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Department of Molecular Microbiology and Immunology, University of Southern California/Norris Comprehensive Cancer Center, Keck School of Medicine, Los Angeles, CA 90089
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Abstract |
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 Top Abstract IntroductionMaterials and MethodsResultsDiscussionReferences |
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, inhibitable by their elimination before infection. NK cells
are shown to be necessary for optimal priming of virus-specific T
cells, assessed by delayed-type hypersensitivity response and CTL
activity, consistent with their ability to secrete IFN-. The
conclusion is drawn that NK cells mediate two important functions in
the liver: they induce cell death in the infected organ and
concomitantly stimulate the induction of T cell-mediated immunity by
release of IFN-. |
Introduction |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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(13). One
could therefore hypothesize that the liver is an organ in which NK
cells play an important role in the early response to infections. To
test this hypothesis, a viral infection model was used that consists of
a replication-defective type 5 adenovirus with deletions in the E1 and
E3 regions. Intravenous injection of this virus causes a high
efficiency infection in the liver, reflected in the expression of
virus-coded genes (14). Using this model we show that NK
cells exert a dual function: they induce apoptosis in hepatocytes and
concomitantly stimulate the induction of a T cell response. It is also
shown that elimination of NK cells before the infection leads to
inhibition of liver injury, secretion of IFN-, and induction of
virus-specific T cell responses. |
Materials and Methods |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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Pathogen-free female C57BL/6 (H-2b) and C57BL/6 nude mice, 6–12 wk of age, were obtained from The Jackson Laboratory (Bar Harbor, ME). BALB/C SCID mice were purchased from the National Institutes of Health breeding colony (Frederick Cancer Research Center, Frederick, MD). For immunization type 5 adenovirus with deletions in the E1 and E3 regions and carrying the lacZ gene was purchased from Microbix Biosystems (Toronto, Canada). Virus was propagated in 293 cells as provided by the supplier. In all experiments animals were injected with 2–3 x 109 PFU virus into the tail vein. Experiments were conducted in accordance with guidelines from the University of Southern California institutional animal care and use committee.
To deplete NK or NKT cells, mice were injected i.p. daily with either
15 µl of anti-AsGM1 (Wako Pure Chemicals Industries, Osaka,
Japan) or 250 µg/mouse of anti-NK1.1 mAb PK136 starting on day
-1 until termination of the experiment. To neutralize IFN-, mice
were injected with 250 µg/mouse anti-IFN- R4–6A2 (American
Type Culture Collection, Manassas, VA) on days -1, 2, 4, 6, and 8.
Control rat IgG1 (for anti-IFN- treatment) and control mouse
IgG2a Ab (for anti-NK1.1 treatment) were purchased from PharMingen
(San Diego, CA) and control rabbit Ig (for anti-AsGM1 treatment)
was purchased from Calbiochem (La Jolla, CA). Control Abs were injected
at equivalent doses and schedules. To activate NK cells in nude mice,
animals were injected once i.p. with 150 µg/mouse poly(I:C) (Sigma,
St. Louis, MO) on the day of virus injection (day 0). For assay of DTH
reactivity mice were challenged into the right footpad with
109 PFU of virus in 25–40 µl of PBS and in the
left footpad with PBS 9 days after virus infection. After 18 h
swelling was measured with a caliper gauge (Mitsutoyo, Tokyo, Japan),
and the difference in footpad diameters between left and right footpads
was determined.
For assay of serum ALT, mice were anesthetized with methoxyflurane (Pittman Moore, Mundelein, IL) and bled via the retro-orbital venous plexus. Serum (100 µl) was mixed with 1 ml of ALT assay solution (Sigma) and incubated for 90 s at 30°C. OD340 was measured in a spectrophotometer following the supplier’s protocol.
