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Receptor Plays a Critical Role in T Cell Alloreactivity1
*
Department of Pediatric Oncology, Dana-Farber Cancer Institute, Harvard Medical School, Boston, MA 02115;
Mater Medical Research Institute, South Brisbane, Australia; and
Departments of Internal Medicine and Pediatrics, Bone Marrow Transplant Program, University of Michigan Cancer Center, MI 48109
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Abstract |
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 Top Abstract IntroductionMaterials and MethodsResultsDiscussionReferences |
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is known to be an important mediator of tissue damage
during allograft rejection and graft-vs-host disease (GVHD), but its
role in supporting T cell responses to allogeneic Ags is unclear. We
have studied this question by comparing normal mice with those lacking
the p55 (p55 TNFR-/-) or p75 (p75 TNFR-/-)
TNF- receptors as donors in well-defined bone marrow transplant
(BMT) models. Recipients of p55 TNFR-/- cells had
significantly reduced mortality and morbidity from GVHD compared with
the other two sources of T cells. In vitro, T cells lacking the p55
(but not the p75) TNF- receptor exhibited decreased proliferation
and production of Th1 cytokines in MLC. This defect was only partially
restored by exogenous IL-2 and affected both CD4+ and
CD8+ populations. CD8+ p55
TNFR-/- proliferation was impaired independently of IL-2
whereas CTL effector function was impaired in an IL-2-dependent
fashion. Inhibition of TNF- with TNFR:Fc in primary MLC also
impaired the proliferation and Th1 differentiation of wild-type T
cells. BMT mixing experiments demonstrated that the reduced ability of
p55 TNFR-/- donor cells to induce GVHD was due to the
absence of the p55 TNFR on T cells rather than bone marrow cells. These
data highlight the importance of TNF- in alloreactive T cell
responses and suggest that inhibition of the T cell p55 TNF-
receptor may provide an additional useful therapeutic maneuver to
inhibit alloreactive T cell responses following bone marrow and solid
organ transplantation. |
Introduction |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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binds
to two receptors, p55 and p75, both of which are expressed (in varying
degrees) on all hematopoetic cells with the exception of erythroid and
unstimulated T cells (1, 2). The two TNF- receptors are
type I transmembrane proteins that form part of the TNF superfamily and
include CD30, CD27, CD40, 4-1BB, and Fas. Most human peripheral blood T
cells and CD4+ T cell clones show enhanced
proliferation when the TNF- receptor is bound to its ligand in the
presence of mAbs to CD3. This process of autocrine T cell stimulation
is mediated predominantly through cell-cell contact, particularly for
the CD40-CD40 ligand and 4-1BB-4-1BB ligand pairs (1).
TNF- can also induce cytotoxicity, which is predominantly linked to
the p55 TNF- receptor (3, 4). However, it is clear that
TNF- binding to the p75 receptor can enhance cytotoxicity through
the p55 receptor by a process of ligand passing (5, 6, 7). In
addition, the p75 TNF- receptor appears to be the primary ligand for
membrane-bound TNF-, which can mediate cytotoxicity independently of
soluble TNF- (7).
The role of the TNF- receptors in innate and adaptive immunity is
further complicated by the fact that these proteins act as receptors
for both TNF- and the lymphotoxin-
(LT)3 homotrimer
(8). In contrast, LT/LT
heterodimers bind
specifically to the LT receptor (8). The recent
generation of mice deficient in TNF-, LT and the p55 and p75
TNF- receptors, has clarified their respective roles in the
developing immune system. Mice lacking TNF- lack splenic primary B
cell follicles and cannot form organized follicular dendritic cell
networks and germinal centers (9), whereas mice lacking
the p55 TNF- receptor have normal segregation of T and B lymphocytes
in the lymph nodes and spleen, but lack Peyer’s patches
(8). Lymphoid organs in p75
TNFR-/- animals appear to be normal
(10). Animals without LT do not have lymph nodes or
Peyer’s patches and have disorganized splenic white pulp
(8). Mice lacking either LT or the p55 TNF- receptor
do not possess organized clusters of follicular dendritic cells in the
spleen (11). Interestingly, the organization of follicular
dendritic cells and the formation of germinal centers are controlled by
bone marrow-derived cells that secrete LT and non-bone
marrow-derived cells that express p55 TNF- receptors
(11). Recent evidence suggests that both TNF- and
LT/LT heterodimers are required for the development and function
of B and T zone stromal cells (12). An increasing body of
evidence therefore points to a critical role for TNF- and the p55
TNF- receptor in the optimal development of the immune system.
