Propionibacterium acnes vaccination induces regulatory T cells and Th1 immune responses and improves mouse atopic dermatitis.
ABSTRACT Atopic dermatitis (AD) is a chronic disease characterized by a polarized Th2 immune response. Propionibacterium acnes (P. acnes) has been shown to elicit strong Th1 immune responses. We hypothesized that the host immune response to P. acnes will prevent the development of AD. To demonstrate this hypothesis, we investigated the effect of P. acnes vaccination on AD that occurs in keratin 14/driven caspase-1 transgenic mouse. Vaccination with low dose of P. acnes successfully prevented clinical manifestations in the skin of AD mice associated with systemic and cutaneous increased expression of Th1-type cytokines but without suppression of Th2 cytokines. Interestingly, the numbers of IFN-γ(+) T cells, FoxP3(+) CD4(+) CD25(+) T cells (nTreg) and IL-10(+) T cells (Tr1) were significantly increased in the spleen. P. acnes vaccination has effects to alter the cytokine milieu and may be useful for the improvement of atopic symptom.
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ABSTRACT: Atopic dermatitis (AD) is a recurrent inflammatory skin disease characterized by dominant T-helper (Th) 2 cytokine response. Bacillus Calmette-Guérin (BCG) has been used for preventing tuberculosis, and is regarded as a strong Th1 cytokine inducer. Antigen (Ag) 85B is a secretory protein present in Mycobacterium species that induces Th1 cytokine production. We investigated the effects of combined vaccination of heat-killed BCG (hkBCG) and Mycobacterium kansasii Ag85B in an AD mouse model. For the AD model, keratin 14 promoter-derived caspase-1 overexpressing mice (KCASP1Tg) were used. The mice received a combination therapy of hkBCG at age 3 weeks and Ag85B twice weekly for 11 weeks from the 4th week; Ag85B monotherapy from the 4th week; hkBCG monotherapy at the 3rd week; or control saline. Areas of skin lesions, cytokine mRNA expression and serum interleukin (IL)-18 and immunoglobulin (Ig) E levels were analysed. Inducible Foxp3+ regulatory T cells (iTreg), IL-10-producing T cells (Tr1), and interferon (IFN)-γ/IL-4/IL-17-producing T cells were evaluated in the spleen. Saline-treated mice and hkBCG monotherapy mice spontaneously developed severe dermatitis. However, combined therapy with hkBCG and Ag85B significantly suppressed the development of skin lesions and mast cell infiltrations. Elevations of the serum IgE and IL-18 levels were significantly suppressed with combined therapy. Mice treated with hkBCG and Ag85B had a normal number of iTreg in the spleen, and decreased number of both IL-4- and IL-17-producing CD4+ T cells. The effect of Ag85B monotherapy was limited. Combined vaccination with hkBCG and Ag85B decreases AD skin lesions by inducing regulatory T cells, suggesting that this vaccination is a potent and novel therapeutic strategy for AD.British Journal of Dermatology 12/2011; 166(5):953-63. · 3.76 Impact Factor
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ABSTRACT: Atopic dermatitis (AD) is a chronic and relapsing inflammatory skin disease, and the pathogenesis is not completely understood. Although there are some mouse models of AD, it is not easy to establish model to represent the natural AD development in human. In this study, we developed an AD model based on outside-inside theory and investigated the effect of Lactobacillus rhamnosus (Lcr35), which have known as an immune modulator in allergic diseases. SKH-1 hairless mice underwent three 1-week exposures (separated by 2-week intervals) to an ovalbumin (OVA) or saline (control) patch at the same site to develop the mouse model of AD. Lcr35 (1 × 10(9) CFU) was administered orally every day from 1 week before the first sensitization until the end of the study. The AD model induced erythematous and itchy skin, increasing TEWL and increasing skin inflammation as assessed by histology in the mice. Oral Lcr35 attenuated all disease parameters previously mentioned. OVA-specific IgE and skin expression of interleukin-4 (IL-4) and thymic