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*
Microbiology and Tumor Biology Center, Karolinska Institutet, Stockholm, Sweden;
Department of Microbiology and Immunology, Howard Hughes Medical Institute, Vanderbilt University School of Medicine, Nashville, TN 37232; and
Department of Molecular Immunology, Chiba University Graduate School of Medicine, Chiba, Japan
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Abstract |
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 Top Abstract IntroductionMaterials and MethodsResultsDiscussionReferences |
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ß+ cells have
been described as being either CD4+CD8- or
CD4-CD8-. Most NKT cells interact with the
nonclassical MHC class I molecule CD1 through a largely invariant
V14-J281 TCR chain in conjunction with either a Vß2, -7, or -8
TCR chain. In the present study, we describe the presence of
significant numbers of NK1.1+TCRß+ cells
within lymphokine-activated killer cell cultures from wild-type
C57BL/6, CD1d1-/-, and J281-/- mice that
lack classical NKT cells. Unlike classical NKT cells, 50–60% of these
NK1.1+TCRß+ cells express CD8 and have a
diverse TCR Vß repertoire. Purified
NK1.1-CD8+ T cells from the spleens of B6
mice, upon stimulation with IL-2, IL-4, or IL-15 in vitro, rapidly
acquire surface expression of NK1.1. Many
NK1.1+CD8+ T cells had also acquired expression
of Ly-49 receptors and other NK cell-associated molecules. The
acquisition of NK1.1 expression on CD8+ T cells was a
particular property of the IL-2Rß+ subpopulation of the
CD8+ T cells. Efficient NK1.1 expression on
CD8+ T cells required Lck but not Fyn. The induction of
NK1.1 on CD8+ T cells was not just an in vitro phenomenon
as we observed a 5-fold increase of NK1.1+CD8+
T cells in the lungs of influenza virus-infected mice. These data
suggest that CD8+ T cells can acquire NK1.1 and other NK
cell-associated molecules upon appropriate stimulation in vitro and in
vivo. |
Introduction |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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14-J281 chain associated with either a Vß2, -7 or -8 chain
(4, 5, 6, 7). Mice deficient in CD1 or J281 are largely
devoid of NKT cells, a consequence of their dependence on the CD1
ligand and the invariant V14-J281 chain of the TCR
(6, 7, 8, 9, 10). NKR-P1 (CD161) is a family of disulfide-linked homodimers expressed on the surface of both NK and NKT cells (11). NKR-P1A and C are activating molecules that can trigger both cytokine production and cytolytic activity by NK cells (12, 13), while NKR-P1B appears to have inhibitory activity (14, 15). The Ab PK136 recognizes the NKR-P1C receptor in the mouse, referred to as NK1.1. This Ab depletes NK cells in vivo (16, 17) and induces activation and proliferation of both NK and NKT cells in vitro (18, 19). NK and NKT cells in mice also express C-type lectins of the Ly-49 family (13, 20). Most Ly-49 molecules act as inhibitors for both cytotoxicity and cytokine production (21, 22, 23) but at least two, Ly-49D and Ly-49H, activate NK cells (24, 25). Therefore, the activity of NK and NKT cells appears to be tightly controlled by a series of positive and negative signals.
In the course of the characterization of lymphokine-activated killer
(LAK)3 cell cultures from NKT
cell-deficient J281-/- and
CD1d1-/- mice, we found surprisingly that these
cultures contained a substantial number of cells coexpressing TCRß
and NK1.1. These cells did not resemble "classical" NKT cells,
because a majority of them were CD8+ and
expressed a broad spectrum of TCR Vß-chains. In the present study, we
characterize this subset of "nonclassical" NKT cells from B6,
CD1d1-/-, and
J281-/- mice in vitro and in vivo.
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Materials and Methods |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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B6 mice (6- to 10-wk-old) were obtained from the Microbiology
and Tumor Biology Center of the Karolinska Institutet (Stockholm,
Sweden). CD1d1-/- (9),
J281-/- (6),
Lck-/- (26),
Fyn-/- (27), and
TAP/ß2m-/-
(28) mice have been described previously. Mutant mice were
backcrossed at least three times to a B6 background. All mice were
maintained at the Microbiology and Tumor Biology Center of the
Karolinska Institutet. Animal care was in accordance with national and
institutional guidelines.
Abs and flow cytometry
All Abs were used according to the manufacturers’
recommendations. For flow cytometric analysis, cells were preincubated
with 2.4G2 supernatant for 20 min to block Fc receptors. FITC-, PE- or
biotin-labeled anti-2B4, -CD3
(145-2C11), -CD4 (KH-CD4), -CD8
(53-6.7), -CD8ß.2 (53-5.8), -CD69 (H1.2F3), -CD122
(TM-ß1), -Ly-49A (A1), -Ly-49C/I (5E6), -Ly-49D
(4E5), -NK1.1 (PK136), -ß TCR (H57-597), -Vß2 TCR (B20.6),
-Vß3 TCR (KJ-25), -Vß4 TCR (KT4), -Vß5.1, 5.2 TCR (MR9-4), -Vß6
TCR (RR4-7), -Vß7 TCR (TR310), -Vß8.1, 8.2 TCR (MR5-2), -Vß9 TCR
(MR10-2), -Vß10b (B21.5), -Vß11 (RR3-15),
-Vß12 TCR (MR11-1), -Vß13 TCR (MR12-3), -Vß14 (14-2), and
-Vß17a (KJ-23) were obtained from PharMingen (San Diego, CA).
