DOCSLIB.ORG

TECHNISCHE UNIVERSITÄT MÜNCHEN Lehrstuhl Für

Total Page：16

File Type：pdf, Size：1020Kb

Download full-text PDF Read full-text
TECHNISCHE UNIVERSITÄT MÜNCHEN Lehrstuhl Für TECHNISCHE UNIVERSITÄT MÜNCHEN Lehrstuhl für Physiologie Differential gene expression during pre-implantation pregnancy in bos taurus Andréa Hammerle-Fickinger Vollständiger Abdruck der Fakultät Wissenschaftszentrum Weihenstephan für Ernährung, Landnutzung und Umwelt der Technischen Universität München zur Erlangung des akademischen Grades eines Doktors der Naturwissenschaften genehmigte Dissertation. Vorsitzender: Univ.-Prof. Dr. U. M. Kulozik Prüfer der Dissertation: 1. Univ.-Prof. Dr. H. H. D. Meyer (schriftliche Beurteilung) 2. Priv.-Doz. Dr. K. Kramer 3. Univ.-Prof. Dr. M. W. Pfaffl Die Dissertation wurde am 20.02.2012 bei der Technischen Universität München eingereicht und durch die Fakultät Wissenschaftszentrum Weihenstephan für Ernährung, Landnutzung und Umwelt am 22.05.2012 angenommen. Table of Contents Table of Contents List of abbreviations .......................................................................................................... iv Zusammenfassung............................................................................................................. vi Abstract............................................................................................................................. viii 1 Introduction .................................................................................................................. 1 1.1 Bovine estrous cycle .................................................................................................. 1 1.2 Activation of the complement system during pregnancy............................................. 2 1.3 Early pregnancy in cattle and signaling responses..................................................... 4 1.4 Diapedesis: leukocytes migration from blood into milk................................................ 7 1.5 Gene expression analysis as a potential method for pregnancy diagnosis in cattle.... 8 1.6 Importance of sample processing for gene expression analysis ................................. 9 1.7 Aim of the thesis....................................................................................................... 11 2 Material and methods................................................................................................. 13 2.1 Study design of pregnancy recognition regarding the complement system activation in endometrium ............................................................................................................ 13 2.1.1 Monozygotic twin animals study ..................................................................... 13 2.1.2 Cyclic animals study....................................................................................... 13 2.1.3 In vitro co-culture of glandular epithelial and stroma cells study ..................... 14 2.1.4 Tissue sampling and RNA extraction.............................................................. 14 2.2 Study design of pregnancy recognition in peripheral blood leukocytes ..................... 14 2.2.1 Validation of blood sampling techniques......................................................... 14 2.2.2 Blood leukocytes study................................................................................... 16 2.3 Study design of pregnancy recognition in milk somatic cells..................................... 18 2.3.1 Validation of milk sampling techniques........................................................... 18 2.3.2 Milk somatic cells study.................................................................................. 19 i Table of Contents 2.4 RNA concentration and quality determination........................................................... 20 2.5 Target gene selection and primer design.................................................................. 20 2.6 Quantitative RT-PCR................................................................................................ 23 2.6.1 Gene expression analysis in endometrium ..................................................... 23 2.6.2 Gene expression analysis for validation of blood sampling............................. 23 2.6.3 Gene expression analysis on blood leukocytes and on milk somatic cells ...... 24 2.6.4 Statistical analysis of gene expression data ................................................... 