Effect of PIN1 knockdown and calpain inhibition on COX-2 stability. KD and Control shRNA MAEC were treated with vehicle (DMSO) or 25 μM zVF for 1 h, then with LPS and IFN for 24 h. Cycloheximide (90 μg/ml) was added, and cell extracts were collected at the indicated times and western blotted for COX-2 and α-tubulin. A, Representative images of COX-2 and α-tubulin. Blots were processed in the same reagents for each protein, and exposed on one film for all samples. B, The average COX-2/α-tubulin signal intensity ratio ± SE of 4 independent cultures for each point, as a percent of the value at 0 h after cycloheximide treatment, is shown. The dashed line marks the 50% value. *, p < 0.05 for comparison between vehicle- and zVF-treated KD cells or Control cells at the indicated time. +, p < 0.05 for comparison between similarly treated KD and Control cells at the indicated time.