| First Author | Markadieu N | Year | 2012 |
| Journal | Am J Physiol Renal Physiol | Volume | 303 |
| Issue | 6 | Pages | F886-92 |
| PubMed ID | 22759396 | Mgi Jnum | J:188407 |
| Mgi Id | MGI:5440519 | Doi | 10.1152/ajprenal.00114.2012 |
| Citation | Markadieu N, et al. (2012) A primary culture of distal convoluted tubules expressing functional thiazide-sensitive NaCl transport. Am J Physiol Renal Physiol 303(6):F886-92 |
| abstractText | Studying the molecular regulation of the thiazide-sensitive Na(+)-Cl(-) cotransporter (NCC) is important for understanding how the kidney contributes to blood pressure regulation. Until now, a native mammalian cell model to investigate this transporter remained unknown. Our aim here is to establish, for the first time, a primary distal convoluted tubule (DCT) cell culture exhibiting transcellular thiazide-sensitive Na(+) transport. Because parvalbumin (PV) is primarily expressed in the DCT, where it colocalizes with NCC, kidneys from mice expressing enhanced green-fluorescent protein (eGFP) under the PV gene promoter (PV-eGFP-mice) were employed. The Complex Object Parametric Analyzer and Sorter (COPAS) was used to sort fluorescent PV-positive tubules from these kidneys, which were then seeded onto permeable supports. After 6 days, DCT cell monolayers developed transepithelial resistance values of 630 +/- 33 Omega.cm(2). The monolayers also established opposing transcellular concentration gradients of Na(+) and K(+). Radioactive (22)Na(+) flux experiments showed a net apical-to-basolateral thiazide-sensitive Na(+) transport across the monolayers. Both hypotonic low-chloride medium and 1 muM angiotensin II increased this (22)Na(+) transport significantly by four times, which could be totally blocked by 100 muM hydrochlorothiazide. Angiotensin II-stimulated (22)Na(+) transport was also inhibited by 1 muM losartan. Furthermore, NCC present in the DCT monolayers was detected by immunoblot and immunocytochemistry studies. In conclusion, a murine primary DCT culture was established which expresses functional thiazide-sensitive Na(+)-Cl(-) transport. |
