Regulatory mechanisms of cyclooxygenase-2 gene expression in cultured mouse macula densa cells

Abstract

The kidney macula densa (MD) control the tubuloglomerular feedback (TGF) system for regulation of the total body fluid volume. The MD cells express cyclooxygenase-2 (COX-2) and COX-2-derived prostaglandins stimulate the renin release from the granular cells of afferent artelioles, especially during volume depletion. However, the subcellular signaling mechanism of COX-2 gene expression remains unclear. For this purpose, newly established mouse macula densa cells (NE-MD) were used to directly determine whether COX-2 gene expression was regulated by either low [NaCl] concentration or intracellular cAMP/Ca²⁺. In the present study, COX-2 gene expression was evaluated by real-time quantitative PCR. Real-time PCR showed a significant increase in COX-2 gene expression in the samples with low [Cl⁻] solution and culture media plus furosemide, an inhibitor of Na⁺-K⁺-2Cl⁻ cotransporter, compared to controls. However, no increase was observed in the samples with a low [Na⁺] solution. Low Cl⁻-induced increase in COX-2 expression was completely reversed in the presence of either BAPTA-AM, a membrane-permeable Ca²⁺ chelator, EIPA, an inhibitor of Na⁺/H⁺ exchanger, or H-89, an inhibitor of protein kinase A. Finally, COX-2 gene expression was significantly increased in the presence of forskolin (normal solution), whereas it was reversed by H-89. In conclusion, low [Cl⁻]-induced COX-2 gene expression may be regulated by intracellular cAMP, pH, Ca²⁺, and [NaCl]. [J Physiol Sci. 2007;57 Suppl:S218]