Fig. 2. EGCG inhibits cell growth.

The effect of EGCG on cell growth was estimated using the MTS assay kit, as described in “Supplemental Methods.” (A) The expression of Pin1 in Pin1KO and Pin1WT MEFs. (B) Growth comparison of Pin1KO and Pin1WT MEFs. Cells (1×10³/well) were seeded on 96-well plates then incubated for 24, 48, 72 or 96 h and growth was estimated using the MTS assay. (C) Effect of various concentrations of EGCG (0-40 μM) on the growth of Pin1KO and Pin1WT MEFs. Cells (1×10³/well) were seeded on 96-well plates then incubated in a 37°C/5% CO₂ incubator for 24 h. Cells were treated with EGCG for 24 or 48 h and growth was then estimated by MTS assay. (D) Expression of Pin1 and Neu in Neu/Pin1WT and Neu/Pin1KO MEFs. (E) Growth comparison of Neu/Pin1WT and Neu/Pin1KO MEFs. Cells (1×10³/well) were seeded on 96-well plates then incubated in a 37°C/5% CO₂ incubator for 24, 48 or 72 h and cell growth was then estimated by MTS assay. (F) Effect of EGCG on the growth of Pin1KO and Pin1WT MEFs. Cells (1×10³/well) were seeded on 96 well plates and incubated for 24 h in a 37°C/5% CO₂ incubator. Cells were treated with EGCG for 24 or 48 h and then growth was estimated by MTS assay. Effect of EGCG on the growth of Neu/Pin1KO MEFs re-expressing (G) Mock, (H) Pin1WT, or (I) mutant Pin1R17A. Cells were transfected with a Mock, Pin1WT or mutant Pin1R17A plasmid. After zeocin selection, cells (1×10³/well) were seeded on 96-well plates and incubated for 24 h in a 30°C/5% CO₂ incubator. Cells were treated with various doses of EGCG for different times and growth was then estimated by MTS assay. Data are expressed as means ± S.D. and the asterisk (*) indicates a significant (p <0.001) decrease in proliferation of Pin1WT or Neu/Pin1WT MEFs compared to Pin1KO or Neu/Pin1KO MEFs or the untreated group compared to treated groups (20 or 30 μM EGCG).