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. Author manuscript; available in PMC: 2013 Apr 4.
Published in final edited form as: Oncogene. 2011 Dec 5;31(31):3569–3583. doi: 10.1038/onc.2011.547

Figure 7.

Figure 7

Effects of MFGE8 on invasion of HER2-overexpressing mammary cells. (a) MCF10A and MCF10A/HER2 cells were serum starved for 12 h, stimulated for 10 min with MFGE8 (100 ng/ml) and cell extracts immunoblotted with the indicated antibodies, including an antibody the phosphorylated form of ERK. (b, c) MCF10A/HER2 cells (50 000 cells) were plated on Matrigel-coated Transwell chambers and incubated for 12 h in the absence or presence of EGF (20 ng/ml) and/or MFGE8 (100 ng/ml). Cells that invaded into the lower side of the Transwell chambers were stained using crystal violet, photographed and counted. (c) Quantification of the cell migration data shown in (b). Bars represent s.d. of triplicates. (d) Phase microscopy analysis of MCF10A/HER2 acini plated in Matrigel and grown for 10 days in the absence or presence of EGF (20 ng/ml), ectopic MFGE8 (10 ng/ml) or a blocking antibody to integrin αV (bar, 50 μm). (e) Monolayers of MCF10A/HER2 cells were stably infected with short hairpin RNA (shRNA) particles, either control (C) or shRNA targeting MFGE8 (clones 74, 77 and 78). Cell clones were tested for MFGE8 expression using immunoblotting. (f, g) MCF10A/HER2 cells expressing the indicated shRNAs were cultured for 10 days in Matrigel. Phase contrast micrographs are shown (bar, 50 μm), along with morphometric analysis of acini. Shown are averages±s.d. values of triplicates.