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Fujisaki Institute, Hayashibara Biochemical Laboratories, Inc., Okayama, Japan
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Abstract |
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 Top Abstract IntroductionMaterials and MethodsResultsDiscussionReferences |
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production by Con A-stimulated spleen cells. In contrast, IL-18
treatment partially but significantly restored the production of these
cytokines. Data further suggested that these IL-18-mediated therapeutic
effects may be due to the induction of donor CD8+ CTL, the
decrease in donor CD4+ T cell numbers, and a
down-regulation of host B cell MHC class II expression. Thus, our
results suggest that IL-18 has beneficial effects in the prevention and
treatment of chronic GVHD. |
Introduction |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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One of the principal factors that causes the distinction between chronic and acute GVHD is considered to be the generation of donor anti-host CTL function (5, 7, 8). Chronic GVHD mice show a 2-fold reduction in CD8+ T cells and a 9-fold reduction in anti-host CTL activity as compared with acute GVHD mice (4). The defect in generating anti-host CTL activity required for the elimination of autoreactive B cells which produce pathological autoantibodies leads to a systemic disorder (9, 10). These results suggest that moderate activation of donor CD8+ T cells would result in the prevention or attenuation of chronic GVHD development without causing excessive elimination of host cells and the severe immunodeficiency observed in acute GVHD.
Because it is well known that IL-12 potentiates the cytotoxic activity of CTLs (11, 12), experiments to treat mice with chronic GVHD with IL-12 have been performed by several laboratories (6, 13, 14, 15). None of these attempts using IL-12, however, were satisfactory as a therapeutic approach to chronic GVHD, in that IL-12-treated chronic GVHD mice exhibited several phenotypes associated with acute GVHD, including weight loss, mortality, increased MHC class II expression, and a profound immunodeficiency as shown by a severe defect in the IL-2 production by Con A-stimulated spleen cells (13, 14). Moreover, when IL-12 administration started after the onset of chronic GVHD, this failed to suppress autoimmune responses, including the production of anti-DNA autoantibodies (13). Thus, IL-12 is likely to be insufficient for the treatment of chronic GVHD.
IL-18 is a recently identified and cloned cytokine, which shares some
biological activities with those of IL-12 (15, 16). IL-18
acts as a costimulatory factor for Th1 clones stimulated with Ag on
APC, immobilized anti-CD3 mAb, or Con A to increase IFN-
production and proliferation of the Th1 clones (17, 18).
IL-18, unlike IL-12, does not drive Th1 development, but potentiates
IL-12-driven Th1 development (19). Recently, we have
demonstrated that IL-18 effectively induces the development of
DBA/2-derived CD8+ T cells into type I effector cells in
DBA/2 anti-BDF1 MLC (20).
In the present study, we evaluated the therapeutic and preventive effects of IL-18 administration to chronic GVHD model mice. Our results indicate that IL-18 administration at the time of chronic GVHD induction effectively prevents its onset mainly by the induction of donor anti-host CD8+ CTL without any signs of acute GVHD-like symptoms. Interestingly, IL-18 treatment after the onset of clinical signs of chronic GVHD also suppressed its progression, probably by reducing an allo-specific reaction between donor CD4+ T cells and host B cells.
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Materials and Methods |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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Female DBA/2 and BDF1 mice at 8–10 wk of age were obtained from Japan Charles River Laboratory (Yokohama, Japan) and Japan SLC (Shizuoka, Japan). Mice were housed under standard conditions for at least 1 wk before our experiments.
Induction of chronic GVHD
DBA/2 mouse spleen cells (9 x 107 viable cells) were transferred by i.v. injection into BDF1 mice as described previously (12). Normal BDF1 mice were injected with FCS-free RPMI 1640 medium as controls.
