The multiple coagulopathies of biliary atresia

Detailed coagulation studies were done prospectively on 43 patients with biliary atresia who had undergone Kasai operation (hepatic portoenterostomy). Patients were divided into three groups based on levels of factor V, factor II, and Echis II and/or response to vitamin K: no coagulopathy (46.5% of patients); coagulopathy of liver disease (30.2% of patients); and coagulopathy of vitamin K deficiency (23.3% of patients). Patients with the coagulopathy of liver disease had significantly lower levels of factors XII, V, and antithrombin III as well as longer thrombin times than patients with no coagulopathy or vitamin K deficiency. Factor V levels were decreased only in patients with more advanced liver disease; normal levels of factor V were not usually helpful in differentiating liver disease and vitamin K deficiency. The prothrombin time, factor VII-X levels, and factor II levels were significantly different for all three groups; the most abnormal values occurred in the vitamin K-deficient group. Comparison of the Echis II level to factor II coagulant activity was helpful in deciding whether a coagulopathy was due to liver disease, vitamin K deficiency, or both. Factor VIII levels were elevated in all groups. Factor VIII coagulant activity was significantly higher by the two-stage (TGT) method than by the one-stage (PTT) method. Hypersplenism causing neutropenia and thrombocytopenia was commonly seen after the age of 5 years. Vitamin E deficiency was more common than vitamin K deficiency; however, all vitamin K-deficient patients were vitamin E deficient. Coagulation status correlated well with hepatobiliary scan data, but not serum bilirubin levels. Recommendations for treatment of patients with vitamin K deficiency and/or liver disease are discussed.     

Coagulation factor activity in platelet concentrates stored up to 7 days: an in vitro and in vivo study

The in vitro and in vivo recovery of coagulation factor activity in platelet concentrates stored up to 172 h was studied. In vitro studies revealed that fibrinogen and antithrombin III levels do not change with storage. Factors II, V, VII, VIII, IX, X and XI all showed statistically significant falls from baseline over the 172 h storage period. However, most factor activities remained above 70%, with the major exception of factors V and VIII. These factors fell to less than 30% activity over the storage period, consistent with their known lability during whole blood storage. In vivo studies after platelet concentrate infusion in patients with concurrent thrombocytopenia and coagulation deficiencies revealed that the measured in vitro activity was recoverable in vivo. We conclude that platelet concentrates stored for 172 h are an adequate source of clotting factors. However, like stored whole blood, they may not provide therapeutic doses of factors V and VIII.     

Effect of DDAVP on plasma level of factor XII

The effect of DDAVP on blood coagulation factors was investigated after its intravenous infusion into normal subjects. A marked increase in factor XII was observed in addition to the expected rise of factor VIII coagulant activity (VIII:C), factor VIII related antigen (VIIIR:Ag) and plasminogen activator, DDAVP also produced a concomitant but less pronounced rise of factor VII, but there was no change in factors V, IX, X and XI.     

HYPOFIBRINOGENÆMIA IN ACUTE LEUKÆMIA WITH EXTENSIVE FIBRINOUS PERICARDITIS

A case of acute myeloid leukæmia and hypofibrinogenæmia is described. The hypofibrinogenæmia was associated with decreased levels of factor V and factor VIII, but there was no increase in fibrinolytic activity. The clinical course was marked by a severe hæmorrhagic diathesis and evidence of cardiac tamponade. At autopsy there was extensive fibrinous pericarditis. It is suggested that the extensive pericardial fibrin deposition might have contributed to the coagulation abnormality observed.     

A study of the cation- and pH-dependent stability of factors V and VIII in plasma.

