Measurement of desarginine fibrinopeptide B in human blood.

Abstract

A B S T R A C T Thrombin converts fibrinogen to fibrin in two steps. First fibrinopeptide A and fibrin I are formed and then fibrinopeptide B (B,3 1-14) and fibrin II. Since it is postulated that fibrin II is important in the genesis of thrombosis, it is of interest to measure fibrinopeptide B in peripheral blood samples. Previous difficulties in interpreting fibrinopeptide B immunoreactivity in plasma resulted from crossreaction offibrinogen and of plasmin digest peptides B,3 1-42 and Bf3 1-21 and from rapid loss of fibrinopeptide B immunoreactivity resulting from cleavage of arginine 14 by blood carboxypeptidase B. We have obviated these difficulties by removing fibrinogen from plasma by precipitation with ethanol and peptides Bf3 1-21 and BP 1-42 by adsorption on bentonite. Fibrinopeptide B is then converted to desarginine fibrinopeptide B, which is measured in a new specific assay. Studies of the kinetics of fibrinopeptide cleavage showed that when whole blood was allowed to clot in vitro, fibrinopeptide A was cleaved more rapidly than fibrinopeptide B. In 18 patients on an acute care medical ward, desarginine fibrinopeptide B levels were lower than fibrinopeptide A levels and did not correlate with the levels of fibrinopeptide A or B,B 1-42. Desarginine fibrinopeptide B levels were <1 pmol/ml in all but two patients. In six patients receiving intraamniotic infusions of hypertonic saline to induce abortion, desarginine fibrinopeptide B levels increased 10-fold from the preinfusion mean level of 0.4 pmol/ml and then decreased. The pattern of changes resembled that ofthe fibrinopeptide A levels rather than of the B,8 1-42 levels. On the basis of these data it is suggested that plasma desarginine fibrinopeptide B levels reflect fibrin II formation in vivo.

Human tissue-type plasminogen activator releases fibrinopeptides A and B from fibrinogen.

“…This value is within the reported normal (24) and is consistent with previous reports ofrapid normalization ofFPA levels after heparin therapy for venous thromboembolic disease (33,34). In contrast, the mean plasma value for desarginine FPB before rt-PA infusion was 5.6 nM, whereas normal levels are < 0.6 nM (29).…”

“…The neutrophil proteinase, elastase, also may contribute since this enzyme is released from neutrophils during the blood coagulation process (43) and has the potential to attack the NW2-terminal region of the BB-chain of fibrinogen resulting in increased desarginine FPB immunoreactivity (44) without an increase in FPA (23). The extent of elevation and the persistent increase in fibrinopeptide levels after rt-PA administration is difficult to explain given the short half-lives of t-PA (45,46) and the fibrinopeptides (24,29), and the relatively slow rates of rt-PAmediated fibrinopeptide release in vitro. The binding of functionally active t-PA to endothelial cells (47) however, may serve to localize and protect the enzyme from its inhibitors thereby prolonging its duration of action and increasing its enzymatic potential.…”

Direct Tandem Mass Spectrometry Reveals Limitations in Protein Profiling Experiments for Plasma Biomarker Discovery

“…38 Both of these exopeptidases degrade biologically active peptides in vivo. [39][40][41][42][43][44][45] Heightened activity of AMPN has been reported for cancer patients, 46 and CBPN activity is increased after myocardial infarction. 47 The proteins that were identified here also indicate some specificity in the cleavage patterns, i.e., the abundances of the plasma peptides observed here do not mirror the abundant proteins (see Table 2).…”

Urokinase has direct catalytic activity against fibrinogen and renders it less clottable by thrombin.

“…LMW urokinase (26 nM to 0.7 AM) was incubated with heparinized plasma for 60 min at 370C. The reaction was terminated and unreacted fibrinogen was precipitated with ethanol, and the ethanol supernatants were then assayed for lytic activity of urokinase against fibrinogen was assessed by measuring the levels of desarginine FPB, the stable carboxypeptidase-derived metabolite of FPB (28,29).…”