Blood, Vol. 93 No. 10 (May 15), 1999:
pp. 3558-3564
Prevention of Graft-Versus-Host Disease by Induction of Immune
Tolerance With Ultraviolet B-Irradiated Leukocytes in H-2 Disparate
Bone Marrow Donor
By
M.L.U. del Rosario,
James R. Zucali, and
K.J. Kao
From the University of Florida Departments of Pediatrics, Medicine,
and Pathology, Immunology & Laboratory Medicine, Gainesville, FL.
 |
ABSTRACT |
Transfusions (Tx) of Ultraviolet B (UVB)-irradiated peripheral blood
mononuclear leukocytes (MNL) have been shown to induce humoral immune
tolerance to major histocompatability complex (MHC) antigens
(Blood 88:4375, 1996). To determine whether cellular immune
tolerance to MHC antigens can be induced by the same approach, transplantation of bone marrow and spleen cells from tolerant donors
across the H-2 barrier was conducted to study its effect on prevention
of graft-versus-host disease (GVHD). After immune tolerance induction
by four weekly Tx of UVB-irradiated BALB/c (H-2d)
peripheral blood MNL into CBA/HT6 (H-2k) mice, bone marrow
cells (BMC) and spleen MNL from tolerant or naive CBA mice were
transplanted into lethally irradiated BALB/c mice. The transplanted
mice were followed by measuring body weight, peripheral leukocyte
counts, GVHD, survival, and cytokine response. All BALB/c recipient
mice were fully engrafted with H-2k CBA donor cells after
transplantation. The severity of GVHD was significantly attenuated in
BALB/c mice transplanted with BMC and spleen MNL from tolerant CBA
donor mice. The recovery of peripheral leukocyte and lymphocyte counts
were faster and more complete in mice transplanted with cells from the
tolerant donors. The serum cytokine profile after transplantation with
tolerant donor cells showed increased interleukin-4 and reduced gamma
interferon that are consistent with a polarized Th2 response. The
results pooled from three separate experiments showed that BALB/c mice transplanted with 5 × 106 BMC and 4 × 105
spleen MNL from tolerant CBA donors had better overall survival than
the control group (72% v 17%, P = .018). The
findings show that transplantation with bone marrow and spleen cells
from tolerant H-2 disparate donor mice is associated with significant
attenuation of GVHD and better outcomes. The results also support that
transfusions of UVB-irradiated leukocytes may induce cellular
immune tolerance.
© 1999 by The American Society of Hematology.
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INTRODUCTION |
IN SPITE OF THE WIDESPREAD use of
allogeneic bone marrow transplant (BMT) in the treatment of various
malignant and nonmalignant conditions, graft-versus-host disease (GVHD)
continues to be a serious complication after allogeneic BMT. GVHD,
whether acute or chronic, has been shown to occur in 30% to 60% of
patients receiving histocompatible sibling-matched allogeneic
BMT.1 Mortality attributed to GVHD has been reported in up
to 50% of cases.2-4 The severity of GVHD is related to the
degree of difference between hosts and donors across the major
histocompatibility (MHC) barrier5,6 and the presence of
donor T cells in marrow grafts.7-10 For this reason, T-cell
depletion from marrow grafts has been successfully applied to prevent
GVHD. Unfortunately, this approach has been associated with increased
incidences of graft failure, more severe immunosuppression, and higher
relapse rates of original malignancies.7,8,11-14 Therefore,
developing a method for inducing specific humoral and cellular immune
tolerance across the MHC barrier offers an attractive alternative
approach to prevent or attenuate GVHD.
