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Originally published In Press as doi:10.1074/jbc.M006218200 on September 11, 2000

J. Biol. Chem., Vol. 275, Issue 48, 37922-37929, December 1, 2000
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Low Chloride Stimulation of Prostaglandin E2 Release and Cyclooxygenase-2 Expression in a Mouse Macula Densa Cell Line*

Tianxin YangDagger , John M. Park§, Lois Arend, Yuning HuangDagger , Rezan TopalogluDagger , Anita PasumarthyDagger , Helle Praetorius||, Kenneth Spring||, Josephine P. BriggsDagger , and Jurgen SchnermannDagger **

From the Dagger  NIDDK and || NHLBI, National Institutes of Health, Bethesda, Maryland 20892, the § Department of Urology, University of Michigan, Ann Arbor, Michigan 48109, and the  Department of Pathology and Laboratory Medicine, University of Rochester, Rochester, New York 14642

Reducing luminal NaCl concentration in the macula densa region of the nephron stimulates renin secretion, and this response is blocked by a specific inhibitor of cyclooxygenase-2 (COX-2) (Traynor, T. R., Smart, A., Briggs, J. P., and Schnermann, J. (1999) Am. J. Physiol. Renal Physiol. 277, F706-710). To study whether low NaCl activates COX-2 activity or expression we clonally derived a macula densa cell line (MMDD1 cells) from SV-40 transgenic mice using fluorescence-activated cell sorting of renal tubular cells labeled with segment-specific fluorescent lectins. MMDD1 cells express COX-2, bNOS, NKCC2, and ROMK, but not Tamm-Horsfall protein, and showed rapid 86Rb+ uptake that was inhibited by a reduction in NaCl concentration and by bumetanide or furosemide. Isosmotic exposure of MMDD1 cells to low NaCl (60 mM) caused a prompt and time-dependent stimulation of prostaglandin E2 (PGE2) release that was prevented by the COX-2 specific inhibitor NS-398 (10 µM). Reducing NaCl to 60 and 6 mM for 16 h increased COX-2 expression in a chloride-dependent fashion. Low NaCl phosphorylated p38 kinase within 30 min and ERK1/2 kinases within 15 min without changing total MAP kinase levels. Low NaCl-stimulated PGE2 release and COX-2 expression was inhibited by SB 203580 and PD 98059 (10 µM), inhibitors of p38 and ERK kinase pathways. We conclude that low chloride stimulates PGE2 release and COX-2 expression in MMDD1 cells through activation of MAP kinases.

* This work was supported by NIDDK, National Institutes of Health Grants DK-37448, DK-39255, and DK-40042 and by intramural funds from NIDDK and NHLBI.The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

** To whom correspondence should be addressed: NIDDK, National Institutes of Health, Bldg. 10, Rm. 4D51, 10 Center Dr., MSC 1370, Bethesda, MD 20892. Tel.: 301-435-6580; Fax: 301-435-6587; E-mail: jurgens@intra.niddk.nih.gov.

Copyright © 2000 by The American Society for Biochemistry and Molecular Biology, Inc.
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