25 2.7 Microarray analysis .................................................................................................. 26 2.7.1 Gene expression analysis in endometrium ..................................................... 26 2.7.2 Gene expression analysis on blood leukocytes .............................................. 26 2.8 Determination of C1Q concentration by ELISA......................................................... 27 3 Results ........................................................................................................................ 28 3.1 Analysis of pregnancy recognition regarding the complement system activation in endometrium ............................................................................................................ 28 3.1.1 Expression of complement system factors by RT-qPCR................................. 28 3.1.2 Expression of complement system factors by microarray analysis ................. 29 3.1.3 Expression of complement system factors in an in vitro co-culture system..... 29 3.2 Analysis of pregnancy recognition in peripheral blood leukocytes ............................ 31 3.2.1 Validation of blood sampling techniques......................................................... 31 3.2.2 Identification of a gene expression signature in bovine blood leukocytes ....... 37 3.3 Analysis of pregnancy recognition in milk somatic cells............................................ 45 3.3.1 Validation of milk sampling techniques........................................................... 45 3.3.2 Identification of a gene expression signature in bovine milk somatic cells ...... 50 4 Discussion .................................................................................................................. 55 4.1 Analysis of pregnancy recognition regarding the complement system activation in endometrium ............................................................................................................ 55 ii Table of Contents 4.2 Analysis of pregnancy recognition in peripheral blood leukocytes ............................ 58 4.3 Analysis of pregnancy recognition in milk somatic cells............................................ 64 5 Conclusions and perspectives.................................................................................. 69 6 References.................................................................................................................. 73 Acknowledgements........................................................................................................... 84 Scientific communication ................................................................................................. 85 Curriculum vitae ................................................................................................................ 87 Appendix............................................................................................................................ 88 iii Abbreviations List of abbreviations 18s rRNA 18S ribosomal RNA CFI complement factor I 28S rRNA 28S ribosomal RNA CK8 cytokeratin 8 ACTB actin beta CL corpus luteum AI artificial insemination Cq quantitative cycle bp base pair CR1 complement component BST2 bone marrow stromal cell receptor 1 antigen 2 CSF1 colony stimulating factor 1 C1Q complement component 1, D day of the cycle q subcomponent DF degrees of freedom C1R complement component 1, r EDTA ethylenediaminetetraacetic subcomponent acid C1S complement component 1, s F F-statistic test subcomponent FDR false discovery rate C2 complement component 2 GAPDH glyceraldehyde 3 phosphate C3 complement component 3 dehydrogenase C3AR complement component 3a GE glandular epithelial cells receptor HIS3 histone H3 C3bBb C3 convertase iC3b inactive C3b C3bBb3b C5 convertase IDO indoleamine 2,3- C4 complement component 4 dioxygenase C5 complement component 5 IFI16 interferon-induced protein C5AR complement component 5a 16 receptor IFI44 interferon-induced protein C6 complement component 6 44 C7 complement component 7 IFIT2 interferon-induced protein C8 complement component 8 with tetratricopeptide C9 complement component 9 repeats 2 CD46 membran co factor protein IFNT interferon-tau CD55 decay accelerating factor for IgG immunoglobulin G complement IgM immunoglobulin M CEL cell intensity files IL18 interleukin 18 CFB complement factor B IL1B interleukin 1 beta CFD complement factor D ISG interferon stimulated genes CFH complement factor H ISG15 ubiquitin-like modifier iv Abbreviations LH luteinizing hormone RMA robust multi-array