Irradiation of leukocytes with medium (UVB) or short (UVC) wavelength
ultraviolet light has been shown to produce various immunomodulatory
effects.15-21 These effects include inhibition of
stimulator and responder function of leukocytes in mixed leukocyte culture (MLC),15,16 suppression of cytokine
production,17 downregulation of cell membrane proteins such
as class-II MHC antigens18 and cell adhesion
molecules,19 reduced lymphocyte proliferative responses to
mitogenic lectins,20 and inhibition of expression of
costimulatory signals by antigen presenting cells.21 On the
basis of these findings, UVC and UVB have been applied to inactivate
leukocytes for reducing the immunogenicity of platelet concentrates.22-25 Results from these studies indicate that
transfusions of platelet concentrates irradiated with UVB or UVC could
induce humoral immune tolerance to allogeneic MHC antigens. However, the induced tolerance was either incomplete22 or
inconsistent.24
To overcome the problems of inconsistency and partial tolerance, we
investigated the effect of plasma and platelets on tolerance induction
by UVB-irradiated leukocytes. Our study showed that the presence of
plasma and platelets interfered with tolerance induction and that the
use of highly purified peripheral mononuclear leukocytes (MNL) is
critical for consistent induction of complete humoral immune tolerance
to allogeneic MHC antigens.26 However, it is not known
whether cellular immune tolerance can be induced by the same approach.
To answer this question and to explore the potential application of
this approach to allogeneic BMT, we conducted experiments to determine
whether transplantation with bone marrow and spleen cells from H-2
disparate tolerant donor mice could prevent or attenuate the GVHD
induced by allogeneic BMT in a murine model.
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MATERIALS AND METHODS |
Animals.
Eight-week old CBA/CaH-T6/J (CBA) mice with H-2k MHC
haplotype and BALB/cByJ (BALB/c) mice with H-2d MHC
haplotype were obtained from Jackson Laboratory (Bar Harbor, ME).
BALB/c mice were also obtained from Harlan Sprague Dawley, Inc
(Indianapolis, IN). All mice were housed in a temperature-controlled room (25°C) with a 12-hour interval light/dark cycle and fed ad libitum. All experiments were approved by the Institutional Animal Care
and Use Committee.
Preparation of UVB-irradiated peripheral mononuclear leukocytes.
Peripheral mononuclear leukocytes were prepared from freshly collected
BALB/c venous blood by differential and Ficoll-hypaque density gradient
centrifugation as described.22 MNL were suspended in
phosphate buffered saline (PBS) and their concentrations were determined by hemacytometers after staining cells with propidium iodide
solution.27 For irradiation of leukocytes by UVB, a
Bioslink-UV irradiator (BIOS Corp, New Haven, CT) with a built-in
dosimeter was used. The dose of UVB irradiation was 1,200 mJ/cm2. The irradiation was performed in an open sterile
polypropylene container. The depth of cell suspension during
UVB-irradiation was 1 mm. The cell suspensions were mixed manually by
moving the tray back and forth during irradiation.
Induction of immune tolerance.
Immune tolerance was induced in CBA mice by four weekly intravenous
injections of 2 × 105 UVB-irradiated BALB/c MNL in
100 µL PBS through a tail vein under light anesthesia with inhalation
of methoxyflurane (Pitman-Moore Inc, Mundelein, IL). Preimmune serum
samples were prepared from venous blood that was collected by
retro-orbital bleeding 2 days before the first transfusion. One week
after the last weekly transfusion of UVB-irradiated leukocytes, serum
samples were collected to assess the development of antibodies to donor
MHC antigens by flow cytometry as described.26 As reported
previously,26 CBA mice became tolerant, if they did not
develop any anti-H2d antibodies after four weekly
transfusions of UVB-irradiated BALB/c MNL. To insure the development of
tolerance, two transfused CBA mice were randomly selected and
challenged with two weekly transfusions of 1 × 105
untreated BALB/c MNL. We found that none of the challenged CBA mice
became immunized to H-2d antigens of BALB/c mice as
expected. In contrast, all control naive CBA mice developed
anti-H-2d antibody after two transfusion challenges.
BMT.
