SEARCH · PubMed Health

Results for “variable methylation”

Explore indexed PubMed citations for clinical trials, systematic reviews and public health research. Read source abstracts and follow each citation to its original PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 127 records · Page 7Linked to original sources

PubMed2020

Harsh Parenting Predicts Novel HPA Receptor Gene Methylation and NR3C1 Methylation Predicts Cortisol Daily Slope in Middle Childhood.

Adverse experiences in childhood are associated with altered hypothalamic-pituitary-adrenal (HPA) axis function and negative health outcomes throughout life. It is now commonly accepted that abuse and neglect can alter epigenetic regulation of HPA genes. Accumulated evidence suggests harsh parenting practices such as spanking are also strong predictors of negative health outcomes. We predicted harsh parenting at 2.5 years old would predict HPA gene DNA methylation similarly to abuse and neglect, and cortisol output at 8.5 years old. Saliva samples were collected three times a day across 3 days to estimate cortisol diurnal slopes. Methylation was quantified using the Illumina Infinium MethylationEPIC array BeadChip (850 K) with DNA collected from buccal cells. We used principal components analysis to compute a summary statistic for CpG sites across candidate genes. The first and second components were used as outcome variables in mixed linear regression analyses with harsh parenting as a predictor variable. We found harsh parenting significantly predicted methylation of several HPA axis genes, including novel gene associations with AVPRB1, CRHR1, CRHR2, and MC2R (FDR corrected p < 0.05). Further, we found NR3C1 methylation predicted a steeper diurnal cortisol slope. Our results extend the current literature by demonstrating harsh parenting may influence DNA methylation similarly to more extreme early life experiences such as abuse and neglect. Further, we show NR3C1 methylation is associated with diurnal HPA function. Elucidating the molecular consequences of harsh parenting on health can inform best parenting practices and provide potential treatment targets for common complex disorders.

Child↗

PubMed1975

A comparison of four quantitative cytochemical methods directed toward demonstration of DNA.

Populations of nuclei isolated from mouse brain tissue were stained by the following cytochemical methods considered stoichiometric for DNA: (1) the Feulgen reaction; (2) gallocyanin-chromalum after RNase; (3) pH 4.0 methylene blue after RNase; and (4) methyl green used in the presence of 2M magnesium chloride. Replicate preparations to be stained with gallocyanin-chromalum, methylene blue, and methyl green were acetylated prior to staining. All of these groups were examined by high-resolution scanning microspectrophotometry. The results indicated that of the methods examined, the Feulgen reaction, gallocyanin-chromalum used without prior acetylation, and methylene blue used with prior acetylation were the most useful in revealing differences attributable to variability in chromatin organization. The greatest variability in total extinction measurements was observed in acetylated, methylene blue-stained nuclei, while the least variability was observed in nuclei stained with methyl green in the presence of 2 M magnesium chloride. Acetylation produced different effects on dye-binding in different groups. It greatly increased binding in nuclei stained with methylene blue; it reduced binding in the methyl green-2 M magnesium chloride series.

Acetylation↗

PubMed1976

Characterization of microsomal methyl sterol demethylase in two Morris hepatomas.

Previously, we reported that the rate of metabolism of methyl sterol intermediates of cholesterol biosynthesis by broken-cell preparations of Morriss hepatoma 7777 is very slow, whereas the intact tumors are known to synthesize cholesterol quite efficiently. Active preparations have now been obtained by substitution of pyrophosphate for phosphate buffer. Although substitution of pyrophosphate buffer markedly enhances microsomal methyl sterol demethylation rates 3- to 4-fold in hepatoma 7777, other microsomal enzymes and electron carriers in either liver or a more slowly growing hepatoma appear to be unaffected by pyrophosphate. Several properties of the active microsomal methyl sterol demethylase have now been compared for control rat liver, host liver, tumor 7777, and tumor 5123C. Conditions necessary for the assay of initial velocities of enzymic reactions in the tumor microsomes have been established with respect to the amount of protein, time-course, concentrations of cofactors and substrate, pH, and other variables. The K'm and the responses to the variables studied above are very similar for methyl sterol demethylase of microsomes isolated from control liver, host liver, tumor 5123C, and tumor 7777. The multienzymic demethylase in the various preparations has been found to be inhibited similarly by in vitro additions of cyanide, cytochrome c, and bile salts. Thus, the enzymes of the microsomal-bound 4-methyl sterol demethylase of cholesterol biosynthesis appear to be very similar in liver and these 2 Morris hepatomas. When xenobiotic inducers of microsomal oxidases, such as phenobarbital and methylcholanthrene, are administered to normal and tumor-bearing rats, elevated rates of methyl sterol demethylation are observed with isolated liver microsomes obtained from both normal and tumor-bearing rats. Similar increases are not observed in the tumors. Furthermore, daily administration of an intestinal bile acid sequestrant elevates hepatic methyl sterol demethylase, but statistically significant changes were not observed in tumors 7777 and 5123C. Since the enzymes of methyl sterol demethylase appear to be grossly similar in liver and these hepatomas, regulation of the activity of the multienzymic system contained in the tumors may be altered. On the other hand, these agents in vivo simply may not affect liver and the hepatomas similarly, due to a lack of uptake of the foreign substances by the tumor that has been transplanted to the thighs.

