| Popis: |
While oxidative stress and dysregulation of Ca2+ homeostasis in vascular signalling are hallmarks of hypertension-associated vascular injury, downstream molecular mechanisms that govern reactive oxygen species (ROS)-dependent regulation of ion channels in hypertension are complex and have not yet been fully elucidated. Activation of poly(ADPribose)-polymerase 1 (PARP1) in response to oxidative stress and DNA damage leads to subsequent activation of transient receptor potential melastatin 2 (TRPM2), an ion channel that regulates Na+ and Ca2+ influx. Considering that ROS generation and Ca2+ influx are increased in hypertension we hypothesised that redox regulation of PARP1-TRPM2 may play a role in vascular injury and target organ-damage associated with hypertension. The proof of concept was tested in a TRPM2-overexpressing human embryonic kidney cell line and the relationship between ROS, PARP1 and TRPM2 was studied in vitro, in human vascular smooth muscle cells and endothelial cells. The regulatory role of PARP1 and TRPM2 on vascular function was assessed in isolated resistance arteries from LinA3 mice, a transgenic model expressing human renin gene resulting in chronic hypertension. Finally, the effects of pharmacological inhibition of PARP1 on blood pressure, target organ damage and cellular signalling were evaluated in vivo in hypertensive LinA3 mice. To our knowledge, findings from this study provide the first evidence in clinically-relevant models, that the redox-sensitive PARP1-TRPM2 pathway regulates vascular contraction in the context of hypertension. This is supported by the following findings: i) The in vitro studies demonstrated that PARP1 and TRPM2 facilitate Ang II and oxidant-dependent activation of a pro-contractile protein MLC20 and partially reduce the anti-contractile activity of MYPT1. ii) The ex vivo experiments on mesenteric resistance arteries from LinA3 mice confirmed that the PARP1-TRPM2 pathway exacerbates vascular hypercontractility of the arteries isolated from hypertensive mice. iii) In vivo inhibition of PARP1 had significant effects on renal and cardiac Akt/PKB-dependent signalling, leading to upregulation of prosurvival and anti-apoptotic proteins, effects that were independent of blood pressure lowering. Overall, the studies presented in this thesis highlight a novel pathway linking ROS to vascular signalling pathways through PARP1 and TRPM2. Dysregulation of this system, in the context of oxidative stress in hypertension, may play a role in hypertension-associated vascular injury and target organ damage. While the present studies have opened the field, further investigations to unambiguously prove the importance of the ROS-PARP1-TRPM2-Ca2+ axis in hypertension are required. |