Anti-Ly-49G2 (4D11) was purified from hybridoma culture supernatants
(hybridoma obtained from American Type Culture Collection, Manassas,
VA) on a protein G column (Amersham Pharmacia Biotech, Uppsala,
Sweden). Streptavidin-FITC was obtained from Dakopatts (Glostrup,
Denmark) and streptavidin-RED670 from Life Technologies (Rockville,
MD). Cells were analyzed on a FACScan or a FACSort cytometer (Becton
Dickinson, Mountain View, CA).
Generation of LAK cells
Single cell suspensions from spleens were depleted from
erythrocytes and resuspended in complete medium (MEM, 10 mM HEPES,
2 x 10-7 M 2-ME, 10% FCS, 100 U/ml
penicillin, and 100 U/ml streptomycin) and human rIL-2 at the
concentrations indicated (PeproTech, Rocky Hill, NJ). Murine rIL-4,
rIL-12, rGM-CSF (PeproTech), human rIL-7, and rIL-15 (R&D, Minneapolis,
MN) were used at 10 ng/ml. Human rIFN- and murine rIFN-
(PeproTech) were used at 1000 U/ml. Anti-CD3 Ab (145-2C11) was used at
10 µg/ml. Cells were cultured at 37°C in a humidified 10%
CO2 atmosphere in air for 5 days when not else
noted. Cell sorting was performed using a FACSvantage cell sorter
(Becton Dickinson).
CD8+ cell purification
Two days before purification, B6 mice were depleted of NK cells
by i.p. injection of 50–100 µg purified anti-NK1.1 Ab/mouse. The
CD8+ cells were isolated using the magnetic
activated cell sorting (MACS) separation system (Miltenyi Biotec,
Bergisch Gladbach, Germany), according to the manufacturer’s
guidelines. Briefly, 1–2 x 108
erythrocyte-depleted B6 splenocytes were washed once with buffer (PBS
without Ca2+ supplemented with 0.5% BSA and 2 mM
EDTA). A total of 10 µl CD8 (Ly-2) microbeads and 90 µl buffer
were added per 106 cells and incubated at 8°C
for 15 min and the cells were washed twice.
CD8+ cells were removed from the cell
suspension using an LS+ column. Purity of cells
was determined by flow cytometry. The MACS-separated cells were washed
once and resuspended in complete medium plus 1000 U/ml IL-2 and plated
in six-well plates (0.5–1 x 106 cells/ml).
Cells were cultured at 37°C in a humidified 10%
CO2 atmosphere in air.
For experiments examining IL-2Rß
(CD122)+CD8+ T cells,
purified CD8+ cells were washed once and
stained with IL-2Rß-FITC or -PE and sorted by flow cytometry into
IL-2Rß+CD8+ and
IL-2Rß-CD8+ T cell
populations. The cells were cultured in complete medium with 1000 U/ml
IL-2 and analyzed for NK1.1 expression on day 3 and day 6.
RNA isolation, reverse transcription, and RT-PCR
Total cellular RNA was extracted using the Trizol RNA isolation method (Life Technologies), followed by cDNA synthesis of 2 µg RNA using the First-strand cDNA synthesis kit (Amersham Pharmacia Biotech) according to the protocol using pd(N)6 primers. PCR analysis was performed using DNA polymerase (Boehringer Mannheim, Mannheim, Germany) on a Programmable Thermal Controller PTC-100 (MJ Research, Watertown, MA) with 10-s denaturation at 94°C, 30-s annealing at 55°C, and 2-min extension at 72°C for 34 cycles, with a start at 94°C for 2 min and a final extension at 72°C for 6 min. NKR-P1C transcripts were detected using two murine NKR-P1C-specific primers (14); 5'-TGAAATGGACACAGCAAGTATC and 3'-GAGTCAACGAATGGAAAGGAA (Cybergene, Stockholm, Sweden) encompassing nucleotides 96–1170 of murine NKR-P1C. cDNA quality was confirmed by amplification of a ß-actin gene fragment using specific primers. Photo image of an ethidium bromide-stained gel is shown.
