Mulberry non-engineered silk gland protein vis-à-vis silk cocoon protein engineered by silkworms as biomaterial matrices

Silk fibroin from silk gland of Bombyx mori 5th instar larvae was utilized to fabricate films, which may find possible applications as two-dimensional matrices for tissue engineering. Bombyx mori cocoon fibroin is well characterized as potential biomaterial by virtue of its good mechanical strength, water stability, thermal properties, surface roughness and biocompatibility. The present study aims to characterize the biophysical, thermal, mechanical, rheological, swelling properties along with spectroscopic analysis, surface morphology and biocompatibility of the silk gland fibroin films compared with cocoon fibroin. Fibroin solutions showed increased turbidity and shear thinning at higher concentration. The films after methanol treatment swelled moderately and were less hydrophilic compared to the untreated. The spectroscopic analysis of the films illustrated the presence of various amide peaks and conformational transition from random coil to beta sheet on methanol treatment. X-ray diffraction studies also confirmed the secondary structure. Thermogravimetric analysis showed distinct weight loss of the films. The films were mechanically stronger and AFM studies showed surfaces were rougher on methanol treatment. The matrices were biocompatible and supported L929 mouse fibroblast cell growth and proliferation. The results substantiate the silk gland fibroin films as potential biomaterial matrices.     

Design,fabrication and characterization of silk fibroin-HPMC-PEG blended films as vehicle for transmucosal delivery

This study illustrates the fabrication of stable mucoadhesive films of silk protein fibroin as potential vehicle for transmucosal delivery by blending fibroin with hydroxy propyl methyl cellulose (HPMC) and poly ethylene glycol 400 (PEG). Investigations on mechanical properties, swelling ability in simulated saliva, bioadhesive strength by a specially designed instrument and study of in vitro stability in simulated saliva of goat buccal mucosa as model membrane was undertaken. Molecular interaction between blended materials was evaluated by FTIR spectroscopy. Increase in fibroin content of the blended films not only increased the mechanical properties and water stability but also the degree of swelling and stability of the films in simulated saliva. The FTIR spectrum shows an increase in water stability of the fibroin-HPMC blended films due to the formation of intermolecular hydrogen bonding between the HPMC and fibroin. The conformational transition of the silk fibroin molecule from the amorphous and random coil to β sheet structure has been observed. Fibroin-HPMC-PEG blended films can be used as a vehicle for transmucosal delivery by virtue of its good mechanical strength, water stability, ex vivo bioadhesive strength and ideal swellability as such characteristics are essential for rapid mucoadhesion.     

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

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.     

Surface modification and functionalization of silk fibroin fibers/fabric toward high performance applications

Silk from silkworms is composed of fibrous proteins with remarkable properties that have resulted in wide usage in the textile industry and as a biomaterial in the medical field. However, for some advanced applications, silk still has the disadvantages of wrinkling, photo-induced aging, deformation and even degradation caused by microorganism, biocompatibility problems. Based on sufficient supply of raw materials around the world and encouraged by the market demands for natural and smart materials, numerous attempts have been made in the surface modification and functionalization of silk fibroin fibers (SFFs) and silk fibroin fabric (SFF) using a wide range of functional materials and technology during last decade. Successful modification of silk may not only overcome its intrinsic shortcomings, but also enhance end-use performance. In this review, we summarize the recent progress of basic ideas, methodologies, and treatment techniques for surface modification and functionalization of SFF/SFFs with consideration of their enhanced properties and potential applications. Encouraged by achievements from experimental studies, the authors believe that such successes give silk products a promising future in both textile and biomaterial applications.     

The preparation of insoluble fibroin films induced by degummed fibroin or fibroin microspheres

Blending degummed fibroin (DF) or insoluble fibroin microspheres with concentrated fibroin solution, the insoluble films were obtained through drying the solution at 40–45 ^∘C. The conformation of silk fibroin films was analyzed by infrared spectrum and X-ray diffractometry. The results demonstrated that β-sheet conformation increased rapidly when the degummed silk or insoluble microspheres blended with fibroin, while the pure SF membrane was mainly composed of α/random coil conformation when the other conditions kept same. This suggested that fibroin microspheres and degummed fibroin could induce the formation of β-sheet crystal and the insoluble films, without methanol after-treatment, could be obtained at approximately room temperature. Although the fibroin films blending with DF had many protuberances, the films containing fibroin microspheres had the smooth surface and could be used effectively in biotechnological materials and biomedical application.     

