Protein Z, protein S levels are lower in patients with thrombophilia and subsequent pregnancy complications.

OBJECTIVE: We posit that low levels of protein S (PS) and protein Z (PZ) contribute to adverse pregnancy outcome (APO). PATIENTS: We evaluated 103 women with subsequent normal pregnancy outcome (NPO), 106 women with APO, and 20 women with thrombophilia (TP). METHODS: We compared first trimester (1st TRI) PZ levels in 103 women with NPO, 106 women with APO, and in 20 women with TP. We compared plasma levels of PZ and free PS antigen during the second (2nd TRI) and third trimesters (3rd TRI) of pregnancy in 51 women with APO and 51 matched women with NPO. RESULTS: The mean 1st TRI PZ level was significantly lower among patients with APO, compared to pregnant controls (1.81 +/- 0.7 vs. 2.21 +/- 0.8 microg mL(-1), respectively, P < 0.001). Of patients with known TP, those with APO had a tendency for lower mean PZ levels compared to those TP women with NPO (1.5 +/- 0.6 vs. 2.3 +/- 0.9 microg mL(-1), respectively, P < 0.0631). There was a significant decrease in the PZ levels in patients with APO compared to NPO (2nd TRI 1.5 +/- 0.4 vs. 2.0 +/- 0.5 microg mL(-1), P < 0.0001; and 3rd TRI 1.6 +/- 0.5 vs. 1.9 +/- 0.5 microg mL(-1), P < 0.0002). Protein S levels were significantly lower in the 2nd and 3rd TRIs among patients with APO compared to patients with NPO (2nd TRI 34.4 +/- 11.8% vs. 38.9 +/- 10.3%, P < 0.05, respectively; and 3rd TRI 27.5 +/- 8.4 vs. 31.2 +/- 7.4, P < 0.025, respectively). CONCLUSIONS: We posit that decreased PZ and PS levels are additional risk factors for APO.

The use of coagulation activation markers (soluble fibrin polymer, TpP, prothrombin fragment 1.2, thrombin-antithrombin, and D-dimer) in the assessment of hypercoagulability in patients with inherited and acquired prothrombotic disorders.

A total of 260 consecutive patients, referred for hypercoagulable assessment, was included in this study. Four coagulation activation markers were utilized to assess these patients [enzyme-linked immunosorbent assays for soluble fibrin polymer (TpP), prothrombin fragment 1.2, thrombin-antithrombin complex, and D-dimer]. The mean levels of the activation markers directly correlated with the number of hypercoagulable abnormalities. The percentage of patients with increased TpP levels for each group was lower than the other activation markers. The findings indicate that activation markers reflect the number of underlying thrombophilic abnormalities. Our data suggest that there is a utility in performing a panel of coagulation activation markers to assess the thrombotic risk. The measurement of soluble fibrin polymer may be more reflective of an impending vascular event.

Thrombophilia and pregnancy: review of the literature and some original data.

The association of thrombophilia with pregnancy complications has received increasing attention. It is now apparent that thrombophilia is responsible for a large number of the serious complications of pregnancy such as venous thrombosis, pulmonary embolism, fetal loss, pregnancy loss, intrauterine fetal demise, and preeclampsia. The inherited thrombophilia abnormalities, factor V Leiden mutation, prothrombin gene mutation 20210A, and antithrombin III, protein C, and protein S deficiency, and the acquired disorders, the anticardiolipin syndrome and lupus inhibitor, are responsible for a large share of the incidences of premature termination of pregnancy and many of the above complications. The normal physiology of pregnancy may be prothrombotic, with evidence for increased markers of activated coagulation and coagulation factors. There is a decrease in protein S and resistance to activated protein C occurs in a significant number of pregnancies in the absence of the factor V Leiden mutation. In the following article, we review some of the major studies that have correlated the thrombophilia and other acquired disorders that adversely impact pregnancies.

Thrombosis and cancer.

Thrombosis is a common complication of malignancy. This is felt to be related to increased activity of the coagulation system as evidenced by markers of accelerated thrombin generation and increased platelet reactivity. Alterations in the hemostatic balance have been documented in patients with malignancy with increased tissue factor (TF) generation and the production of a cysteine protease. These can stimulate the coagulation mechanism via the extrinsic pathway and/or by activating factor X. The thrombotic presentations in malignancy are protean and may be venous or arterial. The underlying clinical pictures may be related to varying degrees of consumptive coagulopathy, microangiopathy, and nonbacterial endocarditis. Prophylaxis and management are, to a significant degree, dependent on the underlying malignancy and the prothrombotic mechanism. Specific agents and drugs must be selected from an expanding menu of options that includes unfractionated heparin, low-molecular-weight heparin (LMWH), warfarin, plasma apheresis, and the newer antithrombin agents.

