Objective HIV-associated CNS dysfunction is a significant problem among people with HIV (PWH), who now live longer due to viral suppression from combined anti-retroviral therapy (ART). Over the course of infection, HIV generates toxic viral proteins and induces inflammatory cytokines that have toxic effects on neurons in the CNS. Among these viral proteins, HIV Nef has been found in neurons of postmortem brain specimens from PWH. However, the source of Nef and its impact on neuronal cell homeostasis are still elusive. Methods and results Here, in using a simian immunodeficiency virus (SIV) infected rhesus macaque model of neuroHIV, we find SIV Nef reactivity in the frontal cortex, hippocampus and cerebellum of SIV-infected animals using immunohistochemistry (IHC). Interestingly, SIV-infected macaques treated with ART also showed frequent Nef positive cells in the cerebellum and hippocampus. Using dual quantitative RNAscope and IHC, we observed cells that were positive for Nef, but were not for SIV RNA, suggesting that Nef protein is present in cells that are not actively infected with SIV. Using cell specific markers, we observed Nef protein in microglia/macrophages and astrocytes. Importantly, we also identified a number of NeuN-positive neurons, which are not permissive to SIV infection, but contained Nef protein. Further characterization of Nef-positive neurons showed caspase 3 activation, indicating late stage apoptosis in the CNS neurons. Conclusions Our results suggest that regardless of ART status, Nef is expressed in the brain of SIV infected macaques and may contribute to neurological complications seen in PWH.
Models of macrophage subtypes require molecular characterization to reliably facilitate their differentiation. Although CD16 + (Fc-gamma III receptor) monocytes that express CD163 (a hemoglobin/haptoglobin receptor) have been implicated in a variety of disease states, the conditions necessary to establish lineages of these cell subtypes remains unknown. The current investigations utilize a cell line derived from acute myelogenous leukemia lineage, MonoMac-1, to interrogate the factors that promote the development of CD16 + macrophages that express CD163. Results implicate the glucocorticoid pathway as well as c-fms signaling based on the action of dexamethasone and macrophage colony-stimulating factor-1 in promoting CD16 + expression, in addition to phorbol myristate acetate and lipopolysaccharides treatment. The ability of glucocorticoid and c-fms receptor antagonists to inhibit CD16 + cell formation further establishes the role of these pathways in CD16 expression in this cell line. In view of the inherent difficulty in working with primary cells as well as donor variation, cell lines may be preferable to primary cells for their consistency. We envision that the process we use to induce CD16 expression in this cell type will be useful for screening and identification of drug candidates potentially useful for the treatment of diseases where the etiology involves the expansion of the CD16 + monocytes subset or the accumulation of CD163 + tissue macrophages.
Objective: Antiretroviral therapy (ART) extends the life of people with HIV (PWH), but these individuals are at increased risk for obesity, dyslipidemia, diabetes, and cardiovascular disease. These comorbidities may be a consequence of HIV-related chronic inflammation and/or adverse effects of ART on tissue regulatory adipose tissue macrophages (ATMs). We sought to determine the effects of HIV/ART on metabolically beneficial ATM populations and functions.Design: We examined subcutaneous ATMs from PWH on integrase inhibitor-containing ART (n ¼ 5) and uninfected persons (n ¼ 9). We complemented these studies with ex vivo and in vitro analyses of peripheral blood mononuclear cell (PBMC) and murine macrophage lipid metabolism and fatty acid oxidation gene expression.Methods: ATM populations were examined by flow cytometry. Macrophage lipid metabolism and fatty acid oxidation gene expression were examined by Seahorse assay and quantitative PCR.Results: Adipose tissue from PWH had reduced populations of metabolically activated CD9 þ ATMs compared to that of uninfected controls (P < 0.001). PBMCs of PWH had lower fatty acid metabolism compared to those of uninfected controls (P < 0.01). Analysis of murine macrophages revealed that dolutegravir reduced lipid metabolism (P < 0.001) and increased expression of the fatty acid beta-oxidation enzyme enoyl-CoA hydratase, short chain 1 (P < 0.05). Conclusions:We report the loss of metabolically beneficial ATM populations in PWH on ART, altered fatty acid metabolism of blood immune cells, and evidence that dolutegravir alters macrophage fatty acid metabolism. Future studies should examine direct or indirect effects and mechanisms of dolutegravir, and other integrase inhibitors and ART classes, on fatty acid beta-oxidation.
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