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Journal of Leukocyte Biology logoLink to Journal of Leukocyte Biology
. 2008 Sep 19;84(6):1447–1453. doi: 10.1189/jlb.0708438

Immune activation and IL-12 production during acute/early HIV infection in the absence and presence of highly active, antiretroviral therapy

Adriana A Byrnes *,†, David M Harris *,†, Sowsan F Atabani ‡, Beulah P Sabundayo *,†, Susan J Langan †, Joseph B Margolick †, Christopher L Karp *,†,‡,1
PMCID: PMC2614601  PMID: 18806124

Abstract

Suppressed IL-12 production and maladaptive immune activation, both of which are ameliorated by successful highly active antiretroviral therapy (HAART), are thought to play important roles in the immunopathogenesis of chronic HIV infection. Despite the important effects of the immunological and virological events of early HIV infection on subsequent disease progression, IL-12 production and immune activation in early infection remain under-defined. To quantify IL-12 production and immune activation during acute/early HIV infection, in the presence and absence of HAART, we performed a prospective, longitudinal study of participants in the Baltimore site of the Acute Infection and Early Disease Research Program, with cross-sectional comparison to healthy control subjects. PBMC cytokine productive capacity and plasma immune activation markers [soluble CD8 (sCD8), sCD4, granzyme B, neopterin, β2-microglobulin, sIL-2R, sTNFRI, sTNFRII, and IL-12p70] were quantified by ELISA. Notably, PBMC from patients with acute/early HIV infection exhibited in vivo IL-12p70 production along with increased, maximal in vitro IL-12 production. Further, despite evidence from plasma markers of generalized immune activation, no elevation in plasma levels of sCD4 was observed, suggesting relative blunting of in vivo CD4+ T cell activation from the beginning of HIV infection. Finally, despite successful virological responses to HAART, heightened in vivo CD8+ T cell activation, IL-12 production, and IFN activity were sustained for at least 6 months during primary HIV infection. These data underscore the need for comparative mechanistic analysis of the immunobiology of early and chronic HIV infection.

Keywords: regulation, cytokine, interferon

INTRODUCTION

Despite extensive literature about the immunology of chronic HIV infection, considerably less is known about the immunology of very early infection. Although there are obvious practical reasons for this gap in knowledge, the determining role played by the virological and immunological milieu of early infection on the progression of HIV disease [1,2,3,4] provides a compelling reason for focusing experimental attention on early HIV infection. Dysregulated IL-12 production and maladaptive immune activation, both ameliorated by successful highly active antiretroviral therapy (HAART), are thought to play important roles in the immunopathogenesis of chronic HIV infection. However, IL-12 production and immune activation in early disease, in the presence and absence of HAART, remain under-defined.

IL-12 is a key regulator of cell-mediated immunity (CMI) [5]. The functional cytokine (IL-12p70) is a heterodimer. Expression of the IL-12p40 chain, a subunit also used by the related heterodimeric cytokine IL-23, is tightly regulated in cells (principally, APCs) that secrete IL-12p70. The IL-12p35 chain is constitutively expressed at low levels by many cell types but tightly regulated in cells that produce the functional cytokine. IL-12p40 and IL-12p70 are secreted, the former in considerable excess of the latter. IL-12 secretion is induced by a variety of microbes and microbial products, as well as by interactions between APCs and activated T cells. Among its various activities, IL-12 potently induces IFN-γ production from lymphocytes and NK cells; is central to Th1 differentiation in many systems; amplifies NK cell cytotoxicity and enhances the generation of cytotoxic T lymphocytes; and regulates delayed type hypersensitivity (DTH) responses [5]. As might thereby be expected, IL-12 plays a central role in host resistance in models of infection with numerous pathogens that cause opportunistic infection in HIV/AIDS [6].

Although functional abnormalities are found in essentially every compartment of the immune system in HIV/AIDS, defects in CMI appear to be of the greatest clinical import. Suppression of CMI is evident in vivo from the range of infections to which HIV infection predisposes, as well as from a failure to mount DTH responses to recall antigens [7]. In vitro correlates include progressive failure of proliferation and IFN-γ production in response to recall antigens, alloantigens, and mitogens by CD4+ T cells; decreased cytolytic function of CD8+ T cells; and decreased proliferation and IFN-γ production by NK cells [8,9,10,11,12,13]. Given the close match between the abnormalities of CMI seen in HIV/AIDS and the known functions of IL-12, the role of IL-12 deficiency in HIV pathogenesis has received considerable attention.

In patients with chronic HIV infection, PBMC are specifically impaired in their ability to produce IL-12 in response to several bacterial and protozoal stimuli [14,15,16,17,18]. Moreover, in vitro treatment of PBMC from such patients with IL-12 ameliorates defective T cell-proliferative responses, enhances T cell production of IL-2, enhances T and NK cell production of IFN-γ, increases NK and CD8+ T cell cytotoxic activity, protects T cells from activation-induced apoptosis, and enhances HIV-specific responses [12, 19,20,21,22]. These data have suggested a likely role for defective IL-12 production in the pathogenesis of HIV-associated immunodeficiency, as well as in inefficient immune responses to HIV itself. Although blunted IL-12 responses are evident as early as they have been assessed in chronic HIV infection (i.e., prior to marked declines in peripheral CD4+ T cell counts), there are few reports of IL-12 production or productive capacity during acute/early HIV infection.

As for maladaptive immune activation, there is a growing consensus that dysregulated, generalized T cell activation plays a central role in the pathogenesis of HIV/AIDS [23,24,25,26,27]. Heightened immune activation, as defined by quantification of immunophenotypic markers on T cells (e.g., HLA-DR, CD69, CD38) or of soluble markers in plasma [e.g., soluble (s)CD8, sIL-2R, β2-microglobulin, sTNFRI, sTNFRII, neopterin], has been associated with the CD4+ T cell depletion rate, disease progression, and risk of mortality, independent of plasma viral load. Bystander killing, accelerated senescence, and increased activation-induced apoptosis have all been proposed as mechanisms underlying activation-induced CD4+ T cell depletion [23, 27,28,29,30]. However, the cellular and molecular details of the link(s) between immune activation and CD4+ T cell depletion remain unclear, something underscored by the fact that cellular markers of CD8+ T cell activation appear to have an especially strong (negative) prognostic significance [31]. Generalized immune activation and CD8+ T cell activation decrease with successful HAART, events that correlate with CD4+ T cell reconstitution in chronic infection [32,33,34]. Of interest, although immune activation, broadly taken, is clearly important for the initial host response to HIV infection, the level of CD8+ T cell activation in acute/early infection has been correlated with subsequent disease progression, independently of plasma viral load, in untreated patients [4, 26].

