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*
Institut National de la Santé et de la Recherche Médicale Unité 477, Université René Descartes, Paris, France;
Service de Chirurgie Digestive, Hôpital Henri Mondor, Université Paris XII, Creteil, France;
Laboratoire d’Immunologie, Hôpital Cochin, Université René Descartes, Paris, France;
Service d’anatomie et cytologie pathologiques, Centre Hospitalo-Universitaire de Bicêtre, Le Kremlin-Bicêtre, France; and
¶ Institut National de la Santé et de la Recherche Médicale Unité 257, Université René Descartes, Paris, France
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Abstract |
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 Top Abstract IntroductionMaterials and MethodsResultsDiscussionReferences |
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Introduction |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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cells (8).
These results were highly encouraging as regards using FasL as a mean
of inducing allograft acceptance. In contrast, it has been recently
reported by several groups that expression of FasL in pancreatic
islets, either as a result of retroviral transfection or transgenesis,
did not confer protection from allograft rejection. In fact, FasL
expression has been shown to target FasL transgenic (TG) islets for
rapid destruction (9, 10, 11). Moreover, the accelerated
rejection of allogeneic organs, which express FasL ectopically, has
hampered the investigation of the impact of FasL in alloimmune-specific
responses in such a setting. Furthermore, it has been reported that
hearts transplanted into syngeneic and allogeneic recipients, from FasL
TG mice expressing FasL in heart muscle cells, were more rapidly
rejected than nontransgenic (NTG) hearts (12). Thus to
date, contradictory effects of TG FasL expression in tissue allograft
rejection have been reported. We have investigated the ability of ectopically expressed FasL on thyroid follicular cells (TFC) to interfere with thyroid allograft rejection by grafting a complete thyroid lobe under the kidney capsule in fully allogeneic recipients. The allograft transplantation thyroid model has numerous advantages, including Fas-mediated death resistance in the TFC (13, 14), and the fact that the particular follicular structure of the thyroid makes it easier to distinguish between donor graft and recipient tissue. We also studied the allogeneic immune response both in vitro and in vivo. In the present work, three TG lines, expressing different levels of FasL, were investigated to evaluate the role of this parameter in allograft experimentation. These mice expressed both the soluble and membrane-bound form of FasL. Our results show that FasL expression confers an immune privilege upon thyroid tissue, and they provide experimental evidence for the underlying mechanisms.
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Materials and Methods |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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C57BL/6 and CBA/J male mice were purchased from Iffa Credo (L’Arbresle, France). C57BL/6 lpr/lpr mice were provided by Dr. E. Schneider (Institut National de la Santé et de la Recherche Médicale Unité 25, Paris, France). Male mice between 7 and 10 wk of age were used in all experiments. Animals were maintained in standard environmental conditions, with free access to food and water. They were allowed to adapt to their environment for 1 wk before the experiments were initiated. pTg-FasL TG mice have been described elsewhere (15). Three selected transgene-positive founders were crossed with CBA/J mice. TG H-2k mice (I-Ak and Kk positive, I-Ab and Kb negative) were then crossed four times with CBA/J mice. Mice were screened for pTg-FasL transgenes by PCR analysis of tail DNA using the following primers: pTg sense, 5'-GCCTCCACAAGATTTTCACC-3'; and FasL anti-sense, 5'-TGGTAGTGGTGATGGAGGTG-3'. A 450-bp PCR product was obtained for TG mice.
Western blotting
Total protein was extracted from FasL TG and control thyroids in RIPA buffer. Protein samples were filtered with Kwikspin 100 (Pierce, Rockford, IL) to remove proteins under 100 kDa, and 25 µg were electrophoresed in 12% polyacrylamide gel and blotted onto a nitrocellulose membrane. The membrane was incubated with blocking buffer (TBS, 5%BSA), hybridized with 1 µg/ml anti-mouse FasL (clone H11 manufactured by Alexis Biochemicals; a gift from J. Tschopp), and washed. Signals were detected using an ECL Western blotting detection system (Amersham, Little Chalfont, U.K.).
