Assembly and activation of HK-PK complex on endothelial cells results in bradykinin liberation and NO formation

Prekallikrein (PK) activation on human umbilical endothelial cells (HUVEC) presumably leads to bradykinin liberation. On HUVEC, PK activation requires the presence of cell-bound high-molecular-weight kininogen (HK) and Zn2+. We examined the Zn2+ requirement for HK binding to and the consequences of PK activation on endothelial cells. Optimal HK binding (14 pmol/10(6) HUVEC ) is seen with no added Zn2+ in HEPES-Tyrode buffer containing gelatin vers us 16-32 muM added Zn2+ in the same buffer containing bovine serum albumin. The affinity and number of HK binding sites on HUVEC are a dissociation co nstant of 9.6 +/- 1.8 nM and a maximal binding of 1.08 +/- 0.26 x 10(7) sit es/cell (means +/- SD). PK is activated to kallikrein by an antipain-sensit ive mechanism in the presence of HK and Zn2+ on HUVEC, human microvascular endothelial cells, umbilical artery smooth muscle cells, and bovine pulmona ry artery endothelial cells. Simultaneous with kallikrein formation, bradyk inin (5.0 or 10.3 pmol/10(6) HUVEC in the absence or presence of lisinopril , respectively) is liberated from cell-bound HK. Liberated bradykinin stimu lates the endothelial cell bradykinin B2 receptor to form nitric oxide. Ass embly and activation of PK on endothelial cells modulates their physiologic al activities.