Coenzyme Q deficiency in endothelial mitochondria caused by hypoxia; remodeling of the respiratory chain and sensitivity to anoxia/reoxygenation.

This study examined the effects of hypoxia on coenzyme Q (Q) levels and mitochondrial function in EA. hy926 endothelial cells, shedding light on their responses to changes in oxygen levels. Chronic hypoxia during endothelial cell culture reduced Q synthesis by reducing hydroxy-methylglutaryl-CoA reductase (HMGCR) levels via hypoxia-inducible factor 1α (HIF1α), leading to severe Q deficiency. In endothelial mitochondria, hypoxia led to reorganization of the respiratory chain through upregulation of supercomplexes (I+III2+IV), forming a complete mitochondrial Q (mQ)-mediated electron transfer pathway. Mitochondria of endothelial cells cultured under hypoxic conditions showed reduced respiratory rates and membrane potential, as well as increased production of mitochondrial reactive oxygen species (mROS) as a result of increased mQ reduction levels (mQH2/mQtot). Anoxia/reoxygenation (A/R) in vitro caused impairment of endothelial mitochondria, manifested by reduced maximal respiration, complex III activity, membrane potential, coupling parameters, and increased mQ reduction and mROS production. Weaker A/R-induced changes compared to control mitochondria indicated better tolerance of A/R stress by the mitochondria of hypoxic cells. Moreover, in endothelial mitochondria, hypoxia-induced increases in uncoupling protein 3 (UCP3) and mitochondrial large-conductance Ca2+-activated potassium channel (mitoBKCa) levels and activities appear to have alleviated reoxygenation injury after A/R. These results not only highlight hypoxia-induced changes in mQ redox homeostasis and related mitochondrial function, but also indicate that chronic hypoxia during endothelial cell culture leads to mitochondrial adaptations that help mitochondria better withstand subsequent oxygen fluctuations.

Mitochondrial Coenzyme Q Redox Homeostasis and Reactive Oxygen Species Production.

Mitochondrial coenzyme Q (mtQ) of the inner mitochondrial membrane is a redox active mobile carrier in the respiratory chain that transfers electrons between reducing dehydrogenases and oxidizing pathway(s). mtQ is also involved in mitochondrial reactive oxygen species (mtROS) formation through the mitochondrial respiratory chain. Some mtQ-binding sites related to the respiratory chain can directly form the superoxide anion from semiubiquinone radicals. On the other hand, reduced mtQ (ubiquinol, mtQH2) recycles other antioxidants and directly acts on free radicals, preventing oxidative modifications. The redox state of the mtQ pool is a central bioenergetic patameter that alters in response to changes in mitochondrial function. It reflects mitochondrial bioenergetic activity and mtROS formation level, and thus the oxidative stress associated with the mitochondria. Surprisingly, there are few studies describing a direct relationship between the mtQ redox state and mtROS production under physiological and pathological conditions. Here, we provide a first overview of what is known about the factors affecting mtQ redox homeostasis and its relationship to mtROS production. We have proposed that the level of reduction (the endogenous redox state) of mtQ may be a useful indirect marker to assess total mtROS formation. A higher mtQ reduction level (mtQH2/mtQtotal) indicates greater mtROS formation. The mtQ reduction level, and thus the mtROS formation, depends on the size of the mtQ pool and the activity of the mtQ-reducing and mtQH2-oxidizing pathway(s) of respiratory chain. We focus on a number of physiological and pathophysiological factors affecting the amount of mtQ and thus its redox homeostasis and mtROS production level.

Effects of Endurance Training on the Coenzyme Q Redox State in Rat Heart, Liver, and Brain at the Tissue and Mitochondrial Levels: Implications for Reactive Oxygen Species Formation and Respiratory Chain Remodeling.

