Transglutaminase 2 decreases ischemic insult by attenuating hypoxic signaling.

转谷氨酰胺酶 2 通过减弱缺氧信号来减少缺血性损伤。

基本信息

批准号：
7609278
负责人：
Anthony J Filiano
金额：
$ 1.78万
依托单位：
UNIVERSITY OF ROCHESTER
依托单位国家：
美国
项目类别：
财政年份：
2009
资助国家：
美国
起止时间：
2009-03-30 至 2009-05-16
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7609278
关键词：
ARNT protein Adenoviruses Affect Alteplase Animals Apoptosis Apoptotic Architecture Asses Attenuated BCL2/Adenovirus E1B 19kd Interacting Protein 3-Like Binding Biological Assay Biological Models Blood - brain barrier anatomy C57BL/6 Mouse Calcium Cause of Death Cell Death Cell Line Cell Nucleus Cells Cessation of life Condition Cultured Cells Data Development Effectiveness Environment Enzymes Event Genes Genetic Genetic Transcription Glucose Goals Green Fluorescent Proteins Guanosine Triphosphate Phosphohydrolases HIF1alpha protein Hour Human Hypoxia Hypoxia Inducible Factor Immunoblot Analysis Immunohistochemistry In Situ Infarction Infection Control Injury Ischemia Knockout Mice LacZ Genes Lactate Dehydrogenase Lactate Dehydrogenases Lead Life Ligation Luciferases Magnetic Resonance Imaging Measures Mediating Messenger RNA Modeling Mus Nerve Degeneration Neuraxis Neurites Neuroblastoma Neurons Oxygen Pathway interactions Pattern Permeability Play Point Mutation Polymerase Chain Reaction Predisposition Property Protein Overexpression Proteins Quality of life Rattus Reporter Response Elements Role Scaffolding Protein Signal Transduction Stroke Stroke Volume Techniques Testing Therapeutic Thrombolytic Therapy Time Transgenic Organisms United States Food and Drug Administration Up-Regulation Vascular Endothelial Growth Factors Weight attenuation base caspase-3 deprivation disability hypoxia inducible factor 1 improved middle cerebral artery mouse model mutant neuron loss novel post stroke protein expression response scaffold size small molecule success transcription factor transglutaminase 2 vector

项目摘要

DESCRIPTION (provided by applicant): Stroke is the third leading cause of death and a major cause of long term disability in the US. Inadequate success of current thrombolytic therapy demonstrates a critical need for new targets to improve quality of life post-stroke. Neurons adapt to decreased oxygen delivery subsequent to stroke by up-regulating genes controlled by the hypoxia inducible factor (HIF). This transcription factor is comprised of two subunits: HIF1 alpha and HIF1 beta. Recent studies show that increased HIF1 during ischemia results in upregulation of apoptotic genes. Our preliminary data indicates that Transglutaminase 2 (TG2) binds to HIF1 beta resulting in attenuation of HIF activation and decreased cell death during ischemia. Also, TG2 inhibits the upregulation of the HIF controlled proapoptotic gene Bnip3, but not VEGF. Our hypothesis is that TG2 will facilitate neuronal protection following oxygen and glucose deprivation (OGD) by attenuating apoptotic genes induced by HIF1. We propose to delineate the effects of TG2 on HIF signaling and ischemic cell death with 3 aims. 1) To test the hypothesis that TG2 is upregulated during OGD and decreases HIF-dependent pro-apoptotic signaling. We will use a neuroblastoma cell line (SH-SY5Y) that overexpresses wild type TG2 and two TG2 constructs with point mutations that differentially affect 2 unique functions (transamidating and GTPase) of the enzyme. Using a HIF signaling reporter assay and analysis of mRNA and protein expression patterns of HIF dependent genes, we will distinguish which properties of TG2 are important for regulating HIF signaling. 2) To test the hypothesis that TG2 protects neuronal cells from ischemic insult and which functions of TG2 are important. Using SH-SY5Y cells and primary cortical neurons we will measure OGD-induced cell death and upstream apoptotic events. 3) To test the hypothesis that TG2 leads to overall protection in a mouse model of stroke. We will use a permanent Middle Cerebral Artery ligation model in C57BL/6 mice that express endogenous TG2, overexpress TG2, or lack TG2. We will measure infarct volumes using T2 weighted MRI as well as identifying genes regulated by TG2 with appropriate techniques. Our preliminary data show that overexpression of TG2 decreases stroke infarct volumes by 33%. We hypothesize that TG2 will lead to overall protection against stroke.

描述（由申请人提供）：中风是美国第三大死亡原因，也是导致长期残疾的主要原因。当前溶栓治疗的不充分成功表明迫切需要新的靶标来改善中风后的生活质量。神经元通过上调由缺氧诱导因子（HIF）控制的基因来适应中风后氧输送减少。该转录因子由两个亚基组成：HIF1 α 和 HIF1 β。最近的研究表明，缺血期间 HIF1 的增加会导致凋亡基因的上调。我们的初步数据表明，转谷氨酰胺酶 2 (TG2) 与 HIF1 β 结合，导致 HIF 激活减弱并减少缺血期间的细胞死亡。此外，TG2 抑制 HIF 控制的促凋亡基因 Bnip3 的上调，但不抑制 VEGF。我们的假设是，TG2 将通过减弱 HIF1 诱导的凋亡基因来促进缺氧和缺糖 (OGD) 后的神经元保护。我们建议通过 3 个目标来描述 TG2 对 HIF 信号传导和缺血性细胞死亡的影响。 1) 检验 TG2 在 OGD 期间上调并减少 HIF 依赖性促凋亡信号传导的假设。我们将使用过度表达野生型 TG2 的神经母细胞瘤细胞系 (SH-SY5Y) 和两个具有点突变的 TG2 构建体，这些点突变会差异影响该酶的 2 个独特功能（转酰胺基化和 GTP 酶）。使用 HIF 信号报告基因测定以及 HIF 依赖性基因的 mRNA 和蛋白质表达模式分析，我们将区分 TG2 的哪些特性对于调节 HIF 信号转导很重要。 2) 检验 TG2 保护神经元细胞免受缺血性损伤的假设以及 TG2 的哪些功能很重要。使用 SH-SY5Y 细胞和原代皮质神经元，我们将测量 OGD 诱导的细胞死亡和上游凋亡事件。 3) 检验 TG2 在小鼠中风模型中产生全面保护作用的假设。我们将在表达内源性 TG2、过表达 TG2 或缺乏 TG2 的 C57BL/6 小鼠中使用永久性大脑中动脉结扎模型。我们将使用 T2 加权 MRI 测量梗塞体积，并使用适当的技术识别 TG2 调节的基因。我们的初步数据显示，TG2 的过度表达可使中风梗塞体积减少 33%。我们假设 TG2 将带来针对中风的全面保护。