Selective induction of cytochrome b5 and NADH cytochrome b5 reductase by propylthiouracil

doi:10.1016/0024-3205(84)90524-1

Life Sciences

Volume 35, Issue 23, 3 December 1984, Pages 2327-2334

https://doi.org/10.1016/0024-3205(84)90524-1 Get rights and content

Abstract

Both the cytochrome b₅ level and NADH cytochrome b₅ reductase activity in rat liver microsomes were increased 2-fold by repeated i.p. administration of 1.5 mmol/kg propylthiouracil (PTU) for 2 weeks, but neither the cytochrome P-450 level nor NADPH cytochrome P-450 reductase activity were affected by the treatment. Liver microsomes from PTU-treated rats showed a significant decrease in aminopyrine N-demethylation, but not in benzphetamine N-demethylation, aniline hydroxylation or 7-ethoxycoumarin O-deethylation. A single administration of the compound had no effect on any components of the system. $In vitro$ , drug hydroxylation activities were not affected by PTU up to 1.0 mM. From the above evidence, repeated administration of PTU selectively induced cytochrome b₅ and NADH cytochrome b₅ reductase in rat liver microsomes.

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GGPP depletion initiates metaflammation through disequilibrating CYB5R3-dependent eicosanoid metabolism
2020, Journal of Biological Chemistry
Citation Excerpt :
If CYB5R3 is responsible for the apoptosis of GGPPS-deficient smooth muscle cells, inhibition of CYB5R3 activity should cause smooth muscle apoptosis also. Propylthiouracil (PTU) is a specific (43) and weak inhibitor of CYB5R3 (IC50 ∼275 μm), and 0.25 mm PTU is sufficient to cause a significant decrease in enzyme activity (44, 45). We treated the primary aorta smooth muscle cells with PTU from 1 to 100 μm.
Metaflammation is a primary inflammatory complication of metabolic disorders characterized by altered production of many inflammatory cytokines, adipokines, and lipid mediators. Whereas multiple inflammation networks have been identified, the mechanisms by which metaflammation is initiated have long been controversial. As the mevalonate pathway (MVA) produces abundant bioactive isoprenoids and abnormal MVA has a phenotypic association with inflammation/immunity, we speculate that isoprenoids from the MVA may provide a causal link between metaflammation and metabolic disorders. Using a line with the MVA isoprenoid producer geranylgeranyl diphosphate synthase (GGPPS) deleted, we find that geranylgeranyl pyrophosphate (GGPP) depletion causes an apparent metaflammation as evidenced by abnormal accumulation of fatty acids, eicosanoid intermediates, and proinflammatory cytokines. We also find that GGPP prenylate cytochrome b₅ reductase 3 (CYB5R3) and the prenylated CYB5R3 then translocate from the mitochondrial to the endoplasmic reticulum (ER) pool. As CYB5R3 is a critical NADH-dependent reductase necessary for eicosanoid metabolism in ER, we thus suggest that GGPP-mediated CYB5R3 prenylation is necessary for metabolism. In addition, we observe that pharmacological inhibition of the MVA pathway by simvastatin is sufficient to inhibit CYB5R3 translocation and induces smooth muscle death. Therefore, we conclude that the dysregulation of MVA intermediates is an essential mechanism for metaflammation initiation, in which the imbalanced production of eicosanoid intermediates in the ER serve as an important pathogenic factor. Moreover, the interplay of MVA and eicosanoid metabolism as we reported here illustrates a model for the coordinating regulation among metabolite pathways.
Amidoxime reductase system containing cytochrome b <inf>5</inf> type B (CYB5B) and MOSC2 is of importance for lipid synthesis in adipocyte mitochondria
2012, Journal of Biological Chemistry
Citation Excerpt :
In addition, simultaneous down-regulation of CYB5R1, CYB5R2, and CYB5R3 did not inhibit the amidoxime reductase activity either (data not shown). Finally, the involvement of CYB5R in the amidoxime reductase activity was studied using the specific CYB5R inhibitor PTU (27). Although PTU was able to efficiently inhibit the CYB5R activity with almost 90% efficiency (Fig. 6B), it was not able to significantly inhibit the benzamidoxime reductase activity in the OMM fraction (Fig. 6A), again confirming that CYB5R is not an essential component of the amidoxime reductase enzyme system in these cells.
