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*
Department of Immunology IMM-25, The Scripps Research Institute, La Jolla, CA 92037; and
Immunobiology Department, DNAX Research Institute, Palo Alto, CA 94304
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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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The response of T cells to agonist ligands can be modified by a number
of factors, including adhesion molecule-mediated enhancement of binding
to APC (2), coreceptor CD4 (3) or CD8
(4) binding to MHC, or IL-2 mediated augmentation of
low-affinity responses (5). Inhibition of T cell responses
can also occur through the triggering of molecules such as CTLA4
(6). The activation of NK cells is also regulated by
inhibitory receptors that bind MHC class I ligands (reviewed in Ref.
7). In mice, the Ly-49 family of NK cell receptors are
encoded by 
9 polymorphic genes on chromosome 6, and these molecules
are expressed on overlapping subsets of NK cells (8, 9, 10).
Many of the Ly-49 receptors contain immunoreceptor tyrosine-based
inhibition motifs
(ITIM)3 in their
cytoplasmic domains that are responsible for recruitment of
intracellular tyrosine phosphatases and inhibition of NK cell function
(11, 12, 13). Recent studies have also found that peripheral
mature T cells are inducible for expression of the NK inhibitory
receptors such as Ly-49A, and it is possible that expression of these
molecules in activated T cells may influence subsequent immune
responses (14, 15, 16, 17). Ly-49A expression by T cells is not
found during early stages in T cell development, presumably because its
activation might alter signals important in triggering specific
developmental decisions (18).
In the present work, we have studied the consequences of altering TCR signaling with an Ly-49A transgene. When activated by its target ligand H-2Dd, the Ly-49A molecule inhibits T cell function, in part, through modulation of signaling (11, 12, 13). Because these events are dependent on the presence of the Ly-49A ligand, T cell function appears to be normal in the absence of the ligand, allowing for proper assessment of the selected TCR repertoire. Our results support the conventional affinity model of repertoire selection in which the intensity of TCR signaling determines the outcome of positive selection. In addition, the alterations in thymic selection induced by the Ly-49A transgene suggest that T cell expression of Ly-49A can influence both peripheral activation and the TCR repertoire.
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Materials and Methods |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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The CD4 transgene cassette is construct "i" from Sawada et al. (19), which was modified to remove the SalI site in the promoter leaving two SalI sites flanking the CD2 cDNA insert. The human CD2 cDNA was excised from the cassette using SalI. The plasmid containing the mouse Ly-49A cDNA (20) was obtained from Dr. Wayne Yokoyama (Washington University, St. Louis, MO). The entire Ly-49A coding sequence was PCR amplified using primers that included SalI sites on their 5' ends. The PCR product was cloned into a TA vector (Invitrogen, San Diego), sequence verified, then subcloned into the CD4 promoter cassette. Transgene integration was assessed using a 1.15-kb PstI-BglII fragment from the first intron of the CD4 transgene cassette and PCR using primers from the CD4 cassette vector and the Ly-49A insert (CD4 cassette vector sense 5'-CAGATTCCCAACCAACAAGAG-3' and Ly-49A antisense 5'-CCATGTTTTTCTGTCCATGACG-3').
Mice
C57BL/6, B10.A(3R), B10.D2, and B10.HTG mice were purchased from The Jackson Laboratory (Bar Harbor, ME) and The Scripps Research Institute (TSRI) rodent breeding colony. To make transgenic mice, fertilized embryos from C57BL/6 mice were microinjected with Ly-49A transgene DNA according to standard procedures. Founder mice were screened by Southern blot and PCR of tail DNA. Ly-49A-transgenic mice were bred to B10.D2 mice until homozygosity was achieved at the MHC as determined by microsatellite analysis. The backcross of TCR-SFE mice (21) to B10.D2, generating TCR-SFE x B10.D2-transgenic mice has been described previously (22); these were used to generate SFE x Ly-49A double-transgenic mice. The SFE TCR is restricted to I-Ed and is specific for the hemagglutinin peptide 110–119 (SFERFEIFPK) of influenza (A/PR/8/34 Mt. Sinai). SFE x Ly-49A double-transgenic mice were also crossed to B10.HTG mice (The Jackson Laboratory) until homozygous at the MHC class I H-2D locus as determined by flow cytometry using Dd- and Db-specific Abs (PharMingen, San Diego, CA). All mice were housed in specific pathogen-free conditions at TSRI rodent colony in accordance with National Institutes of Health and TSRI institutional guidelines.
