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Blood, Vol. 95 No. 1 (January 1), 2000:
pp. 249-255
IMMUNOBIOLOGY
T-cell division in human immunodeficiency virus (HIV)-1
infection is mainly due to immune activation: a longitudinal
analysis in patients before and during highly active antiretroviral
therapy (HAART)
Mette D. Hazenberg,
James W. T. Cohen Stuart,
Sigrid A. Otto,
Jan C. C. Borleffs,
Charles A. B. Boucher,
Rob J. de Boer,
Frank Miedema, and
Dörte Hamann
From the Department of Clinical Viro-Immunology, CLB, and the
Laboratory for Experimental and Clinical Immunology and Department of
Human Retrovirology, Academic Medical Center, University of
Amsterdam, Amsterdam; the Department of Internal Medicine, University
Hospital Utrecht; the Department of Virology, Eijkman-Winkler
Institute, University Hospital Utrecht; and the Department of
Theoretical Biology, Utrecht University, Utrecht, The Netherlands.
 |
Abstract |
In human immunodeficiency virus (HIV)-1 infection, highly
increased T-cell turnover was proposed to cause exhaustion of
lymphocyte production and consequently development of AIDS. Here, we
investigated cell proliferation, as measured by expression of the Ki-67
nuclear antigen, in peripheral blood CD4+ and
CD8+ lymphocyte subpopulations before and during highly
active antiretroviral therapy (HAART). In untreated HIV-1
infection, both the percentage and number of Ki-67+
CD4+ and CD8+ lymphocytes were
significantly increased, compared with values obtained from
healthy individuals. A more than 10-fold increase in the percentage of
dividing naive CD4+ T cells in the blood was found
when the number of these cells were below 100 per µL.. HAART
induced an immediate decline in Ki-67 antigen expression, despite often
very low CD4+ T-cell numbers, arguing against increased
proliferation being a homeostatic response. After approximately 24 weeks of HAART treatment, a transient increase in the number of
proliferating cells was seen, but only in the CD4+
CD27+ memory pool. In the CD8+ T-cell
compartment, the number of dividing cells was elevated 20- to
25-fold. This increase was most notable in the CD27+ CD
45RO+ and CD27
CD45RO+ memory CD8+ T-cell pool,
corresponding with the degree of expansion of these subsets. Reduction
of plasma HIV-RNA load by HAART was accompanied by a decrease in
numbers and percentages of dividing cells in all CD8+
T-cell subsets. Taken together, our results indicate that peripheral T-cell proliferation is a consequence of generalized immune activation. (Blood. 2000;95:249-255)
© 2000 by The American Society of Hematology.
| |
Introduction |
Several hypotheses have been presented to
explain the loss of CD4+ T lymphocytes, the major
hallmark of human immunodeficiency virus (HIV)-1 infection that leads
to severe immune depletion and ultimately AIDS and death. The model of
high lymphocyte turnover has received considerable attention over
the past few years.1,2 On the basis of the observation that
during the first few weeks of highly active antiretroviral therapy
(HAART) the number of CD4+ T cells increases, it was
postulated that HIV-1 infection leads to a rapid turnover of
lymphocytes, reflecting a new balance between production and
death of lymphocytes. The increased production rate would eventually
lead to exhaustion of the CD4+ T-cell renewal capacity and
result in CD4+ lymphocyte depletion. Indeed, in
simian immunodeficiency virus (SIV)-infected macaques increased T-cell
production was observed3; however, there was a parallel
rise in the turnover of B cells and NK cells.4 Moreover,
the highest increase was observed in CD8+ T cells,
the compartment that is initially expanded and does not get depleted
until very late in the course of HIV-1 infection.5-9
HIV-induced lymphocyte depletion and subsequent recovery after HAART
appeared to be variable when studied at the level of T-cell subsets.
Infection with HIV-1 induces an early decline in the number of naive
CD4+ and CD8+ and memory CD4+ T
lymphocytes. Conversely, the memory and activated CD8+
T-cell compartments expand initially. Only shortly preceding progression to AIDS, the numbers of these latter cell types fall as well.10-12 HAART induces an early rise of memory
CD4+ and CD8+ T-cell numbers, accompanied by a
slow increase in naive CD4+ and CD8+
T-cell numbers.13,14 From this, it was concluded that
different mechanisms are involved in the dynamics of depletion and
recovery of the various T-lymphocyte subsets. These mechanisms include interference of HIV with de novo T-cell production
capacity15 (D. R. Clark et al, submitted for publication)
and enhanced sequestration of cells in lymphoid
tissues.14,16-18 Recently several groups have reported on
cross-sectional studies of T-cell production levels. Most of these
reports studied total CD4+ and CD8+ T-cell
populations, and did not discriminate between naive and memory
subsets.3,7,8,19,20
To answer the question whether increased T-cell proliferation could be
involved in CD4+ lymphocyte depletion, we measured
expression of the Ki-67 antigen in naive and memory CD4+
T-cell subsets in the blood. Furthermore, we investigated the role of
T-cell proliferation in the expansion and depletion of naive, memory,
and effector CD8+ T lymphocyte subsets. Our approach was
longitudinal: cell division rates were studied from the stage of
chronic untreated HIV-1 infection up to a year of treatment with HAART.
