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Immunobiology and
Digestive Diseases, Yale University School of Medicine, New Haven, CT 06510
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Abstract |
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 Top Abstract IntroductionMaterials and MethodsResultsDiscussionReferences |
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Introduction |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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We and others have reported the accumulation and apoptosis of activated CD8+ T cells in the liver during systemic immune responses (7, 8, 9, 10). ICAM-1 appears to be important in this process. When a mixture of resting and activated T cells were perfused through a mouse liver, the activated CD8+ T were selectively retained (11), but this retention was compromised in an ICAM-1-deficient liver. Most of the retained CD8+ T cells were in contact with Kupffer cells, a highly mobile population of hepatic macrophages. By 14 h, a proportion of the trapped CD8+ T cells had begun to undergo apoptosis. These findings support the idea that the liver is a trap for activated CD8+ T cells and that such trapping results in their apoptosis.
In many experimental and physiological immune responses, Ag presented on liver cells leads to tolerance, which may be linked to T cell apoptosis (12, 13, 14). To explain the tolerogenic property of liver Ags, we propose the hypothesis that recognition of Ag on the liver tissue (i.e., on sinusoidal endothelium and/or hepatocytes) promotes T cell trapping and apoptotic death. This hypothesis predicts that the capacity of hepatic non-bone marrow-derived cells to present Ag will control the trapping and apoptosis of CD8+ T cells in the liver and thus regulate CD8+ T cell removal from the circulating pool.
To test these predictions, we developed a model in which naive CD8+ T cells from the OT-1 transgenic mouse, expressing a TCR specific for the OVA peptide SIINFEKL, were adoptively transferred into chimeric mice which had received lethal irradiation followed by reconstitution with donor bone marrow. These chimeras were constructed so that Ag presentation could occur on all cells, or alternatively was restricted either to bone marrow-derived cells, or to non-bone marrow-derived cells. Systemic activation of CD8+ T cells resulted in accumulation and apoptosis of activating CD8+ T cells in the liver. The ability of non-bone marrow-derived cells to present Ag was essential for the hepatic accumulation of CD8+ T cells. Chimeras in which these tissues were unable to present Ag accumulated fewer T cells in the liver, and this was associated with impaired deletion of activated CD8+ T cells from the spleen. This supports a role for the liver in systemic CD8+ T cell homeostasis.
Immune mediated liver injury occurs in a wide variety of systemic immune responses (15, 16). Such injury of hepatocytes may occur by classical, MHC-restricted recognition of specific antigenic peptides on hepatocytes, e.g., during immunopathology associated with the CD8+ T cell response to hepatitis B virus Ags (17). It has also been proposed that physiological T cell activation can result in hepatic damage simply because of the presence of activated cytotoxic cells T cells in the liver, a mechanism termed collateral damage (18). In the present series of experiments, chimeric mice in which the hepatocytes were unable to support Ag recognition nevertheless showed evidence of liver damage associated with an activated CD8+ T cell influx. This study thus provides evidence for the collateral damage mechanism.
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Materials and Methods |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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Nontransgenic C57BL/6J and H2-Kbm1 mice (on the C57BL/6J background) and C57BL/6 mice of the CD45.1 allotype were purchased from The Jackson Laboratory (Bar Harbor, ME). A colony of OT-1-transgenic mice was maintained on the C57BL/6J background. All animals were housed in a specific pathogen-free environment, in accordance with institutional guidelines for animal care.
To construct chimeras, 6- to 8-wk-old recipient mice were irradiated
with 13 Gy (1300 rad) as a single dose from a cesium irradiator. Mature
T cells were removed from donor bone marrow cell suspensions by
complement-mediated lysis of T cells, and between 10 and 15 x
106 T cell-depleted bone marrow cells were
injected i.v. within 4 h of irradiation. All chimeras were allowed
to reconstitute for 2 mo before further experimentation. Three types of
chimeras were made: C57BL/6J
C57BL/6J;
H2-Kbm1C57BL/6J; and
C57BL/6JH2-Kbm1.
