Role of miR-218-GREM1 axis in epithelial-mesenchymal transition of oral squamous cell carcinoma: An in vivo and vitro study based on microarray data

Abstract

Oral squamous cell carcinoma (OSCC) is a prevalent cancer that develops in the head and neck area and has high annual mortality despite optimal treatment. microRNA-218 (miR-218) is a tumour inhibiting non-coding RNA that has been reported to suppress the cell proliferation and invasion in various cancers. Thus, our study aims to determine the mechanism underlying the inhibitory role of miR-218 in OSCC. We conducted a bioinformatics analysis to screen differentially expressed genes in OSCC and their potential upstream miRNAs. After collection of surgical OSCC tissues, we detected GREM1 expression by immunohistochemistry, RT-qPCR and Western blot analysis, and miR-218 expression by RT-qPCR. The target relationship between miR-218 and GREM1 was assessed by dual-luciferase reporter gene assay. After loss- and gain-of-function experiments, OSCC cell proliferation, migration and invasion were determined by MTT assay, scratch test and Transwell assay, respectively. Expression of TGF-β1, Smad4, p21, E-cadherin, Vimentin and Snail was measured by RT-qPCR and Western blot analysis. Finally, effects of miR-218 and GREM1 on tumour formation and liver metastasis were evaluated in xenograft tumour-bearing nude mice. GREM1 was up-regulated, and miR-218 was down-regulated in OSCC tissues, and GREM1 was confirmed to be the target gene of miR-218. Furthermore, after up-regulating miR-218 or silencing GREM1, OSCC cell proliferation, migration and invasion were reduced. In addition, expression of TGF-β signalling pathway-related genes was diminished by overexpressing miR-218 or down-regulating GREM1. Finally, up-regulated miR-218 or down-regulated GREM1 reduced tumour growth and liver metastasis in vivo. Taken together, our findings suggest that the overexpression of miR-218 may inhibit OSCC progression by inactivating the GREM1-dependent TGF-β signalling pathway.

Keywords: Gremlin1 gene; TGF-β signalling pathway; epithelial-mesenchymal transition; liver metastases nodes; microRNA-218; oral squamous cell carcinoma.