ABSTRACT
This study focused on developing electrically stimulable hyaluronic acid (HA) films incorporating lipid nanoparticles (NPs) designed for the topical administration of lipophilic drugs and macromolecules. Based on beeswax and medium-chain triglycerides, NPs were successfully integrated into silk fibroin/chitosan films containing HA (NP-HA films) at a density of approximately 1011 NP/cm2, ensuring a uniform distribution. This integration resulted in a 40% increase in film roughness, a twofold decrease in Young's modulus, and enhanced film flexibility and bioadhesion work. The NP-HA films, featuring Ag/AgCl electrodes, demonstrated the capability to conduct a constant electrical current of 0.2 mA/cm2 without inducing toxicity in keratinocytes and fibroblasts during a 15-min application. Moreover, the NPs facilitated the homogeneous distribution of lipophilic drugs within the film, effectively transporting them to the skin and uniformly distributing them in the stratum corneum upon film administration. The sustained release of HA from the films, following Higuchi kinetics, did not alter the macroscopic characteristics of the film. Although anodic iontophoresis did not noticeably affect the release of HA, it did enhance its penetration into the skin. This enhancement facilitated the permeation of HA with a molecular weight (MW) of up to 2 × 105 through intercellular and transcellular routes. Confocal Raman spectroscopy provided evidence of an approximate 100% increase in the presence of HA with a MW in the range of 1.5-1.8 × 106 in the viable epidermis of human skin after only 15 min of iontophoresis applied to the films. Combining iontophoresis with NP-HA films exhibits substantial potential for noninvasive treatments focused on skin rejuvenation and wound healing.
Subject(s)
Hyaluronic Acid , Nanoparticles , Hyaluronic Acid/chemistry , Hyaluronic Acid/administration & dosage , Nanoparticles/administration & dosage , Nanoparticles/chemistry , Humans , Skin Absorption , Animals , Skin/metabolism , Chitosan/chemistry , Chitosan/administration & dosage , Administration, Cutaneous , Drug Delivery Systems , Lipids/chemistry , Lipids/administration & dosage , Fibroins/chemistry , Fibroins/administration & dosage , Keratinocytes/drug effects , Polymers/chemistry , Polymers/administration & dosage , LiposomesABSTRACT
Silk fibroin is a fibrillar protein obtained from arthropods such as mulberry and non-mulberry silkworms. Silk fibroin has been used as a dressing in wound treatment for its physical, chemical, mechanical, and biological properties. This systematic review analyzed studies from PubMed, Web of Science, and Scopus databases to identify the molecules preferred for functionalizing silk fibroin-based dressings and to describe their mechanisms of exhibiting anti-inflammatory and antibacterial properties. The analysis of the selected articles allowed us to classify the dressings into different conformations, such as membranes, films, hydrogels, sponges, and bioadhesives. The incorporation of various molecules, including antibiotics, natural products, peptides, nanocomposites, nanoparticles, secondary metabolites, growth factors, and cytokines, has allowed the development of dressings that promote wound healing with antibacterial and immunomodulatory properties. In addition, silk fibroin-based dressings have been established to have the potential to regenerate wounds such as venous ulcers, arterial ulcers, diabetic foot, third-degree burns, and neoplastic ulcers. Evaluation of the efficacy of silk fibroin-based dressings in tissue engineering is an area of great activity that has shown significant advances in recent years.
