scholarly journals Normal titer of functional and immunoreactive protein-C inhibitor in plasma of patients with congenital combined deficiency of factor V and factor VIII

Blood ◽  
1983 ◽  
Vol 62 (6) ◽  
pp. 1266-1270 ◽  
Author(s):  
K Suzuki ◽  
J Nishioka ◽  
S Hashimoto ◽  
T Kamiya ◽  
H Saito
Keyword(s):  
Factor Viii ◽  
Protein C ◽  
Activated Protein C ◽  
Factor V ◽  
Amidolytic Activity ◽  
Normal Plasma ◽  
Coagulant Activity ◽  
Protein C Inhibitor ◽  
Combined Deficiency ◽  
Good Agreement

Protein-C inhibitor (PCI) is a newly described plasma inhibitor directed against a vitamin-K-dependent serine protease, activated protein-C, which is involved in the inactivation of factor V and factor VIII. Marlar and Griffin have reported that PCI activity is absent in the plasma of patients with congenital combined factor V/VIII deficiency. We have measured the levels of PCI in the plasma of seven unrelated patients with this disorder using both functional and immunologic methods. The rate at which the amidolytic activity of activated protein-C was neutralized in the patients' plasma was essentially identical to that observed in normal plasma. The titer of PCI antigen, as measured by an electroimmunoassay using a monospecific anti-PCI serum, was 5.3 +/- 1.6 micrograms/ml in the patients' plasma and was not significantly different from that of normal plasma (5.3 +/- 2.7 micrograms/ml, n = 30). The levels of factor-V-related antigen, factor V coagulant antigen, and factor VIII coagulant antigen were low in all patient plasma and were in good agreement with their respective coagulant activity. Our results do not appear to support the hypothesis that combined factor V/VIII defect is due to a lack of PCI.

scholarly journals Normal titer of functional and immunoreactive protein-C inhibitor in plasma of patients with congenital combined deficiency of factor V and factor VIII

Blood ◽  
1983 ◽  
Vol 62 (6) ◽  
pp. 1266-1270 ◽  
Author(s):  
K Suzuki ◽  
J Nishioka ◽  
S Hashimoto ◽  
T Kamiya ◽  
H Saito
Keyword(s):  
Factor Viii ◽  
Protein C ◽  
Activated Protein C ◽  
Factor V ◽  
Amidolytic Activity ◽  
Normal Plasma ◽  
Coagulant Activity ◽  
Protein C Inhibitor ◽  
Combined Deficiency ◽  
Good Agreement

Abstract Protein-C inhibitor (PCI) is a newly described plasma inhibitor directed against a vitamin-K-dependent serine protease, activated protein-C, which is involved in the inactivation of factor V and factor VIII. Marlar and Griffin have reported that PCI activity is absent in the plasma of patients with congenital combined factor V/VIII deficiency. We have measured the levels of PCI in the plasma of seven unrelated patients with this disorder using both functional and immunologic methods. The rate at which the amidolytic activity of activated protein-C was neutralized in the patients' plasma was essentially identical to that observed in normal plasma. The titer of PCI antigen, as measured by an electroimmunoassay using a monospecific anti-PCI serum, was 5.3 +/- 1.6 micrograms/ml in the patients' plasma and was not significantly different from that of normal plasma (5.3 +/- 2.7 micrograms/ml, n = 30). The levels of factor-V-related antigen, factor V coagulant antigen, and factor VIII coagulant antigen were low in all patient plasma and were in good agreement with their respective coagulant activity. Our results do not appear to support the hypothesis that combined factor V/VIII defect is due to a lack of PCI.

Interaction of Plasma Kallikrein with Protein C Inhibitor in Purified Mixtures and in Plasma

1991 ◽  
Vol 65 (01) ◽  
pp. 046-051 ◽  
Author(s):  
Francisco España ◽  
Amparo Estelles ◽  
John H Griffin ◽  
Justo Aznar
Keyword(s):  
Complex Formation ◽  
Protein C ◽  
Room Temperature ◽  
Activated Protein C ◽  
Physiological Role ◽  
Plasma Kallikrein ◽  
Amidolytic Activity ◽  
Normal Plasma ◽  
Protein C Inhibitor ◽  
Subsequent Addition

