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Laboratory of Experimental Immunology, Division of Basic Sciences, National Cancer Institute-Frederick Cancer Research and Development Center, Frederick, MD 21702
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Abstract |
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 Top Abstract IntroductionMaterials and MethodsResultsDiscussionReferences |
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1/2). Murine T
cells also were cultured in the presence of targets that express
(H-2Dd) which is inhibiting for the Ly-49A and G2
receptors. These cells were examined for a coincident increase in
cytokine production (IFN-
, TNF-, and granulocyte-macrophage CSF).
Abs to Ly-49A and G2 or their respective class I ligands blocked the
negative signals mediated via the Ly-49 receptors and increased IFN-
and granulocyte-macrophage CSF production after interaction of these T
cells with H-2Dd-expressing tumor targets. Furthermore, an
EL-4 T cell line expressing both Ly-49A and G2, when treated with mAb
YE148 and 4D11, demonstrated reduced cytokine production and calcium
mobilization. These results demonstrate for the first time that Ly-49
class I binding receptors, previously thought to be restricted to mouse
NK cells, can mediate important physiological functions of T cell
subsets. |
Introduction |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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The Ly-49G2 subset of NK cells also has been shown to be inhibited by target cells expressing H-2Dd and/or H-2Ld (10). Studies with Ly-49G2+ NK cells have relied primarily on the reversal of target cell inhibition by mAb 4D11, which binds Ly-49G2 and mAb specific for H-2Dd and H-2Ld. The Ly-49C+ subset of NK cells has been shown to bind the class I molecules H-2b, H-2d, H-2k, and H-2s (11). Recent data from Yu et al. demonstrate that Ly-49C+ NK cells from BALB/c and BALB/b mice are inhibited by H-2d and H-2Kb class I Ags. The authors of this study concluded, however, that not all Ly-49C+ NK cells function in the same manner in all mouse strains and suggested that allelic differences may regulate class I recognition by these cells (12). Previous data reported by members of this group have shown that 5E6+ NK cells can reject bone marrow grafts expressing H-2d but not H-2b (13, 14). The recent finding that 5E6 Ab recognizes both Ly-49C and Ly-49I (15) has clouded the interpretation of data obtained using mAb 5E6. However, the recent description of the Ab NK2.1 (which recognizes Ly-49C only) combined with 5E6 (Ly-49C/I) should provide the necessary tools for elucidation of the roles of these receptors. In this context, examination of the regulation of nonlytic, secretory functions by the Ly-49A and G2 molecules may provide important complimentary information about the in vivo functions of these NK receptors in marrow transplantation and other immunoregulatory functions of these cells.
For a number of years, it has been known that there is a unique T cell subset (2% of spleen) that coexpresses CD3 and the NK-associated marker NK1.1. This NK1.1+ T cell subset produces high titers of cytokines and expresses a restricted repertoire of TCRs (14, 15, 16, 17). In addition, most of these NK1.1+ T cells mediate spontaneous IL-2 induced killing similar to that documented for NK cells (16, 18, 19). In the present study we have examined these NK1.1+ T cells for their expression of Ly-49 receptors and the roles of those receptors in their biology.
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Materials and Methods |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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All mice were obtained from the Animal Production Area, Frederick Cancer Research and Development Center (Frederick, MD) or The Jackson Laboratory (Bar Harbor, ME). Mice were between 8 and 20 wk of age when euthanized. Animal care was provided in accordance with the procedures outlined in the Guide for the Care and Use of Laboratory Animals (National Institutes of Health Publication 86–23, 1985).
Flow cytometric analysis
Cells were stained as previously described (15) and analyzed on a FACSort flow cytometer (Becton Dickinson, Mountain View, CA). Cell sorting was performed on either an EPICS 750 (Coulter Electronics, Hialeah, FL) or a FACStar (Becton Dickinson). Cells were directly stained using phycoerythrin-labeled (PE) and FITC-labeled primary Abs and indirectly stained using a primary Ab followed by an isotype-specific FITC- or PE-conjugated secondary Ab or a biotinylated primary Ab followed by streptavidin PerCp (San Jose, CA) or streptavidin-Red 670 (Life Technologies, Gaithersburg, MD).
Complete medium
RPMI 1640 (BioWhittaker, Walkersville, MD) was supplemented with 10% FBS (Atlanta Biologics, Norcross, GA), L-glutamine, penicillin/streptomycin, sodium pyruvate, nonessential amino acids, HEPES, and 2-ME.
NK and T cell isolation
Murine splenic or liver NK and T cells were isolated from nylon wool-nonadherant lymphocytes. The cells were stained with combinations of CD3 and Ly-49s, and were sorted for subpopulations expressing the desired phenotype. Cells then were expanded for 7 to 10 days in complete medium supplemented with 1000 Cetus units (CU)/ml.
Targets
Con A blasts were made using B6 spleen cells that express class I (H-2Db) or BALB/c spleen cells (H-2Dd), treated for 48 h with 10 µg Con A/2 x 106 cells. Tumor targets were maintained in culture as previously described (15). P815 is a rat mastocytoma. WEHI 164 is a methylcholanthrene-induced fibrosarcoma. L5 MF22 is a B cell line that naturally expresses an H-2Db class I molecule. This line has been transfected with a neomycin resistance gene alone (L5neo3) or with the class I (H-2Dd; L5cDd104)-expressing gene (provided by Dr. I. Nakamura, Buffalo, NY).
