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

Postnatal pulmonary arteries experience an abrupt surge in flow that necessitates tightly balanced remodeling of lumen, wall, and compliance. Yet the cellular programs and lineage dynamics orchestrating this coordination remain poorly defined, in part due to the absence of analytical tools capable of integrating gene–cell–tissue scales. Here, we present CausaLink, a cross-scale causal mapping framework that predict 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 spanning five developmental milestones (P2, P10, P21, P42, and P84) by 11,143 proximal pulmonary artery-derived cells from C57BL/6J mice, revealing dynamic vascular lineage transitions. Our analysis highlighted a transient mesenchymal population that undergoes transcriptomic and phenotypic specification into fibroblasts and smooth muscle cells from P2 to P21, coinciding with tunica intima–media–adventitia formation. In this context, CausaLink pinpointed Mgl2⁺ macrophages as central regulator candidates of balanced artery growth. Network-based, multi-trait simulations predicted that Mgl2⁺ macrophages promote lumen expansion while preventing pathological wall thinning, thickening or stiffening. To extend these predictions beyond murine models, we established a human induced pluripotent stem cell (hiPSCs)–derived arterial assembloid enriched with MGLhigh macrophages. In this platform, MGLhigh macrophages supported lumen enlargement while preserving overall wall thickness, accompanied by the formation of an adventitia-like fibroblast layer that recapitulated of the native adventitial fraction. By linking predictive tissue modeling to human-organoid model validation, this study suggests a cross-scale workflow for tracing how gene programs shape vascular architecture, offering mechanistic insights and a foundation for predictive regenerative medicine in diseases of disrupted tissue homeostasis.