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Biomedical subjects

W Levin

Publications and source records attributed to W Levin.

At least 145 records · Page 8Linked to original sources
PubMed1984

Non-invasive evaluation of ventricular function and volumes during atrioventricular sequential and ventricular pacing.

Thirteen patients who all had previously inserted temporary or permanent pacemakers (6, VVI; 7, A-V sequential) were studied by two-dimensional echocardiography and radionuclide gated blood pool ventriculography (RVG) for non-invasive evaluation of cardiac performance. Patients were paced in both the VVI mode and during sinus/atrial or A-V sequential pacing. Although there was no objective change of the ejection fraction during V-pacing and atrial/A-V sequential pacing or sinus rhythm, as has been previously reported, A-V sequential pacing did result in significant improvement in overall cardiac function and output as judged by radionuclide ventriculography and blood pressure response in most of our patients. An appropriately timed atrial contribution to ventricular systole resulted in improved ventricular function in those individuals with pre-existing systolic or diastolic myocardial dysfunction and/or sick sinus syndrome in whom pacemaker therapy was indicated. Radionuclide ventriculography appears to be a reliable, accurate, non-invasive method that can be used to evaluate patients before implantation of a permanent ventricular or atrioventricular pacemaker in order to decide which pacing mode is best for that particular individual.

Aged↗
PubMed1984

Mutagenicity of the enantiomers of the diastereomeric bay-region benzo(c)phenanthrene 3,4-diol-1,2-epoxides in bacterial and mammalian cells.

The mutagenic activities of the enantiomers of the pair of diastereomeric bay-region benzo(c)phenanthrene 3,4-diol-1,2-epoxides were evaluated in histidine-dependent strains of Salmonella typhimurium and in an 8-azaguanine-sensitive Chinese hamster cell line. In strains TA 98 and TA 100 of S. typhimurium, the range in mutagenic activity observed for the four optically active isomers was less than 4- and 2-fold, respectively. The diol-epoxide with (1S,2R,3R,4S) absolute configuration and the benzylic hydroxyl group trans to the epoxide oxygen [(+)-diol epoxide-2] was the most active isomer in both strains. The enantiomeric (-)-diol-epoxide-2 isomer, with (1R,2S,3S,4R) absolute configuration identical to that of the exceptionally tumorigenic (+)-diol-epoxide-2 isomers of benzo(a)pyrene, benz(a)anthracene, and chrysene, was the least active isomer in strain TA 98 (27%) and the second most active isomer in strain TA 100 (90%). In Chinese hamster V79 cells (-)-diol-epoxide-2 was the most active of the four benzo(c)phenanthrene isomers, and a 4- to 5-fold range in mutagenic activity was observed. The differences in mutagenic activity between the four bay-region diol-epoxide isomers of benzo(c)phenanthrene in the three test systems are relatively small when compared with results from similar studies with optically active bay-region diol-epoxide isomers of three other polycyclic aromatic hydrocarbons, and may be explicable, in part, by a tendency of the hydroxyl groups of benzo(c)phenanthrene diol-epoxides to adopt comparable pseudodiequatorial conformations.

Animals↗
PubMed1984

Tumorigenicity of dihydrodiols and diol-epoxides of benz[c]acridine in newborn mice.

