secretion of thymic stromal lymphopoietin from human keratinocytes in response to malassezia yeasts

Letters to the Editor / Journal of Dermatological Science 62 (2011) 128–140134

This work was supported in part by a Grant-in-Aid for ScientificResearch Project for Private Universities (Atopy (Allergy) ResearchCenter, Juntendo University) from the Ministry of Education,Culture, Sports, Science and Technology, Japan.

Acknowledgements

The authors thank Drs. Honda and Yanai for providing us viruses.We also thank the members of the Atopy Research Center and theDepartment of Immunology for fruitful discussions and technicalassistance, and Ms Michiyo Matsumoto for secretarial assistance.

Appendix A. Supplementary data

Supplementary data associated with this article can be found, in

the online version, at doi:10.1016/j.jdermsci.2011.02.005.

References

[1] He R, Geha RS. Thymic stromal lymphopoietin. Ann NY Acad Sci2010;1183:13–24.

[2] Soumelis V, Reche PA, Kanzler H, Yuan W, Edward G, Homey B, Gilliet M, Ho S,Antonenko S, Lauerma A, Smith K, Gorman D, Zurawski S, Abrams J, Menon S,McClanahan T, de Waal-Malefyt Rd R, Bazan F, Kastelein RA, Liu YJ. Humanepithelial cells trigger dendritic cell mediated allergic inflammation by pro-ducing TSLP. Nat Immunol 2002;3:673–80.

[3] Kinoshita H, Takai T, Le TA, Kamijo S, Wang XL, Ushio H, Hara M, Kawasaki J, VuAT, Ogawa T, Gunawan H, Ikeda S, Okumura K, Ogawa H. Cytokine milieumodulates release of thymic stromal lymphopoietin from human keratino-cytes stimulated with double-stranded RNA. J Allergy Clin Immunol2009;123:179–86.

[4] Niyonsaba F, Ushio H, Nakano N, Ng W, Sayama K, Hashimoto K, Nagaoka I,Okumura K, Ogawa H. Antimicrobial peptides human beta-defensins stimulateepidermal keratinocyte migration, proliferation and production of proinflam-matory cytokines and chemokines. J Invest Dermatol 2007;127:594–604.

[5] Takeuchi O, Akira S. Innate immunity to virus infection. Immunol Rev2009;227:75–86.

[6] Weber F, Wagner V, Rasmussen SB, Hartmann R, Paludan SR. Double-strandedRNA is produced by positive-strand RNA viruses and DNA viruses but not indetectable amounts by negative-strand RNA viruses. J Virol 2006;80:5059–64.

[7] Kato H, Takeuchi O, Mikamo-Satoh E, Hirai R, Kawai T, Matsushita K, Hiiragi A,Dermody TS, Fujita T, Akira S. Length-dependent recognition of double-strand-ed ribonucleic acids by retinoic acid-inducible gene-I and melanoma differ-entiation-associated gene 5. J Exp Med 2008;205:1601–10.

[8] Le Goffic R, Pothlichet J, Vitour D, Fujita T, Meurs E, Chignard M, Si-Tahar M.Cutting edge: influenza A virus activates TLR3-dependent inflammatory andRIG-I-dependent antiviral responses in human lung epithelial cells. J Immunol2007;178:3368–72.

[9] Metz M, Maurer M. Innate immunity and allergy in the skin. Curr OpinImmunol 2009;21:687–93.

[10] Prignano G, Ferraro C, Mussi A, Stivali F, Trento E, Bordignon V, Crescimbeni E,Salvati G, Degener AM, Ameglio F. Prevalence of human papilloma virus type 5DNA in lesional and non-lesional skin scales of Italian plaque-type psoriaticpatients: association with disease severity. Clin Microbiol Infect 2005;11:47–51.

Junko Kawasakia,b

aDepartment of Dermatology,

Juntendo University School of Medicine,

Bunkyo-ku, 2-1-1 Hongo,

Tokyo 113-8421, JapanbAtopy (Allergy) Research Center,



Tokyo 113-8421, Japan

Hiroko Ushio*Atopy (Allergy) Research Center,




Hirokazu KinoshitaTatsuo Fukai

Department of Dermatology,




Francois NiyonsabaTshiro Takai

Hideoki OgawaKo Okumura

Atopy (Allergy) Research Center,




Shigaku Ikedaa,b

aDepartment of Dermatology,



Tokyo 113-8421, JapanbAtopy (Allergy) Research Center,




*Corresponding author. Tel.: +81 3 5802 1591;fax: +81 3 3813 5512

E-mail address: [email protected] (H. Ushio).