Preparation of liver lymphocytes and fluorometric analysis
Mononuclear cells were isolated from livers by passing tissue through a 200-gauge stainless steel mesh in serum-free HBSS (4, 6). The cell suspension was centrifuged 500 x g for 5 min, and the supernatant was discarded. The cell pellet from one liver was resuspended by vigorous vortexing in 20 ml of Percoll-HBSS containing 150 U/ml heparin. A Percoll working solution of 100 ml was prepared by mixing 92.5 ml of Percoll stock (Pharmacia Biotech, Uppsala, Sweden), with 7.2 ml of 10x PBS and 1.2 ml of 7.5% sodium bicarbonate, pH 7.2–7.4. Four parts of the Percoll working solution were mixed with 6 parts of HBSS, and the Percoll-HBSS solution was used for resuspension of cell pellets. The cell suspension was centrifuged at 800 x g for 20 min at room temperature, and the cell pellet was resuspended in 10 ml of RBC lysis solution. The lysis solution consists of 155 mM NH4Cl, 10 mM KHCO3, 1 mM EDTA, and 170 mM Tris, pH 7.3. After incubation for 10 min at room temperature (15), cells were harvested by centrifugation and washed twice in HBSS 5% FCS before use.
For FACS analysis 106 cells were stained with mAbs at a concentration of 1 µg/100 µl of PBS containing 0.2% BSA (Roche, Indianapolis, IN) and 0.05% sodium azide for 30 min on ice (4). The following Abs were used: PE- or FITC-conjugated anti-CD3 (145-2C11), PE- or FITC-conjugated anti-CD4 (GK1.5), PE- or FITC-conjugated anti-CD8 (53-6.7), and PE-conjugated anti-NK1.1 (PK136), all purchased from PharMingen (San Diego, CA). FACS analysis was performed on a FACStarPlus (Becton Dickinson, Mountain View, CA). The numbers of CD3+, CD4+, CD8+, NK, and NKT cells per liver were calculated by multiplying the percentage of each population with the total number of mononuclear cells per liver.
Induction of in vitro CTL responses and IFN--specific ELISPOT
assays
To assay CTL priming, spleen cells were harvested 10 days after virus infection and cultured at 5 x 106 cells/ml in complete RPMI 1640 medium containing 10% FCS for 5 days in 24-well plates (14, 16). Cultures received 2 PFU of virus/input cell as immunogen. To prepare targets, 107 C57SV (H-2b) cells were incubated with 50 PFU of virus/cell in 2 ml of DMEM/10% FCS for 2 h at 37°C. Ten milliliters of complete DMEM/10% FCS were added, and the incubation was continued for 24 h at 37°C. Virus-infected C57SV cells (2 x 106) were labeled with 100 µCi Na2[51Cr]O4 (DuPont, Boston, MA) in 5% FCS/RPMI 1640 for 120 min at 37°C. To demonstrate that cell lysis is due to CTL, effector cells were treated with anti-CD8 (AD4) or anti-CD4 (GK1.5) Ab and Low Tox-M rabbit complement (Accurate Chemical, Westbury, NY).
For IFN- ELISPOT assay (17) liver mononuclear cells
were isolated on day 6 after virus infection and seeded with YAC-1
targets into 96-well ELISPOT plates. To prepare the plates, 100 µl of
10 µg/ml anti-IFN- (R4-6A2; PharMingen) in PBS was pipetted
per well into Multiscreen 96-well filtration plates (Millipore,
Bedford, MA), followed by incubation overnight at 4°C. Plates were
washed three times with PBS, and a suspension of 5 x
105 YAC-1 cells and 1 x
105 liver mononuclear cells in 200 µl of RPMI
1640/10% FCS were added to each well and incubated for 24 h at
37°C. Medium was aspirated, and plates were washed three times in PBS
containing 0.05% Tween-20. Into each well 100 µl of a solution
containing 0.05% Tween-20, 1% BSA, and 5 µg/ml biotin-conjugated
anti-IFN- Ab (XMG1.2, PharMingen) in PBS were pipetted, and
plates were incubated overnight at 4°C. A solution containing a 1/400
dilution of 1 mg/ml avidin peroxidase (Sigma) in PBS containing 0.05%
Tween-20 and 1% BSA was prepared, and 100 µl was pipetted into each
well. After incubation for 2 h at room temperature, plates were
washed four times in PBS/0.05% Tween-20. Into each well 200 µl of
ABE solution (Zymed, South San Francisco, CA) were pipetted, followed
by incubation for 15 min in the dark. Plates were washed in
double-distilled H2O and dried for 2 h, and
spots were counted under microscope. To demonstrate that cytokine
secretion is due to NK cells, effector cells were treated before assay
with anti-NK1.1 Ab PK 136 and Low Tox complement
(16).