Reduction of hydroxylamines and amidoximes is important for drug activation and detoxification of aromatic and heterocyclic amines. Such a reductase system was previously found to be of high activity in adipose tissue and liver, and furthermore, in vitro studies using recombinant truncated and purified enzymes suggested the participation of cytochrome b₅ reductase (CYB5R), cytochrome b₅ (CYB5), and molybdenum cofactor sulfurase C-terminal containing 1 and 2 (MOSC1 and -2). Here, we show that purified rat liver outer mitochondrial membrane contains high amidoxime reductase activity and that MOSC2 is exclusively localized to these membranes. Moreover, using the same membrane fraction, we could show direct binding of a radiolabeled benzamidoxime substrate to MOSC2. Following differentiation of murine 3T3-L1 cells into mature adipocytes, the MOSC2 levels as well as the amidoxime reductase activity were increased, indicating that the enzyme is highly regulated under lipogenic conditions. siRNA-mediated down-regulation of MOSC2 and the mitochondrial form of cytochrome b₅ type B (CYB5B) significantly inhibited the reductase activity in the differentiated adipocytes, whereas down-regulation of MOSC1, cytochrome b₅ type A (CYB5A), CYB5R1, CYB5R2, or CYB5R3 had no effect. Down-regulation of MOSC2 caused impaired lipid synthesis. These results demonstrate for the first time the direct involvement of MOSC2 and CYB5B in the amidoxime reductase activity in an intact cell system. We postulate the presence of a novel reductive enzyme system of importance for lipid synthesis that is exclusively localized to the outer mitochondrial membrane and is composed of CYB5B, MOSC2, and a third unknown component (a CYB5B reductase).
The many roles of cytochrome b<inf>5</inf>
2003, Pharmacology and Therapeutics
Four distinct suggestions have been made to explain the mechanism of the cytochrome b₅-imposed positive modifier action of the cytochrome P450 monooxygenase reaction. The first mechanism involves a direct input of an electron into the monooxygenase cycle. This is the second of the two electrons necessary for activation of molecular oxygen, and appears to be a rate-limiting step in the monooxygenase reaction. P450 monooxygenases all appear to be uncoupled to varying extents, releasing superoxide and hydrogen peroxide instead of oxidized substrate. A second mechanism suggests that cytochrome b₅ acts as a positive modifier of the monooxygenase by decreasing the extent of uncoupling of the monooxygenase reaction. The implication is that a slow input of the second electron allows uncoupling of a superoxide anion instead of formation of two-electron reduced oxygen. Faster input of the second electron via cytochrome b₅ would result in formation of more of the activated oxygen that reacts with substrate to form product. A third suggestion involves formation of a two-hemoprotein complex between cytochrome b₅ and cytochrome P450 that allows acceptance of two electrons from NADPH-cytochrome P450 reductase. Uncomplexed cytochrome P450 accepts an electron from the reductase, dissociates from it, binds oxygen, and re-associates with the reductase to accept another electron. Complexation with cytochrome b₅ enhances the rate of formation of the active oxygen by obviating the need for two interactions with reductase. The fourth mechanism has cytochrome b₅ serving as an effector without a reduction-oxidation role in the monooxygenation reaction. This effector function may be to enhance the breakdown of the oxygenated hemoprotein to products or to facilitate flow of electrons through the system.
Comparison of the effects of propylthiouracil, amiodarone, diphenylhydantoin, phenobarbital, and 3-methylcholanthrene on hepatic and renal T4 metabolism and thyroid gland function in rats