Sixteen adult, 4-month-old male Wistar rats were randomly assigned to the training group (n = 8) or the control group (n = 8). We elucidated the effects of 8 weeks of endurance training on coenzyme Q (Q) content and the formation of reactive oxygen species (ROS) at the tissue level and in isolated mitochondria of the rat heart, liver and brain. We demonstrated that endurance training enhanced mitochondrial biogenesis in all tested organs, while a significant increase in the Q redox state was observed in the heart and brain, indicating an elevated level of QH2 as an antioxidant. Moreover, endurance training increased the mQH2 antioxidant pool in the mitochondria of the heart and liver, but not in the brain. At the tissue and isolated mitochondria level, an increase in ROS formation was only observed in the heart. ROS formation observed in the mitochondria of individual rat tissues after training may be associated with changes in the activity/amount of individual components of the oxidative phosphorylation system and its molecular organization, as well as with the size of the oxidized pool of mitochondrial Q acting as an electron carrier in the respiratory chain. Our results indicate that tissue-dependent changes induced by endurance training in the cellular and mitochondrial QH2 pool acting as an antioxidant and in the mitochondrial Q pool serving the respiratory chain may serve important roles in energy metabolism, redox homeostasis and the level of oxidative stress.

The Relationship between Mitochondrial Reactive Oxygen Species Production and Mitochondrial Energetics in Rat Tissues with Different Contents of Reduced Coenzyme Q.

We investigated the relationship between mitochondrial production of reactive oxygen species (ROS) and mitochondrial energetics in various rat tissues with different contents of the reduced coenzyme Q (Q) pool (Q9 + Q10). Our results indicate that similar to the tissue level, mitochondrial H2O2 release under nonphosphorylating conditions was strongly dependent on the amount of the reduced Q pool. Namely, in brain and lung mitochondria, less H2O2 release corresponded to a less reduced Q pool, while in liver and heart mitochondria, higher H2O2 release corresponded to a more reduced Q pool. We can conclude that the differences observed in rat tissues in the size of the reduced Q pool reflect different levels of ROS production and hence may reflect different demands for reduced Q as an antioxidant. Moreover, differences in mitochondrial H2O2 release were observed in different types of rat mitochondria during the oxidation of succinate (complex II substrate), malate plus glutamate (complex I substrate), and their mixture under phosphorylating and nonphosphorylating conditions. Our results indicate the existence of a tissue-specific maximum respiratory chain capacity in ROS production, possibly related to the membrane potential-mediated control of oxidative phosphorylation. We propose the use of a new parameter for the study of isolated mitochondria, RCRROS, the ratio between the formation of mitochondrial ROS under nonphosphorylating and phosphorylating conditions, which represents the maximum factorial increase in mitochondrial ROS formation that can be achieved after all ADP is phosphorylated.

Lung mitochondria adaptation to endurance training in rats.

We elucidated the impact of eight weeks of endurance training on the oxidative metabolism of rat lungs. Adult 3.5-month-old male rats were randomly allocated to a treadmill training group or a sedentary group as control. In the lungs, endurance training raised the expression level of the oxygen sensors hypoxia inducible factor 1α (HIF1α) and lysine-specific demethylase 6A (KDM6A) as well as stimulated mitochondrial oxidative capacity and mitochondrial biogenesis, while lactate dehydrogenase activity was reduced. Endurance training enhanced antioxidant systems (the coenzyme Q content and superoxide dismutase) in lung tissue but decreased them (and uncoupling protein 2) in lung mitochondria. In the lung mitochondria of trained rats, the decreased Q content and Complex I (CI) activity and the enhanced cytochrome pathway activity (CIII + CIV) may account for the diminished Q reduction level, resulting in a general decrease in H2O2 formation by mitochondria. Endurance training enhanced oxidation of glutamate and fatty acids and caused opposite effects in functional mitochondrial properties during malate and succinate oxidation, which were related to reduced activity of CI and increased activity of CII, respectively. In addition, endurance training downregulated CI in supercomplexes and upregulated CIII in the CIII2+CIV supercomplex in the oxidative phosphorylation system. We concluded that the adaptive lung responses observed could be due to hypoxia and oxidative stress induced by strenuous endurance training.

