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* Department of Internal Medicine II, Chiba University School of Medicine, Chiba, Japan; and
Medicine and Biosystemic Science, Graduate School of Medical Sciences, and
Department of Molecular and Cellular Biology, Medical Institute of Bioregulation, Kyushu University, Fukuoka, Japan
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Abstract |
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 Top Abstract IntroductionMaterials and MethodsResultsDiscussionReferences |
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and
IL-12. Tyk2 is a member of the Janus family of protein tyrosine kinases
and is activated by a variety of cytokines: IFN-
, IL-6, IL-10,
IL-12, and IL-13. In this study, we investigated the role of Tyk2 in
the regulation of Ag-induced Th cell differentiation and Ag-induced
allergic inflammation in the airways using Tyk2-deficient
(Tyk2-/-) mice. When splenocytes were stimulated with
antigenic peptide, IL-12-mediated Th1 cell differentiation was
decreased, but IL-4-mediated Th2 cell differentiation was increased in
Tyk2-/- mice. In vivo, Ag-specific IgE and IgG1
production was increased, but Ag-specific IgG2a production was
decreased in Tyk2-/- mice as compared with those in
control mice. In addition, Ag-induced eosinophil and CD4+ T
cell recruitment, as well as the production of Th2 cytokines in the
airways, was increased in Tyk2-/- mice. Adoptive transfer
experiments revealed that CD4+ T cells were responsible for
the enhanced Ag-induced eosinophil recruitment in Tyk2-/-
mice. In contrast, although the level of IL-13 was increased in the
airways of Tyk2-/- mice after Ag inhalation, the number
of goblet cells, as well as Muc5ac mRNA
expression, was decreased in Tyk2-/- mice. Together,
these results indicate that Tyk2 plays a bilateral role in the
regulation of allergic inflammation in the airways: Tyk2 plays a role
in the down-regulation of Th2 cell-mediated Ab production and
eosinophil recruitment in the airways by regulating Th1/Th2 balance
toward Th1-type, while Tyk2 is necessary for the induction of
IL-13-mediated goblet cell hyperplasia in the
airways. |
Introduction |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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(7, 8) and IL-12
(9, 10) down-regulate Ag-induced allergic inflammation in
the airways. Taken together, these observations suggest that Ag-induced
allergic inflammation is regulated by the balance between Th1 and Th2
cells, in which Th2 cells up-regulate but Th1 cells down-regulate the
allergic inflammation by producing cytokines.
The receptor binding of cytokines results in the activation of the
Janus family of protein tyrosine kinases (Janus kinases
(Jaks))3 and
subsequently the activated Jaks phosphorylate signal transducers and
activators of transcription (Stats) (11, 12). There are
four mammalian Jaks, Jak1, Jak2, Jak3, and Tyk2, and they are
differentially activated in response to various cytokines (11, 12). Tyk2 has been initially identified as an essential molecule
for mediating IFN- signaling (13) and subsequently
shown to be activated in response to IL-6 (14), IL-10
(15), IL-12 (16), and IL-13
(17). Recently, to address the specific and nonredundant
role of Tyk2, mutant mice lacking Tyk2 (Tyk2-/-
mice) were generated by homologous recombination, and using
Tyk2-/- mice, it has been demonstrated that
Tyk2 is not essential for many of the biological responses upon
IFN-, IL-6, and IL-10 stimulation (18, 19). In
contrast, Tyk2 has been shown to be required for IL-12-induced IFN-
production in activated T cells (18, 19). However, the
role of Tyk2 in in vivo immune responses to exogenous Ags is
unknown.
To determine the role of Tyk2 in immune responses to exogenous Ags, we examined Ag-specific Ab production and Ag-induced allergic inflammation in the airways in Tyk2-/- mice. We also examined the role of Tyk2 in Ag-induced Th cell differentiation using TCR transgenic Tyk2-/- mice. We found that immune responses to Ag were biased toward Th2-type in Tyk2-/- mice in vitro as well as in vivo. Consequently, Ag-specific IgE production and Ag-induced eosinophil recruitment into the airways were enhanced in Tyk2-/- mice. These results indicate that Tyk2 plays an important role in regulating Th1/Th2 balance toward Th1-type and thus down-regulating Th2 cell-mediated allergic inflammation.