| |
Materials and methods |
Patients
To study T-cell turnover before and during antiretroviral treatment,
sequential cryopreserved peripheral blood samples from HIV-1 infected
participants of the CHEESE study were used. Cryopreservation was
performed using a computerized freezing device that results in optimal
quality of viably frozen cells for functional studies.21 Frozen blood samples were stored in liquid nitrogen. No differences in
Ki-67 expression were found between freshly isolated or frozen cells in
a pilot experiment (data not shown). In the multicenter CHEESE study,
patients received either zidovudine (Retrovir; 200 mg, 3 times daily),
lamivudine (Epivir; 150 mg, twice daily), and saquinavir soft gel
capsules (Fortovase; 1200 mg, 3 times daily), or zidovudine,
lamivudine, and indinavir (Crixivan; 800 mg, 3 times daily). The
inclusion criteria for this trial were no previous treatment with
antiretroviral therapy, except for AZT < 12 months, CD4+
T-cell count < 500 per µL, or HIV RNA > 10 000 copies per mL, or
CDC stage B or C.22 Of 60 participants, 16 were selected on
the basis of baseline CD4+ T-cell numbers and the
availability of frozen peripheral blood mononuclear cell (PBMC)
samples. Patients were equally distributed among both therapy arms.
Expression of the Ki-67 antigen was analyzed at 5 timepoints: during
untreated HIV infection (t = 0) and after 4 (t = 4), 12 (t = 12),
24 (t = 24), and 48 (t = 48) weeks of therapy. As controls,
cryopreserved PBMC from 5 HIV-negative blood bank donors were used.
Monoclonal antibodies
CD4-PerCP, CD8-PerCP, CD45RO-PE mAb, and Streptavidin-APC were
obtained from Becton Dickinson (San Jose, CA). Biotinylated CD27 mAb
was purchased from the CLB (Amsterdam, The Netherlands), and
FITC-labeled Ki-67 mAb was purchased from Immunotech (Marseille, France).
Flow cytometry
CD4+ and CD8+ T cells were subdivided into
naive (CD45RO/ CD27+), CD27+
memory (CD45RO+/CD27+), CD27
memory (CD45RO+/CD27), and
CD27 effector
(CD45RO/CD27) cells, as
previously described.23,24 In contrast to these earlier
studies, CD45RA mAb was replaced by CD45RO mAb and the definition of
the T-cell subsets was adjusted to the used mAb combination. Cell
proliferation was studied by measuring expression of the Ki-67 antigen,
which is expressed by cells in late G1, S, G2, and M phase of the cell
division circle.25-28
Cryopreserved peripheral blood mononuclear cells were thawed and
incubated with CD4 or CD8PerCP mAb, CD45ROPE, and
biotinylated CD27 mAb. After washing with phosphate-buffered saline
(PBS)/0.5% bovine serum albumin (BSA), cells were incubated with
Streptavidin-APC. Red blood cells were lysed and lymphocytes fixated
with fluorescence-activated cell sorter (FACS) Lysing Solution (Becton
Dickinson). Subsequent permeabilization was performed by incubating
cells with FACS permeabilization buffer (Becton Dickinson), after which
cells were stained intracellularly with Ki-67-FITC mAb. Cells were
fixed, using Cellfix (Becton Dickinson), and analyzed on a FACS Calibur
(Becton Dickinson) with Cellquest software. All incubation steps were
performed at 4°C for 20 minutes; for fixation and permeabilization,
samples were kept at room temperature for 10 minutes.
Viral load
Plasma viral load was determined with RT-PCR detecting HIV-1
RNA (Roche Amplicor Monitor Standard Assay, Roche Diagnostics, Branchburg, NJ).
