The microvascular endothelium of the blood-brain barrier is highly restrictive to JC Polyomavirus neuroinvasion.

JC Polyomavirus is the causative agent of progressive multifocal leukoencephalopathy (PML), an often-fatal demyelinating disease. Unfortunately, a diagnosis of PML occurs only after patients have suffered irreversible neuropathologies. The first step in the initiation of PML is viral entry to the brain, but the route and mechanisms responsible for neuroinvasion have not been well established. To gain a better understanding of this, we asked whether purified virus or virus associated with extracellular vesicles (EVs) could penetrate two different cell culture models of the blood-brain barrier. In one model, we used the hCMEC/D3 brain endothelial cell line, and in the other, we used pluripotent stem cells induced to a brain endothelial cell phenotype (iPSC-EC). We found that neither cell type was permissive to viral infection, but the virus bound and was internalized by both in a sialic acid-dependent manner. Despite virus internalization into these cells, very few virions or virus-associated extracellular vesicles (virus-EVs) penetrated the barriers. The small amount of virus or virus-EVs that did pass through either barrier was sufficient to establish infection in human glial cells. Our findings demonstrate that limited amounts of infectious virions and virus-associated EVs can traverse the brain microvascular endothelium and establish infection.IMPORTANCEThe human polyomavirus, JC Polyomavirus (JCPyV), causes a rapidly progressing demyelinating disease in immunocompromised or immunomodulated patients. Demyelinating lesions are often seen surrounding blood vessels in the brain. In this paper, we used two models to recapitulate a minimal blood-brain barrier and found that both were highly restrictive of virus penetration. A small amount of virus succeeded in crossing both barriers and was sufficient to establish infection of human glia. These data have direct implications for mechanisms used by JCPyV to invade the CNS and cause neurological disease.