By
From the Laboratory of Immunology, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Maryland 20892
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Abstract |
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Cells of the innate immune system secrete cytokines early in immune responses that guide maturing T helper (Th) cells along appropriate lineages. This study investigates the role of cytokine networks, bridging the innate and acquired immune systems, in the pathogenesis of an organ specific autoimmune disease. Experimental allergic encephalomyelitis (EAE), a demyelinating
disease of the central nervous system, is widely used as an animal model for multiple sclerosis.
We demonstrate that interleukin (IL)-12 is essential for the generation of the autoreactive Th1
cells that induce EAE, both in the presence and absence of interferon . The disease-promoting effects of IL-12 are antagonized by IL-10 produced by an antigen nonspecific CD4+ T cell
which, in turn, is regulated by the endogenous production of IL-12. This unique immunoregulatory circuit appears to play a critical role in controlling Th cell differentiation and provides a
mechanism by which microbial triggers of the innate immune system can modulate autoimmune disease.
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Introduction |
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Tcells are classified into Th1/Th2 subsets based on the array of cytokines they produce upon antigen stimulation, which, in turn, dictates the nature of the immune response (1). Studies in infectious disease and autoimmunity models have demonstrated that immune responses to both self- and foreign antigenic challenges are frequently dominated by induction of a particular Th subset, with profound consequences for clinical outcome (2, 3). Although the inflammatory effector function of Th1 cells is essential for the clearance of intracellular pathogens, it is also responsible for tissue damage typical of organ-specific autoimmunity (4- 13). Th2 cells, on the other hand, are critical for the clearance of many helminthic infections and have been implicated in the pathogenesis of systemic autoimmune diseases driven by the production of autoreactive antibodies (14, 15). However, they are generally depicted as suppressor cells or ineffectual bystanders in organ-specific autoimmune diseases (16).
Experimental allergic encephalomyelitis (EAE)1 is a demyelinating disease of the central nervous system (CNS)
induced either by active immunization with myelin proteins or by the adoptive transfer of myelin protein-reactive
T cells. The CD4+ T cell lines and clones that transfer EAE
invariably produce IFN- and/or TNF-
/lymphotoxin-
(LT
) on antigenic challenge in vitro (21). Lines and
clones with the same peptide/MHC specificities that have
been manipulated to produce Th2 rather than Th1 cytokines generally lose their encephalitogenic potential and, in
certain circumstances, can act to suppress disease (24).
Furthermore, mRNA encoding Th1 cytokines is found in
the CNS during peak disease with levels falling at the time
of remission (29). Interventions to block the activity of
the Th1 cytokine, TNF-
, by use of neutralizing antibodies (32, 33), soluble TNF I receptors (34, 35), or type 1 phosphodiesterase inhibitors (36, 37), lead to reversal of
EAE, whereas injection of recombinant TNF-
triggers relapses (38). Conversely, EAE has been treated successfully
by the administration of exogenous Th2 cytokines either
infused systemically or delivered locally by genetically engineered myelin-reactive cells (39). Endogenous production of IL-4 and IL-10, which counterregulate and antagonize Th1 cytokines, has been implicated in the initiation of
spontaneous remissions (29).
The prevailing dogma, shaped by such observations, depicts the inducers of EAE as polarized Th1 cells and causally links their Th1 phenotype with encephalitogenicity
(4). However, the regulation and function of individual cytokines in EAE is more complex than the above studies imply. Alteration in the systemic expression and/or activity of
cytokines believed to be important in the pathogenesis of
EAE has yielded paradoxical results with respect to clinical outcome. For example, the injection of neutralizing antibodies to IFN- exacerbated disease in five independent
studies (43) and administration of recombinant IFN-
has repeatedly been found to have a protective effect (43,
47, 48). EAE has also been successfully induced in IFN-
knockout (
/
) and IFN-
-receptor knockout mice, and in
at least two cases the disease was more severe in the knockouts than in wild-type counterparts (49). Taken together,
these results suggest that IFN-
can actually act to suppress
the development of EAE, either during the evolution of
encephalitogenic effectors or at a downstream event critical to the formation of demyelinating plaques. The application
of the Th1/Th2 paradigm to EAE was most recently
brought into question by the successful induction of the
disease in double knockout mice deficient in expression of
the other two Th1 cytokines, TNF-
and LT
(52), and by
the failure to induce a more severe form of EAE in IL-4
/
mice in comparison to their wild-type counterparts (53).