RT-PCR assay for cytokines, histology, and TUNEL staining
Total RNA was extracted from liver tissue by the
phenol/chloroform method using the RNAzol B kit (Tel-Test, Friendswood,
TX). Five micrograms of RNA was reverse transcribed to cDNA in a
50-µl reaction mixture using Superscript II RNase
H- reverse transcriptase and random primers
(Life Technologies, Grand Island, NY) according to the manufacturer’s
instructions. For PCR, the equivalent amount of cDNA product (5 µl)
was amplified in a 50-µl reaction mixture containing 10 mM Tris (pH
8.3), 50 mM KCl, 1.5 mM MgCl2, 0.2 mM dNTP, 2.5 U
Taq DNA polymerase (Perkin-Elmer, Norwalk, CT), and 1 µM of each
specific primer. The amplification was performed in a Thermoline Gene E
thermocycler (Techne, Cambridge, U.K.) set at 1 min each at 94, 58, and
72°C for 35 cycles, followed by an extension at 72°C for 10 min.
After amplification, PCR products were electrophoresed on a 2% agarose
gel and visualized by ethidium bromide staining under UV illumination.
Primers for IFN- and ß-actin were obtained from Stratagene (La
Jolla, CA).
For histology, liver tissue was fixed in 10% neutral buffered formalin and embedded in paraffin. Five-micron sections were affixed to slides, deparaffinized, and stained with hematoxylin and eosin to determine morphologic changes. Apoptotic cells were visualized by TUNEL staining in deparaffinized sections using an in situ cell death detection kit purchased from Roche.
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Results |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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Whereas NK and NKT cells constitute minor cell populations in the
peripheral lymphoid organs, they are relatively abundant in the liver
(5, 7). Fig. 1
shows that
mononuclear cells isolated from livers of normal C57BL/6 mice contain
7.1% NK1.1+ CD3- cells,
16.2% NK1.1+ CD3+ cells,
and 33.6% CD3+ NK1.1-
cells. In contrast spleens contain 3.6% NK1.1+
CD3- cells, 1.4% NK1.1+
CD3+ cells, and 31.8% CD3+
NK1.1- cells. Specific Abs are able to
selectively eliminate the two NK1.1+ cell
populations in vivo. Thus injection of anti-NK1.1 Ab PK136
eliminates both NK1.1+ CD3+
and NK1.1+ CD3- cells
(Fig. 1C), whereas anti-AsGM1 Ab causes almost complete
disappearance of NK1.1+
CD3- cells while sparing most
NK1.1+ CD3+ cells (Fig. 1D). The two Abs can therefore be used to assess the
relative contributions of the two NK cell populations during immune
responses.
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show that
serum ALT levels are suppressed in animals treated with either
anti-NK1.1 or anti-AsGM1 Ab. In contrast, injection of mouse or
rabbit control Abs had no demonstrable effect. These results indicate
that cells expressing Ags NK1.1 or AsGM1 are required for induction of
liver injury. To investigate the mechanism by which liver injury is
induced, livers were sectioned and assayed for apoptotic hepatocytes by
TUNEL staining. The results presented in Table I show that livers of normal mice
injected with virus contain a substantial number of apoptotic
hepatocytes. Treatment with anti-NK1.1 results in suppression of
the number of apoptotic cells to baseline levels in control mice. These
results suggest that hepatocyte death following the viral infection
requires the presence of NK1.1+ cells.
