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Biomedical subjects

T Wolff

Publications and source records attributed to T Wolff.

At least 91 records · Page 5Linked to original sources
PubMed1990

Capability of human blood cells to form the DNA adduct, C8-(N2-aminofluorenyl)-deoxyguanosine-3'-5'-diphosphate from 2-aminofluorene.

Human blood cells, separated by Ficoll-Hypaque centrifugation, were tested for their ability to catalyze the formation of DNA adducts of 2-aminofluorene (AF), using the 32P-postlabeling procedure for adduct analysis. Incubation of neutrophils with AF, hydrogen peroxide and exogenous DNA yielded a single DNA adduct identified as C8-(N2-aminofluorenyl)-deoxyguanosine-3'-5'-diphosphate (AFdG) by cochromatography with a standard sample. AFdG levels in intact cells, lysed cells and in the granule fraction prepared from cell lysates were 102, 894 and 240 AFdG adducts/10(9) nucleotides/30 min respectively. AFdG levels corresponded to the activity of neutrophil peroxidase in these preparations. The monocyte/lymphocyte fraction yielded a low amount of 30 and 40 AFdG/10(9) nucleotides/30 min in the presence of hydrogen peroxide and of NADPH respectively. Erythrocytes did not generate a detectable level of AFdG, neither as intact cells nor as cell lysates. Whole blood samples likewise did not generate AFdG. Our findings reveal that, among blood cells, only neutrophils are capable of forming a biologically significant DNA adduct of aminofluorene in reasonable amounts and suggest that myeloperoxidase was the catalyzing enzyme.

Blood Cells↗
PubMed1989

Organic solvents as modifiers of aldrin epoxidase in reconstituted monooxygenase systems and in microsomes.

To examine the response of individual cytochrome P-450 species catalysing the epoxidation of aldrin (Wolff T and Guengerich FP, Biochem Pharmacol 36: 2581-2588, 1987), monooxygenase systems reconstituted from these species were assayed in the presence of 5% (v/v) = 0.87 M ethanol. The activity of cytochromes P-450PB-B and P-450PB-D, two enzymes inducible by phenobarbital was increased seven-fold. The activity of two other P-450 enzymes purified from these animals was either inhibited by 50%, as observed for cytochrome P-450PB-C or remained unchanged, as noted with cytochrome P-450PCN-E. Two P-450 enzymes purified from untreated rats, cytochromes P-450UT-F and P-450UT-H, showed an inhibition by 50 and 20%, respectively, while the activity of cytochrome P-450UT-A was slightly increased by 50%. Indirect evidence that solvents enhance aldrin epoxidation by interacting with the hemoprotein was obtained by the finding that ethanol stimulated the activity of cytochrome P-450PB-B already, before addition of the lipid component, L-alpha-1,2-dilauroyl-sn-glycero-3-phosphocholine. The Km of cytochrome P-450PB-B for NADPH cytochrome P-450 reductase was not altered by ethanol indicating that the interaction between the two enzymes was not affected by the solvent. Other results indicate that the stimulatory solvent binds to a site, apart from the type I or type II binding site. The potency of various hydrophylic solvents to modify aldrin epoxidase activity was assayed in microsomes of rats pretreated with phenobarbital and of untreated male rats. Ethanol, n-propranol, n-butanol, acetone and tetrahydrofuran enhanced enzyme activity of phenobarbital pretreated rats to a maximal extent of two-fold and, at similar concentrations, inhibited the enzyme activity of untreated rats by 50%. The potency of these solvents correlated with their lipophilicity. Methanol and dimethylsulfoxide only slightly modified the activity of induced and noninduced animals. In the presence of 0.5 M n-propranol as the modifying agent, microsomal epoxidase activity of rats pretreated with pregnenolone-16 alpha-carbonitrile, dexamethasone, 3-methylcholanthrene and of control rats was inhibited by 60-80%, whereas the activity of animals pretreated with phenobarbital, DDT, or the polychlorinated biphenyl mixture, Clophen A 50, was stimulated between two- and three-fold. The results reveal that organic solvents frequently used to dissolve monooxygenase substrates may considerably modify the activity of cytochrome P-450 dependent reactions, in particular when purified enzymes are assayed.

