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Enhancement of IFN-
Production by T Cells1
Cedars-Sinai Inflammatory Bowel Disease Center, Los Angeles, CA 90048
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Abstract |
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 Top Abstract IntroductionMaterials and MethodsResultsDiscussionReferences |
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in the mucosal inflammation of Crohn’s disease
has been demonstrated by the prolonged clinical responses and/or
remissions among patients receiving i.v. infusion of anti-TNF-.
A correlation between TNF- and elevated IFN-
production is
suggested by the reduction in the number of IFN- producing lamina
propria mononuclear cells (LPMC) found in colonic biopsies from
anti-TNF--treated patients. The aim of this study was to define
the mechanism of TNF--augmented mucosal T cell IFN- production.
In this paper we present evidence that cultured LPMC secrete a factor
which acts on preactivated T cells in concert with TNF- to augment
IFN- production. This activity is independent of IL-12 and IL-18,
the well-documented potentiators of IFN- expression, and is not
produced by PBMC. Peripheral blood PHA-activated T cells incubated in
supernatants from LPMC became responsive to TNF- by increasing
IFN- output upon stimulation. These results are consistent with a
model in which LPMC, but not PBMC, release an unidentified substance
when cultured in vitro with low dose IL-2. This substance can act on
preactivated peripheral T cells, as well as on lamina propria T cells,
conditioning them to respond to TNF- by increased IFN- secretion
upon stimulation. Expression of this factor in the gut mucosa could
contribute to up-regulation of the Th1 response in the presence of
TNF-, and could be important for mucosal
immunoregulation. |
Introduction |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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in the mucosal inflammation of Crohn’s disease has
been demonstrated by prolonged clinical responses and/or remissions
among patients treated with an i.v. infusion of a mAb to TNF-
(1). Several studies using mouse models of colitis have
also indicated a central pathogenic role for TNF- (2).
Especially cogent are those studies showing abrogation of colitis
induction in a mouse TNF- knockout model (3) and the
lethal exacerbation of colitis in mouse models harboring a TNF-
transgene (4). A correlation between TNF- and elevated
IFN- production is suggested by the sharp reduction in the number of
IFN- producing lamina propria
(LP)3 mononuclear
cells (LPMC) found in colonic biopsies from anti-TNF- treated
patients (5). Similar evidence of the correlation between
TNF- and IFN- has been obtained from rodent models of colitis in
which anti-TNF- treatment resulted in decreased production of
IFN- by LPMC (4). Recently, we established that in
vitro culture of LPMC with TNF- for 2 days before stimulation
increases the amount of IFN- produced. In this system, IFN-
production was usually strongly up-regulated in the presence of
nonlymphocyte LPMC, implying that TNF- acts on IFN--producing
cells in synergy with soluble factor(s) from nonlymphocyte LPMC
(5).
Although little is known about TNF- potentiation of IFN-
production, there is evidence that TNF- can act on T cells both
directly and indirectly, via induction of other factors, to augment
IFN- production. In mouse T cells, TNF- can activate the p38
mitogen-activated protein kinase pathway, which mediates IFN-
production (6). TNF- has been reported to stimulate
peripheral blood (PB) mononuclear cells to produce the chemokine IP-10,
and recombinant human IP-10 has been shown to increase IFN-
production by human PB T cells following stimulation by several agents
(7). TNF- can act on human dendritic cells in synergy
with PGE2 to stimulate production of IL-12, which
strongly potentiates IFN- production by T cells (8). In
fact, the cytokines IL-1ß, IL-12 and IL-18 are secreted from
nonlymphocytes and act on T cells, singly and in synergy, to potentiate
IFN- production (9, 10, 11, 12); IL-1ß can be induced by
TNF-, but whether expression of IL-12 or IL-18 is stimulated by
TNF- is not known.
Because some cytokines suppress IFN- production, TNF- might act
to increase IFN- production by suppressing the production of such
inhibitors. Thus, IL-4 suppresses polarization of T cells toward the
Th1, IFN--secreting phenotype, and IL-10 inhibits IFN- expression
by activated T cells (13). In the case of IL-10, however,
TNF- is known to induce, not suppress, production (14, 15).
