IL-1
, but not IL-1ß, is required for contact-allergen-specific T cell activation during the sensitization phase in contact hypersensitivity
Susumu Nakae,
Chie Naruse-Nakajima11,
Katsuko Sudo,
Reiko Horai,
Masahide Asano1 and
Yoichiro Iwakura
Center for Experimental Medicine, Institute of Medical Science, University of Tokyo, 4-6-1 Shirokanedai, Minato-ku, Tokyo 108-8639, Japan
1 Present address: Institute for Experimental Animals, School of Medicine, Kanazawa University, 13-1 Takaramachi, Kanazawa 920-8640, Japan
Correspondence to:
Y. Iwakura
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Abstract
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Contact hypersensitivity (CHS) is a T cell-mediated cellular immune response caused by epicutaneous exposure to contact allergens. In this reaction, after the first epicutaneous allergen sensitization, Langerhans cells (LC) catch allergens and migrate from the skin to draining lymph nodes (LN) and activate naive T cells. Although IL-1 is suggested to be involved in these processes, the mechanisms have not been elucidated completely. In this report, to elucidate roles of IL-1
and IL-1ß in CHS, we analyzed ear swelling in 2,4,6-trinitrochlorobenzene (TNCB)-induced CHS using gene-targeted mice. We found that ear swelling was suppressed in IL-1
-deficient (IL-1
/) mice but not in IL-1ß/ mice. LC migration from the skin into LN was delayed in both IL-1
/ and IL-1ß/ mice, suggesting that this defect was not the direct cause for the reduced CHS in these mice. However, we found that the proliferative response of trinitrophenyl (TNP)-specific T cells after sensitization with TNCB was specifically reduced in IL-1
/ mice. Furthermore, adoptive transfer of TNP-conjugated IL-1-deficient epidermal cells (EC) into wild-type mice indicated that only IL-1
, but not IL-1ß, produced by antigen-presenting cells in EC could prime allergen-specific T cells. These observations indicate that IL-1
, but not IL-1ß, plays a crucial role in TNCB-induced CHS by sensitizing TNP-specific T cells.
Keywords: contact hypersensitivity, IL-1, knockout mice, Langerhans cell
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Introduction
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Contact hypersensitivity (CHS) is a T cell-mediated cellular immune response caused by repeated epicutaneous exposure against contact allergens, chemically reactive haptens which are able to bind directly to soluble or cell-associated proteins. This response is divided into two phases, sensitization and elicitation phases. After the first epicutaneous allergen sensitization, Langerhans cells (LC) catch allergens and migrate from the skin to draining lymph nodes (LN) where naive T cells are thought to be primed against allergens through T cellLC interaction (1). In the elicitation phase, allergen-specific T cells in LN are activated upon re-challenging with the same allergen and migrate from LN to the place where the allergen is challenged, resulting in local inflammation. Using MHC class I/ and MHC class II/ mice, it was shown that CD8+ T cells act as effector cells, while CD4+ T cells play a regulatory role in this reaction (2,3). It is known that LC are important as antigen-presenting cells (APC) in the sensitization phase, but they are not essential for the elicitation phase (4).
Both IL-1
and IL-1ß cause inflammation and also augment immune reactions through activation of lymphocytes, although they are encoded by distinct genes and have little amino acid sequence homology (5). The expression of proinflammatory cytokine mRNAs including those of IL-1
, IL-1ß and tumor necrosis factor (TNF)-
is increased in the contact-allergen-sensitized skin (6). In the epidermis, IL-1
is mainly produced by kelatinocytes (KC) while IL-1ß is mainly produced by LC (79). IL-1ß mRNA is expressed earlier than any other proinflamatory cytokine in CHS (9,10).
It was shown that systemic administration of recombinant IL-1
(rIL-1
) or local treatment of rIL-1ß causes activation and migration of LC (1012). LC migration was impaired by systemic administration of anti-IL-1ß antibody (13). Furthermore, treatment with IL-1 receptor antagonist (IL-1ra), which is a negative regulator of IL-1
and IL-1ß, abolished the enhanced migration of LC (14,15) and corneal LC migration was impaired in mice deficient in the IL-1 receptor type 1 (IL-1RI) gene (16). These observations suggest that both IL-1
and IL-1ß can promote LC migration that may be important in CHS. Consistent with these observations, CHS was markedly reduced by the intradermal administration with anti-IL-1ß mAb (10). However, it was reported that CHS was not affected by the treatment with anti-IL-1
mAb (10). On the other hand, it was shown that IL-1ß was not involved in oxazolone-induced CHS using IL-1ß/ mice (17,18). Furthermore, it was shown that low-dose 2,4,6-trinitrochlorobenzene (TNCB)-induced CHS was suppressed in IL-1ß/ mice, whereas the high-dose response was not (18). Thus, these apparently controversial findings claim that the role of IL-1 in CHS still remains to be elucidated.
