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Viuff M, Skakkebæk A, Johannsen EB, Chang S, Pedersen SB, Lauritsen KM, Pedersen MGB, Trolle C, Just J, Gravholt CH. X chromosome dosage and the genetic impact across human tissues. Genome Med 2023; 15:21. [PMID: 36978128 PMCID: PMC10053618 DOI: 10.1186/s13073-023-01169-4] [Citation(s) in RCA: 15] [Impact Index Per Article: 7.5] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/02/2022] [Accepted: 03/06/2023] [Indexed: 03/30/2023] Open
Abstract
BACKGROUND Sex chromosome aneuploidies (SCAs) give rise to a broad range of phenotypic traits and diseases. Previous studies based on peripheral blood samples have suggested the presence of ripple effects, caused by altered X chromosome number, affecting the methylome and transcriptome. Whether these alterations can be connected to disease-specific tissues, and thereby having clinical implication for the phenotype, remains to be elucidated. METHODS We performed a comprehensive analysis of X chromosome number on the transcriptome and methylome in blood, fat, and muscle tissue from individuals with 45,X, 46,XX, 46,XY, and 47,XXY. RESULTS X chromosome number affected the transcriptome and methylome globally across all chromosomes in a tissue-specific manner. Furthermore, 45,X and 47,XXY demonstrated a divergent pattern of gene expression and methylation, with overall gene downregulation and hypomethylation in 45,X and gene upregulation and hypermethylation in 47,XXY. In fat and muscle, a pronounced effect of sex was observed. We identified X chromosomal genes with an expression pattern different from what would be expected based on the number of X and Y chromosomes. Our data also indicate a regulatory function of Y chromosomal genes on X chromosomal genes. Fourteen X chromosomal genes were downregulated in 45,X and upregulated in 47,XXY, respectively, in all three tissues (AKAP17A, CD99, DHRSX, EIF2S3, GTPBP6, JPX, KDM6A, PP2R3B, PUDP, SLC25A6, TSIX, XIST, ZBED1, ZFX). These genes may be central in the epigenetic and genomic regulation of sex chromosome aneuploidies. CONCLUSION We highlight a tissue-specific and complex effect of X chromosome number on the transcriptome and methylome, elucidating both shared and non-shared gene-regulatory mechanism between SCAs.
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Affiliation(s)
- Mette Viuff
- Department of Molecular Medicine, Aarhus University Hospital, Palle-Juul Jensens Boulevard 99, Aarhus N, 8200, Denmark.
- Department of Gynecology and Obstetrics, Aarhus University Hospital, Palle-Juul Jensens Boulevard 99, Aarhus N, 8200, Denmark.
- Department of Clinical Medicine, Aarhus University, Palle-Juul Jensens Boulevard 99, Aarhus N, 8200, Denmark.
| | - Anne Skakkebæk
- Department of Molecular Medicine, Aarhus University Hospital, Palle-Juul Jensens Boulevard 99, Aarhus N, 8200, Denmark.
- Department of Clinical Medicine, Aarhus University, Palle-Juul Jensens Boulevard 99, Aarhus N, 8200, Denmark.
- Department of Clinical Genetics, Aarhus University Hospital, Palle-Juul Jensens Boulevard 99, Aarhus N, 8200, Denmark.
| | - Emma B Johannsen
- Department of Molecular Medicine, Aarhus University Hospital, Palle-Juul Jensens Boulevard 99, Aarhus N, 8200, Denmark
- Department of Clinical Medicine, Aarhus University, Palle-Juul Jensens Boulevard 99, Aarhus N, 8200, Denmark
| | - Simon Chang
- Department of Endocrinology and Internal Medicine and Medical Research Laboratories, Aarhus University Hospital, Palle-Juul Jensens Boulevard 99, Aarhus N, 8200, Denmark
| | - Steen Bønlykke Pedersen
- Department of Endocrinology and Internal Medicine and Medical Research Laboratories, Aarhus University Hospital, Palle-Juul Jensens Boulevard 99, Aarhus N, 8200, Denmark
| | - Katrine Meyer Lauritsen
- Department of Endocrinology and Internal Medicine and Medical Research Laboratories, Aarhus University Hospital, Palle-Juul Jensens Boulevard 99, Aarhus N, 8200, Denmark
| | - Mette Glavind Bülow Pedersen
- Department of Endocrinology and Internal Medicine and Medical Research Laboratories, Aarhus University Hospital, Palle-Juul Jensens Boulevard 99, Aarhus N, 8200, Denmark
| | - Christian Trolle
- Department of Endocrinology and Internal Medicine and Medical Research Laboratories, Aarhus University Hospital, Palle-Juul Jensens Boulevard 99, Aarhus N, 8200, Denmark
| | - Jesper Just
- Department of Molecular Medicine, Aarhus University Hospital, Palle-Juul Jensens Boulevard 99, Aarhus N, 8200, Denmark
- Department of Clinical Medicine, Aarhus University, Palle-Juul Jensens Boulevard 99, Aarhus N, 8200, Denmark
| | - Claus H Gravholt
- Department of Molecular Medicine, Aarhus University Hospital, Palle-Juul Jensens Boulevard 99, Aarhus N, 8200, Denmark
- Department of Clinical Medicine, Aarhus University, Palle-Juul Jensens Boulevard 99, Aarhus N, 8200, Denmark
- Department of Endocrinology and Internal Medicine and Medical Research Laboratories, Aarhus University Hospital, Palle-Juul Jensens Boulevard 99, Aarhus N, 8200, Denmark
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Kumar V, Umapathy G. Development of an enzyme immunoassay to measure urinary and faecal 5α-androst-16-en-3-one in pigs. MethodsX 2023; 10:102178. [PMID: 37122363 PMCID: PMC10133744 DOI: 10.1016/j.mex.2023.102178] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/29/2022] [Accepted: 04/09/2023] [Indexed: 05/02/2023] Open
Abstract
Androstenone, a volatile steroid that possesses pheromonal activity, is responsible for boar taint, sexual interactions, and reproduction in pigs. A wide range of analytical methods has been developed to quantify and detect androstenone in adipose tissue and blood, which are invasive procedures. Therefore, the present study aimed to develop a non-invasive method to detect and quantify the androstenone. We produced group-specific polyclonal androstenone antibody to standardize and validate an enzyme immunoassay to measure faecal and urinary androstenone in Yorkshire boars and sows. Parallelism was performed to determine the immunoreactivity between faecal and urinary immunoreactive androstenone and respective antibody. In boars, urinary and faecal androstenone concentrations were higher on the day of mounting and copulation with sows. In sows, we also measured faecal progesterone metabolites to confirm the oestrus and mating. Faecal androstenone concentrations were peaked on the day of oestrus and mating in sows. Our results suggest that androstenone could be detected and quantified in faecal and urine samples of boars and sows. •Developed an enzyme immunoassay for measuring 5α-androst-16-en-3-one as a marker of boar taint and sex pheromone in urine and faeces of pigs•Detection of 5α-androst-16-en-3-one using a non-invasive method.
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Balhara A, Kumar AR, Unadkat JD. Predicting Human Fetal Drug Exposure Through Maternal-Fetal PBPK Modeling and In Vitro or Ex Vivo Studies. J Clin Pharmacol 2022; 62 Suppl 1:S94-S114. [PMID: 36106781 PMCID: PMC9494623 DOI: 10.1002/jcph.2117] [Citation(s) in RCA: 11] [Impact Index Per Article: 3.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/09/2022] [Accepted: 06/20/2022] [Indexed: 11/06/2022]
Abstract
Medication (drug) use in human pregnancy is prevalent. Determining fetal safety and efficacy of drugs is logistically challenging. However, predicting (not measuring) fetal drug exposure (systemic and tissue) throughout pregnancy is possible through maternal-fetal physiologically based pharmacokinetic (PBPK) modeling and simulation. Such prediction can inform fetal drug safety and efficacy. Fetal drug exposure can be quantified in 2 complementary ways. First, the ratio of the steady-state unbound plasma concentration in the fetal plasma (or area under the plasma concentration-time curve) to the corresponding maternal plasma concentration (ie, Kp,uu ). Second, the maximum unbound peak (Cu,max,ss,f ) and trough (Cu,min,ss,f ) fetal steady-state plasma concentrations. We (and others) have developed a maternal-fetal PBPK model that can successfully predict maternal drug exposure. To predict fetal drug exposure, the model needs to be populated with drug specific parameters, of which transplacental clearances (active and/or passive) and placental/fetal metabolism of the drug are critical. Herein, we describe in vitro studies in cells/tissue fractions or the perfused human placenta that can be used to determine these drug-specific parameters. In addition, we provide examples whereby this approach has successfully predicted systemic fetal exposure to drugs that passively or actively cross the placenta. Apart from maternal-fetal PBPK models, animal studies also have the potential to estimate fetal drug exposure by allometric scaling. Whether such scaling will be successful is yet to be determined. Here, we review the above approaches to predict fetal drug exposure, outline gaps in our knowledge to make such predictions and map out future research directions that could fill these gaps.
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Affiliation(s)
- Ankit Balhara
- Department of Pharmaceutics, University of Washington, Seattle, Washington, USA
| | - Aditya R Kumar
- Department of Pharmaceutics, University of Washington, Seattle, Washington, USA
| | - Jashvant D Unadkat
- Department of Pharmaceutics, University of Washington, Seattle, Washington, USA
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Tiwari S, Rajamanickam G, Unnikrishnan V, Ojaghi M, Kastelic JP, Thundathil JC. Testis-Specific Isoform of Na +-K + ATPase and Regulation of Bull Fertility. Int J Mol Sci 2022; 23:7936. [PMID: 35887284 PMCID: PMC9317330 DOI: 10.3390/ijms23147936] [Citation(s) in RCA: 8] [Impact Index Per Article: 2.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/13/2022] [Revised: 07/08/2022] [Accepted: 07/11/2022] [Indexed: 12/10/2022] Open
Abstract
An advanced understanding of sperm function is relevant for evidence-based male fertility prediction and addressing male infertility. A standard breeding soundness evaluation (BSE) merely identifies gross abnormalities in bulls, whereas selection based on single nucleotide polymorphisms and genomic estimated breeding values overlooks sub-microscopic differences in sperm. Molecular tools are important for validating genomic selection and advancing knowledge on the regulation of male fertility at an interdisciplinary level. Therefore, research in this field is now focused on developing a combination of in vitro sperm function tests and identifying biomarkers such as sperm proteins with critical roles in fertility. The Na+-K+ ATPase is a ubiquitous transmembrane protein and its α4 isoform (ATP1A4) is exclusively expressed in germ cells and sperm. Furthermore, ATP1A4 is essential for male fertility, as it interacts with signaling molecules in both raft and non-raft fractions of the sperm plasma membrane to regulate capacitation-associated signaling, hyperactivation, sperm-oocyte interactions, and activation. Interestingly, ATP1A4 activity and expression increase during capacitation, challenging the widely accepted dogma of sperm translational quiescence. This review discusses the literature on the role of ATP1A4 during capacitation and fertilization events and its prospective use in improving male fertility prediction.
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Affiliation(s)
| | | | | | | | | | - Jacob C. Thundathil
- Department of Production Animal Health, Faculty of Veterinary Medicine, University of Calgary, Calgary, AB T2N 4N1, Canada; (S.T.); (G.R.); (V.U.); (M.O.); (J.P.K.)
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5
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Göschl L, Gmeiner G, Gärtner P, Steinacher M, Forsdahl G. Detection of DHCMT long-term metabolite glucuronides with LC-MSMS as an alternative approach to conventional GC-MSMS analysis. Steroids 2022; 180:108979. [PMID: 35183566 DOI: 10.1016/j.steroids.2022.108979] [Citation(s) in RCA: 4] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 08/17/2021] [Revised: 01/19/2022] [Accepted: 02/07/2022] [Indexed: 10/19/2022]
Abstract
Dehydrochloromethyltestosterone (DHCMT) is one of the most detected illicit used anabolic-androgenic steroids in professional sports. Therefore, a fast and accurate analysis of this substance is of great importance for a constructive fight against doping abuse. The conventional method for the analysis of this drug, GC-MSMS, is very sensitive and selective but also very time- and resource-consuming. With the presented work, a new approach for simple detection with LC-HRMSMS without any sample preparation is introduced. The method is based on the direct analysis of two newly described phase-II metabolites of the DHCMT long-term metabolite 4-chloro-18-nor-17β-hydroxymethyl-17α-methyl-5β-androst-13-en-3α-ol (M3). LC-HRMSMS, GC-MSMS, fractionation and derivatization experiments are combined to identify and characterize for the first time two different glucuronide-acid conjugates of this metabolite in positive human urine samples. In addition, a third glucuronide metabolite was identified, however without isomeric structure determination. The detection of these metabolites is particularly interesting for confirmation analyses, as the method is rapid and requires little sample material.
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Affiliation(s)
- Lorenz Göschl
- Doping Control Laboratory, Seibersdorf Labor GmbH, Seibersdorf, Austria; Department of Pharmacy, University of Tromsø - The Arctic University of Norway, Tromsø, Norway.
| | - Günter Gmeiner
- Doping Control Laboratory, Seibersdorf Labor GmbH, Seibersdorf, Austria
| | - Peter Gärtner
- Institute of Applied Synthetic Chemistry, Technical University of Vienna, Vienna, Austria
| | - Michael Steinacher
- Institute of Applied Synthetic Chemistry, Technical University of Vienna, Vienna, Austria
| | - Guro Forsdahl
- Doping Control Laboratory, Seibersdorf Labor GmbH, Seibersdorf, Austria; Department of Pharmacy, University of Tromsø - The Arctic University of Norway, Tromsø, Norway
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Dembitsky VM. Antitumor and hepatoprotective activity of natural and synthetic neo steroids. Prog Lipid Res 2020; 79:101048. [PMID: 32603672 DOI: 10.1016/j.plipres.2020.101048] [Citation(s) in RCA: 27] [Impact Index Per Article: 5.4] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/23/2020] [Revised: 04/21/2020] [Accepted: 06/12/2020] [Indexed: 02/06/2023]
Abstract
In this review, steroids with a tertiary butyl group, which are usually called neo steroids, are a small group of natural lipids isolated from higher plants, fungi, marine sponges, and yeast. In addition, steroids with a tertiary butyl group have been synthesized in some laboratories in Canada, USA, Europe, and Japan and their biological activity was studied. Some natural neo steroids demonstrate antitumor or hepatoprotective activities. In addition, synthetic neo steroids exhibit anticancer and neuroprotective properties. However, to confirm the above data, both practical and clinical experimental studies are necessary. Nevertheless, the results may be useful for pharmacologists, chemists, biochemists, and the pharmaceutical industry.
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Affiliation(s)
- Valery M Dembitsky
- Centre for Applied Research, Innovation and Entrepreneurship Lethbridge College, 3000 College Drive South, Lethbridge, AB T1K 1L6, Canada.
