A Quantitative and Mechanistic Assessment of Activated Thrombin-Activatable Fibrinolysis Inhibitor and its Role in Pathological Bleeding and Thrombosis

The coagulation and fibrinolytic systems are linked by the thrombin-thrombomodulin complex which regulates each system through activation of protein C and TAFI, respectively. We have used novel assays and techniques to study the enzymology and biochemistry of TAFI and TAFIa, to measure TAFI activation in hemophilia A and protein C deficiency and to determine if enhancing TAFI activation can improve hemostasis in hemophilic plasma and whole blood. We show that TAFIa not TAFI attenuates fibrinolysis in vitro and this is supported by a relatively high catalytic efficiency (16.41μM-1s-1) of plasminogen binding site removal from fibrin degradation products (FDPs) by TAFIa. Since the catalytic efficiency of TAFIa in removing these sites is ~60-fold higher than that for inflammatory mediators such as bradykinin it is likely that FDPs are a physiological substrate of TAFIa. The high catalytic efficiency is primarily a result of a low Km which can be explained by a novel mechanism where TAFIa forms a binary complex with plasminogen and is recruited to the surface of FDPs. The low Km also suggests that TAFIa would effectively cleave lysines from FDPs during the early stages of fibrinolysis (i.e. at low concentrations of FDPs). Since individuals with hemophilia suffer from premature fibrinolysis as a result of insufficient TAFI activation we quantified TAFI activation in whole blood from hemophilic subjects. Both the rate of activation and the area under the TAFI activation time course (termed TAFIa potential) was determined to be reduced in hemophilia A and the TAFIa potential was significantly and inversely correlated with the clinical bleeding iii phenotype. Using a novel therapeutic strategy, we used soluble thrombomodulin to increase TAFI activation which improved the clot lysis time in factor VIII deficient human plasma and hemophilic dog plasma as well as hemophilic dog blood. Finally, we briefly show in a biochemical case study that TAFI activation is enhanced in protein C deficiency and when afflicted individuals are placed on Warfarin anticoagulant therapy, TAFI activation is reduced. Since TAFIa stabilizes blood clots, this suggests that reducing TAFI activation or inhibiting TAFIa may help restore blood flow in vessels with pathological thrombosis.

COGNITIVE ASSESSMENT USING THE KINARM EXOSKELETON ROBOT IN PATIENTS WITH TRANSIENT ISCHEMIC ATTACK

Background: Transient ischemic attack (TIA) is a condition causing focal neurological deficits lasting less than 24hrs. TIA patients present similarly to other conditions with rapid onset of neurological symptoms such as migraine. The accurate diagnosis of TIA is critical because it serves as a warning for subsequent stroke. Furthermore, cognitive deficit associated with TIA may predict the development of dementia. Therefore, characterizing the cognitive symptoms of TIA patients and discriminating these patients from those with similar symptoms is important for proper diagnosis and treatment. Currently the diagnosis of TIA is made on clinical and radiographic evidence. Robotic assessment, with instruments such as the KINARM, may improve the identification of cognitive impairment in TIA patients. Methods: In this prospective cohort study, two KINARM tests, trail making task (TMT) and spatial span task (SST), were used to detect cognitive deficits. Two study groups were made. The TIA group was tested at 5 time points over the span of a year. The migraine active control group had one initial visit and another a year later. Both of these groups were compared to a normative database of approximately 400 healthy volunteers. From this database age and sex matched normative data was used to calculate Z-scores for the TMT. The Montreal Cognitive Assessment (MoCA) was also administered to both groups. Results: 31 participants were recruited, 20 TIA group and 11 active controls (mean ± SD age= 66 ± 11.3 and 62 ± 14.5). There was no significant difference in TIA and active control group MoCA scores. The TMT was able to detect cognitive impairment in TIA and migraine group. Also, both KINARM tasks could detect significant differences in performance between TIA and migraine patients while the MoCA could not. Changes in TIA and migraine performance on the MoCA, TMT, and SST were observed. Conclusions: The robotic KINARM exoskeleton can be used to assess cognitive deficits in TIA patients.