Hepatic Vitamin B12 Release and Transcobalamin II Synthesis in the Rat

1974 ◽  
Vol 47 (6) ◽  
pp. 531-545 ◽  
Author(s):  
W. G. E. Cooksley ◽  
J. M. England ◽  
L. Louis ◽  
M. C. Down ◽  
A. S. Tavill
Keyword(s):  
Vitamin B12 ◽  
Rat Liver ◽  
Plasma Concentrations ◽  
High Plasma ◽  
Free Form ◽  
Isolated Perfused Rat Liver ◽  
Low Concentrations ◽  
High Concentrations ◽  
Transcobalamin Ii

1. The release of 57Co-labelled vitamin B12 ([57Co]B12) and synthesis of transcobalamin II (TCII) by the isolated perfused rat liver were studied 10–42 days after the parenteral administration of a trace dose of 15 pmol (approximately 20 ng) of radioactive cyanocobalamin. 2. The rate of release of [57Co]B12 into plasma and bile was linear and constituted approximately 0.9% and 0.3% respectively of the initial hepatic radioactivity per hour of perfusion. 3. [57Co]B12 released into plasma was bound to TCII. Saturation of the total TCII by the addition of cyanocobalamin before perfusion resulted in the appearance of the hepatic [57Co]B12 in the free form. 4. These data were found to be compatible with the following observations in vivo: (i) rates of [57Co]B12 release as measured by urinary [57Co]B12 excretion after saturation of plasma binders with non-labelled cyanocobalamin; (ii) rates of biliary excretion of [57Co]B12. 5. Liver damage produced by hypoxaemia was associated with a fall in the rate of release of [57Co]B12. 6. TCII release occurred at a linear rate of almost twenty times that required for the binding of newly released hepatic vitamin B12. 7. Cycloheximide at a dose sufficient to inhibit release of TCII did not prevent the release of [57Co]B12 from the liver into plasma or bile. 8. Alteration of perfusate composition to contain either high plasma concentrations of vitamin B12 and low concentrations of unsaturated TCII or high plasma concentrations of vitamin B12 and high concentrations of unsaturated TCII had no effect on the rate of [57Co]B12 release into plasma or bile. 9. It is concluded that the fluxes of hepatic vitamin B12 and TCII are very rapid and that the release of vitamin B12 by the rat liver is controlled in the short term by factors other than the synthesis of TCII and the concentration of vitamin B12 or unsaturated transcobalamin in the plasma.

Relationship of Radioactive Vitamin B12 Release and Transcobalamin II Synthesis by the Isolated Perfused Rat Liver

1974 ◽  
Vol 46 (2) ◽  
pp. 19P-20P
Author(s):  
W. G. E. Cooksley ◽  
J. M. England ◽  
L. Louis ◽  
M. C. Down ◽  
A. S. Tavill
Keyword(s):  
Vitamin B12 ◽  
Rat Liver ◽  
Isolated Perfused Rat Liver ◽  
Perfused Rat Liver ◽  
Relationship Of ◽  
Transcobalamin Ii

Cardiovascular safety of MnDPDP and MnCl2

1997 ◽  
Vol 38 (5) ◽  
pp. 740-749 ◽  
Author(s):  
P. Jynge ◽  
H. Brurok ◽  
A. Asplund ◽  
R. Towart ◽  
H. Refsum ◽  
...  
Keyword(s):  
Ex Vivo ◽  
Plasma Concentrations ◽  
Cardiovascular Responses ◽  
Cellular Level ◽  
Mesenteric Arteries ◽  
Low Concentrations ◽  
Rat Hearts ◽  
Perfused Rat Hearts ◽  
High Concentrations