The tumorigenicity of benz[c]acridine (B[c]ACR) and a number of its derivatives, including the five metabolically possible transdihydrodiols, the diastereomeric bay-region diol-epoxides, two non-bay-region diol-epoxides, and the K-region 5,6-oxide, were assessed in newborn mice. A total dose of 0.50 or 1.05 mumol of compound was administered i.p. to preweanling mice, and tumorigenic activity was determined when the mice were 33 to 37 weeks old. B[c]ACR was a weak carcinogen producing an average of 2.5 lung tumors/mouse and 0.15 liver tumor/male mouse at the 1.05-mumol dose. Of the five metabolically possible trans-dihydrodiols of B[c]ACR, only trans-3,4-dihydroxy-3,4-dihydro-B[c] ACR (B[c]ACR (B[c]ACR 3,4-dihydrodiol) had high tumorigenic activity. B[c]ACR 3,4-dihydrodiol induced 2- and 10-fold more pulmonary and hepatic tumors, respectively, than did the parent compound while the trans-1,2-, 5,6-, 8,9-, and 10,11-dihydrodiols had very little or no tumorigenic activity. Both of the diastereomeric bay-region 3,4-diol-1,2-epoxides, in which the epoxide oxygen is either cis (isomer 1) or trans (isomer 2) to the benzylic hydroxyl group, had tumorigenic activity. Isomer 2 was the most tumorigenic derivative tested, inducing at least 60, 7, and 12 times more lung tumors per mouse than did isomer 1, B[c]ACR 3,4-dihydrodiol and B[c]ACR, respectively. The K-region 5,6-oxide and two non-bay-region diol-epoxides (isomer 2 of B[c]ACR 8,9-diol-10,11-epoxide and B[c]ACR 10,11-diol-8,9-epoxide) were weakly active or inactive at the dose tested. The demonstration that B[c]ACR 3,4-diol-1,2-epoxide-2 is exceptionally tumorigenic and that its metabolic precursor, B[c]ACR 3,4-dihydrodiol, is more active than the parent hydrocarbon, B[c]ACR, support the concept that isomer 2 of the bay-region diol-epoxide may be an ultimate carcinogenic metabolite of B[c]ACR.

Acridines↗
PubMed1984

High stereoselectivity among the optical isomers of the diastereomeric bay-region diol-epoxides of benz(a)anthracene in the expression of tumorigenic activity in murine tumor models.

The tumorigenicity of the (+)- and (-)-enantiomers of the diastereomeric bay-region benz(a)anthracene 3,4-diol-1,2-epoxides was evaluated in two mouse tumor models. In an initiation-promotion experiment on mouse skin, a single topical application of 0.1 or 0.4 mumol of the benz(a)anthracene diol-epoxides was followed by 25 weeks of promotion with 12-O-tetradecanoylphorbol-13-acetate. Of the four isomers of the bay-region diol-epoxides, only (+)-[1R,2S,3S,4R]-3,4-dihydroxy-1,2-epoxy-1,2,3,4- tetrahydrobenz(a)anthracene [(+)-diol-epoxide-2] and (+)-[1R,2S,3S,4S]-3,4-dihydroxy-1,2-epoxy-1,2,3,4- tetrahydrobenz(a)anthracene [(+)-diol-epoxide-1] had significant tumor-initiating activity. (+)-Diol-epoxide-2 was approximately 4-fold more active as a tumor initiator on mouse skin than was (+)-diolepoxide-1 at both doses tested. In newborn mice, a total of 0.14 mumol of compound, divided into three doses, was administered i.p. on the first, eighth, and fifteenth day of life, and tumorigenic activity was determined when the mice were 26 to 32 weeks of age. As was observed in the initiation-promotion experiment on mouse skin, only two of the four optical isomers of the bay-region diol-epoxides produced a significant tumor incidence. (+)-Diol-epoxide-2 induced a 100% incidence of lung tumors, with an average of 23.11 tumors/mouse, and was at least 60-fold more active (average number of tumors per mouse) than was (+)-diol-epoxide-1, which produced a 31% lung tumor incidence and 0.38 lung tumors/mouse. (+)-Diol-epoxide-2 was the only optical isomer that induced a significant incidence of hepatic tumors in male mice (31% incidence, 1.17 tumors/mouse). The highly tumorigenic (+)-diol-epoxide-2 isomer with [R,S,S,R] absolute configuration has the same absolute configuration as does the highly tumorigenic isomer of the bay-region diol-epoxides of benzo(a)pyrene and chrysene.

Animals↗
PubMed1983

Regio- and stereoselective metabolism of two C19 steroids by five highly purified and reconstituted rat hepatic cytochrome P-450 isozymes.

