Wilms' tumor 1 impairs apoptotic clearance of fibroblasts in distal fibrotic lung lesions

Idiopathic pulmonary fibrosis (IPF) is a fatal fibrotic lung disease characterized by impaired fibroblast clearance, accumulation, and excessive extracellular matrix (ECM) protein production. Wilms' Tumor 1 (WT1), a transcription factor, is selectively upregulated in IPF fibroblasts. However, the mechanisms by which WT1 contributes to fibroblast accumulation and ECM production remain unknown. Here, we investigated the heterogeneity of WT1-expressing fibroblasts using single-nucleus RNA sequencing on the distal lung tissues of IPF patients and healthy controls. WT1 was selectively upregulated in a subset of IPF fibroblasts that co-expressed several pro-survival genes. Both the loss-of-function and gain-of-function studies support the idea that WT1 functions as a positive regulator of multiple pro-survival genes to impair apoptotic clearance and promote ECM production. In support, fibroblast-specific overexpression of WT1 augmented fibroproliferation, myofibroblast accumulation, and ECM production during bleomycin-induced pulmonary fibrosis in both young and old mice. Together, these findings identify WT1 as a potential therapeutic target to attenuate fibroblast expansion, and ECM production in the distal areas of fibrosing lungs. Overall design: snRNA-Seq was performed on human lung tissue samples retreived to characterize all cell populations for the first time with nuclear RNA. CST method was used to isolate the nuclei from tissue samples from both IPF and healthy lung tissues and were processed according to instructions provided by the instrument manufacturer.

特发性肺纤维化（Idiopathic Pulmonary Fibrosis, IPF）是一种致命性纤维化性肺部疾病，以成纤维细胞清除障碍、聚集及细胞外基质（extracellular matrix, ECM）蛋白过度产生为核心特征。肾母细胞瘤1（Wilms' Tumor 1, WT1）作为一种转录因子，在IPF患者的成纤维细胞中呈现选择性上调。然而，WT1促进成纤维细胞聚集与ECM产生的具体分子机制仍未阐明。本研究通过对IPF患者与健康对照的远端肺组织开展单细胞核RNA测序，探究了表达WT1的成纤维细胞的异质性。结果显示，WT1在一群共表达多种促生存基因的IPF成纤维细胞亚群中选择性上调。功能缺失与功能获得性实验均证实，WT1可作为多种促生存基因的正向调控因子，抑制细胞凋亡清除并促进ECM产生。进一步验证表明，在年轻与老年小鼠的博莱霉素诱导性肺纤维化模型中，成纤维细胞特异性过表达WT1可增强成纤维细胞增殖、肌成纤维细胞聚集及ECM产生。综上，本研究证实WT1可作为潜在治疗靶点，用于抑制纤维化肺脏远端区域的成纤维细胞扩增与ECM产生。实验设计概述：本研究对获取的人类肺组织样本进行单细胞核RNA测序，首次通过核RNA全面表征其所有细胞群体。采用CST法从IPF患者与健康肺组织的样本中分离细胞核，并严格按照仪器制造商提供的操作指南完成后续样本处理。