Universität zu Köln

Acharya, Aviseka, Nemade, Harshal ORCID: 0000-0002-2704-181X, Prasad, Krishna Rajendra, Khan, Khadija, Hescheler, Juergen, Blackburn, Nick, Hemmersbach, Ruth, Papadopoulos, Symeon and Sachinidis, Agapios (2022). Live-Cell Imaging of the Contractile Velocity and Transient Intracellular Ca2+ Fluctuations in Human Stem Cell-Derived Cardiomyocytes. Cells, 11 (8). BASEL: MDPI. ISSN 2073-4409

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Abstract

Live-cell imaging techniques are essential for acquiring vital physiological and pathophysiological knowledge to understand and treat heart disease. For live-cell imaging of transient alterations of [Ca2+](i) in human cardiomyocytes, we engineered human-induced pluripotent stem cells carrying a genetically-encoded Ca2+-indicator (GECI). To monitor sarcomere shortening and relaxation in cardiomyocytes in real-time, we generated a alpha-cardiac actinin (ACTN2)-copepod (cop) green fluorescent protein (GFP(+))-human-induced pluripotent stem cell line by using the CRISPR-Cas9 and a homology directed recombination approach. The engineered human-induced pluripotent stem cells were differentiated in transgenic GECI-enhanced GFP(+)-cardiomyocytes and ACTN2-copGFP(+)-cardiomyocytes, allowing real-time imaging of [Ca2+](i) transients and live recordings of the sarcomere shortening velocity of ACTN2-copGFP(+)-cardiomyocytes. We developed a video analysis software tool to quantify various parameters of sarcoplasmic Ca2+ fluctuations recorded during contraction of cardiomyocytes and to calculate the contraction velocity of cardiomyocytes in the presence and absence of different drugs affecting cardiac function. Our cellular and software tool not only proved the positive and negative inotropic and lusitropic effects of the tested cardioactive drugs but also quantified the expected effects precisely. Our platform will offer a human-relevant in vitro alternative for high-throughput drug screenings, as well as a model to explore the underlying mechanisms of cardiac diseases.

Item Type:	Journal Article
Creators:	Creators Email ORCID ORCID Put Code Acharya, Aviseka UNSPECIFIED UNSPECIFIED UNSPECIFIED Nemade, Harshal UNSPECIFIED orcid.org/0000-0002-2704-181X UNSPECIFIED Prasad, Krishna Rajendra UNSPECIFIED UNSPECIFIED UNSPECIFIED Khan, Khadija UNSPECIFIED UNSPECIFIED UNSPECIFIED Hescheler, Juergen UNSPECIFIED UNSPECIFIED UNSPECIFIED Blackburn, Nick UNSPECIFIED UNSPECIFIED UNSPECIFIED Hemmersbach, Ruth UNSPECIFIED UNSPECIFIED UNSPECIFIED Papadopoulos, Symeon UNSPECIFIED UNSPECIFIED UNSPECIFIED Sachinidis, Agapios UNSPECIFIED UNSPECIFIED UNSPECIFIED
URN:	urn:nbn:de:hbz:38-671427
DOI:	10.3390/cells11081280
Journal or Publication Title:	Cells
Volume:	11
Number:	8
Date:	2022
Publisher:	MDPI
Place of Publication:	BASEL
ISSN:	2073-4409
Language:	English
Faculty:	Unspecified
Divisions:	Unspecified
Subjects:	no entry
Uncontrolled Keywords:	Keywords Language CALCIUM INDICATORS; SOFTWARE TOOL; MECHANISMS; SARCOMERE; DOXORUBICIN; SIGNATURES; TOXICITY; DISEASE; PROBES; FORCE Multiple languages Cell Biology Multiple languages
URI:	http://kups.ub.uni-koeln.de/id/eprint/67142