Carbon Monoxide-Releasing Molecule-3 Suppresses the Malignant Biological Behavior of Tongue Squamous Cell Carcinoma via Regulating Keap1/Nrf2/HO-1 Signaling Pathway

Figure 3

The regulatory effect of CORM-3 on the Keap1/Nrf2/HO-1 signaling pathway in the CAL27 and SCC4 cells. (a, b) CAL27 and SCC4 cells were treated with 0, 100, 200, and 400 μM CORM-3 for 24 h, and western blot was used to detect the protein expression of HO-1. (c, d) CORM-3(400 μM) stimulated cells at 0, 0.25, 0.5, 1, 3, 6, and 12 h, respectively. Western blot was used to detect the effect of CORM-3 on HO-1 protein expression. (e) The efficacy of HO-1specific shRNA was measured by RT-qPCR. We finally chose sh-HO-1-3# as the interference sequence. (f) The efficacy of shHO-1-3# was measured by western blot. (g, h) The effect of shHO-1 on the inhibitory effect of CORM-3 on the proliferation of CAL27 and SCC4 cells. Cell proliferation was assessed by a CCK-8 kit. (i, j) The effect of shHO-1 on the inhibitory effect of CORM-3 on the migration of CAL27 and SCC4 cells. Quantification of relative migrating distances of cells with indicated treatment. (k, l) The effect of shHO-1 on the inhibitory effect of CORM-3 on the invasion of CAL27 and SCC4 cells. Quantification of the invaded cells with indicated treatment. (m, n) CAL27 and SCC4 cells were treated with CORM-3 for 0, 0.25, 0.5, 1, 3, 6, and 12 h, and the protein expression of Keap1 and Nrf2 was assessed by western blotting. Relative protein level was normalized to the corresponding GAPDH protein expression. , , and vs. the control group, respectively. ^ and ^^ vs. comparison between two groups.

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