control MSC differentiation, alignment and its capacity to remodel the matrix. PMID:24818890

Ghezzi, Chiara E; Marelli, Benedetto; Donelli, Ilaria; Alessandrino, Antonio; Freddi, Giuliano; Nazhat, Showan N

2014-08-01

223

Amperometric biosensor based on tyrosinase immobilized onto multiwalled carbon nanotubes-cobalt phthalocyanine-silk fibroin film and its application to determine bisphenol A  

International Nuclear Information System (INIS)

An amperometric bisphenol A (BPA) biosensor was fabricated by immobilizing tyrosinase on multiwalled carbon nanotubes (MWNTs)-cobalt phthalocyanine (CoPc)-silk fibroin (SF) composite modified glassy carbon electrode (GCE). In MWNTs-CoPc-SF composite film, SF provided a biocompatible microenvironment for the tyrosinase to retain its bioactivity, MWNTs possessed excellent inherent conductivity to enhance the electron transfer rate and CoPc showed good electrocatalytic activity to electrooxidation of BPA. The cyclic voltammogram of BPA at this biosensor exhibited a well defined anodic peak at 0.625 V. Compared with bare GCE, the oxidation signal of BPA significantly increased; therefore, this oxidation signal was used to determine BPA. The effect factors were optimized and the electrochemical parameters were calculated. The possible oxidation mechanism was also discussed. Under optimum conditions, the oxidation current was proportional to BPA concentration in the range from 5.0 x 10-8 to 3.0 x 10-6 M with correlation coefficient of 0.9979 and detection limit of 3.0 x 10-8 M (S/N = 3). The proposed method was successfully applied to determine BPA in plastic products and the recovery was in the range from 95.36% to 104.39%.

2010-02-05

224

Amperometric biosensor based on tyrosinase immobilized onto multiwalled carbon nanotubes-cobalt phthalocyanine-silk fibroin film and its application to determine bisphenol A  

Energy Technology Data Exchange (ETDEWEB)

An amperometric bisphenol A (BPA) biosensor was fabricated by immobilizing tyrosinase on multiwalled carbon nanotubes (MWNTs)-cobalt phthalocyanine (CoPc)-silk fibroin (SF) composite modified glassy carbon electrode (GCE). In MWNTs-CoPc-SF composite film, SF provided a biocompatible microenvironment for the tyrosinase to retain its bioactivity, MWNTs possessed excellent inherent conductivity to enhance the electron transfer rate and CoPc showed good electrocatalytic activity to electrooxidation of BPA. The cyclic voltammogram of BPA at this biosensor exhibited a well defined anodic peak at 0.625 V. Compared with bare GCE, the oxidation signal of BPA significantly increased; therefore, this oxidation signal was used to determine BPA. The effect factors were optimized and the electrochemical parameters were calculated. The possible oxidation mechanism was also discussed. Under optimum conditions, the oxidation current was proportional to BPA concentration in the range from 5.0 x 10{sup -8} to 3.0 x 10{sup -6} M with correlation coefficient of 0.9979 and detection limit of 3.0 x 10{sup -8} M (S/N = 3). The proposed method was successfully applied to determine BPA in plastic products and the recovery was in the range from 95.36% to 104.39%.

Yin Huanshun [College of Chemistry and Material Science, Shandong Agricultural University, Taian 271018, Shandong (China); College of Resources and Environment, Shandong Agricultural University, Taian 271018, Shandong (China); Zhou Yunlei; Xu Jing [College of Chemistry and Material Science, Shandong Agricultural University, Taian 271018, Shandong (China); Ai Shiyun, E-mail: ashy@sdau.edu.cn [College of Chemistry and Material Science, Shandong Agricultural University, Taian 271018, Shandong (China); Cui Lin [College of Chemistry and Material Science, Shandong Agricultural University, Taian 271018, Shandong (China); Zhu Lusheng, E-mail: lushzhu@sdau.edu.cn [College of Resources and Environment, Shandong Agricultural University, Taian 271018, Shandong (China)

2010-02-05

225

Evaluation of an injectable silk fibroin enhanced calcium phosphate cement loaded with human recombinant bone morphogenetic protein-2 in ovine lumbar interbody fusion.  

Science.gov (United States)

The objective of this study was to investigate the efficacy of an injectable calcium phosphate cement/silk fibroin/human recombinant bone morphogenetic protein-2 composite (CPC/SF/rhBMP-2) in an ovine interbody fusion model. Twenty-four mature sheep underwent anterior lumbar interbody fusion at the levels of L1/2, L3/4, and L5/6 with random implantation of CPC/SF, CPC/rhBMP-2, CPC/SF/rhBMP-2, or autogenous iliac bone. After the sheep were sacrificed, the fusion segments were evaluated by manual palpation, CT scan, undestructive biomechanical testing, undecalcified histology, and histomorphology. The fusion rates of CPC/SF/rhBMP-2 were 55.56% and 77.78% at 6 and 12 months, respectively. The fusion was superior to all the biomaterial grafts in stiffness, and reached the same stiffness as the autograft at 12 months. The new bone formation was less than autograft at 6 months, but similar with that at 12 months. However, the ceramic residue volume of CPC/SF/rhBMP-2 was significantly decreased compared with CPC/SF and CPC/rhBMP-2 at both times. The results indicated that CPC/SF/rhBMP-2 composite had excellent osteoconduction and osteoinduction, and balanced degradation and osteogenesis. PMID:21381189

Gu, Yong; Chen, Liang; Yang, Hui-Lin; Luo, Zong-Ping; Tang, Tian-Si

2011-05-01

226

The development of silk fibroin scaffolds using an indirect rapid prototyping approach: morphological analysis and cell growth monitoring by spectral-domain optical coherence tomography.  

Science.gov (United States)

To date, naturally derived biomaterials are rarely used in advanced tissue engineering (TE) methods despite their superior biocompatibility. This is because these native materials, which consist mainly of proteins and polysaccharides, do not possess the ability to withstand harsh processing conditions. Unlike synthetic polymers, natural materials degrade and decompose rapidly in the presence of chemical solvents and high temperature, respectively. Thus, the fabrication of tissue scaffolds using natural biomaterials is often carried out using conventional techniques, where the efficiency in mass transport of nutrients and removal of waste products within the construct is compromised. The present study identified silk fibroin (SF) protein as a suitable material for the application of rapid prototyping (RP) or additive manufacturing (AM) technology. Using the indirect RP method, via the use of a mould, SF tissue scaffolds with both macro- and micro-morphological features can be produced and qualitatively examined by spectral-domain optical coherence tomography (SD-OCT). The advanced imaging technique showed the ability to differentiate the cells and SF material by producing high contrasting images, therefore suggesting the method as a feasible alternative to the histological analysis of cell growth within tissue scaffolds. PMID:22055004

Liu, M J J; Chou, S M; Chua, C K; Tay, B C M; Ng, B K

2013-02-01

227

Silk gland-specific tRNA(Ala) genes interact more weakly than constitutive tRNA(Ala) genes with silkworm TFIIIB and polymerase III fractions.  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Constitutive and silk gland-specific tRNA(Ala) genes from silkworms have very different transcriptional properties in vitro. Typically, the constitutive type, which encodes tRNA(AlaC), directs transcription much more efficiently than does the silk gland-specific type, which encodes tRNA(AlaSG). We think that the inefficiency of the tRNA(AlaCG) gene underlies its capacity to be turned off in non-silk gland cells. An economical model is that the tRNA(AlaSG) promoter interacts poorly, relative t...

Sullivan, H. S.; Young, L. S.; White, C. N.; Sprague, K. U.

1994-01-01

228

Biomaterials Derived from Silk-Tropoelastin Protein Systems  

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A structural protein blend system based on silkworm silk fibroin and recombinant human tropoelastin is described. Silk fibroin, a semicrystalline fibrous protein with beta-sheet crystals provides mechanical strength and controllable biodegradation, while tropoelastin, a noncrystallizable elastic protein provides elasticity. Differential scanning calorimetry (DSC) and temperature modulated DSC (TMDSC) indicated that silk becomes miscible with tropoelastin at different blend ratios, without mac...

2010-01-01

229

The role of irregular unit, GAAS, on the secondary structure of Bombyx mori silk fibroin studied with 13C CP/MAS NMR and wide-angle X-ray scattering  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Bombyx mori silk fibroin is a fibrous protein whose fiber is extremely strong and tough, although it is produced by the silkworm at room temperature and from an aqueous solution. The primary structure is mainly Ala-Gly alternative copolypeptide, but Gly-Ala-Ala-Ser units appear frequently and periodically. Thus, this study aims at elucidating the role of such Gly-Ala-Ala-Ser units on the secondary structure. The sequential model peptides containing Gly-Ala-Ala-Ser units selected from the prim...

Asakura, Tetsuo; Sugino, Rena; Okumura, Tatsushi; Nakazawa, Yasumoto

2002-01-01

230

Antheraea pernyi silk fibroin-coated PEI/DNA complexes for targeted gene delivery in HEK 293 and HCT 116 cells.  

Science.gov (United States)

Polyethylenimine (PEI) has attracted much attention as a DNA condenser, but its toxicity and non-specific targeting limit its potential. To overcome these limitations, Antheraea pernyi silk fibroin (ASF), a natural protein rich in arginyl-glycyl-aspartic acid (RGD) peptides that contains negative surface charges in a neutral aqueous solution, was used to coat PEI/DNA complexes to form ASF/PEI/DNA ternary complexes. Coating these complexes with ASF caused fewer surface charges and greater size compared with the PEI/DNA complexes alone. In vitro transfection studies revealed that incorporation of ASF led to greater transfection efficiencies in both HEK (human embryonic kidney) 293 and HCT (human colorectal carcinoma) 116 cells, albeit with less electrostatic binding affinity for the cells. Moreover, the transfection efficiency in the HCT 116 cells was higher than that in the HEK 293 cells under the same conditions, which may be due to the target bonding affinity of the RGD peptides in ASF for integrins on the HCT 116 cell surface. This result indicated that the RGD binding affinity in ASF for integrins can enhance the specific targeting affinity to compensate for the reduction in electrostatic binding between ASF-coated PEI carriers and cells. Cell viability measurements showed higher cell viability after transfection of ASF/PEI/DNA ternary complexes than after transfection of PEI/DNA binary complexes alone. Lactate dehydrogenase (LDH) release studies further confirmed the improvement in the targeting effect of ASF/PEI/DNA ternary complexes to cells. These results suggest that ASF-coated PEI is a preferred transfection reagent and useful for improving both the transfection efficiency and cell viability of PEI-based nonviral vectors. PMID:24776757

Liu, Yu; You, Renchuan; Liu, Guiyang; Li, Xiufang; Sheng, Weihua; Yang, Jicheng; Li, Mingzhong

2014-01-01

231

A method for studying the structure of uniaxially aligned biopolymers using solid state 15N-nmr: application to Bombyx mori silk fibroin fibers.  

Science.gov (United States)

Recent advances in the application of solid state nmr spectroscopy to uniformly aligned biopolymers have opened a window through which to view the detailed structure of biological macromolecules that are unable to be seen with standard techniques for structure determination such as x-ray diffraction. Atomic resolution structural details are obtained from solid state nmr data in the form of bond orientations, which yield the relative positions of specific atoms within the molecule. For static aligned systems such as fibers, in which rapid reorientation about the axis of alignment does not occur, it has generally been necessary to perform trial and error line-shape simulations to extract structural details from nmr spectra arising from a single type of nuclear spin interaction. In the present work, a new method is developed in which solid state 15N-nmr spectra obtained from uniaxially aligned molecules placed with the axis of alignment both parallel and perpendicular to the magnetic field are analyzed to yield the orientations of specific molecular bonds. Analytical expressions are derived that utilize spectral features read from 15N chemical shift anisotropy line shapes to calculate a discrete number of possible orientations for a specific site. The 15N-1H dipolar interaction is employed to further narrow the number of unique orientations possible for a given site. With this method, a neighborhood of possible orientations is quickly determined, and full line-shape simulations within this region of allowed space can be performed to refine the limits of orientation. This technique demonstrates the use of a single type of isotopic label to determine the orientation of a specific molecular group such as a peptide plane within a protein. Results from the application of this method to the Bombyx mori silk fibroin protein provide structural detail that is consistent with currently accepted structural models based on fiber diffraction studies. PMID:8343580

Nicholson, L K; Asakura, T; Demura, M; Cross, T A

1993-05-01

232

The use of bi-layer silk fibroin scaffolds and small intestinal submucosa matrices to support bladder tissue regeneration in a rat model of spinal cord injury.  

Science.gov (United States)

Adverse side-effects associated with enterocystoplasty for neurogenic bladder reconstruction have spawned the need for the development of alternative graft substitutes. Bi-layer silk fibroin (SF) scaffolds and small intestinal submucosa (SIS) matrices were investigated for their ability to support bladder tissue regeneration and function in a rat model of spinal cord injury (SCI). Bladder augmentation was performed with each scaffold configuration in SCI animals for 10 wk of implantation and compared to non-augmented control groups (normal and SCI alone). Animals subjected to SCI alone exhibited a 72% survival rate (13/18) while SCI rats receiving SIS and bi-layer SF scaffolds displayed respective survival rates of 83% (10/12) and 75% (9/12) over the course of the study period. Histological (Masson's trichrome analysis) and immunohistochemical (IHC) evaluations demonstrated both implant groups supported de novo formation of smooth muscle layers with contractile protein expression [?-smooth muscle actin (?-SMA) and SM22?] as well as maturation of multi-layer urothelia expressing cytokeratin (CK) and uroplakin 3A proteins. Histomorphometric analysis revealed bi-layer SF and SIS scaffolds respectively reconstituted 64% and 56% of the level of ?-SMA+ smooth muscle bundles present in SCI-alone controls, while similar degrees of CK+ urothelium across all experimental groups were detected. Parallel evaluations showed similar degrees of vascular area and synaptophysin+ boutons in all regenerated tissues compared to SCI-alone controls. In addition, improvements in certain urodynamic parameters in SCI animals, such as decreased peak intravesical pressure, following implantation with both matrix configurations were also observed. The data presented in this study detail the ability of acellular SIS and bi-layer SF scaffolds to support formation of innervated, vascularized smooth muscle and urothelial tissues in a neurogenic bladder model. PMID:24917031

Chung, Yeun Goo; Algarrahi, Khalid; Franck, Debra; Tu, Duong D; Adam, Rosalyn M; Kaplan, David L; Estrada, Carlos R; Mauney, Joshua R

2014-08-01

233

Characterization of dielectrophoresis-aligned nanofibrous silk fibroin-chitosan scaffold and its interactions with endothelial cells for tissue engineering applications.  

Science.gov (United States)

Aligned three-dimensional nanofibrous silk fibroin-chitosan (eSFCS) scaffolds were fabricated using dielectrophoresis (DEP) by investigating the effects of alternating current frequency, the presence of ions, the SF:CS ratio and the post-DEP freezing temperature. Scaffolds were characterized with polarized light microscopy to analyze SF polymer chain alignment, atomic force microscopy (AFM) to measure the apparent elastic modulus, and scanning electron microscopy and AFM to analyze scaffold topography. The interaction of human umbilical vein endothelial cells (HUVECs) with eSFCS scaffolds was assessed using immunostaining to assess cell patterning and AFM to measure the apparent elastic modulus of the cells. The eSFCS (50:50) samples prepared at 10MHz with NaCl had the highest percentage of aligned area as compared to other conditions. As DEP frequency increased from 100kHz to 10MHz, fibril sizes decreased significantly. eSFCS (50:50) scaffolds fabricated at 10MHz in the presence of 5mM NaCl had a fibril size of 77.96±4.69nm and an apparent elastic modulus of 39.9±22.4kPa. HUVECs on eSFCS scaffolds formed aligned and branched capillary-like vascular structures. The elastic modulus of HUVEC cultured on eSFCS was 6.36±2.37kPa. DEP is a potential tool for fabrication of SFCS scaffolds with aligned nanofibrous structures that can guide vasculature in tissue engineering and repair. PMID:24821141

Dunne, Lina W; Iyyanki, Tejaswi; Hubenak, Justin; Mathur, Anshu B

2014-08-01

234

Antheraea pernyi Silk Fibroin-Coated PEI/DNA Complexes for Targeted Gene Delivery in HEK 293 and HCT 116 Cells  

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Full Text Available Polyethylenimine (PEI has attracted much attention as a DNA condenser, but its toxicity and non-specific targeting limit its potential. To overcome these limitations, Antheraea pernyi silk fibroin (ASF, a natural protein rich in arginyl-glycyl-aspartic acid (RGD peptides that contains negative surface charges in a neutral aqueous solution, was used to coat PEI/DNA complexes to form ASF/PEI/DNA ternary complexes. Coating these complexes with ASF caused fewer surface charges and greater size compared with the PEI/DNA complexes alone. In vitro transfection studies revealed that incorporation of ASF led to greater transfection efficiencies in both HEK (human embryonic kidney 293 and HCT (human colorectal carcinoma 116 cells, albeit with less electrostatic binding affinity for the cells. Moreover, the transfection efficiency in the HCT 116 cells was higher than that in the HEK 293 cells under the same conditions, which may be due to the target bonding affinity of the RGD peptides in ASF for integrins on the HCT 116 cell surface. This result indicated that the RGD binding affinity in ASF for integrins can enhance the specific targeting affinity to compensate for the reduction in electrostatic binding between ASF-coated PEI carriers and cells. Cell viability measurements showed higher cell viability after transfection of ASF/PEI/DNA ternary complexes than after transfection of PEI/DNA binary complexes alone. Lactate dehydrogenase (LDH release studies further confirmed the improvement in the targeting effect of ASF/PEI/DNA ternary complexes to cells. These results suggest that ASF-coated PEI is a preferred transfection reagent and useful for improving both the transfection efficiency and cell viability of PEI-based nonviral vectors.

Yu Liu

2014-04-01

235

Reinforcing Silk Scaffolds with Silk Particles  

Science.gov (United States)

Silk fibroin is a useful protein polymer for biomaterials and tissue engineering. In this work, porogen leached scaffolds prepared from aqueous and HFIP silk solutions were reinforced through the addition of silk particles. This led to about 40 times increase in the specific compressive modulus and the yield strength of HFIP-based scaffolds. This increase in mechanical properties resulted from the high interfacial cohesion between the silk matrix and the reinforcing silk particles, due to partial solubility of the silk particles in HFIP. The porosity of scaffolds was reduced from ?90% (control) to ?75% for the HFIP systems containing 200% particle reinforcement, while maintaining pore interconnectivity. The presence of the particles slowed the enzymatic degradation of silk scaffolds.

Rajkhowa, Rangam; Gil, Eun Seok; Kluge, Jonathan; Numata, Keiji; Wang, Lijing; Kaplan, David L.

2014-01-01

236

CHANGES IN FINE STRUCTURE DURING SILK PROTEIN PRODUCTION IN THE AMPULLATE GLAND OF THE SPIDER ARANEUS SERICATUS  

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The ampullate silk gland of the spider, Araneus sericatus, produces the silk fiber for the scaffolding of the web. The fine structure of the various parts of the gland is described. The distal portion of the duct consist of a tube of epithelial cells which appear to secrete a substance which forms the tunica intima of the duct wall. At the proximal end of the duct there is a region of secretory cells. The epithelium of the sac portion contains five morphologically distinct types of granules. The bulk of the synthesis of silk occurs in the tail of the gland, and in this region only a single type of secretory droplet is seen in the epithelium. Protein synthesis can be stimulated by the injection of 1 mg/kg acetylcholine into the body fluids. 10 min after injection, much of the protein stored in the cytoplasm of the epithelial cells has been secreted into the lumen. 20 min after stimulation, the ergastoplasmic sacs form large whorls in the cytoplasm. Protein, similar in electron-opacity to protein found in the lumen, begins to form in that portion of the cytoplasm which is enclosed by the whorls. The limiting membrane of these droplets is formed by ergastoplasmic membranes which lose their ribosomes. No Golgi material has been found in these cells. Protein appears to be manufactured in the cytoplasm of the tail cells in a form which is ready for secretion.

Bell, Allen L.; Peakall, David B.

1969-01-01

237

Production of an active feline interferon in the cocoon of transgenic silkworms using the fibroin H-chain expression system  

International Nuclear Information System (INIS)

We constructed the fibroin H-chain expression system to produce recombinant proteins in the cocoon of transgenic silkworms. Feline interferon (FeIFN) was used for production and to assess the quality of the product. Two types of FeIFN fusion protein, each with N- and C-terminal sequences of the fibroin H-chain, were designed to be secreted into the lumen of the posterior silk glands. The expression of the FeIFN/H-chain fusion gene was regulated by the fibroin H-chain promoter domain. The transgenic silkworms introduced these constructs with the piggyBac transposon-derived vector, which produced the normal sized cocoons containing each FeIFN/H-chain fusion protein. Although the native-protein produced by transgenic silkworms have almost no antiviral activity, the proteins after the treatment with PreScission protease to eliminate fibroin H-chain derived N- and C-terminal sequences from the products, had very high antiviral activity. This H-chain expression system, using transgenic silkworms, could be an alternative method to produce an active recombinant protein and silk-based biomaterials

2007-04-20

238

Structure determination of a peptide model of the repeated helical domain in Samia cynthia ricini silk fibroin before spinning by a combination of advanced solid-state NMR methods.  