analysis LIF leukemia inhibitory factor RNA ribonucleic acid LL leukoLOCK RNA extraction Rpm revolutions per minute method RT-qPCR quantitative reverse LY lysis RNA extraction transcription- polymerase method chain reaction MAC membrane attack complex SAM significance analysis of miRNA microRNA microarrays mRNA messenger ribonucleic acid SC stromal cells MS mean square SCC somatic cell count MX1 myxovirus resistance 1 SEM standard error of mean MX2 myxovirus resistance 2 SERPING1 serpin peptidase inhibitor, NP non-pregnant animals clade G (C1 inhibitor), nt nucleotide member 1 OAS1 2',5'-oligoadenylate SS sum of squares synthetase 1 STAT1 signal transducer and OOSP1 oocyte-secreted protein
Load more

Recommended publications
  • Regulation of Lymphocyte Proliferation by Uterine Serpin: Interleukin-2 Mrna Production, CD25 Expression and Responsiveness to Interleukin-2 (44465) 2 1 MORGAN R
    Regulation of Lymphocyte Proliferation by Uterine Serpin: Interleukin-2 mRNA Production, CD25 Expression and Responsiveness to Interleukin-2 (44465) 2 1 MORGAN R. PELTIER,WEN-JUN LIU, AND PETER J. HANSEN Department of Dairy and Poultry Sciences, University of Florida, Gainesville, Florida 32611–0920 Abstract. During pregnancy, the endometrium of the ewe secretes large amounts of a progesterone-induced protein of the serpin superfamily of serine proteinase inhibitors called ovine uterine serpin (OvUS). This protein inhibits lymphocyte proliferation in response to concanavalin A (ConA), phytohemagglutinin (PHA), or mixed lymphocyte reaction. The purpose of these experiments was to characterize the mechanism by which OvUS inhibits lymphocyte proliferation. Ovine US caused dose-dependent in- hibition of lymphocyte proliferation induced by phorbol myristol acetate (PMA), an activator of protein kinase C. The PHA-induced increase in CD25 expression was inhibited in peripheral blood mononuclear leukocytes (PBML) by OvUS. However, no effect of OvUS on Con A-induced expression of CD25 was observed. Further analysis using two-color flow cytometry revealed that OvUS inhibited ConA-induced expres- sion of CD25 in ␥␦-TCR− cells but not ␥␦-TCR+ cells. Stimulation of PBML for 14 hr with ConA resulted in an increase in steady state amounts of interleukin-2 (IL-2) mRNA that was not inhibited by OvUS. Ovine US was also inhibitory to lymphocyte proliferation induced by human IL-2. Results suggest that OvUS acts to inhibit lymphocyte prolif- eration by blocking the upregulation of the IL-2 receptor and inhibiting IL-2–mediated events. Lack of an effect of OvUS on ConA-stimulated CD25 expression in ␥␦-TCR+ cells may reflect a different mechanism of activation of these cells or insensitivity to inhibition by OvUS.
    [Show full text]
  • A Dissertation Presented to the Graduate School of the University of Florida in Partial Fulfillment of the Requirements for the Degree of Doctor of Philosophy
    MATERNAL IMMUNE CHANGES DURING BOVINE PREGNANCY: A FOCUS ON THE ENDOMETRIAL MACROPHAGE By LILIAN DE JESUS OLIVEIRA A DISSERTATION PRESENTED TO THE GRADUATE SCHOOL OF THE UNIVERSITY OF FLORIDA IN PARTIAL FULFILLMENT OF THE REQUIREMENTS FOR THE DEGREE OF DOCTOR OF PHILOSOPHY UNIVERSITY OF FLORIDA 2010 1 © 2010 Lilian de Jesus Oliveira 2 To my parents Luiz Manoel and Maria Aparecida de Jesus, my brother and sister, Rodolfo Manoel and Luciana de Jesus Oliveira, my boyfriend Robson Fortes Giglio, and my major advisor, Dr. Peter J. Hansen 3 ACKNOWLEDGMENTS I would like to express my heartfelt thanks in having Dr. Peter J. Hansen as my major advisor. His guidance, support and challenges throughout my PhD program have made me grow not only as scientist but also as a person. His enthusiasm, knowledge and intelligence inspired me every day during these years. He is more than an advisor; he is a very good friend that I am glad to have the opportunity to meet. I am going to miss our weekly meetings. Also, I would like to thank my committee members, Dr. Nasser Chegini, Dr. William Thatcher, Dr. Daniel Sharp and Dr. Ammon Peck, for their contributions and suggestions for improving my research projects and academic training. I am also grateful to Dr. Joel Yelich his continuous support and the opportunity to teach in his class. This experience was indispensable to my formation as a teacher; his passion for teaching is contagious. I would like to thank my current lab mates, Dr. Silvia Carambula, Sarah Fields, Barbara Loureiro, Luciano and Aline Bonilla, Justin Fear, and Jim Moss, as well as my old lab mates, Dr.
