Quantification of the effects of thrombin activatable fibrinolysis inhibitor and alpha2-antiplasmin on fibrinolysis in normal human plasma.

Two major proteins that inhibit fibrinolysis include thrombin activatable fibrinolysis inhibitor (TAFI) and alpha2-antiplasmin. Our goal was to quantify the contribution of TAFI and alpha2-antiplasmin to antifibrinolytic defenses with thrombelastography. Plasma activated with tissue factor/kaolin was subjected to fibrinolysis with tissue-type plasminogen activator (100 U/ml). Prior to activation, TAFI activity was inhibited with either potato carboxypeptidase inhibitor (25 microg/ml) or an anti-TAFI antibody, and alpha2-antiplasmin activity was inhibited with an anti-alpha2-antiplasmin antibody. Data were collected for 30 min, with the time of onset and rate of fibrinolysis determined. Compared with uninhibited samples, TAFI inhibition significantly (P < 0.05) decreased the time of onset of fibrinolysis by 70% and increased the rate of lysis by 70%. There was no difference between potato carboxypeptidase inhibitor and anti-TAFI antibody inhibition. Inhibition of alpha2-antiplasmin resulted in a significantly (P < 0.05) decreased time of onset (85%) and increased the rate of lysis (557%) compared with uninhibited samples. Inhibition of alpha2-antiplasmin activity resulted in a significantly (P < 0.05) greater fibrinolytic response than TAFI inhibition. In conclusion, utilization of standard inhibitors and thrombelastography permitted quantification of the effects of TAFI and alpha2-antiplasmin on fibrinolysis in plasma. Future investigation of diseases involving hypofibrinolysis (e.g. left ventricular assist devices) could be conducted using this assay system.

Thrombelastographic measures of clot propagation: a comparison of alpha with the maximum rate of thrombus generation.

The alpha angle alpha (degrees) is a thrombelastographic measure of clot propagation. A parametric measurement of clot propagation [maximum rate of thrombus generation (MRTG), dynes/cm2 per s], however, has recently been utilized. Thus, the relationship of changes in alpha with changes in MRTG were determined. alpha and MRTG values obtained from 859 thrombelastograms was collected from nine studies. Data were analyzed and the relationship between alpha and MRTG defined with commercially available software. Additional comparisons were made retrospectively from whole-blood and plasma data obtained from 33 normal individuals. Data from the nine studies demonstrated that MRTG increased in an exponential fashion compared with increases in alpha (R2 = 0.88, P < 0.001). Whole-blood alpha values were in the range 66.7-74.7 whereas MRTG values were 5.5-10.8, and plasma alpha values were 65.1-77.9 with corresponding MRTG values of 3.5-12.0. Assessment of clot propagation utilizing MRTG provides a more parametric evaluation than the determination of alpha. While normal alpha values may vary by only 12-20%, MRTG values vary by approximately 200-300%. The MRTG should be progressively utilized to a greater extent in both laboratory and clinical settings to parametrically quantify clot growth kinetics with thrombelastography.

Thrombelastographic method to quantify the contribution of factor XIII to coagulation kinetics.

Factor XIII (FXIII) plays a critical role in clot strength, and FXIII deficiency or excess is associated with hemorrhage or thrombosis, respectively. Our goal was to design a thrombelastography-based method to characterize the effects of FXIII on plasma clot strength. Normal human plasma was exposed to 0 or 200 mug/ml anti-FXIII antibodies for 20 min prior to celite activation and calcium addition. Other plasma had addition of fibrinogen (625 mg/dl)/FXIII (2 U/ml) or 30% dilution with hydroxyethyl starch before exposure to 0 or 200 mug/ml anti-FXIII antibodies. Thromboelastography was performed and data were collected until stable clot strength was observed. The exposure of normal plasma to anti-FXIII antibodies resulted in a significant (P < 0.05) decrease in clot strength (63%) compared with plasma without antibodies. Further samples exposed to anti-FXIII antibodies had clot strength no different from FXIII-deficient plasma. The FXIII-mediated clot strength varied between 44 and 50% in hypercoagulable and hypocoagulable plasma, respectively. In conclusion, the present investigation successfully demonstrated a novel method to detect the impact of FXIII activity in plasma samples. Further actuarial investigation will be required to determine the utility of this approach in the diagnosis and treatment of patients with either acquired FXIII deficiency or excess and concordant coagulopathy.

Cyclic AMP-dependent protein kinase in subtypes of major depression and normal volunteers.