We have studied the role of TNF- in T cell responses in vivo using
well-characterized mouse models of graft-vs-host disease (GVHD).
TNF- may be important in GVHD in at least three ways. First, TNF-
may cause direct cytotoxicity to host tissues. Second, TNF- is known
to enhance the expression of MHC molecules (13) and
adhesion molecules (14), which are likely to enhance donor
T cell activation by host tissues. Finally, TNF- may act as an
autocrine T cell growth factor (15) and therefore increase
donor T cell clonal expansion. Using TNF- receptor knock-out mice,
we demonstrate that T cells lacking the p55 TNF- receptor have
impaired responses to alloantigen in vitro, including reduced
proliferation, type 1 cytokine production, and cytolytic function. In
vivo, T cells lacking the p55 TNF- receptor have a reduced capacity
to induce GVHD, which is also characterized by impaired IFN-
and
TNF- production, reduced CD8+ expansion, and
impaired cytolytic function. These data suggest that TNF- plays a
critical role in T cell responses to alloantigens.
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Materials and Methods |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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Female C57BL/6 (B6, H-2b), B6C3F1
(H-2bxk), B6SJLF1 (H-2bxs),
and B6D2F1 (H-2bxd) mice were purchased from The
Jackson Laboratories (Bar Harbor, ME). B6 mice deficient in the p55
TNF- receptor and p75 TNF- receptor were supplied by Immunex
(Seattle, WA) and have been previously described (10, 16, 17, 18). The age of mice used as bone marrow transplant (BMT)
recipients ranged between 9 and 15 wk and the age of donors was >24
wk. Mice were housed in sterilized microisolator cages and received
filtered water and normal chow, or autoclaved hyperchlorinated drinking
water for the first 2 wk after BMT.
Bone marrow transplantation
Mice were transplanted according to a standard protocol described previously (19, 20). On day 0, mice received 1300 cGy total body irradiation (TBI) (137Cs source), split into two doses separated by 3 h to minimize gastrointestinal toxicity. A total of 5 x 106 bone marrow cells and 2 x 106 nylon wool purified splenic donor T cells were resuspended in 0.25 ml of Leibovitz’s L-15 medium (Life Technologies, Gaithersburg MD)) and injected i.v. into recipients. In some experiments, bone marrow was T cell depleted with anti-Thy 1.2 and rabbit complement. Survival was monitored daily, and recipient’s body weights and GVHD clinical score were measured weekly.
Assessment of GVHD
The degree of systemic GVHD was assessed by a scoring system which sums changes in five clinical parameters: weight loss, posture (hunching), activity, fur texture, and skin integrity (maximum index = 10) (21). Individual mice were ear-tagged and graded weekly from 0 to 2 for each criterion without knowledge of treatment group.
FACS analysis
FITC-conjugated mAbs to mouse H-2Dd, CD3, CD4, and CD11b, and PE-conjugated CD4, CD8, CD44, CD62L, CD45RB, and B220 were purchased from PharMingen (San Diego, CA). Cells were first incubated with mAb 2.4G2 for 15 min at 4°C, then with the relevant FITC- or PE-conjugated mAb for 30 min at 4°C. Finally, cells were washed twice with PBS/0.2% BSA, fixed with PBS/1% paraformaldehyde, and analyzed by FACScan (Becton Dickinson, San Jose, CA). Donor T cell expansion was determined by multiplying the fraction of CD4+ and CD8+ cells that were H-2Dd negative in the spleen by the total number of spleen cells 2 wk after BMT.
