Opioid-induced transcriptional reprogramming of cerebrospinal fluid immune cells drives neuroinflammation in SIV-infected rhesus macaques

Background. Opioid use is disproportionately high among People with HIV (PWH). Although combined anti-retroviral therapy (ART) can dampen HIV-associated dementia, a large fraction of PWH continue to experience neurocognitive deficits which are further exacerbated by opioid use. In the present study, we performed single cell transcriptomic profiling of cerebrospinal fluid (CSF) immune cells to explore their functional characteristics in opioid mediated neurological disorders among PWH using the SIV/rhesus macque model. Methods. In this study, we utilized CSF cells from morphine- and saline-administered, SIV-infected, ART-treated rhesus macaques (RMs). The CSF scRNA-Seq was performed longitudinally at baseline, post ramp-up with morphine (pre-infection), during acute infection, and after suppression of viremia to profile cell-specific transcriptomic signatures that mirror the CNS pathogenesis observed in opioid-dependent PWH. Results. We observed the presence of all major immune cells in CSF, including CD4 + T _CM cells, CD4 + T _EM cells, CD8+ naïve T cells, CD8 + T _CM cells, CD8 + T _EM cells, CD14 + Monocytes, CD16 + Monocytes, NK cells, and B cells. Additionally, we also demonstrated morphine-mediated relative increase in CD4 + T _CM , T _reg, and a reduction in CD8 + T _EM cell population prior to SIV infection. Chronic use of morphine was associated with a Th1/Th2 T-cell imbalance with a dominance of the Th2 population. In CSF cells from morphine-dependent RMs, there was dysregulation of genes involved in T-cell receptor signaling pathways, apoptosis, PI3K-Akt signaling pathway, cellular senescence, oxidative phosphorylation, and multiple neurodegenerative disorders. The contribution of different cell populations in these processes evolved along with different stages of disease pathogenesis. In the chronic stage of the disease, the expression of disease-associated microglia (DAM) signature genes were significantly upregulated in monocytes. Further, cell-cell receptor-ligand interaction analysis revealed an altered number of intercellular interactions and signaling strength in morphine vs. saline-administered animals. Specifically, for the CD14 + monocyte populations, the intra/inter-cell communication involving ligand-receptor pairs, including APOE-TREM2, APP(TREM2 + TYROBP), APP-CD74, SPP1−(ITGA4 + ITGB1), and CCL signaling pathways, remains significantly altered in the morphine-dependent macaques. Conclusion. Chronic opioid exposure reprograms CSF monocytes toward a DAM state that persists despite ART-mediated viral suppression, driving maladaptive immune–glial crosstalk and progressive neurocognitive dysfunction in morphine-dependent macaques with possible implications for neuroinflammation and neurodegenerative disorders that are observed in PWH.