197 EVALUATION OF EMBRYONIC AND INDUCED PLURIPOTENT STEM CELLS FOR BLADDER TISSUE ENGINEERING STRATEGIES

You have accessJournal of UrologyStem Cell Research1 Apr 2012197 EVALUATION OF EMBRYONIC AND INDUCED PLURIPOTENT STEM CELLS FOR BLADDER TISSUE ENGINEERING STRATEGIES Abhishek Seth, Debra Franck, Eun Seok Gil, David l. Kaplan, Carlos R. Estrada, and Joshua R. Mauney Abhishek SethAbhishek Seth Boston, MA More articles by this author , Debra FranckDebra Franck Boston, MA More articles by this author , Eun Seok GilEun Seok Gil Medford, MA More articles by this author , David l. KaplanDavid l. Kaplan Medford, MA More articles by this author , Carlos R. EstradaCarlos R. Estrada Boston, MA More articles by this author , and Joshua R. MauneyJoshua R. Mauney Boston, MA More articles by this author View All Author Informationhttps://doi.org/10.1016/j.juro.2012.02.250AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookTwitterLinked InEmail INTRODUCTION AND OBJECTIVES Autologous primary smooth muscle and urothelial cells employed in current methods for bladder regeneration are often derived from diseased tissues and therefore their use in defect repair is suboptimal due to alterations in normal cellular physiology. Pluripotent populations such as embryonic stem cells (ESCs) and induced pluripotent stem (iPS) cells may fulfill the need for alternative cell sources capable of generating organ and patient-specific cells for bladder tissue engineering. In this study, we assessed the ability of ESC and iPS cells to undergo smooth muscle and urothelial differentiation on silk-based biomaterials in order to create tissue engineered constructs for potential use in urologic tissue repair. METHODS Murine iPS cell lines (WP5 and T1b, Stemgent, Cambridge, MA) and ESCs were seeded independently on fibronectin-coated silk scaffolds and maintained up to 14 d in DMEM/15% fetal calf serum in the presence of all trans retinoic acid (RA, 10μM) or as spontaneously differentiating controls. The ability of silk biomaterials to support cell attachment was determined by alamar blue analysis. The extent of pluripotent cell differentiation was determined by monitoring expression of pluripotency markers, urothelial-specific uroplakins (UP) and cytokeratins, as well as smooth muscle associated contractile proteins including α-actin, γ-actin, SM22α, and smooth muscle myosin heavy chain by real time RT-PCR (qPCR) and immunohistochemical (IHC) analyses. RESULTS Alamar blue analysis revealed that fibronectin coating of silk scaffolds significantly improved levels of initial cell attachment in all pluripotent lines tested. Over the course of the 14 d cultivation period, qPCR analysis demonstrated that iPS cell lines and ESCs in response to RA stimulation elicited significant downregulation of pluripotency factors, Oct4, Rex1, and Nanog, but marked upregulation of all uroplakin and smooth muscle contractile gene mRNA transcript levels tested in comparison to undifferentiated and spontaneously differentiating controls. IHC analyses of RA-treated iPS and ESC cultures revealed smooth muscle and urothelial cell populations within silk scaffolds which displayed SM22α and α-actin protein expression as well as pan-UP and pan-cytokeratin markers. CONCLUSIONS Silk matrices provide an effective substrate for RA-mediated smooth muscle and urothelial differentiation of pluripotent stem cells and therefore represent potential options for urologic tissue repair. © 2012 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 187Issue 4SApril 2012Page: e83 Advertisement Copyright & Permissions© 2012 by American Urological Association Education and Research, Inc.MetricsAuthor Information Abhishek Seth Boston, MA More articles by this author Debra Franck Boston, MA More articles by this author Eun Seok Gil Medford, MA More articles by this author David l. Kaplan Medford, MA More articles by this author Carlos R. Estrada Boston, MA More articles by this author Joshua R. Mauney Boston, MA More articles by this author Expand All Advertisement Advertisement PDF downloadLoading ...