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Multivascular networks and functional intravascular topologies within biocompatible hydrogels

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Science  03 May 2019:
Vol. 364, Issue 6439, pp. 458-464
DOI: 10.1126/science.aav9750
  • Fig. 1 Monolithic hydrogels with functional intravascular topologies.

    (A) Monolithic hydrogels with a perfusable channel containing integrated fin elements of alternating chirality. These static elements rapidly promote fluid dividing and mixing (as shown by fluorescence imaging), consistent with a computational model of flow (scale bars, 1 mm). (B) Hydrogels with a functional 3D bicuspid valve integrated into the vessel wall under anterograde and retrograde flows (scale bars, 500 μm). Particle image velocimetry demonstrates stable mirror image vortices in the sinus region behind open valve leaflets.

  • Fig. 2 Entangled vascular networks.

    (A to D) Adaptations of mathematical space-filling curves to entangled vessel topologies within hydrogels (20 wt % PEGDA, 6 kDa): (A) axial vessel and helix, (B) interpenetrating Hilbert curves, (C) bicontinuous cubic lattice, and (D) torus and (3,10) torus knot (scale bars, 3 mm). (E) Tessellation of the axial vessel and its encompassing helix along a serpentine pathway. The photograph is a top-down view of a fabricated hydrogel with oxygen and RBC delivery to respective vessels. During perfusion, RBCs change color from dark red (at the RBC inlet) to bright red (at the RBC outlet) (scale bar, 3 mm). Boxed regions are magnified in (F) (scale bar, 1 mm). (G) Perfused RBCs were collected at the outlet and quantified for So2 and Po2. Oxygen flow increased So2 and Po2 of perfused RBCs compared with deoxygenated RBCs perfused at the inlet (dashed line) and a nitrogen flow negative control (N ≥ 3 replicates, data are mean ± SD, *P < 2 × 10−7 by Student’s t test).

  • Fig. 3 Tidal ventilation and oxygenation in hydrogels with vascularized alveolar model topologies.

    (A) (Top) Architectural design of an alveolar model topology based on a Weaire-Phelan 3D tessellation and topologic offset to derive an ensheathing vasculature. (Bottom) Cutaway view illustrates the model alveoli (alv.) with a shared airway atrium. Convex (blue) and concave (green) regions of the airway are highlighted. (B) Photograph of a printed hydrogel during RBC perfusion while the air sac was ventilated with O2 (scale bar, 1 mm). (C) Upon airway inflation with oxygen, concave regions of the airway (dashed black circles) squeeze adjacent blood vessels and cause RBC clearance (scale bar, 500 μm). (D) A computational model of airway inflation demonstrates increased displacement at concave regions (dashed yellow circles). (E) Oxygen saturation of RBCs increased with decreasing RBC flow rate (N = 3, data are mean ± SD, *P < 9 × 10−4 by Student’s t test). The dashed line indicates So2 of deoxygenated RBCs perfused at the inlet. (F) Elaboration of a lung-mimetic design through generative growth of the airway, offset growth of opposing inlet and outlet vascular networks, and population of branch tips with a distal lung subunit. (G) The distal lung subunit is composed of a concave and convex airway ensheathed in vasculature by 3D offset and anisotropic Voronoi tessellation. (H) Photograph of a printed hydrogel containing the distal lung subunit during RBC perfusion while the air sac was ventilated with O2 (scale bar, 1 mm). (I) Threshold view of the area enclosed by the dashed box in (H) demonstrates bidirectional RBC flow during ventilation. (J) Distal lung subunit can stably withstand ventilation for more than 10,000 cycles (24 kPa, 0.5 Hz) and demonstrates RBC sensitivity to ventilation gas (N2 or O2).

  • Fig. 4 Engraftment of functional hepatic hydrogel carriers.

