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 Previous Article | Table of Contents | Next Article 
Blood, Vol. 93 No. 1 (January 1), 1999:
pp. 43-50
Murine T Lymphocytes Incapable of Producing Macrophage Inhibitory
Protein-1 Are Impaired in Causing Graft-Versus-Host Disease Across a
Class I But Not Class II Major Histocompatibility Complex
Barrier
By
Jonathan S. Serody,
Donald N. Cook,
Suzanne L. Kirby,
Elizabeth Reap,
Thomas C. Shea, and
Jeffrey A. Frelinger
From the Lineberger Comprehensive Cancer Center, and the Departments
of Medicine, Microbiology, and Immunology, University of North Carolina
at Chapel Hill, Chapel Hill, NC; and the Schering Plough Research
Institute, Kenilworth, NJ.
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ABSTRACT |
The routine use of bone marrow transplantation is limited by the
occurrence of acute and chronic graft-versus-host disease (GVHD).
Current approaches to decreasing the occurrence of GVHD after
allogeneic transplantation use T-cell depletion, use immunosuppressive agents, or block costimulatory molecule function. The role of proteins
in the recruitment of alloreactive lymphocytes has not been well
characterized. Chemokines are a large family of proteins that mediate
recruitment of mononuclear cells in vitro and in vivo. To investigate
the role of T-cell production of the chemokine macrophage inhibitory
protein-1 (MIP-1) in the occurrence of GVHD, splenocytes either
from wild-type or from MIP-1 / mice were administered to class
I (B6.C-H2bm1) and class II disparate mice
(B6-C-H2bm12). The incidence and severity of GVHD was
markedly reduced in bm1 mice receiving splenocytes from MIP-1/
mice as compared with mice receiving wild-type splenocytes. Bm1 mice
receiving MIP-1/ splenocytes had significantly less weight
loss and markedly reduced inflammatory responses in the lung and liver
than mice receiving C57BL/6 splenocytes. Bm1 mice receiving
MIP-1/ splenocytes had a markedly decreased production of
antichromatin autoantibodies and impaired generation of bm1-specific T
lymphocytes versus wild-type mice. However, MIP-1/ splenocytes
easily induced GVHD when administered to bm12 mice. This data show that
blockade of chemokine production or function may provide a new approach
to the prevention or treatment of GVHD but that chemokines that recruit
both CD4+ and CD8+ lymphocytes may need to
be targeted.
© 1999 by The American Society of Hematology.
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INTRODUCTION |
ALLOGENEIC BONE marrow transplantation
(ABMT) has been increasingly used in the treatment of individuals with
acute and chronic leukemia,1-3 lymphoma,4-10
and congenital blood disorders.11 However, the widespread
application of this therapy is currently limited by the occurrence of
acute graft-versus-host disease (aGVHD) and chronic GVHD
(cGVHD).12-16 Because of this, patients more than 55 years
of age and individuals that do not have a family or unrelated six-antigen-matched donor are infrequently offered this treatment. As
a result, only 25% to 35% of patients that could benefit from this
therapy are eligible for ABMT.
GVHD is due to the recognition by alloreactive T lymphocytes of minor
or major major histocompatibility complex (MHC) antigens.12 In the context of an MHC-identical six-antigen sibling transplant, the
antigens recognized are peptides from polymorphic proteins complexed to
class I or II MHC molecules.17 T lymphocytes interact with
MHC proteins on antigen-presenting cells through the T-cell receptor
complex. This interaction can lead to the death of the antigen-presenting cell by the elaboration of proteins such as perforin
and granzymes or to the production of proinflammatory cytokines such as
tumor necrosis factor- (TNF-).18 In animal models,
both of these effector mechanisms are important in the pathology of
GVHD.19-21 Current methods to decrease the occurrence of
GVHD after allogeneic BMT include the use cyclosporine or tacrolimus, which inhibit the transcription of genes that occur after T-cell signaling. Alternatively, cytotoxic compounds such as methotrexate have
been used to kill cells undergoing DNA synthesis.