Preparation of cells from lungs of influenza-infected mice
Mice were infected intranasally with influenza A virus Japan/305/57 (a kind gift from Dr. A. D. Diehl (Microbiology and Tumor Biology Center, Karolinska Institute, Stockholm, Sweden). Ten days postinfection, both infected and control mice were sacrificed by cervical dislocation and their lungs were removed. The lungs were minced into small pieces and incubated in RPMI 1640 (10% FCS) with 250 U/ml collagenase type IV (Sigma, St. Louis, MO) at 37°C for 90 min. The erythrocytes were lysed and single cell suspensions were prepared from the digested lungs. Adherent cells were removed from the lung cellular suspension by incubating the cells on tissue culture petri dishes at 37°C for 90 min.
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Results |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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In LAK cell cultures from B6, NKT cell-deficient
CD1d1-/-, and
J281-/- mice, NK1.1+ T
cells were detected after 5 days of culture in IL-2. The numbers of
NK1.1+ T cells in LAK cell cultures from the NKT
cell-deficient mice were similar to those from B6 mice (Fig. 1
). Using three-color flow cytometry,
50–60% of these
NK1.1+TCRß+ cells were
found to express CD8 which is normally not expressed on classical
NKT cells (Fig. 2). A majority of the
CD8+ T cells (70–90%) also coexpressed
CD8ß. The remaining
CD8-NK1.1+TCRß+
population was predominantly CD4- but a minor
population expressed CD4 (Fig. 2; data not shown). Further analysis of
the
NK1.1+TCR+CD8+
T cells (referred to as
NK1.1+CD8+ T cells)
revealed that they expressed a diverse TCR repertoire because they were
polyclonal with regards to Vß expression (Fig. 3). This observation, and the fact that
these cells could be generated from NKT cell-deficient mice, suggested
that these cells were not classical NKT cells.
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Among fresh splenocytes, 1–3% of all CD8+
cells expressed NK1.1. After treating mice with anti-NK1.1 Ab to
remove NK1.1+ cells in vivo,
NK1.1+CD8+ T cells were no
longer detected in the spleen cell population. To demonstrate that
CD8+ T cells could acquire expression of NK1.1,
CD8+ cells were purified from mice pretreated
with anti-NK1.1 Ab (Fig. 4A, left panel).
When these CD8+ cells were examined after 5
days of culture with IL-2, up to 40% of the cells had acquired
expression of NK1.1 (Fig. 4A, right panel). To
extend these observations, RT-PCR analyses for NKR-P1C (NK1.1) was
performed using RNA from freshly isolated CD8+
cells from B6 mice pretreated with anti-NK1.1 Ab and similar
CD8+ cells after culture with IL-2 for 5 days.
Before culture, no NKR-P1C transcripts were detected in the
CD8+ cells, whereas a marked increase in NKR-P1C
transcripts was detected after 5 days of culture in IL-2 (Fig. 4B). To further support the notion that
CD8+ T cells could acquire expression of NK1.1,
B6-derived LAK cells were sorted by flow cytometry on day 4 into
subpopulations of
NK1.1+CD8-,
NK1.1-CD8+, and
NK1.1+CD8+ T cells and
further cultured in the presence of IL-2. After 48 h, 
25% of
the cells from the
NK1.1-CD8+ population had
acquired NK1.1 expression (data not shown).
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D). This observation argues against the possibility that NK
cells would have been the predominant source of the
NK1.1+CD8+ T cell
population. Taken together, all these observations are consistent with
the notion that CD8+ T cells in the LAK cell
cultures can acquire expression of NK1.1, and that the transcriptional
activation precedes expression at the cell surface. Dose responses and kinetics of NK1.1 expression
Because the initial experiments were performed with relatively
high concentrations of IL-2, we determined the minimal IL-2
concentration required for NK1.1 induction on
CD8+ T cells. NK1.1 expression was readily
observed on CD8+ T cells at a concentration of 10
U/ml (Fig. 5), and occasionally at levels
as low as 1 U/ml (data not shown). There was also a gradual increase in
the number of NK1.1+CD8+ T
cells when CD8+ cells were exposed to
increasing concentrations of IL-2. At IL-2 concentrations between 10
and 1000 U/ml, we observed induction of NK1.1 expression already within
the first 48 h of culture (Fig. 5).
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Because up-regulation of NK1.1 was observed on IL-2-stimulated
B6-derived CD8+ T cells, we examined whether
other NK cell-associated molecules such as the Ly-49 molecules were
induced on these cells. Only a small percentage of purified
CD8+ T cells from young B6 mice (<3 mo old)
expressed Ly-49 molecules and the expression levels were generally low
(Table I; data not shown). However, a
significant proportion of
NK1.1+CD8+ T cells acquired
expression of Ly-49A, Ly-49C/I, and Ly-49G2 after 72 h of culture
with IL-2, whereas most
NK1.1-CD8+ T cells
remained Ly-49 negative (Table I). This suggests that the acquisition
of NK1.1 occurs in parallel with the acquisition of the inhibitory
Ly-49 molecules, Ly-49 A, C/I, and G2. We also examined expression of
the activating molecule Ly-49D on
NK1.1+CD8+ T cells.
