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Journal of Virology, May 1999, p. 3623-3629, Vol. 73, No. 5
0022-538X/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.
Bystander Target Cell Lysis and Cytokine Production by Dengue
Virus-Specific Human CD4+ Cytotoxic T-Lymphocyte
Clones
Susan J.
Gagnon,
Francis A.
Ennis, and
Alan L.
Rothman*
Center for Infectious Disease and Vaccine
Research, University of Massachusetts Medical Center, Worcester,
Massachusetts 01655
Received 17 July 1998/Accepted 19 January 1999
 |
ABSTRACT |
Dengue hemorrhagic fever, the severe form of dengue virus
infection, is believed to be an immunopathological response to a secondary infection with a heterologous serotype of dengue virus. Dengue virus capsid protein-specific CD4+ cytotoxic
T-lymphocyte (CTL) clones were shown to be capable of mediating
bystander lysis of non-antigen-presenting target cells. After
activation by anti-CD3 or in the presence of unlabeled antigen-presenting target cells, these clones could lyse both Jurkat
cells and HepG2 cells as bystander targets. Lysis of HepG2 cells
suggests a potential role for CD4+ CTL in the liver
involvement observed during dengue virus infection. Three
CD4+ CTL clones were demonstrated to lyse cognate,
antigen-presenting target cells by a mechanism that primarily involves
perforin, while bystander lysis occurred through Fas/Fas ligand
interactions. In contrast, one clone used a Fas/Fas ligand mechanism to
lyse both cognate and bystander targets. Cytokine production by the CTL
clones was also examined. In response to stimulation with D2 antigen,
CD4+ T-cell clones produced gamma interferon, tumor
necrosis factor alpha (TNF-
) and TNF-
. The data suggest that
CD4+ CTL clones may contribute to the immunopathology
observed upon secondary dengue virus infections through direct
cytolysis and/or cytokine production.
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INTRODUCTION |
Dengue virus (DV) is a
mosquito-borne flavivirus which is prevalent in many tropical and
subtropical areas of the world. Infecting up to 100 million individuals
yearly (10), DV infection can be either asymptomatic or it
can present as one of two forms of disease. The most common form,
classical dengue fever (DF), presents as a flu-like syndrome,
with symptoms including fever, retroorbital headache, muscle aches, and
bone pain. Liver involvement is also a frequent manifestation of DV
infection, characterized by hepatomegaly and increased plasma
levels of liver transaminases (19, 24, 35). Patients with
dengue hemorrhagic fever (DHF), the more severe form of disease,
develop hemoconcentration, thrombocytopenia, and increased
capillary permeability, resulting in plasma leakage and sometimes
hemorrhage. At its most severe, DHF can culminate in circulatory shock
and death.
DV exists as four distinct serotypes, called types 1, 2, 3, and 4. After infection with DV, lifelong protective immunity is induced
against the infecting serotype, although immunity to heterologous serotypes is short-lived (38). Studies have suggested that
the immune response mounted upon reinfection with a heterologous
serotype of DV may have detrimental consequences for the host. The vast majority of DHF cases have been shown to occur in individuals who are
undergoing a secondary DV infection, suggesting that preexisting immunity to one serotype of DV may predispose an individual to the
development of more severe disease (10).
A number of research studies from our laboratory have shown that DV
serotype-cross-reactive memory CD4+ and CD8+ T
cells exist after primary DV infection (reviewed in reference 28). Reactivation of serotype-cross-reactive memory
T cells upon secondary infection possibly results in the increased
level of T-cell activation often observed in individuals with DHF
(27). Induction of immunopathology by CD4+ T
lymphocytes may occur by various mechanisms, including cell-mediated cytotoxicity and/or cytokine production.
CD4+ T-cell-mediated cytotoxicity is thought to occur via
two main pathways: release of perforin and granzymes from the activated cytotoxic T lymphocytes (CTL) (32, 44) or the interaction of
Fas ligand on the T cell with Fas on the target cell (18, 32, 42,
44). The Fas/FasL pathway could contribute to the destruction of
both cells presenting viral antigens as well as non-antigen-presenting
"innocent bystander" cells which are expressing Fas, as has been
demonstrated in other systems (4, 31, 40). Although the
existence of virus-specific CD4+ CTL in humans has been
demonstrated after viral infection or immunization (16, 26, 36,
49), the mechanism of cytolysis utilized by human virus-specific
CD4+ CTL clones has not been extensively characterized. The
direct destruction of certain cell types, either by cognate or
bystander lysis, may result in some of the pathology which occurs in DHF.
Liver disease is commonly observed in patients with DV infections.
Elevated levels of liver enzymes are detectable in serum, indicating
hepatocyte injury, and frequently the liver is enlarged. Kupffer cells
appear to be the primary cell type supporting DV infection in the liver
(9); however, there is some debate as to whether hepatocytes
can be infected with DV (9, 15). The cause of hepatocyte
injury during DV infection is unknown, and we postulate that
CD4+ CTL may mediate liver damage through a mechanism
involving bystander lysis.
In addition to direct cytolysis, cytokine production by activated T
cells may contribute to severe dengue disease. Some reports have
suggested that production of tumor necrosis factor alpha (TNF-) is
increased in cases of severe dengue illness (23, 48),
although production by DV-specific T lymphocytes has not been
demonstrated. When administered experimentally to human volunteers, TNF- has been shown to induce symptoms similar to those observed in
DHF, including hypotension, capillary leakage, and a flu-like syndrome
consisting of headache, nausea, and fatigue (14, 39, 41).
Additionally, it has been demonstrated that both interleukin-2 (IL-2)
and gamma interferon (IFN-
) are elevated in DV-infected individuals
compared to healthy controls or children with other febrile illnesses
(27).