Statistical analysis
Patient characteristics at baseline and after various timepoints
during treatment with HAART were compared using Wilcoxon Signed Ranks
test and Mann-Whitney U test for comparison with healthy control
values. Correlations were calculated using Spearman's correlation coefficients.
| |
Results |
Ki-67 antigen expression in chronic HIV-1 infection
During chronic untreated HIV-1 infection, both the percentage and
the absolute number of Ki-67+ CD4+ and
CD8+ T cells was significantly increased (Figure
1, Tables
1 and 2; P = .001), compared with
healthy controls. The percentage of Ki-67 expression was significantly
elevated in all subsets (P < .005), except for the
CD8+ CD27 effector T-cell population
(Figure 1). Naive T cells showed a 10- (CD4) to 20- (CD8) fold
elevation in Ki-67+ T-cell percentage, and in the memory
and effector T-lymphocyte compartment, Ki-67 expression was increased
up to 7-fold (Figure 1). When absolute numbers instead of
percentages of dividing cells were analyzed, cell proliferation in the
CD4+ T-cell subsets was elevated < 3-fold, whereas in the
CD8+ compartment, a 20- to more than 50-fold rise was
observed (Tables 1 and 2). This increase was most notable in the
CD27+ memory and CD27 memory lymphocyte
population (see t = 0, Figure 4B).
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| Fig 1.
Percentage of Ki-67+ T lymphocytes within
the CD4+ (A) and CD8+ (B) peripheral blood
T-cell populations of healthy individuals (gray bars) and untreated
HIV-infected patients (black bars).
An asterisk represents statistical difference compared with control
value (P < .005; Mann-Whitney U test).
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Table 1.
Number of Ki-67 antigen expressing cells within the
total CD4+ T-cell population and T-cell subsets of
untreated HIV-1 infected patients and healthy
individuals
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Table 2.
Number of Ki-67 antigen expressing cells within the
total CD8+ T-cell population and T-cell subsets of
untreated HIV-1 infected patients and healthy
individuals
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The percentage of naive Ki-67+ CD4+ T
lymphocytes was inversely correlated with the number of naive cells
(Figure 2), but did not correlate with
plasma viral load (data not shown). Strikingly, only when the number of
naive CD4+ T cells dropped below 100 cells per µL, the
proportion of Ki-67 expressing cells in this subset exceeded 2% (naive
CD4 count > 100/µL: r = 0.79, P = .004; naive
CD4 count < 100/µL: r = 0.73, P = .025;
stratification arbitrary). The same overall trend, but not a
significant correlation, was observed for the naive CD8 T-cell
compartment (Figure 2). The size of the CD27+ memory
CD4+ T-cell pool was negatively correlated with the
percentage of proliferating cells in this compartment (Figure 2;
r = 0.68, P = .001). In the CD8+ T-cell
compartment, the opposite was observed. Here, an increase in the
percentage of Ki-67 expressing cells correlated positively with the
size of the CD27+ memory T-lymphocyte subset (Figure 2;
r = 0.50, P = .024) and the size of the
CD27 memory T-cell subset (data not shown;
r = 0.52, P = .016). No correlation was found between the
same parameters in the CD4+ CD27 memory
or the CD8+ CD27 effector T-cell
population, or between Ki-67 antigen expression and viral load (data
not shown).
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| Fig 2.
Correlation between the number of CD4+ and
CD8+ naive or CD27+ memory T cells and the
percentage of Ki-67+ naive or CD27+ memory
cells of untreated HIV-infected and noninfected individuals
(Spearman's correlation coefficients).
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Longitudinal analysis of immune recovery during HAART
Infection with HIV-1 resulted in CD4+ T-cell depletion
and expansion of the CD8+ T-cell pool as has been described
previously (t = 0 in Figure 4A and B). Treatment with antiretroviral
therapy resulted in a significant decrease of plasma HIV-RNA to levels
below the limits of detection (lower limit of detection 400 RNA
copies/mL). In parallel, total numbers of CD4+ T cells
increased, and a fall in total numbers of CD8+ T
lymphocytes was observed (Figure 3). Figure
4A shows that the therapy-induced rise in
CD4+ T cells observed in the first 4 weeks of HAART was
accounted for by an increase of both CD27+ memory and naive
T lymphocytes. Stratification of patients into 2 groups, based on their
baseline naive CD4+ T-cell counts, revealed that the
HAART-induced restoration of the naive subset was better in patients
with higher baseline naive CD4+ T-cell values
(Figure 5). After 1 year of treatment,
numbers of naive and CD27+ memory CD4+ T cells
had risen significantly compared with pretreatment levels, but were
still lower compared with control levels (Figure 4A, P < .05). In the same period, as depicted in Figure 4B, the
number of naive CD8+ T cells increased. The
CD27+ and CD27 memory and
CD27 effector CD8+ T-lymphocyte subsets
remained significantly expanded, compared with control values
(P < .05), despite an initial decrease in cell number that
was significant for CD27+ and CD27
memory T cells after 1 and 3 months, respectively
(P < .05).
