In situ spatial characterization of CNS HIV-host cellular interactions from victims of sudden death

SUMMARY/ABSTRACT In people with HIV (PWH) the central nervous system (CNS) harbors an HIV reservoir that may lead to ongoing inflammation and tissue damage. PWH also have high rates of methamphetamine use which may act in synergy with HIV persistence in the CNS to increase myeloid immune inflammasome activation, astrocyte dysfunction, and neuroinflammation. However, the extent to which HIV-infected cells persist in the CNS in the setting of ART and interact with local tissue environments that mediate inflammation with or without methamphetamine use in vivo is largely unknown. Direct access to CNS tissue in healthy PWH on ART is nearly impossible, yet in-depth spatial viral-host characterization within brain from PWH on ART with minimal comorbidities is crucially needed to facilitate research on HIV-mediated neuroinflammation in the setting of drug use. We established the longitudinal POstmortem Systematic InvesTigation of Sudden Cardiac Death (POST SCD) Study, a prospective, countywide postmortem study to bank extensive samples and autopsy data on all victims of sudden death in San Francisco County, including PWH. To date we have collected extensive samples including brain from >85 PWH, ~80% on ART, and >700 uninfected individuals without major underlying morbidity at the time of their unexpected out-of-hospital sudden death. As a result, the HIV POST SCD cohort is a one-of-kind resource for the in-situ study HIV infection in the CNS in the setting of controlled HIV disease. Well-curated brain tissue from multiple locations from PWH on ART have been collected and we have generated preliminary directly in these tissues using high-dimensional spatial genomics demonstrating that myeloid cells harboring HIV transcripts have unique genomic and transcriptomic phenotypes. We plan to leverage the highly unique and comprehensive POST SCD cohort to provide unparalleled insight into the nature and extent of HIV and methamphetamine use in the brain and host-immune factors that facilitate inflammation and brain injury. In the R61 phase, we will develop and implement innovative single-cell immunohistochemical, in situ hybridization, digital spatial gene expression, and quantitative drug assays. In the R33 phase these analytical frameworks will be applied to brain tissue from PWH on ART and matched HIV- sudden death cases with and without methamphetamine use. Our aims are to (1) implement innovative spatial multi-omic assays that combine analysis of the localization, distribution, and transcriptional and translational activity of HIV-infected cells and cellular host responses in multiple brain regions from PWH that died suddenly out of hospital with minimal comorbidities; (2) test the hypothesis that in-situ immunohistochemistry can be used in combination with high-parameter spatial profiling to quantify methamphetamine and ART levels in-situ and to determine the impact of drug levels on HIV-host responses; and (3) test the hypothesis that methamphetamine use acts in synergy with HIV to enhance myeloid immune cell inflammatory signaling and altered astrocyte function leading to neuroinflammation and neural injury.