Comparison of DNA Complexation with Antitumor Agent cis-DDP and Binuclear Bivalent Platinum Compound Containing Pyrazine

Conformational properties of DNA molecule upon its complexation with binuclear compounds of bivalent platinum in the cis configuration containing pyrazine ligand were studied by circular dichroism, viscometry, and dynamic birefringence. Comparison with an active antitumor agent cis-diamminedichloroplatinum (cis-DDP) was made. Experimental data indicate that interaction of these compounds with DNA results in the formation of a coordination bond of platinum with nucleic bases. The structure of the complex depends on the ratio of platinum and DNA concentrations in the initial solution. The study of DNA protonation in complex with the binuclear coordination compound showed that the binding of platinum with DNA bases involves the N7 atom of guanine. Competition was observed between the studied compound and cis-DDP for the binding site on DNA. The macromolecule binds stronger with the binuclear platinum compound than with cis-DDP.     

In Vivo and in Vitro binding of platinum to metallothionein

The in vivo binding of platinum to metallothionein (MT) has been observed in rat tissues following injections of the cis and trans isomers of DDP (dichlorodiammine-platinum(II)). Platinum in either cis-DDP or trans-DDP does not directly induce MT; platinum-MT is produced by the replacement of previously bound zinc in the protein. The binding of Pt(II) to MT depends on the availability of SH groups in MT. Preinjection with CdCl₂ significantly enhances the association of Pt(II) with MT fractions compared to the degree of association resulting from injections with either cis-DDP or trans-DDP without CdCl₂ pretreatment. In vitro experiments in which tissue extracts including a known (Cd,Zn)-MT were incubated with either cis-DDP or trans-DDP show that these isomers differ with respect to the transfer of Pt to MT; the equilibrium in both cases was reached when approximately 40% of the available Pt is bound to MT but with this equilibrium value attained in 2 h in the case of trans-DDP and only after 72 h in the case of cis-DDP. Pt-MTs were also formed by a series of incubation steps in which a native MT was used to prepare the apoprotein which was subsequently incubated with either cis-DDP or trans-DDP. Spectrophotometry established that a shoulder occurs at 285 nm for the Pt-MTs resulting from the incubation with either isomer. A competitive double-antibody radioimmunoassay for MT demonstrated that these Pt-MTs had complete cross-reactivity with a native (Cd,Zn)-MT. Gel filtration of tissue extracts after either in vivo or in vitro treatment with DDP showed that Pt was bound to a molecular species with properties characteristic of MT. These results were verified by atomic absorption spectrophotometry and polyacrylamide gel electrophoresis assays.     

Influence of cis-dichlorodiamineplatinum on glioma cell morphology and cell cycle kinetics in tissue culture

Summary C6 glioma cells (CCL 107) were cultured for three days and then treated withcis-dichlorodiamineplatinum (cis-DDP) at doses of 0.2–10 µg/ml medium. Changes in DNA synthesis and DNA content, as well as morphology of cells and chromatin distribution, were examined from the first post-treatment day onwards. The number of cells labelled with [³H]thymidine, detected autoradiographically, decreased after treatment with 0.2–10 µg/ml by approximately one half on post-treatment day 1 and diminished further by the third day after treatment. The labelled cells were entirely absent only after treatment with 10µg/ml, 7 days post-treatment. Mitoses decreased from 1.4–0.6% by post-treatment day 1 and completely disappeared by day 3 (1 µg/ml). Feulgen cytophotometry and propidium iodide cytofluorimetry revealed accumulation of cells in the S-phase, especially the latter part (0.5 and 1.0 µg/ml, post-treatment day 1) and subsequently also in G2 phase (post-treatment day 3). Incomplete cyto- and karyokinesis in some cycling cells was indicated by an increased number of binucleate cells and nuclei of higher ploidy classes. Labelled cells with intermediate DNA values were, on average, labelled less intensively, as was revealed by simultaneous measurements of DNA content and [³H]thymidine incorporation. Some cells displayed reduction in grain density over heterochromatin clumps. This would be in agreement with the late S-phase block of DNA replication. After post-treatment day 3 the density of cells in cultures was substantially lower. This was due to slowed transversing through the cell cycle and cell death occurring after post-treatment day 1 with higher doses or after day 2 with lower doses (up to 1 µg/ml). The size of the nuclei of surviving cells enlarged initially (post-treatment day 1) and later (day 7) giant cells with long, branched fibres similar to those of reactive astrocytes occurred. Texture analysis of Feulgen-stained nuclei revealed that the chromatin of cells treated withcis-DDP became less evenly distributed. This might be due either to the direct influence ofcis-DDP on the DNA molecule, or mediated by changes in cytoskeleton and cAMP levels described earlier.     

