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PubMed1996

Isozyme-specific glutathione S-transferase inhibitors potentiate drug sensitivity in cultured human tumor cell lines.

Novel glutathione (GSH) analogs, previously shown to inhibit glutathione S-transferase (GST) activity at about 1 microM in vitro, were tested for their ability to potentiate the killing of cultured tumor cells by chemotherapeutic drugs. When tested at doses up to 200 microM, the analogs were neither toxic nor capable of potentiating drug toxicity unless the diethyl ester (DEE) form was used for treatment of the cells. HPLC analysis revealed rapid internalization of the DEE and intracellular conversion to a monoethyl ester form that accumulated in the cell, followed by a more gradual loss of the second ester to generate the active parent form. For the four GSH analogs tested, the ability of the DEE forms to potentiate chlorambucil (CMB) toxicity in HT-29 human colon adenocarcinoma cells strongly correlated with the in vitro ability of the parent form to inhibit recombinant human P1-1. This isozyme is the dominant form of GST present in HT-29 cells. Of the four analog DEEs tested, gamma-glutamyl-S-(benzyl)cysteinyl-R(-)-phenyl glycine (TER 117) DEE was the most effective in potentiating CMB toxicity in several cell lines: HT-29, HT4-1 (HT-29 subclone), SKOV-3 ovarian carcinoma, and SK VLB (vinblastine-resistant variant of SKOV-3) cells. gamma-Glutamyl-S-(octyl)cysteinyl-glycine (TER 143) DEE potentiated mitomycin C (MTC) toxicity in HT4-1 and SK VLB cells while TER 117 DEE did not. TER 117 DEE enhanced melphalan effects on xenografts of HT4-1 in mice to a similar extent as that achieved with the previously described nonspecific GST inhibitor, ethacrynic acid. Taken together, our results indicate that cell-permeable analogs of GSH can potentiate cytotoxicity of common chemotherapeutic drugs and this effect has a strong positive correlation with the ability of the analogs to inhibit specific GST isozymes.

Adenocarcinoma↗
PubMed2000

Identification and localization of glutathione S-transferase as a potential target enzyme in Brugia species.

Brugia filarial nematodes are pathogenic lymphatic-dwelling parasites that, like other helminths, may modify the host's defense mechanisms by a major detoxification process involving glutathione-binding proteins such as glutathione S-transferases (GSTs). In the present study, soluble extracts of third-stage larvae, adult male and female worms, microfilariae of either B. pahangi or B. malayi or the adult worm excretory-secretory products of B. malayi were used to determine GST activity. These extracts and affinity-purified fractions of B. pahangi adult worms had a specific enzymatic activity when 1-chloro-2,4-dinitrobenzene was used as a substrate. The observance of this enzyme in all life cycle stages of Brugia spp. demonstrates its ubiquitous nature. Lavage of intraperitoneally infected jirds, but not that of uninfected jirds, also showed increased enzymatic activity, suggesting that GST is secreted in vivo. Soluble proteins of both Brugia spp. were strongly recognized by antibodies in sera from rabbits immunized with affinity-purified native GST of Onchocerca volvulus. Immunohistochemical studies localized these proteins in adult worms, demonstrating cross-reactivity between the GST of these two filarial nematodes. The effect of this enzyme on the motility and viability of adult worms, microfilariae, and larvae was tested in vitro using a battery of known GST inhibitors. Of all those tested, ethacrynic acid, N-ethylmalemide, 4-nitropyridine-oxide, or 1-chloro-2,4-dinitrobenzene at micromolar concentrations reduced the viability and motility of microfilariae, third-stage larvae, and adult worms. These results suggest that Brugia GSTs are major metabolic enzymes and may play an important role in the parasite's survival.

Animals↗
PubMed1975

Magnesium deficiency and cardiac disorders.

