Figure 3

ALK+ ALCL cell lines with GzB knock-down show significant reduction in GzB activity. A. Cleavage of the synthetic GzB substrate, Ac-IEPD-pNA, was examined using lysates from the indicated ALK+ ALCL cell lines expressing either control or GzB shRNA. The results represent the average and standard deviation of 3 independent experiments. p values comparing cells expressing GzB shRNA to cells expressing control shRNA were obtained by performing paired, one-tailed t-tests. *p < 0.05, **p < 0.01, ***p < 0.005. B. Lysates from Karpas 299 cells (above) or SUP-M2 cells (below) expressing the indicated shRNAs were incubated with purified vitronectin for the indicated times and vitronectin cleavage was examined by western blotting. Arrows indicate the major observed vitronectin cleavage product. Cleavage of vitronectin was also examined when incubated with lysates from the GzB-negative, Hodgkin lymphoma cell line, KM-H2, which served as a negative control. Anti-β-actin blots were performed to demonstrate that equivalent amounts of lysate were added to the reactions. The percentage of cleaved vitronectin (% cleaved vitronectin) was determined by densitometry, and represents the percentage of cleaved vitronectin in each lane relative to cleaved vitronectin in the lane where vitronectin was incubated with control shRNA lysate for 6 h. Molecular mass standards are indicated to the left of the western blots.