Biological Significance and Regulation of Podoplanin Expression in Oral Squamous Cell Carcinomas

Statement of the problem: Although recent advances in surgery, radiotherapy, and chemotherapy have produced modest improvements in local and regional tumor control, the 5-year survival rate for oral cancer has not improved significantly over the past several decades, remaining at about 50% to 55%. Molecular biological approaches for diagnosis and treatment may lead to better prognosis for oral cancer patients in the future. Podoplanin, a transmembrane sialomucin-like glycoprotein, is known not only to possess metastasis-promoting activity but also one of the cancer stem cell (CSC) markers for head and neck squamous cell carcinoma (HNSCC). However, the biological significance and regulatory mechanism of podoplanin expression in oral squamous cell carcinoma (OSCC) remains unknown. The present study examined the potential mechanism of podoplanin and its association with epithelial-mesenchymal transition (EMT) and CSC.

Materials and methods: We immunohistochemically examined the podoplanin expression in surgically resected clinical specimens from 80 OSCC patients. Western blot analysis and quantitative RT-PCR were also employed to investigate the regulatory mechanism of podoplanin expression and its significance as a stemness marker using 2 OSCC cell lines in vitro.

Methods of data analysis: The statistical significance of the differences in data between each group was determined by applying Student’s t-test. P values of less than 0.05 were considered significant.

Results: Immunohistochemical analysis of OSCC specimens revealed high podoplanin expression in 70% of OSCC cases with localization primarily in the basal layer of the squamous cancer nest. The expression was inversely correlated with the squamous cell differentiation. In vitro analysis of OSCC cell lines revealed that podoplanin expression was down-regulated by treatment with histone deacetylase (HDAC) inhibitors such as sodium butyrate and trichostatin, whereas expressions of differentiation markers such as E-cadherin and Cytokeratin 10 were up-regulated by the same HDAC treatment. Moreover, knockdown of podoplanin expression by siRNA induced increased expressions of E-cadherin and CK 13 and suppressed cell growth in vitro. In addition, transforming growth factor-β (TGF-β) significantly enhanced podoplanin expression in OSCC cell lines in proportion with increased phosphorylation of Smad2. Conversely, TGF-β type I receptor inhibitor (SB431542) significantly reduced such podoplanin expression at both the protein and mRNA levels. Most importantly, TGF-β also induced epithelial-mesenchymal transition (EMT) showing decreased E-cadherin expression and increased vimentin expression.

Conclusions: These results suggest that podoplanin expression is both positively and negatively regulated by TGF-β receptor/Smad signaling pathway and epigenetic mechanisms such as histone acetylation and that podoplanin may play some role in suppressing squamous differentiation and maintaining cancer stemness or induction of EMT in OSCC cells. Podoplanin should therefore be a new target for molecular therapy against OSCC.

References:

1. Yuan P, Temam S, El-Naggar A, et al: Overexpression of podoplanin in oral cancer and its association with poor clinical outcome. Cancer. 2006; 107: 563–569.

2. Maseki S, Ijichi K, Tanaka H, et al: Acquisition of EMT phenotype in the gefitinib-resistant cells of a head and neck squamous cell carcinoma cell line through Akt/GSK-3β/snail signalling pathway. Br J Cancer. 2012; 106: 1196 – 1204.