Effect of apocynin on blood pressure and oxidative stress parameters in young WKY and SHR rats

Pedro Gomes, Elisabete Silva, Sónia Simão, Vanda Pinto, Maria João Pinho, João Amaral, Joana Afonso, Paula Serrão, Patrício Soares-da-Silva

Institute of Pharmacology and Therapeutics, Faculty of Medicine, University of Porto, Porto, Portugal

Although numerous factors including genetic and environmental influences have been implicated in the pathophysiology of hypertension, recent studies indicate a tight relationship between oxidative stress and the development and maintenance of hypertension. Increased oxidative stress has been reported in animal models of hypertension (Dobrian et al., 2004; Nishiyama et al., 2004; Tanito et al., 2004) and in human hypertension (Higashi et al., 2002; Redon et al., 2003). Some studies have shown that antioxidant treatment reduced the elevated blood pressure (Baumer et al., 2007; Schnackenberg et al., 1999) indicating a role of oxidative stress in the etiology of hypertension. This study tested the hypothesis that antioxidant treatment with the NADPH oxidase inhibitor apocynin may reduce the severity of hypertension in spontaneously hypertensive rats (SHR) and attenuate oxidative stress. The study further sought to explore the effect of apocynin on renal dopaminergic activity. Twelve-week old male SHR and their normotensive controls, Wistar Kyoto (WKY) rats, were treated for 10 days with apocynin (100 mg/Kg/day, s.c.). Systolic and diastolic blood pressure were monitored by the tail-cuff method. Urinary 8-isoprostanes, hydrogen peroxide (H2O2) and malondialdehyde (MDA) were used as parameters of oxidative stress. The status of the renal dopaminergic system activity was based on the urinary excretion of L-DOPA, dopamine and its deaminated metabolite 3,4-dihydroxyphenylacetic acid (DOPAC). In some experiments, immortalized proximal tubule cells from WKY and SHR and also cortical tissue from Wistar rats were used to determine the effect of apocynin on L-DOPA uptake and aromatic L-amino acid decarboxylase (AADC) activity, respectively. Apocynin treatment failed to attenuate high blood pressure in the SHR. Urinary levels of 8-isoprostanes and H2O2 were significantly (P<0.05) elevated in the SHR, which indicates the presence of oxidative stress; however, no significant changes in these parameters were observed after antioxidant therapy. Urinary L-DOPA excretion was markedly increased (P<0.05) in response to apocynin in both rat strains, without changes in dopamine and DOPAC excretion. Subsequent in vitro studies using proximal tubule cells from WKY and SHR showed that apocynin did not interfere with L-DOPA uptake and outflow. However, apocynin inhibited, in a competitive manner, the conversion of L-DOPA to dopamine (IC50 = 5.4 mM; 95% confidence interval 3.6 to 8.2) in renal cortical extracts from Wistar rats. In conclusion, neither high blood pressure nor oxidative stress was affected by antioxidant treatment with apocynin. Finally, our results unveil a novel function for apocynin serving as a competitive inhibitor of AADC activity.

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