Previously, a panel of seven CD4+ CTL clones was generated
from a D4-immune donor (7). These clones were shown to
recognize the DV capsid protein, as well as DV-infected
B-lymphoblastoid cell lines (BLCL). Six of the seven clones were
demonstrated to be cross-reactive between the D2 and D4 serotypes of
DV, indicating their potential to be reactivated in vivo in the event
of a secondary D2V infection. In this study, a functional analysis of
these CD4+ T-cell clones was performed, in which we
examined both their mechanisms of target cell lysis and secretion of
IFN-, TNF-, and TNF- after stimulation. Results of these
analyses suggest several potential pathological roles of DV-specific
CD4+ T cells activated during DV infection.
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MATERIALS AND METHODS |
Establishment of CTL clones.
Leukocytes were obtained by
leukopheresis from a human volunteer at 6 months postvaccination with
an experimental live-attenuated D4V vaccine, 314750, and DV-specific
CD4+ CTL clones were established by using a limiting
dilution technique as previously described (7). Clones were
restimulated every 14 days with autologous peripheral blood mononuclear
feeder cells, recombinant human IL-2 (Collaborative Biomedical
Products) at a final concentration of 20 U/ml, and the anti-CD3
antibody 12F6, kindly supplied by Johnson Wong (Massachusetts General
Hospital, Boston, Mass.).
Recognition of the DV capsid protein by these T-cell clones was
determined as described earlier (7). Clones 8G5, 6E2, and 7E4 are D2/D4-cross-reactive and have been shown to recognize an
epitope located between amino acids (aa) 83 and 92 of the D4V capsid
protein. An additional clone (5C8) was identified that was specific for
D4, which recognized aa 43 to 55 of the D4V capsid protein.
Bystander lysis assay.
The expression of functional FasL on
the T-cell clones was assessed by bioassay for lysis of Jurkat cells as
described earlier (37). Briefly, a 24-well plate was coated
with anti-CD3 antibody by adding monoclonal antibody 12F6 (Johnson
Wong) at 5 µg/ml in phosphate-buffered saline (PBS) to the wells
followed by incubation at 4°C overnight. Wells were then washed twice
with PBS to remove unbound antibody, and 1 × 106 to
2 × 106 CTL clones were added to the wells for 4 h at 37°C for activation. After activation, clones were collected and
used in a CTL assay against 5,000 51Cr-labeled Jurkat
target cells per well at various effector/target (E/T) ratios. Lysis
was assessed after 6 and 18 h of incubation (44). To
assess the bystander lysis of liver cells, a similar experiment was
performed in which the human hepatocellular carcinoma cell line HepG2
was used as a bystander target at 2,000 cells per well.
Analysis of cytotoxic mechanisms.
CTL assays were performed
in which 2,500 Jurkat or HepG2 cells and 2,500 autologous BLCL were
present in the same well with 2.5 µg of the relevant peptide per ml
to examine the mechanism of lysis used by CTL clones when both
bystander and cognate target cells are present in the same well. For
analysis of cognate killing, BLCL were 51Cr-labeled and
Jurkat cells were unlabeled, and the assay was harvested after a 4-h
incubation. To analyze bystander lysis, Jurkat cells were
51Cr labeled, BLCL were left unlabeled, and lysis was
measured after an 8-h incubation. Clones were used directly or
preincubated in 96-well plates with either brefeldin A (BFA; catalog
number B7651; Sigma, St. Louis, Mo.) or concanamycin A (CMA; catalog
number C9705; Sigma) at various concentrations for 2 h at 37°C
prior to the addition of target cells.
The anti-human FasL antibody 4H9 (Medical & Biological Laboratories
Co., Ltd., Tokyo, Japan) was used to inhibit Fas/FasL-mediated
lysis of
D4 antigen-pulsed BLCL by adding between 0.1 and 10 µg/ml
to each
well at the start of a 5-h cytotoxicity
assay.
RT-PCR for perforin and FasL gene expression.
Reverse
transcriptase PCR (RT-PCR) to determine perforin gene expression was
performed as described by Van Voorhis et al. (43). RT-PCR
for Fas ligand gene expression was performed as described by Hargreaves
et al. (11).
Cytokine production by CTL clones.
CTL clones were tested
for cytokine production by a method similar to that used by Jassoy et
al. (17). T-cell clones (105) were plated in
96-well V-bottom plates (Costar) in a final volume of 200 µl in AIM-V
media with 10% human AB serum. Cells were stimulated with DV antigens
or Vero antigen at 1:200 or 1:320 or with peptide (capsid protein, aa
84 to 92) at 6.25 µg/ml in the presence of 105
gamma-irradiated autologous PBMC as feeder cells. DV and Vero antigens
were prepared as previously described (25). Cells were incubated at 37°C for 24 h, at which point plates were spun at 200 × g for 5 min, and 150 µl of supernatant was
collected from each well. Supernatants from replicate wells were pooled
and then divided into aliquots and stored at 
70°C until use.
Production of cytokines was analyzed with commercially available
enzyme-linked immunosorbent assay (ELISA) kits (Endogen, Boston, Mass.)
according to the manufacturers' instructions.
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RESULTS |
DV-specific CD4+ CTL clones lyse bystander target
cells.
In contrast to CD8+ T cells, which are believed
to mediate lysis of only cognate target cells presenting the relevant
peptide in the appropriate major histocompatibility complex molecule, studies indicate that CD4+ CTL can efficiently lyse both
cognate and bystander target cells (40). Bystander target
cells are defined as cells that neither present antigen nor activate
CTL but instead are lysed due to their proximity to the
antigen-presenting target cell.
