Temiz Artmann, Aysegül;
Kurulgan Demirci, Eylem;
Fırat, Ipek Seda;
Oflaz, Hakan;
Artmann, Gerhard M.
Recombinant Activated Protein C (rhAPC) Affects Lipopolysaccharide-Induced Mechanical Compliance Changes and Beat Frequency of mESC-Derived Cardiomyocyte Monolayers
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Medientyp:
E-Artikel
Titel:
Recombinant Activated Protein C (rhAPC) Affects Lipopolysaccharide-Induced Mechanical Compliance Changes and Beat Frequency of mESC-Derived Cardiomyocyte Monolayers
Beteiligte:
Temiz Artmann, Aysegül;
Kurulgan Demirci, Eylem;
Fırat, Ipek Seda;
Oflaz, Hakan;
Artmann, Gerhard M.
Beschreibung:
<jats:title>ABSTRACT</jats:title>
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<jats:title>Background:</jats:title>
<jats:p>Septic cardiomyopathy increases mortality by 70% to 90% and results in mechanical dysfunction of cells.</jats:p>
</jats:sec>
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<jats:title>Methods:</jats:title>
<jats:p>Here, we created a LPS-induced <jats:italic toggle="yes">in-vitro</jats:italic> sepsis model with mouse embryonic stem cell-derived cardiomyocytes (mESC-CM) using the CellDrum technology which simultaneously measures mechanical compliance and beat frequency of mESCs. Visualization of reactive oxygen species (ROS), actin stress fibers, and mRNA quantification of endothelial protein C receptor (EPCR) and protease-activated receptor 1 (PAR1) before/after LPS incubation were used for method validation. Since activated protein C (APC) has cardioprotective effects, samples were treated with human recombinant APC (rhAPC) with/-out LPS predamage to demonstrate the application in therapeutic studies.</jats:p>
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<jats:title>Results:</jats:title>
<jats:p>Twelve hours LPS treatment (5 μg/mL) increased ROS and decreased actin stress fiber density and significantly downregulated EPCR and PAR1 compared to control samples (0.26, 0.39-fold respectively). rhAPC application (5 μg/mL, 12 h) decreased ROS and recovered actin density, EPCR, and PAR1 levels were significantly upregulated compared to LPS predamaged samples (4.79, 3.49-fold respectively). The beat frequencies were significantly decreased after 6- (86%) and 12 h (73%) of LPS application. Mechanical compliance of monolayers significantly increased in a time-dependent manner, up to eight times upon 12-h LPS incubation compared to controls. rhAPC incubation increased the beat frequency by 127% (6h-LPS) and 123% (12h-LPS) and decreased mechanical compliance by 68% (12h-LPS) compared to LPS predamaged samples.</jats:p>
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<jats:title>Conclusion:</jats:title>
<jats:p>LPS-induced contraction dysfunction and the reversal effects of rhAPC were successfully assessed by the mechanical properties of mESC-CMs. The CellDrum technology proved a decent tool to simulate sepsis <jats:italic toggle="yes">in-vitro</jats:italic>.</jats:p>
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