Center for Biomedical Engineering, School of Engineering, Brown University, Providence, RI, USA.
Stem Cells Transl Med. 2022 Mar 3;11(1):97-106. doi: 10.1093/stcltm/szab002.
Human-induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) have profound utility in generating functional human engineered cardiac tissues (ECT) for heart repair. However, the field at large is concerned about the relative immaturity of these hiPSC-CMs as we aim to develop clinically relevant models for regenerative therapy and drug testing. Herein, we develop a novel calcium (Ca2+) conditioning protocol that maintains ECTs in a physiological range of Ca2+ and assesses contractility in increasing calcium environments. Lactate-based selection served as a method to purify and shift the metabolic profile of hiPSC-CMs to evaluate the role of metabolism on Ca2+ sensitivity. After 2 weeks, we observe 2-fold greater peak twitch stress in high-Ca2+ conditioned ECTs, despite having lower stiffness and no change in Ca2+ sensitivity of twitch force. Interestingly, the force-calcium relationship reveals higher Ca2+ sensitivity in lactate conditioned tissues, suggesting that metabolic maturation alters mitochondrial Ca2+ buffering and regulation. Ca2+ sensitivity and force amplitude are not coupled, as lactate conditioned tissues produce force comparable to that of controls in high calcium environments. An upregulation of calcium handling protein gene expression likely contributes to the greater Ca2+ sensitivity in lactate conditioned hiPSC-CMs. Our findings support the use of physiological Ca2+ to enhance the functional maturation of excitation-contraction coupling in hiPSC-CMs and demonstrate that metabolic changes induced by lactate conditioning alter cardiomyocyte sensitivity to external Ca2+. These conditioning methods may be used to advance the development of engineered human cardiac tissue for translational applications in vitro and in vivo as a regenerative therapy.
人诱导多能干细胞衍生的心肌细胞（hiPSC-CMs）在生成用于心脏修复的功能性人类工程心脏组织（ECT）方面具有深远的应用价值。然而，由于我们旨在为再生治疗和药物测试开发临床相关模型，该领域普遍关注这些 hiPSC-CMs 的相对不成熟性。在此，我们开发了一种新的钙（Ca2+）调理方案，可使 ECT 保持在生理范围内的 Ca2+，并在增加 Ca2+环境中评估收缩性。基于乳酸盐的选择用作纯化和改变 hiPSC-CMs 代谢谱的方法，以评估代谢对 Ca2+敏感性的作用。两周后，我们观察到高 Ca2+调理 ECT 的峰值收缩力增加了 2 倍，尽管其刚性降低且收缩力对 Ca2+敏感性没有变化。有趣的是，力-钙关系显示出乳酸盐调理组织中更高的 Ca2+敏感性，表明代谢成熟改变了线粒体 Ca2+缓冲和调节。钙敏感性和力振幅不耦合，因为在高钙环境中，乳酸盐调理组织产生的力与对照组织相当。钙处理蛋白基因表达的上调可能有助于乳酸盐调理的 hiPSC-CMs 中更大的 Ca2+敏感性。我们的研究结果支持使用生理 Ca2+来增强 hiPSC-CMs 兴奋-收缩偶联的功能成熟，并表明乳酸盐调理引起的代谢变化改变了心肌细胞对外部 Ca2+的敏感性。这些调理方法可用于推进用于体外和体内再生治疗的工程化人类心脏组织的开发。
