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*
Nuffield Department of Surgery, University of Oxford, John Radcliffe Hospital, Headington, Oxford, United Kingdom;
Department of Surgery, Emory University School of Medicine, Atlanta, GA 30322;
Bristol-Myers Squibb Pharmaceutical Research Institute, Seattle, WA 98121;
§
Immunex Inc., Seattle, WA 98101; and
¶
Children’s Hospital, Boston, MA 02115
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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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B7-1 (CD80), B7-2 (CD86), and CD40 are important costimulatory molecules expressed on APCs (6, 7, 8, 9, 10, 11). Binding of B7-1 and B7-2 to their counterreceptor, CD28, on resting T cells increases lymphokine production, promotes T cell expansion, and thereby prevents the induction of T cell anergy (6, 12, 13, 14). The capacity of APCs to induce T cell activation may depend on the relative level of expression, as well as the density of these costimulatory molecules. Some professional APCs have been demonstrated to express high densities of B7-1 and B7-2 molecules (15, 16, 17). In contrast, resting B cells (rB cells)6 possess Ag-specific Ig receptors and MHC class II molecules, but do not express B7-1 or B7-2 (18, 19). It has therefore been proposed that rB cells are incompetent or nonprofessional APCs (20, 21).
It is clear that rB cells are ineffective stimulators of T cell proliferation in vitro and can render peptide specific T cell clones unresponsive to subsequent antigenic challenge in vitro (13). Moreover, when T cells encounter rB cells it has been shown that this can result in the production of Th2 cytokines (18, 22, 23, 24, 25). These data lead to the suggestion that rB cells might also be potent tolerogens in vivo. This hypothesis has been confirmed for tolerance induction to minor histocompatibility (miH) Ags, as demonstrated by the indefinite survival of skin (26) and cardiac allografts mismatched for H-Y Ag (27). The immune response to MHC Ag may be more complex and the ability and effectiveness of rB cells to switch off the response to these Ags is less clear. With the exception of a single report (28), rB cells have not been shown to be capable of inducing unresponsiveness to MHC Ags (27, 29).
The CD40 pathway plays an important role in B cell activation, including proliferation and Ig isotype switching (30, 31, 32). CD40 is a member of the TNF receptor superfamily and is expressed on B cells, endothelial cells, macrophages, dendritic cells, T cells, and fibroblasts (33, 34, 35). The ligand for CD40, CD40L (also known as gp39 or CD154) is expressed on activated, but not resting, T cells (33, 36). Buhlmann et al. (37) showed that in vivo administration of a mAb specific for CD40L, MR1, combined with allogeneic B cells (mismatched for MHC and miH Ags) diminished the response to alloantigen in vitro. In addition, Parker et al. (29) have also shown that pretreatment with donor small lymphocytes in combination with MR1 prolonged the survival of fully allogeneic pancreatic islets. The recent finding that immunization with rB cells from CD40 knockout mice induce tolerance to alloantigen as determined by the mixed lymphocyte reaction and the cytoxic T cell assay corroborates these findings (38). The latter study also provides evidence that the critical role of the CD40 pathway in this model may be via its role in the up-regulation of B7 expression on the APC.
The aim of the current study was to investigate the role of the CD40 and B7/CD28 costimulatory pathways in the induction of hyporesponsiveness to alloantigen by rB cells in vivo. The experiments are designed to specifically test the hypothesis that pretreatment with a combination of CD40 pathway blockade and rB cells will prolong allograft survival. Furthermore, we hypothesize that the effect of this treatment on allograft survival will be associated with an inhibition of B7 molecule expression on the donor cells. Thus, donor cells will present Ag to the recipient immune system in the absence of an effective costimulatory signal and thereby induce a state of donor-specific hyporesponsiveness to a subsequent allograft.
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Materials and Methods |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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C3H/He (H2k), CBA.Ca (H2k), C57BL/6 (H2b), C57BL/10 (H2b), and BALB/c (H2d) mice were purchased from Harlan (Bicester, U.K.) or The Jackson Laboratory (Bar Harbor, ME). All mice were housed in conventional facilities of the Biomedical Services Unit in John Radcliffe Hospital (Oxford, U.K.) or Emory University (Atlanta, GA) and used between 8 and 12 wk old in accordance with the Animals (Scientific Procedures) Act 1986.