IL-18 treatment
Murine rIL-18 was generated by expression of IL-18 cDNA in Escherichia coli and was purified by several steps of column chromatography, as described previously (20). The endotoxin content was <1 ng/mg, as measured by the Limulus amebocyte lysate assay. Experimental mice received IL-18 i.p. at 0.1, 1, and 10 µg/0.1 ml PBS containing 0.1% mouse serum albumin on days 0–5 or days 8–13 after the induction of chronic GVHD. Control chronic GVHD mice received PBS containing 0.1% mouse serum albumin.
IL-12 treatment
Murine rIL-12 was expressed in Chinese hamster ovary (CHO) cells and was purified from the supernatants of the CHO cell cultures by several steps of column chromatography, as described previously (20). The specific activity was 5.5 x 106 U/mg. The endotoxin content was <30 ng/mg. IL-12 was injected i.p. into chronic GVHD mice at 0.1 µg/0.1 ml PBS containing 0.1% mouse serum albumin on days 8–13 after the induction of chronic GVHD.
Flow cytometry
Spleen cells from GVHD mice and normal mice were analyzed for the percentages of donor CD8+ T cells (H-2Kb-, CD8+), donor CD4+ T cells (H-2Kb-, CD4+), and host B cells (H-2Kb+, CD19+) using anti-CD4 mAb (GK1.5), anti-CD8 mAb (53-6.7), or anti-CD19 mAb (1D3) followed by FITC-conjugated F (ab')2 fragments of goat anti-rat IgG and PE-conjugated anti-H-2Kb mAb (AF6-88.5), all of which were obtained from PharMingen (San Diego, CA). Isotype-matched control Abs were used for background staining. Spleen cells were also assessed for the intensity of host B cell MHC class II expression using PE-conjugated anti-I-Ab mAb (AF6-120.1; PharMingen). Cells were washed three times in PBS supplemented with 1% FCS and 0.05% sodium azide after incubation with each mAb on ice for 30 min. After viable lymphocytes had been gated by forward and side scatter, stained cells were analyzed on an EPICS Profile II flow cytometer (Coulter Electronics, Hialeah, FL).
Induction and measurement of cytokine production in vitro
Spleen cells (5 x 106/well) from experimental
mice were cultured in 24-well plates with 5 µg/ml Con A in RPMI 1640
medium containing 10% FCS, 5 x 10-5 M 2-ME (Life
Technologies, Grand Island, NY), 60 µg/ml penicillin, and 50 µg/ml
streptomycin. Cells were cultured for 24 h or 48 h at 37°C
to assess the levels of IL-2 and IFN- production, respectively. The
cell-free supernatants were collected and frozen at -20°C until
being assayed. Sandwich ELISAs were used to determine IL-2 and IFN-
levels. The mAbs for plate coating and biotinylated secondary mAb were
as follows: for IL-2, rat anti-mouse IL-2 mAb (JES6-1A12;
PharMingen) and biotinylated rat anti-mouse IL-2 mAb (JES6-5H4;
PharMingen); for IFN-, rabbit anti-mouse IFN- polyclonal Ab
prepared in our laboratory and biotinylated rat anti-mouse IFN-
mAb (XMG1.2; PharMingen).
Measurement of serum Igs
Sera were collected from individual mice and serum levels of IgG1 and IgE were determined by sandwich ELISA. Briefly, purified rat anti-mouse IgG1 mAb (A85-3; PharMingen) and HRP-conjugated rat anti-mouse IgG1 (Zymed, San Francisco, CA) for IgG1, or rat anti-mouse IgE mAb (6HD5, Yamasa, Chiba, Japan) and biotinylated rat anti-mouse IgE mAb (HMK-12, Yamasa) for IgE, were used for plate coating and secondary Abs for ELISA. Mouse IgG1 (S1-68.1; PharMingen) and mouse IgE (IgE-3; PharMingen) were used as standards in these assays.
Serum titers of IgG antinuclear Abs were also assessed by ELISA as described previously (21). Briefly, microtiter plates were coated with calf thymus DNA (Sigma, St. Louis, MO) at 5 µg/ml in PBS, and were incubated with serially diluted serum. Plates were then developed by the addition of biotinylated rat anti-mouse IgG (Vector Laboratories, Burlingame, CA) followed by the addition of HRP-labeled streptavidin (Zymed).