Animals↗

PubMed1993

DNA methylation and mutation.

5-Methylcytosine (5mC) in DNA is produced by post-synthetic modification of cytosine residues, and it occurs primarily in CpG doublets in the mammalian genome. 5mC is a mutable site, because it can undergo spontaneous deamination to thymine. There is a repair mechanism which specifically recognises G.T mispairs, and replaces thymine with cytosine. However, this repair is not fully efficient, because the 5mC-->T transition mutation occurs about 10 times as frequently as other transitions. Such mutations are frequently seen in inherited diseases, and mutations in the p53 gene in tumours are also very commonly in 5mCpG doublets. As well as mutations, there can also be heritable changes in DNA methylation, known as epimutations, which may be of particular significance in somatic cells. Whereas the pattern of DNA methylation is very constant for any one cell type, the pattern becomes very variable in tumour cells. The breakdown of the normal controls of DNA methylation in tumorigenesis can lead to increased gene expression or to gene silencing. DNA damage increases not only mutation, but also heritable changes in methylation. At present, little is known about the ability of DNA repair to preserve the normal pattern of methylation in somatic cells.

5-Methylcytosine↗

PubMed1984

DNA methylation versus gene expression.

Vertebrate DNA is methylated at a high proportion of cytosine residues in the sequence CpG, and it has been suggested that the distribution of methylated and non-methylated CpGs in a given cell type influences the pattern of gene expression in those cells. Since a DNA methylation pattern is normally transmitted faithfully to daughter cells via cell division, this idea suggests an origin for stable, clonally inherited patterns of gene expression. This article discusses some of the current evidence for a relationship between DNA methylation and gene expression. Although the evidence is incomplete, it appears already that the relationship is variable: transcription of some genes is repressed by the presence of 5-methylcytosine at certain CpGs, and may be controlled by methylation, while transcription of other genes is indifferent to methylation. In attempting to explain this variability it is helpful to adopt an evolutionary perspective.

5-Methylcytosine↗

PubMed1995

Elementary forms of synaptic plasticity in the visual cortex.

The neocortex is an important site of memory storage, and memories are believed to be formed in the cortex by the activity-dependent modification of synaptic connections. However, in contrast to the hippocampus where there has been an increasingly sophisticated analysis of synaptic plasticity, relatively little is known about the mechanisms of synaptic modification in neocortex. Here we summarize the results of a series of experiments conducted on slices of visual cortex in vitro, aimed at elucidating the elementary mechanisms of synaptic plasticity in the superficial layers of neocortex. We show that long-term potentiation (LTP) and depression (LTD) result from high- and low-frequency conditioning stimulation, respectively, of the middle layers of cortex. Both forms of synaptic plasticity are input-specific and dependent on activation of postsynaptic N-methyl-D-aspartate (NMDA) receptors. The critical variable in determining the sign of the synaptic modification appears to be the level of postsynaptic depolarization during conditioning stimulation. The data support a model in which the state of correlation of pre- and post-synaptic activity is converted by the voltage-dependent NMDA receptor channel into a graded postsynaptic Ca2+ signal. LTD is triggered by a modest but sustained elevation in postsynaptic Ca2+, while LTP is elicited by larger changes in Ca2+. An important variable that regulates synaptic plasticity in the neocortex is intracortical inhibition, which constrains the patterns of activity that can reach the modifiable synapses in layer III.