Adoptive transfer and in vivo activation
Donor OT-1 mice were killed by CO2 narcosis, and a cell suspension was obtained by mechanical homogenization of axillary and abdominal lymph nodes. RBC were removed by density gradient separation (Lymphocyte Separation Medium; ICN Biomedicals, Aurora, OH). MHC class II-positive dendritic cells and B cells were removed using a primary Ab (clone 212.A1 specific for MHC class II molecules, and clone 2.4-G2 specific for FcRs). Magnetic beads coated with secondary Abs were used to remove the cells coated with primary Ab. CD4+ T cells were removed by magnetic beads directly coupled to anti-CD4+ Ab (clone MT30). A suspension of 5 x 106 lymphocytes, containing >95% CD8+ T cells, was injected i.v. into each recipient.
OT-1 T cells were activated after adoptive transfer by daily i.p. injections of 25 nM SIINFEKL peptide for 3 days starting on day 2, as previously described (9). Control mice received saline. To control for any liver toxicity of the SIINFEKL peptide, chimeras without OT-1 cells received the same dosage of the peptide.
Cell isolation, staining, and flow cytometric analysis
At days 3, 5, and 7 after the first peptide or saline injections, mice were anesthetized with methoxyflurane (Schering-Plough Animal Health, Kenilworth, NJ), and exsanguinated by cutting the abdominal aorta and vena cava. Blood was collected from the abdominal cavity using a heparinized 1-ml syringe, and the serum was assayed for the enzymes aspartate aminotransminase (AST) and alanine aminotransaminase (ALT) using a multichannel analyzer.
Intrahepatic lymphocytes were isolated by perfusion of the liver with digestion buffer consisting of Bruff’s medium containing 0.02% collagenase IV (Sigma, St. Louis, MO), 0.002% DNase I (Sigma), and 5% FCS. The digestion buffer was infused into the portal vein using a 5-ml syringe and a 21-gauge needle during 1–2 min. Care was taken to minimize injection of air bubbles into the portal vein, and blanching of the whole liver was used as an indicator of adequate perfusion. After perfusion, the liver was dissected out of the abdominal cavity and homogenized by forcing through a fine metal strainer. The homogenized liver was incubated with 10 ml digestion buffer at 37°C for 30 min in a shaking water bath. The enzymatically digested liver cell suspension was centrifuged at 10 x g for 3 min at 4°C to remove hepatocytes and cell clumps. The supernatant was then centrifuged at 120 x g for 10 min to obtain a pellet of cells depleted of hepatocytes. The volume of the pellet was typically 0.3–0.5 ml, and it was suspended with Bruff’s medium to a final volume of 1 ml, before being mixed with 4 ml 30% metrizamide in Bruff’s medium. This procedure resulted in 5 ml cell suspension in 24% metrizamide, which was layered under 1 ml serum-free Bruff’s medium and centrifuged at 1500 x g for 20 min at 4°C in 15-ml conical centrifuge tubes (Falcon, Franklin Lake, NJ). The cells at the interface were collected, washed with PBS, and counted before analysis using a FACS.
As a control to discriminate liver-specific effects on T cell accumulation and apoptosis from generic properties of nonlymphoid organs, kidney T cells were analyzed in parallel. Kidneys were cut into 2- to 3-mm slices, homogenized by forcing through a metal strainer, and then digested with collagenase. Renal lymphocytes were isolated using metrizamide, as described above. The spleen was dissected, homogenized and RBC were lysed using RBC lysing buffer (Sigma). Lymphocytes were washed and counted before staining for FACS analysis. Sections of liver and kidney were cut and fixed in 1% paraformaldehyde in PBS for histological analysis before homogenization.