51Cr release assay to test cytotoxic activity
Spleen cells from chronic GVHD mice treated with IL-18 or PBS were harvested on day 10 after chronic GVHD induction. The spleen cells (5 x 106/well) were restimulated with mitomycin C (Sigma)-treated BDF1 spleen cells (5 x 106/well) for 5 days. In some experiments, CD8+ T cells were depleted from the effector cells by panning using anti-CD8a mAb (53-6.7; PharMingen). The remaining CD8+ T cells among the effector cells after depletion were <0.8% as determined by FACS analysis. Effector cells were harvested and washed with medium. After centrifugation on Ficoll, various numbers of effector cells were cocultured with 5 x 103 51Cr-labeled target cells in 96-well round-bottomed plates for 4 h at 37°C. The percentage of specific lysis was calculated according to the following formula: percent specific lysis = [(cpm experimental - cpm spontaneous)/(cpm maximum - cpm spontaneous)] x 100.
Statistical analysis
All statistical analyses were performed using Student’s t test. The p values <0.05 were considered statistically significant.
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Results |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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To determine whether IL-18 administration inhibits the
development of chronic GVHD, chronic GVHD was induced by injecting
DBA/2 spleen cells into BDF1 mice. IL-18 was administered
in three doses (0.1, 1, and 10 µg/day) on days 0–5 after the
induction of chronic GVHD. On day 16 after the induction, control
PBS-treated mice exhibited the following findings typical of chronic
GVHD, as previously described (3, 13): splenomegaly; an
increase on host B cells; elevated serum Ab levels of IgG1, IgE, and
IgG antinuclear Abs; and an increase in the level of MHC class II
(I-Ab) expression on host B cells, a sign of host B cell
activation, as compared with those of normal BDF1 mice
(Fig. 1
and Table I). Although host macrophages also
expressed the I-Ab molecule, their percentages in the
spleen cells were <3% in all experiments and were not affected by the
IL-18 administration (data not shown). The expression intensity of the
I-Ab molecule was therefore regarded as reflecting the
activation levels of host splenic B cells and is used as an activation
marker of host B cells in the literature (3, 13). In
addition to the features of chronic GVHD described above, the IL-2 and
IFN- productions by Con A-stimulated spleen cells from control
chronic GVHD mice were reduced to 20% and 12% of those produced by
spleen cells from normal BDF1 mice, respectively (Table II).
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), and
a minimal increase in serum Igs levels (Fig. 1), although these changes
were not significant. However, as evaluated by flow cytometric analysis
shown in Table I, these IL-18-treated BDF1 mice did not
show any significant changes in the percentages and in the number of
CD4+ and CD8+ T cells, or of B cells in the
spleen when compared with PBS-treated BDF1 mice. Thus, it
is unlikely that IL-18 administration into normal BDF1 mice
seriously affects their immunological phenotypes.
IL-18 treatment for 6 days starting on the day of chronic GVHD
induction inhibited the serum titers of IgG1, IgE, and IgG antinuclear
Abs in a dose-dependent manner (Fig. 1) (1 µg/day, p
< 0.05; 10 µg/day, p < 0.01 vs PBS control group),
although the lowest dose of IL-18 (0.1 µg/day) had a minimal effect
that was not statistically significant (Fig. 1). IL-18 treatment (10
µg/day) almost completely suppressed the elevation of these serum Abs
to levels equal to those observed in normal BDF1 mice. In
parallel with these results, the IL-18 treatment significantly
decreased the number of host B cells, and the mean fluorescence
intensity of the MHC class II expression on host B cells (1 µg/day,
p < 0.05; 10 µg/day, p < 0.005 vs
PBS control group) (Table I). In another report, the appearance of the
acute GVHD phenotype was defined as a >50% reduction in host-derived
cells in the spleen (7). However, none of the
IL-18-treated mice displayed a severe decrease in the ratio of host
cells to donor cells (the percentages of host cells was 79.6 ±
11.4% of all spleen cells in mice receiving 10 µg/day of IL-18).
