engleski

The role of angiotensin II AT-1 type receptors in maintenance of the vascular oxidative stress balance of Sprague-Dawley rats

INTRODUCTION: Blockade of angiotensin II type 1 receptor (AT1R) signaling contributes to impaired vascular reactivity ; however, the mechanism is not clear. Presently, we aimed to determine the effects of AT1R blockade by losartan on flow-induced vascular reactive oxygen species production (ROS) in rat middle cerebral arteries (MCA) and on systemic oxidative stress. METHODS: Healthy male Sprague-Dawley rats (N=6-9 per group) were fed low salt diet for 7 days (0.4% NaCl ; LS group) or LS+Losartan (losartan 40 mg/day in water ad libidum). Following dietary protocol, rats were anesthetized with ketamine (75 mg/kg) and midazolam (2.5 mg/kg) and decapitated. MCA were isolated and cannulated on pressure myograph, under flow (Δ80 mmHg) or no flow conditions, and in the absence/presence of TEMPOL (superoxide scavenger, 100μM final concentration) in vitro. ROS production was determined by DHE (dihydroethidine) fluorescence measurements. Systemic oxidative stress was assessed by measurement of 8-iso prostaglandin concentration (ELISA) and antioxidative enzymes (SOD, GPx and CAT) activity assay in serum by spectrophotometry. mRNA levels of antioxidative enzymes and NADPH oxidase 1 (NOX1) were determined by qPCR in surface cerebral vessels. Data were analysed by t-test ; p<0.05 was considered significant. All experimental procedures are conformed to the European Guidelines for the Care and Use of Laboratory Animals (directive 86/609) and were approved by the local and national Ethical Committee (Class: UP/I-322-01/14-01/36, No. 525-10/0255-15-6). RESULTS: Flow-induced vascular ROS production was significantly increased in LS+Losartan group compared to LS group (p=0.0023).No differences in ROS productions were observed in the absence of flow. TEMPOL in vitro in LS+Losartan group significantly decreased vascular ROS production compared to LS+Losartan in the absence of TEMPOL [flow (p=0.004) and no flow (p=0.0012) conditions], and restored ROS to basal values compared to LS group in flow conditions. 17 8-iso prostaglandin concentration was significantly increased in LS+Losartan group while expression of GPx1 and Cu/Zn SOD was significantly decreased in LS+Losartan group compared to LS group. Expression of MnSOD, EC- SOD, CAT and NOX1 was significantly increased in LS+Losartan group compared to LS. Serum enzymes’ activity of SOD and CAT) was significantly decreased in LS+Losartan group. CONCLUSIONS: Blockade of AT1R enhances flow- induced vascular ROS production in the MCA possible due to increased expression of NOX1 (a source of superoxide), and decreased activity of antioxidative enzymes despite their increased expression in vessels. Additionally, systemic oxidative stress is increased. Acknowledgment: Supported by Croatian Science Foundation under the project #IP-2014-09-6380.

ANG II, oxidative stress, Sprague-Dawley rats

nije evidentirano