Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.1.27.5 (RNase)
17,967 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Mechanisms regulating responses of the ovine uterus to endocrine and paracrine signals during the estrous cycle and pregnancy are likely to require tissue- and cell-specific regulation of steroid hormone receptor gene expression. To determine effects of day and pregnancy status (cyclic or pregnant) on uterine estrogen receptor (ER) and progesterone receptor (PR) gene expression, ewes were hysterectomized either on Day 1 (Day 0 = estrus/mating), 6, 11, 13, or 15 of the estrous cycle (n = 3/day) or on Day 11, 13, 15, 17, or 25 of early pregnancy (n = 5/day). Steady state levels of ER and PR mRNA were determined in endometrial and myometrial tissues by slot-blot hybridization and ribonuclease protection assays, respectively, using homologous ovine ER and PR cRNA probes. Changes in spatial expression of ER and PR mRNA and protein in uterine tissue sections were determined by in situ hybridization and immunocytochemical analyses. In cyclic ewes, steady state levels of endometrial ER mRNA were highest on Day 1, declined between Days 1 and 6, and increased between Days 11 and 15. However in pregnant ewes, endometrial ER mRNA levels decreased between Days 11 and 15 and increased slightly between Days 15 and 25. In cyclic ewes, levels of myometrial ER mRNA were highest on Day 1, decreased to Day 6, and remained low thereafter. In cyclic ewes, endometrial PR mRNA levels were highest on Day 1, decreased between Days 1 and 11, and then increased between Days 13 and 15. In cyclic ewes, myometrial PR mRNA levels were highest on Day 1 and declined thereafter. Endometrial PR mRNA levels were not different between cyclic and pregnant ewes on Days 11, 13, and 15. In pregnant ewes, PR mRNA levels were low on Day 11, increased between Days 11 and 17, and decreased between Days 17 and 25. In pregnant ewes, myometrial PR mRNA levels were low and did not change between Days 11 and 25. In situ hybridization and immunocytochemical analyses revealed distinct tissue- and cell type-specific alterations in uterine ER and PR mRNA and protein expression during the estrous cycle and early pregnancy that generally paralleled overall changes in steady state levels of ER and PR mRNAs. In the endometrium, the most striking observation was that PR mRNA and protein expression disappeared from the luminal and shallow glandular epithelium between Days 6 and 13 of the estrous cycle, whereas ER mRNA and protein expression was low on Days 6 and 11 and increased between Days 11 and 15 in the luminal and shallow glandular epithelium. During early pregnancy, expression of ER and PR mRNAs, as well as ER and PR protein, was very low or absent in the luminal and shallow glandular epithelium between Days 13 and 25 of pregnancy. Moreover, ER and PR mRNA and protein were consistently present at low levels in the stroma and deep glandular epithelium in both cyclic (Days 11-15) and pregnant (Days 11-25) ewes. Collectively, results suggest that uterine ER and PR gene expression is regulated in a tissue- and cell type-specific manner during the estrous cycle and early pregnancy.
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PMID:Temporal and spatial alterations in uterine estrogen receptor and progesterone receptor gene expression during the estrous cycle and early pregnancy in the ewe. 856 11