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Göschl L, Gmeiner G, Enev V, Kratena N, Gärtner P, Forsdahl G. Development and validation of a simple online‐SPE method coupled to high‐resolution mass spectrometry for the analysis of stanozolol‐N‐glucuronides in urine samples. Drug Test Anal 2020; 12:1031-1040. [DOI: 10.1002/dta.2805] [Citation(s) in RCA: 6] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/15/2019] [Revised: 04/01/2020] [Accepted: 04/11/2020] [Indexed: 01/24/2023]
Affiliation(s)
- Lorenz Göschl
- Doping Control Laboratory, Seibersdorf Labor GmbH Seibersdorf Austria
- Department of Pharmacy University of Tromsø – The Arctic University of Norway Tromsø Norway
| | - Günter Gmeiner
- Doping Control Laboratory, Seibersdorf Labor GmbH Seibersdorf Austria
| | - Valentin Enev
- Institute of Applied Synthetic Chemistry Technical University of Vienna Austria
| | - Nicolas Kratena
- Institute of Applied Synthetic Chemistry Technical University of Vienna Austria
| | - Peter Gärtner
- Institute of Applied Synthetic Chemistry Technical University of Vienna Austria
| | - Guro Forsdahl
- Doping Control Laboratory, Seibersdorf Labor GmbH Seibersdorf Austria
- Department of Pharmacy University of Tromsø – The Arctic University of Norway Tromsø Norway
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8
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Nazmeen A, Chen G, Ghosh TK, Maiti S. Breast cancer pathogenesis is linked to the intra-tumoral estrogen sulfotransferase (hSULT1E1) expressions regulated by cellular redox dependent Nrf-2/NF κβ interplay. Cancer Cell Int 2020; 20:70. [PMID: 32158360 PMCID: PMC7057506 DOI: 10.1186/s12935-020-1153-y] [Citation(s) in RCA: 8] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/31/2019] [Accepted: 02/24/2020] [Indexed: 12/20/2022] Open
Abstract
BACKGROUND Estrogen sulfotransferase catalyzes conjugation of sulfuryl-group to estradiol/estrone and regulates E2 availability/activity via estrogen-receptor or non-receptor mediated pathways. Sulfoconjugated estrogen fails to bind estrogen-receptor (ER). High estrogen is a known carcinogen in postmenopausal women. Reports reveal a potential redox-regulation of hSULT1E1/E2-signalling. Further, oxidatively-regulated nuclear-receptor-factor 2 (Nrf2) and NFκβ in relation to hSULT1E1/E2 could be therapeutic-target via cellular redox-modification. METHODS Here, oxidative stress-regulated SULT1E1-expression was analyzed in human breast carcinoma-tissues and in rat xenografted with human breast-tumor. Tumor and its surrounding tissues were obtained from the district-hospital. Intracellular redox-environment of tumors was screened with some in vitro studies. RT-PCR and western blotting was done for SULT1E1 expression. Immunohistochemistry was performed to analyze SULT1E1/Nrf2/NFκβ localization. Tissue-histoarchitecture/DNA-stability (comet assay) studies were done. RESULTS Oxidative-stress induces SULT1E1 via Nrf2/NFκβ cooperatively in tumor-pathogenesis to maintain the required proliferative-state under enriched E2-environment. Higher malondialdehyde/non-protein-soluble-thiol with increased superoxide-dismutase/glutathione-peroxidase/catalase activities was noticed. SULT1E1 expression and E2-level were increased in tumor-tissue compared to their corresponding surrounding-tissues. CONCLUSIONS It may be concluded that tumors maintain a sustainable oxidative-stress through impaired antioxidants as compared to the surrounding. Liver-tissues from xenografted rat manifested similar E2/antioxidant dysregulations favoring pre-tumorogenic environment.
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Affiliation(s)
- Aarifa Nazmeen
- Dept. of Biochemistry, Cell & Molecular Therapeutics Lab, Oriental Institute of Science & Technology, Midnapore, 721101 India
| | - Guangping Chen
- Venture I OSU Laboratory, Oklahoma Technology & Research Park, 1110 S. Innovation Way, Stillwater, OK 74074 USA
| | - Tamal Kanti Ghosh
- Special Secretary, Higher Medical Education, Health and Family Welfare Dept, Govt. of West Bengal, Salt Lake, Calcutta, India
| | - Smarajit Maiti
- Dept. of Biochemistry, Cell & Molecular Therapeutics Lab, Oriental Institute of Science & Technology, Midnapore, 721101 India
- Department of Biochemistry and Biotechnology, Cell & Molecular Therapeutics Lab, OIST, Midnapore, 721102 India
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Surmak AJ, Wong KP, Cole GB, Hirata K, Aabedi AA, Mirfendereski O, Mirfendereski P, Yu AS, Huang SC, Ringman JM, Liebeskind DS, Barrio JR. Probing Estrogen Sulfotransferase-Mediated Inflammation with [11C]-PiB in the Living Human Brain. J Alzheimers Dis 2020; 73:1023-1033. [DOI: 10.3233/jad-190559] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/15/2022]
Affiliation(s)
- Andrew J. Surmak
- University of California, Los Angeles, David Geffen School of Medicine, Department of Molecular and Medical Pharmacology, Los Angeles, CA, USA
| | - Koon-Pong Wong
- University of California, Los Angeles, David Geffen School of Medicine, Department of Molecular and Medical Pharmacology, Los Angeles, CA, USA
| | - Graham B. Cole
- University of California, Los Angeles, David Geffen School of Medicine, Department of Molecular and Medical Pharmacology, Los Angeles, CA, USA
| | - Kenji Hirata
- University of California, Los Angeles, David Geffen School of Medicine, Department of Molecular and Medical Pharmacology, Los Angeles, CA, USA
- Department of Nuclear Medicine, Hokkaido University, Sapporo, Japan
| | - Alexander A. Aabedi
- University of California, Los Angeles, David Geffen School of Medicine, Department of Molecular and Medical Pharmacology, Los Angeles, CA, USA
| | - Omid Mirfendereski
- University of California, Los Angeles, David Geffen School of Medicine, Department of Molecular and Medical Pharmacology, Los Angeles, CA, USA
| | - Payam Mirfendereski
- University of California, Los Angeles, David Geffen School of Medicine, Department of Molecular and Medical Pharmacology, Los Angeles, CA, USA
| | - Amy S. Yu
- University of California, Los Angeles, David Geffen School of Medicine, Department of Molecular and Medical Pharmacology, Los Angeles, CA, USA
- Department of Radiation Oncology, Stanford University, Stanford, CA, USA
| | - Sung-Cheng Huang
- University of California, Los Angeles, David Geffen School of Medicine, Department of Molecular and Medical Pharmacology, Los Angeles, CA, USA
| | - John M. Ringman
- University of California, Los Angeles, David Geffen School of Medicine, Department of Molecular and Medical Pharmacology, Los Angeles, CA, USA
- University of Southern California, Department of Neurology, Los Angeles, CA, USA
| | - David S. Liebeskind
- Department of Neurology, David Geffen School of Medicine at UCLA, Los Angeles, CA, USA
| | - Jorge R. Barrio
- University of California, Los Angeles, David Geffen School of Medicine, Department of Molecular and Medical Pharmacology, Los Angeles, CA, USA
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Li T, Zhang W, Lin SX. Steroid enzyme and receptor expression and regulations in breast tumor samples - A statistical evaluation of public data. J Steroid Biochem Mol Biol 2020; 196:105494. [PMID: 31610224 DOI: 10.1016/j.jsbmb.2019.105494] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 05/10/2019] [Revised: 09/20/2019] [Accepted: 10/07/2019] [Indexed: 12/16/2022]
Abstract
In spite of the significant progress of estrogen-dependent breast cancer (BC) treatment, aromatase inhibitor resistance is a major problem limiting the clinical benefit of this frontier endocrine-therapy. The aim of this study was to determine the differential expression of steroid-converting enzymes between tumor and adjacent normal tissues, as well as their correlation in modulating intratumoral steroid-hormone levels in post-menopausal estrogen-dependent BC. RNA sequencing dataset (n = 1097) of The-Cancer-Genome-Atlas (Breast Invasive Carcinoma) retrieved through the data portal of Genomic Data Commons was used for differential expressions and expression correlation analyses by Mann-Whitney U and Spearman's rank test, respectively. The results showed significant up-regulation of 17β-HSD7 (2.50-fold, p < 0.0001) in BC, supporting its effect in sex-hormone control. Besides, suppression of 11β-HSD1 expression (-8.29-fold, p < 0.0001) and elevation of 11β-HSD2 expression (2.04-fold, p < 0.0001) provide a low glucocorticoid environment diminishing BC anti-proliferation. Furthermore, 3α-HSDs were down-regulated (-1.59-fold, p < 0.01; -8.18-fold, p < 0.0001; -33.96-fold, p < 0.0001; -31.85-fold, p < 0.0001 for type 1-4, respectively), while 5α-reductases were up-regulated (1.41-fold, p < 0.0001; 2.85-fold, p < 0.0001; 1.70-fold, p < 0.0001 for type 1-3, respectively) in BC, reducing cell proliferation suppressers 4-pregnenes, increasing cell proliferation stimulators 5α-pregnanes. Expression analysis indicates significant correlations between 11β-HSD1 with 3α-HSD4 (r = 0.605, p < 0.0001) and 3α-HSD3 (r = 0.537, p < 0.0001). Significant expression correlations between 3α-HSDs were also observed. Our results systematically present the regulation of steroid-converting enzymes and their roles in modulating the intratumoral steroid-hormone levels in BC with a vivid 3D-schema, supporting novel therapy targeting the reductive 17β-HSD7 and proposing a new combined therapy targeting 11β-HSD2 and 17β-HSD7.
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MESH Headings
- 17-Hydroxysteroid Dehydrogenases/genetics
- 17-Hydroxysteroid Dehydrogenases/metabolism
- Breast Neoplasms/epidemiology
- Breast Neoplasms/genetics
- Breast Neoplasms/metabolism
- Carcinoma, Ductal, Breast/epidemiology
- Carcinoma, Ductal, Breast/genetics
- Carcinoma, Ductal, Breast/metabolism
- Cohort Studies
- Cytochrome P-450 Enzyme System/genetics
- Cytochrome P-450 Enzyme System/metabolism
- Databases, Factual/statistics & numerical data
- Estradiol/pharmacology
- Female
- Gene Expression Regulation, Enzymologic/drug effects
- Gene Expression Regulation, Neoplastic/drug effects
- Gonadal Steroid Hormones/genetics
- Gonadal Steroid Hormones/metabolism
- Humans
- Public Sector/statistics & numerical data
- Receptors, Cytoplasmic and Nuclear/genetics
- Receptors, Cytoplasmic and Nuclear/metabolism
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Affiliation(s)
- Tang Li
- Axe Molecular Endocrinology and Nephrology, CHU Research Center and Department of Molecular Medicine, Laval University, 2705 Boulevard Laurier, Québec City, Québec G1V 4G2, Canada
| | - Wenfa Zhang
- Axe Molecular Endocrinology and Nephrology, CHU Research Center and Department of Molecular Medicine, Laval University, 2705 Boulevard Laurier, Québec City, Québec G1V 4G2, Canada
| | - Sheng-Xiang Lin
- Axe Molecular Endocrinology and Nephrology, CHU Research Center and Department of Molecular Medicine, Laval University, 2705 Boulevard Laurier, Québec City, Québec G1V 4G2, Canada.
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Afzal S, Ramzan K, Ullah S, Wakil SM, Jamal A, Basit S, Waqar AB. A novel nonsense mutation in the STS gene in a Pakistani family with X-linked recessive ichthyosis: including a very rare case of two homozygous female patients. BMC MEDICAL GENETICS 2020; 21:20. [PMID: 32005174 PMCID: PMC6995215 DOI: 10.1186/s12881-020-0964-y] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 09/04/2019] [Accepted: 01/24/2020] [Indexed: 01/29/2023]
Abstract
Background X-linked ichthyosis (XLI; OMIM# 308100) is a recessive keratinization disorder characterized by the presence of dark brown, polygonal, adherent scales on different parts of the body surface. It almost exclusively affects males and the estimated prevalence ranges from 1:2000–6000 in males worldwide. Extracutaneous manifestations are frequent including corneal opacities, cryptorchidism, neuropsychiatric symptoms or others. Up to 90% of XLI cases are caused by recurrent hemizygous microdeletion encompassing entire STS gene on chromosome Xp22.3, while only a minority of patients shows partial deletions or loss of function point mutations in STS. Larger deletions also involving contiguous genes are identified in syndromic patients. Methods Here, we report clinical and genetic findings of a large Pakistani family having 16 affected individuals including 2 females with XLI. Molecular karyotyping and direct DNA sequencing of coding region of the STS gene was performed. Results The clinical manifestations in affected individuals involved generalized dryness and scaling of the skin with polygonal, dark scales of the skin on scalp, trunk, limbs, and neck while sparing face, palms and soles. There were no associated extra-cutaneous features such as short stature, cryptorchidism, photophobia, corneal opacities, male baldness, and behavioral, cognitive, or neurological phenotypes including intellectual disability, autism or attention deficit hyperactivity disorder. Molecular karyotyping was normal and no copy number variation was found. Sanger sequencing identified a novel hemizygous nonsense mutation (c.287G > A; p.W96*), in exon 4 of STS gene in all affected male individuals. In addition, two XLI affected females in the family were found to be homozygous for the identified variant. Conclusions This study is useful for understanding the genetic basis of XLI in the patients studied, for extending the known mutational spectrum of STS, diagnosis of female carriers and for further application of mutation screening in the genetic counseling of this family.
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Affiliation(s)
- Sibtain Afzal
- Faculty of Allied and Health Sciences, Imperial College of Business Studies, Lahore, Pakistan
| | - Khushnooda Ramzan
- Department of Genetics, King Faisal Specialist Hospital and Research Centre, PO Box 3354, Riyadh, 11211, Saudi Arabia
| | - Sajjad Ullah
- Faculty of Allied and Health Sciences, Imperial College of Business Studies, Lahore, Pakistan
| | - Salma M Wakil
- Department of Genetics, King Faisal Specialist Hospital and Research Centre, PO Box 3354, Riyadh, 11211, Saudi Arabia
| | - Arshad Jamal
- Faculty of Allied and Health Sciences, Imperial College of Business Studies, Lahore, Pakistan
| | - Sulman Basit
- Center for Genetics and Inherited Diseases, Taibah University, Madinah Al-Munawarah, Medina, Saudi Arabia
| | - Ahmed Bilal Waqar
- Faculty of Allied and Health Sciences, Imperial College of Business Studies, Lahore, Pakistan.
