Targeting T lymphocyte Keap1 for acute kidney injury

靶向 T 淋巴细胞 Keap1 治疗急性肾损伤

• 批准号: 9333374
• 负责人: HAMID RABB
• 金额: $ 44.66万
• 依托单位: JOHNS HOPKINS UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-08-16 至 2020-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9333374
• 关键词: Acute Renal Failure with Renal Papillary Necrosis; Adoptive Transfer; Antioxidants; Apoptosis; CD4 Positive T Lymphocytes; CD8-Positive T-Lymphocytes; CRISPR/Cas technology; Cell Therapy; Cells; Chalcones; Cisplatin; Clinical; Cues; Data; Disease; Environment; FOXP3 gene; Frequencies; Future; Health Care Costs; Homeostasis; Human; Immune; Inflammation; Inflammatory; Inflammatory Response; Injury; Interferon Type II; Interleukin-2; Intervention; Ischemia; Kidney; Kidney Diseases; Knockout Mice; Link; Mediating; MicroRNAs; Modeling; Morbidity - disease rate; Mus; Nephrotoxic; Outcome; Pathogenesis; Pathogenicity; Pathway interactions; Patients; Pharmacology; Phenotype; Population; Predisposition; Process; Production; Recovery; Regulatory T-Lymphocyte; Renal function; Reperfusion Therapy; Role; Small Interfering RNA; Specificity; Sulforaphane; T cell response; T-Cell Immunologic Specificity; T-Lymphocyte; TNF gene; Technology; Testing; Therapeutic; Therapeutic Studies; Thymus Gland; Wild Type Mouse; clinical translation; cytokine; design; experimental study; improved; in vitro Assay; in vivo; inhibitor/antagonist; insight; mortality; mouse model; nephrotoxicity; novel; novel therapeutics; overexpression; prevent; protective effect; repaired; response; targeted treatment; transcription factor

PROJECT SUMMARY /ABSTRACT Acute kidney Injury (AKI)-associated morbidity and mortality is a major clinical problem that involves multiple overlapping pathophysiological mechanisms. Our lab and others have established the modulatory roles of T lymphocytes as well as the protective role of transcription factor Nrf2 in ischemia reperfusion (IR) and cisplatin induced AKI. We recently demonstrated that T cell deletion of Keap1 which augments specific Nrf2 activity provides significant protection from IR induced AKI in knockout mice, and that adoptive transfer of T cells with augmented Nrf2 activity improves kidney function and survival following AKI in wild type mice. These findings were accompanied by a significant increase in regulatory T (Treg) cell frequency and numbers, and reduced proinflammatory cytokine production by T cells in the kidneys of mice with Keap1 deficient T cells. These novel observations reveal an unexpected relationship between Keap1 and T cell homeostasis and function in AKI, however the underlying mechanisms are unknown. The overarching aim of this proposed study is to test the hypothesis that Keap1 regulates the expansion of Treg cell population that subsequently suppresses harmful inflammatory responses during AKI. In order to test our hypotheses, we will determine whether Treg cells with Keap1 deletion have enhanced proliferation or decreased apoptosis as compared to Treg cells from wild type (WT) mice. We will also evaluate if Keap1 deletion increases the suppressive function of Treg cells, enhances TCR sensitivity in T cells and regulates response to IL-2 under steady state and in two different models (IR and cisplatin) of AKI in mice. To further establish the specificity of Keap1 deletion and to investigate any Nrf2 independent effects of Keap1 in T cells we will generate mice lacking Keap1 specifically in Treg, CD4 and CD8 T cells and determine whether Treg specific Keap1 deletion is sufficient to prevent ischemic and nephrotoxic AKI. We will further generate mice with T cell specific deletion of either Nrf2 alone or Keap1 and Nrf2 together (double KO) to delineate Nrf2 dependent and independent effects. In the final aim we will transition from mechanistic studies to experiments that will set the stage for clinical translation. We will explore therapeutic approaches including pharmacologic Nrf2 activators to activate Nrf2 and Keap1 specific CRISPR/Cas9 technology, siRNAs and miRNAs to delete Keap1 in mouse T cells ex vivo. We will then examine the effect of these Nrf2 activation/Keap1 deletion strategies on phenotypic and functional T cell responses as well as AKI outcome in IR and cisplatin AKI models. We will test the most promising Nrf2 activation/Keap1 deletion strategy in human primary T lymphocytes and examine its effects on Nrf2 regulated antioxidant response, proinflammatory cytokine production, and T cell dynamics. Successful completion of these studies should result in major advances regarding our understanding on the role of Keap1/Nrf2 in T cell homeostasis and function, and also set the stage for future clinical interventions targeting Keap1-Nrf2 pathway for AKI and other inflammatory diseases.