1-Propanol↗
PubMed1989

Ciprofloxacin-caffeine: a drug interaction established using in vivo and in vitro investigations.

The inhibitory effects of ciprofloxacin and other quinolone derivatives on the hepatic cytochrome P450-dependent metabolism of caffeine have been investigated in humans. In vivo studies involved an intraindividual comparison of the single-dose kinetics of caffeine before and during quinolone administration in 12 healthy men. Changes of enzymatic caffeine degradation by the quinolones were studied in vitro using human liver microsomes from three donors. Enoxacin and pipemidic acid markedly prolonged caffeine elimination in vivo. A positive correlation exists between the doses of enoxacin or ciprofloxacin and the prolongation (increases) in the caffeine elimination half-life. Decreases in caffeine elimination, using doses of ciprofloxacin in the upper part of the recommended dose range, were approximately 1.5-fold in comparison with untreated control subjects, whereas in the case of enoxacin there was a sixfold change. In vitro results with enoxacin, ofloxacin, ciprofloxacin, and pipemidic acid show a competitive inhibition (Dixon plots) of caffeine 3-demethylation. Ciprofloxacin and enoxacin showed the strongest inhibitory effects in vitro, whereas ofloxacin had the lowest inhibitory effect. These results are qualitatively reflected in the in vivo results; however, the clinical effects may be dependent on pharmacokinetic disposition of the quinolone and this could explain the weak inhibitory action of ciprofloxacin in vivo.

Adult↗
PubMed1989

Organization and decentralization of the Malawi National Bilharzia Control Programme.

The Bilharzia Control Project, supported by GTZ in collaboration with the Ministry of Health, began in late 1981 with two pilot areas, namely Nkoma and Pirimiti. The aims of the programme are to decrease the level of morbidity due to schistosomiasis through health facilities, chemotherapy, sanitation and health education. The glass sandwich diagnostic technique is used for the diagnosis of S. mansoni infection. The Nytrel urine syringe filtration technique, with Lugol's solution to stain eggs, and urine test strips are used for diagnosis of S. haematobium infection. Chemotherapy using the "test and treat" (TAT) approach, is carried out in a community, whether at a school or in a village. It is possible to treat between 100 and 300 people per day for S. mansoni infection using this approach with three bilharzia assistants (health surveillance assistants). This is also possible in S. haematobium areas. The "test and treat" teams can spend more than 3 weeks in a village and can treat up to 300 persons per day (using a microscope). Mapping and census of a village is done on the afternoon before the day of treatment. Health education talks are conducted on this same afternoon. Health education is to be the number one priority in the schistosomiasis control programme in Malawi. The role of man in the cycle of the infection will have to be emphasized in the informational component of schistosomiasis control. The Bilharzia Control Programme needs to join forces with other ministries or organizations through the primary health care approach, to motivate and mobilize the communities in its control activities.

Animals↗
PubMed1988

Analgesic treatment in acute myocardial infarction. A double-blind comparison of ketobemidone + the spasmolytic A29 (Ketogan) and morphine.

The analgesic effect of ketobemidone hydrochloride + the spasmolytic component A29 (Ketogan) and morphine hydrochloride was compared double-blindly in patients with suspect acute myocardial infarction. The test drugs were administered i.v. in an initial dose of 0.5 ml (2.5 mg Ketogan, 5 mg morphine) followed, if necessary, by additional injections of 0.25 ml. Altogether, 309 patients participated in the trial. The total consumption of the test drugs showed that 5 mg Ketogan was equipotent with 10 mg of morphine. Treatment with Ketogan resulted in a significantly higher proportion of patients who were completely free of pain 15 and 30 min after the initial injection: 16% and 15% more, respectively, compared to morphine. Within 2 hours after the initial injection, approximately 15% of the patients in both treatment groups had reported nausea and about 7% had vomited (patients who vomited or were nauseated before treatment were not included in this analysis). The frequency of other side-effects was low, with no differences between the two treatment groups. Morphine caused a greater reduction of the systolic blood pressure than Ketogan. The effect of both drugs on pulse rate and respiration was the same.

