DOCSLIB.ORG

Functions of Mmrn1 in Platelet Adhesion & Thrombus Formation

Total Page：16

File Type：pdf, Size：1020Kb

Download full-text PDF Read full-text
Functions of Mmrn1 in Platelet Adhesion & Thrombus Formation FUNCTIONS OF MMRN1 IN PLATELET ADHESION & THROMBUS FORMATION FUNCTIONS OF MULTIMERIN 1 (MMRN1) IN PLATELET ADHESION AND THROMBUS FORMATION, THROUGH INTERACTIONS WITH VON WILLEBRAND FACTOR (VWF) By: D’ANDRA PARKER, HBSc A Thesis Submitted to the School of Graduate Studies in Partial Fulfilment of the Requirements for the Degree Doctorate of Philosophy in Health Science McMaster University © Copyright by D’Andra Parker, April 2017 McMaster University DOCTORATE OF PHILOSOPHY (2017) Hamilton, Ontario (Health Science) TITLE: Functions of multimerin 1 (MMRN1) in platelet adhesion and thrombus formation, through interactions with von Willebrand factor (VWF) AUTHOR: D’Andra Parker, HBSc (McMaster University) SUPERVISOR: Dr. Catherine P.M. Hayward NUMBER OF PAGES: xiv, 209 ii ABSTRACT Multimerin 1 (MMRN1) is a massive, homopolymeric platelet and endothelial cell protein with functions that are emerging to support platelet adhesive processes. MMRN1 supports platelet adhesion under arterial flow conditions by a mechanism dependent on interactions with von Willebrand factor (VWF). The goals of this thesis were to further define the platelet adhesive functions of MMRN1 by: 1) characterizing the molecular mechanisms of VWF interactions with MMRN1; and 2) investigating if multimerin 1 is important for platelet adhesive functions using mice with and without a selective multimerin 1 (Mmrn1) deficiency. Studies of the mechanism of MMRN1-VWF binding indicated that MMRN1 bound to shear exposed VWF, and that MMRN1 interacted with the A1 and A3 domains in the VWF A1A2A3 region. VWF A1A2A3 also bound to MMRN1 with a physiologically relevant binding affinity, and supported platelet adhesion to MMRN1 at a high shear rate. The selective loss of Mmrn1 in mice had limited effects on tail bleeding times, although it impaired collagen-induced aggregation of washed platelets, as well as high shear platelet adhesion of whole blood on collagen surfaces, in vitro. Additionally, the selective loss of Mmrn1 in mice was associated with impaired and delayed platelet-rich thrombus formation in vivo, in arterioles treated with ferric chloride. These findings provide new insights on platelet adhesive, haemostatic functions at arterial shear rates, and the involvement of the platelet and endothelial cell protein, multimerin 1, to support these processes. iii ACKNOWLEDGMENTS I would like to express my gratitude to individuals that provided me with guidance and support throughout my doctoral degree. I would like to begin by thanking my PhD supervisor, Dr. Catherine P.M. Hayward for continued mentorship, guidance, support and encouragement throughout my graduate studies. Her drive and determination constantly inspired me. I am grateful to my committee members, Drs. Peter Gross and William Sheffield for their continued guidance, feedback, and support. I thank my colleagues in the research laboratories of Drs. Hayward, Kelton, Sheffield, Khan, and Gross who were instrumental in the completion of my doctoral project. A special thank you is owed to Dr. Subia Tasneem for mentorship and support; her openness and willingness to help facilitated my achievements. I would like to extend my gratitude to the collaborators that were involved in my studies. I thank Drs. J. Evan Sadler, Philip G. de Groot, Richard W. Farndale, Dominique Bihan, Eric G. Huizinga, David Lillicrap and Ms. Silvie Sebastian for providing important materials for my studies, and for their guidance with data interpretation. I am grateful to Dr. Ran Ni for providing technical guidance for flow cytometry analysis of mouse samples. A special thank you is owed to Drs. Yiming Wang and Heyu Ni for conducting ferric chloride injury studies on mice, and for providing me with guidance on data interpretation. iv I also thank Dr. Lisa A. Westfield and Mrs. Nola Fuller for providing general technical guidance. During my time at McMaster, I had the pleasure of forming lasting friendships with incredible individuals that helped bring balance to my life, and provided me with moments of comic relief. I would like to especially like to thank soon-to-be Drs. Janice Kim, Zainab Motala and Sharif Shajib for their encouragement and support throughout my graduate studies, and for the memories that I will cherish forever. Finally, I would like to thank my family, by beginning with the woman that brought me into this world and raised me into the woman I am today. A special thank you is owed to my mom for being my rock and safe place. Thank you for pushing me to strive for more; I am forever grateful for your encouragement and support throughout this entire process. I also thank my grandma for being my shield, safe place, and prayer warrior: she kept me focused and reminded me that everything is possible, if we have faith. I owe many thanks to my little brothers D’Vantae Stewart and Enrique Parker, little sister Alecia Beckford- Stewart, and my step-father Denroy Stewart. I also thank the members of my larger than life family, including the Lewis, Hall, Seaton, Mckenzie, Osei and Odamatten clan – I hope I can make you proud. Last, but definitely not least, I need to thank the love of my life, my life partner, Kwaku Osei. Thank you Kwaku for your patience, support, and encouragement. You believe in me like no other, and I am eternally grateful for that. v TABLE OF CONTENTS TITLE PAGE………………………………………………….