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The finding that injection of anti-NK1.1 and anti-AsGM1
Abs inhibits virus-induced liver injury raises the possibility that NK
cells, rather than T cells, are the principal effectors causing initial
liver injury. To examine this, use was made of mouse strains devoid of
T cells but possessing NK cells. Fig. 3A shows that injection
of virus into nude mice induces serum ALT values. However, to
reproducibly generate this effect, animals were primed with poly(I:C)
to activate NK cells, as different batches of mice, not primed with
poly(I:C), gave variable results. Treatment of poly(I:C)-primed mice
with either anti-NK1.1 or anti-AsGM1 Ab suppressed the
virus-induced increase in serum ALT (Fig. 3A), consistent
with the hypothesis that in nude mice effector cells expressing NK1.1
and AsGM1 cause injury in the infected liver.
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B that SCID mice respond with a substantial increase
in serum ALT after the viral infection. Interestingly, this response
does not require activation of NK cells by poly(I:C) and is completely
suppressed by injection of anti-AsGM1. TUNEL staining of liver
sections reveals an abundance of apoptotic hepatocytes in the liver of
infected SCID mice that is significantly suppressed in mice treated
with anti-AsGM1 (Table I). Therefore, neither T cells, B cells, nor
NKT cells are required for NK-mediated liver injury. Moreover, the
finding that in SCID mice NK cells do not have to be activated by
poly(I:C) suggests that in nude mice regulatory mechanisms can cause
suppression of NK activity. It is interesting that in SCID mice viral
infection initiates a slow increase in ALT values with kinetics similar
to those in normal mice (Figs. 2 and 3). It therefore appears that even
in SCID mice NK cells are activated by the infection before they induce
liver injury. NK1.1+AsGM1+ cells stimulate virus-specific T cell responses
The demonstration that NK cells mediate virus-induced liver injury
in nude and SCID mice raises the question as to the function of these
cells in normal mice, in particular as it relates to the putative role
of cytotoxic T cells in liver injury. Fluorometric analysis of stained
cells from liver infiltrates of infected normal mice reveals that virus
infection induces a dramatic increase in mononuclear cells in the liver
(Fig. 4A). Whereas the
increase in NK1.1+ cells appears to be quite
moderate, there is a dramatic increase in CD4+
and CD8+ T cells (Fig. 4B). It was
therefore important to test infected animals for priming of
virus-specific T cells. To monitor priming of
CD4+ cells, infected mice were challenged into
the footpad with virus, followed by evaluation of footpad
swelling to assess DTH reactivity. Fig. 5A shows that virus-primed
mice mount a robust DTH response, pointing to sensitization of Th1
cells. To assay for priming of cytotoxic T cells, spleens from
virus-injected mice were restimulated in vitro, followed by assay of
cytolytic activity. It is shown in Fig. 5B that cultures
from primed mice express high virus-specific cytolytic activity.
Treatment of the effector cells with anti-CD8 and complement before
the cytotoxicity assay inhibited cytolytic activity; in contrast,
treatment with anti-CD4 Ab had no detectable effect. These results
demonstrate that virus infection sensitizes virus-specific
CD8+ CTL.
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). A plausible explanation would be that NK cells mediate a dual
function, namely the early lysis of hepatocytes in infected liver,
followed by stimulation of virus-specific T cell responses. To examine
this, mice were treated with anti-NK1.1 or anti-AsGM1, injected
with virus, and tested for DTH and CTL responses. The results in Fig. 5C show that Ab-treated mice respond with a severely
suppressed DTH reaction. Similarly, spleen cells from Ab-treated mice
mount a lower virus-specific CTL response than cells from control mice
(Fig. 5D). These results suggest that cells expressing Ags NK1.1 and
AsGM1 play a stimulatory role in the induction of virus-specific
CD4+ and CD8+ cells.