Adult↗
PubMed1988

Localization of 99mTc-diphosphonates in newly formed bone matrix as a measure of bone lesion detectability.

The lesion-to-normal-bone ratios of DBA-MDP (dibutylamino-methylene-diphosphonate), DPD (dicarboxypropane-diphosphonate) and MDP (methylene-diphosphonate) each labeled with 99mTc, were evaluated in experimental bone lesions. In 3-day old lesions this ratio was increased twofold for DBA-MDP in comparison with MDP and DPD which showed nearly equal ratios. Later on these differences became negligibly small. It is concluded that 99mTc-DBA-MDP is fixed more strongly in the immature bone matrix and that this will lead to an improvement in the detectability of lesions containing larger amounts of immature bone matrix.

Animals↗
PubMed1987

Rat liver cytochrome P-450 isozymes as catalysts of aldrin epoxidation in reconstituted monooxygenase systems and microsomes.

To explore which rat liver cytochrome P-450 species are involved in aldrin epoxidation, we have studied the catalytic activities of a series of cytochrome P-450 isozymes purified from untreated and inducer-treated Sprague-Dawley rats. Of ten cytochrome P-450 forms analyzed, seven isozymes, listed in order of decreasing activity, catalyzed aldrin epoxidation: P-450UT-A, P-450PB-C, P-450UT-H, P-450PB-B, P-450PCN-E, P-450UT-F, and P-450PB-D. P-450UT-I, P-450BNF-B, and P-450ISF-G were not very active at all. A novel aldrin metabolite, endo-dieldrin, was formed by cytochrome P-450UT-F in a 6-fold excess over dieldrin, which is the exo-isomer. The activity of aldrin epoxidase furthermore was assayed in liver microsomes from Sprague-Dawley rats of diverse physiological status and after pretreatment with various inducers resulting in a peculiar pattern of cytochrome P-450 isozymes. Untreated animals, at an age of 3 weeks, showed similar enzyme activities in both genders. During maturation, the activity of males increased by 3-fold, while the activity in females did not significantly change during this period. Pretreatment with pregnenolone-16-alpha-carbonitrile or dexamethasone strongly increased the activity in females. Pretreatment with dexamethasone did not increase the activity of males. A 50% depression of epoxidase activity was noted for males pretreated with 5,6-benzoflavone. Phenobarbital pretreatment increased the activity of females by 12-fold and of males by 2-fold. Males responded to pretreatment with polychlorinated biphenyls in a strain dependent fashion: enzyme activity was increased 2-fold in Sprague-Dawley rats but was not altered in Wistar rats. "Theoretical" values of microsomal epoxidase activity were calculated for weanling and adult Sprague-Dawley rats from turnover numbers and published data on the relative abundance of aldrin epoxidizing P-450 isozymes (Waxmann et al., Biochemistry 24, 4409, 1985). These values agreed with the activities determined. A similar statement can be made for male rats of both strains pretreated with inducers, when the ratio of enzyme activity of pretreated to control animals was used as a basis of comparison. The activity ratio of females pretreated with pregnenolone-16-alpha-carbonitrile, dexamethasone and phenobarbital, however, was much higher than the ratio calculated. Our results reveal that aldrin epoxidation is a reaction indicative of male specific and of phenobarbital-inducible cytochrome P-450 isozymes in rat liver.(ABSTRACT TRUNCATED AT 400 WORDS)

Aldrin↗
PubMed1987

Human liver debrisoquine 4-hydroxylase: test for specificity toward various monooxygenase substrates and model of the active site.