The CD40 knockout mice (H2b) and the CD40L knockout mice (H2b) have been previously described (39, 40). The (C57BL/6 x 129)F1 (H2b) mice used as wild-type controls for the CD40 and CD40L knockout mice were purchased from The Jackson Laboratory.
Preparation of rB cells
Cell preparation was performed in PBS supplemented with 10% heat-inactivated FCS (Life Technologies, Paisley, U.K.). Splenocytes were depleted of erythrocytes with ammonium chloride buffer and of T cells with Abs to CD4 (YTS191) and CD8 (YTS169) (the hybridomas were kindly provided by Prof. H. Waldmann, Oxford, U.K.). Rabbit serum was used as a source of complement (Oxford Transplant Centre, Churchill Hospital, Oxford, U.K.). The remaining cells were passed over two sequential Sephadex G-10 columns (Pharmacia Biotech, Uppsala, Sweden) and the eluted cells were centrifuged over a discontinuous Percoll gradient (Pharmacia Biotech). Cells at the 60–70% interface were characterized and used as rB cells. In all cases, cells were washed three times before use. Typical preparation of rB cells as analyzed by flow cytometry was found to be 95.7% MHC class II+, 1.2% B7-1+, 2.5% B7-2+, and 92.3% surface Ig+.
Preparation of LPS-activated (aB) cells
For preparation of aB cells, the resting population (1 x 106 cells/ml) was cultured with LPS (30 µg/ml) (Escherichia coli O55:B5, Sigma-Aldrich, Poole, U.K.) for 3 days in complete RPMI 1640; RPMI 1640 (Life Technologies) was supplemented with 10% heat-inactivated FCS, penicillin G (75 U/ml), streptomycin (45 mg/ml), kanamycin sulfate (90 mg/ml), glutamine (2 mM), and 5 x 10-5 M 2-ME (Sigma-Aldrich). Typical preparation of aB cells as analyzed by flow cytometry was found to be 96.1% MHC class II+, 49.5% B7-1+, 77.7% B7-2+, and 96.9% sIg+.
Preparation of low density adherent cells (LODACs)
LODACs were prepared as described by Nussenzweig and Steinman (41). Splenocytes were centrifuged over "dense" BSA. The interface was collected and layered onto a plastic dish (1 x 108 cells/dish; Falcon 3003, Lincoln Park, NJ) in 10 ml of complete RPMI 1640. After incubating the plate for 90 min at 37°C, nonadherent cells were removed, the plates rinsed, and the medium replaced. After incubation for another 16 h at 37°C, nonadherent cells were collected and used as LODACs after washing. Typical preparation as analyzed by flow cytometry was found to be >70% CD11c+, 92.2% MHC class II+, 68.8% B7-1+, 88.2% B7-2+, and 9.4% sIg+. Therefore, LODACs were used as a dendritic cell-enriched population.
Flow cytometry
A total of 25 µl of viable cells (107 cells/ml) were incubated for 30 min on ice with primary Abs in PBS supplemented with 2% FCS and 0.02% sodium azide. If second-stage Abs were used, cells were washed twice before being incubated for a further 30 min on ice with a FITC-labeled goat anti-rat Ig mAb (F-6258, Sigma-Aldrich) blocked with 10% mouse serum. If biotinylated Ab was used in the first incubation, Streptavidin-phycoerythrin (PE) (7100-09; Southern Biotechnology Associates, Birmingham, AL), was used for the second incubation. After two more washes, the fluorescence intensities of 10,000 cells were determined using a FACSort (Becton Dickinson, San Jose, CA) flow cytometer. The primary Abs used in this study were as follows: anti-B7-1 FITC (1G10) rat IgG2a (01944D, PharMingen, San Diego, CA) (42); anti-B7-2 FITC (GL1) rat IgG2a (09274D, PharMingen) (8, 43); anti-IA(b,d) FITC (B21.2) rat IgG2b (hybridoma obtained from American Type Culture Collection, Manassas, VA); goat anti-mouse Ig FITC (F-0257, Sigma-Aldrich); anti-CD3 (KT3) rat IgG2a (kindly provided by Dr. K. Tomanari, Fukiu, Japan); anti-CD11c (N418) hamster IgG (kindly provided by Dr. J. M. Austyn, Oxford, U.K.) (44). Secondary Abs utilized included: goat anti-rat IgG FITC (F-6258, Sigma-Aldrich); goat anti-rat IgG PE (B-7139, Sigma-Aldrich); goat anti-hamster IgG biotinylated (BA-9100, Vector, U.K.).