Interfacial bonding and degumming effects on silk fibre/polymer biocomposites

Silk fibre has been popularly used for bio-medical engineering and surgically-operational applications for centuries because of its biocompatible and bioresorbable properties. Using silk fibre as reinforcement for bio-polymers could enhance the stiffness of scaffoldings and bone implants. However, raw silk fibre consists of silk fibroin that is bound together by a hydrophilic glued-liked protein layer called “sericin”. Degumming is a surface modification process for sericin removal which allows a wide control of the silk fibre’s properties, making the silk fibre possible to be properly used for the development and production of novel bio-composites with specific mechanical and biodegradable properties. Some critical issues such as wettability, bonding efficiency and biodegradability at the fibre/matrix interface are of interesting topics in the study of the degumming process. Therefore, it is a need to detailedly study the effect on different degumming processes to the properties of the silk fibre for real-life applications.     

丝素共混膜的结构、性能及应用研究进展

丝素膜是一种性能优良、用途广泛的天然高分子生物材料、但纯丝素膜存在着在干燥情况下力学性能很差等缺点，这些缺点可以通过与其他高分子化合物共混得到改善，从而拓宽丝素膜的应用范围，综述了丝素与其它天然或合成高分子形成的共混膜的种类，结构、性能及应用发展状况。     

丝素蛋白柔性电子器件材料的研究进展

丝素蛋白材料凭借良好的生物相容性、可控生物降解性、再生形貌多样性等已被制成柔性电子器件在电子领域进行了应用研究.本文首先综述不同溶解方法对蚕丝再生材料制备的影响,同时对丝素蛋白材料的(微球、膜、纤维、凝胶、支架等)制备方法、材料性能进行分析,最后总结了近年来丝素蛋白基柔性电子材料的应用研究进展.尽管已有研究表明可获得各...     

A quicker degradation rate is yielded by a novel kind of transgenic silk fibroin consisting of shortened silk fibroin heavy chains fused with matrix metalloproteinase cleavage sites

Degradation performance of silk fibroin is an important property for its medical applications. Herein we constructed a shortened silk fibroin heavy chain protein fused with a matrix metalloproteinase cleavage site (SSFH-MMP) along with a glutathione S-transferase tag ahead. The digestion assay shows it can be cut by matrix metalloproteinase-2 (MMP-2) at its MMP cleavage site. Furthermore, we introduced the SSFH-MMP into silk fibroin by genetic modification of silkworms in order to increase the degradation rate of the silk fibroin. After acquisition of a race of transgenic silkworms with the coding sequence of the MMP cleavage site in their genomic DNA, we tested some properties of their silk fibroin designated TSF-MMP. The results show that the TSF-MMP has MMP cleavage sites and yields a quicker degradation rate during dilution in MMP-2 enzyme buffer or implantation into tumor tissues compared with that of normal silk fibroin. Moreover, the TSF-MMP is in vitro non-toxic to human bone marrow mesenchymal stem cells (hBM-MSCs) indicating that the TSF-MMP may become a biomaterial with a quicker degradation rate for its medical applications.     

Bovine pericardium coated with biopolymeric films as an alternative to prevent calcification: In vitro calcification and cytotoxicity results

Bovine pericardium, for cardiac valve fabrication, was coated with either chitosan or silk fibroin film. In vitro calcification tests of coated and non coated bovine pericardium were performed in simulated body fluid solution in order to investigate potential alternatives to minimize calcification on implanted heart valves. Complementary, morphology was assessed by scanning electron microscopy — SEM; X-ray diffraction (XRD) and infrared spectroscopy (FTIR-ATR) were performed for structural characterization of coatings and biocompatibility of chitosan. Silk fibroin films were assayed by in vitro cytotoxicity and endothelial cell growth tests. Bovine pericardium coated with silk fibroin or chitosan did not present calcification during in vitro calcification tests, indicating that these biopolymeric coatings do not induce bovine pericardium calcification. Chitosan and silk fibroin films were characterized as non cytotoxic and silk fibroin films presented high affinity to endothelial cells. The results indicate that bovine pericardium coated with silk fibroin is a potential candidate for cardiac valve fabrication, since the affinity of silk fibroin to endothelial cells can be explored to induce the tissue endothelization and therefore, increase valve durability by increasing their mechanical resistance and protecting them against calcification.     

