Neutron and cobalt-60 gamma irradiation produce similar changes in DNA supercoiling

The effects of 60Co gamma-ray and d(20 MeV)Be neutron irradiation on DNA supercoiling have been studied using a nucleoid rewinding technique. Irradiation of viable CHO AA8 cells on ice with from 4 to 25 Gy of either radiation produced a similar resistance to rewinding of nuclear supercoils after treatment with ethidium bromide. The restitution from the effects of 12 Gy of either radiation was also similar, leaving no detectable residual damage. The discrepancy between these data and the reduced ability of neutrons to produce DNA breaks, as defined by the alkaline elution assay, is explained by the discontinuous deposition of dose associated with neutron irradiation. It is suggested from a microdosimetric analysis that the neutron radiation interacts with DNA at sites on average 5-10 times further apart than the interactions with gamma rays. The long DNA sequences which results after neutron irradiation are consequently eluted inefficiently during alkaline elution, giving a reported RBE of approximately 0.3. Restrictions in the rewinding of individual supercoils are not dependent on the interionization distance and thus give rise to an RBE of approximately 1. Furthermore, the complete removal of DNA damage, as measured by this technique, supports the hypothesis that neutron toxicity is associated with incorrect, not incomplete, rejoining of the DNA molecule.     

Myricetin and quercetin are naturally occurring co-substrates of cyclooxygenases in vivo

Bioflavonoids are ubiquitously present in the plant kingdom, and some of them are presently being sold as healthy dietary supplements around the world. Recently, it was shown that some of the dietary polyphenols were strong stimulators of the catalytic activity of cyclooxygenase I and II, resulting in increased formation of certain prostaglandin (PG) products in vitro and also in intact cells in culture. In the present study, we investigated the effect of two representative dietary compounds, quercetin and myricetin, on plasma and tissue levels of several PG products in normal Sprague-Dawley rats. We found that these two dietary bioflavonoids could strongly stimulate the formation of PG products in vivo in a time-dependent manner, and the stimulatory effect of these two bioflavonoids was dose-dependent with a unique biphasic pattern. At lower doses (<0.3 mg/kg b.w.), they strongly stimulated the formation of PGE₂, but at higher doses (>0.3 mg/kg b.w.), there was a dose-dependent reduction of the stimulatory effect. These results provide support for the hypothesis that some of the bioflavonoids are naturally occurring physiological co-substrates for the cyclooxygenases in vivo.     

Systemic metabolic reactions are obtained by singular value decomposition of genome-scale stoichiometric matrices

Genome-scale metabolic networks can be reconstructed. The systemic biochemical properties of these networks can now be studied. Here, genome-scale reconstructed metabolic networks were analysed using singular value decomposition (SVD). All the individual biochemical conversions contained in a reconstructed metabolic network are described by a stoichiometric matrix (S). SVD of S led to the definition of the underlying modes that characterize the overall biochemical conversions that take place in a network and rank-ordered their importance. The modes were shown to correspond to systemic biochemical reactions and they could be used to identify the groups and clusters of individual biochemical reactions that drive them. Comparative analysis of the Escherichia coli, Haemophilus influenzae, and Helicobacter pylori genome-scale metabolic networks showed that the four dominant modes in all three networks correspond to: (1) the conversion of ATP to ADP, (2) redox metabolism of NADP, (3) proton-motive force, and (4) inorganic phosphate metabolism. The sets of individual metabolic reactions deriving these systemic conversions, however, differed among the three organisms. Thus, we can now define systemic metabolic reactions, or eigen-reactions, for the study of systems biology of metabolism and have a basis for comparing the overall properties of genome-specific metabolic networks.     

Redox reactions related to indoleamine 2,3-dioxygenase and tryptophan metabolism along the kynurenine pathway

The heme enzyme indoleamine 2,3-dioxygenase (IDO) oxidizes the pyrrole moiety of L-tryptophan (Trp) and other indoleamines and represents the initial and rate-limiting enzyme of the kynurenine (Kyn) pathway. IDO is a unique enzyme in that it can utilize superoxide anion radical (O2*- ) as both a substrate and a co-factor. The latter role is due to the ability of O2*- to reduce inactive ferric-IDO to the active ferrous form. Nitrogen monoxide (*NO) and H2O2 inhibit the dioxygenase and various inter-relationships between the nitric oxide synthase- and IDO-initiated amino acid degradative pathways exist. Induction of IDO and metabolism of Trp along the Kyn pathway is implicated in a variety of physiological and pathophysiological processes, including anti-microbial and anti-tumor defense, neuropathology, immunoregulation and antioxidant activity. Antioxidant activity may arise from O2*- scavenging by IDO and formation of the potent radical scavengers and Kyn pathway metabolites, 3-hydroxyanthranilic acid and 3-hydroxykynurenine. Under certain conditions, these aminophenols and other Kyn pathway metabolites may exhibit pro-oxidant activities. This article reviews findings indicating that redox reactions are involved in the regulation of IDO and Trp metabolism along the Kyn pathway and also participate in the biological activities exhibited by Kyn pathway metabolites.     

