ObjectiveOmega-3 fatty acids, particularly eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), likely prevent cardiovascular disease, however their mechanisms remain unclear. Recently, the role of DNA damage in atherogenesis has been receiving considerable attention. Here, we investigated the effects of EPA and DHA on DNA damage in vascular endothelial cells to clarify their antiatherogenic mechanisms.Methods and resultsWe determined the effect of EPA and DHA on H2O2-induced DNA damage response in human aortic endothelial cells. Immunofluorescence staining showed that γ-H2AX foci formation, a prominent marker of DNA damage, was significantly reduced in the cells treated with EPA and DHA (by 47% and 48%, respectively). H2O2-induced activation of ATM, a major kinase orchestrating DNA damage response, was significantly reduced with EPA and DHA treatment (by 31% and 33%, respectively). These results indicated EPA and DHA attenuated DNA damage independently of the DNA damage response. Thus the effects of EPA and DHA on a source of DNA damage were examined. EPA and DHA significantly reduced intracellular reactive oxygen species under both basal condition and H2O2 stimulation. In addition, the mRNA levels of antioxidant molecules, such as heme oxygenase-1, thioredoxin reductase 1, ferritin light chain, ferritin heavy chain and manganese superoxide dismutase, were significantly increased with EPA and DHA. Silencing nuclear factor erythroid 2-related factor 2 (NRF2) remarkably abrogated the increases in mRNA levels of antioxidant molecules and the decrease in intracellular reactive oxygen species. Furthermore, EPA and DHA significantly reduced H2O2-induced senescence-associated β-galactosidase activity in the cells (by 31% and 22%, respectively), which was revoked by NRF2 silencing.ConclusionsOur results suggested that EPA and DHA attenuate oxidative stress-induced DNA damage in vascular endothelial cells through upregulation of NRF2-mediated antioxidant response. Therefore omega-3 fatty acids likely help prevent cardiovascular disease, at least in part, by their genome protective properties.

ObjectiveCigarette smoking is a major risk factor for atherosclerotic cardiovascular disease, which is responsible for a significant proportion of smoking-related deaths. However, the precise mechanism whereby smoking induces this pathology has not been fully delineated. Based on observation of DNA double-strand breaks (DSBs), the most harmful type of DNA damage, in atherosclerotic lesions, we hypothesized that there is a direct association between smoking and DSBs. The goal of this study was to investigate whether smoking induces DSBs and smoking cessation reverses DSBs in vivo through examination of peripheral mononuclear cells (MNCs).Approach and ResultsImmunoreactivity of oxidative modification of DNA and DSBs were increased in human atherosclerotic lesions but not in the adjacent normal area. DSBs in human MNCs isolated from the blood of volunteers can be detected as cytologically visible “foci” using an antibody against the phosphorylated form of the histone H2AX (γ-H2AX). Young healthy active smokers (n = 15) showed increased γ-H2AX foci number when compared with non-smokers (n = 12) (foci number/cell: median, 0.37/cell; interquartile range [IQR], 0.31–0.58 vs. 4.36/cell; IQR, 3.09–7.39, p<0.0001). Smoking cessation for 1 month reduced the γ-H2AX foci number (median, 4.44/cell; IQR, 4.36–5.24 to 0.28/cell; IQR, 0.12–0.53, p<0.05). A positive correlation was noted between γ-H2AX foci number and exhaled carbon monoxide levels (r = 0.75, p<0.01).ConclusionsSmoking induces DSBs in human MNCs in vivo, and importantly, smoking cessation for 1 month resulted in a decrease in DSBs to a level comparable to that seen in non-smokers. These data reinforce the notion that the cigarette smoking induces DSBs and highlight the importance of smoking cessation.

L ung cancer is the leading cause of cancer deaths (1). Its early detection and treatment are important to reduce the mortality rate. According to the National Lung Screening Trial, annual lung cancer screening by using low-dose (LD) chest CT reduced the number of deaths in patients at high risk (2). LD CT screening is not without risk and although the associated radiation dose is about one-fourth that of standard-dose (SD) CT (3), on the basis of the linear nonthreshold theory, the cancer risk from radiation exposure at LD CT cannot be ignored. Annual screening has been suggested to increase the risk for lung cancer (4,5), although the radiation-induced cancer risk from exposure to less than 100 mSv remains unclear (6,7).g-H2AX analysis to evaluate the biologic effects of lowdose radiation including CT revealed that exposure even at the diagnostic level induces DNA damage (8-14). Dicentric and ring chromosome analyses are the reference standard in biodosimetry. Immunofluorescence analysis of nuclear foci formed by the phosphorylated histone H2AX (g-H2AX), a technically simple and highly sensitive method, yields a marker of DNA double-strand breaks (15). The effects of LD CT on DNA damage and chromosome aberrations (CAs) in adults remain to be clarified before the implementation of