Naive and tolerant CBA mice were used as donors of bone marrow and
spleen cells. BALB/c mice were used as bone marrow recipients. BMC from
CBA mice were prepared by flushing cells out of femurs and tibias using
25-gauge needles and syringes filled with RPMI 1640 medium as
described.28 Spleen MNL were prepared from bone marrow
donor mice by Ficoll gradient centrifugation.22 Fifty µL
of spleen MNL containing specified numbers of cells were mixed with 5 × 106 bone marrow cells in a final volume of 200 µL. These cells were transplanted through a tail vein into each
BALB/c mouse that had been lethally irradiated with 750 cGy gamma ray 5 to 6 hours before transplantation. The transplanted mice were fed with
sterilized laboratory chow and acidified water. They were followed for
engraftment of donor cells by immunofluorescence flow cytometry. Body
weight, peripheral total and differential white cell count, GVHD, and survival were measured.
Immunofluorescence flow cytometry for detection of engraftment and
characterization of lymphocyte subsets.
H-2 phenotypes of peripheral blood leukocytes were determined by
immunofluorescence flow cytometry using fluorescein isothiocyanate (FITC)-labeled anti-H2d antibody and phycoerythrin
(PE)-labeled anti-H2k antibody. Different subsets of
lymphocytes including B cells, CD4+ helper T cells, and
CD8+ cytotoxic T cells were measured using PE-labeled
anti-B220, FITC-labeled anti-CD4, and PE-labeled anti-CD8 antibodies.
All antibodies were purchased from PharMingen Corp (San Diego, CA).
Peripheral leukocytes for flow-cytometric analysis were prepared by
washing 20µL of whole blood with 1 mL PBS and lysing red cells with
1.5 mL of 0.85% ammonium chloride solution. After lysis of red cells,
the remaining cells were washed twice with 0.5 mL PBS and resuspended in 100 µL PBS-0.02% azide-0.5% bovine serum albumin (BSA). Fifty µL of washed leukocytes were stained with appropriate concentrations of fluorescent antibodies for 30 minutes. After two washes, the cells
were analyzed using a FACscan flow cytometer (Becton Dickinson, San
Jose, CA).
Enzyme-linked immunoassays (EIA) for interleukin-4 (IL-4) and
-interferon (-IFN).
For the assays, plastic wells of a microtiter plate were coated with 50 µL of 5µg/mL antimouse IL-4 or -IFN antibody in PBS-azide overnight at 4°C. The plates were washed three times using
PBS-azide-0.2% Tween-20 and blocked with 150 µL PBS containing 10%
newborn calf serum (NCS) for 30 minutes at room temperature.
Thereafter, 50 µL of cytokine standards and diluted serum samples
were added and incubated overnight at 4°C. Serum samples were
diluted with equal volume of PBS-10% NCS. After overnight incubation,
plates were washed and incubated sequentially with 50 µL of 5 µg/mL
biotinylated anticytokine antibody diluted in PBS-Tween-1% BSA,
strepavidin-peroxidase (Sigma Co, St Louis, MO), and o-phenylenediamine
dihydrochloride peroxidase substrate (Sigma Co). Paired anti-IL-4 and
anti-IFN antibodies, recombinant murine IL-4 and -IFN used in the
assays were obtained from PharMingen (San Diego, CA) and Endogen
(Woburn, MA), respectively. The lowest concentrations of IL-4 and
-IFN that can be measured by these two assays were 0.2 pg/mL, and 50 pg/mL, respectively.
GVHD scoring.
A scoring system was devised to grade GVHD in the transplanted mice.
Hair and skin changes involving the head/neck, abdomen and tail, and
changes in anal and perianal mucosa were scored according to severity
from 0 to 3 as defined in Table 1. Mice with a total score of less than 3 were regarded as having mild GVHD,
3 to 
5 as having moderate GVHD and total score of greater than 5 as
severe GVHD.
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RESULTS |
Induction of GVHD.
