scholarly journals Control of ovarian cholesterol ester biosynthesis

1973 ◽  
Vol 132 (2) ◽  
pp. 313-321 ◽  
Author(s):  
A. P. F. Flint ◽  
D. L. Grinwich ◽  
D. T. Armstrong
Keyword(s):  
Luteinizing Hormone ◽  
Cyclic Amp ◽  
Cholesterol Ester ◽  
Cholesterol Esterase ◽  
Interstitial Tissue ◽  
Steroid Synthesis ◽  
Precursor Pool ◽  
Ester Synthetase

1. Experimental evidence is presented for a role of progesterone and 20α-hydroxypregn-4-en-3-one as inhibitors of cholesterol ester synthetase in the acute depletion of ovarian cholesterol ester after trophic stimulation. 2. Luteinizing hormone in vitro decreased by 84% the rate of esterification of cholesterol with added [14C]oleate by slices of rabbit ovarian interstitial tissue; this effect was mimicked by cyclic AMP (adenosine 3′:5′-cyclic monophosphate) in vitro, and occurred without large changes in precursor pool sizes or membrane permeability. 3. Cyclic AMP was shown to have no direct effect on cholesterol ester synthetase or cholesterol esterase in cell-free extracts of rabbit ovarian interstitial tissue, but decreased the activity of cholesterol ester synthetase (not that of cholesterol esterase) in extracts prepared from slices previously incubated with it. 4. The inhibitory effect of cyclic AMP on esterification of cholesterol with added [14C]-oleate was prevented by both cycloheximide and aminoglutethimide phosphate (which also inhibited steroid synthesis in response to cyclic AMP). 5. Cyclic AMP raised the intracellular concentrations of progesterone and 20α-hydroxypregn-4-en-3-one in incubated slices by factors of 2.8 and 3.9 respectively. 6. Cycloheximide and aminoglutethimide phosphate administered in vivo blocked cholesterol ester depletion in response to luteinizing hormone in rats; in these ovaries cycloheximide and aminoglutethimide phosphate decreased the concentrations of progesterone and 20α-hydroxypregn-4-en-3-one and luteinizing hormone raised them. 7. Progesterone and 20α-hydroxypregn-4-en-3-one added to cell-free extracts of rabbit ovarian interstitial tissue in vitro (at concentrations comparable with those found in incubated slices) inhibited cholesterol ester synthetase by up to 85%. 8. The results are discussed with reference to the acute control of cholesterol ester concentrations in the ovary and adrenal cortex.

scholarly journals Activities of enzymes responsible for steroid biosynthesis and cholesterol ester metabolism in rabbit ovarian interstitial tissue and corpora lutea. A comparison of enzyme activities with flow rates

1973 ◽  
Vol 132 (2) ◽  
pp. 301-311 ◽  
Author(s):  
A. P. F. Flint ◽  
D. T. Armstrong
Keyword(s):  
Cholesterol Ester ◽  
Cholesterol Esterase ◽  
Interstitial Tissue ◽  
Corpora Lutea ◽  
Chain Cleavage ◽  
Cleavage Enzyme ◽  
Storage Granules ◽  
Ester Synthetase

A method involving the use of isolated cholesterol ester-storage granules as substrate is described for the assay of cholesterol esterase in rabbit ovarian tissues. Activities of cholesterol esterase 100–200-fold higher than those previously reported in ovarian tissues were measured by using this method. In addition to that of cholesterol esterase, activities of cholesterol ester synthetase, cholesterol side-chain cleavage enzyme and 3β-hydroxy steroid dehydrogenase were determined in rabbit ovarian interstitial tissue and corpora lutea. Activities of these enzymes are in general compatible with the flows through them measured under a variety of conditions both in vivo and in vitro. It is concluded that, in the rabbit ovarian tissues investigated, these enzymes are capable of catalysing the conversions usually attributed to them.