MLC
T cells prepared by nylon wool purification were used as responders (2.5 x 105 cells/well). Stimulator cells were irradiated with 3000 rad before use. Cells were cultured in 200 µl of complete RPMI 1640 for 4 days. [3H]Thymidine was added for the last 16 h of the incubation. Cultures were harvested onto glass fiber filter mats (Wallac, Turku, Finland) and counted for thymidine incorporation using a Betaplate counter (Wallac).
CTL assay
CTL were assayed using a 51Cr release assay. CBA (H2k) splenocytes as effector cells were cultured with irradiated allogeneic C57BL/10 (H2b) stimulators, either LODACs, aB cells, or rB cells for 4 days. Effector cells were added in triplicate to 5000 51Cr-labeled RMA (H2b), the mouse T lymphoma cell line, and P815 (H2d), the mastocytoma cell line, as specific and nonspecific targets, respectively, at different E:T ratios in a total volume of 0.2 ml of complete RPMI 1640. Targets were also incubated with medium alone or 10% Triton X-100 detergent (Sigma-Aldrich) as spontaneous and maximum lysis control, respectively. Plates were incubated for 4 h at 37°C in 5% CO2. A total of 20 µl of the cell-free supernatant was then removed carefully from each of the wells onto a filter mat (Wallac) and assayed in a Betaplate counter. Mean values were calculated from replicate wells, and the results were expressed as percentage of specific lysis, according to the formula [(experimental counts - spontaneous counts)/(maximum counts - spontaneous counts)] x100.
Isolation and analysis of donor cells in recipient mice
Single cell suspensions from the spleens of CBA mice were prepared 24 h after i.v. injection of 1 x 107 rB cells from C57BL/10 mice. Iab-positive cells were detected with biotinylated B21.2 Ab and Streptavidin-PE. B7-2 expression on these cells was determined by staining with the FITC-conjugated GL1 Ab. At least 5000 events were counted per analysis.
Transplant recipient treatment
Recipient mice were pretreated i.v. with either 1 x 107 donor rB cells, aB cells, or 5 x 105 LODACs in combination with 250 µg of a hamster mAb specific for mouse CD40L, MR1 (Bristol-Myers Squibb, Princeton, NJ), and/or 200 µg of human CTLA4Ig (Bristol-Myers Squibb) i.p. 14 days before transplantation.
Heart transplantation
Under anesthesia, fully vascularized heterotopic hearts were grafted into the abdomen using microsurgical techniques (45). Second hearts were grafted into the neck. Graft survival was followed by palpation. Rejection was confirmed by electrocardiogram (46) and/or direct visualization of the graft.
Statistical analysis
Allograft survival between two groups was compared by the Mann-Whitney U test using StatView software (StatView, Abacus Concepts, Berkeley, CA).
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Results |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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The phenotypic and functional characteristics of the rB cells, aB
cells, and LODACs (a dendritic cell-enriched population) were analyzed.
