Mitochondrion-targeted antioxidant SkQ1 prevents rapid animal death caused by highly diverse shocks.

The response to stress involves the activation of pathways leading either to protection from the stress origin, eventually resulting in development of stress resistance, or activation of the rapid death of the organism. Here we hypothesize that mitochondrial reactive oxygen species (mtROS) play a key role in stress-induced programmed death of the organism, which we called "phenoptosis" in 1997. We demonstrate that the synthetic mitochondria-targeted antioxidant SkQ1 (which specifically abolishes mtROS) prevents rapid death of mice caused by four mechanistically very different shocks: (a) bacterial lipopolysaccharide (LPS) shock, (b) shock in response to intravenous mitochondrial injection, (c) cold shock, and (d) toxic shock caused by the penetrating cation C12TPP. Importantly, under all these stresses mortality was associated with a strong elevation of the levels of pro-inflammatory cytokines and administration of SkQ1 was able to switch off the cytokine storms. Since the main effect of SkQ1 is the neutralization of mtROS, this study provides evidence for the role of mtROS in the activation of innate immune responses mediating stress-induced death of the organism. We propose that SkQ1 may be used clinically to support patients in critical conditions, such as septic shock, extensive trauma, cooling, and severe infection by bacteria or viruses.

Comparison of the Effects of Mitochondria-Targeted Antioxidant 10-(6'-Plastoquinonyl)Decyltriphenylphosphonium Bromide (SkQ1) and a Fragment of its Molecule Dodecyltriphenylphosphonium on Carrageenan-Induced Acute Inflammation in Mouse Model of Subcuteneous Air Pouch.

The effect of mitochondria-targeted antioxidant 10-(6'-plastoquinonyl) decyltriphenylphosphonium bromide (SkQ1) and its fragment dodecyltriphenylphosphonium (C12TPP), weak uncouplers of respiration and oxidative phosphorylation, was studied using a mouse model of carrageenan-induced acute inflammation in the subcutaneous air pouch. In our model, SkQ1 demonstrated a strong anti-inflammatory effect that manifested in a decrease in the absolute number of inflammatory cells, mainly neutrophils, and their relative number in parallel with an increase in macrophages and mast cell content in the inflammatory exudate. The concentration of proinflammatory cytokine IL-6 in the exudate also tended to decrease. C12TPP produced no significant effect on the inflammation process.

Emerging role of Cdc42-specific guanine nucleotide exchange factors as regulators of membrane trafficking in health and disease.

It is widely accepted that the Golgi complex operates as a main sorting station in the biosynthetic pathway. On the other hand, the Golgi complex harbors numerous signaling molecules that generate the platform for the coordination of the transduction of specific signals and of membrane transport events. A part of these processes, which require the complex integration of transport-, cytoskeleton- and polarity-associated mechanisms, is tightly regulated by molecular machineries comprising guanine nucleotide exchange factors (GEF) and their down-stream effectors, such as the small GTPase Cdc42. Dysfunction of several Cdc42-specific GEFs has been shown to cause a number of human diseases, which are associated with impaired intracellular trafficking at the level of the Golgi complex as well as in other compartments. Here we briefly overview how mutations in Cdc42-specific GEFs have an impact on the organization of intracellular trafficking fluxes and how such trafficking aberrations could be associated with a number of human disorders.

[Changes in Spatial Organization of Cortical Rhythm Vibrations in Children uner the Influence of Music].

Changes is systemic brain activity under influence of classical music (minor and major music) were studied at two groups of healthy children aged 5-6 years (n = 53). In 25 of studied children the Luscher test showed increased level of anxiety which significantly decreased after music therapy sessions. Bioelectrical cortical activity registered from 20 unipolar leads was subjected to correlation, coherence and factor analysis. Also the dynamics of the power spectrum for each of the EEG was studied. According to EEG all children after listening to both minor and major tones showed reorganization of brain rhythm structure accompanied by a decrease in the level of coherence and correlation of EEG; also was found significant and almost universal decrease in the EEG power spectrum. Registered EEG changes under the influence of classical music seems to reflect a decrease in excess of "internal tension" and weakening degree of "stiffness" to ensure the activity of cerebral structures responsible for mechanisms of "basic integration" which maintain constant readiness of brain to rapid and complete inclusion in action.