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Materials and Methods |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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As the degree of Ag-induced eosinophil recruitment into the airways differs depending on the genetic background of the mice (20), Tyk2-deficient mice (Tyk2-/- mice; Ref. 18) were backcrossed for >4 generations onto BALB/c mice (Charles River Laboratories, Atsugi, Japan). All mice were H-2d/d and littermate wild-type (WT) mice were used as controls. Backcrossing onto BALB/c mice for four generations results in responses that are indistinguishable to those seen in BALB/c mice in our assays (Ref. 21 and our unpublished data). OVA-specific DO11.10 (DO10+) TCR transgenic mice (22) were backcrossed over ten generations onto BALB/c mice and then crossed with Tyk2-/- mice. DO10+Tyk2-/- mice (H-2d/d) and the littermate DO10+ mice (H-2d/d) were analyzed in the following experiments. Mice were housed in microisolator cages under pathogen-free conditions. All experiments were performed according to the guidelines of Chiba University.
Immunization
Mice (aged 7–8 wk) were immunized i.p. twice with 4 µg of OVA (Sigma-Aldrich, St. Louis, MO) in 4 mg of aluminum hydroxide (alum) at a 2-wk interval. As a control, mice were injected i.p. with 4 mg of alum alone.
Ag-induced allergic inflammation in the airways
Allergic airway inflammation was induced by the inhalation of OVA in sensitized mice as described previously (3). Briefly, 12–14 days after the second immunization, the sensitized mice were given aerosolized OVA (50 mg/ml) dissolved in 0.9% saline by a DeVilbiss 646 nebulizer (DeVilbiss, Somerset, PA) for 20 min. OVA solution contained <1 ng/ml endotoxin. As a control, 0.9% saline alone was administered by the nebulizer. In experiments shown in Figs. 3B and 6, the sensitized mice were given aerosolized OVA (50 mg/ml) three times at a 24-h interval. At indicated times after the inhalation, trachea and a sagittal block of right lung were excised, fixed in 10% buffered-formalin, and embedded in paraffin. The specimens (3-µm thick) of the trachea were stained with Luna and H&E solutions. The number of eosinophils in the submucosal tissue of trachea was counted in Luna-stained sections and expressed as the number of eosinophils per the length of the basement membrane of trachea, which was measured with a digital curvimeter. Lung sections were stained with H&E and periodic acid-Schiff (PAS) according to standard protocols. A semiquantitative scoring system was used to grade the magnitude of inflammatory cell infiltration in the perivascular and peribronchial spaces on H&E-stained lung sections as described previously (23). Goblet cells were counted on PAS-stained lung sections using a scoring system as described elsewhere (6).
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Cytokine levels in BALF
The amounts of IL-4, IL-5, IFN-, and TNF- in the BALF were
determined by the enzyme immunoassay using murine IL-4, IL-5, IFN-,
and TNF- ELISA kits from BD PharMingen (San Diego, CA). The amounts
of IL-13 in the BALF were determined by ELISA kit from R&D Systems
(Minneapolis, MN). The assays were performed in duplicate according to
the manufacturers’ instruction. The detection limits of these assays
were 15 pg/ml IL-4 and IL-5, 50 pg/ml IFN- and TNF-, and 30 pg/ml
IL-13.
Determination of Ag-specific IgE, IgG1, and IgG2a Abs in serum
Two weeks after the second immunization, the titer of OVA-specific IgE Ab in mouse serum was assessed by a 24-h passive cutaneous anaphylaxis reaction as described previously (21). The amount of OVA-specific IgG1 and IgG2a Abs in serum was measured by ELISA as described previously (21).
Flow cytometric analysis
Cells from the BALF and spleen were stained and analyzed on a FACSCalibur (BD Biosciences, San Jose, CA) using CellQuest software. For direct staining, the following conjugated Abs were purchased from BD PharMingen: anti-CD4 FITC, PE, APC (H129.19), anti-CD8 PE (53.6.7), and anti-B220 FITC (RA3-6B2). Before staining, FcRs were blocked with anti-CD16/32 Ab (2.4G2; BD PharMingen).
