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14 NKT Cells in Lungs and Their Roles in Th1 Response and Host Defense in Cryptococcal Infection1
*
First Department of Internal Medicine, Faculty of Medicine, University of the Ryukyus, Okinawa, Japan; and
Core Research for Evolutional Science and Technology Project, Department of Molecular Immunology, Graduate School of Medicine, Chiba University, Chiba, Japan
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Abstract |
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 Top Abstract IntroductionMaterials and MethodsResultsDiscussionReferences |
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14 NKT
cell-deficient mice, such increase was significantly attenuated. The
proportion of V14 NKT cells, detected by binding to
-galactosylceramide-loaded CD1d tetramer, and the expression of
V14 mRNA increased after infection with a similar kinetics. The
delayed-type hypersensitivity response and differentiation of the
fungus-specific Th1 cells was reduced in V14 NKT cell-deficient
mice, compared with control mice. Additionally, elimination of this
fungal pathogen from lungs was significantly delayed in V14 NKT
cell-deficient mice. Production of monocyte chemoattractant protein
(MCP)-1 in lungs, detected at both mRNA and protein levels,
increased on day 1, reached a peak level on day 3, and decreased
thereafter, which preceded the increase in NKT cells. Finally, the
increase of total and V14+ subset of NKT cells after
infection was significantly reduced in MCP-1-deficient mice. Our
results demonstrated that NKT cells, especially V14+
subset, accumulated in a MCP-1-dependent manner in the lungs after
infection with C. neoformans and played an important
role in the development of Th1 response and host resistance to this
fungal pathogen. |
Introduction |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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chain consisting of
V14-J281 gene segment and highly skewed V
chains, V8.2,
V7, and V2 in mice (1, 2). These cells are
found in large numbers in the liver, thymus, and bone marrow and in
small numbers in the spleen and lung (2). Although the
natural ligand for NKT cells remains to be defined, GPIs and a
synthetic glycolipid, -galactosylceramide
(-GalCer),3 have
been demonstrated to be presented in the context of CD1d
(1, 2, 3, 4). NKT cells secrete large amounts of IFN-
and
IL-4 in a prompt manner after engagement of the Ag receptor (1, 2, 4, 5) and contribute to the differentiation of both Th1 and
Th2 cells (1, 2, 6, 7, 8, 9, 10). It has been reported that NKT
cells play an important role in various aspects of the regulation and
effector arms of the immune response, including the regulation of
allergic and autoimmune diseases (11, 12, 13, 14) and
prevention of tumor metastasis (15, 16, 17, 18).
The role of NKT cells in host defense to infectious pathogens has
been elucidated by several investigators, and three different results
were reported. First, the clinical course of Mycobacterium
tuberculosis infection is not much different in CD1d-deficient and
control mice, the former of which lack NKT cells (19), and
is minimally affected by treatment with anti-CD1d mAb
(20). Similarly, genetic depletion of V14 NKT cells did
not result in worsening of Salmonella choleraesuis infection
(21). Second, infection with Listeria
monocytogenes or Toxoplasma gondii improved by
manipulations designed to suppress the activity of NKT cells (22, 23). Finally, mice lacking V14 NKT cells were more
susceptible to Leishmania major infection than were control
mice (24). A ligand-specific activation of V14 NKT
cells by -GalCer has been shown to protect mice against protozoan
and fungal infections (25, 26). Thus, the role of NKT
cells seems different among infectious pathogens.
Cryptococcus neoformans causes granulomatous lesions in the
lungs, a primary infected organ, and hematogenously disseminates to the
central nervous system, frequently leading to lethal
meningoencephalitis, particularly in AIDS patients. The host defense
against this pathogen is critically regulated by cell-mediated immunity
(27), and CD4+ T cells play a
central role in limiting infection (28, 29). The balance
between Th1 and Th2 cytokines markedly influences the outcome of
infection; the predominant synthesis of Th1 cytokines over Th2 protects
mice against infection (30, 31), whereas infection is
exacerbated under Th2-dominant conditions (32, 33). In
recent studies (31), targeted disruption of the gene for
IL-12 or IL-18, both of which play important roles in the
differentiation of Th1 cells and IFN- synthesis by T and NK cells,
resulted in reduced host resistance and Th1 response to C.
neoformans. However, the role of NKT cells in the development of
fungus-specific Th1 cells and host defense remains to be elucidated,
although -GalCer-induced activation of NKT cells resulted in the
protection of mice against this infection (26).