……………………...………....…..i DESCRIPTIVE NOTE……………………………………………..…………………………....…ii ABSTRACT……………………………………………………………………………….………iii ACKNOWLEDGMENT………………………………………………..………………………....iv TABLE OF CONTENTS……………… …………………………………………..………........vi LIST OF FIGURES …………………………………………………………………….……........ix LIST OF TABLES…………………………… …………………………………….…...………...xi LIST OF ABBREVIATIONS AND SYMBOLS……………………………………………….....xii Chapter 1: Introduction .................................................................................................................... 1 1.1 Overview of haemostasis ................................................................................................. 2 1.1.1 Protein wave ............................................................................................................. 6 1.1.2 Platelet adhesive functions (primary haemostasis) .................................................. 7 1.1.3 Platelet procoagulant function (secondary haemostasis) and fibrinolysis .............. 18 1.2 Overview of haemostatic functions of plasma and platelet adhesive proteins .............. 20 1.2.1 Fibrinogen and fibrin ............................................................................................. 22 1.2.2 Fibronectin ............................................................................................................. 26 1.2.3 Thrombospondin-1 ................................................................................................. 28 1.3 Overview of von Willebrand factor (human, VWF; mouse, Vwf) ............................... 30 1.3.1 Background on von Willebrand factor ................................................................... 31 1.3.2 VWF protein structure-function relationship ......................................................... 34 1.3.3 Deficiencies in von Willebrand factor, in humans and mice, and its effect on haemostasis ............................................................................................................ 45 1.4 Overview of multimerin 1 (human, MMRN1; mouse, Mmrn1) .................................... 48 1.4.1 Background on multimerin 1 ................................................................................. 49 1.4.2 MMRN1, the main FV/Va binding protein in platelets, and its functions in coagulation ............................................................................................................. 54 1.4.3 MMRN1 functions in platelet adhesion ................................................................. 56 1.4.4 MMRN1 deficiencies in inherited and acquired disease states .............................. 59 1.4.5 Haemostatic functions of Mmrn1 in mice with spontaneous tandem deletion of genes for multimerin 1 and -synuclein ................................................................ 60 1.5 Thesis hypothesis and objectives ................................................................................... 63 vi 1.5.1. Rationale ................................................................................................................ 63 1.5.2 Hypotheses ............................................................................................................. 65 1.5.3 Research objectives ................................................................................................ 65 CHAPTER 2: MATERIALS AND METHODS............................................................................ 67 2.1 Materials ........................................................................................................................ 67 2.1.1 Reagents ................................................................................................................. 67 2.1.2 Antibody sources .................................................................................................... 68 2.1.3 Recombinant proteins ............................................................................................ 70 2.2. Methods .......................................................................................................................... 75 2.2.1 Helsinki Declaration on human research ..............................................................