rB cells did not express either B7-1 or B7-2 at the cell surface, as
determined by FACS analysis (1.2% and 2.5%, respectively). In
contrast, aB cells and LODACs expressed B7-1 (49.5% and 68.8%,
respectively) and B7-2 (77.7% and 88.2%, respectively). The LODAC
population contained more than 70% dendritic cells as determined by
FACS analysis using an anti-CD11c Ab (N418) (47). In
functional assays, aB cells and LODACs, but not rB cells, were able to
induce proliferation of naive allogeneic T cells and to generate
cytotoxic T cells in vitro (Fig. 1
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Cardiac allograft recipients, C3H (H2k) mice,
were pretreated with 1 x 107 donor-specific (C57BL/6)
(H2b) or third-party (BALB/c) (H2d) rB cells
and a single 250-µg dose of MR1 (anti-CD40L mAb) 14 days before
grafting. Donor rB cells combined with MR1 induced indefinite
prolongation of allograft survival (>100 days) in 45% of recipients
and the median survival time (MST) of this group, 56 days, was
significantly greater than that of the untreated control group
(MST = 9 days), (p = 0.0023, Fig. 2). Cardiac allograft survival in mice
treated with rB cells plus MR1 was also significantly greater than in
mice treated with rB cells alone (MST = 11 days, p =
0.0007), or with MR1 alone (MST = 18 days, p =
0.0023). This effect was donor specific as the survival of C57BL/6
cardiac allografts after treatment with rB cells from BALB/c mice in
combination with MR1 (MST = 18 days) was significantly less than
the group treated with donor-specific rB cells and MR1 (MST = 56
days, p = 0.0016).
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). B7-2 expression
on rB cells 1 day after injection was identical in the presence or
absence of MR1 (data not shown). While the expression of B7-1 is
delayed as compared with B7-2 after B cell activation in vitro
(48, 49), we unfortunately could not assess the kinetics
of B7-1 expression on donor rB cells in vivo due to the inability to
isolate a sufficient number of donor cells for analysis at 2 or 3 days
after injection of 107 rB cells.
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As MR1 did not prevent the expression of B7-2 on allogeneic rB
cells in vivo (Fig. 3
) and pretreatment with rB cells in combination
with MR1 did not induce indefinite cardiac graft survival in all
recipients (Fig. 2), CTLA4-Ig was added to the pretreatment protocol to
inhibit the B7-CD28 costimulatory pathway. Pretreatment with CTLA4-Ig
and MR1 in combination with rB cells induced indefinite graft
prolongation (>100 days) in 66% of the recipients and the MST for
this group was significantly longer than that of the untreated control
group (MST = 9 days, p = 0.0072) (Fig. 4). However, graft survival after
treatment with rB cells and MR1 was not statistically significantly
augmented by the addition of CTLA4-Ig (p = 0.39).
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Competent APCs induced hyporesponsiveness to alloantigens when delivered with MR1
The above results suggest that the expression of B7-2 by rB
cells when administered in combination with MR1 does not prevent the
induction of hyporesponsiveness to alloantigens in vivo. To further
test this hypothesis we next examined the ability of other APCs that
express B7-1 and B7-2 constitutively to induce unresponsiveness to
alloantigens. CBA (H2k) mice were pretreated with either
1 x 107 aB cells, 1 x 107 rB cells,
or 5 x 105 LODACs (dendritic cell-enriched
population), from C57BL/10 (H2b) mice either alone or in
combination with 250 µg MR1 14 days before a transplantation of a
C57BL/10 cardiac allograft. Mice pretreated with rB cells, aB cells,
LODACs, or MR1 alone promptly rejected the allografts (MST = 9, 7,
8, and 9 days, respectively). However, when the APC pretreatment was
combined with MR1, rB cells (p = 0.0072), aB cells
(p = 0.0062), and LODACs (p = 0.0046)
were all found to be capable of inducing significant prolongation of
cardiac graft survival as compared with untreated control recipients
(MST = 8 days, n = 5) (Fig. 5). While the expression of B7 molecules
by LODACs in vivo after injection could not be documented because of
the inability to isolate sufficient donor cells from the recipient for
analysis, LODACs are known to express high levels of these molecules
(17). These results suggest that the mechanism of action
of CD40 pathway blockade in this model is not the prevention of B7
expression on the donor APC cells.