Calcium↗

PubMed1995

Temporal and spatial regulation of H19 imprinting in normal and uniparental mouse embryos.

The mouse H19 gene is imprinted so that the paternal copy is both methylated and repressed during fetal development. However, the CpG-rich promoter region encompassing the transcription start is not methylated in sperm; this region must therefore become methylated postzygotically. We first examined the timing of DNA methylation of this region and the corresponding expression of H19. Both parental copies are initially undermethylated in blastocysts and the paternal copy then becomes fully methylated in the embryo around implantation; this methylation is more protracted in the extraembryonic lineages, especially in the trophoblast. By contrast to the lineage-dependent methylation, we observed exclusive expression of the maternal copy in preimplantation embryos and in all the lineages of early postimplantation embryos although variability may exist in cultured embryos. This indicates that methylation of the CpG-rich promoter is not a prerequisite for the paternal repression. We then examined whether methylation and expression occurs appropriately in the absence of a maternal or a paternal genome. Both H19 copies in androgenetic embryos are fully methylated while they are unmethylated in parthenogenetic embryos. This correlates with the lack of expression in androgenetic embryos but expression in parthenogenetic embryos. However, the androgenetic trophoblast was exceptional as it shows reduced methylation and expresses H19. These results suggest that promoter methylation is not the primary inactivation mechanism but is a stabilizing factor. Differential methylation in the more upstream region, which is established in the gametes, is a likely candidate for the gametic signal and may directly control H19 activity.

Animals↗

PubMed1991

The sequence specificity domain of cytosine-C5 methylases.

Prokaryotic DNA[cytosine-C5]methyltransferases (m5C-methylases) share a common architectural arrangement of ten conserved sequence motifs. A series of eleven hybrids have been constructed between the HpaII (recognition sequence: Cm5CGG) and HhaI (recognition sequence: Gm5CGC) DNA-methylases. The hybrids were over-expressed in E.coli and their in vivo methylation phenotypes investigated. Six were inactive by our assay while five of them retained partial methylation activity and full specificity. In all five cases the specificity matched that of the parent methylase which contributed the so-called variable region, located between conserved motifs VIII and IX. This was the only sequence held in common between the active hybrids and for the first time provides unequivocal evidence that the specificity determinants of the mono-specific m5C-methylases are located within the variable region. Correlation of the hybrid methylase structure with the efficiency of methylation suggests that conserved motif IX may interact with the variable region whereas motif X most probably interacts with the N-terminal half of the molecule.

Amino Acid Sequence↗

PubMedDate not supplied

The hair-organ relationship in mercury concentration in contemporary Japanese.

The hair-organ relationship of mercury concentration was investigated in 46 autopsy samples in Tokyo, Japan. Hair mercury levels were highly significantly correlated with organ Hg levels in the cerebrum, cerebellum, heart, spleen, liver, kidney cortex, and kidney medulla, when the total mercury or methyl mercury value in the organ was compared with the hair total mercury or organic mercury, respectively. When the inorganic mercury value was tested, significant correlations remained, with weaker coefficients in all the organs but the spleen. Stepwise multiple regression analysis evidenced that the hair organic mercury value was the major explanatory variable for the organ total mercury or organ methyl mercury value in all the organs. To explain the organ inorganic mercury value, the hair organic mercury value was the major variable for the cerebrum and kidney (both cortex and medulla), the hair inorganic mercury value was the major variable for the cerebellum and heart, and the hair phosphorous and hair organic mercury were the major variables for the liver; no explanatory variable existed for the spleen. Auxiliary explanatory variables accounted for the organ total mercury and inorganic mercury levels, among which the hair selenium value was conspicuous with negative regression coefficients.

Adolescent↗

PubMed1990

Intestinal absorption of menhaden and rapeseed oils and their fatty acid methyl and ethyl esters in the rat.

