Dataset Information

Contractile work contributes to maturation of energy metabolism in hiPSC-derived cardiomyocytes

ABSTRACT: Energy metabolism is a key aspect of cardiomyocyte biology. Human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CM) are a promising tool for biomedical application, but they are immature and have not undergone metabolic shift related to early postnatal development. Cultivation of hiPSC-CM in 3D engineered heart tissue (EHT) format leads to morphological maturation. This study compared the mitochondrial and metabolic state of hiPSC-CM in standard 2D culture and the EHT format and determined the influence of contractile activity. HiPSC-CM in EHTs showed ~2-fold higher number of mitochondria (electron microscopy), mitochondrial mass (mitotracker), DNA (Mt-ND1, Mt-ND2), and protein abundance (proteome) than in 2D culture. While hiPSC-CM exhibited the principal ability to use glucose, lactate and fatty acids as energy substrates irrespective of culture format, hiPSC-CM in 3D performed more oxidation of glucose, lactate and fatty acid, and less anaerobic glycolysis. The increase in mitochondrial mass and DNA in 3D was diminished by pharmacological inhibition of contractile force, suggesting that contractile work participates in mitochondrial development hiPSC-CM. In conclusion, contractile work in the EHT format contributes to metabolic maturation of hiPSC-CM.

INSTRUMENT(S): LTQ Orbitrap

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Heart

SUBMITTER: Andrea Stoehr

LAB HEAD: Arne Hansen

PROVIDER: PXD008205 | Pride | 2018-03-06

REPOSITORIES: Pride

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Contractile Work Contributes to Maturation of Energy Metabolism in hiPSC-Derived Cardiomyocytes.

Ulmer Bärbel M BM Stoehr Andrea A Schulze Mirja L ML Patel Sajni S Gucek Marjan M Mannhardt Ingra I Funcke Sandra S Murphy Elizabeth E Eschenhagen Thomas T Hansen Arne A

Stem cell reports 20180301 3

Energy metabolism is a key aspect of cardiomyocyte biology. Human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) are a promising tool for biomedical application, but they are immature and have not undergone metabolic maturation related to early postnatal development. To assess whether cultivation of hiPSC-CMs in 3D engineered heart tissue format leads to maturation of energy metabolism, we analyzed the mitochondrial and metabolic state of 3D hiPSC-CMs and compared it with 2D cu ...[more]

PMID: 29503093

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Project description:Background: The phospholamban (PLN) p.Arg14del mutation belongs to the established causes of dilated cardiomyopathy, with the molecular disease mechanisms incompletely understood. We used human induced pluripotent stem cell (hiPSC)-derived cardiomyocytes, CRISPR/Cas9 gene editing and patient heart samples to investigate the molecular pathomechanisms of PLN p.Arg14del cardiomyopathy. Methods and results: Patient dermal fibroblasts carrying the PLN p.Arg14del mutation were reprogrammed into hiPSC, isogenic controls were established by CRISPR/Cas9. Cells were differentiated into cardiomyocytes and characterized in 2D and 3D-engineered heart tissue (EHT) format. Mutant cardiomyocytes revealed significantly prolonged Ca2+ transient decay time, Ca2+-load dependent irregular beating pattern and lower peak force compared to isogenic controls. While this data support the reported super-inhibitory effect of PLN p.Arg14del on the sarcoplasmic/endoplasmic reticulum Ca2+ ATPase, an unchanged SR Ca2+ content argued against disturbed SR Ca2+ cycling as the sole cause of contractile dysfunction. Label-free proteomic analysis of p.Arg14del EHTs revealed less endoplasmic reticulum (ER), ribosomal and mitochondrial proteins. Electron microscopy showed dilation of the rough ER, large lipid droplets in close association with mitochondria and reduced mitochondrial number. Follow-up experiments confirmed impairment of the rough ER/mitochondria compartment and enhanced oxidative stress in PLN p.Arg14del. Relevance for human disease was demonstrated by immunohistochemistry of human heart samples. PLN p.Arg14del end-stage heart failure samples revealed perinuclear aggregates positive for ER marker proteins and oxidative stress in comparison to ischemic heart failure - and non-failing donor heart samples. Surprisingly, transduction of PLN p.Arg14del EHTs with the Ca2+ binding protein GCaMP6f reversed the contractile, molecular and morphological disease phenotype. Conclusion: This study identified impairment of the rough ER/mitochondria compartment without SR dysfunction as a novel disease mechanism underlying PLN p.Arg14del cardiomyopathy. The pathology was improved by Ca2+-scavenging, suggesting impaired local Ca2+ cycling as an important disease culprit.

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