Human Neural Stem Cell-Derived Neuron/Astrocyte Co-Cultures as a Model System to Study Emerging Encephalitic Viruses

Many emerging viral infections target the central nervous system. Such diseases often cause high fatality rates and high rates of neurologic sequelae such as behavioral changes, cognitive deficits, seizures, paralysis, or persistent infection. Relatively little is known about viral infections of cells within the central nervous system such as neurons and astrocytes. One challenge in studying central nervous system infection is the lack of suitable model systems to study infection in neurons and astrocytes. Animal based systems often require immunocompromise and may not be suitable for molecular mechanistic studies. In vitro systems often rely on cell lines of cancerous origin, which often have specific signaling defects that may impact cellular responses to infection. This study describes the establishment of a human neural stem cell derived neuron/astrocyte co-cultures system as a model to study encephalitic virus infection. This system provides a more physiologically relevant model that maintains critical interactions between neurons and astrocytes. Two emerging encephalitic viruses were studied using this system, La Crosse virus (LACV) and Nipah virus (NiV). LACV causes pediatric encephalitis in the United States, while NiV causes severe encephalitic and respiratory disease in Southern Asia. Neuron/astrocyte co-cultures were susceptible to LACV infection and displayed apoptotic responses as reported in previous in vitro and in vivo studies. Neurons and astrocytes were both targets of LACV infection, with neurons becoming the predominant target later in infection possibly due to astrocytic responses to interferon. Additionally, neuron/astrocyte co-cultures responded to LACV infection with strong proinflammatory cytokines and chemokines. In contrast, NiV replicated in both neurons and astrocytes, inhibited inflammatory cytokine/chemokine production, and produced delayed interferon-β responses. Interferon-β was shown to significantly reduce viral titer, apoptosis, and cytotoxicity. Interferon-β preferentially protected astrocytes from NiV infection. Additionally, while the type I interferon response reduced NiV infection, it was not able to fully eliminate it. These results suggest that the type I interferon response may play a role in establishing a persistent NiV infection. Together, these studies demonstrate the ability of this neuron/astrocyte system to respond to viral infection in a manner consistent with in vivo observations, and its usefulness in studying encephalitis pathogenesis.