Showing papers on "Steroid biosynthesis published in 2002"
TL;DR: It is concluded that CAR has broad functions in xenobiotic responses, some are specific to CAR but others, including induction of the important drug metabolizing enzyme CYP3A, overlap with those of PXR.
Abstract: The products of the cytochrome P450 (CYP) genes play an important role in the detoxification of xenobiotics and environmental contaminants, and many foreign chemicals or xenobiotics can induce their expression We have previously shown that the nuclear hormone receptor CAR (Constitutive Androstane Receptor, NR113) mediates the well studied induction of CYP2B10 gene expression by phenobarbital (PB) and 1, 4-bis-[2-(3, 5,-dichloropyridyloxy)] benzene (TCPOBOP) We have used the CAR knockout mouse model to explore the broader functions of this xenobiotic receptor In addition to the liver, CAR is expressed in the epithelial cells of the villi in the small intestine, and this expression is required for CYP2B10 induction in response to PB and TCPOBOP in those cells In agreement with previous observations that CAR can bind to regulatory elements in CYP3A genes, CAR is also required for induction of expression of CYP3A11 in response to both PB and TCPOBOP in liver In males, CAR is also required for induction of liver CYP2A4 expression In wild type animals, pretreatment with the CAR inverse agonist androstenol blocks the response of both the CYP2B10 and CYP3A11 genes to PB and TCPOBOP, and decreases basal CYP3A11 expression CAR is also required for the response of CYP2B10 to several additional xenobiotic inducers, including chlorpromazine, clotrimazole and dieldrin, but not dexamethasone, an agonist for both the xenobiotic receptor PXR (Pregnane X Receptor NR112) and the glucocorticoid receptor Chlorpromazine induction of CYP3A11 is also absent in CAR-deficient animals, but the responses to clotrimazole and dieldrin are retained, indicating that both of these inducers can also activate PXR (Pregnane X Receptor NR112) We conclude that CAR has broad functions in xenobiotic responses Some are specific to CAR but others, including induction of the important drug metabolizing enzyme CYP3A, overlap with those of PXR
236 citations
TL;DR: Findings strongly indicate that neurosteroids are synthesized in a region‐specific manner in the brain in the same manner as in the adrenal gland.
Abstract: Steroidogenic acute regulatory protein (StAR) is a 30-kDa protein involved in the transport of cholesterol to the inner mitochondrial membrane and thus plays a key role in steroid biosynthesis. To clarify the implications of this protein in neurosteroid biosynthesis, we examined the possible expression of a StAR transcript in the adult rat CNS and detected it. cDNA cloning and sequencing analysis revealed that two forms of StAR mRNAs are expressed in the brain in the same manner as in the adrenal gland, indicating that they are fully functional and not minor gene transcripts. An RNase protection assay quantitatively revealed that the amount of the rat StAR transcript in brain was two to three orders of magnitude lower than that in the adrenal gland. An in situ hybridization study, involving antisense riboprobes, revealed that StAR transcripts were abundant in the cerebral cortex, hippocampus, dentate gyrus, olfactory bulb, cerebellar granular layer, and Purkinje cells. Furthermore, other steroidogenic enzymes, side-chain cleavage cytochrome P-450SCC (CYP XIA1) and 3beta-hydroxysteroid dehydrogenase/delta5-delta4 isomerase (EC 1.1.1.145), were found to be coexpressed in the hippocampus, dentate gyrus, cerebellar granular layer, and Purkinje cells. These findings strongly indicate that neurosteroids are synthesized in a region-specific manner in the brain.
235 citations
TL;DR: Both naturally occurring disorders in humans and genetic manipulation in mice have demonstrated that the StAR protein is an absolute requirement in the rate-limiting step in steroidogenesis, the transfer of cholesterol into the mitochondria.
108 citations
TL;DR: It was found that overexpression of this gene significantly enhances steroid hydroxylase activity of CYP11B2 expressing fission yeast cells, and a gene was identified that codes for a protein with an amino terminal domain homologous to COX15 of Saccharomyces cerevisiae and a carboxy terminal ferredoxin domain to be named etp1 (electron-transfer protein 1).