To further define immunoregulatory cytokine production and immune activation in acute/early HIV infection and the effects of HAART on these immunological events, we prospectively studied a cohort of adults with acute or early HIV infection.

MATERIALS AND METHODS

Subjects

Forty-eight patients were enrolled from the Baltimore site of the Acute Infection and Early Disease Research Program (Baltimore, MD, USA) [35], in which the following criteria were used to document recent infection: a negative ELISA with detectable plasma HIV RNA; a positive ELISA with a documented negative ELISA in the preceding 5 weeks; a positive ELISA with a negative or evolving Western blot; an indeterminate Western blot, evolving in the presence of detectable plasma HIV RNA; a value <0.75 in the de-tuned (less-sensitive) ELISA [36] with a history consistent with the acute retroviral syndrome in the preceding 4 weeks; a positive ELISA in the presence of a negative (OD <0.75), de-tuned ELISA; and/or a positive ELISA with a documented negative ELISA within 1 year. Fourteen patients meeting ≥1 of criteria “a–e” were judged to have acute HIV infection (≤2 months); 34 patients meeting criteria “f” and/or “g” were judged to have early HIV infection (2–12 months). Patient characteristics, including baseline HIV viral loads and CD4+ T cell counts, and presumptive length of infection prior to enrollment are listed in Table 1. There were no significant differences in age, sex distribution, or CD4+ T cell counts between patients with acute and early HIV infection; patients with acute HIV infection had significantly higher plasma viral loads. As expected [24], there was a significant, negative correlation between plasma HIV RNA levels and CD4+ T cell counts in patients (Pearson correlation=–0.413; P=0.0035).

TABLE 1.

Baseline Subject Characteristics

Study population Number Age Sex (M/F) Plasma HIV RNA level, log10 copies/ml CD4+ T cells/ul
All HIV patients 48 38.5 (33.5–44.5) 29/19 4.69 (4.08–5.29) 442.5 (374.5–579)
Acute Infection 14 40.5 (34–45) 7/7 5.44 (5.00–5.91)a 420 (380–484)
Early Infection 34 37.5 (33–44) 22/12 4.33 (3.96–5.08) 485.5 (372–594)
Control subjects 21 31 (26–35.3)b 11/10 N/A NP
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Median and interquartile ranges are shown. 

a

P < 0.05 compared with patients with early HIV infection; 

b

P < 0.05 compared with all patient groups. P values were calculated with the Mann-Whitney U-test. Acute infection: <2 months; early infection: 2–12 months. N/A, Not applicable; NP, not performed. 

All enrolled patients were offered HAART; approximately half elected such therapy. Those electing HAART had similar demographics as those declining HAART but tended to have higher plasma HIV RNA levels [median 5.20 (interquartile range 4.11–5.56) vs. 4.40 (interquartile range 4.08–5.22) log10 copies/ml; P=0.06] and had significantly lower CD4+ T cell counts [median 381 (interquartile range 280–692) vs. median 487 (interquartile range 414–588) CD4+ T cells/μl; P=0.027]. Any HAART regimen recommended by the U.S. Department of Health and Human Services during the period of this study (1998–2002) was allowed. Eighteen patients on HAART were followed for at least 6 months after HAART initiation; 13 were responders, achieving undetectable plasma HIV RNA levels by 6 months of therapy.

A cohort of healthy controls was enrolled from among the staff of the Johns Hopkins University Schools of Medicine and Public Health (Baltimore, MD, USA). As shown in Table 1, controls were somewhat (if significantly) younger than patients.

Written, informed consent was obtained from all subjects; studies were approved by the Johns Hopkins University Bloomberg School of Public Health Institutional Review Board (patients) and the Johns Hopkins University School of Medicine Institutional Review Board (controls and patients).

Measurement of HIV viral load, HIV serologies, peripheral CD4+ T cell count

Plasma HIV viral load was quantified using the Amplicor HIV-1 monitor assay (Roche Molecular Systems, Branchburg, NJ, USA) with standard (400–750,000 copies/ml) and/or ultrasensitive (50–75,000 copies/ml) ranges of quantification. CD4+ T cell percentages were measured by flow cytometry; absolute CD4+ T cell numbers were calculated from an automated, complete blood count and differential.

Isolation of plasma; isolation and culture of PBMC

Phlebotomy was performed at baseline and 6 months after enrollment. Plasma was isolated from EDTA anticoagulated blood and stored at –80°C until assayed. PBMC were isolated from heparinized blood by centrifugation over Ficoll-Hypaque (Nycomed, Princeton, NJ, USA). Freshly isolated cells were cultured in 48-well plates at a density of 1.5 × 106/ml in RPMI 1640 containing 2 mM L-glutamine (both from Invitrogen, Carlsbad, CA, USA), and 50 mM gentamicin and 5% human AB serum (both from Sigma Chemical Co., St. Louis, MO, USA). All tissue-culture reagents were endotoxin-free to the limits of detection of the Limulus amoebocyte lysate assay (Cambrex Bioscience, Walkersville, MD, USA). Cells were stimulated for 24 h with 0.01% fixed Staphylococcus aureus Cowan strain I (SAC; Calbiochem, San Diego, CA, USA) or 1 μg/ml Salmonella typhimurium LPS (Sigma Chemical Co.) in the presence or absence of 300 IU/ml human recombinant (r)IFN-γ (BD PharMingen, San Diego, CA, USA). Cell-free supernatants were harvested and stored at –80°C until assayed. As samples were limiting, not all assays were performable on all samples.

