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P A Wilce

Publications and source records attributed to P A Wilce.

88 records · Page 5Linked to original sources
PubMed1982

Regulation of 3-hydroxy-3-methylglutaryl-coenzyme A reductase in rat liver and Morris hepatomas 5123C, 9618A and 5123t.c.

Characteristics of 3-hydroxy-3-methylglutaryl-CoA reductase from normal liver, Morris hepatomas 5123C, 5123t.c. and 9618A, and host liver were studied. Animals were fed on control and 5%-cholesterol diets. Microsomal membranes from all tissues were found to accumulate cholesterol after 3 days on the 5%-cholesterol diet. The enzyme of the tumours showed no feedback inhibition by dietary cholesterol, and that of host liver gave a variable response, whereas that of control liver was constantly inhibited by 90% or more. Arrhenius-plot analysis was conducted on the microsomal enzyme isolated from the various tissues. Control animals showed that the phase transition present at 27 degrees C was removed when animals were fed on 5%-cholesterol diet for 12 h. The hepatomas failed to show this change even after 3 days of 5%-cholesterol diet and a significant increase in microsomal cholesterol. This failure to remove the break in Arrhenius plots also occurred in host liver, even though enzyme inhibition occurred. The reason why hepatomas fail to regulate 3-hydroxy-3-methylglutaryl-CoA reductase activity in response to dietary cholesterol may be a decreased membrane-enzyme interaction.

Animals↗
PubMed1981

Metabolism of poly(A)(+)RNA in toad bladder epithelial cells.

Previous studies have characterized the induction of poly(A)(+)RNA synthesis by aldosterone during the latent period, preceding the increased active transepithelial sodium transport (measured as short-circuit current, SCC). To assess the role of aldosterone in the maintenance of the response in general and the metabolism of this RNA in particular, the decay of the increased SCC and of the newly synthesized poly(A)(+)RNA was monitored. On removal of the hormone, the SCC decayed with a half-life of 6.5 hr after a lag period of 2-3 hr. Studies on the disappearance from the cytoplasm of poly(A)(+)RNA synthesized in the first two hours after addition of aldosterone revealed a number of RNA species with diverse size decaying at a relatively slow rate after removal of aldosterone, and RNA sedimenting in the 10-14 S region decaying at a faster rate closely related to the decay in SCC. Maintenance of aldosterone in the media resulted in a much slower rate of decay of this 10-14 S. It is concluded that the decay of the 10-14 S poly(A)(+)RNA is closely related to the decay in SCC and the stability of this RNA is influenced by the retention of aldosterone in the medium.

Aldosterone↗
PubMed1977

Spironolactone antagonism of aldosterone action on Na+ transport and RNA metabolism in toad bladder epithelium.

In earlier studies, aldosterone increased the incorporation of precursors into a class of cytoplasmic RNA with the characteristics of messenger RNA (mRNA), in toad bladder epithelium. In the present studies, this effect was analyzed further with a competitive antagonist, spironolactone (SC-9420). Paired hemibladders were labeled with 3H-uridine (30 min pulse - 140 min chase), with or without aldosterone (3.5 x 10(-8) M, 7 X 10(-8) M) in the presence or absence of SC-9420 (7 X 10(-6) M, 2.5 X 10(-5) M) at molar ratios of 200:1 to 280:1. Cytoplasmic RNA, either the total phenol-SDS extract or polyadenylated-RNA (poly(A)(+)-RNA) obtained by oligo-deoxythymidylate-cellulose (oligo(dT)-cellulose) chromatography was analyzed in linear 5 -- 20% sucrose gradients. Eight sets of experiments were completed in which the short-circuit current (scc) was monitored for 180 min and the incorporation of 3H-uridine (30 min pulse -- 150 min chase) was simultaneously determined on pools of epithelia from 5 to 10 hemibladders. The fractional change in scc correlated linearly with the fractional change in 3H-uridine of 12S cytoplasmic RNA (r=0.95, p less than 0.001). The poly(A)(+)-RNA fraction had no detectable rRNA or tRNA and gave a heterogeneous pattern, typical of mRNA, in the sucrose gradients. In the presence of exogenous aldosterone, SC-9420 inhibited the incorporation of 3H-uridine into poly(A)(+)-RNA (particularly 12S). These results support the inference that induction of mRNA mediates the action of aldosterone on Na+ transport.

