Abstract
We created a murine model of delayed-type hypersensitivity (DTH) to 1-chloro-2, 4-dinitrobenzene (DNCB). Using this murine model, we compared oral mucosal sensitization and skin sensitization for the difference in reaction during the elicitation phase. Evaluation of sensitizability, using the mouse ear swelling test (MEST) after oral mucosal or skin sensitization, showed that the ear swelling response peaked 24 h after challenge. The optimal induction concentration was 1·0% in both oral mucosal and skin sensitization, resulting in a positive reaction rate of 100%. However, the ear swelling response 24 h after challenge with the optimal concentration of DNCB (1·0%) was significantly lower in oral mucosal than in skin sensitization. We compared the oral mucosal and skin sensitization sites for the number of Langerhans’ cells (LC) and the antigen-presenting capability in the induction phase. The numbers of F4/80+ major histocompatibility complex (MHC) class II+ LC before induction did not differ significantly between the oral mucosa and the skin. After induction, F4/80+ MHC class II+ LC increased in number, but the increase was significantly smaller in the oral mucosa than in the skin. MEST on anti-CD86 antibody-administered mice showed that ear swelling was similarly suppressed after oral mucosal or skin sensitization. In murine models of DTH after oral mucosal sensitization, the number of F4/80+CD86+ LC increased after induction, but the increase was significantly smaller than that in murine models of DTH after skin sensitization. This study showed that, in murine models of DTH, oral mucosal sensitization elicited a weaker reaction than skin sensitization. This was presumably because oral mucosal sensitization induced fewer LC, resulting in lower antigen-presenting capability.
Keywords: oral mucosa, DNCB, Langerhans' cells, DTH
INTRODUCTION
The mechanism of delayed type hypersensitivity (DTH) is explained by the concept of skin-associated lymphoid tissue (SALT) in dermatology. After the hapten combines with the carrier protein, the hapten–carrier complex is processed by Langerhans’ cells (LC) or other antigen-presenting cells (APC). In the inductive phase, the LC take antigen from the skin and migrate through lymphoducts to the skin regional lymph nodes, where they, together with major histocompatibility complex class II (MHC class II) molecules, present the antigen to naive T cells [1,2].
Recently the concept of mucosa-associated lymphoid tissue (MALT) has been proposed in the oral mucosa [3,4]. The oral mucosa, like the skin, is subject to allergic sensitization. In general, the oral mucosa is not as readily sensitized as the skin, possibly because the keratin layer of the skin may contain proteins that more readily combine with simple chemicals to form allergen [5]. The investigation of DTH in human oral mucosa with 1-chloro-2, 4-dinitrobenzene (DNCB) indicates that sensitization via the buccal mucosa may increase skin tolerance [6]. In other words, the oral mucosa, in contrast to the skin, is a site where the immune response is inhibited.
Formerly, immune reactivity in the oral mucosa (in particular) and at mucosal surfaces, was mainly studied in terms of T-cell- and/or B-cell-mediated antibody responses in humans and animals [3,4]. Studies in a murine model of DTH to oxazolone (OXA) concluded that the murine buccal mucosa has the capacity to serve as both an inductive and an expression site for local and remote T-cell-mediated DTH reactions [7–9]. A recent study on APC, including LC, has also concluded that MHC class II+ dendritic cells (DC) represent efficient APC of the buccal mucosa [10]. However, few studies have compared the oral mucosa and the skin for their numbers of LC and their function. It is well known that T-cell activation requires the engagement of the T-cell receptor (TCR) with antigen/MHC as well as the engagement of an appropriate costimulatory molecule. The most extensively characterized costimulatory molecules are CD80 and CD86 on professional APC. CD80 and CD86 are expressed on professional APC, such as DC, monocytes and macrophages [11–14]. CD86 is especially strongly involved in the antigen-presentation capability of DTH [13,14].