Newer approaches to the prevention of GVHD have included the blockade
of the costimulatory molecules CD80 and CD8612,22 and the
blockade of interaction of CD4 with the 2 domain of the class II MHC
protein.12 However, none of these methods has been validated in a large clinical trial, and additional approaches are
needed to mitigate both the occurrence of GVHD as well as treatment of
established GVHD. One perplexing aspect regarding the occurrence of
GVHD is that, despite the presence of class I MHC molecules on all
nonneuronal cells, GVHD typically involves specific target organs. In
the setting of an MHC-matched sibling allograft, these differences may
represent differences in the tissue distribution of minor MHC antigens.
However, in the setting of an MHC-disparate transplant, other
mechanisms must be involved in the occurrence of GVHD in specific
target organs. Thus, there may be specific mechanisms involved in
either the presentation of peptides by MHC molecules in certain tissue
sites or in the recruitment of alloreactive T lymphocytes to the skin,
liver, and gastrointestinal tract.
Chemokines are a group of predominantly small molecular weight proteins
that are involved in the recruitment of multiple effector cells
including mononuclear leukocytes and neutrophils.23-29 The C-C chemokine, macrophage inhibitory protein-1 (MIP-1), has been
shown to recruit multiple different cell types, including activated
CD8+ lymphocytes in vitro.30 If GVHD involves
the specific recruitment of activated lymphocytes to target organs,
chemokines may play a role in this process. Additionally, previous
investigators have shown that chemokines effect proliferation and
cytotoxicity in vitro of T lymphocytes and natural killer
(NK) cells. Therefore, we were interested in studying the
role of specific chemokines in the pathophysiology of GVHD.
Using gene targeting, we produced mice with a disrupted gene encoding
MIP-1.31 These mice have a decreased inflammatory response after infection with influenza and Coxsackie viruses. More
importantly, we have shown that CD8+ lymphocytes from these
mice are not effective in the adoptive transfer of protection against
Listeria monocytogenes because of both impaired killing and
recruitment by these lymphocytes to the site of
infection.31a
In the present studies, we have explored the role of MIP-1
production by T lymphocytes in the occurrence of GVHD across both a
class I and class II MHC barrier. Splenocytes from
MIP-1/ were impaired in the ability to cause GVHD
across a class I but not class II MHC barrier. Blockade of T-lymphocyte
production of specific different chemokines may provide a new approach
to the prevention or treatment of GVHD.
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MATERIALS AND METHODS |
Mice.
B6.C-H2bm1/By (bm1), B6.C.H-2bm12/KhEg (bm12),
DBA/2J (H-2d), and C57BL/6 (H-2b) female
mice were purchased from Jackson Laboratories (Bar Harbor, ME). B6-129-Scya3tm1unc
(MIP-1/) mice were backcrossed to the eighth
generation onto a C57BL/6 background (MIP-1/ B6). All
mice were maintained specific-pathogen free in our colony under
microisolator cage tops. Mice received food ad libitum. After
irradiation, mice received neomycin-treated, acidified water. Female
mice between 7 and 12 weeks of age were used.
Splenocyte transfer.
Recipient bm1 or bm12 mice received either 500 or 600 cGy of
irradiation from a 137cesium source (Atomic Energy of
Canada, Ottawa, Ontario, Canada) in the evening before the splenocyte
transfer. The following morning, donor mice were killed by cervical
transection and single-cell suspensions of splenocytes were prepared.
Splenocytes were depleted of red blood cells using ACK
lysis buffer and resuspended in nonbacteriostatic saline. Recipient
mice received either 1 × 107 splenocytes or 5 × 106 CD8+ lymphocytes that had been selected
using anti-CD8 monoclonal antibody (Ly-2; Miltenyi Biotec Bergishch,
Gladbach, Germany) coupled to magnetic beads using the manufacturer's
protocol from either C57BL/6 or MIP-1/ B6 mice. After
immunomagnetic separation, 98% of the cells infused expressed CD8 as
shown by flow cytometry (data not shown). All mice were observed either
until death or day 250. Moribund mice were killed and the data were
censored from that point.
GVHD grading.
