Interaction of benzanthrone with cytochrome P450: altered patterns of hepatic xenobiotic metabolism in rats

Benzanthrone, an anthraquinone dye intermediate, is commonly used for the synthesis of a number of polycyclic vat and disperse dyes. Our prior studies have shown that benzanthrone can be metabolized by rat hepatic microsomal cytochrome P450 (P450) (Biochem. Int., 18, 1989, 1237). In this study, the interaction of benzanthrone with rat hepatic microsomal P-450 and its effect on xenobiotic metabolism have been investigated. Parenteral administration of benzanthrone (40 mg/kg body weight) for 3, 7, or 21 days caused no change in the relative body weight or organ weight of rats. The levels of P450 were found to be reduced (33%-50%) in all the benzanthrone-exposed animals at all the time periods. In vitro addition of benzanthrone caused a spectral change with oxidized P450 and concentration-dependent reduction in the carbon monoxide spectrum of dithionite-reduced P450. The addition of benzanthrone to hepatic microsomes prepared from phenobarbital-treated rats resulted in spectral changes characterized by an absorbance maximum at 397 nm indicative of type I binding. In vitro addition of benzanthrone showed a concentration-dependent inhibition of hepatic aminopyrine N-demethylase (APD) and ethoxyresorufin-O-deethylase (ERD) activities with respective I50 values of 9.5 x 10(-4) and 8.0 x 10(-5) M. However, the inhibition of aryl hydrocarbon hydroxylase (AHH) even at the highest concentration of benzanthrone (10(-2) M), was of the order of only 29%. In vivo administration of benzanthrone also led to the inhibition of APD, AHH, and ERD activities at all treatment times although the magnitude of inhibition was of a lower order.(ABSTRACT TRUNCATED AT 250 WORDS)     

Turnip yellow mosaic virus RNA is aminoacylated in vivo in Chinese cabbage leaves

Turnip yellow mosaic virus (TYMV) contains a tRNA-like structure as an integral part of its genome. This structure is located at the extreme 3' end of the viral RNA and is the acceptor of valine after 3'-terminal adenylation. It is known that in vitro (with bacterial, yeast, or plant systems) and in vivo (upon microinjection into Xenopus laevis oocytes) a series of tRNA-specific enzymes can recognize this structure in the viral RNA. We report that TYMV RNA is valylated and consequently adenylated in vivo in its natural host, Chinese cabbage leaves. This suggests that the acylated form of the viral RNA could play an important role in the life-cycle of the virus.     

Cyclosporin A as prophylaxis against graft-versus-host disease in 36 patients.

Oral cyclosporin A was used as prophylaxis against graft-versus-host disease in (a) 31 patients with acute leukaemia or aplastic anaemia given transplants of HLA-matched bone marrow and (b) five patients with inborn errors of metabolism given transplants of haplotype-identical (parental) bone marrow. Twenty-six patients survived longer than two months after the operation. Despite the cyclosporin A, 31 patients (86%) suffered an acute form of graft-versus-host disease and 22 (61%) a chronic form. Nevertheless, the disease was usually treatable with immunosuppressive agents and caused the death of only one patient. Cyclosporin A caused renal toxicity in all cases; occasionally this was associated with a "capillary leak" syndrome, fatal in two patients. In children hypertension, fits, and fluid retention were common side effects. Blood concentrations of cyclosporin A correlated with blood urea values and blood pressure but did not predict the occurrence of graft-versus-host disease. Four different dose schedules were used to find the optimum way to administer this drug. Oral cyclosporin A is extremely effective at reducing the severity of graft-versus-host disease, but prevention of the disease is limited by toxicity of the drug and variable absorption. Better results might be achieved with parenteral administration or by using the drug in combination with other methods.     

Isolation of a highly active H+-ATPase from beef heart mitochondria

The lysolecithin extraction procedure originally described by Sadleret al. (1974) has been modified to yield a H⁺-ATPase with high levels of P_i-ATP exchange activity (400–600 nmol × min^–1 × mg^–1). This activity is further enhanced (1400–1600 nmol × min^–1 × mg^–1) following sucrose density gradient centrifugation in the presence of asolectin. This enhancement results in part from a lipid-dependent activation and in part from removal of inactive complexes. The H⁺ translocating activity of the complex has been determined spectrophotometrically using binding of oxonol VI as an indicator of membrane potential. P_i-ATP exchange, ATP hydrolysis, and oxonol binding are sensitive to energy-transfer inhibitors (oligomycin, rutamycin) and/or uncouplers (DNP, FCCP).     