This study aimed to compare oral mucosal sensitization and skin sensitization for the difference in the reaction elicited during the elicitation phase in murine models of DTH induced by the hapten DNCB [15,16], and to demonstrate the difference in sensitizability by examining the number of LC and CD86+ cells (as an indicator of antigen-presenting capability) in sites of sensitization in the induction phase.
MATERIALS AND METHODS
Mice
Seven-week-old-female BALB/cA Jcl mice (BALB/c mice) were purchased from CLEA Japan Inc. (Tokyo, Japan). All animals were bred at the Tokyo Dental College, Ichikawa General Hospital, research-building animal breeding room. Each mouse weighed ≈20 g, and each experimental group was divided into five groups.
Chemicals
DNCB (Wako Pure Chemicals Inc, Osaka, Japan) was dissolved in a 4 : 1 (v/v) acetone/olive oil solution (A-O).
Monoclonal antibody (MoAb)
Anti-CD86 antibody [clone PO3, rat immunoglobulin (Ig)G2b] [13,14] was used for mouse treatment, and anti-I-A (clone M5/114.15.2, rat IgG2b) and anti-CD86 (clone PO3, rat IgG2b) [14] were used for immunohistochemistry. All of these reagents were purchased from PharMingen (San Diego, CA). Anti-F4/80 (clone CI: A3-1, rat IgG2b) [17] was also used for immunohistochemistry. This reagent was purchased from Cambrex (East Rutherford, NJ). Rat IgG (Sigma Aldrich, Tokyo, Japan) was used for control groups.
Irritation dose–response
It is important to use a challenge dose that causes no measurable irritation at 12, 24, 48 and 72 h. Therefore, the highest non-irritation dose was determined from the irritation dose–response results for DNCB (Table 1). We topically applied DNCB on both sides of the right ear on day 0. The ear skin patches contained 20 µl of DNCB at concentrations of 0·03%, 0·1%, 0·3%, 1·0% or 3·0% (w/v) in A-O. The ear-thickness increase for each mouse (n = 5) was then determined by averaging the swelling measured before, and 12, 24, 48 and 72 h after, application of the ear skin patch. Ear swelling was expressed as the difference in ear thickness before and after challenge in units of 10−2/mm ± standard error (s.e.) [18].
Table 1.
Irritation dose–response results for 1-chloro-2, 4-dinitrobenzene (DNCB)
| Irritation dose (DNCB % v/v) | Mean ear thickness increase (× 10−2 mm, ±s.e.m.) | Maximal ear thickness increase (× 10−2 mm) |
|---|---|---|
| 0·03 | 0·6 | (0·50) 1 |
| 0·10 | 0·6 | (0·90) 2 |
| 0·30 | 0·8 | (0·11) 3 |
| 1·00 | 2·6 | (0·11) 4 |
| 3·00 | 8·6 | (0·26) 10 |
Ear thickness increase was computed by averaging the change from the pre-challenge value at 12 h after exposure.
Oral mucosa and skin sensitization
We sensitized BALB/c mice, without adjuvant, according to Garrigue et al. and Natsuaki et al. [15,18]. We shaved the middle back skin (≈2 cm2), using an electric hair clipper, on day 0. The back skin patches (≈1 cm2) contained 20 µl of DNCB at concentrations of 0·03%, 0·1%, 0·3%, 1·0%, 3·0% or 10·0% (w/v) in A-O. Control-group mice were treated, in the same manner, with A-O only.
For oral mucosa sensitization, we topically applied 20 µl of DNCB at concentrations of 0·03%, 0·1%, 0·3%, 1·0%, 3·0%, or 10·0% (w/v), in A-O, on the left side of dry buccal mucosa (≈1 cm2) on day 0, in the same manner as for the shaved back skin. Control mice were treated in the same manner with A-O only. White Vaseline was applied on the mouth to prevent DNCB or A-O from contacting the skin surrounding the mouth.