Cytokine and activation marker assays

SAC-stimulated IL-12p70 secretion was measured using the high-sensitivity ELISA (Quantikine) kit from R&D Systems (Minneapolis, MN, USA), having a sensitivity of 0.1–0.4 pg/ml. SAC plus IFN-γ- (SAC/IFN-γ) and LPS/IFN-γ-stimulated IL-12p70 secretion was measured by commercially available antibody pairs and recombinant standards from R&D Systems, with a limit of sensitivity of 2–4 pg/ml. Although different rIL-12p70 standards were used in these different IL-12p70 ELISAs, side-by-side comparison of dilutional curves of such standards using the Quantikine kit revealed their essential equivalence (data not shown). IL-12p40 secretion was measured by the OptEIA kit (BD PharMingen). IL-10 and TNF were measured by ELISA using commercially available antibody pairs and recombinant standards from BD PharMingen.

ELISA assays were used to quantify plasma levels of sCD4, sCD8, sIL-2R (Endogen, Rockford, IL, USA), sTNFR-I, sTNFR-II, β2-microglobulin, IL-12p70 (Quantikine, R&D Systems), neopterin (Immuno-Biological Laboratories, Hamburg Germany), and granzyme B (CLB, Amsterdam, Netherlands).

Statistical analysis

Statistical comparisons between patient groups and between patient and control groups were performed using the nonparametric Mann-Whitney U test or the Kruskal-Wallis test, as appropriate, using Statview software (SAS, Cary, NC, USA). Baseline associations between variables were evaluated using Pearson correlation statistics (SAS software). Values below the ELISA threshold (a consideration only for plasma IL-12p70) were assigned a value = 50% of the highest threshold value for analytical purposes.

RESULTS

Baseline IL-12 production in acute/early HIV infection

To characterize the IL-12-productive capacity of PBMC from patients with acute/early HIV infection at the time of enrollment, we used bacterial stimuli: SAC, a fixed preparation of S. aureus, and LPS from S. typhimurium. These stimuli were used in the absence and/or presence of IFN-γ, which primes for peak IL-12p70 secretion. As shown in Table 2, although SAC-stimulated production of IL-12p70 was blunted in PBMC from patients with acute HIV infection compared with uninfected controls (mirroring findings in chronic HIV infection), PBMC from patients with acute infection had no deficit in IL-12p70 production in response to IFN-γ/SAC, as well as significantly increased production of IL-12p70 in response to IFN-γ/LPS; PBMC from patients with early HIV infection secreted normal amounts of IL-12p70 in response to SAC alone, along with increased amounts of IL-12p70 in response to IFN-γ/SAC and IFN-γ/LPS; IL-12p40 production largely followed that of IL-12p70; and no differences in TNF or IL-10 production were seen between either patient group and healthy controls. Notably, this increased, maximal, IL-12-productive capacity by PBMC ex vivo was mirrored in vivo: There was significant elevation in plasma IL-12p70 levels in the groups of patients with acute (n=12) and early (n=30) HIV infection compared with healthy controls (n=21)—none of the latter of which had detectable plasma IL-12p70 (Fig. 1A). Thus, unlike the published findings in chronic HIV infection, patients with acute/early HIV infection exhibited evidence of in vivo IL-12p70 production, as well as increased, maximal IL-12 production by their PBMC ex vivo.

TABLE 2.

Cytokine-Productive Capacity of PBMC in Patients and Control Subjects

Cytokine Stimulus Controls Acute HIV Early HIV
IL-12p70 SAC 8.6 (4.2–19.8) 2.3 (0.6–4.5)a 5.1 (1.5–16.5)
IFN/SAC 12.8 (2.7–51.0) 20.5 (3.4–33.7)b 118.9 (65.1–185.2)c
IFN/LPS 9.8 (2.0–28.4) 80.3 (39.7–92.1)c 86.6 (37.8–397.7)d
IL-12p40 SAC 1312 (1078–1486) 603 (499–995)c 886 (718–1878)
IFN/SAC 3086 (2292–3846) 2196 (1794–2943) 2394 (1885–4240)
IFN/LPS 1284 (908–1547) 3000 (2314–3796)a 2658 (1554–3838)c
TNF SAC 701 (342–1153) 623 (410–788) 699 (234–1008)
IL-10 SAC 2836 (1694–5663) 3616 (2946–9644) 4957 (2132–8562)
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Median and interquartile ranges of secreted cytokines (pg/ml) are shown. 

a

P < 0.005 compared with controls; 

b

P < 0.005 compared with patients with early HIV infection. 

c

P < 0.05; 

d

P = 0.05 compared with controls. P values were calculated with the Mann-Whitney U-test. 

Fig. 1.

Fig. 1.

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Baseline plasma levels of IL-12 and other markers of immune activation in acute and early HIV infection. (A) IL-12p70; (B) sCD8; (C) sCD4; (D) granzyme B; (E) neopterin; (F) β2-microglobulin; (G) sIL-2R; (H) sTNFRI; (I) sTNFRII. These cross-sectional data, stratified by length of infection at the time of enrollment, are shown as box plots representing medians; 10th, 25th, 75th, and 90th percentiles; and outlying values (○). ‡, P < 0.0001; *, P < 0.05; §, P < 0.0005; , P < 0.0005 compared with controls; acute, Acute HIV infection (<2 months); early, early HIV infection (2–12 months); NC, normal controls.

Baseline plasma activation markers in acute/early HIV infection

We also quantified baseline plasma levels of a panel of commonly studied immune activation markers, including markers of in vivo: CD8+ T cell activation (sCD8); CD4+ T cell activation (sCD4); cytolytic activity (granzyme B); IFN activation of monocyte/macrophages and dendritic cells (neopterin) [37,38,39]; hematopoietic cell activation in response to remarkably diverse stimuli (β2-microglobulin, sIL-2R); and TNF production (sTNFRI and sTNFRII). Compared with healthy controls, patients with acute HIV infection had significantly increased levels of all activation markers except for sCD4 (Fig. 1, B–I), providing evidence for a relative deficit in CD4+ T cell activation, even at the onset of HIV infection. Compared with acute HIV infection, early HIV infection was associated with a general decline in all activation markers, something observed with plasma viral load as well (Table 1). Levels of only two such markers—granzyme B and sTNFRI—became statistically indistinguishable from controls, however (Fig. 1). Thus, CD4+ T cell activation appears to be relatively deficient even in acute/early infection, and the progression from acute to early infection is associated with a general decline in markers of immune activation—although few such markers normalize.

