Abstract Rationale Recent advances in pulmonary arterial hypertension (PAH) research have leveraged multi-omics approaches to uncover molecular mechanisms that may yield new therapeutic targets. We recently identified potentially protective gene modules through co-expression network analysis of a large PAH lung biobank and experimentally validated one hub gene, ASPN. Building on this framework, we sought to identify and functionally characterize an additional hub gene with potential relevance to PAH pathobiology. Methods Potential hub genes were prioritized based on their correlation with the eigengene of a previously identified PAH-related gene module of 266 genes. Expression of the top candidate was evaluated in human PAH versus control bulk lung tissue, vascular cells in vitro, single-nucleus RNA-seq, and spatial transcriptomics datasets. Gain- and loss-of-function studies were performed in vitro followed by RNA-seq. Differential gene expression was analyzed using DESeq2, and pathway enrichment was assessed with fgsea using the Hallmark gene set collection. Results ANTXR1, a cell-surface receptor implicated in extracellular matrix remodeling and proliferative processes such as cancer, was identified as the top hub gene within the PAH-related gene module. It was upregulated in human PAH lung tissue and in adventitial fibroblasts in vitro, as well as by single-nucleus RNA-seq and spatial transcriptomics. Knockdown of ANTXR1 by siRNA in PAH adventitial fibroblasts increased proliferation as measured by CCK-8 and Ki67, whereas recombinant ANTXR1 treatment reduced proliferation. RNA-seq after ANTXR1 knockdown showed significant negative enrichment of the Hallmark p53 pathway with decreased expression of canonical p53 targets, including CDKN1A (p21) and BTG2. Conclusion These findings identify ANTXR1 as a novel, transcriptionally upregulated hub gene in PAH and implicate it as a modulator of fibroblast proliferation through the p53 pathway. Loss of ANTXR1 suppresses canonical p53 targets, including CDKN1A (p21) and BTG2, consistent with release of a cell-cycle checkpoint brake. Together, the data suggest that ANTXR1 exerts a protective, growth-restraining role in PAH adventitial fibroblasts and highlight the p53-ANTXR1 axis as a potential regulator of vascular remodeling. This abstract is funded by: NHLBI K08HL169982
Thangaratnam et al. (Fri,) studied this question.