Magnesium deficiency can occur in congestive heart failure, after diuresis with furoxemide, ethacrynic acid and mercurials, and with digitalis intoxication, diabetic acidosis, acute and chronic alcoholism, delerium tremens, cirrhosis, malabsorption syndromes, protracted postoperative cases, open heart surgery, the diuretic phase of acute tubular necrosis, and with hypoparathyroidism, primary aldosteronism, juxta-glomerular hyperplasia and pancreatitis. Two cases of serious ventricular arrhythmias associated with magnesium depletion are described. Clinical manifestations are vague but center around neurologic symptoms such as weakness, tremors, stupor, coma, nausea, vomiting and anorexia. Serious cardiac arrhythmias also occur with magnesium depletion. Magnesium appears to be very useful in hypomagnesemic or digitalis-toxic tachyarrhythmias. Magnesium may also be valuable in normomagnesemic tachyarrhythmias. Ten to fifteen milliliters of a 20 percent magnesium sulfate solution, given intravenously over 1 minute, followed by a slow 4 to 6 hour infusion of 500 ml of 2 per cent magnesium sulfate in 5 per cent dextrose in water is recommended. Recurrence of arrhythmias is common and a second infusion of magnesium sulfate may be necessary. Hypermagnesemia occurs frequently in renal insufficiency, and magnesium therapy may then be contraindicated. Serum levels above 5.5 meq/liter should be avoided. Loss of deep tendon reflexes and a decrease in respiratory rate can be used as guides to magnesium therapy. A plea is made for frequent analysis of serum magnesium so that more knowledge can be gained regarding this important biologic element in cardiovascular disorders.

Alcoholism↗
PubMed1987

The purification and characterization of glutathione S-transferase from the hepatopancreas of the blue crab, Callinectes sapidus.

High glutathione S-transferase activity was found in the cytosol of F-cells from the hepatopancreas of the blue crab (Callinectes sapidus). Purification of glutathione S-transferase from hepatopancreas extracts by Sephadex G-200, DEAE-Sephacel, and chromatofocusing resulted in the isolation of two isozymes with isoelectric points of 5.9 and 5.7, as determined by analytical isoelectric focusing. Using 1-chloro-2,4-dinitrobenzene as the substrate the specific activities of the two purified isozymes were 222 and 182 mumol/min/mg, respectively. There was no evidence for basic transferase isozymes. In addition to 1-chloro-2,4-dinitrobenzene the purified glutathione S-transferase isozymes showed activity with p-nitrophenyl acetate, p-nitrobenzyl chloride, bromosulfophthalein, and benzopyrene oxide. Thus, both substitution and addition reactions associated with vertebrate glutathione S-transferase were found in the crab transferases. There was no when ethacrynic acid, methyl iodide, trans-4-phenyl-3-buten-2-one, 1,2-epoxy-(p-nitrophenoxy)propane, cumene hydroperoxide, and t-butyl hydroperoxide were used as substrates. The lack of peroxidase activity is of interest since this activity is commonly found in vertebrate transferase isozymes. The two transferases had a dimeric Mr of 40,800 with similar amino acid compositions and similar kinetic parameters (Vmax, Km, and pH maxima) with 1-chloro-2,4-dinitrobenzene as substrate. The two transferases could be distinguished by their isoelectric points, molecular mass of the monomers (22,300 for GST 1 and 22,300 and 22,400 for GST 2), and different inhibitor mechanisms with hematin and bromosulfophthalein.

Amino Acids↗
PubMed1988

Purification of three cytosolic glutathione S-transferases from adult Schistosoma mansoni.

The cytosolic fraction of adult Schistosoma mansoni contains glutathione S-transferase (EC 2.5.1.18) activity, determined with the prototype substrate 1-chloro-2,4-dinitrobenzene, that is 5- to 50-fold greater than that found in other metazoan parasites. A survey of several model substrates revealed that enzymes in male and female schistosomes have distinct but overlapping substrate specificities. Four forms of glutathione S-transferase were detected, three of which, SmGST-1, SmGST-2, and SmGST-3, were purified to apparent homogeneity by glutathione affinity chromatography and HPLC chromatofocusing. The purified enzymes displayed very similar catalytic and physicochemical properties. They could be distinguished by differences in activity with ethacrynic acid and trans-4-phenyl-3-buten-2-one, but not with aryl halide substrates. The isoelectric points of SmGST-1, SmGST-2, and SmGST-3 were estimated to be 7.2, 7.1, 6.9, respectively. A polyclonal antiserum to SmGST-3 cross-reacted with the other two forms, but not with other soluble schistosome proteins. Each of the purified enzymes displayed an apparent subunit molecular weight of 28,500 by polyacrylamide gel electrophoresis under denaturing conditions. Gel filtration chromatography yielded a molecular weight of 30,800 for the catalytically active form of the enzyme. Unlike all known glutathione S-transferases, the three enzyme forms purified from S. mansoni appear to be catalytically active monomeric proteins.

