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Am J Physiol Renal Physiol 283: F963-F970, 2002. First published July 16, 2002; doi:10.1152/ajprenal.00108.2002
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Vol. 283, Issue 5, F963-F970, November 2002

Calcium-sensing receptor-mediated TNF production in medullary thick ascending limb cells

Dairong Wang, Paulina L. Pedraza, Huda Ismail Abdullah, John C. McGiff, and Nicholas R. Ferreri

Department of Pharmacology, New York Medical College, Valhalla, New York 10595

Medullary thick ascending limb (mTAL) cells in primary culture express the Ca2+-sensing receptor (CaR), a G protein-coupled receptor that senses changes in extracellular Ca2+ (Ca<UP><SUB>o</SUB><SUP>2+</SUP></UP>) concentration, resulting in increases of intracellular Ca2+ concentration and PKC activity. Exposure of mTAL cells to either Ca<UP><SUB>o</SUB><SUP>2+</SUP></UP> or the CaR-selective agonist poly-L-arginine increased TNF-alpha synthesis. Moreover, the response to Ca<UP><SUB>o</SUB><SUP>2+</SUP></UP> was enhanced in mTAL cells transfected with a CaR overexpression vector. Transfection of mTAL cells with a TNF promoter construct revealed an increase in reporter gene activity after exposure of the cells to Ca<UP><SUB>o</SUB><SUP>2+</SUP></UP>, suggesting that intracellular signaling pathways initiated by means of activation of a CaR contribute to TNF synthesis by a mechanism that involves transcription of the TNF gene. Neutralization of TNF activity with an anti-TNF antibody attenuated Ca2+-mediated increases in cyclooxygenase-2 (COX-2) protein expression and PGE2 synthesis, suggesting that TNF exerts an autocrine effect in the mTAL, which contributes to COX-2-mediated PGE2 production. Preincubation with the PKC inhibitor bisindolylmaleimide I inhibited Ca2+-mediated TNF production. Significant inhibition of COX-2 protein expression and PGE2 synthesis also was observed when cells were challenged with Ca<UP><SUB>o</SUB><SUP>2+</SUP></UP> in the presence of bisindolylmaleimide I. The data suggest that increases in TNF production subsequent to activation of the CaR may be the basis of an important renal mechanism that regulates salt and water excretion.

tumor necrosis factor; cyclooxygenase-2; kidney


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