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To further define the role of the CD40 pathway in the prolongation
of allograft survival in this model, C3H (H2k) mice were
pretreated with 1 x 107 rB cells from CD40 knockout
mice (H2b) and transplanted with wild-type (C57BL/6 x
129)F1 (H2b) hearts. Pretreatment with
CD40-/- rB cells failed to prolong allograft survival
(MST = 11 days) (Fig. 6). C3H mice
pretreated with (C57BL/6 x 129)F1 rB cells rejected
(C57BL/6 x 129)F1 grafts at the same rate as
untreated recipients (MST = 9 and 11 days, respectively). Thus
CD40-/- rB cells alone failed to induce unresponsiveness
to alloantigen, in contrast to the marked prolongation of graft
survival obtained after combined treatment with wild-type rB cells and
MR1 (Fig. 2). This result further supports the hypothesis that the
critical function of CD40 pathway blockade by MR1 in this model is not
to prevent the up-regulation of the B7 molecules on the donor rB cells
used for treatment.
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An alternative hypothesis is that blockade of the CD40 pathway
within the recipient is the critical factor for the induction of
hyporesponsiveness in this model. To directly investigate the role of
CD40L expression by recipient cells in the induction of
unresponsiveness after pretreatment with rB cells, CD40L knockout mice
were used as recipients. Pretreatment of CD40L knockout mice with rB
cells from C3H mice resulted in significant prolongation of C3H cardiac
allografts (MST = 26 days) when compared with graft survival in
untreated CD40L knockout mice (MST = 8 days, p =
0.0171). Wild-type control recipients (C57BL/6 x
129)F1 recipients rejected C3H grafts acutely when either
untreated or following pretreatment with C3H rB cells (MST = 8 and
13 days, respectively) (Fig. 7).
Pretreatment with C3H rB cells produced a significantly greater
prolongation of C3H cardiac allograft survival in CD40L knockout mice
(MST = 26 days) as compared with the survival of grafts in
wild-type control recipients after treatment with rB cells alone
(MST = 13 days, p = 0.0339) (Fig. 7).
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Discussion |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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rB cells do not express the costimulatory molecules, B7-1 and B7-2 (6, 8) and are ineffective stimulators of T cell responses in vitro (13, 20, 21, 38). Therefore, it has been suggested that they may induce hyporesponsiveness to Ags in vivo, a hypothesis that has been confirmed for the induction of unresponsiveness to miH Ags and peptides (26, 27, 52, 53). However, with the exception of a single report (28), rB cells have been incapable of inducing unresponsiveness to MHC and multiple miH Ags in vivo (27, 29).
Pretreatment with rB cells in combination with anti-CD40L mAb (MR1)
induced clear specific prolongation of fully allogeneic cardiac
allografts (Fig. 2). These results are consistent with those of others
who have shown the ability of this strategy to inhibit alloimmune
responses in vitro (37) and in a mouse pancreatic islet
transplant model in vivo (29).
The CD40 pathway plays an important role in B cell activation
(54, 55), in part due to its ability to increase
expression of B7 on the APC (54, 56, 57); thus, we
hypothesized that blockade of CD40 engagement by MR1 would prevent
up-regulation of B7 on the rB cells in vivo, thereby augmenting their
ability to induce unresponsiveness due to lack of costimulation at the
time of Ag recognition. Surprisingly, examination of B7 expression by
the allogeneic rB cells 24 h after in vivo delivery in combination
with MR1 revealed increased expression of B7-2 as compared with the
expression on rB cells prior to infusion (Fig. 3). In addition, the
expression of B7-2 on rB cells 1 day after injection in the presence or
absence of MR1 was identical (data not shown). This suggests that MR1
does not prevent the expression of B7-2 molecules on the rB cells in
this model; however, as we were not able to isolate donor cells from
recipients at later time points, we cannot completely exclude the
possibility that CD40L blockade modulated B7 expression on the B cells
after 24 hours in vivo. These results are consistent with the
previously reported finding that expression of B7-1 and B7-2
transcripts in cardiac allografts is not inhibited by blockade of the
CD40 pathway (58). Taken together, these data suggest that
B7 expression, at least in these models, may be regulated by
CD40-independent factors.
