MicroRNAs (miRNAs) are small RNAs that regulate gene expression by translational repression and/or mRNA degradation through binding to the 3'UTR of target mRNAs. There is increasing evidence indicating that miRNAs play a major role in fundamental cellular processes, the regulation of the immune system, as well as in the onset and progression of many diseases. Although the role of miRNAs in virus-host interactions is emerging, there is little evidence that viral modulation of cellular miRNAs actually impacts neuro-inflammatory responses in vivo. Unraveling the roles of miRNAs in viral encephalitis in humans is challenging. The study of animal models of viral encephalitis is key to our understanding of how viruses drive disease in the CNS. Differences in the severity of viral encephalitis may reflect the differential ability of viruses to stimulate innate immune responses within the CNS. Coronaviruses (CoVs) are enveloped, positive-sense, single-stranded, polyadenylated RNA viruses. Different murine CoVs (Mouse Hepatitis Virus, MHV) isolates induce acute lethal encephalitis and encephalomyelitis associated with acute and chronic demyelination. Here, we will compare two strains differing markedly in their neurovirulence: MHV-JHM, a highly neurovirulent strain that produces lethal encephalitis;and MHV-A59, a mildly neurovirulent strain that induces acute meningoencephalitis and chronic. In preliminary studies, we have used miRNA microarray analysis and validation by real-time RT-PCR to provide evidence of the differential expression of selected cellular miRNAs in macrophages upon in vitro infection with MHV-A59 and MHV-JHM. Changes in the macrophage miRNAs profiles after murine CoVs infection were overlapping but distinct, and were observed early after infection in the absence of type I IFNs, IFN-?, and pro-inflammatory cytokine secretion. Our data suggest that MHV infection can modify the pattern of cellular miRNA expression. We have identified two miRNAs that are significantly up-regulated by the highly neurovirulent MHV-JHM (but not by MHV-A59). Interestingly, we also found that these JHM-up-regulated miRNAs potentially target the 3'UTR of IL-1 receptor antagonist (IL-1Ra), a cytokine involved in both neuroprotection and neuropathogenesis, as well as IFN?. We hypothesize that CoVs inducing different CNS disease outcome promote distinct miRNAs expression profiles in macrophages and microglia, and that virus specific changes in miRNAs expression contribute to the differences in encephalitis outcome by modulation of the inflammatory response. We propose to: 1) systematically identify miRNAs and mRNAs regulatory networks that correlate with lethal encephalitis progression in the infected brain;2) Define the role of those JHM-up-regulated miRNAs (identified in preliminary studies and in Specific Aim 1) in IL-1Ra and IFN? induction by macrophages and glial cells ex vivo, as well as in vivo using antagomir-lentivirus vectors. Overall, there is significant translational potential in the proposed investigations since they will facilitate the development of therapeutic approaches to prevent and/or treat neuro-inflammatory diseases of viral etiology.
The relative contribution of microRNAs to virus-host interactions in the context of the CNS, and to neuroinflammatory responses and/or neuropathogenesis is poorly understood. Our experimental model will provide a distinctive opportunity to define the role of selected microRNAs in neuro-inflammatory responses that result in viral-induced lethal encephalitis, and could potentially lead to novel therapeutic approaches to prevent and/or treat acute neuro-inflammatory diseases of viral etiology.
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