Mitochondrial ATP synthase inhibition and nitric oxide are involved in muscle weakness that occurs in acute exposure of rats to monocrotophos

Organophosphate poisoning in the context of self-harm is a common medical emergency in Asia. Prolonged muscle weakness is an important but poorly understood cause of morbidity and mortality of the poisoning. This study examined mitochondrial function and its modulation by nitric oxide in muscle weakness of rats exposed to an acute, oral (0.8LD50) dose of monocrotophos. Muscle mitochondrial ATP synthase activity was inhibited in the rat in acute exposure to monocrotophos while respiration per se was not affected. This was accompanied by decreased mitochondrial uptake of calcium and increased levels of nitric oxide. Reactive cysteine groups of ATP synthase subunits were reduced in number, which may contribute to decreased enzyme activity. The decrease in ATP synthase activity and reactive cysteine groups of ATP synthase subunits was prevented by treatment of animals with the nitric oxide synthase inhibitor, L-NG Nitroarginine methyl ester, at 12 mg/kg body weight for 9 days in drinking water, prior to monocrotophos exposure. This indicated a role for nitric oxide in the process. The alterations in mitochondrial calcium uptake may influence cytosolic calcium levels and contribute to muscle weakness of acute organophosphate exposure

Cyclodextrin Induces Calcium-Dependent Lysosomal Exocytosis

Cyclodextrins (CDs) have long been used to manipulate cellular cholesterol levels both in vitro and in vivo, but their direct effects at a cellular level are not well characterized. Recently, CDs have garnered much interest because of their ability to clear stored cholesterol from Niemann Pick Type C (NPC) cells and markedly prolong the life of NPC1 disease mice. Here, we investigate the hypothesis that treatment with 2-hydroxypropyl- β-cyclodextrin (HPB-CD) stimulates lysosomal exocytosis in a calcium-enhanced manner. We propose that this exocytosis is the mechanism by which HPB-CD ameliorates the endolysosomal cholesterol storage phenotype in NPC cells. These findings have significant implications for the use of HPB-CD in biochemical assays and data interpretation as well as for their use for the treatment for NPC and other disorders

Physical and Functional Interaction of NCX1 and EAAC1 Transporters Leading to Glutamate-Enhanced ATP Production in Brain Mitochondria

Glutamate is emerging as a major factor stimulating energy production in CNS. Brain mitochondria can utilize this neurotransmitter as respiratory substrate and specific transporters are required to mediate the glutamate entry into the mitochondrial matrix. Glutamate transporters of the Excitatory Amino Acid Transporters (EAATs) family have been previously well characterized on the cell surface of neuronal and glial cells, representing the primary players for glutamate uptake in mammalian brain. Here, by using western blot, confocal microscopy and immunoelectron microscopy, we report for the first time that the Excitatory Amino Acid Carrier 1 (EAAC1), an EAATs member, is expressed in neuronal and glial mitochondria where it participates in glutamate-stimulated ATP production, evaluated by a luciferase-luciferin system. Mitochondrial metabolic response is counteracted when different EAATs pharmacological blockers or selective EAAC1 antisense oligonucleotides were used. Since EAATs are Na+-dependent proteins, this raised the possibility that other transporters regulating ion gradients across mitochondrial membrane were required for glutamate response. We describe colocalization, mutual activity dependency, physical interaction between EAAC1 and the sodium/calcium exchanger 1 (NCX1) both in neuronal and glial mitochondria, and that NCX1 is an essential modulator of this glutamate transporter. Only NCX1 activity is crucial for such glutamate-stimulated ATP synthesis, as demonstrated by pharmacological blockade and selective knock-down with antisense oligonucleotides. The EAAC1/NCX1-dependent mitochondrial response to glutamate may be a general and alternative mechanism whereby this neurotransmitter sustains ATP production, since we have documented such metabolic response also in mitochondria isolated from heart. The data reported here disclose a new physiological role for mitochondrial NCX1 as the key player in glutamate-induced energy production

Genetic Structure of Modern Durum Wheat Cultivars and Mediterranean Landraces Matches with Their Agronomic Performance

A collection of 172 durum wheat landraces from 21 Mediterranean countries and 20 modern cultivars were phenotyped in 6 environments for 14 traits including phenology, biomass, yield and yield components. The genetic structure of the collection was ascertained with 44 simple sequence repeat markers that identified 448 alleles, 226 of them with a frequency lower than 5%, and 10 alleles per locus on average. In the modern cultivars all the alleles were fixed in 59% of the markers. Total genetic diversity was HT = 0.7080 and the genetic differentiation value was GST = 0.1730. STRUCTURE software allocated 90.1% of the accessions in five subpopulations, one including all modern cultivars, and the four containing landrace related to their geographic origin: eastern Mediterranean, eastern Balkans and Turkey, western Balkans and Egypt, and western Mediterranean. Mean yield of subpopulations ranged from 2.6 t ha-1 for the western Balkan and Egyptian landraces to 4.0 t ha-1 for modern cultivars, with the remaining three subpopulations showing similar values of 3.1 t ha-1. Modern cultivars had the highest number of grains m-2 and harvest index, and the shortest cycle length. The diversity was lowest in modern cultivars (HT = 0.4835) and highest in landraces from the western Balkans and Egypt (HT = 0.6979). Genetic diversity and AMOVA indicated that variability between subpopulations was much lower (17%) than variability within them (83%), though all subpopulations had similar biomass values in all growth stages. A dendrogram based on simple sequence repeat data matched with the clusters obtained by STRUCTURE, improving this classification for some accessions that have a large admixture. landraces included in the subpopulation from the eastern Balkans and Turkey were separated into two branches in the dendrogram drawn with phenotypic data, suggesting a different origin for the landraces collected in Serbia and Macedonia. The current study shows a reliable relationship between genetic and phenotypic population structures, and the connection of both with the geographic origin of the landraces.The research was funded by the Ministerio de Economía y competitividad project AGL-2006-09226-C02-01, and Dr. Jose Miguel Soriano is funded by Instituto Nacional de Investigación y Tecnología Agraria y Alimentaria (http://www.mineco.gob.es/)