Abstract: Mitochondrial cytochrome P450 enzymes play a crucial role in the steroid biosynthesis in human adrenals, catalyzing regio- and stereospecific hydroxylations. In search of a new model system for the study of these enzymes, we expressed the human CYP11B2 (aldosterone synthase, P450aldo) in fission yeast Schizosaccharomyces pombe. Analysis of the subcellular localization of the P450 enzyme by Western blot analysis, fluorescence microscopy, and electron microscopy demonstrated that the mitochondrial localization signal of the human protein is functional in S. pombe. The transformed yeasts show the inducible ability to convert in vivo considerable amounts of 11-deoxycortisol to cortisol and 11-deoxycorticosterone to corticosterone, 18-hydroxycorticosterone, and aldosterone, respectively. Although in mammalian cells, mitochondrial steroid hydroxylases depend for their activity on an electron transport chain that consists of two proteins, adrenodoxin and adrenodoxin reductase, no coexpression of these proteins i...
95 citations
TL;DR: It was determined that the arachidonic acid (especially lipoxygenase metabolites) and mitogen-activated protein kinase pathways were also involved in the mEGF-induced steroidogenic response, and involvement of the latter pathway was further assessed in nonsteroidogenic COS-1 cells transfected with the Elk1 trans-reporting plasmids.
Abstract: Steroid hormone biosynthesis in the adrenals and gonads is regulated by the steroidogenic acute regulatory (StAR) protein through its action in mediating the intramitochondrial transport of cholesterol. A role for epidermal growth factor (EGF) in modulating steroidogenesis has been previously determined, but the mechanism of its action remains unknown. The present investigation was designed to explore the potential mechanism of action of mouse EGF (mEGF) in the regulation of steroid biosynthesis and StAR protein expression in mLTC-1 mouse Leydig tumor cells. We show that treatment of mLTC-1 cells with mEGF significantly increased the levels of progesterone (P), StAR protein, and StAR mRNA in a time- and dose-dependent manner. The coordinate induction of P synthesis and StAR gene expression by mEGF was effectively inhibited by cycloheximide, indicating a requirement for de novo protein synthesis. Also, longer exposure of mLTC-1 cells to mEGF produced a marked decrease in LH-receptor mRNA expressio...
65 citations
TL;DR: It is demonstrated that AA and cAMP act in a highly synergistic manner to increase the sensitivity of steroid production to trophic hormone stimulation and probably do so by increasing StAR gene expression.
59 citations
TL;DR: The data demonstrate that the rodent pubertal female assay is useful for identifying potential EDs having not only estrogenic/antiestrogenic but also androgenic/antiandrogenic activities, however, further validation study is necessary to identify chemicals that operate through other action mechanisms, including steroid biosynthesis inhibitors and thyroid inhibitors.
56 citations
TL;DR: Based on the current data, a Tier I screening battery consisting of in vitro receptor binding assays, a 3-day uterotrophic assay, and a 15-day intact male assay are recommended as the preferred approach on which future validation efforts should be focused.
Abstract: In 1996, Congress passed legislation requiring the U.S. Environmental Protection Agency (EPA) to implement screening/testing strategies for endocrine-active compounds (EACs). In response, EPA convened the Endocrine Disruptor Screening and Testing Advisory Committee (EDSTAC) to advise the agency on a strategy to screen and test xenobiotics for endocrine disruption. EDSTAC completed their charter in 1998 by recommending a tiered screening and testing scheme to evaluate compounds for their potential to act as agonists or antagonists to the estrogen or androgen receptors, steroid biosynthesis inhibitors, or their ability to alter thyroid function. For Tier I, the EDSTAC-recommended screening battery comprised eight different assays, but EDSTAC also proposed two alternative batteries that were deemed worthy of further evaluation. The challenge currently confronting EPA is to choose among the Tier I screening options and then to standardize protocols, validate the assays, and determine the criteria for judging ...
55 citations
TL;DR: It was shown that TCDD disrupts steroidogenesis through its influence on the activity of enzymes involved in the steroid biosynthesis cascade and is probably independent of estrogen receptor (ER) stimulation in luteal cells.
Abstract: To characterize the action of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) during both the follicular and luteal phases of the ovarian cycle, the direct effect of TCDD was investigated in vitro using a system of primary monolayer cell culture. Granulosa and theca cells were collected from the preovulatory follicles and cultured as a co-culture, thus resembling follicles in vivo. Luteal cells were isolated from the corpora lutea collected during the midluteal phase. In both cases cells were isolated from the ovaries of animals exhibiting natural estrus cycle. Results of these experiments suggest that TCDD decreases estradiol secretion by follicular cells and progesterone secretion by luteal cells in a dose-dependent manner. It was also shown that TCDD disrupts steroidogenesis through its influence on the activity of enzymes involved in the steroid biosynthesis cascade. In luteal cells, its action is mediated via the aryl hydrocarbon receptor (AhR) and is probably independent of estrogen receptor (ER) stimulation. Endocrine disruptors that interfere with estradiol production in the follicles can act as ovulatory disruptors, and while interfering with progesterone production by luteal cells they can act as abortifacients.