In the present study, the role of V14 NKT cells in Th1 response and
host resistance to C. neoformans infection was elucidated.
For this purpose, we examined the kinetics of accumulation of this
subset in the lungs after intratracheal inoculation and the effects of
V14 NKT cell deficiency on the development of Th1 cells and
delayed-type hypersensitivity (DTH) response and elimination of
microorganisms from infected organ. We further addressed the mechanism
of NKT cell increase in lungs after intratracheal infection with
C. neoformans by testing the role of monocyte
chemoattractant protein (MCP)-1, a -chemokine.
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Materials and Methods |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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V14 NKT cell-deficient mice were established by targeted
deletion of the J281 gene segment (34) and back-crossed
eight times with C57BL/6 mice. MCP-1-deficient mice with a genetic
background of C57BL/6 mice (35) were kindly provided by
B. J. Rollins (Harvard Medical School, Boston, MA). These mice
were bred in a pathogen-free environment in the Laboratory Animal
Center for Biomedical Science, University of the Ryukyus (Okinawa,
Japan). C57BL/6 mice were purchased from Charles River Breeding
Laboratories (Osaka, Japan) and used as a control wild-type (WT)
animal. All mice were used at 8–15 wk of age. All experimental
protocols described in the present study were approved by the Ethics
Review Committee for Animal Experimentation of University of the
Ryukyus.
Microorganisms
A serotype A-encapsulated strain of C. neoformans, designated as YC-13, was established from a patient with pulmonary cryptococcosis (36). Infection with this pathogen was self-limited in the lungs of WT mice and did not disseminate to the brain. The yeast cells were cultured on potato dextrose agar plates for 2–3 days before use. To induce pulmonary infection, mice were anesthetized by i.p. injection of 70 mg/kg pentobarbital (Abbott Laboratories, North Chicago, IL) and restrained on a small board. Live C. neoformans (1 x 106 cells) were inoculated at 50 µl per mouse by insertion of a 25-gauge blunt needle into and parallel to the trachea.
Preparation of pulmonary intraparenchymal leukocytes
Pulmonary intraparenchymal leukocytes were prepared as described previously (37). Briefly, the chest of the mouse was opened and the lung vascular bed was flushed by injecting 3 ml of chilled physiological saline into the right ventricle. The lungs were then excised and washed in physiological saline. The lungs, teased with the stainless mesh, were incubated in RPMI 1640 (Life Technologies, Grand Island, NY) with 5% FCS (Cansera, Rexdale, Ontario, Canada), 100 U/ml penicillin G, 100 µg/ml streptomycin, 10 mM HEPES, 50 µM 2-ME, and 2 mM L-glutamine, containing 20 U/ml collagenase (Sigma-Aldrich, St. Louis, MO) and 1 µg/ml DNase I (Sigma-Aldrich). After incubation for 60 min at 37°C with vigorous shaking, the tissue fragments and the majority of dead cells were removed by passing through the 50-µm nylon mesh. After centrifugation, the cell pellet was resuspended in 4 ml of 40% (v/v) Percoll (Pharmacia Biotech, Uppsala, Sweden) and layered onto 4 ml of 80% (v/v) Percoll. After centrifugation at 600 x g for 20 min at 15°C, the cells at the interface were collected, washed three times, and counted with a hemocytometer. The obtained cells contain lymphocytes, macrophages, and neutrophils.
Flow cytometric analysis
The following Abs were used for flow cytometry: FITC-conjugated
anti-CD3 and PE-conjugated anti-NK1.1 mAbs (clones 145-2C11 and
PK136, respectively; BD PharMingen, San Diego, CA). Cells were
preincubated with anti-FcRIII mAb (clone 2.4G2; BD PharMingen)
on ice for 15 min in PBS containing 1% FCS and 0.1% sodium azide,
stained with these Abs for 25 min, and then washed three times in the
same buffer. Isotype-matched irrelevant Abs were used as a control
staining. In some experiments, cells were stained with PE-conjugated
-GalCer-loaded or unloaded CD1d tetramer (kind gifts of M.
