The Roles of Microbiota and Innate Immunity during Rotavirus Infection and Humoral Immunity 公开
Uchiyama, Robin Cathleen (2014)
Published
Abstract
Rotavirus (RV) infection is the leading cause of viral gastroenteritis amongst children globally and is responsible for upwards of 500,000 deaths annually. While 2 vaccines are available, they do not adequately protect against disease amongst children in developing countries. With this in mind, we aimed to determine if environmental factors, specifically gut microbiota, contribute to infection and induction of humoral responses. Using an infection model, we found that microbiota-ablation, achieved through antibiotic treatment or germ-free (GF) conditions, experienced a delay in virus shedding. Postponed shedding correlated with less virus at the small intestine, demonstrating that microbiota speeds infection. Microbiota promoted RV entry as antibiotics had no effect on replication. Antibiotics also protected against RV diarrhea, demonstrating a negative role for microbiota during disease. Microbiota inhibited humoral immunity as antibiotics enhanced RV antibody production and small intestinal, IgA-producing antibody-secreting cells (ASCs) frequencies. To examine if microbiota exposure reversed the antibody-enhancing effects of antibiotics, dextran sodium sulfate (DSS) was administered. DSS increased inflammation and weakened RV antibody responses, indicating that microbiota-derived inflammation inhibits humoral immunity. Thus, the microbiota could be an environmental factor involved in vaccine inefficacy and serve as a target for therapeutics.
We also aimed to elucidate how innate immunity controls RV infection and humoral immunity, as innate immunity to RV is understudied. When mice lacking MyD88, an adaptor protein for most TLRs and inflammasome cytokine receptors, were infected, they shed more virus and experienced virus spread, demonstrating MyD88 controls infection and spread. Control of primary infection was independent of inflammasome cytokines IL-1 and -18, indicating that TLRs were responsible for limiting infection. MyD88-deficient neonates experienced greater incidence of and days with diarrhea, demonstrating that MyD88 protects against disease. Mice lacking MyD88 also experienced slowed systemic antibody responses and skewed IgG subisotype switching, with a bias towards Th2-associated IgG1 and away from Th1-associated IgG2c. MyD88's influence on antibody responses originated from bone marrow-derived MyD88, but not epithelial MyD88, and IL-1 and -18 signaling prompted proper subisotype switching. Insights into RV-specific innate immunity uncovered novel therapeutic targets, including MyD88, TLRs, and inflammasome cytokines, during RV infection and vaccination.
Table of Contents
TABLE OF CONTENTS
CHAPTER 1: INTRODUCTION 1
CHAPTER 2: ANTIBIOTIC TREATMENT SUPPRESSES ROTAVIRUS INFECTION AND ENHANCES SPECIFIC HUMORAL IMMUNITY 24
Abstract 25
Introduction 26
Materials and Methods 28
Results 31
Discussion 37
Figures 41
Figure Legends 51
CHAPTER 3: MYD88-MEDIATED TLR SIGNALING PROTECTS AGAINST ACUTE ROTAVIRUS INFECTION WHILE INFLAMMASOME CYTOKINES DIRECT ANTIBODY RESPONSE 55
Abstract 56
Introduction 57
Materials and Methods 60
Results 63
Discussion 73
Figures 74
Figure Legends 86
CHAPTER 4: DISCUSSION 92
REFERENCES 103
CHAPTER 1: INTRODUCTION
1.1 Rotavirus Virology 2
1.1.1 Rotavirus structure and classification 2
1.1.2 Rotavirus replication 3
1.2 Rotavirus Pathogenesis 4
1.2.1 Rotavirus transmission, target cells, and kinetics on infection 4
1.2.2 Rotavirus-mediated induction of diarrhea 4
1.3 Rotavirus Treatment and Prevention 5
1.4 Rotavirus Vaccine Efficacy in Developing Nations 5