44 citations
TL;DR: In this paper, the authors used differential display RT-PCR to identify genes that are rapidly induced by ACTH in the bovine adrenal cortex, reaching a maximal level after 8 h and 3 h of treatment, respectively.
Abstract: ACTH is the major trophic factor regulating and maintaining adrenocortical function, affecting such diverse processes as steroidogenesis, cell proliferation, cell migration, and cell survival We used differential display RT-PCR to identify genes that are rapidly induced by ACTH in the bovine adrenal cortex Of 42 PCR products differentially amplified from primary cultures of bovine adrenocortical cells treated with 10 nM ACTH, six identified mRNAs that were confirmed by Northern blot analysis to be induced by ACTH Four of these amplicons encoded noninformative repetitive sequences Of the other two sequenced amplicons, one encoded a partial sequence for mitochondrial manganese-dependent superoxide dismutase (SOD2), an enzyme that is likely to protect adrenocortical cells from the cytotoxic effects of radical oxygen species generated during steroid biosynthesis The second was identified as TIS11b (phorbol-12-myristate-13-acetate-inducible sequence 11b)/ERF-1/cMG, a member of the CCCH double-zinc finger protein family SOD2 induction by ACTH was independent of extracellular steroid concentration or oxidative stress SOD2 and TIS11b mRNA expressions were rapidly induced by ACTH, reaching a maximal level after 8 h and 3 h of treatment, respectively These ACTH effects were mimicked by forskolin but appeared independent of cortisol secretion Upon ACTH treatment, induction of TIS11b expression closely followed the previously characterized peak of vascular endothelial growth factor (VEGF) expression Transfection of a TIS11b expression plasmid into 3T3 fibroblasts induced a decrease in the expression of a reporter gene placed upstream of the VEGF 3'-untranslated region, indicating that TIS11b may be an important regulator of VEGF expression through interaction with its 3'-untranslated region
40 citations
TL;DR: The time course of change in the gene expression profile compared with changes in pathology and clinical chemistry shows the potential of this approach for modeling causative, predictive, and adaptive changes in gene expression during pathologic change.
Abstract: Erythropoietic protoporphyria patients can develop cholestasis, severe hepatic damage, fibrosis, and cirrhosis. We modeled this hepatic pathology in C57BL/6J and BALB/c mice using griseofulvin and analyzed 3,127 genes for alteration of expression in the liver before and during the onset of protoporphyria, cholestasis, inflammation, and hepatic fibrosis. The two mouse strains developed different levels of pathologic damage in response to the griseofulvin. Characteristic gene expression profiles could be associated with griseofulvin-induced gene expression, disruption of lipid metabolism, and the pathologic states of inflammation, early fibrosis, and cholestasis. Additionally, some genes individually indicated an alteration of homeostasis. or pathologic state; for example, fibroblast proliferation was potentially indicated by increased calcyclin (SA100a6) expression. Changes in cytochrome P450 (Cyp) gene expression were particularly pronounced, with increased expression of the Cyp2a, Cyp2b, and Cyp3a families. Decreased Cyp4a10 and Cyp4a14 expression was observed that could be associated with early pathologic change. A potential decrease in bile acid and steroid biosynthesis was indicated by the decreased expression of Cyp7b1 and Hsd3b4, respectively. DNA damage was indicated by induction of GADD45. This study illustrates how transcriptional programs can be associated with different stimuli in the same experiment. The time course of change in the gene expression profile compared with changes in pathology and clinical chemistry shows the potential of this approach for modeling causative, predictive, and adaptive changes in gene expression during pathologic change.
TL;DR: It is indicated that endogenous PBR agonist-receptor interaction is critical for hormone-induced steroidogenesis and leads for the development of potent PBR antagonists are yield leads.