2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) exhibits remarkably potent antiestrogenic activity. To further elucidate the role of estrogen receptor (ER) regulation in this response, we examined the effects of exposure to TCDD in MCF-7 human breast cancer cells on ER mRNA levels by using an RNase protection assay, on ER accumulation by using an ER immunocytochemical essay (ER-ICA), and on ER function by competitive binding assays under conditions of saturating 17 beta-estradiol (E2). Comparative studies were conducted with E2 and 12-O-tetradecanoylphorbol-13-acetate (TPA), as both compounds are known to suppress ER expression. Our results indicate that 1 nM E2 and 100 nM TPA both suppress ER mRNA levels as early as 4 h after exposure and to 33.6% and 16.5% of control levels, respectively, after 72 h. In contrast, no significant effect on ER mRNA levels was attributed to exposure to 10 nM TCDD. A greater than 50% reduction in positive staining was observed by ER-ICA after 72 h exposure to 1 nM E2 and to 100 nM TPA, while only an 11% reduction in positive staining was observed with 10 nM TCDD. Specific binding of [3H]E2 under saturating conditions (10 nM E2) in whole cells was reduced by 50% in cultures exposed to 100 nM TPA, although no effect on binding was observed with exposure to 10 nM TCDD. In contrast, specific binding using subsaturating 1 nM [3H]E2 was depressed by 49% in MCF-7 cells exposed to 10 nM TCDD for 72 h. This depression was inhibited by a 1-h treatment with 5 microM alpha-naphthoflavone, which inhibits TCDD-induced, P450-mediated, E2 metabolism, and subsequent E2 depletion. In conclusion, while TPA and E2 effectively down-regulate ER expression, TCDD, under antiestrogenic conditions, has little if any effect on total ER levels in MCF-7 cells, and thus ER modulation is probably not necessary for the suppression of estrogenic activity in MCF-7 cells by TCDD.
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PMID:Effects of 2,3,7,8-tetrachlorodibenzo-p-dioxin, 12-O-tetradecanoylphorbol-13-acetate and 17 beta-estradiol on estrogen receptor regulation in MCF-7 human breast cancer cells. 865 28

Several studies in the past few years have supported the hypothesis that oxytocin (OT) is synthesized in a paracrine system within the pregnant human uterus and that this paracrine system may be an important regulator of the timing of human parturition. Using ribonuclease protection assays, we have demonstrated a three-fold increase in the rate of synthesis of OT mRNA in human decidua around the time of parturition. We also have shown that a similar increase in OT mRNA and peptide synthesis can be stimulated in vitro by physiological concentrations of estradiol. This increase is inhibited by concomitant use of the estrogen receptor (ER) blocker tamoxifen or by transcription inhibitors. Progesterone had little, if any effect. We also detected mRNAs for ER and progesterone receptor (PR) in amnion, chorion and decidua with the same relative tissue concentrations as OT mRNA. The concentrations of ER but not PR increased significantly around the time of labour onset. To determine if local OT concentrations may be regulated by changes in OT metabolism, we determined kinetic parameters for OT metabolism in decidua, chorion and placenta. [3H]tyrosyl-OT was used as substrate. Metabolites were separated using HPLC and identified using amino acid analysis and mass spectrometry. Metabolism in decidua and chorion occurred predominantly via a cytosolic post-proline endopeptidase and the activity was comparable to placenta. In microsomal fractions, cystine aminopeptidase activity predominated and placenta had significantly more activity than decidua and chorion. There were no changes in any Km or apparent vmax values around the time of parturition. These findings support the existence of a paracrine system within human decidua that involves sex steroids regulating synthesis of OT and that undergoes significant changes around the time of parturition. Changes in local OT concentrations are controlled by rates of synthesis rather than rates of metabolism.
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PMID:Synthesis and metabolism of oxytocin in late gestation in human decidua. 871 92

The environmental pollutant 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD; dioxin) induces severe reproductive defects in male rats when exposure occurs in utero and during lactation. Yet there is currently a paucity of information regarding the effects of this exposure paradigm in females. In the current study, we examine the effects of TCDD during fetal and perinatal development on the estrogen-signaling system in peripubertal female rats. Pregnant Holtzman rats were given 1 microgram/kg TCDD or vehicle control by gavage on gestational Day 15. Body weights were reduced, though not significantly, on postnatal Day 21. While ovarian and uterine wet weights were not increased by TCDD exposure, the percentage of body weight attributed to the ovary was increased significantly. Through use of ribonuclease protection and gel-shift assays, exposed females were compared with nonexposed counterparts for estrogen receptor (ER) mRNA and DNA-binding activity in the following tissues: hypothalamus, pituitary (mRNA only), uterus, and ovary. ER mRNA levels increased in the hypothalamus, uterus, and ovary, and decreased in the pituitary. The results of the DNA-binding assays paralleled the mRNA results in the uterus, while DNA-binding activity was decreased in the hypothalamus and was unchanged in ovarian protein extracts. Circulating concentrations of estrogen were significantly lower in TCDD-exposed rats than in controls. These data suggest that the decrease in serum estrogen may be a cause of the alterations in ER mRNA; the changes in ER DNA-binding activity may indicate alterations in either translation or posttranslational receptor processing. Overall, this study shows that TCDD may act systemically in this model, and these effects should not necessarily be characterized as antiestrogenic.
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PMID:In utero and lactational exposure of female Holtzman rats to 2,3,7,8-tetrachlorodibenzo-p-dioxin: modulation of the estrogen signal. 879 59