Cell cultures
All culture media and incubation conditions were as previously
described (19, 20). In studies of naive mice, splenic T
cells were enriched by passage through nylon wool. T cell fractions
after this process ranged from 60 to 70% and were similar in each
group. These responder T cells were then plated in 96 flat-bottom
plates (Falcon, Lincoln park, NJ) at a concentration of 1 or 2 x
105 T cells (CD4+ plus
CD8+)/well with 0.5 or 1 x
105 irradiated (2000 rad) peritoneal macrophages
lavaged from naive B6D2F1 (allogeneic) or B6 (syngeneic) animals. At
72 h, cultures were pulsed with
[3H]thymidine (1 µCi/well), and proliferation
was determined 20 h later on a 1205 Betaplate reader (Wallac,
Turku, Finland). Purified CD4+ and
CD8+ T cell subsets were obtained by selecting
CD4+ and CD8+ cells from
nylon wool purified splenic cells using minimacs columns (Miltenyi
Biotech, Bergisch Gladbach, Germany). Supernatant was removed at
48 h for IL-2 determination and 72 h for IFN- and IL-4
determination. To block TNF-, 100 µg/ml of TNFR:Fc (Immunex) or
control human Ig was added to MLC. In experiments analyzing T cell
responses after BMT, splenocytes were removed from animals 14 days
after transplant, and three to six spleens were combined from each
group. Single cell suspensions were then layered over Ficoll-Paque
(Pharmacia Biotech, Piscataway, NJ) and centrifuged at 800 x
g for 15 min. Cells were collected from the interface and
washed twice before suspension in supplemented 10% FCS/RPMI. Cells
were plated in the same concentrations as above and were pulsed with
[3H]thymidine at 48 h; plates were
harvested 20 h later. Supernatant was removed 48 h after
culture for determination of cytokines. Mitogenic responses were
determined by stimulating 105 T cells with 10
µg/ml of plate-bound CD3 (145-2C11, PharMingen) or 2.5 µg/ml of Con
A. In some cultures, CD28 (37.51, PharMingen) was added at a
concentration of 1 µg/ml. [3H]Thymidine was
added to these cultures at 40 h, and plates were harvested 20
h later.
Cytokine ELISA
The Abs used in the TNF- assay were purchased from Genzyme
(Cambridge, MA). Abs used in the IFN-, IL-2, and IL-4 assays were
purchased from PharMingen. All assays were performed according to the
manufacturer’s protocol.
51Cr release assays
A total of 2 x 106 P815 (H-2d) or EL4 (H-2b) tumor targets were labeled with 100 µCi of 51Cr for 2 h. After washing three times, labeled targets were plated at 104 cells per well in U-bottom plates (Costar, Cambridge, MA). Splenocytes from allogeneic BMT recipients (prepared as described above) were added to quadruplicate wells at varying E:T ratios and incubated for 5 h. Maximal and background release was determined by the addition of Triton X-100 (Sigma, St. Louis, MO) or medium alone to targets, respectively. 51Cr activity in supernatants taken 5 h later were determined in a Cobra autogamma counter (Cobra, Meriden, CT), and lysis was expressed as a percentage of maximum. Lytic units were calculated as the number of CD8+ cells per million required to lyse 20% of P815 (H2d) targets (106/E:T ratio at 20% lysis x number P815 targets per well). Because P815 express class I, little, or no class II and are resistant to Fas-induced apoptosis (22), these assays measure predominantly CD8+ perforin-mediated cytotoxicity.
Statistical analysis
Survival curves were plotted using Kaplan-Meier estimates. The Mann-Whitney U test was used for the statistical analysis of cytokine data, LPS levels, clinical scores, and weight loss, whereas the Mantel-Cox log-rank test was used to analyze survival data. A p value of <0.05 was considered statistically significant.
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Results |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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To study the responses of TNFR-/- T cells
to alloantigen in vivo, we used well established murine models of GVHD
that are directed to both major and minor histocompatibility Ags (B6
B6D2F1 and B6 B6C3F1). TNF- production in these BMT models
is high and neutralization of TNF- at the time of BMT with
recombinant human TNF- receptor:Fc (TNFR:Fc) effectively prevents
GVHD (19). We transplanted bone marrow and nylon wool
purified T cells from wild-type, p55 TNFR-/-,
or p75 TNFR-/- B6 animals into B6D2F1 or B6C3F1
recipients after 1300 cGy of TBI. Fig. 1
A shows that GVHD mortality
was significantly reduced in B6C3F1 recipients of p55
TNFR-/- cells compared with recipients of
wild-type and p75 TNFR-/- cells (day 70
survival: 90% vs 55% and 40%, p < 0.05). Although
moderate GVHD (clinical scores between 2 and 4) was present in animals
transplanted with p55 TNFR-/- cells, this was
significantly less than recipients of wild-type or p75
TNFR-/- cells. Interestingly, recipients of p75
TNFR-/- cells in this model tended to have more
severe GVHD than recipients of wild-type cells. In the B6 B6D2F1
strain combination, recipients of p55 TNFR -/- cells had less
mortality from GVHD (survival at day 70: 80% vs 45%) and an
impressive reduction in the severity of GVHD was seen (clinical scores
<2) in surviving animals (Fig. 1B).