Animals↗
PubMed1975

Asymmetric membrane expansion and modification of active and passive cation permeability of human red cells by the fluorescent probe 1-anilino-8-napththalene sulfonate.

1. The membrane perturbations induced by the interaction of the fluorescent probe 1-anilino-8-naphthalene sulfonate (ANS) with human red blood cells were studied. 2. ANS below 0.5 mM inhibits partially (20% maximum) the ouabain-insensitive Na+ and K+ influx and efflux. Above 0.5 mM ANS increases both Na+ and K+ leak fluxes. The increased cation leaks are larger for Na+ than K+. 3. The (Na+ +K+)-ATPase and ouabain-sensitive Na+ and K+ fluxes are inhibited by ANS. Ouabain-insensitive, Mg2+-dependent ATPase activity of ghosts is stimulated by [ANS] less than 0.3 mM and inhibited by [ANS] greater than 0.3 mM. 4. ANS also inhibits the Na+-dependent, ouabain-insensitive K+ influx that is inhibited by ethacrynic acid and furosemide. 5. Red cells become crenated with [ANS] less than 1 mM and sphere at [ANS] greater than 1 mM. In the former conditions hypotonic hemolysis is decreased whereas the latter increase osmotic fragility. 6. It is suggested that ANS expands the membrane asymmetrically by binding preferentially to the external membrane surface. 7. It is concluded that ANS is a general inhibitor of ion transport, particularly of those processes thought to involve facilitated-diffusion mechanisms. The increased cation leaks observed at high ANS concentrations may be related to prehemolytic membrane disruption. 8. The membrane perturbations caused by ANS are compared to those caused by other reversible inhibitors of anion exchange in red blood cells. Their possible modes of action are discussed.

Adenosine Triphosphatases↗
PubMed1977

Transport parameters and stoichiometry of active calcium ion extrusion in intact human red cells.

Ca2+-transport and its energy consumption were studied in intact human red cells loaded with Ca2+ by the aid of the ionophore A23187. After the complete elimination of the ionophore the passive Ca2+-permeability of the membrane returned to its normal low value, except when the intracellular Ca2+-concentration was higher than 3 mM or the ATP level fell below 100 muM. Within these limits the rate of Ca2+-extrusion was independent of the cellular ATP content but was greatly enhanced by increasing [Ca2+]i and reached a plateau at about 1 mM intracellular Ca2+-concentration. The maximum rate of Ca2+-efflux was about 85 mumol/l of cells per min at 37 degrees C, pH 7.4. The activation energy of active Ca2+-extrusion was found to be 15 200 cal/mol, and the optimum pH in the suspension was 7.7. Ca2+-efflux was not connected with the counter-transport of cations. The Ca2+-pump was not affected by ouabain or oligomycin and only partial inhibition could be achieved by the SH-reagents: ethacrynic acid, N-ethylmaleimide and p-chloromercuribenzoate or with propranolol and ruthenium red. An 80 to 95% inhibition of the active Ca2+-extrusion was brought about by 50-250 muM lanthanum, which in the above concentrations caused no aggregation or haemolysis. The inhibition of the Ca2+-pump by lanthanum was found to be reversible, the site of inhibition being at the external surface of the cell membrane. To examine the energy consumption of the Ca2+-extrusion, ATPase activity was assessed by measuring inorganic phosphate liberation in Ca2+-loaded red cells the metabolism of which was inhibited by iodoacetamide + Na+-tetrathionate. Ca2+-activated ATPase activity connected with the Ca2+-pump was distinguished from other Ca2+-ATPases by using the non-penetrating inhibitor, lanthanum. The molar ratio of Ca2+-transported per ATP split was found to be 2 : 1.