Flow cytometric analysis
Mononuclear single-cell suspensions were stained with
combinations of anti-CD4-PE, anti-CD8-allophycocyanin,
anti-heat-stable Ag-FITC, anti-Ly-49A (YEI/48)-FITC,
anti-CD62L-FITC, V
3-FITC,
V5-FITC, or V8-FITC
(PharMingen). Peripheral blood and splenic mononuclear cells were
prepared using Lympholyte-M (Accurate Chemical and Scientific,
Westbury, NY) before staining. Determination of apoptotic cell death
was performed using FITC-conjugated Annexin V (Clontech, Palo Alto,
CA), which binds to phosphatidylserine that becomes translocated to the
outer cell membrane shortly after apoptosis initiation. Dual staining
with Annexin V and propidium iodide (PI), following the manufacturer’s
instructions, allows for discrimination between apoptotic
(AnnexinV+/PI+/-) and
necrotic cell death (Annexin
V-/PI+).
Immunofluorescence analysis was performed using a FACS calibur flow
cytometer with CellQuest version 3.2 software (Becton Dickinson, San
Jose, CA).
In vitro stimulation of CD4 T cells with staphylococcal enterotoxin B (SEB)
Spleens from male C57BL/6-transgenic or -nontransgenic mice from each lineage were harvested and pooled. Dispersed cells were treated with ammonium chloride solution, washed, and incubated with the following mixture of Abs at 10 µg/ml for 25 min at 4°C: Anti-I-Ab/Eb (2G9), CD11b (Mac-1, M1/70), CD45R (B220, RA3–6B2), TER-119, Ly-6G (Gr-1, RB6–8C5), and CD8 (53-6.7; PharMingen). After washing, cells were incubated with anti-mouse IgG and anti-rat IgG beads (Dynal, Great Neck, NY) for 25 min, and unwanted cells were removed with a magnet. The remaining cells were 65–75% CD4+ T cells. APC were prepared from C57BL/6 or B10.A(3R) mice, but depleted using anti-CD4 (GK1.5), anti-CD8 (53-6.7), and anti-CD90 (Thy-1, G7), followed by bead depletion and irradiation (3000 rad). A total of 1 x 105 CD4-enriched cells were cultured with 4 x 105 APC and dilutions of SEB (Toxin Technology, Sarasota, FL) for 4.5 days. Proliferation was assessed by addition of 1 µCi [3H]thymidine during the last 24 h of culture.
Generation of bone marrow chimeric mice
Bone marrow was harvested from TCR-SFE and SFE x Ly-49A donor mice on a B10.D2 background. Irradiated (1100 rad) 8-wk-old wild-type B10.D2 or B10.HTG recipients were injected i.v. with 2–3 x 107 donor bone marrow cells. Seven weeks after transfer, thymuses were analyzed by flow cytometry to determine the percentages of CD4+ and CD8+ cells.
Proliferation assays
Lymph node CD4+ responder T cells were obtained from 6- to 8-wk-old Ly-49A (B10.D2) single-transgenic or wild-type B10.D2 mice. CD4+ T cells were purified by negative selection using goat anti-rat IgG-conjugated magnetic beads (BioMag; Polysciences, Warrington, PA) following incubation of mononuclear cells with Abs to remove B (B220), CD8+ T (CD8), and monocyte/macrophage (F4/80 and M5/114) cell populations. Irradiated (2500 rad) stimulator cells were prepared from spleens of 6- to 8-wk-old wild-type B10.D2, B10.HTG, and C57BL/6 mice. CD4+ responder cells (1.0 x 105 cells/well) were incubated with irradiated splenic stimulator cells (5 x 105 cells/well) for 72 h at 37°C in 5% CO2 in 96-well plates. Plates were pulsed with 1 µCi/well of [3H]thymidine for the last 24 h before harvest and counted in a Microbeta 1450 liquid scintillation counter (LKB Instruments, Gaithersburg, MD). All samples were assayed in triplicate. Error bars represent 1 SD from the mean cpm.
Histology
Tissues from Ly-49A (B10.D2) and wild-type B10.D2 mice were paraffin embedded, sectioned, and stained with hematoxylin and eosin. Photographs were taken at x 200 (original magnification).
