Ethanol Consumption Induces Vascular Oxidative Stress and Alters SOD Antioxidant Activity through NAD(P)H Oxidase

to 16-20 % of normal). These preliminary results suggest that even under SK\VLRORJLFDO FRQGLWLRQV +'/¶V FDUGLR-protective PON1 activity may be particularly sensitive to detrimental effects of carbamylation. 6XSSRUWHG E\ 0LGZHVWHUQ 8QLYHUVLW\¶V 2IILFH of Research and Sponsored Programs (SML) and Biomedical Sciences Research Program (JP).

doi: 10.1016/j.freeradbiomed.2014.10.412

98 Oxidized Low-Density Lipoprotein Accelerates the Degradation of EC-SOD mRNA during Foam Cell Formation in THP-1 Macrophages Junya Makino1, Miyuki Nii1, Tetsuro Kamiya1, Hirokazu Hara1, and Tetsuo Adachi1 1 Gifu Pharmaceutical University, Japan Purpose: Extracellular-SOD (EC-SOD) protects the cells from damaging effects of superoxide and the development of atherosclerosis, suggesting that EC-SOD might have an important protective role in vascular systems. We investigated the regulation of EC-SOD expression during oxidized LDL (oxLDL)induced foam cell formation of THP-1-derived macrophages. Method: THP-1 cells were differentiated into macrophages by the treatment with TPA, and then treated with oxLDL. RT-PCR, Western blotting and ChIP assay were carried out. Results and Discussion: The uptake of oxLDL into THP-1-derived macrophages was confirmed by Oil red O staining and the expression of EC-SOD was decreased by the treatment with oxLDL but not with native LDL. It has been reported that the expression of EC-SOD is regulated by histone acetylation and Sp1/3 binding to EC-SOD promoter region. However, oxLDL did not affect these processes. On the other hand, the stability of ECSOD mRNA was decreased by oxLDL in the presence of actinomycin D. Interestingly, the degradation of ectopically expressed EC-SOD mRNA without 3¶-untranslated region (3¶UTR) was not suppressed. Conclusion: oxLDL induces the destabilization of EC-SOD mRNA and 3¶UTR is important for the destabilization of its mRNA. doi: 10.1016/j.freeradbiomed.2014.10.413

99 Ethanol Consumption Induces Vascular Oxidative Stress and Alters SOD Antioxidant Activity through NAD(P)H Oxidase Katia Colombo Marchi1 and Carlos Renato Tirapelli1 1 University of Sao Paulo, Brazil The aim of our study was to evaluate the participation of NAD(P)H oxidase in the cardiovascular effects induced by chronic ethanol consumption through its inhibition by apocynin (APO). All protocols were approved by the Ethics Committee on Animal Use (11.1.1648.53.8). Male Wistar rats were divided into 4 groups: Control: 0.9% saline intraperitoneal (ip) injection; Control+APO: APO injection (10mg/kg/day,ip); Ethanol: ethanol 20% (v/v) for 2 weeks and 0.9% saline ip injection; Ethanol+APO group: ethanol 20% and APO. Ethanol treatment increased the expression (arbitrary units) of Nox1 (1.66±0.16,n=4) and nNOS (0.46±0.07,n=4) in the rat aorta when compared to control animals (Nox1=0.63±0.10,n=4;nNOS=0.18± 0.03,n=4). Treatment with APO prevented these responses (Nox1=0.64±0.32,n=4;

S50

nNOS=0.15±0.03,n=4). There was no difference in the expression of Nox2, Nox4, eNOS and iNOS after treatment with ethanol. Ethanol treatment did not cause changes in the translocation of p47phox in rat aorta. No difference was found in the levels of H2O2 (μmol/L/mg protein) between groups. Plasma 8-isoprostane (pg/ml) levels increased in Ethanol (200.9±12.9,n=8), Control+APO (170.3±14.0,n=6) and Ethanol+APO (180.8±16.2, n=8) compared to Control group (123.3±12.4,n=9). A decrease in plasma superoxide dismutase (SOD) activity (U/ml) in Ethanol (4.4±0.5,n=8) compared to Control group (6.2±0.4,n=8) was found. Treatment with APO prevented this response (7.8±0.7,n=8). Aortic reduced glutathione (GSH) levels (μg/ml/mg protein) increased in Control+APO (6.0±0.8, n=8) compared to Control group (2.7±0.6, n=8). In plasma, GSH levels was increased in Control+APO (4.5±1.0, n=9) and Ethanol+APO (4.0±0.4, n=10) compared to Control (1.0±0.2, n=7) and Ethanol (1.7±0.3, n=8) groups. No difference was observed in aortic glutathione peroxidase (GPx) activity. Aortic catalase (CAT) activity (nmol/min/mg protein) was increased in Control+APO (48.7±2.5, n=9) and Ethanol+APO (52.2±5.6, n=6) compared to Control (35.4±1.8, n=6).Ethanol treatment increased superoxide anion levels in the rat aorta when compared to control animals and treatment with APO prevented this response. Furthermore, there was a decrease in nitrate and nitrite both in plasma and aorta of the animals treated with ethanol and APO treatment did not prevent this response. Chronic ethanol consumption induces vascular oxidative stress, lipidic peroxidation, increases expression of Nox1 and decreases SOD activity, being these responses mediated by NAD(P)H oxidase. doi: 10.1016/j.freeradbiomed.2014.10.414

100 Myeloperoxidase-Deficiency Protects from Ventricular Tachycardia in a Murine Model of Ischemic Cardiomyopathy Martin Mollenhauer1, Max Lange1, Johanna Schneider1, Anna Klinke1, Lisa Remane1, Kai Friedrichs1, Volker Rudolph1, Tanja Rudolph1, Stephan Baldus1, and Volker Rudolph1 1 University Heart Center Cologne, Germany Background: Leukocytes have emerged as important mediators of adverse myocardial remodeling following myocardial infarction. Myeloperoxidase (MPO), a heme enzyme abundantly expressed and released by polymorphonuclear neutrophils (PMN), exerts pro-inflammatory and pro-fibrotic properties and has been identified to be causally linked to the formation and propagation of various cardiovascular disorders. We disclosed recently, that MPO critically promotes atrial fibrosis and arrhythmias. Thus, we sought to investigate whether MPO might also influence structural and electrical remodeling in ventricular myocardium. Methods and Results: Wild-type C57bl/6J (WT) and MPOdeficient (Mpo-/-) mice underwent either sham surgery or ligation of the left descending coronary artery (LAD). Right ventricular electrophysiological investigations 3 weeks after LAD ligation disclosed profoundly increased vulnerability for ventricular tachycardia (VT) in infarcted WT as compared to Mpo-/- mice. Epicardial mapping analyses indicated a delayed velocity and enhanced heterogeneity of electrical conduction in WT as compared to Mpo-/- mice. This was accompanied by a diminished area of fibrosis and a reduced number of activated fibroblasts, identified by positive alpha-VPRRWK PXVFOH DFWLQ Į-SMA) immunoreactivity in infarction and peri-infarcted regions. In vitro experimeQWVZLWKLVRODWHGFDUGLDFILEUREODVWVUHYHDOHGWKDWĮ-SMA expression is augmented upon MPO treatment and co-incubation with isolated MPO secreting PMNs. Conclusions: The current data demonstrate, that MPO increases

SFRBM 2014