Acute Renal Failure: An Endotoxin Hyper-Responsive State

急性肾衰竭：内毒素高反应状态

基本信息

批准号：
8118789
负责人：
KAROL BOMSZTYK
金额：
$ 35.13万
依托单位：
UNIVERSITY OF WASHINGTON
依托单位国家：
美国
项目类别：
财政年份：
2010
资助国家：
美国
起止时间：
2010-08-01 至 2014-07-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8118789
关键词：
Acute Acute Kidney Failure Acute Renal Failure with Renal Papillary Necrosis Address Bacterial Toxins Bilateral Binding Binding Sites Biological Assay Biological Markers Biology Blood Circulation CCL2 gene Cause of Death Cell model Cells Chromatin Chromatin Remodeling Factor Chromatin Structure Cisplatin Complement Contralateral DNA Modification Process Data Development Docking Endotoxemia Endotoxins Enzymes Epigenetic Process Event Exposure to Failure Functional disorder Future Gene Activation Gene Targeting Genes Genetic Transcription Genomics Goals Histone Acetylation Histones Human Hypersensitivity In Vitro Infection Inflammation Mediators Inflammatory Inflammatory Response Injury Ischemia Ischemic Preconditioning Kidney Knowledge Laboratories Lead Left Ligands Ligation Lipopolysaccharides Maintenance Mediating Mediator of activation protein Memory Messenger RNA Methylation Modeling Modification Molecular Nature Nucleosomes Organ Organ failure Pathway interactions Patients Phase Polymerase Polymerase Gene Positioning Attribute Preventive Process Production RNA Polymerase II Reagent Recruitment Activity Renal Tissue Reperfusion Therapy Right kidney SMARCA4 gene Sepsis Sequence Analysis Series Severities Small Interfering RNA Specific qualifier value Staging TNF gene Testing Transcription Coactivator Transcription Factor AP-1 Trauma Tubular formation Ureteral obstruction Variant Venous base bisulfite cell injury chemokine chromatin remodeling clinically relevant cytokine demethylation histone modification in vivo kidney cortex knock-down mRNA Stability memory process mortality novel promoter public health relevance receptor research study response response to injury septic spatiotemporal transcription factor translational approach

项目摘要

DESCRIPTION (provided by applicant): In the aftermath of acute renal injury (AKI), proximal tubular cells manifest dramatic hyper-responsiveness to Toll receptor ligands, most notably, endotoxin. This exaggerates renal cytokine (e.g., TNF-1) / chemokine (e.g., MCP-1) production which can worsen the severity of acute renal failure (ARF). With renal venous cytokine efflux, extra-renal injury may also result. This LPS hyper-responsiveness is transcriptionally regulated, and mediated, in part, by gene activating chromatin events. The overall goal of this proposal is to: i) further define the pathways which induce the underlying chromatin changes; and ii) delineate how they mediate an exaggerated LPS responsive state. To focus the application, one ARF model (ischemia/reperfusion) 1 exposure to one Toll ligand (LPS) will be tested in three Specific Aims: Aim#1: Define the temporal sequence of chromatin changes along the TNF-1 gene in AKI. We will characterize chromatin changes, including CpG methylation/demethylation, histone acetylation, methylation, histone variants, and alterations in nucleosome positions. Both renal injury, and proximal tubule-specific injury (in cultured tubular HK-2 cells), will be studied following reversible ischemia + LPS. These in vivo and in vitro experiments will set the stage for mechanistic assessments (Aim 3). Aim #2: Ascertain which transcription and chromatin remodeling factors / enzymes are recruited to the TNF-1 gene in AKI. We have already shown that the chromatin remodeler BRG1 is recruited to the TNF-1 gene and is required for injury-induced TNF-1 transcription. We have also demonstrated that NF-:B and AP-1 transcription factors are recruited to the TNF-1 gene in AKI. Using Matrix ChIP, we will more fully define the spatiotemporal sequence of chromatin and transcription events at the TNF-1 gene in response to reversible ischemia + LPS. This will help to ascertain which factors are hyper-recruited to the TNF-1 gene and may thus drive the LPS hyper-responsive state. Aim #3 Test the mechanistic relevance of the above defined changes in the induction of the LPS hyper-responsive state. The mechanistic relevance of the above defined changes in establishing LPS hyper-responsiveness will be tested both in vitro and in vivo using siRNA targeting (as we have already done to knock-down BRG1 in HK-2 cells). We will complement these in vivo and in vitro siRNA approaches with the use of pharmacologic reagents directed at the putative molecular mediators of the hyper-responsive chromatin state. The latter approach may lead to therapeutically relevant translational approaches for modulating experimental AKI and associated multiorgan failure. The proposed studies are highly novel in that, with the exception of data obtained by the PIs, virtually no information exists as to the nature, and consequences of, chromatin alterations in response to AKI. Hence, new perspectives on mechanisms of AKI and its downstream consequences should result. PUBLIC HEALTH RELEVANCE: Acute kidney injury (AKI) leads to renal production of inflammatory mediators and sensitizes/primes the kidney to bacterial toxins. These renal inflammatory mediators gain access to the systemic circulation and can induce extra-renal tissue damage, a common cause of mortality in patients with AKI, due to infections, trauma and other illness. We propose to define the chromatin/transcriptional mechanisms for these renal responses with the goal to develop preventive treatments in the future.