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Results |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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We generated an Ly-49A transgene driven by a modified CD4 promoter
that lacks the sequences suppressing expression in CD8 T cells
(19). As a result, in mice harboring the transgene, Ly-49A
was expressed early in T cell development, and expression continued
throughout T cell differentiation and emigration to the periphery in
both CD4 and CD8 T cells (Fig. 1
A). In mice expressing the
specific ligand H-2Dd, cell surface expression of
the transgene was decreased, as described for NK cells in other similar
transgenic models (23, 24), but was still present on
nearly all T cells (Fig. 1A). The Ly-49A molecule is
believed to inhibit NK responses by activating an intracellular
phosphatase (SHP-1) when triggered by its ligand (11, 12, 13).
In similar fashion, Ly-49A activation also inhibited T cell responses,
as responses of T cells from Ly-49A-transgenic mice (on the
ligand-negative C57BL/6 strain) to the superantigen SEB were inhibited
when the APC expressed the Ly-49A ligand (Fig. 1B). This
inhibitory effect is also consistent with previous reports with another
Ly-49A transgene (25).
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A). When gated on
CD4 single positive populations, the absolute number of
HSAlow cells (mature CD4 thymocytes) was 4.6
x 105 and 0.4 x 105
from Db and Dd thymuses,
respectively. Similarly, 1.6 x 105 and
0.9 x 105 CD8 single positive
HSAlow thymocytes were recovered from
Db and Dd thymuses,
respectively. The minimal effect on CD8 thymocytes may be due to
competition between Ly-49A and CD8 for binding to
H-2Dd. Alternatively, CD8 thymocytes may be less
sensitive to Ly-49A-mediated inhibition than CD4 thymocytes. The
current studies cannot distinguish these possibilities. When stained
with AnnexinV, the percentage of apoptotic cells
(AnnexinV+) was much higher in B10.D2 backcrossed
mice expressing the Ly-49A ligand H-2Dd as
compared with transgenic mice backcrossed to B10.HTG expressing the
same class II I-Ad but different class I
H-2Db instead of H-2Dd
(Fig. 2B).
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As a first indicator of alterations in thymic-negative selection, we
studied the representation of T cells expressing the
superantigen-reactive TCR V5 (Fig. 2
C). In the presence
of the endogenous retroviral superantigens mtv-8 and 9 and the class II
I-E molecule, V5 cells were deleted in the thymus (29).
In C57BL/6 mice, the percentage of TCR V5 cells reflected the
nondeleted numbers, whereas normal B10.D2 mice showed the
I-E/superantigen-mediated deletion of V5 cells. Interestingly,
Ly-49A-transgenic mice on the B10.D2 background showed a failure in
superantigen-mediated deletion. This was evident in thymic single
positive subsets, but also in the peripheral lymph nodes and spleen
(Fig. 2C and data not shown).
Although the Ly-49A transgene may modulate TCR signaling, the presence
of autoreactive receptors may still allow for an initial triggering of
the T cells, followed by Ly-49A-mediated suppression of the responding
cells. Although this might prevent widespread persistent T cell
activation and autoimmune disease, it does appear that a high frequency
of the peripheral T cells in the Ly-49A-transgenic mice show evidence
for prior activation. The peripheral T cells continued to express the
transgene Ly-49A, although at significantly lower levels in the B10.D2
mice (Fig. 1A), indicative of their prior engagement with
the ligand. Also in both spleen and lymph node, a very high proportion
of the peripheral CD4 and CD8 T cells are negative for the lymph node
addressin CD62L (Fig. 2D). However, since the thymus output
may be impaired (suggested by the increased intrathymic apoptosis),
some or most of this peripheral T cell activation may also be due to
Ag-independent expansion to compensate for the lymphopenia.
For the reasons discussed, memory or activation markers might not be
reliable indicators of autoreactive T cells. Therefore, to reveal the
presence of any functional high-affinity autoreactive cells, lymphocyte
proliferation assays were done using stimulators presenting self-MHC
class II molecules in the absence of the Ly-49A ligand. In this
situation, purified CD4 T cells from Ly-49A-transgenic mice were
stimulated with APC from B10.D2 or B10.HTG mice (Fig. 2E).