The inhibitory effect of sodium selenite on N-nitrosodiethylamine-induced and phenobarbital promoted liver tumourigenesis in rats based on the modulation of polyamine levels

In the present study, we have evaluated the effects of dietary selenite (Se) on polyamine levels and its influence on N-nitrosodiethylamine (DEN) initiated and Phenobarbital (PB) promoted in rat liver carcinogenesis. Dietary selenite at a concentration of 4 ppm (through drinking water) was administered in rats either before initiation (4 weeks), or during promotion (16 weeks) and entire experimental period (20 weeks). Male Wistar strain of albino rats was treated with single intra peritoneal dose of DEN (200 mg kg⁻¹ body weight), after 2 weeks the carcinogenic effect was promoted by PB (0.05%; through diet). Alpha fetoprotein (AFP) was investigated after the 20th-week of experimental period. Selenite-treated animals markedly reduced the AFP during the time of pre-selenite [before initiation (4 weeks)] and entire experimental period (20 weeks), administration rather than the promotion period. This infers that anticancer property of selenite depends on the stage of carcinogenesis, rather than duration of treatment. Evaluation of polyamine levels in hepatoma and surrounding liver tissue showed significant difference in the selenite-treated groups compared with pair-fed control groups. Furthermore, histopathological examination showing remarkable difference between control and treated groups. These results demonstrate that selenite can modulate the development of DEN-induced and PB-promoted rat liver carcinogenesis through a polyamine-dependent mechanism. (Mol Cell Biochem xxx: 165–172, 2005)     

Removal and assessment of toxicity of Cu and Fe toAnabaena doliolum andChlorella vulgaris using free and immobilized cells

Alginate-immobilized and free cells ofAnabaena doliolum andChlorella vulgaris were compared for their use in the removal and toxicity bioassays of Cu and Fe. A decrease in toxicity with regard to growth and uptake of NO ₃ ^– and NH ₄ ⁺ was noticed following immobilization of both the organisms. In contrast, immobilized cells had higher uptake rates of Cu and Fe suggesting that immobilization offers protection against metal toxicity. Compared with free cells, the immobilized cells showed greater efficiency for metal removal, even over three repeated cycles, though with a gradual decrease in efficiency in the second and third cycles. This reduction in removal efficiency was, however, more pronounced for Fe withA. doliolum and for Cu withC. vulgaris. The ease in harvesting and potential for repeated use makes the immobilized cells good tools for scavenging heavy metals from metal-contaminated environments.     

