title: Aldosterone stimulates superoxide production in macula densa cells.
abstract: Two major factors which regulate tubuloglomerular feedback (TGF)-mediated constriction of the afferent arteriole are release of superoxide (O(2)(-)) and nitric oxide (NO) by macula densa (MD) cells. MD O(2)(-) inactivates NO; however, among the factors that increase MD O(2)(-) release, the role of aldosterone is unclear. We hypothesize that aldosterone activates the mineralocorticoid receptor (MR) on MD cells, resulting in increased O(2)(-) production due to upregulation of cyclooxygenase-1 (COX-2) and NOX-2, and NOX-4, isoforms of NAD(P)H oxidase. Studies were performed on MMDD1 cells, a renal epithelial cell line with properties of MD cells. RT-PCR and Western blotting confirmed the expression of MR. Aldosterone (10(-8) mol/l for 30 min) doubled MMDD1 cell O(2)(-) production, and this was completely blocked by MR inhibition with 10(-5) mol/l eplerenone. RT-PCR, real-time PCR, and Western blotting demonstrated aldosterone-induced increases in COX-2, NOX-2, and NOX-4 expression. Inhibition of COX-2 (NS398), NADPH oxidase (apocynin), or a combination blocked aldosterone-induced O(2)(-) production to the same degree. These data suggest that aldosterone-stimulated MD O(2)(-) production is mediated by COX-2 and NADPH oxidase. Next, COX-2 small-interfering RNA (siRNA) specifically decreased COX-2 mRNA without affecting NOX-2 or NOX-4 mRNAs. In the presence of the COX-2 siRNA, the aldosterone-induced increases in COX-2, NOX-2, and NOX-4 mRNAs and O(2)(-) production were completely blocked, suggesting that COX-2 causes increased expression of NOX-2 and NOX-4. In conclusion 1) MD cells express MR; 2) aldosterone increases O(2)(-) production by activating MR; and 3) aldosterone stimulates COX-2, which further activates NOX-2 and NOX-4 and generates O(2)(-). The resulting balance between O(2)(-) and NO in the MD is important in modulating TGF.
authors.affiliation: Department of Cardiac Surgery, Shadong Provincial Hospital, Shandong University, Jinan, China.
authors.forename: Xiaolong
authors.initials: X
authors.lastname: Zhu
authors.forename: R Davis
authors.initials: RD
authors.lastname: Manning
authors.forename: Deyin
authors.initials: D
authors.lastname: Lu
authors.forename: Celso E
authors.initials: CE
authors.lastname: Gomez-Sanchez
authors.forename: Yiling
authors.initials: Y
authors.lastname: Fu
authors.forename: Luis A
authors.initials: LA
authors.lastname: Juncos
authors.forename: Ruisheng
authors.initials: R
authors.lastname: Liu
mesh_terms: D000098:Acetophenones / Q000494:pharmacology; D000450:Aldosterone / Q000494:pharmacology*; D000818:Animals; D002460:Cell Line; D051546:Cyclooxygenase 2 / Q000187:drug effects / Q000378:metabolism; D052246:Cyclooxygenase 2 Inhibitors / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004847:Epithelial Cells / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism*; D007668:Kidney / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism*; D008562:Membrane Glycoproteins / Q000378:metabolism; D051379:Mice; D008822:Mice, Transgenic; D023421:Models, Animal; D000074662:NADPH Oxidase 2; D000074663:NADPH Oxidase 4; D019255:NADPH Oxidases / Q000037:antagonists & inhibitors / Q000378:metabolism; D009578:Nitrobenzenes / Q000494:pharmacology; D034741:RNA, Small Interfering / Q000494:pharmacology; D018161:Receptors, Mineralocorticoid / Q000378:metabolism; D013449:Sulfonamides / Q000494:pharmacology; D013481:Superoxides / Q000378:metabolism*
chemical_list: D000098:Acetophenones; D052246:Cyclooxygenase 2 Inhibitors; D004791:Enzyme Inhibitors; D008562:Membrane Glycoproteins; D009578:Nitrobenzenes; D034741:RNA, Small Interfering; D018161:Receptors, Mineralocorticoid; D013449:Sulfonamides; D013481:Superoxides; C080955:N-(2-cyclohexyloxy-4-nitrophenyl)methanesulfonamide; D000450:Aldosterone; C056165:acetovanillone; C077623:Ptgs2 protein, mouse; D051546:Cyclooxygenase 2; C488842:Cybb protein, mouse; D000074662:NADPH Oxidase 2; D000074663:NADPH Oxidase 4; D019255:NADPH Oxidases; C422943:Nox4 protein, mouse