Ex-Vivo Skin Explant Culture Is a Model for TSLP-Mediated Skin Barrier Immunity
Abstract
:1. Introduction
2. Materials and Methods
2.1. Mice
2.2. Serum Isolation
2.3. Epidermal Ear Sheets
2.4. Histological Analysis and Immunofluorescence Microscopy
2.5. Epidermal Single Cell Suspension and Flow Cytometry Analysis
2.6. Skin Explant Cultures
2.7. ELISA
2.8. Skin Protein Isolation
2.9. Statistical Methods
3. Results
3.1. TSLP Is Induced and Released from EGFR∆ep Skin In Vivo and Ex Vivo
3.2. TSLP Expression from Skin Explants Cultured at 32 °C Represents an Ex-Vivo Model for Skin Barrier Immunity
3.3. TSLP Transcriptional Regulator NFAT Is Not Up-Regulated in EGFR∆ep Mice, and an NFAT Pathway Inhibitor Does Not Block TSLP Expression Ex Vivo
3.4. JNK Pathway Inhibitor Blocks TSLP Expression Ex Vivo
4. Discussion
5. Conclusions
Supplementary Materials
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Acknowledgments
Conflicts of Interest
References
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Antibodies | SOURCE | IDENTIFIER |
---|---|---|
Armenian hamster anti-mouse TCR γ/δ Antibody, FITC | Biolegend | Cat# 118106, RRID:AB_313830 |
Rat anti-mouse/human CD11b Antibody, Pacific Blue | Biolegend | Cat# 101224, RRID:AB_755986 |
Armenian hamster anti-mouse CD11c Antibody, PE/Cy5 | Biolegend | Cat# 117316, RRID:AB_493566 |
Rat anti-mouse I-A/I-E Antibody, APC/Cy7 | Biolegend | Cat# 107628, RRID:AB_2069377 |
Rat anti-mouse CD45 Antibody, APC | Biolegend | Cat# 103112, RRID:AB_312977 |
Rat anti-Langerin/CD207 Antibody (929F3.01), Alexa Fluor 488 | Dendritics | Cat# DDX0362A488, RRID:AB_1148740 |
Rat anti-mouse I-A/I-E Antibody, PE | Biolegend | Cat# 107608, RRID:AB_313323 |
Rat anti-mouse/human Langerin/CD207, Alexa Fluor 488 | Dendritics | Cat# DDX0362 929F3.01 |
Rat anti-mouse EpCAM/CD326, FITC | Biolegend | Cat# 118208, RRID:AB_1134107 |
Rat anti-mouse CD45, PE | Biolegend | Cat# 103106, RRID:AB_312971 |
Rat anti-mouse CD45, APC | Biolegend | Cat# 103112, RRID:AB_312977 |
Anti-beta III Tubulin Antibody (TUJ-1) | abcam | ab14545 |
JunD (329) | Santa cruz | sc-74 |
NFATc1 | Santa cruz-sc-7294 | 7A6 |
Fra2 CN10 | home made | |
Phospho-cJun Ser73 | Cell Signalling | D47G9 |
cJun | Cell Signalling | 60A8 |
JunB | Cell Signalling | C37F9 |
cFos | THP-Medical Products | Y075085 |
CD104-Integrin β4 chain | BD Pharmingen™ | Cat# 553745 |
Donkey anti-rabbit 594/488 Alexa Fluor | Invitrogen-Life Technologies | Cat# A21206 |
Donkey anti-rat 594/488 Alexa Fluor | Invitrogen-Life Technologies | Cat# 21208 |
Donkey anti-Guinea Pig 594 Alexa Fluor | Invitrogen-Life Technologies | Cat# A11076 |
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Bauer, T.; Gubi, D.; Klufa, J.; Novoszel, P.; Holcmann, M.; Sibilia, M. Ex-Vivo Skin Explant Culture Is a Model for TSLP-Mediated Skin Barrier Immunity. Life 2021, 11, 1237. https://doi.org/10.3390/life11111237
Bauer T, Gubi D, Klufa J, Novoszel P, Holcmann M, Sibilia M. Ex-Vivo Skin Explant Culture Is a Model for TSLP-Mediated Skin Barrier Immunity. Life. 2021; 11(11):1237. https://doi.org/10.3390/life11111237
Chicago/Turabian StyleBauer, Thomas, Daniela Gubi, Jörg Klufa, Philipp Novoszel, Martin Holcmann, and Maria Sibilia. 2021. "Ex-Vivo Skin Explant Culture Is a Model for TSLP-Mediated Skin Barrier Immunity" Life 11, no. 11: 1237. https://doi.org/10.3390/life11111237