The mucosal immune system has several characteristic features that
sharply differentiate it from the systemic immune compartment
(16). Most mucosal T cells possess surface markers
associated with prior activation (17), yet they are
relatively unresponsive to stimulation in vitro via the TCR/CD3
complex. However, intestinal mucosal T cells can be strongly activated
via the CD2 pathway, especially with CD28 costimulation
(18). Mice with functional inactivation of diverse genes
(IL-2, IL-10, TCR, G-protein i2) show severe
chronic intestinal inflammation, often the predominant phenotypic
result of the knockout (19). In fact, animal models of
intestinal inflammation are almost completely dependent on the presence
of T cells (19, 20). Such findings, among others, suggest
that T cells within the intestinal mucosa may be poised to produce a
strong inflammatory response. Our previous study documented substantial
augmentation of IFN- production by TNF- in LPMC (5). The purpose
of this study was to define the mechanism by which TNF- augments
mucosal T cell IFN- production. We have found that LPMC cultured in
vitro secrete a factor other than IL-12 or IL-18, which acts on
preactivated T cells to augment IFN- production in the presence of
TNF-. Production of this factor appears to be restricted to the gut
mucosa, and, once identified, its neutralization could be a useful part
of an anti-inflammatory strategy to treat inflammatory bowel
disease (IBD).
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Materials and Methods |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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Intestinal specimens were obtained from patients undergoing surgical resection of the colon (with colon carcinoma or treatment-resistant Crohn’s disease or ulcerative colitis) at Cedars-Sinai Medical Center (Los Angeles). Approval for the use of human subjects was obtained from the Institutional Review Board at Cedars-Sinai Medical Center. In this study, all tissue specimens were taken from an uninvolved area of resected colon from patients with colonic carcinoma (normal), involved areas of patients with ulcerative colitis, as well as from uninvolved and involved areas of patients with Crohn’s disease. LPMC were isolated using a technique modified from that described previously (18). Briefly, the intestinal specimen was washed with HBSS, and the mucosa was dissected away from the underlying layers. The mucosal layer was incubated, in a shaking water bath (37°C, 100 rpm), in calcium- and magnesium-deficient HBSS containing 1 mM EDTA, 50 µg/ml gentamicin, 100 U/ml penicillin, 100 µg/ml streptomycin, and 50 µg/ml Fungizone, with the solution changed every 30 min until the supernatant was free of epithelial cells. The remaining LP was minced into 1- to 2-mm pieces and digested for 10 min in RPMI 1640 containing 10% FCS, 0.5 mg/ml collagenase B (Boehringer Mannheim, Indianapolis, IN), 1 mg/ml hyaluronidase (Sigma, St. Louis, MO), 0.1 mg/ml DNase I (Sigma), 50 µg/ml gentamicin, 100 U/ml penicillin, 100 µg/ml streptomycin, and 50 µg/ml Fungizone in the water bath (100 rpm). The supernatant was collected, filtered through 110-µm nylon mesh (Spectrum Laboratory Products, Houston, TX), and centrifuged at 500 x g for 5 min. The cell pellet was resuspended in 15 ml and centrifuged at 30 x g for 5 min to remove epithelial and other large cells. The supernatant was removed and lymphocytes were isolated by separation on Ficoll-Hypaque gradients. The cells were then washed three times with HBSS and resuspended in RPMI 1640 containing 10% FCS. PBMC were isolated from normal healthy volunteers by separation on Ficoll-Hypaque gradients. Resting T cells were enriched using immunomagnetic beads (Miltenyi Biotech, Auburn, CA) and anti-CD2 Ab (T11, Coulter Immunology, Hialeah, FL).
Induction of LP-like T cells
Mononuclear cells from Ficoll-Hypaque gradients were incubated in medium on nylon wool at 37°C for 2 h to enrich for T cells, then were cultured in RPMI 1640 with 10% FCS with a 1:5 ratio Daudi:lymphocytes plus 10 U/ml rIL-2 for 5 days (18). Daudi were irradiated with 3000 rad and washed three times in HBSS before addition to cultures. Following a 5-day culture, LP-like cells were washed with HBSS to remove dead Daudi cells (at this point, there were virtually no live Daudi cells in the culture as determined by flow cytometry).
