Article: Characterization of a murine renal distal convoluted tubule cell line for the study of transcellular calcium transport.
American journal of physiology. Renal physiology
2004 Volume 286, Issue 3, Page(s) F483–9
Abstract: To unravel the molecular regulation of renal transcellular Ca(2+) transport, a murine distal convoluted tubule (mpkDCT) cell line derived from distal convoluted tubules (DCT) microdissected from a SV-PK/Tag transgenic mouse was characterized. This cell ... ...
| Abstract | To unravel the molecular regulation of renal transcellular Ca(2+) transport, a murine distal convoluted tubule (mpkDCT) cell line derived from distal convoluted tubules (DCT) microdissected from a SV-PK/Tag transgenic mouse was characterized. This cell line originated from DCT only, as mRNA encoding for the DCT marker thiazide-sensitive Na(+)/Cl(-) cotransporter was expressed, whereas mRNA encoding for the connecting tubule and collecting duct marker aquaporin-2 was not detected, as determined by reverse-transcriptase PCR. mpkDCT cells expressed mRNA encoding the Ca(2+) channels TRPV5 and TRPV6 and other key players necessary for transcellular Ca(2+) transport, i.e., calbindin-D(9k), calbindin-D(28k), plasma membrane Ca(2+)-ATPase isoform 1b, and Na(+)/Ca(2+) exchanger 1. Primary cultures of DCT cells exhibited net transcellular Ca(2+) transport of 0.4 +/- 0.1 nmol.h(-1).cm(-2), whereas net transcellular Ca(2+) transport across mpkDCT cells was significantly higher at 2.4 +/- 0.4 nmol.h(-1).cm(-2). Transcellular Ca(2+) transport across mpkDCT cells was completely inhibited by ruthenium red, an inhibitor of TRPV5 and TRPV6, but not by the voltage-operated Ca(2+) channel inhibitors felodipine and verapamil. With the use of patch-clamp analysis, the IC(50) of ruthenium red on Na(+) currents was between the values measured for TRPV5- and TRPV6-expressing HEK 293 cells, suggesting that TRPV5 and/or TRPV6 is possibly active in mpkDCT cells. Forskolin in combination with IBMX, 1,25-dihydroxyvitamin D(3), and 1-deamino-8-d-arginine vasopressin increased transcellular Ca(2+) transport, whereas PMA and parathyroid hormone had no significant effect. In conclusion, the murine mpkDCT cell line provides a unique cell model in which to study the molecular regulation of transcellular Ca(2+) transport in the kidney in vitro. |
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| MeSH term(s) | Animals ; Calcitriol/pharmacology ; Calcium/metabolism ; Cell Line ; Cyclic AMP/metabolism ; Electric Conductivity ; Female ; Ion Transport ; Kidney Tubules, Distal/cytology ; Kidney Tubules, Distal/drug effects ; Kidney Tubules, Distal/metabolism ; Mice ; Mice, Inbred C57BL ; RNA, Messenger/metabolism ; Ruthenium Red/pharmacology | |||||
| Chemical Substances | RNA, Messenger ; Ruthenium Red (11103-72-3) ; Cyclic AMP (E0399OZS9N) ; Calcitriol (FXC9231JVH) ; Calcium (SY7Q814VUP) | |||||
| Language | English | |||||
| Publishing date | 2004-03 | |||||
| Publishing country | United States | |||||
| Document type | Journal Article ; Research Support, Non-U.S. Gov't | |||||
| ZDB-ID | 603837-2 | |||||
| ISSN | 1522-1466 ; 1931-857X ; 0363-6127 | |||||
| ISSN (online) | 1522-1466 | |||||
| ISSN | 1931-857X ; 0363-6127 | |||||
| DOI | 10.1152/ajprenal.00231.2003 | |||||
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| Database | MEDical Literature Analysis and Retrieval System OnLINE |
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