Cytotoxicity assays
Tumor targets were labeled with 51Cr and used in 6-h cytotoxicity assays as previously described (20). Assays involving mAb included the specific Abs at a concentration of 20 µg/ml for the duration of the cytotoxicity assay. Data are either presented as lytic units per 107 cells or as percent specific lysis.
Abs used
The following mAbs to Ly-49 receptors were used: YEl/48 (Ly-49A,
rat IgG2c), SW5E6 (Ly-49C, mouse IgG2a), and 4D11 (Ly-49G2, rat IgG2a)
(6, 8, 9, 10, 11, 12, 13). Ly-49D (4E5) is a rat IgG2a Ab specific for Ly-49D and has
been characterized previously (J. R. Ortaldo, A. T. Mason, R.
Winkler-Pickett, W. E. Murphy, and L. H. Mason, manuscript in
preparation). Abs to MHC class I were 34.5.8S (H-2Dd
1/2 mouse IgG2a), 28.8.6 (H-2Kb/H-2Db
mouse IgG2a), and 11.4.1 (H2Kk mouse IgG2a). T cell
reagents were CD3
(500A2), TCRß, and TCR
and were
purchased from PharMingen, Inc. (San Diego, CA).
Isolation of liver lymphocytes
Liver cells were isolated as described previously (21) after mice were injected 3 days, twice daily, with 33 µg of rIL-2 from either Chiron (Emeryville, CA) or Hoffmann-La Roche, Inc. (Nutley, NJ). Cells were then used immediately for functional studies.
Cell treatments for cytokine induction
Cytokines were induced by coculture of IL-2-propagated NK or T cells with tumor target cells or mAb after 18-h incubations. Briefly, cells were plated in 1 ml of complete medium with 100 CU/ml IL-2 at 0.5 or 1 x 106 cells/well in either 48- or 24-well microtiter plates, respectively. Tumor targets, when added, consisted of a final ratio of one tumor per 10 lymphocytes. Purified Abs were added at 20 µg/106 lymphocytes. PMA was added at 10 ng/ml, and ionomycin was added at 1 µg/ml. Cells were cultured at 37°C in a humidified CO2 incubator, and supernatants were harvested after centrifugation and then frozen at -20°C until tested in ELISA.
Cytokine measurement
Cytokines were measured using IFN- and GM-CSF ELISA kits (R&D
Systems, Minneapolis, MN). All samples were measured in duplicate
against the standard curve of the assay and are reported as picograms
per milliliter. In all assays, the SD of the cytokine measurement was
<25 pg/ml.
Calcium mobilization
Analysis of changes in the intracellular Ca2+ concentration ([Ca2+]i) of T cells was conducted using a FACSort flow cytometer (Becton Dickinson) and the calcium-sensitive fluorochrome fluo-3 (Molecular Probes, Eugene, OR) (9). [Ca2+]i was monitored with fluo-3-loaded cells suspended at 37°C in Dulbecco’s PBS with Ca2+ and Mg2+ supplemented with 5 mM glucose. Increases in [Ca2+]i were detected as increases in fluo-3 fluorescence vs time. Data were analyzed using Multitime Analysis Software (Phoenix Flow Systems, San Diego, CA).
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Results |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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These can be detected and their function evaluated by the
following Abs: Ly-49A (A1; YE148), Ly-49C/I (5E6), Ly-49G2 (4D11), and
Ly-49D (4E5). Since Ly-49A and Ly-49G2 together are expressed on
75% of B6 NK cells, we examined T cells for coexpression of CD3,
DX5 (a marker that costains most NK1.1-positive cells) and Ly-49.
Figure 1
demonstrates the expression of
CD3, DX5, and Ly-49 receptors on freshly isolated spleen cells. Most
DX5-positive lymphocytes were CD3-; however, 1 to 2%
of lymphocytes were CD3+, DX5+ T cells. A
similar percentage of CD3+, NK1.1+ T cells was
observed in B6 splenic lymphocytes (see Fig. 2A). Generally,
<3% of T cells express the DX5 marker in normal spleen. As can be
seen in Figure 1, the T cell population also contains small subsets of
cells that express Ly-49 receptors in B6 mice, with the exception of
Ly-49D.
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demonstrates the
advantage of this approach. As can be seen in Figure 2a, the
percentage of splenic NK1.1+ cells, both CD3+
and CD3-, remained similar, while in the liver, the
percentage of NK1.1+ cells dramatically increased after
IL-2 treatment. CD3-, NK1.1+ NK cells can
reach 50% of the total leukocyte isolate, while CD3+,
NK1.1+ T cells range from 9 to 20% of liver lymphocytes.
In addition, as shown in Figure 2B, not only was the
percentage of NK1.1+ increased in liver lymphocytes, but
the liver was also a site that sustained a dramatic increase in NK1.1
cell numbers. The number of NK1.1+ T cells was <1 x
106 cells/liver and increased to >1 x
106 after IL-2 treatment. The increase in percentage
coupled with the increase in lymphocyte yield resulted in this method
being an excellent procedure to recover high percentages and yields of
both CD3+ and CD3-, NK1.1+
lymphocytes for functional studies. Although the spleen contained a
similar number of total NK1.1+ cells, their frequency was
considerably lower (see Fig. 2A). Therefore, we
performed studies to determine whether these liver-derived cells could
be a good source of Ly-49-expressing T cells. As shown in Figure 3, the percentage of T cells expressing
Ly-49s was very low (often <1%) in spleen, but their percentage was
significantly increased (to 3–5%) in IL-2-treated mice. The
patterns of expression of Ly-49 molecules in normal and IL-2-treated
liver were similar; T cells expressed Ly-49A, C/I, and G2 but failed to
express the activating Ly-49D. These results demonstrate that increased
numbers of Ly-49+ T cells can be obtained from the livers
of IL-2-treated mice, offering a distinct advantage over normal
splenocytes.