19 October 2010

doi:10.1016/j.jdermsci.2011.02.005

Letter to the Editor

Secretion of thymic stromal lymphopoietin from humankeratinocytes in response to Malassezia yeasts

Thymic stromal lymphopoietin (TSLP) plays an important rolein the pathogenesis of atopic dermatitis (AD) by promotingdendritic cell-mediated activation of the Th2 inflammatoryresponse [1]. The lipophilic yeasts Malassezia globosa andMalassezia restricta are members of the cutaneous microfloraand act as exacerbating factors in AD [2]. Despite the in vivo

expression and potential importance of keratinocyte-derived TSLP

in AD [1], no studies have tested the influence of the presence ofMalassezia yeast cells on the production of TSLP by humankeratinocytes. In this study, we determined the TSLP expressionprofiles of human keratinocytes after stimulation with Malassezia

yeast cells.M. globosa CBS7966 and M. restricta CBS7877 were cultured at

32 8C on modified Leeming and Notman agar [2]. Acapsular cells ofMalassezia species were prepared by treatment with Triton X-100(TX-100) or n-octyl-b-D-glucoside (OG) [3]. Primary normalhuman epidermal keratinocytes (NHEKs) were obtained from

http://dx.doi.org/10.1016/j.jdermsci.2011.02.005

mailto:[email protected]


[()TD$FIG]

Fig. 1. (A) Malassezia globosa and M. restricta induce thymic stromal lymphopoietin (TSLP) secretion from human keratinocytes. Normal human epidermal keratinocytes (NHEKs)

(1� 105 cells well�1) were exposed to M. globosa or M. restricta at a multiplicity of infection (MOI) of 20. The keratinocyte culture supernatants were collected after 24 h, and the

levels of TSLP in these supernatants were measured by an enzyme-linked immunosorbent assay (ELISA). The detection limit for TSLP was 3.8 pg mL�1. Data are expressed as the

mean (SE) of 4 experiments. Each experiment was performed in duplicate. nd, Less than the minimum detection limit. Statistical significance was determined by using a two-tailed

Student’s t-test; **P < 0.01. (B) Malassezia-induced upregulation of TSLP mRNA expression in keratinocytes. Total RNA was extracted after a 4-h incubation period, and TSLP mRNA

expression was analyzed by reverse transcription polymerase chain reaction (RT-PCR). Glyceraldehyde 3-phosphate dehydrogenase (GAPDH) was used as the internal mRNA

control. (C) Calcium-induced differentiation of keratinocytes enhances Malassezia-induced TSLP secretion. NHEK cells (1� 105 cells well�1) were cultured in the medium

containing 0.15 mM (low-calcium) or 1.35 mM CaCl2 (high-calcium) for 24 h. The cells were then exposed to M. globosa or M. restricta at an MOI of 20. Data are expressed as the

mean (SE) of 4 experiments; **P < 0.01 vs. the low calcium conditions. (D) Hydrocortisone impairs Malassezia-induced TSLP secretion. NHEK cells (1� 105 cells well�1) were

cultured in the presence or absence of 0.5 mg mL�1 hydrocortisone for 24 h. The cells were then exposed to M. globosa or M. restricta at an MOI of 20. Data are expressed as the mean

(SE) of 4 experiments; **P < 0.01 vs. the presence of hydrocortisone. (E) Treatment of Malassezia yeasts with detergents attenuates Malassezia-induced secretion of TSLP from

human keratinocytes. M. globosa or M. restricta samples were treated with 10% (v/v) Triton X-100 (TX-100) or 1.0% (v/v) n-octyl-b-D-glucoside (OG) for 1 h at 37 8C. After washing,

these microorganisms were added to NHEK cells (1� 105 cells well�1) at an MOI of 20, andthe TSLP secretion was analyzed after 24 h. As a control, NHEK cells were incubated with

culture medium only. Data are expressed as the mean (SE) of 4 experiments; **P < 0.01 vs. untreated Malassezia yeasts. (F) OG-extracted fractions of Malassezia yeasts induce the

secretion of TSLP from human keratinocytes. An OG-extracted fraction was prepared by incubating Malassezia yeasts with 1.0% OG in PBS for 1 h at 37 8C. The soluble fraction was

collected and dialyzed against PBS to remove the detergent. NHEK cells (1� 105 cells well�1) were incubated without (control) or with various concentrations of OG-extracted

fractions prepared from M. grobosa or M. restricta. TSLP secretion was analyzed after 24 h. A concentration of 1 U mL�1 of the OG-extracted fraction is equivalent to

2� 106 yeast cells mL�1. Data are expressed as the mean (SE) of 4 experiments. **P < 0.01 vs. control.

