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Biomedical subjects

U Muller-Eberhard

Publications and source records attributed to U Muller-Eberhard.

At least 91 records · Page 5Linked to original sources
PubMed1983

Hemin binding to serum proteins and the catalysis of interprotein transfer.

The reaction of hemin (Hm) with human hemopexin (Hx) has been studied in a mixed dimethyl sulfoxide (Me2SO)-water solvent system and in aqueous caffeine solutions. In both media, the kinetics could be described by a single, second-order process: (formula - see text) with k = 1.8 X 10(6) M-1 s-1 in 40% Me2SO-water [pH 7.4, mu = 0.2 M (NaCl)] and k = 3.9 X 10(7) M-1 s-1 in water [pH 7.4 mu = 0.2 M (NaCl), [caffeine] = 0.025 M]. The reaction shows an ionic strength dependence consistent with a residual 1+ to 2+ charge in the vicinity of the binding region of the protein. The kinetics of the transfer of hemin from albumin to hemopexin (formula - see text) were studied as a function of concentration, ionic strength, pH, and temperature. In experiments conducted at 3 less than or equal to [Alb]0/[Hx]0 less than or equal to 20 where the transfer kinetics are first order, k' = 5 X 10(-3) S-1 at mu = 0.3 M (NaCl), pH 7.1; the reaction is strongly dependent on ionic strength and choice of electrolyte. The addition of imidazole catalyzes this transfer process via a ligand-mediated pathway with k' = 5 X 10(-3) + 21[Im]T2. At [Alb]0/[Hx]0 = 92, the noncatalyzed transfer reaction is second order. From the kinetic analysis of the reaction under these conditions, an estimate is made of the distribution of hemin between the two proteins at concentration levels which are characteristic of serum. The association of hemin and hemopexin is approximately 30 times faster than that of hemin and albumin, a finding consistent with the recycling function of hemopexin during heme transport to the liver parenchymal cells.

Blood Proteins↗
PubMed1983

The effect in vivo and in vitro of allylisopropylacetamide on the content of hepatic microsomal cytochrome P-450 2 of phenobarbital treated rabbits.

Rabbits treated with phenobarbital were given a single injection of allylisopropylacetamide (AIA) s.c. and/or heme i.v. Hepatic microsomes were isolated 1, 5 and 24 hours post injection and the microsomal contents of both total cytochrome P-450 chromophore, and the protein moiety of P-450 2 were determined by spectrophotometric and immunochemical methods respectively. AIA caused the levels of total P-450 chromophore and of P-450 2 protein to decline to 30% of the control values at 5 hours post-injection. Concurrent administration of heme with AIA diminished the decrease in the total microsomal content of P-450 chromophore but not in that of P-450 2 protein. These findings suggest that the destruction of the heme prosthetic group of P-450 by suicide substrates such as AIA may lead to an enhanced degradation of the apo-P-450.

Acetamides↗
PubMed1983

Hemopexin metabolism in patients with altered serum levels.

The rates of synthesis and degradation of hemopexin (Hx) were studied in vivo to determine the cause of altered serum levels of this protein as seen in hemolytic anemias, chronic neuromuscular diseases, and acute intermittent porphyria. The synthetic and fractional catabolic rates of Hx were measured in patients exhibiting low, normal, or elevated serum Hx levels. It was found that the elevated levels were mainly due to increased synthesis rather than decreased catabolism of Hx. In patients with elevated serum Hx levels, the mean synthetic rate of Hx (13 +/- 1.0 mg/kg/day) was twice that of the patients with normal Hx levels (6.6 +/- 0.3), whereas the fractional catabolic rate was 35.3 +/- 7.1% of the i.v. pool per day vs. 26.5 +/- 0.8 for controls. The low serum Hx levels observed in patients with sickle cell anemia appeared to be due to increased Hx catabolism (36.0 and 40.0% of the i.v. pool per day vs. 26.5 +/- 0.8 for controls) with no compensatory increase in synthesis. This latter finding is in agreement with a study in rhesus monkeys in which repeated administration of a large dose of heme caused an increase in the catabolism of hemopexin without a concurrent increase in its synthesis (J LAB CLIN MED 100:451, 1982). Our results indicate that although both synthesis and catabolism are increased in patients with elevated Hx levels, only catabolism is increased in patients with sickle cell anemia.

Adolescent↗
PubMed1982

Rabbit hepatic progesterone 21-hydroxylase exhibits a bimodal distribution of activity.