Additionally, neither weight loss nor mortality was observed in any of
the mice in the IL-18-treated groups during the 16 days (changes in
body weight were within 10% of the starting weight; data not shown),
and all five mice treated with IL-18 were still alive at 100 days (data
not shown). Importantly, IL-18 treatment dose-dependently partially
restored both IL-2 and IFN- production from spleen cells upon
stimulation with Con A (Table II) (p < 0.05).
This IL-18-induced restoration of IL-2 and IFN- does not seem to be
caused by a nonspecific immunomodulatory effect of IL-18, because the
augmentation of both IL-2 and IFN- production was not observed with
spleen cells from normal BDF1 mice injected with IL-18 on
days 0–5 (Table II). Furthermore, these findings contrasted with the
effect of IL-12 treatment as shown by Via et al. (13).
They reported that IL-12 treatment (0.1 µg/day) for 5 days starting
on the day of chronic GVHD induction further compromised the IL-2
production by Con A-stimulated spleen cells compared with that of
control chronic GVHD mice, indicating the development of an acute GVHD
phenotype. Taken together, these results suggest that the
administration of IL-18 effectively prevents the onset of chronic GVHD
without any features of acute GVHD.
IL-18 treatment of chronic GVHD mice changes the balance of donor CD4+ and CD8+ T cell numbers, and selectively decreases host B cells
Interestingly, 1 µg/day of IL-18 treatment for 6 days, starting
on the day of cell transfer, decreased the number of host B cells by
54% compared with those in the PBS control group, although this
decrease corresponds to only an 8% reduction in total spleen cells
(Table I). On the other hand, 10 µg/day of IL-18 treatment resulted
in a further 17% reduction in the number of host B cells and a 54%
reduction in the number of total spleen cells as compared with those in
the 1 µg/day of the IL-18 treatment group. These results suggest that
in the chronic GVHD mice treated with IL-18, host B cells are first
eliminated, followed by the elimination of other host-derived cells,
including host T cells depending on the dose of IL-18. To ascertain
this possibility, we examined the effect of IL-18 treatment (1
µg/day) on donor and host T cell numbers. As shown in Table III, donor CD4+ T cells were
reduced by 46% in the spleens of IL-18-treated mice, whereas the
number of donor CD8+ T cells increased 1.8-fold compared
with those in PBS-treated chronic GVHD mice. On the other hand, as we
expected, little or no change in the number of both CD4+
and CD8+ host T cells was observed in IL-18-treated mice,
whereas the number of host B cells was reduced to 25%. In contrast, in
the case of 10 µg/day of IL-18 treatment, both CD4+ and
CD8+ host T cells were also significantly reduced in number
(data not shown). These results suggest that host B cells were
selectively eliminated in the case of administration with 1 µg/day of
IL-18 in DBA/2 cell-injected BDF1 mice. These results
further suggest that IL-18 may activate donor CD8+ T cells
and induce the development of CD8+ CTL, resulting in the
elimination of activated host B cells.
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It was reported that delayed IL-12 treatment (days 8–12) of
chronic GVHD mice cannot decrease the serum anti-DNA Ab levels and
host B cell numbers (13). To address the question of
whether IL-18 treatment can suppress ongoing chronic GVHD, we started
the administration of IL-18 (1 or 10 µg/day) to chronic GVHD mice 8
days after induction. In kinetic studies, on day 8, DBA/2 cell-injected
BDF1 mice developed clinical signs of chronic GVHD
characterized by splenomegaly (1.2-fold increase in the total number of
spleen cells), together with an elevated serum IgG antinuclear Abs,
IgG1, and IgE (p < 0.05 vs normal
BDF1 mice) (data not shown). During the administration of
IL-18 (days 8–13), neither weight loss nor death was observed (data
not shown). All mice were studied 16 days after cell transfer. In the
IL-18-treated groups, the total number of spleen cells tended to
decrease, although there were no statistically significant differences
(Table IV). Host B cell numbers and their
MHC class II expression intensity were significantly decreased in the
IL-18-treated group by 28% and 23%, respectively, compared with the
PBS control group (10 µg/day, p < 0.05) (Table IV).
