Adrenomedullin Enhances Vascular Integrity and Endothelial Cell Maturation in Bioengineered Lung Models

Rationale: The study of pulmonary vasculature diseases has been hindered by the limitations of current ex vivo vascular models, which often fail to accurately replicate the functionality and maturity of native vasculature. Paracrine signaling plays a key role in regulating endothelial maturation and maintaining vascular homeostasis, yet the understanding of these signals in engineered vascular systems remains incomplete. In this study, we investigated how adrenomedullin (ADM), a paracrine factor enriched in the lung vascular niche, can improve endothelial cell maturity and vascular barrier function in bioengineered lung models. Methods: Single-cell RNA sequencing (scRNAseq) was utilized to profile ligand-receptor (L/R) interactions in the human lung, mapping paracrine signaling within the microvascular niche. We then evaluated the impact of these ligands on vascular barrier function and inflammatory response using in vitro assays, including electrical cell impedance sensing (ECIS) and transwell assays. ADM was further tested under physiological conditions in both microfluidic chips and whole-organ bioengineered lung vascular models to assess its impact on endothelial cell integrity. Results: We applied Connectome to map the averaged gene expression per cell type to known L/R interactions cataloged in the NicheNet database. Using microvascular ECs as signal receivers (Senders) and the putative adjacent cell types in the microvascular niche as signal senders (Ligands), we identified several specific and abundant L/R pairs based on their expression levels in the niche, including SLIT2-ROBO4, ANGPT1-TEK, ADM-RAMP2, VEGFD-VEGFR2, BMP5-BMPR2, etc. Among the top 17 soluble ligands identified, ADM, VEGFD, and Angiopoietin 1 (ANGPT1) significantly increased endothelial cell barrier function, with ADM showing the strongest effect. Also, ADM mitigated TNF-a induced ICAM1 expression and IκBα degradation, indicating an anti-inflammatory response. The receptor for ADM, RAMP2, was highly expressed in both microfluidic and bioengineered lung platforms. Under physiological shear stress (15 dynes/cm 2 ), ADM further prolonged enhanced vascular resistance, and two days of shear stress upregulated EDNRB (2.89-fold compared with shear stress with vehicle), an aerocyte endothelial cell marker, while one day shear stress had no effect on ADM treatment. In the endothelial repopulated lung vascular models, treatment of ADM could improve cellular phenotype toward native lung features and further enhance vascular barrier functions. Conclusions: This study demonstrates that native paracrine cues, such as ADM, can significantly enhance endothelial cell maturation and vascular integrity in bioengineered lung models with physical fluid flow. The findings provide a promising avenue for developing advanced in vitro models that closely mimic native pulmonary vasculature, ultimately contributing to improved disease modeling and therapeutic development. R00HL159261 (YY) This abstract was presented at the American Physiology Summit 2025 and is only available in HTML format. There is no downloadable file or PDF version. The Physiology editorial board was not involved in the peer review process.