IFN- is produced by NK cells and is required for optimal priming
of virus-specific CTL
The finding that cells expressing NK1.1 and AsGM1 stimulate the
priming of virus-specific CD4+ and
CD8+ cells raises the question as to the
mechanism by which this may occur. A plausible mode of action would be
if NK cells secrete a Th1 cytokine, e.g., IFN-, which facilitates T
cell priming. To examine this possibility, mononuclear cells isolated
from the livers of infected mice were assayed for secretion of IFN-
by ELISPOT assay. The results presented in Fig. 6 demonstrate that cells isolated from
the livers of virus-infected mice secrete IFN- upon incubation with
NK target YAC-1. In contrast, cells isolated from control mice contain
very few cells able to secrete IFN- when incubated with YAC-1
targets. Treatment of liver cell infiltrate from infected mice with
anti-NK1.1 and complement before incubation with YAC-1 targets
significantly decreased the number of IFN--secreting cells.
Therefore, the majority of cytokine-secreting cells were
NK1.1+ cells. Based on these results one would
predict that virus infection should induce IFN- transcripts in the
liver, and elimination of NK cells should inhibit this effect. It is
shown in Fig. 7A that in the
liver of normal mice, IFN- mRNA was induced by day 2 and even more
so on day 6 after infection. Treatment of mice with either
anti-NK1.1 or anti-AsGM1 before infection strongly suppressed
the induction of IFN- transcripts, consistent with the idea that NK
cells are producers of IFN- at early times after infection. Data
from T cell-deficient mice support this conclusion. In the livers of
nude mice IFN- mRNA was increased by the infection well above the
level present in poly(I:C)-primed mice, and this effect was suppressed
by injection of anti-NK1.1 and anti-AsGM1 Ab (Fig. 7B). Similar results were seen in SCID mice. Here again,
virus injection induced IFN- mRNA in the liver, and this effect was
suppressed in anti-AsGM1-treated animals (Fig. 7C).
Taken together these results suggest that NK cells constitute a source
of IFN- secretion at early times after the infection.
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and that elimination
of these cells leads to an inefficient priming of virus-specific T
cells supports the hypothesis that NK cells act via the secretion of
IFN-. For this case, then, elimination of this cytokine should
interfere with an efficient priming of CTL. To examine this, mice were
injected with anti-IFN- Ab and infected with virus, and their
spleens were restimulated with virus in vitro. The results presented in
Fig. 8 show that spleen cells from
anti-IFN--treated mice express lower CTL activity than spleen
cells from mice injected with control Ab. Therefore, the presence of
IFN- facilitates the priming of virus-specific CTL.
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Discussion |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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It is shown that NK cells in SCID mice can cause significant liver injury, reflected in high serum ALT and hepatocyte apoptosis, as demonstrable by TUNEL staining. These results clearly identify NK cells as a cell population capable of causing liver injury in the absence of T cells, B cells, and NKT cells. Given the demonstration that adenoviral gene expression in liver reaches a maximum between 36 and 48 h after infection (28), it is surprising that no elevated ALT values are demonstrable by day 3. This suggests that liver NK cells exist in a dormant state and have to be activated before they cause hepatocyte injury. It is likely that IL-12 and IL-18, produced by APC in the liver, are involved in this process (19, 29, 30). A surprising finding in this respect is that NK cells in nude mice require preactivation by poly(I:C) to reproducibly cause virus-induced liver injury. This may point to regulatory mechanisms, perhaps by NKT cells or CD3dim-staining cells, that modulate NK cell activity.
Our observation that serum ALT values increase in SCID mice with time kinetics similar to those in normal mice raises the possibility that even in the presence of T cells an early increase in ALT values is to a large extent due to the action of NK cells. In support it is shown that injection of Abs anti-NK1.1 and anti-AsGM1 into normal mice inhibits the increase in ALT. The demonstration that anti-AsGM1 leaves NKT cells relatively unaffected while efficiently eliminating NK cells suggests that NK cells constitute a major cell type responsible for early injury in the virus-infected liver of immunocompetent animals.