不同亚硒酸钠浓度对党参生长、生理特性和品质的影响

为综合了解党参的施硒效应，该研究基于水培试验探究不同亚硒酸钠浓度处理条件下党参幼苗的硒积累、生长、生理和品质的变化规律。结果表明：(1)适宜硒浓度(0.2 mg·L^-1)可以促进党参的叶片面积、株高、生物量的增长，而高硒浓度(10 mg·L^-1)则抑制党参生长。(2)增加亚硒酸钠浓度和硒暴露时间均可提高党参幼苗在根和叶中的硒含量，不同器官硒积累表现为根>叶>茎。(3)适宜硒浓度(0.2 mg·L^-1)处理可提高光合色素含量和根系活力，并减少丙二醛、脯氨酸和过氧化氢的积累，而高硒浓度(10 mg·L^-1)则与之相反。(4)适宜硒浓度(0.2 mg·L^-1)有利于党参炔苷、多糖、可溶性蛋白的积累，而高硒浓度(10 mg·L^-1)可以对多糖和可溶性蛋白的积累产生不利影响。综上认为，亚硒酸钠对党参具有双重效应，施加适量浓度对党参的生长、生理和品质有益，并以0.2 mg·L^-1硒浓度处理的效果最佳。该研究结果有助于了解亚硒酸钠浓度对...     

The role of root-associatedKlebsiella pneumoniae in the nitrogen nutrition ofPoa pratensis andTriticum aestivum as estimated by the method of15N isotope dilution

Summary The technique of¹⁵N isotope dilution was used to verify that nitrogen was fixed and transferred to the plant byKlebsiella pneumoniae strain Pp in association withPoa pratensis orTriticum aestivum. Surface sterilized, sprouting seeds were inoculated withK. pneumoniae and grown in sand in modified Leonard jars. Potassium nitrate enriched with¹⁵N was used to provide N concentrations ranging from 10–40 mg Nl^–1 nutrient solution. After 10–18 weeks the shoots and roots were analyzed separately for dry matter, N content, total N, and atom %¹⁵N excess. The acetylene reduction technique was used to test for the presence of N₂-fixing organisms on the roots. The data from¹⁵N isotope dilution demonstrated that up to 33.8% of N in the shoots ofP. pratensis and 15.9% in those ofT. aestivum were derived from associative N₂ fixation byK. pneumoniae. In most experiments the dry matter yield, N content, and total N yield of the shoots ofP. pratensis were increased byK. pneumoniae inoculation, whereas inoculation had no significant effect on the dry matter yield, N content or total N of the shoots ofT. aestivum.     

Subcellular distribution of selenium during uptake and its influence on mitochondrial oxidations in germinatingVigna radiata L

The metabolic significance of Se in plants is not well documented, though the presence of many selenoenzymes in bacteria and the essentiality of Se in higher animals is established. Since germination is an active process in plant growth and metabolism, the effect of Se was investigated in germinatingVigna radiata L, a nonaccumulating Sedeficient legume. Growth and protein were enhanced in seedlings supplemented with selenium (Se) as sodium selenite in the medium up to 1 μg/mL. The pattern of uptake of⁷⁵Se in the differentiating tissues and the subcellular distribution were investigated. The percentage of incorporation of⁷⁵Se was greater in the mitochondria at the lowest level (0.5 μg/mL) of Se supplementation compared to higher levels of Se exposure. Proteins precipitated from the postmitochondrial supernatant fractions, when separated by means of polyacrylamide gel electrophoresis (PAGE), indicated a major selenoprotein in the seedlings germinated at 2.0 μg/mL Se. In seedlings grown with supplemented Se, enhanced respiratory control ratio and succinate dehydrogenase activity were observed in the mitochondria of tissues, indicative of a role for Se in mitochondrial membrane functions.     

Selenium in the anterior pituitary of the rat after a single injection of75Se sodium selenite

In order to investigate the selenite metabolism in the anterior pituitary and compare it with other endocrine organs, rats were injected intraperitoneally with⁷⁵Se sodium selenite (5 mg/kg). The rats were whole body counted shortly after injection and recounted just before sacrifice, which was performed 2, 24, 48 h, and 4, 10, 20, 30, 40, 60, and 80 d after injection. Besides the anterior pituitary, the selenium content was also estimated in the thyroid gland, testis, adrenals, liver, kidney, and blood. The maximum selenium content was observed in all organs 2 h after injection, at which time the anterior pituitary contained 2.9 μg/g wet wt, compared to 13.5 μ/g wet wt in liver and .6 μg/mg wet wt in testis. The excretion of selenite from the anterior pituitary resembled that seen in most other organs investigated, i.e., an initial rapid excretion and a slower secondary phase resembling a first order reaction. Practically all selenium was excreted by 60 d after injection.     

