Role of extracellular vesicles in HSV-1-driven brain damage

Extracellular vesicles (EVs) are involved in cell-to cell communication and spreading of toxic proteins, miRNA as well as used by viruses to pass from cell to another one. Microvesicles shed from plasma membrane whereas exosomes are smaller vesicles, generated from multivesicular bodies and released by neurons and glial cells. A growing body of evidence supports the role of EVs in transferring neurotoxic proteins such as A? and Tau, key players in Alzheimer's disease (AD) pathogenesis. Recent studies reported that aggregated forms of Tau can be transmitted among neurons and that exogenous aggregates of Tau could enter inside cells acting as seeds for the aggregation of the endogenous protein, thus propagating Tau-dependent damages in the brain. Several data support the role of Herpes Simplex Virus-1 (HSV-1) infection in AD pathogenesis. We recently demonstrated the effect of multiple viral reactivations on the accumulation of Tau-related brain damages in an in vivo mouse model. Interestingly, brain slices from HSV-1 infected animals showed higher number of phosphorylated Tau (pTau) positive cells with respect to virus positive one. Here, we investigated whether HSV-1 infection in the brain could promote Tau spreading among neurons via EVs, as well as its own spreading. To this aim, EVs were isolated from supernatants of human neuroblastoma and primary cultures of rat neurons following 24-48h of HSV-1- or Mock-infection, analysed in western blotting (WB) or incubated on uninfected cells for 24h following UV-treatment. Cell lysates were then analysed in WB for pTau content and compared to untreated cells. Results showed that EVs derived from HSV-1-infected cells contained: a) viral proteins, suggesting that the virus exploits them for its spreading among cells; b) increased levels of pTau, indicating that the virus can promote pTau propagation among neurons via exosomes. Accordingly, cells incubated with EVs isolated from HSV-1- infected cells showed: a) the occurrence of HSV-1 productive infection, as visualized by the use of fluorescent recombinant HSV-1 virus; b) higher levels of pTau with respect to those detected in untreated cells, indicating that exosomes derived from infected cells can promote Tau aggregation. Overall, these data indicate that the virus can promote pTau propagation among neurons via EVs, as well as its own spreading, and support the hypothesis that repeated HSV-1 reactivations into the brain may concur to neurodegeneration.