单血管转录组图谱描绘病理景观并揭示NR2F2介导的毛细血管畸形中平滑肌细胞表型获得。
Single-vessel transcriptome map pathological landscapes and reveal NR2F2-mediated smooth muscle cell phenotype acquisition in capillary malformations.
作者信息
Nguyen Vi, Mao Irving, He Siwuxie, Castellanos Isabella, Azuero Mackenzie, Hochman Marcelo L, Rong Yin, Pernomian Laena, Chen Elliott H, Friedman Harold I, Chen Yan-Hua, Lu Qun, Fan Daping, Wenceslau Camilla F, Chen Dong-Bao, Nelson J Stuart, Jegga Anil G, Wang Yunguan, Tan Wenbin
机构信息
Department of Cell Biology and Anatomy, School of Medicine, University of South Carolina, Columbia, South Carolina 29209, USA.
Department of Pediatrics, University of Cincinnati College of Medicine, Cincinnati, Ohio 45229, USA.
出版信息
bioRxiv. 2025 Sep 7:2025.09.02.673874. doi: 10.1101/2025.09.02.673874.
BACKGROUND
Capillary malformation (CM) is a congenital vascular anomaly affecting the skin, mucosa, and brain, yet the understanding of its vascular pathogenesis remains limited.
METHODS
We applied spatial whole-transcriptome profiling (GeoMx) and gene set enrichment analysis within CM lesions at single vasculature level. Differentially expressed genes were validated by immunofluorescence staining. Phosphoproteomics was profiled to uncover lesion-wide phosphorylation sites on proteins. Single-cell RNA sequencing was performed on CM-derived induced pluripotent stem cells (iPSCs) to determine differentiation trajectories of lesional vascular lineages. In silico gene perturbation was used to predict candidate genes for modulating vascular pathological progression, followed by functional validation in CM iPSC-derived endothelial cells (ECs) using a Tet-on system.
RESULTS
A spatial transcriptomic atlas was constructed, and pathological landscape of individual CM vasculature was delineated. CM vessels exhibited hallmarks of endothelial-to-mesenchymal transition (EndMT), including disruption of adherens junctions (AJs), vascular identity transitions, and metabolic remodeling. Phosphoproteomics confirmed that differentially phosphorylated proteins were enriched in EndMT- and AJ-related pathways. Aberrant expression of venous transcriptional factor NR2F2 was observed in lesional ECs and correlated with progressive enlargement from capillaries to larger-caliber vessels containing multiple layers of smooth muscle cells (SMCs). In CM iPSCs, differentiation course yielded reduced ECs but increased SMCs. knockout simulation predicted NR2F2 as a crucial regulator of facilitating SMC phenotype in CM. Consistently, enforced NR2F2 expression during iPSC differentiation suppressed endothelial markers while inducing SMC-associated genes.
CONCLUSIONS
Single CM vasculature displays pathological hallmarks characterized by EndMT and AJ disruption, leading to progressive vascular remodeling. NR2F2 functions as a central regulatory factor orchestrating the acquisition of the SMC phenotype, thereby representing a potential therapeutic target in CM.
背景
毛细血管畸形（CM）是一种影响皮肤、黏膜和大脑的先天性血管异常，但对其血管发病机制的了解仍然有限。
方法
我们在单个脉管系统水平对CM病变应用空间全转录组分析（GeoMx）和基因集富集分析。通过免疫荧光染色验证差异表达基因。进行磷酸化蛋白质组学分析以揭示病变范围内蛋白质的磷酸化位点。对CM来源的诱导多能干细胞（iPSC）进行单细胞RNA测序，以确定病变血管谱系的分化轨迹。利用计算机基因扰动预测调节血管病理进展的候选基因，随后使用Tet-on系统在CM iPSC来源的内皮细胞（EC）中进行功能验证。
结果
构建了一个空间转录组图谱，并描绘了单个CM脉管系统的病理格局。CM血管表现出内皮向间充质转化（EndMT）的特征，包括黏附连接（AJ）的破坏、血管身份转变和代谢重塑。磷酸化蛋白质组学证实差异磷酸化蛋白在EndMT和AJ相关途径中富集。在病变EC中观察到静脉转录因子NR2F2的异常表达，且与从毛细血管到含有多层平滑肌细胞（SMC）的较大口径血管的逐渐扩大相关。在CM iPSC中，分化过程产生的EC减少但SMC增加。基因敲除模拟预测NR2F2是促进CM中SMC表型的关键调节因子。一致地，在iPSC分化过程中强制表达NR2F2可抑制内皮标记物，同时诱导SMC相关基因。
结论
单个CM脉管系统表现出以EndMT和AJ破坏为特征的病理特征，导致进行性血管重塑。NR2F2作为协调SMC表型获得的核心调节因子发挥作用，因此是CM的一个潜在治疗靶点。
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