To further complicate matters, a recent study demonstrated that T cells that express a TCR transgene specific for myelin basic protein (MBP) induce EAE in immunodeficient
recipients after culture under either Th1 or Th2 polarizing
conditions (54). This study, as well as one illustrating a similar phenomenon in an animal model of diabetes (55), suggests that T cells that do not fall into the classic Th1 subset
can nevertheless act as mediators of organ-specific autoimmunity.
Thus far, most studies have been designed to investigate the role of highly differentiated, polarized Th1/Th2 effector cells and their signature cytokines in EAE. An alternative strategy is to examine autoreactive cells during formative stages in their development in order to gain insight into the factors that promote the evolution of naive autoreactive precursors into proinflammatory autoimmune effectors. Among the factors currently known to influence patterns of Th cell development, cytokines produced by cells of the innate immune system are the most important. The production of IL-12 by monocytes and dendritic cells results in Th1 differentiation, whereas the production of IL-10 by macrophages and a subset of B cells antagonizes the activities of IL-12 (56). Hence, cells populating the innate component of the immune system, stimulated by conserved microbial products and structural elements, can secrete cytokines early in immune responses that can potentially guide maturing Th cells along appropriate lineages (60, 61). In autoimmune disease, this interplay between the innate and adaptive responses may be subverted to promote the development of autoreactive effectors. We have previously demonstrated that MBP-reactive cells can be converted from a quiescent state into autoimmune effectors by exposure to exogenous IL-12 or to microbial products that induce IL-12 production by macrophages (62, 63). Other studies have also demonstrated a disease-promoting effect of IL-12 in EAE as well as other models of organ-specific autoimmunity (7, 64, 65). Conversely, anti-IL-12 was found to delay the onset of disease when administered short term to recipients of primed Th1 EAE effectors, but severe disease ensued immediately after withdrawal of therapy. More prolonged administration of the neutralizing antiserum resulted in protective effects that persisted after cessation of treatment, but the treated animals were not followed for a long enough period to determine whether they would experience relapses. No effort was made in these studies to analyze the role of IL-12 in the induction of the Th1 effectors and the authors postulated that the protective effects of anti-IL-12 at the late stage of disease pathogenesis may have involved an interference with the ability of the mature autoreactive cells to home to the CNS (64).
This study focuses on the contribution of cytokine production by cells of the innate immune system to the generation and differentiation of Th1 autoreactive effector cells. Initially, we use cytokine-deficient mice to demonstrate that
IL-12 is absolutely essential for the pathogenesis of EAE,
both in the presence and absence of IFN-. In a parallel
approach involving the neutralization of IL-12 in cytokine-sufficient mice, we characterize a unique immunoregulatory circuit in which endogenous production of IL-12 suppresses IL-10 production by an antigen nonspecific CD4+ T
cell. This latter cell, which may represent a new member of the innate immune system, appears to play a critical role in
regulating Th cell differentiation.
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Materials and Methods |
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Mice.
SJL/J, C57BL/6, C57BL/10, BALB/c, BALB/c nu/ nu, and C.B-17 SCID mice were all obtained from the National Cancer Institute (Frederick, MD). Breeding pairs of C57BL/6 IL-12Induction of EAE.
