Figure 9. Ubqln1 overexpression promotes proteasomal degradation of oxidized proteins in cultured NRVMs.

Twenty-four hours after plating, NRVMs were infected with Ad-Ubqln1 or Ad–β-Gal. Further experiments were initiated at 48 hours after the adenoviral infection. (A and B) Representative images of Western blot analyses for DNP-derivatized protein carbonyls. The Ubqln1 overexpressing and control NRVMs were treated with H₂O₂ at the indicated doses. The cells were harvested at either 15 minutes (A) or 3 hours (B) after the H₂O₂ treatment for DNPH derivatization to measure the level of protein carbonyls. D/G, DNP/GAPDH. Representatives of 3 biological repeats are shown. (C and D) Proteasome inhibition attenuates the Ubqln1 overexpression-induced reduction of oxidized proteins. The cultured NRVMs were treated with proteasome inhibitor bortezomib (BZM, +) or vehicle control (saline, –) at 10 minutes before H₂O₂ treatment was initiated. Three hours later, cells were harvested for DNPH derivatization. Representative images (C) and pooled densitometry data from 3 biological repeats (D) are shown. *P < 0.05 vs. the H₂O₂ group; 1-way ANOVA followed by Tukey’s test.