Effect of hydrogel stiffness on morphology and gene expression pattern of CD44^high oral squamous cell carcinoma cells

The stiffness of extracellular matrix (ECM) has been associated with tumor growth, phenotypic plasticity, and invasion through modulation of the intracellular signaling pathway. However, the effect of ECM stiffness on oral cancer stem cells (CSCs) has not been fully elucidated. Therefore, we preliminarily investigated changes in phenotype and gene expression in CD44 positive-oral squamous cell carcinoma (OSCC) cells (i.e., CD44^high OM-1 cells) that were cultured on laminin-coated hydrogel with various degrees of stiffness. Mesenchymal-like morphology was observed when cells were cultured on 4.0 kPa laminin-coated hydrogel; amoeboid-like morphology was observed when cells were cultured on 1.0 kPa and 0.5 kPa laminin-coated hydrogel. These results indicated that CD44^high OM-1 cells underwent mesenchymal to amoeboid transition (MAT) when cultured on laminin-coated softer hydrogel. E-cadherin and ESA mRNA expression levels were significantly reduced in CD44^high OM-1 cells cultured on 0.5 and 1.0 kPa laminin-coated hydrogel, compared with their levels in control cells cultured in laminin-coated dishes. Significant changes in CD44 mRNA expression were not found in CD44^high OM-1 cells that were cultured on different stiff hydrogels, compared with expression in control cells. Microarray analysis revealed that expression of cofilin, an intracellular actin-modulating protein, was increased by 8.19-fold in amoeboid-like CD44^high OM-1 cells, compared with mesenchymal-like CD44^high OM-1 cells; this suggested that cofilin was associated with MAT in CD44^high OSCC cells. Further studies are needed to clarify the relationship between cofilin and invasion ability in CD44^high amoeboid-like OSCC cells.