Intrinsic Gata4 expression sensitizes the aortic root to dilation in a Loeys-Dietz syndrome mouse model

Loeys-Dietz syndrome (LDS) is a hereditary aneurysm disorder caused by mutations that impair transforming growth factor-ß (TGF-ß) signaling. Although LDS patients develop aneurysms throughout the arterial tree, the aortic root is a site of increased risk. In order to identify molecular determinants of this regional vulnerability, we investigated the transcriptional heterogeneity of vascular smooth muscle cells (VSMCs) in the aorta of Tgfbr1M318R/+ LDS mouse models by single cell RNA sequencing (scRNAseq) and spatial transcriptomics. Downregulation of transcripts coding for components of the extracellular matrix-receptor mechanosensing apparatus and upregulation of transcripts related to stress and inflammation were observed in all Tgfbr1M318R/+ VSMCs. However, regardless of genotype, a subset of Gata4-expressing VSMCs predominantly located in the aortic root intrinsically displayed a less differentiated, pro-inflammatory transcriptional profile. A similar population was also identified in a published scRNAseq dataset of the aorta of LDS patients via the Coordinated Gene Activity in Pattern Sets (CoGAPS)/ProjectR pipeline. Postnatal VSMC-specific Gata4 deletion resulted in reduced aortic root dilation in LDS mice, in association with decreased levels of Agtr1a and other pro-inflammatory regulators. We propose that widespread dysregulation of mechanosensitive pathways may act on regionally restricted factors that “prime” specific aortic locations to increased risk of aneurysm. Overall design: Tgfbr1+/+ and Tgfbr1M318R/+ (The Jackson Laboratory, strain #036511) mice, some bearing the EGFP-L10a (The Jackson Laboratory, strain #024750) conditional tracer allele and a CNC-specific CRE recombinase expressed under the control of Wnt2 promoter (The Jackson Laboratory, strain #003829) were used for scRNAseq at 16 weeks of age. All mice were maintained on a 129-background strain (Taconic, 129SVE).

洛伊迪茨综合征（Loeys-Dietz syndrome, LDS）是一类由损伤转化生长因子-β（transforming growth factor-β, TGF-β）信号通路的突变所引发的遗传性动脉瘤疾病。尽管LDS患者的全身动脉树均可发生动脉瘤，但主动脉根部是风险升高的病变位点。为鉴定该区域易感性的分子决定因素，我们通过单细胞RNA测序（single cell RNA sequencing, scRNAseq）与空间转录组学，探究了Tgfbr1M318R/+ LDS小鼠模型主动脉中血管平滑肌细胞（vascular smooth muscle cells, VSMCs）的转录异质性。
在所有Tgfbr1M318R/+ VSMCs中，均观察到编码细胞外基质-受体机械感知装置组分的转录本表达下调，而与应激及炎症相关的转录本表达上调。然而，无论基因型如何，一群主要定位于主动脉根部、表达Gata4的VSMCs亚群，其转录谱本就呈现低分化、促炎症的特征。通过模式集协调基因活性分析（Coordinated Gene Activity in Pattern Sets, CoGAPS）/ProjectR分析流程，我们在已发表的LDS患者主动脉scRNAseq数据集中也鉴定到了相似的细胞亚群。
出生后特异性在VSMCs中敲除Gata4，可减轻LDS小鼠的主动脉根部扩张，该表型与Agtr1a及其他促炎症调控因子的表达水平降低相关。我们提出假说：机械敏感通路的广泛失调，可通过作用于区域限制性因子，“预致敏”特定主动脉部位，使其动脉瘤风险升高。
总体实验设计：本研究使用16周龄的Tgfbr1+/+与Tgfbr1M318R/+小鼠（杰克逊实验室，品系编号#036511）开展scRNAseq实验，部分小鼠携带EGFP-L10a条件示踪等位基因（杰克逊实验室，品系编号#024750），以及由Wnt2启动子调控表达的CNC特异性CRE重组酶（杰克逊实验室，品系编号#003829）。所有小鼠均饲养于129背景品系（塔科尼公司，129SVE）中。