Neonatal neural progenitor cells and their neuronal and glial cell derivatives are fully permissive for human cytomegalovirus infection

Congenital human cytomegalovirus (HCMV) infection causes central nervous system structural abnormalities and functional disorders, affecting both astroglia and neurons with a pathogenesis that is only marginally understood. To better understand HCMV's interactions with such clinically important cell types, we utilized neural progenitor cells (NPCs) derived from neonatal autopsy tissue, which can be differentiated down either glial or neuronal pathways. Studies were performed using two viral isolates, Towne (laboratory adapted) and TR (a clinical strain), at a multiplicity of infection of 3. NPCs were fully permissive for both strains, expressing the full range of viral antigens (Ags) and producing relatively large numbers of infectious virions. NPCs infected with TR showed delayed development of cytopathic effects (CPE) and replication centers and shed less virus. This pattern of delay for TR infections held true for all cell types tested. Differentiation of NPCs was carried out for 21 days to obtain either astroglia (>95% GFAP⁺) or a 1:5 mixed neuron/astroglia population (β-tubulin III⁺/GFAP⁺). We found that both of these differentiated populations were fully permissive for HCMV infection and produced substantial numbers of infectious virions. Utilizing a difference in plating efficiencies, we were able to enrich the neuron population to ~80% β-tubulin III⁺ cells. These β-tubulin III⁺-enriched populations remained fully permissive for infection but were very slow to develop CPE. These infected enriched neurons survived longer than either NPCs or astroglia, and a small proportion were alive until at least 14 days postinfection. These surviving cells were all β-tubulin III⁺ and showed viral Ag expression. Surprisingly, some cells still exhibited extended processes, similar to mock-infected neurons. Our findings strongly suggest neurons as reservoirs for HCMV within the developing brain.