stromal lymphopoietin (TSLP) increased in AD mice, but were reduced in AD mice treated with Lcr35. Moreover, Lcr35 treatment led to an increase in CD4(+) CD25(+) Foxp3(+) Treg cells in the mesenteric lymph nodes of AD mice. In conclusions, based on the 'outside-inside' theory, topical allergen may induce AD without skin injury. Oral application of Lcr35 prevented the development of AD in this model by suppressing production of the inflammatory cytokines, IL-4 and TSLP in the skin via a mechanism that may involve CD4(+) CD25(+) Foxp3(+) Treg cells.Experimental Dermatology 05/2012; 21(9):672-5. · 3.58 Impact Factor
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ABSTRACT: There is a high unmet clinical need for new and better treatments in acne vulgaris. Propionibacterium acnes has a strong proinflammatory activity and targets molecules involved in the innate cutaneous immunity, keratinocytes and sebaceous glands of the pilosebaceous follicle. The role of P. acnes in acne confers legitimacy on the possible benefits of immunization-based approaches, which may represent a solution for limiting the development of antibiotic-resistant P. acnes. Various immunization-based approaches have been developed over the last decades, including killed pathogen-based vaccines, vaccination against cell wall-anchored sialidase, monoclonal antibodies to the Christie, Atkins, Munch-Peterson factor of P. acnes, anti-Toll-like receptors vaccines and natural antimicrobial peptides. This review summarizes the current evidence and explores the challenges to making this a realistic treatment option for the future.American Journal of Clinical Dermatology 09/2013; · 2.52 Impact Factor
Propionibacterium acnes vaccination induces regulatory T cells and
Th1 immune responses and improves mouse atopic dermatitis
Hiroshi Kitagawa1, Keiichi Yamanaka1, Masato Kakeda1, Hiroyasu Inada2, Yasutomo Imai3,
Esteban C Gabazza4, Ichiro Kurokawa1and Hitoshi Mizutani1
1Department of Dermatology, Mie University, Graduate School of Medicine, Tsu, Mie, Japan;2Department of Pathology and Matrix Biology, Mie
University, Graduate School of Medicine, Tsu, Mie, Japan;3Department of Dermatology, Hyogo College of Medicine, Nishinomiya, Japan;
4Department of Immunology, Mie University, Graduate School of Medicine, Tsu, Mie, Japan
Correspondence: Hitoshi Mizutani, MD, PhD, Department of Dermatology, Mie University, Graduate School of Medicine, 2-174 Edobashi, Tsu,
Mie 514-8507, Japan, Tel.: 81-59-231-5025, Fax: 81-59-231-5206, e-mail: email@example.com
Abstract: Atopic dermatitis (AD) is a chronic disease
characterized by a polarized Th2 immune response.
Propionibacterium acnes (P. acnes) has been shown to elicit
strong Th1 immune responses. We hypothesized that the host
immune response to P. acnes will prevent the development of
AD. To demonstrate this hypothesis, we investigated the effect of
P. acnes vaccination on AD that occurs in keratin 14⁄driven
caspase-1 transgenic mouse. Vaccination with low dose of
P. acnes successfully prevented clinical manifestations in the skin
of AD mice associated with systemic and cutaneous increased
expression of Th1-type cytokines but without suppression of Th2
cytokines. Interestingly, the numbers of IFN-c+T cells,
FoxP3+CD4+CD25+T cells (nTreg) and IL-10+T cells (Tr1) were
significantly increased in the spleen. P. acnes vaccination has
effects to alter the cytokine milieu and may be useful for the
improvement of atopic symptom.
Key words: atopic dermatitis – IFN-c-regulatory T cell – K14⁄caspase-1
transgenic mouse – Propionibacterium acnes
Accepted for publication 30 July 2010
Atopic dermatitis (AD) is a chronic disease characterized by a pre-
dominant expression of Th2-type cytokines. Th2-type-dominant
cytokine expression is associated with increased cellular infiltration
and overproduction of IgE (1). On the other hand, Th1 cells mainly
secrete IL-12 and IFN-c that counteract the effect of Th2 cells, sup-
press IgE production and stimulate IgG expression. This observa-
tion suggests that control of Th1⁄Th2 imbalance may improve AD.