The in vitro lability of factors V and VIII in plasma has been studied. In agreement with previous reports, an increase in anticoagulant concentration renders both factors more labile (cation-deficient decay), as does an increase in the pH above 7.3 (alkaline-decay). Calcium appears to be the plasma cation which protects factors V and VIII against in vitro loss of activity. The protection obtained by the addition of other divalent cations depended on the type of plasma used. When resin-EDTA plasma was made cation free by dialysis at 4 degrees C and then incubated at 37 degrees C, the rapid loss of factors V and VIII activity could be prevented by prior addition of strontium, manganese and magnesium. In oxalate plasma, nickel, manganese, cadmium and strontium were effective. The alkaline decay of both factors V and VIII is irreversible. Partial reversibility of the cation-deficient decay was demonstrated for factor V, but not for factor VIII. The temperature coefficient for both the cation-deficient and alkaline decay is 2-3, suggesting an enzymatic rather than a physical reaction. There was no evidence to implicate thrombin, plasmin or trypsin since inhibitors of these enzymes failed to modify either type of decay.     

Combined factor V/VIII deficiency: a case report including levels of factor V and factor VIII coagulant and antigen as well as protein C inhibitor

Comprehensive coagulation studies were performed on members of a family with combined factor V/VIII deficiency. The purpose of these studies was to investigate the hypothesis that combined factor V/VIII deficiency is due to a lack of the inhibitor to activated protein C. The analyses performed included routine APTT and PT, factor V and VIII coagulant activity and antigen levels, von Willebrand factor levels, protein C antigen assay, and both protein C inhibitor activity and antigen levels. Three of the 19 family members studied were found to have a deficiency of both factors V and VIII. These three individuals showed prolonged APTTs and PTs and decreased levels of factor V and factor VIII coagulant activity and antigen. Factor VIII related antigen and ristocetin cofactor (von Willebrand factor) levels were normal. Protein C and both protein C inhibitor activity and antigen levels were also found to be normal. These findings confirm the results of other recent investigators and indicate that the autosomal, inherited combined factor V/VIII deficiency is not due to a protein C inhibitor deficiency. The real defect in this combined deficiency remains to be determined.     

Hemostatic changes in active pulmonary tuberculosis.

OBJECTIVE: Severe pulmonary tuberculosis (PTB) is sometimes complicated by deep vein thrombosis (DVT). We have searched for possible hemostatic disturbances that are predisposing factors for venous thrombosis in patients with PTB. DESIGN: Coagulation and platelet function tests were studied in 45 patients with active PTB and 20 healthy control volunteers before therapy. Findings were compared with results at 30 days. RESULTS: Analysis in patients with active PTB showed anemia, leucocytosis, thrombocytosis, elevation in plasma fibrinogen, factor VIII, plasminogen activator inhibitor 1 (PAI-1) with depressed antithrombin III (AT III) and protein C (PC) levels. On the 30th day of treatment, anemia, leucocytosis and thrombocytosis were improved. Fibrinogen and factor VIII levels had decreased to normal levels, PC and AT III levels had increased to normal levels, and there was no difference in PAI-1 levels. We found no activated protein C resistance. Platelet aggregation studies demonstrated increased platelet activation. However, DVT was not detected in patients during the follow-up period. CONCLUSION: Decreased AT III, PC and elevated plasma fibrinogen levels and increased platelet aggregation appear to induce a hypercoagulable state seen in PTB and improve with treatment.     

DDAVP administration in a case of congenital combined factor V and factor VIII deficiency

Combined deficiency of factor V and factor VIII, a rare bleeding disorder, was found in a 43 year-old male. He had often presented manifestations of easy bruising since childhood, but none of his family had shown evidence of a bleeding tendency. We examined him and his family as far as we could and his abnormality of blood coagulation was apparent, but the members of his family were normal. The prothrombin time and activated partial thromboplastin time of this patient were prolonged, but his thrombin time was normal. Factor V and factor VIII coagulant activity were low, but von Willebrand factor antigen and activity (ristocetin cofactor activity) levels were normal. Protein C and Protein C inhibitor antigen and activity levels were also found to be normal. Following 1-deamino-8-D-arginine vasopressin (DDAVP) injection, he had immediate increases in factor VIII coagulant activity, but both von Willebrand factor antigen, activity levels and factor V coagulant activity remained low. Moreover, there was no rapid decline in factor VIII complex activity. These findings suggest that the endogenous factor VIII in this patient is metabolized normally and that at least the deficiency of factor VIII does not result from accelerated degradation in plasma.     

