Download Human Immunodeficiency Virus Pathogenesis

MAJOR ARTICLE
HIV/AIDS
Human Immunodeficiency Virus Pathogenesis:
Insights from Studies of Lymphoid Cells
and Tissues
J. Michael Kilby
Division of Infectious Diseases, Department of Medicine, University of Alabama at Birmingham
Although plasma virus load is invaluable for monitoring human immunodeficiency virus (HIV) infection, key
pathogenesis events and most viral replication take place in lymphoid tissues. Decreases in virus load associated
with therapy occur in plasma and tissues, but persistent latent infection and ongoing viral replication are
evident. Many unanswered questions remain regarding mechanisms of HIV-associated lymphocyte depletion,
but partial CD4+ cell reconstitution after therapy likely reflects retrafficking from inflamed tissues, increased
thymic or peripheral production, and decreased destruction. Rapid establishment of latent infection and the
follicular dendritic cell–associated viral pool within lymphoid tissues suggest that only early intervention could
substantially alter the natural history of HIV. If therapy is started prior to seroconversion, some individuals
retain potent HIV-specific cellular immune responsiveness that is suggestive of delayed progression. Although
complete virus eradication appears out of reach at present, more attention is being directed toward the prospect
of boosting HIV-specific immune responses to effect another type of “clinical cure”: immune-mediated virus
suppression in the absence of therapy.
The ability to quantify HIV genetic material in plasma
(virus load) has provided a clinical assay with practical
applications [1] and an experimental tool for studying
pathogenesis [2, 3]. In many cases, highly active antiretroviral therapy (HAART) results in a reduction of
plasma virus load to less than the current assay detection thresholds. Because an “undetectable” virus load
that occurs during therapy does not reflect a clinical
cure, novel approaches are necessary to determine the
nature of persistent immunodeficiency in this setting.
In parallel with these developments, a different set
Received 21 November 2000; revised 28 January 2001; electronically published
6 August 2001.
Financial support: National Institutes of Health Acute Infection and Early Disease
Research Network (grant U01 AI 41530) and University of Alabama at Birmingham
General Clinical Research Center (grant NCRR M01 RR00032).
Reprints or correspondence: Dr. J. Michael Kilby, Div. of Infectious Diseases,
Dept. of Medicine, University of Alabama at Birmingham, 908 20th St. South,
Birmingham, AL 35294-2050 ([email protected]).
Clinical Infectious Diseases 2001; 33:873–84
2001 by the Infectious Diseases Society of America. All rights reserved.
1058-4838/2001/3306-0018$03.00
of questions has evolved regarding obstacles on the path
to long-term control over viral replication: Which early
immunologic events are critical determinants for the
course of chronic disease? What is the mechanism of
HIV-associated CD4⫹ depletion, and what is the source
and potential extent of the “partial immune reconstitution” of CD4⫹ cells after HAART? What are the reservoirs for persistent infection in the face of potent
plasma virus load suppression? What role will immunebased strategies play in improving long-term outcomes?
This review focuses on recent insights from studies involving lymphoid cells and tissues that shed light on
HIV disease mechanisms and provide new perspectives
on these unresolved questions.
CLINICAL COURSE OF PROMINENT
LYMPHOID TISSUE
Generalized lymphadenopathy, tonsillar enlargement,
and splenomegaly were prominent features in the earHIV/AIDS • CID 2001:33 (15 September) • 873
liest clinical descriptions of acute HIV infection [4], and French
investigators initially referred to the causative agent of AIDS
as “lymphadenopathy-associated virus” [5]. Lymphadenopathy
is described in 175% of reported cases of acute HIV syndrome;
it is the second-most-common reported feature after fever [6].
Although there is often a gradual decline in lymphadenopathy after seroconversion, enlarged lymph nodes may persist
throughout the course of disease. Enlarged lymph nodes within
the salivary glands can result in salivary duct obstruction and
the formation of large lymphoepithelial cysts [7, 8]. Previously,
generalized adenopathy was believed to be associated with disease progression and served as a clinical marker for HIV staging
[9], but this is an outmoded concept, because there is no evidence that chronic adenopathy has prognostic implications. In
fact, in late-stage HIV disease, the architecture of lymph nodes
is completely disrupted, germinal centers involute, and adenopathy may become less prominent [10]. Precipitous or asymmetric lymph node enlargement in later stages of disease often
suggests an opportunistic infection or neoplastic condition
rather than HIV disease progression alone. In early or advanced
HIV infection, substantial intrathoracic, intra-abdominal, or
retroperitoneal adenopathy should also prompt an evaluation
for other infectious or neoplastic complications.
