Generation of vasoactive peptide bradykinin from human umbilical vein endothelium-bound high molecular weight kininogen by plasma kallikrein

Abstract

High molecular weight kininogen (HK) is a multifunctional plasma glycoprotein that occupies a critical position in pathways that link inflammation and coagulation. It is an inhibitor of sulfhydryl proteases and has procoagulant properties. It is also a source of the vasoactive peptide bradykinin (BK). It has been previously shown that HK binds to human umbilical vein endothelial cells (HUVEC) in culture. We have further characterized that interaction herein. Immunohistochemical experiments have indicated that when freshly obtained umbilical vein segments were treated with HK, washed, and probed with anti-HK antibodies, HK was localized on the endothelium. We next determined whether HUVEC-bound HK can be cleaved by plasma kallikrein to release BK. Cultured HUVEC were incubated with unlabeled HK for varying times, washed, and the kinetics of BK release by plasma kallikrein were assayed by radioimmunoassay. Results indicated that kallikrein released BK from HUVEC in proportion to the initial amount of bound HK. No release of BK occurred in the absence of kallikrein. Also, there was no BK release upon kallikrein treatment of the HUVEC not treated with exogenous HK. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis and autoradiography of HUVEC-bound 125I-HK indicated that addition of kallikrein resulted in cleavage of HK, thus corroborating the BK release experiments. Comparison of cleavage patterns has also indicated that cell-bound HK is slightly less susceptible to digestion by kallikrein than free HK. Therefore, our data suggest that human HK can bind to vascular endothelium in situ and that plasma kallikrein can recognize endothelial-bound HK as a substrate and liberate the vasoactive peptide BK.

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

“…Previous studies (19) had shown that when PK binds to bound HK on HUVEC, the formed kallikrein cleaves its receptor, HK, in a pattern consistent with that associated with liberated BK. Furthermore, Nishikawa et al (21) showed that kallikrein incubated with endothelial cellbound HK will liberate BK. The present investigations advance those investigations by showing that simultaneously with the generation of kallikrein activity on HUVEC, BK is liberated.…”

“…When PK binds to HK on endothelial cells, kallikrein is formed by a cell-associated, antipain-sensitive activator (16,19). Kallikrein formation has the potential to liberate BK from its cell receptor HK (18,19,21). BK, a potent vasoactive agent, has the potential to induce endothelial cells to release nitric oxide (NO) (22).…”

Platelet Glycoprotein Ib: A Zinc-Dependent Binding Protein for the Heavy Chain of High-Molecular-Weight Kininogen

“…HK was iodinated with Na'251 by iodobeads (Pierce) according to manufacturer's recommendations as modified by Nishikawa et al (13).…”

Regulation of leukocyte recruitment by polypeptides derived from high molecular weight kininogen

“…HK is composed of six domains and is present in plasma at a concentration of 0.67 M (13)(14)(15). Kallikrein can liberate the short-lived vasodilator peptide bradykinin from HK (16), thereby generating stoichiometric concentrations of two-chain, kinin-free HKa, which lacks most of its domain 4. Domain 5 in HKa is rich in His, Gly, and Lys, which enables HKa to bind to anionic surfaces, zinc ions, or heparin as well as to cell surface proteoglycans (17)(18)(19)(20).…”