Avaliação dos efeitos vasculares do sildenafil na aterosclerose experimental

• Autor(a) principal: Balarini, Camille de Moura
• Data de Publicação: 2013
• Tipo de documento: Tese
• Idioma: por
• Título da fonte: Repositório Institucional da Universidade Federal do Espírito Santo (riUfes)
• Texto Completo: http://repositorio.ufes.br/handle/10/8054
• topic: atherosclerosis; endothelial dysfunction; nitric oxide; sildenafil; aterosclerose; disfunção endotelial; óxido nítrico; Fisiologia; 612

Endothelial dysfunction is a sine qua non condition to the development of atherosclerosis. Experimentally, this can be demonstrated by an impaired endothelium-dependent vasodilator response to acetylcholine (ACh) that involve nitric oxide (NO). Thus, pharmacological agents that potentiate NO action are considered promising strategies to improve vascular function and reduce atherosclerosis. Among these agents, sildenafil appears to be a good option, once it inhibits phosphodiesterase 5, the enzyme responsible for degrade cGMP, the most important second messenger of NO. So, the aim of this study was to test if sildenafil can ameliorates endothelial dysfunction in experimental atherosclerosis. Male wildtype C57BL/6 (WT) and apolipoprotein E knockout (apoE-/-) mice, which received Western-type diet, were used. ApoE-/- were divided in two groups: treated animals, which received orally sildenafil citrate (40 mg/Kg/day, n=3-10), and vehicle animals, which received vehicle only (n=3-8); WT animals (n=3-10) were used as controls. At the end of treatment, animals were euthanized and had the thoracic aorta removed. Rings were mounted for vascular studies. Vascular function was accessed by concentration-responses curves to cumulative concentrations of ACh (100 pM – 30 µM) or sodium nitroprusside (SNP; 10 pM – 30 µM) , after pre-contraction with phenylephrine (10 µM). To test the influence of NO and reactive oxygen species (ROS) in relaxation responses, rings were pre-incubated with L-NAME (100 µM) or apocynin (300 µM), respectively. In a different set of animals, after the experimental period, mice had their aorta excised and processed to histological evaluation of plaque deposition (dye with Oil Red), ROS production (label with dihydroethidium - DHE) and NO (label with diaminofluorescein - DAF). Responses were expressed as the percentage of dilation relative to the maximal pre-contraction. The maximum effect (Rmax) and the log of the dose of agonist that produced half of Rmax (log EC50) were calculated. Results of pharmacological blockage were expressed as differences in the area under the concentration-responses curves (dAUC) with and without blockage. Values are expressed as means ± SEM. Statistical comparisons were done by ANOVA, followed by Tukey’s post hoc test. Values of *p<0.05 or **p<0.01 vs. WT; # p<0.05 or ##p<0.01 vs. apoE-/- vehicle and § p<0.05 or §§p<0.01 vs. the same group without blockage were regarded as statistically significant. ApoE-/- animals showed markedly vascular dysfunction (Rmax: 66±9.7* e pEC50: 6.1±0.1**) when 10 compared to WT (Rmax: 87±3.6 e pEC50: 7.3±0.1), which was reversed by sildenafil treatment (Rmax: 95±3.1# e pEC50: 7.2±0.3##). This dysfunction was not due to reduction in vascular smooth muscle sensitivity to NO, once no differences were found in SNP responses. The role of NO in relaxation of apoE-/- was diminished (dAUC: 58.3±16.8** vs. WT: 230±10.6) but was restored by sildenafil (233±10.2##). Also, the influence of ROS in reduced vasodilation of apoE-/- was reversed by sildenafil (CT Rmax: 84±5.2 e pEC50: 7.3±0.2; apoE-/- Rmax: 101±4.6§§ e pEC50: 7.3±0.2§§; apoE-/- sildenafil Rmax: 94±3.7 e pEC50: 7.2±0.2). Atherosclerotic plaque evaluation reveled a markedly plaque deposition in aorta of apoE-/- when compared to WT (37.7±3.4** vs. 0.4±0.4%). There was a reduction of 40% in plaque deposition in apoE-/- mice which received sildenafil (37.7±3.4 vs. 21.3±5.0# ). Also, even in control group, a basal level of ROS production was observed (2.25±0.12). In apoE-/- animals there was a increase in oxidative stress (3.47±0.41*), which was restored to the control levels by sildenafil treatment (2.42±0.21# ). Atherosclerotic animals treated with vehicle showed a reduction in NO production when compared to WT (38±5.2 vs. 17.8±1.2*). Chronic sildenafil treatment was able to revert this situation (17.8±1.2 vs. 38.2±6.8# ). Thus, sildenafil treatment restores endothelial function in experimental atherosclerosis. The mechanisms involved in this response do not involve increase in sensitivity of vascular smooth muscle to NO, but ratter restores NO role in endothelium-dependent dilation, probably due to increase in bioavailability of this molecule due to reduced oxidative stress and increased NO production. This amelioration in endothelial function reflects on reduced atherosclerotic plaque deposition in aorta.