Purpose: To investigate the apparent discrepancy between expected basic physiological responses at the cellular level and the in vivo behaviour of both MnDPDP and MnCl2 adminstered i.v. prompted parallel investigations of these substances. Material and Methods: Studies were performed in isolated perfused rat hearts, isolated bovine mesenteric arteries, conscious dogs, and dogs with acute ischaemic heart failure. Results: These studies confirmed that Mn++ at high concentrations acted as a calcium antagonist inducing negative inotropy. Mn++ at low concentrations was an effective su-peroxide scavenger, conserving nitric oxide and facilitating vasodilation. Mn++ maintained or elevated heart rate (HR) and blood pressure (BP), and did not worsen existing cardiac failure. MnDPDP was about 10 times less potent than MnCl2 in eliciting these cardiovascular responses. Conclusion: The ex vivo properties of Mn++, inducing vasodilation and negative inotropy, are counter-balanced in vivo through the action of 2 mechanisms: extensive plasma protein binding reducing active M++, and the release of catecholamines which maintain or even raise HR and BP. Taken together with pharmacokinetic factors, including maximal plasma concentrations in humans given the recommended 5 μmol/kg dose, it is concluded that MnDPDP in normal clinical use represents no safety risk to the cardiovascular system.

Heterogeneity of Hepatic Vitamin B12 in the Rat after Parenteral Cyanocobalamin

1975 ◽  
Vol 49 (3) ◽  
pp. 257-264
Author(s):  
W. G. E. Cooksley ◽  
A. S. Tavill
Keyword(s):  
Vitamin B12 ◽  
Rat Liver ◽  
Intramuscular Injection ◽  
Isolated Perfused Rat Liver ◽  
Vitamin B ◽  
Double Labelling ◽  
Time Intervals ◽  
Rapid Release ◽  
Biphasic Release

1. The rate of radioactive vitamin B12 excretion into plasma and bile from the isolated perfused rat liver and into bile in vivo has been measured after the intramuscular injection of radioactive cyanocobalamin at various time-intervals before study. 2. With an interval of labelling of 10 or more days, the release of radioactive vitamin B12 over 4 h by the isolated perfused liver was linear and con-stituted approximately 35% and 1% of the hepatic radioactivity into plasma and bile respectively. 3. In contrast, after a shorter period of prelabelling (less than 7 days), there was a biphasic release of radioactive vitamin Blz: an initial rapid rate followed by a slower rate after about 1 h of perfusion. The total radioactive vitamin Blz was considerably increased (e.g. 25% and 5% of hepatic radioactivity into plasma and bile respectively during a 4 h perfusion of a liver labelled 18 h previously). 4. Confirmation of coexisting stable and labile pools of intrahepatic vitamin B12 was provided by: (a) the patterns of hepatic release ifter double labelling with 57Co-labelled and 58Co-labelled cyanocobalamin at different times before liver per-fusion; (b) the rates of biliary excretion of 57Co-labelled and 58Co-labelled vitamin B12 in vivo after different periods of prelabelling. 5. The rate and pattern of release were not altered by changes in the quantity of precursor cyanocobala-min, by phenobarbitone treatment or by the addition of cycloheximide to the perfusion. 6. The injection into rats of subcellular preparations of rat liver labelled in viuo with radioactive vitamin B12 demonstrated that hepatic heterogeneity did not depend on physical compartmentation. 7. Despite the rapid release rate from the liver recently administered radioactive cyanocobalamin, the hepatic radioactivity increased progressively with time after labelling in vivo, in contrast to the other tissues, where it decreased. In the presence of rapid bidirectional fluxes the ability of the liver to store vitamin B can be largely explained by the reduction in the rate of hepatic release that continues for about 10 days after parenteral administration of cyano-cobalamin.

scholarly journals Metabolism of inorganic sulphate in the isolated perfused rat liver. Effect of sulphate concentration on the rate of sulphation by phenol sulphotransferase

1978 ◽  
Vol 176 (3) ◽  
pp. 959-965 ◽  
Author(s):  
Gerard J. Mulder ◽  
Katja Keulemans
Keyword(s):  
Steady State ◽  
Plasma Concentration ◽  
Rat Liver ◽  
Isolated Perfused Rat Liver ◽  
Perfused Rat Liver ◽  
Perfusion Medium ◽  
Physiological Concentration ◽  
Sulphate Concentration ◽  
Inorganic Sulphate