REGULATION OF STEROID SYNTHESIS IN RABBIT OVARIAN HOMOGENATES BY LUTEINIZING HORMONE, CYCLIC AMP AND PYRIDINE NUCLEOTIDES

1972 ◽  
Vol 52 (1) ◽  
pp. 147-159 ◽  
Author(s):  
V. SCOON ◽  
PATRICIA W. MAJOR
Keyword(s):  
Luteinizing Hormone ◽  
Cyclic Amp ◽  
Reduced Form ◽  
Interstitial Tissue ◽  
Pyridine Nucleotides ◽  
Steroid Synthesis ◽  
Luteal Tissue

SUMMARY Luteinizing hormone and cyclic AMP stimulate synthesis of progestational steroids in rabbit ovarian homogenates. Both progesterone and 20α-hydroxypregn-4-en-3-one are produced and, in contrast to ovarian slice preparations, progesterone is the predominant steroid formed. Steroid synthesis in homogenates from interstitial tissue is stimulated to a greater extent than in homogenates of luteal tissue. Synthesis of these steroids by the homogenates can also be increased by addition of NADP+ and NADPH. The amount and nature of the steroid synthesized varies with the amount of cofactor added and with its form (oxidized or reduced). When both forms of cofactor were added to the system in vitro, the ratio of the oxidized to the reduced form appeared to be important in determining the form of the steroid produced.

scholarly journals Isolation and properties of cholesterol esterstorage granules from ovarian tissues

1973 ◽  
Vol 134 (2) ◽  
pp. 399-406 ◽  
Author(s):  
David T. Armstrong ◽  
Anthony P. F. Flint
Keyword(s):  
Endoplasmic Reticulum ◽  
Luteinizing Hormone ◽  
Cholesterol Ester ◽  
Microsomal Fraction ◽  
Cholesterol Esterase ◽  
Interstitial Tissue ◽  
Unesterified Cholesterol ◽  
Rat Ovary ◽  
Soluble P

Cholesterol ester-storage granules were isolated from luteinized rat ovary and rabbit ovarian interstitial tissue by centrifugal flotation and were investigated with regard to their structure and function. Cholesterol ester, protein, phospholipid and unesterified cholesterol accounted for the dry weight of granules from luteinized rat ovary. The protein and the phospholipid were resistant to removal by washing. Substrate specificities of nucleotide phosphatase and specific radioactivities of lipid-soluble P (determined after administration of [32P]Piin vivo) were the same in granules and in a microsomal fraction from the same tissue. After administration of [32P]Piin vivo, luteinizing hormone increased the specific radioactivity of lipid-soluble P in granules, mitochondria and the microsomal fraction. Since granules did not swell in hypo-osmotic media, whereas microsomal particles did, it is suggested that adherent phospholipid and protein in granule suspensions is unlikely to result from contamination with endoplasmic reticulum. Luteinizing hormone administered in vivo increased the phospholipid and unesterified cholesterol contents of isolated granules relative to their cholesterol ester content, and also tended to raise their protein content. This treatment decreased the ability of isolated granules to act as a substrate for cholesterol esterase in vitro and increased the activity of cholesterol esterase. Cycloheximide in vivo also raised the unesterified cholesterol/cholesterol ester ratio of isolated granules, and when administered with luteinizing hormone acted synergistically to bring about a further increase. These results are considered compatible with evidence obtained by microscopy which suggests that granules may be surrounded by a membrane, that they arise by pinching off from the endoplasmic reticulum, and that they shrink on trophic stimulation of the tissue.

ACTION OF KI, THYROXINE AND CYCLIC AMP ON [3H]URIDINE INCORPORATION INTO THE RNA OF THYROID SLICES

1976 ◽  
Vol 83 (2) ◽  
pp. 313-320 ◽  
Author(s):  
Mario A. Pisarev ◽  
Leonardo O. Aiello ◽  
Diana L. Kleiman de Pisarev
Keyword(s):  
Cyclic Amp ◽  
Inhibitory Action ◽  
Rna Synthesis ◽  
Actinomycin D ◽  
Protein Biosynthesis ◽  
Rna Degradation ◽  
Precursor Pool ◽  
Rna Precursor