Unlike humans, only a small number of T cells are present in murine
bone marrow. According to our flow cytometric study, CD3+ T
cells account for less than 1% of total bone marrow cells harvested from CBA mice. Therefore, transplantation of allogeneic bone marrow cells alone does not always result in clinically apparent GVHD in
BALB/c recipient mice. Addition of appropriate numbers of spleen MNL to
BMC before transplantation is necessary to induce clinically apparent
GVHD. To determine the optimal numbers of spleen MNL that are required
for inducing clinically significant GVHD, we first studied how
different numbers of spleen MNL mixed with 5 × 106
BMC affected the severity of GVHD. The results of our study indicate that moderate to severe GVHD can be induced by including 2.5 to 4 × 105 donor spleen MNL with 5 × 106
BMC. Transplantation of 1 × 106 or more spleen MNL
was associated with hyperacute GVHD in which most recipient BALB/c mice
died within 8 days after transplantation. Inclusion of less than
1 × 105 spleen MNL was associated with nil
to mild GVHD. GVHD was judged by the presence of hair loss, skin
changes, body weight loss, and stunted weight gain and scored according
to the criteria described in Table 1.
GVHD after transplantation with bone marrow and spleen cells from
tolerant donors.
To determine whether GVHD can be prevented or attenuated by
transplantation of cells from CBA mice tolerant to MHC antigens of
BALB/c mice, we studied how the addition of 4 × 105
or 2.5 × 105 spleen MNL from naive or tolerant CBA
mice to their respective bone marrow cells affected the transplantation
associated GVHD in recipient BALB/c mice. All transplanted BALB/c mice
were followed weekly for changes in body weight and development of
GVHD. As shown in Fig 1 and
2, recipient BALB/c mice transplanted
with two different doses of naive control spleen cells and 5 × 106 BMC had poor body weight gain and moderate to severe
GVHD. Most BALB/c mice transplanted with two different doses of
tolerant spleen MNL and 5 × 106 BMC had better body
weight recovery (Fig 1) and significantly reduced GVHD (Fig
2) when they were compared with recipients of naive control cells.
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| Fig 1.
Changes of body weight after transplantation of lethally
irradiated BALB/c mice with 5 × 106 bone marrow cells and
either 2.5 × 105 (Experiment I) or 4 × 105 (Experiment II) spleen MNL from naive (Experiment Ia
and IIa) or tolerant (Experiment Ib and IIb) CBA mice. There were four
mice in each group. ( ): Death of transplanted BALB/c mouse.
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| Fig 2.
Development of GVHD after transplantation of lethally
irradiated BALB/c mice with bone marrow and spleen MNL from naive
( , ) or tolerant ( , ) CBA mice as described in Fig 1.
Experiment I: 5 × 106 bone marrow cells + 2.5 × 105 spleen MNL. Experiment II: 5 × 106 bone
marrow cells + 4 × 105 spleen MNL. Each value is mean ± SD.
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Engraftment and recovery of peripheral leukocytes.
Two weeks after BMT, all H-2d recipient mice became
completely engrafted with donor H-2k+ leukocytes in
peripheral blood as assessed by immunofluorescence flow cytometry. To
determine whether there was any difference in the rate of engraftment,
peripheral total and differential leukocyte counts were followed.
Peripheral lymphocyte counts including T and B cells subsets were
measured according to peripheral blood total and differential leukocyte
counts and immunophenotyping results for T and B cells by flow
cytometry. As shown in Fig 3, there was no
significant difference in the initial recovery of peripheral leukocyte
counts between BALB/c recipient mice transplanted with either naive or
tolerant donor bone marrow and spleen cells. Nevertheless, lymphocyte
recovery was faster especially in recipients of 4 × 105 tolerant spleen MNL (Fig 4,
Experiment II). BALB/c mice transplanted with 5 × 106
BMC and 4 × 105 spleen MNL from tolerant donors also
showed a faster and more complete recovery of T and B cells in the
peripheral blood (Fig 5). In
addition, it was noted that the recovery of CD8+ T cells
was also faster after transplantation with bone marrow and spleen cells
from tolerant donor mice (Fig 6). As shown
in Fig 6, the recovery of CD8+ T cells in general lagged
behind CD4+ T cells in our murine bone marrow
transplantation model.
