Efficient Generation of Human Embryonic Stem Cell-Derived Cardiac Progenitors Based on Tissue-Specific Enhanced Green Fluorescence Protein Expression

Szebényi, Kornélia; Péntek, Adrienn; Erdei, Zsuzsa; Várady, György; Orbán, Tamás I.; Sarkadi, Balázs; Apáti, Ágota

doi:10.1089/ten.tec.2013.0646

Cited by 3 publications

(2 citation statements)

References 38 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Utilizing the constitutive nature of this promoter, a combined genotype/phenotype screening approach was applied to establish a homozygous rat strain in a relatively short period of time. In accordance with our earlier findings with embryonic stem cells 30 , the highest GCaMP2 expression was found in the cardiac tissues, especially in the atria, while appreciable transgene expression was found in the kidney and in the liver. Anatomical analyses found no differences in the heart size and weight between WT and TG animals, while slight left ventricular wall thickening was observed at the apex of the heart in transgenic rats.…”

Section: Discussionsupporting

confidence: 93%

Generation of a Homozygous Transgenic Rat Strain Stably Expressing a Calcium Sensor Protein for Direct Examination of Calcium Signaling

Szebényi

Füredi

Kolacsek

et al. 2015

Sci Rep

Self Cite

View full text Add to dashboard Cite

In drug discovery, prediction of selectivity and toxicity require the evaluation of cellular calcium homeostasis. The rat is a preferred laboratory animal for pharmacology and toxicology studies, while currently no calcium indicator protein expressing rat model is available. We established a transgenic rat strain stably expressing the GCaMP2 fluorescent calcium sensor by a transposon-based methodology. Zygotes were co-injected with mRNA of transposase and a CAG-GCaMP2 expressing construct, and animals with one transgene copy were pre-selected by measuring fluorescence in blood cells. A homozygous rat strain was generated with high sensor protein expression in the heart, kidney, liver, and blood cells. No pathological alterations were found in these animals, and fluorescence measurements in cardiac tissue slices and primary cultures demonstrated the applicability of this system for studying calcium signaling. We show here that the GCaMP2 expressing rat cardiomyocytes allow the prediction of cardiotoxic drug side-effects, and provide evidence for the role of Na+/Ca2+ exchanger and its beneficial pharmacological modulation in cardiac reperfusion. Our data indicate that drug-induced alterations and pathological processes can be followed by using this rat model, suggesting that transgenic rats expressing a calcium-sensitive protein provide a valuable system for pharmacological and toxicological studies.

show abstract

Section: Discussionsupporting

confidence: 93%

Generation of a Homozygous Transgenic Rat Strain Stably Expressing a Calcium Sensor Protein for Direct Examination of Calcium Signaling

Szebényi

Füredi

Kolacsek

et al. 2015

Sci Rep

Self Cite

View full text Add to dashboard Cite

show abstract

“…The CAG promoter driven eGFP expressing human HUES9 stem cell culture (Ethic license: Hungarian Committee of Human Reproduction; 31681-1/2004-1016EHR12534-0/2009-1016EHR; ES2HEART consortium) [ 18 , 19 ] was dispersed by 0.5 mg/mL collagenase type IV (Gibco, Invitrogen; Carlsbad, CA, USA) dissolved in KnockOut ™ Dulbecco's Modified Eagle Medium (Gibco). Subsequently, cells were maintained in cell suspension culture for 6 days in KnockOut Dulbecco's Modified Eagle Medium (Gibco), supplemented with 20% embryonic stem cell-qualified fetal bovine serum (Gibco), 1% nonessential amino acids, 1% L-glutamine (Gibco), and 0.2% beta-mercaptoethanol (Gibco).…”

Section: Methodsmentioning

confidence: 99%

Exogenous Nitric Oxide Protects Human Embryonic Stem Cell‐Derived Cardiomyocytes against Ischemia/Reperfusion Injury