The ability of anti-CD40L mAb to augment the hyporesponsiveness
induced by rB cell treatment therefore appears not to be due to the
complete inhibition of B7 expression on the rB cells. This hypothesis
was further supported by the finding that pretreatment with rB cells in
combination with CTLA4-Ig to block the B7 pathway produced only minimal
graft prolongation (MST = 22 days, Fig. 4). While this observation
might be explained by incomplete blockade of the B7/CD28 interaction
following a single dose of CTLA4-Ig in vivo, we believe this is
unlikely as CTLA4-Ig has high avidity for both B7-1 and B7-2 and it has
a long serum half-life (59, 60). Alternatively, the CD40
and B7/CD28 pathways may have significant independent function in the
response to alloantigens in vivo, a hypothesis that is supported by
observations that have been made in other systems
(61).
If the latter hypothesis is correct, then an increased ability to
induce hyporesponsiveness should be seen with combined blockade of the
B7/CD28 and CD40 pathways when combined with rB cell treatment.
Pretreatment with this combination was not significantly more effective
at inducing indefinite allograft prolongation than when MR1 was used
alone in combination with allogeneic rB cells (Fig. 4). The reasons for
the lack of a dramatic effect of combination treatment in this system,
as compared with others (61), are not clear, but the
results do suggest a dominant role for the CD40 pathway in determining
the response to alloantigen in this system.
Our results are consistent with those of others that demonstrate the importance of the CD40 pathway in T cell immune responses. The CD40 interaction is known to be an important element for the initiation of humoral immune response to thymus-dependent Ags (54), and has recently been reported to play an important role in cell-mediated immunity (30, 31, 32, 62, 63, 64). While the predominant effect of the CD40 pathway in some experimental systems seems to be via its effect on the up-regulation of the B7 molecules on APCs (32, 62, 65), the results of the current study support our previous observation in alloimmune systems (C.P.L., S. Cowan, S. Waitze, D.Z.A, E. Elwood, M. Corbascio, and T.C.P., unpublished observations), which suggest that disruption of the CD40 signals between the recipient T cells may be functionally important for the down-regulation of the cell-mediated immune response to alloantigen.
The failure of pretreatment with rB cells from CD40 knockout mice to
prolong graft survival was surprising in light of the findings of
Hollander et al. (38), which demonstrated unresponsiveness
in the MLR (which measures the response of recipient T cells to donor
APC) after treatment with CD40-/- rB cells. Our results
suggest that blockade of the CD40 pathway between donor cells and
responding T cells is not critical in this model (Fig. 6); thus, the
CD40 pathway may play a more significant role in the interaction
between cells in the recipient’s immune system, which play a critical
role in allograft rejection in vivo. These interactions could occur
during the process of indirect presentation between recipient APCs and
T cells (66), but could also represent functional
CD40-CD40L interactions between T cells. T cells are known to express
CD40 (67). Our observation that pretreatment with donor rB
cells was more effective in inducing graft prolongation in CD40L
knockout mice than in wild-type, littermate controls (Fig. 7) supports
the hypothesis that disruption of the CD40 pathway between recipient
cells may be an important element for the down-regulation of the immune
response following alloantigen encounter in vivo.
It is also interesting to note that MR1 combined with C57BL/6 rB cells
appears to prolong allograft survival in C3H mice to a greater degree
(Fig. 2) than does wild-type C3H rB cells in CD40L knockout
(C57BL/6 x 129)F1 mice (Fig. 6). This may simply
represent a differential role of the CD40 pathway in these two strain
combinations. This possibility could be addressed with a direct
comparison in the C3H to (C57BL/6 x 129)F1 strain
combination. A second hypothesis to explain this observation is that
MR1 interacts with a second ligand that is functional in the CD40L
knockout mice. We consider this less likely because of the similar
phenotype of the CD40 and CD40L gene knock-out mice and because
searches by several groups have failed to identify additional receptors
or ligands for CD40L and CD40 (R. Geha, unpublished observations). A
third possibility is that CD40L is expressed in a functionally
important manner on donor cells. Therefore, mAb therapy could block
both recipient and donor CD40L-bearing cells, whereas use of
CD40L-/- recipients would affect only recipient
cells.