1. The metabolism of inorganic [35S]sulphate (Na235SO4) was studied in the isolated perfused rat liver at three initial concentrations of inorganic sulphate in the perfusion medium (0, 0.65 and 1.30mm), in relation to sulphation and glucuronidation of a phenolic drug, harmol (7-hydroxy-1-methyl-9H-pyrido[3,4-b]indole). 2. [35S]Sulphate rapidly equilibrated with endogenous sulphate in the liver. It was excreted in bile and reached, at the lowest concentration in the perfusion medium, concentrations in bile that were much higher than those in the perfusion medium; at the higher sulphate concentrations, these concentrations were equal. The physiological concentration of inorganic sulphate in the liver, available for sulphation of drugs, is similar to the plasma concentration. 3. At zero initial inorganic sulphate in the perfusion medium, the rate of sulphation was very low and harmol was mainly glucuronidated. At 0.65mm-sulphate glucuronidation was much decreased and considerable sulphation took place, indicating efficient competition of conjugation by sulphation. At 1.30mm-sulphate the sulphation increased still further. 4. The results suggest that an important factor in sulphation is the relatively high Km of synthesis of adenosine 3′-phosphate 5′-sulphatophosphate (the co-substrate of sulphation) for inorganic sulphate, which is of the order of the plasma concentration of inorganic sulphate. The steady-state adenosine 3′-phosphate 5′-sulphatophosphate concentration may determine the rate of sulphate conjugation of drugs in the rat in vivo.

Bioassay of endotoxin clearance in vivo and by perfused rat liver

1976 ◽  
Vol 231 (1) ◽  
pp. 258-264 ◽  
Author(s):  
BJ Buchanan ◽  
JP Filkins
Keyword(s):  
Linear Relationship ◽  
Rat Liver ◽  
Salt Solution ◽  
Isolated Perfused Rat Liver ◽  
Perfused Rat Liver ◽  
Half Time ◽  
Rat Blood ◽  
Endotoxin Concentration ◽  
Induction Of Tolerance

Endotoxin clearances in vivo and by the isolated perfused rat liver were evaluated via bioassay in lead-sensitized rats. A linear relationship between the probit of shock lethality and the endotoxin dose in the probit range of 4-6 was validated. Endotoxin clearance in normal, fed rats displayed a linear relationship between the logarithm of the blood endotoxin concentration and time throughout the period of 15-240 min at doses of 500 and 1,000 mug/ rat; the half-time values were 58-63 min. Decreasing the endotoxin dose to 250 mug resulted in multiphasic clearance curves. Induction of tolerance to endotoxin resulted in marked acceleration of endotoxin clearance. Endotoxin clearance from the isolated perfused rat liver was not influenced by serum or rat blood as compared to clearance from a balanced salt solution. These data suggest that a physiologically stressful dose of endotoxin is slowly cleared from the blood and, therefore, circulates for prolonged periods.

scholarly journals Vitamin B12 transport in blood. I. Congenital deficiency of transcobalamin II

Blood ◽  
1975 ◽  
Vol 45 (1) ◽  
pp. 71-82 ◽  
Author(s):  
E Gimpert ◽  
M Jakob ◽  
WH Hitzig
Keyword(s):  
Vitamin B12 ◽  
Binding Capacity ◽  
Polyacrylamide Gel Electrophoresis ◽  
Congenital Deficiency ◽  
Complete Clinical Remission ◽  
High Concentrations ◽  
High Doses ◽  
Vitamin B12 Binder ◽  
Very High ◽  
Transcobalamin Ii

Abstract Some characteristics of vitamin B12 binding and transport in the serum of an infant with congenital hereditary transcobalamin II (TC II) deficiency were studied using the following parameters and methods: vitamin B12 level and binding capacity; electrophoretic mobility in polyacrylamide gel electrophoresis; various immunodiffusion and absorption experiments, using a specific anti-TC II antiserum and the patient's serum as antigen. The results of these studies point to a deficient synthesis of TC II. Parenteral administration of high doses of vitamin B12 was followed by rapid and complete clinical remission and the appearance of vitamin B12 binder in the alpha 2 region which is similar to “fetal binder.” Thus, very high concentrations of vitamin B12, either carrier free or bound to this alpha 2 binder, were able to correct the disturbed physiology of TC II deficiency, presumably by normalization of DNA-thymine synthesis.

scholarly journals Real-time study of the urea cycle using 15N n.m.r. in the isolated perfused rat liver

1992 ◽  
Vol 287 (3) ◽  
pp. 813-820 ◽  
Author(s):  
A Geissler ◽  
K Kanamori ◽  
B D Ross
Keyword(s):  
Rat Liver ◽  
Urea Cycle ◽  
Isolated Perfused Rat Liver ◽  
15N Recovery ◽  
Urea Synthesis ◽  
Perfused Liver ◽  
Biochemical Assays ◽  
Low Concentrations ◽  
15N Urea ◽  
Rate Limiting