However, no significant Ly-49D expression was detected on these cells
whereas 20% of normal LAK cell-derived NK cells expressed Ly-49D (data
not shown).
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The ability of IL-2 to induce expression of NK1.1 on
CD8+ T cells may not be a unique property of this
cytokine. Thus, other cytokines were examined with respect to their
ability to induce NK1.1 expression on CD8+ T
cells. CD8+ T cells cultured with IL-4 or
IL-15 for 5 days were found to express NK1.1 at levels similar to cells
cultured with IL-2 (Fig. 6, A–C), whereas cells cultured with IL-7, IL-12, GM-CSF,
IFN-, IFN-, or anti-CD3 Ab did not (Fig. 6D; data
not shown). This observation suggests that IL-2, IL-4, and IL-15
up-regulated NK1.1 on the cell surface of CD8+ T
cells through a common pathway such as via the common -chain.
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Although sorted CD8+ T cells were found to
acquire expression of NK1.1 after cytokine stimulation, it was not
entirely clear whether this was a function of all
CD8+ T cells or a subpopulation of
CD8+ T cells. Because 10–20% of freshly
derived CD8+ T cells expressed IL-2Rß (data not
shown), CD8+ T cells were sorted by flow
cytometry into IL-2Rß+ and
IL-2Rß- populations and cultured in IL-2. When
the cells were examined after 6 days of culture, 35% of the
IL-2Rß+CD8+ T cells
coexpressed NK1.1 (Fig. 7A),
whereas only 3% of the
IL-2Rß-CD8+ T cells were
NK1.1+ (Fig. 7B). These observations
suggest that expression of IL-2Rß was a prerequisite for IL-2-induced
NK1.1 expression on CD8+ T cells.
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IL-2Rß can transduce signals through soluble protein kinases
such as Lck and Fyn. When LAK cells were prepared from
Fyn-/- mice, the number of
NK1.1+CD8+ T cells
generated was equivalent to the number found in B6 LAK cell cultures.
However, LAK cell cultures from Lck-/- mice had
a significantly reduced number of
NK1.1+CD8+ T cells compared
with B6 LAK cell cultures (data not shown). Because this reduced number
of NK1.1+CD8+ T cells could
be related to the low frequency of T cells in
Lck-/- mice, purified
CD8+ T cells from
Lck-/- mice were cultured with IL-2 to
determine whether Lck signaling was required for up-regulation of NK1.1
expression. Even though 50% of the purified
CD8+ T cells from the
Lck-/- mice expressed IL-2Rß (data not
shown), only 8% of the purified CD8+ T cells
from Lck-deficient mice expressed NK1.1 whereas 25% of the
corresponding cells from B6 mice expressed NK1.1 after 5 days of
culture with IL-2 (Fig. 8). Therefore,
these observations suggest that Lck is required for efficient
IL-2-mediated expression of NK1.1 on CD8+ T
cells.
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The fact that CD8+ T cells could up-regulate
NK1.1 and other NK cell-associated molecules upon cytokine stimulation
in vitro led us to address whether this could occur in vivo. NK1.1 was
expressed on 1–3% of CD8+ T cells in the
lungs of untreated B6 mice (Fig. 9A). However, 10 days after
infection with influenza A virus Japan/305/57, elevated numbers of
NK1.1+CD8+ T cells were
detected in the lungs. At this time point, 10% of the
CD8ß+ T cells coexpressed NK1.1 (Fig. 9B). This observation suggests that NK1.1 can be
up-regulated on CD8+ T cells in vivo.
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Discussion |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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NK1.1+CD8+ T cells were
also found in the lungs of mice infected with influenza A virus. These
cells were detected at low numbers in the lungs of uninfected mice, but
represented up to 10% of the total CD8+
population at day 10 postinfection. The majority of the
NK1.1+CD8+ T cells were
CD8ß+TCRß+ and
also expressed the NK cell-associated Ly-49 receptors. Interestingly,
up to 20% of these
NK1.1+CD8+ T cells were
virus-specific as determined by MHC class I tetramer staining. As
observed with the IL-2 derived
NK1.1+CD8+ T cells, these
cells were distinct from conventional NKT cells because they were also
found in influenza-infected CD1-/- mice
(29).
The definition of NKT cells is not entirely clear. Classically, NKT
cells are defined by their coexpression of TCR and NK cell-associated
markers such as NK1.1. Most NKT cells have been characterized as being
either CD4-CD8- or
CD4+CD8-, and express a
restricted TCR repertoire that is specific for CD1 (1, 2, 3, 4).
However, not all NKT cells are CD1-restricted and some have been
reported to express CD8.
NK1.1+CD8+ T cells have
been found previously in LAK cell cultures from B6 mice (30, 31). In one of these studies,
NK1.1+CD8+ T cells
were described to exist in IL-4-derived LAK cell cultures but not in
IL-2-derived LAK cell cultures (30).
