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Biomedical subjects

R N Kurl

Publications and source records attributed to R N Kurl.

At least 37 records · Page 2Linked to original sources
PubMed1988

Effect of tryptophan on toxic cirrhosis induced by intermittent carbon tetrachloride intoxication in the rat.

The effects of the administration of tryptophan on toxic cirrhosis induced by intermittent carbon tetrachloride (CCl4) intoxication in the rat were investigated. Rats received CCl4 (0.45 ml/100 g body wt ip) twice weekly for 10-14 weeks. Tryptophan (30 mg/100 g body wt) by stomach tube was administered 1 hr before killing. Tryptophan improved hepatic polyribosomal aggregation and [14C]leucine incorporation into protein in vitro of control rats as well as long-term CCl4-treated rats that had developed toxic cirrhosis. However, the effects were more marked in control than in experimental rats. Tryptophan administration induced an increase in labeled nuclear RNA release in vitro and a decrease in labeled tryptophan binding to nuclear protein in vitro of livers of rats receiving long-term CCl4 and of control rats. The results indicate that the stimulatory effects of a single administration of tryptophan in toxic cirrhotic livers are similar to, but somewhat less than, those which occur in livers of normal, control rats.

Animals↗
PubMed1987

Effect of tryptophan on isolated hepatocytes of rats.

The addition of tryptophan to adult rat hepatocyte cultures stimulated DNA synthesis. The increase in DNA synthesis as measured by 3H-thymidine incorporation into DNA was observed on treatment of the cultures with tryptophan for 48 h but also as short as for 6 h in comparison with control cultures. An increase was also apparent at 30 h which was maintained for up to 48 h post treatment with tryptophan. The increase in DNA synthesis by tryptophan cannot be attributed to cell injury or to increased DNA degradation. Of the degradative enzymes added after harvesting the hepatocytes, only DNase decreased incorporation of 3H-thymidine. The observed effect was specific for tryptophan since treatment with kynurenine, isoleucine, methionine or serine failed to show a significant effect. Pretreatment of cultured hepatocytes with hydroxyurea prevented the tryptophan stimulated increase in DNA synthesis suggesting that the latter was due to replicative and not to reparative DNA synthesis. Experiments performed with the addition of diethylnitrosamine also alluded to tryptophan's role in replicative DNA synthesis. The mechanism of tryptophan-induced DNA synthesis is discussed.

Animals↗
PubMed1986

A cytosolic binding protein for 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) in the uterus and deciduoma of rats.

The cytosol fraction of the uterus of proestrous rats and of deciduoma specifically binds 3H-2,3,7,8-tetrachlorodibenzo-p-dioxin (3H-TCDD). The 3H-TCDD binding protein has a sedimentation coefficient of 9 S in low ionic 10-40% sucrose density gradients. The binding of 3H-TCDD in the 9 S region is abolished by a 500-fold molar excess of unlabeled benzo[a]pyrene, 3-methylcholanthrene or by a 100-fold molar excess of unlabeled TCDD. Incubation of the binding protein with TCDD in amounts in excess of 500 nM causes aggregation of the TCDD binding protein. Neither estradiol, progesterone, testosterone, dihydrotestosterone nor cortisol competed with TCDD in binding to this 9 S protein. The decidual tissue contains two binding components for TCDD as shown by Scatchard analysis. One of the components has a high affinity for TCDD (Kd = 1.68 nM) and is saturable. The number of binding sites is about 75 fmol/mg protein. The TCDD binding protein eluted through a DEAE-cellulose column using a gradient of 0.25 M KCl. The binding of estradiol and progesterone to their respective receptors was not affected by TCDD or by other polycyclic aromatic hydrocarbons as shown by sucrose density gradients and by microtiter competition assays. These results suggest that TCDD acts by binding to its own receptor system in the target tissue and not by competing with estrogen or progesterone for binding to their receptors. The possible role of the receptor in teratogenesis is discussed.

Animals↗
PubMed1986

Characterization and control of cytosolic binding proteins for 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) in the rat lung.