High pressure liquid chromatographic systems capable of resolving at least 28 known and potential metabolites of 17 beta-hydroxy-4-androsten-3-one (testosterone) and 4-androstene-3,17-dione (androstenedione) were used to quantitatively assess the metabolism of the two steroids in monooxygenase systems reconstituted with five purified rat liver cytochrome P-450 isozymes. Cytochromes P-450a, -b, -c, -d, and -e catalyzed the oxidation of testosterone at overall rates of 21, 27, 2, 0.7, and 3 nmol/min/nmol of cytochrome P-450, respectively; while the corresponding rates for total androstenedione metabolism were 12, 62, 1.5, 0.3, and 5. Cytochrome P-450a catalyzed the oxidation of testosterone and androstenedione almost exclusively to their respective 7 alpha-hydroxy metabolites. Cytochrome P-450b catalyzed the oxidation of testosterone to androstenedione and 16 alpha- and 16 beta-hydroxytestosterone in approximately equal molar ratios. However, this same hemoprotein exhibited a marked stereoselectivity in the metabolism of androstenedione since the molar ratio of 16 alpha- and 16 beta-hydroxyandrostenedione was greater than 1:10. Cytochrome P-450e catalyzed the oxidation of both steroids to the same products as cytochrome P-450b, but at approximately 10% of the rate. Cytochromes P-450c and P-450d catalyzed the oxidation of testosterone and androstenedione regio- and stereospecifically to their respective 6 beta-hydroxy metabolites. These results indicate that certain cytochrome P-450 isozymes show marked positional specificity in the metabolism of both testosterone and androstenedione, and that the rate as well as stereoselectivity of the oxidative reactions can be markedly dependent on subtle differences in the structure of the steroid substrate.

Androstenedione↗
PubMed1983

Metabolism of benzo[c]phenanthrene by rat liver microsomes and by a purified monooxygenase system reconstituted with different isozymes of cytochrome P-450.

Metabolism of the environmental pollutant and weak carcinogen benzo[c]-phenanthrene (B[c]Ph) by rat liver microsomes and by a purified and reconstituted cytochrome P-450 system is examined. B[c]Ph proved to be one of the best polycyclic aromatic hydrocarbon substrates for rat liver microsomes. It is metabolized by microsomes from control rats and by rats treated with phenobarbital or 3-methylcholanthrene at 3.9, 4.2 and 7.8 nmol/nmol cytochrome P-450/min, respectively. Principal metabolites are dihydrodiols along with small amounts (less than 10%) of phenols. The K-region 5,6-dihydrodiol is the major metabolite and accounts for 77-89% of the total metabolites. The 3,4-dihydrodiol with a bay-region 1,2-double bond is formed in much smaller amounts and accounts for only 6-17% of the total metabolites, the highest percentage being formed by microsomes from control rats. Highly purified monooxygenase systems reconstituted with cytochrome P-450a, P-450b and P-450c and epoxide hydrolase form predominantly the 5,6-dihydrodiol (95-97% of total metabolites) and only a small percentage of the 3,4-dihydrodiol (3-5% of total metabolites). The 3,4-dihydrodiol is formed with higher enantiomeric purity by microsomes from 3-methylcholanthrene-treated rats (88%) than by microsomes from control rats (78%) or phenobarbital-treated rats (60%). In each case the (3R,4R)-enantiomer predominates. B[c]Ph 5,6-dihydrodiol formed by all three microsomal preparations is nearly racemic.

Animals↗
PubMed1983

Induction of two immunochemically related rat liver cytochrome P-450 isozymes, cytochromes P-450c and P-450d, by structurally diverse xenobiotics.

We have previously shown that purified rat liver cytochromes P-450c and P-450d share some but not all immunochemical determinants (Reik, L. M., Levin, W., Ryan, D.E., and Thomas, P.E. (1982) J. Biol. Chem. 257, 3950-3957). Antibody to cytochrome P-450d cross-reacts with cytochrome P-450c to form an immunoprecipitin band in the Ouchterlony test, but no detectable immunoprecipitin ring is formed in a radial immunodiffusion assay. However, the addition of purified cytochrome P-450c to purified cytochrome P-450d in the radial immunodiffusion assay alters the cytochrome P-450d standard curve. Appropriate corrections have been made for the interference of cytochrome P-450c in the immunoquantitation of cytochrome P-450d. Twelve structurally diverse xenobiotics have been examined for their capacity to modulate the levels of cytochrome P-450d, as well as cytochromes P-450a, P-450b, and P-450c, in rat liver microsomes. Five compounds (isosafrole, 3-methylcholanthrene, beta-naphthoflavone, 2,3,7,8-tetrachlorodibenzo-p-dioxin, and phenothiazine) and the polychlorinated biphenyl mixture Aroclor 1254 are potent in vivo inducers of cytochrome P-450d (0.44-0.89 nmol of cytochrome P-450d/mg of microsomal protein). Control rats have low levels of this microsomal hemoprotein (0.04-0.05 nmol of cytochrome P-450d/mg of microsomal protein). Isosafrole induces cytochrome P-450d to a greater extent than cytochrome P-450c, Aroclor 1254 induces both hemoproteins to similar extents, and the remaining four compounds preferentially induce cytochrome P-450c relative to cytochrome P-450d. All of these structurally diverse compounds induce cytochromes P-450d and P-450c, suggesting that the inducibility of these cytochrome P-450 isozymes, but not cytochromes P-450a and P-450b, is linked.