Rat embryo cells immortalized with transfected oncogenes are transformed by gamma irradiation.

Cesium-137 gamma rays were used to transform rat embryo cells (REC) which were first transfected with activated c-myc or c-Ha-ras oncogenes to produce immortal cell lines (REC:myc and REC:ras). When exposed to 6 Gy of 137Cs gamma rays, some cells became morphologically transformed with focus formation frequencies of approximately 3 x 10(-4) for REC:myc and approximately 1 x 10(-4) for REC:ras, respectively. Cells isolated from foci of gamma-ray-transformed REC:myc (REC:myc:gamma) formed anchorage-independent colonies and were tumorigenic in nude mice, but foci from gamma-ray-transformed REC:ras (REC:ras:gamma) did not exhibit either of these criteria of transformation. Similar to the results with gamma irradiation, we observed a sequence-dependent phenomenon when myc and ras were transfected into REC, one at a time. REC immortalized by ras transfection were not converted to a tumorigenic phenotype by secondary transfection with myc, but REC transfected with myc were very susceptible to transformation by subsequent ras transfection. This suggests that myc-immortalized cells are more permissive to transformation via secondary treatments. In sequentially transfected REC, myc expression was high whether it was transfected first or second, whereas ras expression was highest when the ras gene was transfected secondarily into myc-containing REC. Molecular analysis of REC:ras:gamma transformants showed no alterations in structure of the transfected ras or of the endogenous ras, myc, p53, or fos genes. The expression of ras and p53 was increased in some isolates of REC:ras:gamma, but myc and fos expression were not affected. Similarly, REC:myc:gamma transformants did not demonstrate rearrangement or amplification of the transfected or the endogenous myc genes, or of the potentially cooperating Ha-, Ki-, or N-ras genes. Northern hybridization analysis revealed increased expression of N-ras in two isolates, REC:myc:gamma 33 and gamma 41, but no alterations in the expression of myc, raf, Ha-ras, or Ki-ras genes in any REC:myc transformant. DNA from several transformed REC:myc:gamma cell lines induced focus formation in recipient C3H 10T1/2 and NIH 3T3 cells. The NIH 3T3 foci tested positive when hybridized to a probe for rat repetitive DNA. A detailed analysis of the NIH 3T3 transformants generated from REC:myc:gamma 33 and gamma 41 DNA failed to detect Ha-ras, Ki-ras, raf, neu, trk, abl, fms, or src oncogenes of rat origin.(ABSTRACT TRUNCATED AT 400 WORDS)     

Influence of whole-body gamma irradiation upon arachidonic acid metabolism in rat platelets

The effects of whole-body gamma irradiation (8.4 Gy) were studied on arachidonic acid (AA) metabolism in rats' blood platelets, from day D + 1 to day D + 10 after irradiation. AA conversion into thromboxane B2 (TxB2) increased at D + 1 and then gradually decreased to very low values from D + 7 to D + 10. This decrease in the conversion of exogenous AA into TxB2 was due to a lower AA incorporation into platelets and not to a decrease of cyclooxygenase and thromboxane-synthetase activities. AA incorporation into membrane phospholipids of blood platelets was much more decreased than AA incorporation into whole platelets; moreover, the lipid composition of the platelet membranes was markedly modified after irradiation, which must have resulted in structural and functional changes in these membranes; from these effects of whole-body gamma irradiation on platelets, the latter's membranes appeared as a major site of in vivo radiation damage in these cells.     

Adaptation to irradiation in vivo

The data about the increasing of radioresistance of cells and organism after the acute, chronic and prolonged irradiation in vivo were presented. The possible mechanisms of adaptation to irradiation connected with selection, stimulation of proliferative activity, forming of protecting system (protected proteins, antioxidant enzymes e.a.), activation of DNA repair and cAMP-, and Ca(2+)-dependent phosphorylation systems were observed. The conclusion about complex mechanism of radiation adaptation was drawn.     