Progesterone 21-hydroxylase activity varies extensively among liver microsomes prepared from individual New Zealand White (NZW) rabbits. The 21-hydroxylase activities are distributed between two groupings that differ by more than tenfold in mean activity. Both male and female animals are represented in the two groupings. However, females exhibited the higher activity more frequently than males. The 21-hydroxylation of progesterone is catalyzed by one of the liver microsomal cytochrome P-450 isozymes, form 1, and these differences in activity are suggestive of differences in the occurrence of this isozyme among NZW rabbits.

Animals↗
PubMed1982

The role of cytosolic proteins in the intracellular transport of haem in rat liver. A dual-label approach.

After consecutive injections of delta-amino[3H]- and -[14C]-laevulinic acid, the incorporation of the two labels into haem associated with different subfractions of the liver was determined. Marked differences in the 14C/3H ratios were observed between haem associated loosely and tightly with microsomes and mitochondria and haem associated with three subfractions of the cytosol obtained by gel filtration. The effect of changing the amounts of delta-aminolaevulinic acid injected and of changing the interval between injections and killing of the animal on the ratios of labels in the haem of each subfraction was studied. The results are discussed in terms of the flow of haem from the mitochondria to other parts of the cell via putative cytosolic carrier proteins.

Aminolevulinic Acid↗
PubMed1982

Effects of haem infusion on biliary secretion of porphyrins, haem and bilirubin in man.

Employing a continuous bile collection, we measured the bile secretion of porphyrins, haem (iron protoporphyrin IX regardless of oxidation state) and bilirubin in five healthy subjects. The baseline values for the flow of porphyrins in the bile were: 4.7 +/- 1.9 nmol/h uroporphyrin, 27.3 +/- 3.8 nmol/h coproporphyrin and 39.2 +/- 11.7 nmol/h protoporphyrin. Bile haem flow was 59.7 +/- 12.6 nmol/h, and that of bilirubin 23.8 +/- 8.2 mumol/h. Following haem injection (6.4 mumol/kg) the flow of protoporphyrin but not of the other porphyrins was reduced, and the bile haem flow increased (232 +/- 109.5 nmol/h), while the flow of bilirubin did not increase significantly. A few patients with representative porphyrias showed the expected increase in copro- and protoporphyrin in the bile. The patient with coproporphyria exhibited a bile flow of coproporphyrin of 1470 +/- 133 nmol/h and of protoporphyrin of 334 +/- 29 nmol/h; haem infusion significantly reduced the bile flow of both porphyrins (to 649 +/- 101 for copro- and 215 +/- 36 nmol/for protoporphyrin). The patient with protoporphyria had an increased protoporphyrin flow, yet haem infusion caused no reduction in protoporphyrin flow (106 +/- 7 after v. 81.4 +/- 13 nmol/h before haem). In conclusion, we found that haem and porphyrins are normal constituents of bile, and that injected haem appears in bile. Bile bilirubin did not rise within 12 h after haem infusion a finding which warrants further investigation.

Adult↗
PubMed1982

Effect of 2,3,7,8-tetrachlorodibenzo-p-dioxin and phenobarbital on the occurrence and distribution of four cytochrome P-450 isozymes in rabbit kidney, lung, and liver.

The effect of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) and of phenobarbital (PB) on the distribution and occurrence of four cytochrome P-450 isozymes, Forms 2, 3, 4, and 6, in the kidney, lung, and liver of adult male rabbits was investigated using immunofluorescence. In the kidney, Forms 2 and 3 were localized in the proximal tubules of both untreated and PB-treated animals, while antibodies to Forms 4 and 6 showed weak to negative staining. In TCDD-pretreated animals, Forms 4 and 6 appeared in the renal endothelium, in addition to staining the proximal tubular epithelium intensely. Form 2 was the only isoenzyme of those studied found to be present in the lungs of normal and PB-pretreated rabbits; it was also present in lungs of TCDD-pretreated rabbits. Form 3 was not detected in any of the rabbit lungs examined. Forms 4 and 6, while not apparent in the lungs of normal or PB-treated animals, were found in the lungs of TCDD-treated animals and also appeared in the endothelium of the pulmonary arteries and veins. All forms tested were present in control liver. The staining for Form 2 was intense in the livers of PB-pretreated animals, as was the staining for Forms 4 and 6 in the livers of TCDD-pretreated animals. Our results indicate that, while PB altered the intensity of staining for Form 2 in the liver and kidney, TCDD altered both the staining intensity and distribution of the isozymes in kidney, lung, and liver, producing, for example, a localization of Forms 4 and 6 in the endothelium of both the kidney and lung which was not seen in either untreated or PB-pretreated rabbits.