Two DV-specific CD4
+ CTL clones were examined to determine
whether they were capable of lysing Jurkat cells as bystander target
cells. Jurkat cells are a human acute T-cell leukemia line that
has
been shown to be susceptible to Fas-mediated killing by T
cells
(
37). The T-cell clones were activated on anti-CD3-coated
plates for 4 h, and the lysis of Jurkat cells was then measured
in
a cytotoxicity assay. Figure
1 shows that
clones 6E2 and 7E4
lysed Jurkat target cells after 6 h of
incubation, and levels
of lysis increased after 18 h of
incubation. In the absence of
preactivation, the clones showed no
appreciable lysis (<5%) of
Jurkat target cells (data not
shown). These results indicate that
clones 6E2 and 7E4 were able to
lyse bystander target cells after
nonspecific activation with anti-CD3.
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FIG. 1.
DV-specific CD4+ CTL clones exhibit
bystander lysis of Jurkat target cells. Clones 6E2 and 7E4 were
preactivated for 4 h on an anti-CD3-coated plate and then used in
either a 6- or an 18-h cytotoxicity assay with Jurkat target cells at
the indicated E/T ratios. ND, not done. The spontaneous release was 9%
at 6 h and 18% at 18 h.
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Evidence suggests that the macrophage-like Kupffer cells of the liver
are sites of DV infection (
9). Activated T cells
responding
to the infected Kupffer cells may also damage neighboring
hepatocytes,
potentially contributing to the liver damage observed
in some cases of
DV infection. In light of this possibility, the
human hepatocellular
carcinoma cell line HepG2 was tested for
its susceptibility to
bystander lysis by the CD4
+ CTL clones. Figure
2 shows that three CD4
+ CTL
clones preactivated on anti-CD3-coated plates exhibited bystander
lysis
of
51Cr-labeled HepG2 cells, while no lysis was observed in
the absence
of preactivation.
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FIG. 2.
DV-specific CD4+ CTL clones exhibit
bystander lysis of HepG2 target cells. Clones 6E2, 7E4, and 5C8 were
either unactivated or preactivated for 4 h on an anti-CD3-coated
plate and then used in either a 5- or a 10-h cytotoxicity assay with
HepG2 target cells. E/T ratios were 15:1 for 7E4, 9:1 for 6E2, and 4:1
for 5C8. The spontaneous release was 12% at 5 h and 18% at
10 h.
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DV-specific CD4+ CTL clones kill antigen-presenting
targets by either FasL or perforin.
Experiments were performed to
determine whether lysis of cognate, antigen-presenting target cells by
the DV-specific CD4+ CTL clones was mediated by FasL or
perforin by using chemical inhibitors of either pathway. BFA is an
inhibitor of intracellular glycoprotein transport that has been
shown to selectively inhibit Fas-based cytotoxicity (20).
CMA is a specific inhibitor of vacuolar-type H+
ATPase, which acidifies vacuolar organelles (47). It
inhibits the activity of perforin in dense granules, mostly due to the accelerated degradation of the protein (20). CMA was shown
to almost completely inhibit the cytotoxicity of a human
CD8+ CTL clone against peptide-presenting target cells
(1). Figure 3 presents results
from two assays in which a representative clone, 6E2, was preincubated
with increasing concentrations of CMA or BFA to determine whether
either inhibitor could decrease target cell lysis. Figure 3A
demonstrates that a 2-h preincubation of clone 6E2 with 2 nM CMA
completely abrogated lysis of dengue antigen-pulsed target cells. In
contrast, Fig. 3B shows that a 2-h preincubation with BFA, even at a
concentration of 100 µM, only inhibited lysis of cognate target cells
by 50%. These results indicate that clone 6E2 lyses antigen-presenting
target cells by a mechanism that is primarily mediated by perforin.
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FIG. 3.
CD4+ CTL clone 6E2 lyses autologous D4
antigen-pulsed BLCL by a mechanism primarily involving perforin. (A)
Clone 6E2 was preincubated with the indicated concentrations of CMA for
2 h prior to the addition of target cells. The E/T ratio was 35:1.
(B) Clone 6E2 was preincubated with the indicated concentration of BFA
for 2 h prior to the addition of target cells. The E/T ratio was
25:1. In both assays, lysis was assessed after a 5-h incubation. The
spontaneous release was  20% for all conditions.
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Experiments were performed to determine whether one mechanism of
killing would be preferentially employed by the clones when
both
cognate and bystander target cells were present in the same
well. To
assess cognate antigen-presenting-cell lysis, autologous
BLCL were
51Cr labeled, while Jurkat bystander cells present in the
same well
were unlabeled. Lysis was measured after 4 h of
incubation. Figure
4 shows that, in the
absence of inhibitors, all four clones tested
were able to lyse cognate
BLCL presenting the epitope peptide.
CMA inhibited the lysis of BLCL
81% by clone 5C8, 94% by clone
7E4, and 100% by clone 6E2.
Preincubation of clones with BFA blocked
the lysis of BLCL to a lesser
degree: 38% by clone 5C8, 20% by
clone 7E4, and 41% by clone 6E2.
The results indicate that these
clones lyse cognate antigen-presenting
cells by a mechanism primarily
mediated by perforin. In contrast, lysis
of labeled BLCL by the
clone 8G5 was inhibited by 100% after
preincubation with BFA,
while CMA only decreased lysis by 46%,
suggesting that this clone
principally causes lysis of cognate,
peptide-presenting BLCL via
Fas/FasL interactions. Additionally, a
monoclonal anti-human FasL
antibody (4H9) also effectively blocked
lysis of antigen-presenting
target cells by clone 8G5. At 1 µg/ml,
lysis was blocked by 43%
and at 10 µg/ml an 86% inhibition of lysis
was observed (14% lysis
reduced to 8 and 2%, respectively).