Proliferative responses of lymph node CD4 T cells were very low against
B10.D2. Interestingly, CD4 T cells showed significant proliferation
against the B10.HTG stimulators, although not as high as to the
positive control C57BL/6 stimulators. Similar results were obtained
with CD4 T cells from spleen (data not shown). Thus, triggering of the
transgene Ly-49A during positive selection in the thymus may permit
positive selection of a significant proportion of T cells that would
otherwise be deleted due to autoreactivity.
Autoimmune inflammatory syndrome in Ly-49A-transgenic mice: autograft-vs-host-disease?
The high-affinity autoreactive T cells selected by the thymus in
the presence of the Ly-49A transgene appear to be generally held in
check by the triggering of the Ly-49A in the periphery. However,
various mechanisms (persistent TCR activation, decreased activation
thresholds, or other unknown mechanism) might cause the autoreactive
cells to escape regulation by the Ly-49A transgene. Moreover, as shown
above, the level of expression of the transgene can be significantly
decreased in the presence of the ligand (Fig. 1A), and, in
some T cells, transgene Ly-49A expression might be entirely lost. In
any case, autoreactive T cells might escape regulation and produce
autoimmune disease. Interestingly, in some lines of Ly-49A-transgenic
mice, a significant proportion of mice develop a chronic wasting
disease associated with skin keratitis and mixed infiltrates in several
tissues. Penetrance is less than complete, for reasons that remain
unclear, but it may be related to variable expression of Ly-49A. This
occurred only in mice with the Ly-49A transgene on the B10.D2
background, and was never seen in backcrosses onto the B10.HTG
backcrossed (i.e., lacking the Ly-49 ligand). The syndrome was variably
found in mice from two different viral pathogen-free mouse colonies,
but the affected individuals generally showed symptoms by 2 wk of age.
Finally, in one line of Ly-49A-transgenic mice (line 6343) backcrossed
to the RAG-1 knockout, the syndrome was also absent, suggesting that
the pathology was dependent on the presence of T or B lymphocytes (data
not shown).
The histology of the most severely affected mice showed significant
mononuclear infiltrates in the portal tracts of the liver and
perivascular/peribronchial spaces of the lung (Fig. 3). Occasional granulocytes, including
both neutrophils and eosinophils, could also be seen. Skin also showed
diffuse dermal mononuclear infiltrates, and intestinal epithelium
showed scattered submucosal accumulations of mononuclear cells.
Finally, the spleen showed extramedullary hematopoiesis and loss of
white pulp definition, in part due to replacement by lymphoblasts (data
not shown). Interestingly, this pattern of infiltrates is similar to
the histopathological picture seen in graft-vs-host disease
(30, 31, 32), consistent with the notion that the disease may
represent the activation of autoreactive T cells.
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One disadvantage of studies on polyclonal T lymphocyte
populations is that the effects on Ag-specific receptors cannot be
observed in detail. The studies above predict the shifting of receptor
thresholds for both positive and negative selection. To examine these
effects with a known receptor, we crossed the Ly-49A transgene to
TCR-SFE-transgenic mice that express a receptor specific for an
influenza hemagglutinin peptide presented on I-Ed
(21). The expression of the Ly-49A transgene was
unaffected by the TCR transgene; therefore, expression was evident in
all thymocyte subsets (Fig. 4A). As predicted, the skewing
of single positive subsets toward CD4 is lost in the presence of the
Ly-49A transgene (Fig. 4B), suggesting that the positive
selection signals normally present in B10.D2 thymus fail to influence
thymocyte development. This loss of positive selection was also evident
in the periphery, shown as the absence of CD4 skewing and reduced total
numbers of peripheral T cells (Table I).
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C). Thus, the defect in CD4
T cell production appeared to be during positive selection on thymic
cortical epithelium and was independent of T cell or APC expression of
H-2Dd. |
Discussion |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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In these studies, we assessed the effects of Ly-49A activation on T cell selection in the thymus. The alterations in positive and negative selection observed are consistent with a general shift in the sensitivity of the signal transduction mechanisms associated with TCR triggering. That is, the Ly-49A transgene appears to provide us with a reversible (+/- ligand) adjustment in the "gain" setting of the TCR complex. However, as simple and useful as this effect appears, we must take into account the complexity of TCR-mediated signaling and phosphatase action.