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| Fig 3.
Changes of WBC counts in peripheral blood after
transplantation of lethally irradiated BALB/c mice with bone marrow and
spleen cells from naive (,) or tolerant (,) CBA mice as
described in Fig 1. Experiment I: 5 × 106 bone marrow
cells + 2.5 × 105 spleen MNL. Experiment II: 5 × 106 bone marrow cells + 4 × 105 spleen MNL.
Each value is mean ± SD.
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| Fig 4.
Changes of lymphocyte counts in peripheral blood after
transplantation of lethally irradiated BALB/c mice with bone marrow and
spleen cells from naive (,) or tolerant (,) CBA mice as
described in Fig 1. Experiment I: 5 × 106 bone marrow
cells + 2.5 × 105 spleen MNL. Experiment II: 5 × 106 bone marrow cells + 4 × 105 spleen MNL.
Each value is mean ± SD.
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| Fig 5.
Changes of B and T cell counts in peripheral blood after
transplantation of lethally irradiated BALB/c mice with 5 × 106 bone marrow and 4 × 105 spleen cells from
naive () or tolerant () CBA mice. Each value is mean ± SD.
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| Fig 6.
Changes of CD4/CD8 T cell ratio in peripheral blood after
transplantation of lethally irradiated BALB/c mice with 5 × 106 bone marrow and 4 × 105 spleen cells from
naive () or tolerant () CBA mice. Each value is mean ± SD.
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Survival after BMT.
Because the initial results of our study (Fig 1) showed that
transplantation of tolerant BMC and spleen MNL had a significant positive effect on overall survival, two additional bone marrow transplantation experiments using 5 × 106 BMC and 4 × 105 spleen MNL were conducted to confirm this
finding. There were five mice in each experimental group. The survival
data from these three separate experiments were pooled and analyzed. As
shown in Fig 7, the overall long-term
survival of mice transplanted with cells from tolerant donors was 72%
versus 17% (P =.018) for those transplanted with naive cells.
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| Fig 7.
Kaplan-Meier survival curve of lethally irradiated BALB/c
mice transplanted with 5 × 106 bone marrow cells and 4 × 105 spleen MNL from naive () or tolerant () CBA
mice. The data pooled from three separate experiments were analyzed.
There were five mice in each treatment group in two experiments and
four mice in one experiment. P value was determined by logrank
test. A total of 14 mice were included in each treatment group.
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T-cell cytokine response after BMT.
According to our recent study,29 tolerance induction by
transfusions with UVB-irradiated BALB/c leukocytes polarizes T cells of
CBA mice to produce of type-2 cytokines during in vitro stimulation by
BALB/c spleen cells. It was of interest to study the serum levels of
-IFN and IL-4 in BALB/c mice after transplantation with cells from
tolerant CBA donors. In this study, three mice from each treatment
group were randomly selected and sacrificed every other day until day 9 after BMT. Equal volumes of serum samples from three mice were pooled
to obtain sufficient amounts of samples for assays of IL-4 and -IFN.
The results show that recipient mice transplanted with tolerant BMC and
spleen MNL had lower peak serum levels of -IFN and higher levels of
IL-4 than those transplanted with cells from naive CBA donor mice after BMT, respectively (Fig 8).
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| Fig 8.
Serum IL-4 and gamma interferon levels after
transplantation of lethally irradiated BALB/c mice with 5 × 106 bone marrow and 4 × 105 spleen cells from
naive () or tolerant () CBA mice. Three recipient mice from each
treatment group were sacrificed every other day and equal volumes of
serum samples were pooled for the assays. The sera were assayed in
duplicate. Variations between duplicates were less than 17%. Similar
results were obtained when the experiment was repeated.
