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Acta Physiologica 2012; Volume 204, Supplement 689
91st Annual Meeting of The German Physiological Society
3/22/2012-3/25/2012
Dresden, Germany
HIGH-THROUGHPUT SCREENING OF INDUCED PLURIPOTENT STEM CELL-DERIVED CARDIOMYOCYTES WITH LONG QT-SYNDROME 3
Abstract number: P189
Friedrichs1 *S., Malan1 D., Matuscheck1 Y., Fleischmann1 B.K., Sasse1 P.
1University Bonn, Institute of Physiology I, Bonn, Germany
Long QT-syndromes (LQTS) are inheritable cardiac diseases and can be investigated in cardiomyocytes derived from disease-specific induced pluripotent stem (iPS) cells. LQTS3 in humans is mainly caused by the ࢞KPQ deletion of the cardiac sodium channel. Recently, we showed that LQTS3-specific iPS cell-derived cardiomyocytes carrying this mutation recapitulate the disease-specific phenotypes, namely prolonged action potentials (APs) and early after depolarizations (EADs) at low heart rates (Circ Res. 2011,109:8417).
These results were obtained using manual patch clamp. For automated patch-clamp analysis purified cardiomyocytes are required. Therefore we have genetically engineered wild-type and LQTS3-specific iPS cell lines using a lentivirus that expresses a puromycin-resistance and GFP under the control of the a-myosin heavy chain promoter. During differentiation of these iPS cells, GFP-positive beating clusters appeared around day 10. For purification puromycin was added, which lead to a strong enrichment of cardiomyocytes (wild-type iPS: 96.2±1.9% (n=3); LQTS3-specific iPS: 93.9±5.3% (n=2)). Conventional patch-clamp experiments revealed AP prolongation and EADs at low pacing rates only in LQTS3-specific but not in wild-type cardiomyocytes. The possibility of drug screening was investigated by application of the sodium channel-blocker mexitilene, which reduced the disease-specific AP prolongation. In order to establish automated drug screening from purified cardiomyocytes, we recorded APs with a planar patch-clamp robot (Patchliner, Nanion). Our first, preliminary data revealed the characteristic AP prolongation as well as EADs in LQTS3-specific but not in wild-type cardiomyocytes.
Hence, purified cardiomyocytes from disease-specific iPS cells can be generated by a lentiviral purification strategy and are suitable for automated drug screening.
To cite this abstract, please use the following information:
Acta Physiologica 2012; Volume 204, Supplement 689 :P189