Human immunodeficiency virus type 1-infected persons with residual disease and virus reservoirs on suppressive highly active antiretroviral therapy can be stratified into relevant virologic and immunologic subgroups

A significant percentage of human immunodeficiency virus type 1 (HIV-1)-infected persons treated with highly active antiretroviral therapy (HAART) will develop plasma HIV-1-specific virion RNA levels <50 copies/mL. HIV-1-infected persons receiving virally suppressive HAART were studied with a viral outgrowth assay of the patients' peripheral blood mononuclear cells (PBMC), and a quantitative polymerase chain reaction assay was used to analyze HIV-1 2-long terminal repeat (2-LTR) circular DNA in PBMC, which indicates new HIV-1 infections of cells in vivo. Viral outgrowth in vitro correlated inversely with the level of peripheral blood CD4(+) T lymphocytes. Detection and quantitation of 2-LTR circular DNA correlated strongly with viral outgrowth patterns and inversely with CD4(+) T lymphocyte counts. Relevant subgroups of HIV-1-infected subjects on suppressive HAART with residual viral disease and reservoirs can now be stratified.     

Cell-associated HIV-1 messenger RNA and DNA in T-helper cell and monocytes in asymptomatic HIV-1-infected subjects on HAART plus an inactivated HIV-1 immunogen.

OBJECTIVE: We examined the effect of an HIV-1-specific immune-based therapy on cell-associated HIV-1 DNA and RNA. DESIGN: Five HIV-1-infected subjects receiving HIV-1 immunogen plus HAART were compared with three HIV-1-infected subjects who received incomplete Freund's adjuvant (IFA) plus HAART. METHODS: Cell-associated HIV-1 RNA or DNA in lymphocytes and monocytes was determined using a dual immunophenotyping/in situ hybridization assay with or without in situ PCR amplification. RESULTS: Cell-associated HIV-1 RNA in CD4 cells correlated with plasma RNA overall. CD4, HIV-1 gag-pol messenger (m)RNA+ cells decreased in the immunogen plus HAART group compared with the IFA plus HAART group. Decreases in HIV-1 DNA+ CD4 cells were observed in the immunogen plus HAART compared with the IFA plus HAART group. Decreases in HIV-1 gag-pol mRNA+ monocytes were observed in the immunogen plus HAART group compared with the IFA plus HAART group. Consistent with the findings in CD4 cells, decreases in HIV-1 DNA+ monocytes were observed in the immunogen plus HAART group compared with the IFA plus HAART group. CONCLUSIONS: These preliminary observations support the rationale for examining the combination of immune-based therapies and antiretroviral drugs for effective HIV-1 control.     

Unexpected coreceptor usage of primary human immunodeficiency virus type 1 isolates from viremic patients under highly active antiretroviral therapy

Recently, combinations of antiretroviral drugs (highly active antiretroviral therapy [HAART]) have led to a dramatic reduction of human immunodeficiency virus type 1 (HIV-1)-related clinical symptoms. Success of treatment is defined as almost complete suppression of plasma viremia, although in a sizable fraction of patients this goal is not achieved. We characterized primary HIV-1 isolates from 2 cohorts of patients in which HAART failed in terms of viral suppression. One cohort showed clinical benefit and stable or increasing CD4+ T cell numbers despite high viral load. The second viremic cohort had no CD4+ T cell recovery and exhibited typical AIDS-related symptoms. Primary isolates from HAART patients with minor clinical symptoms used CXCR4 as the most relevant receptor on primary cells. Thus, for the first time, it is shown that patients improving clinically under HAART harbor relatively high viral loads with viruses preferring CXCR4 as coreceptor.     

