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Figure 2


Fig. 2. Wounding-induced but not EGFR-ligand-induced AKT and ERK phosphorylation is ATP-dependent. (A) Growth factor-starved THCE cells were pretreated with a combination of apyrase and adenosine deaminase (5 units/ml each) for 30 minutes, and were then either extensively injured or stimulated with HB-EGF (50 ng/ml) as a positive control. After removing debris, cells were further incubated in fresh KBM containing the combination of apyrase and adenosine deaminase or HB-EGF for 2 minutes. Cell lysates were subjected to western blotting with antibodies against phosphorylated AKT (P-AKT), phosphorylated ERK 1/2 (P-ERK), AKT and ERK2. (B) Growth factor-starved THCE cells were pretreated with the apyrase and adenosine deaminase mix as in A and wounded with a 0.1-10 µl pipette tip. Cells were then fixed 15 minutes post wounding and incubated with antibody against P-ERK, followed by visualization with FITC-conjugated secondary antibody. Corresponding nuclear staining was performed with DAPI. Arrows indicate nuclear staining of P-ERK at the wound edge (arrowhead).





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