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provides a summary of our studies
investigating the expression of the Ly-49 molecules on CD3+
spleen cells in various mouse strains. Similar to that in B6 mice, the
percentage of DX5+, CD3+ lymphocytes in these
various strains was quite low compared with that of the typical
DX5+, CD3- NK cells. In addition, the lack of
Ly-49D expression on T cells was consistent within all observed
strains. The regulation of expression of Ly-49 molecules is not
understood. Once expressed, Ly-49 receptors appear not to be regulated
on lymphocytes. The signal(s) that induces expression also is not
understood. Therefore, we sought to examine and compare T cells with NK
cells for the relative expression on DX5+ lymphocytes.
Figure 4 compares the percentages of
DX5+ T and NK cells expressing the various Ly-49 receptors.
Of interest was that both T and NK cells from the various haplotypes
appeared to have a similar level of expression. Strains whose NK cells
were high or low for expression of C/I (high: B6, DBA/2; low: AKR or
129/J) and G2 (high: B6, C3H/HeJ; low: AKR or 129/J) also demonstrated
concordant expression of T cell-associated Ly-49s. The exception was as
previously noted, that Ly-49D was not expressed on T cells. These
results are consistent with the hypothesis that these class I binding
receptors are similarly regulated in both DX5+ subsets.
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, both
TCRß+ and TCR+ cells express Ly-49
receptors. In addition, IL-2-treated mice exhibited small percentages
(0.7%) of TCR, Ly-49D+ cells. When the CD4 and CD8
coexpression with Ly-49 was examined, most Ly-49 molecules were
expressed preferentially on the CD8 T cell subset (2–4 times higher),
although some Ly-49 family members were found on the CD4 subset.
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presents two typical experiments using CD3+,
Ly-49G2+ cells as effectors for the lysis of P815
(H-2Dd) targets (often used to study the role of Ly-49
molecules in NK-mediated lysis). The effector function of the
CD3+, Ly-49G2+ cells against P815 was tested
either alone or in the presence of mAb specific for Ly-49G2 or
H-2Dd. Both T and NK effector cells potently killed the
prototypic NK target, YAC-1 (data not shown). As can be seen in Table III, the basal level of NK (CD3-, Ly-49G2+)
and T (CD3+ Ly-49G2+) cell-mediated lysis of
P815 was increased by the inclusion of Abs that reversed the inhibitory
signals mediated through Ly-49A and Ly-49G2 (Expt. I and II) or by Abs
to the appropriate class I, H-2Dd (Expt. II). Since the two
major inhibitory Ly-49s on H-2Db (B6) effectors were Ly-49A
and Ly-49G2 (both recognizing H-2Dd), they had a
complementary effect, reversing inhibition of lysis. In addition, an
increase in lytic capability was seen for both NK and T cells when
anti-class I Abs to the 1/2 domain of H-2Dd were
used, but not with other anti-class I Abs (17). Expt. III examines
the ability of CD3+, NK1.1+ lymphocytes to lyse
a B cell line, L5 MF22 (H-2Db), and an
H-2Dd-transfected line, L5cDd104
(H-2Db/d). When these NK1.1+ T cells were
tested against the parental H-2Db cell line, considerable
lysis was observed. However, this lysis was diminished in the presence
of H-2Dd (reduction from 46.6 to 31.4% lysis). The
inclusion of individual Abs to either Ly-49A or Ly-49G2 reversed the
lysis; however, the addition of both anti-Ly-49A and anti-G2
together or anti-H-2Dd 1/2 inhibited the
negative signal and reversed the lysis to the control
(H-2Db) level. Similar effects on the
H-2Dd-expressing line L5cDd104 were not seen
with mAb to Ly-49C or H-2Db. Therefore, the functional
inhibition of NK1.1+ T cells also was seen with an
H-2Dd-transfected B cell line. The inhibitory effect
mediated by Ly-49 molecules in NK cells also occurred with the
IL-2-induced spontaneous lysis in T cells. The inhibitory effects of
Ly-49A and G2 receptors on both NK and T cells were specifically
induced by Ab binding to effector cells, since anti-CD2 and
anti-H-2Db did not result in similar effects, thus
ruling out a reverse ADCC effect. In addition, anti-CD3 was used as
a positive control for T cells, since it has a potent, demonstrated
ability to cross-link the TCR and activate lysis.
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were used to test the ability of Ly-49A and Ly-49G2 to
regulate cytokine production during the effector-target interaction. T
cells obtained from H-2Db (B6) mice were cultured with
P815, either alone or in the presence of Abs specific for Ly-49A
(YE1–48), Ly-49G2 (4D11), Ly-49A and Ly-49G2, or H-2Dd.
The results of representative experiments are shown in Table IV. The pharmacologic stimuli, PMA and
ionomycin, were used as a positive control. The production of IFN-
and GM-CSF is reported as micrograms per milliliter produced after
18 h. Considerable amounts of IFN-, but not GM-CSF, were
induced by the coculture of both CD3- Ly-49+
and CD3+, Ly-49+ cells, presumably due to the
continued exposure to IL-2, as detected in effector cells cultured with
or without P815 target cells.
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production by lymphocytes cocultured with inhibitory targets is
frequently suboptimal because of the class I inhibitory interactions.
