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PubMed1987

Inactivation of the maize transposable element Activator (Ac) is associated with its DNA modification.

The Activator (Ac) element at the waxy locus (wx-m7 allele) has the ability to undergo changes in its genetic activity and cycles between an active and inactive phase. Comparison of active Ac elements at several loci and the inactive Ac at wx-m7 by Southern blot analysis revealed that the inactive Ac sequence was not susceptible to digestion by the methylation sensitive enzyme PvuII while active elements were susceptible to PvuII digestion. Restriction digest comparisons between the clones of the active and inactive Ac elements were indistinguishable. Further analyses with the enzymes SstII and the methylation sensitive and insensitive isoschizomers EcoRII and BstNI showed the inactive Ac sequence was methylated at these sites, whereas the active Ac was hypomethylated. Although the active Ac at the wx-m7 allele in different genetic backgrounds showed differences in the Ac DNA modification pattern, at least a fraction of genomic DNA contained Ac sequences that were unmethylated at all of the internal sites we assayed. These data may suggest a role for DNA modification in the ability of Ac to transpose from the waxy locus and to destabilize unlinked Ds elements.

Alleles↗
PubMed1997

Hydroxyl radical-induced decline in motility and increase in lipid peroxidation and DNA modification in human sperm.

We employed the xanthine-xanthine oxidase system to produce H2O2 or simply used commercially available H2O2 solution to investigate the effects of exogenous hydroxyl radicals on the motility characteristics and on lipid peroxidation and DNA modification of human sperm. The functional parameters of sperm motility declined concomitantly upon incubation of sperm with hydroxyl radicals. After incubation of freshly ejaculated human sperm with 0.23 mM H2O2 in the presence of 1.8 mM ADP and 2.7 mM FeSO4 for 1 hr at 37 degrees C, 90% reduction of motility was observed. Effect of hydroxyl radicals on sperm motility was dependent on the concentrations of FeSO4 and H2O2, respectively. The remaining motility of sperm after 1 hr incubation showed negative linear correlation with FeSO4 concentration. The response of sperm motility to FeSO4 was also dependent on the concentration of H2O2. Except for the amplitude of lateral head displacement, functional parameters of sperm declined with the increase of H2O2 concentration. Moreover, we found that lipid peroxidation measured as malondialdehyde (MDA) and accumulation of modified DNA indicated by 8-hydroxy-2'-deoxyguanosine (8-OH-dG) in human sperm were significantly accelerated by exogenous hydroxyl radicals. The contents of lipid peroxides and 8-OH-dG in the spermatozoa were increased from 24.6 +/- 2.4 nmol MDA/1 x 10(7) sperm and 0.17 +/- 0.02% in the untreated group to 30.6 +/- 1.2 nmol MDA/1 x 10(7) sperm and 1.9 +/- 0.47%, respectively, in the sperm treated at 37 degrees C for 1 hr with 2.03 mM H2O2, 1.8 mM ADP and 4.5 mM FeSO4. Taken together, these results suggest that the detrimental effects of hydroxyl radicals on human sperm functions may be mediated, at least partly, through lipid peroxidation and DNA modification.

8-Hydroxy-2'-Deoxyguanosine↗
PubMed1986

DNA modification of a maize transposable element correlates with loss of activity.

An unstable allele of the bronze 2 (bz2) locus was isolated from a Robertson's Mutator Zea mays line containing a family of active transposable Mu elements. This mutation is somatically unstable, resulting in numerous revertant purple sectors on a bronze kernel. By following the variegated kernel phenotype through two generations, several lineages have been identified that have a distorted transmission of the mutant phenotype (fewer variegated kernels are produced than expected). Southern blot analysis of Mu elements in these plants demonstrates a correlation between an inhibition of digestion of Mu elements by certain restriction enzymes and the loss of somatic reversion at the mutant allele. The DNA modification can occur in all the Mu elements in a plant within one generation; however, plants have been identified that contain both modified and unmodified elements, suggesting that the modification can occur in a progressive manner. We hypothesize that the DNA modification results in nonfunctional elements.

5-Methylcytosine↗
PubMed1989

Single cell analysis of DNA modifications induced by chemical carcinogens and cytostatic drugs.

Antibodies recognizing specific DNA modifications allow the immunocytochemical visualization and quantification of these modifications at the level of the individual cell. Thus, the formation and repair of DNA adducts induced by chemical mutagens and carcinogens and by cytostatic drugs can be studied in very small samples in relation to e.g. cell type and tissue localization. A number of existing and potential applications in the fields of chemical carcinogenesis, chemical mutagenesis, experimental chemotherapy and molecular epidemiology are illustrated.