CHS develops through several distinct steps including LC migration from the skin into LN, allergen-specific T cell activation and cell infiltration into inflamed regions. It is suggested that various cytokines, chemokines, adhesion molecules and co-signal molecules are involved in these processes (1927). Recently, we have shown that IL-1 produced by APC plays a crucial role in antigen-specific T cell priming and clonal expansion using IL-1
/ß/ and IL-1ra/ mice (28). In this study we tried to elucidate roles of IL-1 in CHS using these IL-1/ mice. We found that TNCB-induced CHS at both low and high doses was greatly reduced in IL-1
/ and IL-1
/ß/ mice, but not in IL-1ß/ mice. The induction of trinitrophenyl (TNP)-specific T cells was abolished in IL-1
/ and IL-1
/ß/ mice, whereas that in IL-1ß/ mice was normal, indicating that IL-1
produced by mature LC in LN is required to prime naive T cells against contact allergens in the sensitization phase of CHS.
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Methods
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Mice
IL-1
/, IL-1ß/ and IL-1
/ß/ mice were generated by homologous recombination as described previously and backcrossed to C57BL/6J or BALB/cA mice for eight generations (29). DO11.10 transgenic mice (BALB/cA background) were kindly provided by Dr Dennis Y. Loh (30). All the mice were housed under specific pathogen-free conditions in an environmentally controlled clean room at the Center for Experimental Medicine, Institute of Medical Science, University of Tokyo. The experiments were conducted according to the institutional ethical guidelines for animal experiments and the safety guidelines for gene manipulation experiments. Sex- and age-matched adult mice (812 weeks old) were used for the experiments.
CHS response
TNCB (Tokyo Kasei, Tokyo, Japan)-induced CHS was assayed as described previously (17,18). Briefly, the abdomen of mice was shaved and sensitized epicutaneously with 25 µl of low-dose (0.3%) or high-dose (3.0%) TNCB dissolved in acetone and olive oil mixture (4:1). On day 5 after sensitization, the outside of one ear (auricle) of mice was challenged with 25 µl of 1.0% TNCB and the outside of the other ear was treated with 25 µl of vehicle alone. At 24 h after the second challenge, mice were euthanized and a disk of ear tissue was removed from both ears using a 6-mm biopsy punch, then each of ear disk was weighed. The difference between TNCB-treated and vehicle-treated ear weights of each mouse is shown as the amount of swelling in TNCB-induced CHS. Ear swelling is calculated as follows: increment of ear swelling = weight of challenged ear weight of vehicle-treated ear)/weight of vehicle-treated earx100 (%).
Migration and maturation of LC
Mice were shaved at the dorsal and abdominal area, and painted with 50 µl of 0.5% FITC isomer I (Sigma, St Louis, MO) dissolved in acetone and dibutylphthalate mixture (1:1). At 24 h after FITC painting, inguinal, axillary and brachial LN were harvested and pooled. Single-cell suspension was prepared from collagenase-treated LN and stained with biotinylated anti-mouse CD11c mAb (HL3; PharMingen, San Diego, CA) after pre-incubation with anti-Fc receptor mAb (2.4G2; PharMingen). To assess the maturation of LC, LN cells were stained with phycoerythrin (PE)anti-mouse CD40 mAb (3.23; Immunotech, Marseilles, France) and PerCPstreptavidin (PharMingen). The frequency of CD11c+ FITC+ cells or expression levels of CD40 on CD11c+ FITC+ cells in LN were analyzed with a FACScan (Becton Dickinson, Mountain View, CA) using Lysys II software (Becton Dickinson). Viable cells were determined by forward and side scatter.
Intracellular staining of IL-1
in peritoneal exudate cells (PEC) and mature LC
Thioglycolate (TGC)-induced PEC were prepared as described previously (29). LN cells from FITC-painted mice (at 24 h after painting) and PEC were harvested and stimulated with 5 µg/ml of LPS for 6 h, then cells were suspended in a staining buffer (HBBS containing 2% FCS and 0.1% sodium azide). After blocking with anti-Fc
RII/III receptor mAb (2.4G2; PharMingen), PEC were treated with FITCanti-mouse Mac-1 mAb and LN cells from FITC-painted mice were treated with biotinylated anti-CD11c mAb (HL3; PharMingen). Then, cells were incubated with PerCPstreptavidin (PharMingen), followed by fixation with PBS containing 4% paraformaldehyde for 20 min. After washing with a permeabilization buffer [0.1% saponin (Sigma) in the staining buffer], cells were incubated with PEhamster anti-mouse IL-1
mAb (ALF-161; PharMingen) or isotype-matched control mAb (PEhamster IgG; Immunotech) in the permeabilization buffer for 30 min at 4°C. Cells were washed with the permeabilization buffer and analyzed using a FACSCalibur (Becton Dickinson) and CellQuest software (Becton Dickinson).