Kronenberg, La Jolla Institute for Allergy and Immunology, San Diego,
CA) (38). The stained cells were analyzed using a EPICS XL
flow cytometer (Beckman Coulter, Fullerton, CA). Data were collected
from 15,000–20,000 individual cells using parameters of forward
scatter and side scatter to set a gate on lymphocyte population.
Extraction of RNA and RT-PCR
Total cellular RNA was extracted from the lungs after various
time periods of infection using Isogen (Wako Pure Chemical,
Osaka, Japan) followed by reverse transcription (30). The
obtained cDNA was then amplified in an automatic DNA thermal cycler
(PerkinElmer/Cetus, Norwalk, CT) using specific primers
5'-TCCATGCAGGTCCCTGTCATGCTT-3' (sense) and 5'-CTAGTTCACTGTCACACTGGTC-3'
(antisense) for MCP-1, 5'-CTAAGCACAGCACGCTGCACA-3' (sense) and
5'-AGGTATGACAATCAGCTGAGTCCC-3' (antisense) for TCR V14, and
5'-GTTGGATACAGGCCAAGACTTTGTTG-3' (sense) and
5'-GATTCAACTTGCGCTCATCTTAGGC-3' (antisense) for hypoxanthine
phosphoribosyl transferase (HPRT). We added 1.0 µl of the sample cDNA
solution to 49 µl of the reaction mixture, which contained the
following concentrations: 10 mM Tris-HCl (pH 8.3), 50 mM KCl, 1.5 mM
MgCl2, 10 µg/ml gelatin, dNTP (each at a concentration of 200 µM),
1.0 µM sense and antisense primer, and 1.25 U of AmpliTaq DNA
polymerase (PerkinElmer/Cetus). The mixture was incubated for 1 min at
94°C, 1 min at 62°C, and 1 min 45 s at 72°C for MCP-1 and
incubated for 1 min at 94°C, 1 min at 54°C, and 1 min 30 s at
72°C for HPRT. The number of cycles was determined for samples not
reaching the amplification plateau (28 cycles for HPRT and 32 cycles
for MCP-1). For TCRV14, the samples were heated at 94°C for 5 min
and then subjected to 35 cycles of incubation for 45 s at 94°C,
45 s at 55°C, and 90 s at 72°C, followed by a final
extension for 7 min at 72°C. The PCR products were electrophoresed on
2% agarose gels, stained with 0.5 µg/ml ethidium bromide, and
observed with a UV transilluminator. The obtained bands of amplified
DNA were quantitated using NIH Image software (version 1.61;
National Institutes of Health, Bethesda, MD), and the production of
V14 mRNA was expressed as a relative value to that of HPRT
mRNA.
In vitro stimulation of lung lymphocytes
Pulmonary leukocytes were prepared from three mice on day 7
after infection with C. neoformans and cultured at 4 x
105/well in flat-bottom microculture plates
(Falcon no. 3072; BD Biosciences, Franklin Lakes, NJ) with various
doses of viable organisms for 48 h. The culture supernatants were
collected and measured for the concentration of IFN- and IL-4 using
ELISA kits.
Measurement of cytokines
Murine IFN-, IL-4, and MCP-1 were measured by respective
ELISA kits (Endogen, Cambridge, MA, for IFN- and IL-4; and Toyobo,
Osaka, Japan, for MCP-1). The detection limits of assays for IFN-,
IL-4, and MCP-1 were 15, 5, and 9 pg/ml, respectively.
DTH response
Seven days after pulmonary infection, 1 x 106 yeast cells of C. neoformans (50 µl) were injected into the left hind foot pads of the mice and the same volume of PBS was injected into the right hind foot pads. The foot pad swelling was blindly measured 24 h later and expressed as the difference of thickness between left and right foot pads.
Enumeration of viable C. neoformans
Mice were sacrificed 1, 2, and 3 wk after infection, and lungs were dissected carefully and excised, then separately homogenized in 10 ml of distilled water by teasing with a stainless mesh at room temperature. The homogenates, appropriately diluted with distilled water, were inoculated at 100 µl on potato dextrose agar plates and cultured for 2–3 days, followed by counting the number of colonies.
