Mitochondrial Recovery after Acute Kidney Injury Needs Ribonucleotide Reductase

急性肾损伤后的线粒体恢复需要核糖核苷酸还原酶

基本信息

批准号：
10396045
负责人：
MANJERI A VENKATACHALAM
金额：
$ 23.25万
依托单位：
UNIVERSITY OF TEXAS HLTH SCIENCE CENTER
依托单位国家：
美国
项目类别：
财政年份：
2020
资助国家：
美国
起止时间：
2020-05-01 至 2024-04-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10396045
关键词：
2&apos -Deoxythymidine Acute Renal Failure with Renal Papillary Necrosis Affect Apoptosis Atrophic Brain Bypass Catalytic Domain Cell Culture Techniques Cell Hypoxia Cells Chronic Kidney Failure Code Cultured Cells Cytosol DNA DNA Damage DNA Repair DNA biosynthesis Data Deoxycytidine Deoxyguanosine Development Dialysis procedure Disease Enzymes Failure Genes Growth Health Healthcare Hypoxia Impairment Incidence Injury Injury to Kidney Interphase Cell Investigation Kidney Kidney Diseases Kidney Failure Knockout Mice Lead Lesion LoxP-flanked allele Mediating Membrane Proteins Mitochondria Mitochondrial DNA Mitochondrial DNA depletion syndromes Modeling Molecular Morbidity - disease rate Mus Muscular Atrophy Mutation Normal Cell Nuclear Null Lymphocytes Oxidants Oxygen PINK1 gene Parkin Pathologic Pathology Pathway interactions Phenotype Process Production Proteins RRM1 gene RRM2 gene Recovery Regimen Regulation Reperfusion Injury Reporter Research Retina Ribonucleotide Reductase Ribonucleotide Reductase Subunit Risk Role Stress Superoxides Supplementation TP53 gene Therapeutic Therapeutic Agents Transgenes Transplantation Treatment Efficacy Tubular formation Undifferentiated cancer cell cell growth deoxyribonucleoside triphosphate efficacy testing improved in vivo insight molecular pathology mortality mouse model novel overexpression oxidant stress preservation prevent renal hypoxia repaired respiratory protein response tripolyphosphate

项目摘要

PROJECT SUMMARY/ABSTRACT Significance: Tubule atrophy underlies progression of acute kidney injury (AKI) to chronic kidney disease. The basis for tubule atrophy after AKI is unknown. We identified a pathology involving RRM2B, alternate regulatory subunit of ribonucleotide reductase (RNR) as a basis for tubule atrophy after AKI. Cell culture data suggest that molecular bypass of RNR by deoxynucleosides can produce deoxynucleoside triphosphates (dNTPs) for DNA synthesis and repair by oxygen independent salvage pathways after AKI to promote tubule recovery. RNR produces dNTPs. Its classical regulatory subunit RRM2 is inhibited by hypoxia, but the hypoxia tolerant RRM2B can substitute for RRM2 to maintain dNTPs during hypoxia to prevent nuclear and mitochondrial DNA damage and preserve cell integrity. RRM2B is induced by hypoxia in cultured cells. Hypoxia mediated increase of mitochondrial superoxide may induce RRM2B as a beneficial adaptation. A regulatory role for mitochondrial superoxide was suggested by our finding that MitoPQ, which increases superoxide selectively in mitochondria, markedly increased cellular RRM2B protein content. The benefits afforded by RRM2B in hypoxic cells would be abrogated if RRM2B is not available. In support, we showed that deletion of RRM2B from cultured tubule cells increases DNA damage during hypoxia, and prevents recovery during reoxygenation. Deoxynucleosides rescued such cells from DNA damage, decreased injury and promoted recovery. We showed also that tubule atrophy after AKI is accompanied by marked RRM2B depletion and DNA damage. Since recovering kidneys are hypoxic, RNR inhibition is expected. In such kidneys RRM2B loss in tubules (rather than an adaptive increase) will have deleterious consequences. Thus, RRM2B loss after AKI may be the cause for tubule atrophy. While the cause for RRM2B decline in tubules after AKi is unclear, its importance for recovery requires investigation. To this end, we have three Specific Aims. Aim 1. We will utilize mouse models of RRM2B deletion and overexpression in tubules to investigate its role in recovery from AKI. Aim 2. We will use cultured tubule cells with RRM2B deletion and overexpression, and deoxynucleoside supplementation, to investigate the role of RNR activity, RRM2B levels and salvage synthesis of dNTPs in cellular responses to hypoxia. To examine the role of mitochondrial oxidants in adaptation to stress, we will use Mito-PQ, a selective inducer of mitochondrial superoxide, to elucidate mechanisms of RRM2B regulation. Aim 3. We will test the efficacy of treatment with deoxynucleosides to promote recovery from ischemic AKI. Impact: The studies we propose will yield new insights into cellular mechanisms that determine successful or failed tubule recovery from AKI and possibly identify deoxynucleosides as novel useful therapeutic agents.

项目摘要/摘要意义：小管萎缩是急性肾脏损伤（AKI）向慢性肾脏疾病的进展。这 AKI后小管萎缩的基础是未知的。我们确定了一种涉及RRM2B的病理学核糖核苷酸还原酶（RNR）的亚基作为AKI后小管萎缩的基础。细胞培养数据表明脱氧核苷的RNR分子旁路可以产生DNA的脱氧核苷三磷酸（DNTP） AKI后通过氧气独立打捞途径的合成和修复，以促进小管恢复。 RNR产生DNTP。它的经典调节亚基RRM2受到缺氧的抑制，但缺氧耐受性 RRM2B可以代替RRM2在缺氧期间维持DNTP，以防止核和线粒体DNA 损坏并保留细胞完整性。 RRM2B是由培养细胞中缺氧诱导的。缺氧介导的增加线粒体超氧化物可能诱导RRM2B作为有益的适应。线粒体的调节作用我们的发现提出了超氧化物，即在线粒体中有选择地增加超氧化物的mitopq，明显增加了细胞RRM2B蛋白含量。如果RRM2B不可用，RRM2B在低氧细胞中提供的好处将被废除。在支持下，我们表明从培养小管细胞中删除RRM2B会增加缺氧期间的DNA损伤，并防止DNA损伤在重氧中恢复。脱氧核苷使此类细胞免于DNA损伤，减少损伤和促进恢复。我们还表明，AKI后的小管萎缩伴随着明显的RRM2B耗竭和DNA损伤。由于恢复肾脏是缺氧的，因此预计RNR抑制作用。在这样的肾脏RRM2B中小管的损失（而不是自适应增加）将带来有害的后果。因此，RRM2B损失 AKI可能是小管萎缩的原因。虽然AKI之后小管下降的RRM2B的原因尚不清楚，但对恢复的重要性需要调查。为此，我们有三个具体的目标。 AIM 1。我们将利用小管中RRM2B缺失和过表达的鼠标模型来研究其在从AKI恢复。 AIM 2。我们将使用带有RRM2B缺失和过表达的培养的小管细胞以及脱氧核苷补充，研究RNR活性的作用，RRM2B水平和DNTP的挽救合成细胞对缺氧的反应。为了检查线粒体氧化剂在适应压力中的作用，我们将使用 Mito-PQ是线粒体超氧化物的选择性诱导剂，以阐明RRM2B调节的机制。 AIM 3。我们将测试脱氧核苷治疗的功效，以促进缺血性AKI的恢复。影响：我们提出的研究将产生对决定成功或的细胞机制的新见解小管从AKI中恢复失败，并可能识别出脱氧核苷是新型有用的治疗剂。