Cell culture conditions
LPMC and PBMC were cultured (1 x
106/ml) for 2 days in RPMI 1640 with 10%
heat-inactivated FBS, 25 mM HEPES, 2 mM L-glutamine and 50
µg/ml gentamicin. A total of 10 U/ml rIL-2 (R&D Systems, Minneapolis,
MN) was added to LPMC and PBT/Daudi cultures to maintain viability
(18), and 1 µg/ml PHA (Sigma, St. Louis, MO) was added
to PBMC for experiments using PHA-activated PB T cells. Most
experiments followed a standard protocol: cells were incubated in cell
culture plates for 2 days, with and without TNF-, both conditions
including any blocking or control Abs or added cytokines. In indicated
experiments, adherent PBMC were prestimulated with PMA (10 ng/ml) plus
ionomycin (1 µg/ml), or with LPS (2 µg/ml, all from Sigma), or with
1,25-dihydroxyvitamin D3 (10 nM, Biomol, Plymouth
Meeting, PA) for 2–20 h before washing and addition of T cells for the
2-day incubation. After incubation, cells were collected, washed with
medium, and resuspended in medium containing the stimulatory monoclonal
anti-CD2 pair (GD10 and CB6, gift of Dr. C. D. Benjamin,
Biogen, Cambridge, MA; 0.2 µg/ml each). Anti-CD3 cross-linking was
delivered by OKT3 mAb incubation in a culture well coated with goat
anti-mouse IgG (Jackson ImmunoResearch, West Grove, PA). After
18 h, supernatants were frozen for later analysis by ELISA.
Neutralizing Abs
Except for anti-TNF-, Abs were added to both control and
TNF- incubation conditions at 2–10 µg/ml as stated in figure
legends. Appropriate species IgG (mouse and goat, Jackson
ImmunoResearch; rabbit, Peprotech, Rocky Hill, NJ) at the same
concentrations were used for controls. Goat anti-IL-12 IgG was the
gift of Dr. M. Gately (Hoffmann-LaRoche, Nutley, NJ), who demonstrated
that at 0.5 µg/ml it neutralized the bioactivity of IL-12 at 200
pg/ml (21). Commercial Ab were as follows: anti-IL-18
mAb (MAB318, R&D Systems). The data sheet provided shows a
neutralization curve in which 50% of the bioactivity of recombinant
human IL-18 at 50 ng/ml or 2.7 nM was neutralized by the monoclonal at
0.4–0.8 µg/ml. (We used the Ab at 10 µg/ml), anti-IL-4 IgG
(Peprotech), anti-IL-10 IgG (mAb JES3-9D7, Biosource International,
Camarillo, CA), anti-TNF- IgG (Genzyme, Cambridge, MA; and R&D
Systems), all of which have been shown to be neutralizing by the
vendors.
Cytokines
Recombinant cytokines were used at the following final
concentrations: 10 U/ml IL-2, 20–100 pM IL-12, 200 U/ml IFN-, and
20 ng/ml TNF- (all from R&D Systems) and 5 nM IL-18 (Peprotech).
IFN- ELISA
IFN- was measured by an amplified ELISA assay
(18). Dynatech (Chantilly, VA) Immulon 3 microtiter plates
were coated overnight with 100 µl of 2.5 µg/ml monoclonal
anti-IFN- (PharMingen, San Diego, CA). Samples and standards
were added for 24 h, followed by washing and addition of 100 µl
of 2.5 µg/ml polyclonal rabbit anti-IFN- (Endogen, Woburn, MA)
for 2 h. This was followed after washing by addition of 100 µl
of 1:1000 diluted mouse anti-rabbit alkaline phosphatase-conjugated
Ab (Jackson ImmunoResearch) for 2 h. Substrate, 0.2 mM NADP
(Sigma), was added for 30 min followed by addition of amplifier (3%
2-propanol, 1 mM iodonitrotetrazolium violet, 75 µg/ml alcohol
dehydrogenase, and 50 µg/ml diaphorase; Sigma) for 30 min. Plates
were read at 490 nm using an Emax plate reader (Molecular Devices,
Menlo Park, CA).
