- Email: [email protected]
VEGF resistance and its role in cardiovascular disease
316
Abstracts
(CARIM), Department of Pharmacology, Maastricht, The Netherlands E-mail address: [email protected] Oxidative stress has been suggested for decades as a key mechanism of many human pathologies, ranging from chronic inflammation to carcinogenesis and ischemia–reperfusion damage, to name a few. None of these hypotheses has reached clinical relevance in particular for chronic disease states, where antioxidants have been unsuccessful as therapeutic approach. Is the oxidative stress hypothesis wrong? It may be too early to dismiss it. Instead of using antioxidants that will interfere also with beneficial effects of reactive oxygen species (ROS) a more targeted and more successful approach may be to identify the enzymatic sources of oxidative stress and specifically interfere with them. Amongst many possible such sources NADPH oxidases (NOX) stand out. They represent the only known enzyme family that forms ROS as their sole enzymatic function; all other known sources form ROS as a side-product or subsequent to uncoupling processes. We have recently identified the NOX4 isoform as a major source of ROS in ischemic stroke and essential for both breakdown of the blood brain barrier and neurodegeneration. Mice in which the NADPH binding site essential for activity was deleted were protected from both transient and permanent ischemic damage. Since NOX4 is expressed in endothelial and vascular smooth muscle cells a primary vascular or neuronal mechanism was possible. We therefore tested other models of neurodegeneration as well as other models of ischemia–reperfusion damage. Traumatic brain injury is a model of primarily non-vascular driven neurodegeneration and here NOX4 mice were not protected. Mice subjected to ischemia–reperfusion injury of the lung or the heart were not or only partially protected, respectively. The reduction in infarct size observed in NOX4−/− mice was moderate compared to that in different forms of ischemic stroke and not accompanied by a similar functional protection. Thus induction of NOX4 and resulting oxidative stress appears neither to be a general mechanism of neurotoxicity nor of ischemia-reperfusion damage, despite the fact that NOX4 is consistently induced in ischemia and hypoxia. With respect to this vascular phenotype, in particular in more long-term models, a complicating factor may be a possible beneficial role of NOX4 in angiogenesis as has been suggested. Indeed in more artificial in vitro models of angiogenesis, NOX4 was essential e.g. for endothelial tube formation. However, this did not translate into more in vivo situations. Thus, for now ischemic stroke appears to be the ischemic condition with the most significant involvement of any NOX. Further studies using both pharmacological and cell-specific KO experiments are under way to elucidate the underlying mechanisms. doi:10.1016/j.vph.2011.08.030
L.4.3 VEGF resistance and its role in cardiovascular disease J. Waltenberger, E. Pardali Department of Cardiology and Angiology, University of Münster, Münster, Germany E-mail address: [email protected] (J. Waltenberger) VEGF is an important vascular growth factor for both development and maintenance of the vascular system. VEGF targets mostly endothelial cells and monocytes via VEGF receptors. Monocytes are circulating leukocytes participating in pathological processes of the cardiovascular system, namely during atherogenesis, i.e. atherosclerotic plaque development, as well as in repair processes such as arteriogenesis, i.e. collateral vessel growth. Monocytes hereby act as bioreactors to release bioactive molecules that contribute to local inflammation and growth. One of the crucial properties of monocyte function is their chemotactic responsiveness towards growth factors
and chemokines such as MCP-1 or VEGF, which allows migration towards sites where they are needed, or alternatively, to prevent that they are trapped where they can be harmful. We have previously established the molecular basis of VEGFinduced chemotaxis. Using ex-vivo analysis of freshly isolated monocytes from patients with metabolic and cardiovascular diseases, we previously demonstrate that pathological conditions such as diabetes mellitus (DM), hypercholesterolemia (HC) or smoking negatively affect the function of VEGF in monocytes. This VEGF resistance is based on a signal transduction defect. In case of isolated CD14+ monocytes, diabetes mellitus resulted in the unspecific preactivation of several signalling pathways including PI-3-kinase, AKT and Erk1/2. This preactivation is mediated by ROS, involves RAGE and PTPases and may be regarded as the molecular equivalent of VEGF resistance in diabetes mellitus. Smoking- and HC-related VEGFresistance can be reverted in vivo by using high doses of either antioxidants (vitamin C) or statins. Based on these and other unpublished data, monocytes appear to be suitable living biosensors for identifying VEGF resistance and for assessing the integrity of vascular cell function. Other novel data will be shown to demonstrate novel aspects of monocyte dysfunction and their relation to the integrity of the cardiovascular system. References Waltenberger, J., Lange, J., Kranz, A., 2000. Circulation 102, 185–190. Tchaikovski, V., Fellbrich, G., Waltenberger, J., 2008. ATVB 28, 322–328. Tchaikovski, et al., 2009. Circulation 120, 150–159. Dunaeva, et al., 2010. Basic. Res. Cardiol 105, 61–71. Czepluch, et al., 2011. Thromb. Haemost. 