Endothelial Dysfunction and Vascular Inflammation

内皮功能障碍和血管炎症

基本信息

批准号：
7733531
负责人：
ROBERT L DANNER
金额：
$ 7.73万
依托单位：
CLINICAL CENTER
依托单位国家：
美国
项目类别：
财政年份：
资助国家：
美国
起止时间：
至
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/7733531
关键词：
2,4-thiazolidinedione Acetates Acute Affect Anti-Inflammatory Agents Anti-inflammatory Antibiotics Appearance Arginine Ascaridil Atherosclerosis Binding Sites Blood Blood Circulation Blood Platelets Blood Vessels CDKN1A gene Calcium Cardiovascular system Cause of Death Cell Line Cells Cessation of life Chronic Clinical Trials Condition Constriction procedure Development Down-Regulation Drug usage Endothelial Cells Endothelium Failure Feedback Functional disorder Genomics Human Hybrids Hypotension Illness Days Immune system Infection Inflammation Inflammatory Injury Intensive Care Units Investigation Ionophores Knowledge Link MAP Kinase Gene MAPK14 gene Mediator of activation protein Messenger RNA Metabolism Mutation Myristates NADP Nitric Oxide Non-Insulin-Dependent Diabetes Mellitus Nuclear Receptors Output Oxidants PPAR gamma Pathologic Patients Peroxisome Proliferator-Activated Receptors Phenotype Phorbol Phorbols Play Production Reactive Oxygen Species Refractory Relaxation Rest Risk Role Sepsis Septic Shock Series Shock Sickle Cell Anemia Signal Pathway Signal Transduction Pathway Site Stress Superoxide Dismutase Superoxides Supportive care Survivors Testing Thiazolidinediones Transfection U937 Cells Umbilical vein United States Vasoconstrictor Agents Work base human NOS3 protein human PLK1 protein inhibitor/antagonist mecarzole myristate oncoprotein p21 prevent promoter response sensor vascular inflammation

项目摘要

Transfection of monoblastoid U937 cells with human eNOS (endothelial nitric oxide synthase) resulted in a cell line that produces nitric oxide (NO) in response to a calcium ionophore, but little or no NO in the resting state (Blood, 1997). However, after differentiation with phorbol-12-acetate-13-myristate, eNOS expressing cells produced increased amounts of both TNFalpha and reactive oxygen species by mechanisms that were independent of NO. Neither N-omega-methyl-L-arginine, a NOS inhibitor, nor mutation of the L-arginine binding site of eNOS (rendering it incapable of producing nitric oxide) blocked the ability of eNOS to upregulate TNFalpha (J Biol Chem, 2000). Conversely, co-transfection with superoxide dismutase or deletion of the NADPH binding site of eNOS completely prevented eNOS from upregulating TNFalpha production. Ultimately, superoxide produced by eNOS was shown to upregulate TNFalpha via Erk1/2 activation (J Biol Chem, 2001). This series of investigations demonstrated that while high output NO from inducible NOS can enhance inflammation (J Immunol, 1994; J Biol Chem, 1997), dysregulation of eNOS can also contribute to an inflammatory phenotype through reactive oxygen species-based signal transduction pathways. Sp1 was shown to act as a sensor of high-output NO, down regulating eNOS in endothelial cells via a proximal Sp promoter-binding site (J Biol Chem, 2003). This work defined a key negative feedback mechanism that suppresses eNOS and reinforces vascular dysfunction in septic shock. Sickle cell disease was characterized by oxidant and inflammatory stress in the vasculature, even in the absence of an acute crisis (Blood, 2004). Circulatory stress in sickle cell disease was associated with abnormalities of arginine metabolism in platelets (Circulation, 2007), further characterizing this condition as an NO-deficient state. Anti-proliferative effects of NO that prevent the development of a chronic vascular injury phenotype was linked to p38 MAPK activation and p21 mRNA stabilization with subsequent down regulation of polo-like kinase 1 through a CDE/CHR proximal promoter site (BMC Genomics, 2005; J Biol Chem, 2006). This and later investigations (see below) indicate that p38 MAPK may play a protective role in the vasculature under some conditions. Both NO and peroxisome proliferator-activated receptors (PPARs) protect the endothelium and regulate its function. Therefore, we tested for crosstalk between these signaling pathways using human umbilical vein and hybrid EA.hy926 endothelial cells (FASEB J, 2007). PPARgamma was activated by NO through a p38 MAPK dependent signal transduction pathway. This crosstalk mechanism may contribute to the anti-inflammatory and cytoprotective effects of NO in the vasculature. Recent work has shown that p38 MAPK is also necessary for the optimal activation of PPAR gamma by thiazolidinediones (TZDs), drugs used in the treatment of type 2 diabetes mellitus. TZDs, like homeostatic NO production, reduce markers of cardiovascular inflammation. Collectively, these findings suggest that PPAR gamma as well as other nuclear receptors with transrepression activity may be useful targets for reducing endothelial dysfunction and vascular inflammation in sepsis and septic shock.