On day 5, all mice were challenged by the topical application of 20 µl of 0·3% DNCB onto both sides of the right ear and with 20 µl of A-O onto both sides of the left ear. The ear thickness of mice was measured using a digimatic thickness gauge (Mitutoyo, Kanagawa, Japan) before, and 12, 24, 48 and 72 h after challenge. Ear swelling was expressed as the difference in ear thickness before and after challenge in units of 10−2/mm ± s.e.m. [18].
Antibody treatment
Each group (n = 5) received intraperitoneal (i.p.) injection with 250 µg/mouse of anti-CD86 MoAb diluted with phosphate-buffered saline (PBS), in a total volume of 0·5 ml, 2 h prior to induction [14]. Control mice received rat IgG. The induction dose was 1·0% DNCB and the challenge dose was 0·3% DNCB.
Immunohistochemical techniques
Tissue specimens of the ear skin and buccal mucosa were removed from sensitized or non-sensitized mice 24 h after induction with DNCB. Each biopsy specimen was immediately snap-frozen, embedded in OCT compound (Tissue-tek, Miles, IN) and frozen in liquid nitrogen. Cryostat serial sections of 4-µm thickness were fixed in cold acetone for 10 min. The sections were incubated for 60 min with rat anti-mouse F4/80, MHC class II, or CD86 MoAbs after blocking with 10% normal rabbit serum, and then incubated for 30 min with secondary biotinylated rabbit anti-rat IgG (Vector, Burlingame, CA). Then, sections were treated with peroxidase-conjugated ABC complex (Vectastain ABC; Vector). The sections were developed using diaminobenzidine-H2O2 as a substrate and counterstained with Mayer's haematoxylin.
F4/80+, MHC class II+ and CD86+ cells in serial sections were counted in a blinded manner through a ×400 objective on a light microscope. MHC class II+ and CD86+ cells among F4/80+ DC were counted in the corresponding fields in serial sections. Three fields were examined for each specimen. We analysed the skin from epidermis to dermis and the oral mucosa from the epithelium to lamina propria, corresponding to a surface of 0·0625 mm2. Data were expressed as mean cell count for F4/80+ MHC class II+ cells or F4/80+ CD86+ cells ± s.e.m. per tissue unit. Cell counts were determined on each test animal, and individual cell counts were estimated [8].
Statistical analysis
Statistical analysis was performed using the Student's t-test.
RESULTS
Determination of the challenge dose
Data for the DNCB irritation study are presented in Fig. 1. Five doses were studied for irritation potency: 0·03, 0·1, 0·3, 1·0 and 3·0%. The ear swelling was minimal or non-existent at all time-points, for all doses, except for 1·0 and 3·0% DNCB. As shown in Table 1, the mean ear thickness increase for these five doses, 12 h after exposure, was 0·6, 0·6, 0·8, 2·6 and 8·6 × 10−2 mm, respectively. Primary irritation by up to 0·3% DNCB did not cause any observable ear swelling. Therefore, the highest concentration (0·3%) causing no primary irritation was selected as the challenge concentration, and the greatest increase (≥ 2 × 10−2 mm) in ear thickness caused by 0·3% DNCB was regarded as a positive response.
Fig. 1.
Irritation response time-course for 1-chloro-2, 4-dinitrobenzene (DNCB). Responses from mice (n = 5) were obtained upon induction to five different doses of DNCB. The 0·3% dose was chosen as the challenge concentration for the sensitization studies. Each value represents the mean ± standard error (s.e.m.).
Dose–response and time-course of ear swelling after challenge by oral mucosa sensitization
The time-course data for DNCB sensitization dose–response studies are shown in Fig. 2(a). Mice sensitized with 0·1, 0·3, 1·0 and 3·0% DNCB showed maximal ear swelling 24 h after challenge with 0·3% DNCB. All mice sensitized with 10·0% DNCB died 1–2 days after challenge(Table 2). Mice sensitized with 0·03% DNCB showed no ear swelling at any time-point. Ear swelling decreased at 72 h after challenge with six sensitization doses. The DTH induction with DNCB was optimal at an induction dose of 1·0%, as the positive-reaction rates with a 1·0% induction dose were 100% (Table 2).