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FIG. 4.
CD4+ CTL clones lyse autologous target cells
by two different mechanisms. Clones were incubated with CMA or BFA for
2 h prior to the addition of target cells, and lysis of
autologous, peptide-presenting targets was measured in a 4-h
cytotoxicity assay in the presence of unlabeled Jurkat cells as
described in Materials and Methods. The E/T ratio was 12:1 for all
clones. Clones 8G5 and 5C8 were tested in one experiment; 6E2 and 7E4
were tested in separate experiments. The spontaneous release was 25%
in all experiments.
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DV-specific CD4+ CTL clones lyse bystander target cells
primarily via Fas/FasL interactions.
In the same experiments in
which the mechanism of cognate target cell lysis was assessed (Fig. 4),
the mechanism of bystander lysis was also investigated. In these
assays, Jurkat or HepG2 bystander cells were 51Cr labeled,
and autologous BLCL present in the same well were unlabeled.
Lysis was measured after an 8-h incubation. These experiments could also determine whether bystander target cells would be
killed if clones were activated by antigen-presenting cells, as well as
after anti-CD3 stimulation as shown in Fig. 1.
Results presented in Fig.
5 show that the
CD4
+ CTL clones differ in their ability to kill Jurkat
bystander target cells in
this system. Clones 8G5 and 5C8 are
able to mediate a significant
degree of bystander lysis of Jurkat
cells when activated by antigen-presenting
BLCL; however, 7E4 and 6E2
show a much lower degree of bystander
lysis in this experiment. Jurkat
cell lysis by clones 8G5 and
5C8 was completely abrogated by
preincubation of the clones with
BFA, but preincubation with CMA only
inhibited the lysis by 24
and 47%, respectively. This suggests that
bystander lysis of Jurkat
cells by clones 8G5 and 5C8 is dependent on
Fas/FasL interactions.
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FIG. 5.
CD4+ CTL clones 8G5 and 5C8 lyse bystander
target cells via a FasL-mediated mechanism. Clones were incubated with
CMA or BFA for 2 h prior to the addition of target cells, and
lysis of labeled Jurkat cells was assessed after an 8-h incubation in
the presence of unlabeled, autologous, peptide-presenting BLCL. The E/T
ratio was 12:1 for all clones tested. Clones 8G5 and 5C8 were tested in
one experiment; clones 6E2 and 7E4 were tested in separate experiments.
The spontaneous release was 30% in all experiments.
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The mechanism of bystander lysis of HepG2 cells was assessed in the
same manner. Figure
6 demonstrates that,
in the presence
of unlabeled BLCL and the epitope peptide, lysis of
HepG2 cells
was inhibited by greater than 74% for all clones tested
after
preincubation with BFA, but preincubation of clones with CMA
resulted
in only 15% inhibition of lysis by clone 7E4 and no
inhibition
with clones 8G5 and 6E2. These results indicate that,
similar
to Jurkat cells, bystander lysis of this human hepatoma cell
line
is likely to occur by a Fas/FasL-mediated mechanism.
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FIG. 6.
CD4+ CTL clones lyse HepG2 bystander target
cells via a Fas/FasL-mediated mechanism. Clones were incubated with
either CMA at 10 nM or BFA at 10 µM for 2 h prior to the
addition of target cells. Lysis of labeled HepG2 bystander targets in
the presence of unlabeled peptide-presenting BLCL was assessed after an
8-h incubation. The E/T ratio was 10:1. The spontaneous release was
27% for all conditions.
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IFN-, TNF-, and TNF- are produced by DV-specific
CD4+ CTL clones upon activation.
Production of
cytokines by serotype-cross-reactive memory T cells reactivated during
a secondary DV infection could contribute to the immunopathology of
DHF. In light of this possibility, CD4+ CTL clones were
examined for cytokine production by ELISA after stimulation with
heterologous DV antigen. Five of five serotype-cross-reactive CD4+ T-cell clones tested produced IFN- (700 to >1,000
pg/ml) after stimulation with D2 antigen. No IFN- production was
observed upon stimulation with a Vero cell control antigen. IFN-
production could also be observed after D2 antigen stimulation of the
donor's peripheral blood mononuclear cells in bulk culture.
In separate experiments, we also detected the production of TNF-
(9 to 412 pg/ml) by four of five CD4
+ CTL clones and
production of TNF-
(14 to 462 pg/ml) by five
of five
CD4
+ CTL clones after stimulation with D2 antigen. In
comparison,
the levels of TNF-
and TNF-
after incubation with the
control
Vero cell Ag were <40 and 0 pg/ml,
respectively.
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DISCUSSION |
Experiments presented here functionally characterize a panel of DV
capsid-protein-specific CD4+ CTL clones with regard to
their mechanism of target cell lysis and cytokine secretion. Chromium
release assays with both Jurkat and HepG2 cells as bystander targets
indicate that the DV capsid-specific CD4+ CTL clones are
capable of mediating bystander lysis. Bystander lysis by human
virus-specific CD4+ T-cell clones has not previously been
demonstrated. In contrast, CD4+ CTL specific for human
immunodeficiency virus (32) and herpes simplex virus
(50) have been shown to kill only antigen-presenting targets. Our results are consistent with the observation that bystander
lysis generally occurs via interaction of FasL on the T-cell clone with
Fas, which is constitutively expressed on a number of cell types,
including Jurkat and HepG2 cells (5, 31). The DV-specific
CD4+ CTL clones express FasL mRNA after activation (data
not shown), and the killing of both Jurkat and HepG2 target cells was
abrogated by preincubation of the clones with BFA, an inhibitor of
FasL-mediated lysis.
