Urinary bladder reconstruction using autologous collagenous connective tissue membrane "Biosheet®" induced by in-body tissue architecture: A pilot study

Abstract

Introduction: In-body tissue architecture (iBTA) technology, based on cell-free tissue engineering, can produces collagenous tissues for implantation by subcutaneous embedding a designed mold. The aim of this study was to evaluate the biocompatibility of iBTA-induced "Biosheet®" collagenous sheets, as scaffold materials for bladder reconstruction.

Methods: Canine Biosheet® implants were prepared by embedding molds into subcutaneous pouches in beagles for 8 weeks. A part of canine bladder wall was excised (2 × 2 cm) and repaired by patching the same sized autologous Biosheet®. The Biosheet® implants were harvested 4 weeks (n = 1) and 12 weeks (n = 3) after the implantation and evaluated histologically.

Results: No disruption of the patched Biosheet® implants or urinary leakage into the peritoneal cavity was observed during the entire observation periods. There were no signs of chronic inflammation or Biosheet® rejection. The urine-contacting surface of luminal surface of the Biosheet® was covered with a multicellular layer of urothelium cells 4 weeks after implantation. α-SMA-positive muscle cells were observed at the margin of the Biosheet® implants at 12 weeks after the implantation. In addition, in the center of the Biosheet® implants, the formation of microvessels stained as α-SMA-positive was observed.

Conclusion: Biosheet® implants have biocompatibility as a scaffold for bladder reconstruction, indicating that they may be applicable for full-thickness bladder wall substitution. Further studies are required for definitive evaluation as a scaffold for bladder reconstruction.

Keywords: BAM, bladder acellular matrices; Biosheet®; Bladder reconstruction; In body tissue architecture; Regenerative medicine; SIS, small intestinal submucosa; Tissue engineering; Urinary bladder; iBTA, in-body tissue architecture.

Fig. 1

Photographs of the preparation and implantation of Biosheet® implants. (a) Mold for Biosheet® preparation (b) Mold encapsulated completely with Biosheet® tissue 8 weeks after implantation. (c) Biosheet® covered the mold after trimming to remove fragile and redundant tissue. (d) Urinary bladder wall after dissection (2 × 2 cm) (e) Trephined Biosheet® (2 cm in diameter) (f) Urinary bladder after implantation of Biosheet®.

Fig. 2

Typical images of urography and ultrasonography after implantation of Biosheet® implants. (a, d) Images of the urinary bladder at 0 days after implantation. (b, e) Images of the urinary bladder at 4 weeks after implantation. (c, f) Images of the urinary bladder at 12 weeks after implantation. White arrows indicate the implantation site of the Biosheet® implants.

Fig. 3

Macroscopic view of Biosheet® implants in the urinary bladder. (a, c) Urinary bladder at 4 and 12 weeks after implantation. (b, d) Mucous membrane of the Biosheet® implants at 4 and 12 weeks after implantation. The broken line indicates the implantation site of the Biosheet® implants.

Fig. 4

Histology of Biosheet® at 4 weeks after implantation. (a) Hematoxylin–eosin staining. Complete view of the Biosheet®. (b, c) Masson's Trichrome staining. Collagen is stained blue; nuclei are stained black. Mucosa and submucosa of the Biosheet® and native bladder are shown. Stratified urothelium covered the entire graft surface of the Biosheet® (black arrows). Neovascularization was observed in the loose connective tissue on the luminal (white arrows).

Fig. 5

Histology of Biosheet® implants at 12 weeks after implantation. (a) Complete view of the Biosheet®. (b, c) Mucosa and submucosa of the Biosheet® and native bladder (d, f) Margin of the Biosheet® and bladder. (e) Center of the Biosheet®. (g–i) Immunohistochemical staining for α-smooth muscle actin (α-SMA). Red: α-SMA, Blue: DAPI staining for cell nuclei. (g, i) Cells which stained positive for α-SMA recruited from native muscle tissues to the Biosheet®. (h) Microvessels that stained α-SMA-positive were observed at the center of the Biosheet® (white arrow heads).