1. Isolated rat liver was perfused with 10 mM-15NH4Cl, 5 mM-lactate and 1 mM-ornithine, or with 3 mM-[15N]alanine and 1 mM-ornithine, in haemoglobin-free medium. The liver was physiologically stable for over 3 h and synthesized urea at the rate of 1.15 mumol.min-1.g of liver-1 (15NH4(+)-perfused) or 0.41 mumol.min-1.g-1 ([15N]alanine-perfused). 2. The perfused liver was continuously monitored by 15N n.m.r. spectroscopy at 20.27 MHz for 15N. Well-resolved 15N resonances of precursors and intermediates of the urea cycle, present at tissue concentrations of 0.2-3.0 mumol/g, were observed from the intact liver in 5-40 min of acquisition. Key metabolites in liver extract and the final perfusion medium were analysed by n.m.r. and by biochemical assays to determine fractional 15N enrichment and the total 15N recovery. 3. In 15NH4(+)-perfused liver (n = 6), 15N incorporation into glutamate and alanine (1.0-1.3 mumol/g), as well as progressive formation of [15N2]urea, was observed during the first 2 h of perfusion. In the second and third hour, hepatic concentrations of [omega-15N]citrulline and [omega, omega'-15N]argininosuccinate increased to n.m.r.-detectable levels (0.3-0.9 mumol/g). The [15N]aspartate pool was large in the absence of added ornithine, but on its addition was rapidly incorporated into argininosuccinate (n = 3). 4. In [15N]alanine-perfused liver, major metabolites were [15N]glutamate, [gamma-15N]glutamine and [15N]urea. Urea-cycle intermediates were undetectable. 5. The results suggest that, in intact liver provided with excess ammonia, low concentrations of cytosolic argininosuccinate synthetase and argininosuccinate lyase limited the rate of metabolite flux in the urea cycle. By contrast, in alanine-perfused liver at a physiological rate of urea synthesis, mitochondrial carbamoylphosphate synthetase was rate-limiting. 6. The potential utility of 15N n.m.r. for study of metabolite channelling through urea-cycle enzymes in intact liver is discussed.

Mechanisms of vanadium action: insulin-mimetic or insulin-enhancing agent?

2000 ◽  
Vol 78 (10) ◽  
pp. 829-847 ◽  
Author(s):  
Margaret C Cam ◽  
Roger W Brownsey ◽  
John H McNeill
Keyword(s):  
Lipid Metabolism ◽  
Metabolic Effects ◽  
Isolated Cells ◽  
Insulin Mimetic ◽  
Endogenous Insulin ◽  
Carbohydrate And Lipid Metabolism ◽  
Low Concentrations ◽  
Tissue Sensitivity ◽  
High Concentrations

The demonstration that the trace element vanadium has insulin-like properties in isolated cells and tissues and in vivo has generated considerable enthusiasm for its potential therapeutic value in human diabetes. However, the mechanisms by which vanadium induces its metabolic effects in vivo remain poorly understood, and whether vanadium directly mimics or rather enhances insulin effects is considered in this review. It is clear that vanadium treatment results in the correction of several diabetes-related abnormalities in carbohydrate and lipid metabolism, and in gene expression. However, many of these in vivo insulin-like effects can be ascribed to the reversal of defects that are secondary to hyperglycemia. The observations that the glucose-lowering effect of vanadium depends on the presence of endogenous insulin whereas metabolic homeostasis in control animals appears not to be affected, suggest that vanadium does not act completely independently in vivo, but augments tissue sensitivity to low levels of plasma insulin. Another crucial consideration is one of dose-dependency in that insulin-like effects of vanadium in isolated cells are often demonstrated at high concentrations that are not normally achieved by chronic treatment in vivo and may induce toxic side effects. In addition, vanadium appears to be selective for specific actions of insulin in some tissues while failing to influence others. As the intracellular active forms of vanadium are not precisely defined, the site(s) of action of vanadium in metabolic and signal transduction pathways is still unknown. In this review, we therefore examine the evidence for and against the concept that vanadium is truly an insulin-mimetic agent at low concentrations in vivo. In considering the effects of vanadium on carbohydrate and lipid metabolism, we conclude that vanadium acts not globally, but selectively and by enhancing, rather than by mimicking the effects of insulin in vivo.Key words: vanadium, insulin-mimetic, insulin-like, insulin-enhancing.

