| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH |
|
|
|
|||||||
|
|||||||||
| ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
1 Physiology, Emory University School of Medicine, Atlanta, GA, USA
2 Medicine/Renal, Emory University School of Medicine, Atlanta, GA, USA
3 Physiology, Emory University School of Medicine, Atlanta, GA, USA; Medicine/Renal, Emory University School of Medicine, Atlanta, GA, USA
* To whom correspondence should be addressed. E-mail: rbgunn{at}emory.edu.
Progress in understanding the cell biology of urea transporter proteins has been hampered by the lack of an appropriate cell culture system. The goal of this study was to create a polarized epithelial cell line that stably expresses the largest of the rat renal urea transporter UT-A isoforms, UT-A1. The gene for UT-A1 was cloned into pcDNA5/FRT and transfected into MDCK cells with an integrated Flp recombination target site. The cells from a single clone were grown to confluence on collagen-coated membranes until the resistance was greater than 1,500 Ohm-cm2. Transepithelial 14C-urea fluxes were measured at 37°C in a bicarbonate/CO2 buffer, pH 7.4, with 5 mM urea. The baseline fluxes were not different between unstimulated UT-A1 transfected MDCK cells and non-transfected or sham-transfected MDCK cells. However, only in the UT-A1 transfected cells was UT-A1 protein expressed (as measured by western analysis) and urea transport stimulated by forskolin or vasopressin. Forskolin and vasopressin also increased the phosphorylation of UT-A1. Thionicotinamide, dimethylurea, and phloretin inhibited the forskolin-stimulated 14C-urea fluxes in the UT-A1 transfected MDCK cells. These characteristics mimic those seen in rat terminal inner medullary collecting ducts. This new polarized epithelial cell line stably expresses UT-A1 and reproduces several of the physiologic responses observed in rat terminal inner medullary collecting ducts.
This article has been cited by other articles:
|
|
 |
|
  Y. Wang, J. D. Klein, M. A. Blount, C. F. Martin, K. J. Kent, V. Pech, S. M. Wall, and J. M. Sands Epac Regulates UT-A1 to Increase Urea Transport in Inner Medullary Collecting Ducts J. Am. Soc. Nephrol., September 1, 2009; 20(9): 2018 - 2024. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 P. Tickle, A. Thistlethwaite, C. P. Smith, and G. S. Stewart Novel bUT-B2 urea transporter isoform is constitutively activated Am J Physiol Regulatory Integrative Comp Physiol, August 1, 2009; 297(2): R323 - R329. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 G. S. Stewart, A. Thistlethwaite, H. Lees, G. J. Cooper, and C. Smith Vasopressin regulation of the renal UT-A3 urea transporter Am J Physiol Renal Physiol, March 1, 2009; 296(3): F642 - F648. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
G. Chen, H. Huang, O. Frohlich, Y. Yang, J. D. Klein, S. R. Price, and J. M. Sands MDM2 E3 ubiquitin ligase mediates UT-A1 urea transporter ubiquitination and degradation Am J Physiol Renal Physiol, November 1, 2008; 295(5): F1528 - F1534. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 G. S. Stewart, J. H. O'Brien, and C. P. Smith Ubiquitination regulates the plasma membrane expression of renal UT-A urea transporters Am J Physiol Cell Physiol, July 1, 2008; 295(1): C121 - C129. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
B. MacIver, C. P. Smith, W. G. Hill, and M. L. Zeidel Functional characterization of mouse urea transporters UT-A2 and UT-A3 expressed in purified Xenopus laevis oocyte plasma membranes Am J Physiol Renal Physiol, April 1, 2008; 294(4): F956 - F964. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
O. Frohlich, D. Aggarwal, J. D. Klein, K. J. Kent, Y. Yang, R. B. Gunn, and J. M. Sands Stimulation of UT-A1-mediated transepithelial urea flux in MDCK cells by lithium Am J Physiol Renal Physiol, March 1, 2008; 294(3): F518 - F524. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. A. Blount, J. D. Klein, C. F. Martin, D. Tchapyjnikov, and J. M. Sands Forskolin stimulates phosphorylation and membrane accumulation of UT-A3 Am J Physiol Renal Physiol, October 1, 2007; 293(4): F1308 - F1313. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
G. S. Stewart, S. L. King, E. A. Potter, and C. P. Smith Acute regulation of mUT-A3 urea transporter expressed in a MDCK cell line Am J Physiol Renal Physiol, April 1, 2007; 292(4): F1157 - F1163. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
J. D. Klein, B. P. Murrell, S. Tucker, Y.-H. Kim, and J. M. Sands Urea transporter UT-A1 and aquaporin-2 proteins decrease in response to angiotensin II or norepinephrine-induced acute hypertension Am J Physiol Renal Physiol, November 1, 2006; 291(5): F952 - F959. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
O. Frohlich, J. D. Klein, P. M. Smith, J. M. Sands, and R. B. Gunn Regulation of UT-A1-mediated transepithelial urea flux in MDCK cells Am J Physiol Cell Physiol, October 1, 2006; 291(4): C600 - C606. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
J. D. Klein, O. Frohlich, M. A. Blount, C. F. Martin, T. D. Smith, and J. M. Sands Vasopressin Increases Plasma Membrane Accumulation of Urea Transporter UT-A1 in Rat Inner Medullary Collecting Ducts J. Am. Soc. Nephrol., October 1, 2006; 17(10): 2680 - 2686. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
E. A. Potter, G. Stewart, and C. P. Smith Urea flux across MDCK-mUT-A2 monolayers is acutely sensitive to AVP, cAMP, and [Ca2+]i Am J Physiol Renal Physiol, July 1, 2006; 291(1): F122 - F128. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
H. Inoue, S. D. Kozlowski, J. D. Klein, J. L. Bailey, J. M. Sands, and S. M. Bagnasco Regulated expression of renal and intestinal UT-B urea transporter in response to varying urea load Am J Physiol Renal Physiol, August 1, 2005; 289(2): F451 - F458. [Abstract] [Full Text] [PDF]
|
|
| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH |
| Visit Other APS Journals Online |
