Molecular mechanisms of esophageal epithelial regeneration following repair of surgical defects with acellular silk fibroin grafts.
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ABSTRACT:
SUBMITTER: Gundogdu G
PROVIDER: S-EPMC8007829 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Bi-layer silk fibroin (BLSF) scaffolds represent an emerging technology in the development of acellular biomaterials for esophageal repair. Their therapeutic potential is attributed to their robust mechanical properties, elasticity, and low immunogenicity. However, the underlying molecular mechanisms of scaffold-mediated wound healing processes in the esophagus remain unclear. Previous research has identified signaling cascades involved in neoepithelial regeneration in a rat model of onlay esophagoplasty with BLSF grafts, including c-MET, TrkA, and PI3K-Akt signaling. Interestingly, these pro-survival signaling cascades are largely governed by DNA methylation (DNAme). Such findings motivate us to apply reduced-representation bisulfite sequencing (RRBS) to characterize the temporal dynamics of DNAme in host and regenerated tissues up to 1 week post-operation. Globally, we observe hypermethylation at post-surgical repair timepoints and an inverse correlation between DNAme and the expression levels of differentially expressed proteins during regeneration. Site-specific hypomethylation targets genes associated with immune activation, which may be indicative of immune cell infiltration at the surgical implant site, while site-specific hypermethylation appears to target PI3K-Akt signaling. Our work provides mechanistic insight into the molecular processes governing esophageal regeneration, which can motivate therapeutic innovations toward temporal modulation of tissue regeneration during scaffold-mediated esophageal repair via epigenetic regulators that target key pathways.
2023-05-12 | GSE203028 | GEO
Project description:DNA methylation dynamics during esophageal epithelial regeneration following repair with acellular silk fibroin grafts in rat
| PRJNA838203 | ENA
Project description:Partial circumferential, full thickness defects of the esophagus can occur as a result of organ perforation or tumour resection, or during surgical reconstruction of strictured segments. Complications associated with autologous tissue flaps conventionally utilized for defect repair necessitate the development of new graft options. In this study, bi-layer silk fibroin (BLSF) scaffolds were investigated for their potential to support functional restoration of partial circumferential defects in a porcine model of esophageal repair. Onlay thoracic esophagoplasty with BLSF matrices (~3 x 1.5 cm) was performed in adult swine (N = 6) for 3 months of implantation. All animals receiving BLSF grafts survived with no complications and were capable of solid food consumption. Radiographic esophagrams revealed preservation of organ continuity with no evidence of contrast extravasation or strictures. Fluoroscopic analysis demonstrated peristaltic contractions. Ex vivo tissue bath studies displayed contractile responses to carbachol, electric field stimulation, and KCl while isoproterenol produced tissue relaxation. Histological and immunohistochemical evaluations of neotissues showed a stratified, squamous epithelium, a muscularis mucosa composed of smooth muscle bundles, and a muscularis externa organized into circular and longitudinal layers, with a mix of striated skeletal muscle fascicles interspersed with smooth muscle. De novo innervation and vascularization were observed throughout the graft sites and consisted of synaptophysin-positive neuronal boutons and vessels lined with CD31-positive endothelial cells. The results of this study demonstrate that BLSF scaffolds can facilitate constructive remodeling of partial circumferential, full thickness esophageal defects in a large animal model. Copyright © 2017 John Wiley & Sons, Ltd.
| S-EPMC5544592 | biostudies-literature
Project description:Successful repair of a damaged corneal surface is a great challenge and may require the use of a scaffold that supports cell growth and differentiation. Amniotic membrane is currently used for this purpose, in spite of its limitations. A thin transparent silk fibroin film from non-mulberry Antheraea mylitta (Am) has been developed which offers to be a promising alternative. The silk scaffolds provide sufficient rigidity for easy handling, the scaffolds support the sprouting, migration, attachment and growth of epithelial cells and keratocytes from rat corneal explants; the cells form a cell sheet, preserve their phenotypes, express cytokeratin3 and vimentin respectively. The films also support growth of limbal stem cell evidenced by expression of ABCG2. The cell growth on the silk film and the amniotic membrane is comparable. The implanted film within the rabbit cornea remains transparent, stable. The clinical examination as well as histology shows absence of any inflammatory response or neovascularization. The corneal surface integrity is maintained; tear formation, intraocular pressure and electroretinography of implanted eyes show no adverse changes. The silk fibroin film from non-mulberry silk worms may be a worthy candidate for use as a corneal scaffold.
