Biology subjects

Himmler, G.

Publications and source records attributed to Himmler, G..

3 recordsLinked to original sources

bioRxiv2023

Age-dependent Powassan Virus Lethality is Directed by Glial Cell Activation and Divergent Neuroinflammatory Cytokine Responses in a Murine Model

Powassan virus (POWV) is an emergent tick-borne flavivirus that causes fatal encephalitis in the elderly and long-term neurologic sequelae in survivors. How age contributes to severe POWV encephalitis remains an enigma and there are currently no animal models that reflect age-dependent POWV neuropathology. Inoculating C57BL/6 mice with a POWV strain (LI9) currently circulating in Ixodes ticks, resulted in age-dependent POWV lethality with overt spongiform brain damage 10-15 dpi. Infection of 50 week old mice resulted in 82% lethality 10-15 dpi that was sequentially reduced by age to 7.1% in 10 week old mice. LI9 encephalitis resulted in early neuronal depletion, with severe CNS damage, persistent inflammatory gliosis and long-term spongiform pathology in survivors (30 dpi). In all mice POWV LI9 was neuroinvasive and reached maximum POWV loads in the CNS 10 dpi. Coincident with murine lethality, in 50 week old mice maximum POWV CNS levels persisted 15 dpi, while instead decreasing by 2-4 logs in 10-30 week old mice. Although glial cells were highly activated in all POWV infected mice, differences in age-dependent CNS cytokine responses were striking 15 dpi. In 50 week old mice POWV induced Th1-type cytokines (IFN{gamma}, IL-2, IL-12, IL-4, TNF, IL-6), suggesting a pro-inflammatory M1 microglial activation cascade. In contrast, POWV induced Th2-type cytokines (IL-10, TGF{beta}, IL-4) in 10 week old mice consistent with a neuroprotective M2 microglial phenotype. These findings reflect differences in neurodegenerative versus neuroprotective glial cell responses that correlate with divergent CNS viral clearance and age-dependent POWV LI9 lethality. Discrete age-dependent CNS cytokine responses suggest neuroinflammatory targets as potential POWV therapeutics. These studies establish a highly lethal POWV murine model and reveal a hyperinflammatory mechanism of age-dependent POWV lethality that mirrors human POWV severity and long-term CNS sequelae in the elderly. ImportancePowassan virus is an emerging tick-borne flavivirus causing lethal encephalitis in aged individuals. We reveal an age-dependent POWV murine model that mirrors human POWV encephalitis and long-term CNS damage in the elderly. Findings demonstrate that POWV load and discrete glial cell cytokine responses in the CNS are critical determinants of age-dependent POWV lethality. POWV age-independently activates microglia and astrocytes, but directs neuroprotective Th2 cytokine responses in 10 week old mice and distinct pro-inflammatory Th1 cytokine responses in the CNS of 50 week old mice. This reveals roles for a hyperinflammatory CNS cytokine cascade in age-dependent POWV lethality, and protective anti-inflammatory cytokines in murine survival. Notably, results define potential therapeutic targets and rationalize approaches for preventing severe POWV encephalitis that may be broadly applicable to neurodegenerative diseases. This age-dependent murine POWV model permits analysis of vaccines, and therapeutics that prevent POWV neuroinvasion or resolve severe POWV encephalitis in the elderly.

