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Departments of
*
Molecular Immunology and
Molecular Embryology, Graduate School of Medicine, Chiba University, Inohana, Chiba, Japan;
Immunology Program, Virginia Mason Research Center, Seattle, WA 98101; and
Department of Immunology, University of Washington School of Medicine, Seattle, WA 98195
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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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In addition to the changes in the expression of cell surface markers, thymocytes traverse through the thymus as they develop. Immature thymocytes reside in the cortex, and as they begin the selection process they traffic from the cortex to the medulla. This transition is marked by the expression of other specific molecules. For example, the chemokine receptor CCR4 is up-regulated in thymocytes as they migrate from the cortex to the medulla (6, 7, 8). This in contrast to CCR9, which is expressed on both cortical and medullary thymocytes, but is down-modulated as thymocytes leave the thymus for the periphery (6, 8). These data suggest that chemokine/chemokine receptor interactions are involved in the migration of thymocyte subsets.
In addition to chemokine receptors, other cell surface molecules are expressed in a restricted fashion on developing thymocytes. One of these is the activation marker CD69, which is expressed at high levels on approximately 10–15% of thymocytes. Expression of CD69 is first seen on DP thymocytes as they begin positive selection, and experiments using TCR transgenic mice showed that only those thymocytes being selected express CD69 (9, 10). Several lines of evidence strongly suggest an active role for CD69 in TCR-mediated positive selection of thymocytes. All TCR+ thymocytes are CD69+, including both TCRlow DP cells and TCRhigh SP cells (5). These CD69+ cells can be further subdivided by expression of heat-stable Ag (HSA; HSA+TCRlow, HSA+TCRhigh, and HSA-TCRhigh); however, CD69+ cells do not express the mature T cell marker Qa-2 (5). Also, using an in vitro reaggregation system, Hare et al. (11) showed that CD69 may regulate an MHC-independent aspect of positive selection, suggesting that CD69 is not merely a marker for cells that have begun the selective process. Thus, CD69-expressing cells represent a population that is undergoing positive selection or has just completed that process.
Despite the compelling circumstantial evidence that CD69 is actively involved in thymocyte development and selection, there is no direct evidence. We have used overexpression and blockade of CD69 to directly test its role in thymocyte development, and demonstrate that CD69 plays a role in thymocyte selection.
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Materials and Methods |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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The Abs used in this study were as follows:
anti-TCR
-FITC (H57-597), anti-CD3
-FITC (145-2C11),
anti-CD69-FITC (H1.2F3), anti-CD25-FITC (7D4), anti-CD8-APC
(53-6.72), anti-CD4-PE (GK1.5), anti-CD44-FITC (KM114),
anti-V5-biotin (MR9-4), and anti-Qa-2-biotin (1-1-2) from BD
PharMingen (La Jolla, CA); and anti-CD62L-FITC (MEL-14),
anti-CD45RB-PE (16A), and anti-HSA-FITC (J11d) from Caltag
Laboratories (Burlingame, CA). Clonotype anti-DO11.10 TCR (KJ1-26)
was a gift from Dr. P. Marrack (National Jewish Hospital, Denver,
CO).
For Ab staining, single-cell suspensions were prepared from freshly isolated lymphoid organs from control and experimental animals and suspended in PBS supplemented with 2% FCS and 0.1% sodium azide. In general, 106 cells were incubated on ice for 30 min with appropriate staining reagents as previously described (12). For direct staining, cells were first incubated with 2.4G2 (from ascites) to prevent nonspecific binding of mAbs via FcR interactions. Flow cytometric analysis was performed on either FACSCalibur or FACSvantage (BD Immunocytometry Systems, Mountain View, CA).
In vivo administration of Abs
Newborn C57BL/6 mice were treated daily for 7 days beginning at day 0 (within 24 h of birth) by the i.p. injection of 200 µg purified an anti-I-Ak mAb (10-2.16, BD PharMingen, as a negative control), an anti-I-Ab mAb (M5/114, BD PharMingen), or an anti-CD69 mAb (H1.2F3, BD PharMingen) suspended in PBS. In some experiments 2-wk-old C57BL/6 mice were treated daily for 7 days with 500 µg anti-CD69 (three mice per group) for 7 days. Then, the thymocytes were prepared and subjected to FCM analysis.
