| dc.description.abstract | Background: There are very limited data on the impact of commonly-prescribed antiplatelet regimens on von Willebrand Factor antigen (VWF:Ag), von Willebrand Factor propeptide (VWFpp) and A Disintegrin And Metalloprotease with ThromboSpondin type 1 motif number 13 (ADAMTS13) activity levels in patients with non-TTP related TIA or ischaemic stroke. Aims: The aims of this thesis were to simultaneously investigate the profiles of VWF:Ag, VWFpp and ADAMTS13 activity in patients within 4 weeks of TIA or ischaemic stroke onset (baseline) and then at 14 +/- 7 days (14d) and ≥ 90 days (90d) after commencing or altering antiplatelet therapy. We also aimed to assess the relationship between these endothelial +/- platelet biomarkers and platelet function/reactivity on user-friendly tests of platelet function at high and low shear stress, and whether blood group status influenced the expression of these biomarkers in CVD patients Methods: We performed a prospective, longitudinal observational analytical study in a clinically well-categorised population of CVD patients (N = 51) who were starting or changing to aspirin monotherapy or clopidogrel/ADP receptor antagonist monotherapy, or changing to aspirin-dipyridamole combination therapy. VWF:Ag and VWFpp levels were quantified with an enzyme-linked immunosorbent assay (ELISA) on thawed, double-spun platelet poor plasma. ADATMS13 activity was assessed with a Fluorescence Resonance Energy Transfer (FRET) assay on thawed double-spun platelet poor plasma. We also assessed the relationship between these biomarkers and relevant platelet function/reactivity tests using a moderately high shear stress test of platelet adhesion and aggregation (PFA-100®) and low shear stress tests of platelet aggregation (VerifyNow® and Multiplate®). Results: VWF:Ag levels significantly decreased between baseline and 90d (17.99 vs. 15.62µg/mL; P = 0.001) and the VWFpp/VWF:Ag ratio significantly increased at both 14d (0.09; P = 0.025) and 90d (0.10; P = 0.005) vs. baseline (0.08) in our overall CVD patient population. In the subgroup of patients who commenced or changed to clopidogrel monotherapy, VWF:Ag levels significantly decreased during follow-up at both 14d (17.43µg/mL; P = 0.007) and 90d (17.17µg/mL; P < 0.001) compared with baseline (20.29µg/mL). VWFpp levels remained stable over time, but the VWFpp/VWF:Ag ratio also significantly increased at both 14d (0.08; P = 0.014) and 90d (0.08; P = 0.024) vs. baseline (0.07), and ADAMTS13 activity was significantly lower at baseline vs. 90d (134.55% vs 147.2%; P = 0.047) in this subgroup starting clopidogrel. There were no significant changes in the levels of VWF:Ag (P ≥ 0.23) or VWFpp (P ≥ 0.34), or in the VWFpp/VWF:Ag ratio (P ≥ 0.08) in the subgroups who started aspirin or changed to aspirin-dipyridamole combination therapy. There was a positive relationship between VWF:Ag and VWFpp levels at all 3 timepoints in the overall CVD patient population (baseline R2 = 13.6%, P = 0.004; 14d R2 = 14.2%, P = 0.004; 90d R2 = 18.5%, P = 0.001), and in the aspirin-dipyridamole (baseline R2 = 20.7%, P = 0.029; 14d R2 = 27.7%, P = 0.012; 90d R2 = 38.4%, P = 0.003) and clopidogrel monotherapy subgroups (baseline R2 = 32.3%, P = 0.005; 14d R2 = 33.7%, P = 0.004; 90d R2 = 31.6%, P = 0.006). There was also a weak but significant inverse relationship between VWF:Ag levels and ADAMTS13 activity in the overall patient population at 90d only (R2 = 8.7%, P = 0.02). There was no obvious relationship between levels of VWF:Ag, VWFpp or ADAMTS13 activity and platelet function/reactivity or antiplatelet-HTPR status according to either cross-sectional or longitudinal definitions on the relevant assays in the aspirin monotherapy or clopidogrel monotherapy groups in OATS. However, there was a significant inverse relationship between VWF:Ag and PFA-100 C-ADP closure times at baseline (R2 = 32.0%, P = 0.008) and 14d (R2 = 46.4%, P = 0.001), and also a significant inverse relationship between VWFpp and PFA-100 C-ADP closure times at baseline (R2 = 18.3%, P = 0.043) and 14d (R2 = 24.3%, P = 0.018) in the subgroup of patients who changed to aspirin-dipyridamole combination treatment. In addition, there was a significant inverse relationship between VWF:Ag levels and PFA-100 INNOVANCE P2Y closure times (R2 = 55.5%, P < 0.001) and between VWFpp levels and PFA-100 INNOVANCE P2Y closure times (R2 = 20.9%, P = 0.028) only at 14d in this treatment subgroup. There was a positive relationship between ADAMTS13 activity and C-ADP closure times at baseline only in the starting aspirin-dipyridamole subgroup (R2 = 19.9%, P = 0.036), most likely mediated via VWF:Ag based on prior literature on this topic. The proportion of patients with blood group O and non-blood group O was similar in the overall CVD patient population and in all 3 treatment subgroups (P ≥ 0.194). However, mean VWF:Ag levels were lower in the overall patient population and in the subgroup of patients who were starting aspirin monotherapy who had blood group O vs. those with non-blood group O at baseline (P ≤ 0.043) and at 90d (P ≤ 0.01). Discussion and Conclusions: The prospective pilot studies outlined in this thesis have shown that starting or changing antiplatelet therapy may reduce circulating VWF:Ag levels and increase the VWFpp/VWF:Ag ratio without altering VWFpp expression in an overall CVD patient population, mainly attributed to commencement of clopidogrel monotherapy in this study. Taken together with the data on ADAMTS13 activity, our results support the hypothesis that these findings appear to be attributed to increased clearance of VWF:Ag over time, particularly in the subgroup who commenced clopidogrel. These data improve our understanding of the protean effects of clopidogrel on haemostatic and thrombotic profiles in CVD patients aside from direct platelet P2Y12 receptor blockade. Blood group O is associated with lower circulating VWF:Ag levels, thus adding to the limited literature on this topic in patients with TIA or ischaemic stroke. The significant inverse relationship between both VWF:Ag and VWFpp with PFA-100 C-ADP closure times early after TIA/ischaemic stroke in the subgroup of patients who changed to aspirin-dipyridamole combination treatment also improves our understanding of the endothelial +/- platelet biomarkers which can influence the results of these assays. Larger studies in a well-phenotyped CVD patient population which simultaneously assess all of these biomarkers, including the VWFpp/VWF:Ag ratio, are warranted to confirm these findings and to understand the precise mechanisms by which certain antiplatelet regimens, including aspirin-clopidogrel combination therapy, might affect VWF:Ag clearance. Furthermore, comprehensive assessment of these biomarkers should improve our understanding of the mechanisms influencing the ex vivo response to commonly-prescribed antiplatelet therapy on relevant high shear stress tests of platelet function/reactivity to potentially aid risk-stratification and tailor antiplatelet treatment to suit individual CVD patients in future. We aim to address this issue in local and multicentre studies which have been prospectively planned by our research group. | en |