The conclusion that in normal mice NK cells are responsible for early
liver injury is unexpected, as NKT cells constitute a far larger
population of cells in liver with well-documented cytolytic activity
(20, 21, 31, 32, 33). It is therefore important to stress that
Ab ablation experiments do not completely exclude participation of NKT
cells in cytotoxic liver injury. It is possible, for example, that
activated NKT cells with high cytolytic activity increase cell surface
expression of AsGM1, thereby making them sensitive to elimination by
anti-AsGM1 in vivo. A most intriguing possibility regarding the
function of NKT cells in liver is raised by the recent observation that
NKT cells may stimulate NK cells by secretion of IFN-
(34). Therefore, NKT cells could provide helper functions
for NK cells by stimulating their activity in the liver. These
functions could provide a rationale for the relative abundance of NKT
cells in this organ.
The demonstration that NK cells play an important role in the early response to infection of the liver is in agreement with results from related models. Thus, NK cells have been shown to mediate the clearance of vaccinia/IL-2 infections in nude mice (35), and removal of NK cells from murine CMV-infected normal mice prompts an increase in the severity of viral hepatitis (2). In these experiments replication-competent virus had been employed, leaving open the question of how NK cells eliminate the infectious pathogen, i.e., by inhibiting viral replication via cytokine secretion or by cytotoxicity. Our approach of using a replication-defective virus clearly shows that hepatocyte apoptosis is induced by NK cells. The question, however, that remains to be resolved is whether hepatocyte lysis by NK cells is specifically directed against virus-infected cells. Our attempts at demonstrating that ex vivo NK cells from the liver of infected mice are virus specific have not been successful (unpublished observations). It is therefore quite possible that liver injury induced by NK cells is not specifically directed against virus-infected cells.
Demonstration of NK-mediated cytotoxicity in infected liver raises the
question of whether this constitutes the principal function of these
cells. We show that elimination of NK1.1+ and
AsGM1+ cells interferes with efficient priming of
virus-specific CTL and DTH responses, suggesting that NK cells mediate
additional functions. Using an ELISPOT assay we demonstrate that NK
cells from virus-infected livers, when stimulated with NK target YAC-1,
secrete IFN-. Moreover, infection of normal or T cell-deficient mice
with virus induces IFN- mRNA, which is inhibited in NK-depleted
mice. Therefore, NK cells produce IFN- in response to the infection,
raising the question of whether it is this process that causes
stimulation of T cell priming. Support for this mechanism is provided
by the finding that injection of anti-IFN- Ab inhibits CTL
priming. This effect, however, is not complete, leaving open the
possibility that additional cytokines, released by NK cells, are also
involved.
Another unresolved question is the possible role of NKT cells in
cytokine secretion. The observation that NKT cells, when stimulated via
their TCR, very rapidly engage in the secretion of IFN-, which, in
turn, stimulates NK cells to secrete this cytokine, could provide a
mechanism for activation of NK cells in the stimulation of T cell
responses (34, 36, 37). There are a limited number of
reports supporting this attractive hypothesis. NKT cells have been
shown to stimulate the induction of CD8+ effector
cells in Toxoplasma infection (36), and priming
of influenza virus-specific CTL was found to be inhibited in NK
cell-depleted mice (38). The in vivo induction of mouse
CTL, specific for Plasmodium, involving infection of
hepatocytes was also reported to depend on the presence of NK cells
(39). These findings together with the ones reported here
provide strong support for the idea that NK cells in conjunction with
NKT cells may play an important stimulatory role in the induction of
Th1 and CTL responses.
Our finding that NK cells secrete IFN- following adenovirus
injection is in line with previous reports showing that NK cells
secrete IFN- in response to viral infections, and this causes
suppression of virus replication (2, 40, 41). This, then,
raises the question of the mechanism by which IFN- stimulates
priming of T cell responses. Several possibilities exist. It is well
documented that MHC expression on hepatocytes is almost undetectable
and is stimulated by IFN- (13). Therefore, NK
cell-derived IFN- could increase MHC expression on hepatocytes,
which, in turn, could inhibit NK function while stimulating T cell
responses. In support of this, it has been shown that isolated
hepatocytes are able to induce CTL responses in vitro; hence,
hepatocytes can act as APC (42). An alternative action of
IFN- could be direct stimulation of CD4+ or
CD8+ cells during priming or restimulation in the
liver. Yet another mechanism is that IFN- can stimulate immature
liver dendritic cells to migrate to the peripheral lymphatics and to
induce a Th1 response (43).
