Abstract
People living with HIV suffer multiple comorbid conditions related to chronic inflammation at increased rates compared to the general population, even when on effective antiretroviral therapy. In particular, current data indicate that the increased incidence and severity of neurocognitive impairment (NCI) are associated with unresolved neuroinflammation. Attempts to treat NCI in people living with HIV by reducing inflammation have thus far been unsuccessful, suggesting that a more mechanistic understanding of inflammatory processes in the CNS during HIV is necessary. Here, we use iPSC-derived microglia (iMg) and astrocytes (iAst) to model HIV infection in the CNS. We show that our iMg robustly express markers associated with microglial identity and are susceptible to HIV infection, but exhibit lower HIV replication rates and weaker immune response to HIV challenge compared to monocyte-derived macrophages. Coculture of iAst with iMg leads to a much stronger pro-inflammatory immune response, and, surprisingly, a robust increase in rates of HIV replication. Increased replication in iMg/iAst cocultures is associated with higher levels of multiple pro-inflammatory cytokines, including TNFα, which is produced by iAst upon exposure to HIV-infected iMg. Addition of exogenous TNFα to iMg during HIV infection is also sufficient to increase rates of replication, and neutralization of TNFα via adalimumab/Humira treatment in iMg/iAst cocultures reduces replication. Blocking NF-kB signaling with iKK inhibitor Bay-11-7082 (Bay-11) demonstrates that increased HIV replication in iMg/iAst cocultures is due to increased NF-kB activity. Finally, we show that in HIV-infected iMg there is movement of lysosomes to the periphery of the cell membrane and release of lysosomal content into the extracellular space, suggesting that this dysregulated lysosomal flux could further contribute to the pro-inflammatory microenvironment. We propose that this altered lysosomal trafficking and increased cytokine production drives a pro-inflammatory phenotype in glia and represents a potential source of unresolved neuroinflammation in people living with HIV.
Data availability
All sequencing data are available in the NCBI GEO database (https://www.ncbi.nlm.nih.gov/geo/) with the accession number GSE143685. Other data used and/or analyzed during the current study are available from the corresponding author on reasonable request.
Abbreviations
- HIV:
-
Human Immunodeficiency virus
- NCI:
-
Neurocognitive impairment
- HAND:
-
HIV-associated neurocognitive disorder
- HABI:
-
HIV-associated brain injury
- PLWH:
-
People living with HIV
- ART:
-
Anti-retroviral therapy
- CNS:
-
Central nervous system
- CSF:
-
Cerebrospinal fluid
- iPSCs:
-
Induced pluripotent stem cells
- iMg:
-
iPSC-derived microglia
- iAst:
-
iPSC-derived astrocytes
- CC:
-
Coculture
- CM:
-
Conditioned media
- MDM:
-
Monocyte-derived macrophages
- CMP:
-
Common myeloid progenitor
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Acknowledgments
We would like to thank all the members of the Gaskill lab and myeloid working group for invaluable discussion on the data. The authors thank Max Eldabbas, Emileigh Maddox, Tanishk Sinha, and Jiayi Shu of the Human Immunology Core at the Perelman School of Medicine at the University of Pennsylvania for assistance with isolation and differentiation of primary human monocytes. The HIC is supported in part by NIH P30 AI045008 and P30 CA016520. The following reagent was obtained through the NIH HIV Reagent Program, NIAID, NIH: Monoclonal Anti-Human Immunodeficiency Virus Type 1 (HIV-1) p24 Gag protein, Clone AG3.0 (produced in vitro), HRP-20068, contributed by Creative Biolabs.
Funding
This research was funded by the National Institutes of Health grant no. T32 (JG), F31 MH131486(JG), R01DA057337(PJG), R61DA058501 (PJG), R21MH129193 (CAE), R01DA049514 and R01DA052826 (KLJS), and P30AI045008 (KJS, CAE).
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JG, CAE, and KLJS contributed study conceptualization and methodology. JG, JP, SS, EB, JS contributed experimental design, data acquisition, and data analysis. JG contributed original draft preparation. All authors contributed to writing, review, and editing. JG, PJG, CAE, and KJS contributed funding. All authors have read and agreed to the submitted version of the manuscript.
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Gesualdi, J., Prah, J., Solomon, S. et al. Neuroinflammatory crosstalk between microglia and astrocytes increases viral replication in an iPSC-derived model of CNS HIV infection. Sci Rep (2026). https://doi.org/10.1038/s41598-026-43248-7
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DOI: https://doi.org/10.1038/s41598-026-43248-7