Acellular bi-layer silk fibroin scaffolds support functional tissue regeneration in a rat model of onlay esophagoplasty

Abstract

Surgical management of long-gap esophageal defects with autologous gastrointestinal tissues is frequently associated with adverse complications including organ dysmotility, dysphagia, and donor site morbidity. In order to develop alternative graft options, bi-layer silk fibroin (SF) scaffolds were investigated for their potential to support functional tissue regeneration in a rodent model of esophageal repair. Onlay esophagoplasty was performed with SF matrices (N=40) in adult rats for up to 2 m of implantation. Parallel groups consisted of animals implanted with small intestinal submucosa (SIS) scaffolds (N=22) or sham controls receiving esophagotomy alone (N=20). Sham controls exhibited a 100% survival rate while rats implanted with SF and SIS scaffolds displayed respective survival rates of 93% and 91% prior to scheduled euthanasia. Animals in each experimental group were capable of solid food consumption following a 3 d post-op liquid diet and demonstrated similar degrees of weight gain throughout the study period. End-point μ-computed tomography at 2 m post-op revealed no evidence of contrast extravasation, fistulas, strictures, or diverticula in any of the implant groups. Ex vivo tissue bath studies demonstrated that reconstructed esophageal conduits supported by both SF and SIS scaffolds displayed contractile responses to carbachol, KCl and electrical field stimulation while isoproterenol produced tissue relaxation. Histological (Masson’s trichrome and hematoxylin and eosin) and immunohistochemical (IHC) evaluations demonstrated both implant groups produced de novo formation of skeletal and smooth muscle bundles positive for contractile protein expression [fast myosin heavy chain (MY32) and α-smooth muscle actin (α-SMA)] within the graft site. However, SF matrices promoted a significant 4-fold increase in MY32+ skeletal muscle and a 2-fold gain in α-SMA+ smooth muscle in comparison to the SIS cohort as determined by histomorphometric analyses. A stratified squamous, keratinized epithelium expressing cytokeratin 5 and involucrin proteins was also present at 2 m post-op in all experimental groups. De novo innervation and vascularization were evident in all regenerated tissues indicated by the presence of synaptophysin (SYP38)+ boutons and vessels lined with CD31 expressing endothelial cells. In respect to SIS, the SF group supported a significant 4-fold increase in the density of SYP38+ boutons within the implant region. Evaluation of host tissue responses revealed that SIS matrices elicited chronic inflammatory reactions and severe fibrosis throughout the neotissues, in contrast to SF scaffolds. The results of this study demonstrate that bi-layer SF scaffolds represent promising biomaterials for onlay esophagoplasty, capable of producing superior regenerative outcomes in comparison to conventional SIS scaffolds.

Mode of Surgical Injury Influences the Source of Urothelial Progenitors during Bladder Defect Repair

“…Sample visualization was performed with an Axioplan-2 microscope (Carl Zeiss MicroImaging, Thornwood, NY) and representative fields were acquired with Axiovision software (version 4.8). Histomorphometric evaluations (n = 3–4 animals per group) were performed on three independent microscopic fields (20× magnification) using published methods ( Algarrahi et al., 2015 ) to quantify urothelial subpopulations in experimental cohorts.…”

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

“…Over the past three decades, a number of acellular biodegradable biomaterials have been explored as alternatives to autologous tissue flaps for repair of partial circumferential, full thickness esophageal defects (Tan et al ., 2012). Patch grafts derived from decellularized tissues such as small intestinal submucosa and urinary bladder submucosa have been utilized for esophageal defect consolidation in a variety of preclinical animal models and short-term clinical trials (Badylak et al ., 2000; Lopes et al ., 2006a, 2006b; Urita et al, 2007; Clough et al ., 2011; Nieponice et al ., 2014; Algarrahi et al ., 2015). However, suboptimal regenerative responses and complications such as scaffold leakage have been reported with these matrix designs, therefore raising doubts about their widespread translation into clinical settings (Urita et al, 2007; Nieponice et al ., 2014; Algarrahi et al ., 2015).…”

“…Clinically viable biomaterial configurations for patch esophagoplasty must possess optimal structural, mechanical and degradative characteristics sufficient to provide for initial defect reinforcement, but allow for gradual scaffold dissipation and subsequent constructive remodeling without eliciting adverse immune reactions. A recent study from our laboratory has shown that bi-layer silk fibroin (BLSF) matrices derived from Bombyx mori silkworm cocoons exhibited such properties in a rat model of onlay esophagoplasty (Algarrahi et al ., 2015). This biodegradable scaffold design displays dual functionality during wound healing by providing a porous foam compartment permissive for host tissue integration while a buttressing film layer prevents leakage of luminal contents (Seth et al ., 2013).…”

Polyurethane scaffolds seeded with autologous cells can regenerate long esophageal gaps: An esophageal atresia treatment model

“…These studies describe regeneration of the esophagus in up to 4 weeks with the presence of an epithelial layer and a disorganized or absent muscle layer. However, more recent studies using silk scaffolding [39][40][41], Poly lactic-co-glycolic acid (PLGA) [42,43], and polyurethane (PU) [28] describe smooth muscle and fibroblast regeneration followed by a squamous epithelial lining. The second objective of this study was to evaluate the feasibility of bridging a long intrathoracic esophageal gap with either an autologous EEC seeded or AF-MSC seeded polyurethane (Biostage™) graft in a porcine model of esophageal loss.…”