Biomedical subjects

Raymond F Burk

Publications and source records attributed to Raymond F Burk.

12 recordsLinked to original sources

PubMed2004

Neurological dysfunction occurs in mice with targeted deletion of the selenoprotein P gene.

Brain function and selenium concentration are well maintained in rodents under conditions of selenium deficiency. Recently, however, targeted deletion of the selenoprotein P gene (Sepp) has been associated with a decrease in brain selenium concentration and with neurological dysfunction. Studies were conducted with Sepp(-/-) and Sepp(+/+) mice to characterize the neurological dysfunction and to correlate it with dietary selenium level. When weanling Sepp(-/-) mice were fed the basal diet (<0.01 mg/kg selenium) supplemented with 0, 0.05 or 0.10 mg selenium/kg, they developed spasticity that progressed and required euthanasia. Supplementing the diet with > or =0.25 mg selenium/kg prevented the neurological dysfunction. To determine whether neurological dysfunction would occur in more mature Sepp(-/-) mice deprived of selenium, Sepp(-/-) mice that had been fed the basal diet supplemented with 1.0 mg selenium/kg for 4 wk were switched to a selenium-deficient diet. Within 3 wk they had developed neurological dysfunction and weight loss. At 3 wk, the 1.0 mg selenium/kg diet was reinstituted. Neurological function stabilized but did not return to normal. Brain selenium concentration did not increase. Weight gain resumed. This study shows that neurological dysfunction occurs when selenium supply to the brain is curtailed and that the dysfunction is not readily reversible. Both the absence of selenoprotein P and a low dietary selenium supply are necessary for the dysfunction to occur, indicating that selenoprotein P and at least one other form of selenium supply the element to the brain.

Animals↗

PubMed2004

Sequential development of flagellar defects in spermatids and epididymal spermatozoa of selenium-deficient rats.

In this study cauda epididymal spermatozoa of rats maintained on a selenium-deficient diet for 5 and 7 months exhibited an array of flagellar defects. Spermatids and spermatozoa were analyzed by light and electron microscopy to define the appearance of flagellar abnormalities during spermiogenesis and post-testicular sperm development. Late spermatids of selenium-deficient rats displayed normal structural organization of the flagellar plasma membrane, axoneme, outer dense fibers, fibrous sheath and annulus, but they exhibited a premature termination of the mitochondrial sheath. A comparison of late spermatids and caput epididymal spermatozoa revealed that a late step in flagellar differentiation was the structural remodeling of the annulus and its accompanying fusion with both the fibrous sheath and the mitochondrial sheath. In selenium-deficient animals, however, the annulus failed to fuse with the mitochondrial sheath, generating an apparent weak point in the flagellum. After epididymal passage, cauda epididymal spermatozoa of selenium-deficient animals also exhibited extensive flagellar disorganization resulting from the apparent sliding and extrusion of specific outer dense fiber-doublet microtubule complexes from the proximal and the distal ends of the mitochondrial sheath and the accompanying loss of the midpiece plasma membrane. Only fiber complex number 4 was extruded proximally, whereas fibers 4, 5, 6 and 7 were extruded from the mitochondrial sheath-deficient posterior midpiece. Axonemal fibers 8, 9, 1, 2 and 3 retained their normal geometric relationships. These data suggest that the known loss of male fertility in selenium deficiency results from the sequential development of sperm defects expressed during both spermiogenesis and maturation in the epididymis.

Animals↗

PubMed2003

Mass spectrometric identification of N- and O-glycosylation sites of full-length rat selenoprotein P and determination of selenide-sulfide and disulfide linkages in the shortest isoform.