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DISCUSSION |
T cells in allogeneic bone marrow are responsible for BMT-associated
GVHD.7-10 Therefore, rendering donor T cells tolerant toward recipient MHC antigens offers an attractive approach to prevent
or reduce GVHD. Our recent study showed that humoral immune tolerance
to allogeneic MHC antigens can be induced without nonspecific immunosuppression by transfusions of UVB-irradiated leukocytes from H-2
disparate mice.26 It was therefore of interest to learn whether GVHD can be prevented by transplantation of bone marrow and
spleen mononuclear cells from H-2 disparate donor mice that have been
tolerized towards MHC antigens of BMT recipient mice. To answer this
question, we conducted such a study. The results reported herein show
that transplantation with bone marrow and spleen cells from tolerant
CBA (H2k) donor mice to BALB/c (H2d) recipient
mice is associated with significant attenuation of GVHD, better
long-term survival, faster reconstitution of peripheral lymphocytes,
and Th2 cytokine response immediately after transplantation. Because
cell-mediated immunity plays an important role in the pathogenesis of
GVHD, these findings suggest that transfusions of UVB-irradiated
leukocytes may induce cellular immune tolerance. This conclusion is
further substantiated by our recent study of cytotoxic T-cell activity
against H-2d+ target cells in the CBA mice that had been
tolerized by transfusions of UVB-irradiated BALB/c mononuclear
leukocytes and subsequently challenged with two fully immunogenic doses
of untreated BALB/c leukocytes (unpublished observation).
In our study, two different doses of spleen MNL (2.5 × 105 and 4 × 105) were used to induce
GVHD. These two doses were selected for their ability to elicit
significant degrees of acute GVHD without causing premature death so
that the transplanted mice could be followed for an extended period of
time. Among BALB/c recipient mice transplanted with cells from naive
donor mice, we noted that recipient BALB/c mice transplanted with a
lower number (2.5 × 105) spleen MNL had less severe
and lasting GVHD (Fig 2). This finding confirms the quantitative
importance of donor lymphocytes in eliciting GVHD as previously
reported.30-33 The results of our study also showed that 1 month after BMT, peripheral leukocyte counts recovered to normal levels
in most recipient mice regardless of whether donor cells were from
naive or tolerant CBA mice. However, during the next 4 months
peripheral leukocyte counts began to decline until GVHD largely
resolved 150 days after BMT. In addition to GVHD, this decline of
peripheral leukocyte counts was likely a result of delayed repopulation
by more primitive long-term repopulating hematopoietic stem cells as
observed in bone marrow transplant recipients.34,35 We also
noted that recovery of peripheral T- and B-lymphocyte counts was faster
in mice transplanted with bone marrow and spleen cells from the
tolerized CBA donors (Fig 5). The cell counts and immunophenotyping
results are consistent with the subjective clinical assessment of GVHD.
All these findings support the beneficial effect of using cells from
the tolerized donor mice for bone marrow transplantation across major
MHC barrier. However, it is not known whether faster quantitative
lymphoid reconstitution would translate to better immune function.
Further studies in this regard are needed.
According to cytokine production profile, two subsets of helper (Th)
and cytotoxic (Tc) T cells have been identified.36,37 Secretion of -IFN and tumor necrosis factor-
(TNF-) defines type-1 Th cells (Th1), and secretion of IL-4, IL-5, and IL-10 defines
type-2 Th cells (Th2). Th1 cells are functionally involved in
cell-mediated immunity, delayed-type hypersensitivity, production of
IgG2a and IgG2b antibodies, and host defense to infection of intracellular pathogens.38-42 Th2 cells are involved in
production of IgG1 and IgE antibodies, allergic conditions, and host
defense to infection of extracellular pathogens.38-42 Th1
and Th2 responses during host-immune response have been shown to be
mutually inhibitory.42 Thus, immune responses can be
characterized by polarization toward Th1 or Th2 response.