T cell proliferative response assay
For TNP-specific T cell proliferative response, inguinal, axillary and brachial LN were harvested and pooled 5 days after the sensitization with 3.0% TNCB. Single-cell suspension was prepared, and T cells were purified by passing through MACS separation columns (Miltenyi Biotec, Bergisch Gladbach, Germany) to remove anti-B220 and anti-Mac-1 mAb-reactive cells. To prepare TNP-conjugated APC, the spleen was harvested from wild-type mice and single-cell suspension was prepared. After treatment with hemolysis buffer (17 mM TrisHCl, 140 mM NH4Cl, pH 7.2), T cells were depleted by passing through MACS columns using anti-Thy1.2, anti-CD4 and anti-CD8 magnetic beads (Miltenyi Biotec). T cell-depleted spleen cells were incubated in PBS containing 100 mM trinitrobenzene sulfonate (TNBS; Wako, Osaka, Japan) at 37°C for 5 min and irradiated with
-rays (3500 rad). LN T cells from TNCB-sensitized mice (5x105 cells/well) and TNP-conjugated APC (2x105 cells/well) were cultured in 200 µl of RPMI 1640 (Sigma) containing 50 mM 2-mercaptoethanol (Gibco/BRL, Gaithersburg, MD), 50 µg/ml streptomycin (Meiji, Tokyo, Japan), 50 µg/ml penicillin (Meiji) and 10% heat-inactivated FCS (Sigma) using 96-well flat-bottom plates for 72 h. For ovalbumin (OVA)-specific primary T cell proliferative responses, inguinal, axillary and brachial LN of DO11.10 transgenic mice were incubated with anti-B220, anti-Mac-1 and anti-CD8 magnetic beads, and CD4+ T cells were purified by passing through MACS column. To collect skin LC, ear skin was incubated with 0.15% trypsin (Gibco) and 50 U/ml of dispase (Godoshusei, Tokyo, Japan) in PBS for 1 h at 37°C and epidermal sheets were prepared. A single-cell suspension from epidermal sheets was prepared and CD11c+ LC were isolated with the MACS system by positive selection using biotinylated anti-CD11c and streptavidinmagnetic beads (Miltenyi Biotec). DO11.10 CD4+ T cells (5x105 cells/well) were cultured with TGC-induced PEC or CD11c+ skin LC (5x103 cells/well) in the absence or presence of the OVA peptide (0.1 µM) with or without rIL-1
(100 pg/ml) for 72 h. After 72 h, cells were labeled with [3H]thymidine (0.25 µCi/ml; Amersham, Little Chalfont, UK) for 6 h, then harvested using a Micro 96 cell harvester (Skatron, Lier, Norway) and 3H radioactivity was measured using Micro Beta System (Pharmacia Biotech, Piscataway, NJ).
Preparation of epidermal cells (EC)
Mice were shaved at the dorsal and abdominal areas, and the hair completely removed with a hair-remover cream at 2 days before experiments. The shaved skin were harvested and the hypodermal tissue removed. The skin was incubated with 0.15% trypsin (Gibco) and 50 U/ml of dispase (Godoshusei) in PBS for 1 h at 37°C and epidermal sheets were prepared. Epidermal sheets were stirred in PBS containing 2% FCS for 15 min at room temperature. EC suspension was obtained by filtering the epidermal sheet suspension with the nylon mesh. TNP-conjugated EC were prepared by incubating the cell suspension with 100 mM TNBS for 10 min at 37°C as described above.
Adoptive transfer of TNP-conjugated splenocytes and induction of CHS
TNP-conjugated EC from wild-type or IL-1-deficient mice were suspended in PBS and injected into wild-type mice s.c. (2x106 cells/mouse). At 7 days after injection, the outside of one ear of mice was challenged with 25 µl of 1.0% TNCB and the outside of the other ear was applied with 25 µl of vehicle alone. At 24 h after the challenge, ear swelling was measured as described above.
Measurement of antibody titers
Mice were sensitized and challenged with TNCB as described above. Four days after the challenge, the sera were collected. TNP-specific Ig levels in the sera were measured by sandwich ELISA. To TNP-specific antibodies, TNP-BSA in PBS (10 µg/ml) was coated on Falcon 3912 Micro Test III flexible assay plates (Becton Dickinson, Oxnard, CA) at 37°C overnight. After washing with TBS, serial diluted serum samples were applied and incubated at room temperature for 1 h. After incubation for 1 h the well was washed with TBS + 0.05% Tween 20, followed by the addition of alkaline phosphatase-conjugated goat anti-mouse IgG (Zymed, San Francisco, CA). Alkaline phosphatase activity was measured using Substrate Phosphatase SIGMA104 (Sigma) as the substrate. Results are expressed by the absorbancy at 415 nm.
Statistics
Student's t-test was used for statistical evaluation of the results.
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Results
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CHS with high-dose TNCB
It was previously reported that CHS was markedly reduced in IL-1ß/ mice when mice were sensitized with low-dose TNCB, whereas high-dose sensitization induced comparable CHS between IL-1ß/ and wild-type mice (17). These results suggest that IL-1ß-deficiency can be compensated by IL-1
at high-dose TNCB sensitization, but not at low-dose sensitization. In order to identify the differential roles of IL-1
and IL-1ß, we re-examined CHS at both low and high TNCB doses using IL-1
/, IL-1ß/ and IL-1
/ß/ mice on the C57BL/6J background (Fig. 1A and B
). In contrast to the previous report, we found that similar levels of CHS were induced in IL-1ß/ and wild-type mice with both low and high doses of TNCB. Furthermore, we found that CHS was significantly reduced in IL-1
/ mice as well as IL-1
/ß/ mice at 24 h after the challenge (Fig. 1A and B
). In our assay system, ear inflammation in wild-type and IL-1/ mice calmed down to the basal levels at 48 h after the challenge (data not shown). We assessed the effect of genetic background using IL-1-deficient mice on the BALB/cA background, because genetic background affects CHS (31). Similar results were obtained using IL-1
/ß/ and IL-1ß/ mice on the BALB/cA background (data not shown). However, the effect of IL-1
deficiency was only small on the BALB/cA background compared to that on the C57BL/6J background. Since we observed more pronounced effects of the deficiency in IL-1
/ß/ mice compared with IL-1
/ mice on the BALB/cA background, it was suggested that IL-1ß also plays some role in a synergistic manner with IL-1
in mice of this background.