Mice were graded for the presence of skin and hair changes, hunched
posture, diarrhea, and weight loss. Mice had to have skin lesions and
hair loss consistent with GVHD and then either diarrhea or weight loss
to be considered to have GVHD. Weight loss greater than 10% of initial
weight was considered significant. Mice were weighed weekly for 50 days
and then every 28 days until day 250.
Histology.
Mice were killed at day 150 after splenocyte transfer and tissues were
placed in 10% neutral buffered formalin. Tissues were sectioned with a
microtome and stained with hematoxylin and eosin. Tissues were
evaluated by one of us (S.L.K.) blinded to the treatment administered.
Autoantibody production.
Surviving mice were bled at day 100 from the lateral tail vein. Blood
was diluted 1:20 in 4 mol/L citric acid and tested for IgG to chromatin
as previously reported.32 Control animals were irradiated
C57BL/6 mice that received splenocytes from C57BL/6 mice.
Detection of antihost cytotoxic T-cell activity in vitro.
At day 35 after transfer, spleen cells were isolated from bm1 recipient
mice and incubated with irradiated (2,500 cGy) splenocytes isolated
from either bm1, DBA/2J, or C57BL/6 mice in the presence of 25%
concavalin A supernatant. Four days later, effector cells were harvested and tested for the ability to lyse con A blasts from
bm1, C57BL/6 (H-2b), or DBA/2J (H-2d) mice in a
standard 4-hour 51Chromium release assay. Target to
effector ratios of 2.5:1 and 25:1 were used. The percentage of lysis
was calculated using the following formula: ([cpm sample cpm
spontaneous]/[cpm total - cpm spontaneous]) × 100. Spontaneous release was measured in wells that contained only target
cells. Total release was measured by the addition of Triton X-100 to a
final concentration of 5% to target cells. All samples were run in
triplicate. Results are shown for an E:T ratio of 25:1.
Engraftment in an MHC-mismatched setting.
To demonstrate that splenocytes from MIP-1/ B6
(H-2b) mice were capable of engrafting in completely
mismatched DBA/2J mice (H-2d), splenocytes were isolated
from MIP-1/ B6 mice and 1 × 107 were
administered intravenously to DBA/2J mice that had received 500 cGy of
irradiation the previous evening. Splenocytes from DBA/2J mice isolated
at day 7 were analyzed by flow cytometry for the expression of
H-2Kb (transferred population) and H-2Kd
(endogenous population). H-2Kb and H-2Kd
antibodies were purchased from Pharmingen (LaJolla, CA). Mice were
killed at day 14 and splenocytes were isolated and
stimulated for 3 days with irradiated splenocytes from either C57BL/6
or DBA/2J mice. After this, T cells were tested for the ability to lyse
con A blasts from either DBA/2J or MIP-1/ B6 mice in a standard 51Chromium-release assay.
Statistical analysis.
Groups were compared for the occurrence of GVHD using Fisher's exact
test. Groups were compared for differences in the production of
autoantibodies and weight loss using the Student's t-test. Estimates of the probability of survival for all groups were determined using the method of Kaplan and Meier.33 P values
less than .05 were considered significant.
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RESULTS |
Incidence of GVHD.
To examine the role of MIP-1 produced by T lymphocytes in the
occurrence of GVHD caused by a single class I MHC difference, we
transferred spleen cells from either B6 or MIP-1/ B6
mice to bm1 recipients. Previously, we have shown that
MIP-1/ mice have similar numbers of total white blood
cells and red blood cells in the bloodstream and similar numbers of CD4
and CD8 cells in lymph nodes and spleen as compared with control
animals.31 Therefore, the splenocyte transfers should
contain similar number of CD4+ and
CD8+lymphocytes regardless of donor used. The results are
shown in Table 1. For recipients receiving
500 cGy of irradiation before the splenocyte transfer, there was a
significant difference in the incidence of GVHD at days 150 (P = .04) and a trend toward a decreased incidence in GVHD at day 100 (P = .054) in mice that received MIP-1/ B6
splenocytes (incidence of GVHD, 40%) as compared with those that
received C57BL/6 splenocytes (incidence of GVHD, 75%).