SummaryThe interaction between plasma kallikrein (KK) and protein C inhibitor (PCI) and the influence of KK on the complex formation between activated protein C (APC) and PCI was studied in purified systems as well as in plasma in order to assess the significance of these reactions in the plasma milieu. PCI complexed to KK (KK: PCI) or to APC (APC: PCI) was measured by sandwich ELISA’s using antibodies directed against each protein in the complexes. The formation of KK: PCI complexes assayed by this method paralleled the inhibition of KK amidolytic activity by PCI in purified system. Incubation of normal plasma (NHP) at 4 °C, which can induce prekallikrein activation due to cold activation, resulted in PCI inactivation and appearance of KK: PCI complexes. PCI activity fell to 35% of the NHp and 1.2 μ/ml of KK: PCI complex was formed. However, incubation of NHP at room temperature or of prekallikrein deficient plasma at 4 °C did not result in significant decrease of PCI activity. Thus the PCI inactivation was associated with prekallikrein activation and complexation to PCI following cold activation. Incubation of exogenous purified KK with NHP resulted in PCI inactivation and complexation with KK in a temperaturedependent manner. Addition of 2.8 μ/ml KK to plasma at 4 °C resulted in the inactivation of 55% of plasma PCI and the formation of 0.9 μ/ml KK: PCI which represents 21% of the KK added, whereas at 37 °C PCI was inactivated to 30% and only 0.30 μg/ml KK: PCI complexes were measured. These results indicate that PCI is a major KK inhibitor at 4 °C. At 37 °C, PCI accounted for aborfi 7% of the inhibition of the KK added. In separate experiments, following addition of 2.5 μg/ml APC to NHR more than 1 μg/ml of APC: PCI complex was formed in 3 h. When NHP was prior incubated with KK, PCI activity decreased to 10% of that of the normal plasma. Subsequent addition of APC to the plasma treated with KK resulted in formation of only 35 μg/ml of APC: PCI complex compared to 1,350 μg/ml when plasma was not previously incubated with KK. These results indicate that PCI could play a physiological role in the inhibition of plasma KK, and that, in turn, plasma KK can either complex to or inactivate plasma PCI. Thus, KK could modulate the PCI inhibition of APC in plasma.

Inhibition Of Activated Protein C In Combined Factor V/VIII Deficiency

1981 ◽  
Author(s):  
J C Giddings ◽  
A L Bloom
Keyword(s):  
Protein C ◽  
Activated Protein C ◽  
Haemophilia A ◽  
Factor V ◽  
Plasma Samples ◽  
Normal Human Plasma ◽  
Amidolytic Activity ◽  
Normal Plasma ◽  
Factor V Deficiency ◽  
Normal Human

Human prothrombin complex concentrate was chromatographed on a dextran sulphate/sepharose column to provide fractions with the recognised properties of protein C. Fractions treated with thrombincoupled sepharose yielded samples which prolonged the activated partial thromboplastin time of normal human plasma and which had amidolytic activity against the chromogenic substrate S2238. A recent report indicated that normal human plasma inhibited this amidolytic activity whereas plasma from patients with hereditary combined factor V/VIII deficiency failed to do so (Marlar, R.A, and Griffin, J.H., J. Clin. Invest, 66: 1186, 1980). In the present study 21 plasma samples from patients of 11 different families with combined factor V/VIII defect were compared with normal plasma for their ability to inhibit the amidolytic action of thrombin- treated protein C. Six patients with classical severe haemophilia A and two patients with severe, isolated factor V deficiency were also studied. Plasma was mixed with activated protein C together with heparin and antikallikrein serum. Aliquots were removed at intervals and incubated with substrate S2238. Hydrolysis was detected in a spectrophotometer at 405 nm. In eight separate experiments normal plasma inhibited an average 63.5% (±15.6) of the amidolytic activity in 30 mins, incubation. Plasma from patients with haemophilia A or isolated factor V deficiency gave results which were not significantly different from normal (67.2% and 61% respectively). However plasma from patients with the combined defect inhibited an average of 24.5% (±13.6) of the amidolytic activity (p <0.01). Two of these plasma samples failed to inhibit any protein C activity. The results confirm that normal plasma contains an inhibitor of activated protein C and this appears to be deficient in patients with hereditary combined factor V/VIII defect.

scholarly journals Proteolytic inactivation of human factor VIII procoagulant protein by activated human protein C and its analogy with factor V

Blood ◽  
1984 ◽  
Vol 63 (2) ◽  
pp. 486-489 ◽  
Author(s):  
CA Fulcher ◽  
JE Gardiner ◽  
JH Griffin ◽  
TS Zimmerman
Keyword(s):  
Factor Viii ◽  
Protein C ◽  
Human Factor ◽  
Activated Protein C ◽  
Light Chains ◽  
Factor V ◽  
Human Factor Viii ◽  
Thrombin Activation ◽  
A Site ◽  
Polypeptide Chains

