A Cross-scale Causal Mapping Framework Pinpoints Macrophage Orchestrators of Balanced Arterial Development

Abstract

Postnatal pulmonary arteries experience an abrupt surge in flow that demands tightly coordinated remodeling of vascular structure and mechanical compliance. However, the cellular programs orchestrating these multiscale adaptation remain poorly defined, partly due to the absence of analytical tools integrating gene-cell-tissue scales. Here, we present CausaLink, a cross-scale causal mapping framework that predicts how altered gene expression propagates through gene-cell-tissue networks by integrating time-course transcriptomic and tissue morpho-mechanical data. We constructed a single-cell transcriptomic atlas of 11,143 proximal pulmonary artery-derived cells from C57BL/6J mice across five developmental stages (P2, P10, P21, P42, and P84), revealing dynamic lineage transitions and transient mesenchymal population specifying into fibroblasts and smooth muscle cells from P2 to P21. Using this framework, we identified Mgl2 macrophages as central regulators promoting lumen expansion while preventing pathological wall thinning or thickening. To validate these predictions, we established a human induced pluripotent stem cell-derived arterial assembloid enriched in MGLhigh macrophages. In this model, MGLhigh macrophages supported lumen enlargement while maintaining overall wall thickness and induced formation of an adventitia-like fibroblast layer that recapitulated approximately 80% of the native adventitial fraction with balanced collagen-elastin remodeling. By linking predictive modeling to human organoid validation, this study establishes a cross-scale workflow for tracing how gene programs shape vascular architecture, offering mechanistic insights and a foundation for predictive regenerative medicine.