1.4.1 Rates of vaccine efficacy in developing nations 5
1.4.2 Proposed explanations for lack of vaccine efficacy 6
1.4.2.1 Environmental enteropathy 6
1.4.2.2 Pathogenic organism co-infection 6
1.4.2.3 Undernutrition 7
1.4.2.4 Maternal antibody interference 8
1.5 The Immune Response to Rotavirus Infection 8
1.5.1 The innate immune response 9
1.5.1.1 TLRs 9
1.5.1.2 PKRs 10
1.5.1.3 RLRs 10
1.5.2 The adaptive immune response 10
1.5.2.1 CD4 T cells 10
1.5.2.2 CD8 T cells 11
1.5.2.3 T regulatory and γδ T cells 12
1.5.2.4 B cells 12
1.5.2.5 Antibody response 13
1.5.3 IFN response to rotavirus infection 13
1.5.3.1 Type I IFN 13
1.5.3.2 Type II IFN 14
1.5.3.3 Type III IFN 14
1.6 The Importance of Microbiota on Immunity and Immune System
Development 14
1.6.1 Intestinal architecture and gut-associated lymphoid tissue 15
1.6.2 Innate immune cells 16
1.6.2.1 Macrophages 16
1.6.2.2 Dendritic cells 16
1.6.2.3 Group 3 innate lymphoid cells 16
1.6.2.4 Intestinal epithelial cells 17
1.6.3 Adaptive immune cells 17
1.6.3.1 B cells 17
1.6.3.2 Th17-T regulatory immunity bias 18
1.6.3.3 Th1-Th2 immunity bias 18
1.7 Interactions Between the Microbiota and Viruses 18
1.7.1 PR8 influenza A virus 19
1.7.2 Poliovirus, reovirus, and mouse mammary tumor virus 19
1.7.3 HIV 20
1.8 Probiotics in Rotavirus Disease, Infection, and Immunity 21
1.8.1 Preventing disease 21
1.8.2 Treating disease 22
1.8.3 Limiting infection 23
1.8.4 Enhancing virus-specific B cell and antibody responses 23
LIST OF FIGURES
(Listed in order of appearance in the chapter)
CHAPTER 2
2-1 Ablation of microbiota retards RV infectivity 41
2-2 Antibiotic treatment reduces RV-induced diarrhea in neonatal mice 42
2-3 Antibiotic treatment enhances the durability of the antibody response to RV 43
2-4 Germ-free mice exhibit enhanced serum antibody response to RV 44
2-5 Antibiotic-treated neonatal mice exhibit enhanced serum IgA production following RV inoculation 45
2-6 Antibiotic treatment results in greater maintenance of RV-specific antibody-producing cells in the intestine 46
2-7 Increasing basal immune activation impairs RV-induced antibody generation 47
S2-1 Antibiotic treatment reduces fecal bacterial load by 99% and cessation of antibiotics results in rapid restoration of bacterial loads in feces 48
S2-2 Antibiotic treatment lowers total fecal IgA 49
S2-3 A 5-day antibiotic treatment modestly enhances RV antibody generation 50
CHAPTER 3
3-1 MyD88 signaling contributes to the control of RV infection in adult mice 73
3-2 MyD88 limits RV spread to the colon and blood 74
3-3 MyD88 signaling protects neonatal mice from RV disease 75
3-4 MyD88 signaling in adult mice enhances anti-RV Ab generation and proper Ab sub-type switch 76
3-5 MyD88 signaling in neonatal mice enhances anti-RV Ab generation and proper Ab sub-type switch 77
3-6 MyD88 signaling in radio-sensitive/bone marrow derived cells but not radio-resistant cells contributes to the production of RV-specific IgG and IgA 78
3-7 MyD88 in radio-sensitive/bone marrow derived cells but not radio-resistant cells assist in proper Ab sub-type switch 79
3-8 Anti-RV IgG and IgA is independent of bone marrow-derived IL-1 and-18, however proper Ab sub-type switch is dependent on both IL-1 and -18 80
S3-1 IL-1 or -18 alone is not important for control of primary RV infection 81
S3-2 MyD88 signaling in either radio-sensitive/bone marrow-derived cells or radio-resistant cells contributes to the control of infection 82
S3-3 MyD88 induced non-specific (total) IgA production in the serum 83
S3-4 IgG sub-type switch is not dependent on either IL-1 or -18 signaling 84
S3-5 IgG sub-type switch is not dependent on the NLRP3 inflammasome 85
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