Measurement of airway reactivity
Two weeks after the second immunization, sensitized mice were challenged with the inhaled OVA three times at a 24-h interval. Twenty-four hours after the final OVA challenge, airway reactivity to aerosolized methacholine (3–50 mg/ml) was measured using the Buxco whole body plethysmograph (Buxco Electronics, Sharon, CT) as described by Hamelmann et al. (24).
RT-PCR
Total RNA was isolated from lung tissues using Isogen solution
(Nippon Gene, Tokyo, Japan), and RT-PCR for Muc5ac mRNA was
performed using primer pairs as described elsewhere (25).
RT-PCR for -actin mRNA was also performed to control the
sample-to-sample variation in RNA isolation and integrity, RNA input,
and reverse transcription. All PCR amplifications were performed at
least three times with multiple sets of experimental RNAs.
Adoptive transfer experiments for Ag-induced eosinophil and T cell infiltration in the airways
Splenocytes from OVA-sensitized Tyk2-/- mice and littermate WT mice were stimulated with OVA (200 µg/ml) at 37°C for 72 h. After dead cells were removed by centrifugation on Ficoll-Paque (Amersham Pharmacia Biotech, Piscataway, NJ), cells (1 x 107 cells/mouse) were transferred to nonimmunized BALB/c SCID mice. The frequency of cell populations of transferred cells was 60–70% of CD4+ T cells, 10–15% of CD8+ T cells, and 20–25% of B220+ cells, and no significant difference was observed between WT mice and Tyk2-/- mice. In some experiments, CD4+ T cells were removed by magnetic cell sorting using FITC-labeled anti-CD4 Ab (RM4-5; BD PharMingen) and anti-FITC magnetic microbeads (Miltenyi Biotec, Sunnyvale, CA), and the CD4+ T cell-depleted populations were transferred to BALB/c SCID mice. These mice were then challenged with inhaled OVA (50 mg/ml) at 0, 24, and 48 h after the cell transfer, and at 24 h after the last OVA inhalation, the number of eosinophils and CD4+ T cells in the BALF was evaluated as described above. In preliminary experiments, we found that the in vitro Ag stimulation of splenocytes was required for the induction of Ag-induced eosinophil recruitment in the airways.
Ag-induced T cell differentiation
Splenocytes (1 x 106/ml) from
DO10+Tyk2-/- mice and the
littermate DO10+ mice were stimulated with
OVA323–339 peptide (50 µM) in a 24-well
microtiter plate at 37°C for 48 h. Where indicated, IL-12 (7.5
ng/ml; R&D Systems) was added to polarize toward Th1 cells (Th1
condition), and IL-4 (7.5 ng/ml; R&D Systems) was added to polarize
toward Th2 cells (Th2 condition). Cells were washed with PBS and
cultured for another 3 days in Th0 (nonpolarizing), Th1, or Th2
condition in the presence of IL-2 (5 ng/ml). Intracellular cytokine
analyses for IL-4 vs IFN- were performed as described previously
(26).
Data analysis
Data are summarized as mean ± SD. The statistical analysis of the results was performed by the unpaired t test. Values of p < 0.05 were considered significant.
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Results |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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It has been demonstrated that IL-4 and IL-12 mediate Th2 and Th1
cell differentiation, respectively (27, 28). IL-4
stimulation results in the activation of Jak1 and Jak3 and subsequently
in the phosphorylation of Stat6, whereas IL-12 activates Jak2 and Tyk2
and then phosphorylates Stat4 (11, 12). Although the
importance of Stat4 and Stat6 in the differentiation of Th1 cells and
Th2 cells is well documented (27), the roles of Jaks in
the Th cell differentiation are still largely unknown. To address the
role of Tyk2 in Ag-induced Th cell differentiation,
Tyk2-/- mice were crossed with OVA-specific
DO10+ TCR transgenic mice, and Ag-induced Th cell
differentiation in
DO10+Tyk2-/- mice was
compared with that in the littermate DO10+ mice.