The effect of liver disease on factors V, VIII and protein C

The components of the factor VIII complex were estimated by immuno- and bioassays in 85 patients with liver disease. The plasma concentrations of the antigens were elevated in 65% (VIII:CAg) and in 76% (VIIIR:Ag) of patients while the biological activities were elevated in only 14% (VIII:C) and 15% (VIII:RiCof). There was no correlation with C-reactive protein, used as a measure of an acute phase reaction (X2 = 0.7; P = 0.1); or with severity of liver disease as judged by prothrombin ratio (P = 1.0) but highest values were observed in patients with cholestatic liver disease. Following parenteral vitamin K there was a significant fall in both the biological activity of VIIIC (36%) and of VIII:CAg (38%) in 13 vitamin K deficient patients (P less than 0.001) but no change in 23 vitamin K replete patients or in the VIIIR:Ag levels in either group. Factor V levels were lower in patients with parenchymal liver disease (0.54 +/- 0.1 units/ml, mean +/- SEM, n = 12; normal range 0.5-1.5 units/ml) than in patients with extrahepatic cholestasis who were vitamin K deficient (1.2 +/- 0.1 units/ml, P less than 0.0001). The levels of protein C antigen, the vitamin K dependent protease which inactivates factors VIII:C and V, was at the lower end of the range in both groups (0.7 +/- 0.1, mean +/- SEM, n = 18, normal range 0.74-1.4 units/ml). There was no significant change in either protein C antigen or factor V following vitamin K. The discrepancy between the biological activity of factor VIII and the antigen levels could represent accumulation of partially degraded factor VIII or production of a hypoactive form. There is no evidence that the reduction in VIIIC and VIII:CAg following vitamin K was mediated by protein C.     

Factor V antigen levels and venous thrombosis: risk profile, interaction with factor V leiden, and relation with factor VIII antigen levels

Clotting factor V has a dual function in coagulation: after activation, procoagulant factor V stimulates the formation of thrombin, whereas anticoagulant factor V acts as a cofactor for activated protein C (APC) in the degradation of factor VIII/VIIIa, thereby reducing thrombin formation. In the present study, we evaluated whether plasma factor V levels, either decreased or increased, are associated with venous thrombosis. High procoagulant factor V levels may enhance prothrombinase activity and increase the thrombosis risk. Low anticoagulant factor V levels could reduce APC-cofactor activity in the factor VIII inactivation (APC-resistant phenotype), which might also promote thrombosis. Low factor V levels in combination with factor V Leiden could lead to a more severe APC-resistant phenotype (pseudohomozygous APC resistance). To address these issues, we have measured factor V antigen (factor V:Ag) levels in 474 patients with thrombosis and 474 control subjects that were part of the Leiden Thrombophilia Study (LETS). Factor V:Ag levels increased by 7.6 U/dL for every successive 10 years of age. Mean factor V:Ag levels were 134 (range 41 to 305) U/dL in patients and 132 (range 47 to 302) U/dL in controls. Neither high nor low factor V:Ag levels were associated with venous thrombosis. We found that factor V:Ag and factor VIII antigen levels in plasma were correlated, but factor V did not modify the thrombotic risk of high factor VIII levels. The normalized APC ratio was not influenced by the factor V:Ag level in subjects with or without factor V Leiden. We conclude that neither low nor high factor V:Ag levels are associated with venous thrombosis and that factor V:Ag levels do not mediate the thrombotic risk associated with high factor VIII levels.     