Statistical analysis
Analysis of data was conducted using StatView, Japanese version (Hulinks, Tokyo, Japan), on a Macintosh computer (Apple Computer, Cupertino, CA). Data are expressed as mean ± SD. Differences between groups were examined for statistical significance using one-way ANOVA with a posthoc analysis (Fisher PLSD test). A p value < 0.05 was considered significant.
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Results |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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14 NKT cells in the lungs after infection with
C. neoformans
Initially, we elucidated whether NKT cells accumulated in the
lungs after infection with C. neoformans by determining the
proportion of these cells, identified as a lymphocyte subset double
positive for CD3 and NK1.1, among lung parenchymal leukocytes obtained
from mice infected intratracheally with this pathogen. As shown in Fig. 1
A, NKT cells formed only
0.5–0.7% of the lung lymphocytes before infection, but their
proportion commenced to increase on day 1, reached a peak level on day
7, and then decreased on day 14 postinfection. During the observation
period, the proportions of NK and T cells, identified as lymphocyte
subsets positive for either NK1.1 or CD3, respectively, increased with
similar kinetics as in NKT cells, although such increase was less
pronounced than in the latter cells (Fig. 1B). The absolute
number of NKT cells also markedly increased with a peak level on day 6
postinfection (Fig. 1C).
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14+ subset to
the increase of NKT cells, we compared the kinetics in the expansion of
NK1.1+CD3+ cells in the
lungs after infection with C. neoformans between V14 NKT
cell-deficient and WT mice. As shown in Fig. 2A, the proportion of NKT
cells increased even in V14 NKT cell-deficient mice, with a peak
level on day 6, although such accumulation was significantly lower on
day 6 compared with the control mice. These results suggested increased
proportion of V14 NKT cells in infected lungs. To confirm this
conclusion, two different experiments were conducted. First, using the
RT-PCR method, we examined the expression of V14 mRNA in the lungs
during the course of infection. As shown in Fig. 2, B and
C, such expression was weak in uninfected lungs, while a
significant increase of V14 mRNA was observed on days 3 and 6 after
infection with C. neoformans. Second, -GalCer-reactive
cells were examined by detecting the cells bound to
fluorescence-labeled -GalCer-loaded CD1d tetramer, which has been
reported to bind to V14 NKT cells in a specific manner
(38). Significant increase of positive cells was observed
in the lungs with a similar kinetics as in NKT cells postinfection,
while there was no change in the proportion of cells bound to
-GalCer-unloaded CD1d tetramer (Fig. 2, D and
E).
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14 NKT cell-deficient mice
Previous studies demonstrated the involvement of V14 NKT cells
in both Th1 and Th2 responses by producing large amounts of IFN- and
IL-4 upon stimulation through Ag receptors (1, 2, 4, 5),
but the role of these cells in host defense to infection remains to be
fully understood (19, 20, 21, 22, 23, 24, 25, 26). Therefore, we elucidated the
role of V14 NKT cells in Th1 response and host defense in
cryptococcal infection. In the first step, we compared the
concentration of IFN- in the culture supernatants of lung
lymphocytes derived from infected WT and V14 NKT cell-deficient mice
upon stimulation with various doses of live microorganisms. The
proportion of T cells was not different in the two strains of mice
(38.3 ± 0.4% vs 38.9 ± 4.3% in WT and V14 NKT
cell-deficient mice, respectively). As shown in Fig. 3, production of IFN- by lung
lymphocytes was significantly lower in V14 NKT cell-deficient mice
compared with WT mice. In contrast, there was no significant difference
in the production of IL-4 by restimulated lung lymphocytes between
these mice (data not shown). Second, we examined the DTH response to
C. neoformans by measuring the foot pad swelling of mice
rechallenged with Ags 7 days after infection. As shown in Fig. 4, such response was strongly reduced in
V14 NKT cell-deficient mice compared with WT mice. Finally, to test
the role of V14 NKT cells in host defense against infection with
C. neoformans, the clinical course of this infection was
compared between V14 NKT cell-deficient and WT mice by measuring the
number of live microorganisms in lungs. As shown in Fig. 5, elimination of C.
neoformans was significantly delayed in the former type of mice
compared with that in the latter.
