Target organ-specific inactivation of drug metabolizing enzymes in kidney of hamsters treated with estradiol

Chronic treatment of hamsters with estradiol for several months has previously been shown to decrease the specific content of cytochrome P450 in the kidney, a target of hormonal carcinogenesis, but not in liver. The reason for this decrease in metabolic enzyme activity is unknown and has been examined in this investigation. We now report that the decrease in specific content of renal cytochrome P450 by 73% in response to estradiol was not affected by co-treatment with tamoxifen for 1 month. The subcutaneous infusion of 250 μg/day estradiol for 7 days lowered renal cytochrome P450 by 71% from control values and was therefore used for further mechanistic studies. This treatment decreased renal activities of estradiol 2- or 4-hydroxylase by 77 to 80%, of 7-ethoxycoumarin-O-deethylase by 66% of control values, respectively, and completely eliminated aryl hydrocarbon hydroxylase activities, whereas liver enzymes remained unaffected. After 7 days of infusion of estradiol, fluorescent products of lipid peroxidation were more than doubled in hamster kidney but remained unchanged in liver. The possibility of enzyme destruction by binding of estradiol 2,3-quinone to metabolizing enzymes was investigatedin vitro. In the presence of 2-hydroxyestradiol, cumene hydroperoxide, and microsomes, conditions known to favor the oxidation of the steroid to quinone, the binding of catechol estrogen metabolite to microsomal protein increased 60 fold over control values in the absence of cofactor. Purified rat liver cytochrome P450c also oxidized 2-hydroxyestradiol to 2,3-estradiol quinone. The rate of oxidation was linear for the first 2–3 min, but thereafter decreased with time. Under these incubation conditions, irreversible binding of catechol estrogen metabolite to cytochrome P450c increased for the first 2–3 min and then remained at this plateau level. It was concluded that enzyme destruction by a reactive estrogen metabolite or by lipid peroxides may be a major reason for the organ-specific decrease in cytochrome P450 enzymes in kidneys of estrogen-treated hamsters.     

Effects of α2- and β-adrenoceptor agonists on growth hormone secretion following lesion of the noradrenergic system of the rat

The aim of the present investigation was to lesion the noradrenergic system and to measure the effect on growth hormone (GH) secretion following peripheral administration of ₂- and -adrenoceptor agonists. Direct injection of these agonists into the paraventricular nucleus of the hypothalamus (PVN) and its effect on GH secretion were also investigated. Systemic administration of N-2-chloroethyl-N-ethyl-2-bromobenzylamine (DSP₄, 60 mg/kg, injected i.p. 10 days prior to experimentation) significantly decreased the noradrenaline (NA) content of the hippocampus, frontal cortex and hypothalamus but had no effect on the dopamine (DA) or serotonin (5-HT) content of these areas. Bilateral injection of 6-hydroxydopamine (6-OHDA, 10 g/l, 14 days prior to experimentation) into the medial forebrain bundle (MFB) caused a greater reduction of NA and also decreased the DA and 5-HT content of the hypothalamus. Analysis of the PVN of the hypothalami of rats following 6-OHDA lesion of the MFB showed significantly decreased NA and 5-HT content. Neither DSP₄ treatment nor 6-OHDA lesion of the MFB affected the clonidine (250 g/kg, i.p.) induced stimulation of GH secretion. Injection of isoproterenol (1 mg/kg, i.p.) had varying effects on GH secretion. It stimulated GH release in control rats but not in DSP₄ or MFB lesioned rats. Direct injection of clonidine (0.1 g/l) into the PVN significantly stimulated GH secretion, whereas injection of isoproterenol (2.5 g/l) into the PVN did not affect GH levels when compared to controls. The results of the present study do not support the hypothesis that hypoactivity of the central noradrenergic system may be the cause of the blunted GH response to clonidine observed in depressed patients.     

Translocation of photoassimilate from leaves of two popyploid genotypes of tall fescue differing in photosynthetic rates

The photosynthetic rate of a decaploid genotype (1-16-2) of tall fescue ( Festuca arundinacea Schreb.) is about twice that of a common hexaploid genotype (V6-802) (Plant Physiol. 72: 16–21, 1983). Translocation of photosynthate out of the leaves is a possible means of regulating carbon assimilation. To evaluate this possibility, we have examined a) translocation velocity, b) time course of translocation from leaves, c) photoassimilate partitioning pattern into whole plants in pulse and chase experiments, and d) interveinal distances between two ploidy genotypes. Most of the ¹⁴C accumulated in sucrose, and the labelled carbon moved down the leaf blades at similar velocities (6 to 10 cm h⁻¹) in both genotypes. Recent ¹⁴C assimilate was rapidly translocated from the fed area of the leaf blade. For example, the decaploid and the common hexaploid had translocated 40 and 26% of the ¹⁴C, respectively, at 6 h, and 79 and 49% of the ¹⁴C, respectively, at 24 h. Partitioning of ¹⁴C among plant organs was considerably different between the genotypes after a 24 h chase. For example, out of the total ¹⁴C recovered from the whole plant, the decaploid had retained 40% in the labelled leaf with 10, 33 and 29% in other leaves, stem bases and roots, respectively; whereas the hexaploid had retained 91% in the labelled leaf with 4, 3 and 2% in other leaves, stem bases and roots, respectively. However, the higher rate of translocation was correlated with greater interveinal distances in the decaploid genotype. These results suggested that the higher translocation percentage in the decaploid than the hexaploid genotype was due to greater sink activity.     