Subject(s)
Anti-Bacterial Agents , Anti-Inflammatory Agents , Bandages , Fibroins , Wound Healing , Fibroins/chemistry , Anti-Bacterial Agents/chemistry , Anti-Bacterial Agents/pharmacology , Humans , Anti-Inflammatory Agents/chemistry , Anti-Inflammatory Agents/pharmacology , Animals , Wound Healing/drug effectsABSTRACT
The aim of this work was to develop a dense lamellar scaffold, as a biomimetic material with potential applications in the regeneration of tracheal tissue after surgical tumor resection. The scaffolds were produced by plastic compression technique, exploiting the use of total phenolic compounds (TPC) from Psidium guajava Linn as a potential cross-linking agent in a polymeric mixture based on collagen (COL), silk fibroin (SF), and polyethylene glycol 400 (PEG 400). Fourier transform infrared spectroscopy (FTIR) and differential scanning calorimetry (DSC) confirmed the chemical interactions between the polymers and the cross-linking of TPC between COL and SF. Morphological analyses showed scaffolds with porosity, interconnectivity, and a porous surface structure with a gyroid-like geometry. The analysis of the anisotropic degree resulted in anisotropic structures (0.1% TFC and 0.3% TFC) and an isotropic structure (0.5% TFC). In the mechanical properties, it was evidenced greater resistance for the 0.3% TFC formulation. The addition of TPC percentages did not result in a significant difference (p > 0.05) in swelling capacity and disintegration rate. The results confirmed that TPC were able to modulate the morphological, morphometric, and mechanical properties of scaffolds. Thus, this study describes a potential new material to improve the regeneration of major tracheal structures after surgical tumor removal.
Subject(s)
Fibroins , Neoplasms , Psidium , Tissue Engineering/methods , Tissue Scaffolds/chemistry , Fibroins/chemistry , Collagen/chemistry , Porosity , Spectroscopy, Fourier Transform InfraredABSTRACT
Electrospinning is a versatile technique for fabricating polymeric fibers with diameters ranging from micro- to nanoscale, exhibiting multiple morphologies and arrangements. By combining silk fibroin (SF) with synthetic and/or natural polymers, electrospun materials with outstanding biological, chemical, electrical, physical, mechanical, and optical properties can be achieved, fulfilling the evolving biomedical demands. This review highlights the remarkable versatility of SF-derived electrospun materials, specifically focusing on their application in tissue regeneration (including cartilage, cornea, nerves, blood vessels, bones, and skin), disease treatment (such as cancer and diabetes), and the development of controlled drug delivery systems. Additionally, we explore the potential future trends in utilizing these nanofibrous materials for creating intelligent biomaterials, incorporating biosensors and wearable sensors for monitoring human health, and also discuss the bottlenecks for its widespread use. This comprehensive overview illuminates the significant impact and exciting prospects of SF-derived electrospun materials in advancing biomedical research and applications.
Subject(s)
Fibroins , Nanofibers , Humans , Fibroins/chemistry , Tissue Engineering/methods , Biocompatible Materials/chemistry , Drug Delivery Systems , Nanofibers/chemistry , Polymers , Silk/chemistry , Tissue Scaffolds/chemistryABSTRACT
There are many challenges in the development of 3D-tissue models for studying bone physiology and disease. Silk fibroin (SF), a natural fibrous protein used in biomedical applications has been studied for bone tissue engineering (TE) due to its mechanical properties, biocompatibility and biodegradability. However, low osteogenic capacity as well as the necessity to reinforce the protein mechanically for some orthopedic applications prompts the need for further designs for SF-based materials for TE bone. Concentric mineralized porous SF-based scaffolds were developed to improve mechanics and mineralization towards osteoregeneration. Hybrid SF silica microparticles (MP) or calcium carbonate nano-structured microparticles (NMP) were seeded with hMSCs co-cultured under osteogenic and osteoclastic conditions with THP-1 human monocytes up to 10 weeks to simulate and recapitulate bone regeneration. Scaffolds with appropriate pore size for cell infiltration, resulted in improved compressive strength, increased cell attachment and higher levels of expression of osteogenic markers and mineralization after adding the NMPs, compared to controls systems without these particles. These hybrid SF-based 3D-structures can provide improved scaffold designs for in vitro bone TE.