Fig. 2.
The time-course of ear swelling of 1-chloro-2, 4-dinitrobenzene (DNCB)-sensitized mice. Responses upon induction to six different sensitization doses were evaluated, in addition to the response of the control group (0%) that was given acetone/olive oil (n = 5). All groups of mice were challenged with 0·3% DNCB on day 5. (a) One sensitizing application was given on the buccal mucosa; (b) one sensitizing application was given on the back skin. Each value represents the mean ± standard error (s.e.m.).
Table 2.
The positive-response ratios to 1-chloro-2, 4-dinitrobenzene (DNCB), as assessed by the mouse ear swelling test (MEST)
| Sensitization dose (%) | |||||||
|---|---|---|---|---|---|---|---|
| A-0 | 0·03 | 0·1 | 0·3 | 1·0 | 3·0 | 10·0 | |
| Oral sensitization | 0/5 | 0/5 | 1/5 | 4/5 | 5/5 | 3/5 | Died |
| Skin sensitization | 0/5 | 0/5 | 2/5 | 2/5 | 5/5 | 4/5 | 2/5 |
All mice were challenged by the topical application of 20 µl of 0·3% DNCB. Each group consisted of five mice.
Dose–response and time-course of ear swelling after challenge by skin sensitization
The time-course data for DNCB sensitization dose–response studies are shown in Fig. 2(b). Maximal ear swelling was observed 24 h after challenge with 1·0, 3·0 or 10·0% DNCB. Ear swelling decreased at 72 h after challenge, at all doses of DNCB. The DTH induction with DNCB was optimal at an induction dose of 1·0%. The positive-reaction rates with a 1·0% induction dose were 100% (Table 2).
Response curve at 24 h after challenge
The response curve at 24 h after challenge is shown in Fig. 3. Three types of dose–response were identified: a low-dose no-effect at concentrations of <0·1% DNCB; a positive dose–response at concentrations of 0·1–1·0% DNCB; and a high-dose down-regulated response at concentrations of >1·0% DNCB.
Fig. 3.
Sensitization dose–response for 1-chloro-2, 4-dinitrobenzene (DNCB). A dose–response range was observed at DNCB concentrations from 0 to 1·0%, and a high-dose reduced-response range was observed at DNCB concentrations of >1·0%. (a) One sensitizing application was given on the buccal mucosa and (b) one sensitizing application was given on the back skin, and the ear thickness was evaluated 24 h later. Data points are group means and error bars represent the standard error (s.e.m.).
Ear swelling after challenge in anti-CD86 MoAb-treated mice
Anti-CD86 MoAb treatment before induction completely inhibited ear swelling by both oral mucosa sensitization and skin sensitization (Fig. 4).
Fig. 4.
The time-course of delayed-type hypersensitivity (DTH) to 1-chloro-2, 4-dinitrobenzene (DNCB) in BALB/cA mice with or without anti-CD86 monoclonal antibody (MoAb) treatment. (a) One sensitizing application was given on the buccal mucosa and (b) one sensitizing application was given on the back skin. All groups of mice were challenged with 0·3% DNCB on day 5 (n = 5) and the ear thickness was evaluated 24 h later. Each value represents the mean ± standard error (s.e.m.).
MHC class II-expressing LC (F4/80+ MHC class II+ cells) in the oral mucosa and skin
Before induction, MHC class II-expressing LC were detected in the oral mucosa, mainly from the intermediate layer to the basal layer of the oral epithelium (Fig. 5c). In the skin, MHC class II-expressing LC were detected in the basal layer of the epidermis, as well as in the shallow layer of the dermis (Fig. 5d). After induction, the numbers of MHC class II-expressing LC were clearly increased in the suprabasal layer of oral mucosa (Fig. 5i). In the skin, most of these cells were found in the dermis (Fig. 5j). Regardless of induction site, the numbers of MHC class II-expressing LC increased significantly after induction, as compared with before induction (Table 3). However, the number of MHC class II-expressing LC in the oral mucosa, after induction, was significantly smaller than that in the skin (Table 3).