Adenine Nucleotides↗
PubMed1978

Selective alteration of erythrocyte deformabiliby by SH-reagents: evidence for an involvement of spectrin in membrane shear elasticity.

In order to elucidate the molecular basis of membrane shear elasticity, the effect of membrane protein modification by SH-reaents on the deformability of human erythrocytes was studied. Deformability was quuantified by measuring the elongation of erythrocytes subjected to viscometric flow in a transparent cone plate viscometer. Impermeable SH-reagents proved to have no mechanical effect. Many, but not all, permeable SH-reagents markedly decreased the elongation. Among these, bifunctional SH-reagents (e.g. diamide, tetrathionate and N, N' -p-phenylenedimaleimide) able to cross-link membrane SH-groups were more effective than monofunctional SH-reagents (e.g. N-ethylmaleimide and ethacrynic acid). The bifunctional SH-reagents produced a 50% decrease of elongation after modification of less than 5% of the membrane SH-groups. In contrast, for a comparable effect, more than 20% of the SH-groups had to be modified by the monofunctional reagents. The effect of SH-oxidizing agents was fully reversible after treatment with disulfide-reducing agents. All bifunctional SH-reagents induced a dimerization of a small fraction of spectrin. Anaalysis of the distribution of the diamide-induced disulfide bonds among the various membrane protein fractions showed that this agent preferentially acts on the spectrin polypeptides. The results provide direct experimental evidence that the native arrangement of spectrin is essential for the shear resistance of the erythrocyte membrane and that introduction of small numbers of intermolecular cross-links as well as modification within the molecule lead to a rapid loss of this function.

Diamide↗
PubMed1979

Stimulation of monovalent cation fluxes by electron donors in the human red cell membrane.

When human red cells are incubated at 37 degrees C with the artificial electron donor system ascorbate + phenazine methosulphate the fluxes of Rb+ (K+) through the cell membrane are increased. The effect of this donor system is much stronger in energy-depleted than in normal cells. The same effects are produced by HS-glutathione, NADH or NADPH loaded into resealed ghosts, but these electron donors were ineffective when added to the incubation medium. The Rb+ (K+) fluxes induced by electron donors resemble closely those induced by an increase of intracellular Ca2+ (Gardos effect). The electron donors require the presence of intracellular Ca2+ to be effective, but at levels that do not stimulate by themselves the fluxes of K+. Flavoenzyme inhibitors (atebrin and chlorpromazine), oligomycin and quinine prevented the effects of both electron donors and Ca2+ alone; antimycin, upcouplers and ethacrynic acid inhibited them partially; ouabain, furosemide, and rotenone had no effect. The results could be explained if the effect of electron donors is to bring about a change in the redox state of some membrane component(s) that makes intracellular Ca2+ more effective to elicit rapid K+ movements. Plasma membrane oxidoreductase activities could be engaged in this change.

Ascorbic Acid↗
PubMed1981

Occurrence of passive furosemide-sensitive transmembrane potassium transport in cultured cells.

Furosemide (1 x 10(-4) M) inhibits a proportion of the total passive (ouabain-insensitive) K+ influx into primary chick heart cell cultures (85%), BC3H1 cells (75%), MDCK cells (40%) and HeLa cells (57%). This action of furosemide upon K+ influx is independent of (Na+ + K+)-pump inhibition since the furosemide-sensitive component of the K+ influx is identical in the presence and absence of ouabain (1 x 10(-3) M). For HeLa cells the passive, furosemide-sensitive component of K+ influx is markedly dependent upon the external K+, Na+ and Cl- content. Acetate, iodide and nitrate are ineffective as substitutes for Cl-, whereas Br- is partially effective. Partial Cl- replacement by NO3- gave an apparent affinity of 100 mM [Cl]. Na+ replacement by choline+ abolishes the furosemide-sensitive component, whereas Li+ replacement reduces this component by 48%. Partial Na+ replacement by choline+ gives an apparent affinity of 25 mM [Na+]. Variation in the external K+ content gives an affinity for the furosemide-sensitive component of approx. 1.0 mM. Furosemide inhibition of the passive K+ influx is of high affinity, half-maximal inhibition being observed at 5 x 10(-6) M furosemide. Piretanide (1 x 10(-4) M) and phloretin (1 x 10(-4) M) inhibit the same component of passive K+ influx as furosemide; ethacrynic acid and amiloride (both 1 x 10(-4) M) partially so. The stilbene, SITS (1 x 10(-6) M), was ineffective as an inhibitor for the furosemide-sensitive component.