    [Show full text]
  • Biological Role of Conceptus Derived Factors During Early Pregnancy In
    Biological Role of Conceptus Derived Factors During Early Pregnancy in Ruminants A dissertation submitted in partial fulfillment of the requirements for the degree of DOCTOR OF PHILOSOPHY IN ANIMAL SCIENCES UNIVERSITY OF MISSOURI- COLUMBIA Division of Animal Science By KELSEY BROOKS Dr. Thomas Spencer, Dissertation Supervisor August 2016 The undersigned have examined the dissertation entitled, BIOLOGICAL ROLE OF CONCEPTUS DERIVED FACTORS DURING EARLY PREGNANCY IN RUMINANTS presented by Kelsey Brooks, a candidate for the degree of doctor of philosophy, and hereby certify that, in their opinion, it is worthy of acceptance. __________________________________ Chair, Dr. Thomas Spencer ___________________________________ Dr. Rodney Geisert ___________________________________ Dr. Randall Prather ___________________________________ Dr. Laura Schulz ACKNOWLEDGMENTS I would like to acknowledge all the students, faculty and staff at Washington State University and the University of Missouri for their help and support throughout my doctoral program. I am grateful for the opportunity to work with Dr. Thomas Spencer, and thank him for his input and guidance not only in planning experiments and completing projects but for helping me turn my love of science into a career in research. I would also like to acknowledge the members of my graduate committee at Washington State University for their help and input during the first 3 years of my studies. A special thanks to Dr. Jim Pru and Cindy Pru for providing unlimited entertainment, and the occasional missing reagent. Thank you to my committee members at the University of Missouri for adopting me late in my program and helping shape my future as an independent scientist. Thanks are also extended to members of the Prather lab and Wells lab for letting me in on the secrets of success using the CRISPR/Cas9 system.
    [Show full text]
  • Spatio-Specific Regulation of Endocrine-Responsive Gene Transcription by Periovulatory Endocrine Profiles in the Bovine Reproductive Tract
    CSIRO PUBLISHING Reproduction, Fertility and Development, 2016, 28, 1533–1544 http://dx.doi.org/10.1071/RD14178 Spatio-specific regulation of endocrine-responsive gene transcription by periovulatory endocrine profiles in the bovine reproductive tract Estela R. Arau´joA, Mariana SponchiadoA, Guilherme PugliesiA, Veerle Van HoeckA, Fernando S. MesquitaB, Claudia M. B. MembriveCand Mario BinelliA,D ADepartment of Animal Reproduction, School of Veterinary Medicine and Animal Science, University of Sa˜o Paulo, Avenida Duque de Caxias Norte, 225, Pirassununga, SP, 13635-900, Brazil. BSchool of Veterinary Medicine, Federal University of Pampa, Rodovia BR 472, 592, Uruguaiana, RS, 97508-000, Brazil. CCollege of Animal Science, University of Sa˜o Paulo State ‘Ju´lio de Mesquita Filho’, DracenaRodovia Comandante Joa˜o Ribeiro de Barros, km 651, Dracena, SP, 17900-000, Brazil. DCorresponding author. Email: [email protected] Abstract. In cattle, pro-oestrous oestradiol and dioestrous progesterone concentrations modulate endometrial gene expression and fertility. The aim was to compare the effects of different periovulatory endocrine profiles on the expression of progesterone receptor (PGR), oestrogen receptor 2 (ESR2), oxytocin receptor (OXTR), member C4 of aldo–keto reductase family 1 (AKR1C4), lipoprotein lipase (LPL), solute carrier family 2, member 1 (SLC2A1) and serpin peptidase inhibitor, clade A member 14 (SERPINA14): (1) between uterine horns ipsi- and contralateral to the corpus luteum (CL), (2) between regions of the ipsilateral horn and (3) in the vagina. Endometrium and vagina tissue samples were collected from cows that ovulated a larger (large follicle-large CL, LF-LCL; n ¼ 6) or smaller follicle (small follicle-small CL, SF-SCL; n ¼ 6) 7 days after oestrus.