Abstract: Peripheral-type benzodiazepine receptor (PBR) is an 18-kDa high-affinity cholesterol and drug ligand-binding protein involved in various cell functions, including cholesterol transport and steroid biosynthesis. To aid our investigation of the biological function of PBR, we have set out to identify functional antagonists. By screening phage display libraries, we have identified peptides that displace the high-affinity PBR benzodiazepine drug ligand, Ro5-4864 (4′-chlorodiazepam). Among these peptides, STPHSTP was the most potent (IC 50 = 10 μM). All of the isolated peptides showed a conserved motif STXXXXP. The role of these peptides in Leydig cell steroidogenesis was examined using a transducible peptide composed of the TAT domain of human immunodeficiency virus and the peptides under investigation. Synthesized peptides efficiently transduced into MA-10 Leydig cells, and the peptide TAT-STPHSTP inhibited Ro5-4864- and human chorionic gonadotropin-stimulated steroid production in a dose-dependent manner (ED 50 = 5 μM). TAT-STPHSTP behaved as a competitive PBR antagonist, which did not affect 22 R -hydroxycholesterol-supported steroidogenesis. These results yield leads for the development of potent PBR antagonists and indicate that endogenous PBR agonist-receptor interaction is critical for hormone-induced steroidogenesis.
TL;DR: In amphibians, Bufo arenarum showed that during the reproductive season, the production of progesterone, 5alpha-pregnan-3alpha,20alpha-diol, 3alpha-hydroxy-5alpha-propnan-20-one, and 5-androstane-3beta,17beta,diol as intermediate in testosterone biosynthesis increases significantly as mentioned in this paper.
TL;DR: A search for genetic disorders of the mineralocorticoid axis should be an integral part of the diagnostic work-up, particularly in young adults with hypertension, where only moderate hypertension with or without hypokalemia as the sole feature is evident.
Abstract: Abnormalities in steroid biosynthesis have been known for years to cause hypertension in some cases of congenital adrenal hyperplasia. In these patients hypertension usually accompanies a characteristic phenotype with abnormal sexual differentiation. Recently, the molecular basis of four forms of severe hypertension transmitted on an autosomal basis but without additional phenotypic features has been elucidated. All these conditions are characterized primarily by low plasma renin, normal or low serum potassium, and salt-sensitive hypertension, indicating an increased mineralocorticoid effect. These four disorders, the glucocorticoid remediable aldosteronism, the syndrome of apparent mineralocorticoid excess, the activating mutation of the mineralocorticoid receptor, and the Liddle syndrome are a consequence of either abnormal biosynthesis, metabolism, or action of steroid hormones, and are ultimately characterized by an overactivation of the epithelial sodium channel in distal renal tubules. Hyperactivity of this channel results in increased sodium reabsorption and volume expansion leading to an increase in blood pressure as well as potassium loss. With the advent of molecular biology in clinical practice, it has become evident that some genetic defect may present with a more discrete phenotype, with only moderate hypertension with or without hypokalemia as the sole feature. A search for genetic disorders of the mineralocorticoid axis should be an integral part of the diagnostic work-up, particularly in young adults with hypertension.
TL;DR: Both oestradiol and glucocorticoids influence the secretion of prolactin by novel direct and indirect mechanisms, in addition to their much better understood effects on transcription via classical intracellular steroid receptors.
Abstract: Prolactin secretion is controlled by the hypothalamus, and by circulating steroids; oestrogens stimulate, but glucocorticoids inhibit prolactin release. Lactotrophs express intracellular receptors for oestrogens, but apparently not glucocorticoids. Therefore, a genomic effect of oestrogens could be direct, but that of glucocorticoids appears to be indirect. Lactotrophs are not a homogeneous cell population: some have large irregular dense-cored vesicles, others have small round vesicles, but the functional significance of this inhomogeneity is far from clear. Oestradiol and testosterone can stimulate rapid release of prolactin selectively from type II lactotrophs characterised by small round vesicles. Progesterone and other steroids do not exert this effect, which results from a non-genomic action of oestradiol and testosterone. Glucocorticoid inhibition of secretagogue-induced prolactin secretion is mimicked by annexin 1 (lipocortin 1), a protein induced by glucocorticoids in the pituitary and many other tissues, and can be blocked by annexin 1 immunoneutralisation and antisense. Glucocorticoid inhibition of ACTH and growth hormone secretion also involves annexin 1. Pituitary annexin 1 is located in folliculo-stellate cells; these express glucocorticoid receptors, and glucocorticoids induce annexin-1 synthesis. Annexin 1 is externalised from folliculo-stellate cells in response to glucocorticoids, despite the fact that it lacks a secretory signal sequence and is not packaged in vesicles. Inhibition of annexin 1 externalisation by glyburide suggests involvement of an ABC (ATP-binding cassette) transporter in externalisation. Both oestradiol and glucocorticoids therefore influence the secretion of prolactin by novel direct and indirect mechanisms, in addition to their much better understood effects on transcription via classical intracellular steroid receptors.