The modulation of TCR signaling in our studies is likely to be dependent in part on Ly-49A recruitment of the tyrosine phosphatase SHP-1 (11, 12, 13). Activated SHP-1 has many substrates, but studies have suggested that its ability to regulate ZAP-70 may be the most relevant to the suppression of TCR-mediated activation (33, 34). However, before SHP-1 can be recruited, Ly-49A must first be phosphorylated on its intracytoplasmic ITIM (35), and it is not clear whether this occurs simultaneously with TCR activation, or whether it can only follow an initial TCR trigger with activation of the associated signaling kinases. The exact sequence of events may have an important influence on the outcome during thymic selection, especially with respect to how the affinity of the specific ligand-TCR interaction is perceived by the T cell. In this regard, it is important to note that in studies on NK cells, the ligation of Ly-49A and recruitment of SHP-1 inhibits early signaling events, including polyphosphoinositide turnover and tyrosine phosphorylation (13). Thus, although TCR-mediated signaling may involve different signaling kinases from those required in NK activation, SHP-1 action may act early in TCR responses as well.
It is not yet clear how the affinity of the ligand/TCR binding is conveyed to the T cell, whether it is through the amount of peak intracellular kinase activity, the kinetics of TCR recycling, or some other mechanism. In addition, it has been shown recently that TCR signaling involves both the assembly of supramolecular activation complexes at the interface with the APC (36) and the reorganization of kinase-rich raft microdomains in response to costimulation through CD28 (37). Since positive selection in the thymus is likely costimulation (CD28) independent, it is not clear how these complex structures would be affected by Ly-49A triggering. Because SHP-1 can inhibit both cytokine receptor signaling and chemokine responsiveness (38, 39), the ability of SHP-1 activation to modulate TCR signaling might not be through a simple reduction in a single parameter. In fact the effect of Ly-49A signaling may be through its action on other signaling pathways distinct from those triggered directly by the T cell Ag receptor. However, despite the potential complications listed here, we were still able to demonstrate predictable shifts in the dose response of T cells in several situations, including thymic positive and negative selection and peripheral T cell responses to self and superantigen.
Does control over the levels of phosphorylation of signaling kinases cause directly proportional effects on signal strength? If activation of SHP-1 is indeed able to affect the dose response of lymphocyte signaling, then a similar (but reciprocal) effect should be evident in mice lacking SHP-1, as in mice carrying the viable motheaten (mev) mutation. That is, T cells from motheaten mice would be predicted to be hyperresponsive to weak stimuli. Consistent with predictions, both thymocytes and mature T cells from motheaten mice showed increased proliferative responses to TCR activation (40, 41), although Fas-mediated apoptosis also appeared to be defective (42). In the case of B cells, autoreactive cells carrying the motheaten mutation have been shown to be far more sensitive to negative selection by low levels of Ag (43). The consequences with regard to TCR repertoire selection have only recently been examined and show a role for SHP-1 in regulating both positive and negative selection (44). Thus, assuming that Ly-49A acts primarily through SHP-1 recruitment, then it appears that control over transgene Ly-49A activation can effectively serve as a rheostat to regulate the signal intensity generated through lymphocyte Ag receptors.
This effect may have important consequences in normal peripheral T cells with inducible expression of Ly-49 molecules or killer inhibitory receptors (KIR) on human cells (14, 15, 16, 17). Activation of naive T cells (negative for Ly-49) would be dependent primarily on the strength of the APC and initial local concentration of target ligand. By contrast, in the activated or memory cell population, cells that have induced expression of Ly-49 molecules would now have acquired a new level of regulation. In effect, Ly-49 activation would raise the threshold for reactivation of these cells and potentially counteract any other mechanisms lowering their signaling threshold. Although the mechanisms regulating inducible Ly-49 or KIR expression on T cells are not yet well defined, manipulation of this phenomenon may be very helpful in modifying ongoing immune responses in vivo.
The affinity model for TCR repertoire selection
Selection of the self-MHC-restricted TCR repertoire in the thymus has to solve a number of potentially incompatible problems simultaneously. First, the repertoire has to be selected with a preference for the recognition of self-MHC molecules as restricting elements. To bind more effectively to the Ag-presenting molecules in the face of great peptide diversity, the receptor repertoire is selected on the basis of effective interaction with exposed surfaces on the MHC molecule. Many of these surfaces include amino acid residues that are polymorphic; therefore, recognition of peptide Ag plus MHC tends to be very much dependent on recognition not only of peptide and MHC class-specific determinants, but also the polymorphic determinants ("restriction determinants"), a phenomenon now described as MHC restriction.