Fig. 5.
Expression of F4/80 (a, b, g, h), major histocompatibility complex (MHC) class II (c, d, i, j), and CD86 (e, f, k, l), in the buccal mucosa (a, c, e, g, i, k) and in the ear skin (b, d, f, h, j, l) sections from mice 24 h before (a–f) or after (g–l) sensitization with 1-chloro-2, 4-dinitrobenzene (DNCB). (Original magnification: 200×); the inset depicts F4/80-expressing Langerhans’ cells (LC) in such an infiltrate (original magnification 400×).
Table 3.
Comparison of the cell density of F4/80+ major histocompatibility complex (MHC) class II+ cells or F4/80+ CD86+ cells between the normal non-sensitized) oral mucosa and skin and in the sensitized oral mucosa and skin
| F4/80+ MHC class II+ | F4/80+ CD86+ | |
|---|---|---|
| Non-sensitized | ||
| Oral mucosa | 3·7 ± 1·5 | 3·3 ± 1·5 |
| Skin | 5·3 ± 1·5 | 4·7 ± 1·8 |
| Sensitized | ||
| Oral mucosa | 28·7 ± 1·9 | 13·7 ± 1·9 |
| Skin | 39·7 ± 1·5* | 35·3 ± 3·9* |
Five mice were used in each group and a minimum of three random fields was examined for each mouse. The field size was 0·0625 mm2 (400× magnification). Data are expressed as mean numbers of F4/80+ MHC class II+ cells or F4/80+ CD86+ cells ± standard error of the mean (s.e.m.) per field.
CD86-expressing LC (F4/80+ CD86+ cells) in the oral mucosa and skin
CD86-expressing LC were not observed in the oral mucosa and skin before induction (Fig. 5e, 5f). After induction, the numbers of CD86-expressing LC in the oral mucosa increased, and most of these cells were observed in the suprabasal layer (Fig. 5k). After induction, most CD86-expressing LC were found in the dermis; in particular, strongly positive cells were observed in the upper layer of the dermis (Fig. 5l). Overall, the numbers of CD86-expressing LC in the oral mucosa and skin increased significantly after induction as compared with before induction (Table 3). However, after sensitization, the numbers of CD86-expressing LC were significantly lower in the mucosa than in the skin (Table 3).
DISCUSSION
We created a murine model of DTH to DNCB to evaluate and compare the oral mucosa and the skin for sensitizability to DNCB. Among many induction methods, the experimental method of Natsuaki et al. [18] was used to induce skin DTH by painting the skin on the back of mice with a single dose of DNFB and evaluating the response by MEST. This was because painting the same site with antigen was reported to suppress skin DTH [19,20], and because repeated painting with antigen was reported to induce immediate-type hypersensitivity, but not DTH [21]. The MEST showed that the optimal induction concentration was 1% DNCB for both the oral mucosa and the skin (Fig. 2a, 2b). However, the ear swelling was lower with oral mucosal sensitization as compared with skin sensitization. At a higher concentration of 3·0 and 10·0%, however, the ear swelling decreased, irrespective of the site of sensitization, which indicates suppressed sensitization (Fig. 3a, 3b). In one study using DNCB, the amount of antigen per unit area (antigen concentration) was shown to be the major determinant in sensitization [22]. Accordingly, the ear swelling may be suppressed by antigen administered at high concentrations. In another sensitization study with DNCB, animals were rated as positive for sensitization if their epidermis showed reddening and swelling, and negative if their epidermis showed damage with suppressed inflammatory reaction [23]. This finding was attributed to the possible failure of sensitization or suppression of reactions owing to injury of LC at the induction site. Therefore, suppression of the ear swelling may be caused by tissue damage at 3·0% and 10·0% concentrations of DNCB.
