Figure 2. KLF4 is increased in α-actin-positive cells in Chop^fl/flSM22α-CreKI⁺Apoe^−/− lesions and is a mechanism of decreased VSMC proliferation.

A, Representative Z series projection images of KLF4 staining (red) and α-actin staining (green) in Chop^fl/flSM22α-CreKI⁺Apoe^−/− lesions and quantification of KLF4⁺;α-actin⁺ cells or KLF4⁺;α-actin⁻ cells in Chop^fl/flApoe^−/− and Chop^fl/flSM22α-CreKI⁺Apoe^−/− lesions (n=8). The upper left image shows the DAPI+ cells in this projection. Scale bar, 100 µm. B, Klf4 mRNA relative to Actb in Chop^fl/flApoe^−/− and Chop^fl/flSM22α-CreKI⁺Apoe^−/− VSMCs treated with 2.5 µg/ml tunicamycin for 12 h (n=3). C, Klf4 mRNA relative to Actb (n=6, graph) and immunoblot of KLF4 and CHOP protein (n=3) in Chop^fl/flApoe^−/− and Chop^fl/flSM22α-CreKI⁺Apoe^−/− VSMCs cultured in 10% FBS medium for 24 hours after serum starvation. β-actin was included as loading control. D, Left panel: Proliferation curve of scrambled (Scr) or Klf4 siRNA-transfected Chop^fl/flApoe^−/− and Chop^fl/flSM22α-CreKI⁺Apoe^−/− VSMCs cultured in 10% FBS medium 0–4 days after starvation (n=3; at day 4, the Cre+ siKlf4 value is different from the Cre+ ScrRNA value at P=0.017). Right panel: KLF4 protein expression in Scr or Klf4 siRNA-transfected Chop^fl/flApoe^−/− and Chop^fl/flSM22α-CreKI⁺Apoe^−/− VSMCs.