Second, given the random nature of TCR gene rearrangement, the selection of T cells with effective binding to self-MHC molecules necessarily includes a great range of affinities. Among the receptors finally chosen for export, affinities for self-MHC cannot be so high as to trigger T cell responses to the array of normal self-peptides, but still must allow for the potential binding with high affinity to self-MHC-presenting target Ags. In addition, during development and selection of the thymocytes, signaling through the TCR during thymocyte selection is absolutely required to inform the cell that it has a functional receptor with a functional affinity for self-MHC molecules. Thus, through this process, the receptor must be sophisticated enough to distinguish between reasonable affinity and pathologically high affinity. How can this be achieved?
According to the conventional "affinity model," the TCR is a sophisticated gauge of receptor affinity that only permits selection at a middle range of affinities. This mechanism requires a shift in either the stimulation threshold or signaling pathway during final maturation so that only the higher affinity ligands will trigger mature T cells in the periphery. Alternatively, thymocyte development could separate the high and low thresholds temporally (45, 46, 47). That is, during positive selection, the stimulation threshold is set fairly low to allow for positive selection for any receptor above a minimum affinity. Selection of high-affinity autoreactive cells is permitted during this first step. This is followed by an efficient negative selection step, where high-affinity signaling in the presence of a costimulatory signal (e.g., CD80, CD86, or other molecules) induces deletion. In this case, the developing thymocyte perceives the first TCR-signaling event (low threshold) as positive selection, and after a maturation step, the next TCR-signaling event (high threshold with costimulation) triggers negative selection. A final maturation step in this case allows for any signal (again high threshold with costimulation) to be perceived in the peripheral immune system as a positive stimulation.
The events described in the Ly-49A model can be placed on a map
representing the affinity model of selection (Fig. 5). Under normal conditions (Fig. 5, top), selection of the randomly rearranged TCR repertoire
gives rise to a distribution of receptors with varying affinities for
self -MHC. Most receptors will have no functional affinity for thymic
MHC, and therefore clones with these receptors are lost (death by
neglect). Above a certain threshold, receptors will signal the
developing thymocyte to continue its development (positive selection),
and, at a later step, a strong signal above a certain threshold will
initiate deletion of the thymocyte (negative selection).
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, bottom
graph). The frequency of nonfunctional receptors is increased, and
this would predict the observed increase in steady-state apoptosis in
the Ly-49A-transgenic thymus (the expanded death by neglect region in
Fig. 5, bottom graph). With a decreased pool of receptors
with acceptably high affinity for positive selection, thymic output is
also predicted to be less efficient. Finally, the modulation of
receptor signaling now permits, indeed, it requires, the positive
selection of receptors with high affinity for self-MHC
("high-affinity autoreactive receptors" in Fig. 5, bottom
graph). When the modulating phosphatase is removed by presenting
the TCR ligand in the absence of the Ly-49A ligand, the autoreactivity
is revealed. In summary, the results show that by manipulating TCR-mediated signaling, we can alter the fates of specific T cell clones during thymic selection. Specifically, in our Ly-49A-transgenic mouse model, the modulation of TCR signaling causes an increased rate of thymocyte loss due to impaired positive selection. At the same time, many high-affinity autoreactive cells fail to be deleted, and instead proceed through normal positive selection and export to the periphery.
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Acknowledgments |
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Footnotes |
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2 Address correspondence and reprint requests to Dr. David Lo, Department of Immunology IMM-25, The Scripps Research Institute, 10550 North Torrey Pines Road, La Jolla, CA 92037. E-mail address:
3 Abbreviations used in this paper: ITIM, immunoreceptor tyrosine-based inhibition motifs; SEB, staphylococcal enterotoxin B; SFE, hemagglutinin peptide 110–119; PI, propidium iodide.
Received for publication July 28, 1999. Accepted for publication October 27, 1999.
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References |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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5+ T lymphocytes. Proc. Natl. Acad. Sci. USA 89:5138.
in murine thymocytes and lymph node T cells. Immunity 1:675.[Medline]
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P. Brawand, F. A. Lemonnier, H. R. MacDonald, J.-C. Cerottini, and W. Held Transgenic Expression of Ly49A on T Cells Impairs a Specific Antitumor Response J. Immunol., August 15, 2000; 165(4): 1871 - 1876. [Abstract] [Full Text] [PDF]
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