Poly (ADP-Ribose) synthetase in expression of endothelia

内皮损伤，以及随之而来的中性粒细胞粘附血管内膜，是心肌缺血再灌注损伤的早期病理事件。中性粒细胞向损伤组织转运的确切分子机制尚不清楚。实验证据表明，氧化应激是白细胞内皮粘附分子表达的重要信号转导事件。在初步的体外和体内研究中，我们发现氧化应激通过核酶聚核糖合成酶（PARS）介导的途径引起细胞功能障碍和损伤。在体外内皮细胞中，过氧亚硝酸盐和过氧化氢诱导DNA单链断裂并激活PARS，导致细胞高能磷酸盐耗竭，细胞功能障碍和死亡。我们发现，在大鼠和小鼠心肌缺血和再灌注期间，体内药理或遗传抑制PARS可减少梗死大小和中性粒细胞在损伤组织中的积累。本研究旨在评估再灌注依赖性氧化应激诱导内皮细胞表达p -选择素、e-选择素、细胞间粘附分子-1 （ICAM-1）和血管粘附分子-1 （VCAM-1）的分子基础，特别关注PARS通路。pars介导的内皮细胞粘附分子表达调控的病理生理学相关性将在小鼠心肌缺血再灌注损伤的体内实验模型中进行评估。具体来说，实验将在缺乏PARS基因的敲除动物的小鼠心肌缺血和再灌注模型中进行，并在窝代野生型对照中进行。为了临床研究，我们将在大鼠心肌再灌注损伤模型中研究PARS药理抑制对内皮细胞损伤和中性粒细胞积累的影响。此外，实验将在体外培养的人脐静脉内皮细胞中进行氧化应激和非氧化应激。这些体外实验是为了更准确地评估细胞内PARS在成人心肌梗死相关的内皮功能障碍中的作用。该项目的结果将为研究PARS作为内皮屏障功能调节剂的作用提供重要信息，并将有助于开发以内皮再灌注损伤为特征的心血管疾病的有效治疗干预措施。

Endothelial injury, and consequent adhesion of neutrophils to the vascular intima, is an early pathologic event in myocardial ischemia and reperfusion injury. The precise molecular mechanisms involved in neutrophil trafficking to injured tissue are not clear. Experimental evidence shows that oxidative stress is an important signal transduction event for the expression of endothelial adhesion molecules for leukocytes. In preliminary in vitro and in vivo studies we have found that oxidative stress causes cellular dysfunction and injury through a pathway mediated by the nuclear enzyme poly (ADP ribose) synthetase (PARS). In in vitro endothelial cells, peroxynitrite and hydrogen peroxide induce DNA single strand breaks and activate PARS, resulting in the depletion of cellular high-energy phosphates, cellular dysfunction and death. We have found that in vivo pharmacological or genetic inhibition of PARS reduces infract size and neutrophil accumulation into the injured tissue during myocardial ischemia and reperfusion in the rat and in the mouse. The present investigation is designed to evaluate the molecular basis of endothelial expressions of P-selectin, E-selectin, intercellular adhesion molecule-1 (ICAM-1) and vascular adhesion molecular-1 (VCAM-1) induced by reperfusion-dependent oxidant stress, with special focus on the PARS pathway. The pathophysiological relevance of a PARS-mediated regulation of endothelial expression of adhesion molecules will be evaluated in in vivo experimental rodent models of myocardial ischemia and reperfusion injury. Specifically, experiments will be performed in a murine model of myocardial ischemia and reperfusion in knock-out animals lacking the gene of PARS and in littermate wild-type controls. For clinical relevance, the effect of pharmacological inhibition of PARS on endothelial injury and neutrophil accumulation will be studied in a rat model of myocardial reperfusion injury. Furthermore, experiments will be performed in cultured human umbilical vein endothelial cells subjected to oxidative and non oxidative stress. These in vitro experiments are proposed in order to obtain a more precise assessment of the role of intracellular PARS on the adult human endothelial dysfunction relevant to the human myocardial infarction. The results of the proposed project will provide important information on the role of PARS as a modulator of the endothelial barrier function and will contribute to the development of effective therapeutic interventions of cardiovascular diseases characterized by endothelial reperfusion injury.