Abstract
Primary rabbit kidney epithelial cell cultures can be obtained that express renal proximal tubule functions. Toward these ends, renal proximal tubules are purified from the rabbit kidney by the method of Brendel and Meezan. To summarize, each kidney is perfused with iron oxide, which becomes associated with glomeruli. The renal cortex is sliced and homogenized to liberate nephron segments. Renal proximal tubules and glomeruli are purified by sieving. The glomeruli, covered with iron oxide, are removed using a magnet. After a brief collagenase treatment (to disrupt basement membrane), the tubules are plated in hormonally defined serum-free medium supplemented with 5 µg/mL bovine insulin, 5 µg/mL human transferrin, and 5 × 10−8 M hydrocortisone. After 5–6 d of incubation, confluent monolayers are obtained that possess multicellular domes, indicative of their capacity for transepithelial solute transport.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
References
Taub, M., Chuman, L., Saier, M. H., Jr., and Sato, G. (1979) Growth of Madin Darby Canine Kidney epithelial cell (MDCK) line in hormone-supplemented serum-free medium. Proc. Natl. Acad. Sci. USA 76, 3338–3342.
Chuman, L., Fine, L. G., Cohen, A. I., and Saier, M. H., Jr. (1982) Continuous growth of proximal tubular epithelial cells in hormone-supplemented serum-free medium. J. Cell Biol. 94, 506–510.
Taub, M. and Sato, G. (1979) Growth of functional primary cultures of kidney epithelial cells in defined medium. J. Cell. Physiol. 105, 369–378.
Chung, S. D., Alavi, N., Livingston, D., Hiller, S., and Taub, M. (1982) Characterization of primary rabbit kidney cultures that express proximal tubule functions in a hormonally defined medium. J. Cell Biol. 95, 118–126.
Taub, M. (1985) Primary culture of proximal tubule cells in defined medium. J. Tissue Culture Methods 9, 67–72.
Waqar, M. A., Seto, J., Chung, S. D., Hiller-Grohol, S., and Taub, M. (1985) Phosphate uptake by primary renal proximal tubule cells grown in hormonally defined medium. J. Cell. Physiol. 124,411–423.
Brendel, K. and Meezan, E. (1975) Isolation and properties of a pure preparation of proximal kidney tubules obtained without collagenase treatment. Fed. Proc. 34, 803.
Meezan, E. K., Brendel, J., Ulreich, J., and Carlson, E. C. (1973) Properties of a pure metabolically active glomerular preparation from rat kidneys. I. Isolation. J. Pharm. Exp. Ther. 187, 332–341.
Taub, M., Axelson, E., and Park, J. H. (1998) Improved method for primary cell cultures: use of tissue culture dishes with a colloidal silica surface. Biotechniques 25, 990–994.
Sakhrani, L. M., Badie-Dezfooly, B., Trizna, W., et al. (1984) Transport and metabolism of glucose by renal proximal tubular cells in primary culture. Am. J. Physiol. 246, F757–F764.
Matsuo, S., Fukatsu, A., Taub, M. L., Caldwell, P. R. B., Brentjens, J. R., and Andres, G. (1987) Nephrotoxic glomerulonephritis induced in the rabbit by antiendothelial antibodies. J. Clin. Invest. 79, 1798–1811.
Yang, I. S., Goldinger, J. M., Hong, S. K., and Taub, M. (1987) The preparation of basolateral membranes that transport p-aminohippurate from primary cultures of rabbit kidney proximal tubule cells. J. Cell. Physiol. 135,481–487.
Fine, L. G. and Sakhrani, L. M. (1986) Proximal tubular cells in primary culture. Miner. Electrolyte Metab. 12, 51–57.
Park, S. H., Taub, M., and Han, H. J. (2001) Regulation of phosphate uptake in primary cultured rabbit renal proximal tubule cells by glucocorticoids: evidence for nongenomic as well as genomic mechanisms. Endocrinology 142, 710–720.
Han, H. J., Lee, Y. H., Park, S. H., and Taub, M. (2002) Estradiol-17β stimulates phosphate uptake and is mitogenic for primary rabbit renal proximal tubule cells. Exp. Nephrol. 10, 355–364.
Han, H. J., Park, S. H., Koh, H. J., and Taub, M. (2000) Mechanism of regulation of Na+ transport by angiotensin II in primary renal cells. Kidney Int. 57, 2457–2467.
Aleo, M. D., Taub, M. L., Olson, J. R., and Kostyniak, P. J. (1990) Primary cultures of rabbit renal proximal tubule cells: II. Selected phase I and phase II metabolic capacities. Toxicol. In Vitro 4, 727–733.
Aleo, M. D., Taub, M. L., and Kostyniak, P. J. (1992) Primary cultures of rabbit renal proximal tubule cells: III. Comparative cytotoxicity of inorganic and organic mercury. J. Toxicol. Appl. Pharm. 112, 310–317.
Aleo, M. D., Taub, M. L., Olson, J. R., Nickerson, P. A., and Kostyniak, P. J. (1987) Primary cultures of rabbit renal proximal tubule cells as an in vitro model of nephrotoxicity: effects of 2 mercurials, in In vitro Toxicology: Approaches to Validation (Goldberg, A. M., ed.), Mary Ann Liebert, Inc., New York, Vol. 5, pp. 211–225.
Wang, Y. and Taub, M. (1991) Insulin and other regulatory factors modulate the growth and the phosphoenolpyruvate carboxykinase (PEPCK) activity of primary rabbit kidney proximal tubule cells in serum free medium. J. Cell. Physiol. 147, 374–382.
Jung, J. C., Lee, S. M., Kadakia, N., and Taub, M. (1992) Growth and function of primary rabbit kidney proximal tubule cells in glucose-free serum-free medium. J. Cell. Physiol. 150, 243–250.
Han, H. J., Jung, J. C., and Taub, M. (1999) Response of primary rabbit kidney proximal tubule cells to estrogens. J. Cell. Physiol. 178, 35–43.
Springate, J., Davies, S., Chen, K., and Taub, M. (1999) Toxicity of ifosfamide and its metabolite chloroacetaldehyde in cultured renal tubule cells. In Vitro Cell Dev. Biol. 35, 314–317.
Zaki, E. L., Springate, J. E., and Taub, M. (2003) Comparative toxicity of ifosfamide metabolites and protective effect of mesna and amifostine in cultured renal tubule cells. Toxicol. In Vitro 17, 397–402.
Cook, W. F. and Pickering, G. W. (1958) A rapid method for separating glomeruli from rabbit kidney. Nature 182, 1103–1104.
Bradford, M. M. (1976) A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal. Biochem. 72, 248–254.
Author information
Authors and Affiliations
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2005 Humana Press Inc., Totowa, NJ
About this protocol
Cite this protocol
Taub, M. (2005). Primary Kidney Proximal Tubule Cells. In: Helgason, C.D., Miller, C.L. (eds) Basic Cell Culture Protocols. Methods in Molecular Biology™, vol 290. Humana Press. https://doi.org/10.1385/1-59259-838-2:231
Download citation
DOI: https://doi.org/10.1385/1-59259-838-2:231
Publisher Name: Humana Press
Print ISBN: 978-1-58829-284-1
Online ISBN: 978-1-59259-838-0
eBook Packages: Springer Protocols