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. 2019 Jun 7;20(11):2792. doi: 10.3390/ijms20112792

Figure 2.

Figure 2

Integrin α5 and integrin β1 silencing alleviated fibronectin effects in 786-O cells. (A) 786-O cells were seeded onto fibronectin (0 and 50 μg/mL)-coated 96-well plates. Twenty-four hours later, cell growth was measured by MTS reduction assay. (B) 786-O cells were seeded onto fibronectin (0 and 50 μg/mL)-coated Transwell inserts and subjected to Transwell migration assay for 24 h. The lower chambers were filled with DMEM containing 10% FBS. (C) 786-O cells were seeded onto Transwell inserts and subjected to Transwell migration assay for 24 h. The lower chambers were filled with DMEM containing fibronectin (0 or 50 μg/mL). (D) 786-O cells were first incubated with indicated IgG (5 μg/mL) for 30 min before seeding to the Transwell inserts for migration assay (24 h). The lower chambers were filled with DMEM containing fibronectin (0 or 50 μg/mL). (E) 786-O cells were transfected with control siRNA, integrin α5 siRNA, and integrin β1 siRNA for 48 h. Proteins were extracted and subjected to Western blot analysis with indicated antibodies. Representative blots are shown. (F) The resultant transfected cells were seeded onto six-well plates for 24 h. When confluence was reached, cell movement was evaluated by a wound-healing assay for 16 h in the presence of 0.5% FBS. Representative photomicrographs are shown. Bar graphs showed relative wound closure among groups. (G) The resultant transfected cells were seeded onto Transwell inserts and subjected to Transwell migration assay for 24 h. The lower chambers were filled with DMEM containing 10% FBS. (H) The resultant transfected cells were seeded onto fibronectin (0 and 50 μg/mL)-coated Transwell inserts and subjected to Transwell migration assay for 24 h. The lower chambers were filled with DMEM containing 10% FBS. (I) The resultant transfected cells were seeded onto Transwell inserts and subjected to Transwell migration assay for 24 h. The lower chambers were filled with DMEM containing fibronectin (0 and 50 μg/mL). Bar graphs show quantitative results among groups and the value in fibronectin (0 μg/mL)/control siRNA group was defined as 100% (A–D, G–I). * p < 0.05 vs. fibronectin (0 μg/mL)/control siRNA group and # p < 0.05 vs. fibronectin (50 μg/mL)/control siRNA group, n = 3.