Effects of IL-1-deficiency on LC migration and maturation
To elucidate roles of IL-1 in CHS, we first examined the migration ability of LC from the skin into draining LN. After sensitization with 0.5% FITC, CD11c+ FITC+ cell counts in draining LN were measured by flow cytometry analysis (Fig. 2A
). The content of CD11c+ FITC+ cells in LN from both IL-1
/, IL-1ß/ and IL-1
/ß/ mice was significantly reduced compared with wild-type mice after FITC treatment. The migration in IL-1
/ß/ mice was most severely affected at 24 h after the treatment. However, the FITC+ LC content in LN of both IL-1
/ and IL-1ß/ mice became similar to that of wild-type mice at 36 h after FITC treatment (Fig. 2B
).

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Fig. 2. Effects of IL-1-deficiency on LC migration and maturation. Wild-type mice and IL-1-deficient mice were epicutaneously sensitized with 0.5% FITC. After FITC painting, draining LN were harvested and analyzed for FITC and CD40 expression by flow cytometry. (A) Content of FITC+ cells among CD11c+ cells at 24 h after FITC painting. (B) Kinetics of CD11c+ FITC+ LC migration. (C) Expression of CD40 on CD11c+ FITC+ cells. One set of representative data from six independent experiments is shown.
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It is known that skin LC are as immature as dendritic cells and they maturate during migration from the skin into draining LN. To examine whether IL-1 is involved in LC maturation, expression levels of CD40 on CD11c+ FITC+ cells were examined by flow cytometry analysis (Fig. 2C
), because CD40 is expressed on mature LC, but not on immature LC (32). Comparable expression levels of CD40 were observed on FITC+ cells from IL-1-deficient and wild-type mice. Furthermore, we could not detect any difference in the expression levels of CD80, CD86 and CD54 on CD11c+ FITC+ cells from these mice (data not shown). These results indicate that both IL-1
and IL-1ß are involved in LC migration, but not in LC maturation.
Effect of IL-1 deficiency on the induction of antigen-specific T cells
We have recently reported that IL-1 produced by APC plays an important role in antigen-specific T cell priming (28). Thus, we examined the role of IL-1 in the induction of antigen-specific T cells by sensitizing with high-dose TNCB. LN T cells from IL-1-deficient mice sensitized with TNCB were cultured with TNP-conjugated and T cell-depleted wild-type splenocytes (Fig. 3
). Proliferative responses of IL-1
/ T cells as well as IL-1
/ß/ T cells were markedly impaired, while those of IL-1ß/ T cells were comparable with those of wild-type T cells. Previously, we showed that T cell development and intrinsic T cell function is normal in IL-1-deficient mice (28). Thus, the reduced proliferative responses of IL-1
/ and IL-1
/ß/ T cells suggest that LN T cells from these IL-1-deficient mice were not sensitized sufficiently in vivo, because APC were derived from wild-type mice.

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Fig. 3. Proliferation of TNCB-sensitized LN T cells after in vitro re-stimulation. On day 5 after sensitization with 3.0% TNCB, LN were harvested and T cells were purified through MACS columns. These T cells and irradiated TNP-conjugated APC or non-treated APC from wild-type splenocytes were cultured for 72 h, and then the proliferative response was assessed by the incorporation of [3H]thymidine. The data were reproducible in three independent experiments.
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IL-1
expression on LC migrating into draining LN
Although LC are known as APC in the sensitization phase of CHS, in immature skin, LC do not produce IL-1
(32, 33). It is not known whether or not mature LC which migrate from the skin into draining LN produce IL-1
. We carried out flow cytometric analysis of the LN cells from FITC-painted wild-type mice after stimulation with LPS for 12 h. As shown in Fig. 4
(A and B), both CD11c+ FITC+ LN cells and CD11b+ PEC produced IL-1
. Cells from IL-1
/ mice (Fig. 4A and B
) and non-stimulated TGC-induced CD11b+ PEC (data not shown) did not produce IL-1
. Agonistic anti-CD40 mAb-treated CD11c+FITC+ cells also produced IL-1
(data not shown).
When CD4+ T cells from DO11.10 transgenic mice were cultured with CD11c+ cells from the skin or CD11b+ PEC in the presence of the OVA peptide, CD11c+ cells from the skin could activate DO.11.10 CD4+ T cells ~5 times more strongly than TGC-induced CD11b+ PEC (Fig. 4C
), suggesting that LC are the major APC in this reaction. Under these conditions, the proliferative response of T cells cultured with CD11c+ cells from IL-1
/ß/ mice was low compared with that of wild-type mice and recovered to wild-type levels in the presence of rIL-1
(Fig. 4C
). These results showed that mature LC in draining LN could produce IL-1
, which may play an important role in contact-allergen-specific T cell activation.
