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CORRESPONDENCE Recently, Lu and Andrieu1 stated that the HIV-1
protease inhibitors (PIs) indinavir and saquinavir used in
concentrations at least 30-fold lower than those needed for 90% viral
inhibition apparently do not influence sensitivity of peripheral blood
T cells from HIV-1-infected individuals toward apoptosis induced either by T-cell receptor/CD3 ligation or by direct triggering of the
CD95 receptor in vitro.1 They conclude from their data that the beneficial effect of PI treatment on clinical and
immunological parameters in HIV-1-infected patients, which can be
encountered even in the absence of relevant virological effects, are
not related to changes in T-cell apoptosis. In our own study in a cohort of HIV-1-infected children and
adolescents, we had observed an increased sensitivity of freshly isolated T cells toward apoptosis,2 which was rapidly
down-regulated following initiation of highly active antiretroviral
therapy including PIs (HAART).3 Here we report that
prolonged PI treatment in vivo reduced sensitivity of peripheral blood
T cells toward apoptosis in vitro even when plasma viral load levels
were not decreased and susceptibility for T-cell activation in these
patients was not down-regulated. Between January 1996 and January 1999, a total of 47 HIV-1-infected
children and adolescents and 28 age-matched healthy controls were
studied. Patients were stratified according to treatment modalities at
the time of the immunological investigation (Table 1). The study was conducted according to
the Declaration of Helsinki and was approved by the ethical committee
of the University Hospital in Heidelberg. T-cell phenotyping, plasma
viral load quantification, and assessment of T-cell sensitivity toward
apoptosis and activation was performed as previously
described.3 Patients and controls and/or their relatives
gave informed consent prior to venipuncture.
We compared T-cell apoptosis and T-cell activation (CD69 expression) in
patients on HAART for more than 6 months (group III; n = 16) to
patients without therapy (group I; n = 29), patients treated with
HIV-1 reverse transcriptase inhibitors (RTIs) alone (group II;
n = 20) and healthy controls (CTRLs; n = 28) by analysis of
variance using the InStat statistical analysis software package (GraphPad Software, San Diego, CA). As shown in Figure
1, the sensitivity of T cells toward
anti-CD95-induced apoptosis was significantly lower in group III than
in group II (Figure 1B; CD4+ T cells, 3 ± 1% versus
26 ± 5%, P < .001; CD8+ T cells,
5 ± 1% versus 24 ± 5%, P < .001) despite similar
plasma viral load levels in both patient groups. Spontaneous T-cell
death did not differ between these groups (Figure 1A; CD4+
T cells, 10 ± 2% versus 17 ± 4%, P = NS;
CD8+ T cells, 12 ± 3% versus 15 ± 2%,
P = NS).
Although T cells from HAART-treated patients (group III) had a normal sensitivity toward anti-CD95-induced apoptosis in vitro (P = NS when compared to the control; Figure 1B), the sensitivity toward anti-CD3-induced CD69 expression in both CD4+ and CD8+ T-cell subpopulations was significantly higher in group III than in all other groups of patients and the controls (Figure 1E, P < .001). Anti-CD3-induced CD69 expression on CD4+ and CD8+ T cells tended to increase in HAART-treated patients (group III) with detectable, compared to those with undetectable, plasma viral load, although these differences were statistically not significant (CD4+ T cells, 71 ± 8% versus 51 ± 7%, P = NS; CD8+ T cells, 69 ± 8% versus 52 ± 6%, P = NS). Spontaneous CD69 expression in T cells from PI-treated patients in group III was increased compared to other treatment groups and healthy controls (28 ± 7% on CD4+ and 27 ± 4% on CD8+ T cells; P < .001); see Figure 1D. Our observations suggest that the reduced T-cell sensitivity toward CD95-induced apoptosis in vitro might partly explain the observation that an increasing number of patients with virological rebound on PI therapy maintain stable CD4 counts and do not experience clinical deterioration as fast as one could expect from the increased viral load. The CD95-resistant phenotype of peripheral blood T cells in these patients may be due to a decreased viral "fitness" or a direct inhibition of apoptosis pathways (eg, at the level of proapoptotic proteases).4 The fact that these T cells are still hypersensitive toward anti-CD3-induced activation argues against the hypothesis of Lu and Andrieu that the restoration of a normal apoptosis sensitivity is simply an epiphenomenon of the general immunological recovery in HIV-infected individuals receiving HAART.
