Expression of Lipogenic Factors Galectin-12, Resistin, SREBP-1, and SCD in Human Sebaceous Glands and Cultured Sebocytes Wesley J. Harrison, Jonathan.

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Expression of Lipogenic Factors Galectin-12, Resistin, SREBP-1, and SCD in Human Sebaceous Glands and Cultured Sebocytes Wesley J. Harrison, Jonathan J. Bull, Holger Seltmann, Christos C. Zouboulis, Michael P. Philpott Journal of Investigative Dermatology Volume 127, Issue 6, Pages 1309-1317 (June 2007) DOI: 10.1038/sj.jid.5700743 Copyright © 2007 The Society for Investigative Dermatology, Inc Terms and Conditions

Figure 1 RT-PCR analysis. RT-PCR analysis of galectin-12, resistin, SREBP-1a, SREBP-1c, and SCD in human SZ95 sebocytes, pre-adipocytes, N/TERT keratinocytes, and whole human skin. Agarose/ethidium bromide gel analysis of RT-PCR amplicons of galectin-12, resistin, SREBP-1a, SREBP-1c, and SCD in total RNA extracts from human SZ95 sebocytes, pre-adipocytes, N/TERT keratinocytes, and whole human skin (n=3). Journal of Investigative Dermatology 2007 127, 1309-1317DOI: (10.1038/sj.jid.5700743) Copyright © 2007 The Society for Investigative Dermatology, Inc Terms and Conditions

Figure 2 Immunohistochemical analysis reveals that C/EBPα, galectin-12, resistin, SREBP-1, and SCD are located within specific cell populations in human scalp skin. Immunoreactivity in the distal human hair follicle is shown for (a) C/EBPα, (d) galectin-12, (g) resistin, (j) SREBP-1, and (m) SCD. Immunoreactivity in the interfollicular epidermis is shown for (b) C/EBPα, (e) galectin-12, (h) resistin, (k) SREBP-1, and (n) SCD. Immunoreactivity in the sebaceous gland is shown for (c) C/EBPα, (f) galectin-12, (i) resistin, (l) SREBP-1, and (o) SCD. (p–r) Absence of specific immunolocalization observed when the primary antibody is pre-incubated with a specific blocking peptide. Bars=(a, d, g, j, m, p) 200μm; (b, e, h, k, n, q) 50μm; (c, f, i, l, o, r) 100μm, (n=6). Immunolocalization for all samples is summarized in Table 2. Journal of Investigative Dermatology 2007 127, 1309-1317DOI: (10.1038/sj.jid.5700743) Copyright © 2007 The Society for Investigative Dermatology, Inc Terms and Conditions

Figure 3 Immunocytochemical analysis. Immunocytochemical analysis reveals subcellular localization of C/EBPα, galectin-12, resistin, SREBP-1, and SCD in cultured human SZ95 sebocytes. Immunocytostaining of SZ95 sebocytes with (a) anti-C/EBPα, (b) anti-galectin, (c) anti-resistin, (d) anti-SREBP-1, (e) anti-SCD antibodies as used for immunohistochemistry, and (f) absence of specific immunolocalisation observed when the primary antibody is pre-incubated with a specific blocking peptide (n=6). Bar=50μm. Journal of Investigative Dermatology 2007 127, 1309-1317DOI: (10.1038/sj.jid.5700743) Copyright © 2007 The Society for Investigative Dermatology, Inc Terms and Conditions

Figure 4 Western blot analysis. Western blot analysis shows the pattern of expression of C/EBPα, galectin-12, resistin, SREBP-1, and SCD in SZ95 sebocytes, pre-adipocytes, N/TERT keratinocytes, and whole skin lysates. Using the same antibodies used for immunohistochemistry and immunocytochemistry C/EBPα, galectin-12, resistin, SREBP-1, and SCD isoforms were detected in SZ95 sebocytes, human pre-adipocytes, N/TERT keratinocytes, and whole skin lysates. Glyceraldehyde-3-phosphate dehydrogenase was detected to indicate equal protein loading (n=6). Journal of Investigative Dermatology 2007 127, 1309-1317DOI: (10.1038/sj.jid.5700743) Copyright © 2007 The Society for Investigative Dermatology, Inc Terms and Conditions

Figure 5 Western blot analysis shows that the overexpression of C/EBPα does not increase the expression of resistin, SCD, PPARγ, or involucrin. Increased expression of 42 and 30kDa C/EBPα isoforms detected in SZ95 sebocytes transfected with pCMV-C/EBPα (lane 3) and 30kDa isoform in SZ95 sebocytes transfected with pCMV-30kDa (lane 4), compared with untransfected (lane 1) or pCMV-STOP-transfected (lane 2) SZ95 sebocytes. No changes in expression level were detected for resistin, SCD, PPARγ, or involucrin (n=3). Journal of Investigative Dermatology 2007 127, 1309-1317DOI: (10.1038/sj.jid.5700743) Copyright © 2007 The Society for Investigative Dermatology, Inc Terms and Conditions