Research ArticleHEALTH AND MEDICINE

4D physiologically adaptable cardiac patch: A 4-month in vivo study for the treatment of myocardial infarction

See allHide authors and affiliations

Science Advances  24 Jun 2020:
Vol. 6, no. 26, eabb5067
DOI: 10.1126/sciadv.abb5067
  • Fig. 1 Design of a physiologically adaptable cardiac patch.

    (A) Photograph of the anatomical heart and the fiber structure of the LV visualized by DTI data. (B) Schematic illustration of a short-sectioned LV that illustrates the variation of fiber angle from the epicardium to the endocardium. The orientation (2D mesh pattern projection) of the fiber angles varies continuously with the position across the wall and distribution changes from the apical region to the basal region. (C) Curvature change of cardiac tissue at two different phases (diastole and systole) of the cardiac cycle, which occurs as the heartbeat and pumping blood. (D) CAD design of 3D stretchable architecture on the heart. It provides dynamic stretchability without material deformation or failure when the heart repeatedly contracts and relaxes. (E) Representation of a simplified geometric model of the fibers in the printed object. In the selected region, the angle (α), the length of fiber (L), spatial displacement (D) and the ventricular curvature (κ) are defined with systole (1) and diastole (2) states. (F) Mechanism of the internal stress-induced morphing process. Uneven cross-linking density results in different volume shrinkage after stress relaxation. Photo credit: Haitao Cui, The George Washington University (GWU).

  • Fig. 2 Printing of smart cardiac patch and optimization.

    (A) Curvature change of 4D morphing versus printing speed (means ± SD, n ≥ 6, *P < 0.05). (B) Printing accuracy of the hydrogel patches versus fiber width for different fill density (fd; means ± SD, n ≥ 6, *P < 0.05, **P < 0.01, and ***P < 0.001). (C) Color map of tensile moduli of the patches with varying GelMA and PEGDA concentrations. (D) Optical and 3D surface plot images of the patches. Scale bars, 200 μm. (E) Average elasticity values of the wave-patterned patches in horizontal (x) and vertical (y) directions. Number sign (#) shows the statistical comparison between the horizontal and vertical directions (means ± SD, n ≥ 6, **P < 0.01 and ##P < 0.01). (F) Uniaxial tensile stress-strain curves of 5% GelMA and 15% PEGDA. Immunostaining of cell morphology (F-actin; red), sarcomeric structure (α-actinin; green), gap junction [connexin 43 (Cx43); red], and contractile protein [cardiac troponin I (cTnI); red] on the patches on (G) day 1 and (H) day 7. Scale bars, 20 μm. (I) Beating rate of hiPSC-CMs on the patch and well plate on day 3 and day 7 (means ± SD, n ≥ 6, *P < 0.05; n.s. no significant difference). BPM, beats per minute. Photo credit: Haitao Cui, GWU.

  • Fig. 3 In vitro characterization of the printed cellularized patches.

    Cell distribution of tricultured hiPSC-CMs (green), hECs (red), and hMSCs (blue) on the cardiac patches using cell tracker staining after (A) 1 day of confluence and (B) 7 days of culture. Scale bars, 200 μm. (C) Autofluorescence 3D images of GFP+ hiPSC-CMs on the wave-patterned patch on day 1 and day 7. Scale bars, 100 μm. (D) Immunostaining of capillary-like hEC distribution (CD31; red) on the hydrogel patches. Scale bars, 200 μm. Immunostaining (3D images) of cTnI (red) and vascular protein (vWf; green) on the (E) wave-patterned and (F) mesh-patterned patches. Scale bars, 200 μm (3D image) and 20 μm (2D inset). Calcium transients of hiPSC-CMs on the hydrogel patches recorded on (G) day 3 and (H) day 7. (I) Peak amplitude of the calcium transients of hiPSC-CMs on the mesh- and wave-patterned patches on day 3, day 7, and day 10 (means ± SD, n ≥ 30 cells, *P < 0.05).

  • Fig. 4 Biomechanical stimulation and functional maturation of printed cellularized patches.