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| Fig 3.
Longitudinal analysis of lymphocyte recovery and plasma
HIV-RNA load decline.
White dots represent HIV-RNA load, black dots CD4+ T cells
and gray dots CD8+ T cells.
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| Fig 4.
CD4+ and CD8+ T-cell subset
recovery.
Naive (circles), CD27+ memory (triangles),
CD27 memory (squares), and CD27
effector (diamonds) lymphocytes within the CD4+ (A) and
CD8+ (B) T-cell compartments are shown. Sequential patient
values were compared using the Wilcoxon Signed Ranks test, and the
Mann-Whitney U test was used for comparison with healthy individuals (*
P < .05).
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| Fig 5.
Early recovery of CD4+ naive and
CD27+ memory T lymphocytes.
Patients were stratified according to baseline naive T-cell values.
Black and white dots represent naive and CD27+ memory
cells, respectively.
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Ki-67 antigen expression during HAART-induced immune recovery
Immediately on reduction of plasma HIV-RNA load by HAART, the total
number (Figure 6A and B) and the percentage
(data not shown) of CD4+ and CD8+
Ki-67+ T cells and of Ki-67+ T lymphocytes in
all subsets declined. In the CD4+ T-cell compartment, this
decline was relatively slow when compared with the rapid decline
observed in the CD8+ T-cell population. Furthermore, after
6 months of HAART, a temporary increase in the number of
Ki-67+ lymphocytes was observed in the total
CD4+ T-cell population. After 1 year of treatment, numbers
of dividing CD4+ T cells had returned to normal.
Interestingly, the percentage of proliferating CD4+ naive
and CD27+ memory T lymphocytes was still significantly
elevated at that time point (P < .05; data not shown). In
the CD8+ T-cell compartment, the steepest decline in the
number of Ki-67+ T cells was observed in the first 4 weeks
of treatment (figure 6B; P < .05 for t = 0 vs t = 4 in
total CD8+ T cells and in naive, CD27+ and
CD27-memory T-cell subsets). After 1 year, the percentage (data not
shown), but not the absolute number of Ki-67+
CD27+ and CD27 memory and
Ki-67+ CD27 effector T lymphocytes had
returned to normal values (P < .008).
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| Fig 6.
Longitudinal analysis of Ki-67 antigen expression in
CD4+ (A) and CD8+ (B) T lymphocyte subsets
and total populations.
Decline of the number of total (gray circles), naive (black circles),
CD27+ memory (black triangles), CD27
memory (black squares), and CD27 effector (black
diamonds) Ki-67+ T cells is shown. Sequential patient
values were compared using the Wilcoxon Signed Ranks test, and the
Mann-Whitney U test was used for comparison with healthy individuals (*
P < .05).
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Despite the severe depletion of the naive T-cell pool, the number of
Ki-67+ naive T lymphocytes fell immediately on introduction
of HAART (Figure 6A and B). Figure 7
depicts data of the individual Ki-67 decline in naive CD4+
T lymphocytes. Reduction of Ki-67 expression was relatively slow in 2 patients who had less than 100 naive CD4+ T cells per µL
and the highest percentage of dividing naive CD4+ T cells
at baseline. However, as is shown in Figure
8A, no correlation was found between early
naive, CD27+ memory or CD27 memory (data
not shown) CD4+ T-cell recovery with the baseline size of
these subsets or with the number of dividing lymphocytes. In the
CD8+ T-cell compartment (Figure 8B), recovery of the naive
subset correlated negatively with its baseline size and with the number of dividing cells within this pool (r = 0.511,
P = .043 and r = 0.743, P = .001
respectively). Finally, patients with the most pronounced reduction in
size of the CD27+ or CD27 memory T-cell
subset had highest numbers of CD27+ or
CD27 memory CD8+ T lymphocytes
(r = 0.568, P = .022 and r = 0.591,
P = .016), and highest numbers of dividing cells
(r = 0.776, P = 0.000 and r = 0.618,
P = .011) at baseline. A similar trend, albeit not significant, was observed for the CD27 effector
T-cell subset (data not shown).
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| Fig 7.
HAART-induced decline in the percentage of proliferating
Ki-67+ naive CD4+ T cells.
The courses of individual patients are shown. Black dots represent
patients with baseline naive CD4+ T-cell numbers < 300/µL (n = 9), gray dots represent individuals with baseline naive
CD4+ T-cell counts > 300/µL but < 400/µL (n = 5)
and white dots represent patients with baseline naive CD4+
T-cell count > 600/µL (n = 2).
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| Fig 8.