Letters to the Editor / Journal of Dermatological Science 62 (2011) 128–140 135

[()TD$FIG]

Fig. 2. (A) Blocking of lysophosphatidic acid (LPA) receptors on keratinocytes impairs Malassezia-induced production of TSLP. NHEK cells (1 � 105 cells well�1) were

pretreated with various concentrations of DGPP or VPC32183 for 1 h, and then exposed to M. globosa or M. restricta at an MOI of 20. As a control, NHEK cells were incubated

with culture medium only. Data are expressed as the mean (SE) of 4 experiments. **P < 0.01 vs. without LPA antagonists. (B) Inhibitors of LPA receptors abolish the Malassezia

extract-induced production of TSLP from keratinocytes. NHEK cells (1 � 105 cells well�1) were pretreated with 20 mM DGPP or VPC32183 for 1 h, and then exposed to

1 U mL�1 (equivalent to 2 � 106 yeast cells mL�1) of M. globosa- or M. restricta-extracted fractions. Data are expressed as the mean (SE) of 4 experiments. **P < 0.01 vs.

without LPA antagonists. (C) Monoclonal antibodies against toll-like receptors do not influence the Malassezia extract-induced production of TSLP from keratinocytes. NHEK

cells (1 � 105 cells well�1) were pretreated with 25 mg mL�1 of anti-human mAbs against TLR2, TLR3, TLR5, TLR6, or isotype control IgG, and then exposed to 1 U mL�1

(equivalent to 2 � 106 yeast cells mL�1) of M. globosa- or M. restricta-extracted fractions. Data are expressed as the mean (SE) of 4 experiments.


Letters to the Editor / Journal of Dermatological Science 62 (2011) 128–140 137

PromoCell (Germany). The cells were cultured at 37 8C in 5% CO2 inKBM-2 medium (Clonetics Inc.) with supplements including0.5 mg mL�1 hydrocortisone. NHEK cells (1 � 105 cells well�1)were exposed to M. globosa or M. restricta at a multiplicity ofinfection (MOI) of 20 in vitro. The levels of TSLP protein in culturesupernatants were measured using the human TSLP DuoSet ELISADevelopment kit (R&D Systems Inc., MN). The expression of TSLPmRNA was determined using a one-step reverse transcriptionpolymerase chain reaction (RT-PCR) kit (Qiagen).

The exposure of NHEK cells to M. globosa or M. restricta induceda marked increase in TSLP secretion (Fig. 1A). RT-PCR analysisconfirmed that upregulation of TSLP mRNA expression occurs inparallel with enhanced TSLP secretion from NHEK cells duringexposure to M. globosa or M. restricta (Fig. 1B). Keratinocytesundergo differentiation under high-calcium culture conditions.Malassezia-induced TSLP secretion from keratinocytes was foundto be markedly enhanced under high-calcium conditions relativeto low-calcium conditions (Fig. 1C). This is consistent with thehistological finding that TSLP production occurs in the differenti-ated suprabasal layer of the skin epidermis [1]. Removal ofhydrocortisone from the culture medium enhances Malassezia-induced TSLP secretion from keratinoyctes (Fig. 1D). Thisobservation is consistent with a previous report demonstratinginhibition of dsRNA-induced TSLP in human keratinocytes byglucocorticoids [4]. Malassezia species are surrounded by a lipid-rich capsular-like layer [3]. Malassezia-induced TSLP secretion wasabrogated by removal of the lipid layer of Malassezia yeasts bytreatment with TX-100 or OG (Fig. 1E). These findings suggest thatdetergent-extractable components of Malassezia yeasts may beneeded to stimulate TSLP secretion from human keratinocytes.Indeed, the addition of an OG-extracted fraction to the NHEKculture dishes induces TSLP secretion from NHEK cells in aconcentration-dependent manner (Fig. 1F).