Adoptive transfer of TNP-conjugated EC
As shown above, IL-1, especially IL-1
, may play an important role not only in LC migration but also in contact-allergen-specific T cell activation. To examine roles of IL-1 produced by LC in T cell activation, we performed adoptive transfer of TNP-conjugated wild-type or IL-1-deficient EC, which contain ~2% LC, into wild-type mice which were not sensitized with contact allergens. When TNP-conjugated IL-1
/ EC as well as IL-1
/ß/ EC were transferred into wild-type mice, their ear swelling after treatment with 1.0% TNCB was markedly reduced compared with those transferred with TNP-conjugated wild-type EC (Fig. 5
). On the other hand, when transferred with TNP-conjugated IL-1ß/ EC, ear swelling was similar to that of wild-type mice (Fig. 5
). These results suggest that IL-1
, but not IL-1ß, produced by APC in EC, most likely to be LC, is required for TNP-specific T cell activation.

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Fig. 5. CHS induction in wild-type mice transferred with TNP-conjugated EC from IL-1-deficient mice. TNP-conjugated EC from wild-type or IL-1-deficient mice were injected into wild-type mice s.c. After 7 days, the ear was challenged with high-dose TNCB and ear swelling was measured 24 h later. Each circle represents an individual mouse and an average ± SD are shown. *P < 0.005, #P < 0.001.
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Distinctive roles of IL-1
and IL-1ß in allergen-specific antibody production
To examine the possibility that IL-1 is involved in CHS through contact-allergen-specific antibody production, TNCB-specific antibody production was measured in those IL-1-deficient mice. TNCB-specific antibody production was measured in those IL-1-deficient mice. As shown in Fig. 6
, serum TNP-specific IgG levels in IL-1ß/ mice as well as in IL-1
/ß/ mice were lower than those in wild-type mice, while those in IL-1
/ mice were comparable. These results are consistent with our previous report (38) showing that IL-1ß, but not IL-1
, is mainly involved in the antibody production. Thus, it was shown that molecular species of IL-1 differ between CHS and antibody production, suggesting different mechanisms are involved in those reactions.
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Discussion
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Previously, IL-1ß rather than IL-1
was thought of as a mediator of CHS, because CHS was suppressed by the administration of anti-IL-1ß mAb, but not by anti-IL-1
mAb (10). Furthermore, low-dose TNCB-induced CHS was suppressed in IL-1ß/ mice, although high-dose TNCB-induced CHS and oxazolone-induced CHS were normal in these mice (17, 18). Here, we demonstrated that TNCB-induced CHS was markedly impaired in IL-1
/ mice as well as in IL-1
/ß/ mice, while that in IL-1ß/ mice was not significantly affected, indicating that IL-1
rather than IL-1ß plays a crucial role in this CHS. Our results are consistent with the previous report using IL-1ß/ mice in which no effect was observed at a high dose (18), but contrary to the results obtained by using IL-1 mAb (10). However, not only high-dose but also low-dose TNCB-induced CHS was normal in IL-1ß/ mice in our system. We think that probably the difference in the experimental conditions and genetic background of the mice may have affected the results (31). With regard to this, we found that suppression of ear swelling in IL-1
/ mice of the C57BL/6J background was similar to that of IL-1
/ß/ mice, whereas that of IL-1
/ mice of the BALB/cA background was milder compared with that of IL-1
/ß/ mice (data not shown). Thus, the action of each IL-1 molecular species may be different in 129xB6 background mice, which were used in the previous reports (17, 18), from that in the C57BL/6J background mice, which we used in this study.
LC migration was suppressed both in IL-1
/ and IL-1ß/ mice. However, ear swelling was only suppressed in IL-1
/ mice, indicating that the delayed LC migration may not completely explain the suppression of CHS in IL-1
/ mice. On the other hand, we found that allergen-specific T cell priming was reduced in IL-1
/ mice, but not in IL-1ß/ mice (Figs 2 and 4C
). Furthermore, we showed that ear swelling upon treatment with TNCB was significantly reduced in mice previously transferred with TNP-conjugated EC from IL-1
/ or IL-1
/ß/ mice compared with the mice transferred with those cells from wild-type mice or IL-1ß/ mice. These observations suggest that IL-1
produced by APC plays a crucial role in the initiation of a primary immune response by activating allergen-specific T cells. In this context, we showed that IL-1
was produced by mature CD11c+ FITC+ LC in LN, although so far it was only believed that IL-1
was produced in skin KC, but not in immature skin LC (68). Taken together, these observations suggest that IL-1
produced by mature LC in LN migrating from the skin plays an important role in allergen-specific T cell priming.