Load more

Recommended publications
  • MMRN1 Antibody (Monoclonal) (M02) Mouse Monoclonal Antibody Raised Against a Partial Recombinant MMRN1
    10320 Camino Santa Fe, Suite G San Diego, CA 92121 Tel: 858.875.1900 Fax: 858.622.0609 MMRN1 Antibody (monoclonal) (M02) Mouse monoclonal antibody raised against a partial recombinant MMRN1. Catalog # AT2883a Specification MMRN1 Antibody (monoclonal) (M02) - Product Information Application WB, E Primary Accession Q13201 Other Accession NM_007351 Reactivity Human Host mouse Clonality Monoclonal Isotype IgG2b Kappa Calculated MW 138110 MMRN1 Antibody (monoclonal) (M02) - Additional Information Antibody Reactive Against Recombinant Protein.Western Blot detection against Gene ID 22915 Immunogen (36.74 KDa) . Other Names Multimerin-1, EMILIN-4, Elastin microfibril interface located protein 4, Elastin microfibril interfacer 4, Endothelial cell multimerin, Platelet glycoprotein Ia*, 155 kDa platelet multimerin, p-155, p155, MMRN1, ECM, EMILIN4, GPIA*, MMRN Target/Specificity MMRN1 (NP_031377, 291 a.a. ~ 390 a.a) partial recombinant protein with GST tag. MW of the GST tag alone is 26 KDa. Detection limit for recombinant GST tagged MMRN1 is approximately 0.1ng/ml as a Dilution capture antibody. WB~~1:500~1000 Format MMRN1 Antibody (monoclonal) (M02) - Clear, colorless solution in phosphate Background buffered saline, pH 7.2 . Multimerin is a massive, soluble protein found Storage in platelets and in the endothelium of blood Store at -20°C or lower. Aliquot to avoid vessels. It is comprised of subunits linked by repeated freezing and thawing. interchain disulfide bonds to form large, variably sized homomultimers. Multimerin is a Precautions factor V/Va-binding protein and may function MMRN1 Antibody (monoclonal) (M02) is for as a carrier protein for platelet factor V. It may research use only and not for use in also have functions as an extracellular matrix diagnostic or therapeutic procedures.
    [Show full text]
  • The EMILIN/Multimerin Family
    View metadata, citation and similar papers at core.ac.uk brought to you by CORE REVIEW ARTICLE published: 06 Januaryprovided 2012 by Frontiers - Publisher Connector doi: 10.3389/fimmu.2011.00093 The EMILIN/multimerin family Alfonso Colombatti 1,2,3*, Paola Spessotto1, Roberto Doliana1, Maurizio Mongiat 1, Giorgio Maria Bressan4 and Gennaro Esposito2,3 1 Experimental Oncology 2, Centro di Riferimento Oncologico, Istituto di Ricerca e Cura a Carattere Scientifico, Aviano, Italy 2 Department of Biomedical Science and Technology, University of Udine, Udine, Italy 3 Microgravity, Ageing, Training, Immobility Excellence Center, University of Udine, Udine, Italy 4 Department of Histology Microbiology and Medical Biotechnologies, University of Padova, Padova, Italy Edited by: Elastin microfibrillar interface proteins (EMILINs) and Multimerins (EMILIN1, EMILIN2, Uday Kishore, Brunel University, UK Multimerin1, and Multimerin2) constitute a four member family that in addition to the Reviewed by: shared C-terminus gC1q domain typical of the gC1q/TNF superfamily members contain a Uday Kishore, Brunel University, UK Kenneth Reid, Green Templeton N-terminus unique cysteine-rich EMI domain. These glycoproteins are homotrimeric and College University of Oxford, UK assemble into high molecular weight multimers. They are predominantly expressed in *Correspondence: the extracellular matrix and contribute to several cellular functions in part associated with Alfonso Colombatti, Division of the gC1q domain and in part not yet assigned nor linked to other specific regions of the Experimental Oncology 2, Centro di sequence. Among the latter is the control of arterial blood pressure, the inhibition of Bacil- Riferimento Oncologico, Istituto di Ricerca e Cura a Carattere Scientifico, lus anthracis cell cytotoxicity, the promotion of cell death, the proangiogenic function, and 33081 Aviano, Italy.