However, the addition of Abs specific for Ly-49G2 and Ly-49A resulted
in an increased level of cytokine production. In addition, elimination
of the negative signal was confirmed using anti-class I Abs to
H-2Dd. These effects were qualitatively similar for all
experiments, with differences of 
100 pg/ml IFN- or GM-CSF induced
by the addition of Ly-49A and Ly-49G2 or anti-H-2Dd.
These effects on T cells were not simply due to Ab binding to effector
cells, since Abs to other Ly-49 molecules, CD2 (not shown) and
H-2Db, did not result in similar effects, thus excluding a
nonspecific binding or reverse ADCC effect. In addition, anti-CD3
served as a positive control for the T cells.
The above data using short term cultured T cells demonstrate that minor
subsets of T cells express functional Ly-49 molecules. Since these
cells are minor subsets (<2%) of T cells, and it is difficult to
obtain adequate numbers of fresh cells for experiments, we examined
EL-4, a murine T cell line that expresses Ly-49 molecules. We found
that this cell line expressed high levels of Ly-49A and low levels of
Ly-49G2, with >99% of the cells being Ly-49A+ (not
shown). Therefore, we used EL-4 as a model to study the effects of the
Ly-49 molecules. Preliminary experiments demonstrated that
phosphorylation of Ly-49A and Ly-49G2 was induced by pervanadate
treatment (not shown), an effect similar to that observed for NK cells
and NK cell lines. Since EL-4 can produce IL-2, we also evaluated EL-4
for the ability of Ly-49A and Ly-49G2 receptors to inhibit IL-2
production during the interaction of EL-4 with P815 cells. A
representative experiment is shown in Figure 5. EL-4 spontaneously produces IL-2, and
this production was not altered by addition of rat IgG2a,
anti-Ly-49A, anti-Ly-49G2 (not shown), or anti-Ly-49A plus
anti-Ly-49G2 or anti-Ly-49C/I. However, the addition of P815
expressing an inhibitory class I molecule (H-2Dd) resulted
in diminished spontaneous production of IL-2 by EL-4 cells. The
addition of anti-Ly-49A and G2 reversed this inhibition, while
irrelevant IgG or anti-Ly-49C/I failed to block the inhibitory
signal being delivered from the H-2Dd on the P815 target.
An anti-CD3 Ab that cross-linked to P815 via its FcR served as a
positive control.
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. Calcium flux was induced by CD2 on
these EL-4 cells using anti-CD2. If the EL-4 cells were pretreated
with anti-Ly-49A (YE1/48; YE1/32) or anti-Ly-49G2 (4D11), a
delay in the kinetics and diminished intensity of calcium flux were
observed. However, the addition of irrelevant rat IgG or
anti-Ly-49D (4E5; not expressed on EL-4) had no effect on the
calcium mobilization. Thus, consistent with the functional alterations
observed above, the biochemical signaling and regulation in T cells via
Ly-49 molecules may be similar to those observed in NK cells.
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Discussion |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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Our examination of lymphocytes from normal mice indicated that the
Ly-49A, C/I, and G2 receptors may be coexpressed on T cells. When
Ly-49+ T cells are examined, they are primarily NK1.1
and/or DX5 positive. Expression of Ly-49 on T cells has been
demonstrated on both TCRß and TCR cells. In addition, when T
cells coexpress Ly-49 receptors, they are predominantly found in the
CD8+ subset, however, some CD4+ T cells can
also express Ly-49 receptors. Finally, analysis of the expression of
Ly-49 receptors in 11 strains of murine T cells strongly indicates that
the Ly-49 genes are similarly regulated in both NK and T cells. Strains
that exhibit high percentages of Ly-49 receptors on NK cells are
expressed in a parallel fashion in DX5+ T cells. Of
considerable interest was the apparent lack of expression of Ly-49D on
murine T cells. This potentially activating Ly-49D receptor is
abundantly expressed on NK cells from several mouse strains, but is
absent on their DX5+ T cells. The functional relevance of
this observation is presently unclear. Also, this study has
demonstrated for the first time the presence of functional Ly-49
molecules on B6 T cells. Ly-49G2 and Ly-49A, which have strong
inhibitory effects on NK cell functions, appear to function similarly
in T cells. As a percentage of the total T cell population, Ly-49
expression is quite low (<1%), and the functional role of these
unique cells in vivo is not understood. However, the selection and the
expansion of T cell populations in which high percentages of the cells
are NK1.1+ or DX5+ demonstrate that the
spontaneous lytic process of these cells can be regulated by
interactions with class I molecules. This effect was shown by addition
of Abs specific for Ly-49A and G2, which are known to inhibit
H-2Dd target cell lysis by B6 effectors, and also by
inclusion of Abs to H-2Dd on the target.
We have recently reported that receptors can inhibit cytokine
production by NK cells (28). In the present report we have examined
several activation pathways in T cells and analyzed the possible
inhibitory role of Ly-49 class I-binding receptors. The induction of
cytokines as a result of target cell interactions was shown to be
regulated by Ly-49 family members for both IFN- and GM-CSF. While
the addition of H-2Dd-expressing targets had no effect on
stimulation by anti-CD3 or by pharmacologic triggering with PMA and
ionomycin, no conclusions can be drawn from these limited data.
Although these findings were of interest, detailed studies with
anti-CD3 were beyond the scope of this study, and the present
experiments were not designed to examine the complex TCR signaling by
Ly-49s. In the present study anti-CD3 was used only as a positive
control for T cells, and the emphasis of the present study is target
cell interactions that result in cytokine production.