Pálóczi

Varga

Apáti

et al. 2016

Oxidative Medicine and Cellular Longevity

Self Cite

View full text Add to dashboard Cite

Background and Aims. Human embryonic stem cell- (hESC-) derived cardiomyocytes are one of the useful screening platforms of potential cardiocytoprotective molecules. However, little is known about the behavior of these cardiomyocytes in simulated ischemia/reperfusion conditions. In this study, we have tested the cytoprotective effect of an NO donor and the brain type natriuretic peptide (BNP) in a screening platform based first on differentiated embryonic bodies (EBs, 6 + 4 days) and then on more differentiated cardiomyocytes (6 + 24 days), both derived from hESCs. Methods. Both types of hESC-derived cells were exposed to 150 min simulated ischemia, followed by 120 min reperfusion. Cell viability was assessed by propidium iodide staining. The following treatments were applied during simulated ischemia in differentiated EBs: the NO-donor S-nitroso-N-acetylpenicillamine (SNAP) (10−7, 10−6, and 10−5 M), BNP (10−9, 10−8, and 10−7 M), and the nonspecific NO synthase inhibitor Nω-nitro-L-arginine (L-NNA, 10−5 M). Results. SNAP (10−6, 10−5 M) significantly attenuated cell death in differentiated EBs. However, simulated ischemia/reperfusion-induced cell death was not affected by BNP or by L-NNA. In separate experiments, SNAP (10−6 M) also protected hESC-derived cardiomyocytes. Conclusions. We conclude that SNAP, but not BNP, protects differentiated EBs or cardiomyocytes derived from hESCs against simulated ischemia/reperfusion injury. The present screening platform is a useful tool for discovery of cardiocytoprotective molecules and their cellular mechanisms.

show abstract

Myocardial commitment from human pluripotent stem cells: Rapid production of human heart grafts

et al. 2016

View full text Add to dashboard Cite

Genome editing on human pluripotent stem cells (hPSCs) together with the development of protocols for organ decellularization opens the door to the generation of autologous bioartificial hearts. Here we sought to generate for the first time a fluorescent reporter human embryonic stem cell (hESC) line by means of Transcription activator-like effector nucleases (TALENs) to efficiently produce cardiomyocyte-like cells (CLCs) from hPSCs and repopulate decellularized human heart ventricles for heart engineering. In our hands, targeting myosin heavy chain locus (MYH6) with mCherry fluorescent reporter by TALEN technology in hESCs did not alter major pluripotent-related features, and allowed for the definition of a robust protocol for CLCs production also from human induced pluripotent stem cells (hiPSCs) in 14 days. hPSCs-derived CLCs (hPSCs-CLCs) were next used to recellularize acellular cardiac scaffolds. Electrophysiological responses encountered when hPSCs-CLCs were cultured on ventricular decellularized extracellular matrix (vdECM) correlated with significant increases in the levels of expression of different ion channels determinant for calcium homeostasis and heart contractile function. Overall, the approach described here allows for the rapid generation of human cardiac grafts from hPSCs, in a total of 24 days, providing a suitable platform for cardiac engineering and disease modeling in the human setting.

show abstract

Efficient Generation of Human Embryonic Stem Cell-Derived Cardiac Progenitors Based on Tissue-Specific Enhanced Green Fluorescence Protein Expression

Cited by 3 publications

References 38 publications

Generation of a Homozygous Transgenic Rat Strain Stably Expressing a Calcium Sensor Protein for Direct Examination of Calcium Signaling

Generation of a Homozygous Transgenic Rat Strain Stably Expressing a Calcium Sensor Protein for Direct Examination of Calcium Signaling

Exogenous Nitric Oxide Protects Human Embryonic Stem Cell‐Derived Cardiomyocytes against Ischemia/Reperfusion Injury

Myocardial commitment from human pluripotent stem cells: Rapid production of human heart grafts

Contact Info

Product

Resources

About