Splenomegaly is also very common among patients with
early, asymptomatic HIV infection (prevalence, 23%, as determined by means of physical examination, and 66%, as determined by means of ultrasound). Like adenopathy, uncomplicated chronic splenomegaly in this setting does not appear to
correlate with specific clinical events or to predict more rapid
disease progression [11].
LYMPHOID TISSUES IN TRANSMISSION
AND EARLY INFECTION
Expansion of lymph nodes in patients with acute infection
corresponds with proliferation of HIV-specific lymphocytes and
follicular hyperplasia; however, inflammatory cell recruitment
from the circulation is probably largely responsible. Virus likely
spreads to regional lymphoid tissues within hours of mucosal
acquisition (figure 1). In primate simian immunodeficiency
virus (SIV) models, virus replication is detectable in iliac lymph
nodes within a few days of vaginal inoculation [12] and numerous infected cells are present in tissues in !2 weeks [13].
Detectable viremia likely follows lymph node involvement; it
typically occurs in humans within 1–2 weeks of genital acquisition [6, 14].
Individual HIV isolates may use different pathways to reach
lymphoid tissues. The affinities of HIV strains for distinct host
receptors influence cellular tropism and, therefore, impact on
the patterns of HIV tissue involvement. Previously, the only
known receptor for HIV was CD4⫹, which normally mediates
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Figure 1. Proposed pathogenesis mechanisms of acute HIV infection,
from initial mucosal exposure to establishment in lymph node tissues
(memory cells shown with darker nuclei; HIV-infected lymphocytes contain
white triangles).
interactions between T helper cells and major histocompatibility complex class II molecules on antigen-presenting cells.
Recent discoveries, however, reveal HIV entry into cells also
requires one of the chemokine receptors (CCR5 and CXCR4)
[15–17]. HIV transmission frequently involves virus strains that
use CCR5 (R5 viruses, previously designated “macrophage-tropic”) [18]; a specific CCR5 mutation confers relative protection
from infection [19]. Although viruses that bind CXCR4 (X4
or “T-cell tropic” viruses that induce syncytium formation in
cell cultures) may predominate the late-stage disease virus pool
[20], there is a selective advantage in the transmission of R5
over X4 variants in the setting of bloodborne, sexual, and maternal or fetal HIV exposures [21, 22].
Following mucosal exposure, HIV or SIV may first come
into contact with monocyte/macrophage cells [12, 23]. Particularly noteworthy are Langerhans’ cells, which are potent T
cell–activating interdigitating dendritic cells found in the lamina propria [24, 25] that express CD4⫹ and CCR5 [26]. HIVinfected tonsil and adenoid tissues contain dendritic cells that
are capable of transferring HIV infection to CD4⫹ cells [27],
which relates to transmission via oral sexual contact and may
be analogous to the pathogenesis of genital transmission. Gen-
ital and rectal mucosa dendritic cells express a receptor that
enables transfer of HIV infection to CD4⫹ cells without necessitating virus entry into the mononuclear cells [28].
Langerhans’ cells that bear HIV particles may be carried from
genital mucosa to lymph nodes via afferent lymphatics (figure
1). Although the interdigitating dendritic cells in skin are a different type of cell than are follicular dendritic cells (FDC) that
are found in lymph nodes (figure 2), there is evidence from
murine models that certain mucosal monocytes are capable of
migrating to lymph nodes where they mature into potent T
cell–activating, antigen-presenting cells [29, 30]. Activated T
helper cells can then facilitate humoral and cellular immune
responses against the invading pathogen. HIV-specific T cells
interact with antigen-specific B cells on the border of lymphoid
follicles in the cortex, and B cell proliferation results in follicular
hyperplasia. This interaction results in HIV-specific antibody responses. Once activated in this process, however, T cells are much
more susceptible to HIV infection than are resting T cells [31].
This model, which involves transfer of HIV-1 from monocytes
to T helper cells, may not be essential to HIV-1 pathogenesis,
because CD4⫹ T cells that express CCR5 are also present at
mucosal surfaces during inflammatory disease [30]. This is potentially pertinent to the increased risk of sexual HIV transmission in the presence of ulcerative genital infections [32]. Once
T cells become infected by this direct route, HIV dissemination
via the bloodstream would quickly follow without the need for
monocyte intermediaries. When SIV-infected primates were euthanized on selected days after the vaginal inoculation of a dualtropic virus strain, ∼10% of infected lymphoid tissue cells were
CD68⫹ (monocyte/macrophage lineage), but the majority were
CD4⫹ lymphocytes [13]. Studies of human lymph nodes in cases
of established HIV infection suggest that the majority of infected
cells are lymphocytes [33], but monocyte/macrophages in tissue
may harbor virus when plasma viremia is relatively low [34], and
they may increase virus production during opportunistic infections [35].