Cytokine production by NK cells is likely to be an important
physiologic event in a number of settings. For example, IFN-
production by NK cells is critical for antiviral effects against
leukochoriomeningitis virus (29). Recent studies have demonstrated
important marrow-regulating effects of NK cells in perforin and Fas
knockout mice (30). Collectively, these data support the hypothesis
that cytokine production by NK cells and perhaps by T cells might be
paramount in the regulation of hybrid resistance and marrow
transplantation. Our current data with NK and T cells also suggest that
Ly-49 receptors can regulate cytokines such as IFN- and GM-CSF,
consistent with the findings of these previous studies. The abilities
of Ly-49+ lymphocytes to recognize various class I
molecules on hemopoietic stem cells and subsequently regulate their
cytokine production could have a dramatic influence on stem cell
repopulation. However, the in vivo function of CD3+,
NK1.1+/DX5+ T cells and their potential role in
marrow transplantation are unclear. Most studies to date have employed
reagents such as NK1.1 and Abs to Ly-49 family members to modify marrow
engraftment in preclinical models. It is clear that these reagents will
not discriminate between NK and T cells in vivo. Therefore, a potential
role for the CD3+/DX5+ T cells may be a
redundancy with the classical NK cells for marrow engraftment, or these
cells may have some unique, as yet undiscovered, function. Further
studies to directly examine these alternatives are required. In humans,
class I binding killer cell-inhibitory receptors that regulate human T
cell functions have been reported on memory CD45RO+ T cells
(31). It is possible that the expression of Ly-49 receptors on T cells
during Ag stimulation might cause select subsets of cells to escape
apoptosis and become memory lymphocytes. Current studies are underway
in virus and tumor model systems to test this hypothesis.
In summary, the present study describes for the first time that Ly-49
class I-binding molecules that are known to inhibit both spontaneous
killing and cytokine production in NK cells also mediate these effects
on a subset of murine T cells. Our studies demonstrate that the
production of GM-CSF and IFN-, which are important hemopoietic
regulatory cytokines, is inhibited in this subset of T cells and T cell
lines via their interaction with class I on target cells. We speculate
that inhibitory effects mediated via these interactions may be
important for the success or failure of marrow grafts.
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Acknowledgments |
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Footnotes |
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2 Address correspondence and reprint requests to Dr. John R. Ortaldo, National Cancer Institute-Frederick Cancer Research and Development Center, Building 560, Room 31-93, Frederick, MD 21702-1201.
3 Abbreviations used in this paper: B6, C57BL/6; ADCC, Ab-dependent cellular cytotoxicity; FcR, Fc receptor; PE, phycoerythrin; CU, Cetus units; GM-CSF, granulocyte macrophage-CSF; [Ca2+]i, intracellular calcium concentration.
Received for publication June 23, 1997. Accepted for publication October 17, 1997.
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References |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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/ß+ cells: new clues to their origin, specificity, and function. J. Exp. Med. 182:633.This article has been cited by other articles:
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  S. Laffont, C. Seillet, J. Ortaldo, J. D. Coudert, and J.-C. Guery Natural killer cells recruited into lymph nodes inhibit alloreactive T-cell activation through perforin-mediated killing of donor allogeneic dendritic cells Blood, August 1, 2008; 112(3): 661 - 671. [Abstract] [Full Text] [PDF]
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 C. M. Watte, T. Nakamura, C. H. Lau, J. R. Ortaldo, and J. Stein-Streilein Ly49 C/I-dependent NKT cell-derived IL-10 is required for corneal graft survival and peripheral tolerance J. Leukoc. Biol., April 1, 2008; 83(4): 928 - 935. [Abstract] [Full Text] [PDF]
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 V. Soulard, J. Roland, C. Sellier, A. C. Gruner, M. Leite-de-Moraes, J.-F. Franetich, L. Renia, P.-A. Cazenave, and S. Pied Primary Infection of C57BL/6 Mice with Plasmodium yoelii Induces a Heterogeneous Response of NKT Cells Infect. Immun., May 1, 2007; 75(5): 2511 - 2522. [Abstract] [Full Text] [PDF]
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H. Kishimoto, T. Ohteki, N. Yajima, K. Kawahara, M. Natsui, S. Kawarasaki, K. Hamada, Y. Horie, Y. Kubo, S. Arase, et al. The Pten/PI3K pathway governs the homeostasis of V{alpha}14iNKT cells Blood, April 15, 2007; 109(8): 3316 - 3324. [Abstract] [Full Text] [PDF]
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 T. L. Denning, S. Granger, D. Mucida, R. Graddy, G. Leclercq, W. Zhang, K. Honey, J. P. Rasmussen, H. Cheroutre, A. Y. Rudensky, et al. Mouse TCR{alpha}beta+CD8{alpha}{alpha} Intraepithelial Lymphocytes Express Genes That Down-Regulate Their Antigen Reactivity and Suppress Immune Responses J. Immunol., April 1, 2007; 178(7): 4230 - 4239. [Abstract] [Full Text] [PDF]
