Acellular Bi-Layer Silk Fibroin Scaffolds Support Tissue Regeneration in a Rabbit Model of Onlay Urethroplasty

Abstract

Acellular scaffolds derived from Bombyx mori silk fibroin were investigated for their ability to support functional tissue regeneration in a rabbit model of urethra repair. A bi-layer silk fibroin matrix was fabricated by a solvent-casting/salt leaching process in combination with silk fibroin film casting to generate porous foams buttressed by homogeneous silk fibroin films. Ventral onlay urethroplasty was performed with silk fibroin grafts (Group 1, N = 4) (Width×Length, 1×2 cm2) in adult male rabbits for 3 m of implantation. Parallel control groups consisted of animals receiving small intestinal submucosa (SIS) implants (Group 2, N = 4) or urethrotomy alone (Group 3, N = 3). Animals in all groups exhibited 100% survival prior to scheduled euthanasia and achieved voluntary voiding following 7 d of initial catheterization. Retrograde urethrography of each implant group at 3 m post-op revealed wide urethral calibers and preservation of organ continuity similar to pre-operative and urethrotomy controls with no evidence of contrast extravasation, strictures, fistulas, or stone formation. Histological (hematoxylin and eosin and Masson's trichrome), immunohistochemical, and histomorphometric analyses demonstrated that both silk fibroin and SIS scaffolds promoted similar extents of smooth muscle and epithelial tissue regeneration throughout the original defect sites with prominent contractile protein (α-smooth muscle actin and SM22α) and cytokeratin expression, respectively. De novo innervation and vascularization were also evident in all regenerated tissues indicated by synaptophysin-positive neuronal cells and vessels lined with CD31 expressing endothelial cells. Following 3 m post-op, minimal acute inflammatory reactions were elicited by silk fibroin scaffolds characterized by the presence of eosinophil granulocytes while SIS matrices promoted chronic inflammatory responses indicated by mobilization of mononuclear cell infiltrates. The results of this study demonstrate that bi-layer silk fibroin scaffolds represent promising biomaterials for onlay urethroplasty, capable of promoting similar degrees of tissue regeneration in comparison to conventional SIS scaffolds, but with reduced immunogenicity.

Silk structure and degradation

“…Since sericin may have a stronger immunogenicity, silk fibroin [4][5][6][7][8][9][10][11][12][13][14][15] or degummed raw silk [17][18][19] is used in biomedical research. In this study, raw silk and degummed raw silk were implanted subcutaneously into ten Sprague-Dawley rats.…”

“…In the degradation process, cells play a significant role in both the in vitro and in vivo environments [13,20,25]. At present, only a few studies have focused directly on investigating the degradation mechanism of the materials or the role tissue solution and cells play during the in vivo degradation process.…”

“…Degradation of degummed silk [17,18], degummed silkprepared surgical sutures [19], or regenerated silk fibroin [1,2,[4][5][6][7][8][9][10][11][12][13][14][15][16][17][18][19][20]25,26] is attributed to the degradation of silk fibroin. While the degradation properties of silk fibroin have been the subject of intensive investigation, the degradation properties of raw silk remain unclear.…”

Biomatrices in Urethral Reconstruction

“…Silk fibroin (SF) is a novel protein obtained from Bombyx mori cocoons that have good biocompatibility and low immunogenicity [26,27]. The mechanical property and microstructure of was improved after it was stretched in 90% ethanol [28].…”

“…The mechanical property and microstructure of was improved after it was stretched in 90% ethanol [28]. Both accellular and cell seeded SF showed good efficacy in urethral reconstruction in animal model [26,29,30]. Recently, Lv et al [31] reported a novel oxygen-generating material composed of SF, keratin, calcium peroxide and gelatin.…”