Abnormally stiff substrates have been shown to trigger cancer progression. However, the detailed molecular mechanisms underlying this trigger are not clear. In this study, we cultured T84 human colorectal cancer cells on plastic dishes to create a stiff substrate or on collagen-I gel to create a soft substrate. The stiff substrate enhanced the expression of matrix metalloproteinase-7 (MMP-7), an indicator of poor prognosis. In addition, we used polyacrylamide gels (2, 67 and 126 kPa) so that the MMP-7 expression on the 126-kPa gel was higher compared with that on the 2-kPa gel. Next, we investigated whether yes-associated protein (YAP) affected the MMP-7 expression. YAP knockdown decreased MMP-7 expression. Treatment with inhibitors of epidermal growth factor receptor (EGFR) and myosin regulatory light chain (MRLC) and integrin-α2 or integrin-β1 knockdown downregulated MMP-7 expression. Finally, we demonstrated that YAP, EGFR, integrin-α2β1 and MRLC produced a positive feedback loop that enhanced MMP-7 expression. These findings suggest that stiff substrates enhanced colorectal cancer cell viability by upregulating MMP-7 expression through a positive feedback loop.

Activating transcription factor 5 (ATF5) is a member of the ATF/cAMP response element-binding protein family. Our research group recently revealed that ATF5 expression increases the invasiveness of human lung carcinoma cells. However, the effects of ATF5 on the invasive potential of other cancer cells lines remain unclear.Therefore, in this study, we investigated the role of ATF5 in the invasive activity of diverse human cancer cell lines. Invasiveness was assessed using Matrigel invasion assays. ATF5 knockdown resulted in decreased invasiveness in seven of eight cancer cell lines tested. These results suggest that ATF5 promotes invasiveness in several cancer cell lines. Furthermore, the roles of ATF5 in the invasiveness were evaluated in three-dimensional (3D) culture conditions. In 3D collagen gel, HT-1080 and MDA-MB-231 cells exhibited high invasiveness, with spindle morphology and high invasion speed. In both cell lines, knockdown of ATF5 resulted in rounded morphology and decreased invasion speed. Next, we showed that ATF5 induced integrin-2 and integrin-1 expression and that the depletion of integrin-2 or integrin-1 resulted in round morphology and decreased invasion speed. Our results suggest that ATF5 promotes invasion by inducing the expression of integrin-2 and integrin-1 in several 3 human cancer cell lines. ATF5 regulates cell morphology in a 3D collagen gel via integrin-21 activity. Keywords

Astrocytes, which can be obtained from neural stem cells (NSCs) by adding serum and/or recombinant proteins in culture media or by passaging NSCs repeatedly, are expected to be applicable in regenerative medicine for the treatment of neurodegenerative diseases. However, astrocytes obtained using existing methods are costly and have poor quality. The stiffness of culture surfaces has been reported to affect astrocytic differentiation of adult NSCs. However, the influence of surface stiffness on astrocytic differentiation of embryonic NSCs has not yet been reported. In this study, we showed that astrocytic differentiation of embryonic NSCs was increased on soft surfaces (1 kPa and 12 kPa) compared with the NSCs on stiff surfaces (2.8 GPa) in serum-free condition. Furthermore, di-phosphorylated myosin regulatory light chain (PP-MRLC) was decreased in embryonic NSCs cultured on the soft surfaces than the cells on the stiff surfaces. Additionally, astrocytic differentiation of embryonic NSCs was induced by a Ras homolog associated kinase (ROCK) inhibitor, which decreased PP-MRLC in NSCs. These results suggest that decreasing the PP-MRLC of embryonic NSCs on soft surfaces or treating NSCs with a ROCK inhibitor is a good method to prepare astrocytes for application in regenerative medicine.