Aldosterone↗
PubMed1977

Effects of 3'-deoxycytidine on rRNA synthesis in toad bladder: analysis of response to aldosterone.

Previous studies showed that aldosterone augments transepithelial active Na+ transport and the incorporation of [3H]uridine into polyadenylated RNA (poly(A)(+)-RNA) (putatively mRNA) early in the latent period. Soon thereafter, incorporation of [methyl-14C] groups, as well as [3H]uridine into rRNA is also increased. To evaluate the role of rRNA in mineralocorticoid action, the inhibitor 3'-deoxycytidine was used in studies on the urinary bladder of the toad Bufo marinus. 3'-deoxycytidine suppressed the incorporation of [methyl-14C] and [3H]uridine into nuclear precursors of rRNA and subunits of cytoplasmic rRNA. In contrast, 3'-deoxycytidine inhibited incorporation of ]3H]uridine into cytoplasmic poly(A)(+)-RNA minimally. In control experiments, 3'-deoxycytidine had no significant effect on Na+ transport, measured as the short-circuit current (scc), when given alone. 3'-Deoxycytidine also had no significant effect on the aldosterone-dependent increase in scc. In the presence of 3'-deoxycytidine, aldosterone enhanced both the scc and the incorporation of [3H]uridine into poly(A)(+)-RNA significantly. We conclude that during the first 3 h, the mineralocorticoid action of aldosterone is not sensitive to inhibition of rRNA synthesis. Previous studies, however, implicate mRNA synthesis in this early response.

Aldosterone↗
PubMed1976

Actions of aldosterone on polyadenylated ribonucleic acid and Na+ transport in the toad bladder.

Polyadenylated ribonucleic acid [poly(A)(+)-RNA] has been isolated from the cytoplasm of the epithelium of the urinary bladder of the toad (Bufo marinus) by oligo-(deoxythymidylate)cellulose chromatography. Aldosterone increased the incorporation of [3H]uridine, and of [3H]uridine and [3H]adenosine (given in combination) into 7S, 12S, and 18S poly(A)(+)-RNA during the first 30 min of the action of the hormone, as defined by either a pulse or pulse-chase sequence. The quantity of cytoplasmic poly(A)(+)-RNA that hybridized to [3H]poly(uridine) was also increased by aldosterone. These results are consistent with the inference of induction of messenger RNA synthesis. This effect was most marked during the first 30 min of the action of the hormone.

Aldosterone↗
PubMed1976

Actions of aldosterone on rRNA and Na+ transport in the toad bladder.

Aldosterone increased methylation by [methyl-3H]methionine of nuclear ribosomal ribonucleic acid (rRNA) sedimenting at 18S, 28S, and 40S within 90 min and of 28S cytoplasmic RNA within 240 min of continuous exposure to the precursor and the hormone, in the toad bladder. In addition, incorporation of [14C]uridine into cytoplasmic 4S transfer RNA, and 18S and 28S rRNA was enhanced after 240 min of continuous exposure to the precursor and the hormone. Aldosterone had minimal effects on the 3H or 14C-labeled, acid-soluble pools. These results suggest that aldosterone augments the synthesis of rRNA at the transcriptional level. Ribosomes isolated 240 min after treatment of the toad bladder with aldosterone showed increased incorporation of [3H]-phenylalanine into peptides with both endogenous messenger RNA (mRNA) and exogenous mRNA (i.e., poly(uridylic acid) in an in vitro assay. Inhibition of reinitiation with NaF or poly (AUG) reduced the rate of amino acid polymerization by 45% but the aldosterone to control ratio remained significantly high. These results imply an increase in active ribosomes and perhaps in endogenous mRNA content. Our findings, however, do not distinguish between a steroid-dependent increase in the total number of active ribosomes and an increase in translational activity per ribosome.