Age-associated murine cardiac lesions are attenuated by the mitochondria-targeted antioxidant SkQ1.

Age-related changes in mammalian hearts often result in cardiac hypertrophy and fibrosis that are preceded by inflammatory infiltration. In this paper, we show that lifelong treatment of BALB/c and C57BL/6 mice with the mitochondria-targeted antioxidant SkQ1 retards senescence-associated myocardial disease (cardiomyopathy), cardiac hypertrophy, and diffuse myocardial fibrosis. To investigate the molecular basis of the action of SkQ1, we have applied DNA microarray analysis. The global gene expression profile in heart tissues was not significantly affected by administration of SkQ1. However, we found some small but statistically significant modifications of the pathways related to cell-to-cell contact, adhesion, and leukocyte infiltration. Probably, SkQ1-induced decrease in leukocyte and mesenchymal cell adhesion and/or infiltration lead to a reduction in age-related inflammation and subsequent fibrosis. The data indicate a causative role of mitochondrial reactive oxygen species in cardiovascular aging and imply that SkQ1 has potential as a drug against age-related cardiac dysfunction.

Effect of the mitochondria-targeted antioxidant SkQ1 on development of spontaneous tumors in BALB/c mice.

The mitochondria-targeted antioxidant SkQ1 (10-(6'-plastoquinonyldecyl)triphenylphosphonium) is a new pharmaceutical substance with a wide spectrum of effects including increase in lifespan of laboratory animals (for example, of BALB/c mice males) and inhibition of development of some experimental tumors and also of tumor cell growth. In this work, the effects of SkQ1 on development of spontaneous tumors in female and male BALB/c mice housed in an SPF-class vivarium were studied. We found that the addition of SkQ1 to drinking water at the dose of 1 and 30 nmol/kg body weight per day throughout the lifespan modified the spectrum of spontaneous tumors in the female mice, decreasing the incidence of follicular lymphomas. SkQ1 at the dose of 1 nmol/kg per day also suppressed the dissemination of these neoplasms, but it did not significantly influence the overall incidence of benign and malignant tumors (including primary multiple tumors) or the lifespan of the tumor-bearing mice (both males and females). Hence, the previously described ability of SkQ1 to increase the lifespan of laboratory BALB/c mice is not related to its anticarcinogenic activity.

Mitochondrial-targeted plastoquinone derivatives. Effect on senescence and acute age-related pathologies.

Plastoquinone, a very effective electron carrier and antioxidant of chloroplasts, was conjugated with decyltriphenylphosphonium to obtain a cation easily penetrating through membranes. This cation, called SkQ1, is specifically targeted to mitochondria by electrophoresis in the electric field formed by the mitochondrial respiratory chain. The respiratory chain also regenerates reduced SkQ1H(2) from its oxidized form that appears as a result of the antioxidant activity of SkQ1H(2). SkQ1H(2) prevents oxidation of cardiolipin, a mitochondrial phospholipid that is especially sensitive to attack by reactive oxygen species (ROS). In cell cultures, SkQ1 and its analog plastoquinonyl decylrhodamine 19 (SkQR1) arrest H(2)O(2)-induced apoptosis. When tested in vivo, SkQs (i) prolong the lifespan of fungi, crustaceans, insects, fish, and mice, (ii) suppress appearance of a large number of traits typical for age-related senescence (cataract, retinopathies, achromotrichia, osteoporosis, lordokyphosis, decline of the immune system, myeloid shift of blood cells, activation of apoptosis, induction of ß-galactosidase, phosphorylation of H2AX histones, etc.) and (iii) lower tissue damage and save the lives of young animals after treatments resulting in kidney ischemia, rhabdomyolysis, heart attack, arrhythmia, and stroke. We suggest that the SkQs reduce mitochondrial ROS and, as a consequence, inhibit mitochondria-mediated apoptosis, an obligatory step of execution of programs responsible for both senescence and fast "biochemical suicide" of an organism after a severe metabolic crisis.

Novel mitochondria-targeted antioxidants, "Skulachev-ion" derivatives, accelerate dermal wound healing in animals.