The relative cellular uptake and incorporation into prechylomicrons and chylomicrons was investigated for the menhaden and rapeseed oil fatty acids, when given by stomach tube as the original oils or the corresponding methyl and ethyl esters. The intermediates and final products of cellular acylation were determined by chromatographic methods at various times over a period of 1-24 h. There was little selectivity in the uptake among the oligo- and poly-unsaturated fatty acids of menhaden oil, when either oil or esters were fed. In contrast, the long-chain saturated and monounsaturated fatty acids of rapeseed oil were discriminated against during both cellular uptake and reacylation (60% overall reduction in utilization). Also, there was detectable discrimination against the long-chain polyunsaturated monoacylglycerols of menhaden oil and against the long-chain saturated and monounsatured monoacylglycerols of rapeseed oil during both cellular uptake and reacylation (30% overall reduction in utilization). Evidence was obtained for an indiscriminate cellular uptake of variable amounts (4-22%) of intact dietary methyl and ethyl esters of fatty acids, which, however, appeared in the chylomicrons only to a very limited extent (0.1-1.0% of total lipid). During peak absorption the cellular and lymphatic appearance of fatty acids from the digestion and absorption of the alkyl esters was nearly 50% lower than that from the corresponding triacylglycerols. The slower absorption of the fatty acids from the alkyl ester feeding is hypothetically attributed to a lower efficiency of the phosphatidic acid pathway, which is required in the absence of dietary 2-monoacylglycerols, but other mechanisms cannot be excluded.

Animals↗

PubMed1977

Temperature-sensitive variants for saturation density and anchorage dependencey of a simian virus 40-transformed human X mouse hybrid cell line.

Several variant clones temperature sensitive for some parameters of transformation from a hybrid cell line containing a stable diploid mouse genome and two human chromosomes 7 carrying an integrated defective simian virus 40 have been isolated. Like the wild-type (wt) cells, at the permissive temperature the temperature-sensitive (ts) clones grew to high saturation densities and readily formed colonies in methyl cellulose. In contrast to the wt cells, at the nonpermissive temperature they had variable but always lower saturation densities and were unable to form colonies in methyl cellulose. The fraction of cells that synthesized DNA decreased at both temperatures when the cells reached saturation density, but it represented always at least 20% of the total population. All of the ts clones so far tested had a near triploid chromosome number, contained from one to three human chromosomes 7, and were T-antigen-positive both at the permissive and nonpermissive temperatures. The ts clones maintained low saturation density at the nonpermissive temperature because of a decrease in the number of cells that were actively proliferating and because of the shedding of cells into the medium. Temperature downshifts and refeeding allowed for expression of the permissive phenotype. In most of the isolated clones anchorage independency was not correlated with unrestricted cell proliferation. These variant clones may provide useful systems for a better understanding of the number of interdependent pathways involved in the expression of the phenotype of a transformed cell and elucidate the molecular mechanism required for the maintenance of a normal state in a cell population.

Antigens, Viral↗

PubMed1978

Inhibition of morphological transformation induced with N-methyl-N'-nitro-N-nitrosoguanidine in cultures of hamster embryo cells by 5'-bromo-2'-deoxyuridine-photolysis.

The present study was performed in order to determine whether type III transformed foci induced by N-methyl-N'-nitro-N-nitrosoguanidine originate from the small subpopulation of cells stimulated by the carcinogen to enter DNA synthesis. During the last 30 min of variable treatment periods using different doses of N-methyl-N'-nitro-N-nitrosoguanidine, administered alone or in association with the thymidine analogue, 5'-bromo-2'-deoxyuridine (0.98 x 10(-5)M), the density-inhibited monolayers of hamster embryo cells were exposed to fluorescent light and then assayed for abnormal growth patterns by the focus formation method. Mock-irradiated cultures as well as monolayers whose medium lacked N-methyl-N'-nitro-N-nitro-soguanidine, 5'-bromo-2'-deoxyuridine, or both, served as controls. The cytotoxicity of 5'-bromo-2'- deoxyuridine + N-methyl-N'-nitro-N-nitrosoguanidine + photolysis (BMP) protocol on confluent as well as logarithmically growing hamster embryo cells was estimated in single-cell survival experiments. Plating efficiency determinations have demonstrated that, unlike their actively growing counterparts, confluent hamster embryo cell monolayers are extremely resistant to the cytotoxic effects of the BMP protocol. The quantitative transformation assays indicated that: (1) in non-illuminated cultures addition of 5'-bromo-2'-deoxyuridine to carcinogen-containing medium does affect transformation frequency of hamster embryo cells in the sense that the incidence of type III foci did not subside at later intervals during the post-carcinogen administration period as it did in the absence of the analogue; (2) irradiation of N-methyl-N'-nitro-N-nitrosoguanidine and halogenated pyrimidine analogue-treated cultures with fluorescent light practically suppressed transformation; (3) analogue-added and analogue-removed experiments pointed out that the event(s) on which 5'-bromo-2'-deoxyuridine fluorescent light sensitization of morphological transformation largely depends, takes place between 5 and 15 h after N-methyl-N'-nitro-N-nitrosoguanidine administration, i.e., during the period of maximal carcinogen-stimulated DNA synthesis; and (4) neither fluorescent light nor 5'-bromo-2'-deoxyuridine, singly or in combination, were able to transform cultures of hamster embryo cells. These findings are strong indirect arguments for the concept that carcinogen-induced DNA synthesis and the initiation of transformed clones are causally related.