Th1 cells may be induced by some infectious agents including Propi-
onibacterium acnes (P. acnes). P. acnes is an aerotolerant anaerobic
gram-positive bacteria, present in skin flora of healthy individuals,
that plays an important role in the pathogenesis of acne (2). P. acnes
is known as an inducer of Th1-type immune response (3).
K14⁄caspase-1 transgenic mouse (KCASP1Tg) develops AD char-
acterized by recalcitrant itching dermatitis with increased expres-
sion of Th2-type cytokines (4–6). We investigated the therapeutic
effects of vaccination with low-dose P. acnes upon AD manifesta-
tions, cytokine profile and the involvement of regulatory T cells in
Vaccination of AD model mice with P. acnes
KCASP1Tg was treated with heat-killed P. acnes (66 lg) injection
twice a week between 4 and 14 weeks of age (P. acnes-Tg). PBS-
treated KCASP1Tg (Mock) and wild-type C57⁄BL6 (WT) were
used as control mice (Appendix S1). Skin manifestation usually
appears in KCASP1Tg mice after the age of 8 weeks. Skin manifes-
tations were examined every 2 weeks between 8 and 14 weeks, and
the affected skin areas were calculated as percentage of the whole
body area. Skin specimens were also sampled at 14th week and
stained with haematoxylin & eosin and toluidine blue.
Data were expressed as the mean ± standard error of the mean
(SEM). The statistical difference between variables was calculated
by the Kruskal–Wallis one-way analysis of variance. P < 0.05 was
considered as significant.
Cutaneous manifestations and histopathological findings
KCASP1Tg (Mock) developed erosive dermatitis on the face at
8th week and showed severe scratching. Skin lesions spread rapidly
to the ears, neck and trunk (Fig. 1a). The erosive lesions worsened
and became hairless lichenoid dermatitis mixed with acute lesions.
In contrast, P. acnes-Tg developed significantly milder dermatitis,
and the total involved skin area was significantly less than in
Mock after 10 weeks (Fig. 1b). Histopathologically, the 14-week-
old Mock showed marked inflammatory reactions with acanthosis,
hyperkeratosis and marked cellular infiltration. The inflammatory
changes and the number of mast cells were significantly decreased
in the P. acnes-Tg compared to Mock (Fig. 1c).
This study demonstrated a novel candidate for the improvement of
AD by vaccination with P. acnes. Vaccination with the extract of this
microorganism improved skin lesions by acting as an immune
response modifier. Antigen from P. acnes activates IL-12p40 pro-
moter after binding to TLR2 (2,7). IL-12 binds to its receptor (IL-
12R) and activates the transcription factors STAT-4 and T-bet, and
then T-bet binding to the promoter of IFN-c gene stimulates the
production of this Th1-polarizing cytokine (8,9). In this study, the
cytokine production was evaluated in spleen cells from each group
of mice, and it was found that IFN-c-positive T cells are significantly
elevated in P.acnes-Tg (Appendix S1 and Fig. S2). In addition, the
mRNA expressions of IFN-c, IL-12 and T-bet were also significantly
Letter to the Editor
ª 2011 John Wiley & Sons A/S, Experimental Dermatology, 20, 145–158
elevated in the spleen cells from P. acnes-Tg compared to control
mice (Appendix S1, Figs S1 and S3). In our previous report, injec-
tion of 1mg of P.acnes into KCASP1Tg caused death of mouse
owing to severe liver injury (4). However, in this experiment, vacci-
nation with low dose of P. acnes increased the differentiation of T
cells expressing Th1-type cytokines without liver injury (data not
shown). P. acnes treatment did not affect the elevated IL-4 mRNA
expression in KCASP1Tg, suggesting that P. acnes therapy does not
simply switch Th2 into Th1-type cells.
Th17 and Treg cell are also important immunoregulators. IL-
17A mRNA and IL-17A-positive cells are significantly elevated in
Mock, suggesting the involvement of Th17 immune response in
the pathogenesis of dermatitis (Figs S1 and S2). However, we
found that Th17 expression is not affected by P. acnes treatment.