The Influence of Statins on the Aerobic Metabolism of Endothelial Cells.

Endothelial mitochondrial dysfunction is considered to be the main cause of cardiovascular disease. The aim of this research was to elucidate the effects of cholesterol-lowering statins on the aerobic metabolism of endothelial cells at the cellular and mitochondrial levels. In human umbilical vein endothelial cells (EA.hy926), six days of exposure to 100 nM atorvastatin (ATOR) induced a general decrease in mitochondrial respiration. No changes in mitochondrial biogenesis, cell viability, or ATP levels were observed, whereas a decrease in Coenzyme Q10 (Q10) content was accompanied by an increase in intracellular reactive oxygen species (ROS) production, although mitochondrial ROS production remained unchanged. The changes caused by 100 nM pravastatin were smaller than those caused by ATOR. The ATOR-induced changes at the respiratory chain level promoted increased mitochondrial ROS production. In addition to the reduced level of mitochondrial Q10, the activity of Complex III was decreased, and the amount of Complex III in a supercomplex with Complex IV was diminished. These changes may cause the observed decrease in mitochondrial membrane potential and an increase in Q10 reduction level as a consequence, leading to elevated mitochondrial ROS formation. The above observations highlight the role of endothelial mitochondria in response to potential metabolic adaptations related to the chronic exposure of endothelial cells to statins.

[Different faces of the mitochondrial coenzyme Q]. / Rózne oblicza mitochondrialnego koenzymu Q.

Coenzyme Q is a fat-soluble molecule present in all cell membranes, including the inner mitochondrial membrane. Mitochondrial Q (mQ) is a key electron carrier in the respiratory chain and an important antioxidant. On the other hand, mQ participates in the production by respiratory chain of mitochondrial reactive oxygen species (mROS) that are formed as a byproduct of oxygen metabolism or under oxidative stress conditions. Increased mROS production can lead to a series of oxidative damage that underlies cell aging or a number of diseases. In addition, mROS act as signaling molecules. Respiratory chain electron carriers, primarily mQ-related protein complexes, are considered the main mROS production sites. With age, the level of Q, and in particular its reduced form, decreases in the body. Disorders associated with coenzyme Q deficiency are mainly associated with excessive mROS production and a decrease in ATP production, which may result in mitochondrial, cardiovascular or neurodegenerative diseases.

The interplay between mitochondrial reactive oxygen species formation and the coenzyme Q reduction level.

Our aim was to elucidate the relationship between the rate of mitochondrial reactive oxygen species (mROS) formation and the reduction level of the mitochondrial coenzyme Q (mQ) pool under various levels of engagement of the mQ-reducing pathway (succinate dehydrogenase, complex II) and mQH2-oxidizing pathways (the cytochrome pathway and alternative oxidase pathway, (AOX)) in mitochondria isolated from the amoeba Acanthamoeba castellanii. The mQ pool was shifted to a more reduced state by inhibition of mQH2-oxidizing pathways (complex III and complex IV of the cytochrome pathway, and AOX) and the oxidative phosphorylation system. The mQ reduction level was lowered by decreasing the electron supply from succinate dehydrogenase and by stimulating the activity of the cytochrome or AOX pathways. The results indicate a direct dependence of mROS formation on the reduction level of the mQ pool for both mQH2-oxidizing pathways. A higher mQ reduction level leads to a higher mROS formation. For the cytochrome pathway, mROS generation depends nonlinearly upon the mQ reduction level, with a stronger dependency observed at values higher than the mQ reduction level of the phosphorylating state (~ 35%). AOX becomes more engaged at higher mQ pool reduction levels (above 40%), when mROS production via the cytochrome pathway increases. We propose that the mQ pool reduction level (endogenous mQ redox state) could be a useful endogenous reporter that allows indirect assessment of overall mROS production in mitochondria.