BackgroundOxidative stress and DNA damage have been implicated in the pathogenesis of renovascular hypertension induced by renal artery stenosis in the two-kidney, one-clip (2K1C) Goldblatt model. Considering our previous report indicating that the chronic blockade of phosphodiesterase 5 with sildenafil (Viagra®) has marked beneficial effects on oxidative stress and DNA damage, we tested the hypothesis that sildenafil could also protect the stenotic kidneys of 2K1C hypertensive mice against oxidative stress and genotoxicity.MethodsThe experiments were performed with C57BL6 mice subjected to renovascular hypertension by left renal artery clipping. Two weeks after clipping, the mice were treated with sildenafil (40 mg/kg/day for 2 weeks, 2K1C-sildenafil group) or the vehicle (2K1C). These mice were compared with control mice not subjected to renal artery clipping (Sham). After hemodynamic measurements, the stenotic kidneys were assessed using flow cytometry to evaluate cell viability and the comet assay to evaluate DNA damage. Measurements of intracellular superoxide anions and hydrogen peroxide levels as well as nitric oxide bioavailability were also obtained.ResultsSildenafil treatment significantly reduced mean arterial pressure (15%), heart rate (8%), intrarenal angiotensin II (50%) and renal atrophy (36%). In addition, it caused a remarkable decrease of reactive oxygen species production. On the other hand, sildenafil increased nitric oxide levels relative to those in the nontreated 2K1C mice. Sildenafil treatment also significantly reduced the high level of kidney DNA damage that is a characteristic of renovascular hypertensive mice.ConclusionsOur data reveal that sildenafil has a protective effect on the stenotic kidneys of 2K1C mice, suggesting a new use of phosphodiesterase 5 inhibitors for protection against the DNA damage observed in the hypoperfused kidneys of individuals with renovascular hypertension. Further translational research is necessary to delineate the mechanisms involved in the prevention of renal stenosis in the clinical setting.

BackgroundStem cells of intensely regenerative tissues are susceptible to cellular damage. Although the response to this process in hematopoietic stem cells (HSCs) is crucial, the mechanisms by which hematopoietic homeostasis is sustained are not completely understood. Aging increases reactive oxygen species (ROS) levels and inflammation, which contribute to increased proliferation, senescence and/or apoptosis, leading to self-renewal premature exhaustion. In this study, we assessed ROS production, DNA damage, apoptosis, senescence and plasticity in young, middle and aged (2-, 12- and 24-month-old, respectively) C57BL/6 J mice.ResultsAged HSCs showed an increase in intracellular superoxide anion (1.4-fold), hydrogen peroxide (2-fold), nitric oxide (1.6-fold), peroxynitrite/hidroxil (2.6-fold) compared with young cells. We found that mitochondria and NADPHox were the major sources of ROS production in the three groups studied, whereas CYP450 contributed in middle and aged, and xanthine oxidase only in aged HSCs. In addition, we observed DNA damage and apoptosis in the middle (4.2- and 2-fold, respectively) and aged (6- and 4-fold, respectively) mice; aged mice also exhibited a significantly shorter telomere length (−1.8-fold) and a lower expression of plasticity markers.ConclusionThese data suggest that aging impairs the functionality of HSCs and that these age-associated alterations may affect the efficacy of aged HSC recovery and transplantation.

BackgroundIt is well known that enhanced production of reactive oxygen species (ROS) leads to oxidative stress observed in atherosclerosis and that ROS can also cause damage in cellular macromolecules, including DNA. Considering previous report that sildenafil, an inhibitor of phosphodiesterase 5 (PDE5), has antioxidant effects, in the present study we evaluated the effect of this drug on genotoxicity of blood mononuclear cells (MNC) and liver cells from atherosclerotic apolipoprotein E knockout mice (apoE-/-).MethodsROS production in MNC was evaluated by flow cytometry with the fluorescent dye dihydroethidium (DHE), a method that has been used to quantify the production of superoxide anion, and DNA damage was evaluated in both MNC and liver cells using the alkaline comet assay. Sildenafil-administered apoE-/- mice were compared with strain-matched mice administered with vehicle and with C57BL/6 wild-type (WT) mice.ResultsMNC from apoE-/- vehicle exhibited a 2-fold increase in production of superoxide anion in comparison with WT. In contrast, sildenafil-administered apoE-/- mice showed superoxide anion levels similar to those observed in WT mice. Similarly, MNC and liver cells from apoE-/- vehicle mice showed a 4-fold and 2-fold augmented DNA fragmentation compared with WT, respectively, and sildenafil-administered apoE-/- mice exhibited minimal DNA damage in those cells similar to WT mice.ConclusionsApoE-/- mice chronically administered with sildenafil exhibited reduced levels of superoxide anion in MNC and less DNA fragmentation in MNC and liver cells, which are biomarkers of genotoxicity. Therefore, sildenafil may offer a new perspective to the use of PDE5 inhibitors to protect against DNA damage, in cells involved in the inflammatory and dyslipidemic processes that accompany atherosclerosis.

BackgroundStem cell function is compromised with aging, especially when a cardiovascular disease is associated. The aim of this study was to evaluate how aging and atherosclerosis affect bone marrow hematopoietic stem cell (HSC) function in mice.Methods: Male mice were divided into C57 and apoE‐/‐ (2, 12 and 24‐month‐old) groups. Oxidative stress was assessed by 2,7‐dichlorofluorescein (DCF) by flow cytometer. For apoptosis detection, HSC were incubated with Annexin V‐FITC and propidium iodide (PI). Plasma inflammation markers, IL‐6 and MCP‐1, were measured. Also, senescence was evaluated by acridine orange protocol. Lipid profile and aortic lipid deposition were also analyzed. Data are mean±SEM. Statistical analysis was performed with two way ANOVA test and Tukey pos hoc.Results: Aging and atherosclerosis increased ROS levels and apoptosis in HSC. Senescence was also increased with aging and atherosclerosis, especially in 24‐month‐old mice. Atherosclerotic adult and aged animals showed higher levels of IL‐6 and MCP‐1. As expected, atherosclerosis augmented total serum cholesterol levels and aortic lipid deposition.ConclusionThe present study provides evidence that aging and atherosclerosis can cause oxidant stress and impair HSC function. Such findings could lead to new insights into the mechanisms by which aging and atherosclerosis can affect immunity and inflammation.