Protamine Sulfate Used in Cardiac Surgery Influences Mitochondrial Bioenergetics Profile and Induces Reactive Oxygen Species Production

Objective: Protamine sulfate is widely used albeit its pharmacological effects are not fully understood and applications are often accompanied by unwanted side effects. Here, we show the effect of protamine sulfate (PS) on mitochondrial bioenergetics profile, and the resulting mitochondrial reactive oxygen species (ROS) production.

Methods: Polarographic measurements were performed in parallel to membrane potential and ROS measurements by FACS analyzer using tetramethylrhodamine ethyl ester (TMRE) and MitoSOX fluorescent dyes, respectively.

Results: Protamine sulfate inhibited intact rat heart mitochondrial respiration stimulated by ADP to 76% (p < 0.001) from the baseline of 51.6 ± 6.9 to 12.4 ± 2.3 nmol O₂ min⁻¹·mL⁻¹. Same effect was found when respiration was priorly inhibited by antimycin A (101.0 ± 8.9 vs. 38.0 ± 9.9 nmol O₂ min⁻¹·mL⁻¹, p < 0.001) and stimulated by the addition of substrates, ascorbate and tetramethyl phenylene diamine (TMPD) for cytochrome oxidase (CytOx), suggesting that PS exerted its effect through inhibition of CytOx activity. Furthermore, the inhibition of mitochondrial respiration by PS occurred in a concentration-dependent manner and accompanied by hyperpolarization of the mitochondrial membrane potential (Δψ_m), that is, 17.4% and further 4.8% increase to control, respectively. This effect was associated with a strong consequent increase in the production of reactive oxygen species, that is, 86.4 and 89.5% compared with control, respectively.

Conclusion: We propose, the excessive increase in ROS concentrations results in mitochondrial dysfunction and thus might relate to the “protamine reaction” contributing to the development of various cardiovascular adverse findings appearing after administration of the drug.

No conflict of interest has been declared by the author(s).