Science.gov (United States)

Fibrous proteins unlike globular proteins, contain repetitive amino acid sequences, giving rise to very regular secondary protein structures. Silk fibroin from a wild silkworm, Samia cynthia ricini, consists of about 100 repeats of alternating polyalanine (poly-Ala) regions of 12-13 residues in length and Gly-rich regions. In this paper, the precise structure of the model peptide, GGAGGGYGGDGG(A)(12)GGAGDGYGAG, which is a typical repeated sequence of the silk fibroin, was determined using a combination of three kinds of solid-state NMR studies; a quantitative use of (13)C CP/MAS NMR chemical shift with conformation-dependent (13)C chemical shift contour plots, 2D spin diffusion (13)C solid-state NMR under off magic angle spinning and rotational echo double resonance. The structure of the model peptide corresponding to the silk fibroin structure before spinning was determined. The torsion angles of the central Ala residue, Ala(19), in the poly-Ala region were determined to be (phi, psi) = (-59 degrees, -48 degrees ) which are values typically associated with alpha-helical structures. However, the torsion angles of the Gly(25) residue adjacent to the C-terminal side of the poly-Ala chain were determined to be (phi, psi) = (-66 degrees, -22 degrees ) and those of Gly(12) and Ala(13) residues at the N-terminal of the poly-Ala chain to be (phi, psi) = (-70 degrees, -30 degrees ). In addition, REDOR experiments indicate that the torsion angles of the two C-terminal Ala residues, Ala(23) and Ala(24), are (phi, psi) = (-66 degrees, -22 degrees ) and those of N-terminal two Ala residues, Ala(13) and Ala(14) are (phi, psi) = (-70 degrees, -30 degrees ). Thus, the local structure of N-terminal and C-terminal residues, and also the neighboring residues of alpha-helical poly-Ala chain in the model peptide is a more strongly wound structure than found in typical alpha-helix structures. PMID:12797796

Nakazawa, Yasumoto; Asakura, Tetsuo

2003-06-18

239

Expression system for production of bioactive compounds, recombinant human adiponectin, in the silk glands of transgenic silkworms.  

Science.gov (United States)

Adiponectin is an adipocyte hormone involved in glucose and lipid metabolism. The aim of this study was to develop a human adiponectin expression system in transgenic silkworm using a human adiponectin expression vector. The silk gland of the silkworm is a highly specialized organ that has the wonderful ability to synthesize and secrete silk protein. To express human adiponectin in the silk gland of transgenic silkworm, targeting vectors pB-A3-adiponectin-IRES-RFP and pB-Ser1-adiponectin-IRES-RFP were constructed and then introduced into the silkworm pupa. The transgenic silkworms were verified by PCR and then generated. The level of adiponectin in the transgenic silkworm was 6-10 ng/50 mg of freeze-dried powder, and western blotting using an antibody against human adiponectin demonstrated a specific band with a molecular weight of 30 kDa in the silkworm. These results showed that human adiponectin introduced into the silkworm genome was expressed successfully on a large-scale. PMID:24272890

Shin, Seulmee; Kim, Bong-Yoon; Jeon, Hyung-Yook; Lee, Aeri; Lee, Sungwon; Sung, Seung-Hyun; Park, Chan-Su; Lee, Chong-Kil; Kong, Hyunseok; Song, Youngcheon; Kim, Kyungjae

2014-05-01

240

Synthetic spider silk fibers spun from Pyriform Spidroin 2, a glue silk protein discovered in orb-weaving spider attachment discs.  

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Spider attachment disc silk fibers are spun into a viscous liquid that rapidly solidifies, gluing dragline silk fibers to substrates for locomotion or web construction. Here we report the identification and artificial spinning of a novel attachment disc glue silk fibroin, Pyriform Spidroin 2 (PySp2), from the golden orb weaver Nephila clavipes . MS studies support PySp2 is a constituent of the pyriform gland that is spun into attachment discs. Analysis of the PySp2 protein architecture reveals sequence divergence relative to the other silk family members, including the cob weaver glue silk fibroin PySp1. PySp2 contains internal block repeats that consist of two subrepeat units: one dominated by Ser, Gln, and Ala and the other Pro-rich. Artificial spinning of recombinant PySp2 truncations shows that the Ser-Gln-Ala-rich subrepeat is sufficient for the assembly of polymeric subunits and subsequent fiber formation. These studies support that both orb- and cob-weaving spiders have evolved highly polar block-repeat sequences with the ability to self-assemble into fibers, suggesting a strategy to allow fiber fabrication in the liquid environment of the attachment discs. PMID:21053953

Geurts, Paul; Zhao, Liang; Hsia, Yang; Gnesa, Eric; Tang, Simon; Jeffery, Felicia; La Mattina, Coby; Franz, Andreas; Larkin, Leah; Vierra, Craig

2010-12-13

 
 
 
 
241

Piriform spider silk sequences reveal unique repetitive elements.  

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Orb-weaving spider silk fibers are assembled from very large, highly repetitive proteins. The repeated segments contain, in turn, short, simple, and repetitive amino acid motifs that account for the physical and mechanical properties of the assembled fiber. Of the six orb-weaver silk fibroins, the piriform silk that makes the attachment discs, which lashes the joints of the web and attaches dragline silk to surfaces, has not been previously characterized. Piriform silk protein cDNAs were isolated from phage libraries of three species: A. trifasciata , N. clavipes , and N. cruentata . The deduced amino acid sequences from these genes revealed two new repetitive motifs: an alternating proline motif, where every other amino acid is proline, and a glutamine-rich motif of 6-8 amino acids. Similar to other spider silk proteins, the repeated segments are large (>200 amino acids) and highly homogenized within a species. There is also substantial sequence similarity across the genes from the three species, with particular conservation of the repetitive motifs. Northern blot analysis revealed that the mRNA is larger than 11 kb and is expressed exclusively in the piriform glands of the spider. Phylogenetic analysis of the C-terminal regions of the new proteins with published spidroins robustly shows that the piriform sequences form an ortholog group. PMID:20954740

Perry, David J; Bittencourt, Daniela; Siltberg-Liberles, Jessica; Rech, Elibio L; Lewis, Randolph V

2010-11-01

242

Assembly mechanism of recombinant spider silk proteins  

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Spider silk threads are formed by the irreversible aggregation of silk proteins in a spinning duct with dimensions of only a few micrometers. Here, we present a microfluidic device in which engineered and recombinantly produced spider dragline silk proteins eADF3 (engineered Araneus diadematus fibroin) and eADF4 are assembled into fibers. Our approach allows the direct observation and identification of the essential parameters of dragline silk assembly. Changes in ionic conditions and pH resu...

Rammensee, S.; Slotta, U.; Scheibel, T.; Bausch, A. R.

2008-01-01

243

Silk Coatings on PLGA and Alginate Microspheres for Protein Delivery  

Science.gov (United States)

Bombyx mori silk fibroin self-assembles on surfaces to form ultrathin nanoscale coatings based on our prior studies using layer-by-layer deposition techniques driven by hydrophobic interactions between silk fibroin protein molecules. In the present study, polylactic-co-glycolic acid (PLGA) and alginate microspheres were used as substrates and coated with silk fibroin. The coatings were visualized by confocal laser scanning microscopy using fluorescein-labeled silk fibroin. On PLGA microspheres the coating was ~1 ?m and discontinuous, reflecting the porous surface of these microspheres determined by SEM. In contrast, on alginate microspheres the coating was ~10 ?m thick and continuous. The silk fibroin penetrated into the alginate gel matrix. The silk coating on the PLGA microspheres delayed PLGA degradation. The silk coating on the alginate microspheres survived ethylenediamine tetraacetic acid (EDTA) treatment used to remove the Ca+2-cross-links in the alginate gels to solubilize the alginate. This suggests that alginate microspheres can be used as templates to form silk microcapsules. Horseradish peroxidase (HRP) and tetramethylrhodamine-conjugated bovine serum albumin (BSA) as model protein drugs were encapsulated in the PLGA and alginate microspheres with and without the silk fibroin coatings. Drug release was significantly retarded by the silk coatings when compared to uncoated microsphere controls, and was retarded further by methanol-treated silk coating when compared to silk water-based coatings on alginate microspheres. Silk coatings on PLGA and alginate microspheres provide mechanically stable shells as well as a diffusion barrier to the encapsulated protein drugs. This coating technique has potential for biosensor and drug delivery applications due to the aqueous process employed, the ability to control coating thickness and crystalline content, and the biocompatibility of the silk fibroin protein used in the process.

Wang, Xiaoqin; Wenk, Esther; Hu, Xiao; Castro, Guillermo R.; Meinel, Lorenz; Wang, Xianyan; Li, Chunmei; Merkle, Hans; Kaplan, David L.

2009-01-01

244

DNA supercoiling facilitates formation of the transcription initiation complex on the fibroin gene promoter.  

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Transcription of the Bombyx mori fibroin gene in a posterior silk gland extract can be separated into three functional steps on the basis of sensitivity to Sarkosyl: 1) formation of an initiation complex, which is blocked by 0.025% Sarkosyl; 2) conversion of the initiation complex to an elongation complex, a step sensitive to 0.05% Sarkosyl; 3) the subsequent elongation of RNA chain which occurs in the presence of 0.05% Sarkosyl. Whereas the last two steps are rapid and unaffected by template topology, the first step is slow and affected by DNA conformation. In the posterior silk gland extract, closed circular DNA forms a superhelical state and supports more rapid assembly of the initiation complex than linear DNA does. Both DNA supercoiling and rapid assembly of the initiation complex require ATP and are abolished by the addition of a topoisomerase II inhibitor VP16. These results suggest that DNA supercoiling enhances the fibroin gene transcription by facilitating formation of the initiation complex. PMID:2459121

Tabuchi, H; Hirose, S

1988-10-25

245

Fabrication and Biocompatibility of Electrospun Silk Biocomposites  

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Full Text Available 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.

Ick-Soo Kim

2011-10-01

246

Selection of silk-binding peptides by phage display  

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Peptides that bind to silkworm-derived silk fibroin fiber were selected from a phage-displayed random peptide library. The selected silk-binding peptides contained a consensus sequence QSWS which is important for silk-binding as confirmed by binding assays using phage and synthetic peptides. With further optimization, we anticipate that the silk-binding peptides will be useful for functionalization of silk for biomaterial applications.

2011-01-01

247

Piriform Spider Silk Sequences Reveal Unique Repetitive Elements  

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Orb-weaving spider silk fibers are assembled from very large, highly repetitive proteins. The repeated segments contain, in turn, short, simple repetitive amino acid motifs that account for the physical and mechanical properties of the assembled fiber. Of the six orb-weaver silk fibroins, the piriform silk that makes the attachment discs, which lashes the joints of the web and attaches dragline silk to surfaces has not been previously characterized. Piriform silk protein cDNAs were isolated f...

Perry, David J.; Bittencourt, Daniela; Siltberg-liberles, Jessica; Rech, Elibio L.; Lewis, Randolph V.

2010-01-01

248

Radiation degradation of silk protein  

International Nuclear Information System (INIS)

Silk fibroin fiber from the domesticated silkworm Bombyx mori was irradiated using an electron beam accelerator to investigate the application of the radiation degradation technique as a means to solubilize fibroin. The irradiation caused a significant degradation of the fiber. The tensile strength of fibroin fiber irradiated up to 2500 kGy decreased rapidly with increasing dose. The presence of oxygen in the irradiation atmosphere enhanced degradation of the tensile strength. The solubilization of irradiated fibroin fiber was evaluated using the following three kinds of solutions: a calcium chloride solution(CaCl2/C2H5OH/H2O=1:2:8 in mole ratio), a hydrochloric acid (0.5 N) and a distilled water. Dissolution of fibroin fiber into these solutions was significantly enhanced by irradiation. Especially, an appreciable amount of water soluble proteins was extracted by a distilled water. (author)

2000-03-01

249

Radiation degradation of silk protein  

International Nuclear Information System (INIS)

Silk fibroin fiber from the domesticated silkworm Bombyx mori was irradiated in the dose range up to 2500 kGy using an electron beam accelerator to apply the radiation degradation technique as a means to solubilize fibroin. The tensile strength of irradiated fibroin fiber decreased with increasing dose and the presence of oxygen in the irradiation atmosphere enhanced the degradation. The solubilization of irradiated fibroin fiber was evaluated using the following three kinds of solutions: calcium chloride solution (CaCl2/C2H5OH/H2O = 1 : 2 : 8 in mole ratio), hydrochloric acid (0.5N) and distilled water. Dissolution of fibroin fiber into these solutions was significantly enhanced by irradiation. Especially, an appreciable amount of water-soluble protein was extracted by distilled water. (author)

2000-09-01

250

Effect of Silk Protein Processing on Drug Delivery from Silk Films  

Science.gov (United States)

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.

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

2013-01-01

251

TATA-Binding Protein–TATA Interaction Is a Key Determinant of Differential Transcription of Silkworm Constitutive and Silk Gland-Specific tRNAAla Genes  

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We have investigated the contribution of specific TATA-binding protein (TBP)–TATA interactions to the promoter activity of a constitutively expressed silkworm tRNACAla gene and have also asked whether the lack of similar interactions accounts for the low promoter activity of a silk gland-specific tRNASGAla gene. We compared TBP binding, TFIIIB-promoter complex stability (measured by heparin resistance), and in vitro transcriptional activity in a series of mutant tRNACAla promoters and found...

2000-01-01

252

Engineering Adipose-like Tissue in vitro and in vivo Utilizing Human Bone Marrow and Adipose-derived Mesenchymal Stem Cells with Silk Fibroin 3D Scaffolds  

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Biomaterials derived from silk fibrion prepared by aqueous (AB) and organic (HFIP) solvent based processes, along with collagen (COL) and poly-lactic acid (PLA) based scaffolds were studied in vitro and in vivo for their utility in adipose tissue engineering strategies. For in vitro studies, human bone marrow and adipose-derived mesenchymal stem cells (hMSCs and hASCs) were seeded on the various biomaterials and cultured for 21 days in the presence of adipogenic stimulants (AD) or maintained ...

2007-01-01

253

Clay-Enriched Silk Biomaterials for Bone Formation  

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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 ...

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

2011-01-01

254

Effect of silk protein processing on drug delivery from silk films.  

Science.gov (United States)

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 a silk film by protease. Model compounds spanning a range of physical-chemical properties generally show 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

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

2013-03-01

255

Determination of the mutual orientation of the 15N and 13C NMR chemical shift tensors of 13- 15N double labeled model peptides for silk fibroin from the dipolar-coupled powder patterns  

Science.gov (United States)

The 15N and 13C chemical shift tensors, and the orientation of the principal axis system relative to the molecular symmetry axes were determined for 15N and 13C carbonyl carbon sites of 13C? 15N double labeled model peptides for Bombyx mori silk fibroin, that is, Boc-[1- 13C]Ala[ 15N]Gly-OMe, Boc-[1- 13C]Ala[ 15N]GlyAlaGly-OPac, Boc-AlaGly[1- 13C]Ala[ 15N]GlyAlaGly-OPac, Boc-[1- 13C]Gly[ 15N]AlaGlyAla-OPac, Boc-GlyAla[1- 13C]Gly[ 15N]AlaGlyAla-OPac and Boc-[1- 13C]Gly[ 15N]ValGlyAla-OPac, where Boc is t-butoxycarbonyl, OMe is methyl ester, OPac is phenacyl ester, Ala is alanine, Gly is glycine and Val is valine. From the comparisons of the 15N chemical shift tensors and the orientations of the principal axis system relative to the molecular symmetry axes among three compounds having [1- 13C]Ala[ 15N]Gly units, it is concluded that the intermolecular interactions such as hydrogen bonding are different between Boc-[1- 13C]Ala[ 15N]Gly-OMe and two compounds, Boc-[1- 13C]Ala[ 15N]GlyAlaGly-OPac and Boc-AlaGly[1- 13C]Ala[ 15N]GlyAlaGly-OPac although the latter two compounds have similar structures. A similar conclusion has also been obtained from the 13C chemical shift tensors of these compounds.

Asakura, Tetsuo; Yamazaki, Yasunobu; Seng, Koo Wey; Demura, Makoto

1998-05-01

256

Production of scFv-Conjugated Affinity Silk Powder by Transgenic Silkworm Technology  

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Bombyx mori (silkworm) silk proteins are being utilized as unique biomaterials for medical applications. Chemical modification or post-conjugation of bioactive ligands expand the applicability of silk proteins; however, the processes are elaborate and costly. In this study, we used transgenic silkworm technology to develop single-chain variable fragment (scFv)-conjugated silk fibroin. The cocoons of the transgenic silkworm contain fibroin L-chain linked with scFv as a fusion protein. After di...

Sato, Mitsuru; Kojima, Katsura; Sakuma, Chisato; Murakami, Maria; Aratani, Eriko; Takenouchi, Takato; Tamada, Yasushi; Kitani, Hiroshi

2012-01-01

257

Nongenomic and genomic actions of an insect steroid coordinately regulate programmed cell death of anterior silk glands of Bombyx mori  

Directory of Open Access Journals (Sweden)

Full Text Available The insect steroid hormone 20-hydroxyecdysone (20E induces programmed cell death of larva-specific tissues at pupal metamorphosis. In the silkworm Bombyx mori, the anterior silk gland undergoes cell death in response to the metamorphic peak titer of ecdysteroids in vivo and also to 20E in vitro. Although 20E elicits early gene activation, an additional 20E stimulus is required for completion of cell death. This additional stimulus involves caspase-3-like protease activation, indicating that 20E also acts through a nongenomic mechanism. Studies using various inhibitors, agonists, and antagonists have shown that cell condensation is under the control of 20E genomic action, and that 20E nongenomic action begins with 20E binding to the putative membrane-bound ecdysone receptor, which is probably a G-protein-coupled receptor. This step is followed by a signaling pathway comprising phospholipase C/inositol 3,4,5-triphosphate/Ca2+/protein kinase C/caspase-3-like protease, which induces DNA and nuclear fragmentation. Nuclear condensation is regulated by signaling of calmodulin/calmodulin-dependent protein kinase II (CaMKII, but CaMKII activation is independent of intracellular Ca2+ elevation. In addition, the genomic action of 20E is indispensable for driving its nongenomic action, indicating that crosstalk between genomic and nongenomic action plays a significant role in 20E-induced cell death.

S Sakurai

2008-02-01

258

Synthesis and characterization of chimeric silkworm silk.  

Science.gov (United States)

A synthetic gene encoding a chimeric silklike protein was constructed that combined a polyalanine encoding region (Ala)(18), a sequence slightly longer than the (Ala)(12-13) found in the silk fibroin from the wild silkworm Samia cynthia ricini, and a sequence encoding GVGAGYGAGAGYGVGAGYGAGVGYGAGAGY, found in the silk fibroin from the silkworm Bombyx mori. A tetramer of the chimeric repeat sequence encoding a approximately 29 kDa protein was expressed as a fusion protein in Escherichia coli. In comparison to S. c. ricini silk, the chimeric protein demonstrated improved solubility because it could be dissolved in 8 M urea. The purified protein assumed an alpha-helical structure based on solid-state (13)C CP/MAS NMR and was less prone to conformational transition to a beta-sheet, unlike native silk proteins from S. c. ricini. Model peptides representing the crystalline region of S. c. ricini silk fibroin, (Ala)(12) and (Ala)(18), formed beta-sheet structures. Therefore, the solubility and structural transitions of the chimeric protein were significantly altered through the formation of this chimeric silk. This experimental strategy to the study of silk structure and function can be used to develop an improved understanding of the contributions of protein domains in repetitive silkworm and spider silk sequences to structure development and structural transitions. PMID:12741803

Asakura, Tetsuo; Nitta, Koji; Yang, Mingying; Yao, Juming; Nakazawa, Yasumoto; Kaplan, David L

2003-01-01

259

Intrinsically Functionalized Silk (Bombyx Mori)  

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The goal of the thesis is to incorporate materials with either fluorescent or conductive properties in silk fibers, by feeding silkworms with a diet containing these materials. To achieve this, one would have to breed (rear) silkworm from eggs into larvaes, then to feed the silkworms with this special diet containing fluorescent or conductive materials. Samples of silk were then collected either from spun cocoons or via removing the silk producing organs (silk glands) from the silkworms via d...