Although most HIV studies have focused on blood and
lymph node compartments, the majority of lymphoid cells are
contained in gut-associated lymphoid tissue. Because there are
higher concentrations of activated CD4⫹ T cells in the gut than
there are in peripheral nodes, this tissue may be a major target
during acute HIV infection. In the rhesus macaque, a decrease
in the ratio of CD4⫹ cells to total T cells occurs near the
intestinal epithelium within days of acute SIV infection, before
similar shifts are observed in peripheral lymph nodes or the
spleen [36]. The same phenomenon may occur in the human
gut during early HIV infection [37].
Changes in lymph node architecture and virus distribution
in lymphoid tissue during the prolonged course of disease could
reflect changes in viral tropism. The transition from R5 viruses
to X4 or dual-tropic viruses during the course of years in some
Figure 2. Changes in lymphoid architecture during the course of HIV
infection and treatment. Lymph node enlargement is a common manifestation of acute HIV infection. Note follicular dendritic cells (FDC) networks where antigen-presenting cells interact with naı̈ve T cells (inset;
viral particles/proteins represented by white triangles) and high concentrations of virus concentrated around germinal centers (represented by
gray-white “clouds”). In cases of advanced disease, the architecture tends
to involute, with fewer discrete follicles, loss of the FDC network, and
more scattered distribution of infected cells (white circles). Successful
virus suppression during therapy results in a partial reversal of this
disorganization.
patients appears to herald more rapidly progressive disease [20],
but what drives this evolution in cellular tropism remains unclear [38–40]. Ex vivo lymphoid tissue models suggest that R5
virus causes less cytopathicity than does X4 virus [41, 42].
Because CCR5⫹ cells make up a minority of total lymph node
cells, an alternative view is that loss of these cells is simply not
as disruptive to lymph node architecture [43, 44].
EARLY DETERMINANTS OF VIRAL
PERSISTENCE AND STEADY STATE VIRUS
LOAD
Lymphoid tissue biopsies obtained from animal models and
humans with acute infection reveal infected “activated” and
“resting” CD4⫹ T cells [13]. The latter cells may have been
infected while “activated” and then returned to a “resting” state.
HIV-infected resting T cells, while not actively contributing to
viremia, may represent the major challenge to curative therapy.
Inhibitors of viral enzymes (reverse transcriptase and protease)
have no effect on a cell with integrated but quiescent viral
genetic material. The rapid establishment of a latently HIVinfected cell pool, persisting long after initiation of suppressive
antiretroviral therapy, has been well documented [45–47]. The
life span of latently infected cells may be quite long [48, 49],
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because they are also not conspicuous targets for immune
surveillance.
It has been estimated that T cells in the circulating blood
represent only ∼1%–2% of the total body pool, whereas the vast
majority of potential HIV target cells are localized to tissue. Although SIV-infected primate models demonstrate that occasional
infected cells can be found in virtually every organ system in
subjects with late-stage disease, T cells in lymphoid tissues apparently remain the predominant contributors to the total body
burden of viral replication [33, 50]. Patterns of infected cell
distribution in nodes and the spleen suggest localized propagation of HIV takes place within lymphoid tissues [33, 51].
Increasing evidence points to the greater importance of HIVspecific cellular immune responses, relative to humoral responses, in determining the course of early events in HIV disease. Because T helper cells, which are the orchestrators of
adaptive immune responses, are the principal targets for HIV,
an infected host may rapidly lose the ability to mount potent
and durable HIV-specific cellular immune responses. HIV-infected individuals generally do not maintain HIV-specific CD4⫹
responses detectable by conventional methods [52]. Rare individuals who maintain low virus levels without therapy and
who have delayed clinical progression tend to possess unusually
strong in vitro HIV-specific CD4⫹ lymphoproliferative responses [52, 53]. When aggressive antiviral therapy is instituted
very early, before seroconversion, selected individuals retain
potent HIV-specific CD4⫹ responsiveness analogous to the situation in “long-term nonprogressors” [52]. Preserving adequate T cell help may be critical in the promotion or stabilization of clonal expansion of effector cells, particularly
antigen-specific CD8⫹ cytotoxic T cells (CTL) that are known
to limit dissemination in other viral infections.