Angiogenesis is a significant prognostic factor in breast cancer, but the factors that control angiogenesis in vivo are not well defined. Multiple angiogenic polypeptides are known, and we have determined the expression of seven of these in primary human breast cancers; the relationship of expression to estrogen receptor and vascular density was also examined. Vascular endothelial growth factor (VEGF) and its four isoforms (121, 165, 189, and 206 amino acids), transforming growth factor (TGF)-beta1, pleiotrophin, acidic and basic fibroblast growth factor (FGF), placental growth factor, and thymidine phosphorylase (platelet-derived endothelial cell growth factor) were quantitated by RNase protection analysis. beta-FGF was also measured by ELISA. The estrogen receptor (ER), epidermal growth factor receptor, and vascular density were analyzed in 64 primary breast cancers. All tumors expressed at least six different vascular growth factors. VEGF was most abundant, and the transcript for the 121-amino acid form predominated. Other angiogenic factors expressed at high levels were thymidine phosphorylase and TGF-beta1. Expression of most of the angiogenic factors did not correlate with that of ER or vascular density. However, thymidine phosphorylase did, with a correlation coefficient of 0.3 (P = 0.03). There were significant associations of pleiotrophin with acidic FGF expression (P = 0.001) and TGF-beta with platelet-derived endothelial cell growth factor expression (P = 0.001). Thus, angiogenesis may involve a coordinate regulation of some vascular growth factors. High VEGF expression correlated with poor prognosis in univariate analysis (P = 0.03), as did ER and epidermal growth factor receptor expression. Basic FGF was also assessed by ELISA and was more highly expressed in tumors than normal breast tissues (median, 346 microg/ml cytosol; range, 54-1323 versus median, 149; range, 32-509; P = 0.01). Implications for therapy are that broad spectrum agents that block features common to these factors may be useful (e.g., antagonism of heparin-binding activity agents), because so many angiogenic factors are expressed. Inhibiting endothelial migration or agents directly toxic to endothelium would be of value in a combined approach to therapy.
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PMID:Expression of the angiogenic factors vascular endothelial cell growth factor, acidic and basic fibroblast growth factor, tumor growth factor beta-1, platelet-derived endothelial cell growth factor, placenta growth factor, and pleiotrophin in human primary breast cancer and its relation to angiogenesis. 904 Dec 2

Studies have shown that the pineal gland via its hormone, melatonin, induces the involution of male and female reproductive systems in seasonally reproducing animals. Melatonin has direct inhibitory effects on both hypothalamic and pituitary functions, which are also exquisitely sensitive to the feedback effects of estradiol. Since melatonin can modulate estrogen receptor (ER) expression in other tissues, immunocytochemical and ribonuclease protection analyses were used to examine the effects of 12 weeks of daily late afternoon injections of melatonin on ER protein and mRNA levels in the hypothalamus of Lak.LVG golden hamsters. Significant decreases in ER-immunoreactivity were noted in the medial preoptic area (MPOA) and bed nucleus of the stria terminalis (BNST) in response to melatonin, while other hypothalamic areas which express ER, e.g. the anterior hypothalamus, showed less dramatic changes. Hypothalamic ER mRNA was decreased in response to melatonin in both intact and ovariectomized animals by 25%. In intact, cycling female hamsters, there was a significant reduction in uterine weight after melatonin treatment. These results suggest that melatonin exerts its anti-reproductive effects in hamsters by modulating ER levels in neurons of the MPOA and BNST, thereby influencing steroid feedback mechanisms.
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PMID:Effects of melatonin on estrogen receptor expression in the forebrain of outbred (Lak.LVG) golden hamsters. 911 84