As shown in Fig. 1, when splenocytes from
DO10+Tyk2-/- mice were
stimulated with antigenic peptide (OVA323–339)
in nonpolarizing Th0 condition, Th2 cells
(IL-4+IFN-- cells) were
increased as compared with those in DO10+ mice
(DO10+Tyk2-/- mice,
11.2 ± 3.0%, vs DO10+ mice, 5.8 ±
2.2%, mean ± SD; n = 5 mice in each group;
p < 0.01) (Fig. 1, A and B).
Moreover, IL-4 and IL-5 levels were increased in the culture
supernatant in
DO10+Tyk2-/- mice (data
not shown). In contrast, the number of Th1 cells
(IL-4-IFN-+ cells) was
not affected by the absence of Tyk2 in Th0 condition. However,
interestingly, when IL-12 was added to the culture (Th1 condition),
IL-12-mediated Th1 cell differentiation was significantly reduced in
DO10+Tyk2-/- mice
(DO10+Tyk2-/- mice,
18.3 ± 4.5%, vs DO10+ mice, 55.3 ±
8.3%; n = 5; p < 0.001) (Fig. 1, C vs D). The finding of reduced IL-12-mediated
Th1 cell differentiation is consistent with a previous finding using
anti-TCR Ab-stimulated CD4+ T cells in
Tyk2-/- mice (18). Furthermore, in
the presence of IL-4 (Th2 condition), the number of Th2 cells in
DO10+Tyk2-/- mice was
more increased than that in DO10+ mice
(DO10+Tyk2-/- mice,
34.6 ± 5.9%, vs DO10+ mice, 19.8 ±
4.2%; n = 5; p < 0.01) (Fig. 1, E and F). Taken together, these results indicate
that Tyk2 regulates Th1/Th2 balance toward Th1-type during Ag-induced
Th cell differentiation.
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Because Th cell differentiation is biased toward Th2-type in Tyk2-/- mice in vitro (Fig. 1), we next attempted to determine whether Tyk2 regulates in vivo Th1/Th2 response to exogenous Ag. Given that Ig class-switching is regulated by cytokines from Th cells (29, 30), we analyzed Ag-specific Ab production in Tyk2-/- mice. Tyk2-/- mice and the littermate WT mice were immunized i.p. twice with OVA in alum at a 2-wk interval, and 14 days after the second immunization, the levels of Ag-specific IgE, IgG1, and IgG2a in sera were evaluated. In agreement with the enhanced Th2 cell differentiation by the absence of Tyk2 (Fig. 1), Tyk2-/- mice exhibited increased levels of OVA-specific IgE and IgG1 production as compared with those in WT mice (n = 10; p < 0.02 and p < 0.01, respectively) (Fig. 2). In contrast, OVA-specific IgG2a production was decreased in Tyk2-/- mice (n = 10, p < 0.01) (Fig. 2). These results suggest that systemic immune response to exogenous Ag is biased toward Th2-type in Tyk2-/- mice.
