Background/Purpose: Slug, a Snail family of zinc finger transcription factor, plays a critical role in tumor proliferation, invasion and metastasis. We have previously shown that Slug is overexpressed in RA synovial tissue and suppression of Slug facilitates apoptosis of RA FLS. However, the precise mechanism through which Slug is induced in RA FLS remains unclear. Smoking is an important environmental risk factor for RA and influences RA disease susceptibility and severity. Benzo(a)pyrene (BaP) is a polycyclic aromatic hydrocarbon found in cigarette smoke. BaP can be metabolized to active compounds that induce the production of reactive oxygen species. The effect of Bap on Slug expression in RA FLS has not been studied thus far.

The present study was undertaken to investigate the biological effects of BaP on the expression of Slug, signaling pathway through which Slug is expressed, and the effect of BaP/Slug on the invasive properties of RA FLS.

Methods:  Expression of Slug was measured by real-time PCR following stimulation of FLS with different concentrations of BaP, EGF, H₂O₂, TGF-β, and TNF-α (n=7, each). Phosphorylation of the key enzymes in the signal transduction pathway was analyzed by Western blot. Inhibitors targeting the PI3K/Akt/mTOR pathway were used to confirm critical signaling pathway for Slug expression (n=7). An in vitro cell invasion assay was performed using RA FLS treated with Slug siRNA or with control siRNA (n=4).

Results:  Slug expression was significantly increased following treatment with BaP (4μM, p=0.0014), EGF (100ng/ml, p=0.027) and H₂O₂ (0.25mM, p=0.029), but not with TGF-β and TNF-α. Stimulation with BaP and EGF induced phosphorylation of Akt kinase activity, but no significant change was observed in ERK, JNK and p38 activity. Slug mRNA expression by BaP and EGF decreased significantly following treatment with PI3K/Akt/mTOR inhibitors (p=0.026, p=0.002, respectively). Slug siRNA treatment induced a significant reduction in the invasive function of FLS compared to those treated with control siRNA (p=0.029). In addition, stimulation with BaP increased invasive function of control siRNA-treated FLS (p=0.01), but no such change was observed in Slug siRNA-treated FLS.

Conclusion: Our data show that BaP, one of major toxic components in cigarette smoke, induce Slug expression in RA FLS through PI3K/Akt/mTOR pathway, and may regulate invasive properties of RA FLS. This mechanism may provide a novel explanation for the increased RA susceptibility and severe clinical phenotype in those who smoke.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/slug-is-induced-by-benzoapyrene-and-egf-through-pi3kaktmtor-pathway-and-is-closely-involved-in-the-regulation-of-the-invasive-properties-of-fls-in-rheumatoid-arthritis/