    [Show full text]
  • The Neuroprotective Role of the GM1 Oligosaccharide, Ii3neu5ac-Gg4, In
    Molecular Neurobiology (2019) 56:6673–6702 https://doi.org/10.1007/s12035-019-1556-8 The Neuroprotective Role of the GM1 Oligosaccharide, 3 II Neu5Ac-Gg4, in Neuroblastoma Cells Elena Chiricozzi1 & Margherita Maggioni1 & Erika di Biase1 & Giulia Lunghi1 & Maria Fazzari1 & Nicoletta Loberto 1 & Maffioli Elisa2 & Francesca Grassi Scalvini2 & Gabriella Tedeschi 2,3 & Sandro Sonnino1 Received: 10 January 2019 /Accepted: 13 March 2019 /Published online: 26 March 2019 # Springer Science+Business Media, LLC, part of Springer Nature 2019 Abstract 3 Recently, we demonstrated that the GM1 oligosaccharide, II Neu5Ac-Gg4 (OligoGM1), administered to cultured murine Neuro2a neuroblastoma cells interacts with the NGF receptor TrkA, leading to the activation of the ERK1/2 downstream pathway and to cell differentiation. To understand how the activation of the TrkA pathway is able to trigger key biochemical signaling, we performed a proteomic analysis on Neuro2a cells treated with 50 μM OligoGM1 for 24 h. Over 3000 proteins were identified. Among these, 324 proteins were exclusively expressed in OligoGM1-treated cells. Interestingly, several proteins expressed only in OligoGM1-treated cells are involved in biochemical mechanisms with a neuroprotective potential, reflecting the GM1 neuroprotective effect. In addition, we found that the exogenous administration of OligoGM1 reduced the cellular oxidative stress in Neuro2a cells and conferred protection against MPTP neurotoxicity. These results confirm and reinforce the idea that the molecular mechanisms underlying the GM1 neurotrophic and neuroprotective effects depend on its oligosaccharide chain, suggesting the activation of a positive signaling starting at plasma membrane level. Keywords GM1 ganglioside . GM1 oligosaccharide chain . TrkA neurotrophin receptor . Plasma membrane signaling . Neuroprotection .
    [Show full text]
  • Investigating the Effect of Chronic Activation of AMP-Activated Protein
    Investigating the effect of chronic activation of AMP-activated protein kinase in vivo Alice Pollard CASE Studentship Award A thesis submitted to Imperial College London for the degree of Doctor of Philosophy September 2017 Cellular Stress Group Medical Research Council London Institute of Medical Sciences Imperial College London 1 Declaration I declare that the work presented in this thesis is my own, and that where information has been derived from the published or unpublished work of others it has been acknowledged in the text and in the list of references. This work has not been submitted to any other university or institute of tertiary education in any form. Alice Pollard The copyright of this thesis rests with the author and is made available under a Creative Commons Attribution Non-Commercial No Derivatives license. Researchers are free to copy, distribute or transmit the thesis on the condition that they attribute it, that they do not use it for commercial purposes and that they do not alter, transform or build upon it. For any reuse or redistribution, researchers must make clear to others the license terms of this work. 2 Abstract The prevalence of obesity and associated diseases has increased significantly in the last decade, and is now a major public health concern. It is a significant risk factor for many diseases, including cardiovascular disease (CVD) and type 2 diabetes. Characterised by excess lipid accumulation in the white adipose tissue, which drives many associated pathologies, obesity is caused by chronic, whole-organism energy imbalance; when caloric intake exceeds energy expenditure. Whilst lifestyle changes remain the most effective treatment for obesity and the associated metabolic syndrome, incidence continues to rise, particularly amongst children, placing significant strain on healthcare systems, as well as financial burden.