Generation of CD69 transgenic mice
CD69FL. A cDNA clone encoding the entire coding region of mouse CD69 (13) was subcloned into p1017, which contains the proximal mouse lck gene promoter and the human growth hormone gene (14). The transgene was excised with NotI and used for pronuclear injections.
CD69 
cyt.
A cDNA construct encoding a cytoplasmic-deletion mutant of mouse CD69
was generated by PCR using the primers
5'-agatctATGGAAGGATCCATTCAAGTT-3' (amino terminus) and
5'-agatctTCATCTGGAGGGCTTGCTGCA-3' (carboxyl terminus). The amino
terminal primer begins, after the initiating ATG, at codon 34 of mouse
CD69. The final construct contains three codons from the cytoplasmic
domain of mouse CD69 (the initiating ATG and the final two amino acids
of the cytoplasmic domain) preceding the membrane-spanning sequence and
extracellular domain. The resulting product was cloned into p1017 and
injected as described above.
Double-transgenic mice (CD69FL and either the OT-II or DO11.10 TCR transgene) were generated by crossing the appropriate single-transgenic lines and screening for the individual transgenes. In all cases transgene-positive mice were identified by PCR using primers to the human growth hormone gene as previously described (14).
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Results |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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The up-regulation of CD69 expression during thymocyte selection
suggests a role for CD69 in the generation of mature, SP thymocytes. To
begin an analysis of the role of CD69 in thymocyte development, newborn
C57BL/6 mice were injected daily for 1 wk with 200 µg anti-CD69
mAb. Control mice were injected with PBS, an irrelevant MHC class II Ab
(anti-I-Ak), or the relevant MHC class II Ab
(anti-I-Ab). The latter mice served as a
control for the ability to block the generation of CD4SP cells. As
shown in Fig. 1
the control mice
displayed the anticipated phenotypes following Ab treatment: the
anti-I-Ab-treated mice showed a decrease in
the number of CD4SP cells relative to the PBS-treated mice, while the
anti-I-Ak-treated mice showed no difference
compared with the PBS controls (Fig. 1A). The effect of
anti-I-Ab treatment was specific to CD4SP
development as the number of CD8SP cells was unaffected.
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A). Further characterization of these mice showed
that the numbers of
Qa-2+HSA- cells as well as
TCR+HSA- cells, were
reduced, consistent with a reduction of SP thymocytes (Fig. 1B). In all cases thymic cellularity was unchanged,
suggesting that the reduction seen in anti-CD69-treated mice was
not due to nonspecific lysis of CD69-expressing cells. Overexpression of CD69 in the thymus results in an increase in SP thymocytes
The data presented above support a role for CD69 in the generation of SP thymocytes. We next determined the effect of CD69 overexpression on thymocyte development. A cDNA encoding full-length mouse CD69 was expressed from the lck-proximal promoter in transgenic animals (these mice will be referred to as CD69FL). Several founder lines were established that expressed varying levels of CD69 in the thymus, as judged by cell surface expression.
As shown in Table I, increased levels of
CD69 expression in the thymus tended to skew the developmental profile
seen in these mice, with the lines expressing the highest levels of
CD69 showing increased numbers of CD4SP and CD8SP cells. Fig. 2 shows the analysis of a representative
line, CD69FL-1. Examination of CD69 expression in this line showed that
it was markedly increased relative to that in littermate controls, with
approximately 99% of thymocytes expressing CD69 (Fig. 2A).
When CD3 levels on thymocytes from CD69FL-1 and NLC mice were analyzed,
the CD69FL-1 line displayed a higher percentage of
CD3high cells (Fig. 2B). Also, there
appeared to be two populations of CD3high cells
in CD69FL-1 mice, with a small shoulder of cells with slightly lower
CD3 levels and a second population with higher CD3 levels (also see
Fig. 4). The peak representing the highest level of CD3 expression was
CD4SP (gate 1) cells, while the lower peak contained CD8SP cells (gate
2). This pattern of CD3 expression is similar to that seen when CD3
levels are analyzed on mature splenic T cells. We next examined the
CD4/CD8 profile of CD69FL-1 and NLC mice. There was a dramatic increase
in the number of CD4SP and CD8SP cells and a diminution of CD4CD8DP
cells in CD69FL-1 mice (Fig. 2C), with approximately 37% of
the thymocytes from CD69FL-1 mice showing an SP phenotype compared with
17% for the NLC. There was a concomitant decrease in DP cells in
CD69Fl-1 mice, and overall thymic cellularity was comparable between
the two sets of mice, suggesting that an increase in total cell numbers
cannot account for this difference in SP cells. We have now examined
four CD69FL lines, and all show a similar phenotype, with the number of
SP thymocytes proportional to the level of CD69 expression (number of
SP cells varies from 25 to 58%; data not shown).
