Fig. 2.

Suppression of KLF4 transcriptional activity and nuclear localization by GSNO. (A-C) ECs were infected with Ad-KLF4 in the presence or absence of tetracycline (Tc, 0.1 μg/mL) for 24 h and then exposed to GSNO. The mRNA levels of KLF4 target genes (eNOS, ASS1 and thrombomodulin) were analyzed by qRT-PCR. *p < 0.05, by one-way ANOVA. n = 4–6. (D) After pretreatment with or without MG132 (1 μmol/L, 1 h), HUVECs were exposed to GSNO for indicated times. Nuclear proteins were extracted and analyzed by western blotting. *p < 0.05 vs. 0 h, by one-way ANOVA. n = 3. (E) Immunoblots of nuclear and cytoplasmic fractions from HUVECs after treatment with GSNO (0.5 mmol/L) for indicated times. *p < 0.05 vs. 0 h, by one-way ANOVA. n = 3. (F) Immunofluorescence of HUVECs stimulated with GSNO (0.5 mmol/L, 24 h) depicting the subcellular distribution of KLF4. Images displayed KLF4 in green and DAPI in blue. Rabbit IgG was used as negative control. n = 3. (For interpretation of the references to color in this figure legend, the reader is referred to the web version of this article).