We confirmed these findings using survival as the end-point for
analysis. bm1 mice receiving splenocytes from
MIP-1/ B6 mice had a statistically increased survival
(median survival, >250 v 105 days) over the 250 days after
the transfer compared with those that received splenocytes from C57BL/6
mice (P = .02; Fig 1). We performed
this same experiment using CD8+-selected splenocytes
transferred into bm1 recipients who had received 600 cGy of
irradiation. bm1 mice in this model die early of aGVHD when
CD8+ lymphocytes from C57BL/6 animals were
administered.34 As shown in Fig
2, we saw a significant difference in mortality due to aGVHD in favor
of bm1 recipients of MIP-1/ B6 splenocytes (median survival, 36 days) as compared with C57BL/6 splenocytes (median survival, 16.5 days; P = .04).
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| Fig 1.
Survival of 10 bm1 mice that received splenocytes from
MIP-1/ or C57BL/6 mice. Bm1 mice were irradiated the day
before transfer of 1 × 107 splenocytes depleted of
red blood cells. Mice were observed until day 250 or death. This is one
of three experiments (the pooled number of mice used was 30 for each
condition).
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| Fig 2.
Survival of bm1 mice that received CD8+
lymphocytes from MIP-1/ or C57BL/6 mice. Bm1 mice received 600 cGy irradiation on the evening before splenocyte
transfer. Splenocytes were prepared as indicated in the text. All mice
were observed until day 100 or death. Sixteen mice were treated in each
group.
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When similar experiments were performed using bm12 mice as
the recipient, no difference in outcome was observed using splenocytes from either MIP-1/ mice or C57BL/6 mice
(Fig 3; median survival, 125 v 145 days). Interestingly, the shape of the curves were different, with most
of the deaths occurring early in bm12 mice receiving MIP-1/ B6 splenocytes, suggesting an enhanced effect
on aGVHD in the setting of a class II MHC mismatch. Therefore,
splenocytes from MIP-1/ B6 mice were impaired in the
ability to cause GVHD across a class I but not class II MHC barrier.
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| Fig 3.
Survival of 10 bm12 mice that received splenocytes from
MIP-1/ or C57BL/6 mice. Bm12 mice were irradiated as described
in Fig 1. Splenocytes were prepared as indicated in the text and mice
were observed until day 250 or death.
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Histology and weight loss.
Tissue sections from bm1 mice that received either
splenocytes from C57BL/6 or MIP-1/ B6 mice were
evaluated for the presence of inflammatory foci in the lungs, liver,
spleen, and gastrointestinal tract. We found a decreased inflammatory
response in the liver and lungs of bm1 mice that received
MIP-1/ B6 splenocytes as compared with C57BL/6
splenocytes (Fig 4a, A and B v D
and E, and Fig 4b, B). Less striking was the decrease in the
inflammatory response in the gastrointestinal tract of bm1 mice
receiving MIP-1/ B6 splenocytes compared with C57BL/6
splenocytes (Fig 4b, A). The inflammatory infiltrate was made up
primarily of mononuclear leukocytes (Fig 4b, C). Furthermore, bm1 mice
that received MIP-1/ B6 splenocytes had significantly
less weight loss at day 75 (P = .04) and at day 150 (P = .007) compared with mice that received splenocytes from C57BL/6 mice
(Fig 5).
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| Fig 4.
(a and b) Histopathology from bm1 mice after transfer of
C57BL/6 or MIP-1/ B6 splenocytes. Bm1 mice that received
splenocytes from C57BL/6 or MIP-1/ B6 were killed 150 days
after transfer. Tissues were prepared as indicated in the text. (a)
shows (A and B) sections of liver from bm1 mouse that received
MIP-1/ B6 splenocytes compared with sections from an animal
that received C57BL/6 splenocytes (D and E). Sections (A and B) and (D
and E) represent two different magnifications of the same liver zone.
(a, C) is a section of the lung of a bm1 mouse that received
MIP-1/ B6 splenocytes compared with (F) and (b, B) sections of
lung from a bm1 mouse that received C57BL/6 splenocytes (figures are of
the same section at different magnifications). Foci of inflammation are
indicated by the arrows. The arrowheads indicate extravasation of
material into the lungs of bm1 mice that received C57BL/6 splenocytes.
