Abstract
Genetic studies on hereditary kidney diseases and in vivo experimental model studies have revealed a critical role for the podocyte in glomerular function and disease. Primary podocyte cultures as well as immortalized podocyte cell lines have been used extensively to study podocyte function. Although, primary cells often more closely resemble the in vivo cells, they may have only a finite replicative life span before they reach senescence. Therefore, the success of studies using primary cell cultures depends on standardized isolation and culture protocols that allow reproducible generation of stable primary cultures.
This chapter describes the isolation of primary podocytes with a proven origin using the novel technology of cell-specific genetic tagging. Podocytes are isolated from glomeruli from a podocyte-specific transgenic reporter mouse. The podocyte-specific reporter gene beta-galactosidase is used to identify and specifically isolate the labeled podocytes from other glomerular cells by FACS.
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Acknowledgments
This work was supported by the German Research Foundation (Grant BO 3755/1-1 to BS), by the Genzyme Renal Innovation Program (GRIP, to BS). Additional support came from the eRARE consortium “Rare-G” (01 GM 1208A to MJM) and TP17 of SFB/Transregio 57 of the German Research Foundation (to MJM). MJM is a member of the SFB/Transregio 57 DFG consortium “mechanisms of organ fibrosis .” We thank Regina Lanzmich for excellent technical support and helpful discussions, and we thank the Q1 platform of the SFB/Transregio 57 for implementing the above-described protocol into routine practice.
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Smeets, B., Kabgani, N., Moeller, M.J. (2016). Isolation and Primary Culture of Murine Podocytes with Proven Origin. In: Hewitson, T., Smith, E., Holt, S. (eds) Kidney Research. Methods in Molecular Biology, vol 1397. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-3353-2_1
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DOI: https://doi.org/10.1007/978-1-4939-3353-2_1
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Publisher Name: Humana Press, New York, NY
Print ISBN: 978-1-4939-3351-8
Online ISBN: 978-1-4939-3353-2
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