Subject(s)
Fibroins , Tissue Engineering , Humans , Tissue Engineering/methods , Silk/chemistry , Tissue Scaffolds/chemistry , Bone and Bones , Osteogenesis , Fibroins/chemistryABSTRACT
Bone regeneration and natural repair are long-standing processes that can lead to uneven new tissue growth. By introducing scaffolds that can be autografts and/or allografts, tissue engineering provides new approaches to manage the major burdens involved in this process. Polymeric scaffolds allow the incorporation of bioactive agents that improve their biological and mechanical performance, making them suitable materials for bone regeneration solutions. The present work aimed to create chitosan/beta-tricalcium phosphate-based scaffolds coated with silk fibroin and evaluate their potential for bone tissue engineering. Results showed that the obtained scaffolds have porosities up to 86%, interconnectivity up to 96%, pore sizes in the range of 60-170 µm, and a stiffness ranging from 1 to 2 MPa. Furthermore, when cultured with MC3T3 cells, the scaffolds were able to form apatite crystals after 21 d; and they were able to support cell growth and proliferation up to 14 d of culture. Besides, cellular proliferation was higher on the scaffolds coated with silk. These outcomes further demonstrate that the developed structures are suitable candidates to enhance bone tissue engineering.
Subject(s)
Chitosan , Fibroins , Calcium Phosphates , Cell Proliferation , Fibroins/chemistry , Porosity , Silk/chemistry , Tissue Engineering/methods , Tissue Scaffolds/chemistryABSTRACT
This review addresses composites prepared from cellulose (Cel) and silk fibroin (SF) to generate multifunctional, biocompatible, biodegradable materials such as fibers, films and scaffolds for tissue engineering. First, we discuss briefly the molecular structures of Cel and SF. Their structural features explain why certain solvents, e.g., ionic liquids, inorganic electrolyte solutions dissolve both biopolymers. We discuss the mechanisms of Cel dissolution because in many cases they also apply to (much less studied) SF dissolution. Subsequently, we discuss the fabrication and characterization of Cel/SF composite biomaterials. We show how the composition of these materials beneficially affects their mechanical properties, compared to those of the precursor biopolymers. We also show that Cel/SF materials are excellent and versatile candidates for biomedical applications because of the inherent biocompatibility of their components.
Subject(s)
Biocompatible Materials/chemistry , Cellulose/chemistry , Fibroins/chemistry , Skin , Tissue Engineering , Animals , Biopolymers/chemistry , Bombyx , Chemical Phenomena , Humans , Ionic Liquids , Structure-Activity Relationship , Tissue Scaffolds/chemistryABSTRACT
Fabrication of functional silk fibroin microstructures has extensive applications in biotechnology and photonics. Considerable progress has been made based on lithographic methods and self-assembly approaches. However, most methods require chemical modification of silk fibroin, which restricts the functionalities of the designed materials. At the same time, femtosecond laser-induced forward transfer (fs-LIFT) has been explored as a simple and attractive processing tool for microprinting of high-resolution structures. In this paper, we propose the use of LIFT with fs-pulses for creating high-resolution structures of regenerated silk fibroin (SF). Furthermore, upon adding Eu3+/Tb3+ complexes to SF, we have been able to demonstrate the printing by LIFT of luminescent SF structures with a resolution on the order of 2 µm and without material degradation. This approach provides a facile method for printing well-defined two-dimensional (2D) micropatterns of pure and functionalized SF, which can be used in a wide range of optical and biomedical applications.
Subject(s)
Fibroins/chemistry , Lasers , Organometallic Compounds/chemistry , Printing, Three-Dimensional , Fibroins/isolation & purification , Organometallic Compounds/chemical synthesis , Particle Size , Surface Properties , Time FactorsABSTRACT
In this study, we proposed the use of the biopolymers silk fibroin, chitosan and alginate, which are recognized for their biocompatibility and biodegradability, for the preparation of multilayer membranes aiming at high performance wound dressings with controlled drug delivery. The rationale was to combine in one material the mechanical properties of fibroin, the antimicrobial action of chitosan and the ideal exudate absorption of alginate, reaching a synergic effect of each biopolymer, without losing their individual intrinsic properties. The membranes were prepared by casting and diclofenac sodium was incorporated as model drug into the chitosan solution before the solvent evaporation, being retained in the middle layer of the membrane. Morphological, thermal, mechanical, solubility and barrier properties of the membranes were evaluated, as well as cytotoxicity and microbiological permeation. Results show that the incorporation of the drug did not affect mechanical and barrier properties, as well as microbiological permeation. Drug release was evaluated in vitro using simulated solution of wound exudate at 37 °C and diclofenac sodium was released from the multilayer membrane in 7 h, in which Fickian diffusion was the main mechanism associated. The results show the potential application of the biopolymer multilayer membranes as high-performance wound dressings.
