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The iron-sulfur cluster containing protein mitoNEET is known to modulate the oxidative capacity of cardiac mitochondria but its function during myocardial reperfusion injury after transient ischemia is unknown. The purpose of this study was to analyze the impact of mitoNEET on oxidative stress induced cell death and its relation to the glutathione-redox system in cardiomyocytes in an in vitro model of hypoxia and reoxygenation (H/R). Our results show that siRNA knockdown (KD) of mitoNEET caused an 1.9-fold increase in H/R induced apoptosis compared to H/R control while overexpression of mitoNEET caused a 53% decrease in apoptosis. Necrosis was not affected. Apoptosis of both, mitoNEET-KD and control cells was diminished to comparable levels by using the antioxidants Tiron and glutathione compound glutathione reduced ethyl ester (GSH-MEE), indicating that mitoNEET-dependent apoptosis is mediated by oxidative stress. The interplay between mitoNEET and glutathione redox system was assessed by treating cardiomyocytes with 2-acetylamino-3-[4-(2-acetylamino-2-carboxyethylsulfanylthio-carbonylamino) phenylthiocarbamoylsulfanyl] propionic acid (2-AAPA), known to effectively inhibit glutathione reductase (GSR) and to decrease the GSH/GSSG ratio. Surprisingly, inhibition of GSR-activity to 20% by 2-AAPA decreased apoptosis of control and mitoNEET-KD cells to 23% and 25% respectively, while at the same time mitoNEET-protein was increased 4-fold. This effect on mitoNEET-protein was not accessible by mitoNEET-KD but was reversed by GSH-MEE. In conclusion we show that mitoNEET protects cardiomyocytes from oxidative stress-induced apoptosis during H/R. Inhibition of GSH-recycling, GSR-activity by 2-AAPA increased mitoNEET-protein, accompanied by reduced apoptosis. Addition of GSH reversed these effects suggesting that mitoNEET can in part compensate for imbalances in the antioxidative glutathione-system and therefore could serve as a potential therapeutic approach for the oxidatively stressed myocardium. [ABSTRACT FROM AUTHOR]

Antibody-mediated rejection (AMR) is a key limiting factor for long-term graft survival in solid organ transplantation. Human leukocyte antigen (HLA) class I (HLA I) antibodies (Abs) play a major role in the pathogenesis of AMR via their interactions with HLA molecules on vascular endothelial cells (ECs). The antioxidant enzyme heme oxygenase (HO)-1 has anti-inflammatory functions in the endothelium. As complement-independent effects of HLA I Abs can activate ECs, it was the goal of the current study to investigate the role of HO-1 on activation of human ECs by HLA I Abs. In cell cultures of various primary human macro- and microvascular ECs treatment with monoclonal pan- and allele-specific HLA I Abs up-regulated the expression of inducible proinflammatory adhesion molecules and chemokines (vascular cell adhesion molecule-1 [VCAM-1], intercellular cell adhesion molecule-1 [ICAM-1], interleukin-8 [IL-8] and monocyte chemotactic protein 1 [MCP-1]). Pharmacological induction of HO-1 with cobalt-protoporphyrin IX reduced, whereas inhibition of HO-1 with either zinc-protoporphyrin IX or siRNA-mediated knockdown increased HLA I Ab-dependent up-regulation of VCAM-1. Treatment with two carbon monoxide (CO)-releasing molecules, which liberate the gaseous HO product CO, blocked HLA I Ab-dependent EC activation. Finally, in an in vitro adhesion assay exposure of ECs to HLA I Abs led to increased monocyte binding, which was counteracted by up-regulation of HO-1. In conclusion, HLA I Ab-dependent EC activation is modulated by endothelial HO-1 and targeted induction of this enzyme may be a novel therapeutic approach for the treatment of AMR in solid organ transplantation. [ABSTRACT FROM AUTHOR]

Objective: Atherosclerosis, a chronic inflammatory disease, arises from metabolic disorders and is driven by inappropriate recruitment and proliferation of monocytes / macrophages and vascular smooth-muscle-cells. The receptor for the urokinase-type plasminogen activator (uPAR, Plaur) regulates the proteolytic activation of plasminogen. It is also a coactivator of integrins and facilitates leukocyte-endothelial interactions and vascular smooth-muscle-cell migration. The role of uPAR in atherogenesis remains elusive. Methods and Results: We generated C57Bl6/J low-density lipoprotein receptor (LDL) and uPAR double knockout (uPAR-/-/LDLR-/-) mice to test the role of uPAR in two distinct atherosclerosis models. In LDLR-/- mice, hepatic overexpression following hydrodynamic transfection of soluble uPAR that competes with endogenous membrane-bound uPAR was performed as an interventional strategy. Aortic root atherosclerotic lesions induced by feeding a high-fat diet were smaller and comprised less macrophages and vascular smooth-muscle-cells in double knockout mice and animals overexpressing soluble uPAR when compared to controls. In contrast, lesion size, lipid-, macrophage-, and vascular smooth muscle cell content of guide-wire-induced intima lesions in the carotid artery were not affected by uPAR deficiency. Adhesion of uPAR-/--macrophages to TNFα-stimulated endothelial cells was decreased in vitro accompanied by reduced VCAM-1 expression on primary endothelial cells. Hepatic overexpression of soluble full-length murine uPAR in LDLR-/- mice led to a reduction of diet-induced atherosclerotic lesion formation and monocyte recruitment into plaques. Ex vivo incubation with soluble uPAR protein also inhibited adhesion of macrophages to TNFα-stimulated endothelial cells in vitro. Conclusion: uPAR-deficiency as well as competitive soluble uPAR reduced diet-promoted but not guide-wire induced atherosclerotic lesions in mice by preventing monocyte recruitment and vascular smooth-muscle-cell infiltration. Soluble uPAR may represent a therapeutic tool for the modulation of hyperlipidemia-associated atherosclerotic lesion formation. [ABSTRACT FROM AUTHOR]

