β3-endonexin decreases hypoxic tube formation. (A) HMEC-1 cells were transfected with expression vectors coding for either β3-endonexin short (EN-S) or long (EN-L) isoforms or with control vector (Ctr). Cells were seeded on matrigel and exposed to hypoxia (1% oxygen, Hx) for 6 h or remained under normoxia (Ctr). (B) HMEC-1 cells were transfected with siRNA against β3-endonexin (siEN) or scrambled RNA (siCtr) and, in addition, transfected with an expression vector coding for IκBdn or with control vector or with siRNA against HIF-1α (siHIF-1α). Cells were seeded on matrigel and exposed to hypoxia as described earlier. (C) For quantification of tube formation assays, total tube lengths were evaluated. Normoxic controls were set equal to 1 (n=3, *p<0.05 vs. normoxic Ctr/siCtr; #p<0.05 vs. hypoxic Ctr/siCtr). IκBα and HIF-1α levels were determined in cells expressing IκBdn or siHIF-1α and the respective control cells using antibodies against IκBα and HIF-1α. Actin levels served as loading control. Representative figures are shown.