Reconstitution of a Regulated Transepithelial Water Pathway in Cells Transfected with AQP2 and an AQP1/AQP2 Hybrid Containing the AQP2-C Terminus
| Title: | Reconstitution of a Regulated Transepithelial Water Pathway in Cells Transfected with AQP2 and an AQP1/AQP2 Hybrid Containing the AQP2-C Terminus |
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| Authors: | Toriano, R.; Ford, P.; Rivarola, V.; Tamarappoo, B.K.; Verkman, Alan S.; Parisi, Valerie M. |
| Publisher: | Journal of Membrane Biology |
| Date Published: | January 01, 1998 |
| Reference Number: | 189 |
Transepithelial water permeability was measured in LLC-PK1 cells stably transfected with aquaporins (AQPs): AQP1, AQP2, and a chimera of AQP1 and AQP2 containing 41 amino acids of the C-terminus of AQP2. Transepithelial water fluxes (Jw) were not previously reported in cells transfected with aquaporins. Jw were now recorded each minute using a specially developed experimental device. A significant increase in Posm after forskolin (FK) plus vasopressin (VP) was found in AQP2 transfected cells (39.9 +/- 8.2 vs. 12.5 +/- 3.3 cm.sec-1.10(-3)), but not in cells transfected with AQP1 (15.3 +/- 3.6 vs. 13.4 +/- 3.6 cm.sec-1.10(-3)). In the case of the AQP1/2 cells (chimera) the FK plus VP-induced Posm was smaller than in AQP2 cells but significantly higher than in mock cells at rest (18.1 +/- 4.8 vs. 6.7 +/- 1.0 cm.sec-1.10(-3)). The increases in Posm values were not paralleled by increases in 14C-Mannitol permeability. HgCl2 inhibited the hydrosmotic response to FK plus VP in AQP2 transfected epithelia. Results were comparable to those observed, in parallel experiments, in a native ADH-sensitive water channel containing epithelial barrier (the toad urinary bladder). Electron microscopy showed confluent LLC-PK1 cells with microvilli at the mucosal border. The presence of spherical or elongated intracellular vacuoles was observed in AQP2 transfected cells, specially after FK plus VP stimulus and under an osmotic gradient. These results demonstrate regulated transepithelial water permeability in epithelial cells transfected with AQP2.
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This translation by the NDI Foundation is to assist the lay reader. To provide a clear, accessible interpretation of the original article, we eliminated or simplified some technical detail and complicated scientific language. We concentrated our translation on those aspects of the article dealing directly with NDI. The NDI Foundation thanks the researchers for their work toward understanding and more effectively treating this disorder.
© Copyright NDI Foundation 2007 (JC)
Toriano, et al., used a newly developed experimental device that could measure water flow across cell membranes on a minute-to-minute basis to determine the effect on the osmotic permeability (and hence the water flow) of cells in laboratory cell cultures under specific conditions. They used some cell cultures transfected with aquaporin-1 (AQP1), some transfected with AQP2, and some with an aquaporin they designed to be part AQP1 and part AQP2 (AQP1/2). One set of cells was not transfected with anything and these served as a control group.
The authors injected one set of all four cell cultures with the antidiuretic hormone, arginine vasopressin (AVP), another set with forskolin (FK), and another set with AVP + FK. They recorded a significant increase in osmotic permeability in the cells with AQP2 when they injected them with AVP + FK, but no such increase in cells with AQP1. In the case of cells with AQP1/2, they found an osmotic permeability increase less than in the AQP2 set, but greater than in the control set.
The authors' article presents the first measurement of water permeability across cell membranes in aquaporin transfected cells. The results demonstrate that an increase in water permeability occurred in the membranes of cell cultures after transfection with AQP2 and, to a lesser extent, with AQP1/2.