Combined deficiency of factor V and factor VIII. A report of another case

Summary A patient with combined factor V and factor VIII deficiency is presented. The bleeding manifestations were: easy bruising, post-traumatic bleeding, bleeding after tooth extractions. The main laboratory feature was a prolonged partial thromboplastin time which was corrected by the addition of adsorbed normal plasma but not by the addition of normal serum, hemophilia A plasma or plasma of another patient with combined factor V and factor VIII deficiency. The thromboplastin generation test was clearly abnormal and was corrected by the addition of adsorbed normal plasma but not by the addition of normal serum. Prothrombin consumption was also defective.Prothrombin time was slightly prolonged too, Thrombin time, platelet and vascular tests were within normal limits and there was no hyperfibrinolysis. Factor VIII was 8% of normal, whereas factor V was 14% of normal. Factor VIII associated antigen was normal. All other clotting factors were within normal limits.The parents of the propositus were consanguineous (first cousins) but had normal factor V and factor VIII activity and normal factor VIII antigen. The same was true for other family members. The hereditary transmission of the condition appears autosomal recessive.This study was supported in part by a grant from the C.N.R. (grant CT. 74.00189.04).     

Increased factor VIII/vWf levels in patients with reduced platelet number

Summary Factor VIII/von Willebrand factor (VIII/vWf) related properties were studied in twenty six patients with thrombocytopenia. Fifteen patients were affected by idiophatic thrombocytopenic purpura (ITP) and 11 patients by thrombocytopenia of a different nature or non-ITP (n-ITP). All patients showed an enhancement of platelet associated IgG (PAIgG). A significant increase of factor VIII ristocetin cofactor (VIII R: RCoF) and factor VIII related antigen (VIII R: Ag) was found in ITP patients while normal values were observed for factor VIII coagulant (VIII: C). All factor VIII/vWf components, on the contrary, were increased in n-ITP group with a prevalence of VIII R: RCoF as observed in ITP group even though with lower mean values. Multimeric analysis of VIII/vWf demonstrated a higher concentration of all multimeric components, with major representation of higher molecular weight multimers (HMWM) in patients of both groups.Two patients were studied before and after improvement in platelet count. A decrease of vWf related properties (VIII R : RCoF and VIII R : Ag) concomitant with the increase in platelet count was found. In n-ITP patients a statistical correlation between VIII R : RCoF and PAIgG was also observed while no correlation was found between other factor VIII/vWf components and PAIgG both in ITP and n-ITP patients.     

Type IIB von Willebrand''s disease presenting as thrombocytopenia during pregnancy

Type IIb von Willebrand's disease has been found to be associated with the development of thrombocytopenia following the infusion of DDAVP (desmopressin). It has also been associated with sporadic thrombocytopenia and evidence of spontaneous platelet aggregation. A family with documented Type IIb von Willebrand's disease is described, where two of the affected females presented with moderate to severe thrombocytopenia developing during pregnancy with reversal to normal or minimally reduced platelet counts in the early post gestational period. In each case, the levels of factor VIII:C, von Willebrand factor antigen and von Willebrand factor ristocetin co-factor activity rose during pregnancy but there were notable discrepancies between the levels of each in any one individual. It is suggested that pregnancy resulted in increased synthesis of the variant form of von Willebrand factor resulting in progressively increasing platelet/variant form von Willebrand factor interaction and subsequent thrombocytopenia. Whether this reflects consumption or sequestration remains uncertain. Although spontaneous platelet aggregation was observed in some family members, the majority did not exhibit this phenomenon. Circulating platelet aggregates could not be detected. Both pregnancies were relatively uneventful and there is no history of unusual bleeding associated with pregnancy in the family. These observations suggest that Type IIb von Willebrand's disease should be considered in the differential diagnosis of thrombocytopenia developing during pregnancy, particularly in those individuals where evidence supporting the diagnosis of immune mediated thrombocytopenia is not forthcoming. Where the diagnosis of Type IIb von Willebrand's disease is established, active intervention other than confinement in a hospital with experience in haemostatic disorders is probably not required as the development of thrombocytopenia does not appear to exert an additive effect on the underlying defect relating to the variant form of von Willebrand's disease.     

