Clin Res Cardiol (2021). 10.1007/s00392-021-01933-9
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An integrated platform for molecular phenotypic analysis in DCM patient-specific iPSC-cardiomyocytes and CRISPR-Cas9-engineered controls as a diagnostic research tool in cardiovascular disease
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A. Ebert1, A. Malkovskiy2, Y. Dai1, N. Ignatyeva1, J. Rajadas2, G. Hasenfuß1, for the study groups: AG31, AG 8, AG12
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1Herzzentrum, Klinik für Kardiologie und Pneumologie, Universitätsmedizin Göttingen, Göttingen; 2School of Medicine, Stanford University, Stanford, US;
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Human induced pluripotent stem
cell-derived cardiomyocytes (iPSC-CMs) emerged as an alternative tool in precision
medicine. New strategies are needed for realizing the full potential of molecular
disease-phenotypic data derived from patient-specific iPSC-CMs and their
utilization to evolve additional treatments for cardiovascular disease.
We developed a new approach for
combined analysis of calcium (Ca2+) handling and beating force in contractile cardiomyocytes. We
employed human iPSC-CMs from dilated cardiomyopathy (DCM) patients carrying a inherited,
familial point mutation in the sarcomeric protein troponin T (TnT), as well as
isogenic TnT-KO iPSC-CMs generated via CRISPR/Cas9 gene editing. Non-invasive single-cell
atomic force microscopy (AFM) was utilized to assess beating forces and -rates,
which were compared to Ca2+ handling capacities in these cells. We report
impaired Ca2+ handling and reduced contractile force in DCM iPSC-CMs compared
to healthy WT controls. We found TnT-KO iPSC-CMs not to display any contractile
force or Ca2+ transients but to generate Ca2+ sparks. We applied our analysis
strategy to Ca2+ traces and AFM deflection recordings and established maximum
rising rate, decay time, and duration of contraction with a multi-step background
correction. We present here an integrated approach for adaptive computing of
signal peaks for different Ca2+ flux or force levels, as well as analysis of
Ca2+ sparks, in DCM patient-specific iPSC-CM models combined with CRISPR/Cas9-engineering.
Moreover, we report long-term measurements of contractile force dynamics on
human single-cell iPSC-CMs.
Our platform enables deeper and more accurate
profiling of disease-specific differences in cardiomyocyte contraction profiles
using patient-derived iPSC-CMs and could facilitate development of new therapeutic
approaches in cardiovascular disease.
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https://dgk.org/kongress_programme/ht2021/BS888.htm
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