Antibodies
The following biotinylated Abs were used in this study: anti-mouse CD4 (rat IgG2a; CT-CD4); B220 (rat IgG2a; RA3-6B2); Mac-1 (rat IgG2b; M1/70-15); and control (rat IgG2a; LO-DNP-16 and rat IgG2b; LO-DNP-57) (Caltag, South San Francisco, CA). Anti-mouse Thy 1.2 (rat IgG2a; 53-2.1), CD8 (rat IgG2a; 53-6.7), Gr-1 (rat IgG2b; RB6-8C5), MHC II I-Ak (mouse IgG2b; 11-5.2), CD11c (hamster IgG; HL-3), and control (mouse IgG2b; 49.2) were all obtained from BD PharMingen (San Diego, CA). For depletion of CD4 or CD8 subsets, a mixture of nonbiotinylated anti-mouse CD4 (GK 1-5 and RM 4-5) or a mixture of nonbiotinylated anti-mouse CD8 (53-6.7 and 3.168) was given as three 2-mg doses on days 3 and 1 before grafting, and day 3 postgrafting. Control groups received PBS.
Thyroid graft transplantation, determination of graft survival, and histopathological studies of grafted specimens
A thyroid lobe was transplanted under the kidney capsule of recipient mice in accordance with the method described by Lafferty et al. (16). Graft survival was evaluated from days 7 to 42 postgrafting. The graft-bearing kidneys were immersed in paraformaldehyde solution and embedded in paraffin, and the 3-µm-thick serial sections taken were stained with hematoxylin and eosin. Sections were coded and the lesions (cell infiltration and graft rejection) were scored by two independent observers.
Immunohistochemistry
The graft-bearing kidneys were immediately covered in optimal temperature medium (Tissue-Tek; Bayer, Elkhart, IN), slowly frozen by floating in isopentane on liquid nitrogen, and stored at -80°C until cut. Sections of 5–7 µm were cut on a cryostat at -18°C and collected onto SuperFrost-plus slides (Roth Sochiel, Lauterbourg, France). Sections were dried overnight and stored at -80°C until use. Before staining, sections were fixed for 10 min in acetone and incubated for 20 min in 0.05 M TBS, pH 7.6, in the presence of 10% normal horse serum, and stained (45 min) with the appropriate biotin-conjugated primary Ab. Avidin-alkaline phosphatase (Sigma, St. Louis, MO) second stage (30 min), and FastRed substrate (Acros Organics, Noisy-Le-Grand, France) were used to visualize specific staining. Sections were counterstained in hemalum (Fisher Scientific, Pittsburgh, PA) and mounted in an aqueous mount (Aquatex; Merck, West Point, PA).
In vitro cytotoxic responses to allogeneic cells
On the postgraft day indicated, spleen cells from recipient mice
were prepared at 1.5 x 106/ml in RPMI 1640
medium with 10% FCS. Cells were cultured with irradiated spleen cells
(7.5 x 106/ml) in 100-mm petri dishes, and
1 ng/ml of recombinant IL-2 was added on days 0, 2, and 5. At the end
of the culture period (day 6), cells were harvested, purified through a
Ficoll-Hypaque, washed twice in HBSS, and were thereafter referred to
as effector cells. The BW 5147 thymoma cell line (MHC class I positive,
class II negative) or peritoneal macrophages were used as target cells
and labeled with 50 µCi of 51Cr per
106 cells. After a 1-h incubation period at
37°C, with shaking, cells were washed twice in HBSS, and 5 x
103 or 104 target cells
were placed into each well of flat-bottom 96-well plates (no. 3799;
Costar, Cambridge, MA). Thereafter, 100 µl of effector cells at 2.5,
5, 10, and 20 x 105 cells/ml were
added. After 5- and 12-h incubations for BW 5147 and macrophages,
respectively, 100 µl of supernatant was collected and chromium
release measured with a 
-scintillation counter (1450 Microbeta plus;
Wallac, Gaithersburg, MD). Spontaneous and maximal releases were
defined by the incubation of target cells with culture medium in the
absence or presence of 10% HCl, respectively. Spontaneous release was
<21% for BW 5147 and <30% for macrophages. Specific lysis (%) was
calculated as (experimental 51Cr release -
spontaneous 51Cr release)/(maximal
51Cr release - spontaneous
51Cr release).
