A, Cell lysates from KMCH cells treated with cyclopamine (5 µM) for the indicated times were immunoblotted for cIAP-1, cIAP-2 and XIAP. Actin was used as a loading control. B, KMCH cells were transfected with a tetracycline-inducible shRNA expression vector targeting Smoothened (shSMO) or a scrambled shRNA vector; shRNA expression was induced for 48 hours by treatment with tetracycline (1 mg/mL) followed by immunoblotting for cIAP-1, cIAP-2 and XIAP using whole cell lysates. C, Cells were treated with vehicle, cyclopamine (5 µM) for 24 hours, or transfected with shSMO as in panel B, then analyzed by real-time RT-PCR for cIAP-1, cIAP-2 and XIAP using total RNA. D, Total RNA from tissue samples of either cholangiocarcinoma (CCA) or adjacent benign liver was used to assess XIAP, cIAP-1, and cIAP-2 mRNA expression, normalized to 18S and expressed as fold change from benign. Mean ± SEM, *p<0.05; **p<0.01; ***p<0.001 compared to vehicle.