Bladder acellular matrix seeding with autologous cultured vaginal smooth cells repaired rabbit vaginal defect

Abstract

Objective To evaluate the feasibility of bladder extracellular matrix (BACM) seeding with autologous cultured vaginal smooth muscle cells (VSMC) repaired rabbit vaginal defect.Methods This study included 24 female rabbits.BACM and vaginal acellular matrix (VACM) were obtained from 8 rabbits by decellularization process.The other 16 rabbits were randomly divided into experimental and control groups.Vaginal tissue biopsies ( ～ 1 cm2) were harvested from female New Zealand rabbits.VSMC were cultured and stained with a-smooth muscle actin antibodies. Cultured VSMC cells were seeded on BACM ( experimental group) or VACM ( control group) at a cell density of 1 × 107 cells/cm2.The cell-seeded matrixes were cultured for 5days.In experimental group of 8 rabbits,a 2 cm segment of vagina was resected and replaced with BACM seeding with VSMC.Then the regenerative segment was studied with histological technique by hematoxylin-eosin staining after 3,6 and 12 weeks postoperative aud vaginography was performed at 12 weeks postoperative.The 8 rabbits in control group underwent the exact same procedure as above but the vaginal defect was repaired with VACM seeding with VSMC,Results The prepared BACM and VACM were transparent,HE staining and scanning electron microscopy showed the two acellular matrix was both consisted of abundant network of fibers with the regular arrangement,without cellular debris.Primary culture of VSMC was successfully established and passaged,and was uniformly spindle-shaped in the confluent state and showed a characteristic hill and valley'formation.Immunohistochemical staining showed VSMC in culture stained positively with a-smooth muscle actin antibodies.VSMC began to adhere to the BACM 5 hour after implanting and the number of the adhered cells increased with time.Cells gradually expanded and showed the typical morphology of smooth muscle cells.Three weeks after implantation in vivo,the luminal surface of matrix was completely covered by vaginal epithelial tissue,a layer of smooth muscle cells was formed in the outer surface of the matrix.Multilayered vaginal epithelial and improved development of organized muscle bundles was observed after 6 weeks.The regenerative tissue was equivalent to the normal vaginal tissue at 12 weeks postoperatively.Vaginography demonstrated the maintenance of full patency and a wide vaginal caliber without fibrosis and graft rejection.There was no significant difference in all evaluated items between experimental and control groups.Conclusions BACM has the same regenerative process as VACM in the replacement of vaginal defect.Moreover,BACM has wider source and appears to be an suitable material for vaginal replacement. Key words: Urinary bladder; Extracellular matrix; Vagina/cytology; Myocytes, smooth muscle; Vagina/abnormalities; Rabbits