Early vessel destabilization mediated by Angiopoietin-2 and subsequent vessel maturation via Angiopoietin-1 induce functional neovasculature after ischemia.

We assessed whether Angiopoietin-2 (Ang2), a Tie2 ligand and partial antagonist of Angiopoietin-1 (Ang1), is required for early vessel destabilization during postischemic angiogenesis, when combined with vascular growth factors. In vitro, matrigel co-cultures assessed endothelial-cell tube formation and pericyte recruitment after stimulation of VEGF-A, Apelin (APLN), Ang1 with or without Ang2. In a murine hindlimb ischemia model, adeno-associated virus (rAAV, 3×10(12) virusparticles) transduction of VEGF-A, APLN and Ang1 with or without Ang2 (continuous or early expression d0-3) was performed intramuscularly (d-14). Femoral artery ligation was performed at d0, followed by laser doppler perfusion meassurements (LDI) 7 and 14. At d7 (early timepoint) and d14 (late timepoint), histological analysis of capillary/muscle fiber ratio (CMF-R, PECAM-1) and pericyte/capillary ratio (PC-R, NG2) was performed. In vitro, VEGF-A, APLN and Ang1 induced ring formation, but only APLN and Ang1 recruited pericytes. Ang2 did not affect tube formation by APLN, but reduced pericyte recruitment after APLN or Ang1 overexpression. In vivo, rAAV.VEGF-A did not alter LDI-perfusion at d14, consistent with an impaired PC-R despite a rise in CMF-R. rAAV.APLN improved perfusion at d14, with or without continuous Ang2, increasing CMF-R and PC-R. rAAV.Ang1 improved perfusion at d14, when combined with rAAV.Ang2 (d0-3), accompanied by an increased CMF-R and PC-R. The combination of early vessel destabilization (Ang2 d0-3) and continuous Ang1 overexpression improves hindlimb perfusion, pointing to the importance of early vessel destabilization and subsequent vessel maturation for enhanced therapeutic neovascularization

ACE-inhibition prevents postischemic coronary leukocyte adhesion and leukocyte-dependent reperfusion injury

Objective: Polymorphonuclear leukocytes (PMN), retained in the microvascular bed, can contribute to postischemic myocardial reperfusion injury. Since a beneficial effect of ACE-inhibition on reperfusion injury has been reported, we investigated the impact of cilazaprilat on PMN dependent reperfusion injury in isolated guinea pig hearts. Methods: Hearts (n=5 per group) were subjected to 15 min of ischemia. Immediately thereafter, a bolus of PMN was injected into the coronary system. External heart work (EHW) and total cardiac nitric oxide release were measured. For microscopic evaluation, hearts received rhodamine 6G labelled PMN after ischemia, were arrested 5 min later and further perfused with FITC dextran (0.1%). Localization of retained PMN was assessed by fluorescence microscopy. Leukocyte activation was studied by FACS analysis of the adhesion molecule CD11b before and after coronary passage of the PMN. The ACE-inhibitor cilazaprilat (Cila, 2 μM) and the NO-synthase inhibitor nitro-L-arginine (NOLAG, 10 μM) were used to modulate nitric oxide formation of the heart. Results: Postischemic EHW recovered to 67±5% (controls) and 64±6% (Cila) of the preischemic value. Addition of PMN severely depressed recovery of EHW (39±2%) and NO release (39±6% of the preischemic value). Simultaneously, ischemia led to a substantial increase in postcapillary PMN adhesion (from 21±5 to 172±27 PMN/mm² surface) and CD11b-expression of the recovered PMN (3-fold). Cila attenuated postischemic PMN adhesion (83±52 PMN/mm²) and activation of PMN, whereas it improved recovery of work performance (64±4%) and NO release (65±4%) in the presence of PMN. Conversely, NOLAG increased PMN adhesion (284±40 PMN/mm²) and myocardial injury. We conclude that ACE-inhibition prevents leukocyte dependent reperfusion injury mainly by inhibition of postcapillary leukocyte adhesion. The effect may be mediated by NO, given the proadhesive effect of NOLAG