The use of thrombus precursor protein, D-dimer, prothrombin fragment 1.2, and thrombin antithrombin in the exclusion of proximal deep vein thrombosis and pulmonary embolism.

We examined various nonSTAT commercially available coagulation activation markers in an attempt to help diagnose or exclude the often subtle clinical presentations of proximal deep vein thrombosis (PDVT) and pulmonary embolism (PE). Fifty-five patients presenting to the Emergency Department were completely assessed. Eleven patients were diagnosed with PDVT, six patients were diagnosed with PE, and three patients were diagnosed with both PDVT and PE. Thrombus precursor protein (TpP) excluded the diagnosis in 19 of the 35 patients negative for PDVT and/or PE, D-Dimer in 15 patients, prothrombin fragment 1.2 in 17 patients, and thrombin-antithrombin (TAT) in 14 patients. Both the TpP and TAT enzyme-linked immunosorbent assay (ELISA) tests had 100% sensitivity and negative predictive value for evaluating PDVT and/or PE. The TpP ELISA had the highest specificity (54%) of all four markers studied.

Ischemic stroke in a young patient with protein C deficiency and prothrombin gene mutation G20210A.

We report a patient who had an ischemic stroke aged 22 years, an inherited type I protein C deficiency and a heterozygous genotype of prothrombin gene 20210A. In view of recent reports of an increased risk for ischemic cerebral vascular disease in patients with the prothrombin 20210A mutation, we suggest that many of the reported cases of ischemic stroke and protein C deficiency may have had additional prothrombotic disorders such as the prothrombin mutation. The current data concerning the magnified risk for stroke in patients with the prothrombin 20210A mutation suggests the need to study all patients with premature stroke for this mutation and the other risk factors for thrombosis. This would include homocysteine, lupus inhibitor, anticardiolipin antibodies, and possibly the natural inhibitors of coagulation. It is possible that patients with the prothrombin 20210A mutation and ischemic cerebral vascular disease would benefit from long-term anticoagulation therapy in a similar way to patients with the antiphospholipid syndrome.

A highly specific functional test for factor V leiden: A modified tissue factor assay for activated protein C resistance.

We compared the sensitivity and specificity of a tissue factor-based assay (FVR) with the addition of a phospholipid/silica preparation, to the commercially available aPTT-based method, APCR (CoatestTM), and a modified aPTT-based method (APCM) which utilized factor V-depleted plasma, for the detection of the factor V Leiden mutation. A total of 110 patients were included in this study. This included 32 patients on coumadin therapy, 7 patients on heparin therapy, 5 patients on both anticoagulants therapy, and 24 patients who were positive for anticardiolipin antibody (ACL) and/or lupus inhibitor (LI). Our data demonstrate that the FVR is not affected by anticoagulation treatment or ACL/LI antibodies, whereas in the APCR method, 33 patients cannot be determined either due to the anticoagulant therapy or presence of the ACL and/or LI. With the APCM method, the clotting endpoint could not be determined in 1 patient due to the presence of a strong LI. The additional phospholipid/silica material utilized in the FVR enhanced the APC degradation of factor Va and therefore sharpened the demarcation between the factor V Leiden-positive and -negative patients. The sensitivity for the APCR, APCM and FVR was 42, 97 and 100% respectively. The specificity for the APCR, APCM and FVR was 94, 96 and 100% respectively.

Acquired von Willebrand's syndrome in association with a lupus-like anticoagulant corrected by intravenous immunoglobulin.

We are reporting on a 47-year-old man who presented with a prolongation of the activated partial thromboplastin time (APTT) prior to orthopedic surgery. An evaluation suggested an inhibitor when his plasma prolonged a normal control APTT upon 50:50 solution of patients with normal plasma. The platelet-neutralizing procedure (PNP), anticardiolipin antibody, and antinuclear antibody (ANA) were positive. Further studies revealed decreased von Willebrand factor ristocetin cofactor (vWF:RCoF), von Willebrand factor antigen (vWF:Ag), an inhibitor to vWF, and absent high-molecular-weight vWF multimeters. Assays of FVIII:C, FIX, and FXI were nonparallel to the standard curve. Intravenous immunoglobulin (IVIG) corrected the APTT, multimeric pattern, and FVIII:C by the 7th day postinfusion. This case demonstrates the efficacy of IVIG for acquired von Willebrand's syndrome (vWS) and also represents a unique combination of a lupus-like anticoagulant and acquired vWS in a patient without the full serological requirement for systemic lupus erythematosus (SLE). Whether patients with acquired vWS and lupus inhibitors are more or less susceptible to either a thrombotic complication or hemorrhage is not established. Prospective studies for the incidence of lupus inhibitor/antiphospholipid syndromes and vWF deficiencies are needed to assess this question.