Animals↗
PubMed1984

Possible DNA modification in GC dinucleotides of Trypanosoma brucei telomeric sequences; relationship with antigen gene transcription.

Polymorphism in restriction site cleavage (PstI, SphI, PvuII, HindIII) has been noticed in several occasions in the telomeric sequences harbouring trypanosome variant-specific antigen genes (1, 2, 3). This polymorphism has been further investigated and seems best interpreted as due to partial DNA modification in GC dinucleotides. The actively transcribed telomeric genes do not exhibit such a polymorphism; furthermore, in at least three independent cases, gene inactivation is linked to the appearance of polymorphism. It could thus be hypothesized that DNA modification prevents antigen gene transcription, or vice-versa. We report however that at least some telomeric antigen-specific sequences of the procyclic trypanosomes (in vitro culture form) are not polymorphic, although they do not synthesize any variant-specific antigen mRNA. There is thus no absolute relationship between the absence of polymorphism and antigen gene transcription.

Animals↗
PubMed1990

Specificity of restriction endonucleases and DNA modification methyltransferases a review (Edition 3).

The properties and sources of all known class-I, class-II and class-III restriction endonucleases (ENases) and DNA modification methyltransferases (MTases) are listed and newly subclassified according to their sequence specificity. In addition, the enzymes are distinguished in a novel manner according to sequence specificity, cleavage position and methylation sensitivity. Furthermore, new nomenclature rules are proposed for unambiguously defined enzyme names. In the various Tables, the enzymes are cross-indexed alphabetically according to their names (Table I), classified according to their recognition sequence homologies (Table II), and characterized within Table II by the cleavage and methylation positions, the number of recognition sites on the DNA of the bacteriophages lambda, phi X174, and M13mp7, the viruses Ad2 and SV40, the plasmids pBR322 and pBR328, and the microorganisms from which they originate. Other tabulated properties of the ENases include relaxed specificities (integrated within Table II), the structure of the generated fragment ends (Table III), interconversion of restriction sites (Table IV) and the sensitivity to different kinds of DNA methylation (Table V). Table VI shows the influence of class-II MTases on the activity of class-II ENases with at least partially overlapping recognition sequences. Table VII lists all class-II restriction endonucleases and MTases which are commercially available. The information given in Table V focuses on the influence of methylation of the recognition sequences on the activity of ENases. This information might be useful for the design of cloning experiments especially in Escherichia coli containing M.EcodamI and M.EcodcmI [H16, M21, U3] or for studying the level and distribution of site-specific methylation in cellular DNA, e.g., 5'- (M)CpG-3' in mammals, 5'-(M)CpNpG-3' in plants or 5'-GpA(M)pTpC-3' in enterobacteria [B29, E4, M30, V4, V13, W24]. In Table IV a cross index for the interconversion of two- and four-nt 5'-protruding ends into new recognition sequences is complied. This was obtained by the fill-in reaction with the Klenow (large) fragment of the E. coli DNA polymerase I (PolIk), or additional nuclease S1 treatment followed by ligation of the modified fragment termini [P3]. Interconversion of restriction sites generates novel cloning sites without the need of linkers. This should improve the flexibility of genetic engineering experiments [K56, P3].(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMed2026

Dogme: a nextflow pipeline for reprocessing nanopore RNA and DNA modifications.

MOTIVATION: Oxford Nanopore (ONT) sequencing allows for the direct detection of RNA and DNA modifications from unamplified nucleic acids, which is a significant advantage over other platforms. However, the rapid updates to ONT basecalling models and the evolving landscape of computational tools for modification detection bring about challenges for reproducible and standardized analyses. To address these challenges, we developed Dogme to automate basecalling, alignment, modification detection, and transcript quantification. Dogme automates the reprocessing of ONT POD5 files by integrating basecalling using Dorado, read mapping using minimap2 and subsequent analysis steps such as running modkit. The pipeline supports three major types of sequencing data-direct RNA (dRNA), complementary DNA (cDNA), and genomic DNA (gDNA). Dogme facilitates detection of diverse RNA modifications supported by Dorado such as N6-methyladenosine (m6A), 5-methylcytosine (m5C), inosine, pseudouridine, 2'-O-methylation (Nm) and DNA methylation, while concurrently quantifying full-length transcript isoforms LR-Kallisto for transcript quantification for dRNA and cDNA. RESULTS: We applied Dogme to three separate mouse C2C12 myoblast replicates using direct RNA sequencing on MinION flow cells. We detected 96 603 m6A, 43 476 m5C, 8829 inosine, 10 055 pseudouridine, and 30 320 Nm sites in three biological replicates. The pipeline produced reproducible modification profiles and transcript expression levels across replicates, demonstrating its utility for integrative long-read transcriptomic and epigenomic analyses. AVAILABILITY AND IMPLEMENTATION: Dogme is implemented in Nextflow and is freely available under the MIT license at https://github.com/mortazavilab/dogme, with documentation provided for installation and usage.

