Fig. 6.

IGF-I increases fibronectin mRNA expression and fibronectin promoter activity at similar time points. A: PTCs were serum deprived for 24 h and then treated with IGF-I (250 ng/ml) for 12, 24, and 48 h, respectively. RNA was extracted using RNAzole bee method. RT-PCR was performed for fibronectin mRNA using ΔΔCt method. Fibronectin mRNA expression was quantified with SYBR green dye and RT²qPCR primers. Data represent the relative induction of fibronectin mRNA to GAPDH mRNA levels. Data are means and SE of three experiments. B: the reporter plasmid containing fibronectin promoter that drives the expression of the luciferase gene was transfected into the PTCs using GeneJuice. After 24 h of transfection, cells were incubated with serum-free media for 24 h followed by treatment with IGF-I (250 ng/ml) for indicated time points. Luciferase activity was determined and normalized by protein content. pCDNA3 was used as a control vector. Data are means and SE of three experiments. Data demonstrate that IGF-I significantly induces the fibronectin promoter activity at 12 and 24 h. Results are expressed as means ± SE. *P < 0.05 and **P < 0.01 compared with uninduced timed control.