It is shown that the novel mitochondria-targeted antioxidant SkQ1, (10-(6'-plastoquinonyl) decyltriphenylphosphonium) stimulates healing of full-thickness dermal wounds in mice and rats. Treatment with nanomolar doses of SkQ1 in various formulations accelerated wound cleaning and suppressed neutrophil infiltration at the early (7 h) steps of inflammatory phase. SkQ1 stimulated formation of granulation tissue and increased the content of myofibroblasts in the beginning of regenerative phase of wound healing. Later this effect caused accumulation of collagen fibers. Local treatment with SkQ1 stimulated re-epithelization of the wound. Lifelong treatment of mice with SkQ1 supplemented with drinking water strongly stimulated skin wounds healing in old (28 months) animals. In an in vitro model of wound in human cell cultures, SkQ1 stimulated movement of epitheliocytes and fibroblasts into the "wound". Myofibroblast differentiation of subcutaneous fibroblasts was stimulated by SkQ1. It is suggested that SkQ1 stimulates wound healing by suppression of the negative effects of oxidative stress in the wound and also by induction of differentiation. Restoration of regenerative processes in old animals is consistent with the "rejuvenation" effects of SkQ1, which prevents some gerontological diseases.

Mitochondria-targeted plastoquinone derivatives as tools to interrupt execution of the aging program. 2. Treatment of some ROS- and age-related diseases (heart arrhythmia, heart infarctions, kidney ischemia, and stroke).

Effects of 10-(6'-plastoquinonyl) decyltriphenylphosphonium (SkQ1) and 10-(6'-plastoquinonyl) decylrhodamine 19 (SkQR1) on rat models of H2O2- and ischemia-induced heart arrhythmia, heart infarction, kidney ischemia, and stroke have been studied ex vivo and in vivo. In all the models listed, SkQ1 and/or SkQR1 showed pronounced protective effect. Supplementation of food with extremely low SkQ1 amount (down to 0.02 nmol SkQ1/kg per day for 3 weeks) was found to abolish the steady heart arrhythmia caused by perfusion of isolated rat heart with H2O2 or by ischemia/reperfusion. Higher SkQ1 (125-250 nmol/kg per day for 2-3 weeks) was found to decrease the heart infarction region induced by an in vivo ischemia/reperfusion and lowered the blood levels of lactate dehydrogenase and creatine kinase increasing as a result of ischemia/reperfusion. In single-kidney rats, ischemia/reperfusion of the kidney was shown to kill the majority of the animals in 2-4 days, whereas one injection of SkQ1 or SkQR1 (1 micromol/kg a day before ischemia) saved lives of almost all treated rats. Effect of SkQR1 was accompanied by decrease in ROS (reactive oxygen species) level in kidney cells as well as by partial or complete normalization of blood creatinine and of some other kidney-controlled parameters. On the other hand, this amount of SkQ1 (a SkQ derivative of lower membrane-penetrating ability than SkQR1) saved the life but failed to normalize ROS and creatinine levels. Such an effect indicates that death under conditions of partial kidney dysfunction is mediated by an organ of vital importance other than kidney, the organ in question being an SkQ1 target. In a model of compression brain ischemia/reperfusion, a single intraperitoneal injection of SkQR1 to a rat (1 micromol/kg a day before operation) effectively decreased the damaged brain area. SkQ1 was ineffective, most probably due to lower permeability of the blood-brain barrier to this compound.

[Prognostic value of structural-functional characteristics of skin melanoma].

Surgical material is investigated. Morphometric criteria of ultrastructural atypia are considered as a possible prognostic factor of melanoma lymphogenic metastases. This allows detection of skin melanoma with high metastatic potential.

[Expression of ouabain-sensitive H+,K+-ATPase catalytic subunit gene in the rat epidermis]. / Ekspressiia gena kataliticheskoi sub''edinitsy uabainchuvstvitel'noi H+,K+-ATP-azy v épidermise kozhi krysy.

