Gentamicin-Loaded Wound Dressing With Polyvinyl Alcohol/Dextran Hydrogel: Gel Characterization and In Vivo Healing Evaluation

To develop a gentamicin-loaded wound dressing, cross-linked hydrogel films were prepared with polyvinyl alcohol (PVA) and dextran using the freezing–thawing method. Their gel properties such as gel fraction, swelling, water vapor transmission test, morphology, tensile strength, and thermal property were investigated. In vitro protein adsorption test, in vivo wound healing test, and histopathology were performed. Dextran decreased the gel fraction, maximum strength, and thermal stability of hydrogels. However, it increased the swelling ability, water vapor transmission rate, elasticity, porosity, and protein adsorption. The drug gave a little positive effect on the gel properties of hydrogels. The gentamicin-loaded wound dressing composed of 2.5% PVA, 1.13% dextran, and 0.1% drug was more swellable, flexible, and elastic than that with only PVA because of its cross-linking interaction with PVA. In particular, it could provide an adequate level of moisture and build up the exudates on the wound area. From the in vivo wound healing and histological results, this gentamicin-loaded wound dressing enhanced the healing effect more compared to conventional product because of the potential healing effect of gentamicin. Thus, this gentamicin-loaded wound dressing would be used as a potential wound dressing with excellent forming and improved healing effect in wound care.     

Electrospun zwitterionic poly(sulfobetaine methacrylate) for nonadherent, superabsorbent, and antimicrobial wound dressing applications

Zwitterionic poly(sulfobetaine methacrylate) (PSBMA) has been well studied for its superhydrophilic and ultralow biofouling properties, making it a promising material for superabsorbent and nonadherent wound dressings. Electrospinning provides multiple desirable features for wound dressings, including high absorptivity due to high surface-area-to-volume ratio, high gas permeation, and conformability to contour of the wound bed. The goal of this work is to develop a fibrous membrane of PSBMA via electrospinning and evaluate its properties related to wound dressing applications. Being superhydrophilic, PSBMA fibers fabricated by a conventional electrospinning method would readily dissolve in water, whereas if cross-linker is added, the formation of hydrogel would prevent electrospinning. A three-step polymerization-electrospinning-photo-cross-linking process was developed in this work to fabricate the cross-linked electrospun PSBMA fibrous membrane. Such electrospun membrane was stable in water and exhibited high water absorption of 353% (w/w), whereas the PSBMA hydrogel only absorbed 81% water. The electrospun membrane showed strong resistance to protein adsorption and cell attachment. Bacterial adhesion studies using Gram negative P. aeruginosa and Gram positive S. epidermidis showed that the PSBMA electrospun membrane was also highly resistant to bacterial adhesion. The Ag(+)-impregnated electrospun PSBMA membrane was shown microbicidal, against both S. epidermidis and P. aeruginosa. Such electrospun PSBMA membrane is ideal for a novel type of nonadherent, superabsorbent, and antimicrobial wound dressing. The superior water absorption aids in fluid removal from highly exudating wounds while keeping the wound hydrated to support healing. Because of the resistance to protein, cell, and bacterial adhesion, the dressing removal will neither cause patients' pain nor disturb the newly formed tissues. The dressing also prevents the attachment of environmental bacteria and offers broad-spectrum antimicrobial activity. It is the first work to develop the water-stable electrospun PSBMA membrane, which has great potential for wound dressing and other applications.     

The Effects of Sericin Cream on Wound Healing in Rats

Sericin has good hydrophilic properties, compatibility, and biodegradation, it can be used as a wound-healing agent. We evaluated the effects of sericin on wound healing and wound size reduction using rats by generating two full-thickness skin wounds on the dorsum. Group 1 animals were treated with Betadine^® on left-side (control) wounds and, with 8% sericin cream on right-side (treated) wounds. Group 2, cream base (formula control) and 8% sericin cream (treated) were topically applied to left-, and right-side wounds respectively. Sericin-treated wounds had much smaller inflammatory reactions, and wound-size reduction was much greater than in the control throughout the inspection period. Mean time in days for 90% healing from sericin-treated wounds was also much less than for cream base-treated wounds. Histological examination after 15 d of treatment with 8% sericin cream revealed complete healing, no ulceration, and an increase in collagen as compared to cream base-treated wounds, which showed some ulceration and acute inflammatory exudative materials.     