1991, Toxicology and Applied Pharmacology
We studied the effects of propylthiouracil (PTU), amiodarone (AMIO), diphenylhydantoin (DPH), phenobarbital (PB), and 3-methylcholanthrene (MC) on thyroid histomorphology, on the hepatic and renal enzymes involved in endogenous and exogenous metabolism, and on the plasma levels and pharmacokinetics of thyroid hormones after 7 and 14 days of treatment. PTU and PB, by decreasing both serum tetraiodothyronine (T4) and triiodothyronine (T3), induced a massive increase in serum thyrotropin (TSH) and thus induced thyroid hypertrophy. AMIO and MC, by decreasing respectively serum T3 and T4, also induced an increase of TSH, but to a lesser extent, not sufficient to induce thyroid hypertrophy. Hepatic 5′-deiodinase activity was decreased in all treated rats. Inhibition of this enzyme by PTU was demonstrated in vitro: AMIO also decreased the enzyme activity by a still unelucidated mechanism, which probably requires intact cell plasma membranes, whereas in PB- and MC-treated rats the decrease in enzyme activity certainly resulted from decreased serum concentrations of T4. In PTU-treated rats, and probably in MC-treated rats, decreases in circulating thyroid hormones were primarily due to impairment of synthesis and/or of secretion by the thyroid. In contrast, in PB-treated rats, the decrease in serum thyroid hormone levels seems to be due to increased excretion of these hormones, as T4 serum clearance was significantly increased. PB, a microsomal enzyme inducer, increased the cytochrome b₅ and P450 content as well as the cytochrome P450-dependent O-depentylation of pentoxyresorufin. The other type of enzyme inducer, MC, did not affect cytochrome b₅ and P450 levels, but did increase the cytochrome P450 dependent O-deethylation of ethoxyresorufin. PB increased the glucuronidation of morphine, whereas MC increased the glucuronidation of 1-naphthol. However, serum T4 clearance, mainly determined by its hepatic conjugation rate, was increased only in PB-treated rats. It appears from this study that the close metabolic relationship between the liver/kidney and the thyroid should be taken into consideration when the findings of chronic toxicology and carcinogenicity studies are interpreted.
Induction of anionic glutathione transferases in rat liver by propylthiouracil
1987, Comparative Biochemistry and Physiology. Part C, Comparative
1. The treatment of rats with propylthiouracil (PTU) resulted in an induction of not only cationic but also anionic glutathione (GSH) transferases.
2. DE-52 column chromatography of the anionie GSH transferases divided into five main peaks (I–V).
3. Peaks I–IV had a high activity toward l-chloro-2,4-dinitrobenzene (CDNB). Peak V, which is a new form of the enzyme, showed high activity to ethaerynic acid.
4. PTU induced peaks I and V, whereas phenobarbital increased the activity of only peak I.
Propylthiouracil, a selective inhibitor of NADH-cytochrome b<inf>5</inf> reductase
1986, FEBS Letters
Propylthiouracil inhibited the activity of NADH-cytochrome b₅ reductase of rat liver microsomes using potassium ferricyanide as electron acceptor. On the other hand, NADPH-cytochrome P-450 reductase activity was not affected by the compound. NADH-supported reduction of cytochrome b₅ was also inhibited by Propylthiouracil in the reconstituted system consisting of cytochrome b₅ and partially purified NADH-cytochrome b₅ reductase.

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Selective induction of cytochrome b5 and NADH cytochrome b5 reductase by propylthiouracil

Abstract

Biochem. Pharmac.

Biochem. Pharmac.

Toxicol. Appl. Pharmacol.

Biochem. Pharmac.

J. Biol. Chem.

J. Biol. Chem.

J. Biol. Chem.

Biochem. Pharmac.

Biochem. Pharmac.

Biochem. Pharmac.

Biochem. Biophys. Res. Commun.

J. Biol. Chem.

Biochim. Biophys. Res. Commun.

Ann. Rev. Pharmacol. Toxicol.

Res. Commun. Chem. Pathol. Pharmacol.

Experientia

Xenobiotica

J. Pharmacol. Exp. Ther.

Selective induction of cytochrome b₅ and NADH cytochrome b₅ reductase by propylthiouracil