Induction of tolerance by priming neonatal mice with allogeneic
leukocytes has been associated with increased production Th2 cytokines
and reduced secretion of Th1 cytokines.43 The finding indicates that tolerance induction leads the immune system towards an
enhanced Th2 response.44-46 Our recent study of cytokine
production in mixed leukocyte culture reactions using tolerized CBA
spleen T lymphocytes as responder and BALB/c spleen leukocytes as
stimulators showed an increase in IL4 and IL5 production (data not
shown). In allogeneic BMT, donor lymphocytes with enhanced Th2 cytokine response have been associated with a much-reduced acute
GVHD.47 For this reason, we measured serum IL-4 and -IFN
levels after transplantation with bone marrow and spleen cells from
tolerant or naive donor CBA mice. Our results showed an enhanced
transient Th2 cytokine response in recipient mice transplanted with
cells from tolerant donors. The results are consistent with our recent in vitro MLC finding and support the earlier reports that polarization of donor T cells to Th2 response is associated with reduced severity of
acute GVHD.47-49 Nevertheless, we are unable to ascertain
whether the observed differences in cytokine production were due to
donor cells, recipient cells, or both. Although the polarized Th2
response in donors may contribute to protection of acute GVHD as
suggested by various investigators,47-49 it is possible
that donor Th2 cytokine response only reflects changes in the effector
arm of the donor immune system and may not play a direct role in the
prevention of GVHD associated with allogeneic BMT. The exact mechanism
by which Th2 polarization of the bone marrow donor immune system could
attenuate acute GVHD remains to be further elucidated.
In the present study, we found that acute GVHD can not be completely
prevented by transplantation of bone marrow and spleen cells from
tolerant CBA donors (Fig 2). This finding is not unexpected. Our recent
MLC study using T cells from tolerant CBA mice as responders and
gamma-irradiated or mitomycin-treated spleen leukocytes from BALB/c
mice as stimulators indicated that T cells from tolerant CBA mice
remain capable of producing type-1 T-cell cytokines such as -IFN but
at a reduced level.29 Because type-1 T-cell cytokines have
been implicated in the pathogenesis of acute GVHD,50 the reduced production of type-1 T-cell cytokines by T cells from tolerant
donor mice might be responsible for the attenuated GVHD observed in our experiments.
In view of the feasibility and the safety of transfusing patients with
UVB-irradiated platelet concentrates,51 the results of our
study support the potential clinical application of UVB-irradiated leukocytes for tolerance induction in bone marrow donors to prevent GVHD. However, the safety and the ethical concerns of using leukocytes from patients with neoplastic diseases to induce tolerance in healthy
bone marrow donors preclude such an approach. Nevertheless, tolerance
induction in related bone marrow donors may be considered for patients
who suffer from hereditary non-neoplastic hematological disorders, such
as sickle cell anemia and thalassemia and who are not at risk of
transmitting infectious diseases to their bone marrow donors. Another
potential approach is to induce immune tolerance in patients using
UVB-irradiated leukocytes from healthy bone marrow donors. The
tolerized patients would then undergo bone marrow transplant using a
nonmyeloablative conditioning regimen. Such an approach may allow the
establishment of macro-mixed chimerism that is sufficient to ameliorate
non-neoplastic hematological disorders without increasing the risk of
graft rejection. At the same time, high-dose chemoradiotherapy related
toxicity is avoided. Obviously, further animal studies are needed
before the proposed approach can be applied to the clinical setting.
| |
ACKNOWLEDGMENT |
The authors are grateful to Sandra Donahue for her excellent technical assistance.
| |
FOOTNOTES |
Submitted September 22, 1998; accepted January 21, 1999.
Supported by Grant R01 HL-58809 from the National Institutes of Health.
Address all correspondence to K.J. Kao, MD, Box 100275, Department of
Pathology, Immunology & Laboratory Medicine, University of Florida,
Gainesville, FL 32610; e-mail: kjkao{at}ufl.edu.
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.
| |
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ABSTRACT
INTRODUCTION