Early recovery of the naive and CD27+
memory CD4+ (A) and naive, CD27+ memory and
CD27 memory CD8+ (B) T-cell pool.
Correlations are shown between T-cell subset recovery in the first 4 weeks of HAART (delta t = 0 vs t = 4) and baseline size of subset
or number of Ki-67+ cells within each subset (Spearman's
correlation coefficients).
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| |
Discussion |
In this longitudinal study, we measured expression of the Ki-67
antigen, which is taken as a measurement for proliferating cells, in
peripheral blood lymphocyte subsets of HIV-1 infected patients. In
untreated HIV-1 infection, both the percentage and number of
Ki-67+ CD4+ and CD8+ lymphocytes
were significantly increased, compared with values obtained from
healthy individuals. These data confirm previous (mainly
cross-sectional) studies,5,7,8,29 showing that T-cell
turnover is increased maximally 2- to 3- fold in the CD4+,
and 6- to 7- fold in the CD8+ T-cell population. However,
when investigating the role of T-cell proliferation in the pathogenesis
of HIV-1 infection, lymphocyte subsets rather than total
CD4+ or CD8+ T-cell pools should be studied, as
naive, memory, and effector T lymphocytes follow different courses of
depletion or expansion. We therefore analyzed changes in expression of
the Ki-67 antigen within naive, CD27+ and
CD27 memory and CD27 effector
CD4+ and CD8+ lymphocyte populations before and
during the course of HAART.
The finding that in untreated HIV-1 infection, the number of
proliferating cells was elevated 2 to 3 times in all CD4+
T-cell subsets is in good agreement with earlier results demonstrating that the number of apoptotic cells within the CD4 population was equally distributed over all subsets.30 Together, these
data show that, in general, HIV-1 infection affects both naive and memory peripheral blood CD4+ T cells to a similar degree.
The early decrease in the number of naive CD4+ T cells
could therefore reflect interference of the virus with T-cell renewal
capacity rather than with peripheral production of these cells.
The median percentage of dividing naive CD4+ T cells was
elevated 10-fold. Higher increases in percentages of Ki-67+
CD4+ naive T cells were observed only when the number of
naive cells fell below 100 cells per µL. When depletion of
lymphocytes would be the driving force for the increase in peripheral
naive CD4+ T-cell production, one would expect the high
rate of cell division to be maintained during treatment with HAART
until the number of cells would be normalized. We observed, however, an
immediate decline in the number and percentage of Ki-67+
naive CD4+ lymphocytes, although CD4+ T cells
were still severely depleted. From this we conclude that increased
Ki-67 expression in the naive CD4+ T-cell subset was caused
by generalized immune activation rather then being a T-cell homeostatic
response to compensate for T-cell depletion.
Furthermore, our data show that the early therapy-induced increase in
CD4+ T cells is accounted for by a rise in the number of
both naive and CD27+ memory T cells. Our finding that, in
both subsets, the percentage and number of dividing lymphocytes
decreased during antiretroviral treatment points toward other
mechanisms than peripheral expansion involved in recovery of the
CD4+ T-cell pool. It confirms previous reports indicating
that in the first 4 weeks of HAART, redistribution of trapped
CD27+ cells significantly adds to therapy-induced increase
in peripheral blood CD4+ T-cell
numbers.14,16,17
Whereas early restoration of the CD27+ memory
CD4+ T-lymphocyte compartment was fairly constant between
patients, recovery of the naive CD4+ T-cell pool was
variable. In patients with less severe depletion of naive
CD4+ T cells, a relatively steep increase in the number of
naive lymphocytes was observed during the first 4 weeks of HAART,
confirming recently published data by Hengel et al.31 After
this first steep rise, replenishment occurred at a slower rate. Several
factors may contribute to restoration of the naive lymphocyte
compartment, such as reappearance of preexisting naive T cells in the
peripheral blood, as well as de novo T-cell production from a thymic
source. The relative importance of each mechanism in recovery of the
immune system needs to be established. Recently, it was reported that
thymic production of T cells increased substantially during HAART. Its dependence on disease progression remains to be studied.15
In contrast to the changes occurring in the CD4+ T-cell
compartment, in the naive CD8+ T-cell pool, percentages of
dividing cells were elevated regardless of the size of this subset.