ABSTRACT Potassium iodide (KI) has been shown to impair thyroid protein biosynthesis both in vivo and in vitro. The present study was performed in order to clarify its mechanism of action. Ribonucleic acid (RNA) synthesis was studied in beef thyroid slices with either [32P] or [3H]-uridine as labelled precursors. Both KI and thyroxine (T4) at 10−5 m significantly decreased RNA labelling under our conditions. In other experiments RNA degradation was examined in pulse-labelled and actinomycin D-treated slices. KI did not modify the degradation of the [3H]-RNA thus indicating that it interferes with the biosynthesis rather than with the degradation of RNA. Taking the perchloric acid soluble radioactivity as a rough index of the precursor pool the present results would indicate an action at this level. Both KClO4 and methylmercapto-imidazole relieved the gland from the inhibitory action of KI, supporting the view that an intracellular and organified form of iodine is responsible for this action. Since T4 also reproduced the effects of KI on RNA synthesis we would like to propose iodothyronines as the intermediates of this action. Cyclic AMP has been shown to stimulate thyroid protein biosynthesis. The present results demonstrate an action at the RNA level. Cyclic AMP increased both the PCA-soluble and RNA-linked radioactivity, thus suggesting an effect at the RNA precursor pool. KI at 10−5 m blocked the action of 2 mm cyclic AMP.

EFFECTS OF PITUITARY HORMONES ON PROGESTATIONAL HORMONE PRODUCTION BY THE RABBIT OVARY IN VIVO AND IN VITRO

1966 ◽  
Vol 35 (1) ◽  
pp. 53-63 ◽  
Author(s):  
J. H. DORRINGTON ◽  
R. KILPATRICK
Keyword(s):  
Ovarian Tissue ◽  
Venous Blood ◽  
Interstitial Tissue ◽  
Corpora Lutea ◽  
Hormone Production ◽  
Stimulatory Action ◽  
Steroid Synthesis ◽  
Ovine Prolactin

SUMMARY Ovine luteinizing hormone (LH) increased the output of progestational steroids (20α-hydroxypregn-4-en-3-one and progesterone) in rabbit ovarian venous blood. Similar increases were found with ovine follicle-stimulating hormone (FSH) and growth hormone, but much larger amounts were necessary. Ovine prolactin was without effect. The increased output was due to increased synthesis and not only to release of stored steroids. Synthesis of these progestational steroids was stimulated by LH incubated with rabbit ovarian tissue. The stimulation produced by FSH was probably due to contamination by LH since the log dose-response lines for LH and FSH were parallel, and FSH was approximately 100 times less active than LH. Ovine prolactin had no stimulatory activity in concentrations up to 20 μg./ml. The stimulatory action of LH was unrelated to the presence of corpora lutea. Separated corpora lutea showed only a slight response to LH, whereas the response of interstitial tissue was similar to that found with undissected ovaries. Hence LH caused progestational steroid synthesis by stimulating the ovarian interstitial tissue.

REPTILIAN OVARIAN STEROIDOGENESIS AND THE INFLUENCE OF MAMMALIAN GONADOTROPHINS (FOLLICLE-STIMULATING HORMONE AND LUTEINIZING HORMONE) IN VITRO

1974 ◽  
Vol 62 (2) ◽  
pp. 267-275 ◽  
Author(s):  
S. W. C. CHAN ◽  
I. P. CALLARD
Keyword(s):  
Luteinizing Hormone ◽  
Ovarian Function ◽  
Cholesterol Metabolism ◽  
Ovarian Tissue ◽  
Follicle Stimulating Hormone ◽  
Sampling Period ◽  
Steroid Synthesis ◽  
Stimulating Hormone

