A B S T R A C T An asymptomatic woman (Ms. \Vil-liams) was found to have a severe abnormality in the surface-activated intrinsic coagulation, fibrinolvtic, and kinin-generating pathways. Assays for known coagulation factors were normal while Fletcher factor (prekallikrein) was 45%, insufficient to account for the observed markedly prolonged partial thromboplastin time. Plasminogen proactivator was present at 20% of normal levels and addition of highly purified plasminogen proactivator containing 10% plasminogen activator partially corrected the coagulation and fibrinolytic abnormalities but not the kinin-generating defect. This effect was due to its plasminogen activator content. In addition, Williams trait plasma failed to convert prekallikrein to kallikrein or release kiniin upon incubation with kaolin. Kininogen
Patients with hereditary angioedema lack C-1 inhibitor, a plasma alpha 2-glycoprotein that inhibits both the proteolytic action of C1, the activated first component of the complement system, and the activity of components of the contact phase of coagulation: kallikrein, factor XIa, and factor XIIa. Such patients have been shown to have low levels of C4 and C2, the natural substrates for C-1, but the levels were not correlated with the presence of symptoms. We studied three patients with angioedema for evidence of activation of the contact system and found that during a symptomatic period they had decreased levels of prekallikrein, a substrate for the activated forms of factor XII, and reductions in high-molecular-weight kininogen, a substrate for plasma kallikrein. These observations suggest that zymogens of the contact system are activated during attacks of hereditary angioedema and that some of the clinical manifestations may be mediated through products of this pathway, such as kinins.
A B S T R A C T Elastase is released from human neutrophils during the early events of blood coagulation. Human plasma kallikrein has been shown to stimulate neutrophil chemotaxis, aggregation, and oxygen consumption. Therefore, the ability of kallikrein to release neutrophil elastase was investigated. Neutrophils were isolated by dextran sedimentation, and elastase release was measured by both an enzyme-linked immunosorbent assay, and an enzymatic assay using t-butoxycarbonyl-Ala-Ala-Pro-Val-amino methyl coumarin as the substrate. Kallikrein, 0.1-1.0 U/ml, (0.045-0.45 ,M), was incubated with neutrophils that were preincubated with cytochalasin B (5 Mg/ml). The release of elastase was found to be proportional to the kallikrein concentration. Kallikrein released a maximum of 34% of the total elastase content, as measured by solubilizing the neutrophils in the nonionic detergent Triton X-100. A series of experiments was carried out to determine if kallikrein was a major enzyme involved in neutrophil elastase release during blood coagulation. When 10 million neutrophils were incubated in 1 ml of normal plasma in the presence of 30
High Mr kininogen increases the activation rate of prekallikrein by activated factor XII on a surface. The resulting serine protease, plasma kallikrein, Mr 88 000, is inhibited in plasma by C1 inhibitor, Mr 105 000. Since prekallikrein circulates in plasma with high Mr kininogen as a complex and a kallikrein-high Mr kininogen complex can be formed in purified systems, we studied whether the inhibition of kallikrein by C1 inhibitor was influenced by high Mr kininogen. With C1 inhibitor in excess, the inactivation of kallikrein followed pseudo-first-order kinetics. The second-order rate constant for the reaction was 1.7 X 10(4) M-1 s-1, and a kallikrein-C1 inhibitor complex, Mr 190 000 was identified on polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate. Kallikrein and C1 inhibitor formed an irreversible complex without measurable prior equilibrium. The rate of this reaction was decreased by 50% in the presence of high Mr kininogen (1 unit/mL or 0.73 muM). Kinetic analysis indicated that this protection was the result of the formation of a reversible complex between kallikrein and high Mr kininogen, which had a dissociation constant of 0.75 muM. However, low Mr kininogen did not protect kallikrein from inactivation by C1 inhibitor. High Mr kininogen also protected kallikrein from inactivation by diisopropyl fluorophosphate. These findings suggest that the kallikrein-high Mr kininogen complex was formed by noncovalent interactions between the light chains of both kallikrein and high Mr kininogen.
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