Bovine MBP was obtained from Sigma Chemical Co. (St. Louis, MO). MBP87-106, corresponding to residues 87-106 of murine MBP (VVHFFKNIVTPRTPPPQGK), was synthesized by the Laboratory of Molecular Structure, Peptide Synthesis Laboratory (NIAID, NIH, Bethesda, MD) and analyzed and purified by high pressure liquid chromatography. For induction of EAE by active immunization, mice were injected with bovine MBP (400 µg) emulsified in CFA (1:1) by the subcutaneous route over the left flank on day 0 and over the right flank on day 7. Mice were examined daily and rated for severity of neurological impairment as previously described (62, 63). For disease induction by adoptive transfer, donor mice were immunized with MBP87-106 (100 µg) or bovine MBP (400 µg) emulsified in CFA (1:1) by subcutaneous injection at four sites over the flanks. 10-14 d later, draining LN cells (axillary and inguinal) were removed and processed as previously described (62). Cells were cultured for 96 h with MBP87-106 (50 µg/ml) or bovine MBP (50 µg/ml) in RPMI 1640 containing 10% FCS and standard supplements (TCM). Recovered cells (5 × 107) were injected intraperitoneally into naive syngeneic recipients that were examined daily for signs of EAE. In certain studies, the mice were injected intraperitoneally with polyclonal goat anti-mouse IL-12 (0.5 µg per injection; a gift of Drs. D. Presky and M. Gately, Hoffman-La Roche), normal goat IgG (0.5 mg per injection; Sigma Chemical Co.), rat anti-mouse IL-10 (1 mg each of mAbs SXC-1 and SXC-2 per injection; reference 66), or control rat IgG (2 mg; Sigma Chemical Co.).Cell Cultures.
Single-cell suspensions of spleen or LN tissue were prepared by passage through wire mesh and red blood cells lysed with ACK buffer (NIH Media Unit, Bethesda, MD). For detection of cytokine production during primary culture, cells (4 × 106/ml) were cultured in TCM in 24-well plates (Costar Corp., Cambridge, MA) for 96-144 h. In some experiments, spleens were depleted of various subpopulations before culture using sheep antifluorescein biomag beads (Perspective Bioresearch Products, Cambridge, MA) together with FITC-conjugated mAbs specific for CD4, CD8, B220, and Mac-1 (PharMingen, San Diego, CA). In other experiments, purified CD4+ T cells and B220+ B cells were cocultured (2 × 105 of each in 200 µl) in 96-well microtiter plates (Costar Corp.). The CD4+ T cells were isolated using CD4 subset enrichment columns (R & D Systems, Inc., Minneapolis, MN) with 90-95% purity achieved. B cells were positively selected to 93-98% purity by incubation with anti-B220 microbeads followed by magnetic separation using VS+ columns (Miltenyi Biotec, Inc., Auburn, CA). For measurement of cytokine production by MBP-reactive LN cells during secondary cultures, the cells were harvested and washed extensively after 96 h of primary in vitro stimulation. Then they were resuspended in fresh media (1 × 106 cells/ml) and supplemented with irradiated syngeneic splenocytes (3,000 rads; 4 × 106 cells/ml) with or without MBP87-106 (50 µg/ml). Supernatants were assayed 48 h later.Cytokine ELISA.
IFN- ![]() |
Results |
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Previous studies have attempted to address the contribution of individual
cytokines to the pathogenesis of EAE by examining the
susceptibility of cytokine-deficient mice to disease induction. Unfortunately, a comparison of these studies was difficult as different myelin-derived antigens were used and
the genetically deficient mice were only backcrossed onto
"susceptible" backgrounds for two to four generations. To
avoid these problems, we used a single antigen to examine
the susceptibility of wild-type mice and several different
cytokine-deficient mice that had been backcrossed more
completely to either the C57BL/6 or the C57BL/10 backgrounds. C57BL/6 wild-type mice were relatively resistant
to EAE, with only 31% of actively immunized individuals
exhibiting clinical symptoms (Fig. 1). Among the affected
cohort the mean peak clinical score was 2.67 ± 0.8 and the
average day of onset was 8.2 ± 0.4. On the other hand, IL-12/
mice were completely resistant to disease whereas
IFN-
/
animals exhibited severe EAE at 100% incidence (mean peak clinical score, 4.5 ± 0.68; average day of onset, 9.1 ± 1.6). Surprisingly, the induction of EAE in the
IFN-
/
animals was completely prevented by the
administration of neutralizing antibody to IL-12, but not
isotype control antibody, during the course of active immunization (Fig. 1, top). This result suggested that IL-12
promotes the development of EAE by an IFN-
-independent mechanism. The failure of IL-12
/
and anti-IL-12-treated IFN-
/
mice to develop disease was not
secondary to the induction of a Th2 response as neither IL-4
nor IL-10 was detected after stimulation of LN cells from
these animals with MBP in vitro (data not shown). Interestingly, the incidence and severity of disease in IL-4
/
mice was comparable to that of the C57BL/6 wild-type
controls (Fig. 1, bottom; mean peak clinical score, 1.95 ± 0.7) with slightly delayed onset (13 d ± 3). By contrast IL-10
/
mice exhibited enhanced disease incidence and severity in comparison to C57B/10 wild-type controls (Fig.