We demonstrated that IL-1
/, but not IL-1ß/, produced by APC was responsible for the proliferative responses of antigen-specific T cells. This observation suggests that IL-1
is required for the development of antigen-specific memory T cells. Although we tried to demonstrate that the proportion of memory T cells (CD44+ or CD45RB- cells) in LN T cells was decreased in TNCB-sensitized IL-1
/ mice compared to the wild-type mice, we could not detect the difference (data not shown). Probably, the proportion of TNP-specific memory T cells was too small to detect in the total T cell population. At this moment, we cannot exclude the possibility completely that TNP-specific T cells of IL-1
/ mice became anergic because of insufficient activation.
This shows a clear contrast with the essential role of IL-1ß in contact-allergen-specific antibody production. Recently, we have also shown that antibody production against sheep red blood cells is severely impaired in IL-1ß/ mice, in agreement with the present observation (38). Thus, it is shown that IL-1
and IL-1ß play distinct roles in CHS; IL-1
is mainly involved in contact-allergen-specific T cell priming, while IL-1ß is involved in contact-allergen-specific antibody production. In this context, it was reported that, in germinal centers of human tonsils, IL-1ß, but not IL-1
, is strongly expressed in follicular dendritic cells which play important roles in affinity maturation and isotype switching of Ig through interaction with B cells (34). Since Ig class switching depends on CD40CD40 ligand interaction, IL-1ß may be required for the B cellfollicular dendritic cell interaction. Consistent with this notion, we have recently reported that IL-1 is required for the induction of CD40 ligand on naive T cells (28). On the other hand, it is known that T cell priming through T cellLC interactions occurs in the T cell zones in LN. We showed that IL-1
plays a major role in this process. Thus, the functional specificity of IL-1
and IL-1ß seems to depend on the difference of the producing cells and the sites of production.
It was reported that CHS was suppressed in Fc
R/ mice (36), suggesting antibodies were involved in this reaction. This observation apparently contradicts our data that hapten-specific antibody levels were not correlated with the development of CHS. However, since athymic mice displayed a similar neutrophil response in oxazolone-induced contact dermatitis (36) and MHC II/ mice in which CD4+ cells are absent displayed elevated CHS (3), it seems unlikely that hapten-specific antibody is involved in this reaction. Similar dissociation between CHS and allergen-specific antibody production was observed in CD80/CD86/, CD154/ and OX40 ligand/ mice (24,27, 35). However, it is not well resolved how Fc
R is involved in CHS. With regard to this, Zhang and Tinkle have shown that production of inflammatory cytokines, such as TNF-
and IL-1ß, was reduced in Fc
R/ mice (36). Since these cytokines are induced in the skin during the first allergen sensitization in mice due to the irritant effects of the chemicals, it is considered that allergen-specific Ig is not involved in this reaction. Therefore, Fc
R may be involved in the activation of the cutaneous innate immune system that is important for the inflammatory cell infiltration and cytokine response (36). It is possible that this defect of proinflammatory cytokine production is responsible for the diminished CHS in Fc
R/ mice.
In conclusion, we have shown that IL-1
, but not IL-1ß, produced by mature LC migrating into LN is essential for contact-allergen-specific T cell activation during the sensitization phase of CHS. Thus, IL-1
and IL-1ß play clearly distinct roles in the immune system, probably depending on the localization of the producing cells and responding cells in a lymphoid organ. We are now trying to further elucidate these complex regulatory mechanisms of the immune system involving IL-1
and IL-1ß. These studies may provide important cues for the control of CHS.
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Acknowledgments
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We thank Drs Jun Tanaka, Shinobu Saijo and Kaori Hamaguchi for technical support in the experiments, and all the members of the laboratory for their excellent animal care. This work was supported by grants from the Ministry of Education, Culture, Sport, Science and Technology of Japan, Ministry of Health and Welfare of Japan, Core Research for Evolutional Science and Technology (CREST), the Japan Society for the Promotion of Science, and Pioneering Research Project in Biotechnology.
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Abbreviations
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APC antigen-presenting cell |
CHS contact hypersensitivity |
EC epidermal cell |
IL-1ra IL-1 receptor antagonist |
IL-1RI IL-1 receptor type II |
IL-1RII IL-1 receptor type II |
KC keratinocyte |
LC Langerhans cell |
LN lymph node |
OVA ovalbumin |
PE phycoerythrin |
PEC peritoneal exudate cell |
TGC thioglycolate |
TNBS trinitrobenzene sulfonate |
TNCB 2,4,6-trinitrochlorobenzene |
TNF tumor necrosis factor |
TNP trinitrophenyl |
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Notes
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Transmitting editor: K.Sugamura
Received 7 June 2001,
accepted 24 August 2001.