Regulation of rat hepatic stearoyl coenzyme A desaturase. The roles of insulin and carbohydrate.

The rat hepatic stearoyl-CoA desaturation decreased by 3.7-fold in streptozotocin-induced diabetes. Insulin treatment of diabetic rats increased the enzyme activity by 7-fold. In marked contrast to glucose administration, fructose feeding in diabetic rats resulted in 20-fold stimulation of stearoyl-CoA desaturation, although both carbohydrates stimulated stearoyl-CoA desaturation in normal rats. Measurement of the microsomal electron transfer components showed no significant changes in the NADH-cytochrome b5 reductase activity or in the concentration of cytochrome b5. However, the activity of the terminal desaturase changed in a parallel fashion as the amount of terminal desaturase reflect changes in the overall desaturation. Supplementation of various microsomes with the saturating amount of purified terminal desaturase resulted in the formation of similar amounts of catalytically active complex and increased the stearoyl-CoA desaturation to the same level suggesting that the changes in the amount of terminal desaturase reflect changes in the overall desaturation. The results support the suggestion that both insulin and the intermediates of carbohydrate metabolism are involved in the regulation of terminal desaturase.     

E.s.r. of spin-trapped radicals in aqueous solutions of peptides. Reactions of the hydroxyl radical.

The reactions of hydroxyl radicals with 30 dipeptides and several larger peptides were studied in aqueous solutions. The OH radicals were generated by U.V. photolysis of H2O2. The short-lived peptide radicals were spin-trapped using t-nitrosobutane and identified by e.s.r. For dipeptides containing the amino terminal residues glycine, alanine and phenylalanine, abstraction of the hydrogen from the carbon adjacent to the peptide nitrogen was the major process leading to the spin-adducts. Such radicals will be referred to as backbone radicals. Dipeptides with a carbonyl terminal serine residue and also glycylglutamic acid form both backbone and side-chain radicals, with the latter being formed in larger quantities. For dipeptides, side-chain radicals were detected on either the carboxyl or amino terminal residues of both. The effect of pD on the e.s.r. sectrum of the spin-adducts of glycylglycine was studied and the pK of the carboxyl group of this radical was determined to be 2.5. For (Ala)3 and (Ala)n, with an average value of n = 1800, backbone and minor side-chain radicals were observed. For ribonucleases-S-peptide, containing 20 amino acid residues, both backbone and side-chain radicals were detected.     

The Role of Fluoride in Montmorillonite-Catalyzed RNA Synthesis

The montmorillonite-catalyzed reactions of the 5′-phosphorimidazolide of adenosine in the presence of fluoride were investigated to complete our study on the effect of salts on this type of reaction. Both anions and cations have been found to influence the oligomerization reactions of the activated nucleotides, being used here as a model system for pre-biotic RNA synthesis. However, in total contrast to the behavior of the activated nucleotides in the presence of montmorillonite and other salts, alkali metal fluorides did not yield any detectable oligomerization products except in very dilute (<0.005 M) solutions of fluoride. Instead, 5′-phosphorofluoridates were formed. Their identity was confirmed by a combination of HPLC, mass spectrometry, synthesis, and NMR.     

Phosphocaveolin-1 is a mechanotransducer that induces caveola biogenesis via Egr1 transcriptional regulation

Maintenance of epithelial cell adhesion is crucial for epidermal morphogenesis and homeostasis and relies predominantly on the interaction of keratins with desmosomes. Although the importance of desmosomes to epidermal coherence and keratin organization is well established, the significance of keratins in desmosome organization has not been fully resolved. Here, we report that keratinocytes lacking all keratins show elevated, PKC-α–mediated desmoplakin phosphorylation and subsequent destabilization of desmosomes. We find that PKC-α activity is regulated by Rack1–keratin interaction. Without keratins, desmosomes assemble but are endocytosed at accelerated rates, rendering epithelial sheets highly susceptible to mechanical stress. Re-expression of the keratin pair K5/14, inhibition of PKC-α activity, or blocking of endocytosis reconstituted both desmosome localization at the plasma membrane and epithelial adhesion. Our findings identify a hitherto unknown mechanism by which keratins control intercellular adhesion, with potential implications for tumor invasion and keratinopathies, settings in which diminished cell adhesion facilitates tissue fragility and neoplastic growth.