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Modulation of aortic reactivity to angiotensin-(1- 7) with intermittent hyperbaric oxygenation in diabetic rats

We previously reported a facilitating effect of hyperbaric oxygen therapy (HBOT) on responses of rat aortic rings to angiotensin-(1-7) (ANG-(1-7)) in healthy rats (‘ Physiology 2011')[1]. It was suggested that this influence was mediated by changes in production or sensitivity to epoxyeicosatrienoic acids (EETs). The aim was to assess whether HBOT exerts similar effects in diabetic rats and to further explore the role of cytochrome P450 (CYP) enzymes in changes induced by HBOT. Diabetes was induced by streptozocin in 6 weeks old male Sprague-Dawley rats. After 8 weeks, they were divided into control and HBOT group (exposed to 100% O2(2 bars) 2 hours/day for 4 consecutive days). Before decapitation, rats were anesthetized with 75 mg/kg ketamine with 2.5 mg/kg midazolam. Thoracic aortic rings were used to test responses to 1 μ M ANG-(1-7) after precontraction with noradrenaline for 5 minutes (tension was read 3 minutes after ANG-(1-7) addition and the response expressed as percentage of precontraction decrease). Intactness of endothelium was previously tested. A separate series of HBOT rats was used to test ANG-(1-7) responses after the epoxygenases inhibitor MS-PPOH (10 μ M) - inhibitor of the CYP4A2 AND CYP4A3 epoxygenation reactions - was added for 15 minutes to the tissue bath. CYP 4A1, 4A2, 4A3 and 2J3 mRNA expression was determined with Quantitative real time PCR and was normalized to the expression of two housekeeping genes -HPRT and 18S. The study was approved by Ethical Committee of Faculty of Medicine University of Osijek. The route of administration of both drugs is intraperitoneal (i.p.). Mean percentage of precontraction decrease after ANG-(1-7) addition was significantly higher in the HBOT group (19.2% ± 7.3 [STDEV]) (n=18) than in the control group (12.1% ± 6.4)(n=14), P= 0.007 (t-test) . The ANG-(1-7) ring response of HBOT rats after MS- PPOH addition was 2.3% ± 3.3 (n=8), significantly less pronounced compared to HBOT rings when no MS-PPOH was used, P< ; 0.001 (Mann Whitney U). Median CYP2J3/18s expression was 12.2 (control)(n=7) and 94.8 (HBOT)(n=6), significantly different (P=0.008, Mann Whitney U). Median CYP2J3/HPRT expression was 7.5 (control)(n=9) and 49.7(HBOT)(n=9), with significant difference (P= 0.042). CYP4A1, CYP4A2 and CYP4A3 expression was similar between groups. HBOT increases the reactivity of thoracic aortic rings to ANG-(1- 7) in diabetic rats. The epoxygenases inhibitor MS-PPOH reverses these changes. HBOT increases mRNA expression of the epoxygenase CYP2J3 in diabetic rats, further suggesting an important role of EETs in HBOT-induced modulation of vascular function.

hyperbaric oxygenation; angiotensin; diabetes

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