NK1.1+CD8+ T cells have
also been reported to be a minor subset of NKT cells in vivo
(32, 33, 34, 35, 36, 37). In a study on lymphocytic choriomeningitis
virus-infected mice, a marked increase in the numbers of
NK1.1+CD8+ T cells was
observed (37). Whether these
NK1.1+CD8+ T cells are
CD1-restricted is not clear as one study found that their numbers did
not change in CD1-/- mice (32),
while another found a dramatic reduction in the number of
NK1.1+CD8+ T cells from the
bone marrow of CD1-/- mice (35).
It has also been suggested that
NK1.1+CD8+ T cells are a
distinct lineage as they can arise even in thymectomized mice
(33). Coles et al. (36) have demonstrated
that
Ly-49+NK1.1+CD8+
T cells are prevalent in older mice. In their study, it was made clear
that these cells represented a subpopulation of normal T cells rather
than "aberrant" NKT cells. Our results differ from some of the
observations described above but are in line with others. In contrast
to the study of Ballas and Rasmussen (30), we found that
both CD8+ and
CD8ß+ T cells could acquire NK1.1.
Furthermore, NK1.1+CD8+ T
cells were detected in LAK cell cultures from both
CD1d1-/- and J281-/-
mice in equal number to those observed in B6 mice. Our data suggest
that a significant proportion of the
NK1.1+CD8+ T cells observed
in the present study arise from the
IL-2Rß+CD8+ T cells
following activation with IL-2 and during influenza infection
(29). Thus,
NK1.1+CD8+ T cells most
likely represent a state of activation or differentiation of
CD8+ T cells rather than the selective expansion
of a distinct cell lineage. The expression of 2B4 and CD69 on a
majority of the
NK1.1+CD8+ T cells is in
line with this notion. These cells differ from classical NKT cells in a
number of aspects. First, they could be detected even in
CD1d1-/- and J281-/-
mice that are deficient in classical NKT cells. Second, the
NK1.1+CD8+ T cells are
polyclonal with regards to TCR Vß expression and finally, they do not
appear to recognize CD1 (our unpublished observations).
Expression of NK cell-associated molecules on T cells may be induced by
cytokines triggering the expression of proteins encoded within the NK
gene complex. The NK gene complex has been described in both rodents
and humans. In mice, the complex is found on chromosome 6 whereas in
humans it is located on chromosome 12 (38). The NK gene
complex contains the genes for NKR-P1, CD69, and the killer cell
inhibitory receptors (KIRs), Ly-49 (in mouse) and CD94 (in mouse and
human). Because only cytokines that use the common -chain appeared
to up-regulate NK1.1 on the T cells, one may speculate that signaling
through this chain could activate specific genes within the NK gene
complex. Support for this hypothesis comes from the observation that
CD94/NKG2A could be up-regulated on human CD8+ T
cells treated with IL-15 (39). Interestingly, the
up-regulation of the CD94/NKG2A complex was associated with inhibition
of the cytotoxic capability of these T cells (39, 40).
Other studies with human CD8+ T cells have
demonstrated that stimulation with IL-12 up-regulated expression of
NKR-P1A, whereas IL-2 and IL-15 down-regulated NKR-P1A
(41). It should be noted that NKR-P1A may have different
functions than NKR-P1C (NK1.1) and so could be regulated in a different
manner to accommodate its functions. Similarly, the lack of Ly-49D on
the NK1.1+CD8+ T cells may
indicate that the expression of this molecule is regulated differently
than that of the other Ly-49 molecules examined.
The protein kinases Lck and Fyn are critical in the development of T
cells and NKT cells in vivo (26, 27).
Lck-/- mice have reduced numbers of
CD4+CD8+ T cells and
single-positive cells in the thymus and consequently reduced numbers of
peripheral T cells. Fyn-/- mice have normal T
cell development but two recent studies have found that the number of
NKT cells is reduced in these mice (42, 43). It has been
demonstrated that Fyn binds to CD3
, and Lck to CD4 and CD8;
however, both Lck and Fyn can bind the IL-2Rß (44, 45).
LAK cells generated from Fyn-/- mice had
equivalent numbers of
NK1.1+CD8+ T cells to those
derived from B6 mice. In contrast, LAK cells generated from
Lck-/- mice had significantly lower numbers of
NK1.1+CD8+ T cells.
Furthermore, purified CD8+ T cells from
Lck-/- mice could not up-regulate NK1.1 in the
presence of IL-2 despite the fact that 50% of these cells expressed
IL-2Rß compared with 10–20% in B6 mice (our unpublished
observations). Therefore, lack of Lck signaling could be related to the
diminished ability of the
IL-2Rß+CD8+ T cells in
the Lck-/- mice to express NK1.1.