SUMMARY The synthesis of steroids from [7α-3H]cholesterol, [7α-3H]pregnenolone and [7α-3H]progesterone by lizard and turtle ovarian tissues in vitro was studied. Progesterone, 17α-hydroxyprogesterone, dehydroepiandrosterone, androstenedione, testosterone, oestrone and oestradiol were identified as products. In the turtle (Pseudemys), conversion of pregnenolone to progesterone was efficient, but transformation of progesterone to other steroids was relatively slow as indicated by the accumulation of progesterone over the incubation period. In Dipsosaurus, accumulation of radioactivity was greatest in testosterone, the quantities of which continued to increase at each sampling period. The rate of utilization of pregnenolone as a substrate was similar for the two species studied and the quantities of oestrone and oestradiol formed were lower in Pseudemys. The use of progesterone as precursor by Dipsosaurus ovarian tissue revealed a similar pattern of Δ4-steroid metabolism to that obtained with pregnenolone as precursor. The effects of addition of purified follicle-stimulating hormone (FSH) and luteinizing hormone (LH) on the metabolism of [14C]cholesterol in vitro was studied using Pseudemys follicular tissue. The pattern of cholesterol metabolism was similar to that for pregnenolone in this species. The synthesis of pregnenolone, progesterone, dehydroepiandrosterone and androstenedione in vitro was significantly enhanced in the presence of LH. Follicle-stimulating hormone had no effect on steroid synthesis except for a decrease of androstenedione formation. The stimulatory effect of LH on steroidogenesis in vitro is discussed in relation to the literature suggesting that mammalian FSH, but not LH, stimulates all phases of reptilian ovarian function when injected in vivo.

EFFECT OF LUTEINIZING HORMONE ON THE CONCENTRATION OF CYCLIC GMP IN RABBIT OVARIES

1978 ◽  
Vol 79 (2) ◽  
pp. 251-252
Author(s):  
V. V. PATWARDHAN ◽  
A. LANTHIER
Keyword(s):  
Luteinizing Hormone ◽  
Cyclic Amp ◽  
Cyclic Gmp ◽  
Ovarian Tissue ◽  
Chorionic Gonadotrophin ◽  
Human Chorionic Gonadotrophin ◽  
Direction Opposite ◽  
Stimulation Of

Laboratoire d'Endocrinologie, Hôpital Notre-Dame et Département de Medicine, Université de Montréal, Montréal, Canada (Received 28 June 1978) Cyclic AMP has been implicated as an intermediate in some of the actions of luteinizing hormone (LH) on ovarian tissues, such as stimulation of steroidogenesis (LeMaire & Marsh, 1975). Both in vitro (Marsh, Butcher, Savard & Sutherland, 1966) and in vivo (Armstrong, Dorrington & Robinson, 1976), stimulation with LH results in a rapid increase in the amount of cyclic AMP in ovarian tissues, which precedes the LH-induced increase in steroidogenesis. Recently, studies on rat ovaries (Grinwich, Ham, Hichens & Behrman, 1976; Ratner, 1976; Ratner & Sanborn, 1976) have indicated that the ovarian tissue content of cyclic GMP may also be regulated by LH, but in a direction opposite to that of cyclic AMP. In the rabbit, Goff & Major (1975) have shown that administration of human chorionic gonadotrophin (HCG) causes a biphasic increase

Transcription from a murine T-cell receptor V beta promoter depends on a conserved decamer motif similar to the cyclic AMP response element

1989 ◽  
Vol 9 (11) ◽  
pp. 4835-4845
Author(s):  
S J Anderson ◽  
S Miyake ◽  
D Y Loh
Keyword(s):  
Cyclic Amp ◽  
Regulatory Region ◽  
Cell Receptor ◽  
Transcription Activator ◽  
Mobility Shift ◽  
Response Element ◽  
Cyclic Amp Response Element ◽  
Gel Mobility Shift

We identified a regulatory region of the murine V beta promoter by both in vivo and in vitro analyses. The results of transient transfection assays indicated that the dominant transcription-activating element within the V beta 8.3 promoter is the palindromic motif identified previously as the conserved V beta decamer. Elimination of this element, by linear deletion or specific mutation, reduced transcriptional activity from this promoter by 10-fold. DNase I footprinting, gel mobility shift, and methylation interference assays confirmed that the palindrome acts as the binding site of a specific nuclear factor. In particular, the V beta promoter motif functioned in vitro as a high-affinity site for a previously characterized transcription activator, ATF. A consensus cyclic AMP response element (CRE) but not a consensus AP-1 site, can substitute for the decamer in vivo. These data suggest that cyclic AMP response element-binding protein (ATF/CREB) or related proteins activate V beta transcription.

