Hepatic stellate cell plasticity and maladaptive fibrogenic memory in chronic liver disease

项目总结 慢性肝病患者肝星状细胞可塑性与不良适应纤维形成记忆 与慢性肝病相关的纤维化影响着全球数亿患者。肝星状 细胞(HSCs)反过来又代表肝纤维化的主要细胞驱动因素。一个关键的悬而未决的问题是 肝星状细胞激活以促进组织修复以应对急性肝损伤，但过度激活以产生旺盛的能量 细胞外基质对反复肝损伤的反应，导致纤维化。这种行为切换的核心是一个 HSC记住以前的损伤事件以做出不同反应的机制，即过度激活， 当暴露在再次伤害中时。最近的研究，包括我们(Wang等人)Dev Cell 2019)，强烈支持 表观基因组，特别是DNA甲基化模式作为发育和组织中细胞记忆的载体 受伤。这项研究的目的是阐明表观基因组，特别是DNA甲基化模式 编码这种适应不良的细胞记忆，并在再次损伤后放大HSC的纤维化反应。这项建议 建立在单细胞技术和低输入染色质图谱的最新进展基础上，我们对其进行了优化 广泛检测HSC基因在体内的表达和表观基因组的变化 纤维性记忆。在我们的纤维化记忆模型中，肝干细胞在纤维化消退后完全失活， 由于它们的转录组与未损伤的HSCs难以区分，然而，表观基因组的变化仍然存在 染色质可及性的变化。作为对再损伤的反应，退行性肝脏的HSCs高度激活，并 由独特的转录网络(WT1、TEAD1、Tbx20和Pbx1)驱动，在经历初始阶段的HSC中没有发现 受伤。使用我们之前在Dev Cell试纸中生成的uhrf1牙线小鼠专门删除uhrf1，a DNA甲基化机制的关键组成部分，我们发现在HSCs中，这些小鼠表现出增强 纤维性记忆对再损伤的反应。我们的中心假设是通过对先前受伤的记忆 表观遗传改变改变了HSC的可塑性，并放大了它们的激活，以应对再次损伤。我们将解决 这一假设有三个相互关联但又截然不同的具体目的：1)定义控制 HSCs的纤维化记忆；2)确定uhrf1和DNA甲基组是如何控制纤维化记忆的 HSC；3)发现新的调控节点，驱动HSC在再损伤中的不良适应反应。这是一项创新 利用尖端基因组学技术和独特的动物模型的方法意义重大，因为它将 通过揭示纤维化记忆的表观遗传学基础，对星状细胞生物学产生新的基本见解， 特定基因和转录网络在纤维化记忆和肝纤维化中的作用 在肝病患者中保守的致纤维化驱动因素，可以导致潜在的治疗靶点。

PROJECT SUMMARY Hepatic stellate cell plasticity and maladaptive fibrogenic memory in chronic liver disease Fibrosis associated with chronic liver disease affects hundreds of millions of patients worldwide. Hepatic stellate cells (HSCs), in turn, represent the main cellular driver of hepatic fibrosis. A key unanswered question is why HSCs activate to facilitate tissue repair in response to acute liver injury but hyperactivate to produce exuberant extracellular matrix in response to repeated liver injury, leading to fibrosis. Central to this behavior switch is a mechanism for HSCs to remember previous injury episodes in order to respond differently, i.e. hyperactivate, when exposed to re-injury. Recent studies, including ours (Wang et al. Dev Cell 2019), strongly support the epigenome and in particular DNA methylation patterns as the carrier of cell memory in development and in tissue injury. The objective of this research is to clarify how the epigenome and specifically DNA methylation patterns encode this maladaptive cell memory and amplify HSC’s fibrogenic response following re-injury. This proposal builds upon recent advances in single cell technology and low-input chromatin profiling which we optimized extensively to measure HSC gene expression and epigenomic changes in vivo in a novel mouse model of fibrogenic memory. In our fibrogenic memory model, HSCs completely deactivate following fibrosis regression, with their transcriptome indistinguishable from uninjured HSCs, however epigenomic changes persist in the form of chromatin accessibility changes. In response to re-injury, HSCs from regressed liver hyperactivate and are driven by unique transcriptional networks (WT1, TEAD1, TBX20, and PBX1) not found in HSCs undergoing initial injury. Using the Uhrf1 floxed mice generated previously in our Dev Cell paper to specifically remove Uhrf1, a critical component of the DNA methylation machinery, in HSCs, we found that these mice display augmented fibrogenic memory in response to re-injury. Our central hypothesis is that memory of previous injury through epigenetic changes modify HSC plasticity and amplify their activation in response to re-injury. We will address this hypothesis in three interrelated, but distinct specific aims:1) Define the regulatory elements controlling fibrogenic memory in HSCs; 2) Determine how UHRF1 and the DNA methylome control fibrogenic memory in HSCs; 3) Uncover novel regulatory nodes driving HSC’s maladaptive response in re-injury. This innovative approach leveraging cutting-edge genomics technology and unique animal models is significant because it will yield fundamental new insights into stellate cell biology by uncovering the epigenetic basis of fibrogenic memory, the contribution of specific genes and transcriptional networks to fibrogenic memory and hepatic fibrosis, and conserved fibrogenic drivers in liver disease patients that can lead to potential therapeutic targets.