Patients with DV infections often exhibit evidence of hepatocyte
injury, including increased plasma levels of liver enzymes (19,
24), as well as hepatomegaly and mild to moderate paracentral or
zonal necrosis noted upon microscopic examination (3).
Although bystander lysis mediated by T cells has not been demonstrated in vivo, the ability of the human hepatocellular cell line HepG2 to
serve as a bystander target for the DV-specific T-cell clones suggests
a potential role for activated DV-specific T-cell clones in this
liver pathology. HepG2 is a well-differentiated liver cell line which
retains certain liver-specific characteristics, including the synthesis
of hepatic proteins (4, 22). Hepatocyte damage may occur
when DV-specific CTL are activated by DV-infected Kupffer cells and
subsequently lyse hepatocytes via a bystander mechanism. Fas is
expressed constitutively at low levels in normal human liver
(30), and upregulation of Fas in the liver occurs after some
virus infections (8, 12). In DV-infected individuals, Bhamarapravati et al. (3) have histologically detected
Councilman bodies in liver sections, and these structures have been
postulated to be apoptotic cells (21, 29).
Only a limited number of studies have been performed examining the
mechanism of lysis utilized by human CD4+ CTL and, to our
knowledge, this is the first such characterization of human
flavivirus-specific CD4+ T cells. Experiments presented
here indicate that DV-specific CD4+ CTL clones isolated
from the same donor exhibit heterogeneity with respect to their
mechanisms of target cell lysis and further demonstrate that the
perforin- and FasL-mediated mechanisms of target cell lysis are not
mutually exclusive. Clones 7E4, 6E2, and 5C8 lyse cognate target cells
via perforin, while lysis of HepG2 or Jurkat bystander target cells
present in the same well appears to be FasL mediated. Lysis of
antigen-presenting targets by a perforin-dependent mechanism suggests
that some CD4+ CTL clones may be involved in the lysis of
virally infected cells in a manner similar to that of CD8+
CTL. The importance of CD4+ CTL in protection and recovery
from viral infection has been suggested previously by experiments
performed in mice (34, 46).
A fourth CD4+ CTL clone, 8G5, appears to kill both cognate,
peptide-pulsed targets and bystander targets by a FasL-mediated mechanism. This heterogeneity in the usage of killing mechanisms between clones recognizing the same viral epitope has not previously been observed with human virus-specific CD4+ T-cell clones,
although similar results have been obtained with a panel of human
autoreactive CD4+ T-cell clones specific for aa 83 to 99 of
myelin basic protein (44). A recent study of human
CD4+ purified-protein-derivative-specific clones by
Lewinsohn et al. (31) suggested that the killing mechanism
used by the clones was dependent on target cell susceptibility to
Fas-mediated lysis. Although BLCL were shown to express Fas, they were
relatively resistant to killing mediated by an anti-Fas antibody
(31). Although variance in target cell susceptibility to
bystander lysis is likely to explain some of our results as well, our
work also suggests that the CTL clones themselves may have differential abilities to induce target cell death via the FasL- or
perforin-mediated pathways. It is possible that upon activation clone
8G5 expresses a higher level of FasL on its surface than the other
DV-specific CD4+ CTL clones and therefore can more
efficiently cause FasL-mediated lysis of antigen-presenting BLCL.
The DV-specific T-cell clones produced IFN-, TNF-, and TNF-
after antigenic stimulation. Although production of IFN- by DV-specific T-cell clones has been documented by our group (25, 26, 33), production of TNF- and TNF- by DV-specific T-cell clones has not been previously demonstrated. T-cell clones specific for
other viruses, including human immunodeficiency virus (17), hepatitis B virus (2), and cytomegalovirus (6)
have similarly been shown to produce TNF- after stimulation. DV
serotype-cross-reactive T-cell clones from this D4-immune individual
produce cytokines after stimulation with D2 antigen, suggesting that a
secondary infection of this individual with D2V would result in the
production of TNF-, TNF-, and IFN-. Although we cannot exclude
an effect of in vitro propagation on the pattern of cytokines detected, these results are consistent with the observations that plasma levels
of IFN- and TNF- are elevated in patients with DHF (13, 27,
45, 48). These cytokines are thought to contribute to the
immunopathogenesis of DHF.
Overall, our results show that activated DV
serotype-cross-reactive memory CD4+ T lymphocytes
secrete cytokines, including IFN-, TNF-, and TNF-, and
that these T cells exhibit lysis of both cognate
antigen-presenting cells and bytander target cells. The
serotype-cross-reactive nature of these CD4+ T cells
suggests that, in vivo, a secondary infection of this individual with
D2V infection could result in the elaboration of these activities,
which might contribute to the enhanced pathology manifested as DHF.
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ACKNOWLEDGMENT |
This work was supported by a grant from the NIAID (RO1 AI30624).
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FOOTNOTES |
*
Corresponding author. Mailing address: Center for
Infectious Disease and Vaccine Research, University of Massachusetts
Medical Center, 55 Lake Ave. North, Worcester, MA 01655. Phone: (508) 856-4182. Fax: (508) 856-4890. E-mail:
Alan.Rothman{at}umassmed.edu.
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REFERENCES |
| 1.
|
Ando, K.,
K. Hiroishi,
T. Kaneko,
T. Moriyama,
Y. Muto,
N. Kayagaki,
H. Yagita,
K. Okumura, and M. Imawari.
1997.
Perforin, Fas/Fas ligand, and TNF- pathways as specific and bystander killing mechanisms of hepatitis C virus-specific human CTL.
J. Immunol.
158:5283-5291[Abstract].
|
| 2.
|
Barnaba, V.,
A. Franco,
M. Paroli,
R. Benvenuto,
G. De Petrillo,
V. L. Burgio,
I. Santilio,
C. Balsano,
M. S. Bonavita,
G. Cappelli,
V. Colizzi,
G. Cutrona, and M. Ferrarini.
1994.