RNA↗
PubMed1993

Drug-induced DNA modification in buccal cells of cancer patients receiving carboplatin and cisplatin combination chemotherapy, as determined by an immunocytochemical method: interindividual variation and correlation with disease response.

Twenty-six patients with a variety of tumor types were treated according to a phase 1 experimental treatment protocol consisting of repetitive cycles of cis-diammine(1,1-cyclobutanedicarboxylato)platinum(II) (carboplatin, 200-480 mg/m2) at day 1 and cis-diamminedichloroplatinum(II) (cisplatin, 50-100 mg/m2) at day 3. Buccal cells were collected in one or two treatment cycles prior to carboplatin, 24 h after carboplatin, just prior to cisplatin, and approximately 24 h after cisplatin administration. Drug-induced DNA modification was visualized at the single cell level by anti-serum NKI-A59 and quantitated by microdensitometry. All (39 of 39) treatments with carboplatin, and almost all (33 of 35) treatments with cisplatin resulted in an increase in nuclear stain. Interindividual variation in drug-induced, adduct-specific nuclear stain amounted to a factor of 5-8 for carboplatin and 5-12 for cisplatin. This drug-induced increase was, however, not related to the dose of either carboplatin or cisplatin, suggesting that large interindividual differences in DNA adduct formation and/or repair obscured the effects of dose variation within the relatively small range used for the drugs (2.4 for carboplatin and 2.0 for cisplatin). This explanation was strengthened by the good reproducibility of the immunocytochemical assay and by the reasonable correlation between carboplatin-induced nuclear stain in cycles 1 and 2 (correlation coefficient, 0.69; P = 0.009). Mean carboplatin-induced nuclear stain was significantly higher in the first cycle than in the second cycle (P = 0.0001) but this difference was no longer significant when drug-induced nuclear stain was corrected for carboplatin dose. Differences in cisplatin-induced nuclear stain between cycle 1 and cycle 2 were small and not significant. Carboplatin-induced nuclear stain was significantly higher in the partial responders than in the nonresponders (P < 0.0001, two cycles combined); the level of statistical significance remained the same after dose correction. Cisplatin-induced nuclear stain did not differ significantly between partial responders and nonresponders; this result might, however, be confounded to some extent by remaining carboplatin-induced nuclear stain at the moment of cisplatin administration. It is concluded that determination of the extent of platinum-induced DNA modification might be helpful in predicting the tumor response in cancer patients.

Adult↗
PubMed1995

Effects of a single dose of the cytochrome P450 inducer, beta-naphthoflavone, on hepatic and renal covalent DNA modifications (I-compounds).

I-compounds are age-dependent covalent DNA modifications, which occur in rodent tissues without known carcinogen exposure. A number of studies from our laboratory indicate that I-compounds may serve as biomarkers of carcinogenesis. Recently, we demonstrated significant lowering of liver I-compound levels in rats that were exposed to different cytochrome P450 inducers. In order to gain further mechanistic insights into the possible relationship between P450 induction and I-compound reduction, female Sprague-Dawley rats were administered a single dose of the CYP1A1 inducer, beta-naphthoflavone (BNF) (80 mg/kg), in corn oil (CO) (2 ml/kg) or CO only (2 ml/kg) as vehicle control. Liver and kidney microsomal P450 contents and P450-related enzyme activities and DNA I-compounds were determined at 4, 24, and 48 h after treatment. Liver and kidney I-compounds were analyzed by nuclease P1-enhanced 32P-postlabeling. DNA synthesis was determined by measuring [3H]methylthymidine incorporation. Liver and kidney microsomal P450 contents were elevated by BNF at 24 and 48 h. Ethoxyresorufin-O-deethylase (EROD) and methoxyresorufin-O-demethylase (MROD) were significantly elevated at all time points, with the former displaying a much higher extent of induction. BNF treatment resulted in significant diminution of the levels of several individual and total I-compounds in liver at 48 h, but few effects were seen at the earlier time-points. Kidney I-compounds were also markedly affected by BNF at 48 h, albeit to a lesser extent than in liver. In both tissues, P450 induction preceded I-compound reduction. Taken together, the results of this investigation demonstrate significant diminution of I-compound levels by a single dose of BNF, a CYP1A1 inducer, in a time-dependent manner, suggesting the participation of a specific biochemical process, possibly involving CYP1A1, in the metabolic regulation of these endogenous DNA adducts.

