Loss of BMPER decreases eNOS levels and increases the expression of endothelial adhesion molecules ICAM-1 and VCAM-1. (A) SiRNA-based BMPER knockdown reduces eNOS protein levels. HUVECs were transfected with either of 2 specific BMPER siRNAs. After 48 hours, cell lysates were used for Western blotting with an anti–human eNOS (A) and an anti-BMPER antibody (supplemental Figure 2). β-tubulin was used as loading control. BMPER knockdown enhanced VCAM-1 RNA (B) shown real-time PCR, VCAM-1 protein (C), and ICAM-1 (D) expression levels as demonstrated by immunoblotting (*P < .05 versus sinegative). (E) ICAM-1 and VCAM-1 expression is enhanced in HUVECs with TNFα treatment after BMPER knockdown. HUVECs were transfected with scrambled or BMPER specific siRNA and after 36 hours cells were exposed to TNFα for additional 6 hours. Cells were lysed and lysates were used for Western blotting.