High cardiac energy demands are predominantly provided by oxidative respiration within mitochondria. In the failing heart, decreased energy production in cardiomyocytes due to impaired mitochondrial function may result in impaired contractile capacity. Dysregulation of mitochondrial phospholipid metabolism has negative effects on mitochondrial function resulting in energy deficiency, oxidative stress, and apoptosis, phenomena well described in heart failure.

We investigated lipidome changes in cardiomyocytes and their association with mitochondrial function in an isoproterenol (ISO)-induced cardiomyopathy model. Eight weeks old male 129sv mice were subcutaneously injected with ISO/vehicle for four consecutive days. Mice were sacrificed on day four, hearts were removed, and cardiomyocytes were isolated using the Langendorff perfusion apparatus. Shotgun lipidomics analysis using a Q Exactive mass spectrometer in Quadrupole-Orbitrap configuration were conducted in primary isolated cardiomyocytes (PCMs) and subsequent lipidomics-tailored bioinformatics analysis was performed. Mitochondrial function in PCMs was assed using the Seahorse Xfe96 analyzer. In addition, we deleted cardiolipin synthase by RNAi in hearts from Drosophila melanogaster and analyzed cardiac function by high-resolution videomicroscopy.

Our data revealed that four days ISO treatment induces cardiac lipidome changes in mice. Among the regulated lipid species, cardiolipins (CL 72:8, 72:9) where significantly downregulated in comparison to vehicle. CLs are localized in the inner membrane of the mitochondria and play major roles in energy metabolism. Altered CL profile has been reported to result in mitochondrial dysfunction and oxidative stress, which has been linked to various cardiovascular diseases. Assessment of mitochondrial function revealed that ISO leads to elevation in oxygen consumption rates, manifested by increased maximal respiration and spare respiratory capacity. However, the extracellular acidification rates were increased under ISO treatment. Moreover, a significant increase in NADPH oxidase 4 (Nox4) in PCMs was observed upon ISO treatment. Serum levels of malondialdehyde assessed using the thiobarbituric acid reactive substances assay were significantly increased by ISO. Cardiac expression of CLS1 was decreased in ISO-treated mice.

CLS1-knockdown in Drosophila led to an impairment of systolic function and an enlargement of diastolic/ systolic diameters of the fly heart.

In summary, our data indicate that the regulation of mitochondrial CLs is associated with mitochondrial dysfunction and oxidative stress in PCMs during the development of ISO-induced cardiomyopathy in mice. Furthermore, the lack of the CL-synthesizing enzyme CLS1 in flies results in cardiac dysfunction. Taken together, we identify CLs as potential mediators of catecholamine-induced cardiac damage.