Abstract Purified human factor VIII procoagulant protein (VIII:C) was treated with purified human activated protein C (APC) and the loss of VIII:C activity correlated with proteolysis of the VIII:C polypeptides. APC proteolyzed all VIII:C polypeptides with mol wt = 92,000 or greater, but not the doublet at mol wt = 79–80,000. These results and our previous thrombin activation studies of purified VIII:C, are analogous with similar studies of factor V and form the basis for the following hypothesis: activated VIII:C consists of heavy and light chain polypeptides [mol wt = 92,000 and mol wt = 79–80,000 (or 71–72,000), respectively] which are similar in Mr to the heavy and light chains of activated factor V. Thrombin activates VIII:C and V by generating these polypeptide chains from larger precursors and APC inactivates both molecules by cleavage at a site located in the heavy chain region of activated VIII:C and V.

scholarly journals Protein C Inhibitor Acts as a Procoagulant by Inhibiting the Thrombomodulin-Induced Activation of Protein C in Human Plasma

Blood ◽  
1998 ◽  
Vol 91 (5) ◽  
pp. 1542-1547 ◽  
Author(s):  
Marc G.L.M. Elisen ◽  
Peter A.Kr. von dem Borne ◽  
Bonno N. Bouma ◽  
Joost C.M. Meijers
Keyword(s):  
Tissue Factor ◽  
Protein C ◽  
Activated Protein C ◽  
Anticoagulant Effect ◽  
Clotting Time ◽  
Thrombin Inhibition ◽  
Normal Plasma ◽  
Protein C Inhibitor ◽  
Inhibition Of Thrombin

AbstractProtein C inhibitor (PCI), which was originally identified as an inhibitor of activated protein C, also efficiently inhibits coagulation factors such as factor Xa and thrombin. Recently it was found, using purified proteins, that the anticoagulant thrombin-thrombomodulin complex was also inhibited by PCI. The paradoxical inhibitory effect of PCI on both coagulant and anticoagulant proteases raised questions about the role of PCI in plasma. We studied the role of thrombomodulin (TM)-dependent inhibition of thrombin by PCI in a plasma system. Clotting was induced by addition of tissue factor to recalcified plasma in the absence or presence of TM, and clot formation was monitored using turbidimetry. In the absence of TM, PCI-deficient plasma showed a slightly shorter coagulation time compared with normal plasma. Reconstitution with a physiologic amount of PCI gave normal clotting times. Addition of PCI to normal plasma and protein C–deficient plasma resulted in a minor prolongation of the clotting time. This suggested that PCI can act as a weak coagulation inhibitor in the absence of TM. TM caused a strong anticoagulant effect in normal plasma due to thrombin scavenging and activation of the protein C anticoagulant pathway. This effect was less pronounced when protein C–deficient plasma was used, but could be restored by reconstitution with protein C. When PCI was added to protein C–deficient plasma in the presence of TM, a strong anticoagulant effect of PCI was observed. This anticoagulant effect was most likely caused by the TM-dependent thrombin inhibition by PCI. However, when PCI was added to normal plasma containing TM, a strong procoagulant effect of PCI was observed, due to the inhibition of protein C activation. PCI-deficient plasma was less coagulant in the presence of TM. A concentration-dependent increase in clotting time was observed when PCI-deficient plasma was reconstituted with PCI. The combination of these results suggest that the major function of PCI in plasma during coagulation is the inhibition of thrombin. A decreased generation of activated protein C is a procoagulant consequence of the TM-dependent thrombin inhibition by PCI. We conclude that TM alters PCI from an anticoagulant into a procoagulant during tissue factor-induced coagulation.

scholarly journals Proteolytic inactivation of human factor VIII procoagulant protein by activated human protein C and its analogy with factor V

Blood ◽  
1984 ◽  
Vol 63 (2) ◽  
pp. 486-489 ◽  
Author(s):  
CA Fulcher ◽  
JE Gardiner ◽  
JH Griffin ◽  
TS Zimmerman
Keyword(s):  
Factor Viii ◽  
Protein C ◽  
Human Factor ◽  
Activated Protein C ◽  
Light Chains ◽  
Factor V ◽  
Human Factor Viii ◽  
Thrombin Activation ◽  
A Site ◽  
Polypeptide Chains

Purified human factor VIII procoagulant protein (VIII:C) was treated with purified human activated protein C (APC) and the loss of VIII:C activity correlated with proteolysis of the VIII:C polypeptides. APC proteolyzed all VIII:C polypeptides with mol wt = 92,000 or greater, but not the doublet at mol wt = 79–80,000. These results and our previous thrombin activation studies of purified VIII:C, are analogous with similar studies of factor V and form the basis for the following hypothesis: activated VIII:C consists of heavy and light chain polypeptides [mol wt = 92,000 and mol wt = 79–80,000 (or 71–72,000), respectively] which are similar in Mr to the heavy and light chains of activated factor V. Thrombin activates VIII:C and V by generating these polypeptide chains from larger precursors and APC inactivates both molecules by cleavage at a site located in the heavy chain region of activated VIII:C and V.

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