Non-MHC-restricted killing is an in vitro phenomenon that is observed in long-term cultured T cells and in T cell cultures stimulated with high concentrations of IL-2 (46). To date, a molecular explanation for this phenomenon has yet to be elucidated. One may speculate that the up-regulation of NK cell-associated molecules allows T cells to expand their killing capabilities to include innate non-MHC-restricted killing. Indeed, we have observed that some CD8+ T cell lines expressing NK1.1 targeted not only specific peptide-loaded cells but also the prototypic NK cell target, YAC-1, whereas CD8+NK1.1- T cell lines did not kill YAC-1 cells (our unpublished observations). However, because there is no known ligand for NK1.1 on YAC-1 cells, YAC-1 killing might also relate to the expression of other unknown activating molecules on these T cells. Although non-MHC-restricted killing may be an in vitro phenomenon, such killing could possibly also play a critical role in controlling the spread of infections in vivo.
The induction of Ly-49 molecules on CD8+ T cells may be of importance in regulating the fate of activated CD8+ T cells. With respect to human CD8+ T cells, it has been speculated that the induction of KIR expression of human CD8+ T cells may by their inhibitory functions dampen T cell activity and so prevent the CD8+ T cells from becoming "exhausted" by overstimulation. The latter could render them a selective advantage during memory cell development (36, 47). Several other studies with human CD8+ T cells also support a notion where expression of KIR or CD94/NKG2 receptors may regulate T cell function (39, 40, 48). The latter may also be the case for inhibitory Ly-49 molecules. Indeed, studies using mice transgenic for Ly-49 molecules have found that CD8+ T cell function is affected in these mice (49, 52). In relation to Ly-49 molecules, it is of interest to note that the significant population of memory CD8+ T cells in the mouse express Ly-49 receptors (36).
NK cells have become appreciated for their potential roles in a variety of infectious diseases (53, 54) as well as in host responses against some tumors (55). Much of this knowledge has been derived in mice depleted of NK cells using anti-NK1.1 Abs. With the present observation that some CD8+ T cells can acquire expression of NK1.1, there may now be a requirement for the reanalysis of some of the data that have implicated a role for NK cells in host responses to infections and tumors. Indeed, the appearance of NK1.1+CD8+ T cells in the lungs of influenza-infected mice and lymphocytic choriomeningitis virus-infected mice (37) suggests that these cells do appear in vivo upon infection and do not merely represent an in vitro artifact. In particular, T cell responses to influenza were reported to be low in mice treated with anti-NK1.1 Ab (56). One may speculate that in this case, viral-specific T cells had been removed by anti-NK1.1 Ab administration that could explain the unresponsiveness.
In conclusion, the present data describe the induction of expression of NK1.1 and other NK cell-associated molecules on a subpopulation of CD8+ T cells. These data suggest that CD8+ T cells can acquire NK cell-associated receptors upon appropriate stimulation in vitro and in vivo.
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Acknowledgments |
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Footnotes |
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2 Address correspondence to Dr. Benedict J. Chambers, Microbiology and Tumor Biology Center, Karolinska Institutet, Stockholm, S-171 77, Sweden.
3 Abbreviations used in this paper: LAK, lymphokine-activated killer; ß2m, ß2-microglobulin; B6, C57BL/6; KIR, killer cell inhibitory receptor.
Received for publication January 28, 2000. Accepted for publication July 17, 2000.
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References |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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  A. Rouhi, C. B. Lai, T. P. Cheng, F. Takei, W. M. Yokoyama, and D. L. Mager Evidence for high bi-allelic expression of activating Ly49 receptors Nucleic Acids Res., September 1, 2009; 37(16): 5331 - 5342. [Abstract] [Full Text] [PDF]
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 J. G. Aust, F. Gays, K. M. Mickiewicz, E. Buchanan, and C. G. Brooks The Expression and Function of the NKRP1 Receptor Family in C57BL/6 Mice J. Immunol., July 1, 2009; 183(1): 106 - 116. [Abstract] [Full Text] [PDF]
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S. M. Ngoi, M. G. Tovey, and A. T. Vella Targeting Poly(I:C) to the TLR3-Independent Pathway Boosts Effector CD8 T Cell Differentiation through IFN-{alpha}/{beta} J. Immunol., December 1, 2008; 181(11): 7670 - 7680. [Abstract] [Full Text] [PDF]
