Phenylbutyrate restricts murine β-coronavirus infectivity and limits virus-induced demyelination <i>in vivo</i>

High morbidity and mortality associated with human β-coronavirus (CoV) infection highlight the need to determine host responses to infection and develop anti-viral therapies. Gap junction intercellular communication (GJIC), particularly involving Connexin43 (Cx43), is vital for maintaining central nervous system (CNS) homeostasis, and disruption of GJIC is a well-documented pathogenic mechanism among β-coronaviruses. Specifically, murine β-coronavirus, mouse hepatitis virus (MHV-A59) inoculation in the mouse brain causes acute-stage CNS viral spread and chronic neuroinflammatory demyelination while causing pronounced downregulation of Cx43 at the acute stage, reflecting a critical role in CNS pathology. To pharmacologically target Cx43, 4-phenylbutyric acid (4-PBA) has been widely used, since it enhances the expression of ER-resident thioredoxin family protein (ERp29), a molecular chaperone of Cx43. 4-PBA has been shown to have anti-viral effects against MHV-A59 <i>in vitro</i>; however, the <i>in vivo</i> efficacy of 4-PBA is unknown. This study investigates the effect of 4-PBA administration on the pathology of MHV-A59 infection <i>in vivo</i>. We demonstrate that 4-PBA treatment reduced acute MHV-A59 infectivity and viral spread in the brain while modulating the glial cell response and mounting host immunity. MHV-A59 infection downregulated the expression of ERp29 in the CNS, which was rescued by 4-PBA treatment. Furthermore, treatment with 4-PBA effectively preserved the expression of Cx43 and Cx47 in infected CNS cells, counteracting their infection-induced downregulation. 4-PBA treatment not only limits acute viral replication and spread throughout the brain but also protects against severe chronic virus-induced neuroinflammatory demyelination <i>in vivo</i>. These findings highlight 4-PBA's significant anti-viral potential against murine β-CoV and therapeutic potential in mitigating virus-induced neuroinflammatory demyelination.IMPORTANCEPast outbreaks and the emergence of novel coronaviruses pose a serious global health threat, warranting studies on the disease mechanism of these viruses and the development of new anti-viral strategies. In the current study, we demonstrated the antiviral potential of 4-phenylbutyric acid (4-PBA) against a neurotropic murine β-coronavirus, mouse hepatitis virus (MHV-A59). MHV-A59 inoculation in the brain causes virus infection and disruption of gap junction (GJ) communication by the downregulation of GJ proteins connexin 43 (Cx43) and connexin 47 (Cx47), crucial for maintaining CNS homeostasis. We demonstrate that 4-PBA restricts viral spread and infectivity in the mouse brain and improves the reduced levels of ERp29 and, thus, Cx43 and Cx47 in the infected CNS. Furthermore, 4-PBA mitigated virus-induced chronic neuroinflammatory demyelination, the characteristic feature of multiple sclerosis (MS). These findings demonstrate that 4-PBA holds significant therapeutic potential for restricting β-CoV spread and virus-induced neuroinflammatory demyelination.