Figure 2. PolyI:C suppresses oncogenicity, cellular motility, migration and invasion of A549 and NCI-H292 cells.

(A) Soft colony formation of lung cancer cells treated with polyI:C for assessment of anchorage-independent growth over 12 days. Cells were treated with 10 μg/mL polyI:C consecutively every 3 days over a period of 12 days and the viability of the colony formed in the soft agar was measured by Alamar blue assay. Control NT cells were treated with PBS (without polyI:C). Enumerated soft agar colonies are presented as percent polyI:C-treated cells relative to NT cells. (B) 3D matrigel growth of cells treated with 10 μg/mL polyI:C consecutively every 3 days over 7 days. The viability of the cell colony formed in the matrigel was quantified and data presented as described in (A). (C) Scratch wound healing assay to measure cellular motility and wound closure rate of A549 and NCI-H292 cells treated with polyI:C at different time intervals (0, 12, 24 h). The wound closure rate and wound width restoration was analysed using ImageJ software. The wound width is presented as percent polyI:C-treated cells relative to NT cells. The representative images of the wounded areas were taken after polyI:C treatment at different time intervals. (D and E) Migration and invasion of A549 and NCI-H292 were determined by transwell migration /invasion assays. Cells were treated with 10 μg/mL polyI:C for 24 h and the migrated /invaded cells underneath the transwell insert were stained by Hoechst 33342, and counted under fluorescence microscopy. The invasion potency was determined using 2% matrigel pre-coated transwell invasion assay. The data are presented as percent of polyI:C-treated cells relative to NT. Images were taken at 40x magnification. Bar, 100 μM; *P<0.05, **P<0.01, ***P<0.001.