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Several investigators previously demonstrated the involvement of
chemokines in the trafficking of NK cells to the sites of inflammation.
These chemokines included -chemokines such as IP-10 (39, 40), -chemokines including MCP-1, -2, -3, macrophage
inflammatory protein (MIP)-1, MIP-1, and RANTES
(40, 41, 42, 43), and -chemokines such as lymphotactin
(39, 44). In the present study, we focused on MCP-1 to
elucidate the regulatory mechanism for NKT cell accumulation in
infected organs. In the initial experiment, we examined the kinetics in
the production of this chemokine at both mRNA and protein levels in the
lungs after infection with C. neoformans. As shown in Fig. 6A, only a weak band of MCP-1
mRNA was identified in the uninfected lungs and the expression
commenced to increase on day 1, reached a peak level on day 3, and then
decreased on days 6–10. A similar kinetics was observed in MCP-1
protein synthesis in the lungs during the course of infection (Fig. 6B). These results indicated the production of MCP-1
preceded the increase of NKT cells in the lung and suggested that this
chemokine may be involved in the accumulation of NKT cells in the lungs
after infection with C. neoformans.
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A, the increase of NKT and NK
cells in lungs caused by infection was significantly less marked in
MCP-1-deficient mice than in WT mice on days 6 and 10 and days 1, 3, 6,
and 10, respectively, while such difference was not detected when the
numbers of conventional T cells were examined (data not shown). The
absolute number of NKT and NK cells in lungs was also significantly
lower in MCP-1-deficient mice than in control on days 3, 6, and 10
postinfection (Fig. 7B). In addition, we examined the effect
of MCP-1 expression on the increase of V14 NKT cells by measuring
the number of -GalCer-loaded CD1d tetramer-binding cells on day 6
postinfection. As shown in Fig. 7C, the increase of V14
NKT cells caused by infection was significantly less pronounced in
MCP-1-deficient mice compared with WT mice.
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Discussion |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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14
NKT cells in lungs after infection with this pathogen and the role that
NKT cells play in the induction of immunity to cryptococcal infection.
Furthermore, we elucidated the effect of MCP-1 on V14 NKT cells at
the site of infection using mice with a gene disruption for this
chemokine. The major findings of this study were 1) V14 NKT cells
increased in the lungs after intratracheal inoculation of C.
neoformans, 2) V14 NKT cells played an important role in the
induction of Th1 response and host resistance to this pathogen, and 3)
MCP-1 was involved in the recruitment of V14 NKT cells to the lungs
after C. neoformans infection.
Several studies have recently demonstrated the expansion of NKT cells
during infection. In leishmanial infection, the proportion of NKT
cells, as identified by double staining with anti-CD3 and
anti-IL-2R mAbs, increased in regional lymph nodes after s.c.
infection in resistant, but not susceptible, mice (24). In
another study, Pied et al. (45) revealed the expansion of
NKT cells, especially
CD4-CD8- double negative
subset, in the liver of mice after infection with Plasmodium
yoelii. Interestingly, CD4+ NKT cells
were rather decreased in these mice (45). Similar results
were reported by Weerasinghe and coworkers (46). In the
present study, the major subset of expanding NKT cells during
cryptococcal infection was negative for the expression of both CD4 and
CD8 (data not shown). We also showed the expansion of V14 NKT cells,
as identified by specific binding of -GalCer-loaded CD1d tetramer
and expression of V14 mRNA, along with an increase in the total
number of NKT cells. In malaria infection, one study indicated
increased expression of V7 and V2 gene segments, which have been
known to form semi-invariant TCRs with V14 (45),
while another study showed a reduction in the expression of V14 mRNA
(46). These observations suggested a regulatory role for
V14+ in addition to other subsets of NKT cells
in the early host defense to infection, although their contribution
might be distinct among infectious pathogens.