Selective expansion of cytotoxic T lymphocytes with a CD4+ CD56+ surface phenotype and a T helper type 1 profile of cytokine secretion in the liver of patients chronically infected with hepatitis B virus.
J. Immunol.
152:3074-3087[Abstract].
|
| 3.
|
Bhamarapravati, N.,
P. Tuchinda, and V. Boonyapaknavik.
1967.
Pathology of Thailand haemorrhagic fever: a study of 100 autopsy cases.
Ann. Trop. Med. Parasitol.
61:500-510[Medline].
|
| 4.
|
Bouma, M.-E.,
E. Rogier,
N. Verthier,
C. Labarre, and G. Feldmann.
1989.
Further cellular investigation of the human hepatoblastoma-erived cell line HepG2: morphology and immunocytochemical studies of hepatic-secreted proteins.
In Vitro Cell. Dev. Biol.
25:267-275[Medline].
|
| 5.
|
Cruikshank, S. M.,
J. Southgate,
P. J. Selby, and L. K. Trejdosiewicz.
1998.
Expression and cytokine regulation of immune recognition elements by normal human biliary epithelial and established liver cell lines in vitro.
J. Hepatol.
29:550-558[Medline].
|
| 6.
|
Davignon, J.-L.,
P. Castanie,
J. A. Yorke,
N. Gautier,
D. Clement, and C. Davrinche.
1996.
Anti-human cytomegalovirus activity of cytokines produced by CD4+ T-cell clones specifically activated by IE1 peptides in vitro.
J. Virol.
70:2162-2169[Abstract].
|
| 7.
|
Gagnon, S. J.,
W. Zeng,
I. Kurane, and F. A. Ennis.
1996.
Identification of two epitopes on the dengue 4 virus capsid protein recognized by a serotype-specific and a panel of serotype-cross-reactive human CD4+ cytotoxic T-lymphocyte clones.
J. Virol.
70:141-147[Abstract].
|
| 8.
|
Galle, P. R.,
W. J. Hofmann,
H. Walczak,
H. Schaller,
G. Otto,
W. Stremmel,
P. H. Krammer, and L. Runkel.
1995.
Involvement of the CD95 (APO-1/Fas) receptor and ligand in liver damage.
J. Exp. Med.
182:1223-1230[Abstract/Free Full Text].
|
| 9.
|
Hall, W. C.,
T. P. Crowell,
D. M. Watts,
V. L. R. Barros,
H. Kruger,
F. Pinheiro, and C. J. Peters.
1991.
Demonstration of yellow fever and dengue antigens in formalin-fixed paraffin-embedded human liver by immunohistochemical analysis.
Am. J. Trop. Med. Hyg.
45:408-417.
|
| 10.
|
Halstead, S. B.
1988.
Pathogenesis of dengue: challenges to molecular biology.
Science
239:476-481[Abstract/Free Full Text].
|
| 11.
|
Hargreaves, R. G.,
N. J. Borthwick,
M. S. Montani,
E. Piccolella,
P. Carmichael,
R. I. Lechler,
A. N. Akbar, and G. Lombardi.
1997.
Dissociation of T cell anergy from apoptosis by blockade of Fas/Apo-1 (CD95) signaling.
J. Immunol.
158:3099-3107[Abstract].
|
| 12.
|
Hiramatsu, N.,
N. Hayashi,
K. Katayama,
K. Mochizuki,
Y. Kawanishi,
A. Kasahara,
H. Fusamoto, and T. Kamada.
1994.
Immunohistochemical detection of Fas antigen in liver tissues of patients with chronic hepatitis C.
Hepatology
19:1354-1359[Medline].
|
| 13.
|
Hober, D.,
L. Poli,
B. Roblin,
P. Gestas,
E. Chungue,
G. Granic,
P. Imbert,
J.-L. Pecarere,
R. Vergez-Pascal,
P. Wattre, and M. Maniez-Montreuil.
1993.
Serum levels of tumor necrosis factor- (TNF-), interleukin-6 (IL-6), and interleukin-1 (IL-1) in dengue-infected patients.
Am. J. Trop. Med. Hyg.
48:324-331.
|
| 14.
|
Horvath, C. J.,
T. J. Ferro,
G. Jesmok, and A. Malik.
1988.
Recombinant tumor necrosis factor increases pulmonary vascular permeability independent of neutrophils.
Proc. Natl. Acad. Sci. USA
85:9219-9223[Abstract/Free Full Text].
|
| 15.
|
Innis, B. L.,
K. S. A. Myint,
A. Nisalak,
K. G. Ishak,
S. Nimmannitya,
T. Laohapand,
S. Tanprasertsuk,
V. Pongritsakada, and U. Thisyakorn.
1990.
Acute liver failure is one important cause of fatal dengue infection (abstr.).
Southeast Asian J. Trop. Med. Public Health
21:695-696.
|
| 16.
|
Jacobson, S.,
J. R. Richert,
W. E. Biddison,
A. Satinsky,
R. J. Hartzmann, and H. F. McFarland.
1984.
Measles virus specific T4+ human cytotoxic T-cell clones are restricted by class II HLA antigens.
J. Immunol.
133:754-757[Abstract].
|
| 17.
|
Jassoy, C.,
T. Harrer,
T. Rosenthal,
B. A. Navia,
J. Worth,
P. R. Johnson, and B. D. Walker.
1993.
Human immunodeficiency virus type 1-specific cytotoxic T lymphocytes release gamma interferon, tumor necrosis factor alpha (TNF-), and TNF- when they encounter their target antigens.