1, bottom; mean peak clinical scores, 2.4 ± 0.6 vs. 1 ± 0.1 in IL-10
/
and wild-type mice, respectively) with similar kinetics (average day of onset, 11.5 ± 0.5 and 11 ± 1.3, respectively). LN cells from both the IL-4
/
and IL-10
/
strains produced elevated levels of IFN-
after
stimulation with MBP in vitro compared to LN cells from
wild-type mice (data not shown). A deficiency of MBP-specific Th2 cells could, theoretically, be responsible for the
increased incidence of EAE, as well as the propensity for
Th1 cytokine production, in IL-10
/
mice. However,
this explanation is unlikely since the IL-4
/
mice did not
differ significantly from controls with regard to those characteristics and IL-4 is required for Th2 development (67).
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To define IL-12-dependent stage(s)
in the development of EAE effectors, we neutralized IL-12
during individual steps of an adoptive transfer protocol.
Susceptible female SJL mice were immunized once with
MBP87-106 peptide (the immunodominant epitope for H-2s
strains) in CFA; 10 d later, draining LN cells were stimulated with antigen in vitro for 4 d and then transferred into
naive syngeneic recipients. As both the in vivo immunization and the in vitro boost are required for the generation
of encephalitogenic effectors, we treated mice with a polyclonal anti-IL-12 antiserum during the course of priming
and/or added the antiserum to the in vitro cultures. MBP-reactive LN cells exposed to anti-IL-12 only during culture
were mildly inhibited in their ability to produce IFN- on
subsequent antigenic challenge in vitro (Fig. 2 A), but were
unimpaired in their capacity to transfer disease (Fig. 2 B).
In contrast, LN cells from donor mice treated with anti-IL-12 in vivo exclusively were severely compromised in their ability to produce antigen-dependent IFN-
on secondary
challenge in vitro (Fig. 2 A) and their encephalitogenicity
was markedly decreased on adoptive transfer (Fig. 2 B). Administration of anti-IL-12 both in vivo and in vitro abrogated IFN-
production in secondary cultures, but did not
further impair the ability of cells to transfer disease when
compared to cells from animals that were only exposed to
anti-IL-12 in vivo (Fig. 2, A and B).
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To investigate whether the MBP-reactive cells from the susceptible SJL strain had assumed a Th2 phenotype after treatment with anti-IL-12 in vivo and/or in vitro, we measured IL-4 in the supernatants of the primary and secondary cultures of the four experimental groups studied in Fig. 2. We were unable to detect IL-4 in any of the supernatants tested. On the other hand, both LN cells and splenocytes from anti-IL-12 treated donors, but not control IgG-treated donors, produced significant quantities of IL-10 during the primary culture (Fig. 3 A). This result raised the question of whether the suppression of the encephalitogenic phenotype by anti-IL-12 treatment was secondary, in part, to the production of IL-10 during the activation of MBP-specific T cells. Indeed, neutralization of IL-10 using a mixture of mAbs during both immunization and the in vitro culture largely reversed the protection from disease afforded by the treatment of donor mice with anti-IL-12 (Fig. 3 B).
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Initially, we postulated that the source of IL-10 was an MBP-specific T cell population induced during priming in the presence of anti-IL-12. However, similar amounts of IL-10 were produced by cultures in the presence or absence of MBP87-106 (data not shown). In fact, IL-10 production was not at all dependent on immunization with MBP in CFA as splenocytes from naive SJL mice previously treated with anti-IL-12 produced as much IL-10 as splenocytes from identically treated animals immunized with MBP (Fig. 4 A). An anti-IL-12 antiserum from a separate source (sheep anti-mouse IL-12; gift of Genetics Institute, Cambridge, MA) was equally effective (not shown).
