Figure 2

THL inhibits the secretion of MMP-2, MMP-9 and uPA and the activity of ERK1/2 in cancer cells. A) Gelatin zymogram of concentrated conditioned medium from MDA-MB-231, H1299, PC-3, or CT-26 cancer cells treated with various concentrations of THL during 24-h incubation period. B) Gelatin zymogram showing direct inhibitory effect of THL on MMP-2 activity. C) Casein zymogram of concentrated conditioned medium from MDA-MB-231 or H1299 cancer cells treated with various concentrations of THL during 24-h incubation period. D) Western blot analysis shows that THL inhibits the phosphorylation of ERK1/2 but not that of AKT in CT-26 cancer cells. CT-26 cells were treated with various concentrations of THL in serum-free medium for 24 h. Western blot analysis of cell lysates was performed as described previously [66]. The blot was first probed with antibodies against phospho-ERK1/2 (cell signaling, Danvers, MA) and phospho-AKT (cell signaling), and then reprobed with antibodies against ERK1, ERK2 (Santa cruz, Santa Cruz, CA) and AKT (cell signaling). Glyceraldehyde-3-phosphate-dehydrogenase (GAPDH) served as an internal control for amounts of protein loaded on the gel.