| S-EPMC4764817 | biostudies-other
Project description:Current synthetic vascular grafts are not suitable for use in low-diameter applications. Silk fibroin is a promising natural graft material which may be an effective alternative. In this study, we compared two electrospun silk grafts with different manufacturing processes, using either water or hexafluoroisopropanol (HFIP) as solvent. This resulted in markedly different Young's modulus, ultimate tensile strength and burst pressure, with HFIP spun grafts observed to have thicker fibres, and greater stiffness and strength relative to water spun. Assessment in a rat abdominal aorta grafting model showed significantly faster endothelialisation of the HFIP spun graft relative to water spun. Neointimal hyperplasia in the HFIP graft also stabilised significantly earlier, correlated with an earlier SMC phenotype switch from synthetic to contractile, increasing extracellular matrix protein density. An initial examination of the macrophage response showed that HFIP spun conduits promoted an anti-inflammatory M2 phenotype at early timepoints while reducing the pro-inflammatory M1 phenotype relative to water spun grafts. These observations demonstrate the important role of the manufacturing process and physical graft properties in determining the physiological response. Our study is the first to comprehensively study these differences for silk in a long-term rodent model.
| S-EPMC6877724 | biostudies-literature
Project description:Osteoarthritis (OA) is a common joint disease with a high disability rate. In addition, OA not only causes great physiological and psychological harm to patients, but also puts great pressure on the social healthcare system. Pathologically, the disintegration of cartilage and the lesions of subchondral bone are related to OA. Currently, tissue engineering, which is expected to overcome the defects of existing treatment methods, had a lot of research in the field of cartilage/osteochondral repair. Silk fibroin (SF), as a natural macromolecular material with good biocompatibility, unique mechanical properties, excellent processability and degradability, holds great potential in the field of tissue engineering. Nowadays, SF had been prepared into various materials to adapt to the demands of cartilage/osteochondral repair. SF-based biomaterials can also be functionally modified to enhance repair performance further. In this review, the preparation methods, types, structures, mechanical properties, and functional modifications of SF-based biomaterials used for cartilage/osteochondral repair are summarized and discussed. We hope that this review will provide a reference for the design and development of SF-based biomaterials in cartilage/osteochondral repair field.
| S-EPMC9274741 | biostudies-literature
Project description:The dissolution of Bombyx mori silk fibroin (SF) films in formic acid (FA) for the preparation of electrospinning dopes is widely exploited to produce electrospun SF scaffolds. The SILKBridge® nerve conduit is an example of medical device having in its wall structure an electrospun component produced from an FA spinning dope. Though highly volatile, residual FA remains trapped into the bulk of the SF nanofibers. The purpose of this work is to investigate the type and strength of the interaction between FA and SF in electrospun mats, to quantify its amount and to evaluate its possible toxicological impact on human health. The presence of residual FA in SF mats was detected by FTIR and Raman spectroscopy (new carbonyl peak at about 1,725 cm-1) and by solid state NMR, which revealed a new carbonyl signal at about 164.3 ppm, attributed to FA by isotopic 13C substitution. Changes occurred also in the spectral ranges of hydroxylated amino acids (Ser and Thr), demonstrating that FA interacted with SF by forming formyl esters. The total amount of FA was determined by HS-GC/MS analysis and accounted for 247 ± 20 μmol/g. The greatest part was present as formyl ester, a small part (about 3%) as free FA. Approximately 17% of the 1,500 μmol/g of hydroxy amino acids (Ser and Thr) theoretically available were involved in the formation of formyl esters. Treatment with alkali (Na2CO3) succeeded to remove the greatest part of FA, but not all. Alkali-treated electrospun SF mats underwent morphological, physical, and mechanical changes. The average diameter of the fibers increased from about 440 nm to about 480 nm, the mat shrunk, became stiffer (the modulus increased from about 5.5 MPa to about 7 MPa), and lost elasticity (the strain decreased from about 1 mm/mm to about 0.8 mm/mm). Biocompatibility studies with human adult dermal fibroblasts did not show significant difference in cell proliferation (313 ± 18 and 309 ± 23 cells/mm2 for untreated and alkali-treated SF mat, respectively) and metabolic activity. An in-depth evaluation of the possible toxicological impact of residual FA was made using the SILKBridge® nerve conduit as case study, following the provisions of the ISO 10993-1 standard. The Potential Patient Daily Intake, calculated from the total amount of FA determined by HS-GC/MS, was 2.4 mg/day and the Tolerable Exposure level was set to 35.4 mg/day. This allowed to obtain a value of the Margin of Safety of 15, indicating that the amount of FA left on SF mats after electrospinning does not raise concerns for human health.