microbiology↗

bioRxiv2023

Age-dependent Powassan Virus Lethality and Neuropathogenesis in Mice

Powassan viruses (POWV) are emergent tick-borne flaviviruses that cause severe neurologic disease in humans. Subcutaneous inoculation of C57BL/6 mice with POWV (strain LI9) resulted in overt brain damage resembling spongiform encephalitis. Noting higher POWV lethality in older mice, we assessed neurovirulence as a function of age. We found that POWV LI9 inoculation was lethal in 80% of 50 wk old mice, 10-15 dpi, and that lethality was sequentially reduced in 40, 30, 20, 10 wk old mice to <10%. Lethality was conferred by 2-20 POWV FFUs, and POWV neuropathology was evident as early as 5 dpi, with lethal disease 10-15 dpi correlated with sustained POWV RNA levels in brains of aged mice. Histology of POWV infected 50 wk old murine brains revealed severe spongiform neuronal necrosis, microgliosis, and inflammation with increased brainstem and cerebellar damage. These findings delineate an age-dependent murine model of lethal POWV infection that mirrors human POWV disease and permits analysis of age-dependent neurovirulence determinants. SignificanceOur findings establish a novel age-dependent lethal animal model to study encephalitic POWV disease in vivo. These initial findings demonstrate that following peripheral inoculation, non-neuroadapted POWV LI9 is neuroinvasive and enters the brains of young and aged mice. However, POWV LI9 lethality is strictly age-dependent and correlated with increased viral load in the brains of aged mice. POWV rapidly directs neuronal loss and spongiform lesions, microglial activation and causes prolonged inflammation that fails to clear POWV from the brains of aged mice. Our results provide a lethal murine model of POWV neurovirulence that mirrors the prevalence of severe human POWV encephalitis in the elderly. This lethal murine POWV model provides mechanisms for defining POWV protective responses of the young, revealing determinants of age-dependent POWV lethality and evaluating potential POWV therapeutics. SUMMARYPowassan virus is an emerging tick-borne flavivirus linked to severe neurologic disease in aged individuals. Here we describe an age-dependent mouse model of POWV pathogenesis. SUBJECTSPowassan virus, flavivirus, neurovirulence, neuroinvasion, neurotropic, spongiform encephalopathy, microgliosis, neuroinflammation

microbiology↗

bioRxiv2021

Powassan Viruses Spread Cell to Cell During Direct Isolation from IxodesTicks and Persistently Infect Human Brain Endothelial Cells and Pericytes

Powassan viruses (POWVs) are neurovirulent tick-borne flaviviruses emerging in the Northeastern U.S., with a 2% prevalence in Long Island (LI) deer ticks (Ixodes scapularis). POWVs are transmitted in as little as 15 minutes of a tick bite, and enter the CNS to cause encephalitis (10% fatal) and long-term neuronal damage. POWV-LI9 and POWV-LI41 present in LI Ixodes ticks were isolated by directly inoculating VeroE6 cells with tick homogenates and detecting POWV infected cells by immunoperoxidase staining. Inoculated POWV-LI9 and LI41 were exclusively present in infected cell foci, indicative of spread cell to cell, despite growth in liquid culture without an overlay. Cloning and sequencing establish POWV-LI9 as a phylogenetically distinct lineage II POWV strain circulating in LI deer ticks. Primary human brain microvascular endothelial cells (hBMECs) and pericytes form a neurovascular complex that restricts entry into the CNS. We found that POWV-LI9, -LI41 and Lineage I POWV-LB, productively infect hBMECs and pericytes and that POWVs were basolaterally transmitted from hBMECs to lower chamber pericytes without permeabilizing polarized hBMECs. Synchronous POWV-LI9 infection of hBMECs and pericytes induced proinflammatory chemokines, interferon-{beta} (IFN{beta}) and IFN-stimulated genes, with delayed IFN{beta} secretion by infected pericytes. IFN inhibited POWV infection, but despite IFN secretion a subset of POWV infected hBMECs and pericytes remained persistently infected. These findings suggest a potential mechanism for POWVs (LI9/LI41 and LB) to infect hBMECs, spread basolaterally to pericytes and enter the CNS. hBMEC and pericyte responses to POWV infection suggest a role for immunopathology in POWV neurovirulence and potential therapeutic targets for preventing POWV spread to neuronal compartments. ImportanceWe isolated POWVs from LI deer ticks (I. scapularis) directly in VeroE6 cells and sequencing revealed POWV-LI9 as a distinct lineage II POWV strain. Remarkably, inoculating VeroE6 cells with POWV containing tick homogenates resulted in infected cell foci in liquid culture, consistent with cell to cell spread. POWV-LI9, -LI41, and Lineage I POWV-LB strains infected hBMECs and pericytes that comprise neurovascular complexes. POWVs were nonlytically transmitted basolaterally from infected hBMECs to lower chamber pericytes, suggesting a mechanism for POWV transmission across BBB. POWV-LI9 elicited inflammatory responses from infected hBMEC and pericytes that may contribute to immune cell recruitment and neuropathogenesis. This study reveals a potential mechanism for POWVs to enter the CNS by infecting hBMECs and spreading basolaterally to abluminal pericytes. Our findings reveal that POWV-LI9 persists in cells that form a neurovascular complex spanning the BBB, and suggest potential therapeutic targets for preventing POWV spread to neuronal compartments.

microbiology↗