Rat selenoprotein P is an extracellular glycoprotein of 366 amino acid residues that is rich in cysteine and selenocysteine. Plasma contains four isoforms that differ principally by length at the C-terminal end. Mass spectrometry was used to identify sites of glycosylation on the full-length protein. Of the potential N-glycosylation sites, three located at residues 64, 155, and 169 were occupied, while the two at residues 351 and 356 were not occupied. Threonine 346 was variably O-glycosylated. Thus, full-length selenoprotein P is both N- and O-glycosylated. The shortest isoform of selenoprotein P, which terminates at residue 244, was analyzed for selenide-sulfide and disulfide linkages. In this isoform, a single selenocysteine and seven cysteines are present. Mass spectrometric analysis indicated that a selenide-sulfide bond exists between Sec40 and Cys43. Two disulfides were also detected as Cys149-Cys167 and Cys153-Cys156. The finding of a selenide-sulfide bond in the shortest isoform is compatible with a redox function of this pair that might be analogous to the selenol-thiol pair near the C terminus of animal thioredoxin reductase. The disulfide formed by Cys153-Cys156 also has some characteristics of a redox active pair.

Amino Acid Sequence↗

PubMed2003

Loss of activity of the selenoenzyme thioredoxin reductase causes induction of hepatic heme oxygenase-1.

The stress response enzyme heme oxygenase (HO)-1 is induced in livers of selenium-deficient rodents, probably to compensate for loss of certain selenoproteins. We sought to identify those selenoproteins. Selenium-replete mice with genetic deletion of selenoprotein P or glutathione peroxidase-1 did not have elevated hepatic HO activity, thus ruling out involvement of those selenoproteins in HO-1 induction by selenium deficiency. However, inhibition of thioredoxin reductase (TrxR) by a low dose of gold in the form of aurothioglucose led to induction of hepatic HO activity. Moreover, further induction by phenobarbital was observed. This HO-1 induction pattern is also seen in selenium-deficient mice. In the rat hepatoma cell line H4IIE, inhibition of TrxR by aurothioglucose or by 1-chloro-2,4-dinitrobenzene led to induction of HO-1. We conclude that loss of TrxR is responsible for the induction of HO-1 by selenium deficiency.

Animals↗

PubMed2003

Deletion of selenoprotein P alters distribution of selenium in the mouse.

Selenoprotein P (Se-P) contains most of the selenium in plasma. Its function is not known. Mice with the Se-P gene deleted (Sepp(-/-)) were generated. Two phenotypes were observed: 1) Sepp(-/-) mice lost weight and developed poor motor coordination when fed diets with selenium below 0.1 mg/kg, and 2) male Sepp(-/-) mice had sharply reduced fertility. Weanling male Sepp(+/+), Sepp(+/-), and Sepp(-/-) mice were fed diets for 8 weeks containing <0.02-2 mg selenium/kg. Sepp(+/+) and Sepp(+/-) mice had similar selenium concentrations in all tissues except plasma where a gene-dose effect on Se-P was observed. Liver selenium was unaffected by Se-P deletion except that it increased when dietary selenium was below 0.1 mg/kg. Selenium in other tissues exhibited a continuum of responses to Se-P deletion. Testis selenium was depressed to 19% in mice fed an 0.1 mg selenium/kg diet and did not rise to Sepp(+/+) levels even with a dietary selenium of 2 mg/kg. Brain selenium was depressed to 43%, but feeding 2 mg selenium/kg diet raised it to Sepp(+/+) levels. Kidney was depressed to 76% and reached Sepp(+/+) levels on an 0.25 mg selenium/kg diet. Heart selenium was not affected. These results suggest that the Sepp(-/-) phenotypes were caused by low selenium in testis and brain. They strongly suggest that Se-P from liver provides selenium to several tissues, especially testis and brain. Further, they indicate that transport forms of selenium other than Se-P exist because selenium levels of all tissues except testis responded to increases of dietary selenium in Sepp(-/-) mice.

Animals↗

PubMed2003

Loss of selenium from selenoproteins: conversion of selenocysteine to dehydroalanine in vitro.

Characterization of reduced and alkylated rat selenoprotein P by mass spectrometry yielded selenopeptides from which one or more selenium atoms were missing. Predicted selenopeptide mass peaks were accompanied by peaks corresponding to the conversion of one or more selenocysteine residues to dehydroalanine(s). Experiments were carried out to determine whether this loss of selenium occurred in vitro. A selenopeptide was isolated that contained two selenocysteine residues that were both in selenide-sulfide linkages with cysteine residues. After the peptide had been reduced and alkylated, in addition to the predicted mass peak with both selenocysteine residues present, two mass peaks were detected at positions expected for conversion of one and two selenocysteine residues of this selenopeptide to dehydroalanine residues, which was confirmed by tandem mass spectrometry. Similar findings were obtained from a study of another selenoprotein, rat plasma glutathione peroxidase. These results indicate that selenium atoms are lost from selenoproteins during purification and characterization. The loss of selenium from selenoproteins is probably through the mechanism of oxidation of selenocysteine residue to selenoxide followed by syn-beta-elimination of selenenic acid during sample processing.