Effect of HAART on IL-12 production and plasma activation markers

Phlebotomy was repeated 6 months after enrollment in 29 of the 48 patients. This subset included: 18 patients followed while receiving HAART (eight with acute infection; 10 with early infection), of whom 13 (six acute; seven early) had undetectable viral loads by 6 months (responders) and 11 patients (one acute, 10 early) not receiving antiretroviral therapy (natural history).

As shown in Figure 2A, plasma IL-12p70 remained elevated significantly in both groups of HIV-infected patients, compared with healthy controls. Notably, plasma IL-12p70 levels remained elevated even in HAART responders (median: 0.3 pg/ml; P=0.0002). As for ex vivo IL-12 production by PBMC, although SAC- and IFN-γ/SAC-stimulated IL-12p70 secretion were normal in those receiving HAART, IFN-γ/LPS-driven IL-12p70 secretion remained elevated significantly (data not shown), even in HAART responders (median: 157.9 pg/ml; P=0.01). Too few natural history samples were available for analysis of ex vivo IL-12-productive capacity. Thus, the early stages of HIV infection are associated with increased IL-12 production in vivo and ex vivo, despite successful HAART.

Fig. 2.

Fig. 2.

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Plasma IL-12 and activation marker levels 6 months after enrollment, stratified by a therapeutic group. (A) IL-12p70; (B) sCD8; (C) neopterin. Data are shown as box plots representing medians; 10th, 25th, 75th, and 90th percentiles; and outlying values. ‡, P < 0.005; , P < 0.0001; §, P < 0.0005; *, P < 0.05 compared with controls. Nat His, Natural history patients not receiving HAART.

As for other plasma activation markers at 6 months, sCD4, sTNFRI, and granzyme B levels were indistinguishable from controls in HAART and natural history patients (data not shown), as expected from the fact that these markers failed to be activated at time of presentation (sCD4) or were no longer activated in patients with early disease (sTNFRI, granzyme B). Although no other activation markers normalized in the absence of HAART, such therapy led to normalization of plasma levels of sIL-2R and sTNFRII (data not shown). Although plasma levels of β2-microglobulin failed to normalize in patients receiving HAART, taken as a group, such levels did normalize in HAART responders (those with undetectable plasma HIV RNA; data not shown). Of note, plasma concentrations of sCD8 and neopterin remained elevated significantly, even in HAART responders (median sCD8=431 pg/ml; P=0.0002; median neopterin=3.7 ng/ml; P<0.05; Fig. 2, B and C). Thus, although successful HAART leads to normalization of several markers of immune activation, systemic evidence of heightened CD8+ T cell activation and IFN activity persists, despite the lack of detectable viremia.

DISCUSSION

The data presented here provide further insights into immunoregulatory cytokine production and immune activation in acute/early HIV infection. Of note, we demonstrate that patients with acute/early HIV infection exhibit in vivo IL-12p70 production, as well as increased, maximal, IL-12-productive capacity by their PBMC in vitro. Extending previous observations [40], our data show relative blunting of CD4+ T cell activation from the beginning of HIV infection. Finally, our data underscore the fact that, despite clearance of detectable viremia, successful HAART during early stages of infection is associated with continued in vivo CD8+ T cell activation along with IFN activity and IL-12p70 production.

As opposed to the blunted, IL-12-productive capacity observed with chronic HIV infection, acute/early HIV infection is associated with heightened, maximal, IL-12-productive capacity ex vivo and increased IL-12 production in vivo. These in vivo data appear to have been prefigured by a recent report in which plasma IL-12 concentrations were found to be “very low” during acute infection [41]. Despite this interpretation, the fact that healthy controls did not have detectable IL-12p70 in plasma in that or the present study indicates that even the very low levels detected in people with acute HIV infection represent an increase over levels present in healthy controls. Indeed, our data about in vivo IL-12 production mirror recent findings that IL-12p70 concentrations are increased in genital tract fluids of women with acute, heterosexually acquired HIV infection [42], and that gut-associated lymphoid tissue from patients with acute HIV infection has increased numbers of cells expressing IL-12p70 [43]. Unlike the current data, no differences in plasma IL-12 concentrations were detected between patients and controls in the former study [42]. Whether this is a result of the less-sensitive ELISA used in the study of Bebell et al. [42] or a result of differences in immunopathogenesis attendant on route of infection remains unclear. We are unaware of any previous reports about the IL-12-productive capacity of PBMC during early HIV infection.

In chronic HIV infection, defects in bacterially stimulated PBMC production of IL-12 can be overcome by IFN-γ priming and/or costimulation with the CD40 ligand [44]. Although we found that SAC-driven IL-12 production was inhibited in PBMC from patients with acute (but not early) HIV, we also found that IFN-γ priming led to elevated IL-12 production in response to LPS stimulation (in acute and early infection and after HAART) and SAC stimulation (in early infection). This increased, maximal, IL-12-productive capacity, which is not seen during acute infection with the second-most common viral cause of immunosuppression-induced mortality in the world at large—measles virus [45]— remains mechanistically undefined. Given the sustained increases observed in plasma neopterin levels, in vivo priming with type I IFNs is an attractive mechanistic possibility. The IFN-α/β-productive capacity of PBMC is profoundly impaired during primary HIV infection, likely as a result of decreases in circulating plasmacytoid dendritic cells [46]. However, acute HIV infection is associated with in vivo IFN-α/β production, something that increases with chronic and progressive disease [47, 48]. The effects of such type I IFNs on IL-12 are complex. Although TLR-driven induction of IL-12 depends on autocrine/paracrine production of IFN-β, which acts to up-regulate IL-12p35 mRNA expression [49], prolonged, robust exposure to type I IFNs inhibits IL-12p40 transcription and IL-12p70 secretion [50, 51]. It is thus tempting to suggest the hypothesis that type I IFN production is mechanistically linked to early augmentation and later suppression of IL-12 production during HIV infection.