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J. R. Ortaldo, R. Winkler-Pickett, J. Wigginton, M. Horner, E. W. Bere, A. T. Mason, N. Bhat, J. Cherry, M. Sanford, D. L. Hodge, et al. Regulation of ITAM-positive receptors: role of IL-12 and IL-18 Blood, February 15, 2006; 107(4): 1468 - 1475. [Abstract] [Full Text] [PDF]
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D. G. Pellicci, K. J. L. Hammond, J. Coquet, K. Kyparissoudis, A. G. Brooks, K. Kedzierska, R. Keating, S. Turner, S. Berzins, M. J. Smyth, et al. DX5/CD49b-Positive T Cells Are Not Synonymous with CD1d-Dependent NKT Cells J. Immunol., October 1, 2005; 175(7): 4416 - 4425. [Abstract] [Full Text] [PDF]
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F. Gays, K. Martin, R. Kenefeck, J. G. Aust, and C. G. Brooks Multiple Cytokines Regulate the NK Gene Complex-Encoded Receptor Repertoire of Mature NK Cells and T Cells J. Immunol., September 1, 2005; 175(5): 2938 - 2947. [Abstract] [Full Text] [PDF]
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 B.-C. Sheu, S.-H. Chiou, H.-H. Lin, S.-N. Chow, S.-C. Huang, H.-N. Ho, and S.-M. Hsu Up-regulation of Inhibitory Natural Killer Receptors CD94/NKG2A with Suppressed Intracellular Perforin Expression of Tumor-Infiltrating CD8+ T Lymphocytes in Human Cervical Carcinoma Cancer Res., April 1, 2005; 65(7): 2921 - 2929. [Abstract] [Full Text] [PDF]
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S. S. Smith, T. Patterson, and M. E. Pauza Transgenic Ly-49A Inhibits Antigen-Driven T Cell Activation and Delays Diabetes J. Immunol., April 1, 2005; 174(7): 3897 - 3905. [Abstract] [Full Text] [PDF]
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N. Anfossi, S. H. Robbins, S. Ugolini, P. Georgel, K. Hoebe, C. Bouneaud, C. Ronet, A. Kaser, C. B. DiCioccio, E. Tomasello, et al. Expansion and Function of CD8+ T Cells Expressing Ly49 Inhibitory Receptors Specific for MHC Class I Molecules J. Immunol., September 15, 2004; 173(6): 3773 - 3782. [Abstract] [Full Text] [PDF]
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 I. Bubic, M. Wagner, A. Krmpotic, T. Saulig, S. Kim, W. M. Yokoyama, S. Jonjic, and U. H. Koszinowski Gain of Virulence Caused by Loss of a Gene in Murine Cytomegalovirus J. Virol., July 15, 2004; 78(14): 7536 - 7544. [Abstract] [Full Text] [PDF]
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F. D. Finkelman, M. Yang, T. Orekhova, E. Clyne, J. Bernstein, M. Whitekus, D. Diaz-Sanchez, and S. C. Morris Diesel Exhaust Particles Suppress In Vivo IFN-{gamma} Production by Inhibiting Cytokine Effects on NK and NKT Cells J. Immunol., March 15, 2004; 172(6): 3808 - 3813. [Abstract] [Full Text] [PDF]
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L. Saurer, I. Seibold, C. Vallan, W. Held, and C. Mueller Cutting Edge: Stimulation with the Cognate Self-Antigen Induces Expression of the Ly49A Receptor on Self-Reactive T Cells Which Modulates Their Responsiveness J. Immunol., December 15, 2003; 171(12): 6334 - 6338. [Abstract] [Full Text] [PDF]
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B. Johnston, C. H. Kim, D. Soler, M. Emoto, and E. C. Butcher Differential Chemokine Responses and Homing Patterns of Murine TCR{alpha}{beta} NKT Cell Subsets J. Immunol., September 15, 2003; 171(6): 2960 - 2969. [Abstract] [Full Text] [PDF]
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S. H. Robbins, S. C. Terrizzi, B. C. Sydora, T. Mikayama, and L. Brossay Differential Regulation of Killer Cell Lectin-Like Receptor G1 Expression on T Cells J. Immunol., June 15, 2003; 170(12): 5876 - 5885. [Abstract] [Full Text] [PDF]
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 R. B. Voyle, F. Beermann, R. K. Lees, J. Schumann, J. Zimmer, W. Held, and H. R. MacDonald Ligand-dependent Inhibition of CD1d-restricted NKT Cell Development in Mice Transgenic for the Activating Receptor Ly49D J. Exp. Med., April 7, 2003; 197(7): 919 - 925. [Abstract] [Full Text] [PDF]
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J. R. Ortaldo and H. A. Young Expression of IFN-{gamma} Upon Triggering of Activating Ly49D NK Receptors In Vitro and In Vivo: Costimulation with IL-12 or IL-18 Overrides Inhibitory Receptors J. Immunol., February 15, 2003; 170(4): 1763 - 1769. [Abstract] [Full Text] [PDF]
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M. A. Morris, J. Liu, V. Arora, T. C. George, J. Klem, J. D. Schatzle, V. Kumar, and M. Bennett B6 Strain Ly49I Inhibitory Gene Expression on T Cells in FVB.Ly49IB6 Transgenic Mice Fails to Prevent Normal T Cell Functions J. Immunol., October 1, 2002; 169(7): 3661 - 3666. [Abstract] [Full Text] [PDF]
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C. W. McMahon, A. J. Zajac, A. M. Jamieson, L. Corral, G. E. Hammer, R. Ahmed, and D. H. Raulet Viral and Bacterial Infections Induce Expression of Multiple NK Cell Receptors in Responding CD8+ T Cells J. Immunol., August 1, 2002; 169(3): 1444 - 1452. [Abstract] [Full Text] [PDF]