105, 122–130. Olieslagers, et al., 2011 May 9. Cardiovasc. Res. doi:10.1016/j.vph.2011.08.031
L.4.4 High intraluminal pressure induces inflammation Jaye Chin-Dusting Baker IDI Heart and Diabetes Institute, Melbourne, Australia 3004 E-mail address: [email protected] Hypertension causes more deaths and disease globally than any other biomedical risk. Despite the widespread use of antihypertensive drugs, optimal blood pressure (BP) control is achieved in less than 50% of patients. Worse, 20–30% have treatment resistant hypertension. Alterations of the immune system are implicated in the genesis of hypertension. No study has examined whether, paradoxically, hypertension contributes to the genesis of inflammation. Increases in blood pressure can lead to turbulent and low shear stress in the vasculature resulting in endothelial dysfunction. We show that high intraluminal pressure also increases leukocyte to endothelium adhesion and explore the pathways involved in this pressure-induced inflammation. Using a specialised pressure myograph we recorded leukocyte adhesion to endothelium intact vessels in real time. Carotid arteries pressurised for 1 h demonstrate pressure dependent increases in leukocyte adhesion (60, 80, 120 mm Hg= 21 ± 3, 36 ± 6, 93 ± 14 leukocytes/field respectively, n = 3–7; P < 0.05) suggesting a rapid inflammatory response with increased pressure. Furthermore we demonstrate that this pressureinduced response was reduced with the use of the non specific cholesterol acceptor (methyl-β-cylcodextrin treatment: control 93± 14 vs treated 47 ± 13; n = 4–7; P < 0.05), NADPH oxidase (apocynin treatment: control 93± 14 vs treated 25± 3; n = 5–7; P < 0.05) and mitochondrial permeability inhibition (cyclosporin A treatment: control 93± 14 vs treated 50± 14; n = 4–7; P < 0.05) but not influenced by
Abstracts
(CARIM), Department of Pharmacology, Maastricht, The Netherlands E-mail address: [email protected] Oxidative stress has been suggested for decades as a key mechanism of many human pathologies, ranging from chronic inflammation to carcinogenesis and ischemia–reperfusion damage, to name a few. None of these hypotheses has reached clinical relevance in particular for chronic disease states, where antioxidants have been unsuccessful as therapeutic approach. Is the oxidative stress hypothesis wrong? It may be too early to dismiss it. Instead of using antioxidants that will interfere also with beneficial effects of reactive oxygen species (ROS) a more targeted and more successful approach may be to identify the enzymatic sources of oxidative stress and specifically interfere with them. Amongst many possible such sources NADPH oxidases (NOX) stand out. They represent the only known enzyme family that forms ROS as their sole enzymatic function; all other known sources form ROS as a side-product or subsequent to uncoupling processes. We have recently identified the NOX4 isoform as a major source of ROS in ischemic stroke and essential for both breakdown of the blood brain barrier and neurodegeneration. Mice in which the NADPH binding site essential for activity was deleted were protected from both transient and permanent ischemic damage. Since NOX4 is expressed in endothelial and vascular smooth muscle cells a primary vascular or neuronal mechanism was possible. We therefore tested other models of neurodegeneration as well as other models of ischemia–reperfusion damage. Traumatic brain injury is a model of primarily non-vascular driven neurodegeneration and here NOX4 mice were not protected. Mice subjected to ischemia–reperfusion injury of the lung or the heart were not or only partially protected, respectively. The reduction in infarct size observed in NOX4−/− mice was moderate compared to that in different forms of ischemic stroke and not accompanied by a similar functional protection. Thus induction of NOX4 and resulting oxidative stress appears neither to be a general mechanism of neurotoxicity nor of ischemia-reperfusion damage, despite the fact that NOX4 is consistently induced in ischemia and hypoxia. With respect to this vascular phenotype, in particular in more long-term models, a complicating factor may be a possible beneficial role of NOX4 in angiogenesis as has been suggested. Indeed in more artificial in vitro models of angiogenesis, NOX4 was essential e.g. for endothelial tube formation. However, this did not translate into more in vivo situations. Thus, for now ischemic stroke appears to be the ischemic condition with the most significant involvement of any NOX. Further studies using both pharmacological and cell-specific KO experiments are under way to elucidate the underlying mechanisms. doi:10.1016/j.vph.2011.08.030
