Neutrophil gelatinase–associated lipocalin mediates 13-cis retinoic acid–induced apoptosis of human sebaceous gland cells
Amanda M. Nelson, … , Wenlei Liu, Diane M. Thiboutot
Amanda M. Nelson, … , Wenlei Liu, Diane M. Thiboutot
Published March 3, 2008
Citation Information: J Clin Invest. 2008;118(4):1468-1478. https://doi.org/10.1172/JCI33869.
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Neutrophil gelatinase–associated lipocalin mediates 13-cis retinoic acid–induced apoptosis of human sebaceous gland cells

Abstract

13-cis retinoic acid (13-cis RA; also known as isotretinoin) is the most potent agent available for treatment of acne. It is known that the drug induces apoptosis in cells cultured from human sebaceous glands, but its mechanism of action has not been determined. In this study, skin biopsies were taken from 7 patients with acne prior to and at 1 week of treatment with 13-cis RA. TUNEL staining confirmed that 13-cis RA induced apoptosis in sebaceous glands. Transcriptional profiling of patient skin and cultured human sebaceous gland cells (SEB-1 sebocytes) indicated that lipocalin 2 was among the genes most highly upregulated by 13-cis RA. Lipocalin 2 encodes neutrophil gelatinase–associated lipocalin (NGAL), which functions in innate immune defense and induces apoptosis of murine B lymphocytes. Increased immunolocalization of NGAL was noted in patients’ sebaceous glands following treatment with 13-cis RA, and recombinant NGAL induced apoptosis in SEB-1 sebocytes. Furthermore, apoptosis in response to 13-cis RA was inhibited in the presence of siRNA to lipocalin 2. These data indicate that NGAL mediates the apoptotic effect of 13-cis RA and suggest that agents that selectively induce NGAL expression in sebaceous glands might represent therapeutic alternatives to the use of 13-cis RA to treat individuals with acne.

Authors

Amanda M. Nelson, Wei Zhao, Kathryn L. Gilliland, Andrea L. Zaenglein, Wenlei Liu, Diane M. Thiboutot

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Figure 6

siRNA to LCN2 specifically knocks down LCN2 expression in SEB-1 sebocytes in the presence of 13-cis RA.

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siRNA to LCN2 specifically knocks down LCN2 expression in SEB-1 sebocyte...
SEB-1 sebocytes (2 × 106 cells per 100 μl reaction mixture) were nucleofected with 1 μg siCONTROL, GAPDH (as a specificity control), or LCN2 siRNA. 13-cis RA (0.1 μM) was added 24 hours after nucleofection, and cells were incubated for 48 and 72 hours. Extent of siRNA knockdown of gene expression was verified by QPCR and western blotting for LCN2 and GAPDH after 13-cis RA treatment. GAPDH and LCN2 gene expression were successfully inhibited in their respective samples. (A) QPCR analysis of LCN2 and GAPDH mRNA levels at 48 hours of 0.1 μM 13-cis RA treatment. The expression of LCN2 mRNA was successfully decreased 15-fold by the LCN2 siRNA compared with siCONTROL, whereas expression of GAPDH was minimally affected by siRNA to LCN2. GAPDH mRNA expression was decreased 4-fold by the specific GAPDH siRNA when compared with siCONTROL, whereas siRNA to GAPDH had minimal effects on expression of mRNA for LCN2. Data represent mean ± SEM; n = 6. *P < 0.05 as determined by REST-XL program. (B) Western analysis of GAPDH and NGAL protein levels. NGAL protein expression is decreased to undetectable levels by western blotting at 48 and 72 hours of 0.1 μM 13-cis RA treatment with LCN2 siRNA. Representative blots of 4 independent experiments are shown.