You have accessJournal of UrologyBladder and Urethra: Anatomy, Physiology and Pharmacology1 Apr 2011427 COMPARING DIFFERENT DECELLULARIZATION PROTOCOLS OF ANIMALS' BLADDERS FOR MESENCHYMAL STEM CELL-BASED TISSUE ENGINEERING Wally Mahfouz, Oleg Loutochin, Daniel L. Coutu, Jacques Galipeau, and Jacques Corcos Wally MahfouzWally Mahfouz Montreal, Canada More articles by this author , Oleg LoutochinOleg Loutochin Montreal, Canada More articles by this author , Daniel L. CoutuDaniel L. Coutu Montreal, Canada More articles by this author , Jacques GalipeauJacques Galipeau Montreal, Canada More articles by this author , and Jacques CorcosJacques Corcos Montreal, Canada More articles by this author View All Author Informationhttps://doi.org/10.1016/j.juro.2011.02.517AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookTwitterLinked InEmail INTRODUCTION AND OBJECTIVES Artificial bladder tissue is a constant need in reconstructive surgery. Intestine can be used but it has several deleterious complications. We are aiming to produce xenogenic decellularized bladder seeded with autologous mesenchymal stem cells for bladder replacement/augmentation in human. We are reporting the first stage of this work which is decellularization protocol for efficient removal of cells and nuclear debris in three species. We also demonstrate the adherence and differentiation of mesenchymal stem cells on the decellularized scaffolds. METHODS This study compared four decellularization protocols for bladders of three species: rat, rabbit and porcine. We first determined which detergent (1% SDS or 1% Triton X-100 in hypotonic Tris-HCl) was more efficient at removing cytoplasmic debris while preserving the structural anatomy. This was done by H&E staining of paraffin sections. We also determined the optimal duration of decellularization. After extensive washes, we verified whether deoxyribonuclease (DNase) digestion of nuclear debris was necessary and adequate using 4',6-diamidino-2-phenylindole (DAPI) staining. We then analyzed the resultant decellularized extracellular matrices for evidence of preserved active growth factors (VEGF, TGFβ1, EGF, TGFα) and matrix proteins collagen (type 1, 2, 3, 4), laminin, and elastin by histology, immuno-fluorescence staining and confocal microscopy. We then tested the ability of mesenchymal stem cells to adhere and differentiate into smooth muscle cells on these scaffolds. RESULTS Both detergents were equally efficient at removing cytoplasmic debris. The duration of detergent treatment proved to be critical here. Triton X-100 appears to preserve the extracellular matrix better. DNase digestion was always necessary for complete removal of nuclear debris. Mesenchymal stem cells adhered well on the scaffolds. CONCLUSIONS The use of Triton X-100 in hypotonic buffer followed by nuclease digestion is efficient for production of decellularized bladder tissue. This method preserves the structural anatomy, extracellular matrix proteins, and growth factors within bladder tissue. Moreover, mesenchymal stem cells adhere well on these scaffolds and remain viable after 2 weeks in vitro. © 2011 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 185Issue 4SApril 2011Page: e172 Advertisement Copyright & Permissions© 2011 by American Urological Association Education and Research, Inc.MetricsAuthor Information Wally Mahfouz Montreal, Canada More articles by this author Oleg Loutochin Montreal, Canada More articles by this author Daniel L. Coutu Montreal, Canada More articles by this author Jacques Galipeau Montreal, Canada More articles by this author Jacques Corcos Montreal, Canada More articles by this author Expand All Advertisement Advertisement PDF downloadLoading ...