Alanine↗

PubMed2003

Combined deficiency of vitamins E and C causes paralysis and death in guinea pigs.

BACKGROUND: On the basis of in vitro studies, the antioxidant nutrients vitamins E and C are postulated to interact in vivo. OBJECTIVE: We developed a guinea pig model to evaluate the combined deficiency of vitamins E and C in vivo. DESIGN: Weanling guinea pigs were fed a control diet or a vitamin E-deficient diet for 14 d, after which one-half of each group had vitamin C removed from their diet, thus creating 4 diet groups. Some animals were observed for clinical signs. Others were killed for evaluation. RESULTS: Of 21 guinea pigs that were observed after being fed the diet deficient in both vitamins, 8 died 9 +/- 2 d (x +/- SD) after starting the diet. Eight additional guinea pigs developed a characteristic syndrome at 11 +/- 3 d. First, they became paralyzed in the hind limbs. Within a few hours, the paralysis progressed to include all 4 limbs and caused difficulty in breathing, which would have caused death had the animals not been euthanized. Histopathologic evaluation did not identify a lesion in the muscles or nervous system that could account for the paralysis. Biochemical measurements confirmed the deficiencies and indicated that the double deficiency caused lipid peroxidation in the central nervous system. CONCLUSIONS: A distinct clinical syndrome of combined vitamin E and vitamin C deficiency occurs in guinea pigs. This syndrome indicates that these antioxidant vitamins are related in vivo. We speculate that acute oxidative injury in the central nervous system underlies the clinical syndrome.

Animals↗

PubMed2003

Selenoprotein metabolism and function: evidence for more than one function for selenoprotein P.

Biological functions of selenium are exerted by selenoproteins that contain selenocysteine in their primary structure. Selenocysteine is synthesized and inserted into proteins cotranslationally by a complex process. Families of selenoproteins include the glutathione peroxidases, the iodothyronine deiodinases and the thioredoxin reductases. These are redox enzymes that take advantage of the chemical properties of selenium to catalyze, respectively, removal of hydroperoxides by glutathione, deiodination of thyroid hormones and support of cellular processes requiring reduction of disulfides. Approximately 10 additional selenoproteins have been identified. One of them, selenoprotein P, is an extracellular protein that contains most of the selenium in plasma. It associates with endothelial cells, probably through its heparin-binding properties. Selenoprotein P has been postulated to protect against oxidative injury and to transport selenium from the liver to peripheral tissues. Selenium-dependent protection against diquat-induced liver necrosis and lipid peroxidation in the rat correlates with the presence of selenoprotein P. Recent results support a transport function. When (75)SeO(3)(2-) was administered intravenously to rats, liver tissue took up (75)Se within minutes, associated with a rapid decline in plasma (75)Se. Brain tissue did not begin accumulating (75)Se until (75)Se-labeled selenoprotein P had begun appearing in the plasma after 30 min. These results suggest that tissues like liver can take up small-molecule forms of selenium whereas presence of the element in selenoprotein P facilitates uptake by tissues like brain. Thus, there is evidence for both antioxidant and selenium transport functions of selenoprotein P.

Animals↗

PubMed2003

Selective induction of liver parenchymal cell heme oxygenase-1 in selenium-deficient rats.