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Next, we evaluated Ag-induced eosinophil recruitment into the airways, another characteristic of Th2 cell-mediated immune response (2, 3, 4), in Tyk2-/- mice. OVA-sensitized Tyk2-/- mice and littermate WT mice were challenged with inhaled OVA, and eosinophil recruitment into the submucosal tissue of trachea was evaluated at 24 h after Ag inhalation. As shown in Fig. 3A, Ag-induced eosinophil recruitment into the trachea was significantly increased by 97% in Tyk2-/- mice (Tyk2-/- mice, 34.8 ± 6.3, vs WT mice, 17.7 ± 3.5 eosinophils/mm; n = 8 mice in each group; p < 0.01). The inhaled OVA challenge did not significantly induce eosinophil recruitment into the trachea in either unsensitized WT mice or Tyk2-/- mice (Fig. 3A). Ag-induced eosinophil recruitment into the trachea at 48 h after Ag inhalation was also significantly increased in Tyk2-/- mice (data not shown). Histological analysis showed that inflammatory cell infiltration in the lung was also significantly enhanced in Tyk2-/- mice after Ag inhalation as compared with that of WT mice (Fig. 3B). As anticipated, without the inhaled Ag challenge, no inflammatory cell infiltration was observed in either WT mice or Tyk2-/- mice (Fig. 3B). To determine which cell type infiltrates in the airways of Tyk2-/- mice, cell populations in BALF of sensitized mice were determined after Ag inhalation (Fig. 3C). Consistent with the results shown in Fig. 3A, the number of eosinophils recovered in BALF at 36 h after Ag inhalation was significantly increased in Tyk2-/- mice (Tyk2-/- mice, 38.4 ± 7.7, vs WT mice, 25.1 ± 5.4 x 104 cells/mice; n = 8 mice in each group; p < 0.01) (Fig. 3C). The number of lymphocytes recovered in BALF was also increased by 117% in Tyk2-/- mice (n = 8, p < 0.005) (Fig. 3C). FACS analysis revealed that the majority of lymphocytes in BALF were CD4+ T cells, and consequently the number of CD4+ T cells was increased by 110% in Tyk2-/- mice (Fig. 3C; p < 0.005). In contrast, the number of neutrophils or macrophage in the BALF was not affected by the absence of Tyk2 (Fig. 3C). These results indicate that Tyk2 plays a role in the down-regulation of Ag-induced eosinophil and CD4+ T cell recruitment into the airways.
We then examined cytokine levels in the BALF of sensitized
Tyk2-/- mice and WT mice after Ag inhalation.
IL-4 and IL-5 levels in the BALF at 36 h after Ag challenge were
increased in Tyk2-/- mice by 50 and 152%,
respectively (n = 5; p < 0.05 and
p < 0.01, respectively) (Table I). IL-13 levels were also increased in
the BALF of Tyk2-/- mice, by 99%
(n = 5, p < 0.01) (Table I). In
contrast, IFN- and TNF- levels were indistinguishable between WT
mice and Tyk2-/- mice (Table I). As expected,
the inhaled Ag challenge did not significantly induce cytokine
production in the airways in unsensitized WT mice or
Tyk2-/- mice (data not shown). Taken together,
these results suggest that Tyk2 plays a role in the down-regulation of
Ag-induced Th2 cell differentiation in vivo and that the enhanced
Ag-induced eosinophil recruitment in Tyk2-/-
mice results in part from the enhanced IL-5 production in the
airways.
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negatively
regulates Ag-induced eosinophil recruitment into the airways (7, 8). To exclude the possibility that the enhanced eosinophil
recruitment into the airways in Tyk2-/- mice
results from the defect in IFN- production, we examined the effect
of rIFN- on Ag-induced eosinophil recruitment into the airways in
Tyk2-/- mice. Consistent with a previous report
(7), the injection of rIFN- to sensitized WT mice
significantly decreased Ag-induced eosinophil recruitment into the
airways (n = 4, p < 0.001) (Fig. 4). Similarly, the injection of rIFN-
decreased Ag-induced eosinophil recruitment into the airways in
sensitized Tyk2-/- mice (n = 4,
p < 0.001) (Fig. 4). These results suggest that there
is no significant defect in IFN--mediated inhibition of Ag-induced
allergic inflammation in Tyk2-/- mice.
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It has been shown that Tyk2 is expressed not only in lymphoid
cells but also in nonlymphoid cells (11, 12). To determine
which cell type is responsible for the enhanced allergic inflammation
in the airways of Tyk2-/- mice, we performed
the adoptive transfer experiments. To eliminate the interference of
endogenous immune response in recipient mice during the inhaled Ag
challenge, we performed these experiments using BALB/c SCID mice as
recipient mice. Splenocytes from OVA-sensitized
Tyk2-/- mice or littermate WT mice were
stimulated with OVA for 3 days and then transferred i.v. to
nonimmunized BALB/c SCID mice. These mice were challenged with the
inhaled OVA three times at a 24-h interval, and the number of
eosinophils and CD4+ T cells in the BALF was
evaluated at 24 h after the last OVA inhalation. As shown in Fig. 5, when splenocytes from WT mice were
transferred to BALB/c SCID mice, eosinophil recruitment into the
airways was induced by the inhaled Ag challenge (Fig. 5). In the
absence of the cell transfer (PBS alone was injected), few eosinophils
were detected in the BALF (Fig. 5), suggesting that the splenocytes
from donor mice are required for the airway eosinophilia in this
system. Interestingly, Ag-induced eosinophil recruitment into the
airways was significantly increased in mice transferred with
splenocytes from Tyk2-/- mice
(n = 5, p < 0.005) (Fig. 5). In
addition to eosinophils, Ag-induced CD4+ T cell
recruitment was also increased in mice transferred with
splenocytes from Tyk2-/- mice (Fig. 5).