Animals↗
PubMed1992

Conductive chloride flux across amphibian skin: inhibition by indacrinone and cobalt ion.

When amphibian skin was incubated under conditions in which transepithelial sodium transport was abolished, a conductive transepithelial Cl- flux arose when Cl- was removed from one of the compartments. This flux was matched by short-circuit current and it accounted entirely for transepithelial conductance. Cl- influx was larger than efflux; it was linearly related to the magnitude of transepithelial Cl- concentration difference. When applied to the epithelial surface of the tissue, divalent metal cations such as Co2+, and the ethacrynic acid derivative, indacrinone, reduced rapidly and reversibly both transepithelial Cl- (in)flux and short-circuit current. Frog skin proved to be more sensitive to these inhibitors than toad skin. Further characterization of transepithelial Cl- pathway(s) should benefit from the fact that Cl- across amphibian skin can easily be monitored by the short-circuit current method, and from the availability of agents which inhibit this passive flux rapidly and reversibly.

Animals↗
PubMed1984

Identification of a new glutathione S-transferase from rat liver cytosol.

A new glutathione S-transferase has been purified to homogeneity from 105,000 X g supernatant of Sprague-Dawley rat liver homogenates. The purified enzyme exhibited specific activities of approximately 1.8, and 0.12 mumoles X min-1 X mg-1 toward 1-chloro 2,4-dinitrobenzene and cumene hydroperoxide respectively. The SDS gel electrophoresis data on subunit composition revealed that the new transferase is composed of two subunits with an identical Mr of 24,400 (Y alpha Family). Our in vitro translation experiments with rat liver poly(A) RNAs and substrate specificity data suggest that this subunit is different from the previously reported Ya , Yb and Yc subunits of rat liver glutathione S-transferases. Comparatively, the new isozyme showed significant activity toward 1,2 epoxy-3-(P-nitrophenoxy)-propane, ethacrynic acid and P-nitrophenyl acetate, 0.4, 0.34 and 0.18 mumoles. min-1 X mg-1 respectively.

Animals↗
PubMed1992

Comparison of the aflatoxin B1-8,9-epoxide conjugating activities of two bacterially expressed alpha class glutathione S-transferase isozymes from mouse and rat.

The complementary DNAs of rat glutathione S-transferase (GST, EC 2.5.1.18) Yc1 and of mouse Yc were expressed from a prokaryotic expression vector in E. coli. The purified proteins were analyzed for their activity toward aflatoxin B1-8,9-epoxide (AFBO), the reactive intermediate of the fungal mycotoxin aflatoxin B1 (AFB). The mouse Yc isozyme had about 50-fold higher conjugating activity toward AFBO than the rat Yc1 isozyme (144 nmol/mg/min versus 3.3 nmol/mg/min). The rat Yc1 isozyme had specific activities toward 1-chloro-2,4-dinitrobenzene, cumene hydroperoxide and ethacrynic acid of 10.7, 0.98 and 0.92 mumol/mg/min, respectively, whereas the mouse Yc isozyme had specific activities of 5.7, 2.1 and 0.1 mumol/mg/min for these substrates, respectively. These data provide further support for the hypothesis that the constitutive presence of the alpha class GST Yc isozyme in mouse liver protects mice from the hepatocarcinogenic effects of aflatoxin B1.