Cells were adjusted to 2 x 107/ml in
staining buffer (saline with 1% bovine albumin). Fifty microliters of
the cell suspension were incubated with Ab on ice for 30 min, washed
with staining buffer, and fixed with 2% paraformaldehyde. FACS data
were acquired using a FACSCalibur flow cytometer (BD Biosciences, San
Jose, CA), set to acquire all events. The Abs used for staining were
TCR
(clone H57-597), CD45.1 (clone A20), CD45.2 (clone 104),
L-selectin (clone MEL-14), LFA-1 (clone 2D7), V2 (clone B20.1), and
CD8 (clone 53-6.7). For TUNEL staining of cell suspensions and tissue
sections, the In Situ Cell Death Detection Kit-Fluorescein
(Boehringer Mannheim, Indianapolis, IN) was used according to the
manufacturer’s instructions. FACS data were analyzed using CellQuest
software (BD Biosciences).
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Results |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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Peripheral deletion and massive liver accumulation and apoptosis of the responding cells were previously reported using TCR-transgenic mice (7, 9), leaving open the possibility that these effects might be limited to mice with a very high frequency of responding T cells. To test this and to establish a system to determine the role of hepatic Ag presentation in this phenomenon, we adoptively transferred 5 x 106 CD8+ T cells from OT-1 mice on the C57BL/6J background into unmanipulated nontransgenic C57BL/6J mice (19). The OT-1 mice are transgenic for a TCR that recognizes the 8-mer OVA peptide, SIINFEKL, in association with the MHC molecule H2-Kb. In all of these experiments, the adoptively transferred T cells were CD45.1/CD45.2 heterozygous, permitting their identification by FACS.
After adoptive transfer, the OT-1 cells were present at low frequency
(1% or less) in the spleen, liver, and kidney. Daily i.p. injections
of saline did not alter the total numbers of lymphocytes or the
percentage of OT-1 cells during 7 days (Fig. 1
). Daily injections of 250 µl 100 µM SIINFEKL peptide (i.e., 25 nM
peptide) beginning on day 2 after adoptive transfer resulted in no
significant change in cell numbers in the spleen or kidney, but an
7-fold increase in the number of intrahepatic lymphocytes. The
percentage of OT-1 cells increased in all three organs after peptide
administration, but the largest increase was in the liver at 25-fold,
compared with the spleen and kidney where the increase was 7- and
10-fold, respectively (Fig. 1). The number of OT-1 cells in each organ
was calculated by multiplying the total number of lymphocytes by the
percentage of OT-1 cells. The maximum number of OT-1 cells in the
spleen was at day 3 after peptide injection, at 12.4 ± 3.8
x 106, decreasing to 5.2 ± 3.1 x
106 on day 5, whereas maximum accumulation in the
liver was later, with 6.3 ± 2.4 x 106
OT-1 cells at day 5. By comparison the kidney contained relatively few
OT-1 cells, 7.4 ± 1.8 x 104. In all,
there was a 170-fold increase in the total number of OT-1 cells in the
liver, compared with 13- and 24-fold in the spleen and kidney,
respectively.
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A key finding in earlier studies was the high frequency of apoptotic CD8+ T cells accumulating in the liver (7, 8, 9, 10). This was confirmed in the present study, in which the percentage of OT-1 cells which were TUNEL positive in the spleen, liver, and kidney were 8.4 ± 2.7%, 18.3 ± 5.2% and 9.6 ± 4.3%, respectively. In the PBS-injected control chimeras, there were too few OT-1 cells to obtain TUNEL information.
Ag presentation by non-bone marrow-derived cells enhances intraheptic accumulation of activated CD8+ T cells
The OT-1-transgenic TCR recognizes the SIINFEKL peptide in association with H2-Kb. The H2 molecule H2-Kbm1 differs from H2-Kb by three amino acids at positions 152, 155, and 156, and this difference is sufficient to prevent effective presentation of the SIINFEKL peptide (21).
