Figure 1.

KLF4 was up-regulated by rapamycin in vitro and in vivo. (A) VSMCs were treated with rapamycin (100 ng·mL⁻¹) or control (DMSO) for 12 and 24 h. KLF4 mRNA level was assessed with qRT-PCR and expressed as fold induction compared with the controls after normalizing to GAPDH. (B) After treatment with a range of concentrations of rapamycin (2–200 ng·mL⁻¹) for 24 h, nuclear extracts were immunoblotted with antibodies against KLF4 and histone. Data are representative of five independent experiments. (C) VSMCs were transfected with plasmids expressing mTOR, mTOR-KD or control plasmid. Total RNA was extracted 36 h later and subjected to qRT-PCR for KLF4. (D) KLF4 mRNA levels were detected in VSMCs transfected with TSC2, TSC2-1080 or control plasmid. Total RNA was extracted 36 h later and subjected to qRT-PCR for KLF4. (E) Rat carotid arteries were balloon-injured. Total RNA was extracted 72 h later and subjected to qRT-PCR for KLF4 (n= 6 for each group). Balloon-injured arteries were treated perivascularly with pluronic gel containing rapamycin (100 µg per artery) or DMSO, for 72 h before RNA extraction (n= 3 for each group). *P < 0.05, **P < 0.01, ***P < 0.001, significantly different from control.