A comparative localization of Na+,K(+)-ATPase and ouabain-sensitive H+,K(+)-ATPase in rat skin was performed using in situ RNA hybridization and immunohistochemistry. Na+,K(+)-ATPase was predominantly detected in the basal layer of epithelium, whereas the ouabain-sensitive H+,K(+)-ATPase, in the granular and prickle cell layers. The genes of these ATPases are thus expressed in epithelial cells at different stages of their development. The hypothesis was advanced that the ouabain-sensitive H+,K(+)-ATPase is involved in maintaining the skin pH value. The probes specific to the mRNAs of the full-size alpha-subunit of the ouabain-sensitive H+,K(+)-ATPase and its truncated form were used to establish a similar distribution of both mRNA variants in skin. The English version of the paper: Russian Journal of Bioorganic Chemistry, 2002, vol. 28, no. 4; see also http://www.maik.ru.

[Identification of Escherichia coli nitrate reductase as an antigen for a monoclonal antibody with previously unknown specificity]. / Identifikatsiia nitratreduktazy Escherichia coli kak antigena monoklonal'nogo antitela neizvestnoi ranee spetsifichnosti.

The immunoaffinity chromatography of total membrane proteins from Escherichia coli helped determine the specificity of the monoclonal antibody 3A6 that was obtained upon immunization of mice with nicotinamide nucleotide transhydrogenase preparations and reacted with an unknown E. coli antigen. Proteins with apparent molecular masses of 150, 45, and 20 kDa were isolated and identified by N-terminal sequencing as the subunits of nitrate reductase. This conclusion was confirmed by immunoblotting with the 3A6 antibody of the proteins from the E. coli cells grown upon induction of nitrate reductase. It was shown that the 3A6 antibody specifically recognizes the alpha subunit of nitrate reductase, and the formation of the enzyme-antibody complex does not result in a loss of the enzyme catalytic activity.

Interactions between the soluble domain I of nicotinamide nucleotide transhydrogenase from Rhodospirillum rubrum and transhydrogenase from Escherichia coli. Effects on catalytic and H+-pumping activities.

Nicotinamide nucleotide transhydrogenase from Escherichia coli is composed of two subunits, the alpha and the beta subunits, each of which contains a hydrophilic domain, domain I and III, respectively, as well as several transmembrane helices, collectively denoted domain II. The interactions between domain I from Rhodospirillum rubrum (rrI) and the intact or the protease-treated enzyme from E. coli was investigated using the separately expressed and purified domain I from R. rubrum, and His-tagged intact and trypsin-treated E. coli transhydrogenase. Despite harsh treatments with, e.g. detergents and denaturing agents, the alpha and beta subunits remained tightly associated. A monoclonal antibody directed towards the alpha subunit was strongly inhibitory, an effect that was relieved by added rrI. In addition, rrI also reactivated the trypsin-digested E. coli enzyme in which domain I had been partly removed. This suggests that the hydrophilic domains I and III are not in permanent contact but are mobile during catalysis while being anchored to domain II. Replacement of domain I of intact, as well as trypsin-digested, E. coli transhydrogenase with rrI resulted in a markedly different pH dependence of the cyclic reduction of 3-acetyl-pyridine-NAD+ by NADH in the presence of NADP(H), suggesting that the protonation of one or more protonable groups in domain I is controlling this reaction. The reverse reaction and proton pumping showed a less pronounced change in pH dependence, demonstrating the regulatory role of domain II in these reactions.

Ouabain-sensitive H,K-ATPase: tissue-specific expression of the mammalian genes encoding the catalytic alpha subunit.

Human ATP1AL1 and corresponding genes of other mammals encode the catalytic alpha subunit of a non-gastric ouabain-sensitive H,K-ATPases, the ion pump presumably involved in maintenance of potassium homeostasis. The tissue specificity of the expression of these genes in different species has not been analyzed in detail. Here we report comparative RT-PCR screening of mouse, rat, rabbit, human, and dog tissues. Significant expression levels were observed in the skin, kidney and distal colon of all species (with the exception of the human colon). Analysis of rat urogenital organs also revealed strong expression in coagulating and preputial glands. Relatively lower expression levels were detected in many other tissues including brain, placenta and lung. In rabbit brain the expression was found to be specific to choroid plexus and cortex. Prominent similarity of tissue-specific expression patterns indicates that animal and human non-gastric H,K-ATPases are indeed products of homologous genes. This is also consistent with the high sequence similarity of non-gastric H,K-ATPases (including partial sequences of hitherto unknown cDNAs for mouse and dog proteins).