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Role of the MUC1/HIF-1a Complex in the kidney proximal tubule during ischemia-reperfusion injury.

MUC1/HIF-1a 复合物在缺血再灌注损伤期间肾近曲小管中的作用。

基本信息

批准号：
10541885
负责人：
Mohammad Al-bataineh
金额：
$ 11.74万
依托单位：
UNIVERSITY OF PITTSBURGH AT PITTSBURGH
依托单位国家：
美国
项目类别：
财政年份：
2021
资助国家：
美国
起止时间：
2021-12-15 至 2024-11-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10541885
关键词：
2-oxoglutarate 3-dioxygenase proline ARNT protein Acute Renal Failure with Renal Papillary Necrosis Adult Affect Apical Award Binding Biological Assay Cell Line Cell Nucleus Cell surface Cells Cessation of life Chimera organism Chronic Kidney Failure Clinical Co-Immunoprecipitations Complex Data Dimerization Distal Elements Epithelial Cells Epithelium Excess Mortality Financial Hardship Functional disorder Genes Genetic Genetic Transcription Glucose Goals Grant Human Hydroxylation Hypotension Hypoxia Immunofluorescence Microscopy In Vitro Incidence Injury Injury to Kidney Ischemia Kidney Knock-out Knockout Mice Limb structure Location Mediator Membrane Glycoproteins Mercuric chloride Metabolic dysfunction Metabolism Morbidity - disease rate Morphology Mucin 1 protein Mus Names Nephrons Nuclear Oxygen Pathway interactions Patients Phosphorylation Play Proteins Proximal Kidney Tubules Publishing Pyruvate Kinase Recovery Regulation Renal function Reperfusion Injury Reporting Research Research Personnel Resistance Role Sepsis Severities Signal Transduction Surface Testing Therapeutic Intervention Thick Tissues Transactivation Transcriptional Activation Transfection Tubular formation Work beta catenin career congenic coping experimental study hypoxia inducible factor 1 inhibitor ischemic injury kidney cell mortality mouse model mutant novel novel therapeutics pancreatic cancer cells prevent renal damage repaired response therapy design trafficking transcription factor urinary

项目摘要

Project Abstract. Acute kidney injury (AKI) is a common and devastating clinical problem. Despite the morbidity, mortality, and financial drain associated, there are no established therapies outside of dialytic treatment. Recent work has identified adaptive responses in the tubular epithelium to cope with ischemia-reperfusion injury (IRI). There is a critical need to identify how adaptive responses are regulated in AKI so that therapeutic interventions to prevent and treat AKI can be developed. Hypoxia inducible factor-1 (HIF-1) is a transcription factor regarded as the most significant mediator of cellular adaptive responses to hypoxic insult. There is emerging evidence that the transmembrane glycoprotein mucin 1 (MUC1) expressed on the apical surface of kidney epithelia plays a novel and important role in enhancing HIF-1 activity. We have strong preliminary data suggesting that genetic deletion of Muc1 from mouse tubular epithelial cells exacerbates damage especially in the proximal tubule (PT) from IRI, limits adaptive HIF-1 responses, alters tubular metabolism, and inhibits recovery. The central hypothesis of this application is that MUC1 protects the kidney during IRI by transactivation of the HIF-1 adaptive response specifically in the recovering proximal tubule. The specific aims are: 1. To determine if MUC1 protection of the kidney during IRI requires MUC1-dependent HIF-1a nuclear targeting and subsequently transcriptional activation of the HIF-1 pathway specifically in the recovering PT. I will test the hypothesis that MUC1 protects the kidney during IRI by transactivation of the HIF-1 adaptive response in the recovering PT. Experiments are proposed to stabilize HIF-1a in the proximal tubule in the Muc1 KO mice during IRI by introducing a PT KO of VHL, and treatment with vehicle or a specific MUC1 inhibitor to block MUC1 internalization and nuclear delivery. Results of these studies will reveal if MUC1 simply stabilizes HIF-1a levels, or if MUC1 nuclear targeting is essential for transduction of the protective HIF-1 pathway. 2. To characterize the MUC1-HIF-1a complex in cultured kidney cells. I will test the hypothesis that MUC1 stabilizes HIF-1a by direct binding. Experiments are proposed here to characterize the MUC1-HIF-1a complex using both in vitro pull-down assays and co-IP studies in primary PT cells and human kidney cell lines with wild type and mutant constructs, and to determine if MUC1 binding to HIF-1a results in its trafficking to the nucleus. The goal of my research is to fully characterize the MUC1-HIF-1a interaction and determine the mechanism of its protective role in the kidney PT during IRI. This information can be used to design therapies to induce MUC1 and limit the severity of AKI. This award will provide me with additional support to generate preliminary data for an R01-level grant to further investigate whether increased levels of renal MUC1 provide resistance to IRI.

项目摘要。急性肾损伤（阿基）是一种常见的和毁灭性的临床问题。尽管发病率很高，死亡率和相关的经济损失，除透析治疗外，还没有确定的治疗方法。最近工作已经确定了肾小管上皮中的适应性反应以科普缺血-再灌注损伤（IRI）。迫切需要确定阿基中的适应性反应是如何调节的，预防和治疗阿基的方法。缺氧诱导因子-1（Hypoxia inducible factor-1，HIF-1）是一种转录因子，作为细胞对缺氧损伤的适应性反应的最重要的介质。有新的证据表达于肾上皮细胞顶面的跨膜糖蛋白粘蛋白1（MUC 1）在肾上皮细胞分化过程中起着重要的作用。在增强HIF-1活性方面具有新颖且重要的作用。我们有充分的初步数据表明，小鼠肾小管上皮细胞Muc 1的缺失会加重损伤，尤其是近端小管（PT）从IRI，限制适应性HIF-1反应，改变肾小管代谢，并抑制恢复。的本申请的中心假设是MUC 1通过HIF-1的反式激活在IRI期间保护肾脏适应性反应，特别是在恢复近端小管。具体目标是： 1.为了确定IRI期间MUC 1对肾脏的保护是否需要MUC 1依赖性HIF-1a核表达，靶向并随后转录激活HIF-1通路，特异性地在恢复的 PT.我将检验MUC 1在IRI期间通过HIF-1适应性转录因子的反式激活来保护肾脏的假设。恢复中PT的反应。提出实验以稳定Muc 1中近端小管中的HIF-1a。在IRI期间通过引入VHL的PT KO和用媒介物或特异性MUC 1抑制剂处理以在IRI期间通过VHL的PT KO治疗KO小鼠，阻断MUC 1内化和核递送。这些研究的结果将揭示MUC 1是否只是稳定 HIF-1a水平，或者MUC 1核靶向是否对保护性HIF-1通路的转导至关重要。 2.鉴定培养的肾细胞中MUC 1-HIF-1a复合物。我将检验MUC 1 通过直接结合稳定HIF-1a。本文提出实验来表征MUC 1-HIF-1a复合物在原代PT细胞和人肾细胞系中使用体外下拉测定和co-IP研究，类型和突变体构建体，并确定MUC 1与HIF-1a的结合是否导致其运输到细胞核。本研究的目的是充分表征MUC 1-HIF-1a相互作用，并确定MUC 1-HIF-1a相互作用的机制。在IRI期间其在肾脏PT中的保护作用。该信息可用于设计诱导MUC 1的疗法。并限制阿基的严重程度。该奖项将为我提供额外的支持，以生成初步数据申请R 01级资助，以进一步研究肾脏MUC 1水平的增加是否提供对IRI的抵抗。