Assembly and degradation of low-fouling click-functionalized poly(ethylene glycol)-based multilayer films and capsules

Nano-/micrometer-scaled films and capsules made of low-fouling materials such as poly(ethylene glycol) (PEG) are of interest for drug delivery and tissue engineering applications. Herein, the assembly and degradation of low-fouling, alkyne-functionalized PEG (PEG(Alk) ) multilayer films and capsules, which are prepared by combining layer-by-layer (LbL) assembly and click chemistry, are reported. A nonlinear, temperature-responsive PEG(Alk) is synthesized, and is then used to form hydrogen-bonded multilayers with poly(methacrylic acid) (PMA) at pH 5. The thermoresponsive behavior of PEG(Alk) is exploited to tailor film buildup by adjusting the assembly conditions. Using alkyne-azide click chemistry, PEG(Alk)/PMA multilayers are crosslinked with a bisazide linker that contains a disulfide bond, rendering these films and capsules redox-responsive. At pH 7, by disrupting the hydrogen bonding between the polymers, PEG(Alk) LbL films and PEG(Alk) -based capsules are obtained. These films exhibit specific deconstruction properties under simulated intracellular reducing conditions, but remain stable at physiological pH, suggesting potential applications in controlled drug release. The low-fouling properties of the PEG films are confirmed by incubation with human serum and a blood clot. Additionally, these capsules showed negligible toxicity to human cells.     

Flexible bio-composites based on silks and celluloses

Biomaterials have attracted worldwide attention due to the concerns regarding health and the environment. Silk, a natural protein produced by several species of insects, has been examined as a potential material for applications in many biotechnological and biomedical fields. However, regenerated silk fibroin has poor ductility and mechanical properties. Therefore, in this study, silk fibroin-cellulose composite films were prepared in an aqueous system to increase the ductility of regenerated silk fibroin. The morphology of the silk fibroin-cellulose composite film was observed by field emission scanning electron microscopy. The structure of the silk fibroin-cellulose composite films was examined by Fourier transform-infrared spectroscopy. The flexibility was analyzed using a bending test.     

丝素蛋白膜表面的等离子体磺酸化及体外抗凝血性能

从蚕丝中提取的丝素蛋白（Silk fibroin）制备成薄膜,首次采用二氧化硫等离子体处理在材料表现引入了磺酸基因;同时丝素蛋白膜用氨气等离子体处理后在表面接入氨基,利用1,3－丙磺酸内酯与氨基的反应在材料表面接枝磺酸。采用X光电子能谱和全反射红光谱分析材料的表面性质。材料的体外抗弟血性能由外凝血时间;凝血酶原时间（PT）、部分凝血活酶时间（APTT）和凝血酶时间（TT）作为评价标准。结果表明两种处理方法均能在丝素蛋白膜表面有效地接枝磺酸基因,而且材料的抗凝血性能有显著提高。     

Stem cell response to multiwalled carbon nanotube-incorporated regenerated silk fibroin films

Multiwalled carbon nanotubes (MWCNTs) are considered to be the ideal reinforcements for biorelated applications on account of their remarkable structural, mechanical and thermal properties. However, before MWCNTs can be incorporated into new and existing biomedical devices, their toxicity and biocompatibility need to be investigated thoroughly. In this study, regenerated silk fibroin/MWCNT nanocomposite films were prepared using a solvent system with pre-dispersed MWCNTs. Their biocompatibility was examined in vitro using human bone marrow stem cells. Scanning electron microscopy and a WST-1 assay demonstrated that the silk fibroin/MWCN film supported BMSC attachment and growth over 7 days in culture similar to the silk fibroin only film.     

Synthesis of selenium nanoparticles in the presence of silk fibroin

This paper describes a solution-phase approach to the synthesis of selenium nanoparticles by reducing selenious acid solution with ascorbic acid in the presence of silk fibroin. The monodispersed spherical selenium colloid particles obtained were very stable in silk fibroin solution and characterized by Atomic force microscopy and X-ray techniques. The influences of temperature and ultrasonication on the morphology of selenium nanoparticles were also discussed. The experiments showed that the selenium nanoparticles with various morphologies could be obtained under different temperatures and the appropriate ultrasonication time was 60 min. This result indicated that the silk fibroin molecules intimately associated with the surface of the selenium particles and controlled the growth particles.     

