Summary

We have quantitated the binding of high molecular weightkininogen (HK) to human microvascular endothelial cells oflung and dermal origin as well as to astrocytes and comparedthe results with those reported for human umbilical veinendothelial cells (HUVEC). We also reassessed parameters ofbinding to HUVEC employing cells in suspension as well as cellsattached to the culture plate and report similar numbers ofsites varying from 6.96x10⁵ to 7.71x10⁵ per cell. The presentstudy shows that HK binds with high specificity and affinity tomicrovascular endothelial cells (Kd = 1.86 to 4.5 nM) comparedto HUVEC (Kd = 10.35nM) but with lower affinity to astrocytes(Kd = 23.73 nM). Human cytokeratin 1, urokinase plasminogenactivator receptor and gC1qR were found to be HKbinding proteins present at the surface of microvascularendothelial cells and astrocytes analogous to that seen inHUVEC, as assessed by inhibition of binding with antibody toeach protein. Lung microvascular endothelial cells had approximatelyhalf the number of HK binding sites as HUVEC whiledermal micro vascular endothelial cells and astrocytes had only8-10% of the sites/cell. The affinity of binding to the microvascularendothelial cells was greater than HUVEC, the affinity ofbinding to astrocytes was considerably less, nevertheless bindingto each cell type involves gC1qR, cytokeratin 1 and u-PARto varying degrees.We also demonstrate, for the first time, thatfactor XII binds to all of these cell types in a saturable and Zn⁺²dependent manner. Given that factor XII accelerates the inter-actionsamong cell surfaces and proteins of the contact activa-tioncascade to generate bradykinin, binding of factor XII (andthe prekallikrein-HK complex) may serve as a mechanism bywhich these proteins are concentrated locally to facilitate theirinteractions.