    [Show full text]
  • Molecular and Cellular Studies of Drosophila Neuroserpin Spn4a And
    Molecular and Cellular Studies of Drosophila Neuroserpin Spn4A and its Polymer-Forming Mutants by Christine Chieh-Lin Lai B.Sc. University of British Columbia, 2008 A THESIS SUBMITTED IN PARTIAL FULFILLMENT OF THE REQUIREMENTS FOR THE DEGREE OF MASTER OF SCIENCE in THE FACULTY OF GRADUATE STUDIES (Microbiology & Immunology) The University of British Columbia (Vancouver) August 2010 © Christine Chieh-Lin Lai, 2010 1 Abstract Serpins (Serine Protease Inhibitors) are expressed by most organisms and perform a variety of functions. Most serpins inhibit proteases by undergoing a unique conformational change. They are clinically relevant in two ways. First, introduction of single amino acid point mutations transforms the serpins’ labile conformations into pathogenic, inactive polymers causing “serpinopathies”. In particular, human neuroserpin is a brain-specific serpin that, when mutated, causes a debilitating early onset dementia through unknown cellular pathways. Second, serpins are currently under investigation as therapeutic inhibitors of proprotein convertases (PCs). PCs are associated with some bacterial and viral infections as well as cancer. However, no comprehensive investigation into the cellular effects of PC inhibitor expression in mammalian cells has been performed. This thesis details the use of the Drosophila serpin, Spn4A, to address the cellular pathways mediated by serpin polymers or PC inhibition. Spn4A is a neuron-specific, secretory pathway serpin that inhibits Drosophila or human PCs. We hypothesized that Spn4A mutants, encoding homologous disease-causing mutations in human neuroserpin, would form pathogenic polymers and represent an ideal candidate for generating a cell-based and transgenic Drosophila serpinopathy model. Further, we hypothesized that we could evaluate the cellular response to PC inhibition and polymer accumulation by transcriptome profiling of H4 human neuroglioma cells expressing Spn4A wild-type and mutants.
    [Show full text]
  • Pepsin-Inhibitory Activity of the Uterine Serpins
    Proc. Natl. Acad. Sci. USA Vol. 93, pp. 13653–13658, November 1996 Biochemistry Pepsin-inhibitory activity of the uterine serpins (uterine secretory activityyaspartic proteinase inhibitoryprogesterone-induced uterine proteinyendometrium–trophoblast interaction) NAGAPPAN MATHIALAGAN*† AND THOMAS R. HANSEN‡ *Department of Animal Sciences, University of Missouri, Columbia, MO 65211; and ‡Department of Animal Sciences, University of Wyoming, Laramie, WY 82071 Communicated by Michael Roberts, University of Missouri, Columbia, MO, September 19, 1996 (received for review June 20, 1996) ABSTRACT Among the major products secreted by the distinct (20). Therefore, it was of considerable interest that uteri of cattle, sheep, and pigs during pregnancy are glyco- both species should produce large quantities of structurally proteins with amino acid sequences that place them in the similar progesterone-inducible products during pregnancy. serpin (serine proteinase inhibitor) superfamily of proteins. Hence the studies on uterine serpins have been extended. The inferred amino acid sequences for bovine uterine serpin Herein we demonstrate that these uterine serpins interact with (boUS-1) and ovine uterine serpin (ovUS-1) exhibit about 72% members of the aspartic proteinase family rather than with sequence identity to each other but only about 50% and 56% serine proteinases. They provide another example of serpins identity, respectively, to two distinct porcine uterine serpins with crossover function. (poUS-1 and poUS-2). Despite these differences in primary Because various acronyms were used for these uterine structure, the uterine serpins possess well-conserved reactive serpins before their general relatedness was revealed by mo- center loop regions that contain several motifs present in the lecular cloning studies, it is proposed that the previous desig- propeptide regions of pepsinogens.
