Résumé
Mitochondria are crucial for maintaining the properties of embryonic stem cells (ESCs) and for regulating their subsequent differentiation into diverse cell lineages, including cardiomyocytes. However, mitochondrial regulators that manage the rate of differentiation or cell fate have been rarely identified. This study aimed to determine the potential mitochondrial factor that controls the differentiation of ESCs into cardiac myocytes. We induced cardiomyocyte differentiation from mouse ESCs (mESCs) and performed microarray assays to assess messenger RNA (mRNA) expression changes at differentiation day 8 (D8) compared with undifferentiated mESCs (D0). Among the differentially expressed genes, Pdp1 expression was significantly decreased (27-fold) on D8 compared to D0, which was accompanied by suppressed mitochondrial indices, including ATP levels, membrane potential, ROS and mitochondrial Ca²⁺. Notably, Pdp1 overexpression significantly enhanced the mitochondrial indices and pyruvate dehydrogenase activity and reduced the expression of cardiac differentiation marker mRNA and the cardiac differentiation rate compared to a mock control. In confirmation of this, a knockdown of the Pdp1 gene promoted the expression of cardiac differentiation marker mRNA and the cardiac differentiation rate. In conclusion, our results suggest that mitochondrial PDP1 is a potential regulator that controls cardiac differentiation at an early differentiation stage in ESCs.
Sujets)
Animaux , Souris , Adénosine triphosphate , Lignage cellulaire , Cellules souches embryonnaires , Potentiels de membrane , Mitochondries , Cellules souches embryonnaires de souris , Myocytes cardiaques , Oxidoreductases , Pyruvate dehydrogenase (lipoamide)-phosphatase , Acide pyruvique , ARN messagerRésumé
The pyruvate dehydrogenase complex (PDC) is an emerging target for the treatment of metabolic syndrome. To maintain a steady-state concentration of adenosine triphosphate during the feed-fast cycle, cells require efficient utilization of fatty acid and glucose, which is controlled by the PDC. The PDC converts pyruvate, coenzyme A (CoA), and oxidized nicotinamide adenine dinucleotide (NAD+) into acetyl-CoA, reduced form of nicotinamide adenine dinucleotide (NADH), and carbon dioxide. The activity of the PDC is up- and down-regulated by pyruvate dehydrogenase kinase and pyruvate dehydrogenase phosphatase, respectively. In addition, pyruvate is a key intermediate of glucose oxidation and an important precursor for the synthesis of glucose, glycerol, fatty acids, and nonessential amino acids.
Sujets)
Acétyl coenzyme A , Adénosine triphosphate , Acides aminés , Dioxyde de carbone , Coenzyme A , Diabète , Acides gras , Glucose , Glycérol , NAD , Obésité , Oxidoreductases , Phosphotransferases , Pyruvate dehydrogenase (lipoamide)-phosphatase , Complexe du pyruvate déshydrogénase , Acide pyruviqueRésumé
The effect of methotrexate (MTX) on the activity of pyruvate dehydrogenase from liver mitochondria and from the crude extract of HeLa cells was studied. the activity of the enzyme from liver mitochondria was 50% inhibited with 800nmoles MTX/mg of protein and the enzyme from vrude extract of HeLa cells was 50% inhibited with only 60 nmoles MTX/mg of protein. The specific activity of the former was 0.75nmoles NAD+ reduced.min-1. meg-1 and the latter 0.48nmoles NAD+ reduced.minmg-1