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Since mRNA encoding IL-12 p40 and p35 has been
found in the spleen and lymph node of naive mice (68),
these results are consistent with the possibility that a low
baseline level of IL-12 tonically suppresses an IL-10 producing cell in lymph node and spleen. IL-12 could act directly on a cell bearing IL-12 receptor 1 and
2 chains
or indirectly through the induction of IFN-
. To clarify
the role of IFN-
in IL-12-mediated suppression of IL-10,
C57BL/6 IFN-
/
and wild-type mice were treated with
anti-IL-12 or control IgG. Splenocytes from IFN-
/
mice treated with anti-IL-12, but not control IgG, produced large quantities of IL-10, at levels comparable to that
produced by wild-type splenocytes from identically treated
donors (Fig. 4 B). Thus, IL-12 suppresses IL-10 production by an IFN-
-independent mechanism.
To define which cell type was responsible for IL-10 production after treatment with anti-IL-12 in vivo, we
compared IL-10 production by splenocytes from anti-IL-12-treated nu/nu, C.B-17 SCID, and wild-type BALB/c
mice. Splenocytes from nu/nu and SCID mice failed to
produce detectable IL-10, whereas spleen cells from normal
BALB/c mice produced amounts of IL-10 comparable to those produced by spleen cells from similarly treated SJL
mice (Fig. 5 A). As these results clearly implicate the T cell
as the source of IL-10 production after anti-IL-12 treatment, we depleted various subpopulations from anti-IL-12-treated SJL spleen preparations before culture and measured IL-10 levels in supernatants 96 h later. As expected,
depletion of CD4+ cells abrogated IL-10 production,
whereas depletion of CD8+ and Mac1+ cells had no effect.
Surprisingly, the B220+-depleted splenocytes produced
significantly less IL-10 than the whole splenocyte population (Fig. 5 B). Furthermore, neither CD4+ T cells nor
B220+ cells purified from spleens of anti-IL-12-treated donors produced IL-10 when cultured separately, whereas IL-10
production was restored when the two subpopulations were
recombined (Fig. 5 C). CD4+ T cells from anti-IL-12-treated
donors produced similar levels of IL-10 whether combined
with purified B cells from control IgG or anti-IL-12-treated
wild-type donors (Fig. 5 C), or with B cells from control
IgG-treated IL-10/
mice (Fig. 5 D). Conversely, CD4+ cells from control IgG-treated wild-type donors or
anti-IL-12-treated IL-10
/
donors failed to produce
detectable IL-10 when combined with B cells from any of
the sources mentioned (Fig. 5, C and D).
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The abrogation of the protective effects
of anti-IL-12 by coinjection of anti-IL-10 (Fig. 3 B) and
the increased incidence of EAE in IL-10/
mice (Fig. 1)
strongly suggested that IL-10 plays a downregulatory role
in the generation of EAE effector cells. To directly demonstrate that the IL-10-producing CD4+ T cell, generated in
the absence of antigenic priming, can downregulate the
generation of EAE effectors, we performed an adoptive
transfer study. SJL mice were injected with splenocytes from
anti-IL-12- or control IgG-treated syngeneic donors and
then immunized with bovine MBP according to the schedule shown in Fig. 1. Splenocytes from anti-IL-12-treated,
but not control IgG-treated, SJL mice significantly inhibited the induction of EAE (Fig. 6) thereby directly demonstrating the biologic importance of the IL-10-producing CD4+ T cells whose presence is revealed when animals are
treated with anti-IL-12.
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Discussion |
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These studies demonstrate a number of novel mechanisms by which cytokine production by the innate immune
system controls an autoimmune response mediated by cells
within the adaptive immune system. In the model systems
we have used, IL-12 produced by macrophages and/or
dendritic cells plays a critical role in the generation of autoimmune effectors, while IL-10 production by antigen
nonspecific regulatory CD4+ T cells subserves a counterregulatory or suppressive function. During the induction of
EAE in susceptible strains of mice, the homeostatic balance
maintained between these antagonistic cytokines is upset to
favor IL-12, the production of which is stimulated by mycobacterial components contained in the adjuvant. On the other hand, the enhanced susceptibility to EAE of IL-10/
mice may be secondary to the loss of the dominant suppressive functions of this cytokine.