Subject(s)
Alginates/chemistry , Chitosan/chemistry , Delayed-Action Preparations/chemistry , Fibroins/chemistry , Membranes/chemistry , Silk/chemistry , Bandages , Biocompatible Materials/chemistry , Biopolymers/chemistry , Delayed-Action Preparations/pharmacology , Drug Liberation/drug effects , Wound Healing/drug effectsABSTRACT
Hydrogel scaffolds are important materials in tissue engineering, and their characterization is essential to determine potential biomedical applications according to their mechanical and structural behavior. In this work, silk fibroin hydrogels were synthesized by two different methods (vortex and sonication), and agarose hydrogels were also obtained for comparison purposes. Samples were characterized by scanning electron microscopy, infrared analysis, thermo-gravimetrical analysis, confined compression test, and rheological test. The results indicate that nanofibers can be obtained via both silk fibroin and agarose hydrogels. The mechanical tests showed that the Young's modulus is similar to those found in the literature, with the highest value for agarose hydrogels. All the hydrogels showed a shear-thinning behavior. Additionally, the MTT test revealed that silk fibroin hydrogels had low cytotoxicity in THP-1 and HEK-293 cells, whereas the agarose hydrogels showed high toxicity for the THP-1 cell line. The results indicate that silk fibroin hydrogels obtained from a Colombian silkworm hybrid are suitable for the development of scaffolds, with potential applications in tissue engineering.
Subject(s)
Cell Proliferation , Fibroins/chemistry , Hydrogels/chemistry , Sepharose/chemistry , Tissue Engineering , Animals , Biocompatible Materials , Bombyx , Colombia , HEK293 Cells , Humans , Mice , THP-1 Cells , Tissue ScaffoldsABSTRACT
Cardiovascular disease is a worldwide main cause of morbidity and mortality. Treatment alternatives include the use of cardiovascular implants that have generated a constant search for materials, and transformation processes that provide structures similar to those that need to be replaced. Among the biomaterials available for vascular implants, silk fibroin (SF) is of great interest because it is a natural, biodegradable, biocompatible protein. In addition, SF has outstanding mechanical properties and can be easily processed by various techniques. This article presents a general review of SF, its potential use as a biomaterial for vascular applications, and modifications that improve its hemocompatibility.
Subject(s)
Biocompatible Materials/chemistry , Blood Vessel Prosthesis , Fibroins/chemistry , Animals , Bombyx , Humans , Materials Testing , Prosthesis DesignABSTRACT
In this study, we prepared translucid hydrogels with different concentrations of silk fibroin, extracted from raw silk fibers, and used them as a matrix to incorporate the photosensitizer 5-(4-aminophenyl)-10,15,20-tris-(4-sulphonatophenyl) porphyrin trisodium for application in photodynamic therapy (PDT). The hydrogels obtained were characterized by rheology, spectrophotometry, and scattering techniques to elucidate the factors involved in the formation of the hydrogel, and to characterize the behavior of silk fibroin (SF) after incorporating of the porphyrin to the matrix. The rheology results demonstrated that the SF hydrogels had a shear thinning behavior. In addition, we were able to verify that the structure of the material was able to be recovered over time after shear deformation. The encapsulation of porphyrins in hydrogels leads to the formation of self-assembled peptide nanostructures that prevent porphyrin aggregation, thereby greatly increasing the generation of singlet oxygen. Also, our findings suggest that porphyrin can diffuse out of the hydrogel and permeate the outer skin layers. This evidence suggests that SF hydrogels could be used as porphyrin encapsulation and as a drug carrier for the sustained release of photosensitizers for PDT.