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N. K. Bjorkstrom, V. D. Gonzalez, K.-J. Malmberg, K. Falconer, A. Alaeus, G. Nowak, C. Jorns, B.-G. Ericzon, O. Weiland, J. K. Sandberg, et al. Elevated Numbers of Fc{gamma}RIIIA+ (CD16+) Effector CD8 T Cells with NK Cell-Like Function in Chronic Hepatitis C Virus Infection J. Immunol., September 15, 2008; 181(6): 4219 - 4228. [Abstract] [Full Text] [PDF]
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M. Terabe, Y. Tagaya, Q. Zhu, L. Granger, M. Roederer, T. A. Waldmann, and J. A. Berzofsky IL-15 Expands Unconventional CD8{alpha}{alpha}NK1.1+ T Cells but Not V{alpha}14J{alpha}18+ NKT Cells J. Immunol., June 1, 2008; 180(11): 7276 - 7286. [Abstract] [Full Text] [PDF]
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 K.-S. Choi, T. Webb, M. Oelke, D. G. Scorpio, and J. S. Dumler Differential Innate Immune Cell Activation and Proinflammatory Response in Anaplasma phagocytophilum Infection Infect. Immun., June 1, 2007; 75(6): 3124 - 3130. [Abstract] [Full Text] [PDF]
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D. Kamimura, Y. Sawa, M. Sato, E. Agung, T. Hirano, and M. Murakami IL-2 In Vivo Activities and Antitumor Efficacy Enhanced by an Anti-IL-2 mAb J. Immunol., July 1, 2006; 177(1): 306 - 314. [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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S. T. Smiley, P. A. Lanthier, K. N. Couper, F. M. Szaba, J. E. Boyson, W. Chen, and L. L. Johnson Exacerbated Susceptibility to Infection-Stimulated Immunopathology in CD1d-Deficient Mice J. Immunol., June 15, 2005; 174(12): 7904 - 7911. [Abstract] [Full Text] [PDF]
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 M. Giroux and F. Denis CD1d-unrestricted human NKT cells release chemokines upon Fas engagement Blood, January 15, 2005; 105(2): 703 - 710. [Abstract] [Full Text] [PDF]
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Y. Han, M. Zhang, N. Li, T. Chen, Y. Zhang, T. Wan, and X. Cao KLRL1, a novel killer cell lectinlike receptor, inhibits natural killer cell cytotoxicity Blood, November 1, 2004; 104(9): 2858 - 2866. [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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 J. E. Gumperz CD1d-restricted "NKT" cells and myeloid IL-12 production: an immunological crossroads leading to promotion or suppression of effective anti-tumor immune responses? J. Leukoc. Biol., August 1, 2004; 76(2): 307 - 313. [Abstract] [Full Text] [PDF]
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C. D. Peacock and R. M. Welsh Origin and Fate of Lymphocytic Choriomeningitis Virus-Specific CD8+ T Cells Coexpressing the Inhibitory NK Cell Receptor Ly49G2 J. Immunol., July 1, 2004; 173(1): 478 - 484. [Abstract] [Full Text] [PDF]
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 K. U. Saikh, B. Dyas, T. Kissner, and R. G. Ulrich CD56+-T-Cell Responses to Bacterial Superantigens and Immune Recognition of Attenuated Vaccines Clin. Vaccine Immunol., November 1, 2003; 10(6): 1065 - 1073. [Abstract] [Full Text] [PDF]
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 P. Lundberg, P. Welander, H. Openshaw, C. Nalbandian, C. Edwards, L. Moldawer, and E. Cantin A Locus on Mouse Chromosome 6 That Determines Resistance to Herpes Simplex Virus Also Influences Reactivation, While an Unlinked Locus Augments Resistance of Female Mice J. Virol., November 1, 2003; 77(21): 11661 - 11673. [Abstract] [Full Text] [PDF]
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S. Dhanji and H.-S. Teh IL-2-Activated CD8+CD44high Cells Express Both Adaptive and Innate Immune System Receptors and Demonstrate Specificity for Syngeneic Tumor Cells J. Immunol., October 1, 2003; 171(7): 3442 - 3450. [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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T. Kambayashi, E. Assarsson, A. E. Lukacher, H.-G. Ljunggren, and P. E. Jensen Memory CD8+ T Cells Provide an Early Source of IFN-{gamma} J. Immunol., March 1, 2003; 170(5): 2399 - 2408. [Abstract] [Full Text] [PDF]
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A. B. Geldhof, J. A. Van Ginderachter, Y. Liu, W. Noel, G. Raes, and P. De Baetselier Antagonistic effect of NK cells on alternatively activated monocytes: a contribution of NK cells to CTL generation Blood, December 1, 2002; 100(12): 4049 - 4058. [Abstract] [Full Text] [PDF]
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 Y. Mizukawa, Y. Yamazaki, Y. Teraki, J. Hayakawa, K. Hayakawa, H. Nuriya, M. Kohara, and T. Shiohara Direct Evidence for Interferon-{gamma} Production by Effector-Memory-Type Intraepidermal T Cells Residing at an Effector Site of Immunopathology in Fixed Drug Eruption Am. J. Pathol., October 1, 2002; 161(4): 1337 - 1347. [Abstract] [Full Text] [PDF]