Objective: Precise perioperative risk stratification is important in vascular surgery patients who are at high risk for major adverse cardiovascular events (MACE) peri- and postoperatively. In clinical practice, the patient’s perioperative risk is predicted by various indicators, e.g. revised cardiac index (RCRI) or modifications thereof. Patients suffering from chronic kidney disease (CKD) are stratified into a higher risk category. We hypothesized that Copeptin as a novel biomarker for hemodynamic stress could help to improve the prediction of perioperative cardiovascular events in patients undergoing vascular surgery including patients with chronic kidney disease. Methods: 477 consecutive patients undergoing abdominal aortic, peripheral arterial or carotid surgery from June 2007 to October 2012 were prospectively enrolled. Primary endpoint was 30-day postoperative major adverse cardiovascular events (MACE). Results: 41 patients reached the primary endpoint, including 63.4% aortic, 26.8% carotid, and 9.8% peripheral surgeries. Linear regression analysis showed that RCRI (P< .001), pre- (P< .001), postoperative Copeptin (P< .001) and Copeptin level change (P= .001) were associated with perioperative MACE, but CKD remained independently associated with MACE and Copeptin levels. Multivariate regression showed that increased Copeptin levels added risk predictive information to the RCRI (P= .003). Especially in the intermediate RCRI categories was Copeptin significantly associated with the occurrence of MACE. (P< .05 Kruskal Wallis test). Subdivision of the study cohort into CKD stages revealed that preoperative Copeptin was significantly associated with CKD stages (P< .0001) and preoperative Copeptin measurements could not predict MACE in patients with more severe CKD stages. Conclusion: Preoperative Copeptin loses its risk predictive potential for perioperative MACE in patients with chronic kidney disease undergoing vascular surgery. [ABSTRACT FROM AUTHOR]

Objective: Atherosclerosis, a chronic inflammatory disease, arises from metabolic disorders and is driven by inappropriate recruitment and proliferation of monocytes / macrophages and vascular smooth-muscle-cells. The receptor for the urokinase-type plasminogen activator (uPAR, Plaur) regulates the proteolytic activation of plasminogen. It is also a coactivator of integrins and facilitates leukocyte-endothelial interactions and vascular smooth-muscle-cell migration. The role of uPAR in atherogenesis remains elusive., Methods and Results: We generated C57Bl6/J low-density lipoprotein receptor (LDL) and uPAR double knockout (uPAR-/-/LDLR-/-) mice to test the role of uPAR in two distinct atherosclerosis models. In LDLR-/- mice, hepatic overexpression following hydrodynamic transfection of soluble uPAR that competes with endogenous membrane-bound uPAR was performed as an interventional strategy. Aortic root atherosclerotic lesions induced by feeding a high-fat diet were smaller and comprised less macrophages and vascular smooth-muscle-cells in double knockout mice and animals overexpressing soluble uPAR when compared to controls. In contrast, lesion size, lipid-, macrophage-, and vascular smooth muscle cell content of guide-wire-induced intima lesions in the carotid artery were not affected by uPAR deficiency. Adhesion of uPAR-/--macrophages to TNFα-stimulated endothelial cells was decreased in vitro accompanied by reduced VCAM-1 expression on primary endothelial cells. Hepatic overexpression of soluble full-length murine uPAR in LDLR-/- mice led to a reduction of diet-induced atherosclerotic lesion formation and monocyte recruitment into plaques. Ex vivo incubation with soluble uPAR protein also inhibited adhesion of macrophages to TNFα-stimulated endothelial cells in vitro., Conclusion: uPAR-deficiency as well as competitive soluble uPAR reduced diet-promoted but not guide-wire induced atherosclerotic lesions in mice by preventing monocyte recruitment and vascular smooth-muscle-cell infiltration. Soluble uPAR may represent a therapeutic tool for the modulation of hyperlipidemia-associated atherosclerotic lesion formation.