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shows the expression of
these markers on CD4SP cells from CD69FL-1 and NLC mice. CD4SPs from
the control animals had an
HSA+Qa-2-CD45RBlowCD62Llow
phenotype. In contrast, the cell surface phenotype of the CD4SP cells
from CD69FL-1 mice more closely resembles that of mature, peripheral T
cells
(HSA-Qa-2+CD45RBhighCD62Lhigh).
The CD8SP cells displayed a similar phenotype (data not shown).
Finally, thymocytes from the CD69 transgenic mice have a higher
proliferative capacity, compared with control mice, when stimulated
through the TCR (data not shown). Taken as a whole, these data
demonstrate that overexpression of full-length CD69 in the thymus
results in the accumulation of SP cells in the thymus that more closely
resemble mature, peripheral T cells then SP thymocytes.
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To determine whether the phenotype seen in the CD69-overexpressing
transgenic animals required CD69-mediated signal transduction, we
generated mice expressing a cytoplasmically truncated form of CD69 from
the lck-proximal promoter (see Materials and
Methods for details of the construct). Several founder lines were
generated, and T cell development was studied in four lines
(CD69cyt-1–4) that displayed CD69 levels roughly equivalent to
those seen in the CD69FL lines (Figs. 2A and 4A
and data not shown). As was seen with CD69FL-1 mice, CD69cyt-1 and
littermate control mice displayed similar thymic cellularity. Thymocyte
development in CD69cyt mice was also very similar to that seen in
CD69FL-1 mice. For example, the CD69cyt-1 line showed the two
populations of CD3high cells seen in CD69FL mice
(Figs. 2B and 4B), as well as increases in both
CD4SP and CD8SP cells (Fig. 4C), although the CD4SP/CD8SP
ratio in CD69Dcyt mice differed from that in CD69FL-1 mice. The
significance of this last finding is unclear at this time. Thus, the
ability of CD69 to affect the DP to SP transition does not necessarily
require a signal through CD69. This suggests that overexpression of the
extracellular domain of CD69 is the major cause of the phenotypes seen
in the CD69FL and CD69cyt transgenic mice, possibly by affecting
interactions with its ligand. However, the severity of the phenotype
seen in CD69FL mice was greater than that seen in CD69cyt mice,
demonstrating a possible role for CD69-mediated signals.
Positive and negative selection in CD69FL mice
A possible explanation for the phenotype seen in CD69FL-1 mice is that overexpression of CD69 leads to enhanced positive selection, and thus an increase in the number of SP cells. To directly study the role of CD69 on the positive and negative selection of thymocytes, we crossed CD69FL-1 mice with two TCR transgenic mouse lines, OT-II and DO11.10. Both TCRs are specific for the same chicken OVA peptide (OVA323–332) (15). OT-II-expressing T cells recognize this peptide presented in the context of I-Ab, while DO11.10-expressing T cells see it presented in the context of I-Ad. However, T cells expressing the DO11.10 TCR also are alloreactive against I-Ab (16), and thymocytes bearing the transgenic TCR are negatively selected in H-2b mice (17). Thus, both positive and negative thymocyte selection in CD69FL-1 mice, which are on the C57BL/6 background, can be examined using these two TCR transgenic lines.
To generate CD69FL/OT-II double-transgenic mice, we crossed the
CD69FL-1 line with C57BL/6 mice expressing the OT-II TCR transgene
(these mice will be referred to as OT-II). There was no significant
difference in thymic cellularity between the two strains (Fig. 5). Thymocyte development was studied in
OT-II and CD69FL-1/OT-II F1 mice, using Abs
against CD4, CD8, CD3, and V5 (the -chain of the transgenic TCR).
As shown in Fig. 5, the OT-II mice showed a skewing toward CD4SP cells,
reflecting positive selection of the OT-II TCR-expressing thymocytes.