TSLP production can be triggered and enhanced by a variety ofstimuli, such as cytokines and pathogen-associated molecularpatterns (PAMPs) [1,4]. In addition, lysophosphatidic acid (LPA)has been reported to stimulate TSLP expression in bronchialepithelial cells [5]. LPA receptors, such as LPA1, LPA2, and LPA3,exist in primary keratinocytes as well as in intact skin. DGPP andVPC32183, competitive antagonists of the LPA1/3 receptors, inhibitMalassezia-induced TSLP secretion from NHEK cells (Fig. 2A).Furthermore, Malassezia extract-stimulated TSLP secretion fromNHEK cells was markedly blocked by 20 mM DGPP and VPC32183(Fig. 2B). These results suggested that the lipid layer of Malassezia

may act as a ligand for LPA1/3 receptors to stimulate TSLPsecretion.

A previous report has described toll-like receptor (TLR)-3-dependent TSLP secretion from human keratinocytes in responseto dsRNA [6]. Recently, Staphylococcus aureus membrane anddiacylated lipopeptide have been demonstrated to induce TSLPexpression in keratinocytes through the TLR2-TLR6 pathway [7].Furthermore, flagellin has been reported to induce TSLP expressionin human keratinocytes via TLR5 [8]. Since we observed thatantibodies which block TLR2, TLR3, TLR5, and TLR6 do not inhibitTSLP secretion from keratinocytes in response to OG-extractedfractions from Malassezia (Fig. 2C), it seems unlikely that theseTLRs play a functional role in Malassezia-induced TSLP secretionfrom keratinocytes.

The proposed pathogenic roles of the lipid layer of Malassezia

are paradoxical. Thomas et al. demonstrated that the removal ofthe lipid layer of Malassezia increases the stimulation of proin-flammatory cytokines, such as IL-6, IL-8, and IL-1a, but decreasesthe levels of intracellular IL-10 in human keratinocytes [3]. Thisreport also postulates that the lipid layer of Malassezia may beimportant for evading a keratinocyte-driven inflammatory re-sponse. Our results indicate that the lipid layer of Malassezia could

contain a ligand for LPA receptors to stimulate TSLP secretion fromkeratinocytes. This contradiction may be explained by differencesin the PAMPs and the corresponding pattern recognition receptorsthat are responsible for triggering TSLP and proinflammatorycytokines. Baroni et al. demonstrated that TLR-2 is required forMalassezia furfur-induced IL-8 expression in keratinocytes [9].Since TLR-2-mediated proinflammatory cytokine production isrestored after removal of the lipid layer of M. furfur [10], the lipidlayer may inhibit the interaction between PAMPs of Malassezia andTLR-2 on keratinocytes. Thus, it is likely that the lipid layer ofMalassezia has dual functions in interacting with LPA receptors tostimulate TSLP expression and interfering with the interactionsbetween TLR-2 and PAMPs.

Currently, there is very little information about the molecularcomponents of the lipid layer of Malassezia. Further studiesinvolving physicochemical characterization of lipid layer compo-nents are needed to elucidate the molecular basis of the interactionbetween the lipid layer of Malassezia and LPA receptors on humankeratinocytes.

In conclusion, our results suggest that M. globosa and M. restricta

induce the secretion of TSLP from human keratinocytes throughthe interaction of the lipid layer of Malassezia with the LPAreceptors on keratinocytes. In normal skin, keratinocytes formhorny layer in the epidermis, which blocks interactions of microbeswith living keratinocytes. Dysfunction of this barrier in AD patientsmay allow Malassezia to interact with living keratinocytes.

Malassezia-induced TSLP secretion by keratinocytes may play arole in stimulating the Th2 immune response, which in turn willtrigger or exacerbate AD. Further elucidation of the mechanisms bywhich Malassezia induces TSLP secretion from keratinocytes willclarify the pathogenesis of AD and provide clues for developingnew therapeutic strategies.

References

[1] He R, Geha RS. Thymic stromal lymphopoietin. Ann NY Acad Sci2010;1183:13–24.

[2] Takahata Y, Sugita T, Kato H, Nishikawa A, Hiruma M, Muto M. CutaneousMalassezia flora in atopic dermatitis differs between adults and children. Br JDermatol 2007;157:1178–82.