Animals↗
PubMed2002

DNA modification by 4-aza-3-ene-1,6-diynes: DNA cleavage, pH-dependent cytosine-specific interactions, and cancer cell cytotoxicity.

The (Z)-hex-1,5-diyne-3-ene reactive core common to the enediyne antitumor antibiotics undergoes a Bergman cyclization after proper activation to afford reactive diradical intermediates that are responsible for initiating DNA cleavage. Direct modification of the enediyne core has been proposed as a method to permit cancer cell-specific triggering of the diradical-generating cyclization. For example, 3-aza-3-ene-1,5-diynes undergo an aza-Bergman cyclization to afford the fleeting 2,5-didehydropyridine diradicals. While protonation of these aza-enediynes can afford products of diradical trapping, the hydrolytic instability of the 3-aza-3-ene-1,5-diyne moiety prevents its use in pH-triggered DNA cleaving anticancer agents. Recently, more hydrolytically stable systems incorporating the 4-aza-3-ene-1,6-diyne moiety were developed. We report here studies of the 4-aza-3-ene-1,6-diyne-containing benzimidazolium salt AZB002 [1-methyl-2-(phenylethynyl)-3-(3-phenylprop-2-ynyl)-3H-benzimidazolium tetrafluoroborate] and two structurally related heterocycles that lack the aza-enediyne functionality, AZB016 [1,3-dimethyl-2-(phenylethynyl)-3H-benzimidazolium triflate] and AZB004 [3-methyl-2-(phenylethynyl)benzothiazolium triflate]. The interaction of these compounds with supercoiled DNA, a double-stranded DNA fragment, and a short DNA duplex oligonucleotide was investigated. There are three distinct DNA interactions exhibited by AZB002: a frank strand scission leading to the relaxation of supercoiled DNA and formation of at least two different DNA adducts, one of which leads to cytosine-specific cleavage after piperidine/heat treatment. In contrast, analogues lacking the aza-enediyne functionality either fail to interact with DNA (AZB016) or cleave DNA at guanine residues, presumably through alkylation of the N-7 position (AZB004). We also investigated the cytotoxicity of AZB002 and the related heterocyclic compounds AZB004 and AZB016 and find that only the DNA interactive compounds AZB002 and AZB004 display significant cytotoxicity. In particular, AZB002 is cytotoxic against a wide range of cancer cell lines.

Alkynes↗
PubMed1990

Age-related DNA modifications (I-compounds): modulation by physiological and pathological processes.

I-compounds are covalent DNA modifications that can be detected and measured by 32P-postlabeling assay because of their DNA-adduct like properties. They accumulate in an age-dependent, highly reproducible manner in tissue DNA of untreated animals in the absence of exogenous carcinogens and, therefore, appear to arise via the interaction of DNA with endogenous reactants formed in the course of normal metabolism. Chromatographically, they exhibit a wide range of polarities, indicative of structural diversity. In addition to age-dependent increases, I-compound profiles exhibit prominent species-, sex-, tissue- and diet-dependent qualitative and quantitative differences. Natural-ingredient (chow) diets produce qualitative differences as well as substantially higher I-compound levels in rat liver and kidney, when compared with purified diets. Modified purified diets containing high carbohydrate, protein, or fat concentrations further modulate I-compound profiles. During liver regeneration, I-compounds behave like DNA adducts rather than m5 C in that their levels are not quickly restored. Treatment of rats with the hepatocarcinogens 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), CCl4, and peroxisome proliferators as well as with a choline-devoid hepatocarcinogenic diet depressed the age-related increases of I-compound levels in liver, the target organ. Additional 32P-labeled derivatives were observed only with the peroxisome proliferators and presumably represent DNA adducts of exogenous origin. No I-compounds were detected in a series of Morris hepatomas with different degrees of differentiation. Thus, loss of I-compounds may be associated with altered gene expression/dedifferentiation. On the other hand, the age-dependent accumulation of I-compounds and their adduct-like character suggest potential relations to aging-associated dysdifferentiation and initiation of cancer. Structural complexity indicates different biological roles of I-compounds.

