Abstract
Low-temperature biopreservation and 3D tissue engineering present two differing routes towards eventual on-demand access to transplantable biologics, but recent advances in both fields present critical new opportunities for crossover between them. In this work, we demonstrate sub-zero centigrade preservation and revival of autonomously beating three-dimensional human induced pluripotent stem cell (hiPSC)-derived cardiac microtissues via isochoric supercooling, without the use of chemical cryoprotectants. We show that these tissues can cease autonomous beating during preservation and resume it after warming, that the supercooling process does not affect sarcomere structural integrity, and that the tissues maintain responsiveness to drug exposure following revival. Our work suggests both that functional three dimensional (3D) engineered tissues may provide an excellent high-content, low-risk testbed to study complex tissue biopreservation in a genetically human context, and that isochoric supercooling may provide a robust method for preserving and reviving engineered tissues themselves.
Highlights
Low-temperature biopreservation and 3D tissue engineering present two differing routes towards eventual on-demand access to transplantable biologics, but recent advances in both fields present critical new opportunities for crossover between them
Our data demonstrate that functional 3D microtissues may be effectively preserved at subzero temperatures, which has several important implications
Recent work has demonstrated that the cardiac microphysiological system (MPS) can successfully predict clinical trial arrhythmia outcomes[11], indicating potential prognostic value in predicting human-scale cardiac behavior
Summary
Low-temperature biopreservation and 3D tissue engineering present two differing routes towards eventual on-demand access to transplantable biologics, but recent advances in both fields present critical new opportunities for crossover between them. We demonstrate sub-zero centigrade preservation and revival of autonomously beating threedimensional human induced pluripotent stem cell (hiPSC)-derived cardiac microtissues via isochoric supercooling, without the use of chemical cryoprotectants We show that these tissues can cease autonomous beating during preservation and resume it after warming, that the supercooling process does not affect sarcomere structural integrity, and that the tissues maintain responsiveness to drug exposure following revival. While historical research on the preservation of organs and complex tissues focused largely on the deep cryogenic temperatures that would theoretically enable indefinite storage, the last decade has seen the advent of “high-subzero” preservation efforts, targeting the −20°–0 °C range[5] These efforts aim for multiday preservation, in recognition of the fact that even 24- to 48-h preservation of many complex donor biologics could prove transformative for transplant medicine[1,2,3,4]. We employ a recently developed technique, isochoric supercooling[6], to achieve high-stability and predictable supercooling of the University of Wisconsin (UW) organ preservation solution at −3 °C, in which we successfully preserve a functional 3D hiPSC-derived cardiac microphysiological system (MPS)[11] for multiple days without the addition of nonphysiological cryoprotectants such as dimethyl sulfoxide (DMSO) or glycerol
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