Figure 4

Effects of stable silence of Tβ10 on cell migration and monolayer wound healing in KKU-M055 cell line. Stable cell lines expressing Tβ10 shRNA and control vector (sh-vector) were generated in KKU-M055 cell line. (A) All clones of control and Tβ10 silencing cells were confirmed by real-time RT-PCR. β-actin was used as an internal control. Arrows indicate selected clone for use in subsequent experiments; and insert pictures show immunocytochemistry results of selected clones. (B) Cell migration of sh-Tβ10 and sh-vector cells was studied by the modified Boyden chamber assay at 24 h incubation. Bar graphs represent fold change or percentage of control. n = 3; *P<0.05 versus the control. M055 sh-vector and M055 sh-Tβ10 cells did not express eGFP gene. The green fluorescent signal in these cells was from Calcein-AM staining for the migration assay. (C) Wound healing assay was determined in sh-Tβ10 and sh-vector cells. Representative images were taken from the same field at 0, 24 and 40 h. (D) M055-sh-vector-GFP and M055-sh-Tβ10-GFP cells were pre-treated with a Ras-GTPase inhibitor (FPT inhibitor III) for 2 h. Wound healing assay was performed. Representative images were taken from the same field at 0, 20, and 24 h.