title: Functional and developmental properties of human embryonic stem cells-derived cardiomyocytes.
abstract: Cardiovascular diseases are the most frequent cause of death in the industrialized world, with the main contributor being myocardial infarction. Given the high morbidity and mortality rates associated with congestive heart failure, the shortage of donor hearts for transplantation, complications resulting from immunosuppression, and long-term failure of transplanted organs, regeneration of the diseased myocardium by cell transplantation is an attractive therapeutic modality. Because it is desired that the transplanted cells fully integrate within the diseased myocardium, contribute to its contractile performance, and respond appropriately to various physiological stimuli (eg, beta-adrenergic stimulation), our major long-term goal is to investigate the developmental changes in functional properties and hormonal responsiveness of human embryonic stem cells-derived cardiomyocytes (hESC-CM). Furthermore, because one of the key obstacles in advancing cardiac cell therapy is the low differentiation rate of hESC into cardiomyocytes, which reduces the clinical efficacy of cell transplantation, our second major goal is to develop efficient protocols for directing the cardiomyogenic differentiation of hESC in vitro. To accomplish the first goal, we investigated the functional properties of hESC-CM (<90 days old), respecting the contractile function and the underlying intracellular Ca(2+) handling. In addition, we performed Western blot analysis of the key Ca(2+)-handling proteins SERCA2, calsequestrin, phospholamban and the Na(+)/Ca(2+) exchanger. Our major findings were the following: (1) In contrast to the mature myocardium, hESC-CM exhibit negative force-frequency relationships and do not present postrest potentiation. (2) Ryanodine and thapsigargin do not affect the [Ca(2+)](i) transient and contraction, suggesting that, at this developmental stage, the contraction does not depend on sarcoplasmic reticulum Ca(2+) release. (3) In agreement with the finding that a voltage-dependent Ca(2+) current is present in hESC-CM and contributes to the mechanical function, verapamil completely blocks contraction. (4) Although hESC-CM express SERCA2 and Na(+)/Ca(2+) exchanger at levels comparable to those of the adult human myocardium, calsequestrin and phospholamban are not expressed. (4) In agreement with other reports, hESC-CM are responsive to beta-adrenergic stimulation. These findings show that the mechanical function related to intracellular Ca(2+) handling of hESC-CM differs from the adult myocardium, probably because of immature sarcoplasmic reticulum capacity.
authors.affiliation: Rappaport Family Institute for Research in the Medical Sciences, Ruth & Bruce Rappaport Faculty of Medicine, Technion, Haifa, Israel. binah@tx.technion.ac.il
authors.forename: Ofer
authors.initials: O
authors.lastname: Binah
authors.forename: Katya
authors.initials: K
authors.lastname: Dolnikov
authors.forename: Oshra
authors.initials: O
authors.lastname: Sadan
authors.forename: Mark
authors.initials: M
authors.lastname: Shilkrut
authors.forename: Naama
authors.initials: N
authors.lastname: Zeevi-Levin
authors.forename: Michal
authors.initials: M
authors.lastname: Amit
authors.forename: Asaf
authors.initials: A
authors.lastname: Danon
authors.forename: Joseph
authors.initials: J
authors.lastname: Itskovitz-Eldor
mesh_terms: D000200:Action Potentials / Q000502:physiology*; D002118:Calcium / Q000378:metabolism; D002478:Cells, Cultured; D053595:Embryonic Stem Cells / Q000166:cytology* / Q000502:physiology*; D006801:Humans; D009200:Myocardial Contraction / Q000502:physiology*; D032383:Myocytes, Cardiac / Q000166:cytology* / Q000502:physiology*; D023822:Tissue Engineering / Q000379:methods*
chemical_list: D002118:Calcium