Aging↗
PubMed1990

Differential effect of gestation stage on benzo(a)pyrene-induced micronucleus formation and/or covalent DNA modifications in mice.

Benzo(a)pyrene (BP), an environmental carcinogen, binds ubiquitously to the DNA of maternal and fetal tissues (Lu et al., Cancer Res., 46: 3046-3054, 1986). These studies further investigated the effect of gestation age on the induction of genetic damage by BP. Timed-pregnant ICR mice were treated with one dose of BP on various days of gestation and sacrificed 24-120 h after treatment. At the molecular level, BP covalently bound to the DNA of adult bone marrow and fetal liver of mice at all gestation ages. Compared to the nonpregnant mice (adduct level = 15 adducts/10(8) bases), the adduct levels in the pregnant adult bone marrow were decreased up to 50% during early gestation (days 3-9) and then increased steadily to a 4-fold excess over nonpregnant values during late gestation (days 15-18). In the fetal liver, adduct levels exhibited little variation (3-4 adducts/10(8) bases) between days 11 and 15 of gestation and then increased sharply to 14 adducts/10(8) bases after day 16. At the cellular level, a higher percentage of polychromatic RBCs from adult and fetal mice after BP treatment contained micronuclei (MN) than controls (solvent or untreated). Bone marrow from pregnant mice exhibited greater increases in the formation of MN during early gestation (days 3-9) relative to late gestation (days 15-18), compared to the nonpregnant mice. In the fetuses, the amounts of MN formed were higher than those found in the adult nonpregnant or maternal mice, but these amounts decreased with gestation progression. Thus, MN induction with gestation progression differed from DNA adduction in adults and fetuses. In addition, the dose and time responses of MN formation also differed from those of covalent DNA modifications, when analyzed in the bone marrow of pregnant mice treated on gestation day 5. Collectively, our results showed that pregnancy and development modulate different types of genetic damage in different ways. Fetal tissues may be more sensitive than maternal tissues to genetic damage. Factors in addition to DNA adduct formation may be responsible for MN induction.

Age Factors↗
PubMed2016

A probabilistic generative model for quantification of DNA modifications enables analysis of demethylation pathways.

We present a generative model, Lux, to quantify DNA methylation modifications from any combination of bisulfite sequencing approaches, including reduced, oxidative, TET-assisted, chemical-modification assisted, and methylase-assisted bisulfite sequencing data. Lux models all cytosine modifications (C, 5mC, 5hmC, 5fC, and 5caC) simultaneously together with experimental parameters, including bisulfite conversion and oxidation efficiencies, as well as various chemical labeling and protection steps. We show that Lux improves the quantification and comparison of cytosine modification levels and that Lux can process any oxidized methylcytosine sequencing data sets to quantify all cytosine modifications. Analysis of targeted data from Tet2-knockdown embryonic stem cells and T cells during development demonstrates DNA modification quantification at unprecedented detail, quantifies active demethylation pathways and reveals 5hmC localization in putative regulatory regions.

5-Methylcytosine↗
PubMedDate not supplied

[A method for detecting new strains producing DNA modification and restriction enzymes--isoschizomers and isomethylomers of known restrictases and methylases].

The paper describes a technique for the detection of new strains producing enzymes which mediate DNA modification and restriction, and isoschizomers and isomethylomers of the known restriction endonucleases and methylases. Three Bacillus subtilis strains whose DNA carries a BamH1 modification have been found. Two of these strains exert the restrictase activity with an R BamH1 specificity.

Bacillus subtilis↗
PubMed1988

Tet repressor-tet operator contacts probed by operator DNA-modification interference studies.