Aldosterone↗
PubMed1974

Studies on the form and synthesis of messenger ribonucleic acid in the rat ventral prostate gland, including its tissue-specific stimulation by androgens.

1. When prostate polyribosomes are labelled with radioactive precursors in vivo and subsequently dissociated with sodium dodecyl sulphate, a heterogeneous 6-15S RNA species may be identified that possesses all of the distinctive properties of mRNA. 2. Apart from the selective incorporation of 5'-fluoro-orotic acid into this 6-15S RNA component, it is bound by nitrocellulose filters under experimental conditions where only poly(A)-rich species of RNA are specifically retained. Most importantly, however, only the 6-15S RNA fraction is capable of promoting the incorporation of amino acids into peptide linkage in an mRNA-depleted cell-free system derived from ascites-tumour cells. 3. With the development of a simpler method for labelling the total RNA fraction of the prostate gland in vitro, the poly(A)-enriched RNA fraction may be readily isolated by adsorption and elution from oligo(dT)-cellulose. The synthesis of the poly(A)-enriched 6-15S RNA fraction is stringently controlled by androgens in a highly tissue- and steroid-specific manner. 4. From an analysis of the proteins synthesized in the ascites cell-free system in the presence of the poly(A)-rich RNA fraction, it appears that protein synthesis in the prostate gland is stimulated in a rather general way, even during the earliest phases of the androgenic response. This conclusion may require modification when more specific means of analysis are available than those used in the present investigation. 5. The implications of these findings to the mechanism of action of androgens are discussed.

Amino Acids↗
PubMed1974

Kinetics of RNA labeling in toad bladder epithelium: effects of aldosterone and related steroids.

Sucrose gradient analysis revealed an aldosterone-dependent increase in the incorporation of [(3)H]uridine into a nonmethylated, rapidly labeled RNA that sedimented at 9-12 S. This effect was antagonized by an anti-mineralocorticoid, spironolactone (SC 9420), and was not elicited either by a glucocorticoid, cortisol, or by the inactive isomer, 17-alpha-isoaldosterone. These findings are in accord with the inference that mRNA is induced by aldosterone during the latent period and that the induced mRNA mediates the action on sodium transport.

Aldosterone↗
PubMed1973

The control of the form and function of the ribosomes in androgen-dependent tissues by testosterone.

1. The ribosome content of the rat ventral prostate gland is controlled by the concentrations of circulating androgens and the polyribosomal complement of the total population of ribosomes is acutely dependent on androgenic stimulation. After the administration of testosterone to castrated rats in vivo, there is a pronounced increase in the amounts of heavy (150-240S) polyribosomes. 2. These results are consistent with a pronounced increase in the mRNA and rRNA content of the prostate gland after the administration of testosterone in vivo. 3. From studies conducted both in vitro, the heavy prostate polyribosomes formed after androgenic stimulation are particularly active in protein synthesis. 4. The androgen-stimulated increase in the formation of prostate polyribosomes has a mandatory requirement for sustained RNA and protein synthesis. 5. Since the androgen-mediated increase in prostate polyribosomes may also be suppressed by the concomitant administration of certain anti-androgenic steroids in vivo, the response in polyribosome formation is probably initiated by the binding of a metabolite of testosterone, 5alpha-dihydrotestosterone, in the prostate gland. 6. The relevance of these findings to the pronounced increase in protein synthesis in androgen-dependent tissues after hormonal stimulation is discussed.

Animals↗
PubMed1972

Further studies on the stimulation of protein synthesis in androgen-dependent tissues by testosterone.