High plasma levels of factor VIII: an important risk factor for isolated pulmonary embolism

OBJECTIVE: The aim of the study was to investigate whether factor V Leiden and prothrombin G20210A mutations, elevated levels of factor VIII and factor IX are associated with pulmonary embolism (PE). METHODS: Sixty-four patients with objectively documented PE and 64 control subjects were included in this study. The authors divided the 64 subjects with PE into those with PE and deep vein thrombosis (combined form of venous thromboembolism, n = 26) and those with PE without deep vein thrombosis (isolated PE n = 38). RESULTS: There was no significant difference between the PE groups and the control subjects with regard to the presence of factor V Leiden and prothrombin mutations and elevated levels of factor IX. Using the 90th percentile measured in control subjects (P(90) = 168 U/dL) as a cut-off point for factor VIII levels, the authors found an 11-fold increased risk for both isolated PE patients and patients with a combined form of venous thromboembolism who have factor VIII levels >168 U/dL compared with individuals having factor VIII levels below this cut-off point. The risk was not affected by adjustments for other possible risk factors. CONCLUSIONS: Elevated plasma factor VIII levels were found to be a significant, independent risk factor for PE.     

Factor VIII expression in azoxymethane-induced murine fulminant hepatic failure

Fulminant hepatic failure (FHF) in humans produces a bleeding diathesis due in large part to a reduction in the biosynthesis of liver-derived coagulation factors. Remarkably, factor VIII procoagulant activity is elevated in most of these patients despite widespread liver cell death. FHF can be modeled in mice by administration of azoxymethane, the active ingredient found in cycad palm nuts. We compared the expression of factor VIII to other hepatic hemostatic factors in azoxymethane-induced murine FHF. Mice displayed dose-dependent decreases in all coagulation factor activities measured, including factors V, VII, VIII, and IX. At the highest dose of azoxymethane (50 microg/g body weight), factor VIII activity in plasma decreased by 98% within 36 hours after treatment, which was associated with an 80% reduction in hepatic factor VIII messenger RNA (mRNA). In contrast, factor VIII mRNA levels in spleen, kidney, and lung tissue of azoxymethane-treated mice were unchanged. Cellular damage in these mice appeared to be limited to hepatocytes as evident by histologic examination. This finding is supported by 2 observations. First, hepatic mRNA levels of von Willebrand factor, which is synthesized by liver sinusoidal endothelial cells but not hepatocytes, were unchanged. Second, von Willebrand factor was detected antigenically in liver sections of azoxymethane-treated mice by immunofluorescence. These results indicate that the contribution of the liver to factor VIII biosynthesis is not replaced or significantly supplemented by other tissues in this model of FHF.     

Hemostatic alterations in inflammatory bowel disease

Twelve patients with acute, untreated inflammatory bowel disease (IBD) were followed prospectively for coagulation and platelet function. With no symptomatic coagulopathy, abnormalities were found in all patients. With acute diseases, elevations of fibrinogen (9/12), factor V (8/12), and factor VIII (6/12) were common. Depressions of antithrombin III levels were also observed acutely (8/12). Abnormalities of platelets were both quantitative and qualitative. Thrombocytosis was present (11/12), and abnormalities in the rate and percent platelet aggregation were seen (9/10). During therapy, factors V and VIII, antithrombin III levels, and the quantitative and qualitative platelet abnormalities returned towards normal in direct correlation with sedimentation rate and clinical disease activity.     