Nanocomposite regeneration systems for wound healing

The regeneration system “Biokol” based on the principles of combining synthetic and natural biopolymers has been investigated. The wound dressing consists of “large” (200–250 nm) particles of a synthetic biopolymer and a gel component, which consists of “small” polysaccharide elements 10–20 nm in size. The system can be used both separately and in combination with the gel complex and cell cultures. In properties (vapor and gas permeability, mechanical properties, conductivity, resistance to microbes, etc.), it corresponds to the upper layer of the skin. When applied to the wound, the dressing changes its adhesiveness: first, owing to its hydrophilicity and low hydrophobicity, it closely adheres to the wound surface, and after some time, which corresponds to the time the polysaccharide complex is released from the dressing, it becomes hydrophobic and easily separates from the wound. Owing to these properties, the system can be used at all stages of wound healing.     

Potential of 2D crosslinked sericin membranes with improved biostability for skin tissue engineering

Silk sericin protein is a natural, hydrophilic, macromolecular glycoprotein mainly synthesized in the middle silk gland of the silkworm. It constitutes 25–30% of the silk cocoon. Sericin proteins have antioxidant, antimicrobial, UV-resistant properties, promote wound healing and support cell proliferation even in serum-free media. Most of the sericin is discarded as waste in silk processing industries. This study aims at improving the mechanical strength and stability of sericin extracted from the silk cocoons during processing and utilize it as a biocompatible natural biopolymer in biomedical applications. Crosslinked sericin membranes, from the cocoon of non-mulberry tropical silkworm, Antheraea mylitta, were prepared using gluteraldehyde as the crosslinking agent. Physical and structural characteristics of the membranes were analyzed using scanning electron microscopy, atomic force microscopy, Fourier transform infrared spectroscopy and X-ray diffraction along with swelling and degradation studies. The secondary structure of the membrane indicates that crosslinking provides a more integrated structure that significantly improves the stability and mechanical strength of the membranes. In vitro cytocompatibility of the membranes was evaluated by MTT assay and cell cycle analysis of feline fibroblast cells. The adherence, growth and proliferation patterns of cells on membranes were assessed by confocal microscopy, which demonstrated that the latter is non-toxic and supports cell growth. Cell cycle analyses indicate cytocompatibility with normal cell cycle pattern. This study reveals that silk sericin protein can be used as a biocompatible natural biopolymer for various applications in the biomedical field.     

Immobilization of derivatized dextran nanoparticles on konjac glucomannan/chitosan film as a novel wound dressing

The aim of this study was to prepare konjac glucomannan (KGM)/chitosan (CS) film containing glycidyl methacrylate derivatized dextran (dex-GMA)/acrylic acid(AAc) nanoparticles loaded with antibacterial agent. In this study, An optimized procedure chosen from three methods was used to prepare Erythromycin (EM)-loaded poly(dex-GMA/AAc) nanoparticles and obtained nanoparticles ranged from 50-200 nm. Film was found to have equilibrium water content (EWC) 99.3% which could prevent exudates on wound bed from accumulating and also have excellent water adsorption 2362.3 +/- 55.2%; the water vapor transmission rate (WVTR) was 2335 +/- 36 gm(-2) day(-1) and evaporative water loss from the film (EWL) was approximately 10% after 1 h and within 6 h it increased to 90%. Drug release of film containing nanoparticles or absent was determined, within 22 h accumulative release was 40.3%, 72.5% respectively. In conclusion, KGM/CS film containing nanoparticles could not only maintain a moist environment over wound bed in moderate to heavily exuding wound but also provide a continuous and sustained release of the antibacterial agent on the wound surface, which could be potential wound dressing.     

