Journal: Small methods

Article Title: A Robust Method for Perfusable Microvascular Network Formation In Vitro

doi: 10.1002/smtd.202200143

Figure Lengend Snippet: Different types of MVNs made with two-step method. Representative images of two-step seeding method produced MVNs formed by a) ImHUVECs expressing BFP with patient-derived thyroid FBs. Opening percentage (%) is 67.46 ± 15.95%; central perfusable MVN percentage is 77.67 ± 16.81% (n = 3 devices, 5 ROIs each). Green, ECs. Magenta, dextran. b) Two-step brain MVNs formed with brain ECs expressing GFP with brain pericytes and astrocytes. Opening percentage is 80.80 ± 14.02%; central perfusable MVN percentage is 87.20 ± 12.22% (n = 3 devices, 5 ROIs each). Green, ECs. Magenta, dextran. Yellow, GFAP immunofluorescent staining for activated astrocytes. c) Two-step dermal MVNs formed with human dermal blood microvascular ECs (HDBMECs) expressing BFP and dermal FBs. Opening percentage is 93.20 ± 8.621%; central perfusable MVN percentage is 98.93 ± 4.652% (n = 3 devices, 5 ROIs each). Green, ECs. Magenta, dextran. d) Permeability analysis of two-step dermal MVNs treated with or without TNF- α . Bars represent mean ± SD. Two-tailed t -tests were performed for the statistical comparisons (n = 4 devices, 4 ROIs each). e) Two-step dermal MVNs stained with CD31 antibody (red). f) Two-step method MVNs formed with iPSC-derived ECs and lung FBs. Opening percentage is 77.37 ± 15.03%; central perfusable MVN percentage is 76.47 ± 13.43% (n = 3 devices, 5 ROIs each). Green, iPSC-ECs stained with UEA-I. Magenta, dextran. g) Statistical analysis of permeability of two-step iPSC-EC MVNs treated with or without TNF- α . Bars represent mean ± SD. Two-tailed t -tests were performed for the statistical comparisons (n = 4 devices, 4 ROIs each). h) CD31 immunostaining (red) of two-step MVNs made with iPSC-ECs and lung FBs. In all MVN images, white arrows point to the perfusable narrow microvessels. Dashed rectangles identify regions shown in inserts at higher magnification. 10 kDa dextran was used for brain MVNs perfusion. The 70 kDa dextran was used for all other perfusion and permeability experiments.

Article Snippet: Human brain microvascular endothelial cells (Angioproteomie, P4–P5) were cultured on fibronectin (30 μg mL −1 ) coated flasks in Vasculife VEGF endothelial medium with 5% FBS.

Techniques: Produced, Expressing, Derivative Assay, Staining, Permeability, Two Tailed Test, Immunostaining