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T-cell activation positively correlates with cell-associated HIV-DNA level in viremic patients with primary or chronic HIV-1 infection

Weiss, Laurencea,b,c; Chevalier, Mathieu F.b,d,*; Assoumou, Lamberte,f; Didier, Célineb; Girard, Pierre-Marieg; Piketty, Christophec; Costagliola, Dominiquee,f; Rouzioux, Christinehthe ANRS 116 SALTO Study Group

doi: 10.1097/QAD.0000000000000319
Research Letters

We investigated the relationship between the size of blood HIV reservoirs and T-cell activation in patients with primary HIV infection (PHI) and chronic HIV infection (CHI) before and after antiretroviral therapy (ART) interruption. Levels of T-cell activation strongly positively correlated with HIV-DNA levels in viremic PHI and CHI patients. In ART-treated CHI patients, residual immune activation was not associated with HIV-DNA levels. Interestingly, early levels of HIV-DNA in PHI predicted the extent of residual T-cell proliferation under ART.

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aUniversité Paris Descartes, Sorbonne Paris Cité

bInstitut Pasteur, Régulation des infections rétrovirales

cAP-HP Hôpital Européen Georges Pompidou

dUniversité Paris Diderot, Sorbonne Paris Cité


fUniversité Pierre et Marie Curie

gAP-HP Hôpital Saint-Antoine

hUniversité Paris Descartes, EA 3620, Paris, France.

*Present address: Urology Research Unit, Lausanne University Hospital, Lausanne, Switzerland.

Correspondence to Laurence Weiss, MD, PhD, Institut Pasteur, 25 rue du Dr Roux, 75015 Paris, France. Tel: +33 156 093 297; fax: +33 156 093 026; e-mail:

Received 24 October, 2013

Revised 22 April, 2014

Accepted 24 April, 2014

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Although most HIV-infected patients receiving antiretroviral therapy (ART) exhibit an effective and sustained control of plasma viral load, HIV-1 persists in latently infected cells in the blood [1,2], as well as in tissues [3]. The persistence of T-cell activation in virally suppressed patients is associated with poor CD4+ cell reconstitution and may result in increased mortality/morbidity [4,5]. Low-level viremia supported by latently infected cells might contribute to this persistent chronic T-cell activation [6], whereas immune activation/T-cell proliferation may in turn be involved in HIV reservoir seeding [7]. Few data are available regarding the relationship between the size of HIV reservoirs and the level of residual immune activation in virally suppressed patients and inconsistent results have been reported [8,9].

The aim of this study was to evaluate the relationship between the size of blood HIV reservoirs and T-cell activation, in viremic patients with primary HIV infection (PHI) and in patients with chronic HIV infection (CHI) before and after ART treatment interruption (TI). We also investigated the impact of the HIV reservoir size on residual immune activation after 6 months of ART in PHI patients. We used levels of total HIV-DNA in peripheral blood mononuclear cells (PBMCs) measured in whole blood by the ANRS real-time PCR assay (Biocentric, Bandol, France) [10] to estimate the HIV reservoir size, as this marker is representative and predictive of disease progression [11].

Twenty-two individuals with PHI (median estimated time postinfection: 41 days) were recruited in a prospective study [12,13]. Patients were enrolled before ART might be initiated. Ten patients received ART after enrollment in the study.

Twenty-five patients with CHI were enrolled in a substudy [14] of the ANRS 116 SALTO trial, a prospective multicentre study of TI in patients who had started treatment at CD4+ cell counts above 350/μl (NCT00118677) [15]. ART was interrupted at day 0 (baseline). After 12 months of TI, none of the patients had resumed ART. Written informed consent was provided by the study participants, according to French ethical laws. Both studies were approved by French ethical committees. Ex-vivo T-cell activation was assessed by flow cytometry measuring expression of HLA-DR, CD38 and Ki-67 markers on CD4+ and CD8+ T cells (Fig. S1,

Figure 1 a depicts patients’ characteristics for both cohorts. At baseline, in untreated PHI patients, HIV-DNA levels strongly correlated with the level of CD8+ T-cell activation as measured by the proportion of CD8+ T cells expressing CD38 (R = 0.64, P = 0.001), Ki-67 (R = 0.71, P = 0.0002) and moderately with the proportion of CD8+ T cells coexpressing HLA-DR and CD38 (R = 0.47, P = 0.034) (Fig. 1 e, data not shown). Moreover, HIV-DNA levels also correlated with the proportion of CD4+ T cells expressing CD38 (R = 0.51, P = 0.016), HLA-DR (R = 0.53, P = 0.013), and Ki67 (R = 0.61, P = 0.003) (data not shown). In multivariable analysis (assessing HLA-DR, CD38 and Ki-67 expression on CD8+ T cells), CD38 expression was the only activation marker independently associated with HIV-DNA levels (β = 0.62, P = 0.003).