Animals↗
PubMed1982

Effect of microsomal cytochrome P-450 isozyme induction on the mutagenic activation of 2-aminoanthracene.

Two rabbit microsomal cytochrome P-450 isozymes. Forms 4 and 6, which are differentially induced by 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) in the liver in an age-dependent fashion, catalyze the activation of 2-aminoanthracene to a mutagen in the Ames Salmonella mutagenesis assay. We have shown previously that in the presence of saturating concentrations of 2-aminoanthracene, Form 4 is 15-fold more active than Form 6 in the activation of this mutagen. Similar differences in the activation of 2-aminoanthracene are observed between liver microsomes isolated from TCDD-treated adult rabbits, in which Form 4 predominates, and microsomes from rabbit neonates exposed transplacentally to TCDD prior to birth, in which Form 6 predominates. However, when the extent of mutagenesis is limited by the amount of 2-aminoanthracene and is independent of the rate of activation, the number of revertants produced is similar for microsomes isolated from either newborn or adult TCDD-treated rabbits. Under these conditions, the extent of mutagenesis obtained for a given amount of 2-aminoanthracene will depend on the balance between activation and competing reaction pathways leading to detoxication. Thus, differences in the rate of activation between adult and newborn microsomes are probably offset by similar differences in the rates of competing pathways of metabolism. This is consistent with the finding that the overall rate of 2-aminoanthracene metabolism by the adult microsomes is greater than that of the neonate. In order for the extent of mutagenesis to be independent of rate, the half-life of 2-aminoanthracene was seen to be less than approximately 12 min.

Animals↗
PubMed1982

Effect of heme administration on hemopexin metabolism in the rhesus monkey.

Clinical conditions such as hemolytic anemias and certain neuromuscular diseases in which serum hemopexin levels are either increased or decreased were simulated in rhesus monkeys by administering heme intravenously daily at three dose levels over a period of 10 days. At the lower dose of heme (0.02 to 0.04 mg/kg/day), serum hemopexin levels were elevated to 150% of control (control = 53.3 +/- 2.8 U/100 ml). At the higher dose of heme (5.0 mg/kg/day), hemopexin levels decreased to 60% of control. After an intermediate dose of heme (0.6 mg/kg/day), no change was seen in the circulating hemopexin levels. These changes appeared to be specific for hemopexin, since neither the serum haptoglobin levels nor the transferrin level was affected by the heme administration at any of the dose levels. Parameters of hemopexin metabolism revealed that after administration of the low dose of heme there was a 76% increase in the net rate of hemopexin synthesis, resulting in a 65% increase in the intravascular pool size of hemopexin. At the intermediate dose there was a 43% increase in the rate of hemopexin synthesis accompanied by a 33% increase in catabolism, resulting in no net change in serum hemopexin concentrations. At the high dose of heme there was a 57% increase in catabolism of hemopexin without a concurrent increase in synthesis, resulting in lowered circulating hemopexin levels. These findings seem to indicate a relationship between the amount of heme presented to the liver and net hemopexin synthesis.

Animals↗
PubMed1981

Effects of hexachlorobenzene and iron loading on rat liver mitochondria.

The effects of hexachlorobenzene treatment and simultaneous iron-overload on the iron and porphyrin content of rat liver and rat liver mitochondria have been examined. In order to assess damages to the mitochondrial membrane occurring with these treatments, the content of malondialdehyde and selected functional properties of mitochondria were compared with those from control animals. Prolonged intake of hexachlorobenzene (8 weeks) resulted in a strikingly increased level of porphyrins together with a moderate increase in iron concentration. Simultaneous administration of hexachlorobenzene and iron-dextran caused the porphyrin level to reach 25% of the amount induced by hexachlorobenzene alone. The iron concentrations in liver as well as in liver mitochondria are also decreased under these conditions, as compared to the effect of iron-dextran. In contrast, the effects of hexachlorobenzene combined with iron-dextran on mitochondrial oxidative phosphorylation and malondialdehyde content are greater than those of either hexachlorobenzene or iron-dextran. These data suggest that porphyrin accumulation per se causes little deleterious effect and that both agents administered together act synergistically in causing damage to the mitochondrial membrane.

Animals↗
PubMed1981

Specificity in the activation and inhibition by flavonoids of benzo[a]pyrene hydroxylation by cytochrome P-450 isozymes from rabbit liver microsomes.