Antagonistic effects of soil bacteria onFusarium oxysporum Schlecht f. sp.dianthi (Prill and Del.) Snyd. and Hans

Summary Fusarium oxysporum f. sp.dianthi, pathogenic on carnation plants is very sensitive toBacillus subtilis M51 inhibition.Fusarium oxysporum disease (fusariosis) is prevented for a period of two months after treatment of plants withBacillus subtilis M51. The persistence ofB. subtilis M51, marked for selenomycin resistance (MZ51) and inoculated on the roots of carnation cuttings was studied. Soil used was two types: naturally infested withFusarium oxysporum and free from this pathogen. Bacterial cells presence on the roots was detected by direct plating and the presence of the pathogen in the roots was investigated by histological assays. Evidence gathered by these procedures suggest that plant protection is dependent on the physical presence ofB. subtilis M51 cells on the roots.     

A new point of view on the taxonomy ofPottia starckeana agg. (Musci,Pottiaceae)

A new taxonomic treatment is proposed for thePottia starckeana species complex. The peristome development is not considered to be a useful feature to separate the taxa. On the basis of spore morphology only two species are accepted:P. starckeana, with spores wavy in outline, andP. davalliana, with variously-shaped and developed processes on the spores.Pottia starckeana var.brachyoda is reduced to synonymy withP. starckeana; P. conica andP. commutata are treated as synonyms ofP. davalliana. The speciesP. mutica, P. affinis, P. salina, P. microphylla, P. texana, andP. arizonica (included var.mucronulata) are considered taxa of doubtful affinity, as they have spore features intermediate between the two spore types established for the group. The identity ofP. appertii andP. recurvifolia has not been elucidated because the type material has been destroyed.     

A role for yeast glutaredoxin genes in selenite-mediated oxidative stress

Since the double Δgrx1Δgrx2 mutant is hypersensitive to selenite we decided to evaluate mechanisms underlying this phenomenon and establish the roles of other components of yeast glutaredoxin system, in particular glutaredoxin 5 in the selenite resistance. We found elevation in the intracellular and mitochondrial superoxide production in the Δgrx1Δgrx2 and Δgrx5 mutants after Se(IV) treatment. The last effect was more pronounced for cells lacking the mitochondrial Grx5 protein. We also recorded selenite-induced increase in the peroxide production in all strains tested. Nonfermentable carbon sources, glycerol and ethanol, augmented selenite toxicity. Hypo- and anoxia protected against the harmful effects of Se(VI). Augmentation of the intracellular levels of two endogenous antioxidants, erythroascorbic acid and glutathione confers resistance to selenite. We recorded a strain-unspecific, selenite-mediated decrease in the level of acid-soluble thiols. Collectively, our data demonstrate that hypersensitivity to the Δgrx1Δgrx2 and Δgrx5 disruptants to selenite is mediated by altered intracellular redox equilibrium.     

Production of the taste/odor-causing compound,trans-2,cis-6-nonadienal,within the Synurophyceae

Although several species of the Synurophyceae have been associated with taste and odor problems in potable water supplies, electron microscopic-based field studies linked problematic blooms only toSynura petersenii Korshikov. Eventually, the organic compoundtrans-2,cis-6-nonadienal was implicated to cause the associated cucumberlike odors. The objective of this study was to survey unialgal cultures of various Synurophycean species for the occurrence oftrans-2,cis-6-nonadienal. The compound was detected throughout a 24-day growth assay with aS. petersenii isolate, but was not detected in an identical assay withSynura sphagnicola (Korshikov) Korshikov. In separate 24-day cultures,trans-2,cis-6-nonadienal was detected in two isolates from theS. petersenii species complex, but was not detected in isolates of twoMallomonas or fourSynura taxa not from theS. petersenii complex. These results support the hypothesis that production oftrans-2,cis-6-nonadienal is unique to taxa within theS. petersenii complex. When contrast-enhancing optics and specific specimen preparation techniques are employed, light microscopy can be used to distinguish taxa in theS. petersenii complex from all other Synurophycean taxa. These methods are suggested as an efficient way to monitortrans-2,cis-6-nonadienal-producing taxa in potable water supplies.Author for correspondence     