Donor CD4+ T cells decreased in number after delayed
treatment with IL-18 (10 µg/day) (69% reduction, p
< 0.05 vs PBS control group), although unlike in early IL-18 treatment
(days 0–5), donor CD8+ T cells in the spleen did not
increase. As shown already in Tables I and III, similar results were
also obtained when IL-18 (1 µg/day) was administered on days 0–5
after the chronic GVHD induction (MHC class II expression, 39%
reduction; donor CD4+ T cell number, 46% reduction). Thus,
regardless of the timing of the treatment, IL-18 significantly
decreased the number of donor CD4+ T cells and the level of
host B cell MHC class II expression.
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, delayed IL-18
treatment (days 8–13) significantly suppressed the serum IgG
antinuclear Ab levels as compared with those in the PBS control group
(p < 0.05). The IL-18 treatment also
significantly decreased the serum IgG1 levels (1 and 10 µg/day) and
the IgE levels (10 µg/day) compared with those in the PBS control
group (p < 0.05). To compare the effect of
IL-18 treatment on the serum IgG antinuclear Ab levels with that of
IL-12 treatment, chronic GVHD mice were administered 0.1 µg/day of
IL-12 according to the same schedule as that for IL-18 treatment (days
8–13). We confirmed that the delayed administration of IL-12 failed to
decrease serum IgG antinuclear Ab levels, although it suppressed serum
IgG1 and IgE levels, as previously reported (Fig. 3 and data not shown) (13).
Moreover, we observed that chronic GVHD mice injected with IL-12 (0.1
µg/day) exhibited 1.7-fold greater splenomegaly (3.1 ± 0.5
x 108 cells vs 5.3 ± 1.1 x 108
cells, p < 0.05) and tended to have increased host B
cell numbers compared with those in the PBS control group (no
significant difference; data not shown). We next assessed whether
delayed IL-18 treatment will also restore the impaired IL-2 and IFN-
production upon stimulation of the spleen cells with Con A. As shown in
Table V, both IL-2 and IFN- production
in the culture supernatants were significantly increased in the
IL-18-treated group (10 µg/day groups, p < 0.05). No
clear IL-18 dose response in these data may suggest that the effect of
IL-18 at a dose ranging from 1 to 10 µg/day has reached a plateau
level. Taken together, our results indicate that even after the onset
of chronic GVHD, IL-18 treatment significantly ameliorates the
progression of chronic GVHD.
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In the preceding sections, we showed that increased donor
CD8+ T cells and decreased host B cells were observed in
IL-18-treated chronic GVHD mice when compared with control PBS-treated
mice, especially when treatment started immediately after the chronic
GVHD induction. In addition, our previous in vitro study demonstrated
that IL-18 induces the development of DBA/2-derived naive
CD8+ T cells into type I effector cells in DBA/2
anti-BDF1 MLC (20). Therefore, we
investigated whether the in vivo preventive effect of IL-18 on the
development of chronic GVHD was due to the elimination of host B cells
by IL-18-induced CD8+ CTL. To examine the anti-host CTL
activity in chronic GVHD mice treated with IL-18 (1 µg/day, days
0–5), spleen cells harvested 10 days after induction were stimulated
with mitomycin C-treated BDF1 spleen cells for 5 days, and
then the cytotoxic function of the cultured cells was analyzed using
51Cr-labeled EL-4 (H-2b) and P815
(H-2d) as target cells. No lysis against donor-type P815
was observed in any group (<1% specific lysis). The spleen cells from
IL-18-treated chronic GVHD mice showed CTL activity specific for
host-type EL-4 cells (15% at E:T ratio of 50:1, Fig. 4), whereas a minimal CTL response was
detected in the PBS control group (3.3% at E:T ratio of 50:1), as
reported previously in similar experiments (4). This
result indicates that IL-18 treatment is able to elicit donor
anti-host CTL activity, suggesting that the IL-18-induced donor
anti-host CTL then eliminates activated host B cells.