Despite the likely role of sustained immune activation in the pathogenesis of HIV disease, prompt mobilization of the immune system is likely necessary for any successful host response to HIV infection. Our demonstration of widespread immune activation during acute HIV infection, along with a general decline in immune activation markers with progression to early infection, is not novel [4, 28, 37, 52,53,54,55,56]. However, our demonstration that acute/early HIV infection is not accompanied by an evident increase in plasma sCD4 is of interest. Notably, significant (approximately twofold mean) increases in plasma concentrations of sCD4 have been found during acute infection with viruses that are cleared by the host (e.g., measles virus [57, 58]) as well as with viruses that are not cleared but are controlled by the host (e.g., EBV [59]). Our finding that CD4+ T cell activation appears blunted from the beginning of HIV infection is consistent with previous reports of absent ex vivo HIV protein-specific, proliferative responses in acute disease [40], and decreased CD38 expression on CD4+ compared with CD8+ T cells in acute/early HIV infection [4, 28]; however, further interpretation of these latter studies is limited by the lack of a direct comparison with CD38 expression levels on CD4 cells from healthy controls. Given the necessity for CD4+ T cell help for effective, sustained CD8+ T cell cytolytic function, any early decrement in CD4+ T cell activation is likely of pathogenetic significance. As with our finding of increased IL-12 production, the mechanisms responsible for blunted CD4+ T cell activation in early disease remain to be defined.

Although successful virological responses to HAART during chronic HIV infection are associated with normalization of many markers of immune activation, markers of CD8+ T cell activation remain elevated [32]. Our data indicate that elevated CD8+ T cell activation persists after successful virological responses to HAART during acute/early HIV infection as well and is accompanied by increased IFN activity. This continued CD8/IFN signature, despite a lack of detectable viremia, may well be a reflection of ongoing anti-HIV immune responses in tissue viral compartments not accessible by phlebotomy. The recent demonstration that HIV infection is associated with sustained systemic exposure to microbial products [60] suggests an additional, complementary hypothesis: that such products themselves drive IFN production and CD8+ T cell activation. Mechanistic attention to this possibility as well as to the link among CD8+ T cell activation, CD4+ T cell loss, and HIV disease progression may well lead to novel, therapeutic approaches to HIV infection.

Acknowledgments

This study was supported by grants from the National Institutes of Health (R01DE12167 to C. L. K.; U01AI141532 to J. B. M.). We thank the subjects for their participation; Dr. Joel Gallant and Arlette Lindsay, PA, for patient care; and Stacey Meyerer for laboratory assistance.

