Fig. 1. Inhibitory effect of plant polyphenols on the HGF-induced motility of ERK2-MDCK cells. ERK2-MDCK cells were mock-treated (control), or treated with 10 μM PD184352 or 30 μM of respective plant polyphenol for 1 h, followed by stimulation with 4 ng/ml of HGF for 12 h. Data shown are representative of three separate experiments that gave essentially the same results.

Fig. 2. Plant polyphenols do not inhibit the activation of ERK1/2, activation of Akt, or the expression of MMP-2/-9 in HT1080 cells. HT1080 cells were mock-treated (control), or treated with 10 μM PD184352, 10 μM LY294002, or 30 μM of respective plant polyphenol for 1 h. Cell lysates (30 μg of protein) were resolved by SDS–PAGE, followed by immunoblot analysis using the anti-phospho-ERK1/2 antibody, the anti-ERK1/2 antibody, the anti-phospho-Akt antibody, or the anti-Akt antibody. The expression of MMP-2/-9 was analyzed by gelatin zymography, which was performed on media conditioned by 2 × 10⁵ HT1080 cells treated with 10 μM PD184352, 10 μM LY294002, or 30 μM of respective plant polyphenol for 2 h. Data shown are representative of three separate experiments that gave essentially the same results.

Fig. 3. Inhibitory effect of plant polyphenols on the MMP-2 or MMP-9 activity. (A) Media conditioned by HT1080 cells were loaded on SDS–polyacrylamide gels that had been co-polymerized with 0.1% gelatin, separated by electrophoresis, and then gelatin zymography was performed in the presence of various concentrations of respective plant polyphenol for 24 h. The presence of MMP-2/-9 is shown as an opaque unstained band. Data shown are representative of three separate experiments that gave essentially the same results. (B) Density of each of MMP-2 bands (□) or MMP-9 bands (■) was quantified by using the NIH Image 1.62 software. Percentages were calculated by comparison with corresponding mock-treated controls. Each data point represents average ± SD of three separate experiments (two determinations/experiment).

Fig. 4. Inhibitory effect of plant polyphenols on the invasion of HT1080 cells through Matrigel. (A) The effect of plant polyphenols on the invasiveness of HT1080 cells through Matrigel-coated filters was examined in a Boyden chamber. HT1080 cells (3 × 10⁴ cells) were placed into the upper chamber of a Boyden chamber and mock-treated (control) or treated with 10 μM PD184352 or varying concentrations of plant polyphenols. After 24 h, cells that had migrated to the lower surface of the filter were fixed, stained, and photographed. Data shown are representative of three separate experiments that gave essentially the same results. (B) Values for invasion were determined by calculating the average number of migrated cells per mm² over three fields per assay. Percentages were calculated by comparison with corresponding mock-treated controls (485.4 ± 47.1 cells). Each data point represents average ± SD of three separate experiments (two determinations/experiment).

Structure and inhibitory activity of hydrolyzable tannins

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