A?berg, Gabriel

2013-01-01

260

Production of fine powder from silk by radiation  

Energy Technology Data Exchange (ETDEWEB)

To produce silk fine powder, silk fibroin fibers were irradiated with an accelerated electron beam. Though unirradiated silk fibers were not pulverized at all, irradiated silk fibers were able to be crushed only by physical means using a ball mill. In the dose range of 250- 1000 kGy, as the dose to silk fibroin increased, the conversion rate from fiber to powder was increased. The electron microscope observation showed that the particle size of silk powder was less than 10 micrometers, and that those particles composed aggregate. Although silk fibroin fiber was not soluble in water, silk powders from fiber irradiated 500 kGy dissolved more than 50 weight %. Amino acid analysis of soluble fraction in silk powder showed that Gly content is lower than in normal silk and other amino acids contents except for Ala are 1.5 times as large as in normal silk. From these results, it is reasonable to suppose that irradiation and pulverization decomposed Gly and Ala. (author)

Takeshita, Hidefumi; Yoshii, Fumio; Kume, Tamikazu [Japan Atomic Energy Research Inst., Takasaki, Gunma (Japan). Takasaki Radiation Chemistry Research Establishment; Ishida, Kazushige; Kamiishi, Youichi [Textile Research Inst. of Gunma, Kiryu, Gunma (Japan)

2000-09-01

 
 
 
 
261

Negative supercoiling of DNA facilitates an interaction between transcription factor IID and the fibroin gene promoter  

Energy Technology Data Exchange (ETDEWEB)

Transcription of the fibroin gene can be reconstituted with partially purified components from HeLa cells. Transcription factors IIB, IID, and IIE and RNA polymerase II are required for accurate initiation of transcription. Linear and relaxed closed circular DNA show a similar level of template activity. However, transcription of closed circular DNA is stimulated when negative supercoils are introduced by the addition of DNA topoisomerase II and supercoiling factor purified from the posterior silk gland of Bombyx mori. Dissection of transcription into pre- and postinitiation steps by the use of Sarkosyl reveals that DNA supercoiling promotes formation of a preinitiation complex. Furthermore, order of addition experiements suggest that DNA supercoiling facilitates a functional binding of transcription factor IID to the promoter.

Mizutani, Mitsuko; Ohta, Tsutomu; Hirose, Susumu (National Inst. of Genetics, Mishima (Japan)); Watanabe, Hajime; Handa, Hiroshi (Univ. of Tokyo (Japan))

1991-02-01

262

Characteristics of Silk Fiber with and without Sericin Component: A Comparison between Bombyx mori and Philosamia ricini Silks  

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Full Text Available The study aimed to investigate and compare some characteristics of Bombyx mori and Eri (Philosamia ricini silks in different forms; with and without sericin. The protein contents were measured and find out the composition of the silk fibroin and sericin proteins by Lowry method. The secondary structure and thermal behavior of all kind of silk were determined by FT-IR and TA instrument, respectively. The B. mori composed of more amount of sericin content than that of Eri silk. FT-IR spectra indicated that the Eri silk was similar profile of silk with and without sericin, whereas B. mori silk showed dramatically differed. With sericin, B. mori composed of higher ratio random coil and ?-helix structures than ?-structure. With thermogravimetric analysis, both B. mori and Eri silk fibers without sericin showed higher stability than that silk fiber with sericin. This is due to the crystalline region of hydrophobic amino acid composed in the fibroin core protein. The differential scanning calorimetry thermogram of B. mori was differed from Eri silk fiber. It is a promising that characteristics of the silk were influenced by both silk components and silk varieties.

S. Prasong

2009-01-01

263

Conformation transition kinetics of Bombyx mori silk protein.  

Science.gov (United States)

Time-resolved FTIR analysis was used to monitor the conformation transition induced by treating regenerated Bombyx mori silk fibroin films and solutions with different concentrations of ethanol. The resulting curves showing the kinetics of the transition for both films and fibroin solutions were influenced by the ethanol concentration. In addition, for silk fibroin solutions the protein concentration also had an effect on the kinetics. At low ethanol concentrations (for example, less than 40% v/v in the case of film), films and fibroin solutions showed a phase in which beta-sheets slowly formed at a rate dependent on the ethanol concentration. Reducing the concentration of the fibroin in solutions also slowed the formation of beta-sheets. These observations suggest that this phase represents a nucleation step. Such a nucleation phase was not seen in the conformation transition at ethanol concentrations > 40% in films or > 50% in silk fibroin solutions. Our results indicate that the ethanol-induced conformation transition of silk fibroin in films and solutions is a three-phase process. The first phase is the initiation of beta-sheet structure (nucleation), the second is a fast phase of beta-sheet growth while the third phase represents a slow perfection of previously formed beta-sheet structure. The nucleation step can be very fast or relatively slow, depending on factors that influence protein chain mobility and intermolecular hydrogen bond formation. The findings give support to the previous evidence that natural silk spinning in silkworms is nucleation-dependent, and that silkworms (like spiders) use concentrated silk protein solutions, and careful control of the pH value and metallic ion content of the processing environment to speed up the nucleation step to produce a rapid conformation transition to convert the water soluble spinning dope to a tough solid silk fiber. PMID:17436322

Chen, Xin; Shao, Zhengzhong; Knight, David P; Vollrath, Fritz

2007-07-01

264

Structure of a protein superfiber: spider dragline silk.  

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Spider major ampullate (dragline) silk is an extracellular fibrous protein with unique characteristics of strength and elasticity. The silk fiber has been proposed to consist of pseudocrystalline regions of antiparallel beta-sheet interspersed with elastic amorphous segments. The repetitive sequence of a fibroin protein from major ampullate silk of the spider Nephila clavipes was determined from a partial cDNA clone. The repeating unit is a maximum of 34 amino acids long and is not rigidly co...

1990-01-01

265

Tough silk fibers prepared in air using a biomimetic microfluidic chip.  

Science.gov (United States)

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. PMID:24613677

Luo, Jie; Zhang, Lele; Peng, Qingfa; Sun, Mengjie; Zhang, Yaopeng; Shao, Huili; Hu, Xuechao

2014-05-01

266

Influence of antioxidants on thermal properties of fibroins of natural polymer  

International Nuclear Information System (INIS)

Full Text: The termophysical properties of fibroins of natural polymer are investigated. It is established, that at injecting of antioxidants N-(3,5-ditret-buty-4-hydroxyfenil) salicylaldimin into the structure of fibroins, by additional fertilizing of caterpillars by last-named amorphous sites increase and thus the stability of silk to external influences raises, i.e. the stabilization occurs due to sewing action of antioxidant

2002-02-01

267

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

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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.

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

2006-02-01

268

Vibrational spectroscopic study of sulphated silk proteins  

Science.gov (United States)

Degummed Bombyx mori ( B. m.) silk fibroin fabric and mutant naked pupa cocoons (Nd-s) consisting of almost pure silk sericin were treated with chlorosulphonic acid in pyridine and investigated by FT-IR and FT-Raman spectroscopies. Untreated silk fibroin and sericin displayed typical spectral features due to characteristic amino acid composition and molecular conformation (prevailing ?-sheet with a less ordered structure in sericin). Upon sulphation, the degree of molecular disorder increased in both proteins and new bands appeared. The IR bands at 1049 and 1014 cm -1 were attributed to vibrations of sulphate salts and that at 1385 cm -1 to the ?asSO 2 mode of organic covalent sulphates. In the 1300-1180 cm -1 range various contributions of alkyl and aryl sulphate salts, sulphonamides, sulphoamines and organic covalent sulphates, fell. Fibroin covalently bound sulphate groups through the hydroxyl groups of tyrosine and serine, while sericin through the hydroxyl groups of serine, since the ?OH vibrations at 1399 cm -1 in IR and at 1408 cm -1 in Raman disappeared almost completely. Finally, the increase of the I850/ I830 intensity ratio of Raman tyrosine doublet in fibroin suggested a change towards a more exposed state of tyrosine residues, in good agreement with the more disordered conformation taken upon sulphation.

Monti, P.; Freddi, G.; Arosio, C.; Tsukada, M.; Arai, T.; Taddei, P.

2007-05-01

269

Bio-inspired fabrication of fibroin cryogels from the muga silkworm Antheraea assamensis for liver tissue engineering.  

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Conventional scaffold fabrication techniques result in narrow pore architectures causing a limited interconnectivity and use of porogens, which affects the bio- or cyto-compatibility. To ameliorate this, cryogels are immensely explored due to their macro-porous nature, ease in fabrication, using ice crystals as porogens, the shape property, easy reproducibility and cost-effective fabrication technique. Cryogels in the present study are prepared from nonmulberry Indian muga silk gland protein fibroin of Antheraea assamensis using two different fabrication temperatures (-20 and -80 °C). Anionic surfactant sodium dodecyl sulfate is used to solubilize fibroin, which in turn facilitates gelation by accelerating the ß-sheet formation. Ethanol is employed to stabilize the 3D network and induces bimodal porosity. The gels thus formed demonstrate increased ß-sheet content (FTIR) and a considerable effect of pre-freezing temperatures on 3D micro-architectures. The cryogels are capable of absorbing large amounts of water and withstanding mechanical compression without structure deformation. Further, cell impregnated cryogels well support the viability of human hepatocarcinoma cells (live/dead assay). The formation of cellular aggregates (confocal laser and scanning electron microscope), derivation in metabolic activity and proliferation rate are obtained in constructs fabricated at different temperatures. In summary, the present work reveals promising insights in the development of a biomimetic functional template for biomedical therapeutics and liver tissue engineering. PMID:24002731

Kundu, Banani; Kundu, S C

2013-10-01

270

Fluid Mechanical Properties of Silkworm Fibroin Solutions  

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The aqueous solution behavior of silk fibroin is of interest due to the assembly and processing of this protein related to the spinning of protein fibers that exhibit remarkable mechanical properties. To gain insight into the origins of this functional feature, it is desired to determine how the protein behaves under a range of solution conditions. Pure fibroin at different concentrations in water was studied for surface tension, as a measure of surfactancy. In addition, shear induced changes on these solutions in terms of structure and morphology was also determined. Fibroin solutions exhibited shear rate-sensitive viscosity changes and precipitated at a critical shear rate where a dramatic increase of 75-150% of the initial value was observed along with a decrease in viscosity. In surface tension measurements, critical micelle concentrations were in the range of 3-4% w/v. The influence of additional factors, such as sericin protein, divalent and monovalent cations, and pH on the solution behavior in relation to structural and morphological features will also be described.

Matsumoto, Akira

2005-11-01

271

Solubilization of silk protein by radiation  

International Nuclear Information System (INIS)

Gamma irradiated silk fibroin at doses of 0, 5, 10, 20, 40, 60, 80, 100, 125, 250, 500, 750 and 1000 kGy were soaked in water for 1 hr. Silk fibroin solubilized percentage was investigated from lost weight of sample (dried at 1050C), they were 0, 0, 0.7, 0, 0.11, 0.11, 0, 0.73, 0.77, 4.38, 8.32, 10.22 and 18.52 respectively. It showed that at the higher dose up to 250 kGy had direct effect to solubility, and increased with increasing dose. In addition, silk sericin dissolved 77.76, 82.22, 83.55, 84.31, 86.04, 86.67 and 87.37% after gamma irradiation at the doses of 0, 50, 100, 200, 500, 750 and 1000 kGy respectively. It presents that radiation can cause silk protein, fibroin and sericin dissolve because of their degradation. (author)

2002-03-01

272

Silk-pectin hydrogel with superior mechanical properties, biodegradability, and biocompatibility.  

Science.gov (United States)

A new method is developed to prepare silk hydrogels and silk-pectin hydrogels via dialysis against methanol to obtain hydrogels with high concentrations of silk fibroin. The relationship between the mechanical and biological properties and the structure of the silk-pectin hydrogels is subsequently evaluated. The present results suggest that pectin associates with silk molecules when the silk concentration exceeds 15?wt%, suggesting that a silk concentration of over 15?wt% is critical to construct interacting silk-pectin networks. The silk-pectin hydrogel reported here is composed of a heterogeneous network, which is different from fiber-reinforced, interpenetrated networks and double-network hydrogels, as well as high-stiffness hydrogels (elastic modulus of 4.7?±?0.9?MPa, elastic stress limit of 3.9?±?0.1?MPa, and elastic strain limit of 48.4?±?0.5%) with regard to biocompatibility and biodegradability. PMID:24610718

Numata, Keiji; Yamazaki, Shoya; Katashima, Takuya; Chuah, Jo-Ann; Naga, Naofumi; Sakai, Takamasa

2014-06-01

273

Effect of ?-sheet crystalline content on mass transfer in silk films  

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The material properties of silk are favorable for drug delivery due to the ability to control material structure and morphology under ambient, aqueous processing conditions. Mass transport of compounds with varying physical-chemical characteristics was studied in silk fibroin films with control of ?-sheet crystalline content. Two compounds, vitamin B12 and fluorescein isothiocynate (FITC) labeled lysozyme were studied in a diffusion apparatus to determine transport through silk films. The fi...

2011-01-01

274

De novo bone formation on macro/microporous silk and silk/nano-sized calcium phosphate scaffolds  

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Macro/micro porous silk/nano-sized calcium phosphate scaffolds (SC16) with bioactive and superior physicochemical properties have been recently developed. In this study, we aim at evaluating the new bone formation ability of the SC 16 scaffolds in vivo, using silk fibroin scaffolds (S16) as control. The CaP distribution profile in the scaffolds was characterized by Micro-Computed Tomography. The in vitro mineralization behavior was examined by immersion in Simulated Body Fluid solution from 1...

Yan, Leping; Salgado, A. J.; Oliveira, J. M.; Oliveira, A. L.; Reis, R. L.

2013-01-01

275

Characterization by Raman microspectroscopy of the strain-induced conformational transition in fibroin fibers from the silkworm Samia cynthia ricini.  

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Raman microspectroscopy has been used to quantitatively study the effect of a mechanical deformation on the conformation and orientation of Samia cynthia ricini (S. c. ricini) silk fibroin. Samples were obtained from the aqueous solution stored in the silk gland and stretched at draw ratios (lambda) ranging from 0 to 11. Using an appropriate band decomposition procedure, polarized and orientation-insensitive spectra have been analyzed to determine order parameters and the content of secondary structures, respectively. The data unambiguously show that, in response to mechanical deformation, S. c. ricini fibroin undergoes a cooperative alpha-helix to beta-sheet conformational transition above a critical draw ratio of 4. The alpha-helix content decreases from 33 to 13% when lambda increases from 0 to 11, while the amount of beta-sheets increases from 15 to 37%. In comparison, cocoon silk is devoid of alpha-helical structure and always contains a larger amount of beta-sheets. Although the presence of isosbestic points in different spectral regions reveals that the conformational change induced by mechanical deformation is a two-state process, our results suggest that part of the glycine residues might be incorporated into beta-poly(alanine) structures. The beta-sheets are initially isotropically distributed and orient along the fiber axis as lambda increases, but do not reach the high level of orientation found in the cocoon fiber. The increase in the orientation level of the beta-sheets is found to be concomitant with the alpha --> beta conformational conversion, whereas alpha-helices do not orient under the applied strain but are rather readily converted into beta-sheets. The components assigned to turns exhibit a small orientation perpendicular to the fiber axis in stretched samples, showing that, overall, the polypeptide chains are aligned along the stretching direction. Our results suggest that, in nature, factors other than stretching contribute to the optimization of the amount of beta-sheets and the high degree of orientation found in natural cocoon silk. PMID:16961312

Rousseau, Marie-Eve; Beaulieu, Lilyane; Lefèvre, Thierry; Paradis, Joanie; Asakura, Tetsuo; Pézolet, Michel

2006-09-01

276

TATA-Binding Protein-TATA Interaction Is a Key Determinant of Differential Transcription of Silkworm Constitutive and Silk Gland-Specific tRNAAla Genes  

Science.gov (United States)

We have investigated the contribution of specific TATA-binding protein (TBP)–TATA interactions to the promoter activity of a constitutively expressed silkworm tRNACAla gene and have also asked whether the lack of similar interactions accounts for the low promoter activity of a silk gland-specific tRNASGAla gene. We compared TBP binding, TFIIIB-promoter complex stability (measured by heparin resistance), and in vitro transcriptional activity in a series of mutant tRNACAla promoters and found that specific TBP-TATA contacts are important for TFIIIB-promoter interaction and for transcriptional activity. Although the wild-type tRNACAla promoter contains two functional TBP binding sequences that overlap, the tRNASGAla promoter lacks any TBP binding site in the corresponding region. This feature appears to account for the inefficiency of the tRNASGAla promoter since provision of either of the wild-type TATA sequences derived from the tRNACAla promoter confers robust transcriptional activity. Transcriptional impairment of the wild-type tRNASGAla gene is not due to reduced incorporation of TBP into transcription complexes since both the tRNACAla and tRNASGAla promoters form transcription complexes that contain the same amount of TBP. Thus, the deleterious consequences of the lack of appropriate TBP-TATA contacts in the tRNASGAla promoter must come from failure to incorporate some other essential transcription factor(s) or to stabilize the complete complex in an active conformation.

Ouyang, Ching; Martinez, M. Juanita; Young, Lisa S.; Sprague, Karen U.

2000-01-01

277

Self-assembly in the major ampullate gland of Nephila clavipes  

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We present a tentative interpretation of the origin of nematic liquid crystalline order exhibited by dragline silk fibroin solutions collected from the spider Nephila clavipes. Liquid crystallinity is thought to confer certain rheological properties on the fibroin solution which are exploited during the dragline spinning process. We show that the feasibility of liquid crystallinity under physiological conditions depends critically on parameters characterising the amino-acid sequence of the fibroin molecules.

Braun, F N

2002-01-01

278

Effect of degumming condition on the solution properties and electrospinnablity of regenerated silk solution.  

Science.gov (United States)

The application of silk on tissue engineering scaffolds has been studied intensively because silk has an electrospinning technique using a good blood compatibility, excellent cytocompatibility and biodegradability. Silk consists of two polymers, fibroin and sericin. In spite of importance of sericin, most studies were focused on the fibroin only and the effect of residual sericin on the electrospinning performance of silk has not been considered. In this study, regenerated silk with different residual sericin contents was prepared by controlling the degumming conditions. The effects of the degumming conditions on the solution properties and electrospinning performance of silk were examined. The fast protein liquid chromatography (FPLC) measurements confirmed that the molecular weight of the regenerated silk decreased slightly with increasing residual sericin content. More molecular aggregation of silk occurred with increasing sericin content, resulting in an increase in the solution turbidity of formic acid. All silk formic acid solutions exhibited almost Newtonian fluid behavior and the viscosity increased with increasing sericin content. Interestingly, the dope solution viscosity of silk increased remarkably at sericin contents 25%) leading to significant improvements in electrospinnability and an increase in the fiber diameter of the silk web. PMID:23295206

Ko, Jae Sang; Yoon, Kyunghwan; Ki, Chang Seok; Kim, Hyun Ju; Bae, Do Gyu; Lee, Ki Hoon; Park, Young Hwan; Um, In Chul

2013-04-01

279

Construction of a binary transgenic gene expression system for recombinant protein production in the middle silk gland of the silkworm Bombyx mori.  

Science.gov (United States)

To construct an efficient system for the production of recombinant proteins in silkworm (Bombyx mori), we investigated the promoter activity of the silkworm sericin 1, 2, and 3 genes (Ser1, Ser2, and Ser3) using a GAL4/UAS binary gene expression system in transgenic silkworm. The promoter activity of the upstream region of Ser1 was strong, yielding high expression of an enhanced green fluorescent protein (EGFP) transgene in the middle and posterior regions of the middle silk gland (MSG) after day 2 of the fifth instar. The Ser3 upstream region exhibited moderate promoter activity in the anterior MSG, but the Ser2 upstream region did not exhibit any promoter activity. Since the strongest promoter activity was observed for Ser1, we devised a system for the production of recombinant proteins using a GAL4-Ser1 promoter construct (Ser1-GAL4). Transgenic silkworms harboring both the Ser1-GAL4 construct and the previously reported upstream activating sequence (UAS)-EGFP construct, which contains the TATA box region of the Drosophila hsp70 gene, yielded approximately 100 microg EGFP per larva. When we then analyzed the TATA box region, signal peptide, and intron sequences for their effects on production from the UAS-EGFP construct, we found that the optimization of these sequences effectively increased production to an average of 500 microg EGFP protein per transgenic larva. We conclude that this binary system is a useful tool for the mass production of recombinant proteins of biomedical and pharmaceutical interest in silkworm. PMID:19789990

Tatematsu, Ken-ichiro; Kobayashi, Isao; Uchino, Keiro; Sezutsu, Hideki; Iizuka, Tetsuya; Yonemura, Naoyuki; Tamura, Toshiki

2010-06-01

280

SPIDER SILK  

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Full Text Available The strengthness and toughness of spider fiber and its multifunctional nature is only surpassed in some cases by synthetic high performance fibers. In the world of natural fibers, spider silk has been long time recognized as a wonder fiber for its unique combination of high strength and rupture elongation. Scientists in civil military engineering reveal that the power of biological material (spider silk lies in the geometric configuration of structural protein, and the small cluster of week hydrogen bonds that works together to resist force and dissipate energy. Each spider and each type of silk has a set of mechanical properties optimized for their biological function. Most silks, in particular deagline silk, have exceptional mechanical properties. They exhibit a unique combination of high tensile strength and extensibility (ductility. This enables a silk fiber to absorb a lot of energy before breaking (toughness, the area under a stress- strain curve. A frequent mistake made in the mainstream media is to confuse strength and toughness when comparing silk to other materials. As shown below in detail, weight for weight, silk is stronger than steel, but not as strong as Kevlar. Silk is,however, tougher than both.This paper inform about overview on the today trend in the world of spider silk.

PORAV Viorica

2014-05-01

 
 
 
 
281

Differential Scanning Fluorimetry provides high throughput data on silk protein transitions.  