J. Virol.
67:2844-2852[Abstract/Free Full Text].
|
| 18.
|
Ju, S. T.,
H. Cui,
D. J. Panka,
R. Ettinger, and A. Marshak-Rothstein.
1994.
Participation of target Fas protein in apoptosis pathway induced by CD4+ Th1 and CD8+ cytotoxic T cells.
Proc. Natl. Acad. Sci.
91:4185-4189[Abstract/Free Full Text].
|
| 19.
|
Kalayanarooj, S.,
D. W. Vaughn,
S. Nimmannitya,
S. Green,
S. Suntayakorn,
N. Kunentrasai,
W. Viramitrachai,
S. Ratanachu-eke,
S. Kiatpolpoj,
B. L. Innis,
A. L. Rothman,
A. Nisalak, and F. A. Ennis.
1997.
Early clinical and laboratory indicators of acute dengue illness.
J. Infect. Dis.
176:313-321[Medline].
|
| 20.
|
Katoka, T.,
N. Shinohara,
H. Takayama,
K. Takaku,
S. Kondo,
S. Yonehara, and K. Nagai.
1996.
Concanamycin A, a powerful tool for characterization and estimation of contribution of perforin- and Fas-based lytic pathways in cell-mediated cytotoxicity.
J. Immunol.
156:3678-3686[Abstract].
|
| 21.
|
Kerr, J. F.,
W. G. Cooksley,
J. Searle,
J. W. Halliday,
W. J. Halliday,
I. Holder,
I. Roberts,
W. Burnett, and L. W. Powell.
1979.
The nature of piecemeal necrosis in chronic active hepatitis.
Lancet
ii:827-828.
|
| 22.
|
Knowles, B. B.,
C. C. Howe, and D. P. Aden.
1980.
Human hepatocellular carcinoma cell lines secrete the major plasma proteins and hepatitis B surface antigen.
Science
209:479-499[Free Full Text].
|
| 23.
|
Kuno, G., and R. E. Bailey.
1994.
Cytokine responses to dengue infection among Puerto Rican patients.
Mem. Inst. Oswaldo Cruz
89:179-182[Medline].
|
| 24.
|
Kuo, C.,
D. Tai,
C. Chang-Chien,
C. Lan,
S. Chiou, and Y. Liaw.
1992.
Liver biochemical tests and dengue fever.
Am. J. Trop. Med. Hyg.
47:265-270.
|
| 25.
|
Kurane, I.,
B. L. Innis,
C. Nisalak,
C. Hoke,
S. Nimmanitya,
A. Meager, and F. A. Ennis.
1989.
Human responses to dengue virus antigens. Proliferative responses and interferon gamma production.
J. Clin. Invest.
83:506-513.
|
| 26.
|
Kurane, I.,
A. Meager, and F. A. Ennis.
1989.
Dengue virus-specific human T cell clones: serotype crossreactive proliferation, interferon- production, and cytotoxic activity.
J. Exp. Med.
170:763-775[Abstract/Free Full Text].
|
| 27.
|
Kurane, I.,
B. L. Innis,
S. Nimmannitya,
A. Nisalak,
A. Meager,
J. Janus, and F. A. Ennis.
1991.
Activation of T lymphocytes in dengue virus infections. High levels of soluble interleukin 2 receptor, soluble CD4, soluble CD8, interleukin 2, and interferon- in sera of children with dengue.
J. Clin. Invest.
88:1473-1480.
|
| 28.
|
Kurane, I., and F. A. Ennis.
1994.
Cytotoxic T lymphocytes in dengue virus infection.
Curr. Top. Microbiol. Immunol.
189:93-105[Medline].
|
| 29.
|
Lacronique, V.,
A. Mignon,
M. Fabre,
B. Viollet,
N. Rouquet,
T. Molina,
A. Porteu,
A. Henrion,
D. Bouscary,
P. Varlet,
V. Joulin, and A. Kahn.
1996.
Bcl-2 protects from lethal hepatic apoptosis induced by an anti-Fas antibody in mice.
Nat. Med.
2:80-86[Medline].
|
| 30.
|
Leithauser, F.,
J. Dhein,
G. Mechtersheimer,
K. Koretz,
S. Bruderlein,
C. Henne,
A. Schmidt,
K.-M. Debatin,
P. H. Krammer, and P. Moller.
1993.
Constitutive and induced expression of APO-1, a new member of the nerve growth factor/tumor necrosis factor receptor superfamily, in normal and neoplastic cells.
Lab. Invest.
69:415-429[Medline].
|
| 31.
|
Lewinsohn, D. M.,
T. T. Bemet,
J. Xu,
D. H. Lynch,
K. H. Grabstein,
S. G. Reed, and M. R. Alderson.
1998.
Human purified protein derivative-specific CD4+ T cells use both CD95-dependent and CD95-independent cytolytic mechanisms.
J. Immunol.
160:2374-2379[Abstract/Free Full Text].
|
| 32.
|
Miskovsky, E. P.,
A. Y. Liu,
W. Pavlat,
V. Renate,
P. E. Stanhope,
D. Finzi,
W. M. I. Fox,
R. H. Hruban,
E. R. Podack, and R. F. Siliciano.
1994.
Studies of the mechanism of cytolysis by HIV-1-specific CD4+ human CTL clones induced by candidate AIDS vaccines.
J. Immunol.
153:2787-2799[Abstract].
|
| 33.
|
Mori, M.,
I. Kurane,
J. Janus, and F. A. Ennis.
1997.
Cytokine production by dengue virus antigen-responsive human T lymphocytes in vitro examined using a double immunocytochemical technique.
J. Leukocyte Biol.
61:338-345[Abstract].
|
| 34.
|
Neal, Z. C., and G. A. Splitter.
1995.
