Effect of 2-aminoethoxydiphenyl borate on the functions of mouse skeletal muscle mitochondria.

This work examined the effect of 2-aminoethoxydiphenyl borate (2-APB) on the functioning of isolated mouse skeletal muscle mitochondria and modeled its putative interaction with mitochondrial proteins. We have shown that 2-APB is able to dose-dependently suppress mitochondrial respiration in state 3 and 3UDNP driven by substrates of complex I and II. This effect of 2-APB was accompanied by a slight dose-dependent decrease in mitochondrial membrane potential and appears to be due to inhibition of complex I and complex III of the electron transport chain (ETC) with IC50 values of 200 and 120 µM, respectively. The results of molecular docking identified putative 2-APB interaction sites in these ETC complexes. 2-APB was shown to dose-dependently inhibit both mitochondrial Ca2+ uptake and Ca2+ efflux, which seems to be caused by a decrease in the membrane potential of the organelles. We have found that 2-APB has no significant effect on mitochondrial calcium retention capacity. On the other hand, 2-APB exhibited antioxidant effect by reducing mitochondrial hydrogen peroxide production but without affecting superoxide generation. It is concluded that the effect of 2-APB on mitochondrial targets should be taken into account when interpreting the results of cell and in vivo experiments.

Mitochondrial Transplantation Therapy Ameliorates Muscular Dystrophy in mdx Mouse Model.

Duchenne muscular dystrophy is caused by loss of the dystrophin protein. This pathology is accompanied by mitochondrial dysfunction contributing to muscle fiber instability. It is known that mitochondria-targeted in vivo therapy mitigates pathology and improves the quality of life of model animals. In the present work, we applied mitochondrial transplantation therapy (MTT) to correct the pathology in dystrophin-deficient mdx mice. Intramuscular injections of allogeneic mitochondria obtained from healthy animals into the hind limbs of mdx mice alleviated skeletal muscle injury, reduced calcium deposits in muscles and serum creatine kinase levels, and improved the grip strength of the hind limbs and motor activity of recipient mdx mice. We noted normalization of the mitochondrial ultrastructure and sarcoplasmic reticulum/mitochondria interactions in mdx muscles. At the same time, we revealed a decrease in the efficiency of oxidative phosphorylation in the skeletal muscle mitochondria of recipient mdx mice accompanied by a reduction in lipid peroxidation products (MDA products) and reduced calcium overloading. We found no effect of MTT on the expression of mitochondrial signature genes (Drp1, Mfn2, Ppargc1a, Pink1, Parkin) and on the level of mtDNA. Our results show that systemic MTT mitigates the development of destructive processes in the quadriceps muscle of mdx mice.

Effect of Large-Conductance Calcium-Dependent K+ Channel Activator NS1619 on Function of Mitochondria in the Heart of Dystrophin-Deficient Mice.

Dystrophin-deficient muscular dystrophy (Duchenne dystrophy) is characterized by impaired ion homeostasis, in which mitochondria play an important role. In the present work, using a model of dystrophin-deficient mdx mice, we revealed decrease in the efficiency of potassium ion transport and total content of this ion in the heart mitochondria. We evaluated the effect of chronic administration of the benzimidazole derivative NS1619, which is an activator of the large-conductance Ca2+-dependent K+ channel (mitoBKCa), on the structure and function of organelles and the state of the heart muscle. It was shown that NS1619 improves K+ transport and increases content of the ion in the heart mitochondria of mdx mice, but this is not associated with the changes in the level of mitoBKCa protein and expression of the gene encoding this protein. The effect of NS1619 was accompanied by the decrease in the intensity of oxidative stress, assessed by the level of lipid peroxidation products (MDA products), and normalization of the mitochondrial ultrastructure in the heart of mdx mice. In addition, we found positive changes in the tissue manifested by the decrease in the level of fibrosis in the heart of dystrophin-deficient animals treated with NS1619. It was noted that NS1619 had no significant effect on the structure and function of heart mitochondria in the wild-type animals. The paper discusses mechanisms of influence of NS1619 on the function of mouse heart mitochondria in Duchenne muscular dystrophy and prospects for applying this approach to correct pathology.

BKCa Activator NS1619 Improves the Structure and Function of Skeletal Muscle Mitochondria in Duchenne Dystrophy.

Duchenne muscular dystrophy (DMD) is a progressive hereditary disease caused by the absence of the dystrophin protein. This is secondarily accompanied by a dysregulation of ion homeostasis, in which mitochondria play an important role. In the present work, we show that mitochondrial dysfunction in the skeletal muscles of dystrophin-deficient mdx mice is accompanied by a reduction in K+ transport and a decrease in its content in the matrix. This is associated with a decrease in the expression of the mitochondrial large-conductance calcium-activated potassium channel (mitoBKCa) in the muscles of mdx mice, which play an important role in cytoprotection. We observed that the BKCa activator NS1619 caused a normalization of mitoBKCa expression and potassium homeostasis in the muscle mitochondria of these animals, which was accompanied by an increase in the calcium retention capacity, mitigation of oxidative stress, and improvement in mitochondrial ultrastructure. This effect of NS1619 contributed to the reduction of degeneration/regeneration cycles and fibrosis in the skeletal muscles of mdx mice as well as a normalization of sarcomere size, but had no effect on the leakage of muscle enzymes and muscle strength loss. In the case of wild-type mice, we noted the negative effect of NS1619 manifested in the inhibition of the functional activity of mitochondria and disruption of their structure, which, however, did not significantly affect the state of the skeletal muscles of the animals. This article discusses the role of mitoBKCa in the development of DMD and the prospects of the approach associated with the correction of its function in treatments of this secondary channelopathy.

S-15176 Difumarate Salt Can Impair Mitochondrial Function through Inhibition of the Respiratory Complex III and Permeabilization of the Inner Mitochondrial Membrane.

S-15176 difumarate salt, a derivative of the anti-ischemic metabolic drug trimetazidine, has been intensively studied for its impact on cellular metabolism in animal models of ischemia-reperfusion injury of the liver, heart, spinal cord, and other organs. Despite evidence of some reduction in oxidative damage to cells, the results of therapy with S-15176 have been mostly disappointing, possibly because of the lack of data on its underlying mechanisms. Here, we aimed to investigate in more detail the role of complexes I-IV of the electron transport chain and membrane permeability transition in mitochondrial toxicity associated with S-15176. Using rat thymocyte and liver mitochondria, we demonstrated that: (1) acute exposure to S-15176 (10 to 50 µM) dose-dependently decreased the mitochondrial membrane potential; (2) S-15176 suppressed the ADP-stimulated (State 3) and uncoupled (State 3UDNP) respiration of mitochondria energized with succinate or malate/glutamate, but not ascorbate/TMPD, and increased the resting respiration (State 4) when using all the substrate combinations; (3) S-15176 directly inhibited the activity of the respiratory complex III; (4) low doses of S-15176 diminished the rate of H2O2 production by mitochondria; (5) at concentrations of above 30 µM, S-15176 reduced calcium retention capacity and contributed to mitochondrial membrane permeabilization. Taken together, these findings suggest that S-15176 at tissue concentrations reached in animals can impair mitochondrial function through suppression of the cytochrome bc1 complex and an increase in the nonspecific membrane permeability.