The TNF- ELISA used capture and detection Abs from R&D Systems and
enhanced development as for IFN-. For the IL-12 ELISA, we used the
Quantikine kit from R&D Systems which can detect less than 0.5 pg/ml of
hIL-12.
Data analysis
Data acquisition and reduction were performed using the ELISA Master program for Macintosh computers, developed by R. L. Deem. Statistical analysis was done by Statview using the paired t test or the nonparametric Wilcoxon signed rank test in cases where individual experiments had very large differences in measured values.
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Results |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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potentiates IFN- production by
LPMC
We have previously reported that LPMC from normal colon incubated
with TNF- in vitro for 2 or more days secrete higher levels of
IFN- upon stimulation with anti-CD2 Abs compared with controls
incubated without added TNF- (5). In the present study,
we have confirmed and extended this finding. Incubation of LPMC from
inflamed as well as from normal colon with TNF- increases subsequent
IFN- production upon stimulation with anti-CD2 (see
Figs. 1–4
),
but we have seen no consistent difference in augmentation of IFN-
secretion when comparing LPMC from inflamed and normal tissue (data not
shown). We have found that TNF- is spontaneously generated in
control cultures during incubation (8/15 incubation supernatants had
90–1620 pg/ml TNF-, where 90 pg was the lower limit of detection).
Thus, we might have underestimated the role that TNF- plays in LPMC
IFN- production, because spontaneous release of TNF- during in
vitro culture of LPMC might increase the "control" level of IFN-
production. To test this possibility, LPMC were incubated with
neutralizing anti-TNF- during the 2 days of culture before
stimulating with Abs to CD2. Results in Fig. 1 show that incubation of LPMC cultures
with anti-TNF- Ab decreased the secretion of IFN- upon
stimulation. Thus, if endogenously produced as well as added TNF-
were taken into account, the magnitude of the effect of TNF- on
IFN- secretion was considerably greater than we had previously
found. This anti-TNF- Ab did not induce apoptosis of T cells, as
determined by Annexin V staining and flow cytometry (data not shown).
Therefore, the decrease in IFN- production was due to the
neutralization of TNF-.
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potentiation of LPMC IFN- production is independent of
cytokines known to modulate IFN- secretion
Because TNF- is spontaneously produced in LPMC cultures, it is
possible that other cytokines that can increase IFN- production are
secreted during the 2-day incubation as well. Further, because the
augmentation of IFN- requires prolonged incubation with TNF-,
changes in cytokine levels in the cultures during incubation may
underlie the effect of TNF-. Therefore, we sought to determine
whether IFN- potentiation by TNF- is mediated via other cytokines
known to affect IFN- expression.
The TNF- effect is independent of IL-12 and IL-18.
Augmentation of mucosal Th1 cytokine production appears to require the
presence of non-T, non-B cells (mostly macrophages) and at least 2 days
of culture in the presence of TNF-, suggesting that a soluble factor
from nonlymphocytes is generated over this 2-day period. To begin to
identify such a soluble factor, we performed experiments to determine
whether cytokines known to regulate generation or function of
IFN--producing Th1 cells were responsible for the TNF- effect.