项目总结/摘要 急性肾损伤（阿基）相关的发病率和死亡率是一个主要的临床问题，涉及多个 重叠的病理生理机制。我们的实验室和其他实验室已经确定了T 以及转录因子Nrf 2在缺血再灌注（IR）和顺铂中的保护作用 诱发阿基。我们最近证明，T细胞缺失Keap 1可增强特异性Nrf 2活性， 在敲除小鼠中提供了针对IR诱导的阿基的显著保护，并且具有IR诱导的AKI的T细胞的过继转移在小鼠中是有效的。 增强的Nrf 2活性改善了野生型小鼠阿基后的肾功能和存活率。这些发现 伴随着调节性T（Treg）细胞频率和数量的显着增加， Keap 1缺陷型T细胞小鼠肾脏中T细胞的促炎细胞因子产生。这些新颖 观察结果揭示了阿基中Keap 1和T细胞稳态和功能之间的意想不到的关系， 然而，其潜在机制尚不清楚。这项研究的主要目的是测试 Keap 1调节Treg细胞群的扩增， 抑制阿基期间有害的炎症反应。为了验证我们的假设，我们将确定 Keap 1缺失的Treg细胞是否具有增强的增殖或减少的凋亡， 来自野生型（WT）小鼠的Treg细胞。我们还将评估Keap 1缺失是否会增加抑制功能， Treg细胞，增强T细胞中的TCR敏感性，并在稳态和两种情况下调节对IL-2的应答。 在小鼠中的阿基的不同模型（IR和顺铂）。为了进一步确定Keap 1缺失的特异性， 研究Keap 1在T细胞中的任何Nrf 2独立作用，我们将产生缺乏Keap 1的小鼠，特异性地 Treg、CD 4和CD 8 T细胞，并确定Treg特异性Keap 1缺失是否足以预防 缺血性和肾毒性阿基。我们将进一步产生具有单独的Nrf 2或Nrf 3的T细胞特异性缺失的小鼠。 Keap 1和Nrf 2一起（双KO）描述Nrf 2依赖性和独立性效应。在最后的目标 我们将从机械研究过渡到实验，为临床研究奠定基础。 翻译.我们将探索治疗方法，包括药理学Nrf 2激活剂，以激活Nrf 2 以及Keap 1特异性CRISPR/Cas9技术、siRNA和miRNA，以在小鼠T细胞中离体缺失Keap 1。 然后，我们将研究这些Nrf 2激活/Keap 1缺失策略对表型和功能的影响。 IR和顺铂阿基模型中的T细胞应答以及阿基结果。我们将测试最有前途的NRF 2 激活/Keap 1缺失策略，并检查其对Nrf 2调节的影响。 抗氧化反应、促炎细胞因子产生和T细胞动力学。成功完成 这些研究将使我们对Keap 1/Nrf 2在T细胞中的作用的理解取得重大进展。 体内平衡和功能，并为未来针对Keap 1-Nrf 2通路的临床干预奠定基础 用于阿基和其他炎症性疾病。