Academic literature on the topic 'Oxidative Phosphorilation'

Abstract Jasmonates are a group of plant stress hormones, which play a role in activation of apoptosis and defense mechanisms. In vitro, it has been found that methyl jasmonate MJ induces death of lymphoma and leukemia cells by independent mechanisms involving mitochondrial damage at the level of the oxidative phosphorilation. Since CLL is a suitable model to study the effect of MJ on circulating leukemic cells, we analyzed the in vitro apoptosis and necrosis of B-CLL cells treated with MJ by PI/ Annexin V analysis and cytotoxicity by tetrazolium-based assay (XTT test). In comparison normal lymphocytes from healthy donors were also analyzed. Interestingly, normal lymphocytes were not affected al all by methyl jasmonate. Blood samples from 28 CLL patients in advanced stages not treated or at least 4 weeks after cessation of treatment were selected and incubated with MJ. The proportion of cells undergoing apoptosis or necrosis following 4 hour of incubation was measured by flow cytometry and considered as positive if >35% cells death occurred. Following 24 hours incubation cells vialability was measured using XTT Cell Proliferation kit assay by Elisa. Cytotoxicity (%) was calculated by the ratio of optical density of control cells - drug-treated cells/ control cells ×100 (IC50). The titration of drug showed that 1 mM of MJ was the most effective to induce apoptosis of CLL cells and has no effect on normal lymphocytes. In 90% of samples the specific cells death tested by Annexin V was higher than 35%. A significant cytotoxicity was observed in 60% of the cases with an optimal titration of the drug for XTT test of 0.5 mM. Correlation between the two methods was found in 80% of the cases analyzed. Previous experiments showed that MJ induces cytochrome C release selectively in mitochondria isolated from CLL cells and high glucose levels inhibit MJ-induced ATP depletion. Furthermore MJ induces mitochondrial membrane depolarization in CLL cells. Altogether these experiments showed that MJ kills B-CLL cells through an original pathway inducing bioenergitic cell collapse. The cytotoxic effect and specific cell death tested in the present analysis support the concept that MJ is a potential drug to treat B-CLL. A clinical trial is currently carried out in our Institution.

In the body, 2-methoxyethanol compound may be converted to MAA. MAA is a strong oxidant and may cause oxidation stress in spermatozoa. Oxidation stress is a disturbance on phosphorilation that increases ROS concentration, and it produces lipid peroxide in spermatozoa membrane resulted in high MDA concentration. One of indicator of spermatozoa membrane disturbances is a lack of spermatozoa membrane integrity. The main purpose of this research was to determine the relationship between MDA concentration in sperm and membrane integrity of spermatozoa in rats. The animal of treated groups (n = 40) were divide into 8 groups of 5 each. The rats were given subcutaneous injection with 0,2 ml of 200 mg/kg/day for 1 day (P1), 3 days (P2), 6 days/week (P3), and 12 days/two weeks (P4), respectively the control group was injected with physiological saline of the same volume. The concentration of MDA was measured by spectrophotometer and observing membrane integrity used HOS method to watch the spermatozoa response on hypoosmotic condition. The results of the research indicated that 2-ME caused the increasing in sperm MDA concentration and the decrease of spermatozoa membrane integrity. There was negative correlations between MDA concentration and spermatozoa membrane integrity.

Objective. To investigate in experiment a functional state of mitochondria from the brain cells after its acute ischemic injury and processes, which take place in the brain in conditions of experimental ischemia/reperfusion; to analyze a correlation of the states investigated with the ischemic-reperfusion changes in the brain of patients, having the aortal arch aneurism, operated in environment of hypothermia and artificial cerebral perfusion. Materials and metods. First stage of the investigation consisted of estimation of the cerebral cells mitochondria state in conditions of ischemia/reperfusion of the neurons primary culture, obtained from embryos of rats of the 17 - 18 gestation days, using mechanical and the enzyme dissociation in the brain cortex. The second stage of the investigation was conducted on 14 white male rats (Rattus norvegicus). In 7 animals ischemia/reperfusion of the brain was simulated, using temporary mechanical blockade of blood flow along vessels of aortal arch with its subsequent restoration, as following: thoracotomy was performed along lower edge of the third rib, making surgical access to aortal arch and main vessels with their mechanical occlusion during 30 min. Additionally in 7 experimental animals thoracotomy only without impact on cerebral blood flow was performed. After extraction of animals from the experiment the blood was collected from the heart cavities, the brain was extirpated from the skull in accordance to standard method. After finishing of the experiment MTT-test was accomplished for estimation of vital capacity of the brain neurons. The principal parameters of the mitochondria functioning and velocity of their respiration in States II and III were estimated in accordance to Chance and in a respiratory control - a ratio of respiration velocity in a State III towards respiration velocity in a State IV in accordance to Chance. Transmembranous mitochondrial potential in cellular suspension, consisting of cells of the brain. Activity of lactate dehydrogenase in the culture environment was determined. Immunohistochemical investigation of the neurons culture was conducted. Membranous mitochondrial potential was estimated in the neurons cultivated. Respiration of isolated mitochondria was measured. Energetic state of mitochondria was determined. The development degree of apoptotic processes was determined in accordance to the DNA fragmentation degree. Results. There was established, that index of respiratory control of mitochondria from the ischemized brain, measured in pH 7.4, was lowered more than by 40%, comparing with corresponding index of mitochondria, obtained from a control culture. Velocity of a completely stimulated respiration of mitochondria (State III), which were extracted from the brain in 24 h after ischemia/reperfusion, was significantly lower, than velocity of respiration in mitochondria of control cultures, witnessing presence of damage in respiratory chain and transition of electrons of cerebral cellular mitochondria in ischemia/reperfusion, as well as incapacity of such mitochondria to transit electrons effectively from oxidation substrate towards oxygen due to damage of some complexes of the electron-transporting chain. Thus, in conditions of ischemia several processes, impacting mitochondrial functioning and cellular bioenergetics, occur simultaneously. Due to simultaneous action of these processes the mitochondrial functioning fails so significantly, that becomes one of the key moments in cerebral cellular dysfunction and death. Statistically meaningful difference, taking into account the results, between two groups of investigated animals was not revealed. Conclusion. In environment of ischemia/reperfusion of the brain in its cells a significant disorders of mitochondrial function occur, which are demonstrated by raising of generation of the oxygen active forms, and by lowering of transmembranous mitochondrial potential and velocity of mitochondrial respiration by 40%, what is accompanied by development of lactate-acidosis. It is expedient to apply a mitochondria-directed antioxidants in complex of the treatment measures in the brain ischemia/reperfusion, as well as desintegrators of respiration and oxidated phosphorilation.