Fig. 1

Fig. 1

Fig. 1

Fig. 1

At month 6, HIV-DNA levels significantly decreased in both ART-treated patients (P = 0.004) and untreated patients (P = 0.019) (Wilcoxon rank test; Fig. 1 b, c). The decrease in HIV-DNA levels between baseline and M6 was significantly greater in treated patients compared to untreated patients (−1.19 log versus −0.63 log copies/106 PBMCs, respectively; P = 0.003 according to the Mann–Whitney test). We next analyzed the impact of total HIV-DNA levels at baseline on T-cell activation at M6 in untreated and in successfully ART-treated patients. In untreated patients, HIV-DNA levels at baseline were not predictive of the extent of T-cell activation at M6. In contrast, in ART-treated patients, baseline HIV-DNA levels positively correlated with Ki67-expressing CD8+ T cells at M6 (R = 0.82, P = 0.004). There was also a trend to a positive relationship between baseline HIV-DNA levels and the proportion of Ki-67+ CD4+ T cells (R = 0.54; P = 0.10) (Fig. 1 f, g).

In patients with successfully treated CHI, there was no relationship between HIV-DNA levels and T-cell activation, regardless of whether assessed by the expression of CD38 and/or HLA-DR on CD4+ and CD8+ T cells (Fig. 1 h, data not shown). In contrast, at M12 of TI, HIV-DNA levels strongly correlated with the proportion of CD8+ and CD4+ T cells expressing CD38 (R = 0.77, P < 0.0001; and R = 0.72, P < 0.0001, respectively), but not HLA-DR (Fig. 1 i, data not shown). Interestingly, HIV-DNA levels before TI predicted the level of CD38+CD8+ T cells (R = 0.49, P = 0.013) and of CD38+CD4+ T cells (R = 0.69, P < 0.0001) at M12 of TI (Fig. 1 j, k).

The data presented here show a strong positive correlation between the size of blood HIV reservoirs and the extent of CD4+ and CD8+ T-cell activation in viremic patients with primary or chronic infection. Similar results were found when total HIV-DNA levels in blood (copies/106 PBMCs) were converted into copies/106 CD4+ T lymphocytes, as previously reported [10]. Our HIV assay is a measurement of total HIV-DNA level and did not allow us to distinguish between unintegrated and integrated forms of DNA. Of note, in multivariable analysis, the only marker independently associated with HIV reservoirs was CD38, previously reported to predict HIV disease progression independent of viral load [16,17].

We studied patients with CHI included in a trial of ART interruption to investigate the relationship between HIV reservoirs and T-cell activation before TI when plasma viremia was suppressed and, after TI, in the context of ongoing viral replication. Strikingly, HIV-DNA levels before TI were significantly associated with T-cell activation at M12. These results further support the important role of the viral reservoir on chronic immune activation. Moreover, T-cell activation and HIV replication/reservoir may be reciprocally interrelated; immune activation may be involved in HIV replication and reservoir seeding. Indeed, we previously reported that CD8+ T-cell activation before TI predicted the increase in HIV-DNA levels after 12 months of TI [14].

In ART-treated patients, the size of blood HIV reservoirs was not associated with residual T-cell activation. We did not evaluate the relationship between the degree of residual plasma viremia and T-cell activation. Two previous studies reported on a lack of correlation between low level of viral replication under ART and T-cell activation [8,18]. Although a strong association was reported between the size of HIV reservoir and the frequency of Programmed cell death-1 (PD-1)-expressing CD4+ T cells – a negative regulatory molecule and a marker of T-cell exhaustion – only a modest relationship was found between HIV-DNA levels and residual T-cell activation [8]. Consistent with our results, no association was found between HIV-DNA levels and residual immune activation in patients with sustained viral suppression [9].

In patients with early PHI, the strong relationship between T-cell activation and total HIV-DNA levels underlines the important role of immune activation in the establishment of the HIV reservoir. Interestingly, HIV-DNA levels in PHI patients – prior to ART – was found to predict residual immune activation/T-cell proliferation after 6 months of ART, but not immune activation levels in untreated PHI patients. It can be hypothesized that the size of the reservoir established early in PHI is one of the determinant of the residual T-cell proliferation that persists following ART [19].