We have shown earlier that surgical human breast cancer tissue can be maintained in culture as in culture as intact tissue slices (organ culture). Because tumor organ culture ostensibly preserves the interacting network of tumor cells, stromal fibroblasts, endothelial cells and extracellular matrix, it represents a rather complex culture system. Such a system may be especially useful in preclinical trials, where the objective is to make extrapolations to the even more complex in vivo situation. A classical therapeutic target in breast cancer is the estrogen receptor, and we showed earlier that human breast cancer slices retain expression of this receptor in culture. Retinoic acid, the active form of vitamin A, is also an important (negative) growth regulator in breast cancer. In the present communication, we used in situ hybridization to monitor the expression of retinoic acid receptors in tumor slices cultured for 4 days. We show that both members of the all-trans retinoic acid and 9-cis retinoic acid receptor family (RAR and RXR, respectively) are expressed. Moreover, RNase protection analysis showed that expression of the cellular retinoic acid-binding protein type II gene, a known retinoic acid target gene, is upregulated by treatment with 1 microM all-trans retinoic acid for 2 days. These findings attest to the feasibility of using tumor organ cultures as a preclinical model for the evaluation of synthetic vitamin A derivatives (retinoids).
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PMID:Human breast carcinoma slice cultures retain retinoic acid sensitivity. 918 Oct 52

Oxytocin (OT) and its receptor (OTR) are synthesized in the endometrium and myometrium of the pregnant rat during late gestation. Both are regulated by estrogen and progesterone (P4), and tissue concentrations of both increase markedly before parturition. The P4 antagonist RU486 will induce parturition in the rat. The purpose of the present studies was to investigate changes in OT and OTR messenger RNA (mRNA) and peptide synthesis within the pregnant rat uterus during RU486-induced parturition. Pregnant rats were given a single injection of RU486 (2.5 mg/rat in oil) on day 15 of pregnancy (normal delivery occurs on day 22). Control animals received injections of oil only. Groups of animals (n = 5 in each group) were euthanized at 0, 6, 12, 24, and 48 h after injection and during labor (immediately after delivery of the first pup). Maternal serum estradiol (E2), P4 and uterine OT, and PGE2 concentrations were measured by RIA. Prostaglandin F2alpha and estrogen receptor levels were measured by enzyme immunoassay (EIA). OTR and P4 receptor (PR) were measured using radioligand-binding assays. OT, OTR, and estrogen receptor mRNAs were measured with ribonuclease protection assays. The average time to delivery, after RU486 injection, was 27.0 +/- 1.2 h. Serum E2 and P4 levels were increased slightly, but significantly, at 24 h after RU486. In controls, OT mRNA increased significantly, and this increase was blocked in the RU486 treatment group. OTR mRNA levels increased within 6 h of RU486 and remained elevated until delivery. OTR peptide was increased by 12 h. PGE2 and PGF2alpha were increased 3-fold and 16-fold, respectively, but not until after the increase in OTR had occurred. We conclude that the mechanism of action of RU486 is to inhibit the P4 suppression of OTR synthesis, allowing increased expression of OTR, which may directly stimulate myometrial contractions or act indirectly through increased synthesis of PGs.
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PMID:Effects of RU486 on estrogen, progesterone, oxytocin, and their receptors in the rat uterus during late gestation. 920 15