Effect of intravenous immunoglobulin on inhibition of fibrinogen binding to platelets by sera from patients with immune thrombocytopenia.

We previously described an ELISA to measure the inhibition of platelet glycoprotein IIb/IIIa (GPIIb/IIIa) binding to fibrinogen due to immune complexes and/or anti-platelet antibodies from patients with immune thrombocytopenia (ITP) or HIV-related ITP. Circulating immune complexes (CIC) were the main factor in the inhibition of GPIIb/IIIa binding to fibrinogen in HIV-related ITP, whereas in non-HIV ITP, inhibition was only partially due to CIC; anti-platelet antibodies specific to GPIIIa were also shown to play a role. In this study, we correlated the rise in the platelet count after intravenous immunoglobulin (IVIG) infusion with the decrease in inhibition of fibrinogen binding to GPIIb/IIIa by the sera of patients with ITP and HIV-related ITP. In the majority of the patients' sera tested, as the platelet count increased following the administration of IVIG, the degree of inhibition of GPIIb/IIIa binding to fibrinogen decreased. We also observed a decrease and/or disappearance of the antibodies specific to GPIIb and/or GPIIIa after IVIG administration. In HIV-seronegative ITP patients, the decrease or disappearance of anti-platelet antibodies directly correlated with the decreased inhibition of GPIIb/IIIa binding to fibrinogen by the 2% PEG supernatants of sera which contained anti-platelet antibodies. These findings suggest that IVIG directly affects the binding of CIC and anti-platelet antibodies to platelets and thereby improves platelet survival. Our results also suggest that the anti-idiotypic effect may contribute to IVIG's therapeutic action. In contrast, in the HIV-seropositive group, the decreased inhibition by PEG precipitates after IVIG administration was more strongly associated with an increase in the platelet count.

Dysfunctional platelet glycoprotein IIb/IIIa associated with a platelet release defect: a family study.

We are reporting on a 36-year-old white female with a bleeding history attributed to dysfunctional platelet glycoprotein IIb/IIIa (GPIIb/IIIa) and a coexisting platelet release defect. Platelet aggregation studies (PAS) revealed markedly diminished to absent responses to ADP, epinephrine, collagen and arachidonic acid; the ristocetin response was normal. ATP content was normal with poor release to the agonists as measured by luminescent technique. DDAVP infusion shortened bleeding time from 13.5 min to 8.0 and 12 min (at 1 and 2 hours). Flow cytometry and immunoblotting revealed normal amounts of GPIIb and diminished GPIIIa (50% of control). Using a previously reported ELISA which measures the binding of GPIIb/IIIa to immobilized fibrinogen, the patient's platelet extract showed no binding to fibrinogen. Both the father and mother were found to have decreased PAS responses and normal amounts of GPIIb/IIIa determined by both Western blot and flow cytometry. However, the ELISA showed decreased binding of their GPIIb/IIIa to fibrinogen (71% and 62% as compared to controls, respectively). The patient's dysfunctional fibrinogen receptor was clearly demonstrated by the ELISA. The parents had moderately reduced GPIIb/IIIa function in this assay, but they did not demonstrate a reduced GPIIIa as was noted in the patient. The parents' PAS indicated a platelet release defect. These findings suggest an inherited platelet release defect and a dysfunctional GPIIIa. The partial response to DDAVP would be compatible with the presence of a platelet release defect.

Inhibition of human platelet glycoprotein IIB/IIIA binding to fibrinogen by tumor cell membrane proteins.

Immunochemical and functional characteristics of tumor cell membrane proteins and human platelet glycoproteins were studied. Immunoblotting revealed that membrane proteins of a cultured breast tumor cell line (BT-20) had three protein bands, which were each recognized by monoclonal antibodies to human platelet glycoprotein Ib, IIb, and IIIa, suggesting some immunochemical similarities between the tumor cell membrane proteins and platelet glycoproteins. The monoclonal antibodies failed to bind to an extract of a lung tumor cell line (A549). Neither tumor extract induced platelet aggregation. However, tumor-associated antigens isolated from the breast tumor cells markedly inhibited platelet glycoprotein IIb/IIIa binding to fibrinogen. In contrast, tumor-associated antigens from the lung tumor cells had no effect. These results suggest that tumor cells which have immunological and/or structural similarities to platelets may affect hemostasis and coagulation in vivo.