CD69/OT-II mice displayed a greater skewing toward the development of
CDSP cells, with greater than 60% of thymocytes
CD4+/V5+ (Fig. 5). We
have also examined the ability of CD4SP cells from these mice to
respond to Ag in a proliferation assay. CD4+
cells were purified from OT-II and CD69/OT-II mice and cultured with
irradiated C57BL/6 splenocytes in the presence of the antigenic
peptide. No difference was found between OT-II- and CD69/OT-II-derived
cells (data not shown). Taken as a whole, these data show that the
process of positive selection proceeds at an enhanced rate in mice
overexpressing CD69 in the thymus, and that the selected CD4SP cells
respond normally to antigenic stimulation.
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, with
CD69FL-expressing mice showing an increase in SP cells and a
concomitant decrease in DP cells (data not shown). An analysis of DO11
and CD69/D011 mice is shown in Fig. 6.
DO11 mice displayed a decrease in thymus cellularity (29 x
106 for DO11, compared with 156 x
106 for NLC mice) along with a marked increase in
DN cells and a decrease in DP cells.
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cyt, mice expressing both the DO11.10 TCR transgene and the
CD69cyt transgene displayed the same overall thymic phenotype as
CD69/DO11 mice (data not shown).
We next examined the expression of DO11.10 TCR in these mice. As shown
in Fig. 6, both lines had similar numbers of CD3+
cells. When the expression of DO11.10 TCR was examined using the
clonotypic Ab KJ1-26 (17), both DO11 and CD69/DO11 mice
showed a reduction in the number of clonotype-positive cells. DO11.10
TCR expression was then examined in individual thymic subpopulations of
both sets of animals. CD4SP cells from both DO11 and CD69/DO11 mice
showed a dramatic reduction in KJ1-26 staining, suggesting that the
clonotype-positive cells had been negatively selected. However, as
mentioned above, CD69/DO11 mice had more CD4SP cells than DO11 mice
(Fig. 6). These cells do not express the transgenic TCR and most likely
arise through rearrangement of endogenous TCR genes. This finding is
consistent with the increased accumulation of CD4SP cells in CD69FL
mice (Fig. 2).
We next examined the expression of the transgenic TCR in the DN
compartment in both mice. A majority of DN cells in DO11 mice expressed
CD3 and were KJ1-26+ (Fig. 6). Earlier work had
shown that these KJ1-26+,
CD4-CD8- cells
represented a lineage that bypassed the DP stage (16). In
contrast, most of the DN cells from the CD69/DO11 mice were
CD3- and KJ1-26-.
Mice expressing CD69 transgenes had reduced T cells in peripheral lymphoid organs
As shown above, thymi from mice that overexpress CD69, either
full-length or containing a cytoplasmic truncation, displayed a
dramatic increase in SP cells (Figs. 2 and 4). These cells had the cell
surface characteristics of mature T cells
(TCRhighQa-2+HSA-;
Fig. 3 and data not shown). One possible explanation for this phenotype
is that following maturation the SP cells are incapable of exiting the
thymus. If this explanation were correct, one prediction would be a
reduction in the number of peripheral T cells. We tested this
hypothesis by examining CD3 levels and CD4/CD8 profiles of spleens and
lymph nodes isolated from CD69FL and CD69cyt mice. It was apparent
that the spleens and lymph nodes from both sets of animals were
significantly smaller than those in the control animals, and this
observation was supported by CD3 and CD4/CD8 profiles from each animal.
As shown in Fig. 7A, CD69FL-1
mice had dramatically reduced numbers of T cells in spleen and almost
no T cells in lymph node (data not shown). Concomitantly, the number of
CD3+ cells was severely reduced (data not shown).
We also examined T cells in spleens and lymph nodes of CD69cyt
animals. Similar to what we observed in CD69FL-1 animals, spleens and
lymph nodes in CD69cyt-1 were smaller than those in littermate
control mice. However, CD69cyt-1 mice had a 50% reduction of
peripheral T cells number (Fig. 7B), a less severe loss than
what was seen in CD69FL-1 mice. This finding was consistent with the
less severe thymic phenotype seen in these mice (Fig. 4) and suggests a
role for CD69 signaling in the phenotypes seen in these two sets
of mice.