The authors discuss about the relationships between oculoauriculo- vertebral spectrum (OAVS), a malformative condition affecting eyes, ears and vertebral structures mainly derived from neural crests, and autosomal dominant progressive external ophthalmoplegia (adPEO) – an electron transport chain disorder characterized by palpebral ptosis and muscular symptoms – on the basis of laboratory data and clinical signs in a case belonging to a family in which symptoms of both diseases occur. Hemifacial microsomia and ears abnormalities were the main neural crest cells (NCC) derived signs for OAVS diagnosis. Palpebral ptosis and the associated hyperlactacidemia with high Lactate to Pyruvate ratio were main signs for the suspicion of adPEO, but the diagnosis was proposed by clinical, biochemical, electrophysiological and genetic methods because the biopsy not always is diagnostic, as in our case. The association of two groups of symptoms (malformed, NCCrelated, and myopatic-electron transport related) in the same subject as observed in our case and partially in the family, may be ascribed to nuclear abnormalities and mitochondrial electron transport disorder, respectively, as well as to a possible common etiopathogenetic mechanism. In some cases of OAVS, the malformations may be consequent to oxidative phosphorilation disease (OXPHOS) derangement, as previously demonstrated; the extensive determination of lactacidemia may be important to address the research on OAVS patients in the mitochondrial/OXPHOS genetic domain, with advantage in the knowledge of OAVS pathogenesis.

Beside its apoptotic function, the Apoptosis Inducing Factor (AIF), a highly conserved mitochondrial flavoprotein, plays a pro-vital role in eukaryotic cells through its interaction with CHCHD4, a mitochondrial protein that contributes to oxidative folding of respiratory complexes’ subunits. A unique feature of AIF is the ability to form a tight, air-stable charge-transfer complex (CT complex) upon reaction with NAD(H), which leads to protein dimerization modulating the affinity for its ligands. To date, nine point mutations of the human AIF gene were found to cause rare and severe neurodegenerative mitochondriopathies. To define the molecular bases of the pathogenicity of AIF variants, we selected a set of AIF mutations (G337E, D236G, G261S and F133L) and investigated their effects on both AIF molecular properties and its interaction with CHCHD4. To this aim, a combination of biophysical techniques, Microscale Thermophoresis (MST) and structural biology was used. AIF variants CT complex stability was evaluated studying its reoxidation by O2. Interestingly, CT complex of G337E and G261S forms displayed a faster oxidation compared to wild type AIF, indicating a lower stability. Moreover, the 3D structures of the CT complex of AIF-D236G and of AIF-F133L both in oxidized and CT complex state were obtained, showing, however, no significant structural rearrangements with respect to the wild type protein. Hence, we investigated possible alterations of the interaction with CHCHD4 induced by AIF pathogenic mutations. Through a MST approach, for the first time we quantitatively studied redox-dependent effects of AIF amino acid replacements on its affinity for CHCHD4, revealing a possible involvement of the G337 residue in protein-protein complex formation. We characterized the AIF-CHCHD4 complex also from a structural point of view using the Small-Angle X-ray Scattering (SAXS) technique, through which we identified a putative interaction region between the two proteins. In addition, all SAXS models obtained revealed that the solvent-exposed G337 residue is localized in the neighborhood of the identified region, in agreement with a possible role of this residue in complex formation. Our results, not only shed new light on AIF-CHCHD4 relationship, but also provide a possible explanation of the pathogenicity of the AIF G337E allelic variant.