Humanized Bone Marrow-Liver-Thymus Mice for Studying HIV-1 Persistence in Liver and Lung CD4+ T and Myeloid Cell Subsets during Antiretroviral Therapy

Cattin, A.; Pham, T. N.; Dias, J.; Do Rosario, N. F.; Marchand, L. R.; Volodina, O.; Guimond, J.-V.; Patey, N.; Li, Y.; Beland, K.; Jenabian, M.-A.; Estaquier, J.; Haddad, E.; Cohen, E. A.; Ancuta, P.

doi:10.1101/2025.01.31.635552

bioRxiv · 10.1101/2025.01.31.635552

Humanized Bone Marrow-Liver-Thymus Mice for Studying HIV-1 Persistence in Liver and Lung CD4+ T and Myeloid Cell Subsets during Antiretroviral Therapy

Cattin, A.·Pham, T. N.·Dias, J.·Do Rosario, N. F.·Marchand, L. R.·Volodina, O.·Guimond, J.-V.·Patey, N.·Li, Y.·Beland, K.·Jenabian, M.-A.·Estaquier, J.·Haddad, E.·Cohen, E. A.·Ancuta, P.

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Abstract

BackgroundWhile the role of CD4+ T-cells in HIV-1 reservoir persistence during antiretroviral therapy (ART) is well-established, studies on tissue-resident macrophages (M{Phi}) in people with HIV-1 (PWH) are restricted by difficulties in accessing deep tissue samples. Investigations in myeloid-only humanized mouse models demonstrated the contribution of M{Phi} to viral rebound upon ART interruption. Two distinct M{Phi} subsets exist in mice and humans: one of embryonic origin and self-renewal capacity generating long-lived tissue-resident M{Phi} (LL-TRM), and another one of short-lived M{Phi} (SL-M{Phi}), constantly replenished by bone marrow monocytes. The relative contribution of LL-TRM versus SL-M{Phi} to tissue HIV-1 reservoir persistence during ART remains understudied. Here, we used a humanized BM-liver-thymus (hu-BLT) mouse model to quantify integrative HIV-1 infection in liver/lung M{Phi} versus CD4+ T-cells before/after ART and document their expression of LL-TRM/SL-M{Phi} markers. ResultsLung/liver immune cells were extracted from ART-naive (ART-) and ART-treated (ART+) HIV-infected (HIV+) hu-BLT mice, as well as HIV-uninfected mice (HIV-). M{Phi} were identified as cells expressing the myeloid markers CD33/HLA-DR and/or CD68 and flow-cytometry sorted based on their differential expression of CD14 and/or CCR2. Matched CD3+CD4+ T-cells were sorted in parallel and used as controls. HIV-DNA integration was measured by nested real-time PCR. In contrast to CD4+ T-cells that carried the highest levels of proviral HIV-DNA before and after ART, integrative infection in liver/lung M{Phi} was detected before ART, but was drastically reduced in HIV+ART+ hu-BLT mice, regardless of CD14 or CCR2 expression on M{Phi}. Markers of LL-TRM (CD163/CX3CR1/Ki67/c-Kit) were expressed on a small fraction of liver but not lung M{Phi}, indicative of a deficient LL-TRM development in this hu-BLT model. ConclusionsTogether our results demonstrate that lung/liver M{Phi} in hu-BLT mice support integrative HIV-1 infection in vivo, but their contribution to viral reservoir persistence during ART is minor when compared to CD4+ T-cells. This is consistent with the deficient development of LL-TRM we observed in hu-BLT mice. However, HIV-1 permissive M{Phi} present in this model likely contribute to viral rebound upon ART interruption. Therefore, HIV-1 cure interventions that are tested in such preclinical models should consider targeting HIV-1 replication in both M{Phi} and CD4+ T-cells.

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Cattin, A., Pham, T. N., Dias, J., Do Rosario, N. F., Marchand, L. R., Volodina, O., Guimond, J.-V., Patey, N., Li, Y., Beland, K., Jenabian, M.-A., Estaquier, J., Haddad, E., Cohen, E. A., Ancuta, P.. 2025-02-06. Humanized Bone Marrow-Liver-Thymus Mice for Studying HIV-1 Persistence in Liver and Lung CD4+ T and Myeloid Cell Subsets during Antiretroviral Therapy. https://doi.org/10.1101/2025.01.31.635552

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