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, depletion of CD8+ T cells from the effector cells using
anti-CD8 mAb completely abrogated the IL-18-induced CTL activity
(1.5% at E:T ratio of 50:1). These data show that the administration
of IL-18 into DBA/2 cell-injected BDF1 mice induces
anti-host CD8+ CTL. These results further suggest that
the activation of donor CD8+ T cells is important in the
prevention of chronic GVHD development.
We next examined the effect of IL-18 treatment after the onset of
immunological evidences of chronic GVHD on the induction of donor
anti-host CTL. However, we could not detect any cytotoxic activity
against EL-4 cells in the spleen cells from chronic GVHD mice that
received IL-18 (1 or 10 µg/day) on days 8–13 (<3% at an E:T ratio
of 100:1 in two separate experiments). This result correlated with the
observation that we could not see an increase in the number of donor
CD8+ T cells after delayed IL-18 treatment (Table V). These
results suggest that delayed IL-18 treatment reduces the number of host
B cells by a mechanism other than the development of donor
anti-host CTL.
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Discussion |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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No conversion into acute GVHD after IL-18 treatment may be ascribed to the reduction in the number of donor CD4+ T cells. Our present results show that DBA/2 cell-injected BDF1 mice treated with IL-18 exhibited reduced numbers of donor CD4+ T cells regardless of the timing of the administration. These results were in accordance with our previous in vitro study that CD4+ T cells decreased in DBA/2 anti-BDF1 MLC in the presence of IL-18, whereas CD8+ T cells increased (20). Recently, Buhlmann et al. (22) demonstrated that donor CD4+ T cells that produce IL-2 and express CD40 ligand are required not only for the induction, but also for the sustained expansion of donor effector CD8+ T cells in C57BL/6 cell-injected BDF1 mice with acute GVHD. In addition, donor CD4+ T cells seem to play a pivotal role in causing lethal acute GVHD (23). These findings suggest that the decrease in donor CD4+ T cells in DBA/2 cell-injected BDF1 mice treated with IL-18 may play a role as a negative feedback mechanism in the sustained expansion of donor CD8+ CTL, resulting in the prevention of conversion to acute GVHD.
Another possibility for the mechanism of preventing conversion into an
acute phenotype in IL-18-treated mice could be also considered. IFN-
induced by IL-18 administration to chronic GVHD mice may play a role in
blocking the conversion to acute GVHD. It has been postulated that
IFN- production would lead to exacerbation of acute GVHD
(24, 25, 26). However, IFN- knock-out mice used as donors
in a mouse acute GVHD model accelerated acute GVHD-associated morbidity
(27). Similarly, a recent study demonstrated that the
protection of acute GVHD by IL-12 treatment was dependent on
donor-derived IFN- production (28). We could observe
high levels of IFN- in the serum of chronic GVHD mice treated with
10 µg/day of IL-18 on days 0–5 (sera were obtained from a pool of
three mice) (<0.5, 44.9, 70.5, and 1.9 IU/ml on days 2, 4, 5, and 7,
respectively), whereas IFN- in the serum from control PBS-treated
chronic GVHD mice was undetected. Therefore, it is probable that
IFN- serves to protect from conversion into the acute phenotype in
IL-18-treated chronic GVHD mice.
Our present study is the first demonstration that IL-18 treatment even
after the onset of clinical symptoms (days 8–13) ameliorates the
immunological findings associated with chronic GVHD. However, this
efficacy of the delayed IL-18 treatment was attenuated compared with
that of early IL-18 treatment during the first week (days 0–5). The
attenuated effect on chronic GVHD was similarly observed in delayed
treatment with IL-12 (13). It has been reported that the
progression of chronic GVHD during the first 2 wk is associated with
skewing donor T cells toward a Th2 phenotype (3, 29, 30).