(b, A) shows a crypt abscess in the gastrointestinal tract of a bm1
animal that received C57BL/6 splenocytes; these were not seen in
animals that received MIP-1/ splenocytes. The character of the
mononuclear cell infiltrate in the liver is shown in (b, C).
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| Fig 5.
Weight loss in bm1 recipients of C57BL/6 or
MIP-1/ B6 splenocytes. After splenocyte transfer, bm1 mice
were weighed weekly for the first 7 weeks and then monthly until day
250. After death, the weight just before death of each mouse was
carried until day 250. N = 12 mice in each group.
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Autoantibody production.
cGVHD in mice is characterized by the Th2 cytokine-driven production of
autoantibodies.22 Because our previous experiments had
suggested greater differences later after the transfer of splenocytes,
we wanted to test for differences in the production of autoantibodies
in this model. As demonstrated in Fig 6,
there was a striking decrease in the production of serum antichromatin antibodies in mice that received MIP-1/ B6 splenocytes
as compared with C57BL/6 splenocytes (P < .001). Mice that
received splenocytes from MIP-1/ mice had similar
production of autoantibodies compared with control mice in which
C57BL/6 splenocytes were administered to C57BL/6 mice. Thus,
autoantibody production was increased even in the setting of only a
class I MHC difference.
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| Fig 6.
Autoantibody production after splenocyte transfer. Bm1
mice were transferred splenocytes from either C57BL/6 or
MIP-1/ B6 mice. Mice were bled from the lateral tail vein and
50 µL of blood was diluted in 1 mL of 4 mol/L citric acid. The serum
was collected by centrifugation of the sample at 13,000g and
frozen at 70°C. The serum was tested for the presence of IgG
antichromatin autoantibodies using an enzyme-linked immunosorbent assay
(ELISA). Control mice were C57BL/6 mice that received splenocytes from
C57BL/6 mice.
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Generation of host-specific cytotoxic T lymphocytes
(CTL).
We have previously shown that pathogen-specific T lymphocytes are not
recruited to the site of infection if they do not produce MIP-1
after adoptive transfer.31a We tested for the presence of
donor-specific T lymphocytes in the spleen of bm1 mice after transfer
of either wild-type or MIP-1/ B6 splenocytes. After
the transfer of wild-type splenocytes, anti-host CTL activity was
readily apparent (Fig 7). In contrast,
there was no activity against host after the transfer of
MIP-1/ B6 splenocytes. To evaluate if donor
splenocytes were still present in the spleen, we tested splenocytes
from bm1 recipients of either MIP-1/ or C57BL/6 for
the ability to recognize third-party target cells. Bm1 recipients of
both C57BL/6 and MIP-1/ splenocytes were able to
recognize and kill third-party con A blasts from DBA/2J mice. There was
no lytic activity directed against C57BL/6 con A blasts, showing that
the lytic activity was not due to expansion of endogenous bm1
lymphocytes reacting with donor splenocytes. Thus,
MIP-1/ splenocytes are capable of initiating a
response against host in a completely mismatched setting. However, in
the setting of differences in the Kb allele, splenocytes
from MIP-1/ B6 mice do not cause a detectable antihost
response.
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| Fig 7.
Lytic activity of splenocytes isolated from bm1 mice
against blast cells from bm1, B6, or DBA/2J mice. Bm1 mice were killed
35 days after splenocyte transfer of either C57BL/6 ( ) or
MIP-1/ ( ) mice. Splenocytes were stimulated
in vitro as indicated and tested for lytic activity in a conventional
51Cr-release assay. This is one of three representative
experiments.
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Engraftment studies.