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receptor proliferate poorly to
alloantigen
We next tested splenic T cells from mice lacking either the p55
TNF- receptor or p75 TNF- receptor for responses to alloantigen
in primary mixed lymphocyte cultures. T cells from p55
TNFR-/- animals had markedly impaired
proliferation and IL-2 production to H2bxd
(B6D2F1) stimulators, whereas responses from p75
TNFR-/- T cells were equivalent to wild type
(Table I). Exogenous IL-2 only partially
restored the proliferative response however, suggesting that the
impaired production of IL-2 was not completely responsible for the
impaired proliferation (Table I). The proportion of apoptotic cells was
not increased in MLC using p55 TNFR-/-
responder T cells compared with wild-type or p75
TNFR-/- cells (4% vs 6% and 17%,
respectively). Despite the reduced response to alloantigen, responses
of TNFR-/- T cells to Con A and CD28 were
normal (Table I). In addition, the numbers of T cells in the spleens of
naive p55 TNFR-/- animals were normal although
there was a significant increase in the proportion of memory T cells
(CD44high, CD62Llow,
CD45RBlow) compared with wild-type animals (Table II). Conversely, there was a significant
reduction in the proportion of memory T cells in p75
TNFR-/- spleens compared with those from
wild-type animals (Table II), suggesting that the p55 and p75 TNF-
receptors may play opposing roles in the generation of T cell memory.
The most pronounced abnormality in the spleens of p55
TNFR-/- mice was a reduction in the number of B
cells (B220+), consistent with the known role of TNF- in B cell
proliferation and differentiation (23, 24).
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To study whether the impairment in proliferation to alloantigen in
p55 TNFR-/- T cells was restricted to either
the CD4+ or CD8+ subsets,
splenic T cells were purified as detailed in Materials and
Methods and stimulated with alloantigen or CD3 and CD28. These
experiments demonstrated that proliferation and IL-2 production to
alloantigen were significantly impaired in both the
CD4+ and CD8+ subsets
whereas responses to mitogen were normal (Table III). Given that IL-2 was detectable in
culture supernatant from wild-type CD8 cells in MLC, the apparent
absence of IL-2 production from mitogen stimulated
wild-type CD8 cells is likely to reflect the sampling of
supernatant after autocrine consumption had exhausted IL-2 in the
culture medium. Surprisingly, the production of IFN- to mitogen was
dramatically increased in both the CD4+ and
CD8+ p55 TNFR-/- T cell
populations (6-fold and 100-fold, respectively). This increase was not
associated with changes in the production of the type 2 cytokine IL-4.
Enhanced proliferation to low concentrations of anti-CD3 was
observed in both CD4+ and
CD8+ p55 TNFR-/- cells
compared with wild-type cells (data not shown). Taken together, these
data show that CD4+ and
CD8+ T cells lacking the p55 TNF- exhibit
specific defects to allogeneic APC even though the TCR and CD28
signaling pathways are intact.