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References
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---|
-
Kripke, M. L., Munn, C. G., Jeevan, A., Tang, J. M. and Bucana, C. 1990. Evidence that cutaneous antigen-presenting cells migrate to regional lymph nodes during contact sensitization. J. Immunol. 145:2833.[Abstract/Free Full Text]
-
Krasteva, M., Kehren, J., Horand, F., Akiba, H., Choquet, G., Ducluzeau, M. T., Tedone, R., Garrigue, J. L., Kaiserlian, D. and Nicolas, J. F. 1998. Dual role of dendritic cells in the induction and down-regulation of antigen-specific cutaneous inflammation. J. Immunol. 160:1181.[Abstract/Free Full Text]
-
Bouloc, A., Cavani, A. and Katz, S. I. 1998. Contact hypersensitivity in MHC class II-deficient mice depends on CD8 T lymphocytes primed by immunostimulating Langerhans cells. J. Invest. Dermatol. 111:44.[Abstract]
-
Grabbe, S., Steinbrink, K., Steinert, M., Luger, T. A. and Schwarz, T. 1995. Removal of the majority of epidermal Langerhans cells by topical or systemic steroid application enhances the effector phase of murine contact hypersensitivity. J. Immunol. 155:4207.[Abstract]
-
Dinarello, C. A. 1991. Interleukin-1 and interleukin-1 antagonism. Blood 77:1627.[Abstract]
-
Enk, A. H. and Katz, S. I. 1992. Early molecular events in the induction phase of contact sensitivity. Proc. Natl Acad. Sci. USA 89:1398.[Abstract]
-
Kupper, T. S., Ballard, D. W., Chua, A. O., McGuire, J. S., Flood, P. M., Horowitz, M. C., Langdon, R., Lightfoot, L. and Gubler, U. 1986. Human keratinocytes contain mRNA indistinguishable from monocyte interleukin 1 alpha and beta mRNA. Keratinocyte epidermal cell-derived thymocyte-activating factor is identical to interleukin 1. J. Exp. Med. 164:2095.[Abstract]
-
Heufler, C., Topar, G., Koch, F., Trockenbacher, B., Kampgen, E., Romani, N. and Schuler, G. 1992. Cytokine gene expression in murine epidermal cell suspensions: interleukin 1 beta and macrophage inflammatory protein 1 alpha are selectively expressed in Langerhans cells but are differentially regulated in culture. J. Exp. Med. 176:1221.[Abstract]
-
Enk, A. H. and Katz, S. I. 1992. Early events in the induction phase of contact sensitivity. J. Invest. Dermatol. 99:39S.[Abstract]
-
Enk, A. H., Angeloni, V. L., Udey, M. C. and Katz, S. I. 1993. An essential role for Langerhans cell-derived IL-1 beta in the initiation of primary immune responses in skin. J. Immunol. 150:3698.[Abstract/Free Full Text]
-
Roake, J. A., Rao, A. S., Morris, P. J., Larsen, C. P., Hankins, D. F. and Austyn, J. M. 1995. Dendritic cell loss from nonlymphoid tissues after systemic administration of lipopolysaccharide, tumor necrosis factor, and interleukin 1. J. Exp. Med. 181:2237.[Abstract]
-
Cumberbatch, M., Dearman, R. J. and Kimber, I. 1997. Langerhans cells require signals from both tumour necrosis factor-alpha and interleukin-1 beta for migration. Immunology 92:388.[ISI][Medline]
-
Stoitzner, P., Zanella, M., Ortner, U., Lukas, M., Tagwerker, A., Janke, K., Lutz, M. B., Schuler, G., Echtenacher, B., Ryffel, B., Koch, F. and Romani, N. 1999. Migration of Langerhans cells and dermal dendritic cells in skin organ cultures: augmentation by TNF-alpha and IL-1beta. J. Leuk. Biol. 66:462.[Abstract]
-
Kondo, S., Pastore, S., Fujisawa, H., Shivji, G. M., McKenzie, R. C., Dinarello, C. A. and Sauder, D. N. 1995. Interleukin-1 receptor antagonist suppresses contact hypersensitivity. J. Invest. Dermatol. 105:334.[Abstract]
-
Wang, B., Zhuang, L., Fujisawa, H., Shinder, G. A., Feliciani, C., Shivji, G. M., Suzuki, H., Amerio, P., Toto, P. and Sauder, D. N. 1999. Enhanced epidermal Langerhans cell migration in IL-10 knockout mice. J. Immunol. 162:277.[Abstract/Free Full Text]
-
Dekaris, I., Zhu, S. N. and Dana, M. R. 1999. TNF-alpha regulates corneal Langerhans cell migration. J. Immunol. 162:4235.[Abstract/Free Full Text]
-
Zheng, H., Fletcher, D., Kozak, W., Jiang, M., Hofmann, K. J., Conn, C. A., Soszynski, D., Grabiec, C., Trumbauer, M. E., Shaw, A., et al. 1995. Resistance to fever induction and impaired acute-phase response in interleukin-1 beta-deficient mice. Immunity 3:9.[ISI][Medline]
-
Shornick, L. P., De Togni, P., Mariathasan, S., Goellner, J., Strauss-Schoenberger, J., Karr, R. W., Ferguson, T. A. and Chaplin, D. D. 1996. Mice deficient in IL-1beta manifest impaired contact hypersensitivity to trinitrochlorobenzone. J. Exp. Med. 183:1427.[Abstract]