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C). Splenic cellularity was comparable in the
two mice. OT-II mice showed an increase in CD4+
cells in the spleen, nearly all of which were
V5+ (data not shown). However,
spleens from CD69/OT-II mice were nearly devoid of T cells. Subsequent
analysis determined that the cellularity in the spleen in these animals
was maintained by an increase in the number of B cells (data not
shown). The reduction in the number of peripheral T cells in OT-II/CD69
mice was greater than that in CD69FL-1 mice. |
Discussion |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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). While these data strongly suggest that blockade
of CD69 inhibits SP development, we cannot rule out that cross-linking
CD69 with the mAb results in the deletion of SP thymocytes, or that the
lower levels of SP cells reflects phagocytosis of Ab-coated cells. To
address the effect of CD69 blockade more directly, we have generated
transgenic mice expressing a secreted form of the CD69 extracellular
domain. A preliminary analysis of thymocyte development in these mice
has suggested that they are similar to what was observed in Ab-treated
animals, supporting the hypothesis that CD69 blockade inhibits the
development of SP thymocytes (data not shown).
In contrast to what was observed in mice in which CD69 interactions
were blocked, mice that overexpressed CD69 in the thymus displayed a
dramatic increase in the numbers of CD4SP and CD8SP cells (Figs. 2 and 4 and Table I). In addition, these cells had the phenotype of mature T
cells in that they were
TCRhighQa-2+HSA-.
This is the phenotype seen on T cells as they leave the thymus and seed
the periphery (5). The fact that this population is
present in large numbers in the thymus of CD69 transgenic mice, but not
in littermate control mice, suggests that it is the continued
expression of CD69 that affects the developmental profile of these
cells. Consistent with a more mature phenotype, CD4SP cells from CD69FL
mice were more responsive to TCR engagement (D. J. Kasprowicz and
S. F. Ziegler, unpublished observation).
The accumulation of SP cells, at the expense of DP cells, in CD69FL
mice suggests differences in thymic selection between these animals. We
tested this hypothesis by examining the role of CD69 in the selection
events using mice expressing transgenic TCRs that were either
positively or negatively selected on the C57BL/6 background. The data
generated from these double-transgenic mice supports the hypothesis
that overall thymic selection was more efficient in mice overexpressing
CD69 (Figs. 5 and 6). For example, double-transgenic mice expressing
either positively or negatively selected TCRs had similar cellularity
as their single TCR transgenic littermates. The positively selecting
mice (CD69/OT-II) had twice as many CD4SP cells as the OT-II mice, all
of which expressed the TCR transgene (Fig. 5, bottom panel).
The negatively selecting mice (CD69/DO11) also displayed an increase in
CD4SP cells relative to the single TCR transgenic line. However, these
CD4SP cells did not express the transgenic TCR (as shown by staining
with KJ1-26; Fig. 6, bottom panel), suggesting that they
arose from cells that rearranged endogenous TCR genes. Those
thymocytes expressing the DO11.10 TCR were deleted. These data suggest
that, similar to the CD69FL-1 mice, thymocytes in both sets of double
transgenic mice were being driven to the SP stage. In the CD69/OT-II
mice, these SP cells were selected and accumulated. In CD69/DO11.10
mice those thymocytes that expressed the TCR transgene were deleted,
and those that were able to rearrange and express endogenous TCR
-chains accumulated. Consistent with this was our finding that the
spleens of DO11 mice contained
CD4-CD8-DO11.10+
cells, as has been previously reported when this TCR transgene is
expressed on a H-2b background (16).
In CD69/DO11 mice these cells were not present in the periphery, and
these mice, similar to CD69FL-1, had a dramatic reduction in the number
of peripheral T cells (data not shown).
To assess whether CD69 signal transduction was involved in thymocyte
development, we generated mice overexpressing a cytoplasmic mutant of
CD69 (CD69cyt). In cell culture studies we have shown that
expression of this construct in cells inhibited the signaling of
endogenous CD69 (data not shown). An analysis of thymocyte development
in these mice showed that cell-surface expression of CD69 is sufficient
to lead to an increase in SP cells. However, the phenotype in mice
overexpressing functional CD69 (CD69FL) is more dramatic than that in
mice expressing an equivalent level of CD69cyt (compare Figs. 2 and 4). Again, similar to the CD69FL mice, the phenotypes seen in mice
expressing the CD69cyt transgene correlated with levels of transgene
expression. These data, taken together, suggest a model by which a
combination of CD69 interaction with an as yet unknown ligand, coupled
with CD69-mediated signal transduction, contributes to the generation
of SP thymocytes.
