Cd11b Regulation of B Cell Receptor Signaling in Health and Systemic Lupus Erythematosus
Total Page：16
File Type：pdf, Size：1020Kb
University of Louisville ThinkIR: The University of Louisville's Institutional Repository Electronic Theses and Dissertations 12-2018 CD11b regulation of B cell receptor signaling in health and systemic lupus erythematosus. Paul Lorenzo Dascani University of Louisville Follow this and additional works at: https://ir.library.louisville.edu/etd Part of the Immunity Commons Recommended Citation Dascani, Paul Lorenzo, "CD11b regulation of B cell receptor signaling in health and systemic lupus erythematosus." (2018). Electronic Theses and Dissertations. Paper 3076. https://doi.org/10.18297/etd/3076 This Doctoral Dissertation is brought to you for free and open access by ThinkIR: The University of Louisville's Institutional Repository. It has been accepted for inclusion in Electronic Theses and Dissertations by an authorized administrator of ThinkIR: The University of Louisville's Institutional Repository. This title appears here courtesy of the author, who has retained all other copyrights. For more information, please contact [email protected]. CD11B REGULATION OF B CELL RECEPTOR SIGNALING IN HEALTH AND SYSTEMIC LUPUS ERYTHEMATOSUS By Paul Lorenzo Dascani B.S., University of Pittsburgh, 2011 M.S., University of Louisville, 2015 A Dissertation Submitted to the Faculty of the School of Medicine in Partial Fulfillment of the Requirements for the Degree of Doctor of Philosophy in Microbiology and Immunology Department of Microbiology and Immunology University of Louisville Louisville, Kentucky December 2018 Copyright 2018 by Paul Lorenzo Dascani All rights reserved CD11B REGULATION OF B CELL RECEPTOR SIGNALING IN HEALTH AND SYSTEMIC LUPUS ERYTHEMATOSUS By Paul Lorenzo Dascani B.S., University of Pittsburgh, 2008 M.S., University of Louisville, 2015 A Dissertation Approved on December 3, 2018 by the following Dissertation Committee: _____________________________ Jun Yan, M.D., Ph.D. Dissertation Director _____________________________ Michele Kosiewicz, Ph.D. _____________________________ Pascale Alard, Ph.D. _____________________________ Venkatakrishna Jala, Ph.D. ____________________________ David Powell, Ph.D. ii ABSTRACT CD11B REGULATION OF B CELL RECEPTOR SIGNALING IN HEALTH AND SYSTEMIC LUPUS ERYTHEMATOSUS Paul Dascani December 3, 2018 Loss of function mutation in CD11b has been associated with incidence of systemic lupus erythematosus (SLE), a disease driven by B cell production of pathogenic autoimmune antibody. Our previous work has revealed the ability of CD11b to regulate B cell receptor (BCR) signaling and control autoimmune disease in mice. However, how CD11b regulates the immune response under normal conditions remains unknown. Through the use of a CD11b knockout model, we demonstrated that CD11b-deficient mice have an elevated antigen-specific humoral response in a Th2 type immunization. Deletion of CD11b resulted in elevated serum IgM and IgG antibody, increases in antigen-specific germinal center (GC) B cells and plasma cells (PCs), and increased expression of survival and regulatory factors Bcl-XL and Blimp1 within these subsets. Subsequent experiments using a tissue-specific CD11b deletion model revealed this effect to be B cell intrinsic, and not altered by myeloid cell CD11b expression. Examination of BCR signaling in GC B cells of CD11b knockouts revealed defects in association of negative regulators SHP-1 and pLyn with the BCR following stimulation, as well as alterations in nuclear iii localization of GC regulators cMyc and FOXO1. Through the use of a CD11b-reporter mouse model, we identified multiple novel CD11b-expressing B cell subsets that are dynamically altered during immunization and autoimmune conditions. Lastly, we investigated the incidence of CD11b mutation rs1143679 in SLE patients and examined its effect on the clinical outcome of disease and impact on B cell function. These studies describe a novel role for CD11b in regulation of the healthy humoral response at the GC stage and reveal previously unknown populations of CD11b expressing B cell subsets, suggesting a complex function for CD11b in B cells during development and maturation. iv TABLE OF CONTENTS PAGE SIGNATURE PAGE ........................................................................................................ii ABSTRACT .......................................................................................................iii LIST OF FIGURES .......................................................................................................vii INTRODUCTION .........................................................................................................1 General Introduction ......................................................................1 