The naive CD4+ count in HIV-1-infected patients at time of initiation of highly active antiretroviral therapy is strongly associated with the level of immunological recovery

Current antiretroviral therapy can induce considerable, sustained viral suppression followed by immunological recovery, in which naive CD4 + cells are important. Long-term immunological recovery was investigated during the first 3 y of highly active antiretroviral therapy (HAART) in 210 HIV-1-infected patients. The focus was on the naive CD4 + cell time course and associations between naive CD4 + cell counts and established prognostic markers. Total and naive CD4 + cell counts were measured using flow cytometry. The HIV-RNA detection limit was 20 copies/ml. During 36 months of HAART, the total CD4 + count followed a triphasic pattern, reflecting an initial phase of rapid redistribution from lymphoid tissues, followed by a slow increase, partially due to an increase in naive CD4+ cell count. From Month 18 onwards, both naive and total CD4 + cell counts stabilized, although viral suppression was sustained. There was no association between plasma viral load and the increase in naive CD4 + cell count. Importantly, baseline naive CD4 + cell count was significantly associated with the change in naive CD4 + cell count, suggesting that the naive cell count at baseline does influence the immunological recovery that can be obtained from treatment. Surprisingly, the naive CD4 + cell count tended to stabilize at a subnormal level after 18 months of HAART. This finding merits further investigation.     

抗病毒治疗对HIV/AIDS患者自然杀伤细胞和γδT细胞的重建作用研究

目的 研究高效联合抗逆转录病毒治疗(HAART)对HIV/AIDS患者NK细胞和γδT细胞的重建作用,比较HAART对获得性免疫和固有免疫重建规律的异同.方法 对59例接受规律HAART的HIV/AIDS患者于治疗前、治疗后1、3、6、9、12个月末进行随访,采用分支DNA(bDNA)法检测血浆病毒载量,经流式细胞仪检测患者外周血淋巴细胞亚群.以56例健康献血员作为正常对照.结果 基线时HIV/AIDS患者CD4+T细胞中位数为45(164,13)个/μl,NK细胞和γδT细胞中位数分别为96(135,56)个/μl和30(45,19)个/μl,与正常对照组相比均显著减低(P<0.001).HAART开始后1个月,血浆病毒载量降低了2.33 lg拷贝/ml(P<0.001),CD4+T细胞数增长了111个/μl(P<0.001),主要以记忆CD4+T细胞的快速增长为主,而NK细胞和γδT细胞与基线相比无显著变化.在HAART后3个月NK细胞和.γδT细胞数量出现增长,分别较基线增加26个/μl(P=0.007)和15个/μl(P<0.001).治疗12个月,CD4+T细胞、NK细胞及γδT细胞计数分别达到271(365,134)个/μl、166(281,108)个/μl和46(66,34)个/μl,但仍低于正常对照.结论 HAART可以重建HIV/AIDS患者固有免疫,但NK细胞和γδT细胞数量的增加晚于CD4+ T细胞,HAART对固有免疫的重建作用机制有待进一步研究.     

Peripheral blood Dendritic cells are not a major reservoir for HIV type 1 in infected individuals on virally suppressive HAART

Dendritic cells (DCs) are potent antigen-presenting cells, and their physiological localization in tissues that interact with the external environment is important as a first barrier against pathogens such as human immunodeficiency virus type I (HIV-1). Several models have been proposed to explain the possible role of DCs as a reservoir for HIV-1 in patients on virally suppressive highly active antiretroviral therapy (HAART). However, the low yield of cell isolates has made this evaluation a difficult task. The present study analyzes whether peripheral blood DCs from HIV-1-infected individuals on virally suppressive HAART, with plasma HIV-1 RNA levels of less than 50 copies/ml, carry either HIV-1 provirus and/or HIV-1 virions. Peripheral blood DCs were isolated from a cohort of 10 HIV-1-seropositive men taking suppressive HAART. In five patients, plasmacytoid and myeloid dendritic cells were isolated to attempt to identify their respective roles in HIV-1 residual disease. Viral out-growth assays were performed in vitro, as well as gag and R/U5 polymerase chain reaction (PCR) amplification of viral RNA and DNA, respectively, from DC and peripheral blood mononuclear cell (PBMC) extracts. Fluorescence activated cell-sorting (FACS) data revealed cellular yields from 85.90 to 95.18%, of relatively pure DCs isolated from patients' PBMCs. Although HIV-1 RNA gag and DNA RU/5 were detected in all PBMC samples isolated from the patients, proviral DNA and viral RNA forms were not detected in any of the DC isolates. In addition, no replication-competent virus was demonstrated in DC coculture assays, while virus was isolated from each patients' CD8+ T-lymphocyte-depleted PBMC cocultures. Furthermore, HIV-1 gag proviral DNA was not detected in either plasmacytoid or myeloid DC subfractions. The current study suggests that in HIV-1-infected individuals treated with suppressive HAART, peripheral blood DCs do not carry HIV-1 proviral DNA or viral particles attached to their surface. These populations of peripheral blood DCs are likely not a major HIV-1 reservoir in patients on HAART with clinically undetectable plasma viral RNA.     

