Use this URL to cite or link to this record in EThOS: http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.502000
Title: The role of AMP-activated protein kinase in endothelial VEGF signalling
Author: Reihill, James Anthony
Awarding Body: University of Glasgow
Current Institution: University of Glasgow
Date of Award: 2009
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Abstract:
The endothelium acts to maintain vascular homeostasis, including the regulation of vascular tone, blood fluidity and coagulation. Endothelial dysfunction, a condition largely characterised by reduced NO bioavailability, is an important feature associated with the aetiology of several pathophysiological disorders including type 2 diabetes and cardiovascular disease. AMPK is the downstream component of a protein kinase cascade important in the regulation of cellular and whole body metabolism. AMPK has been demonstrated to mediate a number of physiological responses in the endothelium, including the stimulation of eNOS phosphorylation and NO synthesis; and as such AMPK represents a therapeutic target in the dysfunctional endothelium. VEGF has been established as the prime angiogenic molecule during development, adult physiology and pathology. VEGF stimulates NO production, proposed to be a result of phosphorylation of Ser-1177 on eNOS, a residue also phosphorylated upon AMPK activation in cultured endothelial cells. The present study, utilising HAEC as a model, provides the first demonstration that AMPK is activated by physiological concentrations of VEGF; and furthermore, partially mediates VEGF-stimulated phosphorylation of eNOS on Ser-1177 and subsequent NO production. In addition, the present investigation demonstrates that the upstream AMPK kinase CaMKK is responsible for these VEGF-mediated effects. VEGF is known to increase intracellular calcium levels in endothelial cells via the generation of DAG and IP3. DAG increases Ca2+ influx through a family of non-selective cation channels, whereas IP3 promotes the release of Ca2+ from intracellular stores. High potassium-induced depolarisation, which reduces the driving force for Ca2+ entry through non-selective cation channels in endothelial cells, abolished VEGF-mediated AMPK activation, whereas the IP3 receptor blocker 2-APB was without effect. Exposure of HAEC to a DAG mimetic (OAG) also stimulated AMPK, an effect which was sensitive to the CaMKK inhibitor STO-609 and high potassium induced depolarization. The functional effects of VEGF-stimulated AMPK were also assessed in HAEC. Ablation of AMPK abrogated VEGF-stimulated HAEC migration and proliferation, two key features of the angiogenic process. While AMPK was necessary for VEGF-stimulated endothelial cell proliferation direct activation of the kinase was insufficient to induce this process. AICAR-stimulated AMPK activation has been demonstrated to stimulate fatty acid oxidation in endothelial cells. However, exposure of HAEC to VEGF did not alter fatty acid oxidation in the present study. Together, the current investigation suggests that a VEGF-Ca2+-CaMKK-AMPK-eNOS- NO pathway is present in HAEC, and furthermore, that AMPK is required, albeit insufficient, for the VEGF-stimulated angiogenic response.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID: uk.bl.ethos.502000  DOI: Not available
Keywords: QD Chemistry ; QP Physiology
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