Iron Deficiency Impairs Contractility of Human Cardiomyocytes Through Decreased Mitochondrial Function

Figures

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  Figure 1. In vitro iron deficiency is obtained through deferoxamine (DFO) incubation. Ferritin levels decrease in a time‐dependent fashion during DFO incubation (A). Low iron levels lead to induced gene transcription levels of genes involved in iron uptake, transport, and storage (B). (C) Hypoxia‐inducible factor 1 alpha (HIF1α) protein levels in relation with α‐tubulin levels during DFO incubation. N.D., not determined; SLC11A2, solute carrier family 11 member 2; SLC39A14, solute carrier family 39 member 14; TfRC, transferrin receptor. **P < 0.01; ***P < 0.001; ****P < 0.0001.

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  Figure 2. The effect of iron deficiency on mitochondrial function. Decreasing levels of intracellular iron correlate with ATP levels (A). Representative traces for control cardiomyocytes and cardiomyocytes treated with deferoxamine (DFO) for 2 and 4 days in a Mito Stress test (B). Effects of iron deficiency on ATP‐linked respiration and respiratory reserve are shown in (C). The enzymatic activity of each individual mitochondrial complex was analysed (D). OCR, oxygen consumption rate. **P < 0.01; ***P < 0.001; ****P < 0.0001.

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  Figure 3. Effects of iron depletion are reversible by transferrin (Tf) administration. Following Tf‐bound iron supplementation, levels of ferritin (A), gene expression (B), mitochondrial respiration (C), of which ATP‐linked respiration and respiratory reserve shown in detail (D), hypoxia‐inducible factor 1 alpha (HIF1α) protein (E), and ATP (F) were mostly found to be restored. AntA, antimycin A; FCCP, carbonyl cyanide p‐tri‐fluoromethoxy‐phenyl‐hydrazone; OCR, oxygen consumption rate; Oligo, oligomycin; Rot, rotenone. *P < 0.05; ***P < 0.001; ****P < 0.0001.

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  Figure 4. Mitochondrial morphology is also affected by iron deficiency. Mitochondria in iron‐deficient cardiomyocytes appear swollen and contain electron dense inclusion bodies (A), which were found to contain nitrogen and sulfur based on energy dispersive X‐ray (EDX) analysis (B). Scale bar = 1 μm. DFO, deferoxamine.

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  Figure 5. Low iron levels resulted in reduced contractile force and impaired systolic and diastolic velocity. The fractional area change (FAC) of a single contraction for each condition (A) show that iron deficiency impairs contractile force (B). FAC, and maximum systolic and diastolic velocities (Vmax) are affected by low iron levels, but are restored upon addition of transferrin (Tf)‐bound iron. *P < 0.05; **P < 0.01; ***P < 0.001. DFO, deferoxamine.

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  Figure 6. Reversible morphological aberrations during iron deficiency. Electron micrographs of (A) control, (B) iron‐deficient, and (C) transferrin‐treated cardiomyocytes. N, nucleus; G, glycogen; V, vacuole. Scale bar = 2 μm.