In vitro proliferative and cytokine responses to allogeneic cells
In vitro proliferative responses to alloantigen were measured by
culturing 2 x 105 recipient spleen cells
with 2 x 105 irradiated CBA spleen cells
for 72 h. Cells were pulsed with 0.5 µCi of
[3H]thymidine for the final 12 h, and then
harvested for liquid scintillation counting. Culture supernatants were
collected on days 1–3 for IL-2 assay using the IL-2-dependent CTLL-2
cell line, and IFN- assay using a two-site ELISA with R46-A2 mAb as
the coating Ab, and -galactosidase-coupled XMG1-2 mAb as the
developing Ab. The detection limit for IFN- was 100 pg/ml.
Levels and isotypes of Abs to allogeneic cells
Mice were bled either by retro-orbital puncture or by cardiac puncture at the time of sacrifice. Serum samples were stored at -20°C until use. Allogeneic specific Abs were analyzed by FACS. Target cells (BW 5147) were incubated with individual mouse serum at a 1/20 dilution in FACS medium (PBS, 2% FCS, 0.01% NaN3). After washings with FACS medium, secondary staining was performed using FITC-conjugated anti-mouse IgM Ab (Caltag) or anti-mouse IgG, IgG1, IgG2a, IgG2b, and IgG3 Abs (Southern Biotechnology Associates, Birmingham, AL). The cells were washed and analyzed by FACS (EPICS; Coulter, Hialeah, FL).
Statistical analysis
Statistical analysis was performed using a two tailed Student’s t test (p < 0.05).
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Results |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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pTg-FasL TG mice.
We have previously reported the generation of TG mice expressing
functional FasL cDNA under the control of the bovine thyroglobulin
promoter (pTg-FasL mice) (15). Western blotting analysis
confirmed TG FasL expression and showed that TG thyroid expressed both
the soluble and membrane-bound FasL (Fig. 1
). The levels of expression of both
forms correlated. Among the numerous Abs against FasL that have been
tested (N-20 and C-178 from Santa Cruz Biotechnology, Santa Cruz, CA;
clone 33 from Transduction Laboratories, Lexington, KY; AB1665 from
Chemicon International, Temecula, CA; and clone H11 from Alexis
Biochemicals, Paris, France), only the clone H11 gave
reproducible results. The three different lines were named
TG6low, TG9med, and
TG11high in accordance with the level of
functional and protein FasL expressed.
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and Fig. 2. Syngeneic grafts from NTG donors were
recovered at various times postgraft, from days 7 to 42, and
demonstrated a 100% acceptance rate. On day 7 postgraft, NTG and
TG6low grafts were found to be massively
infiltrated, and large numbers of follicles were already destroyed
(Table I). In those cases of severe rejection, inflammation extended
throughout the graft. The infiltration was confluent, containing
numerous activated and blastic lymphocytes, some macrophages, and only
very few granulocytes. The majority of follicles were completely
destroyed. Others were damaged, showing vacuolization, breaks in the
basement membrane, and necrosis. By contrast, in
TG9med and TG11high grafts
there were no signs of rejection, and only a mild lymphocytic
infiltration was observed. On day 14, NTG and
TG6low grafts had been completely rejected,
whereas those from TG9med and
TG11high mice were still protected (Fig. 2). At
this time, the lymphocytic infiltrate was generally confined within the
margin of the graft. Follicular inflammation mostly involved apoptotic
lymphocytes. This result was confirmed by in situ TUNEL analysis (data
not shown). A minority of follicles was cuffed and showed only mild
reactive changes (Table I, and Fig. 2, C and D).
By day 21, TG9med grafts had been completely
rejected. TG11high grafts were mainly intact up
to 42 days postgraft. By contrast, TG11high
grafts were rejected by day 17 when transplanted under the kidney
capsule of lpr/lpr mice, further demonstrating the
importance of FasL in allograft survival (Table I). Thus, expression of
FasL on thyroid cells inhibited acute allograft rejection, in a
dose-dependent manner.