01 Jan 2002
TL;DR: In this article, the authors present a model for modeling and simulation of nanoparticle and molecular nanosystems, and present an interpretation of STM images of carbon-nanotube systems.
Abstract: Preface. Photograph of Participants. Part I. Modeling and Simulation of Nanoparticle and Molecular Nanosystems. Multiscale computer simulations in physics, chemistry and biology: the example of silica J. Horbach, et al. Application of the IMOMM (Integrated Molecular Orbital Molecular Mechanics) method for biopolymers I. Komaromi, L. Muszbek. Molecular orbital simulation of semiconductor and metal clusters V. Gurin. Modeling and interpretation of STM images of carbon nanosystems G.I. Mark, et al. Carbon Nanotubes under internal pressure B.A. Galanov, et al. Part II. Nanotechnology of Nanoparticle and Molecular Nanosystems. Recognition templates for biomaterials with engineered bioreactivity B. Ratner, et al. Layer-by-layer method for immobilization of protein molecules on biochip surface G. Zhavnerko, et al. Enzyme electrodes with enzyme immobilised by sol-gel technique M. Przybyt, M. Bialkowska. Template-directed lattices of nanostructures preparation and physical properties S. Romanov. Templates for metal nanowire self-assembly M. Brust, et al. Layer-by-layer assembly of nanotubes and nanofilms from nanoparticle and polymer blocks for electronic applications N. Kovtyukhova, et al. Non-thermal plasma synthesis of nanocarbons A. Huczko, et al. A novel network structure of organometallic clusters in gas phase A. Nakajima, K. Kaya. Nanotechnology of DNA/nano-Si and DNA/Carbon nanotubes/nano-Si chips E. Buzaneva, et al. Part III. Fundamental Properties of Nanosystems. Fundamental properties and applications of fullerene and carbon nanotube systems P. Scharff. Fundamental properties of DNA: some lessons from studies on the molecular basis of drug binding M. Waring. DNA modifications by novel antitumorplatinum drugs V. Brabec. Studies on protein electron carrier complexes: adrenodoxin reductase-adrenodoxin complex in steroid biosynthesis S. Mardanyan, Y. Sargisova. Interaction of nucleic acids and lipids from tumour cells with anticancer drugs: an SEIRA spectroscopy data G. Dovbeshko, et al. Aggregation of fullerenes in pyridine/water solutions V.L. Aksenov, et al. Infrared spectrum of fullerene C60 aggregates in water solution A.A. Golub, et al. Electrical and magnetic properties of undoped fullerene polymers T. Makarova, B. Sundqvist. Direct transition in the porous nanosilicon measured by electroflectance R.Y. Holiney, et al. Part IV. Single and Assembled Molecules Experiments. Scanning probe microscopy of biomacromolecules: nucleic acids, proteins and their complexes O.I. Kisolyova, et al. Application of atomic force microscopy in protein and DNA biochips development O. Stukalov. Peculiarities of Th. Terrestris spores surface ultrastructure investigated by AFM E.H. Gromozova, et al. Part V. Multifunctional Nanosystems. Relaxation of nanostructured molecular materials under the infuence of solvent vapors Y. Shirshov, et al. Biospecific interactions on the optical transducer surface -- the base of infection diagnostics N. Starodub, et al. Thin film biotermosensors A. Shmyryeva, N. Starodub. Porous silicon as transducer for immune sensors: from theory to practice V. Starodub. A porous silicon microcavity as an optical and electrical multiparametric chemical sensor Z. Gaburro, et al. Composite silicon-based photonic crystals and light emission and sensor elements L. Karachevtseva. Optical transmission of macroporous silicon A. Remenyuk, et al. Magnetoresistive sensors and memory
TL;DR: A subset of new ligands bind the PBR (rat brain and testis) with picomolar affinity, representing the most potent ligands that have been identified to date, and elicited effects on endogenous rate of steroidogenesis in MA10 Leydig cells, having similar potency and effect as PK11195.