In the C57BL/6J (H2-Kb) mice, all cells of the
liver were able to present the SIINFEKL peptide to OT-1 T cells. To
study the effect on liver accumulation of OT-1 T cells in the absence
of Ag presentation by non-bone marrow-derived cells, we generated
chimeras in which C57BL/6J bone marrow was infused into
B6.C-H2bm1 (H2-Kbm1)
hosts (B6bm1). In these chimeras, Ag presentation to OT-1 cells was
possible by bone marrow-derived cells only. C57BL/6J into C57BL/6J
(B6B6) and B6.C-H2bm1 into C57BL/6 of the
CD45.1 allotype (bm1B6.CD45.1) provided the controls. The negative
control of B6.C-H2bm1 into
B6.C-H2bm1 chimeras are not suitable recipients
for OT-1 cells, because they mount an alloimmune response to the
H2-Kb molecule expressed on the T cells. In
control experiments, OT-1 cells transferred into intact
B6.C-H2bm1 hosts simply disappeared, probably
due to allorejection. The donors and recipients for B6B6 and
B6bm1 were CD45.2, but the recipients for bm1B6.CD45.1 chimeras
were C57BL/6J mice with the CD45.1 allotype. This allowed analysis of
the percentage of donor bone marrow-derived cells (CD45.2), recipient
bone marrow-derived cells (CD45.1), and donor OT-1 T cells (CD45.1/CD
45.2 heterozygous) (see Fig. 2).
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shows clonal expansion of OT-1 cells
in the spleens of all three groups of chimeras at day 5 after peptide
injection. OT-1 T cell expansion in B6B6 and B6bm1 chimeras was
expected, due to the presence of H2-Kb on bone
marrow cells. Expansion in the bm1B6.CD45.1 was likely due to the
persistence of recipient bone marrow-derived cells despite irradiation,
and these are identifiable by FACS analysis as CD45.1 positive but
CD45.2 negative (Fig. 2). They constituted 4.3 ± 1.4% of all the
bone marrow-derived cells in the spleens of bm1B6 chimeras injected
with PBS. Therefore, the full activation of OT-1 cells in these
chimeras should not be taken to imply that activation was independent
of bone marrow-derived cells.
Fig. 3
show the percentage and total number of OT-1 T cells in the livers of
the three groups of chimeras after 5 days of saline or peptide
injection. Peptide injection resulted in accumulation of OT-1 cells in
the livers of mice in all three groups of chimeras. B6B6 and
bm1B6.CD45.1 chimeras were able to present SIINFEKL peptide on
non-bone marrow-derived cells, and the number of OT-1 cells were
similar to those seen in normal C57BL/6 mice. These numbers were
6.4 x 106 ± 1.7 x
106 in the B6B6 and 5.9 x
106 ± 1.1 x 106 in
the bm1B6 chimeras, and these numbers were not significantly
different by an unpaired t test (p =
0.5). In B6bm1 chimeras, hepatocytes and endothelial cells were
unable to present Ag, and the numbers of OT-1 cells in the livers of
these chimeras was 2.4 x 106 ± 0.9 x
106, which is significantly different from both
the value in B6B6 chimeras (p = 0.002) and
the value in bm1B6 chimeras (p = 0.001).
There were no significant differences among the three groups of
chimeras in the number of OT-1 cells in the kidney after peptide
injection (Fig. 3, E and F, p >
0.8 in all cases).
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The greater liver accumulation of OT-1 cells in B6B6 chimeras
relative to B6bm1 was associated with lower percentages and total
numbers of OT-1 cells in the spleen of B6B6 chimeras (Fig. 3, C and D). Thus, the B6B6 chimeras contained
3.22 x 106 ± 1.78 x
106 OT-1 cells, whereas the B6bm1 chimeras
contained 10.5 x 106 ± 2.0 x
106 OT-1 T cells, a difference that was
significant based on an unpaired t test
(p = 0.001). This suggests that the removal of
activated OT-1 cells from the spleen was compromised because of the
reduced accumulation of OT-1 cells in the liver of the B6bm1
chimeras.