Moreover, IL-4 production in the BALF was increased, but IFN-
production was decreased in mice transferred with splenocytes from
Tyk2-/- mice (data not shown). Furthermore,
when CD4+ T cell-depleted splenocytes from either
WT mice or Tyk2-/- mice were transferred to
BALB/c SCID mice, not only Ag-induced CD4+ T cell
recruitment but also Ag-induced eosinophil recruitment into the airways
was significantly decreased (Fig. 5). These results indicate that the
Ag-induced eosinophil recruitment into the airways is mediated by
CD4+ T cells and thus suggest that
CD4+ T cells are responsible for the enhanced
eosinophil recruitment into the airways in
Tyk2-/- mice.
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Because the levels of IL-13, a key cytokine that induces goblet cell hyperplasia and airway hyperreactivity (5, 6), were increased by inhaled Ag challenge in Tyk2-/- mice (Table I), we next examined the number of epithelial goblet cells as well as airway hyperreactivity to methacholine after Ag inhalation in Tyk2-/- mice. Surprisingly, Ag-induced epithelial goblet cell hyperplasia was significantly decreased in Tyk2-/- mice as compared with that in WT mice (n = 8–10, p < 0.02) (Fig. 6, A and B). In addition, Ag-induced mRNA expression of Muc5ac, one of the mucin genes that are mainly produced by goblet cells in response to IL-13 (31), was decreased in Tyk2-/- mice as compared with that in WT mice (Fig. 6C). Given the increased IL-13 production in the airways of Tyk2-/- mice (Table I), these results suggest that Tyk2 may be involved in IL-13-mediated hyperplasia of epithelial goblet cells. In contrast, no significant difference was observed in the responses to methacholine-induced airway hyperreactivity between Tyk2-/- mice and WT mice (n = 6 for each group; Fig. 6D).
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Discussion |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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Tyk2 was first identified as an essential component in IFN-
signaling (13). However, recent studies with
Tyk2-/- mice have shown that the defective
IFN- responses in Tyk2-/- embryonic
fibroblasts are overcome when high doses of IFN- are provided
(18, 19). Moreover, Tyk2-/- mice
respond normally to IL-10 and IL-6 (18, 19), both of which
activate Tyk2 (14, 15). Thus, unlike other Jaks, the
nonredundant functions of Tyk2 in IFN-, IL-10, and IL-6 signaling
are modest. In contrast, IL-12-induced IFN- production was severely
impaired in Tyk2-/-CD4+ T
cells (Fig. 1 and Refs. 18 and 19), and the
defect was not overcome even by a high concentration of IL-12 (20
ng/ml) (data not shown), suggesting that IL-12 signaling depends more
profoundly on Tyk2 than IFN- signaling.