In summary, we show here that NK cells play a dual role in the
cell-mediated immune response to adenoviral infection in the liver.
They lyse hepatocytes in the virus-infected liver and stimulate,
probably by their ability to secrete IFN-, the induction of a
virus-specific, T cell-mediated immune response. It is therefore
suggested that NK cells constitute a very important component required
for optimal responses to viral liver infections and thereby may hold
the key to a successful cell-mediated immune response in this
organ.
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Footnotes |
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2 Address correspondence and reprint requests to Dr. Gunther Dennert, University of Southern California/Norris Comprehensive Cancer Center, 1441 Eastlake Avenue, M/S #73, Los Angeles, CA 90089-9176.
3 Abbreviations used in this paper: DTH, delayed-type hypersensitivity; AsGM1, asialo-GM1; ALT, alanine aminotransferase; ELISPOT, enzyme-linked immunospot; lacZ, ß-galactosidase.
Received for publication December 20, 1999.
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M. J. Cotter, A. K. Zaiss, and D. A. Muruve Neutrophils Interact with Adenovirus Vectors via Fc Receptors and Complement Receptor 1 J. Virol., December 1, 2005; 79(23): 14622 - 14631. [Abstract] [Full Text] [PDF]
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T. Delale, A. Paquin, C. Asselin-Paturel, M. Dalod, G. Brizard, E. E. M. Bates, P. Kastner, S. Chan, S. Akira, A. Vicari, et al. MyD88-Dependent and -Independent Murine Cytomegalovirus Sensing for IFN-{alpha} Release and Initiation of Immune Responses In Vivo J. Immunol., November 15, 2005; 175(10): 6723 - 6732. [Abstract] [Full Text] [PDF]
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M. Biburger and G. Tiegs {alpha}-Galactosylceramide-Induced Liver Injury in Mice Is Mediated by TNF-{alpha} but Independent of Kupffer Cells J. Immunol., August 1, 2005; 175(3): 1540 - 1550. [Abstract] [Full Text] [PDF]
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 R. J. McKallip, M. Fisher, U. Gunthert, A. K. Szakal, P. S. Nagarkatti, and M. Nagarkatti Role of CD44 and Its v7 Isoform in Staphylococcal Enterotoxin B-Induced Toxic Shock: CD44 Deficiency on Hepatic Mononuclear Cells Leads to Reduced Activation-Induced Apoptosis That Results in Increased Liver Damage Infect. Immun., January 1, 2005; 73(1): 50 - 61. [Abstract] [Full Text] [PDF]
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A. Ahmad and F. Alvarez Role of NK and NKT cells in the immunopathogenesis of HCV-induced hepatitis J. Leukoc. Biol., October 1, 2004; 76(4): 743 - 759. [Abstract] [Full Text] [PDF]
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K. A. Muhlen, J. Schumann, F. Wittke, S. Stenger, N. van Rooijen, L. van Kaer, and G. Tiegs NK Cells, but Not NKT Cells, Are Involved in Pseudomonas aeruginosa Exotoxin A-Induced Hepatotoxicity in Mice J. Immunol., March 1, 2004; 172(5): 3034 - 3041. [Abstract] [Full Text] [PDF]
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K. Herzer, C. S. Falk, J. Encke, S. T. Eichhorst, A. Ulsenheimer, B. Seliger, and P. H. Krammer Upregulation of Major Histocompatibility Complex Class I on Liver Cells by Hepatitis C Virus Core Protein via p53 and TAP1 Impairs Natural Killer Cell Cytotoxicity J. Virol., August 1, 2003; 77(15): 8299 - 8309. [Abstract] [Full Text] [PDF]
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J. Schumann, K. Muhlen, A. K. Kiemer, A. M. Vollmar, and G. Tiegs Parenchymal, But Not Leukocyte, TNF Receptor 2 Mediates T Cell-Dependent Hepatitis in Mice J. Immunol., February 15, 2003; 170(4): 2129 - 2137. [Abstract] [Full Text] [PDF]
