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EMBRYONIC STEM CELLS/INDUCED PLURIPOTENT STEM CELLS

1,4,5-Inositol Trisphosphate-Operated Intracellular Ca²⁺ Stores and Angiotensin-II/Endothelin-1 Signaling Pathway Are Functional in Human Embryonic Stem Cell-Derived Cardiomyocytes

^aRuth and Bruce Rappaport Faculty of Medicine, Technion-Israel Institute of Technology, Haifa, Israel;
^bRambam Medical Center, Haifa, Israel

Key Words. Human embryonic stem cell-derived cardiomyocytes • 1,4,5-Inositol trisphosphate • Angiotensin-II • Endothelin-1 • Intracellular Ca²⁺ transient • Contraction

Correspondence: Correspondence: Ofer Binah, Ph.D., Rappaport Institute, P.O. Box 9697, Haifa 31096, Israel. Telephone: 972-4-8295262; Fax: 972-4-8513919; e-mail: binah{at}tx.technion.ac.il; or Joseph Itskovitz-Eldor, M.D., Ph.D., Rambam Medical Center, Haifa 31096, Israel. Telephone: 972-4-8542536; e-mail: itskovitz{at}rambam.health.gov.il

Received on August 12, 2008; accepted for publication on September 11, 2008.

First published online in STEM CELLS EXPRESS  September 25, 2008.

On the basis of previous findings suggesting that in human embryonic stem cell-derived cardiomyocytes (hESC-CM) the sarcoplasmic reticulum Ca²⁺-induced release of calcium machinery is either absent or immature, in the present study we tested the hypothesis that hESC-CM contain fully functional 1,4,5-inositol trisphosphate (1,4,5-IP₃)-operated intracellular Ca²⁺ ([Ca²⁺]_i) stores that can be mobilized upon appropriate physiological stimuli. To test this hypothesis we investigated the effects of angiotensin-II (AT-II) and endothelin-1 (ET-1), which activate the 1,4,5-IP₃ pathway, on [Ca²⁺]_i transients and contractions in beating clusters of hESC-CM. Our major findings were that in paced hESC-CM both AT-II and ET-1 (10^–9 to 10^–7 M) increased the contraction amplitude and the maximal rates of contraction and relaxation. In addition, AT-II (10^–9 to 10^–7 M) increased the [Ca²⁺]_i transient amplitude. The involvement of 1,4,5-IP₃-dependent intracellular Ca²⁺ release in the inotropic effect of AT-II was supported by the findings that (a) hESC-CM express AT-II, ET-1, and 1,4,5-IP₃ receptors determined by immunofluorescence staining, and (b) the effects of AT-II were blocked by 2 µM 2-aminoethoxyphenyl borate (a 1,4,5-IP₃ receptor blocker) and U73122 [GenBank] (a phospholipase C blocker). In conclusion, these findings demonstrate for the first time that hESC-CM exhibit functional AT-II and ET-1 signaling pathways, as well as 1,4,5-IP₃-operated releasable Ca²⁺ stores.

Disclosure of potential conflicts of interest is found at the end of this article.