Estrogen receptor-like 1a (ESRL1a; same as estrogen receptor-related orphan receptors, ERR1) belongs to a subfamily of the nuclear receptor superfamily. We have previously shown that human ESRL1a modulates estrogen responsiveness of the lactoferrin gene promoter in transiently transfected endometrial carcinoma RL95-2 cells. In this study, we cloned and characterized the human ESRL1 gene. Through the fluorescence in situ hybridization method, the ESRL1 gene was localized to the centromere region of chromosome 11q12. Partial sequencing, restriction mapping, and PCR analysis revealed that the ESRL1 gene consists of seven exons and is approximately 20 kb in length. We found that the smallest exon (exon 3) contains 117 bp and the largest exon (exon 7) has 1032 bp. The smallest intron (intron 5) is only 88 bp long and the largest intron (intron 2) is 8 kb long. All introns have the conserved GT and AG dinucleotides present at the donor and acceptor sites, respectively. Like the estrogen receptor, the highly conserved DNA-binding domain of hESRL1a is encoded by exon 2 and exon 3, and the intron/exon junctions (2 and 3) are well conserved between the two genes. Primer extension analysis revealed multiple transcription initiation start sites in human uterine (HeLa, HEC, and RL95-2) cell lines. However, one major initiation start site was found by RNase protection assay. The hESRL1a mRNA is differentially expressed in various human tissues. The nucleotide sequence adjacent to the transcription start sites of the ESRL1 lacks the typical TATA and CAAT boxes but is GC rich and contains 10 consensus Sp1-binding elements and two E boxes. The region that contains these transcription factor-binding elements showed a high level of promoter activity when transiently transfected into RL95-2 cells.
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PMID:Human estrogen receptor-like 1 (ESRL1) gene: genomic organization, chromosomal localization, and promoter characterization. 928

Until recently, only a single type of estrogen receptor (ER) was thought to exist and mediate the genomic effects of the hormone 17beta-estradiol in mammalian tissues. However, the cloning of a gene encoding a second type of ER, termed ERbeta, from the mouse, rat, and human has prompted a reevaluation of the estrogen signaling system. Based on in vitro studies, the ERbeta protein binds estradiol with an affinity similar to that of the classical ER (now referred to as ERalpha) and is able to mediate the effects of estradiol in transfected mammalian cell lines. Essential to further investigations of the possible physiological roles of ERbeta, and its possible interactions with ERalpha, are data on the tissue distribution of the two ER types. Herein, we have described the optimization and use of an RNase protection assay able to detect and distinguish messenger RNA (mRNA) transcripts from both the ERalpha and ERbeta genes in the mouse. Because this assay is directly quantitative, a comparison of the levels of expression within various tissues was possible. In addition, the effect of disruption of the ERalpha gene on the expression of the ERbeta gene was also investigated using the ERalpha-knockout (ERKO) mouse. Transcripts encoding ERalpha were detected in all the wild-type tissues assayed from both sexes. In the female reproductive tract, the highest expression of ERbeta mRNA was observed in the ovary and showed great variation among individual animals; detectable levels were observed in the uterus and oviduct, whereas mammary tissue was negative. In the male reproductive tract, significant expression of ERbeta was seen in the prostate and epididymis, whereas the testes were negative. In other tissues of both sexes, the hypothalamus and lung were clearly positive for both ERalpha and ERbeta mRNA. The ERKO mice demonstrated slightly reduced levels of ERbeta mRNA in the ovary, prostate, and epididymis. These data, in combination with the several described phenotypes in both sexes of the ERKO mouse, suggest that the biological functions of the ERbeta protein may be dependent on the presence of ERalpha in certain cell types and tissues. Further characterization of the physiological phenotypes in the ERKO mice may elucidate possible ERbeta specific actions.
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PMID:Tissue distribution and quantitative analysis of estrogen receptor-alpha (ERalpha) and estrogen receptor-beta (ERbeta) messenger ribonucleic acid in the wild-type and ERalpha-knockout mouse. 934 86


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