References

  1. Lyles R H, Munoz A, Yamashita T E, Bazmi H, Detels R, Rinaldo C R, Margolick J B, Phair J P, Mellors J W. Natural history of human immunodeficiency virus type 1 viremia after seroconversion and proximal to AIDS in a large cohort of homosexual men. Multicenter AIDS Cohort Study. J Infect Dis. 2000;181:872–880. doi: 10.1086/315339. [DOI] [PubMed] [Google Scholar]
  2. Schacker T W, Hughes J P, Shea T, Coombs R W, Corey L. Biological and virologic characteristics of primary HIV infection. Ann Intern Med. 1998;128:613–620. doi: 10.7326/0003-4819-128-8-199804150-00001. [DOI] [PubMed] [Google Scholar]
  3. Pantaleo G, Demarest J F, Schacker T, Vaccarezza M, Cohen O J, Daucher M, Graziosi C, Schnittman S S, Quinn T C, Shaw G M, Perrin L, Tambussi G, Lazzarin A, Sekaly R P, Soudeyns H, Corey L, Fauci A S. The qualitative nature of the primary immune response to HIV infection is a prognosticator of disease progression independent of the initial level of plasma viremia. Proc Natl Acad Sci USA. 1997;94:254–258. doi: 10.1073/pnas.94.1.254. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Deeks S G, Kitchen C M, Liu L, Guo H, Gascon R, Narvaez A B, Hunt P, Martin J N, Kahn J O, Levy J, McGrath M S, Hecht F M. Immune activation set point during early HIV infection predicts subsequent CD4+ T-cell changes independent of viral load. Blood. 2004;104:942–947. doi: 10.1182/blood-2003-09-3333. [DOI] [PubMed] [Google Scholar]
  5. Watford W T, Moriguchi M, Morinobu A, O'Shea J J. The biology of IL-12: coordinating innate and adaptive immune responses. Cytokine Growth Factor Rev. 2003;14:361–368. doi: 10.1016/s1359-6101(03)00043-1. [DOI] [PubMed] [Google Scholar]
  6. Trinchieri G. Interleukin-12: a cytokine at the interface of inflammation and immunity. Adv Immunol. 1998;70:83–243. doi: 10.1016/s0065-2776(08)60387-9. [DOI] [PubMed] [Google Scholar]
  7. Gordin F M, Hartigan P M, Klimas N G, Zolla-Pazner S B, Simberkoff M S, Hamilton J D. Delayed-type hypersensitivity skin tests are an independent predictor of human immunodeficiency virus disease progression. Department of Veterans Affairs Cooperative Study Group. J Infect Dis. 1994;169:893–897. doi: 10.1093/infdis/169.4.893. [DOI] [PubMed] [Google Scholar]
  8. Lane H C, Depper J M, Greene W C, Whalen G, Waldmann T A, Fauci A S. Qualitative analysis of immune function in patients with the acquired immunodeficiency syndrome. Evidence for a selective defect in soluble antigen recognition. N Engl J Med. 1985;313:79–84. doi: 10.1056/NEJM198507113130204. [DOI] [PubMed] [Google Scholar]
  9. Smolen J S, Bettelheim P, Koller U, McDougal S, Graninger W, Luger T A, Knapp W, Lechner K. Deficiency of the autologous mixed lymphocyte reaction in patients with classic hemophilia treated with commercial factor VIII concentrate. Correlation with T cell subset distribution, antibodies to lymphadenopathy-associated or human T lymphotropic virus, and analysis of the cellular basis of the deficiency. J Clin Invest. 1985;75:1828–1834. doi: 10.1172/JCI111896. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Clerici M, Stocks N I, Zajac R A, Boswell R N, Lucey D R, Via C S, Shearer G M. Detection of three distinct patterns of T helper cell dysfunction in asymptomatic, human immunodeficiency virus-seropositive patients. Independence of CD4+ cell numbers and clinical staging. J Clin Invest. 1989;84:1892–1899. doi: 10.1172/JCI114376. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Maggi E, Macchia D, Parronchi P, Mazzetti M, Ravina A, Milo D, Romagnani S. Reduced production of interleukin 2 and interferon-γ and enhanced helper activity for IgG synthesis by cloned CD4+ T cells from patients with AIDS. Eur J Immunol. 1987;17:1685–1690. doi: 10.1002/eji.1830171202. [DOI] [PubMed] [Google Scholar]
  12. Chehimi J, Starr S E, Frank I, Rengaraju M, Jackson S J, Llanes C, Kobayashi M, Perussia B, Young D, Nickbarg E. Natural killer (NK) cell stimulatory factor increases the cytotoxic activity of NK cells from both healthy donors and human immunodeficiency virus-infected patients. J Exp Med. 1992;175:789–796. doi: 10.1084/jem.175.3.789. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Ullum H, Gotzsche P C, Victor J, Dickmeiss E, Skinhoj P, Pedersen B K. Defective natural immunity: an early manifestation of human immunodeficiency virus infection. J Exp Med. 1995;182:789–799. doi: 10.1084/jem.182.3.789. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Chehimi J, Starr S E, Frank I, D'Andrea A, Ma X, MacGregor R R, Sennelier J, Trinchieri G. Impaired interleukin 12 production in human immunodeficiency virus-infected patients. J Exp Med. 1994;179:1361–1366. doi: 10.1084/jem.179.4.1361. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Chougnet C, Wynn T A, Clerici M, Landay A L, Kessler H A, Rusnak J, Melcher G P, Sher A, Shearer G M. Molecular analysis of decreased interleukin-12 production in persons infected with human immunodeficiency virus. J Infect Dis. 1996;174:46–53. doi: 10.1093/infdis/174.1.46. [DOI] [PubMed] [Google Scholar]
  16. Gazzinelli R T, Bala S, Stevens R, Baseler M, Wahl L, Kovacs J, Sher A. HIV infection suppresses type 1 lymphokine and IL-12 responses to Toxoplasma gondii but fails to inhibit the synthesis of other parasite-induced monokines. J Immunol. 1995;155:1565–1574. [PubMed] [Google Scholar]
  17. Meyaard L, Hovenkamp E, Pakker N, van der Pouw Kraan T C, Miedema F. Interleukin-12 (IL-12) production in whole blood cultures from human immunodeficiency virus-infected individuals studied in relation to IL- 10 and prostaglandin E2 production. Blood. 1997;89:570–576. [PubMed] [Google Scholar]
  18. Marshall J D, Chehimi J, Gri G, Kostman J R, Montaner L J, Trinchieri G. The interleukin-12-mediated pathway of immune events is dysfunctional in human immunodeficiency virus-infected individuals. Blood. 1999;94:1003–1011. [PubMed] [Google Scholar]
  19. Clerici M, Lucey D R, Berzofsky J A, Pinto L A, Wynn T A, Blatt S P, Dolan M J, Hendrix C W, Wolf S F, Shearer G M. Restoration of HIV-specific cell-mediated immune responses by interleukin-12 in vitro. Science. 1993;262:1721–1724. doi: 10.1126/science.7903123. [DOI] [PubMed] [Google Scholar]
