Role of slug in TGF-β1 mediated MMP-9 expression in human oral squamous cell cancer

Abstract

21000 Background: Oral squamous cell cancer (OSCC) causes significant morbidity and mortality as a result of loco-regional invasion and distant metastasis. Invasive OSCC tumors demonstrate increased expression of matrix metalloproteinases (MMPs), including MMP-9. TGF-β1, which promotes MMP-dependent invasion, is also frequently overexpressed in OSCC. Recent reports indicate that TGF-β1 can signal through Snail family of transcription factors, including Slug, which are key regulators of epithelial to mesenchymal transition. Methods: To examine the relationship between TGF-β1 and MMP-9 in human OSCC tumors, phospho-Smad2 and MMP-9 expression was quantified in an oral cancer tissue microarray, and then the relationship determined using Pearson correlation coefficient. To understand the role of Slug in TGF-β1-mediated MMP-9 expression, premalignant oral keratinocyte cell lines (OKF4 and OKF6) and malignant OSCC lines (SCC25 and UMSCC1) were treated with TGF-β1 (10 ng/ml) and analyzed for Slug and MMP-9 expression at the message and protein levels. The specific role of Slug was determined by transfecting OSCC cells with Slug specific siRNA or control siRNA, treating with TGF-β1 for 24h, and determining the effect on MMP-9 expression. Results: There was a statistically significant correlation between phospho-Smad2 and MMP- 9 expression in human OSCC samples, with a Pearson correlation coefficient of 0.4392 (p=0.0007). In vitro, treatment of premalignant and OSCC cells with TGF-β1 induced both Slug and MMP-9 expression at the message and protein level. Interestingly, transfection of Slug siRNA in OKF4 and OKF6 cells increased both basal and TGF-β1-mediated MMP-9 expression, while transfection of Slug siRNA in SCC25 and UMSCC1 cells decreased basal and TGF-β1-mediated MMP-9 expression. Conclusions: Overall, our data indicate that TGF-β1 may regulate MMP-9 expression in human oral squamous cell cancer. Additionally, our data indicate a role for Slug in TGF-β1-mediated MMP-9 expression in oral cancer cells. Future experiments will examine the mechanism by which Slug differentially regulates MMP-9 expression in premalignant and malignant OSCC cells. No significant financial relationships to disclose.