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S. Korten, L. Volkmann, M. Saeftel, K. Fischer, M. Taniguchi, B. Fleischer, and A. Hoerauf Expansion of NK Cells with Reduction of Their Inhibitory Ly-49A, Ly-49C, and Ly-49G2 Receptor-Expressing Subsets in a Murine Helminth Infection: Contribution to Parasite Control J. Immunol., May 15, 2002; 168(10): 5199 - 5206. [Abstract] [Full Text] [PDF]
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K. Van Beneden, A. De Creus, F. Stevenaert, V. Debacker, J. Plum, and G. Leclercq Expression of Inhibitory Receptors Ly49E and CD94/NKG2 on Fetal Thymic and Adult Epidermal TCR V{gamma}3 Lymphocytes J. Immunol., April 1, 2002; 168(7): 3295 - 3302. [Abstract] [Full Text] [PDF]
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A. O. Dokun, D. T. Chu, L. Yang, A. S. Bendelac, and W. M. Yokoyama Analysis of In Situ NK Cell Responses During Viral Infection J. Immunol., November 1, 2001; 167(9): 5286 - 5293. [Abstract] [Full Text] [PDF]
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M. Maeda, S. Lohwasser, T. Yamamura, and F. Takei Regulation of NKT Cells by Ly49: Analysis of Primary NKT Cells and Generation of NKT Cell Line J. Immunol., October 15, 2001; 167(8): 4180 - 4186. [Abstract] [Full Text] [PDF]
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 S. H. More, M. Breloer, and A. von Bonin Eukaryotic heat shock proteins as molecular links in innate and adaptive immune responses: Hsp60-mediated activation of cytotoxic T cells Int. Immunol., September 1, 2001; 13(9): 1121 - 1127. [Abstract] [Full Text] [PDF]
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K. J. L. Hammond, D. G. Pellicci, L. D. Poulton, O. V. Naidenko, A. A. Scalzo, A. G. Baxter, and D. I. Godfrey CD1d-Restricted NKT Cells: An Interstrain Comparison J. Immunol., August 1, 2001; 167(3): 1164 - 1173. [Abstract] [Full Text] [PDF]
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J. Roger, A. Chalifour, S. Lemieux, and P. Duplay Cutting Edge: Ly49A Inhibits TCR/CD3-Induced Apoptosis and IL-2 Secretion J. Immunol., July 1, 2001; 167(1): 6 - 10. [Abstract] [Full Text] [PDF]
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L. Fahlen, U. Lendahl, and C. L. Sentman MHC Class I-Ly49 Interactions Shape the Ly49 Repertoire on Murine NK Cells J. Immunol., June 1, 2001; 166(11): 6585 - 6592. [Abstract] [Full Text] [PDF]
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P. C. Sandel, M. Gendelman, G. Kelsoe, and J. G. Monroe Definition of a Novel Cellular Constituent of the Bone Marrow That Regulates the Response of Immature B Cells to B Cell Antigen Receptor Engagement J. Immunol., May 15, 2001; 166(10): 5935 - 5944. [Abstract] [Full Text] [PDF]
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B. Kunz and W. Held Positive and Negative Roles of the Trans-Acting T Cell Factor-1 for the Acquisition of Distinct Ly-49 MHC Class I Receptors by NK Cells J. Immunol., May 15, 2001; 166(10): 6181 - 6187. [Abstract] [Full Text] [PDF]
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J. R. Ortaldo, E. W. Bere, D. Hodge, and H. A. Young Activating Ly-49 NK Receptors: Central Role in Cytokine and Chemokine Production J. Immunol., April 15, 2001; 166(8): 4994 - 4999. [Abstract] [Full Text] [PDF]
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K. Van Beneden, F. Stevenaert, A. De Creus, V. Debacker, J. De Boever, J. Plum, and G. Leclercq Expression of Ly49E and CD94/NKG2 on Fetal and Adult NK Cells J. Immunol., April 1, 2001; 166(7): 4302 - 4311. [Abstract] [Full Text] [PDF]
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T. Hanke and D. H. Raulet Cumulative Inhibition of NK Cells and T Cells Resulting from Engagement of Multiple Inhibitory Ly49 Receptors J. Immunol., March 1, 2001; 166(5): 3002 - 3007. [Abstract] [Full Text] [PDF]
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J. R. Ortaldo, R. Winkler-Pickett, and G. Wiegand Activating Ly-49D NK receptors: expression and function in relation to ontogeny and Ly-49 inhibitor receptors J. Leukoc. Biol., November 1, 2000; 68(5): 748 - 756. [Abstract] [Full Text]
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E. Assarsson, T. Kambayashi, J. K. Sandberg, S. Hong, M. Taniguchi, L. Van Kaer, H.-G. Ljunggren, and B. J. Chambers CD8+ T Cells Rapidly Acquire NK1.1 and NK Cell-Associated Molecules Upon Stimulation In Vitro and In Vivo J. Immunol., October 1, 2000; 165(7): 3673 - 3679. [Abstract] [Full Text] [PDF]
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E. T. Silver, D.-E. Gong, C. S. Chang, A. Amrani, P. Santamaria, and K. P. Kane Ly-49P Activates NK-Mediated Lysis by Recognizing H-2Dd 1 J. Immunol., August 15, 2000; 165(4): 1771 - 1781. [Abstract] [Full Text] [PDF]
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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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C. D. Peacock, M. Y. Lin, J. R. Ortaldo, and R. M. Welsh The Virus-Specific and Allospecific Cytotoxic T-Lymphocyte Response to Lymphocytic Choriomeningitis Virus Is Modified in a Subpopulation of CD8+ T Cells Coexpressing the Inhibitory Major Histocompatibility Complex Class I Receptor Ly49G2 J. Virol., August 1, 2000; 74(15): 7032 - 7038. [Abstract] [Full Text]
