Probes for CD8

Follicular helper CD4 T cells, TFH, residing in B-cell follicles within secondary lymphoid tissues, are readily infected by AIDS viruses and are a major source of persistent virus despite relative control of viral replication. This persistence is due at least in part to a relative exclusion of effective antiviral CD8 T cells from B-cell follicles. To determine whether CD8 T cells could be engineered to enter B-cell follicles, we genetically modified unselected CD8 T cells to express CXCR5, the chemokine receptor implicated in cellular entry into B-cell follicles. Engineered CD8 T cells expressing CXCR5 (CD8hCXCR5) exhibited ligand specific signaling and chemotaxis in vitro. Six infected rhesus macaques were infused with differentially fluorescent dye-labeled autologous CD8hCXCR5 and untransduced CD8 T cells and necropsied 48h later. Flow cytometry of both spleen and lymph node samples revealed higher frequencies of CD8hCXCR5 than untransduced cells, consistent with preferential trafficking to B-cell follicle-containing tissues. Confocal fluorescence microscopy of thin-sectioned lymphoid tissues demonstrated strong preferential localization of CD8hCXCR5 T cells within B-cell follicles with only rare cells in extrafollicular locations. CD8hCXCR5 T cells were present throughout the follicles with some observed near infected TFH In contrast, untransduced CD8 T cells were found in the extrafollicular T-cell zone. Our ability to direct localization of unselected CD8 T cells into B-cell follicles using CXCR5 expression provides a strategy to place highly effective virus-specific CD8 T cells into these AIDS virus sanctuaries and potentially suppress residual viral replication.IMPORTANCE AIDS virus persistence in individuals under effective drug therapy or those who spontaneously control viremia remains an obstacle to definitive treatment. Infected follicular helper CD4 T cells, TFH, present inside B-cell follicles represent a major source of this residual virus. While effective CD8 T-cell responses can control viral replication in conjunction with drug therapy or in rare cases spontaneously, most antiviral CD8 T cells do not enter B-cell follicles and those that do fail to robustly control viral replication in the TFHpopulation. Thus, these sites are a sanctuary and a reservoir for replicating AIDS viruses. Here, we demonstrate that engineering unselected CD8 T cells to express CXCR5, a chemokine receptor on TFH associated with B-cell follicle localization, redirects them into B-cell follicles. These proof of principle results open a pathway for directing engineered antiviral T cells into these viral sanctuaries to help eliminate this source of persistent virus.

African green monkeys (AGMs) are a natural host of SIV that do not develop simian AIDS. Adult AGMs naturally have low numbers of CD4+T cells and a large population of MHC class II-restricted CD8αα T cells that are generated through CD4 downregulation in CD4+ T cells. In this article, we study the functional profiles and SIV infection status in vivo of CD4+ T cells, CD8αα T cells, and CD8αβ T cells in lymph nodes, peripheral blood, and bronchoalveolar lavage fluid of AGMs and rhesus macaques (in which CD4 downregulation is not observed). We show that, although CD8αα T cells in AGMs maintain functions associated with CD4+ T cells (including Th follicular functionality in lymphoid tissues and Th2 responses in bronchoalveolar lavage fluid), they also accumulate functions normally attributed to canonical CD8+ T cells. These hyperfunctional CD8αα T cells are found to circulate peripherally, as well as reside within the lymphoid tissue. Due to their unique combination of CD4 and CD8 T cell effector functions, these CD4- CD8αα T cells are likely able to serve as an immunophenotype capable of Th1, follicular Th, and CTL functionalities, yet they are unable to be infected by SIV. These data demonstrate the ambiguity of CD4/CD8 expression in dictating the functional capacities of T cells and suggest that accumulation of hyperfunctional CD8αα T cells in AGMs may lead to tissue-specific antiviral immune responses in lymphoid follicles that limit SIV replication in this particular anatomical niche.

Infection with HIV persists despite suppressive antiretroviral therapy (ART), and treatment interruption results in rapid viral rebound. Antibody-mediated CD8(+) lymphocyte depletion in simian immunodeficiency virus (SIV)-infected rhesus macaques (RMs) shows that these cells contribute to viral control in untreated animals. However, the contribution of CD8(+) lymphocytes to maintaining viral suppression under ART remains unknown. Here, we have shown that in SIV-infected RMs treated with short-term (i.e., 8-32 week) ART, depletion of CD8(+) lymphocytes resulted in increased plasma viremia in all animals and that repopulation of CD8(+) T cells was associated with prompt reestablishment of virus control. Although the number of SIV-DNA-positive cells remained unchanged after CD8 depletion and reconstitution, the frequency of SIV-infected CD4(+) T cells before depletion positively correlated with both the peak and area under the curve of viremia after depletion. These results suggest a role for CD8(+) T cells in controlling viral production during ART, thus providing a rationale for exploring immunotherapeutic approaches in ART-treated HIV-infected individuals.