描述（由申请人提供）：在急性肾脏损伤（AKI）之后，近端管状细胞表现出对收费受体配体的显着性超反应性，最著名的是内毒素。这夸大了肾细胞因子（例如TNF-1） /趋化因子（例如MCP-1）的产生，这可能会使急性肾衰竭（ARF）的严重程度恶化。随着肾静脉细胞因子外排，可能会导致肾上腺外损伤。该LPS超反应性受到转录调节，并部分通过激活染色质事件的基因介导。该提案的总体目标是：i）进一步定义诱导潜在染色质变化的途径； ii）描述它们如何介导夸张的LPS响应状态。为了集中应用，将在三个特定的目标中测试一个ARF模型（缺血/再灌注）1接触一个收费配体（LPS）：AIM＃1：定义AKI中TNF-1基因的染色质变化的时间序列。我们将表征染色质的变化，包括CpG甲基化/脱甲基化，组蛋白乙酰化，甲基化，组蛋白变异以及核小体位置的改变。在可逆性缺血 + LPS之后，将研究肾脏损伤和近端小管特异性损伤（在培养的管状HK-2细胞中）。这些体内和体外实验将为机械评估奠定阶段（AIM 3）。目标＃2：确定哪些转录和染色质重塑因子 /酶被募集到AKI中的TNF-1基因。我们已经表明，将染色质重塑剂BRG1募集到TNF-1基因，是损伤诱导的TNF-1转录所必需的。我们还证明了NF-：B和AP-1转录因子被募集到AKI中的TNF-1基因。使用矩阵芯片，我们将更全面地定义TNF-1基因在TNF-1基因上的时空序列，以响应可逆性缺血 + LP。这将有助于确定哪些因素已超过TNF-1基因，因此可能会驱动LPS超反应态。 AIM＃3测试了上述LPS超响应状态诱导上上述定义变化的机理相关性。上述确定LPS超反应性变化的机理相关性将在体外和体内使用siRNA靶向进行测试（就像我们已经在HK-2细胞中敲低BRG1的那样）。我们将使用针对高反应性染色质状态的推定分子介质的药理试剂来补充这些体内和体外siRNA方法。后一种方法可能会导致用于调节实验AKI和相关多机器人失败的治疗相关的翻译方法。提出的研究是高度新颖的，因为除了PIS获得的数据外，几乎不存在有关染色质改变对AKI的性质和后果的信息。因此，应该导致对AKI机制及其下游后果的新观点。公共卫生相关性：急性肾脏损伤（AKI）导致炎症介质的肾脏产生，并使肾脏对细菌毒素进行敏感/素。这些肾脏炎症介质可以进入全身循环，并可能诱导肾外组织损伤，这是由于感染，创伤和其他疾病而导致AKI患者死亡的常见原因。我们建议定义这些肾脏反应的染色质/转录机制，以未来开发预防性治疗的目标。