We evaluated the ability of MIP-1/ T cells to engraft
in a completely MHC-mismatched setting. We transferred 1 × 107 MIP-1/ B6 splenocytes into completely
mismatched irradiated (500 cGy) DBA/2J mice. We performed flow
cytometry on splenocytes from DBA/2J mice 7 days after splenocyte
transfer. Two percent to 4% of the splenocytes expressed the
transferred Kb protein. Thus, of 1 × 107
splenocytes transferred, 20% to 30% of these cells were either still
present or had expanded to this number at day 7 after transfer in the
spleen alone (data not shown). We also evaluated whether DBA/2J-specific lymphocytes were present in the spleen at day 14 after
transfer of splenocytes from C57BL/6 or MIP-1/ B6 mice. As shown in Fig 8, we were able to
show DBA/2J-specific T-cell activity after transfer of
MIP-1/ B6 splenocytes. Thus, because con A blasts do
not express detectable class II MHC complexes, we have shown that class
I-reactive MIP-1/ B6 lymphocytes can engraft using
sublethal irradiation in a completely MHC-mismatched setting.
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| Fig 8.
Lytic activity from splenocytes of DBA/2J mice after
transfer of MIP-1/ B6 splenocytes. DBA/2J mice were irradiated
as indicated in the text and the following day received 1 × 107 splenocytes from MIP-1/ B6 or C57BL/6 mice.
Fourteen days later, mice were killed, and their splenocytes were
isolated and stimulated for 48 hours with irradiated (2,500 cGy)
C57BL/6 or DBA/2J splenocytes. Lymphocytes were tested for lytic
activity on con A blasts from C57BL/6 and DBA/2J mice as indicated in
the text. Spontaneous release for each target cell was less than 20%
of maximal. This is one of two experiments.
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DISCUSSION |
GVHD is the limiting factor in the applicability of mismatched and
unrelated allogeneic BMT for the treatment of tumors and congenital
diseases. Current prevention and treatment of GVHD is not adequate to
allow the majority of patients to receive this form of therapy. In the
present study, we have investigated the role of T-cell production of
the chemokine MIP-1 in the occurrence of GVHD. In our model system,
MIP-1/ B6 splenocytes were markedly impaired in the
ability to cause aGVHD and cGVHD across a class I but not class II MHC
barrier. This was confirmed by demonstrating differences in overall
survival, histopathological changes in the liver and lungs, weight
loss, production of autoantibodies, and antihost CTL reactivity when
comparing bm1 recipients receiving wild-type or MIP-1/
splenocytes.
The present work was undertaken to investigate if alloreactive
lymphocytes are actively recruited out of the vasculature in GVHD and
if the production of chemokines impacted this recruitment. Because of
our previous finding that T-cell production of MIP-1 was important
in the recruitment of listeria-specific T lymphocytes, we focused on
the role of MIP-1 produced by T cells in GVHD. Our current data
suggest that T-cell production of MIP-1 is important in the
recruitment of alloreactive T lymphocytes. This was demonstrated by
showing the lack of bm1-specific T lymphocytes in the spleens of bm1
mice that received splenocytes from MIP-1/ mice. We believe that the best explanation for the data presented is the lack of
recruitment of alloreactive T lymphocytes to the spleen. However,
although we have found that T lymphocytes that do not produce MIP-1
can function as allospecific cytolytic lymphocytes, these cells do not
kill as well as wild-type cells. This would be consistent with data
from Taub et al35 that suggest enhanced killing by T
lymphocytes in the presence of  chemokines. The decreased ability to
kill target cells may also play a role in the decreased incidence of
GVHD in this setting.
We do not believe that the decreased incidence of GVHD after the
transfer of MIP-1/ splenocytes is due to the lack of
engraftment of the transferred splenocytes. This is based on the
following observations. (1) The model that we have used has been
previously shown to enable engraftment using sublethal
irradiation.34 (2) We have shown that
MIP-1/ B6 splenocytes engraft in the more stringent
MHC-mismatched transfer and that CD8+ lymphocytes mediate
lysis of host splenocytes after this transfer. (3) We have shown that
third-party-specific lymphocytes are present in bm1 recipients after
transfer of MIP-1/ B6 splenocytes. Because these
lymphocytes do not lyse splenocytes from C57BL/6 mice, they cannot be
from expanded endogenous cells but must come from the transferred
population of splenocytes. Finally, (4) the incidence of GVHD is always
greater than the severity of GVHD using splenocytes from
MIP-1/ B6 mice. This finding is not consistent with
enhanced survival solely being due to lack of engraftment and therefore
occurrence of GVHD.