1. By using centrifugation through a discontinuous sucrose gradient, four microsomal fractions are obtained from the prostate gland. 2. Administration of androgens to castrated rats stimulates protein synthesis in all fractions, particularly in the heavy rough fraction. 3. Androgens also increase the content of protein, RNA and phospholipid in the heavy rough fraction. 4. Time-course experiments in vivo show that androgens induce a rapid increase in the synthesis of ribosomal precursor RNA preceding the synthesis of new microsomal fraction and the increase in protein synthesis.

Animals↗
PubMedDate not supplied

Chronic ethanol has region-selective effects on Egr-1 and Egr-3 DNA-binding activity and protein expression in the rat brain.

This study focused on the DNA-binding activity and protein expression of the transcription factors Egr-1 and Egr-3 in the rat brain cortex and hippocampus after chronic or acute ethanol exposure. DNA-binding activity was reduced in both regions after chronic ethanol exposure and was restored to the level of the pair-fed group at 16 h of withdrawal. Cortical Egr-1 protein levels were not altered by chronic ethanol exposure but increased 16 h after withdrawal, thus mirroring DNA-binding activity. In contrast, Egr-3 protein levels did not undergo any change. There was no change in the level of either protein in the hippocampus. Immunohistochemistry revealed a region-selective change in immunopositive cells in the cortex and hippocampus. Finally, an acute bolus dose of ethanol did not affect Egr DNA-binding activity and ethanol treatment did not alter the DNA-binding activity or protein levels of the transcription factor Sp1. These observations suggest that chronic exposure to ethanol has region-selective effects on the DNA-binding activity and protein expression of Egr-1 and Egr-3 transcription factors in the rat brain. These changes occur after prolonged ethanol exposure and may thus reflect neuroadaptive changes associated with physical dependency and withdrawal. These effects are also transcription factor-selective. Clearly, protein expression is not the sole mediator of the changes in DNA-binding activity and chronic ethanol exposure must have effects on modulatory agents of Egr DNA-binding activity.

Animals↗
PubMedDate not supplied

Glutamate-mediated transmission, alcohol, and alcoholism.

Glutamate-mediated neurotransmission may be involved in the range of adaptive changes in brain which occur after ethanol administration in laboratory animals, and in chronic alcoholism in human cases. Excitatory amino acid transmission is modulated by a complex system of receptors and other effectors, the efficacy of which can be profoundly affected by altered gene or protein expression. Local variations in receptor composition may underlie intrinsic regional variations in susceptibility to pathological change. Equally, ethanol use and abuse may bring about alterations in receptor subunit expression as the essence of the adaptive response. Such considerations may underlie the regional localization characteristic of the pathogenesis of alcoholic brain damage, or they may form part of the homeostatic change that constitutes the neural substrate for alcohol dependence.

Alcoholism↗
PubMedDate not supplied

Comparison of the formation of proteins modified by direct and indirect ethanol metabolites in the liver and blood of rats fed the Lieber-DeCarli liquid diet.

It has been proposed that proteins modified by ethanol metabolites, such as acetaldehyde (AcH) or alpha-hydroxyethyl radicals (HER) may be an important step in the aetiology of alcoholic liver disease. Furthermore, it has also been suggested that these modified proteins may act as a marker of ethanol intake. In this study, we have measured the generation of various types of modified proteins in the liver and blood of ethanol-fed rats. Multiple types of protein modification were observed in the livers of the ethanol-fed rats. In each case, the level of modification increased over the first 6 weeks of ethanol feeding, but reached a plateau by 10 weeks. In contrast to the liver, elevated levels of proteins modified by malondialdehyde were not seen in the plasma of ethanol-fed animals, whereas elevated levels of modification due to AcH and HER were observed. In haemolysates from these animals, only modification due to AcH was seen. Further investigation of the modification of plasma proteins showed that albumin, a protein produced in the liver, carried all the types of modification investigated, whereas immunoglobulin G, a protein derived from an extra-hepatic source, only carried modifications due to acetaldehyde. This study demonstrates for the first time that modification of plasma proteins by ethanol metabolites can occur at both intra- and extra-hepatic sites.

Acetaldehyde↗