Although one might have predicted that IFN- would
be indispensable for the generation and function of autoimmune effector cells, studies using anti-IFN-
mAbs and
IFN-
/
and IFN-
-receptor
/
strains have uniformly
demonstrated that IFN-
is not required and in many cases
exerts a protective effect in EAE (43, 45, 49, 51). It was
therefore somewhat surprising that IL-12
/
mice were
resistant to EAE and that treatment of IFN-
/
mice with anti-IL-12 prevented EAE. Furthermore, the generation of EAE effectors could be markedly inhibited by treatment of highly susceptible SJL mice with anti-IL-12 during
the course of priming alone. Taken together, these results
demonstrate for the first time that a cytokine implicated
in the pathogenesis of EAE, namely IL-12, is indispensable
for its manifestation, whereas the effector molecules IFN-
,
TNF-
, and LT
appear to be redundant (50, 52). These
results should be contrasted with the requirements for both
IL-12 and IFN-
in mediating resistance to a number of intracellular pathogens including Toxoplasma gondii, Mycobacterium tuberculosis, and Listeria monocytogenes (69). Although IFN-
-independent actions have been attributed
to IL-12 in the past (72, 73, 57), in only one other case has
such an action been reported to affect the pathogenesis of
disease. Taylor and Murray (74) found that treatment of
IFN-
/
mice infected with Leishmania donovani with
exogenous IL-12 induced leishmanicidal activity and also
partially restored the near-absent tissue granulomatous response. The action of IL-12 against L. donovani was TNF-
-dependent and involved upregulation of inducible nitric
oxide synthase (iNOS). It was unclear from this study
whether the protective effects of IL-12 were mediated by
antigen-specific CD4+ T cells or by NK cells. Nevertheless, induction of the TNF-
/iNOS pathway may also be
partially responsible for the IFN-
-independent actions of
IL-12 in EAE. We have detected high levels of mRNA encoding both TNF-
and iNOS in the spinal cords of actively immunized C57BL/6 IFN-
/
mice and markedly
reduced levels after treatment with anti-IL-12 (unpublished
data). Although treatment of mice with anti-TNF-
has
been shown to prevent EAE, the contribution of TNF-
to the pathogenesis of demyelination has been difficult to
define because the therapeutic effects of anti-TNF-
appeared to have been mediated by prevention of entry of
pathogenic T cells into the CNS (32). It would therefore
be of interest to assess whether treatment with anti-TNF-
and/or anti-LT
are able to protect IFN-
/
mice
from disease at a time point when pathogenic effector cells
have already entered the CNS. It would also be of interest
to determine whether administration of recombinant TNF-
reverses the protection afforded by the anti-IL-12
treatment of actively immunized IFN-
/
mice.
Because it has previously been reported that IL-12/
mice mount a Th2 response when infected with L. major,
whereas wild-type mice mount a Th1 response (75), we
intensively investigated whether anti-IL-12-treated SJL
mice develop a Th2 response when immunized with MBP
in CFA. Although no evidence for antigen-specific IL-4
production was observed, we consistently observed antigen-independent IL-10 production. The IL-10-producing
population implicated has several unique properties that
distinguish it from conventional Th2 memory cells. After
anti-IL-12 treatment, it was as readily obtained from naive
mice as from mice that had been previously immunized.
The tonic inhibition of IL-10 production by the constitutive presence of IL-12 in vivo is an IFN-
-independent activity as upregulation of IL-10 production was also seen
when IFN-
/
mice were treated with anti-IL-12. Cell
purification studies demonstrated that the IL-10 producing
cell was a CD4+ T cell, but required coculture with B cells
for induction of IL-10 production in vitro; we have not yet
evaluated other antigen-presenting cell types for their potential to activate IL-10 production by these CD4+ T cells.