scholarly journals Hypogonadism associated withCyp19a1 (Aromatase) post-transcriptional upregulation inCelf1-KO mice

2015 ◽  
pp. MCB.00074-15 ◽  
Author(s):  
Gaella Boulanger ◽  
Marie Cibois ◽  
Justine Viet ◽  
Alexis Fostier ◽  
Stéphane Deschamps ◽  
...  
Keyword(s):  
Rna Binding ◽  
Hypogonadotropic Hypogonadism ◽  
Type I ◽  
Male Gametes ◽  
Low Concentrations ◽  
Reporter Assays ◽  
High Concentrations ◽  
In Vitro Interaction

CELF1 is a multifunctional RNA-binding protein that controls several aspects of RNA fate. The targeted disruption of theCelf1gene in mice causes male infertility due to impaired spermiogenesis, the post-meiotic differentiation of male gametes. Here, we investigated the molecular reasons that underlie this testicular phenotype. By measuring sex hormone levels, we detected low concentrations of testosterone inCelf1-null mice. We investigated the effect ofCelf1disruption on the expression levels of steroidogenic enzyme genes, and we observed thatCyp19a1was upregulated.Cyp19a1encodes aromatase, which transforms testosterone into estradiol. Administration of testosterone or the aromatase inhibitor Letrozole partly rescued the spermiogenesis defects, indicating that a lack of testosterone associated with excessive aromatase contributes to the testicular phenotype. In vivo and in vitro interaction assays demonstrated that CELF1 binds toCyp19a1mRNA, and reporter assays supported the conclusion that CELF1 directly repressesCyp19a1translation. We conclude that CELF1 downregulatesCyp19a1/Aromatasepost-transcriptionally to achieve high concentrations of testosterone compatible with spermiogenesis completion. We discuss the implications of these findings with respect to reproductive defects in men, including patients suffering from isolated hypogonadotropic hypogonadism and myotonic dystrophy type I.

Effects of polymethoxylated flavone metabolites on apoB100 secretion and MTP activity in Huh7.5 cells

2021 ◽  
Vol 18 ◽  
Author(s):  
Danielle R. Gonçalves ◽  
Thais B. Cesar ◽  
John A. Manthey ◽  
Paulo I. Costa
Keyword(s):  
Lipid Synthesis ◽  
Dependent Manner ◽  
Transfer Protein ◽  
Low Concentrations ◽  
Wide Range ◽  
Cell Assays ◽  
Polymethoxylated Flavones ◽  
High Concentrations

Background: Citrus polymethoxylated flavones (PMFs) reduce the synthesis of liver lipoproteins in animal and in vitro cell assays, but few studies have evaluated the direct effects of their metabolites on this highly regulated process. Objective: To investigate the effects of representative metabolites of PMF on the secretion of liver lipoproteins using the mammalian cell Huh7.5. Method: In this study, the influences of three PMFs and five previously isolated PMF metabolites on hepatic apoB-100 secretion and microsomal transfer protein (MTP) activity were evaluated. Tangeretin (TAN), nobiletin (NOB) and 3,5,6,7,8,3′,4′-heptamethoxyflavone (HMF), and their glucuronides (TAN-Gluc, NOB-Gluc and HMF-Gluc) and oxidatively demethylated metabolites (TAN-OH, NOB-OH, HMF-OH) were incubated with Huh7.5 cells to measure their inhibitory effects on lipid synthesis. Results: The results showed that TAN, HMF and TAN-OH reduced the secretion of apoB-100 in a dose-dependent manner, while NOB and the other tested metabolites showed no inhibition. MTP activity in the Huh7.5 cells was significantly reduced in the presence of low concentrations of TAN, and in high concentrations of NOB-OH. This study also showed that PMFs and PMF metabolites produced a wide range of effects on apoB-100 secretion and MTP activity. Conclusion: The results suggest that while PMFs and their metabolites control dyslipidemia in vivo, the inhibition of MTP activity cannot be the only pathway influenced by these compounds.

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