Animals↗
PubMed1983

The accumulation of distinct mRNAs for the immunochemically related cytochromes P-450c and P-450d in rat liver following 3-methylcholanthrene treatment.

Treatment of rats with 3-methylcholanthrene leads not only to a marked accumulation in the liver of translatable mRNA coding for a 56-kilodalton polypeptide representing cytochrome P-450c, the major 3-methylcholanthrene-induced cytochrome P-450 of rat liver, but also to the accumulation of comparable amounts of mRNA encoding a 52-kilodalton polypeptide which is immunoprecipitated with antibodies prepared against rat liver cytochrome P-450c. Further electrophoretic and immunochemical characterization of the latter translation product demonstrates that it corresponds to cytochrome P-450d, the major isosafrole-induced form of rat liver cytochrome P-450. The mRNAs for cytochromes P-450c and P-450d can be completely separated by electrophoresis in denaturing agarose gels and have chain lengths of approximately 4000 and 2000 nucleotides, respectively. These two mRNAs do not show detectable sequence homology to the mRNAs coding for the major phenobarbital-induced forms of cytochrome P-450 (P-450b and P-450e) since in Northern blotting experiments they fail to hybridize under conditions of low to moderate stringency to cloned probes for the latter mRNAs.

Animals↗
PubMed1983

On the glycosylation state of five rat hepatic microsomal cytochrome P-450 isozymes.

The glycosylation states of five rat hepatic microsomal cytochrome P-450 isozymes (cytochromes P-450a, P-450b, P-450c, P-450d, and P-450e) were examined by quantitative carbohydrate analysis. Carbohydrate content of the purified enzymes as determined by acid hydrolysis, reduction, and gas chromatography of the alditol acetates revealed only trace amounts of neutral and amino hexoses in each of the five isozymes. Levels of mannose ranged from 0.3 to 1.7 mol/mol of cytochrome P-450 whereas levels of galactose were less than or equal to 0.2 mol/mol of cytochrome P-450 for the five hemoproteins. The amino sugars glucosamine and galactosamine were usually present at levels less than or equal to 0.2 mol/mol of cytochrome P-450, although one preparation of cytochrome P-450b had as much as 0.5 mol of glucosamine/mol of cytochrome P-450. Other carbohydrate residues (xylose and arabinose) were not detected in significant quantities. Since N- and O-glycosylation of proteins occurs primarily through N-acetylglucosaminyl and N-acetylgalactosaminyl residues, respectively, the lack of significant amounts of these amino sugars indicates that these five cytochrome P-450 isozymes are not normally glycosylated in the native state. Purified NADPH-cytochrome c reductase, which functions as an electron donor for microsomal cytochrome P-450, contained no detectable quantities of hexose sugars.

Animals↗
PubMed1983

Distinct rat hepatic microsomal epoxide hydrolases catalyze the hydration of cholesterol 5,6 alpha-oxide and certain xenobiotic alkene and arene oxides.

