Ye Ri Park

Wound healing effect of electrospun silk fibroin nanomatrix in burn-model.

Silk fibroin has recently become an important biomaterial for tissue engineering application. In this study, silk fibroin nanomatrix was fabricated by electrospinning and evaluated as wound dressing material in a burn rat model. The wound size reduction, histological examination, and the quantification of transforming growth factor TGF-β1 and interleukin IL-1α, 6, and 10 were measured to evaluate the healing effects. The silk fibroin nanomatrix treatment exhibited effective performance in decreasing the wound size and epithelialization. Histological finding also revealed that the deposition of collagen in the dermis was organized by covering the wound area in the silk fibroin nanomatrix treated group. The expression level of pro-inflammatory cytokine (IL-1α) was significantly reduced in the injured skin following the silk fibroin nanomatrix treatment compared to the medical gauze (control) at 7 days after burn. Also, the expression level of TGF-β1 in the wound treated with silk fibroin nanomatrix peaked 21-days post-treatment whereas expression level of TGF-β1 was highest at day 7 in the gauze treated group. In conclusion, this data demonstrates that silk fibroin nanomatrix enhances the burn wound healing, suggesting it is a good candidate for burn wound treatment.

Three-dimensional electrospun silk-fibroin nanofiber for skin tissue engineering.

Tissue-engineered skin substitutes may offer an effective therapeutic option for the treatment of patients with skin damages. In this study, a novel three-dimensional (3D) scaffold composed of electrospun silk fibroin (SF) nanofiber was fabricated using electrospinning with the addition of NaCl crystals. It has well known that the electrospun SF nanofibers were excellent scaffold for tissue. However, it is generally difficult for cells to infiltrate the electrospun silk fibroin due to its small pore size. To resolve this problem, we dropped the NaCl crystals above the rotating collector, which become incorporated into the nanofibers. Three methods (freeze-drying, salt-leaching, and electrospinning with NaCl) for fabrication of SF scaffolds were compared to the difference of their characteristics using scanning electron microscopy (SEM), Fourier transform infrared spectroscopy (FTIR), mechanical strength, porosity, swelling abilities, and cell proliferation. Additionally, using air-liquid culture system, keratinocytes were co-cultured with fibroblasts in each type of SF scaffolds to construct an artificial bilayer skin in vitro. In our experimental results, histologic findings in only electrospun SF scaffolds showed more proliferation of fibroblasts in deep layer and more differentiation of keratinocytes in superficial layer. The present study suggests that 3D electrospun SF scaffolds might be a suitable for skin tissue engineering.

Fabrication of duck’s feet collagen–silk hybrid biomaterial for tissue engineering

Collagen constituting the extracellular matrix has been widely used as biocompatible material for human use. In this study, we have selected ducks feet for extracting collagen. A simple method not utilizing harsh chemical had been employed to extract collagen from ducks feet. We fabricated ducks feet collagen/silk hybrid scaffold for the purpose of modifying the degradation rate of ducks feet collagen. This study suggests that extracted collagen from ducks feet is biocompatible and resembles collagen extracted from porcine which is commercially used. Ducks feet collagen is also economically feasible and it could therefore be a good candidate as a tissue engineering material. Further, addition of silk to fabricate a ducks feet collagen/silk hybrid scaffold could enhance the biostability of ducks feet collagen scaffold. Ducks feet collagen/silk scaffold increased the cell viability compared to silk alone. Animal studies also showed that ducks feet collagen/silk scaffold was more biocompatible than silk alone and more biostable than ducks feet or porcine collagen alone. Additionally, the results revealed that ducks feet collagen/silk hybrid scaffold had high porosity, cell infiltration and proliferation. We suggest that ducks feet collagen/silk hybrid scaffold could be used as a dermal substitution for full thickness skin defects.

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

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

Novel fabrication method of the peritoneal dialysis filter using silk fibroin with urease fixation system

During the last decade, there has been a great advance in the kidney dialysis system by wearable artificial kidney (WAK) system for end-stage renal disease patients. Uremic solute removal and water regeneration system are the most prerequisite for WAK to work properly. In this study, we designed a filtering membrane system by using immobilized urease silk fibroin filter and evaluated its comparative effectiveness with a PVDF filtering system in peritoneal dialysate regeneration system by urea removal efficacy. We evaluated this membranes characteristic and performances by conducting SEM-EDX analyze, water-binding abilities and porosity test, removal abilities of urea, cytotoxicity assay and enzyme activity assay. Under the condition for optimization of urease, the percentage removal of urea was about 40% and 60% in 50 mg/dL urea solution by urease immobilized PVDF and silk fibroin scaffolds, respectively. The batch experimental result showed that immobilized filter removed more than 50% of urea in 50 mg/dL urea solution. In addition silk fibroin with urease filter removed 90 percent of urea in the peritoneal dialysate after 24 h filtration. We suggest that silk fibroin with urease fixation filter can be used more effectively for peritoneal dialysate regeneration system, which have hydrophilic property and prolonged enzyme activity.