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12
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Giudice A, Barbieri A, Bimonte S, Cascella M, Cuomo A, Crispo A, D'Arena G, Galdiero M, Della Pepa ME, Botti G, Caraglia M, Capunzo M, Arra C, Montella M. Dissecting the prevention of estrogen-dependent breast carcinogenesis through Nrf2-dependent and independent mechanisms. Onco Targets Ther 2019; 12:4937-4953. [PMID: 31388303 PMCID: PMC6607693 DOI: 10.2147/ott.s183192] [Citation(s) in RCA: 11] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/08/2018] [Accepted: 12/14/2018] [Indexed: 12/19/2022] Open
Abstract
Breast cancer is the most common malignancy among women worldwide. Various studies indicate that prolonged exposure to elevated levels of estrogens is associated with development of breast cancer. Both estrogen receptor-dependent and independent mechanisms can contribute to the carcinogenic effects of estrogens. Among them, the oxidative metabolism of estrogens plays a key role in the initiation of estradiol-induced breast cancer by generation of reactive estrogen quinones as well as the associated formation of oxygen free radicals. These genotoxic metabolites can react with DNA to form unstable DNA adducts which generate mutations leading to the initiation of breast cancer. A variety of endogenous and exogenous factors can alter estrogen homeostasis and generate genotoxic metabolites. The use of specific phytochemicals and dietary supplements can inhibit the risk of breast cancer not only by the modulation of several estrogen-activating enzymes (CYP19, CYP1B1) but also through the induction of various cytoprotective enzymes (eg, SOD3, NQO1, glutathione S-transferases, OGG-1, catechol-O-methyltransferases, CYP1B1A, etc.) that reestablish the homeostatic balance of estrogen metabolism via nuclear factor erythroid 2-related factor 2 (Nrf2)-dependent and independent mechanisms.
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Affiliation(s)
- Aldo Giudice
- Epidemiology Unit, Istituto Nazionale Tumori “Fondazione G. Pascale”, IRCCS, Naples, Italy
| | - Antonio Barbieri
- S.S.D Sperimentazione Animale, Istituto Nazionale Tumori “Fondazione G. Pascale”, IRCCS, Naples, Italy
| | - Sabrina Bimonte
- Division of Anesthesia and Pain Medicine, Istituto Nazionale Tumori “Fondazione G. Pascale”, IRCCS, Naples, Italy
| | - Marco Cascella
- Division of Anesthesia and Pain Medicine, Istituto Nazionale Tumori “Fondazione G. Pascale”, IRCCS, Naples, Italy
| | - Arturo Cuomo
- Division of Anesthesia and Pain Medicine, Istituto Nazionale Tumori “Fondazione G. Pascale”, IRCCS, Naples, Italy
| | - Anna Crispo
- Epidemiology Unit, Istituto Nazionale Tumori “Fondazione G. Pascale”, IRCCS, Naples, Italy
| | - Giovanni D'Arena
- Hematology and Stem Cell Transplantation Unit, IRCCS Centro di Riferimento Oncologico della Basilicata, Rionero in Vulture, Italy
| | - Massimiliano Galdiero
- Department of Experimental Medicine, Università della Campania “Luigi Vanvitelli”, 80134Naples, Italy
| | - Maria Elena Della Pepa
- Department of Experimental Medicine, Università della Campania “Luigi Vanvitelli”, 80134Naples, Italy
| | - Gerardo Botti
- Scientific Direction, Istituto Nazionale Tumori-IRCCS “Fondazione G. Pascale”, Naples, Italy
| | - Michele Caraglia
- Department of Biochemistry, Biophysics and General Pathology, University of Campania “Luigi Vanvitelli”, 80138Naples, Italy
| | - Mario Capunzo
- Department of Medicine, Surgery and Dentistry “Scuola Medica Salernitana”, University of Salerno, Baronissi, 84081Salerno, Italy
| | - Claudio Arra
- S.S.D Sperimentazione Animale, Istituto Nazionale Tumori “Fondazione G. Pascale”, IRCCS, Naples, Italy
| | - Maurizio Montella
- Epidemiology Unit, Istituto Nazionale Tumori “Fondazione G. Pascale”, IRCCS, Naples, Italy
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13
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Laderoute H, Bone C, Brewer D, Squires EJ. The synthesis of 16-androstene sulfoconjugates from primary porcine Leydig cell culture. Steroids 2019; 146:14-20. [PMID: 30904503 DOI: 10.1016/j.steroids.2019.03.007] [Citation(s) in RCA: 8] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 01/09/2019] [Revised: 02/19/2019] [Accepted: 03/14/2019] [Indexed: 11/22/2022]
Abstract
Increased public interest in the welfare of pigs reared for pork production has led to an enhanced effort in finding alternatives to castration for controlling the unpleasant odour and flavour from heated pork products known as boar taint. The purpose of this study was to investigate the testicular metabolism of androstenone, one of the major components of boar taint. Leydig cells were isolated from mature boars and incubated with radiolabeled androstenone for 10 min, 1 h, 4 h, 8 h, and 12 h. Steroid profiles were analyzed by high-performance liquid chromatography (HPLC) and liquid chromatography-mass spectrometry (LC-MS/MS). Sulfoconjugated, but not glucuronidated steroids were produced by Leydig cells. Approximately 85% of androstenone was converted into sulfoconjugated metabolites in Leydig cell incubations after 8 h. This sulfoconjugate fraction included androstenol-3-sulfate and two major sulfated forms of androstenone. Following removal of the sulfate group, these two sulfated forms of androstenone returned the parent compound androstenone, and not a hydroxylated metabolite. These findings provided direct evidence for the testicular production of sulfoconjugated forms of androstenone and androstenol in the boar. The high proportion of sulfoconjugates produced by the Leydig cells emphasizes the importance of steroid conjugation, which serves to regulate the amount of unconjugated steroid hormones available for accumulation in adipose tissue.
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Affiliation(s)
- Heidi Laderoute
- Department of Animal Biosciences, University of Guelph, Guelph, Ontario N1G2W1, Canada
| | - Christine Bone
- Department of Animal Biosciences, University of Guelph, Guelph, Ontario N1G2W1, Canada
| | - Dyanne Brewer
- Advanced Analysis Centre, University of Guelph, Guelph, Ontario N1G2W1, Canada
| | - E James Squires
- Department of Animal Biosciences, University of Guelph, Guelph, Ontario N1G2W1, Canada.
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14
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Tissue steroid levels in response to reduced testicular estrogen synthesis in the male pig, Sus scrofa. PLoS One 2019; 14:e0215390. [PMID: 30986232 PMCID: PMC6464225 DOI: 10.1371/journal.pone.0215390] [Citation(s) in RCA: 9] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/02/2018] [Accepted: 04/01/2019] [Indexed: 12/17/2022] Open
Abstract
Production of steroid hormones is complex and dependent upon steroidogenic enzymes, cofactors, receptors, and transporters expressed within a tissue. Collectively, these factors create an environment for tissue-specific steroid hormone profiles and potentially tissue-specific responses to drug administration. Our objective was to assess steroid production, including sulfated steroid metabolites in the boar testis, prostate, and liver following inhibition of aromatase, the enzyme that converts androgen precursors to estrogens. Boars were treated with the aromatase inhibitor, letrozole from 11 to 16 weeks of age and littermate boars received the canola oil vehicle. Steroid profiles were evaluated in testes, prostate, and livers of 16, 20, and 40 week old boars using liquid chromatography/mass spectrometry. Testis, prostate, and liver had unique steroid profiles in vehicle-treated animals. Only C18 steroid hormones were altered by treatment with the aromatase inhibitor, letrozole; no significant differences were detected in any of the C19 or C21 steroids evaluated. Testis was the only tissue with significantly decreased free estrogens following treatment with the aromatase inhibitor; estrone and estradiol concentrations were lower (p < 0.05) in testes from 16, 20, and 40 week letrozole-treated boars. However, concentrations of the sulfated conjugates, estrone-sulfate and estradiol-sulfate, were significantly decreased (p<0.05) in 16 and 20 week boar testes, prostates, and livers from letrozole-treated boars. Hence, the distribution of estrogens between the free and conjugated forms was altered in a tissue-specific manner following inhibition of aromatase. The results suggest sulfated testicular estrogens are important estrogen precursors for the prostate, potentially enabling peripheral target tissues to synthesize free estrogens in the male pig.
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15
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Pranata A, Fitzgerald CC, Khymenets O, Westley E, Anderson NJ, Ma P, Pozo OJ, McLeod MD. Synthesis of steroid bisglucuronide and sulfate glucuronide reference materials: Unearthing neglected treasures of steroid metabolism. Steroids 2019; 143:25-40. [PMID: 30513322 DOI: 10.1016/j.steroids.2018.11.017] [Citation(s) in RCA: 10] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 08/13/2018] [Revised: 10/24/2018] [Accepted: 11/26/2018] [Indexed: 02/06/2023]
Abstract
Doubly or bisconjugated steroid metabolites have long been known as minor components of the steroid profile that have traditionally been studied by laborious and indirect fractionation, hydrolysis and gas chromatography-mass spectrometry (GC-MS) analysis. Recently, the synthesis and characterisation of steroid bis(sulfate) (aka disulfate or bis-sulfate) reference materials enabled the liquid chromatography-tandem mass spectrometry (LC-MS/MS) study of this metabolite class and the development of a constant ion loss (CIL) scan method for the direct and untargeted detection of steroid bis(sulfate) metabolites. Methods for the direct LC-MS/MS detection of other bisconjugated steroids, such as steroid bisglucuronide and mixed steroid sulfate glucuronide metabolites, have great potential to reveal a more complete picture of the steroid profile. However, access to steroid bisglucuronide or sulfate glucuronide reference materials necessary for LC-MS/MS method development, metabolite identification or quantification is severely limited. In this work, ten steroid bisglucuronide and ten steroid sulfate glucuronide reference materials were synthesised through an ordered combination of chemical sulfation and/or enzymatic glucuronylation reactions. All compounds were purified and characterised using NMR and MS methods. Chemistry for the preparation of stable isotope labelled steroid {13C6}-glucuronide internal standards has also been developed and applied to the preparation of two selectively mono-labelled steroid bisglucuronide reference materials used to characterise more completely MS fragmentation pathways. The electrospray ionisation and fragmentation of the bisconjugated steroid reference materials has been studied. Preliminary targeted ultra-high-performance liquid chromatography-tandem mass spectrometry (UHPLC-MS/MS) analysis of the reference materials prepared revealed the presence of three steroid sulfate glucuronides as endogenous human urinary metabolites.
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Affiliation(s)
- Andy Pranata
- Research School of Chemistry, Australian National University, Canberra, ACT 2601, Australia
| | | | - Olha Khymenets
- Integrative Pharmacology and Systems Neuroscience Group, IMIM, Hospital del Mar, Doctor Aiguader 88, Barcelona, Spain
| | - Erin Westley
- Research School of Chemistry, Australian National University, Canberra, ACT 2601, Australia
| | - Natasha J Anderson
- Research School of Chemistry, Australian National University, Canberra, ACT 2601, Australia
| | - Paul Ma
- Research School of Chemistry, Australian National University, Canberra, ACT 2601, Australia
| | - Oscar J Pozo
- Integrative Pharmacology and Systems Neuroscience Group, IMIM, Hospital del Mar, Doctor Aiguader 88, Barcelona, Spain
| | - Malcolm D McLeod
- Research School of Chemistry, Australian National University, Canberra, ACT 2601, Australia.
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16
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Atwood CS, Ekstein SF. Human versus non-human sex steroid use in hormone replacement therapies part 1: Preclinical data. Mol Cell Endocrinol 2019; 480:12-35. [PMID: 30308266 DOI: 10.1016/j.mce.2018.10.003] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 06/29/2016] [Revised: 03/12/2018] [Accepted: 10/04/2018] [Indexed: 11/24/2022]
Abstract
Prior to 2002, hormone replacement therapy (HRT) was considered to be an important component of postmenopausal healthcare. This was based on a plethora of basic, epidemiological and clinical studies demonstrating the health benefits of supplementation with human sex steroids. However, adverse findings from the Women's Health Initiative (WHI) studies that examined the 2 major forms of HRT in use in the US at that time - Premarin (conjugated equine estrogens; CEE) and Prempro (CEE + medroxyprogesterone acetate; MPA), cast a shadow over the use of any form of HRT. Here we review the biochemical and physiological differences between the non-human WHI study hormones - CEE and MPA, and their respective human counterparts 17β-estradiol (E2) and progesterone (P4). Preclinical data from the last 30 years demonstrate clear differences between human and non-human sex steroids on numerous molecular, physiological and functional parameters in brain, heart and reproductive tissue. In contrast to CEE supplementation, which is not always detrimental although certainly not as optimal as E2 supplementation, MPA is clearly not equivalent to P4, having detrimental effects on cognitive, cardiac and reproductive function. Moreover, unlike P4, MPA is clearly antagonistic of the positive effects of E2 and CEE on tissue function. These data indicate that minor chemical changes to human sex steroids result in physiologically distinct actions that are not optimal for tissue health and functioning.
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Affiliation(s)
- Craig S Atwood
- Division of Geriatrics and Gerontology, Department of Medicine, University of Wisconsin-Madison School of Medicine and Public Health, USA; Geriatric Research, Education and Clinical Center, Veterans Administration Hospital, Madison, WI, 53705, USA; School of Exercise, Biomedical and Health Sciences, Edith Cowan University, Joondalup, 6027, WA, Australia.
| | - Samuel F Ekstein
- Division of Geriatrics and Gerontology, Department of Medicine, University of Wisconsin-Madison School of Medicine and Public Health, USA
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17
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Jackson EN, Rowland-Faux L, James MO, Wood CE. Administration of low dose triclosan to pregnant ewes results in placental uptake and reduced estradiol sulfotransferase activity in fetal liver and placenta. Toxicol Lett 2018; 294:116-121. [PMID: 29772265 DOI: 10.1016/j.toxlet.2018.05.014] [Citation(s) in RCA: 11] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/13/2017] [Revised: 04/26/2018] [Accepted: 05/11/2018] [Indexed: 01/04/2023]
Abstract
Sulfonation is a major pathway of estrogen biotransformation with a role in regulating estrogen homeostasis in humans and sheep. Previous in vitro studies found that triclosan is an especially potent competitive inhibitor of ovine placental estrogen sulfotransferase, with Kic of <0.1 nM. As the placenta is the main organ responsible for estrogen synthesis in pregnancy in both women and sheep, and the liver is another site of estrogen biotransformation, this study examined the effects of triclosan exposure of pregnant ewes on placental and hepatic sulfotransferase activity. Triclosan, 0.1 mg/kg/day, or saline vehicle was administered to late gestation fetal sheep for two days either by direct infusion into the fetal circulation or infusion into the maternal blood. On the third day, fetal liver and placenta were harvested and analyzed for triclosan and for cytosolic estradiol sulfotransferase activity. Placenta contained higher concentrations of triclosan than liver in each individual sheep in both treatment groups. There was a negative correlation between triclosan tissue concentration (pmol/g tissue) and cytosolic sulfotransferase activity (pmol/min/mg protein) towards estradiol. These findings demonstrated that in the sheep exposed to very low concentrations of triclosan, this substance is taken up into placenta and reduces estrogen sulfonation.