Exploratory clinical trials that have evaluated the effects of
infusing HIV-specific cytotoxic T lymphocytes (CTL) in HIVinfected subjects suggest that the interactions between HIVinfected cells and HIV-specific CTL occur predominantly
within lymph nodes [54]. Therefore, the key immunologic
events in HIV infection—the complex interplay of viruses infecting T cells, priming T cells for antiviral responses, and the
targeted lysis of infected cells via CTLs—all are centered in
lymphoid tissues.
as others, demonstrates that the majority of viral signal in lymphoid tissues during early disease is present on the surfaces of
FDC (figures 2 and 3), compared with a relatively small number
of productively infected tissue cells.
PCR assays for HIV RNA reveal high levels of virus in blood
at all stages of untreated disease [56]. Furthermore, the large
FDC-associated viral pool appears to be fully established extremely early—within weeks of initial infection—rather than
developing gradually [57]. The increasing level of free virus in
plasma during the course of untreated disease likely reflects the
progeny from an increasing number of infected lymphoid tissue
cells [58] rather than the transition of viral replication from
tissues to blood.
Although some investigations suggest slower virologic clearance from lymph nodes than from plasma during therapy [59,
60], it is now generally accepted that highly potent combination
therapy suppresses virus levels in lymphoid tissues to a similar
degree as that observed in plasma [61, 62]. In fact, the frequency
of productively infected tissue cells falls with an initial half-life
of ∼1 day [63]. When computerized image analysis was applied
to serial paraffin-fixed tonsil biopsies obtained from patients
who were commencing therapy, not only were productively
infected cells noted to decrease dramatically, but FDC-associated HIV RNA also decreased 11000-fold [63]. After therapy,
rare positive cells were detected in tissues, but they appeared
to contain relatively few copies of viral RNA. Another study
attempted to analyze virus distribution and tissue architecture
in more advanced patients, before and several weeks after initiating HAART [58]. Cervical lymph nodes were excised intact
with minimal local tissue damage and “snap frozen” with liquid
VIRAL DYNAMICS AND LYMPHOID TISSUES
A quantitative technique, which is based on in situ hybridization for HIV RNA and computerized image analysis, has been
developed to study the distribution and dynamics of HIV-infected cells in tissues. Diffuse signal in germinal centers is interpreted as virus in the network of extracellular dendrites surrounding FDC, whereas a discretely localized signal denotes
intracellular infection [55]. This experimental approach, as well
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Figure 3. Photomicrograph of a germinal center within an HIV-infected
lymph node tissue biopsy specimen. An in situ hybridization assay has
been used which stains HIV RNA a darker color than surrounding cells.
To the left and slightly above the center, a single lymphocyte with discreet
staining suggests intracellular infection. On the right side of the figure,
there is a more diffuse “cloud” of staining consistent with extracellular
viral material trapped in the follicular dendritic cell network. (Photo courtesy of Dr. R. Pat Bucy.)
nitrogen in the operating room. A strategy involving multiple
analyses of the same tissue sample—in situ hybridization for
HIV RNA, limiting dilution, and bulk tissue PCR—was used.
Unlike the findings in a cohort that included acutely infected
subjects [64], this study, which involved cases of more advanced
disease, found a direct correlation between the frequency of
infected cells in tissue and plasma virus load. Regardless of the
frequency of cellular infection in tissue or the plasma virus
load, the mean viral RNA per infected cell remained relatively
constant.
It has been proposed that the FDC-associated virus represents a latent reservoir with potential to resume infectiousness
after prolonged sequestration in immune complexes [33]; however, the rapid establishment of this lymph node virus pool
during acute infection [57] and its rapid clearance on therapy
[63] suggest that it consists of viral particles in a high-flux
equilibrium, which is analogous to the kinetic characteristics
of plasma virus load, rather than a relatively static reservoir.
Interruption of therapy frequently results in brisk viral rebound. Infected cells, albeit at extremely low frequency, remain
in tissue when plasma virus load has been suppressed below
detectable levels [58]. Indeed, cells that contain full-length viral
transcripts can be detected within lymph node tissue specimens
from patients with “undetectable” plasma virus [58, 59, 63, 65].
Residual productive viral infection that occurs after prolonged
plasma virus load suppression has also been described recently
in blood [48, 66–68] and semen [69]. A latently infected cell
pool, made up of T cells that can be stimulated in vitro to
produce infectious virus, persists in blood even after years of
effective suppression of plasma viremia [46, 47]. The interplay
between actively replicating and latently infected cell pools is
not understood, but distinct lymphocyte populations exist in
peripheral blood, as determined by the expression of viral
mRNA with or without in vitro activation [48].
