Protective role of pinacidil against adrenaline-induced myocardium injury in guinea pig liver mitochondria

We investigated the role of the ATP-sensitive potassium channel opener pinacidil and blocker glibenclamide on guinea pig liver mitochondrial function, and a possible significance of pinacidil in the pharmacological treatment during myocardium dystrophy. First, a series of experiments was performed to determine the effect of pinacidil and glibenclamide on mitochondrial oxygen consumption. We found that pinacidil increased the rate of mitochondrial respiration for FAD-generated substrate (succinate oxidation), but was most effective for α-ketoglutarate oxidation with enhancement of respiratory control ratio. Oxidation of FAD-generated substrate inhibited efficiency of phosphorylation for α-ketoglutarate oxidation in pinacidil-treated animals. Glibenclamide decreased the rate of respiration with the lowest value of efficiency of phosphorylation, especially for α-ketoglutarate oxidation. A second series of experiments was performed to determine the effects of pinacidil and glibenclamide on oxidative phosphorylation during adrenaline-induced myocardium dystrophy. The increase in respiratory control ratio and efficiency of phosphorylation for α-ketoglutarate oxidation was greater than for succinate oxidation in mitochondria of pinacidil-pretreated animals during myocardium dystrophy. Inhibitory analysis with malonate suggested that endogenous succinate increased oxidation of NADH-generated substrates in mitochondria. Pinacidil is mainly involved in the adrenaline-induced alterations of mitochondrial function due to elevation of phosphorylation efficiency for α-ketoglutarate oxidation and a decreased level of lipid peroxidation.     

Caenorhabditis elegans integrates food and reproductive signals in lifespan determination

Dietary restriction extends lifespan and inhibits reproduction in many species. In Caenorhabditis elegans, inhibiting reproduction by germline removal extends lifespan. Therefore, we asked whether the effect of dietary restriction on lifespan might proceed via changes in the activity of the germline. We found that dietary restriction could increase the lifespan of animals lacking the entire reproductive system. Thus, dietary restriction can extend lifespan independently of any reproductive input. However, dietary restriction produced little or no increase in the long lifespan of animals that lack germ cells. Thus, germline removal and dietary restriction may potentially activate lifespan-extending pathways that ultimately converge on the same downstream longevity mechanisms. In well-fed animals, the somatic reproductive tissues are generally completely required for germline removal to extend lifespan. We found that this was not the case in animals subjected to dietary restriction. In addition, in these animals, loss of the germline could either further lengthen lifespan or shorten lifespan, depending on the genetic background. Thus, nutrient levels play an important role in determining how the reproductive system influences longevity.     

Frequency of hemochromatosis gene (HFE) mutations in Russian healthy women and patients with estrogen-dependent cancers

Possible association between the C282Y and H63D mutations in the HFE gene and estrogen-dependent cancer risk was assessed. Genotyping was performed using PCR amplification followed by digestion of products with specific restrictases. In a population of 260 healthy women (permanent residents of the southwest European Russia), mutant allele frequencies at the C282Y and H63D sites were evaluated as 3.3 and 16.3%, respectively. In patients with breast, ovarian, and endometrial cancer, C282Y frequencies were also low (1.0, 1.3, and 3.8%, respectively), and no cancer risk associated with the C282Y mutation was found. Odds ratios for breast cancer risk associated with the H63D mutation increased significantly with age: 0.5 in women below 48 years old, 1.0 in a range of 48-57 years, and 4.4 in older women (P(trend)=0.002). The latter value was statistically significant (95% CI, 1.4-14.1), indicating that women bearing the H63D mutation may be at an increased breast cancer risk at an age above 57 years. Preliminary results obtained in patients with two other estrogen-dependent malignancies revealed the same tendency to OR increase with age in ovarian cancer patients (P(trend)=0.008), but no age-related OR differences in endometrial cancer patients.     