Interestingly, we found that the levels of IL-4 and IL-5 in the BALF of
Ag-inhaled Tyk2-/- mice were increased as
compared with those in WT mice but IFN- levels were comparable to
those in WT mice (Table I). Because it has been shown that IL-12 and
subsequent Stat4 activation is required for IFN- production in
CD4+ T cells but not in
CD8+ T cells (32), Tyk2 may not be
essential for IFN- production from CD8+ T
cells. Therefore, it is possible that IFN- production from
non-CD4+ T cell populations, including
CD8+ T cells, masks the possible defect in
IFN- production of CD4+ T cells in the airways
of Tyk2-/- mice. However, we also found that,
when Ag-specific CD4+ T cells were stimulated
with antigenic peptide in the absence of exogenous cytokines, Th2 cells
were increased but Th1 cells were not decreased by the absence of Tyk2
(Fig. 1). These results suggest that Tyk2 may be involved in the
regulation of the balance between Th1 cells and Th2 cells beyond its
role in IL-12 signaling for IFN- production. In this regard, it has
been demonstrated that some of the IFN--dependent signaling is
impaired in Tyk2-/- embryonic fibroblasts
(19), although no association of Tyk2 with the IFN-R
has been reported. Therefore, it is plausible that Tyk2 may be involved
in IFN--mediated inhibition of Th2 cell differentiation independent
of IFN- production, thus accounting for the enhanced Th2 cell
differentiation without the decreased IFN- production in
Tyk2-/-CD4+ T cells. This
possibility is under investigation in our laboratory.
In contrast to the enhanced eosinophil and CD4+ T
cell recruitment in Tyk2-/- mice, we also found
that Ag-induced goblet cell hyperplasia and Muc5ac mucin
gene expression were decreased in Tyk2-/- mice
(Fig. 6). Because IL-13, a key cytokine for epithelial goblet cell
hyperplasia and the induction of mucin gene expression (2, 5, 6, 31), was increased in the BALF of
Tyk2-/- mice after Ag inhalation, these results
suggest that Tyk2 may be involved in IL-13-mediated goblet cell
hyperplasia in the airways. This observation is in agreement with the
previous finding that Tyk2 is associated with IL-13R1 chain and
phosphorylated upon IL-13 stimulation (17, 33).
In contrast, we found that Tyk2-/- mice exhibited a response to methacholine-induced airway hyperreactivity similar to that of WT mice (Fig. 6D), whereas Ag-induced goblet cell hyperplasia was decreased in Tyk2-/- mice. Because airway hyperreactivity is believed to be mediated not only by epithelial damage but also by the inflammatory change in the airways (34), the increased inflammatory cell infiltration in the airways may compensate for the decreased epithelial damage in Tyk2-/- mice (Fig. 6A) and then Tyk2-/- mice exhibit a response to methacholine-induced airway hyperreactivity similar to that of WT mice (Fig. 6D).
Although there has been no information available on the expression and/or activation levels of Tyk2 protein in allergic diseases, our results raise the possibility that the impaired expression and/or activation of Tyk2 may be involved in the pathogenesis of Th2 cell-mediated allergic diseases. Further studies are required to address the role of Tyk2 in the pathogenesis of allergic diseases in humans.
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Acknowledgments |
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Footnotes |
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2 Address correspondence and reprint requests to Dr. Hiroshi Nakajima, Department of Internal Medicine II, Chiba University School of Medicine, 1-8-1 Inohana, Chiba City, Chiba 260-8670, Japan. E-mail address: nakajimh{at}intmed02.m.chiba-u.ac.jp
3 Abbreviations used in this paper: Jak, Janus kinase; Stat, signal transducer and activator of transcription; WT, wild type; alum, aluminum hydroxide; PAS, periodic acid-Schiff; BALF, bronchoalveolar lavage fluid.
Received for publication June 24, 2002. Accepted for publication November 6, 2002.
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  K. M. Spach, R. Noubade, B. McElvany, W. F. Hickey, E. P. Blankenhorn, and C. Teuscher A Single Nucleotide Polymorphism in Tyk2 Controls Susceptibility to Experimental Allergic Encephalomyelitis J. Immunol., June 15, 2009; 182(12): 7776 - 7783. [Abstract] [Full Text] [PDF]
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 O. Simma, E. Zebedin, N. Neugebauer, C. Schellack, A. Pilz, S. Chang-Rodriguez, K. Lingnau, E. Weisz, E. M. Putz, W. F. Pickl, et al. Identification of an Indispensable Role for Tyrosine Kinase 2 in CTL-Mediated Tumor Surveillance Cancer Res., January 1, 2009; 69(1): 203 - 211. [Abstract] [Full Text] [PDF]
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