The B6B6 and B6bm1 chimeras do not differ only in Ag presentation
by hepatocytes and endothelial cells, because non-bone marrow-derived
cells of other organs in the B6bm1 chimeras would also be unable to
present the SIINFEKL peptide to OT-1 cells. To study the effect of Ag
presentation by non-bone marrow-derived cells in another organ, the
percentage and total number of OT-1 cells in the kidneys were
determined in chimeras of all three groups. There was an 15- to
24-fold increase in the number of OT-1 cells in the kidney in
peptide-injected mice compared with PBS-injected controls (from
2 x 103 to 40 x
103). There were, however, no significant
differences between different groups of chimeras in the total number of
OT-1 cells in the kidneys. The kidneys of B6B6 chimeras contained
48 x 103 ± 18 x
103 OT-1 cells, the kidneys of B6bm1 chimeras
contained 49 x 103 ± 21 x
103 OT-1 cells, and the kidneys of bm1B6
chimeras contained 45 x 103 ± 21 x
103 OT-1 cells. None of these differences was
significant by an unpaired t test (p
> 0.8 in every case).
Thus, in the absence of an inflammatory stimulus, the presence of Ag on
tissue cells did not enhance the localization of activated T cells to
the kidney. An experiment conducted in transgenic mice that expressed
hepatitis B Ags in many tissues supports this interpretation, because
the introduction of Ag-specific CTL into these mice resulted in
immunopathology only in the liver (22). The liver may be
unique in its capacity to allow such interactions, because both Kupffer
cells and sinusoidal endothelial cells express a high resting level of
ICAM-1 (5, 6). In our mice, the systemic vasculature was
not inflamed, and based on this argument plus the kidney data, we would
argue that the lack of Ag presentation by non-bone marrow-derived cells
in B6bm1 chimeras most likely did not affect the localization of
OT-1 cells to organs other than the liver.
Increased apoptosis of activated CD8+ T cells retained in livers with Ag presentation limited to bone marrow-derived cells
The percentage of TUNEL-positive OT-1 cells was consistently
higher in the liver, compared with the spleen and kidney, in all three
groups of chimeras (Fig. 4). This percentage was similar to that observed in nonchimeric mice
(7, 8, 9, 10). The total number of TUNEL-positive OT-1 cells
was, however, lower in B6bm1 chimeras (0.5 ± 0.3 x
106) compared with chimeras B6B6 and
bm1B6.CD45. (1.2 ± 0.49, 1.1 ± 0.4 x
106). This difference was because there were many
fewer OT-1 cells in the livers of B6bm1 chimeras than B6B6 or
bm1B6.CD45 (Fig. 3). However, although there was significant
variation in the percentage of TUNEL-positive OT-1 cells between
experiments, the mean percentage of TUNEL-positive cells was elevated
1.6-fold (range, 1.1–2.0-fold) in the B6bm1 chimeras, relative to
the B6B6 chimeras. This increase was statistically significant
(paired t test, n = 4 experiments,
p = 0.04).
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The B6bm1 chimeras allowed us to test the requirement for
hepatocyte Ag presentation in the induction of hepatocyte injury by
CD8+ T cells. Serum aminotransaminases were
elevated in all three groups of chimeras after peptide injection (Table I). The aminotransaminase levels were at least 10 times greater than in
the PBS injected controls, and this elevation was statistically
significant by an unpaired t test (p
= 0.001). The serum aminotransaminases in B6bm1 chimeras after
peptide injection appeared to be twofold lower than in the B6B6 and
bm1B6 chimeras, and despite the wide variation in individual values,
this difference was statistically significant
(p = 0.02). Injection of peptide into chimeras
that had not received OT-1 cells did not result in significant
elevation of aminotransaminases (AST 34 ± 10.5 U/L, ALT 23
± 7.2 U/L). Fig. 5
shows liver histology in chimeras injected with PBS and SIINFEKL
peptide. In all three groups of chimeras, there was a mononuclear
infiltrate around the portal tract, around and central veins, and in
liver lobules. This was associated with the eosinophilic bodies that
are characteristic of hepatocyte apoptosis.
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and 6). There were no TUNEL-positive nuclei in the livers of PBS-injected
mice (Fig. 5, b, f, and j). However,
TUNEL-positive nuclei were evident in the livers of all three groups of
chimeras injected with peptide (Fig. 5, d, h, and
l). The majority of TUNEL-positive nuclei were of small
mononuclear cells, but TUNEL-positive hepatocyte nuclei were also
present (Fig. 6, b, d, and f). Identical TUNEL
staining of kidney tissues from the same chimeras revealed few
TUNEL-positive nuclei (Fig. 7, b, d, and f). This is consistent with published
data showing CD8+ T cell apoptosis in the kidney
(20), but our quantitative analysis of OT-1 T cells shows
that in absolute numbers very few OT-1 cells are undergoing apoptosis
in this organ.