In conclusion, hyporesponsiveness to alloantigens in vivo may be obtained by pretreatment with donor cells combined with blockade of the CD40 pathway by anti-CD40L mAb. These observations demonstrate the importance of the CD40 interaction in the recipient’s immune response to alloantigen, and suggest that the effect of this pathway may be independent of its effect on the B7/CD28 pathway. Moreover, our results suggest that the CD40 pathway in cells of the recipient’s immune system plays a critical role in regulating the outcome of the immune response to alloantigen in vivo.
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Acknowledgments |
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Footnotes |
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2 Address correspondence and reprint requests to Dr. Thomas C. Pearson, Emory University Transplantation Immunology Laboratory, Suite 5105, WMB, 1639 Pierce Drive, Atlanta, GA 30322. E-mail address:
3 Address correspondence and reprint requests to Dr. Christian P. Larsen, Emory University Transplantation Immunology Laboratory, Suite 5105, WMB, 1639 Pierce Drive, Atlanta, GA 30322. E-mail address:
4 Current address: Bristol-Myers Squibb Pharmaceutical Research Institute, Princeton, NJ 08543-4000.
5 Current address: Rosetta Inpharmatics, Kirkland, WA 98034.
6 Abbreviations used in this paper: rb cells, resting B cells; miH, minor histocompatibility; aB, activated B cell; LODACs, low density adherent cells; PE, phycoerythrin; CD40L, CD40 ligand; MST, median survival time.
Received for publication December 1, 1997. Accepted for publication July 14, 1998.
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References |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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stimulated monocytes: a potential mechanism for amplification of T cell activation through the CD28 pathway. Cell. Immunol. 137:429.[Medline]
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K. Namba, K. Ogasawara, N. Kitaichi, T. Morohashi, Y. Sasamoto, S. Kotake, H. Matsuda, K. Iwabuchi, C. Iwabuchi, S. Ohno, et al. Amelioration of Experimental Autoimmune Uveoretinitis by Pretreatment with a Pathogenic Peptide in Liposome and Anti-CD40 Ligand Monoclonal Antibody J. Immunol., September 15, 2000; 165(6): 2962 - 2969. [Abstract] [Full Text] [PDF]
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N. D. Jones, A. Van Maurik, M. Hara, B. M. Spriewald, O. Witzke, P. J. Morris, and K. J. Wood CD40-CD40 Ligand-Independent Activation of CD8+ T Cells Can Trigger Allograft Rejection J. Immunol., July 15, 2000; 165(2): 1111 - 1118. [Abstract] [Full Text] [PDF]
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P. A. Taylor, A. Panoskaltsis-Mortari, R. J. Noelle, and B. R. Blazar Analysis of the Requirements for the Induction of CD4+ T Cell Alloantigen Hyporesponsiveness by Ex Vivo Anti-CD40 Ligand Antibody J. Immunol., January 15, 2000; 164(2): 612 - 622. [Abstract] [Full Text] [PDF]
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N. N. Iwakoshi, J. P. Mordes, T. G. Markees, N. E. Phillips, A. A. Rossini, and D. L. Greiner Treatment of Allograft Recipients with Donor-Specific Transfusion and Anti-CD154 Antibody Leads to Deletion of Alloreactive CD8+ T Cells and Prolonged Graft Survival in a CTLA4-Dependent Manner J. Immunol., January 1, 2000; 164(1): 512 - 521. [Abstract] [Full Text] [PDF]
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M. E. Ozaki, B. A. Coren, T. N. Huynh, D. J. Redondo, H. Kikutani, and S. R. Webb CD4+ T Cell Responses to CD40-Deficient APCs: Defects in Proliferation and Negative Selection Apply Only with B Cells as APCs J. Immunol., November 15, 1999; 163(10): 5250 - 5256. [Abstract] [Full Text] [PDF]
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D. M. Shepherd and N. I. Kerkvliet Disruption of CD154:CD40 Blocks Generation of Allograft Immunity Without Affecting APC Activation J. Immunol., September 1, 1999; 163(5): 2470 - 2477. [Abstract] [Full Text] [PDF]
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