Raman spectroscopy and WAXS method as a tool for analysing ion-exchange properties of alginate hydrogels

Hydrogels are cross-linked three-dimensional macromolecular networks that contain a large fraction of water within their structure. One of the most important properties of alginate hydrogels, leading to their broad versatility, is their ability for controlled uptake, release and retention of molecules. This ability, in turn, is due to specific interactions of the macromolecular network with the diffusing or retained molecule. Raman spectroscopy has been employed to characterize the diffusion properties of solutes in hydrogels. Besides their application in the food sector, they are used in many biomedical, pharmaceutical and technical areas; for example, as a natural tissue or drug carriers. In the latter case, controlled release of drugs from a wound dressing is of particular interest-or ion exchange between the drug and the structure of the dressing. Raman active vibrations were used to show the areas responsible for the penetration of the model azo-dyes (based on non-genotoxic benzidine analogs) within Ca-alginate/carboxymethylcellulose Medisorb A wound dressing. In this case, the intensity of the stretching bands was used to obtain the concentration profiles of the model dye in alginate/carboxymethylcellulose gel (Medisorb A). The characteristic band at 1511 cm(-1) indicates that new band positions were observed following dye adsorption on wound dressing. The Raman spectra of alginate immersed for different times in Ringer's solution reveal peak shifts. Differences in peak shapes and the appearance of new bands are observed as the sodium content increased. Raman spectra give direct information on the exchange process. There are also new peaks appearing at 1034-1016 and 850 cm(-1) regions in the spectra after the release studies. This could, therefore, correspond to a partial bonding between sodium and oxygen atoms (the guluronic units originate a band at approximately 1025 cm(-1)). The aim of the examination in this paper also was to investigate the crystallinity index of Medisorb A wound dressing dyed (or undyed) and Medisorb A wound dressing after the release process in Ringer's solution (the crystallinity index is about 65%). In WAXS curves we can observed additional peaks (2theta at 32 degrees and 45 degrees ).     

The effects of sericin cream on wound healing in rats

Sericin has good hydrophilic properties, compatibility, and biodegradation, it can be used as a wound-healing agent. We evaluated the effects of sericin on wound healing and wound size reduction using rats by generating two full-thickness skin wounds on the dorsum. Group 1 animals were treated with Betadine on left-side (control) wounds and, with 8% sericin cream on right-side (treated) wounds. Group 2, cream base (formula control) and 8% sericin cream (treated) were topically applied to left-, and right-side wounds respectively. Sericin-treated wounds had much smaller inflammatory reactions, and wound-size reduction was much greater than in the control throughout the inspection period. Mean time in days for 90% healing from sericin-treated wounds was also much less than for cream base-treated wounds. Histological examination after 15 d of treatment with 8% sericin cream revealed complete healing, no ulceration, and an increase in collagen as compared to cream base-treated wounds, which showed some ulceration and acute inflammatory exudative materials.     

Characterization of sericin powder prepared from citric acid-degraded sericin polypeptides of the silkworm, Bombyx Mori

Acid-degraded sericin powder (AC-SP) was prepared from aqueous solution containing citric acid-degraded sericin polypeptides of Bombyx mori. The morphological and biochemical properties of AC-SP were compared with those of alkali-degraded sericin powder (AL-SP) and hot-water degraded sericin powder (HW-SP). Based on an SEM analysis, AC-SP showed a thin film structure of 10-100 microm with good dispersity while AL-SP and HW-SP had a much larger thin film structure (<500 microm). The extract of AC-SP showed stronger trypsin inhibitor activity due to cocoon shell trypsin inhibitor (CSTI-IV) than that of HW-SP. The extract of AL-SP showed no CSTI-IV activity. It was found that AC-SP was a trypsin inhibitor complex powder and that the release of CSTI-IV from AC-SP depended on pH and ion strength. Similar powder materials were obtained when such organic acids as tartaric acid and succinic acid were used. These results suggest that the acid-degraded sericin polypeptides work as a protein matrix to which CSTI-IV may bind ionically.     

Interfacial behavior of fatty-acylated sericin prepared by lipase-catalyzed solid-phase synthesis

Fatty-acylated sericin {1:0.7 molar ratio of sericin (Mr 18,700) to oleic acid} was prepared by lipase-catalyzed solid-phase synthesis in n-hexane containing oleic acid to endow sericin with interfacial properties. Acylation with oleic acid was confirmed by 1H-NMR. The fatty-acylated sericin exhibited superior emulsifying activity index and emulsion stability in the presence of 0-0.5 M NaCl, in a temperature range of 30-80 degrees C and pH range of 2-7, as compared with the control sericin. The fatty-acylated sericin (1:0.4 molar ratio) prepared by using low-molecular-weight sericin (Mr 5,000) also exhibited superior emulsifying properties. The affinity of the fatty-acylated sericin to a hydrophobic surface as evaluated by a biomolecular interaction analyzer was about twice as much as that of the control sericin. The fatty-acylated sericin showed retarded water vaporization, similar to the control sericin, indicating good retention of moistness, and was adsorbed four times as much to defatted wool with little desorption as compared with the control sericin.     

