Efeitos hipotensor e anti-hipertensivo do nitrato orgânico 1,3-bis (hexiloxi) propano-2-ila (NDHP) sobre o sistema cardiovascular de ratos normotensos e hipertensos

Abstract

Arterial hypertension (AH) is a chronic degenerative disease of multifactorial etiology responsible for thousands of deaths each year. One of the main pathophysiological processes in AH is endothelial dysfunction, which is caused primarily by a decrease in the bioavailability of nitric oxide (NO). NO is an important molecule responsible for the maintenance of vascular homeostasis. The use of compounds that increase NO bioavailability present as a therapeutic strategy in the treatment of AH. Thus, the aim of the present study was to characterize the 1,3-bis (hexyloxy) propane-2-yl nitrate (NDHP) as a novel NO donor and to evaluate its effects on the cardiovascular system of normotensive and hypertensive rats. All results were considered significant when p <0.05. Using in vitro methodologies, it was observed that Xanthine oxidoreductase (XOR) catalyzes the generation of NO from NDHP under anaerobic conditions at the flavin adenine dinucleotide (FAD) and molybdenum sites. NDHP promoted vasorelaxation in mesenteric arteries of rats and mice, using myograph. The pre-exposure of the mesenteric artery rings to the NDHP (10-3) for 30 minutes did not decrease the effect (ME = 124 ± 4% vs. 100 ± 3%, n = 7 and 6, respectively). In the presence of nicotinamide adenine dinucleotide (NADH) and XOR, the administration of NDHP (0.1, 1 and 10 mM) increased NO production in a concentration-dependent manner. Treatment with NDHP increased NO levels in mesenteric artery sections compared to control (95.7 ± 3.4 vs. 10.9 ± 0.8 a.u., respectively, p<0.05). The blockade of endothelial nitric oxide synthase (eNOS) by L-NG-Nitroarginine (L-NNA, 100 μM) did not inhibit the increase in NO levels (94.2 ± 3.6 vs. 95.7 ± 3.4 a.u., respectively). The increase in NO levels after NDHP treatment was blocked by the XOR inhibitor, febuxostat. NDHP induced a vasorelaxative effect on the mesenteric artery with functional endothelium (ME = 83.3 ± 6.7%, n = 6). This maximal effect (ME) was higher in rings without functional endothelium (ME = 113.4 ± 5.3%, n = 7, p<0.05). Pre-incubation with the NOS inhibitor N(ω)-nitro-L-arginine methyl ester (L-NAME) in mesenteric artery with functional endothelium increased NDHP-induced relaxation (ME = 109.7 ± 1.5%; n = 6, p<0.05). Pretreatment with 2-(4-phenyl)-4,4,5,5-tetramethylimidazoline-1-oxy-3-oxide (PTIO, 300 μM), an NO sequestrant along with hydroxocobalamin (HDX) promoted a decrease in the vasorelaxative effect of NDHP (ME = 90.8 ± 0.8%, n = 6, p<0.05). Futhermore, a decrease in the vasorelaxative effect of NDHP was observed in rings without functional endothelium when 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one (ODQ, 10 μM) was used, a selective inhibitor of soluble guanylyl cyclase (sGC) enzyme (84.9 ± 9.5%, n = 6, p<0.05). The potency of the vasorelaxative effect of NDHP was decreased using a non-specific blocker of potassium (K+) channels, 3mM tetramethylammonium chloride (TEA) (pD2 = 4.8 ± 0.1, n = 6, p <0, 05), using a specific BKCa blocker, TEA (1 mM) (pD2 = 5.1 ± 0.1, n = 6, p <0.05) and KV, using 4-aminopyridine (4-AP, 1 mM) (pD2 = 5,7 ± 0,1; n = 6, p < 0,05). Acute treatment with NDHP in high doses (300 and 2000 mg/kg) did not present toxic effect in the toxicity test. In in vivo models, the acute administration of NDHP (1, 5, 10, 20 mg/kg, i.v) induced hypotension in normotensive animals (-11 ± 1, -19 ± 2, -28 ± 2, -44 ± 5 mmHg respectively, n = 8) and hypertensive patients (-16 ± 3, -23 ± 4, -50 ± 1, and -71 ± 8 mmHg respectively, n = 8). Chronic treatment (15 days) with NDHP (10 mg/kg/day), via i.p., significantly attenuated endothelial dysfunction and increased mean arterial pressure in animals with angiotensin II-induced hypertension (MAPvehicle: 49 mmHg vs. MAPNDHP 18 mmHg) when compared to vehicle-treated hypertensive animals (MAPvehicle: 98 ± 2 mm Hg vs. MAPNDHP 99 ± 3 mm Hg, baseline). Plasma nitrate levels were four times higher in NDHP-treated animals compared to the control group (21.6 ± 2.9 vs 4.4 ± 1.1 μM, p <0.05). These results demonstrate that NDHP acts as an NO donor, induces vasorelaxation through the NO-cGMP-PKG pathway and activation of specific subtypes of K+ channels (Kv and BKca), does not cause vascular tolerance in the mesenteric artery, has an antihypertensive effect in animals with renovascular hypertension and by infusion of angiotensin II, requires XOR for the formation of NO and presents low acute toxicity.