J Korean Med Sci.  2008 Dec;23(6):1039-1045. 10.3346/jkms.2008.23.6.1039.

Expression of NAD(P)H Oxidase Subunits and Their Contribution to Cardiovascular Damage in Aldosterone/Salt-Induced Hypertensive Rat

• Park YM
• Lim BH
• Touyz RM
• Park JB

• KMID: 1107531
• DOI: http://doi.org/10.3346/jkms.2008.23.6.1039

Abstract

NAD(P)H oxidase plays an important role in hypertension and its complication in aldosterone-salt rat. We questioned whether NAD(P)H oxidase subunit expression and activity are modulated by aldosterone and whether this is associated with target- organ damage. Rats were infused with aldosterone (0.75 microgram/hr/day) for 6 weeks and were given 0.9% NaCl+/-losartan (30 mg/kg/day), spironolactone (200 mg/kg/ day), and apocynin (1.5 mM/L). Aldosterone-salt hypertension was prevented completely by spironolactone and modestly by losartan and apocynin. Aldosterone increased aortic NAD(P)H oxidase activity by 34% and spironolactone and losartan inhibited the activity. Aortic expression of the subunits p47(phox), gp91(phox), and p22(phox) increased in aldosterone-infused rats by 5.5, 4.7, and 3.2-fold, respectively, which was decreased completely by spironolactone and partially by losartan and apocynin. Therefore, the increased expression of NAD(P)H oxidase may contribute to cardiovascular damage in aldosterone-salt hypertension through the increased expression of each subunit.

Keyword

Aldosterone; Oxidative Stress; NAD(P)H Oxidase; Hypertension

MeSH Terms

Acetophenones/administration & dosage
Aldosterone/administration & dosage/*toxicity
Aldosterone Antagonists/administration & dosage
Angiotensin II Type 1 Receptor Blockers/administration & dosage
Animals
Anti-Inflammatory Agents, Non-Steroidal/administration & dosage
Aorta/metabolism/pathology
Blood Pressure/drug effects
Hypertension/chemically induced/drug therapy/*enzymology
Kidney/metabolism/pathology
Losartan/administration & dosage
Male
NADPH Oxidase/antagonists & inhibitors/*metabolism
Organ Size
Oxidative Stress
Protein Subunits/metabolism
RNA, Messenger/metabolism
Rats
Rats, Sprague-Dawley
Sodium Chloride/administration & dosage
Spironolactone/administration & dosage
Superoxides/metabolism

Figure

• 

  Fig. 1 Systolic BP of aldosterone-infused hypertensive rats was slightly but significantly elevated relative to that of normotensive control rats after 6 weeks of treatment, and showed further elevation thereafter. Administration of spironolactone treatment prevented development of hypertension. Losartan and apocynin showed a trend of decrease. *, p<0.01 aldosterone vs. aldosterone+losartan or aldosterone+spironolactone or aldosterone+apocynin; †, p<0.01 aldosterone+spironolactone vs. aldosterone+losartan or aldosterone+apocynin (mean±SEM).

• 

  Fig. 2 NAD(P)H oxidase activity is increased in aorta from aldosterone-infused rats. (A) Activity of NAD(P)H oxidase in aortic segments from aldosterone-infused rats were detected by lucigenin chemiluminescence. *, p<0.01 vs. control rats; †, p<0.05 vs. aldosterone-infused rats. (B) Aldosterone-indused superoxide (O2-) production is attenuated in kidney of spironolactone, losartan and apocynin treated rats. O2- levels were determined by fluorescent dihydroethidium (DHE [10 µM/L]) and visualized by confocal microscopy. Arrow indicates intense O2- production. RLU, relative light units.

• 

  Fig. 3 Treatment with spironolactone attenuated NAD(P)H oxidase mRNA expression in aorta (A) and kidney (B) of aldosterone-infused rats. The mRNA expression of the NAD(P)H oxidase subunits p47phox, gp91phox, and p22phox was markedly increased in aldosterone-infused rats. The treatment with spironolactone significantly reduced NAD(P)H oxidase subunits mRNA expression. Losartan and apocynin decreased NAD(P)H oxidase subunits mRNA expression in aorta and kidney from aldosterone-infused rats. *, p<0.05 vs. control rats; †, p< 0.05 vs. aldosterone-infused rats.

• 

  Fig. 4 Immunostaining for NAD(P)H oxidase subunit, p47phox in aorta (A) and kidney (B) of control rats, aldosterone-salt rat, aldosterone-salt rat treated with losartan, spironolactone and apocynin. (A) The expression of p47phox was increased in aorta of aldosterone-infused rats. Spironolactone significantly decreased p47phox expression in aorta of aldosterone-infused rats. (B) The expression of p47phox was increased in kidney aldosterone-infused rats. Spironolactone significantly decreased p47phox expression in aorta of aldosterone-infused rats.

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