The expression of IL-17A receptor is also a critical factor for IL-17
signal transduction. The expression of IL-17A receptor was
decreased in the skin of Mock compared to that of WT (Fig. S3),
suggesting that there is a negative feedback following increased IL-
17A production in KCASP1Tg. P. acnes vaccination showed no
effects on IL-17A receptor expression. Therefore, it appears that
IL-17 is involved in the development of dermatitis, but it is not
affected by P. acnes therapy.
Regarding regulatory T cells, nTreg cells were slightly decreased
in the spleen of Mock compared to WT. However, P. acnes ther-
apy increased nTreg in KCASP1Tg (Fig. S2). A bacterial TLR2
agonist has been shown to induce nTreg in vivo (10); therefore,
P. acnes vaccination may increase nTreg via TLR2 (7).
TGF-b is a pleiotropic cytokine that may increase the number of
Treg. This study showed that vaccination with P. acnes significantly
augmented TGF-b mRNA expression compared to untreated mice
(Figs S1 and S3), suggesting that the suppression of TGF-b expres-
sion plays a role in the development of dermatitis in KCASP1Tg
and that TGF-b is a key factor to control dermatitis.
P. acnes treatment significantly increased the number of Tr1
cells compared to WT (Fig. S2). We also analysed mRNA expres-
sion in spleen cells and skins (Figs S1 and S3) and found that the
mRNA expression of IL-10 is significantly enhanced in P. acnes-Tg
compared to that in Mock and WT. In humans, IL-10 released by
Tr1 cells is critical for the immune control of atopic diseases
(11,12). A recent study has shown that P. acnes-soluble polysac-
charide suppresses type1 hypersensitivity late-phase reaction in
mice (13). However, further investigation is warranted to clarify
the mechanism of Treg induction by P. acnes.
In brief, we showed in this study that P. acnes vaccination ame-
liorates AD by generating natural and induced Tregs and by
inducing Th1 cytokines in a mouse model that spontaneously
develops intrinsic AD. This study indicates that P. acnes immuno-
therapy may be a potent therapeutic approach for AD.
The authors have no financial conflict of interest.
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Additional Supporting Information may be found in the
online version of this article:
Figure S1. mRNA expression in spleen cells.
Figure S2. Intracellular cytokine and Foxp3 staining
in spleen cells.
Figure S3. mRNA expression in the skin lesions.
Appendix S1. Materials and methods.
Please note: Wiley-Blackwell are not responsible for
the content or functionality of any supporting materials
supplied by the authors. Any queries (other than missing
material) should be directed to the corresponding author
for the article.
Kitagawa et al.
KCASP1 Tg mice
KCASP1 Tg mice
Skin eruption area
The number of toluidine
P.acnes-Tg Mock WT
Figure 1. (a) Skin facial manifestations at 14th week of age. Severe dermatitis
was observed on the face in Mock. In contrast, P. acnes-treated mice (P. acnes-Tg)
showed mild dermatitis limited to the periocular areas. Histologically, Mock showed
severe skin lesions including acanthosis and hyperkeratosis in the epidermis and
marked fibrosis with dense mononuclear cell infiltration. P. acnes-Tg showed mild
inflammatory reactions and limited epidermal acanthosis (H&E section). The
infiltration of mast cells was also increased in the skin from Mock compared to
P. acnes-Tg (toluidine blue stain). (b) Time course and skin areas with disease. The
diseased skin area increased from 8 weeks after birth in Mock, but it was less in
P. acnes-Tg. Data are expressed as the mean ± SE *P < 0.05. (c) The toluidine-
blue-positive mast cell numbers per HPF were significantly increased in Mock
compared to WT; the degree decreased to the level of WT in animals treated with
P. acnes. Data are expressed as the mean ± SE *P < 0.05, **P < 0.01.
Letter to the Editor
ª 2011 John Wiley & Sons A/S, Experimental Dermatology, 20, 145–158