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Bi-layer silk fibroin grafts support functional tissue regeneration in a porcine model of onlay esophagoplasty.

ABSTRACT: 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.

SUBMITTER: Algarrahi K

PROVIDER: S-EPMC5544592 | biostudies-literature | 2018 Feb

REPOSITORIES: biostudies-literature

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Bi-layer silk fibroin grafts support functional tissue regeneration in a porcine model of onlay esophagoplasty.

Algarrahi Khalid K Franck Debra D Cristofaro Vivian V Yang Xuehui X Savarino Alyssa A Affas Saif S Schäfer Frank-Mattias FM Ghezzi Chiara C Jennings Russell R Nedder Arthur A Kaplan David L DL Kaplan David L DL Sullivan Maryrose P MP Estrada Carlos R CR Mauney Joshua R JR

Journal of tissue engineering and regenerative medicine 20170620 2

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 p ...[more]

PMID: 28084044

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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.

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.

Dataset Information

Bi-layer silk fibroin grafts support functional tissue regeneration in a porcine model of onlay esophagoplasty.

Publications

Bi-layer silk fibroin grafts support functional tissue regeneration in a porcine model of onlay esophagoplasty.

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