Reprogramming of mature SMCs to vascular progenitor cells: Focus on Vascular Fibrosis

摘要 过度的血管周围纤维化是由正常组织修复过程的高度动态和失调引起的 并且由细胞外基质（ECM）材料的过度积累定义，导致细胞外基质（ECM）的减少。 血管顺应性和增加的血管硬度。相比之下，纤维化反应对于形成 和动脉粥样硬化情况下纤维帽的稳定性。虽然我们的研究取得了进展， 在对纤维化病理生物学的理解方面，仍然存在重大差距。特别是，虽然间充质细胞与 转变成活化的肌成纤维细胞的潜力被认为是过度ECM沉积的关键来源， 其起源仍有争议。利用细胞特异性谱系追踪和RNA测序，我们的研究小组制作了 一个改变范式的发现，即在血管内存在的常驻血管干细胞亚群， 外膜来源于成熟的血管平滑肌细胞（SMC）（称为AdvSca 1-SM细胞）。SMC 重编程和AdvSca 1-SM细胞维持依赖于转录的诱导和活性 因子，Klf 4。相反，血管损伤诱导的或AdvSca 1-SM细胞特异性的遗传Klf 4缺失促进了细胞凋亡。 转化为促纤维化肌成纤维细胞，其因此作为血管周围纤维化的主要贡献者。 然而，Klf 4决定AdvSca 1-SM的维持和分化的分子机制， 细胞仍然不清楚。利用高度特异性的AdvSca 1-SM细胞报告系统和单细胞RNA- 通过scRNA-seq测序，我们证明了AdvSca 1-SM细胞的促纤维化分化轨迹。 分化的特征在于Klf 4及其下游效应子PI 16的缺失，PI 16是细胞分化的负调节因子。 表观遗传蛋白，组蛋白去乙酰化酶1（HDAC 1）和lncRNA，Meg 3，但获得的表达促纤维化 表型。重要的是，这些变化在人类纤维化组织中重现，这是翻译的基础。 意义他汀类药物治疗以前被证明可以减少组织纤维化;然而， 不明用AdvSca 1-SM细胞及其促纤维化的基因特征查询连接性图谱数据库 衍生物表明他汀类药物是拮抗AdvSca 1-SM的促纤维化转变的潜在候选药物 细胞最后，虽然炎症和纤维化驱动大多数成年哺乳动物的伤口愈合过程， 已经报道了Acomys物种的成年啮齿动物的再生伤口愈合。我们最近的发现显示 在Acomys的两种肾损伤模型中，完全没有纤维化并快速再生。这里我们 提出Klf 4-Meg 3轴在维持干细胞表型中起核心作用的新概念; Klf 4和/或Meg 3的缺失激活促进AdvSca 1-SM的表型转变的信号级联（Aim 一）。此外，我们认为Acomys对病理性血管重塑和纤维化具有抗性，部分原因是 AdvSca 1-SM特异性再生基因表达标签的激活（目的二）。最后，他汀类药物 治疗将抑制或逆转AdvSca 1-SM细胞的促纤维化转变（目的三）。

ABSTRACT Excessive perivascular fibrosis results from the highly dynamic and dysregulated process of normal tissue repair and is defined by the excessive accumulation of extracellular matrix (ECM) material resulting in decreased vascular compliance and increased vessel stiffness. In contrast, a fibrotic response is critical to the formation and stability of a fibrous cap in the setting of atherosclerosis. While advances have been made in our understanding of fibrosis pathobiology, significant gaps remain. In particular, while mesenchymal cells with the potential to transition into activated myofibroblasts are believed to be key sources of excessive ECM deposition, their origin remain debated. Using cell-specific lineage tracing and RNA sequencing, our group made the paradigm-shifting discovery that a subpopulation of resident vascular stem cells residing within the vascular adventitia originate from mature vascular smooth muscle cells (SMC)(termed AdvSca1-SM cells). SMC reprogramming and AdvSca1-SM cell maintenance is dependent on induction and activity of the transcription factor, Klf4. In contrast, vascular injury-induced or AdvSca1-SM cell-specific genetic Klf4 depletion promote the transition to profibrotic myofibroblasts, which consequently serve as major contributors to perivascular fibrosis. However, the molecular mechanism whereby Klf4 dictates the maintenance and differentiation of AdvSca1-SM cells remains unclear. Leveraging a highly specific AdvSca1-SM cell reporter system and single-cell RNA- sequencing (scRNA-seq), we demonstrate the profibrotic differentiation trajectory of AdvSca1-SM cells. Differentiation is characterized by loss of Klf4 and its downstream effector PI16, a negative regulator of the epigenetic protein, histone deacetylase 1 (HDAC1), and the lncRNA, Meg3, but gain of expression of a profibrotic phenotype. Importantly, these changes were recapitulated in human fibrotic tissue, underlying the translational significance. Statin treatment was previously shown to reduce tissue fibrosis; however, the molecular mechanism is unknown. Querying Connectivity Map database with gene signatures of AdvSca1-SM cells and their profibrotic derivatives indicate that statins are potential candidates to antagonize the profibrotic transition of AdvSca1-SM cells. Finally, while inflammation and fibrosis drive the wound healing process in most adult mammals, scarless regenerative wound healing in adult rodents of the species Acomys has been reported. Our recent findings show a complete absence of fibrosis with rapid regeneration in two models of kidney injury in Acomys. Here, we propose a novel concept that a Klf4-Meg3 axis plays a central role in the maintenance of the stem cell phenotype; loss of Klf4 and/or Meg3 activates a signaling cascade facilitating the phenotypic transition of AdvSca1-SM (Aim One). Further, we propose that Acomys is resistant to pathological vascular remodeling and fibrosis in part due to activation of an AdvSca1-SM-specific regenerative gene expression signature (Aim Two). Finally, statin treatment will inhibit or reverse the profibrotic transition of AdvSca1-SM cells (Aim Three).