The finding that T lymphocytes' production of MIP-1 is important in
the setting of a class I but not class II MHC mismatch was not entirely
unexpected. Previous investigators have also shown that
CD8+ lymphocytes are the predominant source of C-C
chemokines that block infectivity with human immunodeficiency
virus.36-39 MIP-1 has been shown to have less of a role
in the recruitment of activated CD4+
lymphocytes.40-42
Our data also show that the blockade of T-cell production of only
MIP-1 will have little effect on the occurrence of GVHD in the
setting of an MHC class II mismatch. Interestingly, the incidence of
lethal early GVHD was greater in bm12 mice that received splenocytes
from MIP-1/ mice as compared with B6 mice. The reason(s) for the enhanced effects on GVHD of splenocytes from MIP-1/ mice has not been addressed in the current
work. Quite possibly, splenocytes from MIP-1/ mice
overexpress other chemokines that may be more important in the
recruitment of activated CD4+ T lymphocytes. Alternatively,
the loss of MIP-1 may have less of an effect on the production of
cytokines or the lytic activity of activated CD4+
lymphocytes. We are currently investigating these possibilities.
Our finding of enhanced autoantibody production after transfer of
C57BL/6 splenocytes into irradiated bm1 recipients is intriguing. Previous investigators have shown that in the class II mismatched setting autoantibodies are produced by host B cells.43-45
If production of autoantibodies is similar in this model, the data
suggest that donor CD8+ lymphocytes either directly or
indirectly are capable of stimulating host B cells to produce
autoantibodies. We are currently performing work to examine this
possibility.
We have focused on the production of chemokines by lymphocytes because
of our data in the adoptive transfer model using Listeria monocytogenes. Current models suggest that local production of chemokines in the lung by alveolar macrophages may be quite important in the pathophysiology of asthma and infectious
pneumonitis.46-48 Our work does not eliminate the
possibility that local production of chemokines by macrophages,
endothelial cells, fibroblasts, and other cell types contribute to the
pathology in GVHD. Our findings of a marked decrease in the
inflammatory infiltrate in the lungs and liver of recipients of
MIP-1/ B6 splenocytes show that lymphocyte production
of MIP-1 may be more important in the incidence and severity of GVHD
in specific target organs than previously suspected.
We chose to use a nonlethal irradiation model for induction of GVHD for
two reasons. MIP-1 is expressed in the bone marrow microenvironment
and plays a critical role in stem cell kinetics.30 Therefore, bone marrow transplanted from MIP-1/ mice
is qualitatively different than bone marrow transplanted from C57BL/6
mice. This variable would be difficult to control and could lead to
differences in outcome that are independent of the occurrence of GVHD.
In addition, the system that we have used allows for differences in the
timing of GVHD, which is dependent on the number of lymphocytes infused
and the amount of irradiation administered to the recipients. This
allowed us to study the role of MIP-1 in the early and late events
associated with GVHD.
In summary, we have shown that blockade of T-cell production of a
chemokine may offer a new avenue for the prevention of GVHD. We have
shown that recipients that receive T cells that do not produce the C-C
chemokine, MIP-1, have a statistically improved survival and
decreased incidence of GVHD across a class I MHC barrier. However,
preventing T-cell production of MIP-1 had little effect on the
occurrence of GVHD when splenocytes were administered to class
II-mismatched recipients. Thus, an approach that blocks the function of
multiple different chemokines may provide a new avenue of prevention in
allogeneic BMT.
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FOOTNOTES |
Submitted April 7, 1998;
accepted August 24, 1998.
Supported by Public Health Service Grants No. CA66715 (J.S.S.) and
AI20288 (J.A.F.).
The publication costs of this
article were defrayed in part by
page charge payment. This article
must therefore be hereby marked
"advertisement"
in accordance with 18 U.S.C. section
1734 solely to indicate this fact.
Address reprint requests to Jonathan S. Serody, MD, Division of
Medical Oncology, Campus Box #7305, University of North Carolina School
of Medicine, Chapel Hill, NC 27599-7305; e-mail: serody{at}med.unc.edu.
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Abstract
Introduction