    [Show full text]
  • Evolution and Function of the Uterine Serpins (SERPINA14) Maria B
    REVIEW ARTICLE Evolution and Function of the Uterine Serpins (SERPINA14) Maria B. Padua1, Peter J. Hansen2 1Department of Obstetrics and Gynecology, College of Medicine, University of Florida, Gainesville, FL, USA; 2Department of Animal Sciences, University of Florida, Gainesville, FL, USA Keywords Uterine serpins (recently designated as SERPINA14) are hormonally Immunoregulation, pregnancy, progesterone, induced proteins secreted in large quantities by the endometrial epithe- uterine milk proteins, uterine serpin lium during pregnancy. The SERPINA14 proteins belong to the serine proteinase inhibitor (serpin) superfamily, but their apparent lack of Correspondence Dr Maria B. Padua, PO Box 100294, inhibitory activity toward serine proteinases suggests that these proteins Department of Obstetrics and Gynecology, evolved a different function from the anti-proteinase activity typically University of Florida, Gainesville, found in most members of the serpin superfamily. The gene is present FL 32610-0294, USA. in a limited group of mammals in the Laurasiatheria superorder (rumi- E-mail: mpadua@ufl.edu nants, horses, pigs, dolphins and some carnivores) while being absent in primates, rodents, lagomorphs and marsupials. Thus, the gene is likely Submitted June 7, 2010; to have evolved by gene duplication after divergence of Laurasiatheria accepted June 8, 2010. and to play an important role in pregnancy. That role may vary between Citation species. In sheep, SERPINA14 probably serves an immunoregulatory role Padua MB, Hansen PJ. Evolution and function to prevent rejection of the fetal allograft. It is inhibitory to lymphocyte of the uterine serpins (SERPINA14). Am J proliferation and natural killer cell function. In the pig, SERPINA14 is Reprod Immunol 2010; 64: 265–274 involved in iron transport to the fetus by binding to and stabilizing the iron-binding protein uteroferrin.
    [Show full text]
  • Corticosteroid-Binding Globulin (Cbg): Deficiencies and the Role
    CORTICOSTEROID-BINDING GLOBULIN (CBG): DEFICIENCIES AND THE ROLE OF CBG IN DISEASE PROCESSES by LESLEY ANN HILL B.Sc. University of Western Ontario, 2011 A DISSERTATION SUBMITTED IN PARTIAL FULFILLMENT OF THE REQUIREMENT FOR THE DEGREE OF DOCTOR OF PHILOSOPHY in THE FACULTY OF GRADUATE AND POSTDOCTORAL STUDIES (Reproductive and Developmental Sciences) THE UNIVERSITY OF BRITISH COLUMBIA (Vancouver) July 2017 © Lesley Ann Hill, 2017 Abstract Corticosteroid-binding globulin (CBG, SERPINA6) is a serine protease inhibitor family member produced by hepatocytes. Plasma CBG transports biologically active glucocorticoids, determines their bioavailability to target tissues and acts as an acute-phase negative protein with a role in the delivery of glucocorticoids to sites of inflammation. A few CBG-deficient individuals have been identified, yet the clinical significance of this remain unclear. In this thesis, I investigated 1) the biochemical consequences of naturally occurring single nucleotide polymorphisms in the SERPINA6 gene, 2) the role of human CBG during infections and acute inflammation and 3) CBG as a biomarker of inflammation in rats. A comprehensive analysis of functionally relevant naturally occurring SERPINA6 SNP revealed 11 CBG variants with abnormal production and/or function, diminished responses to proteolytic cleavage of the CBG reactive center loop (RCL) or altered recognition by monoclonal antibodies. In a genome-wide association study, plasma cortisol levels were most closely associated with SERPINA6 SNPs and plasma CBG-cortisol binding capacity. These studies indicate that human CBG variants need to be considered in clinical evaluations of patients with abnormal cortisol levels. In addition, I obtained evidence that discrepancies in CBG values obtained by the 9G12 ELISA compared to CBG binding capacity and 12G2 ELISA are likely due to differential N-glycosylation rather than proteolysis, as recently reported.