We have shown a reduction in beta adrenoceptor-linked, cyclic AMP-dependent protein kinase [protein kinase A (PKA)] activity in fibroblasts of patients with major depression with melancholic features relative to normal volunteers. We evaluated a group of 35 patients with major depression subtyped by DSM-IV criteria as melancholic, atypical, and those not meeting either subtype designation ('non-subtyped') and 21 normal volunteers to ascertain whether or not the PKA activity abnormality was specific to melancholia. The melancholics showed marked reduction in cyclic AMP-stimulated PKA activity relative to normal volunteers. Although the atypicals were statistically significantly lower, almost all fell into the range for the normals. The non-subtyped group fell between the atypicals and the melancholics. Basal activity was significantly lower in atypical and melancholic groups. The data suggest that reduced PKA activity is consistently found in melancholic major depression and may not be seen with other depressive subtypes.

Fat embolus in femur fractures: a comparison of two reaming systems.

OBJECTIVE: To measure the amount of fat presented to the right heart during reaming and nail placement using two different reamer systems. DESIGN: Prospective, randomized clinical trial. SETTING: University-based Level I Trauma Center. PATIENTS: 20 patients with femur fractures. INTERVENTION: Patients with femur fractures were treated with intramedullary nailing using either reamer-irrigator-aspirator or a conventional reamer. MAIN OUTCOME MEASURE: four-chamber trans-esophageal echocardiogram was used to quantify the amount of fat presented to the right atrium. RESULTS: There were 2 female and 18 male patients enrolled, 10 in each group. There was no significant difference (p = 0.10) between reaming systems on the opening reamer, which was expected since both trial limbs used the same opening reamer. However, during the first pass of the reamer, the RIA showed a nearly-significant decrease in the volume of fat in the right atrium (p = 0.06). During passage of the nail, there was a significant difference with Group B having less fat embolus than Group A (p = 0.01). The power of this study is 0.81. The mean ISS was not significantly different between the two groups, nor was the sex, age or race. There was one death from cardiac complications in a patient who showed no fat during any phase of the procedure. This patient had significant mitral and aortic regurgitation pre-operatively. There was one patient with clinical fat embolism syndrome and one patient with a nonunion. CONCLUSIONS: There is a statistically significant difference in the amount of fat presented to the lungs using a RIA versus conventional reamer.

Patient with hypofibrinolysis-mediated thromboembolism converted to a hypercoagulable/hyperfibrinolytic state via ventricular assist device placement.

Mechanical circulatory support with ventricular assist devices (VADs) for end-stage heart failure has been a focus of intense interest for nearly four decades. Despite improvements in VAD design and biomaterial composition, it is common to administer anti-coagulation (e.g., warfarin, anti-platelet agents) to prevent both device thrombosis and thromboembolism. We present a patient with pre-operative thrombophilia (pulmonary embolism, intracardiac thrombus) and hypofibrinolysis, who subsequently developed hypercoagulability with hyperfibrinolysis with normalization of clot lifespan after left VAD placement. Such complex patient-VAD-hemostatic-state interactions serve as the rationale for continuing investigation of the effects of mechanical circulation on the fibrinolytic system and thrombophilia.

Mechanical circulatory device thrombosis: a new paradigm linking hypercoagulation and hypofibrinolysis.

This review considers the perhaps unappreciated role of contact pathway proteins in the pathogenesis of thrombotic/thromboembolic morbidity associated with mechanical circulatory support. Placement of ventricular assist devices (VADs) has been associated with consumption of circulating contact proteins and persistent generation of activated contact proteins such as Factor XII and high molecular weight kininogen. Importantly, activated contact proteins are absorbed to the surface of VADs via the Vroman effect. Further, hyperfibrinogenemia and persistent platelet activation exist in patients with VADs, likely contributing to speed of clot growth. Using thrombelastographic-based analyses, it has been determined that contact pathway protein activated coagulation results in a thrombus that develops strength at a significantly faster rate that tissue factor initiated coagulation. Further, thrombelastographic analyses that include the addition of tissue-type plasminogen activator have demonstrated that contact protein pathway activation results in thrombin activatable fibrinolysis inhibitor activation to a far greater extent than that observed with tissue factor initiated coagulation, resulting in a thrombus that takes significantly longer to lyse. These observations serve as the rational basis for clinical investigation to determine if regional suppression of thrombin generation with FXII/high molecular weight kininogen inhibition in concert with thrombin-activatable fibrinolysis inhibitor inhibition may decrease mechanical circulatory support-associated thrombotic morbidity.