Figure 2. Loss of c-Kit function in primary smooth muscle cells (SMC) is associated with increased NF-κB activity.

(A) NF-κB transcriptional activity in c-Kit deficient (Kit^W∕W−v), control (Kit^+∕+), and c-Kit rescued SMC (Kit^R) after 24-hour treatment with POVPC (50 µg/ml), as determined by dual-luciferase reporter assay. Transcriptional activity is represented as the mean ± standard deviation (SD) of the Firefly/Renilla luciferase ratio normalized with respect to the control group (Kit^+∕+) (n = 3 independent experiments). (B) Phosphorylated (pS536) protein levels of the NF-κB p65 subunit in POVPC-treated c-Kit deficient, control, and c-Kit rescued SMC as determined by ELISA. Values are expressed as the mean ± SD of the p-p65/total p65 ratio normalized with respect to the control group (Kit^+∕+) (n = 3 independent experiments). (C) Protein expression of the NF-κB related pro-inflammatory mediators MMP-2 and MCP-1 in POVPC-treated c-Kit deficient, control, and c-Kit rescued SMC as determined by Western blot. Molecular weight markers are shown on the right side of the gel. Protein expression is expressed as the mean ± SD of the MMP-2∕β-actin and MCP-1∕β-actin signal ratios normalized with respect to the control group (Kit^+∕+) (n = 3 per cell type). ^∗p < 0.05 and ^∗∗p < 0.01 using a one-way ANOVA followed by a Newman-Keuls test.