Persistent HIV-1 infection of natural killer cells in patients receiving highly active antiretroviral therapy

We have identified a subset of CD56(+)CD3(-) human natural killer (NK) cells that express CD4 and the HIV coreceptors CCR5 and CXCR4. These cells can be productively infected in vitro by both CCR5- and CXCR4-using molecular clones of HIV-1 in a CD4-dependent manner. Analysis of HIV-infected persons showed that viral DNA is present in purified NK cells, and virus could be rescued from these cells after in vitro cultivation. Longitudinal analysis of the HIV-1 DNA levels in NK cells from patients after 1-2 years of highly active antiretroviral therapy indicated that NK cells remain persistently infected and account for a substantial amount of the viral DNA in peripheral blood mononuclear cells. These results demonstrate that a subset of non-T cells with NK markers are persistently infected and suggest that HIV infection of NK cells is important for virus persistence. The properties of the virus reservoir in these cells should be considered in attempts to further optimize antiretroviral therapies.     

Longitudinal analysis of lymphocyte ratios and HIV-1 intracellular DNA levels in children

The associations between human immunodeficiency virus type 1 (HIV-1) intracellular DNA and immunological markers were analyzed longitudinally for children with sustained, undetectable RNA levels while receiving highly active antiretroviral therapy (HAART) for >2 years. When DNA levels reached a plateau at week 104 of therapy, in contrast to findings for adults, there was no correlation between the CD4(+) : CD8(+) ratio and DNA levels (r=-0.02; P=.95), and naive CD4(+)CD45RA(+) lymphocytes predominated. These data suggest that the increased proportion of naive lymphocytes found in children are less susceptible to HIV-1 infection than are the memory lymphocytes that dominate immune reconstitution in adults.     

Recovery of replication-competent virus from CD4 T cell reservoirs and change in coreceptor use in human immunodeficiency virus type 1-infected children responding to highly active antiretroviral therapy

Highly active antiretroviral therapy (HAART) suppresses plasma viremia in most patients with human immunodeficiency virus (HIV) infection. Prospective study of HIV-infected children (n=27) shows that, in 8 of 12 who responded to HAART (>/=0.5 log reduction in plasma HIV RNA), HAART restricted the number of coreceptors used by the predominant HIV isolate (mean number of coreceptors used at baseline was 4, vs. 1 coreceptor used at 6 months after treatment). This decrease was most striking in 6 of 8 children whose HIV coreceptor tropism changed from X4-tropic at baseline to R5-tropic. In 6 of 10 children tested, with plasma HIV RNA levels of <50 copies/mL, R5-tropic virus was isolated from CD4 T cell reservoirs. All the responding children had a significant increase in naive CD4 T cells (P<.05). These results show that persistent HIV T cell reservoirs are present in children and that HAART may influence the number and type of coreceptors used by the predominant virus isolate.     