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). CD4+ T cells
and a few B lymphocytes were also present. TG9med
and TG11high grafts showed a smaller number of
infiltrating cells (up to 5 times fewer than NTG grafts) with a major
reduction in the CD8+ T cell subset. In every
case, no or only a few granulocytes were detected. Thus, TG FasL
expression resulted in a strong reduction of all infiltrating cell
subpopulations, which was particularly obvious for
CD8+ T cells.
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for the CD4 T cell-depleted hosts
demonstrate for both TG and NTG thyroid recipients the persistence of
thyroid follicles at the graft site, although all grafts were
undergoing a rejection process. When compared with the full rejection
of NTG thyroid observed in untreated recipients on day 14 (Table I
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CD4 T cell depletion resulted in a delay of graft rejection. By
contrast, CD4 T cell depletion abolished the protective effect of TG
FasL expression.
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Effect of FasL on allogeneic cell-mediated immunity
Consequences of TG thyroid graft on the allospecific, cytotoxic T
cell response.
Allospecific, cytotoxic T cells appear as important effectors in
thyroid allograft rejection (Tables II and III). Therefore, we examined
the development of allogeneic, cytotoxic T cell responses in recipient
animals. A splenic, cytotoxic T cell response in C57BL/6 grafted mice
toward H-2k allogeneic cells was measured on day
14 postgraft, when thyroids of CBA/J and TG6low
were rejected whereas those of TG9med and
TG11high were still accepted. To minimize
variability between different experiments, all animals were grafted at
the same time.
Cytotoxic T cells were always detected among splenocytes from all
groups, and TG6low-immunized mice had a similar
level of cytotoxicity as mice grafted with CBA thyroid (Fig. 3A). Conversely, high
expression of functional FasL (TG11high) induced
a lower allospecific, cytotoxic T cell response compared with NTG mice
(Fig. 3A). Intermediate expression of FasL by
TG9med TFC resulted in a less pronounced effect
(Fig. 3A). The role of Fas/FasL interaction in the
allogeneic, cytotoxic response was further demonstrated by similar
levels of cytotoxic T cells among splenocytes of lpr/lpr
mice grafted with TG or NTG thyroid (data not shown).
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The measurement of the frequency of anti-H-2k
CTL at day 14 postgraft by limiting dilution analysis showed a 3- to
5-fold reduction, in that the frequency of reactive CTL was reduced by
3–5 times in C57BL/6 mice grafted with TG11high
thyroid compared with mice grafted with NTG thyroid (Fig. 3B). Furthermore, the frequency of the IFN--producing
cells was also markedly reduced in mice grafted with TG compared with
NTG thyroid (data not shown). A decreased number of CTL- and
IFN--producing cells was observed in TG6low
and TG9med thyroid-grafted animals, although this
decrease was not statistically significant. Thus, TG FasL expression in
the grafted thyroid inhibited the development of alloreactive,
cytotoxic T cells.
Consequences of CD4 and CD8 T cell depletion on allocytotoxic and
alloproliferative T cell responses following TG and NTG grafting.
Experiments were performed to gain further insight into the respective
roles of CD4 and CD8 T cell responses in thyroid allografts. Mice were
depleted of CD4 or CD8 T cells before transplantation with CBA/J or
TG11high thyroid, and the development of the
alloproliferative response was evaluated. Without depletion, the level
of alloresponse was lower in TG compared with NTG graft recipient
animals (p < 0.02) (Fig. 4, left). The lower
alloreactive T cell proliferation observed in control mice grafted with
TG thyroid could be totally restored by adding IL-2 to the culture
(Fig. 4, right). MLR proliferation of spleen cells from NTG
and TG mice was similar when tested in the presence of
non-H-2k cells, and no differences in
proliferation and IL-2 production were noted between NTG and TG spleen
cells stimulated with Con A (data not shown).
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, right). Conversely,
proliferation of CD8-depleted T cells was still completely abolished
despite the presence of IL-2. Hence, the allospecific CD8 T cells
represent the major proliferative T cell population in our model.
We also investigated whether depletion of the CD4 or CD8 T cell
population affected the cytotoxic alloresponse in the grafted animals.