纳米TiO_2改性丝素膜制备工艺优化及其性能研究

设计正交试验法并采用溶胶一凝胶法制备了性能良好纳米TiO2溶胶丝素复合膜,并利用SEM、FTIR及XRD等分析手段对丝素膜进行表征,结果表明:纳米TiO2均匀分散在丝素膜中,纳米TiO2与丝素分子相互作用,导致丝素分子的重排,并诱导丝素构象从Silk Ⅰ向SilkⅡ转变.     

纳米CaCO3/柞蚕丝素复合膜的制备及结构表征

制备了一种含有纳米CaCO3的新型柞蚕丝素蛋白膜,通过SEM、DSC、TG、XRD和IR研究了纳米CaCO3的加入对这种丝素蛋白膜性能的影响。SEM测试表明,纳米CaCO3粒子能均匀分散在丝素膜中不团聚;DSC测试表明,适量加入纳米CaCO3复合膜的Tm有所提高,但随着纳米CaCO3含量增加,Tm又有所降低;TG测试结果表明,随着纳米CaCO3加入量增加,复合膜的热稳定性得到提高;XRD测试结果表明,丝素结晶结构从silk I向silk II转化,结晶度得到提高。IR测试结果佐证了前面的推断。     

EDC/NHS交联改性再生桑蚕丝素纳米纤维

用静电纺丝技术制备了再生桑蚕丝素纳米纤维,并用1-(3-二甲基氨基丙基)-3-乙基碳化二亚胺(EDC)和N-羟基丁二酰亚胺(NHS)进行交联改性,考察了交联改性前后,桑蚕丝素纳米纤维微观形貌及聚集态结构等的变化,采用扫描电镜(SEM)、X射线衍射(XRD)及红外光谱(FT-IR)等测试方法对纳米纤维进行表征。研究结果表明,经EDC/NHS交联改性后,纤维直径由250 nm增加到320 nm,纤维表面变粗糙,且纤维发生弯曲变形;丝素的结构以Silk II为主,并含有部分无规卷曲或Silk I构象;桑蚕丝纳米纤维的力学性能和亲水性有所提高,且交联改性后的纳米纤维具有良好的细胞相容性。     

Surface properties and cytocompatibillity of silk fibroin films cast from aqueous solutions in different concentrations

Silk fibroin film (SFF) has been widely used in biomaterials. SFF is usually prepared from a regenerated silk aqueous solution and its properties depend remarkably on the preparation conditions. However, the effect of the silk fibroin concentration (C₀) on the SFF surface properties as well as the cytocompatibility has rarely been investigated. In this work we prepared a series of Bombyx mori SFFs by casting SF aqueous solutions with the concentration from 10⁰ to 10²mg/mL on TCPS substrate at 60°C. The test results of atomic force microscopy, attenuated total reflection Fourier transform infrared and contact angles analysis showed that the film surface roughness and β-sheet structure increased with the increase of C₀, whereas the surface hydrophilicity increased with the decrease of C₀. The in vitro clotting time measurement results revealed that the SFFs prepared from the thinner solution showed a longer APTT (activated partial thromboplastin time) and TT (thrombin time). The results of microscopy and MTT assay also revealed that cell adhesion and growth were enhanced on the SFF cast from lower C₀ for fibroblasts. In contrast, endothelial cells showed a similar behavior on all those films that were prepared from the solution in different concentrations.     

Processing methods to control silk fibroin film biomaterial features

Control of silk structural and morphological features is reported for fibroin protein films via all aqueous processing, with and without polyethylene oxide (PEO). Silk films with thicknesses from 500 nm to 50 μm were generated with controllable surface morphologies by employing soft-lithography surface patterning or by adjusting PEO concentrations. FTIR analysis indicated that water-annealing, used to cure or set the films, resulted in increased β-sheet and α-helix content within the films. Steam sterilization provided an additional control point by increasing β-sheet content, while reducing random coil and turn structures, yet retaining film transparency and material integrity. Increased PEO concentration used during processing resulted in decreased sizes of surface globule structures, while simultaneously increasing uniformity of these features. The above results indicate that both surface and bulk morphologies and structures can be controlled by adjusting PEO concentration. The combined tool set for controlling silk film geometry and structure provides a foundation for further study of novel silk film biomaterial systems. These silk film biomaterials have potential applicability for a variety of optical and regenerative medicine applications due to their optical clarity, impressive mechanical properties, slow degradability, and biocompatibility.