Therefore, the diminished effect of delayed treatment may be, in part,
accounted for by a decrease in the responsiveness of T cells to IL-18
or IL-12 in chronic GVHD mice during the first 2 wk, because expression
of the IL-18 receptor is known to be down-regulated in Th2 cells
(31, 32). In fact, IFN- production by spleen cells from
chronic GVHD mice (n = 3) on day 8 in response to 10
ng/ml of IL-18 was reduced to about one-third of that produced by
spleen cells on day 2 (day 2,378 ± 45 IU/ml; day 8, 132 ±
15 IU/ml).
The preventive and therapeutic effects of IL-18 on chronic GVHD may be
explained by two additional findings, besides the induction of donor
anti-host CD8+ CTL as important effector cells for
eliminating host B cells. First, chronic GVHD mice treated with IL-18
suppressed the level of MHC class II (I-Ab) expression,
thus leading to a reduction in Ag presentation to donor
CD4+ T cells. In contrast, IL-12 treatment up-regulated
this expression, as shown by Via et al. (13). Recently, it
has been reported that IFN- down-regulates MHC class II expression
on B cells, whereas it up-regulates its expression on macrophages
(33). Furthermore, as described above, we could detect
significant amounts of IFN- in the serum of chronic GVHD mice
treated with IL-18 on days 0–5. Thus, IFN- induced by IL-18 may
suppress host B cell MHC class II expression. However, we could detect
little IFN- production in the sera of chronic GVHD mice after
delayed treatment with IL-18 (data not shown), whereas we could observe
down-regulation of MHC class II expression on B cells by this
treatment. Thus, because we were not able to thoroughly rule out the
possibility that IFN- induced by IL-18 treatment suppressed MHC
class II expression, another mechanism may also be responsible for the
IL-18-mediated suppression of MHC class II expression. Second, IL-18
administration suppressed the engraftment of donor CD4+ T
cells by as yet unknown mechanisms. A recent study suggests that the
onset of chronic GVHD is caused by the immune response of donor
CD4+ T cells which recognize host alloantigen on host APC,
and then the donor CD4+ T cells help host B cells to
produce autoantibody (1, 4). Moreover, Via et al.
(34) have shown that blockade of CD28/CTLA-4: B7
interaction by CTLA4-Ig in ongoing chronic GVHD mice prevents the
development of Th2-deriven responses, along with down-regulation of B
cell MHC class II expression and reduced numbers of donor
CD4+ T cells. In their studies, they also postulated that
systemic autoimmune disease, such as chronic GVHD, requires continuous
CD4+ T cell help for B cells. Thus, it is possible that the
decrease in donor CD4+ T cell numbers and in host B cell
MHC class II expression resulted in the suppression of
alloantigen-specific immune responses, such as Ab production by host B
cells. Therefore, a reduced interaction between donor CD4+
T cells and host B cells by altering the surface expression of MHC
class II or costimulatory molecules may be a beneficial approach for
the treatment of chronic GVHD in mice.
It is well recognized that human SLE patients manifest immunological features such as autoreactive B cell activation and autoantibodies production that are similar to those observed in the chronic GVHD mice used in the present study (1, 4, 10). Furthermore, it has been shown that both in human SLE and in chronic GVHD mice, CD4+ T cell-B cell interaction is required for producing autoantibodies by B cells (10, 34, 35). These results together with the findings obtained from our present study suggest that IL-18 may have beneficial effects on the treatment of human SLE.
It has been shown that Th1-mediated immune responses are dominated in acute GVHD mice (3, 5). In addition, anti-host CTL effectors play an important role in mediating acute GVHD (5, 7). Furthermore, we have shown that Th1 response-related cytokine IL-18 induces CD8+ CTL development in in vitro (20) and the present in vivo study. These results suggest that in acute GVHD model mice IL-18 may be involved in its pathogenesis through the induction of CTL development. To clarify this possibility, further studies with acute GVHD mice need to be performed.