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Affiliation(s)
- Erin N Jackson
- Department of Medicinal Chemistry, University of Florida, Gainesville, FL, 32610, USA
| | - Laura Rowland-Faux
- Department of Medicinal Chemistry, University of Florida, Gainesville, FL, 32610, USA
| | - Margaret O James
- Department of Medicinal Chemistry, University of Florida, Gainesville, FL, 32610, USA.
| | - Charles E Wood
- Department of Physiology and Functional Genomics, University of Florida, Gainesville, FL, 32610, USA
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18
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Influence of SULT1A1 genetic variation on age at menopause, estrogen levels, and response to hormone therapy in recently postmenopausal white women. Menopause 2018; 23:863-9. [PMID: 27300114 PMCID: PMC4961269 DOI: 10.1097/gme.0000000000000648] [Citation(s) in RCA: 22] [Impact Index Per Article: 3.1] [Reference Citation Analysis] [Abstract] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/25/2022]
Abstract
Objective: Onset and symptoms of menopause, and response to hormone therapy (HT) show large interindividual variability. SULT1A1 encodes for a highly expressed enzyme that metabolizes estrogens. We evaluated the relationship between genetic variation in SULT1A1, menopause age, symptoms, and response to HT. Methods: Women enrolled in the Kronos Early Estrogen Prevention Study at Mayo Clinic were randomized to 48 months of treatment with oral conjugated equine estrogen (n = 34), transdermal 17β-estradiol (E2) (n = 33), or placebo (n = 35). Linear regression models and ANOVA were used to test for association of SULT1A1 copy number, rs3760091, rs750155, and rs9282861 (SULT1A1∗2), with age at menopause and symptoms, levels of estrogens (estrone [E1], estrone sulfate [E1S], E2, and estradiol sulfate [E2S]), before and after HT. Results: SULT1A1 gene copy number affected the minor allele frequency for each single nucleotide polymorphisms tested. Before administration of exogenous hormones, increasing number of G alleles at rs9282861 was associated with earlier age at menopause (P = 0.014), lower frequency of night sweats (P = 0.009), and less severe insomnia (P = 0.046). After 48 months of treatment, SULT1A1 genotype was not associated with the presence of menopausal symptoms. In women randomized to oral conjugated equine estrogen, increasing number of the A allele at rs750155 was associated with lower E1S and E2S (P = 0.004 and 0.017), whereas increasing number of the C allele at rs3760091 was associated with lower E2S/E2 (P = 0.044). Conclusions: Interindividual variability in onset of menopause and symptoms before initiation of HT is explained in part by genetic variation in SULT1A1 and may represent a step toward individualizing HT treatment decisions.
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19
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Garbacz WG, Jiang M, Xie W. Sex-Dependent Role of Estrogen Sulfotransferase and Steroid Sulfatase in Metabolic Homeostasis. ADVANCES IN EXPERIMENTAL MEDICINE AND BIOLOGY 2017; 1043:455-469. [PMID: 29224107 DOI: 10.1007/978-3-319-70178-3_21] [Citation(s) in RCA: 19] [Impact Index Per Article: 2.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 12/13/2022]
Abstract
Sulfonation and desulfation are two opposing processes that represent an important layer of regulation of estrogenic activity via ligand supplies. Enzymatic activities of families of enzymes, known as sulfotransferases and sulfatases, lead to structural and functional changes of the steroids, thyroids, xenobiotics, and neurotransmitters. Estrogen sulfotransferase (EST) and steroid sulfatase (STS) represent negative and positive regulation of the estrogen activity, respectively. This is because EST-mediated sulfation deactivates estrogens, whereas STS-mediated desulfation converts the inactive estrogen sulfates to active estrogens. In addition to the known functions of estrogens, EST and STS in reproductive processes, regulation of estrogens and other signal molecules especially at the local tissue levels has gained increased attention in the context of metabolic disease in recent years. EST expression is detectable in the subcutaneous adipose tissue in both obese women and men, and the expression of EST is markedly induced in the livers of rodent models of obesity and type 2 diabetes. STS was found to be upregulated in patients with chronic inflammatory liver diseases. Interestingly, the tissue distribution and the transcriptional regulation of EST and STS exhibit obvious sex and species specificity. EST ablation produces completely opposite metabolic phenotype in female and male obese mice. Adipogenesis is also differentially regulated by EST in murine and human adipocytes. This chapter focuses on the recent progress in our understanding of the expression and regulation EST and STS in the context of metabolic homeostasis.
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Affiliation(s)
- Wojciech G Garbacz
- Center for Pharmacogenetics and Department of Pharmaceutical Sciences, University of Pittsburgh, Pittsburgh, PA, USA
| | - Mengxi Jiang
- Center for Pharmacogenetics and Department of Pharmaceutical Sciences, University of Pittsburgh, Pittsburgh, PA, USA
| | - Wen Xie
- Center for Pharmacogenetics and Department of Pharmaceutical Sciences, University of Pittsburgh, Pittsburgh, PA, USA. .,Department of Pharmacology and Chemical Biology, University of Pittsburgh, Pittsburgh, PA, USA.
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20
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Sánchez-Guijo A, Neunzig J, Gerber A, Oji V, Hartmann MF, Schuppe HC, Traupe H, Bernhardt R, Wudy SA. Role of steroid sulfatase in steroid homeostasis and characterization of the sulfated steroid pathway: Evidence from steroid sulfatase deficiency. Mol Cell Endocrinol 2016; 437:142-153. [PMID: 27531568 DOI: 10.1016/j.mce.2016.08.019] [Citation(s) in RCA: 35] [Impact Index Per Article: 3.9] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 04/08/2016] [Revised: 08/09/2016] [Accepted: 08/11/2016] [Indexed: 11/23/2022]
Abstract
The impact of steroid sulfatase (STS) activity in the circulating levels of both sulfated and unconjugated steroids is only partially known. In addition, the sulfated steroid pathway, a parallel pathway to the one for unconjugated steroids, which uses the same enzymes, has never been characterized in detail before. Patients with steroid sulfatase deficiency (STSD) are unable to enzymatically convert sulfated steroids into their unconjugated forms, and are a good model to elucidate how STS affects steroid biosynthesis and to study the metabolism of sulfated steroids. We quantified unconjugated and sulfated steroids in STSD serum, and compared these results with data obtained from serum of healthy controls. Most sulfated steroids were increased in STSD. However, androstenediol-3-sulfate and epiandrosterone sulfate showed similar levels in both groups, and the concentrations of androsterone sulfate were notably lower. Hydroxylated forms of DHEAS and of pregnenolone sulfate were found to be increased in STSD, suggesting a mechanism to improve the excretion of sulfated steroids. STSD testosterone concentrations were normal, but cholesterol and DHEA were significantly decreased. Additionally, serum bile acids were three-fold higher in STSD. Correlations between concentrations of steroids in each group indicate that 17α-hydroxy-pregnenolone-3-sulfate in men is mainly biosynthesized from the precursor pregnenolone sulfate and androstenediol-3-sulfate from DHEAS. These findings confirm the coexistence of two steroidogenic pathways: one for unconjugated steroids and another one for sulfated steroids. Each pathway is responsible for the synthesis of specific steroids. The equal levels of testosterone, and the reduced level of unconjugated precursors in STSD, support that testosterone is primarily synthesized from sulfated steroids. In consequence, testosterone synthesis in STSD relies on an enzyme with sulfatase activity other than STS. This study reveals that STS is a key player of steroid biosynthesis regulating the availability of circulating cholesterol.
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Affiliation(s)
- Alberto Sánchez-Guijo
- Steroid Research & Mass Spectrometry Unit, Division of Pediatric Endocrinology & Diabetology, Center of Child and Adolescent Medicine, Justus Liebig University, Feulgenstrasse 12, 35392, Giessen, Germany.
| | - Jens Neunzig
- Department of Biochemistry, Faculty of Technical and Natural Sciences III, Saarland University, 66123, Saarbrücken, Germany
| | - Adrian Gerber
- Department of Biochemistry, Faculty of Technical and Natural Sciences III, Saarland University, 66123, Saarbrücken, Germany
| | - Vinzenz Oji
- Department of Dermatology, University of Münster, 48149, Münster, Germany
| | - Michaela F Hartmann
- Steroid Research & Mass Spectrometry Unit, Division of Pediatric Endocrinology & Diabetology, Center of Child and Adolescent Medicine, Justus Liebig University, Feulgenstrasse 12, 35392, Giessen, Germany
| | - Hans-Christian Schuppe
- Clinic of Urology, Pediatric Urology and Andrology, Justus-Liebig-University, 35385, Giessen, Germany
| | - Heiko Traupe
- Department of Dermatology, University of Münster, 48149, Münster, Germany
| | - Rita Bernhardt
- Department of Biochemistry, Faculty of Technical and Natural Sciences III, Saarland University, 66123, Saarbrücken, Germany
| | - Stefan A Wudy
- Steroid Research & Mass Spectrometry Unit, Division of Pediatric Endocrinology & Diabetology, Center of Child and Adolescent Medicine, Justus Liebig University, Feulgenstrasse 12, 35392, Giessen, Germany
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21
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Franchi A, Cubilla M, Guidobaldi HA, Bravo AA, Giojalas LC. Uterosome-like vesicles prompt human sperm fertilizing capability. Mol Hum Reprod 2016; 22:833-841. [PMID: 27678485 DOI: 10.1093/molehr/gaw066] [Citation(s) in RCA: 8] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/16/2015] [Revised: 09/09/2016] [Accepted: 09/22/2016] [Indexed: 12/20/2022] Open
Abstract
STUDY QUESTION Does the rapid transit through the uterine environment modulate the sperm physiological state? SUMMARY ANSWER The uterosome-like vesicles (ULVs) secreted by endometrial epithelial cells (EECs) in vitro are able to fuse with human spermatozoa, prompting their fertilizing capacity. WHAT IS KNOWN ALREADY Early studies suggest that sperm capacitation begins in the uterus and ends in the oviduct, and that a synergistic effect of both female organs may accelerate this process. Although it has been reported that co-incubation of human spermatozoa with endometrial cell-conditioned medium (CM) stimulates sperm capacitation, the mechanism mediating this communication is unknown. STUDY DESIGN, SIZE, DURATION Human ULVs secreted by EECs were characterized and their effect on human sperm physiology was analysed. Spermatozoa were incubated with EEC-derived CM or ULV, after which sperm capacitation was evaluated at different time points. In addition, the interaction of spermatozoa with ULV was analysed. PARTICIPANTS/MATERIALS, SETTING, METHODS ULVs were isolated by ultracentrifugation and identified using electron microscopy and Western blotting to assess the presence of specific protein markers. Following seminal plasma removal, human spermatozoa were incubated CM or ULV, after which sperm capacitation was evaluated as the ability of the sperm to undergo the induced acrosome reaction and the level of protein tyrosine phosphorylation (PY) determined by Western blot and immunocytochemistry. The interaction of spermatozoa with labelled ULV was analysed by fluorescence microscopy. In all cases, at least three biological replicates from different sperm donors were performed for each set of experiments. Significant differences between mean values were determined by one-way ANOVA followed by Tukey's post hoc test. Differences between treatments were considered statistically significant at P ≤ 0.05. MAIN RESULTS AND THE ROLE OF CHANCE The level of capacitated spermatozoa and those recruited by chemotaxis increased 3- to 4-fold when spermatozoa were incubated in the presence of CM for 4 h. Even a 15 min incubation of spermatozoa with CM was also enough to increase the level of capacitated cells 3- to 4-fold (P < 0.05). Furthermore, a short co-incubation of spermatozoa with ULV stimulates sperm capacitation, as determined by the increase in the level of induced acrosome reaction and the induction of PY. In addition, after the co-incubation of spermatozoa with fluorescent labelled ULV, the sperm cells acquired the fluorescent staining which indicates that ULV might be transferred to the sperm surface by a fusion mechanism. LIMITATIONS, REASONS FOR CAUTION This is an in vitro study performed with human biological material, spermatozoa and endometrial derived cells; the latter being a cell line originally isolated from a uterine adenocarcinoma. WIDER IMPLICATIONS OF THE FINDINGS The capability of spermatozoa to briefly interact with ULVs supports the hypothesis that any step of sperm transport may have physiological consequences, despite the interaction lasting for only a limited period of time. This way of communication of spermatozoa with cell products of uterine origin opens new frontiers of investigation (e.g. the signalling molecules involved), shedding light on the sperm processes that prepare the male gamete for fertilization, which might have implications for human infertility treatment. LARGE SCALE DATA N/A. STUDY FUNDING AND COMPETING INTERESTS The project was financially supported by SECyT-UNC. The authors declare no conflict of interest.