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Discussion |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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It is not possible to test the effect of liver T cell retention on a systemic immune response by removing or bypassing the liver. We therefore chose to determine whether the capacity of the liver to present Ag regulates hepatic trapping and apoptosis of activated CD8+ T cells. If such regulation were present, it would allow us to address whether there were any consequences for the behavior of T cells during a systemic CD8+ T cell response. Our experimental approach to this issue was to construct three sets of radiation bone marrow chimeras, which allows us to determine whether the presentation of the SIINFEKL peptide on bone marrow-derived cells, non-bone marrow-derived cells or both contributes to intrahepatic trapping of CD8+ T cells. The three main cellular components of the liver are the hepatocytes, the sinusoidal endothelial cells, and the bone marrow-derived Kupffer cells. In addition, there are bone marrow-derived intrahepatic lymphocytes, which are unlikely to play a key role in Ag presentation, and dendritic cells, which could play such a role. Previous studies have shown that isolated hepatocytes (28, 29), sinusoidal endothelial cells (30, 31), and Kupffer cells (31, 32) are all competent to present Ag in vitro. We therefore base our interpretation on the extrapolation that all of these cell types can present antigenic peptide in vivo.
The data in Fig. 3 show that Ag presentation by the non-bone
marrow-derived cells of the liver is important in the trapping of
CD8+ T cells, whereas our previous studies
implicate ICAM-1 and by implication its main ligand, LFA-1. In the
intact liver, these two systems are likely to work together. It seems
unlikely that the binding affinity of Ag-specific TCR-mediated
recognition would contribute directly to adhesion between T cells and
either endothelium or hepatocytes. Instead, such interactions would
come into play after an activated T cell has formed an initial adhesion
to endothelium or hepatocytes, most likely through a nonspecific
adhesion molecule such as ICAM-1. The effect of TCR ligation on
LFA-1/ICAM-1 interactions has been studied extensively in vitro.
Stimulation of T cells by receptor cross-linking, or phorbol esters,
increases both the level of LFA-1 expression and the affinity of LFA-1
for ICAM-1 (33, 34). Thus, in activated
CD8+ T cells traversing liver sinusoids, Ag
recognition would be expected to increase the avidity of LFA-1,
promoting firm adhesion and the retention of the T cell in the
liver.
The sinusoidal endothelium resembles inflamed vascular endothelium,
because it constitutively expresses a high level of ICAM-1, but unlike
inflamed vascular endothelium there is no expression of the B7-1 and
B7-2 costimulatory molecules. This high level of expression of ICAM-1
on sinusoidal endothelium may contribute to the apoptosis of
CD8+ T cell seen in the liver. Stimulation of
naive and memory CD8+, but not
CD4+, T cells by anti-TCR Ab and ICAM-1 has
been shown to result in both proliferation and apoptosis of the
responding T cells (35). This contrasts with anti-TCR
and B7-1 stimulation, which results in a similar level of proliferation
but reduced apoptosis. It is of interest that the costimulatory effect
of ICAM-1 preferentially acts on CD8+ T cells
(35, 36, 37). Thus, it is possible that ICAM-1 is the basis of
both intrahepatic CD8+ T cell retention and
CD8+ T cell apoptosis. However, other potential
proapoptotic mechanisms are present in the liver. Our data suggest that
bone marrow-derived Kupffer cells may be important in the induction of
apoptosis, and in addition to ICAM-1 these cells express several
proapoptotic molecules, including membrane bound FasL and soluble
TNF-, whereas sinusoidal endothelium expresses galectin-1
(38, 39, 40, 41). Furthermore, systemic T cell activation promotes
FasL expression in several tissues, including liver (4).
The contribution of these various cell types and proapoptotic molecules
is not yet defined.
