Figure 4.

Attenuation of IPO5 levels with siRNA results in reduced levels of IL-33 when overexpressed in NHLFs. A, NHLFs treated with IPO5-targeting siRNA then electroporated with the FLIL33HA-encoding plasmid show a reduction in FLIL33HA protein as detected by anti-HA antibody, with two separate experiments shown (left panel). By contrast, MIL33HA levels were not affected by IPO5 attenuation (right panel). This experiment was repeated in two different primary NHLF cultures, with similar results. B, ELISA for IL-33 was performed with NHLF lysates following siRNA-mediated depletion of IPO5 and electroporation with the FLIL33HA-encoding (left panel) or vehicle control (NULL, right panel). Mean ± S.D. values pooled from three separate experiments with each condition tested in duplicates are shown. Significantly decreased (p < 0.05) IL-33 levels were observed with IPO5 attenuation in FLIL33HA-overexpressing cells as indicated with asterisks; there was a tendency to lower intracellular endogenous IL-33 in NULL-transfected cells, but it did not reach statistical significance. Treatment with the 20S proteasome inhibitor, bortezomib, was used as indicated and resulted in partial restoration of IL-33 levels.