    (A to C) Albumin promoter activity was enhanced in hydrogel carriers containing hepatic aggregates after implantation in nude mice. Data from all time points for each condition are shown in (B) [N = 4, *P < 0.05 by two-way analysis of variance (ANOVA) followed by Tukey’s post-hoc test]. Cumulative bioluminescence for each condition is shown in (C) (N = 4, *P < 0.05 by one-way ANOVA followed by Tukey’s post-hoc test). Error bars indicate SEM. GelMA, gelatin methacrylate. (D) Gross images of hydrogels upon resection (scale bars, 5 mm). (E) (Left) Prevascularized hepatic hydrogel carriers are created by seeding endothelial cells (HUVECs) in the vascular network after printing. (Right) Confocal microscopy observations show that hydrogel anchors physically entrap fibrin gel containing the hepatocyte aggregates (Hep) (scale bar, 1 mm). (F) Hepatocytes in prevascularized hepatic hydrogel carriers exhibit albumin promoter activity after implantation in mice with chronic liver injury. Graft sections stained with H&E show positioning of hepatic aggregates (black arrows) relative to printed (case, anchor) and nonprinted (fibrin) components of the carrier system (scale bar, 50 μm). (G) Hydrogel carriers are infiltrated with host blood (gross, H&E). Carriers contain aggregates that express the marker cytokeratin-18 (Ck-18) and are in close proximity to Ter-119–positive RBCs (scale bars, 40 μm).

Supplementary Materials

  • Multivascular networks and functional intravascular topologies within biocompatible hydrogels

    Bagrat Grigoryan, Samantha J. Paulsen, Daniel C. Corbett, Daniel W. Sazer, Chelsea L. Fortin, Alexander J. Zaita, Paul T. Greenfield, Nicholas J. Calafat, John P. Gounley, Anderson H. Ta, Fredrik Johansson, Amanda Randles, Jessica E. Rosenkrantz, Jesse D. Louis-Rosenberg, Peter A. Galie, Kelly R. Stevens, Jordan S. Miller

    Materials/Methods, Supplementary Text, Tables, Figures, and/or References

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    • Materials and Methods
    • Figs. S1 to S15
    • Table S1
    • Captions for Movies S1 to S5
    • References

    Images, Video, and Other Media

    Movie S1
    Movie compilation to help explain the functional intravascular bicuspid valve design and performance. We first illustrate the design of the bicuspid valve, followed by phase contrast imaging and fluorescence imaging videos of anterograde and retrograde flow of fluorescent beads. The valve is open under anterograde flow and closes under retrograde flow. Note also the mirror image vortices in the sinus region during anterograde flows. This movie supplements Fig. 1.
    Movie S2
    Movie compilation of fabricated hydrogels with entangled vascular networks. This movie supplements Fig. 2. Perfusion video of red dye injected into helical channel wrapped around axial vessel filled with blue dye. Rotational video of 1°, 2° Hilbert Curve filled with di↵erent colored dyes into individual channels. Animation of layers used for stereolithographic fabrication of torus model, followed by perfusion of red dye through the (3,10) torus knot. Filling of the channels in the torus model with colored dyes enabled μCT acquisition, reconstruction, and visualization of perfusable channels.
    Movie S3
    Movie compilation illustrating the design and function of the bioinspired alveolar model based on the Weaire-Phelan foam topology. Cyclic ventilation of the airway during RBC perfusion through ensheathing vasculature demonstrates valving of the vasculature by nearby concave airway regions. This movie supplements Fig. 3.
    Movie S4
    Movie compilation illustrating the design and function of the scalable voronoi lungmimetic model. We show an animation explaining the generative lung topology algorithm where an airway is grown, inlet and outlet vasculature track the airway on opposing sides, and the tips of all branches are then populated with the distal lung subunit. Movie footage of ventilation and perfusion experiments are shown of the distal lung subunit as well as color-filtered views of bidirectional blood flow observed during ventilation and perfusion. This movie supplements Fig. 3.
    Movie S5
    Luminescence imaging of fabricated tissue constructs perfused with media containing luciferin substrate and either deoxygenated or oxygenated RBCs. Luminescence of entrapped Luc2P-expressing HEK cells is boosted in tissues perfused with oxygenated RBCs. Images from complete time course shown in fig. S10.

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