Thomas Böhler and Klaus-Michael Debatin
Uwe Wintergerst
References
1.
Lu W, Andrieu JM.
HIV protease inhibitors restore impaired T-cell proliferative response in vivo and in vitro: a viral-suppression-independent mechanism.
Blood.
2000;96:250-258
2.
Bäumler CB, Böhler T, Herr I, Benner A, Krammer PH, Debatin KM.
Activation of the CD95 (APO-1/Fas) system in T cells from human immunodeficiency virus type-1-infected children.
Blood.
1996;88:1741-1746 3. Böhler T, Walcher J, Hölzl-Wenig G, et al. Early effects of antiretroviral combination therapy on activation, apoptosis and regeneration of T cells in HIV-1-infected children and adolescents. AIDS. 1999;13:779-789[CrossRef][Medline] [Order article via Infotrieve].
4.
Sloand EM, Kumar PN, Kim S, Chaudhuri A, Weichold FF, Young NS.
Human immunodeficiency virus type 1 protease inhibitor modulates activation of peripheral blood CD4+ T cells and decreases their susceptibility to apoptosis in vitro and in vivo.
Blood.
1999;94:1021-1027
Response:T-cell recovery in HIV-infected patients experiencing virologic failure under highly active antiretroviral therapyBöhler et al's comments invoke the fundamental issue: what is the interrelation between T-cell activation, anergy, and apoptosis (AAA) in HIV-infected patients treated with highly active antiretroviral therapy (HAART). We agree that this issue is important enough to deserve a deeper discussion. Recently, we1 and others2-5 have reported an
apparent dissociation between T-cell recovery and plasma viral-load
suppression in HIV-infected patients under HAART. In addition,
Böhler et al also observed a reduced sensitivity of HAART-treated
patients' T cells to apoptosis that was independent from viral-load
response. This dissociation implies a direct effect of HAART on
patients' immune systems rather than "a decreased viral fitness"
as suggested by Böhler et al, because the immune recovery in
patients experiencing virologic failure could not be maintained after
withdrawal of HAART (Table 1).
In general, the immune recovery under HAART is characterized by an increase of CD4+ and CD8+ T-cell counts associated with an overall down-regulation of T-cell AAA in vivo. To learn what the effects of HAART on the AAA are, we have conducted an in vitro study to investigate the AAA kinetics of patients' peripheral blood mononuclear cells (PBMCs) following immune stimuli in the absence or the presence of HAART components (nucleoside reverse trascriptase inhibitors [NRTIs] or protease inhibitors [PIs]).1 Results showed that PIs (indinavir or saquinavir), even at nonantiviral doses, allowed an early (4 days after activation) reversal of T-cell anergy (ie, restoration of T-cell proliferation) but a later (14 days after activation) down-regulation of T-cell apoptosis. Therefore, we concluded that reversal of T-cell anergy should be the primary action of PIs responsible for the overall immune improvements observed in patients treated with HAART. This finding is in keeping with the increasing evidence that the progression of HIV disease is associated with an impaired T-cell proliferation/production (anergic mechanism) coupled with chronic immune activation and apoptosis (ie, reduced half-life of T cells) rather than with an increased T-cell turnover (overproduction or exhaustion hypothesis).6-10 An increased proportion of HIV-infected patients' T cells undergoing apoptosis following APO-1/Fas (CD95) ligation reflects, indeed, a subpopulation of T cells chronically activated in vivo, because purified quiescent T cells (G0) showed nearly 100% resistance to apoptosis triggered by APO-1/Fas ligation while the proportion of T cells susceptible to Fas-mediated apoptosis increased gradually after 7 days of stimulation with anti-CD3 and anti-CD28 monoclonal antibodies (Lu and Andrieu, unpublished observation, 2000). Normalized sensitivity of T cells to apoptosis in PI-treated (but not NRTI-treated) patients as reported by Böhler et al could thus be the consequence of early restoration of T-cell proliferation and subsequent down-regulation of T-cell activation by PIs. Böhler et al's key argument challenging the normalization of T-cell proliferation/activation as the primary event involved in HAART (ie, PI)-induced immune recovery is based on their observation that CD69 expression on T cells following TCR/CD3 stimulation was significantly enhanced in HAART-treated patients. In fact, this observation supports further the PI-mediated enhancement of T-cell proliferation/differentiation following immune stimuli, because CD69 expression on T cells upon to TCR/CD3 stimulation was shown to be correlated positively with proliferation and effector function of T cells.11 In conclusion, anergized status of HIV-infected patients' T cells
permits unresolved chronic activation and apoptosis throughout the
course of infection, thus contributing to the eventual development of
AIDS. PIs, even at nonantiviral concentrations, are capable of
restoring T-cell proliferation and in consequence
normalizing
Wei Lu and Jean-Marie Andrieu
References 1. Lu W, Andrieu JM. HIV protease inhibitors restore impaired T-cell proliferative response in vivo and in vitro: a viral-suppression-independent mechanism. Blood. 2000;96:250-258. 2. Kaufmann D, Pantaleo G, Sudre P, Telenti A. CD4-cell count in HIV-1-infected individuals remaining viraemic with highly active antiretroviral therapy (HAART): Swiss HIV cohort study. Lancet. 1998;351:723-724[CrossRef][Medline] [Order article via Infotrieve].