| S-EPMC8829063 | biostudies-literature
Project description:As the incidence of cardiovascular diseases has been growing in recent years, the need for small-diameter vascular grafts is increasing. Considering the limited success of synthetic grafts, vascular tissue engineering/repair/regeneration aim to find novel solutions. Silk fibroin (SF) has been widely investigated for the development of vascular grafts, due to its good biocompatibility, tailorable biodegradability, excellent mechanical properties, and minimal inflammatory reactions. In this study, a new generation of three-layered SF vascular scaffolds has been produced and optimized. Four designs of the SILKGraft vascular prosthesis have been developed with the aim of improving kink resistance and mechanical strength, without compromising the compliance with native vessels and the proven biocompatibility. A more compact arrangement of the textile layer allowed for the increase in the mechanical properties along the longitudinal and circumferential directions and the improvement of the compliance value, which approached that reported for the saphenous and umbilical veins. The higher braid density slightly affected the grafts' morphology, increasing surface roughness, but the novel design mimicked the corrugation approach used for synthetic grafts, causing significant improvements in kink resistance.
| S-EPMC9147556 | biostudies-literature
Project description:Surgical reconstruction of tubular esophageal defects with autologous gastrointestinal segments is the gold standard treatment to replace damaged or diseased esophageal tissues. Unfortunately, this approach is associated with adverse complications, including dysphagia, donor-site morbidity, and in some cases patient death. Bilayer silk fibroin (BLSF) scaffolds were investigated as alternative, acellular grafts for tubular esophagoplasty in a porcine defect model for 3 months of implantation. Adult Yucatan mini-swine (n = 5) were subjected to esophageal reconstruction with tubular BLSF grafts (2 cm in length) in combination with transient esophageal stenting for 2 months followed by a 1-month period, where the graft site was unstented. All animals receiving BLSF grafts survived and were capable of solid food consumption, however strictures were noted at graft regions in 60% of the experimental cohort between 2 and 3 months postop and required balloon dilation. In addition, fluoroscopic analysis showed peristaltic function in only 1/5 neotissues. Following swine harvest at 3 months, ex vivo tissue bath evaluations revealed that neoconduits exhibited contractile responses to carbachol, electric field stimulation, and KCl, whereas sodium nitroprusside and isoproterenol induced relaxation effects. Histological (Masson's Trichrome) and immunohistochemical analyses of regenerated tissue conduits showed a stratified, squamous epithelium expressing pan-cytokeratins buttressed by a vascularized lamina propria containing a smooth muscle-rich muscularis mucosa surrounded by a muscularis externa. Neuronal density, characterized by the presence of synaptophysin-positive boutons, was significantly lower in neotissues in comparison to nonsurgical controls. BLSF scaffolds represent a promising platform for the repair of tubular esophageal defects, however improvements in scaffold design are needed to reduce the rate of complications and improve the extent of constructive tissue remodeling. Impact statement The search for a superior "off-the-shelf" scaffold capable of repairing tubularesophageal defects as well as overcoming limitations associated with conventional autologous gastrointestinal segments remains elusive. The purpose of this study was to investigate the performance of an acellular, bilayer silk fibroin graft (BLSF) for tubular esophagoplasty in a porcine model. Our results demonstrated that BLSF scaffolds supported the formation of tubular neotissues with innervated, vascularized epithelial and muscular components capable of contractile and relaxation responses. BLSF scaffolds represent a promising platform for esophageal tissue engineering.
| S-EPMC7826443 | biostudies-literature
Project description:Cartilage defects frequently occur around the knee joint yet cartilage has limited self-repair abilities. Hydrogel scaffolds have excellent potential for use in tissue engineering. Therefore, the aim of the present study was to assess the ability of silk fibroin (SF) hydrogel scaffolds incorporated with chitosan (CS) nanoparticles (NPs) to repair knee joint cartilage defects. In the present study, composite systems of CS NPs incorporated with transforming growth factor-β1 (TGF-β1; TGF-β1@CS) and SF incorporated with bone morphogenetic protein-2 (BMP-2; TGF-β1@CS/BMP-2@SF) were developed and characterized with respect to their size distribution, zeta potential, morphology, and release of TGF-β1 and BMP-2. Bone marrow stromal cells (BMSCs) were co-cultured with TGF-β1@CS/BMP-2@SF extracts to assess chondrogenesis in vitro using a cell counting kit-8 assay, which was followed by in vivo evaluations in a rabbit model of knee joint cartilage defects. The constructed TGF-β1@CS/BMP-2@SF composite system was successfully characterized and showed favorable biocompatibility. In the presence of TGF-β1@CS/BMP-2@SF extracts, BMSCs exhibited normal cell morphology and enhanced chondrogenic ability both in vitro and in vivo, as evidenced by the promotion of cell viability and the alleviation of cartilage defects. Thus, the TGF-β1@CS/BMP-2@SF hydrogel developed in the present study promoted chondrogenic ability of BMSCs both in vivo and in vitro by releasing TGF-β1 and BMP-2, thereby offering a novel therapeutic strategy for repairing articular cartilage defects in knee joints.
| S-EPMC7856775 | biostudies-literature