Animals↗

PubMed1992

Abnormalities of insulin-like growth factor (IGF-I and IGF-II) genes in human tumor tissue.

Recent findings have indicated that Insulin-like growth factors (IGF-I and IGF-II) may play a role in neoplasia. Expression of their genes, which are highly complex structures, is tissue-specific and developmentally regulated. The purpose of the present study was to determine whether a relationship exists between tumorigenesis and the structure and expression of IGF genes. The structures of the IGF-I and IGF-II genes were investigated in 40 tumors by Southern blot analysis but no obvious re-arrangements (such as amplification or deletion) were observed in any of the tissues investigated. DNA methylation was also studied, using the enzyme Avall. The extent of DNA methylation of the IGF genes was highly variable in most of the tumors, as was the level of mRNA expression. A relationship could be detected between IGF over-expression and gene demethylation in tumors associated with hypoglycemia and in certain hepatocarcinomas. Loss of heterozygosity has been reported in the 11p15 region of some childhood tumors. The present findings provide further evidence of this loss of heterozygosity for the IGF-II gene and show an imbalance in the leukocyte alleles in several childhood tumors. Likewise, an imbalance in the alleles was noted in several adult tumors, including hepatocarcinomas and breast cancers. This suggests that in certain adult tumors alterations of the IGF-II gene may be associated with tumorigenesis, but in other tumors another mechanism may be involved.

Alleles↗

PubMed1993

Structural characterization and expression analysis of the Neurospora conidiation gene con-6.

The gene con-6 of Neurospora crassa is expressed during the formation of asexual spores (conidia), but it is not expressed in mycelium. con-6 mRNA appears upon induction of conidiation and reaches high levels at the late stages of conidiation, and in mature conidia. The CON6 polypeptide and a CON6-beta-Gal fusion protein were present at high levels only in free conidia. Shortly after spore germination con-6 mRNA disappears and the CON6 polypeptide is degraded. CON6 is a small, hydrophilic polypeptide containing a repeat sequence; it not homologous to any known protein but has features resembling the late embryogenesis abundant proteins of maize. Inactivation of con-6 by the repeat-induced point mutation process had no demonstrable effect on formation or germination of conidia. Upstream sequence comparisons for con-6 and other con genes identified a common potential regulatory sequence, designated CRS-B. DNA mobility shift analyses with cell extracts identified a factor that bound to synthetic DNA fragments containing this sequence. This binding factor was present in mycelium but not in conidiating cultures. Experiments with independent integrated con-6'-'lacZ translational fusions revealed substantial variability of expression among transformants carrying identical fusion constructs: This variability may be due to the differential methylation of transformant DNA noted by others.

Amino Acid Sequence↗

PubMed1977

Non-equilibrium method for the radioimmunoassay of clozapine in the presence of metabolites.