In earlier studies, Yoshimoto and coworkers (47)
demonstrated that activation of NKT cells by in vivo administration of
anti-CD3 mAb resulted in a rapid production of IL-4 and proposed
that this population may be the major source of early IL-4 production
which contributes to the differentiation of Th2 cells. However, several
studies subsequently indicated that Th2 response was not hampered in
2-microglobulin- or CD1d-deficient mice, which have
markedly reduced numbers of NKT cells (48, 49, 50, 51). Although
these findings made the role of these cells in Th2 cell development
obscure, such possibility was not completely excluded. For example,
V14 TCR-transgenic mice showed elevated serum levels of IgE and IL-4
(52), and activation of V14 NKT cells by -GalCer
induced T cell response to OVA polarized toward Th2-dominant condition
(9). In contrast, accumulating evidence emphasizes the
positive role of NKT cells in the development of Th1 cells. Activation
of V14 NKT cells by -GalCer led to the rapid production of
IFN- by themselves and other bystander cells, such as NK cells, in
vitro (53), and suppressed in vivo the Th2 differentiation
and subsequent IgE synthesis induced by OVA immunization or infection
with Nippostrongylus brasiliensis through the induction of
IFN- production (7). In other studies, NKT cells were
found to contribute to Th1-mediated responses, including granuloma
formation caused by mycobacterial lipid Ag (54) and the
IFN--mediated protection of mice against infection with malaria
parasites (25) and C. neoformans
(26) through the ligand-specific activation of NKT cells
by -GalCer. In the present study, we demonstrated that V14 NKT
cells played a critical role in the induction of Th1 response, such as
IFN- production and DTH response, during cryptococcal infection.
Thus, NKT cells are likely to contribute to the development of both Th1
and Th2 responses under distinct situations. Further studies are
necessary to clarify the precise mechanism of this regulation.
Several studies addressed the role of NKT cells in the host defense to
infectious pathogens. Behar and coworkers (19) compared
the clinical course of M. tuberculosis infection between
CD1d-deficient and control mice. They indicated that the lack of CD1d
expression, leading to deficiency of NKT cells, did not result in
exacerbation of mycobacterial infection. In another study, a
significant but small inhibition of host resistance to the same
pathogen was observed in mice treated with anti-CD1d mAb
(20). However, these authors concluded that NKT cells had
a minimal contribution, if any, to the host defense against
mycobacterial infection (55). Similar results were also
reported in S. choleraesuis infection by Ishigami et al.
(21); that is, the clinical course of this infection was
not different between V14 NKT cell-deficient and control mice,
although the hepatic injury was reduced in the former mice
(21). Szalay et al. (22) reported that
treatment with anti-CD1d mAb led to a transient amelioration of
infection with L. monocytogenes through enhanced production
of Th1-type cytokines. Similarly, depletion of NKT cells by
anti-IL-2R mAb treatment, but not of NK cells by anti-asialo
GM1 Ab, resulted in the protection of mice against T. gondii
infection, suggesting the negative regulatory role of NKT cells in host
defense to this pathogen (23). In contrast, the recent
studies of Ishikawa and colleagues (24) showed opposite
results; V14 NKT cells played a crucial role in the early stage of
protective immunity against L. major infection. Thus, the
role of NKT cells varies depending on the infectious pathogen. Our
study demonstrated the protective role of these cells in the host
defense against infection with C. neoformans. At present,
the precise mechanism of their different roles among infectious
pathogens remains to be elucidated, and further studies will be
required to determine the ligand of pathogens responsible for
activation of NKT cells before understanding such a mechanism.
There are two possible mechanisms for the increase of V14 NKT cells
in lungs after infection with C. neoformans: 1) local growth
at the infected sites and 2) recruitment from the peripheral
circulation. IL-15 is known to act as a major growth factor for NKT
cells, because mice deficient of IL-15R or IL-2/IL-15R lacked
such cells (56). In the present study, we did not examine
the expression of this cytokine at the site of infection and its
contribution to the increase of V14 NKT cells. Thus, the first
mechanism remains open for further study. In contrast, migration of NKT
cells into the lungs is a plausible mechanism, because we found that
NKT cells did not migrate to lungs of MCP-1-deficient mice in response
to cryptococcal infection. Compatibly, the expression of MCP-1 mRNA
preceded the increase of NKT cells after infection. In addition, we
also identified accumulation of NKT cells in the lungs after
intratracheal instillation of rMCP-1 (our unpublished data). To
our best knowledge, this is the first report showing the involvement of
MCP-1 in the in vivo accumulation of NKT cells, although MIP-2 has been
recently demonstrated as a chemoattractant for these cells
(57). At the same time, we also demonstrated the
contribution of MCP-1 to the recruitment of NK cells to the lungs
infected with C. neoformans. Similarly, other investigators
have reported the ability of this chemokine to attract resting or
activated NK cells under various conditions (41, 42, 43).