-
Sligh, J. E., Jr, Ballantyne, C. M., Rich, S. S., Hawkins, H. K., Smith, C. W., Bradley, A. and Beaudet, A. L. 1993. Inflammatory and immune responses are impaired in mice deficient in intercellular adhesion molecule 1. Proc. Natl Acad. Sci. USA 90:8529.[Abstract/Free Full Text]
-
Tedder, T. F., Steeber, D. A. and Pizcueta, P. 1995. L-selectin-deficient mice have impaired leukocyte recruitment into inflammatory sites. J. Exp. Med. 181:2259.[Abstract]
-
Subramaniam, M., Saffaripour, S., Watson, S. R., Mayadas, T. N., Hynes, R. O. and Wagner, D. D. 1995. Reduced recruitment of inflammatory cells in a contact hypersensitivity response in P-selectin-deficient mice. J. Exp. Med. 181:2277.[Abstract]
-
Staite, N. D., Justen, J. M., Sly, L. M., Beaudet, A. L. and Bullard, D. C. 1996. Inhibition of delayed-type contact hypersensitivity in mice deficient in both E-selectin and P-selectin. Blood 88:2973.[Abstract/Free Full Text]
-
Tang, A., Judge, T. A. and Turka, L. A. 1997. Blockade of CD40CD40 ligand pathway induces tolerance in murine contact hypersensitivity. Eur. J. Immunol. 27:3143.[ISI][Medline]
-
Rauschmayr-Kopp, T., Williams, I. R., Borriello, F., Sharpe, A. H. and Kupper, T. S. 1998. Distinct roles for B7 costimulation in contact hypersensitivity and humoral immune responses to epicutaneous antigen. Eur. J. Immunol. 28:4221.[ISI][Medline]
-
Grabbe, S. and Schwarz, T. 1998. Immunoregulatory mechanisms involved in elicitation of allergic contact hypersensitivity. Immunol. Today 19:37.[ISI][Medline]
-
Berlin-Rufenach, C., Otto, F., Mathies, M., Westermann, J., Owen, M. J., Hamann, A. and Hogg, N. 1999. Lymphocyte migration in lymphocyte function-associated antigen (LFA)-1-deficient mice. J. Exp. Med. 189:1467.[Abstract/Free Full Text]
-
Chen, A. I., McAdam, A. J., Buhlmann, J. E., Scott, S., Lupher, M. L., Jr, Greenfield, E. A., Baum, P. R., Fanslow, W. C., Calderhead, D. M., Freeman, G. J. and Sharpe, A. H. 1999. Ox40-ligand has a critical costimulatory role in dendritic cell:T cell interactions. Immunity 11:689.[ISI][Medline]
-
Nakae, S., Asano, M., Horai, R., Sakaguchi, N. and Iwakura, Y. 2001. IL-1 enhances T cell-dependent antibody production through induction of CD40 ligand and OX40 on T cells. J. Immunol. 167:90.[Abstract/Free Full Text]
-
Horai, R., Asano, M., Sudo, K., Kanuka, H., Suzuki, M., Nishihara, M., Takahashi, M. and Iwakura, Y. 1998. Production of mice deficient in genes for interleukin (IL)-1alpha, IL-1beta, IL-1alpha/beta, and IL-1 receptor antagonist shows that IL-1beta is crucial in turpentine-induced fever development and glucocorticoid secretion. J. Exp. Med. 187:1463.[Abstract/Free Full Text]
-
Murphy, K. M., Heimberger, A. B. and Loh, D. Y. 1990. Induction by antigen of intrathymic apoptosis of CD4+CD8+TCRlo thymocytes in vivo. Science 250:1720.[ISI][Medline]
-
Nagai, H., Ueda, Y., Ochi, T., Hirano, Y., Tanaka, H., Inagaki, N. and Kawada, K. 2000. Different role of IL-4 in the onset of hapten-induced contact hypersensitivity in BALB/c and C57BL/6 mice. Br. J. Pharmacol. 129:299.[Abstract/Free Full Text]
-
Wang, B., Amerio, P. and Sauder, D. N. 1999. Role of cytokines in epidermal Langerhans cell migration. J. Leuk. Biol. 66:33.[Abstract]
-
Dieu-Nosjean, M. C., Vicari, A., Lebecque, S. and Caux, C. 1999. Regulation of dendritic cell trafficking: a process that involves the participation of selective chemokines. J. Leuk. Biol. 66:252.[Abstract]
-
Toellner, K. M., Scheel-Toellner, D., Sprenger, R., Duchrow, M., Trumper, L. H., Ernst, M., Flad, H. D. and Gerdes, J. 1995. The human germinal centre cells, follicular dendritic cells and germinal centre T cells produce B cell-stimulating cytokines. Cytokine 7:344.[ISI][Medline]
-
Gorbachev, A. V., Heeger, P. S. and Fairchild, R. L. 2001. CD4+ and CD8+ T cell priming for contact hypersensitivity occurs independently of CD40CD154 interactions. J. Immunol. 166:2323.[Abstract/Free Full Text]
-
Zhang, L. and Tinkle, S. S. 2000. Chemical activation of innate and specific immunity in contact dermatitis. J. Invest. Dermatol. 115:168.[Abstract/Free Full Text]
-
Gosgrove, D., Gray, D., Dierich, A., Kaufman, J., Lemeur, M., Benoist, C. and Mathis, D. 1991. Mice lacking MHC class II molecules. Cell 66:1051.[ISI][Medline]
-
Nakae, S., Asano, M., Horai, R. and Iwakura, Y. 2001. Interleukin (IL)-1ß, but not IL-1
, is required for T cell dependent antibody production. Immunology (in press).