An electrophoretic analysis of sheep plasma protein labeled with Na2 75SeO3 in vivo

Following an intravenous injection of 75Se, sodium selenite plasma samples were analyzed by two-dimensional electrophoresis. 75Se was detected by indirect autoradiography. From 0.5 to 53 hr postinjection of 75Se, 21 75Se peptides were detected. Both the isoelectric points and molecular weights of these peptides are reported. The molecular weights of the peptides ranged from 20,000 to 70,000 daltons.     

The effects of dietary L-ascorbic acid on the absorption and utilization of Na75SeO3 of silver-treated rats

Male hooded lister rats of the Rowett Institute strain, treated with silver in the drinking water, were used to study the effects of L-ascorbic acid on the utilization of Na75SeO3. Silver treatment dramatically decreased the absorption of 75Se from the rat GI tract. An attempt was made to utilize the reducing ability of L-ascorbic acid upon the absorption of 75Se from the rat GI tract. Increasing concentrations of L-ascorbic acid slightly increased the amount of 75Se absorbed from the rat gut.     

Na2SeO3对蚕豆根尖细胞遗传损伤作用的研究

采用蚕豆根尖微核实验和姊妹染色单体交换实验,研究亚硒酸钠(0.01～10.0 mg·L-1)对蚕豆根尖细胞的遗传损伤效应.结果表明一定浓度的亚硒酸钠(Na2SeO3)可导致蚕豆根尖细胞有丝分裂指数下降,间期细胞微核频率明显增高,且Na2SeO3的上述效应具有一定的剂量效应关系;同时Na2SeO3可诱导细胞姊妹染色单体交换(SCE)频率显著增高.研究表明,亚硒酸钠对蚕豆根尖细胞具有遗传毒害作用,并有可能对人体产生遗传损伤.     

Metabolism of Na2 75SeO4 in Astragalus bisulcatus Lima Bean, and Wheat: a Comparative Study

A comparative study of the metabolism of Na₂ ⁷⁵SeO₄ in Astragalusbisulcatus, luma bean, and wheat has been carried out. The resultsindicate that all three plants metabolize selenium extensively.Important differences were observed in the distribution of radioactivitybetween the various fractions isolated from the plants. Comparedto the protein fraction, the free amino acid fraction from A.bisulcatus contained a higher percentage of radioactivity. Theconverse was true for wheat and lima bean. As A. bisulcatusproteins contained a significant percentage of radioactivity,it is suggested, that the differences in the toxicity of seleniumtowards wheat, lime bean, and A. bisulcatus are difficult toexplain in terms of the differences in its incorporation intothe protein of the three species.     

Mechanisms of mutagenicity and toxicity of sodium selenite (Na2SeO3) in Salmonella typhimurium

The mechanisms of selenite toxicity and mutagenicity in S. typhimurium have been characterized. In contrast to previous reports, selenite toxicity was shown not to involve nonspecific incorporation into protein via the sulfur metabolic pathways. Selenite toxicity was, however, shown to involve its ability to act as an oxidizing agent, primarily through reactions with sulfhydryls. Strains which lack glutathione (GSH) are more sensitive to killing by sulfhydryl reagents. The selenite sensitivity of such a mutant was a biphasic phenomenon. The mutant was much more sensitive than a strain which contained GSH at lower selenite concentrations whereas, at higher concentrations, the mutant was much more resistant to selenite. The mechanism of selenite toxicity at lower concentrations in this mutant thus appeared to involve damage to intracellular sulfhydryls. The sensitization to higher doses of selenite by GSH could be explained by the generation of toxic oxygen species. The in vitro reactions of selenite with both cysteine and GSH readily produced H2O2 and O2-. A S. typhimurium strain which overproduces superoxide dismutase (SOD) and catalase was more resistant to high concentrations of selenite, but not killing by the lower doses. Pretreatment of cells with a nonlethal dose of selenite induced the synthesis of proteins which protected the cells from killing by H2O2 or high doses of selenite. Selenite was also a mutagen in the tester strain TA104, in which a number of other oxidizing agents have also been found to be mutagens. These results were consistent with a model in which the reactions of selenite and intracellular thiols with concomitant production of active oxygen species are the primary causal agents of selenite mutagenicity and toxicity in S. typhimurium.     

Phosphate incorporation into "nuclear" residual proteins of goose erythrocytes

Minor fractions of “residual proteins” from erythroblast-enriched avian erythroid cells were found to incorporate significantly larger amounts of radiophosphate than similar fractions from mature erythrocytes. This higher level of incorporation could not be detected in the bulk, unfractionated residual proteins from cell populations, respectively, enriched in erythroblasts, reticulocytes, or mature erythrocytes, probably because of variable amounts of phosphate-free protein. It was confirmed that when avian erythroid cells incorporated radiophosphate, specific activities of chromosomal “residual proteins” were orders of magnitude greater than those of histones, although levels of alkali-labile phosphate were only a fewfold higher.     