Abstract: In the present study, we have synthesized and tested novel pyridopyrrolo- and pyrrolobenzoxazepine derivatives, as novel and selective peripheral type benzodiazepine receptor (PBR) ligands, and their ability to modulate steroid biosynthesis has been investigated. A subset of new ligands bind the PBR (rat brain and testis) with picomolar affinity, representing the most potent ligands that have been identified to date, and elicited effects on endogenous rate of steroidogenesis in MA10 Leydig cells, having similar potency and effect as PK11195. Several compounds, differently substituted at C-7, were used as molecular yardsticks to probe the spatial dimension of the lipophilic pocket L4 in the receptor binding site.
TL;DR: In this article, the authors investigated whether neonatal exposure affects spermatogenesis in juvenile and pubertal testes, Sprague-Dawley rat pups were given various endocrine disruptors by a single injection on the day of birth at concentrations ranging between 4μM and 40mM and sacrificed on day 21 or 50 (puberty).
TL;DR: Results demonstrate that inhibition of PTP activity affected both StAR protein and mRNA synthesis and suggest that the activity of hormone-regulated PTPs is a requirement in the LH signaling cascade that results in the up-regulation of StARprotein and, subsequently, increased steroid synthesis.
Abstract: The LH signal transduction pathway features the activation of protein tyrosine phosphatases (PTPs) as one of the components of a cascade that includes other well characterized events such as cAMP-dependent protein kinase A (PKA) activation. Moreover, the action of PTPs is required to increase the rate-limiting step in steroid biosynthesis, namely the cAMP-regulated transfer of cholesterol to the inner mitochondrial membrane. Since both PKA activity and steroidogenic acute regulatory (StAR) protein induction are obligatory steps in this transfer of cholesterol, the present study was performed to investigate the role of PTPs in the regulation of PKA activity and StAR expression in response to LH/chorionic gonadotropin (CG) and 8Br-cAMP in MA-10 cells. While the exposure of MA-10 cells to the PTP inhibitor, phenylarsine oxide (PAO), did not modify PKA activity, it partially inhibited the effect of human CG and cAMP analog on StAR protein levels. Time-course studies demonstrated that PAO inhibited cAMP induction of StAR protein and mRNA. At 30 min, the effect on cAMP-stimulated StAR protein levels was a 35% inhibition, progressing to up to 90% inhibition at 120 min of stimulation. The maximal inhibitory effect on cAMP-induced StAR mRNA level was obtained at 60 min (85%). In summary, these results demonstrate that inhibition of PTP activity affected both StAR protein and mRNA synthesis and suggest that the activity of hormone-regulated PTPs is a requirement in the LH signaling cascade that results in the up-regulation of StAR protein and, subsequently, increased steroid synthesis.
23 May 2002
TL;DR: Steroid Biosynthesis: Enzymology, Integration and Control, and Aromatase: Insights into the Roles of Estrogens Revealed by Natural and Targeted Mutations.
Abstract: Steroid Biosynthesis: Enzymology, Integration and Control. Sterol Biosynthesis. Vitamin D Biosynthesis and its Disorders. Bile Acid Biosynthesis. Cholesterol Metabolism in Steroidogenic Tissues. Steroid 21-Hydroxylase. Steroid 11ss-Hydroxylase Isozymes. 3ss-Hydroxysteroid Dehydrogenase/ Isomerase Deficiency. Human 17a-Hydroxylase/17, 20-Lyase. Aromatase: Insights into the Roles of Estrogens Revealed by Natural and Targeted Mutations. 17ss-Hydroxysteroid Dehydrogenase and 5a-Reductase Deficiencies. Steroid Metabolism in Peripheral Tissues. Animal Models of Impaired Steroidgenesis. Regulation of Gene Expression by the Nuclear Receptor Family. Neurosteroids and Brain Sterols.
Patent•
24 Jun 2002TL;DR: In this paper, the authors present methods of assaying for genes, gene products and genes in pathways controlled by such genes and gene products, using RNAi and microarray analysis, that regulate lifespan in eukaryotes.
Abstract: The present invention relates to regulation of adult lifespan in eukaryotes. More particularly, the present invention is directed to methods of assaying for genes, gene products, and genes in pathways controlled by such genes and gene products, using RNAi and microarray analysis, that regulate lifespan (e.g., extend or truncate adult lifespan) in eukaryotes such as invertebrates (e.g., C. elegans), plants, and mammals, e.g., humans. For example, the present invention is directed to genes encoding components of the mitochondrial respiratory chain and genes encoding glycolysis enzymes, which are involved in lifespan regulation, and genes and gene products in pathways controlled by such genes. Other genes and gene products identified as regulating aging and aging pathways include a gene encoding a GTPase; a transcriptional activator; novel genes: llw-1, llw-2, llw-3, and llw-4; genes encoding cytochrome P450 proteins (involved in steroid biosynthesis); a melatonin synthesis gene; genes encoding insulin and insulin-like peptides; genes encoding heat shock factors; genes encoding catalases; stress-response genes; and metabolic genes. The invention further relates to methods for identifying and using agents, including small molecule chemical compositions, antibodies, antisense nucleic acids, and ribozymes, that regulate, e.g., enhance, adult lifespan via modulation of aging associated proteins; as well as to the use of expression profiles, markers, and compositions in diagnosis and therapy related to lifespan extension, life expectancy, and aging. The present invention also relates to gene therapy involving lifespan associated genes.