Mutagenicity of azo dyes in the Salmonella/microsome assay using in vitro and in vivo activation

The mutagenicity of 6 azo dyes, including direct black 38 (DB38), direct black 19 (DB19), direct brown 95 (DB95), solvent yellow 3 (SY3), trypan blue (TPB), and food black 2 (FB2), was examined in the Salmonella/microsome assay. The effect of chemical azo reduction (dithionite) and in vivo metabolism on the mutagenicity of the dyes was also studied. In vivo azo-dye metabolites were isolated from the urine of rats intubated with dyes by XAD-2 column chromatography. Urinary metabolites from all the treated animals, except animals treated with FB2, induced frame-shift mutations in strains TA1538 and TA98 in the presence of liver S9 activation. The control urine did not increase the incidence of revertants in strains TA1538 and TA98. Thus, XAD-2 chromatography can be used to isolate genotoxic metabolites from the urine of animals intubated with azo dyes.     

Effects of combined administration ofl-tryptophan and tricyclic antidepressants on α2- and β-adrenoceptors and monoamine levels in rat brain

In order to test whether co-administration of a serotonin precursor with antidepressant drugs could potentiate the effects of the antidepressants on monoamines or adrenoceptors in rat brain,l-tryptophan (20 mg/kg) was administered to rats daily for 7 or 15 days, either alone or in combination with desipramine (10 mg/ kg) or amitriptyline (10 mg/kg). After treatment withl-tryptophan for 7 days, increases were observed in rat hypothalamic and frontal cortex 5-hydroxy-3-indoleacetic acid levels as well as in hypothalamic dopamine and nucleus accumbens 3,4-dihydroxyphenylacetic acid levels. After 15 days, hippocampal -adrenoceptor density was found to be decreased. There was no evidence of potentiation of desipramine or amitriptyline action whenl-tryptophan was administered in combination with the antidepressants. On the contrary, the antidepressants appeared to interact withl-tryptophan to reduce its effects.     

Late enzymes of vindoline biosynthesis

From differentiated plants of Catharanthus roseus (L.) G. Don we have isolated a specific enzyme of the vindoline biosynthetic pathway catalysing the S-adenosylmethionine-dependent methylation of 11-O-demethyl-17-O-deacetyl-vindoline. The enzyme we named S-adenosyl-L-methionine : 11-O-demethyl-17-O-deacetylvindoline 11-O-methyltransferase. This transferase exhibits a high substrate specificity. Obviously the O-methylation at C-11 precedes the O-acetylation at the C-17 position during the biosynthesis of vindoline.A second enzyme was detected which hydrolyses the acetyl function of vindoline. The distribution of this acetylesterase in C. roseus plants demonstrates that the enzyme is not specifically associated with the vindoline distribution in the plant material. Most probably this enzyme plays no essential role in the biosynthesis of vindoline.     

Late enzymes of vindoline biosynthesis. Acetyl-CoA: 17-O-deacetylvindoline 17-O-acetyl-transferase

From differentiated plants of Catharanthus roseus (L.) G. Don, a specific enzyme was isolated and named acetyl-CoA : 17-O-deacetylvindoline 17-O-acetyltransferase, acting on the biosynthetic formation of the Aspidosperma type alkaloid vindoline.The enzyme shows a high selectivity towards different substrates. The acetyl-CoA-dependent transferase also catalyses the reverse reaction by hydrolysis of the 17-O-acetyl group of vindoline in the presence of free CoA. This enzyme is localized only in vindoline-containing plant parts, but was so far not detectable in cell suspension cultures of C. roseus. The enzyme allows the synthesis of labelled vindoline with high specific activity, applicable for instance as tracer for radioimmunoassays of vindoline.     

Relation between enzyme release and metabolic changes in reversible anoxic injury of myocardial cells

Cultured adult cardiac myocytes were exposed to anoxia under substrate-free conditions. When compared to the metabolic changes in the oxygen deficient organ, those in the anoxic cell culture proceed in a similar, yet prolonged manner. Release of cytosolic enzymes starts with minor energetic disturbances and proceeds in close correlation to the actual ATP decay. Below 2 μmol. ATP/g_WW, an increasing number of cells becomes irreversibly damaged, but above, 30 min reoxygenation leads to extensive recovery of the whole preparation. The results indicate that leakage of cytosolic enzymes during the early stage of anoxia is due to a gradual protein release from the individual cells, related to reversible membrane alterations.     

Expression of L3T4 molecules on Lyt-2+, class II-restricted antigen-specific T suppressor lymphocytes

Three bovine serum albumin-specific Lyt-2⁺ T suppressor (Ts) cell clones from CBA/J mice have been analyzed with regard to expression of L3T4 molecules. All three Ts-cell clones can be stained with monoclonal antibodies (mAb) to L3T4. Tested for the two clones restricted to recognition of E^k determinants, antigen-specific proliferation on antigen-presenting cells, but not the proliferation induced by conditioned medium can be inhibited by L314-specific mAb. In a similar way, Ts-cell cytolytic effector functions can be blocked by L3T4-specific mAb. Thus L3T4 structures seem to play a role in Ts-cell functions. Furthermore, the data support the view that L3T4 expression can be a property of class II-restricted T cells irrespective of their Lyt phenotype.