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. 2017 Mar 2;8(20):32884–32904. doi: 10.18632/oncotarget.15862

Figure 4. TLR3 knockdown by siRNA decreased polyI:C-suppression of monolayer cell proliferation, survival, migration and invasion of A549 and NCI-H292.

Figure 4

(A) TLR3 mRNA level of A549 and NCI-H292 after knockdown with 50 nM TLR3-siRNA (siTLR3) for 24 h, compared to scrambled RNA control (siControl). After knockdown, the cells were treated with 10 μg/ml polyI:C for another 24 h. Data are presented as Tlr3 mRNA level relative to a housekeeping gene (Hprt) compared to NT control (not treated with polyI:C). (B) Monolayer cell proliferation assay (of total cell numbers) shows that TLR3 knockdown decreased the polyI:C-killing of A549 and NCI-H292. (C) TLR3 knockdown cells were treated as described above and caspase 3/7 activity was measured by caspase-glo 3/7 assay, showing decreased the polyI:C-mediated apoptosis. (D) TLR3 knockdown NCI-H292 cells were treated as above, and the cell migration and invasion were assayed, showing decreased the polyI:C-mediated suppression. The migrated and invaded cells which appeared underneath the transwell insert were stained with Hoechst 33342 and counted by fluorescent microscopy. For all of the above functional assays, the results are presented as percent of polyI:C-treated cells with siControl or siTLR3 relative to NT. *P<0.05, **P<0.01, ***P<0.001 indicate polyI:C treated cells vs. NT. n.s, non-significant; #P<0.05, ##P<0.01, ###P<0.001 indicate cells with TLR3 knockdown vs. scrambled control siRNA.