Mechanical, physicochemical and color properties of chitosan based-films as a function of Aloe vera gel incorporation

The objective of the present study was to investigate the effect of Aloe vera gel incorporation at different proportions on chitosan-based films. Consequently, the thickness of films was affected significantly by the addition of the gel and decreased from F0 (plain chitosan film) to F50 (the film containing 50% gel). The gel incorporation did not have a considerable effect on water vapor permeability (WVP); however, a significant difference was observed for F50. Addition of the gel significantly improved the water solubility (WS), wherein the F10 (the film with 10% of gel) showed the lowest. All mechanical properties increased by introducing the gel and, after reaching the peak for F20, started to reduce. Color properties were affected by the gel addition as the higher the gel, the darker the films. Overall, the results showed that incorporating the gel into film-forming solution of chitosan up to 20% (F20) was promising.     

Development and in vitro evaluation of chitosan-Eudragit RS 30D composite wound dressings

The purpose of this research was to design and evaluate chitosan-based films intended for wound dressing application. Required properties for successful wound dressing, such as liquid uptake, vapor and oxygen penetration, bioadhesiveness, and film elasticity, were examined. Water uptake and vapor penetration of the films were determined gravimetrically, while oxygen penetration was determined by Winkler’s method. The bioadhesive properties were determined with an in-house pulley system instrument using a pig gut model. Film elasticity was determined with a stretch test using an Instron apparatus. The results showed that pure chitosan films exhibited relatively high liquid uptake and the adsorption tended to decrease with the addition of Eudragit RS 30D. Moisture vapor and oxygen were found to be able to penetrate through all film formulations in comparable amounts. The bioadhesiveness test tended to show lower bioadhesive properties with the addition of Eudragit RS 30D. The formulation containing only chitosan exhibited low elongation of the film at 2 N, but the film elasticity increased with the addition of Eudragit RS 30D. In conclusion, the addition of Eudragit RS 30D could improve a film’s mechanical properties but lower its bioadhesiveness. Published: March 24, 2006     

Sericin, a protein derived from silkworms, accelerates the proliferation of several mammalian cell lines including a hybridoma

Sericin, a constituent of the silkworm cocoon, was added to the culture of four mammalian cell lines: murine hybridoma 2E3-O,human hepatoblastoma HepG2, human epithelial HeLa and human embryonal kidney 293 cells. The proliferation of all cell lineswas accelerated in the presence of sericin. The hybridoma cellline was further studied. The 2E3-O cell line was so well adapted to serum-free medium that both the proliferation rate and maximum cell density in serum-free ASF103 medium were higher than in RPMI medium supplemented with all lots of FBS tested, and this proliferation was stimulated by the addition of sericin in a dose-dependent manner. Stimulation was observed at sericin concentrations from 0.01 to 0.1 %, although 1% sericin was severely harmful to the culture. In comparison with bovine serum albumin (BSA), a widely used supplement in serum-free medium, sericin had an equivalent effect on the proliferation of the hybridomas and sericin additively stimulated the proliferation with BSA. Although heat easily denatures and inactivates most proteins, the activity of sericin was not affected by autoclaving. In a similar manner to the silkworm-derived sericin, recombinant sericin synthesized in E. coli also stimulated the hybridoma proliferation, irrespective of whether it was autoclaved or filtered. Since BSA is obtained from bovine serum and the risk of infections such as bovine spongiform encephalopathy cannot be eradicated, sericin derived from insects could be a preferable culture medium supplement for stimulating the proliferation of mammalian cells. This revised version was published online in August 2006 with corrections to the Cover Date.     