Application of a Collagen Patch Derived from Duck Feet in Acute Tympanic Membrane Perforation

We investigated the utility of the duck-feet collagen extraction patching procedure in the traumatic tympanic membrane (TM) perforation via a comparison with spontaneous healing or paper patch. Fifty-six ears of adult male Sprague-Dawley rats, each weighing in the range of 250 to 300 g, were used for the animal studies. Sixteen rats had one-side ear in the control group and the opposite-side ear in the treated groups. The remaining twelve rats had a one-side ear with the duck-feet collagen patch and the opposite-side ear with a paper patch. The proliferating cell nuclear antigen (PCNA) expression cells were calculated among the 200 basal cells, and the expression percentage was identified as the labeling index. The healing of the perforation in the duck-feet collagen patch group was confirmed to be more rapid compared to the healing of the other groups. PCNA staining was observed in the migrating portion of PCNA enhanced cell to collagen scaffold in Post operative day (POD) 7 of collagen patch group. Thus, the adhesive effect of the duck-feet collagen patch to perforated margin was better than that of the paper patch. After completing the healing process, the collagen patch shrank and detached from the tympanic membrane (POD 14). In this study, we confirmed that the use of a duck-feet collagen patch had the advantage of early healing, inducing natural TM contour, and disappearing ability after the patch effect is complete.

NF-κB signaling is key in the wound healing processes of silk fibroin

Silk fibroin (SF) is a well-studied biomaterial for tissue engineering applications including wound healing. However, the signaling mechanisms underlying the impact of SF on this phenomenon have not been determined. In this study, through microarray analysis, regulatory genes of NF-ĸB signaling were activated in SF-treated NIH3T3 cells along with other genes. Immunoblot analysis confirmed the activation of the NF-ĸB signaling pathway as SF induced protein expression levels of IKKα, IKKβ, p65, and the degradation of IκBα. The treatment of NIH3T3 cells with SF also increased the expression of cyclin D1, vimentin, fibronectin, and vascular endothelial growth factor (VEGF). The expression of these factors by SF treatment was abrogated when NF-ĸB was inhibited by a pharmacological inhibitor Bay 11-7082. Knockdown of NF-ĸB using siRNA of IKKα and IKKβ also inhibited the SF-induced wound healing response of the NIH3T3 cells in a wound scratch assay. Collectively, these results indicated that SF-induced wound healing through the canonical NF-κB signaling pathway via regulation of the expression of cyclin D1, vimentin, fibronectin, and VEGF by NIH3T3 cells. Using an in vivo study with a partial-thickness excision wound in rats we demonstrated that SF-induced wound healing via NF-κB regulated proteins including cyclin D1, fibronectin, and VEGF. The in vitro and in vivo data suggested that SF induced wound healing via modulation of NF-ĸB signaling regulated proteins. STATEMENT OF SIGNIFICANCE Silk fibroin has been effectively used as a dressing for wound treatment for more than a century. However, mechanistic insight into the basis for wound healing via silk fibroin has not been elucidated. Here we report a key mechanism involved in silk fibroin induced wound healing both in vitro and in vivo. Using genetic- and protein-level analyses, NF-κB signaling was found to regulate silk fibroin-induced wound healing by modulating target proteins. Thus, the NF-κB signaling pathway may be utilized as a therapeutic target during the formulation of silk fibroin-based biomaterials for wound healing and tissue engineering.

In vitro and in vivo evaluation of the duck's feet collagen sponge for hemostatic applications

Recently different hemostatic agents have been developed, but most of them are ineffective in severe bleeding and expensive or cause safety concerns. In this study, we fabricated ducks feet collagen-based porous sponges and investigated its use as a hemostatic agent. We determined the sponges physical and biological characteristics and compared with Avitene via scanning electron microscope analysis, water-uptake abilities and porosity test, and cytotoxicity assay. The ducks feet collagen/silk sponge showed a larger interconnected porous structure compared to others sponges. The ducks feet collagen/silk sponge also exhibited significantly higher porosity than Avitene. Hemostatic properties of the sponges were evaluated by whole blood clotting and rat femoral artery hemorrhage experiment. The addition of silk to ducks feet collagen showed better blood clotting ability than Avitene in vitro. However, rat femoral artery hemorrhage test showed a similar hemostatic property between the ducks feet collagen-based sponges and Avitene. We suggest that ducks feet collagen-based sponge can be effectively used for hemostatic applications.