Macrophage-derived microparticles facilitate the dissemination and spread of Japanese encephalitis virus and offer targets for antiviral intervention.

Japanese encephalitis virus (JEV) is thought to spread systemically; however, its transport and propagation mechanisms remain unclear. This study examined JEV progression through analysis of macrophage response and macrophage-derived microparticles (MPs) dynamics. MPs were isolated from conditioned media of macrophages at 72 h post-infection to assess their role in JEV transmission. Flow cytometry, qRT-PCR and western blotting were performed to analyze the MPs' JEV content and protein profiles. To further determine the impact of MPs on viral spread, MPs were neutralized and cocultured with uninfected macrophages. JEV infection was also mapped in the brain and lymph nodes at 24 h and 7 days post-infection to investigate early viral movement via macrophage-derived MPs. The findings indicate macrophage-derived MPs facilitate JEV transit from the brain to the lymph nodes. JEV infection of macrophages triggers apoptosis, increased viral replication, and a proinflammatory response, leading to the heightened secretion of MPs. These MPs carried JEV RNA, cytokines, and proteins such as HSP90 and flotillin-1. MPs from infected macrophages promoted viral spread to uninfected macrophages, initiating viremia. Neutralizing MPs with annexin V antibodies or heat inactivation significantly reduced JEV RNA levels in cocultures. These results suggest macrophage-derived MPs play a key role in JEV dissemination and presenting potential targets for antiviral intervention.