T-cell activation by antigen-presenting LC requires stimulation with the antigen receptor MHC class II on LC (first signal). This failure, in turn, causes functional inactivation without apoptosis, in the form of the inability to respond to stimulation by the same antigen (clonal anergy). The study of Nuriya et al., using a DTH mouse model, reported that MHC class II-expressing LC clearly increased in the epidermis and dermis after induction [14]. Our study also showed that, after induction, the number of MHC class II-expressing LC clearly increased in the oral mucosa as well as in the skin (Table 3).
CD86 molecules expressed on skin LC in DTH have drawn attention because they are strongly correlated to the cell's antigen-presentation capability [11–14]. However, in LC under normal conditions, CD86 expression is subject to considerable limitations in terms of time and place [24]. Nuriya et al. [14] reported that almost no expression of CD86 is observed in normal skin tissue. In our immunohistochemical investigation, almost no CD86 was detected in normal skin or oral mucosal (Table 3). Nuriya et al. also reported that CD86-expressing LC clearly increased in the sensitized epidermis and dermis. This study also showed that, after induction, the number of CD86-expressing LC clearly increased in the oral mucosa as well as in the skin (Table 3). Nuriya et al. [14] hypothesized that the enhanced expression of CD86 on LC after sensitization, and the significant suppression of DTH by administration of anti-CD86, were functionally mediated by the selective expression of CD86 in LC. Aiba et al. [25] proposed that DNCB-treated LC in vitro showed clearly increased selective expression of CD86, which could be suppressed by treatment with anti-CD86. In the present study, CD86 was expressed in the oral mucosa after sensitization, such as in the skin after skin sensitization (Fig. 5k, 5l). In addition, administration of anti-CD86 significantly suppressed DTH after oral mucosal sensitization (Fig. 4).
In a DTH murine model, oral mucosal sensitization elicited a weaker reaction than skin sensitization, presumably because of the differences in the number of LC and in antigen-presenting capability between the oral mucosa and the skin. Hutchens found fewer LC in the normal monkey oral mucosa than in the normal skin, suggesting that the oral mucosa shows a form of tolerance [26]. However, histological examination in our mouse model showed that the number of LC in the normal oral mucosa did not differ significantly from that in the normal skin (Table 3), which is in agreement with the results of van Loon, showing no significant difference in the number of LC between the normal human oral mucosa and skin [27]. However, the number of LC after induction in the oral mucosa was significantly smaller than that in the skin (Table).
Next, we investigated the expression of CD86 as an indicator of antigen-presenting capability. The numbers of CD86-expressing LC before induction did not differ significantly between the oral mucosa and the skin. However, after induction, the number of CD86-expressing LC was significantly smaller in the oral mucosa than in the skin (Table 3). These results suggest that, in a DTH murine model, oral mucosal sensitization elicited a weaker reaction than skin sensitization because the number of LC and antigen-presenting capability were smaller in the oral mucosa, as a site of induction, than in the skin, as a site of induction.
Unlike the skin, the oral mucosa, as in MALT, is considered to show immune suppression [3,28]. The findings in this study, that the number of LC and antigen-presenting capability in the oral mucosa in the induction phase were decreased, may be associated with the clonal anergy of tolerance.
In conclusion, our study confirms that oral mucosal sensitization can be an inductive site of DTH with DNCB. However, the DTH reaction was lower with oral mucosal sensitization as compared with skin. This study showed that oral mucosal sensitization elicited a lower reaction than skin sensitization, presumably because oral mucosal sensitization induced a smaller number of LC with decreased antigen-presenting capability in comparison with skin sensitization.
Acknowledgments
I would like to thank Dr Sumie Yamanaka and Dr Kaoru Ohta for valuable and technical advice during the pilot phase of this study.
