The Receptor for Advanced Glycation End Products Is Highly Expressed in the Skin and Upregulated by Advanced Glycation End Products and Tumor Necrosis.

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The Receptor for Advanced Glycation End Products Is Highly Expressed in the Skin and Upregulated by Advanced Glycation End Products and Tumor Necrosis Factor-Alpha Christina Lohwasser, Daniel Neureiter, Bernd Weigle, Thomas Kirchner, Detlef Schuppan Journal of Investigative Dermatology Volume 126, Issue 2, Pages 291-299 (February 2006) DOI: 10.1038/sj.jid.5700070 Copyright © 2006 The Society for Investigative Dermatology, Inc Terms and Conditions

Figure 1 Expression of RAGE, HSP47, factor XIIIa, CD31, and CD45 in human skin. Hematoxylin–eosin stainings (a) showed basic morphological changes of the specimen due to aging and/or sun-aging. Factor (b) XIIIa, (c) CD31, (d) CD45, (e/f) HSP47, and (g/h) RAGE protein expression was investigated on consecutive sections of younger sun-protected (a–h, breast), older sun-protected (i–p, breast), older sun-protected (q–x, popliteal fossa), and older sun-exposed (y–ff, face) skin. Expression of RAGE was found mainly in the basal zone of the epidermis. RAGE was overexpressed in fibroblasts and dendrocytes close to areas of older skin damaged by sun exposure. Bar=200μm. Journal of Investigative Dermatology 2006 126, 291-299DOI: (10.1038/sj.jid.5700070) Copyright © 2006 The Society for Investigative Dermatology, Inc Terms and Conditions

Figure 2 RAGE is expressed in HFFs. HFF were extracted, separated with 12% SDS-PAGE, and blotted using the monoclonal anti-RAGE antibody (1:100 dilution). Binding was visualized with a horseradish peroxidase-labeled secondary antibody. The lower band represents β-actin (monoclonal antibody, 1:5,000 dilution), which was used to control for protein loading. This is a representative blot from at least three independent experiments performed in duplicate. Journal of Investigative Dermatology 2006 126, 291-299DOI: (10.1038/sj.jid.5700070) Copyright © 2006 The Society for Investigative Dermatology, Inc Terms and Conditions

Figure 3 CML-BSA upregulates RAGE protein and mRNA expression in HFFs. (a) HFFs were treated with 50μg/ml BSA as control and 10–100μg/ml CML-BSA for 24hours in starvation medium before extraction, separation, and blotting with the anti-RAGE antibody. Bars represent mean RAGE protein expression relative to β-actin (±SD) derived from three independent experiments performed in duplicate. (b) HFFs were incubated with 50μg/ml BSA or CML-BSA for 24hours in serum-free medium before RNA extraction and determination of RAGE mRNA expression by real-time PCR. (c) Bars represent mean RAGE mRNA expression relative to glyceraldehyde-3-phosphate dehydrogenase (GAPDH) mRNA (±SD) from two independent experiments performed in quadruplicate. *P<0.05 versus BSA. Journal of Investigative Dermatology 2006 126, 291-299DOI: (10.1038/sj.jid.5700070) Copyright © 2006 The Society for Investigative Dermatology, Inc Terms and Conditions

Figure 4 TNF-α upregulates RAGE protein and mRNA expression in HFFs. (a) HFF were treated with 0–10ng/ml TNF-α for 24hours in starvation medium before extraction, separation, and blotting with the anti-RAGE antibody. Bars represent mean RAGE protein expression relative to β-actin (±SD) derived from three independent experiments performed in duplicate. (b) HFFs were incubated with 0 or 1ng/ml TNF-α for 24hours in starvation medium before RNA extraction and determination of RAGE mRNA expression by real-time PCR. (c) Bars represent mean RAGE mRNA expression relative to GAPDH (±SD) from two independent experiments performed in quadruplicate. **P<0.01 versus 0ng/ml TNF-α. Journal of Investigative Dermatology 2006 126, 291-299DOI: (10.1038/sj.jid.5700070) Copyright © 2006 The Society for Investigative Dermatology, Inc Terms and Conditions

Figure 5 The specific RAGE ligand CML-BSA modulates ECM-related gene expression in HFFs. HFFs were treated with 50μg/ml BSA or CML-BSA for 24hours in starvation medium, extracted, and expression of CTGF, TGF-β1, PC-α1(I), MMP-1, MMP-2, MMP-3, and MMP-12 mRNAs was quantified by real-time PCR relative to GAPDH mRNA. Bars represent means±SD derived from two independent experiments performed in quadruplicate. *P<0.05 versus BSA. Journal of Investigative Dermatology 2006 126, 291-299DOI: (10.1038/sj.jid.5700070) Copyright © 2006 The Society for Investigative Dermatology, Inc Terms and Conditions