Because IL-2 was necessary for T cell proliferation (Fig. 4, right), it was added to the in vitro T cell allogeneic
stimulation experiment. Allocytotoxicity was almost completely
abolished by CD8 T cell depletion in all grafted recipients (data not
shown). Conversely, CD4 T cell depletion did not affect the cytotoxic,
allogeneic response in TG and NTG thyroid-grafted recipients (data not
shown). These results showed that, in the absence of
CD4+ T cells, but in the presence of IL-2, a
cytotoxic T cell response, which is not affected by thyroid FasL
expression, can occur.
Anti-allogeneic, humoral-mediated immunity following TG and NTG grafting
Inhibition of the early alloantibody response in TG FasL
thyroid-grafted animals.
The presence of alloantibody reactive to H-2k
cells was analyzed in the serum of grafted animals at different times
postgraft (Fig. 5). Mice grafted with
control thyroid developed an alloantibody response characterized by the
presence of IgM and IgG on days 14 and 21. In the case of
TG6low and TG9med grafts,
IgM alloantibodies were transiently detectable on day 14, whereas IgG
alloantibodies were detectable on days 14 and 21. In the case of
TG11high graft, no IgM alloantibodies were
detected. Before day 14, no IgG alloantibody response could be detected
in 7 of 8 TG11high thyroid recipient mice.
Furthermore, in this group of animals, analysis of the alloantibody
response at a later time demonstrated a detectable IgG response in only
11 of the 16 animals analyzed. The analysis of the response in
lpr/lpr grafted mice showed that no difference in IgM and
IgG Ab responses occurred in NTG or TG grafted animals (data not
shown), confirming that the inhibitory effect was mediated through
Fas/FasL interaction. Thus, the FasL expression on grafted organs
resulted either in a complete inhibition or a delay in the development
of the alloantibody response.
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. Among the grafted mice
developing alloantibodies, only 4 of the 13 mice grafted with control
thyroid developed IgG1 alloantibodies, compared with 9 of 11
(p < 0.02) animals grafted with
TG11high thyroid. Conversely, mice grafted with
control thyroid developed IgG2a Ab almost twice as frequently as mice
grafted with TG11high thyroid, and the IgG3
response was observed significantly more frequently
(p < 0.007) in TG11high
compared with control-grafted animals. Thus, when it did not induce a
complete abrogation of the alloantibody response, FasL expression on
the grafted organ resulted in a switch toward a T2-Ab response.
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). Conversely, the
alloantibody response was strongly increased following CD8 T cell
depletion (Fig. 6). In those animals, an isotypic switch toward a T2 Ab
response was still observed in TG11high- compared
with NTG-grafted recipients (data not shown).
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Discussion |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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When TG thyroid was used in allograft experiments, we found that
increased allograft survival was proportional to the level of FasL
expression on transplanted TFC. The involvement of the Fas/FasL
interaction in these findings is confirmed by the rejection of
TG11high grafts in Fas-deficient mice. Several
groups have reported that high FasL expression on pancreatic islet
cells resulted in a more rapid rejection of the islet graft compared
with islets with low or no expression (10, 11). One
explanation could rely on the Fas-induced cell death sensitivity of the
grafted cells. In fact, pancreatic cells have been shown to be
highly susceptible to Fas-mediated death, both in humans
(17) and mice (9), whereas TFC have been
shown to be naturally resistant to this form of cell death (14, 18, 19). The spontaneous apoptosis of the grafted Fas-sensitive
cells, but not thyrocytes, could explain the contrasting outcome of
the grafting of these two endocrine glands, namely thyroid and
pancreas, which both ectopically express FasL. In particular, future
studies should focus on regulation of the neutrophilic infiltration
occurring in FasL-pancreas but not in FasL-thyroid allografts. At least
two explanations have been proposed to explain the discrepancies
between the results of studies using FasL-based immunotherapy in
allografting. One explanation involves the suppressive activity of the
inflammatory response of soluble FasL. Indeed, it has recently been
demonstrated that the natural cleavage product of murine FasL was
unable to induce neutrophil inflammation (20), as opposed
to the previously held view of its neutrophil chemotactic activity
(21, 22). This would imply that eye, testis, and thyroid
might produce more soluble FasL than pancreas and heart tissues, which
would result in an inhibition of neutrophil attraction. It would be of
particular interest to determine whether different expressions of
particular metalloprotease activity able to cleave membrane FasL exist
in these tissues.
