Liensinine- and Neferine-Induced Cardiotoxicity in Primary Neonatal Rat Cardiomyocytes and Human-Induced Pluripotent Stem Cell-Derived Cardiomyocytes

Background: It has been shown that transplantation of engineered cardiac tissue (ECT) derived from human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) into the infarct heart induces electrical communication between the ECT and the native myocardium; however, factors enhancing the electrical integrity and thus the therapeutic effects are not fully understood. We herein hypothesized that content of cardiomyocytes in the ECT may be a key to achieve efficient electrical coupling and functional contribution in chronic rat myocardial infarction (MI) heart. Methods and Results: Neonatal rat cardiomyocytes (NRCM), mimicking the host myocardium, were partially covered by the ECT containing iPSC-CMs produced by thermoresponsive culture dishes in vitro , to explore electrical communication of the ECT with myocardium. As a result, the NRCM and the ECT showed spontaneous, individual contractions for 2 hours, though they gradually showed electrical and motional synchronization, featuring transmitted electrical pulse from the NRCM to the ECT, as assessed by multi-electrode array. Subsequently, the ECT of different ratios (25, 50, 70, and 90%) of iPSC-CMs were generated by magnetic-activated cell sorting using cardiac specific cell surface marker. As a result, the 70% group exhibited the highest contractile and relaxation properties in vitro , as assessed by high-speed video microscopy image-based motion analysis and Ca transient measurement. Finally, the ECTs including 25, 50, 70% CMs were transplanted to immune deficient rat MI model (n=7 each). As a result, ejection fraction was significantly improved in the 50% (52±10%) and 70% (52±12%) groups, but not in the 25% group (35±5%), as compared to the control (35±10%; P <0.05). Epicardial optical mapping of Langendorff perfused heart on day 3 showed that the ECTs of 50% and the 70% groups exhibited electrical activity and synchronization with the native myocardium. Conclusion: Transplantation of the ECT improved cardiac performance associated with synchronization with the myocardium in rat infarction model, dependent upon content of the cardiomyocytes in the ECT. It was thus suggested that transplanted ECT may behave “working cardiac construct” in the damaged heart.

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Abstract 342: Serum Albumin Hydrogels Alter Excitation-Contraction Coupling in Neonatal Rat Myocytes and Human Induced Pluripotent Stem Cell Derived Cardiomyocytes

Circulation Research ◽

10.1161/res.121.suppl_1.342 ◽

2017 ◽

Vol 121 (suppl_1) ◽

Author(s):

Eleanor J Humphrey ◽

Manuel M Mazo ◽

Nadav Amdursky ◽

Nicholas S Peters ◽

Molly M Stevens ◽

...

Keyword(s):

Gene Expression ◽

Stem Cell ◽

Serum Albumin ◽

Pluripotent Stem Cell ◽

Induced Pluripotent Stem Cell ◽

Neonatal Rat ◽

Calcium Handling ◽

Time To Peak ◽

Induced Pluripotent ◽

Reduced Time

Tissue engineering provides a promising method of introducing functional cardiomyocytes (CMs) to damaged myocardium after myocardial infarction; however, finding a biocompatible construct with the chemical and mechanical properties capable of supporting CM function is challenging. Serum albumin hydrogels are novel autogenic scaffolds with elastic properties that can be tailored to mimic the stiffness of native adult myocardium. We assessed the hypothesis that culturing immature CMs on these serum albumin hydrogels would affect CM gene expression and calcium handling. Neonatal cardiomyocyte (NRVM) viability was maintained for at least 14 days on the hydrogels, with clear sarcomeric striations. Cardiac gene expression was quantified using RT-qPCR and demonstrated an up regulation in many genes of cells cultured on hydrogels compared to glass (e.g. relative expression (log 2-ΔΔCt) of ryanodine receptor 2: glass= -2.3±0.5, hydrogel= -0.3±0.1,p<0.01; connexin 43:glass= -1.7±0.5, hydrogel= 0.3±0.1,p<0.01,n=4-6). Compared to glass, NRVMs on hydrogels have an increased time to peak of the calcium transients measured using Fluo-4AM and field stimulated at 1 Hz (tp glass=38±3 ms, tp hydrogel= 54±2 ms, p<0.01,n=4-6). Compared to glass the hydrogels also have a reduced time 50% decay (t50 glass=108±13 ms, t50 hydrogel=78±6 ms, p<0.05,n=4-6) and 80% decay (t80 glass=217±19 ms, t80 hydrogel= 152±10 ms,p<0.05,n=4-6). Human induced pluripotent stem cell derived cardiomyocytes (iPSC-CMs) were cultured on the hydrogels for up to 28 days. Calcium handling was faster in the iPSC-CMs cultured on the hydrogels in comparison to glass with a reduced time to peak (tp glass=281±43 ms, tp hydrogel= 186±8 ms, p<0.05, n=4) and time to 50% decay (t50 glass=269±15 ms, t50 hydrogel=204±10 ms,p<0.01,n=4) and 90% decay (t90 glass=535±33 ms, t90 hydrogel=397±19 ms, p<0.01,n=4). The serum albumin hydrogels are compatible with NRVM and iPSC-CM culture for at least 28 days. We demonstrate that the serum albumin hydrogels have significant effects on CM calcium cycling and have the potential for use in myocardial repair. Further study is required to determine the mechanisms involved in calcium handling alterations and then assess this engineered patch in vivo for cardiac repair.

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