The presence of a high-affinity, low-capacity 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) binding complex was demonstrated in cytosol from rat lung. When analyzed by sucrose density gradient centrifugation, the 3H-TCDD binding complex sedimented at 8-9 S and 6.5 S in low and high ionic media, respectively. The apparent dissociation constant (Kd) of the 3H-TCDD binding complex was determined to be 2.9 nM and the number of binding sites (Bmax) was approximately equal to 69 fmol/mg of cytosolic protein. The entity was sensitive to heat and to proteases but not to DNAse or RNAse, indicating that it was a protein. An excess of unlabeled TCDD, benzo[a]pyrene, 3-methylcholanthrene, 7,12-dimethylbenzo[a]anthracene and 9-hydroxy ellipticine all competed with 3H-TCDD for binding. A single injection of either benzo[a]pyrene (20 mg/kg body weight) or 9-hydroxy ellipticine (20 mg/kg body weight) had different effects on the concentrations of 3H-TCDD binding proteins in the lung but none of the chemicals had a significant effect on the synthesis of surfactant as assayed by total phospholipid content, 72 h after the injection.

Animals↗
PubMed1985

Characterization of a factor that can prevent random transcription of cloned rDNA and its probable relationship to poly(ADP-ribose) polymerase.

A factor which eliminated nonspecific transcription of cloned rat rDNA was extensively purified from rat mammary adenocarcinoma ascites cells by successive fractionations on DEAE-Sephadex and heparin-Sepharose columns. The fractions containing RNA polymerase I (HS-B) and fractions eluting thereafter (HS-C) from the heparin-Sepharose column were pooled separately. Addition of HS-C to HS-B prevented random transcription of rDNA and yielded an accurate rDNA transcript with negligible non-specific transcription. The factor was essentially homogeneous and corresponded to Poly(ADP-ribose) polymerase with respect to molecular weight, dependence on DNA for its activity and its ability to undergo auto ADP-ribosylation. The total amount of protein in the transcription assay was approximately 2 micrograms, which indicates a high degree of purity of all the factors required for specific transcription of rDNA.

Adenocarcinoma↗
PubMed1985

Accurate initiation of rat ribosomal RNA gene transcription using a fractionated nuclear extract from normal liver and a hepatoma.

Nuclear extracts obtained from normal rat liver and from Morris hepatoma 3924A were fractionated by DEAE-Sephadex chromatography. The fraction eluted with 175 mM (NH4)2SO4 (DE-B), which contains greater than 90% of RNA polymerase I activity, supported accurate transcription of cloned rat rDNA. A similar fraction obtained from the cytosol had all of the factors required for rDNA transcription. However, its transcriptional activity was at most one-sixth that of the corresponding nuclear fraction, as determined by the amount of protein needed to produce a similar quantity of the transcript. Unfractionated nuclear or cytosol preparations did not yield an accurate transcript. Optimal KCl and magnesium concentrations for rDNA transcription were 60 mM and 5-7.5 mM, respectively. The extent of transcriptional activity was in the following order: hepatoma nuclear fraction DE-B greater than whole cell extract derived from rat mammary adenocarcinoma cells much greater than normal liver fraction DE-B. The hepatoma preparation produced at least 10 times the amount of transcript produced by the corresponding liver nuclear preparation. Transcriptional activity was proportional to the levels of RNA polymerase I and to the rate of rRNA synthesis in these tissues.

Animals↗
PubMed1985

A metabolite of riboflavin binds to the 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) receptor.

A cytosolic receptor to which TCDD binds with high affinity has been reported in a number of rodent tissues, but the endogenous/natural ligand is not known. However, lumichrome, a metabolite of riboflavin, competed with 3H-TCDD for binding to the cytosolic receptor. Neither riboflavin nor riboflavin-5-phosphate competed for binding to the TCDD receptor in rat liver cytosol. Lumichrome is the first known endogenous chemical in the rat to exhibit affinity for the TCDD receptor.

Animals↗
PubMed1985

Control of 2,3,7,8-tetrachlorodibenzo-p-dioxin binding protein(s) in the hamster kidney.

Hamster renal cytosol binds [3H]-2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) with high specificity. Sucrose density gradient centrifugation revealed two binding entities- one with a low sedimentation coefficient of 4-5 S which was displaced by neither TCDD nor other polycyclic aromatic hydrocarbons (PAHs) and another with a high sedimentation coefficient of 7-8 S which was displaced by TCDD, benzo[a]pyrene (BP), 2-methylcholanthrene (MC), and 7,12-dimethylbenzo[a]anthracene (DMBA) but not by estradiol (E), progesterone (P), cortisol (F), testosterone (T), 5 alpha-dihydrotestosterone (DHT), or methyltrienolone (R-1881), a synthetic androgen. Cytosol from intact male hamsters showed maximum binding of labelled TCDD to the 7-8 S binding site. Castration or hypophysectomy reduced this binding. Pretreatment with DMBA increased binding, whereas diethylstilbestrol (DES) decreased binding. Sex difference was observed in the binding capacity of renal cytosol. This is the first report of endocrine control over TCDD binding and its modulation by other PAHs and steroids.