The lack of association between HIV-DNA levels and T-cell activation in ART-treated patients suggests that residual immune activation is not directly dependent on the size of the latent HIV reservoir, at least in early ART-treated patients. These data highlight the role of other mechanisms, apart from HIV persistence, in maintaining elevated levels of immune activation in these patients, for example, cytomegalovirus coinfection, low thymic output, lymphoid tissue fibrosis, poor immune reconstitution, T and B-cell dysfunction, and microbial translocation [20]. Our data do not preclude a role for HIV reservoir in fueling immune activation. Of note, immune activation may conversely lead to viral persistence through several mechanisms such as increased number of target cells, homeostatic proliferation and up-regulation of PD-1, as PD-1-expressing cells are enriched in HIV-DNA [7]. The present study also provides additional evidence for early treatment of HIV infection as we found that early HIV-DNA levels predicted the level of residual peripheral T-cell proliferation under ART.

Even if the blood HIV reservoir is not directly related to peripheral T-cell activation, immune activation is likely to be involved in both the initial establishment of the reservoir and its maintenance, and is a potential barrier to HIV cure. Results from pathogenesis-oriented therapeutic interventions [21] will provide new clues on the relationship between the HIV reservoir and immune activation.

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We thank all patients involved in this study. We are grateful to Daniel Scott Algara and Françoise Barré-Sinoussi (Pasteur Institute, Paris) for helpful discussions. We thank members of the ANRS PRIMO Cohort study group and especially C. Deveau, as well as N. Desplanques, N. Valin, L. Fonquerine (Hôpital Saint Antoine, Paris); M. Karmochkine, P. Kousignian, J. Derouineau, M. Buisson, I. Pierre, D. Batisse (Hôpital Européen Georges Pompidou, Paris); L. Slama, T. Lyavanc, L. Iordache (Hôpital Tenon, Paris); M.-A. Valantin (Hôpital Pitié-Salpétrière, Paris), for including patients with PHI. We thank S. Bachir-Cherif and T. Bommenel (INSERM U943, Paris) for study monitoring; A. Aouba (Hôpital Européen Georges Pompidou, Paris), D. Bollens (CHU Saint-Antoine, Paris) for help in performing the study; and C. Katlama, A. Simon (CHU Pitié-Salpétrière, Paris), C. Goujard (CHU Bicêtre, Paris), G. Pialoux (CHU Tenon, Paris) for including patients with CHI.

The ANRS116 study group members include M. Bentata (Hôpital Avicenne, Bobigny); P.-M. Girard and J.-L. Meynard (Hôpital Saint Antoine, Paris); C. Katlama, R. Tubiana, A. Simon (Hôpital Pitié-Salpétrière, Paris); J.-F. Delfraissy, C. Gougard (Hôpital Bicêtre); A. Sobel, Y. Levy (Hôpital Henri Mondor, Créteil); G. Pialoux (Hôpital Tenon, Paris); J.-P. Viard (Hôpital Necker, Paris); B. Hoen (Hôpital Saint Jacques, Besançon); P. Yeni, R. Landmann (Hôpital Bichat, Paris); J.A. Gastaut, I. Poizot-Martin (Hôpital Sainte Marguerite, Marseille); J.M. Ragnaut (Hôpital Pellegrin, Bordeaux); P. Morlat (Hôpital Saint Jacques, Bordeaux); T. May (Hôpital Brabois, Nancy); C. Bazin (CHU de Caen); J.M. Lang (CHU Strasbourg); P. Massip (Hôpital Purpan, Toulouse); J.P. Cassuto (Hôpital de l’Archet, Nice); P. Choutet (Hôpital Bretonneau, Tours).

L.W. designed and supervised the study; L.W., M.F.C., L.A., D.C., and C.R. contributed to the experimental design and provided intellectual input; M.F.C. and C.D. performed experiments; P.-M.G. included patients and reviewed the manuscript; C.P. was the principal investigator of the ANRS116 trial; L.W., M.F.C. and L.A. analyzed data; L.W. and M.F.C. wrote the manuscript; and L.A., D.C. and C.R. participated in revision and discussion of the article. All the authors read and approved the final article.

Source of funding: This work was supported by the Agence Nationale de Recherches sur le SIDA et les Hépatites Virales (ANRS) and the Assistance Publique – Hôpitaux de Paris (AP-HP, Paris).

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Conflicts of interest

There are no conflicts of interest.