    [Show full text]
  • The Structure, Function and Evolution of the Extracellular Matrix: a Systems-Level Analysis
    The Structure, Function and Evolution of the Extracellular Matrix: A Systems-Level Analysis by Graham L. Cromar A thesis submitted in conformity with the requirements for the degree of Doctor of Philosophy Department of Molecular Genetics University of Toronto © Copyright by Graham L. Cromar 2014 ii The Structure, Function and Evolution of the Extracellular Matrix: A Systems-Level Analysis Graham L. Cromar Doctor of Philosophy Department of Molecular Genetics University of Toronto 2014 Abstract The extracellular matrix (ECM) is a three-dimensional meshwork of proteins, proteoglycans and polysaccharides imparting structure and mechanical stability to tissues. ECM dysfunction has been implicated in a number of debilitating conditions including cancer, atherosclerosis, asthma, fibrosis and arthritis. Identifying the components that comprise the ECM and understanding how they are organised within the matrix is key to uncovering its role in health and disease. This study defines a rigorous protocol for the rapid categorization of proteins comprising a biological system. Beginning with over 2000 candidate extracellular proteins, 357 core ECM genes and 524 functionally related (non-ECM) genes are identified. A network of high quality protein-protein interactions constructed from these core genes reveals the ECM is organised into biologically relevant functional modules whose components exhibit a mosaic of expression and conservation patterns. This suggests module innovations were widespread and evolved in parallel to convey tissue specific functionality on otherwise broadly expressed modules. Phylogenetic profiles of ECM proteins highlight components restricted and/or expanded in metazoans, vertebrates and mammals, indicating taxon-specific tissue innovations. Modules enriched for medical subject headings illustrate the potential for systems based analyses to predict new functional and disease associations on the basis of network topology.
    [Show full text]
  • Differential Gene Expression by RNA-Seq Analysis of the Primo Vessel in the Rabbit Lymph
    J Acupunct Meridian Stud 2019;12(1):11e19 Available online at www.sciencedirect.com Journal of Acupuncture and Meridian Studies journal homepage: www.jams-kpi.com Research Article Differential Gene Expression by RNA-Seq Analysis of the Primo Vessel in the Rabbit Lymph Jun-Young Shin, Sang-Heon Choi, Da-Woon Choi, Ye-Jin An, Jae-Hyuk Seo, Jong-Gu Choi, Min-Suk Rho, Sang-Suk Lee* Department of Oriental Biomedical Engineering, Sangji University, Wonju, Republic of Korea Available online 28 October 2018 Received: May 31, 2018 Abstract Revised: Jul 26, 2018 For the connectome of primo vascular system, some long-type primo vessels dyed with Accepted: Oct 23, 2018 Alcian blue injected into inguinal nodes, abdominal node, and axially nodes were visual- ized, which passed over around the vena cava of the rabbit. The Alcian blue dye revealed KEYWORDS primo vessels and colored blue in the rabbit lymph vessels. The length of long-type primo e m gene expression level; vessels was 18 cm on average, of which diameters were about 20 30 m, and the lymph e m lymph node; vessels had diameters of 100 150 m. Three different tissues of pure primo vessel, mixed primo connectome; primo þ lymph vessel, and only lymph vessel were made to undergo RNA-Seq analysis by RNA-Seq analysis next-generation sequencing. We also analyzed differentially expressed genes (DEGs) from the RNA-Seq data, in which 30 genes of the primo vessels, primo þ lymph vessels, and lymph vessels were selected for primo marker candidates. From the plot of DEG analysis, 10 genes had remarkably different expression pattern on the Group 1 (primo vessel) vs Group 3 (lymph vessel).