Massive liver retention and apoptosis of activated
CD8+ T cells is accompanied by pathology in the
liver. In the original study of Huang et al. (7), we
observed histological evidence of liver damage, and this was
subsequently confirmed in another experimental model by elevation in
serum aminotransaminases (9). In these experiments,
CD8+ T cell activation was induced by the
injection of antigenic peptide in TCR-transgenic mice, and such
peptides may have been presented by hepatocytes so that hepatocytes
would be targets for the cytotoxic effector function of
CD8+ T cells trapped in the liver. Hepatocyte
injury by CD8+ T cells after MHC-restricted
recognition of hepatocyte Ags occurs, and it requires both perforin and
CD95L cytotoxic mechanisms (23). Because activated
CD8+ T cells express FasL and because hepatocytes
can be killed by Fas ligation (24), it has been
hypothesized that T cell activation can result in hepatocyte injury in
the absence of hepatocyte Ag presentation, a mechanism termed
collateral damage (18). In the present study, the
hepatitis induced by systemic CD8+ T cell
activation when hepatocytes are unable to present the SIINFEKL
peptide proves that activation of a small percentage of T cells can
result in such "collateral" liver injury. The relative
contributions of direct and collateral damage are very difficult to
determine from these limited data. The fact that the mean serum ALT was
2-fold lower in the B6bm1 chimeras could be taken to imply that
about one-half of the liver damage was direct, whereas one-half was
collateral. However, individual variation between experimental mice
makes this difference hard to evaluate. Even if the difference proves
to be real in a more extensive series of experiments, it could simply
be because the absolute number of OT-1 cells in the liver is less in
these chimeras. Whatever the relative contributions of the two
mechanism in intact mice, the present data provide clear evidence for a
novel mechanism of collateral liver injury, which may be involved in
some disease states.
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Acknowledgments |
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Footnotes |
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2 Address correspondence and reprint requests to Dr. I. Nicholas Crispe, The David H. Smith Center for Vaccine Biology and Immunology, The Aab Institute for Biomedical Research. University of Rochester, 601 Elmwood Avenue, Rochester, NY 14620. E-mail address: Nick_crispe{at}urmc.rochester.edu
3 Abbreviations used in this paper: FasL, Fas ligand; AST, aspartate aminotransaminase; ALT, alanine aminotransaminase.
Received for publication December 6, 2000. Accepted for publication May 3, 2001.
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References |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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W. Z. Mehal, S. Z. Sheikh, L. Gorelik, and R. A. Flavell TGF-{beta} signaling regulates CD8+ T cell responses to high- and low-affinity TCR interactions Int. Immunol., May 1, 2005; 17(5): 531 - 538. [Abstract] [Full Text] [PDF]
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D. A. Murray and I. N. Crispe TNF-{alpha} Controls Intrahepatic T Cell Apoptosis and Peripheral T Cell Numbers J. Immunol., August 15, 2004; 173(4): 2402 - 2409. [Abstract] [Full Text] [PDF]
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C. A. Wysocki, S. B. Burkett, A. Panoskaltsis-Mortari, S. L. Kirby, A. D. Luster, K. McKinnon, B. R. Blazar, and J. S. Serody Differential Roles for CCR5 Expression on Donor T Cells during Graft-versus-Host Disease Based on Pretransplant Conditioning J. Immunol., July 15, 2004; 173(2): 845 - 854. [Abstract] [Full Text] [PDF]
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) or peptide (
) injections. There was an increase in the OT-1
population after peptide injection with a peak at day 3 and a rapid
decline by day 7 (A and D). A greater
increase was evident in the liver, with a peak at a later time point,
day 5 (B and E). The kinetics in the
kidney were similar to the spleen, but of a lower magnitude
(C and F). The total numbers of
OT-1 cells (106) follow the same kinetics as the
percentages of OT-1 cells. The very large increase in cell number the
liver was due to a 7-fold increase in total liver lymphocyte count. The
total numbers of lymphocytes in the spleen and kidney did not change
significantly after peptide injection.