Cross-reactions with metabolites are an ever-recurring problem encountered in the use of radioimmunoassay techniques to determine active compounds in biological material. Metabolites may interfere with the assay of the parent drug to a variable extent. Taking 8-chloro-11-(4-methyl-1-piperazinyl)-5H-dibenzo[b,e][1,4]diazepine (clozapine as an example, it was shown that the extent to which the antiserum produced interacts with the parent drug and the metabolites can be estimated by determining the equilibrium constants and the kinetics. In the present case, therefore, it was advantageous to carry out the radioimmunoassay in disequilibrium, i.e. in order to differentiate the metabolites from the parent drug, the sample was incubated with the antiserum for 10 min, after which the labelled antigen was added and the reaction mixture again incubated for a brief, exactly timed interval. It was shown that cross-reactions did not occur in mixtures of clozapine and its N-demethyl and N-oxide metabolites in the propor tions 1:1:2 over a range of concentration of 1.5-48 ng clozapine per 100 microliter human plasma. The equilibrium constants measured with the clozapine goat antiserum were as follows: clozapine 1.2 X 10(8) M-1, the N-demethyl metabolite 4.6 X 10(7) M-1 and the N-oxide metabolite 3.7 X 10(7) M-1 (pH 7.5 and 20 degrees C).

Administration, Oral↗

PubMed1984

Comparison of sigma- and kappa-opiate receptor ligands as excitatory amino acid antagonists.

Using the technique of microelectrophoresis in pentobarbitone-anaesthetized cats and rats, the effects of benzomorphans, with known actions at sigma- and kappa- opioid receptors, were tested on responses of spinal neurones to amino acids and acetylcholine. The racemic mixture and both enantiomers of the sigma opiate receptor agonist, N-allylnormetazocine (SKF 10, 047), and the dissociative anaesthetic, ketamine, reduced or abolished excitation evoked by N-methyl-aspartate (NMA) with only small and variable effects on responses to quisqualate or kainate. (+)-SKF 10, 047 was 1.2 +/- 0.7 times more potent than the (-)-enantiomer in antagonizing NMA. On Renshaw cells, (+)-SKF 10, 047 enhanced responses to acetylcholine whereas the (-) enantiomer produced only a small reduction. The kappa- opiate receptor agonist, ethylketocyclazocine, had no selective effects on responses to amino acids or to acetylcholine. We conclude that actions at sigma- but not kappa-, opiate receptors are responsible for the NMA antagonism observed with benzomorphans.

Acetylcholine↗

PubMed1994

Stereoselective differences in the binding of N-methylated barbiturates to human serum albumin.

The binding to human serum albumin (HSA) of a homologous series of N-methylated chiral barbiturates was studied by means of equilibrium dialysis. The length of the aliphatic side chain at C-5 of the barbiturate ring was variable, and the compounds used were: (+)-(S)- and (-)-(R)-5-methyl-(A), -5-ethyl-(B), -5-propyl-(C) and -5-butyl-(D)-1-methyl-5-phenyl-barbiturate. Binding parameters (numbers of binding classes and of binding sites in each class, affinity constant, total binding constant) were obtained from the Scatchard plot of the percent binding values. For both enantiomers of A and D as well as for (-)-(R)-B 2 classes were obtained; (+)-(S)-B and both enantiomers of C had only 1 class. The total binding constant (K) indicated a more than twofold higher binding of (-)-(R)-B (2.64 x 10(3).mol-1) and C (5.75 x 10(3).mol-1) compared with the corresponding (+)-(S)-enantiomer (1.02 and 2.00 x 10(3).mol-1, respectively); in the case of D the (+)-(S)-enantiomer was preferentially bound (K = 10.14 vs. 5.40 x 10(3).mol-1 for the (-)-(R)-enantiomer). The percent binding values of (+)-(S)-A were higher than those of (-)-(R)-A; however, the K-values of the A-enantiomers were almost identical.

Barbiturates↗

PubMed1985

The effects of some beta lactam antibiotics on (3H)-methyl-yohimbine binding to intact human platelets.

Several antibiotics have been reported to cause a bleeding diathesis in man, characterized by reduced platelet aggregation. We investigated the effects of several of the penicillins and of moxalactam on the binding of (3H)-methyl-yohimbine to intact human platelets. The (3H)-methyl-yohimbine binding met the criteria for interaction at an alpha2 adrenergic binding site and showed low interindividual variability. Penicillin G, ticarcillin, carbenicillin, piperacillin and moxalactam all inhibited (3H)-methyl-yohimbine binding, but at concentrations far in excess of clinically achievable plasma levels. We conclude that these compounds exert their antiplatelet effects by a mechanism other than competitive inhibition of catecholamine binding.

Adrenergic alpha-Antagonists↗