Mitochondria-dependent signalling pathway are involved in the early process of radiation-induced bystander effects

Bystander effects induced by cytoplasmic irradiation have been reported recently. However, the mechanism(s) underlying, such as the functional role of mitochondria, is not clear. In the present study, we used either mtDNA-depleted (ρ0) AL or normal (ρ+) AL cells as irradiated donor cells and normal human skin fibroblasts as receptor cells in a series of medium transfer experiments to investigate the mitochondria-related signal process. Our results indicated that mtDNA-depleted cells or normal AL cells treated with mitochondrial respiratory chain function inhibitors had an attenuated γ-H2AX induction, which indicates that mitochondria play a functional role in bystander effects. Moreover, it was found that treatment of normal AL donor cells with specific inhibitors of NOS, or inhibitor of mitochondrial calcium uptake (ruthenium red) significantly decreased γ-H2AX induction and that radiation could stimulate cellular NO and O2•− production in irradiated ρ+ AL cells, but not in ρ0 AL cells. These observations, together with the findings that ruthenium red treatment significantly reduced the NO and O2•− levels in irradiated ρ+ AL cells, suggest that radiation-induced NO derived from mitochondria might be an intracellular bystander factor and calcium-dependent mitochondrial NOS might play an essential role in the process

Antiinflammatory Therapy with Canakinumab for Atherosclerotic Disease

Background: Experimental and clinical data suggest that reducing inflammation without affecting lipid levels may reduce the risk of cardiovascular disease. Yet, the inflammatory hypothesis of atherothrombosis has remained unproved. Methods: We conducted a randomized, double-blind trial of canakinumab, a therapeutic monoclonal antibody targeting interleukin-1β, involving 10,061 patients with previous myocardial infarction and a high-sensitivity C-reactive protein level of 2 mg or more per liter. The trial compared three doses of canakinumab (50 mg, 150 mg, and 300 mg, administered subcutaneously every 3 months) with placebo. The primary efficacy end point was nonfatal myocardial infarction, nonfatal stroke, or cardiovascular death. RESULTS: At 48 months, the median reduction from baseline in the high-sensitivity C-reactive protein level was 26 percentage points greater in the group that received the 50-mg dose of canakinumab, 37 percentage points greater in the 150-mg group, and 41 percentage points greater in the 300-mg group than in the placebo group. Canakinumab did not reduce lipid levels from baseline. At a median follow-up of 3.7 years, the incidence rate for the primary end point was 4.50 events per 100 person-years in the placebo group, 4.11 events per 100 person-years in the 50-mg group, 3.86 events per 100 person-years in the 150-mg group, and 3.90 events per 100 person-years in the 300-mg group. The hazard ratios as compared with placebo were as follows: in the 50-mg group, 0.93 (95% confidence interval [CI], 0.80 to 1.07; P = 0.30); in the 150-mg group, 0.85 (95% CI, 0.74 to 0.98; P = 0.021); and in the 300-mg group, 0.86 (95% CI, 0.75 to 0.99; P = 0.031). The 150-mg dose, but not the other doses, met the prespecified multiplicity-adjusted threshold for statistical significance for the primary end point and the secondary end point that additionally included hospitalization for unstable angina that led to urgent revascularization (hazard ratio vs. placebo, 0.83; 95% CI, 0.73 to 0.95; P = 0.005). Canakinumab was associated with a higher incidence of fatal infection than was placebo. There was no significant difference in all-cause mortality (hazard ratio for all canakinumab doses vs. placebo, 0.94; 95% CI, 0.83 to 1.06; P = 0.31). Conclusions: Antiinflammatory therapy targeting the interleukin-1β innate immunity pathway with canakinumab at a dose of 150 mg every 3 months led to a significantly lower rate of recurrent cardiovascular events than placebo, independent of lipid-level lowering. (Funded by Novartis; CANTOS ClinicalTrials.gov number, NCT01327846.

Czech Adolescents’ Face-to-Face Meetings With People from the Internet: The Role of Adolescents' Motives and Expectations

Research of face-to-face meetings between adolescents and people met online stands on untested assumptions that these meetings are uniform, and adolescents attend them to expand their social circle. It is also unclear what makes such meetings pleasant or unpleasant. This study examined meetings of 611 Czech adolescents (age 11–16, M(age) = 14.04, SD = 1.67, 47.1% female). Face-to-face meetings attended with friendly, romantic, or instrumental motives differed from each other, emphasizing the need to investigate them separately. Pleasantness of meetings is closely related to disconfirmation of adolescents’ expectations. Unmet expectations related to unpleasant meetings, exceeded expectations to pleasant ones. While present findings uphold existing theories (e.g., social compensation), they also call for new theoretical perspectives for this common adolescents’ activity