IL-12 is a macrophage-derived cytokine that is known to be required for
generation of Th1 responses (10). IL-18 is a recently
discovered and cloned cytokine also derived from macrophages that
alone, but especially in conjunction with IL-12 augments Th1 function
and IFN- production (11, 12). IL-12 and TNF-
synergize to induce IFN- production from splenocytes
(13). Therefore, to determine whether the augmentation of
IFN- production by TNF- is dependent upon enhancement of IL-12 or
IL-18 production and/or interactions with IL-12 present in the in vitro
LPMC culture, we added blocking Abs to both control and TNF-
conditions for the two day cultures. Fig. 2
demonstrates that addition of
anti-IL-12 or anti-IL-18 blocking Abs had no effect on the
TNF- mediated augmentation of IFN- production. Furthermore, if
TNF- acted by inducing IL-12 or IL-18, addition of high levels of
IL-12 and/or IL-18 should eliminate the TNF- effect. When high
levels of recombinant IL-12 or IL-18 were added to the incubations, as
expected, IFN- production was elevated by the added cytokines in
both control and TNF- conditions, but IFN- production was
consistently greater than control in the TNF- condition (Fig. 3). Thus, the addition of either IL-12 or
IL-18 did not abrogate the enhancing effect of TNF-. We measured
IL-12 in LPMC supernatants from six surgical resection samples
incubated with or without TNF-. Values ranged from 0 to 3.5 pg/ml
with most below 0.6 pg/ml, near the detection limit, and did not
correlate with the addition of TNF-. Therefore, taking into account
the results from neutralizing Ab and cytokine addition experiments,
together with results of ELISA, TNF- enhances IFN- production
independently of any effect on IL-12 or IL-18.
TNF- does not act by influencing IL-4 or IL-10
production.
IL-4 and IL-10 are readily produced by LPMC and are prominent
inhibitors of Th1-type cytokines, particularly production of IFN-
(22). These cytokines, however, have been reported to
potentiate IFN- production, as well, in established Th1 cells
(15). The effect of TNF- on IFN- production might be
due to an effect of TNF- on either IL-4 or IL-10. We have shown
previously (5) that no IL-4 or IL-10 is detectable in
incubation supernatants when tested by ELISA. However, to determine
whether either factor could play a role at very low levels,
neutralizing Ab was added to both control and TNF- incubations. The
presence of neutralizing Ab, while having a small suppression in the
case of IL-10, did not block the potentiation of IFN- production by
TNF- (Fig. 4).
Potentiation of IFN- by TNF- does not occur in PBMC
We have demonstrated that incubation of freshly isolated PBMC with
TNF- for 2–5 days did not result in increased IFN- production,
in contrast to the case of LPMC (5). This difference could
relate to the fact that LPMC are a population of highly activated
cells, T and non-T, as compared with the relatively resting state of
PBMC. Thus, to determine whether the effects of TNF- were unique to
the mucosal compartment, PBMC were cultured for 2 days with low-dose
PHA, with or without TNF-, then activated with anti-CD2 Abs;
although high levels of IFN- were produced, no potentiation of
IFN- by TNF- occurred (Fig. 5A). However, the effect could
depend on the activation state of either the T cells or the cells
adherent to plastic. Incubations were performed using preactivated T
cells, with or without adherent cells, which were themselves untreated
or pre-activated. T cells preactivated by coculture with Daudi
("LP-like" T cells; Ref. 18) were cultured with or
without TNF- by themselves or after remixing with plastic adherent
cells preactivated by exposure to IFN-, LPS, or 1,25-dihydroxy
vitamin D3. Cells were incubated for 2 days,
washed, and stimulated via CD2, the same stimulus used for LP-derived
T cells. Although cells cocultured with adherent cells could
produce large amounts of IFN-, no consistent potentiation of IFN-
by TNF- was observed (Fig. 5, B--E). Thus, the
TNF- potentiation of IFN- production, which is so prominent in
LPMC from the intestinal mucosa, is not shared by PBMC, either resting
or preactivated by several modalities commonly used in in vitro
studies.
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with increased IFN-
production
The fact that TNF- does not augment IFN- production in PBMC
suggests that the potentiation in LPMC may be due either to the unique
activation state of LP T cells or to production of a cofactor generated
only in LPMC cultures. To test whether PHA-activated PB T cells could
respond to a LPMC-generated cofactor, PHA-activated PB T cells were
incubated with supernatants from LPMC that had shown a substantial
increase in IFN- production when incubated with TNF-.
