Aleksandra Rak-Raszewska1, Ganna Reint1, Fabienne Geiger1, Florence Naillat1, Seppo J Vainio1,2. 1. Laboratory of Developmental Biology, Disease Networks Researtch Unit, Faculty of Biochemistry and Molecular Medicine, University of Oulu, Oulu, Finland. 2. Kvantum Institute, Infotech Oulu, University of Oulu, Oulu, Finland.
Abstract
BACKGROUND: Tissue organoids derived from primary cells have high potential for studying organ development and diseases in numerous organs. They recreate the morphological structure and mimic the functions of given organ while being compact in size, easy to produce, and suitable for use in various experimental setups. RESULTS: In this study we established the number of cells that form mouse kidney rudiments at E11.5, and generated renal organoids of various sizes from the mouse primary cells of the metanephric mesenchyme (MM). We investigated the ability of renal organoids to undergo nephrogenesis upon Wnt/ β-catenin pathway-mediated tubule induction with a GSK-3 inhibitor (BIO) or by initiation through the ureteric bud (UB). We found that 5000 cells of MM cells are necessary to successfully form renal organoids with well-structured nephrons as judged by fluorescent microscopy, transmission electron microscopy (TEM), and quantitative Polymerase Chain Reaction (qPCR). These mouse organoids also recapitulated renal secretion function in the proximal tubules. CONCLUSIONS: We show that a significant decrease of cells used to generate renal mouse organoids in a dissociation/re-aggregation assay, does not interfere with development, and goes toward 3Rs. This enables generation of more experimental samples with one mouse litter, limiting the number of animals used for studies.
BACKGROUND: Tissue organoids derived from primary cells have high potential for studying organ development and diseases in numerous organs. They recreate the morphological structure and mimic the functions of given organ while being compact in size, easy to produce, and suitable for use in various experimental setups. RESULTS: In this study we established the number of cells that form mouse kidney rudiments at E11.5, and generated renal organoids of various sizes from the mouse primary cells of the metanephric mesenchyme (MM). We investigated the ability of renal organoids to undergo nephrogenesis upon Wnt/ β-catenin pathway-mediated tubule induction with a GSK-3 inhibitor (BIO) or by initiation through the ureteric bud (UB). We found that 5000 cells of MM cells are necessary to successfully form renal organoids with well-structured nephrons as judged by fluorescent microscopy, transmission electron microscopy (TEM), and quantitative Polymerase Chain Reaction (qPCR). These mouse organoids also recapitulated renal secretion function in the proximal tubules. CONCLUSIONS: We show that a significant decrease of cells used to generate renal mouse organoids in a dissociation/re-aggregation assay, does not interfere with development, and goes toward 3Rs. This enables generation of more experimental samples with one mouse litter, limiting the number of animals used for studies.
Authors: Anwar A Palakkan; Julia Tarnick; Martin Waterfall; May Sallam; Fokion Glykofrydis; Mona Elhendawi; Jamie A Davies Journal: Sci Rep Date: 2022-07-22 Impact factor: 4.996