Bradykinin regulates the expression of kininogen binding sites on endothelial cells.
The vasoactive compound bradykinin (BK) is liberated by proteolytic cleavage from high molecular weight kininogen (HK) and low molecular weight kininogen (LK). Expression of kininogens on cell surface receptors may affect the delivery of BK at sites of inflammation. Therefore, we investigated whether BK itself alters the expression of binding sites for its parent molecules, HK and LK, on the surface of cultured human umbilical vein endothelial cells (HUVEC). 125I-LK and 125I-HK each bind to a single class of sites on HUVEC in reactions that are saturable, reversible, and zinc-dependent (Bmax = 9.7 +/- 0.2 x 10(5) sites/cell; kd = 43.3 +/- 8 nmol/L; n = 5 and Bmax = 10.3 +/- 0.4 x 10(5) sites/cell; kd = 40.3 +/- 0.9 nmol/L; n = 3 for LK and HK, respectively). HK and LK compete for the same binding site (Ki = 19.4 +/- 5 nmol/L HK v 125I-LK; Ki = 24.5 +/- 4 nmol/L LK v 125I-HK, n = 3). Moreover, 50-fold molar excess light chain of HK inhibits 125I-LK binding 51% and 50-fold molar excess LK and the heavy chain of HK inhibit 125I-light chain of HK binding 92% and 76%, respectively. Preincubation of HUVEC with BK produces a transient, concentration-dependent increase in the binding of HK and LK, reaching a maximum 3 to 4 hours after addition of BK (46% increase over control for HK; 57% increase over control for LK; P < .005 for each ligand). Des-Arg9-bradykinin, a B1 receptor agonist, increases kininogen binding to the same extent as BK; the upregulation of kininogen binding sites by BK is partially blocked by a B1 but not by a B2 receptor antagonist. The protein kinase C inhibitors (PKC), sphingosine and H7, completely block the induction of HK receptors by BK. Phorbol 12-myristate 13-acetate (PMA), which also activates PKC, stimulates the binding of HK and the purified light chain of HK to HUVEC as well. However, unlike HK and its light chain, binding of LK and the heavy chain of HK are increased by PMA only in the presence of added calcium ion. These studies show that BK upregulates a common binding site for HK, LK, and each chain of HK on HUVEC. Induction of kininogen receptors on endothelial cells by BK may modulate the generation of this vasoactive compound at sites of vascular injury.
Department of Medicine (Hematology/Oncology), Hospital of the University of Pennsylvania, Philadelphia 19104.
J M
JM
Zini
A H
AH
Schmaier
D B
DB
Cines
D001920:Bradykinin / Q000494:pharmacology*; D002118:Calcium / Q000494:pharmacology; D004224:Diterpenes; D004730:Endothelium, Vascular / Q000378:metabolism*; D006801:Humans; D066298:In Vitro Techniques; D007704:Kininogens / Q000378:metabolism*; D011493:Protein Kinase C / Q000037:antagonists & inhibitors; D018002:Receptors, Bradykinin; D017981:Receptors, Neurotransmitter / Q000378:metabolism* / Q000502:physiology*; D018000:Receptors, Peptide; D013729:Terpenes / Q000494:pharmacology; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D013997:Time Factors; D015854:Up-Regulation
D004224:Diterpenes; D007704:Kininogens; D018002:Receptors, Bradykinin; D017981:Receptors, Neurotransmitter; D018000:Receptors, Peptide; D013729:Terpenes; C044865:kininogen receptor; C011309:mezerein; D011493:Protein Kinase C; D013755:Tetradecanoylphorbol Acetate; D001920:Bradykinin; D002118:Calcium