Zika virus induced neuro-ocular pathology correlates with autoreactive anti-ganglioside antibodies and can be ameliorated with cutaneous vaccination Open Access
Beaver, Jacob (Fall 2020)
Abstract
Zika virus (ZIKV) garnered international attention between 2015-2017, initially infecting pregnant women in Brazil and later attacking indiscriminately residents of the Americas. Infection of pregnant population resulted in a newly identified fetal-maternal pathology including severe neurological impairments such as microcephaly, which is now part of the congenital ZIKV syndrome (CZS). Among adults, ZIKV caused moderate to severe neuro-ocular sequelae, with symptoms ranging from conjunctivitis to Guillain-Barré Syndrome (GBS). The lack of case reports of ZIKV since 2017 may be directly attributed to increased herd immunity and better arbovirus vector control. However, children born from mothers infected with ZIKV continue to demonstrate the long-term impacts of fetal-maternal ZIKV infection, as reports of CSZ-related brain and eye developmental abnormalities continue to emerge among children born without microcephaly.
In this dissertation, we used immunocompetent BALB/c mice, which generate robust humoral immune responses, to investigate long-term impacts of ZIKV infection. Using a high infectious dose to overcome natural interferon barriers that may hinder murine infections, we found that immunocompetent mice exhibited motor impairment such as arthralgia, as well as ocular inflammation resulting in retinal vascular damage, and corneal edema. This pathology persisted 100 days after infection with evidence of chronic inflammation in immune-privileged compartments, demyelination in the hippocampus and motor cortex regions of the brain, and retinal/corneal hyperplasia. Pathology in immunocompetent animals coincided with weakly neutralizing antibodies and increased ADE among ZIKV strains (PRVABC59, FLR, and MR766) and all Dengue virus (DENV) serotypes. These antibodies were autoreactive to GBS-associated gangliosides. This study highlights the importance of longevity studies in ZIKV infection and confirms the role of anti-ganglioside antibodies in ZIKV-induced neuro-ocular disease.
Table of Contents
ABSTRACT.. 2
CHAPTER I: INTRODUCTION.. 8
ABSTRACT.. 8
INTRODUCTION.. 9
SECTION I.I - HISTORY OF VIRUS EMERGENCE.. 10
FIGURE 2. Phylogenetic analysis of ZIKV genomes by region. 16
SECTION I.III - ZIKV TRANSMISSION AND TISSUE TROPISM... 19
Vector influence on viral evolution. 19
Lineage-Dependent Differences in Animal and Cell Culture Models. 25
Correlates of Inflammation during ZIKV infection in Eyes and Brain. 29
SECTION I.IV - CURRENT UNDERSTANDING OF ZIKV AND THE HOST IMMUNE RESPONSE.. 30
FIGURE 3. ZIKV activates signaling pathways that promote transcription of IFN genes differentially. 31
Type I interferon responses. 31
Lineage differences. 33
Type II Interferon Responses. 36
Type III Interferon Responses. 38
ZIKV Vaccine development. 40
Knowledge gaps and future studies. 42
CONCLUSIONS. 43
SECTION I.V - Introduction to Dissertation Research. 45
CHAPTER II : Zika virus-induced neuro-ocular pathology in immunocompetent mice correlates with anti-ganglioside autoantibodies. 46
ABSTRACT.. 47
Introduction.. 48
METHODS AND MATERIALS. 50
Results. 60
ZIKV-PRVABC59 infection in immunocompetent mice results in distinct neurological and ocular pathology. 60
ZIKV infection-induced antibodies have limited cross-protection and enhance infection among ZIKV and DENV strains. 64
Corneal and retinal hyperplasia among infected mice coincides with elevated cell death and persistent inflammation in the eye. 66
ZIKV infection reduces myelin expression and results in prolonged inflammation in the brain. 68
Antibodies generated during ZIKV infection are auto-reactive to GBS-associated neural markers. 69
Discussion.. 70
Acknowledgements. 75
FIGURES. 76
Figure 1. PRVABC59 can infect systemic and privileged organs in immunocompetent mice. 77
Figure 2. Competent mice infected with ZIKV-PRVABC59 demonstrate distinct ocular and motor/neural symptomatology that correlate with viral load. 79
Figure 3. PRVABC59-induced antibodies are weakly neutralizing and enhance non-homologous infection. 81
Figure 4. Tenacious corneal and retinal hyperplasia coincides with persistent ocular infection, increasing levels of cell death, and chronic inflammation. 83
Figure 5. PRVABC59 infection in the hippocampus and cortex corresponds to decreased levels of myelin and persistent inflammation. 85
Figure 6. IgG produced during ZIKV infection are cross-reactive to host gangliosides. 87
Supplementary Fig 1. ZIKV-PRVABC59 demyelinates cortex and hippocampus. 89
Supplementary Fig 2. Intraocular infection increases apoptosis and necroptosis. 90
CHAPTER III : Cutaneous vaccination ameliorates Zika virus-induced neuro-ocular pathology via reduction of anti-ganglioside antibodies. 92
ABSTRACT.. 93
Introduction.. 94
METHODS AND MATERIALS. 96
Results. 108
MNP vaccination induced a greater concentration of vaccine-specific antibodies with higher avidity. 108
MNP vaccinations induced sustained B and T cell responses while IM vaccination induced ephemeral T cell secretory responses. 110
Unadjuvanted IM vaccination offered little protection against infectious challenge. 111
IM vaccination exacerbated intraocular pathology without protecting against infection or cell death. 113
MNP vaccination protected against ZIKV-induced demyelination and chronic neuroinflammation. 116
MNP antibodies demonstrate greater breadth of neutralization, less ADE, and are less auto-reactive. 118
Discussion.. 120
Acknowledgements & Figures. 126
Figure 1. Cutaneous vaccinations induced greater antibody concentrations that bind with higher avidity. 129
Figure 2. Cutaneous vaccination generated greater B and T cell responses within 2 weeks after of vaccination. 131
Figure 3. Cutaneous vaccination reduced viral burden and protected against pathogenic weight loss and neuro-ocular symptoms. 133
Figure 4. Low-quality antibodies triggered acute ocular pathology, increased ocular infectivity, and elevated cell death persisting up to 100 DPI. 135
Figure 5. Low-quality antibodies elevated infection, lower myelin expression, and chronically stimulate pro-inflammatory cytokine expression. 137
Figure 6. Antibodies generated by MNP vaccination remain elevated after infection challenge. 139
Figure 7. IM-induced antibodies demonstrated lower neutralizing activity than those generated by MNP. 140
Figure 9. Antibodies generated by MNP vaccination were less auto-reactive to gangliosides. 142
Supplementary Fig 1. ZVIP incorporation into MNP for cutaneous vaccination maintained antigen functionality. 143
Supplementary Fig 2. Gating strategies utilized for flow cytometry experiments. 145
Supplementary Fig 3. MNP conferred protection against intraocular apoptosis and necroptosis induced by ZIKV infection. 147
Supplementary Fig 4. MNP protected against ZIKV-induced demyelination. 149
Supplementary Tables summarizing the neutralization and ADE qualities of vaccination induced antibodies. 151
CHAPTER IV: CONCLUSIONS. 152
REFERENCES. 156
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