There are several possible explanations for the accumulation of thymic
SP cells in the CD69 transgenic mice. One possible explanation is that
there is greater turnover of those SP cells in the thymus. The fact
that overall thymic cellularity is unchanged in these mice argues
against this. Also, we have begun to examine apoptosis in these mice,
using annexin V staining and TUNEL analysis and found no difference
between NLC and CD69 transgenic mice (D. J. Kasprowicz and S.
F. Ziegler, unpublished observations). Another possible explanation is
that CD69 is involved in the trafficking of thymocytes during their
maturation. In this model, unregulated expression of CD69 on thymocytes
increases the movement of thymocytes from the cortex to the medulla
during differentiation. The inability to down-regulate CD69 expression
on these cells causes them to remain in the thymus. Support for this
model comes from indirect immunofluorescence of human thymus using
anti-CD69 mAb. In this study Jung et al. (18) found
that only scattered cells in the cortex expressed CD69, and these cells
tended to cluster in the subcapsular region. These may correspond to
the CD44+CD25- DN
population that we have shown expresses CD69 in mouse thymus (T.
Nakayama and S. F. Ziegler, unpublished observation). In contrast,
most, if not all, medullary thymocytes were
CD69+. These data are consistent with a model in
which CD69 acts as a trafficking molecule, perhaps acting in concert
with CCR4, for cells migrating from the cortex to the medulla during
selection. As these cells complete their developmental program they
down-regulate CD69 and leave the thymus to seed peripheral lymphoid
organs. Overexpression of CD69 on the surface of thymocytes would
therefore be predicted to lead to an increase in mature thymocytes and
a lack of T cells in peripheral organs. This is the phenotype observed
in CD69FL transgenic mice and, to a lesser extent, in CD69cyt mice
(Figs. 2, 4, and 7).
The finding the mice lacking CD69 have normal thymic development (T. Nakayama, D. J. Kasprowicz, M. Yamashita, and S. F. Ziegler, unpublished results) suggests that there are additional molecules expressed on thymocytes capable of interacting with the same ligand and generating related signals. We have searched the public EST databases and have found three novel C-type lectins that are expressed in the thymus (data not shown). We are currently testing the roles of these molecules in thymocyte development. Also, as mentioned above, our interpretation of the data is predicated on the existence of a ligand for CD69 that is expressed in the thymus. Using purified soluble CD69 we have detected specific binding on cell lines that can serve as APC (data not shown). These data are consistent with a CD69 binding partner being expressed in thymus by cells that regulate selection. We are currently examining thymic stromal cell lines for solCD69 binding.
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Acknowledgments |
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Footnotes |
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2 T.N. and D.J.K. contributed equally to this work.
3 Address correspondence and reprint requests to Dr. Steven F. Ziegler, Virginia Mason Research Center, 1201 9th Avenue, Seattle, WA 98101. E-mail address: sziegler{at}vmresearch.org
4 Abbreviations used in this paper: DN, CD8CD4 double negative; DP, CD4CD8 double positive; SP, CD4 or CD8 single positive; HSA, heat-stable Ag.
Received for publication June 6, 2001. Accepted for publication October 22, 2001.
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References |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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T cell receptors. Curr. Opin. Immunol. 9:250.[Medline]
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 C. B.F. Thien, R. M. Scaife, J. M. Papadimitriou, M. A. Murphy, D. D.L. Bowtell, and W. Y. Langdon A Mouse with a Loss-of-function Mutation in the c-Cbl TKB Domain Shows Perturbed Thymocyte Signaling without Enhancing the Activity of the ZAP-70 Tyrosine Kinase J. Exp. Med., February 17, 2003; 197(4): 503 - 513. [Abstract] [Full Text] [PDF]
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T. Berki, L. Palinkas, F. Boldizsar, and P. Nemeth Glucocorticoid (GC) sensitivity and GC receptor expression differ in thymocyte subpopulations Int. Immunol., May 1, 2002; 14(5): 463 - 469. [Abstract] [Full Text] [PDF]
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