PHA-activated PB T cells incubated with supernatants from LPMC cultured
with TNF- did produce more IFN- than those incubated with the
supernatants cultured without TNF- (Fig. 6). Possible explanations for this result
include: 1) TNF- could be necessary to induce LPMC to secrete a
factor into the TNF- supernatants which then acts alone, or in
concert with TNF-, on the PHA-activated PB T cells; or 2) LPMC may
spontaneously elaborate a factor necessary to sensitize PHA-activated
PB T cells to TNF-. TNF- incubated supernatants doubtless
contained residual exogenously added TNF-, but control supernatants
contained only the (presumably) much lower level of endogenous TNF-
produced during incubation (Fig. 1). To test whether a cofactor was
spontaneously produced by LPMC, a high level of TNF- was added to
control supernatants in which PHA-activated PB T cells were incubated.
A lack of effect of addition of TNF- would support the first
hypothesis above, whereas potentiation of IFN- production would
support the second. To further support the second hypothesis, when
TNF- was added to cells incubated in the control supernatants, their
IFN- production was potentiated (Fig. 7, unheated supernatants) and was
equivalent to that of cells cultured with the TNF- supernatant (data
not shown). Thus, while TNF- alone did not influence IFN-
production by PB-PHA cells, TNF- in the presence of supernatant from
LPMC did. To begin to define the subpopulations that produce this
factor, LPMC were depleted of T cells (>94% T cells removed) before
incubating LPMC for 2 days. In seven experiments, supernatants from
four non-T LPMC incubations were tested on PB-PHA. TNF- added to
these supernatants increased IFN- produced in five experiments (from
15 to 125%), with three of the four supernatants. We conclude that
PB-PHA cells, exposed to a factor released by cultured non-T LPMC, are
then able, like LP T cells, to respond to prolonged exposure to TNF-
by expressing more IFN- upon subsequent stimulation by Abs to CD2.
Because we measured low levels of TNF- in most LPMC cultures, the
question of whether or not generation of the factor depends on TNF-
is still open.
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production, but the
differential augmentation of IFN- in the TNF- condition is
maintained
From our results it is apparent that TNF- augments IFN-
production from T cells independently of cytokines known to be involved
in regulation of IFN- production. To begin to identify the nature of
the cofactor involved, LPMC supernatants were heated to 75°C for 40
min, treatment known to eliminate the functions of most, but not all,
bioactive proteins (23, 24). Heated supernatants were then
tested on PB-PHA cells with and without TNF-, as before (Fig. 7).
Although IFN- production was reduced, in four of four experiments,
some TNF- augmentation of IFN- production still occurred. These
results support the hypothesis that at least part of the IFN-
augmenting TNF- cofactor activity is mediated by a heat-stable
soluble factor, which could be an unusually stable protein, a peptide,
or other organic molecule.
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Discussion |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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Ab in treating
Crohn’s disease resulted in marked prolonged amelioration of symptoms
in most patients, and focused attention on TNF- as a key factor
sustaining inflammation. In experiments designed to examine this effect
of TNF-, we have identified a pro-inflammatory action of TNF- in
LPMC cultured in vitro: potentiation of IFN- production. Here we
present evidence that the magnitude of the effect is larger than
previously reported (5) due to endogenous production of
low levels of TNF- in control incubations, which significantly
enhanced IFN- production. In addition, this potentiation of IFN-
by TNF- in LPMC was independent of IL-12 and IL-18, the most
well-documented potentiators of IFN- expression. Further, the effect
of TNF- could not be reproduced in PBMC, whether resting or
preactivated by any of several stimuli, using multiple donors. However,
PHA-activated PB-T cells incubated in supernatants from LPMC became
responsive to TNF- by increasing IFN- output upon stimulation.
Heating of supernatants did not abolish the activity, although the
activity of TNF- itself was abolished by this treatment in control
experiments (data not shown). These results are consistent with a model
in which LPMC, but not PBMC, release an unidentified factor when
cultured in vitro with low dose IL-2 for 2 days. In the presence of
this factor, preactivated peripheral T cells, as well as LP T cells,
respond to incubation with TNF- by increased IFN- output upon
stimulation.