Science.gov (United States)

Here we present a set of measurements using Differential Scanning Fluorimetry (DSF) as an inexpensive, high throughput screening method to investigate the folding of silk protein molecules as they abandon their first native melt conformation, dehydrate and denature into their final solid filament conformation. Our first data and analyses comparing silks from spiders, mulberry and wild silkworms as well as reconstituted ‘silk' fibroin show that DSF can provide valuable insights into details of silk denaturation processes that might be active during spinning. We conclude that this technique and technology offers a powerful and novel tool to analyse silk protein transitions in detail by allowing many changes to the silk solutions to be tested rapidly with microliter scale sample sizes. Such transition mechanisms will lead to important generic insights into the folding patterns not only of silks but also of other fibrous protein (bio)polymers.

Vollrath, Fritz; Hawkins, Nick; Porter, David; Holland, Chris; Boulet-Audet, Maxime

2014-01-01

282

Differential Scanning Fluorimetry provides high throughput data on silk protein transitions.  

Science.gov (United States)

Here we present a set of measurements using Differential Scanning Fluorimetry (DSF) as an inexpensive, high throughput screening method to investigate the folding of silk protein molecules as they abandon their first native melt conformation, dehydrate and denature into their final solid filament conformation. Our first data and analyses comparing silks from spiders, mulberry and wild silkworms as well as reconstituted 'silk' fibroin show that DSF can provide valuable insights into details of silk denaturation processes that might be active during spinning. We conclude that this technique and technology offers a powerful and novel tool to analyse silk protein transitions in detail by allowing many changes to the silk solutions to be tested rapidly with microliter scale sample sizes. Such transition mechanisms will lead to important generic insights into the folding patterns not only of silks but also of other fibrous protein (bio)polymers. PMID:25004800

Vollrath, Fritz; Hawkins, Nick; Porter, David; Holland, Chris; Boulet-Audet, Maxime

2014-01-01

283

The natural silk spinning process. A nucleation-dependent aggregation mechanism?  

Science.gov (United States)

The spinning mechanism of natural silk has been an open issue. In this study, both the conformation transition from random coil to beta sheet and the beta sheet aggregation growth of silk fibroin are identified in the B. mori regenerated silk fibroin aqueous solution by circular dichroism (CD) spectroscopy. A nucleation-dependent aggregation mechanism, similar to that found in prion protein, amyloid beta (Abeta) protein, and alpha-synuclein protein with the conformation transition from a soluble protein to a neurotoxic, insoluble beta sheet containing aggregate, is a novel suggestion for the silk spinning process. We present evidence that two steps are involved in this mechanism: (a) nucleation, a rate-limiting step involving the conversion of the soluble random coil to insoluble beta sheet and subsequently a series of thermodynamically unfavorable association of beta sheet unit, i.e. the formation of a nucleus or seed; (b) once the nucleus forms, further growth of the beta sheet unit becomes thermodynamically favorable, resulting a rapid extension of beta sheet aggregation. The aggregation growth follows a first order kinetic process with respect to the random coil fibroin concentration. The increase of temperature accelerates the beta sheet aggregation growth if the beta sheet seed is introduced into the random coil fibroin solution. This work enhances our understanding of the natural silk spinning process in vivo. PMID:11737214

Li, G; Zhou, P; Shao, Z; Xie, X; Chen, X; Wang, H; Chunyu, L; Yu, T

2001-12-01

284

Light can transform the secondary structure of silk protein  

Science.gov (United States)

Fibroin is the main component of silk and is expected to be used as a novel functional material in medicine and bioelectronics. The main secondary structures of this protein are of the random-coil and the ?-sheet types. In this study, we carried out laser-induced transformation of the secondary structure, from the random-coil type to the ?-sheettype, in solid fibroin films. We prepared two types of fibroin films with the random-coil structure. One is a fibroin film doped with a dye as a photosensitizer with a small amount (1 wt%), and the other is a neat fibroin film. The former was excited at 532 nm and the latter was excited at 266 nm. Irradiations were carried out with fluences much lower than each ablation threshold. The excitation of the dye at 532 nm did not affect the secondary structure of the random-coil type. By contrast, 266-nm laser irradiation, which excites tryptophan (an amino-acid residue) involved in fibroin, created the ?-sheetdomain in the film. The structural transformation was revealed by infrared absorption spectroscopy and atomic force microscopy.

Tsuboi, Y.; Ikejiri, T.; Shiga, S.; Yamada, K.; Itaya, A.

285

Silk-elastinlike protein polymers for matrix-mediated cancer gene therapy.  

Science.gov (United States)

Silk-elastinlike protein polymers (SELPs) are recombinant polymers designed from silk fibroin and mammalian elastin amino acid repeats. These are versatile materials that have been examined as controlled release systems for intratumoral gene delivery. SELP hydrogels comprise monodisperse and tunable polymers that have the capability to control and localize the release and expression of plasmid DNA and viruses. This article reviews recent developments in the synthesis and characterization of SELP hydrogels and their use for matrix-mediated gene delivery. PMID:20430059

Gustafson, Joshua A; Ghandehari, Hamidreza

2010-12-30

286

Integration of silk protein in organic and light-emitting transistors  

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We present the integration of a natural protein into electronic and optoelectronic devices by using silk fibroin as a thin film dielectric in an organic thin film field-effect transistor (OFET) ad an organic light emitting transistor device (OLET) structures. Both n- (perylene) and p-type (thiophene) silk-based OFETs are demonstrated. The measured electrical characteristics are in agreement with high-efficiency standard organic transistors, namely charge mobility of the order of 10-2 cm2/Vs a...

2011-01-01

287

Controlled Release from Multilayer Silk Biomaterial Coatings to Modulate Vascular Cell Responses  

Digital Repository Infrastructure Vision for European Research (DRIVER)

A multilayered silk fibroin protein coating system was employed as a drug carrier and delivery system to evaluate vascular cell responses to heparin, paclitaxel, and clopidogrel. The results demonstrated that the silk coating system was an effective system for drug-eluting coatings, such as for stent applications, based on its useful micromechanical properties and biological outcomes. Cell attachment and viability studies with human aortic endothelial cells (HAECs) and human coronary artery s...

Wang, Xianyan; Zhang, Xiaohui; Castellot, John; Herman, Ira; Iafrati, Mark; Kaplan, David L.

2008-01-01

288

Nanocomposite gold-silk nanofibers  

Science.gov (United States)

Cell-biomaterial interactions can be controlled by modifying the surface chemistry or nanotopography of the material, to induce cell proliferation and differentiation if desired. Here we combine both approaches in forming silk nanofibers (SNFs) containing gold nanoparticles (AuNPs) and subsequently chemically modifying the fibers. Silk fibroin mixed with gold seed nanoparticles was electrospun to form SNFs doped with gold seed nanoparticles (SNFseed). Following gold reduction, there was a two-fold increase in particle diameter confirmed by the appearance of a strong absorption peak at 525 nm. AuNPs were dispersed throughout the AuNP-doped silk nanofibers (SNFsAu). The Young’s modulus of the SNFsAu was almost 70% higher than that of SNFs. SNFsAu were modified with the arginine-glycine-aspartic acid (RGD) peptide. Human mesenchymal stem cells that were cultured on RGD-modified SNFAu had a more than two-fold larger cell area compared to the cells cultured on bare SNFs; SNFAu also increase cell size. We suggest that this approach can be used to alter the cell-material interface in tissue engineering and other applications.

Cohen-Karni, Tzahi; Jeong, Kyung Jae; Tsui, Jonathan H.; Reznor, Gally; Mustata, Mirela; Wanunu, Meni; Graham, Adam; Marks, Carolyn; Bell, David C.; Langer, Robert S.; Kohane, Daniel S.

2012-01-01

289

Moisture sorption as a potential condition marker for historic silks: noninvasive determination by near-infrared spectroscopy.  

Science.gov (United States)

Given their ephemeral nature, the preservation of historic silks can be problematic. Rapid, on-site condition monitoring would offer significant benefits to conservators and museum curators concerned with continued access to collections. In this paper, near-infrared spectroscopy (NIR) is investigated as a noninvasive approach to the characterization of silk fabrics and particularly for determining the moisture content of silks as a potential age-related marker. Bands within the NIR spectrum of silk are assigned to contributions from water and the silk fibroin polymer. The water bands may be deconvolved to show separate contributions from bound and structural water. When silk is exposed to deuterium oxide, the water OH NIR bands are rapidly lost. The accompanying changes in the amide-related NIR absorptions reflect differential accessibility of regions within the semi-crystalline fibroin aggregate. NIR spectra were recorded while silk was maintained at a range of relative humidity; complementary gravimetry provided absolute reference data for moisture sorption. A single spectral parameter, the intensity of the water combination band, is sufficient to indicate the relative moisture content of silk and allows distinction of unaged and heat, light, and humidity aged silks. The results confirm that NIR has significant potential for on-site studies at collections in support of the preservation and access of our silk heritage. PMID:17331315

Zhang, Xiaomei; Wyeth, Paul

2007-02-01

290

Isolation of the smallest component of silk protein.  

Science.gov (United States)

Silk proteins were solubilized from cocoons with ethylenediamine/cupric hydroxide solution. A series of polymers of the smallest component, detected by polyacrylamide-gel electrophoresis, could be converted into the smallest component by reduction and aminoethylation. Fibroin and sericin fractions were separated by precipitation of sericin at pH 5.5. On gel electrophoresis, sericin showed distinct bands but fibroin did not. The components of fibroin and sericin were fractionated by gel filtration on Sepharose 6B. The smallest component in the sericin fraction was purified by rechromatography and showed a single band on gel electrophoresis. Its mol. wt. was 24 000, and its amino acid composition was determined. Images Fig. 1. Fig. 2. Fig. 4.

Tokutake, S

1980-01-01

291

TOWARDS A ROUTINE METHODOLOGY FOR ASSESSING THE CONDITION OF HISTORIC SILK  

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Full Text Available To ensure continued access and long-term preser vation itis essential to understand the condition of an artefact, andthe behaviour of its component materials, in order to setlimits on display and handling. In support of curators andconservators of collections with historic textiles, we aredeveloping rapid and routine technology, involving nearinfrared spectroscopy coupled with multivariate analysis,which, when applied non-invasively on-site, will allow theestimation of the state of deterioration of silk fabrics.Pursuing this aim, we have now carried out further analyticalstudies on silks artificially aged under different regimesfor up to 20 days: dry thermal, high heat and humidity, andsunlight equivalent irradiation. The tensile strengths, yellownessindices, apparent molecular weights of fibroin, andnear infrared spectral sorbed moisture parameters for theaged silks were determined. The mechanical performanceof silk diminished exponentially over each ageing timecourse. For the various silks, the increases in yellownessindices and decreases in fibroin molecular weights andmoisture contents showed similar kinetics to the mechanicalchanges. While each parameter was correlated with thetensile strength, silks exposed to the different acceleratedageing factors exhibited diverse correlations. It is concludedthat, when formulating a general model for characterisingthe condition of silk using chemometrics, the referenceset should include examples of silks which have beenexposed to the full variety of ageing factors.

Jeongjin Kim

2009-01-01

292

Biomaterials Derived from Silk-Tropoelastin Protein Systems  

Science.gov (United States)

A structural protein blend system based on silkworm silk fibroin and recombinant human tropoelastin is described. Silk fibroin, a semicrystalline fibrous protein with beta-sheet crystals provides mechanical strength and controllable biodegradation, while tropoelastin, a noncrystallizable elastic protein provides elasticity. Differential scanning calorimetry (DSC) and temperature modulated DSC (TMDSC) indicated that silk becomes miscible with tropoelastin at different blend ratios, without macrophase separation. Fourier transform infrared spectroscopy (FTIR) revealed secondary structural changes of the blend system (beta-sheet content) before and after methanol treatment. Atomic Force Microscopy (AFM) nano-indentation demonstrated that blending silk and tropoelastin at different ratios resulted in modification of mechanical features, with resilience from ~68% to ~97%, and elastic modulus between 2~9Mpa, depending on the ratio of the two polymers. Some of these values are close to those of native aortic elastin or elastin-like polypeptides. Significantly, during blending and drying silk-tropoelastin form micro- and nano-scale porous morphologies which promote human mesenchymal stem cell attachment and proliferation. These blends offer a new protein biomaterial system for cell support and tailored biomaterial properties to match mechanical needs.

Hu, Xiao; Wang, Xiuli; Rnjak, Jelena; Weiss, Anthony S.; Kaplan, David L.

2010-01-01

293

Biomaterials derived from silk-tropoelastin protein systems.  

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A structural protein blend system based on silkworm silk fibroin and recombinant human tropoelastin is described. Silk fibroin, a semicrystalline fibrous protein with beta-sheet crystals provides mechanical strength and controllable biodegradation, while tropoelastin, a noncrystallizable elastic protein provides elasticity. Differential scanning calorimetry (DSC) and temperature modulated DSC (TMDSC) indicated that silk becomes miscible with tropoelastin at different blend ratios, without macrophase separation. Fourier transform infrared spectroscopy (FTIR) revealed secondary structural changes of the blend system (beta-sheet content) before and after methanol treatment. Atomic Force Microscopy (AFM) nano-indentation demonstrated that blending silk and tropoelastin at different ratios resulted in modification of mechanical features, with resilience from approximately 68%- approximately 97%, and elastic modulus between 2 and 9 Mpa, depending on the ratio of the two polymers. Some of these values are close to those of native aortic elastin or elastin-like polypeptides. Significantly, during blending and drying silk-tropoelastin form micro- and nano-scale porous morphologies which promote human mesenchymal stem cell attachment and proliferation. These blends offer a new protein biomaterial system for cell support and tailored biomaterial properties to match mechanical needs. PMID:20674969

Hu, Xiao; Wang, Xiuli; Rnjak, Jelena; Weiss, Anthony S; Kaplan, David L

2010-11-01

294

The expression of GFP under the control of fibroin promotor in primary ovarian cells of Antheraea pernyi.  

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The fibroin promoter can stably express foreign gene in lepidopteran cells. Total RNA was extracted from the gland of silkworm, Antheraea pernyi and the transcription initiation site of fibroin gene of A. pernyi was identified by RNA ligase mediated rapid amplification of cDNA ends (RLM-RACE). The expression vector (pGFP-N2/Fib) was constructed by use of replacing the CMV promoter with the fibroin promoter. The results of visual screening under a fluorescent inverted microscope and Western blot analysis indicated that the GFP gene was expressed in the primary cells of ovary origins from A. pernyi. PMID:14660868

Li, Wenli; Jin, Liji; Bu, Pengcheng; An, Lijia

2003-12-01

295

Production of scFv-conjugated affinity silk film and its application to a novel enzyme-linked immunosorbent assay.  

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Bombyx mori (silkworm) silk proteins have been utilized as unique biomaterials for various medical applications. To develop a novel affinity silk material, we generated a transgenic silkworm that spins silk protein containing the fibroin L-chain linked with the single-chain variable fragment (scFv) as a fusion protein. Previously, the scFv-conjugated "affinity" silk powder specifically immunoprecipitated its target protein, Wiskott-Aldrich syndrome protein. To expand the applicability of affinity silk materials, we processed the scFv-conjugated silk protein into a thin film by dissolving it in lithium bromide, then drying it in the wells of 96-well plates. Enzyme-linked immunosorbent assay demonstrated specific detection of Wiskott-Aldrich syndrome protein, both as a recombinant protein and in its native form extracted from mouse macrophages. These findings suggest that this scFv-conjugated silk film serves as the basis for an alternative immunodetection system. PMID:24518284

Sato, Mitsuru; Kojima, Katsura; Sakuma, Chisato; Murakami, Maria; Tamada, Yasushi; Kitani, Hiroshi

2014-01-01

296

Silk production in a spider involves acid bath treatment  

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We studied physiological conditions during the spinning of dragline silk by the garden cross spider, Araneus diadematus. Silk is converted from the liquid feedstock in the gland into a solid thread via a tapering tubular duct and exits at a spigot. The distal part of the tubule appears specialized for ion transport and the management of the pH inside the lumen. Thus, it appears that spider silk in vivo, like some industrial polymers in vitro, is spun through an acid bath.

Vollrath, F.; Knight, D. P.; Hu, X. W.

1998-01-01

297

Statistical approaches for investigating silk properties  

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Amino acid repeats or motifs have engendered interest because of their significance for protein physical characteristics as well as folding properties. Spider dragline silk proteins are unique because they are composed of long repetitive sections and relatively short non-repetitive sections that are known to interact to generate the very peculiar mechanical and solubility properties of silk. Computational analysis compared with in vitro measurements suggest that the silks achieve their unique pattern of extreme solubility inside the spider glands/complete insolubility outside by correlating their repetitive hydrophobic regions through a type of stochastic resonance, generated by the addition of the non-repetitive sequences to a basically periodic hydrophobicity pattern.

Zbilut, J. P.; Scheibel, T.; Huemmerich, D.; Webber, C. L., Jr.; Colafranceschi, M.; Giuliani, A.

2006-02-01

298

Integration of silk protein in organic and light-emitting transistors  

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We present the integration of a natural protein into electronic and optoelectronic devices by using silk fibroin as a thin film dielectric in an organic thin film field-effect transistor (OFET) ad an organic light emitting transistor device (OLET) structures. Both n- (perylene) and p-type (thiophene) silk-based OFETs are demonstrated. The measured electrical characteristics are in agreement with high-efficiency standard organic transistors, namely charge mobility of the order of 10-2 cm2/Vs and on/off ratio of 104. The silk-based optolectronic element is an advanced unipolar n-type OLET that yields a light emission of 100nW.

Capelli, R.; Amsden, J. J.; Generali, G.; Toffanin, S.; Benfenati, V.; Muccini, M.; Kaplan, D. L.; Omenetto, F. G.; Zamboni, R.

2012-01-01

299

Reproducing natural spider silks' copolymer behavior in synthetic silk mimics.  

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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 ?-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 as 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. PMID:23110450

An, Bo; Jenkins, Janelle E; Sampath, Sujatha; Holland, Gregory P; Hinman, Mike; Yarger, Jeffery L; Lewis, Randolph

2012-12-10

300

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

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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-1? 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-?? 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-? gene. PMID:23896003

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

 
 
 
 
301

Self-assembly in the major ampullate gland of Nephila clavipes  

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We present a tentative interpretation of the origin of nematic liquid crystalline order exhibited by dragline silk fibroin solutions collected from the spider Nephila clavipes. Liquid crystallinity is thought to confer certain rheological properties on the fibroin solution which are exploited during the dragline spinning process. We show that the feasibility of liquid crystallinity under physiological conditions depends critically on parameters characterising the amino-acid ...

Braun, F. N.; Viney, C.

2002-01-01

302

Ultrastructure of the silk glands in three adult females of sphecid wasps of the genus Microstigmus (Hymenoptera: Pemphredoninae Ultraestructura de las glándulas de seda en hembras adultas de tres especies de avispas del género Microstigmus (Hymenoptera: Sphecidae: Pemphredoninae  

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Full Text Available The ultrastructure of silk glands in adult females of sphecid wasps is described. Individual glands are scattered in the more posterior portion of the metasoma being each gland formed by an enlarged secretory unity with one conducting canal. Each secretory unit is a gland cell containing a large lumen and numerous secretory granules, which are delimited by membrane and filled with middle electron dense content of homogeneous aspect, which are exocytosed into the extra cellular lumen. Cell cytoplasm is filled with rough endoplasmic reticulum, polyribosomes and Golgi apparatuses profiles. The basal plasma membrane has many short infoldings and is lined by a thin basement membrane. The apical membrane is invaginated to form the boundary of the lumen, containing scattered short microvilli. The cells are mononucleated and the nucleus is pleomorphic containing disperse chromatin. The extracellular lumen is separated of the receiving canal by a convoluted sponge-like wall forming many villi, which in their tip are lined by a thin single layered cuticle. In the basal region of the villi a cuticular covering is lacking. In the conducting canal secretion acquire two different electron densities. A thin cuticle lines the lumen of conducting canal and the cell cytoplasm is scarce and the organelles are represented for few rough endoplasmic reticulum and polyribosomes. These results are discussed in relation to function and origin of silk gland in adult SphecidaeSe describe la ultraestructura de las glándulas de la seda en tres hembras adultas de avispas Sphecidae del género Microstigmus. Las glándulas individualmente se dispersan en la porción más posterior del metasoma, y cada una de ellas está formada por una unidad secretora grande y un canal que conduce la secreción. Cada célula de la unidad secretora contiene un gran lumen y numerosos gránulos secretores, los que están delimitados por una membrana y repletos de un contenido denso a los electrones de aspecto homogéneo, el que es eliminado hacia el lumen. El citoplasma de la célula está repleto de perfiles del retículo endoplasmático granular, de polirribosomas y de aparatos de Golgi. La membrana plasmática basal presenta muchos repliegues cortos hacia el interior y está revestida por una membrana basal fina. La membrana plasmática apical está invaginada para formar el límite del lumen extracelular, lleva microvellosidades cortas y dispersas. Las células son mononucleadas y el núcleo es de forma variada y contiene cromatina dispersa. El lumen está separado del canal de recepción por una pared replegada semejante a una esponja, con muchas vellosidades, las que en su extremidad están delimitadas por una cutícula fina. En la región basal, las vellosidades carecen de cubierta cuticular. El canal que conduce la secreción adquiere dos densidades de electrones diferentes. Una cutícula fina reviste el lumen del canal que conduce la secreción, siendo escaso el citoplasma de la célula y conteniendo poco retículo endoplasmático granular y polirribosomas. Estos resultados se discuten en relación a la función y al origen de la glándula de seda en los adultos de Sphecidae

JOSÉ EDUARDO SERRÃO

2005-03-01

303

The use of silk-based devices for fracture fixation  

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Metallic fixation systems are currently the gold standard for fracture fixation but have problems including stress shielding, palpability and temperature sensitivity. Recently, resorbable systems have gained interest because they avoid removal and may improve bone remodelling due to the lack of stress shielding. However, their use is limited to paediatric craniofacial procedures mainly due to the laborious implantation requirements. Here we prepare and characterize a new family of resorbable screws prepared from silk fibroin for craniofacial fracture repair. In vivo assessment in rat femurs shows the screws to be self-tapping, remain fixed in the bone for 4 and 8 weeks, exhibit biocompatibility and promote bone remodelling. The silk-based devices compare favourably with current poly-lactic-co-glycolic acid fixation systems, however, silk-based devices offer numerous advantages including ease of implantation, conformal fit to the repair site, sterilization by autoclaving and minimal inflammatory response.