    (A) Schematic illustration of a custom-made bioreactor to apply dual MS for the maturation of engineered cardiac tissue. PMMA, polymethylmethacrylate. (B) Both the out-of-plane loading and fluid shear stress applied to the patches. (C) Immunostaining of cTnI (red) and vWf (green) on the wave-patterned patch under MS condition (+MS) versus nonstimulated control (−MS). Scale bars, 50 μm. (D) Immunostaining of the α-actinin (green) and Cx43 (red) on the wave-patterned patch under MS condition (+MS) versus nonstimulated control (−MS). Scale bars, 20 μm. (E) Cross-sectional immunostaining of the sarcomeric structure (Desmin; green) and vascular CD31 (red) on the patches under MS condition (+MS). Scale bars, 50 μm. (F) The beating rate of hiPSC-CMs on the printed patches under MS condition (+MS) versus nonstimulated control (−MS) on day 14 (means ± SD, n ≥ 6, *P < 0.05). BPM, beats per minute. Relative gene expression of (G) myocardial structure [myosin light chain 2 (MYL2)], (H) excitation-contraction coupling [ryanodine receptor 2 (RYR2)], and (I) angiogenesis (CD31) on the patches under MS condition (+MS) versus nonstimulated control (−MS) on day 1, day 7, and day 14 (means ± SD, n ≥ 9, *P < 0.05, **P < 0.01, and ***P < 0.001).

  • Fig. 5 In vivo implantation and long-term evaluation of 4D patches.

    (A) Optical image of surgical implantation of the patch. (B) Optical image of a heart I/R MI model after 4 months. (C) Optical image of the implanted cellularized patch at week 3, exhibiting a firm adhesion (inset). (D) H&E image of the cellularized patch at week 3, demonstrating the cell clusters with a high density (yellow arrowhead). Scale bar, 400 μm. (E) Fluorescent image of (GFP+) iPSC-CMs on the patch at week 3, showing a high engraftment rate (yellow arrowhead). Scale bar, 100 μm. (F) Immunostaining of cTnI (red) and vWf (green) on the cellularized patch at week 3. Scale bar, 100 μm. (G) H&E images of mouse MI hearts without treatment (MI) and with cellularized patch (MI + patch) at week 10. Infarct area after MI (yellow circles). Scale bars, 800 μm. (H) Cardiac magnetic resonance imaging (cMRI) images of a mouse heart with patch at week 10. Left (spin echo): the position of the heart and implanted patch. Right (cine): the blood (white color) perfusion from the heart to the patch. Photo credit: Haitao Cui, GWU.

  • Fig. 6 In vivo implantation and long-term evaluation of 4D patches (continuous).

    (A) Immunostaining of cTnI (red) and vWf (green) on the cellularized patch at week 10. Scale bar, 800 μm. Border of the heart (white dashed line) and capillary lumen (white arrow). Scale bar (enlarged), 100 μm. (B) Immunostaining of human-specific CD31 (red) on the cellularized patch at week 10, showing the generated capillaries by hECs. Scale bar, 800 μm. Border of the heart (white dashed line). Scale bar (enlarged), 100 μm. (C) Immunostaining of cTnI (red) and vWf (green) on the cellularized patch for 4 months, showing the increased density of the vessels. Scale bar, 800 μm. Border of the heart (white dashed line) and capillary lumen (white arrow). Scale bar (enlarged), 100 μm. (D) Quantification of capillaries with vWf staining data for 10 weeks and 4 months (means ± SD, n ≥ 6, *P < 0.05 and **P < 0.01). (E) Immunostaining of human-specific CD31 (red) on the cellularized patch for 4 months. Scale bar, 800 μm. Border of the heart (white dashed line). Scale bar (enlarged), 100 μm. (F) Immunostaining of α-actinin (green) and human-specific CD31 (red) on the cellularized patch at month 4. Scale bars, 50 μm.

Supplementary Materials

  • Supplementary Materials

    4D physiologically adaptable cardiac patch: A 4-month in vivo study for the treatment of myocardial infarction

    Haitao Cui, Chengyu Liu, Timothy Esworthy, Yimin Huang, Zu-xi Yu, Xuan Zhou, Hong San, Se-jun Lee, Sung Yun Hann, Manfred Boehm, Muhammad Mohiuddin, John P. Fisher, Lijie Grace Zhang

    Download Supplement

    The PDF file includes:

    • Figs. S1 to S7
    • Table S1
    • Legends for movies S1 to S5

    Other Supplementary Material for this manuscript includes the following:

    Files in this Data Supplement:

Stay Connected to Science Advances

Navigate This Article