Background
Neurologic consequences of HIV linger even after viral suppression, suggesting a molecular “scar” that persists despite antiretroviral therapy (ART). However, whether innate immune cells in the cerebrospinal fluid (CSF) are altered during HIV and how they contribute to neuropathogenesis are poorly understood, owing in part to difficulties in isolating these rare populations of cells before the advent of single cell technologies.
Methods
We enrolled 12 neurologically asymptomatic people with HIV (PWH) on suppressive ART and 11 demographically matched people without HIV (PWoH) for cross-sectional CSF single immune cell epigenome and transcriptome analyses. We utilized the 10X Genomics single nucleus multiome platform (ATAC-Seq + RNA-Seq), modified for low-cellular CSF, for 6 PWH and 5 PWoH (33,353 total nuclei). ATAC-seq data was used to identify open chromatin regions (peaks). We validated the single-nucleus transcriptomic data with additional data previously collected utilizing 10X 5’ single cell RNA-Seq from CSF of 7 PWH also on ART with features similar to the multiome participants and 6 PWoH (102,313 total cells).
Results
CSF of both PWH and PWoH consisted primarily of T cells (median=89.5%; IQR=6.2%), and secondarily of myeloid cells (median=5.9%; IQR=6.5%), including a small percentage of dendritic cells (DCs) (median=2.4%; IQR=1.7%). We identified 555 differentially accessible chromatin regions (DARs) across all cell types between PWH and PWoH. Despite comprising only a minority of CSF cells, DCs contained 32.8% of DARs, most of which were more open (more accessible) in PWH (130/182) (Fig. 1A). By annotating these DARs of DCs to their nearest genes, we identified 145 genes, including interferon-related genes IFI6 and GBP4, both of which exhibit differential RNA expression between PWH and PWoH in DCs. In our validation dataset of 5’ single cell RNA-Seq CSF transcriptomes, we observed a DC transcriptomic signature in CSF of PWH compared to PWoH (Fig.1B). Pathway analysis identified downregulation of antigen processing and presentation, and cytokine (mainly IL-1 and IL-12) signaling in DCs in PWH. Lastly, we detected extremely rare HIV RNA and chromatin-accessible provirus DNA in CSF cells.
Conclusions
Single-cell epigenome and transcriptome profiling of CSF suggests dysregulation of DCs in virally suppressed PWH. Further studies are needed to determine if these changes impair antigen presentation and antiviral responses, contributing to HIV persistence and neuropathogenesis.