The expression of high molecular weight kininogen on human umbilical vein endothelial cells

• Published in: The Journal of biological chemistry 1988-11, Vol.263 (31), p.16327-16333
• Main Authors: Schmaier, A H, Kuo, A, Lundberg, D, Murray, S, Cines, D B
• Format: Article
• Language: English
• Subjects: Biological and medical sciences; Blotting, Northern; Cell physiology; Cells, Cultured; Endothelium, Vascular - metabolism; Fundamental and applied biological sciences. Psychology; Hormonal regulation; Humans; Kinetics; kininogen; Kininogens - biosynthesis; Kininogens - genetics; Kininogens - isolation & purification; Molecular and cellular biology; Molecular Weight; Protein Binding; RNA - genetics; RNA - isolation & purification; umbilical cord; Umbilical Veins
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1016/S0021-9258(18)37596-3

High molecular weight kininogen (HMWK) functions as a cofactor for activation of plasma serine zymogens and as an inhibitor of tissue cysteine proteases. Cell surfaces to which HMWK binds may provide sites for regulation of these systems. Localization of these HMWK-dependent processes at sites of vascular injury may depend on its binding to specific receptors on endothelial cells. In culture, passaged human umbilical vein endothelial cells (HUVEC) bind anti-HMWK antibody to the cell surface and contain 171 +/- 75 ng of HMWK/10(8) cells. [35S]Methionine-labeled HUVEC in culture synthesize a 120-kDa protein immunoisolated using an anti-kininogen antibody, and a 3500-nucleotide message for human HMWK was detected by Northern blot in RNA extracted from HUVEC. HUVEC also express unoccupied binding sites for HMWK on their surface. 125I-HMWK specifically binds to HUVEC in a reaction requiring Zn2+. 125I-HMWK binding to HUVEC is saturable at 4 degrees C but not at 23 degrees C. 125I-HMWK binds to HUVEC with equal affinity as unlabeled HMWK. Kallikrein, factor XII, fibrinogen, fibronectin, and thrombin do not inhibit 125I-HMWK binding to HUVEC. 125I-HMWK-HUVEC binding remains fully reversible at 60 min following the addition of a 50-fold molar excess HMWK. HUVEC express 9.3 +/- 2.0 X 10(5) (mean +/- S.E.) HMWK binding sites/cell (Kd = 52 +/- 13 nM). Both added and cell-bound 125I-HMWK migrate at 120 kDa on sodium dodecyl sulfate gel electrophoresis, suggesting that the protein remains uncleaved upon binding to the HUVEC surface. These studies indicate that HUVEC synthesize HMWK and the HUVEC surface has a site for its expression. By synthesizing and localizing HMWK to the cell surface, endothelial cells may contribute to the activation of plasma's contact serine zymogens and regulation of tissue cysteine proteases.