Liver heme oxygenase (HO) activity is higher in selenium-deficient rats than in control animals under basal conditions and is further increased in them, but not in controls, by phenobarbital treatment. In the present study we characterized liver HO induction by selenium deficiency using molecular methods. Severe selenium deficiency in rats caused a doubling of liver HO activity without affecting spleen, kidney, brain, or testis HO activities. HO-1 protein and mRNA were increased to accompany the increased HO activity, but HO-2 protein and mRNA were not increased. Fractionation of the liver into hepatocyte and Kupffer cell/endothelial cell fractions revealed that the increased HO activity resides in the hepatocyte fraction. Immunohistochemical localization of HO-1 protein confirms the induction of HO-1 taking place solely in hepatocytes and throughout the liver lobule. Phenobarbital treatment sharply increased HO-1 mRNA and protein expression in selenium-deficient liver and HO activity in hepatocytes, but had no effect in control liver or in the Kupffer cell/endothelial cell fraction of selenium-deficient liver. Electrophoretic mobility shift assays showed increased AP-1 binding activity, suggesting an involvement of this redox-sensitive transcription factor in the induction by phenobarbital of HO-1 in selenium deficiency. We speculate that selenium deficiency affects hepatic antioxidant selenoproteins, resulting in an up-regulation of HO-1.

Animals↗

PubMed2002

Thioredoxin reductase reduces lipid hydroperoxides and spares alpha-tocopherol.

We investigated whether and how rat liver thioredoxin reductase spares alpha-tocopherol in biomembranes. Purified hydroperoxides of beta-linoleoyl-gamma-palmitoylphosphatidylcholine were decreased 35% by treatment with thioredoxin reductase and 54% by thioredoxin reductase plus E. coli thioredoxin. Thioredoxin reductase also halved the amount of hydroperoxides that had been formed during photoperoxidation of liposomes composed of beta-linoleoyl-gamma-palmitoylphosphatidylcholine, and of emulsions of both cholesterol and cholesteryl linolenate. In erythrocyte ghosts, thioredoxin reductase spared alpha-tocopherol from oxidation by both soybean lipoxygenase and ferricyanide. Thioredoxin reductase also decreased F(2)-isoprostanes in ghosts oxidized by ferricyanide, suggesting that its ability to spare alpha-tocopherol relates to reduction of lipid hydroperoxides.

Animals↗

PubMed2002

Mass spectrometric characterization of full-length rat selenoprotein P and three isoforms shortened at the C terminus. Evidence that three UGA codons in the mRNA open reading frame have alternative functions of specifying selenocysteine insertion or translation termination.

Selenoprotein P is an abundant extracellular glycoprotein. Its mRNA contains 10 UGAs in an open reading frame terminated by a UAA. This predicts that full-length selenoprotein P will contain 10 selenocysteine residues. Full-length selenoprotein P and three smaller isoforms that have identical N termini have been demonstrated. Selenoprotein P was purified from rat plasma, and the four isoforms were separated by heparin chromatography and SDS-PAGE. Mass spectrometric peptide analysis of the full-length isoform verified 357 of its 366 predicted amino acid residues, including its C terminus and all 10 selenocysteines. The C termini of the smaller isoforms were characterized by mass spectrometry. The shortened isoforms terminated where the second, third, and seventh selenocysteine residues were predicted to be. This suggests that all isoforms arise from the same mRNA and that the UGAs that specify the second, third, and seventh selenocysteines in full-length selenoprotein P can alternatively serve to terminate translation, producing the shorter isoforms.

Amino Acid Sequence↗

PubMedDate not supplied

Selenium, an antioxidant nutrient.

Selenium is an essential constituent of a number of enzymes, some of which have antioxidant functions. Deficiency of the element in animals makes them susceptible to injury by certain types of oxidative stress. At least 1 human disease occurs only in selenium-deficient individuals. Therefore, it seems prudent to avoid selenium deficiency. The plasma (or serum) selenium concentration is often used to assess selenium nutritional status. A plasma selenium concentration of 8 micrograms/dL or greater in a healthy subject indicates that plasma selenoproteins are optimized and the subject is selenium replete. The Third National Health and Nutrition Examination Survey determined plasma selenium in 17,630 subjects in the United States. Its results indicate that more than 99% of the subjects studied were selenium replete. The Institute of Medicine has set the Recommended Dietary Allowance for selenium at 55 micrograms per day for adults. Since most estimates of selenium intake in the United States are 80 micrograms per day or greater, routine selenium supplementation is not recommended in the United States.

Antioxidants↗