The effect of flavone and 7,8-benzoflavone on the metabolism of benzo[a]pyrene to fluorescent phenols by five cytochrome P-450 isozymes obtained from rabbit liver microsomes was determined. Benzo[a]pyrene metabolism was stimulated more than 5-fold by the addition of 600 microM flavone to a reconstituted monooxygenase system consisting of NADPH, cytochrome P-450 reductase, dilauroylphosphatidylcholine, and cytochrome P-450LM3c or cytochrome P-450LM4. In contrast, an inhibitory effect of flavone on benzo[a]pyrene metabolism was observed when cytochrome P-450LM2, cytochrome P-450LM3b, or cytochrome P-450LM6 was used in the reconstituted system. 7,8-Benzoflavone (50-100 microM) stimulated benzo[a]pyrene metabolism by the reconstituted monooxygenase system about 10-fold when cytochrome P-450LM3c was used, but benzo[a]pyrene hydroxylation was strongly inhibited when 7,8-benzoflavone was added to the cytochrome P-450LM6-dependent system. Smaller effects of 7,8-benzoflavone were observed on the metabolism of benzo[a]pyrene by the cytochrome P-450LM2-, cytochrome P-450LM3b-, and cytochrome P-450LM4-dependent monooxygenase systems. These results demonstrate that the activating and inhibiting effects of flavone and 7,8-benzoflavone on benzo[a]pyrene metabolism depend on the type of cytochrome P-450 used in the reconstituted monooxygenase system.

Animals↗
PubMed1981

Fluorimetric study of the binding of protoporphyrin to haemopexin and albumin.

Fluorescence spectra of protoporphyrin bound to its most affinitive site on human serum albumin, bound to human haemopexin and dissolved in human plasma reveal that, when present in plasma, at least 90% of this porphyrin is bound to albumin. Human serum albumin binds protoporphyrin with an affinity KA = 3 X 10(9)M-1 in phosphate-buffered saline. The affinity of haemopexin for protoporphyrin is 4 times smaller. From these data it is concluded that less than 1% of plasma protoporphyrin is bound to haemopexin. Implications of the data for protoporphyrin transport and clearance are discussed.

Animals↗
PubMed1981

The interaction of heme-hemopexin with CO.

The equilibria and kinetics of the reaction of heme-hemopexin with CO were studied. A stoichiometry of one CO/heme was determined, and the affinity of heme-hemopexin for CO was found to be pH-dependent. At pH 8.0, the affinity constant was 4.5 X 10(5) M-1 compared with 4 X 10(6) M-1 at pH 6.1. The kinetics of CO binding were also pH-dependent. A biphasic reaction at neutral pH could be resolved into a faster phase (kon = 2.2 X 10(3) M-1 s-1) solely found at pH 6.0, and a slower phase (kon = 2.0 X 10(2) M-1 s-1) solely found at pH 8.0. The dissociation reaction on the other hand was found to be independent of pH in the range examined (koff = 5 X 10(-4) s-1).

Animals↗
PubMed1980

Catalysis of divergent pathways of 2-acetylaminofluorene metabolism by multiple forms of cytochrome P-450.

Four highly purified forms of rabbit hepatic, microsomal cytochrome P-450 catalyze the N- and ring-hydroxylation of 2-acetylaminofluorene (AAF) at different rates. Form 4, the major form of the cytochrome induced by 2,3,7,8-tetrachlorodibenzo-p-dioxin in adult rabbit liver, catalyzed the N-hydroxylation of AAF more rapidly than did the other three forms. N-Hydroxy-2-acetylaminofluorene accounted for 70% of the metabolites formed by the action of this cytochrome. Form 6, the major form of the cytochrome induced by 2,3,7,8-tetrachlorodibenzo-p-dioxin in neonate rabbit liver, and Form 3, a constitutive form of the cytochrome, both metabolized AAF at one-half the rate observed for Form 5. Phenols accounted for more than 90% of the metabolites produced by these two cytochromes. The major phenobarbital-inducible cytochrome P-450, Form 2, exhibited practically no catalytic activity (< 1% of the other forms) with AAF as a substrate. Since N- and ring-hydroxylation are thought to represent divergent pathways of carcinogen metabolism (activation versus detoxification), the differential occurrence of the various cytochrome forms should affect the balance between these two reaction pathways. In this sense, cytochrome P-450 induction by 2,3,7,8-tetrachlorodibenzo-p-dioxin differentially affects the magnitude and direction of in vitro microsomal metabolism of AAF as a function of age.

2-Acetylaminofluorene↗