Perrone, Gabriel S.; Leisk, Gary G.; Lo, Tim J.; Moreau, Jodie E.; Haas, Dylan S.; Papenburg, Bernke J.; Golden, Ethan B.; Partlow, Benjamin P.; Fox, Sharon E.; Ibrahim, Ahmed M. S.; Lin, Samuel J.; Kaplan, David L.

2014-03-01

304

The use of silk-based devices for fracture fixation.  

Science.gov (United States)

Metallic fixation systems are currently the gold standard for fracture fixation but have problems including stress shielding, palpability and temperature sensitivity. Recently, resorbable systems have gained interest because they avoid removal and may improve bone remodelling due to the lack of stress shielding. However, their use is limited to paediatric craniofacial procedures mainly due to the laborious implantation requirements. Here we prepare and characterize a new family of resorbable screws prepared from silk fibroin for craniofacial fracture repair. In vivo assessment in rat femurs shows the screws to be self-tapping, remain fixed in the bone for 4 and 8 weeks, exhibit biocompatibility and promote bone remodelling. The silk-based devices compare favourably with current poly-lactic-co-glycolic acid fixation systems, however, silk-based devices offer numerous advantages including ease of implantation, conformal fit to the repair site, sterilization by autoclaving and minimal inflammatory response. PMID:24594992

Perrone, Gabriel S; Leisk, Gary G; Lo, Tim J; Moreau, Jodie E; Haas, Dylan S; Papenburg, Bernke J; Golden, Ethan B; Partlow, Benjamin P; Fox, Sharon E; Ibrahim, Ahmed M S; Lin, Samuel J; Kaplan, David L

2014-01-01

305

Carbon dioxide induced silk protein gelation for biomedical applications.  

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We present a novel method to fabricate silk fibroin hydrogels using high pressure carbon dioxide (CO(2)) as a volatile acid without the need for chemical cross-linking agents or surfactants. The simple and efficient recovery of CO(2) post processing results in a remarkably clean production method offering tremendous benefit toward materials processing for biomedical applications. Further, with this novel technique we reveal that silk protein gelation can be considerably expedited under high pressure CO(2) with the formation of extensive ?-sheet structures and stable hydrogels at processing times less than 2 h. We report a significant influence of the high pressure CO(2) processing environment on silk hydrogel physical properties such as porosity, sample homogeneity, swelling behavior and compressive properties. Microstructural analysis revealed improved porosity and homogeneous composition among high pressure CO(2) specimens in comparison to the less porous and heterogeneous structures of the citric acid control gels. The swelling ratios of silk hydrogels prepared under high pressure CO(2) were significantly reduced compared to the citric acid control gels, which we attribute to enhanced physical cross-linking. Mechanical properties were found to increase significantly for the silk hydrogels prepared under high pressure CO(2), with a 2- and 3-fold increase in the compressive modulus of the 2 and 4 wt % silk hydrogels over the control gels, respectively. We adopted a semiempirical theoretical model to elucidate the mechanism of silk protein gelation demonstrated here. Mechanistically, the rate of silk protein gelation is believed to be a function of the kinetics of solution acidification from absorbed CO(2) and potentially accelerated by high pressure effects. The attractive features of the method described here include the acceleration of stable silk hydrogel formation, free of residual mineral acids or chemical cross-linkers, reducing processing complexity, and avoiding adverse biological responses, while providing direct manipulation of hydrogel physical properties for tailoring toward specific biomedical applications. PMID:22657735

Floren, Michael L; Spilimbergo, Sara; Motta, Antonella; Migliaresi, Claudio

2012-07-01

306

Impact of sterilization on the enzymatic degradation and mechanical properties of silk biomaterials.  

Science.gov (United States)

The effect of some sterilization methods (autoclaving and ethanol treatments) on the degradation rate and mechanical properties of two types of porous silk scaffolds (aqueous- and hexafluoroisopropanol-derived) is evaluated. Changes in secondary structure, crystal size, and supramolecular features of silk fibroin, resulting from sterilization, are tracked to elucidate molecular level effects on protease XIV enzymatic degradation and compressive mechanical properties. The structural features and pore sizes of the silk scaffolds remain intact after both sterilization processes. Autoclave sterilization dramatically reduce the degradation rate of the silk scaffolds in response to protease XIV and significantly increase mechanical properties, in contrast to scaffolds sterilized with 70% ethanol. Higher ?-sheet content and larger crystal size are observed after autoclaving, unlike in response to 70% ethanol sterilization, based on examination of Fourier transform (FT) IR spectroscopy and wide-angle X-ray scattering (WAXS). In addition, thermal analysis finds supramolecular features within silk fibroin amorphous regions, including the glass transition temperature (Tg ), heat capacity of glass transition (?Cp-Tg ), and thermal gravimetric degradability. Such supramolecular level changes are related to the shift in enzymatic degradation and mechanical properties due to autoclaving versus treatment with 70% EtOH. The changes in supramolecular organization in amorphous regions can retard enzyme diffusion through the glassy regions of the silk matrix or/and hinder binding of enzymes, while also stiffening these matrices. PMID:24519787

Gil, Eun Seok; Park, Sang-Hyug; Hu, Xiao; Cebe, Peggy; Kaplan, David L

2014-02-01

307

Radiation processing of silk protein (Bilateral research cooperation OAEP and JAERI. December 1998 - December 2002)  

International Nuclear Information System (INIS)

Thailand's production of silk, about 1,200 ton per year, also gives about 10% of silk waste which is expected to be recycled into new material (non-textile application) and to avoid environmental pollution. For this purpose, cooperative program 'radiation processing of silk protein' was conducted between OAEP (Thailand) and JAERI. Among the results already obtained are: radiation degradation of silk protein (fibroin) with gamma rays at 160 kGy, production of fine silk milled powder (<90 microns) by electron beam irradiation at 250-1000 kGy (dry method) using electron accelerator (1 MeV, 1 mA), use of antioxidant effect of silk protein on lipid peroxidation and antibacterial activity of irradiated silk protein powder, and wound dressing hydrogel mixed with silk protein and use of antibacterial activity of cross-linked silk protein/PVA hydrogel. Other topics of interest are gamma irradiation of anionic natural polymer solution for use as latex protein scavenger and gamma radiation degradation of chitosan for use as plant growth promoter and fungicide. (S. Ohno)

2003-10-01

308

Functional expression of a Bombyx mori cocoonase: potential application for silk degumming.  

Science.gov (United States)

Cocoon, a shelter for larva development to silk moth, contains the fibrous protein fibroin, which is coated by the globular protein sericin. Emergence of the silk moth requires the action of cocoonase, a protease secreted by the pupa. The full-length prococoonase cDNA, with 780 bp open reading frame encoding 260 amino acids, was cloned by reverse transcription from total RNA of the head of 6-day-old Thai-silk Bombyx mori pupa. Only the gene fragment lacking the propeptide encoding sequence was successfully expressed in Pichia pastoris, yielding an extracellularly active cocoonase. The recombinant cocoonase was purified to homogeneity by 80% ammonium-sulfate fractionation and CM-Sepharose chromatography, and its internal peptide sequences were analyzed by nano liquid chromatography-mass spectrometry/mass spectrometry. This monomeric protein has native molecular weight of 26 kDa by gel exclusion analysis and 25 kDa subunit size by sodium dodecyl sulphate-polyacrylamide gel electrophoresis. The enzyme hydrolyses sericin but does not hydrolyse fibroin, as shown by radial diffusion on thin-layer enzyme assay (RD-TEA). Scanning electron microscopy showed that purified recombinant cocoonase could remove sericin from natural silk completely in 24 h, without damaging fibroin, using only 1 immobilized sericin unit (ISU) of enzyme as determined by RD-TEA. Natural cocoonase isolated from B. mori pupa could also digest sericin effectively, but required more enzymes (2 ISU) and longer time (48 h). In comparison, a commercial enzyme, alcalase, with the same activity not only showed less complete digestion of sericin but also caused damage of fibroin. These results suggest that recombinant B. mori cocoonase is potentially useful for silk degumming. PMID:23169343

Rodbumrer, Prangprapai; Arthan, Dumrongkiet; Uyen, Utai; Yuvaniyama, Jirundon; Svasti, Jisnuson; Wongsaengchantra, Pramvadee Y

2012-12-01

309

Role of pH and charge on silk protein assembly in insects and spiders  

Science.gov (United States)

Silk fibers possess impressive mechanical properties, dependant, in part, on the crystalline ?-sheets silk II conformation. The transition to silk II from soluble silk I-like conformation in silk glands, is thought to originate in the spinning ducts immediately before the silk is drawn down into a fiber. However the assembly process of these silk molecules into fibers, whether in silkworms or spiders, is not well understood. Extensional flow, protein concentration, pH and metal ion concentrations are thought to be most important in in vivo silk processing and in affecting structural conformations. We look at how parameters such as pH, [Ca2+], [K+], and [Cu2+], and water content, interact with the domain structure of silk proteins towards the successful storage and processing of these concentrated hydrophobic silk proteins. Our recent domain mapping studies of all known silk proteins, and 2D Raman spectroscopy, NMR, and DLS studies performed on sections of silkworm gland, suggest that low pH and gradual water removal promote intermolecular over intramolecular hydrogen bonding. This discussion helps to provide the necessary ground rules towards the design of silk protein analogues with specific hydrophobicity and charge profiles to optimize expression, solubility and assembly with implications in structural biology and material science.

Foo, C. Wong Po; Bini, E.; Hensman, J.; Knight, D. P.; Lewis, R. V.; Kaplan, D. L.

2006-02-01

310

Physical characterization of functionalized spider silk: electronic and sensing properties  

Energy Technology Data Exchange (ETDEWEB)

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 {beta}-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 deg. 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 {beta}-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 {beta}-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.

Steven, Eden; Brooks, James S [Department of Physics and National High Magnetic Field Laboratory, Florida State University, 1800 East Paul Dirac, Tallahassee, FL 32310 (United States); Park, Jin Gyu [FAMU-FSU Department of Industrial and Manufacturing Engineering, High-Performance Materials Institute, Florida State University, 2005 Levy Ave., Tallahassee, FL 32310 (United States); Paravastu, Anant; Siegrist, Theo; Kaner, Papatya; Alamo, Rufina G [FAMU-FSU Department of Chemical and Biomedical Engineering and National High Magnetic Field Laboratory, Florida State University, 1800 East Paul Dirac, Tallahassee, FL 32310 (United States); Branco Lopes, Elsa [Departamento de Quimica, Instituto Tecnologico e Nuclear/CFMC-UL, P-2686-953 Sacavem (Portugal); Englander, Ongi, E-mail: esteven@magnet.fsu.edu [FAMU-FSU Department of Mechanical Engineering and National High Magnetic Field Laboratory, Florida State University, 1800 East Paul Dirac, Tallahassee, Florida 32310 (United States)

2011-10-15

311

Physical characterization of functionalized spider silk: electronic and sensing properties  

International Nuclear Information System (INIS)

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 ?-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 deg. 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 ?-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 ?-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.

2011-10-01

312

Physical characterization of functionalized spider silk: electronic and sensing properties  

Directory of Open Access Journals (Sweden)

Full Text Available 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 ?-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 ?-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 ?-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.

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

2011-01-01

313

New exocrine glands in ants: the hypostomal gland and basitarsal gland in the genus Melissotarsus (Hymenoptera: Formicidae)  

Science.gov (United States)

Fisher and Robertson (Insect Soc 46: 78-83, 1999) discovered the production of silk-like secretions emerging from slit-shaped openings along the anterior margin of the ventral hypostoma of Melissotarsus ant workers. The current histological study describes a hitherto unknown hypostomal gland from which this silk-like substance originates. In addition, this study describes a new basitarsal gland in the three pairs of legs of Melissotarsus workers.

Hölldobler, Bert; Obermayer, Malu; Plowes, Nicola J. R.; Fisher, Brian L.

2014-06-01

314

Controlled release from multilayer silk biomaterial coatings to modulate vascular cell responses.  

Science.gov (United States)

A multilayered silk fibroin protein coating system was employed as a drug carrier and delivery system to evaluate vascular cell responses to heparin, paclitaxel, and clopidogrel. The results demonstrated that the silk coating system was an effective system for drug-eluting coatings, such as for stent applications, based on its useful micromechanical properties and biological outcomes. Cell attachment and viability studies with human aortic endothelial cells (HAECs) and human coronary artery smooth muscle cells (HCASMCs) on the drug-incorporated silk coatings demonstrated that paclitaxel and clopidogrel inhibited smooth muscle cell (SMC) proliferation and retarded endothelial cell proliferation. Heparin-loaded silk multilayers promoted HAEC proliferation while inhibiting HCASMC proliferation, desired outcomes for the prevention of restenosis. The preservation of the phenotype of endothelial cells on silk and heparin-loaded silk coatings was confirmed with the presence of endothelial markers CD-31, CD-146, vWF and VE-Cadherin using immunocytochemistry assays. A preliminary in-vivo study in a porcine aorta showed integrity of the silk coatings after implantation and the reduction of platelet adhesion on the heparin-loaded silk coatings. PMID:18048096

Wang, Xianyan; Zhang, Xiaohui; Castellot, John; Herman, Ira; Iafrati, Mark; Kaplan, David L

2008-03-01

315

Effect of pH value on graft copolymerization of 2-(dimethylamino)ethyl methacrylate onto silk fabrics initiated by UV rays  

International Nuclear Information System (INIS)

Effect of pH value on graft yield (Yg) of 2-(dimethylamino)ethyl methacrylate (DMAEMA) onto silk fabrics initiated by ultraviolet rays are investigated in this paper. Three kinds of grafted silk fabrics obtained in different pH adjustment processes were characterized by DSC (differential scanning calorimetry), TGA/DTG (thermogravimetry analysis and derivative thermogravimetry), DTA (differential thermal analysis) and contact angle measurement. The thermal analysis results indicated that the grafted silk fabrics showed different compatibility between the silk fibroin molecules and 2-(dimethylamino)ethyl methacrylate polymers. And the crystalline structure of the grafted silk fabrics remained unchanged, or changed due to the pH adjustment process. It was found that the gel particles sized differently in solutions with the pH value adjusted in different ways and this might lead to different grafted effects of the silk fabrics. (authors)

2008-03-01

316

Reproducing Natural Spider Silks’ Copolymer Behavior in Synthetic Silk Mimics  

Digital Repository Infrastructure Vision for European Research (DRIVER)

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 ?-sheet nanostructures, while MaSp2 proteins form random coil and helical structures. To test if this...

An, Bo; Jenkins, Janelle E.; Sampath, Sujatha; Holland, Gregory P.; Hinman, Mike; Yarger, Jeffery L.; Lewis, Randolph

2012-01-01

317

NMR analysis of the fibronectin cell-adhesive sequence, Arg-Gly-Asp, in a recombinant silk-like protein and a model peptide.  

Science.gov (United States)

It is well established that by introducing the cell-adhesive sequence Arg-Gly-Asp (RGD) from fibronectin into Bombyx mori silk fibroin by covalent coupling or bioengineering techniques, excellent biomaterials have been developed with the modified silk fibroin. However, there is no report about the structure and dynamics of the RGD moiety in the silk fibroin. To clarify the origin of such a high cell adhesion character and to design new recombinant silk protein with higher cell adhesion ability, it is necessary to characterize the structure and dynamics of the RGD moiety introduced into silk fibroin. In this study, the structure and dynamics of the RGD moiety in a recombinant silk-like protein, SLPF(10), consisting of the repeated silk fibroin sequence (AGSGAG)(3) and the sequence ASTGRGDSPA including the RGD moiety, were studied using solution NMR. The (1)H, (15)N, and (13)C chemical shifts indicate that the RGD moiety, as well as the silk fibroin sequence, takes a random coil form with high mobility in aqueous solution. Next, a (13)C solid-state NMR study was performed on a (13)C selectively labeled model peptide, AGSGAG[3-(13)C]A(7)GSGAGAGSGGT[2-(13)C]G(19)R[1-(13)C]G(21)DSPAGGGAGAGSGAG. After formic acid treatment, an increase in the ?-sheet fraction for the AGSGAG sequence and peak narrowing of the residues around the RGD moiety were observed in the dry state. The latter indicates a decrease in the chemical shift distribution although the RGD moiety is still in random coil. A decrease in the peak intensities of the RGD moiety in the swollen state after immersing it in distilled water was observed, indicating high mobility of the RGD sequence in the peptide in the swollen state. Thus, the random coil state of the RGD moiety in the recombinant silk-like protein is maintained in aqueous solution and also in both dry and swollen state. This is similar to the case of the RGD moiety in fibronectin. The presence of the linker ASTG at the N-terminus and SPAGG at the C-terminus seems important to maintain the random coil form and the flexible state of the RGD sequence in order to permit access for binding to various integrins. PMID:21955288

Asakura, Tetsuo; Nishi, Hirohito; Nagano, Aya; Yoshida, Ai; Nakazawa, Yasumoto; Kamiya, Masakatsu; Demura, Makoto

2011-11-14

318

Identification and synthesis of novel biomaterials based on spider structural silk fibers  

Science.gov (United States)

The diversity in function and mechanical behavior of spider silks, and the ability to produce these silks recombinantly, have tremendous potential in creating a new class of biomimetic materials. Here we investigate the structural and mechanical properties of pyriform silks from the golden orb-weaver, Nephila clavipes. Nanoscale indentation measurements using atomic force microscopy on natural pyriform silk suggests that this biomaterial has high toughness that may be suitable for dissipating high amounts of mechanical energy. We also observed the occurrence of highly organized nanocrystals within the pyriform silk fibers that may contribute to the remarkable energy dissipation capability of these silks. It has been demonstrated that poly-(Gly-Ala) and poly-Ala stretches within the internal block repeat modules of dragline silk fibroins form nanocrystals, and these nanocrystalline structures may be responsible for the high extensibility of the dragline silks. In contrast, amino acid sequence analysis shows that PySp2 does not contain the same motifs. In the absence of poly-(Gly-Ala) and poly-Ala repeats, we hypothesized that PySp2 contains new protein motifs sufficient to polymerize into functional structures. To investigate the functional contributions of these novel motifs during pyriform fiber formation, we expressed different recombinant PySp2 fibroins with various segments spanning its block repeat units. We demonstrate that PySp2 recombinant proteins with the Pro-rich sub-block domain (PXP motifs, where X= sub-set of the amino acids A, L, or R) and/or the Ser + Gln + Ala-rich sub-block domain (QQSSVAQS motifs) are sufficient for artificial fiber formation. Moreover, we show that recombinant PySp2 proteins that contain a single block repeat unit can self-assemble into foam-like nanostructures. Collectively, our findings support the use of PySp2 recombinant proteins for a wide range of biomimetic materials with morphologies ranging from fibers to porous structures.

Hsia, Yang; Gnesa, Eric; Tang, Simon; Jeffery, Felicia; Geurts, Paul; Zhao, Liang; Franz, Andreas; Vierra, Craig

2011-11-01

319

Silk hydrogels from non-mulberry and mulberry silkworm cocoons processed with ionic liquids  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Matrices based on silk fibroin from the non-mulberry silkworm Antheraea mylitta and the mulberry silkworm Bombyx mori have demonstrated good applicability in regenerative medicine. However, the cocoons of A. mylitta are underutilized in part due to their lack of solubility in traditional organic solvents. Therefore, the present work investigates the solubilization and processing of degummed fibers obtained from the cocoons of both silkworm species into hydrogels using ionic liquids (ILs). The...

Silva, S. S.; Popa, Elena G.; Gomes, Manuela E.; Oliveira, M. B.; Nayak, S.; Bano, S.; Mano, J. F.; Kundu, S. C.; Reis, R. L.

2013-01-01

320

Air Filter Devices Including Nonwoven Meshes of Electrospun Recombinant Spider Silk Proteins  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Based on the natural sequence of Araneus diadematus Fibroin 4 (ADF4), the recombinant spider silk protein eADF4(C16) has been engineered. This highly repetitive protein has a molecular weight of 48kDa and is soluble in different solvents (hexafluoroisopropanol (HFIP), formic acid and aqueous buffers). eADF4(C16) provides a high potential for various technical applications when processed into morphologies such as films, capsules, particles, hydrogels, coatings, fibers and nonwoven meshes. Due ...