Polyclonal antibodies raised toward a debrisoquine 4-hydroxylating cytochrome P-450 species purified from rat liver (P-450UT-A) were used to determine which monooxygenase reactions are linked to debrisoquine hydroxylation in human liver. Anti P-450UT-A did not inhibit the oxidation of dimethylnitrosamine, morphine, diazepam, vinylidene chloride, trichloroethylene, benzo(a)pyrene and its 7.8-dihydrodiol, but was inhibitory for the hydroxylation of debrisoquine, (+/-)-bufuralol, lasiocarpine and monocrotaline. A model interpreting the substrate specificity of the human liver enzyme is presented.

Animals↗
PubMed1986

Characterization of rat and human liver microsomal cytochrome P-450 forms involved in nifedipine oxidation, a prototype for genetic polymorphism in oxidative drug metabolism.

The metabolism of the dihydropyridine calcium antagonist and vasodilator nifedipine has been reported to exhibit polymorphism among individual humans (Kleinbloesem, C. H., van Brummelen, P., Faber, H., Danhof, M., Vermeulen, N. P. E., and Breimer, D.D. (1984) Biochem. Pharmacol. 33, 3721-3724). Nifedipine oxidation has been shown to be catalyzed by cytochrome P-450 (P-450) enzymes. Reconstitution, immunoinhibition, and induction studies with rat liver indicated that the forms designated P-450UT-A and P-450PCN-E are the major contributors to microsomal nifedipine oxidation. The P-450 which oxidizes nifedipine (P-450NF) was purified to electrophoretic homogeneity from several human liver samples. Antibodies raised to P-450NF were highly specific as judged by immunoblotting analysis and inhibited greater than 90% of the nifedipine oxidase activity in human liver microsomes. A monoclonal antibody raised to the human P-450 preparation reacted with both human P-450NF and rat P-450PCN-E. Immunoblotting analysis of 39 human liver microsomal samples using anti-P-450NF antibodies revealed the same 52,000-dalton polypeptide, corresponding to P-450NF, with only one of the microsomal samples showing an additional immunoreactive protein. The level of nifedipine oxidase activity was highly correlated with the amount of P-450NF thus detected using either polyclonal (r = 0.78) or monoclonal (r = 0.65) antibodies, suggesting that the amount of the P-450NF polypeptide may be a major factor in influencing the level of catalytic activity in humans as well as rats. Cytochrome b5 enhanced the catalytic activity of reconstituted P-450NF, and anti-cytochrome b5 inhibited nifedipine oxidase activity in human liver microsomes. P-450NF also appears to be a major contributor to human liver microsomal aldrin epoxidation, d-benzphetamine N-demethylation, 17 beta-estradiol 2- and 4-hydroxylation, and testosterone 6 beta-hydroxylation, the major pathway for oxidation of this androgen in human liver microsomes.

Animals↗
PubMed1986

Human-liver cytochromes P-450 involved in polymorphisms of drug oxidation.

Nine forms of cytochrome P-450 have been purified to electrophoretic homogeneity from human-liver microsomes. These include the enzymes involved in debrisoquine 4-hydroxylation, phenacetin O-deethylation and mephenytoin 4-hydroxylation, three reactions which are characterized by genetic polymorphism in humans. Evidence for the involvement of the above enzymes comes from reconstituted immunochemical inhibition studies with human-liver microsomes. These and other lines of evidence are consonant with the view that different forms of cytochrome P-450 are involved in the three reactions. The debrisoquine 4-hydroxylase has been studied most extensively in terms of its substrate specificity. In addition, an analogous rat enzyme shows some homology and serves as a useful model. The use of antibodies raised to the rat-liver enzyme in immuno-inhibition studies with human-liver microsomes provides a means of determining the extent to which this enzyme participates in other reactions. Translation of rat-liver mRNA in vitro yields the intact debrisoquine 4-hydroxylase; studies with human mRNA suggest a lower frequency than in rats. The basis for impaired catalytic activity in phenotypically poor human metabolizers appears to be an altered enzyme in all three cases, as opposed to a decreased level of a single enzyme. Using antibody screening of fusion proteins expressed in a cDNA library, it has been possible to isolate cDNA probes for all three of these cytochromes P-450 for use in screening individuals and ultimately determining the basis of these polymorphisms.