L.4.3 VEGF resistance and its role in cardiovascular disease J. Waltenberger, E. Pardali Department of Cardiology and Angiology, University of Münster, Münster, Germany E-mail address: [email protected] (J. Waltenberger) VEGF is an important vascular growth factor for both development and maintenance of the vascular system. VEGF targets mostly endothelial cells and monocytes via VEGF receptors. Monocytes are circulating leukocytes participating in pathological processes of the cardiovascular system, namely during atherogenesis, i.e. atherosclerotic plaque development, as well as in repair processes such as arteriogenesis, i.e. collateral vessel growth. Monocytes hereby act as bioreactors to release bioactive molecules that contribute to local inflammation and growth. One of the crucial properties of monocyte function is their chemotactic responsiveness towards growth factors
and chemokines such as MCP-1 or VEGF, which allows migration towards sites where they are needed, or alternatively, to prevent that they are trapped where they can be harmful. We have previously established the molecular basis of VEGFinduced chemotaxis. Using ex-vivo analysis of freshly isolated monocytes from patients with metabolic and cardiovascular diseases, we previously demonstrate that pathological conditions such as diabetes mellitus (DM), hypercholesterolemia (HC) or smoking negatively affect the function of VEGF in monocytes. This VEGF resistance is based on a signal transduction defect. In case of isolated CD14+ monocytes, diabetes mellitus resulted in the unspecific preactivation of several signalling pathways including PI-3-kinase, AKT and Erk1/2. This preactivation is mediated by ROS, involves RAGE and PTPases and may be regarded as the molecular equivalent of VEGF resistance in diabetes mellitus. Smoking- and HC-related VEGFresistance can be reverted in vivo by using high doses of either antioxidants (vitamin C) or statins. Based on these and other unpublished data, monocytes appear to be suitable living biosensors for identifying VEGF resistance and for assessing the integrity of vascular cell function. Other novel data will be shown to demonstrate novel aspects of monocyte dysfunction and their relation to the integrity of the cardiovascular system. References Waltenberger, J., Lange, J., Kranz, A., 2000. Circulation 102, 185–190. Tchaikovski, V., Fellbrich, G., Waltenberger, J., 2008. ATVB 28, 322–328. Tchaikovski, et al., 2009. Circulation 120, 150–159. Dunaeva, et al., 2010. Basic. Res. Cardiol 105, 61–71. Czepluch, et al., 2011. Thromb. Haemost. 105, 122–130. Olieslagers, et al., 2011 May 9. Cardiovasc. Res. doi:10.1016/j.vph.2011.08.031
L.4.4 High intraluminal pressure induces inflammation Jaye Chin-Dusting Baker IDI Heart and Diabetes Institute, Melbourne, Australia 3004 E-mail address: [email protected] Hypertension causes more deaths and disease globally than any other biomedical risk. Despite the widespread use of antihypertensive drugs, optimal blood pressure (BP) control is achieved in less than 50% of patients. Worse, 20–30% have treatment resistant hypertension. Alterations of the immune system are implicated in the genesis of hypertension. No study has examined whether, paradoxically, hypertension contributes to the genesis of inflammation. Increases in blood pressure can lead to turbulent and low shear stress in the vasculature resulting in endothelial dysfunction. We show that high intraluminal pressure also increases leukocyte to endothelium adhesion and explore the pathways involved in this pressure-induced inflammation. Using a specialised pressure myograph we recorded leukocyte adhesion to endothelium intact vessels in real time. Carotid arteries pressurised for 1 h demonstrate pressure dependent increases in leukocyte adhesion (60, 80, 120 mm Hg= 21 ± 3, 36 ± 6, 93 ± 14 leukocytes/field respectively, n = 3–7; P < 0.05) suggesting a rapid inflammatory response with increased pressure. Furthermore we demonstrate that this pressureinduced response was reduced with the use of the non specific cholesterol acceptor (methyl-β-cylcodextrin treatment: control 93± 14 vs treated 47 ± 13; n = 4–7; P < 0.05), NADPH oxidase (apocynin treatment: control 93± 14 vs treated 25± 3; n = 5–7; P < 0.05) and mitochondrial permeability inhibition (cyclosporin A treatment: control 93± 14 vs treated 50± 14; n = 4–7; P < 0.05) but not influenced by