Theoretically, the most favorable situation to evaluate the
possibility of viral eradication might involve an individual who
initiates potent therapy during acute infection. Unfortunately,
even among subjects who initiate HAART within weeks of exposure and who achieve durable plasma virus load suppression,
HIV RNA expression remains detectable by in situ hybridization studies in peripheral blood cells and in lymph node, tonsil,
and gut tissues [65, 70]. Therefore, novel approaches to therapy
are needed in addition to potent antiviral drugs. The optimal
design of immune-based strategies is dependent on an understanding of the cellular dynamics underlying HIV infection.
LYMPHOID CELLULAR DYNAMICS
IN PATIENTS WITH HIV INFECTION
One approach to the study of HIV-associated cellular dynamics
is to evaluate the myriad toxic effects of the virus on CD4⫹
lymphocytes in vitro, including direct cytopathicity; CTL-mediated lysis of infected cells; indirect mechanisms, such as triggering of apoptosis by viral components (envelope glycoproteins or TAT); and interference with maturation of T cells from
stem cells (figure 4). The challenge for future research is not
merely to document whether these mechanisms exist in vitro,
but to quantify relative contributions of each mechanism to
HIV immunopathogenesis in different clinical settings.
Another approach to the problem is to develop accurate,
reproducible methods to estimate lymphocyte “turnover” rates
in HIV-infected patients, compared with uninfected controls.
CD4⫹ T cell depletion typically occurs very gradually, leading
to increased risk for opportunistic infections and neoplasms
only after many years. It has been difficult to discern whether
HIV-induced CD4⫹ T cell depletion is predominately related
Figure 4. Proposed mechanisms of altered lymphocyte dynamics in
the setting of HIV infection and treatment. HIV infection results in gradual
qualitative and quantitative deficits in CD4⫹ lymphocyte function, but the
mechanism(s) involved are unclear. Functioning lymphopoietic epithelium
is gradually lost during childhood (box at top). The degree of residual
thymic function in adults and the potential for compensatory hypertrophy
in the setting of HIV infection have been controversial issues. The initial
increase in lymphocytes that occurs after virus load suppression on therapy may reflect redistribution of memory cells (darker nuclei; infected
cells contain white triangles) from inflamed tissues, followed by gradual
return of naı̈ve T cells (lighter nuclei).
HIV/AIDS • CID 2001:33 (15 September) • 877
to increased destruction (which might be secondary to direct
cytopathicity or spontaneous apoptosis, for example) or to decreased production of lymphocytes. Establishment of the circulating T cell pool begins when pluripotent lymphocyte precursors produced in fetal bone marrow migrate to the thymus
to undergo maturation. The key to this process is the rearrangement of T cell–receptor genes so that lymphocytes develop the ability to recognize specific foreign antigens, and
distinguish them from self-antigens, before migrating to secondary lymphoid organs, including lymph nodes, the spleen,
and gut-associated lymphoid tissue [71].
The role of the spleen in the pathogenesis of HIV remains
poorly understood. HIV-infected patients who undergo splenectomy (because of immune-mediated thrombocytopenia, for
example) typically have an abrupt increase in the absolute number of circulating CD4⫹ T cells [72, 73]. This likely reflects the
release of a large number of CD4⫹ cells from splenic sequestration [72] rather than renewed production of T cells. Recommendations have been made to monitor the percentage of
total lymphocytes that are CD4⫹ as a clinical benchmark in
this setting because the absolute CD4⫹ count may be misleading
after splenectomy [73]. Small, preliminary reports suggest the
possibility that splenectomy may improve the course of HIV
disease beyond merely elevating the absolute CD4⫹ cell count
[74, 75]; however, larger studies are needed to confirm this
observation.
The “tap and drain” hypothesis of HIV pathogenesis derived
from an assumption that ongoing CD4⫹ cell losses over the
prolonged course of HIV infection are nearly balanced on a
daily basis by production of new uninfected CD4⫹ cells [2]. If
viral cytopathic effects were concentrated on mature, activated
T cells, then a major research focus should be the capacity to
maintain substantial circulating CD4⫹ cell numbers over many
years in the face of rapid HIV turnover. If the only source of
adult T cell replenishment were proliferation of the differentiated peripheral T cell pool, then this would result in an inability to develop efficient immune responses to newly encountered pathogens (and also an inability to replace
HIV-specific lymphocytes, which are apparently lost during initial HIV infection).