[3] Thomas DS, Ingham E, Bojar RA, Holland KT. In vitro modulation of humankeratinocyte pro- and anti-inflammatory cytokine production by the capsuleof Malassezia species. FEMS Immunol Med Microbiol 2008;54:203–14.

[4] Le TA, Takai T, Vu AT, Kinoshita H, Ikeda S, Ogawa H, et al. Glucocorticoidsinhibit double-stranded RNA-induced thymic stromal lymphopoietin releasefrom keratinocytes in an atopic cytokine milieu more effectively than tacro-limus. Int Arch Allergy Immunol 2010;153:27–34.

[5] Medoff BD, Landry AL, Wittbold KA, Sandall BP, Derby MC, Cao Z, et al. CARMA3mediates lysophosphatidic acid-stimulated cytokine secretion by bronchialepithelial cells. Am J Respir Cell Mol Biol 2009;40:286–94.

[6] Kinoshita H, Takai T, Le TA, Kamijo S, Wang XL, Ushio H, et al. Cytokine milieumodulates release of thymic stromal lymphopoietin from human keratino-cytes stimulated with double-stranded RNA. J Allergy Clin Immunol2009;123:179–86.

[7] Vu AT, Baba T, Chen X, Le TA, Kinoshita H, Xie Y, et al. Staphylococcus aureusmembrane and diacylated lipopeptide induce thymic stromal lymphopoietinin keratinocytes through the Toll-like receptor 2-Toll-like receptor 6 pathway.J Allergy Clin Immunol 2010;126:985–93.

[8] Le TA, Takai T, Vu AT, Kinoshita H, Chen X, Ikeda S, et al. Flagellin induces theexpression of thymic stromal lymphopoietin in human keratinocytes via Toll-like receptor 5. Int Arch Allergy Immunol 2010;155:31–7.

[9] Baroni A, Orlando M, Donnarumma G, Farro P, Iovene MR, Tufano MA, et al.Toll-like receptor 2 (TLR2) mediates intracellular signalling in human kerati-nocytes in response to Malassezia furfur. Arch Dermatol 2006;297:280–8.

[10] Oh C, Kim A, Kuo I, Krutzik SR, Liu PT, Sieling PA, et al. Lipids on the Malasseziafurfur cell wall inhibit proinflammatory cytokine production in human mono-cytes by downregulating the toll-like receptor 2: immunomodulatory role ofMalassezia furfur. J Invest Dermatol 2005;124:A127.

Yoshio Ishibashi*Kazuya Sugawara

Department of Immunobiology, Meiji Pharmaceutical University,

Noshio, Kiyose, Tokyo 204-8588, Japan


Takashi SugitaDepartment of Microbiology, Meiji Pharmaceutical University, Noshio,

Kiyose, Tokyo 204-8588, Japan

Akemi NishikawaDepartment of Immunobiology, Meiji Pharmaceutical University,

Noshio, Kiyose, Tokyo 204-8588, Japan

§ This work was supported by the Bundesminsterium fur Umwelt (BMU), the

Deutsche Forschungsgemeinschaft SFB 728, an unrestricted grant by Estee Lauder

Inc and by a grant from the Ministry of Education, Culture, Sports, Science and

Technology Japan. We would like to thank all study participants.

*Corresponding author. Tel.: +81 42 495 8741;fax: +81 42 495 8612

E-mail address: [email protected] (Y. Ishibashi)

24 December 2010

doi:10.1016/j.jdermsci.2011.02.012

Letter to the Editor

Table 1Antioxidant levels in fasting blood samples of German and Japanese women.

GM (95% CI) p

German women Japanese women

Carotenoids in mmol/l

a-Carotene 0.05 (0.04–0.07) 0.16 (0.12–0.20) <0.001

ß-Carotene 0.32 (0.32–0.46) 0.79 (0.64–0.98) <0.001

ß-Cryptoxanthin 0.19 (0.14–0.25) 0.32 (0.26–0.40) 0.005

Lycopene 0.29 (0.21–0.40) 0.48 (0.41–0.58) 0.005

Vitamins in mmol/l

Retinol 0.94 (0.71–1.25) 1.25 (1.17–1.33) 0.091

Association between sun-exposure, smoking behaviour andplasma antioxidant levels with the different manifestation ofskin ageing signs between Japanese and German women—Apilot study

To the Editor,

The clinical manifestation of extrinsic skin ageing differsbetween Asian and Caucasian women. It has been observed thatAsian women develop pigment spots (lentigines) much earlier thanage-matched Caucasian women, whereas the opposite is true forthe formation of coarse wrinkles [1–5]. Underlying reasons haveremained enigmatic. In the present study we tested the hypothesiswhether differences in sun-exposure, smoking or plasma antioxi-dant levels, which are all known influencing factors of skin ageing[6–8], might be associated with the regional difference in skinageing manifestation.

