Mechanisms of active renal repair: Single-cell trajectories of chromatin accessibility and gene expression changes in renal tubule cells

主动肾脏修复机制：肾小管细胞染色质可及性和基因表达变化的单细胞轨迹

• 批准号: 427245510
• 负责人: Dr. Louisa Gerhardt
• 金额: --
• 依托单位: Center for Regenerative Medicine and Stem Cell Research
• 依托单位国家: 德国
• 项目类别: Research Fellowships
• 财政年份: 2019
• 资助国家: 德国
• 起止时间: 2018-12-31 至 2021-12-31
• 项目状态: 已结题
• 来源: https://gepris.dfg.de/gepris/projekt/427245510?language=en
• 关键词: Mechanisms; active; renal; repair; Single

Acute kidney injury (AKI), an abrupt loss of kidney function, is an increasing global health burden associated with high morbidity and mortality, for which no targeted therapy exits to date. Common causes of AKI are renal hypoperfusion, sepsis and nephrotoxicity. They result in an acute tubular injury and loss of kidney function, while simultaneously triggering the intrinsic repair processes of the kidney. Tubule cells that survive the acute insult re-enter the cell cycle and proliferate to regenerate the epithelial cells that are essential for the recovery of kidney function. Yet, surprisingly, tubule cells can also become maladaptive and contribute to the development of chronic kidney disease by continuous profibrotic signaling. The development of therapeutics to prevent such maladaptive processes is currently hampered by an insufficient understanding of the underlying molecular mechanisms. Therefore, the aim of the proposed project is to obtain a comprehensive understanding of the molecular underpinnings of renal repair after AKI at a single-cell resolution and to identify key regulatory factors governing adaptive and maladaptive repair processes. We will characterize both chromatin accessibility and associated gene expression of all nephron cells at various time points after both mild and severe AKI in transgenic mice, using a novel combination of single-nucleus ATAC- and single-nucleus RNA-sequencing. In addition, we will acquire the same data from maladaptive tubule cells specifically by isolating their nuclei based on the expression of a marker gene. Together, these data sets will provide a comprehensive regulatory map of the nephron at various stages after mild and severe AKI. We will further analyze these data using state-of-the-art bioinformatic tools to reconstruct the single-cell trajectories that lead to both maladaptive and adaptive tubule cells. From these trajectories, we can identify the pivotal regulatory factors determining both reparative and maladaptive repair processes. We will employ additional methods like RNAScope in situ hybridization and immunostaining of the kidney to study these regulatory factors in even greater detail. Ultimately, the goal of this study is to achieve a detailed understanding of renal repair processes that could facilitate targeted drug development to treat AKI in the future.

急性肾损伤（AKI）是肾功能的突然丧失，是一种日益严重的全球健康负担，发病率和死亡率很高，迄今为止尚无针对性的治疗方法。 AKI 的常见原因是肾脏灌注不足、败血症和肾毒性。它们会导致急性肾小管损伤和肾功能丧失，同时触发肾脏的内在修复过程。在急性损伤中幸存下来的肾小管细胞重新进入细胞周期并增殖以再生对肾功能恢复至关重要的上皮细胞。然而，令人惊讶的是，肾小管细胞也可能变得适应不良，并通过持续的促纤维化信号传导导致慢性肾病的发展。目前，由于对潜在的分子机制了解不足，阻碍了预防这种适应不良过程的治疗方法的发展。因此，该项目的目的是在单细胞分辨率下全面了解 AKI 后肾脏修复的分子基础，并确定控制适应性和适应不良修复过程的关键调节因素。我们将使用单核 ATAC 和单核 RNA 测序的新型组合，描述转基因小鼠轻度和重度 AKI 后不同时间点所有肾单位细胞的染色质可及性和相关基因表达。此外，我们将从适应不良的小管细胞中获取相同的数据，特别是根据标记基因的表达分离其细胞核。这些数据集将共同提供轻度和重度 AKI 后不同阶段肾单位的全面调控图。我们将使用最先进的生物信息学工具进一步分析这些数据，以重建导致适应不良和适应性小管细胞的单细胞轨迹。从这些轨迹中，我们可以确定决定修复和适应不良修复过程的关键调节因素。我们将采用 RNAScope 原位杂交和肾脏免疫染色等其他方法来更详细地研究这些调节因素。最终，这项研究的目标是详细了解肾脏修复过程，以促进未来治疗 AKI 的靶向药物开发。