In summary, the effects of IL-18 on the development of chronic GVHD may be ascribed to the following IL-18-mediated triple action; the induction of donor anti-host CD8+ CTL, the down-regulation of host B cell MHC class II expression, and the decrease in donor CD4+ T cell numbers. To ascertain the mechanism for these IL-18-mediated effects, further experiments are necessary. Our findings presented in this study would provide new strategies using IL-18 for the treatment of chronic GVHD in mice and human autoimmune diseases, such as SLE.
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Acknowledgments |
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Footnotes |
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2 Abbreviations used in this paper: GVHD, graft-versus-host disease; BDF1, (C57BL/6 x DBA/2)F1; SLE, systemic lupus erythematosus.
Received for publication December 20, 1999. Accepted for publication March 20, 2000.
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  G. Socie and B. R. Blazar Acute graft-versus-host disease: from the bench to the bedside Blood, November 12, 2009; 114(20): 4327 - 4336. [Abstract] [Full Text] [PDF]
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C.-K. Min, Y. Maeda, K. Lowler, C. Liu, S. Clouthier, D. Lofthus, E. Weisiger, J. L. M. Ferrara, and P. Reddy Paradoxical effects of interleukin-18 on the severity of acute graft-versus-host disease mediated by CD4+ and CD8+ T-cell subsets after experimental allogeneic bone marrow transplantation Blood, November 15, 2004; 104(10): 3393 - 3399. [Abstract] [Full Text] [PDF]
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 T. Iwasaki Recent Advances in the Treatment of Graft-Versus-Host Disease Clin. Med. Res., November 1, 2004; 2(4): 243 - 252. [Abstract] [Full Text] [PDF]
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 W. Leung, R. Iyengar, V. Turner, P. Lang, P. Bader, P. Conn, D. Niethammer, and R. Handgretinger Determinants of Antileukemia Effects of Allogeneic NK Cells J. Immunol., January 1, 2004; 172(1): 644 - 650. [Abstract] [Full Text] [PDF]
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P. Reddy, T. Teshima, G. Hildebrandt, D. L. Williams, C. Liu, K. R. Cooke, and J. L.M. Ferrara Pretreatment of donors with interleukin-18 attenuates acute graft-versus-host disease via STAT6 and preserves graft-versus-leukemia effects Blood, April 1, 2003; 101(7): 2877 - 2885. [Abstract] [Full Text] [PDF]
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 D. Arnold, C. Wasem, P. Juillard, P. Graber, I. Cima, C. Frutschi, S. Herren, S. Jakob, S. Alouani, C. Mueller, et al. IL-18-independent cytotoxic T lymphocyte activation and IFN-{gamma} production during experimental acute graft-versus-host disease Int. Immunol., May 1, 2002; 14(5): 503 - 511. [Abstract] [Full Text] [PDF]
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 J. R. Wingard, G. B. Vogelsang, and H. J. Deeg Stem Cell Transplantation: Supportive Care and Long-Term Complications Hematology, January 1, 2002; 2002(1): 422 - 444. [Abstract] [Full Text]
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 P. Reddy, T. Teshima, M. Kukuruga, R. Ordemann, C. Liu, K. Lowler, and J. L.M. Ferrara Interleukin-18 Regulates Acute Graft-Versus-Host Disease by Enhancing Fas-mediated Donor T Cell Apoptosis J. Exp. Med., November 12, 2001; 194(10): 1433 - 1440. [Abstract] [Full Text] [PDF]
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D. H. Kono, M. S. Park, A. Szydlik, K. M. Haraldsson, J. D. Kuan, D. L. Pearson, P. Hultman, and K. M. Pollard Resistance to Xenobiotic-Induced Autoimmunity Maps to Chromosome 1 J. Immunol., August 15, 2001; 167(4): 2396 - 2403. [Abstract] [Full Text] [PDF]
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