用人 eNOS（内皮一氧化氮合酶）转染单母细胞 U937 细胞，产生了一种细胞系，该细胞系响应钙离子载体产生一氧化氮（NO），但在静息状态下很少或不产生 NO（Blood，1997）。然而，在用佛波醇12-乙酸酯-13-肉豆蔻酸酯分化后，表达eNOS的细胞通过独立于NO的机制产生增加量的TNFα和活性氧。 N-omega-甲基-L-精氨酸（一种 NOS 抑制剂）和 eNOS 的 L-精氨酸结合位点的突变（使其无法产生一氧化氮）都不会阻止 eNOS 上调 TNFα 的能力（J Biol Chem，2000）。相反，与超氧化物歧化酶共转染或删除 eNOS 的 NADPH 结合位点完全阻止了 eNOS 上调 TNFα 的产生。最终，eNOS 产生的超氧化物被证明可以通过 Erk1/2 激活上调 TNFα（J Biol Chem，2001）。这一系列的研究表明，虽然诱导型 NOS 的高输出 NO 可以增强炎症（J Nutrition，1994；J Biol Chem，1997），但 eNOS 的失调也可以通过基于活性氧的信号转导途径导致炎症表型。 Sp1 被证明可作为高输出 NO 的传感器，通过近端 Sp 启动子结合位点下调内皮细胞中的 eNOS（J Biol Chem，2003）。这项工作定义了一种关键的负反馈机制，可抑制 eNOS 并强化感染性休克中的血管功能障碍。镰状细胞病的特征是脉管系统中的氧化和炎症应激，即使没有急性危象也是如此（Blood，2004）。镰状细胞病中的循环应激与血小板中精氨酸代谢异常有关（Circulation，2007），进一步将这种情况描述为 NO 缺乏状态。 NO 预防慢性血管损伤表型发展的抗增殖作用与 p38 MAPK 激活和 p21 mRNA 稳定有关，随后通过 CDE/CHR 近端启动子位点下调 Polo 样激酶 1（BMC Genomics，2005；生物化学杂志，2006）。这项研究和后来的研究（见下文）表明，在某些条件下，p38 MAPK 可能在脉管系统中发挥保护作用。 NO 和过氧化物酶体增殖物激活受体 (PPAR) 均可保护内皮并调节其功能。因此，我们使用人脐静脉和杂交 EA.hy926 内皮细胞测试了这些信号传导途径之间的串扰（FASEB J，2007）。 PPARγ 通过 p38 MAPK 依赖性信号转导途径被 NO 激活。这种串扰机制可能有助于血管系统中 NO 的抗炎和细胞保护作用。最近的研究表明，p38 MAPK 对于噻唑烷二酮类 (TZD)（用于治疗 2 型糖尿病的药物）最佳激活 PPAR γ 也是必需的。 TZD 与稳态 NO 产生一样，可以减少心血管炎症标志物。总的来说，这些发现表明 PPAR γ 以及其他具有反式阻抑活性的核受体可能是减少脓毒症和脓毒性休克中内皮功能障碍和血管炎症的有用靶标。