Subject(s)
Fibroins/chemistry , Hydrogels/chemistry , Circular Dichroism , Drug Carriers/chemistry , Light , Photochemotherapy , Photosensitizing Agents/chemistry , Photosensitizing Agents/metabolism , Porphyrins/chemistry , Porphyrins/metabolism , Rheology , Singlet Oxygen/metabolismABSTRACT
The capture spiral of web from N. clavipes spider consists of a single type of spidroin - the flagelliform silk protein, a natural material representing a combination of strength and high elasticity. Flagelliform spider silk is the most extensible silk fibre produced by orb weaver spiders and the structure of this remarkable material is still largely unknown. In the present study we used a proteomic approach to elucidate the complete sequence and the post-translational modifications of flagelliform silk proteins. The long sequence of flagelliform silk protein presents 45 hydroxylated proline residues, which may contribute to explain the mechanoelastic property of these fibres, since they are located in the GPGGX motif. The 3D-structure of the protein was modelled considering the three domains together, i.e., the N- and C-terminal non-repetitive domains, and the central repetitive domain. In the resulting molecular model there is a predominance of random structures in the solid fibres of the silk protein. The N-terminal domain is composed of three α-helices and the C-terminal domain is composed of one small helical section. Proteomic data reported herein may be relevant for the development of novel approaches for the synthetic or recombinant production of novel silk-based spider polymers.
Subject(s)
Fibroins/chemistry , Silk/chemistry , Spiders/chemistry , Animals , Biomechanical Phenomena , Fibroins/metabolism , Fibroins/ultrastructure , Microscopy, Electron, Scanning , Models, Molecular , Protein Conformation , Protein Processing, Post-TranslationalABSTRACT
Silk fibroin (SF) films containing a peptide, neurotensin (NT), stimulated by iontophoresis were developed aiming to modulate the inflammatory process and prevent the growth of microorganisms typical of wounds. NT-loaded SF films composition shows predominance of ß-sheet structures that conferred adequate mechanical properties, transparency, moderate roughness and low swelling index to fibroin films. Infrared spectroscopy and thermal analysis suggested the presence of non-covalent interactions between NT and fibroin. Using the MALDI imaging technique, it was possible to visualize the homogeneous NT distribution throughout the film surface, in addition to its prolonged release for up to 72â¯h. In vitro studies in E. coli liposaccharide-stimulated macrophages showed a significant reduction of interleukins production after NT-loaded film application, whereas NT solution did not reduce them. Bi-laminated NT-loaded fibroin films containing silver electrodes provided a burst release of NT when anodic iontophoresis was applied, enabling a rapid onset of drug action. In addition, anodic iontophoresis presented a bacteriostatic effect against gram-positive microorganisms. Different iontophoresis densities, from 0.2 to 0.6â¯mA/cm2, did not significantly reduce fibroblast viability after 30â¯min of application. In conclusion, iontophoretic-stimulated peptide-loaded fibroin films could be a promising platform for the treatment of wounds.
Subject(s)
Anti-Bacterial Agents/administration & dosage , Drug Delivery Systems/methods , Fibroins/chemistry , Neurotensin/administration & dosage , Wound Healing/drug effects , Animals , Anti-Bacterial Agents/chemistry , Bacteria/drug effects , Bacteria/metabolism , Bombyx/chemistry , Cell Survival , Fibroblasts , Interleukins/metabolism , Iontophoresis/methods , Lipopolysaccharides/pharmacology , Macrophages/drug effects , Macrophages/metabolism , Microbial Sensitivity Tests , Neurotensin/chemistry , Spectrometry, Mass, Matrix-Assisted Laser Desorption-IonizationABSTRACT
In the present study, a new electrospun silk fibroin coating of silicone breast implants with improved biocompatibility and mechanical properties was obtained. Fibrous scaffolds were produced by electrospinning a solution containing silk fibroin, derived from Bombyx mori cocoons, and polyethylene oxide (PEO) to be used as a coating of breast implants. A randomly oriented structure of fibroin/PEO was electrospun on implants as assessed by SEM analysis, roughness measurements and ATR-FTIR spectroscopy. The scaffold showed 0.25 µm diameter fibres, 0.76 µm size superficial pores, arithmetic roughness of 0.632 ± 0.12 µm and texture aspect ratio of 0.893 ± 0.04. ATR-FTIR spectroscopy demonstrates the presence of PEO and fibroin in the coating. The mechanical characterisation of the implants before and after being coated with fibroin/PEO demonstrated that the fibroin/PEO scaffold contributes to the increase in the elastic modulus from 0.392 ± 0.02 to 0.560 ± 0.03 MPa and to a more elastic behaviour of the breast implants. Using the fibroin/PEO coating, human fibroblasts seeded on this matrix increased viability up to 30% compared to conventional breast implants. Electrospun silk fibroin could represent a clinically compatible, viable form to coat breast implants. Low cytotoxicity by the fibroin coating and its physico-chemical and mechanical properties may find application in improving breast implants biocompatibility. © 2017 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 106B: 1655-1661, 2018.