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Affiliation(s)
- A Franchi
- Centro de Biología Celular y Molecular (FCEFN-UNC) and Instituto de Investigaciones Biológicas y Tecnológicas (CONICET-UNC), Facultad de Ciencias Exactas, Físicas y Naturales, Universidad Nacional de Córdoba, Av. Vélez Sarsfield 1611, 5016 Córdoba, Argentina
| | - M Cubilla
- Centro de Biología Celular y Molecular (FCEFN-UNC) and Instituto de Investigaciones Biológicas y Tecnológicas (CONICET-UNC), Facultad de Ciencias Exactas, Físicas y Naturales, Universidad Nacional de Córdoba, Av. Vélez Sarsfield 1611, 5016 Córdoba, Argentina
| | - H A Guidobaldi
- Centro de Biología Celular y Molecular (FCEFN-UNC) and Instituto de Investigaciones Biológicas y Tecnológicas (CONICET-UNC), Facultad de Ciencias Exactas, Físicas y Naturales, Universidad Nacional de Córdoba, Av. Vélez Sarsfield 1611, 5016 Córdoba, Argentina
| | - A A Bravo
- Centro de Biología Celular y Molecular (FCEFN-UNC) and Instituto de Investigaciones Biológicas y Tecnológicas (CONICET-UNC), Facultad de Ciencias Exactas, Físicas y Naturales, Universidad Nacional de Córdoba, Av. Vélez Sarsfield 1611, 5016 Córdoba, Argentina
| | - L C Giojalas
- Centro de Biología Celular y Molecular (FCEFN-UNC) and Instituto de Investigaciones Biológicas y Tecnológicas (CONICET-UNC), Facultad de Ciencias Exactas, Físicas y Naturales, Universidad Nacional de Córdoba, Av. Vélez Sarsfield 1611, 5016 Córdoba, Argentina
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Wang Q, Mesaros C, Blair IA. Ultra-high sensitivity analysis of estrogens for special populations in serum and plasma by liquid chromatography-mass spectrometry: Assay considerations and suggested practices. J Steroid Biochem Mol Biol 2016; 162:70-9. [PMID: 26767303 PMCID: PMC4931956 DOI: 10.1016/j.jsbmb.2016.01.002] [Citation(s) in RCA: 26] [Impact Index Per Article: 2.9] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 09/14/2015] [Revised: 12/15/2015] [Accepted: 01/04/2016] [Indexed: 11/24/2022]
Abstract
Estrogen measurements play an important role in the clinical evaluation of many endocrine disorders as well as in research on the role of hormones in human biology and disease. It remains an analytical challenge to quantify estrogens and their metabolites in specimens from special populations including older men, children, postmenopausal women and women receiving aromatase inhibitors. Historically, immunoassays have been used for measuring estrogens and their metabolites in biological samples for risk assessment. However, the lack of specificity and accuracy of immunoassay-based methods has caused significant problems when interpreting data generated from epidemiological studies and across different laboratories. Stable isotope dilution (SID) methodology coupled with liquid chromatography-selected reaction monitoring-mass spectrometry (LC-SRM/MS) is now accepted as the 'gold-standard' to quantify estrogens and their metabolites in serum and plasma due to improved specificity, high accuracy, and the ability to monitor multiple estrogens when compared with immunoassays. Ultra-high sensitivity can be obtained with pre-ionized derivatives when using triple quadruple mass spectrometers in the selected reaction monitoring (SRM) mode coupled with nanoflow LC. In this review, we have examined the special issues related to utilizing ultra-high sensitivity SID LC-SRM/MS-based methodology to accurately quantify estrogens and their metabolites in the serum and plasma from populations with low estrogen levels. The major issues that are discussed include: sample preparation for both unconjugated and conjugated estrogens, derivatization, chromatographic separation, matrix effects, and assay validation.
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Affiliation(s)
- Qingqing Wang
- Center of Excellence in Environmental Toxicology and Penn SRP Center, Perelman School of Medicine, University of Pennsylvania Philadelphia, PA 19104, United States; Department of Systems Pharmacology and Translational Therapeutics, Perelman School of Medicine, University of Pennsylvania Philadelphia, PA 19104, United States; Department of Pharmacology and Toxicology, Beijing Institute of Radiation Medicine, Beijing 100850, China
| | - Clementina Mesaros
- Center of Excellence in Environmental Toxicology and Penn SRP Center, Perelman School of Medicine, University of Pennsylvania Philadelphia, PA 19104, United States; Department of Systems Pharmacology and Translational Therapeutics, Perelman School of Medicine, University of Pennsylvania Philadelphia, PA 19104, United States
| | - Ian A Blair
- Center of Excellence in Environmental Toxicology and Penn SRP Center, Perelman School of Medicine, University of Pennsylvania Philadelphia, PA 19104, United States; Department of Systems Pharmacology and Translational Therapeutics, Perelman School of Medicine, University of Pennsylvania Philadelphia, PA 19104, United States.
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Ding W, Bishop ME, Lyn-Cook LE, Davis KJ, Manjanatha MG. In Vivo Alkaline Comet Assay and Enzyme-modified Alkaline Comet Assay for Measuring DNA Strand Breaks and Oxidative DNA Damage in Rat Liver. J Vis Exp 2016:53833. [PMID: 27166647 PMCID: PMC4942029 DOI: 10.3791/53833] [Citation(s) in RCA: 9] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/14/2023] Open
Abstract
Unrepaired DNA damage can lead to genetic instability, which in turn may enhance cancer development. Therefore, identifying potential DNA damaging agents is important for protecting public health. The in vivo alkaline comet assay, which detects DNA damage as strand breaks, is especially relevant for assessing the genotoxic hazards of xenobiotics, as its responses reflect the in vivo absorption, tissue distribution, metabolism and excretion (ADME) of chemicals, as well as DNA repair process. Compared to other in vivo DNA damage assays, the assay is rapid, sensitive, visual and inexpensive, and, by converting oxidative DNA damage into strand breaks using specific repair enzymes, the assay can measure oxidative DNA damage in an efficient and relatively artifact-free manner. Measurement of DNA damage with the comet assay can be performed using both acute and subchronic toxicology study designs, and by integrating the comet assay with other toxicological assessments, the assay addresses animal welfare requirements by making maximum use of animal resources. Another major advantage of the assays is that they only require a small amount of cells, and the cells do not have to be derived from proliferating cell populations. The assays also can be performed with a variety of human samples obtained from clinically or occupationally exposed individuals.
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Affiliation(s)
- Wei Ding
- Division of Genetic and Molecular Toxicology, US FDA/National Center for Toxicological Research;
| | - Michelle E Bishop
- Division of Genetic and Molecular Toxicology, US FDA/National Center for Toxicological Research
| | - Lascelles E Lyn-Cook
- Division of Genetic and Molecular Toxicology, US FDA/National Center for Toxicological Research
| | - Kelly J Davis
- Toxicologic Pathology Associates, US FDA/National Center for Toxicological Research
| | - Mugimane G Manjanatha
- Division of Genetic and Molecular Toxicology, US FDA/National Center for Toxicological Research
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24
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del Refugio Rivera Vega M, Murillo-Vilches MR, Toral-Lopez J, Sanchez EG, Sanchez AT, González-Huerta LM, Cuevas-Covarrubias SA. X-linked ichthyosis in a patient with a novel nonsense mutation in the STS gene. J Dermatol Sci 2015; 80:160-2. [PMID: 26421812 DOI: 10.1016/j.jdermsci.2015.09.004] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/12/2015] [Revised: 09/02/2015] [Accepted: 09/10/2015] [Indexed: 11/16/2022]
Affiliation(s)
| | | | - Jaime Toral-Lopez
- Departamento de Genética Médica, Centro Médico Ecatepec, ISSEMYM Edo. Mexico, Mexico
| | | | | | - Luz M González-Huerta
- Departamento de Genética Médica, Hospital General de México, Facultad de Medicina, UNAM, Mexico City, Mexico
| | - Sergio A Cuevas-Covarrubias
- Departamento de Genética Médica, Hospital General de México, Facultad de Medicina, UNAM, Mexico City, Mexico.
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25
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Seneff S, Davidson RM, Lauritzen A, Samsel A, Wainwright G. A novel hypothesis for atherosclerosis as a cholesterol sulfate deficiency syndrome. Theor Biol Med Model 2015; 12:9. [PMID: 26014131 PMCID: PMC4456713 DOI: 10.1186/s12976-015-0006-1] [Citation(s) in RCA: 20] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/09/2015] [Accepted: 05/18/2015] [Indexed: 12/31/2022] Open
Abstract
BACKGROUND Despite a vast literature, atherosclerosis and the associated ischemia/reperfusion injuries remain today in many ways a mystery. Why do atheromatous plaques make and store a supply of cholesterol and sulfate within the major arteries supplying the heart? Why are treatment programs aimed to suppress certain myocardial infarction risk factors, such as elevated serum homocysteine and inflammation, generally counterproductive? METHODS Our methods are based on an extensive search of the literature in atherosclerotic cardiovascular disease as well as in the area of the unique properties of water, the role of biosulfates in the vascular wall, and the role of electromagnetic fields in vascular flow. Our investigation reveals a novel pathology linked to atherosclerosis that better explains the observed facts than the currently held popular view. RESULTS We propose a novel theory that atherosclerosis can best be explained as being due to cholesterol sulfate deficiency. Furthermore, atheromatous plaques replenish the supply of cholesterol and sulfate to the microvasculature, by exploiting the inflammatory agent superoxide to derive sulfate from homocysteine and other sulfur sources. We argue that the sulfate anions attached to the glycosaminoglycans in the glycocalyx are essential in maintaining the structured water that is crucial for vascular endothelial health and erythrocyte mobility through capillaries. Sulfate depletion leads to cholesterol accumulation in atheromas, because its transport through water-based media depends on sulfurylation. We show that streaming potential induces nitric oxide (NO) release, and NO derivatives break down the extracellular matrix, redistributing sulfate to the microvasculature. We argue that low (less negative) zeta potential due to insufficient sulfate anions leads to hypertension and thrombosis, because these responses can increase streaming potential and induce nitric-oxide mediated vascular relaxation, promoting oxygen delivery. Our hypothesis is a parsimonious explanation of multiple features of atherosclerotic cardiovascular disease. CONCLUSIONS If our interpretation is correct, then it would have a significant impact on how atherosclerosis is treated. We recommend a high intake of sulfur-containing foods as well as an avoidance of exposure to toxicants that may impair sulfate synthesis.
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Affiliation(s)
- Stephanie Seneff
- Computer Science and Artificial Intelligence Laboratory, MIT, Cambridge, MA, 02139, USA.
| | - Robert M Davidson
- Internal Medicine Group Practice, PhyNet, Inc, 4002 Technology Center, Longview, TX, 75605, USA.
| | | | - Anthony Samsel
- Research Scientist and Consultant, Deerfield, NH, 03037, USA.
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Toral-López J, González-Huerta LM, Cuevas-Covarrubias SA. X linked recessive ichthyosis: Current concepts. World J Dermatol 2015; 4:129. [DOI: 10.5314/wjd.v4.i3.129] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 11/22/2014] [Revised: 01/31/2015] [Accepted: 05/28/2015] [Indexed: 02/06/2023] Open
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Sex-specific dose-response analysis of genotoxicity in cyproterone acetate-treated F344 rats. MUTATION RESEARCH-GENETIC TOXICOLOGY AND ENVIRONMENTAL MUTAGENESIS 2014; 774:1-7. [DOI: 10.1016/j.mrgentox.2014.08.005] [Citation(s) in RCA: 7] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Track Full Text] [Subscribe] [Scholar Register] [Received: 07/14/2014] [Revised: 08/18/2014] [Accepted: 08/23/2014] [Indexed: 11/19/2022]
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Zschockelt L, Amelkina O, Siemieniuch MJ, Koster S, Jewgenow K, Braun BC. Corpora lutea of pregnant and pseudopregnant domestic cats reveal similar steroidogenic capacities during the luteal life span. J Steroid Biochem Mol Biol 2014; 144 Pt B:373-81. [PMID: 25138635 DOI: 10.1016/j.jsbmb.2014.08.010] [Citation(s) in RCA: 20] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 04/23/2014] [Revised: 07/31/2014] [Accepted: 08/13/2014] [Indexed: 01/22/2023]
Abstract
In domestic cats, luteal phases of pregnancy and pseudopregnancy (non-pregnant luteal phase) differ in the course and level of plasma progesterone (P4). Therefore, we assumed differences in luteal steroidogenic capacities. Here we present a comprehensive analysis of intraluteal steroid biogenesis in the domestic cat. We quantitatively measured relative mRNA levels of steroidogenic acute regulatory protein (STAR), cytochrome P450 oxidases (CYP), hydroxysteroid dehydrogenases (HSD), steroid reductase (SRD) and enzymes involved in sulfoconjugation of steroids, i.e. sulfotransferase (SULT) and sulfatase (STS). Protein expression was analysed by Western Blot for HSD3B. Additionally, intraluteal steroid contents were determined. During the pseudopregnant luteal phase, expression of STAR (p=0.005), HSD3B1 (p<0.0001), CYP19A1 (p<0.0001) and HSD17B7 (p=0.008) decreased from formation of the corpus luteum (CL) onwards. HSD3B protein expression was highest in the development/maintenance stage of CL and declined during the subsequent luteal phase of pregnancy and pseudopregnancy. This was in accordance with decreasing intraluteal levels of P4, oestrogens and androgens. In contrast, expression of SRD5A1 (p<0.001) increased with progression through stages of the pseudopregnant CL, being indicative of P4 metabolism via an alternate pathway to dihydrotestosterone (DHT). Compared to the formation stage, expression of SULT1E1 was higher in all other luteal stages of pseudopregnancy (p=0.004), implying a potential sulfoconjugation of oestrogens. Expression of CYP11A1 and CYP17A1 was unaffected by the luteal stage (p>0.05), suggesting a permanent capacity of cat CL to convert progestogens via androgen and oestrogen pathways. In general, mRNA expression profiles of steroidogenic enzymes during the pregnant luteal phase reflected the pseudopregnancy profiles. Intraluteal oestrogen (p<0.0001) and androgen (p=0.008) levels were higher in the formation stage compared to the following luteal stages of pseudopregnancy. Concentrations of P4 were higher in the development/maintenance compared to the regression stages (p=0.01). We conclude that cat CL of the same histomorphological stage are characterised by identical steroidogenic capacities independently of an on-going pregnancy.
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Affiliation(s)
- Lina Zschockelt
- Leibniz Institute for Zoo and Wildlife Research, Department of Reproduction Biology, PF700430, 10324 Berlin, Germany.
| | - Olga Amelkina
- Leibniz Institute for Zoo and Wildlife Research, Department of Reproduction Biology, PF700430, 10324 Berlin, Germany.
| | - Marta J Siemieniuch
- Institute of Animal Reproduction and Food Research of the Polish Academy of Sciences, Department of Reproductive Immunology and Pathology, Tuwima Street, Olsztyn 10-748, Poland.
| | - Stefanie Koster
- Leibniz Institute for Zoo and Wildlife Research, Department of Reproduction Biology, PF700430, 10324 Berlin, Germany.
| | - Katarina Jewgenow
- Leibniz Institute for Zoo and Wildlife Research, Department of Reproduction Biology, PF700430, 10324 Berlin, Germany.
| | - Beate C Braun
- Leibniz Institute for Zoo and Wildlife Research, Department of Reproduction Biology, PF700430, 10324 Berlin, Germany.
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Samavat H, Kurzer MS. Estrogen metabolism and breast cancer. Cancer Lett 2014; 356:231-43. [PMID: 24784887 DOI: 10.1016/j.canlet.2014.04.018] [Citation(s) in RCA: 239] [Impact Index Per Article: 21.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/22/2014] [Revised: 04/01/2014] [Accepted: 04/19/2014] [Indexed: 01/18/2023]
Abstract
There is currently accumulating evidence that endogenous estrogens play a critical role in the development of breast cancer. Estrogens and their metabolites have been studied in both pre- and postmenopausal women with more consistent results shown in the latter population, in part because of large hormonal variations during the menstrual cycle and far fewer studies having been performed in premenopausal women. In this review we describe in detail estrogen metabolism and associated genetic variations, and provide a critical review of the current literature regarding the role of estrogens and their metabolites in breast cancer risk.
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Affiliation(s)
- Hamed Samavat
- Department of Food Science and Nutrition, University of Minnesota, St. Paul, MN, USA
| | - Mindy S Kurzer
- Department of Food Science and Nutrition, University of Minnesota, St. Paul, MN, USA.