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enhances T cell proliferation and Th1 differentiation to
alloantigen
TNF- enhances T cell proliferation to both mitogenic
(25) and allogeneic stimuli (15),
independently of its effects on APC (15), and TNF- may
therefore function as an autocrine growth factor for T cells. Mitogenic
responses can be enhanced by p75 TNF- receptor agonists
(4), but the specific TNF- receptors involved in
responses to alloantigen have not been elucidated. To study the role of
TNF- in the defects seen in p55 TNFR-/- T
cells, splenic T cells from wild-type, p55
TNFR-/-, and p75
TNFR-/- animals were cultured in primary MLC
with a TNF- inhibitor (recombinant TNF- receptor linked to the
human Fc Ig, TNFR:Fc) or control human Ig. As shown in Fig. 2, the neutralization of TNF- in these
cultures impaired proliferation of wild-type T cells, p55
TNFR-/- T cells, and p75
TNFR-/- T cells. TNFR:Fc did not affect IL-2
production (data not shown) but inhibited IFN- production T cells
from all animals (Fig. 2). Importantly, the inhibition of TNF- in
these cultures did not reduce T cell proliferation or IFN-
production in wild-type T cells down to the levels of p55
TNFR-/- cells, suggesting a component of the
defect in the p55 TNFR-/- T cells was
independent of exogenous TNF-. As shown in Fig. 3, the defect in proliferation and
cytokine production to three allogeneic stimuli was similar, confirming
the general nature of the deficit. To investigate whether these defects
also affected T cell expansion and cytolytic function, respective
CD8+ cells were examined after primary MLC. The
addition of exogenous IL-2 to the primary MLC restored defective CTL
lysis by p55 TNFR-/- CD8+
cells but did not improve their poor expansion (Table IV). Interestingly, T cells from both old
(>24 wk) and young (<12 wk) p55 TNFR-/-
animals had impaired proliferation and T cell expansion in vitro. T
cells from old p55 TNFR-/- animals also had
marked impairment in cytokine production and cytotoxicity to
alloantigen, which was only mild in T cells from young p55
TNFR-/- animals. The reasons for the more
pronounced defect in T cells from older animals is not clear but is
likely to relate to a progressive T cell defect as a result of the
prolonged absence of TNF- signaling. In support of this we have also
noted a progressive age-related stem cell defect in p55
TNFR-/- animals (26).
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Given the defects of p55 TNFR-/- T cells
to alloantigens in vitro, we measured this response in vivo by
analyzing systemic IFN- levels after allogeneic BMT. All animals
received wild-type T cell-depleted bone marrow together with either
wild-type, p55 TNFR-/-, or p75
TNFR-/- T cells. No IFN- was detectable in
the sera of recipients of p55 TNFR-/- T cells 4
days after BMT, and it remained significantly depressed 7 days after
BMT (Fig. 4). We next examined donor T
cell expansion and function 2 wk after BMT, a time of maximal T cell
expansion in this model. The total number and percentage of splenic
CD4+ cells from p55
TNFR-/- donors were not different from
wild-type donors 14 days after BMT, although there was a significant
decrease in the number and percentage of CD8+
cells (Table V). By 2 wk after BMT, donor
p55 TNFR-/- T cells proliferated normally and
produced equivalent amounts of the type I cytokines IL-2 and IFN- to
wild-type cells in vitro (Table V). In view of the impaired
CD8+ T cell expansion, we next studied the
effector function of these cells in standard 51Cr
assays. As shown in Fig. 5, in two
separate experiments there was a consistent reduction in the capacity
of p55 TNFR-/- T cells to lyse host type P815
targets after allogeneic BMT.
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is important for priming mononuclear cells to secrete
TNF- after BMT, we were interested in whether the reduced production
of IFN- following transplantation of p55
TNFR-/- T cells would effect subsequent TNF-
production. As shown in Fig. 4
, the levels of TNF- 7 days after BMT
were 10- to 20-fold lower following transplantation of p55
TNFR-/- T cells than those seen in recipients
of wild-type or p75 TNFR-/- T cells. These data
confirm that p55 TNFR-/-
CD8+ T cells expand poorly and function
suboptimally in vivo, and demonstrate that T cells lacking the p55
TNF- receptor are unable to amplify the proinflammatory cytokine
cascade that is known to be critical in the pathophysiology of GVHD
(27). The reduced ability of p55 TNFR-/- cells to induce GVHD is due solely to defects in the T cell compartment
To study whether the reduction in GVHD seen after transplantation
of p55 TNFR-/- donor bone marrow and T cells
(Fig. 1) was due primarily to defects in donor T cell function, we
performed mixing studies. As shown in Fig. 6, survival was significantly lower in
recipients of p55 TNFR-/- bone marrow and
wild-type T cells than in those receiving p55
TNFR-/- bone marrow and p55
TNFR-/- T cells (50% vs 100%,
p < 0.01), confirming that p55
TNFR-/- mononuclear cells did not play a role
in reducing GVHD. Furthermore, the severity of GVHD in surviving
recipients of p55 TNFR-/- T cells was minimal
(clinical scores, <2), regardless of the origin of the bone marrow. By
contrast, recipients of wild-type T cells had moderately severe GVHD
(clinical scores, >3), regardless of the origin of the bone marrow.
Therefore the expression of the p55 TNF- receptor on donor T cells
was critical to the induction of acute GVHD, confirming the earlier in
vitro studies.
