CARDIOLOGY
Bioprinting 3D heart pumps A concept that is gaining traction BY MOLLY KUPFER, PHD, AND BRENDA OGLE, PHD
H
eart disease is the leading cause of death worldwide, due in large part to the low regenerative capacity of the heart. With recent advances in stem cell biology, cardiac tissue engineering with human cells has emerged as an avenue to replace lost muscle after a cardiac event and to produce human models in vitro that can be used for disease modeling and testing of drugs and medical devices. Early engineered heart tissues, pioneered in the late 1990s and early 2000s, consisted of geometrically simple structures (strips or rings) made by casting cardiomyocytes in a protein-based gel. While such tissues can recapitulate the contractility of cardiac muscle, their lack of geometric complexity limits their capacity to reflect clinically relevant characteristics of the heart. That is, while they can generate force, they possess no internal chambered structure with which to pump fluid.
“Printing” cardiac tissue 3D bioprinting, wherein structures are fabricated layer-by-layer utilizing a cellladen “bio-ink” as a substrate, has been proposed as a means to generate more geometrically complex tissues from the bottom up. The concept is gaining
traction, as the ability to print tissues composed entirely of native proteins, cells, and/or biocompatible synthetic components is possible and accessible to many laboratories. Further, robust protocols have been developed for differentiating human-induced pluripotent stem cells (hiPSCs) into a variety of cell types, making it relatively easy to obtain cardiomyocytes ex vivo. However, while researchers have demonstrated the capacity to 3D-print entire heart organ models using biological materials, no one has yet demonstrated electromechanical function of cardiomyocytes within such a tissue. The fact that macroscale contractile function has not yet been achieved in a 3D-printed, perfusable, chambered heart model reflects the challenges associated with handling mature cardiac muscle cells. More specifically, cardiomyocytes do not proliferate or migrate readily. For this reason, it is challenging to achieve the high cell density required for the formation of functional cell-cell junctions while maintaining the structural support needed for an enclosed chamber. Macroscale cardiac function relies on the electromechanical coupling of individual cardiomyocytes to form an organized, synchronously contracting tissue. Traditionally, researchers have taken the approach of differentiating hiPSCs into cardiomyocytes in a tissue culture dish, and then collecting the differentiated cardiomyocytes and 3D printing with them. However, when hiPSCs are differentiated into cardiomyocytes this way, they tend to couple to each other and form a beating monolayer. To collect the cells from such an environment for further downstream applications typically requires one to break up these connections. Hence, to incorporate these cells into an engineered tissue, it is necessary to place them in a context where they can reform these interrupted connections. This is feasible in smaller, millimeterscale tissues, but it becomes challenging in larger, centimeter-scale tissues where the physical distance between cardiomyocytes after printing is too large to overcome.
A new strategy Our alternative approach is to print stem cells, which are highly proliferative, and then induce differentiation of cardiomyocytes in situ following cell expansion. To enable this approach, we sought to develop a bio-ink formulation that: 1. Promotes hiPSC viability; 2. Enables hiPSC proliferation and subsequent differentiation into cardiomyocytes; and 3. Is amenable to printing complex structures. Building on our understanding of how native extracellular matrix proteins modulate cell behavior, we developed an optimized bioink formulation composed of native proteins found in the heart. Some of these proteins were chemically modified to enable photo-crosslinking of the printed construct in order to maintain its geometric shape and structural integrity. To generate the printing template, an MRI scan of a human heart was obtained and scaled to the size of a mouse heart such that the longest axis was
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SEPTEMBER 2020 MINNESOTA PHYSICIAN