Promotive effects of a silk film on epidermal recovery from full-thickness skin wounds

We examined the effects of the transparent fibroin film (silk film) on full-thickness skin wounds. Full-thickness dermatotomies (15 mm x 9 mm) were prepared on the dorsal wall of CRJ:CD-1 nu/nu (ICR nu/nu) mice. The area of the wounds dressed with silk film was reduced to 10% of that made by the dermatotomy 14 days after the dermatotomy and were covered with regenerated epidermis 21 days after the dermatotomy. In contrast, less recovery and epidermal regeneration were found 14 days after dermatotomy in the wounds dressed with a conventional hydrocolloid dressing (Duro Active). Furthermore, only partial incomplete epidemal growth was obtained 21 days after dermatotomy. Most importantly, the healing time of wounds dressed with silk film was 7 days shorter than those dressed with DuoActive dressing. The silk film showed an almost similar or slightly better promotive effect as the lyophilized porcine dermis (Alloask D), which is used as a dressing for burns, ulcers, and decubitis. Histologic findings revealed that there was greater collagen regeneration and less inflammation and neutrophil-lymphocyte infiltration of the wounds dressed with silk film than with DuoActive dressing. It is clear that regeneration of the epidermis and dermis of the wound beds covered with silk film was faster than with DuoActive dressing. Finally, silk film is easily obtainable, sterilizable, and transparent, and it allows easy observation of tissue recovery. Therefore, silk film offers advantages over other dressings and may be clinically useful for wound treatment.     

A biomimetic chitosan composite with improved mechanical properties in wet conditions

Chitosan is one of the most widely used structural polymers for biomedical applications because it has many favorable properties. However, one of the most critical drawbacks regarding the use of chitosan as a biomedical material is its poor mechanical properties in wet conditions. Here, we designed a method to improve the mechanical properties of chitosan in wet conditions and minimized the swelling behavior of chitosan film due to water adsorption by mimicking the sclerotization of insect cuticles and squid beaks, that is, catechol‐meditated crosslinking. The biomimetic chitosan composite film was prepared by mixing chitosan with l ‐3,4‐dihydroxyphenylalanine (DOPA) as a catecholic crosslinker and sodium periodate as an oxidant. The catechol‐meditated crosslinking provided a sevenfold enhancement in the stiffness in wet conditions compared to pure chitosan films and reduced the swelling behavior of the chitosan film. This strategy expands the possible applications for the use of chitosan composites as load‐bearing biomaterials. © 2013 American Institute of Chemical Engineers Biotechnol. Prog., 29: 505–512, 2013     

一种可止血的富血小板血浆壳聚糖/丝素蛋白多孔伤口敷料的制备及性能表征

为了进一步提高伤口敷料的止血性能,文中在生物相容性良好的壳聚糖溶液中引入含有多种生长因子的人源性富血小板血浆(Humanplatelet-richplasma,hPRP),并加入不同体积比例(1∶1、1∶3、3∶1、1∶0)的丝素蛋白溶液以提高材料的多孔性与止血性,通过冷冻干燥法制备不同配比的hPRP-壳聚糖/丝素蛋白敷料,并将纯壳聚糖敷料作为对照组,研究hPRP和丝素蛋白对敷料的止血性能的影响以及丝素蛋白对PRP中生长因子控制释放的影响。结果表明,在壳聚糖敷料中引入hPRP对敷料的止血性有所提高,但对敷料的多孔结构及吸水率无明显改善,若在hPRP-壳聚糖溶液中按照体积比为1∶1的比例加入丝素蛋白溶液,会得到具有较为均匀的多孔结构的敷料,敷料的孔隙率与吸水率分别可达到86.83%±3.84%与1 474%±114%,且该比例的敷料在快速止血性能上表现优异。此外,加入丝素蛋白与壳聚糖比例为1∶1的PRP敷料能有效减少PRP中生长因子在初始阶段的爆裂释放。因此,含hPRP的壳聚糖/丝素蛋白复合敷料有望成为一种能快速止血且能促进伤口愈合的新型伤口敷料。     

Fibroin and Sericin from Bombyx mori Silk Stimulate Cell Migration through Upregulation and Phosphorylation of c-Jun