Metabolism of cholesterol 5,6 alpha-oxide to the 5,6-glycol is catalyzed by a rat liver microsomal epoxide hydrolase that is distinct from the microsomal epoxide hydrolase that metabolizes a wide range of xenobiotic alkene and arene oxides. The two enzymes are antigenically distinct, and the purified microsomal epoxide hydrolase that metabolizes xenobiotic oxides does not catalyze the hydration of cholesterol 5,6 alpha-oxide. In vivo treatment of rats with inducers of microsomal epoxide hydrolase does not enhance the activity of cholesterol 5,6 alpha-oxide hydrolase and, in some cases, actually depresses enzyme activity in the resultant microsomal preparations. Octene 1,2-oxide and benz[a]anthracene 5,6-oxide, both good substrates for xenobiotic epoxide hydrolase, are not competitive inhibitors of cholesterol oxide hydration by rat liver microsomes. The above results establish the existence of a liver microsomal epoxide hydrolase that is under different regulatory control and that appears to have a different substrate specificity than the well-characterized microsomal epoxide hydrolase involved in the metabolism of a widely diverse group of alkene and arene oxides.

Alkenes↗
PubMed1983

Differential time course of induction of rat liver microsomal cytochrome P-450 isozymes and epoxide hydrolase by Aroclor 1254.

The time course of induction of rat liver microsomal cytochromes P-450a, P-450b + P-450e, P-450c, and P-450d and epoxide hydrolase has been determined in immature male rats administered a single large dose [1500 mumol (500 mg)/kg body wt] of the polychlorinated biphenyl mixture Aroclor 1254. Differential regulation of these xenobiotic-metabolizing enzymes was indicated by their characteristic patterns of induction. The rate of induction of cytochrome P-450a and epoxide hydrolase was relatively slow, and steady-state levels of these enzymes were maintained from approximately Days 9 to 15 after Aroclor 1254 treatment. In contrast, cytochrome P-450c was maximally induced 2 days after Aroclor 1254 treatment and remained at a constant level through Day 15. Steady-state levels of cytochrome P-450d, beginning 1 week after Aroclor 1254 treatment, were preceded by a fairly rapid rate of induction and possibly by a small decline from maximal levels observed around Days 4 to 5. Like those of the other cytochrome P-450 isozymes and epoxide hydrolase, the levels of cytochromes P-450b + P-450e were constant from Day 9 to 15 after Aroclor 1254 treatment. However, an unexpected but reproducible decline (approximately 25%) in total cytochrome P-450 content observed between Days 4 and 9 after Aroclor 1254 treatment principally reflected a dramatic and totally unanticipated decrease (approximately 45%) in the level of cytochromes P-450b + P-450e. This transient decline in the level of cytochromes P-450b + P-450e was not due to an unusual effect of a mixture of polychlorinated biphenyls, since identical results were obtained with two individual congeners, namely 2,3,4,5,4'-penta- and 2,3,4,5,3',4'-hexachlorobiphenyl, that induced the same isozymes as Aroclor 1254. In contrast, when rats were treated with 2,4,5,2',4',5'-hexachlorobiphenyl, which induces cytochromes P-450a and P-450b + P-450e and epoxide hydrolase but not cytochromes P-450c or P-450d, maximal levels of cytochromes P-450b + P-450e were attained on Day 4 and no decrease was observed over the next 11 days. These results suggest that there may be an interaction in the regulation of induction of certain individual cytochrome P-450 isozymes.

Animals↗
PubMed1983

The in vivo turnover of rat liver microsomal epoxide hydrolase and both the apoprotein and heme moieties of specific cytochrome P-450 isozymes.

The in vivo turnover rates of liver microsomal epoxide hydrolase and both the heme and apoprotein moieties of cytochromes P-450a, P-450b + P-450e, and P-450c have been determined by following the decay in specific radioactivity from 2 to 96 h after simultaneous injections of NaH14CO3 and 3H-labeled delta-aminolevulinic acid to Aroclor 1254-treated rats. Total liver microsomal protein was characterized by an apparent biphasic exponential decay in specific radioactivity, with half-lives of 5-9 and 82 h for the fast- and slow-phase components, respectively. Most (approximately 90%) of the rapidly turning over microsomal protein fraction was immunologically distinct from membrane-associated serum protein, and thus appeared to represent integral membrane proteins. The existence of two distinct populations of cytochrome P-450a was suggested by the apparent biphasic turnover of both the heme and apoprotein moieties of the holoenzyme. The half-lives of the apoprotein were estimated to be 12 and 52 h for the fast- and slow-phase components, respectively, and 7 and 34 h for the heme moiety. The turnover of cytochromes P-450b + P-450e was identical to that of cytochrome P-450c, with half-lives of 37 and 28 h for the apoprotein and heme moieties, respectively. In all cases, the shorter half-lives of the heme component compared to the protein component were statistically significant. In contrast to the cytochrome P-450 isozymes, epoxide hydrolase (t1/2 = 132 h) turned over slower than the "average" microsomal protein (t1/2 = 82 h). The differential rates of degradation of these major integral membrane proteins during both the rapid and slow phases of total microsomal protein turnover argue against the concepts of unit membrane degradation and unidirectional membrane flow of liver endoplasmic reticulum.