This points toward a distinct regulation of peripheral T-cell
production rates in CD4+ and CD8+ cells. The
high proliferation rate in the latter population most likely reflects
antigen-induced expansion. The degree of expansion of the
CD8+ T-cell subset correlated with the degree of cell
proliferation during chronic HIV-1 infection observed in that
particular subset. Ongoing antigenic pressure during untreated HIV-1
infection could lead to continuous recruitment of naive
CD8+ T cells, thereby maintaining increased percentages of
Ki-67 antigen expression in this subset. During HAART, both the
percentage and the total numbers of dividing cells declined, suggesting
that activation-related cell division was the driving force for
expansion of the primed subset during chronic HIV-1 infection. The
early therapy-induced drop in the number of primed CD8+ T
cells reached a plateau after 6 months, at which the CD27+
or CD27CD45RO+ memory and
CD27CD45RO effector
CD8+ lymphocyte pool remained significantly expanded.
Although the absolute number of dividing primed CD8+ T
cells remained elevated, the percentage of Ki-67+ T cells
within this subset returned to normal values. Using tetrameric HLA-peptide complexes, it has been shown that HAART reduced the frequency of HIV-1 specific cytotoxic T lymphocytes (CTLs)
several fold.32 The early decline in primed
CD8+ T cells that we observed could therefore reflect
apoptosis of HIV-specific CTLs.
The numbers of dividing cells in healthy individuals found in this
study correspond well to the range of earlier results obtained with
Ki-67 staining7,8 and with deuterated glucose labeling of
dividing cells.19 However, when studying HIV-1 infected
patients, in these reports, the average total peripheral blood
production of proliferating CD4+ T cells was
5.2 × 107 (range 2.7-8.3 × 107)
and 10.8 × 107 (range
7.1-12.9 × 107) of CD8+ T cells. These
values are lower than those reported in our study (11.4 × 107 [range
2.4-21.0 × 107] and 49.2 × 107
[range 7.0-648.5 × 107], respectively), most
likely because of the more advanced disease stage of our patients (see
Table 3).
After the initial HAART-induced decline, we observed a transient rise
in the number of proliferating CD4+ T cells after 6 months
of therapy, after which these numbers declined further. Because this
rise was only due to increased numbers of proliferating
CD27+ memory cells, and coincided with dissolvement of the
depletion of this subset, it could reflect improvement of the
functional capacity of these cells. Indeed, T-cell function, as
measured by the in vitro proliferative response to CD3 and CD28 mAb,
improved and reached a plateau phase after 6 months of HAART (data not shown). Whereas in our study, the rise in T-cell proliferation did not
exceed pretreatment values, in a study reported by Fleury et
al,7 a clear increase in cell production was seen over
baseline after 6 months of HAART. In that report, the baseline
CD4+ T-cell counts were relatively high, on average over
600 cells per µL. We found the transient rise in numbers of dividing
cells to be more pronounced in patients with higher baseline
CD4+ T-cell counts (data not shown), pointing toward
variances in the stage of HIV-1 infection as the cause of the observed differences.
Taken together, our data are compatible with the idea that most of the
elevated cell division and cell death in HIV-1 infection is related to
strong persistent immune activation30,33 rather than a
response to or the cause of lymphocyte depletion. Therefore, alternative factors leading to T-cell decline must be involved in HIV-1
pathogenesis. As has been proposed previously, CD4+ T-cell
depletion may be due to a diminished capacity for T-cell renewal as a
direct or indirect result of HIV-1 infection or could be related to an
intrinsically low capacity for renewal in adults, which cannot even
compensate for a slightly enhanced T-cell death due to HIV-1 infection
and generalized immune activation.34-37 Furthermore, we
observed a clear distinction between CD4+ and
CD8+ T cells, with respect to proliferation rates. A
similar dichotomy was found before, when much more cell activation
induced death and shortened telomeres were found in the
CD8+ T-cell fraction.5,38 This may fit well
with differential requirements for clonal expansion to exercise
CD4+ helper and CD8+ effector cell functions.
Because cell division diminished rapidly with HAART even when T-cell
numbers were still very low, our results suggest that the
increased peripheral lymphocyte proliferation is not a homeostatic
response to T-cell depletion.
| |
Acknowledgments |
We thank patients and physicians who participated in the
CHEESE study, and Dr R. A. W. van Lier for critical reading of the manuscript.
| |
Footnotes |
Submitted July 26, 1999; accepted August 31, 1999.
Reprints: Mette D. Hazenberg, Department of Clinical
Viro-Immunology, Plesmanlaan 125, 1066 CX Amsterdam, The Netherlands.
The publication costs of this
article were defrayed in part by
page charge payment. Therefore,
and solely to indicate this fact,
this article is hereby marked
"advertisement"
in accordance with 18 U.S.C.
section 1734.
| |
References |
1.
Ho DD, Neumann AU, Perelson AS, Chen W, Leonard JM, Markowitz M.
Rapid turnover of plasma virions and CD4 lymphocytes in HIV-1 infection.
