H₂O₂ inhibits proteasome activity and proteasome inhibitor lactacystin induces p62 up-regulation.

(A) ARPE-19 cells were treated with 200 or 400μM H₂O₂ and then chymotrypsin-like proteasome activity was measured at 24h post-exposure using a fluorogenic substrate. Control (Ctrl) = No H₂O₂ (B) Chymotrypsin-like proteasome activity was detected in RPE cells treated with the proteasome specific inhibitor lactacystin (4nM and 25μM, 12h). Ctrl = DMSO control (C) qRT-PCR was used to measure the p62 mRNA level following lactacystin treatment (25μM, 12h). Ctrl = DMSO control (D) p62 protein level was examined by western blot after proteasome specific inhibitor lactacystin (25μM, 12h) exposure. Ctrl = DMSO control. Data represent the mean + S.E.M. for three samples per group. Differences in means were considered statistically significant when p<0.05.