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Establishment of metabolism and transport pathways in the rodent and human fetal liver. Int J Mol Sci 2013; 14:23801-27. [PMID: 24322441 PMCID: PMC3876079 DOI: 10.3390/ijms141223801] [Citation(s) in RCA: 20] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/05/2013] [Revised: 11/25/2013] [Accepted: 11/26/2013] [Indexed: 12/16/2022] Open
Abstract
The ultimate fate of drugs and chemicals in the body is largely regulated by hepatic uptake, metabolism, and excretion. The liver acquires the functional ability to metabolize and transport chemicals during the perinatal period of development. Research using livers from fetal and juvenile rodents and humans has begun to reveal the timing, key enzymes and transporters, and regulatory factors that are responsible for the establishment of hepatic phase I and II metabolism as well as transport. The majority of this research has been limited to relative mRNA and protein quantification. However, the recent utilization of novel technology, such as RNA-Sequencing, and the improved availability and refinement of functional activity assays, has begun to provide more definitive information regarding the extent of hepatic drug disposition in the developing fetus. The goals of this review are to provide an overview of the early regulation of the major phase I and II enzymes and transporters in rodent and human livers and to highlight potential mechanisms that control the ontogeny of chemical metabolism and excretion pathways.
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31
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Srivastava DP, Woolfrey KM, Penzes P. Insights into rapid modulation of neuroplasticity by brain estrogens. Pharmacol Rev 2013; 65:1318-50. [PMID: 24076546 PMCID: PMC3799233 DOI: 10.1124/pr.111.005272] [Citation(s) in RCA: 95] [Impact Index Per Article: 7.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/22/2022] Open
Abstract
Converging evidence from cellular, electrophysiological, anatomic, and behavioral studies suggests that the remodeling of synapse structure and function is a critical component of cognition. This modulation of neuroplasticity can be achieved through the actions of numerous extracellular signals. Moreover, it is thought that it is the integration of different extracellular signals regulation of neuroplasticity that greatly influences cognitive function. One group of signals that exerts powerful effects on multiple neurologic processes is estrogens. Classically, estrogens have been described to exert their effects over a period of hours to days. However, there is now increasing evidence that estrogens can rapidly influence multiple behaviors, including those that require forebrain neural circuitry. Moreover, these effects are found in both sexes. Critically, it is now emerging that the modulation of cognition by rapid estrogenic signaling is achieved by activation of specific signaling cascades and regulation of synapse structure and function, cumulating in the rewiring of neural circuits. The importance of understanding the rapid effects of estrogens on forebrain function and circuitry is further emphasized as investigations continue to consider the potential of estrogenic-based therapies for neuropathologies. This review focuses on how estrogens can rapidly influence cognition and the emerging mechanisms that underlie these effects. We discuss the potential sources and the biosynthesis of estrogens within the brain and the consequences of rapid estrogenic-signaling on the remodeling of neural circuits. Furthermore, we argue that estrogens act via distinct signaling pathways to modulate synapse structure and function in a manner that may vary with cell type, developmental stage, and sex. Finally, we present a model in which the coordination of rapid estrogenic-signaling and activity-dependent stimuli can result in long-lasting changes in neural circuits, contributing to cognition, with potential relevance for the development of novel estrogenic-based therapies for neurodevelopmental or neurodegenerative disorders.
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Affiliation(s)
- Deepak P Srivastava
- Department of Neuroscience & Centre for the Cellular Basis of Behaviour, 125 Coldharbour Lane, The James Black Centre, Institute of Psychiatry, King's College London, London, SE5 9NU, UK.
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32
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Bai X, Casey FXM, Hakk H, DeSutter TM, Oduor PG, Khan E. Dissipation and transformation of 17β-estradiol-17-sulfate in soil-water systems. JOURNAL OF HAZARDOUS MATERIALS 2013; 260:733-9. [PMID: 23846123 DOI: 10.1016/j.jhazmat.2013.06.036] [Citation(s) in RCA: 9] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Received: 03/26/2013] [Revised: 06/10/2013] [Accepted: 06/16/2013] [Indexed: 05/22/2023]
Abstract
In the environment, estrogen conjugates can be precursors to the endocrine-disrupting free estrogens, 17β-estradiol (E2) and estrone (E1). Compared to other estrogen conjugates, 17β-estradiol-17-sulfate (E2-17S) is detected at relatively high concentrations and frequencies in animal manure and surface runoff from fields receiving manure. To elucidate the lifecycle of manure-borne estrogens and their conjugates in the environment, the fate of radiolabelled E2-17S in agricultural soils was investigated using laboratory batch studies with soils of different organic carbon (OC) content (1.29% for topsoil versus 0.26% for subsoil). E2-17S was found relatively persistent in the aqueous phase throughout the duration of the 14 d experiment. The aqueous E2-17S persisted longer in the subsoil (half-lives (DT₅₀)=64-173 h) than the topsoil (DT₅₀=4.9-26 h), and the aqueous persistence of E2-17S depended on its initial concentration. The major transformation pathway was hydroxylation, yielding mono- and di-hydroxy-E2-17S (OH-E2-17S and diOH-E2-17S). Free estrogens, E2 and E1, were only observed in the sorbed phase of the soil at low concentrations (∼1% of applied dose), which demonstrated that deconjugation and subsequent oxidation had occurred. Although deconjugation was not a major pathway, E2-17S could be a precursor of free estrogens in the environment.
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Affiliation(s)
- Xuelian Bai
- Department of Soil Science, North Dakota State University, Fargo, ND 58108-6050, USA
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Roy J, Lefebvre J, Maltais R, Poirier D. Inhibition of dehydroepiandosterone sulfate action in androgen-sensitive tissues by EM-1913, an inhibitor of steroid sulfatase. Mol Cell Endocrinol 2013; 376:148-55. [PMID: 23806558 DOI: 10.1016/j.mce.2013.06.022] [Citation(s) in RCA: 14] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 02/06/2013] [Revised: 05/09/2013] [Accepted: 06/17/2013] [Indexed: 11/23/2022]
Abstract
Steroid sulfatase (STS) plays an important role in the formation of estrogens and androgens by allowing the conversion of inactive circulating sulfated steroids into active hormones. These steroids support the development and growth of a number of hormone-dependent cancers, including prostate cancer. Here, we tested a non-estrogenic and non-androgenic inhibitor of steroid STS, namely EM-1913, with special attention to its potential use in the treatment of prostate cancer. After determining the required dosage of dehydroepiandrosterone sulfate (DHEAS) needed to stimulate the ventral prostate and seminal vesicles in castrated rats, we measured that EM-1913 partially (26%) and almost entirely blocked (81%) the stimulating effect of DHEAS on ventral prostates and seminal vesicles, respectively. In addition, the homogenization of these two tissues allowed us to confirm that they were completely deprived of STS activity following a treatment with EM-1913. This effect is also reflected in blood, since the plasma level of DHEAS was increased in animals treated with EM-1913, whereas the levels of dehydroepiandrosterone (DHEA) and dihydrotestosterone (DHT), two DHEAS metabolites, meanwhile decreased. From these results, we concluded that STS inhibitor EM-1913 is a good candidate for additional preclinical studies.
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Affiliation(s)
- Jenny Roy
- Laboratory of Medicinal Chemistry, CHU de Québec - Research Center Endocrinology and Nephrology Unit and Faculty of Medicine, Université Laval, Québec, Canada
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Li HH, Zhao YJ, Li Y, Dai CF, Jobe SO, Yang XS, Li XF, Patankar MS, Magness RR, Zheng J. Estradiol 17β and its metabolites stimulate cell proliferation and antagonize ascorbic acid-suppressed cell proliferation in human ovarian cancer cells. Reprod Sci 2013; 21:102-11. [PMID: 23757313 DOI: 10.1177/1933719113492211] [Citation(s) in RCA: 21] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/09/2023]
Abstract
Estradiol 17β (E2β) and ascorbic acid (AA) have been implicated in cancer progression. However, little is known about the actions of biologically active metabolites of E2β, 2-hydroxyestradiol (2OHE2), 4-hydroxyestradiol (4OHE2), 2-methoxyestradiol (2ME2), and 4-methoxyestradiol (4ME2) synthesized sequentially by cytochrome P450, family 1, subfamily A (CYP1A1) and B (CYP1B1), polypeptide 1, and catechol-O-methyltransferase (COMT) on ovarian cancer. Herein, we examined the expression of CYP1A1, CYP1B1, COMT, and estrogen receptor α (ERα) and β (ERβ) in human ovarian surface epithelial (IOSE-385) and cancer cell lines (OVCAR-3, SKOV-3, and OVCA-432). We also investigated the roles of E2β, 2OHE2, 4OHE2, 2ME2, and 4ME2 in cell proliferation, and their interactive effects with AA on ovarian cells. We found the expression of CYP1A1, CYP1B1, COMT, ERα, and ERβ in most cell lines tested. Treating cells with physiological concentrations of E2β and its metabolites promoted (13%-42% of the control) IOSE-385 and OVCAR-3 proliferation. The ER blockade inhibited IOSE-385 (∼76%) and OVCAR-3 (∼87%) proliferative response to E2β but not to its metabolites. The ERα blockade inhibited (∼85%) E2β-stimulated OVCAR-3 proliferation, whereas ERβ blockade attenuated (∼83%) E2β-stimulated IOSE-385 proliferation. The AA at ≥250 μmol/L completely inhibited serum-stimulated cell proliferation in all cell lines tested; however, such inhibition in IOSE-385, OVCAR-3, and OVCA-432 was partially (∼10%-20%) countered by E2β and its metabolites. Thus, our findings indicate that E2β and its metabolites promote cell proliferation and antagonize the AA-suppressed cell proliferation in a subset of ovarian cancer cells, suggesting that blocking the actions of E2β and its metabolites may enhance AA's antiovarian cancer activity.
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Affiliation(s)
- Hui-Hui Li
- 1Department of Obstetrics and Gynecology, Qilu Hospital, Shandong University, Shandong, China
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Abstract
Estradiol (E(2)) is an important modifier of the activity of the fetal hypothalamus-pituitary-adrenal axis. We have reported that estradiol-3-sulfate (E(2)SO(4)) circulates in fetal blood in far higher concentrations than E(2) and that the fetal brain expresses steroid sulfatase, required for local deconjugation of E(2)SO(4). We performed the present study to test the hypothesis that chronic infusion of E(2)SO(4) chronically increases ACTH and cortisol secretion and that it shortens gestation. Chronically catheterized fetal sheep were treated with E(2)SO(4) intracerebroventricular (n = 5), E(2)SO(4) iv (n = 4), or no steroid infusion (control group, n = 5). Fetuses were subjected to arterial blood sampling every other day until spontaneous birth for plasma hormone analysis. Treatment with E(2)SO(4) attenuated preparturient increases in ACTH secretion near term without affecting the ontogenetic rise in plasma cortisol. Infusion of E(2)SO(4) intracerebroventricularly significantly increased plasma E(2), plasma E(2)SO(4), and plasma progesterone and shortened gestation compared with all other groups. These results are consistent with the conclusion that E(2)SO(4): 1) interacts with the hypothalamus-pituitary-adrenal axis primarily by stimulating cortisol secretion and inhibiting ACTH and pro-ACTH secretion by negative feedback; and 2) stimulates the secretion of E(2) and E(2)SO(4). We conclude that the endocrine response to E(2)SO(4) in the fetus is not identical with the response to E(2).
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Affiliation(s)
- Charles E Wood
- Department of Physiology and Functional Genomics, University of Florida College of Medicine, Gainesville, Florida 32610-0274, USA. mail
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Abstract
A female hormone, estrogen, is linked to breast cancer incidence. Estrogens undergo phase I and II metabolism by which they are biotransformed into genotoxic catechol estrogen metabolites and conjugate metabolites are produced for excretion or accumulation. The molecular mechanisms underlying estrogen-mediated mammary carcinogenesis remain unclear. Cell proliferation through activation of estrogen receptor (ER) by its agonist ligands and is clearly considered as one of carcinogenic mechanisms. Recent studies have proposed that reactive oxygen species generated from estrogen or estrogen metabolites are attributed to genotoxic effects and signal transduction through influencing redox sensitive transcription factors resulting in cell transformation, cell cycle, migration, and invasion of the breast cancer. Conjuguation metabolic pathway is thought to protect cells from genotoxic and cytotoxic effects by catechol estrogen metabolites. However, methoxylated catechol estrogens have been shown to induce ER-mediated signaling pathways, implying that conjugation is not a simply detoxification pathway. Dual action of catechol estrogen metabolites in mammary carcinogenesis as the ER-signaling molecules and chemical carcinogen will be discussed in this review.
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Affiliation(s)
- Minsun Chang
- Department of Medical and Pharmaceutical Science, College of Science, Sookmyung Women's University, Seoul, Korea.
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Winikor J, Schlaerth C, Rabaglino MB, Cousins R, Sutherland M, Wood CE. Complex actions of estradiol-3-sulfate in late gestation fetal brain. Reprod Sci 2011; 18:654-65. [PMID: 21273638 PMCID: PMC3235910 DOI: 10.1177/1933719110395400] [Citation(s) in RCA: 11] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/16/2023]
Abstract
The most abundant form of estrogen circulating in fetal plasma is sulfo-conjugated estrogen; for example, estradiol-3-sulfate (E(2)SO(4)) is more highly abundant than estradiol (E(2)). The present study investigated the ontogeny of the deconjugating (steroid sulfatase [STS]) and conjugating (estrogen sulfotransferase [STF]) enzymes in ovine fetal brain and tested the hypothesis that treatment with E(2)SO(4) would alter the expression of one or both enzymes. Steroid sulfatase was more highly expressed than STF, and both changed as a function of gestational age. Estradiol-3-sulfate infused intracerebroventricularly (icv) significantly increased plasma adrenocorticotropic hormone (ACTH) and cortisol concentrations. Plasma E(2) and E(2)SO(4) were increased, and brain expression of estrogen receptor α was decreased. The proteins STS and STF were up- and downregulated, respectively. Pituitary proopiomelanocortin (POMC) and follicle-stimulating hormone (FSH) and hypothalamic corticotrophin-releasing hormone (CRH) messenger RNA (mRNA) was decreased. We conclude that E(2)SO(4) has complex actions on the fetal brain, which might involve deconjugation by STS, but that the net result of direct E(2)SO(4) icv infusion is more complex than can be accounted for by infusion of E(2) alone.
