Generation of vasoactive peptide bradykinin from human umbilical vein endothelium-bound high molecular weight kininogen by plasma kallikrein.
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.
Nishikawa
K
K
Department of Medicine, SUNY-Stony Brook 11794-8161.
Shibayama
Y
Y
Kuna
P
P
Calcaterra
E
E
Kaplan
A P
AP
Reddigari
S R
SR
D000595:Amino Acid Sequence; D001920:Bradykinin / Q000378:metabolism*; D002478:Cells, Cultured; D004730:Endothelium, Vascular / Q000378:metabolism*; D006801:Humans; D007124:Immunoenzyme Techniques; D007610:Kallikreins / Q000378:metabolism*; D007700:Kinetics; D007704:Kininogens / Q000378:metabolism*; D007705:Kinins; D008969:Molecular Sequence Data; D011498:Protein Precursors / Q000378:metabolism*; D014471:Umbilical Veins
D007704:Kininogens; D007705:Kinins; D011498:Protein Precursors; C046022:prekininogens; D007610:Kallikreins; D001920:Bradykinin