Other chemokines, including MCP-2 and -3, MIP-1, RANTES, IP-10, and
lymphotactin, were also identified as trafficking chemokines of NK
cells (39, 40, 41, 42, 43, 44), suggesting that these chemokines may act
on NKT cells in addition to MCP-1.
In conclusion, we demonstrated in the present study the MCP-1-dependent
increase of V14 NKT cells in the lungs after infection with C.
neoformans and their important roles in the development of Th1
response and host resistance to this infection. The natural ligand of
NKT cells remains to be elucidated, although some glycolipids, such as
GPIs, represent a potential candidate (3). At present, it
is not clear whether this fungal pathogen produces glycolipids
recognized by NKT cells. In this regard, Doering and coworkers
(58) recently provided evidence indicating the secretion
of GPIs from C. neoformans, as previously reported in
malaria parasite and Trypanosoma brucei (59).
To understand the precise mechanism of how NKT cells are involved in
the host defense to this infection, identification of pathogen-derived
ligands of these cells is desired.
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Acknowledgments |
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-GalCer-loaded and unloaded CD1d tetramers. |
Footnotes |
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2 Address correspondence and reprint requests to Dr. Kazuyoshi Kawakami, First Department of Internal Medicine, Faculty of Medicine, University of the Ryukyus, 207 Uehara, Nishihara, Okinawa 903-0215, Japan. E-mail address: kawakami{at}med.u-ryukyu.ac.jp
3 Abbreviations used in this paper: -GalCer, -galactosylceramide; DTH, delayed-type hypersensitivity; WT, wild type; MCP, monocyte chemoattractant protein; MIP, macrophage inflammatory protein.
Received for publication July 19, 2001. Accepted for publication October 5, 2001.
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14 NKT cells in IL-12-mediated rejection of tumors. Science 278:1623.14 NKT cells. Proc. Natl. Acad. Sci. USA 95:5690. T cells activated by IL-12 as a major effector in inhibition of experimental tumor metastasis. J. Immunol. 156:3366.[Abstract]
-galactosylceramide. J. Immunol. 163:2387.14+NK1.1+ T cells for liver injury induced by Salmonella infection in mice. Hepatology 29:1799.[Medline]
2 and Th1 cytokines and ameliorates listeriosis in mice. J. Immunol. 162:6955.14 NKT cells play a crucial role in an early stage of protective immunity against infection with Leishmania major. Int. Immunol. 12:1267.-Galactosylceramide-activated V14 natural killer T cells mediate protection against murine malaria. Proc. Natl. Acad. Sci. USA 97:8461.14+ natural killer T cells by -galactosylceramide results in development of Th1 response and local host resistance in mice infected with Cryptococcus neoformans. Infect. Immun. 69:213. production by NK cells. J. Immunol. 165:941.14 NKT cells. Proc. Natl. Acad. Sci. USA 95:5690.
and chemokines induce NK cell migration and enhance NK-mediated cytolysis. J. Immunol. 155:3877.[Abstract]
intermediate cells increase during experimental malaria infection and are able to exhibit inhibitory activity against the parasite liver stage in vitro. J. Immunol. 164:1463.2-microglobulin-dependent NK1.1+ T cells are not essential for T helper cell 2 immune responses. J. Exp. Med. 184:1295.2-microglobulin-dependent T cells are dispensable for allergen-induced T helper 2 responses. J. Exp. Med. 184:1507.This article has been cited by other articles:
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), and T cells (
) in lymphocyte population were
analyzed (B), and the absolute number of NKT cells was
calculated (C). Each symbol represents the mean ±
SD of four mice.
) tetramer. The lymphocyte
population was analyzed by flow cytometry. Each histogram shows the
representative staining profile of four mice on day 6 postinfection.
The number described in the histogram represents the percentage of
positive cells (D). Each symbol represents the mean
± SD of four mice (E).