    [Show full text]
  • Supplementary Figure 1. Estimation of the Percentage of Volume of Each Population of Extracellular Vesicles by Dynamic Light Scattering
    Supplementary Figure 1. Estimation of the percentage of volume of each population of extracellular vesicles by Dynamic light scattering. Each line represents an independent run. Supplementary Table 1. Differential protein expressionidentified by SWATH MS in neonatalexosomescompared to adultsexosomes (p < 0.05; FCh≠1). Ig: Immunoglobulin. Fold Uniprot Name Gene Name Protein Name p value Change P02786 TFR1 Transferrin receptor protein 1 0.00104 15.32 Q99808 S29A1 Equilibrative nucleide transporter 1 0.01424 14.01 Solute carrier family 2, facilitated glucose P11166 GTR1 0.00106 10.15 transporter member 1 P69892 HBG2 Hemoglobin subunit γ-2 0 9.43 P02792 FRIL Ferritin light chain 0.00438 9.22 Q16819 MEP1A Meprin A subunitα 0.01481 7.13 P02776 PLF4 Platelet factor 4 0.00809 7.01 Q02094 RHAG Ammonium transporter Rh type A 0.00772 6.71 P27918 PROP Properdin 0 6.26 P16671 CD36 Platelet glycoprotein 4 0.00001 5.99 G-protein coupled receptor family C group 5 Q9NQ84 GPC5C 0.00467 5.87 member C P08195 4F2 4F2 cell-surface antigen heavy chain 0.00224 5.77 Q658P3 STEA3 Metalloreductase STEAP3 0.00262 5.34 Q6UX06 OLFM4 Olfactomedin-4 0.00008 5.08 P02787 TRFE Serotransferrin 0.00101 5.05 P08514 ITA2B Integrin αIIb 0 4.88 P02730 B3AT Band 3 anion transport protein 0.00334 4.71 P11142 H7C Heat shock cognate 71 kDa protein 0.00239 4.7 2 of 6 Q15758 AAAT Neutral amino acid transporter B(0) 0.00829 4.56 P05164 PERM Myeloperoxidase 0.00003 4.46 P0DMV9 HS71B Heat shock 70 kDa protein 1B 0.0012 4.36 Erythrocyte band 7 integral P27105 STOM 0.00326 4.18 membrane
    [Show full text]
  • Complexin-1 and Foxp1 Expression Changes Are Novel Brain Effects of Alpha-Synuclein Pathology
    Mol Neurobiol DOI 10.1007/s12035-014-8844-0 Complexin-1 and Foxp1 Expression Changes Are Novel Brain Effects of Alpha-Synuclein Pathology Suzana Gispert & Alexander Kurz & Nadine Brehm & Katrin Rau & Michael Walter & Olaf Riess & Georg Auburger Received: 15 April 2014 /Accepted: 31 July 2014 # The Author(s) 2014. This article is published with open access at Springerlink.com Abstract As the second most frequent neurodegenerative Introduction disorder of the aging population, Parkinson’s disease (PD) is characterized by progressive deficits in spontaneous move- Parkinson’s disease (PD) is the second most frequent age- ment, atrophy of dopaminergic midbrain neurons and aggre- associated brain degeneration disorder, affecting about 1 % gation of the protein alpha-synuclein (SNCA). To elucidate of the population over 65 years of age. The PD-specific molecular events before irreversible cell death, we studied progressive movement deficit is mostly due to the severe synucleinopathy-induced expression changes in mouse brain affliction and cell death of midbrain nigrostriatal dopaminer- and identified 49 midbrain/brainstem-specific transcriptional gic neurons [1]. Surviving neurons in vulnerable regions dysregulations. In particular complexin-1 (Cplx1), Rabl2a and exhibit aggregates predominantly consisting of the protein 14-3-3epsilon (Ywhae) downregulation, as well as upregula- alpha-synuclein, which are visualized as Lewy neurites and tion of the midbrain-specific factor forkhead box P1 (Foxp1) Lewy bodies, both in sporadic late-onset and most familial and of Rabgef1, were interesting as early mRNA level effects early onset PD variants [2]. of alpha-synuclein triggered pathology. The protein levels of Autosomal dominant PD with early clinical manifestation complexin-1 were elevated in midbrain/brainstem tissue of was observed in rare families, leading to the identification of mice with A53T-SNCA overexpression and of mice with alpha-synuclein (SNCA) protein missense mutations such as SNCA-knockout.
