Speaker
Description
Objectives: Progressive multifocal leukoencephalopathy (PML) is an often-fatal demyelinating disease of the central nervous system, due to reactivation of JC virus (JCV) in the context of severe immune dysfunction. Pathophysiology of JCV infection is complex, and many open questions remain regarding how the virus accumulates genomic rearrangements to reactivate and specifically infects glia. Next generation sequencing technologies provide unprecedented tools to comprehensively survey host-pathogen interactions and create a detailed map integrating JCV intra-host activity and host immune response.
Methods: Frozen brain sections from two patients who died of PML were analyzed. Ante-mortem clinical MR imaging was used to identify four regions of interest (ROI) for transcriptomic characterization, including leukocortical junction, edge of an active PML lesion, center of a lesion, and normal appearing white matter. Sequencing reads were aligned to the annotated human and the JCV genomes.
Results: Regions of active infection have increased numbers of JCV-infected cells compared to normal appearing white matter. Most highly infected cells cluster separate from healthy cells indicating clearly distinct molecular phenotypes. Highly infected cells generally have less diverse host transcriptional profiles and are more likely classified in S- or G2M-phase, consistent with viral cooption of host cell transcriptional machinery. A minority of infected cells still cluster with healthy cells, primarily oligodendrocytes, neurons, and myeloid cells, likely depicting earlier stages of infection and phagocytosing myeloid cells. Preliminary differential gene expression analysis highlights that pathways associated with innate immunity and antigen presentation are enriched in infected compared to uninfected cells.
Conclusions: We have established an initial workflow for MRI-guided tissue sampling and single cell sequencing that can effectively capture changes in host and viral biology during JCV infection. Current challenges include interpretation of variability observed between patients and optimization of ROI selection. Next steps are to increase sample numbers and complete histopathologic validation of ROIs.
