Role of KLF15 in proximal tubule metabolism

KLF15 在近曲小管代谢中的作用

• 批准号: 10481366
• 负责人: Sandeep K Mallipattu
• 金额: --
• 依托单位: NORTHPORT VA MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-01-01 至 2026-09-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10481366
• 关键词: Acetyl Coenzyme A; Acute Renal Failure with Renal Papillary Necrosis; Agonist; American; Anabolism; Aristolochic Acids; Attenuated; Binding Sites; Cardiovascular Diseases; Cell Cycle Arrest; Cell Differentiation process; Cell physiology; Cells; ChIP-seq; Chronic Kidney Failure; Cisplatin; Clinical; Compensation; DNA Damage; DNA Repair; Data; Diglycerides; Down-Regulation; Fatty Acids; Fibroblasts; Funding; General Population; Generations; Genes; Glycolysis; Goals; Human; Injury; Injury to Kidney; Ischemia; Kidney; Lipids; Mediating; Metabolic; Metabolic Pathway; Metabolism; Mitochondria; Mus; Myofibroblast; Nonesterified Fatty Acids; Organoids; Oxidative Stress; PPAR alpha; Palmitates; Pathway interactions; Phase; Pre-Clinical Model; Predisposition; Prevalence; Production; Profibrotic signal; Proliferating; Reactive Oxygen Species; Recovery; Regression Analysis; Regulation; Renal glomerular disease; Research; Respiration; Risk Factors; Role; Secondary to; Signal Transduction; Signaling Molecule; Source; Testing; Therapeutic; Toxin; Transcriptional Regulation; Triglycerides; United States Department of Veterans Affairs; Veterans; cardiovascular risk factor; cell dedifferentiation; cell injury; chemotherapeutic agent; clinically relevant; extracellular; in silico; inducible gene expression; interest; kidney biopsy; knock-down; macromolecule; mouse model; novel; novel therapeutic intervention; overexpression; oxidation; peroxidation; prevent; repaired; single nucleus RNA-sequencing; transcription factor; transcriptome sequencing

Chronic kidney disease (CKD) is a leading risk factor for cardiovascular disease, with a disproportionate burden on U.S. Veterans. Recent data demonstrates that acute kidney injury (AKI), despite initial renal recovery, is a major risk factor for CKD. The proximal tubule (PT) is the primary target in AKI due to its high susceptibility to ischemia and DNA-damaging nephrotoxins such as chemotherapeutic agents. Damaged PT cells dedifferentiate, and initially undergo cell cycle arrest, predominantly at the G2/M checkpoint. This cell cycle arrest may allow repair of DNA damage caused by reactive oxygen species secondary to mitochondrial damage or directly by DNA-damaging toxins. Sustained cell cycle arrest is associated with a switch to secretion of pro-fibrotic signaling molecules, inducing resident fibroblasts to proliferate and differentiate to myofibroblasts, beginning the transition to a fibrotic injury. PT cells also undergo metabolic reprograming, with severe downregulation of fatty acid b-oxidation (FAO), and limited compensation by anerobic glycolysis. While restoring FAO either by overexpressing Ppara or by using a peroxisome proliferator activated receptor alpha (PPARa) agonist attenuates AKI and CKD in murine models, this has not translated to use in clinical AKI, suggesting additional factors are required to mitigate the progression from AKI to CKD. Krüppel-Like Factor 15 (KLF15) is a kidney-enriched transcription factor, involved in a diverse range of cellular processes, including cell differentiation and FAO. In the initial funding period of the VA Merit, we demonstrated the salutary role of KLF15 in glomerular disease leading to a composition-of-matter IP on KLF15 agonists by the Veterans Affairs. During this initial period, we also identified that KLF15 is highly expressed in differentiated PT cells, but is significantly reduced in murine models of PT injury. Utilizing a murine model of PT-specific injury secondary to DNA damage, we observed that PT-specific knockdown of Klf15 exacerbated AKI as well as CKD. PT-specific knockdown of Klf15 also increased pathways involving cell cycle arrest, oxidative stress, pro-fibrotic signaling and a decrease in pathways utilizing FA for the generation of acetyl-CoA, a central metabolic intermediate in macromolecule biosynthesis and energy production. We also observed an enrichment of genes critical for FA utilization with putative and proximal KLF15- and PPARa-binding sites, suggesting potential KLF15-PPARa co-operativity in the regulation of FA utilization. In addition, we demonstrated a significant increase in glycerolipid synthesis pathways and lipid droplet formation in the setting of suppressed FAO, suggesting a potential compensatory mechanism post-DNA damage. KLF15 expression was also associated with PPARA expression in human kidney biopsies with and without CKD. In addition, multivariate regression analysis demonstrated that a decrease in KLF15 expression was independently associated with eGFR decline, suggesting that the loss of KLF15 might be a key driver of PT injury. Based on these preliminary data and strong scientific rigor of prior research, we hypothesize that KLF15-PPARa co-operativity drives the utilization of excess free fatty acids for acetyl-CoA and glycerolipid synthesis to prevent maladaptive PT repair post-DNA damage. We propose to test this hypothesis by (1) determining the mechanism by which KLF15-PPARa co-operativity restores PT metabolism after PT injury secondary to DNA-damage and (2) to investigate the requisite role of KLF15-PPARa in PT injury secondary to DNA-damage. This proposal will address a current gap in the field by investigating the mechanisms mediating transcriptional regulation of FA utilization in the PT cells post-DNA damage. The long-term goal of our project is to demonstrate that the combination of KLF15 and PPARa agonists is a novel therapeutic strategy to mitigate PT injury post-DNA damage. Identification of novel targets for the treatment of AKI is of major interest to the VA, given the high burden of CKD among U.S. Veterans.