Inactivation of Coxiella burnetii by gamma irradiation

The gamma radiation inactivation kinetics for Coxiella burnetii at -79 degrees C were exponential. The radiation dose needed to reduce the number of infective C. burnetii by 90% varied from 0.64 to 1.2 kGy depending on the phase of the micro-organism, purity of the culture and composition of suspending menstruum. The viability of preparations containing 10(11) C. burnetii ml-1 was completely abolished by 10 kGy without diminishing antigenicity or ability to elicit a protective immune response in vaccinated mice. Immunocytochemical examinations using monoclonal antibodies and electron microscopy demonstrated that radiation doses of 20 kGy did not alter cell-wall morphology or cell-surface antigenic epitopes.     

Redox balances in the metabolism of sugars by yeasts

Abstract The central role of the redox couples NAD⁺/NADH and NADP⁺/NADPH in the metabolism of sugars by yeasts is discussed in relation to energy metabolism and product formation. Besides their physical compartmentation in cytosol and mitochondria, the two coenzyme systems are separated by chemical compartmentation as a consequence of the absence of transhydrogenase activity. This has considerable consequences for the redox balances of both coenzyme systems and hence for sugar metabolism in yeasts.
As examples, the competition between respiration and fermentation of glucose, the Crabtree effect, the Custers effect, adaptation to anaerobiosis, the activities of the hexose monophosphate pathway, and the fermentation of xylose in yeast are discussed.     

Redox reactions in hydrocortisone transformation by Arthrobacter globiformis cells

Hydrocortisone and prednisolone transformation by Arthrobacter globiformis cells in aerobic and anaerobic conditions was studied. 3-Ketosteroid-1-en-dehydrogenase activity was shown to be the major factor regulating the direction of transformation. When it is high (aerobic conditions), the end products of hydrocortisone transformation are prednisolone or its 20 beta-hydroxy derivative. The latter is produced via 1-en-dehydrogenation, which is not a limiting stage of the process. Low 3-ketosteroid-1-en-dehydrogenase activity (in the presence of cyanide) or its complete inhibition (strictly anaerobic conditions) result in the direct reduction of 20-keto group of hydrocortisone.     

Blood-brain barrier permeability after gamma whole-body irradiation: an in vivo microdialysis study

The effects of total-body irradiation on the permeability of rat striatal blood-brain barrier (BBB) to [3H]alpha-aminoisobutyric acid (AIBA) and [14C]sucrose were investigated using the microdialysis technique. Seven days, 3 and 6 weeks, and 3, 5, and 8 months after gamma exposure at a dose of 4.5 Gy, no modification of the permeability to both [3H]AIBA and [14C]sucrose was observed. But, in the course of the initial syndrome, we observed a significant but transient increase in the BBB permeability to the two markers between 3 and 17 h after exposure. A secondary transient "opening" of the BBB to [14C]sucrose was noticed about 28 h following irradiation without the corresponding increase in BBB permeability to [3H]AIBA. On the contrary, the transport of [3H]AIBA through the BBB was decreased between 33 and 47 h postradiation. In conclusion, our experiments showed early modifications of BBB permeability after a moderate-dose whole-body exposure. Confirmation of these results with other tracers, in another experimental model or in humans, would have clinical applications for designing appropriate pharmacotherapy in radiotherapy and treatment of accidental overexposure.     

Measurements of nucleic bases released after gamma irradiation of DNA in solution in air.

The release of unaltered nucleic bases from gamma-irradiated DNA in a dilute buffered aqueous solution was studied in both salmon sperm and superhelical viral DNA. Analyses of freed bases were made by high-performance liquid chromatography. An elution protocol was developed for maximum separation of the four nucleic bases and nucleosides with a sensitivity of 10-20 pmol of nucleic base. It was found that: (i) both prompt and delayed release of bases postirradiation occur in both types of DNA; (ii) these yields (G-values) were measured to be 10-15 times higher for the salmon sperm DNA in comparison to the SV40 DNA; (iii) the A-T/G-C ratio in the DNA was not reflected in the ratios of the released base; and (iv) based on measurements made by us of DNA strand breaks in SV40 DNA (unpublished results), less than half of all breaks result in the release of an undamaged base.     

In vivo nuclear magnetic resonance studies of the metabolism of methanol and pyruvate by Methanosarcina barkeri

Abstract The metabolism of methanol and pyruvate by cells of Methanosarcina barkeri was probed in vivo by NMR taking advantage of the non-invasive characteristics of this technique. Upon administration of substrates, the kinetics of substrate consumption, the product formation and the energetic state of the cells was monitored using carbon-13, phosphorus-31 or proton NMR. The effects of several inhibitors and uncouplers were investigated. Cells supplied with pyruvate developed considerable levels of nucleotide triphosphate; methane production was monitored, as well as CO₂ and H₂ formation. Most of the pyruvate was utilized for the synthesis of valine or intermediates of the valine pathway. The origin of the carbon atom in methane was elucidated using ¹³C-labelled pyruvate.