TL;DR: Results indicate that PTP activity participates in StAR protein induction and led us to attribute to the PKA-mediated PTP activation in steroidogenic systems a functional role, as mediator of Star protein induction.
Abstract: A key regulatory step in the steroidogenic hormones signaling pathway is the synthesis of steroidogenic acute regulatory protein (StAR). This protein facilitates the delivery of cholesterol to the inner mitochondrial membrane, the rate-limiting step in steroidogenesis. ACTH and LH pathway also includes tyrosine dephosphorylation processes. Indeed, our previous studies have demonstrated that both hormones increase protein tyrosine phosphatase (PTP) activity by a PKA-dependent mechanism and that the action of PTPs is required for the stimulation of steroid biosynthesis in adrenal and Leydig cells. In order to test the putative relationship between PTP activity and StAR protein induction in adrenocortical cells, in the present study we evaluated steroid production and StAR protein level in Y1 adrenocortical cells under PTP inhibition. Phenylarsine oxide (PAO), a powerful cell permeable PTP inhibitor, reduced ACTH-stimulated steroidogenesis in a concentration-dependent fashion. A concentration of 2.5 µM of th...
TL;DR: The results provide no evidence for an association between prostate cancer risk and CYP17 T/C polymorphism.
Abstract: CYP17 gene encodes the enzyme cytochrome p450c17alpha, which mediates two steps in the steroid biosynthesis pathway. Steroid hormones are believed to play a key role in the etiology of prostate cancer. A polymorphic T-->C transition in the 5(') promoter region of CYP17 creates an additional Sp1-type (CCACC box) promoter site (allele A2). We have evaluated the genotypic and allelic distribution of this polymorphism among 92 prostate cancer patients in order to assess risk by comparison with a population-based series of 200 healthy individuals from Brazil. Our results provide no evidence for an association between prostate cancer risk and CYP17 T/C polymorphism.
TL;DR: The changes in the maternal steroids were associated with a single common factor, which strongly correlated with all the steroids except dehydroepiandrosterone sulfate, and a scheme of steroid biosynthesis in maternal organism during the critical stages around parturition is proposed.
TL;DR: Toad CypP450c17 activity is higher in the non-reproductive period than the reproductive period, suggesting that this enzyme is an important factor in toad steroidogenic changes, and could explain the predominance of the 5-ene pathway for testosterone biosynthesis.
Abstract: In Bufo arenarum, the biosynthesis of testosterone and 5α-dihydrotestosterone takes place through a complete 5-ene pathway, 5-androsten-3β,17β-diol being the immediate precursor of testosterone. Besides androgens, testes are able to synthesise 5α-pregnan-3,20-dione and several 3α and 20α reduced derivatives. During the breeding season, steroid biosynthesis turns from androgen to C21-steroid production. As a consequence, the cytochrome P450 17-hydroxylase, C17,20-lyase (CypP450c17) could be a key enzyme in that metabolic shift. The present study demonstrates that in testes of B. arenarum, CypP450c17 co-localises with glucose-6-phosphatase in the microsomal fraction. CypP450c17 possesses more affinity for pregnenolone than for progesterone in both non-reproductive (Km=43.76±4.63 nM and 2,170±630 nM, respectively) and reproductive (Km=37.46±4.19 nM and 3,060±190 nM, respectively) seasons. These results could explain the predominance of the 5-ene pathway for testosterone biosynthesis. Toad CypP450c17 activity is higher in the non-reproductive period than the reproductive period, suggesting that this enzyme is an important factor in toad steroidogenic changes. Animals in reproductive conditions showed a significant reduction in circulating androgens. This is in agreement with the decrease in Vmax of cytochrome P450 17-hydroxylase activity, enhancing the physiological relevance of these in vitro results.