Interdependence of Angiogenesis and Arteriogenesis in Development and Disease

The structure of arterial networks is optimized to allow efficient flow delivery to metabolically active tissues. Optimization of flow delivery is a continuous process involving synchronization of the structure and function of the microcirculation with the upstream arterial network. Risk factors for ischemic cardiovascular diseases, such as diabetes mellitus and hyperlipidemia, adversely affect endothelial function, induce capillary regression, and disrupt the micro- to macrocirculation cross-talk. We provide evidence showing that this loss of synchronization reduces arterial collateral network recruitment upon arterial stenosis, and the long-term clinical outcome of current revascularization strategies in these patient cohorts. We describe mechanisms and signals contributing to synchronized growth of micro- and macrocirculation in development and upon ischemic challenges in the adult organism and identify potential therapeutic targets. We conclude that a long-term successful revascularization strategy should aim at both removing obstructions in the proximal part of the arterial tree and restoring “bottom-up” vascular communication

Tumor Necrosis Factor-α Contributes to Ischemia- and Reperfusion-Induced Endothelial Activation in Isolated Hearts

During myocardial reperfusion, polymorphonuclear neutrophil (PMN) adhesion involving the intercellular adhesion molecule-1 (ICAM-1) may lead to aggravation and prolongation of reperfusion injury. We studied the role of early tumor necrosis factor-α (TNF-α) cleavage and nuclear factor-κB (NF-κB) activation on ICAM-1 expression and venular adhesion of PMN in isolated hearts after ischemia (15 minutes) and reperfusion (30 to 480 minutes). NF-κB activation (electromobility shift assay) was found after 30 minutes of reperfusion and up to 240 minutes. ICAM-1 mRNA, assessed by Northern blot, increased during the same interval. Functional effect of newly synthesized adhesion molecules was found by quantification (in situ fluorescence microscopy) of PMN, given as bolus after ischemia, which became adherent to small coronary venules (10 to 50 mm in diameter). After 480 minutes of reperfusion, ICAM-1–dependent PMN adhesion increased 2.5-fold compared with PMN adhesion obtained during acute reperfusion. To study the influence of NF-κB on PMN adhesion, we inhibited NF-κB activation by transfection of NF-κB decoy oligonucleotides into isolated hearts using HJV-liposomes. Decoy NF-κB but not control oligonucleotides blocked ICAM-1 upregulation and inhibited the subacute increase in PMN adhesion. Similar effects were obtained using BB 1101 (10 μg), an inhibitor of TNF-α cleavage enzyme. These data suggest that ischemia and reperfusion in isolated hearts cause liberation of TNF-α, activation of NF-κB, and upregulation of ICAM-1, an adhesion molecule involved in inflammatory response after ischemia and reperfusion

Endothelial preconditioning by transient oxidative stress reduces inflammatory responses of cultured endothelial cells to TNF-α

Brief episodes of ischemia can render an organ resistant to subsequent severe ischemia. This ‘ischemic preconditioning’ is ascribed to various mechanisms, including oxidative stress. We investigated whether preconditioning exists on an endothelial level. Human umbilical vein endothelial cells (HUVECs) were transiently confronted with oxidative stress (1 mM H2O2, 5 min). Adhesion molecules ICAM-1 and E-selectin and release of cytokines IL-6 and IL-8 to subsequent stimulation with TNF-α (2.5 ng/ml, 4 h) were measured (flow cytometry and immunoassay), as were nuclear translocation of the transcription factor NFkB (Western blotting, confocal microscopy) and redox status of HUVECs (quantification of glutathione by HPLC). TNF-α elevated IL-6 in the cell supernatant from 8.8 ± 1 to 41 ± 3 pg/ml and IL-8 from 0.5 ± 0.03 to 3 ± 0.2 ng/ml. ICAM-1 was increased threefold and E-selectin rose eightfold. Oxidative stress (decrease of glutathione by 50%) reduced post-TNF-α levels of IL-6 to 14 ± 3 and IL-8 to 1 ± 0.2; the rise of ICAM-1 was completely blocked and E-selectin was only doubled. The anti-inflammatory effects of preconditioning via oxidative stress were paralleled by reduction of the translocation of NFkB on stimulation with TNF-α, and antagonized by the intracellular radical scavenger N-acetylcysteine. ‘Anti-inflammatory preconditioning’ of endothelial cells by oxidative stress may account for the inhibitory effects of preconditioning on leukocyte adhesion in vivo

Disparate effects of adhesion and degranulation of platelets on myocardial and coronary function in postischaemic hearts