REFERENCES
- 1.Sauder DN. Allergic contact dermatitis. In: Bruce H, Thiers MD, editors. Pathogenesis of Skin Disease. 1. New York: Churchill Livingstone; 1986. pp. 3–12. [Google Scholar]
- 2.Streilein JW. Skin-associated lymphoid tissues (SALT): origins and functions. J Invest Dermatol. 1983;80:12s–16s. doi: 10.1111/1523-1747.ep12536743. [DOI] [PubMed] [Google Scholar]
- 3.Brandtzaeg P. History of oral tolerance and mucosal immunity. Ann N Y Acad Sci. 1996;778:1–27. doi: 10.1111/j.1749-6632.1996.tb21110.x. [DOI] [PubMed] [Google Scholar]
- 4.Brandtzaeg P. Overview of the mucosal immune system. Curr Top Microbiol Immunol. 1987;146:13–25. doi: 10.1007/978-3-642-74529-4_2. [DOI] [PubMed] [Google Scholar]
- 5.Robert LR, Joseph FF. Contact stomatitis and cheilitis. In: Jonathan WP, editor. Fisher's Contact Dermatitis. 4. Maryland: Williams & Wilkins; 1995. pp. 886–919. [Google Scholar]
- 6.Lowney ED. Unresponsiveness to a contact sensitizer in man. J Invest Dermatol. 1968;51:411. [PubMed] [Google Scholar]
- 7.Ahlfors E, Czerkinsky C. Contact sensitivity in the murine oral mucosa. I. An experimental model of delayed-type hypersensitivity reactions at mucosal surfaces. Clin Exp Immunol. 1991;86:449–56. doi: 10.1111/j.1365-2249.1991.tb02952.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 8.Ahlfors E, Jonsson R, Czerkinsky C. Experimental T cell mediated inflammatory reactions in the murine oral mucosa. II. Immunohistochemical characterization of resident and infiltrating cells. Clin Exp Immunol. 1996;104:297–305. doi: 10.1046/j.1365-2249.1996.967659.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 9.Ahlfors E, Lyberg T. Contact sensitivity reactions in the oral mucosa. Acta Odontol Scand. 2001;59:248–54. doi: 10.1080/00016350152509283. [DOI] [PubMed] [Google Scholar]
- 10.Eriksson K, Ahlfors E, George-Chandy A, et al. Antigen presentation in the murine oral epithelium. Immunology. 1996;88:147–52. doi: 10.1046/j.1365-2567.1996.d01-647.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 11.Hathcock KS, Laszlo G, Pucillo C, et al. Comparative analysis of B7-1 and B7-2 costimulatory ligands: expression and function. J Exp Med. 1994;180:631–40. doi: 10.1084/jem.180.2.631. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 12.Inaba K, Witmer-Pack M, Inaba M, et al. The tissue distribution of the B7-2 costimulator in mice: abundant expression on dendritic cells in situ and during maturation in vitro. J Exp Med. 1994;180:1849–60. doi: 10.1084/jem.180.5.1849. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 13.Nakajima A, Azuma M, Kodera S, et al. Preferential dependence of autoantibody production in murine lupus on CD86 costimulatory molecule. Eur J Immunol. 1995;25:3060–9. doi: 10.1002/eji.1830251112. [DOI] [PubMed] [Google Scholar]
- 14.Nuriya S, Yagita H, Okumura K, et al. The differential role of CD86 and CD80 co-stimulatory molecules in the induction and the effector phases of contact hypersensitivity. Int Immunol. 1996;8:917–26. doi: 10.1093/intimm/8.6.917. [DOI] [PubMed] [Google Scholar]