Expansion of CD1d-restricted NKT cells in patients with primary HIV-1 infection treated with interleukin-2

Innate CD1d-restricted natural killer T (NKT) cells are infected and lost in HIV-1-infected patients, and this could contribute to HIV-1 pathogenesis because NKT cells play an important role in directing both adaptive and innate immunity. Administration of interleukin-2 (IL-2) to HIV-1-infected patients leads to substantial and sustained CD4+ T-cell expansion, involving both naive and memory cells. We investigated whether IL-2 treatment could restore the NKT cell compartment in patients with primary HIV-1 infection. We show that IL-2 combined with effective antiretroviral therapy (ART) resulted in significant expansion of CD1d-restricted NKT cells. Expansion occurred in both the CD4- and CD4+ subsets of NKT cells, and expanded cells expressed the CD161 maturation marker while expression of the HIV coreceptor CCR5 was reduced. These data indicate that IL-2 treatment in combination with effective ART is beneficial for the restoration of innate NKT cell immunity in patients with primary HIV-1 infection.     

Amplification of low-frequency antiviral CD8 T cell responses using autologous dendritic cells.

OBJECTIVE: To utilize the potent antigen-presenting capacity of mature dendritic cells (MDC) in order to develop a rapid, sensitive method for quantifying antigen-specific CD8 T cells present at low frequency in peripheral blood. DESIGN: Peripheral blood mononuclear cells (PBMC) were obtained from seven HIV-1-positive individuals with low to moderate CD8 T cell responses, including five on highly active antiretroviral therapy (HAART). IFN-gamma ELISPOT assays were performed using either monocytes or MDC to present antigens expressed by recombinant vaccinia viruses (r-VV). METHODS: Peripheral blood-derived monocytes were cultured for 5-6 days in the presence of IL-4 and granulocyte macrophage colony-stimulating factor, then matured in monocyte-conditioned medium. MDC were infected with r-VV and co-cultured in an ELISPOT assay with autologous monocyte-depleted PBMC. RESULTS: Relative to autologous monocytes, MDC amplified detection of antigen-specific CD8 T cells by 2-30-fold in response to antigens from HIV-1, Epstein-Barr virus and cytomegalovirus. Furthermore, antigenic specificities were revealed that had not been detected using standard ELISPOT of PBMC. CONCLUSION: This assay will prove useful for the detection of memory T cells present at low frequency, and may be of interest for identifying subdominant cytotoxic T lymphocyte epitopes. This method may have broad applications for the detection of antiviral CD8 T cell responses in patient populations in whom such responses have been difficult to detect, including HIV-1-seropositive individuals with advanced disease or undergoing HAART.     

Molecular evidence for drug-induced compartmentalization of HIV-1 quasispecies in a patient with periodic changes to HAART

OBJECTIVE: To perform molecular analysis of the predominant viral populations and drug-resistance mutations from plasma and peripheral blood mononuclear cell (PBMC) compartments over time from an HIV infected patient, who experienced virological failure while on different HAART regimens. MATERIALS AND METHODS: In a longitudinal study proviral and plasma HIV-1 sequences were amplified in the pol, protease and env genes and were sequenced directly and analysed phylogenetically. Virus was recovered from time points corresponding to viral load peaks using co-culturing techniques. The periodic failure of different highly active antiretroviral therapy (HAART) regimens was analysed sequencing. RESULTS: Longitudinal follow-up studies revealed four inflection peaks of plasma viraemia associated with the recovery of culturable virus in vitro, which indicated failure of the concurrent HAART regimen. Molecular analysis of viral strains revealed evidence of continual evolution and compartmentalization of drug-resistance mutants/quasispecies between plasma and PBMC, with the widest spectrum of mutations isolated from plasma. Importantly, these data show the periodic appearance and clearance of drug-resistance mutants concomitant with the introduction and withdrawal of zidovudine over time. CONCLUSION: This report is unique in showing drug-induced compartmentalization of viral quasispecies under the control of different HAART regimens in both plasma and PBMC. Introduction and withdrawal of zidovudine from the HAART regimen had direct bearing on the appearance and disappearance of specific zidovudine drug-resistance mutations in plasma-derived virus. This data has important implications for the management of HIV-infected patients with poor compliance with certain HAART regimens, and also in predicting the late emergence of drug-resistance mutations via the latent integrated provirus.     