TL;DR: It is demonstrated that, in blowfly ovaries, calcium ions inhibit cAMP-stimulated steroidogenesis by activating a calmodulin-sensitive (type I) PDE.
Abstract: Calcium is frequently involved in the stimulation of steroidogenesis in gonads and endocrine glands, generally in association with cAMP. However, our present observations show that it has the opposite effect in the ovary of the blowfly Phormia regina. Our in vitro experiments first showed that extracellular calcium does not play a role during the stimulation of steroidogenesis in fly ovaries; indeed steroidogenesis was activated in vitro as efficiently in a medium with or without calcium, either by pharmacological compounds mimicking cAMP signaling or by active brain extracts. When calcium was experimentally introduced into biosynthetic cells by ionophores or liberated from internal stores by thapsigargin, it did not activate, but clearly inhibited both basal and acute steroidogenesis respectively in previtellogenic and in vitellogenic ovaries. Our experiments also demonstrated that calcium decreases cAMP concentrations in the ovaries of Phormia, by stimulating its degradation, without modifying its biosynthesis. Moreover, inhibitors of calcium-calmodulin phosphodiesterases (PDEs) increased steroid biosynthesis in vitro, whereas inhibitors of calcium-insensitive PDEs did not. These data thus demonstrate that, in blowfly ovaries, calcium ions inhibit cAMP-stimulated steroidogenesis by activating a calmodulin-sensitive (type I) PDE.
TL;DR: In steroidogenic cells the steroidogenic acute regulatory (StAR) protein plays a key role in the transport of cholesterol to the inner mitochondrial membrane, where the first step of steroidogenesis, the conversion ofolesterol to pregnenolone, takes place.
Abstract: In steroidogenic cells the steroidogenic acute regulatory (StAR) protein plays a key role in the transport of cholesterol to the inner mitochondrial membrane, where the first step of steroidogenesis, the conversion of cholesterol to pregnenolone, takes place. cAMP is a known positive regulator of StAR gene expression and steroid biosynthesis in steroidogenic cells. As some steroids, such as progesterone, can also be synthesized de novo in the central and peripheral nervous systems and display neuroprotective and neurotrophic effects, we decided to verify the effect of cAMP on StAR gene expression in cultured Schwann cells. We observed that (1) in the presence of serum, forskolin, an agent known to elevate intracellular cAMP, induced both a morphological change and proliferation of cultured Schwann cells; (2) StAR mRNA and protein were expressed in Schwann cells; (3) unexpectedly, forskolin and 8 Br-cAMP, a cell-permeant analogue of cAMP, extinguishcd StAR gene expression; and (4) this response was similar in the presence or absence of serum.
TL;DR: An unusual polar steroid is detected as a major end product of gonadal steroid biosynthesis in male and female blotched blue-tongued lizards, Tiliqua nigrolutea and is more polar than 17beta-estradiol, which was not detected, but less polar than all tri-hydroxylated estrogens to which it was compared.
TL;DR: By characterizing the regulatory systems mediating both metabolic and steroidogenic processes in normal and PCOS theca cells, it is anticipated that new information will be obtained that will provide insight into the relations between insulin resistance, hyperinsulinemia, and hyperandrogenemia in patients with PCOS.
Abstract: The cellular mechanisms underlying excess ovarian androgen production in patients with polycystic ovary syndrome (PCOS) are presently unknown. Examination of the steroid biosynthesis by theca interna cells isolated from ovaries of normal women and ovaries from women with PCOS has recently advanced our understanding of the steroidogenic abnormalities that result in increased androgen biosynthesis in PCOS. In the past few years, the combined data derived from studies utilizing freshly isolated and propagated theca cells have established that both progestin and androgen biosynthesis are increased in theca cells obtained from PCOS ovaries. This increase in steroid biosynthesis results from selectively increased cholesterol side-chain cleavage (CYP11A), 3β-hydroxysteroid dehydrogenase type II (HSD3B2) and 17α-hydroxylase (CYP17) gene expression. The finding that increased androgen biosynthesis is a stable characteristic of propagated PCOS theca cells maintained in long-term culture has provided new opportunities to examine the molecular and cellular basis for increased ovarian androgen biosynthesis in patients with PCOS. By characterizing the regulatory systems mediating both metabolic and steroidogenic processes in normal and PCOS theca cells, it is anticipated that new information will be obtained that will provide insight into the relations between insulin resistance, hyperinsulinemia, and hyperandrogenemia in patients with PCOS.