Inoculated common beans are protected againstMacrophomina phaseolina byBurkholderia cepacia UPR 5C

Ashy stem blight (ASB), caused byMacrophomina phaseolina (Tassi) Goid., is a common severe disease affectingPhaseolus vulgaris in Puerto Rico and the Dominican Republic. The effect of inoculating seeds ofP. vulgaris withBurkholderia cepacia strain UPR 5C on the severity of ASB was studied under greenhouse conditions. Results of this study showed thatB. cepacia reduced the ASB disease severity by 71% and was compatible withRhizobium phaseoli CIAT 632.     

Protection of maize seedlings byFusarium moniliforme against infection byFusarium graminearum in the soil

The incidence ofFusarium moniliforme in surface-sterilized kernels in two commercial South African white maize cultivars was 64% and 6%, respectively. Heat treatment completely eliminated seedborneF. moniliforme from kernels of both cultivars. Heat treated, uncontaminated maize germlings were pre-inoculated with different isolates ofF. moniliforme and planted in steam-treated soil containing inoculum of different isolates ofF. graminearum Group 1 and Group 2. Seedling weights of germlings pre-inoculated with some isolates ofF. moniliforme were significantly higher than those of controls when exposed to some isolates ofF. graminearum in the soil. The protective effect of pre-inoculation withF. moniliforme was particularly evident in maize seedlings exposed to inoculum of an aggressive isolate ofF. graminearum Group 1. This is the first report of the protection of maize seedlings byF. moniliforme against infection byF. graminearum in the soil.     

Intergeneric hybridization and C-banding patterns inHordelymus (Triticeae,Poaceae)

Crosses ofHordelymus europaeus (2n = 4x = 28) with four genera in theTriticeae were attempted. Adult hybrids were obtained in combinations withHordeum bogdanii (2x),Hordeum depressum (4x), andSecale cereale (2x). The meiotic pairing was very low in the hybrids withH. bogdanii andSecale cereale (0.12 and 0.30 chiasmata/cell, respectively), whereas high pairing (9.90 chiasmata/cell) was found in hybrids withH. depressum due to autosyndetic pairing ofH. depressum chromosomes. The chromosome complement ofHordelymus europaeus comprised 16 metacentrics, 8 submetacentrics, and 4 SAT-chromosomes. The Giemsa C-banding patterns of the chromosomes were characterized by small to minute bands at no preferential positions. It is hypothesized thatHordelymus europaeus may genomically be closest related toTaeniatherum andPsathyrostachys spp.     

Selenium enhances the antimutagenic activity of probiotic bacterium Enterococcus faecium M-74

The antibacterial activity of the probiotic bacterium Enterococcus faecium M-74 was assessed on De Man–Rogosa–Sharpe (MRS), Todd–Hewitt (T–H), M17 (M-17) and brain heart infusion (BHI) media with sodium selenite pentahydrate (+Se) and without sodium selenite pentahydrate (–Se) under aerobic or anaerobic conditions against nine bacterial pathogens. The highest antibacterial activity was found to be in the MRS medium under anaerobic conditions. There were no differences in the antibacterial activity between MRS(+Se) and MRS(–Se) media. The antimutagenic activity of MRS(+Se) and MRS(–Se) extracts after culture with E. faecium M-74 as well as of live and killed cells of E. faecium M-74 grown in the presence or absence of Se against the genotoxicity of ofloxacin (OFL) and acridine orange (AO) was determined in the Euglena gracilis assay. The MRS(+Se) extracts showed a significantly higher activity in reducing the genotoxicity of OFL and AO than MRS(–Se) extracts. The live cells of the probiotic strain M-74 exhibited higher antimutagenic activity than the killed bacterial cells, but differed depending on the mutagen used. However, the live bacterial cells grown in the presence of Se showed significantly higher antimutagenic activity. These results suggest a potential benefit for the future development of new Se-enriched probiotics exhibiting higher antimutagenic properties.     