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1. Chun TW, Stuyver L, Mizell SB, Ehler LA, Mican JA, Baseler M, et al. Presence of an inducible HIV-1 latent reservoir during highly active antiretroviral therapy. Proc Natl Acad Sci U S A 1997; 94:13193–13197.
2. Finzi D, Hermankova M, Pierson T, Carruth LM, Buck C, Chaisson RE, et al. Identification of a reservoir for HIV-1 in patients on highly active antiretroviral therapy. Science 1997; 278:1295–1300.
3. Lewin SR, Rouzioux C. HIV cure and eradication: how will we get from the laboratory to effective clinical trials?. AIDS 2011; 25:885–897.
4. Deeks SG, Phillips AN. HIV infection, antiretroviral treatment, ageing, and non-AIDS related morbidity. Br Med J 2009; 338:a3172.
5. Kaplan RC, Sinclair E, Landay AL, Lurain N, Sharrett AR, Gange SJ, et al. T cell activation and senescence predict subclinical carotid artery disease in HIV-infected women. J Infect Dis 2011; 203:452–463.
6. Massanella M, Esteve A, Buzon MJ, Llibre JM, Puertas MC, Gatell JM, et al. Dynamics of CD8 T-cell activation after discontinuation of HIV treatment intensification. J Acquir Immune Defic Syndr 2013; 63:152–160.
7. Chomont N, El-Far M, Ancuta P, Trautmann L, Procopio FA, Yassine-Diab B, et al. HIV reservoir size and persistence are driven by T cell survival and homeostatic proliferation. Nat Med 2009; 15:893–900.
8. Hatano H, Jain V, Hunt PW, Lee TH, Sinclair E, Do TD, et al. Cell-based measures of viral persistence are associated with immune activation and programmed cell death protein 1 (PD-1)-expressing CD4+ T cells. J Infect Dis 2013; 208:50–56.
9. Poizot-Martin I, Faucher O, Obry-Roguet V, Nicolino-Brunet C, Ronot-Bregigeon S, Dignat-George F, Tamalet C. Lack of correlation between the size of HIV proviral DNA reservoir and the level of immune activation in HIV-infected patients with a sustained undetectable HIV viral load for 10 years. J Clin Virol 2013; 57:351–355.
10. Avettand-Fenoel V, Boufassa F, Galimand J, Meyer L, Rouzioux C. HIV-1 DNA for the measurement of the HIV reservoir is predictive of disease progression in seroconverters whatever the mode of result expression is. J Clin Virol 2008; 42:399–404.
11. Rouzioux C, Hubert JB, Burgard M, Deveau C, Goujard C, Bary M, et al. Early levels of HIV-1 DNA in peripheral blood mononuclear cells are predictive of disease progression independently of HIV-1 RNA levels and CD4+ T cell counts. J Infect Dis 2005; 192:46–55.
12. Petitjean G, Chevalier MF, Tibaoui F, Didier C, Manea ME, Liovat AS, et al. Level of double negative T cells, which produce TGF-beta and IL-10, predicts CD8 T-cell activation in primary HIV-1 infection. AIDS 2012; 26:139–148.
13. Chevalier MF, Petitjean G, Dunyach-Remy C, Didier C, Girard PM, Manea ME, et al. The Th17/Treg ratio, IL-1RA and sCD14 levels in primary HIV infection predict the T-cell activation set point in the absence of systemic microbial translocation. PLoS Pathog 2013; 9:e1003453.
14. Weiss L, Piketty C, Assoumou L, Didier C, Caccavelli L, Donkova-Petrini V, et al. Relationship between regulatory T cells and immune activation in human immunodeficiency virus-infected patients interrupting antiretroviral therapy. PLoS One 2010; 5:e11659.
15. Piketty C, Weiss L, Assoumou L, Burgard M, Melard A, Ragnaud JM, et al. A high HIV DNA level in PBMCs at antiretroviral treatment interruption predicts a shorter time to treatment resumption, independently of the CD4 nadir. J Med Virol 2010; 82:1819–1828.
16. Giorgi JV, Hultin LE, McKeating JA, Johnson TD, Owens B, Jacobson LP, et al. Shorter survival in advanced human immunodeficiency virus type 1 infection is more closely associated with T lymphocyte activation than with plasma virus burden or virus chemokine coreceptor usage. J Infect Dis 1999; 179:859–870.
17. Deeks SG, Kitchen CM, Liu L, Guo H, Gascon R, Narvaez AB, et al. Immune activation set point during early HIV infection predicts subsequent CD4+ T-cell changes independent of viral load. Blood 2004; 104:942–947.
18. Chun TW, Murray D, Justement JS, Hallahan CW, Moir S, Kovacs C, Fauci AS. Relationship between residual plasma viremia and the size of HIV proviral DNA reservoirs in infected individuals receiving effective antiretroviral therapy. J Infect Dis 2011; 204:135–138.
19. Anthony KB, Yoder C, Metcalf JA, DerSimonian R, Orenstein JM, Stevens RA, et al. Incomplete CD4 T cell recovery in HIV-1 infection after 12 months of highly active antiretroviral therapy is associated with ongoing increased CD4 T cell activation and turnover. J Acquir Immune Defic Syndr 2003; 33:125–133.
20. Klatt NR, Chomont N, Douek DC, Deeks SG. Immune activation and HIV persistence: implications for curative approaches to HIV infection. Immunol Rev 2013; 254:326–342.
21. Douek DC. Immune activation, HIV persistence, and the cure. Top Antivir Med 2013; 21:128–132.

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