2013-01-01

 
 
 
 
321

Material quality assessment of silk nanofibers based on swarm intelligence  

International Nuclear Information System (INIS)

In this paper, we propose a novel approach for texture analysis based on artificial crawler model. Our method assumes that each agent can interact with the environment and each other. The evolution process converges to an equilibrium state according to the set of rules. For each textured image, the feature vector is composed by signatures of the live agents curve at each time. Experimental results revealed that combining the minimum and maximum signatures into one increase the classification rate. In addition, we pioneer the use of autonomous agents for characterizing silk fibroin scaffolds. The results strongly suggest that our approach can be successfully employed for texture analysis.

2013-02-08

322

Material quality assessment of silk nanofibers based on swarm intelligence  

Science.gov (United States)

In this paper, we propose a novel approach for texture analysis based on artificial crawler model. Our method assumes that each agent can interact with the environment and each other. The evolution process converges to an equilibrium state according to the set of rules. For each textured image, the feature vector is composed by signatures of the live agents curve at each time. Experimental results revealed that combining the minimum and maximum signatures into one increase the classification rate. In addition, we pioneer the use of autonomous agents for characterizing silk fibroin scaffolds. The results strongly suggest that our approach can be successfully employed for texture analysis.

Brandoli Machado, Bruno; Nunes Gonçalves, Wesley; Martinez Bruno, Odemir

2013-02-01

323

Spider Silk Proteins.  

Science.gov (United States)

Spider silk proteins from major and minor ampullate silk have been sequenced via their cDNAs. One of the proteins from major ampullate silk has been expressed in bacteria to levels of 10-20 mg/mL. Studies using fiber X-ray diffraction and solid state NMR ...

R. V. Lewis

1994-01-01

324

Human amniotic fluid stem cells seeded in fibroin scaffold produce in vivo mineralized matrix.  

Science.gov (United States)

This study investigated the potential of amniotic fluid stem cells (AFSCs) to synthesize mineralized extracellular matrix (ECM) within different porous scaffolds of collagen, poly-D,L-lactic acid (PDLLA), and silk fibroin. The AFSCs were initially differentiated by using an osteogenic medium in two-dimensional culture, and expression of specific bone proteins and the physiologic mineral production by the AFSCs were analyzed. In particular, during differentiation process, AFSCs expressed proteins like Runt-related transcription factor 2 (Runx2), Osterix, Osteopontin, and Osteocalcin with a sequential expression, analogous to those occurring during osteoblast differentiation, and produced extracellular calcium stores. AFSCs were then cultured on three-dimensional (3D) scaffolds and evaluated for their ability to differentiate into osteoblastic cells in vivo. Stem cells were cultured in vitro for 1 week in collagen, fibroin, and PDLLA scaffolds. The effect of predifferentiation of the stem cells in scaffolds on the subsequent bone formation in vivo was determined in a rat subcutaneous model. With the addition of a third dimension, osteogenic differentiation and mineralized ECM production by AFSCs were significantly higher. This study demonstrated the strong potential of AFSCs to produce 3D mineralized bioengineered constructs in vivo and suggests that fibroin may be an effective scaffold material for functional repair of critical size bone defects. PMID:21864161

Maraldi, Tullia; Riccio, Massimo; Resca, Elisa; Pisciotta, Alessandra; La Sala, Giovanni B; Ferrari, Adriano; Bruzzesi, Giacomo; Motta, Antonella; Migliaresi, Claudio; Marzona, Laura; De Pol, Anto

2011-11-01

325

Pulsed laser deposition of silk protein: Effect of photosensitized-ablation on the secondary structure in thin deposited films  

Science.gov (United States)

Silk fibroin is a simple protein expected to have functional applications in medicine and bioelectronics. The primary structure of this protein is quite simple, and the main secondary structures are ?-sheet crystals and amorphous random coils. In the present study, we investigated pulsed laser deposition (PLD) of fibroin with the ?-sheet structures as targets. The primary and secondary structures in films deposited were analyzed using infrared spectroscopy. Normal laser deposition at 351 nm using neat fibroin targets produced thin films of fibroin with a random coiled structure. Ablation was triggered by two-photonic excitation of the peptide chains, which resulted in the destruction of ?-sheet structure in PLD. In order to avoid the two-photonic excitation, we adopted a PLD method utilizing anthracene (5-0.1 wt %) in a photosensitized reaction involving doped fibroin targets. Laser light (351 or 355 nm) was absorbed only by anthracene, which plays an important role converting photon energy to thermal energy with great ablation efficiency. Thin fibroin films deposited by this method had both random coil and ?-sheet structures. As the dopant concentration and laser fluence decreased, the ratio of ?-sheet domain to random coil increased in thin deposited films.

Tsuboi, Yasuyuki; Goto, Masaharu; Itaya, Akira

2001-06-01

326

Protein families, natural history and biotechnological aspects of spider silk.  

Science.gov (United States)

Spiders are exceptionally diverse and abundant organisms in terrestrial ecosystems and their evolutionary success is certainly related to their capacity to produce different types of silks during their life cycle, making a specialized use on each of them. Presenting particularly tandemly arranged amino acid repeats, silk proteins (spidroins) have mechanical properties superior to most synthetic or natural high-performance fibers, which makes them very promising for biotechnology industry, with putative applications in the production of new biomaterials. During the evolution of spider species, complex behaviors of web production and usage have been coupled with anatomical specialization of spinning glands. Spiders retaining ancestral characters, such as the ones belonging to the Mygalomorph group, present simpler sorts of webs used mainly to build burrows and egg sacs, and their silks are produced by globular undifferentiated spinning glands. In contrast, Araneomorphae spiders have a complex spinning apparatus, presenting up to seven morphologically distinct glands, capable to produce a more complex set of silk polymers with different degrees of rigidness and elasticity associated with distinct behaviors. Aiming to provide a discussion involving a number of spider silks' biological aspects, in this review we present descriptions of members from each family of spidroin identified from five spider species of the Brazilian biodiversity, and an evolutionary study of them in correlation with the anatomical specialization of glands and spider's spinning behaviors. Due to the biotechnological importance of spider silks for the production of new biomaterials, we also discuss about the new possible technical and biomedical applications of spider silks and the current status of it. PMID:22911606

Bittencourt, D; Oliveira, P F; Prosdocimi, F; Rech, E L

2012-01-01

327

Integration of silk protein in organic and light-emitting transistors.  

Science.gov (United States)

We present the integration of a natural protein into electronic and optoelectronic devices by using silk fibroin as a thin film dielectric in an organic thin film field-effect transistor (OFET) ad an organic light emitting transistor device (OLET) structures. Both n- (perylene) and p-type (thiophene) silk-based OFETs are demonstrated. The measured electrical characteristics are in agreement with high-efficiency standard organic transistors, namely charge mobility of the order of 10(-2) cm(2)/Vs and on/off ratio of 10(4). The silk-based optolectronic element is an advanced unipolar n-type OLET that yields a light emission of 100nW. PMID:22899899

Capelli, R; Amsden, J J; Generali, G; Toffanin, S; Benfenati, V; Muccini, M; Kaplan, D L; Omenetto, F G; Zamboni, R

2011-07-01

328

PLGA nanofiber-coated silk microfibrous scaffold for connective tissue engineering.  

Science.gov (United States)

A modified degumming technique, involving boiling in 0.25% Na2CO3 with addition of 1% sodium dodecyl sulphate and intermittent ultrasonic agitation, was developed for knitted silk scaffolds. Sericin was efficiently removed, while mechanical and structural properties of native silk fibroin were preserved. Biocompatible and mechanically robust hybrid nano-microscaffolds were fabricated by coating these degummed silk scaffolds with an intervening adhesive layer of silk solution followed by electrospun poly-lactic-co-glycolic acid (PLGA) nanofibers. Cell proliferation on the hybrid silk scaffolds was improved by seeding cells on both surfaces of the flat scaffolds. Rolling up and continued culture of the cell-seeded hybrid scaffolds yielded cylindrical constructs that permitted cell proliferation, extracellular matrix deposition, and generated ligament/tendon graft analogs. Although PLGA-based hybrid scaffolds have earlier been proposed for dense connective tissue engineering, rapid biodegradation of PLGA was a drawback. In contrast, the underlying strong and slowly-degrading microfibrous silk scaffold used in this study ensured that the hybrid scaffold maintained adequate mechanical properties for longer periods, which is vital for continued support to the injured ligament/tendon throughout its healing period. PMID:20665681

Sahoo, Sambit; Toh, Siew Lok; Goh, James Cho Hong

2010-10-01

329

Enhanced cellular adhesion on titanium by silk functionalized with titanium binding and RGD peptides.  

Science.gov (United States)

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. A 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 arginine-glycine-aspartic acid (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

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

2013-01-01

330

Study of protein conformation and orientation in silkworm and spider silk fibers using Raman microspectroscopy.  

Science.gov (United States)

Raman microspectroscopy has been used for the first time to determine quantitatively the orientation of the beta-sheets in silk monofilaments from Bombyx mori and Samia cynthia ricini silkworms, and from the spider Nephila edulis. It is shown that, for systems with uniaxial symmetry such as silk, it is possible to determine the order parameters P2 and P4 of the orientation distribution function from intensity ratios of polarized Raman spectra. The equations allowing the calculation of P2 and P4 using polarized Raman microspectroscopy for a vibration with a cylindrical Raman tensor were first derived and then applied to the amide I band that is mostly due to the C=O stretching vibration of the peptide groups. The shape of the Raman tensor for the amide I vibration of the beta-sheets was determined from an isotropic film of Bombyx mori silk treated with methanol. For both the Bombyx mori and Samia cynthia ricini fibroin fibers, the values of P2 and P4 obtained are equal to -0.36 +/- 0.03 and 0.19 +/- 0.02, respectively, even though the two types of silkworm fibroins strongly differ in their primary sequences. For the Nephila edulis dragline silk, values of P2 and P4 of -0.32 +/- 0.02 and 0.13 +/- 0.02 were obtained, respectively. These results clearly indicate that the carbonyl groups are highly oriented perpendicular to the fiber axis and that the beta-sheets are oriented parallel to the fiber axis, in agreement with previous X-ray and NMR results. The most probable distribution of orientation was also calculated from the values of P2 and P4 using the information entropy theory. For the three types of silk, the beta-sheets are highly oriented parallel to the fiber axis. The orientation distributions of the beta-sheets are nearly Gaussian functions with a width of 32 degrees and 40 degrees for the silkworm fibroins and the spider dragline silk, respectively. In addition to these results, the comparison of the Raman spectra recorded for the different silk samples and the polarization dependence of several bands has allowed to clarify some important band assignments. PMID:15530039

Rousseau, Marie-Eve; Lefèvre, Thierry; Beaulieu, Lilyane; Asakura, Tetsuo; Pézolet, Michel

2004-01-01

331

Elastic Spider Silk Proteins.  

Science.gov (United States)

Flagelliform silk proteins have been studied by cloning the cDNAs for the major protein in that silk as well as the gene. The protein consists of three sequence segments which compose a repeat These repeats appear numerous times in the protein The three s...

R. V. Lewis

1998-01-01

332

Microscopic structural analysis of fractured silk fibers from Bombyx mori and Samia cynthia ricini using 13C CP/MAS NMR with a 1mm microcoil MAS NMR probehead.  

Science.gov (United States)

Conformational changes have been studied in silk fibers from the domestic silkworm Bombyx mori and a wild silkworm Samia cynthia ricini as a result of fractured by stretching. About 300 samples consisting of only the fractured regions of [1-13C]Ala or [1-13C]Gly labeled silk fibers were collected and observed by 13C CP/MAS NMR spectra. The total amount of these fractured fibers is only about 1mg and therefore we used a home-built 1mm microcoil MAS NMR probehead. A very small increase in the fraction of random coil was noted for the alanine regions of both silk fibroins and for the glycine region of B. mori silk fibroin. However, there is no difference in the spectra before and after fractured for the glycine region of S. c. ricini silk fibroin. Thus, the influence of fracture occurs exclusively at the Ala region for S. c. ricini. The relationship between sequence, fracture and structure is discussed. PMID:20579856

Yamauchi, Kazuo; Yamasaki, Shizuo; Takahashi, Rui; Asakura, Tetsuo

2010-07-01

333

13 C solid-state NMR study of the 13 C-labeled peptide, (E)8 GGLGGQGAG(A)6 GGAGQGGYGG as a model for the local structure of Nephila clavipes dragline silk (MaSp1) before and after spinning.  

Science.gov (United States)

We prepared the water soluble model peptide, (E)(8) GGLGGQGAG(A)(6) GGAGQGGYGG, to throw light on the local structure of spidroin 1 (MaSpl) protein in spider dragline silk of Nephila clavipes before and after spinning. Solution (13) C NMR showed that the conformation of the peptide in aqueous solution was essentially random coil. Solid-state NMR was used to follow conformation-dependent (13) C chemical shifts in (13) C selectively labeled versions of the peptide. The peptide lyophilized from an aqueous solution at neutral pH (hereafter referred to as "without acid treatment)"was used to mimic the state of the spidroin stored in the spider's silk gland while the peptide precipitated from the acidic solution ("with acid treatment") was used to simulate the role of acid treatment in inducing conformation change in the natural spinning process. In without acid treatment, the fraction of random coil conformation was lowest in the N-terminal region (residues 15-18) when compared with the C-terminus. The conformational change produced by the acid treatment occurred in the sequence, G(15) AG(A)(6) GGAG(27), interposed between pairs of Gly residues pairs, Gly(12,13), and Gly(29,30). The acid treated peptide showed a remarkable decrease in the fraction of random coil conformation from A(20) to A(23) in the poly-Ala region when compared with the peptide without acid treatment. These observations taken together suggest that the peptide can be used as a model for studying the localization of the conformation change in spider silk fibroin in the natural spinning and the role of acid treatment in this process. PMID:21913180

Yazawa, Koji; Yamaguchi, Erika; Knight, David; Asakura, Tetsuo

2012-06-01

334

Structure of a protein superfiber: spider dragline silk.  

Science.gov (United States)

Spider major ampullate (dragline) silk is an extracellular fibrous protein with unique characteristics of strength and elasticity. The silk fiber has been proposed to consist of pseudocrystalline regions of antiparallel beta-sheet interspersed with elastic amorphous segments. The repetitive sequence of a fibroin protein from major ampullate silk of the spider Nephila clavipes was determined from a partial cDNA clone. The repeating unit is a maximum of 34 amino acids long and is not rigidly conserved. The repeat unit is composed of three different segments: (i) a 6 amino acid segment that is conserved in sequence but has deletions of 3 or 6 amino acids in many of the repeats; (ii) a 13 amino acid segment dominated by a polyalanine sequence of 5-7 residues; (iii) a 15 amino acid, highly conserved segment. The latter is predominantly a Gly-Gly-Xaa repeat with Xaa being alanine, tyrosine, leucine, or glutamine. The codon usage for this DNA is highly selective, avoiding the use of cytosine or guanine in the third position. A model for the physical properties of fiber formation, strength, and elasticity, based on this repetitive protein sequence, is presented.

Xu, M; Lewis, R V

1990-01-01

335

Protein Secondary Structure and Orientation in Silk as Revealed by Raman Spectromicroscopy  

Science.gov (United States)

Taking advantage of recent advances in polarized Raman microspectroscopy, and based on a rational decomposition of the amide I band, the conformation and orientation of proteins have been determined for cocoon silks of the silkworms Bombyx mori and Samia cynthia ricini and dragline silks of the spiders Nephila clavipes and Nephila edulis. This study distinguished between band components due to ?-sheets, ?-turns, 31-helices, and unordered structure for the four fibers. For B. mori, the ?-sheet content is 50%, which matches the proportion of residues that form the GAGAGS fibroin motifs. For the Nephila dragline and S. c. ricini cocoon, the ?-sheet content (36–37% and 45%, respectively) is higher than the proportion of residues that belong to polyalanine blocks (18% and 42%, respectively), showing that adjacent GGA motifs are incorporated into the ?-sheets. Nephila spidroins contain fewer ?-sheets and more flexible secondary structures than silkworm fibroins. The amorphous polypeptide chains are preferentially aligned parallel to the fiber direction, although their level of orientation is much lower than that of ?-sheets. Overall, the results show that the four silks exhibit a common molecular organization, with mixtures of different amounts of ?-sheets and flexible structures, which are organized with specific orientation levels.

Lefevre, Thierry; Rousseau, Marie-Eve; Pezolet, Michel

2007-01-01

336

Protein secondary structure and orientation in silk as revealed by Raman spectromicroscopy.  

Science.gov (United States)

Taking advantage of recent advances in polarized Raman microspectroscopy, and based on a rational decomposition of the amide I band, the conformation and orientation of proteins have been determined for cocoon silks of the silkworms Bombyx mori and Samia cynthia ricini and dragline silks of the spiders Nephila clavipes and Nephila edulis. This study distinguished between band components due to beta-sheets, beta-turns, 3(1)-helices, and unordered structure for the four fibers. For B. mori, the beta-sheet content is 50%, which matches the proportion of residues that form the GAGAGS fibroin motifs. For the Nephila dragline and S. c. ricini cocoon, the beta-sheet content (36-37% and 45%, respectively) is higher than the proportion of residues that belong to polyalanine blocks (18% and 42%, respectively), showing that adjacent GGA motifs are incorporated into the beta-sheets. Nephila spidroins contain fewer beta-sheets and more flexible secondary structures than silkworm fibroins. The amorphous polypeptide chains are preferentially aligned parallel to the fiber direction, although their level of orientation is much lower than that of beta-sheets. Overall, the results show that the four silks exhibit a common molecular organization, with mixtures of different amounts of beta-sheets and flexible structures, which are organized with specific orientation levels. PMID:17277183

Lefèvre, Thierry; Rousseau, Marie-Eve; Pézolet, Michel

2007-04-15

337

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

Science.gov (United States)

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. PMID:21558082

Aramwit, Pornanong; Siritientong, Tippawan; Srichana, Teerapol

2012-03-01

338

Solid-state NMR determination of the secondary structure of Samia cynthia ricini silk.  

Science.gov (United States)

Silks are fibrous proteins that form heterogeneous, semi-crystalline solids. Silk proteins have a variety of physical properties reflecting their range of functions. Spider dragline silk, for example, has high tensile strength and elasticity, whereas other silks are better suited to making housing, egg sacs or the capture spiral of spiders' webs. The differing physical properties arise from variation in the protein's primary and secondary structure, and their packing in the solid phase. The high mechanical performance of spider dragline silk, for example, is probably due to a beta-sheet conformation of poly-alanine domains, embedded as small crystallites within the fibre. Only limited structural information can be obtained from diffraction of silks, so further characterization requires spectroscopic studies such as NMR. However, the classical approach to NMR structure determination fails because the high molecular weight, repetitive primary structure and structural heterogeneity of solid silk means that signals from individual amino-acid residues cannot be resolved. Here we adapt a recently developed solid-state NMR technique to determine torsion angle pairs (phi, psi) in the protein backbone, and we study the distribution of conformations in silk from the Eri silkworm, Samia cynthia ricini. Although the most probable conformation in native fibres is an anti-parallel beta-sheet, film produced from liquid directly extracted from the silk glands appears to be primarily alpha-helical. PMID:10890452

van Beek, J D; Beaulieu, L; Schäfer, H; Demura, M; Asakura, T; Meier, B H

2000-06-29

339

Transparent and flexible resistive switching memory devices with a very high ON/OFF ratio using gold nanoparticles embedded in a silk protein matrix  

Science.gov (United States)

The growing demand for biomaterials for electrical and optical devices is motivated by the need to make building blocks for the next generation of printable bio-electronic devices. In this study, transparent and flexible resistive memory devices with a very high ON/OFF ratio incorporating gold nanoparticles into the Bombyx mori silk protein fibroin biopolymer are demonstrated. The novel electronic memory effect is based on filamentary switching, which leads to the occurrence of bistable states with an ON/OFF ratio larger than six orders of magnitude. The mechanism of this process is attributed to the formation of conductive filaments through silk fibroin and gold nanoparticles in the nanocomposite. The proposed hybrid bio-inorganic devices show promise for use in future flexible and transparent nanoelectronic systems.

Gogurla, Narendar; Mondal, Suvra P.; Sinha, Arun K.; Katiyar, Ajit K.; Banerjee, Writam; Kundu, Subhas C.; Ray, Samit K.

2013-08-01

340

Transparent and flexible resistive switching memory devices with a very high ON/OFF ratio using gold nanoparticles embedded in a silk protein matrix  

International Nuclear Information System (INIS)

The growing demand for biomaterials for electrical and optical devices is motivated by the need to make building blocks for the next generation of printable bio-electronic devices. In this study, transparent and flexible resistive memory devices with a very high ON/OFF ratio incorporating gold nanoparticles into the Bombyx mori silk protein fibroin biopolymer are demonstrated. The novel electronic memory effect is based on filamentary switching, which leads to the occurrence of bistable states with an ON/OFF ratio larger than six orders of magnitude. The mechanism of this process is attributed to the formation of conductive filaments through silk fibroin and gold nanoparticles in the nanocomposite. The proposed hybrid bio-inorganic devices show promise for use in future flexible and transparent nanoelectronic systems. (paper)

2013-08-30

 
 
 
 
341

Liquid crystals and flow elongation in a spider's silk production line  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Our observations on whole mounted major ampullate silk glands suggested that the thread is drawn from a hyperbolic die using a pre-orientated lyotropic liquid crystalline feedstock. Polarizing microscopy of the gland's duct revealed two liquid crystalline optical textures: a curved pattern in the feedstock within the ampulla of the gland and, later in the secretory pathway, the cellular texture previously identified in synthetic nematic liquid crystals. The behaviour of droplet inclusions wit...