Aflatoxin B1↗
PubMed1992

Tyrosine-7 is an essential residue for the catalytic activity of human class PI glutathione S-transferase: chemical modification and site-directed mutagenesis studies.

The glutathione (GSH)-conjugating activity of human class Pi glutathione S-transferase (GST pi) toward 1-chloro-2,4-dinitrobenzene (CDNB) was significantly lowered by reaction with N-acetylimidazole, an O-acetylating reagent for tyrosine residues. Further, the replacement of Tyr7 in GST pi, which is conserved in all cytosolic GSTs, with phenylalanine by site-directed mutagenesis also lowered the activities toward CDNB and ethacrynic acid. The Km values of the mutant for both GSH and CDNB were almost equivalent to those of the wild type, while the Vmax of the former was about 55-fold smaller than that of the latter. Therefore, Tyr7 is considered to be an essential residue for the catalytic activity of GST pi.

Base Sequence↗
PubMed1983

Selenium and drug metabolism--I. Multiple modulations of mouse liver enzymes.

Male albino mice were raised on diets containing less than 10 ppb selenium (Se-) or supplemented with 0.5 ppm selenium (Se+) for 6 months. In the (Se-) group total liver selenium was less than 10% of the control, liver selenium-dependent glutathione peroxidase (GSH-Px) less than 2%. The specific activities of catalase and superoxide dismutase showed essentially no differences between the dietary groups. Several phase I-related specific enzyme activities were measured in liver microsomes. No significant differences between the two animal groups were found for cytochrome P-450 and b 5 content, NADH-cytochrome b 5 reductase, as well as for aniline hydroxylation and aminopyrine dealkylation rates. In (Se-) microsomes, NADPH-cytochrome P-450 reductase activity was about half that found in (Se+) microsomes. An increase in microsomes from (Se-) mice was found for 7-ethoxycoumarine deethylation rate (460%), cytochrome P-450 hydroperoxidase activity (170%), and heme oxygenase (276%). The N-oxidation rate of the flavin-containing monooxygenase decreased by 35%, the N-demethylation rate by 50% in (Se-) animals. Stopped-flow measurements of the reduction rates of microsomal pigments did not support evidence for limitations in microsomal electron supply during selenium deficiency. Among the phase II reactions examined, sulfotransferase activity towards 4-nitrophenol was 47% of the controls in Se-deficient liver cytosols while UDP-glucuronyl transferase activity towards this substrate increased to 215%. Glutathione-S-transferase activity was much higher in (Se-) livers than in (Se+): 310% with 1,2-dichloro-4-nitrobenzene, 255% with 1-chloro-2,4-dinitrobenzene and 120% with ethacrynic acid as substrate. The data indicate that in addition to GSH-Px many other enzyme activities in mouse liver are affected by prolonged dietary selenium deficiency. These effects might be useful in assessing the severity of selenium deficiency. A microsomal selenium-dependent metabolic modulator is discussed as a possible mechanism.

Cytochrome P-450 Enzyme System↗
PubMed1983

Human kidney thiopurine methyltransferase. Purification and biochemical properties.

Thiopurine methyltransferase (TPMT) catalyzes the S-methylation of thiopurines and thiopyrimidines. Human kidney TPMT was purified over 300-fold and its biochemical properties were determined. TPMT was "soluble" and had a molecular weight of approximately 36,000 daltons as estimated by gel filtration chromatography. The pH optimum of the purified TPMT was 6.7. "True" Km values for 6-mercaptopurine (6-MP) and S-adenosyl-L-methionine (SAM), the two cosubstrates for the reaction, were 0.30 mM and 2.7 microM respectively. "Apparent" Km values for 6-thioguanine and 2-thiouracil, two other methyl acceptor substrates, were 0.55 and 2.0 mM respectively. Aliphatic thiol compounds were either poor substrates for TPMT or were not methylated. S-Adenosyl-L-homocysteine was a competitive inhibitor of TPMT when the varied substrate was SAM, and 6-methylmercaptopurine was a noncompetitive inhibitor with respect to 6-MP. Purified TPMT was neither activated nor inhibited by 1 mM Ca2+ or Mg2+, but exposure to reagents such as N-ethylmaleimide and ethacrynic acid that interact with sulfhydryl groups inactivated the enzyme. Tropolone inhibited TPMT with a Ki of approximately 0.85 mM. Finally, human kidney TPMT activity could be distinguished from human kidney thiol methyltransferase (EC 2.1.1.9) activity on the basis of subcellular distribution, substrate specificity, kinetic characteristics and differential sensitivity to inhibitors.