Expression of such a factor specifically in the mucosa may account for
an increased pro-inflammatory effect of TNF- via augmentation of
IFN- production in the gut. In light of the normalization of IFN-
production by stimulated LPMC from anti-TNF--treated patients,
the presence of such a protein in the gut mucosa may explain, at least
in part, the strong anti-inflammatory action of TNF- blockade
(5). If expression of the factor is largely specific to
the mucosa, as our data suggest, its role and even its identity may not
have been evident from studies on cells from PB. Indeed, the lack of
effect of TNF- on IFN- production in preactivated PBMC implies
that factors known to be produced by activated PBMC are probably not
responsible for the effect seen in LPMC. These factors include the
chemokine IP-10, expected to be produced by activated PBMC
(7), and for which a high percentage of PB T cells possess
receptors (25). However, because preactivated PB cells can
respond to the factor, a convenient assay is at hand to aid in its
identification.
Accumulating evidence points to a primary role for TNF- in the
pathogenesis of Crohn’s disease as well as in the maintenance of
inflammation in Crohn’s disease patients (1, 26, 27, 28), and
this role for TNF- is supported by findings in several animal models
of intestinal inflammation (2, 3, 4, 20). A strong case can
now be made that TNF- acts by shifting the mucosal immune response
away from production of anti-inflammatory factors such as IL-10 and
TGF-ß (29) and toward production of pro-inflammatory
factors, IFN- (as we have documented) and chemokines, such as RANTES
(30). Interestingly, several studies have demonstrated
that shorter-term exposure to TNF- can enhance T cell responses
(Ref. 31 and references therein). Direct effect of TNF-
on T cell activation has been shown by studies in which TNF-
enhanced the expression of CD69 (32) or CD70 (CD27 ligand)
(33) during acute activation. Furthermore, it has been
shown that TNF- can enhance T cell cytotoxic activity by inhibiting
TGF-ß, a cytokine known to inhibit cytotoxic function (34, 35). The results of these studies suggested the possibility that
TNF- augments Th1 cytokine secretion and IFN- production by
modulating accessory populations, or by selectively down-regulating
inhibitory cytokine(s). The present study demonstrates a direct effect
of TNF- on IFN- production by cells stimulated via CD2, a
predominantly T cell stimulatory receptor. However, this direct effect
required another factor, produced by LPMC during culture. This factor
alone does not promote increased IFN- production, because LPMC
supernatants containing anti-TNF- Ab added to PB-PHA did not
increase IFN- over control (data not shown). But because the strong
stimulation of the Th1 arm (as measured by IFN- production) of
regulatory T cells by TNF- requires the factor, its identification
and further study will provide further insight into an important factor
involved in mucosal immunoregulation.
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Acknowledgments |
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and 6. |
Footnotes |
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2 Address correspondence and reprint requests to Dr. Stephan R. Targan, Inflammatory Bowel Disease Research Center, Cedars-Sinai Medical Center, 8700 Beverly Boulevard, D4063, Los Angeles, CA 90048. E-mail address:
3 Abbreviations used in this paper: LP, lamina propria; LPMC, LP mononuclear cells; IBD, inflammatory bowel disease; PB, peripheral blood.
Received for publication December 22, 1998. Accepted for publication August 9, 1999.
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responsive elements. J. Biol. Chem. 270:21545.This article has been cited by other articles:
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K. A. Papadakis, J. L. Prehn, C. Landers, Q. Han, X. Luo, S. C. Cha, P. Wei, and S. R. Targan TL1A Synergizes with IL-12 and IL-18 to Enhance IFN-{gamma} Production in Human T Cells and NK Cells J. Immunol., June 1, 2004; 172(11): 7002 - 7007. [Abstract] [Full Text] [PDF]
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 A. Q. Khan, Y. Shen, Z.-Q. Wu, T. A. Wynn, and C. M. Snapper Endogenous Pro- and Anti-Inflammatory Cytokines Differentially Regulate an In Vivo Humoral Response to Streptococcus pneumoniae Infect. Immun., February 1, 2002; 70(2): 749 - 761. [Abstract] [Full Text] [PDF]
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N.-J. Chen, M.-W. Huang, and S.-L. Hsieh Enhanced Secretion of IFN-{{gamma}} by Activated Th1 Cells Occurs Via Reverse Signaling Through TNF-Related Activation-Induced Cytokine J. Immunol., January 1, 2001; 166(1): 270 - 276. [Abstract] [Full Text] [PDF]
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