ADR1c mutations enhance the ability of ADR1 to activate transcription by a mechanism that is independent of effects on cyclic AMP-dependent protein kinase phosphorylation of Ser-230

1992 ◽  
Vol 12 (4) ◽  
pp. 1507-1514
Author(s):  
C L Denis ◽  
S C Fontaine ◽  
D Chase ◽  
B E Kemp ◽  
L T Bemis
Keyword(s):  
Protein Kinase ◽  
Cyclic Amp ◽  
Transcriptional Activation ◽  
Growth Conditions ◽  
Dependent Protein Kinase ◽  
Inactive Form ◽  
Dependent Protein ◽  
Kinase Phosphorylation

Four ADR1c mutations that occur close to Ser-230 of the Saccharomyces cerevisiae transcriptional activator ADR1 and which greatly enhance the ability of ADR1 to activate ADH2 expression under glucose-repressed conditions have been shown to reduce or eliminate cyclic AMP-dependent protein kinase (cAPK) phosphorylation of Ser-230 in vitro. In addition, unregulated cAPK expression in vivo blocks ADH2 depression in an ADR1-dependent fashion in which ADR1c mutations display decreased sensitivity to unregulated cAPK activity. Taken together, these data have suggested that ADR1c mutations enhance ADR1 activity by blocking cAPK phosphorylation and inactivation of Ser-230. We have isolated and characterized an additional 17 ADR1c mutations, defining 10 different amino acid changes, that were located in the region defined by amino acids 227 through 239 of ADR1. Three observations, however, indicate that the ADR1c phenotype is not simply equivalent to a lack of cAPK phosphorylation. First, only some of these newly isolated ADR1c mutations affected the ability of yeast cAPK to phosphorylate corresponding synthetic peptides modeled on the 222 to 234 region of ADR1 in vitro. Second, we observed that strains lacking cAPK activity did not display enhanced ADH2 expression under glucose growth conditions. Third, when Ser-230 was mutated to a nonphosphorylatable residue, lack of cAPK activity led to a substantial increase in ADH2 expression under glucose-repressed conditions. Thus, while cAPK controls ADH2 expression and ADR1 is required for this control, cAPK acts by a mechanism that is independent of effects on ADR1 Ser-230. It was also observed that deletion of the ADR1c region resulted in an ADR1c phenotype. The ADR1c region is, therefore, involved in maintaining ADR1 in an inactive form. ADR1c mutations may block the binding of a repressor to ADR1 or alter the structure of ADR1 so that transcriptional activation regions become unmasked.

scholarly journals Overexpression of the Matrix Metalloproteinase Matrilysin Results in Premature Mammary Gland Differentiation and Male Infertility

1998 ◽  
Vol 9 (2) ◽  
pp. 421-435 ◽  
Author(s):  
Laura A. Rudolph-Owen ◽  
Paul Cannon ◽  
Lynn M. Matrisian
Keyword(s):  
Matrix Metalloproteinase ◽  
Transgenic Animals ◽  
Reproductive Organs ◽  
Mammary Development ◽  
Seminiferous Tubules ◽  
Interstitial Tissue ◽  
Wild Type ◽  
The Matrix

To examine the role of matrilysin (MAT), an epithelial cell-specific matrix metalloproteinase, in the normal development and function of reproductive tissues, we generated transgenic animals that overexpress MAT in several reproductive organs. Three distinct forms of human MAT (wild-type, active, and inactive) were placed under the control of the murine mammary tumor virus promoter/enhancer. Although wild-type, active, and inactive forms of the human MAT protein could be produced in an in vitro culture system, mutations of the MAT cDNA significantly decreased the efficiency with which the MAT protein was produced in vivo. Therefore, animals carrying the wild-type MAT transgene that expressed high levels of human MAT in vivo were further examined. Mammary glands from female transgenic animals were morphologically normal throughout mammary development, but displayed an increased ability to produce β-casein protein in virgin animals. In addition, beginning at approximately 8 mo of age, the testes of male transgenic animals became disorganized with apparent disintegration of interstitial tissue that normally surrounds the seminiferous tubules. The disruption of testis morphology was concurrent with the onset of infertility. These results suggest that overexpression of the matrix-degrading enzyme MAT alters the integrity of the extracellular matrix and thereby induces cellular differentiation and cellular destruction in a tissue-specific manner.

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