Knight, D. P.; Vollrath, F.

1999-01-01

342

Analysis of a new type of major ampullate spider silk gene, MaSp1s.  

Science.gov (United States)

Spider dragline silk, which is secreted from the major ampullate silk glands, is a unique fibrous protein with a combination of tensile strength and elasticity. Here, we describe a new short type of dragline silk gene, Cyrtophora moluccensis MaSp1s. The full-length gene is only 1320 base pairs (bp), which encodes 439 amino acids that includes the intact non-repetitive N-terminal (149 residues), C-terminal (98 residues) and so-called repetitive regions (192 residues); the deduced molecular weight is approximately 40 kDa. The sequence analysis demonstrated that the two termini are highly homologous to the other characterized dragline silk genes but that the so-called repetitive region is different. Our results suggest that MaSp1s is a possible new characteristic dragline gene; the discovery of this gene should enhance our understanding of the major ampullate spider silk genes. PMID:23403024

Han, Leng; Zhang, Lei; Zhao, Tianfu; Wang, Yujun; Nakagaki, Masao

2013-05-01

343

Pituitary gland  

Science.gov (United States)

The pituitary gland is often referred to as the "master gland" of the body, since it regulates many activities of other endocrine glands. Located above the pituitary gland is the hypothalamus. The hypothalamus decides which ...

344

Supercontraction of dragline silk spun by lynx spiders (Oxyopidae).  

Science.gov (United States)

Supercontraction is commonly considered as a functional adaptation of major ampullate gland (MA) silk to its role as the main structural material in orb-webs. However, the observation of supercontraction in the dragline silk of a lynx spider species, as it is shown in this work, offers a strong support to the hypothesis that the appearance of supercontraction preceded the advent of capture webs. Moreover, the absence of proline in the sequence of dragline silk spidroin in Oxyopidae and related spiders indicates that the presence of this amino acid may not be required for the existence of supercontraction. In this regard, the presence of particular subrepeats--in orb-web and non-orb-web building spiders--adds new clues for the understanding of supercontraction and associated effects. PMID:20359492

Pérez-Rigueiro, J; Plaza, G R; Torres, F G; Hijar, A; Hayashi, C; Perea, G B; Elices, M; Guinea, G V

2010-06-01

345

Untangling spider silk evolution with spidroin terminal domains  

Directory of Open Access Journals (Sweden)

Full Text Available Abstract Background Spidroins are a unique family of large, structural proteins that make up the bulk of spider silk fibers. Due to the highly variable nature of their repetitive sequences, spidroin evolutionary relationships have principally been determined from their non-repetitive carboxy (C-terminal domains, though they offer limited character data. The few known spidroin amino (N-terminal domains have been difficult to obtain, but potentially contain critical phylogenetic information for reconstructing the diversification of spider silks. Here we used silk gland expression data (ESTs from highly divergent species to evaluate the functional significance and phylogenetic utility of spidroin N-terminal domains. Results We report 11 additional spidroin N-termini found by sequencing ~1,900 silk gland cDNAs from nine spider species that shared a common ancestor > 240 million years ago. In contrast to their hyper-variable repetitive regions, spidroin N-terminal domains have retained striking similarities in sequence identity, predicted secondary structure, and hydrophobicity. Through separate and combined phylogenetic analyses of N-terminal domains and their corresponding C-termini, we find that combined analysis produces the most resolved trees and that N-termini contribute more support and less conflict than the C-termini. These analyses show that paralogs largely group by silk gland type, except for the major ampullate spidroins. Moreover, spidroin structural motifs associated with superior tensile strength arose early in the history of this gene family, whereas a motif conferring greater extensibility convergently evolved in two distantly related paralogs. Conclusions A non-repetitive N-terminal domain appears to be a universal attribute of spidroin proteins, likely retained from the origin of spider silk production. Since this time, spidroin N-termini have maintained several features, consistent with this domain playing a key role in silk assembly. Phylogenetic analyses of the conserved N- and C-terminal domains illustrate dramatic radiation of the spidroin gene family, involving extensive duplications, shifts in expression patterns and extreme diversification of repetitive structural sequences that endow spider silks with an unparalleled range of mechanical properties.

Garb Jessica E

2010-08-01

346

Characterization on Eri (Philosamia ricini Fibroin Film: Morphology, Structure and Thermal Properties  

Directory of Open Access Journals (Sweden)

Full Text Available This study aims to prepare Eri Silk Fibroin (SF solution and construct of its film. The Eri cocoons were firstly removed of sericin and then dissolved by 6 M Ca(NO32. The SF solution was dialyzed before measurement of silk percentage by weight. The solvate protein was then taken to prepare of SF film using 0.5 and 1% weight. The 10 mL of SF solution was cast on the polystyrene plates, left in room temperature for 2 days. The obtained SF films were then observed under Scanning Electron Microscope (SEM and Fourier transform infrared spectroscopy (FTIR for morphological and conformational studies, respectively. The results found that Eri SF films were rougher surfaces than Bombyx mori (B. mori SF film. In addition, the film of 0.5% weight SF solution was also rougher surfaces than 1% weight. It contained the ?-helix structure in higher ratio than 1% weight film since the 1% weight SF solution film composed in higher ?-sheet structure. However, the later conformation could affect the SF film to be more hard and brittle than low percent weight content.

Y. Srisuwan

2009-01-01

347

Tarantulas cling to smooth vertical surfaces by secreting silk from their feet.  

Science.gov (United States)

Like all spiders, tarantulas (family Theraphosidae) synthesize silk in specialized glands and extrude it from spinnerets on their abdomen. In one species of large tarantula, Aphonopelma seemanni, it has been suggested that silk can also be secreted from the tarsi but this claim was later refuted. We provide evidence of silk secretion directly from spigots (nozzles) on the tarsi of three distantly related tarantula species: the Chilean rose, Grammostola rosea; the Indian ornamental, Poecilotheria regalis; and the Mexican flame knee, Brachypelma auratum, suggesting tarsal silk secretion is widespread among tarantulas. We demonstrate that multiple strands of silk are produced as a footprint when the spider begins to slip down a smooth vertical surface. The nozzle-like setae on the tarsi responsible for silk deposition have shanks reinforced by cuticular thickenings, which serve to prevent the shanks' internal collapse while still maintaining their flexibility. This is important as the spigots occur on the ventral surface of the tarsus, projecting beyond the finely divided setae of the dry attachment pads. We also reveal the structure and disposition of the silk-secreting spigots on the abdominal spinnerets of the three tarantula species and find they are very similar to those from the earliest known proto-spider spinneret from the Devonian period, giving another indication that silk secretion in tarantulas is close to the ancestral condition. PMID:21562174

Rind, F Claire; Birkett, Chris Luke; Duncan, Benjamin-James A; Ranken, Alexander J

2011-06-01

348

Silk Road policy  

Energy Technology Data Exchange (ETDEWEB)

Landlocked Asia is blessed with rich mineral and energy resources and has great potential to supply these resources to Europe and eastern Asia. With globalisation of markets progressing rapidly, central Asia is now in the spotlight for its economic activity, as a route for international trade and as the heartland of the `Modern Silk Road` of the 21st century. The article discusses the mineral resources of central Asia, covering Kazakhstan, Uzbekistan, Turkmenistan, Kyrghystan, Xingjiang, and Mongolia. Energy resources of the region, coal, oil and natural gas, are mentioned. International organizations and industrial companies like Japan are supporting the transformation of these vast largely unexplored areas to market economies. Construction of the New Silk Road has begun, under the UNDP, and improvement of the Silk Road railway is also underway, with Japanese funding. 3 figs.

Nishikawa, Y. [Mitsui Mineral Development Engineering Co., Tokyo (Japan)

1999-04-01

349

The molecular structures of major ampullate silk proteins of the wasp spider, Argiope bruennichi: a second blueprint for synthesizing de novo silk.  

Science.gov (United States)

The dragline silk of orb-weaving spiders possesses extremely high tensile strength and elasticity. To date, full-length sequences of only two genes encoding major ampullate silk protein (MaSp) in Latrodectus hesperus have been determined. In order to further understand this gene family, we utilized in this study a variety of strategies to isolate full-length MaSp1 and MaSp2 cDNAs in the wasp spider Argiope bruennichi. A. bruennichi MaSp1 and MaSp2 are primarily composed of remarkably homogeneous ensemble repeats containing several complex motifs, and both have highly conserved C-termini and N-termini. Two novel amino acid motifs, GGF and SGR, were found in MaSp1 and MaSp2, respectively. Amino acid composition analysis of silk, luminal contents and predicted sequences indicates that MaSp1 and MaSp2 are two major components of major ampullate glands and that the ratio of MaSp1 to MaSp2 is approximately 3:2 in dragline silk. Furthermore, both the MaSp1:MaSp2 ratio and the conserved termini are closely linked with the production of high quality synthetic fibers. Our results make an important contribution to our understanding of major ampullate silk protein structure and provide a second blueprint for creating new composite silk which mimics natural spider dragline silk. PMID:23262065

Zhang, Yang; Zhao, Ai-Chun; Sima, Yang-Hu; Lu, Cheng; Xiang, Zhong-Huai; Nakagaki, Masao

2013-03-01

350

Silk-hyaluronan-based composite hydrogels: a novel, securable vehicle for drug delivery.  

Science.gov (United States)

A new, biocompatible hyaluronic acid (HA)-silk hydrogel composite was fabricated and tested for use as a securable drug delivery vehicle. The composite consisted of a hydrogel formed by cross-linking thiol-modified HA with poly(ethylene glycol)-diacrylate, within which was embedded a reinforcing mat composed of electrospun silk fibroin protein. Both HA and silk are biocompatible, selectively degradable biomaterials with independently controllable material properties. Mechanical characterization showed the composite tensile strength as fabricated to be 4.43 ± 2.87 kPa, two orders of magnitude above estimated tensions found around potential target organs. In the presence of hyaluronidase (HAse) in vitro, the rate of gel degradation increased with enzyme concentration although the reinforcing silk mesh was not digested. Composite gels demonstrated the ability to store and sustainably deliver therapeutic agents. Time constants for in vitro release of selected representative antibacterial and anti-inflammatory drugs varied from 46.7 min for cortisone to 418 min for hydrocortisone. This biocomposite showed promising mechanical characteristics for direct fastening to tissue and organs, as well as controllable degradation properties suitable for storage and release of therapeutically relevant drugs. PMID:22090427

Elia, Roberto; Newhide, Danny R; Pedevillano, Paul D; Reiss, G Russell; Firpo, Matthew A; Hsu, Edward W; Kaplan, David L; Prestwich, Glenn D; Peattie, Robert A

2013-02-01

351

Silk hydrogels from non-mulberry and mulberry silkworm cocoons processed with ionic liquids.  

Science.gov (United States)

Matrices based on silk fibroin from the non-mulberry silkworm Antheraea mylitta and the mulberry silkworm Bombyx mori have demonstrated good applicability in regenerative medicine. However, the cocoons of A. mylitta are underutilized in part due to their lack of solubility in traditional organic solvents. Therefore, the present work investigates the solubilization and processing of degummed fibers obtained from the cocoons of both silkworm species into hydrogels using ionic liquids (ILs). The developed hydrogels exhibited a rubbery consistency, viscoelastic behavior and rapid degradation in the presence of protease XIV. Scanning electron and confocal microscopy images suggest that human adipose stem cells (hASCs) are able to adhere to and migrate at different levels within the hydrogel structures. Moreover, the MTS assay demonstrated the maintenance of cell metabolic activity for up to 28 days, while DNA quantification showed that hASCs were able to proliferate on the seeded hydrogels. The findings indicate that complete IL removal from the fabricated hydrogels results in a positive hASCs cellular response. Thus the present approach provides a unique opportunity to broaden the processability and application of silk fibroin obtained from A. mylitta cocoons for regenerative medicine, namely cartilage regeneration. PMID:23845228

Silva, Simone S; Popa, Elena G; Gomes, Manuela E; Oliveira, Mariana B; Nayak, Sunita; Subia, Bano; Mano, João F; Kundu, Subhas C; Reis, Rui L

2013-11-01

352

Silk composition in Lepidoptera.  

Czech Academy of Sciences Publication Activity Database

. Londrina : Embrapa Soja, 2000. s. 864.[International Congress of Entomology /21./. 20.08.2000-26.08.2000, Foz do Iguassu]Grant CEP: GA ?R GA204/00/0019Výzkumný zám?r: CEZ:AV0Z5007907Klí?ová slova: lepidopteran silkKód oboru RIV: EB - Genetika a molekulární biologie

Sehnal, František; Žurovec, Michal

353

Structure, composition and mechanical properties of the silk fibres of the egg case of the Joro spider, Nephila clavata (Araneae, Nephilidae).  

Science.gov (United States)

The silk egg case and orb web of spiders are elaborate structures that are assembled from a number of components. We analysed the structure, the amino acid and fibre compositions, and the tensile properties of the silk fibres of the egg case of Nephila clavata. SEM shows that the outer and inner covers of the egg case consist of thick, medium and thin silk fibres. The silk fibres of the outer cover of the egg case are probably produced by the major and minor ampullate glands. The silk fibres of the inner cover of the egg case from cylindrical glands appears to be distinct from the silk fibres of the major ampullate glands based on their micro-morphology, mole percent amino acid composition and types, and tensile behaviour and properties. Collectively, our investigations show that N. clavata uses silk fibres from relatively few glands in varying combinations to achieve different physical and chemical properties (e.g., color, diameter, morphology and amino acid composition) and functional and mechanical properties in the different layers of the egg case. PMID:22116288

Jiang, Ping; Guo, Cong; Lv, Taiyong; Xiao, Yonghong; Liao, Xinjun; Zhou, Bing

2011-12-01

354

Structural Characterization of Minor Ampullate Spidroin Domains and Their Distinct Roles in Fibroin Solubility and Fiber Formation  

Science.gov (United States)

Spider silk is protein fibers with extraordinary mechanical properties. Up to now, it is still poorly understood how silk proteins are kept in a soluble form before spinning into fibers and how the protein molecules are aligned orderly to form fibers. Minor ampullate spidroin is one of the seven types of silk proteins, which consists of four types of domains: N-terminal domain, C-terminal domain (CTD), repetitive domain (RP) and linker domain (LK). Here we report the tertiary structure of CTD and secondary structures of RP and LK in aqueous solution, and their roles in protein stability, solubility and fiber formation. The stability and solubility of individual domains are dramatically different and can be explained by their distinct structures. For the tri-domain miniature fibroin, RP-LK-CTDMi, the three domains have no or weak interactions with one another at low protein concentrations (<1 mg/ml). The CTD in RP-LK-CTDMi is very stable and soluble, but it cannot stabilize the entire protein against chemical and thermal denaturation while it can keep the entire tri-domain in a highly water-soluble state. In the presence of shear force, protein aggregation is greatly accelerated and the aggregation rate is determined by the stability of folded domains and solubility of the disordered domains. Only the tri-domain RP-LK-CTDMi could form silk-like fibers, indicating that all three domains play distinct roles in fiber formation: LK as a nucleation site for assembly of protein molecules, RP for assistance of the assembly and CTD for regulating alignment of the assembled molecules.

Gao, Zhenwei; Lin, Zhi; Huang, Weidong; Lai, Chong Cheong; Fan, Jing-song; Yang, Daiwen

2013-01-01

355

Review the role of terminal domains during storage and assembly of spider silk proteins.  

Science.gov (United States)

Fibrous proteins in nature fulfill a wide variety of functions in different structures ranging from cellular scaffolds to very resilient structures like tendons and even extra-corporal fibers such as silks in spider webs or silkworm cocoons. Despite their different origins and sequence varieties many of these fibrous proteins share a common building principle: they consist of a large repetitive core domain flanked by relatively small non-repetitive terminal domains. Amongst protein fibers, spider dragline silk shows prominent mechanical properties that exceed those of man-made fibers like Kevlar. Spider silk fibers assemble in a spinning process allowing the transformation from an aqueous solution into a solid fiber within milliseconds. Here, we highlight the role of the non-repetitive terminal domains of spider dragline silk proteins during storage in the gland and initiation of the fiber assembly process. PMID:22057429

Eisoldt, Lukas; Thamm, Christopher; Scheibel, Thomas

2012-06-01

356

Spider silk reduces insect herbivory  

Science.gov (United States)

The role of predators in food webs extends beyond their ability to kill and consume prey. Such trait-mediated effects occur when signals of the predator influence the behaviour of other animals. Because all spiders are silk-producing carnivores, we hypothesized that silk alone would signal other arthropods and enhance non-lethal effects of spiders. We quantified the herbivory inflicted by two beetle species on green bean plants (Phaseolus vulgaris) in the presence of silkworm silk and spider silk along with no silk controls. Single leaflets were treated and enclosed with herbivores in the laboratory and field. Another set of leaflets were treated and left to experience natural herbivory in the field. Entire plants in the field were treated with silk and enclosed with herbivores or left exposed to herbivory. In all cases, the lowest levels of herbivory occurred with spider silk treatments and, in general, silkworm silk produced intermediate levels of leaf damage. These results suggest that silk may be a mechanism for the trait-mediated impacts of spiders and that it might contribute to integrated pest management programmes.

Rypstra, Ann L.; Buddle, Christopher M.

2013-01-01

357

Nutritional value of silk powder from irradiated silk waste  

International Nuclear Information System (INIS)

Silk waste from Thai reeling factory was developed into purified silk protein. Cleanliness of silk filaments by boiling in water was firstly needed. After air drying, they were irradiated with Gamma ray using Cobalt-60 at doses of 750 and 1,000 kGy, then the irradiated silk filaments were dissolved in calcium chloride-ethanol-water solution. The next steps would be dialysis followed by freezed dried process to obtain irradiated silk powder. Two samples of 750 and 1,000 kGy irradiated silk powder were analyzed for nutritional components compared with non irradiated silk filaments. It was found that, the fat content in two irradiated samples was distinctive lower than the non irradiated one, however the protein content was nearly the same in three samples. In addition the moisture content in two irradiated samples was distinctive higher than the non irradiated one. These results show that irradiation technique is useful for development of valuable silk protein as biomaterial. (author)

2002-03-01

358

The science of silks  

Digital Repository Infrastructure Vision for European Research (DRIVER)

Comparative silk research has begun to provide us with valuable insights into a class of biopolymers that have evolved an enormous range of material performance based on highly adapted structure-property interactions. As outlined in this article, such insights cover the analysis of biological and bioinspired spinning technologies, the discovery of a novel type of melt spinning at ambient temperatures, and an advanced predictive modeling framework that uses ab initio calculations. Importantly,...

Vollrath, F.; Porter, D.; Holland, C.

2013-01-01

359

Molecular spring: from spider silk to silkworm silk  

CERN Document Server

In this letter, we adopt a new approach combining theoretical modeling with silk stretching measurements to explore the mystery of the structures between silkworm and spider silks, leading to the differences in mechanical response against stretching. Hereby the typical stress-strain profiles are reproduced by implementing the newly discovered and verified "$\\beta$-sheet splitting" mechanism, which primarily varies the secondary structure of protein macromolecules; our modeling and simulation results show good accordance with the experimental measurements. Hence, it can be concluded that the post-yielding mechanical behaviors of both kinds of silks are resulted from the splitting of crystallines while the high extensibility of spider dragline is attributed to the tiny $\\beta$-sheets solely existed in spider silk fibrils. This research reveals for the first time the structural factors leading to the significant difference between spider and silkworm silks in mechanical response to the stretching force. Addition...

Wu, Xiang; Du, Ning; Xu, Gang-Qin; Li, Bao-Wen

2009-01-01

360

Thermovoltaic properties of hornet silk.  

Science.gov (United States)

In silk from the larval silk caps of the Oriental hornet Vespa orientalis (Hymenoptera, Vespinae), temperature-dependent changes in the electric voltage have been recorded, with rise in the voltage occurring mainly upon rise in the temperature between 10-36 degrees C. The peak voltage was measured between 32-38 degrees C and attained 240-360 mV, but with further increase in temperature, the voltage decreased, dropping to 0 mV at about 45-50 degrees C. Upon second measurement (of same silk specimen), the voltage peak usually occurred later (by 8-9 degrees C) and at higher temperature than in the first measurement. Continuous measurements during warming up to 30 degrees C followed by cooling down to 15 degrees C yielded an hysteresis between the warming "line" and the cooling "line", the former often straight and the latter usually curved. Maintaining the silk specimen at a fixed temperature for a prolonged period (hours) initially causes the voltage to rise, then remain steady, and finally drop. Boiling the silk caps in tap water for 7-10 min exerts some changes in the silk properties, mainly a decrease in voltage level. The general behavior of the silk suggests that it is a polymer endowed with the qualities of an organic semiconductor. The various properties of the larval silk are discussed in great detail. PMID:17029238

Volynchik, S; Plotkin, M; Ermakov, N Y; Bergman, D J; Ishay, J S

2006-11-01

 
 
 
 
361

Recombinant DNA production of spider silk proteins.  