  20. Paganin C, Frank I, Trinchieri G. Priming for high interferon-γ production induced by interleukin-12 in both CD4+ and CD8+ T cell clones from HIV-infected patients. J Clin Invest. 1995;96:1677–1682. doi: 10.1172/JCI118209. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Landay A L, Clerici M, Hashemi F, Kessler H, Berzofsky J A, Shearer G M. In vitro restoration of T cell immune function in human immunodeficiency virus-positive persons: effects of interleukin (IL)-12 and anti-IL-10. J Infect Dis. 1996;173:1085–1091. doi: 10.1093/infdis/173.5.1085. [DOI] [PubMed] [Google Scholar]
  22. Estaquier J, Idziorek T, Zou W, Emilie D, Farber C M, Bourez J M, Ameisen J C. T helper type 1/T helper type 2 cytokines and T cell death: preventive effect of interleukin 12 on activation-induced and CD95 (FAS/APO-1)-mediated apoptosis of CD4+ T cells from human immunodeficiency virus-infected persons. J Exp Med. 1995;182:1759–1767. doi: 10.1084/jem.182.6.1759. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Hazenberg M D, Hamann D, Schuitemaker H, Miedema F. T cell depletion in HIV-1 infection: how CD4+ T cells go out of stock. Nat Immunol. 2000;1:285–289. doi: 10.1038/79724. [DOI] [PubMed] [Google Scholar]
  24. Lederman M M, Kalish L A, Asmuth D, Fiebig E, Mileno M, Busch M P. “Modeling” relationships among HIV-1 replication, immune activation and CD4+ T-cell losses using adjusted correlative analyses. AIDS. 2000;14:951–958. doi: 10.1097/00002030-200005260-00006. [DOI] [PubMed] [Google Scholar]
  25. Rodriguez B, Sethi A K, Cheruvu V K, Mackay W, Bosch R J, Kitahata M, Boswell S L, Mathews W C, Bangsberg D R, Martin J, Whalen C C, Sieg S, Yadavalli S, Deeks S G, Lederman M M. Predictive value of plasma HIV RNA level on rate of CD4 T-cell decline in untreated HIV infection. JAMA. 2006;296:1498–1506. doi: 10.1001/jama.296.12.1498. [DOI] [PubMed] [Google Scholar]
  26. Hazenberg M D, Otto S A, van Benthem B H, Roos M T, Coutinho R A, Lange J M, Hamann D, Prins M, Miedema F. Persistent immune activation in HIV-1 infection is associated with progression to AIDS. AIDS. 2003;17:1881–1888. doi: 10.1097/00002030-200309050-00006. [DOI] [PubMed] [Google Scholar]
  27. McCune J M. The dynamics of CD4+ T-cell depletion in HIV disease. Nature. 2001;410:974–979. doi: 10.1038/35073648. [DOI] [PubMed] [Google Scholar]
  28. Papagno L, Spina C A, Marchant A, Salio M, Rufer N, Little S, Dong T, Chesney G, Waters A, Easterbrook P, Dunbar P R, Shepherd D, Cerundolo V, Emery V, Griffiths P, Conlon C, McMichael A J, Richman D D, Rowland-Jones S L, Appay V. Immune activation and CD8+ T-cell differentiation towards senescence in HIV-1 infection. PLoS Biol. 2004;2:E20. doi: 10.1371/journal.pbio.0020020. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Holm G H, Gabuzda D. Distinct mechanisms of CD4+ and CD8+ T-cell activation and bystander apoptosis induced by human immunodeficiency virus type 1 virions. J Virol. 2005;79:6299–6311. doi: 10.1128/JVI.79.10.6299-6311.2005. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Bofill M, Mocroft A, Lipman M, Medina E, Borthwick N J, Sabin C A, Timms A, Winter M, Baptista L, Johnson M A, Lee C A, Phillips A N, Janossy G. Increased numbers of primed activated CD8+CD38+CD45RO+ T cells predict the decline of CD4+ T cells in HIV-1-infected patients. AIDS. 1996;10:827–834. doi: 10.1097/00002030-199607000-00005. [DOI] [PubMed] [Google Scholar]
  31. Liu Z, Cumberland W G, Hultin L E, Prince H E, Detels R, Giorgi J V. Elevated CD38 antigen expression on CD8+ T cells is a stronger marker for the risk of chronic HIV disease progression to AIDS and death in the Multicenter AIDS Cohort Study than CD4+ cell count, soluble immune activation markers, or combinations of HLA-DR and CD38 expression. J Acquir Immune Defic Syndr Hum Retrovirol. 1997;16:83–92. doi: 10.1097/00042560-199710010-00003. [DOI] [PubMed] [Google Scholar]
  32. Benito J M, Lopez M, Lozano S, Ballesteros C, Martinez P, Gonzalez-Lahoz J, Soriano V. Differential upregulation of CD38 on different T-cell subsets may influence the ability to reconstitute CD4+ T cells under successful highly active antiretroviral therapy. J Acquir Immune Defic Syndr. 2005;38:373–381. doi: 10.1097/01.qai.0000153105.42455.c2. [DOI] [PubMed] [Google Scholar]
  33. Benito J M, Lopez M, Lozano S, Martinez P, Gonzalez-Lahoz J, Soriano V. CD38 expression on CD8 T lymphocytes as a marker of residual virus replication in chronically HIV-infected patients receiving antiretroviral therapy. AIDS Res Hum Retroviruses. 2004;20:227–233. doi: 10.1089/088922204773004950. [DOI] [PubMed] [Google Scholar]
  34. Rosenblatt H M, Stanley K E, Song L Y, Johnson G M, Wiznia A A, Nachman S A, Krogstad P A. Immunological response to highly active antiretroviral therapy in children with clinically stable HIV-1 infection. J Infect Dis. 2005;192:445–455. doi: 10.1086/431597. [DOI] [PubMed] [Google Scholar]
  35. Blankson J N, Finzi D, Pierson T C, Sabundayo B P, Chadwick K, Margolick J B, Quinn T C, Siciliano R F. Biphasic decay of latently infected CD4+ T cells in acute human immunodeficiency virus type 1 infection. J Infect Dis. 2000;182:1636–1642. doi: 10.1086/317615. [DOI] [PubMed] [Google Scholar]
  36. Janssen R S, Satten G A, Stramer S L, Rawal B D, O'Brien T R, Weiblen B J, Hecht F M, Jack N, Cleghorn F R, Kahn J O, Chesney M A, Busch M P. New testing strategy to detect early HIV-1 infection for use in incidence estimates and for clinical and prevention purposes. JAMA. 1998;280:42–48. doi: 10.1001/jama.280.1.42. [DOI] [PubMed] [Google Scholar]
  37. Wirleitner B, Schroecksnadel K, Winkler C, Fuchs D. Neopterin in HIV-1 infection. Mol Immunol. 2005;42:183–194. doi: 10.1016/j.molimm.2004.06.017. [DOI] [PubMed] [Google Scholar]
  38. Wirleitner B, Reider D, Ebner S, Bock G, Widner B, Jaeger M, Schennach H, Romani N, Fuchs D. Monocyte-derived dendritic cells release neopterin. J Leukoc Biol. 2002;72:1148–1153. [PubMed] [Google Scholar]
  39. Meyer K C, Cornwell R, Carlin J M, Powers C, Irizarry A, Byrne G I, Borden E C. Effects of interferons β or γ on neopterin biosynthesis and tryptophan degradation by human alveolar macrophages in vitro: synergy with lipopolysaccharide. Am J Respir Cell Mol Biol. 1992;6:639–646. doi: 10.1165/ajrcmb/6.6.639. [DOI] [PubMed] [Google Scholar]