T cells are generally understood to be long-lived cells that
are not constantly replenished like other blood cells [76–79].
Thymic epithelial tissue gradually shrinks with age, whereas the
perivascular space, containing adipose tissue and inflammatory
cells that migrate in from the periphery, increases during adulthood (figure 4) [80]. There is a corresponding age-dependent
decline in the capacity for proliferation of naı̈ve T cells following myeloablative chemotherapy [81], which supports the hypothesis that most T cells in an older adult represent original
fetal thymus cells or daughter cells derived from clonal proliferation of these cells.
878 • CID 2001:33 (15 September) • HIV/AIDS
CT scans suggest compensatory thymic hypertrophy in response to depletion of CD4⫹ cells during the course of HIV
disease in some adults [82]; however, autopsy materials from
adults with AIDS have not always shown evidence of significant
thymopoiesis, regardless of the thymic CT appearance [83].
Furthermore, myasthenia patients who undergo thymectomy
before becoming HIV infected or early in the course of infection
manifest gradual CD4⫹ cell depletion typical of HIV-infected
subjects without thymectomy and are not precluded from favorable CD4⫹ count responses to antiretroviral therapy [83].
The length of telomeres, DNA segments on the ends of chromosomes, inversely correlates with the number of previous cell
proliferation cycles in a cell line. Cellular turnover estimates
based on telomere length suggest the CD4⫹ cell pool is not
turning over rapidly in HIV-infected patients [84, 85]. Other
surrogate assays for heightened cellular activation and proliferation, using isotope-labeled glucose uptake [86] or the nuclear antigen Ki67 [87, 88], however, arrive at the opposite
conclusion. Cellular proliferation assays based on bromodeoxyuridine lymphocyte labeling suggest that SIV-infected macaques
have heightened CD4⫹ cell proliferation compared with uninfected controls [89, 90]. The majority of increased T cell
proliferation may be in the “memory” pools of CD4⫹, and
especially CD8⫹, populations [91]. This “turnover” could reflect
clonal proliferation of preexisting T cells rather than production
of “new” lymphocytes as a homeostatic response to depletion.
A novel assay for excisional DNA products of T cell receptor
rearrangement may distinguish between peripheral expansion
of existing naı̈ve T cells and thymic production of new cells
[92]. Theoretically, T cell–receptor excision circles (TRECs) are
present preferentially in cells recently emigrated from the thymus and then progressively dilute out as cells undergo cycles
of clonal proliferation. Predictably, adults have an age-related
decrease in recent thymic emigrants. HIV-infected adults have
a greater deficit in residual thymic function by this assay. Although the mean number of excisional DNA by-products is
lower in HIV-infected adults than in age-matched controls,
there is considerable overlap in pooled results, which suggests
that limited thymic regenerative capacity is not the universal
cause of CD4⫹ depletion in patients with AIDS [93].
CHANGES IN LYMPHOID TISSUES AFTER
THERAPY
T helper lymphoproliferative responses to specific antigens are
significantly improved in many cases following a few months
of effective virus suppression [94–96]. The levels of immune
activation markers (such as b-2–microglobulin and neopterin)
and proinflammatory cytokines (such as tumor necrosis factor–a, interferon g, interleukin (IL)-1, IL-6, macrophage inflammatory protein–1 a) decrease in blood and tissues as the
virus load decreases [97–99]. The susceptibility of lymphocytes
to virally mediated apoptosis decreases during therapy as well
[100], which may correlate with improved CD4⫹ T cell numbers and decreased disease progression [101]. Lymphoid tissue
architecture returns toward normal following potent therapy,
with more organized lymphoid follicles and FDC networks (figure 2) [102]. These immunologic improvements are reflected
by decreasing numbers of opportunistic infections [103], the
potential to safely interrupt prophylactic antibiotics for certain
infections [104, 105], and, most important, decreased AIDSrelated mortality [106, 107].
The initial increase in cell counts that occurs when patients
begin antiretroviral therapy consists of all lymphocyte populations (B cells, CD4⫹ T cells, CD8⫹ T cells), not just CD4⫹
cells. The majority of the increase in the CD4⫹ cell fraction
represents “memory” cells and not “naı̈ve” cells, which suggests
that initial improvement is due to redistribution of cells from
tissues [108]. The anti-inflammatory effects of the suppression
of virus load are associated with decreased expression of adhesion molecules (VCAM and ICAM) that normally mediate
homing of lymphocytes to inflamed tissues [98]. It is possible
that the increase in blood lymphocytes is commensurate with
the decrease in total lymphocyte populations in lymph node
specimens after therapy.