    [Show full text]
  • Genetic Background: Understanding Its Importance in Mouse-Based Biomedical Research a Jackson Laboratory Resource Manual
    Genetic Background: Understanding its importance in mouse-based biomedical research A Jackson Laboratory Resource Manual This resource manual highlights the importance of using genetically well-defined mice for biomedical research. It briefly describes the following: • The importance of genetic background • Resources for helping researchers choose the appropriate mouse model • Proper nomenclature to communicate the genetic makeup of mouse models • The Jackson Laboratory’s Genetic Quality Control and Genetic Stability programs Cover Photos Front cover, left: JAX® Mice strain C3H/HeJ (000659) Front cover, middle: Technician displaying holders with straws in the liquid nitrogen storage tank in our Cryopreservation Repository. Front cover, right: JAX® Mice strain C57BL/6J (000664) Table of Contents Introduction ........................................................................................................................ 1 Genetic Background Definition and Examples ............................................................................................ 2 Genetic Background Makes a Difference ................................................................. 2 The Influence of 129 Substrain Backgrounds on Targeted Mutations ................. 4 Consequences of Using Inappropriate Backgrounds .............................................. 4 Minimizing the Confounding Effects of Genetic Background .............................. 5 How Substrains Arise .................................................................................................
    [Show full text]
  • Genetic-Linkage Mapping of Complex Hereditary Disorders to a Whole-Genome Molecular-Interaction Network
    Downloaded from genome.cshlp.org on September 28, 2021 - Published by Cold Spring Harbor Laboratory Press Methods Genetic-linkage mapping of complex hereditary disorders to a whole-genome molecular-interaction network Ivan Iossifov,1 Tian Zheng,2 Miron Baron,3 T. Conrad Gilliam,4 and Andrey Rzhetsky4,5,6 1Department of Biomedical Informatics, Center for Computational Biology and Bioinformatics, Columbia University, New York, New York 10032, USA; 2Department of Statistics, Columbia University, New York, New York 10027, USA; 3Department of Psychiatry, Columbia University, New York, New York 10032, USA; 4Department of Human Genetics, University of Chicago, Chicago, Illinois 60637, USA; 5Department of Medicine, Institute for Genomics & Systems Biology, Computation Institute, University of Chicago, Chicago, Illinois 60637, USA Common hereditary neurodevelopmental disorders such as autism, bipolar disorder, and schizophrenia are most likely both genetically multifactorial and heterogeneous. Because of these characteristics traditional methods for genetic analysis fail when applied to such diseases. To address the problem we propose a novel probabilistic framework that combines the standard genetic linkage formalism with whole-genome molecular-interaction data to predict pathways or networks of interacting genes that contribute to common heritable disorders. We apply the model to three large genotype–phenotype data sets, identify a small number of significant candidate genes for autism (24), bipolar disorder (21), and schizophrenia (25), and
    [Show full text]
  • Characterization of Lymphatic Vessels and Lymphatic Endothelial Cells in Type 2 Diabetes Mellitus