Regulation of B Cells and the Germinal Center Reaction .............2 Classic Functions CD11b .............................................................16 CD11b Regulates B Cell Receptor Signaling ..............................18 CHAPTER I: CD11B REGULATION OF B CELL RECEPTOR SIGNALING IN THE NORMAL IMMUNE RESPONSE ..........................................................................22 Introduction ..................................................................................23 Materials and Methods .................................................................27 Results ..........................................................................................31 Global CD11b Deficiency Alters Antibody Production and GC Formation in Non-autoimmune Mice ...........................................31 CD11b Regulates Germinal Center B Cell BCR Signaling .........37 B Intrinsic CD11b Regulates the Th2 Antibody Response ….....45 Myeloid Cell Loss of CD11b Does Not Impact the Th2 Antibody Response……………………………………………...50 Discussion ....................................................................................54 CHAPTER II: DYNAMIC EXPRESSION OF CD11B ON B CELLS IN NAÏVE AND IMMUNE SETTINGS ............................................................................................62 Introduction ..................................................................................63 Materials and Methods .................................................................67 Results………………………………………………………...…70 B cell CD11b Expression is Altered in a Th2 Immunization Setting ...........................................................................................70 Progression of Lupus-Like Disease in Mice Impacts B Cell Expression of CD11b ...................................................................81 Comparison of the CD11b Expressing Plasma Cell Repertoire ...87 Discussion .....................................................................................91 CHAPTER III: THE IMPACT OF CD11B POLYMORPHISM ON B CELLS IN SYSTEMIC LUPUS ERYTHEMATOSUS......................................................................96 Introduction ..................................................................................97 Materials and Methods ...............................................................106 v Results ........................................................................................109 A Phenotypic Survey of ITGAM Mutation rs1143679 in SLE Patients of the Louisville Region ................................................109 Discussion ...................................................................................116 CONCLUSIONARY REMARKS AND FUTURE PERSPECTIVES ..........................119 CHAPTER IV: TRASNCRIPTION FACTOR STAT3 SERVES AS A NEGATIVE REGULATOR CONTROLLING IGE CLASS SWITCHING ......................................123 Introduction .................................................................................124 Materials and Methods ................................................................132 Results .........................................................................................134 STAT3 Deficiency in B and T Cell Subsets Disrupts the Regulation of Class Switching in a Th2-type Immunization ......137 STAT3-Dependent Regulation of Germinal Center Class Switching is a B cell Intrinsic effect ...........................................142 Depletion of STAT3 Regulates Germinal Center Class Switching In Vitro ........................................................................................151 Target Genes of STAT3 in the GC B Cells .................................156 STAT3 in B Cells Regulates Lung Inflammation During Inhaled Allergy Reactions ........................................................................160 Discussion ...................................................................................164 REFERENCES .....................................................................................................169 CURRICULUM VITAE .....................................................................................................200 vi LIST OF FIGURES Figure 1: Increased antibody response in CD11b-knockout mice following Immunization ....................................................................................................................33 Figure 2: Germinal center B cells are increased in CD11b-knockout mice .....................34 Figure 3: Plasma cells are increased in CD11b-knockout mice .......................................35 Figure 4: CD11b deficiency alters GC and PC regulator expression ...............................36 Figure 5: Loss of CD11b alters SHP-1 colocalization with the BCR ...............................39 Figure 6: Loss of CD11b alters pLyn colocalization