Immunological and virological responses in HIV-1-infected adults at early stage of established infection treated with highly active antiretroviral therapy

OBJECTIVE: To evaluate the immunological and virological responses to highly active antiretroviral therapy (HAART) in blood and lymphoid compartments of HIV-1-infected patients at an early stage of infection. DESIGN: An open-label, observational, non-randomized, prospective trial of outpatients attending the Centre of Clinical Investigation in Infectious Diseases, Centre Hospitalier Universitaire Vaudois, University of Lausanne, Switzerland. SUBJECTS: Forty-one antiretroviral-naive HIV-1-infected adults with 400 CD4 T cells/microl or greater and 5000 plasma HIV-1-RNA copies/ml or greater were enrolled, and 32 finished the study. Forty-nine HIV-negative individuals were included as controls. All subjects gave written informed consent. INTERVENTIONS: All patients received abacavir 300 mg by mouth every 12 h and amprenavir 1200 mg by mouth every 12 h for 72 weeks. MAIN OUTCOME MEASURES: The extent of immune reconstitution in blood and lymph nodes after 72 weeks of HAART was evaluated, and compared with immunological measures of 49 HIV-negative subjects. RESULTS: Virus replication was effectively suppressed (-3.5 log10 at week 72). Substantial increments of CD4 T cell count in blood and percentage in lymph nodes were observed over time, and these measures were comparable to HIV-negative subjects by week 24 in blood and by week 48 in lymph nodes. The increase was equally distributed between naive and memory CD4 T cells. Recovery of HIV-specific CD4 responses occurred in 40% of patients. CONCLUSION: The initiation of HAART at an early stage of established HIV infection induces systemic quantitative normalization of CD4 T cells, a partial recovery of HIV-specific CD4 cell responses, and effective and durable suppression of virus replication.     

Therapeutic immune reconstitution in HIV-1-infected children is independent of their age and pretreatment immune status.

OBJECTIVE: To evaluate long-term immune reconstitution of children treated with highly active antiretroviral therapy (HAART). METHODS: The long-term immunological response to HAART was studied in 71 HIV-1-infected children (aged 1 month to 18 years) in two prospective, open, uncontrolled national multicentre studies. Blood samples were taken before and after HAART was initiated, with a follow-up of 96 weeks, and peripheral CD4 and CD8 T cells plus naive and memory subsets were identified in whole blood samples. Relative cell counts were calculated in relation to the median of the age-specific reference. RESULTS: The absolute CD4 cell count and percentage and the CD4 cell count as a percentage of normal increased significantly (P < 0.001) to medians of 939 x 106 cells/l (range, 10-3520), 32% (range, 1-50) and 84% (range, 1-161), respectively, after 48 weeks. This increase was predominantly owing to naive CD4 T cells. There was a correlation between the increase of absolute naive CD4 T cell counts and age. However, when CD4 T cell restoration was studied as percentage of normal values, the inverse correlation between the increase of naive CD4 T cell count and age was not observed. In addition, no difference in immunological reconstitution was observed at any time point between virological responders and non-responders. CONCLUSIONS: Normalization of the CD4 cell counts in children treated with HAART is independent of age, indicating that children of all age groups can meet their CD4 T cell production demands. In general, it appears that children restore their CD4 T cell counts better and more rapidly than adults, even in a late stage of HIV-1 infection.     

