A critical role of vascular endothelium is as a semi-permeable barrier, dynamically regulating the flux of solutes between blood and the surrounding tissue. Existing platforms that quantify endothelial function in vitro are either significantly throughput limited or overlook physiologically relevant extracellular matrix (ECM) interactions and thus do not recapitulate in vivo function. Leveraging droplet microfluidics, we developed a scalable platform to measure endothelial function in nanoliter-volume, ECM-based microtissues. In this study, we describe our high-throughput method for fabricating endothelial-coated collagen microtissues that incorporate physiologically relevant cell-ECM interactions. We showed that the endothelial cells had characteristic morphology, expressed tight junction proteins, and remodeled the ECM via compaction and deposition of basement membrane. We also measured macromolecular permeability using two optical modalities, and found the cell layers: (1) had permeability values comparable to in vivo measurements and (2) were responsive to physiologically-relevant modulators of endothelial permeability (TNF-α and TGF-β). This is the first demonstration, to the authors' knowledge, of high-throughput assessment (n > 150) of endothelial permeability on natural ECM. Additionally, this technology is compatible with standard cell culture equipment (e.g. multi-well plates) and could be scaled up further to be integrated with automated liquid handling systems and automated imaging platforms. Overall, this platform recapitulates the functions of traditional transwell inserts, but extends application to high-throughput studies and introduces new possibilities for interrogating cell-cell and cell-matrix interactions.
Bibliographical noteFunding Information:
We thank the American Heart Association (Scientist Development Grant 13SDG6450000), the National Heart, Lung, and Blood Institute (NHLBI R21 HL132256), the National Institute of Environmental Health Sciences (NIEHS R21 ES027622), the National Science Foundation (CBET 1704332) for support. ALC was supported by the National Science Foundation GRFP (Grant 00039202). Portions of this work were conducted in the Minnesota Nano Center, which is supported by the National Science Foundation through the National Nano Coordinated Infrastructure Network (NNCI) under Award Number ECCS-1542202. We also thank Dr Paolo Provenzano for the use of his multiphoton microscope, and Julia Nguyen and Dr Gregory Vercellotti for isolating and generously donating HUVECs.
© 2018 The Royal Society of Chemistry.