Figure 6

TGF-β1 up-regulates mRNA and protein level of TIMP-3 in h PSCs and TGF-β1 induces TIMP-3 expression via Akt pathway in h PSCs. (A) h PSCs were incubated with 10 ng/ml of TGF-β1 in serum-free DMEM for 1, 2, 4, and 6 hours following starvation for 24 hours. TIMP-3 mRNA was significantly up-regulated after 1, 2, 4, and 6 hours and had the maximal effect in the time of 4th hour (n = 3). Lane 1 as basal control and lane 2 to lane 5 were compared with basal control. (B) h PSCs were incubated with 10 ng/ml of TGF-β1 in serum-free DMEM for 6, 12, and 24 hours. TIMP-3 protein was significantly induced after 6, 12, and 24 hours (n = 3). Lane 1 as basal control and lane 2 to lane 4 were compared with basal control. (C) h PSCs were incubated in starvation medium for 24 hours and then treated with 10 ng/ml of TGF-β1 in serum-free DMEM for 15, 30, 60, 120, and 240 minutes (n = 3). Lane 1 as basal control and lane 2 to lane 6 were compared with basal control. (D) h PSCs were incubated in starvation medium for 24 hours and then treated with 10 μM of SH-6 for 30 minutes followed by incubated with 10 ng/ml of TGF-β1 in serum-free DMEM for 4 hours (n = 3). Lane 1 as basal control and lane 2 as positive control, lane 3 was compared with positive control. (E) h PSCs were treated with 10 μM of SH-6 for 30 minutes followed by incubated with 10 ng/ml of TGF-β1 in serum-free DMEM for 24 hours (n = 3). Lane 1 as basal control and lane 2 as positive control, lane 3 was compared with positive control.