Impact of EBV infection on cortical neurodevelopment

The Epstein-Barr Virus (EBV) is a DNA virus belonging to the Herpesvirus family. EBV infections are extremely common, and linked to various diseases. Recent studies have highlighted the role of herpesviruses in the central nervous system (CNS) and some reports suggest a possible link between EBV infection and Rasmussen’s encephalitis. This is a progressive neurological disease that primarily affects children between the ages of 6 and 10 and it suggests a potential involvement of early-stage EBV infection. Due to EBV's inability to infect rodent cells lacking the necessary receptors, iPSC-derived cortical cultures provide a unique opportunity to study the interactions between EBV and human brain cells across neurodevelopmental stages. To this end, 2D cell cultures were infected at a neurodevelopmental stage in which mature neurons and neural progenitors coexist. Real-time PCR analysis confirmed that EBV successfully infected cortical cultures, detecting latent and lytic transcripts, including BZLF1 LMP1 and GP220. Strikingly, infected cultures displayed higher levels of the neural progenitor marker PAX6, suggesting an alteration in neuronal differentiation. Focusing on an earlier neurodevelopmental stage, we observed an increase in LAMP1 and CLEAVED CASPASE 3, associated with an abnormal structuring of neural rosettes after EBV infection. Typically, neural rosettes recapitulate neural tube closure with a well-defined organization, however, EBV infection profoundly disrupted the morphology of neural rosettes in 2D cultures leading to chaotic formations. To further investigate this phenomenon, we employed a 3D micropatterned system (RosetteArray technology) that forces cell aggregation into 3D neural rosettes. Across three different control cell lines, we observed that EBV infection impaired neural rosette formation, leading to a reduced number of rosettes with significantly smaller areas supporting the idea that EBV infection impacts neurodevelopment. To further explore its effects on neuronal networks, we will analyze morphological, molecular, and functional changes using multielectrode array recordings and calcium imaging.