Cd4+ T cells programmed to traffic to lymph nodes account for increases in numbers of cd4+ T cells up to 1 year after the initiation of highly active antiretroviral therapy for human immunodeficiency virus type 1 infection

Cells programmed to traffic through lymph nodes dominate initial increases in total CD4(+) T cell numbers after highly active antiretroviral therapy (HAART) is begun for human immunodeficiency virus type 1 (HIV-1) infection. However, it is unknown whether this dominance continues throughout the first year of treatment. To examine this question, 10 subjects who had a positive response to HAART for 1 year were selected from a cohort of 20 who were receiving this treatment. Flow cytometry, which was used to characterize CD4(+) T cell subsets by immunophenotype, demonstrated that cells programmed to traffic through lymph nodes, irrespective of their memory or naive phenotype, continued to best account for increases in CD4(+) T cells, even 1 year after starting HAART. This suggests that, although this pool is preferentially depleted during HIV-1 infection, HAART allows for reaccumulation of these cells for at least 1 year. Furthermore, it suggests that phenotypic differences based on markers of lymphocyte trafficking may be more relevant for understanding HIV-1 pathogenesis than are naive and memory markers alone.     

Comparative analysis of total and integrated HIV-1 DNA in peripheral CD4 lymphocytes and monocytes after long treatment with HAART

OBJECTIVE: To determine the level of total and integrated HIV-1 DNA load in CD4 lymphocytes and monocytes of patients undergoing HAART treatment for at least 2 years. METHODS: CD4 lymphocytes were isolated by subjecting monocyte-depleted blood samples to immune-purging carried out with M-450 Dynabeads. Monocytes were separated by blood through a combined procedure of cell adherence to dishes and complement induced immune lysis with anti-CD3 Mab. HIV DNA in CD4 lymphocytes and monocytes was quantified by polymerase chain reaction (PCR) based limit dilution assay with two PCR protocols, specific for total (LTR PCR) and integrated (Alu PCR) forms of HIV DNA. The replication competence of the provirus harboured in monocyte-depleted peripheral blood mononuclear cells (PBMC) and adherent monocytes was assayed by measuring HIV-1 p24 antigen produced by in-vitro cultures established with these cells. RESULTS: The CD4 lymphocytes of all patients contained a consistent number of HIV DNA copies. Most patients were also positive for HIV DNA in monocytes. The Alu PCR analysis detected, integrated provirus in CD4 lymphocytes of 9 patients and in the monocytes of only three. Four patients had replication-competence virus in their PBL. The monocytes of all patients did not produce virus in vitro. CONCLUSION: The HIV infection of CD4 lymphocytes and monocytes is maintained even after HAART related, apparent, and durable suppression of viral replication. We suggest that the viral persistent infection of monocytes may play a role in maintaining the residual HIV activity found in patients undergoing HAART.     

Quantification of integrated and total HIV-1 DNA after long-term highly active antiretroviral therapy in HIV-1-infected patients.

OBJECTIVE: To assess the impact of long-term virus suppression on the peripheral blood CD4 T cells integrated and total HIV-1 DNA loads in patients receiving highly active antiretroviral therapy (HAART). METHODS: A total of 10 HIV-1-infected patients receiving a triple combination therapy (two nucleoside analogues and one protease inhibitor) were longitudinally studied to compare integrated and total HIV-1 DNA loads. HIV-1 DNA quantification was performed using a quantitative nested polymerase chain reaction (PCR) on genomic peripheral blood mononuclear cell (PBMC) DNA obtained at baseline and at 48 weeks of HAART. RESULTS: All the study patients showed an early and sustained decrease in plasma HIV-1 RNA to below the limit of detection (200 copies/ml). Concordant with the plasma viral decline, a significant increase in the CD4 T cell count was observed (P = 0.007). A statistically significant fivefold decrease in total HIV-1 DNA was detected after 48 weeks of HAART (P = 0.005). However, no statistically significant change was noted after the therapy when the integrated HIV-1 DNA copy number was compared (P = 0.333). Taken together, these results suggest that in the patients analysed the integrated HIV-1 DNA does not decay rapidly after HAART. CONCLUSION: Within the study cohort the total amount of PBMC HIV-1 DNA decreased drastically after 48 weeks of HAART. Nevertheless, the integrated HIV-1 DNA did not significantly decay during this period. Although the data presented here are limited by the number of patients analysed, our findings suggest that 48 weeks of HAART does not significantly reduce the integrated HIV-1 proviral DNA load in the latently infected CD4 T cell reservoir.     