3.
Levitz SM.
Improvement in CD4+ cell counts despite persistently detectable HIV load.
N Engl J Med.
1998;338:1074-1075 4. Ledergerber B, Egger M, Opravil M, et al. Clinical progression and virological failure on highly active antiretroviral therapy in HIV-1 patients: a prospective cohort study: SwissHIV cohort study. Lancet. 1999;353:863-868[CrossRef][Medline] [Order article via Infotrieve].
5.
Grabar S, Moing VL, Goujard C, et al.
Clinical outcome of patients with HIV-1 infection according to immunologic and virologic response after 6 months of highly active antiretroviral therapy.
Ann Intern Med.
2000;133:401-410 6. Andersson J, Fehniger TE, Patterson BK, et al. Early reduction of immune activation in lymphoid tissue following highly active HIV therapy. AIDS. 1998;12:F123-F129[Medline] [Order article via Infotrieve]. 7. Bofill M, Borthwick NJ, Simmonds HA. Novel mechanism for the impairment of cell proliferation in HIV-1 infection. Immunol Today. 1999;20:258-261[CrossRef][Medline] [Order article via Infotrieve]. 8. Hellerstein M, Hanley MB, Cesar D, et al. Directly measured kinetics of circulating T lymphocytes in normal and HIV-1-infected humans. Nat Med. 1999;5:83-89[CrossRef][Medline] [Order article via Infotrieve]. 9. McCune JM, Hanley MB, Cesar D, et al. Factors influencing T-cell turnover in HIV-1-seropositive patients. J Clin Invest. 2000;105:R1-R8.
10.
Fleury S, Rizzardi GP, Chapuis A, et al.
Long-term kinetics of T cell production in HIV-infected subjects treated with highly active antiretroviral therapy.
Proc Natl Acad Sci U S A.
2000;97:5393-5398 11. Nielsen SD, Afzelius P, Ersboll AK, Nielsen JO, Hansen JE. Expression of the activation antigen CD69 predicts functionality of in vitro expanded peripheral blood mononuclear cells (PBMC) from healthy donors and HIV-infected patients. Clin Exp Immunol. 1998;114:66-72[CrossRef][Medline] [Order article via Infotrieve].   CiteULike  Connotea  Del.icio.us  Digg  Reddit  Technorati What's this?
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| Copyright © 2001 by American Society of Hematology Online ISSN: 1528-0020 | |||||||||
indicates CD4+ T
cell subpopulations, and 
, CD8+ T cell subpopulations.
Horizontal bars indicate the group median. Data from HIV-1-infected
patients receiving either no effective antiretroviral therapy (group I,
n = 26) or at least 6 months of treatment with 2 inhibitors of HIV-1
reverse transcriptase (RTIs; group II, n = 20) or 2 RTIs plus at
least 1 inhibitor of HIV-1 protease (HAART; group III, n = 16) were
compared to 28 age-matched healthy controls (CTRLs). The respective
patient numbers for measurements of CD69 expression were group I,
n = 11; group II, n = 20; group III, n = 16; and CTRL, n = 12.
Methods: Freshly isolated peripheral blood mononuclear cells (PBMCs)
(2 × 105 cells/well) were cultured in the presence or
absence of either anti-CD3 monoclonal antibodies (moAbs; OKT3; 10 µg/mL) or anti-CD95 moAb (anti-APO-1; 10 µg/mL) and protein A (5 ng/mL) as described.
% of spontaneous cell
death)/(100