FIGURE 8:

LARG and GEF-H1 overexpression after EMT partially rescues cell stiffness and stiffness response and attenuates migration and invasion. (A) NMuMGs were treated with 100 pM TGF-β for 48 h. Cell were transfected with plasmid containing LARG-GFP or GEF-H1-GFP for the final 16–24 h of treatment. Quantifications are shown normalized to β-actin and relative to control, confirming protein expression. (B) Average cell stiffness (G₁) for cells transfected with GFP vector control (n = 65), TGF-β and GFP vector control (n = 55), and GFP-DNA constructs to overexpress GEF-H1 (n = 32) or LARG (n = 54). *p < 0.05 denotes stiffness difference relative to cells treated with TGF-β and GFP vector control. Error bars represent SEM; data were collected from three independent experiments. (C) Average force response at pulls 2 (G₂/G₁) and 8 (G₈/G₁) for NMuMG cells transfected with GFP vector control (n = 25), TGF-β and GFP vector control (n = 15), and GFP-DNA constructs to overexpress GEF-H1 (n = 16) or LARG (n = 23). ^#p < 0.05 denotes stiffness difference from G₁; *p < 0.05 denotes stiffness response difference relative to cells treated with TGF-β and GFP vector control. Error bars represent SEM; data were collected from three independent experiments. (D, E) Average migration and invasion for cells treated with TGF-β and transfected with plasmid containing empty vector control (GFP), LARG-GFP, or GEF-H1 GFP for the final 16–24 h of treatment and sorted by flow cytometry for GFP expression. Quantifications given as fold migration or invasion relative to control (n = 3). *p < 0.05.