Reactive oxygen species from mitochondria mediate SW480 cells apoptosis induced by Na2SeO3

A number of selenium compounds have been found to inhibit tumorigenesis in a variety of animal and cell models. In order to explore the molecular mechanism involved in the anticarcinogenesis activity of selenium, we examined the effects of sodium selenite on cell viabilty, generation of reactive oxygen species (ROS), and mitochondrial transmembrane potential (Δω _m ) in human colonic carcinoma cells SW480. The result from MTT test showed that sodium selenite reduced cell viability. Morophologic and flow cytometric results indicated that Na₂SeO₃ induced the apoptosis of SW480 cells. Na₂SeO₃ increased the generation of intracellular ROS, whereas BAPTA-AM, rotenone, and NaCN completely inhibited the increase of ROS induced by Na₂SeO₃. Na₂SeO₃ also caused the disruption of Δω _m . The intracellular ROS increase and apoptosis induced by Na₂SeO₃ were significantly decreased by superoxide dismutase (SOD), catalase. These data suggest that the ROS mediate apoptosis induced by Na₂SeO₃ and mitochondria may be a major source of Na₂SeO₃-induced ROS.     

The incorporation of P into spectrin aggregates following incubation of erythrocytes in P-labelled inorganic phosphate

³²P was incorporated into spectrin by incubation of fresh erythrocytes with ³²P_i and glucose. The dimer and tetramer aggregates revealed only covalently-bound incorporation of phosphorus, while a higher aggregate of spectrin revealed both covalent and non-covalent incorporation. The specific activity of the covalently-bound phosphorus in all oligomers was identical, suggesting that the state of association is independent of phosphorylation. The non-covalent incorporation was shown to be due to the association of ATP with this higher aggregate. The nucleotide appears not to be bound directly to spectrin but rather to component 5 (erythrocyte actin) which is also found to be associated with this highly aggregated spectrin structure.     

Microcalorimetric studies of the action of Na2SeO3 on the growth of Halobacterium halobium R1

The effect of Na₂SeO₃ on the growth of Halobacterium halobium R1 was investigated by means of microcalorimetry at 37°C. The biological response to toxicants is observed as the inhibition of the rate constant of growth of living cells. A low concentration of Na₂SeO₃ stimulated the growth of H. halobium R1, and a high concentration of Na₂SeO₃ inhibited the growth of H. halobium R1. Toxicity may be expressed as the half-inhibition concentration (IC₅₀). The rate constants of growth (k) and the concentrations of Na₂SeO₃ (c) shows a linear relationship: k=1.790 × 10⁻⁶ − 2.27 × 10⁻³ c. The value of IC₅₀ obtained from the accompanying figure of I-c is 679 μg/mL.     

Asymmetric incorporation of Na+, K+-ATPase into phospholipid vesicles

Purified lamb kidney Na⁺, K⁺-ATPase, consisting solely of the Mτ = 95,000 catalytic subunit and the M_τ～- 44,000 glycoprotein, was solubilized with Triton X-100 and incorporated into unilamellar phospholipid vesicles. Freeze-fracture electron microscopy of the vesicles showed intramembranous particles of approximately 90–100 Å in diameter, which are similar to those seen in the native Na⁺,K⁺-ATPase fraction. Digestion of the reconstituted proteins with neuraminidase indicated that the glycoprotein moiety of the Na⁺,K⁺-ATPase was asymmetrically oriented in the reconstituted vesicles, with greater than 85% of the total sialic acid directed toward the outside of the vesicles. In contrast, in the native Na⁺,K⁺-ATPase fraction, the glycoprotein was symmetrically distributed. Purified glycoprotein was also asymmetrically incorporated into phospholipid vesicles using Triton X-100 and without detergents as described by R. I. MacDonald and R. L. MacDonald (1975, J. Biol. Chem.250, 9206–9214). The glycoprotein-containing vesicles were 500–1000 Å in diameter, unilamellar, and, in contrast to the vesicles containing the Na⁺,K⁺-ATPase, did not contain the 90- to 100-Å intramembranous particles. These results indicate that the intramembranous particles observed in the native Na⁺,K⁺-ATPase and in the reconstituted Na⁺,K⁺-ATPase are not due to the glycoprotein alone, but represent either the catalytic subunit, or the catalytic plus the glycoprotein subunit.     

The incorporation of a purified, membrane-bound form of guanylate cyclase into phospholipid vesicles and erythrocytes

The purified membrane-bound form of guanylate cyclase was incorporated into artificial unilamellar phospholipid vesicles. The rate and extent of enzyme incorporation into the vesicles was dependent upon the phospholipid concentration and the time period of incubation. The enzyme was incorporated at a significantly faster rate after removal of carbohydrate with endoglycosidase H. The incorporation of the enzyme led to a 10-fold decrease in the apparent maximal velocity and a 2-fold increase in the apparent Michaelis constant for MnGTP. Extraction of liposomes containing guanylate cyclase with 0.2% Lubrol PX resulted in the recovery of 85% of the original amount of added activity, suggesting that the decrease in maximal velocity was not due to enzyme denaturation. Phosphatidylcholine liposomes differentially effected the activity of the membrane-form of guanylate cyclase, dependent on the nature of the fatty acid present on the phospholipid. Specific activities ranged between 458 nmol/min per mg and 2.6 mumol/min per mg, dependent upon the fatty acids present. Liposomes containing the membrane-bound form of guanylate cyclase were subsequently fused with erythrocytes using poly(ethylene glycol) 4000 in attempts to introduce the enzyme into intact cells. The enzyme was successfully introduced into the erythrocytes; greater than 90% of the enzyme activity was subsequently shown to be associated with erythrocyte membranes. Cyclic GMP concentrations of erythrocytes increased from essentially nondetectable to 4 pmol/10(9) cells after introduction of the enzyme. These results demonstrate that guanylate cyclase can be incorporated into liposomes in an active state and that such liposomes can be used to introduce the enzyme into cells where it can subsequently function to generate cyclic GMP.