Nature.
1995;373:123-126[Medline]
[Order article via Infotrieve].
2.
Wei X, Ghosh SK, Taylor ME, et al.
Viral dynamics in human immunodeficiency virus type 1 infection.
Nature.
1995;373:117-122[Medline]
[Order article via Infotrieve].
3.
Rosenzweig M, DeMaria MA, Harper DM, Friedrich S, Jain RK, Johnson RP.
Increased rates of CD4+ and CD8+ T lymphocyte turnover in simian immunodeficiency virus-infected macaques.
Proc Natl Acad Sci U S A.
1998;95:6388-6393[Abstract/Free Full Text].
4.
Mohri H, Bonhoeffer S, Monard S, Perelson AS, Ho DD.
Rapid turnover of T lymphocytes in SIV-infected rhesus macaques.
Science.
1998;279:1223-1227[Abstract/Free Full Text].
5.
Wolthers KC, Wisman GBA, Otto SA, et al.
T-cell telomere length in HIV-1 infection: No evidence for increased CD4+ T cell turnover.
Science.
1996;274:1543-1547[Abstract/Free Full Text].
6.
Wolthers KC, Otto SA, Wisman GBA, et al.
Normal T cell telomerase activity and upregulation in HIV-1 infection.
Blood.
1998;93:1011-1019[Abstract/Free Full Text].
7.
Fleury S, De Boer RJ, Rizzardi GP, et al.
Limited CD4+ T-cell renewal in early HIV-1 infection: effect of highly active antiretroviral therapy.
Nat Med.
1998;4:794-801[Medline]
[Order article via Infotrieve].
8.
Sachsenberg N, Perelson AS, Yerly S, et al.
Turnover of CD4+ and CD8+ T Lymphocytes in HIV-1 infection as measured by Ki-67 antigen.
J Exp Med.
1998;187:1295-1303[Abstract/Free Full Text].
9.
Clark DR, De Boer RJ, Wolthers KC, Miedema F.
T cell dynamics in HIV-1 infection.
Adv Immunol.
1999;73:301-327[Medline]
[Order article via Infotrieve].
10.
Roederer M, Gregson Dubs J, Anderson MT, Raju PA, Herzenberg LA, Herzenberg L.
CD8 naive T cell counts decrease progressively in HIV-infected adults.
J Clin Invest.
1995;95:2061-2066.
11.
Roederer M.
T cell dynamics of immunodeficiency.
Nat Med.
1995;1:621-622[Medline]
[Order article via Infotrieve].
12.
Margolick JB, Mu-oz A, Donnenberg AD, et al.
for the Multicenter AIDS Cohort Study. Failure of T-cell homeostasis preceding AIDS in HIV-1 infection.
Nat Med.
1995;1:674-680[Medline]
[Order article via Infotrieve].
13.
Autran B, Carcelain G, Li TS, et al.
Positive effects of combined antiretroviral therapy on CD4+ T cell homeostasis and function in advanced HIV disease.
Science.
1997;277:112-116[Abstract/Free Full Text].
14.
Pakker NG, Notermans DW, De Boer RJ, et al.
Biphasic kinetics of peripheral blood T cells after triple combination therapy in HIV-1 infection: a composite of redistribution and proliferation.
Nat Med.
1998;4:208-214[Medline]
[Order article via Infotrieve].
15.
Douek DC, McFarland RD, Keiser PH, et al.
Changes in thymic function with age and during the treatment of HIV infection.
Nature.
1998;396:690-695[Medline]
[Order article via Infotrieve].
16.
Kelleher AD, Carr A, Zaunders J, Cooper DA.
Alterations in the immune response of human immunodeficiency virus (HIV) infected subjects treated with an HIV-specific protease inhibitor, Ritonavir.
J Infect Dis.
1996;173:321-329[Medline]
[Order article via Infotrieve].
17.
Bucy RP, Hockett RD, Derdeyn CA, et al.
Initial increase in blood CD4+ lymphocytes after HIV antiretroviral therapy reflects redistribution from lymphoid tissues.
J Clin Invest.
1999;103:1391-1398[Medline]
[Order article via Infotrieve].
18.
Wang L, Chen JJY, Gelman BB, Konig R, Cloyd MW.
A novel mechanism of CD4 lymphocyte depletion involves effects of HIV on resting lymphocytes: induction of lymph node homing and apoptosis upon secondary signaling through homing receptors.
J Immunol.
1999;162:268-276[Abstract/Free Full Text].
19.
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[Medline]
[Order article via Infotrieve].
20.
Zhang ZQ, Notermans DW, Sedgewick G, et al.
