UC Irvine UC Irvine Electronic Theses and Dissertations Title Generation and application of a chimeric model to examine human microglia responses to amyloid pathology Permalink https://escholarship.org/uc/item/38g5s0sj Author Hasselmann, Jonathan Publication Date 2021 Supplemental Material https://escholarship.org/uc/item/38g5s0sj#supplemental Peer reviewed|Thesis/dissertation eScholarship.org Powered by the California Digital Library University of California UNIVERSITY OF CALIFORNIA, IRVINE Generation and application of a chimeric model to examine human microglia responses to amyloid pathology DISSERTATION Submitted in partial satisfaction of the requirements for the degree of DOCTOR OF PHILOSOPHY In Biological Sciences By Jonathan Hasselmann Dissertation Committee: Associate Professor Mathew Blurton-Jones, Chair Associate Professor Kim Green Professor Brian Cummings 2021 Portions of the Introduction and Chapter 2 © 2020 Wiley Periodicals, Inc. Chapter 1 and a portion of Chapter 2 © 2019 Elsevier Inc. All other materials © 2021 Jonathan Hasselmann DEDICATION To Nikki and Isla: During this journey, I have been fortunate enough to have the love and support of two of the strongest ladies I have ever met. I can say without a doubt that I would not have made it here without the two of you standing behind me. I love you both with all my heart and this achievement belongs to the two of you just as much as it belongs to me. Thank you for always having my back. ii TABLE OF CONTENTS LIST OF FIGURES .......................................................................................................................v LIST OF TABLES ...................................................................................................................... vii ACKNOWLEDGEMENTS ...................................................................................................... viii CURRICULUM VITAE .............................................................................................................. xi ABSTRACT OF THE DISSERTATION ................................................................................ xiv INTRODUCTION..........................................................................................................................1 A. MICROGLIA AND THEIR ROLES IN ALZHEIMER’S DISEASE ...................................3 History of Microglia ............................................................................................................3 Microglia in Alzheimer’s Disease .......................................................................................5 B. CURRENT METHODS OF STUDYING MICROGLIA ......................................................8 Mouse Microglia ..................................................................................................................8 Primary Human Microglia ...................................................................................................9 iPSC-derived Human Microglia ........................................................................................11 Organoid Microglia ............................................................................................................14 C. PITFALLS ASSOCIATED WITH THE CURRENT MODELS ........................................16 D. MICROGLIA XENOTRANSPLANTATION INTO THE MOUSE BRAIN .....................19 CHAPTER 1: TRANSPLANTATION OF HUMAN IPSC-DERIVED MICROGLIA INTO THE MOUSE BRAIN INDUCES APPROPRIATE TRANSCRIPTOMIC PROGRAMMING AND REVEALS HUMAN SPECIFIC AMYLOID RESPONSES ........21 INTRODUCTION ......................................................................................................................21 MATERIALS AND METHODS ...............................................................................................23 RESULTS ...................................................................................................................................40 DISCUSSION ............................................................................................................................69 CHAPTER 2: THE ALZHEIMER’S DISEASE-ASSOCIATED TREM2 R47H GENETIC POLYMORPHISM DRIVES TRANSCRIPTOMIC AND FUNCTIONAL ALTERATIONS IN XENOTRANSPLANTED MICROGLIA ..............................................71 INTRODUCTION ......................................................................................................................71 MATERIALS AND METHODS ...............................................................................................72 RESULTS ...................................................................................................................................79 DISCUSSION ............................................................................................................................85 CHAPTER 3: ALTERATIONS TO CHROMATIN STRUCTURE AND TRANSCRIPTOMIC PROGRAMMING UNDERLIE THE MICROGLIA RESPONSE TO AMYLOID PATHOLOGY ..................................................................................................87 INTRODUCTION ......................................................................................................................87 iii MATERIALS AND METHODS ...............................................................................................88 RESULTS ...................................................................................................................................94 DISCUSSION ..........................................................................................................................101 CONCLUSIONS AND FUTURE DIRECTIONS ...................................................................105 REFERENCES ...........................................................................................................................111 iv LIST OF FIGURES INTRODUCTION FIGURE 1: Homology between human and mouse proteins associated with Alzheimer’s disease risk ...................................................................................................................................1 FIGURE 2: In vivo and in vitro microglia ontogeny .................................................................12 CHAPTER 1 FIGURE 1.1: iPSC-derived human HPCs differentiate into microglia and display robust engraftment within the forebrain of MITRG mice .....................................................................41 FIGURE 1.2: Xenotransplanted human cells express microglia markers ..................................43 FIGURE 1.3: Chimeric distribution of human xMG is influenced by age of transplantation and hCSF1 expression level ..............................................................................................................44 FIGURE 1.4: Transplanted iHPCs adapt to diverse niches within the brain and differentiate into the four CNS macrophage subtypes in a context-dependent manner .................................45 FIGURE 1.5: Methods of tissue dissociation and validation of the human microglia flow cytometry paradigm ....................................................................................................................48 FIGURE 1.6: Transplantation of human iHPCs into the murine brain recapitulates an in vivo human transcriptome ..................................................................................................................49 FIGURE 1.7: xMG cluster closely with brain-derived human microglia ..................................52 FIGURE 1.8: xMG survey their surroundings, rapidly respond to laser ablation, and interact with neuronal components after trauma .....................................................................................55 FIGURE 1.9: Differential responses of xMG and iMG to LPS administration .........................57 FIGURE 1.10: LPS treatment upregulates numerous inflammatory pathways and induces a transcriptomic profile resembling disease-affected microglia ...................................................59 FIGURE 1.11: xMG gene expression profiles are indicative of known microglial activation signatures ....................................................................................................................................61 FIGURE 1.12: xMG down-regulate homeostatic markers and upregulate activation markers around Aβ plaques ......................................................................................................................63 FIGURE 1.13: Single-cell sequencing of xMG from MITRG and MITRG-5X mice reveals altered population distributions and human-specific genetic responses ....................................65 FIGURE 1.14: Analysis of scRNA-seq by individual sex or genes reveals additional aspects of xMG transcriptomics ..................................................................................................................67 CHAPTER 2 FIGURE 2.1: Deletion of TREM2 suppresses the development of Disease-Associated Microglia in vivo ........................................................................................................................80 FIGURE 2.2: CRISPR/Cas9 editing to create a homozygous TREM2 R47H mutant human iPSC line .....................................................................................................................................81 v FIGURE 2.3: xMG response to Aβ plaques is altered by the TREM2 R47H mutation .............82 FIGURE 2.4: The TREM2 R47H mutation alters the population-level response to amyloid in xMG ..........................................................................................................................................83