Cigarette smoking is a major risk factor for atherosclerosis. We previously reported that DNA damage was accumulated in atherosclerotic plaque, and was increased in human mononuclear cells by smoking. As vascular endothelial cells are known to modulate inflammation, we investigated the mechanism by which smoking activates innate immunity in endothelial cells focusing on DNA damage. Further, we sought to characterize the plasma level of cell-free DNA (cfDNA), a result of mitochondrial and/or genomic DNA damage, as a biomarker for atherosclerosis. Cigarette smoke extract (CSE) increased DNA damage in the nucleus and mitochondria in human endothelial cells. Mitochondrial damage induced minority mitochondrial outer membrane permeabilization, which was insufficient for cell death but instead led to nuclear DNA damage. DNA fragments, derived from the nucleus and mitochondria, were accumulated in the cytosol, and caused a persistent increase in IL-6 mRNA expression via the cGAS-STING pathway. cfDNA, quantified with quantitative PCR in culture medium was increased by CSE. Consistent with in vitro results, plasma mitochondrial cfDNA (mt-cfDNA) and nuclear cfDNA (n-cfDNA) were increased in young healthy smokers compared to age-matched non-smokers. Additionally, both mt-cfDNA and n-cfDNA were significantly increased in patients with atherosclerosis compared to the normal controls. Our multivariate analysis revealed that only mt-cfDNA predicted the risk of atherosclerosis. In conclusion, accumulated cytosolic DNA caused by cigarette smoke and the resultant activation of the cGAS-STING pathway may be a mechanism of atherosclerosis development. The plasma level of mt-cfDNA, possibly as a result of DNA damage, may be a useful biomarker for atherosclerosis.

• DNA damage was induced by radiation exposure from cardiac CT. • The γ-H2AX foci number was correlated with the CT radiation dose. • Physical exposure parameters reflect the DNA damage by CT radiation exposure.

Deficiency of X-ray repair cross-complementing protein 3 (XRCC3), a DNA-damage repair molecule, and the 241Met variant of XRCC3 have been reported to increase endoreduplication, which induces polyploidy. The aims of this study were to determine the impact of the XRCC3 polymorphism on the incidence of hypertension-induced left ventricular hypertrophy (LVH) and to investigate the mechanisms underlying any potential relationship. Patients undergoing chronic hemodialysis (n = 77) were genotyped to assess for the XRCC3 Thr241Met polymorphism. The XRCC3 241Thr/Met genotype was more frequent in the LVH (+) group than in the LVH (-) group (42.3 vs. 13.7%, χ2 = 7.85, p = 0.0051). To investigate possible mechanisms underlying these observations, human XRCC3 cDNA of 241Thr or that of 241Met was introduced into cultured CHO cells. The surface area of CHO cells expressing XRCC3 241Met was larger than that expressing 241Thr. Spontaneous DNA double-strand breaks accumulated to a greater degree in NIH3T3 cells expressing 241Met (3T3-241Met) than in those expressing 241Thr (3T3-241Thr). DNA damage caused by radiation induced cell senescence more frequently in 3T3-241Met. The levels of basal and TNF-α-stimulated MCP-1 mRNA and protein secretion were higher in 3T3-241Met. Finally, FACS analysis revealed that the cell percentage in G2/M phase including polyploidy was significantly higher in 3T3-241Met than in 3T3-241Thr. Furthermore, the basal level of MCP-1 mRNA positively correlated with the cell percentage in G2/M phase and polyploidy. These data suggest that the XRCC3 241Met increases the risk of LVH via accumulation of DNA damage, thereby altering cell cycle progression and inducing cell senescence and a proinflammatory phenotype.

Aortic aneurysm refers to dilatation of the aorta due to loss of elasticity and degenerative weakening of its wall. A preventive role for osteoprotegerin (Opg) in the development of abdominal aortic aneurysm has been reported in the CaCl 2-induced aneurysm model, whereas Opg was found to promote suprarenal aortic aneurysm in the AngII-induced ApoE knockout mouse aneurysm model. To determine whether there is a common underlying mechanism to explain the impact of Opg deficiency on the vascular structure of the two aneurysm models, we analyzed suprarenal aortic tissue of 6-month-old ApoE-/-Opg-/mice after AngII infusion for 28 days. Less aortic dissection and aortic lumen dilatation, more adventitial thickening, and higher expression of collagen I and Trail were observed in ApoE-/-Opg-/mice relative to ApoE-/-Opg +/+ mice. An accumulation of α-smooth muscle actin and vimentin double-positive myofibroblasts was noted in the thickened adventitia of ApoE-/-Opg-/mice. Our results suggest that fibrotic remodeling of the aorta induced by myofibroblast accumulation might be an important pathological event which tends to limit AngIIinduced aortic dilatation in ApoE-/-Opg-/mice.

No abstract