MP81-04 3D RENAL TUBULAR ORGANOIDS GENERATED FROM URINE-DERIVED STEM CELLS

Abstract

You have accessJournal of UrologyStem Cell Research: Stem Cell Research II1 Apr 2018MP81-04 3D RENAL TUBULAR ORGANOIDS GENERATED FROM URINE-DERIVED STEM CELLS Fangpong Shu, Haibin Guo, Xiong Geng, Anthony Atala, and Yuanyuan Zhang Fangpong ShuFangpong Shu More articles by this author , Haibin GuoHaibin Guo More articles by this author , Xiong GengXiong Geng More articles by this author , Anthony AtalaAnthony Atala More articles by this author , and Yuanyuan ZhangYuanyuan Zhang More articles by this author View All Author Informationhttps://doi.org/10.1016/j.juro.2018.02.2712AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookTwitterLinked InEmail INTRODUCTION AND OBJECTIVES In vitro 3D organoids provide an alternative model for studying of renal toxicology. However, to obtain human renal tubular epithelial cells for these propose, a kidney biopsy is required, with potential risks and complications due to this invasive procedure. Development of in vitro renal tissue model has been limited due to the challenges of differentiating stem cells into various functional renal cells. Our previous studies have demonstrated that human urine-derived stem cells (USCs) originate from glomerular parietal stem cells, possess self-renewal and multipotent capacity2. The goal of this study is to induce human USCs to differentiate into renal tubule epithelial cells when grown on kidney extracellular matrix (k-ECM) to generate in vitro 3D renal tubular organoids for potential use as biological models for nephrotoxic drug development and for renal repair. METHODS USCs obtained from 4 healthy individuals were isolated and expanded in vitro. Normal human renal cells were used as control. Porcine-derived k-ECM hydrogen was produced (k-ECM gel) when mixed with hydraulic acid gel (1:9). To generate the renal tubular organoids, USCs were induced within 1% k-ECM gel for one week, compared to USC aggregates without k-ECM gel. The organoids of USCs+k-ECM gel were cultured in Hanging Drop 96-well plates for 4 days and then transferred to ultra-low attachment plates. To optimize the protocols and measure cell viability, different cell concentrations were assessed by ATPase and LIVE/DEAD Kit. The size of organoids and the renal tubule structures were assessed by histology, immunofluorescence staining and Western blot using renal tubular epithelial cell markers. Renal tubular cell function was measured by ?-Glutamyltransferase (GGT) activity after Ethanol as nephrotoxic agent was added into the culture medium. RESULTS 3D organoids (about 350μm at diameter) were generated by 4,000 USCs cultured for 7 days. ATPase and LIVE/DEAD assays showed that majority of USCs (>95%) remained viable up to 4 weeks. H.E. staining demonstrated the renal tubular-like structures formed within the USCs+k-ECM gel, but no within the USC aggregates without k-ECM gel. Induced USCs expressed proximal renal tubule epithelial cells marker (Aquaporin 1, i.e. AQP1) within whole mount staining. In addition, induced USCs expressing GGT was similar to normal renal cells after ethanol treatment. CONCLUSIONS Human USCs induced by k-ECM formed renal tubular organoids, which could provide an alternative tool for nephrotoxic drug screening, renal disease research and kidney tissue repair. © 2018FiguresReferencesRelatedDetails Volume 199Issue 4SApril 2018Page: e1098 Advertisement Copyright & Permissions© 2018MetricsAuthor Information Fangpong Shu More articles by this author Haibin Guo More articles by this author Xiong Geng More articles by this author Anthony Atala More articles by this author Yuanyuan Zhang More articles by this author Expand All Advertisement Advertisement PDF downloadLoading ...