RESUMEN
Here, we present a protocol for producing a microfluidic vessel-on-chip platform using human pluripotent stem cell-derived endothelial cells (SC-ECs). We describe steps for manufacturing the 3D-printed chip, cell culturing to generate SC-ECs, hydrogel patterning, and the formation and cultivation of barrier-forming vessels. We then detail procedures for the retrieval of cells and media from the open microfluidic chip platform to enable multi-omics analysis. For complete details on the use and execution of this protocol, please refer to Marder et al.1.
RESUMEN
The metabolic syndrome is accompanied by vascular complications. Human in vitro disease models are hence required to better understand vascular dysfunctions and guide clinical therapies. Here, we engineered an open microfluidic vessel-on-chip platform that integrates human pluripotent stem cell-derived endothelial cells (SC-ECs). The open microfluidic design enables seamless integration with state-of-the-art analytical technologies, including single-cell RNA sequencing, proteomics by mass spectrometry, and high-resolution imaging. Beyond previous systems, we report SC-EC maturation by means of barrier formation, arterial toning, and high nitric oxide synthesis levels under gravity-driven flow. Functionally, we corroborate the hallmarks of early-onset atherosclerosis with low sample volumes and cell numbers under flow conditions by determining proteome and secretome changes in SC-ECs stimulated with oxidized low-density lipoprotein and free fatty acids. More broadly, our organ-on-chip platform enables the modeling of patient-specific human endothelial tissue and has the potential to become a general tool for animal-free vascular research.