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Discussion |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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receptor have impaired responses to alloantigen, characterized by
reduced proliferation, type I cytokine production, and cytolytic
function, despite the normal response of these cells to mitogen and
CD28. In vivo, T cells lacking the p55 TNF- receptor have a reduced
capacity to induce GVHD, characterized by impaired IFN- and TNF-
production, reduced CD8+ expansion, impaired
cytolysis of host tissues, and milder systemic disease. The
interpretation of the effects of TNF- neutralization on donor T cell
function is complex because TNF- is known to induce cytokine
production by APCs (such as IL-12 (28)), which may effect
T cell responses indirectly. Indeed, we cannot exclude this as a
mechanism for the reduced proliferation and IFN- production by T
cells in the presence of TNF- inhibitors that we observed in vitro.
For these reasons the direct effects of TNF- on T cells can best be
studied by the use of T cells lacking appropriate TNF-
receptors.
Agonists of the p75 TNF- receptor are known to enhance T cell
proliferation in response to mitogen (29, 30). TNF- has
been shown to enhance proliferative responses in MLC (15),
probably through increased IL-2 expression (31). The role
of the p55 receptor during autocrine T cell stimulation is unclear
because the surface expression of p75 is dominant following T cell
activation (15, 32). However, the reduced apoptosis of p55
TNFR-/- CD8+ cells
following activation in vivo (33) and the synergistic
effect of both receptors on T lymphoblast apoptosis (34)
confirms a role for the p55 TNF- receptor on T cells. TNF- is
also known to play a role in the survival of activated T cells
(35, 36), although normal negative selection induced by
superantigen has been shown in p55- and p75-deficient mice (10, 17). The p55 receptor appears to control the deletion of
peripheral lymphocytes following their activation by peptide in vivo
(33). The prolonged persistence of activated p55
TNFR-/- T cells, associated with resistance to
activation-induced cell death (33), is consistent with the
increased numbers of memory T cells seen in the spleen of the p55
TNFR-/- animals in this study.
To our knowledge, abnormalities in the proliferative function of p55 TNFR-/- T cells have not been previously described. In this study we noted an increased sensitivity of p55 TNFR-/- T cells to activation by mitogen, which is consistent with the lower activation threshold of memory T cells compared with naive T cells (37, 38). The differences seen between responses to alloantigen and to mitogen may relate to differences in the intensity of TCR signaling or alternatively may reflect enhanced sensitivity to inhibitory signals from the APC itself. In support of this latter possibility, p55 TNFR-/- T cells also respond to mitogen suboptimally in the presence of syngeneic APCs (data not shown). Supernatants taken from primary MLC of p55 TNFR-/- T cells and allogeneic APC do not inhibit responses of naive wild-type or p55 TNFR-/- T cells to alloantigen, suggesting that this process requires cell-cell contact. Studies have shown that memory CD4+ T cells have an increased expression of CTLA-4 and impaired proliferative responses in the presence APC bearing B7 (39). However the proliferative defect in p55 TNFR-/- T cells was not rescued by CTLA-4 inhibition or simultaneous CD28 stimulation and B7 inhibition with CTLA-4Ig (data not shown). Further studies are in progress to delineate the mechanisms by which APCs inhibit the responses of T cells lacking the p55 receptor.
TNF- is also required for the optimal generation of human CTL in
vitro; when added to the sensitizing phase of the primary MLC, TNF-
causes a selective up-regulation of the IL-2R on CD8-positive T cells
(40). TNF- can also increase granzyme activity induced
by IL-2 as determined by benzyloxycarbonyl-L-lysine
thiobenzylester-esterase (BLT-E) activity (40). Our data
are consistent with the ability of TNF- to amplify CTL activity by
increasing IL-2 production. This study demonstrates that TNF-
signaling through the p55 receptor controls both the generation and the
function of CTL in vivo. In agreement with this observation, we and
others have recently confirmed that TNF- neutralization inhibits the
graft-vs-leukemia effect after allogeneic BMT (41, 42).