Subject(s)
Breast Implants , Coated Materials, Biocompatible/chemistry , Fibroblasts/metabolism , Fibroins/chemistry , Materials Testing , Polyethylene Glycols/chemistry , Female , Fibroblasts/cytology , Humans , Surface PropertiesABSTRACT
BACKGROUND: Current treatment of inflammatory bowel disease is based on the use of immunosuppressants or anti-inflammatory drugs, which are characterized by important side effects that can limit their use. Previous research has been performed by administering these drugs as nanoparticles that target the ulcerated intestinal regions and increase their bioavailability. It has been reported that silk fibroin can act as a drug carrier and shows anti-inflammatory properties. PURPOSE: This study was designed to enhance the interaction of the silk fibroin nanoparticles (SFNs) with the injured intestinal tissue by functionalizing them with the peptide motif RGD (arginine-glycine-aspartic acid) and to evaluate the intestinal anti-inflammatory properties of these RGD-functionalized silk fibroin nanoparticles (RGD-SFNs) in the trinitrobenzenesulfonic acid (TNBS) model of rat colitis. MATERIALS AND METHODS: SFNs were prepared by nanoprecipitation in methanol, and the linear RGD peptide was linked to SFNs using glutaraldehyde as the crosslinker. The SFNs (1 mg/rat) and RGD-SFNs (1 mg/rat) were administered intrarectally to TNBS-induced colitic rats for 7 days. RESULTS: The SFN treatments ameliorated the colonic damage, reduced neutrophil infiltration, and improved the compromised oxidative status of the colon. However, only the rats treated with RGD-SFNs showed a significant reduction in the expression of different pro-inflammatory cytokines (interleukin [IL]-1ß, IL-6, and IL-12) and inducible nitric oxide synthase in comparison with the TNBS control group. Moreover, the expression of both cytokine-induced neutrophil chemoattractant-1 and monocyte chemotactic protein-1 was significantly diminished by the RGD-SFN treatment. However, both treatments improved the intestinal wall integrity by increasing the gene expression of some of its markers (trefoil factor-3 and mucins). CONCLUSION: SFNs displayed intestinal anti-inflammatory properties in the TNBS model of colitis in rats, which were improved by functionalization with the RGD peptide.
Subject(s)
Colitis/drug therapy , Fibroins/chemistry , Fibroins/pharmacology , Intestines/drug effects , Nanoparticles/chemistry , Oligopeptides/chemistry , Trinitrobenzenesulfonic Acid/pharmacology , Animals , Anti-Inflammatory Agents/chemistry , Anti-Inflammatory Agents/pharmacology , Anti-Inflammatory Agents/therapeutic use , Colitis/chemically induced , Colitis/immunology , Colitis/metabolism , Cytokines/metabolism , Disease Models, Animal , Female , Fibroins/therapeutic use , Intestinal Mucosa/metabolism , Intestines/immunology , Nanomedicine , Neutrophil Infiltration/drug effects , Nitric Oxide Synthase Type II/metabolism , Rats , Rats, WistarABSTRACT
Layer-by-Layer (LbL) films containing silk fibroin (SF) and the 40 aminoacid-long amyloid-ß peptide (Aß1-40) were prepared with the purpose of developing a new prototype of an electrochemical immunosensor. The film showed a satisfactory growth in quartz substrate and screen-printed carbon electrodes, as observed by UV-vis spectroscopy and cyclic voltammetric, respectively. The peptide immobilized in LbL films in junction with SF shows secondary structure induced, as shown by circular dichroism measurements, favoring the interaction SF/peptide LbL film with the specific antibody. Immunosensor showed a linear response in the presence of the antibody with concentrations from 0 to 10ngmL(-1) both analyzed by current changes in 0.3V and voltammogram area. This system can be applied as a new prototype for preliminary diagnosis of Alzheimer's disease.