Effects of interleukin-2 plus highly active antiretroviral therapy on HIV-1 replication and proviral DNA (COSMIC trial)

BACKGROUND: The effect of interleukin-2 (IL-2) in combination with antiretroviral therapy on HIV-1 replication and reservoirs was investigated. METHODS: In a prospective, open-label trial, 56 asymptomatic HIV-1-infected subjects (CD4 T cell count > 350 x 10(6) cells/l) were randomized to highly active antiretroviral therapy (HAART: stavudine, lamivudine, nelfinavir, saquinavir) with or without IL-2 (9 megaunits daily for 5 days in 6-weekly intervals for a total of eight cycles). Productive and latent infection were analysed in peripheral blood, and residual virus replication in the lymphoid tissue and in the cerebrospinal fluid. The influence of IL-2 on viral rebound after treatment discontinuation was studied. RESULTS: Virus replication was detected in 21 of 31 on-treatment lymph nodes despite undetectable plasma viraemia. Viral RNA was found in resting as well as in proliferating cells. RNA-negative patients tended towards more rapid proviral DNA elimination. Supplementary IL-2 led to a greater increase in CD4 T cell counts than HAART alone (P < 0.001), resulting in normalization in approximately 90% of IL-2-treated patients compared with approximately 50% HAART-only subjects. IL-2 had no beneficial effect on virus replication and on proviral DNA in peripheral blood. CONCLUSIONS: Viral persistence during HAART is partly a result of continued low-level replication, calling for more active regimens. IL-2 accelerates the normalization of CD4 T cell counts but does not impact on virus production or latency.     

Switch from inhibitory to activating NKG2 receptor expression in HIV-1 infection: lack of reversion with highly active antiretroviral therapy

BACKGROUND: HIV-1 infection is characterized by increase in inhibitory receptors and loss of activating receptors on natural killer (NK) cells, resulting in loss of cell activity. Exceptionally, for an inhibitory receptor, the proportion of NK cells bearing CD94-NKG2A decreases during HIV-1 infection. It is not understood whether HIV-1 itself or other concomitant infections drive these changes. OBJECTIVES: To investigate the relationship between HIV-1 viraemia and changes in C-type lectin-like receptor expression in NK cells and to investigate the effect of highly active antiretroviral therapy (HAART) on these changes. METHODS: Three cohorts of patients were studied: (1) before, during and after treatment interruption in aviraemic and viraemic patients receiving HAART (n = 15); (2) HIV-1-positive treatment-naive individuals (n = 13); and (3) HIV-1-positive individuals receiving successful HAART for a minimum of 1 year without interruption (n = 11). Flow cytometry was used to study the expression of NKG2A before and after treatment interruption and to define expanded populations of NK cells in untreated and treated HIV-1-positive individuals. Assays were performed in vitro to assess the cytotoxicity of the expanded populations. RESULTS: Increases in plasma HIV-1 RNA during treatment interruption in aviraemic HAART-treated individuals did not influence the proportion of NK cells carrying the complex CD94-NKG2A. Loss of NKG2A NK cells corresponded to the dramatic expansion of a distinct population of cells expressing a functional activating CD94-NKG2C receptor with skewed expression of killer cell immunoglobulin-like receptor family and natural cytotoxicity receptors. CONCLUSION: Changes in the NK cell repertoire during HIV-1 infection were not a result of HIV-1 viraemia alone but resembled those associated with concomitant infections.