  40. Rosenberg E S, Altfeld M, Poon S H, Phillips M N, Wilkes B M, Eldridge R L, Robbins G K, D'Aquila R T, Goulder P J, Walker B D. Immune control of HIV-1 after early treatment of acute infection. Nature. 2000;407:523–526. doi: 10.1038/35035103. [DOI] [PubMed] [Google Scholar]
  41. Norris P J, Pappalardo B L, Custer B, Spotts G, Hecht F M, Busch M P. Elevations in IL-10, TNF-α, and IFN-γ from the earliest point of HIV type 1 infection. AIDS Res Hum Retroviruses. 2006;22:757–762. doi: 10.1089/aid.2006.22.757. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. Bebell L M, Passmore J A, Williamson C, Koleka M, Iriogbe I, van Loggerenberg F, Karim Q A, Karim S A. Relationship between levels of inflammatory cytokines in the genital tract and CD4+ cell counts in women with acute HIV-1 infection. J Infect Dis. 2008;198:710–714. doi: 10.1086/590503. [DOI] [PubMed] [Google Scholar]
  43. Nilsson J, Kinloch-de-Loes S, Granath A, Sonnerborg A, Goh L E, Andersson J. Early immune activation in gut-associated and peripheral lymphoid tissue during acute HIV infection. AIDS. 2007;21:565–574. doi: 10.1097/QAD.0b013e3280117204. [DOI] [PubMed] [Google Scholar]
  44. Chougnet C, Thomas E, Landay A L, Kessler H A, Buchbinder S, Scheer S, Shearer G M. CD40 ligand and IFN-γ synergistically restore IL-12 production in HIV-infected patients. Eur J Immunol. 1998;28:646–656. doi: 10.1002/(SICI)1521-4141(199802)28:02<646::AID-IMMU646>3.0.CO;2-6. [DOI] [PubMed] [Google Scholar]
  45. Atabani S F, Byrnes A A, Jaye A, Kidd I M, Magnusen A F, Whittle H, Karp C L. Natural measles causes prolonged suppression of interleukin-12 production. J Infect Dis. 2001;184:1–9. doi: 10.1086/321009. [DOI] [PubMed] [Google Scholar]
  46. Kamga I, Kahi S, Develioglu L, Lichtner M, Maranon C, Deveau C, Meyer L, Goujard C, Lebon P, Sinet M, Hosmalin A. Type I interferon production is profoundly and transiently impaired in primary HIV-1 infection. J Infect Dis. 2005;192:303–310. doi: 10.1086/430931. [DOI] [PubMed] [Google Scholar]
  47. Cordiali Fei P, Massa A, Prignano G, Pietravalle M, Alemanno L, Vitelli G, Palamara G, Giglio A, Gandolfo G M, Gentili G. Behavior of several “progression markers” during the HIV-Ab seroconversion period. Comparison with later stages. J Biol Regul Homeost Agents. 1992;6:57–64. [PubMed] [Google Scholar]
  48. Von Sydow M, Sonnerborg A, Gaines H, Strannegard O. Interferon-α and tumor necrosis factor-α in serum of patients in various stages of HIV-1 infection. AIDS Res Hum Retroviruses. 1991;7:375–380. doi: 10.1089/aid.1991.7.375. [DOI] [PubMed] [Google Scholar]
  49. Gautier G, Humbert M, Deauvieau F, Scuiller M, Hiscott J, Bates E E, Trinchieri G, Caux C, Garrone P. A type I interferon autocrine-paracrine loop is involved in Toll-like receptor-induced interleukin-12p70 secretion by dendritic cells. J Exp Med. 2005;201:1435–1446. doi: 10.1084/jem.20041964. [DOI] [PMC free article] [PubMed] [Google Scholar]
  50. Byrnes A A, Ma X, Cuomo P, Park K, Wahl L, Wolf S F, Zhou Z, Trinchieri G, Karp C L. Type I interferons and IL-12: convergence and cross-regulation among mediators of cellular immunity. Eur J Immunol. 2001;31:2026–2034. doi: 10.1002/1521-4141(200107)31:7&#60;2026::aid-immu2026&#62;3.0.co;2-u. [DOI] [PubMed] [Google Scholar]
  51. Byrnes A A, McArthur J C, Karp C L. Interferon-β therapy for multiple sclerosis induces reciprocal changes in interleukin-12 and interleukin-10 production. Ann Neurol. 2002;51:165–174. doi: 10.1002/ana.10084. [DOI] [PubMed] [Google Scholar]
  52. Biglino A, Sinicco A, Forno B, Pollono A M, Sciandra M, Martini C, Pich P, Gioannini P. Serum cytokine profiles in acute primary HIV-1 infection and in infectious mononucleosis. Clin Immunol Immunopathol. 1996;78:61–69. doi: 10.1006/clin.1996.0009. [DOI] [PubMed] [Google Scholar]
  53. Nubling C M, Chudy M, Volkers P, Lower J. Neopterin levels during the early phase of human immunodeficiency virus, hepatitis C virus, or hepatitis B virus infection. Transfusion. 2006;46:1886–1891. doi: 10.1111/j.1537-2995.2006.00994.x. [DOI] [PubMed] [Google Scholar]
  54. Sinicco A, Biglino A, Sciandra M, Forno B, Pollono A M, Raiteri R, Gioannini P. Cytokine network and acute primary HIV-1 infection. AIDS. 1993;7:1167–1172. doi: 10.1097/00002030-199309000-00003. [DOI] [PubMed] [Google Scholar]
  55. Barcellini W, Rizzardi G P, Poli G, Tambussi G, Velati C, Meroni P L, Dalgleish A G, Lazzarin A. Cytokines and soluble receptor changes in the transition from primary to early chronic HIV type 1 infection. AIDS Res Hum Retroviruses. 1996;12:325–331. doi: 10.1089/aid.1996.12.325. [DOI] [PubMed] [Google Scholar]
  56. Gaines H, von Sydow M A, von Stedingk L V, Biberfeld G, Bottiger B, Hansson L O, Lundbergh P, Sonnerborg A B, Wasserman J, Strannegaard O O. Immunological changes in primary HIV-1 infection. AIDS. 1990;4:995–999. doi: 10.1097/00002030-199010000-00008. [DOI] [PubMed] [Google Scholar]
  57. Griffin D E, Ward B J. Differential CD4 T cell activation in measles. J Infect Dis. 1993;168:275–281. doi: 10.1093/infdis/168.2.275. [DOI] [PubMed] [Google Scholar]
  58. Ryon J J, Moss W J, Monze M, Griffin D E. Functional and phenotypic changes in circulating lymphocytes from hospitalized Zambian children with measles. Clin Diagn Lab Immunol. 2002;9:994–1003. doi: 10.1128/CDLI.9.5.994-1003.2002. [DOI] [PMC free article] [PubMed] [Google Scholar]
  59. Yoneyama A, Nakahara K, Higashihara M, Kurokawa K. Increased levels of soluble CD8 and CD4 in patients with infectious mononucleosis. Br J Haematol. 1995;89:47–54. doi: 10.1111/j.1365-2141.1995.tb08912.x. [DOI] [PubMed] [Google Scholar]
  60. Brenchley J M, Price D A, Schacker T W, Asher T E, Silvestri G, Rao S, Kazzaz Z, Bornstein E, Lambotte O, Altmann D, Blazar B R, Rodriguez B, Teixeira-Johnson L, Landay A, Martin J N, Hecht F M, Picker L J, Lederman M M, Deeks S G, Douek D C. Microbial translocation is a cause of systemic immune activation in chronic HIV infection. Nat Med. 2006;12:1365–1371. doi: 10.1038/nm1511. [DOI] [PubMed] [Google Scholar]

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