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Affiliation(s)
- Jared Winikor
- Department of Physiology and Functional Genomics, University of Florida College of Medicine, FL, USA
| | - Christine Schlaerth
- Department of Physiology and Functional Genomics, University of Florida College of Medicine, FL, USA
| | - Maria Belen Rabaglino
- Department of Physiology and Functional Genomics, University of Florida College of Medicine, FL, USA
| | - Roderick Cousins
- Department of Physiology and Functional Genomics, University of Florida College of Medicine, FL, USA
| | - Monique Sutherland
- Department of Physiology and Functional Genomics, University of Florida College of Medicine, FL, USA
| | - Charles E. Wood
- Department of Physiology and Functional Genomics, University of Florida College of Medicine, FL, USA
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Zalata A, Hassan A, Christophe A, Comhaire F, Mostafa T. Cholesterol and desmosterol in two sperm populations separated on Sil-Select gradient. INTERNATIONAL JOURNAL OF ANDROLOGY 2010; 33:528-535. [PMID: 19490187 DOI: 10.1111/j.1365-2605.2009.00961.x] [Citation(s) in RCA: 23] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 02/05/2023]
Abstract
Sperm lipids are important for sperm viability, maturity and function. This study aimed to identify cholesterol and desmosterol composition of human spermatozoa of two sperm populations separated on Sil-Select gradient. Forty-eight males were divided into four groups namely healthy men (n = 13), asthenozoospermia (n = 11), asthenoteratozoospermia (n = 10) and oligoasthenoteratozoospermia (n = 14). Sperm cholesterol and desmosterol were estimated in two human sperm population separated by centrifugation in a discontinuous Sil-Select gradient. The results showed that cholesterol and desmosterol were the major sterols in human spermatozoa. Spermatozoa recovered from upper/lower layer interface (fraction I) had low fertilization potential, while those from the base (fraction II) had high fertilization potential. Median values of cholesterol and desmosterol in fraction I were 2.55 micromol and 0.77 micromol/10(9) spermatozoa and in fraction II were 1.16 micromol and 0.27 micromol/10(9) spermatozoa. Cholesterol/desmosterol ratio was significantly higher in fraction II than I (4.8 vs. 3.2, p < 0.01). Cholesterol, desmosterol, total phospholipids and sterols/phospholipids were negatively correlated with sperm concentrations, sperm motility, linear velocity, normal sperm morphology and acrosome reaction percentage whereas cholesterol/desmosterol ratio was positively correlated with these parameters. It is concluded that the difference in sterol composition of sperm subpopulations separated on Sil-Select gradient suggests that composition of sterols is related to sperm functions.
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Affiliation(s)
- A Zalata
- Medical Biochemistry Department, Faculty of Medicine, Mansoura University, Mansoura, Egypt
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Cole GB, Keum G, Liu J, Small GW, Satyamurthy N, Kepe V, Barrio JR. Specific estrogen sulfotransferase (SULT1E1) substrates and molecular imaging probe candidates. Proc Natl Acad Sci U S A 2010; 107:6222-7. [PMID: 20304798 PMCID: PMC2852016 DOI: 10.1073/pnas.0914904107] [Citation(s) in RCA: 49] [Impact Index Per Article: 3.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/24/2022] Open
Abstract
This work focuses on the development of specific substrates for estrogen sulfotransferase (SULT1E1) to produce molecular imaging probes for this enzyme. SULT1E1 is a key enzyme in estrogen homeostasis, playing a central role in the prevention and development of human disease. In vitro sulfation assays showed alkyl and aryl substitutions to a fused heterocyclic system modeled after beta-naphthol (betaN), based on compounds that interact with the estrogen receptor, rendered several molecules with enhanced specificity for SULT1E1 over SULT1A1*1, SULT1A1*2, SULT1A3, and SULT2A1. Several 6-hydroxy-2-arylbenzothiazoles tested demonstrated excellent affinity--V(max)/K(m) ratios-and specificity for SULT1E1. K(m) values ranged from 0.12-2.36 microM. A strong correlation was observed between polarity of the 4'-sustituent on the 2-aryl moiety (Hammett sigma(p)) and the log(V(max)/K(m)) (r = 0.964). Substrate sensitivity is influenced by the acidity of the 6-phenolic group demonstrated by correlating its (1)H NMR chemical shift (delta(OH)) with the log(V(max)/K(m)) (r = 0.963). Acidity is mediated by the electron withdrawing capacity of the 4'-substituent outlined by the correlation of the C-2 (13)C NMR chemical shift (delta(C2)) with the log(V(max)/K(m)) (r = 0.987). 2-[4-(Methylamino)phenyl]-6-hydroxybenzothiazole (2b) was radiolabeled with carbon-11 ((11)C-(2b)) and used in vivo for microPET scanning and tissue metabolite identification. High PET signal was paralleled with the presence of radiolabeled (11)C-(2b)-6-O-sulfate and the SULT1E1 protein detected by western blot. Because this and other members of this family presenting specificity for SULT1E1 can be labeled with carbon-11 or fluorine-18, in vivo assays of SULT1E1 functional activity are now feasible in humans.
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Affiliation(s)
- Graham B. Cole
- Departments of Molecular and Medical Pharmacology and Psychiatry and Biobehavioral Sciences, David Geffen School of Medicine at University of California, Los Angeles, CA 90095-6948
| | - Gyochang Keum
- Departments of Molecular and Medical Pharmacology and Psychiatry and Biobehavioral Sciences, David Geffen School of Medicine at University of California, Los Angeles, CA 90095-6948
| | - Jie Liu
- Departments of Molecular and Medical Pharmacology and Psychiatry and Biobehavioral Sciences, David Geffen School of Medicine at University of California, Los Angeles, CA 90095-6948
| | - Gary W. Small
- Departments of Molecular and Medical Pharmacology and Psychiatry and Biobehavioral Sciences, David Geffen School of Medicine at University of California, Los Angeles, CA 90095-6948
| | - Nagichettiar Satyamurthy
- Departments of Molecular and Medical Pharmacology and Psychiatry and Biobehavioral Sciences, David Geffen School of Medicine at University of California, Los Angeles, CA 90095-6948
| | - Vladimir Kepe
- Departments of Molecular and Medical Pharmacology and Psychiatry and Biobehavioral Sciences, David Geffen School of Medicine at University of California, Los Angeles, CA 90095-6948
| | - Jorge R. Barrio
- Departments of Molecular and Medical Pharmacology and Psychiatry and Biobehavioral Sciences, David Geffen School of Medicine at University of California, Los Angeles, CA 90095-6948
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Stute P, Szuwart T, Schlueter M, Götte M, Packeisen J, Kiesel L. Effects of hormone therapy on estrogen synthesis from E1S in the mammary gland of postmenopausal women. Maturitas 2008; 59:163-73. [DOI: 10.1016/j.maturitas.2007.12.005] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/31/2007] [Revised: 12/07/2007] [Accepted: 12/13/2007] [Indexed: 10/22/2022]
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Stute P, Register TC, Blair RM, Cline JM. Effects of tibolone on estrogen biosynthesis in the mammary tissue of postmenopausal monkeys. Menopause 2008; 13:232-40. [PMID: 16645537 DOI: 10.1097/01.gme.0000198487.55456.0e] [Citation(s) in RCA: 11] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 10/24/2022]
Abstract
OBJECTIVE To evaluate the long-term effects of tibolone on estrone sulfate (E1S)-sulfatase activity in breast tissue in a primate model (Macaca fascicularis) in comparison with conventional hormone therapies. DESIGN Ovariectomized female animals (n = 112) were randomized into five groups and treated for 2 years. Treatment included tibolone at 0.05 mg/kg (LoTib, n = 23) or 0.2 mg/kg (HiTib, n = 23), conjugated equine estrogens at 0.042 mg/kg (CEE, n = 24), CEE + medroxyprogesterone acetate at 0.167 mg/kg (CEE+MPA, n = 21), or placebo (controls, n = 21). E1S-sulfatase activity was evaluated by incubating homogenized breast tissue with [H]-E1S. Thin-layer chromatography was performed to separate the products estrone (E1) and estradiol (E2). Histomorphometry was performed to measure the amount of epithelial and fat tissue in the mammary gland. RESULTS Significantly more E2 than E1 was produced in all groups. E1S-sulfatase activity did not differ among the groups. E1S-sulfatase activity was highest in HiTib animals with less fatty breasts (5.9 fmol total estrogen/mg of protein/min; P < or =0.05) and lowest in HiTib animals with more fatty breasts (2.8 fmol total estrogen/mg of protein/min; P = 0.004 relative to less fatty breasts). CONCLUSIONS We conclude that tibolone had a differential effect on local estrogen biosynthesis in animals with high and low breast fat content. Therefore, breast tissue composition affects the steroidogenic response to hormonal treatment.
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Affiliation(s)
- Petra Stute
- Department of Obstetrics and Gynecology, University of Münster, Münster, Germany
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Stute P, Nisslein T, Götte M, Kamischke A, Kiesel L, Klockenbusch W. Effects of black cohosh on estrogen biosynthesis in normal breast tissue in vitro. Maturitas 2007; 57:382-91. [PMID: 17548177 DOI: 10.1016/j.maturitas.2007.04.007] [Citation(s) in RCA: 16] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/21/2006] [Revised: 04/16/2007] [Accepted: 04/20/2007] [Indexed: 11/17/2022]
Abstract
OBJECTIVES To investigate the effect of black cohosh on the estrogen biosynthesis in the breast in vitro. METHODS Steroid sulfatase (STS) activity was studied in normal breast tissue obtained from pre- and postmenopausal women undergoing reduction mammoplasty. STS protein expression was studied by immunohistochemistry and western blotting. Breast tissue was incubated in vitro without or with black cohosh (iCR) at concentrations ranging from 0.1mg/ml to 1 ng/ml. STS activity was evaluated by incubating homogenized breast tissue with [3H]-estrone sulfate, separating the formed products, estrone (E1) and estradiol (E2), by thin layer chromatography and measuring the amounts of E1 and E2 by scintillation counting. RESULTS STS protein expression and enzymatic activity were detected in all specimens investigated. In all groups, significantly more E1 than E2 was produced. Local estrogen formation was decreased in premenopausal breast tissue by treatment with iCR at 0.1mg/ml (p<or=0.05). CONCLUSIONS iCR decreases local estrogen formation in normal human breast tissue in vitro. This may contribute to the lack of hormonal effects of black cohosh in breast tissue observed in previous studies.
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Affiliation(s)
- Petra Stute
- Department of Obstetrics and Gynecology, University Clinic of Muenster, 48149 Muenster, Germany.
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Aulenta F, Berndt M, Brüdgam I, Hartl H, Sörgel S, Reissig HU. A New, Efficient and Stereoselective Synthesis of Tricyclic and Tetracyclic Compounds by Samarium Diiodide Induced Cyclisations of Naphthyl-Substituted Arylketones—An Easy Access to Steroid-Like Skeletons. Chemistry 2007; 13:6047-62. [PMID: 17487910 DOI: 10.1002/chem.200700057] [Citation(s) in RCA: 36] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/09/2022]
Abstract
In this report, we present the application of samarium diiodide induced cyclisations of naphthyl-substituted ketones towards an easy and stereoselective access to tri- and tetracyclic-functionalised compounds. Typical naphthalene derivatives were studied to investigate the scope and limitations of this novel cyclisation process. The model substrates studied demonstrate that the samarium ketyl cyclisations are essentially restricted to the formation of six-membered rings. The diastereoselectivity of these reactions is strongly influenced by the connection of the alkyl side chain to the naphthalene core. Gamma-naphth-1-yl-substituted ketones furnished cyclisation products, such as 17 or 22-26, as single diastereomers, whereas gamma-naphth-2-yl-substituted precursors gave mixtures of diastereomers--as demonstrated by the conversion of model compound 10 into tricyclic products 18 a/18 b, or that of cyclohexanone derivative 33 into tetracyclic diastereomers 34 a/34 b. Cyclic ketones as ketyl precursors furnished steroid-like tetracyclic skeletons; however, due to the cis/cis fusion of rings B/C and C/D these products have an "unnatural" bowl-like shape. Several of the cyclisation products have been identified by X-ray analyses, which not only proved the constitutions, but also the relative configurations and the preferred conformations. Steroid analogue 23 was subjected to subsequent transformations, which demonstrate that the styrene-like double bond of such compounds can be used for further structural diversification. First attempts to synthesise related azasteroids by incorporating nitrogen atoms into the ketone moiety are also reported. Thus, pyrrolidine derivatives 44 and 47 as well as piperidine derivatives 50 and 52 were subjected to samarium diiodide induced cyclisations. The expected tetracyclic products 48, 49 a/49 b, 51 and 53 a/53 b were obtained in moderate to good yields. The stereoselectivities observed follow the rules already established for the all-carbon precursors. The resulting products, bearing a nitrogen atom in ring D, are interesting azasteroid analogues with "unnatural" configuration.
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Affiliation(s)
- Francesca Aulenta
- Institut für Chemie und Biochemie, Freie Universität Berlin, Takustrasse 3, 14195 Berlin, Germany
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44
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Stute P, Götte M, Kiesel L. Differential effect of hormone therapy on E1S-sulfatase activity in non-malignant and cancerous breast cells in vitro. Breast Cancer Res Treat 2007; 108:363-74. [PMID: 17546497 DOI: 10.1007/s10549-007-9615-7] [Citation(s) in RCA: 8] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/09/2007] [Accepted: 05/07/2007] [Indexed: 11/28/2022]
Abstract
Breast tissue possesses the enzymes for local estrogen biosynthesis. We measured the effect of Estradiol (E2), Tibolone (OrgOD14) and its metabolite Org4094 on estrone sulfate (E1S)-sulfatase (STS) using breast cancer (MCF-7) and non-malignant breast cells (HBL-100). Cells were cultured in 5% steroid depleted fetal calf serum for 3 days and subsequently incubated with each steroid for either 24 h or directly in cell extracts. STS mRNA and protein expression, and its subcellular localization were determined by semi-quantitative RT-PCR, immunoblotting, and confocal immunofluorescence microscopy. STS activity was evaluated by incubating homogenized breast cells with [(3)H]-E1S. The products E1 and E2 were separated by thin layer chromatography. STS was co-localized with the Golgi marker protein GM130 and the endoplasmic reticulum marker protein calnexin. Treatment did not significantly alter STS mRNA expression. STS protein expression was increased by each steroid in HBL-100 cells but by E2 only in MCF-7 cells. 24 h incubation with OrgOD14 and Org4094 did not alter STS activity in both cell lines. However, STS activity was significantly diminished in HBL-100 but slightly increased in MCF-7 cells by 24 h treatment with E2. "Direct" incubation of cell extracts, eliminating cellular regulation of metabolism, reduced estrogen biosynthesis regardless of cell line and treatment. In conclusion, the immediate reduction of estrogen biosynthesis by OrgOD14 is counteracted by an increased STS protein expression. On the contrary, E2 exerts a differential effect on STS in HBL-100 and MCF-7 cells. The transition from normal to malignant breast cells may be accompanied by an abolished autoregulation of local estrogen formation.
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Affiliation(s)
- Petra Stute
- Department of Obstetrics and Gynecology, Münster University Hospital, Muenster, Germany.
