Natural Suppression of Human Immunodeﬁciency Virus Type 1 Replication Is Mediated by Transitional Memory CD8 (cid:1) T Cells (cid:1)

HIV replication is suppressed in vitro by a CD8 (cid:1) cell noncytotoxic antiviral response (CNAR). This activity directly correlates with an asymptomatic clinical state. The objective of this study was to identify the phenotype of CD8 (cid:1) cell subsets having strong CNAR activity. CD8 (cid:1) cell subset frequencies and CNAR levels were measured for human immunodeﬁciency virus (HIV)-uninfected individuals and three groups of HIV type 1 (HIV-1)-infected individuals: asymptomatic individuals with low-level viremia (vHIV), antiretroviral-drug-treated subjects with undetectable virus levels (TxHIV), and therapy-naïve aviremic elite controllers (EC). CD8 (cid:1) cells from the vHIV individuals exhibited the highest HIV-suppressing activity and had elevated frequencies of CD45RA (cid:2) CD27 (cid:1) and PD-1 (cid:1) (CD279 (cid:1) ) cells. Functional assessments of CD8 (cid:1) cells sorted into distinct subsets established that maximal CNAR activity was mediated by CD45RA (cid:2) CCR7 (cid:2) CD27 (cid:1) and PD-1 (cid:1) CD8 (cid:1) cells. T cell receptor (TCR) repertoire proﬁles of CD8 (cid:1) cell subsets having strong CNAR activity exhibited increased perturbations in comparison to those of inactive subsets. Together, these studies suggest that CNAR is driven by HIV replication and that this antiviral activity is associated with oligoclonally expanded activated CD8 (cid:1) cells expressing PD-1 and having a transitional memory cell phenotype. The ﬁndings better describe the identity of CD8 (cid:1) cells showing CNAR and should facilitate the evaluation of this important immune response in studies of HIV pathogenesis, resistance to infection, and vaccine development. CNAR assays. To determine the respective levels of CNAR activity, incremen- tal numbers of CD8 (cid:1) cells (without in vitro stimulation) were cocultured with acutely HIV-infected autologous or heterologous CD4 (cid:1) cells and the ensuing level of HIV replication were measured. Brieﬂy, CD4 (cid:1) cells were resuspended (3 (cid:7) 10 6 cells/ml) in growth medium (RPMI 1640 medium supplemented with fetal calf serum [heat inactivated at 56°C for 30 min; 10%, vol/vol], penicillin [100 U/ml], streptomycin [100 (cid:6) g/ml], L -glutamine [2 mM], and recombinant inter- leukin 2 [IL-2; 100 U/ml; Invitrogen]) and stimulated for 3 days in the presence of phytohemagglutinin-leucoagglutinin (PHA-L; 3 (cid:6) g/ml; Sigma) in a 37°C hu- midiﬁed incubator. Subsequently, 10 (cid:7) 10 6 cells were treated with Polybrene (2 (cid:6) g/ml; Sigma) for 20 min at 37°C and pelleted. The pelleted cells were resus- pended in 1 ml of HIV-1 SF33 (10,000 50% tissue culture infective doses [TCID 50 ]/ml in PBMC) for 1 h at 37°C with periodic mixing. HIV-1 SF33 , a syncytium-inducing (SI), CXCR4-tropic (X4) strain, has been maintained in primary cells since its isolation, exhibits rapid replication kinetics with a high degree of cytopathicity in cell culture, and is not sensitive to (cid:8) -chemokine-mediated antiviral effects (31). The acutely infected CD4 (cid:1) cells were then washed and resuspended at 10 6 cells/ml of growth medium, and 200- (cid:6) l aliquots were placed into a ﬂat-bottom 96-well tissue culture plate (Falcon 3072; BD) cation CD45RA CD45RA CD28 Comparisons of bulk CD8 (cid:1) cells, which were unstained or stained with antibodies, revealed no substantial differences in CNAR activity (data not shown). Thus, activation of the cells and/or blocking of the surface antigens due to antibody binding was not involved. These results show that is associated with memory CD8 (cid:1) cells, particularly those with a transitional memory phenotype.