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M. Thomson, M. Nascimbeni, M. B. Havert, M. Major, S. Gonzales, H. Alter, S. M. Feinstone, K. K. Murthy, B. Rehermann, and T. J. Liang The Clearance of Hepatitis C Virus Infection in Chimpanzees May Not Necessarily Correlate with the Appearance of Acquired Immunity J. Virol., December 20, 2002; 77(2): 862 - 870. [Abstract] [Full Text] [PDF]
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H.-G. Zhang, J. Xie, L. Xu, P. Yang, X. Xu, S. Sun, Y. Wang, D. T. Curiel, H.-C. Hsu, and J. D. Mountz Hepatic DR5 Induces Apoptosis and Limits Adenovirus Gene Therapy Product Expression in the Liver J. Virol., May 3, 2002; 76(11): 5692 - 5700. [Abstract] [Full Text] [PDF]
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C. L. Thio, X. Gao, J. J. Goedert, D. Vlahov, K. E. Nelson, M. W. Hilgartner, S. J. O'Brien, P. Karacki, J. Astemborski, M. Carrington, et al. HLA-Cw*04 and Hepatitis C Virus Persistence J. Virol., April 16, 2002; 76(10): 4792 - 4797. [Abstract] [Full Text] [PDF]
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Z.-X. Liu, H. Nishida, J.-W. He, M. M. C. Lai, N. Feng, and G. Dennert Hepatitis C Virus Genotype 1b Core Protein Does Not Exert Immunomodulatory Effects on Virus-Induced Cellular Immunity J. Virol., February 1, 2002; 76(3): 990 - 997. [Abstract] [Full Text] [PDF]
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Y. Peng, E. Falck-Pedersen, and K. B. Elkon Variation in Adenovirus Transgene Expression between BALB/c and C57BL/6 Mice Is Associated with Differences in Interleukin-12 and Gamma Interferon Production and NK Cell Activation J. Virol., May 15, 2001; 75(10): 4540 - 4550. [Abstract] [Full Text]
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 M. J. Smyth, E. Cretney, K. Takeda, R. H. Wiltrout, L. M. Sedger, N. Kayagaki, H. Yagita, and K. Okumura Tumor Necrosis Factor-related Apoptosis-inducing Ligand (TRAIL) Contributes to Interferon {{gamma}}-dependent Natural Killer Cell Protection from Tumor Metastasis J. Exp. Med., March 12, 2001; 193(6): 661 - 670. [Abstract] [Full Text] [PDF]
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Z.-X. Liu, S. Govindarajan, S. Okamoto, and G. Dennert Fas-Mediated Apoptosis Causes Elimination of Virus-Specific Cytotoxic T Cells in the Virus-Infected Liver J. Immunol., March 1, 2001; 166(5): 3035 - 3041. [Abstract] [Full Text] [PDF]
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) or 2 x
109 PFU virus (
) on day 0. On day 10 mice were
challenged in the footpad with virus, and DTH was assayed. Averages
from groups of five mice, including SD values, are plotted. Spleens
from control or virus-injected mice were harvested on day 10 and
restimulated with virus in vitro for 5 days. Virus-specific
cytotoxicity is plotted. Where indicated, effector cells were treated
with anti-CD8 or anti-CD4 and complement before assay. Error
bars constitute SD values from triplicate wells. C and
D, Effect of anti-NK1.1 and anti-AsGM1 on T cell
sensitization. Mice were infected with 2 x 109 PFU
virus on day 0 and injected with Abs from days -1 to 9. After virus
challenge in the footpad on day 10, DTH was assayed. Spleen cells from
animals injected with Ab from days -1 to 9 were harvested on day 10,
restimulated with virus in vitro for 5 days, and assayed for
virus-specific cytotoxicity. Responses in cultures from normal
virus-primed mice and anti-AsGM1 or anti-NK1.1 treated mice are
shown. Error bars constitute SD values from triplicate wells in the
cytotoxicity assay.