    CHARACTERIZATION OF LYMPHATIC VESSELS AND LYMPHATIC ENDOTHELIAL CELLS IN TYPE 2 DIABETES MELLITUS Structural, morphological and molecular analysis DOCTORAL THESIS for obtaining the academic degree of Doctor of Philosophy (Ph.D.) submitted by Monika Hämmerle, MD within the thematic program: Cell communication in health and disease (CCHD) supervised by Prof. Dr. Dontscho Kerjaschki & Dr. Brigitte Hantusch Clinical Institute of Pathology Vienna, August 2012 Acknowledgements First of all, I would like to thank my supervisors Prof. Dontscho Kerjaschki und Dr. Brigitte Hantusch for giving me the opportunity to do my PhD in the research laboratory of the Clinical Institute of Pathology and who supported me throughout the years. I would like to thank my cooperation partners at the Department of Rheumatology, Carl‐Walter Steiner, for excellent technical assistence in FACS sorting and at the Department of General Surgery, Dr. Christoph Neumayer, for guaranteeing me that I could use my material as fresh as possible. Moreover, I would like to thank Dr. Stefan Thurner and especially Dejan Stokic for helping me with the bioinformatical data analysis. I thank all my friends and colleagues from the CCHD PhD program, especially my lab mate Tom without whom the day would not have been so much fun. I would like to announce a big thank to Bernhard Höfle, who triggered my enthusiasm for science. Last but not least, I would like to thank my family and friends for their incessant support, love and motivation. Die Wissenschaft, richtig verstanden, heilt den Menschen von seinem Stolz; denn sie zeigt ihm seine Grenzen. Albert Schweitzer Summary Background ‐ Small vessel disease of kidney, nerves, retina and skin, referred to as microangiopathy, is a major cause of morbidity in type 2 diabetes mellitus (T2DM).
    [Show full text]
  • Investigation of the Interaction of Multimerin 1 with Components of Prothrombinase, in Vitro
    INVESTIGATION OF THE INTERACTION OF MULTIMERIN 1 WITH COMPONENTS OF PROTHROMBINASE, IN VITRO By ZAINAB A MOTALA, BSc HONOURS A Thesis Submitted to the School of Graduate Studies in Partial Fulfilment of the Requirements for the Degree Master of Science McMaster University © Copyright by Zainab A Motala, December 2011 MASTER OF SCIENCE (2011) (Medical Sciences) TITLE: Investigation of the interaction of multimerin 1 with components of prothrombinase, in vitro AUTHOR: Zainab A Motala, BSc Honours (McMaster University) SUPERVISOR: Professor Catherine P. M. Hayward NUMBER OF PAGES: xiv, 135 ii ABSTRACT Prothrombinase is an enzymatic complex that accelerates the conversion of prothrombin to thrombin for efficient blood clot formation at sites of vessel injury. Prothrombinase consists of the enzyme factor Xa and its cofactor factor Va, assembled on a phosphatidyl serine-containing membrane, in the presence of calcium. Multimerin 1 (MMRN1) is a polymeric, factor V/Va-, prothrombin-, and phosphatidyl serine-binding protein that is stored in platelet and endothelial cell secretion granules. When released, MMRN1 binds to their cell surface and to the extracellular matrix. Unlike plasma factor Va, platelet factor Va is stored complexed to MMRN1 in platelet α-granules, and is resistant to inactivation by activated protein C (APC). Previous studies revealed that exogenous MMRN1 inhibits thrombin generation in plasma. My thesis investigated the interaction of MMRN1 with components of prothrombinase in order to elucidate the molecular mechanisms by which MMRN1 modulates coagulation. ELISA binding assays that used prothrombin derivatives revealed that the prothrombin gamma-carboxyglutamic acid and kringle domains have potential MMRN1 binding sites. Thrombin generation assays that used purified proteins and/or phospholipid vesicles revealed that MMRN1 inhibits thrombin generation in the presence and absence of factor Va, but not in the absence of phospholipid vesicles.
    [Show full text]