Subject(s)
Alzheimer Disease , Amyloid beta-Peptides/metabolism , Antibodies/chemistry , Biosensing Techniques/methods , Fibroins/chemistry , Membranes, Artificial , Peptide Fragments/metabolism , Alzheimer Disease/diagnosis , Alzheimer Disease/metabolism , Animals , HumansABSTRACT
Most reports about the 3-D structure of spidroin-1 have been proposed for the protein in solid state or for individual domains of these proteins. A gel-based mass spectrometry strategy using collision-induced dissociation (CID) and electron-transfer dissociation (ETD) fragmentation methods was used to completely sequence spidroins-1A and -1B and to assign a series of post-translational modifications (PTMs) on to the spidroin sequences. A total of 15 and 16 phosphorylation sites were detected on spidroin-1A and -1B, respectively. In this work, we present the nearly complete amino acid sequence of spidroin-1A and -1B, including the nonrepetitive N- and C-terminal domains and a highly repetitive central core. We also described a fatty acid layer surrounding the protein fibers and PTMs in the sequences of spidroin-1A and -1B, including phosphorylation. Thus, molecular models for phosphorylated spidroins were proposed in the presence of a mixture fatty acids/water (1:1) and submitted to molecular dynamics simulation. The resulting models presented high content of coils, a higher percentage of α-helix, and an almost neglected content of 310-helix than the previous models. Knowledge of the complete structure of spidroins-1A and -1B would help to explain the mechanical features of silk fibers. The results of the current investigation provide a foundation for biophysical studies of the mechanoelastic properties of web-silk proteins.
Subject(s)
Fibroins/chemistry , Models, Molecular , Silk/chemistry , Spiders/chemistry , Amino Acid Sequence , Animals , Microscopy, Electron, Scanning , Molecular Dynamics Simulation , Molecular Sequence Data , Sequence Homology, Amino AcidABSTRACT
A previous experiment demonstrated that fibroin protein and chitosan mixed in proper proportion presented good physical and chemical properties and biological characteristics, which can make up for their respective disadvantages. To observe the growth of bone marrow mesenchymal stem cells (BMSCs) on these fibroin protein/chitosan 3D scaffolds, induced rabbit BMSCs were seeded on fibroin protein/chitosan scaffolds. The cell adhesion rate was measured, and cell growth was observed under an inverted microscope and a scanning electron microscope. The cell adhesion rate increased with time. The inverted microscope observations showed that the cells on fibroin protein/chitosan scaffolds could not be seen clearly. As time passed, the number of cells around the stent increased and some cells stretched inside the scaffolds. Electron microscopy showed active cell growth and normal proliferation, and the granular and filamentous matrix substances could be seen around cells. The microfilaments of cell and scaffold materials were tightly connected. The cells not only grew on the surface of the adherent material, but also stretched inside of the materials. These results indicated that the fibroin protein/ chitosan mixed scaffolds have good biocompatibility.
Subject(s)
Cell Culture Techniques , Chitosan/chemistry , Fibroins/chemistry , Mesenchymal Stem Cells/cytology , Tissue Scaffolds/chemistry , Animals , Cell Adhesion , Female , Male , Mesenchymal Stem Cells/metabolism , Mesenchymal Stem Cells/ultrastructure , RabbitsABSTRACT
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 25/75 wt.% 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.