Fig. 6.

Proliferative effects of Shh induction on SMCs is inhibited by AKTI IV, cyclopamine, and 3-methyladenine (3-MA). A: Shh cells with or without Dox induction were treated with cyclopamine (Cyclo) for 24 h. Dimethyl sulfoxide (DMSO, control) was the vehicle control for cyclopamine. Cell numbers were determined using the water-soluble tetrazolium salt (Wst-1) assay (*P < 0.05 vs. control and #P < 0.05 vs. Dox treated, n = 4). B: Shh cells with or without Dox induction were incubated with AKTI IV for 24 h, followed by determination of cell numbers (*P < 0.05 vs. control and #P < 0.05 vs. Dox treated, n = 4). C and D: Western blots showing the effects of Shh induction on the expression of proliferating cell nuclear antigen (PCNA, C) and p27 (D) (top: representative Western blots). The expression of PCNA (C) or p27 (D) was normalized to α-tubulin (*P < 0.05 vs. without Dox treated, n = 5). E: Shh cells with or without Dox induction were treated with 3-MA or rapamycin for 24 h, followed by determination of cell numbers (*P < 0.05 vs. control and #P < 0.05 vs. Dox treated, n = 4). F: TR cells were treated with N-Shh, bafilomycin A1, or N-Shh and bafilomycin A1 for 24 h, followed by determination of cell numbers (*P < 0.05 vs. control and #P < 0.05 vs. N-Shh treated, n = 4). G: TR cells were transfected with NC siRNA or ATG7 siRNA in the presence of N-Shh for 24 h, followed by determination of cell numbers (*P < 0.05 vs. NC siRNA control and #P < 0.05 vs. N-Shh + NC siRNA, n = 4).