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F. Gays, M. Unnikrishnan, S. Shrestha, K. P. Fraser, A. R. Brown, C. M. G. Tristram, Z. M. A. Chrzanowska-Lightowlers, and C. G. Brooks The Mouse Tumor Cell Lines EL4 and RMA Display Mosaic Expression of NK-Related and Certain Other Surface Molecules and Appear to Have a Common Origin J. Immunol., May 15, 2000; 164(10): 5094 - 5102. [Abstract] [Full Text] [PDF]
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H. R.C. Smith, H. H. Chuang, L. L. Wang, M. Salcedo, J. W. Heusel, and W. M. Yokoyama Nonstochastic Coexpression of Activation Receptors on Murine Natural Killer Cells J. Exp. Med., April 17, 2000; 191(8): 1341 - 1354. [Abstract] [Full Text] [PDF]
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M. K. Slifka, R. R. Pagarigan, and J. L. Whitton NK Markers Are Expressed on a High Percentage of Virus-Specific CD8+ and CD4+ T Cells J. Immunol., February 15, 2000; 164(4): 2009 - 2015. [Abstract] [Full Text] [PDF]
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L. Fahlen, L. Oberg, T. Brannstrom, N. K. S. Khoo, U. Lendahl, and C. L. Sentman Ly49A expression on T cells alters T cell selection Int. Immunol., February 1, 2000; 12(2): 215 - 222. [Abstract] [Full Text] [PDF]
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S. Pied, J. Roland, A. Louise, D. Voegtle, V. Soulard, D. Mazier, and P.-A. Cazenave Liver CD4-CD8- NK1.1+ TCR{alpha}{beta} Intermediate Cells Increase During Experimental Malaria Infection and Are Able to Exhibit Inhibitory Activity Against the Parasite Liver Stage In Vitro J. Immunol., February 1, 2000; 164(3): 1463 - 1469. [Abstract] [Full Text] [PDF]
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L. H. Mason, J. Willette-Brown, A. T. Mason, D. McVicar, and J. R. Ortaldo Interaction of Ly-49D+ NK Cells with H-2Dd Target Cells Leads to Dap-12 Phosphorylation and IFN-{gamma} Secretion J. Immunol., January 15, 2000; 164(2): 603 - 611. [Abstract] [Full Text] [PDF]
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M. Pauza, K. M. Smith, H. Neal, C. Reilly, L. L. Lanier, and D. Lo Transgenic Expression of Ly-49A in Thymocytes Alters Repertoire Selection J. Immunol., January 15, 2000; 164(2): 884 - 892. [Abstract] [Full Text] [PDF]
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J. R. Ortaldo, R. Winkler-Pickett, J. Willette-Brown, R. L. Wange, S. K. Anderson, G. J. Palumbo, L. H. Mason, and D. W. McVicar Structure/Function Relationship of Activating Ly-49D and Inhibitory Ly-49G2 NK Receptors ,2 J. Immunol., November 15, 1999; 163(10): 5269 - 5277. [Abstract] [Full Text] [PDF]
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A. J. Zajac, R. E. Vance, W. Held, D. J. D. Sourdive, J. D. Altman, D. H. Raulet, and R. Ahmed Impaired Anti-Viral T Cell Responses Due to Expression of the LY49A Inhibitory Receptor J. Immunol., November 15, 1999; 163(10): 5526 - 5534. [Abstract] [Full Text] [PDF]
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A. M. Moodycliffe, S. Maiti, and S. E. Ullrich Splenic NK1.1-Negative, TCR{alpha}{beta} Intermediate CD4+ T Cells Exist in Naive NK1.1 Allelic Positive and Negative Mice, with the Capacity to Rapidly Secrete Large Amounts of IL-4 and IFN-{gamma} Upon Primary TCR Stimulation J. Immunol., May 1, 1999; 162(9): 5156 - 5163. [Abstract] [Full Text] [PDF]
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 I. Apostolou, Y. Takahama, C. Belmant, T. Kawano, M. Huerre, G. Marchal, J. Cui, M. Taniguchi, H. Nakauchi, J.-J. Fournie, et al. Murine natural killer cells contribute to the granulomatous reaction caused by mycobacterial cell walls PNAS, April 27, 1999; 96(9): 5141 - 5146. [Abstract] [Full Text] [PDF]
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C. S. Chang, L. Brossay, M. Kronenberg, and K. P. Kane The Murine Nonclassical Class I Major Histocompatibility Complex-like CD1.1 Molecule Protects Target Cells from Lymphokine-activated Killer Cell Cytolysis J. Exp. Med., February 1, 1999; 189(3): 483 - 491. [Abstract] [Full Text] [PDF]
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N. Seo, Y. Tokura, F. Furukawa, and M. Takigawa Down-Regulation of Tumoricidal NK and NK T Cell Activities by MHC Kb Molecules Expressed on Th2-Type {gamma}{delta} T and {alpha}{beta} T Cells Coinfiltrating in Early B16 Melanoma Lesions J. Immunol., October 15, 1998; 161(8): 4138 - 4145. [Abstract] [Full Text] [PDF]
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H. Robson MacDonald, R. K. Lees, and W. Held Developmentally Regulated Extinction of Ly-49 Receptor Expression Permits Maturation and Selection of NK1.1+ T Cells J. Exp. Med., June 15, 1998; 187(12): 2109 - 2114. [Abstract] [Full Text] [PDF]
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) and
DX5+ (CD3+, DX5+; 
) T cells were
examined for expression of Ly-49s. Data represent the percentage of
Ly-49s that are expressed on the DX5+ subsets for Ly-49A
(A), Ly-49C/I (B),
Ly-49D (C), and Ly-49G2
(D).