The CD4+ IL-10-producing cell resembles the IL-4 producing CD4+, NK1.1+ cell in that it produces cytokines in
the absence of priming and therefore appears to be a member of the newly recognized category of unconventional
"natural" T cells in the innate immune system that guide
adaptive responses through the production of Th1/2-modulating cytokines (76, 77). The induction of IL-4 production by CD4+NK1.1+ T cells in response to CD1d stimulation (78, 61) raises the possibility that a nonclassical MHC
molecule may also serve as the ligand for the IL-10 producer and that the IL-10 producing T cell is responding
to an autoantigen presented by the B cell. Currently we
are conducting experiments to clarify the nature of the interaction between the B and T cells responsible for IL-10 production. It should be noted that we did not observe enhanced IL-10 production by spleen cells from IL-12
/
mice, but this may be secondary to adaptive processes that
have taken place in the animal in the absence of IL-12.
The production of IL-10 by the CD4+ T cells is highly
significant biologically as coinjection of anti-IL-10 largely
reversed the autoimmune suppressive effects of anti-IL-12.
Most importantly, IL-10 producing cells from anti-IL-12-
treated naive mice markedly suppressed the induction of
EAE after transfer into sensitized recipients. It is also likely
that the increased incidence of EAE that we observed in
IL-10/
mice is due to the functional absence of this
immunoregulatory cell. The concept that exposure of developing autoimmune effector cells to IL-10 hinders their development is supported by an earlier study in which EAE
was suppressed by the administration of recombinant IL-10
to rats during priming (40). It remains to be seen if this
CD4+ IL-10-producing T cell plays a broader role in immune surveillance by preventing organ-specific autoimmunity. The high incidence of spontaneous autoimmune phenomena in IL-10
/
mice suggest that this might be the
case (79). The innate immune system appears to be biased
towards the development of inflammatory immune responses as IL-10 production is constitutively suppressed by
IL-12. Therefore, CD4+ IL-10-producing T cells may
be needed to curtail overzealous responses to acute infections that would otherwise lead to immunopathology. The
slightly delayed expression of IL-10, as compared to IL-12 and other proinflammatory cytokines, in several infectious
disease models make it a particularly effective downregulator
of the IL-12 response (58). The importance of this negative
regulatory role of IL-10 is highlighted by the emergence of
a lethal immune response, characterized by overproduction
of IL-12, IFN-
, and TNF-
, in IL-10
/
mice infected
with T. gondii (80).
Thus far efforts to manipulate cytokine production to alter the course of autoimmune disease have been focused on modification of the adaptive immune response. Although positive results have been reported, these interventions were mainly effective when administered early in the evolution of the disease before the expression of clinical signs (39, 42, 64). Furthermore, recent reports of Th2-mediated autoimmunity suggest that approaches to induce immune deviation of Th1 effector cells may also ultimately result in immunopathology (54, 55). Collectively, our studies strongly suggest that manipulation of the cytokine milieu, the IL-12/ IL-10 balance, maintained by cells of the innate immune system, can have profound effects on the incidence of autoimmune disease. In chronic autoimmune diseases that progress as a result of the continuous reactivation of autoantigen-specific memory T cells, it would be unlikely that attempts to block IL-12 production or enhance IL-10 production would be therapeutically useful. On the other hand, relapsing remitting autoimmune disease may provide a unique opportunity for effective intervention employed during periods when new effectors, capable of initiating relapses, are recruited from a naive precursor pool. Studies documenting the phenomenon of determinant spreading in EAE and multiple sclerosis suggest that relapses are attributable in large part to the activation of naive T cells specific for cryptic epitopes during or after the peak of the initial episode (81). We have previously suggested that IL-12 production in response to an infectious insult during disease remission may lead to recruitment of Th1 cells (63); consideration should be given to the prophylactic use of IL-12 antagonists and/or IL-10 in patients with autoimmune diseases during epidemic outbreaks that have been associated with autoimmune sequelae.
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Footnotes |
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Received for publication Received for publication 23 September 1997 and in revised form 3 December 1997..
The authors would like to thank D. Presky and M.K. Gately and J. Magram for generously providing anti- IL-12 antiserum and the IL-12 ![]() |
References |
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