Figure 1. sPLA₂-V and -X suppress adenovirus-mediated gene delivery into human lung-derived cells.

(A) Human bronchial epithelial BEAS-2B cells, which had been stably transfected with various PLA₂s and COX-2 were subjected to infection with Ad-mPGES-1 or Ad-iPLA₂β at MOI (multiplicity of infection)=10. After 24 h the cells were harvested and taken for Northern-blotting with probes against mPGES-1, iPLA₂β and G3PDH. (B) NHPF were infected with adenoviruses for various PLA₂s at MOI=10 for 24 h and were then infected with Ad-mPGES-1 or Ad-iPLA₂β at MOI=2 for an additional 24 h. The cells were harvested and taken for Northern-blotting with probes against mPGES-1, iPLA₂β and G3PDH. (C) BEAS-2B cells stably expressing different amounts of sPLA₂-V (left panel) or -X (right panel) or mock-transfected cells (−) were infected with Ad-mPGES-1 at MOI=10 for 24 h. Then the cells were harvested and taken for Northern-blotting with probes for sPLA₂-V or -X, mPGES-1 and G3PDH. sPLA₂ activity released into the supernatants from individual clones is shown in the bottom panel. (D) Control BEAS-2B cells and those stably transfected with sPLA₂-V or -X were infected for the indicated periods with Ad-mPGES-1 at MOI=10. Then the cells were subjected to Northern-blotting with probes against mPGES-1 and G3PDH. The bottom panel shows the ratio of signal intensities between mPGES-1 and G3PDH. (E) AA release from BEAS-2B cells transfected with various PLA₂s. Cells prelabelled with [³H]AA for 24 h were stimulated for 4 h with 100 units/ml TNFα to assess [³H]AA release. (F) BEAS-2B cells stably expressing various PLA₂s with or without cotransfection of COX-2 were incubated with TNFα for 4 h to assess generation of PGE₂. (A) to (D) representative results of 3–5 experiments and values are mean±S.E.M. for three independent experiments in (E) and (F).