- 15.Garrigue JL, Nicolas JF, Fraginals R, et al. Optimization of the mouse ear swelling test for in vivo and in vitro studies of weak contact sensitizers. Contact Dermatitis. 1994;30:231–7. doi: 10.1111/j.1600-0536.1994.tb00650.x. [DOI] [PubMed] [Google Scholar]
- 16.Gad SC, Dunn BJ, Dobbs DW, et al. Development and validation of an alternative dermal sensitization test: the mouse ear swelling test (MEST) Toxicol Appl Pharmacol. 1986;84:93–114. doi: 10.1016/0041-008x(86)90419-9. [DOI] [PubMed] [Google Scholar]
- 17.Kraal G, Rep M, Janse M. Macrophages in T and B cell compartments and other tissue macrophages recognized by monoclonal antibody MOMA-2; an immunohistochemical study. Scand J Immunol. 1987;26:653–61. doi: 10.1111/j.1365-3083.1987.tb02301.x. [DOI] [PubMed] [Google Scholar]
- 18.Yamashita N, Natsuaki M, Sagami S. Flare-up reaction on murine contact hypersensitivity. Immunology. 1989;67:365–9. [PMC free article] [PubMed] [Google Scholar]
- 19.Nakano Y. Antigenic competition in the induction of contact sensitivity in mice. Immunology. 1977;33:167–78. [PMC free article] [PubMed] [Google Scholar]
- 20.Nakano K, Nakano Y. Suppressor cells in antigenic competition in contact allergy in mice. Immunology. 1978;34:981–7. [PMC free article] [PubMed] [Google Scholar]
- 21.Natsuaki M, Yano N, Yamaya K, et al. Immediate contact hypersensitivity induced by repeated hapten challenge in mice. Contact Dermatitis. 2000;43:262–72. doi: 10.1034/j.1600-0536.2000.043005267.x. [DOI] [PubMed] [Google Scholar]
- 22.Friedmann PS. The immunology of allergic contact dermatitis: the DNCB story. Adv Dermatol. 1990;5:175–95. [PubMed] [Google Scholar]
- 23.Bergstresser PR, Toews GB, Streilein JW. Natural and perturbed distributions of Langerhans’ cells: responses to ultraviolet light, heterotopic skin grafting, and dinitrofluorobenzene sensitization. J Invest Dermatol. 1980;75:73–7. doi: 10.1111/1523-1747.ep12521261. [DOI] [PubMed] [Google Scholar]
- 24.Simon J, Dietrich A, Mielke V, et al. Expression of the B7/BB1 antigen and its ligand CD28 in T-cell-mediated skin diseases. J Invest Dermatol. 1994;103:539–43. doi: 10.1111/1523-1747.ep12395743. [DOI] [PubMed] [Google Scholar]
- 25.Aiba S, Terunuma A, Manome H, et al. Dendritic cells differently respond to haptens and irritants by their production of cytokines and expression of co- stimulatory molecules. Eur J Immunol. 1997;27:3031–8. doi: 10.1002/eji.1830271141. [DOI] [PubMed] [Google Scholar]
- 26.Hutchens L. Oral epithelial dendric cells of the rhesus monkey: histologic demonstration, fine structure and quantitative destruction. J Invest Dermatol. 1971;56:325–36. doi: 10.1111/1523-1747.ep12261091. [DOI] [PubMed] [Google Scholar]
- 27.van Loon LAJ, Krieg SR, Davidson CL, Bos JD. Quantification and distribution of lymphocyte subsets and Langerhans’ cells in normal human oral mucosa and skin. J Oral Pathol Med. 1989;18:197–201. doi: 10.1111/j.1600-0714.1989.tb00762.x. [DOI] [PubMed] [Google Scholar]
- 28.Brandtzaeg P, Farstad IN, Haraldsen G, Jahnsen FL. Cellular and molecular mechanisms for induction of mucosal immunity. In: Brown F, Haaheim LR, editors. Modulation of Immune Response to Vaccine Antigens. Vol. 92. Basel: Karger; 1998. pp. 93–108. Dev Biol Stand. [PubMed] [Google Scholar]