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 . 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 .
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 . Consistent with our results, no association was found between HIV-DNA levels and residual immune activation in patients with sustained viral suppression .
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 .
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 . 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 . 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  will provide new clues on the relationship between the HIV reservoir and immune activation.
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).
Conflicts of interest
There are no conflicts of interest.
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