Europe's population is aging, with declining health conditions among older groups. Studies have linked aging mechanisms to cardiovascular diseases, the leading cause of death worldwide. However, aging mechanisms vary between species, making animal model data hard to translate to humans. A 3D organ-specific model of the human aging microvasculature is a valuable tool to study these mechanisms, identify biomarkers, and accelerate therapeutic development. Our goal is to design a high-throughput (HT) platform embedding human microvascular networks (MVNs) to study basic mechanisms of vascular aging.

We microfabricated a HT platform hosting up to 32 samples through digital-light-processing 3D printing. The system allows to generate measurable, unidirectional flows through integration with a custom rocker. It is compatible with automated seeding of millimeter-size 3D hydrogels, culturing under physiological flow, and parallel analysis via high-content confocal imaging and RNAseq. MVNs were biofabricated by seeding human microvascular endothelial cells (ECs) and fibroblasts obtained from biopsies of old/young donors into 3D fibrin hydrogels. MVN self-assembly and stabilization was promoted with factors like VEGF-A and monodirectional interstitial flow.

We isolated and sorted human ECs and fibroblasts from dermal biopsies and confirmed their identity by analyzing CD31, CD49, PDPN, CD45 and NG2. By optimizing device architecture, and seeding conditions (e.g. cell density, matrix concentration, flow rate), we obtained self-assembled MVNs with physiological diameters compatible with arterioles/venules and capillaries (5-30 micrometers). Perfusability tests demonstrated flow of 70kDa dextran through the lumens of the MVN spanning the entire matrix (1200 micrometers). Confocal Images showed asymmetry of the MVN, suggesting that monodirectional flow is affecting EC behavior. Validation of aging markers (e.g. p16, H2AX, SIRT1) and MVN architecture against dermal biopsies from donors of different age, as well as treatments to counteract aging-related endothelial dysfunction are ongoing.

This work introduces a novel HT platform coupled with a mesoscale 3D model of functional organotypic MVNs. Our robust in vitro system is suitable for studying vascular aging, laying the foundation for targeted therapeutic strategies, and accelerated clinical translation.