In the years 2007/2008 we investigated healthy femalevolunteers: 39 German women living in Dusseldorf and 48Japanese women living in Nagoya. Both cities have similar climaticconditions. Volunteers did not apply cosmetic or treatmentproducts to their face the day of assessment so that facial skinageing symptoms could be properly evaluated according toSCINEXA (SCore for INtrinsic and EXtrinsic skin Ageing) [9].SCINEXA evaluations were performed by two different personstrained by the same investigator. Inter-evaluator concordanceranges from moderate to high. Fasting blood samples wereanalyzed by HPLC to measure plasma antioxidants. In aninterview-based lifestyle questionnaire subjects were asked aboutthe environmental factors in question, such as diet, sun exposureand smoking behaviour. The medical ethics committee of theHeinrich-Heine University Dusseldorf, Germany as well as of theNagoya City University, Japan approved the entire study. Thedeclaration of Helsinki Principles was followed and all studyparticipants were informed in detail by written form and gavewritten consent.

Multiple linear regressions were used to determine thecontribution of multiple factors on skin ageing. We first calculatedthe age-independent regional difference in wrinkle and lentigines

manifestation. Then we stepwise adjusted for further influencingfactors, in order to see if these factors impacted the difference in aspecific ageing sign. Statistical significance was defined as p < 0.05.The analyses were performed with statistical software SAS 9.2 (SASInstitute Inc., Cary, NC, USA, 2002–2003).

The mean age of German women was 52.2 � 9.1 years and ofJapanese women 45.7 � 7.6 years. There was no significant differencein the menopausal status of the two populations after adjusting for

age. German women reported a higher exposure to UV radiation, asmore than 30% of the German women used sunbeds but none of theJapanese women did and German women reported spending morehours per day outside in summer time than Japanese women. Therewas no significant difference between German or Japanese women inthe number of weeks spent in sun-rich regions per year. Japanesewomen had a lower rate of smoking than German women.

The skin ageing signs of wrinkles under the eyes, wrinkles onupper lip and lentigines on cheeks showed significant differences inthe manifestation between German and Japanese women withwrinkles more pronounced in German women and lentigines beingmore frequent in Japanese women. Measurements of plasmaantioxidants revealed striking differences between the two groups(Table 1). In comparison to German women, Japanese women hadsignificantly higher concentrations of all investigated antioxidantsexcept retinol independent of age. More than 60% of all womenreported regular consumption of fruit and vegetables in the recenttime and this was positively associated with the plasmaantioxidant concentration. However, it was not possible toaccurately evaluate differences in intake of specific nutrients, inabsorption or in metabolism between the two regional groupswhich might further influence the plasma antioxidant concentra-tion. Nevertheless, plasma antioxidant levels significantly andinversely correlated with signs of skin ageing. In German womenthe protective effect of antioxidants on skin ageing signs was mostpronounced for a-tocopherol and retinol. In Japanese women b-carotene, lycopene and retinol were most protective. In Germanand older Japanese (51–70 years) women antioxidants had also aprotective effect on the occurrence of lentigines on cheeks. Finally,we tested how the regional differences in specific skin ageing signschange when additionally adjust for further influencing factors(Table 2). As only German women used sunbeds, we excludedGerman sunbed users from this analysis. The age adjusted

a-Tocopherol 19.75 (15.25–25.57) 28.97 (26.27–32.06) 0.002

g-Tocopherol 1.21 (0.98–1.48) 2.96 (2.54–3.47) <0.001

GM (95% CI), geometric mean with 95% confidence interval; p, p-value of test for

differences in antioxidant level between German and Japanese women adjusted

for age

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secretion of thymic stromal lymphopoietin from human keratinocytes in response to malassezia yeasts

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