The current study suggests that TNF- may be required in the
differentiation of CTL from precursors rather than as an effector
molecule by mature CTL. We noted a more profound reduction in CTL
generation in vitro than in vivo, and we believe this is likely to
reflect the ability of other compensatory factors to support CTL
generation in the absence of TNF- in vivo. It is likely that IL-12
is important in this regard (43) because the production of
IL-12 in MLC after BMT was high and similar between animals
transplanted with wild-type or p55 TNFR-/- T
cells. Interestingly, preliminary data suggest that the addition of
exogenous IL-12 to primary MLC also improves the proliferative and
cytokine defects in p55 TNFR-/- T cells in
secondary cultures. This may, in part, explain the apparent equivalent
proliferation and cytokine production of p55
TNFR-/- T cells and wild-type T cells after BMT
(Table V). Unfortunately, interpretation of T cell function at this
time is complicated by the presence of moderately severe GVHD and its
associated immunosuppression which is absent in p55
TNFR-/- T cells. This impairment in function of
wild-type T cells in mice with GVHD will therefore mask differences in
relation to p55 TNFR-/- T cells in mice
without GVHD.
The role of TNF- as an effector of host cytotoxicity during acute
GVHD is well established (19, 44), and the presence of the
p55 TNF- receptor on recipient tissues has been shown to be critical
for the induction of GVHD (45). The present study confirms
that the p55 receptor plays a critical role in T cell responses to
alloantigens. The fact that donor p55 TNFR-/- T
cells failed to expand and produce IFN- in response to alloantigen
suggests that TNF- also alters T cell alloreactivity directly. The
reduced levels of TNF- produced in vivo despite a normal wild-type
bone marrow compartment confirm the importance of IFN- from donor T
cells in the priming of mononuclear cells to produce inflammatory
cytokines (46). This study delineates an additional
mechanism by which neutralization of TNF- early after allogeneic BMT
may reduce acute GVHD. The apparent requirement for TNF- in the
differentiation of cytotoxic T cells suggests that complete
neutralization of TNF- early after BMT may impair the
graft-vs-leukemia (GVL) effect and highlights the complexity of
inflammatory cytokine involvement in both GVHD and GVL.
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Footnotes |
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2 Address correspondence and reprint requests to Dr. James L. M. Ferrara, Departments of Internal Medicine and Pediatrics, Bone Marrow Transplant Program, University of Michigan Cancer Center, MI 48109-0560. E-mail address:
3 Abbreviations used in this paper: LT, lymphotoxin; GVHD, graft-vs-host disease; BMT, bone marrow transplant; TBI, total body irradiation.
Received for publication July 29, 1999. Accepted for publication November 1, 1999.
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References |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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-deficient mice: a critical requirement for TNF in the formation of primary B cell follicles, follicular dendritic cell networks and germinal centers, and in the maturation of the humoral immune response. J. Exp. Med. 184:1397. and the type I tumor necrosis factor (TNF) receptor in the establishment of follicular dendritic cells from non-marrow-derived cells. J. Exp. Med. 186:1997./ and tumor necrosis factor are required for stromal cell expression of homing chemokines in B and T cell areas of the spleen. J. Exp. Med. 189:403. and IL-1. J. Immunol. 151:5239.[Abstract]
in dendritic cell-mediated primary mixed leukocyte reactions. Bone Marrow Transplant. 15:163.[Medline]
: Autocrine and paracrine cytokines involved in B cell function. J. Immunol. 146:3462.[Abstract]
and interleukin-1. J. Immunol. 139:2970.[Abstract]
. J. Immunol. 140:531.[Abstract]
involvement in the optimal induction of class I allospecific cytotoxic T cells. J. Immunol. 144:4555.[Abstract]
on graft-versus-host disease and graft-versus-leukemia. J. Clin. Invest. 104:459.[Medline]
production to LPS stimulation by donor cells predicts the severity of experimental acute graft-versus-host disease. J. Clin. Invest. 102:1882.[Medline]
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) or TNFR:Fc (
) as detailed in Materials
and Methods. *, p < 0.02. Results
represent one of four experiments. The range of reductions (1 -
(TNFR:Fc/control) x 100) in proliferation from cultures with
TNFR:Fc were as follows: wild-type cells, 28–38%
(p < 0.02); p55 TNFR-/-, 39–95%
(p < 0.03); and p75 TNFR-/-,
0–51%. The range of reductions in IFN-
), or
p75 TNFR-/- (
) animals were cultured with
B6D2F1 (H2bxd), B6C3F1 (H2bxk), or B6SJF1
(H2bxs) peritoneal macrophages in MLC. Proliferation, IL-2
and IFN-
