Neurological complications are common in COVID-19. Although SARS-CoV-2 has been detected in patients' brain tissues, its entry routes and resulting consequences are not well understood. Here, we show a pronounced upregulation of interferon signaling pathways of the neurovascular unit in fatal COVID-19. By investigating the susceptibility of human induced pluripotent stem cell (hiPSC)-derived brain capillary endothelial-like cells (BCECs) to SARS-CoV-2 infection, we found that BCECs were infected and recapitulated transcriptional changes detected in vivo. While BCECs were not compromised in their paracellular tightness, we found SARS-CoV-2 in the basolateral compartment in transwell assays after apical infection, suggesting active replication and transcellular transport of virus across the blood-brain barrier (BBB) in vitro. Moreover, entry of SARS-CoV-2 into BCECs could be reduced by anti-spike-, anti-angiotensin-converting enzyme 2 (ACE2)-, and anti-neuropilin-1 (NRP1)-specific antibodies or the transmembrane protease serine subtype 2 (TMPRSS2) inhibitor nafamostat. Together, our data provide strong support for SARS-CoV-2 brain entry across the BBB resulting in increased interferon signaling.

Background Genome-wide DNA methylation profiling has recently been developed into a tool that allows tumor classification in central nervous system tumors. Extracellular vesicles (EVs) are released by tumor cells and contain high molecular weight DNA, rendering EVs a potential biomarker source to identify tumor subgroups, stratify patients and monitor therapy by liquid biopsy. We investigated whether the DNA in glioblastoma cell-derived EVs reflects genome-wide tumor methylation and mutational profiles and allows non-invasive tumor subtype classification. Methods DNA was isolated from EVs secreted by glioblastoma cells as well as from matching cultured cells and tumors. EV-DNA was localized and quantified by direct stochastic optical reconstruction microscopy. Methylation and copy number profiling was performed using 850k arrays. Mutations were identified by targeted gene panel sequencing. Proteins were differentially quantified by mass spectrometric proteomics. Results Genome-wide methylation profiling of glioblastoma-derived EVs correctly identified the methylation class of the parental cells and original tumors, including the MGMT promoter methylation status. Tumor-specific mutations and copy number variations (CNV) were detected in EV-DNA with high accuracy. Different EV isolation techniques did not affect the methylation profiling and CNV results. DNA was present inside EVs and on the EV surface. Proteome analysis did not allow specific tumor identification or classification but identified tumor-associated proteins that could potentially be useful for enriching tumor-derived circulating EVs from biofluids. Conclusions This study provides proof of principle that EV-DNA reflects the genome-wide methylation, CNV and mutational status of glioblastoma cells and enables their molecular classification.

To date the exact neuronal implementation of decision confidence has been subject to little research. Here we explore electroencephalographic correlates of human choice certainty in a visual motion discrimination task for either spatial attention or motor effector cue instructions. We demonstrate electrophysiological correlates of choice certainty that evolve as early as 300 ms after stimulus onset and resemble the primary visual motion representations in early visual cortex. These correlates do not emerge unless or until the subject unambiguously knows which of the competing visual stimuli is actually relevant to behaviour. They extend beyond stimulus presentation up to the motor response but are independent of the motor effector. Our findings suggest that perceptual confidence evolves in parallel with representations of stimulus properties and is dedicated to one specific aspect of the visual world. Its electroencephalographic correlates can be disentangled from representations of sensory evidence, objective discrimination performance and overt motor behaviour.