9,10-Dimethyl-1,2-benzanthracene↗
PubMed1984

Phosphorylation of purified glucocorticoid receptor from rat liver by an endogenous protein kinase.

Glucocorticoid receptor was purified from rat liver cytosol using a dexamethasone affinity column. The receptor thus purified displayed a single protein band when subjected to SDS-polyacrylamide gel electrophoresis. It had a molecular weight of 90,000 which was consistent with the reported value for other glucocorticoid receptor preparations. Incubation of the purified preparation with [gamma 32P] ATP and Mg2+ resulted in transfer of [32P] to the receptor protein indicating the presence of an endogeneous protein kinase activity capable of phosphorylating the receptor molecule. Phosphorylation of the glucocorticoid receptor by the endogenous protein kinase might serve as a direct mechanism for the activation of the receptor.

Adrenalectomy↗
PubMed1984

A purified fraction containing RNA polymerase I that can accurately transcribe rat ribosomal RNA gene.

Transcription of a cloned rat rDNA containing the transcription initiation region was studied using purified fractions derived from whole cell extract. The cell extract obtained from rat mammary adenocarcinoma cells was fractionated successively on DEAE-Sephadex and heparin-Sepharose columns. The fractions eluted by 175 mM (NH4)2SO4 (DE-B) from the DEAE-Sephadex column and those eluting with RNA polymerase I on the subsequent heparin-Sepharose column (HS-B) were used in a run-off transcription assay using Xho I-cleaved cloned rat rDNA. Both fractions that contained greater than 90% of the RNA polymerase I activity produced the anticipated 635-nucleotide-long transcript. Fractionation of fraction DE-B on a phosphocellulose column also resulted in a RNA polymerase I-containing complex that, by itself, could transcribe the rDNA accurately. The major polypeptides obtained after NaDodSO4/PAGE of fraction HS-B had Mr values of 190,000, 120,000, 65,000, 42,000, and 25,000, corresponding to the major subunits of RNA polymerase I. Purified nuclear RNA polymerase I did not produce the correct transcript. These studies have shown that a purified fraction containing RNA polymerase I from whole cells can support accurate transcription of rDNA.

Adenocarcinoma↗
PubMed1983

Melanocyte-stimulating hormone and prolactin secretion in cell culture of estrogen-induced pituitary tumors in the hamster.

Pituitary cells from hamsters bearing diethylstilbestrol induced renal adenocarcinomas were cultured in vitro. Dispersed cells in plastic dishes were viable for up to two weeks in Dulbecco's modified Eagle's medium supplemented with 17.5% of 6:1 horse serum to fetal calf serum. The secretion of alpha-melanocyte stimulating hormone and prolactin into the medium were measured by radioimmunoassay. The concentrations of both were elevated by day 3 in the medium from the hyperplastic pituitaries obtained from the estrogen treated, tumor bearing hamsters. Neither DES (10(-8)M) nor tamoxifen (10(-7)M) influenced the secretion of either hormone and neither altered either cell number or DNA synthetic activity as measured by thymidine incorporation. The secretion of hormones and the growth of the pituitary cells were, however, decreased by charcoal treatment of the serum. The results suggest that the elevation of serum alpha-MSH and prolactin observed in DES implanted hamsters is due to pituitary secretion of the hormones but that DES probably does not act directly on the pituitary to control the secretion.

Adenocarcinoma↗
PubMed1983

Action of testosterone on RNA synthesis in the rat uterus.

The effect of testosterone on transcription was investigated in the uterus of the ovariectomized rat. Testosterone rapidly depressed both RNA polymerase A and B activities for up to 2 h after administration. Thereafter, RNA polymerase A activity increased, reaching control values at 4 h, and remained constant. On the contrary, RNA polymerase B activity remained depressed at 6 and 18 h after the administration of testosterone. The cellular entities or mechanisms which testosterone uses to alter transcription in the cell nucleus remain to be determined.

Animals↗
PubMed1983

Further characterization of the dioxin receptor in the rat deciduoma.