    [Show full text]
  • Understanding the Influence of Synchronization of Ovulation On
    University of Tennessee, Knoxville TRACE: Tennessee Research and Creative Exchange Doctoral Dissertations Graduate School 8-2018 Understanding the influence of synchronization of ovulation on steroid bioavailability and its association with the presence of Pseudomonas aeruginosa within the bovine reproductive tract Sierra Ashley Lockwood University of Tennessee, [email protected] Follow this and additional works at: https://trace.tennessee.edu/utk_graddiss Recommended Citation Lockwood, Sierra Ashley, "Understanding the influence of synchronization of ovulation on steroid bioavailability and its association with the presence of Pseudomonas aeruginosa within the bovine reproductive tract. " PhD diss., University of Tennessee, 2018. https://trace.tennessee.edu/utk_graddiss/5060 This Dissertation is brought to you for free and open access by the Graduate School at TRACE: Tennessee Research and Creative Exchange. It has been accepted for inclusion in Doctoral Dissertations by an authorized administrator of TRACE: Tennessee Research and Creative Exchange. For more information, please contact [email protected]. To the Graduate Council: I am submitting herewith a dissertation written by Sierra Ashley Lockwood entitled "Understanding the influence of synchronization of ovulation on steroid bioavailability and its association with the presence of Pseudomonas aeruginosa within the bovine reproductive tract." I have examined the final electronic copy of this dissertation for form and content and recommend that it be accepted in partial fulfillment of the equirr ements for the degree of Doctor of Philosophy, with a major in Animal Science. Henry G. Kattesh, Major Professor We have read this dissertation and recommend its acceptance: Peter D. Krawczel, Ky G. Pohler, Justin D. Rhinehart, Brynn H. Voy, Brian K.
    [Show full text]
  • Antiproliferative Actions of Ovine Uterine Serpin
    Copyright Ó Blackwell Munksgaard, 2005 AJRI 2005; 53: 136–143 American Journal of Reproductive Immunology Antiproliferative Actions of Ovine Uterine Serpin Tekin S¸, Padua MB, Brad AM, Hansen PJ. Antiproliferative actions S¸aban Tekin1,2, Maria B. Padua1, of ovine uterine serpin. AJRI 2005; 53:136–143 Ó Blackwell Amber M. Brad1, Peter J. Hansen1 Munksgaard, 2005 1Department of Animal Sciences, University of Florida, Gainesville, FL, USA; 2Department of Biology, University PROBLEM: Ovine uterine serpin (OvUS) is a member of the serine of Gaziosmanpas¸a, Tokat, Turkey proteinase inhibitor superfamily and is the major protein produced by luminal and glandular epithelium of the sheep endometrium during mid to late pregnancy. The protein does not have prototypical proteinase inhibitory activity but can inhibit a wide variety of lymphocyte functions such as mitogen-induced proliferation and natural killer cell cytotoxicity. METHOD OF STUDY: The antiproliferative actions of OvUS were studied. RESULTS: It was demonstrated that, in addition to inhibiting lymphocyte proliferation, OvUS inhibits growth of two tumor cell lines (D17 and PC-3). The protein also interrupts development of pre- implantation embryos. Inhibition of cell proliferation is not universal, however, as OvUS did not inhibit growth of two non-tumorigenic cell lines (MDBK and BEND). The mechanism of action of inhibitory Key words: Lymphocytes, pre-implantation embryos, effects of OvUS is not known although experiments with inhibitors of proliferation, uterine serpin protein kinase A indicate that the protein does not inhibit lymphocyte proliferation through this pathway. Moreover, the protein does not Address reprint requests to Peter J. Hansen, PO Box 110910, Department of Animal Sciences, University induce apoptosis.