Picornavirus-specific CD4+ T lymphocytes possessing cytolytic activity confer protection in the absence of prophylactic antibodies.
J. Virol.
69:4914-4923[Abstract].
|
| 35.
|
Nimmannitya, S.
1987.
Clinical spectrum and management of dengue haemorrhagic fever.
Southeast Asian J. Trop. Med. Public Health
18:392-397[Medline].
|
| 36.
|
Orentas, R. J.,
J. E. K. Hildreth,
E. Obah,
M. Polydefkis,
G. E. Smith,
M. L. Clements, and R. F. Siliciano.
1990.
Induction of CD4+ human cytolytic T cells specific for HIV-infected cells by a gp120 subunit vaccine.
Science
248:1234-1237[Abstract/Free Full Text].
|
| 37.
|
Ramsdell, F.,
M. S. Seaman,
R. E. Miller,
K. S. Picha,
M. K. Kennedy, and D. H. Lynch.
1994.
Differential ability of Th1 and Th2 T cells to undergo activation-induced cell death.
Int. Immunol.
6:1545-1553[Abstract/Free Full Text].
|
| 38.
|
Sabin, A. B.
1952.
Research on dengue during World War II.
Am. J. Trop. Med. Hyg.
1:30-50.
|
| 39.
|
Sherman, M. L.,
D. R. Spriggs,
K. A. Arthur,
K. Imamura,
E. I. Frei, and D. W. Kufe.
1988.
Recombinant human tumor necrosis factor administered as a five-day continuous infusion in cancer patients: phase I toxicity and effects on lipid metabolism.
J. Clin. Oncol.
6:344-350[Abstract].
|
| 40.
|
Smyth, M. J.
1997.
Fas Ligand-mediated bystander lysis of syngeneic cells in response to an allogeneic stimulus.
J. Immunol.
158:5765-5772[Abstract].
|
| 41.
|
Spriggs, D. R.,
M. L. Sherman,
H. Michie,
K. A. Arthur,
K. Imamura,
D. Wilmore,
E. I. Frei, and D. W. Kufe.
1988.
Recombinant human tumor necrosis factor administered as a 24-hour intravenous infusion. A phase I and pharmacologic study.
J. Nat. Cancer Inst.
80:1039-1044[Abstract/Free Full Text].
|
| 42.
|
Stalder, T.,
S. Hahn, and P. Erb.
1994.
Fas antigen is the major target molecule for CD4+ T cell-mediated cytotoxicity.
J. Immunol.
152:1127-1133[Abstract].
|
| 43.
|
Van Voorhis, W. C.,
L. K. Barrett,
J. M. Nasio,
F. A. Plummer, and S. A. Lukehart.
1996.
Lesions of primary and secondary syphilis contain activated cytolytic T cells.
Infect. Immun.
64:1048-1050[Abstract].
|
| 44.
|
Vergelli, M.,
B. Hemmer,
P. A. Muraro,
L. Tranquill,
W. E. Biddison,
A. Sarin,
H. F. McFarland, and R. Martin.
1997.
Human autoreactive CD4+ T cell clones use perforin- or Fas/Fas ligand-mediated pathways for target cell lysis.
J. Immunol.
158:2756-2761[Abstract].
|
| 45.
|
Vitarana, T.,
H. de Silva,
N. Withana, and C. Gunasekera.
1991.
Elevated tumour necrosis factor in dengue fever and dengue haemorrhagic fever.
Ceylon Med. J.
36:63-65[Medline].
|
| 46.
|
Wijburg, O. L. C.,
M. H. M. Heemskerk,
A. Sanders,
C. J. P. Boog, and N. Van Rooijen.
1996.
Role of virus-specific CD4+ cytotoxic T cells in recovery from mouse hepatitis virus infection.
Immunology
87:34-41[Medline].
|
| 47.
|
Woo, J.-T.,
C. Shinohara,
K. Sakai,
K. Hasumi, and A. Endo.
1992.
Isolation, characterization, and biological activities of concanamycins as inhibitors of lysosomal acidification.
J. Antibiot.
45:1108[Medline].
|
| 48.
|
Yadav, M.,
K. R. Kamath,
N. Iyngkaran, and M. Sinniah.
1991.
Dengue haemorrhagic fever and dengue shock syndrome: are they tumour necrosis factor-mediated disorders?
FEMS Microbiol. Immunol.
89:45-50.
|
| 49.
|
Yasukawa, M.,
A. Inatsuki, and Y. Kobayashi.
1989.
Differential in vitro activation of CD4+ CD8 and CD8+ CD4 herpes simplex virus-specific human cytotoxic T cells.
J. Immunol.
143:2051-2057[Abstract].
|
| 50.
|
Yasukawa, M.,
Y. Yakushijin, and S. Fujita.
1996.
Two distinct mechanisms of cytotoxicity mediated by herpes simplex virus-specific CD4+ human cytotoxic T cell clones.
Clin. Immunol. Immunopathol.
78:70-76[Medline].
|
Journal of Virology, May 1999, p. 3623-3629, Vol. 73, No. 5
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Copyright © 1999, American Society for Microbiology. All rights reserved.
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-
Herbein, G., O'brien, W. A.
(2000). Tumor Necrosis Factor (TNF)-{alpha} and TNF Receptors in Viral Pathogenesis. Exp. Biol. Med.
223: 241-257
[Abstract]
[Full Text]
-
Zivny, J., DeFronzo, M., Jarry, W., Jameson, J., Cruz, J., Ennis, F. A., Rothman, A. L.
(1999). Partial Agonist Effect Influences the CTL Response to a Heterologous Dengue Virus Serotype. J. Immunol.
163: 2754-2760
[Abstract]
[Full Text]
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Abstract
Introduction