Original Article: Changes in Platelet Function, von Willebrand Factor and Factor VIII in Patients Undergoing Aorto-bifemoral By-pass with Dacron Grafts

Platelets play a major role in the development of patency complications in vascular grafts. The aim of this study was to evaluate changes in platelet count and function, and also in factor VIII:C (FVIII:C) and von Willebrand factor (vWF) plasma levels, induced by aorto-bifemoral by-pass with Dacron grafts in seven patients. Platelet count, platelet aggregate ratio (PAR), and platelet aggregability induced by several stimuli, as well as FVIII:C and vWF plasma levels were evaluated before and on days 1,4,9 and 11 after surgery. We observed a mild thrombocytopenia on day 1, followed by a progressive increase in platelet count, which attained a relative thrombocytosis on the 11th day. PAR did not vary significantly during the whole observation period. Platelet aggregation, assayed by the optical method using ADP, epinephrine, arachidonic acid, collagen and ristocetin, (decreased on days 1 and 4). Thereafter, an increase in aggregation was observed until day 11 when hyperaggregability was verified. FVIII:C and vWF peaked on the 4th day, decreasing progressively to pre-surgery values on day 11.     

Original Article: Changes in Platelet Function,von Willebrand Factor and Factor VIII in Patients Undergoing Aorto-bifemoral By-pass with Dacron Grafts

Platelets play a major role in the development of patency complications in vascular grafts. The aim of this study was to evaluate changes in platelet count and function, and also in factor VIII:C (FVIII:C) and von Willebrand factor (vWF) plasma levels, induced by aorto-bifemoral by-pass with Dacron grafts in seven patients. Platelet count, platelet aggregate ratio (PAR), and platelet aggregability induced by several stimuli, as well as FVIII:C and vWF plasma levels were evaluated before and on days 1,4,9 and 11 after surgery. We observed a mild thrombocytopenia on day 1, followed by a progressive increase in platelet count, which attained a relative thrombocytosis on the 11th day. PAR did not vary significantly during the whole observation period. Platelet aggregation, assayed by the optical method using ADP, epinephrine, arachidonic acid, collagen and ristocetin, (decreased on days 1 and 4). Thereafter, an increase in aggregation was observed until day 11 when hyperaggregability was verified. FVIII:C and vWF peaked on the 4th day, decreasing progressively to pre-surgery values on day 11.     

Combined factor V-VIII deficiency: a case report with studies of factor V and VIII activation by thrombin

A new case of combined factor V-VIII deficiency is reported with in vitro studies of factors V and VIII activation by thrombin. The normal activation of factors V and VIII demonstrated in the patient's plasma and the equivalent levels of factor VIII coagulant activity and coagulant antigen support the hypothesis that a quantitative rather than qualitative defect in factors V and VIII is present in this disorder.     

Selective factor VIII and V inactivation by iminodiacetate ion exchange resin through metal ion adsorption

The procoagulant activity of factors VIII and V depends on the presence of metal ion(s). We examined the effect of cation-exchange resins with different functional groups on both factors, of which only reaction with iminodiacetate resin resulted in the complete loss of their activity levels in plasma. However, the antigen level of factor VIII was preserved by >95%. This resin reduced divalent cations content present in factor VIII preparations, indicating that it inactivated this factor by direct deprivation of predominant Ca(2+) (>Mn(2+)>Cu(2+)), rather than adsorption of the factor itself. The antigen level of recombinant factor VIII alone was decreased by >95% by reaction with resin, whilst that complexed with von Willebrand factor was preserved by >95%. Iminodiacetate resin-treated plasma was evaluated by measuring factor VIII and V activity in plasma with various levels of either activity. These were significantly correlated to the values obtained using factor VIII- or V-deficient plasma prepared commercially by immunodepletion. We demonstrated that iminodiacetate resin-induced factors VIII and V inactivation is because of direct deprivation of metal ions, predominantly Ca(2+), which is more essential for the functional structure of their molecules. Furthermore, iminodiacetate resin-treated plasma would be useful as a substrate for measuring the activity of these factors.