The rat deciduoma cytosol contains a protein which binds radio-labeled dioxin and sediments as an 8-9 S entity on 10-40 per cent (w/v) sucrose density gradients. Incubation of cytosol with [3H]-TCDD at 37 degrees C for ten minutes shifted the sedimentation coefficient from 8-9 S to 4-5 S, whereas incubation of cytosol at room temperature for 30 minutes has no effect on the S value. At high concentrations of [3H]-TCDD the dioxin receptor complex tends to aggregate. Cytosol eluted with 0.25 M KCl from a DEAE cellulose column and complexed with [3H]-TCDD sedimented at 9S in low ionic strength buffer, but sedimented slower (5S) in the presence of 0.4 M KCl in the buffer. The effect of salt and temperature on the sedimentation behavior is compared with that of steroid receptors. The usage of DEAE cellulose column chromatography in purification of the receptor complex is also discussed.

Animals↗
PubMed1982

Differential effect of 2,3,7,8-tetrachlorodibenzo-p-dioxin on nuclear RNA polymerase activity in the rat liver and thymus.

AN increase in RNA polymerase B activity in hepatic cell nuclei was observed after a single intravenous injection 1 microgram/rat of TCDD. The enzyme activity was about above the control value at 1 hr after injection and then declined rapidly. A secondary increase was evident at 24 hr. Prior to the secondary increase in RNA polymerase B activity, there was an increase in RNA polymerase A activity which was about 125% above the control value. The initial increase in RNA polymerase B activity was sensitive to both alpha-amanitin and actinomycin D injected min before TCDD administration. In the thymus, an increase in RNA polymerase B activity was observed 4 hr after injection (25% above control value) but thereafter it declined and at 24 hr it was about 30% below the control value. RNA polymerase A activity was inhibited as early as 1 hr after injection but had returned to the control value at 4 hr and then paralleled RNA polymerase B activity. Thus, TCDD stimulated RNA synthesis in the rat liver but the drug inhibited RNA synthesis in the rat thymus. Since these effects were detected as early as 1 hr after TCDD administration it is proposed that TCDD action requires transcriptional response and that this response may represent a primary site of TCDD action in the cell. The cellular entities or mechanisms which TCDD uses to alter transcription in the cell nucleus remain to be determined.

Animals↗
PubMed1982

High-affinity binding of 2,3,7,8-tetrachlorodibenzo-p-dioxin in cell nuclei from rat liver.

The intranuclear binding of radioactive 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) in rat liver has been studied both in vivo and in vitro. Following the intravenous administration of [1,6-3H]TCDD, a maximum uptake by cell nuclei could be observed at 2 h after injection with a concurrent decrease in the cytosolic uptake. Using linear sucrose density gradient centrifugation, dextran-coated charcoal adsorption assay, DEAE-Sepharose ion-exchange chromatography, competition, enzymatic and saturation studies, a high-affinity binding protein for TCDD in liver cell nuclei could be demonstrated both in vivo and after an exchange in vitro of intravenously administered unlabelled 2,3,7,8- tetrachlorodibenzofuran (TCDBF) for [3H]TCDD. Sucrose density gradient analysis showed a size of 4-5 S for both the cytosolic and nuclear TCDD binding entity. The specific binding of [3H]TCDD to nuclear components was heat labile and saturable and had an equilibrium dissociation constant of 1.05 nM. Based on a differential susceptibility to specific hydrolases, i.e. DNAase, RNAase, trypsin and pronase, the binding entity appears to be a 4-5 S salt-extractable protein.

Animals↗
PubMed1982

Progesterone receptors in human endometrium.

Progesterone receptor levels were determined in the cytoplasm obtained from endometrial tissue of 21 patients. In normal women, the levels of progesterone receptors were related to the stage of the menstrual cycle with highest levels occurring in the early secretory stage. In the non-secretory endometrium 5 out of 11 patients had no detectable progesterone receptors. The absence of progesterone receptors may be related to the distribution of non-responders to progestogen therapy.

Adult↗
PubMed1981

Effects of dioxane on RNA synthesis in the rat liver.

The effect of dioxane on transcription was investigated. On intravenous administration of the lower dose (10 mg/rat) both RNA polymerase A and B activities were initially depressed but recovered after 1 h. A secondary effect on RNA polymerase A was observed but the enzyme activity recovered at 24 h when it was about 35% above control values whereas RNA polymerase B activity had achieved control values. With a higher dose (100 mg/rat) the pattern was similar but the suppressive effects were more pronounced - RNA polymerase B activity never attained control values during the times studied whereas there was no increase in RNA polymerase A activity at 24 h.

Animals↗