Contacts between tet operator DNA and Tet repressor protein are characterized by modification interference studies. The modified DNA fragments are separated into fractions with high, intermediate and low affinities for Tet repressor by polyacrylamide gel electrophoresis. Ethylation of the phosphates with N-ethylnitrosourea reveals 12 contacts of a repressor dimer to tet operator. Eight of these contacts appear to be important for Tet repressor binding, as judged by the strong interference at these positions, while four contacts are probably less important. All of the phosphate contacts are located on the same side of the B-DNA structure. The sequences of tet operators proposed to interact with the recognition alpha-helix of Tet repressor are TCTATC in three cases and CCTATC in one case. After methylation of N-7 with dimethylsulfate, strong interference is observed at the guanine residues at positions +/- 2. None of the N-7 functions of other guanine residues seems to be involved in tight contacts to Tet repressor. Tet repressor subunits form identical phosphate and guanine N-7 contacts with each half side of the two tet operators indicating twofold dyad symmetry of the complexes. Attempts to analyze the methylation interference at the adenine N-3 sites reveal different results for the operators. Modification of DNA fragments with diethylpyrocarbonate yields hypersensitive sites in the tet operators, indicating different local DNA structures. Carbethoxylation interference studies confirm the contacts at the purines found by methylation interference. All of the sequence-specific protein-DNA contacts detected in this study are centered at the inside four base-pairs in each tet operator half side. The contacts are discussed with respect to the structure of the repressor-operator complex.

Autoradiography↗
PubMed1991

Modulation by dietary vitamin E of I-compounds (putative indigenous DNA modifications) in rat liver and kidney.

I(indigenous)-compounds are age-related, carcinogen adduct-like, putative indigenous DNA modifications detectable by 32P-postlabeling assay in untreated animals. To investigate the origins of these DNA derivatives, we examined the effects of dietary vitamin E, a natural antioxidant, on I-compounds of rat liver and kidney DNA. Weanling female Sprague-Dawley rats were fed Draper's diets containing 0, 100, 1000, or 10,000 mg/kg alpha-tocopheryl acetate for 6 mo. The DNA from four individual rats of each group was analyzed by a nuclease P1-enhanced version of the 32P-postlabeling assay for DNA adducts. The amount of vitamin E in the liver was measured by high performance liquid chromatography. Rats fed vitamin E-deficient diet (0 mg/kg) showed identical profiles and similar levels of I-compounds as those fed the 100 mg/kg diet. Most I-spots were significantly intensified and one tissue-specific extra spot was found in both liver and kidney DNA of rats fed the 1000 or 10,000 mg/kg vitamin E diet. However, one of the five major I-spots detected in the kidney was weaker in the 1000 and 10,000 mg/kg groups than in the 0 and 100 mg/kg groups. These results show that formation of most I-compounds was not affected by vitamin E-deficient diet, and that long-term feeding of diet containing high levels of vitamin E may cause metabolic alterations leading to an increased formation of DNA-reactive (potentially mutagenic or carcinogenic) electrophiles.

Animals↗
PubMed1992

Sex-specific modulation of hepatic covalent DNA modifications (I-compounds) by the cytochrome P450 inducer, pregnenolone-16 alpha-carbonitrile.

I-compounds are recently discovered, age-dependent covalent DNA modifications, which are detectable by 32P-postlabeling assay for DNA adducts. The effects of the catatoxic antiglucocorticoid, pregnenolone-16 alpha-carbonitrile (PCN), on hepatic and renal I-compound levels have been studied in male and female Sprague-Dawley rats together with the levels of microsomal cytochrome P450 and rates of ethylmorphine N-demethylation. PCN (50 mg/kg ip) was dissolved in corn oil and administered to rats once daily for 4 days, and animals were killed at 1 day or 8 days after the last treatment. Hepatic and renal I-compounds were analyzed by 32P-postlabeling in control and PCN-treated animals at both time points. Microsomal cytochrome P450 and ethylmorphine N-demethylase activities were also determined. Total levels of liver nonpolar and polar I-compounds were reduced in female rats by 37 and 51%, respectively, compared to controls, at 1 day. Ten out of sixteen individual I-compounds were also markedly reduced in female rat liver DNA as a result of PCN administration. In contrast to females, total levels of liver I-compounds were not significantly altered in males by PCN at 1 day; however, two individual I-compounds were lowered. I-compound levels recovered 8 days after termination of PCN treatment in both males and females. Total levels of renal I-compounds were not affected by PCN treatment in either males or females. [3H]Methylthymidine incorporation studies showed an increase in mean DNA synthesis rate at 1 day in liver of both males and females, but this was significant in males only. Marked induction of hepatic microsomal cytochrome P450 (2.2-fold) and ethylmorphine N-demethylase (4.0-fold) activity was observed in female rats treated with PCN at 1 day as compared to controls. The extent of induction of these enzymes was much higher in females than males. At 8 days the levels of cytochrome P450 and ethylmorphine N-demethylase activity had returned to uninduced values. The results are consistent with a pivotal role for PCN-inducible cytochrome P450 in the metabolism of I-compounds.

Animals↗