The vascular endothelium forms the inner lining of blood vessels and actively regulates vascular permeability in response to chemical and physical stimuli. Understanding the molecular pathways and mechanisms that regulate the permeability of blood vessels is of critical importance for developing therapies for cardiovascular dysfunction and disease. Recently, we developed a novel microfluidic human engineered microvessel (hEMV) platform to enable controlled blood flow through a human endothelial lumen within a physiologic 3D extracellular matrix (ECM) into which pericytes and other stromal cells can be introduced to recapitulate tissue-specific microvascular physiology. This protocol describes how to design and fabricate the silicon hEMV device master molds (takes ~1 week) and elastomeric substrates (takes 3 d); how to seed, culture, and apply calibrated fluid shear stress to hEMVs (takes 1–7 d); and how to assess vascular barrier function (takes 1 d) and perform immunofluorescence imaging (takes 3 d).
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The datasets generated during and/or analyzed during the current study are available from the corresponding author upon reasonable request.
The custom code used for the current study is provided in the Supplementary Data.
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This work was supported in part by grants from the National Institutes of Health (EB00262, EB08396, UH3EB017103, and HL115553) and the National Science Foundation Center for Engineering MechanoBiology (CMMI15-48571). W.J.P. acknowledges support from a Ruth L. Kirchstein National Research Service Award (F32 HL129733) and from the NIH through the Organ Design and Engineering Training program (T32 EB16652); M.L.K. acknowledges support from the Hartwell Foundation and from the National Institutes of Health (K99-CA226366-01A1); and J.B.T. acknowledges support from the NIH through the Translational Research in Biomaterials Training Program (T32 EB006359).
The authors declare no competing interests.
Publisher’s note: Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.
Key references using this protocol
Polacheck, W. J. et al. Nature 552, 258–262 (2017): https://doi.org/10.1038/nature24998
Alimperti, S. et al. Proc. Natl. Acad. Sci. USA 114, 8758–8763 (2017): https://doi.org/10.1073/pnas.1618333114
McCurley, A. et al. J. Am. Soc. Nephrol. 28, 1741–1752 (2017): https://doi.org/10.1681/ASN.2016020200
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Polacheck, W.J., Kutys, M.L., Tefft, J.B. et al. Microfabricated blood vessels for modeling the vascular transport barrier. Nat Protoc 14, 1425–1454 (2019). https://doi.org/10.1038/s41596-019-0144-8
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