Science.gov (United States)

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

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

2013-11-01

362

Antheraea pernyi Silk Fiber: A Potential Resource for Artificially Biospinning Spider Dragline Silk  

Digital Repository Infrastructure Vision for European Research (DRIVER)

The outstanding properties of spider dragline silk are likely to be determined by a combination of the primary sequences and the secondary structure of the silk proteins. Antheraea pernyi silk has more similar sequences to spider dragline silk than the silk from its domestic counterpart, Bombyx mori. This makes it much potential as a resource for biospinning spider dragline silk. This paper further verified its possibility as the resource from the mechanical properties and the structures of t...

Yaopeng Zhang; Hongxia Yang; Huili Shao; Xuechao Hu

2010-01-01

363

Determination of the torsion angles of alanine and glycine residues of model compounds of spider silk (AGG)10 using solid-state NMR methods  

International Nuclear Information System (INIS)

Spiders synthesize several kinds of silk fibers. In the primary structure of spider silk, one of the major ampullate (dragline, frame) silks, spidroin 1, and flagelliform silk (core fibers of adhesive spiral), there are common repeated X-Gly-Gly (X = Ala, Leu, Pro, Tyr, Glu, and Arg) sequences, which are considered to be related to the elastic character of these fibers. In this paper, two dimensional spin diffusion solid-state NMR under off magic angle spinning (OMAS), 13C chemical shift contour plots, and Rotational Echo DOuble Resonance (REDOR) were applied to determine the torsion angles of one Ala and two kinds of Gly residues in the Ala-Gly-Gly sequence of 13C=O isotope-labeled (Ala-Gly-Gly)10. The torsion angles were determined to be (?, ?) = (-90 deg., 150 deg.) within an experimental error of ±10 deg. for each residue. This conformation is characterized as 31 helix which is in agreement with the structure proposed from the X-ray powder diffraction pattern of poly(Ala-Gly-Gly). The 31 helix of (Ala-Gly-Gly)10 does not change by formic acid treatment although (Ala-Gly)15 easily changes from the silk I conformation (the structure of Bombyx mori silk fibroin before spinning in the solid state) to silk II conformation (the structure of the silk fiber after spinning) by such treatment. Thus, the 31 helix conformation of (Ala-Gly-Gly)10 is considered very stable. Furthermore, the torsion angles of the 16th Leu residue of (Leu-Gly-Gly)10 were also determined as (?, ?) = (-90 deg., 150 deg.) and this peptide is also considered to take 31 helix conformation

2003-02-01

364

Determination of the torsion angles of alanine and glycine residues of model compounds of spider silk (AGG){sub 10} using solid-state NMR methods  

Energy Technology Data Exchange (ETDEWEB)

Spiders synthesize several kinds of silk fibers. In the primary structure of spider silk, one of the major ampullate (dragline, frame) silks, spidroin 1, and flagelliform silk (core fibers of adhesive spiral), there are common repeated X-Gly-Gly (X = Ala, Leu, Pro, Tyr, Glu, and Arg) sequences, which are considered to be related to the elastic character of these fibers. In this paper, two dimensional spin diffusion solid-state NMR under off magic angle spinning (OMAS), {sup 13}C chemical shift contour plots, and Rotational Echo DOuble Resonance (REDOR) were applied to determine the torsion angles of one Ala and two kinds of Gly residues in the Ala-Gly-Gly sequence of {sup 13}C=O isotope-labeled (Ala-Gly-Gly){sub 10}. The torsion angles were determined to be ({phi}, {psi}) = (-90 deg., 150 deg.) within an experimental error of {+-}10 deg. for each residue. This conformation is characterized as 3{sub 1} helix which is in agreement with the structure proposed from the X-ray powder diffraction pattern of poly(Ala-Gly-Gly). The 3{sub 1} helix of (Ala-Gly-Gly){sub 10} does not change by formic acid treatment although (Ala-Gly){sub 15} easily changes from the silk I conformation (the structure of Bombyx mori silk fibroin before spinning in the solid state) to silk II conformation (the structure of the silk fiber after spinning) by such treatment. Thus, the 3{sub 1} helix conformation of (Ala-Gly-Gly){sub 10} is considered very stable. Furthermore, the torsion angles of the 16th Leu residue of (Leu-Gly-Gly){sub 10} were also determined as ({phi}, {psi}) = (-90 deg., 150 deg.) and this peptide is also considered to take 3{sub 1} helix conformation.

Ashida, Jun; Ohgo, Kosuke; Komatsu, Kohei; Kubota, Ayumi; Asakura, Tetsuo [Tokyo University of Agriculture and Technology, Department of Biotechnology (Japan)], E-mail: asakura@cc.tuat.ac.jp

2003-02-15

365

Split gland  

Energy Technology Data Exchange (ETDEWEB)

A split gland having only three parts is described. The gland has substantially the same stability to the relative motion of the constituent half-gland members during the attachment process to a female fitting as have more complicated designs. Ease of manufacture and use result from the reduction in complexity of the present invention.

Petranto, Joseph J. (Los Alamos, NM)

1989-01-01

366

Split gland  

Energy Technology Data Exchange (ETDEWEB)

A split gland having only three parts is described. The gland has substantially the same stability to the relative motion of the constituent half-gland members during the attachment process to a female fitting as have more complicated designs. Ease of manufacture and use result from the reduction in complexity of the present invention. 15 figs.

Petranto, J.J.

1989-09-05

367

?-Sheet nanocrystalline domains formed from phosphorylated serine-rich motifs in caddisfly larval silk: a solid state NMR and XRD study.  

Science.gov (United States)

Adhesive silks spun by aquatic caddisfly (order Trichoptera) larvae are used to build both intricate protective shelters and food harvesting nets underwater. In this study, we use (13)C and (31)P solid-state NMR and wide angle X-ray diffraction (WAXD) as tools to elucidate molecular protein structure of caddisfly larval silk from the species Hesperophylax consimilis . Caddisfly larval silk is a fibroin protein based biopolymer containing mostly repetitive amino acid motifs. NMR and X-ray results provide strong supporting evidence for a structural model in which phosphorylated serine repeats (pSX)4 complex with divalent cations Ca(2+) and Mg(2+) to form rigid nanocrystalline ?-sheet structures in caddisfly silk. (13)C NMR data suggests that both phosphorylated serine and neighboring valine residues exist in a ?-sheet conformation while glycine and leucine residues common in GGX repeats likely reside in random coil conformations. Additionally, (31)P chemical shift anisotropy (CSA) analysis indicates that the phosphates on phosphoserine residues are doubly ionized, and are charge-stabilized by divalent cations. Positively charged arginine side chains also likely play a role in charge stabilization. Finally, WAXD results finds that the silk is at least 7-8% crystalline, with ?-sheet interplane spacings of 3.7 and 4.5 Å. PMID:23452243

Addison, J Bennett; Ashton, Nicholas N; Weber, Warner S; Stewart, Russell J; Holland, Gregory P; Yarger, Jeffery L

2013-04-01

368

Mechanical improvements to reinforced porous silk scaffolds.  

Science.gov (United States)

Load-bearing porous biodegradable scaffolds are required to engineer functional tissues such as bone. Mechanical improvements to porogen leached scaffolds prepared from silk proteins were systematically studied through the addition of silk particles in combination with silk solution concentration, exploiting interfacial compatibility between the two components. Solvent solutions of silk up to 32 w/v % were successfully prepared in hexafluoroisopropanol (HFIP) for the study. The mechanical properties of the reinforced silk scaffolds correlated to the material density and matched by a power law relationship, independent of the ratio of silk particles to matrix. These results were similar to the relationships previously shown for cancellous bone. From these data we conclude that the increased mechanical properties were due to a densification effect and not due to the inclusion of stiffer silk particles into the softer silk matrix. A continuous interface between the silk matrix and the silk particles, as well as homogeneous distribution of the silk particles within the matrix was observed. Furthermore, we note that the roughness of the pore walls was controllable by varying the ratio of the particles matrix, providing a route to control topography. The rate of proteolytic hydrolysis of the scaffolds decreased with increase in mass of silk used in the matrix and with increasing silk particle content. PMID:21793193

Gil, Eun Seok; Kluge, Jonathan A; Rockwood, Danielle N; Rajkhowa, Rangam; Wang, Lijing; Wang, Xungai; Kaplan, David L

2011-10-01

369

Effect of Pretreatment on Fabrication of Natural FibroinFiber/Apatite Composites Using Alternate Soaking Method  

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Full Text Available To develop biomimetic fibroin/apatite composites with bone―like structure byalternate soaking method, the degummed natural fibroin fibers were pretreatedby soaking into CaCl2:EtOH:H2O (<>n(CaCl2):<>n(EtOH):<>n(H2O)=1:2:8) solution for given time and then soaked into calciumsolution and phosphate solution, respectively, to fabricate fibroin/apatite composites. The effects of pretreatment time andthe alternate soaking times on fibroin microstructure and fibroin/apatitecomposites were examined by SEM, FTIR, XRD and TGA techniques. The pretreatmentusing CaCl2:EtOH:H2O(<>n(CaCl2):<>n(EtOH):<>n(H2O)=1:2:8) solution makes fibroin fiber exhibit rougher surface and higher internalporosity, which is beneficial to interaction of calcium ion and residue groupssuch as ―COOH, –OHand ―CONH― from fibroin, and then improvethe deposition of apatite on fibroin fibers. The homogeneous apatites withoriented <>c―axis were formed on the pretreated fibroin fibers. Moreover,there are more than 30wt% apatites deposited on fibroin fibers after alternatesoaking for more than 3 times. The fibroin/apatite composite has potential tobe applied as bone repair materials.

LI Zhi-Hong, WU Ji-Min, HUANG Shu-Jie, GUAN Jing, ZHANGXi-Zheng

2010-07-01

370

Air filter devices including nonwoven meshes of electrospun recombinant spider silk proteins.  

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Based on the natural sequence of Araneus diadematus Fibroin 4 (ADF4), the recombinant spider silk protein eADF4(C16) has been engineered. This highly repetitive protein has a molecular weight of 48kDa and is soluble in different solvents (hexafluoroisopropanol (HFIP), formic acid and aqueous buffers). eADF4(C16) provides a high potential for various technical applications when processed into morphologies such as films, capsules, particles, hydrogels, coatings, fibers and nonwoven meshes. Due to their chemical stability and controlled morphology, the latter can be used to improve filter materials. In this protocol, we present a procedure to enhance the efficiency of different air filter devices, by deposition of nonwoven meshes of electrospun recombinant spider silk proteins. Electrospinning of eADF4(C16) dissolved in HFIP results in smooth fibers. Variation of the protein concentration (5-25% w/v) results in different fiber diameters (80-1,100 nm) and thus pore sizes of the nonwoven mesh. Post-treatment of eADF4(C16) electrospun from HFIP is necessary since the protein displays a predominantly ?-helical secondary structure in freshly spun fibers, and therefore the fibers are water soluble. Subsequent treatment with ethanol vapor induces formation of water resistant, stable ?-sheet structures, preserving the morphology of the silk fibers and meshes. Secondary structure analysis was performed using Fourier transform infrared spectroscopy (FTIR) and subsequent Fourier self-deconvolution (FSD). The primary goal was to improve the filter efficiency of existing filter substrates by adding silk nonwoven layers on top. To evaluate the influence of electrospinning duration and thus nonwoven layer thickness on the filter efficiency, we performed air permeability tests in combination with particle deposition measurements. The experiments were carried out according to standard protocols. PMID:23685883

Lang, Gregor; Jokisch, Stephan; Scheibel, Thomas

2013-01-01

371

THE INHERENT ACIDIC CHARACTERISTICS OF SILK, PART II – WEIGHTED SILKS  

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Full Text Available Silk is found in many textiles of cultural and historic significance,but these fibres may deteriorate rapidly underadverse conditions, so are of particular concern to conservatorsand curators. Measurement of pH is common practicewhen assessing condition, as enhanced acidity is oftenascribed to previous processing and treatment, adsorbedpollutants and oxidative deterioration, but recent researchhas demonstrated that the increased acidity of silk is anexpected consequence of ageing, irrespective of the causeof deterioration. The study presented here builds on thisearlier work to assess the role that common silk processingmethods (bleaching and metal salt weighting have onthis phenomenon. Tensile strength was used as a marker ofdeterioration, independent of mechanism, and was comparedwith pH values of saline extracts of various silk samples.These samples included surrogates, prepared usinghistorically accurate methods and subject to acceleratedageing, as well as a range of historic materials. It was seenthat most samples followed the same correlation betweenpH and strength as seen in the previous work. However,silks weighted with tin salts appeared to be buffered (in therange pH 6.75-7.05, irrespective of their mechanicalintegrity.

Paul Garside

2010-01-01

372

Diversity of molecular transformations involved in the formation of spider silks.  

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Spiders that spin orb webs secrete seven types of silk. Although the spinning process of the dragline thread is beginning to be understood, the molecular events that occur in spiders' opisthosomal glands, which produce the other fibers, are unknown due to a lack of data regarding their initial and final structures. Taking advantage of the efficiency of Raman spectromicroscopy in investigating micrometer-sized biological samples, we have determined the secondary structure of proteins in the complete set of glands of the orb-weaving spider Nephila clavipes. The major and minor ampullate silks in the sac of their glands have identical secondary structures typical of natively unfolded proteins. Spidroins are converted into fibers containing highly oriented ?-sheets. The capture spiral represents a distinct structural singleton. The proteins are highly disordered prior to spinning and undergo no molecular change or alignment upon spinning. The cylindrical, aciniform, and piriform proteins are folded in their initial state with a predominance of ?-helices, but whereas the cylindrical gland forms a fiber similar to the major ampullate thread, the aciniform and piriform glands produce fibers dominated by moderately oriented ?-sheets and ?-helices. The conformation of the proteins before spinning is related to intrinsic characteristics of their primary structure. Proteins that are unfolded in the gland have repeat sequences composed of submotifs and display no sequence regions with aggregation propensity. By contrast, the folded proteins have neither submotifs nor aggregation-prone sequence regions. Taken together, the Raman data show a remarkable diversity of molecular transformations occurring upon spinning. PMID:21050860

Lefèvre, Thierry; Boudreault, Simon; Cloutier, Conrad; Pézolet, Michel

2011-01-01

373

A silk hydrogel-based delivery system of bone morphogenetic protein for the treatment of large bone defects.  

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The use of tissue grafting for the repair of large bone defects has numerous limitations including donor site morbidity and the risk of disease transmission. These limitations have prompted research efforts to investigate the effects of combining biomaterial scaffolds with biochemical cues to augment bone repair. The goal of this study was to use a critically-sized rat femoral segmental defect model to investigate the efficacy of a delivery system consisting of an electrospun polycaprolactone (PCL) nanofiber mesh tube with a silk fibroin hydrogel for local recombinant bone morphogenetic protein 2 (BMP-2) delivery. Bilateral 8 mm segmental femoral defects were formed in 13-week-old Sprague Dawley rats. Perforated electrospun PCL nanofiber mesh tubes were fitted into the adjacent native bone such that the lumen of the tubes contained the defect (Kolambkar et al., 2011b). Silk hydrogels with or without BMP-2 were injected into the defect. Bone regeneration was longitudinally assessed using 2D X-ray radiography and 3D microcomputed topography (?CT). Following sacrifice at 12 weeks after surgery, the extracted femurs were either subjected to biomechanical testing or assigned for histology. The results demonstrated that silk was an effective carrier for BMP-2. Compared to the delivery system without BMP-2, the delivery system that contained BMP-2 resulted in more bone formation (p<0.05) at 4, 8, 12 weeks after surgery. Biomechanical properties were also significantly improved in the presence of BMP-2 (p<0.05) and were comparable to age-matched intact femurs. Histological evaluation of the defect region indicated that the silk hydrogel has been completely degraded by the end of the study. Based on these results, we conclude that a BMP-2 delivery system consisting of an electrospun PCL nanofiber mesh tube with a silk hydrogel presents an effective strategy for functional repair of large bone defects. PMID:22658161

Diab, Tamim; Pritchard, Eleanor M; Uhrig, Brent A; Boerckel, Joel D; Kaplan, David L; Guldberg, Robert E

2012-07-01

374

A silk hydrogel-based delivery system of bone morphogenic protein for the treatment of large bone defects  

Science.gov (United States)

The use of tissue grafting for the repair of large bone defects has numerous limitations including donor site morbidity and the risk of disease transmission. These limitations have prompted research efforts to investigate the effects of combining biomaterial scaffolds with biochemical cues to augment bone repair. The goal of this study was to use a critically-sized rat femoral segmental defect model to investigate the efficacy of a delivery system consisting of an electrospun polycaprolactone (PCL) nanofiber mesh tube with a silk fibroin hydrogel for local recombinant bone morphogenetic protein 2 (BMP-2) delivery. Bilateral 8 mm segmental femoral defects were formed in 13-week-old Sprague Dawley rats. Perforated electrospun PCL nanofiber mesh tubes were fitted into the adjacent native bone such that the lumen of the tubes contained the defect (Kolambkar et al., 2011b). Silk hydrogels with or without BMP-2 were injected into the defect. Bone regeneration was longitudinally assessed using 2D X-ray radiography and 3D microcomputed topography (µCT). Following sacrifice at 12 weeks after surgery, the extracted femurs were either subjected to biomechanical testing or assigned for histology. The results demonstrated that silk was an effective carrier for BMP-2. Compared to the delivery system without BMP-2, the delivery system that contained BMP-2 resulted in more bone formation (p < 0.05) at 4, 8, 12 weeks after surgery. Biomechanical properties were also significantly improved in the presence of BMP-2 (p < 0.05) and were comparable to age-matched intact femurs. Histological evaluation of the defect region indicated that the silk hydrogel have completely been degraded by the end of the study. Based on these results, we conclude that a BMP-2 delivery system consisting of an electrospun PCL nanofiber mesh tube with a silk hydrogel presents an effective strategy for functional repair of large bone defects.

Diab, Tamim; Pritchard, Eleanor M.; Uhrig, Brent A.; Boerckel, Joel D.; Kaplan, David L.; Guldberg, Robert E.

2011-01-01

375

Effects of ultraviolet rays and acid rain on strength of spider silk; Kumo ito no kyodo ni oyobosu shigaisen oyobi sanseiu no eikyo  

Energy Technology Data Exchange (ETDEWEB)

Spider silk, differing to the synthetic fibers yarned at high temperature or using hazardous solvents, is a fiber in the body of animal and the process itself is safe. Again, the silk is made of up protein and is kind to the environment because does not cause pollution even after disposal. Also, it is a material of interest as a strong fiber having high tensile strength reaching to 2000 MPa. Accordingly, spider silk can be taken as an effective industrial material if the mass production of the fiber is possible by genetic engineering. In previous report, the mechanical properties of dragline silk spewed out from the same silk gland as the warp, a kind of silk spewed out by Nephila clavata, so called Jorogumo, were studied. In this report, the deformation behavior of circular net made by Jorogumo and applicability to environmental evaluation are under study, and effect of acid environment and ultraviolet rays on the mechanical properties of spider silk are studied. As a result, it was revealed that ultraviolet rays degraded the spider silk, and further, promoted the degradation when the acid rain acted at the same time. 16 refs., 11 figs.

Kitagawa, M.; Sasagawa, H. [Kanazawa Univ., Ishikawa (Japan). Faculty of engineering; Kawagoe, M. [Toyama Prefectural Univ., Toyama (Japan)

1995-12-15

376

Silk structure in the hornet cocoon.  

Science.gov (United States)

This study deals with the development of a silk coat around the growing larva of the Oriental hornet (Vespa orientalis). Prior to its metamorphosis into a pupa, the larva secretes a silk weave, which enwraps it completely. This silk coat is not uniform throughout, but rather varies along the extent of the larval body. First, it is possible to discern in the spun silk weave fibers whose thickness is approximately 10 microm; in which case such a fiber is actually a duplet of two individual fibers, each with its own silk coat and an inner fibril (core). Additionally, there are silk plaque-like surfaces of a greater diameter and of variable shape, depending on the contour that they cover. In the cap region, i.e. the region where the head of the larva (and later of the developing imago) is located, the fibers are quite numerous and arranged in many (up to 10) layers, while the silk plaques are small. For the remainder of its body, the pupating larva produces a silk sleeve in which there are few fibers with numerous plaques in between, occupying most of the area. The larval silk fibers proper are of fairly uniform diameter. Apart from the pupating larvae, the adult hornets also secrete a type of silk, which acts as a glue holding together the components of the larval cell wall and their sticky silk is of variable thickness, depending on the configuration of the cell. The temperature of the silk varies in accordance with age of the pupating larva. Thus, the closer the developing hornet to eclosion, the lower the temperature. The present study reviews and summarizes this transformation process in the silk weave from its inception until eclosion of the imago. PMID:15332657

Joseph, Zmira; Ishay, Jacob S

2004-01-01

377

Thromboelastometric and platelet responses to silk biomaterials.  

Science.gov (United States)

Silkworm's silk is natural biopolymer with unique properties including mechanical robustness, a