DNA cleavage activity of VO(acac)2 and derivatives

The DNA cleavage activity of several β-diketonate vanadyl complexes is examined. Vanadyl acetylacetonate, V^IVO(acac)₂, 1, shows a remarkable activity in degrading plasmid DNA in the absence of any activating agents, air and photoirradiation. The cleaving activity of several related complexes V^IVO(hd)₂ (2, Hhd = 3,5-heptanedione), V^IVO(acac-NH₂)₂ (3, Hacac-NH₂ = acetoacetamide) and V^IVO(acac-NMe₂)₂ (4, Hacac-NMe₂ = N,N-dimethylacetoacetamide) is also evaluated. It is shown that 2 exhibits an activity similar to 1, while 3 and 4 are much less efficient cleaving agents. The different activity of the complexes is related to their stability towards hydrolysis in aqueous solution, which follows the order 1∼2 ? 3∼4. The nature of the pH buffer was also found to be determinant in the nuclease activity of 1 and 2. In a phosphate buffered medium DNA cleavage by these agents is much more efficient than in tris, hepes, mes or mops buffers. The reaction seems to take place through a mixed mechanism, involving the formation of reactive oxygen species (ROS), namely OH radicals, and possibly also direct cleavage at phosphodiester linkages induced by the vanadium complexes.     

In Vitro Effects of Cholesterol β-d-Glucoside, Cholesterol and Cycad Phytosterol Glucosides on Respiration and Reactive Oxygen Species Generation in Brain Mitochondria

The cluster of neurodegenerative disorders in the western Pacific termed amyotrophic lateral sclerosis-parkinsonism dementia complex (ALS-PDC) has been repeatedly linked to the use of seeds of various species of cycad. Identification and chemical synthesis of the most toxic compounds in the washed cycad seeds, a variant phytosteryl glucosides, and even more toxic cholesterol β-D-glucoside (CG), which is produced by the human parasite Helicobacter pylori, provide a possibility to study in vitro the mechanisms of toxicity of these compounds. We studied in detail the effects of CG on the respiratory activities and generation of reactive oxygen species (ROS) by nonsynaptic brain and heart mitochondria oxidizing various substrates. The stimulatory effects of CG on respiration and ROS generation showed strong substrate dependence, suggesting involvement of succinate dehydrogenase (complex II). Maximal effects on ROS production were observed with 1 μmol CG/1 mg mitochondria. At this concentration the cycad toxins β-sitosterol-β-D-glucoside and stigmasterol-β-D-glucoside had effects on respiration and ROS production similar to CG. However, poor solubility precluded full concentration analysis of these toxins. Cholesterol, stigmasterol and β-sitosterol had no effect on mitochondrial functions studied at concentrations up to 100 μmol/mg protein. Our results suggest that CG may influence mitochondrial functions through changes in the packing of the bulk membrane lipids, as was shown earlier by Deliconstantinos et al. (Biochem Cell Biol 67:16-24, 1989). The neurotoxic effects of phytosteryl glucosides and CG may be associated with increased oxidative damage of neurons. Unlike heart mitochondria, in activated neurons mitochondria specifically increase ROS production associated with succinate oxidation (Panov et al., J Biol Chem 284:14448-14456, 2009).     

1-Bromo-3-(1',1'-dimethylalkyl)-1-deoxy-Δ(8)-tetrahydrocannabinols: New selective ligands for the cannabinoid CB(2) receptor

Δ(8)-Tetrahydrocannabinol (26), 3-(1',1'-dimethylbutyl)- (12), 3-(1',1'-dimethylpentyl)- (13), 3-(1',1'-dimethylhexyl)- (14) and 3-(1',1'-dimethylheptyl)-Δ(8)-tetrahydrocannabinol (15) have been converted into the corresponding 1-bromo-1-deoxy-Δ(8)-tetrahydrocannabinols (25, 8-11). This was accomplished using a protocol developed in our laboratory in which the trifluoromethanesulfonate of a phenol undergoes palladium mediated coupling with pinacolborane. Reaction of this dioxaborolane with aqueous-methanolic copper(II) bromide provides the aryl bromide. The affinities of these bromo cannabinoids for the cannabinoid CB(1) and CB(2) receptors were determined. All of these compounds showed selectivity for the CB(2) receptor and one of them, 1-bromo-1-deoxy-3-(1',1'-dimethylhexyl)-Δ(8)-tetrahydrocannabinol (10), exhibits 52-fold selectivity for this receptor with good (28nM) affinity.