The second explanation relies on the possible different levels of IL-8-induced expression in the various transplanted tissues. Because of the well known IL-8 neutrophil chemotaxis activity, different levels of IL-8 expression could form part of the explanation. Production of other chemokines with neutrophil chemotaxis activity could also explain such results. The pro- or anti-inflammatory consequences of FasL expression might certainly be influenced by local environmental factors that vary from tissue to tissue. It is highly probable that FasL expression might not be sufficient in all transplantation settings to ensure graft survival. For example, corneas grafted heterotopically to the skin are rapidly rejected (23). The results reported herein show that FasL expression confers immune privilege status upon thyroid, providing protection against allograft rejection, a property that is probably enhanced by the kidney capsule microenvironment.
The induction of graft acceptance certainly depends on the ability to reduce the alloimmune response, particularly in a fully MHC-mismatched combination as in our experiments. Rejection of NTG, allogeneic grafts in CD4 T cell-depleted mice was delayed by up to 20 days after grafting, and two of the six NTG, allogeneic grafts in CD8 T cell-depleted mice were not rejected. These results show that both CD4 and CD8 T cells were involved in the rejection process. Depletion of CD8 T cells did not significantly modify rejection of TG grafts, whereas CD4 T cell depletion induced a severe rejection process involving all TG grafts. These results suggest that FasL-induced protection was mainly mediated by CD4 T cells.
We then analyzed the consequences of thyroid-FasL expression on
alloreactive CD4 and CD8 T cell responses. Analysis of the
allospecific, cytotoxic T cell responses following grafting
demonstrated that TG grafts induced a lower response compared with
animals grafted with NTG thyroid. The systemic elimination of
alloreactive CD8 T cells by TFC-FasL could explain these results.
Nevertheless, we did not observe any significant difference in the
alloreactive, cytotoxic response after in vivo depletion of CD4 T
cells, arguing against a direct, dominant effect of TFC-FasL on
alloreactive, CD8 cytotoxic T cells. Furthermore, alloreactive CD8 T
cells have been demonstrated to be relatively resistant to Fas-induced
cell death (24, 25, 26). A second possibility might be a
change in the help necessary to develop a full alloreactive, cytotoxic
response following the grafting of FasL thyroid. Indeed, no significant
differences in the cytotoxic response (as mentioned above) or in
IFN- production between hosts receiving NTG or TG thyroid were
observed after in vivo depletion of CD4 T cells. Moreover, adding IL-2
to the culture completely restored the deficient CD8 T
cell-proliferative response in mice grafted with TG thyroid, further
showing that the CD8 T cells had not been eliminated. We concluded
that, in our experiments, the effect of TFC FasL expression on CD4 T
cells was probably directly responsible for the inhibition of the
alloreactive, cytotoxic T cell response in mice grafted with FasL TG
thyroid. Because Th1 cells are a population of CD4 T cell that are
known to promote the development of IFN--producing cytotoxic T
lymphocytes, our results suggest their inhibition in the host by
FasL.
Analysis of humoral responses demonstrated that TFC-FasL expression resulted in an alteration of the anti-H-2k-allospecific B cell response. This resulted either in a highly significant diminution of all classes of the anti-H-2k Abs or a specific diminution of the Th1-associated class of Abs. In C57BL/6 lpr/lpr mice, FasL did not alter the alloantibody response, further confirming that a FasL interaction is required for these effects. Moreover, this result was dependent on the level of FasL expression on the TFC. Mice grafted with NTG, TG6low, or TG9med thyroid developed very similar alloantibody responses, whereas mice grafted with TG11high preferentially developed a T2 type Ab response. These data could be explained either by the preferential survival of Th2 cells or by the inhibition of Th1 cells in TG11high graft recipients. Indeed, it has been reported that activation-induced cell death in CD4 Th cells could be responsible for imbalances in Th1 and Th2 subsets because Th1, but not Th2, undergo rapid Fas/FasL-mediated apoptosis (27, 28, 29). Moreover, during the Th2 response, B cells are protected from Fas/FasL-induced apoptosis, and IgG1 production is elicited (30). Because we have shown that the B cell response was totally abrogated in the absence of CD4 T cells, these results further demonstrate the shift of the T cell response away from a Th1 type. Nevertheless, we cannot formally rule out a direct intragraft selection of the ongoing B cell response, particularly during the early response when alloantibodies were undetectable.