    [Show full text]
  • Regulation of DNA Synthesis and the Cell Cycle in Human Prostate Cancer Cells and Lymphocytes by Ovine Uterine Serpin Maria B Padua and Peter J Hansen*
    BMC Cell Biology BioMed Central Research article Open Access Regulation of DNA synthesis and the cell cycle in human prostate cancer cells and lymphocytes by ovine uterine serpin Maria B Padua and Peter J Hansen* Address: Department of Animal Sciences, University of Florida, Gainesville, FL 32611-0910, USA Email: Maria B Padua - [email protected]; Peter J Hansen* - [email protected] * Corresponding author Published: 24 January 2008 Received: 17 August 2007 Accepted: 24 January 2008 BMC Cell Biology 2008, 9:5 doi:10.1186/1471-2121-9-5 This article is available from: http://www.biomedcentral.com/1471-2121/9/5 © 2008 Padua and Hansen; licensee BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. Abstract Background: Uterine serpins are members of the serine proteinase inhibitor superfamily. Like some other serpins, these proteins do not appear to be functional proteinase inhibitors. The most studied member of the group, ovine uterine serpin (OvUS), inhibits proliferation of several cell types including activated lymphocytes, bovine preimplantation embryos, and cell lines for lymphoma, canine primary osteosarcoma and human prostate cancer (PC-3) cells. The goal for the present study was to evaluate the mechanism by which OvUS inhibits cell proliferation. In particular, it was tested whether inhibition of DNA synthesis in PC-3 cells involves cytotoxic actions of OvUS or the induction of apoptosis. The effect of OvUS in the production of the autocrine and angiogenic cytokine interleukin (IL)-8 by PC-3 cells was also determined.
    [Show full text]
  • RESEARCH ARTICLE Intrinsic Expression of Transcortin in Neural Cells of the Mouse Brain: a Histochemical and Molecular Study
    245 The Journal of Experimental Biology 216, 245-252 © 2013. Published by The Company of Biologists Ltd doi:10.1242/jeb.076893 RESEARCH ARTICLE Intrinsic expression of transcortin in neural cells of the mouse brain: a histochemical and molecular study Elena Sivukhina1, Jean-Christophe Helbling2,3, Amandine M. Minni2,3, H. Hendrik Schäfer1, Véronique Pallet2,3, Gustav F. Jirikowski1 and Marie-Pierre Moisan2,3,* 1Institute of Anatomy II, Friedrich-Schiller University, Jena University Hospital, Jena, Germany, 2INRA, Nutrition and Integrative Neurobiology, UMR 1286, 33076 Bordeaux, France and 3Université de Bordeaux, Nutrition and Integrative Neurobiology, UMR 1286, 33076 Bordeaux, France *Address for correspondence ([email protected]) SUMMARY Corticosteroid binding globulin (CBG, transcortin) has been shown to be expressed in the brain of rat and human species. In this study, we examined the CBG brain expression and cDNA structure in mice, comparing wild-type (Cbg+/+) and Cbg knockout mice (Cbg–/–, obtained by genetic disruption of the SerpinA6 alias Cbg gene). We used double immunofluorescence labeling with specific neuronal and glial markers to analyze the cellular localization of CBG in various regions of the mouse brain. In wild-type (Cbg+/+) mice, we found CBG immunoreactivity in neuronal perikarya of the magnocellular hypothalamic nuclei, amygdala, hippocampus, cerebral cortex, cerebellum and pituitary. A portion of glial cells (astrocytes, oligodendrocytes) contained CBG immunoreactivity, including some of the ependymal cells and choroid plexus cells. No CBG immunoreactivity was detected in Cbg–/– brain tissues. Using RT-PCR, we showed that the full-length Cbg mRNA is present in those regions, indicating an intrinsic expression of the steroid-binding globulin.
    [Show full text]