Studies of mycobacterial infections, which also involve sequestration of T cells at inflamed tissue sites, provide interesting
parallels [109–111]. Patients with active tuberculosis may have
“anergy” to mycobacterial antigen skin testing, whereas cells
with tuberculosis-specific lymphoproliferative responses can be
detected simultaneously in inflamed tissues. After antibiotic
treatment and decreased inflammation in infected tissues,
pathogen-specific lymphocytes in local tissue sites decrease simultaneously with the reversal of systemic “anergy.”
Improved immune responses to preexisting pathogens may
also bring about heightened clinical manifestations of infection
soon after initiation of HAART. Patients with advanced immunosuppression may develop localized adenitis due to Mycobacterium avium complex infection soon after commencing
HAART [103, 112, 113]. Examination of biopsy specimens of
infected lymphoid tissue reveal organized granulomas and purulent drainage from the site, which is quite different from
what is observed during the unchecked bacteremia typically
associated with Mycobacterium avium complex infection in patients with AIDS. Presumably, these patients had subclinical
systemic infections that manifested as inflammatory localized
masses only after the resurgence in circulating T cell numbers
and function on HAART. Similarly, some HIV-infected patients
with tuberculosis who start to receive HAART have paradoxical
expansion of inflammatory masses [114–116], whereas those
with a history of cytomegalovirus retinitis may develop inflammatory vitreitis [103, 117, 118]. High fevers and localized in-
flammatory reactions in this special setting may respond to
corticosteroid therapy [114]. Therefore, although the decrease
in HIV-induced immune activation releases sequestered lymphocytes from lymphoid tissue back into the circulation, the
improvement in cellular immune responses may also allow localized inflammatory manifestations to develop where there was
virtually no immune response to an opportunistic pathogen
before.
TREATMENT AND LYMPHOCYTE
“TURNOVER RATES”
After several months of therapy, many patients have a gradual
return of T cells with “naı̈ve” markers, which suggests much
more than just an incidental redistribution of cells. It appears
that residual thymic (or extrathymic) productivity contributes
to a gradual return of “naı̈ve” T cells in blood [94] and in
lymphoid tissues [119] after therapy. It is unclear to what degree
the detection of new “naı̈ve” T cells is actually due to the
transition of “memory” cells back to resting status [77, 79, 120].
Studies that have attempted to resolve whether a therapeutic
increase in lymphocyte counts represents clonal proliferation
of peripheral cells versus thymic production of cells have provided conflicting conclusions [79, 121, 122].
After virus suppression occurs during HAART, there is a
return to supranormal levels of recent thymic emigrants detectable in blood and lymphoid tissues [92], which suggests
that HIV-infected adults maintain some capacity to produce
“new” T cells with novel receptor specificities. Use of TRECs
as an estimate of turnover, however, suggests significant variability from patient to patient. Though HIV-infected individuals who had normal range values for recent thymic emigrants
before therapy may have no change after therapy, some patients
with a baseline deficit demonstrate a modest increase while
receiving therapy [93]. Furthermore, mathematical modeling
suggests the key factor in determining TREC frequency could
be the rate of naı̈ve T cell proliferation, rather than changes in
thymic output [91]. In other words, the assumption that TRECs
decrease at a nearly constant rate may be flawed. High levels
of HIV antigen may drive ongoing cellular proliferation, resulting in a relative decrease in TREC frequency secondary to
a dilutional effect (TRECs are not passed on to each daughter
cell), regardless of the status of thymic output. Conversely, the
relative increase in the frequency of TREC-containing cells after
therapy may reflect the decrease in antigen-driven cellular proliferation rather than a reversal of thymic impairment.
In the setting of untreated HIV infection, the half-life of T
cells, as determined by deuterated glucose labeling, is diminished without compensatory increases in production [123].
Soon after therapy is started, regenerative production rates
gradually increase [123]. There is evidence from experiments
HIV/AIDS • CID 2001:33 (15 September) • 879
that have involved fetal thymus organ cultures that HIV-1 can
interfere with T cell production at the level of progenitor cells,
and that, even among adults, HAART administration can improve the capacity to renew this activity [124]. At the same
time, however, the much more dynamic transitional proliferation driven by immune activation may be decreased in the
presence of therapy [51, 91]. Naı̈ve T cells that become activated
or that are caught in the transitional state of becoming effector
cells also express the Ki67 marker, which may have previously
resulted in erroneous assertions about overall turnover of the
T cell population rather than short-term fluctuations in immune activation [51].