Binding Sites↗
PubMed1985

Lack of relationship between debrisoquine 4-hydroxylation and other cytochrome P-450 dependent reactions in rat and human liver.

The effect of various inhibitors and inducers of cytochrome P-450 on the activity of microsomal debrisoquine 4-hydroxylation (DQH) was investigated in rat liver. DQH was strongly inhibited by SKF 525-A, 7,8-benzoflavone, and metyrapone. Pretreatment of animals with common inducers, such as phenobarbital, 3-methylcholanthrene, the commercial PCB mixture, Clophen A-50, dexamethasone and pregenenolone-16 alpha-carbonitrile did not lead to induction of DQH, while most reference reactions, i.e. aldrin epoxidation, ethylmorphine demethylation, and benzo(a)-pyrene hydroxylation were induced under these conditions. DQH likewise was not induced by pretreatment of animals with the enzyme substrate DQ. The relationship between DQH and other cytochrome P-450 functions was also studied in untreated animals. Both genders of Wistar rats exhibited similar rates of DQH, but different activities of the reference reactions. DQH activity in Wistar females however, was 10 times higher than in females of the DA-strain, whereas the reference activities were similar in both strains. The studies were also extended to human liver. DQH activity in homogenates of various biopsy samples was neither correlated to the activity of aldrin epoxidation nor to AHH activity. The results indicate that DQH in the rat and in man reflects the activity of a cytochrome P-450 species not related to various other known cytochrome P-450 functions.

Animals↗
PubMed1985

Dealkylation of pentoxyresorufin: a rapid and sensitive assay for measuring induction of cytochrome(s) P-450 by phenobarbital and other xenobiotics in the rat.

The O-dealkylation of pentoxyresorufin (7-pentoxyphenoxazone) by rat liver microsomes was examined. The reaction appeared highly specific for certain phenobarbital inducible forms of cytochrome P-450 and was increased 95- to 140-fold by animal pretreatment with phenobarbital (75 mg/kg/day, four ip injections) and approximately 50-fold by Aroclor 1254 (500 mg/kg, one ip injection) while animal pretreatment with 3-methylcholanthrene (50 mg/kg/day, three ip injections) resulted in less than a 2-fold increase over the rate detected in control microsomes. It was observed that this activity, in microsomes for Aroclor-pretreated rats, was dependent on O2 and was inhibited by metyrapone and SKF 525-A, indicative of cytochrome(s) P-450 mediation in the reaction. When antibodies directed against purified cytochrome(s) P-450s were employed to inhibit the pentoxyresorufin O-dealkylation reaction, antibodies to P-450PB-B greatly inhibited the reaction (greater than 90%), while antibodies to P-450PB-C or P-450PB/PCN-E had minimal effects. Assay of hepatic microsomes from rats which were pretreated with varying doses of phenobarbital (0.9-75 mg/kg/day, four ip injections) indicated that while aminopyrine-N-demethylase activity was induced only 2-fold at the maximum dose (75 mg/kg/day), pentoxyresorufin O-dealkylase activity was induced approximately 140-fold at this dose and approximately 4-fold by a dose of phenobarbital as low as 0.9 mg/kg.

Aminopyrine N-Demethylase↗
PubMed1985

Infectious complications of percutaneously inserted central venous catheters.

The frequency of early and late complications of percutaneously inserted central venous catheters (CVC) was prospectively assessed. One hundred and fifty-one periods of catheterization (siliconized and heparinized polyethylene and teflon catheters), inserted mainly infraclavicularly and on the right side, were evaluated. Strict aseptic technique was used during insertion and daily care of the CVC. The only early complications noted were three cases of pneumothorax. No nerve injuries or arterial punctures were observed. Of 124 evaluated cases (96 arm-phlebography, 28 post-mortem) 16% developed thrombosis, and in one patient this probably resulted in fatal pulmonary embolism. Septicaemia was seldom seen: two cases were considered as proved and another two cases as probable. This study and other studies indicate that an aseptic technique is essential and perhaps even more important than using tunnelled versus percutaneously inserted catheters in respect of reducing infectious complications of central venous catheters.

Adult↗