Figure 7

Effects of Tβ10 silence on ERK1/2 phosphorylation and the expression of EGR1, Snail and MMPs in KKU-M055 and KKU-M214 cell lines. (A) M055 sh-Tβ10 cells and M214 sh-Tβ10 cells as well as their vector control cells were grown in the complete medium for 24 h. Phosphorylated form and total form of ERK1/2 were determined by Western blot analysis. β-actin was used as a loading control. Intensity of the immunoreactive bands was measured by ImageJ software. This assay was repeated with pretreatment of a Ras-GTPase inhibitor (FPT inhibitor III, 100 μM, 2 h). (B) EGR1 and Snail protein levels in M055 sh-Tβ10 cells and M214 sh-Tβ10 cells as well as their vector control cells were determined by Western blot. Histone H1 was also included as a loading control. Intensity of the immunoreactive bands was measured by ImageJ software. This assay was repeated with pretreatment of a Ras-GTPase inhibitor (FPT inhibitor III, 100 μM, 2 h). The mRNA levels of Tβ10, EGR1 and Snail in M055 sh-Tβ10 cells (C) and M214 sh-Tβ10 cells (D) as well as their vector control cells were determined by real time RT-PCR analysis. Also, the mRNA levels of MMP3, MMP7 and MMP9 in M055 sh-Tβ10 cells (E) and M214 sh-Tβ10 cells (F) as well as their vector control cells were determined by real time RT-PCR analysis. All data were normalized to β-actin mRNA levels in these cells.