Beside the major effect of acute thrombus formation, little is known about the interaction of platelets with the coronary endothelium in an ischaemia–reperfusion situation. The present study was designed to investigate, separately, the consequences of platelet adhesion and degranulation during myocardial reperfusion. Methods: Isolated guinea pig hearts perfused with Krebs–Henseleit buffer and performing pressure–volume work were used. We infringed myocardial function by imposing ischaemia (20 min of low-flow perfusion with 1 ml/min and 10 min of global ischaemia) and reperfusion (15 min with 5 ml/min). During low-flow perfusion, the coronary endothelium was stimulated by thrombin before and during infusion of a bolus: 108 washed human platelets±the Arg–Gly–Asp (RGD) analogon lamifiban, the supernatant of 108 thrombin-stimulated platelets, fibrinogen (2 μM), lamifiban (2 μM) or Tyrode’s solution (control group). The parameter external heart work (EHW), determined pre- and postischaemically, served as criterion for recovery of myocardial function. Additionally, the formation of capillary transudate was measured during the reperfusion phase to assess coronary permeability. Coronary perfusion pressure was monitored continuously and myocardial production of lactate and consumption of pyruvate were measured. Electron microscopy of hearts was performed after platelet application to verify platelet adhesion in the coronary system. Results: Recovery of EHW by hearts without platelet application was 64±3% and was significantly reduced to 49±5% by platelet infusion (n=8 each). Infusion of supernatant of thrombin-stimulated platelets did not impair recovery of heart work. In the reperfusion phase (6th–10th min), hearts that either had received platelets or supernatant of platelets exhibited a significantly reduced production of capillary transudate (70 μl/min vs. 180 μl/min for the controls). Intracoronary bolus application of fibrinogen or lamifiban also reduced coronary leak. Coronary perfusion pressure and metabolic parameters were not statistically different between the groups at any time. Conclusions: Platelet adhesion to the coronary endothelium in a situation of myocardial ischaemia impairs cardiac recovery, whereas constituents released by platelets may have beneficial effects on the integrity of the coronary endothelium. In particular, fibrinogen seems to contribute to the permeability reducing effect, possibly by interaction with endothelial receptors recognising the RGD sequence

Transcutaneous aortic valve replacement with the Edwards SAPIEN XT and Medtronic CoreValve prosthesis under fluoroscopic guidance and local anaesthesia only

Objective: To assess the feasibility of transcatheter aortic valve implantation (TAVI; Medtronic CoreValve and Edwards SAPIEN XT) under local anaesthesia with only mild analgesic medication and fluoroscopic guidance. Methods: 461 patients underwent TAVI under local anaesthesia with lidocaine. The procedure was performed successfully in 459 of the cases. All patients were also treated with piritramide, metoclopramide hydrochloride and 62 mg dimenhydrinate. Monitoring consisted of a six-electrode, virtual 12-lead ECG, pulse oximetry, and invasive arterial pressure measurement. There was no continuous surveillance by an anaesthesiologist. Results: There was no need for conversion to general anaesthesia except in four patients who required cardiopulmonary resuscitation. Conscious sedation with intravenous administration of midazolam for agitation or inotropic medication for prolonged hypotension was necessary in only seven of the 461 patients. The combined safety end point according to the Valve Academic Research Consortium consensus document was reached in 12.6%. Conclusions: Our results show that TAVI performed under local anaesthesia with only mild analgesic medication and under fluoroscopic guidance is feasible, with good outcome comparable to published data

Transcatheter Aortic Valve Implantation in Dialysis Patients

Background/Aims: Transcatheter aortic valve implantation (TAVI) has emerged as a new therapeutic option for high-risk patients. However, dialysis patients were excluded from all previous studies. The aim of this study is to compare the outcomes of TAVI for dialysis patients with those for patients with chronic kidney disease (CKD) stages 3 and 4 and to compare TAVI with open surgery in dialysis patients. Methods: Part I: comparison of 10 patients on chronic hemodialysis with 116 patients with non-dialysis-dependent CKD undergoing TAVI. Part II: comparison of transcatheter (n = 15) with open surgical (n = 24) aortic valve replacement in dialysis patients. Results: Part I: dialysis patients were significantly younger (72.3 vs. 82.0 years; p < 0.01). Hospital stay was significantly longer in dialysis patients (21.8 vs. 12.1 days; p = 0.01). Overall 30-day mortality was 3.17%, with no deaths among dialysis patients. Six-month survival rates were similar (log-rank p = 0.935). Part II: patient age was comparable (66.5 vs. 69.5 years; p = 0.42). Patients in the surgical group tended to stay longer in hospital than TAVI patients (29.5 vs. 22.5 days; p = 0.35). Conclusion: TAVI is a safe procedure in patients on chronic hemodialysis. Until new data become available, we find no compelling reason to refuse these patients TAVI. Copyright (C) 2012 S. Karger AG, Base