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Mouse Footpad Inoculation Model to Study Viral-Induced Neuroinflammatory Responses
Published on: June 14, 2020
Complexity of the microglial activation pathways that drive innate host responses during lethal alphavirus
Nilufer Esen1, Pennelope K Blakely, Emily K Rainey-Barger
1Department of Neurology, University of Michigan Medical School, Ann Arbor, MI, USA.
Abstract:
Microglia express multiple TLRs (Toll-like receptors) and provide important host defence against viruses that invade the CNS (central nervous system). Although prior studies show these cells become activated during experimental alphavirus encephalitis in mice to generate cytokines and chemokines that influence virus replication, tissue inflammation and neuronal survival, the specific PRRs (pattern recognition receptors) and signalling intermediates controlling microglial activation in this setting remain unknown. To investigate these questions directly in vivo, mice ablated of specific TLR signalling molecules were challenged with NSV (neuroadapted Sindbis virus) and CNS viral titres, inflammatory responses and clinical outcomes followed over time. To approach this problem specifically in microglia, the effects of NSV on primary cells derived from the brains of wild-type and mutant animals were characterized in vitro. From the standpoint of the virus, microglial activation required viral uncoating and an intact viral genome; inactivated virus particles did not elicit measurable microglial responses. At the level of the target cell, NSV triggered multiple PRRs in microglia to produce a broad range of inflammatory mediators via non-overlapping signalling pathways. In vivo, disease survival was surprisingly independent of TLR-driven responses, but still required production of type-I IFN (interferon) to control CNS virus replication. Interestingly, the ER (endoplasmic reticulum) protein UNC93b1 facilitated host survival independent of its known effects on endosomal TLR signalling. Taken together, these data show that alphaviruses activate microglia via multiple PRRs, highlighting the complexity of the signalling networks by which CNS host responses are elicited by these infections.
Insights
Microglia activate via multiple pattern recognition receptors during alphavirus infections. Host survival depends on type-I interferon, not Toll-like receptor signaling, and involves endoplasmic reticulum protein UNC93b1.
Area of Science:
- Neuroimmunology
- Virology
- Infectious Disease
Background:
- Microglia are key immune cells in the central nervous system (CNS) that express Toll-like receptors (TLRs) and defend against viral infections.
- Previous research indicates microglial activation during alphavirus encephalitis influences inflammation and neuronal survival, but specific pattern recognition receptors (PRRs) and signaling pathways remain unclear.
Purpose of the Study:
- To identify the specific PRRs and signaling intermediates that control microglial activation during neuroadapted Sindbis virus (NSV) infection in vivo and in vitro.
- To elucidate the roles of TLR signaling and other host factors in controlling viral replication and host survival during alphavirus encephalitis.
Main Methods:
- Mice lacking specific TLR signaling molecules were infected with NSV, and CNS viral loads, inflammatory markers, and clinical outcomes were monitored.
- Primary microglia from wild-type and mutant mice were infected with NSV in vitro to characterize PRR activation and mediator production.
- Viral uncoating and genome integrity were assessed for their necessity in triggering microglial activation.
Main Results:
- Microglial activation by NSV requires viral uncoating and an intact viral genome; inactivated virus did not induce activation.
- NSV triggered multiple PRRs in microglia, leading to diverse inflammatory mediator production through distinct signaling pathways.
- In vivo, host survival was independent of TLR signaling but dependent on type-I interferon production for controlling CNS viral replication.
- The endoplasmic reticulum protein UNC93b1 promoted host survival independently of its known role in endosomal TLR signaling.
Conclusions:
- Alphaviruses activate microglia through a complex network of multiple PRRs.
- Type-I interferon and UNC93b1 play critical roles in host defense against alphavirus CNS infections, highlighting intricate neuroimmune responses.