Cations, Divalent↗
PubMed1984

Selenium and drug metabolism--II. Independence of glutathione peroxidase and reversibility of hepatic enzyme modulations in deficient mice.

Male mice were fed a diet containing less than 0.01 ppm selenium (Se-) for 6 months. A control group received the same diet containing 0.5 ppm selenium (Se+). In the livers of the Se- animals a drastic decrease in glutathione peroxidase (GSH-Px) activity was observed. It reached undetectable levels after 17 days of the Se- diet. At that time, GSH-transferase activity began to increase significantly, followed by changes in many other enzyme activities. After the 60th day, these enzyme modulations had reached a plateau with the following percentage changes compared to controls: GSH-transferases: 320% (1,2-dichloro-4-nitrobenzene), 218% (1-chloro-2,4-dinitrobenzene); glutathione reductase: 160%; ethoxycoumarin deethylase: 330%; cytochrome P-450-hydroperoxidase: 230%; heme oxygenase: 240%; UDP-glucuronyltransferase: 200%; GSH-thioltransferase: 64%; sulphotransferase: 62%; NADPH-cytochrome-P-450-reductase: 65%; flavin-containing mono-oxygenase: 57%. No significant changes were observed for GSH-transferase activity assayed with ethacrynic acid or for microsomal H2O2 formation and aniline hydroxylase activity. In single-pulse repletion experiments by injection of 250 micrograms selenium/kg body wt, different individual time constants for the recovery process of the enzymatic perturbations were observed. The half-times for the recovery ranged from 5.7 hr for the microsomal NADPH-cytochrome-P-450 reductase to over 29 hr for GSH-Px up to 44 hr for part of the GSH-transferase activity. 250 micrograms selenium/kg body wt were needed to restore 50% of GSH-Px activity in the long-term Se- mice compared to Se+ controls. All other enzymatic changes in the Se- mice needed a dose of 7 micrograms selenium/kg body wt for 50% restorage . The results demonstrate that processes other than those related to GSH-Px take place in a later phase of selenium deficiency in mouse liver with a chronologically common beginning. The different repletion and depletion kinetics as well as the different need of these processes for the trace element are discussed with respect to the existence of two separate selenium pools.

Animals↗
PubMed1985

Differential regulation of hepatic glutathione transferase and glutathione peroxidase activities in the rat.

The effects of the xenobiotics, i.e. butylated hydroxytoluene, beta-naphthoflavone, isosafrole, pregnenolone-16 alpha-carbonitrile, trans-stilbene oxide, 3-methylcholanthrene, phenobarbital, 3,3',4,4'-tetrachlorobiphenyl, 2,2',4,4',5,5'-hexachlorobiphenyl, on rat liver cytosolic glutathione transferase and glutathione peroxidase activities have been investigated. Although the glutathione transferase isozymes (measured by the specific substrates ethacrynic acid and delta 5-androstene-3,17-dione) which have been shown to possess peroxidase activity were significantly increased, little or no increase in peroxidase activity (toward cumene hydroperoxide, tert-butyl hydroperoxide or hydrogen peroxide) was observed. Likewise during a 16-day time course following the administration of Aroclor 1254 or fireMaster BP-6 (each 500 mg/kg, i.p.), potent induction of glutathione transferase activities was seen without any significant increases in peroxidase activities. In fact during the second week of the time course, there were significant decreases in selenium-dependent glutathione peroxidase activity (toward hydrogen peroxide). The inverse regulation of these activities, i.e. the depression of selenium-dependent glutathione peroxidase activity following sustained induction of glutathione transferases, may have direct implications for the toxicity of the polyhalogenated aromatic hydrocarbons.

Animals↗