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45
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Marsolais F, Boyd J, Paredes Y, Schinas AM, Garcia M, Elzein S, Varin L. Molecular and biochemical characterization of two brassinosteroid sulfotransferases from Arabidopsis, AtST4a (At2g14920) and AtST1 (At2g03760). PLANTA 2007; 225:1233-44. [PMID: 17039368 DOI: 10.1007/s00425-006-0413-y] [Citation(s) in RCA: 64] [Impact Index Per Article: 3.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Received: 07/31/2006] [Revised: 09/18/2006] [Accepted: 09/18/2006] [Indexed: 05/12/2023]
Abstract
Mammalian sulfotransferases (EC 2.8.2) are involved in many important facets of steroid hormone activity and metabolism. In this study, Arabidopsis AtST4a and AtST1 were identified and characterized as brassinosteroid sulfotransferases that appear to be involved in different aspects of hormone regulation. The two proteins share 44% identity in amino acid sequence, and belong to different plant sulfotransferase families. AtST4a was specific for biologically active end products of the brassinosteroid pathway. The enzyme sulfated brassinosteroids with diverse side-chain structures, including 24-epibrassinosteroids and the naturally occurring (22R, 23R)-28-homobrassinosteroids. AtST4a belongs to a small subfamily of sulfotransferases having two other members, AtST4b and -c. Among the three recombinant enzymes, only AtST4a was catalytically active with brassinosteroids. Transcript expression of AtST4 subfamily members was largely specific to the root. AtST4b- and -c transcript levels were induced by treatment with trans-zeatin, while AtST4a was repressed under the same conditions, supporting a divergent function of AtST4a. Co-regulation of AtST4b and -c correlated with their location in tandem on chromosome 1. AtST1 was stereospecific for 24-epibrassinosteroids, with a substrate preference for the metabolic precursor 24-epicathasterone, and exhibited catalytic activity with hydroxysteroids and estrogens. To gain more insight into this dual activity with plant and mammalian steroids, enzymatic activities of human steroid sulfotransferases toward brassinosteroids were characterized. The dehydroepiandrosterone sulfotransferase SULT2A1 displayed catalytic activity with a selected set of 24-epibrassinolide precursors, including 24-epicathasterone, with specific activities comparable to that measured for the endogenous substrate dehydroepiandrosterone. The comparable activity profiles of AtST1 and SULT2A1 suggest a similar architecture of the acceptor-binding site between the two enzymes, and may potentially reflect a common ability to conjugate certain xenobiotics.
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Affiliation(s)
- Frédéric Marsolais
- Agriculture and Agri-Food Canada, Southern Crop Protection and Food Research Centre, 1391 Sandford St., London, ON, N5V 4T3, Canada.
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46
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Gershon E, Hourvitz A, Reikhav S, Maman E, Dekel N. Low expression of COX-2, reduced cumulus expansion, and impaired ovulation in SULT1E1-deficient mice. FASEB J 2007; 21:1893-901. [PMID: 17341680 DOI: 10.1096/fj.06-7688com] [Citation(s) in RCA: 37] [Impact Index Per Article: 2.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/23/2023]
Abstract
The SULT1E1-encoded estrogen sulfotransferase (EST) catalyzes sulfation of estrogen, resulting in its inactivation. Reduced fertility observed in SULT1E1 knockout (KO) female mice has previously been attributed to the deleterious effect of chronic exposure to high levels of circulating estrogen on placental function. We herein suggest that, in addition to placental dysfunction, this phenotype demonstrates that an excess of estrogen impairs ovulation. The role of SULT1E1 in ovulation is suggested by the substantially low ovulatory response in hCG-treated SULT1E1 KO mice; a similar effect was observed when 17beta-estradiol was administered to wild-type (WT) females. The normal rate of ovulation in SULT1E1 KO females may be restored by PGE2. Along this line, ovaries of human Chorionic Gonadotropin (hCG)-treated SULT1E1 KO mice expressed low levels of cyclooxygenase-2 (COX-2) and its downstream TSG6; moreover, their ovaries contained a reduced number of expanded cumuli. Our results demonstrate, for the first time, that estrogen inactivation may allow the expression of COX-2 and subsequent cumulus expansion, enabling normal ovulation. Our findings may be applied to novel treatments of human ovulatory failure.
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Affiliation(s)
- Eran Gershon
- Department of Biological Regulation, Weizmann Institute of Science, P.O.B. 26, Rehovot 76100, Israel
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47
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Starlard-Davenport A, Xiong Y, Bratton S, Gallus-Zawada A, Finel M, Radominska-Pandya A. Phenylalanine(90) and phenylalanine(93) are crucial amino acids within the estrogen binding site of the human UDP-glucuronosyltransferase 1A10. Steroids 2007; 72:85-94. [PMID: 17174996 PMCID: PMC1829494 DOI: 10.1016/j.steroids.2006.11.016] [Citation(s) in RCA: 26] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 10/27/2006] [Revised: 11/08/2006] [Accepted: 11/13/2006] [Indexed: 11/26/2022]
Abstract
Human UDP-glucuronosyltransferase 1A10 has been identified as the major isoform involved in the biotransformation of a wide range of phenolic substrates, including native estrogens and their oxidized metabolites. Our recent studies point to the F(90)-M(91)-V(92)-F(93) amino acid motif of UGT1A10, which was identified using photoaffinity labeling followed by LC-MS/MS analysis, as a key determinant of the binding of phenolic substrates. In this report, we have evaluated the role of F(90), V(92), and F(93) in the recognition of estrogens by UGT1A10 using site-directed mutagenesis. Kinetic studies using five mutants revealed that F(90) and F(93) are critical residues for the recognition of all estrogen substrates. The substitution of F(90) with alanine totally abolished the activity of this enzyme toward all the estrogens investigated. Overall, sequential removal for the aromatic ring (F to L) and of the hydrophobic chain (F to A and V to A) from amino acids 90, 92, and 93 effectively alters estrogen recognition. This demonstrates that individual features of the native and hydroxylated estrogens determine the specific binding properties of the compound within the binding site of the human UGT1A10 and the mutants. The resulting activities are completely abolished, unchanged, increased, or decreased depending on the structures of both the mutant and the substrate. The novel identification of UGT1A10 as the major isoform involved in the glucuronidation of all estrogens and the discovery of the importance of the FMVF motif in the binding of steroids will help to elucidate the molecular mechanism of glucuronidation, resulting in the design of more effective estrogen-based therapies.
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Affiliation(s)
- Athena Starlard-Davenport
- Department of Biochemistry and Molecular Biology, University of Arkansas for Medical Sciences, Little Rock, AR 72205, USA
| | - Yan Xiong
- Department of Biochemistry and Molecular Biology, University of Arkansas for Medical Sciences, Little Rock, AR 72205, USA
| | - Stacie Bratton
- Department of Biochemistry and Molecular Biology, University of Arkansas for Medical Sciences, Little Rock, AR 72205, USA
| | - Anna Gallus-Zawada
- Department of Biochemistry and Molecular Biology, University of Arkansas for Medical Sciences, Little Rock, AR 72205, USA
| | - Moshe Finel
- DDTC, Faculty of Pharmacy, University of Helsinki, Helsinki, Finland
| | - Anna Radominska-Pandya
- Department of Biochemistry and Molecular Biology, University of Arkansas for Medical Sciences, Little Rock, AR 72205, USA
- Corresponding author: Anna Radominska-Pandya, Ph.D. Department of Biochemistry and Molecular Biology University of Arkansas for Medical Sciences 4301 W. Markham, Slot 516 Little Rock, AR 72205 Tel: (501) 686-5414 Fax: (501) 603-1146
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48
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Abstract
Sulfotransferases (Sults) are phase-II conjugation enzymes that catalyze the transfer of a sulfonate group from 3'-phosphoadenosine 5'-phosphosulfate (PAPS) to target endo and xenobiotics. PAPS is formed from inorganic sulfate by the action of the enzyme PAPS synthase (PAPSs). In the present study, the tissue distribution and developmental changes in the mRNA expression of 11 Sult isozymes and 2 PAPSs isoforms in mice were quantified. Sult1a1, 1b1, 1c1, 1c2, 1d1, 1e1, 2a1/2, 2b1, 3a1, 4a1, 5a1, PAPSs1, and PAPSs2 mRNA expression was quantified in 14 tissues from male and female mice using the branched DNA signal amplification assay. Sult2a1/2 and 3a1 expression were highest in liver; Sult1b1, 2b1, and PAPSs2 in small intestine; Sult1a1 in large intestine; Sult1c2 in stomach; Sult1d1 in kidney; Sult1e1 in placenta; and Sult4a1 in brain. Sult1c1, 5a1, and PAPSs1 were ubiquitously expressed in most tissues. These enzymes demonstrated three different ontogenic expression patterns in liver. Sult1a1, 1c2, 1d1, 2a1/2, and PAPSs2 hepatic expression gradually increased from birth until about 3 weeks of age and then declined somewhat thereafter, Sult1c1 expression was highest before birth and declined after that, and Sult3a1 mRNA expression was very low in fetal livers and remained low until 30 days of age, when expression in females dramatically increased, whereas it never increased in males. The organ-specific distribution of Sults as well as the different expression of the Sults in young animals may affect the pharmacokinetic behavior and organ-specific toxicity of xenobiotics.
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Affiliation(s)
- Yazen Alnouti
- Department of Pharmacology, Toxicology and Therapeutics, University of Kansas Medical Center, Kansas City, Kansas 66160, USA
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49
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Martina V, Benso A, Gigliardi VR, Masha A, Origlia C, Granata R, Ghigo E. Short-term dehydroepiandrosterone treatment increases platelet cGMP production in elderly male subjects. Clin Endocrinol (Oxf) 2006; 64:260-4. [PMID: 16487434 DOI: 10.1111/j.1365-2265.2006.02454.x] [Citation(s) in RCA: 14] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 01/06/2023]
Abstract
OBJECTIVE Several clinical and population-based studies suggest that dehydroepiandrosterone (DHEA) and its sulphate (DHEA-S) play a protective role against atherosclerosis and coronary artery disease in human. However, the mechanisms underlying this action are still unknown. It has recently been suggested that DHEA-S could delay atheroma formation through an increase in nitric oxide (NO) production. STUDY DESIGN AND METHODS Twenty-four aged male subjects [age (mean +/- SEM): 65.4 +/- 0.7 year; range: 58.2-67.6 years] underwent a blinded placebo controlled study receiving DHEA (50 mg p.o. daily at bedtime) or placebo for 2 months. Platelet cyclic guanosine-monophosphate (cGMP) concentration (as marker of NO production) and serum levels of DHEA-S, DHEA, IGF-I, insulin, glucose, oestradiol (E(2)), testosterone, plasminogen activator inhibitor (PAI)-1 antigen (PAI-1 Ag), homocysteine and lipid profile were evaluated before and after the 2-month treatment with DHEA or placebo. RESULTS At the baseline, all variables in the two groups were overlapping. All parameters were unchanged after treatment with placebo. Conversely, treatment with DHEA (a) increased (P < 0.001 vs. baseline) platelet cGMP (111.9 +/- 7.1 vs. 50.1 +/- 4.1 fmol/10(6) plts), DHEA-S (13.6 +/- 0.8 vs. 3.0 +/- 0.3 micromol/l), DHEA (23.6 +/- 1.7 vs. 15.3 +/- 1.4 nmol/l), testosterone (23.6 +/- 1.0 vs. 17.7 +/- 1.0 nmol/l) and E(2) (72.0 +/- 5.0 vs. 60.0 +/- 4.0 pmol/l); and (b) decreased (P < 0.05 vs. baseline) PAI-1 Ag (27.4 +/- 3.8 vs. 21.5 +/- 2.5 ng/ml) and low-density lipoprotein (LDL) cholesterol (3.4 +/- 0.2 vs. 3.0 +/- 0.2 mmol/l). IGF-I, insulin, glucose, triglycerides, total cholesterol, HDL cholesterol, HDL2 cholesterol, HDL3 cholesterol, apolipoprotein A1 (ApoA1), apolipoprotein B (ApoB) and homocysteine levels were not modified by DHEA treatment. CONCLUSIONS This study shows that short-term treatment with DHEA increased platelet cGMP production, a marker of NO production, in healthy elderly subjects. This effect is coupled with a decrease in PAI-1 and LDL cholesterol levels as well as an increase in testosterone and E(2) levels. These findings, therefore, suggest that chronic DHEA supplementation would exert antiatherogenic effects, particularly in elderly subjects who display low circulating levels of this hormone.
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Affiliation(s)
- Valentino Martina
- Division of Endocrinology and Metabolism, Department of Internal Medicine, University of Turin, Italy.
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Miyauchi S, Srinivas SR, Fei YJ, Gopal E, Umapathy NS, Wang H, Conway SJ, Ganapathy V, Prasad PD. Functional characteristics of NaS2, a placenta-specific Na+-coupled transporter for sulfate and oxyanions of the micronutrients selenium and chromium. Placenta 2005; 27:550-9. [PMID: 16129486 DOI: 10.1016/j.placenta.2005.05.004] [Citation(s) in RCA: 20] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 01/25/2005] [Revised: 05/06/2005] [Accepted: 05/10/2005] [Indexed: 11/29/2022]
Abstract
NaS2 is a Na+-coupled transporter for sulfate that belongs to the SLC13 gene family. This transporter was originally cloned from high endothelial venule endothelial cells, but nothing is known about the functional characteristics of this transporter except that it transports sulfate in a Na+-coupled manner. Northern blot analysis indicates that NaS2 is expressed most robustly in placenta. In the present study, we cloned NaS2 from rat placenta and characterized its transport function in detail using the Xenopus laevis oocyte expression system. Rat NaS2 consists of 629 amino acids and is highly similar to human NaS2. In situ hybridization studies with mouse placental sections show that NaS2 transcripts are expressed primarily in trophoblasts of the labyrinth zone. The expression of the transporter is confirmed in primary cultures of trophoblasts isolated from human placenta. When expressed in X. laevis oocytes, rat NaS2 mediates Na+-coupled transport of sulfate. The transport of sulfate is inhibited by oxyanions of selenium, chromium, arsenic, molybdenum, and phosphorous, suggesting that the transporter may mediate the transport of these oxyanions in addition to sulfate. The Kt for sulfate is 153+/-30 microM and the Na+:sulfate stoichiometry is 3:1. The transport process is electrogenic as evidenced from the inhibition of the uptake process by K+-induced depolarization. We conclude that NaS2 is a placenta-specific Na+-coupled, electrogenic, transporter for sulfate expressed in trophoblasts and that it is also responsible for the transport of oxyanions of the micronutrients selenium and chromium.
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Affiliation(s)
- S Miyauchi
- Department of Biochemistry and Molecular Biology, Medical College of Georgia, Augusta, GA 30912, USA
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