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. 2020 Mar 20;40(3):BSR20200164. doi: 10.1042/BSR20200164

Figure 3. GREM1 increases EMT in human breast cancer cells.

Figure 3

Cells were starved overnight and stimulated with GREM1 (10 and 50 ng/ml) for an additional 48 h. RNA was analyzed by qPCR analysis (A). Cells were starved overnight and stimulated with GREM1 (50 ng/ml) for an additional 48 h. Protein lysates were subjected to immunoblot analysis (B). Each cell line was established by using lentiviral shRNA system. RNA was collected and indicated genes were quantitated by qPCR analysis (C). MDA-MB-453-shCtrl or MDA-MB-453-shGREM1 cells were seeded into 6-well plates and wounded with 1 ml pipette tip. Each cell line was incubated with vehicle or TGF-β (10 ng/ml) for 48 h. The wound closure was monitored by photography at the indicated time (D and E). Cells were starved overnight and treated with GREM1 (50 ng/ml) for 30 min. Protein lysates were subjected to immunoblot analysis (F). Cells were starved overnight and co-treated with GREM1 (50 ng/ml) and U0126 (10 μM) for 30 min (pERK and ERK) or 48 h (N-cadherin, vimentin, and Slug). Protein lysates were subjected to immunoblot analysis (G). *, P<0.05; **, P<0.01; ***, P<0.001, and NS = non-significant.