Kinetics of CD4+ T cell repopulation of lymphoid tissues after treatment of HIV-1 infection.
Proc Natl Acad Sci U S A.
1998;95:1154-1159[Abstract/Free Full Text].
21.
Bom-van Noorloos AA, Van Beek AAM, Melief CJM.
Cryopreservation of cells for immunological typing of non-Hodgkin lymphomas.
Cancer Res.
1980;40:2890-2894.
22.
Cohen Stuart JWT, Schuurman R, Burger D, et al.
Randomized trial comparing saquinavir soft gelatin capsules versus indinavir as part of triple therapy (CHEESE study).
AIDS.
1999;13:F53-F58[Medline]
[Order article via Infotrieve].
23.
Hamann D, Baars PA, Rep MHG, et al.
Phenotypic and functional separation of memory and effector human CD8+ T cells.
J Exp Med.
1997;186:1407-1418[Abstract/Free Full Text].
24.
Baars PA, Maurice MM, Rep M, Hooibrink B, Van Lier RAW.
Heterogeneity of the circulating human CD4+ T-cell population: further evidence that the CD4+ D45RACD27 T-cell subset contains specialized primed cells.
J Immunol.
1995;154:17-25[Abstract].
25.
Gerdes J, Lemke H, Baisch H, Wacker H-H, Schwab U, Stein H.
Cell cycle analysis of a cell proliferation-associated human nuclear antigen defined by the monoclonal antibody Ki-67.
J Immunol.
1984;133:1710-1715[Abstract].
26.
Gerdes J, Schlueter L-LC, Duchrow M, et al.
Immunobiochemical and molecular biologic characterization of the cell proliferation-associated nuclear antigen that is defined by monoclonal antibody Ki-67.
Am J Pathol.
1991;138:867-873[Abstract].
27.
Schwarting R, Gerdes J, Niehus J, Jaeschke L, Stein H.
Determination of the growth fraction in cell suspensions by flow cytometry using the monoclonal antibody Ki-67.
J Immunol Meth.
1986;90:65-70[Medline]
[Order article via Infotrieve].
28.
Bruno S, Darzynkiewicz Z.
Cell cycle dependent expression and stability of the nuclear protein detected by Ki-67 antibody in HL-60 cells.
Cell Prolif.
1992;25:31-40[Medline]
[Order article via Infotrieve].
29.
Wolthers KC, Noest AJ, Otto SA, Miedema F, DeBoer RJ.
Normal telomere lengths in naive and memory CD4+ T cells in HIV-1 infection: a mathematical interpretation.
AIDS Res Hum Retroviruses.
1999;15:1053-1062[Medline]
[Order article via Infotrieve].
30.
Meyaard L, Otto SA, Keet IPM, Roos MThL, Miedema F.
Programmed death of T cells in HIV-1 infection: no correlation with progression to disease.
J Clin Invest.
1994;93:982-988.
31.
Hengel RL, Jones BM, Kenndey MS, Hubbard MR, McDougal JS.
Lymphocyte kinetics and precursor frequency-dependent recovery of CD4+ CD45RA+ CD62L+ naive T cells following triple-drug therapy for HIV type 1 infection.
AIDS Res Hum Retroviruses.
1999;15:435-443[Medline]
[Order article via Infotrieve].
32.
Ogg GS, Jin X, Bonhoeffer S, et al.
Quantitation of HIV-1-specific cytotoxic T lymphocytes and plasma load of viral RNA.
Science.
1998;279:2103-2106[Abstract/Free Full Text].
33.
Grossman Z, Herberman RB, Dimitrov DS.
T cell turnover in SIV infection.
Science.
1999;284:555a[Free Full Text].
34.
Wolthers KC, Schuitemaker H, Miedema F.
Rapid CD4+ T-cell turnover in HIV-1 infection: a paradigm revisited.
Immunol Today.
1998;19:44-48[Medline]
[Order article via Infotrieve].
35.
Clark DR, De Boer RJ, Wolthers KC, Miedema F.
T cell dynamics in HIV-1 infection.
Adv Immunol.
1999;73:301-327.
36.
Hazenberg MD, Clark DR, Miedema F.
Tilted balance of T cell renewal in HIV-1 infection.
AIDS Rev.
1999;1:67-73.
37.
Rowland-Jones S.
HIV infection: where have all the T cells gone?
Lancet.
1999;354:5-7[Medline]
[Order article via Infotrieve].
38.
Meyaard L, Otto SA, Jonker RR, Mijnster MJ, Keet RPM, Miedema F.
Programmed death of T cells in HIV-1 infection.
Science.
1992;257:217-219[Abstract/Free Full Text].
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Abstract
Introduction