Response ofSetaria italica to inoculation withAzospirillum brasilense as compared toAzotobacter chroococcum

Summary Under controlled conditions in pots filled with sand, vermiculite and field soil, inoculation withA. brasilense-Cd ATCC 29729 or withAzotobacter chroococcum caused increases above controls in the weight and N content of panicles ofSetaria italica. In no case, however, did N increases in test plants exceed the initial total N content. High acetylene reduction activities (1,000–2,000 nmole/h/pot) could be found only in plants inoculated withAzospirillum. Inoculation withAzospirillum (strain-Cd) in the field caused a significant increase above noninoculated controls of 18.5% in shoot dry weight, ofSetaria italica. Azotobacter caused a non significant increase of 8%. No significant differences were found between yields ofSetaria italica grown in soil inoculated withA. chroococcum, and those of plants grown in the presence ofA. vinelandii. A. brasilense-Cd was more effective in the field thanA. brasilense Sp-7 ATCC 29145. The results suggest that Azospirillum may increase yields ofS. italica more efficiently than Azotobacter under local field conditions.     

The antimutagenic effect of selenium on 7,12-dimethylbenz(a)anthracene and metabolites in the amesSalmonella/microsome system

The antimutagenic effect of selenium as sodium selenite, sodium selenate, selenium dioxide, and seleno-methionine was studied in the AmesSalmonella/microsome mutagenicity test using 7,12-dimethylbenz(a)anthracene (DMBA) and some of its metabolites. Selenium (20 ppm) as sodium selenite reduced the number of histidine revertants on plates containing up to 100 μg DMBA/plate. Increasing concentrations of selenium as sodium selenite, sodium selenate, and selenium dioxide up to 40 ppm Se progressively decreased the number of revertants caused by 50 μg DMBA. DMBA and its metabolites 7-hydroxymethyl-12-methylbenz(a)anthracene, 12-hydroxymethyl-7-methylbenz(a)anthracene, and 3-hydroxy-7,12-dimethylbenz(a)anthracene were mutagenic forSalmonella typhimurium TA100 in the presence of an S-9 mixture. Selenium supplementation as Na₂SeO₃ reduced the number of revertants induced by these metabolites to background levels. The antimutagenic effect of inorganic selenium compounds cannot be explained by toxicity of selenium as determined by viability tests withSalmonella typhimurium TA100. Selenium supplementation in all forms examined, except sodium selenate, decreased the rate of spontaneous reversion. Selenium as sodium selenate was slightly mutagenic at concentrations of 4 ppm or less. Higher concentration of Na₂SeO₄ inhibited the mutagenicity of DMBA. The present studies support the anticarcinogenic potential of selenium and indicate that form and concentration are important factors in this trace element's efficacy.     

The branching of the inflorescence and vegetative shoot and taxonomy of the genusKummerowia (Leguminosae)

A study of the branching of the inflorescence and the vegetative shoot of the genusKummerowia, consisting ofK. stipulacea (Maxim.) Makino andK. striata (Thunb.) Schindler, has led to the following conclusions: (1) the inflorescences of both species are reduced compound cymes, (2) the branching system of the inflorescence ofKummerowia is not clearly different from that of the vegetative shoot and there are some transitional forms between both systems, and (3) the inflorescence ofKummerowia is different from the racemose inflorescences ofLespedeza andCampylotropis. Based on the differences found in the branching system of the inflorescence,Kummerowia is distinctly separated fromLespedeza andCampylotropis and is more correctly treated as a distinct genus from the latter two.