The exact mechanism by which FasL expression on the transplanted tissue leads to control of the CD8 T cell-alloreactive effector population is presently unclear. The apoptosis of thyroid-infiltrating T cells that we observed could be necessary to eliminate Th1 cells and/or to create local immunoregulation, at least partially through the production of immunoregulatory cytokines, as shown in other models (31, 32, 33, 34, 35, 36, 37). Recently, the control of CD8 T cells in a tolerant state by a CD4 T cell-mediated mechanism has been shown to occur in cancer (38) and in autoimmunity (39). To determine whether this also occurs in our graft model, and how to tie together these complementary mechanisms, is the next challenge.
In conclusion, our findings indicate that the ectopic expression of FasL in a Fas-induced death-resistant tissue may induce tolerance to this tissue following allogeneic transplantation. Our results show that the level of FasL expression is of crucial importance in the success, or otherwise, of the graft. In addition, we have provided evidence that the protection provided by FasL expression in the grafted tissue might be explained by a shift away from a T1 toward a T2 alloreactive response, and through a mechanism involving allospecific CD8 T cell tolerance mediated by a CD4 T cell population.
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Acknowledgments |
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Footnotes |
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2 Address correspondence and reprint requests to Dr. Gilles Chiocchia, Institut National de la Santé et de la Recherche Médicale Unité 477, Hôpital Cochin, 27 ruedu Faubourg Saint-Jacques, 75674 Paris Cedex 14, France. E-mail address: chiocchia{at}cochin.inserm.fr
3 Abbreviations used in this paper: FasL, Fas ligand; TFC, thyroid follicular cell; TG, transgenic; NTG, nontransgenic.
Received for publication February 5, 2001. Accepted for publication May 30, 2001.
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References |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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cells induces a granulocytic infiltration but does not confer immune privilege upon islet allografts. Proc. Natl. Acad. Sci. USA 94:3943. cells for Fas-mediated destruction in insulin-dependent diabetes mellitus. J. Exp. Med. 186:1193., PGE2, and PAF. J. Clin. Invest. 101:890.[Medline]
in macrophages that have ingested apoptotic cells. J. Immunol. 163:6164.This article has been cited by other articles:
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K. A. Whartenby, E. E. Straley, H. Kim, F. Racke, V. Tanavde, K. S. Gorski, L. Cheng, D. M. Pardoll, and C. I. Civin Transduction of donor hematopoietic stem-progenitor cells with Fas ligand enhanced short-term engraftment in a murine model of allogeneic bone marrow transplantation Blood, October 16, 2002; 100(9): 3147 - 3154. [Abstract] [Full Text] [PDF]
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K. J. Young, L. Yang, M. J. Phillips, and L. Zhang Donor-lymphocyte infusion induces transplantation tolerance by activating systemic and graft-infiltrating double-negative regulatory T cells Blood, October 16, 2002; 100(9): 3408 - 3414. [Abstract] [Full Text] [PDF]
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B. Zhu, L. Luo, Y. Chen, D. W. Paty, and M. S. Cynader Intrathecal Fas Ligand Infusion Strengthens Immunoprivilege of Central Nervous System and Suppresses Experimental Autoimmune Encephalomyelitis J. Immunol., August 1, 2002; 169(3): 1561 - 1569. [Abstract] [Full Text] [PDF]
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Y. Wei, K. Chen, G. C. Sharp, H. Yagita, and H. Braley-Mullen Expression and Regulation of Fas and Fas Ligand on Thyrocytes and Infiltrating Cells During Induction and Resolution of Granulomatous Experimental Autoimmune Thyroiditis J. Immunol., December 1, 2001; 167(11): 6678 - 6686. [Abstract] [Full Text] [PDF]
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