Rosiglitazone inhibits RANTES expression by human epidermal keratinocytes

ARTICLE

Auteur(s) : Min Zhang¹, Jing Chen², Zaipei Guo¹, Li Li¹, Yizhi Zhang¹, Jianwei Li³

¹Department of Dermatovenereology, West China Hospital, Sichuan University, Chengdu, China
²Department of Pathology, Immunology, and Laboratory Medicine, University of Florida College of Medicine, Gainesville, FL, USA
³Department of Endocrinology, West China Hospital, Sichuan University, number 37 Guoxue Lane, Chengdu, 610041, Sichuan province, China

The etiology of psoriasis is not known, it can be treated but not cured [1]. The chemokine RANTES (regulated upon activation, normal T cell expressed and secreted), which is secreted by keratinocytes and induces chemotaxis and activation of T cells, may play a pathogenic role. Thiazolidinediones have been shown to have anti-psoriatic effects after either oral [2] or topical [3] administration. We questioned whether, and by which signaling pathways, these drugs might affect the production of RANTES by epidermal keratinocytes.

We used the immortalized human epidermal keratinocyte cell line HaCaT (China Center for Type Culture collection, Wuhan, China), and first stimulated it to produce RANTES, using tumor necrosis factor alpha (TNF-α) and interferon gamma (IFN-γ). RANTES secretion, initially low, was increased moderately by 20 ng/mL TNF-α, and strongly when TNF-α was increased to 50 ng/mL or combined with 20 ng/mL IFN-γ, with which it is known to act synergistically (figure 1A). We then applied the drug rosiglitazone (Taiji Industry, Chongqing, China) as an inhibitor. We used real-time PCR to measure the effect of rosiglitazone on RANTES mRNA, and ELISA to measure the effect on RANTES protein levels (RANTES ELISA kit, Biosource, USA). The expression of RANTES was inhibited, in a dose-dependent manner, by rosiglitazone at a range of concentrations between 12.5 and 50 μM; at 25 μM rosiglitazone RANTES post-stimulatory mRNA levels fell by 50% (p < 0.02) (figure 1B).

We next examined which signal transduction pathways might be mediating the observed inhibition of RANTES expression. Transcriptional control of RANTES in keratinocytes is mediated through six, variably expressed, short regulatory elements that are responsive to both STAT1 (signal transducer and activator of transcription 1) and nuclear factor kappa B (NFκB) [4-6]. We used western blotting to measure protein levels of both STAT1 (rabbit anti-human-STAT1 Ab, Boster Biotechnology, Wuhan, China) and NFκBp65 (mouse anti-human-NFκBp65 mAb, Santa Cruz Biotechnology, USA) in HaCaT cell nuclear extracts (Nuclear extract kit, KeyGen Biotechnology, Nanjing, China). Nuclear levels of both proteins, initially low, were strongly increased after 0.5 h stimulation by TNF-α and IFN-γ (p < 0.02). Rosiglitazone (25 μM) inhibited the effect on STAT1 by 25.42% (p < 0.01), but had no effect on NFκBp65 (p > 0.05).

In summary, we have shown that the antipsoriatic drug rosiglitazone appears to inhibit the production of RANTES by human keratinocytes, an effect that may be mediated by an inhibition of nuclear translocation of STAT1. We intend to extend these studies by measuring the phosphorylation of STAT1 under the conditions we have described here, and to compare the DNA-binding abilities of STAT1 and NFκB in normal and psoriatic primary keratinocytes.

Acknowledgements

References

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