Hhat KD inhibits PANC1 proliferation and matrix invasion.

<p>A. PANC1 cells were labeled with CFSE at the start of the experiment and treated with Shh neutralizing antibody 5E1 on Day 1 (5E1 D1) to test whether PANC1 proliferation is Shh dependent. 5E1 treatment on Day 4 (5E1 D4) was used to mimic the kinetics of siRNA KD. Cells were allowed to grow for 8 days during which CFSE dilution gave a measure of cell division. 8 days after transfection with Hhat-#1 siRNA the cells were analyzed by flow cytometry. The Y-axis shows the CFSE mean fluorescence intensity (MFI) observed from 30,000 cells in each condition. The experiments were repeated three times in triplicate, and statistical significance was measured by using the two-tailed t test (***, <i>P</i><0.001). B, C. 72 h after transfection with Hhat-1 siRNA or control Mutated Hhat siRNA PANC1 cells were plated onto Matrigel invasion chambers for 24 h. Cells that had migrated from the upper to the lower side of the filter were photographed (B) and counted with a light microscope (40 fields/filter, C). Non-treated (NT) and 5E1-treated cells are shown for comparison in B. The experiments were repeated four times, and statistical significance was measured by using the two-tailed t test (***, <i>P</i><0.001).