Wound healing is a biological process directed to the restoration of tissue that has suffered an injury. An important phase of wound healing is the generation of a basal epithelium able to wholly replace the epidermis of the wound. A broad range of products derived from fibroin and sericin from Bombyx mori silk are used to stimulate wound healing. However, so far the molecular mechanism underlying this phenomenon has not been elucidated. The aim of this work was to determine the molecular basis underlying wound healing properties of silk proteins using a cell model. For this purpose, we assayed fibroin and sericin in a wound healing scratch assay using MDA-MB-231 and Mv1Lu cells. Both proteins stimulated cell migration. Furthermore, treatment with sericin and fibroin involved key factors of the wound healing process such as upregulation of c-Jun and c-Jun protein phosphorylation. Moreover, fibroin and sericin stimulated the phosphorylation of ERK 1/2 and JNK 1/2 kinases. All these experiments were done in the presence of specific inhibitors for some of the cell signalling pathways referred above. The obtained results revealed that MEK, JNK and PI3K pathways are involved in fibroin and sericin stimulated cells migration. Inhibition of these three kinases prevented c-Jun upregulation and phosphorylation by fibroin or sericin. Fibroin and sericin were tested in the human keratinocyte cell line, HaCaT, with similar results. Altogether, our results showed that fibroin and sericin initiate cell migration by activating the MEK, JNK and PI3K signalling pathways ending in c-Jun activation.     

Proteomic analysis of sericin in <Emphasis Type="Italic">Bombyx mori</Emphasis> cocoons

Cocoon sericin plays an important role in the reeling of silk and serves as a valuable biomaterial in the field of biomedicine, skincare, and food industries; however, knowledge about cocoon sericin proteins has been limited. For a comprehensive study on sericin, cocoons of eight varieties of silkworm of different geographic origin and with varied cocoon color were analyzed utilizing proteomics and bioinformatics approaches. The electrophoresis pattern demonstrated some common protein bands for all silkworm varieties and distinctive protein bands for some of those examined in the present study. The Ser2 protein, a new Ser3 protein, and four other novel sericin proteins were identified in cocoons for the first time. Products of both Ser1 and Ser3 genes appear to be ubiquitous in the cocoon shell of Bombyx mori. In addition, cocoons with especially high-reelability produced by the mutant strain B84 had an unique protein product of the Ser2 gene, indicating that the protein may play an important role in cocoon reelability. A series of sequence conflicts and post-translational modifications (PTMs) were also revealed in sericin proteins. Lipid modifications of sericin proteins, which promote waterproofing of the cocoon shell, were observed. Further, hydroxylation was identified, which provided evidence for intermolecular bonds among neighboring molecules of sericin as found in collagens. The sericin proteome data obtained from this study illuminated the molecular complexity of cocoon sericin and contributed to our understanding of the properties of sericin in filature and biomaterials.     

Flexible chitin films: structural studies

Chitin gels were transformed into thin, flexible chitin films with minimal dimensional shrinkage and maximum flexibility and thickness in the range of 25-80 microm by a cold-press process. Solvent residue was removed by heating the films at 50 degrees C for 12 h, followed by rinsing in 95% ethanol. The crystallinity and mechanical properties of the flexible chitin films were found to be a function of the amount of shrinkage from the gel to the final film that was obtained. For 28-microm thick films with 30% shrinkage, transparency of up to 90% was found. X-ray diffractometry (XRD) showed that the number of diffraction peaks appearing at 2theta;=23 degrees and 2theta;=27 degrees became increasingly sharper with shrinkage. Topographical information obtained from scanning electron microscopy (SEM) and atomic force microscopy (AFM) attributed the structural morphology of the films to the formation of sub-microscopic micelles. Scanning transmission electron microscopy (STEM) showed that shrinkage resulted in coarser microstructure, affecting tensile properties, where the ductility and toughness were proportional to the amount of shrinkage. These flexible chitin films have potential as wound dressing materials.     

Latest Advances on Bacterial Cellulose‐Based Materials for Wound Healing,Delivery Systems,and Tissue Engineering

Bacterial cellulose (BC) is a nanocellulose form produced by some nonpathogenic bacteria. BC presents unique physical, chemical, and biological properties that make it a very versatile material and has found application in several fields, namely in food industry, cosmetics, and biomedicine. This review overviews the latest state‐of‐the‐art usage of BC on three important areas of the biomedical field, namely delivery systems, wound dressing and healing materials, and tissue engineering for regenerative medicine. BC will be reviewed as a promising biopolymer for the design and development of innovative materials for the mentioned applications. Overall, BC is shown to be an effective and versatile carrier for delivery systems, a safe and multicustomizable patch or graft for wound dressing and healing applications, and a material that can be further tuned to better adjust for each tissue engineering application, by using different methods.