Beyond 12 months of therapeutic virus suppression, the
“turnover rate” and the absolute production rate have returned
toward normal, determined by use of the radiolabeled glucose
uptake assay [123]. In general, “naı̈ve” T cells have a longer
half-life (in the range of 6 months) than do “memory” cells
(∼1–2 months). The latter half-life estimate, however, is complicated by the heterogeneity of the “memory” cell population,
which contains a wide spectrum of cells from effector cells
recently exposed to antigen (soon to undergo activation-induced cell death) to long-lived resting memory cells. There is
also a heterogeneous response from patient to patient, and this
appears to be at least partially explained by the thymic capacity.
That is, patients who have a substantial thymic shadow seen
on radiographic imaging have a larger number of “naı̈ve” T
cells, and this preponderance of “naı̈ve” cells, in turn, is associated with a slower turnover of the overall circulating T cell
population [123]. This observation is supported by other recent
studies that have suggested that children may have a substantial
increase in both TRECs and “naı̈ve” T cells while receiving
therapy [125, 126], and that, among adults, those with larger
CT thymic shadows have a tendency to demonstrate more
abrupt and significant “naı̈ve” T cell increases while receiving
therapy [127].
Therefore, the effects of HIV on cellular dynamics are a
complex interplay of increased destruction and interference
with production. The interpretation of cellular dynamics is further complicated by heterogeneity both at the intercellular and
interpatient levels. Any reconstitution of the naı̈ve T cell pool
that occurs while the patient is receiving HAART alone is likely
to be gradual, limited in diversity, and potentially skewed relative to the preinfection T cell repertoire.
cytokine, have significantly fewer detectable HIV-infected blood
and lymph node cells than do those patients who receive HAART
alone [128]. Nonetheless, interrupting HAART in a small number
of patients who were treated with adjunctive IL-2 still resulted
in virus rebound [129]. Although the possibility remains that
virus resurgence could be delayed compared with control subjects
when comparisons are made in larger clinical trials, concern has
been raised that activation events may replenish the latent cell
pool rather than purge it [130].
It remains plausible that another type of “clinical cure” can
be approached without necessarily resulting in absolute clearance of the latently infected cell pool. Recent reports demonstrate that intermittent therapeutic interruptions may restimulate effective HIV-specific immune responses, at least in the
setting of very early infection, to the degree that immediate
virus rebound that occurs while the patient is not receiving
therapy may not be observed [131–133]. Studies are underway
to investigate the impact of immune-based strategies—
cytokines, vaccinations, and scheduled treatment interruptions—on virus rebound kinetics when therapy is discontinued.
What is necessary is a better understanding of a viral antigen
threshold that might generate and sustain efficient HIV-specific
immune responses without causing irreversible damage in the
process. A further challenge is that this elusive threshold may
differ for each individual case on the basis of reciprocal relationships between different viral isolates and host immunologic
factors.
FUTURE TREATMENT STRATEGIES
Acknowledgments
Where do these immunologic insights lead us in terms of future
treatment strategies? One hypothesis proposes immune activation
strategies may “flush out” latently infected cell pools. Patients
with established HIV infection who receive HAART plus cycles
of IL-2, which is a T cell growth factor and immunoactivating
We thank Dr. R. Pat Bucy and Dr. Michael S. Saag, for
constant guidance and collaboration; Dr. Edward Hook, Dr.
John Gnann, Dr. Craig Hoesley, Dr. Cynthia Derdeyn, and other
members of the “UAB Pathogenesis Group,” for helpful discussions and critical readings; and the many patient volunteers
880 • CID 2001:33 (15 September) • HIV/AIDS
CONCLUSIONS
Improvements in HIV therapy have benefited patients directly
while also serving as a research probe in studies of viral replication and lymphocyte dynamics. Tissue sites of viral replication represent the driving force behind the total body viral
burden. Lymphoid tissue analysis provides a perspective on
patterns of viral distribution and cellular interaction not revealed with blood assays alone. Unique opportunities to study
the critical first encounters between the virus and the immune
system during acute infection may be particularly instructive
for designing future therapeutic strategies. The evaluation of
concurrently obtained blood and tissue samples will likely continue to be an important analytical approach to deciphering
the complexities of HIV pathogenesis.
who provide blood and tissue specimens for HIV pathogenesis
research.
23.
24.
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