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J. D. Miller, M. Peters, A. E. Oran, G. W. Beresford, L. Harrington, J. M. Boss, and J. D. Altman CD94/NKG2 Expression Does Not Inhibit Cytotoxic Function of Lymphocytic Choriomeningitis Virus-Specific CD8+ T Cells J. Immunol., July 15, 2002; 169(2): 693 - 701. [Abstract] [Full Text] [PDF]
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N. Ohta, T. Hiroi, M.-N. Kweon, N. Kinoshita, M. H. Jang, T. Mashimo, J.-I. Miyazaki, and H. Kiyono IL-15-Dependent Activation-Induced Cell Death-Resistant Th1 Type CD8{alpha}{beta}+NK1.1+ T Cells for the Development of Small Intestinal Inflammation J. Immunol., July 1, 2002; 169(1): 460 - 468. [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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 J. E. Gumperz, S. Miyake, T. Yamamura, and M. B. Brenner Functionally Distinct Subsets of CD1d-restricted Natural Killer T Cells Revealed by CD1d Tetramer Staining J. Exp. Med., March 4, 2002; 195(5): 625 - 636. [Abstract] [Full Text] [PDF]
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E. Y. Chiang, M. Henson, and I. Stroynowski The Nonclassical Major Histocompatibility Complex Molecule Qa-2 Protects Tumor Cells from NK Cell- and Lymphokine-Activated Killer Cell-Mediated Cytolysis J. Immunol., March 1, 2002; 168(5): 2200 - 2211. [Abstract] [Full Text] [PDF]
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M. S. Duthie, M. Wleklinski-Lee, S. Smith, T. Nakayama, M. Taniguchi, and S. J. Kahn During Trypanosoma cruzi Infection CD1d-Restricted NK T Cells Limit Parasitemia and Augment the Antibody Response to a Glycophosphoinositol-Modified Surface Protein Infect. Immun., January 1, 2002; 70(1): 36 - 48. [Abstract] [Full Text] [PDF]
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T. Kambayashi, E. Assarsson, B. J. Chambers, and H.-G. Ljunggren Cutting Edge: Regulation of CD8+ T Cell Proliferation by 2B4/CD48 Interactions J. Immunol., December 15, 2001; 167(12): 6706 - 6710. [Abstract] [Full Text] [PDF]
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J. A. Hobbs, S. Cho, T. J. Roberts, V. Sriram, J. Zhang, M. Xu, and R. R. Brutkiewicz Selective Loss of Natural Killer T Cells by Apoptosis following Infection with Lymphocytic Choriomeningitis Virus J. Virol., November 15, 2001; 75(22): 10746 - 10754. [Abstract] [Full Text] [PDF]
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L. M. Alonso-C., J. J. Munoz, and A. G. Zapata Delineation of Intrathymic T, NK, and Dendritic Cell (DC) Progenitors in Fetal and Adult Rats: Demonstration of a Bipotent T/DC Intermediate Precursor J. Immunol., October 1, 2001; 167(7): 3635 - 3641. [Abstract] [Full Text] [PDF]
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J. Dunne, S. Lynch, C. O'Farrelly, S. Todryk, J. E. Hegarty, C. Feighery, and D. G. Doherty Selective Expansion and Partial Activation of Human NK Cells and NK Receptor-Positive T Cells by IL-2 and IL-15 J. Immunol., September 15, 2001; 167(6): 3129 - 3138. [Abstract] [Full Text] [PDF]
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C. Karnbach, M. R. Daws, E. C. Niemi, and M. C. Nakamura Immune Rejection of a Large Sarcoma Following Cyclophosphamide and IL-12 Treatment Requires Both NK and NK T Cells and Is Associated with the Induction of a Novel NK T Cell Population J. Immunol., September 1, 2001; 167(5): 2569 - 2576. [Abstract] [Full Text] [PDF]
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Z. Trobonjaca, F. Leithauser, P. Moller, H. Bluethmann, Y. Koezuka, H. R. MacDonald, and J. Reimann MHC-II-Independent CD4+ T Cells Induce Colitis in Immunodeficient RAG-/- Hosts J. Immunol., March 15, 2001; 166(6): 3804 - 3812. [Abstract] [Full Text] [PDF]
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M. Uhrberg, N. M. Valiante, N. T. Young, L. L. Lanier, J. H. Phillips, and P. Parham The Repertoire of Killer Cell Ig-Like Receptor and CD94:NKG2A Receptors in T Cells: Clones Sharing Identical {{alpha}}{{beta}} TCR Rearrangement Express Highly Diverse Killer Cell Ig-Like Receptor Patterns J. Immunol., March 15, 2001; 166(6): 3923 - 3932. [Abstract] [Full Text] [PDF]
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D. G. Pellicci, K. J.L. Hammond, A. P. Uldrich, A. G. Baxter, M. J. Smyth, and D. I. Godfrey A Natural Killer T (NKT) Cell Developmental Pathway Involving a Thymus-dependent NK1.1-CD4+ CD1d-dependent Precursor Stage J. Exp. Med., March 25, 2002; 195(7): 835 - 844. [Abstract] [Full Text] [PDF]
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| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
, Freshly
isolated CD8+ spleen cells from B6 mice; 
,
NK1.1-CD8+ cells after 5 days of culture in
1000 U/ml IL-2; 
, NK1.1+CD8+ cells after 5
days of culture in 1000 U/ml IL-2.