慢性肾脏病（CKD）是心血管疾病的主要危险因素， 给美国退伍军人带来了沉重的负担。最近的数据表明，急性肾损伤（阿基），尽管 初始肾脏恢复是CKD的主要风险因素。近端小管（PT）是阿基的主要靶点， 其对缺血和DNA损伤性肾毒素如化疗剂的高度敏感性。 受损的PT细胞去分化，最初经历细胞周期停滞，主要在G2/M检查点。 这种细胞周期停滞可能允许修复由继发于细胞凋亡的活性氧引起的DNA损伤。 线粒体损伤或直接通过DNA损伤毒素。持续的细胞周期停滞与 转换为促纤维化信号分子的分泌，诱导驻留的成纤维细胞增殖， 细胞分化成肌成纤维细胞，开始向纤维化损伤转变。PT细胞也经历代谢 重编程，严重下调脂肪酸b-氧化（FAO），和有限的补偿， 无氧糖酵解在通过过表达Ppara或使用过氧化物酶体增殖剂来恢复FAO的同时， 活化受体α（PPARa）激动剂在鼠模型中减弱阿基和CKD，这尚未转化为 用于临床阿基，表明需要其他因素来缓解从阿基到CKD的进展。 Krüppel样因子15（KLF 15）是一种富含肾脏的转录因子，参与多种肾脏疾病的发生。 细胞过程，包括细胞分化和FAO。在VA Merit的初始资助期间，我们 证明了KLF 15在肾小球疾病中的有益作用，导致KLF 15的物质组成IP 退伍军人事务部的激动剂。在这一初始阶段，我们还发现KLF 15在细胞中高度表达， 分化的PT细胞，但在PT损伤的鼠模型中显著降低。利用小鼠模型， PT特异性损伤继发于DNA损伤，我们观察到PT特异性敲除Klf 15加重了 阿基和CKD。PT特异性敲除Klf 15也增加了涉及细胞周期停滞的途径， 氧化应激、促纤维化信号传导和利用FA产生乙酰辅酶A的途径减少， 大分子生物合成和能量产生中的一种中心代谢中间体。我们还观察到， 富集具有推定的和近端的KLF 15-和PPARa-结合位点的对FA利用至关重要的基因， 提示KLF 15-PPARa在调节FA利用方面具有潜在的协同性。另外我们 证明了甘油脂质合成途径和脂滴形成的显着增加， 抑制粮农组织，这表明一个潜在的补偿机制后DNA损伤。KLF 15表达 在有和没有CKD的人肾活检组织中也与PPARA表达相关。此外，本发明还提供了一种方法， 多变量回归分析表明，KLF 15表达的降低是独立的， 与eGFR下降相关，表明KLF 15的丢失可能是PT损伤的关键驱动因素。 基于这些初步数据和先前研究的强大科学严谨性，我们假设， KLF 15-PPARa协同作用驱动过量游离脂肪酸用于乙酰辅酶A和甘油脂质 合成以防止适应不良PT修复后DNA损伤。我们建议通过（1）来检验这一假设。 确定PT损伤后KLF 15-PPARa协同作用恢复PT代谢的机制 （2）研究KLF 15-PPARa在继发于DNA损伤的PT损伤中的必要作用， DNA损伤这项建议将通过调查调解机制， DNA损伤后PT细胞中FA利用的转录调节。我们项目的长期目标是 是为了证明KLF 15和PPARa激动剂的组合是一种新的治疗策略， 减轻DNA损伤后的PT损伤。用于治疗阿基的新靶点的鉴定是主要的兴趣 鉴于CKD在美国退伍军人中的高负担，