Table 1.
Characteristics of study subjects.
Note: CAD = coronary artery disease, CABG = coronary artery bypass grafting, COPD = chronic obstructive pulmonary disease, CPR = cardiopulmonary resuscitation, MI = myocardial infarction, T1DM/T2DM = Diabetes Mellitus, type 1 or 2, respectively.
Fig 1.
Step-by-step flowchart of cardiac muscle tissue homogenate preparation.
Table 2.
Mitochondrial functional indices.
Fig 2.
Mitochondrial functional indices measured by high-resolution respirometry on 2.5% homogenate of human cardiac muscle (left ventricle).
Bioenergetic parameters were assessed at baseline (State 1) and by sequential addition of 2.5 mM malate and 15 mM glutamate (so-called pseudo-State 2 respiration representing LEAK state), 2.5 mM ADP (complex I linked State 3 respiration), 10 μM cytochrome c (outer mitochondrial membrane integrity), 10 mM succinate (complex I+II linked State 3 respiration), 2.5 μM oligomycin (State 4), 1.5 μM FCCP (State 3u representing electron transfer capacity), 3.5 μM rotenone (complex I linked uncoupled respiration), 4 μM antimycin A (residual oxygen consumption representing non-mitochondrial respiration) and 1 mM KCN (to confirm that antimycin A provides an adequate non-mitochondrial background). Each concentration refers to final concentration of the agent in the chamber. Stepwise titration of the uncoupler is possible and recommended if time permits. We also verified that 2.5 μM oligomycin does not inhibit uncoupled respiration as compared to 0.5 μM oligomycin. Note: AA = antimycin A; ADP Mg = Adenosine diphosphate with Mg2+; Cyt c = cytochrome c; FCCP = uncoupler; oligo = oligomycin (F1FOATPase inhibitor); KCN = potassium cyanide.
Fig 3.
Detail of intact mitochondria in human heart muscle homogenate (from left ventricle) on electron microscopy.
Scale bar: 0.5 μm.
Table 3.
Raw data from 6 duplicate measurements (homogenate A vs homogenate B) from right atrial appendages.
Mitochondrial functional parameters. Note: ETS = Electron Transfer System, OXPHOS = oxidative phosphorylation, RCR = Respiratory Control Ratio, CI = Complex I, CII = Complex II, ROX = Residual Oxygen Consumption, ET = Electron Transfer.
Fig 4.
Comparison of atrial and ventricular mitochondrial functional indices (left atrium; left ventricle).
A) Dimensionless functional indices. B) Raw data (absolute values). C) Values corrected to citrate synthase activity. D) Values corrected to baseline respiration.
Fig 5.
A-left: Dot-plot of interclass variability of electron transfer capacity normalised to citrate synthase activity (Denoted as state 3u = uncoupled). B-right: Bland-Altman plot showing the individual differences in the same parameter between atria and ventricles. Ventricles have generally a bit higher 3u/CS than atria, but there is no dependency on absolute value.