Animals↗
PubMed1983

The effect of steric strain in the bay-region of polycyclic aromatic hydrocarbons: tumorigenicity of alkyl-substituted benz[a]anthracenes.

3,6-Dimethylcholanthrene (3,6-DMC) and 7,11,12-trimethylbenz[a]anthrene (7,11,12-TMBA) were tested for tumor-initiating activity on mouse skin as an approach to evaluate the potential role of steric strain in the bay-region on tumorigenic potency. Methyl-substitution at the 6-position of 3-methylcholanthrene (3-MC) increases steric strain in the bay-region of the hydrocarbon as it does at the 12-position of benz[a]anthracene (BA) causing both hydrocarbons to become non-planar. 3,6-DMC had at least 2- to 3-fold higher tumor-initiating activity than did 3-MC. Introduction of an 11-methyl group in 7,12-dimethylbenz[a]anthracene (7,12-DMBA) results in the formation of a more highly hindered (buttressing effect) hydrocarbon. 7,11,12-TMBA had 5% or less of the tumor-initiating activity of 7,12-DMBA, although the hydrocarbon still had relatively high tumorigenic activity on mouse skin. The results obtained with 3,6-DMC and studies reported previously with other methyl-substituted hydrocarbons, show that hydrocarbons possessing steric strain in the bay-region of the molecule can have enhanced tumorigenic activity. The basis of this steric effect remains unclear, however, as a result of the decreased tumorigenic activity of the 11-methyl-substituted derivative of 7,12-DMBA. The weak tumor-initiating activity of BA was enhanced at least 4- to 8-fold as a result of methyl-substitution at the 6- and 8-positions (6,8-dimethylbenz[a]anthracene). The higher tumorigenic activity of 6,8-dimethylbenz[a]anthracene compared to BA is consistent with a presumed decrease in metabolic detoxification of the dimethyl-substituted derivative at the 5,6- and 8,9-double bonds.

9,10-Dimethyl-1,2-benzanthracene↗
PubMed1983

Identification and localization of amino acid substitutions between two phenobarbital-inducible rat hepatic microsomal cytochromes P-450 by micro sequence analyses.

Two isozymes of rat liver microsomal cytochrome P-450--P-450b and P-450e--were compared by micro sequence analyses of their NH2 termini and tryptic fragments. These two phenobarbital-inducible hemoproteins, which are immunochemically indistinguishable with antibody against cytochrome P-450b, have extensive sequence homology. Automated Edman degradation of the native proteins revealed identical amino acids for the first 35 residues. Sequence determinations of the tryptic peptides, which constitute approximately 75% of each protein molecule, have thus far shown 10 amino acid differences between the two isozymes. Results of our amino acid sequence analyses established that two of the cDNAs, pcP-450pb1 and pcP-450pb4, reported by Fujii-Kuriyama et al. [Fujii-Kuriyama, Y., Mizukami, Y., Kamajiri, K., Sogawa, K. & Muramatsu, M. (1982) Proc. Natl. Acad. Sci. USA 79, 2793-2797] encode cytochrome P-450b whereas pcP-450pb2, a third cDNA whose nucleotide sequence differed slightly from that of the other two (six amino acid substitutions), encodes cytochrome P-450e. In addition to establishing the identity of these cloned cDNAs we provide direct evidence for seven additional amino acid differences between cytochromes P-450b and P-450e that occur beyond the region (Arg358) encoded by the cloned cDNA for cytochrome P-450e. Together, the amino acid sequences determined by micro sequence analysis and recombinant DNA techniques reveal 13 amino acid differences between these two isozymes. This report highlights the complementary nature of two different molecular approaches to elucidation of the amino acid sequences of isozymes with extensive structural homology.

Amino Acid Sequence↗